# AMR-English alignment release (generated on Mon Mar 14, 2016 at 23:18:32) # ::id a_pmid_2094_2929.7 ::amr-annotator SDL-AMR-09 ::preferred # ::tok 1 @- RT @-@ PCR and western blot analyses confirmed the strong up @-@ regulation of serpinE2 expression and secretion by IECs expressing oncogenic MEK , Ras or BRAF . # ::alignments 0-1.1 2-1.2.1.1.1.1 4-1.2.1.1.1.1 5-1.2 6-1.2.2 7-1.2.2 8-1.2.1 9-1 11-1.3.1.2 12-1.3.1 13-1.3.1 14-1.3.1 15-1.3.1.1.r 16-1.3.1.1.1.1.1 17-1.3.1.1 18-1.3 19-1.3.2 20-1.3.2.1.r 21-1.3.2.1.1.1 22-1.3.2.1.2 23-1.3.2.1.2.1.4 23-1.3.2.1.2.1.4.1.2.1 24-1.3.2.1.2.1.1.1.1 26-1.3.2.1.2.1.2.1.1 27-1.3.2.1.2.1 28-1.3.2.1.2.1.3.1.1 (c / confirm-01~e.9 :li 1~e.0 :ARG0 (a / and~e.5 :op1 (a2 / analyze-01~e.8 :instrument (t / thing :name (n4 / name :op1 "RT-PCR"~e.2,4))) :op2 (i / immunoblot-01~e.6,7)) :ARG1 (a4 / and~e.18 :op1 (u / upregulate-01~e.12,13,14 :ARG1~e.15 (e3 / express-03~e.17 :ARG2 (p / protein :name (n6 / name :op1 "serpinE2"~e.16))) :ARG1-of (s / strong-02~e.11)) :op2 (s2 / secrete-01~e.19 :ARG0~e.20 (c2 / cell :name (n7 / name :op1 "IEC"~e.21) :ARG3-of (e4 / express-03~e.22 :ARG2 (o2 / or~e.27 :op1 (e / enzyme :name (n2 / name :op1 "MEK"~e.24)) :op2 (e2 / enzyme :name (n3 / name :op1 "Ras"~e.26)) :op3 (e5 / enzyme :name (n8 / name :op1 "BRAF"~e.28)) :ARG0-of (c3 / cause-01~e.23 :ARG1 (d / disease :wiki "Cancer" :name (n / name :op1 "cancer"~e.23)))))) :ARG1 p))) # ::id a_pmid_2094_2929.8 ::amr-annotator SDL-AMR-09 ::preferred # ::tok 2 @- Interestingly , serpinE2 mRNA and protein were also markedly enhanced in human CRC cells exhibiting mutation in KRAS @ and BRAF @ . # ::alignments 0-1.1 2-1.6 2-1.6.r 4-1.2.2.1.1 5-1.2.1.1.1 6-1.2 7-1.2.2 9-1.4 10-1.3 10-1.3.r 11-1 12-1.5.r 13-1.5.2 14-1.5.3.1.1 15-1.5 16-1.5.1 17-1.5.1.1 20-1.5.1.1.1.1.1.1 22-1.5.1.1.1 24-1.5.1.1.1.2.1.1 (e / enhance-01~e.11 :li 2~e.0 :ARG1 (a3 / and~e.6 :op1 (n6 / nucleic-acid :name (n / name :op1 "mRNA"~e.5) :ARG0-of (e2 / encode-01 :ARG1 p)) :op2 (p / protein~e.7 :name (n2 / name :op1 "serpinE2"~e.4))) :manner~e.10 (m / marked~e.10) :mod (a2 / also~e.9) :location~e.12 (c / cell~e.15 :ARG0-of (e3 / exhibit-01~e.16 :ARG1 (m2 / mutate-01~e.17 :ARG1 (a4 / and~e.22 :op1 (g / gene :name (n4 / name :op1 "KRAS"~e.20)) :op2 (g2 / gene :name (n5 / name :op1 "BRAF"~e.24))))) :mod (h / human~e.13) :mod (d / disease :name (n3 / name :op1 "CRC"~e.14))) :manner~e.2 (i / interesting~e.2)) # ::id a_pmid_2094_2929.9 ::amr-annotator SDL-AMR-09 ::preferred # ::tok 3 @- RNAi directed against serpinE2 in caMEK @-@ transformed rat IECs or in human CRC cell lines HCT116 and LoVo markedly decreased foci formation , anchorage @-@ independent growth in soft agarose , cell migration and tumor formation in nude mice . # ::alignments 0-1.1 3-1.2.2 4-1.2.2.1 5-1.2.2.1.1.1.1.1 5-1.2.2.1.1.2.1.1 9-1.2.2.1.1.1.2.3 10-1.2.2.1.1.1.2.2 11-1.2.2.1.1.1.2.1.1 12-1.2.2.1.1 13-1.2.2.1.1.r 14-1.2.2.1.1.2.2.2 15-1.2.2.1.1.2.2.3.1.1 16-1.2.2.1.1.2.2 17-1.2.2.1.1.2.2.1.1.1 17-1.2.2.1.1.2.2.1.1.2 18-1.2.2.1.1.2.2.1.1.1.1.1 19-1.2.2.1.1.2.2.1.1 20-1.2.2.1.1.2.2.1.1.2.1.1 21-1.4 21-1.4.r 22-1 23-1.3.1.1 24-1.3.1 26-1.3.2.1.2 28-1.3.2.1 28-1.3.2.1.1 28-1.3.2.1.1.r 29-1.3.2 30-1.3.2.2.r 31-1.3.2.2.1 32-1.3.2.2 34-1.3.3.1 35-1.3.3 36-1.3 37-1.3.4.1 38-1.3.4 39-1.3.4.2.r 40-1.3.4.2.1 41-1.3.4.2 (d / decrease-01~e.22 :li 3~e.0 :ARG0 (i2 / interfere-01 :ARG1 (n / nucleic-acid :name (n11 / name :op1 "RNA")) :ARG1-of (d2 / direct-01~e.3 :ARG2 (o2 / oppose-01~e.4 :ARG1~e.13 (o / or~e.12 :op1 (p / protein :name (n2 / name :op1 "serpinE2"~e.5) :location (c / cell :name (n3 / name :op1 "IEC"~e.11) :mod (r / rat~e.10) :ARG1-of (t / transform-01~e.9 :ARG0 (e / enzyme :name (n4 / name :op1 "MEK") :ARG2-of (m5 / mutate-01) :mod (c3 / constitutive) :ARG1-of (a4 / activate-01))))) :op2 (p2 / protein :name (n5 / name :op1 "serpinE2"~e.5) :location (c6 / cell~e.16 :ARG2-of (i / include-91 :ARG1 (a3 / and~e.19 :op1 (c2 / cell-line~e.17 :name (n6 / name :op1 "HCT116"~e.18)) :op2 (c4 / cell-line~e.17 :name (n8 / name :op1 "LoVo"~e.20)))) :mod (h / human~e.14) :mod (d4 / disease :name (n10 / name :op1 "CRC"~e.15)))))))) :ARG1 (a / and~e.36 :op1 (f / form-01~e.24 :ARG1 (f2 / focus~e.23)) :op2 (g / grow-01~e.29 :ARG1-of (d3 / depend-01~e.28 :polarity~e.28 -~e.28 :ARG0 (a2 / anchorage~e.26)) :location~e.30 (a5 / agarose~e.32 :ARG1-of (s / soft-02~e.31))) :op3 (m3 / migrate-01~e.35 :ARG0 (c5 / cell~e.34)) :op4 (f3 / form-01~e.38 :ARG1 (t2 / tumor~e.37) :location~e.39 (m4 / mouse~e.41 :mod (n9 / nude~e.40)))) :manner~e.21 (m2 / marked~e.21)) # ::id a_pmid_2094_2929.10 ::amr-annotator SDL-AMR-09 ::preferred # ::tok 4 @- Treatment of CRC cell lines with U0126 markedly reduced serpinE2 @ mRNA levels , indicating that expression of serpinE2 @ is likely dependent of ERK activity . # ::alignments 0-1.1 2-1.2 3-1.2.1.r 4-1.2.1.1.1.1 5-1.2.1 6-1.2.1 7-1.2.2.r 8-1.2.2.1.1 9-1.4 9-1.4.r 10-1 12-1.3.1.2.1.1.1 14-1.3.1.1.1 15-1.3 17-1.5 18-1.5.1.r 19-1.5.1.1.1 22-1.5.1.1.1.1 25-1.5.1 26-1.5.1.1 27-1.5.1.1.2.r 28-1.5.1.1.2.1.1.1 29-1.5.1.1.2 (r / reduce-01~e.10 :li 4~e.0 :ARG0 (t / treat-04~e.2 :ARG1~e.3 (c / cell-line~e.5,6 :mod (d2 / disease :name (n3 / name :op1 "CRC"~e.4))) :ARG2~e.7 (s / small-molecule :name (n / name :op1 "U0126"~e.8))) :ARG1 (l / level~e.15 :quant-of (n6 / nucleic-acid :name (n4 / name :op1 "mRNA"~e.14) :ARG0-of (e2 / encode-01 :ARG1 (p / protein :name (n5 / name :op1 "serpinE2"~e.12))))) :manner~e.9 (m / marked~e.9) :ARG0-of (i / indicate-01~e.17 :ARG1~e.18 (l2 / likely-01~e.25 :ARG1 (d / depend-01~e.26 :ARG0 (e3 / express-03~e.19 :ARG2 p~e.22) :ARG1~e.27 (a / activity-06~e.29 :ARG0 (e / enzyme :name (n2 / name :op1 "ERK"~e.28))))))) # ::id a_pmid_2094_2929.11 ::amr-annotator SDL-AMR-09 ::preferred # ::tok 5 @- Finally , Q @-@ PCR analyses demonstrated that mRNA levels of serpinE2 were markedly increased in human adenomas in comparison to healthy adjacent tissues and in colorectal tumors , regardless of tumor stage and grade . # ::alignments 0-1.1 2-1.1.r 2-1.2 2-1.2.r 4-1.3.1.1.1 6-1.3.1.1.1 7-1.3 8-1 9-1.4.r 10-1.4.1.1.1.1 11-1.4.1 13-1.4.1.1.2.1.1.1 15-1.4.4 15-1.4.4.r 16-1.4 17-1.4.2.r 18-1.4.2.1.1 19-1.4.2.1 21-1.4.5.r 23-1.4.5.2 24-1.4.5.1 25-1.4.5 26-1.4.2 28-1.4.2.2.1 29-1.4.2.2 31-1.4.3 33-1.4.3.1.1.1.1 34-1.4.3.1.1 34-1.4.3.1.1.1 34-1.4.3.1.1.1.r 35-1.4.3.1 36-1.4.3.1.2 (d / demonstrate-01~e.8 :li~e.2 5~e.0 :li~e.2 -1~e.2 :ARG0 (a / analyze-01~e.7 :mod (t / thing :name (n / name :op1 "Q-PCR"~e.4,6))) :ARG1~e.9 (i / increase-01~e.16 :ARG1 (l / level~e.11 :quant-of (n4 / nucleic-acid :name (n2 / name :op1 "mRNA"~e.10) :ARG0-of (e / encode-01 :ARG1 (p / protein :name (n3 / name :op1 "serpinE2"~e.13))))) :location~e.17 (a2 / and~e.26 :op1 (a3 / adenomas~e.19 :mod (h / human~e.18)) :op2 (t3 / tumor~e.29 :mod (c / colon~e.28))) :ARG1-of (r / regardless-91~e.31 :ARG2 (a5 / and~e.35 :op1 (t5 / thing~e.34 :ARG2-of~e.34 (s / stage-02~e.34 :ARG1 (t4 / tumor~e.33))) :op2 (g / grade~e.36 :mod t4))) :manner~e.15 (m / marked~e.15) :compared-to~e.21 (t2 / tissue~e.25 :mod (a4 / adjacent~e.24) :mod (h2 / healthy~e.23)))) # ::id a_pmid_2094_2929.39 ::amr-annotator SDL-AMR-09 ::preferred # ::tok SerpinE2 is overexpressed in intestinal epithelial cells transformed by activated MEK1 and oncogenic RAS and BRAF # ::alignments 2-1 3-1.2.r 4-1.2.2 5-1.2.1 6-1.2 7-1.2.3 8-1.2.3.1.r 9-1.2.3.1.1.2 10-1.2.3.1.1.1.1 11-1.2.3.1 12-1.2.3.1.2 12-1.2.3.1.2.2 12-1.2.3.1.2.2.1.2.1 12-1.2.3.1.2.2.r 13-1.2.3.1.2.1.1 14-1.2.3.1 15-1.2.3.1.3.1.1 (o / overexpress-01~e.2 :ARG1 (p / protein :name (n2 / name :op1 "serpinE2")) :location~e.3 (c / cell~e.6 :mod (e2 / epithelium~e.5) :part-of (i / intestine~e.4) :ARG1-of (t / transform-01~e.7 :ARG0~e.8 (a / and~e.11,14 :op1 (e3 / enzyme :name (n3 / name :op1 "MEK1"~e.10) :ARG1-of (a2 / activate-01~e.9)) :op2 (e4 / enzyme~e.12 :name (n4 / name :op1 "RAS"~e.13) :ARG0-of~e.12 (c2 / cause-01~e.12 :ARG1 (d / disease :wiki "Cancer" :name (n / name :op1 "cancer"~e.12)))) :op3 (e / enzyme :name (n5 / name :op1 "BRAF"~e.15) :ARG0-of c2))))) # ::id a_pmid_2094_2929.40 ::amr-annotator SDL-AMR-09 ::preferred # ::tok Among the most harmful of all genetic abnormalities that appear in CRC development are mutations of KRAS and its downstream effector BRAF as they result in abnormal ERK signaling . # ::alignments 0-1 2-1.2.4.1 3-1.2.4 5-1.2.3 6-1.2.2 9-1.2.5 11-1.2.5.1.1.1.1.1 12-1.2.5.1 14-1.1.1 15-1.1.1.1.r 16-1.1.1.1.1.1 17-1.1 18-1.1.2.3 18-1.1.2.3.r 19-1.1.2.1 20-1.1.2 21-1.1.2.2.1.1 22-1.2.5.1.r 24-1.1.3 26-1.1.3.1.2 26-1.1.3.1.2.1 26-1.1.3.1.2.1.r 26-1.2 26-1.2.1 26-1.2.1.r 27-1.1.3.1.1.1.1 28-1.1.3.1 (i / include-91~e.0 :ARG1 (a4 / and~e.17 :op1 (m2 / mutate-01~e.14 :ARG1~e.15 (e4 / enzyme :name (n3 / name :op1 "KRAS"~e.16))) :op2 (e2 / effector~e.20 :mod (d2 / downstream~e.19) :mod (e3 / enzyme :name (n4 / name :op1 "BRAF"~e.21)) :poss~e.18 e4~e.18) :ARG1-of (r / result-01~e.24 :ARG2 (s / signal-07~e.28 :ARG0 (e / enzyme :name (n / name :op1 "ERK"~e.27)) :ARG1-of (n6 / normal-02~e.26 :polarity~e.26 -~e.26)))) :ARG2 (n5 / normal-02~e.26 :polarity~e.26 -~e.26 :ARG2 (g / genetics~e.6) :mod (a2 / all~e.5) :ARG0-of (h / harmful-02~e.3 :degree (m / most~e.2)) :ARG1-of (a3 / appear-01~e.9 :time~e.22 (d / develop-02~e.12 :ARG1 (c / cell :mod (d3 / disease :name (n2 / name :op1 "CRC"~e.11))))))) # ::id a_pmid_2094_2929.41 ::amr-annotator SDL-AMR-09 ::preferred # ::tok In a previous report , we had shown that expression of a constitutive active mutant of MEK1 ( caMEK ) in the intestinal epithelial cell line IEC @-@ 6 induced morphological transformation and growth in soft agar ; in marked contrast , wtMEK overexpression had no effect on IEC @-@ 6 phenotype [ @ 3 @ ] . # ::alignments 2-1.1.3.1 3-1.1.3 5-1.1.1 7-1.1 8-1.1.2.r 9-1.1.2.1 10-1.1.2.1.1.r 12-1.1.2.1.1.2 13-1.1.2.1.1.3 14-1.1.2.1.1 15-1.1.2.1.1.1.r 16-1.1.2.1.1.1.1.1 20-1.1.2.1.2.r 22-1.1.2.1.2.2.1 23-1.1.2.1.2.2 24-1.1.2.1.2 25-1.1.2.1.2 25-1.2.3.1 26-1.1.2.1.2.1.1 28-1.1.2.1.2.1.1 29-1.1.2 30-1.1.2.2.1.1 31-1.1.2.2.1 32-1.1.2.2 33-1.1.2.2.2 34-1.1.2.2.2.1.r 35-1.1.2.2.2.1.1 36-1.1.2.2.2.1 39-1.2.5.1 40-1.2.5 43-1.2.2 45-1.2.1 45-1.2.1.r 46-1.2 47-1.2.3.r 48-1.2.3.1.1.1 50-1.2.3.1.1.1 51-1.2.3 54-1.2.4.1.1.1 (a / and :op1 (s / show-01~e.7 :ARG0 (w / we~e.5) :ARG1~e.8 (i / induce-01~e.29 :ARG0 (e / express-03~e.9 :ARG2~e.10 (m / mutate-01~e.14 :ARG2~e.15 (e2 / enzyme :name (n / name :op1 "MEK1"~e.16)) :mod (c / constitutive~e.12) :ARG0-of (a2 / activity-06~e.13)) :ARG3~e.20 (c2 / cell-line~e.24,25 :name (n3 / name :op1 "IEC-6"~e.26,28) :mod (e4 / epithelium~e.23 :part-of (i2 / intestine~e.22)))) :ARG2 (a3 / and~e.32 :op1 (t / transform-01~e.31 :mod (m3 / morphological~e.30)) :op2 (g / grow-01~e.33 :location~e.34 (a4 / agar~e.36 :ARG1-of (s2 / soft-02~e.35))))) :medium (r / report~e.3 :time (p / previous~e.2))) :op2 (a5 / affect-01~e.46 :polarity~e.45 -~e.45 :ARG0 (o / overexpress-01~e.43 :ARG1 (e5 / enzyme :name (n4 / name :op1 "MEK") :mod (w2 / wild-type))) :ARG1~e.47 (p2 / phenotype~e.51 :mod (c3 / cell-line~e.25 :name (n5 / name :op1 "IEC-6"~e.48,50))) :ARG1-of (d / describe-01 :ARG0 (p3 / publication :ARG1-of (c4 / cite-01 :ARG2 3~e.54))) :ARG2-of (c5 / contrast-01~e.40 :ARG1-of (m4 / mark-01~e.39)))) # ::id a_pmid_2094_2929.42 ::amr-annotator SDL-AMR-09 ::preferred # ::tok In order to understand the mechanisms by which activated MEK1 induces intestinal cell tumorigenesis , the pattern of gene expression was analyzed by microarray in IEC @-@ 6 cells overexpressing activated MEK1 . # ::alignments 0-1.4.r 1-1.4.r 2-1.4.r 3-1.4 5-1.4.1 8-1.4.1.1.1.2 9-1.4.1.1.1.1.1 10-1.4.1.1 11-1.4.1.1.2.1.1.1 12-1.4.1.1.2.1.1 13-1.4.1.1.2 13-1.4.1.1.2.1 13-1.4.1.1.2.1.r 16-1.2 17-1.2.1.r 18-1.2.1.1 19-1.2.1 21-1 22-1.1.r 23-1.1 24-1.3.r 25-1.3.1.1 27-1.3.1.1 28-1.3 29-1.3.2 30-1.3.2.1 31-1.3.2.1 (a3 / analyze-01~e.21 :ARG0~e.22 (m2 / microarray~e.23) :ARG1 (p / pattern-01~e.16 :ARG1~e.17 (e2 / express-03~e.19 :ARG2 (g / gene~e.18))) :location~e.24 (c2 / cell-line~e.28 :name (n2 / name :op1 "IEC-6"~e.25,27) :ARG0-of (o / overexpress-01~e.29 :ARG1 e~e.30,31)) :purpose~e.0,1,2 (u / understand-01~e.3 :ARG1 (m / mechanism~e.5 :instrument-of (i / induce-01~e.10 :ARG0 (e / enzyme :name (n / name :op1 "MEK1"~e.9) :ARG1-of (a / activate-01~e.8)) :ARG2 (c3 / create-01~e.13 :ARG1~e.13 (t / tumor~e.13 :mod (c / cell~e.12 :part-of (i2 / intestine~e.11)))))))) # ::id a_pmid_2094_2929.43 ::amr-annotator SDL-AMR-09 ::preferred # ::tok Results from microarrays comparing control ( wtMEK ) to caMEK @-@ expressing IEC @-@ 6 cells identified the Serpin clade E member 2 @ ( serpinE2 @ or PN @-@ 1 ) gene as a potential target of activated MEK1 . # ::alignments 0-1.1 0-1.1.1 0-1.1.1.r 1-1.1.2.r 2-1.1.2 3-1.1.2.1 4-1.1.2.1.1 11-1.1.2.1.2.2 12-1.1.2.1.2.1.1 14-1.1.2.1.2.1.1 15-1.1.2.1.2 16-1 19-1.2.1.1 20-1.2.1.2 21-1.2.1.3 22-1.2.1.4 23-1.2.1.5 27-1.2.2.1.1.1.1 31-1.2.2.1.2.1.1 33-1.2.2.1.2.1.1 36-1.2 36-1.2.2.1.1 36-1.2.2.1.2 37-1.3.r 39-1.3.3 40-1.3 42-1.1.2.1.2.2.1.4 42-1.3.1.2 43-1.3.1.1.1 (i / identify-01~e.16 :ARG0 (t / thing~e.0 :ARG2-of~e.0 (r / result-01~e.0) :source~e.1 (m / microarray~e.2 :ARG0-of (c / compare-01~e.3 :ARG1 (c2 / control~e.4 :ARG1-of (m2 / mean-01 :ARG2 (e / enzyme :name (n / name :op1 "MEK") :mod (w / wild-type)))) :ARG2 (c3 / cell-line~e.15 :name (n2 / name :op1 "IEC-6"~e.12,14) :ARG3-of (e2 / express-03~e.11 :ARG2 (e3 / enzyme :name (n3 / name :op1 "MEK") :ARG2-of (m4 / mutate-01) :mod (c4 / constitutive) :ARG1-of (a4 / activate-01~e.42))))))) :ARG1 (g / gene~e.36 :name (n4 / name :op1 "Serpin"~e.19 :op2 "clade"~e.20 :op3 "E"~e.21 :op4 "member"~e.22 :op5 2~e.23) :ARG1-of (m3 / mean-01 :ARG2 (a / and :op1 (g2 / gene~e.36 :name (n5 / name :op1 "serpinE2"~e.27)) :op2 (g3 / gene~e.36 :name (n6 / name :op1 "PN-1"~e.31,33))))) :ARG2~e.37 (t2 / target-01~e.40 :ARG0 (e4 / enzyme :name (n7 / name :op1 "MEK1"~e.43) :ARG1-of (a2 / activate-01~e.42)) :ARG1 g :mod (p / potential~e.39))) # ::id a_pmid_2094_2929.44 ::amr-annotator SDL-AMR-09 ::preferred # ::tok Indeed , serpinE2 @ expression was significantly induced by more that 28 @-@ fold ( p < 0.05 ) in cells overexpressing activated MEK1 in comparison to cells expressing wtMEK ( data not shown ) . # ::alignments 0-1.2 3-1.1.1.1.1 5-1.1 7-1.3 8-1 9-1.7.r 10-1.7 12-1.7.1.1 14-1.7.1 16-1.8 17-1.8.1 18-1.8.1.1 20-1.4.r 21-1.4 22-1.4.1 23-1.4.1.1.2 24-1.4.1.1.1.1 26-1.5.r 28-1.5 29-1.5.1 32-1.6.1 33-1.6.1.1.1 33-1.6.1.1.1.r 34-1.6.1.1 (i / induce-01~e.8 :ARG2 (e / express-03~e.5 :ARG1 (g / gene :name (n / name :op1 "serpinE2"~e.3))) :mod (i2 / indeed~e.0) :ARG1-of (s / significant-02~e.7) :location~e.20 (c / cell~e.21 :location-of (o / overexpress-01~e.22 :ARG1 (e2 / enzyme :name (n2 / name :op1 "MEK1"~e.24) :ARG1-of (a / activate-01~e.23)))) :compared-to~e.26 (c2 / cell~e.28 :ARG3-of (e3 / express-03~e.29 :ARG2 (e4 / enzyme :name (n3 / name :op1 "MEK") :mod (w / wild-type)))) :ARG1-of (d / describe-01 :ARG0 (d2 / data~e.32 :ARG1-of (s2 / show-01~e.34 :polarity~e.33 -~e.33))) :quant~e.9 (m2 / more-than~e.10 :op1 (p / product-of~e.14 :op1 28~e.12)) :ARG1-of (s3 / statistical-test-91~e.16 :ARG2 (l / less-than~e.17 :op1 0.05~e.18))) # ::id a_pmid_2094_2929.45 ::amr-annotator SDL-AMR-09 ::preferred # ::tok Overexpression of serpinE2 @ in caMEK @-@ expressing IECs was furthermore confirmed following RT @-@ PCR analysis as shown in Figure 1A @ . # ::alignments 0-1.1.1 3-1.1.1.1.1.1 8-1.1.1.2.2 9-1.1.1.2.1.1 11-1 12-1.1 13-1.1.2 14-1.1.2.1.1.1.1 16-1.1.2.1.1.1.1 17-1.1.2.1 18-1.1.2.r 19-1.2 20-1.2.1.r 21-1.2.1 23-1.2.1.1 (a / and~e.11 :op2 (c / confirm-01~e.12 :ARG1 (o / overexpress-01~e.0 :ARG1 (g / gene :name (n / name :op1 "serpinE2"~e.3)) :location (c2 / cell :name (n2 / name :op1 "IEC"~e.9) :ARG3-of (e / express-03~e.8 :ARG2 (e2 / enzyme :name (n3 / name :op1 "MEK") :mod (c3 / constitutive) :ARG2-of (m / mutate-01) :ARG1-of (a4 / activate-01))))) :time~e.18 (f / follow-01~e.13 :ARG2 (a2 / analyze-01~e.17 :mod (t / thing :name (n4 / name :op1 "RT-PCR"~e.14,16))))) :ARG1-of (s / show-01~e.19 :ARG0~e.20 (f2 / figure~e.21 :mod "1A"~e.23))) # ::id a_pmid_2094_2929.46 ::amr-annotator SDL-AMR-09 ::preferred # ::tok SerpinE2 expression was also markedly enhanced in IEC @-@ 6 cells transformed by oncogenic RAS ( 26 @-@ fold ) or BRAF ( 12 @-@ fold after 12 h of 4 @-@ hydroxytamoxifen ( 4 @-@ OHT ) ( Figure 1B and 1C ) . # ::alignments 0-1.1.1.1.1.1 1-1.1.1 3-1.1.4 4-1.1.3 4-1.1.3.r 5-1.1 5-1.2 7-1.1.5.1.1 7-1.2.4.1.1 9-1.1.5.1.1 10-1.1.5 10-1.2.4 11-1.1.5.2 11-1.2.4.2 12-1.1.5.2.1.r 13-1.1.5.2.1 13-1.1.5.2.1.2 13-1.1.5.2.1.2.1.2.1 13-1.1.5.2.1.2.r 14-1.1.5.2.1.1.1 16-1.1.2.1 18-1.1.2 20-1 21-1.2.4.2.1.1.1 23-1.2.2.1 25-1.2.2 26-1.2.3 27-1.2.3.2.1 28-1.2.3.2.2 30-1.2.3.1.1.1.1 32-1.2.3.1.1.1.1 34-1.2.3.1.1.1.1 39-1.3.1.1 39-1.3.1.2 41-1.3.1.1.1 43-1.3.1 45-1.3.1.2.1 (o / or~e.20 :op1 (e2 / enhance-01~e.5 :ARG1 (e3 / express-03~e.1 :ARG2 (p / protein :name (n2 / name :op1 "SerpinE2"~e.0))) :ARG3 (p2 / product-of~e.18 :op1 26~e.16) :manner~e.4 (m / marked~e.4) :mod (a / also~e.3) :location (c / cell-line~e.10 :name (n3 / name :op1 "IEC-6"~e.7,9) :ARG1-of (t / transform-01~e.11 :ARG0~e.12 (e / enzyme~e.13 :name (n / name :op1 "Ras"~e.14) :ARG0-of~e.13 (c2 / cause-01~e.13 :ARG1 (d2 / disease :wiki "Cancer" :name (n6 / name :op1 "cancer"~e.13))))))) :op2 (e4 / enhance-01~e.5 :ARG1 e3 :ARG3 (p4 / product-of~e.25 :op1 12~e.23) :time (a2 / after~e.26 :op1 (s / stimulate-01 :ARG2 (s2 / small-molecule :name (n5 / name :op1 "4-hydroxytamoxifen"~e.30,32,34))) :quant (t2 / temporal-quantity :quant 12~e.27 :unit (h / hour~e.28))) :location (c4 / cell-line~e.10 :name (n7 / name :op1 "IEC-6"~e.7) :ARG1-of (t3 / transform-01~e.11 :ARG0 (e5 / enzyme :name (n4 / name :op1 "BRAF"~e.21) :ARG0-of c2)))) :ARG1-of (d / describe-01 :ARG0 (a3 / and~e.43 :op1 (f / figure~e.39 :mod "1B"~e.41) :op2 (f2 / figure~e.39 :mod "1C"~e.45)))) # ::id a_pmid_2094_2929.47 ::amr-annotator SDL-AMR-09 ::preferred # ::tok Of note , the induction of serpinE2 was induced within 1 h following ERK activation as observed in cells expressing the inducible BRAF : ER fusion protein stimulated with 4 @-@ OHT ( Figure 1C ) . # ::alignments 1-1 4-1.1 6-1.1.1.1.1.1 8-1.1 9-1.1.2 9-1.1.2.2 9-1.1.2.2.1.1.r 9-1.1.2.2.r 10-1.1.2.2.1.1 11-1.1.2.2.1.2 12-1.1.2 12-1.1.3 13-1.1.2.1.1.1.1 14-1.1.2.1 15-1.1.2.r 15-1.1.4.r 16-1.1.4 17-1.1.4.1.r 18-1.1.4.1 19-1.1.4.1.1 21-1.1.1 21-1.1.4.1.1.1.2 22-1.1.4.1.1.1.1.1 24-1.1.4.1.1.1.1.1 25-1.1.4.1.1.1.1.2 26-1.1.1.1 26-1.1.4.1.1.1 27-1.1.4.1.1.1.3 28-1.1.4.1.1.1.3.1.r 29-1.1.4.1.1.1.3.1.1.1 31-1.1.4.1.1.1.3.1.1.1 33-1.2.1 35-1.2.1.1 (n2 / note-02~e.1 :ARG1 (i / induce-01~e.4,8 :ARG2 (i2 / induce-01~e.21 :ARG2 (p / protein~e.26 :name (n3 / name :op1 "serpinE2"~e.6))) :time~e.15 (a / after~e.9,12 :op1 (a2 / activate-01~e.14 :ARG1 (e / enzyme :name (n / name :op1 "ERK"~e.13))) :quant~e.9 (u / up-to~e.9 :op1 (t / temporal-quantity :quant~e.9 1~e.10 :unit (h / hour~e.11)))) :ARG1-of (f / follow-01~e.12) :ARG1-of~e.15 (o / observe-01~e.16 :location~e.17 (c / cell~e.18 :ARG3-of (e2 / express-03~e.19 :ARG2 (p3 / protein~e.26 :name (n5 / name :op1 "BRAF:ER"~e.22,24 :op2 "fusion"~e.25) :ARG1-of (i3 / induce-01~e.21 :ARG1-of (p2 / possible-01)) :ARG1-of (s / stimulate-01~e.27 :ARG2~e.28 (s2 / small-molecule :name (n4 / name :op1 "4-OHT"~e.29,31)))))))) :ARG1-of (d / describe-01 :ARG0 (f2 / figure~e.33 :mod "1C"~e.35))) # ::id a_pmid_2094_2929.48 ::amr-annotator SDL-AMR-09 ::preferred # ::tok Treatment with the MEK @-@ inhibitor U0126 completely abrogated serpinE2 gene expression induced by oncogenic MEK1 ( Figure 1A ) and BRAF ( Figure 1C ) , indicating that induction of serpinE2 is an early and direct event occurring following the activation of ERK signaling . # ::alignments 0-1.2 1-1.2.1.r 3-1.2.1.2.1.1.1 5-1.2.1 5-1.2.1.2 5-1.2.1.2.r 6-1.2.1.1.1 7-1.3 8-1 9-1.1.1.1.1 10-1.1.1 11-1.1 12-1.1.2 12-1.1.3 13-1.1.2.1.r 14-1.1.2.1 14-1.1.2.1.2 14-1.1.2.1.2.1.2.1 14-1.1.2.1.2.r 15-1.1.2.1.1.1 17-1.1.2.2.1 19-1.1.2.2.1.1 23-1.1.3.1.1.1 25-1.1.3.2.1 27-1.1.3.2.1.1 31-1.4 32-1.4.1.r 33-1.4.1.3 34-1.4.1.3.1.r 35-1.4.1.3.1 36-1.4.1.3.r 38-1.4.1.1 40-1.4.1.2 41-1.4.1 43-1.4.1.4 45-1.4.1.4.1 46-1.4.1.4.1.1.r 47-1.4.1.4.1.1.1.1.1 48-1.4.1.4.1.1 (a / abrogate-01~e.8 :ARG1 (e3 / express-03~e.11 :ARG2 (g / gene~e.10 :name (n5 / name :op1 "serpinE2"~e.9)) :ARG2-of (i2 / induce-01~e.12 :ARG0~e.13 (e4 / enzyme~e.14 :name (n6 / name :op1 "MEK1"~e.15) :ARG0-of~e.14 (c2 / cause-01~e.14 :ARG1 (d4 / disease :wiki "Cancer" :name (n / name :op1 "cancer"~e.14)))) :ARG1-of (d / describe-01 :ARG0 (f / figure~e.17 :mod "1A"~e.19))) :ARG2-of (i3 / induce-01~e.12 :ARG0 (e7 / enzyme :name (n7 / name :op1 "BRAF"~e.23) :ARG0-of c2) :ARG1-of (d2 / describe-01 :ARG0 (f2 / figure~e.25 :mod "1C"~e.27)))) :ARG2 (t / treat-04~e.0 :ARG2~e.1 (s2 / small-molecule~e.5 :name (n3 / name :op1 "U0126"~e.6) :ARG0-of~e.5 (i / inhibit-01~e.5 :ARG1 (p2 / protein-family :name (n4 / name :op1 "MEK"~e.3))))) :ARG1-of (c / complete-02~e.7) :ARG0-of (i4 / indicate-01~e.31 :ARG1~e.32 (e5 / event~e.41 :mod (e6 / early~e.38) :ARG1-of (d3 / direct-02~e.40) :domain~e.36 (i5 / induce-01~e.33 :ARG1~e.34 g~e.35) :ARG1-of (f3 / follow-01~e.43 :ARG2 (a2 / activate-01~e.45 :ARG1~e.46 (s3 / signal-07~e.48 :ARG0 (p / protein-family :name (n2 / name :op1 "ERK"~e.47)))))))) # ::id a_pmid_2094_2929.49 ::amr-annotator SDL-AMR-09 ::preferred # ::tok Since serpinE2 protein is known to be secreted [ @ 22 , 33 @ ] , we easily confirmed its presence in conditioned culture medium of caMEK @-@ expressing IECs whereas no serpinE2 protein was detected in the culture medium of wtMEK @-@ expressing or parental IECs ( Figure 1D ) . # ::alignments 0-1.5 1-1.2.1.1.1 2-1.2.1 4-1.5.1 7-1.5.1.1 10-1.5.2.1.1.1.1 14-1.5.2.1.1.1.2 18-1.1 19-1.4 20-1 22-1.2 23-1.2.2.r 24-1.2.2.2 25-1.2.2.1 26-1.2.2 30-1.2.2.3.2 31-1.2.2.3.1.1 32-1.6 33-1.6.1.1 33-1.6.1.1.r 34-1.6.1.2 35-1.6.1.2 37-1.6.1 38-1.6.1.3.r 40-1.6.1.3.1.1 40-1.6.1.3.2.1 41-1.6.1.3.1 41-1.6.1.3.2 45-1.6.1.3.1.2.2 46-1.6.1.3 47-1.6.1.3.2.2.2 48-1.6.1.3.1.2.1.1 48-1.6.1.3.2.2.1.1 50-1.3.1 52-1.3.1.1 (c / confirm-01~e.20 :ARG0 (w2 / we~e.18) :ARG1 (p / present-02~e.22 :ARG1 (p2 / protein~e.2 :name (n / name :op1 "serpinE2"~e.1)) :ARG2~e.23 (m / medium~e.26 :mod (c3 / culture~e.25) :ARG1-of (c4 / condition-01~e.24) :poss (c7 / cell :name (n4 / name :op1 "IEC"~e.31) :ARG3-of (e2 / express-03~e.30 :ARG2 (e / enzyme :name (n3 / name :op1 "MEK") :ARG2-of (m2 / mutate-01) :mod (c6 / constitutive) :ARG1-of (a3 / activate-01)))))) :ARG1-of (d2 / describe-01 :ARG0 (f / figure~e.50 :mod "1D"~e.52)) :ARG1-of (e5 / easy-05~e.19) :ARG0-of (c10 / cause-01~e.0 :ARG1 (k / know-01~e.4 :ARG1 (s2 / secrete-01~e.7 :ARG1 p2)) :ARG1-of (d3 / describe-01 :ARG0 (p5 / publication :ARG1-of (c9 / cite-01 :ARG2 (a2 / and :op1 22~e.10 :op2 33~e.14))))) :ARG1-of (c2 / contrast-01~e.32 :ARG2 (d / detect-01~e.37 :polarity~e.33 -~e.33 :ARG1 p2~e.34,35 :location~e.38 (o / or~e.46 :op1 (m3 / medium~e.41 :mod (c5 / culture-01~e.40) :poss (c12 / cell :name (n2 / name :op1 "IEC"~e.48) :ARG3-of (e4 / express-03~e.45 :ARG2 (e3 / enzyme :name (n5 / name :op1 "MEK") :mod (w / wild-type))))) :op2 (m4 / medium~e.41 :mod (c11 / culture-01~e.40) :poss (c8 / cell :name (n6 / name :op1 "IEC"~e.48) :mod (p4 / parent~e.47))))))) # ::id a_pmid_2094_2929.50 ::amr-annotator SDL-AMR-09 ::preferred # ::tok Again , treatment with the MEK @-@ inhibitor U0126 completely abrogated serpinE2 secretion ( Figure 1D ) . # ::alignments 0-1.3 2-1.2 3-1.2.1.r 5-1.2.1.2.1.1.1 7-1.2.1 7-1.2.1.2 7-1.2.1.2.r 8-1.2.1.1.1 9-1.5 10-1 11-1.1.1.1.1 12-1.1 14-1.4.1 16-1.4.1.1 (a / abrogate-01~e.10 :ARG1 (s2 / secrete-01~e.12 :ARG1 (p / protein :name (n3 / name :op1 "serpinE2"~e.11))) :ARG2 (t / treat-04~e.2 :ARG2~e.3 (s / small-molecule~e.7 :name (n / name :op1 "U0126"~e.8) :ARG0-of~e.7 (i / inhibit-01~e.7 :ARG1 (p2 / protein-family :name (n2 / name :op1 "MEK"~e.5))))) :mod (a2 / again~e.0) :ARG1-of (d / describe-01 :ARG0 (f / figure~e.14 :mod "1D"~e.16)) :ARG1-of (c / complete-02~e.9)) # ::id a_pmid_2094_2929.51 ::amr-annotator SDL-AMR-09 ::preferred # ::tok Interestingly , serpinE2 protein was difficult to detect in total cell lysates ( Figure 1E , lane 2 ) . # ::alignments 0-1.2 2-1.3.1.1.1 3-1.3.1 4-1.3.r 5-1 7-1.3 8-1.3.2.r 9-1.3.2.2 10-1.3.2.1 11-1.3.2 13-1.1.1.1 15-1.1.1.1.1 18-1.1.1.1.2 19-1.1.1.1.2.1 (d / difficult~e.5 :ARG1-of (d3 / describe-01 :ARG0 (a / and :op1 (f / figure~e.13 :mod "1E"~e.15 :part (l2 / lane~e.18 :mod 2~e.19)))) :mod (i / interesting~e.0) :domain~e.4 (d2 / detect-01~e.7 :ARG1 (p / protein~e.3 :name (n / name :op1 "serpinE2"~e.2)) :location~e.8 (l / lysate~e.11 :mod (c / cell~e.10) :mod (t / total~e.9)))) # ::id a_pmid_2094_2929.52 ::amr-annotator SDL-AMR-09 ::preferred # ::tok However , serpinE2 was easily observed in lysates prepared from foci of post @-@ confluent caMEK @-@ expressing cells ( Figure 1E , lane 4 ) , while it was not detectable in the surrounding monolayer ( Figure 1E , lane 3 ) . # ::alignments 0-1 0-1.1 0-1.1.r 2-1.1.1.1.1.1 3-1.1.1.3.1.1.1.1.1.1.r 4-1.1.1.2 5-1.1.1 6-1.1.1.3.r 7-1.1.1.3 8-1.1.1.3.1 9-1.1.1.3.1.1.r 10-1.1.1.3.1.1 11-1.1.1.3.1.1.1.r 12-1.1.1.3.1.1.1.1 14-1.1.1.3.1.1.1.1.1 17-1.1.1.3.1.1.1.2 18-1.1.1.3.1.1.1 20-1.1.1.4.1.1 22-1.1.1.4.1.1.1 25-1.1.1.4.1.1.2 26-1.1.1.4.1.1.2.1 29-1.1.1.3.1.1.1.1.r 30-1.1.2.1.2 31-1.1.1.3.1.1.1.1.1.1.r 32-1.1.2.1.1 32-1.1.2.1.1.r 33-1.1.2.1 34-1.1.2.1.3.r 36-1.1.2.1.3.1 37-1.1.2.1.3 39-1.1.2.1.4.1.1 41-1.1.2.1.4.1.1.1 44-1.1.2.1.4.1.1.2 45-1.1.2.1.4.1.1.2.1 (h / have-concession-91~e.0 :ARG1~e.0 (c / contrast-01~e.0 :ARG1 (o / observe-01~e.5 :ARG1 (p / protein :name (n / name :op1 "serpinE2"~e.2)) :ARG1-of (e / easy-05~e.4) :location~e.6 (l / lysate~e.7 :ARG1-of (p2 / prepare-01~e.8 :ARG2~e.9 (f / focus~e.10 :part-of~e.11 (c2 / cell~e.18 :time~e.29 (a / after~e.12 :op1 (c4 / confluent~e.14 :domain~e.3,31 c2)) :ARG3-of (e3 / express-03~e.17 :ARG2 (e2 / enzyme :name (n2 / name :op1 "MEK") :mod (c3 / constitutive) :ARG2-of (m / mutate-01) :ARG1-of (a4 / activate-01))))))) :ARG1-of (d / describe-01 :ARG0 (a2 / and :op1 (f2 / figure~e.20 :mod "1E"~e.22 :part (l2 / lane~e.25 :mod 4~e.26))))) :ARG2 (p4 / possible-01 :ARG1 (d2 / detect-01~e.33 :polarity~e.32 -~e.32 :ARG1 p~e.30 :location~e.34 (m2 / monolayer~e.37 :ARG1-of (s / surround-01~e.36)) :ARG1-of (d3 / describe-01 :ARG0 (a3 / and :op1 (f3 / figure~e.39 :mod "1E"~e.41 :part (l3 / lane~e.44 :mod 3~e.45)))))))) # ::id a_pmid_2094_2929.53 ::amr-annotator SDL-AMR-09 ::preferred # ::tok This indicates a stronger expression of serpinE2 protein by the transformed IECs forming the foci . # ::alignments 0-1.1 1-1 3-1.2.3 3-1.2.3.1 3-1.2.3.1.r 4-1.2 5-1.2.1.r 6-1.2.1.1.1 7-1.2.1 8-1.2.2.r 10-1.2.2.2 11-1.2.2.1.1 12-1.2.2.3 14-1.2.2.3.1 (i / indicate-01~e.1 :ARG0 (t / this~e.0) :ARG1 (e / express-03~e.4 :ARG2~e.5 (p / protein~e.7 :name (n / name :op1 "serpinE2"~e.6)) :ARG3~e.8 (c / cell :name (n2 / name :op1 "IEC"~e.11) :ARG1-of (t2 / transform-01~e.10) :ARG0-of (f / form-01~e.12 :ARG1 (f2 / focus~e.14))) :ARG1-of (s / strong-02~e.3 :degree~e.3 (m / more~e.3)))) # ::id a_pmid_2094_2929.54 ::amr-annotator SDL-AMR-09 ::preferred # ::tok Gene silencing of serpinE2 decreases foci formation , growth in soft agarose and migration induced by activated MEK # ::alignments 0-1.1.2 1-1.1 2-1.1.1.r 3-1.1.1.1.1 4-1 5-1.2.1.1 6-1.2.1 8-1.2.2 9-1.2.2.1.r 10-1.2.2.1.1 11-1.2.2.1 12-1.2 13-1.2.3 14-1.2.3.1 15-1.2.3.1.1.r 16-1.2.3.1.1.2 17-1.2.3.1.1.1.1 (d / decrease-01~e.4 :ARG0 (s / silence-01~e.1 :ARG1~e.2 (p / protein :name (n2 / name :op1 "serpinE2"~e.3)) :mod (g / gene~e.0)) :ARG1 (a / and~e.12 :op1 (f / form-01~e.6 :ARG1 (f2 / focus~e.5)) :op2 (g2 / grow-01~e.8 :location~e.9 (a3 / agarose~e.11 :ARG1-of (s2 / soft-02~e.10))) :op3 (m2 / migrate-01~e.13 :ARG2-of (i / induce-01~e.14 :ARG0~e.15 (e2 / enzyme :name (n4 / name :op1 "MEK"~e.17) :ARG1-of (a2 / activate-01~e.16)))))) # ::id a_pmid_2094_2929.55 ::amr-annotator SDL-AMR-09 ::preferred # ::tok In order to determine the contribution of serpinE2 in intestinal transformation induced by activated MEK , foci from post @-@ confluent caMEK @-@ expressing IECs were retrieved by aspiration with a pipette and pooled as one caMEK @-@ expressing cell population . # ::alignments 0-1.3.r 1-1.3.r 2-1.3.r 3-1.3 5-1.3.1 6-1.3.1.1.r 7-1.3.1.1.1.1 8-1.3.1.2.r 9-1.3.1.2.1 10-1.3.1.2 11-1.3.1.2.2 13-1.1.2.2.1.4 13-1.3.1.2.2.1.2 14-1.1.2.2.1.1.1 14-1.3.1.2.2.1.1.1 16-1.1.1 18-1.1.2.3 20-1.1.2.3.1 23-1.1.2.2 24-1.1.2.1.1 25-1.1.2.3.1.1.r 26-1.1 27-1.1.3.r 28-1.1.3 29-1.1.3.3.r 31-1.1.3.3 32-1 33-1.2 34-1.1.2.3.r 38-1.2.2.1.1 39-1.2.2.1 40-1.2.2 (a5 / and~e.32 :op1 (r / retrieve-01~e.26 :ARG1 (f / focus~e.16) :ARG2 (c2 / cell :name (n3 / name :op1 "IEC"~e.24) :ARG3-of (e2 / express-03~e.23 :ARG2 (e3 / enzyme :name (n4 / name :op1 "MEK"~e.14) :mod (c3 / constitutive) :ARG2-of (m / mutate-01) :ARG1-of (a4 / activate-01~e.13))) :time~e.34 (a2 / after~e.18 :op1 (c5 / confluent~e.20 :domain~e.25 c2))) :manner~e.27 (a3 / aspirate-101~e.28 :ARG1 f :ARG2 c2 :instrument~e.29 (p3 / pipette~e.31))) :op2 (p4 / pool-01~e.33 :ARG1 f :ARG2 (p5 / population~e.40 :mod (c4 / cell~e.39 :ARG3-of (e4 / express-03~e.38 :ARG2 e3)))) :purpose~e.0,1,2 (d / determine-01~e.3 :ARG1 (c / contribute-01~e.5 :ARG0~e.6 (p / protein :name (n2 / name :op1 "serpinE2"~e.7)) :ARG2~e.8 (t / transform-01~e.10 :mod (i / intestine~e.9) :ARG2-of (i2 / induce-01~e.11 :ARG0 (e / enzyme :name (n / name :op1 "MEK"~e.14) :ARG1-of (a / activate-01~e.13))))))) # ::id a_pmid_2094_2929.56 ::amr-annotator SDL-AMR-09 ::preferred # ::tok All further experiments were performed with this previously characterized caMEK @-@ expressing IEC population [ @ 14 @ ] and compared with wtMEK @-@ expressing cell populations . # ::alignments 0-1.1.1.3 1-1.1.1.2 2-1.1.1 4-1.1 5-1.1.1.1.r 6-1.1.1.1.3 7-1.1.1.1.1.1 8-1.1.1.1.1 11-1.1.1.1.2.2 12-1.1.1.1.2.1.1 13-1.1.1.1 16-1.1.2.1.1.1 19-1 20-1.2 24-1.2.2.1.1 25-1.2.2.1 26-1.2.2 (a3 / and~e.19 :op1 (p / perform-02~e.4 :ARG1 (e / experiment-01~e.2 :ARG1~e.5 (p2 / population~e.13 :ARG1-of (c / characterize-01~e.8 :time (p3 / previous~e.7)) :mod (c2 / cell :name (n / name :op1 "IEC"~e.12) :ARG3-of (e2 / express-03~e.11 :ARG2 (e3 / enzyme :name (n2 / name :op1 "MEK") :mod (c3 / constitutive) :ARG2-of (m / mutate-01) :ARG1-of (a4 / activate-01)))) :mod (t / this~e.6)) :degree (f / further~e.1) :mod (a / all~e.0)) :ARG1-of (d / describe-01 :ARG0 (p4 / publication :ARG1-of (c4 / cite-01 :ARG2 14~e.16)))) :op2 (c5 / compare-01~e.20 :ARG1 e :ARG2 (p5 / population~e.26 :mod (c6 / cell~e.25 :ARG3-of (e4 / express-03~e.24 :ARG2 (e5 / enzyme :name (n3 / name :op1 "MEK") :mod (w / wild-type))))))) # ::id a_pmid_2094_2929.57 ::amr-annotator SDL-AMR-09 ::preferred # ::tok Recombinant lentiviruses encoding anti @- @ serpinE2 @ short hairpin RNA ( shRNA ) were therefore developed to stably suppress serpinE2 levels in these cells . # ::alignments 0-1.1.1.1 2-1.1.1.2 2-1.1.1.2.1.2 3-1.1.1.2.1.2.1.1 6-1.1.1.2.1.2.1.1.1.1.1 8-1.1.1.2.1.1.1 9-1.1.1.2.1.1.2 10-1.1.1.2.1 10-1.1.1.2.1.3.1 12-1.1.1.2.1.3.1.1.1 15-1 16-1.1 18-1.1.2.2 19-1.1.2 20-1.1.2.1.1 21-1.1.2.1 22-1.1.2.3.r 23-1.1.2.3.1 24-1.1.2.3 (c / cause-01~e.15 :ARG1 (d / develop-02~e.16 :ARG1 (l / lentivirus :ARG3-of (r / recombine-01~e.0) :ARG0-of (e / encode-01~e.2 :ARG1 (n4 / nucleic-acid~e.10 :name (n / name :op1 "short"~e.8 :op2 "hairpin"~e.9) :ARG0-of (e2 / encode-01~e.2 :ARG1 (a / antibody :ARG0-of (o / oppose-01~e.3 :ARG1 (p / protein :name (n2 / name :op1 "serpinE2"~e.6))))) :ARG1-of (m / mean-01 :ARG2 (n5 / nucleic-acid~e.10 :name (n3 / name :op1 "shRNA"~e.12)))))) :ARG4 (s / suppress-01~e.19 :ARG1 (l2 / level~e.21 :quant-of p~e.20) :ARG1-of (s2 / stable-03~e.18) :location~e.22 (c2 / cell~e.24 :mod (t / this~e.23))))) # ::id a_pmid_2094_2929.58 ::amr-annotator SDL-AMR-09 ::preferred # ::tok Several lentiviral constructs were generated and tested for their ability to knock down serpinE2 protein . # ::alignments 0-1.1.1.2 1-1.1.1.1 2-1.1.1 4-1.1 5-1 6-1.2 9-1.2.2 10-1.2.2.2.r 11-1.2.2.2 12-1.2.2.2 13-1.2.2.2.2.1.1 14-1.2.2.2.2 (a / and~e.5 :op1 (g / generate-01~e.4 :ARG1 (c / construct-01~e.2 :ARG1 (l / lentiviral~e.1) :quant (s / several~e.0))) :op2 (t / test-01~e.6 :ARG1 c :ARG2 (c2 / capable-01~e.9 :ARG1 c :ARG2~e.10 (k / knock-down-02~e.11,12 :ARG0 c :ARG1 (p / protein~e.14 :name (n / name :op1 "serpinE2"~e.13)))))) # ::id a_pmid_2094_2929.59 ::amr-annotator SDL-AMR-09 ::preferred # ::tok One of these viral shRNAs was selected and designated as shSerpinE2 . # ::alignments 2-1.1.1.1.1.3 3-1.1.1.1.1.2 4-1.1.1.1.1.1.1 4-1.2.2.1.1 6-1.1 7-1 8-1.2 (a / and~e.7 :op1 (s / select-01~e.6 :ARG1 (n6 / nucleic-acid :ARG1-of (i / include-91 :ARG2 (n4 / nucleic-acid :name (n / name :op1 "shRNA"~e.4) :mod (v / virus~e.3) :mod (t / this~e.2))))) :op2 (d / designate-01~e.8 :ARG1 n6 :ARG2 (n5 / nucleic-acid :name (n2 / name :op1 "shRNA"~e.4) :ARG0-of (e / encode-01 :ARG1 (p / protein :name (n3 / name :op1 "serpinE2")))))) # ::id a_pmid_2094_2929.60 ::amr-annotator SDL-AMR-09 ::preferred # ::tok caMEK @-@ expressing cells were henceforth infected with shSerpinE2 lentiviruses or with lentiviruses expressing a control shRNA ( shScrambled ) . # ::alignments 2-1.1.1 3-1.1 5-1.3 6-1 10-1.2 13-1.2.2.1 15-1.2.2.1.1 15-1.2.2.1.1.2 15-1.2.2.1.1.2.r 16-1.2.1.1.1.1 16-1.2.2.1.1.1.1 18-1.2.2.1.1.3.1.1.1 (i / infect-01~e.6 :ARG1 (c / cell~e.3 :ARG3-of (e / express-03~e.2 :ARG2 (e2 / enzyme :name (n / name :op1 "MEK") :mod (c2 / constitutive) :ARG2-of (m / mutate-01) :ARG1-of (a2 / activate-01)))) :ARG2 (o / or~e.10 :op1 (l / lentivirus :mod (n6 / nucleic-acid :name (n2 / name :op1 "shRNA"~e.16) :ARG0-of (e3 / encode-01 :ARG1 (p / protein :name (n3 / name :op1 "serpinE2"))))) :op2 (l2 / lentivirus :ARG1-of (e4 / express-03~e.13 :ARG2 (n7 / nucleic-acid~e.15 :name (n4 / name :op1 "shRNA"~e.16) :ARG0-of~e.15 (c3 / control-01~e.15) :ARG1-of (m2 / mean-01 :ARG2 (n8 / nucleic-acid :name (n5 / name :op1 "shScrambled"~e.18))))))) :time (h / henceforth~e.5)) # ::id a_pmid_2094_2929.61 ::amr-annotator SDL-AMR-09 ::preferred # ::tok Secretion of serpinE2 protein was analyzed 14 days after selection with blasticidin S in these populations . # ::alignments 0-1.1 1-1.1.1.r 2-1.1.1.1.1 3-1.1.1 5-1 6-1.2.2.1 7-1.2.2.2 8-1.2 9-1.2.1 10-1.2.1.2.r 11-1.2.1.2.1.1 12-1.2.1.2.1.2 13-1.2.1.1.r 14-1.2.1.1.1 15-1.2.1.1 (a / analyze-01~e.5 :ARG1 (s / secrete-01~e.0 :ARG1~e.1 (p / protein~e.3 :name (n / name :op1 "serpinE2"~e.2))) :time (a2 / after~e.8 :op1 (s2 / select-01~e.9 :ARG2~e.13 (p2 / population~e.15 :mod (t2 / this~e.14)) :instrument~e.10 (s3 / small-molecule :name (n2 / name :op1 "blasticidin"~e.11 :op2 "S"~e.12))) :quant (t / temporal-quantity :quant 14~e.6 :unit (d2 / day~e.7)))) # ::id a_pmid_2094_2929.62 ::amr-annotator SDL-AMR-09 ::preferred # ::tok As shown in Figure 2A , secreted serpinE2 levels were markedly reduced (> 60 %) in cells @-@ expressing shSerpinE2 ; in contrast , shScrambled had no effect on the secretion of serpinE2 ( data not shown ) . # ::alignments 1-1.1.5 2-1.1.5.1.r 3-1.1.5.1 5-1.1.5.1.1 8-1.1.1.2 9-1.1.1.1.1.1 10-1.1.1 12-1.1.3 12-1.1.3.r 13-1.1 15-1.1.2.1.1 17-1.1.4.r 18-1.1.4 20-1.1.4.1 24-1.2 26-1.2.1.2.1.1 28-1.2.1.1 28-1.2.1.1.r 29-1.2.1 30-1.2.1.3.r 32-1.2.1.3 33-1.2.1.3.1.r 34-1.2.1.3.1 36-1.2.2.1 37-1.2.2.1.1.1 37-1.2.2.1.1.1.r 38-1.2.2.1.1 (a / and :op1 (r / reduce-01~e.13 :ARG1 (l / level~e.10 :quant-of (p2 / protein :name (n / name :op1 "serpinE2"~e.9)) :ARG1-of (s / secrete-01~e.8)) :ARG2 (m2 / more-than :op1 (p / percentage-entity :value 60~e.15)) :manner~e.12 (m / marked~e.12) :location~e.17 (c / cell~e.18 :ARG3-of (e2 / express-03~e.20 :ARG2 (n4 / nucleic-acid :name (n2 / name :op1 "shRNA") :ARG0-of (e / encode-01 :ARG1 p2)))) :ARG1-of (s3 / show-01~e.1 :ARG0~e.2 (f / figure~e.3 :mod "2A"~e.5))) :op2 (c2 / contrast-01~e.24 :ARG2 (a2 / affect-01~e.29 :polarity~e.28 -~e.28 :ARG0 (n5 / nucleic-acid :name (n3 / name :op1 "shScrambled"~e.26)) :ARG1~e.30 (s2 / secrete-01~e.32 :ARG1~e.33 p2~e.34)) :ARG1-of (d / describe-01 :ARG0 (d2 / data~e.36 :ARG1-of (s4 / show-01~e.38 :polarity~e.37 -~e.37))))) # ::id a_pmid_2094_2929.63 ::amr-annotator SDL-AMR-09 ::preferred # ::tok To determine the functional role of serpinE2 in caMEK @-@ expressing cells , the proliferation rate of these cell populations was assessed when cultured on plastic . # ::alignments 1-1.3 3-1.3.1.2 4-1.3.1 5-1.3.1.1.r 6-1.3.1.1.1.1 10-1.3.1.3.1 11-1.3.1.3 14-1.1.1 15-1.1 16-1.1.1.1.r 17-1.1.1.1.2 18-1.1.1.1.1 19-1.1.1.1 21-1 22-1.2.r 23-1.2 24-1.2.2.r 25-1.2.2 (a3 / assess-01~e.21 :ARG1 (r2 / rate~e.15 :degree-of (p2 / proliferate-01~e.14 :ARG0~e.16 (p3 / population~e.19 :mod (c3 / cell~e.18) :mod (t / this~e.17)))) :time~e.22 (c4 / culture-01~e.23 :ARG1 p3 :manner~e.24 (p4 / plastic~e.25)) :purpose (d / determine-01~e.1 :ARG1 (r / role~e.4 :topic~e.5 (p / protein :name (n / name :op1 "serpinE2"~e.6)) :ARG1-of (f / function-01~e.3) :location (c / cell~e.11 :ARG3-of (e / express-03~e.10 :ARG2 (e2 / enzyme :name (n2 / name :op1 "MEK") :mod (c2 / constitutive) :ARG2-of (m / mutate-01) :ARG1-of (a2 / activate-01))))))) # ::id a_pmid_2094_2929.64 ::amr-annotator SDL-AMR-09 ::preferred # ::tok No difference was observed in the proliferation rate of subconfluent caMEK @-@ expressing cells when serpinE2 expression was downregulated ( Figure 2B ) . # ::alignments 0-1.1.1 0-1.1.1.r 1-1.1 1-1.1.1 1-1.1.1.r 3-1 4-1.1.2.r 6-1.1.2.1 7-1.1.2 8-1.1.2.1.1.r 9-1.1.2.1.1.2 10-1.1.2.1.1.1.1.1.1 12-1.1.2.1.1.1 13-1.1.2.1.1 14-1.2.r 15-1.2.1.1.1.1 16-1.2.1 18-1.2 20-1.3.1 22-1.3.1.1 (o / observe-01~e.3 :ARG1 (d / differ-02~e.1 :polarity~e.0,1 -~e.0,1 :ARG1~e.4 (r / rate~e.7 :degree-of (p / proliferate-01~e.6 :ARG0~e.8 (c / cell~e.13 :ARG3-of (e / express-03~e.12 :ARG2 (e2 / enzyme :name (n / name :op1 "caMEK"~e.10))) :mod (s / subconfluent~e.9))))) :time~e.14 (d2 / downregulate-01~e.18 :ARG1 (e3 / express-03~e.16 :ARG2 (p2 / protein :name (n2 / name :op1 "serpinE2"~e.15)))) :ARG1-of (d3 / describe-01 :ARG0 (f / figure~e.20 :mod "2B"~e.22))) # ::id a_pmid_2094_2929.65 ::amr-annotator SDL-AMR-09 ::preferred # ::tok In a previous study , we had shown that expression of activated MEK in intestinal epithelial cells resulted in loss of cell @-@ cell contact growth inhibition and produced colonies or multilayered domes which grew to increased saturation density and formed tumors when transplanted into nude mice [ @ 14 @ ] . # ::alignments 2-1.3.1 3-1.3 5-1.1 7-1 8-1.2.r 9-1.2.1.1 10-1.2.1.1.1.r 11-1.2.1.1.1.2 12-1.2.1.1.1.1.1 13-1.2.1.1.2.r 14-1.2.1.1.2.1.1 15-1.2.1.1.2.1 16-1.2.1.1.2 17-1.2.1 18-1.2.1.2.r 19-1.2.1.2 20-1.2.1.2.1.r 21-1.2.1.2.1.1.1 23-1.2.1.2.1.1.1 23-1.2.1.2.1.1.2 24-1.2.1.2.1.1 25-1.2.1.2.1.2 26-1.2.1.2.1 27-1.2 28-1.2.2 29-1.2.2.2.1 30-1.2.2.2 31-1.2.2.2.2.1 32-1.2.2.2.2 34-1.2.2.2.3 35-1.2.2.2.3.1.r 36-1.2.2.2.3.1.2 37-1.2.2.2.3.1.1 38-1.2.2.2.3.1 40-1.2.2.2.4 41-1.2.2.2.4.1 42-1.2.2.2.4.2.r 43-1.2.2.2.4.2 44-1.2.2.2.4.2.2.r 45-1.2.2.2.4.2.2.1 46-1.2.2.2.4.2.2 49-1.4.1.1.1 (s / show-01~e.7 :ARG0 (w / we~e.5) :ARG1~e.8 (a2 / and~e.27 :op1 (r / result-01~e.17 :ARG1 (e / express-03~e.9 :ARG2~e.10 (e2 / enzyme :name (n / name :op1 "MEK"~e.12) :ARG1-of (a / activate-01~e.11)) :ARG3~e.13 (c / cell~e.16 :source (e3 / epithelium~e.15 :part-of (i / intestine~e.14)))) :ARG2~e.18 (l / lose-02~e.19 :ARG1~e.20 (i2 / inhibit-01~e.26 :ARG0 (c2 / contact-01~e.24 :ARG0 (c4 / cell~e.21,23) :ARG1 (c3 / cell~e.23)) :ARG1 (g / grow-01~e.25)))) :op2 (p / produce-01~e.28 :ARG0 e :ARG1 (o / or~e.30 :op1 (c5 / colony~e.29) :op2 (d / dome~e.32 :mod (m / multilayered~e.31)) :ARG1-of (g2 / grow-01~e.34 :ARG4~e.35 (d2 / density~e.38 :mod (s2 / saturate-01~e.37) :ARG1-of (i3 / increase-01~e.36))) :ARG0-of (f / form-01~e.40 :ARG1 (t / tumor~e.41) :time~e.42 (t2 / transplant-01~e.43 :ARG1 o :ARG2~e.44 (m2 / mouse~e.46 :mod (n2 / nude~e.45))))))) :medium (s3 / study-01~e.3 :time (p2 / previous~e.2)) :ARG1-of (d3 / describe-01 :ARG0 (p3 / publication :ARG1-of (c6 / cite-01 :ARG2 14~e.49)))) # ::id a_pmid_2094_2929.66 ::amr-annotator SDL-AMR-09 ::preferred # ::tok Of note , focus formation assays performed herein revealed that initially , there was little difference in the number of foci obtained between control cells and serpinE2 @-@ depleted cells ( data not shown ) . # ::alignments 1-1.3 3-1.1.1.1 4-1.1.1 5-1.1 6-1.1.2 7-1.1.2.1 8-1 9-1.2.r 10-1.2.5 14-1.2.4 15-1.2 16-1.2.3.r 18-1.2.3 19-1.2.3.1.r 20-1.2.3.1 21-1.2.3.2 23-1.2.1.1 24-1.2.1 26-1.2.2.1.1.1.1 28-1.2.2.1 29-1.2.2 31-1.4.1 32-1.4.1.1.1 32-1.4.1.1.1.r 33-1.4.1.1 (r / reveal-01~e.8 :ARG0 (a / assay-01~e.5 :ARG1 (f2 / form-01~e.4 :ARG1 (f3 / focus~e.3)) :ARG1-of (p / perform-02~e.6 :medium (h / herein~e.7))) :ARG1~e.9 (d / differ-02~e.15 :ARG1 (c / cell~e.24 :mod (c2 / control~e.23)) :ARG2 (c3 / cell~e.29 :ARG1-of (d2 / deplete-01~e.28 :ARG2 (p2 / protein :name (n3 / name :op1 "serpinE2"~e.26)))) :ARG3~e.16 (n2 / number~e.18 :quant-of~e.19 (f / focus~e.20) :ARG1-of (o / obtain-01~e.21)) :degree (l / little~e.14) :time (i / initial~e.10)) :ARG1-of (n4 / note-02~e.1) :ARG1-of (d3 / describe-01 :ARG0 (d4 / data~e.31 :ARG1-of (s / show-01~e.33 :polarity~e.32 -~e.32)))) # ::id a_pmid_2094_2929.67 ::amr-annotator SDL-AMR-09 ::preferred # ::tok However , serpinE2 @ silencing markedly reduced the size of foci ( Figure 2C ) suggesting a reduced capacity of these foci to grow . # ::alignments 0-1 3-1.1.1.1.1.1 5-1.1.1 6-1.1.3 6-1.1.3.r 7-1.1 9-1.1.2 10-1.1.2.1.r 11-1.1.2.1 13-1.1.4.1 15-1.1.4.1.1 18-1.1.5 20-1.1.5.1.3 21-1.1.5.1 24-1.1.5.1.2.1 26-1.1.5.1.2 (c2 / contrast-01~e.0 :ARG2 (r / reduce-01~e.7 :ARG0 (s / silence-01~e.5 :ARG1 (g2 / gene :name (n / name :op1 "serpinE2"~e.3))) :ARG1 (s2 / size~e.9 :poss~e.10 (f / focus~e.11)) :manner~e.6 (m / marked~e.6) :ARG1-of (d / describe-01 :ARG0 (f2 / figure~e.13 :mod "2C"~e.15)) :ARG0-of (s3 / suggest-01~e.18 :ARG1 (c / capable-01~e.21 :ARG1 f :ARG2 (g / grow-01~e.26 :ARG1 f~e.24) :ARG1-of (r2 / reduce-01~e.20))))) # ::id a_pmid_2094_2929.68 ::amr-annotator SDL-AMR-09 ::preferred # ::tok Indeed , phase @-@ contrast microscopy revealed that the colonies were smaller when serpinE2 was downregulated ( Figure 2D ) . # ::alignments 0-1.4 2-1.1.1.1 4-1.1.1.1 5-1.1.1.2 6-1 7-1.2.r 9-1.2.2 10-1.2.2.r 11-1.2 11-1.2.1 11-1.2.1.r 12-1.2.3.r 13-1.2.3.1.1.1 15-1.2.3 17-1.3.1 19-1.3.1.1 (r / reveal-01~e.6 :ARG0 (t / thing :name (n / name :op1 "phase-contrast"~e.2,4 :op2 "microscopy"~e.5)) :ARG1~e.7 (s / small~e.11 :degree~e.11 (m / more~e.11) :domain~e.10 (c / colony~e.9) :time~e.12 (d / downregulate-01~e.15 :ARG1 (p / protein :name (n2 / name :op1 "serpinE2"~e.13)))) :ARG1-of (d2 / describe-01 :ARG0 (f / figure~e.17 :mod "2D"~e.19)) :mod (i / indeed~e.0)) # ::id a_pmid_2094_2929.69 ::amr-annotator SDL-AMR-09 ::preferred # ::tok Finally , expression of shSerpinE2 led to a significant decrease in the ability of caMEK @-@ expressing cells to grow under anchorage @-@ independent conditions in soft agarose ( Figure 2E ) . # ::alignments 0-1.1 0-1.1.r 2-1.2 3-1.2.1.r 4-1.2.1.1.1 5-1 8-1.3.2 9-1.3 10-1.3.1.r 12-1.3.1 13-1.3.1.1.r 14-1.3.1.1.1.1.1.1 16-1.3.1.1.1 17-1.3.1.1 19-1.3.1.2 21-1.3.1.2.3.3 23-1.3.1.2.3 23-1.3.1.2.3.1 23-1.3.1.2.3.1.r 24-1.3.1.2.3.r 25-1.3.1.2.2.r 26-1.3.1.2.2.1 27-1.3.1.2.2 29-1.4.1 31-1.4.1.1 (l / lead-03~e.5 :li~e.0 -1~e.0 :ARG0 (e / express-03~e.2 :ARG2~e.3 (p / protein :name (n / name :op1 "shSerpinE2"~e.4))) :ARG1 (d / decrease-01~e.9 :ARG1~e.10 (c / capable-01~e.12 :ARG1~e.13 (c2 / cell~e.17 :ARG3-of (e2 / express-03~e.16 :ARG2 (e3 / enzyme :name (n2 / name :op1 "caMEK"~e.14)))) :ARG2 (g / grow-01~e.19 :ARG1 c2 :location~e.25 (a2 / agarose~e.27 :ARG1-of (s2 / soft-02~e.26)) :condition~e.24 (d2 / depend-01~e.23 :polarity~e.23 -~e.23 :ARG0 g :ARG1 (a / anchorage~e.21)))) :ARG2 (s / significant-02~e.8)) :ARG1-of (d3 / describe-01 :ARG0 (f / figure~e.29 :mod "2E"~e.31))) # ::id a_pmid_2094_2929.70 ::amr-annotator SDL-AMR-09 ::preferred # ::tok Cell migration is an important process of tumorigenesis and metastasis . # ::alignments 0-1.1.1 1-1.1 4-1.2 5-1 6-1.3.r 7-1.3.1 7-1.3.1.1 7-1.3.1.1.r 8-1.3 9-1.3.2 (p / process-02~e.5 :ARG1 (m / migrate-01~e.1 :ARG0 (c / cell~e.0)) :mod (i / important~e.4) :subevent-of~e.6 (a / and~e.8 :op1 (c2 / create-01~e.7 :ARG1~e.7 (t / tumor~e.7)) :op2 (m2 / metastasis~e.9))) # ::id a_pmid_2094_2929.71 ::amr-annotator SDL-AMR-09 ::preferred # ::tok Moreover , we recently reported that intestinal epithelial cells expressing activated MEK1 clearly acquire an increased capacity to migrate as compared to wtMEK @-@ expressing cells [ @ 14 @ ] . # ::alignments 0-1 2-1.1.1 3-1.1.3 4-1.1 5-1.1.2.r 6-1.1.2.1.1.1 7-1.1.2.1.1 8-1.1.2.1 8-1.1.2.2.4.1 9-1.1.2.1.2 9-1.1.2.2.4.1.1 10-1.1.2.1.2.1.2 11-1.1.2.1.2.1.1.1 12-1.1.2.3 13-1.1.2 15-1.1.2.2.3 16-1.1.2.2 16-1.1.2.2.4 18-1.1.2.2.2 19-1.1.3.r 20-1.1.2.2.4.r 24-1.1.2.2.1 25-1.1.2.2.1 28-1.2.1.1.1 (a / and~e.0 :op2 (r / report-01~e.4 :ARG0 (w / we~e.2) :ARG1~e.5 (a2 / acquire-01~e.13 :ARG0 (c2 / cell~e.8 :source (e / epithelium~e.7 :part-of (i / intestine~e.6)) :ARG3-of (e2 / express-03~e.9 :ARG2 (e3 / enzyme :name (n / name :op1 "MEK1"~e.11) :ARG1-of (a3 / activate-01~e.10)))) :ARG1 (c3 / capable-01~e.16 :ARG1 c2~e.24,25 :ARG2 (m / migrate-01~e.18 :ARG0 c2) :ARG1-of (i2 / increase-01~e.15) :compared-to~e.20 (c4 / capable-01~e.16 :ARG1 (c5 / cell~e.8 :ARG3-of (e4 / express-03~e.9 :ARG2 (e5 / enzyme :name (n2 / name :op1 "MEK") :mod (w2 / wild-type)))) :ARG2 m)) :ARG1-of (c / clear-06~e.12)) :time~e.19 (r2 / recent~e.3)) :ARG1-of (d / describe-01 :ARG0 (p / publication :ARG1-of (c6 / cite-01 :ARG2 14~e.28)))) # ::id a_pmid_2094_2929.72 ::amr-annotator SDL-AMR-09 ::preferred # ::tok Herein , in an in vitro @ transwell migration assay , serpinE2 deficiency significantly reduced caMEK @-@ expressing IEC migration to the undersurface of the polycarbonate membrane of Boyden chambers coated with fibronectin or vitronectin ( Figure 2F ) , two extracellular matrix proteins which can interact with serpinE2 [ @ 34 , 35 @ ] . # ::alignments 0-1.4 2-1.6.2 5-1.6.2 6-1.6.2 8-1.6.1.1 9-1.6.1 10-1.6 12-1.1.1.1.1 14-1.3 15-1 16-1.2.1.2.1.1.1 18-1.2.1.2 19-1.2.1.1.1 20-1.2 21-1.2.2.r 23-1.2.2 24-1.2.2.1.r 26-1.2.2.1.1 27-1.2.2.1 28-1.2.2.1.2.r 29-1.2.2.1.2.1.1 30-1.2.2.1.2.1.2 31-1.2.2.2 32-1.2.2.2.1.r 33-1.2.2.2.1.1.1.1 34-1.2.2.2.1 35-1.2.2.2.1.2.1.1 37-1.5.1 39-1.5.1.1 43-1.2.2.2.1.3.1.1 44-1.2.2.2.1.3.1.4.1 45-1.2.2.2.1.3.1.4 46-1.2.2.2.1.3.1 48-1.2.2.2.1.3.1.3.2 49-1.2.2.2.1.3.1.3 50-1.2.2.2.1.3.1.3.1.r 51-1.2.2.2.1.3.1.3.1 54-1.2.2.2.1.3.1.2.1.1.1.1 58-1.2.2.2.1.3.1.2.1.1.1.2 (r / reduce-01~e.15 :ARG0 (l / lack-01 :ARG1 (p / protein :name (n / name :op1 "serpinE2"~e.12))) :ARG1 (m / migrate-01~e.20 :ARG0 (c / cell :name (n2 / name :op1 "IEC"~e.19) :ARG3-of (e / express-03~e.18 :ARG2 (e2 / enzyme :name (n3 / name :op1 "caMEK"~e.16)))) :ARG1~e.21 (u / undersurface~e.23 :part-of~e.24 (m2 / membrane~e.27 :consist-of (p2 / polycarbonate~e.26) :poss~e.28 (t / thing :name (n4 / name :op1 "Boyden"~e.29 :op2 "chamber"~e.30))) :ARG1-of (c2 / coat-01~e.31 :ARG2~e.32 (o / or~e.34 :op1 (p3 / protein :name (n5 / name :op1 "fibronectin"~e.33)) :op2 (p4 / protein :name (n6 / name :op1 "vitronectin"~e.35)) :ARG1-of (m3 / mean-01 :ARG2 (p5 / protein~e.46 :quant 2~e.43 :ARG1-of (d / describe-01 :ARG0 (p7 / publication :ARG1-of (c3 / cite-01 :ARG2 (a / and :op1 34~e.54 :op2 35~e.58)))) :ARG0-of (i / interact-01~e.49 :ARG1~e.50 p~e.51 :ARG1-of (p6 / possible-01~e.48)) :mod (m4 / matrix~e.45 :mod (e3 / extracellular~e.44)))))))) :ARG2 (s / significant-02~e.14) :medium (h / herein~e.0) :ARG1-of (d2 / describe-01 :ARG0 (f / figure~e.37 :mod "2F"~e.39)) :condition (a2 / assay-01~e.10 :ARG1 (m5 / migrate-01~e.9 :mod (t3 / transwell~e.8)) :manner (i2 / in-vitro~e.2,5,6))) # ::id a_pmid_2094_2929.73 ::amr-annotator SDL-AMR-09 ::preferred # ::tok Taken together , these results support a role of serpinE2 in MEK1 @-@ induced transformation whereby serpinE2 activates anchorage @-@ independent growth and cell migration . # ::alignments 0-1.2 1-1.2.2 3-1.1.1.1 4-1.1.1 4-1.1.1.2 4-1.1.1.2.r 5-1.1 7-1.1.2 8-1.1.2.1.r 9-1.1.2.1.1.1 10-1.1.2.2.r 11-1.1.2.2.1.1.1.1 13-1.1.2.2.1 14-1.1.2.2 16-1.1.2.2.2.1 17-1.1.2.2.2 18-1.1.2.2.2.2.1.2.2 20-1.1.2.2.2.2.1.2 20-1.1.2.2.2.2.1.2.1 20-1.1.2.2.2.2.1.2.1.r 21-1.1.2.2.2.2.1 23-1.1.2.2.2.2.1.1 24-1.1.2.2.2.2.2 (h / have-condition-91 :ARG1 (s / support-01~e.5 :ARG0 (t / thing~e.4 :mod (t2 / this~e.3) :ARG2-of~e.4 (r / result-01~e.4)) :ARG1 (r2 / role~e.7 :poss~e.8 (p / protein :name (n / name :op1 "serpinE2"~e.9)) :purpose~e.10 (t3 / transform-01~e.14 :ARG2-of (i / induce-01~e.13 :ARG0 (e / enzyme :name (n2 / name :op1 "MEK1"~e.11))) :subevent (a / activate-01~e.17 :ARG0 p~e.16 :ARG1 (a2 / and :op1 (g / grow-01~e.21 :ARG1 (c2 / cell~e.23) :ARG0-of (d / depend-01~e.20 :polarity~e.20 -~e.20 :ARG1 (a3 / anchorage~e.18))) :op2 (m / migrate-01~e.24 :ARG0 c2)))))) :ARG2 (t4 / take-01~e.0 :ARG1 t :mod (t5 / together~e.1))) # ::id a_pmid_2094_2929.74 ::amr-annotator SDL-AMR-09 ::preferred # ::tok Expression of serpinE2 in colorectal cancer cells is dependent on MEK @/@ ERK activity # ::alignments 0-1.1 1-1.1.1.r 2-1.1.1.1.1 3-1.1.2.r 4-1.1.2.1.1.1 5-1.1.2.1.1.2 6-1.1.2 8-1 9-1.2.r 10-1.2.1.1.1 12-1.2.1.1.1 13-1.2 (d / depend-01~e.8 :ARG0 (e / express-03~e.0 :ARG2~e.1 (p / protein :name (n / name :op1 "serpinE2"~e.2)) :ARG3~e.3 (c / cell~e.6 :source (d2 / disease :name (n3 / name :op1 "colorectal"~e.4 :op2 "cancer"~e.5)))) :ARG1~e.9 (a / activity-06~e.13 :ARG0 (p2 / pathway :name (n2 / name :op1 "MEK/ERK"~e.10,12)))) # ::id a_pmid_2094_2929.75 ::amr-annotator SDL-AMR-09 ::preferred # ::tok To assess the contribution of serpinE2 in human colorectal cancer , serpinE2 expression was first examined in various CRC cell lines including Caco @-@ 2 @/@ 15 as well as others exhibiting mutation in KRAS @ ( HCT @-@ 116 , DLD @-@ 1 , LoVo , SW480 , T84 ) or BRAF @ ( Colo @-@ 205 , HT @-@ 29 ) [ @ 36 @ ] . # ::alignments 1-1.5 3-1.5.1 4-1.5.1.1.r 5-1.5.1.1 6-1.5.1.2.r 7-1.5.1.2.2 8-1.5.1.2.1.1 9-1.5.1.2.1.2 11-1.1.1.1.1 12-1.1 14-1.2 15-1 16-1.3.r 17-1.3.1 18-1.3.3.1.1 19-1.3 20-1.3 20-1.3.2.1.2 20-1.3.2.1.3 21-1.3.2 22-1.3.2.1.1.1.1 24-1.3.2.1.1.1.1 26-1.3.2.1.1.1.1 27-1.3.2.1 27-1.3.2.1.2.3.1 27-1.3.2.1.3.3.1 28-1.3.2.1.2.3.1 29-1.3.2.1.2.1.r 29-1.3.2.1.2.3.1 30-1.3.2.1.2.1 31-1.3.2.1.2.2 31-1.3.2.1.3.1 32-1.3.2.1.2.2.1 32-1.3.2.1.3.1.1 35-1.3.2.1.2.2.1.1.1.1 38-1.3.2.1.2.3.1.1.1.1 40-1.3.2.1.2.3.1.1.1.1 42-1.3.2.1.2.3.1.2.1.1 44-1.3.2.1.2.3.1.2.1.1 46-1.3.2.1.2.3.1.3.1.1 48-1.3.2.1.2.3.1.4.1.1 50-1.3.2.1.2.3.1.5.1.1 54-1.3.2.1.3.1.1.1.1.1 57-1.3.2.1.3.3.1.1.1.1 59-1.3.2.1.3.3.1.1.1.1 61-1.3.2.1.3.3.1.2.1.1 63-1.3.2.1.3.3.1.2.1.1 67-1.4.1.1.1 (e / examine-01~e.15 :ARG1 (e2 / express-03~e.12 :ARG2 (p / protein :name (n / name :op1 "serpinE2"~e.11))) :time (f / first~e.14) :location~e.16 (c / cell-line~e.19,20 :mod (v / various~e.17) :ARG2-of (i / include-01~e.21 :ARG1 (a / and~e.27 :op1 (c2 / cell-line :name (n3 / name :op1 "Caco-2/15"~e.22,24,26)) :op2 (c3 / cell-line~e.20 :mod~e.29 (o / other~e.30) :ARG0-of (e3 / exhibit-01~e.31 :ARG1 (m / mutate-01~e.32 :ARG1 (g / gene :name (n4 / name :op1 "KRAS"~e.35)))) :ARG1-of (m3 / mean-01 :ARG2 (a2 / and~e.27,28,29 :op1 (c5 / cell-line :name (n6 / name :op1 "HCT-116"~e.38,40)) :op2 (c6 / cell-line :name (n7 / name :op1 "DLD-1"~e.42,44)) :op3 (c7 / cell-line :name (n8 / name :op1 "LoVo"~e.46)) :op4 (c8 / cell-line :name (n9 / name :op1 "SW480"~e.48)) :op5 (c9 / cell-line :name (n10 / name :op1 "T84"~e.50))))) :op3 (c4 / cell-line~e.20 :ARG0-of (e4 / exhibit-01~e.31 :ARG1 (m2 / mutate-01~e.32 :ARG1 (g2 / gene :name (n5 / name :op1 "BRAF"~e.54)))) :mod o :ARG1-of (m4 / mean-01 :ARG2 (a3 / and~e.27 :op1 (c10 / cell-line :name (n11 / name :op1 "Colo-205"~e.57,59)) :op2 (c11 / cell-line :name (n12 / name :op1 "HT-29"~e.61,63))))))) :mod (d3 / disease :name (n2 / name :op1 "CRC"~e.18))) :ARG1-of (d / describe-01 :ARG0 (p2 / publication :ARG1-of (c12 / cite-01 :ARG2 36~e.67))) :purpose (a4 / assess-01~e.1 :ARG1 (c13 / contribute-01~e.3 :ARG0~e.4 p~e.5 :ARG2~e.6 (d2 / disease :name (n13 / name :op1 "colorectal"~e.8 :op2 "cancer"~e.9) :mod (h / human~e.7))))) # ::id a_pmid_2094_2929.76 ::amr-annotator SDL-AMR-09 ::preferred # ::tok As shown in Figure 3A , serpinE2 @ mRNA levels were barely detectable in the Caco @-@ 2 @/@ 15 cell line while being markedly expressed in all other CRC cell lines tested . # ::alignments 1-1.3 2-1.3.1.r 3-1.3.1 5-1.3.1.1 9-1.1.1.1.1.2.1.1.1 11-1.1.1.1.1.1.1 12-1.1.1.1 14-1.1.1.2 15-1.1.1 16-1.1.1.3.r 18-1.1.1.3.1.1 20-1.1.1.3.1.1 22-1.1.1.3.1.1 23-1.2.2 24-1.2.2 25-1 27-1.2.3 27-1.2.3.r 28-1.2 30-1.2.2.1 31-1.2.2.2 32-1.2.2.4.1.1 33-1.2.2 34-1.2.2 35-1.2.2.3 (c / contrast-01~e.25 :ARG1 (p / possible-01 :ARG1 (d / detect-01~e.15 :ARG1 (l / level~e.12 :quant-of (n5 / nucleic-acid :name (n / name :op1 "mRNA"~e.11) :ARG0-of (e / encode-01 :ARG1 (g / gene :name (n2 / name :op1 "serpinE2"~e.9))))) :degree (b / bare~e.14) :location~e.16 (c2 / cell-line :name (n3 / name :op1 "Caco-2/15"~e.18,20,22)))) :ARG2 (e2 / express-03~e.28 :ARG1 n5 :ARG3 (c3 / cell-line~e.23,24,33,34 :mod (a / all~e.30) :mod (o / other~e.31) :ARG1-of (t / test-01~e.35) :mod (d2 / disease :name (n4 / name :op1 "CRC"~e.32))) :manner~e.27 (m / marked~e.27)) :ARG1-of (s / show-01~e.1 :medium~e.2 (f / figure~e.3 :mod "3A"~e.5))) # ::id a_pmid_2094_2929.77 ::amr-annotator SDL-AMR-09 ::preferred # ::tok Two human CRC cell lines , namely HCT116 and LoVo , which have an activating mutation in the KRAS @ gene resulting in elevated MEK @/@ ERK activities [ @ 37 @ ] , were thereby chosen to further analyze the regulation and role of serpinE2 expression in human colorectal cancer cells . # ::alignments 0-1.1.1.1 1-1.1.1.2 2-1.1.1.5.1.1 3-1.1.1 4-1.1.1 6-1.1.1.4 7-1.1.1.4.1.1.1.1 8-1.1.1.4.1 9-1.1.1.4.1.2.1.1 14-1.1.1.3.3 15-1.1.1.3 19-1.1.1.3.1.1.1 21-1.1.1.3.1 22-1.1.1.3.2 23-1.1.1.3.2.1.r 24-1.1.1.3.2.1.2 25-1.1.1.3.2.1.1.1.1 27-1.1.1.3.2.1.1.1.1 28-1.1.1.3.2.1 31-1.1.1.3.4.1.1.1 37-1.1 39-1.1.2.2 40-1.1.2 42-1.1.2.1.1 43-1.1.2.1 44-1.1.2.1.2 45-1.1.2.1.1.1.r 46-1.1.2.1.1.1.1.1.1 47-1.1.2.1.1.1 49-1.1.2.1.1.1.2.1.2 50-1.1.2.1.1.1.2.1.1.1 51-1.1.2.1.1.1.2.1.1.2 52-1.1.1 52-1.1.2.1.1.1.2 (c9 / cause-01 :ARG1 (c / choose-01~e.37 :ARG1 (c2 / cell-line~e.3,4,52 :quant 2~e.0 :mod (h / human~e.1) :location-of (m2 / mutate-01~e.15 :ARG1 (g / gene~e.21 :name (n4 / name :op1 "KRAS"~e.19)) :ARG1-of (r / result-01~e.22 :ARG2~e.23 (a3 / act-02~e.28 :ARG0 (p3 / pathway :name (n8 / name :op1 "MEK/ERK"~e.25,27)) :ARG1-of (e / elevate-01~e.24))) :ARG0-of (a2 / activate-01~e.14) :ARG1-of (d / describe-01 :ARG0 (p / publication :ARG1-of (c5 / cite-01 :ARG2 37~e.31)))) :ARG1-of (m / mean-01~e.6 :ARG2 (a / and~e.8 :op1 (c3 / cell-line :name (n2 / name :op1 "HCT116"~e.7)) :op2 (c4 / cell-line :name (n3 / name :op1 "LoVo"~e.9)))) :mod (d3 / disease :name (n / name :op1 "CRC"~e.2))) :purpose (a4 / analyze-01~e.40 :ARG1 (a5 / and~e.43 :op1 (r2 / regulate-01~e.42 :ARG1~e.45 (e4 / express-03~e.47 :ARG2 (p2 / protein :name (n7 / name :op1 "serpinE2"~e.46)) :ARG3 (c6 / cell~e.52 :source (d2 / disease :name (n5 / name :op1 "colorectal"~e.50 :op2 "cancer"~e.51) :mod (h2 / human~e.49))))) :op2 (r3 / role~e.44 :poss e4)) :degree (f / further~e.39)))) # ::id a_pmid_2094_2929.78 ::amr-annotator SDL-AMR-09 ::preferred # ::tok In addition , the impact of U0126 treatment was also investigated to evaluate the contribution of endogenous MEK @/@ ERK activities in serpinE2 expression in human cell models . # ::alignments 0-1 1-1 4-1.1.1 5-1.1.1.1.r 6-1.1.1.1.1.1.1 7-1.1.1.1 9-1.1.3 10-1.1 12-1.1.2 14-1.1.2.1 15-1.1.2.1.1.r 16-1.1.2.1.1.1.2 17-1.1.2.1.1.1.1.1 19-1.1.2.1.1.1.1.1 20-1.1.2.1.1 21-1.1.2.1.2.r 22-1.1.2.1.2.1.1.1 23-1.1.2.1.2 24-1.1.2.1.2.2.r 25-1.1.2.1.2.2.1.1 26-1.1.2.1.2.2.1 27-1.1.2.1.2.2 (a / and~e.0,1 :op2 (i2 / investigate-01~e.10 :ARG1 (i / impact-01~e.4 :ARG0~e.5 (t / treat-04~e.7 :ARG2 (s / small-molecule :name (n / name :op1 "U0126"~e.6)))) :ARG2 (e / evaluate-01~e.12 :ARG1 (c / contribute-01~e.14 :ARG0~e.15 (a3 / act-02~e.20 :ARG0 (p / pathway :name (n2 / name :op1 "MEK/ERK"~e.17,19) :mod (e2 / endogenous~e.16))) :ARG2~e.21 (e3 / express-03~e.23 :ARG2 (p2 / protein :name (n4 / name :op1 "serpinE2"~e.22)) :ARG3~e.24 (m / model~e.27 :mod (c2 / cell~e.26 :mod (h / human~e.25)))))) :mod (a2 / also~e.9))) # ::id a_pmid_2094_2929.79 ::amr-annotator SDL-AMR-09 ::preferred # ::tok Forty @-@ eight @-@ hour treatment of HCT116 and LoVo cell lines with U0126 efficiently blocked endogenous MEK activity as confirmed by the marked inhibition of ERK1 @/@ 2 phosphorylation ( data not shown ) [ @ 14 @ ] . # ::alignments 4-1.1.3.2 5-1.1 6-1.1.1.r 7-1.1.1.1.1.1 8-1.1.1 9-1.1.1.2.1.1 10-1.1.1.1 10-1.1.1.2 11-1.1.1.1 12-1.1.2.r 13-1.1.2.1.1 14-1.3 15-1 16-1.2.1.2 17-1.2.1.1.1 18-1.2 19-1.4.r 20-1.4 21-1.4.1.r 23-1.4.1.2 24-1.4.1 25-1.4.1.1.r 26-1.4.1.1.1.1.1.1 27-1.4.1.1.1 29-1.4.1.1 31-1.5.1.1 32-1.5.1.1.1.1 32-1.5.1.1.1.1.r 33-1.5.1.1.1 37-1.5.1.2.1.1 (b2 / block-01~e.15 :ARG0 (t2 / treat-04~e.5 :ARG1~e.6 (a / and~e.8 :op1 (c / cell-line~e.10,11 :name (n3 / name :op1 "HCT116"~e.7)) :op2 (c2 / cell-line~e.10 :name (n4 / name :op1 "LoVo"~e.9))) :ARG2~e.12 (s / small-molecule :name (n / name :op1 "U0126"~e.13)) :duration (t / temporal-quantity :quant 48 :unit (h / hour~e.4))) :ARG1 (a2 / activity-06~e.18 :ARG0 (e / enzyme :name (n2 / name :op1 "MEK"~e.17) :mod (e3 / endogenous~e.16))) :ARG2-of (e2 / efficient-01~e.14) :ARG1-of~e.19 (c3 / confirm-01~e.20 :ARG0~e.21 (i / inhibit-01~e.24 :ARG1~e.25 (p / phosphorylate-01~e.29 :ARG1 (s2 / slash~e.27 :op1 (e4 / enzyme :name (n5 / name :op1 "ERK1"~e.26)) :op2 (e5 / enzyme :name (n6 / name :op1 "ERK2")))) :mod (m / marked~e.23))) :ARG1-of (d / describe-01 :ARG0 (a3 / and :op1 (d2 / data~e.31 :ARG1-of (s3 / show-01~e.33 :polarity~e.32 -~e.32)) :op2 (p2 / publication :ARG1-of (c4 / cite-01 :ARG2 14~e.37))))) # ::id a_pmid_2094_2929.80 ::amr-annotator SDL-AMR-09 ::preferred # ::tok As shown in Figure 3B , treatment of these CRC cell lines with U0126 markedly and significantly reduced serpinE2 @ mRNA levels , indicating that expression of serpinE2 @ is likely dependent of ERK activity in these cell lines . # ::alignments 1-1.5 2-1.5.1.r 3-1.5.1 5-1.5.1.1 8-1.1 9-1.1.1.r 10-1.1.1.1 11-1.1.1.2.1.1 12-1.1.1 13-1.1.1 14-1.1.2.r 15-1.1.2.1.1 16-1.3.1 17-1.3 18-1.3.2 19-1 21-1.2.1.2.1.1.1 23-1.2.1.1.1 24-1.2 26-1.4 27-1.4.1.r 28-1.4.1.1.1 31-1.4.1.1.1.1 34-1.4.1 35-1.4.1.1 36-1.4.1.1.2.r 37-1.4.1.1.2.1.1.1 38-1.4.1.1.2 40-1.1.1.1 41-1.1.1 42-1.1.1 (r / reduce-01~e.19 :ARG0 (t / treat-04~e.8 :ARG1~e.9 (c / cell-line~e.12,13,41,42 :mod (t2 / this~e.10,40) :mod (d2 / disease :name (n3 / name :op1 "CRC"~e.11))) :ARG2~e.14 (s / small-molecule :name (n / name :op1 "U0126"~e.15))) :ARG1 (l / level~e.24 :quant-of (n6 / nucleic-acid :name (n4 / name :op1 "mRNA"~e.23) :ARG0-of (e2 / encode-01 :ARG1 (g / gene :name (n5 / name :op1 "serpinE2"~e.21))))) :ARG2 (a / and~e.17 :op1 (m / marked~e.16) :op2 (s2 / significant-02~e.18)) :ARG0-of (i / indicate-01~e.26 :ARG1~e.27 (l2 / likely-01~e.34 :ARG1 (d / depend-01~e.35 :ARG0 (e3 / express-03~e.28 :ARG1 g~e.31 :ARG3 c) :ARG1~e.36 (a2 / activity-06~e.38 :ARG0 (p / protein-family :name (n2 / name :op1 "ERK"~e.37)))))) :ARG1-of (s3 / show-01~e.1 :medium~e.2 (f / figure~e.3 :mod "3B"~e.5))) # ::id a_pmid_2094_2929.81 ::amr-annotator SDL-AMR-09 ::preferred # ::tok Down @-@ regulation of serpinE2 expression in human colorectal cancer cells inhibits soft agarose colony formation , migration and tumor growth in nude mice # ::alignments 0-1.1 1-1.1 2-1.1 3-1.1.1.r 4-1.1.1.1.1.1 5-1.1.1 6-1.1.1.2.r 7-1.1.1.2.1.2 8-1.1.1.2.1.1.1 9-1.1.1.2.1.1.2 10-1.1.1.2 11-1 12-1.2.1.2.1 13-1.2.1.2 14-1.2.1.1 15-1.2.1 17-1.2.2 18-1.2 19-1.2.3.1 20-1.2.3 21-1.3.r 22-1.3.1 23-1.3 (i / inhibit-01~e.11 :ARG0 (d / downregulate-01~e.0,1,2 :ARG1~e.3 (e / express-03~e.5 :ARG2 (p / protein :name (n / name :op1 "serpinE2"~e.4)) :ARG3~e.6 (c / cell~e.10 :source (d2 / disease :name (n3 / name :op1 "colorectal"~e.8 :op2 "cancer"~e.9) :mod (h / human~e.7))))) :ARG1 (a / and~e.18 :op1 (f / form-01~e.15 :ARG1 (c4 / colony~e.14) :location (a2 / agarose~e.13 :ARG1-of (s / soft-02~e.12))) :op2 (m / migrate-01~e.17) :op3 (g / grow-01~e.20 :ARG1 (t / tumor~e.19))) :location~e.21 (m2 / mouse~e.23 :mod (n2 / nude~e.22))) # ::id a_pmid_2094_2929.82 ::amr-annotator SDL-AMR-09 ::preferred # ::tok We next investigated the effect of serpinE2 knockdown on anchorage independent growth and cell migration after downregulation of serpinE2 @ gene expression by RNA interference in HCT116 and LoVo cells . # ::alignments 0-1.1 1-1.3 1-1.4 2-1 4-1.2 5-1.2.1.r 6-1.2.1.1.1.1 7-1.2.1 8-1.2.2.r 9-1.2.2.1.1.2 10-1.2.2.1.1 10-1.2.2.1.1.1 10-1.2.2.1.1.1.r 11-1.2.2.1 12-1.2.2 13-1.2.2.2.1 14-1.2.2.2 15-1.4 16-1.4.1 19-1.4.1.1.1.1.1 21-1.4.1.1.1 22-1.4.1.1 23-1.4.1.2.r 24-1.4.1.2.1.1.1 25-1.4.1.2 26-1.4.1.1.2.r 27-1.4.1.1.2.1.1.1 28-1.4.1.1.2 29-1.4.1.1.2.2.1.1 30-1.4.1.1.2.1 (i / investigate-01~e.2 :ARG0 (w / we~e.0) :ARG1 (a / affect-01~e.4 :ARG0~e.5 (k / knock-down-02~e.7 :ARG1 (p / protein :name (n2 / name :op1 "serpinE2"~e.6))) :ARG1~e.8 (a2 / and~e.12 :op1 (g / grow-01~e.11 :ARG0-of (d / depend-01~e.10 :polarity~e.10 -~e.10 :ARG1 (a3 / anchorage~e.9))) :op2 (m / migrate-01~e.14 :ARG0 (c / cell~e.13)))) :time (n / next~e.1) :time (a4 / after~e.1,15 :op1 (d2 / downregulate-01~e.16 :ARG1 (e / express-03~e.22 :ARG1 (g2 / gene~e.21 :name (n3 / name :op1 "serpinE2"~e.19)) :ARG3~e.26 (a5 / and~e.28 :op1 (c2 / cell-line~e.30 :name (n4 / name :op1 "HCT116"~e.27)) :op2 (c3 / cell-line :name (n5 / name :op1 "LoVo"~e.29)))) :ARG2~e.23 (i2 / interfere-01~e.25 :ARG0 (n6 / nucleic-acid :name (n7 / name :op1 "RNA"~e.24)))))) # ::id a_pmid_2094_2929.83 ::amr-annotator SDL-AMR-09 ::preferred # ::tok As shown in Figure 4A , serpinE2 @ mRNA were significantly reduced by respectively 37 % and 88 % in LoVo cells expressing shSerpinE2(#15 ) or shSerpinE2(#16 ) and by 77 % and 92 % in HCT116 expressing shSerpinE2(#15 ) or shSerpinE2(#16 ) ; conversely , expression of the control shRNA ( shTGFP ) had no effect on endogenous serpinE2 @ expression ( data not shown ) . # ::alignments 1-1.1.5 2-1.1.5.1.r 3-1.1.5.1 5-1.1.5.1.1 9-1.1.1.1.2.1.1.1 11-1.1.1.1.1.1 13-1.1.1.3 14-1.1.1 14-1.1.2 14-1.1.3 14-1.1.4 17-1.1.1.2.1 18-1.1.1.2 19-1.1 20-1.1.2.2.1 21-1.1.2.2 23-1.1.1.4.1.1 23-1.1.2.3.1.1 24-1.1.1.4 24-1.1.2.3 24-1.1.3.3 24-1.1.4.4 25-1.1.1.4.2 25-1.1.2.3.2 31-1.1 32-1.1.3.2.r 33-1.1.3.2.1 34-1.1.3.2 35-1.1 36-1.1.4.2.1 37-1.1.4.2 39-1.1.3.3.1.1 39-1.1.4.4.1.1 40-1.1.1.4.2 49-1.2.1.2 52-1.2.1.2.1.2 53-1.2.1.2.1.1.1 55-1.2.1.2.1.3.1.1.1 58-1.2.1.1 58-1.2.1.1.r 59-1.2.1 61-1.2.1.3.1.2 63-1.2.1.3.1.1.1 65-1.2.1.2 65-1.2.1.3 67-1.2.2.1 68-1.2.2.1.1.1 68-1.2.2.1.1.1.r 69-1.2.2.1.1 (c5 / contrast-01 :ARG1 (a5 / and~e.19,31,35 :op1 (r / reduce-01~e.14 :ARG1 (n12 / nucleic-acid :name (n / name :op1 "mRNA"~e.11) :ARG0-of (e / encode-01 :ARG1 (g / gene :name (n2 / name :op1 "serpinE2"~e.9)))) :ARG2 (p / percentage-entity~e.18 :value 37~e.17) :ARG1-of (s / significant-02~e.13) :location (c / cell-line~e.24 :name (n3 / name :op1 "LoVo"~e.23) :ARG3-of (e2 / express-03~e.25,40 :ARG2 (p5 / protein :name (n5 / name :op1 "shSerpinE2") :ARG1-of (l / label-01 :ARG2 15))))) :op2 (r8 / reduce-01~e.14 :ARG1 n12 :ARG2 (p2 / percentage-entity~e.21 :value 88~e.20) :location (c6 / cell-line~e.24 :name (n10 / name :op1 "LoVo"~e.23) :ARG3-of (e6 / express-03~e.25 :ARG2 (p6 / protein :name (n4 / name :op1 "shSerpinE2") :ARG1-of (l2 / label-01 :ARG2 16)))) :ARG1-of s) :op3 (r9 / reduce-01~e.14 :ARG1 n12 :ARG2~e.32 (p3 / percentage-entity~e.34 :value 77~e.33) :location (c2 / cell-line~e.24 :name (n6 / name :op1 "HCT116"~e.39) :ARG3-of e2) :ARG1-of s) :op4 (r10 / reduce-01~e.14 :ARG1 n12 :ARG2 (p4 / percentage-entity~e.37 :value 92~e.36) :ARG1-of s :location (c7 / cell-line~e.24 :name (n11 / name :op1 "HCT116"~e.39) :ARG3-of e6)) :ARG1-of (s3 / show-01~e.1 :medium~e.2 (f / figure~e.3 :mod "4A"~e.5))) :ARG2 (c3 / contrast-01 :ARG1 (a4 / affect-01~e.59 :polarity~e.58 -~e.58 :ARG0 (e3 / express-03~e.49,65 :ARG2 (n15 / nucleic-acid :name (n7 / name :op1 "shRNA"~e.53) :mod (c4 / control~e.52) :ARG1-of (d / describe-01 :ARG2 (p7 / protein :name (n8 / name :op1 "shTGFP"~e.55))))) :ARG1 (e4 / express-03~e.65 :ARG1 (g2 / gene :name (n9 / name :op1 "serpinE2"~e.63) :mod (e5 / endogenous~e.61)))) :ARG1-of (d2 / describe-01 :ARG0 (d3 / data~e.67 :ARG1-of (s2 / show-01~e.69 :polarity~e.68 -~e.68))))) # ::id a_pmid_2094_2929.84 ::amr-annotator SDL-AMR-09 ::preferred # ::tok Again , the proliferation rate of these cell populations was assessed when cultured on plastic . # ::alignments 0-1.3 3-1.1.1 4-1.1 5-1.1.1.1.r 6-1.1.1.1.1 7-1.1.1.1.2 8-1.1.1.1 10-1 11-1.2.r 12-1.2 13-1.2.2.r 14-1.2.2 (a / assess-01~e.10 :ARG1 (r / rate~e.4 :degree-of (p / proliferate-01~e.3 :ARG0~e.5 (p2 / population~e.8 :mod (t / this~e.6) :mod (c / cell~e.7)))) :time~e.11 (c2 / culture-01~e.12 :ARG1 p2 :location~e.13 (p3 / plastic~e.14)) :mod (a2 / again~e.0)) # ::id a_pmid_2094_2929.85 ::amr-annotator SDL-AMR-09 ::preferred # ::tok No difference was observed in the proliferation rate of subconfluent cells when serpinE2 expression was downregulated ( Figure 4B ) . # ::alignments 0-1.1.1 0-1.1.1.r 1-1.1 1-1.1.1 1-1.1.1.r 3-1 4-1.1.2.r 6-1.1.2.1 7-1.1.2 8-1.1.2.1.1.r 9-1.1.2.1.1.1 10-1.1.2.1.1 11-1.2.r 12-1.2.1.1.1.1 13-1.2.1 15-1.2 17-1.3.1 19-1.3.1.1 (o / observe-01~e.3 :ARG1 (d / differ-02~e.1 :polarity~e.0,1 -~e.0,1 :ARG1~e.4 (r / rate~e.7 :degree-of (p / proliferate-01~e.6 :ARG0~e.8 (c / cell~e.10 :mod (s / subconfluent~e.9))))) :time~e.11 (d2 / downregulate-01~e.15 :ARG1 (e / express-03~e.13 :ARG2 (p2 / protein :name (n / name :op1 "serpinE2"~e.12)))) :ARG1-of (d3 / describe-01 :ARG0 (f / figure~e.17 :mod "4B"~e.19))) # ::id a_pmid_2094_2929.86 ::amr-annotator SDL-AMR-09 ::preferred # ::tok We then verified whether the reduction in serpinE2 @ expression alters the ability of colon cancer cells to form colonies in soft agarose . # ::alignments 0-1.1 1-1.3 2-1 3-1.2.1 3-1.2.1.r 5-1.2.2 8-1.2.2.1.1.1.1 10-1.2.2.1 11-1.2 13-1.2.3 14-1.2.3.1.r 15-1.2.3.1.1.1.1 16-1.2.3.1.1.1.2 17-1.2.3.1 19-1.2.3.2 20-1.2.3.2.2 21-1.2.3.2.3.r 22-1.2.3.2.3.1 23-1.2.3.2.3 (v / verify-01~e.2 :ARG0 (w / we~e.0) :ARG1 (a / alter-01~e.11 :mode~e.3 interrogative~e.3 :ARG0 (r / reduce-01~e.5 :ARG1 (e / express-03~e.10 :ARG1 (g / gene :name (n / name :op1 "serpinE2"~e.8)))) :ARG1 (c / capable-01~e.13 :ARG1~e.14 (c2 / cell~e.17 :source (d / disease :name (n2 / name :op1 "colon"~e.15 :op2 "cancer"~e.16))) :ARG2 (f / form-01~e.19 :ARG0 c2 :ARG1 (c5 / colony~e.20) :location~e.21 (a2 / agarose~e.23 :ARG1-of (s / soft-02~e.22))))) :time (t / then~e.1)) # ::id a_pmid_2094_2929.87 ::amr-annotator SDL-AMR-09 ::preferred # ::tok As shown in Figure 4C , expression of both shRNA against SerpinE2 (#15 and #16 ) decreased the ability of HCT116 and LoVo cells to form colonies in soft agarose . # ::alignments 1-1.3 2-1.3.1.r 3-1.3.1 5-1.3.1.1 8-1.1 9-1.1.1.r 10-1.1.1.2 11-1.1.1.1.1 12-1.1.1 12-1.1.1.3 12-1.1.1.3.r 13-1.1.1.3.1.1.1 13-1.1.1.3.1.2.1.1.1.1 13-1.1.1.3.1.2.1.2.1.1 15-1.1.1.3.1.2.1 18-1 20-1.2 21-1.2.1.r 22-1.2.1.1.1.1 23-1.2.1 24-1.2.1.2.1.1 25-1.2.1.1 25-1.2.1.2 27-1.2.2 28-1.2.2.2 29-1.2.2.3.r 30-1.2.2.3.1 31-1.2.2.3 (d / decrease-01~e.18 :ARG0 (e / express-03~e.8 :ARG2~e.9 (n7 / nucleic-acid~e.12 :name (n / name :op1 "shRNA"~e.11) :mod (b / both~e.10) :ARG0-of~e.12 (c / counter-01~e.12 :ARG1 (p / protein :name (n2 / name :op1 "SerpinE2"~e.13) :ARG1-of (m / mean-01 :ARG2 (a / and~e.15 :op1 (p2 / protein :name (n3 / name :op1 "SerpinE2"~e.13) :ARG1-of (l / label-01 :ARG2 15)) :op2 (p3 / protein :name (n4 / name :op1 "SerpinE2"~e.13) :ARG1-of (l2 / label-01 :ARG2 16)))))))) :ARG1 (c2 / capable-01~e.20 :ARG1~e.21 (a2 / and~e.23 :op1 (c3 / cell-line~e.25 :name (n5 / name :op1 "HCT116"~e.22)) :op2 (c4 / cell-line~e.25 :name (n6 / name :op1 "LoVo"~e.24))) :ARG2 (f / form-01~e.27 :ARG0 a2 :ARG1 (c5 / colony~e.28) :location~e.29 (a3 / agarose~e.31 :ARG1-of (s / soft-02~e.30)))) :ARG1-of (s2 / show-01~e.1 :ARG0~e.2 (f2 / figure~e.3 :mod "4C"~e.5))) # ::id a_pmid_2094_2929.88 ::amr-annotator SDL-AMR-09 ::preferred # ::tok Of note , shSerpinE2(#15 ) which was less efficient than the shRNA (#16 ) to reduce serpinE2 @ gene expression ( Figure 4A ) was also less efficient to reduce colony formation . # ::alignments 1-1.1.4 7-1.1.2.2 8-1.1 8-1.1.2 8-1.1.2.r 9-1.1.2.4.r 11-1.1.2.4.1.1 15-1.1.2.1 17-1.1.2.1.2.1.1.1 19-1.1.2.1.2.1 20-1.1.2.1.2 22-1.1.2.3.1 24-1.1.2.3.1.1 28-1.3 29-1.4 30-1 30-1.1 30-1.1.2 30-1.1.2.r 30-1.1.r 32-1.2 33-1.2.2.1 34-1.2.2 (e / efficient-01~e.30 :ARG1~e.30 (p / protein~e.8,30 :name (n / name :op1 "shSerpinE2") :ARG1-of~e.8,30 (e2 / efficient-01~e.8,30 :ARG2 (r4 / reduce-01~e.15 :ARG0 p :ARG1 (e3 / express-03~e.20 :ARG1 (g / gene~e.19 :name (n3 / name :op1 "serpinE2"~e.17)))) :degree (l / less~e.7) :ARG1-of (d / describe-01 :ARG0 (f2 / figure~e.22 :mod "4A"~e.24)) :compared-to~e.9 (n6 / nucleic-acid :name (n2 / name :op1 "shRNA"~e.11) :ARG1-of (l3 / label-01 :ARG2 16))) :ARG1-of (l2 / label-01 :ARG2 15) :ARG1-of (n4 / note-02~e.1)) :ARG2 (r / reduce-01~e.32 :ARG0 p :ARG1 (f / form-01~e.34 :ARG1 (c / colony~e.33))) :mod (a / also~e.28) :degree l~e.29) # ::id a_pmid_2094_2929.89 ::amr-annotator SDL-AMR-09 ::preferred # ::tok This indicates that serpinE2 controls anchorage @-@ independent growth of human colon carcinoma cells . # ::alignments 0-1.1 1-1 2-1.2.r 3-1.2.1.1.1 4-1.2 5-1.2.2.2.2 7-1.2.2.2 7-1.2.2.2.1 7-1.2.2.2.1.r 8-1.2.2 9-1.2.2.1.r 10-1.2.2.1.1.1 11-1.2.2.1.1.2 12-1.2.2.1.1 13-1.2.2.1 (i / indicate-01~e.1 :ARG0 (t / this~e.0) :ARG1~e.2 (c / control-01~e.4 :ARG0 (p / protein :name (n / name :op1 "serpinE2"~e.3)) :ARG1 (g / grow-01~e.8 :ARG1~e.9 (c2 / cell~e.13 :source (c3 / carcinoma~e.12 :mod (h / human~e.10) :mod (c4 / colon~e.11))) :ARG0-of (d / depend-01~e.7 :polarity~e.7 -~e.7 :ARG1 (a / anchorage~e.5))))) # ::id a_pmid_2094_2929.90 ::amr-annotator SDL-AMR-09 ::preferred # ::tok Additionally , as observed in caMEK @-@ expressing IECs , the size of foci formed at post @-@ confluency was significantly decreased in serpinE2 @-@ depleted LoVo cells ( Figure 4D ) . # ::alignments 0-1 2-1.1.1.1.1.1.r 3-1.1.4 5-1.1.4.1.2.1.1.1 7-1.1.4.1.2 8-1.1.4.1.1.1 11-1.1.1 13-1.1.1.1 14-1.1.1.1.1 16-1.1.1.1.1.1 18-1.1.1.1.1.1.1 20-1.1.2 21-1.1 23-1.1.3.2.1.1.1 25-1.1.3.2 26-1.1.3.1.1 27-1.1.3 27-1.1.4.1 29-1.2.1 31-1.2.1.1 (a / and~e.0 :op2 (d / decrease-01~e.21 :ARG1 (s2 / size~e.11 :poss (f / focus~e.13 :ARG1-of (f2 / form-01~e.14 :time~e.2 (a2 / after~e.16 :op1 (c3 / confluency~e.18))))) :ARG2 (s / significant-02~e.20) :location (c / cell-line~e.27 :name (n / name :op1 "LoVo"~e.26) :ARG1-of (d2 / deplete-01~e.25 :ARG2 (p2 / protein :name (n2 / name :op1 "serpinE2"~e.23)))) :ARG1-of (o / observe-01~e.3 :location (c2 / cell~e.27 :name (n3 / name :op1 "IEC"~e.8) :ARG3-of (e / express-03~e.7 :ARG2 (e2 / enzyme :name (n4 / name :op1 "caMEK"~e.5)))))) :ARG1-of (d3 / describe-01 :ARG0 (f3 / figure~e.29 :mod "4D"~e.31))) # ::id a_pmid_2094_2929.91 ::amr-annotator SDL-AMR-09 ::preferred # ::tok The tumorigenicity of colorectal cell lines was next assessed after subcutaneous ( s.c.) injection into the flank of nude mice . # ::alignments 1-1.1 1-1.1.1 1-1.1.1.2 1-1.1.1.2.r 1-1.1.1.r 2-1.1.1.1.r 3-1.1.1.1.1 4-1.1.1.1 5-1.1.1.1 7-1.2 7-1.3 8-1 9-1.3 10-1.3.1.2 13-1.3.1 14-1.3.1.1.r 16-1.3.1.1 17-1.3.1.1.1.r 18-1.3.1.1.1.1 19-1.3.1.1.1 (a / assess-01~e.8 :ARG1 (p / possible-01~e.1 :ARG1~e.1 (c3 / create-01~e.1 :ARG0~e.2 (c / cell-line~e.4,5 :source (c2 / colorectal~e.3)) :ARG1~e.1 (t / tumor~e.1))) :time (n / next~e.7) :time (a2 / after~e.7,9 :op1 (i / inject-01~e.13 :ARG2~e.14 (f / flank~e.16 :poss~e.17 (m / mouse~e.19 :mod (n2 / nude~e.18))) :mod (s / subcutaneous~e.10)))) # ::id a_pmid_2094_2929.92 ::amr-annotator SDL-AMR-09 ::preferred # ::tok As shown in Figure 5A and 5B , HCT116 and LoVo cell lines induced palpable tumors with a short latency period of respectively 15 and 10 days after their injection . # ::alignments 0-1.2.2.3.r 1-1.3 2-1.3.1.r 3-1.3.1.1 3-1.3.1.2 5-1.3.1.1.1 7-1.3.1 9-1.3.1.2.1 12-1.1.1.1.1 13-1.1 14-1.1.2.1.1 15-1.1.1 15-1.1.2 16-1.1.1 17-1 18-1.2.1 19-1.2 22-1.2.2.2 23-1.2.2.1 24-1.2.2 26-1.2.2.3.2.3 27-1.2.2.3.2.1.1 28-1.2.2.3.2 29-1.2.2.3.2.2.1 30-1.2.2.3.2.1.2 30-1.2.2.3.2.2.2 31-1.2.2.3 32-1.2.2.3.1.1 32-1.2.2.3.1.1.r 33-1.2.2.3.1 (i / induce-01~e.17 :ARG0 (a / and~e.13 :op1 (c / cell-line~e.15,16 :name (n / name :op1 "HCT116"~e.12)) :op2 (c2 / cell-line~e.15 :name (n2 / name :op1 "LoVo"~e.14))) :ARG2 (t2 / tumor~e.19 :mod (p / palpable~e.18) :mod (p2 / period~e.24 :mod (l / latency~e.23) :ARG1-of (s / short-07~e.22) :time~e.0 (a3 / after~e.31 :op1 (i2 / inject-01~e.33 :ARG1~e.32 a~e.32) :quant (a2 / and~e.28 :op1 (t3 / temporal-quantity :quant 15~e.27 :unit (d2 / day~e.30)) :op2 (t4 / temporal-quantity :quant 10~e.29 :unit (d / day~e.30)) :mod (r / respective~e.26))))) :ARG1-of (s2 / show-01~e.1 :medium~e.2 (a4 / and~e.7 :op1 (f / figure~e.3 :mod "5A"~e.5) :op2 (f2 / figure~e.3 :mod "5B"~e.9)))) # ::id a_pmid_2094_2929.93 ::amr-annotator SDL-AMR-09 ::preferred # ::tok More importantly , downregulation of serpinE2 expression with shSerpinE2(#16 ) in these cell lines severely impaired their capacity to grow as tumors in nude mice . # ::alignments 0-1.4.1 1-1.4 3-1.1 4-1.1.1.r 5-1.1.1.1.1.1 6-1.1.1 10-1.1.1.2.r 11-1.1.1.2.1 12-1.1.1.2 13-1.1.1.2 14-1.3 14-1.3.r 15-1 16-1.2.1 16-1.2.1.r 17-1.2 19-1.2.2 20-1.2.2.2.r 21-1.2.2.2 22-1.2.2.3.r 23-1.2.2.3.1 24-1.2.2.3 (i / impair-01~e.15 :ARG0 (d / downregulate-01~e.3 :ARG1~e.4 (e / express-03~e.6 :ARG2 (p / protein :name (n / name :op1 "serpinE2"~e.5)) :ARG3~e.10 (c / cell-line~e.12,13 :mod (t / this~e.11))) :ARG2 (p2 / protein :name (n2 / name :op1 "shSerpinE2") :ARG1-of (l / label-01 :ARG2 16))) :ARG1 (c2 / capable-01~e.17 :ARG1~e.16 c~e.16 :ARG2 (g / grow-02~e.19 :ARG1 c :ARG2~e.20 (t2 / tumor~e.21) :location~e.22 (m / mouse~e.24 :mod (n3 / nude~e.23)))) :manner~e.14 (s / severe~e.14) :mod (i2 / important~e.1 :degree (m2 / more~e.0))) # ::id a_pmid_2094_2929.94 ::amr-annotator SDL-AMR-09 ::preferred # ::tok Finally , in vitro @ transwell migration assays were performed to verify the importance of serpinE2 in colon carcinoma cell migration . # ::alignments 0-1.1 0-1.1.r 3-1.2.2 4-1.2.2 6-1.2.1.1 7-1.2.1 8-1.2 10-1 12-1.3 14-1.3.1 15-1.3.1.1.r 16-1.3.1.1.1.1 17-1.2.2 17-1.3.1.2.r 18-1.3.1.2.1.1.1 19-1.3.1.2.1.1 20-1.3.1.2.1 21-1.3.1.2 (p / perform-02~e.10 :li~e.0 -1~e.0 :ARG1 (a / assay-01~e.8 :ARG1 (m2 / migrate-01~e.7 :mod (t2 / transwell~e.6)) :manner (i / in-vitro~e.3,4,17)) :purpose (v / verify-01~e.12 :ARG1 (i2 / important~e.14 :domain~e.15 (p2 / protein :name (n2 / name :op1 "serpinE2"~e.16)) :purpose~e.17 (m / migrate-01~e.21 :ARG0 (c / cell~e.20 :source (c2 / carcinoma~e.19 :mod (c3 / colon~e.18))))))) # ::id a_pmid_2094_2929.95 ::amr-annotator SDL-AMR-09 ::preferred # ::tok As illustrated in Figure 6A , serpinE2 deficiency significantly reduced HCT116 ( not shown ) and LoVo cell migration to the undersurface of the membrane coated or not with fibronectin or vitronectin ( data not shown ) . # ::alignments 1-1.4 2-1.4.1.r 3-1.4.1 5-1.4.1.1 8-1.1.1.1.1 10-1.3 11-1 12-1.2.1.1.1.1 14-1.2.1.1.2.1 14-1.2.1.1.2.1.r 15-1.2.1.1.2 17-1.2.1 18-1.2.1.2.1.1 19-1.2.1.1 19-1.2.1.2 20-1.2 21-1.2.2.r 23-1.2.2 24-1.2.2.1.r 26-1.2.2.1 27-1.2.2.2 27-1.2.2.2.2.1 28-1.2.2.2.1 28-1.2.2.2.2 29-1.2.2.2.2.1.1 29-1.2.2.2.2.1.1.r 30-1.2.2.2.1.r 31-1.2.2.2.1.1.1.1 32-1.2.2.2.1 33-1.2.2.2.1.2.1.1 35-1.5.1 36-1.5.1.1 37-1.5.1.1 (r / reduce-01~e.11 :ARG0 (l / lack-01 :ARG1 (p / protein :name (n / name :op1 "serpinE2"~e.8))) :ARG1 (m / migrate-01~e.20 :ARG0 (a / and~e.17 :op1 (c2 / cell-line~e.19 :name (n2 / name :op1 "HCT116"~e.12) :ARG1-of (s2 / show-01~e.15 :polarity~e.14 -~e.14)) :op2 (c4 / cell-line~e.19 :name (n3 / name :op1 "LoVo"~e.18))) :ARG2~e.21 (u / undersurface~e.23 :part-of~e.24 (m2 / membrane~e.26) :ARG1-of (c5 / coat-01~e.27 :ARG2~e.30 (o / or~e.28,32 :op1 (p2 / protein :name (n4 / name :op1 "fibronectin"~e.31)) :op2 (p3 / protein :name (n5 / name :op1 "vitronectin"~e.33))) :op1-of (o2 / or~e.28 :op2 (c6 / coat-01~e.27 :polarity~e.29 -~e.29 :ARG1 u :ARG2 o))))) :ARG2 (s / significant-02~e.10) :ARG1-of (i / illustrate-01~e.1 :medium~e.2 (f / figure~e.3 :mod "6A"~e.5)) :ARG1-of (d / describe-01 :ARG0 (d2 / data~e.35 :ARG1-of s2~e.36,37))) # ::id a_pmid_2094_2929.96 ::amr-annotator SDL-AMR-09 ::preferred # ::tok The net effect of serpinE2 knockdown was also determined on invasion by using BD Biocoat Matrigel invasion chambers , in presence of hydroxyurea . # ::alignments 1-1.1.2 2-1.1 3-1.1.1.r 4-1.1.1.1.1.1 5-1.1.1 7-1.2 8-1 9-1.3.r 10-1.3 11-1.4.r 12-1.4 13-1.4.1.1.1 14-1.4.1.1.2 15-1.4.1.1.3 16-1.4.1.1.4 17-1.4.1.1.5 19-1.5.r 20-1.5 21-1.5.1.r 22-1.5.1.1.1 (d / determine-01~e.8 :ARG1 (a / affect-01~e.2 :ARG0~e.3 (k / knock-down-02~e.5 :ARG1 (p / protein :name (n2 / name :op1 "serpinE2"~e.4))) :mod (n / net~e.1)) :mod (a2 / also~e.7) :time~e.9 (i / invade-01~e.10) :manner~e.11 (u / use-01~e.12 :ARG1 (t / thing :name (n3 / name :op1 "BD"~e.13 :op2 "Biocoat"~e.14 :op3 "Matrigel"~e.15 :op4 "invasion"~e.16 :op5 "chamber"~e.17))) :condition~e.19 (p2 / present-02~e.20 :ARG1~e.21 (s / small-molecule :name (n4 / name :op1 "hydroxyurea"~e.22)))) # ::id a_pmid_2094_2929.97 ::amr-annotator SDL-AMR-09 ::preferred # ::tok As shown in Figure 6B , the capacity of LoVo cells to invade Matrigel was also altered by serpinE2 silencing # ::alignments 1-1.4 2-1.4.1.r 3-1.4.1 5-1.4.1.1 9-1.2 10-1.2.1.r 11-1.2.1.1.1 12-1.2.1 14-1.2.2 15-1.2.2.2.1.1 17-1.3 18-1 19-1.1.r 20-1.1.1.1.1 21-1.1 (a / alter-01~e.18 :ARG0~e.19 (s / silence-01~e.21 :ARG1 (p2 / protein :name (n3 / name :op1 "serpinE2"~e.20))) :ARG1 (c / capable-01~e.9 :ARG1~e.10 (c2 / cell-line~e.12 :name (n / name :op1 "LoVo"~e.11)) :ARG2 (i / invade-01~e.14 :ARG0 c2 :ARG1 (p / protein :name (n2 / name :op1 "Matrigel"~e.15)))) :mod (a2 / also~e.17) :ARG1-of (s2 / show-01~e.1 :medium~e.2 (f / figure~e.3 :mod "6B"~e.5))) # ::id a_pmid_2094_2929.98 ::amr-annotator SDL-AMR-09 ::preferred # ::tok To test the hypothesis that this altered migration and invasion capacity could result from a defect in cell adhesion , adhesion strength to the substrate was examined for control and shSerpinE2(#16 ) - expressing LoVo cells . # ::alignments 1-1.2 3-1.2.1 4-1.2.1.1.r 5-1.2.1.1.1.2.3 6-1.2.1.1.1.2.2 7-1.2.1.1.1.2.1.1 8-1.2.1.1.1.2.1 9-1.2.1.1.1.2.1.2 10-1.2.1.1.1.2 11-1.2.1.1 12-1.2.1.1.1 13-1.2.1.1.1.1.r 15-1.2.1.1.1.1 16-1.2.1.1.1.1.1.r 17-1.2.1.1.1.1.1.1 18-1.2.1.1.1.1.1 20-1.1.1 21-1.1 22-1.1.1.2.r 24-1.1.1.2 26-1 27-1.1.1.1.r 28-1.1.1.1.1.2 29-1.1.1.1 33-1.1.1.1.2.2 34-1.1.1.1.1.1.1 34-1.1.1.1.2.1.1 35-1.1.1.1.1 35-1.1.1.1.2 (e / examine-01~e.26 :ARG1 (s / strong-02~e.21 :degree-of (a / adhere-01~e.20 :ARG1~e.27 (a2 / and~e.29 :op1 (c / cell-line~e.35 :name (n / name :op1 "LoVo"~e.34) :mod (c2 / control~e.28)) :op2 (c3 / cell-line~e.35 :name (n2 / name :op1 "LoVo"~e.34) :ARG3-of (e2 / express-03~e.33 :ARG2 (p2 / protein :name (n3 / name :op1 "shSerpinE2") :ARG1-of (l / label-01 :ARG2 16))))) :ARG2~e.22 (s2 / substrate~e.24))) :ARG2 (t2 / test-01~e.1 :ARG1 (h / hypothesize-01~e.3 :ARG1~e.4 (p / possible-01~e.11 :ARG1 (r2 / result-01~e.12 :ARG1~e.13 (d / defect~e.15 :topic~e.16 (a5 / adhere-01~e.18 :ARG1 (c5 / cell~e.17))) :ARG2 (c4 / capable-01~e.10 :ARG2 (a3 / and~e.8 :op1 (m / migrate-01~e.7) :op2 (i / invade-01~e.9)) :ARG1-of (a4 / alter-01~e.6) :mod (t3 / this~e.5))))))) # ::id a_pmid_2094_2929.99 ::amr-annotator SDL-AMR-09 ::preferred # ::tok Using a trypsin @-@ mediated de @-@ adhesion assay , downregulation of serpinE2 significantly delayed LoVo cell detachment after trypsinization ( Figure 6C ) , suggesting that serpinE2 expression decreases adhesion of colorectal carcinoma cells to the substrate . # ::alignments 0-1.5 2-1.5.1.2.1.1.1 4-1.5.1.2 7-1.5.1.1 8-1.5.1 10-1.1 11-1.1.1.r 12-1.1.1.1.1 13-1.3 14-1 15-1.2.1.1.1 16-1.2.1 17-1.2 18-1.4 21-1.6.1 23-1.6.1.1 27-1.7 28-1.7.1.r 29-1.7.1.1.1 30-1.7.1.1 31-1.7.1 32-1.7.1.2 33-1.7.1.2.1.r 34-1.7.1.2.1.1.1 35-1.7.1.2.1.1 36-1.7.1.2.1 37-1.7.1.2.2.r 39-1.7.1.2.2 (d / delay-01~e.14 :ARG0 (d2 / downregulate-01~e.10 :ARG1~e.11 (p / protein :name (n / name :op1 "serpinE2"~e.12))) :ARG1 (d3 / detach-01~e.17 :ARG1 (c / cell-line~e.16 :name (n2 / name :op1 "LoVo"~e.15))) :ARG2 (s / significant-02~e.13) :time (a / after~e.18 :op1 (t / trypsinize-00)) :manner (u / use-01~e.0 :ARG1 (a2 / assay-01~e.8 :ARG1 (a3 / adhere-01~e.7 :polarity -) :ARG1-of (m / mediate-01~e.4 :ARG0 (e / enzyme :name (n4 / name :op1 "trypsin"~e.2))))) :ARG1-of (d4 / describe-01 :ARG0 (f / figure~e.21 :mod "6C"~e.23)) :ARG0-of (s2 / suggest-01~e.27 :ARG1~e.28 (d5 / decrease-01~e.31 :ARG0 (e2 / express-03~e.30 :ARG2 p~e.29) :ARG1 (a4 / adhere-01~e.32 :ARG1~e.33 (c2 / cell~e.36 :source (c3 / carcinoma~e.35 :mod (c4 / colorectal~e.34))) :ARG2~e.37 (s3 / substrate~e.39))))) # ::id a_pmid_2094_2929.100 ::amr-annotator SDL-AMR-09 ::preferred # ::tok SerpinE2 @ gene expression is up @-@ regulated in human colorectal cancers # ::alignments 3-1.1.1 4-1.1 9-1.1.2.r 10-1.1.2.3 11-1.1.2.4 12-1.1.2.2.1 (u / upregulate-01 :ARG1 (e / express-03~e.4 :ARG1 (g / gene~e.3 :name (n / name :op1 "serpinE2")) :ARG3~e.9 (d / disease :wiki "Cancer" :name (n2 / name :op1 "cancer"~e.12) :mod (h / human~e.10) :mod (c2 / colorectal~e.11)))) # ::id a_pmid_2094_2929.101 ::amr-annotator SDL-AMR-09 ::preferred # ::tok We next analyzed serpinE2 @ gene expression in a series of human paired specimens ( resection margins and primary tumors ) by Q @-@ PCR analysis . # ::alignments 0-1.1 1-1.3 2-1 2-1.4.1.2 4-1.2.1.1.1 6-1.2.1 7-1.2 8-1.2.2.r 10-1.2.2 11-1.2.2.1.r 12-1.2.2.1.1 13-1.2.2.1.2 14-1.2.2.1 16-1.2.2.1.3.1.1.1 17-1.2.2.1.3.1.1 18-1.2.2.1.3.1 19-1.2.2.1.3.1.2.1 20-1.2.2.1.3.1.2 23-1.4.1.1 25-1.4.1.1 26-1 26-1.4.1.2 (a / analyze-01~e.2,26 :ARG0 (w / we~e.0) :ARG1 (e / express-03~e.7 :ARG1 (g / gene~e.6 :name (n2 / name :op1 "serpinE2"~e.4)) :ARG3~e.8 (s / series~e.10 :consist-of~e.11 (s2 / specimen~e.14 :mod (h / human~e.12) :ARG1-of (p / pair-01~e.13) :ARG1-of (m / mean-01 :ARG2 (a2 / and~e.18 :op1 (m2 / margin~e.17 :mod (r / resection~e.16)) :op2 (t / tumor~e.20 :mod (p2 / primary~e.19))))))) :time (n / next~e.1) :manner (t2 / thing :name (n3 / name :op1 "Q-PCR"~e.23,25 :op2 "analysis"~e.2,26))) # ::id a_pmid_2094_2929.102 ::amr-annotator SDL-AMR-09 ::preferred # ::tok As shown in Figure 7 , mRNA levels of serpinE2 were markedly increased in human adenomas in comparison to healthy adjacent tissues . # ::alignments 1-1.3 2-1.3.1.r 3-1.3.1 5-1.3.1.1 8-1.1.1.1.1 9-1.1 9-1.4 11-1.1.1.2.1.1.1 13-1.2 13-1.2.r 14-1 15-1.5.r 16-1.5.2 17-1.5.1.1 19-1.4.r 20-1.4.1.r 21-1.4.1.1 22-1.4.1.2 23-1.4.1 (i / increase-01~e.14 :ARG1 (l / level~e.9 :quant-of (n4 / nucleic-acid :name (n / name :op1 "mRNA"~e.8) :ARG0-of (e / encode-01 :ARG1 (p / protein :name (n2 / name :op1 "serpinE2"~e.11))))) :manner~e.13 (m / marked~e.13) :ARG1-of (s / show-01~e.1 :medium~e.2 (f / figure~e.3 :mod 7~e.5)) :compared-to~e.19 (l2 / level~e.9 :location~e.20 (t / tissue~e.23 :mod (h2 / healthy~e.21) :mod (a2 / adjacent~e.22)) :quant-of n4) :location~e.15 (m2 / medical-condition :name (n3 / name :op1 "adenoma"~e.17) :mod (h / human~e.16))) # ::id a_pmid_2094_2929.103 ::amr-annotator SDL-AMR-09 ::preferred # ::tok Furthermore , serpinE2 expression was also significantly enhanced in colorectal tumors , regardless of tumor stage and grade . # ::alignments 0-1 2-1.1.1.1.1.1 3-1.1.1 5-1.1.3 6-1.1.2 7-1.1 8-1.1.1.2.r 9-1.1.1.2.1 10-1.1.1.2 12-1.1.4 14-1.1.4.1.1.1.1 15-1.1.4.1.1 15-1.1.4.1.1.1 15-1.1.4.1.1.1.r 16-1.1.4.1 17-1.1.4.1.2 (a / and~e.0 :op2 (e / enhance-01~e.7 :ARG1 (e2 / express-03~e.3 :ARG2 (p / protein :name (n / name :op1 "serpinE2"~e.2)) :ARG3~e.8 (t / tumor~e.10 :mod (c / colorectal~e.9))) :ARG3 (s / significant-02~e.6) :mod (a2 / also~e.5) :ARG1-of (r / regardless-91~e.12 :ARG2 (a3 / and~e.16 :op1 (t2 / thing~e.15 :ARG2-of~e.15 (s2 / stage-02~e.15 :ARG1 t~e.14)) :op2 (g / grade~e.17 :mod t))))) # ::id a_pmid_2139_2397.12 ::amr-annotator SDL-AMR-09 ::preferred # ::tok The average level of phosphorylation of YB @-@ 1 in the breast cancer cell lines SKBr3 , MCF @-@ 7 , HBL100 and MDA @-@ MB @-@ 231 was significantly higher than that in normal cells . # ::alignments 1-1.1.1 2-1.1 3-1.1.2.r 4-1.1.2 5-1.1.2.1.r 6-1.1.2.1.1.1 8-1.1.2.1.1.1 9-1.1.3.r 11-1.1.3.5.2.1 12-1.1.3.5.2.2 13-1.1.3.1 14-1.1.3.1 14-1.1.3.2 14-1.1.3.3 14-1.1.3.4 15-1.1.3.1.1.1 17-1.1.3.2.1.1 19-1.1.3.2.1.1 21-1.1.3.3.1.1 22-1.1.3 23-1.1.3.4.1.1 25-1.1.3.4.1.1 27-1.1.3.4.1.1 29-1.3 30-1 30-1.2 30-1.2.r 31-1.4.r 34-1.4.1 35-1.4 (h / high-02~e.30 :ARG1 (l / level~e.2 :ARG1-of (a / average-04~e.1) :degree-of~e.3 (p / phosphorylate-01~e.4 :ARG1~e.5 (p2 / protein :name (n / name :op1 "YB-1"~e.6,8))) :location~e.9 (a2 / and~e.22 :op1 (c3 / cell-line~e.13,14 :name (n3 / name :op1 "SKBr3"~e.15)) :op2 (c4 / cell-line~e.14 :name (n4 / name :op1 "MCF-7"~e.17,19)) :op3 (c5 / cell-line~e.14 :name (n5 / name :op1 "HBL100"~e.21)) :op4 (c6 / cell-line~e.14 :name (n6 / name :op1 "MDA-MB-231"~e.23,25,27)) :mod (d / disease :wiki "Breast_cancer" :name (n2 / name :op1 "breast"~e.11 :op2 "cancer"~e.12)))) :degree~e.30 (m / more~e.30) :ARG1-of (s / significant-02~e.29) :compared-to~e.31 (c7 / cell~e.35 :ARG1-of (n7 / normal-02~e.34))) # ::id a_pmid_2139_2397.13 ::amr-annotator SDL-AMR-09 ::preferred # ::tok Exposure to IR and stimulation with erbB1 ligands resulted in phosphorylation of YB @-@ 1 in K @-@ RAS @ wt @ SKBr3 , MCF @-@ 7 and HBL100 cells , which was shown to be K @-@ Ras @-@ independent . # ::alignments 0-1.1.1 1-1.1.1.1.r 2-1.1.1.1 2-1.1.1.1.1 2-1.1.1.1.1.r 3-1.1 4-1.1.2 5-1.1.2.1.r 6-1.1.2.1.1.1 7-1.1.2.1 8-1 9-1.2.r 10-1.2 11-1.2.1.r 12-1.2.1.1.1 14-1.2.1.1.1 17-1.2.2.4.1.1 22-1.2.2.4.2 24-1.2.2.1.1.1 26-1.2.2.2.1.1 28-1.2.2.2.1.1 29-1.2.2 30-1.2.2.3.1.1 31-1.2.2.1 31-1.2.2.2 31-1.2.2.3 35-1.2.3.3 38-1.2.3.2 39-1.2.3.2 40-1.2.3.2 42-1.2.3 42-1.2.3.1 42-1.2.3.1.r (r / result-01~e.8 :ARG1 (a / and~e.3 :op1 (e / expose-01~e.0 :ARG2~e.1 (r2 / radiate-01~e.2 :ARG0-of~e.2 (i / ionize-01~e.2))) :op2 (s / stimulate-01~e.4 :ARG2~e.5 (l / ligand~e.7 :name (n2 / name :op1 "erbB1"~e.6)))) :ARG2~e.9 (p / phosphorylate-01~e.10 :ARG1~e.11 (p2 / protein :name (n3 / name :op1 "YB-1"~e.12,14)) :location (a2 / and~e.29 :op1 (c / cell-line~e.31 :name (n4 / name :op1 "SKBr3"~e.24)) :op2 (c2 / cell-line~e.31 :name (n5 / name :op1 "MCF-7"~e.26,28)) :op3 (c3 / cell-line~e.31 :name (n6 / name :op1 "HBL100"~e.30)) :mod (g / gene :name (n7 / name :op1 "K-Ras"~e.17) :mod (w / wild-type~e.22))) :ARG1-of (d / depend-01~e.42 :polarity~e.42 -~e.42 :ARG0 g~e.38,39,40 :ARG1-of (s2 / show-01~e.35)))) # ::id a_pmid_2139_2397.14 ::amr-annotator SDL-AMR-09 ::preferred # ::tok In contrast , lack of YB @-@ 1 phosphorylation after stimulation with either IR or erbB1 ligands was observed in K @-@ RAS @ mt @ MDA @-@ MB @-@ 231 cells . # ::alignments 1-1 3-1.1.1 4-1.1.1.1.r 5-1.1.1.1.1.1.1 7-1.1.1.1.1.1.1 8-1.1.1.1 9-1.1.1.2 10-1.1.1.2.1 13-1.1.1.2.1.1.1 13-1.1.1.2.1.1.1.1 13-1.1.1.2.1.1.1.1.r 14-1.1.1.2.1.1 15-1.1.1.2.1.1.2.1.1 16-1.1.1.2.1.1.2 18-1.1 21-1.1.2.2.1.1 23-1.1.2.2.1.1 28-1.1.2.1.1 30-1.1.2.1.1 32-1.1.2.1.1 33-1.1.2 (c / contrast-01~e.1 :ARG2 (o / observe-01~e.18 :ARG1 (l / lack-01~e.3 :ARG1~e.4 (p / phosphorylate-01~e.8 :ARG1 (p2 / protein :name (n / name :op1 "YB-1"~e.5,7))) :time (a / after~e.9 :op1 (s / stimulate-01~e.10 :ARG2 (o2 / or~e.14 :op1 (r / radiate-01~e.13 :ARG0-of~e.13 (i / ionize-01~e.13)) :op2 (l3 / ligand~e.16 :name (n3 / name :op1 "erbB1"~e.15)))))) :location (c2 / cell-line~e.33 :name (n4 / name :op1 "MDA-MB-231"~e.28,30,32) :mod (g / gene :name (n5 / name :op1 "K-RAS"~e.21,23) :ARG2-of (m / mutate-01))))) # ::id a_pmid_2139_2397.15 ::amr-annotator SDL-AMR-09 ::preferred # ::tok Similarly to MDA @-@ MB @-@ 231 cells , YB @-@ 1 became constitutively phosphorylated in K @-@ RAS @ wt @ cells following the overexpression of mutated K @-@ RAS , and its phosphorylation was not further enhanced by IR. # ::alignments 0-1.3 1-1.3.1.r 2-1.3.1.1.1 4-1.3.1.1.1 6-1.3.1.1.1 7-1.3.1 9-1.1.1.1.1 11-1.1.1.1.1 12-1.1 13-1.1.2.2 14-1.1.2 17-1.1.2.3.1.1.1 19-1.1.2.3.1.1.1 22-1.1.2.3.1.2 24-1.1.2.3 25-1.1.3 27-1.1.3.1 28-1.1.3.1.1.r 29-1.1.3.1.1.2 31-1.1.3.1.1.1.1 33-1.1.3.1.1.1.1 37-1.1.2.1 37-1.1.2.1.r 38-1.1.2 40-1.2.1 40-1.2.1.r 41-1.2.4 42-1.2 (a / and :op1 (b / become-01~e.12 :ARG1 (p / protein :name (n / name :op1 "YB-1"~e.9,11)) :ARG2 (p2 / phosphorylate-01~e.14,38 :ARG1~e.37 p~e.37 :mod (c / constitutive~e.13) :location (c2 / cell~e.24 :mod (g2 / gene :name (n2 / name :op1 "K-RAS"~e.17,19) :mod (w / wild-type~e.22)))) :ARG2-of (f / follow-01~e.25 :ARG1 (o / overexpress-01~e.27 :ARG1~e.28 (g / gene :name (n3 / name :op1 "K-RAS"~e.31,33) :ARG2-of (m / mutate-01~e.29))))) :op2 (e3 / enhance-01~e.42 :polarity~e.40 -~e.40 :ARG1 p2 :ARG2 (r2 / radiate-01 :ARG0-of (i / ionize-01)) :degree (f2 / further~e.41)) :ARG1-of (r / resemble-01~e.0 :ARG2~e.1 (c3 / cell-line~e.7 :name (n5 / name :op1 "MDA-MB-231"~e.2,4,6)))) # ::id a_pmid_2139_2397.16 ::amr-annotator SDL-AMR-09 ::preferred # ::tok Phosphorylation of YB @-@ 1 as a result of irradiation or K @-@ RAS @ mutation was dependent on erbB1 and its downstream pathways , PI3K and MAPK/ERK . # ::alignments 0-1.1 1-1.1.1.r 2-1.1.1.1.1 4-1.1.1.1.1 5-1.1.2.r 7-1.1.2 8-1.1.2.1.r 9-1.1.2.1.1 12-1.1.2.1.2.1.1.1 14-1.1.2.1.2.1.1.1 16-1.1.2.1.2 18-1 19-1.2.r 20-1.2.1.1.1 21-1.2 22-1.2.2.3 22-1.2.2.3.r 23-1.2.2.1 24-1.2.2 26-1.2.2.2.1.1.1.1 27-1.2.2.2.1 28-1.2.2.2.1.2.1.1 (d / depend-01~e.18 :ARG0 (p2 / phosphorylate-01~e.0 :ARG1~e.1 (p3 / protein :name (n2 / name :op1 "YB-1"~e.2,4)) :ARG2-of~e.5 (r / result-01~e.7 :ARG1~e.8 (a / and :op1 (i / irradiate-01~e.9) :op2 (m / mutate-01~e.16 :ARG1 (g / gene :name (n3 / name :op1 "K-RAS"~e.12,14)))))) :ARG1~e.19 (a3 / and~e.21 :op1 (p / protein :name (n6 / name :op1 "erbB1"~e.20)) :op2 (p4 / pathway~e.24 :mod (d2 / downstream~e.23) :ARG2-of (i2 / include-91 :ARG1 (a2 / and~e.27 :op1 (p5 / pathway :name (n4 / name :op1 "PI3K"~e.26)) :op2 (p6 / pathway :name (n5 / name :op1 "MAPK/ERK"~e.28)))) :poss~e.22 p~e.22))) # ::id a_pmid_2139_2397.17 ::amr-annotator SDL-AMR-09 ::preferred # ::tok In K @-@ RAS @ mt @ cells K @-@ RAS @ siRNA as well as YB @-@ 1 siRNA blocked repair of DNA @-@ DSB . # ::alignments 2-1.1.1.2.1.1.1 2-1.3.1.1.1 4-1.3.1.1.1 9-1.3 11-1.3.1.1.1 13-1.3.1.1.1 15-1.1.1.1.1 15-1.1.2.1.1 16-1.1 17-1.1 18-1.1 19-1.1.2.2.1.1.1 21-1.1.2.2.1.1.1 22-1.1.1.1.1 22-1.1.2.1.1 23-1 24-1.2 25-1.2.1.r 26-1.2.1.1.1 28-1.2.1.1.1 (b / block-01~e.23 :ARG0 (a / and~e.16,17,18 :op1 (n7 / nucleic-acid :name (n2 / name :op1 "siRNA"~e.15,22) :ARG0-of (e2 / encode-01 :ARG1 (g / gene :name (n / name :op1 "K-Ras"~e.2)))) :op2 (n8 / nucleic-acid :name (n3 / name :op1 "siRNA"~e.15,22) :ARG0-of (e3 / encode-01 :ARG1 (p / protein :name (n4 / name :op1 "YB-1"~e.19,21))))) :ARG1 (r3 / repair-01~e.24 :ARG1~e.25 (t / thing :name (n5 / name :op1 "DNA-DSB"~e.26,28))) :location (c / cell~e.9 :mod (g2 / gene :name (n6 / name :op1 "K-RAS"~e.2,4,11,13) :ARG2-of (m / mutate-01)))) # ::id a_pmid_2139_2397.18 ::amr-annotator SDL-AMR-09 ::preferred # ::tok Likewise , YB @-@ 1 siRNA increased radiation sensitivity . # ::alignments 0-1.3 2-1.1.2.1.1.1 4-1.1.2.1.1.1 5-1.1.1.1 6-1 7-1.2.1 8-1.2 (i / increase-01~e.6 :ARG0 (n3 / nucleic-acid :name (n / name :op1 "siRNA"~e.5) :ARG0-of (e / encode-01 :ARG1 (p / protein :name (n2 / name :op1 "YB-1"~e.2,4)))) :ARG1 (s / sensitive-03~e.8 :ARG1 (r2 / radiate-01~e.7)) :manner (l / likewise~e.0)) # ::id a_pmid_2139_2397.115 ::amr-annotator SDL-AMR-09 ::preferred # ::tok Stimulation of YB @-@ 1 phosphorylation in breast cancer cells by IR and exposure to erbB1 ligands # ::alignments 0-1.1 1-1.1.2.r 2-1.1.2.1.1.1 4-1.1.2.1.1.1 5-1.1.2 6-1.1.3.r 7-1.1.3.1.2.1 8-1.1.3.1.2.2 9-1.1.3 10-1.1.1.r 11-1.1.1 11-1.1.1.1 11-1.1.1.1.r 12-1 13-1.2 14-1.2.1.r 15-1.2.1.1.1 16-1.2.1 (a / and~e.12 :op1 (s / stimulate-01~e.0 :ARG0~e.10 (r / radiate-01~e.11 :ARG0-of~e.11 (i / ionize-01~e.11)) :ARG1~e.1 (p2 / phosphorylate-01~e.5 :ARG1 (p3 / protein :name (n2 / name :op1 "YB-1"~e.2,4))) :location~e.6 (c / cell~e.9 :mod (d / disease :wiki "Breast_cancer" :name (n / name :op1 "breast"~e.7 :op2 "cancer"~e.8)))) :op2 (e / expose-01~e.13 :ARG1~e.14 (l / ligand~e.16 :name (n4 / name :op1 "erbB1"~e.15)))) # ::id a_pmid_2139_2397.116 ::amr-annotator SDL-AMR-09 ::preferred # ::tok The level of basal YB @-@ 1 phosphorylation at S102 in a panel of breast cancer cells ( MDA @-@ MB @-@ 231 , MCF @-@ 7 , HBL100 and SKBr3 ) was compared to the level of YB @-@ 1 phosphorylation in normal cells , that is , human skin and lung fibroblasts ( HSF1 , HSF7 and HFL ) as well as normal mammary epithelial cells ( MCF @-@ 10A ) ( Figures 1A and 1B ) . # ::alignments 1-1.1 3-1.1.1.3 4-1.1.1.1.3.1.1 6-1.1.1.1.3.1.1 7-1.1.1 10-1.1.1.2.r 12-1.1.1.2 13-1.1.1.2.1.r 14-1.1.1.2.1.2.2.1 15-1.1.1.2.1.2.2.2 16-1.1.1.2.1 18-1.1.1.2.1.1.1.1.1.1 20-1.1.1.2.1.1.1.1.1.1 22-1.1.1.2.1.1.1.1.1.1 24-1.1.1.2.1.1.1.2.1.1 26-1.1.1.2.1.1.1.2.1.1 28-1.1.1.2.1.1.1.3.1.1 29-1.1.1.2.1.1.1 30-1.1.1.2.1.1.1.4.1.1 33-1 36-1.1 36-1.2 37-1.2.1.r 38-1.2.1.1 39-1.2.1.1 40-1.2.1.1 41-1.2.1 42-1.2.1.2.r 43-1.2.1.2.1 44-1.2.1.2 49-1.2.1.2.2.1.1.1 50-1.2.1.2.2.1.1 51-1.2.1.2.2.1 52-1.2.1.2.2.1.2.1 53-1.2.1.2.2.1.2 55-1.2.1.2.2.1.4.1.1.1.1 57-1.2.1.2.2.1.4.1.2.1.1 59-1.2.1.2.2.1.4.1.3.1.1 61-1.1.1.2.1.1.1 62-1.1.1.2.1.1.1 63-1.1.1.2.1.1.1 63-1.2.1.2.2.1.4.1 64-1.2.1.2.2.1.3.4 65-1.2.1.2.2.1.3.2 66-1.2.1.2.2.1.3.1 67-1.2.1.2.2.1.3 69-1.2.1.2.2.1.3.3.1.1.1 71-1.2.1.2.2.1.3.3.1.1.1 74-1.3.1.1 74-1.3.1.2 76-1.3.1.1.1 78-1.3.1 80-1.3.1.2.1 (c / compare-01~e.33 :ARG1 (l / level~e.1,36 :degree-of (p / phosphorylate-01~e.7 :ARG1 (a / amino-acid :mod 102 :name (n2 / name :op1 "serine") :part-of (p2 / protein :name (n3 / name :op1 "YB-1"~e.4,6))) :location~e.10 (p3 / panel~e.12 :consist-of~e.13 (c2 / cell-line~e.16 :ARG2-of (i / include-91 :ARG1 (a2 / and~e.29,61,62,63 :op1 (c4 / cell-line :name (n4 / name :op1 "MDA-MB-231"~e.18,20,22)) :op2 (c5 / cell-line :name (n5 / name :op1 "MCF-7"~e.24,26)) :op3 (c6 / cell-line :name (n6 / name :op1 "HBL100"~e.28)) :op4 (c7 / cell-line :name (n7 / name :op1 "SKBr3"~e.30)))) :mod (d / disease :wiki "Breast_cancer" :name (n / name :op1 "breast"~e.14 :op2 "cancer"~e.15)))) :mod (b2 / basal~e.3))) :ARG2 (l2 / level~e.36 :degree-of~e.37 (p4 / phosphorylate-01~e.41 :ARG1 p2~e.38,39,40 :location~e.42 (c8 / cell-line~e.44 :ARG1-of (n8 / normal-02~e.43) :ARG2-of (m3 / mean-01 :ARG1 (a4 / and~e.51 :op1 (s / skin~e.50 :mod (h / human~e.49)) :op2 (f / fibroblast~e.53 :mod (l3 / lung~e.52)) :op3 (c12 / cell~e.67 :mod (e / epithelium~e.66) :mod (m / mammary~e.65) :ARG1-of (m2 / mean-01 :ARG2 (c13 / cell-line :name (n14 / name :op1 "MCF-10A"~e.69,71))) :ARG1-of n8~e.64) :ARG2-of (i3 / include-91 :ARG1 (a3 / and~e.63 :op1 (c10 / cell-line :name (n10 / name :op1 "HSF1"~e.55)) :op2 (c9 / cell-line :name (n11 / name :op1 "HSF7"~e.57)) :op3 (c11 / cell-line :name (n12 / name :op1 "HFL"~e.59))))))))) :ARG1-of (d2 / describe-01 :ARG0 (a5 / and~e.78 :op1 (f2 / figure~e.74 :mod "1A"~e.76) :op2 (f3 / figure~e.74 :mod "1B"~e.80)))) # ::id a_pmid_2139_2397.117 ::amr-annotator SDL-AMR-09 ::preferred # ::tok As shown in Figure 1C , the ratio of P @-@ YB @-@ 1/YB @-@ 1 is significantly higher in tumor cells than in fibroblasts . # ::alignments 1-1.2 2-1.2.1.r 3-1.2.1 5-1.2.1.1 9-1.1 10-1.1 11-1.1.1.2 13-1.1.1.1.1 13-1.1.2.1.1 17-1.1.1.1.1 17-1.1.2.1.1 18-1.1.r 19-1.1.3.2 20-1 20-1.1.3 20-1.1.3.1 20-1.1.3.1.r 21-1.1.4.r 22-1.1.4.1 23-1.1.4 24-1.1.5.r 26-1.1.5 (h2 / high~e.20 :domain~e.18 (r / ratio-of~e.9,10 :op1 (p2 / protein :name (n / name :op1 "YB-1"~e.13,17) :ARG3-of (p / phosphorylate-01~e.11)) :op2 (p3 / protein :name (n2 / name :op1 "YB-1"~e.13,17)) :ARG1-of (h / high-02~e.20 :degree~e.20 (m / more~e.20) :ARG1-of (s2 / significant-02~e.19)) :location~e.21 (c / cell~e.23 :mod (t / tumor~e.22)) :compared-to~e.24 (f2 / fibroblast~e.26)) :ARG1-of (s / show-01~e.1 :ARG0~e.2 (f / figure~e.3 :mod "1C"~e.5))) # ::id a_pmid_2139_2397.118 ::amr-annotator SDL-AMR-09 ::preferred # ::tok The comparisons of the ratio of P @-@ YB @-@ 1/YB @-@ 1 in tumor cells and normal mammary epithelial cells indicated an even stronger significant difference as tested for MDA @-@ MB @-@ 231 and MCF @-@ 10A cells ( Figures 1B and 1C ) . # ::alignments 1-1.1 2-1.1.1 4-1.1.1 5-1.1.1 6-1.1.1.1.2 8-1.1.1.1.1.1 8-1.1.1.2.1.1 12-1.1.1.1.1.1 12-1.1.1.2.1.1 13-1.1.2.r 14-1.1.2.1.1 15-1.1.2.1 15-1.1.2.2 16-1.1.2 17-1.1.2.2.3 18-1.1.2.2.2 19-1.1.2.2.1 20-1.1.2.2 21-1 23-1.2.1.2 24-1.2.1 24-1.2.1.1 24-1.2.1.1.r 25-1.2.2 26-1.2 27-1.2.3.r 28-1.2.3 29-1.2.3.1.r 30-1.2.3.1.1.1.1 32-1.2.3.1.1.1.1 34-1.2.3.1.1.1.1 35-1.2.3.1 36-1.2.3.1.2.1.1 38-1.2.3.1.2.1.1 39-1.2.3.1.1 41-1.3.1.1 41-1.3.1.2 43-1.3.1.1.1 45-1.3.1 47-1.3.1.2.1 (i / indicate-01~e.21 :ARG0 (c / compare-01~e.1 :ARG2 (r / ratio-of~e.2,4,5 :op1 (p2 / protein :name (n / name :op1 "YB-1"~e.8,12) :ARG3-of (p / phosphorylate-01~e.6)) :op2 (p3 / protein :name (n2 / name :op1 "YB-1"~e.8,12))) :location~e.13 (a3 / and~e.16 :op1 (c2 / cell~e.15 :mod (t / tumor~e.14)) :op2 (c3 / cell~e.15,20 :mod (e / epithelium~e.19) :mod (m / mammary~e.18) :ARG1-of (n5 / normal-02~e.17)))) :ARG1 (d / differ-02~e.26 :mod (s / strong~e.24 :degree~e.24 (m2 / more~e.24) :mod (e2 / even~e.23)) :ARG1-of (s2 / significant-02~e.25) :ARG1-of~e.27 (t2 / test-01~e.28 :ARG2~e.29 (a / and~e.35 :op1 (c4 / cell-line~e.39 :name (n3 / name :op1 "MDA-MB-231"~e.30,32,34)) :op2 (c5 / cell-line :name (n4 / name :op1 "MCF-10A"~e.36,38))))) :ARG1-of (d2 / describe-01 :ARG0 (a2 / and~e.45 :op1 (f / figure~e.41 :mod "1B"~e.43) :op2 (f2 / figure~e.41 :mod "1C"~e.47)))) # ::id a_pmid_2139_2397.119 ::amr-annotator SDL-AMR-09 ::preferred # ::tok YB @-@ 1 has been identified as a direct substrate of Akt [ @ 12 , 35 @ ] . # ::alignments 0-1.1.1.1 2-1.1.1.1 5-1 6-1.2.r 8-1.2.2 9-1.2 10-1.2.1.r 11-1.2.1.1.1 14-1.3.1.1.1.1 18-1.3.1.1.1.2 (i / identify-01~e.5 :ARG1 (p / protein :name (n / name :op1 "YB-1"~e.0,2)) :ARG2~e.6 (s / substrate~e.9 :mod~e.10 (e / enzyme :name (n2 / name :op1 "Akt"~e.11)) :ARG1-of (d / direct-02~e.8)) :ARG1-of (d2 / describe-01 :ARG0 (p2 / publication :ARG1-of (c / cite-01 :ARG2 (a / and :op1 12~e.14 :op2 35~e.18))))) # ::id a_pmid_2139_2397.120 ::amr-annotator SDL-AMR-09 ::preferred # ::tok As previously reported , IR can activate the Akt ligand independently [ @ 30 , 36 @ ] . # ::alignments 0-1.3.1.r 1-1.3.1 2-1.3 4-1.1.1 4-1.1.1.1 4-1.1.1.1.r 5-1 6-1.1 8-1.1.2.1.1.1 9-1.1.2 10-1.1.3.1 13-1.2.1.1.1.1 17-1.2.1.1.1.2 (p / possible-01~e.5 :ARG1 (a / activate-01~e.6 :ARG0 (r2 / radiate-01~e.4 :ARG0-of~e.4 (i / ionize-01~e.4)) :ARG1 (l / ligand~e.9 :mod (e / enzyme :name (n3 / name :op1 "Akt"~e.8))) :manner (d2 / depend-01 :polarity -~e.10)) :ARG1-of (d / describe-01 :ARG0 (p3 / publication :ARG1-of (c / cite-01 :ARG2 (a2 / and :op1 30~e.13 :op2 36~e.17)))) :ARG1-of (r / report-01~e.2 :time~e.0 (p2 / previous~e.1))) # ::id a_pmid_2139_2397.121 ::amr-annotator SDL-AMR-09 ::preferred # ::tok Therefore , we asked whether IR could induce YB @-@ 1 phosphorylation as well . # ::alignments 0-1 2-1.1.2 3-1.1 4-1.1.1 4-1.1.1.r 5-1.1.3.1.1 5-1.1.3.1.1.1 5-1.1.3.1.1.1.r 6-1.1.3 7-1.1.3.1 8-1.1.3.1.2.1.1.1 10-1.1.3.1.2.1.1.1 11-1.1.3.1.2 12-1.1.3.1.3 13-1.1.3.1.3 (c / cause-01~e.0 :ARG1 (a / ask-01~e.3 :mode~e.4 interrogative~e.4 :ARG0 (w / we~e.2) :ARG1 (p3 / possible-01~e.6 :ARG1 (i / induce-01~e.7 :ARG0 (r / radiate-01~e.5 :ARG0-of~e.5 (i2 / ionize-01~e.5)) :ARG2 (p2 / phosphorylate-01~e.11 :ARG1 (p4 / protein :name (n2 / name :op1 "YB-1"~e.8,10))) :mod (a2 / as-well~e.12,13))))) # ::id a_pmid_2139_2397.122 ::amr-annotator SDL-AMR-09 ::preferred # ::tok As shown in Figure 1D , IR induces YB @-@ 1 phosphorylation differentially . # ::alignments 1-1.3 2-1.3.1.r 3-1.3.1 5-1.3.1.1 8-1.1 8-1.1.1 8-1.1.1.r 9-1 10-1.2.1.1.1 12-1.2.1.1.1 13-1.2 14-1.4 14-1.4.r (i / induce-01~e.9 :ARG0 (r / radiate-01~e.8 :ARG0-of~e.8 (i2 / ionize-01~e.8)) :ARG2 (p / phosphorylate-01~e.13 :ARG1 (p2 / protein :name (n2 / name :op1 "YB-1"~e.10,12))) :ARG1-of (s / show-01~e.1 :ARG0~e.2 (f / figure~e.3 :mod "1D"~e.5)) :manner~e.14 (d / differential~e.14)) # ::id a_pmid_2139_2397.123 ::amr-annotator SDL-AMR-09 ::preferred # ::tok A strong phosphorylation signal was observed in SKBr3 , whereas HBL100 showed moderate phosphorylation of YB @-@ 1 and phosphorylation in MCF @-@ 7 was weak . # ::alignments 1-1.1.1.2 2-1.1.1.1 3-1.1.1 5-1.1 6-1.1.1.3.r 7-1.1.1.3.1.1 9-1 10-1.2.1.1.1.1 11-1.2.1 12-1.2.1.2.2 13-1.2.1.2 14-1.2.1.2.1.r 15-1.2.1.2.1.1.1 17-1.2.1.2.1.1.1 18-1.2 19-1.2.2 20-1.2.2.1.r 21-1.2.2.1.1.1 23-1.2.2.1.1.1 25-1.2.2.2 (c / contrast-01~e.9 :ARG1 (o / observe-01~e.5 :ARG1 (s / signal-07~e.3 :ARG1 (p / phosphorylate-01~e.2) :mod (s2 / strong~e.1) :location~e.6 (c2 / cell-line :name (n / name :op1 "SKBr3"~e.7)))) :ARG2 (a / and~e.18 :op1 (s3 / show-01~e.11 :ARG0 (c3 / cell-line :name (n2 / name :op1 "HBL100"~e.10)) :ARG1 (p2 / phosphorylate-01~e.13 :ARG1~e.14 (p3 / protein :name (n3 / name :op1 "YB-1"~e.15,17)) :ARG1-of (m / moderate-03~e.12))) :op2 (p4 / phosphorylate-01~e.19 :location~e.20 (c4 / cell-line :name (n4 / name :op1 "MCF-7"~e.21,23)) :ARG1-of (w / weak-02~e.25)))) # ::id a_pmid_2139_2397.124 ::amr-annotator SDL-AMR-09 ::preferred # ::tok However , in MDA @-@ MB @-@ 231 cells , a lack of IR @-@ induced YB @-@ 1 phosphorylation was observed . # ::alignments 0-1 3-1.1.1.2.1.1 5-1.1.1.2.1.1 7-1.1.1.2.1.1 8-1.1.1.2 11-1.1.1 12-1.1.1.1.r 13-1.1.1.1.2.1 13-1.1.1.1.2.1.1 13-1.1.1.1.2.1.1.r 15-1.1.1.1.2 16-1.1.1.1.1.1.1 18-1.1.1.1.1.1.1 19-1.1.1.1 21-1.1 (h / have-concession-91~e.0 :ARG1 (o / observe-01~e.21 :ARG1 (l / lack-01~e.11 :ARG1~e.12 (p / phosphorylate-01~e.19 :ARG1 (p2 / protein :name (n / name :op1 "YB-1"~e.16,18)) :ARG2-of (i / induce-01~e.15 :ARG0 (r / radiate-01~e.13 :ARG0-of~e.13 (i2 / ionize-01~e.13)))) :location (c / cell-line~e.8 :name (n3 / name :op1 "MDA-MB-231"~e.3,5,7))))) # ::id a_pmid_2139_2397.125 ::amr-annotator SDL-AMR-09 ::preferred # ::tok In this cell line , stimulation with the erbB1 ligand EGF , AREG or TGFα did not induce YB @-@ 1 phosphorylation , whereas strong phosphorylation at the indicated times after stimulation was observed in the cell lines SKBr3 , HBL100 and MCF @-@ 7 ( Figure 1D ) . # ::alignments 1-1.1.4.1 2-1.1.4 3-1.1.4 5-1.1.2 6-1.1.2.1.r 8-1.1.2.1.1.1 9-1.1.2.1 10-1.1.2.1.2.1.1.1.1 12-1.1.2.1.2.1.2.1.1 14-1.1.2.1.2.1.3.1.1 16-1.1.1 16-1.1.1.r 17-1.1 18-1.1.3.1.1.1 20-1.1.3.1.1.1 21-1.1.3 21-1.2.1 23-1 24-1.2.1.1 25-1.2.1 28-1.2.1.2.1 29-1.2.1.2 29-1.2.1.2.r 29-1.2.1.3.r 30-1.2.1.3 31-1.2.1.3.1 33-1.2 34-1.2.2.r 36-1.2.2.1 37-1.2.2.1 37-1.2.2.2 37-1.2.2.3 38-1.2.2.1.1.1 40-1.2.2.2.1.1 41-1.2.2 42-1.2.2.3.1.1 44-1.2.2.3.1.1 46-1.3.1 48-1.3.1.1 (c / contrast-01~e.23 :ARG1 (i / induce-01~e.17 :polarity~e.16 -~e.16 :ARG0 (s / stimulate-01~e.5 :ARG2~e.6 (l / ligand~e.9 :name (n / name :op1 "erbB1"~e.8) :ARG2-of (i2 / include-91 :ARG1 (a / and :op1 (p6 / protein :name (n2 / name :op1 "EGF"~e.10)) :op2 (p2 / protein :name (n3 / name :op1 "AREG"~e.12)) :op3 (p3 / protein :name (n4 / name :op1 "TGFα"~e.14)))))) :ARG2 (p / phosphorylate-01~e.21 :ARG1 (p4 / protein :name (n5 / name :op1 "YB-1"~e.18,20))) :location (c5 / cell-line~e.2,3 :mod (t2 / this~e.1))) :ARG2 (o / observe-01~e.33 :ARG1 (p5 / phosphorylate-01~e.21,25 :mod (s3 / strong~e.24) :time~e.29 (t / time~e.29 :ARG1-of (i3 / indicate-01~e.28)) :time~e.29 (a2 / after~e.30 :op1 s~e.31)) :location~e.34 (a3 / and~e.41 :op1 (c2 / cell-line~e.36,37 :name (n6 / name :op1 "SKBr3"~e.38)) :op2 (c3 / cell-line~e.37 :name (n7 / name :op1 "HBL100"~e.40)) :op3 (c4 / cell-line~e.37 :name (n8 / name :op1 "MCF-7"~e.42,44)))) :ARG1-of (d / describe-01 :ARG0 (f / figure~e.46 :mod "1D"~e.48))) # ::id a_pmid_2139_2397.126 ::amr-annotator SDL-AMR-09 ::preferred # ::tok Although the MCF @-@ 7 and HBL100 cell lines have K @-@ RAS @ wt @ status , these cells presented high basal YB @-@ 1 phosphorylation . # ::alignments 0-1 2-1.2.1.1.1.1 4-1.2.1.1.1.1 5-1.2.1 6-1.2.1.2.1.1 7-1.2.1.1 7-1.2.1.2 8-1.2.1.1 9-1.2 11-1.2.2.1.1.1 13-1.2.2.1.1.1 16-1.2.2.1.2 18-1.2.2 21-1.2.1.1 22-1.1 23-1.1.2.2 24-1.1.2.3 25-1.1.2.1.1.1 27-1.1.2.1.1.1 28-1.1.2 (h / have-concession-91~e.0 :ARG1 (p2 / present-01~e.22 :ARG0 a :ARG1 (p / phosphorylate-01~e.28 :ARG1 (p3 / protein :name (n4 / name :op1 "YB-1"~e.25,27)) :ARG1-of (h3 / high-02~e.23) :mod (b / basal~e.24))) :ARG2 (h2 / have-03~e.9 :ARG0 (a / and~e.5 :op1 (c / cell-line~e.7,8,21 :name (n2 / name :op1 "MCF-7"~e.2,4)) :op2 (c2 / cell-line~e.7 :name (n3 / name :op1 "HBL100"~e.6))) :ARG1 (s / status~e.18 :mod (g / gene :name (n / name :op1 "K-RAS"~e.11,13) :mod (w / wild-type~e.16))))) # ::id a_pmid_2139_2397.127 ::amr-annotator SDL-AMR-09 ::preferred # ::tok To prove whether the high basal phosphorylation status of YB @-@ 1 was due to stimulation by growth factors in the culture medium , P @-@ YB @-@ 1 was compared under serum supplementation and serum depletion in MCF @-@ 7 cells . # ::alignments 0-1.3.1 1-1.3 2-1.3.1.1 2-1.3.1.1.r 4-1.3.1.3.1.2 5-1.3.1.3.1.3 6-1.3.1.3.1 7-1.3.1.3 8-1.3.1.3.1.1.r 9-1.3.1.3.1.1.1.1 11-1.3.1.3.1.1.1.1 13-1.3.1 14-1.3.1 15-1.3.1.2 16-1.3.1.2.1.r 17-1.3.1.2.1 18-1.3.1.2.1 19-1.3.1.2.2.r 21-1.3.1.2.2.1 22-1.3.1.2.2 24-1.2.2 26-1.2.1.1 28-1.2.1.1 30-1 32-1.1.1.1 33-1.1.1 35-1.1.1.1 36-1.1.2 37-1.1.r 38-1.1.3.1.1 40-1.1.3.1.1 41-1.1.3 (c5 / compare-01~e.30 :ARG0~e.37 (a / and :op1 (s4 / supplement-01~e.33 :ARG1 (s5 / serum~e.32,35)) :op2 (d / deplete-01~e.36 :ARG1 s5) :location (c4 / cell-line~e.41 :name (n3 / name :op1 "MCF-7"~e.38,40))) :ARG1 (p4 / protein :name (n4 / name :op1 "YB-1"~e.26,28) :ARG3-of (p5 / phosphorylate-01~e.24)) :purpose (p2 / prove-01~e.1 :ARG1 (c / cause-01~e.0,13,14 :mode~e.2 interrogative~e.2 :ARG0 (s2 / stimulate-01~e.15 :ARG0~e.16 (g / growth-factor~e.17,18) :location~e.19 (m / medium~e.22 :mod (c2 / culture~e.21))) :ARG1 (s / status~e.7 :mod (p / phosphorylate-01~e.6 :ARG1~e.8 (p3 / protein :name (n / name :op1 "YB-1"~e.9,11)) :degree (h / high-02~e.4) :mod (b / basal~e.5)))))) # ::id a_pmid_2139_2397.128 ::amr-annotator SDL-AMR-09 ::preferred # ::tok As shown in Figure 1F , P @-@ YB @-@ 1 was markedly reduced when cells were incubated in serum @-@ free medium for 24 hours . # ::alignments 0-1.3.r 1-1.4 2-1.4.1.r 3-1.4.1 5-1.4.1.1 8-1.1.2 10-1.1.1.1 12-1.1.1.1 14-1.2 14-1.2.r 15-1 16-1.3.r 17-1.3.1 19-1.3 20-1.3.2.r 21-1.3.2.1.1 23-1.3.2.1 24-1.3.2 25-1.3.3.r 26-1.3.3.1 27-1.3.3.2 (r / reduce-01~e.15 :ARG1 (p / protein :name (n / name :op1 "YB-1"~e.10,12) :ARG3-of (p2 / phosphorylate-01~e.8)) :manner~e.14 (m / marked~e.14) :time~e.0,16 (i / incubate-01~e.19 :ARG1 (c / cell~e.17) :ARG2~e.20 (m2 / medium~e.24 :ARG1-of (f / free-04~e.23 :ARG2 (s / serum~e.21))) :duration~e.25 (t2 / temporal-quantity :quant 24~e.26 :unit (h2 / hour~e.27))) :ARG1-of (s2 / show-01~e.1 :ARG0~e.2 (f2 / figure~e.3 :mod "1F"~e.5))) # ::id a_pmid_2139_2397.129 ::amr-annotator SDL-AMR-09 ::preferred # ::tok In contrast , serum depletion did not reduce basal YB @-@ 1 phosphorylation in K @-@ RAS @ mt @ MDA @-@ MB @-@ 231 cells ( Figure 1F ) . # ::alignments 1-1 3-1.1.2.1 4-1.1.2 6-1.1.1 6-1.1.1.r 7-1.1 8-1.1.3.2 9-1.1.3.1.1.1 11-1.1.3.1.1.1 12-1.1.3 15-1.1.4.2.1.1 17-1.1.4.2.1.1 22-1.1.4.1.1 24-1.1.4.1.1 26-1.1.4.1.1 27-1.1.4 29-1.2.1 31-1.2.1.1 (c / contrast-01~e.1 :ARG2 (r / reduce-01~e.7 :polarity~e.6 -~e.6 :ARG0 (d / deplete-01~e.4 :ARG1 (s / serum~e.3)) :ARG1 (p / phosphorylate-01~e.12 :ARG1 (p2 / protein :name (n2 / name :op1 "YB-1"~e.9,11)) :mod (b / basal~e.8)) :location (c2 / cell-line~e.27 :name (n3 / name :op1 "MDA-MB-231"~e.22,24,26) :mod (g / gene :name (n / name :op1 "K-RAS"~e.15,17) :ARG2-of (m / mutate-01)))) :ARG1-of (d2 / describe-01 :ARG0 (f / figure~e.29 :mod "1F"~e.31))) # ::id a_pmid_2139_2397.130 ::amr-annotator SDL-AMR-09 ::preferred # ::tok Constitutive phosphorylation of YB @-@ 1 in MDA @-@ MB @-@ 231 cells is K @-@ Ras @-@ dependent # ::alignments 0-1.1.2 1-1.1 2-1.1.1.r 3-1.1.1.1.1 5-1.1.1.1.1 6-1.1.3.r 7-1.1.3.1.1 9-1.1.3.1.1 11-1.1.3.1.1 12-1.1.3 14-1.2.1.1 16-1.2.1.1 18-1 (d / depend-01~e.18 :ARG0 (p / phosphorylate-01~e.1 :ARG1~e.2 (p2 / protein :name (n / name :op1 "YB-1"~e.3,5)) :mod (c / constitutive~e.0) :location~e.6 (c2 / cell-line~e.12 :name (n2 / name :op1 "MDA-MB-231"~e.7,9,11))) :ARG1 (e / enzyme :name (n3 / name :op1 "K-Ras"~e.14,16))) # ::id a_pmid_2139_2397.131 ::amr-annotator SDL-AMR-09 ::preferred # ::tok MDA @-@ MB @-@ 231 cells are characterized by a point mutation at codon 13 in the K @-@ RAS @ gene [ @ 37 @ ] . # ::alignments 0-1.2.1.1 2-1.2.1.1 4-1.2.1.1 5-1.2 7-1 8-1.1.r 10-1.1.2 11-1.1 12-1.1.1.r 13-1.1.1 14-1.1.1.1 18-1.1.1.2.1.1 20-1.1.1.2.1.1 22-1.1.1.2 25-1.3.1.1.1 (c / characterize-01~e.7 :ARG0~e.8 (m / mutate-01~e.11 :ARG1~e.12 (c3 / codon~e.13 :mod 13~e.14 :part-of (g / gene~e.22 :name (n3 / name :op1 "K-RAS"~e.18,20))) :mod (p / point~e.10)) :ARG1 (c2 / cell-line~e.5 :name (n2 / name :op1 "MDA-MB-231"~e.0,2,4)) :ARG1-of (d / describe-01 :ARG0 (p2 / publication :ARG1-of (c4 / cite-01 :ARG2 37~e.25)))) # ::id a_pmid_2139_2397.132 ::amr-annotator SDL-AMR-09 ::preferred # ::tok This mutation is responsible for the constitutive phosphorylation of ERK1 @/@ 2 [ @ 30 @ ] . # ::alignments 0-1.1.1 1-1.1 3-1 4-1.2.r 6-1.2.2 7-1.2 8-1.2.1.r 9-1.2.1.1.1 11-1.2.1.1.1 14-1.3.1.1.1 (r / responsible-01~e.3 :ARG0 (m / mutate-01~e.1 :mod (t / this~e.0)) :ARG1~e.4 (p / phosphorylate-01~e.7 :ARG1~e.8 (e / enzyme :name (n / name :op1 "ERK1/2"~e.9,11)) :mod (c / constitutive~e.6)) :ARG1-of (d / describe-01 :ARG0 (p2 / publication :ARG1-of (c2 / cite-01 :ARG2 30~e.14)))) # ::id a_pmid_2139_2397.133 ::amr-annotator SDL-AMR-09 ::preferred # ::tok In addition to ERK1 @/@ 2 phosphorylation , these cells also present a constitutive phosphorylation of YB @-@ 1 , which is not further modified after exposure to IR or stimulation with erbB1 ligands ( Figures 1D and 1E ) . # ::alignments 0-1.5.1 1-1.4 3-1.4.1.1.1.1 5-1.4.1.1.1.1 6-1.2 6-1.4.1 8-1.1.1 9-1.1 10-1.3 11-1 13-1.2.2 14-1.2 15-1.2.1.r 16-1.2.1.1.1 18-1.2.1.1.1 22-1.2.3.1 22-1.2.3.1.r 23-1.2.3.3 24-1.2.3 26-1.2.3.2.1 27-1.2.3.2.1.2.r 28-1.2.3.2.1.2 28-1.2.3.2.1.2.1 28-1.2.3.2.1.2.1.r 29-1.2.3.2 30-1.2.3.2.2 31-1.2.3.2.2.2.r 32-1.2.3.2.2.2.1.1 33-1.2.3.2.2.2 35-1.5.1.1 35-1.5.1.2 37-1.5.1.1.1 39-1.5.1 41-1.5.1.2.1 (p5 / present-01~e.11 :ARG0 (c / cell~e.9 :mod (t / this~e.8)) :ARG1 (p3 / phosphorylate-01~e.6,14 :ARG1~e.15 (p4 / protein :name (n2 / name :op1 "YB-1"~e.16,18)) :mod (c2 / constitutive~e.13) :ARG1-of (m / modify-01~e.24 :polarity~e.22 -~e.22 :ARG0 (o / or~e.29 :op1 (e2 / expose-01~e.26 :ARG1 c :ARG2~e.27 (r / radiate-01~e.28 :ARG0-of~e.28 (i / ionize-01~e.28))) :op2 (s / stimulate-01~e.30 :ARG1 c :ARG2~e.31 (l / ligand~e.33 :name (n4 / name :op1 "erbB1"~e.32)))) :degree (f3 / further~e.23))) :mod (a2 / also~e.10) :ARG1-of (a4 / add-02~e.1 :ARG2 (p / phosphorylate-01~e.6 :ARG1 (e / enzyme :name (n / name :op1 "ERK1/2"~e.3,5)))) :ARG1-of (d / describe-01 :ARG0 (a3 / and~e.0,39 :op1 (f / figure~e.35 :mod "1D"~e.37) :op2 (f2 / figure~e.35 :mod "1E"~e.41)))) # ::id a_pmid_2139_2397.134 ::amr-annotator SDL-AMR-09 ::preferred # ::tok Thus , we investigated whether the constitutive phosphorylation of YB @-@ 1 in MDA @-@ MB @-@ 231 cells is due to the described endogenous expression of mutated K @-@ RAS @ [ @ 37 @ ] . # ::alignments 0-1.1.2 2-1.1.1 3-1.1 4-1.1.2.1 4-1.1.2.1.r 6-1.1.2.3.3 7-1.1.2.3 8-1.1.2.3.1.r 9-1.1.2.3.1.1.1 11-1.1.2.3.1.1.1 12-1.1.2.3.2.r 13-1.1.2.3.2.1.1 15-1.1.2.3.2.1.1 17-1.1.2.3.2.1.1 18-1.1.2.3.2 20-1 21-1 23-1.1.2.2.2 23-1.2 24-1.1.2.2.3 25-1.1.2.2 26-1.1.2.2.1.r 27-1.1.2.2.1.2 29-1.1.2.2.1.1.1 31-1.1.2.2.1.1.1 35-1.2.1.1.1 (c / cause-01~e.20,21 :ARG1 (i / investigate-01~e.3 :ARG0 (w / we~e.2) :ARG1 (c2 / cause-01~e.0 :mode~e.4 interrogative~e.4 :ARG0 (e2 / express-03~e.25 :ARG2~e.26 (g / gene :name (n / name :op1 "K-RAS"~e.29,31) :ARG2-of (m2 / mutate-01~e.27)) :ARG1-of (d / describe-01~e.23) :mod (m / monocot~e.24)) :ARG1 (p / phosphorylate-01~e.7 :ARG1~e.8 (p2 / protein :name (n2 / name :op1 "YB-1"~e.9,11)) :location~e.12 (c3 / cell-line~e.18 :name (n3 / name :op1 "MDA-MB-231"~e.13,15,17)) :mod (c4 / constitutive~e.6)))) :ARG1-of (d2 / describe-01~e.23 :ARG0 (p3 / publication :ARG1-of (c5 / cite-01 :ARG2 37~e.35)))) # ::id a_pmid_2139_2397.135 ::amr-annotator SDL-AMR-09 ::preferred # ::tok Therefore , K @-@ Ras expression was downregulated by siRNA , and the level of P @-@ YB @-@ 1 was investigated . # ::alignments 0-1 2-1.1.1.1.1.1.1 4-1.1.1.1.1.1.1 5-1.1.1.1 7-1.1.1 8-1.1.1.2.r 9-1.1.1.2.1.1 11-1.1 13-1.1.2.1 14-1.1.2.1.1.r 15-1.1.2.1.1.2 17-1.1.2.1.1.1.1 19-1.1.2.1.1.1.1 21-1.1.2 (c / cause-01~e.0 :ARG1 (a / and~e.11 :op1 (d / downregulate-01~e.7 :ARG1 (e2 / express-03~e.5 :ARG2 (e / enzyme :name (n / name :op1 "K-Ras"~e.2,4))) :ARG2~e.8 (n4 / nucleic-acid :name (n2 / name :op1 "siRNA"~e.9))) :op2 (i / investigate-01~e.21 :ARG1 (l / level~e.13 :quant-of~e.14 (p / protein :name (n3 / name :op1 "YB-1"~e.17,19) :ARG3-of (p2 / phosphorylate-01~e.15)))))) # ::id a_pmid_2139_2397.136 ::amr-annotator SDL-AMR-09 ::preferred # ::tok Using a similar approach , we analyzed the effect of ERK1 on YB @-@ 1 phosphorylation downstream of mutated K @-@ Ras . # ::alignments 0-1.3 2-1.3.1.1 3-1.3.1 5-1.1 6-1 8-1.2 9-1.2.1.r 10-1.2.1.1.1 11-1.2.2.r 12-1.2.2.1.1.1 14-1.2.2.1.1.1 15-1.2.2 16-1.2.2.2.2 18-1.2.2.2.1.2 19-1.2.2.2.1.1.1 21-1.2.2.2.1.1.1 (a4 / analyze-01~e.6 :ARG0 (w / we~e.5) :ARG1 (a3 / affect-01~e.8 :ARG0~e.9 (e2 / enzyme :name (n2 / name :op1 "ERK1"~e.10)) :ARG1~e.11 (p / phosphorylate-01~e.15 :ARG1 (p2 / protein :name (n3 / name :op1 "YB-1"~e.12,14)) :location (r2 / relative-position :op1 (e / enzyme :name (n / name :op1 "K-Ras"~e.19,21) :ARG2-of (m / mutate-01~e.18)) :direction (d / downstream~e.16)))) :manner (u / use-01~e.0 :ARG1 (a / approach-02~e.3 :ARG1-of (r / resemble-01~e.2)))) # ::id a_pmid_2139_2397.137 ::amr-annotator SDL-AMR-09 ::preferred # ::tok As shown in Figure 2A , K @-@ RAS @ siRNA led to a strong reduction in P @-@ ERK1 @/@ 2 and P @-@ YB @-@ 1 ( Figure 2A ) . # ::alignments 1-1.3 2-1.3.1.r 3-1.3.1 5-1.3.1.1 9-1.1.2.1.1.1 13-1.1.1.1 14-1 15-1.2.r 17-1.2.2 18-1.2 19-1.2.1.r 20-1.2.1.1 20-1.2.1.1.2 20-1.2.1.1.2.r 20-1.2.1.2.2 22-1.2.1.1.1.1 24-1.2.1.1.1.1 25-1.2.1 26-1.2.1.1 26-1.2.1.1.2 26-1.2.1.1.2.r 26-1.2.1.2.2 28-1.2.1.2.1.1 30-1.2.1.2.1.1 32-1.3.1 34-1.3.1.1 (l / lead-03~e.14 :ARG0 (n5 / nucleic-acid :name (n2 / name :op1 "siRNA"~e.13) :ARG0-of (e2 / encode-01 :ARG1 (g / gene :name (n / name :op1 "K-Ras"~e.9)))) :ARG2~e.15 (r2 / reduce-01~e.18 :ARG1~e.19 (a / and~e.25 :op1 (e3 / enzyme~e.20,26 :name (n3 / name :op1 "ERK1/2"~e.22,24) :ARG1-of~e.20,26 (p / phosphorylate-01~e.20,26)) :op2 (p2 / protein :name (n4 / name :op1 "YB-1"~e.28,30) :ARG3-of (p3 / phosphorylate-01~e.20,26))) :mod (s / strong~e.17)) :ARG1-of (s2 / show-01~e.1 :ARG0~e.2 (f / figure~e.3,32 :mod "2A"~e.5,34))) # ::id a_pmid_2139_2397.138 ::amr-annotator SDL-AMR-09 ::preferred # ::tok Yet , ERK1 @/@ 2 and YB @-@ 1 protein levels were not affected . # ::alignments 2-1.1.2.1.1.1.1 4-1.1.2.1.1.1.1 5-1.1.2 6-1.1.2.2.1.1.1 8-1.1.2.2.1.1.1 9-1.1.2.2.1 10-1.1.2.1 10-1.1.2.2 12-1.1.1 12-1.1.1.r 13-1.1 (h / have-concession-91 :ARG1 (a / affect-01~e.13 :polarity~e.12 -~e.12 :ARG1 (a2 / and~e.5 :op1 (l / level~e.10 :quant-of (e / enzyme :name (n / name :op1 "ERK1/2"~e.2,4))) :op2 (l2 / level~e.10 :quant-of (p / protein~e.9 :name (n2 / name :op1 "YB-1"~e.6,8)))))) # ::id a_pmid_2139_2397.139 ::amr-annotator SDL-AMR-09 ::preferred # ::tok Likewise , a marked reduction of P @-@ YB @-@ 1 was observed when ERK1 was targeted with siRNA . # ::alignments 0-1.2 3-1.1.3 4-1.1 5-1.1.1.r 6-1.1.1.2 8-1.1.1.1.1 10-1.1.1.1.1 12-1 13-1.1.2.r 14-1.1.2.1.1.1 16-1.1.2 17-1.1.2.2.r 18-1.1.2.2.1.1 (o / observe-01~e.12 :ARG1 (r / reduce-01~e.4 :ARG1~e.5 (p / protein :name (n / name :op1 "YB-1"~e.8,10) :ARG3-of (p2 / phosphorylate-01~e.6)) :time~e.13 (t / target-01~e.16 :ARG1 (e / enzyme :name (n2 / name :op1 "ERK1"~e.14)) :ARG2~e.17 (n4 / nucleic-acid :name (n3 / name :op1 "siRNA"~e.18))) :degree (m / marked~e.3)) :manner (l / likewise~e.0)) # ::id a_pmid_2139_2397.140 ::amr-annotator SDL-AMR-09 ::preferred # ::tok The role of stimulated ERK1 @/@ 2 phosphorylation on YB @-@ 1 phosphorylation was further supported by the results when a MEK inhibitor was used . # ::alignments 1-1.2 2-1.2.1.r 3-1.2.1.2 4-1.2.1.1.1.1 6-1.2.1.1.1.1 7-1.2.1 7-1.2.2 8-1.2.2.1.r 9-1.2.2.1.1.1 11-1.2.2.1.1.1 12-1.2.1 14-1.3 15-1 16-1.1.r 18-1.1 18-1.1.1 18-1.1.1.r 19-1.4.r 21-1.4.1.1.1.1.1 22-1.4.1 22-1.4.1.1 22-1.4.1.1.r 24-1.4 (s / support-01~e.15 :ARG0~e.16 (t2 / thing~e.18 :ARG2-of~e.18 (r2 / result-01~e.18)) :ARG1 (r / role~e.1 :poss~e.2 (p / phosphorylate-01~e.7,12 :ARG1 (e2 / enzyme :name (n2 / name :op1 "ERK1/2"~e.4,6)) :ARG1-of (s2 / stimulate-01~e.3)) :topic (p2 / phosphorylate-01~e.7 :ARG1~e.8 (p3 / protein :name (n3 / name :op1 "YB-1"~e.9,11)))) :degree (f / further~e.14) :time~e.19 (u / use-01~e.24 :ARG1 (m / molecular-physical-entity~e.22 :ARG0-of~e.22 (i / inhibit-01~e.22 :ARG1 (p4 / protein-family :name (n / name :op1 "MEK"~e.21)))))) # ::id a_pmid_2139_2397.141 ::amr-annotator SDL-AMR-09 ::preferred # ::tok As shown in Figure 2B , pretreatment of MDA @-@ MB @-@ 231 cells with the MEK inhibitor PD98059 markedly blocked YB @-@ 1 phosphorylation . # ::alignments 1-1.4 2-1.4.1.r 3-1.4.1 5-1.4.1.1 8-1.1 9-1.1.1.r 10-1.1.1.1.1 12-1.1.1.1.1 14-1.1.1.1.1 15-1.1.1 16-1.1.2.r 18-1.1.2.2.1.1.1 19-1.1.2 19-1.1.2.2 19-1.1.2.2.r 20-1.1.2.1.1 21-1.3 22-1 23-1.2.1.1.1 25-1.2.1.1.1 26-1.2 (b / block-01~e.22 :ARG0 (p2 / pretreat-01~e.8 :ARG1~e.9 (c / cell-line~e.15 :name (n2 / name :op1 "MDA-MB-231"~e.10,12,14)) :ARG3~e.16 (s2 / small-molecule~e.19 :name (n3 / name :op1 "PD98059"~e.20) :ARG0-of~e.19 (i2 / inhibit-01~e.19 :ARG1 (p4 / protein-family :name (n / name :op1 "MEK"~e.18))))) :ARG1 (p / phosphorylate-01~e.26 :ARG1 (p3 / protein :name (n4 / name :op1 "YB-1"~e.23,25))) :degree (m2 / marked~e.21) :ARG1-of (s / show-01~e.1 :ARG0~e.2 (f / figure~e.3 :mod "2B"~e.5))) # ::id a_pmid_2139_2397.142 ::amr-annotator SDL-AMR-09 ::preferred # ::tok Similar to the data shown in Figure 1D , exposure to IR did not induce YB @-@ 1 phosphorylation . # ::alignments 0-1.4 1-1.4.1.r 3-1.4.1 4-1.5 5-1.5.1.r 6-1.5.1 8-1.5.1.1 11-1.2 12-1.2.1.r 13-1.2.1 13-1.2.1.1 13-1.2.1.1.r 15-1.1 15-1.1.r 16-1 17-1.3.1.1.1 19-1.3.1.1.1 20-1.3 (i2 / induce-01~e.16 :polarity~e.15 -~e.15 :ARG0 (e / expose-01~e.11 :ARG2~e.12 (r2 / radiate-01~e.13 :ARG0-of~e.13 (i / ionize-01~e.13))) :ARG2 (p / phosphorylate-01~e.20 :ARG1 (p2 / protein :name (n2 / name :op1 "YB-1"~e.17,19))) :ARG1-of (r / resemble-01~e.0 :ARG2~e.1 (d / data~e.3)) :ARG1-of (s / show-01~e.4 :ARG0~e.5 (f / figure~e.6 :mod "1D"~e.8))) # ::id a_pmid_2139_2397.143 ::amr-annotator SDL-AMR-09 ::preferred # ::tok These results indicates that the constitutive YB @-@ 1 phosphorylation in MDA @-@ MB @-@ 231 cells is a consequence of mutated K @-@ Ras @-@ mediated ERK1 @/@ 2 phosphorylation . # ::alignments 0-1.1.2 1-1.1 1-1.1.1 1-1.1.1.r 2-1 3-1.2.r 5-1.2.1.2 6-1.2.1.1.2.1 8-1.2.1.1.2.1 9-1.2.1 10-1.2.1.3.r 11-1.2.1.3.2.1 13-1.2.1.3.2.1 15-1.2.1.3.2.1 16-1.2.1.3 17-1.2.1.r 19-1.2 19-1.2.2 19-1.2.2.r 20-1.2.2.1.r 21-1.2.2.1.2.1.3 22-1.2.2.1.2.1.2.1 24-1.2.2.1.2.1.2.1 26-1.2.2.1.2 27-1.2.2.1.1.2.1 29-1.2.2.1.1.2.1 30-1.2.2.1 (i / indicate-01~e.2 :ARG0 (t / thing~e.1 :ARG2-of~e.1 (r / result-01~e.1) :mod (t2 / this~e.0)) :ARG1~e.3 (c / consequence~e.19 :domain~e.17 (p / phosphorylate-01~e.9 :ARG1 (p2 / protein :wiki "Y_box_binding_protein_1" :name (n / name :op1 "YB-1"~e.6,8)) :mod (c2 / constitutive~e.5) :location~e.10 (c3 / cell-line~e.16 :wiki - :name (n2 / name :op1 "MDA-MB-231"~e.11,13,15))) :ARG1-of~e.19 (c4 / cause-01~e.19 :ARG0~e.20 (p3 / phosphorylate-01~e.30 :ARG1 (e / enzyme :wiki "Extracellular_signal-regulated_kinases" :name (n3 / name :op1 "ERK1/2"~e.27,29)) :ARG1-of (m / mediate-01~e.26 :ARG0 (e2 / enzyme :wiki "KRAS" :name (n4 / name :op1 "K-Ras"~e.22,24) :ARG2-of (m2 / mutate-01~e.21))))))) # ::id a_pmid_2139_2397.144 ::amr-annotator SDL-AMR-09 ::preferred # ::tok Overexpression of mutated K @-@ RAS @ V12 @ enhances basal YB @-@ 1 phosphorylation # ::alignments 0-1.1 1-1.1.1.r 2-1.1.1.2 4-1.1.1.1.1 6-1.1.1.1.1 9-1.1.1.2.1 11-1 12-1.2.2 13-1.2.1.1.1 15-1.2.1.1.1 16-1.2 (e2 / enhance-01~e.11 :ARG0 (o / overexpress-01~e.0 :ARG1~e.1 (g / gene :name (n / name :op1 "K-RAS"~e.4,6) :ARG2-of (m / mutate-01~e.2 :value "V12"~e.9))) :ARG1 (p / phosphorylate-01~e.16 :ARG1 (p2 / protein :name (n2 / name :op1 "YB-1"~e.13,15)) :mod (b / basal~e.12))) # ::id a_pmid_2139_2397.145 ::amr-annotator SDL-AMR-09 ::preferred # ::tok To investigate the role of K @-@ Ras in the constitutive phosphorylation of YB @-@ 1 , we further analyzed the status of K @-@ RAS @ in SKBr3 , MCF @-@ 7 and HBL100 cells . # ::alignments 1-1.4 3-1.4.2 4-1.4.2.2.r 5-1.4.2.2.1.1 7-1.4.2.2.1.1 8-1.4.2.1.r 10-1.4.2.1.2 11-1.4.2.1 12-1.4.2.1.1.r 13-1.4.2.1.1.1.1 15-1.4.2.1.1.1.1 17-1.1 18-1.3 19-1 21-1.2 24-1.2.2.1.1 26-1.2.2.1.1 28-1.2.1.r 29-1.2.1.1.1.1 31-1.2.1.2.1.1 33-1.2.1.2.1.1 34-1.2.1 35-1.2.1.3.1.1 36-1.2.1.1 36-1.2.1.2 36-1.2.1.3 (a / analyze-01~e.19 :ARG0 (w / we~e.17) :ARG1 (s / status~e.21 :location~e.28 (a2 / and~e.34 :op1 (c / cell-line~e.36 :name (n2 / name :op1 "SKBr3"~e.29)) :op2 (c2 / cell-line~e.36 :name (n3 / name :op1 "MCF-7"~e.31,33)) :op3 (c3 / cell-line~e.36 :name (n4 / name :op1 "HBL100"~e.35))) :poss (g / gene :name (n6 / name :op1 "K-RAS"~e.24,26))) :degree (f / further~e.18) :purpose (i / investigate-01~e.1 :ARG0 w :ARG1 (r / role~e.3 :topic~e.8 (p / phosphorylate-01~e.11 :ARG1~e.12 (p2 / protein :name (n5 / name :op1 "YB-1"~e.13,15)) :mod (c4 / constitutive~e.10)) :poss~e.4 (e / enzyme :name (n / name :op1 "K-Ras"~e.5,7))))) # ::id a_pmid_2139_2397.146 ::amr-annotator SDL-AMR-09 ::preferred # ::tok Sequencing of the K @-@ RAS @ gene revealed that none of these cell lines presents a K @-@ RAS @ point mutation in codon 12 , codon 13 or 61 . # ::alignments 0-1.1 4-1.1.1.1.1 6-1.1.1.1.1 8-1.1.1 9-1 10-1.2.r 11-1.2.1.1 12-1.2.1.1.r 13-1.2.1.2 14-1.2.1 15-1.2.1 16-1.2 19-1.2.2.1 20-1.2.2.1 21-1.2.2.1 23-1.2.2.2 24-1.2.2 25-1.2.2.3.r 26-1.2.2.3.1 27-1.2.2.3.1.1 29-1.2.2.3.2 29-1.2.2.3.3 30-1.2.2.3.2.1 31-1.2.2.3 32-1.2.2.3.3.1 (r / reveal-01~e.9 :ARG0 (s / sequence-01~e.0 :ARG1 (g / gene~e.8 :name (n2 / name :op1 "K-RAS"~e.4,6))) :ARG1~e.10 (p / present-01~e.16 :ARG0 (c / cell-line~e.14,15 :quant~e.12 (n3 / none~e.11) :mod (t / this~e.13)) :ARG1 (m / mutate-01~e.24 :ARG1 g~e.19,20,21 :mod (p2 / point~e.23) :location~e.25 (o / or~e.31 :op1 (c2 / codon~e.26 :mod 12~e.27) :op2 (c3 / codon~e.29 :mod 13~e.30) :op3 (c4 / codon~e.29 :mod 61~e.32))))) # ::id a_pmid_2139_2397.147 ::amr-annotator SDL-AMR-09 ::preferred # ::tok To investigate whether mutated K @-@ RAS @ V12 @ could upregulate YB @-@ 1 phosphorylation , we introduced mutated K @-@ RAS @ into K @-@ RAS @ wt , SKBr3 and MCF @-@ 7 cells . # ::alignments 1-1.4 2-1.4.2.1 2-1.4.2.1.r 3-1.4.2.2.2.2 5-1.4.2.2.2.1.1 7-1.4.2.2.2.1.1 10-1.4.2.2.2.2.1 12-1.4.2 13-1.4.2.2 14-1.4.2.2.1.1.1.1 16-1.4.2.2.1.1.1.1 17-1.4.2.2.1 19-1.1 20-1 21-1.2 23-1.3.3.1.1 23-1.4.2.2.2.1.1 25-1.3.3.1.1 25-1.4.2.2.2.1.1 29-1.3.3.1.1 29-1.4.2.2.2.1.1 31-1.3.3.1.1 31-1.4.2.2.2.1.1 34-1.3.3.2 37-1.3.1.1.1 38-1.3 39-1.3.2.1.1 41-1.3.2.1.1 42-1.3.1 42-1.3.2 (i / introduce-02~e.20 :ARG0 (w / we~e.19) :ARG1 g~e.21 :ARG2 (a / and~e.38 :op1 (c / cell-line~e.42 :name (n4 / name :op1 "SKBr3"~e.37)) :op2 (c2 / cell-line~e.42 :name (n5 / name :op1 "MCF-7"~e.39,41)) :mod (g2 / gene :name (n3 / name :op1 "K-RAS"~e.23,25,29,31) :mod (w2 / wild-type~e.34))) :purpose (i2 / investigate-01~e.1 :ARG0 w :ARG1 (p3 / possible-01~e.12 :mode~e.2 interrogative~e.2 :ARG1 (u / upregulate-01~e.13 :ARG1 (p2 / phosphorylate-01~e.17 :ARG1 (p / protein :name (n2 / name :op1 "YB-1"~e.14,16))) :ARG2 (g / gene :name (n / name :op1 "K-RAS"~e.5,7,23,25,29,31) :ARG2-of (m / mutate-01~e.3 :value "V12"~e.10)))))) # ::id a_pmid_2139_2397.148 ::amr-annotator SDL-AMR-09 ::preferred # ::tok Cells were transiently transfected with either a control pEGFP @-@ C1 vector ( indicated as con . @-@ vector ) or a vector expressing mutated K @-@ RAS , pEGFP @-@ C1/ @ K @-@ RAS @ V12 @ ( indicated as K @-@ RAS @ V12 ) . # ::alignments 0-1.1 2-1.3 3-1 7-1.2.1.2 8-1.2.1.1.1 8-1.2.2.1.1 10-1.2.1.1.1 10-1.2.2.1.1 11-1.2.1 11-1.2.1.3.1 11-1.2.1.3.1.1.1 11-1.2.2.3.1 13-1.2.1.3 13-1.2.2.3 18-1.2.1.3.1.1.1 18-1.2.2 20-1.2 22-1.2.1.3.1.1.1 22-1.2.2 23-1.2.2.2 24-1.2.2.2.1.2 26-1.2.2.2.1.1.1 28-1.2.2.2.1.1.1 31-1.2.2.1.1 35-1.2.2.1.1 35-1.2.2.2.1.1.1 35-1.2.2.3.1.1.1 37-1.2.2.2.1.1.1 43-1.2.1.3 46-1.2.2.1.1 46-1.2.2.2.1.1.1 46-1.2.2.3.1.1.1 48-1.2.2.2.1.1.1 (t / transfect-01~e.3 :ARG1 (c / cell~e.0) :ARG2 (o / or~e.20 :op1 (v / vector~e.11 :name (n2 / name :op1 "pEGFP-C1"~e.8,10) :mod (c2 / control~e.7) :ARG1-of (i / indicate-01~e.13,43 :ARG0 (v4 / vector~e.11 :name (n5 / name :op1 "con.-vector"~e.11,18,22)))) :op2 (v2 / vector~e.18,22 :name (n4 / name :op1 "pEGFP-C1/K-RASV12"~e.8,10,31,35,46) :ARG3-of (e2 / express-03~e.23 :ARG1 (g / gene :name (n / name :op1 "K-RAS"~e.26,28,35,37,46,48) :ARG2-of (m / mutate-01~e.24))) :ARG1-of (i2 / indicate-01~e.13 :ARG0 (v3 / vector~e.11 :name (n3 / name :op1 "K-RASV12"~e.35,46))))) :ARG1-of (t2 / transient-02~e.2)) # ::id a_pmid_2139_2397.149 ::amr-annotator SDL-AMR-09 ::preferred # ::tok Fluorescence images of living cells transfected with con . @-@ vector and K @-@ RAS @ V12 @ revealed that GFP in K @-@ RAS @ V12 @ vector @-@ transfected cells was localized to the plasma membrane , but that in con . @-@ vector @-@ transfected cells it was not ( Figure 3A ) . # ::alignments 0-1.3 1-1.1 3-1.1.1.1.1 4-1.1.1.1 4-1.1.1.2 4-1.2.1.3 5-1.1.1.2.2 10-1.1.1.2.2.1 13-1.1.1.2.2.1.1.1 20-1 21-1.2.r 22-1.2.1.1.1.1 25-1.1.1.2.2.1.1.1 32-1.2.2.3.1 33-1.2.2.3.1 34-1.2.2.3.1 35-1.2.2.3 37-1.2.1 37-1.2.2 38-1.2.1.2.r 40-1.2.1.2.1 41-1.2.1.2 43-1.2 49-1.2.2.3.1 50-1.2.2.3.1 51-1.2.2.3.1 52-1.2.2.3 55-1.2.2.1 55-1.2.2.1.r 57-1.4.1 59-1.4.1.1 (r / reveal-01~e.20 :ARG0 (i / image-101~e.1 :ARG1 (a / and :op1 (c / cell~e.4 :ARG0-of (l / live-01~e.3) :ARG1-of (t / transfect-01 :ARG2 (v / vector :name (n2 / name :op1 "con.-vector")))) :op2 (c3 / cell~e.4 :ARG0-of l :ARG1-of (t2 / transfect-01~e.5 :ARG2 (v2 / vector~e.10 :name (n / name :op1 "K-RASV12"~e.13,25)))))) :ARG1~e.21 (c2 / contrast-01~e.43 :ARG1 (l2 / localize-01~e.37 :ARG1 (p / protein :name (n3 / name :op1 "GFP"~e.22)) :location~e.38 (m / membrane~e.41 :mod (p2 / plasma~e.40)) :location (c4 / cell~e.4 :ARG2-of t2)) :ARG2 (l3 / localize-01~e.37 :polarity~e.55 -~e.55 :ARG1 p :location (c5 / cell~e.35,52 :ARG2-of t~e.32,33,34,49,50,51) :location m)) :mod (f / fluorescence~e.0) :ARG1-of (d / describe-01 :ARG0 (f2 / figure~e.57 :mod "3A"~e.59))) # ::id a_pmid_2139_2397.150 ::amr-annotator SDL-AMR-09 ::preferred # ::tok This is due to posttranslational modification and membrane association of K @-@ Ras ( Figure 3A ) . # ::alignments 0-1.2 2-1 3-1 4-1.1.1.1.1.1.r 4-1.1.1.2 4-1.1.1.2.1 4-1.1.1.2.1.r 4-1.1.1.r 5-1.1.1 6-1.1 7-1.1.2.2 8-1.1.2 9-1.1.1.1.r 10-1.1.1.1.1.1 12-1.1.1.1.1.1 14-1.3.1 16-1.3.1.1 (c / cause-01~e.2,3 :ARG0 (a / and~e.6 :op1~e.4 (m / modify-01~e.5 :ARG1~e.9 (e / enzyme :name (n / name :op1~e.4 "K-Ras"~e.10,12)) :time (a2 / after~e.4 :op1~e.4 (t2 / translate-02~e.4 :ARG1 e))) :op2 (a3 / associate-01~e.8 :ARG1 e :ARG2 (m2 / membrane~e.7))) :ARG1 (t / this~e.0) :ARG1-of (d / describe-01 :ARG0 (f / figure~e.14 :mod "3A"~e.16))) # ::id a_pmid_2139_2397.151 ::amr-annotator SDL-AMR-09 ::preferred # ::tok In con . @-@ vector @-@ transfected cells , GFP expression was not accumulated at the cell membrane , but rather it was equally distributed throughout the cytoplasm . # ::alignments 4-1.2.3.2.1.1 4-1.2.3.2.1.1.1.1 6-1.2.3.2.1 7-1.2.3.2 9-1.2.3.1.1.1 10-1.2.3 12-1.2.1 12-1.2.1.r 13-1.2 14-1.2.2.r 16-1.2.2.1 17-1.2.2 20-1 20-1.1.3 21-1.1.1 23-1.1.4 24-1.1 27-1.1.2 (i / instead-of-91~e.20 :ARG1 (d / distribute-01~e.24 :ARG1 e~e.21 :location (c / cytoplasm~e.27) :mod (r / rather~e.20) :ARG1-of (e2 / equal-01~e.23)) :ARG2 (a / accumulate-01~e.13 :polarity~e.12 -~e.12 :ARG0~e.14 (m / membrane~e.17 :part-of c2~e.16) :ARG1 (e / express-03~e.10 :ARG2 (p / protein :name (n / name :op1 "GFP"~e.9)) :ARG3 (c2 / cell~e.7 :ARG1-of (t / transfect-01~e.6 :ARG2 (v / vector~e.4 :name (n2 / name :op1 "con.-vector"~e.4))))))) # ::id a_pmid_2139_2397.152 ::amr-annotator SDL-AMR-09 ::preferred # ::tok The efficiency of transfection was verified by immunoblotting as well ( Figure 3B ) . # ::alignments 1-1.1 2-1.1.1.r 3-1.1.1 5-1 6-1.4.r 7-1.4 8-1.2 9-1.2 11-1.3.1 13-1.3.1.1 (v / verify-01~e.5 :ARG1 (e / efficient-01~e.1 :ARG1~e.2 (t / transfect-01~e.3)) :mod (a / as-well~e.8,9) :ARG1-of (d / describe-01 :ARG0 (f / figure~e.11 :mod "3B"~e.13)) :manner~e.6 (i / immunoblot-01~e.7)) # ::id a_pmid_2139_2397.153 ::amr-annotator SDL-AMR-09 ::preferred # ::tok In cells transfected with K @-@ RAS @ V12 @ vector , the expression of K @-@ Ras ( 21 kDa ) resulted in a shift of GFP from 27 kDa to 48 kDa ( Figure 3B ) . # ::alignments 1-1.1.2 2-1.1.2.1 5-1.1.2.1.1.1.1 12-1.1.2.1.1 15-1.1 17-1.1.1.1.1 17-1.1.2.1.1.1.1 19-1.1.1.1.1 21-1.1.1.2.1 22-1.1.1.2.2 24-1 25-1.2.r 27-1.2 28-1.2.1.r 29-1.2.1.1.1 30-1.2.3.r 31-1.2.3.1 32-1.1.1.2.2 33-1.2.2.r 34-1.2.2.1 35-1.2.2.2 37-1.3.1 39-1.3.1.1 (r / result-01~e.24 :ARG1 (e2 / express-03~e.15 :ARG2 (e / enzyme :name (n / name :op1 "K-Ras"~e.17,19) :quant (m2 / mass-quantity :quant 21~e.21 :unit (k / kilodalton~e.22,32))) :ARG3 (c / cell~e.1 :ARG1-of (t / transfect-01~e.2 :ARG2 (v / vector~e.12 :name (n2 / name :op1 "K-RASV12"~e.5,17))))) :ARG2~e.25 (s / shift-01~e.27 :ARG1~e.28 (p / protein :name (n3 / name :op1 "GFP"~e.29)) :ARG2~e.33 (m4 / mass-quantity :quant 48~e.34 :unit k~e.35) :ARG3~e.30 (m3 / mass-quantity :quant 27~e.31 :unit k)) :ARG1-of (d / describe-01 :ARG0 (f / figure~e.37 :mod "3B"~e.39))) # ::id a_pmid_2139_2397.154 ::amr-annotator SDL-AMR-09 ::preferred # ::tok The expression of GFP @-@ tagged K @-@ Ras with a molecular weight of 48 kDa was further confirmed by stripping the anti @-@ GFP antibody from the membrane and reincubating the blots with a K @-@ Ras antibody . # ::alignments 1-1.1 2-1.1.1.r 3-1.1.1.2.1.1.1 5-1.1.1.2 6-1.1.1.1.1 8-1.1.1.1.1 11-1.1.1.3.1 12-1.1.1.3 13-1.1.1.3.2.r 14-1.1.1.3.2.1 15-1.1.1.3.2.2 17-1.2 18-1 19-1.3.r 20-1.3.1 22-1.3.1.1.1 22-1.3.2.2.1 24-1.1.1.2.1.1.1 25-1.3.1.1 25-1.3.2.2 26-1.3.1.2.r 28-1.3.1.2 29-1.3 32-1.3.2.1 35-1.1.1.1.1 37-1.1.1.1.1 38-1.3.1.1 (c / confirm-01~e.18 :ARG1 (e2 / express-03~e.1 :ARG2~e.2 (e / enzyme :name (n / name :op1 "K-Ras"~e.6,8,35,37) :ARG1-of (t / tag-01~e.5 :ARG2 (p / protein :name (n2 / name :op1 "GFP"~e.3,24))) :mod (w / weight~e.12 :mod (m / molecule~e.11) :quant~e.13 (m2 / mass-quantity :quant 48~e.14 :unit (k / kilodalton~e.15))))) :degree (f / further~e.17) :manner~e.19 (a2 / and~e.29 :op1 (s / strip-01~e.20 :ARG1 (a3 / antibody~e.25,38 :ARG0-of (c2 / counter-01~e.22 :ARG1 p)) :ARG2~e.26 (m3 / membrane~e.28)) :op2 (i / incubate-01 :ARG1 (b / blot~e.32) :ARG2 (a4 / antibody~e.25 :ARG0-of (c3 / counter-01~e.22 :ARG1 e)) :mod (a5 / again)))) # ::id a_pmid_2139_2397.155 ::amr-annotator SDL-AMR-09 ::preferred # ::tok In line with our observations of MDA @-@ MB @-@ 231 cells , exogenous expression of K @-@ RAS @ V12 @ in K @-@ RAS @ wt , SKBr3 and MCF @-@ 7 cells resulted in markedly enhanced basal phosphorylation of YB @-@ 1 at S102 ( Figure 3B ) , which prevents further enhancement of phosphorylation by IR ( Figure 3C ) . # ::alignments 1-1.3.1.2 3-1.3.1.1 3-1.3.1.1.r 4-1.3.1 6-1.3.1.2.1.1 8-1.3.1.2.1.1 10-1.3.1.2.1.1 11-1.3.1.2 13-1.1.3 14-1.1 17-1.1.1.1.1 17-1.1.2.3.1.1 19-1.1.1.1.1 19-1.1.2.3.1.1 22-1.1.1.2.1 26-1.1.1.1.1 26-1.1.2.3.1.1 28-1.1.1.1.1 28-1.1.2.3.1.1 31-1.1.2.3.2 34-1.1.2.1.1.1 35-1.1.2 36-1.1.2.2.1.1 38-1.1.2.2.1.1 39-1.1.2.1 39-1.1.2.2 40-1 41-1.2.r 42-1.2.3.1 42-1.2.3.1.r 43-1.2.3 44-1.2.2 45-1.2 47-1.2.1.3.1.1 49-1.2.1.3.1.1 53-1.4.1 55-1.4.1.1 60-1.2.4 61-1.2.4.1.3 62-1.2.4.1 64-1.2 65-1.2.4.1.2.r 66-1.2.4.1.2 66-1.2.4.1.2.1 66-1.2.4.1.2.1.r 68-1.2.4.2.1 70-1.2.4.2.1.1 (r / result-01~e.40 :ARG1 (e2 / express-03~e.14 :ARG1 (g / gene :name (n / name :op1 "K-RAS"~e.17,19,26,28) :ARG2-of (m / mutate-01 :value "V12"~e.22)) :ARG3 (a / and~e.35 :op1 (c / cell-line~e.39 :name (n3 / name :op1 "SKBr3"~e.34)) :op2 (c2 / cell-line~e.39 :name (n4 / name :op1 "MCF-7"~e.36,38)) :mod (g2 / gene :name (n2 / name :op1 "K-RAS"~e.17,19,26,28) :mod (w / wild-type~e.31))) :mod (e3 / exogenous~e.13)) :ARG2~e.41 (p / phosphorylate-01~e.45,64 :ARG1 (a2 / amino-acid :mod 102 :name (n5 / name :op1 "serine") :part-of (p2 / protein :name (n6 / name :op1 "YB-1"~e.47,49))) :mod (b / basal~e.44) :ARG1-of (e5 / enhance-01~e.43 :manner~e.42 (m2 / marked~e.42)) :ARG0-of (p3 / prevent-01~e.60 :ARG1 (e6 / enhance-01~e.62 :ARG1 p :ARG2~e.65 (r3 / radiate-01~e.66 :ARG0-of~e.66 (i / ionize-01~e.66)) :degree (f2 / further~e.61)) :ARG1-of (d2 / describe-01 :ARG0 (f3 / figure~e.68 :mod "3C"~e.70)))) :ARG1-of (r2 / resemble-01 :ARG2 (o / observe-01~e.4 :ARG0~e.3 (w2 / we~e.3) :ARG1 (c3 / cell-line~e.1,11 :name (n7 / name :op1 "MDA-MB-231"~e.6,8,10)))) :ARG1-of (d / describe-01 :ARG0 (f / figure~e.53 :mod "3B"~e.55))) # ::id a_pmid_2139_2397.156 ::amr-annotator SDL-AMR-09 ::preferred # ::tok Thus , these data support the hypothesis that in cells expressing mutated K @-@ RAS , the basal phosphorylation of YB @-@ 1 is constitutively enhanced and cannot be further stimulated by IR. # ::alignments 2-1.1.1.1 3-1.1.1 4-1.1 6-1.1.2 8-1.1.2.1.r 9-1.1.2.1.3 10-1.1.2.1.3.1 11-1.1.2.1.3.1.1.2 13-1.1.2.1.3.1.1.1.1 15-1.1.2.1.3.1.1.1.1 19-1.1.2.1.1.1.2 20-1.1.2.1.1.1 21-1.1.2.1.1.1.1.r 22-1.1.2.1.1.1.1.1.1 24-1.1.2.1.1.1.1.1.1 26-1.1.2.1.1.2 27-1.1.2.1.1 28-1.1.2.1 29-1.1.2.1.2 29-1.1.2.1.2.1 29-1.1.2.1.2.1.r 31-1.1.2.1.2.2.3 32-1.1.2.1.2.2 (i / infer-01 :ARG1 (s / support-01~e.4 :ARG0 (d / data~e.3 :mod (t2 / this~e.2)) :ARG1 (h2 / hypothesize-01~e.6 :ARG1~e.8 (a / and~e.28 :op1 (e2 / enhance-01~e.27 :ARG1 (p / phosphorylate-01~e.20 :ARG1~e.21 (p3 / protein :name (n2 / name :op1 "YB-1"~e.22,24)) :mod (b / basal~e.19)) :mod (c / constitutive~e.26)) :op2 (p2 / possible-01~e.29 :polarity~e.29 -~e.29 :ARG1 (s2 / stimulate-01~e.32 :ARG1 p :ARG2 (r / radiate-01 :ARG0-of (i2 / ionize-01)) :degree (f / further~e.31))) :location (c2 / cell~e.9 :ARG3-of (e3 / express-03~e.10 :ARG1 (g / gene :name (n / name :op1 "K-RAS"~e.13,15) :ARG2-of (m / mutate-01~e.11)))))))) # ::id a_pmid_2139_2397.157 ::amr-annotator SDL-AMR-09 ::preferred # ::tok IR @-@ induced YB @-@ 1 phosphorylation is mediated by erbB1 @-@ dependent PI3K @/@ Akt and MAPK @/@ ERK pathways # ::alignments 0-1.2.2.1 0-1.2.2.1.1 0-1.2.2.1.1.r 2-1.2.2 3-1.2.1.1.1 5-1.2.1.1.1 6-1.2 8-1 9-1.1.r 10-1.1.3.1.1.1 12-1.1.3 13-1.1.1.1.1 16-1.1 17-1.1.2.1.1 19-1.1.2.1.1 20-1.1.1 20-1.1.2 (m / mediate-01~e.8 :ARG0~e.9 (a / and~e.16 :op1 (p2 / pathway~e.20 :name (n / name :op1 "PI3K/AKT"~e.13)) :op2 (p4 / pathway~e.20 :name (n4 / name :op1 "MAPK/ERK"~e.17,19)) :ARG0-of (d / depend-01~e.12 :ARG1 (p5 / protein :name (n5 / name :op1 "erbB1"~e.10)))) :ARG1 (p / phosphorylate-01~e.6 :ARG1 (p3 / protein :name (n3 / name :op1 "YB-1"~e.3,5)) :ARG2-of (i / induce-01~e.2 :ARG0 (r / radiate-01~e.0 :ARG0-of~e.0 (i2 / ionize-01~e.0))))) # ::id a_pmid_2139_2397.158 ::amr-annotator SDL-AMR-09 ::preferred # ::tok The phosphorylation of YB @-@ 1 at S102 in response to stimulation with EGF has been described as being dependent on p90 ribosomal S6 kinase [ @ 11 @ ] . # ::alignments 1-1.1 2-1.1.1.r 3-1.1.1.3.1.1 5-1.1.1.3.1.1 8-1.1.2.r 9-1.1.2 10-1.1.2.1.r 11-1.1.2.1 12-1.1.2.1.2.r 13-1.1.2.1.2.1.1 16-1 16-1.3 16-1.3.r 19-1.2 20-1.2.2.r 21-1.2.2.1.1 22-1.2.2.1.2 23-1.2.2.1.3 24-1.2.2.1.4 27-1.3.1.1.1 (d / describe-01~e.16 :ARG1 (p / phosphorylate-01~e.1 :ARG1~e.2 (a / amino-acid :mod 102 :name (n2 / name :op1 "serine") :part-of (p2 / protein :name (n / name :op1 "YB-1"~e.3,5))) :ARG2-of~e.8 (r / respond-01~e.9 :ARG1~e.10 (s / stimulate-01~e.11 :ARG1 p2 :ARG2~e.12 (p4 / protein :name (n3 / name :op1 "EGF"~e.13))))) :ARG2 (d2 / depend-01~e.19 :ARG0 p :ARG1~e.20 (e / enzyme :name (n5 / name :op1 "p90"~e.21 :op2 "ribosomal"~e.22 :op3 "S6"~e.23 :op4 "kinase"~e.24))) :ARG1-of~e.16 (d3 / describe-01~e.16 :ARG0 (p3 / publication :ARG1-of (c / cite-01 :ARG2 11~e.27)))) # ::id a_pmid_2139_2397.159 ::amr-annotator SDL-AMR-09 ::preferred # ::tok In that study [ @ 11 @ ] , Stratford et al @ . showed that the stimulation of SUM149 breast cancer cells with serum , EGF and phorbol 12 @-@ myristate 13 @-@ acetate ( PMA ) leads to phosphorylation of YB @-@ 1 at S102 , which is dependent on the MAP kinase pathway [ @ 11 @ ] . # ::alignments 1-1.3.1 2-1.3 5-1.3.2.1.1.1 9-1.1.1.1.1 11-1.1 12-1.1.2.1 15-1 16-1.2.r 16-1.3.1 18-1.2.1 19-1.2.1.1.r 20-1.2.1.1.1.1 21-1.2.1.1.2.2.1 22-1.2.1.1.2.2.2 23-1.2.1.1 24-1.2.1.2.r 25-1.2.1.2.1 27-1.2.1.2.2.1.1 28-1.2.1.2 29-1.2.1.2.3.1.1 30-1.2.1.2.3.1.2 32-1.2.1.2.3.1.2 33-1.2.1.2.3.1.3 35-1.2.1.2.3.1.3 37-1.2.1.2.3.2.1.1.1 39-1.2 40-1.2.2.r 41-1.2.2 42-1.2.2.1.r 43-1.2.2.1.3.1.1 45-1.2.2.1.3.1.1 51-1.2.2.2 52-1.2.2.2.1.r 54-1.2.2.2.1.1.1 55-1.2.2.2.1.1.2 56-1.2.2.2.1 59-1.4 (s / show-01~e.15 :ARG0 (a / and~e.11 :op1 (p2 / person :name (n2 / name :op1 "Stratford"~e.9)) :op2 (p3 / person :mod (o / other~e.12))) :ARG1~e.16 (l / lead-03~e.39 :ARG0 (s2 / stimulate-01~e.18 :ARG1~e.19 (c / cell-line~e.23 :name (n3 / name :op1 "SUM149"~e.20) :source (d4 / disease :wiki "Breast_cancer" :name (n9 / name :op1 "breast"~e.21 :op2 "cancer"~e.22))) :ARG2~e.24 (a2 / and~e.28 :op1 (s3 / serum~e.25) :op2 (p7 / protein :name (n4 / name :op1 "EGF"~e.27)) :op3 (s5 / small-molecule :name (n5 / name :op1 "phorbol"~e.29 :op2 "12-myristate"~e.30,32 :op3 "13-acetate"~e.33,35) :ARG1-of (d2 / describe-01 :ARG2 (s6 / small-molecule :name (n6 / name :op1 "PMA"~e.37)))))) :ARG2~e.40 (p / phosphorylate-01~e.41 :ARG1~e.42 (a3 / amino-acid :mod 102 :name (n7 / name :op1 "serine") :part-of (p4 / protein :name (n / name :op1 "YB-1"~e.43,45))) :ARG0-of (d / depend-01~e.51 :ARG1~e.52 (p5 / pathway~e.56 :name (n8 / name :op1 "MAP"~e.54 :op2 "kinase"~e.55))))) :medium (s7 / study-01~e.2 :mod (t / that~e.1,16) :ARG1-of (d3 / describe-01 :ARG0 (p6 / publication :ARG1-of (c3 / cite-01 :ARG2 11~e.5)))) :ARG1-of d3~e.59) # ::id a_pmid_2139_2397.161 ::amr-annotator SDL-AMR-09 ::preferred # ::tok Because we and others have shown that IR induces activation of erbB1 in a ligand @-@ independent manner [ @ 24 , 25 @ ] , we tested whether the IR @-@ induced YB @-@ 1 phosphorylation shown in Figure 1D could be blocked by erbB1 tyrosine kinase inhibitors . # ::alignments 0-1.3 1-1.3.1.1.1 2-1.3.1.1 3-1.3.1.1.2.1 5-1.3.1 7-1.2.2.1.2.1 7-1.2.2.1.2.1.1 7-1.2.2.1.2.1.1.r 8-1.2.2.1.2 8-1.3.1.2 9-1.3.1.2.2 14-1.3.1.2.3.3 16-1.3.1.2.3 16-1.3.1.2.3.1 16-1.3.1.2.3.1.r 17-1.3.1.2.3.r 20-1.3.1.3.1.1.1.1 24-1.3.1.3.1.2.1.1 28-1.1 29-1 30-1.2.1 30-1.2.1.r 32-1.2.2.1.2.1 32-1.2.2.1.2.1.1 32-1.2.2.1.2.1.1.r 34-1.2.2.1.2 35-1.2.2.1.1.1.1 37-1.2.2.1.1.1.1 38-1.2.2.1 39-1.2.2.1.3 40-1.2.2.1.3.1.r 41-1.2.2.1.3.1 43-1.2.2.1.3.1.1 45-1.2 47-1.2.2 50-1.2.2.2.1.1.1.2 51-1.2.2.2.1.1.1.3 52-1.2.2.2 52-1.2.2.2.1 52-1.2.2.2.1.r (t2 / test-01~e.29 :ARG0 (w / we~e.28) :ARG1 (p2 / possible-01~e.45 :mode~e.30 interrogative~e.30 :ARG1 (b / block-01~e.47 :ARG1 (p / phosphorylate-01~e.38 :ARG1 (p3 / protein :name (n2 / name :op1 "YB-1"~e.35,37)) :ARG2-of (i2 / induce-01~e.8,34 :ARG0 (r / radiate-01~e.7,32 :ARG0-of~e.7,32 (i4 / ionize-01~e.7,32))) :ARG1-of (s / show-01~e.39 :medium~e.40 (f / figure~e.41 :mod "1D"~e.43))) :ARG3 (m3 / molecular-physical-entity~e.52 :ARG0-of~e.52 (i / inhibit-01~e.52 :ARG1 (e / enzyme :name (n4 / name :op1 "erB1" :op2 "tyrosine"~e.50 :op3 "kinase"~e.51)))))) :ARG1-of (c / cause-01~e.0 :ARG0 (s2 / show-01~e.5 :ARG0 (a / and~e.2 :op1 (w2 / we~e.1) :op2 (p5 / person :mod (o / other~e.3))) :ARG1 (i3 / induce-01~e.8 :ARG0 r :ARG2 (a2 / activate-01~e.9 :ARG1 e) :manner~e.17 (d / depend-01~e.16 :polarity~e.16 -~e.16 :ARG0 a2 :ARG1 (l / ligand~e.14))) :ARG1-of (d2 / describe-01 :ARG0 (a3 / and :op1 (p6 / publication :ARG1-of (c2 / cite-01 :ARG2 24~e.20)) :op2 (p7 / publication :ARG1-of (c3 / cite-01 :ARG2 25~e.24))))))) # ::id a_pmid_2139_2397.162 ::amr-annotator SDL-AMR-09 ::preferred # ::tok To test this hypothesis , the effect of the erbB1 @-@ RTK inhibitor erlotinib on YB @-@ 1 phosphorylation was analyzed in whole cell extracts as well as in cytoplasmic and nuclear fractions . # ::alignments 1-1.3 2-1.3.1.2 3-1.3.1 3-1.3.1.1 3-1.3.1.1.r 6-1.1 7-1.1.1.r 9-1.1.1.2.1.1.1 11-1.1.1.2.1.1.1 12-1.1.1 12-1.1.1.2 12-1.1.1.2.r 13-1.1.1.1.1 14-1.1.2.r 15-1.1.2.1.1.1 17-1.1.2.1.1.1 18-1.1.2 20-1 21-1.2.r 22-1.2.1.1.1 23-1.2.1.1 24-1.2.1 25-1.2 26-1.2 27-1.2 29-1.2.2.1 30-1.2 31-1.2.3.1 32-1.2.2 32-1.2.3 (a / analyze-01~e.20 :ARG1 (a2 / affect-01~e.6 :ARG0~e.7 (s / small-molecule~e.12 :name (n / name :op1 "erlotinib"~e.13) :ARG0-of~e.12 (i / inhibit-01~e.12 :ARG1 (p3 / pathway :name (n4 / name :op1 "erbB1-RTK"~e.9,11)))) :ARG1~e.14 (p / phosphorylate-01~e.18 :ARG1 (p2 / protein :name (n2 / name :op1 "YB-1"~e.15,17)))) :location~e.21 (a3 / and~e.25,26,27,30 :op1 (e / extract-01~e.24 :ARG1 (c / cell~e.23 :mod (w / whole~e.22))) :op2 (f / fraction~e.32 :part-of (c2 / cytoplasm~e.29)) :op3 (f2 / fraction~e.32 :part-of (n3 / nucleus~e.31))) :purpose (t3 / test-01~e.1 :ARG1 (t / thing~e.3 :ARG1-of~e.3 (h / hypothesize-01~e.3) :mod (t4 / this~e.2)))) # ::id a_pmid_2139_2397.163 ::amr-annotator SDL-AMR-09 ::preferred # ::tok Pretreatment of SKBr3 cells with erlotinib resulted in complete inhibition of YB @-@ 1 phosphorylation in whole cell extract ( Figure 4A ) as well as in cytoplasmic and nuclear fractions ( Figure 4B ) . # ::alignments 0-1.1 1-1.1.1.r 2-1.1.1.1.1 3-1.1.1 4-1.1.2.r 5-1.1.2.1.1 6-1 7-1.2.r 8-1.2.2 9-1.2 10-1.2.1.r 11-1.2.1.1.1.1 13-1.2.1.1.1.1 14-1.2.1 15-1.2.3.r 16-1.2.3.1.1.1 17-1.2.3.1.1 18-1.2.3.1 20-1.2.3.1.2.1 22-1.2.3.1.2.1.1 25-1.2.3 25-1.2.3.2 25-1.2.3.2.r 26-1.2.3.2 27-1.2.3.2 29-1.2.3.2.1.1 30-1.2.3.2 31-1.2.3.2.2.1 32-1.2.3.2.1 32-1.2.3.2.2 34-1.2.3.2.3.1 36-1.2.3.2.3.1.1 (r / result-01~e.6 :ARG1 (p3 / pretreat-01~e.0 :ARG1~e.1 (c / cell-line~e.3 :name (n / name :op1 "SKBr3"~e.2)) :ARG3~e.4 (s / small-molecule :name (n2 / name :op1 "erlotinib"~e.5))) :ARG2~e.7 (i / inhibit-01~e.9 :ARG1~e.10 (p / phosphorylate-01~e.14 :ARG1 (p2 / protein :name (n3 / name :op1 "YB-1"~e.11,13))) :ARG1-of (c2 / complete-02~e.8) :location~e.15 (a / and~e.25 :op1 (e / extract-01~e.18 :ARG1 (c3 / cell~e.17 :mod (w / whole~e.16)) :ARG1-of (d / describe-01 :ARG0 (f / figure~e.20 :mod "4A"~e.22))) :op2~e.25 (a2 / and~e.25,26,27,30 :op1 (f2 / fraction~e.32 :part-of (c4 / cytoplasm~e.29)) :op2 (f3 / fraction~e.32 :part-of (n4 / nucleus~e.31)) :ARG1-of (d2 / describe-01 :ARG0 (f4 / figure~e.34 :mod "4B"~e.36)))))) # ::id a_pmid_2139_2397.164 ::amr-annotator SDL-AMR-09 ::preferred # ::tok As expected , erlotinib also blocked basal @- and radiation @-@ induced P @-@ Akt and P @-@ ERK1 @/@ 2 in these cells ( Figure 4A ) . # ::alignments 1-1.4 3-1.1.1.1 4-1.6 5-1 6-1.2.1.3 8-1.2.1 9-1.2.2.4.1 11-1.2.2.4 12-1.2.1.4 14-1.2.1.1.1.1 15-1.2.1 16-1.2.1.4 18-1.2.1.2.1.1.1 19-1.2.1.2 21-1.3.r 22-1.3.1 23-1.3 25-1.5.1 27-1.5.1.1 (b / block-01~e.5 :ARG0 (s / small-molecule :name (n / name :op1 "erlotinib"~e.3)) :ARG1 (a / and :op1 (a3 / and~e.8,15 :op1 (e / enzyme :name (n2 / name :op1 "Akt"~e.14)) :op2 (s2 / slash~e.19 :op1 (e2 / enzyme :name (n3 / name :op1 "ERK1"~e.18)) :op2 (e3 / enzyme :name (n4 / name :op1 "ERK2"))) :mod (b2 / basal~e.6) :ARG1-of (p / phosphorylate-01~e.12,16)) :op2 (a4 / and :op1 e :op2 s2 :ARG1-of p :ARG2-of (i / induce-01~e.11 :ARG0 (r / radiate-01~e.9)))) :location~e.21 (c / cell~e.23 :mod (t / this~e.22)) :ARG1-of (e4 / expect-01~e.1) :ARG1-of (d / describe-01 :ARG0 (f / figure~e.25 :mod "4A"~e.27)) :mod (a2 / also~e.4)) # ::id a_pmid_2139_2397.165 ::amr-annotator SDL-AMR-09 ::preferred # ::tok To rule out off @-@ target effects of erlotinib , the efficacy of the highly specific erbB1 @-@ RTK inhibitor BIBX1382BS [ @ 38 @ ] on radiation @-@ induced YB @-@ 1 phosphorylation was tested in cytoplasmic and nuclear fractions . # ::alignments 1-1.3 2-1.3 3-1.3.1.2 5-1.3.1.2 6-1.3.1 7-1.3.1.1.r 8-1.3.1.1.2.1 14-1.1.1.5.1 15-1.1.1.5 16-1.1.1.3.1.2.1 18-1.1.1.3.1.2.1 19-1.1.1 19-1.1.1.3 19-1.1.1.3.r 20-1.1.1.2.1 23-1.1.1.4.1.1.1 27-1.1.2.2.1 29-1.1.2.2 30-1.1.2.1.2.1 32-1.1.2.1.2.1 33-1.1.2 35-1 36-1.2.r 37-1.2.1.1 38-1.2 39-1.2.2.1 40-1.2.1 40-1.2.2 (t2 / test-01~e.35 :ARG1 (e / efficient-01 :ARG1 (s3 / small-molecule~e.19 :wiki - :name (n2 / name :op1 "BIBX1382BS"~e.20) :ARG0-of~e.19 (i / inhibit-01~e.19 :ARG1 (e2 / enzyme :wiki - :name (n5 / name :op1 "erbB1-RTK"~e.16,18))) :ARG1-of (d / describe-01 :ARG0 (p3 / publication :ARG1-of (c / cite-01 :ARG2 38~e.23))) :ARG1-of (s / specific-02~e.15 :ARG1-of (h / high-02~e.14))) :ARG2 (p / phosphorylate-01~e.33 :ARG1 (p2 / protein :wiki "Y_box_binding_protein_1" :name (n / name :op1 "YB-1"~e.30,32)) :ARG2-of (i2 / induce-01~e.29 :ARG0 (r / radiate-01~e.27)))) :location~e.36 (a2 / and~e.38 :op1 (f / fraction~e.40 :part-of (c2 / cytoplasm~e.37)) :op2 (f2 / fraction~e.40 :mod (n3 / nucleus~e.39))) :purpose (r2 / rule-out-02~e.1,2 :ARG1 (a3 / affect-01~e.6 :ARG0~e.7 (s2 / small-molecule :wiki "Erlotinib" :name (n4 / name :op1 "erlotinib"~e.8)) :mod (o / off-target~e.3,5)))) # ::id a_pmid_2139_2397.166 ::amr-annotator SDL-AMR-09 ::preferred # ::tok EGF was included as positive control . # ::alignments 0-1.1.1.1 2-1 3-1.2.r 4-1.2.1 5-1.2 (i / include-01~e.2 :ARG1 (p2 / protein :name (n / name :op1 "EGF"~e.0)) :condition~e.3 (c / control~e.5 :mod (p / positive~e.4))) # ::id a_pmid_2139_2397.167 ::amr-annotator SDL-AMR-09 ::preferred # ::tok As shown at the bottom of Figure 4B , in both cytoplasmic and nuclear protein fractions treatment with BIBX1382BS resulted in a marked reduction of YB @-@ 1 phosphorylation stimulated by IR as well as EGF treatment . # ::alignments 1-1.3 2-1.3.1.r 4-1.3.1 5-1.3.1.1.r 6-1.3.1.1 8-1.3.1.1.1 12-1.4.3 13-1.4.1.1.1 14-1.4 15-1.4.2.1.1 16-1.4.1.1 16-1.4.2.1 17-1.4.1 17-1.4.2 18-1.1 19-1.1.1.r 20-1.1.1.1.1 21-1 22-1.2.r 24-1.2.2 25-1.2 26-1.2.1.r 27-1.2.1.1.1.1 29-1.2.1.1.1.1 30-1.2.1 31-1.2.3 32-1.2.3.1.r 33-1.2.3.1.1 33-1.2.3.1.1.1 33-1.2.3.1.1.1.r 34-1.2.3.1 35-1.2.3.1 36-1.2.3.1 37-1.2.3.1.2.1.1.1 38-1.2.3.1.2 (r / result-01~e.21 :ARG1 (t / treat-04~e.18 :ARG2~e.19 (s2 / small-molecule :name (n / name :op1 "BIBX1382BS"~e.20))) :ARG2~e.22 (r2 / reduce-01~e.25 :ARG1~e.26 (p / phosphorylate-01~e.30 :ARG1 (p2 / protein :name (n2 / name :op1 "YB-1"~e.27,29))) :mod (m2 / marked~e.24) :ARG1-of (s / stimulate-01~e.31 :ARG0~e.32 (a / and~e.34,35,36 :op1 (r3 / radiate-01~e.33 :ARG0-of~e.33 (i / ionize-01~e.33)) :op2 (t2 / treat-04~e.38 :ARG2 (p5 / protein :name (n4 / name :op1 "EGF"~e.37)))))) :ARG1-of (s4 / show-01~e.1 :medium~e.2 (b / bottom~e.4 :mod~e.5 (f / figure~e.6 :mod "4B"~e.8))) :location (a2 / and~e.14 :op1 (f2 / fraction~e.17 :part-of (p3 / protein~e.16 :mod (c / cytoplasm~e.13))) :op2 (f3 / fraction~e.17 :part-of (p4 / protein~e.16 :mod (n5 / nucleus~e.15))) :mod (b2 / both~e.12))) # ::id a_pmid_2139_2397.168 ::amr-annotator SDL-AMR-09 ::preferred # ::tok These data indicate that erbB1 @-@ RTK activity is necessary for radiation @-@ induced YB @-@ 1 phosphorylation , and this is most likely due to activation of the PI3K @/@ Akt and MAPK @/@ ERK pathways . # ::alignments 0-1.1.1.1 1-1.1.1 2-1.1 3-1.1.2.r 4-1.1.2.2.1.1.1 6-1.1.2.2.1.1.1 9-1.1.2 10-1.1.2.1.r 11-1.1.2.1.2.1 13-1.1.2.1.2 14-1.1.2.1.1.1.1 16-1.1.2.1.1.1.1 17-1.1.2.1 20-1.1.1.1 22-1.2.2 23-1.2 24-1.2.1 25-1.2.1 26-1.1.2.2 26-1.2.1.1 27-1.2.1.1.1.r 29-1.2.1.1.1.1.1.1 32-1.2.1.1.1 33-1.2.1.1.1.2.1.1 35-1.2.1.1.1.2.1.1 36-1.1.2.2.1 36-1.2.1.1.1.1 36-1.2.1.1.1.2 (a2 / and :op1 (i / indicate-01~e.2 :ARG0 (d / data~e.1 :mod (t / this~e.0,20)) :ARG1~e.3 (n / need-01~e.9 :ARG0~e.10 (p / phosphorylate-01~e.17 :ARG1 (p3 / protein :name (n3 / name :op1 "YB-1"~e.14,16)) :ARG2-of (i2 / induce-01~e.13 :ARG0 (r / radiate-01~e.11))) :ARG1 (a / activate-01~e.26 :ARG1 (p5 / pathway~e.36 :name (n5 / name :op1 "erbB1-RTK"~e.4,6))))) :op2 (l / likely-01~e.23 :ARG1 (c / cause-01~e.24,25 :ARG0 (a3 / activate-01~e.26 :ARG1~e.27 (a4 / and~e.32 :op1 (p2 / pathway~e.36 :name (n2 / name :op1 "PI3K/AKT"~e.29)) :op2 (p4 / pathway~e.36 :name (n4 / name :op1 "MAPK/ERK"~e.33,35)))) :ARG1 n) :degree (m / most~e.22))) # ::id a_pmid_2139_2397.169 ::amr-annotator SDL-AMR-09 ::preferred # ::tok To test the function of PI3K @/@ Akt and MAPK @/@ ERK pathways in YB @-@ 1 phosphorylation , we further investigated whether the inhibitors of PI3K , Akt and MAPK affect YB @-@ 1 phosphorylation in irradiated cells . # ::alignments 1-1.4 3-1.4.2 4-1.4.2.1.r 5-1.4.2.1.1.1.1 7-1.4.2.1.1.1.1 8-1.4.2.1 9-1.4.2.1.2.1.1 11-1.4.2.1.2.1.1 12-1.4.2.1.1 12-1.4.2.1.2 13-1.4.2.2.r 14-1.4.2.2 15-1.4.2.2 16-1.4.2.2 17-1.4.2.2 19-1.1 20-1.3 21-1 22-1.2.1 22-1.2.1.r 24-1.2.2.1 24-1.2.2.1.1 24-1.2.2.1.1.r 24-1.2.2.2 24-1.2.2.2.1 24-1.2.2.2.1.r 24-1.2.2.3 24-1.2.2.3.1 24-1.2.2.3.1.r 25-1.2.2.1.1.1.r 26-1.2.2.1.1.1.1.1 28-1.2.2.2.1.1.1.1 29-1.2.2 30-1.2.2.3.1.1.1.1 31-1.2 32-1.2.3.1.1.1 34-1.2.3.1.1.1 35-1.2.3 36-1.2.4.r 37-1.2.4.1 38-1.2.4 (i2 / investigate-01~e.21 :ARG0 (w / we~e.19) :ARG1 (a / affect-01~e.31 :mode~e.22 interrogative~e.22 :ARG0 (a4 / and~e.29 :op1 (m / molecular-physical-entity~e.24 :ARG0-of~e.24 (i / inhibit-01~e.24 :ARG1~e.25 (e / enzyme :name (n3 / name :op1 "PI3K"~e.26)))) :op2 (m2 / molecular-physical-entity~e.24 :ARG0-of~e.24 (i4 / inhibit-01~e.24 :ARG1 (e2 / enzyme :name (n4 / name :op1 "Akt"~e.28)))) :op3 (m3 / molecular-physical-entity~e.24 :ARG0-of~e.24 (i5 / inhibit-01~e.24 :ARG1 (p5 / protein-family :name (n5 / name :op1 "MAPK"~e.30))))) :ARG1 (p2 / phosphorylate-01~e.35 :ARG1 (p4 / protein :name (n6 / name :op1 "YB-1"~e.32,34))) :location~e.36 (c / cell~e.38 :ARG1-of (i3 / irradiate-01~e.37))) :degree (f / further~e.20) :purpose (t / test-01~e.1 :ARG0 w :ARG1 (f2 / function-01~e.3 :ARG0~e.4 (a3 / and~e.8 :op1 (p / pathway~e.12 :name (n / name :op1 "PI3K/Akt"~e.5,7)) :op2 (p3 / pathway~e.12 :name (n2 / name :op1 "MAPK/ERK"~e.9,11))) :ARG1~e.13 p2~e.14,15,16,17))) # ::id a_pmid_2139_2397.170 ::amr-annotator SDL-AMR-09 ::preferred # ::tok The data shown in Figures 4C and 4D indicate that treatment with either of the inhibitors markedly reduced the phosphorylation of YB @-@ 1 at S102 . # ::alignments 1-1.1 2-1.1.1 3-1.1.1.1.r 4-1.1.1.1.1 4-1.1.1.1.2 6-1.1.1.1.1.1 8-1.1.1.1 10-1.1.1.1.2.1 12-1 13-1.2.r 14-1.2.1 15-1.2.1.1.r 16-1.2.1.1.1 19-1.2.1.1 19-1.2.1.1.2 19-1.2.1.1.2.r 20-1.2.3 20-1.2.3.r 21-1.2 23-1.2.2 24-1.2.2.1.r 25-1.2.2.1.3.1.1 27-1.2.2.1.3.1.1 (i2 / indicate-01~e.12 :ARG0 (d / data~e.1 :ARG1-of (s2 / show-01~e.2 :medium~e.3 (a / and~e.8 :op1 (f / figure~e.4 :mod "4C"~e.6) :op2 (f2 / figure~e.4 :mod "4D"~e.10)))) :ARG1~e.13 (r / reduce-01~e.21 :ARG0 (t / treat-04~e.14 :ARG2~e.15 (m2 / molecular-physical-entity~e.19 :mod (e / either~e.16) :ARG0-of~e.19 (i / inhibit-01~e.19))) :ARG1 (p / phosphorylate-01~e.23 :ARG1~e.24 (a2 / amino-acid :mod 102 :name (n / name :op1 "serine") :part-of (p2 / protein :name (n2 / name :op1 "YB-1"~e.25,27)))) :manner~e.20 (m / marked~e.20))) # ::id a_pmid_2139_2397.171 ::amr-annotator SDL-AMR-09 ::preferred # ::tok However , optimal inhibition was observed when cells were treated with a combination of PI3K and MEK inhibitors . # ::alignments 0-1 2-1.1.1.1 3-1.1.1 3-1.1.2.2.1 3-1.1.2.2.1.1 3-1.1.2.2.1.1.r 5-1.1 6-1.1.2.r 7-1.1.2.1 9-1.1.2 10-1.1.2.2.r 12-1.1.2.2 13-1.1.2.2.1.1.1.r 14-1.1.2.2.1.1.1.1.1 16-1.1.2.2.2.1.1.1.1 17-1.1.1 17-1.1.2.2.2 17-1.1.2.2.2.1 17-1.1.2.2.2.1.r (h / have-concession-91~e.0 :ARG1 (o / observe-01~e.5 :ARG1 (i / inhibit-01~e.3,17 :mod (o2 / optimal~e.2)) :time~e.6 (t2 / treat-04~e.9 :ARG1 (c / cell~e.7) :ARG2~e.10 (c2 / combine-01~e.12 :ARG1 (m / molecular-physical-entity~e.3 :ARG0-of~e.3 (i2 / inhibit-01~e.3 :ARG1~e.13 (p / protein-family :name (n2 / name :op1 "PI3K"~e.14)))) :ARG2 (m2 / molecular-physical-entity~e.17 :ARG0-of~e.17 (i3 / inhibit-01~e.17 :ARG1 (p2 / protein-family :name (n / name :op1 "MEK"~e.16)))))))) # ::id a_pmid_2139_2397.172 ::amr-annotator SDL-AMR-09 ::preferred # ::tok Constitutive YB @-@ 1 phosphorylation due to K @-@ RAS @ mutation depends on erbB1 and downstream PI3K @/@ Akt and MAPK @/@ ERK pathways # ::alignments 0-1.1.2 1-1.1.1.1.1 3-1.1.1.1.1 4-1.1 5-1.1.3 6-1.1.3 8-1.1.3.1.1.1.1 10-1.1.3.1.1.1.1 12-1.1.3.1 13-1 14-1.2.r 15-1.2.1.1.1 16-1.2 17-1.2.2.2 18-1.2.2.1.1 21-1.2 22-1.2.3.1.1 24-1.2.3.1.1 25-1.2.2 25-1.2.3 (d / depend-01~e.13 :ARG0 (p / phosphorylate-01~e.4 :ARG1 (p3 / protein :name (n3 / name :op1 "YB-1"~e.1,3)) :mod (c / constitutive~e.0) :ARG1-of (c2 / cause-01~e.5,6 :ARG0 (m / mutate-01~e.12 :ARG1 (g / gene :name (n4 / name :op1 "K-RAS"~e.8,10))))) :ARG1~e.14 (a / and~e.16,21 :op1 (p4 / protein :name (n5 / name :op1 "erbB1"~e.15)) :op2 (p2 / pathway~e.25 :name (n2 / name :op1 "PI3K/AKT"~e.18) :location (d2 / downstream~e.17)) :op3 (p5 / pathway~e.25 :name (n6 / name :op1 "MAPK/ERK"~e.22,24) :location d2))) # ::id a_pmid_2139_2397.173 ::amr-annotator SDL-AMR-09 ::preferred # ::tok As IR @-@ induced YB @-@ 1 phosphorylation was shown to be dependent on erbB1 , PI3K @/@ Akt and MAPK/ERK , we further investigated whether K @-@ RAS @ mt @ -@ dependent constitutive phosphorylation of YB @-@ 1 might be sensitive to the inhibition of erbB1 , PI3K and MEK . # ::alignments 1-1.4.1.1.1.2.1 1-1.4.1.1.1.2.1.1 1-1.4.1.1.1.2.1.1.r 3-1.4.1.1.1.2 4-1.4.1.1.1.1 5-1.4.1.1.1.1 6-1.4.1.1.1.1 7-1.4.1.1.1 9-1.4.1 12-1.4.1.1 14-1.2.2.2.1.1.1.1 16-1.2.2.2.1.2.1.1 18-1.4.1.1.2.2.1.1 19-1.4.1.1.2 20-1.4.1.1.2.3.1.1 22-1.1 23-1.3 24-1 25-1.2.1 25-1.2.1.r 27-1.2.2.1.3.1.1.1 29-1.2.2.1.3.1.1.1 35-1.2.2.1.3 36-1.2.2.1.2 37-1.2.2.1 38-1.2.2.1.1.r 39-1.2.2.1.1.1.1 41-1.2.2.1.1.1.1 42-1.2 44-1.2.2 47-1.2.2.2 49-1.2.2.2.1.1.1.1 51-1.2.2.2.1.2.1.1 52-1.2.2.2.1 53-1.2.2.2.1.3.1.1 (i / investigate-01~e.24 :ARG0 (w / we~e.22) :ARG1 (p3 / possible-01~e.42 :mode~e.25 interrogative~e.25 :ARG1 (s / sensitive-03~e.44 :ARG0 (p / phosphorylate-01~e.37 :ARG1~e.38 (p4 / protein :name (n4 / name :op1 "YB-1"~e.39,41)) :mod (c / constitutive~e.36) :ARG0-of (d / depend-01~e.35 :ARG1 (g / gene :name (n2 / name :op1 "K-RAS"~e.27,29) :ARG1-of (m / mutate-01)))) :ARG1 (i2 / inhibit-01~e.47 :ARG1 (a / and~e.52 :op1 (p5 / protein :name (n5 / name :op1 "erbB1"~e.14,49)) :op2 (e / enzyme :name (n6 / name :op1 "PI3K"~e.16,51)) :op3 (e2 / enzyme :name (n3 / name :op1 "MEK"~e.53)))))) :degree (f / further~e.23) :ARG1-of (c2 / cause-01 :ARG0 (s2 / show-01~e.9 :ARG1 (d2 / depend-01~e.12 :ARG0 (p6 / phosphorylate-01~e.7 :ARG1 p4~e.4,5,6 :ARG2-of (i3 / induce-01~e.3 :ARG0 (r / radiate-01~e.1 :ARG0-of~e.1 (i4 / ionize-01~e.1)))) :ARG1 (a2 / and~e.19 :op1 p5 :op2 (p2 / pathway :name (n / name :op1 "PI3K/Akt"~e.18)) :op3 (p7 / pathway :name (n8 / name :op1 "MAPK/ERK"~e.20))))))) # ::id a_pmid_2139_2397.174 ::amr-annotator SDL-AMR-09 ::preferred # ::tok To this end , K @-@ RAS @ wt @ MCF @-@ 7 cells were transiently transfected with con . @-@ vector or K @-@ RAS @ V12 @ vector , and 48 hours after transfection the cells were treated with the erbB1 inhibitor erlotinib , the PI3K inhibitor LY294002 or the MEK inhibitor PD98059 for 2 hours . # ::alignments 1-1.3 5-1.2.3.1 6-1.2.3.1 7-1.2.3.1 8-1.2.3.1 9-1.2.3.1 10-1.2.3.1 11-1.2.3.1 12-1.2.3.1 13-1.2.3.1 14-1.2.3.1 15-1.2.3.1 16-1.2.3.1 17-1.2.3.1 18-1.2.3.1 19-1.2.3.1 20-1.2.3.1 21-1.2.3.1 22-1.2.3.1 23-1.2.3.1 24-1.2.3.1 25-1.2.3.1 26-1.2.3.1 27-1.2.3.1 28-1.2.3.1 29-1.2.3.1 30-1.2.3.1 31-1.2.3.1 32-1.2.3.1 33-1.2.3.1 35-1 36-1.2.3.2.1 37-1.2.3.2.2 38-1.2.3 39-1.1 41-1.1.1 43-1.2 44-1.2.2.r 46-1.2.2.1.2.1.1.1 47-1.2.2.1 47-1.2.2.1.2 47-1.2.2.1.2.r 48-1.2.2.1.1.1 51-1.2.2.2.2.1.1.1 52-1.2.2.2 52-1.2.2.2.2 52-1.2.2.2.2.r 53-1.2.2.2.1.1 54-1.1.2 56-1.2.2.3.2.1.1.1 57-1.2.2.3 57-1.2.2.3.2 57-1.2.2.3.2.r 58-1.2.2.3.1.1 59-1.2.4.r 60-1.2.4.1 61-1.2.4.2 (a / and~e.35 :op1 (t4 / transfect-01~e.39 :ARG1 (c / cell-line~e.41 :name (n3 / name :op1 "MCF-7") :mod (g / gene :name (n / name :op1 "K-RAS") :mod (w / wild-type))) :ARG2 (o / or~e.54 :op1 (v2 / vector :name (n2 / name :op1 "con.-vector")) :op2 (v / vector :name (n4 / name :op1 "K-RASV12"))) :ARG1-of (t5 / transient-02)) :op2 (t / treat-04~e.43 :ARG1 c :ARG2~e.44 (a2 / and :op1 (s / small-molecule~e.47 :name (n5 / name :op1 "erlotinib"~e.48) :ARG0-of~e.47 (i / inhibit-01~e.47 :ARG1 (p / protein :name (n6 / name :op1 "erbB1"~e.46)))) :op2 (s3 / small-molecule~e.52 :name (n7 / name :op1 "LY294002"~e.53) :ARG0-of~e.52 (i2 / inhibit-01~e.52 :ARG1 (p2 / protein-family :name (n9 / name :op1 "PI3K"~e.51)))) :op3 (s2 / small-molecule~e.57 :name (n8 / name :op1 "PD98059"~e.58) :ARG0-of~e.57 (i3 / inhibit-01~e.57 :ARG1 (p3 / protein-family :name (n10 / name :op1 "MEK"~e.56))))) :time (a3 / after~e.38 :op1 t4~e.5,6,7,8,9,10,11,12,13,14,15,16,17,18,19,20,21,22,23,24,25,26,27,28,29,30,31,32,33 :quant (t2 / temporal-quantity :quant 48~e.36 :unit (h / hour~e.37))) :duration~e.59 (t3 / temporal-quantity :quant 2~e.60 :unit h~e.61)) :purpose (t6 / this~e.1)) # ::id a_pmid_2139_2397.175 ::amr-annotator SDL-AMR-09 ::preferred # ::tok Similar to the results shown in Figure 3 , overexpression of K @-@ RAS @ V12 @ resulted in an about 2.5 @-@ fold stimulation of YB @-@ 1 phosphorylation . # ::alignments 0-1.3 1-1.3.1.r 3-1.3.1.1 4-1.3.1 4-1.3.1.2 4-1.3.1.2.r 5-1.3.1.2.1.r 6-1.3.1.2.1 8-1.3.1.2.1.1 11-1.1 14-1.1.1.1.1 16-1.1.1.1.1 19-1.1.1.2.1 21-1 21-1.3.1.1 22-1.2.r 24-1.2.2 25-1.2.2.1.1 27-1.2.2.1 28-1.2 29-1.2.1.r 29-1.2.2.1 30-1.2.1.1.1.1 32-1.2.1.1.1.1 33-1.2.1 (r2 / result-01~e.21 :ARG1 (o / overexpress-01~e.11 :ARG1 (g / gene :name (n2 / name :op1 "K-RAS"~e.14,16) :ARG2-of (m / mutate-01 :value "V12"~e.19))) :ARG2~e.22 (s / stimulate-01~e.28 :ARG1~e.29 (p / phosphorylate-01~e.33 :ARG1 (p2 / protein :name (n / name :op1 "YB-1"~e.30,32))) :degree (a / about~e.24 :op1 (p3 / product-of~e.27,29 :op1 2.5~e.25))) :ARG1-of (r / resemble-01~e.0 :ARG2~e.1 (t / thing~e.4 :ARG2-of (r3 / result-01~e.3,21) :ARG1-of~e.4 (s2 / show-01~e.4 :medium~e.5 (f / figure~e.6 :mod 3~e.8))))) # ::id a_pmid_2139_2397.176 ::amr-annotator SDL-AMR-09 ::preferred # ::tok Erlotinib reduced mutated K @-@ RAS @ V12 @ -@ induced YB @-@ 1 phosphorylation by about 50 % , while the PI3K inhibitor and the MEK inhibitor reduced K @-@ RAS @ V12 @ -@ induced YB @-@ 1 phosphorylation to the control level . # ::alignments 0-1.1.1.1.1 1-1.1 2-1.1.2.2.1.2 4-1.1.2.2.1.1.1 6-1.1.2.2.1.1.1 9-1.1.2.2.1.2.1 12-1.1.2.2 13-1.1.2.1.1.1 15-1.1.2.1.1.1 16-1.1.2 19-1.1.3.1 20-1.1.3 22-1 24-1.2.1.1.1.1.1.1 25-1.2.1.1 25-1.2.1.1.1 25-1.2.1.1.1.r 25-1.2.1.2 25-1.2.1.2.1 25-1.2.1.2.1.r 26-1.2.1 28-1.2.1.2.1.1.1.1 29-1.2.1.1 29-1.2.1.1.1 29-1.2.1.1.1.r 29-1.2.1.2 29-1.2.1.2.1 29-1.2.1.2.1.r 30-1.1 30-1.2 32-1.2.2 33-1.2.2 34-1.2.2 35-1.2.2 36-1.2.2 37-1.2.2 38-1.2.2 39-1.2.2 40-1.2.2 41-1.2.2 42-1.2.2 43-1.2.2 44-1.2.2 45-1.2.3.r 47-1.2.3.1 48-1.2.3 (c / contrast-01~e.22 :ARG1 (r / reduce-01~e.1,30 :ARG0 (s / small-molecule :name (n3 / name :op1 "erlotinib"~e.0)) :ARG1 (p2 / phosphorylate-01~e.16 :ARG1 (p3 / protein :name (n4 / name :op1 "YB-1"~e.13,15)) :ARG2-of (i2 / induce-01~e.12 :ARG0 (g / gene :name (n / name :op1 "K-RAS"~e.4,6) :ARG2-of (m / mutate-01~e.2 :value "V12"~e.9)))) :ARG2 (p4 / percentage-entity~e.20 :value 50~e.19)) :ARG2 (r2 / reduce-01~e.30 :ARG0 (a / and~e.26 :op1 (m2 / molecular-physical-entity~e.25,29 :ARG0-of~e.25,29 (i / inhibit-01~e.25,29 :ARG1 (p / protein-family :name (n5 / name :op1 "PI3K"~e.24)))) :op2 (m3 / molecular-physical-entity~e.25,29 :ARG0-of~e.25,29 (i3 / inhibit-01~e.25,29 :ARG1 (p5 / protein-family :name (n2 / name :op1 "MEK"~e.28))))) :ARG1 p2~e.32,33,34,35,36,37,38,39,40,41,42,43,44 :ARG4~e.45 (l / level~e.48 :mod (c2 / control~e.47)))) # ::id a_pmid_2139_2397.177 ::amr-annotator SDL-AMR-09 ::preferred # ::tok However , the combination of PD98059 and LY294002 ( PD/LY ) blocked basal and K @-@ RAS @ V12 @ -@ induced YB @-@ 1 phosphorylation completely ( Figure 5A ) . # ::alignments 0-1 3-1.1.1 4-1.1.1.1.r 5-1.1.1.1.1.1 7-1.1.1.2.1.1 11-1.1 12-1.1.2.1.2 13-1.1.2 15-1.1.2.2.2.1.1.1 17-1.1.2.2.2.1.1.1 20-1.1.2.2.2.1.2.1 23-1.1.2.2.2 24-1.1.2.1.1.1.1 26-1.1.2.1.1.1.1 27-1.1.2.1 27-1.1.2.2 28-1.1.3 30-1.2.1 32-1.2.1.1 (h / have-concession-91~e.0 :ARG1 (b / block-01~e.11 :ARG0 (c / combine-01~e.3 :ARG1~e.4 (s / small-molecule :name (n2 / name :op1 "PD98059"~e.5)) :ARG2 (s2 / small-molecule :name (n3 / name :op1 "LY294002"~e.7))) :ARG1 (a / and~e.13 :op1 (p / phosphorylate-01~e.27 :ARG1 (p2 / protein :name (n4 / name :op1 "YB-1"~e.24,26)) :mod (b2 / basal~e.12)) :op2 (p3 / phosphorylate-01~e.27 :ARG1 p2 :ARG2-of (i / induce-01~e.23 :ARG0 (g / gene :name (n / name :op1 "K-RAS"~e.15,17) :ARG2-of (m3 / mutate-01 :value "V12"~e.20))))) :ARG1-of (c2 / complete-02~e.28)) :ARG1-of (d2 / describe-01 :ARG0 (f / figure~e.30 :mod "5A"~e.32))) # ::id a_pmid_2139_2397.178 ::amr-annotator SDL-AMR-09 ::preferred # ::tok These data indicate that phosphorylation of YB @-@ 1 due to mutation of K @-@ RAS @ in part depends on activation of erbB1 . # ::alignments 0-1.1.1 1-1.1 2-1 3-1.2.r 4-1.2.1 5-1.2.1.1.r 6-1.2.1.1.1.1 8-1.2.1.1.1.1 9-1.2.1.2 10-1.2.1.2 11-1.2.1.2.1 14-1.2.1.2.1.1.1.1 16-1.2.1.2.1.1.1.1 18-1.2.3.r 19-1.2.3 19-1.2.3.r 20-1.2 21-1.2.2.r 22-1.2.2 23-1.2.2.1.r 24-1.2.2.1.1.1 (i / indicate-01~e.2 :ARG0 (d / data~e.1 :mod (t / this~e.0)) :ARG1~e.3 (d2 / depend-01~e.20 :ARG0 (p / phosphorylate-01~e.4 :ARG1~e.5 (p2 / protein :name (n2 / name :op1 "YB-1"~e.6,8)) :ARG1-of (c / cause-01~e.9,10 :ARG0 (m / mutate-01~e.11 :ARG1 (g / gene :name (n / name :op1 "K-RAS"~e.14,16))))) :ARG1~e.21 (a / activate-01~e.22 :ARG1~e.23 (p3 / protein :name (n3 / name :op1 "erbB1"~e.24))) :degree~e.18,19 (p4 / part~e.19))) # ::id a_pmid_2139_2397.179 ::amr-annotator SDL-AMR-09 ::preferred # ::tok This is most likely mediated by autocrine production of ligands and is in part independent of erbB1 , but it is dependent on activation of the PI3K @/@ Akt and MAPK @/@ ERK pathways . # ::alignments 0-1.1.1.1.2 2-1.1.1.2 3-1.1.1 4-1.1.1.1 5-1.1.1.1.1.r 6-1.1.1.1.1.2 7-1.1.1.1.1 8-1.1.1.1.1.1.r 9-1.1.1.1.1.1 10-1.1 12-1.1.2.4.r 13-1.1.2.4 13-1.1.2.4.r 14-1.1.2 14-1.1.2.1 14-1.1.2.1.r 15-1.1.2.3.r 16-1.1.2.3.1.1 18-1 19-1.2.1 21-1.2 22-1.2.2.r 23-1.2.2 24-1.2.2.1.r 26-1.2.2.1.1.1.1 28-1.2.2.1.1.1.1 29-1.2.2.1 30-1.2.2.1.2.1.1 32-1.2.2.1.2.1.1 33-1.2.2.1.1 33-1.2.2.1.2 (c / contrast-01~e.18 :ARG1 (a / and~e.10 :op1 (l / likely-01~e.3 :ARG1 (m2 / mediate-01~e.4 :ARG0~e.5 (p2 / produce-01~e.7 :ARG1~e.8 (l2 / ligand~e.9) :mod (a2 / autocrine~e.6)) :ARG1 (t / this~e.0)) :degree (m / most~e.2)) :op2 (d / depend-01~e.14 :polarity~e.14 -~e.14 :ARG0 t :ARG1~e.15 (p3 / protein :name (n2 / name :op1 "erbB1"~e.16)) :degree~e.12,13 (p5 / part~e.13))) :ARG2 (d2 / depend-01~e.21 :ARG0 t~e.19 :ARG1~e.22 (a3 / activate-01~e.23 :ARG1~e.24 (a4 / and~e.29 :op1 (p / pathway~e.33 :name (n / name :op1 "PI3K/Akt"~e.26,28)) :op2 (p4 / pathway~e.33 :name (n3 / name :op1 "MAPK/ERK"~e.30,32)))))) # ::id a_pmid_2139_2397.180 ::amr-annotator SDL-AMR-09 ::preferred # ::tok Because K @-@ Ras strongly induces YB @-@ 1 phosphorylation when it is mutated ( Figures 3 and 5A ) , we next analyzed whether phosphorylation of YB @-@ 1 in K @-@ RAS @ wt @ cells after irradiation or stimulation with EGF depends on K @-@ Ras expression . # ::alignments 0-1.4 1-1.4.1.1 2-1.4.1.1 3-1.4.1.1 4-1.4.1.4 4-1.4.1.4.r 5-1.4.1 6-1.4.1.2 7-1.4.1.2 8-1.4.1.2 9-1.4.1.2 10-1.2.4.r 10-1.3.r 13-1.4.1.3 15-1.4.2.1.1 15-1.4.2.1.2 17-1.4.2.1.1.1 19-1.4.2.1 21-1.4.2.1.2.1 25-1.1 26-1.3 27-1 28-1.2.1 28-1.2.1.r 29-1.2.2 30-1.2.2.1.r 31-1.2.2.1.1.1 33-1.2.2.1.1.1 36-1.2.2.2.1.1.1 38-1.2.2.2.1.1.1 41-1.2.2.2.1.2 43-1.2.2.2 44-1.2.4 45-1.2.4.1.1 46-1.2.4.1 47-1.2.4.1.2 48-1.2.4.1.2.2.r 49-1.2.4.1.2.2.1.1 50-1.2 51-1.2.3.r 52-1.2.3.1.1.1 54-1.2.3.1.1.1 55-1.2.3 (a / analyze-01~e.27 :ARG0 (w / we~e.25) :ARG1 (d / depend-01~e.50 :mode~e.28 interrogative~e.28 :ARG0 (p / phosphorylate-01~e.29 :ARG1~e.30 (p2 / protein :name (n2 / name :op1 "YB-1"~e.31,33)) :location (c / cell~e.43 :mod (g / gene :name (n3 / name :op1 "K-RAS"~e.36,38) :mod (w2 / wild-type~e.41)))) :ARG1~e.51 (e2 / express-03~e.55 :ARG2 (e / enzyme :name (n / name :op1 "K-Ras"~e.52,54))) :time~e.10 (a2 / after~e.44 :op1 (o / or~e.46 :op1 (i / irradiate-01~e.45 :ARG1 c) :op2 (s / stimulate-01~e.47 :ARG1 c :ARG2~e.48 (p3 / protein :name (n7 / name :op1 "EGF"~e.49)))))) :time~e.10 (n5 / next~e.26) :ARG1-of (c2 / cause-01~e.0 :ARG0 (i2 / induce-01~e.5 :ARG0 e~e.1,2,3 :ARG2 p~e.6,7,8,9 :condition (m / mutate-01~e.13 :ARG2 e) :manner~e.4 (s3 / strong~e.4)) :ARG1-of (d2 / describe-01 :ARG0 (a3 / and~e.19 :op1 (f / figure~e.15 :mod 3~e.17) :op2 (f2 / figure~e.15 :mod "5A"~e.21))))) # ::id a_pmid_2139_2397.181 ::amr-annotator SDL-AMR-09 ::preferred # ::tok Therefore , following downregulation of K @-@ Ras by siRNA , SKBr3 cells were irradiated or stimulated with EGF . # ::alignments 0-1 2-1.1.3 3-1.1.3.1 4-1.1.3.1.1.r 5-1.1.3.1.1.1.1 7-1.1.3.1.1.1.1 8-1.1.3.1.2.r 9-1.1.3.1.2.1.1 11-1.1.1.1.1.1 12-1.1.1.1 14-1.1.1 15-1.1 16-1.1.2 17-1.1.2.2.r 18-1.1.2.2.1.1 (i / infer-01~e.0 :ARG1 (o / or~e.15 :op1 (i2 / irradiate-01~e.14 :ARG1 (c / cell-line~e.12 :name (n2 / name :op1 "SKBr3"~e.11))) :op2 (s / stimulate-01~e.16 :ARG1 c :ARG2~e.17 (p / protein :name (n3 / name :op1 "EGF"~e.18))) :ARG1-of (f / follow-01~e.2 :ARG2 (d / downregulate-01~e.3 :ARG1~e.4 (e / enzyme :name (n / name :op1 "K-Ras"~e.5,7)) :ARG2~e.8 (n5 / nucleic-acid :name (n4 / name :op1 "siRNA"~e.9)))))) # ::id a_pmid_2139_2397.182 ::amr-annotator SDL-AMR-09 ::preferred # ::tok As shown in Figure 5B , downregulation of K @-@ Ras did not affect either IR @- or EGF @-@ induced YB @-@ 1 phosphorylation . # ::alignments 1-1.4 2-1.4.1.r 3-1.4.1 5-1.4.1.1 8-1.2 9-1.2.1.r 10-1.2.1.1.1 12-1.2.1.1.1 14-1.1 14-1.1.r 15-1 17-1.3.1.2.1 17-1.3.1.2.1.1 17-1.3.1.2.1.1.r 19-1.3 20-1.3.2.2.1.1.1 22-1.3.1.2 22-1.3.2.2 23-1.3.1.1.1.1 25-1.3.1.1.1.1 26-1.3.1 26-1.3.2 (a / affect-01~e.15 :polarity~e.14 -~e.14 :ARG0 (d / downregulate-01~e.8 :ARG1~e.9 (e / enzyme :name (n / name :op1 "K-Ras"~e.10,12))) :ARG1 (o / or~e.19 :op1 (p / phosphorylate-01~e.26 :ARG1 (p2 / protein :name (n2 / name :op1 "YB-1"~e.23,25)) :ARG2-of (i / induce-01~e.22 :ARG0 (r / radiate-01~e.17 :ARG0-of~e.17 (i3 / ionize-01~e.17)))) :op2 (p3 / phosphorylate-01~e.26 :ARG1 p2 :ARG2-of (i2 / induce-01~e.22 :ARG0 (p4 / protein :name (n4 / name :op1 "EGF"~e.20))))) :ARG1-of (s3 / show-01~e.1 :medium~e.2 (f / figure~e.3 :mod "5B"~e.5))) # ::id a_pmid_2139_2397.183 ::amr-annotator SDL-AMR-09 ::preferred # ::tok A lack of effect of K @-@ RAS @ -@ siRNA on P @-@ ERK1 @/@ 2 was observed as well ( Figure 5B ) . # ::alignments 1-1.1 2-1.1.1.r 3-1.1.1 6-1.1.1.1.2.1.1 8-1.1.1.1.2.1.1 11-1.1.1.1.1.1 12-1.1.1.2.r 13-1.1.1.2.3 15-1.1.1.2.1.1.1 16-1.1.1.2 19-1 20-1.2 21-1.2 23-1.3.1 25-1.3.1.1 (o / observe-01~e.19 :ARG1 (l / lack-01~e.1 :ARG1~e.2 (a / affect-01~e.3 :ARG0 (n / nucleic-acid :name (n2 / name :op1 "siRNA"~e.11) :mod (g / gene :name (n3 / name :op1 "K-RAS"~e.6,8))) :ARG1~e.12 (s / slash~e.16 :op1 (e3 / enzyme :name (n4 / name :op1 "ERK1"~e.15)) :op2 (e4 / enzyme :name (n5 / name :op1 "ERK2")) :ARG1-of (p / phosphorylate-01~e.13)))) :manner (a2 / as-well~e.20,21) :ARG1-of (d / describe-01 :ARG0 (f / figure~e.23 :mod "5B"~e.25))) # ::id a_pmid_2139_2397.184 ::amr-annotator SDL-AMR-09 ::preferred # ::tok YB @-@ 1 regulates repair of IR @-@ induced DNA @-@ DSB and postirradiation survival # ::alignments 0-1.1.1.1 2-1.1.1.1 3-1 4-1.2.1 5-1.2.1.1.r 6-1.2.1.1.3.1 6-1.2.1.1.3.1.1 6-1.2.1.1.3.1.1.r 8-1.2.1.1.3 9-1.2.1.1.2.2.1 11-1.2.1.1.1.1 12-1.2 14-1.2.2 (r / regulate-01~e.3 :ARG0 (p / protein :name (n / name :op1 "YB-1"~e.0,2)) :ARG1 (a / and~e.12 :op1 (r2 / repair-01~e.4 :ARG1~e.5 (e / event :name (n2 / name :op1 "DSB"~e.11) :mod (n3 / nucleic-acid :wiki "DNA" :name (n4 / name :op1 "DNA"~e.9)) :ARG2-of (i / induce-01~e.8 :ARG0 (r3 / radiate-01~e.6 :ARG0-of~e.6 (i3 / ionize-01~e.6))))) :op2 (s2 / survive-01~e.14 :time (a2 / after :op1 (i2 / irradiate-01))))) # ::id a_pmid_2139_2397.185 ::amr-annotator SDL-AMR-09 ::preferred # ::tok In addition to its function as a transcription factor , YB @-@ 1 is also involved in DNA repair , that is , base excision repair and mismatch repair [ @ 39 @ ] . # ::alignments 0-1.2.2.1 1-1.2.2.1 1-1.4 3-1.4.1.1 3-1.4.1.1.r 4-1.4.1 5-1.4.1.2.r 7-1.4.1.2.1 8-1.4.1.2 10-1.1.1.1 12-1.1.1.1 14-1.3 15-1 17-1.2.1.2.1 18-1.2 23-1.2.2.1.1.1.1 24-1.2.2.1.1.1.2 25-1.2.2.1.1.1.3 26-1.2.2.1 27-1.2.2.1.2.1.1 28-1.2.2.1.2.1.2 31-1.5.1.1.1 (i / involve-01~e.15 :ARG1 (p / protein :name (n / name :op1 "YB-1"~e.10,12)) :ARG2 (r / repair-01~e.18 :ARG1 (n4 / nucleic-acid :wiki "DNA" :name (n5 / name :op1 "DNA"~e.17)) :ARG1-of (m / mean-01 :ARG2 (a3 / and~e.0,1,26 :op1 (e / event :name (n2 / name :op1 "base"~e.23 :op2 "excision"~e.24 :op3 "repair"~e.25)) :op2 (e2 / event :name (n3 / name :op1 "mismatch"~e.27 :op2 "repair"~e.28))))) :mod (a / also~e.14) :ARG1-of (a2 / add-02~e.1 :ARG2 (f / function-01~e.4 :ARG0~e.3 p~e.3 :ARG1~e.5 (f2 / factor~e.8 :ARG0-of (t / transcribe-01~e.7)))) :ARG1-of (d2 / describe-01 :ARG0 (p2 / publication :ARG1-of (c / cite-01 :ARG2 39~e.31)))) # ::id a_pmid_2139_2397.186 ::amr-annotator SDL-AMR-09 ::preferred # ::tok In line with this function , it has been demonstrated that YB @-@ 1 binds to double @-@ stranded , single @-@ stranded and DNA @-@ containing abasic sites [ @ 40 @ ] . # ::alignments 3-1.3.1.1 4-1.3.1 9-1 10-1.1.r 11-1.1.1.1.1 13-1.1.1.1.1 14-1.1 15-1.1.2.r 16-1.1.2.1.3.1 18-1.1.2.1.3 20-1.1.2.2.3.1 22-1.1.2.1.3 22-1.1.2.2.3 23-1.1.2 24-1.1.2.1.2.1 24-1.1.2.2.2.1 24-1.1.2.3.2.1.2.1 26-1.1.2.3.2 27-1.1.2.3.1.1 28-1.1.2.3.1.2 31-1.2.1.1.1 (d / demonstrate-01~e.9 :ARG1~e.10 (b / bind-01~e.14 :ARG1 (p / protein :name (n / name :op1 "YB-1"~e.11,13)) :ARG2~e.15 (a / and~e.23 :op1 (n2 / nucleic-acid :wiki "DNA" :name (n4 / name :op1 "DNA"~e.24) :mod (s / strand~e.18,22 :mod (d3 / double~e.16))) :op2 (n5 / nucleic-acid :wiki "DNA" :name (n6 / name :op1 "DNA"~e.24) :mod (s2 / strand~e.22 :ARG1-of (s3 / single-02~e.20))) :op3 (d2 / dna-sequence :name (n3 / name :op1 "abasic"~e.27 :op2 "site"~e.28) :ARG0-of (c / contain-01~e.26 :ARG1 (n7 / nucleic-acid :wiki "DNA" :name (n8 / name :op1 "DNA"~e.24)))))) :ARG1-of (d7 / describe-01 :ARG0 (p2 / publication :ARG1-of (c2 / cite-01 :ARG2 40~e.31))) :ARG1-of (r / resemble-01 :ARG2 (f / function~e.4 :mod (t / this~e.3)))) # ::id a_pmid_2139_2397.187 ::amr-annotator SDL-AMR-09 ::preferred # ::tok So far , however , no data demonstrating the function of YB @-@ 1 in repair of IR @-@ induced DNA @-@ DSB and postirradiation survival exist . # ::alignments 0-1.2 1-1.2 3-1 5-1.1.1.1 5-1.1.1.1.r 6-1.1.1 7-1.1.1.2 9-1.1.1.2.1 10-1.1.1.2.1.1.r 11-1.1.1.2.1.1.1.1 13-1.1.1.2.1.1.1.1 14-1.1.1.2.1.2.r 15-1.1.1.2.1.2.1 16-1.1.1.2.1.2.1.2.r 17-1.1.1.2.1.2.1.2.1 17-1.1.1.2.1.2.1.2.1.1 17-1.1.1.2.1.2.1.2.1.1.r 19-1.1.1.2.1.2.1.2 20-1.1.1.2.1.2.1.1.2.2.1 22-1.1.1.2.1.2.1.1.1.1 23-1.1.1.2.1.2 25-1.1.1.2.1.2.2 26-1.1 (h / have-concession-91~e.3 :ARG1 (e / exist-01~e.26 :ARG1 (d / data~e.6 :polarity~e.5 -~e.5 :ARG0-of (d2 / demonstrate-01~e.7 :ARG1 (f / function-01~e.9 :ARG0~e.10 (p / protein :name (n / name :op1 "YB-1"~e.11,13)) :ARG1~e.14 (a / and~e.23 :op1 (r / repair-01~e.15 :ARG1 (e2 / event :name (n2 / name :op1 "DSB"~e.22) :mod (n3 / nucleic-acid :wiki "DNA" :name (n4 / name :op1 "DNA"~e.20))) :ARG2-of~e.16 (i / induce-01~e.19 :ARG0 (r2 / radiate-01~e.17 :ARG0-of~e.17 (i3 / ionize-01~e.17)))) :op2 (s3 / survive-01~e.25 :time (a2 / after :op1 (i2 / irradiate-01)))))))) :time (s / so-far~e.0,1)) # ::id a_pmid_2139_2397.188 ::amr-annotator SDL-AMR-09 ::preferred # ::tok The function of erbB1 and its downstream pathways and the impact of mutated K @-@ RAS @ on repair of DNA @-@ DSB have been demonstrated previously [ @ 15 , 34 , 41 , 42 @ ] . # ::alignments 1-1.1.1 2-1.1.1.1.r 3-1.1.1.1.1.1.1 4-1.1.1.1 5-1.1.1.1.2.1 5-1.1.1.1.2.1.r 6-1.1.1.1.2.2 7-1.1.1.1.2 8-1.1 8-1.1.1.1 10-1.1.2 12-1.1.2.1.2 14-1.1.2.1.1.1 16-1.1.2.1.1.1 19-1.1.2.2 20-1.1.2.2.1.r 21-1.1.2.2.1.2.2.1 23-1.1.2.2.1.1.1 24-1.1.2 26-1 27-1.2 30-1.3.1.1.1.1 34-1.3.1.2.1.1 38-1.3.1.3.1.1 42-1.3.1.4.1.1 (d / demonstrate-01~e.26 :ARG1 (a / and~e.8 :op1 (f / function-01~e.1 :ARG0~e.2 (a2 / and~e.4,8 :op1 (p2 / protein :name (n2 / name :op1 "erbB1"~e.3)) :op2 (p3 / pathway~e.7 :poss~e.5 p2~e.5 :location (d2 / downstream~e.6)))) :op2 (i / impact-01~e.10,24 :ARG0 (g / gene :name (n / name :op1 "K-RAS"~e.14,16) :ARG2-of (m / mutate-01~e.12)) :ARG1 (r / repair-01~e.19 :ARG1~e.20 (e / event :name (n3 / name :op1 "DSB"~e.23) :mod (n4 / nucleic-acid :wiki "DNA" :name (n5 / name :op1 "DNA"~e.21)))))) :time (p / previous~e.27) :ARG1-of (d4 / describe-01 :ARG0 (a3 / and :op1 (p4 / publication :ARG1-of (c / cite-01 :ARG2 15~e.30)) :op2 (p5 / publication :ARG1-of (c2 / cite-01 :ARG2 34~e.34)) :op3 (p6 / publication :ARG1-of (c3 / cite-01 :ARG2 41~e.38)) :op4 (p7 / publication :ARG1-of (c4 / cite-01 :ARG2 42~e.42))))) # ::id a_pmid_2139_2397.189 ::amr-annotator SDL-AMR-09 ::preferred # ::tok Therefore , we next asked whether the cells presenting a differential pattern of basal @- and radiation @-@ induced YB @-@ 1 phosphorylation additionally exert a differential sensitivity to IR. # ::alignments 0-1 2-1.1.1 3-1.1.3 4-1.1 5-1.1.2.1 5-1.1.2.1.r 7-1.1.2.2 8-1.1.2.2.1 10-1.1.2.2.1.1.1 11-1.1.2.2.1.1 13-1.1.2.2.1.1.2.1.2 15-1.1.2.2.1.1.2 16-1.1.2.2.1.1.2.2.2.1 16-1.1.2.3.2 18-1.1.2.2.1.1.2.2.2 19-1.1.2.2.1.1.2.1.1.1.1 21-1.1.2.2.1.1.2.1.1.1.1 22-1.1.2.2.1.1.2.1 22-1.1.2.2.1.1.2.2 23-1.1.2.2.1.1.2 23-1.1.2.4 23-1.1.2.4.r 24-1.1.2 26-1.1.2.3.3 27-1.1.2.3 28-1 (c2 / cause-01~e.0,28 :ARG1 (a / ask-01~e.4 :ARG0 (w / we~e.2) :ARG1 (e / exert-01~e.24 :mode~e.5 interrogative~e.5 :ARG0 (c / cell~e.7 :ARG0-of (p2 / present-01~e.8 :ARG1 (p3 / pattern~e.11 :mod (d / differential~e.10) :topic (a3 / and~e.15,23 :op1 (p / phosphorylate-01~e.22 :ARG1 (p4 / protein :name (n2 / name :op1 "YB-1"~e.19,21)) :mod (b / basal~e.13)) :op2 (p5 / phosphorylate-01~e.22 :ARG1 p4 :ARG2-of (i2 / induce-01~e.18 :ARG0 (r / radiate-01~e.16))))))) :ARG1 (s / sensitive-03~e.27 :ARG0 c :ARG1 (r2 / radiate-01~e.16 :ARG0-of (i / ionize-01)) :mod d~e.26) :manner~e.23 (a2 / additional~e.23)) :time (n / next~e.3))) # ::id a_pmid_2139_2397.190 ::amr-annotator SDL-AMR-09 ::preferred # ::tok The results obtained by clonogenic assay indicate a differential response in terms of postirradiation survival of the cell lines analyzed . # ::alignments 1-1.1 1-1.1.1 1-1.1.1.r 2-1.1.2 3-1.1.2.1.r 4-1.1.2.1.1 5-1.1.2.1 6-1 8-1.2.2 9-1.2 14-1.2.1 15-1.2.1.1.r 17-1.2.1.1 18-1.2.1.1 19-1.2.1.1.1 (i / indicate-01~e.6 :ARG0 (t / thing~e.1 :ARG2-of~e.1 (r / result-01~e.1) :ARG1-of (o / obtain-01~e.2 :manner~e.3 (a3 / assay-01~e.5 :mod (c2 / clonogenic~e.4)))) :ARG1 (r2 / respond-01~e.9 :ARG2 (s / survive-01~e.14 :ARG0~e.15 (c / cell-line~e.17,18 :ARG1-of (a2 / analyze-01~e.19)) :time (a / after :op1 (i2 / irradiate-01))) :ARG1-of (d / differ-02~e.8))) # ::id a_pmid_2139_2397.191 ::amr-annotator SDL-AMR-09 ::preferred # ::tok The radiation dose , D @ 37 , which is required to reduce cell survival to 37 % , is 1.95 Gy for SKBr3 , 1.65 Gy for MDA @-@ MB @-@ 23 , 1.35 Gy for MCF @-@ 7 and 1.10 Gy for HBL100 cells . # ::alignments 1-1.1.1 1-1.2.1 1-1.2.2 1-1.2.3 1-1.2.4 2-1.1 6-1.1.2.1.2.1 11-1.1.2 13-1.1.2.1 14-1.2.2.3 14-1.2.3.3 14-1.2.4.3 15-1.1.2.1.1 17-1.1.2.1.2.1 18-1.1.2.1.2 21-1.2.1.1 23-1.2.1.3.r 24-1.2.1.3.1.1 26-1.2.2.1 29-1.2.2.3.1.1 31-1.2.2.3.1.1 33-1.2.2.3.1.1 35-1.2.3.1 38-1.2.3.3.1.1 40-1.2.3.3.1.1 41-1.2 42-1.2.4.1 45-1.2.4.3.1.1 46-1.2.1.3 (e / equal-01 :ARG1 (d / dose-01~e.2 :ARG1 (r5 / radiate-01~e.1) :ARG1-of (r6 / require-01~e.11 :ARG0 (r7 / reduce-01~e.13 :ARG1 (s / survive-01~e.15 :ARG0 c) :ARG4 (p2 / percentage-entity~e.18 :value 37~e.6,17))) :ARG1-of (l / label-01 :ARG2 (s2 / string-entity :value "D37"))) :ARG2 (a / and~e.41 :op1 (r / radiation-quantity~e.1 :quant 1.95~e.21 :unit (g / gray) :location~e.23 (c / cell-line~e.46 :name (n / name :op1 "SKBr3"~e.24))) :op2 (r2 / radiation-quantity~e.1 :quant 1.65~e.26 :unit g :location (c2 / cell-line~e.14 :name (n2 / name :op1 "MDA-MB-23"~e.29,31,33))) :op3 (r3 / radiation-quantity~e.1 :quant 1.35~e.35 :unit g :location (c3 / cell-line~e.14 :name (n3 / name :op1 "MCF-7"~e.38,40))) :op4 (r4 / radiation-quantity~e.1 :quant 1.10~e.42 :unit g :location (c4 / cell-line~e.14 :name (n4 / name :op1 "HBL100"~e.45))))) # ::id a_pmid_2139_2397.192 ::amr-annotator SDL-AMR-09 ::preferred # ::tok We further investigated whether YB @-@ 1 activity is involved in the process of DNA @-@ DSB repair and postirradiation survival . # ::alignments 0-1.1 1-1.3 2-1 3-1.2.1 3-1.2.1.r 4-1.2.2.1.1.1 6-1.2.2.1.1.1 7-1.2.2 9-1.2 10-1.2.3.r 12-1.2.3.1 13-1.2.3.1.1.r 14-1.2.3.1.1.1.2.2.1 16-1.2.3.1.1.1.1.1 17-1.2.3.1.1 18-1.2.3 20-1.2.3.2 (i / investigate-01~e.2 :ARG0 (w / we~e.0) :ARG1 (i2 / involve-01~e.9 :mode~e.3 interrogative~e.3 :ARG1 (a / activity-06~e.7 :ARG0 (p / protein :name (n / name :op1 "YB-1"~e.4,6))) :ARG2~e.10 (a2 / and~e.18 :op1 (p2 / process-02~e.12 :ARG1~e.13 (r / repair-01~e.17 :ARG1 (e / event :name (n2 / name :op1 "DSB"~e.16) :mod (n3 / nucleic-acid :wiki "DNA" :name (n4 / name :op1 "DNA"~e.14))))) :op2 (s / survive-01~e.20 :time (a3 / after :op1 (i3 / irradiate-01))))) :degree (f / further~e.1)) # ::id a_pmid_2139_2397.193 ::amr-annotator SDL-AMR-09 ::preferred # ::tok For this purpose , a siRNA approach was used . # ::alignments 1-1.2 2-1.2.r 5-1.1.1.1.1 6-1.1 8-1 8-1.2.r (u / use-01~e.8 :ARG1 (a / approach-02~e.6 :mod (n2 / nucleic-acid :name (n / name :op1 "siRNA"~e.5))) :purpose~e.2,8 (t / this~e.1)) # ::id a_pmid_2139_2397.194 ::amr-annotator SDL-AMR-09 ::preferred # ::tok As shown in Figure 6 , downregulation of YB @-@ 1 by siRNA , either in K @-@ RAS @ mt @ MDA @-@ MB @-@ 231 or in K @-@ RAS @ wt @ SKBr3 cells , resulted in impaired repair of DNA @-@ DSB as shown by enhanced residual γ-H2AX foci 24 hours after irradiation . # ::alignments 0-1.4.2.r 1-1.3 2-1.3.1.r 3-1.3.1 5-1.3.1.1 8-1.1 9-1.1.1.r 10-1.1.1.1.1 12-1.1.1.1.1 13-1.1.2.r 14-1.1.2.1.1 19-1.1.3.1.2.1.1 19-1.1.3.2.2.1.1 21-1.1.3.1.2.1.1 21-1.1.3.2.2.1.1 26-1.1.3.1.1.1 28-1.1.3.1.1.1 30-1.1.3.1.1.1 31-1.1.3 34-1.1.3.1.2.1.1 34-1.1.3.2.2.1.1 36-1.1.3.1.2.1.1 36-1.1.3.2.2.1.1 39-1.1.3.2.2.2 41-1.1.3.2.1.1 42-1.1.3.1 42-1.1.3.2 44-1 45-1.2.r 46-1.2.2 47-1.2 48-1.2.1.r 49-1.2.1.2.2.1 51-1.2.1.1.1 52-1.4.2.r 53-1.4 54-1.4.1.r 55-1.4.1.3 56-1.4.1.2 57-1.4.1.1.1.1 58-1.4.1 59-1.4.2.2.1 60-1.4.2.2.2 61-1.4.2 62-1.4.2.1 (r / result-01~e.44 :ARG1 (d / downregulate-01~e.8 :ARG1~e.9 (p / protein :name (n2 / name :op1 "YB-1"~e.10,12)) :ARG2~e.13 (n11 / nucleic-acid :name (n3 / name :op1 "siRNA"~e.14)) :location (o / or~e.31 :op1 (c / cell-line~e.42 :name (n4 / name :op1 "MDA-MB-231"~e.26,28,30) :mod (g / gene :name (n6 / name :op1 "K-RAS"~e.19,21,34,36) :ARG2-of (m / mutate-01))) :op2 (c2 / cell-line~e.42 :name (n5 / name :op1 "SKBr3"~e.41) :mod (g2 / gene :name (n7 / name :op1 "K-RAS"~e.19,21,34,36) :mod (w / wild-type~e.39))))) :ARG2~e.45 (r4 / repair-01~e.47 :ARG1~e.48 (e2 / event :name (n8 / name :op1 "DSB"~e.51) :mod (n / nucleic-acid :wiki "DNA" :name (n10 / name :op1 "DNA"~e.49))) :ARG1-of (i / impair-01~e.46)) :ARG1-of (s2 / show-01~e.1 :medium~e.2 (f2 / figure~e.3 :mod 6~e.5)) :ARG1-of (s / show-01~e.53 :ARG0~e.54 (f / focus~e.58 :mod (p2 / protein :name (n9 / name :op1 "γ-H2AX"~e.57)) :mod (r3 / residual~e.56) :ARG1-of (e3 / enhance-01~e.55)) :time~e.0,52 (a / after~e.61 :op1 (i2 / irradiate-01~e.62) :quant (t / temporal-quantity :quant 24~e.59 :unit (h / hour~e.60))))) # ::id a_pmid_2139_2397.195 ::amr-annotator SDL-AMR-09 ::preferred # ::tok Interestingly , downregulating K @-@ Ras resulted in enhanced frequency of residual DSB to the level observed with YB @-@ 1 siRNA . # ::alignments 0-1.3 2-1.1 3-1.1.1.1.1 5-1.1.1.1.1 6-1 8-1.2.2 9-1.2 11-1.2.1.2 12-1.2.1.1.1 13-1.2.2.1.r 15-1.2.2.1 16-1.2.2.1.1 17-1.2.2.1.1.1.r 18-1.2.2.1.1.1.2.1.1 20-1.2.2.1.1.1.2.1.1 21-1.2.2.1.1.1.1.1 (r / result-01~e.6 :ARG1 (d / downregulate-01~e.2 :ARG1 (e / enzyme :name (n / name :op1 "K-Ras"~e.3,5))) :ARG2 (h / have-frequency-91~e.9 :ARG1 (e3 / event :name (n2 / name :op1 "DSB"~e.12) :mod (r3 / residual~e.11)) :ARG1-of (e2 / enhance-01~e.8 :ARG3~e.13 (l / level~e.15 :ARG1-of (o / observe-01~e.16 :location~e.17 (n5 / nucleic-acid :name (n3 / name :op1 "siRNA"~e.21) :mod (p / protein :name (n4 / name :op1 "YB-1"~e.18,20))))))) :ARG2-of (i / interest-01~e.0)) # ::id a_pmid_2139_2397.196 ::amr-annotator SDL-AMR-09 ::preferred # ::tok Likewise , siRNA targeting of YB @-@ 1 increased radiation sensitivity tested in MDA @-@ MB @-@ 231 cells . # ::alignments 2-1.1.1.1.1 3-1.1 4-1.1.2.r 5-1.1.2.1.1 7-1.1.2.1.1 8-1 9-1.2.1 10-1.2 11-1.2.2 12-1.2.2.1.r 13-1.2.2.1.1.1 15-1.2.2.1.1.1 17-1.2.2.1.1.1 18-1.2.2.1 (i / increase-01~e.8 :ARG0 (t / target-01~e.3 :ARG0 (n4 / nucleic-acid :name (n2 / name :op1 "siRNA"~e.2)) :ARG1~e.4 (p / protein :name (n / name :op1 "YB-1"~e.5,7))) :ARG1 (s / sensitive-03~e.10 :ARG1 (r / radiate-01~e.9) :ARG2-of (t2 / test-01~e.11 :ARG1~e.12 (c / cell-line~e.18 :name (n3 / name :op1 "MDA-MB-231"~e.13,15,17)))) :ARG1-of (r3 / resemble-01)) # ::id a_pmid_2169_9731.11 ::amr-annotator SDL-AMR-09 ::preferred # ::tok Primary macrophages from both naïve and lung @-@ tumor bearing mice stimulated epithelial cell proliferation . # ::alignments 0-1.1.1 1-1.1 2-1.1.2.r 4-1.1.2.1.1 5-1.1.2 6-1.1.2.2.1.1.1 8-1.1.2.2.1.1 9-1.1.2.2.1 10-1.1.2.1 10-1.1.2.2 11-1 12-1.2.1.1 13-1.2.1 14-1.2 (s / stimulate-01~e.11 :ARG0 (m / macrophage~e.1 :mod (p2 / primary~e.0) :source~e.2 (a / and~e.5 :op1 (m2 / mouse~e.10 :mod (n / naive~e.4)) :op2 (m3 / mouse~e.10 :ARG0-of (b / bear-01~e.9 :ARG1 (t / tumor~e.8 :mod (l / lung~e.6)))))) :ARG1 (p / proliferate-01~e.14 :ARG0 (c / cell~e.13 :mod (e / epithelium~e.12)))) # ::id a_pmid_2169_9731.12 ::amr-annotator SDL-AMR-09 ::preferred # ::tok The lungs of tumor @-@ bearing mice contained 3.5 @-@ times more IGF @-@ 1 than naïve littermates , and media conditioned by freshly isolated tumor @-@ educated macrophages contained more IGF @-@ 1 than media conditioned by naïve macrophages ; IL @-@ 4 stimulated IGF @-@ 1 production by both macrophage subsets . # ::alignments 1-1.1.1.1 2-1.1.1.1.1.r 3-1.1.1.1.1.1.1 5-1.1.1.1.1.1 6-1.1.1.1.1 7-1.1.1 8-1.1.1.4.1.1 10-1.1.1.4.1 11-1.1.1.4 12-1.1.1.2.1.1 14-1.1.1.2.1.1 15-1.1.1.3.r 16-1.1.1.3.1 17-1.1.1.3 19-1.1 20-1.1.2.1 21-1.1.2.1.1 22-1.1.2.1.1.1.r 23-1.1.2.1.1.1.2.1 24-1.1.2.1.1.1.2 25-1.1.2.1.1.1.1.1 27-1.1.2.1.1.1.1 28-1.1.2.1.1.1 29-1.1.1 29-1.1.2 30-1.1.2.4 31-1.1.2.2 32-1.1.2.2 33-1.1.2.2 34-1.1.2.3.r 35-1.1.2.3 36-1.1.2.3.1 37-1.1.2.3.1.1.r 38-1.1.2.3.1.1.1 39-1.1.2.3.1.1 41-1.2.1.1.1 43-1.2.1.1.1 44-1.2 45-1.2.2.2.1.1 47-1.2.2.2.1.1 48-1.2.2 49-1.2.2.1.r 50-1.2.2.1.2 51-1.2.2.1.1 52-1.2.2.1 (m / multi-sentence :snt1 (a / and~e.19 :op1 (c / contain-01~e.7,29 :ARG0 (l / lung~e.1 :part-of~e.2 (m2 / mouse~e.6 :ARG0-of (b / bear-01~e.5 :ARG1 (t / tumor~e.3)))) :ARG1 (p2 / protein :name (n / name :op1 "IGF-1"~e.12,14)) :compared-to~e.15 (l2 / littermate~e.17 :mod (n2 / naive~e.16)) :quant (m3 / more~e.11 :degree (p / product-of~e.10 :op1 3.5~e.8))) :op2 (c2 / contain-01~e.29 :ARG0 (m4 / medium~e.20 :ARG1-of (c3 / condition-01~e.21 :ARG0~e.22 (m5 / macrophage~e.28 :ARG1-of (e / educate-01~e.27 :ARG0 (t2 / tumor~e.25)) :ARG1-of (i / isolate-01~e.24 :ARG1-of (f / fresh-04~e.23))))) :ARG1 p2~e.31,32,33 :compared-to~e.34 (m6 / medium~e.35 :ARG1-of (c4 / condition-01~e.36 :ARG0~e.37 (m7 / macrophage~e.39 :mod (n3 / naive~e.38)))) :quant (m9 / more~e.30))) :snt2 (s / stimulate-01~e.44 :ARG0 (p3 / protein :name (n4 / name :op1 "IL-4"~e.41,43)) :ARG1 (p5 / produce-01~e.48 :ARG0~e.49 (s2 / subset~e.52 :part-of (m8 / macrophage~e.51) :mod (b2 / both~e.50)) :ARG1 (p4 / protein :name (n5 / name :op1 "IGF-1"~e.45,47))))) # ::id a_pmid_2169_9731.13 ::amr-annotator SDL-AMR-09 ::preferred # ::tok The ability of macrophage conditioned media to stimulate neoplastic proliferation correlated with media IGF @-@ 1 levels , and recombinant IGF @-@ 1 alone was sufficient to induce epithelial proliferation in all cell lines evaluated . # ::alignments 1-1.1 2-1.1.2.r 3-1.1.2.1.1.1 4-1.1.2.1.1 5-1.1.2.1 7-1.1.2 9-1.1.2.2 10-1.1.2.2.2 11-1.1.2.2.2.1.r 12-1.1.2.2.2.1.1.1.2 13-1.1.2.2.2.1.1.1.1.1 13-1.1.2.2.2.1.2.1.1 15-1.1.2.2.2.1.1.1.1.1 15-1.1.2.2.2.1.2.1.1 16-1.1.2.2.2.1.1 18-1.1.2.2.2.1 19-1.1.2.2.2.1.2.3 20-1.1.2.2.2.1.2.1.1 22-1.1.2.2.2.1.2.1.1 23-1.1.2.2.2.1.2.2 25-1 27-1.2 28-1.2.2.1 29-1.2.2 30-1.2.3.r 31-1.2.3.2 32-1.2.3 33-1.2.3 34-1.2.3.1 (s / suffice-01~e.25 :ARG0 (c4 / capable-01~e.1 :ARG1 m :ARG2~e.2 (s2 / stimulate-01~e.7 :ARG0 (m / medium~e.5 :ARG1-of (c / condition-01~e.4 :ARG0 (m2 / macrophage~e.3))) :ARG1 (p2 / proliferate-01~e.9 :ARG0 (t / tumor) :ARG1-of (c2 / correlate-01~e.10 :ARG2~e.11 (a / and~e.18 :op1 (l / level~e.16 :quant-of (p3 / protein :name (n2 / name :op1 "IGF-1"~e.13,15) :mod (m3 / medium~e.12))) :op2 (p4 / protein :name (n3 / name :op1 "IGF-1"~e.13,15,20,22) :mod (a2 / alone~e.23) :ARG3-of (r2 / recombine-01~e.19))))))) :ARG1 (i / induce-01~e.27 :ARG0 c4 :ARG2 (p5 / proliferate-01~e.29 :ARG0 (e / epithelium~e.28)) :location~e.30 (c5 / cell-line~e.32,33 :ARG1-of (e2 / evaluate-01~e.34) :mod (a3 / all~e.31)))) # ::id a_pmid_2169_9731.14 ::amr-annotator SDL-AMR-09 ::preferred # ::tok Macrophage @-@ conditioned media and IGF @-@ 1 stimulated lung tumor cell growth in an additive manner , while EGF had no effect . # ::alignments 0-1.1.1.1.1.1 2-1.1.1.1.1 3-1.1.1.1 4-1.1.1 5-1.1.1.2.1.1 7-1.1.1.2.1.1 8-1.1 9-1.1.2.1.1.1 10-1.1.2.1.1 11-1.1.2.1 12-1.1.2 15-1.1.3 16-1.1.3.r 18-1 19-1.2.2.1.1 21-1.2.1 21-1.2.1.r 22-1.2 (c3 / contrast-01~e.18 :ARG1 (s / stimulate-01~e.8 :ARG0 (a / and~e.4 :op1 (m / medium~e.3 :ARG1-of (c / condition-01~e.2 :ARG0 (m2 / macrophage~e.0))) :op2 (p / protein :name (n / name :op1 "IGF-1"~e.5,7))) :ARG1 (g / grow-01~e.12 :ARG1 (c2 / cell~e.11 :mod (t / tumor~e.10 :mod (l / lung~e.9)))) :manner~e.16 (a2 / additive~e.15)) :ARG2 (a3 / affect-01~e.22 :polarity~e.21 -~e.21 :ARG0 (p2 / protein :name (n2 / name :op1 "EGF"~e.19)) :ARG1 g)) # ::id a_pmid_2169_9731.15 ::amr-annotator SDL-AMR-09 ::preferred # ::tok Macrophage @-@ derived factors increased p @-@ Erk1 @/@ 2 , p @-@ Akt and cyclin D1 levels in neoplastic cells , and the combined inhibition of both MEK and PI3K ablated macrophage @-@ mediated increases in epithelial growth . # ::alignments 0-1.1.1.1.1 2-1.1.1.1 3-1.1.1 4-1.1 5-1.1.2.1.1.2 7-1.1.2.1.1.1.1 9-1.1.2.1.1.1.1 11-1.1.2.1.1.2 13-1.1.2.2.1.1.1 14-1.1.2 15-1.1.2.3.1.1.1 16-1.1.2.3.1.1.2 17-1.1.2.1 17-1.1.2.2 17-1.1.2.3 20-1.1.3 22-1.1.2 24-1.2.2.2 25-1.2.2 28-1.2.2.1.1.1.1 30-1.2.2.1.2.1.1 31-1.2 32-1.2.1.2.1 34-1.2.1.2 35-1.2.1 36-1.2.1.1.r 37-1.2.1.1.1 38-1.2.1.1 (a / and :op1 (i / increase-01~e.4 :ARG0 (f / factor~e.3 :ARG1-of (d / derive-01~e.2 :ARG2 (m3 / macrophage~e.0))) :ARG1 (a2 / and~e.14,22 :op1 (l / level~e.17 :quant-of (e4 / enzyme :name (n / name :op1 "Erk1/2"~e.7,9) :ARG3-of (p / phosphorylate-01~e.5,11))) :op2 (l2 / level~e.17 :quant-of (e5 / enzyme :name (n2 / name :op1 "Akt"~e.13) :ARG3-of p)) :op3 (l3 / level~e.17 :quant-of (p3 / protein :name (n3 / name :op1 "cyclin"~e.15 :op2 "D1"~e.16)))) :location (c / cell~e.20 :mod (t / tumor))) :op2 (a3 / ablate-01~e.31 :ARG1 (i3 / increase-01~e.35 :ARG1~e.36 (g / grow-01~e.38 :ARG1 (e3 / epithelium~e.37)) :ARG1-of (m / mediate-01~e.34 :ARG0 m3~e.32)) :ARG3 (i2 / inhibit-01~e.25 :ARG1 (a4 / and :op1 (e / enzyme :name (n5 / name :op1 "MEK"~e.28)) :op2 (e2 / enzyme :name (n6 / name :op1 "PI3K"~e.30))) :ARG1-of (c2 / combine-01~e.24)))) # ::id a_pmid_2169_9731.60 ::amr-annotator SDL-AMR-09 ::preferred # ::tok Macrophage conditioned media profoundly stimulates the anchorage @-@ independent growth of lung tumor cells # ::alignments 0-1.1.1.1 1-1.1.1 2-1.1 3-1.3 3-1.3.r 4-1 6-1.2.2.2 8-1.2.2 8-1.2.2.1 8-1.2.2.1.r 9-1.2 10-1.2.1.r 11-1.2.1.1.1 12-1.2.1.1 13-1.2.1 (s / stimulate-01~e.4 :ARG0 (m / medium~e.2 :ARG1-of (c2 / condition-01~e.1 :ARG0 (m2 / macrophage~e.0))) :ARG1 (g / grow-01~e.9 :ARG1~e.10 (c / cell~e.13 :mod (t / tumor~e.12 :mod (l / lung~e.11))) :ARG0-of (d / depend-01~e.8 :polarity~e.8 -~e.8 :ARG1 (a / anchorage~e.6))) :manner~e.3 (p / profound~e.3)) # ::id a_pmid_2169_9731.61 ::amr-annotator SDL-AMR-09 ::preferred # ::tok Despite the correlation between lung macrophage content and lung tumor growth , the direct contribution of alveolar macrophages to lung tumor growth is unclear [ @ 6 , 7 , 13 @ ] . # ::alignments 0-1.4.r 2-1.4 4-1.4.1.1.1 5-1.4.1.1 6-1.4.1 7-1.3.1.1.1 8-1.2.2.1.1 9-1.2.2.1 10-1.2.2 13-1.2.3 14-1.2 15-1.2.1.r 16-1.2.1.1 17-1.2.1 19-1.2.2.1.1 20-1.2.2.1 21-1.2.2 23-1 23-1.1 23-1.1.r 26-1.3.1.1.1.1 30-1.3.1.1.1.2 34-1.3.1.1.1.3 (c / clear-06~e.23 :polarity~e.23 -~e.23 :ARG1 (c2 / contribute-01~e.14 :ARG0~e.15 (m / macrophage~e.17 :mod (a / alveolus~e.16)) :ARG2 (g / grow-01~e.10,21 :ARG1 (t / tumor~e.9,20 :mod (l / lung~e.8,19))) :ARG1-of (d / direct-02~e.13)) :ARG1-of (d2 / describe-01 :ARG0 (p / publication :ARG1-of (c5 / cite-01 :ARG2 (a2 / and~e.7 :op1 6~e.26 :op2 7~e.30 :op3 13~e.34)))) :concession~e.0 (c3 / correlate-01~e.2 :ARG1 (c4 / content~e.6 :mod (m2 / macrophage~e.5 :mod (l2 / lung~e.4))) :ARG2 g)) # ::id a_pmid_2169_9731.62 ::amr-annotator SDL-AMR-09 ::preferred # ::tok Media conditioned by an immortalized lung macrophage cell line , MH @-@ S , has been previously reported to stimulate the migration of lung epithelial cells harboring Kras @ mutations [ @ 18 @ ] . # ::alignments 0-1.1.1 1-1.1.1.1 2-1.1.1.1.1.r 4-1.1.1.1.1.1 5-1.1.1.1.1.3.1 6-1.1.1.1.1.3 7-1.1.1.1.1 8-1.1.1.1.1 10-1.1.1.1.1.2.1.1.1 12-1.1.1.1.1.2.1.1.1 16-1.2 17-1 19-1.1 21-1.1.2 22-1.1.2.1.r 23-1.1.2.1.3 24-1.1.2.1.1 25-1.1.2.1 26-1.1.2.1.2 28-1.1.2.1.2.1.1.1.1 30-1.1.2.1.2.1 33-1.3.1.1.1 (r / report-01~e.17 :ARG1 (s / stimulate-01~e.19 :ARG0 (m3 / medium~e.0 :ARG1-of (c2 / condition-01~e.1 :ARG0~e.2 (c6 / cell-line~e.7,8 :ARG1-of (i / immortalize-03~e.4) :ARG1-of (m5 / mean-01 :ARG2 (c4 / cell-line :name (n2 / name :op1 "MH-S"~e.10,12))) :mod (m4 / macrophage~e.6 :mod (l / lung~e.5))))) :ARG1 (m / migrate-01~e.21 :ARG0~e.22 (c / cell~e.25 :mod (e / epithelium~e.24) :ARG0-of (h / harbor-01~e.26 :ARG1 (m2 / mutate-01~e.30 :ARG1 (g / gene :name (n / name :op1 "Kras"~e.28)))) :mod l~e.23))) :time (p / previous~e.16) :ARG1-of (d / describe-01 :ARG0 (p2 / publication :ARG1-of (c5 / cite-01 :ARG2 18~e.33)))) # ::id a_pmid_2169_9731.63 ::amr-annotator SDL-AMR-09 ::preferred # ::tok To determine if MH @-@ S conditioned media directly stimulates neoplastic growth , we first evaluated neoplastic colony formation and cell number after long @-@ term conditioned media exposure . # ::alignments 1-1.5 3-1.5.2.1.2.1.1 5-1.5.2.1.2.1.1 6-1.5.2.1.1 7-1.5.2.1 8-1.5.2.3 9-1.5.2 11-1.5.2.2 13-1.1 14-1.3 14-1.3.1 14-1.3.1.r 15-1 17-1.2.1.1 18-1.2.1 19-1.2 20-1.2.2.1 21-1.2.2 22-1.4 23-1.4.1.2 23-1.4.1.2.r 25-1.4.1.2.r 26-1.4.1.1.1 27-1.4.1.1 28-1.4.1 (e / evaluate-01~e.15 :ARG0 (w / we~e.13) :ARG1 (a / and~e.19 :op1 (f / form-01~e.18 :ARG1 (c / colony~e.17 :mod (t / tumor))) :op2 (n2 / number~e.21 :quant-of (c2 / cell~e.20))) :ord (o / ordinal-entity~e.14 :value~e.14 1~e.14) :time (a2 / after~e.22 :op1 (e2 / expose-01~e.28 :ARG2 (m / medium~e.27 :ARG1-of (c3 / condition-01~e.26)) :duration~e.23,25 (l / long-03~e.23))) :purpose (d / determine-01~e.1 :ARG0 w :ARG1 (s / stimulate-01~e.9 :ARG0 (m2 / medium~e.7 :ARG1-of (c4 / condition-01~e.6) :mod (c5 / cell-line :name (n4 / name :op1 "MH-S"~e.3,5))) :ARG1 (g / grow-01~e.11 :mod t) :ARG1-of (d2 / direct-02~e.8)))) # ::id a_pmid_2169_9731.64 ::amr-annotator SDL-AMR-09 ::preferred # ::tok In both the classic model of anchorage @-@ independent neoplastic growth on soft agar ( Figure 1A @-@ C ) , and colonization on new ultra @-@ low adherence , neutrally @-@ charged plastic ( Figure 1D @-@ F ) , macrophage @-@ conditioned media potently stimulated the proliferation of two Kras @ mutant lung tumor @-@ derived cell lines ( LM2 and JF32 ) . # ::alignments 3-1.4.1.1 4-1.4.1 5-1.4.1.2.r 6-1.4.1.2.2.2 8-1.4.1.2.2 8-1.4.1.2.2.1 8-1.4.1.2.2.1.r 10-1.4.1.2 11-1.4.1.2.1.r 12-1.4.1.2.1.1 13-1.4.1.2.1 15-1.4.1.3.1.1 15-1.4.1.3.1.2 15-1.4.1.3.1.3 15-1.4.2.3.1.2 15-1.4.2.3.1.3 17-1.4.1.3.1.1.1 23-1.4 23-1.4.1.3.1 24-1.4.2 25-1.4.2.2.r 26-1.4.2.2.2.1 27-1.4.2.2.2.2.1 29-1.4.2.2.2.2 34-1.4.2.2.1 35-1.4.2.2 37-1.4.2.3.1.1 39-1.4.2.3.1.1.1 45-1.1.1.1 47-1.1.1 48-1.1 49-1.3 49-1.3.r 50-1 52-1.2 53-1.2.1.r 54-1.2.1.1 56-1.2.1.5.1.1 58-1.2.1.2 59-1.2.1.4.1.1 60-1.2.1.4.1 60-1.4.1.2.3 62-1.2.1.4 63-1.2.1 64-1.2.1 66-1.2.1.3.1.1.1.1 67-1.2.1.3.1 68-1.2.1.3.1.2.1.1 (s / stimulate-01~e.50 :ARG0 (m / medium~e.48 :ARG1-of (c / condition-01~e.47 :ARG0 (m2 / macrophage~e.45))) :ARG1 (p2 / proliferate-01~e.52 :ARG0~e.53 (c2 / cell-line~e.63,64 :quant 2~e.54 :ARG1-of (m3 / mutate-01~e.58) :ARG2-of (m5 / mean-01 :ARG1 (a / and~e.67 :op1 (c3 / cell-line :name (n2 / name :op1 "LM2"~e.66)) :op2 (c4 / cell-line :name (n3 / name :op1 "JF32"~e.68)))) :ARG1-of (d / derive-01~e.62 :ARG2 (t / tumor~e.60 :mod (l / lung~e.59))) :mod (e / enzyme :name (n / name :op1 "Kras"~e.56)))) :manner~e.49 (p / potent~e.49) :location (a2 / and~e.23 :op1 (m4 / model~e.4 :mod (c5 / classic~e.3) :topic~e.5 (g / grow-01~e.10 :location~e.11 (a3 / agar~e.13 :ARG1-of (s2 / soft-02~e.12)) :ARG0-of (d2 / depend-01~e.8 :polarity~e.8 -~e.8 :ARG1 (a4 / anchorage~e.6)) :mod (t2 / tumor~e.60)) :ARG1-of (d3 / describe-01 :ARG0 (a7 / and~e.23 :op1 (f / figure~e.15 :mod "1A"~e.17) :op2 (f5 / figure~e.15 :mod "1B") :op3 (f6 / figure~e.15 :mod "1C")))) :op2 (c6 / colonize-01~e.24 :ARG0 c2 :location~e.25 (p3 / plastic~e.35 :ARG1-of (c7 / charge-03~e.34 :ARG0-of (n4 / neutral-02)) :ARG1-of (a5 / adhere-01 :ARG1-of (n5 / new-01~e.26) :ARG1-of (l2 / low-04~e.29 :degree (u / ultra~e.27)))) :ARG1-of (d4 / describe-01 :ARG0 (a6 / and :op1 (f2 / figure~e.37 :mod "1D"~e.39) :op2 (f3 / figure~e.15 :mod "1E") :op3 (f4 / figure~e.15 :mod "1F")))))) # ::id a_pmid_2169_9731.65 ::amr-annotator SDL-AMR-09 ::preferred # ::tok Thus , macrophages secrete soluble molecules capable of greatly stimulating neoplastic colony formation and proliferation in vitro , which may shed light on the role of macrophage recruitment to lung cancer in vivo @ . # ::alignments 0-1 2-1.1.1 3-1.1 5-1.1.2 6-1.1.2.1 7-1.1.2.1.1.r 8-1.1.2.1.1.4 8-1.1.2.1.1.4.r 9-1.1.2.1.1 11-1.1.2.1.1.2.1.1 12-1.1.2.1.1.2.1 13-1.1.2.1.1.2 14-1.1.2.1.1.2.2 16-1.1.2.1.1.3 17-1.1.2.1.1.3 21-1.1.2.2.1 22-1.1.3 23-1.1.3.1 24-1.1.3.2.r 26-1.1.3.2 27-1.1.3.2.1.r 28-1.1.3.2.1.1 29-1.1.3.2.1 30-1.1.3.2.1.2.r 31-1.1.3.2.1.2.2.1 32-1.1.3.2.1.2.2.2 34-1.1.3.2.1.3 35-1.1.3.2.1.3 (c / cause-01~e.0 :ARG1 (s / secrete-01~e.3 :ARG0 (m / macrophage~e.2) :ARG1 (m2 / molecule~e.5 :ARG1-of (c2 / capable-01~e.6 :ARG2~e.7 (s2 / stimulate-01~e.9 :ARG0 m2 :ARG1 (a / and~e.13 :op1 (f / form-01~e.12 :ARG1 (c3 / colony~e.11 :mod (t / tumor))) :op2 (p / proliferate-01~e.14 :ARG0 c3)) :manner (i / in-vitro~e.16,17) :manner~e.8 (g / great~e.8))) :ARG1-of (d / dissolve-01 :ARG1-of (p3 / possible-01~e.21))) :ARG0-of (s3 / shed-03~e.22 :ARG1 (l / light~e.23) :ARG2~e.24 (r / role~e.26 :topic~e.27 (r2 / recruit-01~e.29 :ARG1 m~e.28 :ARG2~e.30 (d2 / disease :wiki "Lung_cancer" :name (n / name :op1 "lung"~e.31 :op2 "cancer"~e.32)) :manner (i2 / in-vivo~e.34,35))) :ARG1-of p3))) # ::id a_pmid_2169_9731.66 ::amr-annotator SDL-AMR-09 ::preferred # ::tok Naïve and tumor @-@ educated primary macrophage co @-@ culture stimulates the proliferation of neoplastic and non @-@ neoplastic pulmonary epithelial cells # ::alignments 0-1.1.1.1 1-1.1 2-1.1.2.2.1.1 2-1.2.1.2.3 4-1.1.2.2.1 5-1.1.1.2 6-1.1.1.3 6-1.1.2.2 7-1.1.1 7-1.1.2 9-1.1.1 10-1 12-1.2 15-1.2.1 16-1.2.1.2.3.1 16-1.2.1.2.3.1.r 20-1.2.1.1.1 21-1.2.1.1 21-1.2.1.2 (s / stimulate-01~e.10 :ARG0 (a / and~e.1 :op1 (c / co-culture~e.7,9 :mod (n / naive~e.0) :mod (p / primary~e.5) :mod (m / macrophage~e.6)) :op2 (c2 / co-culture~e.7 :mod p :mod (m2 / macrophage~e.6 :ARG1-of (e / educate-01~e.4 :ARG0 (t / tumor~e.2))))) :ARG1 (p2 / proliferate-01~e.12 :ARG0 (a2 / and~e.15 :op1 (c3 / cell~e.21 :mod (e2 / epithelium~e.20) :mod (l / lung) :mod t) :op2 (c4 / cell~e.21 :mod e2 :mod l :mod (t2 / tumor~e.2 :polarity~e.16 -~e.16))))) # ::id a_pmid_2169_9731.67 ::amr-annotator SDL-AMR-09 ::preferred # ::tok The relative ability of naïve vs. tumor @-@ educated alveolar macrophages to directly stimulate lung epithelial cell proliferation not been reported . # ::alignments 1-1.2.3 2-1.2 4-1.2.1.1 5-1.2.4.r 6-1.2.4.1.1 8-1.2.4.1 9-1.2.1.2 10-1.2.1 10-1.2.4 12-1.2.2.3 13-1.2.2 14-1.2.2.2.1.2 15-1.2.2.2.1.1 16-1.2.2.2.1 17-1.2.2.2 18-1.1 18-1.1.r 20-1 (r / report-01~e.20 :polarity~e.18 -~e.18 :ARG1 (c2 / capable-01~e.2 :ARG1 (m / macrophage~e.10 :mod (n / naive~e.4) :mod (a / alveolus~e.9)) :ARG2 (s / stimulate-01~e.13 :ARG0 m :ARG1 (p2 / proliferate-01~e.17 :ARG0 (c / cell~e.16 :mod (e2 / epithelium~e.15) :mod (l / lung~e.14))) :ARG1-of (d / direct-02~e.12)) :ARG2-of (r2 / relative-05~e.1) :compared-to~e.5 (m2 / macrophage~e.10 :ARG1-of (e / educate-01~e.8 :ARG0 (t / tumor~e.6)) :mod a))) # ::id a_pmid_2169_9731.68 ::amr-annotator SDL-AMR-09 ::preferred # ::tok To determine if macrophages from the lungs of tumor @-@ bearing mice could directly stimulate neoplastic cell growth in a co @-@ culture system , neoplastic LM2 cells were co @-@ cultured with bronchoalveolar lavage ( BAL ) macrophages ( MØ) isolated from tumor @-@ bearing mice , and monolayer growth was assessed ( Figure 2A ) . # ::alignments 1-1.3 3-1.3.1.1.1 4-1.3.1.1.1.1.r 6-1.3.1.1.1.1 7-1.3.1.1.1.1.1.r 8-1.3.1.1.1.1.1 9-1.3.1.1.1.1.1 10-1.3.1.1.1.1.1 11-1.3.1.1.1.1.1 12-1.3.1 12-1.3.1.1.3 13-1.3.1.1.4 14-1.3.1.1 15-1.3.1.1.2.1.1 16-1.3.1.1.2.1 17-1.3.1.1.2 18-1.3.1.1.2.2.r 20-1.3.1.1.2.2.1 22-1.3.1.1.2.2.1 23-1.3.1.1.2.2 25-1.1.1.1.2 26-1.1.1.1.1.1 27-1.1.1.1 29-1.3.1.1.2.2.1 31-1.1 31-1.3.1.1.2.2.1 32-1.1.1.r 33-1.1.1.2.1 34-1.1.1.2.1.1 36-1.1.1.2.1.1 38-1.1.1.2 41-1.1.1.2.2 42-1.3.1.1.1.1.r 43-1.3.1.1.1.1.1 44-1.3.1.1.1.1.1 45-1.3.1.1.1.1.1 46-1.3.1.1.1.1.1 48-1 48-1.1.1 50-1.2.1 52-1.2 54-1.4.1 56-1.4.1.1 (a2 / and~e.48 :op1 (c / culture-01~e.31 :ARG1~e.32 (a / and~e.48 :op1 (c2 / cell-line~e.27 :name (n / name :op1 "LM2"~e.26) :mod (n2 / neoplastic~e.25)) :op2 (m / macrophage~e.38 :mod (b / bronchoalveolar~e.33 :mod (l / lavage~e.34,36)) :ARG1-of (i / isolate-01~e.41 :ARG2 (m2 / mouse :ARG0-of (b2 / bear-01 :ARG1 (t / tumor))))))) :op2 (a3 / assess-01~e.52 :ARG1 (g / grow-01~e.50 :ARG1 (l2 / layer :quant 1))) :purpose (d / determine-01~e.1 :ARG1 (p2 / possible-01~e.12 :ARG1 (s / stimulate-01~e.14 :ARG0 (m3 / macrophage~e.3 :source~e.4,42 (l3 / lung~e.6 :part-of~e.7 m2~e.8,9,10,11,43,44,45,46)) :ARG1 (g2 / grow-01~e.17 :mod (c3 / cell~e.16 :mod (n3 / neoplastic~e.15)) :location~e.18 (s2 / system~e.23 :mod (c4 / co-culture~e.20,22,29,31))) :ARG1-of (p / possible-01~e.12) :ARG1-of (d3 / direct-02~e.13)))) :ARG1-of (d2 / describe-01 :ARG0 (f / figure~e.54 :mod "2A"~e.56))) # ::id a_pmid_2169_9731.69 ::amr-annotator SDL-AMR-09 ::preferred # ::tok Growth in standard tissue culture conditions measures proliferation per se , and not cell motility or the requirement for solid support , and permits the evaluation of non @-@ neoplastic epithelial cells which do not proliferate in anchorage @-@ independent systems . # ::alignments 0-1.1.1.1 1-1.1.1.1.1.r 2-1.1.1.1.1.1.1 3-1.1.1.1.1.1 4-1.1.1.1.1 6-1.1.1 6-1.1.2 7-1.1.1.2 9-1.1.1.3 10-1.1.1.3 14-1.1.2.1 14-1.1.2.1.r 15-1.1.2.3.1.1 16-1.1.2.3.1 17-1.1.2.3 19-1.1.2.3.2 20-1.1.2.3.2.1.r 21-1.1.2.3.2.1.1 22-1.1.2.3.2.1 24-1 25-1.2 27-1.2.2 28-1.2.2.1.r 29-1.2.2.1.2.1 29-1.2.2.1.2.1.r 32-1.2.2.1.1 33-1.2.2.1 36-1.2.2.1.2.1 36-1.2.2.1.2.1.r 36-1.2.2.1.3.1 36-1.2.2.1.3.1.r 37-1.2.2.1.3 38-1.2.2.1.3.2.r 39-1.2.2.1.3.2.1.2 41-1.2.2.1.3.2.1 41-1.2.2.1.3.2.1.1 41-1.2.2.1.3.2.1.1.r 42-1.2.2.1.3.2 (a / and~e.24 :op1 (c2 / contrast-01 :ARG1 (m / measure-01~e.6 :ARG0 (g / grow-01~e.0 :ARG1~e.1 (c / culture-01~e.4 :ARG1 (t / tissue~e.3 :ARG1-of (s / standard-02~e.2)))) :ARG1 (p4 / proliferate-01~e.7) :manner (p / per-se~e.9,10)) :ARG2 (m2 / measure-01~e.6 :polarity~e.14 -~e.14 :ARG0 g :ARG1 (o / or~e.17 :op1 (m3 / motility~e.16 :mod (c3 / cell~e.15)) :op2 (r / require-01~e.19 :ARG1~e.20 (s2 / support-01~e.22 :ARG1-of (s3 / solid-02~e.21)))))) :op2 (p2 / permit-01~e.25 :ARG0 g :ARG1 (e / evaluate-01~e.27 :ARG1~e.28 (c4 / cell~e.33 :mod (e2 / epithelium~e.32) :mod (t2 / tumor :polarity~e.29,36 -~e.29,36) :ARG0-of (p3 / proliferate-01~e.37 :polarity~e.36 -~e.36 :location~e.38 (s4 / system~e.42 :ARG0-of (d / depend-01~e.41 :polarity~e.41 -~e.41 :ARG1 (a2 / anchorage~e.39)))))))) # ::id a_pmid_2169_9731.70 ::amr-annotator SDL-AMR-09 ::preferred # ::tok LM2 cell number significantly increased with BAL macrophage co @-@ culture at 48 ( 2.3 vs. 4.1 @-@ fold ) and 72 hrs ( 3.5 vs. 7.5 @-@ fold ) ( Figure 2A ) . # ::alignments 0-1.1.2.1.1.1 1-1.1.2.1 2-1.1.2 3-1.1.3 4-1.1 4-1.2 5-1.1.1.r 6-1.1.1.1.1 6-1.1.1.1.1.1 6-1.1.1.1.1.1.1 6-1.1.1.1.1.1.1.r 6-1.1.1.1.1.1.r 7-1.1.1.1 8-1.1.1 10-1.1.1 12-1.1.6.1.1 14-1.1.4.1 16-1.1.5.1 18-1.1.4 18-1.1.5 20-1 21-1.2.6.1.1 22-1.1.6.1.2 22-1.2.6.1.2 24-1.2.4.1 26-1.2.5.1 28-1.2.4 28-1.2.5 31-1.3.1 33-1.3.1.1 (a / and~e.20 :op1 (i / increase-01~e.4 :ARG0~e.5 (c / co-culture~e.8,10 :mod (m / macrophage~e.7 :mod (l / lavage~e.6 :mod~e.6 (a4 / alveolus~e.6 :mod~e.6 (b / bronchus~e.6))))) :ARG1 (n2 / number~e.2 :quant-of (c2 / cell-line~e.1 :name (n3 / name :op1 "LM2"~e.0))) :ARG2 (s / significant-02~e.3) :ARG3 (p / product-of~e.18 :op1 2.3~e.14) :ARG4 (p2 / product-of~e.18 :op1 4.1~e.16) :time (a2 / after :op1 (t2 / temporal-quantity :quant 48~e.12 :unit (h / hour~e.22)))) :op2 (i2 / increase-01~e.4 :ARG0 c :ARG1 n2 :ARG2 s :ARG3 (p3 / product-of~e.28 :op1 3.5~e.24) :ARG4 (p4 / product-of~e.28 :op1 7.5~e.26) :time (a3 / after :op1 (t3 / temporal-quantity :quant 72~e.21 :unit (h2 / hour~e.22)))) :ARG1-of (d / describe-01 :ARG0 (f / figure~e.31 :mod "2A"~e.33))) # ::id a_pmid_2169_9731.71 ::amr-annotator SDL-AMR-09 ::preferred # ::tok As 72 hrs of macrophage co @-@ culture resulted in ≥ 2 @-@ times more tumor cells , this time point was used in subsequent experiments . # ::alignments 0-1.2.1.r 1-1.3.1.1.2.1 2-1.3.1.1.2.2 4-1.3.1.1.1 5-1.3.1.1 7-1.3.1.1 8-1.3.1 11-1.3.1.2.2.1.1.1.1 11-1.3.1.2.2.1.2.1 13-1.3.1.1.2 13-1.3.1.2.2.1.2 14-1.3.1.2.2 14-1.3.1.2.2.1.1 15-1.3.1.2.1 16-1.3.1.2 18-1.1.2 19-1.1.1 20-1.1 22-1 24-1.2.1 24-1.2.1.r 25-1.2 (u / use-01~e.22 :ARG0 (p / point~e.20 :mod (t / time~e.19) :mod (t4 / this~e.18)) :ARG1 (e / experiment-01~e.25 :time~e.0,24 (s / subsequent~e.24)) :ARG1-of (c / cause-01 :ARG0 (r / result-01~e.8 :ARG1 (c2 / co-culture~e.5,7 :mod (m / macrophage~e.4) :duration (t2 / temporal-quantity~e.13 :quant 72~e.1 :unit (h / hour~e.2))) :ARG2 (c3 / cell~e.16 :mod (t3 / tumor~e.15) :quant (m2 / more~e.14 :degree (v / value-interval :op1 (m3 / more-than~e.14 :op1 (p2 / product-of :op1 2~e.11)) :op2 (p3 / product-of~e.13 :op1 2~e.11))))))) # ::id a_pmid_2169_9731.72 ::amr-annotator SDL-AMR-09 ::preferred # ::tok To determine if tumor @-@ educated macrophages stimulated neoplastic growth more effectively than naïve , BAL macrophages from either naïve or tumor @-@ bearing mice were co @-@ cultured with neoplastic LM2 ( Figure 2B ) and JF32 ( Figure 2C ) cells . # ::alignments 1-1.2 3-1.2.1.2.1.1 5-1.2.1.2.1 6-1.2.1.2 7-1.2.1 9-1.2.1.3 10-1.2.1.4.1 11-1.2.1.4 12-1.2.1.4.2.r 13-1.2.1.4.2.1 15-1.1.1.2 15-1.1.1.2.1 15-1.1.1.2.1.1 15-1.1.1.2.1.1.r 15-1.1.1.2.1.r 16-1.1.1 16-1.2.1.4.2 17-1.1.1.1.r 19-1.1.1.1.1.1 20-1.1.1.1 21-1.1.1.1.2.1.1 23-1.1.1.1.2.1 24-1.1.1.1.1 24-1.1.1.1.2 28-1 31-1.1.2.1.1 33-1.1.2.2.1 35-1.1.2.2.1.1 38-1.1 39-1.1.3.1.1 41-1.1.3.2.1 43-1.1.3.2.1.1 46-1.1.2 46-1.1.3 (c / culture-01~e.28 :ARG1 (a / and~e.38 :op1 (m / macrophage~e.16 :source~e.17 (o / or~e.20 :op1 (m2 / mouse~e.24 :mod (n / naive~e.19)) :op2 (m3 / mouse~e.24 :ARG0-of (b / bear-01~e.23 :ARG1 (t / tumor~e.21)))) :mod (l / lavage~e.15 :mod~e.15 (a2 / alveolus~e.15 :mod~e.15 (b2 / bronchus~e.15)))) :op2 (c2 / cell-line~e.46 :name (n2 / name :op1 "LM2"~e.31) :ARG1-of (d / describe-01 :ARG0 (f / figure~e.33 :mod "2B"~e.35)) :mod t) :op3 (c3 / cell-line~e.46 :name (n3 / name :op1 "JF32"~e.39) :ARG1-of (d2 / describe-01 :ARG0 (f2 / figure~e.41 :mod "2C"~e.43)) :mod t)) :purpose (d3 / determine-01~e.1 :ARG1 (s / stimulate-01~e.7 :mode interrogative :ARG0 (m4 / macrophage~e.6 :ARG1-of (e / educate-01~e.5 :ARG0 t~e.3)) :ARG1 (g / grow-01~e.9 :ARG1 t) :ARG1-of (e2 / effective-04~e.11 :degree (m5 / more~e.10) :compared-to~e.12 (m6 / macrophage~e.16 :mod (n6 / naive~e.13)))))) # ::id a_pmid_2169_9731.73 ::amr-annotator SDL-AMR-09 ::preferred # ::tok LM2 growth was equally stimulated by both naïve and tumor @-@ educated BAL macrophages , while the growth of JF32 cells was enhanced slightly upon co @-@ culture with tumor @-@ educated BAL macrophages ( Figure 2C ) . # ::alignments 0-1.1.1.1.1.1 1-1.1.1 3-1.1.3 4-1.1 7-1.1.2.1.1 8-1.1.2 9-1.1.2.2.1.1 11-1.1.2.2.1 12-1.1.2.1.2 12-1.1.2.1.2.1 12-1.1.2.1.2.1.1 12-1.1.2.1.2.1.1.r 12-1.1.2.1.2.1.r 13-1.1.2.1 13-1.1.2.2 15-1 17-1.1.1 17-1.2.1 18-1.2.r 19-1.2.1.1.1.1 20-1.1.1.1 20-1.2.1.1 22-1.2 23-1.2.2 23-1.2.2.r 27-1.2.3.1 28-1.2.3.1.1.r 29-1.2.3.1.1.2 30-1.2.3.1.1.2 31-1.2.3.1.1.2 32-1.1.2.1.2 32-1.1.2.1.2.1 32-1.1.2.1.2.1.1 32-1.1.2.1.2.1.1.r 32-1.1.2.1.2.1.r 33-1.1.2.1 35-1.3.1 37-1.3.1.1 (c / contrast-01~e.15 :ARG1 (s / stimulate-01~e.4 :ARG0 (g / grow-01~e.1,17 :ARG1 (c2 / cell-line~e.20 :name (n2 / name :op1 "LM2"~e.0))) :ARG1 (a / and~e.8 :op1 (m / macrophage~e.13,33 :mod (n / naive~e.7) :mod (l / lavage~e.12,32 :mod~e.12,32 (a4 / alveolus~e.12,32 :mod~e.12,32 (b / bronchus~e.12,32)))) :op2 (m2 / macrophage~e.13 :ARG1-of (e / educate-01~e.11 :ARG0 (t / tumor~e.9)))) :ARG1-of (e2 / equal-01~e.3)) :ARG2~e.18 (e3 / enhance-01~e.22 :ARG1 (g2 / grow-01~e.17 :ARG1 (c3 / cell-line~e.20 :name (n3 / name :op1 "JF32"~e.19))) :manner~e.23 (s2 / slight~e.23) :time (a3 / after :op1 (c4 / culture-01~e.27 :ARG1~e.28 (a2 / and :op1 c3 :op2 m2~e.29,30,31)))) :ARG1-of (d / describe-01 :ARG0 (f / figure~e.35 :mod "2C"~e.37))) # ::id a_pmid_2169_9731.74 ::amr-annotator SDL-AMR-09 ::preferred # ::tok To determine if primary alveolar macrophages also stimulated the proliferation of non @-@ tumor cells , the non @-@ neoplastic E10 cell line was co @-@ cultured with naïve and tumor @-@ educated BAL macrophages . # ::alignments 1-1.2 3-1.2.1.2.1 4-1.2.1.2.2 5-1.2.1.2 6-1.2.1.4 7-1.2.1 9-1.2.1.3 11-1.1.1.2.1 11-1.1.1.2.1.r 13-1.1.1.2 14-1.2.1.3.1 17-1.1.1.2.1.r 20-1.1.1.1.1 21-1.1.1 22-1.1.1 26-1 28-1.1.2.1 29-1.1 30-1.1.3.1.1 32-1.1.3.1 33-1.1.3.2 33-1.1.3.2.1 33-1.1.3.2.1.1 33-1.1.3.2.1.1.r 33-1.1.3.2.1.r 34-1.1.2 34-1.1.3 (c / culture-01~e.26 :ARG1 (a / and~e.29 :op1 (c2 / cell-line~e.21,22 :name (n / name :op1 "E10"~e.20) :mod (t3 / tumor~e.13 :polarity~e.11,17 -~e.11)) :op2 (m / macrophage~e.34 :mod (n2 / naive~e.28)) :op3 (m2 / macrophage~e.34 :ARG1-of (e / educate-01~e.32 :ARG0 (t / tumor~e.30)) :mod (l / lavage~e.33 :mod~e.33 (a4 / alveolus~e.33 :mod~e.33 (b / bronchus~e.33))))) :purpose (d / determine-01~e.1 :ARG1 (s / stimulate-01~e.7 :mode interrogative :ARG0 (m3 / macrophage~e.5 :mod (p2 / primary~e.3) :mod (a2 / alveolus~e.4)) :ARG1 (p / proliferate-01~e.9 :ARG0 (c3 / cell~e.14 :mod t3)) :mod (a3 / also~e.6)))) # ::id a_pmid_2169_9731.75 ::amr-annotator SDL-AMR-09 ::preferred # ::tok Both macrophage types increased E10 cell number 3.5 @-@ fold ( Figure 2D ) when maintained in serum @-@ free conditions ; only tumor @-@ educated macrophages stimulated E10 proliferation when cultured in the presence of serum ( Figure 2E ) . # ::alignments 0-1.1.1.2 1-1.1.1.1 2-1.1.1 3-1.1 4-1.1.2.1.1.1 5-1.1.2.1 5-1.2.2.1 6-1.1.2 7-1.1.3.1 9-1.1.3 11-1.1.4.1 13-1.1.2.1.r 13-1.1.4.1.1 16-1.1.5.r 17-1.1.5 18-1.1.5.2.r 19-1.1.5.2.1.1 21-1.1.5.2.1 22-1.1.5.2 24-1.2.1.2 25-1.2.1.1.1 27-1.2.1.1 28-1.2.1 29-1.2 30-1.2.2.1.1.1 31-1.2.2 32-1.2.3.r 33-1.2.3 34-1.2.4.r 36-1.2.4 37-1.2.4.1.r 38-1.2.4.1 40-1.2.5.1 42-1.2.5.1.1 (m / multi-sentence :snt1 (i / increase-01~e.3 :ARG0 (t / type-03~e.2 :ARG1 (m2 / macrophage~e.1) :mod (b / both~e.0)) :ARG1 (n / number~e.6 :quant-of~e.13 (c / cell-line~e.5 :name (n2 / name :op1 "E10"~e.4))) :ARG2 (p / product-of~e.9 :op1 3.5~e.7) :ARG1-of (d / describe-01 :ARG0 (f / figure~e.11 :mod "2D"~e.13)) :time~e.16 (m3 / maintain-01~e.17 :ARG1 t :ARG2~e.18 (c2 / condition~e.22 :ARG1-of (f2 / free-04~e.21 :ARG2 (s / serum~e.19))))) :snt2 (s2 / stimulate-01~e.29 :ARG0 (m4 / macrophage~e.28 :ARG1-of (e / educate-01~e.27 :ARG0 (t2 / tumor~e.25)) :mod (o / only~e.24)) :ARG1 (p2 / proliferate-01~e.31 :ARG0 (c3 / cell-line~e.5 :name (n3 / name :op1 "E10"~e.30))) :time~e.32 (c4 / culture-01~e.33 :ARG1 c3) :condition~e.34 (p3 / present-02~e.36 :ARG1~e.37 (s3 / serum~e.38)) :ARG1-of (d2 / describe-01 :ARG0 (f3 / figure~e.40 :mod "2E"~e.42)))) # ::id a_pmid_2169_9731.76 ::amr-annotator SDL-AMR-09 ::preferred # ::tok Both types of primary macrophages equally stimulated LM2 proliferation in the presence of serum , though the magnitude was reduced when compared to serum @-@ free co @-@ culture ( data not shown ) . # ::alignments 0-1.1.2 1-1.1 2-1.1.1.r 3-1.1.1.1 4-1.1.1 5-1.3 6-1 7-1.2.1.1.1 8-1.2 9-1.4.r 11-1.4 12-1.4.1.r 13-1.4.1 15-1.6.r 17-1.6.1 19-1.6 20-1.6.2.r 21-1.6.2 22-1.6.2.2.r 23-1.6.2.2.1.1 25-1.6.2.2.1 26-1.6.2.2 28-1.6.2.2 30-1.5.1 31-1.5.1.1.1 31-1.5.1.1.1.r 32-1.5.1.1 (s / stimulate-01~e.6 :ARG0 (t / type-03~e.1 :ARG1~e.2 (m / macrophage~e.4 :mod (p3 / primary~e.3)) :mod (b / both~e.0)) :ARG1 (p / proliferate-01~e.8 :ARG0 (c / cell-line :name (n / name :op1 "LM2"~e.7))) :ARG1-of (e / equal-01~e.5) :condition~e.9 (p2 / present-02~e.11 :ARG1~e.12 (s2 / serum~e.13)) :ARG1-of (d / describe-01 :ARG0 (d3 / data~e.30 :ARG1-of (s3 / show-01~e.32 :polarity~e.31 -~e.31))) :concession~e.15 (r / reduce-01~e.19 :ARG1 (m2 / magnitude~e.17) :time~e.20 (c2 / compare-01~e.21 :ARG1 m2 :ARG2~e.22 (c3 / co-culture~e.26,28 :ARG1-of (f / free-04~e.25 :ARG2 s2~e.23))))) # ::id a_pmid_2169_9731.77 ::amr-annotator SDL-AMR-09 ::preferred # ::tok To determine if MH @-@ S macrophages could recapitulate the effects of primary alveolar macrophages in this in vitro @ model , we co @-@ cultured MH @-@ S macrophages with both neoplastic and non @-@ neoplastic lung epithelial cells . # ::alignments 1-1.3 3-1.2.1.1.1.1 5-1.2.1.1.1.1 6-1.2.1 7-1.3.2 8-1.3.2.1 10-1.3.2.1.2 11-1.3.2.1.2.1.r 12-1.3.2.1.2.1.2 13-1.3.2.1.2.1.1 14-1.3.2.1.2.1 15-1.3.2.1.3.1 16-1.3.2.1.3.2 18-1.3.2.1.3.1 19-1.3.2.1.3.1 21-1.3.2.1.3 23-1.1 26-1 27-1.2.1.1.1.1 29-1.2.1.1.1.1 30-1.2.1 33-1.2.3.2 35-1.2.3.2.1 35-1.2.3.2.1.r 37-1.2.3.2 38-1.2.2.1 39-1.2.2.2 40-1.2.2 40-1.2.3 (c / culture-01~e.26 :ARG0 (w / we~e.23) :ARG1 (a / and :op1 (m / macrophage~e.6,30 :mod (c2 / cell-line :name (n / name :op1 "MH-S"~e.3,5,27,29))) :op2 (c3 / cell~e.40 :mod (l / lung~e.38) :mod (e / epithelium~e.39) :mod (t2 / tumor)) :op3 (c4 / cell~e.40 :mod l :mod (n3 / neoplastic~e.33,37 :polarity~e.35 -~e.35) :mod t2)) :purpose (d / determine-01~e.1 :ARG0 w :ARG1 (p / possible-01~e.7 :ARG1 (r / recapitulate-01~e.8 :ARG0 m :ARG1 (a2 / affect-01~e.10 :ARG0~e.11 (m2 / macrophage~e.14 :mod (a3 / alveolar~e.13) :mod (p2 / primary~e.12))) :location (m3 / model~e.21 :mod (i / in-vitro~e.15,18,19) :mod (t / this~e.16)))))) # ::id a_pmid_2169_9731.78 ::amr-annotator SDL-AMR-09 ::preferred # ::tok MH @-@ S co @-@ culture increased the growth rate of all pulmonary epithelial cell lines similar to co @-@ culture with tumor @-@ educated BAL macrophages ( Figure 2B @-@ E ) . # ::alignments 0-1.1.1.1.1 2-1.1.1.1.1 3-1.1 5-1.1 6-1 8-1.2.1 9-1.2 10-1.2.1.1.r 11-1.2.1.1.1 13-1.2.1.1.2 14-1.2.1.1 15-1.2.1.1 16-1.2.1.1.4 18-1.1 20-1.2.1.1.4.1 21-1.2.1.1.4.1.1.r 22-1.2.1.1.4.1.1.1.1 24-1.2.1.1.4.1.1.1 25-1.2.1.1.4.1.1.2 25-1.2.1.1.4.1.1.2.1 25-1.2.1.1.4.1.1.2.1.1 25-1.2.1.1.4.1.1.2.1.1.r 25-1.2.1.1.4.1.1.2.1.r 26-1.2.1.1.4.1.1 28-1.3.1.1 28-1.3.1.2 28-1.3.1.3 28-1.3.1.4 30-1.3.1.1.1 (i / increase-01~e.6 :ARG0 (c / co-culture~e.3,5,18 :mod (c2 / cell-line :name (n / name :op1 "MH-S"~e.0,2))) :ARG1 (r / rate~e.9 :degree-of (g / grow-01~e.8 :ARG1~e.10 (c3 / cell-line~e.14,15 :mod (a / all~e.11) :mod (e / epithelium~e.13) :mod (l / lung) :ARG1-of (r2 / resemble-01~e.16 :ARG2 (c4 / culture-01~e.20 :ARG1~e.21 (m / macrophage~e.26 :ARG1-of (e2 / educate-01~e.24 :ARG0 (t / tumor~e.22)) :mod (l2 / lavage~e.25 :mod~e.25 (a3 / alveolus~e.25 :mod~e.25 (b / bronchus~e.25))))))))) :ARG1-of (d / describe-01 :ARG0 (a2 / and :op1 (f / figure~e.28 :mod "2B"~e.30) :op2 (f2 / figure~e.28 :mod "2C") :op3 (f3 / figure~e.28 :mod "2D") :op4 (f4 / figure~e.28 :mod "2E")))) # ::id a_pmid_2169_9731.79 ::amr-annotator SDL-AMR-09 ::preferred # ::tok These results indicate that primary lung macrophages produce diffusible signals which can augment the proliferation of both non @-@ neoplastic and neoplastic cells in vitro @ . # ::alignments 0-1.1.2 1-1.1 1-1.1.1 1-1.1.1.r 2-1 3-1.2.r 4-1.2.1.1 5-1.2.1.2 6-1.2.1 7-1.2 9-1.2.2 11-1.2.2.1.1 12-1.2.2.2 14-1.2.2.2.1 17-1.2.2.2.1.1.1.1.1 17-1.2.2.2.1.1.1.1.1.r 19-1.2.2.2.1.1.2.1 21-1.2.2.2.1.1.2.1 22-1.2.2.2.1.1.1 22-1.2.2.2.1.1.2 24-1.2.2.2.1.2 25-1.2.2.2.1.2 (i / indicate-01~e.2 :ARG0 (t / thing~e.1 :ARG2-of~e.1 (r / result-01~e.1) :mod (t2 / this~e.0)) :ARG1~e.3 (p / produce-01~e.7 :ARG0 (m / macrophage~e.6 :mod (p4 / primary~e.4) :mod (l / lung~e.5)) :ARG1 (s / signal-07~e.9 :ARG1-of (d / diffuse-01 :ARG1-of (p2 / possible-01~e.11)) :ARG0-of (a / augment-01~e.12 :ARG1 (p3 / proliferate-01~e.14 :ARG0 (a2 / and :op1 (c / cell~e.22 :mod (t3 / tumor :polarity~e.17 -~e.17)) :op2 (c2 / cell~e.22 :mod (n2 / neoplastic~e.19,21))) :manner (i2 / in-vitro~e.24,25)) :ARG1-of p2)))) # ::id a_pmid_2169_9731.80 ::amr-annotator SDL-AMR-09 ::preferred # ::tok Further , we observed that in vivo @ tumor education of primary lung macrophages slightly enhances this ability to stimulate epithelial proliferation , an effect similar to co @-@ culture with MH @-@ S macrophages . # ::alignments 0-1.3 2-1.1 3-1 6-1.2.1.3 7-1.2.1.3 9-1.2.1.1 10-1.2.1 12-1.2.1.2.2 13-1.2.1.2.1 14-1.2.1.2 15-1.2.2 15-1.2.2.r 16-1.2 18-1.2.3 20-1.2.3.1 21-1.2.3.1.1.1 22-1.2.3.1.1 25-1.2.4 26-1.2.4.2 30-1.2.4.2.1 32-1.2.4.2.1.1.2.1.1.1 34-1.2.4.2.1.1.2.1.1.1 35-1.2.1.2 35-1.2.4.2.1.1.2 (o / observe-01~e.3 :ARG0 (w / we~e.2) :ARG1 (e / enhance-01~e.16 :ARG0 (e2 / educate-01~e.10 :ARG0 (t / tumor~e.9) :ARG1 (m / macrophage~e.14,35 :mod (l / lung~e.13) :mod (p / primary~e.12)) :manner (i / in-vivo~e.6,7)) :manner~e.15 (s / slight~e.15) :ARG1-of (c / capable-01~e.18 :ARG2 (s2 / stimulate-01~e.20 :ARG1 (p2 / proliferate-01~e.22 :ARG0 (e3 / epithelium~e.21)))) :ARG2-of (a / affect-01~e.25 :ARG1 c :ARG1-of (r / resemble-01~e.26 :ARG2 (c2 / culture-01~e.30 :ARG1 (a2 / and :op1 m :op2 (m2 / macrophage~e.35 :mod (c3 / cell-line :name (n / name :op1 "MH-S"~e.32,34)))))))) :mod (f / further~e.0)) # ::id a_pmid_2169_9731.81 ::amr-annotator SDL-AMR-09 ::preferred # ::tok Macrophage co @-@ culture stimulates epithelial proliferation through kinase activation # ::alignments 0-1.1.1 1-1.1 2-1.1 3-1.1 4-1 5-1.2.1 6-1.2 8-1.3.1 9-1.3 (s / stimulate-01~e.4 :ARG0 (c / coculture-01~e.1,2,3 :ARG1 (m / macrophage~e.0)) :ARG1 (p / proliferate-01~e.6 :ARG0 (e / epithelium~e.5)) :ARG2 (a / activate-01~e.9 :ARG1 (k / kinase~e.8))) # ::id a_pmid_2169_9731.82 ::amr-annotator SDL-AMR-09 ::preferred # ::tok Since MH @-@ S macrophages and tumor @-@ educated primary macrophages stimulated epithelial proliferation to a similar degree , MH @-@ S macrophages were used to elucidate the mechanisms of increased epithelial proliferation . # ::alignments 0-1.3 1-1.3.1.1.1 2-1.3.1.1.1 3-1.3.1.1.1 4-1.3.1.1.1 5-1.3.1.1 6-1.3.1.1.2.2.1 8-1.3.1.1.2.2 9-1.3.1.1.2.1 10-1.3.1.1.2 11-1.3.1 12-1.3.1.2 13-1.3.1.2 14-1.3 16-1.3.1.3 17-1.3.1.3.r 19-1.1.1.1.1 21-1.1.1.1.1 22-1.1 24-1 25-1.3 26-1.2 28-1.2.2 29-1.2.2.1.r 30-1.2.2.1.2 31-1.2.2.1.1 32-1.2.2.1 (u / use-01~e.24 :ARG1 (m2 / macrophage~e.22 :mod (c2 / cell-line :name (n / name :op1 "MH-S"~e.19,21))) :ARG2 (e / elucidate-01~e.26 :ARG0 m2 :ARG1 (m / mechanism~e.28 :poss~e.29 (p / proliferate-01~e.32 :ARG0 (e2 / epithelium~e.31) :ARG1-of (i / increase-01~e.30)))) :ARG1-of (c / cause-01~e.0,14,25 :ARG0 (s / stimulate-01~e.11 :ARG0 (a / and~e.5 :op1 m2~e.1,2,3,4 :op2 (m3 / macrophage~e.10 :mod (p2 / primary~e.9) :ARG1-of (e3 / educate-01~e.8 :ARG0 (t / tumor~e.6)))) :ARG1 p~e.12,13 :degree~e.17 (r / resemble-01~e.16)))) # ::id a_pmid_2169_9731.83 ::amr-annotator SDL-AMR-09 ::preferred # ::tok Because Kras pathways are commonly hyper @-@ activated in lung tumorigenesis [ @ 22 , 23 @ ] , and the tumorigenic lines examined herein contain Kras @ mutations , activities of downstream mediators Erk and Akt were examined . # ::alignments 0-1.2 2-1.2.1.1.1 4-1.2.1.1.5 4-1.2.1.1.5.r 5-1.2.1.1.2 7-1.2.1.1 8-1.2.1.1.3.r 9-1.2.1.1.3.1.1 10-1.2.1.1.3 10-1.2.1.1.3.1 10-1.2.1.1.3.1.r 13-1.2.1.1.4.1.1.1.1 17-1.2.1.1.4.1.1.1.2 21-1.2.1 21-1.2.1.1.4.1.1.1 23-1.2.1.2.1.2 23-1.2.1.2.1.2.1 23-1.2.1.2.1.2.1.r 23-1.2.1.2.1.2.2 23-1.2.1.2.1.2.2.r 24-1.2.1.2.1 25-1.2.1.2.1.1 26-1.2.1.2.1.1.1 27-1.2.1.2 31-1.2.1.2.2 33-1.1 34-1.1.1.r 35-1.1.1.3.1 36-1.1.1.3 37-1.1.1.1.1.1 38-1.1.1 39-1.1.1.2.1.1 41-1 (e / examine-01~e.41 :ARG1 (a / act-02~e.33 :ARG0~e.34 (a2 / and~e.38 :op1 (e2 / enzyme :name (n / name :op1 "Erk"~e.37)) :op2 (e3 / enzyme :name (n2 / name :op1 "Akt"~e.39)) :ARG0-of (m / mediate-01~e.36 :direction (d / downstream~e.35)))) :ARG1-of (c / cause-01~e.0 :ARG0 (a3 / and~e.21 :op1 (a4 / activate-01~e.7 :ARG1 (p / pathway~e.2 :name (n3 / name :op1 "K-Ras")) :degree (h / hyper~e.5) :time~e.8 (c2 / create-01~e.10 :ARG1~e.10 (t / tumor~e.10 :mod (l / lung~e.9))) :ARG1-of (d2 / describe-01 :ARG0 (p2 / publication-91 :ARG1-of (c3 / cite-01 :ARG2 (a5 / and~e.21 :op1 22~e.13 :op2 23~e.17)))) :manner~e.4 (c4 / common~e.4)) :op2 (c5 / contain-01~e.27 :ARG0 (l2 / line~e.24 :ARG1-of (e5 / examine-01~e.25 :medium (h2 / herein~e.26)) :ARG0-of (c6 / create-01~e.23 :ARG1~e.23 (t2 / tumor~e.23) :ARG1-of~e.23 (p3 / possible-01~e.23))) :ARG1 (m2 / mutate-01~e.31 :ARG2 (g / gene :name (n4 / name :op1 "K-Ras"))))))) # ::id a_pmid_2169_9731.84 ::amr-annotator SDL-AMR-09 ::preferred # ::tok Cytosolic Raf functionally links the Erk and Akt pathways ; activated Akt can phosphorylate cRaf at S259 , placing Erk regulation downstream of Akt activation [ @ 32 , 33 @ ] . # ::alignments 0-1.1.1.2 1-1.1.1.1.1 2-1.1.3 3-1.1 5-1.1.2.1.1.1 6-1 6-1.1.2 7-1.1.2.2.1.1 7-1.2.1.2.1.1 8-1.1.2.1 8-1.1.2.2 8-1.2.1.2 10-1.2.1.2.2 11-1.2.1.2.1.1 12-1.2 13-1.2.1 18-1.2.1.3.1 19-1.2.1.3.1.1.1 20-1.2.1.3.1.1 21-1.2.1.3.1.2.2 23-1.2.1.3.1.2.1 24-1.2.1.3.1.2.1 27-1.2.2.1.1.1.1 31-1.2.2.1.1.1.2 (a / and~e.6 :op1 (l / link-01~e.3 :ARG0 (e / enzyme :name (n / name :op1 "Raf"~e.1) :location (c / cytosol~e.0)) :ARG1 (a2 / and~e.6 :op1 (p2 / pathway~e.8 :name (n2 / name :op1 "Erk"~e.5)) :op2 (p3 / pathway~e.8 :name (n3 / name :op1 "Akt"~e.7))) :ARG0-of (f / function-01~e.2)) :op2 (p4 / possible-01~e.12 :ARG1 (p / phosphorylate-01~e.13 :ARG1 (a4 / amino-acid :mod 259 :name (n6 / name :op1 "serine") :part-of (e2 / enzyme :name (n4 / name :op1 "C-Raf"))) :ARG2 (p7 / pathway~e.8 :name (n5 / name :op1 "Akt"~e.7,11) :ARG1-of (a3 / activate-01~e.10)) :ARG0-of (c2 / cause-01 :ARG1 (p5 / place-01~e.18 :ARG1 (r / regulate-01~e.20 :ARG1 p2~e.19) :ARG2 (r3 / relative-position :op1 p7~e.23,24 :direction (d / downstream~e.21))))) :ARG1-of (d2 / describe-01 :ARG0 (p6 / publication-91 :ARG1-of (c3 / cite-01 :ARG2 (a6 / and :op1 32~e.27 :op2 33~e.31)))))) # ::id a_pmid_2169_9731.85 ::amr-annotator SDL-AMR-09 ::preferred # ::tok MH @-@ S co @-@ culture stimulated cRaf phosphorylation at S259 in all three cell lines , resulting in significantly higher levels of p @-@ cRaf ( Figure 3A @-@ C ) . # ::alignments 0-1.1.1.1.1.1 2-1.1.1.1.1.1 3-1.1 4-1.1 5-1.1 6-1 8-1.2 8-1.3.2.2 11-1.4.r 12-1.4.2 13-1.4.1 14-1.4 15-1.4 18-1.3.r 19-1.3.1.2 20-1.3.1 20-1.3.1.1 20-1.3.1.1.r 21-1.3 23-1.2 23-1.3.2.2 27-1.5.1.1 27-1.5.1.2 27-1.5.1.3 29-1.5.1.1.1 31-1.2.1.3.1.1 31-1.3.2.1.1 (s / stimulate-01~e.6 :ARG0 (c / coculture-01~e.3,4,5 :ARG1 (m / macrophage :mod (c3 / cell-line :name (n / name :op1 "MH-S"~e.0,2)))) :ARG1 (p / phosphorylate-01~e.8,23 :ARG1 (a / amino-acid :mod 259 :name (n2 / name :op1 "serine") :part-of (e / enzyme :name (n3 / name :op1 "C-Raf"~e.31)))) :ARG3~e.18 (l / level~e.21 :ARG1-of (h / high-02~e.20 :degree~e.20 (m2 / more~e.20) :ARG1-of (s2 / significant-02~e.19)) :quant-of (e2 / enzyme :name (n4 / name :op1 "C-Raf"~e.31) :ARG3-of (p2 / phosphorylate-01~e.8,23))) :location~e.11 (c2 / cell-line~e.14,15 :quant 3~e.13 :mod (a2 / all~e.12)) :ARG1-of (d / describe-01 :ARG0 (a3 / and :op1 (f / figure~e.27 :mod "3A"~e.29) :op2 (f2 / figure~e.27 :mod "3B") :op3 (f3 / figure~e.27 :mod "3C")))) # ::id a_pmid_2169_9731.86 ::amr-annotator SDL-AMR-09 ::preferred # ::tok The smaller (~ 74 kDa ) p @-@ cRaf isoform was most highly abundant and its phosphorylation significantly increased with macrophage co @-@ culture in the LM2 and E10 cells , but a larger (~ 100 kDa ) isoform was heavily phosphorylated at the expense of the 74 kDa isoform in neoplastic JF32 cells ( Figure 3A ) . # ::alignments 1-1.1.1.1.3 1-1.1.1.1.3.1 1-1.1.1.1.3.1.r 3-1.1.1.1.2.1.1.1 4-1.1.1.1.2.1.1.2 6-1.1.1.1.1.2 9-1.1.1.1 10-1.1.1.1.r 11-1.1.1.2.1 12-1.1.1.2 13-1.1.1 14-1.1 16-1.1.1.1.1.2 16-1.1.2.1 17-1.1.2.2 18-1.1.2 19-1.1.2.3.r 20-1.1.2.3.1 21-1.1.2.3 22-1.1.2.3 23-1.1.2.3 24-1.1.2.3.2.r 26-1.1.2.3.2.1.1.1 27-1.1.2.3.2 28-1.1.2.3.2.2.1.1 29-1.1.2.3.2.1 29-1.1.2.3.2.2 31-1 33-1.2.1.1 33-1.2.1.1.1 33-1.2.1.1.1.r 35-1.2.1.2.1.1.1 36-1.2.1.2.1.1.2 38-1.2.1 40-1.2.2 40-1.2.2.r 41-1.2 42-1.2.3.r 44-1.2.3 45-1.2.3 47-1.2.3.1.1 48-1.2.3.1.1 49-1.2.3.1 50-1.2.4.r 51-1.2.4.2 52-1.2.4.1.1 53-1.2.4 55-1.3.1 57-1.3.1.1 (c / contrast-01~e.31 :ARG1 (a / and~e.14 :op1 (a2 / abundant~e.13 :domain~e.10 (i / isoform~e.9 :mod (e / enzyme :name (n / name :op1 "C-Raf") :ARG3-of (p2 / phosphorylate-01~e.6,16)) :ARG1-of (e2 / equal-01 :ARG2 (a4 / approximately :op1 (m2 / mass-quantity :quant 74~e.3 :unit (k / kilodalton~e.4)))) :mod (s / small~e.1 :degree~e.1 (m3 / more~e.1))) :ARG1-of (h / high-02~e.12 :degree (m / most~e.11))) :op2 (i2 / increase-01~e.18 :ARG1 (p3 / phosphorylate-01~e.16 :ARG1 e) :ARG2 (s2 / significant-02~e.17) :instrument~e.19 (c2 / coculture-01~e.21,22,23 :ARG1 (m4 / macrophage~e.20) :location~e.24 (a3 / and~e.27 :op1 (c3 / cell-line~e.29 :name (n2 / name :op1 "LM2"~e.26)) :op2 (c4 / cell-line~e.29 :name (n3 / name :op1 "E10"~e.28)))))) :ARG2 (p / phosphorylate-01~e.41 :ARG1 (i3 / isoform~e.38 :mod (l / large~e.33 :degree~e.33 (m5 / more~e.33)) :ARG1-of (e3 / equal-01 :ARG2 (a5 / approximately :op1 (m6 / mass-quantity :quant 100~e.35 :unit (k2 / kilodalton~e.36))))) :manner~e.40 (h2 / heavy~e.40) :ARG0-of~e.42 (c5 / compromise-02~e.44,45 :ARG1 (i4 / isoform~e.49 :quant m2~e.47,48)) :location~e.50 (c6 / cell-line~e.53 :name (n5 / name :op1 "JF32"~e.52) :mod (n6 / neoplastic~e.51))) :ARG1-of (d / describe-01 :ARG0 (f / figure~e.55 :mod "3A"~e.57))) # ::id a_pmid_2169_9731.87 ::amr-annotator SDL-AMR-09 ::preferred # ::tok The 74 kDa isoform was the most abundant in total cRaf immunoblots from all three cell lines . # ::alignments 1-1.3.1.1 2-1.3.1.2 3-1.3 4-1.3.r 6-1.1 7-1 9-1.2.3 13-1.2.2.2 14-1.2.2.1 15-1.2.2 16-1.2.2 (a / abundant~e.7 :degree (m / most~e.6) :location (i2 / immunoblot-01 :ARG1 (e / enzyme :name (n2 / name :op1 "C-Raf")) :ARG2 (c / cell-line~e.15,16 :quant 3~e.14 :mod (a2 / all~e.13)) :mod (t / total~e.9)) :domain~e.4 (i / isoform~e.3 :quant (m2 / mass-quantity :quant 74~e.1 :unit (k / kilodalton~e.2)))) # ::id a_pmid_2169_9731.88 ::amr-annotator SDL-AMR-09 ::preferred # ::tok MH @-@ S co @-@ culture significantly increased the levels of active Erk1 @/@ 2 ( p @-@ Erk ) in LM2 and JF32 cells , as well as non @-@ neoplastic E10 cells , when normalized either to total Erk ( panErk ) or β-actin levels ( Figure 3A , D and 3E ) , which correlates with the observed increases in proliferation ( Figure 2 ) . # ::alignments 0-1.1.1.1.1.1 2-1.1.1.1.1.1 3-1.1 4-1.1 5-1.1 6-1.3 7-1 9-1.2 10-1.2.1.r 11-1.2.1.3 12-1.2.1.1.1.1 13-1.2.1 14-1.7.2.1.1 16-1.2.1.4.1.2 18-1.2.1.4.1.1.1 20-1.4.r 21-1.4.1.1.1 22-1.4 23-1.4.2.1.1 24-1.4.1 24-1.4.2 24-1.4.3 26-1.4 27-1.4 28-1.4 29-1.4.3.2.1 29-1.4.3.2.1.r 31-1.4.3.2 32-1.4.3.1.1 33-1.1.1.1 33-1.4.3 36-1.5 38-1.5.2 39-1.5.2.1.1.1.2 40-1.5.2.1.1.1.1.1 40-1.5.2.1.1.1.3.1.1.1 44-1.5.2.1 45-1.5.2.1.2.1.1.1 46-1.5.2.1.1 46-1.5.2.1.2 48-1.6.1.1 48-1.6.1.2 48-1.6.1.3 50-1.6.1.1.1 54-1.6.1 56-1.6.1.3.1 61-1.7 62-1.7.1.r 64-1.7.1.2 65-1.7.1 66-1.7.1.1.r 67-1.7.1.1 69-1.7.2.1 71-1.7.2.1.1 (i / increase-01~e.7 :ARG0 (c / coculture-01~e.3,4,5 :ARG1 (m / macrophage :mod (c7 / cell-line~e.33 :name (n / name :op1 "MH-S"~e.0,2)))) :ARG1 (l / level~e.9 :quant-of~e.10 (s2 / slash~e.13 :op1 (e3 / enzyme :name (n10 / name :op1 "Erk1"~e.12)) :op2 (e4 / enzyme :name (n11 / name :op1 "Erk2")) :ARG0-of (a / activity-06~e.11) :ARG1-of (m2 / mean-01 :ARG2 (e / enzyme :name (n2 / name :op1 "Erk"~e.18) :ARG3-of (p3 / phosphorylate-01~e.16))))) :ARG2 (s / significant-02~e.6) :location~e.20 (a2 / and~e.22,26,27,28 :op1 (c2 / cell-line~e.24 :name (n3 / name :op1 "LM2"~e.21)) :op2 (c3 / cell-line~e.24 :name (n4 / name :op1 "JF32"~e.23)) :op3 (c4 / cell-line~e.24,33 :name (n5 / name :op1 "E10"~e.32) :mod (n6 / neoplastic~e.31 :polarity~e.29 -~e.29))) :condition (n7 / normalize-01~e.36 :ARG1 s2 :ARG1-of (c6 / conform-01~e.38 :ARG2 (o / or~e.44 :op1 (l3 / level~e.46 :quant-of (e2 / enzyme :name (n8 / name :op1 "Erk"~e.40) :mod (t / total~e.39) :ARG1-of (m3 / mean-01 :ARG2 (e5 / enzyme :name (n12 / name :op1 "Erk"~e.40) :mod (p4 / pan))))) :op2 (l2 / level~e.46 :quant-of (p / protein :name (n9 / name :op1 "β-actin"~e.45)))))) :ARG1-of (d / describe-01 :ARG0 (a3 / and~e.54 :op1 (f / figure~e.48 :mod "3A"~e.50) :op2 (f2 / figure~e.48 :mod "3D") :op3 (f3 / figure~e.48 :mod "3E"~e.56))) :ARG1-of (c5 / correlate-01~e.61 :ARG2~e.62 (i2 / increase-01~e.65 :ARG1~e.66 (p2 / proliferate-01~e.67) :ARG1-of (o2 / observe-01~e.64)) :ARG1-of (d2 / describe-01 :ARG0 (f4 / figure~e.69 :mod 2~e.14,71)))) # ::id a_pmid_2169_9731.89 ::amr-annotator SDL-AMR-09 ::preferred # ::tok E10 cells expressed lower basal p @-@ Erk @/@ panErk vs. the neoplastic cell lines , consistent with previous observations [ @ 21 @ ] . # ::alignments 0-1.2.1.1 1-1.2 2-1 2-1.3.1 3-1.1.4 3-1.1.4.1 3-1.1.4.1.r 4-1.1.3 5-1.1.1.2 7-1.1.1.1.1 7-1.1.2.1.1 8-1.1 10-1.3 12-1.3.1.2.1 13-1.3.1.2 14-1.3.1.2 16-1.4 17-1.4.1.r 18-1.4.1.1 19-1.4.1 22-1.5.1.1.1 (e / express-03~e.2 :ARG2 (s / slash~e.8 :op1 (e3 / enzyme :name (n4 / name :op1 "Erk"~e.7) :ARG3-of (p4 / phosphorylate-01~e.5)) :op2 (e4 / enzyme :name (n5 / name :op1 "Erk"~e.7) :mod (p5 / pan)) :mod (b / basal~e.4) :ARG1-of (l / low-04~e.3 :degree~e.3 (m / more~e.3))) :ARG3 (c / cell-line~e.1 :name (n / name :op1 "E10"~e.0)) :ARG1-of (c2 / contrast-01~e.10 :ARG2 (e2 / express-03~e.2 :ARG2 s :ARG3 (c3 / cell-line~e.13,14 :mod (n3 / neoplastic~e.12)))) :ARG1-of (c4 / consistent-01~e.16 :ARG2~e.17 (o / observe-01~e.19 :time (p2 / previous~e.18))) :ARG1-of (d / describe-01 :ARG0 (p3 / publication-91 :ARG1-of (c5 / cite-01 :ARG2 21~e.22)))) # ::id a_pmid_2169_9731.90 ::amr-annotator SDL-AMR-09 ::preferred # ::tok Total Erk remained unchanged in both neoplastic cell lines , while macrophage co @-@ culture caused Erk2 ( 42 kDa ) to nearly disappear in the E10 cells , with little effect on Erk1 ( Figure 3A , D and 3E ) . # ::alignments 0-1.1.1.2 1-1.1.1.1.1 2-1.1 3-1.1.2 3-1.1.2.1 3-1.1.2.1.r 4-1.1.2.3.r 5-1.1.2.3.1 6-1.1.2.3.2 7-1.1.2.3 8-1.1.2.3 10-1 11-1.2.1.1 12-1.2.1 13-1.2.1 14-1.2.1 15-1.2 16-1.2.2.2.1.1 22-1.2.2.3 23-1.2.2 24-1.2.2.4.r 26-1.2.2.4.1.1 27-1.2.2.4 29-1.2.3.r 30-1.2.3.2 31-1.2.3 32-1.2.3.1.r 33-1.2.3.1.1.1 35-1.3.1.1 35-1.3.1.2 35-1.3.1.3 37-1.3.1.1.1 41-1.3.1 43-1.3.1.3.1 (c / contrast-01~e.10 :ARG1 (r / remain-01~e.2 :ARG1 (e / enzyme :name (n / name :op1 "Erk"~e.1) :mod (t / total~e.0)) :ARG3 (c2 / change-01~e.3 :polarity~e.3 -~e.3 :ARG1 e :location~e.4 (c6 / cell-line~e.7,8 :mod (b / both~e.5) :mod (n6 / neoplastic~e.6)))) :ARG2 (c3 / cause-01~e.15 :ARG0 (c4 / coculture-01~e.12,13,14 :ARG1 (m / macrophage~e.11)) :ARG1 (d / disappear-01~e.23 :ARG0 c4 :ARG1 (e2 / enzyme :name (n2 / name :op1 "Erk2"~e.16)) :mod (n3 / near~e.22) :location~e.24 (c5 / cell-line~e.27 :name (n4 / name :op1 "E10"~e.26))) :ARG0-of~e.29 (a / affect-01~e.31 :ARG1~e.32 (e3 / enzyme :name (n5 / name :op1 "Erk1"~e.33)) :degree (l / little~e.30))) :ARG1-of (d2 / describe-01 :ARG0 (a2 / and~e.41 :op1 (f / figure~e.35 :mod "3A"~e.37) :op2 (f2 / figure~e.35 :mod "3D") :op3 (f3 / figure~e.35 :mod "3E"~e.43)))) # ::id a_pmid_2169_9731.91 ::amr-annotator SDL-AMR-09 ::preferred # ::tok Activated Akt ( p @-@ Akt ) levels rose significantly in both neoplastic cell lines when normalized to either total Akt ( panAkt ) or β-actin , but macrophage co @-@ culture caused both p @-@ Akt and panAkt levels to rise to similar extents in E10 cells ( Figure 3A and 3F ) . # ::alignments 0-1.1.1.1.2 1-1.1.1.1.1.1 1-1.1.1.1.3.1.1.1 3-1.1.1.1.3.1.2 5-1.1.1.1.1.1 5-1.1.1.1.3.1.1.1 7-1.1.1 8-1.1 9-1.1.2 10-1.1.3.r 11-1.1.3.1 12-1.1.3.2 13-1.1.3 14-1.1.3 16-1.1.4 17-1.1.4.2 19-1.1.4.2.1.1.2 20-1.1.4.2.1.1.1.1 24-1.1.4.2.1 25-1.1.4.2.1.2.1.1 27-1 28-1.2.1.1 29-1.2.1 30-1.2.1 31-1.2.1 32-1.2 33-1.1.3.1 34-1.2.2.2.1.1 35-1.2.2.2.1.1 36-1.2.2.2.1.1 37-1.2.2.2.1 39-1.2.2.2 41-1.2.2 42-1.2.2.3.r 43-1.2.2.3.1 44-1.2.2.3 47-1.2.2.4 49-1.3.1.1 49-1.3.1.2 51-1.3.1.1.1 53-1.3.1 55-1.3.1.2.1 (c / contrast-01~e.27 :ARG1 (r / rise-01~e.8 :ARG1 (l / level~e.7 :quant-of (e / enzyme :name (n / name :op1 "Akt"~e.1,5) :ARG1-of (a / activate-01~e.0) :ARG1-of (m2 / mean-01 :ARG2 (e3 / enzyme :name (n6 / name :op1 "Akt"~e.1,5) :ARG3-of (p2 / phosphorylate-01~e.3))))) :ARG2 (s / significant-02~e.9) :location~e.10 (c3 / cell-line~e.13,14 :mod (b / both~e.11,33) :mod (n2 / neoplastic~e.12)) :condition (n3 / normalize-01~e.16 :ARG1 e :ARG1-of (c5 / conform-01~e.17 :ARG2 (o / or~e.24 :op1 (e2 / enzyme :name (n4 / name :op1 "Akt"~e.20) :mod (t / total~e.19)) :op2 (p / protein :name (n5 / name :op1 "β-actin"~e.25)))))) :ARG2 (c2 / cause-01~e.32 :ARG0 (c4 / coculture-01~e.29,30,31 :ARG1 (m / macrophage~e.28)) :ARG1 (r3 / rise-01~e.41 :ARG0 c4 :ARG1 (l2 / level~e.39 :quant-of (a2 / and~e.37 :op1 e~e.34,35,36 :op2 e2)) :ARG2~e.42 (e4 / extent~e.44 :ARG1-of (r4 / resemble-01~e.43 :ARG2 l)) :location c3~e.47)) :ARG1-of (d / describe-01 :ARG0 (a3 / and~e.53 :op1 (f / figure~e.49 :mod "3A"~e.51) :op2 (f2 / figure~e.49 :mod "3F"~e.55)))) # ::id a_pmid_2169_9731.92 ::amr-annotator SDL-AMR-09 ::preferred # ::tok When p @-@ Akt was normalized to panAkt expression , there was no change in E10 cells with MH @-@ S co @-@ culture ( Figure 3F ) . # ::alignments 0-1.3.r 1-1.3.1.2 3-1.3.1.1.1 3-1.3.2.1.1.1 5-1.3 6-1.3.2 8-1.3.2.1 8-1.3.2.1.3 8-1.3.2.1.3.r 12-1.1 12-1.1.r 13-1 14-1.2.r 15-1.2.1.1 16-1.2 16-1.2.2.1.1.1 18-1.2.2.1.1.1.1.1 20-1.2.2.1.1.1.1.1 21-1.2.2.1 22-1.2.2.1 23-1.2.2.1 25-1.4.1 27-1.4.1.1 (c / change-01~e.13 :polarity~e.12 -~e.12 :ARG1~e.14 (c2 / cell-line~e.16 :name (n / name :op1 "E10"~e.15) :ARG0-of (c3 / contain-01 :ARG1 (c4 / coculture-01~e.21,22,23 :ARG1 (m / macrophage :mod (c6 / cell-line~e.16 :name (n2 / name :op1 "MH-S"~e.18,20)))))) :time~e.0 (n3 / normalize-01~e.5 :ARG1 (e2 / enzyme :name (n4 / name :op1 "Akt"~e.3) :ARG3-of (p / phosphorylate-01~e.1)) :ARG1-of (c5 / conform-01~e.6 :ARG2 (e3 / enzyme~e.8 :name (n6 / name :op1 "Akt"~e.3) :mod (p3 / pan) :ARG2-of~e.8 (e / express-03~e.8)))) :ARG1-of (d / describe-01 :ARG0 (f / figure~e.25 :mod "3F"~e.27))) # ::id a_pmid_2169_9731.93 ::amr-annotator SDL-AMR-09 ::preferred # ::tok Total Akt expression increased slightly in LM2 cells but decreased in JF32 cells ( Figure 3A ) . # ::alignments 0-1.1.1.1.2 1-1.1.1.1.1.1 2-1.1.1 3-1.1 4-1.1.2 5-1.1.3.r 6-1.1.3.1.1 7-1.1.3 8-1 9-1.2 10-1.2.2.r 11-1.2.2.1.1 12-1.2.2 14-1.3.1 16-1.3.1.1 (c / contrast-01~e.8 :ARG1 (i2 / increase-01~e.3 :ARG1 (e / express-03~e.2 :ARG2 (e2 / enzyme :name (n / name :op1 "Akt"~e.1) :mod (t / total~e.0))) :ARG2 (s / slight~e.4) :location~e.5 (c2 / cell-line~e.7 :name (n2 / name :op1 "LM2"~e.6))) :ARG2 (d / decrease-01~e.9 :ARG1 e :location~e.10 (c3 / cell-line~e.12 :name (n3 / name :op1 "JF32"~e.11))) :ARG1-of (d2 / describe-01 :ARG0 (f / figure~e.14 :mod "3A"~e.16))) # ::id a_pmid_2169_9731.94 ::amr-annotator SDL-AMR-09 ::preferred # ::tok When normalized to β-actin , p @-@ Akt levels significantly increased upon MH @-@ S co @-@ culture in all three cell lines ( Figure 3A and 3G ) . # ::alignments 1-1.4 2-1.4.2 3-1.4.2.1.1.1 5-1.2.1.2 7-1.2.1.1.1 8-1.2 9-1.3 10-1 12-1.1.1.1.1.1 14-1.1.1.1.1.1 15-1.1 16-1.1 17-1.1 18-1.5.r 19-1.5.2 20-1.5.1 21-1.5 22-1.5 24-1.6.1.1 24-1.6.1.2 26-1.6.1.1.1 28-1.6.1 30-1.6.1.2.1 (i / increase-01~e.10 :ARG0 (c / coculture-01~e.15,16,17 :ARG1 (m / macrophage :mod (c4 / cell-line :name (n4 / name :op1 "MH-S"~e.12,14)))) :ARG1 (l / level~e.8 :quant-of (e / enzyme :name (n3 / name :op1 "Akt"~e.7) :ARG3-of (p2 / phosphorylate-01~e.5))) :ARG2 (s / significant-02~e.9) :condition (n / normalize-01~e.1 :ARG1 e :ARG1-of (c3 / conform-01~e.2 :ARG2 (p / protein :name (n2 / name :op1 "β-actin"~e.3)))) :location~e.18 (c2 / cell-line~e.21,22 :quant 3~e.20 :mod (a / all~e.19)) :ARG1-of (d / describe-01 :ARG0 (a2 / and~e.28 :op1 (f / figure~e.24 :mod "3A"~e.26) :op2 (f2 / figure~e.24 :mod "3G"~e.30)))) # ::id a_pmid_2169_9731.95 ::amr-annotator SDL-AMR-09 ::preferred # ::tok Increased p @-@ S473 Akt content suggests increased enzymatic activity , which can be confirmed by enhanced phosphorylation of downstream substrates . # ::alignments 0-1.1.2 1-1.1.1.2 1-1.1.1.2.3 1-1.1.1.2.3.r 4-1.1.1.1.1 5-1.1 6-1 7-1.1.2 8-1.2.1 9-1.2 12-1.2.3.2 14-1.2.3 15-1.2.3.1.r 16-1.2.3.1.2 17-1.2.3.1 18-1.2.3.1.1.r 19-1.2.3.1.1.1 20-1.2.3.1.1 (s / suggest-01~e.6 :ARG0 (c / contain-01~e.5 :ARG1 (e / enzyme :name (n / name :op1 "Akt"~e.4) :part (a / amino-acid~e.1 :mod 473 :name (n2 / name :op1 "serine") :ARG1-of~e.1 (p / phosphorylate-01~e.1))) :ARG1-of (i / increase-01~e.0,7)) :ARG1 (a2 / activity-06~e.9 :ARG0 (e2 / enzyme~e.8) :ARG1-of i :ARG1-of (c2 / confirm-01~e.14 :ARG0~e.15 (p3 / phosphorylate-01~e.17 :ARG1~e.18 (s2 / substrate~e.20 :direction (d / downstream~e.19)) :ARG1-of (e3 / enhance-01~e.16)) :ARG1-of (p2 / possible-01~e.12)))) # ::id a_pmid_2169_9731.96 ::amr-annotator SDL-AMR-09 ::preferred # ::tok To determine if macrophage co @-@ culture increases Akt activity , we measured levels of p @-@ GSK @-@ 3β, a known target of Akt [ @ 32 @ ] . # ::alignments 1-1.3 3-1.3.2.1.1 4-1.3.2.1 5-1.3.2.1 6-1.3.2.1 7-1.3.2 8-1.3.2.2.1.1.1 9-1.3.2.2 11-1.1 12-1 13-1.2 14-1.2.1.r 15-1.2.1.3 17-1.2.1.1.1 21-1.2.1.2.2 22-1.2.1 22-1.2.1.2 22-1.2.1.2.r 23-1.2.1.2.1.r 24-1.2.1.2.1 27-1.4.1.1.1 (m / measure-01~e.12 :ARG0 (w / we~e.11) :ARG1 (l / level~e.13 :quant-of~e.14 (e2 / enzyme~e.22 :name (n2 / name :op1 "GSK-3β"~e.17) :ARG1-of~e.22 (t / target-01~e.22 :ARG0~e.23 e~e.24 :ARG1-of (k / know-01~e.21)) :ARG3-of (p / phosphorylate-01~e.15))) :purpose (d / determine-01~e.1 :ARG0 w :ARG1 (i / increase-01~e.7 :ARG0 (c / coculture-01~e.4,5,6 :ARG1 (m2 / macrophage~e.3)) :ARG1 (a / activity-06~e.9 :ARG0 (e / enzyme :name (n / name :op1 "Akt"~e.8))))) :ARG1-of (d2 / describe-01 :ARG0 (p2 / publication-91 :ARG1-of (c2 / cite-01 :ARG2 32~e.27)))) # ::id a_pmid_2169_9731.97 ::amr-annotator SDL-AMR-09 ::preferred # ::tok Consistent with the elevation in p @-@ Akt , MH @-@ S co @-@ culture significantly increased p @-@ GSK @-@ 3β in both LM2 and E10 cells and trended towards an increase in JF32 cells ( Figure 3A and 3H ) ; panGSK @-@ 3β levels were unchanged ( data not shown ) . # ::alignments 0-1.2.3 1-1.2.3.1.r 3-1.2.3.1 4-1.2.3.1.1.r 5-1.2.3.1.1.2 7-1.2.3.1.1.1.1 9-1.2.2.2 10-1.2.2.2 11-1.2.2.2 12-1.2.2.2 13-1.2.2.2 14-1.2.2.2 15-1.2.2.2 16-1.2.2.2 17-1.2.2.2 18-1.2.2.2 19-1.2.2.2 20-1.2.2.2 21-1.2.2.2 22-1.2.2.2 23-1.2.2.2 24-1.2.2.2 25-1.2.2.2 26-1.2.2.2 27-1.2.2.2 28-1.2 28-1.2.1.4 29-1.2.2 32-1.2.2.2 34-1.2.2.3.1.1 35-1.2.1.1.1.1 35-1.2.2.3 37-1.2.4.1.1 37-1.2.4.1.2 39-1.2.4.1.1.1 41-1.2.4.1 43-1.2.4.1.2.1 49-1.1.2.1.1.1 50-1.1.2 52-1.1 52-1.1.1 52-1.1.1.r 54-1.1.3.1 55-1.1.3.1.1.1 55-1.1.3.1.1.1.r 56-1.1.3.1.1 (m / multi-sentence :snt2 (c / change-01~e.52 :polarity~e.52 -~e.52 :ARG1 (l / level~e.50 :quant-of (e2 / enzyme :name (n6 / name :op1 "GSK-3β"~e.49) :mod (p2 / pan))) :ARG1-of (d2 / describe-01 :ARG0 (d3 / data~e.54 :ARG1-of (s2 / show-01~e.56 :polarity~e.55 -~e.55)))) :snt1 (a2 / and~e.28 :op1 (i / increase-01 :ARG0 (c2 / coculture-01 :ARG1 (m2 / macrophage :mod (c7 / cell-line~e.35 :name (n / name :op1 "MH-S")))) :ARG1 (e / enzyme :name (n2 / name :op1 "GSK-3β") :ARG3-of (p / phosphorylate-01)) :ARG2 (s / significant-02) :location (a / and~e.28 :op1 (c3 / cell-line :name (n3 / name :op1 "LM2")) :op2 (c4 / cell-line :name (n4 / name :op1 "E10")))) :op2 (t / trend-01~e.29 :ARG1 c2 :ARG2 i~e.9,10,11,12,13,14,15,16,17,18,19,20,21,22,23,24,25,26,27,32 :location (c5 / cell-line~e.35 :name (n5 / name :op1 "JF32"~e.34))) :ARG1-of (c6 / consistent-01~e.0 :ARG2~e.1 (e3 / elevate-01~e.3 :ARG1~e.4 (e4 / enzyme :name (n7 / name :op1 "Akt"~e.7) :ARG3-of p~e.5))) :ARG1-of (d / describe-01 :ARG0 (a3 / and~e.41 :op1 (f / figure~e.37 :mod "3A"~e.39) :op2 (f2 / figure~e.37 :mod "3H"~e.43))))) # ::id a_pmid_2169_9731.98 ::amr-annotator SDL-AMR-09 ::preferred # ::tok Phospho @-@ S259 cRaf is another measure of Akt activity , and p @-@ cRaf levels increased in all three cell lines with macrophage co @-@ culture ( Figure 3A @-@ C ) . # ::alignments 0-1.1.1.2.3 5-1.1.3 6-1.1 7-1.1.2.r 8-1.1.2.1.1.1 9-1.1.2 12-1.1.1.2.3 15-1.2.1 16-1.2 17-1.2.2.r 18-1.2.2.2 19-1.2.2.1 20-1.2.2 21-1.2.2 23-1.2.2.3.1.1 24-1.2.2.3.1 25-1.2.2.3.1 26-1.2.2.3.1 28-1.3.1.1 28-1.3.1.2 30-1.3.1.1.1 32-1.1.1.1.1 (a / and :op1 (m / measure-01~e.6 :ARG0 (e / enzyme :name (n / name :op1 "C-Raf"~e.32) :part (a2 / amino-acid :mod 259 :name (n2 / name :op1 "serine") :ARG3-of (p / phosphorylate-01~e.0,12))) :ARG1~e.7 (a3 / activity-06~e.9 :ARG0 (e2 / enzyme :name (n3 / name :op1 "Akt"~e.8))) :mod (a4 / another~e.5)) :op2 (i / increase-01~e.16 :ARG1 (l / level~e.15 :quant-of e) :location~e.17 (c / cell-line~e.20,21 :quant 3~e.19 :mod (a5 / all~e.18) :ARG0-of (c2 / contain-01 :ARG1 (c3 / coculture-01~e.24,25,26 :ARG1 (m2 / macrophage~e.23))))) :ARG1-of (d / describe-01 :ARG0 (v / value-interval :op1 (f / figure~e.28 :mod "3A"~e.30) :op2 (f2 / figure~e.28 :mod "3C")))) # ::id a_pmid_2169_9731.99 ::amr-annotator SDL-AMR-09 ::preferred # ::tok Together , the observed increases in epithelial proliferation and the known roles for Erk and Akt in neoplastic lung cell division suggest that macrophage co @-@ culture stimulates lung cell proliferation through increased Erk and Akt activity [ @ 34 @ ] . # ::alignments 0-1.1.3 3-1.1.1.2 4-1.1.1 6-1.1.1.1.1 7-1.1.1.1 8-1.1 10-1.1.2.1 11-1.1.2 12-1.1.2.2.r 13-1.1.2.2.1.1.1 15-1.1.2.2.2.1.1 16-1.1.2.3.r 17-1.1.2.3.1.2 18-1.1.2.3.1.1 19-1.1.2.3.1 20-1.1.2.3 21-1 22-1.2.r 23-1.2.1.1 24-1.2.1 25-1.2.1 26-1.2.1 27-1.2 28-1.2.2.1.1 29-1.2.2.1 30-1.2.2 32-1.1.1 32-1.2.3.2 33-1.2.3.1 34-1.2.3.1 35-1.2.3.1 36-1.2.3 39-1.3.1.1.1 (s / suggest-01~e.21 :ARG0 (a / and~e.8 :op1 (i / increase-01~e.4,32 :ARG1 (p / proliferate-01~e.7 :ARG0 (e / epithelium~e.6)) :ARG1-of (o / observe-01~e.3)) :op2 (r / role~e.11 :ARG1-of (k / know-01~e.10) :poss~e.12 (a3 / and :op1 (e2 / enzyme :name (n / name :op1 "Erk"~e.13)) :op2 (e3 / enzyme :name (n2 / name :op1 "Akt"~e.15))) :topic~e.16 (d / divide-02~e.20 :ARG1 (c / cell~e.19 :mod (l / lung~e.18) :mod (n3 / neoplasm~e.17)))) :mod (t / together~e.0)) :ARG1~e.22 (s2 / stimulate-01~e.27 :ARG0 (c2 / coculture-01~e.24,25,26 :ARG1 (m / macrophage~e.23)) :ARG1 (p2 / proliferate-01~e.30 :ARG0 (c4 / cell~e.29 :mod l~e.28)) :manner (a2 / activity-06~e.36 :ARG0 a3~e.33,34,35 :ARG1-of (i2 / increase-01~e.32))) :ARG1-of (d2 / describe-01 :ARG0 (p3 / publication-91 :ARG1-of (c3 / cite-01 :ARG2 34~e.39)))) # ::id a_pmid_2169_9731.100 ::amr-annotator SDL-AMR-09 ::preferred # ::tok Combined inhibition of MEK and PI3K abrogates macrophage stimulation of neoplastic growth # ::alignments 0-1.1.2 1-1.1 2-1.1.1.r 3-1.1.1.1.1.1 4-1.1.1 5-1.1.1.2.1.1 6-1 7-1.2.1 8-1.2 9-1.2.2.r 10-1.2.2.1 11-1.2.2 (a / abrogate-01~e.6 :ARG0 (i / inhibit-01~e.1 :ARG1~e.2 (a2 / and~e.4 :op1 (e / enzyme :name (n / name :op1 "MEK"~e.3)) :op2 (e2 / enzyme :name (n2 / name :op1 "PI3K"~e.5))) :ARG3-of (c / combine-01~e.0)) :ARG1 (s / stimulate-01~e.8 :ARG0 (m / macrophage~e.7) :ARG1~e.9 (g / grow-01~e.11 :ARG1 (n3 / neoplasm~e.10)))) # ::id a_pmid_2169_9731.101 ::amr-annotator SDL-AMR-09 ::preferred # ::tok Erk and Akt regulate both proliferation and resistance to apoptotic cell death , are more active in lung tumors than in normal tissue [ @ 21 , 35 @ ] , and were activated with macrophage co @-@ culture . # ::alignments 0-1.1.1.1.1.1 1-1.1.1 2-1.1.1.2.1.1 3-1.1 5-1.1.2.1 6-1.1.2 7-1.1.2.2 8-1.1.2.2.1.r 9-1.1.2.2.1.2 10-1.1.2.2.1.1 11-1.1.2.2.1 14-1.2.2 15-1.2 16-1.2.4.r 17-1.2.4.1 18-1.2.4 19-1.2.4.2.r 21-1.2.4.2.1 22-1.2.4.2 25-1.2.3.1.1.1.1 29-1.2.3.1.1.1.2 33-1 33-1.2.3.1.1.1 35-1.3 36-1.3.1.r 37-1.3.1.1 38-1.3.1 39-1.3.1 40-1.3.1 (a / and~e.33 :op1 (r / regulate-01~e.3 :ARG0 (a4 / and~e.1 :op1 (e / enzyme :name (n / name :op1 "Erk"~e.0)) :op2 (e2 / enzyme :name (n2 / name :op1 "Akt"~e.2))) :ARG1 (a5 / and~e.6 :op1 (p / proliferate-01~e.5) :op2 (r2 / resist-01~e.7 :ARG1~e.8 (d / die-01~e.11 :ARG1 (c / cell~e.10) :mod (a6 / apoptosis~e.9))))) :op2 (a2 / activity-06~e.15 :ARG0 a4 :degree (m / more~e.14) :ARG1-of (d2 / describe-01 :ARG0 (p2 / publication-91 :ARG1-of (c3 / cite-01 :ARG2 (a7 / and~e.33 :op1 21~e.25 :op2 35~e.29)))) :location~e.16 (t / tumor~e.18 :mod (l / lung~e.17) :compared-to~e.19 (t2 / tissue~e.22 :ARG1-of (n3 / normal-02~e.21)))) :op3 (a3 / activate-01~e.35 :ARG0~e.36 (c2 / coculture-01~e.38,39,40 :ARG1 (m2 / macrophage~e.37)) :ARG1 a4)) # ::id a_pmid_2169_9731.102 ::amr-annotator SDL-AMR-09 ::preferred # ::tok Since combined MEK and PI3K inhibition slowed mutant Kras @- @ driven lung tumor growth in vivo @ [ @ 25 @ ] , we determined whether selective inhibition of MEK and PI3K affected macrophage @-@ stimulated proliferation in these Kras @ mutant lung tumor cell lines . # ::alignments 0-1.3 1-1.3.1.1.1 2-1.2.2.1.1.1.1 3-1.2.2.1 4-1.2.2.1.2.1.1 5-1.3.1.1 6-1.3.1 7-1.3.1.2.1.2.1 12-1.3.1.2.1.2 13-1.3.1.2.1.1 14-1.3.1.2.1 15-1.3.1.2 17-1.3.1.2.2 18-1.3.1.2.2 22-1.3.1.3.1.1.1 26-1.1 27-1 28-1.2.1 28-1.2.1.r 29-1.2.2.2 30-1.2.2 32-1.2.2.1.1.1.1 33-1.2.2.1 34-1.2.2.1.2.1.1 35-1.2 36-1.2.3.1.1 38-1.2.3.1 39-1.2.3 40-1.3.1.2.2 45-1.2.3.2.1 45-1.2.3.2.1.2 45-1.2.3.2.1.2.r 46-1.2.3.2.3 47-1.2.3.2.2 48-1.2.3.2 49-1.2.3.2 (d / determine-01~e.27 :ARG0 (w / we~e.26) :ARG1 (a / affect-01~e.35 :mode~e.28 interrogative~e.28 :ARG0 (i / inhibit-01~e.30 :ARG1 (a2 / and~e.3,33 :op1 (e / enzyme :name (n / name :op1 "MEK"~e.2,32)) :op2 (e2 / enzyme :name (n2 / name :op1 "PI3K"~e.4,34))) :manner (s / selective~e.29)) :ARG1 (p / proliferate-01~e.39 :ARG1-of (s2 / stimulate-01~e.38 :ARG0 (m / macrophage~e.36)) :location (c / cell-line~e.48,49 :mod (g2 / gene~e.45 :name (n3 / name :op1 "K-Ras") :ARG2-of~e.45 (m2 / mutate-01~e.45)) :mod (t / tumor~e.47) :mod (l / lung~e.46)))) :ARG1-of (c2 / cause-01~e.0 :ARG0 (s3 / slow-01~e.6 :ARG0 (i4 / inhibit-01~e.5 :ARG3-of (c4 / combine-01~e.1)) :ARG1 (g / grow-01~e.15 :ARG1 (t2 / tumor~e.14 :mod (l2 / lung~e.13) :ARG1-of (d2 / drive-02~e.12 :ARG0 g2~e.7)) :manner (i2 / in-vivo~e.17,18,40)) :ARG1-of (d3 / describe-01 :ARG0 (p2 / publication-91 :ARG1-of (c3 / cite-01 :ARG2 25~e.22)))))) # ::id a_pmid_2169_9731.103 ::amr-annotator SDL-AMR-09 ::preferred # ::tok Selective inhibition of either MEK ( by U0126 ) or PI3K ( by LY294002 ) significantly decreased basal proliferation , and blocked growth stimulated by macrophage co @-@ culture to different extents in LM2 and JF32 cells ( Figure 4A and 4B , respectively ) . # ::alignments 0-1.1.1.3 1-1.1.1.1 1-1.1.1.2 4-1.1.1.1.2.1.1 6-1.1.1.1.1.r 7-1.1.1.1.1.1.1 9-1.1.1 10-1.1.1.2.2.1.1 12-1.1.1.2.1.r 13-1.1.1.2.1.1.1 15-1.1.3 16-1.1 17-1.1.2.1 18-1.1.2 20-1 21-1.2 22-1.2.2 23-1.2.2.1 24-1.2.2.1.1.r 25-1.2.2.1.1.1 26-1.2.2.1.1 27-1.2.2.1.1 28-1.2.2.1.1 29-1.2.2.1.2.r 30-1.2.2.1.2.1 31-1.2.2.1.2 32-1.2.3.r 33-1.2.3.1.1.1 34-1.2.3 35-1.2.3.2.1.1 36-1.2.3.1 36-1.2.3.2 38-1.3.1.1 38-1.3.1.2 40-1.3.1.1.1 42-1.3.1 44-1.3.1.2.1 47-1.3.2 (a / and~e.20 :op1 (d / decrease-01~e.16 :ARG0 (o2 / or~e.9 :op1 (i2 / inhibit-01~e.1 :ARG0~e.6 (s2 / small-molecule :name (n3 / name :op1 "U0126"~e.7)) :ARG1 (e / enzyme :name (n / name :op1 "MEK"~e.4))) :op2 (i3 / inhibit-01~e.1 :ARG0~e.12 (s3 / small-molecule :name (n4 / name :op1 "LY294002"~e.13)) :ARG1 (e2 / enzyme :name (n2 / name :op1 "PI3K"~e.10))) :manner (s / selective~e.0)) :ARG1 (p / proliferate-01~e.18 :mod (b2 / basal~e.17)) :ARG2 (s4 / significant-02~e.15)) :op2 (b / block-01~e.21 :ARG0 o2 :ARG1 (g / grow-01~e.22 :ARG1-of (s5 / stimulate-01~e.23 :ARG0~e.24 (c / coculture-01~e.26,27,28 :ARG1 (m / macrophage~e.25)) :degree~e.29 (e3 / extent~e.31 :ARG1-of (d2 / differ-02~e.30)))) :location~e.32 (a2 / and~e.34 :op1 (c2 / cell-line~e.36 :name (n5 / name :op1 "LM2"~e.33)) :op2 (c3 / cell-line~e.36 :name (n6 / name :op1 "JF32"~e.35)))) :ARG1-of (d3 / describe-01 :ARG0 (a3 / and~e.42 :op1 (f / figure~e.38 :mod "4A"~e.40) :op2 (f2 / figure~e.38 :mod "4B"~e.44)) :mod (r / respective~e.47))) # ::id a_pmid_2169_9731.104 ::amr-annotator SDL-AMR-09 ::preferred # ::tok Only the combined inhibition of both kinases ablated the stimulatory effect of macrophage co @-@ culture on neoplastic proliferation ( U0 + LY , Figure 4 ) . # ::alignments 0-1.2.4 2-1.2.3 3-1.2 4-1.2.2.r 5-1.2.2.1 6-1.2.2 7-1 10-1.1 11-1.1.1.r 12-1.1.1.1 13-1.1.1 14-1.1.1 15-1.1.1 16-1.1.2.r 17-1.1.2.1 18-1.1.2 20-1.2.3.1.1.1.1.1 21-1.2.3.1.1 22-1.2.3.1.1.2.1.1 24-1.2.3.1.2.1 26-1.2.3.1.2.1.1 (a / ablate-01~e.7 :ARG1 (a2 / affect-01~e.10 :ARG0~e.11 (c / coculture-01~e.13,14,15 :ARG1 (m / macrophage~e.12)) :ARG1~e.16 (p / proliferate-01~e.18 :ARG0 (n / neoplasm~e.17)) :ARG2 (s / stimulate-01)) :ARG3 (i / inhibit-01~e.3 :ARG0 a3 :ARG1~e.4 (k / kinase~e.6 :mod (b / both~e.5)) :ARG3-of (c2 / combine-01~e.2 :ARG1-of (m2 / mean-01 :ARG2 (a3 / and~e.21 :op1 (s3 / small-molecule :name (n2 / name :op1 "U0"~e.20)) :op2 (s4 / small-molecule :name (n3 / name :op1 "LY"~e.22))) :ARG1-of (d / describe-01 :ARG0 (f / figure~e.24 :mod 4~e.26)))) :mod (o / only~e.0))) # ::id a_pmid_2169_9731.105 ::amr-annotator SDL-AMR-09 ::preferred # ::tok Kinase inhibitors were applied at concentrations reported to be cytostatic and not cytotoxic [ @ 34 , 36 , 37 @ ] , and none of these treatments significantly increased LM2 or JF32 cell death ( data not shown ) . # ::alignments 0-1.1.1.1.1.1 1-1.1.1.1 1-1.1.1.1.1 1-1.1.1.1.1.r 3-1.1 5-1.1.1 6-1.1.1.2 9-1.1.1.3 11-1.1.1.4.1 11-1.1.1.4.1.r 12-1.1.1.4 15-1.1.2.1.1.1.1 19-1.1.2.1.1.1.2 23-1.1.2.1.1.1.3 27-1 27-1.1.2.1.1.1 28-1.2.1.2 29-1.2.1.2.r 30-1.2.1.1 31-1.2.1 32-1.2.3 33-1.2 34-1.2.2.1.1.1.1 35-1.2.2.1 36-1.2.2.1.2.1.1 37-1.2.2.1.1 37-1.2.2.1.2 38-1.2.2 40-1.3.1 41-1.3.1.1.1 41-1.3.1.1.1.r 42-1.3.1.1 (a / and~e.27 :op1 (a2 / apply-02~e.3 :ARG1 (c / concentrate-02~e.5 :ARG1 (m / molecular-physical-entity~e.1 :ARG0-of~e.1 (i2 / inhibit-01~e.1 :ARG1 (k / kinase~e.0))) :ARG1-of (r / report-01~e.6) :mod (c2 / cytostatic~e.9) :mod (c3 / cytotoxic~e.12 :polarity~e.11 -~e.11)) :ARG1-of (d4 / describe-01 :ARG0 (p / publication-91 :ARG1-of (c6 / cite-01 :ARG2 (a3 / and~e.27 :op1 34~e.15 :op2 36~e.19 :op3 37~e.23))))) :op2 (i / increase-01~e.33 :ARG0 (t / treatment~e.31 :mod (t2 / this~e.30) :quant~e.29 (n / none~e.28)) :ARG1 (d3 / die-01~e.38 :ARG1 (o / or~e.35 :op1 (c4 / cell-line~e.37 :name (n2 / name :op1 "LM2"~e.34)) :op2 (c5 / cell-line~e.37 :name (n3 / name :op1 "JF32"~e.36)))) :ARG2 (s2 / significant-02~e.32)) :ARG1-of (d / describe-01 :ARG0 (d2 / data~e.40 :ARG1-of (s / show-01~e.42 :polarity~e.41 -~e.41)))) # ::id a_pmid_2169_9731.106 ::amr-annotator SDL-AMR-09 ::preferred # ::tok These results suggest that both the MEK and PI3K pathways must be blocked to effectively inhibit macrophage @-@ stimulated neoplastic growth . # ::alignments 0-1.3.1.2 1-1.3.1 1-1.3.1.1 1-1.3.1.1.r 2-1.3 6-1.1.1.1.1 7-1.1 8-1.1.2.1.1 9-1.1.1 9-1.1.2 10-1 12-1.2 14-1.2.2.3 15-1.2.2 16-1.2.2.2.2.1 18-1.2.2.2.2 19-1.2.2.2.1 20-1.2.2.2 (o / obligate-01~e.10 :ARG1 (a / and~e.7 :op1 (p / pathway~e.9 :name (n / name :op1 "MEK"~e.6)) :op2 (p2 / pathway~e.9 :name (n2 / name :op1 "PI3K"~e.8))) :ARG2 (b / block-01~e.12 :ARG1 a :purpose (i / inhibit-01~e.15 :ARG0 a :ARG1 (g / grow-01~e.20 :ARG1 (n3 / neoplasm~e.19) :ARG1-of (s2 / stimulate-01~e.18 :ARG0 (m / macrophage~e.16))) :ARG1-of (e / effective-04~e.14))) :ARG1-of (s / suggest-01~e.2 :ARG0 (t2 / thing~e.1 :ARG2-of~e.1 (r / result-01~e.1) :mod (t / this~e.0)))) # ::id a_pmid_2169_9731.107 ::amr-annotator SDL-AMR-09 ::preferred # ::tok Macrophage conditioned media contains 3 @-@ 10 kDa factors which stimulate neoplastic proliferation # ::alignments 0-1.1.1.1 1-1.1.1 2-1.1 3-1 4-1.2.2.1.1 6-1.2.2.2.1 7-1.2.2.1.2 7-1.2.2.2.2 8-1.2 10-1.2.1 11-1.2.1.1.1 12-1.2.1.1 (c / contain-01~e.3 :ARG0 (m / medium~e.2 :ARG1-of (c2 / condition-01~e.1 :ARG2 (m2 / macrophage~e.0))) :ARG1 (f / factor~e.8 :ARG0-of (s / stimulate-01~e.10 :ARG1 (p / proliferate-01~e.12 :ARG0 (n / neoplasm~e.11))) :quant (b / between :op1 (m3 / mass-quantity :quant 3~e.4 :unit (k3 / kilodalton~e.7)) :op2 (m4 / mass-quantity :quant 10~e.6 :unit (k4 / kilodalton~e.7))))) # ::id a_pmid_2169_9731.108 ::amr-annotator SDL-AMR-09 ::preferred # ::tok Macrophages produce numerous cytokines , eicosanoids and other soluble factors depending upon tissue location and environmental stimuli [ @ 4 , 18 @ ] , any number of which could be responsible for the observed neoplastic growth stimulation described above . # ::alignments 0-1.1 1-1 2-1.2.1.1 3-1.2.1 6-1.2 7-1.2.3.2 8-1.2.3.1 9-1.2.3 10-1.3 12-1.3.1.1.1 13-1.3.1.1 14-1.3.1 15-1.3.1.2.1 16-1.3.1.2 19-1.4.1.1.1.1 23-1.4.1.1.1.2 27-1.2.5.1 28-1.2.5 31-1.2.4.2 33-1.2.4 34-1.2.4.1.r 36-1.2.4.1.1.2 37-1.2.4.1.1.1 38-1.2.4.1.1 39-1.2.4.1 40-1.2.4.1.2 40-1.4 41-1.2.4.1.2.1 (p / produce-01~e.1 :ARG0 (m / macrophage~e.0) :ARG1 (a / and~e.6 :op1 (c / cytokine~e.3 :quant (n / numerous~e.2)) :op2 (e / eicosanoid) :op3 (f / factor~e.9 :mod (s / soluble~e.8) :mod (o / other~e.7)) :ARG0-of (r / responsible-01~e.33 :ARG1~e.34 (s3 / stimulate-01~e.39 :ARG1 (g / grow-01~e.38 :ARG1 (n2 / neoplasm~e.37) :ARG1-of (o2 / observe-01~e.36)) :ARG1-of (d2 / describe-01~e.40 :location (a4 / above~e.41))) :ARG1-of (p2 / possible-01~e.31)) :quant (n3 / number~e.28 :mod (a3 / any~e.27))) :ARG0-of (d / depend-01~e.10 :ARG1 (a2 / and~e.14 :op1 (l / location~e.13 :mod (t / tissue~e.12)) :op2 (s2 / stimulus~e.16 :mod (e2 / environment~e.15)))) :ARG1-of (d3 / describe-01~e.40 :ARG0 (p3 / publication-91 :ARG1-of (c2 / cite-01 :ARG2 (a5 / and :op1 4~e.19 :op2 18~e.23))))) # ::id a_pmid_2169_9731.109 ::amr-annotator SDL-AMR-09 ::preferred # ::tok Media conditioned by primary BAL macrophages ( MØCM ) stimulated the proliferation of LM2 cells , albeit to a lesser extent than primary macrophage co @-@ culture ( Figure 5 " Total " vs. Figure 2B ) . # ::alignments 0-1.1 1-1.1.1 2-1.1.1.1.r 3-1.1.1.1.2 4-1.1.1.1.1 4-1.1.1.1.1.1 4-1.1.1.1.1.1.r 5-1.1.1.1 9-1 9-1.4 9-1.4.r 11-1.2 12-1.2.1.r 13-1.2.1.1.1 14-1.2.1 17-1.4.3.r 19-1.4.3 19-1.4.3.1 19-1.4.3.1.r 21-1.4.5.r 22-1.4.5.1.1 23-1.4.5.1 24-1.4.5 25-1.4.5 26-1.4.5 28-1.3.1 30-1.3.1.1 33-1.3.2 35-1.4.5.r 36-1.4.4.1 38-1.4.4.1.1 (s / stimulate-01~e.9 :ARG0 (m / medium~e.0 :ARG1-of (c / condition-01~e.1 :ARG0~e.2 (m2 / macrophage~e.5 :source (a / alveolus~e.4 :mod~e.4 (b / bronchus~e.4)) :mod (p / primary~e.3)))) :ARG1 (p2 / proliferate-01~e.11 :ARG0~e.12 (c2 / cell-line~e.14 :name (n2 / name :op1 "LM2"~e.13))) :ARG1-of (d / describe-01 :ARG0 (f / figure~e.28 :mod 5~e.30) :ARG2 (t / total~e.33)) :concession~e.9 (s2 / stimulate-01~e.9 :ARG0 m :ARG1 p2 :degree~e.17 (l / less~e.19 :degree~e.19 (m4 / more~e.19)) :ARG1-of (d2 / describe-01 :ARG0 (f2 / figure~e.36 :mod "2B"~e.38)) :compared-to~e.21,35 (c3 / coculture-01~e.24,25,26 :ARG1 (m5 / macrophage~e.23 :mod (p3 / primary~e.22))))) # ::id a_pmid_2169_9731.110 ::amr-annotator SDL-AMR-09 ::preferred # ::tok When size @-@ fractionated MØCM was added to LM2 cells , molecules between 3 and 10 kDa stimulated LM2 growth to the greatest extent ( Figure 5 ) . # ::alignments 1-1.4.1.1.1 3-1.4.1.1 6-1.4 7-1.4.2.r 8-1.4.2 9-1.4.2 11-1.1 12-1.1.1.1 13-1.1.1.1.1 14-1.1.1.1 15-1.1.1.1.2 16-1.1.1.2 17-1 18-1.2.1.1.1 19-1.2 20-1.3.r 22-1.3 22-1.3.1 22-1.3.1.r 25-1.5.1 27-1.5.1.1 (s / stimulate-01~e.17 :ARG0 (m2 / molecule~e.11 :mod (m3 / mass-quantity :quant (b / between~e.12,14 :op1 3~e.13 :op2 10~e.15) :unit (k / kilodalton~e.16))) :ARG1 (g2 / grow-01~e.19 :ARG1 (c / cell-line :name (n / name :op1 "LM2"~e.18))) :degree~e.20 (g / great~e.22 :degree~e.22 (m / most~e.22)) :condition (a / add-02~e.6 :ARG1 (m4 / medium :ARG1-of (f / fractionate-00~e.3 :mod (s2 / size~e.1)) :ARG1-of (c2 / condition-01 :ARG0 (m5 / macrophage))) :ARG2~e.7 c~e.8,9) :ARG1-of (d / describe-01 :ARG0 (f2 / figure~e.25 :mod 5~e.27))) # ::id a_pmid_2169_9731.111 ::amr-annotator SDL-AMR-09 ::preferred # ::tok Thus , factors of this size mediated the majority of MØCM effects on LM2 growth . # ::alignments 2-1.1.1 3-1.1.1.1.r 4-1.1.1.1.1 5-1.1.1.1 6-1.1 8-1.1.2.3 9-1.1.2.3.r 11-1.1.2 12-1.1.2.2.r 13-1.1.2.2.1.1.1 14-1.1.2.2 (i / infer-01 :ARG1 (m / mediate-01~e.6 :ARG0 (f / factor~e.2 :mod~e.3 (s / size~e.5 :mod (t / this~e.4))) :ARG1 (a / affect-01~e.11 :ARG0 (m3 / medium :ARG1-of (c2 / condition-01 :ARG0 (m4 / macrophage))) :ARG1~e.12 (g / grow-01~e.14 :ARG0 (c / cell-line :name (n2 / name :op1 "LM2"~e.13))) :quant~e.9 (m2 / majority~e.8)))) # ::id a_pmid_2169_9731.112 ::amr-annotator SDL-AMR-09 ::preferred # ::tok Alveolar macrophages produce numerous growth factors in this size range , including IGF @-@ 1 , GM @-@ CSF and EGF [ @ 11 , 18 @ ] . # ::alignments 0-1.1.1 1-1.1 2-1 3-1.2.2 4-1.2 5-1.2 6-1.3.r 7-1.3.2 8-1.3.1 9-1.3 11-1.2.1 12-1.2.1.1.1.1.1 14-1.2.1.1.1.1.1 16-1.2.1.1.2.1.1 18-1.2.1.1.2.1.1 19-1.2.1.1 20-1.2.1.1.3.1.1 23-1.4.1.1.1.1 27-1.4.1.1.1.2 (p / produce-01~e.2 :ARG0 (m / macrophage~e.1 :mod (a / alveolus~e.0)) :ARG1 (g / growth-factor~e.4,5 :ARG2-of (i / include-01~e.11 :ARG1 (a2 / and~e.19 :op1 (p5 / protein :name (n2 / name :op1 "IGF-1"~e.12,14)) :op2 (p4 / protein :name (n3 / name :op1 "GM-CSF"~e.16,18)) :op3 (p3 / protein :name (n4 / name :op1 "EGF"~e.20)))) :quant (n5 / numerous~e.3)) :prep-in~e.6 (r / range~e.9 :mod (s2 / size~e.8) :mod (t / this~e.7)) :ARG1-of (d / describe-01 :ARG0 (p2 / publication :ARG1-of (c / cite-01 :ARG2 (a3 / and :op1 11~e.23 :op2 18~e.27))))) # ::id a_pmid_2169_9731.113 ::amr-annotator SDL-AMR-09 ::preferred # ::tok To further narrow down the list of possible candidates , an in silico @ analysis was performed for each fraction size as described in Materials and Methods . # ::alignments 1-1.2.2 2-1.2 3-1.2 5-1.2.1 6-1.2.1.1.r 7-1.2.1.1.1 8-1.2.1.1 12-1.1.2 13-1.1.2 15-1.1 17-1 18-1.1.1.r 19-1.1.1.2 20-1.1.1.1 21-1.1.1 22-1.3.r 23-1.3 24-1.1.2 25-1.3.1.1.1 26-1.3.1.1.2 27-1.3.1.1.3 (p / perform-02~e.17 :ARG1 (a / analyze-01~e.15 :ARG1~e.18 (s / size~e.21 :mod (f / fraction~e.20) :mod (e / each~e.19)) :manner (i / in-silico~e.12,13,24)) :purpose (n / narrow-down-03~e.2,3 :ARG1 (l / list~e.5 :consist-of~e.6 (c / candidate~e.8 :ARG1-of (p2 / possible-01~e.7))) :degree (f2 / further~e.1)) :ARG1-of~e.22 (d / describe-01~e.23 :ARG0 (s2 / section :name (n2 / name :op1 "Materials"~e.25 :op2 "and"~e.26 :op3 "Methods"~e.27)))) # ::id a_pmid_2169_9731.114 ::amr-annotator SDL-AMR-09 ::preferred # ::tok The resulting data points were separately fit for each fraction size to the general equation y @ = y @ 0 @ + a @ ( 1 @- @ e @ -@ bx @ ) as described , with regression r @ 2 @ = 0.997 , 0.842 and 0.918 for the > 3 , > 10 and > 30 kDa fractions , respectively . # ::alignments 1-1.1.2 2-1.1.1 3-1.1 5-1.3 6-1 7-1.4.r 8-1.4.2 9-1.4.1 10-1.4 11-1.2.r 13-1.2.1 14-1.2 25-1.5 33-1.2.2.1 40-1.6.r 41-1.6 50-1.5.1.1 52-1.5.2.1 53-1.5 54-1.5.3.1 57-1.5.1.2.1 58-1.5.1.2.1.1.1 60-1.5.1.2.1 61-1.5.2.2.1.1.1 62-1.5 63-1.5.1.2.1 63-1.5.2.2.1 63-1.5.3.2.1 64-1.5.3.2.1.1.1 65-1.5.1.2.1.1.2 65-1.5.2.2.1.1.2 65-1.5.3.2.1.1.2 66-1.5.1.2 66-1.5.2.2 66-1.5.3.2 (f / fit-06~e.6 :ARG1 (p / point~e.3 :mod (d / data~e.2) :ARG1-of (r / result-01~e.1)) :ARG2~e.11 (e2 / equation~e.14 :ARG1-of (g / general-02~e.13) :mod (s6 / string-entity :value "y=y0+a(1-e-bx)"~e.33)) :manner (s / separate-02~e.5) :beneficiary~e.7 (s2 / size~e.10 :mod (f2 / fraction~e.9) :mod (e / each~e.8)) :condition (a / and~e.25,53,62 :op1 (s3 / statistical-test-91 :ARG3 0.997~e.50 :ARG1 (f3 / fraction~e.66 :mod (m4 / more-than~e.57,60,63 :op1 (m / mass-quantity :quant 3~e.58 :unit (k / kilodalton~e.65))))) :op2 (s4 / statistical-test-91 :ARG3 0.842~e.52 :ARG1 (f4 / fraction~e.66 :mod (m5 / more-than~e.63 :op1 (m2 / mass-quantity :quant 10~e.61 :unit (k2 / kilodalton~e.65))))) :op3 (s5 / statistical-test-91 :ARG3 0.918~e.54 :ARG1 (f5 / fraction~e.66 :mod (m6 / more-than~e.63 :op1 (m3 / mass-quantity :quant 30~e.64 :unit (k3 / kilodalton~e.65)))))) :ARG1-of~e.40 (d2 / describe-01~e.41)) # ::id a_pmid_2169_9731.115 ::amr-annotator SDL-AMR-09 ::preferred # ::tok From regression analysis , the responsible factor( s ) appeared to be 7.23 @-@ 10.8 kDa in size , suggesting that growth factors such as IGF @-@ 1 ( 7.5 kDa ) may be responsible for the MØCM @-@ stimulated neoplastic proliferation . # ::alignments 2-1.2.1 5-1.1.3.1.1 9-1 12-1.1.2.1.2.1 14-1.1.2.1.2.2 15-1.1.2.1.1 17-1.1.2 19-1.1.3 21-1.1.3.1.1.1 22-1.1.1 22-1.1.3.1.1.1 23-1.1.3.1.1.1.1.r 24-1.1.3.1.1.1.1.r 25-1.1.3.1.1.1.1.1.1 27-1.1.3.1.1.1.1.1.1 29-1.1.3.1.1.1.1.2.1 30-1.1.3.1.1.1.1.2.2 32-1.1.3.1 34-1.1.1.1 34-1.1.3.1.1 39-1.1.3.1.1.2.2 41-1.1.3.1.1.2 (a / appear-02~e.9 :ARG1 (h / have-03 :ARG0 (f / factor~e.22 :ARG0-of (r / responsible-01~e.34)) :ARG1 (s / size~e.17 :mod (m / mass-quantity :unit (k / kilodalton~e.15) :quant (b / between :op1 7.23~e.12 :op2 10.8~e.14))) :ARG0-of (s2 / suggest-01~e.19 :ARG1 (p / possible-01~e.32 :ARG1 (r3 / responsible-01~e.5,34 :ARG0 (g / growth-factor~e.21,22 :example~e.23,24 (p3 / protein :name (n2 / name :op1 "IGF-1"~e.25,27) :mod (m2 / mass-quantity :quant 7.5~e.29 :unit (k2 / kilodalton~e.30)))) :ARG1 (p2 / proliferate-01~e.41 :ARG0 (t / tumor) :ARG1-of (s4 / stimulate-01~e.39 :ARG0 (m3 / medium :ARG1-of (c / condition-01 :ARG0 (m4 / macrophage))))))))) :ARG1-of (d2 / deduce-01 :ARG2 (a2 / analysis~e.2 :mod (r2 / regress-01)))) # ::id a_pmid_2169_9731.116 ::amr-annotator SDL-AMR-09 ::preferred # ::tok Macrophage @-@ conditioned media IGF @-@ 1 levels correlate to effects on neoplastic proliferation # ::alignments 0-1.1.2.1.1 2-1.1.2.1 3-1.1.2 4-1.1.1.1.1 6-1.1.1.1.1 7-1.1 8-1 9-1.2.r 10-1.2 13-1.2.1 (c / correlate-01~e.8 :ARG1 (l / level~e.7 :quant-of (p2 / protein :name (n / name :op1 "IGF-1"~e.4,6)) :source (m / medium~e.3 :ARG1-of (c2 / condition-01~e.2 :ARG0 (m2 / macrophage~e.0)))) :ARG2~e.9 (a / affect-01~e.10 :ARG1 (p / proliferate-01~e.13 :ARG0 (t / tumor)))) # ::id a_pmid_2169_9731.117 ::amr-annotator SDL-AMR-09 ::preferred # ::tok IGF @-@ 1 has a well @-@ established role in the metastasis of cancer cells in vivo , as well as stimulating growth in vitro @ [ @ 27 @ ] , and alveolar macrophages produce high levels of IGF @-@ 1 in response to quartz dust @-@ induced lung injury [ @ 30 @ ] . # ::alignments 0-1.1.1.1.1 2-1.1.1.1.1 3-1.1 5-1.1.2.1.1 7-1.1.2.1 8-1.1.2 9-1.1.2.2.r 11-1.1.2.2.1 12-1.1.2.2.1.1.r 13-1.1.2.2.1.1.1.2.1 14-1.1.2.2.1.1 16-1.1.2.2.1.2.r 16-1.1.2.2.r 17-1.1.2.2.1.2 20-1.1.2.2 21-1.1.2.2 22-1.1.2.2 23-1.1.2.2.2 24-1.1.2.2.2.2 26-1.1.2.2.2.3 27-1.1.2.2.2.3 31-1.1.2.3.1.1.1 35-1 36-1.2.1.1 37-1.2.1 38-1.2 39-1.2.2.1 40-1.2.2 41-1.2.2.2.r 42-1.2.2.2 43-1.2.2.2 44-1.2.2.2 45-1.1.2.2.r 45-1.2.4.r 46-1.2.4 47-1.2.4.1.r 48-1.2.4.1.2.1.1.1 49-1.2.4.1.2.1.1.2 51-1.2.4.1.2 52-1.2.4.1.1 53-1.2.4.1 56-1.2.3.1.1.1 (a / and~e.35 :op1 (h / have-03~e.3 :ARG0 (p4 / protein :name (n2 / name :op1 "IGF-1"~e.0,2)) :ARG1 (r / role~e.8 :ARG1-of (e2 / establish-01~e.7 :mod (w / well~e.5)) :prep-in~e.9,16,45 (a2 / and~e.20,21,22 :op1 (m / metastasize-101~e.11 :ARG1~e.12 (c / cell~e.14 :mod (d / disease :wiki "Cancer" :name (n / name :op1 "cancer"~e.13))) :manner~e.16 (i / in-vivo~e.17)) :op2 (s / stimulate-01~e.23 :ARG0 p4 :ARG1 (g / grow-01~e.24) :manner (i2 / in-vitro~e.26,27))) :ARG1-of (d2 / describe-01 :ARG0 (p / publication :ARG1-of (c3 / cite-01 :ARG2 27~e.31))))) :op2 (p2 / produce-01~e.38 :ARG0 (m2 / macrophage~e.37 :mod (a3 / alveolus~e.36)) :ARG1 (l / level~e.40 :ARG1-of (h2 / high-02~e.39) :quant-of~e.41 p4~e.42,43,44) :ARG1-of (d3 / describe-01 :ARG0 (p3 / publication :ARG1-of (c4 / cite-01 :ARG2 30~e.56))) :ARG2-of~e.45 (r2 / respond-01~e.46 :ARG1~e.47 (i3 / injure-01~e.53 :ARG1 (l2 / lung~e.52) :ARG2-of (i4 / induce-01~e.51 :ARG0 (s2 / small-molecule :name (n3 / name :op1 "quartz"~e.48 :op2 "dust"~e.49))))))) # ::id a_pmid_2169_9731.118 ::amr-annotator SDL-AMR-09 ::preferred # ::tok While alveolar macrophages are an important component of the chronic inflammatory milieu responsible for promoting lung tumorigenesis , IGF @-@ 1 has not been examined as a possible connection between macrophage recruitment and lung cancer progression . # ::alignments 0-1 1-1.2.3.1 2-1.2.3 3-1.2.3.r 5-1.2.1 6-1.2 9-1.2.2.1.1 10-1.2.2.1.2 11-1.2.2.1 12-1.2.2.1.3 13-1.2.2.1.3.1.r 14-1.2.2.1.3.1 15-1.2.2.1.3.1.1.1.1 16-1.2.2.1.3.1.1 16-1.2.2.1.3.1.1.1 16-1.2.2.1.3.1.1.1.r 18-1.1.2.1.1 20-1.1.2.1.1 22-1.1.1 22-1.1.1.r 24-1.1 25-1.1.3.r 27-1.1.3.4 28-1.1.3 30-1.1.3.2.1 31-1.1.3.2 33-1.1.3.3.1.2.1 34-1.1.3.3.1.2.2 35-1.1.3.3 (c / contrast-01~e.0 :ARG1 (e / examine-01~e.24 :polarity~e.22 -~e.22 :ARG1 (p / protein :name (n2 / name :op1 "IGF-1"~e.18,20)) :ARG2~e.25 (c2 / connect-01~e.28 :ARG0 p :ARG1 (r / recruit-01~e.31 :ARG1 (m / macrophage~e.30)) :ARG1 (p3 / progress-01~e.35 :ARG1 (d / disease :wiki "Lung_cancer" :name (n / name :op1 "lung"~e.33 :op2 "cancer"~e.34))) :ARG1-of (p2 / possible-01~e.27))) :ARG2 (c4 / component~e.6 :mod (i3 / important~e.5) :ARG1-of (i / include-91 :ARG2 (m3 / milieu~e.11 :mod (c3 / chronic~e.9) :mod (i2 / inflame-01~e.10) :ARG0-of (r2 / responsible-01~e.12 :ARG1~e.13 (p4 / promote-01~e.14 :ARG1 (c5 / create-01~e.16 :ARG1~e.16 (t2 / tumor~e.16 :mod (l / lung~e.15))))))) :domain~e.3 (m2 / macrophage~e.2 :mod (a / alveolus~e.1)))) # ::id a_pmid_2169_9731.119 ::amr-annotator SDL-AMR-09 ::preferred # ::tok BALF from tumor @-@ bearing lungs contained 3.5 @-@ times more IGF @-@ 1 than BALF from naïve mice , while EGF levels were unchanged ( Figure 6A ) . # ::alignments 0-1.1.1 0-1.1.3.2.1 0-1.1.3.2.1.1 0-1.1.3.2.1.1.r 1-1.1.1.1.r 2-1.1.1.1.1.1 4-1.1.1.1.1 5-1.1.1.1 6-1.1 7-1.1.2.2.1.1 9-1.1.2.2.1 10-1.1.2.2 11-1.1.2.1.1 13-1.1.2.1.1 14-1.1.3.r 15-1.1.3 15-1.1.3.2 15-1.1.3.2.1 15-1.1.3.2.1.1 15-1.1.3.2.1.1.r 15-1.1.3.2.1.r 15-1.1.3.2.r 16-1.1.1.1.r 16-1.1.3.1.r 17-1.1.3.1.1 18-1.1.3.1 20-1 21-1.2.2.1.1.1 22-1.2.2 24-1.2 24-1.2.1 24-1.2.1.r 26-1.3.1 28-1.3.1.1 (c / contrast-01~e.20 :ARG1 (c3 / contain-01~e.6 :ARG0 (f3 / fluid~e.0 :source~e.1,16 (l2 / lung~e.5 :ARG0-of (b / bear-01~e.4 :ARG1 (t / tumor~e.2))) :mod l3) :ARG1 (p2 / protein :name (n3 / name :op1 "IGF-1"~e.11,13) :quant (m2 / more~e.10 :degree (p3 / product-of~e.9 :op1 3.5~e.7))) :compared-to~e.14 (f2 / fluid~e.15 :part-of~e.16 (m4 / mouse~e.18 :mod (n5 / naive~e.17)) :mod~e.15 (l3 / lavage~e.15 :mod~e.15 (a / alveolus~e.0,15 :mod~e.0,15 (b2 / bronchus~e.0,15))))) :ARG2 (c2 / change-01~e.24 :polarity~e.24 -~e.24 :ARG1 (l / level~e.22 :quant-of (p / protein :name (n / name :op1 "EGF"~e.21)))) :ARG1-of (d / describe-01 :ARG0 (f / figure~e.26 :mod "6A"~e.28))) # ::id a_pmid_2169_9731.120 ::amr-annotator SDL-AMR-09 ::preferred # ::tok Even after normalizing to total BALF protein levels , BALF IGF @-@ 1 was significantly higher in tumor @-@ bearing animals than naïve controls ( 1.81 ± 0.33 vs. 0.95 ± 0.36 pg IGF @-@ 1/ug BALF protein , respectively , P @ < 0.01 , mean ± SD ) , suggesting that more IGF @-@ 1 is produced in the lungs of tumor @-@ bearing mice . # ::alignments 2-1.7 3-1.7.2.r 4-1.7.2.2 5-1.1.1.2 5-1.1.1.2.1 5-1.1.1.2.1.1 5-1.1.1.2.1.1.1 5-1.1.1.2.1.1.1.r 5-1.1.1.2.1.1.r 5-1.1.1.2.1.r 6-1.7.2.1 7-1.1 7-1.2 7-1.7.2 9-1.1.1.2.1.1 9-1.1.1.2.1.1.1 9-1.1.1.2.1.1.1.r 10-1.1.1.1.1 12-1.1.1.1.1 14-1.6 15-1 15-1.5 15-1.5.r 16-1.3.r 17-1.3.1.1 19-1.3.1 20-1.3 21-1.2.r 22-1.2.1.2.1 23-1.2.1.2 25-1.1.2.1.2.1.2 25-1.1.2.1.2.2.2 27-1.1.2.1.2.1.1 27-1.1.2.1.2.2.1 28-1.2.r 29-1.2.2.1.2.1.2 29-1.2.2.1.2.2.2 31-1.2.2.1.2.1.1 31-1.2.2.1.2.2.1 33-1.1.1.1.1 36-1.1.1.2 36-1.1.1.2.1 36-1.1.1.2.1.1 36-1.1.1.2.1.1.1 36-1.1.1.2.1.1.1.r 36-1.1.1.2.1.1.r 36-1.1.1.2.1.r 37-1.1.1 42-1.1.2.2.1 44-1.1.2.2.1.1 45-1.1.2.2.1.1.1 47-1.1.2.2.2.1 52-1.4 54-1.4.1.2 54-1.5 55-1.2.1.1.1 57-1.2.1.1.1 59-1.4.1 60-1.4.1.3.r 62-1.4.1.3 63-1.1.2.1.1 63-1.4.1.3.1.r 64-1.4.1.3.1.1 65-1.4.1.3.1.1 66-1.4.1.3.1.1 67-1.4.1.3.1 (h / high-02~e.15 :ARG1 (l3 / level~e.7 :quant-of (p / protein~e.37 :name (n / name :op1 "IGF-1"~e.10,12,33) :source (f / fluid~e.5,36 :mod~e.5,36 (l5 / lavage~e.5,36 :mod~e.5,36 (a3 / alveolus~e.5,9,36 :mod~e.5,9,36 (b2 / bronchus~e.5,9,36))))) :ARG1-of (e2 / equal-01 :ARG2 (m4 / mass-quantity :unit (r / ratio-of~e.63 :op1 (p5 / picogram :mod p) :op2 (m6 / microgram :mod f)) :quant (v / value-interval :op1 (s4 / subtract-01 :ARG1 0.33~e.27 :ARG2 1.81~e.25) :op2 (a4 / add-02 :ARG1 0.33~e.27 :ARG2 1.81~e.25))) :condition (a2 / and :op1 (s6 / statistical-test-91~e.42 :ARG2 (l6 / less-than~e.44 :op1 0.01~e.45)) :op2 (d / deviate-01 :mod (m8 / mean~e.47) :ARG1-of (s3 / standard-02))))) :compared-to~e.21,28 (l4 / level~e.7 :quant-of (p4 / protein :name (n5 / name :op1 "IGF-1"~e.55,57) :source (c / control~e.23 :mod (n3 / naive~e.22))) :ARG1-of (e3 / equal-01 :ARG2 (m7 / mass-quantity :unit r :quant (v2 / value-interval :op1 (s5 / subtract-01 :ARG1 0.36~e.31 :ARG2 0.95~e.29) :op2 (a5 / add-02 :ARG1 0.36~e.31 :ARG2 0.95~e.29))) :condition a2)) :location~e.16 (a / animal~e.20 :ARG0-of (b / bear-01~e.19 :ARG1 (t / tumor~e.17))) :ARG0-of (s2 / suggest-01~e.52 :ARG1 (p3 / produce-01~e.59 :ARG1 p :quant (m2 / more~e.54) :location~e.60 (l2 / lung~e.62 :part-of~e.63 (m3 / mouse~e.67 :ARG0-of b~e.64,65,66)))) :degree~e.15 (m5 / more~e.15,54) :ARG1-of (s / significant-02~e.14) :concession (n4 / normalize-01~e.2 :ARG1 p :prep-to~e.3 (l / level~e.7 :quant-of (p2 / protein~e.6 :mod f) :mod (t2 / total~e.4)))) # ::id a_pmid_2169_9731.121 ::amr-annotator SDL-AMR-09 ::preferred # ::tok Measurement of IGF @-@ 1 levels in MØCM from primary naïve and tumor @-@ educated BAL macrophages showed that tumor @-@ educated macrophages produced significantly more IGF @-@ 1 than naïve macrophages ( Figure 6B , grey bars ) . # ::alignments 0-1.1 2-1.1.1.1.1.1 4-1.1.1.1.1.1 5-1.1.1 5-1.2.2 8-1.1.2.1.r 9-1.1.2.1.4 10-1.1.2.1.1.1 11-1.1.2.1 12-1.1.2.1.2.1.1 14-1.1.2.1.2.1 15-1.1.2.1.3.1 15-1.1.2.1.3.1.1 15-1.1.2.1.3.1.1.r 16-1.1.2.1.2 16-1.1.2.2.1 17-1 19-1.1.2.1.2.1.1 21-1.1.2.1.2.1 22-1.1.2.1.1 22-1.1.2.1.2 22-1.1.2.2.1 23-1.2 24-1.2.2.2.1 25-1.2.2.2 26-1.2.2.1 27-1.2.2.1 28-1.2.2.1 29-1.2.3.r 30-1.2.3 31-1.1.2.1.1 31-1.1.2.2.1 33-1.3.1 35-1.3.1.1 38-1.3.2.1 39-1.3.2 (s / show-01~e.17 :ARG0 (m / measure-01~e.0 :ARG1 (l / level~e.5 :quant-of (p / protein :name (n / name :op1 "IGF-1"~e.2,4))) :location (m2 / medium :source~e.8 (a / and~e.11 :op1 (m3 / macrophage~e.22,31 :mod (n4 / naive~e.10)) :op2 (m5 / macrophage~e.16,22 :ARG1-of (e / educate-01~e.14,21 :ARG0 (t / tumor~e.12,19))) :mod (f2 / fluid :mod (a2 / alveolus~e.15 :mod~e.15 (b2 / bronchus~e.15))) :mod (p2 / primary~e.9)) :ARG1-of (c / condition-01 :ARG0 (m7 / macrophage~e.16,22,31)))) :ARG1 (p3 / produce-01~e.23 :ARG0 m5 :ARG1 (l2 / level~e.5 :quant-of p~e.26,27,28 :quant (m6 / more~e.25 :ARG1-of (s2 / significant-02~e.24))) :compared-to~e.29 m3~e.30) :ARG1-of (d / describe-01 :ARG0 (f / figure~e.33 :mod "6B"~e.35) :ARG2 (b / bar~e.39 :ARG1-of (g / gray-02~e.38)))) # ::id a_pmid_2169_9731.122 ::amr-annotator SDL-AMR-09 ::preferred # ::tok IL @-@ 4 potently stimulates alternative macrophage activation , and is more abundant in tumor @-@ bearing lungs than naïve [ @ 38 @ ] . # ::alignments 0-1.1.1.1.1 2-1.1.1.1.1 3-1.1.3 3-1.1.3.r 4-1.1 5-1.1.2.2 6-1.1.2.1 7-1.1.2 9-1 10-1.2.1.r 11-1.2.2 12-1.2 13-1.2.3.r 14-1.2.3.1.1 16-1.2.3.1 17-1.2.3 18-1.2.4.r 19-1.2.4 22-1.3.1.1.1 (a / and~e.9 :op1 (s / stimulate-01~e.4 :ARG0 (p2 / protein :name (n / name :op1 "IL-4"~e.0,2)) :ARG1 (a2 / activate-01~e.7 :ARG1 (m / macrophage~e.6) :mod (a3 / alternative~e.5)) :manner~e.3 (p / potent~e.3)) :op2 (a4 / abundant~e.12 :domain~e.10 p2 :degree (m2 / more~e.11) :location~e.13 (l / lung~e.17 :ARG0-of (b / bear-01~e.16 :ARG1 (t / tumor~e.14))) :compared-to~e.18 (n2 / naive~e.19)) :ARG1-of (d / describe-01 :ARG0 (p3 / publication :ARG1-of (c / cite-01 :ARG2 38~e.22)))) # ::id a_pmid_2169_9731.123 ::amr-annotator SDL-AMR-09 ::preferred # ::tok Alternative macrophage polarization is associated with tumorigenesis [ @ 6 @ ] and increased macrophage IGF @-@ 1 production [ @ 39 @ ] . # ::alignments 0-1.1.2 1-1.1.1 2-1.1 4-1 5-1.2.r 6-1.2.1 6-1.2.1.1 6-1.2.1.1.r 9-1.2.1.2.1.1.1 12-1.2 13-1.2.2.3 14-1.2.2.1 15-1.2.2.2.1.1 17-1.2.2.2.1.1 18-1.2.2 21-1.2.2.4.1.1.1 (a / associate-01~e.4 :ARG1 (p / polarize-01~e.2 :ARG1 (m / macrophage~e.1) :mod (a2 / alternative~e.0)) :ARG2~e.5 (a3 / and~e.12 :op1 (c3 / create-01~e.6 :ARG1~e.6 (t / tumor~e.6) :ARG1-of (d / describe-01 :ARG0 (p2 / publication :ARG1-of (c / cite-01 :ARG2 6~e.9)))) :op2 (p3 / produce-01~e.18 :ARG0 m~e.14 :ARG1 (p4 / protein :name (n / name :op1 "IGF-1"~e.15,17)) :ARG1-of (i / increase-01~e.13) :ARG1-of (d2 / describe-01 :ARG0 (p5 / publication :ARG1-of (c2 / cite-01 :ARG2 39~e.21)))))) # ::id a_pmid_2169_9731.124 ::amr-annotator SDL-AMR-09 ::preferred # ::tok Therefore , IL @-@ 4 was added to wells containing primary naïve and tumor @-@ educated BAL macrophages to determine if alternative activation could increase IGF @-@ 1 production in either macrophage group . # ::alignments 0-1 2-1.1.1.1.1 4-1.1.1.1.1 6-1.1 7-1.1.2.r 8-1.1.2 9-1.1.2.1 10-1.1.2.1.1.3 11-1.1.2.1.1.1.1 12-1.1.2.1.1 13-1.1.2.1.1.2.1.1 15-1.1.2.1.1.2.1 17-1.1.2.1.1.1 17-1.1.2.1.1.2 17-1.1.2.1.1.4.1.1 19-1.1.3 21-1.1.3.1.1.1.1 22-1.1.3.1.1.1 23-1.1.3.1 24-1.1.3.1.1 28-1.1.3.1.1.2 31-1.1.3.1.1.3 (c / cause-01~e.0 :ARG1 (a / add-02~e.6 :ARG1 (p2 / protein :name (n / name :op1 "IL-4"~e.2,4)) :ARG2~e.7 (w / well~e.8 :ARG0-of (c2 / contain-01~e.9 :ARG1 (a2 / and~e.12 :op1 (m2 / macrophage~e.17 :mod (n3 / naive~e.11)) :op2 (m3 / macrophage~e.17 :ARG1-of (e / educate-01~e.15 :ARG0 (t / tumor~e.13))) :mod (p3 / primary~e.10) :mod (m / medium :ARG1-of (c3 / condition-01 :ARG0 (m4 / macrophage~e.17)))))) :purpose (d / determine-01~e.19 :ARG1 (p4 / possible-01~e.23 :ARG1 (i / increase-01~e.24 :ARG0 (a3 / activate-01~e.22 :mod (a4 / alternative~e.21)) :ARG1 (p5 / produce-01~e.28 :ARG1 p2) :location a2~e.31))))) # ::id a_pmid_2169_9731.125 ::amr-annotator SDL-AMR-09 ::preferred # ::tok Both naïve and tumor @-@ educated macrophages produced significantly more IGF @-@ 1 after IL @-@ 4 treatment ; tumor @-@ educated macrophages more than doubled IGF @-@ 1 output compared to naïve samples ( Figure 6B , green bars ) . # ::alignments 1-1.1.1.1.1 2-1.1.1 3-1.1.1.2.1.1 5-1.1.1.2.1 6-1.1.1.1 6-1.1.1.2 7-1.1 8-1.1.2.2.1 9-1.1.2.2 10-1.1.2.1.1 12-1.1.2.1.1 14-1.1.3.1.1.1 16-1.1.3.1.1.1 17-1.1.3 19-1.2.1.1.1 21-1.2.1.1 22-1.2.1 23-1.2.3 24-1.2.3 24-1.2.4.r 25-1.2 26-1.2.2.1.1.1 28-1.2.2.1.1.1 29-1.2.2 30-1.2.4.r 32-1.2.4.1 33-1.2.4 33-1.2.4.2 33-1.2.4.2.r 35-1.3.1 37-1.3.1.1 40-1.3.2.1 41-1.3.2 (m / multi-sentence :snt1 (p / produce-01~e.7 :ARG0 (a / and~e.2 :op1 (m2 / macrophage~e.6 :mod (n / naive~e.1)) :op2 (m3 / macrophage~e.6 :ARG1-of (e / educate-01~e.5 :ARG0 (t / tumor~e.3)))) :ARG1 (p2 / protein :name (n2 / name :op1 "IGF-1"~e.10,12) :quant (m4 / more~e.9 :degree (s / significant~e.8))) :condition (t2 / treat-04~e.17 :ARG2 (p3 / protein :name (n3 / name :op1 "IL-4"~e.14,16)))) :snt2 (d / double-01~e.25 :ARG0 (m6 / macrophage~e.22 :ARG1-of (e2 / educate-01~e.21 :ARG0 (t3 / tumor~e.19))) :ARG1 (o / output~e.29 :mod (p4 / protein :name (n4 / name :op1 "IGF-1"~e.26,28))) :degree (m5 / more-than~e.23,24) :compared-to~e.24,30 (t4 / thing~e.33 :mod (n5 / naive~e.32) :ARG1-of~e.33 (s2 / sample-01~e.33))) :ARG1-of (d2 / describe-01 :ARG0 (f / figure~e.35 :mod "6B"~e.37) :ARG2 (b / bar~e.41 :ARG1-of (g / green-02~e.40)))) # ::id a_pmid_2169_9731.126 ::amr-annotator SDL-AMR-09 ::preferred # ::tok MH @-@ S macrophages produced 20 @-@ times more IGF @-@ 1 than either non @-@ neoplastic or neoplastic lung cell lines , and all three cell lines produced only trace amounts (< 2 pg/mL ) of EGF ( Figure 6E ) . # ::alignments 0-1.1.1.1.1.1 2-1.1.1.1.1.1 3-1.1.1 4-1.1 5-1.1.2.2.1.1 7-1.1.2.2.1 8-1.1.2.2 9-1.1.2.1.1 11-1.1.2.1.1 12-1.1.3.r 12-1.2.2.3.1 14-1.1.3.1.1.1 14-1.1.3.1.1.1.r 17-1.1.3 19-1.1.3.3 20-1.1.3.1 21-1.1.3.1 21-1.1.3.2 26-1.1.3.1 27-1.1.3.1 28-1.2 29-1.2.2.4 30-1.2.2.1 31-1.2.2 33-1.2.2.3.1.1.1 36-1.2.2.2.r 37-1.2.2.2.1.1 39-1.3.1 41-1.3.1.1 (a / and :op1 (p / produce-01~e.4 :ARG0 (m / macrophage~e.3 :source (c / cell-line :name (n / name :op1 "MH-S"~e.0,2))) :ARG1 (p2 / protein :name (n2 / name :op1 "IGF-1"~e.9,11) :quant (m2 / more~e.8 :degree (p3 / product-of~e.7 :op1 20~e.5))) :compared-to~e.12 (o / or~e.17 :op1 (c5 / cell-line~e.20,21,26,27 :mod (t / tumor :polarity~e.14 -~e.14)) :op2 (c6 / cell-line~e.21 :mod (t2 / tumor)) :source (l2 / lung~e.19))) :op2 (p4 / produce-01~e.28 :ARG0 (a2 / and :op1 m :op2 c5 :op3 c6) :ARG1 (a3 / amount~e.31 :mod (t3 / trace~e.30) :mod~e.36 (p5 / protein :name (n3 / name :op1 "EGF"~e.37)) :ARG1-of (e / equal-01 :ARG2 (l5 / less-than~e.12 :op1 (c4 / concentration-quantity :quant 2~e.33 :unit (p6 / picogram-per-milliliter)))) :mod (o2 / only~e.29))) :ARG1-of (d / describe-01 :ARG0 (f / figure~e.39 :mod "6E"~e.41))) # ::id a_pmid_2169_9731.127 ::amr-annotator SDL-AMR-09 ::preferred # ::tok In order to determine whether the growth effects of MØCM from samples generated in Figure 6B correlated with their IGF @-@ 1 content , MØCM was added to neoplastic LM2 cells . # ::alignments 0-1.3.r 1-1.3.r 2-1.3.r 3-1.3 4-1.3.1.1 4-1.3.1.1.r 6-1.3.1.2.2 7-1.3.1.2 10-1.3.1.2.3.r 11-1.3.1.2.3 11-1.3.1.2.3.1 11-1.3.1.2.3.1.r 12-1.3.1.2.4 13-1.3.1.2.4.1.r 14-1.3.1.2.4.1 16-1.3.1.2.4.1.1 18-1.3.1 19-1.3.1.3.r 20-1.3.1.3.1 20-1.3.1.3.1.r 21-1.3.1.3.2.1.1 23-1.3.1.3.2.1.1 24-1.3.1.3 28-1 31-1.2.1.1 32-1.2 (a / add-02~e.28 :ARG1 (m / medium :ARG1-of (c4 / condition-01 :ARG0 (m2 / macrophage))) :ARG2 (c / cell-line~e.32 :name (n2 / name :op1 "LM2"~e.31) :mod (t / tumor)) :purpose~e.0,1,2 (d / determine-01~e.3 :ARG1 (c2 / correlate-01~e.18 :mode~e.4 interrogative~e.4 :ARG1 (a2 / affect-01~e.7 :ARG0 m :ARG1 (g / grow-01~e.6) :location~e.10 (t2 / thing~e.11 :ARG1-of~e.11 (s / sample-01~e.11)) :ARG1-of (g2 / generate-01~e.12 :location~e.13 (f / figure~e.14 :mod "6B"~e.16))) :ARG2~e.19 (c3 / contain-01~e.24 :ARG0~e.20 t2~e.20 :ARG1 (p / protein :name (n3 / name :op1 "IGF-1"~e.21,23)))))) # ::id a_pmid_2169_9731.128 ::amr-annotator SDL-AMR-09 ::preferred # ::tok IL @-@ 4 stimulated naïve and tumor @-@ educated MØCM significantly augmented LM2 proliferation ( Figure 6C , green bars ) , with IL @-@ 4 treated tumor @-@ educated MØCM being the most potent . # ::alignments 0-1.1.1.3.1.1.1 2-1.1.1.3.1.1.1 3-1.1.1.3 4-1.1.1.1.1 5-1.1.1 6-1.1.1.2.1.1 8-1.1.1.2.1 10-1.1.3 11-1.1 12-1.1.2.1.1.1 13-1.1.2 15-1.1.4.1 17-1.1.4.1.1 20-1.1.4.2.1 21-1.1.4.2 25-1.1.1.3.1.1.1 27-1.1.1.3.1.1.1 29-1.1.1.2.1.1 31-1.1.1.2.1 33-1.2.2.r 35-1.2.1 36-1.2 (a / and :op1 (a2 / augment-01~e.11 :ARG0 (a3 / and~e.5 :op1 (m / medium :mod (n3 / naive~e.4) :ARG1-of (c2 / condition-01 :ARG0 (m4 / macrophage))) :op2 (m2 / medium :ARG1-of (e / educate-01~e.8,31 :ARG0 (t / tumor~e.6,29)) :ARG1-of c2) :ARG1-of (s / stimulate-01~e.3 :ARG0 (p / protein :name (n4 / name :op1 "IL-4"~e.0,2,25,27)))) :ARG1 (p2 / proliferate-01~e.13 :ARG0 (c / cell-line :name (n5 / name :op1 "LM2"~e.12))) :ARG1-of (s2 / significant-02~e.10) :ARG1-of (d / describe-01 :ARG0 (f / figure~e.15 :mod "6C"~e.17) :ARG2 (b / bar~e.21 :ARG1-of (g / green-02~e.20)))) :op2 (p3 / potent~e.36 :degree (m3 / most~e.35) :domain~e.33 m2)) # ::id a_pmid_2169_9731.129 ::amr-annotator SDL-AMR-09 ::preferred # ::tok MØCM from untreated tumor @-@ educated macrophages did not stimulate LM2 growth significantly more than untreated naïve MØCM ( Figure 6C , grey bars ) , corresponding to previous co @-@ culture results ( Figure 2B ) . # ::alignments 1-1.2.1.r 2-1.2.1.1 2-1.2.1.1.1 3-1.2.1.2.1 5-1.2.1.2 6-1.2.1 6-1.7.3.1 8-1.1 8-1.1.r 8-1.2.1.1.1.r 9-1 10-1.3.1.1.1 11-1.3 12-1.4.1 13-1.4 14-1.7.r 15-1.7.1 16-1.7.2 19-1.5.1 21-1.5.1.1 24-1.5.2.1 25-1.5.2 28-1.6 29-1.6.1.r 30-1.6.1.1.2 31-1.6.1.1.1 33-1.6.1.1.1 34-1.6.1 34-1.6.1.1 34-1.6.1.1.r 36-1.6.1.1.3.1 38-1.6.1.1.3.1.1 (s / stimulate-01~e.9 :polarity~e.8 -~e.8 :ARG0 (m / medium :source~e.1 (m2 / macrophage~e.6 :ARG1-of (t / treat-04~e.2 :polarity~e.8 -~e.2) :ARG1-of (e / educate-01~e.5 :ARG0 (t2 / tumor~e.3))) :ARG1-of c4) :ARG1 (g / grow-01~e.11 :ARG1 (c / cell-line :name (n2 / name :op1 "LM2"~e.10))) :degree (m3 / more~e.13 :degree (s2 / significant~e.12)) :ARG1-of (d / describe-01 :ARG0 (f / figure~e.19 :mod "6C"~e.21) :ARG2 (b / bar~e.25 :ARG1-of (g2 / gray-02~e.24))) :ARG1-of (c2 / correspond-02~e.28 :ARG2~e.29 (t3 / thing~e.34 :ARG2-of~e.34 (r / result-01~e.34 :ARG1 (c3 / co-culture~e.31,33) :time (p / previous~e.30) :ARG1-of (d2 / describe-01 :ARG0 (f2 / figure~e.36 :mod "2B"~e.38))))) :compared-to~e.14 (m4 / medium :ARG1-of t~e.15 :mod (n4 / naive~e.16) :ARG1-of (c4 / condition-01 :ARG0 (m5 / macrophage~e.6)))) # ::id a_pmid_2169_9731.130 ::amr-annotator SDL-AMR-09 ::preferred # ::tok As the growth @-@ stimulating ability of MØCM appeared to correlate to media IGF @-@ 1 levels , the levels of IGF @-@ 1 present were plotted against the fold @-@ change in LM2 cell number after MØCM addition ( Figure 6D ) . # ::alignments 2-1.3.1.1.1.2.2 4-1.3.1.1.1.2 5-1.3.1.1.1 8-1.3.1 10-1.3.1.1 11-1.3 12-1.2.2.1 13-1.1.1.1.1 15-1.1.1.1.1 16-1.1 19-1.1 21-1.1.1.1.1 23-1.1.1.1.1 24-1.1.2 26-1 29-1.2.3 31-1.2 32-1.2.1.r 33-1.2.1.1.1.1 34-1.2.1.1 35-1.2.1 38-1.2.2 40-1.4.1 42-1.4.1.1 (p / plot-01~e.26 :ARG1 (l / level~e.16,19 :quant-of (p2 / protein :name (n / name :op1 "IGF-1"~e.13,15,21,23) :location m) :ARG1-of (p4 / present-02~e.24)) :ARG2 (c / change-01~e.31 :ARG1~e.32 (n2 / number~e.35 :quant-of (c2 / cell-line~e.34 :name (n3 / name :op1 "LM2"~e.33))) :condition (a / add-02~e.38 :ARG1 (m / medium~e.12 :ARG1-of (c6 / condition-01 :ARG0 (m3 / macrophage)))) :mod (p3 / product-of~e.29)) :ARG1-of (c3 / cause-01~e.11 :ARG0 (a2 / appear-02~e.8 :ARG1 (c4 / correlate-01~e.10 :ARG1 (c5 / capable-01~e.5 :ARG1 m :ARG2 (s / stimulate-01~e.4 :ARG0 m :ARG1 (g / grow-01~e.2))) :ARG2 l))) :ARG1-of (d / describe-01 :ARG0 (f2 / figure~e.40 :mod "6D"~e.42))) # ::id a_pmid_2169_9731.131 ::amr-annotator SDL-AMR-09 ::preferred # ::tok The correlation between IGF @-@ 1 levels and neoplastic growth stimulation was highly significant ( p < 0.001 ) , indicating that MØCM IGF @-@ 1 levels were directly related to the ability of MØCM to stimulate neoplastic proliferation . # ::alignments 1-1.1 3-1.1.1.1.1.1 5-1.1.1.1.1.1 6-1.1.1 9-1.1.2.1 10-1.1.2 12-1.2 13-1 16-1.4.1 17-1.4.1.1 20-1.3 23-1.3.1.1.1 24-1.3.1.1.1 25-1.3.1.1.1 26-1.3.1.1 28-1.3.1.3 29-1.3.1 30-1.3.1.2.r 32-1.3.1.2 36-1.3.1.2.2 38-1.3.1.2.2.2 (s / significant-02~e.13 :ARG1 (c / correlate-01~e.1 :ARG1 (l / level~e.6 :quant-of (p / protein :name (n / name :op1 "IGF-1"~e.3,5))) :ARG2 (s2 / stimulate-01~e.10 :ARG1 (g / grow-01~e.9 :ARG1 (t / tumor)))) :ARG1-of (h / high-02~e.12) :ARG0-of (i / indicate-01~e.20 :ARG1 (r / relate-01~e.29 :ARG1 (l2 / level~e.26 :quant-of p~e.23,24,25 :location (m / medium :ARG1-of (c3 / condition-01 :ARG0 (m3 / macrophage)))) :ARG2~e.30 (c2 / capable-01~e.32 :ARG1 m :ARG2 (s3 / stimulate-01~e.36 :ARG0 m :ARG1 (p2 / proliferate-01~e.38 :ARG0 t))) :ARG1-of (d / direct-02~e.28))) :ARG1-of (m2 / mean-01 :ARG2 (l4 / less-than~e.16 :op1 0.001~e.17))) # ::id a_pmid_2169_9731.132 ::amr-annotator SDL-AMR-09 ::preferred # ::tok IGF @-@ 1 stimulates lung epithelial cell proliferation and is additive with MØCM # ::alignments 0-1.1.1.1.1 2-1.1.1.1.1 3-1 4-1.2.1.1 5-1.2.1.2 6-1.2.1 7-1.2 8-1.1 (s / stimulate-01~e.3 :ARG0 (a / and~e.8 :op1 (p / protein :name (n / name :op1 "IGF-1"~e.0,2)) :op2 (m / medium :ARG1-of (c2 / condition-01 :ARG0 (m2 / macrophage)))) :ARG1 (p2 / proliferate-01~e.7 :ARG0 (c / cell~e.6 :source (l / lung~e.4) :mod (e / epithelium~e.5)))) # ::id a_pmid_2169_9731.133 ::amr-annotator SDL-AMR-09 ::preferred # ::tok While IGF @-@ 1 levels correlated strongly with the ability of MØCM to stimulate neoplastic growth , IGF @-@ 1 induced proliferation of these non @-@ neoplastic and neoplastic mouse lung cell lines has not been demonstrated . # ::alignments 0-1 1-1.2.1.1 2-1.2.1.1 3-1.2.1.1 4-1.2.1 5-1.2 6-1.2.3 7-1.2.2.r 9-1.2.2 13-1.2.2.2 15-1.2.2.2.2 17-1.1.2.2.1.1.1 19-1.1.2.2.1.1.1 20-1.1.2.2 21-1.1.2 22-1.1.2.1.r 23-1.1.2.1.4 24-1.1.2.1.1.1.1 24-1.1.2.1.1.1.1.r 27-1.1.2.1 29-1.1.2.1.3.1 30-1.1.2.1.3 31-1.1.2.1.1 31-1.1.2.1.2 32-1.1.2.1.1 34-1.1.1 34-1.1.1.r 36-1.1 (c3 / contrast-01~e.0 :ARG1 (d / demonstrate-01~e.36 :polarity~e.34 -~e.34 :ARG1 (p / proliferate-01~e.21 :ARG0~e.22 (a / and~e.27 :op1 (c / cell-line~e.31,32 :mod (t / tumor :polarity~e.24 -~e.24)) :op2 (c2 / cell-line~e.31 :mod (t2 / tumor)) :source (l / lung~e.30 :part-of (m / mouse~e.29)) :mod (t3 / this~e.23)) :ARG2-of (i / induce-01~e.20 :ARG0 (p2 / protein :name (n / name :op1 "IGF-1"~e.17,19))))) :ARG2 (c4 / correlate-01~e.5 :ARG1 (l2 / level~e.4 :quant-of p2~e.1,2,3) :ARG2~e.7 (c5 / capable-01~e.9 :ARG1 (m2 / medium :ARG1-of (c6 / condition-01 :ARG0 (m3 / macrophage))) :ARG2 (s2 / stimulate-01~e.13 :ARG0 m2 :ARG1 (g / grow-01~e.15 :ARG1 t2))) :ARG1-of (s / strong-02~e.6))) # ::id a_pmid_2169_9731.134 ::amr-annotator SDL-AMR-09 ::preferred # ::tok Recombinant mouse IGF @-@ 1 or MH @-@ S macrophage @-@ conditioned media was sufficient to stimulate the proliferation of neoplastic LM2 , JF32 and E9 cells and non @-@ neoplastic E10 cells ( Figure 7A @-@ D ) . # ::alignments 0-1.1.1.1.1.3 1-1.1.1.1.1.2 2-1.1.1.1.1.1.1 4-1.1.1.1.1.1.1 5-1.1 6-1.1.2.1.1.1.1.1 8-1.1.2.1.1.1.1.1 9-1.1.2.1.1 11-1.1.1.1 11-1.1.2.1 12-1.1.1 12-1.1.2 14-1 16-1.2 18-1.2.2 21-1.2.2.1.1.1.1.1 23-1.2.2.1.1.2.1.1 24-1.2.2.1.1 25-1.2.2.1.1.3.1.1 26-1.1.2.1.1.1 26-1.2.2.1.1.1 26-1.2.2.1.1.2 26-1.2.2.1.1.3 26-1.2.2.1.2 27-1.2.2.1 27-1.2.2.1.1 27-1.2.2.1.1.r 28-1.2.2.1.2.2.1 28-1.2.2.1.2.2.1.r 31-1.2.2.1.2.1.1 32-1.2.2.1.1.1 34-1.3.1.1 34-1.3.1.2 34-1.3.1.3 34-1.3.1.4 36-1.3.1.1.1 (s / suffice-01~e.14 :ARG0 (o / or~e.5 :op1 (m3 / medium~e.12 :ARG1-of (c3 / condition-01~e.11 :ARG0 (p / protein :name (n2 / name :op1 "IGF-1"~e.2,4) :source (m4 / mouse~e.1) :ARG3-of (r / recombine-01~e.0)))) :op2 (m / medium~e.12 :ARG1-of (c / condition-01~e.11 :ARG0 (m2 / macrophage~e.9 :source (c2 / cell-line~e.26 :name (n / name :op1 "MH-S"~e.6,8)))))) :ARG1 (s2 / stimulate-01~e.16 :ARG0 o :ARG1 (p2 / proliferate-01~e.18 :ARG0 (a / and~e.27 :op1~e.27 (a2 / and~e.24,27 :op1 (c4 / cell-line~e.26,32 :name (n3 / name :op1 "LM2"~e.21)) :op2 (c5 / cell-line~e.26 :name (n4 / name :op1 "JF32"~e.23)) :op3 (c6 / cell-line~e.26 :name (n5 / name :op1 "E9"~e.25)) :mod (t / tumor)) :op2 (c7 / cell-line~e.26 :name (n6 / name :op1 "E10"~e.31) :mod (t2 / tumor :polarity~e.28 -~e.28))))) :ARG1-of (d / describe-01 :ARG0 (a3 / and :op1 (f / figure~e.34 :mod "7A"~e.36) :op2 (f2 / figure~e.34 :mod "7B") :op3 (f3 / figure~e.34 :mod "7C") :op4 (f4 / figure~e.34 :mod "7D")))) # ::id a_pmid_2169_9731.135 ::amr-annotator SDL-AMR-09 ::preferred # ::tok The degree of growth stimulated by 50 ng @/@ mL IGF @-@ 1 was similar to that of MØCM in each line ( Figure 7A @-@ D ) . # ::alignments 3-1.1 3-1.2 4-1.1.1 5-1.1.1.1.r 6-1.1.1.1.2.1 7-1.1.1.1.2.2 9-1.1.1.1.2.2 10-1.1.1.1.1.1 12-1.1.1.1.1.1 14-1 19-1.3.r 20-1.3.1 21-1.3 23-1.4.1.1 23-1.4.1.2 23-1.4.1.3 23-1.4.1.4 25-1.4.1.1.1 (r / resemble-01~e.14 :ARG1 (g / grow-01~e.3 :ARG1-of (s / stimulate-01~e.4 :ARG0~e.5 (p / protein :name (n / name :op1 "IGF-1"~e.10,12) :quant (c / concentration-quantity :quant 50~e.6 :unit (n2 / nanogram-per-milliliter~e.7,9))))) :ARG2 (g2 / grow-01~e.3 :ARG1 (m / medium :ARG1-of (c2 / condition-01 :ARG0 (m2 / macrophage)))) :location~e.19 (l / line~e.21 :mod (e / each~e.20)) :ARG1-of (d / describe-01 :ARG0 (a / and :op1 (f / figure~e.23 :mod "7A"~e.25) :op2 (f2 / figure~e.23 :mod "7B") :op3 (f3 / figure~e.23 :mod "7C") :op4 (f4 / figure~e.23 :mod "7D")))) # ::id a_pmid_2169_9731.136 ::amr-annotator SDL-AMR-09 ::preferred # ::tok These results confirm that IGF @-@ 1 alone can stimulate the growth of long @-@ established neoplastic and non @-@ neoplastic cell lines , as well as cells isolated more recently from primary mouse lung tumors ( JF32 ) , consistent with previous reports on human cancer cell lines [ @ 27 @ ] . # ::alignments 0-1.1.2 1-1.1 1-1.1.1 1-1.1.1.r 2-1 3-1.2.r 4-1.2.1.1.1.1 6-1.2.1.1.1.1 7-1.2.1.1.2 8-1.2 9-1.2.1 11-1.2.1.2 12-1.2.1.2.1.r 13-1.2.1.2.1.1.3.1 15-1.2.1.2.1.1.3 17-1.2.1.2.1.1 18-1.2.1.2.1.1.2.1.1 18-1.2.1.2.1.1.2.1.1.r 21-1.2.1.2.1.2 22-1.2.1.2.1.1.1 24-1.2.1.2.1.1 25-1.2.1.2.1.1 26-1.2.1.2.1 26-1.2.1.2.1.1 26-1.2.1.2.1.1.r 27-1.2.1.2.1.1.1 27-1.2.1.2.1.1.2 27-1.2.1.2.1.2 28-1.2.1.2.1.2.1 29-1.2.1.2.1.2.1.2.1 30-1.2.1.2.1.2.1.2 31-1.1.1.1.1.3.1.r 31-1.2.1.2.1.2.1.1.r 32-1.2.1.2.1.2.1.1.2 33-1.2.1.2.1.2.1.1.1.1 34-1.2.1.2.1.2.1.1.1 35-1.2.1.2.1.1.1.1 35-1.2.1.2.1.1.2.1 35-1.2.1.2.1.2.1.1 37-1.2.1.2.1.2.1.1.3.1.1.1 40-1.1.1.1 42-1.1.1.1.1.2 43-1.1.1.1.1 43-1.1.1.1.1.1 43-1.1.1.1.1.1.r 45-1.1.1.1.1.3.1.3 46-1.1.1.1.1.3.1.2.1 47-1.1.1.1.1.3 48-1.1.1.1.1.3 51-1.1.1.1.1.4.1.1.1 (c / confirm-01~e.2 :ARG0 (t / thing~e.1 :ARG1-of~e.1 (r / result-01~e.1 :ARG1-of (c6 / consistent-01~e.40 :ARG2 (t6 / thing~e.43 :ARG1-of~e.43 (r3 / report-01~e.43) :time (p4 / previous~e.42) :topic (c7 / cell-line~e.47,48 :source~e.31 (d / disease :wiki "Cancer" :name (n / name :op1 "cancer"~e.46) :mod (h / human~e.45))) :ARG1-of (d2 / describe-01 :ARG0 (p5 / publication :ARG1-of (c9 / cite-01 :ARG2 27~e.51)))))) :mod (t2 / this~e.0)) :ARG1~e.3 (p / possible-01~e.8 :ARG1 (s / stimulate-01~e.9 :ARG0 (p2 / protein :name (n2 / name :op1 "IGF-1"~e.4,6) :mod (a / alone~e.7)) :ARG1 (g / grow-01~e.11 :ARG1~e.12 (a2 / and~e.26 :op1~e.26 (a3 / and~e.17,24,25,26 :op1 (c2 / cell-line~e.22,27 :mod (t3 / tumor~e.35)) :op2 (c3 / cell-line~e.27 :mod (t4 / tumor~e.35 :polarity~e.18 -~e.18)) :ARG1-of (e / establish-01~e.15 :ARG1-of (l / long-03~e.13))) :op2 (c4 / cell~e.21,27 :ARG1-of (i / isolate-01~e.28 :ARG2~e.31 (t5 / tumor~e.35 :mod (l2 / lung~e.34 :part-of (m2 / mouse~e.33)) :mod (p3 / primary~e.32) :ARG1-of (m3 / mean-01 :ARG2 (c5 / cell-line :name (n3 / name :op1 "JF32"~e.37)))) :time (r2 / recent~e.30 :degree (m / more~e.29))))))))) # ::id a_pmid_2169_9731.137 ::amr-annotator SDL-AMR-09 ::preferred # ::tok In order to determine any relevant role of EGFR in mediating macrophage @-@ stimulated tumor cell proliferation in these cell lines , recombinant mouse EGF was added at 2 ng/mL . # ::alignments 0-1.2.r 1-1.2.r 2-1.2.r 3-1.2 5-1.2.1 6-1.2.1.1 7-1.2.1.1.1.r 8-1.2.1.1.1.1.1 9-1.2.1.2.r 10-1.2.1.2 11-1.2.1.2.2.2.1 13-1.2.1.2.2.2 14-1.2.1.2.2.1.1 15-1.2.1.2.2.1 15-1.2.1.2.2.3 16-1.2.1.2.2 18-1.2.1.2.2.3.1 19-1.2.1.2.2.3 20-1.2.1.2.2.3 22-1.2.1.2.1 23-1.2.1.2.1 24-1.2.1.2.1 26-1 27-1.1.r 28-1.1.2.1 (a / add-02~e.26 :ARG1~e.27 (p / protein :name (n2 / name :op1 "EGF") :quant (c / concentration-quantity :quant 2~e.28 :unit (n3 / nanogram-per-milliliter)) :source (m / mouse) :ARG3-of (r / recombine-01)) :purpose~e.0,1,2 (d / determine-01~e.3 :ARG1 (r3 / relevant-01~e.5 :ARG1 (r4 / role~e.6 :poss~e.7 (e / enzyme :name (n / name :op1 "EGFR"~e.8))) :ARG2~e.9 (m2 / mediate-01~e.10 :ARG0 p~e.22,23,24 :ARG1 (p2 / proliferate-01~e.16 :ARG0 (c2 / cell~e.15 :mod (t / tumor~e.14)) :ARG1-of (s / stimulate-01~e.13 :ARG0 (m3 / macrophage~e.11)) :location (c3 / cell-line~e.15,19,20 :mod (t2 / this~e.18))))))) # ::id a_pmid_2169_9731.138 ::amr-annotator SDL-AMR-09 ::preferred # ::tok This is roughly 500 @-@ times the reported EC @ 50 @ for growth stimulation and 20 @-@ times higher than levels found in the BALF from tumor @-@ bearing animals ( Figure 6A ) . # ::alignments 0-1.1.1 2-1.1.2 2-1.2.2 3-1.1.2.1.1 5-1.1.2.1 7-1.1.2.1.2.2 10-1.1.2.1.2.1.1 13-1.1.2.1.2.1.2.1 14-1.1.2.1.2.1.2 15-1 16-1.2.2.1.3.1 18-1.2.2.1.3 19-1.2.2.1 19-1.2.2.1.2 19-1.2.2.1.2.r 20-1.2.2.1.4.r 21-1.2.2.1.4 22-1.2.2.1.4.1 23-1.2.2.1.4.1.1.r 25-1.2.2.1.4.1.1 25-1.2.2.1.4.1.1.1 25-1.2.2.1.4.1.1.1.1 25-1.2.2.1.4.1.1.1.1.1 25-1.2.2.1.4.1.1.1.1.1.r 25-1.2.2.1.4.1.1.1.1.r 25-1.2.2.1.4.1.1.1.r 26-1.2.2.1.4.1.1.2.r 27-1.2.2.1.4.1.1.2.1.1 29-1.2.2.1.4.1.1.2.1 30-1.2.2.1.4.1.1.2 32-1.3.1 34-1.3.1.1 (a / and~e.15 :op1 (e / equal-01 :ARG1 (t / this~e.0) :ARG2 (r2 / roughly~e.2 :op1 (p / product-of~e.5 :op1 500~e.3 :op2 (c / concentration-quantity :ARG4-of (h2 / have-percentage-maximal-effective-concentration-01 :ARG2 50~e.10 :ARG3 (s / stimulate-01~e.14 :ARG1 (g / grow-01~e.13))) :ARG1-of (r / report-01~e.7))))) :op2 (e2 / equal-01 :ARG1 t :ARG2 (r3 / roughly~e.2 :op1 (h / high-02~e.19 :ARG1 t :degree~e.19 (m / more~e.19) :degree (p2 / product-of~e.18 :op1 20~e.16) :compared-to~e.20 (l / level~e.21 :ARG1-of (f / find-01~e.22 :location~e.23 (f3 / fluid~e.25 :mod~e.25 (l2 / lavage~e.25 :mod~e.25 (a3 / alveolus~e.25 :mod~e.25 (b2 / bronchus~e.25))) :source~e.26 (a2 / animal~e.30 :ARG0-of (b / bear-01~e.29 :ARG1 (t3 / tumor~e.27))))))))) :ARG1-of (d / describe-01 :ARG0 (f2 / figure~e.32 :mod "6A"~e.34))) # ::id a_pmid_2169_9731.139 ::amr-annotator SDL-AMR-09 ::preferred # ::tok EGF had no significant effect on tumor cell proliferation when added alone , and did not significantly affect the ability of either IGF @-@ 1 or MØCM to stimulate neoplastic growth ( Figure 7E , F ) . # ::alignments 0-1.1.2.1.1 2-1.1.1 2-1.1.1.r 3-1.1.4 4-1.1 5-1.1.3.r 6-1.1.3.1.1 7-1.1.3.1 8-1.1.3 10-1.1.5 11-1.1.5.2 13-1 13-1.3.1 15-1.2.1 15-1.2.1.r 16-1.2.4 17-1.2 19-1.2.3 22-1.2.3.2.1.1.1.1 24-1.2.3.2.1.1.1.1 25-1.2.3.2.1 28-1.2.3.2 29-1.2.3.2.2.1 30-1.2.3.2.2 32-1.3.1.1 32-1.3.1.2 34-1.3.1.1.1 (a4 / and~e.13 :op1 (a / affect-01~e.4 :polarity~e.2 -~e.2 :ARG0 (p / protein :name (n / name :op1 "EGF"~e.0)) :ARG1~e.5 (p2 / proliferate-01~e.8 :ARG0 (c / cell~e.7 :mod (t / tumor~e.6))) :ARG1-of (s / significant-02~e.3) :condition (a2 / add-02~e.10 :ARG1 p :manner (a3 / alone~e.11))) :op2 (a5 / affect-01~e.17 :polarity~e.15 -~e.15 :ARG0 p :ARG1 (c3 / capable-01~e.19 :ARG1 o :ARG2 (s3 / stimulate-01~e.28 :ARG0 (o / or~e.25 :op1 (p4 / protein :name (n3 / name :op1 "IGF-1"~e.22,24)) :op2 (m / medium :ARG1-of (c2 / condition-01 :ARG0 (m2 / macrophage)))) :ARG1 (g / grow-01~e.30 :ARG1 (n2 / neoplasm~e.29)))) :ARG1-of s~e.16) :ARG1-of (d / describe-01 :ARG0 (a6 / and~e.13 :op1 (f / figure~e.32 :mod "7E"~e.34) :op2 (f2 / figure~e.32 :mod "7F")))) # ::id a_pmid_2169_9731.140 ::amr-annotator SDL-AMR-09 ::preferred # ::tok This is not surprising in view of recent studies showing that EGFR inhibitors do not inhibit growth of lung cells with KRAS @ mutations [ @ 40 @ ] . # ::alignments 0-1.2 2-1.1 2-1.1.r 3-1 4-1.4.r 5-1.4 6-1.4.1.r 7-1.4.1.1.1 8-1.4.1.1 9-1.4.1 10-1.4.1.2.r 11-1.4.1.2.2.1.1.1.1 12-1.4.1.2 12-1.4.1.2.2 12-1.4.1.2.2.1 12-1.4.1.2.2.1.r 12-1.4.1.2.2.r 13-1.4.1.1 14-1.4.1.2.1 14-1.4.1.2.1.r 15-1.4.1.2 15-1.4.1.2.2 15-1.4.1.2.2.1 15-1.4.1.2.2.1.r 15-1.4.1.2.2.r 16-1.4.1.2.3 17-1.4.1.2.3.1.r 18-1.4.1.2.3.1.1 19-1.4.1.2.3.1 22-1.4.1.2.3.1.2.1.1 24-1.4.1.2.3.1.2 24-1.4.1.2.3.1.2.2 24-1.4.1.2.3.1.2.2.r 27-1.3.1.1.1 (s / surprise-01~e.3 :polarity~e.2 -~e.2 :ARG0 (t2 / this~e.0) :ARG1-of (d / describe-01 :ARG0 (p2 / publication :ARG1-of (c3 / cite-01 :ARG2 40~e.27))) :ARG2-of~e.4 (v / view-02~e.5 :ARG1~e.6 (s2 / show-01~e.9 :ARG0 (s3 / study-01~e.8,13 :time (r / recent~e.7)) :ARG1~e.10 (i2 / inhibit-01~e.12,15 :polarity~e.14 -~e.14 :ARG0~e.12,15 (m / molecular-physical-entity~e.12,15 :ARG0-of~e.12,15 (i3 / inhibit-01~e.12,15 :ARG1 (e / enzyme :name (n / name :op1 "EGFR"~e.11)))) :ARG1 (g / grow-01~e.16 :ARG1~e.17 (c2 / cell~e.19 :mod (l / lung~e.18) :mod (g2 / gene~e.24 :name (n2 / name :op1 "KRAS"~e.22) :ARG2-of~e.24 (m2 / mutate-01~e.24)))))))) # ::id a_pmid_2169_9731.141 ::amr-annotator SDL-AMR-09 ::preferred # ::tok As IGF @-@ 1 was sufficient to induce neoplastic proliferation , we determined whether the IGF @-@ 1 and MØCM growth effects were additive . # ::alignments 1-1.3.1.1 2-1.3.1.1 3-1.3.1.1 5-1.3.1 7-1.3.1.2 8-1.3.1.2.2.1 9-1.3.1.2.2 11-1.1 12-1 13-1.2.1 13-1.2.1.r 15-1.2.2.1.1.1.1 17-1.2.2.1.1.1.1 20-1.2.2.1.2 21-1.2.2.1 21-1.2.2.2 23-1.2 (d / determine-01~e.12 :ARG0 (w / we~e.11) :ARG1 (a / add-02~e.23 :mode~e.13 interrogative~e.13 :ARG1 (a5 / and :op1 (a2 / affect-01~e.21 :ARG0 (p / protein :name (n / name :op1 "IGF-1"~e.15,17)) :ARG2 (g / grow-01~e.20)) :op2 (a3 / affect-01~e.21 :ARG0 (m / medium :ARG1-of (c / condition-01 :ARG0 (m2 / macrophage))) :ARG2 g))) :ARG1-of (c2 / cause-01 :ARG0 (s / suffice-01~e.5 :ARG0 p~e.1,2,3 :ARG1 (i / induce-01~e.7 :ARG0 p :ARG2 (p2 / proliferate-01~e.9 :ARG0 (n3 / neoplasm~e.8)))))) # ::id a_pmid_2169_9731.142 ::amr-annotator SDL-AMR-09 ::preferred # ::tok A dose of 50 ng @/@ ml IGF @-@ 1 stimulated neoplastic growth to a similar extent as MØCM ( Figure 7A @-@ D ) ; 2 ng @/@ mL IGF is the reported EC @ 50 @ for IGF @-@ 1 stimulated proliferation in vitro @ as well as the concentration detected in the BALF of tumor @-@ bearing mice in vivo @ ( Figure 6A ) . # ::alignments 1-1.1.1 2-1.1.1.1.r 3-1.1.1.1.2.1 4-1.1.1.1.2.2 6-1.1.1.1.2.2 7-1.1.1.1.1.1 9-1.1.1.1.1.1 10-1.1 10-1.1.3.1.1 11-1.1.2.1 12-1.1.2 13-1.1.3.r 15-1.1.3.1 16-1.1.3 20-1.1.4.1.1 22-1.1.4.1.1.1 28-1.2.2.1 29-1.2.2.2 30-1.2.2.2 31-1.2.2.2 32-1.2.1.1 35-1.2.3.1.1.2 38-1.2.3.1.1.1 41-1.2.1.1 43-1.2.4.1.1.1.1 44-1.2.4.1 45-1.2.4 47-1.2.4.2 48-1.2.4.2 50-1 50-1.2.3.1 51-1.2.3.1 52-1.1.4.1 52-1.2.3.1 54-1.1.1.1.2 54-1.2.2 54-1.2.3.1.1 54-1.2.3.1.2 55-1.2.3.1.2.2 56-1.2.3.1.2.1.r 56-1.2.3.1.2.3 58-1.2.3.1.2.1 58-1.2.3.1.2.1.1 58-1.2.3.1.2.1.1.1 58-1.2.3.1.2.1.1.1.1 58-1.2.3.1.2.1.1.1.1.r 58-1.2.3.1.2.1.1.1.r 58-1.2.3.1.2.1.1.r 59-1.2.3.1.2.1.2.r 60-1.2.3.1.2.1.2.1.1 62-1.2.3.1.2.1.2.1 63-1.2.3.1.2.1.2 65-1.2.3.1.2.3 66-1.2.3.1.2.3 69-1.1.4.1.2 69-1.1.4.1.3 69-1.1.4.1.4 69-1.3.1 71-1.3.1.1 (a2 / and~e.50 :op1 (s / stimulate-01~e.10 :ARG0 (d / dose-01~e.1 :ARG2~e.2 (p / protein :name (n / name :op1 "IGF-1"~e.7,9) :quant (c / concentration-quantity~e.54 :quant 50~e.3 :unit (n5 / nanogram-per-milliliter~e.4,6)))) :ARG1 (g / grow-01~e.12 :ARG1 (n2 / neoplasm~e.11)) :ARG3~e.13 (e / extent~e.16 :ARG1-of (r2 / resemble-01~e.15 :ARG2 (s2 / stimulate-01~e.10 :ARG0 (m / medium :ARG1-of (c2 / condition-01 :ARG0 (m4 / macrophage))) :ARG1 g))) :ARG1-of (d2 / describe-01 :ARG0 (a3 / and~e.52 :op1 (f / figure~e.20 :mod "7A"~e.22) :op2 (f3 / figure~e.69 :mod "7B") :op3 (f4 / figure~e.69 :mod "7C") :op4 (f5 / figure~e.69 :mod "7D")))) :op2 (p2 / protein :name (n4 / name :op1 "IGF"~e.32,41) :quant (c3 / concentration-quantity~e.54 :quant 2~e.28 :unit n5~e.29,30,31) :ARG1-of (e2 / equal-01 :ARG2 (a / and~e.50,51,52 :op1 (h / have-percentage-maximal-effective-concentration-01~e.54 :ARG2 50~e.38 :ARG1-of (r3 / report-01~e.35)) :op2 (c4 / concentrate-02~e.54 :ARG1~e.56 (f6 / fluid~e.58 :mod~e.58 (l / lavage~e.58 :mod~e.58 (a4 / alveolus~e.58 :mod~e.58 (b2 / bronchus~e.58))) :source~e.59 (m2 / mouse~e.63 :ARG0-of (b / bear-01~e.62 :ARG1 (t / tumor~e.60)))) :ARG1-of (d3 / detect-01~e.55) :manner (i2 / in-vivo~e.56,65,66)))) :condition (p3 / proliferate-01~e.45 :ARG1-of (s3 / stimulate-01~e.44 :ARG0 (p5 / protein :name (n6 / name :op1 "IGF-1"~e.43))) :manner (i / in-vitro~e.47,48))) :ARG1-of (d4 / describe-01 :ARG0 (f2 / figure~e.69 :mod "6A"~e.71))) # ::id a_pmid_2169_9731.143 ::amr-annotator SDL-AMR-09 ::preferred # ::tok IGF @-@ 1 dose @-@ dependently stimulated the proliferation of both LM2 and JF32 cells , and augmented the growth @-@ stimulating effects of MØCM when added in combination . # ::alignments 0-1.1.1.1.1 2-1.1.1.1.1 3-1.1.3.1 5-1.1.3 6-1.1 8-1.1.2 11-1.1.2.1.1.1.1 12-1.1.2.1 13-1.1.2.1.2.1.1 14-1.1.2.1.1 14-1.1.2.1.2 16-1 17-1.2 19-1.2.2.2.1 21-1.2.2.2 22-1.2.2 26-1.2.3 27-1.2.3.2.r 28-1.2.3.2 (a / and~e.16 :op1 (s / stimulate-01~e.6 :ARG0 (p / protein :name (n / name :op1 "IGF-1"~e.0,2)) :ARG1 (p2 / proliferate-01~e.8 :ARG0 (a3 / and~e.12 :op1 (c / cell-line~e.14 :name (n2 / name :op1 "LM2"~e.11)) :op2 (c2 / cell-line~e.14 :name (n3 / name :op1 "JF32"~e.13)))) :ARG0-of (d2 / depend-01~e.5 :ARG1 (d3 / dose~e.3))) :op2 (a2 / augment-01~e.17 :ARG0 p :ARG1 (a5 / affect-01~e.22 :ARG0 (m / medium :ARG1-of (c3 / condition-01 :ARG0 (m2 / macrophage))) :ARG2 (s2 / stimulate-01~e.21 :ARG1 (g / grow-01~e.19))) :condition (a4 / add-02~e.26 :ARG1 p :ARG2~e.27 (c4 / combine-01~e.28)))) # ::id a_pmid_2169_9731.144 ::amr-annotator SDL-AMR-09 ::preferred # ::tok To determine if IGF @-@ 1R signaling mediates both IGF @-@ 1 and MØCM stimulation , lung cancer cells were pre @-@ treated with vehicle or 5 μM NVP @-@ AEW541 (- or +, respectively ) , and cell numbers determined as indicated . # ::alignments 1-1.3 3-1.3.1.2.1.1.1 3-1.3.1.3.1.1.1.1 5-1.3.1.2.1.1.1 6-1.3.1.2 7-1.3.1 9-1.3.1.3.1.1.1.1 11-1.3.1.3.1.1.1.1 12-1.3.1.3.1 14-1.3.1.3 16-1.1.1.1.2.1 17-1.1.1.1.2.2 18-1.1.1 24-1.1.2.1 25-1.1.2 26-1.1.2.2.2.1 27-1.1.2.2.2.2 28-1.1.2.2.1.1 30-1.1.2.2.1.1 32-1.1.2 37-1 38-1.2.1.1 39-1.2.1 40-1.2 41-1.2.2.r 42-1.2.2 (a / and~e.37 :op1 (p3 / pretreat-01 :ARG1 (c / cell~e.18 :source (d / disease :wiki "Lung_cancer" :name (n / name :op1 "lung"~e.16 :op2 "cancer"~e.17))) :ARG3 (o / or~e.25,32 :op1 (v / vehicle~e.24) :op2 (s3 / small-molecule :name (n2 / name :op1 "NVP-AEW541"~e.28,30) :quant (c3 / concentration-quantity :quant 5~e.26 :unit (m / micromolar~e.27))))) :op2 (d2 / determine-01~e.40 :ARG1 (n3 / number~e.39 :quant-of (c4 / cell~e.38)) :ARG1-of~e.41 (i / indicate-01~e.42)) :purpose (d3 / determine-01~e.1 :ARG1 (m2 / mediate-01~e.7 :mode interrogative :ARG0 (s / signal-07~e.6 :ARG0 (p / protein :name (n4 / name :op1 "IGF-1R"~e.3,5))) :ARG1 (s2 / stimulate-01~e.14 :ARG1 (a2 / and~e.12 :op1 (p2 / protein :name (n5 / name :op1 "IGF-1"~e.3,9,11)) :op2 (m3 / medium :ARG1-of (c2 / condition-01 :ARG0 (m4 / macrophage)))))))) # ::id a_pmid_2169_9731.145 ::amr-annotator SDL-AMR-09 ::preferred # ::tok IGF @-@ 1 and MØCM each significantly increased cell numbers after 48 and 72 hrs , while pharmacological inhibition of IGF @-@ 1R signaling blocked IGF @-@ 1 and MØCM growth effects in both neoplastic lines ( Figure 7G , H ) . # ::alignments 0-1.1.1.1.1.1 2-1.1.1.1.1.1 3-1.1.1 6-1.1.3 7-1.1 8-1.1.2.1 9-1.1.2 10-1.1.4 11-1.1.4.1.1.1 12-1.1.4.1 13-1.1.4.1.2.1 14-1.1.4.1.1.2 16-1 16-1.1.4.r 17-1.2.1.2 18-1.2.1 19-1.2.1.1.r 20-1.2.1.1.1.1.1 22-1.2.1.1.1.1.1 23-1.2.1.1 24-1.2 25-1.2.2.1.1.1 26-1.2.2.1.1.1 27-1.2.2.1.1.1 28-1.2.2.1.1 28-1.3.1 30-1.2.2.1 31-1.2.2 32-1.2.3.r 33-1.2.3.2 34-1.2.3.1 35-1.2.3 37-1.3.1.1 37-1.3.1.2 39-1.3.1.1.1 (c / contrast-01~e.16 :ARG1 (i / increase-01~e.7 :ARG0 (a / and~e.3 :op1 (p / protein :name (n / name :op1 "IGF-1"~e.0,2)) :op2 (m / medium :ARG1-of (c2 / condition-01 :ARG0 (m2 / macrophage)))) :ARG1 (n3 / number~e.9 :quant-of (c3 / cell~e.8)) :ARG1-of (s / significant-02~e.6) :time~e.16 (a2 / after~e.10 :quant (a3 / and~e.12 :op1 (t / temporal-quantity :quant 48~e.11 :unit (h / hour~e.14)) :op2 (t2 / temporal-quantity :quant 72~e.13 :unit h)))) :ARG2 (b / block-01~e.24 :ARG0 (i2 / inhibit-01~e.18 :ARG1~e.19 (s2 / signal-07~e.23 :ARG0 (p3 / protein :name (n4 / name :op1 "IGF-1R"~e.20,22))) :mod (p2 / pharmacology~e.17)) :ARG1 (a6 / affect-01~e.31 :ARG2 (g / grow-01~e.30 :ARG0 (a4 / and~e.28 :op1 p~e.25,26,27 :op2 m))) :location~e.32 (l / line~e.35 :mod (n5 / neoplasm~e.34) :mod (b2 / both~e.33))) :ARG1-of (d / describe-01 :ARG0 (a5 / and~e.28 :op1 (f / figure~e.37 :mod "7G"~e.39) :op2 (f2 / figure~e.37 :mod "7H")))) # ::id a_pmid_2169_9731.146 ::amr-annotator SDL-AMR-09 ::preferred # ::tok Parallel comparison of MTS values indicated a highly significant correlation between live cell numbers and relative MTS scores ( r @ 2 @ = 0.7912 and 0.8201 for LM2 and JF32 , respectively , p < 0.0001 , data not shown ) . # ::alignments 0-1.1.3 1-1.1 2-1.1.1.r 3-1.1.1.1.1.1 4-1.1.1 5-1 7-1.2.3.1 8-1.2.3 9-1.2 11-1.2.1.1.1 12-1.2.1.1 13-1.2.1 14-1.2.2.3.1 15-1.2.2.2 16-1.2.2.1 17-1.2.2 24-1.2.2.3.1.1.1 25-1.2.2.3.1 26-1.2.2.3.1.2.1 28-1.2.2.3.1.1.2.1.1 29-1.2.2.3.1 30-1.2.2.3.1.2.2.1.1 34-1.2.2.3.1.1 34-1.2.2.3.1.2 34-1.2.2.3.1.3 35-1.2.2.3.1.3.1 36-1.2.2.3.1.3.1.1 38-1.2.2.3.2.1 39-1.2.2.3.2.1.1.1 39-1.2.2.3.2.1.1.1.r 40-1.2.2.3.2.1.1 (i / indicate-01~e.5 :ARG0 (c / compare-01~e.1 :ARG1~e.2 (v / value~e.4 :mod (t / thing :name (n / name :op1 "MTS"~e.3))) :ARG2 v :manner (p / parallel~e.0)) :ARG1 (c2 / correlate-01~e.9 :ARG1 (n3 / number~e.13 :quant-of (c3 / cell~e.12 :ARG0-of (l / live-01~e.11))) :ARG2 (s / score-01~e.17 :ARG1 t~e.16 :ARG1-of (r / relative-05~e.15) :ARG1-of (m / mean-01 :ARG2 (a / and~e.14,25,29 :op1 (s4 / statistical-test-91~e.34 :ARG3 0.7912~e.24 :ARG1 (c4 / cell-line :name (n5 / name :op1 "LM2"~e.28))) :op2 (s5 / statistical-test-91~e.34 :ARG3 0.8201~e.26 :ARG1 (c5 / cell-line :name (n6 / name :op1 "JF32"~e.30))) :op3 (s6 / statistical-test-91~e.34 :ARG2 (l2 / less-than~e.35 :quant 0.0001~e.36))) :ARG1-of (d / describe-01 :ARG0 (d2 / data~e.38 :ARG1-of (s3 / show-01~e.40 :polarity~e.39 -~e.39))))) :ARG1-of (s2 / significant-02~e.8 :ARG1-of (h / high-02~e.7)))) # ::id a_pmid_2169_9731.147 ::amr-annotator SDL-AMR-09 ::preferred # ::tok Furthermore , both IGF @-@ 1 and MØCM increased the fraction of BrdU @ + @ LM2 cells 12 @-@ 24 hrs after treatment , corresponding with significantly increased cell numbers ( data not shown ) . # ::alignments 0-1 3-1.1.1.1.1.1 5-1.1.1.1.1.1 6-1.1.1 8-1.1 10-1.1.2 11-1.1.2.1.r 12-1.1.2.1.2.1.1 14-1 16-1.1.2.1.1.1 17-1.1.2.1 18-1.1.3.2.1.1 20-1.1.3.2.1.2 21-1.1.3.2.2 22-1.1.3 23-1.1.3.1 25-1.1.4 26-1.1.4.1.r 27-1.1.4.1.2.1 28-1.1.4.1.2 29-1.1.4.1.1 30-1.1.4.1 32-1.2.1 33-1.2.1.1.1 33-1.2.1.1.1.r 34-1.2.1.1 (a / and~e.0,14 :op2 (i / increase-01~e.8 :ARG0 (a2 / and~e.6 :op1 (p / protein :name (n / name :op1 "IGF-1"~e.3,5)) :op2 (m / medium :ARG1-of (c2 / condition-01 :ARG0 (m2 / macrophage)))) :ARG1 (f / fraction~e.10 :quant-of~e.11 (c / cell-line~e.17 :name (n4 / name :op1 "LM2"~e.16) :mod (s3 / small-molecule :name (n3 / name :op1 "BrdU"~e.12) :mod (w / wild-type)))) :time (a3 / after~e.22 :op1 (t / treat-04~e.23) :quant (t2 / temporal-quantity :quant (v / value-interval :op1 12~e.18 :op2 24~e.20) :unit (h / hour~e.21))) :ARG1-of (c3 / correspond-02~e.25 :ARG2~e.26 (n5 / number~e.30 :quant-of (c4 / cell~e.29) :ARG1-of (i2 / increase-01~e.28 :ARG2 (s / significant-02~e.27))))) :ARG1-of (d / describe-01 :ARG0 (d2 / data~e.32 :ARG1-of (s2 / show-01~e.34 :polarity~e.33 -~e.33)))) # ::id a_pmid_2169_9731.148 ::amr-annotator SDL-AMR-09 ::preferred # ::tok These observations suggest that IGF @-@ 1 , but not EGF , plays a major role in macrophage stimulated neoplastic growth in vitro , consistent with the elevated IGF @-@ 1 levels observed in lung @-@ tumor bearing animals in vivo @ . # ::alignments 0-1.1.1 1-1.1 2-1 3-1.2.r 4-1.2.1.1.1.1 6-1.2.1.1.1.1 8-1.2 9-1.2.2.1 9-1.2.2.1.r 10-1.2.2.2.1.1 12-1.2.1 12-1.2.2 14-1.2.1.4 16-1.2.1.2.r 16-1.2.1.3 17-1.2.1.2.1.1.1 18-1.2.1.2.1.1 19-1.2.1.2.1 20-1.2.1.2 22-1.2.1.3 23-1.2.1.3 26-1.3 27-1.3.1.r 29-1.3.1.2 30-1.3.1.3 31-1.3.1.3 32-1.3.1.3 33-1.3.1 34-1.3.1.1 35-1.3.1.1.1 35-1.3.1.1.2.r 36-1.3.1.1.2.1.1.1 38-1.3.1.1.2.1.1 39-1.3.1.1.2.1 40-1.3.1.1.2 42-1.3.1.1.1 43-1.3.1.1.1 (s / suggest-01~e.2 :ARG0 (o / observe-01~e.1 :mod (t / this~e.0)) :ARG1~e.3 (c / contrast-01~e.8 :ARG1 (p / play-08~e.12 :ARG0 (p2 / protein :name (n / name :op1 "IGF-1"~e.4,6)) :ARG1~e.16 (g / grow-01~e.20 :ARG1 (n2 / neoplasm~e.19 :ARG1-of (s2 / stimulate-01~e.18 :ARG0 (m2 / macrophage~e.17)))) :manner (i / in-vitro~e.16,22,23) :ARG1-of (m / major-02~e.14)) :ARG2 (p3 / play-08~e.12 :polarity~e.9 -~e.9 :ARG0 (p4 / protein :name (n3 / name :op1 "EGF"~e.10)))) :ARG1-of (c2 / consistent-01~e.26 :ARG2~e.27 (l / level~e.33 :ARG1-of (o2 / observe-01~e.34 :manner (i2 / in-vivo~e.35,42,43) :location~e.35 (a / animal~e.40 :ARG0-of (b / bear-01~e.39 :ARG1 (t2 / tumor~e.38 :mod (l2 / lung~e.36))))) :ARG1-of (e / elevate-01~e.29) :quant-of p2~e.30,31,32))) # ::id a_pmid_2169_9731.149 ::amr-annotator SDL-AMR-09 ::preferred # ::tok MØCM stimulation of neoplastic growth is diminished when IGF @-@ 1 content is decreased # ::alignments 1-1.1 2-1.1.2.r 3-1.1.2.1 4-1.1.2 6-1 7-1.2.r 8-1.2.1.1.1.1 10-1.2.1.1.1.1 11-1.2.1 13-1.2 (d / diminish-01~e.6 :ARG1 (s / stimulate-01~e.1 :ARG0 (m / medium :ARG1-of (c / condition-01 :ARG0 (m2 / macrophage))) :ARG1~e.2 (g / grow-01~e.4 :ARG1 (n2 / neoplasm~e.3))) :time~e.7 (d2 / decrease-01~e.13 :ARG1 (c2 / content~e.11 :quant-of (p / protein :name (n3 / name :op1 "IGF-1"~e.8,10))))) # ::id a_pmid_2169_9731.150 ::amr-annotator SDL-AMR-09 ::preferred # ::tok In order to determine if IGF @-@ 1 was a molecular mediator directly responsible for growth stimulated by MØCM , we decreased MØCM IGF @-@ 1 content through two independent avenues : immuno @-@ depletion and siRNA interference . # ::alignments 0-1.4.r 1-1.4.r 2-1.4.r 3-1.4 5-1.4.2.2.1.1 7-1.4.2.2.1.1 10-1.4.2.2.3 11-1.4.2.2 11-1.4.2.2.2 11-1.4.2.2.2.r 12-1.4.2.4 13-1.4.2 14-1.4.2.3.r 15-1.4.2.3 16-1.4.2.3.1 20-1.1 20-1.4.1 21-1 23-1.2.2 24-1.2.2 25-1.2.2 26-1.2 28-1.3.1 29-1.3.2 29-1.3.2.1 29-1.3.2.1.r 30-1.3 35-1.3.3.1 36-1.3.3.1.2.1.1.1 37-1.3.3.1.2 (d / decrease-01~e.21 :ARG0 (w / we~e.20) :ARG1 (c / contain-01~e.26 :ARG0 m4 :ARG1 p3~e.23,24,25) :instrument (a2 / avenue~e.30 :quant 2~e.28 :ARG0-of (d2 / depend-01~e.29 :polarity~e.29 -~e.29) :ARG1-of (m2 / mean-01 :ARG2 (a3 / and~e.35 :op1 (i / immunodeplete-00) :op2 (i2 / interfere-01~e.37 :ARG1 (n / nucleic-acid :name (n3 / name :op1 "siRNA"~e.36)))))) :purpose~e.0,1,2 (d3 / determine-01~e.3 :ARG0 (w2 / we~e.20) :ARG1 (r2 / responsible-01~e.13 :mode interrogative :ARG0 (p3 / protein~e.11 :name (n4 / name :op1 "IGF-1"~e.5,7) :ARG0-of~e.11 (m / mediate-01~e.11) :mod (m3 / molecule~e.10)) :ARG1~e.14 (g / grow-01~e.15 :ARG1-of (s / stimulate-01~e.16 :ARG0 (m4 / medium :ARG1-of (c2 / condition-01 :ARG0 (m5 / macrophage))))) :ARG1-of (d4 / direct-02~e.12)))) # ::id a_pmid_2169_9731.151 ::amr-annotator SDL-AMR-09 ::preferred # ::tok MØCM was concentrated to contain ~ 3.5 ng @/@ mL IGF @-@ 1 , and then incubated with control IgG ( Con IgG ) or α-IGF @-@ 1 IgG coated resin , as described [ @ 39 @ ] . # ::alignments 2-1.1 2-1.1.2.2.2.1 4-1.1.2 5-1.1.2.2.2 6-1.1.2.2.2.1.1 7-1.1.2.2.2.1.2 9-1.1.2.2.2.1.2 10-1.1.2.2.1.1 12-1.1.2.2.1.1 14-1 15-1.2.3 16-1.2 17-1.2.2.r 18-1.2.2.1 18-1.2.2.1.2 18-1.2.2.1.2.r 19-1.2.2.1.1.1 22-1.2.2.1.1.1 24-1.2.2 27-1.2.2.2.1.1.1.1.1 28-1.2.2.2.1.1.2 29-1.2.2.2.1 30-1.2.2.2 32-1.2.3.r 33-1.3.1.1 36-1.3.1.1.1.1.1 (a / and~e.14 :op1 (c / concentrate-02~e.2 :ARG1 (m / medium :ARG1-of (c2 / condition-01 :ARG0 (m2 / macrophage))) :purpose (c3 / contain-01~e.4 :ARG0 m :ARG1 (p4 / protein :name (n5 / name :op1 "IGF-1"~e.10,12) :quant (a3 / approximately~e.5 :op1 (c4 / concentration-quantity~e.2 :quant 3.5~e.6 :unit (n6 / nanogram-per-milliliter~e.7,9)))))) :op2 (i / incubate-01~e.16 :ARG1 m :ARG2~e.17 (o / or~e.24 :op1 (p / protein~e.18 :name (n2 / name :op1 "IgG"~e.19,22) :ARG0-of~e.18 (c5 / control-01~e.18)) :op2 (r2 / resin~e.30 :ARG1-of (c6 / coat-01~e.29 :ARG2 (a2 / and :op1 (p2 / protein :name (n4 / name :op1 "α-IGF-1"~e.27)) :op2 p~e.28)))) :time~e.32 (t / then~e.15)) :ARG1-of (r / resemble-01 :ARG2 (t2 / thing :ARG1-of (d / describe-01~e.33 :ARG0 (p3 / publication :ARG1-of (c7 / cite-01 :ARG2 39~e.36)))))) # ::id a_pmid_2169_9731.152 ::amr-annotator SDL-AMR-09 ::preferred # ::tok This procedure successfully decreased MØCM IGF @-@ 1 levels to 40 @-@ 50 % of control ( Figure 8A ) . # ::alignments 0-1.1.1 1-1.1 2-1.5 3-1 5-1.2.1.1.1 7-1.2.1.1.1 8-1.2 9-1.3.r 10-1.3.1.1.1 12-1.3.1.1.2 13-1.3.1 14-1.3.1.r 15-1.3 17-1.4.1 19-1.4.1.1 (d / decrease-01~e.3 :ARG0 (p2 / procedure~e.1 :mod (t / this~e.0)) :ARG1 (l / level~e.8 :quant-of (p3 / protein :name (n2 / name :op1 "IGF-1"~e.5,7) :mod (m / medium :ARG1-of (c / condition-01 :ARG0 (m2 / macrophage))))) :ARG4~e.9 (c3 / control~e.15 :quant~e.14 (p4 / percentage-entity~e.13 :value (v / value-interval :op1 40~e.10 :op2 50~e.12))) :ARG1-of (d2 / describe-01 :ARG0 (f / figure~e.17 :mod "8A"~e.19)) :ARG1-of (s / succeed-01~e.2)) # ::id a_pmid_2169_9731.153 ::amr-annotator SDL-AMR-09 ::preferred # ::tok When this IGF @-@ 1 depleted media was added to LM2 and JF32 cells , growth stimulation was significantly decreased compared to untreated MØCM or IgG controls , which were identical ( Figure 8B ) . # ::alignments 0-1.3.r 1-1.3.1.2 2-1.3.1.1.1.1.1 4-1.3.1.1.1.1.1 5-1.3.1.1 6-1.3.1 6-1.4.1.1 8-1.3 9-1.3.2.r 10-1.3.2.1.1.1 11-1.3.2 12-1.3.2.2.1.1 13-1.3.2.1 13-1.3.2.2 15-1.1.1 16-1.1 18-1.2 19-1 20-1.4.r 21-1.4.1.1.1.r 22-1.4.1.1.1 22-1.4.1.1.1.1 22-1.4.1.1.1.1.r 24-1.4.1 25-1.4.1.2.1.1 26-1.4 30-1.4.1.2 30-1.4.1.2.2 30-1.4.1.2.2.r 32-1.5.1 34-1.5.1.1 (d4 / decrease-01~e.19 :ARG1 (s / stimulate-01~e.16 :ARG1 (g / grow-01~e.15)) :ARG2 (s2 / significant-02~e.18) :time~e.0 (a / add-02~e.8 :ARG1 (m / media~e.6 :ARG1-of (d2 / deplete-01~e.5 :ARG2 (p2 / protein :name (n3 / name :op1 "IGF-1"~e.2,4))) :mod (t2 / this~e.1)) :ARG2~e.9 (a2 / and~e.11 :op1 (c / cell-line~e.13 :name (n4 / name :op1 "LM2"~e.10)) :op2 (c4 / cell-line~e.13 :name (n5 / name :op1 "JF32"~e.12)))) :compared-to~e.20 (c3 / control-01~e.26 :ARG0 (o / or~e.24 :op1 (m2 / medium~e.6 :ARG1-of~e.21 (t / treat-04~e.22 :polarity~e.22 -~e.22) :ARG1-of (c2 / condition-01 :ARG0 (m3 / macrophage))) :op2 (p / protein~e.30 :name (n2 / name :op1 "IgG"~e.25) :ARG1-of~e.30 (i / identical-01~e.30 :ARG2 m2) :ARG1-of t))) :ARG1-of (d3 / describe-01 :ARG0 (f / figure~e.32 :mod "8B"~e.34))) # ::id a_pmid_2169_9731.154 ::amr-annotator SDL-AMR-09 ::preferred # ::tok In addition , MH @-@ S macrophage IGF @-@ 1 secretion was interrupted by transfection with scrambled control ( scr siRNA ) or siRNA constructs designed against mouse IGF @-@ 1 (α-IGF siRNA ) . # ::alignments 0-1 1-1 3-1.1.2.1.1.1.1 5-1.1.2.1.1.1.1 6-1.1.2.1 7-1.1.2.2.1.1 9-1.1.2.2.1.1 10-1.1.2 12-1.1 13-1.1.1.r 14-1.1.1 15-1.1.1.1.r 16-1.1.1.1.1.1 17-1.1.1.1.1 20-1.1.1.1.1.2.1.1.1 20-1.1.1.1.2.1.1.1 22-1.1.1.1 23-1.1.1.1.1.2.1.1.1 23-1.1.1.1.2.1.1.1 24-1.1.1.1.2 25-1.1.1.1.2.2 26-1.1.1.1.2.2.1 27-1.1.1.1.2.2.1.1.2 28-1.1.1.1.2.2.1.1.1.1 28-1.1.1.1.2.2.1.1.3.1.2.1.1 30-1.1.1.1.2.2.1.1.1.1 32-1.1.1.1.1.2.1.1.1 32-1.1.1.1.2.1.1.1 32-1.1.1.1.2.2.1.1.3.1.1.1 (a / and~e.0,1 :op2 (i / interrupt-01~e.12 :ARG0~e.13 (t / transfect-01~e.14 :ARG2~e.15 (o / or~e.22 :op1 (c / control~e.17 :ARG1-of (s2 / scramble-02~e.16) :ARG1-of (m4 / mean-01 :ARG2 (n8 / nucleic-acid :name (n7 / name :op1 "siRNA"~e.20,23,32) :ARG1-of s2))) :op2 (c2 / construct~e.24 :mod (n9 / nucleic-acid :name (n3 / name :op1 "siRNA"~e.20,23,32)) :ARG1-of (d / design-01~e.25 :purpose (o2 / oppose-01~e.26 :ARG1 (p2 / protein :name (n4 / name :op1 "IGF-1"~e.28,30) :source (m2 / mouse~e.27) :ARG1-of (m3 / mean-01 :ARG2 (n10 / nucleic-acid :name (n5 / name :op1 "siRNA"~e.32) :mod (p3 / protein :name (n6 / name :op1 "α-IGF"~e.28)))))))))) :ARG1 (s / secrete-01~e.10 :ARG0 (m / macrophage~e.6 :mod (c3 / cell-line :name (n / name :op1 "MH-S"~e.3,5))) :ARG1 (p / protein :name (n2 / name :op1 "IGF-1"~e.7,9))))) # ::id a_pmid_2169_9731.155 ::amr-annotator SDL-AMR-09 ::preferred # ::tok One α-IGF siRNA construct was more effective than the scr siRNA , and significantly decreased MØCM IGF @-@ 1 levels to 25 % of control ( Figure 8C ) . # ::alignments 0-1.1.1.1 1-1.1.1.2.2.1.1 2-1.1.1.2.1.1 3-1.1.1 5-1.1.2 6-1.1 7-1.1.3.r 10-1.1.3.1.1 12-1 13-1.2.2 14-1.2 16-1.2.1.1.1.1 18-1.2.1.1.1.1 19-1.2.1 20-1.2.3.r 21-1.2.3.1.1 22-1.2.3.1 23-1.2.3.1.r 24-1.2.3 26-1.3.1 28-1.3.1.1 (a / and~e.12 :op1 (e / effective-04~e.6 :ARG0 (c / construct-01~e.3 :quant 1~e.0 :ARG2 (n6 / nucleic-acid :name (n / name :op1 "siRNA"~e.2) :mod (p2 / protein :name (n2 / name :op1 "α-IGF"~e.1)))) :degree (m / more~e.5) :compared-to~e.7 (n7 / nucleic-acid :name (n3 / name :op1 "siRNA"~e.10) :ARG1-of (s / scramble-02))) :op2 (d / decrease-01~e.14 :ARG1 (l / level~e.19 :quant-of (p3 / protein :name (n4 / name :op1 "IGF-1"~e.16,18) :mod (m2 / medium :ARG1-of (c2 / condition-01 :ARG0 (m3 / macrophage))))) :ARG2 (s2 / significant-02~e.13) :ARG4~e.20 (c3 / control~e.24 :quant~e.23 (p / percentage-entity~e.22 :value 25~e.21))) :ARG1-of (d2 / describe-01 :ARG0 (f / figure~e.26 :mod "8C"~e.28))) # ::id a_pmid_2169_9731.156 ::amr-annotator SDL-AMR-09 ::preferred # ::tok The scr siRNA construct decreased macrophage IGF @-@ 1 secretion to a lesser extent ( Figure 8C ) . # ::alignments 2-1.1.1.1.1 3-1.1 4-1 5-1.2.1 6-1.2.2.1.1 8-1.2.2.1.1 9-1.2 10-1.3.r 12-1.3.1 13-1.3 15-1.4.1 17-1.4.1.1 (d / decrease-01~e.4 :ARG0 (c / construct~e.3 :topic (n3 / nucleic-acid :name (n / name :op1 "siRNA"~e.2) :ARG1-of (s / scramble-02))) :ARG1 (s2 / secrete-01~e.9 :ARG0 (m / macrophage~e.5) :ARG1 (p / protein :name (n2 / name :op1 "IGF-1"~e.6,8))) :ARG4~e.10 (e / extent~e.13 :degree (l / less~e.12)) :ARG1-of (d2 / describe-01 :ARG0 (f / figure~e.15 :mod "8C"~e.17))) # ::id a_pmid_2169_9731.157 ::amr-annotator SDL-AMR-09 ::preferred # ::tok Transfection reagents and conditions were chosen to minimize cellular toxicity , and media IGF @-@ 1 content significantly decreased when normalized to MH @-@ S viability ( media IGF @-@ 1/MTS relative viability , data not shown ) . # ::alignments 0-1.1.1.1.1 1-1.1.1.1 2-1.1.1 3-1.1.1.2 5-1.1 7-1.1.2 8-1.1.2.2.1 9-1.1.2.2 11-1.1.1 12-1.2.1.1.2 13-1.2.1.1.1.1 15-1.2.1.1.1.1 16-1.2.1 17-1.2.2 18-1.2 19-1.2.3.r 20-1.2.3 22-1.2.3.2.1.1.1.1 24-1.2.3.2.1.1.1.1 27-1.2.3.2.2.1.1 28-1.2.3.2.2.1.1 31-1.2.3.2.2.1.2.1 34-1.2.3.2.2.1.3.2 35-1.2.3.2.2.1.3.1 35-1.2.3.2.2.1.3.1.r 36-1.2.3.2.2.1.3 (a / and :op1 (c / choose-01~e.5 :ARG1 (a2 / and~e.2,11 :op1 (r / reagent~e.1 :ARG2-of (t / transfect-01~e.0)) :op2 (c2 / condition~e.3 :mod t)) :purpose (m / minimize-01~e.7 :ARG0 a2 :ARG1 (t2 / toxicity~e.9 :mod (c3 / cell~e.8)))) :op2 (d / decrease-01~e.18 :ARG1 (c4 / content~e.16 :quant-of (p / protein :name (n / name :op1 "IGF-1"~e.13,15) :mod (m2 / media~e.12))) :ARG2 (s / significant-02~e.17) :time~e.19 (n2 / normalize-01~e.20 :ARG1 c4 :topic (v / viable :domain (m3 / macrophage :mod (c5 / cell-line :name (n3 / name :op1 "MH-S"~e.22,24))) :ARG1-of (m4 / mean-01 :ARG2 (s2 / slash :op1 p~e.27,28 :op2 (v2 / viable :ARG2-of (r2 / relative-05~e.31)) :ARG1-of (s3 / show-01~e.36 :polarity~e.35 -~e.35 :ARG0 (d2 / data~e.34)))))))) # ::id a_pmid_2169_9731.158 ::amr-annotator SDL-AMR-09 ::preferred # ::tok Neoplastic growth reflected the level of IGF @-@ 1 in the media conditioned by siRNA @-@ treated macrophages , with all three groups differing significantly in JF32 cells ( Figure 8D ) . # ::alignments 0-1.1.1.1 1-1.1.1 2-1.1 4-1.1.2 5-1.1.2.1.r 6-1.1.2.1.1.1 8-1.1.2.1.1.1 9-1.1.3.r 11-1.1.3 12-1.1.3.1 13-1.1.3.1.1.r 14-1.1.3.1.1.1.1.1.1 16-1.1.3.1.1.1 17-1.1.3.1.1 20-1.2.1.2 21-1.2.1.1 22-1.2.1 23-1.2 24-1.2.2 25-1.2.3.r 26-1.2.3.1.1 27-1.2.3 29-1.1.4.1 31-1.1.4.1.1 (a / and :op1 (r / reflect-01~e.2 :ARG1 (g / grow-01~e.1 :ARG1 (n / neoplasm~e.0)) :ARG2 (l / level~e.4 :quant-of~e.5 (p / protein :name (n2 / name :op1 "IGF-1"~e.6,8))) :location~e.9 (m / media~e.11 :ARG1-of (c / condition-01~e.12 :ARG0~e.13 (m2 / macrophage~e.17 :ARG1-of (t / treat-04~e.16 :ARG2 (n5 / nucleic-acid :name (n3 / name :op1 "siRNA"~e.14)))))) :ARG1-of (d2 / describe-01 :ARG0 (f / figure~e.29 :mod "8D"~e.31))) :op2 (d / differ-02~e.23 :ARG1 (g2 / group~e.22 :quant 3~e.21 :mod (a2 / all~e.20)) :ARG1-of (s / significant-02~e.24) :location~e.25 (c2 / cell~e.27 :name (n4 / name :op1 "JF32"~e.26)))) # ::id a_pmid_2169_9731.159 ::amr-annotator SDL-AMR-09 ::preferred # ::tok While scr siRNA @-@ treated media did not significantly lower LM2 cell growth , the correlation of media IGF @-@ 1 levels vs. LM2 proliferation was highly significant , as in JF32 cells ( Figure 8D @-@ F ) . # ::alignments 0-1 2-1.1.2.1.1.1.1 4-1.1.2.1 5-1.1.2 7-1.1.1 7-1.1.1.r 8-1.1.4 9-1.1 10-1.1.3.1.1.1 11-1.1.3.1 12-1.1.3 15-1.2 16-1.2.1.r 17-1.2.1.1.2 18-1.2.1.1.1.1 20-1.2.1.1.1.1 21-1.2.1 22-1 23-1.1.3.1.1.1 24-1.2.2 24-1.2.4.1 26-1.2.3.1 27-1.2.3 31-1.2.4.1.1.1.1 32-1.1.3.1 32-1.2.4.1.1 34-1.3.1.1 34-1.3.1.2 34-1.3.1.3 36-1.3.1.1.1 (c / contrast-01~e.0,22 :ARG1 (l2 / lower-05~e.9 :polarity~e.7 -~e.7 :ARG0 (m2 / medium~e.5 :ARG1-of (t / treat-04~e.4 :ARG2 (n5 / nucleic-acid :name (n / name :op1 "siRNA"~e.2) :ARG1-of (s2 / scramble-02)))) :ARG1 (g / grow-01~e.12 :ARG1 (c2 / cell-line~e.11,32 :name (n2 / name :op1 "LM2"~e.10,23))) :ARG1-of (s / significant-02~e.8)) :ARG2 (c3 / correlate-01~e.15 :ARG1~e.16 (l3 / level~e.21 :quant-of (p / protein :name (n3 / name :op1 "IGF-1"~e.18,20) :mod (m / medium~e.17))) :ARG2 (p2 / proliferate-01~e.24 :ARG0 c2) :ARG1-of (s3 / significant-02~e.27 :ARG1-of (h / high-02~e.26)) :ARG1-of (r2 / resemble-01 :ARG2 (p3 / proliferate-01~e.24 :ARG0 (c4 / cell-line~e.32 :name (n4 / name :op1 "JF32"~e.31))))) :ARG1-of (d / describe-01 :ARG0 (a / and :op1 (f / figure~e.34 :mod "8D"~e.36) :op2 (f2 / figure~e.34 :mod "8E") :op3 (f3 / figure~e.34 :mod "8F")))) # ::id a_pmid_2169_9731.160 ::amr-annotator SDL-AMR-09 ::preferred # ::tok Taken together , these experiments demonstrate that IGF @-@ 1 is responsible for the majority of neoplastic growth stimulated by MØCM . # ::alignments 0-1 1-1.2 3-1.1.2 4-1.1 5-1.1.1 6-1.1.1.1.r 7-1.1.1.1.1.1.1 9-1.1.1.1.1.1.1 11-1.1.1.1 12-1.1.1.1.2.r 14-1.1.1.1.2.2 16-1.1.1.1.2.1 17-1.1.1.1.2 18-1.1.1.1.2.3 (t / take-01~e.0 :ARG1 (e / experiment-01~e.4 :ARG0-of (d / demonstrate-01~e.5 :ARG1~e.6 (r / responsible-01~e.11 :ARG0 (p / protein :name (n / name :op1 "IGF-1"~e.7,9)) :ARG1~e.12 (g / grow-01~e.17 :ARG1 (n2 / neoplasm~e.16) :mod (m / majority~e.14) :ARG1-of (s / stimulate-01~e.18 :ARG0 (m2 / medium :ARG1-of (c / condition-01 :ARG0 (m3 / macrophage))))))) :mod (t3 / this~e.3)) :mod (t2 / together~e.1)) # ::id a_pmid_2169_9731.161 ::amr-annotator SDL-AMR-09 ::preferred # ::tok Combined MEK and PI3K inhibition blocks IGF @-@ 1 and MØCM induced neoplastic proliferation by decreasing cyclin D1 expression # ::alignments 0-1.1.1 1-1.1.1.1.1.1.1 3-1.1.1.2.1.1.1 4-1.1.1.1 4-1.1.1.2 5-1.1 6-1.1.2.1.1 8-1.1.2.1.1 9-1 11-1.2 12-1.2.2.1 13-1.2.2 14-1.2.3.r 15-1.2.3 16-1.2.3.1.1.1.1 17-1.2.3.1.1.1.1 18-1.2.3.1 (a / and~e.9 :op1 (b / block-01~e.5 :ARG0 (c / combine-01~e.0 :ARG1 (i2 / inhibit-01~e.4 :ARG1 (e2 / enzyme :name (n2 / name :op1 "MEK"~e.1))) :ARG2 (i3 / inhibit-01~e.4 :ARG1 (e3 / enzyme :name (n3 / name :op1 "PI3K"~e.3)))) :ARG1 (p / protein :name (n4 / name :op1 "IGF-1"~e.6,8))) :op2 (i / induce-01~e.11 :ARG0 (m / medium :ARG1-of (c2 / condition-01 :ARG0 (m2 / macrophage))) :ARG2 (p2 / proliferate-01~e.13 :ARG0 (n6 / neoplasm~e.12)) :instrument~e.14 (d / decrease-01~e.15 :ARG1 (e4 / express-03~e.18 :ARG2 (p3 / protein :name (n7 / name :op1 "cyclin-D1"~e.16,17)))))) # ::id a_pmid_2169_9731.162 ::amr-annotator SDL-AMR-09 ::preferred # ::tok IGF @-@ 1 stimulated neoplastic proliferation and mediated a significant portion of macrophage @-@ induced tumor cell growth in culture . # ::alignments 0-1.1.1.1.1 2-1.1.1.1.1 3-1.1 4-1.1.2.1 5-1.1.2 6-1 7-1.2 9-1.2.2.2 10-1.2.2 12-1.2.2.1.1.2.1 14-1.2.2.1.1.2 15-1.2.2.1.1.1.1 16-1.2.2.1.1.1 17-1.2.2.1.1 18-1.2.3.r 19-1.2.3 (a / and~e.6 :op1 (s / stimulate-01~e.3 :ARG0 (p / protein :name (n / name :op1 "IGF-1"~e.0,2)) :ARG1 (p2 / proliferate-01~e.5 :ARG0 (n2 / neoplasm~e.4))) :op2 (m / mediate-01~e.7 :ARG0 p :ARG1 (p3 / portion~e.10 :ARG1-of (i / include-01 :ARG2 (g2 / grow-01~e.17 :ARG1 (c2 / cell~e.16 :mod (t2 / tumor~e.15)) :ARG2-of (i3 / induce-01~e.14 :ARG0 (m3 / macrophage~e.12)))) :ARG1-of (s2 / significant-02~e.9)) :location~e.18 (c3 / culture~e.19))) # ::id a_pmid_2169_9731.163 ::amr-annotator SDL-AMR-09 ::preferred # ::tok To determine if MØCM and @/@ or IGF @-@ 1 were similarly blocked by MEK and PI3K inhibition , LM2 and JF32 cells were treated with combinations of MEK and @/@ or PI3K inhibitors , in the presence of IGF @-@ 1 or MØCM . # ::alignments 1-1.4 2-1.3.r 4-1.4.1.2 6-1.4.1.2 7-1.4.1.2.2 8-1.4.1.2.2 9-1.4.1.2.2 11-1.4.1.3 12-1.4.1 14-1.2.1.1.1.1.1.1 16-1.2.1.2.1.1.1.1 17-1.2.1.1 17-1.2.1.1.1 17-1.2.1.1.1.r 17-1.2.1.2 17-1.2.1.2.1 17-1.2.1.2.1.r 17-1.4.1.1 19-1.1.1.1.1 20-1.1 21-1.1.2.1.1 22-1.1.1 22-1.1.2 24-1 26-1.2 28-1.2.1.1.1.1.1.1 29-1.2.1 31-1.2.1 32-1.2.1.2.1.1.1.1 33-1.2.1.2 33-1.2.1.2.1 33-1.2.1.2.1.r 33-1.4.1.1 37-1.3 38-1.3.1.r 39-1.3.1.1.1.1 41-1.3.1.1.1.1 42-1.3.1 (t / treat-04~e.24 :ARG1 (a / and~e.20 :op1 (c / cell-line~e.22 :name (n2 / name :op1 "LM2"~e.19)) :op2 (c2 / cell-line~e.22 :name (n3 / name :op1 "JF32"~e.21))) :ARG2 (c3 / combine-01~e.26 :ARG1 (a4 / and-or~e.29,31 :op1 (m / molecular-physical-entity~e.17 :ARG0-of~e.17 (i / inhibit-01~e.17 :ARG1 (e / enzyme :name (n7 / name :op1 "MEK"~e.14,28)))) :op2 (m3 / molecular-physical-entity~e.17,33 :ARG0-of~e.17,33 (i2 / inhibit-01~e.17,33 :ARG1 (e2 / enzyme :name (n8 / name :op1 "PI3K"~e.16,32)))))) :condition~e.2 (p / present-02~e.37 :ARG1~e.38 (o / or~e.42 :op1 (p2 / protein :name (n5 / name :op1 "IGF-1"~e.39,41)) :op2 (m2 / medium :ARG1-of (c4 / condition-01 :ARG0 (m4 / macrophage))))) :purpose (d / determine-01~e.1 :ARG1 (b / block-01~e.12 :ARG0 (i3 / inhibit-01~e.17,33 :ARG1 (a3 / and :op1 e :op2 e2)) :ARG1 (a2 / and-or~e.4,6 :op1 m2 :op2 p2~e.7,8,9) :ARG1-of (r / resemble-01~e.11)))) # ::id a_pmid_2169_9731.164 ::amr-annotator SDL-AMR-09 ::preferred # ::tok Analogous to previous results with macrophage co @-@ culture , growth stimulated by either IGF @-@ 1 or MØCM was blocked by combined inhibition of MEK and PI3K , to a greater extent than either pathway by itself ( Figure 9A , B ) . # ::alignments 0-1.5 1-1.5.1.r 2-1.5.1.1.2 3-1.5.1 3-1.5.1.1 3-1.5.1.1.r 5-1.2.1.1.2.1.1 5-1.5.1.1.1.1 6-1.5.1.1.1 8-1.5.1.1.1 10-1.2 10-1.4.2.1 11-1.2.1 11-1.4.2.1.1 14-1.2.1.1.1.1.1 16-1.2.1.1.1.1.1 17-1.2.1.1 17-1.4.2.1.1.1 20-1 20-1.4.2 21-1.1.r 22-1.1.1 23-1.1 25-1.1.1.1.1.1 27-1.1.1.2.1.1 29-1.4.r 31-1.4 31-1.4.1 31-1.4.1.r 33-1.4.2.r 35-1.1.1.1 35-1.1.1.2 39-1.3.1.1 39-1.3.1.2 41-1.3.1.1.1 (b3 / block-01~e.20 :ARG0~e.21 (i / inhibit-01~e.23 :ARG3-of (c2 / combine-01~e.22 :ARG1 (p2 / pathway~e.35 :name (n / name :op1 "MEK"~e.25)) :ARG2 (p3 / pathway~e.35 :name (n4 / name :op1 "PI3K"~e.27)))) :ARG1 (g2 / grow-01~e.10 :ARG1-of (s / stimulate-01~e.11 :ARG0 (o / or~e.17 :op1 (p / protein :name (n2 / name :op1 "IGF-1"~e.14,16)) :op2 (m3 / medium :ARG1-of (c / condition-01 :ARG0 (m4 / macrophage~e.5)))))) :ARG1-of (d / describe-01 :ARG0 (a / and :op1 (f / figure~e.39 :mod "9A"~e.41) :op2 (f2 / figure~e.39 :mod "9B"))) :degree~e.29 (g / great~e.31 :degree~e.31 (m / more~e.31) :compared-to~e.33 (b2 / block-01~e.20 :ARG1 (g3 / grow-01~e.10 :ARG1-of (s2 / stimulate-01~e.11 :ARG0 (o2 / or~e.17 :op1 p :op2 m3))))) :mod (a2 / analogous~e.0 :topic~e.1 (t / thing~e.3 :ARG2-of~e.3 (r2 / result-01~e.3 :ARG1 (c3 / co-culture~e.6,8 :mod (m2 / macrophage~e.5)) :mod (p4 / previous~e.2))))) # ::id a_pmid_2169_9731.165 ::amr-annotator SDL-AMR-09 ::preferred # ::tok Consistent with the proliferation results , cyclin D1 content ( a biochemical correlate of lung cell proliferation [ @ 41 @ ]) was reduced by these inhibitors ( Figure 9C @-@ E ) . # ::alignments 0-1 1-1.2.r 3-1.2.1.1 4-1.2 4-1.2.1 4-1.2.1.r 6-1.1.2.1.1.1 7-1.1.2.1.1.1 8-1.1.2 11-1.1.2.1.2.1.2 12-1.1.2.1.2.1 12-1.1.2.1.2.1.1 12-1.1.2.1.2.1.1.r 13-1.1.2.1.2.1.1.1.r 14-1.1.2.1.2.1.1.1.1.1 15-1.1.2.1.2.1.1.1.1 16-1.1.2.1.2.1.1.1 19-1.1.2.1.2.1.1.2.1.1.1 23-1.1 24-1.1.1.r 25-1.1.1.2 26-1.1.1 26-1.1.1.1 26-1.1.1.1.r 28-1.3.1.1 28-1.3.1.2 28-1.3.1.3 30-1.3.1.1.1 (c / consistent-01~e.0 :ARG1 (r2 / reduce-01~e.23 :ARG0~e.24 (m / molecular-physical-entity~e.26 :ARG0-of~e.26 (i / inhibit-01~e.26) :mod (t / this~e.25)) :ARG1 (c2 / content~e.8 :quant-of (p2 / protein :name (n / name :op1 "cyclin-D1"~e.6,7) :ARG1-of (m2 / mean-01 :ARG2 (t3 / thing~e.12 :ARG1-of~e.12 (c3 / correlate-01~e.12 :ARG2~e.13 (p3 / proliferate-01~e.16 :ARG0 (c4 / cell~e.15 :mod (l / lung~e.14))) :ARG1-of (d / describe-01 :ARG0 (p4 / publication :ARG1-of (c5 / cite-01 :ARG2 41~e.19)))) :mod (b / biochemistry~e.11)))))) :ARG2~e.1 (t2 / thing~e.4 :ARG2-of~e.4 (r / result-01~e.4 :mod (p / proliferate-01~e.3))) :ARG1-of (d2 / describe-01 :ARG0 (a / and :op1 (f / figure~e.28 :mod "9C"~e.30) :op2 (f2 / figure~e.28 :mod "9D") :op3 (f3 / figure~e.28 :mod "9E")))) # ::id a_pmid_2169_9731.166 ::amr-annotator SDL-AMR-09 ::preferred # ::tok MØCM induced early increases in cRaf , Akt and GSK @-@ 3β phosphorylation , and Erk1 @/@ 2 phosphorylation peaked at 24 hrs ( Figure 9C , D ) . # ::alignments 1-1.1 2-1.1.2.2 3-1.1.2 4-1.1.2.1.r 5-1.1.2.1.1.1.1.1 7-1.1.2.1.1.2.1.1 8-1.1.2.1.1 9-1.1.2.1.1.3.1.1 11-1.1.2.1.1.3.1.1 12-1.1.2.1 14-1 14-1.3.1 15-1.2.1.1.1.1.1 16-1.2.1.1 18-1.2.1 19-1.2 21-1.2.2.1.1 22-1.2.2.1.2 24-1.3.1.1 24-1.3.1.2 26-1.3.1.1.1 (a / and~e.14 :op1 (i / induce-01~e.1 :ARG0 (m / medium :ARG1-of (c / condition-01 :ARG0 (m2 / macrophage))) :ARG2 (i2 / increase-01~e.3 :ARG1~e.4 (p / phosphorylate-01~e.12 :ARG1 (a2 / and~e.8 :op1 (e2 / enzyme :name (n2 / name :op1 "cRaf"~e.5)) :op2 (e3 / enzyme :name (n3 / name :op1 "Akt"~e.7)) :op3 (e4 / enzyme :name (n4 / name :op1 "GSK-3β"~e.9,11)))) :time (e / early~e.2))) :op2 (p2 / peak-01~e.19 :ARG1 (p3 / phosphorylate-01~e.18 :ARG1 (s / slash~e.16 :op1 (e5 / enzyme :name (n5 / name :op1 "Erk1"~e.15)) :op2 (e6 / enzyme :name (n6 / name :op1 "Erk2")))) :time (a3 / after :quant (t / temporal-quantity :quant 24~e.21 :unit (h / hour~e.22)))) :ARG1-of (d / describe-01 :ARG0 (a4 / and~e.14 :op1 (f / figure~e.24 :mod "9C"~e.26) :op2 (f2 / figure~e.24 :mod "9D")))) # ::id a_pmid_2169_9731.167 ::amr-annotator SDL-AMR-09 ::preferred # ::tok In both LM2 and JF32 cells , increased Akt phosphorylation corresponded to more phosphorylation of the Akt substrate , pGSK @-@ 3β ( Figure 9H and 9I ) . # ::alignments 2-1.3.1.1.1 3-1.3 4-1.3.2.1.1 5-1.3.1 5-1.3.2 7-1.1.2 8-1.1.1.1.1 9-1.1 10-1 11-1.2.r 12-1.2.2 13-1.2 13-1.2.1.2.1.2 14-1.2.1.r 16-1.2.1.1 17-1.2.1 21-1.2.1.2.1.1.1 23-1.4.1.1 23-1.4.1.2 25-1.4.1.1.1 27-1.4.1 29-1.4.1.2.1 (c / correspond-02~e.10 :ARG1 (p / phosphorylate-01~e.9 :ARG1 (e / enzyme :name (n / name :op1 "Akt"~e.8)) :ARG1-of (i / increase-01~e.7)) :ARG2~e.11 (p2 / phosphorylate-01~e.13 :ARG1~e.14 (s / substrate~e.17 :poss e~e.16 :ARG1-of (m2 / mean-01 :ARG2 (e2 / enzyme :name (n2 / name :op1 "GSK-3β"~e.21) :ARG3-of (p3 / phosphorylate-01~e.13)))) :degree (m / more~e.12)) :location (a / and~e.3 :op1 (c2 / cell-line~e.5 :name (n3 / name :op1 "LM2"~e.2)) :op2 (c3 / cell-line~e.5 :name (n4 / name :op1 "JF32"~e.4))) :ARG1-of (d / describe-01 :ARG0 (a2 / and~e.27 :op1 (f / figure~e.23 :mod "9H"~e.25) :op2 (f2 / figure~e.23 :mod "9I"~e.29)))) # ::id a_pmid_2169_9731.168 ::amr-annotator SDL-AMR-09 ::preferred # ::tok Phospho @-@ cRaf levels , another marker of Akt activity , also increased in concert with heightened increased Akt activity from 4 @-@ 24 hrs ; although p @-@ cRaf abruptly dropped at 48 hrs , pAkt and pGSK @-@ 3β levels remained highly elevated ( Figure 9F , H and 9I ) . # ::alignments 0-1.1.1.1.2 2-1.1.1.1.1.1 3-1.1.1 5-1.1.1.1.3.1.2 6-1.1.1.1.3.1 7-1.1.1.1.3.1.1.r 8-1.1.1.1.3.1.1.1.1.1 9-1.1.1.1.3.1.1 11-1.1.2 12-1.1 13-1.1.3 14-1.1.3 15-1.1.3.2.r 16-1.1.3.2.2 17-1.1.3.2.3 18-1.1.3.2.1 19-1.1.3.2 21-1.1.4.1.1 23-1.1.4.2.1 24-1.2.2.3.1.2 26-1.2 27-1.1.1.1.2 29-1.1.1.1.1.1 30-1.2.2.2 30-1.2.2.2.r 31-1.2.2 33-1.2.2.3.1.1 34-1.1.4.1.2 34-1.2.2.3.1.2 36-1.2.1.1.1.1.1.1 37-1.2.1.1 40-1.2.1.1.2.1.1.1 41-1.2.1.1.1 41-1.2.1.1.2 42-1.2.1 43-1.2.1.2.1 44-1.2.1.2 46-1.3.1.1 46-1.3.1.2 46-1.3.1.3 48-1.3.1.1.1 52-1.3.1 54-1.3.1.3.1 (a9 / and :op1 (i / increase-01~e.12 :ARG1 (l / level~e.3 :quant-of (e / enzyme :name (n / name :op1 "cRaf"~e.2,29) :ARG3-of (p / phosphorylate-01~e.0,27) :ARG1-of (m2 / mean-01 :ARG2 (m3 / marker~e.6 :mod~e.7 (a / activity-06~e.9 :ARG0 (e2 / enzyme :name (n2 / name :op1 "Akt"~e.8))) :mod (a2 / another~e.5))))) :mod (a3 / also~e.11) :manner (i2 / in-concert~e.13,14 :op1 i :op2~e.15 (a5 / activity-06~e.19 :ARG0 e2~e.18 :ARG1-of (h / heighten-01~e.16) :ARG1-of (i3 / increase-01~e.17))) :time (b / between :op1 (t / temporal-quantity :quant 4~e.21 :unit (h6 / hour~e.34)) :op2 (t4 / temporal-quantity :quant 24~e.23 :unit h6))) :op2 (h2 / have-concession-91~e.26 :ARG1 (r / remain-01~e.42 :ARG1 (a8 / and~e.37 :op1 (l2 / level~e.41 :quant-of (e3 / enzyme :name (n3 / name :op1 "Akt"~e.36) :ARG3-of p)) :op2 (l3 / level~e.41 :quant-of (e4 / enzyme :name (n4 / name :op1 "GSK-3β"~e.40) :ARG3-of p))) :ARG3 (e5 / elevate-01~e.44 :ARG1-of (h4 / high-02~e.43))) :ARG2 (d / drop-01~e.31 :ARG1 e :manner~e.30 (a6 / abrupt~e.30) :time (a7 / after :quant (t2 / temporal-quantity :quant 48~e.33 :unit (h3 / hour~e.24,34))))) :ARG1-of (d2 / describe-01 :ARG0 (a4 / and~e.52 :op1 (f / figure~e.46 :mod "9F"~e.48) :op2 (f2 / figure~e.46 :mod "9H") :op3 (f3 / figure~e.46 :mod "9I"~e.54)))) # ::id a_pmid_2169_9731.169 ::amr-annotator SDL-AMR-09 ::preferred # ::tok We observed reciprocal changes in the Erk and Akt pathways in response to their respective enzyme inhibitors . # ::alignments 0-1.1 1-1 2-1.2.2 3-1.2 6-1.2.1.1.1.1 6-1.2.3.1.1.1.1.1.1 7-1.2.1 8-1.2.1.2.1.1 8-1.2.3.1.1.1.2.1.1 9-1.2.1.1 9-1.2.1.2 10-1.2.3.r 11-1.2.3 14-1.2.3.1.2 15-1.2.3.1.1.1.1 15-1.2.3.1.1.1.2 16-1.2.3.1 16-1.2.3.1.1 16-1.2.3.1.1.r (o / observe-01~e.1 :ARG0 (w / we~e.0) :ARG1 (c / change-01~e.3 :ARG1 (a / and~e.7 :op1 (p / pathway~e.9 :name (n2 / name :op1 "Erk"~e.6)) :op2 (p2 / pathway~e.9 :name (n3 / name :op1 "Akt"~e.8))) :mod (r / reciprocal~e.2) :ARG2-of~e.10 (r2 / respond-01~e.11 :ARG1 (t / thing~e.16 :ARG0-of~e.16 (i / inhibit-01~e.16 :ARG1 (a2 / and :op1 (p3 / protein-family~e.15 :name (n4 / name :op1 "Erk"~e.6)) :op2 (p4 / protein-family~e.15 :name (n5 / name :op1 "Akt"~e.8)))) :mod (r3 / respective~e.14))))) # ::id a_pmid_2169_9731.170 ::amr-annotator SDL-AMR-09 ::preferred # ::tok In LM2 cells , MEK inhibition ( by U0126 ) suppressed early Erk1 @/@ 2 phosphorylation while p @-@ Akt levels increased . # ::alignments 1-1.3.1.1 2-1.3 4-1.1.1.2.1.1 5-1.1.1 7-1.1.1.1.r 8-1.1.1.1.1.1 10-1.1 11-1.1.2.2 12-1.1.2.1.1.1.1 13-1.1.2.1 15-1.1.2 15-1.2.1.1 15-1.2.1.1.2 15-1.2.1.1.2.r 16-1 17-1.1.2 17-1.2.1.1 17-1.2.1.1.2 17-1.2.1.1.2.r 19-1.2.1.1.1.1 20-1.2.1 21-1.2 (c / contrast-01~e.16 :ARG1 (s2 / suppress-01~e.10 :ARG0 (i / inhibit-01~e.5 :ARG0~e.7 (s / small-molecule :name (n2 / name :op1 "U0126"~e.8)) :ARG1 (e / enzyme :name (n / name :op1 "MEK"~e.4))) :ARG1 (p2 / phosphorylate-01~e.15,17 :ARG1 (s3 / slash~e.13 :op1 (e3 / enzyme :name (n3 / name :op1 "Erk1"~e.12)) :op2 (e4 / enzyme :name (n4 / name :op1 "Erk2"))) :time (e2 / early~e.11))) :ARG2 (i2 / increase-01~e.21 :ARG1 (l / level~e.20 :quant-of (e5 / enzyme~e.15,17 :name (n5 / name :op1 "Akt"~e.19) :ARG1-of~e.15,17 (p / phosphorylate-01~e.15,17)))) :location (c2 / cell-line~e.2 :name (n6 / name :op1 "LM2"~e.1))) # ::id a_pmid_2169_9731.171 ::amr-annotator SDL-AMR-09 ::preferred # ::tok Conversely , PI3K inhibition ( by LY294002 ) increased basal p @-@ Erk1 @/@ 2 levels at the expense of p @-@ Akt ( 4 hrs time point ; Figure 9C ) . # ::alignments 2-1.1.1.2.1.1 3-1.1.1 5-1.1.1.1.r 6-1.1.1.1.1.1 8-1.1 9-1.1.2.2 10-1.1.2.1.1.2 12-1.1.2.1.1.1.1 13-1.1.2.1 15-1.1.2 16-1.1.3.r 18-1.1.3 19-1.1.3 20-1.1.3.1.2 22-1.1.3.1.1.1 24-1.2.1.1 24-1.2.1.2 26-1.2.1 26-1.2.r 29-1.3.1 31-1.3.1.1 (c / contrast-01 :ARG2 (i / increase-01~e.8 :ARG0 (i2 / inhibit-01~e.3 :ARG0~e.5 (s / small-molecule :name (n2 / name :op1 "LY294002"~e.6)) :ARG1 (e2 / enzyme :name (n / name :op1 "PI3K"~e.2))) :ARG1 (l / level~e.15 :quant-of (s2 / slash~e.13 :op1 (e3 / enzyme :name (n3 / name :op1 "Erk1"~e.12) :ARG3-of (p / phosphorylate-01~e.10)) :op2 (e4 / enzyme :name (n4 / name :op1 "Erk2") :ARG3-of p)) :mod (b / basal~e.9)) :ARG0-of~e.16 (c2 / compromise-02~e.18,19 :ARG1 (e5 / enzyme :name (n5 / name :op1 "Akt"~e.22) :ARG3-of p~e.20))) :time~e.26 (a / after :quant (t2 / temporal-quantity~e.26 :quant 4~e.24 :unit 4~e.24)) :ARG1-of (d / describe-01 :ARG0 (f / figure~e.29 :mod "9C"~e.31))) # ::id a_pmid_2169_9731.172 ::amr-annotator SDL-AMR-09 ::preferred # ::tok MEK inhibition raised p @-@ Erk1 @/@ 2 and total Erk1 @/@ 2 levels at 24 and 48 hrs , while PI3K inhibition caused a compensatory increase in cellular p @-@ Akt levels from 24 @-@ 48 hrs . # ::alignments 0-1.1.1.1.1.1 1-1.1.1 2-1.1 3-1.1.2.1.1.1.2 3-1.1.2.1.2.1.2 5-1.1.2.1.1.1.1.1 5-1.1.2.2.1.1.1.1 6-1.1.2.2 9-1.1.2.2.3 10-1.1.2.2.1.1.1.1 11-1.1.2.2 13-1.1.2.1.1 13-1.1.2.1.2 13-1.1.2.2.1 13-1.1.2.2.2 15-1.1.3.1.1.1 16-1.1.2.1 17-1.1.3.2.1.1 18-1.1.3.1.1.2 18-1.1.3.2.1.2 20-1 20-1.1.3.r 21-1.2.1.1.1.1 22-1.2.1 23-1.2 26-1.2.2 27-1.2.2.1.r 28-1.2.2.1.1.2 29-1.1.2.1.1.1.2 29-1.1.2.1.2.1.2 31-1.2.2.1.1.1.1 32-1.2.2.1 34-1.1.3.1.1.1 34-1.2.2.3.1.1 36-1.1.3.2.1.1 37-1.1.3.1.1.2 37-1.1.3.2.1.2 37-1.2.2.3.1.2 (c / contrast-01~e.20 :ARG1 (r / raise-01~e.2 :ARG0 (i / inhibit-01~e.1 :ARG1 (e2 / enzyme :name (n2 / name :op1 "MEK"~e.0))) :ARG1 (a2 / and :op1 (a / and~e.16 :op1 (l / level~e.13 :quant-of (e3 / enzyme :name (n3 / name :op1 "Erk1"~e.5) :ARG3-of (p / phosphorylate-01~e.3,29))) :op2 (l2 / level~e.13 :quant-of (e4 / enzyme :name (n4 / name :op1 "Erk2") :ARG3-of (p2 / phosphorylate-01~e.3,29)))) :op2 (s / slash~e.6,11 :op1 (l3 / level~e.13 :quant-of (e5 / enzyme :name (n5 / name :op1 "Erk1"~e.5,10))) :op2 (l4 / level~e.13 :quant-of (e6 / enzyme :name (n6 / name :op1 "Erk2"))) :mod (t / total~e.9))) :time~e.20 (a4 / and :op1 (a5 / after :quant (t2 / temporal-quantity :quant 24~e.15,34 :unit (h / hour~e.18,37))) :op2 (a6 / after :quant (t3 / temporal-quantity :quant 48~e.17,36 :unit (h2 / hour~e.18,37))))) :ARG2 (c2 / cause-01~e.23 :ARG0 (i2 / inhibit-01~e.22 :ARG1 (e7 / enzyme :name (n7 / name :op1 "PI3K"~e.21))) :ARG1 (i3 / increase-01~e.26 :ARG1~e.27 (l5 / level~e.32 :quant-of (e8 / enzyme :name (n8 / name :op1 "Akt"~e.31) :mod (c4 / cell~e.28) :ARG3-of p)) :ARG0-of (c3 / compensate-01) :time (b / between :op1 (t5 / temporal-quantity :quant 24~e.34 :unit (h4 / hour~e.37)) :op2 (t6 / temporal-quantity :quant 28 :unit h4))))) # ::id a_pmid_2169_9731.173 ::amr-annotator SDL-AMR-09 ::preferred # ::tok JF32 cell growth was also suppressed by each drug ; although MEK inhibition did not affect p @-@ Erk1 @/@ 2 levels at 4 hrs , p @-@ Erk1 @/@ 2 levels decreased at 48 hrs ( Figure 9D ) . # ::alignments 0-1.1.2.1.1.1 1-1.1.2.1 2-1.1.2 4-1.1.3 5-1.1 6-1.1.1.r 7-1.1.1.1 8-1.1.1 10-1.2 11-1.2.2.2.1.1.1 12-1.2.2.2 14-1.2.2.1 14-1.2.2.1.r 15-1.2.2 16-1.2.2.3.1.1.2 18-1.2.2.3.1.1.1.1 19-1.2.2.3 21-1.2.2.3.1 21-1.2.2.3.2 23-1.2.2.4.1.1 24-1.2.2.4.1.2 26-1.2.2.3.1.1.2 28-1.2.2.3.1.1.1.1 29-1.2.2.3 31-1.2.2.3.1 31-1.2.2.3.2 32-1.2.1 34-1.2.1.2.1.1 35-1.2.1.2.1.2 37-1.3.1 39-1.3.1.1 (a5 / and :op1 (s / suppress-01~e.5 :ARG0~e.6 (d / drug~e.8 :mod (e2 / each~e.7)) :ARG1 (g / grow-01~e.2 :ARG1 (c / cell-line~e.1 :name (n2 / name :op1 "JF32"~e.0))) :mod (a / also~e.4)) :op2 (h / have-concession-91~e.10 :ARG1 (d2 / decrease-01~e.32 :ARG1 s2 :time (a4 / after :quant (t2 / temporal-quantity :quant 48~e.34 :unit (h3 / hour~e.35)))) :ARG2 (a2 / affect-01~e.15 :polarity~e.14 -~e.14 :ARG0 (i / inhibit-01~e.12 :ARG1 (e3 / enzyme :name (n3 / name :op1 "MEK"~e.11))) :ARG1 (s2 / slash~e.19,29 :op1 (l / level~e.21,31 :quant-of (e4 / enzyme :name (n4 / name :op1 "Erk1"~e.18,28) :ARG3-of (p / phosphorylate-01~e.16,26))) :op2 (l2 / level~e.21,31 :quant-of (e5 / enzyme :name (n5 / name :op1 "Erk2") :ARG3-of p))) :time (a3 / after :quant (t / temporal-quantity :quant 4~e.23 :unit (h2 / hour~e.24))))) :ARG1-of (d3 / describe-01 :ARG0 (f / figure~e.37 :mod "9D"~e.39))) # ::id a_pmid_2169_9731.174 ::amr-annotator SDL-AMR-09 ::preferred # ::tok PI3K inhibition stimulated Erk1 @/@ 2 phosphorylation from 4 @-@ 24 hrs , and increased Akt phosphorylation throughout the treatment time @-@ course ( Figure 9G , H ) . # ::alignments 0-1.1.1.1.1.1 1-1.1.1 2-1.1 3-1.1.2.1.1.1.1 4-1.1.2.1 6-1.1.2 8-1.1.3.1.1 10-1.1.3.2.1 11-1.1.3.1.2 13-1 13-1.1.3 13-1.3.1 14-1.2 15-1.2.1.1.1.1 16-1.2.1 19-1.2.2 20-1.1.3.1 20-1.1.3.2 20-1.1.3.r 20-1.2.2.r 24-1.3.1.1 24-1.3.1.2 26-1.3.1.1.1 (a / and~e.13 :op1 (s / stimulate-01~e.2 :ARG0 (i2 / inhibit-01~e.1 :ARG1 (e / enzyme :name (n / name :op1 "PI3K"~e.0))) :ARG1 (p2 / phosphorylate-01~e.6 :ARG1 (s2 / slash~e.4 :op1 (e2 / enzyme :name (n2 / name :op1 "Erk1"~e.3)) :op2 (e3 / enzyme :name (n3 / name :op1 "Erk2")))) :time~e.20 (b / between~e.13 :op1 (t3 / temporal-quantity~e.20 :quant 4~e.8 :unit (h2 / hour~e.11)) :op2 (t4 / temporal-quantity~e.20 :quant 24~e.10 :unit h2))) :op2 (i / increase-01~e.14 :ARG1 (p3 / phosphorylate-01~e.16 :ARG1 (e4 / enzyme :name (n4 / name :op1 "Akt"~e.15))) :time~e.20 (t2 / treat-04~e.19)) :ARG1-of (d / describe-01 :ARG0 (a2 / and~e.13 :op1 (f / figure~e.24 :mod "9G"~e.26) :op2 (f2 / figure~e.24 :mod "9H")))) # ::id a_pmid_2169_9731.175 ::amr-annotator SDL-AMR-09 ::preferred # ::tok While each inhibitor decreased basal proliferation rates ( Figure 9A , B ) , combinations of kinase inhibitors and MØCM increased cRaf , Erk1 @/@ 2 , Akt and GSK @-@ 3β phosphorylation in an additive manner , with the highest levels observed in cells treated with both kinase inhibitors and MØCM (" U+L +", Figure 9C @-@ I ) . # ::alignments 0-1 1-1.1.1.2 2-1.1.1 2-1.1.1.1 2-1.1.1.1.r 3-1.1 4-1.1.2.2 5-1.1.2.1 6-1.1.2 8-1.1.3.1.1 10-1.1.3.1.1.1 16-1.2.1 17-1.2.1.1.r 18-1.2.1.1.1.1 19-1.2.1.1 19-1.2.1.1.1 19-1.2.1.1.1.r 22-1.2 23-1.2.2.1.1.1.1 25-1.2.2.1.2.1.1.1 26-1.2.2.1.2 29-1.2.2.1.3.1.1 30-1.2.2.1 31-1.2.2.1.4.1.1 33-1.2.2.1.4.1.1 34-1.2.2 35-1.2.3.r 37-1.2.3 38-1.2.4.r 42-1.2.4.1.1 42-1.2.4.1.1.1 42-1.2.4.1.1.1.r 43-1.2.4.1 44-1.2.4 45-1.2.4.2.r 46-1.2.4.2 47-1.2.4.2.1 50-1.2.4.2.1.1.1 51-1.2.4.2.1.1.1 52-1.1.3.1 52-1.3.1 57-1.1.3.1.2 57-1.3.1.1 57-1.3.1.2 57-1.3.1.3 57-1.3.1.4 57-1.3.1.5 57-1.3.1.6 57-1.3.1.7 59-1.3.1.1.1 (c / contrast-01~e.0 :ARG1 (d / decrease-01~e.3 :ARG0 (m2 / molecular-physical-entity~e.2 :ARG0-of~e.2 (i2 / inhibit-01~e.2) :mod (e / each~e.1)) :ARG1 (r / rate~e.6 :mod (p2 / proliferate-01~e.5) :mod (b / basal~e.4)) :ARG1-of (d2 / describe-01 :ARG0 (a / and~e.52 :op1 (f / figure~e.8 :mod "9A"~e.10) :op2 (f2 / figure~e.57 :mod "9B")))) :ARG2 (i3 / increase-01~e.22 :ARG0 (c2 / combine-01~e.16 :ARG1~e.17 (m3 / molecular-physical-entity~e.19 :ARG0-of~e.19 (i4 / inhibit-01~e.19 :ARG1 (k / kinase~e.18))) :ARG2 (m / medium :ARG1-of (c3 / condition-01 :ARG0 (m4 / macrophage)))) :ARG1 (p3 / phosphorylate-01~e.34 :ARG1 (a2 / and~e.30 :op1 (e3 / enzyme :name (n3 / name :op1 "cRaf"~e.23)) :op2 (s / slash~e.26 :op1 (e4 / enzyme :name (n4 / name :op1 "Erk1"~e.25)) :op2 (e5 / enzyme :name (n5 / name :op1 "Erk2"))) :op3 (e6 / enzyme :name (n6 / name :op1 "Akt"~e.29)) :op4 (e7 / enzyme :name (n7 / name :op1 "GSK-3β"~e.31,33)))) :manner~e.35 (a3 / add-02~e.37) :manner~e.38 (o / observe-01~e.44 :ARG1 (l / level~e.43 :ARG1-of (h / high-02~e.42 :degree~e.42 (m5 / most~e.42))) :location~e.45 (c4 / cell~e.46 :ARG1-of (t / treat-04~e.47 :ARG2 (a4 / and :op1 m3~e.50,51 :op2 m))))) :ARG1-of (d3 / describe-01 :ARG0 (a5 / and~e.52 :op1 (f3 / figure~e.57 :mod "9C"~e.59) :op2 (f4 / figure~e.57 :mod "9D") :op3 (f5 / figure~e.57 :mod "9E") :op4 (f6 / figure~e.57 :mod "9F") :op5 (f7 / figure~e.57 :mod "9G") :op6 (f8 / figure~e.57 :mod "9H") :op7 (f9 / figure~e.57 :mod "9I")))) # ::id a_pmid_2169_9731.176 ::amr-annotator SDL-AMR-09 ::preferred # ::tok Total and p @-@ cRaf , p @-@ Akt and p @-@ GSK @-@ 3β were each significantly higher after 4 @-@ 24 hrs of treatment in all groups receiving any combination of drug and MØCM , and p @-@ Erk1 @/@ 2 levels spiked after 24 hrs of treatment ( Figure 9F @-@ I ) . # ::alignments 0-1.1.1.1.1.2 1-1.1.1 1-1.1.1.1 1-1.1.1.1.r 1-1.1.1.2 1-1.1.1.2.r 2-1.1.1.2.1.2 4-1.1.1.1.1.1.1 4-1.1.1.2.1.1.1 6-1.1.1.2.1.2 8-1.1.1.1.2.1.1 8-1.1.1.2.2.1.1 9-1.1.1.2 10-1.1.1.2.1.2 12-1.1.1.1.3.1.1 12-1.1.1.2.3.1.1 14-1.1.1.1.3.1.1 14-1.1.1.2.3.1.1 17-1.1.3 18-1.1 18-1.1.2 18-1.1.2.r 19-1.1.4 20-1.1.4.2.1.1 22-1.1.4.2.1.2 23-1.1.4.2.2 24-1.1.4.1.r 25-1.1.4.1 26-1.1.5.r 27-1.1.5.1 28-1.1.5 29-1.1.5.2 30-1.1.5.2.1.3 31-1.1.5.2.1 32-1.1.5.2.1.1.r 33-1.1.5.2.1.1 34-1.1.1 34-1.1.1.1 34-1.1.1.1.r 34-1.1.1.2 34-1.1.1.2.r 37-1.1.1 37-1.1.1.1 37-1.1.1.1.r 37-1.1.1.2 37-1.1.1.2.r 38-1.1.1.2.1.2 40-1.2.1.1.1.1.1 41-1.2.1.1 43-1.2.1 44-1.2 45-1.2.2 46-1.2.2.2.1 47-1.2.2.2.2 48-1.2.2.1.r 49-1.2.2.1 51-1.3.1.1 51-1.3.1.2 51-1.3.1.3 51-1.3.1.4 53-1.3.1.1.1 (a6 / and :op1 (h / high-02~e.18 :ARG1 (a / and~e.1,34,37 :op1~e.1,34,37 (a2 / and~e.1,34,37 :op1 (e / enzyme :name (n / name :op1 "cRaf"~e.4) :mod (t / total~e.0)) :op2 (e2 / enzyme :name (n2 / name :op1 "Akt"~e.8) :mod t) :op3 (e3 / enzyme :name (n3 / name :op1 "GSK-3β"~e.12,14) :mod t)) :op2~e.1,34,37 (a3 / and~e.1,9,34,37 :op1 (e4 / enzyme :name (n4 / name :op1 "cRaf"~e.4) :ARG3-of (p / phosphorylate-01~e.2,6,10,38)) :op2 (e5 / enzyme :name (n5 / name :op1 "Akt"~e.8) :ARG3-of p) :op3 (e6 / enzyme :name (n6 / name :op1 "GSK-3β"~e.12,14) :ARG3-of p))) :degree~e.18 (m2 / more~e.18) :ARG1-of (s / significant-02~e.17) :time (a4 / after~e.19 :op1~e.24 (t3 / treat-04~e.25) :quant (t2 / temporal-quantity :quant (v / value-interval :op1 4~e.20 :op2 24~e.22) :unit (h3 / hour~e.23))) :location~e.26 (g / group~e.28 :mod (a5 / all~e.27) :ARG0-of (r / receive-01~e.29 :ARG1 (c / combine-01~e.31 :ARG1~e.32 (d / drug~e.33) :ARG2 (m / medium :ARG1-of (c2 / condition-01 :ARG0 (m3 / macrophage))) :mod (a8 / any~e.30))))) :op2 (s2 / spike-04~e.44 :ARG1 (l / level~e.43 :quant-of (s3 / slash~e.41 :op1 (e7 / enzyme :name (n8 / name :op1 "Erk1"~e.40)) :op2 (e8 / enzyme :name (n9 / name :op1 "Erk2")))) :time (a7 / after~e.45 :op1~e.48 (t4 / treat-04~e.49) :quant (t5 / temporal-quantity :quant 24~e.46 :unit (h4 / hour~e.47)))) :ARG1-of (d2 / describe-01 :ARG0 (a9 / and :op1 (f / figure~e.51 :mod "9F"~e.53) :op2 (f2 / figure~e.51 :mod "9G") :op3 (f3 / figure~e.51 :mod "9H") :op4 (f4 / figure~e.51 :mod "9I")))) # ::id a_pmid_2169_9731.177 ::amr-annotator SDL-AMR-09 ::preferred # ::tok Either inhibitor alone partially prevented the increase in cyclin D1 in cells treated with MØCM ; cells receiving both inhibitors had the lowest cyclin D1 levels and were unresponsive to MØCM @-@ induced growth ( Figure 9E ) . # ::alignments 0-1.1.1.2 1-1.1.1 1-1.1.1.1 1-1.1.1.1.r 2-1.1.1.3 3-1.1.3 3-1.1.3.r 4-1.1 6-1.1.2 7-1.1.2.1.r 8-1.1.2.1.1.1 9-1.1.2.1.1.1 10-1.1.2.2.r 11-1.1.2.2 12-1.1.2.2.1 16-1.2.1.1 17-1.2.1.1.1 18-1.2.1.1.1.1.2 19-1.2.1.1.1.1.1 20-1.2.1 22-1.2.1.2.2 22-1.2.1.2.2.1 22-1.2.1.2.2.1.r 23-1.2.1.2.1 24-1.2.1.2.1 25-1.2.1.2 26-1.2 28-1.2.2 28-1.2.2.1 28-1.2.2.1.r 32-1.2.2.3.1 33-1.2.2.3 35-1.3.1 37-1.3.1.1 (a / and :op1 (p / prevent-01~e.4 :ARG0 (m / molecular-physical-entity~e.1 :ARG0-of~e.1 (i2 / inhibit-01~e.1) :mod (e / either~e.0) :mod (a3 / alone~e.2)) :ARG1 (i3 / increase-01~e.6 :ARG1~e.7 (p3 / protein :name (n / name :op1 "cyclin-D1"~e.8,9)) :location~e.10 (c / cell~e.11 :ARG1-of (t / treat-04~e.12 :ARG2 (m4 / medium :ARG1-of (c2 / condition-01 :ARG0 (m5 / macrophage)))))) :degree~e.3 (p2 / part~e.3)) :op2 (a2 / and~e.26 :op1 (h / have-03~e.20 :ARG0 (c3 / cell~e.16 :ARG0-of (r / receive-01~e.17 :ARG1 (m3 / molecular-physical-entity :ARG0-of i2~e.19 :mod (b / both~e.18)))) :ARG1 (l / level~e.25 :quant-of p3~e.23,24 :ARG1-of (l2 / low-04~e.22 :degree~e.22 (m2 / most~e.22)))) :op2 (r2 / respond-01~e.28 :polarity~e.28 -~e.28 :ARG0 c3 :ARG1 (g / grow-01~e.33 :ARG2-of (i4 / induce-01~e.32 :ARG0 m4)))) :ARG1-of (d / describe-01 :ARG0 (f / figure~e.35 :mod "9E"~e.37))) # ::id a_pmid_2169_9731.178 ::amr-annotator SDL-AMR-09 ::preferred # ::tok Taken together , MØCM @-@ induced neoplastic Akt and Erk1 @/@ 2 phosphorylation was magnified several @-@ fold by inhibitor treatment , dissociating kinase activity from proliferation in drug @-@ treated cells ; however , cyclin D1 levels were suppressed by either drug alone , which corresponded to decreased cell proliferation . # ::alignments 0-1.2.1.3 1-1.2.1.3.1 5-1.2.3 6-1.2.2 7-1.2.1.1.1.1 8-1.2.1 9-1.2.1.2.1.1.1 10-1.2.1.2 12-1.2 14-1 15-1.4.1 17-1.4 18-1.1.r 19-1.1.1 19-1.1.1.1 19-1.1.1.1.r 20-1.1 22-1.2.4 23-1.2.4.1.1 24-1.2.4.1 26-1.2.4.2 28-1.2.4.2.2.1.1 30-1.2.4.2.2.1 31-1.2.4.2.2 33-1.3 35-1.3.1.2.1.1.1 36-1.3.1.2.1.1.1 37-1.3.1.2 39-1.3.1 40-1.3.1.1.r 41-1.3.1.1.1 42-1.3.1.1 43-1.3.1.1.2 46-1.3.1.3 48-1.3.1.3.1.2 49-1.2.4.2.2 49-1.3.1.3.1.1 50-1.2.4.2 50-1.3.1.3.1 (m2 / magnify-01~e.14 :ARG0~e.18 (t / treat-04~e.20 :ARG2 (m / molecular-physical-entity~e.19 :ARG0-of~e.19 (i4 / inhibit-01~e.19))) :ARG1 (p2 / phosphorylate-01~e.12 :ARG1 (a / and~e.8 :op1 (e / enzyme :name (n / name :op1 "Akt"~e.7)) :op2 (s / slash~e.10 :op1 (e2 / enzyme :name (n2 / name :op1 "Erk1"~e.9)) :op2 (e3 / enzyme :name (n3 / name :op1 "Erk2"))) :ARG1-of (t2 / take-01~e.0 :mod (t3 / together~e.1))) :mod (n4 / neoplasm~e.6) :ARG2-of (i2 / induce-01~e.5 :ARG0 (m3 / medium :ARG1-of (c / condition-01 :ARG0 (m4 / macrophage)))) :ARG0-of (d / dissociate-01~e.22 :ARG1 (a2 / activity-06~e.24 :ARG0 (k / kinase~e.23)) :ARG2 (p5 / proliferate-01~e.26,50 :ARG0 (p4 / protein) :location (c2 / cell~e.31,49 :ARG1-of (t5 / treat-03~e.30 :ARG3 (d2 / drug~e.28)))))) :ARG1-of (c5 / contrast-01~e.33 :ARG2 (s3 / suppress-01~e.39 :ARG0~e.40 (d4 / drug~e.42 :mod (e4 / either~e.41) :mod (a3 / alone~e.43)) :ARG1 (l / level~e.37 :quant-of (p6 / protein :name (n7 / name :op1 "cyclin-D1"~e.35,36))) :ARG1-of (c3 / correspond-02~e.46 :ARG2 (p7 / proliferate-01~e.50 :ARG0 (c4 / cell~e.49) :ARG1-of (d3 / decrease-01~e.48))))) :quant (p3 / product-of~e.17 :op1 (s2 / several~e.15))) # ::id a_pmid_2169_9731.179 ::amr-annotator SDL-AMR-09 ::preferred # ::tok As with MØCM , IGF @-@ 1 stimulated both Akt and Erk1 @/@ 2 activities . # ::alignments 4-1.1.1.1.1 6-1.1.1.1.1 7-1.1 9-1.1.2.1.1.1.1 10-1.1.2.1 11-1.1.2.1.2.1.1.1 12-1.1.2.1.2 14-1.1.2 (r / resemble-01 :ARG1 (s / stimulate-01~e.7 :ARG0 (p / protein :name (n2 / name :op1 "IGF-1"~e.4,6)) :ARG1 (a / act-02~e.14 :ARG0 (a2 / and~e.10 :op1 (e / enzyme :name (n3 / name :op1 "Akt"~e.9)) :op2 (s2 / slash~e.12 :op1 (e2 / enzyme :name (n4 / name :op1 "Erk1"~e.11)) :op2 (e3 / enzyme :name (n5 / name :op1 "Erk2")))))) :ARG2 (m / medium :ARG1-of (c / condition-01 :ARG0 (m2 / macrophage)))) # ::id a_pmid_2169_9731.180 ::amr-annotator SDL-AMR-09 ::preferred # ::tok Kinase activation was greatest within 4 hrs of treatment , and remained elevated 48 hrs later , corresponding with increased cyclin D1 expression ( Figure 10A @-@ E ) . # ::alignments 0-1.1.1.1 1-1.1.1 2-1.1.1.r 3-1.1 3-1.1.2 3-1.1.2.r 4-1.1.3 4-1.1.3.2 4-1.1.3.2.1.1.r 4-1.1.3.2.r 4-1.2.3 5-1.1.3.2.1.1 6-1.1.3.2.1.2 7-1.1.3.1.r 8-1.1.3.1 10-1 10-1.3.1 11-1.2 12-1.2.2 13-1.2.3.1.1 14-1.2.3.1.2 17-1.2.4 18-1.2.4.1.r 19-1.2.4.1.2 20-1.2.4.1.1.1.1 21-1.2.4.1.1.1.1 22-1.2.4.1 24-1.3.1.1 24-1.3.1.2 24-1.3.1.3 24-1.3.1.4 24-1.3.1.5 26-1.3.1.1.1 (a / and~e.10 :op1 (g2 / great~e.3 :domain~e.2 (a2 / activate-01~e.1 :ARG1 (k / kinase~e.0)) :degree~e.3 (m / most~e.3) :time (a3 / after~e.4 :op1~e.7 (t / treat-03~e.8) :quant~e.4 (u / up-to~e.4 :op1 (t2 / temporal-quantity :quant~e.4 4~e.5 :unit (h / hour~e.6))))) :op2 (r / remain-01~e.11 :ARG1 a2 :ARG3 (e2 / elevate-01~e.12 :ARG1 a2) :time (a4 / after~e.4 :quant (t3 / temporal-quantity :quant 48~e.13 :unit (h2 / hour~e.14))) :ARG1-of (c / correspond-02~e.17 :ARG2~e.18 (e3 / express-03~e.22 :ARG2 (p / protein :name (n2 / name :op1 "cyclin-D1"~e.20,21)) :ARG1-of (i / increase-01~e.19)))) :ARG1-of (d / describe-01 :ARG0 (a5 / and~e.10 :op1 (f / figure~e.24 :mod "10A"~e.26) :op2 (f2 / figure~e.24 :mod "10B") :op3 (f3 / figure~e.24 :mod "10C") :op4 (f4 / figure~e.24 :mod "10D") :op5 (f5 / figure~e.24 :mod "10E")))) # ::id a_pmid_2169_9731.181 ::amr-annotator SDL-AMR-09 ::preferred # ::tok When treated with 2 ng @/@ mL EGF , a concentration 1,000 @-@ times higher than the amount of EGF in cell @-@ conditioned media and 40 @-@ times higher than what is detected in BAL fluid , Erk1 @/@ 2 activity was not significantly elevated and Akt levels were unaffected ( Figure 10C , D ) . # ::alignments 1-1.3 2-1.3.1.r 3-1.3.1.2.1 4-1.3.1.2.2 5-1.1.2.1 6-1.3.1.2.2 7-1.3.1.1.1 10-1.3.1.2 10-1.3.1.3.1 11-1.3.1.3.1.2.1.1.1 13-1.3.1.3.1.2.1.1 14-1.3.1.3.1.2 14-1.3.1.3.1.2.1 14-1.3.1.3.1.2.1.r 15-1.3.1.3.1.2.2.r 17-1.3.1.3.1.2.2.1 18-1.3.1.3.1.2.1.1 19-1.3.1.1.1 20-1.3.1.3.1.2.2.1.2.r 21-1.3.1.3.1.2.2.1.2.1.1 23-1.3.1.3.1.2.2.1.2.1 24-1.3.1.3.1.2.2.1.2 25-1.3.1.3.1.2.2 26-1.3.1.3.1.3.1.1.1 28-1.3.1.3.1.3.1.1 29-1.3.1.3.1.3 29-1.3.1.3.1.3.1 29-1.3.1.3.1.3.1.r 30-1.3.1.3.1.3.2.r 31-1.3.1.3.1.3.2 33-1.3.1.3.1.3.2.1 34-1.3.1.3.1.3.2.1.1.r 35-1.3.1.3.1.3.2.1.1.1 35-1.3.1.3.1.3.2.1.1.1.1 35-1.3.1.3.1.3.2.1.1.1.1.1 35-1.3.1.3.1.3.2.1.1.1.1.1.r 35-1.3.1.3.1.3.2.1.1.1.1.r 36-1.3.1.3.1.3.2.1.1 38-1.1.2.1.1.1.1 39-1.1.2.1 40-1.3.1.2.1 41-1.1.2 43-1.1.1 43-1.1.1.r 44-1.1.3 45-1.1 46-1 46-1.4.1 47-1.2.2.1.1.1 48-1.2.2 50-1.2 50-1.2.1 50-1.2.1.r 52-1.4.1.1 52-1.4.1.2 54-1.4.1.1.1 (a2 / and~e.46 :op1 (e / elevate-01~e.45 :polarity~e.43 -~e.43 :ARG1 (a / activity-06~e.41 :ARG0 (s / slash~e.5,39 :op1 (e2 / enzyme :name (n / name :op1 "Erk1"~e.38)) :op2 (e3 / enzyme :name (n2 / name :op1 "Erk2")))) :ARG1-of (s2 / significant-02~e.44)) :op2 (a3 / affect-01~e.50 :polarity~e.50 -~e.50 :ARG1 (l / level~e.48 :quant-of (e4 / enzyme :name (n3 / name :op1 "Akt"~e.47)))) :condition (t / treat-04~e.1 :ARG2~e.2 (p / protein :name (n4 / name :op1 "EGF"~e.7,19) :quant (c / concentration-quantity~e.10 :quant 2~e.3,40 :unit (n6 / nanogram-per-milliliter~e.4,6)) :ARG1-of (e5 / equal-01 :ARG2 (c2 / concentrate-02~e.10 :ARG1 p :ARG1-of (h / high-02~e.14 :degree~e.14 (m2 / more~e.14 :quant (p2 / product-of~e.13,18 :op1 1000~e.11)) :compared-to~e.15 (a4 / and~e.25 :op1 (a5 / amount~e.17 :quant-of p :location~e.20 (m3 / media~e.24 :ARG1-of (c3 / condition-01~e.23 :ARG2 (c4 / cell~e.21)))))) :ARG1-of (h2 / high-02~e.29 :degree~e.29 (m4 / more~e.29 :quant (p3 / product-of~e.28 :op1 40~e.26)) :compared-to~e.30 (t2 / thing~e.31 :ARG1-of (d2 / detect-01~e.33 :location~e.34 (f / fluid~e.36 :mod (l2 / lavage~e.35 :mod~e.35 (a7 / alveolus~e.35 :mod~e.35 (b / bronchus~e.35))))))))))) :ARG1-of (d / describe-01 :ARG0 (a6 / and~e.46 :op1 (f2 / figure~e.52 :mod "10C"~e.54) :op2 (f3 / figure~e.52 :mod "10D")))) # ::id a_pmid_2169_9731.182 ::amr-annotator SDL-AMR-09 ::preferred # ::tok EGF may partially stimulate Erk1 @/@ 2 activity at supra @-@ physiological levels , but this was not sufficient to stimulate cellular growth . # ::alignments 0-1.1.1.1.1.1 1-1.1 2-1.1.1.3 2-1.1.1.3.r 2-1.1.1.4.1.1.r 3-1.1.1 4-1.1.1.2.1.1.1.1 5-1.1.1.2.1 7-1.1.1.2 8-1.1.1.4.r 9-1.1.1.4.1.1 11-1.1.1.4.1 12-1.1.1.4 14-1 15-1.2.2 17-1.2.1 17-1.2.1.r 18-1.2 20-1.2.3 21-1.2.3.2.1 22-1.2.3.2 (c / contrast-01~e.14 :ARG1 (p / possible-01~e.1 :ARG1 (s / stimulate-01~e.3 :ARG0 (p2 / protein :name (n / name :op1 "EGF"~e.0)) :ARG1 (a / activity-06~e.7 :ARG0 (s2 / slash~e.5 :op1 (e / enzyme :name (n2 / name :op1 "Erk1"~e.4)) :op2 (e2 / enzyme :name (n3 / name :op1 "Erk2")))) :degree~e.2 (p3 / part~e.2) :location~e.8 (l / level~e.12 :mod (p4 / physiological~e.11 :degree~e.2 (s3 / supra~e.9))))) :ARG2 (s4 / suffice-01~e.18 :polarity~e.17 -~e.17 :ARG0 (t / this~e.15) :ARG1 (s5 / stimulate-01~e.20 :ARG0 t :ARG1 (g / grow-01~e.22 :ARG1 (c2 / cell~e.21))))) # ::id a_pmid_2169_9731.183 ::amr-annotator SDL-AMR-09 ::preferred # ::tok When administered at cell @-@ and tissue @-@ relevant levels , IGF @-@ 1 stimulated both Erk1 @/@ 2 and Akt activation , elevated cellular cyclin D1 content , and induced neoplastic proliferation . # ::alignments 1-1.4 3-1.2.2.1.2 5-1.4.2 6-1.4.2.2.1 8-1.4.2.1.2 9-1.4.2.1 9-1.4.2.2 11-1.1.1.1.1 13-1.1.1.1.1 14-1.1 16-1.1.2.1.1.1.1.1 17-1.1.2.1.1 19-1.1.2 20-1.1.2.2.1.1.1 21-1.1.2.1 21-1.1.2.2 23-1.2 24-1.2.2.1.2 25-1.2.2.1.1.1 26-1.2.2.1.1.1 27-1.2.2 29-1 30-1.3 31-1.3.2.1 32-1.3.2 (a4 / and~e.29 :op1 (s / stimulate-01~e.14 :ARG0 (p / protein :name (n / name :op1 "IGF-1"~e.11,13)) :ARG1 (a / and~e.19 :op1 (a2 / activate-01~e.21 :ARG1 (s2 / slash~e.17 :op1 (e / enzyme :name (n2 / name :op1 "Erk1"~e.16)) :op2 (e2 / enzyme :name (n3 / name :op1 "Erk2")))) :op2 (a3 / activate-01~e.21 :ARG1 (e3 / enzyme :name (n4 / name :op1 "Akt"~e.20))))) :op2 (e4 / elevate-01~e.23 :ARG0 p :ARG1 (c / content~e.27 :quant-of (p2 / protein :name (n5 / name :op1 "cyclin-D1"~e.25,26) :source (c2 / cell~e.3,24)))) :op3 (i / induce-01~e.30 :ARG0 p :ARG2 (p3 / proliferate-01~e.32 :ARG0 (n6 / neoplasm~e.31))) :condition (a5 / administer-01~e.1 :ARG1 p :location (a6 / and~e.5 :op1 (l / level~e.9 :mod c2 :ARG1-of (r / relevant-01~e.8)) :op2 (l2 / level~e.9 :mod (t / tissue~e.6) :ARG1-of r)))) # ::id a_pmid_2194_3101.13 ::amr-annotator SDL-AMR-09 ::preferred # ::tok Evidence presented here indicate that BRAF @ V600E @ significantly induces cell migration and invasion properties in vitro @ in colon cancer cells , at least in part through activation of RhoA GTPase . # ::alignments 0-1.1 1-1.1.1 2-1.1.1.1 3-1 4-1.2.r 5-1.2.1.1.1 7-1.2.1.2.1 9-1.2.3 10-1.2 11-1.2.2.4 12-1.2.2.1.1 13-1.2.2 14-1.2.2.2.1 15-1.2.2.1 15-1.2.2.2 17-1.2.2.3 18-1.2.2.3 20-1.2.2.3 20-1.2.2.4.1.r 21-1.2.2.4.1.3 22-1.2.2.4.1.2.1 23-1.2.2.1.1.1 25-1.2.4.3 26-1.2.4.3 27-1.2.4.3.1 27-1.2.4.3.r 28-1.2.4.3.r 30-1.2.4 31-1.2.4.2.r 32-1.2.4.2.1.1 33-1.2.4.2.1.2 (i / indicate-01~e.3 :ARG0 (e / evidence~e.0 :ARG1-of (p / present-01~e.1 :medium (h / here~e.2))) :ARG1~e.4 (i2 / induce-01~e.10 :ARG0 (e3 / enzyme :name (n / name :op1 "BRAF"~e.5) :ARG1-of (m / mutate-01 :value "V600E"~e.7)) :ARG2 (a / and~e.13 :op1 (p2 / property~e.15 :mod (m2 / migrate-01~e.12 :ARG0 (c / cell~e.23))) :op2 (p3 / property~e.15 :mod (i3 / invade-01~e.14 :ARG0 c)) :manner (i4 / in-vitro~e.17,18,20) :location (c2 / cell~e.11 :mod~e.20 (d / disease :wiki "Cancer" :name (n2 / name :op1 "cancer"~e.22) :mod (c3 / colon~e.21)))) :ARG1-of (s / significant-02~e.9) :manner (a2 / activate-01~e.30 :ARG0 e3 :ARG1~e.31 (p5 / pathway :name (n3 / name :op1 "RhoA"~e.32 :op2 "GTPase"~e.33)) :degree~e.27,28 (a3 / at-least~e.25,26 :op1 (p4 / part~e.27))))) # ::id a_pmid_2194_3101.14 ::amr-annotator SDL-AMR-09 ::preferred # ::tok The relationship established between BRAF @ V600E @ and RhoA activation is mediated by the MEK @-@ ERK pathway . # ::alignments 1-1.2 2-1.2.3 4-1.2.1.1.1.1 6-1.2.1.1.2.1 9-1.2.2.1.1.1 10-1.2.1 10-1.2.2 12-1 13-1.1.r 15-1.1.1.1 17-1.1.1.1 18-1.1 (m / mediate-01~e.12 :ARG0~e.13 (p / pathway~e.18 :name (n / name :op1 "MEK-ERK"~e.15,17)) :ARG1 (r / relation-03~e.1 :ARG0 (a / activate-01~e.10 :ARG1 (e2 / enzyme :name (n2 / name :op1 "BRAF"~e.4) :ARG1-of (m2 / mutate-01 :value "V600E"~e.6))) :ARG2 (a2 / activate-01~e.10 :ARG1 (p2 / protein :name (n3 / name :op1 "RhoA"~e.9))) :ARG1-of (e / establish-01~e.2))) # ::id a_pmid_2194_3101.15 ::amr-annotator SDL-AMR-09 ::preferred # ::tok In parallel , KRAS @ G12V @ enhances the ability of colon adenocarcinoma cells Caco @-@ 2 to migrate and invade through filopodia formation and PI3K @-@ dependent Cdc42 activation . # ::alignments 1-1.3 3-1.1.1.1 5-1.1.2.1 7-1 9-1.2 10-1.2.1.r 11-1.2.1.2.1 12-1.2.1.2 13-1.2.1 14-1.2.1.1.1 16-1.2.1.1.1 17-1.2.2.r 18-1.2.2.1 19-1.2.2 20-1.2.2.2 22-1.2.2.3.1.2 23-1.2.2.3.1 24-1.2.2.3 25-1.2.2.3.2.3.1.1.1 27-1.2.2.3.2.3 28-1.2.2.3.2.2.1.1 29-1.2.2.3.2 (e / enhance-01~e.7 :ARG0 (e2 / enzyme :name (n / name :op1 "KRAS"~e.3) :ARG1-of (m / mutate-01 :value "G12V"~e.5)) :ARG1 (c / capable-01~e.9 :ARG1~e.10 (c2 / cell-line~e.13 :name (n2 / name :op1 "Caco-2"~e.14,16) :source (a / adenocarcinoma~e.12 :part-of (c3 / colon~e.11))) :ARG2~e.17 (a2 / and~e.19 :op1 (m2 / migrate-01~e.18 :ARG0 c2) :op2 (i / invade-01~e.20 :ARG0 c2) :manner (a3 / and~e.24 :op1 (f / form-01~e.23 :ARG0 c2 :ARG1 (f2 / filopodia~e.22)) :op2 (a4 / activate-01~e.29 :ARG0 c2 :ARG1 (p2 / protein :name (n3 / name :op1 "Cdc42"~e.28)) :ARG0-of (d / depend-01~e.27 :ARG1 (e3 / enzyme :name (n4 / name :op1 "PI3K"~e.25))))))) :mod (p / parallel~e.1)) # ::id a_pmid_2194_3101.16 ::amr-annotator SDL-AMR-09 ::preferred # ::tok Ultimately increased cell migration and invasion , mediated by Rac1 , along with the mesenchymal morphology obtained through the Epithelial @-@ Mesenchymal Transition ( EMT ) were the main characteristics rendered by HRAS @ G12V @ in Caco @-@ 2 cells . # ::alignments 0-1.1.5.1 0-1.1.5.1.r 1-1.1.5 2-1.1.1.1 3-1.1.1 4-1.1 5-1.1.2 7-1.1.4 8-1.1.4.1.r 9-1.1.4.1.1.1 14-1.1.3.1 15-1.1.3 16-1.1.3.2 19-1.1.3.2.1.2 21-1.1.3.2.1.1 22-1.1.3.2.1 24-1.1.3.2.1 28-1.2 29-1 30-1.3 31-1.3.1.r 32-1.3.1.1.1 34-1.3.1.2.1 36-1.3.2.r 37-1.3.2.1.1 39-1.3.2.1.1 40-1.3.2 (c / characteristic-02~e.29 :ARG2 (a / and~e.4 :op1 (m4 / migrate-01~e.3 :ARG0 (c3 / cell~e.2)) :op2 (i2 / invade-01~e.5 :ARG0 c3) :op3 (m5 / morphology~e.15 :mod (m6 / mesenchyme~e.14) :ARG1-of (o / obtain-01~e.16 :ARG2 (t / transition-01~e.22,24 :ARG2 (m7 / mesenchyme~e.21) :ARG3 (e / epithelium~e.19)))) :ARG1-of (m3 / mediate-01~e.7 :ARG0~e.8 (p / protein :name (n3 / name :op1 "Rac1"~e.9))) :ARG1-of (i / increase-01~e.1 :manner~e.0 (u / ultimate~e.0))) :mod (m2 / main~e.28) :ARG1-of (r / render-01~e.30 :ARG0~e.31 (g / gene :name (n / name :op1 "HRAS"~e.32) :ARG1-of (m / mutate-01 :value "G12V"~e.34)) :location~e.36 (c2 / cell-line~e.40 :name (n2 / name :op1 "Caco-2"~e.37,39)))) # ::id a_pmid_2194_3101.17 ::amr-annotator SDL-AMR-09 ::preferred # ::tok Moreover , BRAF and KRAS oncogenes are shown to cooperate with the TGFβ-1 pathway to provide cells with additional transforming properties . # ::alignments 0-1.1.1.1 2-1.1.1.1.1.1.1 3-1.1.1.1 4-1.1.1.1.2.1.1 7-1.1 8-1.1.1.1.3 9-1.1.1 10-1.1.1.2.r 12-1.1.1.2.1.1 13-1.1.1.2 14-1.1.1.1.3 15-1.1.1.3 16-1.1.1.3.3 17-1.1.1.3.2.r 18-1.1.1.3.2.2 19-1.1.1.3.2.1 20-1.1.1.3.2 (a / and :op2 (s / show-01~e.7 :ARG1 (c / cooperate-01~e.9 :ARG0 (a2 / and~e.0,3 :op1 (g / gene :name (n / name :op1 "BRAF"~e.2)) :op2 (g2 / gene :name (n2 / name :op1 "KRAS"~e.4)) :ARG0-of (c3 / cause-01~e.8,14 :ARG1 (d / disease :wiki "Cancer" :name (n4 / name :op1 "cancer")))) :ARG1~e.10 (p2 / pathway~e.13 :name (n3 / name :op1 "TGFβ-1"~e.12)) :ARG2 (p / provide-01~e.15 :ARG0 (a4 / and :op1 a2 :op2 p2) :ARG1~e.17 (p3 / property~e.20 :mod (t / transform-01~e.19) :mod (a3 / additional~e.18)) :ARG2 (c2 / cell~e.16))))) # ::id a_pmid_2194_3101.131 ::amr-annotator SDL-AMR-09 ::preferred # ::tok BRAF @ V600E @ induces distinct morphological changes in colon adenocarcinoma cells as compared to KRAS @ G12V @ and loss of their epithelial architecture in 3D culture # ::alignments 0-1.1.1.1 2-1.1.2.1 4-1 5-1.2.1.4 6-1.2.1.3 7-1.2.1 8-1.2.1.2.r 9-1.2.1.2.1.1 10-1.2.1.2.1 11-1.2.1.2 13-1.2.1.5.r 15-1.2.1.5.1.1 17-1.2.1.5.2.1 19-1.2 20-1.2.2 21-1.2.2.1.r 22-1.2.2.1 23-1.2.2.2.1 24-1.2.2.2 25-1.2.2.2.2.r 26-1.2.2.2.2.1 26-1.2.2.2.2.1.1 26-1.2.2.2.2.1.1.r 27-1.2.2.2.2 (i / induce-01~e.4 :ARG0 (e2 / enzyme :name (n / name :op1 "BRAF"~e.0) :ARG1-of (m / mutate-01 :value "V600E"~e.2)) :ARG2 (a4 / and~e.19 :op1 (c / change-01~e.7 :ARG0 e2 :ARG1~e.8 (c2 / cell~e.11 :source (a / adenocarcinoma~e.10 :mod (c3 / colon~e.9))) :mod (m2 / morphology~e.6) :mod (d / distinct~e.5) :compared-to~e.13 (e3 / enzyme :name (n2 / name :op1 "KRAS"~e.15) :ARG1-of (m3 / mutate-01 :value "G12V"~e.17))) :op2 (l / lose-02~e.20 :ARG0~e.21 c2~e.22 :ARG1 (a3 / architecture~e.24 :mod (e / epithelium~e.23) :location~e.25 (c4 / culture~e.27 :mod (d2 / dimension~e.26 :quant~e.26 3~e.26)))))) # ::id a_pmid_2194_3101.132 ::amr-annotator SDL-AMR-09 ::preferred # ::tok Previously established Caco @-@ BR cells have adopted a substantially different morphology when compared to the parental Caco @-@ 2 cells [ @ 21 @ ] . # ::alignments 0-1.1.2.1 1-1.1.2 2-1.1.1.1 4-1.1.1.1 5-1.1 7-1 9-1.2.1.1 10-1.2.1 11-1.2 12-1.4.r 13-1.4 14-1.4.2.r 16-1.4.2.2 17-1.4.2.1.1 19-1.4.2.1.1 20-1.4.2 23-1.3.1.1.1 (a / adopt-01~e.7 :ARG0 (c2 / cell-line~e.5 :name (n / name :op1 "Caco-BR"~e.2,4) :ARG1-of (e / establish-01~e.1 :time (p2 / previous~e.0))) :ARG1 (m / morphology~e.11 :ARG1-of (d2 / differ-02~e.10 :degree (s / substantial~e.9))) :ARG1-of (d / describe-01 :ARG0 (p / publication :ARG1-of (c / cite-01 :ARG2 21~e.23))) :time~e.12 (c3 / compare-01~e.13 :ARG1 c2 :ARG2~e.14 (c4 / cell-line~e.20 :name (n2 / name :op1 "Caco-2"~e.17,19) :mod (p3 / parent~e.16)))) # ::id a_pmid_2194_3101.133 ::amr-annotator SDL-AMR-09 ::preferred # ::tok The elongated morphology acquired by Caco @-@ BR cells was characterized by long membrane protrusions ( Figure 1A , Additional Figure 1 ) . # ::alignments 1-1.1.2 2-1.1 3-1.1.1 4-1.1.1.1.r 5-1.1.1.1.1.1 7-1.1.1.1.1.1 8-1.1.1.1 10-1 11-1.2.r 12-1.2.2 13-1.2.1 14-1.2 16-1.3.1.1 18-1.3.1.1.1 21-1.3.1.2.2 22-1.3.1.2 24-1.3.1.2.1 (c / characterize-01~e.10 :ARG1 (m / morphology~e.2 :ARG1-of (a3 / acquire-01~e.3 :ARG0~e.4 (c2 / cell-line~e.8 :name (n / name :op1 "Caco-BR"~e.5,7))) :ARG1-of (e / elongate-01~e.1)) :ARG2~e.11 (p / protrude-01~e.14 :ARG1 (m2 / membrane~e.13) :ARG1-of (l / long-03~e.12)) :ARG1-of (d / describe-01 :ARG0 (a / and :op1 (f / figure~e.16 :mod "1A"~e.18) :op2 (f2 / figure~e.22 :mod 1~e.24 :mod (a2 / additional~e.21))))) # ::id a_pmid_2194_3101.134 ::amr-annotator SDL-AMR-09 ::preferred # ::tok We present evidence that the morphology of Caco @-@ BR13 cells show properties of both Caco @-@ 2 epithelial nature and of the mesenchymal phenotype of Caco @-@ H2 cells . # ::alignments 0-1.1 1-1 2-1.2 3-1.2.1.r 5-1.2.1.1 7-1.2.1.1.1.1.1 9-1.2.1.1.1.1.1 10-1.2.1.1.1 10-1.2.1.2.1.1.2 11-1.2.1 12-1.2.1.2.1 12-1.2.1.2.2 15-1.2.1.2.1.1.2.1.1 17-1.2.1.2.1.1.2.1.1 18-1.2.1.2.1.1.1 19-1.2.1.2.1.1 20-1.2.1.2 21-1.2.1.2.2.1.r 23-1.2.1.2.2.1.1 24-1.2.1.2.2.1 25-1.2.1.2.2.1.2.r 26-1.2.1.2.2.1.2.1.1 28-1.2.1.2.2.1.2.1.1 29-1.2.1.2.2.1.2 (p / present-01~e.1 :ARG0 (w / we~e.0) :ARG1 (e / evidence-01~e.2 :ARG1~e.3 (s / show-01~e.11 :ARG0 (m / morphology~e.5 :poss (c / cell-line~e.10 :name (n / name :op1 "Caco-BR13"~e.7,9))) :ARG1 (a / and~e.20 :op1 (p2 / property~e.12 :poss (n2 / nature~e.19 :mod (e2 / epithelium~e.18) :poss (c2 / cell-line~e.10 :name (n3 / name :op1 "Caco-2"~e.15,17)))) :op2 (p3 / property~e.12 :poss~e.21 (p4 / phenotype~e.24 :mod (m2 / mesenchyme~e.23) :poss~e.25 (c3 / cell-line~e.29 :name (n4 / name :op1 "Caco-H2"~e.26,28)))))))) # ::id a_pmid_2194_3101.135 ::amr-annotator SDL-AMR-09 ::preferred # ::tok On the other hand , Caco @-@ K15 cells , which overexpress KRAS @ G12V , have retained the overall parental morphology of Caco @-@ 2 cells . # ::alignments 5-1.1.1.1.1 7-1.1.1.1.1 8-1.1.1 11-1.1.1.2 12-1.1.1.2.1.1.1 14-1.1.1.2.1.2.1 18-1.1 20-1.1.2.1 21-1.1.2.2 22-1.1.2 23-1.1.2.3.r 24-1.1.2.3.1.1 26-1.1.2.3.1.1 27-1.1.2.3 (c3 / contrast-01 :ARG2 (r2 / retain-01~e.18 :ARG0 (c / cell-line~e.8 :name (n / name :op1 "Caco-K15"~e.5,7) :location-of (o / overexpress-01~e.11 :ARG1 (g / gene :name (n2 / name :op1 "KRAS"~e.12) :ARG1-of (m / mutate-01 :value "G12V"~e.14)))) :ARG1 (m2 / morphology~e.22 :mod (o2 / overall~e.20) :mod (p / parent~e.21) :poss~e.23 (c2 / cell-line~e.27 :name (n3 / name :op1 "Caco-2"~e.24,26))))) # ::id a_pmid_2194_3101.136 ::amr-annotator SDL-AMR-09 ::preferred # ::tok For comparison , established adenocarcinoma cell lines HT29 and DLD @-@ 1 , bearing mutant BRAF @ V600E @ @ and KRAS @ G13D @ @ respectively , have also been analyzed in the present study . # ::alignments 1-1.2 3-1.1.4 4-1.1.3 5-1.1.1 6-1.1.1 6-1.1.2 7-1.1.1.1.1 8-1.1 9-1.1.2.1.1 11-1.1.2.1.1 13-1.1.1.2 13-1.1.2.2 14-1.1.1.2.1 14-1.1.1.2.1.2 14-1.1.1.2.1.2.r 14-1.1.2.2.1 14-1.1.2.2.1.2 14-1.1.2.2.1.2.r 16-1.1.1.2.1.1.1 18-1.1.1.2.1.2.1 21-1.1 23-1.1.2.2.1.1.1 25-1.1.2.2.1.2.1 28-1.1.5 31-1.4 33-1 34-1.3.r 36-1.3.1 37-1.3 (a / analyze-01~e.33 :ARG1 (a3 / and~e.8,21 :op1 (c2 / cell-line~e.5,6 :name (n / name :op1 "HT29"~e.7) :ARG0-of (b / bear-01~e.13 :ARG1 (g / gene~e.14 :name (n3 / name :op1 "BRAF"~e.16) :ARG1-of~e.14 (m / mutate-01~e.14 :value "V600E"~e.18)))) :op2 (c3 / cell-line~e.6 :name (n2 / name :op1 "DLD-1"~e.9,11) :ARG0-of (b2 / bear-01~e.13 :ARG1 (g2 / gene~e.14 :name (n4 / name :op1 "KRAS"~e.23) :ARG1-of~e.14 (m2 / mutate-01~e.14 :value "G13D"~e.25)))) :mod (a4 / adenocarcinoma~e.4) :ARG1-of (e / establish-01~e.3) :mod (r / respective~e.28)) :purpose (c / compare-01~e.1) :medium~e.34 (s / study-01~e.37 :time (p / present~e.36)) :mod (a2 / also~e.31)) # ::id a_pmid_2194_3101.137 ::amr-annotator SDL-AMR-09 ::preferred # ::tok It is of interest that the phenotype of Caco @-@ BR cells resembles that of DLD @-@ 1 cells ( KRAS @ G13D ) , especially since both of these cell types share high levels of p @-@ BRAF ( later analyzed ) . # ::alignments 3-1 6-1.1.1 6-1.1.2 7-1.1.1.1.r 8-1.1.1.1.1.1 10-1.1.1.1.1.1 11-1.1.1.1 11-1.1.2.1 12-1.1 15-1.1.2.1.1.1 17-1.1.2.1.1.1 18-1.1.1.1 20-1.1.2.1.2.1.1 22-1.1.2.1.2.2.1 26-1.2.2 27-1.2 31-1.1.1.1 33-1.2.1 34-1.2.1.2.1 35-1.2.1.2 36-1.2.1.2.2.r 37-1.2.1.2.2.2 39-1.2.1.2.2.1.1 41-1.2.1.2.2.3.1 41-1.2.1.2.2.3.1.1 41-1.2.1.2.2.3.1.1.r 42-1.2.1.2.2.3 (i / interest-01~e.3 :ARG0 (r / resemble-01~e.12 :ARG1 (p / phenotype~e.6 :poss~e.7 (c2 / cell-line~e.11,18,31 :name (n / name :op1 "Caco-BR"~e.8,10))) :ARG2 (p2 / phenotype~e.6 :poss (c3 / cell-line~e.11 :name (n2 / name :op1 "DLD-1"~e.15,17) :mod (e3 / enzyme :name (n3 / name :op1 "KRAS"~e.20) :ARG1-of (m2 / mutate-01 :value "G13D"~e.22))))) :ARG1-of (c / cause-01~e.27 :ARG0 (s / share-01~e.33 :ARG0 (a / and :op1 c2 :op2 c3) :ARG1 (l / level~e.35 :ARG1-of (h / high-02~e.34) :quant-of~e.36 (e2 / enzyme :name (n4 / name :op1 "BRAF"~e.39) :ARG3-of (p4 / phosphorylate-01~e.37) :ARG1-of (a2 / analyze-01~e.42 :time (l2 / late~e.41 :degree~e.41 (m3 / more~e.41)))))) :mod (e / especially~e.26))) # ::id a_pmid_2194_3101.138 ::amr-annotator SDL-AMR-09 ::preferred # ::tok Our previous study shows important similarities between Caco @-@ BR and DLD @-@ 1 cells regarding their tumourigenic properties and signaling pathways , suggesting that their transformation process occurs mainly through the constitutive activation of the MAPK ( Mitogen @-@ Activated Protein Kinase ) pathway [ @ 21 @ ] . # ::alignments 0-1.1.1 0-1.1.1.r 1-1.1.2 2-1.1 3-1 4-1.2.4 5-1.2 7-1.2.1.1.1 9-1.2.1.1.1 11-1.2.2.1.1 13-1.2.2.1.1 14-1.2.1 14-1.2.2 15-1.2.3.r 17-1.2.3.1 17-1.2.3.1.1 17-1.2.3.1.1.1 17-1.2.3.1.1.1.r 17-1.2.3.1.1.r 19-1.2.3 20-1.2.3.2.1 21-1.2.3.2 23-1.3 26-1.3.1.1 27-1.3.1 32-1.3.2.2 33-1.3.2 34-1.3.2.1.r 36-1.3.2.1.1.1 38-1.3.2.1.2.1.1.1 40-1.3.2.1.2.1.1.1 41-1.3.2.1.2.1.1.2 42-1.3.2.1.2.1.1.3 44-1.3.2.1 44-1.3.2.1.2.1 47-1.4.1.1.1 (s / show-01~e.3 :ARG0 (s2 / study-01~e.2 :ARG0~e.0 (w / we~e.0) :time (p / previous~e.1)) :ARG1 (r / resemble-01~e.5 :ARG1 (c / cell-line~e.14 :name (n / name :op1 "Caco-BR"~e.7,9)) :ARG2 (c2 / cell-line~e.14 :name (n2 / name :op1 "DLD-1"~e.11,13)) :topic~e.15 (a / and~e.19 :op1 (p2 / possible-01~e.17 :ARG1~e.17 (c5 / create-01~e.17 :ARG1~e.17 (t / tumor~e.17))) :op2 (p3 / pathway~e.21 :ARG0-of (s3 / signal-07~e.20))) :mod (i / important~e.4)) :ARG0-of (s4 / suggest-01~e.23 :ARG1 (p4 / process-02~e.27 :ARG1 (t2 / transform-01~e.26 :ARG1 (a3 / and :op1 c :op2 c2))) :manner (a4 / activate-01~e.33 :ARG1~e.34 (p5 / pathway~e.44 :name (n3 / name :op1 "MAPK"~e.36) :ARG1-of (m / mean-01 :ARG2 (p6 / pathway~e.44 :name (n4 / name :op1 "Mitogen-Activated"~e.38,40 :op2 "Protein"~e.41 :op3 "Kinase"~e.42)))) :mod (c3 / constitutive~e.32))) :ARG1-of (d / describe-01 :ARG0 (p7 / publication :ARG1-of (c4 / cite-01 :ARG2 21~e.47)))) # ::id a_pmid_2194_3101.139 ::amr-annotator SDL-AMR-09 ::preferred # ::tok Staining with phalloidin resolved the morphological differences within the cell line panel indicating major actin cytoskeleton changes ( Figure 1A ) . # ::alignments 0-1.1 1-1.1.1.r 2-1.1.1.1.1 3-1 5-1.2.2 6-1.2 9-1.2.1.1 10-1.2.1.1 11-1.2.1 12-1.2.1.2 13-1.2.1.2.1.2 14-1.2.1.2.1.1.1.1.1 15-1.2.1.2.1.1 16-1.2.1.2.1 18-1.3.1 20-1.3.1.1 (r / resolve-01~e.3 :ARG0 (s / stain-01~e.0 :ARG2~e.1 (s2 / small-molecule :name (n / name :op1 "phalloidin"~e.2))) :ARG1 (d2 / differ-02~e.6 :ARG1 (p / panel~e.11 :mod (c / cell-line~e.9,10) :ARG0-of (i / indicate-01~e.12 :ARG1 (c2 / change-01~e.16 :ARG1 (c3 / cytoskeleton~e.15 :mod (p2 / protein :name (n2 / name :op1 "actin"~e.14))) :ARG1-of (m3 / major-02~e.13)))) :mod (m2 / morphology~e.5)) :ARG1-of (d / describe-01 :ARG0 (f / figure~e.18 :mod "1A"~e.20))) # ::id a_pmid_2194_3101.140 ::amr-annotator SDL-AMR-09 ::preferred # ::tok More specifically , in Caco @-@ BR13 cells the formation of stress fibers was enhanced , whereas formation of filopodia @-@ membrane protrusions enriched with actin @- is evident in Caco @-@ K15 cells ( Figure 1A @-@ ii , arrow ) . # ::alignments 0-1.4.1 1-1.4 4-1.1.2.1.1 6-1.1.2.1.1 7-1.1.2 9-1.1.1 10-1.1.1.1.r 11-1.1.1.1.1 12-1.1.1.1 14-1.1 16-1 17-1.2.1 18-1.2.1.1.r 19-1.2.1.1.1.1 21-1.2.1.1.1 22-1.2.1.1 23-1.2.1.1.2 24-1.2.1.1.2.1.r 25-1.2.1.1.2.1.1.1 27-1.2.1.r 28-1.2 29-1.2.2.r 30-1.2.2.1.1 32-1.2.2.1.1 33-1.2.2 35-1.3.1.1 42-1.3.1.2 (c / contrast-01~e.16 :ARG1 (e / enhance-01~e.14 :ARG1 (f4 / form-01~e.9 :ARG1~e.10 (f5 / fiber~e.12 :mod (s2 / stress~e.11))) :location (c3 / cell-line~e.7 :name (n3 / name :op1 "Caco-BR13"~e.4,6))) :ARG2 (e2 / evident~e.28 :domain~e.27 (f / form-01~e.17 :ARG1~e.18 (p / protrusion~e.22 :mod (m2 / membrane~e.21 :mod (f2 / filopodia~e.19)) :ARG1-of (e3 / enrich-01~e.23 :ARG2~e.24 (p2 / protein :name (n / name :op1 "actin"~e.25))))) :location~e.29 (c2 / cell-line~e.33 :name (n2 / name :op1 "Caco-K15"~e.30,32))) :ARG1-of (d / describe-01 :ARG0 (a / and :op1 (f3 / figure~e.35 :mod "1A.ii") :op2 (a2 / arrow~e.42))) :ARG1-of (s / specific-02~e.1 :degree (m / more~e.0))) # ::id a_pmid_2194_3101.141 ::amr-annotator SDL-AMR-09 ::preferred # ::tok In order to study in depth the morphology and architecture of the different cell lines under conditions that resemble the real tissue microenvironment , the three @-@ dimensional ( 3D ) culture system was adopted . # ::alignments 0-1.2.r 1-1.2.r 2-1.2.r 3-1.2 4-1.2.2.r 5-1.2.2 7-1.2.1.1 8-1.2.1 9-1.2.1.2 10-1.2.1.1.1.r 12-1.2.1.1.1.1 13-1.2.1.1.1 14-1.2.1.1.1 16-1.1.2 16-1.1.2.r 18-1.1.2.1 20-1.1.2.1.1.1.1 21-1.1.2.1.1.1 22-1.1.2.1.1 25-1.1.1.1.1 27-1.1.1.1 29-1.1.1.1 29-1.1.1.1.1 29-1.1.1.1.1.r 31-1.1.1 32-1.1 34-1 (a / adopt-01~e.34 :ARG1 (s / system~e.32 :mod (c / culture~e.31 :mod (d3 / dimension~e.27,29 :quant~e.29 3~e.25,29)) :condition~e.16 (c4 / condition~e.16 :ARG1-of (r2 / resemble-01~e.18 :ARG2 (m3 / microenvironment~e.22 :mod (t2 / tissue~e.21 :ARG1-of (r / real-04~e.20)))))) :purpose~e.0,1,2 (s2 / study-01~e.3 :ARG1 (a2 / and~e.8 :op1 (m2 / morphology~e.7 :poss~e.10 (c2 / cell-line~e.13,14 :ARG1-of (d2 / differ-02~e.12))) :op2 (a3 / architecture~e.9 :poss c2)) :ARG1-of~e.4 (d / deep-03~e.5))) # ::id a_pmid_2194_3101.142 ::amr-annotator SDL-AMR-09 ::preferred # ::tok As also previously shown [ @ 22 @ ] , Caco @-@ 2 cells were organized into cyst @-@ like structures that resemble normal colon cell architecture following their growth in Matrigel for about 12 days ( Figure 1B ) . # ::alignments 0-1.2.r 1-1.3 2-1.2 3-1 6-1.4.1.1.1 10-1.1.1.1.1 12-1.1.1.1.1 13-1.1.1 15-1.1 16-1.1.2.r 17-1.1.2.1.1 19-1.1.2.1 19-1.1.2.2 20-1.1.2 22-1.1.2.2 23-1.1.2.2.1.1.1.1 24-1.1.2.2.1.1.1 25-1.1.2.2.1.1 26-1.1.2.2.1 27-1.1.3 28-1.1.3.1.1 28-1.1.3.1.1.r 29-1.1.3.1 30-1.1.3.1.2.r 31-1.1.3.1.2.1.1 32-1.1.3.1.3.r 33-1.1.3.1.3 34-1.1.3.1.3.1.1 35-1.1.3.1.3.1.2 37-1.1.4.1 39-1.1.4.1.1 (s / show-01~e.3 :ARG1 (o / organize-01~e.15 :ARG1 (c2 / cell-line~e.13 :name (n2 / name :op1 "Caco-2"~e.10,12)) :ARG4~e.16 (s2 / structure~e.20 :ARG1-of (r / resemble-01~e.19 :ARG2 (c3 / cyst~e.17)) :ARG1-of (r2 / resemble-01~e.19,22 :ARG2 (a3 / architecture~e.26 :mod (c4 / cell~e.25 :source (c5 / colon~e.24 :ARG1-of (n3 / normal-02~e.23)))))) :ARG1-of (f / follow-01~e.27 :ARG2 (g / grow-03~e.29 :ARG1~e.28 c2~e.28 :medium~e.30 (p3 / protein :name (n / name :op1 "Matrigel"~e.31)) :duration~e.32 (a / about~e.33 :op1 (t / temporal-quantity :quant 12~e.34 :unit (d / day~e.35))))) :ARG1-of (d3 / describe-01 :ARG0 (f2 / figure~e.37 :mod "1B"~e.39))) :time~e.0 (p / previous~e.2) :mod (a2 / also~e.1) :ARG1-of (d2 / describe-01 :ARG0 (p2 / publication :ARG1-of (c / cite-01 :ARG2 22~e.6)))) # ::id a_pmid_2194_3101.143 ::amr-annotator SDL-AMR-09 ::preferred # ::tok In contrast , Caco @-@ H cells ( EMT model ) formed invasive masses with elongated protrusions , an architecture not shared by Caco @-@ BR13 and Caco @-@ K15 cells ( Figure 1B , upper panel , black arrow ) . # ::alignments 1-1 3-1.1.1.1.1 5-1.1.1.1.1 6-1.1.1 8-1.1.1.2 11-1.1 12-1.1.2.1 13-1.1.2 14-1.1.2.2.r 15-1.1.2.2.1 16-1.1.2.2 19-1.1.2.2.2 20-1.1.2.2.2.1.1 20-1.1.2.2.2.1.1.r 21-1.1.2.2.2.1 22-1.1.2.2.2.1.2.r 23-1.1.2.2.2.1.2.1.1.1 25-1.1.2.2.2.1.2.1.1.1 26-1.1.2.2.2.1.2 27-1.1.2.2.2.1.2.1.1.1 27-1.1.2.2.2.1.2.2.1.1 29-1.1.2.2.2.1.2.2.1.1 30-1.1.2.2.2.1.2.1 30-1.1.2.2.2.1.2.2 32-1.2.1.1 34-1.2.1.1.1 37-1.2.1.2.1 38-1.2.1.2 40-1.2.1.3.1 41-1.2.1.3 (c / contrast-01~e.1 :ARG2 (f / form-01~e.11 :ARG0 (c2 / cell-line~e.6 :name (n / name :op1 "Caco-H"~e.3,5) :mod (t / transition-01~e.8 :ARG2 (m2 / mesenchyme) :ARG3 (e2 / epithelium))) :ARG1 (m / mass~e.13 :ARG0-of (i / invade-01~e.12) :ARG1-of~e.14 (p / protrude-01~e.16 :ARG1-of (e / elongate-01~e.15) :mod (a / architecture~e.19 :ARG1-of (s / share-01~e.21 :polarity~e.20 -~e.20 :ARG0~e.22 (a2 / and~e.26 :op1 (c3 / cell-line~e.30 :name (n2 / name :op1 "Caco-BR13"~e.23,25,27)) :op2 (c4 / cell-line~e.30 :name (n3 / name :op1 "Caco-K15"~e.27,29)))))))) :ARG1-of (d / describe-01 :ARG0 (a3 / and :op1 (f2 / figure~e.32 :mod "1B"~e.34) :op2 (p2 / panel~e.38 :mod (u / upper~e.37)) :op3 (a4 / arrow~e.41 :ARG1-of (b / black-04~e.40))))) # ::id a_pmid_2194_3101.144 ::amr-annotator SDL-AMR-09 ::preferred # ::tok During 3D culture conditions , normal epithelial cells are organized into spheroids presenting a characteristic centrally @-@ localized hollow lumen and distinct polarization of cells surrounding this lumen . # ::alignments 1-1.3.1 1-1.3.1.1 1-1.3.1.1.r 2-1.3 3-1.3.r 5-1.1.1 6-1.1.2 7-1.1 9-1 10-1.2.r 11-1.2 12-1.2.1 14-1.2.1.1.1.3 15-1.2.1.1.1.2 18-1.2.1.1.1.1 19-1.2.1.1.1 20-1.2.1.1 21-1.2.1.1.2.2 22-1.2.1.1.2 23-1.2.1.1.2.1.r 24-1.2.1.1.2.1 25-1.2.1.1.2.1.1 27-1.2.1.1.2.1.1.1 (o2 / organize-01~e.9 :ARG1 (c3 / cell~e.7 :ARG1-of (n / normal-02~e.5) :source (e / epithelium~e.6)) :ARG4~e.10 (s / spheroid~e.11 :ARG0-of (p / present-01~e.12 :ARG1 (a / and~e.20 :op1 (l / lumen~e.19 :ARG1-of (h / hollow-02~e.18) :location (c / center~e.15) :ARG1-of (c6 / characteristic-02~e.14)) :op2 (p2 / polarize-01~e.22 :ARG1~e.23 (c4 / cell~e.24 :ARG1-of (s2 / surround-01~e.25 :ARG2 l~e.27)) :mod (d / distinct~e.21))))) :condition~e.3 (c2 / culture~e.2 :mod (d2 / dimension~e.1 :quant~e.1 3~e.1))) # ::id a_pmid_2194_3101.145 ::amr-annotator SDL-AMR-09 ::preferred # ::tok Epithelial cancer cells do not form such structures ; instead they develop non @-@ polarized clusters with limited differentiation [ @ 23 , 24 @ ] . # ::alignments 0-1.3.1.1.2 1-1.3.1.1.1.2.1 2-1.3.1.1 4-1.2.1.1.r 5-1.3.1 6-1.3.1.2.1 7-1.3.1.2 9-1.3 11-1 12-1.2.1.1 12-1.2.1.1.r 14-1.2.1 15-1.2 16-1.2.2.r 17-1.2.2.1 18-1.2.2 21-1.4.1.1.1.1 25-1.4.1.2.1.1 (d / develop-02~e.11 :ARG0 c :ARG1 (c2 / cluster~e.15 :ARG1-of (p / polarize-01~e.14 :polarity~e.4,12 -~e.12) :mod~e.16 (d3 / differentiate-01~e.18 :ARG1-of (l / limit-01~e.17))) :ARG1-of (i / instead-of-91~e.9 :ARG2 (f / form-01~e.5 :ARG0 (c / cell~e.2 :mod (d2 / disease :wiki "Cancer" :name (n / name :op1 "cancer"~e.1)) :mod (e / epithelium~e.0)) :ARG1 (s / structure~e.7 :degree (s2 / such~e.6)))) :ARG1-of (d4 / describe-01 :ARG0 (a / and :op1 (p2 / publication :ARG1-of (c3 / cite-01 :ARG2 23~e.21)) :op2 (p3 / publication :ARG1-of (c4 / cite-01 :ARG2 24~e.25))))) # ::id a_pmid_2194_3101.146 ::amr-annotator SDL-AMR-09 ::preferred # ::tok Following staining with Hoechst and phalloidin the ability of Caco @-@ 2 cells to form spheroids with lumen was observed , a property also retained by Caco @-@ K15 cells but completely absent in Caco @-@ BR13 and Caco @-@ H2 cells ( Figure 1B , lower panel ) . # ::alignments 0-1.2 1-1.2.1 2-1.2.1.1.r 3-1.2.1.1.1.1.1 4-1.2.1.1 5-1.2.1.1.2.1.1 7-1.1 8-1.1.1.r 9-1.1.1.1.1 11-1.1.1.1.1 12-1.1.1 14-1.1.2 15-1.1.2.2 16-1.1.2.2.1.r 17-1.1.2.2.1 19-1 22-1.1.3.1 23-1.1.3.1.1.2 24-1.1.3.1.1 25-1.1.3.1.1.1.r 26-1.1.3.1.1.1.1.1 28-1.1.3.1.1.1.1.1 29-1.1.3.1.1.1 31-1.1.3.1.2.2 32-1.1.3.1.2 33-1.1.3.1.2.1.r 34-1.1.3.1.2.1.1.1.1 36-1.1.3.1.2.1.1.1.1 37-1.1.3.1.2.1 38-1.1.3.1.2.1.1.1.1 38-1.1.3.1.2.1.2.1.1 40-1.1.3.1.2.1.2.1.1 41-1.1.3.1.2.1.1 41-1.1.3.1.2.1.2 43-1.3.1.1 45-1.3.1.1.1 48-1.3.1.2.1 48-1.3.1.2.1.1 48-1.3.1.2.1.1.r 49-1.3.1.2 (o / observe-01~e.19 :ARG1 (c / capable-01~e.7 :ARG1~e.8 (c2 / cell-line~e.12 :name (n3 / name :op1 "Caco-2"~e.9,11)) :ARG2 (f3 / form-01~e.14 :ARG0 c2 :ARG1 (s2 / spheroid~e.15 :mod~e.16 (l / lumen~e.17))) :ARG1-of (m4 / mean-01 :ARG2 (p2 / property~e.22 :ARG1-of (r / retain-01~e.24 :ARG0~e.25 (c3 / cell-line~e.29 :name (n4 / name :op1 "Caco-K15"~e.26,28)) :mod (a2 / also~e.23)) :ARG1-of (a3 / absent-01~e.32 :ARG2~e.33 (a4 / and~e.37 :op1 (c4 / cell-line~e.41 :name (n5 / name :op1 "Caco-BR13"~e.34,36,38)) :op2 (c6 / cell-line~e.41 :name (n6 / name :op1 "Caco-H2"~e.38,40))) :ARG1-of (c5 / complete-02~e.31))))) :ARG2-of (f / follow-01~e.0 :ARG1 (s / stain-01~e.1 :ARG2~e.2 (a / and~e.4 :op1 (s4 / small-molecule :name (n / name :op1 "Hoechst"~e.3)) :op2 (s3 / small-molecule :name (n2 / name :op1 "phalloidin"~e.5))))) :ARG1-of (d / describe-01 :ARG0 (a5 / and :op1 (f2 / figure~e.43 :mod "1B"~e.45) :op2 (p / panel~e.49 :ARG1-of (l2 / low-04~e.48 :degree~e.48 (m3 / more~e.48)))))) # ::id a_pmid_2194_3101.147 ::amr-annotator SDL-AMR-09 ::preferred # ::tok Significantly enlarged and more compact spheroids without lumen were formed by Caco @-@ BR13 cells as compared to Caco @-@ 2 cells . # ::alignments 0-1.2.1.1 1-1.2.1 3-1.2.2.1 4-1.2.2 5-1.2 6-1.2.3.1 6-1.2.3.1.r 7-1.2.3.2 9-1 10-1.1.r 11-1.1.1.1 13-1.1.1.1 14-1.1 14-1.3 16-1.3.r 18-1.3.1.1 20-1.3.1.1 21-1.1 (f / form-01~e.9 :ARG0~e.10 (c2 / cell-line~e.14,21 :name (n / name :op1 "Caco-BR13"~e.11,13)) :ARG1 (s / spheroid~e.5 :ARG1-of (e / enlarge-01~e.1 :ARG2 (s2 / significant-02~e.0)) :ARG1-of (c / compact-01~e.4 :degree (m / more~e.3)) :ARG0-of (h / have-03 :polarity~e.6 -~e.6 :ARG1 (l2 / lumen~e.7))) :compared-to~e.16 (c3 / cell-line~e.14 :name (n2 / name :op1 "Caco-2"~e.18,20))) # ::id a_pmid_2194_3101.148 ::amr-annotator SDL-AMR-09 ::preferred # ::tok In the case of Caco @-@ H2 cells , no typical spheroids were formed , instead large masses with non @-@ canonical shape were observed , typical of cancer cells . # ::alignments 3-1.2 4-1.3.1.1 6-1.3.1.1 7-1.3 9-1.1.2.1.r 10-1.2.1.1.1 11-1.2.1.1 13-1.2.1 15-1.2 16-1.1.1 17-1.1 19-1.1.2.1 19-1.1.2.1.r 21-1.1.2.2 22-1.1.2 24-1 26-1.2.1.1.1 27-1.2 28-1.1.3.1.1.2.1 29-1.1.3.1 (o / observe-01~e.24 :ARG1 (m / mass~e.17 :mod (l / large~e.16) :ARG1-of (s2 / shape-01~e.22 :polarity~e.9,19 -~e.19 :ARG2 (c3 / canonical~e.21)) :ARG0-of (t2 / typify-01 :ARG1 (c4 / cell~e.29 :mod (d / disease :wiki "Cancer" :name (n2 / name :op1 "cancer"~e.28))))) :ARG1-of (i / instead-of-91~e.3,15,27 :ARG2 (f / form-01~e.13 :ARG1 (s / spheroid~e.11 :ARG1-of (t / typical-02~e.10,26)))) :topic (c2 / cell-line~e.7 :name (n / name :op1 "Caco-H2"~e.4,6))) # ::id a_pmid_2194_3101.149 ::amr-annotator SDL-AMR-09 ::preferred # ::tok Therefore , under 2D as well as 3D culture conditions BRAF @ V600E @ overexpression managed to alter the morphology of colon adenocarcinoma cells , rendering them a more mesenchymal @-@ like phenotype , while KRAS @ G12V @ conserved the epithelial architecture of Caco @-@ 2 cells in general . # ::alignments 0-1 3-1.1.3.1.1.1 3-1.1.3.1.1.1.r 4-1.1.3 5-1.1.3 6-1.1.3 7-1.1.3.1.1 7-1.1.3.2.1 7-1.1.3.2.1.1 7-1.1.3.2.1.1.r 8-1.1.3.1 8-1.1.3.2 9-1.1.3.r 10-1.1.1.1.1.1.1 12-1.1.1.1.1.2.1 14-1.1.1.1 15-1.1.1 17-1.1.1.1.2 19-1.1.1.1.2.1 20-1.1.1.1.2.1.1.r 21-1.1.1.1.2.1.1.1.1 22-1.1.1.1.2.1.1.1 23-1.1.1.1.2.1.1 25-1.1.1.2 26-1.1.1.2.2 28-1.1.1.2.1.1.1.1.1 29-1.1.1.2.1.1.1.1 31-1.1.1.2.1.1 32-1.1.1.2.1 32-1.1.1.2.1.1.1 34-1.1 35-1.1.2.1.1.1 37-1.1.2.1.2.1 39-1.1.2 41-1.1.2.2.1 42-1.1.2.2 43-1.1.2.2.2.r 44-1.1.2.2.2.1.1 46-1.1.2.2.2.1.1 47-1.1.2.2.2 48-1.1.2.3.r 49-1.1.2.3 (c / cause-01~e.0 :ARG1 (c10 / contrast-01~e.34 :ARG1 (m / manage-02~e.15 :ARG0 (o / overexpress-01~e.14 :ARG1 (e3 / enzyme :name (n3 / name :op1 "BRAF"~e.10) :ARG1-of (m3 / mutate-01 :value "V600E"~e.12)) :ARG0-of (a2 / alter-01~e.17 :ARG1 (m4 / morphology~e.19 :poss~e.20 (c5 / cell~e.23 :source (a3 / adenocarcinoma~e.22 :part-of (c6 / colon~e.21)))))) :ARG0-of (r / render-02~e.25 :ARG1 (p / phenotype~e.32 :ARG1-of (r2 / resemble-01~e.31 :ARG2 (p2 / phenotype~e.32 :mod (m5 / mesenchyme~e.29 :degree (m6 / more~e.28))))) :ARG2 c5~e.26)) :ARG2 (c2 / conserve-01~e.39 :ARG0 (e2 / enzyme :name (n / name :op1 "KRAS"~e.35) :ARG1-of (m2 / mutate-01 :value "G12V"~e.37)) :ARG1 (a / architecture~e.42 :mod (e / epithelium~e.41) :source~e.43 (c3 / cell-line~e.47 :name (n2 / name :op1 "Caco-2"~e.44,46))) :ARG1-of~e.48 (g2 / general-02~e.49)) :condition~e.9 (a4 / and~e.4,5,6 :op1 (c8 / culture~e.8 :mod (d / dimension~e.7 :quant~e.3 2~e.3)) :op2 (c9 / culture~e.8 :mod (d2 / dimension~e.7 :quant~e.7 3~e.7))))) # ::id a_pmid_2194_3101.150 ::amr-annotator SDL-AMR-09 ::preferred # ::tok BRAF @ V600E @ downregulates E @-@ cadherin at the mRNA level and impairs its distribution in human colon adenocarcinoma cells # ::alignments 0-1.1.2.1.1 2-1.1.2.2.1 4-1.1 5-1.1.1.1.1 7-1.1.1.1.1 8-1.1.3.r 10-1.1.3.1.1.1 11-1.1.3 12-1 13-1.2 15-1.2.2 16-1.2.2.2.r 17-1.2.2.2.1.1.1 18-1.2.2.2.1.1 19-1.2.2.2.1 20-1.2.2.2 (a / and~e.12 :op1 (d / downregulate-01~e.4 :ARG1 (p / protein :name (n2 / name :op1 "E-cadherin"~e.5,7)) :ARG2 (e / enzyme :name (n / name :op1 "BRAF"~e.0) :ARG1-of (m / mutate-01 :value "V600E"~e.2)) :location~e.8 (l / level~e.11 :quant-of (n4 / nucleic-acid :name (n3 / name :op1 "mRNA"~e.10)))) :op2 (i / impair-01~e.13 :ARG0 e :ARG1 (d2 / distribute-01~e.15 :ARG1 p :location~e.16 (c / cell~e.20 :source (a2 / adenocarcinoma~e.19 :mod (c2 / colon~e.18 :mod (h / human~e.17))))))) # ::id a_pmid_2194_3101.151 ::amr-annotator SDL-AMR-09 ::preferred # ::tok It has been previously shown that HRAS @ G12V @ converts Caco @-@ 2 epithelial into mesenchymal cells by inducing loss of E @-@ cadherin and overexpression of vimentin [ @ 25 @ ] . # ::alignments 3-1.3 4-1 5-1.1.r 6-1.1.1.1.1 8-1.1.1.2.1 10-1.1 11-1.1.2.1.1 13-1.1.2.1.1 14-1.1.2.2 15-1.1.3.r 16-1.1.3.1 17-1.1.3 18-1.1.4.r 19-1.1.4 20-1.1.4.2.1 21-1.1.4.2.1.1.r 22-1.1.4.2.1.1.1.1 24-1.1.4.2.1.1.1.1 25-1.1.4.2 26-1.1.4.2.2 27-1.1.4.2.2.1.r 28-1.1.4.2.2.1.1.1 31-1.2.1.1.1 (s / show-01~e.4 :ARG1~e.5 (c2 / convert-01~e.10 :ARG0 (g / gene :name (n3 / name :op1 "HRAS"~e.6) :ARG1-of (m / mutate-01 :value "G12V"~e.8)) :ARG1 (c3 / cell-line :name (n4 / name :op1 "Caco-2"~e.11,13) :mod (e / epithelium~e.14)) :ARG2~e.15 (c4 / cell~e.17 :location (m2 / mesenchyme~e.16)) :manner~e.18 (i / induce-01~e.19 :ARG0 g :ARG2 (a / and~e.25 :op1 (l / lose-02~e.20 :ARG1~e.21 (p4 / protein :name (n2 / name :op1 "E-cadherin"~e.22,24))) :op2 (o / overexpress-01~e.26 :ARG1~e.27 (p3 / protein :name (n / name :op1 "vimentin"~e.28)))))) :ARG1-of (d / describe-01 :ARG0 (p / publication :ARG1-of (c / cite-01 :ARG2 25~e.31))) :time (p2 / previous~e.3)) # ::id a_pmid_2194_3101.152 ::amr-annotator SDL-AMR-09 ::preferred # ::tok In order to examine whether BRAF @ V600E @ had a similar effect on Caco @-@ 2 cells , the expression and localization of E @-@ cadherin was analyzed ( Figure 2A , B ) . # ::alignments 0-1.3.r 1-1.3.r 2-1.3.r 3-1.3 4-1.3.1.1 4-1.3.1.1.r 5-1.3.1.2.1.1.1 7-1.3.1.2.1.2.1 11-1.3.1.2.3 12-1.3.1.2 13-1.3.1.2.2.r 14-1.3.1.2.2.1.1 16-1.3.1.2.2.1.1 17-1.3.1.2.2 20-1.1.1 21-1.1 22-1.1.2 24-1.1.2.1.1.1 26-1.1.2.1.1.1 27-1.1.2 28-1 30-1.2.1.1 30-1.2.1.2 32-1.2.1.1.1 (a / analyze-01~e.28 :ARG1 (a3 / and~e.21 :op1 (e / express-03~e.20 :ARG2 p) :op2 (b / be-located-at-91~e.22,27 :ARG2 (p / protein :name (n / name :op1 "E-cadherin"~e.24,26)))) :ARG1-of (d / describe-01 :ARG0 (a2 / and :op1 (f / figure~e.30 :mod "2A"~e.32) :op2 (f2 / figure~e.30 :mod "2B"))) :purpose~e.0,1,2 (e2 / examine-01~e.3 :ARG1 (p2 / possible-01 :mode~e.4 interrogative~e.4 :ARG1 (a4 / affect-01~e.12 :ARG0 (e3 / enzyme :name (n2 / name :op1 "BRAF"~e.5) :ARG1-of (m / mutate-01 :value "V600E"~e.7)) :ARG1~e.13 (c / cell-line~e.17 :name (n3 / name :op1 "Caco-2"~e.14,16)) :ARG1-of (r / resemble-01~e.11))))) # ::id a_pmid_2194_3101.153 ::amr-annotator SDL-AMR-09 ::preferred # ::tok Transformation of Caco @-@ 2 cells with BRAF @ V600E @ led to a significant decrease in the mRNA levels of E @-@ cadherin but had no significant effect on the actual protein expression ( Figure 2A ) . # ::alignments 0-1.1.1 1-1.1.1.2.r 2-1.1.1.2.1.1 4-1.1.1.2.1.1 5-1.1.1.2 6-1.1.1.1.r 7-1.1.1.1.1.1 9-1.1.1.1.2.1 11-1.1 14-1.1.2.2 15-1.1.2 16-1.1.2.1.r 18-1.1.2.1.1.1.1 19-1.1.2.1 21-1.1.2.1.1.2.1.1 23-1.1.2.1.1.2.1.1 24-1 26-1.2.1 26-1.2.1.r 27-1.2.4 28-1.2 29-1.2.3.r 31-1.2.3.2 32-1.2.3.1 33-1.2.3 35-1.3.1 37-1.3.1.1 (c / contrast-01~e.24 :ARG1 (l / lead-03~e.11 :ARG0 (t / transform-01~e.0 :ARG0~e.6 (e2 / enzyme :name (n2 / name :op1 "BRAF"~e.7) :ARG1-of (m / mutate-01 :value "V600E"~e.9)) :ARG1~e.1 (c2 / cell-line~e.5 :name (n / name :op1 "Caco-2"~e.2,4))) :ARG1 (d / decrease-01~e.15 :ARG1~e.16 (l2 / level~e.19 :quant-of (n5 / nucleic-acid :name (n3 / name :op1 "mRNA"~e.18) :mod (p / protein :name (n4 / name :op1 "E-cadherin"~e.21,23)))) :ARG2 (s / significant-02~e.14))) :ARG2 (a / affect-01~e.28 :polarity~e.26 -~e.26 :ARG0 t :ARG1~e.29 (e / express-03~e.33 :ARG2 p~e.32 :ARG1-of (a2 / actual-02~e.31)) :ARG1-of s~e.27) :ARG1-of (d2 / describe-01 :ARG0 (f / figure~e.35 :mod "2A"~e.37))) # ::id a_pmid_2194_3101.154 ::amr-annotator SDL-AMR-09 ::preferred # ::tok Notably , in Caco @-@ BR cells reduced intensity for E @-@ cadherin was observed mostly in lower molecular weight protein bands representing the mature protein at 120 kDa ( Figure 2A @ -@ lower panel , high exposure ) , whereas the decrease in the actual precursors at 135 kDa ( Figure 2A @ -@ lower panel , low exposure ) , is considerably less . # ::alignments 0-1.3 3-1.1.4.1.1 5-1.1.4.1.1 6-1.1.4 7-1.1.1.1 8-1.1.1 9-1.1.1.2.r 10-1.1.1.2.1.1 12-1.1.1.2.1.1 12-1.1.2.2.1.1.1 14-1.1 15-1.1.3 16-1.1.2.r 17-1.1.2.1.1 17-1.1.2.1.1.2 17-1.1.2.1.1.2.r 18-1.1.2.1.1.1.1 19-1.1.2.1.1.1 20-1.1.2.1 21-1.1.2 22-1.1.2.2 24-1.1.2.2.1.2 25-1.1.2.2.1 26-1.1.2.2.1.3.r 27-1.1.2.2.1.3.1 28-1.1.2.2.1.3.2 30-1.1.5.1.1 30-1.2.3.1.1 32-1.1.5.1.1.1 32-1.2.3.1.1.1 35-1.1.5.1.2.1 35-1.1.5.1.2.1.1 35-1.1.5.1.2.1.1.r 35-1.2.3.1.2.1 35-1.2.3.1.2.1.1 35-1.2.3.1.2.1.1.r 36-1.1.5.1.2 36-1.2.3.1.2 38-1.1.5.1.3.1 39-1.1.5.1.3 42-1 44-1.2 45-1.2.1.r 47-1.2.1.1 48-1.2.1 49-1.2.1.2.r 50-1.2.1.2.1 51-1.2.1.2.2 53-1.2.3.1.1 55-1.2.3.1.1.1 58-1.2.3.1.2.1 58-1.2.3.1.2.1.1 58-1.2.3.1.2.1.1.r 59-1.2.3.1.2 61-1.2.3.1.3.1 62-1.2.3.1.3 66-1.2.2.1 67-1.2.2 (c / contrast-01~e.42 :ARG1 (o / observe-01~e.14 :ARG1 (i / intensity~e.8 :ARG1-of (r2 / reduce-01~e.7) :poss~e.9 (p5 / protein :name (n4 / name :op1 "E-cadherin"~e.10,12))) :location~e.16 (b / band~e.21 :mod (p3 / protein~e.20 :ARG1-of (l4 / low-04~e.17 :ARG2 (w / weight-01~e.19 :ARG1 (m4 / molecule~e.18)) :degree~e.17 (m5 / more~e.17))) :ARG0-of (r / represent-01~e.22 :ARG1 (p4 / protein~e.25 :name (n2 / name :op1 "E-cadherin"~e.12) :ARG1-of (m / mature-02~e.24) :quant~e.26 (m7 / mass-quantity :quant 120~e.27 :unit k~e.28)))) :degree (m3 / most~e.15) :location (c3 / cell-line~e.6 :name (n3 / name :op1 "Caco-BR"~e.3,5)) :ARG1-of (d3 / describe-01 :ARG0 (a3 / and :op1 (f2 / figure~e.30 :mod "2A"~e.32) :op2 (p6 / panel~e.36 :ARG1-of (l5 / low-04~e.35 :degree~e.35 (m8 / more~e.35))) :op3 (e2 / expose-01~e.39 :ARG1-of (h / high-02~e.38))))) :ARG2 (d / decrease-01~e.44 :ARG1~e.45 (p / precursor~e.48 :ARG1-of (a / actual-02~e.47) :quant~e.49 (m9 / mass-quantity :quant 135~e.50 :unit (k / kilodalton~e.51))) :ARG2 (l6 / less~e.67 :degree (c2 / considerable~e.66)) :ARG1-of (d2 / describe-01 :ARG0 (a2 / and :op1 (f / figure~e.30,53 :mod "2A"~e.32,55) :op2 (p2 / panel~e.36,59 :ARG1-of (l / low-04~e.35,58 :degree~e.35,58 (m2 / more~e.35,58))) :op3 (e / exposure~e.62 :ARG1-of (l2 / low-04~e.61))))) :ARG1-of (n / notable-04~e.0)) # ::id a_pmid_2194_3101.155 ::amr-annotator SDL-AMR-09 ::preferred # ::tok It appears that mutant BRAF @ V600E @ but not upstream KRAS @ G12V @ activation is able to suppress the mature E @-@ cadherin , while the precursor remained mostly unaffected . # ::alignments 1-1 2-1.1.r 3-1.1.1.1.1 3-1.1.1.1.1.2 3-1.1.1.1.1.2.r 3-1.1.1.1.2.1.1 3-1.1.1.1.2.1.1.2 3-1.1.1.1.2.1.1.2.r 4-1.1.1.1.1.1.1 6-1.1.1.1.1.2.1 8-1.1.1.1.2 9-1.1.1.1.2.1.2.1 9-1.1.1.1.2.1.2.1.r 10-1.1.1.1.2.1.3 11-1.1.1.1.2.1.1.1.1 13-1.1.1.1.2.1.1.2.1 15-1.1.1.1 15-1.1.1.1.2.1 17-1.1.1 17-1.1.1.1.2.1.2 19-1.1.1.2 21-1.1.1.2.1 21-1.1.1.2.1.2 21-1.1.1.2.1.2.r 22-1.1.1.2.1.1.1 24-1.1.1.2.1.1.1 26-1.1 28-1.1.2.1 29-1.1.2 30-1.1.2.2.2 31-1.1.2.2 31-1.1.2.2.1 31-1.1.2.2.1.r (a / appear-01~e.1 :ARG1~e.2 (c3 / contrast-01~e.26 :ARG1 (c2 / capable-01~e.17 :ARG1 (a3 / activate-01~e.15 :ARG1 (e / enzyme~e.3 :name (n / name :op1 "BRAF"~e.4) :ARG1-of~e.3 (m / mutate-01~e.3 :value "V600E"~e.6)) :ARG1-of (c / contrast-01~e.8 :ARG2 (a4 / activate-01~e.15 :ARG1 (e2 / enzyme~e.3 :name (n2 / name :op1 "KRAS"~e.11) :ARG1-of~e.3 (m2 / mutate-01~e.3 :value "G12V"~e.13)) :ARG1-of (c4 / capable-01~e.17 :polarity~e.9 -~e.9 :ARG2 s) :location (u / upstream~e.10)))) :ARG2 (s / suppress-01~e.19 :ARG1 (p / protein~e.21 :name (n3 / name :op1 "E-cadherin"~e.22,24) :ARG1-of~e.21 (m3 / mature-02~e.21)))) :ARG2 (r / remain-01~e.29 :ARG1 (p2 / precursor~e.28) :ARG3 (a2 / affect-01~e.31 :polarity~e.31 -~e.31 :degree (m4 / most~e.30))))) # ::id a_pmid_2194_3101.156 ::amr-annotator SDL-AMR-09 ::preferred # ::tok Nevertheless , immunostaining with E @-@ cadherin revealed a significant impairment of its distribution at the cell @-@ cell boundaries since staining appeared discontinuous at the adherent junctions ( Figure 2B , upper panel magnification ) . # ::alignments 0-1 2-1.1.2.1 3-1.1.2.1.1.r 4-1.1.2.1.1.1.1 6-1.1.2.1.1.1.1 7-1.1.2 9-1.1.2.2.2 10-1.1.2.2 11-1.1.2.2.1.r 12-1.1.2.2.1.1 12-1.1.2.2.1.1.r 13-1.1.2.2.1 14-1.1.2.2.1.2.r 16-1.1.2.2.1.2.1.2 18-1.1.2.2.1.2.1.2 19-1.1.2.2.1.2 20-1.1 21-1.1.1.1.2 22-1.1.1 23-1.1.1.1 23-1.1.1.1.1 23-1.1.1.1.1.r 24-1.1.1.2.r 26-1.1.1.2.1.1 27-1.1.1.2.1.2 29-1.2.1.1 31-1.2.1.1.1 34-1.2.1.2.1.1 35-1.2.1.2.1 36-1.2.1.2 (h / have-concession-91~e.0 :ARG1 (c2 / cause-01~e.20 :ARG0 (a / appear-01~e.22 :ARG1 (c / continue-01~e.23 :polarity~e.23 -~e.23 :ARG1 (s / stain-01~e.21)) :location~e.24 (m3 / macro-molecular-complex :name (n2 / name :op1 "adherent"~e.26 :op2 "junction"~e.27))) :ARG1 (r / reveal-01~e.7 :ARG0 (i / immunostain-01~e.2 :ARG2~e.3 (p / protein :name (n / name :op1 "E-cadherin"~e.4,6))) :ARG1 (i2 / impair-01~e.10 :ARG1~e.11 (d2 / distribute-01~e.13 :ARG1~e.12 p~e.12 :ARG2~e.14 (b / boundary~e.19 :mod (b2 / between :op1 c3 :op2 (c3 / cell~e.16,18)))) :ARG1-of (s2 / significant-02~e.9)))) :ARG1-of (d3 / describe-01 :ARG0 (a3 / and :op1 (f / figure~e.29 :mod "2B"~e.31) :op2 (m / magnify-01~e.36 :ARG1 (p2 / panel~e.35 :mod (u / upper~e.34)))))) # ::id a_pmid_2194_3101.157 ::amr-annotator SDL-AMR-09 ::preferred # ::tok Expression of E @-@ cadherin in the Caco @-@ BR grown in 3D spheroids was found significantly downregulated with diffused distribution ( Figure 2B , lower panel ) . # ::alignments 0-1.1.1 1-1.1.1.1.r 2-1.1.1.1.1.1 4-1.1.1.1.1.1 5-1.1.1.2.r 7-1.1.1.2.1.1 9-1.1.1.2.1.1 10-1.1.1.2 10-1.1.1.2.2 10-1.1.1.2.2.r 11-1.1.1.2.2.1.r 12-1.1.1.2.2.1.1 12-1.1.1.2.2.1.1.1 12-1.1.1.2.2.1.1.1.r 13-1.1.1.2.2.1 15-1 16-1.1.3 17-1.1 18-1.1.2.r 19-1.1.2.1 20-1.1.2 22-1.2.1.1 24-1.2.1.1.1 27-1.2.1.2.1 27-1.2.1.2.1.1 27-1.2.1.2.1.1.r 28-1.2.1.2 (f / find-01~e.15 :ARG1 (d2 / downregulate-01~e.17 :ARG1 (e / express-03~e.0 :ARG2~e.1 (p2 / protein :name (n / name :op1 "E-cadherin"~e.2,4)) :ARG3~e.5 (c / cell-line~e.10 :name (n2 / name :op1 "Caco-BR"~e.7,9) :ARG1-of~e.10 (g / grow-03~e.10 :location~e.11 (s / spheroid~e.13 :mod (d5 / dimension~e.12 :quant~e.12 3~e.12))))) :manner~e.18 (d3 / distribute-01~e.20 :ARG1-of (d4 / diffuse-01~e.19)) :ARG1-of (s2 / significant-02~e.16)) :ARG1-of (d / describe-01 :ARG0 (a / and :op1 (f2 / figure~e.22 :mod "2B"~e.24) :op2 (p / panel~e.28 :ARG1-of (l / low-04~e.27 :degree~e.27 (m / more~e.27)))))) # ::id a_pmid_2194_3101.158 ::amr-annotator SDL-AMR-09 ::preferred # ::tok In contrast , the epithelial marker E @-@ cadherin was normally localized at the cell @-@ cell junctions of Caco @-@ 2 and Caco @-@ K15 cells ( Figure 2B , lower panel magnification ) . # ::alignments 1-1 4-1.1.1.2 5-1.1.1 6-1.1.1.1.1 8-1.1.1.1.1 9-1.1 10-1.1.3 11-1.1 12-1.1 14-1.1.2.1.1 16-1.1.2.1.1 17-1.1.2 19-1.1.2.1.1.1.1 19-1.1.2.1.2.1.1 21-1.1.2.1.1.1.1 22-1.1.2.1 23-1.1.2.1.1.1.1 23-1.1.2.1.2.1.1 25-1.1.2.1.2.1.1 26-1.1.2.1.1 26-1.1.2.1.2 28-1.2.1.1 30-1.2.1.1.1 33-1.2.1.2.1.1 34-1.2.1.2.1 35-1.2.1.2 (c / contrast-01~e.1 :ARG2 (b / be-located-at-91~e.9,11,12 :ARG1 (m3 / marker~e.5 :name (n2 / name :op1 "E-cadherin"~e.6,8) :mod (e / epithelium~e.4)) :ARG2 (j3 / junction~e.17 :mod (b2 / between~e.22 :op1 (c2 / cell-line~e.14,16,26 :name (n3 / name :op1 "Caco-2"~e.19,21,23)) :op2 (c3 / cell-line~e.26 :name (n4 / name :op1 "Caco-K15"~e.19,23,25)))) :ARG1-of (n / normal-02~e.10)) :ARG1-of (d / describe-01 :ARG0 (a / and :op1 (f / figure~e.28 :mod "2B"~e.30) :op2 (m / magnify-01~e.35 :ARG1 (p / panel~e.34 :ARG1-of (l2 / low-04~e.33)))))) # ::id a_pmid_2194_3101.159 ::amr-annotator SDL-AMR-09 ::preferred # ::tok In order to determine whether Caco @-@ BR cells have acquired more mesenchymal characteristics , RNA and protein levels of the mesenchymal marker Vimentin were examined ( Figure 2C ) . # ::alignments 0-1.2.r 1-1.2.r 2-1.2.r 3-1.2 4-1.2.1.1 4-1.2.1.1.r 5-1.2.1.2.1.1 7-1.2.1.2.1.1 8-1.2.1.2 10-1.2.1 11-1.2.1.3.1 12-1.2.1.3.2.1 13-1.2.1.3 13-1.2.1.3.2 13-1.2.1.3.2.r 15-1.1.1.1.1.1 16-1.1 17-1.1.2.1 18-1.1.1 18-1.1.2 21-1.1.3.2 22-1.1.3 23-1.1.3.1.1 25-1 27-1.3.1 29-1.3.1.1 (e / examine-01~e.25 :ARG1 (a / and~e.16 :op1 (l / level~e.18 :quant-of (n3 / nucleic-acid :name (n4 / name :op1 "RNA"~e.15))) :op2 (l2 / level~e.18 :quant-of (p / protein~e.17)) :poss (m4 / marker~e.22 :name (n / name :op1 "Vimentin"~e.23) :mod (m / mesenchyme~e.21))) :purpose~e.0,1,2 (d / determine-01~e.3 :ARG1 (a2 / acquire-01~e.10 :mode~e.4 interrogative~e.4 :ARG0 (c / cell-line~e.8 :name (n2 / name :op1 "Caco-BR"~e.5,7)) :ARG1 (t / thing~e.13 :quant (m3 / more~e.11) :ARG2-of~e.13 (c2 / characteristic-02~e.13 :ARG1 m~e.12)))) :ARG1-of (d2 / describe-01 :ARG0 (f / figure~e.27 :mod "2C"~e.29))) # ::id a_pmid_2194_3101.160 ::amr-annotator SDL-AMR-09 ::preferred # ::tok An increase of about 3 @-@ fold was observed at the protein level , while confocal images did not show significant difference , as compared to Caco @-@ 2 ( Figure 2D ) , since it is known that some cancer epithelial cells abnormally express N @-@ cadherin which has been shown to promote motility and invasion [ @ 26 , 27 @ ] , N @-@ cadherin expression was examined ( Figure 2E ) . # ::alignments 1-1.1.1.1 2-1.1.1.1.2.1.1 3-1.1.1.1.2 3-1.1.1.1.2.1 3-1.1.1.1.2.1.r 4-1.1.1.1.2.1.1.1 6-1.1.1.1.2.1.1 8-1.1.1 9-1.1.1.1.1.r 11-1.1.1.1.1.1 12-1.1.1.1.1 14-1.1 15-1.1.2.2.1 16-1.1.2.2 18-1.1.2.1 18-1.1.2.1.r 19-1.1.2 20-1.1.2.3.1 21-1.1.2.3 24-1.1.2.4.r 26-1.1.2.4.1.1 28-1.1.2.4.1.1 30-1.1.2.5.1 32-1.1.2.5.1.1 36-1.2 39-1.2.1 40-1.2.1.1.r 41-1.2.1.1.2.1 42-1.2.1.1.2.3.2.1 43-1.2.1.1.2.2 44-1.2.1.1.2 45-1.2.1.1.3 45-1.2.1.1.3.r 46-1.2.1.1 47-1.2.1.1.1.1.1 49-1.2.1.1.1.1.1 53-1.2.1.1.1.2.2 55-1.2.1.1.1 55-1.2.1.1.1.2 55-1.2.1.1.1.2.r 56-1.2.1.1.1.2.1.1 57-1.2.1.1.1.2.1 58-1.2.1.1.1.2.1.2 61-1.2.1.1.1.2.2.1.1.1.1 65-1.2.1.1.1.2.2.1.2.1.1 69-1.2.1.1.1.1.1 71-1.2.1.1.1.1.1 72-1.2.1.1 74-1.2.2 76-1.2.2.2.1 78-1.2.2.2.1.1 (a5 / and :op1 (c2 / contrast-01~e.14 :ARG1 (o / observe-01~e.8 :ARG1 (i2 / increase-01~e.1 :ARG1~e.9 (l / level~e.12 :quant-of (p5 / protein~e.11)) :ARG3 (a6 / about~e.3 :quant~e.3 (a4 / about~e.3 :op1 (p6 / product-of~e.2,6 :op1 3~e.4))))) :ARG2 (s / show-01~e.19 :polarity~e.18 -~e.18 :ARG0 (i3 / image~e.16 :mod (c8 / confocal~e.15)) :ARG1 (d4 / differ-02~e.21 :ARG1-of (s4 / significant-02~e.20)) :compared-to~e.24 (c / cell-line :name (n3 / name :op1 "Caco-2"~e.26,28)) :ARG1-of (d / describe-01 :ARG0 (f / figure~e.30 :mod "2D"~e.32)))) :op2 (c9 / cause-01~e.36 :ARG0 (k / know-01~e.39 :ARG1~e.40 (e2 / express-03~e.46,72 :ARG2 (p / protein~e.55 :name (n2 / name :op1 "N-cadherin"~e.47,49,69,71) :ARG0-of~e.55 (p2 / promote-01~e.55 :ARG1 (a2 / and~e.57 :op1 (m / motility~e.56) :op2 (i / invade-01~e.58)) :ARG1-of (s3 / show-01~e.53 :ARG0 (a3 / and :op1 (p3 / publication :ARG1-of (c6 / cite-01 :ARG2 26~e.61)) :op2 (p4 / publication :ARG1-of (c7 / cite-01 :ARG2 27~e.65)))))) :ARG3 (c5 / cell~e.44 :quant (s2 / some~e.41) :source (e3 / epithelium~e.43) :mod (d2 / disease :wiki "Cancer" :name (n / name :op1 "cancer"~e.42))) :manner~e.45 (a / abnormal~e.45))) :ARG1 (e / examine-01~e.74 :ARG1 e2 :ARG1-of (d3 / describe-01 :ARG0 (f2 / figure~e.76 :mod "2E"~e.78))))) # ::id a_pmid_2194_3101.161 ::amr-annotator SDL-AMR-09 ::preferred # ::tok In Caco @-@ BR cells N @-@ cadherin expression is increased about 2 @-@ fold both at mRNA and protein levels , as compared to Caco @-@ 2 cells . # ::alignments 1-1.1.2.1.1 1-1.4.1.1 3-1.1.2.1.1 4-1.1.2 5-1.1.1.1.1 7-1.1.1.1.1 8-1.1 10-1 11-1.2 12-1.4.1.1 14-1.2.1 16-1.3.r 17-1.3.1.1.1.1 18-1.3 19-1.3.2.1 20-1.3.1 20-1.3.2 23-1.4.r 25-1.4.1.1 27-1.4.1.1 28-1.4 (i / increase-01~e.10 :ARG1 (e / express-03~e.8 :ARG2 (p / protein :name (n2 / name :op1 "N-cadherin"~e.5,7)) :ARG3 (c2 / cell-line~e.4 :name (n3 / name :op1 "Caco-BR"~e.1,3))) :ARG2 (a2 / about~e.11 :op1 (p2 / product-of~e.14 :op1 12)) :prep-at~e.16 (a / and~e.18 :op1 (l2 / level~e.20 :mod (n5 / nucleic-acid :name (n4 / name :op1 "mRNA"~e.17))) :op2 (l / level~e.20 :mod (p3 / protein~e.19))) :compared-to~e.23 (c / cell-line~e.28 :name (n / name :op1 "Caco-2"~e.1,12,25,27))) # ::id a_pmid_2194_3101.162 ::amr-annotator SDL-AMR-09 ::preferred # ::tok Confocal images confirmed this increase , as shown in Figure 2F @ . # ::alignments 0-1.1.1 1-1.1 2-1 3-1.2.1 4-1.2 6-1.3.r 7-1.3 8-1.3.1.r 9-1.3.1 11-1.3.1.1 (c / confirm-01~e.2 :ARG0 (i / images~e.1 :mod (c2 / confocal~e.0)) :ARG1 (i2 / increase-01~e.4 :mod (t / this~e.3)) :ARG1-of~e.6 (s / show-01~e.7 :ARG0~e.8 (f / figure~e.9 :mod "2F"~e.11))) # ::id a_pmid_2194_3101.163 ::amr-annotator SDL-AMR-09 ::preferred # ::tok Taken together these data suggest that BRAF @ V600E @ overexpression failed to induce an integrated EMT phenotype , which is the case with HRAS @ G12V @ overexpression [ @ 25 @ ] , but managed to transform Caco @-@ 2 cells through the loss of some important epithelial characteristics . # ::alignments 0-1.1.2 1-1.1.2.1 2-1.1.1 3-1.1 4-1 5-1.2.r 6-1.2.1.1.1.1.1 8-1.2.1.1.1.2.1 10-1.2.1.1 11-1.2.1 13-1.2.1.2 15-1.2.1.2.2.2 16-1.2.1.2.2.1 17-1.2.1.2.2 24-1.2.1.3.1.1.1.1 26-1.2.1.3.1.1.2.1 28-1.2.1.3.1 31-1.2.1.3.1.2.1.1.1 35-1.2 35-1.2.1.3 36-1.2.2 38-1.2.2.2 39-1.2.2.2.2.1.1 41-1.2.2.2.2.1.1 42-1.2.2.2.2 45-1.2.2.3 46-1.2.2.3.2.r 47-1.2.2.3.2.2.2 48-1.2.2.3.2.1 49-1.2.1.2.2.1.2 49-1.2.2.3.2.2.1 50-1.2.2.3.2 50-1.2.2.3.2.2 50-1.2.2.3.2.2.r (s / suggest-01~e.4 :ARG0 (d / data~e.3 :mod (t2 / this~e.2) :ARG1-of (t / take-01~e.0 :mod (t3 / together~e.1))) :ARG1~e.5 (c4 / contrast-01~e.35 :ARG1 (f / fail-01~e.11 :ARG1 (o / overexpress-01~e.10 :ARG1 (e3 / enzyme :name (n / name :op1 "BRAF"~e.6) :ARG1-of (m / mutate-01 :value "V600E"~e.8))) :ARG2 (i / induce-01~e.13 :ARG0 o :ARG2 (p / phenotype~e.17 :mod (t5 / transition-01~e.16 :ARG2 (m4 / mesenchyme) :ARG3 (e4 / epithelium~e.49)) :ARG1-of (i2 / integrate-01~e.15))) :ARG2-of (c5 / contrast-01~e.35 :ARG1 (o2 / overexpress-01~e.28 :ARG1 (e2 / enzyme :name (n3 / name :op1 "HRAS"~e.24) :ARG1-of (m2 / mutate-01 :value "G12V"~e.26)) :ARG1-of (d2 / describe-01 :ARG0 (p2 / publication :ARG1-of (c / cite-01 :ARG2 25~e.31)))))) :ARG2 (m3 / manage-02~e.36 :ARG0 o :ARG1 (t4 / transform-01~e.38 :ARG0 o :ARG1 (c2 / cell-line~e.42 :name (n4 / name :op1 "Caco-2"~e.39,41))) :manner (l / lose-02~e.45 :ARG0 c2 :ARG1~e.46 (t6 / thing~e.50 :mod (i3 / important~e.48) :ARG2-of~e.50 (c3 / characteristic-02~e.50 :ARG1 (e / epithelium~e.49) :quant (s2 / some~e.47))))))) # ::id a_pmid_2194_3101.164 ::amr-annotator SDL-AMR-09 ::preferred # ::tok Differential BRAF @ V600E , KRAS @ G12V @ and HRAS @ G12V @ effect on the migration and invasion ability of Caco @-@ 2 cells in vitro # ::alignments 0-1.1.4 1-1.1.1.1.1 3-1.1.1.2.1 6-1.1.2.1.1 8-1.1.2.2.1 8-1.1.3.2.1 11-1.1.3.1.1 13-1.1.2.2.1 13-1.1.3.2.1 15-1 16-1.2.r 18-1.2.1.2 19-1.2 20-1.2.2.2 21-1.2.1 21-1.2.2 22-1.2.1.1.r 23-1.2.1.1.1.1 25-1.2.1.1.1.1 26-1.2.1.1 28-1.2.3 29-1.2.3 (a / affect-01~e.15 :ARG0 (a2 / and :op1 (e / enzyme :name (n / name :op1 "BRAF"~e.1) :ARG1-of (m / mutate-01 :value "V600E"~e.3)) :op2 (e2 / enzyme :name (n2 / name :op1 "KRAS"~e.6) :ARG1-of (m2 / mutate-01 :value "G12V"~e.8,13)) :op3 (e3 / enzyme :name (n3 / name :op1 "HRAS"~e.11) :ARG1-of (m3 / mutate-01 :value "G12V"~e.8,13)) :ARG1-of (d / differ-02~e.0)) :ARG1~e.16 (a3 / and~e.19 :op1 (c / capable-01~e.21 :ARG1~e.22 (c3 / cell-line~e.26 :name (n4 / name :op1 "Caco-2"~e.23,25)) :ARG2 (m4 / migrate-01~e.18 :ARG0 c3)) :op2 (c2 / capable-01~e.21 :ARG1 c3 :ARG2 (i / invade-01~e.20 :ARG0 c3)) :manner (i2 / in-vitro~e.28,29))) # ::id a_pmid_2194_3101.165 ::amr-annotator SDL-AMR-09 ::preferred # ::tok To further explore oncogenic effects on the cell cytoskeleton with regard to oncogenic transformation , the invasive and migratory properties of the previously established oncogenic cell models and in colon cancer cell lines HT29 and DLD @-@ 1 were analyzed . # ::alignments 1-1.2.3 2-1.2 4-1.2.1 5-1.2.1.2.r 7-1.2.1.2.1 8-1.2.1.2 13-1.2.2 16-1.1.1.1 17-1.1 17-1.1.1.2 18-1.1.2.1 19-1.1.1 19-1.1.2 20-1.1.1.2.r 22-1.1.1.2.1.1.1 23-1.1.1.2.1.1 25-1.1.1.2.1.3 26-1.1.1.2.1 27-1.1.1.2 29-1.1.1.2.2.2.3 30-1.1.1.2.2.2.2.1 31-1.1.1.2.2 32-1.1.1.2.2 33-1.1.1.2.2.1.1 34-1.1.1.2 35-1.1.1.2.3.1.1 37-1.1.1.2.3.1.1 39-1 (a / analyze-01~e.39 :ARG1 (a3 / and~e.17 :op1 (p / property~e.19 :mod (i / invade-01~e.16) :poss~e.20 (a4 / and~e.17,27,34 :op1 (m2 / model~e.26 :ARG1-of (e2 / establish-01~e.23 :time (p3 / previous~e.22)) :mod o :mod (c7 / cell~e.25)) :op2 (c4 / cell-line~e.31,32 :name (n / name :op1 "HT29"~e.33) :mod (d / disease :wiki "Cancer" :name (n3 / name :op1 "cancer"~e.30) :mod (c6 / colon~e.29))) :op3 (c5 / cell-line :name (n2 / name :op1 "DLD-1"~e.35,37) :mod d))) :op2 (p2 / property~e.19 :mod (m / migrate-01~e.18) :poss a4)) :purpose (e / explore-01~e.2 :ARG1 (a2 / affect-01~e.4 :ARG0 (o / oncogene) :ARG1~e.5 (c / cytoskeleton~e.8 :part-of (c2 / cell~e.7))) :ARG2 (t / transform-01~e.13 :mod o) :degree (f / further~e.1))) # ::id a_pmid_2194_3101.166 ::amr-annotator SDL-AMR-09 ::preferred # ::tok Transformation induced by each of the three oncogenes KRAS @ G12V @ @ ( Caco @-@ K cells ) , BRAF @ V600E @ @ ( Caco @-@ BR cells ) and HRAS @ G12V @ @ ( Caco @-@ H cells ) managed to increase the ability of Caco @-@ 2 cells to migrate and invade in vitro , independently of their proliferating ability , which has been previously analyzed in [ @ 21 @ ] . # ::alignments 0-1.1 1-1.1.1 9-1.1.1.1.1.1.1 15-1.1.1.1.1.2.1.1 17-1.1.1.1.1.2.1.1 18-1.1.1.1.1.2 22-1.1.1.1.2.1.1 28-1.1.1.1.2.2.1.1 30-1.1.1.1.2.2.1.1 31-1.1.1.1.1.2 35-1.1.1.1.3.1.1 41-1.1.1.1.3.2.1.1 43-1.1.1.1.3.2.1.1 44-1.1.1.1.1.2 44-1.1.1.1.3.2 46-1 47-1.1.1.1.4 48-1.2 50-1.2.2 52-1.2.2.1.1.1 54-1.2.2.1.1.1 55-1.1.1.1.2.2 55-1.2.2.1 56-1.2.2.2.r 57-1.2.2.2.1 58-1.2.2.2 59-1.2.2.2.2 61-1.2.2.2.3 62-1.2.2.2.3 65-1.2.2.2.4 65-1.2.2.2.4.r 68-1.2.2.2.4.1.2 69-1.2.2.2.4.1 74-1.1.1.1.2.2.2 75-1.2.2.2.4.1.3 76-1.2.2.2.3 79-1.2.2.2.4.1.3.1.1.1 (m / manage-01~e.46 :ARG0 (t / transform-01~e.0 :ARG2-of (i / induce-01~e.1 :ARG0 (a / and :op1 (g / gene :name (n4 / name :op1 "KRAS"~e.9) :location-of (c2 / cell~e.18,31,44 :name (n / name :op1 "Caco-K"~e.15,17))) :op2 (g2 / gene :name (n5 / name :op1 "BRAF"~e.22) :location-of (c3 / cell-line~e.55 :name (n2 / name :op1 "Caco-BR"~e.28,30) :time (p3 / previous~e.74))) :op3 (g3 / gene :name (n6 / name :op1 "HRAS"~e.35) :location-of (c4 / cell-line~e.44 :name (n3 / name :op1 "Caco-H"~e.41,43))) :ARG0-of (c8 / cause-01~e.47 :ARG1 (d / disease :wiki "Cancer" :name (n8 / name :op1 "cancer")))))) :ARG1 (i2 / increase-01~e.48 :ARG0 t :ARG1 (c5 / capable-01~e.50 :ARG1 (c6 / cell-line~e.55 :name (n7 / name :op1 "Caco-2"~e.52,54)) :ARG2~e.56 (a2 / and~e.58 :op1 (m2 / migrate-01~e.57 :ARG0 c6) :op2 (i3 / invade-01~e.59 :ARG0 c6) :manner (i4 / in-vitro~e.61,62,76) :manner~e.65 (i5 / independent~e.65 :topic (c7 / capable-01~e.69 :ARG1 c6 :ARG2 (p2 / proliferate-01~e.68 :ARG0 c6) :ARG1-of (a3 / analyze-01~e.75 :medium (p / publication :ARG1-of (c / cite-01 :ARG2 21~e.79))))))))) # ::id a_pmid_2194_3101.167 ::amr-annotator SDL-AMR-09 ::preferred # ::tok More specifically , BRAF @ V600E @ @ and HRAS @ G12V @ @ provided Caco @-@ 2 cells with highly migrating and invasive properties , some similar to those in DLD @-@ 1 cells ( Figure 3A , B ) , which is compatible with their more elongated morphology described earlier ( Figure 1 ) . # ::alignments 0-1.5.1 1-1.5 4-1.1.1.1.1 6-1.1.1.2.1 9-1.1 11-1.1.2.1.1 13-1.1.2.2.1 16-1 17-1.3.1.1 19-1.3.1.1 20-1.3 21-1.2.r 21-1.4.1.r 22-1.2.1.1 23-1.2.2 25-1.2.1 26-1.2 26-1.2.3.1 28-1.2.3.1.2 29-1.2.3 32-1.2.3.1.1.r 33-1.2.3.1.1.1.1 35-1.2.3.1.1.1.1 36-1.2.3.1.1 38-1.2.3.2.1.1 38-1.2.3.2.1.2 40-1.2.3.2.1.1.1 48-1.4 49-1.4.1.r 51-1.4.1.2.1 52-1.4.1.2 53-1.4.1 54-1.2.3.2 54-1.4.1.3 55-1.4.1.3.2 57-1.4.1.3.1 59-1.4.1.3.1.1 (p / provide-01~e.16 :ARG0 (a / and~e.9 :op1 (g / gene :name (n / name :op1 "BRAF"~e.4) :ARG2-of (m / mutate-01 :value "V600E"~e.6)) :op2 (g2 / gene :name (n2 / name :op1 "HRAS"~e.11) :ARG2-of (m2 / mutate-01 :value "G12V"~e.13))) :ARG1~e.21 (p2 / property~e.26 :mod (i / invade-01~e.25 :ARG1-of (h / high-02~e.22)) :mod (m3 / migrate-01~e.23 :degree h) :ARG1-of (r / resemble-01~e.29 :ARG2 (p3 / property~e.26 :location~e.32 (c / cell-line~e.36 :name (n3 / name :op1 "DLD-1"~e.33,35)) :quant (s / some~e.28)) :ARG1-of (d / describe-01~e.54 :ARG0 (a2 / and :op1 (f / figure~e.38 :mod "3A"~e.40) :op2 (f2 / figure~e.38 :mod "3B"))))) :ARG2 (c2 / cell-line~e.20 :name (n4 / name :op1 "Caco-2"~e.17,19)) :mod (c3 / compatible~e.48 :prep-with~e.21,49 (m4 / morphology~e.53 :poss c2 :ARG1-of (e3 / elongate-01~e.52 :degree (m5 / more~e.51)) :ARG1-of (d2 / describe-01~e.54 :ARG0 (f3 / figure~e.57 :mod 1~e.59) :time (e4 / early~e.55 :degree m5)))) :ARG1-of (s2 / specific-02~e.1 :degree m5~e.0)) # ::id a_pmid_2194_3101.168 ::amr-annotator SDL-AMR-09 ::preferred # ::tok Moreover , Caco @-@ K cells , that retained typical epithelial morphology of Caco @-@ 2 parental cells also presented enhanced migrating and invasive properties , but to a lesser extent . # ::alignments 0-1 2-1.1.1.1.1 4-1.1.1.1.1 5-1.1.1 8-1.1.1.2 9-1.1.1.2.1.2 10-1.1.1.2.1.1 11-1.1.1.2.1 13-1.1.1.2.1.3.1.1 15-1.1.1.2.1.3.1.1 16-1.1.1.2.1.3.2 17-1.1.1 17-1.1.1.2.1.3 18-1.1.3 19-1.1 19-1.1.4.1 20-1.1.2.3 21-1.1.2.2 23-1.1.2.1 24-1.1.2 26-1.1.4 27-1.1.4.1.3.r 29-1.1.4.1.3 (a / and~e.0 :op2 (p / present-01~e.19 :ARG0 (c / cell~e.5,17 :name (n / name :op1 "Caco-K"~e.2,4) :ARG0-of (r / retain-01~e.8 :ARG1 (m / morphology~e.11 :mod (e / epithelium~e.10) :ARG1-of (t / typical-02~e.9) :poss (c2 / cell-line~e.17 :name (n2 / name :op1 "Caco-2"~e.13,15) :mod (p2 / parent~e.16))))) :ARG1 (p3 / property~e.24 :mod (i / invade-01~e.23) :mod (m2 / migrate-01~e.21) :ARG1-of (e2 / enhance-01~e.20)) :mod (a2 / also~e.18) :ARG1-of (c3 / contrast-01~e.26 :ARG2 (p4 / present-01~e.19 :ARG0 c :ARG1 p3 :degree~e.27 (l / less~e.29))))) # ::id a_pmid_2194_3101.169 ::amr-annotator SDL-AMR-09 ::preferred # ::tok Taken together , morphological properties induced by either BRAF @ V600E @ @ or KRAS @ G12V @ @ oncogene affected the ability of Caco @-@ 2 cells to migrate and invade in vitro , but were not sufficient to fully reverse their epithelial phenotype . # ::alignments 0-1.3 1-1.3.1 3-1.1.1.1 4-1.1.1 5-1.1.1.2 9-1.1.1.2.1.1.1.1 11-1.1.1.2.1.1.2.1 14-1.1.1.2.1 16-1.1.1.2.1.2.1.1 18-1.1.1.2.1.2.2.1 22-1.1 26-1.1.2.1.1.1.1.1 28-1.1.2.1.1.1.1.1 29-1.1.2.1.1.1 31-1.1.2.1.1 32-1.1.2.1 33-1.1.2.1.2 35-1.1.2.1.3 36-1.1.2.1.3 39-1 41-1.2.1 41-1.2.1.r 42-1.2 44-1.2.3.3 45-1.2.3 46-1.2.3.2.2 46-1.2.3.2.2.r 47-1.2.3.2.1 48-1.2.3.2 (c / contrast-01~e.39 :ARG1 (a / affect-01~e.22 :ARG0 (p / property~e.4 :mod (m / morphological~e.3) :ARG2-of (i / induce-01~e.5 :ARG0 (o / or~e.14 :op1 (g / gene :name (n / name :op1 "BRAF"~e.9) :ARG2-of (m2 / mutate-01 :value "V600E"~e.11)) :op2 (g2 / gene :name (n2 / name :op1 "KRAS"~e.16) :ARG2-of (m3 / mutate-01 :value "G12V"~e.18)) :ARG0-of (c3 / cause-01 :ARG1 (d / disease :wiki "Cancer" :name (n4 / name :op1 "cancer")))))) :ARG1 (p2 / possible-01 :ARG1 (a2 / and~e.32 :op1 (m4 / migrate-01~e.31 :ARG0 (c2 / cell-line~e.29 :name (n3 / name :op1 "Caco-2"~e.26,28))) :op2 (i2 / invade-01~e.33 :ARG0 c2) :manner (i3 / in-vitro~e.35,36)))) :ARG2 (s / suffice-01~e.42 :polarity~e.41 -~e.41 :ARG0 p :ARG1 (r / reverse-01~e.45 :ARG0 p :ARG1 (p3 / phenotype~e.48 :mod (e / epithelium~e.47) :poss~e.46 c2~e.46) :degree (f / full~e.44))) :ARG1-of (t / take-01~e.0 :mod (t2 / together~e.1))) # ::id a_pmid_2194_3101.170 ::amr-annotator SDL-AMR-09 ::preferred # ::tok The role of BRAF @ and KRAS @ oncogenes in altering cytoskeletal properties was further emphasized following depletion of BRAF @ V600E @ by shRNA in HT29 cells , where migration ability of HT @-@ ShBR3 cells , with downregulated expression of mt @ BRAF @ gene , was significantly impaired as compared to the empty vector control HT @-@ ps cells . # ::alignments 1-1.1 4-1.1.1.1.1.1 6-1.1.1 8-1.1.1.2.1.1 11-1.1.2.r 12-1.1.2 14-1.1.2.2 16-1.2 17-1 18-1.3 19-1.3.1 20-1.3.1.2.r 21-1.3.1.2.1.1 23-1.3.1.2.2.1 25-1.3.1.1.r 26-1.3.1.1.1.1 27-1.3.1.3.r 28-1.3.1.3.1.1 29-1.3.1.3 31-1.3.1.3.2.r 32-1.3.1.3.2.1.1 34-1.3.1.3.2.1.1.1.r 35-1.3.1.3.2.1.1.1.1.1 37-1.3.1.3.2.1.1.1.1.1 38-1.3.1.3.2.1.1.1 40-1.3.1.3.2.1.2.r 41-1.3.1.3.2.1.2.2 42-1.3.1.3.2.1.2 46-1.3.1.3.2.1.2.1.1.1 48-1.1.1.1 48-1.1.1.2 48-1.3.1.3.2.1.2.1 51-1.3.1.3.2.2 52-1.3.1.3.2 53-1.3.1.3.2.3.r 54-1.3.1.3.2.3 55-1.3.1.3.2.3.1.r 57-1.3.1.3.2.3.1.2.1.1 58-1.3.1.3.2.3.1.2.1 59-1.3.1.3.2.3.1.2 60-1.3.1.3.2.3.1.1.1 62-1.3.1.3.2.3.1.1.1 63-1.3.1.3.2.3.1 (e / emphasize-01~e.17 :ARG1 (r / role~e.1 :poss (a / and~e.6 :op1 (g2 / gene~e.48 :name (n / name :op1 "BRAF"~e.4)) :op2 (g3 / gene~e.48 :name (n2 / name :op1 "KRAS"~e.8)) :ARG0-of (c7 / cause-01 :ARG1 (d3 / disease :wiki "Cancer" :name (n10 / name :op1 "cancer")))) :topic~e.11 (a2 / alter-01~e.12 :ARG0 a :ARG1 (p / property~e.14 :mod (c / cytoskeleton)))) :degree (f / further~e.16) :ARG1-of (f2 / follow-01~e.18 :ARG2 (d / deplete-01~e.19 :ARG0~e.25 (n9 / nucleic-acid :name (n3 / name :op1 "shRNA"~e.26)) :ARG1~e.20 (e2 / enzyme :name (n4 / name :op1 "BRAF"~e.21) :ARG2-of (m / mutate-01 :value "V600E"~e.23)) :location~e.27 (c2 / cell-line~e.29 :name (n5 / name :op1 "HT29"~e.28) :location-of~e.31 (i / impair-01~e.52 :ARG1 (p2 / possible-01 :ARG1 (m2 / migrate-01~e.32 :ARG0~e.34 (c3 / cell-line~e.38 :name (n6 / name :op1 "HT-ShBR3"~e.35,37))) :accompanier~e.40 (e3 / express-03~e.42 :ARG1 (g / gene~e.48 :name (n7 / name :op1 "BRAF"~e.46) :ARG2-of (m3 / mutate-01)) :ARG1-of (d2 / downregulate-01~e.41))) :ARG1-of (s / significant-02~e.51) :ARG1-of~e.53 (c4 / compare-01~e.54 :ARG2~e.55 (c5 / cell-line~e.63 :name (n8 / name :op1 "HT-ps"~e.60,62) :ARG2-of (c6 / control-01~e.59 :ARG0 (v / vector~e.58 :ARG1-of (e4 / empty-02~e.57)))))))))) # ::id a_pmid_2194_3101.171 ::amr-annotator SDL-AMR-09 ::preferred # ::tok Likewise , knock out of KRAS @ G13D @ in DLD @-@ 1 cells ( DKO @-@ 4 ) [ @ 28 @ ] significantly reverted the migration ability of DLD @-@ 1 cells ( Figure 3C ) . # ::alignments 0-1.4 2-1.1 3-1.1 4-1.1.1.r 5-1.1.1.1.1 7-1.1.1.2.1 9-1.1.2.r 10-1.1.2 11-1.1.2 12-1.1.2 13-1.1.3.1 13-1.2.1.1 15-1.1.3.1.1.1 17-1.1.3.1.1.1 21-1.1.4.1.1.1 24-1.3 25-1 27-1.2.1 30-1.2.1.1.1.1 32-1.2.1.1.1.1 33-1.2.1.1 35-1.5.1 37-1.5.1.1 (r / revert-01~e.25 :ARG0 (k / knock-out-03~e.2,3 :ARG1~e.4 (e / enzyme :name (n2 / name :op1 "KRAS"~e.5) :ARG2-of (m2 / mutate-01 :value "G13D"~e.7)) :location~e.9 c~e.10,11,12 :ARG1-of (d / describe-01 :ARG0 (c2 / cell-line~e.13 :name (n3 / name :op1 "DKO-4"~e.15,17))) :ARG1-of (d2 / describe-01 :ARG0 (p2 / publication :ARG1-of (c3 / cite-01 :ARG2 28~e.21)))) :ARG1 (p / possible-01 :ARG1 (m / migrate-01~e.27 :ARG0 (c / cell-line~e.13,33 :name (n / name :op1 "DLD-1"~e.30,32)))) :ARG1-of (s / significant-02~e.24) :mod (l / likewise~e.0) :ARG1-of (d3 / describe-01 :ARG0 (f / figure~e.35 :mod "3C"~e.37))) # ::id a_pmid_2194_3101.172 ::amr-annotator SDL-AMR-09 ::preferred # ::tok BRAF @ V600E @ enhances the ability of Caco @-@ 2 cells to migrate and invade in vitro @ through RhoA activation # ::alignments 0-1.1.1.1 2-1.1.2.1 4-1 8-1.2.1.1.1.1.1 10-1.2.1.1.1.1.1 11-1.2.1.1.1 13-1.2.1.1 14-1.2.1 15-1.2.1.2 17-1.2.1.3 18-1.2.1.3 21-1.3.1.1.1 22-1.3 (e / enhance-01~e.4 :ARG0 (e2 / enzyme :name (n / name :op1 "BRAF"~e.0) :ARG2-of (m / mutate-01 :value "V600E"~e.2)) :ARG1 (p / possible-01 :ARG1 (a / and~e.14 :op1 (m2 / migrate-01~e.13 :ARG0 (c / cell-line~e.11 :name (n2 / name :op1 "Caco-2"~e.8,10))) :op2 (i / invade-01~e.15 :ARG0 c) :manner (i2 / in-vitro~e.17,18))) :manner (a2 / activate-01~e.22 :ARG1 (p2 / protein :name (n3 / name :op1 "RhoA"~e.21)))) # ::id a_pmid_2194_3101.173 ::amr-annotator SDL-AMR-09 ::preferred # ::tok Overexpression of BRAF @ V600E @ in Caco @-@ 2 cells had a profound effect on the RAS effector protein RhoA ( Figure 4 ) . # ::alignments 0-1.1 1-1.1.1.r 2-1.1.1.1.1 4-1.1.1.2.1 6-1.1.2.r 7-1.1.2.1.1 9-1.1.2.1.1 10-1.1.2 13-1.3 14-1 15-1.2.r 17-1.2.2.1.1.1 18-1.2.2 19-1.2 19-1.2.2.1 20-1.2.1.1 22-1.4.1 24-1.4.1.1 (a / affect-01~e.14 :ARG0 (o / overexpress-01~e.0 :ARG1~e.1 (e / enzyme :name (n / name :op1 "BRAF"~e.2) :ARG2-of (m / mutate-01 :value "V600E"~e.4)) :location~e.6 (c / cell-line~e.10 :name (n2 / name :op1 "Caco-2"~e.7,9))) :ARG1~e.15 (p / protein~e.19 :name (n3 / name :op1 "RhoA"~e.20) :mod (e2 / effector~e.18 :mod (p3 / protein-family~e.19 :name (n4 / name :op1 "RAS"~e.17)))) :degree (p2 / profound~e.13) :ARG1-of (d / describe-01 :ARG0 (f / figure~e.22 :mod 4~e.24))) # ::id a_pmid_2194_3101.174 ::amr-annotator SDL-AMR-09 ::preferred # ::tok In Caco @-@ BR cells activation of RhoA is increased ( Figure 4A ) as well as phosphorylation of its downstream target Cofilin , a protein that is related to stress fibre formation ( Figure 4B ) . # ::alignments 1-1.1.2.1.2.1.1.1 3-1.1.2.1.2.1.1.1 4-1.1.2.1.2.1 5-1.1.1 7-1.1.1.1.1.1 9-1 11-1.1.1.2.1 13-1.1.1.2.1.1 16-1.1 17-1.1 18-1.1 19-1.1.2 22-1.1.2.1.2.2 23-1.1.2.1.2 24-1.1.2.1.1.1 27-1.1.1.1 27-1.1.2.1 30-1.1.2.1.3 31-1.1.2.1.3.1.r 32-1.1.2.1.3.1.1.1 34-1.1.2.1.3.1 36-1.1.2.2.1 38-1.1.2.2.1.1 (i / increase-01~e.9 :ARG1 (a / and~e.16,17,18 :op1 (a2 / activate-01~e.5 :ARG1 (p / protein~e.27 :name (n / name :op1 "RhoA"~e.7)) :ARG1-of (d / describe-01 :ARG0 (f / figure~e.11 :mod "4A"~e.13))) :op2 (p2 / phosphorylate-01~e.19 :ARG1 (p3 / protein~e.27 :name (n2 / name :op1 "Cofilin"~e.24) :ARG1-of (t / target-01~e.23 :ARG0 (c / cell-line~e.4 :name (n3 / name :op1 "Caco-BR"~e.1,3)) :location (d2 / downstream~e.22)) :ARG1-of (r / relate-01~e.30 :ARG2~e.31 (f2 / form-01~e.34 :ARG1 (f3 / fiber :mod (s / stress~e.32))))) :ARG1-of (d3 / describe-01 :ARG0 (f4 / figure~e.36 :mod "4B"~e.38)))) :location c) # ::id a_pmid_2194_3101.175 ::amr-annotator SDL-AMR-09 ::preferred # ::tok These findings are closely related to the observation regarding increased stress fibre formation indicated by phalloidin staining in Caco @-@ BR13 cells ( Figure 1 ) . # ::alignments 0-1.1.2 1-1.1 1-1.1.1 1-1.1.1.r 3-1.3 4-1 5-1.2.r 7-1.2 8-1.2.1 9-1.2.1.1.2 10-1.2.1.1.1.1 12-1.2.1.1 13-1.2.1.1.3 14-1.2.1.1.3.1.r 15-1.2.1.1.3.1.1.1.1 16-1.2.1.1.3.1 17-1.2.1.1.3.1.2.r 18-1.2.1.1.3.1.2.1.1 20-1.2.1.1.3.1.2.1.1 21-1.2.1.1.3.1.2 23-1.4.1 25-1.4.1.1 (r / relate-01~e.4 :ARG1 (t / thing~e.1 :ARG1-of~e.1 (f / find-01~e.1) :mod (t2 / this~e.0)) :ARG2~e.5 (o / observe-01~e.7 :ARG0-of (r2 / regard-01~e.8 :ARG1 (f2 / form-01~e.12 :ARG1 (f3 / fiber :mod (s / stress~e.10)) :ARG1-of (i / increase-01~e.9) :ARG1-of (i2 / indicate-01~e.13 :ARG0~e.14 (s2 / stain-01~e.16 :ARG2 (s3 / small-molecule :name (n / name :op1 "phalloidin"~e.15)) :location~e.17 (c / cell-line~e.21 :name (n2 / name :op1 "Caco-BR13"~e.18,20))))))) :ARG1-of (c2 / close-10~e.3) :ARG1-of (d / describe-01 :ARG0 (f4 / figure~e.23 :mod 1~e.25))) # ::id a_pmid_2194_3101.176 ::amr-annotator SDL-AMR-09 ::preferred # ::tok Notably , an extra band of lower molecular weight is detected for RhoA in Caco @-@ BR and DLD @-@ 1 cells , which potentially represents the main active GTPase form ( Figure 4A ) . # ::alignments 0-1.3 3-1.1.1 4-1.1 5-1.1.2.r 6-1.1.2.2 6-1.1.2.2.1 6-1.1.2.2.1.r 7-1.1.2.1 8-1.1.2 10-1 11-1.1.3.r 12-1.1.3.1.1 13-1.2.r 14-1.2.1.1.1 16-1.2.1.1.1 17-1.2 18-1.2.2.1.1 20-1.2.2.1.1 21-1.2.1 21-1.2.2 24-1.1.4.2 25-1.1.4 27-1.1.4.1.3 29-1.1.4.1.1.1.1 30-1.1.4.1 32-1.4.1 34-1.4.1.1 (d / detect-01~e.10 :ARG1 (b / band~e.4 :mod (e / extra~e.3) :ARG1-of~e.5 (w / weight-01~e.8 :mod (m / molecule~e.7) :ARG1-of (l / low-04~e.6 :degree~e.6 (m2 / more~e.6))) :mod~e.11 (p / protein :name (n / name :op1 "RhoA"~e.12)) :ARG0-of (r / represent-01~e.25 :ARG1 (f / form~e.30 :mod (e2 / enzyme :name (n4 / name :op1 "GTPase"~e.29)) :ARG1-of (a / activate-01) :mod (m3 / main~e.27)) :mod (p2 / potential~e.24))) :location~e.13 (a2 / and~e.17 :op1 (c / cell-line~e.21 :name (n2 / name :op1 "Caco-BR"~e.14,16)) :op2 (c2 / cell-line~e.21 :name (n3 / name :op1 "DLD-1"~e.18,20))) :ARG1-of (n5 / notable-04~e.0) :ARG1-of (d2 / describe-01 :ARG0 (f2 / figure~e.32 :mod "4A"~e.34))) # ::id a_pmid_2194_3101.177 ::amr-annotator SDL-AMR-09 ::preferred # ::tok A variant of lower molecular weight for RhoA protein has previously been reported both in colon and breast tissues [ @ 7 @ ] . # ::alignments 1-1.1 2-1.1.1.r 3-1.1.1.2 3-1.1.1.2.1 3-1.1.1.2.1.r 4-1.1.1.1 5-1.1.1 6-1.1.2.r 7-1.1.2.1.1 8-1.1.2 10-1.2 12-1 14-1.3.r 15-1.3.1.1 16-1.3 17-1.3.2.1 18-1.3.1 18-1.3.2 21-1.4.1.1.1 (r / report-01~e.12 :ARG1 (v / variant~e.1 :ARG1-of~e.2 (w / weight-01~e.5 :mod (m / molecule~e.4) :ARG1-of (l / low-04~e.3 :degree~e.3 (m2 / more~e.3))) :poss~e.6 (p / protein~e.8 :name (n / name :op1 "RhoA"~e.7))) :time (p2 / previous~e.10) :location~e.14 (a / and~e.16 :op1 (t / tissue~e.18 :part-of (c / colon~e.15)) :op2 (t2 / tissue~e.18 :part-of (b / breast~e.17))) :ARG1-of (d / describe-01 :ARG0 (p3 / publication :ARG1-of (c2 / cite-01 :ARG2 7~e.21)))) # ::id a_pmid_2194_3101.178 ::amr-annotator SDL-AMR-09 ::preferred # ::tok However , RT @-@ PCR analysis and treatment with the proteasome inhibitor MG @-@ 132 , both in Caco @-@ BR and DLD @-@ 1 cells , suggested no association of this faster migrating RhoA band with alternative splicing or proteasomal degradation ( data not shown ) . # ::alignments 0-1 2-1.1.1.1.1.1.1 4-1.1.1.1.1.1.1 5-1.1.1.1 6-1.1.1 6-1.1.1.3 6-1.1.1.3.r 7-1.1.1.2 8-1.1.1.1.1.r 8-1.1.1.2.1.r 10-1.1.1.2.1.2.1.1.1 11-1.1.1.2.1 11-1.1.1.2.1.2 11-1.1.1.2.1.2.r 12-1.1.1.2.1.1.1 14-1.1.1.2.1.1.1 18-1.1.1.3.1.1.1 20-1.1.1.3.1.1.1 21-1.1.1.3 22-1.1.1.3.2.1.1 24-1.1.1.3.2.1.1 25-1.1.1.3.1 25-1.1.1.3.2 27-1.1 28-1.1.2.1 28-1.1.2.1.r 29-1.1.2 30-1.1.2.2.r 31-1.1.2.2.3 32-1.1.2.2.2.1 32-1.1.2.2.2.1.1 32-1.1.2.2.2.1.1.r 33-1.1.2.2.2 34-1.1.2.2.1.1.1 35-1.1.2.2 36-1.1.1.1.1.r 38-1.1.2.3.1 39-1.1.2.3 41-1.1.2.3.2 43-1.2.1 44-1.2.1.1.1 44-1.2.1.1.1.r 45-1.2.1.1 (h / have-concession-91~e.0 :ARG1 (s / suggest-01~e.27 :ARG0 (a / and~e.6 :op1 (a2 / analyze-01~e.5 :instrument~e.8,36 (t3 / thing :name (n / name :op1 "RT-PCR"~e.2,4))) :op2 (t / treat-04~e.7 :ARG2~e.8 (s2 / small-molecule~e.11 :name (n2 / name :op1 "MG-132"~e.12,14) :ARG0-of~e.11 (i / inhibit-01~e.11 :ARG1 (e / enzyme :name (n3 / name :op1 "proteasome"~e.10))))) :location~e.6 (a3 / and~e.6,21 :op1 (c / cell-line~e.25 :name (n4 / name :op1 "Caco-BR"~e.18,20)) :op2 (c2 / cell-line~e.25 :name (n5 / name :op1 "DLD-1"~e.22,24)))) :ARG1 (a4 / associate-01~e.29 :polarity~e.28 -~e.28 :ARG1~e.30 (b / band~e.35 :mod (p2 / protein :name (n6 / name :op1 "RhoA"~e.34)) :ARG0-of (m / migrate-01~e.33 :ARG1-of (f / fast-02~e.32 :degree~e.32 (m2 / more~e.32))) :mod (t2 / this~e.31)) :ARG2 (o / or~e.39 :op1 (s3 / splice-01~e.38 :ARG1-of (a5 / alternate-01)) :op2 (d / degrade-01~e.41 :ARG1 e)))) :ARG1-of (d2 / describe-01 :ARG0 (d3 / data~e.43 :ARG1-of (s4 / show-01~e.45 :polarity~e.44 -~e.44)))) # ::id a_pmid_2194_3101.179 ::amr-annotator SDL-AMR-09 ::preferred # ::tok These data suggested that the additional band potentially represents a post @-@ translational modification of RhoA protein . # ::alignments 0-1.1.1 1-1.1 2-1 6-1.2.1 7-1.2.3 8-1.2 10-1.2.2.2 12-1.2.2.2.1 13-1.2.2 14-1.2.2.1.r 15-1.2.2.1.1.1 16-1.2.2.1 (s / suggest-01~e.2 :ARG0 (d / data~e.1 :mod (t / this~e.0)) :ARG1 (r / represent-01~e.8 :ARG0 (b / band~e.6 :ARG1-of (a / add-02)) :ARG1 (m / modify-01~e.13 :ARG1~e.14 (p / protein~e.16 :name (n / name :op1 "RhoA"~e.15)) :time (a2 / after~e.10 :op1 (t2 / translate-02~e.12))) :mod (p3 / potential~e.7))) # ::id a_pmid_2194_3101.180 ::amr-annotator SDL-AMR-09 ::preferred # ::tok To further explore the role of BRAF @ V600E @ in the activation of the RhoA pathway , transient transfection of the oncogene in Caco @-@ 2 cells was performed ( Additional Figure 2 ) . # ::alignments 1-1.2.2 2-1.2 4-1.2.1 5-1.2.1.1.r 6-1.2.1.1.1.1 8-1.2.1.1.2.1 10-1.2.1.2.r 12-1.2.1.2 13-1.2.1.2.1.r 15-1.2.1.2.1.1.1 16-1.2.1.2.1 18-1.1.3 19-1.1 20-1.1.2.r 22-1.1.2 23-1.1.1.r 24-1.1.1.1.1 26-1.1.1.1.1 27-1.1.1 29-1 32-1.3.1 34-1.3.1.1 (p / perform-02~e.29 :ARG1 (t / transfect-01~e.19 :ARG1~e.23 (c / cell-line~e.27 :name (n / name :op1 "Caco-2"~e.24,26)) :ARG2~e.20 (o / oncogene~e.22) :ARG1-of (t2 / transient-02~e.18)) :purpose (e / explore-01~e.2 :ARG1 (r / role~e.4 :poss~e.5 (e2 / enzyme :name (n2 / name :op1 "BRAF"~e.6) :ARG2-of (m / mutate-01 :value "V600E"~e.8)) :topic~e.10 (a / activate-01~e.12 :ARG1~e.13 (p2 / pathway~e.16 :name (n3 / name :op1 "RhoA"~e.15)))) :degree (f / further~e.1)) :ARG1-of (d / describe-01 :ARG0 (f2 / figure~e.32 :mod 2~e.34 :ARG1-of (a2 / add-02)))) # ::id a_pmid_2194_3101.181 ::amr-annotator SDL-AMR-09 ::preferred # ::tok Subsequent analysis of the migration and invasion properties showed that moderate RhoA activation induced a partial cell migration and cell invasion response ( Addition Figure 2A , B ) . # ::alignments 0-1.1.2 0-1.1.2.r 1-1.1 2-1.1.1.r 4-1.1.1.2 6-1.1.1.1 7-1.1.1 8-1 9-1.2.r 10-1.2.1.2 11-1.2.1.1.1.1 12-1.2.1 13-1.2 15-1.2.2.3 15-1.2.2.3.r 16-1.2.2.1.1.1 17-1.2.2.1.1 19-1.2.2.1.1.1 20-1.2.2.2.1 21-1.2.2.1 21-1.2.2.2 23-1.3.1.3 24-1.3.1.1 24-1.3.1.2 26-1.3.1.1.1 (s / show-01~e.8 :ARG0 (a / analyze-01~e.1 :ARG1~e.2 (p / property~e.7 :mod (i / invade-01~e.6) :mod (m / migrate-01~e.4)) :time~e.0 (s2 / subsequent~e.0)) :ARG1~e.9 (i2 / induce-01~e.13 :ARG0 (a2 / activate-01~e.12 :ARG1 (p2 / protein :name (n / name :op1 "RhoA"~e.11)) :ARG1-of (m2 / moderate-03~e.10)) :ARG2 (a3 / and :op1 (r / respond-01~e.21 :ARG2 (m3 / migrate-01~e.17 :ARG0 (c / cell~e.16,19))) :op2 (r2 / respond-01~e.21 :ARG2 (i3 / invade-01~e.20 :ARG0 c)) :degree~e.15 (p3 / part~e.15))) :ARG1-of (d / describe-01 :ARG0 (a4 / and :op1 (f / figure~e.24 :mod "2A"~e.26) :op2 (f2 / figure~e.24 :mod "2B") :ARG1-of (a5 / add-02~e.23)))) # ::id a_pmid_2194_3101.182 ::amr-annotator SDL-AMR-09 ::preferred # ::tok Notably in the invasion assay cell phenotype became slightly altered and resembled that of the stable Caco @-@ BR clones ( Additional Figure 2C ) , suggesting that a stable expression of BRAF @ V600E @ is required to achieve complete cell transformation and extensive RhoA activation . # ::alignments 0-1.3 3-1.4.1 4-1.4 5-1.2.2.1.1 6-1.1.1 6-1.2.2 7-1.1 8-1.1.2.2 9-1.1.2 10-1 11-1.2 15-1.2.2.1.2 16-1.2.2.1.1.1.1 18-1.2.2.1.1.1.1 19-1.2.2.1 22-1.5.1 24-1.5.1.1 28-1.6 29-1.6.1.r 31-1.6.1.2.2 32-1.6.1.2 33-1.6.1.2.1.r 34-1.6.1.2.1.1.1 36-1.6.1.2.1.2.1 39-1.6.1 41-1.6.1.1 42-1.6.1.1.1.1.2 43-1.1.1.1 44-1.6.1.1.1.1 45-1.6.1.1.1 46-1.6.1.1.1.2.2 47-1.6.1.1.1.2.1.1.1 48-1.6.1.1.1.2 (a / and~e.10 :op1 (b / become-01~e.7 :ARG1 (p / phenotype~e.6 :mod (c / cell~e.43)) :ARG2 (a2 / alter-01~e.9 :ARG1 p :degree (s / slight~e.8))) :op2 (r / resemble-01~e.11 :ARG1 p :ARG2 (p2 / phenotype~e.6 :poss (c2 / clone-01~e.19 :ARG1 (c3 / cell-line~e.5 :name (n / name :op1 "Caco-BR"~e.16,18)) :ARG1-of (s2 / stable-03~e.15)))) :ARG1-of (n2 / notable-04~e.0) :time (a3 / assay-01~e.4 :ARG1 (i / invade-01~e.3)) :ARG1-of (d / describe-01 :ARG0 (f / figure~e.22 :mod "2C"~e.24 :ARG1-of (a4 / add-02))) :ARG0-of (s3 / suggest-01~e.28 :ARG1~e.29 (r2 / require-01~e.39 :ARG0 (a5 / achieve-01~e.41 :ARG1 (a6 / and~e.45 :op1 (t / transform-01~e.44 :ARG1 c :ARG1-of (c4 / complete-02~e.42)) :op2 (a7 / activate-01~e.48 :ARG1 (p3 / protein :name (n3 / name :op1 "RhoA"~e.47)) :ARG1-of (e / extensive-03~e.46)))) :ARG1 (e2 / express-03~e.32 :ARG2~e.33 (e3 / enzyme :name (n4 / name :op1 "BRAF"~e.34) :ARG2-of (m / mutate-01 :value "V600E"~e.36)) :ARG1-of s2~e.31)))) # ::id a_pmid_2194_3101.183 ::amr-annotator SDL-AMR-09 ::preferred # ::tok Regarding the importance of RhoA activation in the induced cell migration and invasion observed in Caco @-@ BR cells , siRNA against RhoA was performed leading to significant protein depletion in both Caco @-@ 2 and Caco @-@ BR13 cells ( Figure 5A ) . # ::alignments 0-1.2.r 2-1.2 4-1.2.1.1 5-1.2.1 6-1.2.1.2.r 8-1.2.1.2.3 9-1.2.1.2.4.1 9-1.3.1.3.2 10-1.2.1.2.1 11-1.2.1.2 12-1.2.1.2.2 13-1.2.1.2.4 15-1.2.1.2.4.1.1.1 17-1.2.1.2.4.1.1.1 18-1.2.1.2.1.1 20-1.1.1.1 21-1.1 21-1.1.2 21-1.1.2.r 22-1.1.2.1.1.1 23-1.2.1.r 24-1 25-1.3 26-1.3.1.r 27-1.3.1.2 28-1.3.1.1 29-1.3.1 32-1.3.1.3.1.1.1 34-1.3.1.3.1.1.1 35-1.3.1.3 36-1.3.1.3.1.1.1 36-1.3.1.3.2.1.1 38-1.3.1.3.2.1.1 39-1.3.1.3.1 41-1.4.1 43-1.4.1.1 (p / perform-02~e.24 :ARG1 (n6 / nucleic-acid~e.21 :name (n / name :op1 "siRNA"~e.20) :ARG0-of~e.21 (o / oppose-01~e.21 :ARG1 (p2 / protein :name (n2 / name :op1 "RhoA"~e.22)))) :topic~e.0 (i / important~e.2 :domain~e.23 (a / activate-01~e.5 :ARG1 p2~e.4 :location~e.6 (a2 / and~e.11 :op1 (m / migrate-01~e.10 :ARG0 (c / cell~e.18)) :op2 (i2 / invade-01~e.12 :ARG0 c) :ARG2-of (i3 / induce-01~e.8) :ARG1-of (o2 / observe-01~e.13 :location (c2 / cell-line~e.9 :name (n3 / name :op1 "Caco-BR"~e.15,17)))))) :ARG0-of (l / lead-03~e.25 :ARG2~e.26 (d / deplete-01~e.29 :ARG1 (p3 / protein~e.28) :ARG1-of (s / significant-02~e.27) :location (a3 / and~e.35 :op1 (c3 / cell-line~e.39 :name (n4 / name :op1 "Caco-2"~e.32,34,36)) :op2 (c4 / cell-line~e.9 :name (n5 / name :op1 "Caco-BR13"~e.36,38))))) :ARG1-of (d2 / describe-01 :ARG0 (f / figure~e.41 :mod "5A"~e.43))) # ::id a_pmid_2194_3101.184 ::amr-annotator SDL-AMR-09 ::preferred # ::tok Depletion of RhoA substantially impaired both acquired properties with more profound effect in Caco @-@ BR13 cells , further illustrating its central role in the BRAF @ V600E @ oncogene @-@ induced transformation of colon adenocarcinoma cells ( Figure 5B ) . # ::alignments 0-1.1 1-1.1.1.r 2-1.1.1.1.1 3-1.3 4-1 5-1.2.2 6-1.2.1 7-1.2 8-1.4.r 9-1.4.1.1 10-1.4.1 11-1.4 12-1.4.2.r 13-1.4.2.1.1 15-1.4.2.1.1 16-1.4.2 18-1.5.2 19-1.5 20-1.5.1.1 20-1.5.1.1.r 21-1.5.1.3 22-1.5.1 23-1.5.1.2.r 25-1.5.1.2.2.1.1.1 27-1.5.1.2.2.1.2.1 31-1.5.1.2.2 32-1.5.1.2 33-1.5.1.2.1.r 34-1.5.1.2.1.1.1 35-1.5.1.2.1.1 36-1.5.1.2.1 38-1.6.1 40-1.6.1.1 (i / impair-01~e.4 :ARG0 (d / deplete-01~e.0 :ARG1~e.1 (p / protein :name (n / name :op1 "RhoA"~e.2))) :ARG1 (p2 / property~e.7 :ARG1-of (a / acquire-01~e.6) :mod (b / both~e.5)) :degree (s / substantial~e.3) :ARG0-of~e.8 (a2 / affect-01~e.11 :degree (p3 / profound~e.10 :degree (m / more~e.9)) :location~e.12 (c / cell-line~e.16 :name (n2 / name :op1 "Caco-BR13"~e.13,15))) :ARG0-of (i2 / illustrate-01~e.19 :ARG1 (r / role~e.22 :poss~e.20 d~e.20 :topic~e.23 (t / transform-01~e.32 :ARG1~e.33 (c2 / cell~e.36 :mod (a3 / adenocarcinoma~e.35 :mod (c3 / colon~e.34))) :ARG2-of (i3 / induce-01~e.31 :ARG0 (g / gene :name (n3 / name :op1 "BRAF"~e.25) :ARG2-of (m2 / mutate-01 :value "V600E"~e.27) :ARG0-of (c5 / cause-01 :ARG1 (d3 / disease :wiki "Cancer" :name (n4 / name :op1 "cancer")))))) :mod (c4 / central~e.21)) :mod (f / further~e.18)) :ARG1-of (d2 / describe-01 :ARG0 (f2 / figure~e.38 :mod "5B"~e.40))) # ::id a_pmid_2194_3101.185 ::amr-annotator SDL-AMR-09 ::preferred # ::tok Moreover , following RhoA depletion in Caco @-@ 2 cells , the number and size of stress fibres were notably reduced as compared to Caco @-@ BR cells , where no such alteration was observed ( data not shown ) . # ::alignments 0-1.1.1 2-1.1.4 3-1.1.4.1.1.1.1 4-1.1.4.1 5-1.1.4.1.2.r 6-1.1.4.1.2.1.1 8-1.1.4.1.2.1.1 9-1.1.4.1.2 12-1.1.1.1 13-1.1.1 14-1.1.1.2 16-1.1.1.3.1 19-1.1.2 20-1.1 22-1.1.3.r 24-1.1.3.1.1 26-1.1.3.1.1 27-1.1.3 29-1.1.3.2.r 30-1.1.3.2.1.1 30-1.1.3.2.1.1.r 31-1.1.3.2.1.2 32-1.1.3.2.1 34-1.1.3.2 36-1.2.1 37-1.2.1.1.1 37-1.2.1.1.1.r 38-1.2.1.1 (a / and :op2 (r / reduce-01~e.20 :ARG1 (a2 / and~e.0,13 :op1 (n / number~e.12) :op2 (s / size~e.14) :quant-of (f / fiber :mod (s2 / stress~e.16))) :ARG2 (n3 / notable-04~e.19) :compared-to~e.22 (c / cell-line~e.27 :name (n2 / name :op1 "Caco-BR"~e.24,26) :location-of~e.29 (o / observe-01~e.34 :ARG1 (a3 / alter-01~e.32 :polarity~e.30 -~e.30 :mod (s3 / such~e.31)))) :ARG1-of (f2 / follow-01~e.2 :ARG2 (d / deplete-01~e.4 :ARG1 (p / protein :name (n4 / name :op1 "RhoA"~e.3)) :location~e.5 (c2 / cell-line~e.9 :name (n5 / name :op1 "Caco-2"~e.6,8))))) :ARG1-of (d2 / describe-01 :ARG0 (d3 / data~e.36 :ARG1-of (s4 / show-01~e.38 :polarity~e.37 -~e.37)))) # ::id a_pmid_2194_3101.186 ::amr-annotator SDL-AMR-09 ::preferred # ::tok In order to study further the impact of RhoA GTPase on cell migration , silencing of RhoA was performed in DLD @-@ 1 and HT29 cells . # ::alignments 0-1.3.r 1-1.3.r 2-1.3.r 3-1.3 4-1.3.2 6-1.3.1 7-1.3.1.1.r 8-1.3.1.1.1.1 9-1.3.1.1.1.2 10-1.3.1.2.r 11-1.3.1.2.1 12-1.3.1.2 14-1.1 15-1.1.1.r 16-1.1.1.1.1 18-1 19-1.2.r 20-1.2.1.1.1 22-1.2.1.1.1 23-1.2 24-1.2.2.1.1 25-1.2.1 25-1.2.2 (p / perform-02~e.18 :ARG1 (s / silence-01~e.14 :ARG1~e.15 (p2 / protein :name (n / name :op1 "RhoA"~e.16))) :location~e.19 (a / and~e.23 :op1 (c / cell-line~e.25 :name (n2 / name :op1 "DLD-1"~e.20,22)) :op2 (c2 / cell-line~e.25 :name (n3 / name :op1 "HT29"~e.24))) :purpose~e.0,1,2 (s2 / study-01~e.3 :ARG1 (i / impact-01~e.6 :ARG0~e.7 (p3 / pathway :name (n4 / name :op1 "RhoA"~e.8 :op2 "GTPase"~e.9)) :ARG1~e.10 (m / migrate-01~e.12 :ARG0 (c3 / cell~e.11))) :degree (f / further~e.4))) # ::id a_pmid_2194_3101.187 ::amr-annotator SDL-AMR-09 ::preferred # ::tok Considering that these cell lines bear mutation in KRAS @ G13D @ and BRAF @ V600E @ respectively , RhoA depletion was also performed in selected clones where KRAS @ G13D @ ( DKO4 ) or BRAF @ V600E @ ( HTshBR3 ) was knocked out or down regulated via shRNA respectively . # ::alignments 0-1.4 1-1.4.1.r 2-1.4.1.1.1 3-1.4.1.1 4-1.4.1.1 5-1.4.1 6-1.3.1.1 6-1.3.1.1.2 6-1.3.1.1.2.r 6-1.3.2.1 6-1.3.2.1.2 6-1.3.2.1.2.r 6-1.4.1.2 8-1.3.1.1.1.1 10-1.3.1.1.2.1 13-1.3.2.1.1.1 15-1.3.2.1.2.1 17-1.3.3.2 19-1.1.1.1.1 20-1.1 22-1.2 23-1 25-1.3.1.2 26-1.3.1 26-1.3.2 27-1.3.r 28-1.3.1.1.1.1 30-1.3.1.1.2.1 33-1.3.1.1.3.1.1.1 35-1.3 36-1.3.2.1.1.1 38-1.3.2.1.2.1 41-1.3.2.1.3.1.1.1 44-1.3.3 45-1.3.3 46-1.3 50-1.3.3.1.1.1 51-1.3.3.2 (p / perform-02~e.23 :ARG1 (d / deplete-01~e.20 :ARG1 (p2 / protein :name (n / name :op1 "RhoA"~e.19))) :mod (a / also~e.22) :location~e.27 (o / or~e.35,46 :op1 (c / clone-01~e.26 :ARG1 (e / enzyme~e.6 :name (n2 / name :op1 "KRAS"~e.8,28) :ARG2-of~e.6 (m / mutate-01~e.6 :value "G13D"~e.10,30) :ARG1-of (d2 / describe-01 :ARG0 (c2 / cell :name (n3 / name :op1 "DKO4"~e.33)))) :ARG1-of (s / select-01~e.25)) :op2 (c3 / clone-01~e.26 :ARG1 (e2 / enzyme~e.6 :name (n4 / name :op1 "BRAF"~e.13,36) :ARG2-of~e.6 (m2 / mutate-01~e.6 :value "V600E"~e.15,38) :ARG1-of (d3 / describe-01 :ARG0 (c4 / cell :name (n5 / name :op1 "HTshBR3"~e.41)))) :ARG1-of s) :ARG1-of (k / knock-out-03~e.44,45 :ARG2 (n7 / nucleic-acid :name (n6 / name :op1 "shRNA"~e.50)) :mod (r2 / respective~e.17,51)) :ARG1-of (d4 / downregulate-01 :mod r2 :instrument n7)) :ARG2-of (c5 / consider-02~e.0 :ARG1~e.1 (b / bear-01~e.5 :ARG0 (c6 / cell-line~e.3,4 :mod (t / this~e.2)) :ARG1 (m3 / mutate-01~e.6) :location (a2 / and :op1 e :op2 e2 :mod r2)))) # ::id a_pmid_2194_3101.188 ::amr-annotator SDL-AMR-09 ::preferred # ::tok This approach can implement the connection between each oncogene and the small GTPase . # ::alignments 0-1.1.1.1 1-1.1.1 2-1 3-1.1 5-1.1.2 7-1.1.2.1.1 8-1.1.2.1 11-1.1.2.2.2 12-1.1.2.2.1.1 (p / possible-01~e.2 :ARG1 (i / implement-01~e.3 :ARG0 (a / approach-02~e.1 :mod (t / this~e.0)) :ARG1 (c / connect-01~e.5 :ARG1 (o / oncogene~e.8 :mod (e / each~e.7)) :ARG1 (p2 / protein-family :name (n / name :op1 "GTPase"~e.12) :mod (s / small~e.11))))) # ::id a_pmid_2194_3101.189 ::amr-annotator SDL-AMR-09 ::preferred # ::tok After silencing of RhoA , cell migration was significantly reduced in DLD @-@ 1 , while no reduction was observed in DKO4 cells , where mutant KRAS @ G13D @ is knocked out , ( Figure 5C ) . # ::alignments 0-1.3 1-1.3.1 2-1.3.1.1.r 3-1.3.1.1.1.1 5-1.1.1.1 6-1.1.1 8-1.1.2 9-1.1 10-1.1.3.r 11-1.1.3.1.1 13-1.1.3.1.1 15-1 15-1.3.r 16-1.2.1.1 16-1.2.1.1.r 17-1.2.1 19-1.2 20-1.2.2.r 21-1.2.2.1.1 22-1.2.2 24-1.2.2.2.r 25-1.2.2.2.1 25-1.2.2.2.1.2 25-1.2.2.2.1.2.r 26-1.2.2.2.1.1.1 28-1.2.2.2.1.2.1 31-1.2.2.2 32-1.2.2.2 35-1.4.1 37-1.4.1.1 (c / contrast-01~e.15 :ARG1 (r / reduce-01~e.9 :ARG1 (m / migrate-01~e.6 :ARG0 (c2 / cell~e.5)) :ARG2 (s / significant-02~e.8) :location~e.10 (c3 / cell-line :name (n / name :op1 "DLD-1"~e.11,13))) :ARG2 (o / observe-01~e.19 :ARG1 (r2 / reduce-01~e.17 :polarity~e.16 -~e.16) :location~e.20 (c4 / cell-line~e.22 :name (n2 / name :op1 "DKO4"~e.21) :location-of~e.24 (k / knock-out-03~e.31,32 :ARG1 (e / enzyme~e.25 :name (n3 / name :op1 "KRAS"~e.26) :ARG2-of~e.25 (m2 / mutate-01~e.25 :value "G13D"~e.28))))) :time~e.15 (a / after~e.0 :op1 (s2 / silence-01~e.1 :ARG1~e.2 (p / protein :name (n4 / name :op1 "RhoA"~e.3)))) :ARG1-of (d / describe-01 :ARG0 (f / figure~e.35 :mod "5C"~e.37))) # ::id a_pmid_2194_3101.190 ::amr-annotator SDL-AMR-09 ::preferred # ::tok Depletion of RhoA in HTshBR3 ( transfected with shRNA pSUPER BRAF @ V600E ) cells with suppressed BRAF @ V600E @ activity did not reverse the ability of HT29 cell to migrate , while in HTps ( HT29 cells transfected with empty vector pSUPER ) a moderate reduction in cell migration was observed ( Figure 5D ) . # ::alignments 0-1.1.2 1-1.1.2.1.r 2-1.1.2.1.1.1 4-1.1.2.2.1.1 6-1.2.1.3.2.1.2 8-1.1.2.2.2.1.4.1.1.1 9-1.1.2.2.2.1.3.1.1 10-1.1.2.2.2.1.1.1 12-1.1.2.2.2.1.2.1 15-1.2.1.3.2.1 17-1.1.2.2.3.1.2 18-1.1.2.2.3.1.1 19-1.1.2.2.3.1.1 20-1.1.2.2.3.1.1 22-1.1.2.2.3.1 24-1.1.1 24-1.1.1.r 25-1.1 29-1.1.3.1.1.1.1 29-1.2.1.3.2.1.1.1 30-1.2.1.1.1 32-1.2.1.1 34-1 36-1.2.1.3.1.1 38-1.2.1.3.2.1.1.1 39-1.1.2.2 39-1.2.1.3.2.1 40-1.1.2.2.2 40-1.2.1.3.2.1.2 41-1.2.1.3.2.1.2.1.r 42-1.2.1.3.2.1.2.1.2 43-1.1.2.2.2.1.3 43-1.2.1.3.2.1.2.1 44-1.2.1.3.2.1.2.1.1.1 47-1.2.1.2 48-1.2.1 50-1.1.3.1.1 50-1.2.1.3 51-1.1.3.1 53-1.2 55-1.3.1 57-1.3.1.1 (c / contrast-01~e.34 :ARG1 (r / reverse-01~e.25 :polarity~e.24 -~e.24 :ARG0 (d / deplete-01~e.0 :ARG1~e.1 (p / protein :name (n / name :op1 "RhoA"~e.2)) :location (c2 / cell~e.39 :name (n2 / name :op1 "HTshBR3"~e.4) :ARG1-of (t / transfect-01~e.40 :ARG2 (e / enzyme :name (n3 / name :op1 "BRAF"~e.10) :ARG2-of (m / mutate-01 :value "V600E"~e.12) :mod (v / vector~e.43 :name (n4 / name :op1 "pSUPER"~e.9)) :ARG1-of (e2 / encode-01 :ARG0 (n10 / nucleic-acid :name (n5 / name :op1 "shRNA"~e.8))))) :ARG0-of (h / have-03 :ARG1 (a / activity-06~e.22 :ARG0 e~e.18,19,20 :ARG1-of (s / suppress-01~e.17))))) :ARG1 (p2 / possible-01 :ARG1 (m2 / migrate-01~e.51 :ARG0 (c3 / cell-line~e.50 :name (n6 / name :op1 "HT29"~e.29))))) :ARG2 (o / observe-01~e.53 :ARG1 (r3 / reduce-01~e.48 :ARG1 (m3 / migrate-01~e.32 :ARG0 (c4 / cell~e.30)) :ARG1-of (m4 / moderate-03~e.47) :location (c5 / cell-line~e.50 :name (n7 / name :op1 "HTps"~e.36) :ARG1-of (d2 / describe-01 :ARG0 (c6 / cell-line~e.15,39 :name (n8 / name :op1 "HT29"~e.29,38) :ARG1-of (t2 / transfect-01~e.6,40 :ARG2~e.41 (v2 / vector~e.43 :name (n9 / name :op1 "pSUPER"~e.44) :ARG1-of (e3 / empty-02~e.42)))))))) :ARG1-of (d3 / describe-01 :ARG0 (f / figure~e.55 :mod "5D"~e.57))) # ::id a_pmid_2194_3101.191 ::amr-annotator SDL-AMR-09 ::preferred # ::tok Taken together , these results indicate that both BRAF @ and KRAS @ oncogenes utilize RhoA activation to promote cell migration . # ::alignments 0-1.3 1-1.3.1 3-1.1.2 4-1.1 4-1.1.1 4-1.1.1.r 5-1 9-1.2.1.1.1.1 11-1.2.1 13-1.2.1.2.1.1 16-1.2 17-1.2.2.1.1.1 18-1.2.2 19-1.2.1.3 20-1.2.3 21-1.2.3.2.1 22-1.2.3.2 (i / indicate-01~e.5 :ARG0 (t / thing~e.4 :ARG2-of~e.4 (r / result-01~e.4) :mod (t2 / this~e.3)) :ARG1 (u / utilize-01~e.16 :ARG0 (a / and~e.11 :op1 (g / gene :name (n / name :op1 "BRAF"~e.9)) :op2 (g2 / gene :name (n2 / name :op1 "KRAS"~e.13)) :ARG0-of (c2 / cause-01~e.19 :ARG1 (d / disease :wiki "Cancer" :name (n4 / name :op1 "cancer")))) :ARG1 (a2 / activate-01~e.18 :ARG1 (p / protein :name (n3 / name :op1 "RhoA"~e.17))) :purpose (p2 / promote-01~e.20 :ARG0 a :ARG1 (m / migrate-01~e.22 :ARG0 (c / cell~e.21)))) :ARG1-of (t3 / take-01~e.0 :mod (t4 / together~e.1))) # ::id a_pmid_2194_3101.192 ::amr-annotator SDL-AMR-09 ::preferred # ::tok In a different approach , inhibition of RhoA downstream signalling was achieved via treatment of cells with UO126 , a MEK ( mitogen @-@ activated protein kinase @/@ extracellular signal @-@ regulated kinase kinase ) inhibitor targeting the MAPK pathway , which is active in Caco @-@ BR cells [ @ 21 @ ] . # ::alignments 2-1.3.1 3-1.3 5-1.1 6-1.1.1.r 7-1.1.1.1.1.1 8-1.1.1.2 9-1.1.1 11-1 13-1.2 14-1.2.1.r 15-1.2.1 16-1.2.2.r 17-1.2.2.1.1 20-1.2.2.2.1.1.1 22-1.2.2.2.1.2.1.1.1.1 24-1.2.2.2.1.2.1.1.1.1 24-1.2.2.3.1.2 25-1.2.2.2.1.2.1.1.1.2 26-1.2.2.2.1.2.1.1.1.3 26-1.2.2.2.1.2.1.2.1.3 28-1.2.2.2.1.2.1.2.1.1 29-1.2.2.2.1.2.1.2.1.2 31-1.2.2.2.1.2.1.2.1.2 32-1.2.2.2.1.2.1.1.1.3 32-1.2.2.2.1.2.1.2.1.3 33-1.2.2.2.1.2.1.1.1.3 33-1.2.2.2.1.2.1.2.1.3 35-1.2.2 35-1.2.2.2 35-1.2.2.2.r 36-1.2.2.3 38-1.2.2.3.1.1.1 39-1.2.2.3.1 44-1.2.2.3.1.2.1.r 45-1.2.2.3.1.2.1.1.1 47-1.2.2.3.1.2.1.1.1 48-1.2.2.3.1.2.1 51-1.4.1.1.1 (a / achieve-01~e.11 :ARG1 (i / inhibit-01~e.5 :ARG1~e.6 (s / signal-07~e.9 :ARG0 (p / protein :name (n / name :op1 "RhoA"~e.7)) :location (d / downstream~e.8))) :manner (t / treat-04~e.13 :ARG1~e.14 (c / cell~e.15) :ARG2~e.16 (s2 / small-molecule~e.35 :name (n2 / name :op1 "UO126"~e.17) :ARG0-of~e.35 (i2 / inhibit-01~e.35 :ARG1 (p4 / protein-family :name (n3 / name :op1 "MEK"~e.20) :ARG1-of (m / mean-01 :ARG2 (o / or :op1 (p5 / protein-family :name (n4 / name :op1 "mitogen-activated"~e.22,24 :op2 "protein"~e.25 :op3 "kinase"~e.26,32,33)) :op2 (p6 / protein-family :name (n5 / name :op1 "extracellular"~e.28 :op2 "signal-regulated"~e.29,31 :op3 "kinase"~e.26,32,33)))))) :ARG0-of (t2 / target-01~e.36 :ARG1 (p2 / pathway~e.39 :name (n6 / name :op1 "MAPK"~e.38) :ARG1-of (a2 / activate-01~e.24 :location~e.44 (c2 / cell-line~e.48 :name (n7 / name :op1 "Caco-BR"~e.45,47))))))) :ARG1-of (a3 / approach-02~e.3 :ARG1-of (d2 / differ-02~e.2)) :ARG1-of (d3 / describe-01 :ARG0 (p3 / publication :ARG1-of (c3 / cite-01 :ARG2 21~e.51)))) # ::id a_pmid_2194_3101.193 ::amr-annotator SDL-AMR-09 ::preferred # ::tok Treatment with UO126 , at the most optimal treatment condition ( Additional Figure 3 ) , resulted in the decreased activation of RhoA illustrating that mutant BRAF @ V600E @ utilises the MAPK pathway to activate RhoA ( Figure 5E , upper panel ) . # ::alignments 0-1.1 2-1.1.1.1.1 6-1.1.2.1 7-1.1.2 8-1.1 9-1.1.2.r 12-1.1.3.1 14-1.1.3.1.1 18-1 19-1.2.r 21-1.2.3 22-1.2 24-1.2.2.1.1 25-1.3 27-1.3.1.1 27-1.3.1.1.2 27-1.3.1.1.2.r 28-1.3.1.1.1.1 30-1.3.1.1.2.1 34-1.3.1.2.1.1 35-1.3.1.2 37-1.2 37-1.3.1.3 38-1.2.2.1.1 40-1.4.1.1 42-1.4.1.1.1 45-1.4.1.1.2.1 46-1.4.1.1.2 (r / result-01~e.18 :ARG1 (t / treat-04~e.0,8 :ARG2 (s / small-molecule :name (n / name :op1 "UO126"~e.2)) :condition~e.9 (o / optimal~e.7 :degree (m / most~e.6)) :ARG1-of (d / describe-01 :ARG0 (f / figure~e.12 :mod 3~e.14 :ARG1-of (a / add-02)))) :ARG2~e.19 (a2 / activate-01~e.22,37 :ARG0 t :ARG1 (p / protein :name (n2 / name :op1 "RhoA"~e.24,38)) :ARG1-of (d2 / decrease-01~e.21)) :ARG0-of (i / illustrate-01~e.25 :ARG1 (u / utilize-01 :ARG0 (e / enzyme~e.27 :name (n3 / name :op1 "BRAF"~e.28) :ARG2-of~e.27 (m2 / mutate-01~e.27 :value "V600E"~e.30)) :ARG1 (p2 / pathway~e.35 :name (n4 / name :op1 "MAPK"~e.34)) :purpose (a3 / activate-01~e.37 :ARG0 e :ARG1 p))) :ARG1-of (d3 / describe-01 :ARG0 (a4 / and :op1 (f2 / figure~e.40 :mod "5E"~e.42 :part (p3 / panel~e.46 :mod (u2 / upper~e.45)))))) # ::id a_pmid_2194_3101.194 ::amr-annotator SDL-AMR-09 ::preferred # ::tok Alternative regulation of RhoA through the PI3K pathway was analysed in Caco @-@ BR cells , and a mild effect on RhoA downstream components like p @-@ Cofilin and p @-@ Myl was observed ( Figure 5E , lower panel ) . # ::alignments 1-1.1.1 2-1.1.1.2.r 3-1.1.1.2.1.1 6-1.1.1.1.1.1 7-1.1.1.1 9-1.1 10-1.1.2.r 11-1.1.2.1.1 13-1.1.2.1.1 14-1.1.2 16-1 18-1.2.1.2 19-1.2.1 20-1.2.1.1.r 21-1.2.1.1.2 22-1.2.1.1.1 23-1.2.1.1 24-1.2.1.1.3.r 25-1.2.1.1.3.1 25-1.2.1.1.3.1.2 25-1.2.1.1.3.1.2.r 27-1.2.1.1.3.1.1.1 29-1.2.1.1.3.1 29-1.2.1.1.3.1.2 29-1.2.1.1.3.1.2.r 31-1.2.1.1.3.2.1.1 33-1.2 35-1.3.1.1 37-1.3.1.1.1 40-1.3.1.1.2.1 40-1.3.1.1.2.1.1 40-1.3.1.1.2.1.1.r 41-1.3.1.1.2 (a / and~e.16 :op1 (a2 / analyze-01~e.9 :ARG1 (r / regulate-01~e.1 :ARG0 (p / pathway~e.7 :name (n / name :op1 "PI3K"~e.6)) :ARG1~e.2 (p2 / protein :name (n2 / name :op1 "RhoA"~e.3)) :ARG1-of (a3 / alternate-01)) :location~e.10 (c / cell-line~e.14 :name (n3 / name :op1 "Caco-BR"~e.11,13))) :op2 (o / observe-01~e.33 :ARG1 (a4 / affect-01~e.19 :ARG1~e.20 (c2 / component~e.23 :location (d / downstream~e.22) :part-of p2~e.21 :example~e.24 (a5 / and :op1 (p3 / protein~e.25,29 :name (n4 / name :op1 "Cofilin"~e.27) :ARG1-of~e.25,29 (p4 / phosphorylate-01~e.25,29)) :op2 (p5 / protein :name (n5 / name :op1 "Myl"~e.31) :ARG1-of p4))) :degree (m / mild~e.18))) :ARG1-of (d2 / describe-01 :ARG0 (a6 / and :op1 (f / figure~e.35 :mod "5E"~e.37 :part (p6 / panel~e.41 :ARG1-of (l / low-04~e.40 :degree~e.40 (m2 / more~e.40))))))) # ::id a_pmid_2194_3101.195 ::amr-annotator SDL-AMR-09 ::preferred # ::tok Analysis of RhoA @-@ ROCK axis # ::alignments 0-1 1-1.1.r 2-1.1.1.1 4-1.1.1.1 5-1.1 (a / analyze-01~e.0 :ARG1~e.1 (a2 / axis~e.5 :name (n / name :op1 "RhoA-ROCK"~e.2,4))) # ::id a_pmid_2194_3101.196 ::amr-annotator SDL-AMR-09 ::preferred # ::tok Since RhoA appears to be essential for the attained migration in Caco @-@ BR13 cells , RhoA @-@ Rho kinase signalling was inhibited using the selective ROCK ( Rho @-@ associated coiled coil forming protein serine @/@ threonine kinase ) inhibitor Y @-@ 27632 aiming to inhibit cell migration . # ::alignments 0-1 1-1.2.2.1.1.1 2-1.1 3-1.1.1.r 4-1.1.1.1.r 5-1.1.1 6-1.1.1.2.r 8-1.1.1.2.1 9-1.1.1.2 10-1.1.1.2.2.r 11-1.1.1.2.2.1.1 13-1.1.1.2.2.1.1 14-1.1.1.2.2 16-1.1.1.1.1.1 18-1.2.2.1.1.1 19-1.2.2.1 20-1.2.2 22-1.2 22-1.2.1.2 23-1.2.3.r 26-1.2.1.2.1.1.1 28-1.2.2.1.1.1 34-1.1.1.1 38-1.2.2.1 40-1.2 40-1.2.1.2 41-1.2.1.1.1 43-1.2.1.1.1 44-1.2.3 46-1.2.3.1 47-1.2.3.1.1.1 48-1.2.3.1.1 (c / cause-01~e.0 :ARG0 (a / appear-02~e.2 :ARG1~e.3 (e / essential~e.5 :domain~e.4 (p / protein~e.34 :name (n / name :op1 "RhoA"~e.16)) :purpose~e.6 (m / migrate-01~e.9 :ARG1-of (a2 / attain-01~e.8) :location~e.10 (c2 / cell-line~e.14 :name (n2 / name :op1 "Caco-BR13"~e.11,13))))) :ARG1 (i / inhibit-01~e.22,40 :ARG0 (s / small-molecule :name (n4 / name :op1 "Y-27632"~e.41,43) :ARG0-of (i2 / inhibit-01~e.22,40 :ARG1 (e2 / enzyme :name (n5 / name :op1 "ROCK"~e.26))) :ARG0-of (s2 / select-01)) :ARG1 (s3 / signal-07~e.20 :ARG0 (k / kinase~e.19,38 :name (n6 / name :op1 "RhoA-Rho"~e.1,18,28))) :purpose~e.23 (a3 / aim-01~e.44 :ARG1 (i3 / inhibit-01~e.46 :ARG1 (m2 / migrate-01~e.48 :ARG0 (c3 / cell~e.47)))))) # ::id a_pmid_2194_3101.197 ::amr-annotator SDL-AMR-09 ::preferred # ::tok Treatment of Caco @-@ 2 and Caco @-@ BR13 cells with the ROCK inhibitor had a moderate effect on downstream target p @-@ Cofilin , while cell motility was found significantly increased in both cell lines ( Figure 6A @-@ B ) . # ::alignments 0-1.1.1 1-1.1.1.1.r 2-1.1.1.1.1.2.1 2-1.1.1.1.2.2.1 4-1.1.1.1.1.2.1 5-1.1.1.1 6-1.1.1.1.1.2.1 6-1.1.1.1.2.2.1 8-1.1.1.1.2.2.1 9-1.1.1.1.1 9-1.1.1.1.2 10-1.1.1.2.r 12-1.1.1.2.1.1.2.1 13-1.1.1.2 13-1.1.1.2.1 13-1.1.1.2.1.r 16-1.1.3 17-1.1 18-1.1.2.r 19-1.1.2.4.1 20-1.1.2.4 21-1.1.2 21-1.1.2.3 21-1.1.2.3.r 23-1.1.2.2.1 25-1 26-1.2.1.1.1 27-1.2.1.1 29-1.2 30-1.2.1.2 31-1.2.1 34-1.2.1.1.1 35-1.2.2 37-1.3.1.1 37-1.3.1.2 39-1.3.1.1.1 (c / contrast-01~e.25 :ARG1 (a / affect-01~e.17 :ARG0 (t / treat-04~e.0 :ARG1~e.1 (a2 / and~e.5 :op1 (c2 / cell-line~e.9 :wiki "Caco-2" :name (n / name :op1 "Caco-2"~e.2,4,6)) :op2 (c3 / cell-line~e.9 :wiki - :name (n2 / name :op1 "Caco-BR13"~e.2,6,8))) :ARG2~e.10 (m / molecular-physical-entity~e.13 :ARG0-of~e.13 (i / inhibit-01~e.13 :ARG1 (e / enzyme :wiki "ROCK1" :name (n3 / name :op1 "ROCK"~e.12))))) :ARG1~e.18 (p / protein~e.21 :wiki "Cofilin" :name (n4 / name :op1 "Cofilin"~e.23) :ARG1-of~e.21 (p2 / phosphorylate-01~e.21) :ARG1-of (t3 / target-01~e.20 :location (d / downstream~e.19))) :ARG1-of (m2 / moderate-03~e.16)) :ARG2 (f / find-01~e.29 :ARG1 (i2 / increase-01~e.31 :ARG1 (m3 / motility~e.27 :mod (c4 / cell~e.26,34)) :ARG2 (s / significant-02~e.30)) :location a2~e.35) :ARG1-of (d2 / describe-01 :ARG0 (a3 / and :op1 (f2 / figure~e.37 :mod "6A"~e.39) :op2 (f3 / figure~e.37 :mod "6B")))) # ::id a_pmid_2194_3101.198 ::amr-annotator SDL-AMR-09 ::preferred # ::tok To exclude the possibility of this observation being the non @-@ specific effect of the inhibitor targeting several other kinases , siRNA against both ROCK isoforms ( ROCK1 and ROCK2 ) was applied to both Caco @-@ BR clones and parental Caco @-@ 2 cells ( Figure 6C ) . # ::alignments 1-1.3 3-1.3.1 4-1.3.1.1.r 5-1.3.1.1.2 6-1.3.1.1 9-1.3.1.1.1.2.1 9-1.3.1.1.1.2.1.r 11-1.3.1.1.1.2 12-1.3.1.1.1 13-1.3.1.1.1.1.r 15-1.3.1.1.1.1 15-1.3.1.1.1.1.1 15-1.3.1.1.1.1.1.r 16-1.3.1.1.1.1.2 17-1.3.1.1.1.1.2.1.2 18-1.3.1.1.1.1.2.1.1 19-1.3.1.1.1.1.2.1 21-1.1.1.1 22-1.1 22-1.1.2 22-1.1.2.r 25-1.1.2.1.1 25-1.1.2.1.2 27-1.1.2.1.1.1.1 28-1.1.2.1 29-1.1.2.1.2.1.1 32-1 35-1.2.1.1.1.1 37-1.2.1.1.1.1 38-1.2.1 39-1.2 40-1.2.2.2 41-1.2.2.1.1 43-1.2.2.1.1 44-1.2.1.1 44-1.2.2 46-1.4.1 48-1.4.1.1 (a / apply-02~e.32 :ARG1 (n4 / nucleic-acid~e.22 :name (n / name :op1 "siRNA"~e.21) :ARG0-of~e.22 (o / oppose-01~e.22 :ARG1 (a2 / and~e.28 :op1 (i / isoform~e.25 :name (n2 / name :op1 "ROCK1"~e.27)) :op2 (i2 / isoform~e.25 :name (n3 / name :op1 "ROCK2"~e.29))))) :ARG2 (a3 / and~e.39 :op1 (c / clone-01~e.38 :ARG1 (c2 / cell-line~e.44 :name (n5 / name :op1 "Caco-BR"~e.35,37))) :op2 (c3 / cell-line~e.44 :name (n6 / name :op1 "Caco-2"~e.41,43) :mod (p / parent~e.40))) :purpose (e2 / exclude-01~e.1 :ARG1 (p2 / possible-01~e.3 :ARG1~e.4 (o2 / observe-01~e.6 :ARG1 (a4 / affect-01~e.12 :ARG0~e.13 (m2 / molecular-physical-entity~e.15 :ARG0-of~e.15 (i3 / inhibit-01~e.15) :ARG0-of (t2 / target-01~e.16 :ARG1 (k / kinase~e.19 :mod (o3 / other~e.18) :quant (s / several~e.17)))) :ARG1-of (s2 / specific-02~e.11 :polarity~e.9 -~e.9)) :mod (t / this~e.5)))) :ARG1-of (d / describe-01 :ARG0 (f / figure~e.46 :mod "6C"~e.48))) # ::id a_pmid_2194_3101.199 ::amr-annotator SDL-AMR-09 ::preferred # ::tok Besides , the use of siRNA to deplete a protein and especially a small GTPase can prove more promising since the specific protein sequence is targeted . # ::alignments 3-1.1.1.1 4-1.1.1.1.1.r 5-1.1.1.1.1.1.1 7-1.1.1.1.2 9-1.1.1.1.2.2.1 10-1.1.1.1.2.2 11-1.1.1.1.2.2.2.3 13-1.1.1.1.2.2.2.2 14-1.1.1.1.2.2.2.1.1 15-1.1 16-1.1.1 17-1.1.1.2.1 18-1.1.1.2 19-1.1.2 21-1.1.2.1.1.1 22-1.1.2.1.1.2 23-1.1.2.1.1 25-1.1.2.1 (a / and :op2 (p / possible-01~e.15 :ARG1 (p2 / prove-01~e.16 :ARG0 (u / use-01~e.3 :ARG1~e.4 (n3 / nucleic-acid :name (n / name :op1 "siRNA"~e.5)) :ARG2 (d / deplete-01~e.7 :ARG0 n3 :ARG1 (a2 / and~e.10 :op1 (p3 / protein~e.9) :op2 (e / enzyme :name (n2 / name :op1 "GTPase"~e.14) :mod (s / small~e.13) :mod (e2 / especially~e.11))))) :ARG1 (p4 / promise-01~e.18 :degree (m / more~e.17))) :ARG1-of (c / cause-01~e.19 :ARG0 (t / target-01~e.25 :ARG1 (s3 / sequence~e.23 :ARG1-of (s2 / specific-02~e.21) :part-of (p5 / protein~e.22)))))) # ::id a_pmid_2194_3101.200 ::amr-annotator SDL-AMR-09 ::preferred # ::tok In several reported studies , treatment with a selective inhibitor may produce more adverse effect through interaction with other ( associated ) components . # ::alignments 1-1.2.2 2-1.2.1 3-1.2 5-1.1.1 9-1.1.1.1 9-1.1.1.1.1 9-1.1.1.1.1.r 10-1 11-1.1 12-1.1.2.1.1 13-1.1.2.1 14-1.1.2 16-1.1.3 17-1.1.3.2.r 18-1.1.3.2.1 20-1.1.3.2.2 22-1.1.3.2 (p / possible-01~e.10 :ARG1 (p2 / produce-01~e.11 :ARG0 (t / treat-04~e.5 :ARG2 (m / molecular-physical-entity~e.9 :ARG0-of~e.9 (i / inhibit-01~e.9) :ARG0-of (s / select-01))) :ARG1 (a / affect-01~e.14 :mod (a2 / adverse~e.13 :degree (m2 / more~e.12))) :manner (i2 / interact-01~e.16 :ARG0 m :ARG1~e.17 (c / component~e.22 :mod (o / other~e.18) :ARG1-of (a3 / associate-01~e.20)))) :medium (s2 / study-01~e.3 :ARG1-of (r / report-01~e.2) :quant (s3 / several~e.1))) # ::id a_pmid_2194_3101.201 ::amr-annotator SDL-AMR-09 ::preferred # ::tok Regardless efficient ROCK depletion , no inhibition in cell migration or invasion was observed in BRAF @ V600E @ transformed cells ( Figure 6D ) . # ::alignments 0-1.3.r 1-1.3.2 2-1.3.1.1.1 3-1.3 5-1.1.1 5-1.1.1.r 6-1.1 7-1.1.2.r 8-1.1.2.1.1 9-1.1.2.1 10-1.1.2 11-1.1.2.2 13-1 14-1.2.r 15-1.2.1.1.1.1 17-1.2.1.1.2.1 19-1.2.1 20-1.2 22-1.4.1 24-1.4.1.1 (o / observe-01~e.13 :ARG1 (i / inhibit-01~e.6 :polarity~e.5 -~e.5 :ARG1~e.7 (o2 / or~e.10 :op1 (m / migrate-01~e.9 :ARG0 (c / cell~e.8)) :op2 (i2 / invade-01~e.11 :ARG0 c))) :location~e.14 (c2 / cell~e.20 :ARG1-of (t / transform-01~e.19 :ARG0 (e / enzyme :name (n / name :op1 "BRAF"~e.15) :ARG2-of (m2 / mutate-01 :value "V600E"~e.17)))) :concession~e.0 (d / deplete-01~e.3 :ARG1 (e2 / enzyme :name (n2 / name :op1 "ROCK"~e.2)) :ARG1-of (e3 / efficient-01~e.1)) :ARG1-of (d2 / describe-01 :ARG0 (f / figure~e.22 :mod "6D"~e.24))) # ::id a_pmid_2194_3101.202 ::amr-annotator SDL-AMR-09 ::preferred # ::tok Nevertheless increase motility was recorded in Caco @-@ 2 cells suggesting that Rac1 activation may be taking a lead role in the absence of the RhoA @-@ Rho kinase signalling . # ::alignments 0-1 1-1.1.1 2-1.1.1.1 4-1.1 5-1.1.2.r 6-1.1.2.1.1 8-1.1.2.1.1 9-1.1.2 10-1.1.3 11-1.1.3.1.r 12-1.1.3.1.1.1.1.1.1 13-1.1.3.1.1.1 14-1.1.3.1 16-1.1.3.1.1 18-1.1.3.1.1.2.1 19-1.1.3.1.1.2 20-1.1.3.1.1.3.r 22-1.1.3.1.1.3 23-1.1.3.1.1.3.1.r 25-1.1.3.1.1.3.1.1.1.1 27-1.1.3.1.1.3.1.1.1.1 28-1.1.3.1.1.3.1.1 29-1.1.3.1.1.3.1 (h / have-concession-91~e.0 :ARG1 (r / record-01~e.4 :ARG1 (i / increase-01~e.1 :ARG1 (m / motility~e.2)) :location~e.5 (c / cell-line~e.9 :name (n / name :op1 "Caco-2"~e.6,8)) :ARG0-of (s / suggest-01~e.10 :ARG1~e.11 (p / possible-01~e.14 :ARG1 (t / take-01~e.16 :ARG0 (a / activate-01~e.13 :ARG1 (p2 / protein :name (n2 / name :op1 "Rac1"~e.12))) :ARG1 (r2 / role~e.19 :purpose (l / lead-02~e.18)) :condition~e.20 (a2 / absent-01~e.22 :ARG1~e.23 (s2 / signal-07~e.29 :ARG0 (k / kinase~e.28 :name (n4 / name :op1 "RhoA-Rho"~e.25,27))))))))) # ::id a_pmid_2194_3101.203 ::amr-annotator SDL-AMR-09 ::preferred # ::tok KRAS @ G12V @ induces Cdc42 @-@ dependent migration ability and filopodia formation in Caco @-@ 2 cells , partially dependent on PI3K pathway # ::alignments 0-1.1.2.1.1 2-1.1.1 4-1 5-1.2.1.1.2.1.1.1 7-1.2.1.1.2 8-1.2.1.1 10-1.2 12-1.2.2 13-1.2.2.2.r 14-1.2.2.2.1.1 16-1.2.2.2.1.1 17-1.2.2.2 19-1.1.3.2 19-1.1.3.2.r 20-1.1.3 21-1.1.3.1.r 22-1.1.3.1.1.1 23-1.1.3.1 (i / induce-01~e.4 :ARG0 (m2 / mutate-01 :value "G12V"~e.2 :ARG1 (g / gene :name (n4 / name :op1 "KRAS"~e.0)) :ARG0-of (d3 / depend-01~e.20 :ARG1~e.21 (p4 / pathway~e.23 :name (n7 / name :op1 "PI3K"~e.22)) :degree~e.19 (p2 / part~e.19))) :ARG2 (a / and~e.10 :op1 (p3 / possible-01 :ARG1 (m / migrate-01~e.8 :ARG0 n2 :condition (d / depend-01~e.7 :ARG1 (p / protein :name (n / name :op1 "Cdc42"~e.5))))) :op2 (f / form-01~e.12 :ARG1 (f2 / filopodium) :location~e.13 (c / cell-line~e.17 :name (n2 / name :op1 "Caco-2"~e.14,16))))) # ::id a_pmid_2194_3101.204 ::amr-annotator SDL-AMR-09 ::preferred # ::tok Previous studies have indicated that RhoA , Rac1 and Cdc42 signalling is essential for oncogenic Ras transforming capacity [ @ 29 , 30 @ ] . # ::alignments 0-1.1.1 1-1.1 3-1 4-1.2.r 5-1.2.1.1.1.1.1 7-1.2.1.1.2.1.1 8-1.2.1.1 9-1.2.1.1.3.1.1 10-1.2.1 11-1.2.1.r 12-1.2 13-1.2.2.r 14-1.2.2.1 14-1.2.2.1.2 14-1.2.2.1.2.1.2.1 14-1.2.2.1.2.r 15-1.2.2.1.1.1 16-1.2.2.2 17-1.2.2 20-1.3.1.1.1 24-1.3.1.1.2 (i / indicate-01~e.3 :ARG0 (s / study-01~e.1 :time (p / previous~e.0)) :ARG1~e.4 (e / essential~e.12 :domain~e.11 (s2 / signal-07~e.10 :ARG0 (a / and~e.8 :op1 (p2 / protein :name (n / name :op1 "RhoA"~e.5)) :op2 (p3 / protein :name (n2 / name :op1 "Rac1"~e.7)) :op3 (p4 / protein :name (n3 / name :op1 "Cdc42"~e.9)))) :purpose~e.13 (c3 / capable-01~e.17 :ARG1 (e2 / enzyme~e.14 :name (n4 / name :op1 "Ras"~e.15) :ARG0-of~e.14 (c / cause-01~e.14 :ARG1 (d2 / disease :wiki "Disease" :name (n5 / name :op1 "Cancer"~e.14)))) :ARG2 (t / transform-01~e.16 :ARG1 e2))) :ARG1-of (d / describe-01 :ARG0 (c2 / cite-01 :ARG1 (a2 / and :op1 29~e.20 :op2 30~e.24)))) # ::id a_pmid_2194_3101.205 ::amr-annotator SDL-AMR-09 ::preferred # ::tok In the present study , Caco @-@ 2 cells overexpressing mutant KRAS @ G12V @ ( Caco @-@ K ) , selective activation for Cdc42 was detected ( Figure 7A ) . # ::alignments 2-1.2.2 3-1.2 5-1.2.1.2.1.1 5-1.2.1.3.1.1.1 7-1.2.1.2.1.1 8-1.2.1.2 8-1.2.1.3.1 9-1.2.1 10-1.2.1.1 11-1.2.1.1.2.1.1 13-1.2.1.1.1 16-1.2.1.2.1.1 16-1.2.1.3.1.1.1 18-1.2.1.3.1.1.1 21-1.1.2 22-1.1 23-1.1.1.r 24-1.1.1.1.1 26-1 28-1.3.1 30-1.3.1.1 (d / detect-01~e.26 :ARG1 (a / activate-01~e.22 :ARG1~e.23 (p / protein :name (n / name :op1 "Cdc42"~e.24)) :manner (s / selective~e.21)) :location (s2 / study-01~e.3 :ARG1 (o2 / overexpress-01~e.9 :ARG1 (m / mutate-01~e.10 :value "G12V"~e.13 :ARG1 (g / gene :name (n3 / name :op1 "KRAS"~e.11))) :location (c / cell-line~e.8 :name (n2 / name :op1 "Caco-2"~e.5,7,16)) :ARG1-of (m2 / mean-01 :ARG2 (c2 / cell-line~e.8 :name (n4 / name :op1 "Caco-K"~e.5,16,18)))) :time (p2 / present~e.2)) :ARG1-of (d2 / describe-01 :ARG0 (f / figure~e.28 :mod "7A"~e.30))) # ::id a_pmid_2194_3101.206 ::amr-annotator SDL-AMR-09 ::preferred # ::tok The formation of filopodia in these cells , earlier described , was in agreement with the high Cdc42 activity and is illustrated here by staining with antibody against Fascin , a filopodia marker ( Figure 7B , upper panel ) . # ::alignments 1-1.1.1 4-1.1.1.2.r 5-1.1.1.2.1 6-1.1.1.2 8-1.3.2 8-1.3.2.1 8-1.3.2.1.r 9-1.3 13-1.1 14-1.1.2.r 16-1.1.2.2 17-1.1.2.1.1.1 18-1.1.2 19-1 21-1.2 22-1.2.3 23-1.2.2.r 24-1.2.2 25-1.2.2.1.r 26-1.2.2.1 27-1.2.2.1.1 28-1.2.2.1.1.1.1.1 32-1.2.2.1.1.1 34-1.3.1.2 36-1.3.1.2.1 39-1.3.1.1 40-1.3.1 (a / and~e.19 :op1 (a2 / agree-01~e.13 :ARG1 (f / form-01~e.1 :ARG1 (f2 / filopodium) :location~e.4 (c / cell~e.6 :mod (t / this~e.5))) :ARG2~e.14 (a3 / activity-06~e.18 :ARG0 (p / protein :name (n / name :op1 "Cdc42"~e.17)) :ARG1-of (h / high-02~e.16))) :op2 (i / illustrate-01~e.21 :ARG1 f :ARG0-of~e.23 (s / stain-01~e.24 :ARG2~e.25 (a4 / antibody~e.26 :ARG0-of (c2 / counter-01~e.27 :ARG1 (m / marker~e.32 :name (n2 / name :op1 "Fascin"~e.28) :mod (f3 / filopodium))))) :medium (h2 / here~e.22)) :ARG1-of (d / describe-01~e.9 :ARG0 (p2 / panel~e.40 :location (u / upper~e.39) :part-of (f4 / figure~e.34 :mod "7B"~e.36)) :mod (e / early~e.8 :degree~e.8 (m3 / more~e.8)))) # ::id a_pmid_2194_3101.207 ::amr-annotator SDL-AMR-09 ::preferred # ::tok A large number of relatively short filopodia distributed almost exclusively at the cell periphery was evident in Caco @-@ K cells , while Caco @-@ BR and Caco @-@ H cells formed less but longer structures with a rather polarized shape potentially pointing towards the direction of cell migration ( Figure 7B , upper panel ) . # ::alignments 1-1.1.1.2.1 2-1.1.1.2 4-1.1.1.1.1 5-1.1.1.1 7-1.1.1.3 8-1.1.1.3.2.1 9-1.1.1.3.2 10-1.1.1.3.1.r 12-1.1.1.3.1.1 13-1.1.1.3.1 14-1.1.1.3.1.1.r 14-1.1.1.r 15-1.1 16-1.1.2.r 17-1.1.2.1.1 19-1.1.2.1.1 20-1.1.2 22-1 23-1.2.1.1.1.1 25-1.2.1.1.1.1 26-1.2.1 27-1.2.1.1.1.1 27-1.2.1.2.1.1 29-1.2.1.2.1.1 30-1.2.1.1 30-1.2.1.2 31-1.2 32-1.2.2.2 33-1.2.2.2.1 34-1.2.2.2.1.1 34-1.2.2.2.1.1.2 34-1.2.2.2.1.1.2.r 35-1.2.2 38-1.2.2.1.1.1.1 39-1.2.2.1.1.1 40-1.2.2.1.1 41-1.2.2.1.1.2.2 42-1.2.2.1.1.2 43-1.2.2.1.1.2.1.1.r 45-1.2.2.1.1.2.1 45-1.2.2.1.1.2.1.1.r 47-1.2.2.1.1.2.1.1.1 48-1.2.2.1.1.2.1.1 50-1.3.1.2 52-1.3.1.2.1 55-1.3.1.1 56-1.3.1 (c / contrast-01~e.22 :ARG1 (e3 / evident~e.15 :domain~e.14 (f / filopodium :ARG1-of (s / short-07~e.5 :ARG2-of (r / relative-05~e.4)) :quant (n2 / number~e.2 :mod (l2 / large~e.1)) :ARG1-of (d / distribute-01~e.7 :location~e.10 (p6 / periphery~e.13 :domain~e.14 (c7 / cell~e.12)) :ARG0-of (e / exclusive-02~e.9 :degree (a3 / almost~e.8)))) :location~e.16 (c2 / cell~e.20 :name (n / name :op1 "Caco-K"~e.17,19))) :ARG2 (f2 / form-01~e.31 :ARG0 (a2 / and~e.26 :op1 (c4 / cell-line~e.30 :name (n3 / name :op1 "Caco-BR"~e.23,25,27)) :op2 (c5 / cell-line~e.30 :name (n4 / name :op1 "Caco-H"~e.27,29))) :ARG1 (s2 / structure~e.35 :ARG0-of (h / have-03 :ARG1 (s4 / shape~e.40 :ARG1-of (p / polarize-01~e.39 :degree (r3 / rather~e.38)) :ARG0-of (p7 / point-01~e.42 :ARG2 (d4 / direction~e.45 :direction-of~e.43,45 (m2 / migrate-01~e.48 :ARG0 (c3 / cell~e.47))) :mod (p8 / potential~e.41)))) :quant (l4 / less~e.32 :ARG1-of (c6 / contrast-01~e.33 :ARG2 (l / long-03~e.34 :ARG1 s :degree~e.34 (m / more~e.34)) :compared-to f)))) :ARG1-of (d3 / describe-01 :ARG0 (p5 / panel~e.56 :location (u / upper~e.55) :part-of (f3 / figure~e.50 :mod "7B"~e.52)))) # ::id a_pmid_2194_3101.208 ::amr-annotator SDL-AMR-09 ::preferred # ::tok Nevertheless , no changes in Fascin protein expression were recorded in the different cell lines , ( Figure 7B , lower panel ) . # ::alignments 0-1 2-1.1.1.1 2-1.1.1.1.r 3-1.1.1 4-1.1.1.2.r 5-1.1.1.2.1.1.1 6-1.1.1.2.1 7-1.1.1.2 9-1.1 10-1.1.2.r 12-1.1.2.1 13-1.1.2 14-1.1.2 17-1.2.1.2 19-1.2.1.2.1 22-1.2.1.1 22-1.2.1.1.1 22-1.2.1.1.1.r 23-1.2.1 (h / have-concession-91~e.0 :ARG2 (r / record-01~e.9 :ARG1 (c / change-01~e.3 :polarity~e.2 -~e.2 :ARG1~e.4 (e / express-03~e.7 :ARG2 (p / protein~e.6 :name (n / name :op1 "Fascin"~e.5)))) :location~e.10 (c2 / cell-line~e.13,14 :ARG1-of (d / differ-02~e.12))) :ARG1-of (d2 / describe-01 :ARG0 (p2 / panel~e.23 :ARG1-of (l / low-04~e.22 :degree~e.22 (m2 / more~e.22)) :part-of (f / figure~e.17 :mod "7B"~e.19)))) # ::id a_pmid_2194_3101.209 ::amr-annotator SDL-AMR-09 ::preferred # ::tok Increased migration ability in Caco @-@ BR and Caco @-@ H cells may be indicative for the length and the location of filopodia . # ::alignments 0-1.1.1 1-1.1.1.1.1 3-1.1.1.1.1.1.r 4-1.1.1.1.1.1.1.1.1 4-1.1.1.1.1.1.2.1.1 6-1.1.1.1.1.1.1.1.1 7-1.1.1.1.1.1 8-1.1.1.1.1.1.1.1.1 8-1.1.1.1.1.1.2.1.1 10-1.1.1.1.1.1.2.1.1 11-1.1.1.1.1.1.1 11-1.1.1.1.1.1.2 12-1 12-1.1.1.1 17-1.1.2.1 18-1.1.2 20-1.1.2.2 20-1.1.2.2.1.r (p / possible-01~e.12 :ARG1 (i / indicate-01 :ARG0 (i2 / increase-01~e.0 :ARG1 (p2 / possible-01~e.12 :ARG1 (m / migrate-01~e.1 :location~e.3 (a2 / and~e.7 :op1 (c / cell-line~e.11 :name (n / name :op1 "Caco-BR"~e.4,6,8)) :op2 (c2 / cell-line~e.11 :name (n2 / name :op1 "Caco-H"~e.4,8,10)))))) :ARG1 (a3 / and~e.18 :op1 (l / length~e.17 :mod (f / filopodium)) :op2 (l2 / location~e.20 :location-of~e.20 f)))) # ::id a_pmid_2194_3101.210 ::amr-annotator SDL-AMR-09 ::preferred # ::tok It has been previously shown that in CHO @-@ K1 cells ( Chinese hamster ovary fibroblast @- like cells ) RhoA expression down @-@ regulates Cdc42 and Rac1 activity in order to regulate membrane protrusions and cell polarity . # ::alignments 3-1.2 4-1 7-1.1.1.2.1.1 9-1.1.1.2.1.1 10-1.1.1.2.2.1.2.1 12-1.1.1.2.2.1.1.1.1.1.2.1 13-1.1.1.2.2.1.1.1.1 14-1.1.1.2.2.1.1 15-1.1.1.2.2.1 17-1.1.1.2.2.1.2 18-1.1.1.2 20-1.1.1.1.1.1 21-1.1.1 24-1.1.3 25-1.1.2.1.1.1.1 26-1.1.2.1 27-1.1.2.1.2.1.1 28-1.1.2 29-1.1.3.r 30-1.1.3.r 31-1.1.3.r 32-1.1.3 33-1.1.3.2.1.1 34-1.1.3.2.1 35-1.1.3.2 36-1.1.3.2.2.1 37-1.1.3.2.2 (s / show-01~e.4 :ARG1 (d / downregulate-01 :ARG0 (e / express-03~e.21 :ARG2 (p2 / protein :name (n3 / name :op1 "RhoA"~e.20)) :location (c2 / cell-line~e.18 :name (n6 / name :op1 "CHO-K1"~e.7,9) :ARG1-of (m2 / mean-01 :ARG2 (f2 / fibroblast~e.15 :location (o2 / ovary~e.14 :part-of (a5 / animal :mod (h2 / hamster~e.13 :mod (c5 / country :wiki "China" :name (n / name :op1 "China"~e.12))))) :ARG1-of (r3 / resemble-01~e.17 :ARG2 (c6 / cell~e.10)))))) :ARG1 (a / activity-06~e.28 :ARG0 (a2 / and~e.26 :op1 (p3 / protein :name (n4 / name :op1 "Cdc42"~e.25)) :op2 (p4 / protein :name (n5 / name :op1 "Rac1"~e.27)))) :purpose~e.29,30,31 (r2 / regulate-01~e.24,32 :ARG0 e :ARG1 (a4 / and~e.35 :op1 (p5 / protrude-01~e.34 :ARG1 (m / membrane~e.33)) :op2 (p6 / polarity~e.37 :mod (c4 / cell~e.36))))) :time (p / previous~e.3)) # ::id a_pmid_2194_3101.211 ::amr-annotator SDL-AMR-09 ::preferred # ::tok In addition , Rac1 activity may down @-@ regulate Cdc42 activity and promote the formation of stabilized rather than transient protrusions [ @ 31 @ ] . # ::alignments 0-1.1.1 1-1 1-1.1.1 3-1.1.1.1.1.1.1.1 4-1.1.1.1.1 4-1.1.1.1.2 5-1.1 9-1.1.1.1.2.1.1.1 10-1.1.1.1.1 10-1.1.1.1.2 11-1.1.1 12-1.1.1.2 14-1.1.1.2.1 15-1.1.1.2.1.2 16-1.1.1.2.1.1.1 17-1.1.1.2.1.2 19-1.1.1.2.1.2.1.1 20-1.1.1.2.1.1 20-1.1.1.2.1.2.1 23-1.2.1 (a / and~e.1 :op2 (p / possible-01~e.5 :ARG1 (a2 / and~e.0,1,11 :op1 (d2 / downregulate-01 :ARG0 (a3 / activity-06~e.4,10 :ARG0 (p2 / protein :name (n / name :op1 "Rac1"~e.3))) :ARG1 (a4 / activity-06~e.4,10 :ARG0 (p3 / protein :name (n2 / name :op1 "Cdc42"~e.9)))) :op2 (p4 / promote-02~e.12 :ARG1 (f / form-01~e.14 :ARG1 (p5 / protrude-01~e.20 :ARG1-of (s / stabilize-01~e.16)) :ARG1-of (i / instead-of-91~e.15,17 :ARG2 (p6 / protrude-01~e.20 :ARG1-of (t / transient-02~e.19))))))) :ARG1-of (c / cite-01 :ARG2 31~e.23)) # ::id a_pmid_2194_3101.212 ::amr-annotator SDL-AMR-09 ::preferred # ::tok Indeed , low Cdc42 activity was recorded in Caco @-@ BR and Caco @-@ H cells where RhoA signaling is activated . # ::alignments 0-1.3 2-1.1.2 3-1.1.1.1.1 4-1.1 6-1 7-1.2.r 8-1.2.1.1.1 8-1.2.2.1.1 10-1.2.1.1.1 11-1.2 12-1.2.1.1.1 12-1.2.2.1.1 14-1.2.2.1.1 15-1.2.1 15-1.2.2 16-1.2.3.r 17-1.2.3.1.1.1.1 18-1.2.3.1 20-1.2.3 (r / record-01~e.6 :ARG1 (a / activity-06~e.4 :ARG0 (p / protein :name (n / name :op1 "Cdc42"~e.3)) :ARG1-of (l / low-04~e.2)) :location~e.7 (a2 / and~e.11 :op1 (c / cell-line~e.15 :name (n2 / name :op1 "Caco-BR"~e.8,10,12)) :op2 (c2 / cell-line~e.15 :name (n3 / name :op1 "Caco-H"~e.8,12,14)) :location-of~e.16 (a4 / activate-01~e.20 :ARG1 (s2 / signal-07~e.18 :ARG0 (p3 / protein :name (n5 / name :op1 "RhoA"~e.17))))) :mod (i / indeed~e.0)) # ::id a_pmid_2194_3101.213 ::amr-annotator SDL-AMR-09 ::preferred # ::tok To explore the role of Cdc42 in mutant KRAS @ G12V @ induced cell transformation , Caco @-@ 2 and Caco @-@ K15 cells were treated with siRNA against this small GTPase . # ::alignments 1-1.2 3-1.2.1.1.2 5-1.2.1.1.1.1 7-1.2.1.3.1 8-1.2.1.3.1.2.1.1 10-1.2.1.3.1.1 12-1.2.1.3 13-1.2.1.2 14-1.2.1 16-1.1.1.1.1.1 16-1.1.1.2.1.1 18-1.1.1.1.1.1 19-1.1.1 20-1.1.1.1.1.1 20-1.1.1.2.1.1 22-1.1.1.2.1.1 23-1.1.1.1 23-1.1.1.2 25-1.1 30-1.1.2.1.1 30-1.1.3.2.2 31-1.1.3.2.1.1 (h / have-purpose-91 :ARG1 (t / treat-04~e.25 :ARG1 (a / and~e.19 :op1 (c / cell-line~e.23 :name (n2 / name :op1 "Caco-2"~e.16,18,20)) :op2 (c2 / cell-line~e.23 :name (n3 / name :op1 "Caco-K15"~e.16,20,22))) :ARG2 (n7 / nucleic-acid :name (n4 / name :op1 "small"~e.30 :op2 "interfering" :op3 "RNA")) :purpose (d / defend-01 :ARG2 n7 :ARG3 (e2 / enzyme :name (n5 / name :op1 "GTPase"~e.31) :mod (s / small~e.30)))) :ARG2 (e / explore-01~e.1 :ARG1 (t2 / transform-01~e.14 :ARG0 (p / protein :name (n / name :op1 "Cdc42"~e.5) :domain (r / role~e.3)) :ARG1 (c3 / cell~e.13) :ARG2-of (i / induce-01~e.12 :ARG0 (m / mutate-01~e.7 :value "G12V"~e.10 :ARG1 (g / gene :name (n6 / name :op1 "KRAS"~e.8))))))) # ::id a_pmid_2194_3101.214 ::amr-annotator SDL-AMR-09 ::preferred # ::tok Significant downregulation of Cdc42 at the protein level was observed in both cell lines ( Figure 7C @ -@ upper panel ) , that caused a significant decrease of cell migration and invasion ability of Caco @-@ K15 and of Caco @-@ 2 cells but to a lesser extent ( Figure 7C @ -@ lower panel ) . # ::alignments 0-1.1.2 1-1.1 2-1.1.1.r 3-1.1.1.1.1 4-1.1.3.r 6-1.1.3.1 7-1.1.3 9-1.1.4 10-1.1.4.1.r 11-1.1.4.1.1 12-1.1.4.1 13-1.1.4.1 15-1.1.5.1 17-1.1.5.1.1 20-1.1.5.1.2.1 21-1.1.5.1.2 25-1 27-1.2.3 28-1.2 29-1.2.2.r 30-1.2.2.1.1.1 30-1.2.2.2.1.1 31-1.2.2.1.1 31-1.2.2.2.1 32-1.2.2 32-1.2.2.1 32-1.2.2.1.r 32-1.2.2.2 32-1.2.2.2.r 33-1.2.2.1.2.1 33-1.2.2.2.2 36-1.2.2.1.1.1.1.1 36-1.2.2.2.1.1.1.1 38-1.2.2.1.1.1.1.1 39-1.2.2.1 41-1.2.2.1.1.1.1.1 41-1.2.2.2.1.1.1.1 43-1.2.2.2.1.1.1.1 44-1.2.2.1.1.1 45-1.2.2.1.3 46-1.2.2.1.3.2.r 48-1.2.2.1.3.2.1 49-1.2.2.1.3.2 51-1.2.4.1 53-1.2.4.1.1 56-1.2.4.1.2.1 56-1.2.4.1.2.1.1 56-1.2.4.1.2.1.1.r 57-1.2.4.1.2 (c / cause-01~e.25 :ARG0 (d / downregulate-01~e.1 :ARG1~e.2 (p / protein :name (n / name :op1 "Cdc42"~e.3)) :ARG1-of (s / significant-02~e.0) :prep-at~e.4 (l2 / level~e.7 :mod (p2 / protein~e.6)) :ARG1-of (o / observe-01~e.9 :location~e.10 (c2 / cell-line~e.12,13 :mod (b / both~e.11))) :ARG1-of (d2 / describe-01 :ARG0 (f / figure~e.15 :mod "7C"~e.17 :part-of (p3 / panel~e.21 :mod (u / upper~e.20))))) :ARG1 (d3 / decrease-01~e.28 :ARG0 d :ARG1~e.29 (a / and~e.32 :op1~e.32 (a2 / and~e.32,39 :op1 (m3 / migrate-01~e.31 :ARG0 (c3 / cell-line~e.30,44 :name (n2 / name :op1 "Caco-K15"~e.36,38,41))) :op2 (p5 / possible-01 :ARG1 (i3 / invade-01~e.33 :ARG0 c3)) :ARG1-of (c6 / contrast-01~e.45 :ARG2 a4 :mod~e.46 (e / extent~e.49 :degree (l / less~e.48)))) :op2~e.32 (a4 / and~e.32 :op1 (m4 / migrate-01~e.31 :ARG0 (c4 / cell-line~e.30 :name (n3 / name :op1 "Caco-2"~e.36,41,43))) :op2 (i2 / invade-01~e.33 :ARG0 c4))) :ARG2 (s2 / significant-02~e.27) :ARG1-of (d4 / describe-01 :ARG0 (f2 / figure~e.51 :mod "7C"~e.53 :part-of (p4 / panel~e.57 :ARG1-of (l3 / low-04~e.56 :degree~e.56 (m5 / more~e.56))))))) # ::id a_pmid_2194_3101.215 ::amr-annotator SDL-AMR-09 ::preferred # ::tok Depletion of Cdc42 also affected the filopodia formation , when Caco @-@ K cells were treated with siRNA against Cdc42 acquired rounded cell membrane lacking filapodia protrusion suggesting that filopodia formation in Caco @-@ K cells is Cdc42 @-@ dependent ( Figure 7D ) . # ::alignments 0-1.1.1 1-1.1.1.1.r 2-1.1.1.1.1.1 3-1.1.3 4-1.1 7-1.1.2 10-1.1.4.2.1.1 11-1.1.4.2.1.1 12-1.1.4.2.1.1 13-1.1.4.2.1.1 15-1.1.4.2.1 18-1.1.4.2.1.3 19-1.1.4.2.1.3.1 20-1.1.4 20-1.1.4.2.2.1 21-1.1.4.2.2.1.2.1 22-1.1.4.2.2.1.2.2 23-1.1.4.2.2.1.2 24-1.1.4.2.2.1.2.3 26-1.1.4.2.2.1.2.3.1 27-1 30-1.2.1 31-1.2.1.2.r 32-1.2.1.2 33-1.2.1.2 34-1.2.1.2 35-1.2.1.2 37-1.2.2 39-1.2 41-1.3.1 43-1.3.1.1 (s / suggest-01~e.27 :ARG0 (a / affect-01~e.4 :ARG0 (d / deplete-01~e.0 :ARG1~e.1 (p / protein :name (n / name :op1 "Cdc42"~e.2))) :ARG1 (f / form-01~e.7 :ARG1 (f2 / filopodium)) :mod (a2 / also~e.3) :condition (a3 / acquire-01~e.20 :ARG0 (c2 / cell :name (n4 / name :op1 "Caco-K")) :ARG1-of (c3 / condition-01 :ARG0 (t / treat-04~e.15 :ARG1 c2~e.10,11,12,13 :ARG2 (n2 / nucleic-acid :name (n5 / name :op1 "small" :op2 "interfere" :op3 "RNA")) :ARG0-of (c4 / counter-01~e.18 :ARG1 p~e.19)) :ARG1-of (r2 / result-01 :ARG2 (a4 / acquire-01~e.20 :ARG0 c2 :ARG1 (m / membrane~e.23 :ARG1-of (r / round-04~e.21) :mod (c5 / cell~e.22) :ARG0-of (l / lack-01~e.24 :ARG1 (p2 / protrude-01~e.26 :ARG1 (f5 / filopodium))))))))) :ARG2 (d2 / depend-01~e.39 :ARG0 (f3 / form-01~e.30 :ARG1 (f4 / filopodium) :location~e.31 c2~e.32,33,34,35) :ARG1 p~e.37) :ARG1-of (d3 / describe-01 :ARG0 (f6 / figure~e.41 :mod "7D"~e.43))) # ::id a_pmid_2194_3101.216 ::amr-annotator SDL-AMR-09 ::preferred # ::tok These findings suggest that KRAS @ G12V @ regulates motility and invasiveness of colon cancer cells through the Cdc42 GTPase . # ::alignments 0-1.1.1 1-1.1 1-1.1.2 1-1.1.2.r 2-1 3-1.2.r 4-1.2.1.2.1.1 6-1.2.1.1 8-1.2 9-1.2.2.1 12-1.2.2.1.1.r 13-1.2.2.1.1.1.2 14-1.2.2.1.1.1.1.1 15-1.2.2.1.1 18-1.2.3.1.1 19-1.2.3.1.2 (s / suggest-01~e.2 :ARG0 (t2 / thing~e.1 :mod (t / this~e.0) :ARG1-of~e.1 (f / find-01~e.1)) :ARG1~e.3 (r / regulate-01~e.8 :ARG0 (m / mutate-01 :value "G12V"~e.6 :ARG1 (g / gene :name (n3 / name :op1 "KRAS"~e.4))) :ARG1 (a3 / and :op1 (m2 / motility~e.9 :mod~e.12 (c2 / cell~e.15 :mod (d / disease :name (n / name :op1 "Cancer"~e.14) :location (c / colon~e.13)))) :op2 (i / invade-01 :ARG0 c2)) :manner (p / protein :name (n4 / name :op1 "Cdc42"~e.18 :op2 "GTPase"~e.19)))) # ::id a_pmid_2194_3101.217 ::amr-annotator SDL-AMR-09 ::preferred # ::tok Considering that the PI3K pathway is also a KRAS effector pathway , the possibility of a cross @-@ talk between the PI3K signalling pathway and Cdc42 was explored [ @ 16 , 21 @ ] . # ::alignments 0-1.2 1-1.2.1.r 3-1.2.1.2 4-1.2.1 5-1.2.1.2.r 6-1.2.2 8-1.2.1.1.1.1.1 9-1.2.1.1 10-1.2.1 13-1.1 21-1.1.1.1.1.1.1.1 22-1.1.1.1.1 23-1.1.1.1 25-1.1.1.2.1.1 26-1.2.1.2.r 27-1 30-1.3.1.1 34-1.3.1.2 (e / explore-01~e.27 :ARG1 (p / possible-01~e.13 :ARG1 (i / interfere-01 :ARG0 (p2 / pathway~e.23 :ARG1-of (s / signal-07~e.22 :ARG0 (p3 / pathway :name (n / name :op1 "PI3K"~e.21)))) :ARG1 (p4 / protein :name (n2 / name :op1 "Cdc42"~e.25)))) :condition (c / consider-01~e.0 :ARG1~e.1 (p5 / pathway~e.4,10 :mod (e2 / effector~e.9 :mod (g / gene :name (n3 / name :op1 "KRAS"~e.8))) :domain~e.5,26 p3~e.3) :mod (a / also~e.6)) :ARG1-of (c2 / cite-01 :ARG2 (a2 / and :op1 16~e.30 :op2 21~e.34))) # ::id a_pmid_2194_3101.218 ::amr-annotator SDL-AMR-09 ::preferred # ::tok Following treatment with wortmanin at the most optimal treatment condition , as retrieved from inhibition of the active PI3K pathway in Caco @-@ H2 cells that show high p @-@ AKT levels ( Additional Figure 4 ) , resulted in reduced Cdc42 activity . # ::alignments 0-1.2.1 1-1.2.1 2-1.2.1 3-1.2.1 4-1.2.1 5-1.2.1 6-1.2.1 7-1.2.1 8-1.2.1 9-1.2.1 10-1.2.1 11-1.2.1 12-1.2.1 13-1.2.1 14-1.2.1 17-1.2.2 18-1.1.2.1.1.1 19-1.1.2.1 21-1.1.2.3.1.1 23-1.1.2.3.1.1 24-1.1.2.3 26-1.1.2.3.2 27-1.1.2.3.2.1.1 28-1.1.2.3.2.1.2.2 30-1.1.2.3.2.1.2.1.1 31-1.1.2.3.2.1 33-1.1.3.1.2 34-1.1.3.1 36-1.1.3.1.1 40-1 42-1.2 43-1.2.2.1.1.1 44-1.2.2 (r / result-01~e.40 :ARG1 (f3 / follow-01 :ARG2 (t / treat-04 :ARG2 (w2 / wortmanin)) :example (i / inhibit-01 :ARG1 (p2 / pathway~e.19 :name (n2 / name :op1 "PI3K"~e.18) :ARG1-of (a2 / activate-01)) :ARG1-of (r3 / retrieve-01) :location (c / cell-line~e.24 :name (n3 / name :op1 "Caco-H2"~e.21,23) :ARG0-of (s / show-01~e.26 :ARG1 (l / level~e.31 :ARG1-of (h / high-02~e.27) :quant-of (e / enzyme :name (n4 / name :op1 "AKT"~e.30) :ARG3-of (p3 / phosphorylate-01~e.28)))))) :ARG1-of (d / describe-01 :ARG0 (f2 / figure~e.34 :mod 4~e.36 :mod (a4 / additional~e.33))) :condition (t3 / treat-04 :mod (o2 / optimal :degree (m3 / most)))) :ARG2 (r2 / reduce-01~e.42 :ARG0 f3~e.0,1,2,3,4,5,6,7,8,9,10,11,12,13,14 :ARG1 (a / activity-06~e.17,44 :ARG0 (p / protein :name (n / name :op1 "Cdc42"~e.43))))) # ::id a_pmid_2194_3101.219 ::amr-annotator SDL-AMR-09 ::preferred # ::tok This illustrates how Cdc42 activation in response to the KRAS @ G12V @ -@ PI3K signalling pathway can be potentially essential for Cdc42 @-@ dependent cell migration and invasion properties ( Figure 7E ) . # ::alignments 0-1.1 1-1 2-1.2.1.r 3-1.2.1.1.1.1.1.1 4-1.2.1.1.1 5-1.2.1.1.1.2.r 6-1.2.1.1.1.2 14-1.2.1.1.1.2.1.1.1 15-1.2.1.1.1.2.1.2 16-1.2.1.1.1.2.1 17-1.2.1 18-1.2.1.1.1.r 20-1.2.1.1 21-1.2.1.1.2.r 22-1.2.1.1.2.1.1.1.1 24-1.2.1.1.2.1.1.1 25-1.2.1.1.2.1.1 26-1.2.1.1.2.1 27-1.2.1.1.2 28-1.2.1.1.2.2.1 29-1.2.1.1.2.2 31-1.3.1 33-1.3.1.1 (i / illustrate-01~e.1 :ARG0 (t / this~e.0) :ARG1 (t2 / thing :manner-of~e.2 (p4 / possible-01~e.17 :ARG1 (e2 / essential~e.20 :domain~e.18 (a2 / activate-01~e.4 :ARG1 (p6 / protein :name (n3 / name :op1 "Cdc42"~e.3)) :ARG2-of~e.5 (r2 / respond-01~e.6 :ARG1 (p7 / pathway~e.16 :name (n4 / name :op1 "KRASG12V-PI3K"~e.14) :ARG1-of (s2 / signal-07~e.15)))) :purpose~e.21 (a3 / and~e.27 :op1 (m / migrate-01~e.26 :ARG0 (c / cell~e.25 :ARG0-of (d3 / depend-01~e.24 :ARG1 p6~e.22))) :op2 (p8 / property~e.29 :mod (i2 / invade-01~e.28 :ARG0 c)))))) :ARG1-of (d2 / describe-01 :ARG0 (f / figure~e.31 :mod "7E"~e.33))) # ::id a_pmid_2194_3101.220 ::amr-annotator SDL-AMR-09 ::preferred # ::tok HRAS @ G12V @ induces high cell migration and invasion properties mediated by Rac1 associated with acquired EMT # ::alignments 0-1.1.2.1.1 2-1.1.1 4-1 5-1.2.1.2 6-1.2.1.1 7-1.2.1 8-1.2 9-1.2.2.1 10-1.2.2 11-1.2.2.1.1 12-1.2.2.1.1.1.r 13-1.2.2.1.1.1.1.1.1 14-1.2.2.1.1.1 15-1.2.2.1.1.1.2.r 16-1.2.2.1.1.1.2.3 17-1.2.2.1.1.1.2.2.2 (i / induce-01~e.4 :ARG0 (m / mutate-01 :value "G12V"~e.2 :ARG1 (g / gene :name (n / name :op1 "HRAS"~e.0))) :ARG2 (a3 / and~e.8 :op1 (m2 / migrate-01~e.7 :ARG0 (c2 / cell~e.6) :ARG1-of (h / high-02~e.5)) :op2 (p3 / property~e.10 :mod (i2 / invade-01~e.9 :ARG1-of (m3 / mediate-01~e.11 :ARG0~e.12 (a4 / associate-01~e.14 :ARG1 (p2 / protein :name (n4 / name :op1 "Rac1"~e.13)) :ARG2~e.15 (e / event :wiki "Epithelial–mesenchymal_transition" :name (n3 / name :op1 "epithelial−mesenchymal" :op2 "transition"~e.17) :ARG1-of (a6 / acquire-01~e.16)))))))) # ::id a_pmid_2194_3101.221 ::amr-annotator SDL-AMR-09 ::preferred # ::tok Activation of Rac1 , another RAS effector protein , was found slightly increased in Caco @-@ H2 cells with EMT characteristics [ @ 15 , 25 @ ] ( Figure 8A ) . # ::alignments 0-1.1.1 1-1.1.1.1.r 2-1.1.1.1.1.1 4-1.1.1.1.2 5-1.1.1.1.3.1.1.1 6-1.1.1.1.3 7-1.1.1.1 7-1.1.1.1.3.1 9-1.1.1.1.3.r 10-1 11-1.1.2 12-1.1 13-1.1.3.r 14-1.1.3.1.1 16-1.1.3.1.1 17-1.1.3 18-1.1.3.2.r 19-1.1.3.2.1.1.2 20-1.1.3.2 23-1.2.1.1.1.1.1 27-1.2.1.1.1.1.2 31-1.2.1.2 33-1.2.1.2.1 (f / find-01~e.10 :ARG1 (i3 / increase-01~e.12 :ARG1 (a / activate-01~e.0 :ARG1~e.1 (p / protein~e.7 :name (n2 / name :op1 "Rac1"~e.2) :mod (a6 / another~e.4) :domain~e.9 (e2 / effector~e.6 :mod (p2 / protein-family~e.7 :name (n3 / name :op1 "Ras"~e.5))))) :ARG2 (s2 / slight~e.11) :location~e.13 (c / cell-line~e.17 :name (n5 / name :op1 "Caco-H2"~e.14,16) :ARG1-of~e.18 (c6 / characteristic-02~e.20 :ARG2 (e / event :name (n / name :op1 "epithelial−mesenchymal" :op2 "transition"~e.19))))) :ARG1-of (d / describe-01 :ARG0 (a4 / and :op1 (p3 / publication :ARG1-of (c4 / cite-01 :ARG2 (a5 / and :op1 15~e.23 :op2 25~e.27))) :op2 (f2 / figure~e.31 :mod "8A"~e.33)))) # ::id a_pmid_2194_3101.222 ::amr-annotator SDL-AMR-09 ::preferred # ::tok Activation of Rac1 in Caco @-@ H2 cells is in agreement with previous studies that correlate Rac1 with EMT and the inhibition of E @-@ cadherin in mammary epithelial and pancreatic carcinoma cells respectively [ @ 32 @ ] . # ::alignments 0-1.1 1-1.1.1.r 2-1.1.1.1.1 4-1.1.1.2.1.1 6-1.1.1.2.1.1 7-1.1.1.2 10-1 11-1.2.r 12-1.2.2 13-1.2 15-1.2.1 16-1.2.1.1 17-1.2.1.2.r 18-1.2.1.2.1.1.2 19-1.2.1.2 21-1.2.1.2.2 22-1.2.1.2.2.1.r 23-1.2.1.2.2.1.1.1 25-1.2.1.2.2.1.1.1 27-1.2.1.2.2.1.2.1.2 28-1.2.1.2.2.1.2.1.1 29-1.2.1.2.2.1.2 30-1.2.1.2.2.1.2.2.1 31-1.2.1.2.2.1.2.2.2 32-1.2.1.2.2.1.2.1 32-1.2.1.2.2.1.2.2 33-1.2.1.2.2.1.2.3 36-1.3.1.1.1 (a / agree-01~e.10 :ARG0 (a2 / activate-01~e.0 :ARG1~e.1 (p / protein :name (n / name :op1 "Rac1"~e.2) :location (c / cell-line~e.7 :name (n2 / name :op1 "Caco-H2"~e.4,6)))) :ARG2~e.11 (s / study-01~e.13 :ARG0-of (c2 / correlate-01~e.15 :ARG1 p~e.16 :ARG2~e.17 (a3 / and~e.19 :op1 (e3 / event :name (n3 / name :op1 "epithelial−mesenchymal" :op2 "transition"~e.18)) :op2 (i / inhibit-01~e.21 :ARG1~e.22 (p2 / protein :name (n4 / name :op1 "E-cadherin"~e.23,25) :location (a4 / and~e.29 :op1 (c3 / cell~e.32 :mod (e / epithelium~e.28) :mod (m / mammary~e.27)) :op2 (c4 / cell~e.32 :mod (p3 / pancreas~e.30) :mod (c5 / carcinoma~e.31)) :mod (r / respective~e.33)))))) :time (p5 / previous~e.12)) :ARG1-of (d / describe-01 :ARG0 (p4 / publication :ARG1-of (c6 / cite-01 :ARG2 32~e.36)))) # ::id a_pmid_2194_3101.223 ::amr-annotator SDL-AMR-09 ::preferred # ::tok In contrast , a weak effect on Rac1 GTPase was recorded in Caco @-@ BR cells ( Figure 8A ) and could be explained by the known antagonistic effect that exists between RhoA and Rac1 [ @ 33 @ ] . # ::alignments 1-1 4-1.1.1.1.2 5-1.1.1.1 6-1.1.1.1.1.r 7-1.1.1.1.1.1.1 8-1.1.1.1.1.1.2 10-1.1.1 11-1.1.1.2.r 12-1.1.1.2.1.1 14-1.1.1.2.1.1 15-1.1.1.2 17-1.1.1.3.1 19-1.1.1.3.1.1 22-1.1 23-1.1.2 25-1.1.2.1 26-1.1.2.1.1.r 28-1.1.2.1.1.2 30-1.1.2.1.1 34-1.1.2.1.1.1.1.1.1 35-1.1 36-1.1.2.1.1.1.2.1.1 39-1.2.1.1.1 (c / contrast-01~e.1 :ARG2 (a / and~e.22,35 :op1 (r / record-01~e.10 :ARG1 (a2 / affect-01~e.5 :ARG1~e.6 (p2 / protein :name (n / name :op1 "Rac1"~e.7 :op2 "GTPase"~e.8)) :ARG1-of (w / weak-02~e.4)) :location~e.11 (c2 / cell-line~e.15 :name (n2 / name :op1 "Caco-BR"~e.12,14)) :ARG1-of (d / describe-01 :ARG0 (f / figure~e.17 :mod "8A"~e.19))) :op2 (p / possible-01~e.23 :ARG1 (e / explain-01~e.25 :ARG0~e.26 (a3 / affect-01~e.30 :ARG2 (a4 / antagonize-02 :ARG1 (p3 / protein :name (n3 / name :op1 "RhoA"~e.34)) :ARG2 (p4 / protein :name (n4 / name :op1 "Rac1"~e.36))) :ARG1-of (k / know-01~e.28)) :ARG1 a2))) :ARG1-of (d2 / describe-01 :ARG0 (p5 / publication :ARG1-of (c3 / cite-01 :ARG2 33~e.39)))) # ::id a_pmid_2194_3101.224 ::amr-annotator SDL-AMR-09 ::preferred # ::tok As described earlier , HRAS @ G12V @ -@ transfected Caco @-@ 2 cells ( Caco @-@ H2 ) have undergone EMT , followed by the dramatic reduction of E @-@ cadherin expression [ @ 16 @ ] . # ::alignments 0-1.4.1.r 1-1.4 1-1.5 2-1.4.1 2-1.4.1.1 2-1.4.1.1.r 4-1.1.2.1.1.1 6-1.1.2.1.2.1 9-1.1.2 10-1.1.1.1 12-1.1.1.1 13-1.1 15-1.1.1.1 20-1 21-1.2.1.2 23-1.3 24-1.3.1.r 26-1.3.1.2 27-1.3.1 28-1.3.1.1.r 29-1.3.1.1.1.1.1 31-1.3.1.1.1.1.1 32-1.3.1.1 35-1.5.1.1.1 (u / undergo-28~e.20 :ARG1 (c / cell-line~e.13 :name (n / name :op1 "Caco-2"~e.10,12,15) :ARG1-of (t / transfect-01~e.9 :ARG2 (e3 / enzyme :name (n2 / name :op1 "HRAS"~e.4) :ARG2-of (m / mutate-01 :value "G12V"~e.6)))) :ARG2 (e4 / event :name (n3 / name :op1 "epithelial−mesenchymal" :op2 "transition"~e.21)) :ARG2-of (f / follow-01~e.23 :ARG1~e.24 (r / reduce-01~e.27 :ARG1~e.28 (e / express-03~e.32 :ARG2 (p2 / protein :name (n4 / name :op1 "E-cadherin"~e.29,31))) :manner (d / dramatic~e.26))) :ARG1-of (d2 / describe-01~e.1 :time~e.0 (e2 / early~e.2 :degree~e.2 (m2 / more~e.2))) :ARG1-of (d3 / describe-01~e.1 :ARG0 (p3 / publication :ARG1-of (c2 / cite-01 :ARG2 16~e.35)))) # ::id a_pmid_2194_3101.225 ::amr-annotator SDL-AMR-09 ::preferred # ::tok Following PI3K pathway depletion using the specific inhibitor wortmanin at the most optimal treatment condition ( Additional Figure 4 ) , Rac1 activity was successfully inhibited only in Caco @-@ 2 cells , leaving Caco @-@ H2 cells unaffected ( Figure 8B , upper panel ) . # ::alignments 0-1.4 1-1.4.1.1.1.1 2-1.4.1.1 3-1.4.1 4-1.4.1.2 6-1.4.1.2.1.3 7-1.4.1.2.1 7-1.4.1.2.1.2 7-1.4.1.2.1.2.r 8-1.4.1.2.1.1.1 11-1.4.1.2.2.1.1 13-1.4.1.2.2 14-1.4.1.2.2.r 17-1.4.1.3.1 19-1.4.1.3.1.1 23-1.1.1.1.1 24-1.1 26-1.2 27-1 28-1.3.2 30-1.3.1.1 32-1.3.1.1 33-1.3 35-1.1.1.2 36-1.1.1.2.1.2.1.1 38-1.1.1.2.1.2.1.1 39-1.1.1.2.1.2 40-1.1.1.2.1 40-1.1.1.2.1.1 40-1.1.1.2.1.1.r 42-1.5.1 44-1.5.1.1 47-1.5.1.2.1 48-1.5.1.2 (i / inhibit-01~e.27 :ARG1 (a / activity-06~e.24 :ARG0 (p / protein :name (n / name :op1 "Rac1"~e.23) :ARG0-of (l / leave-13~e.35 :ARG1 (a2 / affect-01~e.40 :polarity~e.40 -~e.40 :ARG1 (c2 / cell-line~e.39 :name (n3 / name :op1 "Caco-H2"~e.36,38)))))) :ARG1-of (s / succeed-01~e.26) :location (c / cell-line~e.33 :name (n2 / name :op1 "Caco-2"~e.30,32) :mod (o / only~e.28)) :ARG2-of (f / follow-01~e.0 :ARG1 (d / deplete-01~e.3 :ARG1 (p2 / pathway~e.2 :name (n4 / name :op1 "PI3K"~e.1)) :ARG0-of (u / use-01~e.4 :ARG1 (s2 / small-molecule~e.7 :name (n5 / name :op1 "wortmanin"~e.8) :ARG0-of~e.7 (i2 / inhibit-01~e.7) :ARG1-of (s3 / specific-02~e.6)) :condition~e.14 (t2 / treat-04~e.13 :mod (o2 / optimum :degree (m2 / most~e.11)))) :ARG1-of (d2 / describe-01 :ARG0 (f2 / figure~e.17 :mod 4~e.19 :ARG1-of (a3 / add-02))))) :ARG1-of (d3 / describe-01 :ARG0 (f3 / figure~e.42 :mod "8B"~e.44 :location (p3 / panel~e.48 :mod (u2 / upper~e.47))))) # ::id a_pmid_2194_3101.226 ::amr-annotator SDL-AMR-09 ::preferred # ::tok Notably , under the same treatment conditions RhoA activity was found to be slightly increased , suggesting an involvement of the PI3K pathway in RhoA regulation ( Figure 8B , lower panel ) . # ::alignments 0-1.4 4-1.2.1 5-1.2 6-1.2.r 7-1.1.1.1.2.1 8-1.1.1 10-1 13-1.1.2 14-1.1 16-1.1.3 18-1.1.3.1 19-1.1.3.1.1.r 21-1.1.3.1.1.2.1 22-1.1.3.1.1 23-1.1.3.1.2.r 24-1.1.3.1.2.1 25-1.1.3.1.2 27-1.3.1 29-1.3.1.1 32-1.3.1.2.1 32-1.3.1.2.1.1 32-1.3.1.2.1.1.r 33-1.3.1.2 (f / find-02~e.10 :ARG1 (i / increase-01~e.14 :ARG1 (a / activity-06~e.8 :ARG0 (p / protein :wiki "RHOA" :name (n / name :op1 "RhoA"~e.7))) :ARG2 (s / slight~e.13) :ARG0-of (s3 / suggest-01~e.16 :ARG1 (i2 / involve-01~e.18 :ARG1~e.19 (p2 / pathway~e.22 :wiki - :name (n2 / name :op1 "PI3K"~e.21)) :ARG2~e.23 (r / regulate-01~e.25 :ARG1 p~e.24)))) :condition~e.6 (t / treat-04~e.5 :ARG1-of (s2 / same-01~e.4)) :ARG1-of (d / describe-01 :ARG0 (f2 / figure~e.27 :mod "8B"~e.29 :location (p3 / panel~e.33 :ARG1-of (l / low-04~e.32 :degree~e.32 (m / more~e.32))))) :ARG1-of (n3 / notable-04~e.0)) # ::id a_pmid_2194_3101.227 ::amr-annotator SDL-AMR-09 ::preferred # ::tok It is therefore concluded that in Caco @-@ H2 cells , HRAS @ G12V @ deregulates PI3K @-@ dependent activation of Rac1 as well as mediates RhoA inhibition . # ::alignments 2-1 3-1.1 5-1.1.1.r 6-1.1.1.1.3.1.1 8-1.1.1.1.3.1.1 9-1.1.1.1.3 11-1.1.1.1.1.1.1 13-1.1.1.1.1.2.1 15-1.1.1.1 16-1.1.1.1.2.1.2.1.1.1 18-1.1.1.1.2.1 18-1.1.1.1.2.1.2 18-1.1.1.1.2.1.2.r 19-1.1.1.1.2 21-1.1.1.1.2.1.1.1 22-1.1.1 23-1.1.1 24-1.1.1 25-1.1.1.2 26-1.1.1.2.2.1.1.1 27-1.1.1.2.2 (c2 / cause-01~e.2 :ARG1 (c / conclude-01~e.3 :ARG1~e.5 (a2 / and~e.22,23,24 :op1 (d / deregulate-01~e.15 :ARG0 (e2 / enzyme :name (n / name :op1 "HRAS"~e.11) :ARG2-of (m / mutate-01 :value "G12V"~e.13)) :ARG1 (a / activate-01~e.19 :ARG1 (p2 / protein~e.18 :name (n2 / name :op1 "Rac1"~e.21) :ARG0-of~e.18 (d2 / depend-01~e.18 :ARG1 (e / enzyme :name (n3 / name :op1 "PI3K"~e.16))))) :location (c3 / cell-line~e.9 :name (n5 / name :op1 "Caco-H2"~e.6,8))) :op2 (m2 / mediate-01~e.25 :ARG0 e2 :ARG1 (i / inhibit-01~e.27 :ARG1 (p3 / protein :name (n4 / name :op1 "RhoA"~e.26))))))) # ::id a_pmid_2194_3101.228 ::amr-annotator SDL-AMR-09 ::preferred # ::tok To further explore the involvement of Rac1 activation in the transforming ability of HRAS @ G12V @ in Caco @-@ 2 cells , pharmacological inhibition of Rac1 was established using the selective inhibitor NSC23766 ( Figure 8C ) [ @ 34 @ ] . # ::alignments 1-1.3.2 2-1.3 4-1.3.1 5-1.3.1.1.r 6-1.3.1.1.1 7-1.3.1.1 8-1.3.1.2.r 10-1.3.1.2.2 11-1.3.1.2 12-1.3.1.2.1.r 13-1.3.1.2.1.1.1 15-1.3.1.2.1.2.1 17-1.3.1.2.2.2.r 18-1.3.1.2.2.2.1.1 20-1.3.1.2.2.2.1.1 21-1.3.1.2.2.2 24-1.1 24-1.2.1 24-1.2.1.2 24-1.2.1.2.r 25-1.1.1.r 26-1.1.1.1.1 28-1 29-1.2 29-1.3.r 31-1.2.1.3 32-1.2.1 32-1.2.1.2 32-1.2.1.2.r 33-1.2.1.1.1 35-1.4.1.1 37-1.4.1.1.1 42-1.4.1.2.1.1 (e / establish-01~e.28 :ARG1 (i / inhibit-01~e.24 :ARG1~e.25 (p / protein :name (n / name :op1 "Rac1"~e.26)) :mod (p2 / pharmacologic)) :ARG2-of (u / use-01~e.29 :ARG1 (s2 / small-molecule~e.24,32 :name (n2 / name :op1 "NSC23766"~e.33) :ARG0-of~e.24,32 (i2 / inhibit-01~e.24,32) :mod (s / selective~e.31))) :purpose~e.29 (e2 / explore-01~e.2 :ARG1 (i3 / involve-01~e.4 :ARG1~e.5 (a / activate-01~e.7 :ARG1 p~e.6) :ARG2~e.8 (c / capable-01~e.11 :ARG1~e.12 (e3 / enzyme :name (n3 / name :op1 "HRAS"~e.13) :ARG2-of (m2 / mutate-01 :value "G12V"~e.15)) :ARG2 (t / transform-01~e.10 :ARG0 e3 :location~e.17 (c2 / cell-line~e.21 :name (n4 / name :op1 "Caco-2"~e.18,20))))) :degree (f / further~e.1)) :ARG1-of (d / describe-01 :ARG0 (a2 / and :op1 (f2 / figure~e.35 :mod "8C"~e.37) :op2 (p3 / publication :ARG1-of (c3 / cite-01 :ARG2 34~e.42))))) # ::id a_pmid_2194_3101.229 ::amr-annotator SDL-AMR-09 ::preferred # ::tok Inhibition of Rac1 not only managed to suppress Rac1 activation but also to abolish cell migration and invasion properties in a dose dependent manner ( Figure 8D ) , indicating the critical role of Rac1 in EMT cell properties of Caco @-@ H cells . # ::alignments 0-1.1 1-1.1.1.r 2-1.1.1.1.1 5-1 7-1.2.1 8-1.2.1.2.1 9-1.2.1.2 11-1.2.2.3 13-1.2.2 14-1.2.2.2.1.1 15-1.2.2.2.1.1.1 16-1.2.2.2 17-1.2.2.2.2.1.1 18-1.2.2.2.1 21-1.2.2.4.1 22-1.2.2.4 23-1.2.2.4.r 25-1.3.1 27-1.3.1.1 31-1.4 33-1.4.1.1 34-1.4.1 36-1.1.1.1.1 37-1.4.1.1.1.r 38-1.4.1.1.1.2.1.2 39-1.4.1.1.1.1 40-1.2.2.2.2 40-1.4.1.1.1 42-1.4.1.1.1.1.1.1 44-1.4.1.1.1.1.1.1 45-1.2.2.2.2.1 (m / manage-01~e.5 :ARG0 (i / inhibit-01~e.0 :ARG1~e.1 (p / protein :name (n / name :op1 "Rac1"~e.2,36))) :ARG1 (a / and :op1 (s / suppress-01~e.7 :ARG0 i :ARG1 (a2 / activate-01~e.9 :ARG1 p~e.8)) :op2 (a3 / abolish-01~e.13 :ARG0 i :ARG1 (a4 / and~e.16 :op1 (p2 / property~e.18 :mod (c / cell~e.14 :ARG0-of (m2 / migrate-01~e.15))) :op2 (p3 / property~e.40 :mod (c5 / cell~e.45 :ARG0-of (i2 / invade-01~e.17)))) :mod (a5 / also~e.11) :manner~e.23 (d / depend-01~e.22 :ARG1 (d2 / dose~e.21)))) :ARG1-of (d3 / describe-01 :ARG0 (f / figure~e.25 :mod "8D"~e.27)) :ARG0-of (i3 / indicate-01~e.31 :ARG1 (r / role~e.34 :ARG1-of (c2 / critical-02~e.33 :ARG2~e.37 (p4 / property~e.40 :poss (c3 / cell-line~e.39 :name (n3 / name :op1 "Caco-H"~e.42,44)) :mod (e / event :name (n2 / name :op1 "epithelial−mesenchymal" :op2 "transition"~e.38)))) :poss p))) # ::id a_pmid_2194_3101.230 ::amr-annotator SDL-AMR-09 ::preferred # ::tok TGFβ-1 co @-@ operates with BRAF @ V600E @ @ and KRAS @ G12V @ @ oncogenes to provide Caco @-@ 2 cells with enhanced transformation properties # ::alignments 0-1.1.1.1 6-1.2.1.1.1 8-1.2.1.2.1 11-1.2 13-1.2.2.1.1 15-1.2.2.2.1 19-1.2.3 20-1.3 21-1.3.3.1.1 23-1.3.3.1.1 24-1.3.3 25-1.3.2.r 26-1.3.2.2 27-1.3.2.1 28-1.3.2 (c / cooperate-01 :ARG0 (p / protein :name (n / name :op1 "TGFβ-1"~e.0)) :ARG1 (a / and~e.11 :op1 (g / gene :name (n2 / name :op1 "BRAF"~e.6) :ARG2-of (m / mutate-01 :value "V600E"~e.8)) :op2 (g2 / gene :name (n3 / name :op1 "KRAS"~e.13) :ARG2-of (m2 / mutate-01 :value "G12V"~e.15)) :ARG0-of (c2 / cause-01~e.19 :ARG1 (d / disease :wiki "Cancer" :name (n5 / name :op1 "cancer")))) :ARG2 (p2 / provide-01~e.20 :ARG0 p :ARG1~e.25 (p3 / property~e.28 :ARG0-of (t / transform-01~e.27) :ARG1-of (e / enhance-01~e.26)) :ARG2 (c3 / cell-line~e.24 :name (n4 / name :op1 "Caco-2"~e.21,23)))) # ::id a_pmid_2194_3101.231 ::amr-annotator SDL-AMR-09 ::preferred # ::tok Since BRAF @ V600E @ @ and KRAS @ G12V @ @ oncogenes did not manage to fully transform Caco @-@ 2 cells nor induced an EMT phenotype , as HRAS @ G12V @ @ did , it was further investigated whether co @-@ operation of oncogene @-@ growth factor can produce synergistic effect . # ::alignments 0-1.1.1.2.1.1 0-1.3 2-1.3.1.1.2.1.1.1 4-1.3.1.1.2.1.2.1 7-1.3.1.1.2 9-1.3.1.1.2.2.1.1 11-1.3.1.1.2.2.2.1 16-1.3.1.1.1 16-1.3.1.1.1.r 17-1.3.1.1 19-1.3.1.1.3.3 20-1.3.1.1.3 21-1.3.1.1.3.2.1.1 23-1.3.1.1.3.2.1.1 24-1.3.1.1.3.2 25-1.3.1.2.1 25-1.3.1.2.1.r 26-1.3.1.2 28-1.3.1.2.3.1.1.2 29-1.3.1.2.3 33-1.3.1.3.1.1.1 35-1.3.1.3.1.2.1 42-1.2 43-1 44-1.1.1.1 44-1.1.1.1.r 51-1.1.1.2.1 52-1.1.1.2.1 53-1.1 54-1.1.1 55-1.1.1.3.1 56-1.1.1.3 (i / investigate-01~e.43 :ARG1 (p2 / possible-01~e.53 :ARG1 (p / produce-01~e.54 :mode~e.44 interrogative~e.44 :ARG0 (c / cooperate-01 :ARG0 (g4 / growth-factor~e.51,52 :ARG0-of (c2 / cause-01~e.0 :ARG1 (d / disease :wiki "Cancer" :name (n7 / name :op1 "cancer"))))) :ARG1 (a / affect-01~e.56 :ARG2 (s / synergize-01~e.55)))) :degree (f / further~e.42) :ARG1-of (c4 / cause-01~e.0 :ARG0 (a2 / and :op1 (m / manage-01~e.17 :polarity~e.16 -~e.16 :ARG0 (a3 / and~e.7 :op1 (g / gene :name (n2 / name :op1 "BRAF"~e.2) :ARG2-of (m2 / mutate-01 :value "V600E"~e.4)) :op2 (g2 / gene :name (n3 / name :op1 "KRAS"~e.9) :ARG2-of (m3 / mutate-01 :value "G12V"~e.11)) :ARG0-of c2) :ARG1 (t / transform-01~e.20 :ARG0 a3 :ARG1 (c3 / cell-line~e.24 :name (n4 / name :op1 "Caco-2"~e.21,23)) :degree (f2 / full~e.19))) :op2 (i2 / induce-01~e.26 :polarity~e.25 -~e.25 :ARG0 a3 :ARG2 (p4 / phenotype~e.29 :mod (e / event :name (n5 / name :op1 "epithelial−mesenchymal" :op2 "transition"~e.28)))) :ARG1-of (c8 / contrast-01 :ARG2 (g3 / gene :name (n6 / name :op1 "HRAS"~e.33) :ARG2-of (m4 / mutate-01 :value "G12V"~e.35)))))) # ::id a_pmid_2194_3101.232 ::amr-annotator SDL-AMR-09 ::preferred # ::tok The previously established oncogenic models of BRAF @ V600E @ and KRAS @ G12V @ along with the parental Caco @-@ 2 cells were treated with Transforming Growth Factor beta @-@ 1 ( TGFβ-1 ) for 14 days . # ::alignments 1-1.1.1.3.1 2-1.1.1.3 3-1.1.1.2 3-1.1.1.2.1.2.1 4-1.1.1 4-1.1.2 5-1.1.1.1.r 6-1.1.1.1.1.1 8-1.1.1.1.2.1 10-1.1 11-1.1.2.1.1.1 13-1.1.2.1.2.1 18-1.1.3.2 19-1.1.3.1.1 21-1.1.3.1.1 22-1.1.3 24-1 25-1.2.r 26-1.2.1.1 27-1.2.1.2 28-1.2.1.3 29-1.2.1.4 31-1.2.1.4 35-1.3.r 36-1.3.1 37-1.3.2 (t / treat-04~e.24 :ARG1 (a / and~e.10 :op1 (m / model~e.4 :poss~e.5 (g / gene :name (n2 / name :op1 "BRAF"~e.6) :ARG2-of (m2 / mutate-01 :value "V600E"~e.8)) :ARG0-of (c / cause-01~e.3 :ARG1 (d2 / disease :wiki "Cancer" :name (n5 / name :op1 "cancer"~e.3))) :ARG1-of (e / establish-01~e.2 :time (p3 / previous~e.1))) :op2 (m3 / model~e.4 :poss (g2 / gene :name (n3 / name :op1 "KRAS"~e.11) :ARG2-of (m4 / mutate-01 :value "G12V"~e.13)) :ARG1-of e :ARG0-of c) :op3 (c2 / cell-line~e.22 :name (n4 / name :op1 "Caco-2"~e.19,21) :mod (p2 / parent~e.18))) :ARG2~e.25 (p / protein :name (n / name :op1 "Transforming"~e.26 :op2 "Growth"~e.27 :op3 "Factor"~e.28 :op4 "beta-1"~e.29,31)) :duration~e.35 (t2 / temporal-quantity :quant 14~e.36 :unit (d / day~e.37))) # ::id a_pmid_2194_3101.233 ::amr-annotator SDL-AMR-09 ::preferred # ::tok Staining with phalloidin revealed significant morphological changes in TGFβ-1 treated Caco @-@ K15 cells that were not observed in Caco @-@ 2 cells following treatment with TGFβ-1 , while no morphological changes were recorded in TGFβ-1 treated Caco @-@ BR13 cells ( Figure 9A ) . # ::alignments 0-1.1.1 1-1.1.1.1.r 2-1.1.1.1.1.1 3-1.1 4-1.1.2.4 5-1.1.2.2 6-1.1.2 7-1.1.2.1.r 8-1.1.2.1.2.1.1.1 9-1.1.2.1.2 10-1.1.2.1.1.1 12-1.1.2.1.1.1 13-1.1.2.1 13-1.1.2.3.2 16-1.1.2.3.1 16-1.1.2.3.1.r 17-1.1.2.3 19-1.1.2.3.2.1.1 21-1.1.2.3.2.1.1 22-1.1.2.1 23-1.1.2.3.3 24-1.1.2.3.3.1 25-1.1.2.3.3.1 26-1.1.2.3.3.1 28-1 29-1.2.1.1 29-1.2.1.1.r 30-1.2.1.3 31-1.2.1 33-1.2 34-1.2.1.2.r 35-1.2.1.2.2 36-1.2.1.2.2 37-1.2.1.2.1.1 39-1.2.1.2.1.1 40-1.2.1.2 42-1.3.1 44-1.3.1.1 (c4 / contrast-01~e.28 :ARG1 (r / reveal-01~e.3 :ARG0 (s / stain-01~e.0 :ARG2~e.1 (s3 / small-molecule :name (n / name :op1 "phalloidin"~e.2))) :ARG1 (c / change-01~e.6 :ARG1~e.7 (c2 / cell-line~e.13,22 :name (n2 / name :op1 "Caco-K15"~e.10,12) :ARG1-of (t / treat-04~e.9 :ARG2 (p / protein :name (n3 / name :op1 "TGFβ-1"~e.8)))) :mod (m2 / morphological~e.5) :ARG1-of (o / observe-01~e.17 :polarity~e.16 -~e.16 :location (c3 / cell-line~e.13 :name (n4 / name :op1 "Caco-2"~e.19,21)) :ARG1-of (f / follow-01~e.23 :ARG2 t~e.24,25,26)) :ARG1-of (s2 / significant-02~e.4))) :ARG2 (r2 / record-01~e.33 :ARG1 (c5 / change-01~e.31 :polarity~e.29 -~e.29 :ARG1~e.34 (c6 / cell-line~e.40 :name (n5 / name :op1 "Caco-BR13"~e.37,39) :ARG1-of t~e.35,36) :mod m2~e.30)) :ARG1-of (d / describe-01 :ARG0 (f2 / figure~e.42 :mod "9A"~e.44))) # ::id a_pmid_2194_3101.234 ::amr-annotator SDL-AMR-09 ::preferred # ::tok Protein analysis for E @-@ cadherin , in fractionized soluble ( intracellular ) and insoluble ( bound @-@ membrane E @-@ cadherin ) extracts indicated a reduction of E @-@ cadherin in the insoluble fraction in Caco @-@ 2 and Caco @-@ K15 cells to a greater extend ( Figure 9B , upper panel ) . # ::alignments 0-1.1.2 1-1.1 2-1.1.3.r 3-1.1.3.1.1 5-1.1.3.1.1 9-1.1.1.2.1 11-1.1.1.1.2 13-1.1.1 14-1.1.1.1.1 14-1.1.1.2.1 14-1.1.1.2.1.1 14-1.1.1.2.1.1.r 16-1.1.1.2.2.1.2.1 18-1.1.1.2.2.1.2 19-1.1.1.2.2.1.1.1 21-1.1.1.2.2.1.1.1 23-1.1.1.1 23-1.1.1.2 24-1 26-1.2 28-1.1.1.2.2.1.1.1 28-1.1.3.1.1 30-1.1.1.2.2.1.1.1 30-1.1.3.1.1 33-1.1.1.2.1 33-1.1.1.2.1.1 33-1.1.1.2.1.1.r 34-1.1.1.3 34-1.2.3.1 34-1.2.3.2 35-1.2.3.r 36-1.2.3.1.1.1.1 36-1.2.3.2.1.1.1 38-1.2.3.1.1.1.1 39-1.2.3 40-1.2.3.1.1.1.1 40-1.2.3.2.1.1.1 42-1.2.3.2.1.1.1 43-1.2.3.1.1 43-1.2.3.2.1 44-1.2.2.r 46-1.2.2 46-1.2.2.1 46-1.2.2.1.r 49-1.3.1 51-1.3.1.1 54-1.3.1.2.1 55-1.3.1.2 (i / indicate-01~e.24 :ARG0 (a / analyze-01~e.1 :ARG1 (a2 / and~e.13 :op1 (e / extract~e.23 :mod (s / soluble~e.14) :location (c / cell~e.11)) :op2 (e2 / extract~e.23 :mod (s2 / soluble~e.9,14,33 :polarity~e.14,33 -~e.14,33) :ARG1-of (m / mean-01 :ARG2 (p2 / protein :name (n / name :op1 "E-cadherin"~e.19,21,28,30) :mod (m2 / membrane~e.18 :ARG1-of (b / bind-01~e.16))))) :ARG1-of (f / fraction-01~e.34)) :mod (p / protein~e.0) :purpose~e.2 (p3 / protein :name (n2 / name :op1 "E-cadherin"~e.3,5,28,30))) :ARG1 (r / reduce-01~e.26 :ARG1 p3 :ARG2~e.44 (g / great~e.46 :degree~e.46 (m3 / more~e.46)) :location~e.35 (a3 / and~e.39 :op1 (f2 / fraction~e.34 :part-of (c2 / cell-line~e.43 :name (n3 / name :op1 "Caco-2"~e.36,38,40)) :mod s2) :op2 (f3 / fraction~e.34 :part-of (c3 / cell-line~e.43 :name (n4 / name :op1 "Caco-K15"~e.36,40,42)) :mod s2))) :ARG1-of (d / describe-01 :ARG0 (f4 / figure~e.49 :mod "9B"~e.51 :location (p4 / panel~e.55 :mod (u / upper~e.54))))) # ::id a_pmid_2194_3101.235 ::amr-annotator SDL-AMR-09 ::preferred # ::tok Interestingly , even though levels of E @-@ cadherin were not altered in Caco @-@ BR13 cells , confocal images clearly presented disrupted cell @-@ cell contacts and discontinuous staining which weakens cell junctions allowing cell migration ( Figure 9B , lower panel @-@ arrows ) . # ::alignments 0-1.5 0-1.5.r 2-1.8.r 3-1.8.r 4-1.8.2 5-1.8.2.1.r 6-1.8.2.1.1.1 8-1.8.2.1.1.1 10-1.8.1 10-1.8.1.r 11-1.8 12-1.8.3.r 13-1.8.3.1.1 15-1.8.3.1.1 16-1.8.3 18-1.1.1 19-1.1 20-1.7 21-1 22-1.2.1.3 23-1.2.1.1 25-1.2.1.1 26-1.2.1 27-1.2 28-1.2.2.1 28-1.2.2.1.1 28-1.2.2.1.1.r 29-1.2.2 31-1.3 32-1.3.1.1 33-1.3.1 34-1.4 35-1.4.1.1 36-1.4.1 38-1.6.1 40-1.6.1.1 43-1.6.1.2.1 43-1.6.1.2.1.1 43-1.6.1.2.1.1.r 44-1.6.1.2 46-1.6.1.2 (p / present-102~e.21 :ARG0 (i / image~e.19 :mod (c / confocal~e.18)) :ARG1 (a / and~e.27 :op1 (c2 / contact-01~e.26 :ARG0 (c3 / cell~e.23,25) :ARG1 c3 :ARG1-of (d / disrupt-01~e.22)) :op2 (s / stain-01~e.29 :ARG1-of (c4 / continue-01~e.28 :polarity~e.28 -~e.28))) :ARG0-of (w / weaken-01~e.31 :ARG1 (j / junction~e.33 :mod c3~e.32)) :ARG0-of (a2 / allow-01~e.34 :ARG1 (m / migrate-01~e.36 :ARG0 c3~e.35)) :manner~e.0 (i2 / interesting~e.0) :ARG1-of (d2 / describe-01 :ARG0 (f / figure~e.38 :mod "9B"~e.40 :location (p3 / panel-arrow~e.44,46 :ARG1-of (l2 / low-04~e.43 :degree~e.43 (m2 / more~e.43))))) :ARG1-of (c5 / clear-06~e.20) :concession~e.2,3 (a3 / alter-01~e.11 :polarity~e.10 -~e.10 :ARG1 (l / level~e.4 :quant-of~e.5 (p2 / protein :name (n / name :op1 "E-cadherin"~e.6,8))) :location~e.12 (c6 / cell-line~e.16 :name (n2 / name :op1 "Caco-BR13"~e.13,15)))) # ::id a_pmid_2194_3101.236 ::amr-annotator SDL-AMR-09 ::preferred # ::tok Altered localization of E @-@ cadherin is an important mechanism contributing to cell metastasis [ @ 35 @ ] . # ::alignments 0-1.3.2 1-1.3 2-1.3.1.r 3-1.3.1.1.1 5-1.3.1.1.1 6-1.3.r 8-1.1 9-1 10-1.2 11-1.2.1.r 12-1.2.1.1 13-1.2.1 16-1.4.1.1.1 (m / mechanism~e.9 :mod (i / important~e.8) :ARG0-of (c / contribute-01~e.10 :ARG2~e.11 (m2 / metastasize-101~e.13 :ARG1 (c2 / cell~e.12))) :domain~e.6 (l / localize-01~e.1 :ARG1~e.2 (p / protein :name (n / name :op1 "E-cadherin"~e.3,5)) :ARG1-of (a / alter-01~e.0)) :ARG1-of (d / describe-01 :ARG0 (p2 / publication :ARG1-of (c3 / cite-01 :ARG2 35~e.16)))) # ::id a_pmid_2194_3101.237 ::amr-annotator SDL-AMR-09 ::preferred # ::tok TGFβ-1 was also investigated for its potential effect on cell migration and invasion . # ::alignments 0-1.1.1.1 2-1.3 3-1 4-1.2.r 5-1.2.1 5-1.2.1.r 6-1.2.3 7-1.2 8-1.2.2.r 9-1.2.2.1.1 10-1.2.2.1 11-1.2.2 12-1.2.2.2 (i / investigate-01~e.3 :ARG1 (p / protein :name (n / name :op1 "TGFβ-1"~e.0)) :ARG2~e.4 (a / affect-01~e.7 :ARG0~e.5 p~e.5 :ARG1~e.8 (a2 / and~e.11 :op1 (m / migrate-01~e.10 :ARG0 (c / cell~e.9)) :op2 (i2 / invade-01~e.12 :ARG0 c)) :mod (p2 / potential~e.6)) :mod (a3 / also~e.2)) # ::id a_pmid_2194_3101.238 ::amr-annotator SDL-AMR-09 ::preferred # ::tok Treatment with TGFβ-1 increased the capacity of Caco @-@ BR13 cells to invade in vitro , while no effect in the migrating ability of these cells was recorded ( Figure 9C ) . # ::alignments 0-1.1.1 1-1.1.1.2.r 2-1.1.1.2.1.1 3-1.1 5-1.1.2 6-1.1.2.2.r 7-1.1.2.2.1 8-1.1.2.2.1 9-1.1.2.2.1 10-1.1.2.2.1 12-1.1.2.2 14-1.1.2.2.2 15-1.1.2.2.2 18-1 19-1.2.1.1 19-1.2.1.1.r 20-1.2.1 21-1.1.2.2.2 21-1.2.1.2.r 23-1.2.1.2.2 24-1.2.1.2 27-1.2.1.2.1 29-1.2 31-1.3.1 33-1.3.1.1 (c / contrast-01~e.18 :ARG1 (i / increase-01~e.3 :ARG0 (t / treat-04~e.0 :ARG1 c3 :ARG2~e.1 (p / protein :name (n / name :op1 "TGFβ-1"~e.2))) :ARG1 (c2 / capable-01~e.5 :ARG1 (c3 / cell-line :name (n2 / name :op1 "Caco-BR13")) :ARG2~e.6 (i2 / invade-01~e.12 :ARG0 c3~e.7,8,9,10 :manner (i3 / in-vitro~e.14,15,21)))) :ARG2 (r / record-01~e.29 :ARG1 (a / affect-01~e.20 :polarity~e.19 -~e.19 :ARG1~e.21 (c4 / capable-01~e.24 :ARG1 c3~e.27 :ARG2 (m / migrate-01~e.23 :ARG0 c3)))) :ARG1-of (d / describe-01 :ARG0 (f / figure~e.31 :mod "9C"~e.33))) # ::id a_pmid_2194_3101.239 ::amr-annotator SDL-AMR-09 ::preferred # ::tok This enhanced invasive capacity of Caco @-@ BR13 cells is independent of their cell proliferation ( Additional Figure 5 ) . # ::alignments 0-1.2.4 1-1.2.3 2-1.2.2 3-1.2 4-1.2.1.r 5-1.2.1.1.1 7-1.2.1.1.1 8-1.2.1 10-1 10-1.1 10-1.1.r 13-1.3.1 14-1.3 17-1.4.1 19-1.4.1.1 (d / depend-01~e.10 :polarity~e.10 -~e.10 :ARG0 (c / capable-01~e.3 :ARG1~e.4 (c2 / cell-line~e.8 :name (n / name :op1 "Caco-BR13"~e.5,7)) :ARG2 (i / invade-01~e.2 :ARG0 c2) :ARG1-of (e / enhance-01~e.1) :mod (t / this~e.0)) :ARG1 (p / proliferate-01~e.14 :ARG0 c2~e.13) :ARG1-of (d2 / describe-01 :ARG0 (f / figure~e.17 :mod 5~e.19 :ARG1-of (a / add-02)))) # ::id a_pmid_2194_3101.240 ::amr-annotator SDL-AMR-09 ::preferred # ::tok In contrast , cell migration and invasion of Caco @-@ 2 and Caco @-@ K15 cells were not affected by TGFβ-1 treatment , although KRAS @ G12V @ -@ transfected cells acquired a more elongated morphology and slightly downregulated E @-@ cadherin . # ::alignments 1-1 3-1.1.3.1.1.1 3-1.1.3.1.1.2 4-1.1.3.1 5-1.1.3 5-1.1.3.1.1 6-1.1.3.2 7-1.1.3.2.1.r 8-1.1.3.2.1 9-1.1.3.2.1 10-1.1.3.2.1 11-1.1.3.2.1 12-1.1.3.2.1 13-1.1.3.2.1 14-1.1.3.2.1 15-1.1.3.2.1 17-1.1.1 17-1.1.1.r 18-1.1 19-1.1.2.r 20-1.1.2.1.1.1 21-1.1.2 23-1.1.4.r 24-1.1.4.1.1.1.1.1.1 26-1.1.4.1.1.1.1.2.1 29-1.1.4.1.1.1 30-1.1.4.1.1 31-1.1.4.1 33-1.1.4.1.2.1.1 34-1.1.4.1.2.1 35-1.1.4.1.2 36-1.1.4 37-1.1.4.2.3 38-1.1.4.2 39-1.1.4.2.1.1.1 41-1.1.4.2.1.1.1 (c / contrast-01~e.1 :ARG2 (a / affect-01~e.18 :polarity~e.17 -~e.17 :ARG0~e.19 (t3 / treat-04~e.21 :ARG2 (p3 / protein :name (n6 / name :op1 "TGFβ-1"~e.20))) :ARG1 (a2 / and~e.5 :op1 (m / migrate-01~e.4 :ARG0 (a3 / and~e.5 :op1 (c2 / cell-line~e.3 :name (n / name :op1 "Caco-2")) :op2 (c3 / cell-line~e.3 :name (n2 / name :op1 "Caco-K15")))) :op2 (i / invade-01~e.6 :ARG0~e.7 a3~e.8,9,10,11,12,13,14,15)) :concession~e.23 (a5 / and~e.36 :op1 (a6 / acquire-01~e.31 :ARG0 (c4 / cell~e.30 :ARG1-of (t / transfect-01~e.29 :ARG2 (e2 / enzyme :name (n4 / name :op1 "KRAS"~e.24) :ARG2-of (m2 / mutate-01 :value "G12V"~e.26)))) :ARG1 (m3 / morphology~e.35 :ARG1-of (e / elongate-01~e.34 :degree (m4 / more~e.33)))) :op2 (d / downregulate-01~e.38 :ARG1 (p2 / protein :name (n5 / name :op1 "E-cadherin"~e.39,41)) :ARG2 c4 :degree (s / slight~e.37))))) # ::id a_pmid_2194_3101.241 ::amr-annotator SDL-AMR-09 ::preferred # ::tok Taken together , these results suggest that TGFβ-1 can synergise with KRAS @ G12V @ and BRAF @ V600E @ oncogenes to provide Caco @-@ 2 cells with a more transforming phenotype . # ::alignments 0-1.1.3 1-1.1.3.1 3-1.1.2 4-1.1 4-1.1.1 4-1.1.1.r 5-1 6-1.2.r 7-1.2.1.1.1.1 8-1.2 11-1.2.1.2.1.1.1 13-1.2.1.2.1.2.1 15-1.2.1.2 16-1.2.1.2.2.1.1 18-1.2.1.2.2.2.1 21-1.2.1.2.3 22-1.2.1.3 23-1.2.1.3.3.1.1 25-1.2.1.3.3.1.1 26-1.2.1.3.3 27-1.2.1.3.2.r 29-1.2.1.3.2.1.1 30-1.2.1.3.2.1 31-1.2.1.3.2 (s / suggest-01~e.5 :ARG0 (t / thing~e.4 :ARG2-of~e.4 (r / result-01~e.4) :mod (t2 / this~e.3) :ARG1-of (t4 / take-01~e.0 :mod (t5 / together~e.1))) :ARG1~e.6 (p3 / possible-01~e.8 :ARG1 (s2 / synergize-01 :ARG0 (g / gene :name (n / name :op1 "TGFβ-1"~e.7)) :ARG1 (a / and~e.15 :op1 (g2 / gene :name (n2 / name :op1 "KRAS"~e.11) :ARG2-of (m / mutate-01 :value "G12V"~e.13)) :op2 (g3 / gene :name (n3 / name :op1 "BRAF"~e.16) :ARG2-of (m2 / mutate-01 :value "V600E"~e.18)) :ARG0-of (c / cause-01~e.21 :ARG1 (d / disease :wiki "Cancer" :name (n5 / name :op1 "cancer")))) :ARG2 (p / provide-01~e.22 :ARG0 g :ARG1~e.27 (p2 / phenotype~e.31 :ARG1-of (t3 / transform-01~e.30 :degree (m3 / more~e.29))) :ARG2 (c2 / cell-line~e.26 :name (n4 / name :op1 "Caco-2"~e.23,25)))))) # ::id a_pmid_2194_3101.242 ::amr-annotator SDL-AMR-09 ::preferred # ::tok According to previous studies , the mutation in the C @-@ terminal domain of Smad4 , D351H , that is present in Caco @-@ 2 cells , results in complete Smad4 inactivation [ @ 36 @ ] . # ::alignments 0-1.3 1-1.3 2-1.3.1.1 3-1.3.1 3-1.3.1.2 3-1.3.1.2.r 6-1.1 7-1.1.2.r 9-1.1.2.1.1 11-1.1.2.1.1 14-1.1.2.2.1.1 16-1.1.1 21-1.1.3.r 22-1.1.3.1.1 24-1.1.3.1.1 25-1.1.3 27-1 28-1.2.r 29-1.2.2 30-1.1.2.2.1.1 31-1.2 31-1.2.1 31-1.2.1.r 34-1.4.1.1.1 (r / result-01~e.27 :ARG1 (m / mutate-01~e.6 :value "D351H"~e.16 :ARG1~e.7 (p / protein-segment :name (n / name :op1 "C-terminus"~e.9,11) :part-of (p2 / protein :name (n2 / name :op1 "Smad4"~e.14,30))) :location~e.21 (c2 / cell-line~e.25 :name (n4 / name :op1 "Caco-2"~e.22,24))) :ARG2~e.28 (a / activate-01~e.31 :polarity~e.31 -~e.31 :ARG1-of (c / complete-02~e.29)) :ARG1-of (s / say-01~e.0,1 :ARG0 (t / thing~e.3 :time (p3 / previous~e.2) :ARG1-of~e.3 (s2 / study-01~e.3))) :ARG1-of (d3 / describe-01 :ARG0 (p4 / publication :ARG1-of (c3 / cite-01 :ARG2 36~e.34)))) # ::id a_pmid_2194_3101.243 ::amr-annotator SDL-AMR-09 ::preferred # ::tok However , TGFβ-1 has been shown to act through alternative non @-@ Smad pathways , such as Rho GTPases and MAPK [ @ 37 @ - @ 39 @ ] . # ::alignments 0-1 2-1.1.1.1.1.1 5-1.1 7-1.1.1 9-1.1.1.2.3 10-1.1.1.2.1 10-1.1.1.2.1.r 12-1.1.1.2.2.1 13-1.1.1.2 13-1.1.1.2.4.1 13-1.1.1.2.4.2 15-1.1.1.2.4.r 16-1.1.1.2.4.r 17-1.1.1.2.4.1.1.1 19-1.1.1.2.4 20-1.1.1.2.4.2.1.1 23-1.1.2.1.1.1.1 27-1.1.2.1.1.1.2 (h / have-concession-91~e.0 :ARG1 (s / show-01~e.5 :ARG1 (a / act-01~e.7 :ARG0 (p / protein :name (n / name :op1 "TGFβ-1"~e.2)) :manner (p7 / pathway~e.13 :polarity~e.10 -~e.10 :name (n5 / name :op1 "Smad"~e.12) :mod (a3 / alternative~e.9) :example~e.15,16 (a2 / and~e.19 :op1 (p4 / pathway~e.13 :name (n3 / name :op1 "Rho"~e.17 :op2 "GTPase")) :op2 (p5 / pathway~e.13 :name (n4 / name :op1 "MAPK"~e.20))))) :ARG1-of (d / describe-01 :ARG0 (p6 / publication :ARG1-of (c2 / cite-01 :ARG2 (v / value-interval :op1 37~e.23 :op2 39~e.27)))))) # ::id a_pmid_2194_3101.244 ::amr-annotator SDL-AMR-09 ::preferred # ::tok Indeed , following TGFβ-1 treatment , enhanced activity for RhoA GTPase as well as pERK1 @/@ 2 was recorded in Caco @-@ 2 , Caco @-@ K15 and Caco @-@ BR13 cells . # ::alignments 0-1.4 2-1.3 3-1.3.1.1.1.1 4-1.3.1 6-1.1.2 7-1.1 8-1.1.1.r 9-1.1.1.1.1.1 10-1.1.1.1.1.2 11-1.1.1 12-1.1.1 13-1.2 16-1.1.1.2.1.1 16-1.2.1.1.1 18-1 20-1.2.1.1.1 20-1.2.2.1.1 20-1.2.3.1.1 22-1.2.1.1.1 24-1.2.1.1.1 24-1.2.2.1.1 24-1.2.3.1.1 26-1.2.2.1.1 27-1.2 28-1.2.1.1.1 28-1.2.2.1.1 28-1.2.3.1.1 30-1.2.3.1.1 31-1.2.1 31-1.2.2 31-1.2.3 (r / record-01~e.18 :ARG1 (a / activity-06~e.7 :ARG0~e.8 (a2 / and~e.11,12 :op1 (p / pathway :name (n / name :op1 "RhoA"~e.9 :op2 "GTPase"~e.10)) :op2 (e2 / enzyme :name (n2 / name :op1 "ERK1/2"~e.16) :ARG3-of (p3 / phosphorylate-01))) :ARG1-of (e / enhance-01~e.6)) :location (a3 / and~e.13,27 :op1 (c / cell-line~e.31 :name (n3 / name :op1 "Caco-2"~e.16,20,22,24,28)) :op2 (c2 / cell-line~e.31 :name (n4 / name :op1 "Caco-K15"~e.20,24,26,28)) :op3 (c3 / cell-line~e.31 :name (n5 / name :op1 "Caco-BR13"~e.20,24,28,30))) :ARG1-of (f / follow-01~e.2 :ARG2 (t / treat-04~e.4 :ARG2 (p2 / protein :name (n6 / name :op1 "TGFβ-1"~e.3)))) :mod (i / indeed~e.0)) # ::id a_pmid_2194_3101.245 ::amr-annotator SDL-AMR-09 ::preferred # ::tok Based on these observations other than non @-@ Smad signaling like RhoA GTPase and pERK1 @/@ 2 pathways can be regulated by TGF @-@ beta , to induce the morphological changes observed in the Caco @-@ 2 transformed and parental cells ( Figure 9D ) . # ::alignments 0-1.3 1-1.3.1.r 2-1.3.1.1 3-1.3.1 6-1.1.2.1.1.1 6-1.1.2.1.1.1.r 8-1.1.2.1.1.2.1 9-1.1.2.1.1.3 10-1.1.2.1.1.4.r 11-1.1.2.1.1.4.1.1.1 12-1.1.2.1.1.4.1.1.2 13-1.1.2.1.1.4 14-1.1.2.1.1.4.2.1.1 16-1.1.2.1.1.4.2.1.1 17-1.1.2 18-1 20-1.1 21-1.1.1.r 22-1.1.1.1.1 24-1.1.1.1.1 27-1.1.3 29-1.1.3.2.2 30-1.1.3.2 31-1.1.3.2.1 32-1.1.3.2.1.1.r 34-1.1.3.2.1.1.1.1.1 34-1.1.3.2.1.1.2.1.1 36-1.1.3.2.1.1.1.1.1 36-1.1.3.2.1.1.2.1.1 37-1.1.3.2.1.1.1.2 38-1.1.3.2.1.1 39-1.1.3.2.1.1.2.2 40-1.1.3.2.1.1.1 40-1.1.3.2.1.1.2 42-1.2.1 44-1.2.1.1 (p / possible-01~e.18 :ARG1 (r / regulate-01~e.20 :ARG0~e.21 (p2 / protein :name (n / name :op1 "TGF-beta"~e.22,24)) :ARG1 (p7 / pathway~e.17 :ARG2-of (e / except-01 :ARG1 (p4 / pathway :polarity~e.6 -~e.6 :name (n4 / name :op1 "Smad"~e.8) :ARG0-of (s / signal-07~e.9) :example~e.10 (a2 / and~e.13 :op1 (p5 / pathway :name (n5 / name :op1 "RhoA"~e.11 :op2 "GTPase"~e.12)) :op2 (p6 / pathway :name (n6 / name :op1 "pERK1/2"~e.14,16)))))) :purpose (i / induce-01~e.27 :ARG0 r :ARG2 (c / change-01~e.30 :ARG1-of (o / observe-01~e.31 :location~e.32 (a / and~e.38 :op1 (c2 / cell-line~e.40 :name (n2 / name :op1 "Caco-2"~e.34,36) :ARG1-of (t / transform-01~e.37)) :op2 (c3 / cell-line~e.40 :name (n3 / name :op1 "Caco-2"~e.34,36) :mod (p3 / parent~e.39)))) :mod (m / morphological~e.29)))) :ARG1-of (d / describe-01 :ARG0 (f / figure~e.42 :mod "9D"~e.44)) :ARG1-of (b / base-02~e.0 :ARG2~e.1 (o2 / observe-01~e.3 :mod (t2 / this~e.2)))) # ::id a_pmid_2256_9000.96 ::amr-annotator SDL-AMR-09 ::preferred # ::tok In @-@ vitro inhibition of cell proliferation by selumetinib treatment in NSCLC and CRC cell lines # ::alignments 1-1.4 3-1.4 5-1 6-1.2.r 7-1.2.1 8-1.2 9-1.1.r 10-1.1.1.1.1 11-1.1 12-1.3.r 12-1.4 13-1.3.1.1.1.1 14-1.3 15-1.3.2.1.1.1 16-1.3.1 16-1.3.2 17-1.3.1 (i / inhibit-01~e.5 :ARG0~e.9 (t / treat-04~e.11 :ARG2 (s / small-molecule :name (n / name :op1 "selumetinib"~e.10))) :ARG1~e.6 (p / proliferate-01~e.8 :ARG0 (c / cell~e.7)) :location~e.12 (a / and~e.14 :op1 (c2 / cell-line~e.16,17 :mod (d / disease :name (n2 / name :op1 "NSCLC"~e.13))) :op2 (c3 / cell-line~e.16 :mod (d2 / disease :name (n3 / name :op1 "CRC"~e.15)))) :manner (i2 / in-vitro~e.1,3,12)) # ::id a_pmid_2256_9000.97 ::amr-annotator SDL-AMR-09 ::preferred # ::tok We first evaluated the sensitivity to the selective MEK1 @/@ 2 inhibitor , selumetinib , in a panel of five NSCLC ( GLC82 , H460 , A549 , H1299 , Calu3 ) and six CRC ( GEO , HCT15 , HCT116 , SW480 , SW620 , LS174T ) cell lines by using the MTT assay . # ::alignments 0-1.1 1-1.3 2-1 4-1.2 8-1.2.1.2.1.1.1 10-1.2.1.2.1.1.1 11-1.2.1 11-1.2.1.2 11-1.2.1.2.r 13-1.2.1.1.1 15-1.4.r 17-1.4 18-1.4.1.r 19-1.4.1.1.1 20-1.4.1.1.3.1.1 22-1.4.1.1.2.1.1.1.1 24-1.4.1.1.2.1.2.1.1 26-1.4.1.1.2.1.3.1.1 28-1.4.1.1.2.1.4.1.1 30-1.4.1.1.2.1.5.1.1 32-1.4.1 32-1.4.1.1.2.1 33-1.4.1.2.1 34-1.4.1.2.3.1.1 36-1.4.1.2.2.1.1.1.1 38-1.4.1.2.2.1.2.1.1 40-1.4.1.2.2.1.3.1.1 42-1.4.1.2.2.1.4.1.1 44-1.4.1.2.2.1.5.1.1 46-1.4.1.2.2.1.6.1.1 48-1.4.1.1 48-1.4.1.2 49-1.4.1.2 50-1.5.r 51-1.5 53-1.5.2.1.1.1 54-1.5.2 (e / evaluate-01~e.2 :ARG0 (w / we~e.0) :ARG1 (s / sensitive-03~e.4 :ARG1 (s2 / small-molecule~e.11 :name (n / name :op1 "selumetinib"~e.13) :ARG0-of~e.11 (i / inhibit-01~e.11 :ARG1 (e2 / enzyme :name (n2 / name :op1 "MEK1/2"~e.8,10))) :ARG0-of (s3 / select-01))) :mod (f / first~e.1) :location~e.15 (p / panel~e.17 :consist-of~e.18 (a / and~e.32 :op1 (c / cell-line~e.48 :quant 5~e.19 :ARG1-of (m / mean-01 :ARG2 (a2 / and~e.32 :op1 (c2 / cell-line :name (n4 / name :op1 "GLC82"~e.22)) :op2 (c3 / cell-line :name (n5 / name :op1 "H460"~e.24)) :op3 (c4 / cell-line :name (n6 / name :op1 "A549"~e.26)) :op4 (c5 / cell-line :name (n7 / name :op1 "H1299"~e.28)) :op5 (c6 / cell-line :name (n8 / name :op1 "Calu3"~e.30)))) :mod (d / disease :name (n3 / name :op1 "NSCLC"~e.20))) :op2 (c7 / cell-line~e.48,49 :quant 6~e.33 :ARG1-of (m2 / mean-01 :ARG2 (a3 / and :op1 (c8 / cell-line :name (n10 / name :op1 "GEO"~e.36)) :op2 (c9 / cell-line :name (n11 / name :op1 "HCT15"~e.38)) :op3 (c10 / cell-line :name (n12 / name :op1 "HCT116"~e.40)) :op4 (c11 / cell-line :name (n13 / name :op1 "SW480"~e.42)) :op5 (c12 / cell-line :name (n14 / name :op1 "SW620"~e.44)) :op6 (c13 / cell-line :name (n15 / name :op1 "LS174T"~e.46)))) :mod (d2 / disease :name (n9 / name :op1 "CRC"~e.34))))) :manner~e.50 (u / use-01~e.51 :ARG0 w :ARG1 (a4 / assay-01~e.54 :instrument (s4 / small-molecule :name (n16 / name :op1 "MTT"~e.53))))) # ::id a_pmid_2256_9000.98 ::amr-annotator SDL-AMR-09 ::preferred # ::tok Cancer cells were treated with selumetinib at concentrations ranging from 0.01 to 10 μ @ ℳ for 48 , 72 , and 96 h . # ::alignments 0-1.1.1.2.1 1-1.1 3-1 4-1.2.r 5-1.2.1.1 7-1.2.2 7-1.2.2.1.1 7-1.2.2.1.2 8-1.2.2.1 10-1.2.2.1.1.1 12-1.2.2.1.2.1 17-1.3.r 18-1.3.1.1 20-1.3.1.2 22-1.3.1 23-1.3.1.3 24-1.3.2 (t / treat-04~e.3 :ARG1 (c / cell~e.1 :mod (d / disease :wiki "Cancer" :name (n / name :op1 "cancer"~e.0))) :ARG2~e.4 (s / small-molecule :name (n2 / name :op1 "selumetinib"~e.5) :quant (c2 / concentration~e.7 :ARG1-of (r / range-01~e.8 :ARG3 (c3 / concentration-quantity~e.7 :quant 0.01~e.10 :unit (m2 / micromolar)) :ARG4 (c4 / concentration-quantity~e.7 :quant 10~e.12 :unit m2)))) :duration~e.17 (t2 / temporal-quantity :quant (a / and~e.22 :op1 48~e.18 :op2 72~e.20 :op3 96~e.23) :unit (h / hour~e.24))) # ::id a_pmid_2256_9000.99 ::amr-annotator SDL-AMR-09 ::preferred # ::tok As shown in Figure 1 , there was a wide range of sensitivity , with IC @ 50 values varying between 0.01 and > 10 μ @ ℳ. # ::alignments 1-1.2 4-1.2.1 5-1.2.1.1 11-1.1.1 12-1.1 14-1 17-1.1.2.1.1.1 19-1.1.2.1.1.1.1 21-1.1.2.1.1 22-1.1.2.1 24-1.1.2.1.2.1 26-1.1.2.1.3 27-1.1.2.1.3.1.1 (s / sensitive-03~e.14 :ARG1-of (r / range-01~e.12 :ARG1-of (w / wide-02~e.11) :ARG1-of (m / mean-01 :ARG2 (v / vary-01~e.22 :ARG1 (v2 / value~e.21 :ARG4-of (h / have-percentage-maximal-inhibitory-concentration-01~e.17 :ARG2 50~e.19)) :ARG3 (c / concentration-quantity :quant 0.01~e.24 :unit (m2 / micromolar)) :ARG4 (m3 / more-than~e.26 :op1 (c2 / concentration-quantity :quant 10~e.27 :unit (m4 / micromolar)))))) :ARG1-of (s2 / show-01~e.1 :ARG0 (f / figure~e.4 :mod 1~e.5))) # ::id a_pmid_2256_9000.100 ::amr-annotator SDL-AMR-09 ::preferred # ::tok We classified as sensitive ( S ) or resistant ( R ) the cancer cell lines according to selumetinib IC @ 50 at 96 h ⩽ 1 or > 1 μ @ ℳ, respectively . # ::alignments 0-1.1.1 1-1.1 1-1.2 2-1.1.3.r 3-1.1.3 7-1.1.4.3 8-1.2.3 13-1.1.2.1.2.1 14-1.1.2 15-1.1.2 18-1.1.4.2.1.1 19-1.1.4 19-1.2.4 21-1.1.4.1 21-1.2.4.1 24-1.1.4.4.1.1 25-1.1.4.4.1.2 27-1.1.4.3.1.1 27-1.1.4.3.2.1.1 28-1.1.4.3 29-1.2.4.3 30-1.1.4.3.1.1 30-1.1.4.3.2.1.1 30-1.2.4.3.1.1 (a / and :op1 (c / classify-01~e.1 :ARG0 (w / we~e.0) :ARG1 (c2 / cell-line~e.14,15 :mod (d / disease :wiki "Cancer" :name (n / name :op1 "cancer"~e.13))) :ARG2~e.2 (s / sensitive-03~e.3 :ARG0 c2) :condition (h2 / have-percentage-maximal-inhibitory-concentration-01~e.19 :ARG2 50~e.21 :ARG1 (s2 / small-molecule :name (n3 / name :op1 "selumetinib"~e.18)) :ARG4 (o / or~e.7,28 :op1 (c3 / concentration-quantity :quant 1~e.27,30 :unit (m3 / micromolar)) :op2 (l / less-than :op1 (c4 / concentration-quantity :quant 1~e.27,30 :unit (m / micromolar)))) :time (a2 / after :op1 (t2 / temporal-quantity :quant 96~e.24 :unit (h / hour~e.25))))) :op2 (c5 / classify-01~e.1 :ARG0 w :ARG1 c2 :ARG2 (r / resist-01~e.8 :ARG0 c2) :condition (h3 / have-percentage-maximal-inhibitory-concentration-01~e.19 :ARG2 50~e.21 :ARG1 s2 :ARG4 (m2 / more-than~e.29 :op1 (c7 / concentration-quantity :quant 1~e.30 :unit (m4 / micromolar))) :time a2))) # ::id a_pmid_2256_9000.101 ::amr-annotator SDL-AMR-09 ::preferred # ::tok This concentration was chosen based on the data from phase I studies , which indicated that 1 μ @ ℳ was within the average plasma concentrations of selumetinib achieved in patients at the maximum tolerated dose for this agent ( Adjei et al , 2008 ) . # ::alignments 0-1.1.1 1-1.1 1-1.2.1.2.1.1 3-1 4-1.2 5-1.2.1.r 7-1.2.1 8-1.2.1.1.r 9-1.2.1.1.1 10-1.2.1.1.1.1.1 11-1.2.1.1 14-1.2.1.2 16-1.2.1.2.1.1.1 22-1.2.1.2.1.1.1.r 24-1.2.1.2.1.2.3 25-1.2.1.2.1.2.2 26-1.2.1.2.1.2 27-1.2.1.2.1.2.1.r 28-1.2.1.2.1.2.1.1.1 29-1.2.1.2.1.2.4 30-1.2.1.2.1.2.4.1.r 31-1.2.1.2.1.2.4.1 32-1.2.1.2.1.2.5.r 34-1.2.1.2.1.2.5.1 35-1.2.1.2.1.2.5.2 36-1.2.1.2.1.2.5 37-1.2.1.2.1.2.5.2.1.r 38-1.2.1.2.1.2.5.2.1.1 39-1.2.1.2.1.2.5.2.1 42-1.3.1.1.1.1.1 44-1.3.1.1 45-1.3.1.1.2.1 48-1.3.1.2.1 (c / choose-01~e.3 :ARG1 (c2 / concentrate-02~e.1 :mod (t / this~e.0)) :ARG1-of (b / base-02~e.4 :ARG2~e.5 (d / data~e.7 :source~e.8 (s / study-01~e.11 :mod (p / phase~e.9 :ord (o / ordinal-entity :value 1~e.10))) :ARG0-of (i / indicate-01~e.14 :ARG1 (i2 / include-01 :ARG1 (c3 / concentration-quantity~e.1 :quant~e.22 1~e.16 :unit (m / micromolar)) :ARG2 (c4 / concentrate-02~e.26 :ARG1~e.27 (s2 / small-molecule :name (n / name :op1 "selumetinib"~e.28)) :location (p2 / plasma~e.25) :ARG1-of (a / average-04~e.24) :ARG1-of (a2 / achieve-01~e.29 :location~e.30 (p3 / patient~e.31)) :quant~e.32 (d2 / dose~e.36 :quant (m2 / maximum~e.34) :ARG1-of (t2 / tolerate-01~e.35 :topic~e.37 (a3 / agent~e.39 :mod (t3 / this~e.38))))))))) :ARG1-of (d3 / describe-01 :ARG0 (p4 / publication-91 :ARG0 (a4 / and~e.44 :op1 (p5 / person :name (n2 / name :op1 "Adjei"~e.42)) :op2 (p6 / person :mod (o2 / other~e.45))) :time (d4 / date-entity :year 2008~e.48)))) # ::id a_pmid_2256_9000.102 ::amr-annotator SDL-AMR-09 ::preferred # ::tok Overall , 67 % ( four of six ) CRC and 40 % ( two of five ) NSCLC cell lines had a selumetinib IC @ 50 of ⩽ 1 μ @ ℳ. # ::alignments 0-1.3 2-1.1.1.2.2.1 3-1.1.1.2.2 5-1.1.1.1 7-1.1.1.2.1.1 9-1.1.1.2.1.2.1.1 10-1.1 11-1.1.2.2.2.1 12-1.1.2.2.2 14-1.1.2.1 16-1.1.2.2.1.1 18-1.1.2.2.1.2.1.1 19-1.1.1 19-1.1.1.2.1 19-1.1.2 19-1.1.2.2.1 20-1.1.2.2.1 21-1 23-1.2.1.1 24-1.2 24-1.2.2 24-1.2.2.r 26-1.2.2.1 30-1.2.2.2.1.2.1 30-1.2.2.2.1.1 (h / have-03~e.21 :ARG0 (a / and~e.10 :op1 (c / cell-line~e.19 :quant 4~e.5 :ARG1-of (i / include-91 :ARG2 (c2 / cell-line~e.19 :quant 6~e.7 :mod (d2 / disease :name (n / name :op1 "CRC"~e.9))) :ARG3 (p / percentage-entity~e.3 :value 67~e.2))) :op2 (c3 / cell-line~e.19 :quant 2~e.14 :ARG1-of (i2 / include-91 :ARG2 (c4 / cell-line~e.19,20 :quant 5~e.16 :mod (d / disease :name (n3 / name :op1 "NSCLC"~e.18))) :ARG3 (p2 / percentage-entity~e.12 :value 40~e.11)))) :ARG1 (s / small-molecule~e.24 :name (n6 / name :op1 "selumetinib"~e.23) :ARG1-of~e.24 (h2 / have-percentage-maximal-inhibitory-concentration-01~e.24 :ARG2 50~e.26 :ARG4 (c5 / concentration-quantity :quant (o / or :op2 1~e.30 :op1 (l / less-than :op1 1~e.30)) :unit (m / micromolar)))) :mod (o2 / overall~e.0)) # ::id a_pmid_2256_9000.103 ::amr-annotator SDL-AMR-09 ::preferred # ::tok In particular , among the sensitive cancer cell lines there were four , two NSCLC ( Calu3 and H1299 ) and two CRC ( HCT116 and SW620 ) , extremely sensitive to selumetinib with an IC @ 50 of ⩽ 0.01 μ @ ℳ ( Figure 1A and B ) . # ::alignments 1-1.3 3-1 3-1.1.4 5-1.1.3 6-1.1.2.2.1 7-1.1 7-1.2 8-1.1 8-1.1.4.1.1 8-1.1.4.1.2 11-1.1.1 13-1.1.4.1.2.1 14-1.1.4.1.1.3.1.1 16-1.1.4.1.1.2.1.1.1.1 17-1.1.4.1.1.2.1 18-1.1.4.1.1.2.1.2.1.1 21-1.1.4.1.1.1 22-1.1.4.1.2.3.1.1 24-1.1.4.1.2.2.1.1.1.1 26-1.1.4.1.2.2.1.2.1.1 29-1.1.4.1.3.2 30-1.1.4.1.3 31-1.1.4.1.3.1.r 32-1.1.4.1.3.1.1.1 35-1.1.4.1.3.1 35-1.1.4.1.3.1.2 35-1.1.4.1.3.1.2.r 37-1.1.4.1.3.1.2.1 41-1.1.4.1.3.1.2.2.1.2.1 41-1.1.4.1.3.1.2.2.1.1 48-1.4.1.1 48-1.4.1.2 49-1.4.1.1.1 50-1.4.1 (i / include-91~e.3 :ARG1 (c / cell-line~e.7,8 :quant 4~e.11 :mod (d / disease :wiki "Cancer" :name (n / name :op1 "cancer"~e.6)) :ARG0-of (s / sensitive-03~e.5) :ARG2-of (i2 / include-91~e.3 :ARG1 (a / and :op1 (c2 / cell-line~e.8 :quant 2~e.21 :ARG1-of (m / mean-01 :ARG2 (a2 / and~e.17 :op1 (c3 / cell-line :name (n3 / name :op1 "Calu3"~e.16)) :op2 (c4 / cell-line :name (n4 / name :op1 "H1299"~e.18)))) :mod (d3 / disease :name (n2 / name :op1 "NSCLC"~e.14))) :op2 (c5 / cell-line~e.8 :quant 2~e.13 :ARG1-of (m2 / mean-01 :ARG2 (a3 / and :op1 (c6 / cell-line :name (n6 / name :op1 "HCT116"~e.24)) :op2 (c7 / cell-line :name (n7 / name :op1 "SW620"~e.26)))) :mod (d4 / disease :name (n5 / name :op1 "CRC"~e.22))) :ARG0-of (s2 / sensitive-03~e.30 :ARG1~e.31 (s3 / small-molecule~e.35 :name (n8 / name :op1 "selumetinib"~e.32) :ARG1-of~e.35 (h / have-percentage-maximal-inhibitory-concentration-01~e.35 :ARG2 50~e.37 :ARG4 (c8 / concentration-quantity :quant (o / or :op2 0.01~e.41 :op1 (l / less-than :op1 0.01~e.41)) :unit (m3 / micromolar)))) :degree (e2 / extreme~e.29))))) :ARG2 (c9 / cell-line~e.7 :mod d :ARG0-of s) :mod (p / particular~e.1) :ARG1-of (d2 / describe-01 :ARG0 (a4 / and~e.50 :op1 (f / figure~e.48 :mod "1A"~e.49) :op2 (f2 / figure~e.48 :mod "1B")))) # ::id a_pmid_2256_9000.104 ::amr-annotator SDL-AMR-09 ::preferred # ::tok To investigate the mechanisms underlying the different sensitivities to the drug , we conducted experiments on a group of four cancer cell lines representing both selumetinib @-@ sensitive ( HCT116 and Calu3 ) and selumetinib @-@ resistant ( HCT15 and H460 ) models . # ::alignments 1-1.3 3-1.3.2 4-1.3.2.1 6-1.3.2.1.1.2 7-1.3.2.1.1 8-1.3.2.1.1.1.r 10-1.3.2.1.1.1 12-1.1 13-1 14-1.2 15-1.2.2.r 17-1.2.2 18-1.2.2.1.r 19-1.2.2.1.1 20-1.2.2.1.2.2.1 21-1.2.2.1 22-1.2.2.1 23-1.2.2.1.3 25-1.2.2.1.3.1.1.1.1.1.1 27-1.2.2.1.3.1.1.1 29-1.2.2.1.3.1.1.2.1.1.1.1 30-1.2.2.1.3.1.1.2.1 31-1.2.2.1.3.1.1.2.1.2.1.1 34-1.2.2.1.3.1.2.1.1 36-1.2.2.1.3.1.2.1 38-1.2.2.1.3.1.2.2.1.1.1.1 40-1.2.2.1.3.1.2.2.1.2.1.1 42-1.2.2.1.3.1.1 42-1.2.2.1.3.1.2 (c / conduct-01~e.13 :ARG0 (w / we~e.12) :ARG1 (e / experiment-01~e.14 :ARG0 w :ARG1~e.15 (g / group~e.17 :consist-of~e.18 (c2 / cell-line~e.21,22 :quant 4~e.19 :mod (d / disease :wiki "Cancer" :name (n / name :op1 "cancer"~e.20)) :ARG0-of (r / represent-01~e.23 :ARG1 (a / and :op1 (m / model-01~e.42 :ARG0-of (s / sensitive-03~e.27 :ARG1 (s2 / small-molecule :name (n2 / name :op1 "selumetinib"~e.25))) :ARG1-of (m2 / mean-01 :ARG2 (a2 / and~e.30 :op1 (c3 / cell-line :name (n3 / name :op1 "HCT116"~e.29)) :op2 (c4 / cell-line :name (n4 / name :op1 "Calu3"~e.31))))) :op2 (m3 / model-01~e.42 :ARG0-of (r2 / resist-01~e.36 :ARG1 s2~e.34) :ARG1-of (m4 / mean-01 :ARG2 (a3 / and :op1 (c5 / cell-line :name (n5 / name :op1 "HCT15"~e.38)) :op2 (c6 / cell-line :name (n6 / name :op1 "H460"~e.40)))))))))) :purpose (i / investigate-01~e.1 :ARG0 w :ARG1 (m5 / mechanism~e.3 :ARG0-of (u / underlie-01~e.4 :ARG1 (s3 / sensitive-03~e.7 :ARG1~e.8 (d2 / drug~e.10) :ARG1-of (d3 / differ-02~e.6)))))) # ::id a_pmid_2256_9000.105 ::amr-annotator SDL-AMR-09 ::preferred # ::tok Effects of selumetinib treatment on cell @-@ cycle distribution in NSCLC and CRC cell lines # ::alignments 0-1 1-1.1.r 2-1.1.1.1.1 3-1.1 4-1.2.r 5-1.2.1.1 7-1.2.1 8-1.2 9-1.3.r 10-1.3.1.1.1.1 11-1.3 12-1.3.2.1.1.1 13-1.3.1 13-1.3.2 14-1.3.1 (a / affect-01~e.0 :ARG0~e.1 (t / treat-04~e.3 :ARG2 (s / small-molecule :name (n / name :op1 "selumetinib"~e.2))) :ARG1~e.4 (d / distribute-01~e.8 :ARG1 (c / cycle-02~e.7 :ARG1 (c2 / cell~e.5))) :location~e.9 (a2 / and~e.11 :op1 (c3 / cell-line~e.13,14 :mod (d2 / disease :name (n2 / name :op1 "NSCLC"~e.10))) :op2 (c4 / cell-line~e.13 :mod (d3 / disease :name (n3 / name :op1 "CRC"~e.12))))) # ::id a_pmid_2256_9000.106 ::amr-annotator SDL-AMR-09 ::preferred # ::tok We conducted flow cytometric analysis to compare the cell @-@ cycle distribution following treatment of the selumetinib @-@ sensitive HCT116 and Calu3 cancer cell lines and of the selumetinib @-@ resistant HCT15 and H460 cancer cell lines . # ::alignments 0-1.1 1-1 2-1.2.1.1 4-1.2 6-1.3 8-1.3.2.1.1 10-1.3.2.1 11-1.3.2 12-1.3.2.2 13-1.3.2.2.1 14-1.3.2.2.1.1.r 16-1.3.2.2.1.1.1.4.1.1.1 18-1.3.2.2.1.1.1.4 19-1.3.2.2.1.1.1.1.1.1 20-1.3.2.2.1.1.1 21-1.3.2.2.1.1.1.2.1.1 22-1.3.2.2.1.1.1.3.2.1 23-1.3.2.2.1.1.1.1 24-1.3.2.2.1.1.1.1 24-1.3.2.2.1.1.1.2 24-1.3.2.2.1.1.2.2 25-1.3.2.2.1.1.1 28-1.3.2.2.1.1.2.4.1 30-1.3.2.2.1.1.2.4 31-1.3.2.2.1.1.2.1.1.1 32-1.3.2.2.1.1.2 33-1.3.2.2.1.1.2.2.1.1 34-1.3.2.2.1.1.1.3.2.1 35-1.3.2.2.1.1.2.1 36-1.3.2.2.1.1.2.1 (c / conduct-01~e.1 :ARG0 (w / we~e.0) :ARG1 (a / analyze-01~e.4 :manner (c9 / cytometry :mod (f2 / flow~e.2))) :purpose (c2 / compare-01~e.6 :ARG0 w :ARG1 (d / distribute-01~e.11 :ARG1 (c3 / cycle-02~e.10 :ARG1 (c4 / cell~e.8)) :ARG1-of (f / follow-01~e.12 :ARG2 (t / treat-04~e.13 :ARG1~e.14 (a2 / and :op1 (a3 / and~e.20,25 :op1 (c5 / cell-line~e.23,24 :name (n2 / name :op1 "HCT116"~e.19)) :op2 (c6 / cell-line~e.24 :name (n3 / name :op1 "Calu3"~e.21)) :mod (d2 / disease :wiki "Cancer" :name (n4 / name :op1 "cancer"~e.22,34)) :ARG0-of (s / sensitive-03~e.18 :ARG1 (s2 / small-molecule :name (n5 / name :op1 "selumetinib"~e.16)))) :op2 (a4 / and~e.32 :op1 (c7 / cell-line~e.35,36 :name (n6 / name :op1 "HCT15"~e.31)) :op2 (c8 / cell-line~e.24 :name (n7 / name :op1 "H460"~e.33)) :mod d2 :ARG0-of (r / resist-01~e.30 :ARG1 s2~e.28)))))))) # ::id a_pmid_2256_9000.107 ::amr-annotator SDL-AMR-09 ::preferred # ::tok Cancer cells were treated with selumetinib at the IC @ 50 values for inhibition of cell proliferation for 24 , 48 , and 72 h . # ::alignments 0-1.1.1.2.1 1-1.1 3-1 4-1.2.r 5-1.2.1.1 8-1.2 8-1.2.2 8-1.2.2.r 10-1.2.2.1 13-1.2.2.2.r 14-1.2.2.2 15-1.2.2.2.1.r 16-1.2.2.2.1.1 17-1.2.2.2.1 18-1.3.r 19-1.3.1.1 21-1.3.1.2 23-1.3.1 24-1.3.1.3 25-1.3.2 (t / treat-04~e.3 :ARG1 (c / cell~e.1 :mod (d / disease :wiki "Cancer" :name (n / name :op1 "cancer"~e.0))) :ARG2~e.4 (s / small-molecule~e.8 :name (n2 / name :op1 "selumetinib"~e.5) :ARG1-of~e.8 (h2 / have-percentage-maximal-inhibitory-concentration-01~e.8 :ARG2 50~e.10 :ARG3~e.13 (i / inhibit-01~e.14 :ARG1~e.15 (p / proliferate-01~e.17 :ARG0 (c2 / cell~e.16))))) :duration~e.18 (t3 / temporal-quantity :quant (a / and~e.23 :op1 24~e.19 :op2 48~e.21 :op3 72~e.24) :unit (h / hour~e.25))) # ::id a_pmid_2256_9000.108 ::amr-annotator SDL-AMR-09 ::preferred # ::tok Twenty @-@ four hours selumetinib treatment caused cell accumulation in the G1 phase and concomitant reduction in the S phase as compared with controls in selumetinib @-@ sensitive cancer cell lines ( Figure 2A ) . # ::alignments 3-1.1.2.2 4-1.1.1.1.1 5-1.1 6-1 7-1.2.1.1 8-1.2.1 9-1.2.1.2.r 11-1.2.1.2.1.1 12-1.2.1.2 13-1.2 14-1.2.2.2 15-1.2.2 18-1.2.2.3.1.1 19-1.2.2.3 20-1.2.2.3.r 20-1.2.r 21-1.2.3.r 23-1.2.3 23-1.2.3.1 23-1.2.3.1.r 24-1.2.3.2.r 25-1.2.3.2.2.1 27-1.2.3.2.2 28-1.2.3.2.1.2.1 29-1.2.3.2 30-1.2.3.2 33-1.3.1 34-1.3.1.1 (c / cause-01~e.6 :ARG0 (t / treat-04~e.5 :ARG2 (s / small-molecule :name (n / name :op1 "selumetinib"~e.4)) :duration (t2 / temporal-quantity :quant 24 :unit (h / hour~e.3))) :ARG1~e.20 (a / and~e.13 :op1 (a2 / accumulate-01~e.8 :ARG1 (c2 / cell~e.7) :time~e.9 (p / phase~e.12 :name (n2 / name :op1 "G1"~e.11))) :op2 (r / reduce-01~e.15 :ARG1 c2 :mod (c3 / concomitant~e.14) :time~e.20 (p2 / phase~e.19 :name (n3 / name :op1 "S"~e.18))) :compared-to~e.21 (m / molecular-physical-entity~e.23 :ARG2-of~e.23 (c4 / control-01~e.23) :location~e.24 (c5 / cell-line~e.29,30 :mod (d / disease :wiki "Cancer" :name (n4 / name :op1 "cancer"~e.28)) :ARG0-of (s2 / sensitive-03~e.27 :ARG1 s~e.25)))) :ARG1-of (d2 / describe-01 :ARG0 (f / figure~e.33 :mod "2A"~e.34))) # ::id a_pmid_2256_9000.109 ::amr-annotator SDL-AMR-09 ::preferred # ::tok The arrest in the G1 phase was significantly increased with longer ( 48 and 72 h ) treatment with selumetinib in both HCT116 and Calu3 cells . # ::alignments 1-1.1 2-1.1.1.r 4-1.1.1.1.1 5-1.1.1 7-1.2 8-1 9-1.3.r 10-1.3.2 10-1.3.2.1 10-1.3.2.1.r 12-1.3.2.2.1.1.1 13-1.3.2.2.1 14-1.3.2.2.1.2.1 15-1.3.2.2.1.1.2 15-1.3.2.2.1.2.2 17-1.3 18-1.3.1.r 18-1.3.r 19-1.3.1.1.1 22-1.4.1.1.1 23-1.4 24-1.4.2.1.1 25-1.4.1 25-1.4.2 (i / increase-01~e.8 :ARG1 (a / arrest-02~e.1 :time~e.2 (p / phase~e.5 :name (n / name :op1 "G1"~e.4))) :ARG2 (s / significant-02~e.7) :instrument~e.9,18 (t / treat-04~e.17 :ARG2~e.18 (s2 / small-molecule :name (n2 / name :op1 "selumetinib"~e.19)) :ARG1-of (l / long-03~e.10 :degree~e.10 (m / more~e.10) :ARG1-of (m2 / mean-01 :ARG2 (a4 / and~e.13 :op1 (t2 / temporal-quantity :quant 48~e.12 :unit (h / hour~e.15)) :op2 (t3 / temporal-quantity :quant 72~e.14 :unit (h2 / hour~e.15)))))) :location (a3 / and~e.23 :op1 (c / cell-line~e.25 :name (n3 / name :op1 "HCT116"~e.22)) :op2 (c2 / cell-line~e.25 :name (n4 / name :op1 "Calu3"~e.24)))) # ::id a_pmid_2256_9000.110 ::amr-annotator SDL-AMR-09 ::preferred # ::tok This effect was also paralleled by a time @-@ dependent reduction in the S phase in both selumetinib @-@ sensitive cancer cell lines ( Figure 2A ) . # ::alignments 0-1.1.1 1-1.1 3-1.3 4-1 5-1.2.r 7-1.2.1.1 9-1.2.1 10-1.2 11-1.2.2.r 13-1.2.2.1.1 14-1.2.2 15-1.2.3.r 16-1.2.3.3 17-1.2.3.2.1.1.1 19-1.2.3.2 20-1.2.3.1.2.1 21-1.2.3 22-1.2.3 25-1.4.1 26-1.4.1.1 (p / parallel-01~e.4 :ARG0 (a / affect-01~e.1 :mod (t / this~e.0)) :ARG1~e.5 (r / reduce-01~e.10 :ARG0-of (d / depend-01~e.9 :ARG1 (t2 / time~e.7)) :time~e.11 (p2 / phase~e.14 :name (n / name :op1 "S"~e.13)) :location~e.15 (c / cell-line~e.21,22 :mod (d2 / disease :wiki "Cancer" :name (n2 / name :op1 "cancer"~e.20)) :ARG0-of (s / sensitive-03~e.19 :ARG1 (s2 / small-molecule :name (n3 / name :op1 "selumetinib"~e.17))) :mod (b / both~e.16))) :mod (a2 / also~e.3) :ARG1-of (d3 / describe-01 :ARG0 (f / figure~e.25 :mod "2A"~e.26))) # ::id a_pmid_2256_9000.111 ::amr-annotator SDL-AMR-09 ::preferred # ::tok In contrast , no effect was observed on cell @-@ cycle distribution in the two selumetinib @-@ resistant HCT15 and H460 cells ( Figure 2A ) . # ::alignments 1-1 3-1.1.1.1 3-1.1.1.1.r 4-1.1.1 6-1.1 7-1.1.1.2.r 8-1.1.1.2.1.1 10-1.1.1.2.1 11-1.1.1.2 12-1.1.2.r 14-1.1.2.1 15-1.1.2.4.1.1.1 17-1.1.2.4 18-1.1.2.2.1.1 19-1.1.2 20-1.1.2.3.1.1 21-1.1.2.2 21-1.1.2.3 24-1.2.1 25-1.2.1.1 (c / contrast-01~e.1 :ARG2 (o / observe-01~e.6 :ARG1 (a / affect-01~e.4 :polarity~e.3 -~e.3 :ARG1~e.7 (d / distribute-01~e.11 :ARG1 (c2 / cycle-02~e.10 :ARG1 (c3 / cell~e.8)))) :location~e.12 (a2 / and~e.19 :quant 2~e.14 :op1 (c4 / cell-line~e.21 :name (n / name :op1 "HCT15"~e.18)) :op2 (c5 / cell-line~e.21 :name (n2 / name :op1 "H460"~e.20)) :ARG0-of (r / resist-01~e.17 :ARG1 (s / small-molecule :name (n3 / name :op1 "selumetinib"~e.15))))) :ARG1-of (d2 / describe-01 :ARG0 (f / figure~e.24 :mod "2A"~e.25))) # ::id a_pmid_2256_9000.112 ::amr-annotator SDL-AMR-09 ::preferred # ::tok These results were supported also by Garon et al ( 2010 ) , in which the block in G1 phase inducted by selumetinib is evident only in sensitive cell lines . # ::alignments 0-1.2.2 1-1.2 1-1.2.1 1-1.2.1.r 3-1 4-1.3 7-1.1.1.1.1.1 9-1.1.1 10-1.1.1.2.1 13-1.1.2.1 20-1.1.3.1 21-1.1.3.1.1.r 22-1.1.3.1.1.1.1 23-1.1.3.1.1 26-1.1.3.1.2.1.1.1 27-1.1.3.1.r 28-1.1.3 29-1.1.3.2.2 31-1.1.3.2.1 32-1.1.3.2 33-1.1.3.2 (s / support-01~e.3 :ARG0 (p / publication-91 :ARG0 (a / and~e.9 :op1 (p2 / person :name (n / name :op1 "Garon"~e.7)) :op2 (p3 / person :mod (o / other~e.10))) :time (d / date-entity :year 2010~e.13) :location-of (e / evident~e.28 :domain~e.27 (b / block-01~e.20 :time~e.21 (p4 / phase~e.23 :name (n2 / name :op1 "G1"~e.22)) :ARG2-of (i / induce-01 :ARG0 (s2 / small-molecule :name (n3 / name :op1 "selumetinib"~e.26)))) :location (c / cell-line~e.32,33 :ARG0-of (s3 / sensitive-03~e.31) :mod (o2 / only~e.29)))) :ARG1 (t / thing~e.1 :ARG2-of~e.1 (r / result-01~e.1) :mod (t2 / this~e.0)) :mod (a2 / also~e.4)) # ::id a_pmid_2256_9000.113 ::amr-annotator SDL-AMR-09 ::preferred # ::tok Effects of selumetinib treatment on the induction of apoptosis in NSCLC and CRC cell lines # ::alignments 0-1 1-1.1.r 2-1.1.1.1.1 3-1.1 4-1.2.r 6-1.2 7-1.2.1.r 8-1.2.1 9-1.3.r 10-1.3.1.1.1.1 11-1.3 12-1.3.2.1.1.1 13-1.3.1 13-1.3.2 14-1.3.1 (a / affect-01~e.0 :ARG0~e.1 (t / treat-04~e.3 :ARG2 (s / small-molecule :name (n / name :op1 "selumetinib"~e.2))) :ARG1~e.4 (i / induce-01~e.6 :ARG2~e.7 (a2 / apoptosis~e.8)) :location~e.9 (a3 / and~e.11 :op1 (c / cell-line~e.13,14 :mod (d / disease :name (n2 / name :op1 "NSCLC"~e.10))) :op2 (c2 / cell-line~e.13 :mod (d2 / disease :name (n3 / name :op1 "CRC"~e.12))))) # ::id a_pmid_2256_9000.114 ::amr-annotator SDL-AMR-09 ::preferred # ::tok Several preclinical models have demonstrated that selumetinib act as a cytotoxic drug rather than cytostatic by inducing proapoptotic activity ( Huynh et al , 2007a ; Huynh et al , 2007b ; Meng et al , 2009 ; Meng et al , 2010 ) . # ::alignments 0-1.1.2 1-1.1.1 2-1.1 4-1 5-1.2.r 6-1.2.1.1.1 7-1.2 8-1.2.2.r 8-1.3.1.1.2.r 8-1.3.1.2.2.r 8-1.3.1.4.2.r 10-1.2.2.1 11-1.2.2 11-1.2.2.2.1 12-1.2.2.2 14-1.2.2.2.1.1 15-1.2.3.r 16-1.2.3 18-1.2.3.1 21-1.3.1.1.1.1.1.1 23-1.3.1.1.1 24-1.3.1.1.1.2.1 31-1.3.1.1.1.1.1.1 33-1.3.1.1.1 34-1.3.1.1.1.2.1 41-1.3.1.3.1.1.1.1 43-1.3.1 43-1.3.1.1.1 43-1.3.1.3.1 44-1.3.1.1.1.2.1 47-1.3.1.3.2.1 51-1.3.1.3.1.1.1.1 53-1.3.1 53-1.3.1.1.1 53-1.3.1.3.1 54-1.3.1.1.1.2.1 57-1.3.1.4.2.1 (d / demonstrate-01~e.4 :ARG0 (m / model-01~e.2 :mod (p / preclinical~e.1) :quant (s / several~e.0)) :ARG1~e.5 (a / act-01~e.7 :ARG0 (s2 / small-molecule :name (n / name :op1 "selumetinib"~e.6)) :ARG1~e.8 (d2 / drug~e.11 :mod (c / cytotoxic~e.10) :ARG1-of (i / instead-of-91~e.12 :ARG2 (d3 / drug~e.11 :mod (c2 / cytostatic~e.14)))) :manner~e.15 (i2 / induce-01~e.16 :ARG2 (a2 / activity-06~e.18 :ARG0-of (f / favor-01 :ARG1 (a3 / apoptosis))))) :ARG1-of (d4 / describe-01 :ARG0 (a4 / and~e.43,53 :op1 (p2 / publication-91 :ARG0 (a5 / and~e.23,33,43,53 :op1 (p3 / person :name (n2 / name :op1 "Huynh"~e.21,31)) :op2 (p4 / person :mod (o / other~e.24,34,44,54))) :time~e.8 (d5 / date-entity :year 2007)) :op2 (p5 / publication-91 :ARG0 a5 :time~e.8 d5) :op3 (p6 / publication-91 :ARG0 (a6 / and~e.43,53 :op1 (p7 / person :name (n3 / name :op1 "Meng"~e.41,51)) :op2 p4) :time (d6 / date-entity :year 2009~e.47)) :op4 (p8 / publication-91 :ARG0 a6 :time~e.8 (d7 / date-entity :year 2010~e.57))))) # ::id a_pmid_2256_9000.115 ::amr-annotator SDL-AMR-09 ::preferred # ::tok To confirm this effect , the induction of apoptosis was evaluated using an FACS @-@ based assay for Annexin V and PI staining . # ::alignments 1-1.3 2-1.3.1.1 3-1.3.1 6-1.1 7-1.1.1.r 8-1.1.1 10-1 11-1.2 11-1.3.r 13-1.2.1.1.1.1.1 15-1.2.1.1 16-1.2.1 17-1.2.2.r 18-1.2.2.1.1.1.1 19-1.2.2.1.1.1.2 20-1.2.2.1 21-1.2.2.1.2.1.1 22-1.2.2 (e / evaluate-01~e.10 :ARG1 (i / induce-01~e.6 :ARG2~e.7 (a / apoptosis~e.8)) :manner (u / use-01~e.11 :ARG1 (a2 / assay-01~e.16 :ARG1-of (b / base-02~e.15 :ARG2 (t / thing :name (n / name :op1 "FACS"~e.13)))) :purpose~e.17 (s / stain-01~e.22 :ARG1 (a3 / and~e.20 :op1 (p / protein :name (n2 / name :op1 "Annexin"~e.18 :op2 "V"~e.19)) :op2 (s2 / small-molecule :name (n3 / name :op1 "PI"~e.21))))) :purpose~e.11 (c / confirm-01~e.1 :ARG1 (a4 / affect-01~e.3 :mod (t2 / this~e.2)))) # ::id a_pmid_2256_9000.116 ::amr-annotator SDL-AMR-09 ::preferred # ::tok After 24 h of selumetinib treatment , a significant induction of apoptosis was detected in the sensitive HCT116 and Calu3 cancer cell lines , which was maximal following 72 h of treatment ( Figure 2B ) . # ::alignments 0-1.3 1-1.3.1.2.1 2-1.3.1.2.2 4-1.1.4.1.1.1.1 5-1.3.1 8-1.1.2 9-1.1 10-1.1.1.r 11-1.1.1 13-1 14-1.2.r 16-1.2.3 17-1.2.1.1.1 18-1.2 19-1.2.2.1.1 20-1.2.4.2.1 21-1.2.1 21-1.2.2 22-1.2.1 26-1.1.3 27-1.1.4 28-1.1.4.1.2.1 29-1.1.4.1.2.2 31-1.1.4.1 31-1.3.1 34-1.4.1 35-1.4.1.1 (d / detect-01~e.13 :ARG1 (i / induce-01~e.9 :ARG2~e.10 (a / apoptosis~e.11) :ARG1-of (s / significant-02~e.8) :mod (m / maximal~e.26) :ARG1-of (f / follow-01~e.27 :ARG2 (t / treat-04~e.31 :ARG2 (s2 / small-molecule :name (n / name :op1 "selumetinib"~e.4)) :duration (t2 / temporal-quantity :quant 72~e.28 :unit (h / hour~e.29))))) :location~e.14 (a2 / and~e.18 :op1 (c / cell-line~e.21,22 :name (n2 / name :op1 "HCT116"~e.17)) :op2 (c2 / cell-line~e.21 :name (n3 / name :op1 "Calu3"~e.19)) :ARG0-of (s3 / sensitive-03~e.16) :mod (d2 / disease :wiki "Cancer" :name (n4 / name :op1 "cancer"~e.20))) :time (a3 / after~e.0 :op1 (t3 / treat-04~e.5,31 :ARG2 s2 :duration (t4 / temporal-quantity :quant 24~e.1 :unit h~e.2))) :ARG1-of (d3 / describe-01 :ARG0 (f2 / figure~e.34 :mod "2B"~e.35))) # ::id a_pmid_2256_9000.117 ::amr-annotator SDL-AMR-09 ::preferred # ::tok Moreover , apoptosis was not detectable following treatment with selumetinib in the two resistant HCT15 and H460 cancer cell lines ( Figure 2B ) . # ::alignments 0-1 2-1.1.2.1 4-1.1.1 4-1.1.1.r 5-1.1.2 6-1.1.3 7-1.1.3.1 8-1.1.3.1.1.r 9-1.1.3.1.1.1.1 10-1.1.2.2.r 12-1.1.2.2.1 13-1.1.2.2.4 14-1.1.2.2.2.1.1 15-1.1.2.2 16-1.1.2.2.3.1.1 17-1.1.2.2.5.2.1 18-1.1.2.2.2 18-1.1.2.2.3 19-1.1.2.2.2 22-1.2.1 23-1.2.1.1 (a / and~e.0 :op2 (p / possible-01 :polarity~e.4 -~e.4 :ARG1 (d / detect-01~e.5 :ARG1 (a2 / apoptosis~e.2) :location~e.10 (a3 / and~e.15 :quant 2~e.12 :op1 (c / cell-line~e.18,19 :name (n / name :op1 "HCT15"~e.14)) :op2 (c2 / cell-line~e.18 :name (n2 / name :op1 "H460"~e.16)) :ARG0-of (r / resist-01~e.13) :mod (d2 / disease :wiki "Cancer" :name (n3 / name :op1 "cancer"~e.17)))) :ARG1-of (f / follow-01~e.6 :ARG2 (t / treat-04~e.7 :ARG2~e.8 (s / small-molecule :name (n4 / name :op1 "selumetinib"~e.9))))) :ARG1-of (d3 / describe-01 :ARG0 (f2 / figure~e.22 :mod "2B"~e.23))) # ::id a_pmid_2256_9000.118 ::amr-annotator SDL-AMR-09 ::preferred # ::tok Effects of selumetinib treatment on intracellular signalling in NSCLC and CRC cell lines # ::alignments 0-1 1-1.1.r 2-1.1.1.1.1 3-1.1 4-1.2.r 5-1.2.1 6-1.2 7-1.3.r 8-1.3.1.1.1.1 9-1.3 10-1.3.2.1.1.1 11-1.3.1 11-1.3.2 12-1.3.1 (a / affect-01~e.0 :ARG0~e.1 (t / treat-04~e.3 :ARG2 (s / small-molecule :name (n / name :op1 "selumetinib"~e.2))) :ARG1~e.4 (s2 / signal-07~e.6 :mod (i / intracellular~e.5)) :location~e.7 (a2 / and~e.9 :op1 (c / cell-line~e.11,12 :mod (d / disease :name (n2 / name :op1 "NSCLC"~e.8))) :op2 (c2 / cell-line~e.11 :mod (d2 / disease :name (n3 / name :op1 "CRC"~e.10))))) # ::id a_pmid_2256_9000.119 ::amr-annotator SDL-AMR-09 ::preferred # ::tok To assess the effects of selumetinib treatment on key intracellular downstream signalling pathways , western blots were performed to assess total and phosphorylated EGFR and downstream effectors ( total MEK1 @/@ 2 , phospho @-@ MEK1 @/@ 2 , total MAPK , phospho @-@ MAPK , total AKT , phospho @-@ AKT , total 4EBP1 , and phosho @-@ 4EBP1 ) . # ::alignments 1-1.3 3-1.3.1 4-1.3.1.1.r 5-1.3.1.1.1.1.1 6-1.3.1.1 7-1.3.1.2.r 8-1.3.1.2.3 9-1.3.1.2.2 10-1.3.1.2.1.1 11-1.3.1.2.1 12-1.3.1.2 14-1.1 15-1.1 17-1 19-1.2 19-1.3 20-1.2.1.1.2 21-1.2.1 22-1.2.1.2.2 23-1.2.1.1.1.1 23-1.2.1.2.1.1 24-1.2.1 25-1.2.1.3.1 26-1.2.1.3 28-1.2.1.1.2 29-1.2.1.3.2.1.1.1.1 29-1.2.1.3.2.1.2.1.1 31-1.2.1.3.2.1.1.1.1 31-1.2.1.3.2.1.2.1.1 33-1.2.1.2.2 35-1.2.1.3.2.1.1.1.1 35-1.2.1.3.2.1.2.1.1 37-1.2.1.3.2.1.1.1.1 37-1.2.1.3.2.1.2.1.1 39-1.2.1.1.2 40-1.2.1.3.2.1.3.1.1 40-1.2.1.3.2.1.4.1.1 42-1.2.1.2.2 44-1.2.1.3.2.1.3.1.1 44-1.2.1.3.2.1.4.1.1 46-1.2.1.1.2 47-1.2.1.3.2.1.5.1.1 47-1.2.1.3.2.1.6.1.1 49-1.2.1.2.2 51-1.2.1.3.2.1.5.1.1 51-1.2.1.3.2.1.6.1.1 53-1.2.1.1.2 54-1.2.1.3.2.1.7.1.1 54-1.2.1.3.2.1.8.1.1 56-1.2.1.3.2.1 59-1.2.1.3.2.1.7.1.1 59-1.2.1.3.2.1.8.1.1 (p / perform-01~e.17 :ARG1 (i2 / immunoblot-01~e.14,15) :purpose (a / assess-01~e.19 :ARG1 (a2 / and~e.21,24 :op1 (e / enzyme :name (n2 / name :op1 "EGFR"~e.23) :mod (t2 / total-01~e.20,28,39,46,53)) :op2 (e2 / enzyme :name (n3 / name :op1 "EGFR"~e.23) :ARG3-of (p2 / phosphorylate-01~e.22,33,42,49)) :op3 (e3 / effector~e.26 :location (d / downstream~e.25) :ARG1-of (m / mean-01 :ARG2 (a3 / and~e.56 :op1 (e4 / enzyme :name (n4 / name :op1 "MEK1/2"~e.29,31,35,37) :mod t2) :op2 (e5 / enzyme :name (n5 / name :op1 "MEK1/2"~e.29,31,35,37) :ARG3-of p2) :op3 (e6 / enzyme :name (n6 / name :op1 "MAPK"~e.40,44) :mod t2) :op4 (e7 / enzyme :name (n7 / name :op1 "MAPK"~e.40,44) :ARG3-of p2) :op5 (e8 / enzyme :name (n8 / name :op1 "AKT"~e.47,51) :mod t2) :op6 (e9 / enzyme :name (n9 / name :op1 "AKT"~e.47,51) :ARG3-of p2) :op7 (p3 / protein :name (n10 / name :op1 "4EBP1"~e.54,59) :mod t2) :op8 (p4 / protein :name (n11 / name :op1 "4EBP1"~e.54,59) :ARG3-of p2)))))) :purpose (a4 / assess-01~e.1,19 :ARG1 (a5 / affect-01~e.3 :ARG0~e.4 (t3 / treat-04~e.6 :ARG2 (s / small-molecule :name (n12 / name :op1 "selumetinib"~e.5))) :ARG1~e.7 (p5 / pathway~e.12 :ARG0-of (s2 / signal-07~e.11 :direction (d2 / downstream~e.10)) :mod (i / intracellular~e.9) :ARG1-of (k / key-02~e.8))))) # ::id a_pmid_2256_9000.120 ::amr-annotator SDL-AMR-09 ::preferred # ::tok Cancer cells were evaluated at baseline and at different time points after treatment with selumetinib at 0.05 μ @ ℳ. # ::alignments 0-1.1.1.2.1 1-1.1 3-1 4-1.2.r 5-1.2.1 6-1.2 8-1.2.2.2 9-1.2.2.1 10-1.2.2 11-1.3 12-1.3.1 13-1.3.1.1.r 14-1.3.1.1.1.1 16-1.3.1.1.2.1 (e / evaluate-01~e.3 :ARG1 (c / cell~e.1 :mod (d / disease :wiki "Cancer" :name (n / name :op1 "cancer"~e.0))) :manner~e.4 (a / and~e.6 :op1 (b / baseline~e.5) :op2 (p / point~e.10 :mod (t / time~e.9) :ARG1-of (d2 / differ-02~e.8))) :time (a2 / after~e.11 :op1 (t2 / treat-04~e.12 :ARG2~e.13 (s / small-molecule :name (n2 / name :op1 "selumetinib"~e.14) :quant (c2 / concentration-quantity :quant 0.05~e.16 :unit (m / micromolar)))))) # ::id a_pmid_2256_9000.121 ::amr-annotator SDL-AMR-09 ::preferred # ::tok No change in total and phosphorylated EGFR expression was observed in both selumetinib @-@ sensitive and selumetinib @-@ resistant cancer cell lines ( Figure 3A @–@ D ) . # ::alignments 0-1.1.1 0-1.1.1.r 1-1.1 2-1.1.2.r 3-1.1.2.1.1.2 4-1.1.2.1 5-1.1.2.1.2.2 6-1.1.2.1.1.1.1 6-1.1.2.1.2.1.1 7-1.1.2 9-1 12-1.2.1.1.1.1.1 14-1.2.1.1 16-1.2.1.1.1.1.1 18-1.2.2.1 19-1.2.3.2.1 20-1.2.1 20-1.2.2 21-1.2.2 24-1.3.1.1 24-1.3.1.2 24-1.3.1.3 24-1.3.1.4 25-1.3.1.1.1 (o / observe-01~e.9 :ARG1 (c / change-01~e.1 :polarity~e.0 -~e.0 :ARG1~e.2 (e / express-03~e.7 :ARG2 (a / and~e.4 :op1 (e2 / enzyme :name (n / name :op1 "EGFR"~e.6) :mod (t / total-01~e.3)) :op2 (e3 / enzyme :name (n2 / name :op1 "EGFR"~e.6) :ARG3-of (p / phosphorylate-01~e.5))))) :location (a2 / and :op1 (c2 / cell-line~e.20 :ARG0-of (s / sensitive-03~e.14 :ARG1 (s2 / small-molecule :name (n3 / name :op1 "selumetinib"~e.12,16)))) :op2 (c3 / cell-line~e.20,21 :ARG0-of (r / resist-01~e.18 :ARG1 s2)) :mod (d / disease :wiki "Cancer" :name (n4 / name :op1 "cancer"~e.19))) :ARG1-of (d2 / describe-01 :ARG0 (a3 / and :op1 (f / figure~e.24 :mod "3A"~e.25) :op2 (f2 / figure~e.24 :mod "3B") :op3 (f3 / figure~e.24 :mod "3C") :op4 (f4 / figure~e.24 :mod "3D")))) # ::id a_pmid_2256_9000.122 ::amr-annotator SDL-AMR-09 ::preferred # ::tok Treatment with selumetinib caused a time @-@ dependent inhibition in phospho @-@ MEK1 @/@ 2 with a complete signal suppression within 60 or 15 min of treatment in HCT116 and Calu3 cells , respectively ( Figure 3A and B ) . # ::alignments 0-1.1 1-1.1.1.r 2-1.1.1.1.1 3-1 5-1.2.1.1.1 5-1.2.2.3.1.2.1 5-1.2.3.3.2.1 5-1.2.3.3.r 7-1.2.1.1 8-1.2.1 9-1.3.r 10-1.3.2 12-1.3.1.1 14-1.3.1.1 15-1.2.r 17-1.2.2.2 18-1.2.2.1 19-1.2.2 19-1.2.3 20-1.2.2.3 20-1.2.2.3.1.2 20-1.2.2.3.1.2.1.1.r 20-1.2.3.3 20-1.2.3.3.2 20-1.2.3.3.2.r 21-1.2.2.3.1.2.1.1 23-1.2.3.3.2.1.1 24-1.2.2.3.1.2.1.2 25-1.2.2.3.1.2.r 26-1.2.2.3.1 26-1.2.3.3.1 28-1.2.2.4.1.1 29-1.2 30-1.2.3.4.1.1 31-1.2.2.4 31-1.2.3.4 36-1.4.1.1 36-1.4.1.2 37-1.4.1.1.1 38-1.4.1 (c / cause-01~e.3 :ARG0 (t / treat-04~e.0 :ARG2~e.1 (s / small-molecule :name (n / name :op1 "selumetinib"~e.2))) :ARG1~e.15 (a4 / and~e.29 :op1 (i / inhibit-01~e.8 :ARG0-of (d / depend-01~e.7 :ARG1 (t2 / time~e.5))) :op2 (s2 / suppress-01~e.19 :ARG1 (s3 / signal-07~e.18) :degree (c2 / complete-01~e.17) :time (a / after~e.20 :op1 (t3 / treat-04~e.26 :ARG2 s :quant~e.25 (u / up-to~e.20 :op1 (t4 / temporal-quantity~e.5 :quant~e.20 60~e.21 :unit (m / minute~e.24))))) :location (c3 / cell-line~e.31 :name (n2 / name :op1 "HCT116"~e.28))) :op3 (s4 / suppress-01~e.19 :ARG1 s3 :degree c2 :time~e.5 (a2 / after~e.20 :op1 (t5 / treat-04~e.26 :ARG2 s) :quant~e.20 (u2 / up-to~e.20 :op1 (t6 / temporal-quantity~e.5 :quant 15~e.23 :unit m))) :location (c4 / cell-line~e.31 :name (n3 / name :op1 "Calu3"~e.30)))) :location~e.9 (e / enzyme :name (n4 / name :op1 "MEK1/2"~e.12,14) :ARG3-of (p / phosphorylate-01~e.10)) :ARG1-of (d2 / describe-01 :ARG0 (a3 / and~e.38 :op1 (f / figure~e.36 :mod "3A"~e.37) :op2 (f2 / figure~e.36 :mod "3B")))) # ::id a_pmid_2256_9000.123 ::amr-annotator SDL-AMR-09 ::preferred # ::tok This inhibition was sustained for 24 h of selumetinib treatment in both cancer cell lines . # ::alignments 0-1.1.1 1-1.1 3-1 4-1.2.r 5-1.2.2.1 6-1.2.2.2 8-1.2.1.1.1 9-1.2 10-1.3.r 11-1.3.2 12-1.3.1.2.1 13-1.3 14-1.3 (s / sustain-01~e.3 :ARG1 (i / inhibit-01~e.1 :mod (t / this~e.0)) :duration~e.4 (t2 / treat-04~e.9 :ARG2 (s2 / small-molecule :name (n / name :op1 "selumetinib"~e.8)) :duration (t3 / temporal-quantity :quant 24~e.5 :unit (h / hour~e.6))) :location~e.10 (c / cell-line~e.13,14 :mod (d / disease :wiki "Cancer" :name (n2 / name :op1 "cancer"~e.12)) :mod (b / both~e.11))) # ::id a_pmid_2256_9000.124 ::amr-annotator SDL-AMR-09 ::preferred # ::tok In contrast , MEK1 @/@ 2 inhibition of protein phosphorylation was less effective with only a slight reduction in pospho @-@ MEK1 @/@ 2 in the two selumetinib @-@ resistant HCT15 and H460 cancer cell lines ( Figure 3C and D ) . # ::alignments 1-1 3-1.1.1.1.1.1 3-1.1.2.1.1.1 5-1.1.1.1.1.1 5-1.1.2.1.1.1 6-1.1.1 8-1.1.1.2.1 9-1.1.1.2 9-1.1.2.1.2 11-1.1.3 12-1.1 13-1.1.2.r 14-1.1.2.3 16-1.1.2.2 17-1.1.2 21-1.1.2.1.1.1 23-1.1.2.1.1.1 23-1.1.2.4.1 26-1.1.2.1.1.1 26-1.1.2.4.1 27-1.1.2.4.5.1.1.1 29-1.1.2.4.5 30-1.1.2.4.2.1.1 31-1.1.2.4 32-1.1.2.4.3.1.1 33-1.1.2.4.4.2.1 34-1.1.2.4.2 34-1.1.2.4.3 35-1.1.2.4.2 38-1.2.1.1 38-1.2.1.2 39-1.2.1.1.1 40-1.2.1 (c / contrast-01~e.1 :ARG2 (e / effective-04~e.12 :ARG0 (i / inhibit-01~e.6 :ARG0 (e3 / enzyme :name (n6 / name :op1 "MEK1/2"~e.3,5)) :ARG1 (p2 / phosphorylate-01~e.9 :ARG1 (p3 / protein~e.8))) :ARG1~e.13 (r / reduce-01~e.17 :ARG1 (e2 / enzyme :name (n / name :op1 "MEK1/2"~e.3,5,21,23,26) :ARG3-of (p / phosphorylate-01~e.9)) :ARG2 (s / slight~e.16) :mod (o / only~e.14) :location (a / and~e.31 :quant 2~e.23,26 :op1 (c2 / cell-line~e.34,35 :name (n2 / name :op1 "HCT15"~e.30)) :op2 (c3 / cell-line~e.34 :name (n3 / name :op1 "H460"~e.32)) :mod (d / disease :wiki "Cancer" :name (n4 / name :op1 "cancer"~e.33)) :ARG0-of (r2 / resist-01~e.29 :ARG1 (s2 / small-molecule :name (n5 / name :op1 "selumetinib"~e.27))))) :degree (l / less~e.11)) :ARG1-of (d2 / describe-01 :ARG0 (a2 / and~e.40 :op1 (f / figure~e.38 :mod "3C"~e.39) :op2 (f2 / figure~e.38 :mod "3D")))) # ::id a_pmid_2256_9000.125 ::amr-annotator SDL-AMR-09 ::preferred # ::tok Moreover , MAPK phosphorylation was completely blocked following selumetinib treatment in HCT116 and Calu3 . # ::alignments 0-1 2-1.1.1.1.1.1 3-1.1.1 5-1.1.2 6-1.1 7-1.1.3 8-1.1.3.1.1.1.1 9-1.1.3.1 10-1.1.4.r 11-1.1.4.1.1.1 12-1.1.4 13-1.1.4.2.1.1 (a / and~e.0 :op2 (b / block-01~e.6 :ARG1 (p / phosphorylate-01~e.3 :ARG1 (e / enzyme :name (n / name :op1 "MAPK"~e.2))) :degree (c / complete-01~e.5) :ARG1-of (f / follow-01~e.7 :ARG2 (t / treat-04~e.9 :ARG2 (s / small-molecule :name (n2 / name :op1 "selumetinib"~e.8)))) :location~e.10 (a2 / and~e.12 :op1 (c2 / cell-line :name (n3 / name :op1 "HCT116"~e.11)) :op2 (c3 / cell-line :name (n4 / name :op1 "Calu3"~e.13))))) # ::id a_pmid_2256_9000.126 ::amr-annotator SDL-AMR-09 ::preferred # ::tok In contrast , in the two selumetinib @-@ resistant HCT15 and H460 cells MAPK phosphorylation was only reduced but never completely abrogated ( Figure 3C and D ) . # ::alignments 1-1 1-1.1 3-1.1.r 5-1.1.3.1 6-1.1.3.4.1.1.1 8-1.1.3.4 9-1.1.3.2.1.1 10-1.1.3 11-1.1.3.3.1.1 12-1.1.3.2 12-1.1.3.3 13-1.1.1.1.1.1.1 14-1.1.1.1 16-1.1.1.2 17-1.1.1 18-1.1 19-1.1.2.3 19-1.1.2.3.1 19-1.1.2.3.1.r 20-1.1.2.2 21-1.1.2 24-1.2.1.1 24-1.2.1.2 25-1.2.1.1.1 26-1.2.1 (c / contrast-01~e.1 :ARG2~e.3 (c2 / contrast-01~e.1,18 :ARG1 (r / reduce-01~e.17 :ARG1 (p / phosphorylate-01~e.14 :ARG1 (e / enzyme :name (n / name :op1 "MAPK"~e.13))) :mod (o / only~e.16)) :ARG2 (a2 / abrogate-01~e.21 :ARG1 p :degree (c3 / complete-01~e.20) :time (e2 / ever~e.19 :polarity~e.19 -~e.19)) :location (a / and~e.10 :quant 2~e.5 :op1 (c4 / cell-line~e.12 :name (n2 / name :op1 "HCT15"~e.9)) :op2 (c5 / cell-line~e.12 :name (n3 / name :op1 "H460"~e.11)) :ARG0-of (r2 / resist-01~e.8 :ARG1 (s / small-molecule :name (n4 / name :op1 "selumetinib"~e.6))))) :ARG1-of (d / describe-01 :ARG0 (a3 / and~e.26 :op1 (f / figure~e.24 :mod "3C"~e.25) :op2 (f2 / figure~e.24 :mod "3D")))) # ::id a_pmid_2256_9000.127 ::amr-annotator SDL-AMR-09 ::preferred # ::tok We next assessed the levels of AKT , phosphor @-@ AKT , 4EBP1 and poshospho @-@ 4EBP1 , which are downstream effectors of the PI3 kinase ( PI3K ) pathway . # ::alignments 0-1.1 1-1.3 2-1 4-1.2.1 4-1.2.2 4-1.2.3 4-1.2.4 6-1.2.1.1.1.1 6-1.2.2.1.1.1 10-1.2.1.1.1.1 10-1.2.2.1.1.1 12-1.2.3.1.1.1 12-1.2.4.1.1.1 13-1.2 16-1.2.3.1.1.1 16-1.2.4.1.1.1 20-1.4.2 24-1.4.1.1.1 25-1.4.1.1.2 29-1.4.1 (a / assess-01~e.2 :ARG0 (w / we~e.0) :ARG1 (a2 / and~e.13 :op1 (l / level~e.4 :quant-of (e / enzyme :name (n2 / name :op1 "AKT"~e.6,10))) :op2 (l2 / level~e.4 :quant-of (e2 / enzyme :name (n3 / name :op1 "AKT"~e.6,10) :ARG3-of (p / phosphorylate-01))) :op3 (l3 / level~e.4 :quant-of (p2 / protein :name (n4 / name :op1 "4EBP1"~e.12,16))) :op4 (l4 / level~e.4 :quant-of (p3 / protein :name (n5 / name :op1 "4EBP1"~e.12,16) :ARG3-of p))) :time (n / next~e.1) :ARG0-of (a3 / affect-01 :ARG1 (p5 / pathway~e.29 :name (n6 / name :op1 "PI3"~e.24 :op2 "kinase"~e.25)) :mod (d / downstream~e.20))) # ::id a_pmid_2256_9000.128 ::amr-annotator SDL-AMR-09 ::preferred # ::tok Previous reports have suggested that resistance to selumetinib treatment in CRC and NSCLC cell lines was associated with baseline increased PI3K signalling ( Balmanno et al , 2009 ) . # ::alignments 0-1.1.2 1-1.1 1-1.1.1 1-1.1.1.r 3-1 4-1.2.r 5-1.2.1 6-1.2.1.1.r 7-1.2.1.1.1.1.1 8-1.2.1.1 9-1.2.1.2.r 10-1.2.1.2.1.1.1.1 11-1.2.1.2 12-1.2.1.2.2.1.1.1 13-1.2.1.2.1 13-1.2.1.2.2 14-1.2.1.2.1 16-1.2 17-1.2.2.r 18-1.2.2.1.3 19-1.2.2.1.2 20-1.2.2.1.1.1 21-1.2.2 24-1.3.1.1.1.1.1 26-1.3.1.1 27-1.3.1.1.2.1 30-1.3.1.2.1 (s / suggest-01~e.3 :ARG0 (t / thing~e.1 :ARG1-of~e.1 (r / report-01~e.1) :time (p / previous~e.0)) :ARG1~e.4 (a2 / associate-01~e.16 :ARG1 (r2 / resist-01~e.5 :ARG1~e.6 (t2 / treat-04~e.8 :ARG2 (s4 / small-molecule :name (n5 / name :op1 "selumetinib"~e.7))) :location~e.9 (a / and~e.11 :op1 (c / cell-line~e.13,14 :mod (d4 / disease :name (n / name :op1 "CRC"~e.10))) :op2 (c2 / cell-line~e.13 :mod (d3 / disease :name (n2 / name :op1 "NSCLC"~e.12))))) :ARG2~e.17 (s3 / signal-07~e.21 :ARG0 (p2 / pathway :name (n3 / name :op1 "PI3K"~e.20) :ARG1-of (i / increase-01~e.19) :mod (b / baseline~e.18)))) :ARG1-of (d2 / describe-01 :ARG0 (p3 / publication-91 :ARG0 (a3 / and~e.26 :op1 (p4 / person :name (n4 / name :op1 "Balmanno"~e.24)) :op2 (p5 / person :mod (o / other~e.27))) :time (d / date-entity :year 2009~e.30)))) # ::id a_pmid_2256_9000.129 ::amr-annotator SDL-AMR-09 ::preferred # ::tok However , as depicted in Figure 3E @–@ H , we failed to segregate the panel of 11 NSCLC and CRC cell lines into selumetinib @-@ sensitive and selumetinib @-@ resistant groups , by using the PI3K pathway . # ::alignments 0-1 3-1.2 6-1.2.1.1 6-1.2.1.2 7-1.2.1.1.1 12-1.1.1 13-1.1 15-1.1.2 17-1.1.2.2 18-1.1.2.2.1.r 19-1.1.2.2.1.1 20-1.1.2.2.1.2.1.1.1 21-1.1.2.2.1 22-1.1.2.2.1.3.1.1.1 23-1.1.2.2.1.2 23-1.1.2.2.1.3 24-1.1.2.2.1.2 26-1.1.2.3.1.1.1.1.1 28-1.1.2.3.1.1 30-1.1.2.3.1.1.1.1.1 32-1.1.2.3.2.1 33-1.1.2.3.1 33-1.1.2.3.2 35-1.1.2.4.r 36-1.1.2.4 38-1.1.2.4.1.1.1 39-1.1.2.4.1 (h / have-concession-91~e.0 :ARG1 (f / fail-01~e.13 :ARG1 (w / we~e.12) :ARG2 (s / segregate-01~e.15 :ARG0 w :ARG1 (p / panel~e.17 :consist-of~e.18 (a3 / and~e.21 :quant 11~e.19 :op1 (c / cell-line~e.23,24 :mod (d2 / disease :name (n / name :op1 "NSCLC"~e.20))) :op2 (c2 / cell-line~e.23 :mod (d3 / disease :name (n2 / name :op1 "CRC"~e.22))))) :ARG2 (a2 / and :op1 (g / group~e.33 :ARG0-of (s2 / sensitive-03~e.28 :ARG1 (s4 / small-molecule :name (n4 / name :op1 "selumetinib"~e.26,30)))) :op2 (g2 / group~e.33 :ARG0-of (r / resist-01~e.32 :ARG1 s4))) :manner~e.35 (u / use-01~e.36 :ARG1 (p3 / pathway~e.39 :name (n3 / name :op1 "PI3K"~e.38))))) :ARG1-of (d / depict-01~e.3 :ARG0 (v / value-interval :op1 (f2 / figure~e.6 :mod "3E"~e.7) :op2 (f3 / figure~e.6 :mod "3H")))) # ::id a_pmid_2256_9000.130 ::amr-annotator SDL-AMR-09 ::preferred # ::tok Effects of selumetinib treatment on NSCLC and CRC tumour xenografts in nude mice # ::alignments 0-1 1-1.1.r 2-1.1.1.1.1 3-1.1 4-1.2.r 5-1.2.1.1.1.1.1 6-1.2 7-1.2.2.1.1.1.1 8-1.2.1.1 8-1.2.2.1 9-1.2.1 9-1.2.2 10-1.3.r 11-1.3.1 12-1.3 (a / affect-01~e.0 :ARG0~e.1 (t / treat-04~e.3 :ARG2 (s2 / small-molecule :name (n4 / name :op1 "selumetinib"~e.2))) :ARG1~e.4 (a2 / and~e.6 :op1 (x3 / xenograft~e.9 :source (t2 / tumor~e.8 :mod (d / disease :name (n2 / name :op1 "NSCLC"~e.5)))) :op2 (x / xenograft~e.9 :source (t3 / tumor~e.8 :mod (d2 / disease :name (n / name :op1 "CRC"~e.7))))) :location~e.10 (m / mouse~e.12 :mod (n3 / nude~e.11))) # ::id a_pmid_2256_9000.131 ::amr-annotator SDL-AMR-09 ::preferred # ::tok As shown in Figure 4A and B , HCT116 and Calu3 xenograft growth was significantly inhibited by selumetinib treatment in a dose @-@ dependent manner . # ::alignments 1-1.5 4-1.5.1.1 4-1.5.1.2 5-1.5.1.1.1 6-1.5.1 10-1.2.1.1.1.1.1 11-1.2.1 12-1.2.1.2.1.1.1 13-1.2.1.1 13-1.2.1.2 14-1.2 17-1 18-1.1.r 19-1.1.1.1.1 20-1.1 23-1.4.2 25-1.4 26-1.4.r (i / inhibit-01~e.17 :ARG0~e.18 (t / treat-04~e.20 :ARG2 (s4 / small-molecule :name (n3 / name :op1 "selumetinib"~e.19))) :ARG1 (g / grow-01~e.14 :ARG1 (a / and~e.11 :op1 (x3 / xenograft~e.13 :source (c / cell-line :name (n / name :op1 "HCT116"~e.10))) :op2 (x2 / xenograft~e.13 :source (c2 / cell-line :name (n4 / name :op1 "Calu3"~e.12))))) :ARG1-of (s / signify-01) :manner~e.26 (d / depend-01~e.25 :ARG0 g :ARG1 (d2 / dose~e.23)) :ARG1-of (s3 / show-01~e.1 :ARG0 (a2 / and~e.6 :op1 (f / figure~e.4 :mod "4A"~e.5) :op2 (f2 / figure~e.4 :mod "4B")))) # ::id a_pmid_2256_9000.132 ::amr-annotator SDL-AMR-09 ::preferred # ::tok As an example , at day 50 from the injection of cancer cells , the mean tumour volume in mice bearing HCT116 tumour xenografts and treated with selumetinib , 25 or 50 mg kg @ −1 were 40 % and 15 % , respectively , as compared with control untreated mice . # ::alignments 0-1.3.r 2-1.6 5-1.3.2.2 6-1.3.2.1 7-1.1.3.1.1.1.r 9-1.3.1 10-1.3.1.1.r 11-1.3.1.1.1.2.1 12-1.3.1.1 16-1.1.1.1 17-1.1.1 19-1.1.3 20-1.1.3.1 21-1.1.3.1.1.1.1.1.1 22-1.1.3.1.1.1 23-1.1.3.1.1 24-1 25-1.2.3.1 26-1.2.3.1.1.r 27-1.2.3.1.1.1.1 29-1.2.3.1.1.2.1.1 30-1.2.3.1.1.2 31-1.2.3.1.1.2.2.1 32-1.2.3.1.1.2.1.2 32-1.2.3.1.1.2.2.2 33-1.2.3.1.1.2.1.2 33-1.2.3.1.1.2.2.2 38-1.1.2.1 39-1.1.2 40-1 41-1.2.2.1 42-1.2.2 44-1.4 46-1.3.r 47-1.5.r 49-1.5.2 50-1.5.1 50-1.5.1.1 50-1.5.1.1.r 51-1.2.3 51-1.5 (a4 / and~e.24,40 :op1 (a / average-01 :ARG1 (v / volume~e.17 :mod (t / tumor~e.16)) :ARG2 (p / percentage-entity~e.39 :value 40~e.38) :location (m2 / mouse~e.19 :ARG0-of (b / bear-01~e.20 :ARG1 (x2 / xenograft~e.23 :source~e.7 (t3 / tumor~e.22 :mod (c / cell-line :name (n / name :op1 "HCT116"~e.21))))))) :op2 (a5 / average-01 :ARG1 v :ARG2 (p2 / percentage-entity~e.42 :value 15~e.41) :location (m4 / mouse~e.51 :ARG1-of (t4 / treat-04~e.25 :ARG2~e.26 (s2 / small-molecule :name (n4 / name :op1 "selumetinib"~e.27) :quant (o / or~e.30 :op1 (c3 / concentration-quantity :quant 25~e.29 :unit (m / milligram-per-kilogram~e.32,33)) :op2 (c2 / concentration-quantity :quant 50~e.31 :unit (m3 / milligram-per-kilogram~e.32,33))))))) :time~e.0,46 (a2 / after :op1 (i / inject-01~e.9 :ARG2~e.10 (c4 / cell~e.12 :mod (d / disease :wiki "Cancer" :name (n2 / name :op1 "cancer"~e.11)))) :quant (t5 / temporal-quantity :quant 50~e.6 :unit (d2 / day~e.5))) :mod (r / respective~e.44) :compared-to~e.47 (m5 / mouse~e.51 :ARG1-of (t6 / treat-04~e.50 :polarity~e.50 -~e.50) :mod (c6 / control~e.49)) :ARG0-of (e / exemplify-01~e.2)) # ::id a_pmid_2256_9000.133 ::amr-annotator SDL-AMR-09 ::preferred # ::tok Similarly , at day 50 from the injection of cancer cells , the mean tumour volume in mice bearing Calu3 tumour xenografts and treated with selumetinib , 25 or 50 mg kg @ −1 were 22 % and 8 % , respectively , as compared with control untreated mice . # ::alignments 0-1.5 3-1.6.2.2 4-1.6.2.1 5-1.1.3.1.1.1.r 7-1.6.1 9-1.6.1.1.1.2.1 10-1.1.3.1.1.1.1.1.1 10-1.6.1.1 14-1.1.1.1 15-1.1.1 17-1.1.3 18-1.1.3.1 19-1.1.3.1.1.1.1.1.2 20-1.1.3.1.1.1 21-1.1.3.1.1 22-1 23-1.2.3.1 24-1.2.3.1.1.r 25-1.2.3.1.1.1.1 27-1.2.3.1.1.2.1.1 28-1.2.3.1.1.2 29-1.2.3.1.1.2.2.1 30-1.2.3.1.1.2.1.2 30-1.2.3.1.1.2.2.2 31-1.2.3.1.1.2.1.2 31-1.2.3.1.1.2.2.2 36-1.1.2.1 37-1.1.2 38-1 39-1.2.2.1 40-1.2.2 42-1.3 44-1.6.r 45-1.4.r 47-1.4.2 48-1.4.1 48-1.4.1.1 48-1.4.1.1.r 49-1.2.3 49-1.4 (a / and~e.22,38 :op1 (a2 / average-01 :ARG1 (v / volume~e.15 :mod (t / tumor~e.14)) :ARG2 (p / percentage-entity~e.37 :value 22~e.36) :location (m / mouse~e.17 :ARG0-of (b / bear-01~e.18 :ARG1 (x2 / xenograft~e.21 :source~e.5 (t6 / tumor~e.20 :mod (d3 / disease :name (n / name :op1 "cell-line"~e.10 :op2 "Calu3"~e.19))))))) :op2 (a3 / average-01 :ARG1 v :ARG2 (p2 / percentage-entity~e.40 :value 8~e.39) :location (m2 / mouse~e.49 :ARG1-of (t3 / treat-04~e.23 :ARG2~e.24 (s / small-molecule :name (n3 / name :op1 "selumetinib"~e.25) :quant (o / or~e.28 :op1 (c2 / concentration-quantity :quant 25~e.27 :unit (m3 / milligram-per-kilogram~e.30,31)) :op2 (c3 / concentration-quantity :quant 50~e.29 :unit (m4 / milligram-per-kilogram~e.30,31))))))) :mod (r / respective~e.42) :compared-to~e.45 (m5 / mouse~e.49 :ARG1-of (t2 / treat-04~e.48 :polarity~e.48 -~e.48) :mod (c4 / control~e.47)) :ARG1-of (r2 / resemble-01~e.0) :time~e.44 (a4 / after :op1 (i / inject-01~e.7 :ARG2 (c5 / cell~e.10 :mod (d2 / disease :wiki "Cancer" :name (n2 / name :op1 "cancer"~e.9)))) :quant (t5 / temporal-quantity :quant 50~e.4 :unit (d / day~e.3)))) # ::id a_pmid_2256_9000.134 ::amr-annotator SDL-AMR-09 ::preferred # ::tok In contrast , selumetinib treatment was unable to affect tumour growth in both HCT15 and H460 tumour xenografts ( Figure 4C and D ) . # ::alignments 1-1 3-1.1.2.1.1.1 4-1.1.2 6-1.1.1 8-1.1.3 9-1.1.3.2.1 10-1.1.3.2 13-1.1.3.2.2.1.1.1.1.1 14-1.1.3.2.2 15-1.1.3.2.2.2.1.1.1.1 16-1.1.3.2.2.1.1 16-1.1.3.2.2.2.1 17-1.1.3.2.2.1 17-1.1.3.2.2.2 20-1.2.1.1 20-1.2.1.2 21-1.2.1.1.1 22-1.2.1 (c / contrast-01~e.1 :ARG2 (c2 / capable-01 :polarity -~e.6 :ARG1 (t / treat-04~e.4 :ARG2 (s / small-molecule :name (n / name :op1 "selumetinib"~e.3))) :ARG2 (a / affect-01~e.8 :ARG0 t :ARG1 (g / grow-01~e.10 :ARG1 (t2 / tumor~e.9) :location (a2 / and~e.14 :op1 (x3 / xenograft~e.17 :source (t3 / tumor~e.16 :mod (c4 / cell-line :name (n3 / name :op1 "HCT15"~e.13)))) :op2 (x / xenograft~e.17 :source (t4 / tumor~e.16 :mod (c3 / cell-line :name (n2 / name :op1 "H460"~e.15)))))))) :ARG1-of (d / describe-01 :ARG0 (a3 / and~e.22 :op1 (f / figure~e.20 :mod "4C"~e.21) :op2 (f2 / figure~e.20 :mod "4D")))) # ::id a_pmid_2256_9000.135 ::amr-annotator SDL-AMR-09 ::preferred # ::tok EGFR , RAS , MEK , and AKT protein expression and selumetinib sensitivity in NSCLC and CRC cell lines # ::alignments 0-1.1.1.1.1.1 2-1.1.1.2.1.1 4-1.1.1.3.1.1 7-1.1.1.4.1.1 9-1.1 10-1 11-1.2.1.1.1 12-1.2 13-1.2.2.r 14-1.2.2.1.1.1.1 15-1.2.2 16-1.2.2.2.1.1.1 17-1.2.2.1 17-1.2.2.2 18-1.2.2.1 (a / and~e.10 :op1 (e4 / express-03~e.9 :ARG2 (a2 / and :op1 (e8 / enzyme :name (n4 / name :op1 "EGFR"~e.0)) :op2 (e5 / enzyme :name (n5 / name :op1 "RAS"~e.2)) :op3 (e6 / enzyme :name (n6 / name :op1 "MEK"~e.4)) :op4 (e7 / enzyme :name (n7 / name :op1 "AKT"~e.7))) :ARG3 a3) :op2 (s / sensitive-03~e.12 :ARG1 (s2 / small-molecule :name (n8 / name :op1 "selumetinib"~e.11)) :location~e.13 (a3 / and~e.15 :op1 (c / cell-line~e.17,18 :mod (d / disease :name (n / name :op1 "NSCLC"~e.14))) :op2 (c2 / cell-line~e.17 :mod (d2 / disease :name (n2 / name :op1 "CRC"~e.16)))))) # ::id a_pmid_2256_9000.136 ::amr-annotator SDL-AMR-09 ::preferred # ::tok Identification of predictive biomarkers is becoming a fundamental aspect in the development of targeted agents . # ::alignments 0-1.1 2-1.1.1.1 5-1 7-1.2.2 8-1.2 9-1.2.1.r 11-1.2.1 12-1.2.1.1.r 13-1.2.1.1.1 14-1.2.1.1 (b / become-01~e.5 :ARG1 (i / identify-01~e.0 :ARG1 (b2 / biomarker :ARG0-of (p / predict-01~e.2))) :ARG2 (a / aspect~e.8 :topic~e.9 (d / develop-02~e.11 :ARG1~e.12 (a2 / agent~e.14 :ARG1-of (t / target-01~e.13))) :mod (f / fundamental~e.7))) # ::id a_pmid_2256_9000.137 ::amr-annotator SDL-AMR-09 ::preferred # ::tok So far , several preclinical studies have been published trying to address this aspect for the sensitivity to MEK inhibitors , but , since different results have been reported , no univocal interpretation could be made ( Balmanno et al , 2009 ; Dry et al , 2010 ; Garon et al , 2010 ; Tendler et al , 2010 ) @ . # ::alignments 0-1.1.2 1-1.1.2 3-1.1.1.2 4-1.1.1.1 5-1.1.1 8-1.1 8-1.3.1.1 8-1.3.1.2 8-1.3.1.3 8-1.3.1.4 9-1.1.1.3 11-1.1.1.3.1 12-1.1.1.3.1.2.1 13-1.1.1.3.1.2 14-1.1.1.3.1.2.2.r 16-1.1.1.3.1.2.2 17-1.1.1.3.1.2.2.1.r 18-1.1.1.3.1.2.2.1.1.1.1.1 19-1.1.1.3.1.2.2.1 19-1.1.1.3.1.2.2.1.1 19-1.1.1.3.1.2.2.1.1.r 21-1 23-1.2.2 24-1.2.2.1.1.2 25-1.2.2.1.1 25-1.2.2.1.1.1 25-1.2.2.1.1.1.r 28-1.2.2.1 30-1.2.1.1 30-1.2.1.1.r 31-1.2.1.2 32-1.2.1 33-1.2 35-1.2.2.1 38-1.3.1.1.1.1.1.1 40-1.3.1.1.1 40-1.3.1.2.1 40-1.3.1.3.1 40-1.3.1.4.1 41-1.3.1.1.1.2.1 44-1.3.1.1.2.1 48-1.3.1.2.1.1.1.1 50-1.3.1 51-1.3.1.1.1.2.1 54-1.3.1.3.2 58-1.3.1.3.1.1.1.1 60-1.3.1 61-1.3.1.1.1.2.1 64-1.3.1.4.2 68-1.3.1.4.1.1.1.1 70-1.3.1 71-1.3.1.1.1.2.1 74-1.3.1.2.2.1 (c / contrast-01~e.21 :ARG1 (p2 / publish-01~e.8 :ARG1 (s2 / study-01~e.5 :mod (p3 / preclinical~e.4) :quant (s3 / several~e.3) :ARG0-of (t2 / try-01~e.9 :ARG1 (a / address-02~e.11 :ARG0 s2 :ARG1 (a2 / aspect~e.13 :mod (t3 / this~e.12) :topic~e.14 (s4 / sensitive-03~e.16 :ARG1~e.17 (m / molecular-physical-entity~e.19 :ARG0-of~e.19 (i / inhibit-01~e.19 :ARG1 (p / protein-family :name (n / name :op1 "MEK"~e.18))))))))) :time (s / so-far~e.0,1)) :ARG2 (p4 / possible-01~e.33 :ARG1 (i2 / interpret-01~e.32 :polarity~e.30 -~e.30 :mod (u / univocal~e.31)) :ARG1-of (c2 / cause-01~e.23 :ARG0 (r / report-01~e.28,35 :ARG1 (t4 / thing~e.25 :ARG2-of~e.25 (r2 / result-01~e.25) :ARG1-of (d4 / differ-02~e.24))))) :ARG1-of (d3 / describe-01 :ARG0 (a3 / and~e.50,60,70 :op1 (p5 / publication-91~e.8 :ARG0 (a4 / and~e.40 :op1 (p6 / person :name (n2 / name :op1 "Balmanno"~e.38)) :op2 (p7 / person :mod (o / other~e.41,51,61,71))) :time (d / date-entity :year 2009~e.44)) :op2 (p8 / publication-91~e.8 :ARG0 (a5 / and~e.40 :op1 (p9 / person :name (n3 / name :op1 "Dry"~e.48)) :op2 p7) :time (d2 / date-entity :year 2010~e.74)) :op3 (p11 / publication-91~e.8 :ARG0 (a6 / and~e.40 :op1 (p12 / person :name (n4 / name :op1 "Garon"~e.58)) :op2 p7) :time d2~e.54) :op4 (p14 / publication-91~e.8 :ARG0 (a7 / and~e.40 :op1 (p15 / person :name (n5 / name :op1 "Tendler"~e.68)) :op2 p7) :time d2~e.64)))) # ::id a_pmid_2256_9000.138 ::amr-annotator SDL-AMR-09 ::preferred # ::tok To identify specific profiles , which could allow identifying different molecular patterns of sensitivity or resistance to MEK inhibition , we first screened the intracellular signalling status of each cell lines . # ::alignments 1-1.3 2-1.3.2.1 3-1.3.2 6-1.3.2.2.2 7-1.3.2.2 8-1.3.2.2.1 9-1.3.2.2.1.1.3 10-1.3.2.2.1.1.1.1.1.2 10-1.3.2.2.1.1.2 11-1.3.2.2.1.1 11-1.3.2.2.1.1.1.1.1 12-1.3.2.2.1.1.1.r 13-1.3.2.2.1.1.1.1 14-1.3.2.2.1.1.1 15-1.3.2.2.1.1.1.2 16-1.3.2.2.1.1.1.1.1.1.r 17-1.3.2.2.1.1.1.1.1.1.1.1.1 18-1.3.2.2.1.1.1.1.1.1 20-1.1 21-1.4 22-1 24-1.2.1.1 24-1.2.1.2 25-1.2.1 26-1.2 28-1.2.1.1.1 29-1.2.1.1 30-1.2.1.1 (s / screen-01~e.22 :ARG0 (w / we~e.20) :ARG1 (s2 / status~e.26 :mod (s3 / signal-07~e.25 :ARG1 (c / cell-line~e.24,29,30 :mod (e2 / each~e.28)) :mod (i4 / intracellular~e.24))) :purpose (i / identify-01~e.1 :ARG0 w :ARG1 (p2 / profile~e.3 :ARG1-of (s4 / specific-02~e.2) :ARG0-of (a / allow-01~e.7 :ARG1 (i2 / identify-01~e.8 :ARG1 (p3 / pattern-01~e.11 :ARG1~e.12 (o3 / or~e.14 :op1 (s5 / sensitive-03~e.13 :ARG1 (p / pattern~e.11 :ARG0-of~e.16 (i3 / inhibit-01~e.18 :ARG1 (e / enzyme :name (n / name :op1 "MEK"~e.17))) :mod (m2 / molecule~e.10))) :op2 (r2 / resist-01~e.15 :ARG1 p)) :mod (m / molecule~e.10) :ARG1-of (d / differ-02~e.9))) :ARG1-of (p4 / possible-01~e.6)))) :time (f / first~e.21)) # ::id a_pmid_2256_9000.139 ::amr-annotator SDL-AMR-09 ::preferred # ::tok As depicted in Supplementary Figure 1A , the NCSLC and CRC cell lines displayed highly variable basal levels of total and phosphorylated EGFR , RAS , MEK , and AKT . # ::alignments 1-1.3 4-1.3.1.2 5-1.3.1 6-1.3.1.1 10-1.1.1.1.1.1 11-1.1 12-1.1.2.1.1.1 13-1.1.1 13-1.1.2 14-1.1.1 15-1 16-1.2.9.1 18-1.2.10 19-1.2.1 19-1.2.2 19-1.2.3 19-1.2.4 19-1.2.5 19-1.2.6 19-1.2.7 19-1.2.8 20-1.2.1.1.r 21-1.2.1.1.2 22-1.2 23-1.2.2.1.2 24-1.2.1.1.1.1 24-1.2.2.1.1.1 26-1.2.3.1.1.1 26-1.2.4.1.1.1 28-1.2.5.1.1.1 28-1.2.6.1.1.1 30-1.2 31-1.2.7.1.1.1 31-1.2.8.1.1.1 (d / display-01~e.15 :ARG0 (a / and~e.11 :op1 (c / cell-line~e.13,14 :mod (d4 / disease :name (n4 / name :op1 "NCSLC"~e.10))) :op2 (c2 / cell-line~e.13 :mod (d3 / disease :name (n5 / name :op1 "CRC"~e.12)))) :ARG1 (a2 / and~e.22,30 :op1 (l5 / level~e.19 :quant-of~e.20 (e2 / enzyme :name (n2 / name :op1 "EGFR"~e.24) :mod (t2 / total~e.21))) :op2 (l / level~e.19 :quant-of (e / enzyme :name (n / name :op1 "EGFR"~e.24) :ARG3-of (p / phosphorylate-01~e.23))) :op3 (l6 / level~e.19 :quant-of (e3 / enzyme :name (n3 / name :op1 "RAS"~e.26) :mod t2)) :op4 (l2 / level~e.19 :quant-of (e4 / enzyme :name (n6 / name :op1 "RAS"~e.26) :ARG3-of p)) :op5 (l3 / level~e.19 :quant-of (e5 / enzyme :name (n7 / name :op1 "MEK"~e.28) :mod t2)) :op6 (l7 / level~e.19 :quant-of (e7 / enzyme :name (n9 / name :op1 "MEK"~e.28) :ARG2-of p)) :op7 (l4 / level~e.19 :quant-of (e6 / enzyme :name (n8 / name :op1 "AKT"~e.31) :mod t2)) :op8 (l8 / level~e.19 :quant-of (e8 / enzyme :name (n10 / name :op1 "AKT"~e.31) :ARG3-of p)) :ARG1-of (v / vary-01 :ARG2 (h / high-02~e.16)) :mod (b / basal~e.18)) :ARG1-of (d2 / depict-01~e.1 :ARG0 (f / figure~e.5 :mod "1A"~e.6 :ARG2-of (s / supplement-01~e.4)))) # ::id a_pmid_2256_9000.140 ::amr-annotator SDL-AMR-09 ::preferred # ::tok As illustrated in Supplementary Figure 1B , there was no apparent correlation between basal levels of total and phosphorylated proteins listed above in both tumour types . # ::alignments 1-1.5 4-1.5.1.2 5-1.5.1 6-1.5.1.1 11-1.1 11-1.1.r 12-1.4 13-1 15-1.2.2 16-1.2 16-1.3 17-1.2.1.r 18-1.2.1.1 20-1.3.1 21-1.2.1 22-1.2.3 23-1.2.3.1 24-1.2.4.r 25-1.2.4.2 26-1.2.4.1 27-1.2.4 (c2 / correlate-01~e.13 :polarity~e.11 -~e.11 :ARG1 (l / level~e.16 :quant-of~e.17 (p / protein~e.21 :mod (t / total~e.18)) :mod (b / basal~e.15) :ARG1-of (l3 / list-01~e.22 :location (a2 / above~e.23)) :location~e.24 (t2 / type-03~e.27 :ARG1 (t3 / tumor~e.26) :mod (b2 / both~e.25))) :ARG2 (l2 / level~e.16 :ARG3-of (p3 / phosphorylate-01~e.20) :ARG1-of l3 :mod b :location t2) :mod (a / apparent~e.12) :ARG1-of (i / illustrate-01~e.1 :ARG0 (f / figure~e.5 :mod "1B"~e.6 :ARG2-of (s / supplement-01~e.4)))) # ::id a_pmid_2256_9000.141 ::amr-annotator SDL-AMR-09 ::preferred # ::tok Gene mutations and selumetinib sensitivity in NSCLC and CRC cell lines # ::alignments 0-1.1.1 1-1.1 2-1 2-1.3 3-1.2.1.1.1 4-1.2 5-1.3.r 6-1.3.1.1.1.1 7-1.3 8-1.3.2.1.1.1 9-1.3.1 9-1.3.2 10-1.3.1 (a / and~e.2 :op1 (m / mutate-01~e.1 :ARG2 (g / gene~e.0)) :op2 (s / sensitive-03~e.4 :ARG1 (s2 / small-molecule :name (n / name :op1 "selumetinib"~e.3))) :location~e.5 (a2 / and~e.2,7 :op1 (c / cell-line~e.9,10 :mod (d / disease :name (n2 / name :op1 "NSCLC"~e.6))) :op2 (c2 / cell-line~e.9 :mod (d2 / disease :name (n3 / name :op1 "CRC"~e.8))))) # ::id a_pmid_2256_9000.142 ::amr-annotator SDL-AMR-09 ::preferred # ::tok Activation of the classic MAPK cascade ( RAS @-@ RAF @-@ MEK @-@ ERK ) is a common event in colorectal and lung cancer . # ::alignments 0-1.3 1-1.3.1.r 3-1.3.1.2 4-1.3.1.1.1 7-1.3.1.3.1.1.1.1 9-1.3.1.3.1.2.1.1 11-1.3.1.3.1.3.1.1 13-1.3.1.3.1.4.1.1 15-1.3.r 17-1.1 18-1 19-1.2.r 20-1.2.1.2.1 21-1.2 23-1.2.2.2.1 (e / event~e.18 :mod (c / common~e.17) :location~e.19 (a / and~e.21 :op1 (d / disease :wiki "Colorectal_cancer" :name (n2 / name :op1 "colon"~e.20 :op2 "cancer")) :op2 (d2 / disease :wiki "Lung_cancer" :name (n7 / name :op1 "cancer"~e.23))) :domain~e.15 (a2 / activate-01~e.0 :ARG1~e.1 (p / pathway :name (n / name :op1 "MAPK"~e.4) :mod (c6 / classic~e.3) :ARG1-of (m / mean-01 :ARG2 (m2 / macro-molecular-complex :part (e5 / enzyme :name (n3 / name :op1 "RAS"~e.7)) :part (e2 / enzyme :name (n4 / name :op1 "RAF"~e.9)) :part (e4 / enzyme :name (n5 / name :op1 "MEK"~e.11)) :part (e3 / enzyme :name (n6 / name :op1 "ERK"~e.13))))))) # ::id a_pmid_2256_9000.143 ::amr-annotator SDL-AMR-09 ::preferred # ::tok Some genes within this pathway are mutated or aberrantly expressed . # ::alignments 0-1.1.1.1 1-1.1.1 2-1.1.1.1.r 3-1.1.1.2.1.1 4-1.1.1.2.1 6-1.1 7-1 9-1.2 (o / or~e.7 :op1 (m / mutate-01~e.6 :ARG1 (g / gene~e.1 :quant~e.2 (s / some~e.0) :ARG1-of (i / include-91 :ARG2 (p / pathway~e.4 :mod (t / this~e.3))))) :op2 (e / express-03~e.9 :ARG2 g :manner (a / aberrant))) # ::id a_pmid_2256_9000.144 ::amr-annotator SDL-AMR-09 ::preferred # ::tok Moreover , the MAPK pathway can be indirectly activated by mutations of genes encoding for PI3K/PTEN/AKT and p53 . # ::alignments 0-1 3-1.1.1.2.1.1 4-1.1.1.1.1.1.1.1 4-1.1.1.2 5-1.1 7-1.1.1.3 7-1.1.1.3.1 7-1.1.1.3.1.r 8-1.1.1 9-1.1.1.1.r 10-1.1.1.1 11-1.1.1.1.1.r 12-1.1.1.1.1 13-1.1.1.1.1.1 14-1.1.1.1.1.1.1.r 15-1.1.1.1.1.1.1.1.1.1 16-1.1.1.1.1.1.1 17-1.1.1.1.1.1.1.2.1.1 (a / and~e.0 :op2 (p2 / possible-01~e.5 :ARG1 (a2 / activate-01~e.8 :ARG0~e.9 (m / mutate-01~e.10 :ARG1~e.11 (g / gene~e.12 :ARG0-of (e / encode-01~e.13 :ARG1~e.14 (a3 / and~e.16 :op1 (p3 / pathway~e.4 :name (n2 / name :op1 "PI3K/PTEN/AKT"~e.15)) :op2 (p4 / protein :name (n3 / name :op1 "p53"~e.17)))))) :ARG1 (p / pathway~e.4 :name (n / name :op1 "MAPK"~e.3)) :ARG1-of (d / direct-02~e.7 :polarity~e.7 -~e.7)))) # ::id a_pmid_2256_9000.145 ::amr-annotator SDL-AMR-09 ::preferred # ::tok We have screened the panel of 11 NSCLC and CRC cell lines for mutations in KRAS , NRAS , BRAF , PIK3CA , p53 , PTEN , MEK1 @/@ 2 , AKT , EGFR ( Table 1 ; Supplementary Table 1A @–@ F ) . # ::alignments 0-1.1 2-1 4-1.2 5-1.2.1.r 6-1.2.1.1 7-1.2.1.2.1.1.1 8-1.2.1 9-1.2.1.3.1.1.1 10-1.2.1.2 10-1.2.1.3 11-1.2.1.2 12-1.3.r 13-1.3 16-1.3.1.1.1.1 18-1.3.1.2.1.1 20-1.3.1.3.1.1 22-1.3.1.4.1.1 24-1.3.1.5.1.1 26-1.3.1.6.1.1 28-1.3.1.7.1.1 30-1.3.1.7.1.1 32-1.3.1.8.1.1 34-1.3.1.9.1.1 38-1.4.1.1 38-1.4.1.2.2 39-1.4.1.1.1 43-1.4.1.2.3 44-1.4.1.2.1 44-1.4.1.2.2 45-1.4.1.2.1.1 (s / screen-01~e.2 :ARG0 (w2 / we~e.0) :ARG1 (p / panel~e.4 :consist-of~e.5 (a4 / and~e.8 :quant 11~e.6 :op1 (c / cell-line~e.10,11 :mod (d2 / disease :name (n / name :op1 "NSCLC"~e.7))) :op2 (c2 / cell-line~e.10 :mod (d3 / disease :name (n2 / name :op1 "CRC"~e.9))))) :ARG2~e.12 (m / mutate-01~e.13 :ARG1 (a2 / and :op1 (g2 / gene :name (n5 / name :op1 "KRAS"~e.16)) :op2 (g3 / gene :name (n6 / name :op1 "NRAS"~e.18)) :op3 (g4 / gene :name (n7 / name :op1 "BRAF"~e.20)) :op4 (g5 / gene :name (n8 / name :op1 "PIK3CA"~e.22)) :op5 (g6 / gene :name (n9 / name :op1 "p53"~e.24)) :op6 (g7 / gene :name (n10 / name :op1 "PTEN"~e.26)) :op7 (g8 / gene :name (n11 / name :op1 "MEK1/2"~e.28,30)) :op8 (g9 / gene :name (n12 / name :op1 "AKT"~e.32)) :op9 (g10 / gene :name (n13 / name :op1 "EGFR"~e.34)))) :ARG1-of (d / describe-01 :ARG0 (a3 / and :op1 (t / table~e.38 :mod 1~e.39) :op2 (v / value-interval :op1 (t2 / table~e.44 :mod "1A"~e.45) :op2 (t3 / table~e.38,44 :mod "1F") :ARG2-of (s2 / supplement-01~e.43))))) # ::id a_pmid_2256_9000.146 ::amr-annotator SDL-AMR-09 ::preferred # ::tok In NSCLC cell lines , two out of five ( 40 %) harboured a KRAS mutation , which was located in codon 12 or 13 . # ::alignments 1-1.3.1.1.1 2-1.1 3-1.1 3-1.1.2.1 3-1.3 5-1.1.1 8-1.1.2.1.1 10-1.1.2.2.1 12-1 15-1.2.1.1.1 17-1.2 21-1.2.2 22-1.2.2.1.r 23-1.2.2.1.1 23-1.2.2.1.2 24-1.2.2.1.1.1 25-1.2.2.1 26-1.2.2.1.2.1 (h / harbor-01~e.12 :ARG0 (c4 / cell-line~e.2,3 :quant 2~e.5 :ARG1-of (i / include-91 :ARG2 (c5 / cell-line~e.3 :quant 5~e.8) :ARG3 (p / percentage-entity :value 40~e.10))) :ARG1 (m / mutate-01~e.17 :ARG1 (g / gene :name (n / name :op1 "KRAS"~e.15)) :ARG1-of (l / locate-01~e.21 :location~e.22 (o / or~e.25 :op1 (c / codon~e.23 :mod 12~e.24) :op2 (c2 / codon~e.23 :mod 13~e.26)))) :location (c3 / cell-line~e.3 :mod (d / disease :name (n2 / name :op1 "NSCLC"~e.1)))) # ::id a_pmid_2256_9000.147 ::amr-annotator SDL-AMR-09 ::preferred # ::tok Moreover , three out of five ( 60 %) of NSCLC cells harboured a PI3KCA mutation , which were located in exon 9 or 20 . # ::alignments 0-1 2-1.1.1.1 5-1.1.1.2.1.1 7-1.1.1.2.2.1 10-1.1.1.2.1.2.1.1 11-1.1.1 11-1.1.1.2.1 12-1.1 15-1.1.2.1.1.1 17-1.1.2 21-1.1.2.2 22-1.1.2.2.1.r 23-1.1.2.2.1.1 23-1.1.2.2.1.2 24-1.1.2.2.1.1.1 25-1.1.2.2.1 26-1.1.2.2.1.2.1 (a / and~e.0 :op2 (h / harbor-01~e.12 :ARG0 (c / cell-line~e.11 :quant 3~e.2 :ARG1-of (i / include-91 :ARG2 (c2 / cell-line~e.11 :quant 5~e.5 :mod (d / disease :name (n / name :op1 "NSCLC"~e.10))) :ARG3 (p / percentage-entity :value 60~e.7))) :ARG1 (m / mutate-01~e.17 :ARG1 (g / gene :name (n3 / name :op1 "PI3KCA"~e.15)) :ARG1-of (l / locate-01~e.21 :location~e.22 (o / or~e.25 :op1 (e / exon~e.23 :mod 9~e.24) :op2 (e2 / exon~e.23 :mod 20~e.26)))))) # ::id a_pmid_2256_9000.148 ::amr-annotator SDL-AMR-09 ::preferred # ::tok A concomitant mutation in KRAS and PI3KCA gene was found in two out of five ( 40 %) NSCLC . # ::alignments 1-1.1.2 2-1.1 5-1.1.1.1.1.1 7-1.1.1 9-1.1.1.2.1.1 11-1.1.1.1 11-1.1.1.2 13-1 14-1.2.r 15-1.2.1 18-1.2.2.1.1 20-1.2.2.2.1 22-1.2.2.1.2.1.1 (f / find-01~e.13 :ARG1 (m / mutate-01~e.2 :ARG1 (a / and~e.7 :op1 (g / gene~e.11 :name (n / name :op1 "KRAS"~e.5)) :op2 (g2 / gene~e.11 :name (n2 / name :op1 "PI3KCA"~e.9))) :mod (c / concomitant~e.1)) :location~e.14 (c2 / cell-line :quant 2~e.15 :ARG1-of (i / include-91 :ARG2 (c3 / cell-line :quant 5~e.18 :mod (d / disease :name (n3 / name :op1 "NSCLC"~e.22))) :ARG3 (p / percentage-entity :value 40~e.20)))) # ::id a_pmid_2256_9000.149 ::amr-annotator SDL-AMR-09 ::preferred # ::tok One NSCLC cell line had an NRAS mutation ( Table 1 ; Supplementary Table 1A @–@ F ) . # ::alignments 0-1.1.1 1-1.1.2.1.1 2-1.1 3-1.1 4-1 7-1.2.1.1.1 9-1.2 12-1.3.1.1 12-1.3.1.2.2 13-1.3.1.1.1 17-1.3.1.2.3 18-1.3.1.2.1 18-1.3.1.2.2 19-1.3.1.2.1.1 (h / have-03~e.4 :ARG0 (c / cell-line~e.2,3 :quant 1~e.0 :mod (d2 / disease :name (n / name :op1 "NSCLC"~e.1))) :ARG1 (m / mutate-01~e.9 :ARG1 (g / gene :name (n2 / name :op1 "NRAS"~e.7))) :ARG1-of (d / describe-01 :ARG0 (a / and :op1 (t / table~e.12 :mod 1~e.13) :op2 (v / value-interval :op1 (t2 / table~e.18 :mod "1A"~e.19) :op2 (t3 / table~e.12,18 :mod "1F") :ARG2-of (s / supplement-01~e.17))))) # ::id a_pmid_2256_9000.150 ::amr-annotator SDL-AMR-09 ::preferred # ::tok In the panel of six CRC cell lines , all of them harboured a KRAS gene mutation that was located in codon 12 or 13 . # ::alignments 2-1.3 3-1.3.1.r 4-1.3.1.1 5-1.3.1.2.1.1 6-1.3.1 7-1.3.1 11-1.1 12-1 15-1.2.1.1.1 17-1.2.1 18-1.2 21-1.2.2 22-1.2.2.1.r 23-1.2.2.1.1 23-1.2.2.1.2 24-1.2.2.1.1.1 25-1.2.2.1 26-1.2.2.1.2.1 (h / harbor-01~e.12 :ARG0 c3~e.11 :ARG1 (m / mutate-01~e.18 :ARG1 (g / gene~e.17 :name (n / name :op1 "KRAS"~e.15)) :ARG1-of (l / locate-01~e.21 :location~e.22 (o / or~e.25 :op1 (c / codon~e.23 :mod 12~e.24) :op2 (c2 / codon~e.23 :mod 13~e.26)))) :location (p / panel~e.2 :consist-of~e.3 (c3 / cell-line~e.6,7 :quant 6~e.4 :mod (d / disease :name (n2 / name :op1 "CRC"~e.5))))) # ::id a_pmid_2256_9000.151 ::amr-annotator SDL-AMR-09 ::preferred # ::tok Moreover , half of CRC cell lines had both a PI3KCA mutation in exon 9 or 20 and a KRAS mutation . # ::alignments 0-1 2-1.1.1.1 4-1.1.1.2.1.1.1.1 5-1.1.1 5-1.1.1.2.1 6-1.1.1 6-1.1.1.2.1 7-1.1 11-1.1.2.1.1.1.1 13-1.1.2.1 15-1.1.2.1.2.1 15-1.1.2.1.2.2 16-1.1.2.1.2.1.1 18-1.1.2.1.2.2.1 19-1.1.2 19-1.1.2.1.2 22-1.1.2.2.1.1.1 24-1.1.2.1 24-1.1.2.2 (a / and~e.0 :op2 (h / have-03~e.7 :ARG0 (c / cell-line~e.5,6 :quant "1/2"~e.2 :ARG1-of (i / include-91 :ARG2 (c2 / cell-line~e.5,6 :mod (d / disease :name (n / name :op1 "CRC"~e.4))))) :ARG1 (a2 / and~e.19 :op1 (m / mutate-01~e.13,24 :ARG1 (g / gene :name (n3 / name :op1 "PI3KCA"~e.11)) :location (a3 / and~e.19 :op1 (e / exon~e.15 :mod 9~e.16) :op2 (e2 / exon~e.15 :mod 20~e.18))) :op2 (m2 / mutate-01~e.24 :ARG1 (g2 / gene :name (n4 / name :op1 "KRAS"~e.22)))))) # ::id a_pmid_2256_9000.152 ::amr-annotator SDL-AMR-09 ::preferred # ::tok None of the CRC cells had an NRAS mutation . # ::alignments 0-1.1.1 1-1.1.1.r 3-1.1.2.1.1 4-1.1 5-1 8-1.2.1.1.1 10-1.2 (h / have-03~e.5 :ARG0 (c / cell-line~e.4 :quant~e.1 (n2 / none~e.0) :mod (d / disease :name (n / name :op1 "CRC"~e.3))) :ARG1 (m / mutate-01~e.10 :ARG1 (g / gene :name (n3 / name :op1 "NRAS"~e.8)))) # ::id a_pmid_2256_9000.153 ::amr-annotator SDL-AMR-09 ::preferred # ::tok No BRAF mutations were observed in the whole panel of NSCLC and CRC cells . # ::alignments 0-1.1.1 0-1.1.1.r 2-1.1.2.1.1 4-1.1 6-1 7-1.2.r 9-1.2.1 10-1.2 11-1.2.2.r 12-1.2.2.1.1.1.1 13-1.2.2 14-1.2.2.2.1.1.1 15-1.2.2.1 15-1.2.2.2 (o / observe-01~e.6 :ARG1 (m / mutate-01~e.4 :polarity~e.0 -~e.0 :ARG1 (g / gene :name (n / name :op1 "BRAF"~e.2))) :location~e.7 (p / panel~e.10 :mod (w / whole~e.9) :consist-of~e.11 (a2 / and~e.13 :op1 (c / cell-line~e.15 :mod (d / disease :name (n2 / name :op1 "NSCLC"~e.12))) :op2 (c2 / cell-line~e.15 :mod (d2 / disease :name (n3 / name :op1 "CRC"~e.14)))))) # ::id a_pmid_2256_9000.154 ::amr-annotator SDL-AMR-09 ::preferred # ::tok As reported in Supplementary Table 1E and F , no other gene mutations were found in both NSCLC and CRC cell lines . # ::alignments 1-1.3 4-1.3.1.3 5-1.3.1.1 5-1.3.1.2 6-1.3.1.1.1 7-1.3.1 11-1.1.1 11-1.1.1.r 12-1.1.2.1 13-1.1.2 14-1.1 16-1 19-1.2.1.1.1.1 20-1.2 21-1.2.2.1.1.1 22-1.2.1 22-1.2.2 23-1.2.1 (f / find-01~e.16 :ARG1 (m / mutate-01~e.14 :polarity~e.11 -~e.11 :ARG1 (g / gene~e.13 :mod (o / other~e.12))) :location (a / and~e.20 :op1 (c / cell-line~e.22,23 :mod (d2 / disease :name (n / name :op1 "NSCLC"~e.19))) :op2 (c2 / cell-line~e.22 :mod (d / disease :name (n2 / name :op1 "CRC"~e.21)))) :ARG1-of (r / report-01~e.1 :ARG0 (a2 / and~e.7 :op1 (t / table~e.5 :mod "1E"~e.6) :op2 (t2 / table~e.5 :mod "1F") :ARG2-of (s / supplement-01~e.4)))) # ::id a_pmid_2256_9000.155 ::amr-annotator SDL-AMR-09 ::preferred # ::tok After this screening , we tried to correlate the mutational status with selumetinib sensitivity . # ::alignments 0-1.3 1-1.3.1.1 2-1.3.1 4-1.1 5-1 7-1.2 9-1.2.2.1 10-1.2.2 11-1.2.3.r 12-1.2.3.1.1.1 13-1.2.3 (t / try-01~e.5 :ARG0 (w / we~e.4) :ARG1 (c / correlate-01~e.7 :ARG0 w :ARG1 (s / status~e.10 :mod (m / mutate-01~e.9)) :ARG2~e.11 (s2 / sensitive-03~e.13 :ARG1 (s3 / small-molecule :name (n / name :op1 "selumetinib"~e.12)))) :time (a / after~e.0 :op1 (s4 / screen-01~e.2 :mod (t2 / this~e.1)))) # ::id a_pmid_2256_9000.156 ::amr-annotator SDL-AMR-09 ::preferred # ::tok The analysis was made either considering separately the two sets of cell lines ( data not shown ) or all together ( Supplementary Figure 2A and B ) . # ::alignments 1-1 5-1.1 6-1.1.1.1.3 8-1.1.1.1.1 8-1.1.1.2.1 9-1.1.1.1 9-1.1.1.2 10-1.1.1.1.2.r 11-1.1.1.1.2 12-1.1.1.1.2 14-1.1.1.1.4.1 15-1.1.1.1.4.1.1.1 15-1.1.1.1.4.1.1.1.r 16-1.1.1.1.4.1.1 18-1.1.1 20-1.1.1.2.3 23-1.1.1.2.4.1.3 24-1.1.1.2.4.1.1 24-1.1.1.2.4.1.2 25-1.1.1.2.4.1.1.1 26-1.1.1.2.4.1 (a4 / analyze-01~e.1 :manner (c / consider-01~e.5 :ARG1 (o / or~e.18 :op1 (s / set~e.9 :quant 2~e.8 :consist-of~e.10 (c2 / cell-line~e.11,12) :ARG1-of (s2 / separate-02~e.6) :ARG1-of (d / describe-01 :ARG0 (d2 / data~e.14 :ARG1-of (s3 / show-01~e.16 :polarity~e.15 -~e.15)))) :op2 (s5 / set~e.9 :quant 2~e.8 :consist-of c2 :mod (t2 / together~e.20) :ARG1-of (d3 / describe-01 :ARG0 (a3 / and~e.26 :op1 (f / figure~e.24 :mod "2A"~e.25) :op2 (f2 / figure~e.24 :mod "2B") :ARG2-of (s4 / supplement-01~e.23))))))) # ::id a_pmid_2256_9000.157 ::amr-annotator SDL-AMR-09 ::preferred # ::tok Sensitivity to selumetinib did not seem to correlate with any specific gene mutations in this panel of NSCLC and CRC cell lines . # ::alignments 0-1.2.1 1-1.2.1.1.r 2-1.2.1.1.1.1 4-1.1 4-1.1.r 5-1 7-1.2 8-1.2.2.r 9-1.2.2.3 10-1.2.2.2 11-1.2.2.1 12-1.2.2 13-1.2.3.r 14-1.2.3.2 15-1.2.3 16-1.2.3.1.r 17-1.2.3.1.1.1.1.1 18-1.2.3.1 19-1.2.3.1.2.1.1.1 20-1.2.3.1.1 20-1.2.3.1.2 21-1.2.3.1.1 (s / seem-01~e.5 :polarity~e.4 -~e.4 :ARG1 (c / correlate-01~e.7 :ARG1 (s2 / sensitive-03~e.0 :ARG1~e.1 (s3 / small-molecule :name (n / name :op1 "selumetinib"~e.2))) :ARG2~e.8 (m / mutate-01~e.12 :ARG1 (g / gene~e.11) :ARG1-of (s4 / specific-02~e.10) :mod (a / any~e.9)) :location~e.13 (p / panel~e.15 :consist-of~e.16 (a3 / and~e.18 :op1 (c2 / cell-line~e.20,21 :mod (d / disease :name (n2 / name :op1 "NSCLC"~e.17))) :op2 (c3 / cell-line~e.20 :mod (d2 / disease :name (n3 / name :op1 "CRC"~e.19)))) :mod (t / this~e.14)))) # ::id a_pmid_2256_9000.158 ::amr-annotator SDL-AMR-09 ::preferred # ::tok Identification of gene expression profiles that could be predictive of response to selumetinib in NSCLC and CRC cell lines # ::alignments 0-1 1-1.1.r 2-1.1.1.1 3-1.1.1 4-1.1 6-1.2.2 8-1.2 9-1.2.1.r 10-1.2.1 11-1.2.1.1.r 12-1.2.1.1.1.1 13-1.3.r 14-1.3.1.1.1.1 15-1.3 16-1.3.2.1.1.1 17-1.3.1 17-1.3.2 18-1.3.1 (i2 / identify-01~e.0 :ARG1~e.1 (p / profile~e.4 :topic (e / express-03~e.3 :ARG2 (g / gene~e.2))) :ARG0-of (p5 / predict-01~e.8 :ARG1~e.9 (r / respond-01~e.10 :ARG1~e.11 (s / small-molecule :name (n / name :op1 "selumetinib"~e.12))) :ARG1-of (p2 / possible-01~e.6)) :location~e.13 (a / and~e.15 :op1 (c / cell-line~e.17,18 :mod (d / disease :name (n2 / name :op1 "NSCLC"~e.14))) :op2 (c2 / cell-line~e.17 :mod (d2 / disease :name (n3 / name :op1 "CRC"~e.16))))) # ::id a_pmid_2256_9000.159 ::amr-annotator SDL-AMR-09 ::preferred # ::tok RNAs from the 11 cancer cell lines were extracted and used for microarray gene expression analysis . # ::alignments 1-1.1.1.2.r 3-1.1.1.2.1 4-1.1.1.2.2.2.1 5-1.1.1.2 6-1.1.1.2 8-1.1 9-1 10-1.2 11-1.2.2.r 12-1.2.2.1 13-1.2.2.2.1 14-1.2.2.2 15-1.2.2 (a / and~e.9 :op1 (e / extract-01~e.8 :ARG1 (n3 / nucleic-acid :name (n2 / name :op1 "RNA") :source~e.1 (c / cell-line~e.5,6 :quant 11~e.3 :mod (d / disease :wiki "Cancer" :name (n / name :op1 "cancer"~e.4))))) :op2 (u / use-01~e.10 :ARG1 n3 :ARG2~e.11 (a2 / analyze-01~e.15 :ARG0 (m / microarray~e.12) :ARG1 (e2 / express-03~e.14 :ARG2 (g / gene~e.13))))) # ::id a_pmid_2256_9000.160 ::amr-annotator SDL-AMR-09 ::preferred # ::tok Using Student 's t @ -@ test with Benjamini @–@ Hochberg multiple test correction , 21 and 18 genes were identified as upregulated or downregulated , respectively , in selumetinib @-@ resistant cancer cell lines ( t @ -@ test , P @ < 0.05 ) ( Supplementary Figure 3 ) . # ::alignments 1-1.4.2.2.1 2-1.4.2.2.1 4-1.4.2.2.2 7-1.4.2.2.2 9-1.4.2.3.2.1.1.1.1 11-1.4.2.3.2.1.2.1.1 12-1.4.2.3.1.1 13-1.4.2.3.1 14-1.4.2.3 16-1.1.1.1 17-1 18-1.2.1.1 19-1.1.1 19-1.2.1 21-1.1 21-1.2 22-1.1.2.r 23-1.1.2 25-1.2.2 30-1.3.2.1.1.1 32-1.3.2 33-1.3.1.2.1 34-1.3 35-1.3 38-1.4.2.2.2 41-1.4 41-1.4.2.2.2 44-1.4 46-1.4.1 47-1.4.1.1 51-1.5.1.2 52-1.5.1 53-1.5.1.1 (a2 / and~e.17 :op1 (i / identify-01~e.21 :ARG1 (g / gene~e.19 :quant 21~e.16) :ARG2~e.22 (u / upregulate-01~e.23)) :op2 (i2 / identify-01~e.21 :ARG1 (g2 / gene~e.19 :quant 18~e.18) :ARG2 (d2 / downregulate-01~e.25)) :location (c / cell-line~e.34,35 :mod (d / disease :wiki "Cancer" :name (n / name :op1 "cancer"~e.33)) :ARG0-of (r2 / resist-01~e.32 :ARG1 (s2 / small-molecule :name (n2 / name :op1 "selumetinib"~e.30)))) :ARG1-of (s / statistical-test-91~e.41,44 :ARG2 (l / less-than~e.46 :op1 0.05~e.47) :ARG4 (t / thing :wiki "Student's_t-test" :name (n3 / name :op1 "Student's"~e.1,2 :op2 "t-test"~e.4,7,38,41) :mod (c2 / correct-01~e.14 :mod (t2 / test-01~e.13 :quant (m / multiple~e.12)) :ARG1-of (a3 / accord-02 :ARG2 (a4 / and :op1 (p / person :name (n5 / name :op1 "Benjamini"~e.9)) :op2 (p2 / person :name (n6 / name :op1 "Hochberg"~e.11))))))) :ARG1-of (d3 / describe-01 :ARG0 (f / figure~e.52 :mod 3~e.53 :ARG2-of (s3 / supplement-01~e.51)))) # ::id a_pmid_2256_9000.161 ::amr-annotator SDL-AMR-09 ::preferred # ::tok Table 2 lists the differentially expressed genes in selumetinib @-@ resistant cancer cell lines . # ::alignments 1-1.1 2-1.1.1 4-1 6-1.2.1.2 6-1.2.1.2.r 7-1.2.1 8-1.2 9-1.2.1.1.r 10-1.2.1.1.1.1.1.1 12-1.2.1.1.1 13-1.2.1.1.2.2.1 14-1.2.1.1 15-1.2.1.1 (l / list-01~e.4 :ARG0 (t / table~e.1 :mod 2~e.2) :ARG1 (g / gene~e.8 :ARG2-of (e / express-03~e.7 :ARG3~e.9 (c / cell-line~e.14,15 :ARG0-of (r / resist-01~e.12 :ARG1 (s / small-molecule :name (n2 / name :op1 "selumetinib"~e.10))) :mod (d / disease :wiki "Cancer" :name (n / name :op1 "cancer"~e.13))) :manner~e.6 (d2 / differential~e.6)))) # ::id a_pmid_2256_9000.162 ::amr-annotator SDL-AMR-09 ::preferred # ::tok Among the 21 genes that were upregulated in selumetinib @-@ resistant cancer cell lines , we identified two genes , ADCY7 and AKAP13 , which are involved in the cAMP @-@ dependent protein kinase ( PKA ) pathway ( Table 2A ; Supplementary Figure 3 ) . # ::alignments 0-1.2.4 2-1.2.4.1.1 3-1.2.4.1 6-1.2.4.1.2 7-1.2.4.1.2.1.r 8-1.2.4.1.2.1.1.1.1.1 10-1.2.4.1.2.1.1 11-1.2.4.1.2.1.2.2.1 12-1.2.4.1.2.1 13-1.2.4.1.2.1 15-1.1 16-1 18-1.2.1 18-1.2.2 21-1.2.1.1.1 23-1.2 25-1.2.2.1.1 30-1.2.3 31-1.2.3.1.r 33-1.2.3.1.1.1 35-1.2.3.1.1.1 36-1.2.3.1.1.2 37-1.2.3.1.1.3 41-1.2.3.1 44-1.3.1.1 45-1.3.1.1.1 49-1.3.1.2.2 50-1.3.1.2 51-1.3.1.2.1 (i / identify-01~e.16 :ARG0 (w / we~e.15) :ARG1 (a / and~e.23 :op1 (g / gene~e.18 :name (n / name :op1 "ADCY7"~e.21)) :op2 (g2 / gene~e.18 :name (n2 / name :op1 "AKAP13"~e.25)) :ARG1-of (i2 / involve-01~e.30 :ARG2~e.31 (p / pathway~e.41 :name (n3 / name :op1 "cAMP-dependent"~e.33,35 :op2 "protein"~e.36 :op3 "kinase"~e.37))) :ARG1-of (i3 / include-91~e.0 :ARG2 (g3 / gene~e.3 :quant 21~e.2 :ARG1-of (u / upregulate-01~e.6 :location~e.7 (c / cell-line~e.12,13 :ARG0-of (r / resist-01~e.10 :ARG1 (s / small-molecule :name (n4 / name :op1 "selumetinib"~e.8))) :mod (d2 / disease :wiki "Cancer" :name (n5 / name :op1 "cancer"~e.11))))))) :ARG1-of (d / describe-01 :ARG0 (a2 / and :op1 (t / table~e.44 :mod "2A"~e.45) :op2 (f / figure~e.50 :mod 3~e.51 :ARG2-of (s2 / supplement-01~e.49))))) # ::id a_pmid_2256_9000.163 ::amr-annotator SDL-AMR-09 ::preferred # ::tok The ADCY7 gene encodes a membrane @-@ bound adenylatecyclase that convert ATP into 3 ', 5 ' @-@ adenosine monophosphate ( cAMP ) and pyrophosphate ( Supplementary Figure 4 ) . # ::alignments 2-1.1.1.1 4-1.1 5-1 7-1.2.2.1 9-1.2 9-1.2.2 9-1.2.2.r 10-1.2.1.1 12-1.2.3 13-1.2.3.1.1.1 14-1.2.3.2.r 15-1.2.3.2.1.1.1 17-1.2.3.2.1.1.2 18-1.2.3.2.1.1.1 18-1.2.3.2.1.1.2 20-1.2.3.2.1.1.3 21-1.2.3.2.1.1.4 25-1.2.3.2 26-1.2.3.2.2.1.1 29-1.3.1.2 30-1.3.1 31-1.3.1.1 (e / encode-01~e.5 :ARG0 (g / gene~e.4 :name (n / name :op1 "ADCY7"~e.2)) :ARG1 (e2 / enzyme~e.9 :name (n2 / name :op1 "adenylatecyclase"~e.10) :ARG1-of~e.9 (b / bind-01~e.9 :ARG2 (m2 / membrane~e.7)) :ARG0-of (c / convert-01~e.12 :ARG1 (s / small-molecule :name (n3 / name :op1 "ATP"~e.13)) :ARG2~e.14 (a / and~e.25 :op1 (s2 / small-molecule :name (n4 / name :op1 "3'"~e.15,18 :op2 "5'"~e.17,18 :op3 "adenosine"~e.20 :op4 "monophosphate"~e.21)) :op2 (s3 / small-molecule :name (n5 / name :op1 "pyrophosphate"~e.26))))) :ARG1-of (d / describe-01 :ARG0 (f / figure~e.30 :mod 4~e.31 :ARG2-of (s4 / supplement-01~e.29)))) # ::id a_pmid_2256_9000.164 ::amr-annotator SDL-AMR-09 ::preferred # ::tok The cAMP is a second messenger that has a key role in intracellular signalling transduction . # ::alignments 1-1.1.1.1.1 2-1.1.1.r 4-1.1.2 4-1.1.2.1 4-1.1.2.1.r 5-1.1 7-1 9-1.2.1 10-1.2 11-1.2.2.r 12-1.2.2.1.1 13-1.2.2.1 14-1.2.2 (h / have-03~e.7 :ARG0 (m / messenger~e.5 :domain~e.2 (s3 / small-molecule :name (n / name :op1 "cAMP"~e.1)) :ord (o / ordinal-entity~e.4 :value~e.4 2~e.4)) :ARG1 (r / role~e.10 :ARG1-of (k / key-02~e.9) :prep-in~e.11 (t / transduce-01~e.14 :ARG1 (s2 / signal-07~e.13 :mod (i / intracellular~e.12))))) # ::id a_pmid_2256_9000.165 ::amr-annotator SDL-AMR-09 ::preferred # ::tok A major function in mammalian cells is the activation of the PKA ( Tasken et al , 1997 ) . # ::alignments 1-1.2 2-1 3-1.3.r 4-1.3.1.1.1 5-1.3 8-1.1 9-1.1.1.r 11-1.1.1.1.1 14-1.4.1.1.1.1.1 16-1.4.1.1 17-1.4.1.1.2.1 20-1.4.1.2.1 (f / function-01~e.2 :ARG1 (a / activate-01~e.8 :ARG1~e.9 (e / enzyme :name (n / name :op1 "PKA"~e.11))) :ARG1-of (m / major-02~e.1) :location~e.3 (c / cell~e.5 :part-of (a2 / animal :name (n2 / name :op1 "Mammalia"~e.4))) :ARG1-of (d / describe-01 :ARG0 (p / publication-91 :ARG0 (a3 / and~e.16 :op1 (p3 / person :name (n3 / name :op1 "Tasken"~e.14)) :op2 (p2 / person :mod (o / other~e.17))) :time (d2 / date-entity :year 1997~e.20)))) # ::id a_pmid_2256_9000.166 ::amr-annotator SDL-AMR-09 ::preferred # ::tok The AKAP13 gene encodes the A @-@ kinase anchor protein 13 that has the function of binding to the regulatory subunits of PKA ( Supplementary Figure 4 ) . # ::alignments 2-1.1.1.1 4-1.1 5-1 7-1.2.1.1 9-1.2.1.1 10-1.2.1.2 11-1.2.1.3 12-1.2.1.4 16-1.2 16-1.2.2 16-1.2.2.r 17-1.2.2.1.r 18-1.2.2.1 19-1.2.2.1.2.r 21-1.2.2.1.2.1 22-1.2.2.1.2 23-1.2.2.1.2.2.r 24-1.2.2.1.2.2.1.1 27-1.3.1.2 28-1.3.1 29-1.3.1.1 (e / encode-01~e.5 :ARG0 (g / gene~e.4 :name (n / name :op1 "AKAP13"~e.2)) :ARG1 (e2 / enzyme~e.16 :name (n2 / name :op1 "A-kinase"~e.7,9 :op2 "anchor"~e.10 :op3 "protein"~e.11 :op4 13~e.12) :ARG0-of~e.16 (f / function-01~e.16 :ARG1~e.17 (b / bind-01~e.18 :ARG1 e2 :ARG2~e.19 (s / subunit~e.22 :ARG0-of (r / regulate-01~e.21) :part-of~e.23 (e3 / enzyme :name (n3 / name :op1 "PKA"~e.24)))))) :ARG1-of (d / describe-01 :ARG0 (f2 / figure~e.28 :mod 4~e.29 :ARG2-of (s2 / supplement-01~e.27)))) # ::id a_pmid_2256_9000.167 ::amr-annotator SDL-AMR-09 ::preferred # ::tok Moreover , four genes ( NCOA3 , TAF3 , NR1H2 , and RXRA ) , which are upregulated in selumetinib @-@ resistant cancer cell lines , belong to the retinoic acid pathway , which is also activated by PKA ( Altucci et al , 2007 ) . # ::alignments 0-1.1.3.1.1 2-1.1.1.1 3-1.1.1 6-1.1.1.2.1.1.1.1 8-1.1.1.2.1.2.1.1 10-1.1.1.2.1.3.1.1 13-1.1.1.2.1 15-1.1.1.2.1.4.1.1 21-1.1.1.3 22-1.1.1.3.1.r 23-1.1.1.3.1.1.1.1.1 25-1.1.1.3.1.1 26-1.1.1.3.1.2.2.1 27-1.1.1.3.1 28-1.1.1.3.1 30-1.1 31-1.1.2.r 33-1.1.2.1.1 34-1.1.2.1.2 35-1.1.2 39-1.1.2.2.2 40-1.1.2.2 41-1.1.2.2.1.r 42-1.1.2.2.1.1.1 45-1.1.3.1.1.1.1.1 47-1 47-1.1.3.1.1 48-1.1.3.1.1.2.1 51-1.1.3.1.2.1 (a / and~e.47 :op2 (b / belong-01~e.30 :ARG0 (g5 / gene~e.3 :quant 4~e.2 :ARG1-of (m / mean-01 :ARG2 (a2 / and~e.13 :op1 (g / gene :name (n / name :op1 "NCOA3"~e.6)) :op2 (g2 / gene :name (n2 / name :op1 "TAF3"~e.8)) :op3 (g3 / gene :name (n3 / name :op1 "NR1H2"~e.10)) :op4 (g4 / gene :name (n4 / name :op1 "RXRA"~e.15)))) :ARG1-of (u / upregulate-01~e.21 :location~e.22 (c / cell-line~e.27,28 :ARG0-of (r / resist-01~e.25 :ARG1 (s / small-molecule :name (n5 / name :op1 "selumetinib"~e.23))) :mod (d3 / disease :wiki "Cancer" :name (n9 / name :op1 "cancer"~e.26))))) :ARG1~e.31 (p / pathway~e.35 :name (n6 / name :op1 "retinoic"~e.33 :op2 "acid"~e.34) :ARG1-of (a3 / activate-01~e.40 :ARG0~e.41 (e / enzyme :name (n7 / name :op1 "PKA"~e.42)) :mod (a4 / also~e.39))) :ARG1-of (d / describe-01 :ARG0 (p2 / publication-91 :ARG0 (a5 / and~e.0,47 :op1 (p3 / person :name (n8 / name :op1 "Altucci"~e.45)) :op2 (p4 / person :mod (o / other~e.48))) :time (d2 / date-entity :year 2007~e.51))))) # ::id a_pmid_2256_9000.168 ::amr-annotator SDL-AMR-09 ::preferred # ::tok Treatment with selective PKA inhibitors sensitises selumetinib @-@ resistant cancer cell lines to selumetinib # ::alignments 0-1.1 1-1.1.1.r 2-1.1.1.2 3-1.1.1.1.1.1.1 4-1.1.1 4-1.1.1.1 4-1.1.1.1.r 6-1.2.1.1.1.1 8-1.2.1 9-1.2.2.2.1 10-1.2 11-1.2 13-1.3 (s / sensitize-01 :ARG0 (t / treat-04~e.0 :ARG2~e.1 (m / molecular-physical-entity~e.4 :ARG0-of~e.4 (i / inhibit-01~e.4 :ARG1 (p / protein-family :name (n / name :op1 "PKA"~e.3))) :mod (s2 / selective~e.2))) :ARG1 (c / cell-line~e.10,11 :ARG0-of (r / resist-01~e.8 :ARG1 (s3 / small-molecule :name (n2 / name :op1 "selumetinib"~e.6))) :mod (d / disease :wiki "Cancer" :name (n3 / name :op1 "cancer"~e.9))) :ARG2 s3~e.13) # ::id a_pmid_2256_9000.169 ::amr-annotator SDL-AMR-09 ::preferred # ::tok To functionally evaluate if the cAMP @-@ dependent PKA could mediate resistance to MEK inhibitor treatment , we have treated the panel of NSCLC and CRC with 8 @-@ Cl @-@ cAMP , a site @-@ selective cAMP analogue , which specifically inhibits PKAI , the PKA isoform that is directly involved in mitogenic signalling and the transformed phenotype ( Tortora and Ciardiello , 2000 ; Naviglio et al , 2009 ) . # ::alignments 1-1.4.3 1-1.4.3.r 2-1.4 5-1.3.2.1.1.1 7-1.4.2.2.1 7-1.4.2.2.1.2 7-1.4.2.2.1.2.r 8-1.4.2.2.1.1.1 9-1.4.2 10-1.4.2.2 11-1.4.2.2.2 13-1.4.2.2.2.1.1.1.1.1.1 14-1.3 14-1.3.3 14-1.3.3.r 14-1.4.2.2.2.1.1 14-1.4.2.2.2.1.1.1 14-1.4.2.2.2.1.1.1.r 15-1 15-1.4.2.2.2.1 17-1.1 19-1 21-1.2 22-1.2.1.r 23-1.2.1.1.1.1 25-1.2.2.1.1.1 27-1.3.1.1 29-1.3.1.1 31-1.3.2.1.1.1 34-1.3.2.2.1 36-1.3.2.2 37-1.3.2.1.1.1 38-1.3.2 41-1.3.3.2 42-1.3 42-1.3.3 42-1.3.3.r 43-1.3.3.1.1.1 46-1.3.3.1.2.1.1.1 47-1.3.3.1.2 50-1.3.3.1.2.2.2 51-1.3.3.1.2.2 52-1.3.3.1.2.2.1.r 53-1.3.3.1.2.2.1.1.1 54-1.3.3.1.2.2.1.1 55-1.3.3.1.2.2.1 57-1.3.3.1.2.2.1.2.1 58-1.3.3.1.2.2.1.2 61-1.5.1.1.1.1.1.1 62-1.5.1.1.1 63-1.5.1.1.1.2.1.1 65-1.5.1.1.2.1 69-1.5.1.2.1.1.1.1 71-1.5.1 71-1.5.1.2.1 72-1.5.1.2.1.2.1 75-1.5.1.2.2.1 (t / treat-04~e.15,19 :ARG0 (w / we~e.17) :ARG1 (p / panel~e.21 :part~e.22 (c / cell-line :mod (d6 / disease :name (n / name :op1 "NSCLC"~e.23))) :part (c2 / cell-line :mod (d7 / disease :name (n2 / name :op1 "CRC"~e.25)))) :ARG2 (s / small-molecule~e.14,42 :name (n3 / name :op1 "8-Cl-cAMP"~e.27,29) :mod (a2 / analogue~e.38 :poss (s2 / small-molecule :name (n4 / name :op1 "cAMP"~e.5,31,37)) :mod (s3 / selective~e.36 :mod (s4 / site~e.34))) :ARG0-of~e.14,42 (i / inhibit-01~e.14,42 :ARG1 (e / enzyme :name (n5 / name :op1 "PKAI"~e.43) :mod (i2 / isoform~e.47 :poss (e2 / enzyme :name (n6 / name :op1 "PKA"~e.46)) :ARG1-of (i3 / involve-01~e.51 :ARG2~e.52 (a3 / and~e.55 :op1 (s6 / signal-07~e.54 :mod (m / mitogenic~e.53)) :op2 (p2 / phenotype~e.58 :ARG1-of (t2 / transform-01~e.57))) :ARG1-of (d / direct-02~e.50)))) :ARG1-of (s5 / specific-02~e.41))) :purpose (e3 / evaluate-01~e.2 :ARG0 w :ARG1 (p3 / possible-01~e.9 :mode interrogative :ARG1 (m2 / mediate-01~e.10 :ARG0 (e4 / enzyme~e.7 :name (n7 / name :op1 "PKA"~e.8) :ARG0-of~e.7 (d2 / depend-01~e.7 :ARG1 s2)) :ARG1 (r / resist-01~e.11 :ARG1 (t3 / treat-04~e.15 :ARG2 (m3 / molecular-physical-entity~e.14 :ARG0-of~e.14 (i4 / inhibit-01~e.14 :ARG1 (p10 / protein-family :name (n8 / name :op1 "MEK"~e.13)))))))) :manner~e.1 (f / functional~e.1)) :ARG1-of (d3 / describe-01 :ARG0 (a4 / and~e.71 :op1 (p4 / publication-91 :ARG0 (a5 / and~e.62 :op1 (p5 / person :name (n9 / name :op1 "Tortora"~e.61)) :op2 (p6 / person :name (n10 / name :op1 "Ciardiello"~e.63))) :time (d4 / date-entity :year 2000~e.65)) :op2 (p7 / publication-91 :ARG0 (a6 / and~e.71 :op1 (p8 / person :name (n11 / name :op1 "Naviglio"~e.69)) :op2 (p9 / person :mod (o / other~e.72))) :time (d5 / date-entity :year 2009~e.75))))) # ::id a_pmid_2256_9000.170 ::amr-annotator SDL-AMR-09 ::preferred # ::tok A dose @-@ dependent inhibition of growth was observed in all cancer cell lines with a different degree of sensitivity , as illustrated in Figure 5A @ . # ::alignments 1-1.1.2.1 3-1.1.2 4-1.1 5-1.1.1.r 6-1.1.1 8-1 9-1.2.r 10-1.2.2 11-1.2.3.2.1 12-1.2 13-1.2 16-1.2.1.1 16-1.2.1.1.2 16-1.2.1.1.2.r 17-1.2.1.1.1.r 19-1.2.1.1.1 21-1.3.r 22-1.3 25-1.3.1 26-1.3.1.1 (o / observe-01~e.8 :ARG1 (i / inhibit-01~e.4 :ARG1~e.5 (g / grow-01~e.6) :ARG0-of (d / depend-01~e.3 :ARG1 (d2 / dose~e.1))) :location~e.9 (c / cell-line~e.12,13 :ARG0-of (h / have-03 :ARG1 (t / thing~e.16 :degree-of~e.17 (s / sensitive-03~e.19) :ARG1-of~e.16 (d4 / differ-02~e.16))) :mod (a / all~e.10) :mod (d5 / disease :wiki "Cancer" :name (n / name :op1 "cancer"~e.11))) :ARG1-of~e.21 (i2 / illustrate-01~e.22 :ARG0 (f / figure~e.25 :mod "5A"~e.26))) # ::id a_pmid_2256_9000.171 ::amr-annotator SDL-AMR-09 ::preferred # ::tok To evaluate the interaction between selumetinib and 8 @-@ Cl @-@ cAMP , combination analyses were done . # ::alignments 1-1.2 3-1.2.1 5-1.2.1.1.1.1 7-1.2.1.2.1.1 9-1.2.1.2.1.1 11-1.2.1.2.1.1 13-1.1 14-1 (a / analyze-01~e.14 :mod (c / combine-01~e.13) :purpose (e / evaluate-01~e.1 :ARG1 (i / interact-01~e.3 :ARG0 (s / small-molecule :name (n / name :op1 "selumetinib"~e.5)) :ARG1 (s2 / small-molecule :name (n2 / name :op1 "8-Cl-cAMP"~e.7,9,11))))) # ::id a_pmid_2256_9000.172 ::amr-annotator SDL-AMR-09 ::preferred # ::tok The CRC and NSCLC cancer cells were treated with different concentrations of selumetinib ( range , 0.01 @–@ 10 μ @ ℳ) and 8 @-@ Cl @-@ cAMP ( range , 0.01 @–@ 5 μ @ ℳ) each day , for a total of 3 days at a fixed drug ratio selumetinib to 8 @-@ Cl @-@ cAMP of 1 : 1 . # ::alignments 1-1.1.2.1.1.1 2-1.1 3-1.1.1.1.1.1 4-1.1.3.2.1 5-1.1.1 5-1.1.2 7-1 9-1.2.1.2.3 9-1.2.2.2.1 10-1.2.1.2 10-1.2.2.2 11-1.2.r 12-1.2.1.1.1 16-1.2.1.2.1.1 18-1.2.1.2.1.2 24-1.2.2.1.1 26-1.2.2.1.1 28-1.2.2.1.1 32-1.2.2.2.2.1 34-1.2.2.2.2.2 40-1.3.1.2 40-1.5.2 42-1.5.r 44-1.5.3 46-1.5.1 47-1.3.1.2 47-1.5.2 50-1.4.1.3 52-1.4.1 53-1.4.1.1 55-1.2.2.1.1 57-1.2.2.1.1 59-1.2.2.1.1 60-1.4.2 61-1.3.1.1 61-1.4.2.1 61-1.4.2.2 63-1.3.1.1 63-1.4.2.1 (t / treat-04~e.7 :ARG1 (a / and~e.2 :op1 (c / cell-line~e.5 :mod (d6 / disease :name (n / name :op1 "NSCLC"~e.3))) :op2 (c2 / cell-line~e.5 :mod (d7 / disease :name (n2 / name :op1 "CRC"~e.1))) :mod (d5 / disease :wiki "Cancer" :name (n5 / name :op1 "cancer"~e.4))) :ARG2~e.11 (a2 / and :op1 (s / small-molecule :name (n3 / name :op1 "selumetinib"~e.12) :mod (c4 / concentration-quantity~e.10 :quant (v / value-interval :op1 0.01~e.16 :op2 10~e.18) :unit (m / micromolar) :ARG1-of (d3 / differ-02~e.9))) :op2 (s2 / small-molecule :name (n4 / name :op1 "8-Cl-cAMP"~e.24,26,28,55,57,59) :mod (c5 / concentration-quantity~e.10 :ARG1-of (d4 / differ-02~e.9) :quant (v2 / value-interval :op1 0.01~e.32 :op2 5~e.34) :unit m))) :frequency (r2 / rate-entity-91 :ARG3 (t2 / temporal-quantity :quant 1~e.61,63 :unit (d / day~e.40,47))) :condition (e / equal-01 :ARG1 (r3 / ratio-of~e.52 :op1 s~e.53 :op2 s2 :ARG1-of (f / fix-03~e.50)) :ARG2 (r4 / ratio-of~e.60 :op1 1~e.61,63 :op2 1~e.61)) :duration~e.42 (t3 / temporal-quantity :quant 3~e.46 :unit (d2 / day~e.40,47) :mod (t4 / total~e.44))) # ::id a_pmid_2256_9000.173 ::amr-annotator SDL-AMR-09 ::preferred # ::tok In all selumetinib @-@ resistant cancer cell lines , the combination treatment caused synergistic growth inhibitory effects . # ::alignments 1-1.3.2 2-1.3.1.1.1.1 4-1.3.1 5-1.3.3.2.1 6-1.3 7-1.3 10-1.1.1 11-1.1 12-1 13-1.2.2 14-1.2.1.1 15-1.2.1 16-1.2 (c / cause-01~e.12 :ARG0 (t / treat-04~e.11 :mod (c2 / combine-01~e.10)) :ARG1 (a / affect-01~e.16 :ARG2 (i / inhibit-01~e.15 :ARG1 (g / grow-01~e.14)) :ARG0-of (s2 / synergize-01~e.13)) :location (c3 / cell-line~e.6,7 :ARG0-of (r / resist-01~e.4 :ARG1 (s / small-molecule :name (n / name :op1 "selumetinib"~e.2))) :mod (a2 / all~e.1) :mod (d / disease :wiki "Cancer" :name (n2 / name :op1 "cancer"~e.5)))) # ::id a_pmid_2256_9000.174 ::amr-annotator SDL-AMR-09 ::preferred # ::tok In fact , the CI values for the combinations treatments ranged between 0.013 and 0.350 . # ::alignments 0-1.4 1-1.4 4-1.3.1 4-1.3.1.1 4-1.3.1.1.r 5-1.3 6-1.3.2.r 8-1.3.2.1 9-1.3.2 10-1 12-1.1 14-1.2 (r / range-01~e.10 :ARG3 0.013~e.12 :ARG4 0.350~e.14 :ARG1 (v / value~e.5 :mod (i / interval~e.4 :mod~e.4 (c / confidence~e.4)) :poss~e.6 (t / treat-04~e.9 :mod (c2 / combine-01~e.8))) :mod (i2 / in-fact~e.0,1)) # ::id a_pmid_2256_9000.175 ::amr-annotator SDL-AMR-09 ::preferred # ::tok This was significantly different in the selumetinib @-@ sensitive cancer cells in which the combination treatment was clearly antagonistic with CI between 1.3 and 3.6 ( Figure 5B ) . # ::alignments 0-1.1 2-1.4 3-1 4-1.2.r 6-1.2.1.1.1.1 8-1.2.1 9-1.2.4.2.1 10-1.2 14-1.2.2.1.1 15-1.2.2.1 17-1.2.2.2 20-1.2.3.1 20-1.2.3.1.1 20-1.2.3.1.1.r 20-1.2.3.1.2 20-1.2.3.1.2.r 22-1.2.3.1.2.1 24-1.2.3.1.2.2 27-1.3.1 28-1.3.1.1 (d2 / differ-02~e.3 :ARG1 (t / this~e.0) :location~e.4 (c / cell-line~e.10 :ARG0-of (s2 / sensitive-03~e.8 :ARG1 (s3 / small-molecule :name (n / name :op1 "selumetinib"~e.6))) :location-of (a / antagonize-02 :ARG1 (t2 / treat-04~e.15 :mod (c3 / combine-01~e.14)) :ARG1-of (c2 / clear-06~e.17)) :ARG0-of (h / have-03 :ARG1 (i / interval~e.20 :mod~e.20 (c4 / confidence~e.20) :mod~e.20 (v / value-interval~e.20 :op1 1.3~e.22 :op2 3.6~e.24))) :mod (d3 / disease :wiki "Cancer" :name (n2 / name :op1 "cancer"~e.9))) :ARG1-of (d / describe-01 :ARG0 (f / figure~e.27 :mod "5B"~e.28)) :ARG1-of (s4 / significant-02~e.2)) # ::id a_pmid_2256_9000.176 ::amr-annotator SDL-AMR-09 ::preferred # ::tok We next assessed the phosphorylation state of MEK and MAPK following treatment with selumetinib , 8 @-@ Cl @-@ cAMP as single agents or in combination with HCT15 and H460 cancer cell lines . # ::alignments 0-1.1 1-1.3 2-1 4-1.2.1 5-1.2 6-1.2.1.1.r 7-1.2.1.1.1.1.1 8-1.2.1.1 9-1.2.1.1.2.1.1 10-1.2.2 11-1.2.2.1 13-1.2.2.1.1.1.1.1.1 15-1.2.2.1.1.1.2.1.1 17-1.2.2.1.1.1.2.1.1 19-1.2.2.1.1.1.2.1.1 21-1.2.2.1.1.1.3.1.1 22-1.2.2.1.1.1.3.1 23-1.2.2.1.1 23-1.2.2.1.1.1 23-1.2.2.1.1.1.r 24-1.2.2.1.1.r 25-1.2.2.1.1.2 26-1.2.2.1.1.2.2.r 27-1.2.2.1.1.2.2.1.1.1 28-1.2.2.1.1.2.2 29-1.2.2.1.1.2.2.2.1.1 30-1.2.2.1.1.2.2.3.2.1 31-1.2.2.1.1.2.2.1 31-1.2.2.1.1.2.2.2 32-1.2.2.1.1.2.2.1 (a / assess-01~e.2 :ARG0 (w / we~e.0) :ARG1 (s / state~e.5 :mod (p / phosphorylate-01~e.4 :ARG1~e.6 (a2 / and~e.8 :op1 (e / enzyme :name (n2 / name :op1 "MEK"~e.7)) :op2 (e2 / enzyme :name (n3 / name :op1 "MAPK"~e.9)))) :ARG1-of (f / follow-01~e.10 :ARG2 (t / treat-04~e.11 :ARG2~e.24 (o / or~e.23 :op1~e.23 (o2 / or~e.23 :op1 (s2 / small-molecule :name (n4 / name :op1 "selumetinib"~e.13)) :op2 (s3 / small-molecule :name (n5 / name :op1 "8-Cl-cAMP"~e.15,17,19)) :ARG0-of (a3 / act-01 :ARG1 (a4 / agent~e.22 :ARG1-of (s4 / single-02~e.21)))) :op2 (c / combine-01~e.25 :ARG1 (a5 / and :op1 s2 :op2 s3) :ARG2~e.26 (a6 / and~e.28 :op1 (c2 / cell-line~e.31,32 :name (n6 / name :op1 "HCT15"~e.27)) :op2 (c3 / cell-line~e.31 :name (n7 / name :op1 "H460"~e.29)) :mod (d / disease :wiki "Cancer" :name (n8 / name :op1 "cancer"~e.30)))))))) :mod (n / next~e.1)) # ::id a_pmid_2256_9000.177 ::amr-annotator SDL-AMR-09 ::preferred # ::tok As shown in Figure 3C , single agent selumetinib or 8 @-@ Cl @-@ cAMP slightly inhibited p @-@ MEK and p @-@ MAPK , whereas the combination induced a significant inhibition of both phosphorylated proteins . # ::alignments 1-1.3 4-1.3.1 5-1.3.1.1 8-1.1.1.3.1.1 9-1.1.1.3.1 10-1.1.1.1.1.1 11-1.1.1 12-1.1.1.2.1.1 14-1.1.1.2.1.1 16-1.1.1.2.1.1 17-1.1.3 18-1.1 19-1.1.2.1.2 21-1.1.2.1.1.1 23-1.1.2.1.2 25-1.1.2.2.1.1 27-1 29-1.2.1 30-1.2 32-1.2.2.2 33-1.2.2 36-1.2.2.1 (c / contrast-01~e.27 :ARG1 (i / inhibit-01~e.18 :ARG0 (o / or~e.11 :op1 (s2 / small-molecule :name (n / name :op1 "selumetinib"~e.10)) :op2 (s3 / small-molecule :name (n2 / name :op1 "8-Cl-cAMP"~e.12,14,16)) :ARG0-of (a / act-01 :ARG1 (a2 / agent~e.9 :ARG1-of (s4 / single-02~e.8)))) :ARG1 (a3 / and :op1 (e / enzyme :name (n3 / name :op1 "MEK"~e.21) :ARG3-of (p5 / phosphorylate-01~e.19,23)) :op2 (e2 / enzyme :name (n4 / name :op1 "MAPK"~e.25) :ARG3-of p5)) :degree (s / slight~e.17)) :ARG2 (i2 / induce-01~e.30 :ARG0 (c2 / combine-01~e.29 :ARG1 s2 :ARG2 s3) :ARG1 (i3 / inhibit-01~e.33 :ARG1 a3~e.36 :ARG1-of (s7 / significant-02~e.32))) :ARG1-of (s6 / show-01~e.1 :ARG0 (f / figure~e.4 :mod "3C"~e.5))) # ::id a_pmid_2256_9000.178 ::amr-annotator SDL-AMR-09 ::preferred # ::tok A genetic approach to inhibiting PKAI expression was developed by the use of phosphorothioate antisense oligonucleotides targeting the synthesis of the PKAI regulatory subunit RI @ α @ . # ::alignments 1-1.1.2 2-1.1 3-1.1.1.r 4-1.1.1 5-1.1.1.1.1.1.1 6-1.1.1.1 8-1 9-1.2.r 11-1.2 12-1.2.1.r 13-1.2.1.3 15-1.2.1 16-1.2.1.1 18-1.2.1.1.1 19-1.2.1.1.1.1.r 21-1.2.1.1.1.1.2 22-1.2.1.1.1.1 22-1.2.1.1.1.1.3 22-1.2.1.1.1.1.3.r (d / develop-02~e.8 :ARG1 (a / approach-02~e.2 :ARG1~e.3 (i / inhibit-01~e.4 :ARG1 (e / express-03~e.6 :ARG2 (e2 / enzyme :name (n / name :op1 "PKAI"~e.5)))) :mod (g / genetics~e.1)) :manner~e.9 (u / use-01~e.11 :ARG1~e.12 (o / oligonucleotide~e.15 :ARG0-of (t / target-01~e.16 :ARG1 (s / synthesize-01~e.18 :ARG1~e.19 (p / protein-segment~e.22 :name (n3 / name :op1 "RIα") :part-of e2~e.21 :ARG0-of~e.22 (r / regulate-01~e.22)))) :ARG0-of (c / counter-01 :ARG1 (s2 / sense-01)) :mod (p2 / phosphorothioate~e.13)))) # ::id a_pmid_2256_9000.179 ::amr-annotator SDL-AMR-09 ::preferred # ::tok These oligonucleotides inhibit growth of several human cancer cell lines in vitro and in vivo ( Yokozaki et al , 1993 ; Nesterova and Cho @-@ Chung , 1995 ; Tortora et al , 1997a ) . # ::alignments 0-1.1.1 1-1.1 2-1 3-1.2 4-1.2.1.r 5-1.2.1.2 6-1.2.1.1 7-1.2.1.3.2.1 8-1.2.1 9-1.2.1 11-1.3.1 12-1.3.1 14-1.3 16-1.3.1 16-1.3.2 17-1.3.2 21-1.4.1.1.1.1.1.1 23-1.4.1.1.1 24-1.4.1.1.1.2.1 27-1.4.1.1.2.1 31-1.4.1.2.1.1.1.1 32-1.4.1.2.1 33-1.4.1.2.1.2.1.1 35-1.4.1.2.1.2.1.1 37-1.4.1.2.2.1 41-1.4.1.3.2.1.1.1 43-1.4.1 43-1.4.1.1.1 43-1.4.1.3.2 44-1.4.1.1.1.2.1 (i / inhibit-01~e.2 :ARG0 (o / oligonucleotide~e.1 :mod (t / this~e.0)) :ARG1 (g / grow-01~e.3 :ARG1~e.4 (c / cell-line~e.8,9 :mod (h / human~e.6) :quant (s / several~e.5) :mod (d5 / disease :wiki "Cancer" :name (n5 / name :op1 "cancer"~e.7)))) :manner (a / and~e.14 :op1 (i2 / in-vitro~e.11,12,16) :op2 (i3 / in-vivo~e.16,17)) :ARG1-of (d / describe-01 :ARG0 (a2 / and~e.43 :op1 (p / publication-91 :ARG0 (a3 / and~e.23,43 :op1 (p4 / person :name (n / name :op1 "Yokozaki"~e.21)) :op2 (p5 / person :mod (o2 / other~e.24,44))) :time (d2 / date-entity :year 1993~e.27)) :op2 (p2 / publication-91 :ARG0 (a4 / and~e.32 :op1 (p6 / person :name (n2 / name :op1 "Nesterova"~e.31)) :op2 (p7 / person :name (n3 / name :op1 "Cho-Chung"~e.33,35))) :time (d3 / date-entity :year 1995~e.37)) :op3 (p3 / publication-91 :li "a" :ARG0 (a5 / and~e.43 :op1 (p8 / person :name (n4 / name :op1 "Tortora"~e.41)) :op2 p5) :time (d4 / date-entity :year 1997))))) # ::id a_pmid_2256_9000.180 ::amr-annotator SDL-AMR-09 ::preferred # ::tok Therefore , HYB 190 , an 18 @-@ mer MBO antisense to the N @-@ terminal 8 @–@ 13 codons of the RI @ α subunit of PKAI , and the control HYB 239 , containing four mismatched bases , were tested to study the effect on the growth of NSCLC and CRC cell lines . # ::alignments 0-1 2-1.1.1.1.1.1 3-1.1.1.1.1.2 6-1.1.1.1.4.1.1 10-1.1.1.1.3 11-1.1.1.1.3.1.r 13-1.1.1.1.3.1.2.1.1 15-1.1.1.1.3.1.2.1.1 16-1.1.1.1.3.1.1.1 18-1.1.1.1.3.1.1.2 19-1.1.1.1.3.1 27-1.1.1.1.3.1.3.r 28-1.1.1.1.3.1.3.2.1.1 30-1.1.1 32-1.1.1.2 32-1.1.1.2.3 32-1.1.1.2.3.r 33-1.1.1.2.1.1 34-1.1.1.2.1.2 36-1.1.1.2.2 37-1.1.1.2.2.1.1 38-1.1.1.2.2.1.2 39-1.1.1.2.2.1 42-1.1 44-1.1.2 46-1.1.2.1 47-1.1.2.1.1.r 49-1.1.2.1.1 50-1.1.2.1.1.1.r 51-1.1.2.1.1.1.1.1.1.1 52-1.1.2.1.1.1 53-1.1.2.1.1.1.2.1.1.1 54-1.1.2.1.1.1.1 54-1.1.2.1.1.1.2 55-1.1.2.1.1.1.1 (c3 / cause-01~e.0 :ARG1 (t / test-01~e.42 :ARG1 (a3 / and~e.30 :op1 (o / oligonucleotide :name (n3 / name :op1 "HYB"~e.2 :op2 190~e.3) :mod (b2 / backbone :ARG3-of (m / mix-01)) :mod (a4 / antisense~e.10 :prep-to~e.11 (c4 / codon~e.19 :mod (v / value-interval :op1 8~e.16 :op2 13~e.18) :part-of (p / protein-segment :name (n4 / name :op1 "N-terminus"~e.13,15)) :part-of~e.27 (p2 / protein-segment :name (n5 / name :op1 "RIα") :part-of (e2 / enzyme :name (n6 / name :op1 "PKAI"~e.28))))) :mod (p3 / polymer :ARG1-of (l / long-03 :ARG2 18~e.6))) :op2 (s2 / small-molecule~e.32 :name (n7 / name :op1 "HYB"~e.33 :op2 239~e.34) :ARG0-of (c6 / contain-01~e.36 :ARG1 (b / base~e.39 :quant 4~e.37 :mod (m4 / mismatch~e.38))) :ARG0-of~e.32 (c5 / control-01~e.32))) :purpose (s / study-01~e.44 :ARG1 (a / affect-01~e.46 :ARG1~e.47 (g / grow-01~e.49 :ARG1~e.50 (a2 / and~e.52 :op1 (c / cell-line~e.54,55 :mod (d / disease :name (n / name :op1 "NSCLC"~e.51))) :op2 (c2 / cell-line~e.54 :mod (d2 / disease :name (n2 / name :op1 "CRC"~e.53))))))))) # ::id a_pmid_2256_9000.181 ::amr-annotator SDL-AMR-09 ::preferred # ::tok All cancer cell lines treated with HBY 190 displayed a dose @-@ dependent inhibition of growth by MTT assay ( Figure 6A ) . # ::alignments 0-1.1.2 1-1.1.3.2.1 2-1.1 3-1.1 4-1.1.1 5-1.4.1.r 7-1.1.1.1.1.2 8-1 10-1.2.2.1 12-1.2.2 13-1.2 14-1.2.1.r 15-1.2.1 17-1.4.1.1.1 18-1.4 21-1.3.1 22-1.3.1.1 (d2 / display-01~e.8 :ARG0 (c / cell-line~e.2,3 :ARG1-of (t / treat-04~e.4 :ARG2 (s / small-molecule :name (n2 / name :op1 "HYB" :op2 190~e.7))) :mod (a / all~e.0) :mod (d / disease :wiki "Cancer" :name (n / name :op1 "cancer"~e.1))) :ARG1 (i / inhibit-01~e.13 :ARG1~e.14 (g / grow-01~e.15) :ARG0-of (d3 / depend-01~e.12 :ARG1 (d4 / dose~e.10))) :ARG1-of (d5 / describe-01 :ARG0 (f / figure~e.21 :mod "6A"~e.22)) :manner (a2 / assay-01~e.18 :instrument~e.5 (s2 / small-molecule :name (n3 / name :op1 "MTT"~e.17)))) # ::id a_pmid_2256_9000.182 ::amr-annotator SDL-AMR-09 ::preferred # ::tok Growth inhibition was more pronounced in selumetinib @-@ resistant cancer cell lines ( IC @ 50 between 0.25 and 1 μ @ ℳ) , while the IC @ 50 values were > 5 μ @ ℳ in the selumetinib @-@ sensitive cancer cells . # ::alignments 0-1.1.1.1 1-1.1.1 3-1.1.2 4-1.1 5-1.1.3.r 6-1.1.3.1.1.1.1 8-1.1.3.1 9-1.1.3.2.2.1 10-1.1.3 11-1.2.3 13-1.1.3.1.1 13-1.1.3.1.1.2 13-1.1.3.1.1.2.r 13-1.2 15-1.1.3.1.1.2.1 15-1.2.1 18-1.1.3.1.1.2.2.2.1 20-1.1.3.1.1.2.2.2.2 26-1 28-1.1.3.1.1 28-1.1.3.1.1.2 28-1.1.3.1.1.2.r 28-1.2 30-1.1.3.1.1.2.1 30-1.2.1 32-1.1.3.1.1.2.2.2 34-1.2.2 35-1.2.2.1.1 42-1.2.3.1.1 44-1.2.3.1 45-1.2.3.2 46-1.2.3 (c / contrast-01~e.26 :ARG1 (p / pronounced-02~e.4 :ARG1 (i / inhibit-01~e.1 :ARG1 (g / grow-01~e.0)) :degree (m / more~e.3) :location~e.5 (c2 / cell-line~e.10 :ARG0-of (r / resist-01~e.8 :ARG1 (s / small-molecule~e.13,28 :name (n2 / name :op1 "selumetinib"~e.6) :ARG1-of~e.13,28 (h2 / have-percentage-maximal-inhibitory-concentration-01~e.13,28 :ARG2 50~e.15,30 :ARG4 (c4 / concentration-quantity :unit (m3 / micromolar) :quant (v / value-interval~e.32 :op1 0.25~e.18 :op2 1~e.20))))) :mod (d / disease :wiki "Cancer" :name (n / name :op1 "cancer"~e.9)))) :ARG2 (h3 / have-percentage-maximal-inhibitory-concentration-01~e.13,28 :ARG2 50~e.15,30 :ARG4 (m5 / more-than~e.34 :op1 (c7 / concentration-quantity :quant 5~e.35 :unit m3)) :location (c5 / cell-line~e.11,46 :ARG0-of (s2 / sensitive-03~e.44 :ARG1 s~e.42) :mod d~e.45))) # ::id a_pmid_2256_9000.183 ::amr-annotator SDL-AMR-09 ::preferred # ::tok In contrast , the control oligonucleotide , HYB 239 , at doses up to 5 μ @ ℳ, showed only 5 @–@ 15 % growth inhibition among all of the cell lines tested ( Figure 6A ) . # ::alignments 1-1 4-1.1.1.3 5-1.1.1 7-1.1.1.2.1 8-1.1.1.2.2 11-1.1.3 12-1.1.3.1 13-1.1.3.1 14-1.1.3.1.1.1 19-1.1 20-1.1.2.2.2 21-1.1.2.2.1.1 23-1.1.2.2.1.2 24-1.1.2.2 25-1.1.2.1 26-1.1.2 28-1.1.4.1 31-1.1.4 32-1.1.4 33-1.1.4.2 36-1.2.1 37-1.2.1.1 (c / contrast-01~e.1 :ARG2 (s / show-01~e.19 :ARG0 (o / oligonucleotide~e.5 :wiki - :name (n / name :op1 "HYB"~e.7 :op2 239~e.8) :ARG0-of (c2 / control-01~e.4)) :ARG1 (i / inhibit-01~e.26 :ARG1 (g / grow-01~e.25) :mod (p / percentage-entity~e.24 :value (v / value-interval :op1 5~e.21 :op2 15~e.23) :mod (o2 / only~e.20))) :condition (d / dose~e.11 :quant (u / up-to~e.12,13 :op1 (c4 / concentration-quantity :quant 5~e.14 :unit (m / micromolar)))) :location (c3 / cell-line~e.31,32 :mod (a / all~e.28) :ARG1-of (t / test-01~e.33))) :ARG1-of (d2 / describe-01 :ARG0 (f / figure~e.36 :mod "6A"~e.37))) # ::id a_pmid_2256_9000.184 ::amr-annotator SDL-AMR-09 ::preferred # ::tok To determine whether a combination of selumetinib and oligonucleotide HYB 190 could enhance the antiproliferative effect compared with either agent alone , selumetinib @-@ resistant cells were treated with different combinations of the two agents at a fixed drug ratio of 1 : 1 . # ::alignments 1-1.3 2-1.3.1.1 2-1.3.1.1.r 4-1.3.1.2.1 5-1.3.1.2.1.1.r 6-1.3.1.2.1.1.1.1 8-1.3.1.2.1.2 9-1.3.1.2.1.2.1.1 10-1.3.1.2.1.2.1.2 11-1.3.1 12-1.3.1.2 12-1.3.1.2.3.1 12-1.3.1.2.3.2 14-1.3.1.2.2.1 14-1.3.1.2.2.1.1 14-1.3.1.2.2.1.1.r 15-1.3.1.2.2 16-1.3.1.2.3.r 22-1.1.1.1 24-1.1.1 25-1.1 27-1 28-1.2.r 29-1.2.3 30-1.2 31-1.4.2 37-1.4.1.2 38-1.4.1.1 39-1.4.1 40-1.4.2 41-1.4.2.1 41-1.4.2.2 43-1.4.2.1 (t / treat-04~e.27 :ARG1 (c4 / cell~e.25 :ARG0-of (r3 / resist-01~e.24 :ARG1 s~e.22)) :ARG2~e.28 (c / combine-01~e.30 :ARG1 s :ARG2 o :ARG1-of (d3 / differ-02~e.29)) :purpose (d / determine-01~e.1 :ARG1 (p2 / possible-01~e.11 :mode~e.2 interrogative~e.2 :ARG1 (e / enhance-01~e.12 :ARG0 (c2 / combine-01~e.4 :ARG1~e.5 (s / small-molecule :name (n / name :op1 "selumetinib"~e.6)) :ARG2 (o / oligonucleotide~e.8 :name (n2 / name :op1 "HYB"~e.9 :op2 190~e.10))) :ARG1 (a / affect-01~e.15 :ARG2 (c3 / counter-01~e.14 :ARG1~e.14 (p3 / proliferate-01~e.14))) :compared-to~e.16 (o2 / or :op1 (e2 / enhance-01~e.12 :ARG0 s :ARG1 a) :op2 (e3 / enhance-01~e.12 :ARG0 o :ARG1 a))))) :condition (e4 / equal-01 :ARG1 (r2 / ratio~e.39 :mod (d2 / drug~e.38) :ARG1-of (f / fix-03~e.37)) :ARG2 (r4 / ratio-of~e.31,40 :op1 1~e.41,43 :op2 1~e.41))) # ::id a_pmid_2256_9000.185 ::amr-annotator SDL-AMR-09 ::preferred # ::tok As shown in Figure 6B , the combination treatment caused synergistic growth inhibitory effects . # ::alignments 1-1.3 4-1.3.1 5-1.3.1.1 9-1.1.1 10-1.1 11-1 13-1.2.1.1 14-1.2.1 15-1.2 (c / cause-01~e.11 :ARG0 (t / treat-04~e.10 :mod (c2 / combine-01~e.9)) :ARG1 (a / affect-01~e.15 :ARG2 (i / inhibit-01~e.14 :ARG1 (g / grow-01~e.13))) :ARG1-of (s / show-01~e.1 :ARG0 (f / figure~e.4 :mod "6B"~e.5))) # ::id a_pmid_2256_9000.186 ::amr-annotator SDL-AMR-09 ::preferred # ::tok To determine whether the growth inhibitory effect of oligonucleotide HYB 190 correlated with a reduction in RI @ α protein levels , we performed protein blots on total cell extracts prepared from H460 cells treated with oligonucleotide HYB 190 , oligonucleotide HYB 239 , selumetinib , and combinations . # ::alignments 1-1.3 2-1.3.2.1 2-1.3.2.1.r 4-1.3.2.2.2.2 5-1.3.2.2.2 6-1.3.2.2 8-1.2.2.3.1.2.1.2 8-1.3.2.2.1 9-1.3.2.2.1.1.1 10-1.3.2.2.1.1.2 11-1.3.2 12-1.3.2.3.r 14-1.3.2.3 20-1.3.2.3.1.1 21-1.3.2.3.1 23-1.1 24-1 25-1.2.1 28-1.2.2.2 29-1.2.2.1.1 30-1.2.2 30-1.2.2.1 30-1.2.2.1.r 31-1.2.2.3 32-1.2.2.3.1.r 33-1.2.2.3.1.1.1 34-1.2.2.3.1 35-1.2.2.3.1.2 36-1.2.2.3.1.2.1.r 37-1.2.2.3.1.2.1.1 38-1.2.2.3.1.2.1.1 39-1.2.2.3.1.2.1.1 40-1.2.2.3.1.2.1.1 41-1.2.2.3.1.2.1.1 42-1.2.2.3.1.2.1.2.1.1 43-1.2.2.3.1.2.1.2.1.2 45-1.2.2.3.1.2.1.3.1.1 48-1.2.2.3.1.2.1.4 (p / perform-01~e.24 :ARG0 (w / we~e.23) :ARG1 (i2 / immunoblot-01 :ARG1 (p2 / protein~e.25) :ARG2 (m / molecular-physical-entity~e.30 :ARG1-of~e.30 (e / extract-01~e.30 :ARG2 (c2 / cell~e.29)) :mod (t / total~e.28) :ARG1-of (p4 / prepare-01~e.31 :ARG2~e.32 (c3 / cell-line~e.34 :name (n3 / name :op1 "H460"~e.33) :ARG1-of (t2 / treat-04~e.35 :ARG2~e.36 (o2 / or :op1 o~e.37,38,39,40,41 :op2 (o3 / oligonucleotide~e.8 :name (n4 / name :op1 "HYB"~e.42 :op2 239~e.43)) :op3 (s / small-molecule :name (n5 / name :op1 "selumetinib"~e.45)) :op4 (c4 / combine-01~e.48))))))) :purpose (d / determine-01~e.1 :ARG0 w :ARG1 (c / correlate-01~e.11 :mode~e.2 interrogative~e.2 :ARG1 (a / affect-01~e.6 :ARG0 (o / oligonucleotide~e.8 :name (n / name :op1 "HYB"~e.9 :op2 190~e.10)) :ARG2 (i / inhibit-01~e.5 :ARG0 o3 :ARG1 (g / grow-01~e.4))) :ARG2~e.12 (r / reduce-01~e.14 :ARG1 (l / level~e.21 :quant-of (p3 / protein-segment~e.20 :name (n2 / name :op1 "RIα"))))))) # ::id a_pmid_2256_9000.187 ::amr-annotator SDL-AMR-09 ::preferred # ::tok Compared with untreated H460 cells , RI @ α levels were substantially unchanged , even in cells treated with a higher dose of the control oligonucleotide HYB 239 ( 1 μ @ ℳ) and selumetinib ( 5 μ @ ℳ) . # ::alignments 0-1.2.2.r 2-1.2.2.2 2-1.2.2.2.1 2-1.2.2.2.1.r 3-1.2.2.1.1 4-1.2.2 10-1.2 12-1.3 13-1 13-1.1 13-1.1.r 16-1.4.r 17-1.4.1 18-1.4 19-1.4.2.r 21-1.4.2.1.1 21-1.4.2.1.1.1 21-1.4.2.1.1.1.r 22-1.4.2.1 22-1.4.2.2 23-1.4.2.1.2.r 25-1.4.2.1.2.2 26-1.4.2.1.2 27-1.4.2.1.2.1.1 28-1.4.2.1.2.1.2 30-1.4.2.1.3.1.1 35-1.4.2 36-1.4.2.2.2.1.1 38-1.4.2.2.3.1.1 (c / change-01~e.13 :polarity~e.13 -~e.13 :ARG1 (l / level~e.10 :quant-of (p / protein-segment :name (n / name :op1 "RIα")) :compared-to~e.0 (c2 / cell-line~e.4 :name (n2 / name :op1 "H460"~e.3) :ARG1-of (t / treat-04~e.2 :polarity~e.2 -~e.2))) :degree (s / substantial~e.12) :concession~e.16 (t2 / treat-04~e.18 :ARG1 (c3 / cell~e.17) :ARG2~e.19 (a / and~e.35 :op1 (d / dose~e.22 :ARG1-of (h / high-02~e.21 :degree~e.21 (m / more~e.21)) :quant-of~e.23 (o / oligonucleotide~e.26 :name (n3 / name :op1 "HYB"~e.27 :op2 239~e.28) :ARG0-of (c4 / control-01~e.25)) :ARG1-of (m2 / mean-01 :ARG2 (c5 / concentration-quantity :quant 1~e.30 :unit (m3 / micromolar)))) :op1 (d2 / dose~e.22 :ARG1-of h :quant-of (s2 / small-molecule :name (n4 / name :op1 "selumetinib"~e.36)) :ARG1-of (m5 / mean-01 :ARG2 (m6 / mass-quantity :quant 5~e.38 :unit m3)))))) # ::id a_pmid_2256_9000.188 ::amr-annotator SDL-AMR-09 ::preferred # ::tok However , reduction of RI @ α expression was seen when H460 cells were treated with oligonucleotide HYB 190 ( Figure 6C ) . # ::alignments 0-1 2-1.1.1 8-1.1.1.1 10-1.1 12-1.1.2.1.1.1 13-1.1.2.1 15-1.1.2 16-1.1.2.2.r 17-1.1.2.2 18-1.1.2.2.1.1 19-1.1.2.2.1.2 22-1.1.3.1 23-1.1.3.1.1 (c2 / contrast-01~e.0 :ARG2 (s / see-01~e.10 :ARG1 (r / reduce-01~e.2 :ARG1 (e / express-03~e.8 :ARG2 (p / protein-segment :name (n / name :op1 "RIα")))) :condition (t / treat-04~e.15 :ARG1 (c / cell-line~e.13 :name (n2 / name :op1 "H460"~e.12)) :ARG2~e.16 (o / oligonucleotide~e.17 :name (n3 / name :op1 "HYB"~e.18 :op2 190~e.19))) :ARG1-of (d / describe-01 :ARG0 (f / figure~e.22 :mod "6C"~e.23)))) # ::id a_pmid_2256_9000.189 ::amr-annotator SDL-AMR-09 ::preferred # ::tok Moreover , a complete inhibition of RI @ α expression was seen in the combination treatment ( Figure 6C ) . # ::alignments 0-1 3-1.1.1.2 4-1.1.1 10-1.1.1.1 12-1.1 13-1.1.2.r 15-1.1.2.1 16-1.1.2 19-1.1.3.1 20-1.1.3.1.1 (a / and~e.0 :op2 (s / see-01~e.12 :ARG1 (i / inhibit-01~e.4 :ARG1 (e / express-03~e.10 :ARG2 (p / protein-segment :name (n / name :op1 "RIα"))) :degree (c / complete-02~e.3)) :condition~e.13 (t / treat-04~e.16 :mod (c2 / combine-01~e.15)) :ARG1-of (d / describe-01 :ARG0 (f / figure~e.19 :mod "6C"~e.20)))) # ::id a_pmid_2256_9000.190 ::amr-annotator SDL-AMR-09 ::preferred # ::tok We finally evaluated whether the combined inhibition of both PKA and MEK pathways would have antitumour activity in vivo in selumetinib @-@ resistant HCT15 and H460 xenografts ( Figure 7 ) . # ::alignments 0-1.2 1-1.1 1-1.1.r 2-1 3-1.3.1 3-1.3.1.r 5-1.3.2.2 6-1.3.2 9-1.3.2.1.1.1.1 10-1.3.2.1 11-1.3.2.1.2.1.1 12-1.3.2.1.1 12-1.3.2.1.2 16-1.3 18-1.3.4 19-1.3.4 21-1.3.4 21-1.3.5.r 22-1.3.5.3.1.1.1 24-1.3.5.3 25-1.3.5.1.1.1.1 26-1.3.5 27-1.3.5.2.1.1.1 28-1.3.5.1 28-1.3.5.2 31-1.4.1 32-1.4.1.1 (e2 / evaluate-01~e.2 :li~e.1 -1~e.1 :ARG0 (w2 / we~e.0) :ARG1 (a / activity-06~e.16 :mode~e.3 interrogative~e.3 :ARG0 (i / inhibit-01~e.6 :ARG1 (a2 / and~e.10 :op1 (p / pathway~e.12 :name (n2 / name :op1 "PKA"~e.9)) :op2 (p2 / pathway~e.12 :name (n3 / name :op1 "MEK"~e.11))) :ARG1-of (c / combine-01~e.5)) :ARG1 (c2 / counter-01 :ARG1 (t / tumor)) :manner (i2 / in-vivo~e.18,19,21) :location~e.21 (a3 / and~e.26 :op1 (x / xenograft~e.28 :source (c3 / cell-line :name (n / name :op1 "HCT15"~e.25))) :op2 (x2 / xenograft~e.28 :source (c4 / cell-line :name (n4 / name :op1 "H460"~e.27))) :ARG0-of (r / resist-01~e.24 :ARG1 (s / small-molecule :name (n6 / name :op1 "selumetinib"~e.22))))) :ARG1-of (d / describe-01 :ARG0 (f2 / figure~e.31 :mod 7~e.32))) # ::id a_pmid_2256_9000.191 ::amr-annotator SDL-AMR-09 ::preferred # ::tok At day 50 from cancer cell injection , the mean tumour volume in mice bearing HCT15 and H460 tumour xenografts and treated with selumetinib , 25 mg kg @ −1 , and 8 @-@ Cl @-@ cAMP , 0.5 mg kg @ −1 , were 20 % and 16 % , respectively , as compared with control untreated mice . # ::alignments 1-1.3.2.2 2-1.3.2.1 3-1.1.3.1.1.1.1.r 3-1.1.3.1.1.2.1.r 4-1.3.1.1.1.2.1 5-1.3.1.1 6-1.3.1 9-1.1.1.2 10-1.1.1.1 11-1.1.1 13-1.1.3 14-1.1.3.1 15-1.1.3.1.1.1.1.1.1.1 16-1.1.3.1.1 17-1.1.3.1.1.2.1.1.1.1 18-1.1.3.1.1.1.1 18-1.1.3.1.1.2.1 19-1.1.3.1.1.1 19-1.1.3.1.1.2 20-1.1.3.1.1 21-1.1.3.2 22-1.1.3.2.1.r 23-1.1.3.2.1.1.1 25-1.1.3.2.1.2.1 26-1.1.3.2.1.2.2 27-1.1.3.2.1.2.2 32-1.1.3.1.1 33-1.2.3.2.1.1.1 35-1.2.3.2.1.1.1 37-1.2.3.2.1.1.1 39-1.2.3.2.1.2.1 40-1.2.3.2.1.2.2 41-1.2.3.2.1.2.2 47-1.1.2.1 48-1.1.2 49-1 50-1.2.2.1 51-1.2.2 55-1.3.r 56-1.4.r 58-1.4.1 59-1.2.3.2 59-1.4.2 59-1.4.2.1 59-1.4.2.1.r 60-1.2.3 60-1.4 (a / and~e.49 :op1 (e / equal-01 :ARG1 (v / volume~e.11 :mod (t / tumor~e.10) :mod (m / mean~e.9)) :ARG2 (p / percentage-entity~e.48 :value 20~e.47) :location (m2 / mouse~e.13 :ARG0-of (b / bear-01~e.14 :ARG1 (a2 / and~e.16,20,32 :op1 (x / xenograft~e.19 :source~e.3 (t2 / tumor~e.18 :mod (c6 / cell-line :name (n / name :op1 "HCT15"~e.15)))) :op2 (x2 / xenograft~e.19 :source~e.3 (t3 / tumor~e.18 :mod (c4 / cell-line :name (n2 / name :op1 "H460"~e.17)))))) :ARG1-of (t4 / treat-04~e.21 :ARG2~e.22 (s / small-molecule :name (n4 / name :op1 "selumetinib"~e.23) :mod (c / concentration-quantity :quant 25~e.25 :unit (m3 / milligram-per-kilogram~e.26,27)))))) :op2 (e2 / equal-01 :ARG1 v :ARG2 (p2 / percentage-entity~e.51 :value 16~e.50) :location (m4 / mouse~e.60 :ARG0-of b :ARG1-of (t5 / treat-04~e.59 :ARG2 (s2 / small-molecule :name (n5 / name :op1 "8-Cl-cAMP"~e.33,35,37) :mod (c2 / concentration-quantity :quant 0.5~e.39 :unit (m5 / milligram-per-kilogram~e.40,41)))))) :time~e.55 (a3 / after :op1 (i / inject-01~e.6 :ARG1 (c3 / cell~e.5 :mod (d / disease :wiki "Cancer" :name (n3 / name :op1 "cancer"~e.4)))) :quant (t6 / temporal-quantity :quant 50~e.2 :unit (d2 / day~e.1))) :compared-to~e.56 (m6 / mouse~e.60 :ARG0-of (c5 / control-01~e.58) :ARG1-of (t7 / treat-04~e.59 :polarity~e.59 -~e.59))) # ::id a_pmid_2458_8908.10 ::amr-annotator SDL-AMR-09 ::preferred # ::tok In a cohort of 144 metastatic melanoma patients we found that patients with N @-@ RAS mutant melanoma had a worse prognosis . # ::alignments 2-1.2.1.3.1 3-1.2.1.3.1.1.r 4-1.2.1.3.1.1.1 5-1.2.1.3.1.1.2.1.2 6-1.2.1.1.1.1.1 6-1.2.1.3.1.1.2.1.1.1 7-1.2.1.2.1 8-1.1 9-1 11-1.2.1.2.1 11-1.2.1.3.1.1.3.1 13-1.2.1.1.1.2.1.1 15-1.2.1.1.1.2.1.1 16-1.2.1.1.1.2 16-1.2.1.1.1.2.2 16-1.2.1.1.1.2.2.r 17-1.2.1.1.1.1.1 17-1.2.1.3.1.1.2.1.1.1 18-1.2 20-1.2.2.1 20-1.2.2.1.1 20-1.2.2.1.1.r 21-1.2.2 (f / find-01~e.9 :ARG0 (w / we~e.8) :ARG1 (h / have-03~e.18 :ARG0 (p / person :ARG0-of (h2 / have-03 :ARG1 (m5 / medical-condition :name (n / name :op1 "melanoma"~e.6,17) :mod (e / enzyme~e.16 :name (n2 / name :op1 "N-RAS"~e.13,15) :ARG2-of~e.16 (m / mutate-01~e.16)))) :ARG0-of (h4 / have-rel-role-91 :ARG2 (p4 / patient~e.7,11)) :ARG1-of (i / include-91 :ARG2 (c / cohort~e.2 :consist-of~e.3 (p3 / person :quant 144~e.4 :ARG0-of (h3 / have-03 :ARG1 (m4 / medical-condition :name (n3 / name :op1 "melanoma"~e.6,17) :ARG1-of (m3 / metastasize-101~e.5))) :ARG0-of (h5 / have-rel-role-91 :ARG2 (p5 / patient~e.11)))))) :ARG1 (p2 / prognosis~e.21 :ARG1-of (b / bad-07~e.20 :degree~e.20 (m2 / more~e.20))))) # ::id a_pmid_2458_8908.11 ::amr-annotator SDL-AMR-09 ::preferred # ::tok These patients were more likely to have brain metastases at the time of presentation with metastatic disease than their N @-@ RAS @-@ wild @-@ type counterparts . # ::alignments 0-1.1.1.1 1-1.1.1.3.1 3-1.2 4-1 6-1.1 6-1.1.1.3 7-1.1.2.1 8-1.1.2 11-1.1.3.r 13-1.1.3 14-1.1.3.2.r 15-1.1.3.2.1 16-1.1.3.2 17-1.1.3.1 18-1.1.3.1 19-1.1.3.1 20-1.1.3.1 21-1.1.3.1 22-1.1.3.1 23-1.1.3.1 24-1.1.3.1 25-1.1.3.1 26-1.1.3.1 (l / likely-01~e.4 :ARG1 (h / have-03~e.6 :ARG0 (p / person :mod (t / this~e.0) :compared-to (c / counterpart :mod (e / enzyme :name (n / name :op1 "N-RAS") :mod (w / wild-type))) :ARG0-of (h2 / have-rel-role-91~e.6 :ARG2 (p3 / patient~e.1))) :ARG1 (m / metastasize-101~e.8 :ARG2 (b / brain~e.7)) :time~e.11 (p2 / present-102~e.13 :ARG0 p~e.17,18,19,20,21,22,23,24,25,26 :ARG1~e.14 (d / disease~e.16 :ARG1-of (m2 / metastasize-101~e.15)))) :degree (m3 / more~e.3)) # ::id a_pmid_2458_8908.12 ::amr-annotator SDL-AMR-09 ::preferred # ::tok All N @-@ RAS mutant melanoma cultures tested in our study ( n = 7 ) were sensitive to MEK inhibition162 . # ::alignments 0-1.1.4 1-1.1.2.2.1.1 3-1.1.2.2.1.1 4-1.1.2.2 4-1.1.2.2.2 4-1.1.2.2.2.r 5-1.1.2.1.1 6-1.1 7-1.1.3 8-1.1.3.1.r 9-1.1.3.1.1 9-1.1.3.1.1.r 10-1.1.3.1 14-1.1.1 17-1 19-1.2.1.1.1 (s / sensitive-03~e.17 :ARG0 (c / culture-01~e.6 :quant 7~e.14 :ARG1 (m2 / medical-condition :name (n3 / name :op1 "melanoma"~e.5) :mod (e / enzyme~e.4 :name (n4 / name :op1 "N-RAS"~e.1,3) :ARG2-of~e.4 (m / mutate-01~e.4))) :ARG1-of (t / test-01~e.7 :medium~e.8 (s2 / study-01~e.10 :ARG0~e.9 (w / we~e.9))) :mod (a / all~e.0)) :ARG1 (i / inhibit-01 :ARG0 (s3 / small-molecule :name (n2 / name :op1 "MEK"~e.19 :op2 162)))) # ::id a_pmid_2458_8908.13 ::amr-annotator SDL-AMR-09 ::preferred # ::tok Exposure to MEK162 reduced ERK1 @/@ 2 phosphorylation , and induced apoptosis . # ::alignments 0-1.1.1 1-1.1.1.1.r 2-1.1.1.1.1.1 3-1.1 4-1.1.2.1.1.1 6-1.1.2.1.1.1 7-1.1.2 9-1 10-1.2 11-1.2.2 (a / and~e.9 :op1 (r2 / reduce-01~e.3 :ARG0 (e2 / expose-01~e.0 :ARG2~e.1 (s2 / small-molecule :name (n3 / name :op1 "MEK162"~e.2))) :ARG1 (p / phosphorylate-01~e.7 :ARG1 (e4 / enzyme :name (n4 / name :op1 "ERK1/2"~e.4,6)))) :op2 (i2 / induce-01~e.10 :ARG0 e2 :ARG2 (a3 / apoptosis~e.11))) # ::id a_pmid_2458_8908.14 ::amr-annotator SDL-AMR-09 ::preferred # ::tok Clonogenic survival was significantly reduced in sensitive melanoma cell cultures . # ::alignments 1-1.1 3-1.2 4-1 5-1.3.r 6-1.3.1.1 7-1.3.1.2.1.1 8-1.3.1 9-1.3 (r / reduce-01~e.4 :ARG1 (s2 / survive-01~e.1 :mod (c / clonogen)) :ARG2 (s / significant-02~e.3) :location~e.5 (c2 / culture-01~e.9 :ARG1 (c3 / cell~e.8 :ARG0-of (s3 / sensitive-03~e.6) :mod (m / medical-condition :name (n / name :op1 "melanoma"~e.7))))) # ::id a_pmid_2458_8908.58 ::amr-annotator SDL-AMR-09 ::preferred # ::tok Clinical profiles of patients whose tumors harbor N @-@ RAS and B @-@ RAF mutations # ::alignments 0-1.2 1-1 2-1.1.r 3-1.1.2.1 5-1.1.1.1 6-1.1.1.1.1 7-1.1.1.1.1.1.1.1.1.1 9-1.1.1.1.1.1.1.1.1.1 10-1.1.1.1.1.1.1 11-1.1.1.1.1.1.1.2.1.1 13-1.1.1.1.1.1.1.2.1.1 14-1.1.1.1.1.1 (p / profile-01~e.1 :ARG1~e.2 (p2 / person :ARG0-of (h2 / have-03 :ARG1 (t / tumor~e.5 :ARG0-of (h / harbor-01~e.6 :ARG1 (m / mutate-01~e.14 :ARG1 (a / and~e.10 :op1 (e / enzyme :name (n3 / name :op1 "N-RAS"~e.7,9)) :op2 (e2 / enzyme :name (n4 / name :op1 "B-RAF"~e.11,13))))))) :ARG0-of (h3 / have-rel-role-91 :ARG2 (p3 / patient~e.3))) :mod (c / clinical~e.0)) # ::id a_pmid_2458_8908.59 ::amr-annotator SDL-AMR-09 ::preferred # ::tok Characteristics of our cohort of 144 patients with stage IV melanoma are shown in Table 1 @ . # ::alignments 0-1.2 1-1.2.1.r 2-1.2.1.4.1.1 2-1.2.1.4.1.1.r 3-1.2.1.4.1 5-1.2.1.1 6-1.2.1.3.1 8-1.2.1.2.1 8-1.2.1.2.1.2 8-1.2.1.2.1.2.r 9-1.2.1.2.1.2.1 10-1.2.1.2.1.1.1 12-1 13-1.1.r 14-1.1 16-1.1.1 (s / show-01~e.12 :ARG0~e.13 (t / table~e.14 :mod 1~e.16) :ARG1 (c / characteristic-02~e.0 :ARG1~e.1 (p / person :quant 144~e.5 :ARG0-of (h / have-03 :ARG1 (m / medical-condition~e.8 :name (n / name :op1 "melanoma"~e.10) :ARG1-of~e.8 (s2 / stage-02~e.8 :ARG2 "IV"~e.9))) :ARG0-of (h2 / have-rel-role-91 :ARG2 (p2 / patient~e.6)) :ARG1-of (i / include-91 :ARG2 (c2 / cohort~e.3 :poss~e.2 (w / we~e.2)))))) # ::id a_pmid_2458_8908.60 ::amr-annotator SDL-AMR-09 ::preferred # ::tok Mutations were found in B @-@ RAF in 43.7 % , N @-@ RAS in 27.7 % , and 28.4 % were wild type ( WT ) for both . # ::alignments 0-1.1 2-1 3-1.1.1.r 4-1.1.1.1.1.1 6-1.1.1.1.1.1 8-1.1.1.1.2.1 9-1.1.1.1.2 11-1.1.1.2.1.1 13-1.1.1.2.1.1 15-1.1.1.2.2.1 16-1.1.1.2.2 18-1.1.1 19-1.1.1.3.2.1 20-1.1.1.3.2 22-1.1.1.3.1.1 23-1.1.1.3.1.1 25-1.1.1.3.1.1 (f / find-01~e.2 :ARG1 (m / mutate-01~e.0 :ARG1~e.3 (a2 / and~e.18 :op1 (e / enzyme :name (n3 / name :op1 "B-RAF"~e.4,6) :quant (p4 / percentage-entity~e.9 :value 43.7~e.8)) :op2 (e2 / enzyme :name (n4 / name :op1 "N-RAS"~e.11,13) :quant (p5 / percentage-entity~e.16 :value 27.7~e.15)) :ARG2-of (i / include-91 :ARG1 (e3 / enzyme :mod (w2 / wild-type~e.22,23,25)) :ARG3 (p6 / percentage-entity~e.20 :value 28.4~e.19))))) # ::id a_pmid_2458_8908.61 ::amr-annotator SDL-AMR-09 ::preferred # ::tok The majority of B @-@ RAF mutations were represented by substitution of valine at position 600 to glutamic acid ( 74.6 % were B @-@ RAF @ V600E ) or to lysine ( 19 % were B @-@ RAF @ V600K ) . # ::alignments 1-1.1.2.1.2 3-1.1.2.1.1.1.1.1 5-1.1.2.1.1.1.1.1 6-1.1.2 6-1.1.2.1.1 6-1.1.2.2.1 6-1.2.2 6-1.2.2.2.1 8-1.1 8-1.2 10-1.1.1 10-1.2.1 11-1.1.1.2.r 12-1.1.1.2.2.1 15-1.1.1.2.1 16-1.1.1.1.r 17-1.1.1.1.1.1 18-1.1.1.1.1.2 20-1.1.2.2.2.1 21-1.1.2.2.2 23-1.1.2.2.1.2 24-1.1.2.2.1.2 25-1.1.2.2.1.2 27-1.1.2.2.1.1 30-1 31-1.2.1.1.r 32-1.2.1.1.1.1 34-1.2.2.2.2.1 35-1.2.2.2.2 37-1.2.2.2.1.2 38-1.2.2.2.1.2 39-1.2.2.2.1.2 41-1.2.2.2.1.1 (o2 / or~e.30 :op1 (r / represent-01~e.8 :ARG0 (s / substitute-01~e.10 :ARG1~e.16 (a / amino-acid :name (n2 / name :op1 "glutamic"~e.17 :op2 "acid"~e.18)) :ARG2~e.11 (a2 / amino-acid :mod 600~e.15 :name (n5 / name :op1 "valine"~e.12))) :ARG1 (m / mutate-01~e.6 :ARG1-of (i / include-91 :ARG2 (m5 / mutate-01~e.6 :ARG2 (e2 / enzyme :name (n / name :op1 "B-RAF"~e.3,5))) :ARG3 (m2 / majority~e.1)) :ARG2-of (i2 / include-91 :ARG1 (m3 / mutate-01~e.6 :value "V600E"~e.27 :ARG2 e2~e.23,24,25) :ARG3 (p / percentage-entity~e.21 :value 74.6~e.20)))) :op2 (r2 / represent-01~e.8 :ARG0 (s2 / substitute-01~e.10 :ARG1~e.31 (a4 / amino-acid :name (n7 / name :op1 "lysine"~e.32)) :ARG2 a2) :ARG1 (m6 / mutate-01~e.6 :ARG1-of i :ARG2-of (i3 / include-91 :ARG1 (m4 / mutate-01~e.6 :value "V600K"~e.41 :ARG2 e2~e.37,38,39) :ARG3 (p2 / percentage-entity~e.35 :value 19~e.34))))) # ::id a_pmid_2458_8908.62 ::amr-annotator SDL-AMR-09 ::preferred # ::tok Substitutions of glutamine 61 accounted for 95 % of N @-@ RAS mutations ( most frequently Q61R/K/L/H ) . # ::alignments 0-1.1 1-1.1.1.r 2-1.1.1.2.1 3-1.1.1.1 4-1 5-1.2.r 6-1.2.3.2.1 7-1.2.3.2 9-1.2.1.1.1 11-1.2.1.1.1 12-1.2 12-1.2.2.1.1 12-1.2.2.1.2 12-1.2.2.1.3 12-1.2.2.1.4 12-1.2.3.1 14-1.2.2.2.1 15-1.2.2.2 (a / account-01~e.4 :ARG0 (s / substitute-01~e.0 :ARG2~e.1 (a2 / amino-acid :mod 61~e.3 :name (n / name :op1 "glutamine"~e.2))) :ARG2~e.5 (m / mutate-01~e.12 :ARG1 (e / enzyme :name (n2 / name :op1 "N-RAS"~e.9,11)) :ARG2-of (i / include-91 :ARG1 (a3 / and :op1 (m2 / mutate-01~e.12 :value "Q61R") :op2 (m5 / mutate-01~e.12 :value "Q61K") :op3 (m6 / mutate-01~e.12 :value "Q61L") :op4 (m7 / mutate-01~e.12 :value "Q61H")) :ARG1-of (f / frequent-02~e.15 :degree (m3 / most~e.14))) :ARG1-of (i2 / include-91 :ARG2 (m4 / mutate-01~e.12 :ARG1 e) :ARG3 (p2 / percentage-entity~e.7 :value 95~e.6)))) # ::id a_pmid_2458_8908.63 ::amr-annotator SDL-AMR-09 ::preferred # ::tok The slightly higher percentage of N @-@ RAS mutant melanomas in our population than what is commonly reported may be a result of the relatively small sample size or a reflection of local demographics . # ::alignments 1-1.1.1.2.1.1.1 2-1.1.1.2.1 2-1.1.1.2.1.1 2-1.1.1.2.1.1.r 3-1.1.1.2 4-1.1.1.2.2.r 5-1.1.1.2.2.2.1.1 7-1.1.1.2.2.2.1.1 8-1.1.1.2.2.2 8-1.1.1.2.2.2.2 8-1.1.1.2.2.2.2.r 9-1.1.1.2.2.1.1 10-1.1.1.2.3.r 11-1.1.1.2.3.1 11-1.1.1.2.3.1.r 12-1.1.1.2.3 13-1.1.1.2.4.r 14-1.1.1.1.1 14-1.1.1.2.4 16-1.1.1.2.4.1.1 17-1.1.1.2.4.1 18-1 21-1.1.1 22-1.1.1.1.r 24-1.1.1.1.2.1 25-1.1.1.1.2 26-1.1.1.1.1.1 27-1.1.1.1 28-1.1 30-1.1.2 31-1.1.2.2.r 32-1.1.2.2.1 33-1.1.2.2 (p / possible-01~e.18 :ARG1 (o2 / or~e.28 :op1 (r / result-01~e.21 :ARG1~e.22 (s2 / size-01~e.27 :ARG1 (t2 / thing~e.14 :ARG1-of (s3 / sample-01~e.26)) :ARG2 (s4 / small~e.25 :ARG2-of (r4 / relative-05~e.24))) :ARG2 (p2 / percentage~e.3 :ARG1-of (h / high-02~e.2 :degree~e.2 (m2 / more~e.2 :degree (s / slight~e.1))) :quant-of~e.4 (m / medical-condition :name (n / name :op1 "melanoma"~e.9) :mod (e / enzyme~e.8 :name (n2 / name :op1 "N-RAS"~e.5,7) :ARG2-of~e.8 (m3 / mutate-01~e.8))) :location~e.10 (p3 / population~e.12 :poss~e.11 (w / we~e.11)) :compared-to~e.13 (t / thing~e.14 :ARG1-of (r2 / report-01~e.17 :mod (c / common~e.16))))) :op2 (r3 / reflect-01~e.30 :ARG1 p2 :ARG2~e.31 (d / demographics~e.33 :ARG1-of (l / local-02~e.32))))) # ::id a_pmid_2458_8908.64 ::amr-annotator SDL-AMR-09 ::preferred # ::tok Clinical and pathological characteristics # ::alignments 0-1.1 2-1.2 3-1 (c / characteristic-02~e.3 :mod (c2 / clinical~e.0) :mod (p / pathology~e.2)) # ::id a_pmid_2458_8908.65 ::amr-annotator SDL-AMR-09 ::preferred # ::tok Patients with B @-@ RAF mutations tended to be younger ; median age at initial diagnosis of melanoma was 57.6 in patients with B @-@ RAF mutations , 68.2 in patients with N @-@ RAS mutations and 66.3 years in patients wild @-@ type for both ( P < 0.0001 ) . # ::alignments 0-1.1.1.1.1 1-1.1.1.1.1.1.r 2-1.1.1.1.1.1.1.1 4-1.1.1.1.1.1.1.1 5-1.1.1.1.1.1 5-1.1.1.1.1.1.2 5-1.1.1.1.1.1.2.r 6-1.1 9-1.1.2.1 9-1.1.2.1.1 9-1.1.2.1.1.r 11-1.2.4 12-1.2.1 12-1.2.2 12-1.2.3 14-1.2.5.2 17-1.2.5.1.1.1 19-1.2.1.2.1 20-1.2.1.1.r 21-1.2.1.1 22-1.2.1.1 23-1.2.1.1 24-1.2.1.1 25-1.2.1.1 26-1.2.1.1 28-1.2.2.2.1 29-1.2.2.1.r 30-1.2.2.1.1.1 31-1.2.2.1.1.1.1.r 32-1.2.2.1.1.1.1.1.1 34-1.2.2.1.1.1.1.1.1 35-1.2.2.1.1.1.1 35-1.2.2.1.1.1.1.2 35-1.2.2.1.1.1.1.2.r 36-1.2 37-1.2.3.2.1 38-1.2.1.2.2 38-1.2.2.2.2 38-1.2.3.2.2 39-1.2.3.1.r 40-1.2.3.1.1.1 41-1.2.3.1.1.1.3 43-1.2.3.1.1.1.3 48-1.2.6.1 49-1.2.6.1.1 (a3 / and :op1 (t / tend-02~e.6 :ARG1 (p / person :ARG0-of (h4 / have-rel-role-91 :ARG2 (p3 / patient~e.0 :mod~e.1 (e3 / enzyme~e.5 :name (n / name :op1 "B-RAF"~e.2,4) :ARG2-of~e.5 (m / mutate-01~e.5))))) :ARG2 (p2 / person :mod (y / young~e.9 :degree~e.9 (m3 / more~e.9)))) :op2 (a / and~e.36 :op1 (a4 / age-01~e.12 :ARG1~e.20 p~e.21,22,23,24,25,26 :ARG2 (t2 / temporal-quantity :quant 57.6~e.19 :unit (y2 / year~e.38))) :op2 (a5 / age-01~e.12 :ARG1~e.29 (p4 / person :ARG0-of (h6 / have-rel-role-91 :ARG2 (p7 / patient~e.30 :mod~e.31 (e / enzyme~e.35 :name (n4 / name :op1 "N-RAS"~e.32,34) :ARG2-of~e.35 (m6 / mutate-01~e.35))))) :ARG2 (t3 / temporal-quantity :quant 68.2~e.28 :unit (y3 / year~e.38))) :op3 (a6 / age-01~e.12 :ARG1~e.39 (p5 / person :ARG0-of (h7 / have-rel-role-91 :ARG2 (p8 / patient~e.40 :mod e :mod e3 :mod (w / wild-type~e.41,43)))) :ARG2 (t4 / temporal-quantity :quant 66.3~e.37 :unit (y4 / year~e.38))) :mod (m4 / median~e.11) :time (d / diagnose-01 :ARG2 (m2 / medical-condition :name (n2 / name :op1 "melanoma"~e.17)) :mod (i / initial~e.14)) :ARG1-of (p6 / possible-01 :quant (l / less-than~e.48 :value 0.0001~e.49)))) # ::id a_pmid_2458_8908.66 ::amr-annotator SDL-AMR-09 ::preferred # ::tok Our cohort included 22 patients who received either dabrafenib ( N = 1 ) , vemurafenib ( N = 19 ) , a pan @-@ RAF inhibitor ( N = 1 ) or a pan @-@ RAF inhibitor and vemurafenib ( N = 1 ) . # ::alignments 0-1.2.1 0-1.2.1.r 1-1.2 2-1 2-1.1.2.1.1 3-1.1.1 4-1.1.2.1 4-1.1.2.1.1.1.1.2.1 6-1.1.2.1.1.1.1.3 6-1.1.2.1.1.1.2.3 6-1.1.2.1.1.1.3.3 6-1.1.2.1.1.1.4.3 8-1.1.2.1.1.1.1.3.1.1.1 12-1.1.2.1.1.1.1.1 12-1.1.2.1.1.1.3.1 12-1.1.2.1.1.1.4.1 15-1.1.2.1.1.1.2.3.1.1.1 19-1.1.2.1.1.1.2.1 23-1.1.2.1.1.1.3.3.1.1.1.1.1 25-1.1.2.1.1.1.3.3.1.1.1.1.1 26-1.1.2.1.1.1.3.3.1 26-1.1.2.1.1.1.3.3.1.1 26-1.1.2.1.1.1.3.3.1.1.r 30-1.1.2.1.1.1.3.1 34-1.1.2.1.1.1.3.3.1.1.1.1.1 36-1.1.2.1.1.1.3.3.1.1.1.1.1 37-1.1.2.1.1.1.3.3.1 37-1.1.2.1.1.1.3.3.1.1 37-1.1.2.1.1.1.3.3.1.1.r 38-1.1.2.1.1.1 38-1.1.2.1.1.1.4.3.1 39-1.1.2.1.1.1.2.3.1.1.1 43-1.1.2.1.1.1.1.1 43-1.1.2.1.1.1.3.1 43-1.1.2.1.1.1.4.1 (i2 / include-01~e.2 :ARG1 (p / person :quant 22~e.3 :ARG0-of (h / have-rel-role-91 :ARG2 (p6 / patient~e.4 :ARG2-of (i7 / include-91~e.2 :ARG1 (a2 / and~e.38 :op1 (p2 / person :quant 1~e.12,43 :ARG0-of (h2 / have-rel-role-91 :ARG2 (p7 / patient~e.4)) :ARG0-of (r2 / receive-01~e.6 :ARG1 (s / small-molecule :name (n / name :op1 "dabrafenib"~e.8)))) :op2 (p3 / person :quant 19~e.19 :ARG0-of h2 :ARG0-of (r3 / receive-01~e.6 :ARG1 (s2 / small-molecule :name (n2 / name :op1 "vemurafenib"~e.15,39)))) :op3 (p4 / person :quant 1~e.12,30,43 :ARG0-of h2 :ARG0-of (r4 / receive-01~e.6 :ARG1 (m / molecular-physical-entity~e.26,37 :ARG0-of~e.26,37 (i / inhibit-01~e.26,37 :ARG1 (e / enzyme :name (n4 / name :op1 "pan-RAF"~e.23,25,34,36)))))) :op4 (p5 / person :quant 1~e.12,43 :ARG0-of h2 :ARG0-of (r5 / receive-01~e.6 :ARG2 (a3 / and~e.38 :op1 m :op2 s2)))))))) :ARG2 (c / cohort~e.1 :poss~e.0 (w / we~e.0))) # ::id a_pmid_2458_8908.67 ::amr-annotator SDL-AMR-09 ::preferred # ::tok Patients with N @-@ RAS mutated melanomas appear to have an increased rate of skin and soft tissue involvement ( 70 %) compared to B @-@ RAF mutated counterparts and WT patients ( 37 % and 41 % respectively , P = 0.0025 ) . # ::alignments 0-1.1.3.1.2.2.1.1 2-1.1.2.1.1.1.2.1.1 4-1.1.2.1.1.1.2.1.1 5-1.1.2.1.1.1.2.2 6-1.1.2.1.1.1.1.1 7-1 9-1.1.2 9-1.1.2.1 9-1.1.2.1.1 9-1.1.2.1.1.r 9-1.1.2.1.2 9-1.1.2.1.2.r 9-1.1.2.1.r 11-1.1 12-1.1.1 12-1.1.3.1.1 12-1.1.3.1.2 13-1.1.1.1.r 14-1.1.1.1.1.1 15-1.1.1.1.1 16-1.1.1.1.1.2.1 17-1.1.1.1.1.2 18-1.1.1.1 20-1.1.1.2.1 22-1.1.3 24-1.1.3.1.1.2.1.1.1 24-1.1.3.1.2.2.1.1.1.1.1 26-1.1.3.1.1.2.1.1.1 26-1.1.3.1.2.2.1.1.1.1.1 27-1.1.3.1.1.2.1.2 28-1.1.3.1.1.2 29-1.1.3.1 30-1.1.3.1.2.2.1.1.1.2 31-1.1.2.1.2.1 33-1.1.3.1.1.3.1 34-1.1.1.2 34-1.1.3.1.1.3 35-1.1.3.1 36-1.1.3.1.2.3.1 37-1.1.3.1.2.3 38-1.1.3.1.3 42-1.2.1 (a / appear-01~e.7 :ARG1 (i / increase-01~e.11 :ARG1 (r / rate~e.12 :degree-of~e.13 (i2 / involve-01~e.18 :ARG1 (a2 / and~e.15 :op1 (s / skin~e.14) :op2 (t / tissue~e.17 :ARG1-of (s2 / soft-02~e.16)))) :mod (p6 / percentage-entity~e.34 :value 70~e.20)) :ARG1-of (h / have-03~e.9 :ARG0~e.9 (p2 / person~e.9 :ARG0-of~e.9 (h2 / have-03~e.9 :ARG1 (m2 / medical-condition :name (n3 / name :op1 "melanoma"~e.6) :mod (e / enzyme :name (n4 / name :op1 "N-RAS"~e.2,4) :ARG2-of (m / mutate-01~e.5)))) :ARG0-of~e.9 (h3 / have-rel-role-91~e.9 :ARG2 (p3 / patient~e.31)))) :ARG1-of (c / compare-01~e.22 :ARG2 (a3 / and~e.29,35 :op1 (r3 / rate~e.12 :degree-of i2 :mod (c3 / counterpart~e.28 :mod (e2 / enzyme :name (n5 / name :op1 "B-RAF"~e.24,26) :ARG2-of (m3 / mutate-01~e.27))) :mod (p7 / percentage-entity~e.34 :value 37~e.33)) :op2 (r4 / rate~e.12 :degree-of i2 :mod (p4 / person :ARG0-of (h4 / have-rel-role-91 :ARG2 (p5 / patient~e.0 :mod (e3 / enzyme :name (n / name :op1 "B-RAF"~e.24,26) :mod (w2 / wild-type~e.30))))) :mod (p8 / percentage-entity~e.37 :value 41~e.36)) :mod (r2 / respective~e.38)))) :ARG1-of (p / possible-01 :value 0.0025~e.42)) # ::id a_pmid_2458_8908.68 ::amr-annotator SDL-AMR-09 ::preferred # ::tok The rate of lymph node metastasis was also noted to be higher in patients with N @-@ RAS mutations ( 75 %) compared to B @-@ RAF mutant and WT patients ( 46 % and 61 % , respectively ) ( P = 0.01 ) . # ::alignments 1-1.1 1-1.1.5.1.1 1-1.1.5.1.2 2-1.1.3.r 3-1.1.3.1.1 4-1.1.3.1 5-1.1.3 7-1.3 8-1 11-1.1.2 11-1.1.2.1 11-1.1.2.1.r 12-1.1.4.r 13-1.1.4.1.1 14-1.1.4.1.1.1.r 15-1.1.4.1.1.1.1.1 17-1.1.4.1.1.1.1.1 18-1.1.4.1.1.1 18-1.1.4.1.1.1.2 18-1.1.4.1.1.1.2.r 20-1.1.1.1 22-1.1.5 23-1.1.5.1.1.1.r 23-1.1.5.1.r 24-1.1.5.1.1.1.2.1.1 26-1.1.5.1.1.1.2.1.1 27-1.1.5.1.1.1.2 27-1.1.5.1.1.1.2.2 27-1.1.5.1.1.1.2.2.r 28-1.1.5.1 29-1.1.5.1.2.1.2 30-1.1.5.1.1.1.1.1 32-1.1.5.1.1.2.1 33-1.1.1 33-1.1.5.1.1.2 34-1.1.5.1 35-1.1.5.1.2.2.1 36-1.1.5.1.2.2 43-1.2.1 (n3 / note-02~e.8 :ARG1 (r / rate~e.1 :mod (p4 / percentage-entity~e.33 :value 75~e.20) :ARG1-of (h2 / high-02~e.11 :degree~e.11 (m3 / more~e.11)) :mod~e.2 (m / metastasize-101~e.5 :ARG2 (n4 / node~e.4 :mod (l / lymph~e.3))) :condition~e.12 (p / person :ARG0-of (h / have-rel-role-91 :ARG2 (p2 / patient~e.13 :mod~e.14 (e / enzyme~e.18 :name (n5 / name :op1 "N-RAS"~e.15,17) :ARG2-of~e.18 (m4 / mutate-01~e.18))))) :ARG1-of (c / compare-01~e.22 :ARG2~e.23 (a / and~e.28,34 :op1 (r3 / rate~e.1 :mod~e.23 (p3 / person :ARG0-of (h3 / have-rel-role-91 :ARG2 (p5 / patient~e.30)) :mod (e2 / enzyme~e.27 :name (n6 / name :op1 "B-RAF"~e.24,26) :ARG2-of~e.27 (m5 / mutate-01~e.27))) :mod (p7 / percentage-entity~e.33 :value 46~e.32)) :op2 (r2 / rate~e.1 :mod (p6 / person :ARG0-of h3 :mod (w / wild-type~e.29)) :mod (p8 / percentage-entity~e.36 :value 61~e.35))))) :ARG1-of (p9 / possible-01 :value 0.01~e.43) :mod (a2 / also~e.7)) # ::id a_pmid_2458_8908.69 ::amr-annotator SDL-AMR-09 ::preferred # ::tok No statistically significant difference was seen between the genotypes and other clinical characteristics , such as M stage and LDH levels.Seeing that B @-@ RAF inhibitors can affect survival in patients with B @-@ RAF mutant melanomas , this group of patients was removed from the survival analysis . # ::alignments 0-1.1.1.r 2-1.1.2.3 3-1.1.1 3-1.1.1.r 3-1.1.2 5-1.1 5-1.2 8-1.1.2.1 9-1.1.2.2.1 10-1.1.2.2.3 11-1.1.2.2.2 12-1.1.2.2 15-1.1.2.2.1.r 16-1.1.2.2.1.1.1.1 17-1.1.2.2.1.1.1.2 18-1.1.2.2.1 19-1.1.2.2.1.2.1.1.1 21-1.2.1.r 22-1.2.1.1.1.1.1.1.1 24-1.2.1.1.1.1.1.1.1 25-1.2.1.1.1 25-1.2.1.1.1.1 25-1.2.1.1.1.1.r 26-1.2.1 27-1.2.1.1 28-1.2.1.1.2 29-1.2.1.1.2.1.r 30-1.2.1.1.2.1.2.1 32-1.2.1.1.2.1.1.1.2.1.1 34-1.2.1.1.2.1.1.1.2.1.1 35-1.2.1.1.2.1.1.1.2 35-1.2.1.1.2.1.1.1.2.2 35-1.2.1.1.2.1.1.1.2.2.r 36-1.2.1.1.2.1.1.1.1.1 38-1.2.2.1.1.1 39-1.2.2.1.1 41-1.2.1.1.2.1.2.1 43-1.2.2.1 44-1.2.2.1.2.r 46-1.2.2.1.2.1 47-1.2.2.1.2 (m / multi-sentence :snt1 (s / see-01~e.5 :polarity~e.0,3 -~e.3 :ARG1 (d / differ-02~e.3 :ARG1 (g / genotype~e.8) :ARG2 (c / characteristic-02~e.12 :ARG2~e.15 (a / and~e.9,18 :op1 (e5 / event :name (n4 / name :op1 "M"~e.16 :op2 "stage"~e.17)) :op2 (l / level :quant-of (e3 / enzyme :name (n3 / name :op1 "LDH"~e.19))) :ARG1-of (e / exemplify-01)) :mod (c2 / clinic~e.11) :mod (o / other~e.10)) :ARG1-of (s2 / significant-02~e.2 :mod (s3 / statistic)))) :snt2 (s7 / see-01~e.5 :ARG1~e.21 (p / possible-01~e.26 :ARG1 (a2 / affect-01~e.27 :ARG0 (m3 / molecular-physical-entity~e.25 :ARG0-of~e.25 (i2 / inhibit-01~e.25 :ARG1 (e4 / enzyme :name (n / name :op1 "B-RAF"~e.22,24)))) :ARG1 (s5 / survive-01~e.28 :ARG0~e.29 (p2 / person :ARG0-of (h / have-03 :ARG1 (m2 / medical-condition :name (n5 / name :op1 "melanoma"~e.36) :mod (e2 / enzyme~e.35 :name (n2 / name :op1 "B-RAF"~e.32,34) :ARG2-of~e.35 (m4 / mutate-01~e.35)))) :ARG0-of (h2 / have-rel-role-91 :ARG2 (p3 / patient~e.30,41)))))) :ARG0-of (c4 / cause-01 :ARG1 (r / remove-01~e.43 :ARG1 (g2 / group~e.39 :mod (t / this~e.38) :ARG2-of (i / include-91 :ARG1 p2)) :ARG2~e.44 (a3 / analyze-01~e.47 :ARG1 (s6 / survive-01~e.46)))))) # ::id a_pmid_2458_8908.70 ::amr-annotator SDL-AMR-09 ::preferred # ::tok By Cox univariate analysis we found a trend towards shorter survival in the N @-@ RAS mutant population , compared to the B @-@ RAF and WT groups combined ( p = 0.12 ) . # ::alignments 1-1.4.1.1 2-1.4.2 3-1.4 4-1.1 5-1 7-1.2 9-1.2.1.2 9-1.2.1.2.1 9-1.2.1.2.1.r 10-1.2.1 11-1.2.1.1.r 13-1.2.1.1.1.1.1.1 15-1.2.1.1.1.1.1.1 16-1.2.1.1.1.1 16-1.2.1.1.1.1.2 16-1.2.1.1.1.1.2.r 17-1.2.1.1 19-1.2.1.3 20-1.2.1.3.1.r 22-1.2.1.3.1.1.1.1.1 24-1.2.1.3.1.1.1.1.1 25-1.2.1.3.1 26-1.2.1.3.1.2.1 27-1.2.1.3.1.1 27-1.2.1.3.1.2 28-1.2.1.3.1.3 32-1.3.1 (f / find-01~e.5 :ARG0 (w / we~e.4) :ARG1 (t / trend-01~e.7 :ARG2 (s / survive-01~e.10 :ARG1~e.11 (p / population~e.17 :ARG0-of (h / have-03 :ARG1 (e3 / enzyme~e.16 :name (n3 / name :op1 "N-RAS"~e.13,15) :ARG2-of~e.16 (m / mutate-01~e.16)))) :ARG1-of (s2 / short-07~e.9 :degree~e.9 (m2 / more~e.9)) :ARG1-of (c / compare-01~e.19 :ARG2~e.20 (a / and~e.25 :op1 (g / group~e.27 :mod (e4 / enzyme :name (n4 / name :op1 "B-RAF"~e.22,24))) :op2 (g2 / group~e.27 :mod (w2 / wild-type~e.26)) :ARG1-of (c3 / combine-01~e.28))))) :ARG1-of (p2 / possible-01 :value 0.12~e.32) :manner (a3 / analyze-01~e.3 :name (n / name :op1 "Cox"~e.1) :mod (u / univariate~e.2))) # ::id a_pmid_2458_8908.71 ::amr-annotator SDL-AMR-09 ::preferred # ::tok The median survival was 13 and 19.6 months , respectively . # ::alignments 1-1.1 2-1 4-1.2.1.1 5-1.2 6-1.2.2.1 7-1.2.1.2 9-1.2.3 (s / survive-01~e.2 :mod (m2 / median~e.1) :duration (a / and~e.5 :op1 (t / temporal-quantity :quant 13~e.4 :unit (m / month~e.7)) :op2 (t4 / temporal-quantity :quant 19.6~e.6 :unit m) :mod (r / respective~e.9))) # ::id a_pmid_2458_8908.72 ::amr-annotator SDL-AMR-09 ::preferred # ::tok Kaplan @-@ Meier survival curves are shown in Figure 1 @ a for the three groups of patients ( BRAF or NRAS mutant or WT for both ) and Figure 1 @ b for the two groups ( NRAS compared to BRAF mutant and WT combined ) to visually demonstrate the differences between the groups . # ::alignments 0-1.2.1.1 2-1.2.1.1 3-1.2.2 4-1.2 6-1 7-1.1.r 8-1.1.1 15-1.1.1.2.1 16-1.1.1.2 17-1.1.1.2.2.r 18-1.1.1.2.2.1.1 20-1.1.1.2.2.2.1.1.1 20-1.1.1.2.2.2.2.1.1 21-1.1.1.2.2.2 22-1.1.1.2.2.2.3.1.1 22-1.1.1.2.2.2.4.1.1 23-1.1.1.2.2.2.1 23-1.1.1.2.2.2.1.2 23-1.1.1.2.2.2.1.2.r 23-1.1.1.2.2.2.3 23-1.1.1.2.2.2.3.2 23-1.1.1.2.2.2.3.2.r 24-1.1.1.2.2.2 25-1.1.1.2.2.2.2.2 25-1.1.1.2.2.2.4.2 29-1.1 30-1.1.1 30-1.1.2 35-1.1.2.2.r 37-1.1.2.2.1 38-1.1.2.2 40-1.1.2.2.2.1.1.1 41-1.1.2.2.2 43-1.1.1.2.2.2.1.1.1 43-1.1.1.2.2.2.2.1.1 44-1.1.1.2.2.2.1 44-1.1.1.2.2.2.1.2 44-1.1.1.2.2.2.1.2.r 44-1.1.1.2.2.2.3 44-1.1.1.2.2.2.3.2 44-1.1.1.2.2.2.3.2.r 46-1.1.1.2.2.2.2.2 46-1.1.1.2.2.2.4.2 47-1.1.2.2.2.2 50-1.3.2 51-1.3 53-1.3.1 56-1.3.1.2 (s / show-01~e.6 :ARG0~e.7 (a / and~e.29 :op1 (f / figure~e.8,30 :mod "1a" :beneficiary (g / group-01~e.16 :quant 3~e.15 :ARG1~e.17 (p / person :ARG0-of (h2 / have-rel-role-91 :ARG2 (p2 / patient~e.18)) :mod (o2 / or~e.21,24 :op1 (e / enzyme~e.23,44 :name (n / name :op1 "BRAF"~e.20,43) :ARG2-of~e.23,44 (m4 / mutate-01~e.23,44)) :op2 (e3 / enzyme :name (n4 / name :op1 "BRAF"~e.20,43) :mod (w2 / wild-type~e.25,46)) :op3 (e2 / enzyme~e.23,44 :name (n2 / name :op1 "NRAS"~e.22) :ARG2-of~e.23,44 (m5 / mutate-01~e.23,44)) :op4 (e4 / enzyme :name (n5 / name :op1 "NRAS"~e.22) :mod (w3 / wild-type~e.25,46)))))) :op2 (f2 / figure~e.30 :mod "1b" :beneficiary~e.35 (g2 / group-01~e.38 :quant 2~e.37 :mod (c / compare-01~e.41 :ARG1 (e5 / enzyme :name (n3 / name :op1 "NRAS"~e.40)) :ARG2 (c2 / combine-01~e.47 :ARG1 e :ARG2 e3))))) :ARG1 (c4 / curve~e.4 :name (n6 / name :op1 "Kaplan-Meier"~e.0,2) :mod (s2 / survive-01~e.3)) :purpose (d / demonstrate-01~e.51 :ARG1 (d2 / differ-02~e.53 :ARG1 g :ARG2 g2~e.56) :mod (v / visual~e.50))) # ::id a_pmid_2458_8908.73 ::amr-annotator SDL-AMR-09 ::preferred # ::tok Interestingly , analysis of anatomic sites at the time of initial diagnosis of stage IV disease revealed a higher rate of brain involvement among B @-@ RAF ( 16 %) and N @-@ RAS ( 15 %) mutant melanoma patients , compared with patients with WT disease ( 2.5 %) ( P = 0.04 ) . # ::alignments 0-1.3 2-1.1 3-1.1.1.r 4-1.1.1.1 5-1.1.1 8-1.1.2.r 10-1.1.2.2 13-1.1.2.1.1 14-1.1.2.1.1.1 15-1.1.2.1 16-1 18-1.2.3 18-1.2.3.1 18-1.2.3.1.r 19-1.2.1 19-1.2.2 19-1.2.4.1 20-1.2.1.1.r 21-1.2.1.1.1 22-1.2.1.1 22-1.2.2.1 22-1.2.4.1.1 23-1.2.1.1.2.r 24-1.2.1.1.2.1.1.1.2.1.1 26-1.2.1.1.2.1.1.1.2.1.1 28-1.2.1.2.1 30-1.2 31-1.2.2.1.2.1.1.1.2.1.1 33-1.2.2.1.2.1.1.1.2.1.1 35-1.2.2.2.1 37-1.2.1.1.2.1.1.1.2 37-1.2.1.1.2.1.1.1.2.2 37-1.2.1.1.2.1.1.1.2.2.r 37-1.2.2.1.2.1.1.1.2 37-1.2.2.1.2.1.1.1.2.2 37-1.2.2.1.2.1.1.1.2.2.r 38-1.2.1.1.2.1.1.1.1.1 38-1.2.2.1.2.1.1.1.1.1 39-1.2.4.1.2.1.1 41-1.2.4 43-1.2.1.1.2.1.1 43-1.2.2.1.2.1.1 43-1.2.4.1.2.1.1 44-1.2.4.1.2.1.1.1.r 45-1.2.4.1.2.1.1.1.1 46-1.2.4.1.2.1.1.1 48-1.2.4.1.3.1 53-1.4.1 (r / reveal-01~e.16 :ARG0 (a / analyze-01~e.2 :ARG1~e.3 (s / site~e.5 :mod (a2 / anatomy~e.4)) :time~e.8 (d / diagnose-01 :ARG2 (d2 / disease~e.15 :ARG1-of (s2 / stage-02~e.13 :ARG2 "IV"~e.14)) :mod (i / initial~e.10))) :ARG1 (a4 / and~e.30 :op1 (r3 / rate~e.19 :mod~e.20 (i4 / involve-01~e.22 :ARG1 (b / brain~e.21) :prep-among~e.23 (p3 / person :ARG0-of (h / have-rel-role-91 :ARG2 (p8 / patient~e.43 :mod (m3 / medical-condition :name (n5 / name :op1 "melanoma"~e.38) :mod (e / enzyme~e.37 :name (n / name :op1 "B-RAF"~e.24,26) :ARG2-of~e.37 (m / mutate-01~e.37))))))) :mod (p4 / percentage-entity :value 16~e.28)) :op2 (r4 / rate~e.19 :mod (i5 / involve-01~e.22 :ARG1 b :prep-among (p9 / person :ARG0-of (h5 / have-rel-role-91 :ARG2 (p10 / patient~e.43 :mod (m5 / medical-condition :name (n2 / name :op1 "melanoma"~e.38) :mod (e2 / enzyme~e.37 :name (n6 / name :op1 "N-RAS"~e.31,33) :ARG2-of~e.37 (m4 / mutate-01~e.37))))))) :mod (p5 / percentage-entity :value 15~e.35)) :ARG1-of (h2 / high-02~e.18 :degree~e.18 (m2 / more~e.18)) :ARG1-of (c / compare-01~e.41 :ARG2 (r2 / rate~e.19 :mod (i3 / involve-01~e.22 :ARG1 b) :condition (p6 / person :ARG0-of (h3 / have-rel-role-91 :ARG2 (p7 / patient~e.39,43 :mod~e.44 (d6 / disease~e.46 :mod (w2 / wild-type~e.45))))) :mod (p / percentage-entity :value 2.5~e.48)))) :ARG2-of (i2 / interest-01~e.0) :ARG1-of (p2 / possible-01 :value 0.04~e.53)) # ::id a_pmid_2458_8908.74 ::amr-annotator SDL-AMR-09 ::preferred # ::tok With longitudinal follow @-@ up , however , the rate of development of brain metastases did not differ among the three groups , possibly because the WT groups lived longer and thus developed brain metastases over time . # ::alignments 1-1.2.1.1 2-1.2.1 4-1.2.1 6-1 9-1.1.3 10-1.1.3.1.r 11-1.1.3.1 12-1.1.3.1.1.r 13-1.1.3.1.1.1 14-1.1.3.1.1 16-1.1.1 16-1.1.1.r 17-1.1 20-1.1.2.1 21-1.1.2 23-1.1.4.2 24-1.1.4 26-1.1.4.1.1.1.1 27-1.1.4.1.1.1 28-1.1.4.1.1 29-1.1.4.1.1.2 29-1.1.4.1.1.2.1 29-1.1.4.1.1.2.1.r 30-1.1.4.1 31-1.1.4.1.2.3 32-1.1.4.1.2 33-1.1.4.1.2.1 34-1.1.4.1.2.1 35-1.1.4.1.2.2 36-1.1.4.1.2.2.1 (c3 / contrast-01~e.6 :ARG2 (d / differ-02~e.17 :polarity~e.16 -~e.16 :ARG1 (g / group~e.21 :quant 3~e.20) :ARG3 (r / rate~e.9 :mod~e.10 (d2 / develop-01~e.11 :ARG2~e.12 (m / metastasize-101~e.14 :ARG2 (b / brain~e.13)))) :ARG1-of (c2 / cause-01~e.24 :ARG0 (a2 / and~e.30 :op1 (l2 / live-01~e.28 :ARG0 (g2 / group~e.27 :mod (w / wild-type~e.26)) :ARG1-of (l3 / long-03~e.29 :degree~e.29 (m2 / more~e.29))) :op2 (d3 / develop-01~e.32 :ARG2 m~e.33,34 :duration (o / over~e.35 :mod (t / time~e.36)) :ARG1-of (c4 / cause-01~e.31 :ARG0 l2))) :ARG1-of (p / possible-01~e.23))) :ARG1-of (c / condition-01 :ARG2 (f / follow-up-03~e.2,4 :mod (l / longitude~e.1)))) # ::id a_pmid_2458_8908.75 ::amr-annotator SDL-AMR-09 ::preferred # ::tok In vitro activity of B @-@ RAF and MEK inhibitors in a large panel of melanoma cultures # ::alignments 1-1.2 2-1.2 4-1 5-1.1.r 6-1.1.1.1.1.1.1 8-1.1.1.1.1.1.1 9-1.1 10-1.1.2.1.1.1.1 11-1.1.1 11-1.1.1.1 11-1.1.1.1.r 11-1.1.2 11-1.1.2.1 11-1.1.2.1.r 12-1.2 12-1.3.r 14-1.3.1 15-1.3 16-1.3.2.r 17-1.3.2.1.1.1 18-1.3.2 (a / activity-06~e.4 :ARG0~e.5 (a2 / and~e.9 :op1 (m2 / molecular-physical-entity~e.11 :ARG0-of~e.11 (i / inhibit-01~e.11 :ARG1 (e / enzyme :name (n / name :op1 "B-RAF"~e.6,8)))) :op2 (m3 / molecular-physical-entity~e.11 :ARG0-of~e.11 (i3 / inhibit-01~e.11 :ARG1 (p2 / protein-family :name (n2 / name :op1 "MEK"~e.10))))) :manner (i2 / in-vitro~e.1,2,12) :location~e.12 (p / panel~e.15 :mod (l / large~e.14) :consist-of~e.16 (c / culture~e.18 :mod (m / medical-condition :name (n3 / name :op1 "melanoma"~e.17))))) # ::id a_pmid_2458_8908.76 ::amr-annotator SDL-AMR-09 ::preferred # ::tok To investigate the effect of B @-@ RAF and MEK inhibition in melanoma cultures , we used RAF265 ( a pan @-@ RAF inhibitor ) , MEK162 ( a MEK1 @/@ 2 inhibitor ) and the MEK inhibitor trametinib . # ::alignments 1-1.3 3-1.3.2 5-1.3.2.1.1.1.2.1 7-1.3.2.1.1.1.2.1 8-1.2 8-1.3.2.1.1 9-1.3.2.1.1.2 10-1.2.3 10-1.2.3.3 10-1.2.3.3.r 10-1.3.2.1 11-1.3.2.2.r 12-1.3.2.2.1.2.1 13-1.3.2.2 15-1.1 16-1 17-1.2.1.2.1 20-1.2.1.3.1.2.1 22-1.2.1.3.1.2.1 23-1.2.1 23-1.2.1.3 23-1.2.1.3.r 23-1.2.3 23-1.2.3.3 23-1.2.3.3.r 26-1.2.2.2.1 29-1.2.2.3.1.2.1 31-1.2.2.3.1.2.1 32-1.2.1 32-1.2.1.3 32-1.2.1.3.r 32-1.2.2 32-1.2.2.3 32-1.2.2.3.r 32-1.2.3 32-1.2.3.3 32-1.2.3.3.r 36-1.2.3.3.1.2.1 37-1.2.3 37-1.2.3.3 37-1.2.3.3.r 38-1.2.3.2.1 (u / use-01~e.16 :ARG0 (w / we~e.15) :ARG1 (a / and~e.8 :op1 (s2 / small-molecule~e.23,32 :wiki - :name (n4 / name :op1 "RAF265"~e.17) :ARG0-of~e.23,32 (i2 / inhibit-01~e.23,32 :ARG1 (e / enzyme :wiki - :name (n5 / name :op1 "pan-RAF"~e.20,22)))) :op2 (s3 / small-molecule~e.32 :wiki "Binimetinib" :name (n6 / name :op1 "MEK162"~e.26) :ARG0-of~e.32 (i3 / inhibit-01~e.32 :ARG1 (e7 / enzyme :wiki "Mitogen-activated_protein_kinase_kinase" :name (n7 / name :op1 "MEK1/2"~e.29,31)))) :op3 (s / small-molecule~e.10,23,32,37 :wiki "Trametinib" :name (n8 / name :op1 "trametinib"~e.38) :ARG0-of~e.10,23,32,37 (i4 / inhibit-01~e.10,23,32,37 :ARG1 (p / protein-family :wiki "Mitogen-activated_protein_kinase_kinase" :name (n9 / name :op1 "MEK"~e.36))))) :ARG2 (i5 / investigate-01~e.1 :ARG0 w :ARG1 (a2 / affect-01~e.3 :ARG0 (i6 / inhibit-01~e.10 :ARG0 (a3 / and~e.8 :op1 (e9 / enzyme :wiki - :name (n10 / name :op1 "B-RAF"~e.5,7)) :op2 p~e.9)) :ARG1~e.11 (c / culture~e.13 :mod (m / medical-condition :wiki "Melanoma" :name (n12 / name :op1 "melanoma"~e.12)))))) # ::id a_pmid_2458_8908.77 ::amr-annotator SDL-AMR-09 ::preferred # ::tok A panel of 22 patient @-@ derived melanoma cultures was used ; the IC @ 50 for RAF265 and MEK162 are shown in Table 2 @ . # ::alignments 1-1.1.1 2-1.1.1.1.r 3-1.1.1.1.1 4-1.1.1.1.3.1.1.1 6-1.1.1.1.3 7-1.1.1.1.2.1.1 8-1.1.1.1 10-1.1 13-1.2.2.1 15-1.2.2.1.1 17-1.2.2.1.2.r 18-1.2.2.1.2.1.1.1 19-1.2.2.1.2 20-1.2.2.1.2.2.1.1 22-1.2 23-1.2.1.r 24-1.2.1 (m / multi-sentence :snt1 (u / use-01~e.10 :ARG1 (p / panel~e.1 :consist-of~e.2 (c / culture-01~e.8 :quant 22~e.3 :ARG1 (m2 / medical-condition :name (n / name :op1 "melanoma"~e.7)) :ARG1-of (d / derive-01~e.6 :ARG2 (p2 / person :ARG0-of (h / have-rel-role-91 :ARG2 (p3 / patient~e.4))))))) :snt2 (s / show-01~e.22 :ARG0~e.23 (t2 / table~e.24 :mod 1) :ARG1 (c2 / concentration-quantity :ARG4-of (h2 / have-percentage-maximal-inhibitory-concentration-01~e.13 :ARG2 50~e.15 :ARG1~e.17 (a / and~e.19 :op1 (s2 / small-molecule :name (n3 / name :op1 "RAF265"~e.18)) :op2 (s3 / small-molecule :name (n4 / name :op1 "MEK162"~e.20))))))) # ::id a_pmid_2458_8908.78 ::amr-annotator SDL-AMR-09 ::preferred # ::tok This was compared to the IC @ 50 for trametinib ( Additional file 1 @ : Table S1 ) . # ::alignments 0-1.1 2-1 3-1.2.r 5-1.2 7-1.2.1 9-1.2.2.r 10-1.2.2.1.1 12-1.3.1.2.2 13-1.3.1.2 15-1.3.1.2.1 18-1.3.1 19-1.3.1.1 (c / compare-01~e.2 :ARG1 (t / this~e.0) :ARG2~e.3 (h / have-percentage-maximal-inhibitory-concentration-01~e.5 :ARG2 50~e.7 :ARG1~e.9 (s / small-molecule :name (n2 / name :op1 "trametinib"~e.10))) :ARG1-of (d / describe-01 :ARG0 (t3 / table~e.18 :mod "S1"~e.19 :location (f / file~e.13 :mod 1~e.15 :mod (a / additional~e.12))))) # ::id a_pmid_2458_8908.79 ::amr-annotator SDL-AMR-09 ::preferred # ::tok Patient @-@ derived melanoma cultures with their B @-@ RAF/N @-@ RAS mutational status and sensitivity to RAF265 and MEK162 # ::alignments 0-1.2.1.1 2-1 3-1.1.1.1.1 4-1.1 5-1.3.r 6-1.3.1.2 6-1.3.1.2.r 7-1.3.1.1.1.1.1.1 11-1.3.1.1.1.2.1.1 12-1.3.1.1 13-1.3.1 14-1.3 15-1.3.2 16-1.3.2.2.r 17-1.3.2.2.1.1.1 19-1.3.2.2.2.1.1 (d / derive-01~e.2 :ARG1 (c / culture-01~e.4 :ARG1 (m2 / medical-condition :name (n / name :op1 "melanoma"~e.3))) :ARG2 (p / person :ARG0-of (h / have-rel-role-91 :ARG2 (p2 / patient~e.0))) :accompanier~e.5 (a / and~e.14 :op1 (s / status~e.13 :mod (m / mutate-01~e.12 :ARG1 (s5 / slash :op1 (e / enzyme :name (n5 / name :op1 "B-RAF"~e.7)) :op2 (e2 / enzyme :name (n6 / name :op1 "N-RAS"~e.11)))) :poss~e.6 c~e.6) :op2 (s2 / sensitive-03~e.15 :ARG0 c :ARG1~e.16 (a2 / and :op1 (s3 / small-molecule :name (n3 / name :op1 "RAF265"~e.17)) :op2 (s4 / small-molecule :name (n4 / name :op1 "MEK162"~e.19)))))) # ::id a_pmid_2458_8908.80 ::amr-annotator SDL-AMR-09 ::preferred # ::tok Cells were treated with each drug individually at concentrations ranging from 1 nM to 1000 nM and analyzed three days later . # ::alignments 0-1.1.1 2-1.1 3-1.1.2.r 4-1.1.2.2 5-1.1.2 6-1.1.3 6-1.1.3.r 7-1.1.2.1.r 8-1.1.2.1 8-1.1.2.1.1.1 8-1.1.2.1.1.2 9-1.1.2.1.1 11-1.1.2.1.1.1.1 12-1.1.2.1.1.1.2 14-1.1.2.1.1.2.1 15-1.1.2.1.1.2.2 16-1 17-1.2 18-1.2.2.1.1 19-1.2.2.1.2 20-1.2.2 20-1.2.2.2 20-1.2.2.2.r (a / and~e.16 :op1 (t / treat-04~e.2 :ARG1 (c / cell~e.0) :ARG2~e.3 (d / drug~e.5 :mod~e.7 (c2 / concentration~e.8 :ARG1-of (r / range-01~e.9 :ARG3 (c3 / concentration-quantity~e.8 :quant 1~e.11 :unit (n / nanomolar~e.12)) :ARG4 (c4 / concentration-quantity~e.8 :quant 1000~e.14 :unit n~e.15))) :mod (e / each~e.4)) :manner~e.6 (i / individual~e.6)) :op2 (a2 / analyze-01~e.17 :ARG1 c :time (l3 / late~e.20 :op1 (t3 / temporal-quantity :quant 3~e.18 :unit (d3 / day~e.19)) :degree~e.20 (m4 / more~e.20)))) # ::id a_pmid_2458_8908.81 ::amr-annotator SDL-AMR-09 ::preferred # ::tok As shown in Table 2 , the IC @ 50 for RAF265 ranged from 24 to > 10000 nM , 4 to 2004 nM , and 62 to 2082 nM for WT , B @-@ RAF mutant and N @-@ RAS mutant cultures , respectively . # ::alignments 1-1 2-1.1.r 3-1.1 5-1.1.1 9-1.2.1.1.1 9-1.2.2.1.1 9-1.2.3.1.1 11-1.2.1.1.1.1 11-1.2.2.1.1.1 11-1.2.3.1.1.1 13-1.2.1.1.1.2.r 13-1.2.r 14-1.2.1.1.1.2.1.1 15-1.2.1 15-1.2.2 15-1.2.3 16-1.2.1.2.r 17-1.2.1.2.1 18-1.2.1.3.r 19-1.2.1.3 20-1.2.1.3.1.1 21-1.2.1.2.2 23-1.2.2.2.1 24-1.2.2.3.r 25-1.2.2.3.1 26-1.2.1.2.2 28-1.2 29-1.2.3.2.1 30-1.2.3.3.r 31-1.2.3.3.1 32-1.2.3.3.2 34-1.2.1.4.1 36-1.2.2.4.1.1.1.1 38-1.2.2.4.1.1.1.1 39-1.2.2.4.1 40-1.2 41-1.2.3.4.1.1.1.1 43-1.2.3.4.1.1.1.1 44-1.2.2.4.1 44-1.2.3.4.1 45-1.2.1.4 45-1.2.2.4 45-1.2.3.4 47-1.2.4 (s / show-01~e.1 :ARG0~e.2 (t / table~e.3 :mod 2~e.5) :ARG1~e.13 (a / and~e.28,40 :op1 (r / range-01~e.15 :ARG1 (c10 / concentration-quantity :ARG4-of (h / have-percentage-maximal-inhibitory-concentration-01~e.9 :ARG2 50~e.11 :ARG1~e.13 (s2 / small-molecule :name (n / name :op1 "RAF265"~e.14)))) :ARG3~e.16 (c4 / concentration-quantity :quant 24~e.17 :unit (n5 / nanomolar~e.21,26)) :ARG4~e.18 (m / more-than~e.19 :op1 (c5 / concentration-quantity :quant 10000~e.20 :unit n5)) :location (c / culture-01~e.45 :mod (w / wild-type~e.34))) :op2 (r2 / range-01~e.15 :ARG1 (c11 / concentration-quantity :ARG4-of (h2 / have-percentage-maximal-inhibitory-concentration-01~e.9 :ARG2 50~e.11 :ARG1 s2)) :ARG3 (c6 / concentration-quantity :quant 4~e.23 :unit n5) :ARG4~e.24 (c7 / concentration-quantity :quant 2004~e.25 :unit n5) :location (c2 / culture-01~e.45 :mod (m2 / mutate-01~e.39,44 :ARG2 (e / enzyme :name (n2 / name :op1 "B-RAF"~e.36,38))))) :op3 (r3 / range-01~e.15 :ARG1 (c12 / concentration-quantity :ARG4-of (h3 / have-percentage-maximal-inhibitory-concentration-01~e.9 :ARG2 50~e.11 :ARG1 s2)) :ARG3 (c8 / concentration-quantity :quant 62~e.29 :unit n5) :ARG4~e.30 (c9 / concentration-quantity :quant 2082~e.31 :unit n5~e.32) :location (c3 / culture-01~e.45 :mod (m3 / mutate-01~e.44 :ARG2 (e2 / enzyme :name (n3 / name :op1 "N-RAS"~e.41,43))))) :mod (r4 / respective~e.47))) # ::id a_pmid_2458_8908.82 ::amr-annotator SDL-AMR-09 ::preferred # ::tok The IC @ 50 for MEK162 ranged from 10 to > 10000 nM , < 1 to 150 nM , and 4 to 13 nM for WT , B @-@ RAF mutant and N @-@ RAS mutant melanoma cultures , respectively . # ::alignments 1-1.1.1.1 1-1.2.1.1 1-1.3.1.1 3-1.1.1.1.1 3-1.2.1.1.1 3-1.3.1.1.1 5-1.1.1.1.2.r 6-1.1.1.1.2.1.1 7-1.1 7-1.2 7-1.3 8-1.1.2.r 9-1.1.2.1 10-1.1.3.r 11-1.1.3 12-1.1.3.1.1 13-1.1.3.1.2 15-1.2.2 16-1.2.2.1.1 17-1.2.3.r 18-1.2.3.1 19-1.1.2.2 21-1 22-1.3.2.1 23-1.3.3.r 24-1.3.3.1 25-1.3.3.2 27-1.1.1.2.1 29-1.2.1.2.1.2.1.1 31-1.2.1.2.1.2.1.1 32-1.2.1.2.1.2 32-1.2.1.2.1.2.2 32-1.2.1.2.1.2.2.r 32-1.3.1.2.1.2 32-1.3.1.2.1.2.2 32-1.3.1.2.1.2.2.r 33-1 34-1.3.1.2.1.2.1.1 36-1.3.1.2.1.2.1.1 37-1.2.1.2.1.2 37-1.2.1.2.1.2.2 37-1.2.1.2.1.2.2.r 37-1.3.1.2.1.2 37-1.3.1.2.1.2.2 37-1.3.1.2.1.2.2.r 38-1.2.1.2.1.1.1 39-1.1.1.2 39-1.2.1.2 39-1.3.1.2 41-1.4 (a / and~e.21,33 :op1 (r / range-01~e.7 :ARG1 (c10 / concentration-quantity :ARG4-of (h / have-percentage-maximal-inhibitory-concentration-01~e.1 :ARG2 50~e.3 :ARG1~e.5 (s / small-molecule :name (n3 / name :op1 "MEK162"~e.6))) :location (c / culture-01~e.39 :mod (w / wild-type~e.27))) :ARG3~e.8 (c4 / concentration-quantity :quant 10~e.9 :unit (n2 / nanomolar~e.19)) :ARG4~e.10 (m / more-than~e.11 :op1 (c5 / concentration-quantity :quant 10000~e.12 :unit n2~e.13))) :op2 (r2 / range-01~e.7 :ARG1 (c11 / concentration-quantity :ARG4-of (h2 / have-percentage-maximal-inhibitory-concentration-01~e.1 :ARG2 50~e.3 :ARG1 s) :location (c2 / culture-01~e.39 :mod (m2 / medical-condition :name (n5 / name :op1 "melanoma"~e.38) :mod (e2 / enzyme~e.32,37 :name (n6 / name :op1 "B-RAF"~e.29,31) :ARG2-of~e.32,37 (m3 / mutate-01~e.32,37))))) :ARG3 (l / less-than~e.15 :op1 (c6 / concentration-quantity :quant 1~e.16 :unit n2)) :ARG4~e.17 (c7 / concentration-quantity :quant 150~e.18 :unit n2)) :op3 (r3 / range-01~e.7 :ARG1 (c12 / concentration-quantity :ARG4-of (h3 / have-percentage-maximal-inhibitory-concentration-01~e.1 :ARG2 50~e.3 :ARG1 s) :location (c3 / culture-01~e.39 :mod (m5 / medical-condition :name n5 :mod (e / enzyme~e.32,37 :name (n9 / name :op1 "N-RAS"~e.34,36) :ARG2-of~e.32,37 (m4 / mutate-01~e.32,37))))) :ARG3 (c8 / concentration-quantity :quant 4~e.22 :unit n2) :ARG4~e.23 (c9 / concentration-quantity :quant 13~e.24 :unit n2~e.25)) :mod (r4 / respective~e.41)) # ::id a_pmid_2458_8908.83 ::amr-annotator SDL-AMR-09 ::preferred # ::tok The sensitivity to RAF265 in wild type ( 2 out of 5 ) and N @-@ RAS ( 2 out of 7 ) melanoma cultures was low . # ::alignments 1-1 2-1.2.r 3-1.2.1.1 4-1.1.r 5-1.1.1.2.2 6-1.1.1.2.2 8-1.1.1.1 11-1.1.1.3.1.1 13-1.1 14-1.1.2.2.2.1.1 16-1.1.2.2.2.1.1 18-1.1.2.1 21-1.1.2.3.1.1 23-1.1.1.2.1.1 23-1.1.2.2.1.1 24-1.1.1 24-1.1.1.3.1 24-1.1.2 24-1.1.2.3.1 26-1.3 (s / sensitive-03~e.1 :ARG0~e.4 (a2 / and~e.13 :op1 (c2 / culture-01~e.24 :quant 2~e.8 :ARG1 (m / medical-condition :name (n4 / name :op1 "melanoma"~e.23) :mod (w2 / wild-type~e.5,6)) :ARG1-of (i3 / include-91 :ARG2 (c / culture-01~e.24 :quant 5~e.11))) :op2 (c3 / culture-01~e.24 :quant 2~e.18 :ARG1 (m2 / medical-condition :name (n5 / name :op1 "melanoma"~e.23) :mod (e / enzyme :name (n6 / name :op1 "N-RAS"~e.14,16))) :ARG1-of (i4 / include-91 :ARG2 (c4 / culture-01~e.24 :quant 7~e.21)))) :ARG1~e.2 (s2 / small-molecule :name (n / name :op1 "RAF265"~e.3)) :ARG1-of (l / low-04~e.26)) # ::id a_pmid_2458_8908.84 ::amr-annotator SDL-AMR-09 ::preferred # ::tok Two wild type cultures ( YUROB and YUSOC ) are sensitive to both RAF265 and MEK162 . # ::alignments 0-1.1.1 1-1.1.2 2-1.1.2 3-1.1 5-1.1.3.1.1.1.1 6-1.1.3.1 7-1.1.3.1.2.1.1 10-1 11-1.2.r 12-1.2.3 13-1.2.1.1.1 14-1.2 15-1.2.2.1.1 (s / sensitive-03~e.10 :ARG0 (c / culture-01~e.3 :quant 2~e.0 :mod (w / wild-type~e.1,2) :ARG1-of (m / mean-01 :ARG2 (a / and~e.6 :op1 (c2 / cell-line :name (n / name :op1 "YUROB"~e.5)) :op2 (c3 / cell-line :name (n2 / name :op1 "YUSOC"~e.7))))) :ARG1~e.11 (a2 / and~e.14 :op1 (s2 / small-molecule :name (n3 / name :op1 "RAF265"~e.13)) :op2 (s3 / small-molecule :name (n4 / name :op1 "MEK162"~e.15)) :mod (b / both~e.12))) # ::id a_pmid_2458_8908.85 ::amr-annotator SDL-AMR-09 ::preferred # ::tok Six of ten B @-@ RAF mutant cultures were sensitive to RAF265 , and seven out of ten were sensitive to MEK162 . # ::alignments 0-1.1.1.1 2-1.1.1.3.1.1 3-1.1.1.2.1.1.1 5-1.1.1.2.1.1.1 6-1.1.1.2 7-1.1.1 7-1.1.1.3.1 7-1.2.1 9-1.1 9-1.2 11-1.1.2.1.1 13-1 14-1.2.1.1 17-1.1.1.3.1.1 19-1.1 20-1.2.2.r 21-1.2.2.1.1 (a / and~e.13 :op1 (s / sensitive-03~e.9,19 :ARG0 (c / culture-01~e.7 :quant 6~e.0 :mod (m / mutate-01~e.6 :ARG2 (e3 / enzyme :name (n3 / name :op1 "B-RAF"~e.3,5))) :ARG1-of (i3 / include-91 :ARG2 (c2 / culture-01~e.7 :quant 10~e.2,17))) :ARG1 (s3 / small-molecule :name (n4 / name :op1 "RAF265"~e.11))) :op2 (s2 / sensitive-03~e.9 :ARG0 (c3 / culture-01~e.7 :quant 7~e.14 :ARG1-of (i / include-91 :ARG2 c2) :mod m) :ARG1~e.20 (s4 / small-molecule :name (n5 / name :op1 "MEK162"~e.21)))) # ::id a_pmid_2458_8908.86 ::amr-annotator SDL-AMR-09 ::preferred # ::tok In N @-@ RAS mutant melanoma cultures , 2 out of 7 were sensitive to RAF265 and , strikingly , all were sensitive to MEK162 . # ::alignments 1-1.1.1.3.1.1.1 3-1.1.1.3.1.1.1 4-1.1.1.3 5-1.1.1.2.1.1 6-1.1.1 6-1.1.1.4.1 8-1.1.1.1 11-1.1.1.4.1.1 13-1.1 14-1.1.2.r 15-1.1.2.1.1 16-1 18-1.2.3 20-1.2.1 22-1.2 23-1.2.2.r 24-1.2.2.1.1 (a / and~e.16 :op1 (s / sensitive-03~e.13 :ARG0 (c / culture-01~e.6 :quant 2~e.8 :ARG1 (m2 / medical-condition :name (n3 / name :op1 "melanoma"~e.5)) :mod (m / mutate-01~e.4 :ARG1 (e3 / enzyme :name (n4 / name :op1 "N-RAS"~e.1,3))) :ARG1-of (i2 / include-91 :ARG2 (c2 / culture-01~e.6 :quant 7~e.11))) :ARG1~e.14 (s4 / small-molecule :name (n / name :op1 "RAF265"~e.15))) :op2 (s2 / sensitive-03~e.22 :ARG0 (a2 / all~e.20) :ARG1~e.23 (s5 / small-molecule :name (n2 / name :op1 "MEK162"~e.24)) :ARG1-of (s3 / strike-04~e.18))) # ::id a_pmid_2458_8908.87 ::amr-annotator SDL-AMR-09 ::preferred # ::tok Of the 7 N @-@ RAS mutant cultures , 5 were sensitive to trametinib . # ::alignments 2-1.1.3.1.1 3-1.1.2.1.1.1 5-1.1.2.1.1.1 6-1.1.2 7-1.1 7-1.1.3.1 9-1.1.1 11-1 12-1.2.r 13-1.2.1.1 (s / sensitive-03~e.11 :ARG0 (c / culture-01~e.7 :quant 5~e.9 :mod (m / mutate-01~e.6 :ARG2 (e / enzyme :name (n2 / name :op1 "N-RAS"~e.3,5))) :ARG1-of (i / include-91 :ARG2 (c2 / culture-01~e.7 :quant 7~e.2))) :ARG1~e.12 (s2 / small-molecule :name (n / name :op1 "trametinib"~e.13))) # ::id a_pmid_2458_8908.88 ::amr-annotator SDL-AMR-09 ::preferred # ::tok YUFIC and YUTICA were more resistant . # ::alignments 0-1.1.1.1.1 1-1.1 2-1.1.2.1.1 4-1.2 5-1 (r / resist-01~e.5 :ARG0 (a / and~e.1 :op1 (c / cell-line :name (n / name :op1 "YUFIC"~e.0)) :op2 (c2 / cell-line :name (n2 / name :op1 "YUTICA"~e.2))) :degree (m / more~e.4)) # ::id a_pmid_2458_8908.89 ::amr-annotator SDL-AMR-09 ::preferred # ::tok Molecular effects of MEK162 # ::alignments 0-1.2 1-1 2-1.1.r 3-1.1.1.1 (a / affect-01~e.1 :ARG0~e.2 (s / small-molecule :name (n / name :op1 "MEK162"~e.3)) :ARG1 (m / molecule~e.0)) # ::id a_pmid_2458_8908.90 ::amr-annotator SDL-AMR-09 ::preferred # ::tok Due to the striking sensitivity patterns of MEK162 , we conducted additional studies to verify target down @-@ regulation in the sensitive and resistant cultures . # ::alignments 0-1 1-1 3-1.1.2 4-1.1.1 5-1.1 6-1.1.1.1.r 7-1.1.1.1.1.1 9-1.2.1 10-1.2 11-1.2.2.2 12-1.2.2 14-1.2.3 15-1.2.3.2.1 16-1.2.3.2 17-1.2.3.2 18-1.2.3.2 19-1.2.3.2.2.r 21-1.2.3.2.2.1.1 22-1.2.3.2.2 23-1.2.3.2.2.2.1 24-1.2.3.2.2.1 24-1.2.3.2.2.2 (c / cause-01~e.0,1 :ARG0 (p / pattern-01~e.5 :ARG1 (s / sensitive-03~e.4 :ARG1~e.6 (s5 / small-molecule :name (n / name :op1 "MEK162"~e.7))) :ARG1-of (s2 / strike-04~e.3)) :ARG1 (c2 / conduct-01~e.10 :ARG0 (w / we~e.9) :ARG1 (s3 / study-01~e.12 :ARG0 w :mod (a / additional~e.11)) :purpose (v / verify-01~e.14 :ARG0 w :ARG1 (d / downregulate-01~e.16,17,18 :ARG1 (t / target-01~e.15) :location~e.19 (a2 / and~e.22 :op1 (c3 / culture-01~e.24 :ARG0-of (s6 / sensitive-03~e.21)) :op2 (c4 / culture-01~e.24 :ARG0-of (r2 / resist-01~e.23))))))) # ::id a_pmid_2458_8908.91 ::amr-annotator SDL-AMR-09 ::preferred # ::tok ERK1 @/@ 2 isoforms are the immediate downstream substrates and best studied effectors of dual specificity kinases MEK1 @/@ 2 . # ::alignments 0-1.3.1.1.1 2-1.3.1.1.1 3-1.3 4-1.3.r 6-1.1.2 7-1.1.1 8-1.1 9-1 10-1.2.1.1 10-1.2.1.1.1 10-1.2.1.1.1.r 11-1.2.1 12-1.2 13-1.2.2.r 14-1.2.2.2.1 15-1.2.2.2 16-1.2.2 17-1.2.2.1.1 19-1.2.2.1.1 (a2 / and~e.9 :op1 (s / substrate~e.8 :location (d / downstream~e.7) :mod (i2 / immediacy~e.6)) :op2 (e3 / effector~e.12 :ARG1-of (s4 / study-01~e.11 :ARG1-of (g / good-02~e.10 :degree~e.10 (m / most~e.10))) :poss~e.13 (k / kinase~e.16 :name (n2 / name :op1 "MEK1/2"~e.17,19) :mod (s3 / specificity~e.15 :mod (d2 / dual~e.14)))) :domain~e.4 (i / isoform~e.3 :mod (e2 / enzyme :name (n / name :op1 "ERK1/2"~e.0,2)))) # ::id a_pmid_2458_8908.92 ::amr-annotator SDL-AMR-09 ::preferred # ::tok To assess the effect of MEK1 @/@ 2 inhibition on ERK1 @/@ 2 activation state ( phosphorylation at T202 @/@ Y204 sites ) , melanoma cultures were treated with MEK162 and compared with untreated controls . # ::alignments 1-1.3 3-1.3.1 4-1.3.1.1.r 5-1.3.1.1.1.1.1 6-1.3.1.3.1.1 7-1.3.1.1.1.1.1 7-1.3.1.2.1.1.1.1 8-1.3.1.1 9-1.3.1.2.r 10-1.3.1.2.1.1.1.1 11-1.3.1.3.1.1 12-1.3.1.2.1.1.1.1 13-1.3.1.2.1 14-1.3.1.2 16-1.3.1.3.1 19-1.3.1.3.1.1 24-1.1.1.1.1.1 25-1.1.1 27-1.1 28-1.1.2.r 29-1.1.2.1.1 30-1 31-1.2 32-1.2.2.r 33-1.2.2.1 33-1.2.2.1.1 33-1.2.2.1.1.r 34-1.2.2 (a / and~e.30 :op1 (t / treat-04~e.27 :ARG1 (c2 / culture-01~e.25 :ARG1 (m2 / medical-condition :name (n3 / name :op1 "melanoma"~e.24))) :ARG2~e.28 (s3 / small-molecule :name (n4 / name :op1 "MEK162"~e.29))) :op2 (c / compare-01~e.31 :ARG1 c2 :ARG2~e.32 (c3 / control~e.34 :ARG1-of (t2 / treat-04~e.33 :polarity~e.33 -~e.33))) :purpose (a2 / assess-01~e.1 :ARG1 (a3 / affect-01~e.3 :ARG0~e.4 (i / inhibit-01~e.8 :ARG1 (e / enzyme :name (n / name :op1 "MEK1/2"~e.5,7))) :ARG1~e.9 (s / state~e.14 :mod (a4 / activate-01~e.13 :ARG1 (e2 / enzyme :name (n2 / name :op1 "ERK1/2"~e.7,10,12)))) :ARG2-of (m / mean-01 :ARG1 (p2 / phosphorylate-01~e.16 :ARG1 (s4 / slash~e.6,11,19 :op1 (a5 / amino-acid :mod 202 :name (n6 / name :op1 "threonine")) :op2 (a6 / amino-acid :mod 204 :name (n7 / name :op1 "tyrosine")))))))) # ::id a_pmid_2458_8908.93 ::amr-annotator SDL-AMR-09 ::preferred # ::tok We selected one sensitive and one resistant culture in the WT and B @-@ RAF mutant categories . # ::alignments 0-1.1 1-1 2-1.2.1.1 3-1.2.1.2 4-1.2 5-1.2.2.1 6-1.2.2.2 7-1.2.1 7-1.2.2 10-1.3.1.1 12-1.3.2.1.1.2.1 14-1.3.2.1.1.2.1 15-1.3.2.1 16-1.3.1 16-1.3.2 (s / select-01~e.1 :ARG0 (w / we~e.0) :ARG1 (a3 / and~e.4 :op1 (c3 / culture-01~e.7 :quant 1~e.2 :ARG0-of (s3 / sensitive-03~e.3)) :op2 (c4 / culture-01~e.7 :quant 1~e.5 :ARG0-of (r2 / resist-01~e.6))) :ARG2 (a / and :op1 (c / category~e.16 :mod (w4 / wild-type~e.10)) :op2 (c5 / category~e.16 :mod (m / mutate-01~e.15 :ARG2 (e3 / enzyme :wiki - :name (n3 / name :op1 "B-RAF"~e.12,14)))))) # ::id a_pmid_2458_8908.94 ::amr-annotator SDL-AMR-09 ::preferred # ::tok Seeing that all N @-@ RAS mutant cultures were sensitive to MEK162 , we selected two sensitive cultures for these studies . # ::alignments 0-1.1 1-1.1.1.r 2-1.1.1.1.2 3-1.1.1.1.1.1.1.1 5-1.1.1.1.1.1.1.1 6-1.1.1.1.1 7-1.1.1.1 9-1.1.1 10-1.1.1.2.r 11-1.1.1.2.1.1 13-1.2.1 14-1.2 15-1.2.2.1 16-1.2.2.2 17-1.2.2 18-1.2.3.r 19-1.2.3.1 20-1.2.3 (c / cause-01 :ARG0 (s / see-01~e.0 :ARG1~e.1 (s3 / sensitive-03~e.9 :ARG0 (c2 / culture-01~e.7 :mod (m / mutate-01~e.6 :ARG2 (e / enzyme :name (n / name :op1 "N-RAS"~e.3,5))) :mod (a / all~e.2)) :ARG1~e.10 (s6 / small-molecule :name (n2 / name :op1 "MEK162"~e.11)))) :ARG1 (s2 / select-01~e.14 :ARG0 (w / we~e.13) :ARG1 (c3 / culture-01~e.17 :quant 2~e.15 :ARG0-of (s4 / sensitive-03~e.16)) :ARG3~e.18 (s5 / study-01~e.20 :mod (t / this~e.19)))) # ::id a_pmid_2458_8908.95 ::amr-annotator SDL-AMR-09 ::preferred # ::tok WT ( YUVON and YUROB ) , B @-@ RAF mutant ( YUKSI and YUMAC ) and N @-@ RAS mutant ( YUDOSO and YUKIM ) cells were treated with increasing doses ( 10 @-@ 1000 nM ) of MEK162 or left untreated for 4 and 24 hours . # ::alignments 0-1.1.1.1.3 2-1.1.1.1.1.1.1 3-1.1.1.1 4-1.1.1.1.2.1.1 7-1.1.1.2.3.1.1 9-1.1.1.2.3.1.1 10-1.1.1.2.3 10-1.1.1.2.3.2 10-1.1.1.2.3.2.r 12-1.1.1.2.1.1.1 14-1.1.1.2.2.1.1 16-1.1.1 17-1.1.1.3.3.1.1 19-1.1.1.3.3.1.1 20-1.1.1.3.3 20-1.1.1.3.3.2 20-1.1.1.3.3.2.r 22-1.1.1.3.1.1.1 23-1.1.1.3 24-1.1.1.3.2.1.1 26-1.1.1.1.1 26-1.1.1.1.2 26-1.1.1.2.1 26-1.1.1.2.2 26-1.1.1.3.1 26-1.1.1.3.2 28-1.2.1 30-1.1.2.2.2 31-1.1.2.2 33-1.1.2.2.1.1.1 35-1.1.2.2.1.2.1 36-1.1.2.2.1.1.2 39-1.1.2.1.1 40-1 41-1.2 42-1.1 42-1.2.1.1 43-1.3.r 44-1.3.1.1 45-1.3 46-1.3.2.1 47-1.3.1.2 (o / or~e.40 :op1 (t2 / treat-04~e.42 :ARG1 (a6 / and~e.16 :op1 (a7 / and~e.3 :op1 (c2 / cell-line~e.26 :name (n / name :op1 "YUVON"~e.2)) :op2 (c3 / cell-line~e.26 :name (n2 / name :op1 "YUROB"~e.4)) :mod (w2 / wild-type~e.0)) :op2 (a2 / and :op1 (c4 / cell-line~e.26 :name (n6 / name :op1 "YUKSI"~e.12)) :op2 (c5 / cell-line~e.26 :name (n7 / name :op1 "YUMAC"~e.14)) :mod (e / enzyme~e.10 :name (n8 / name :op1 "B-RAF"~e.7,9) :ARG2-of~e.10 (m3 / mutate-01~e.10))) :op3 (a3 / and~e.23 :op1 (c6 / cell-line~e.26 :name (n3 / name :op1 "YUDOSO"~e.22)) :op3 (c7 / cell-line~e.26 :name (n9 / name :op1 "YUKIM"~e.24)) :mod (e2 / enzyme~e.20 :name (n10 / name :op1 "N-RAS"~e.17,19) :ARG2-of~e.20 (m / mutate-01~e.20)))) :ARG2 (s / small-molecule :name (n5 / name :op1 "MEK162"~e.39) :quant (d / dose~e.31 :quant (b / between :op1 (c8 / concentration-quantity :quant 10~e.33 :unit (n4 / nanomolar~e.36)) :op2 (c9 / concentration-quantity :quant 1000~e.35 :unit n4)) :ARG1-of (i / increase-01~e.30)))) :op2 (l / leave-14~e.41 :ARG1 (t3 / treat-04~e.28 :polarity -~e.42 :ARG1 a6)) :duration~e.43 (a5 / and~e.45 :op1 (t / temporal-quantity :quant 4~e.44 :unit (h / hour~e.47)) :op2 (t4 / temporal-quantity :quant 24~e.46 :unit h))) # ::id a_pmid_2458_8908.96 ::amr-annotator SDL-AMR-09 ::preferred # ::tok Western blot analysis was performed using phospho @-@ ERK1 @/@ 2 , total ERK1 @/@ 2 and β-actin antibodies , and results are shown in Figure 2 @ A.In the MEK162 resistant melanoma cultures ( YUVON and YUKSI ) , the baseline level of phospho @-@ ERK1 @/@ 2 and the ratio of phospho @-@ ERK1 @/@ 2 to total ERK1 @/@ 2 was lower compared to sensitive cultures ( YUROB , YUMAC , YUDOSO , YUKIM ) . # ::alignments 0-1.1.1.1 1-1.1.1.1 4-1.1.1 6-1.1.1.2.1.2 8-1.1.1.2.1.1.1 8-1.1.1.2.2.1.1 10-1.1.1.2.1.1.1 10-1.1.1.2.2.1.1 12-1.1.1.2.2.2 13-1.1.1.2.1.1.1 13-1.1.1.2.2.1.1 15-1.1.1.2.1.1.1 15-1.1.1.2.2.1.1 16-1.1.1.2 17-1.1.1.2.3.1.1.1.1 18-1.1.1.2.3 20-1.1 20-1.1.1.2 21-1.1.2.2 21-1.1.2.2.1 21-1.1.2.2.1.r 23-1.1.2 24-1.1.2.1.r 25-1.1.2.1 27-1.2.1.2.2.1.1 31-1.2.3.2.1.1.1 32-1.2.3.2 33-1.2.3.1.1.1 34-1.2.3 36-1.2.3.3.1.1.1.1 37-1.2.3.3.1 38-1.2.3.3.1.2.1.1 42-1.2.1.1.1 43-1.2.1.1 44-1.2.1.1.2.r 44-1.2.1.2 45-1.2.1.1.2.2 47-1.2.1.1.2.1.1 47-1.2.1.2.2.1.1 49-1.2.1.1.2.1.1 49-1.2.1.2.2.1.1 50-1.2.1 52-1.2.1.2 53-1.2.1.2 54-1.2.1.1.2.2 56-1.2.1.1.2.1.1 56-1.2.1.2.2.1.1 58-1.2.1.1.2.1.1 58-1.2.1.2.2.1.1 60-1.2.1.2.2.2 61-1.2.1.1.2.1.1 61-1.2.1.2.2.1.1 63-1.2.1.1.2.1.1 63-1.2.1.2.2.1.1 65-1.2 65-1.2.2 65-1.2.2.r 66-1.2.1.3 67-1.2.1.3.1.r 68-1.2.1.3.1.1 69-1.2.1.3.1 71-1.2.1.3.1.2.1.1.1.1 73-1.2.1.3.1.2.1.2.1.1 75-1.2.1.3.1.2.1.3.1.1 77-1.2.1.3.1.2.1.4.1.1 (m2 / multi-sentence :snt1 (a / and~e.20 :op1 (p / perform-01~e.4 :ARG1 (i / immunoblot-01~e.0,1) :ARG2 (a3 / and~e.16,20 :op1 (e3 / enzyme :name (n2 / name :op1 "ERK1/2"~e.8,10,13,15) :ARG3-of (p2 / phosphorylate-01~e.6)) :op2 (e4 / enzyme :name (n3 / name :op1 "ERK1/2"~e.8,10,13,15) :mod (t / total~e.12)) :op3 (a4 / antibody~e.18 :ARG0-of (c4 / counter-01 :ARG2 (p7 / protein :name (n4 / name :op1 "β-actin"~e.17)))))) :op2 (s / show-01~e.23 :ARG0~e.24 (f / figure~e.25 :mod "2A") :ARG1 (t2 / thing~e.21 :ARG2-of~e.21 (r / result-01~e.21)))) :snt2 (l2 / low-04~e.65 :ARG1 (a5 / and~e.50 :op1 (l3 / level~e.43 :domain (b2 / baseline~e.42) :quant-of~e.44 (e / enzyme :name (n8 / name :op1 "ERK1/2"~e.47,49,56,58,61,63) :ARG3-of (p8 / phosphorylate-01~e.45,54))) :op2 (r2 / ratio-of~e.44,52,53 :op1 e :op2 (e2 / enzyme :name (n9 / name :op1 "ERK1/2"~e.27,47,49,56,58,61,63) :mod (t3 / total~e.60))) :ARG1-of (c2 / compare-01~e.66 :ARG2~e.67 (c3 / culture-01~e.69 :ARG0-of (s3 / sensitive-03~e.68) :ARG1-of (m4 / mean-01 :ARG2 (a6 / and :op1 (c7 / cell-line :name (n11 / name :op1 "YUROB"~e.71)) :op2 (c8 / cell-line :name (n12 / name :op1 "YUMAC"~e.73)) :op3 (c9 / cell-line :name (n13 / name :op1 "YUDOSO"~e.75)) :op4 (c10 / cell-line :name (n14 / name :op1 "YUKIM"~e.77))))))) :degree~e.65 (m3 / more~e.65) :location (c / culture-01~e.34 :ARG1 (m5 / medical-condition :name (n6 / name :op1 "melanoma"~e.33)) :ARG0-of (r3 / resist-01~e.32 :ARG1 (s2 / small-molecule :name (n7 / name :op1 "MEK162"~e.31))) :ARG1-of (m / mean-01 :ARG2 (a8 / and~e.37 :op1 (c5 / cell-line :name (n5 / name :op1 "YUVON"~e.36)) :op2 (c6 / cell-line :name (n10 / name :op1 "YUKSI"~e.38))))))) # ::id a_pmid_2458_8908.97 ::amr-annotator SDL-AMR-09 ::preferred # ::tok In MEK162 @-@ sensitive melanomas exposure to MEK162 resulted in a significant decrease in the level of ERK1 @/@ 2 phosphorylation ( Figure 2 @ A ) . # ::alignments 1-1.1.1.2.1.1.1 3-1.1.1 3-1.1.1.2 3-1.1.1.2.r 4-1.1.1.1.1 5-1.1 6-1.1.2.r 7-1.1.2 8-1 9-1.2.r 11-1.2.3 12-1.2 13-1.2.2.r 15-1.2.2 16-1.2.2.1.r 17-1.2.2.1.1.1.1 19-1.2.2.1.1.1.1 20-1.2.2.1 22-1.3.1 24-1.2.2.1.1.1.1 (r / result-01~e.8 :ARG1 (e / expose-01~e.5 :ARG1 (m / medical-condition~e.3 :name (n / name :op1 "melanoma"~e.4) :ARG0-of~e.3 (s / sensitive-03~e.3 :ARG1 (s4 / small-molecule :name (n2 / name :op1 "MEK162"~e.1)))) :ARG2~e.6 s4~e.7) :ARG2~e.9 (d2 / decrease-01~e.12 :ARG0 e :ARG1~e.13 (l / level~e.15 :degree-of~e.16 (p2 / phosphorylate-01~e.20 :ARG1 (e2 / enzyme :name (n4 / name :op1 "ERK1/2"~e.17,19,24)))) :ARG2 (s2 / significant-02~e.11)) :ARG1-of (d3 / describe-01 :ARG0 (f / figure~e.22 :mod "2A"))) # ::id a_pmid_2514_2146.9 ::amr-annotator SDL-AMR-09 ::preferred # ::tok Sensitivity fell into three groups : sensitive , 50 % inhibitory concentration ( IC @ 50 ) < 1 μM ; intermediately sensitive , IC @ 50 1 @-@ 2 μM ; and resistant , > 2 μM . Fifteen of 21 ( 71 %) BRAF mutants , including 4 with innate vemurafenib resistance , were sensitive to SCH772984 . # ::alignments 0-1.1.1 0-1.2 1-1.1 2-1.1.2.r 3-1.1.2.1 4-1.1.2 6-1.1.1 8-1.1.2.2.1.2.2.1.1 9-1.2.1.4.2 10-1.1.2.2.1.2.2.1 11-1.1.2.2.1.1.1.1.2.1 11-1.1.2.2.1.2.2.1 11-1.1.2.2.1.2.2.1.2 11-1.1.2.2.1.2.2.1.2.r 11-1.1.2.2.1.3.1.1.2.1 13-1.1.2.2.1.1.1.1 13-1.1.2.2.1.2.2.1 15-1.1.2.2.1.1.1.1.1 15-1.1.2.2.1.2.2.1.1 18-1.1.2.2.1.1.1.1.2 19-1.1.2.2.1.1.1.1.2.1.1 20-1.1.2.2.1.1.1.1.2.1.2 22-1.1.2.2.1.2.1 23-1.1.2.2.1.1 23-1.1.2.2.1.2 25-1.1.2.2.1.1.1.1 25-1.1.2.2.1.3.1.1 27-1.1.2.2.1.1.1.1.1 27-1.1.2.2.1.3.1.1.1 29-1.1.2.2.1.2.2.1.2.1.1 31-1.1.2.2.1.2.2.1.2.1.2 31-1.1.2.2.1.3.1.1.2.1.1 32-1.1.2.2.1.1.1.1.2.1.2 34-1.1.2.2.1 34-1.1.2.2.1.2.2.1.2.1 35-1.1.2.2.1.3 37-1.1.2.2.1.3.1.1.2 38-1.1.2.2.1.2.2.1.2.1.2 39-1.1.2.2.1.2.2.1.2.2 41-1.2.1.1 43-1.2.1.4.1.1 45-1.2.1.4.2.1 48-1.2.1.2.1 48-1.2.1.4.1.2.1 48-1.2.1.5.1.2.1 50-1.2.1 50-1.2.1.3 50-1.2.1.3.r 52-1.2.1.4 52-1.2.1.5 53-1.2.1.5.1.1 55-1.2.1.5.1.3.2 56-1.2.1.5.1.3.1.1.1 57-1.2.1.5.1 57-1.2.1.5.1.3 57-1.2.1.5.1.3.r 60-1.2 61-1.2.2.r 62-1.2.2.1.1 (m6 / multi-sentence :snt1 (f / fall-04~e.1 :ARG1 (s / sensitive-03~e.0,6) :ARG2~e.2 (g / group~e.4 :quant 3~e.3 :ARG1-of (m9 / mean-01 :ARG2 (a / and~e.34 :op1 (s2 / sensitive-03~e.23 :ARG1-of (m / mean-01 :ARG2 (h / have-percentage-maximal-inhibitory-concentration-01~e.13,25 :ARG2 50~e.15,27 :ARG4 (l / less-than~e.18 :op1 (c / concentration-quantity~e.11 :quant 1~e.19 :unit (m2 / micromolar~e.20,32)))))) :op2 (s3 / sensitive-03~e.23 :mod (i / intermediate~e.22) :ARG1-of (m3 / mean-01 :ARG2 (h2 / have-percentage-maximal-inhibitory-concentration-01~e.10,11,13 :ARG2 50~e.8,15 :ARG4~e.11 (c3 / concentration-quantity~e.11 :quant (b / between~e.34 :op1 1~e.29 :op2 2~e.31,38) :unit m2~e.39)))) :op3 (r / resist-01~e.35 :ARG1-of (m4 / mean-01 :ARG2 (h3 / have-percentage-maximal-inhibitory-concentration-01~e.25 :ARG2 50~e.27 :ARG4 (m5 / more-than~e.37 :op1 (c2 / concentration-quantity~e.11 :quant 2~e.31 :unit m2))))))))) :snt2 (s4 / sensitive-03~e.0,60 :ARG0 (g2 / gene~e.50 :quant 15~e.41 :name (n4 / name :op1 "BRAF"~e.48) :ARG2-of~e.50 (m7 / mutate-01~e.50) :ARG1-of (i2 / include-91~e.52 :ARG2 (g3 / gene :quant 21~e.43 :name (n5 / name :op1 "BRAF"~e.48) :ARG2-of m7) :ARG3 (p / percentage-entity~e.9 :value 71~e.45)) :ARG2-of (i3 / include-01~e.52 :ARG1 (g4 / gene~e.57 :quant 4~e.53 :name (n6 / name :op1 "BRAF"~e.48) :ARG0-of~e.57 (r2 / resist-01~e.57 :ARG1 (s5 / small-molecule :name (n7 / name :op1 "vemurafenib"~e.56)) :mod (i4 / innate~e.55)) :ARG2-of m7))) :ARG1~e.61 (s6 / small-molecule :name (n8 / name :op1 "SCH772984"~e.62)))) # ::id a_pmid_2514_2146.10 ::amr-annotator SDL-AMR-09 ::preferred # ::tok All three ( 100 %) BRAF @/@ NRAS double mutants , 11 of 14 ( 78 %) NRAS mutants and 5 of 7 ( 71 %) wild @-@ type melanomas were sensitive . # ::alignments 0-1.1.1.5 1-1.1.1.1.1 3-1.1.1.4.1.1 6-1.1.1.1.2.1 7-1.1.1 8-1.1.1.2.1.1 8-1.1.2.4.1.2.1 10-1.1.1.3.1 11-1.1.1.3 11-1.1.2 11-1.1.2.3 11-1.1.2.3.r 13-1.1.2.1 15-1.1.2.4.1.1 17-1.1.2.4.2.1 20-1.1.1.2.1.1 20-1.1.2.2.1 22-1.1.1.3 22-1.1.2 22-1.1.2.3 22-1.1.2.3.r 23-1.1 24-1.1.3.1 26-1.1.3.4.1.1 28-1.1.3.4.2.1 30-1.1.3.3 32-1.1.3.3 33-1.1.3.2.1 33-1.1.3.4.1.2.1 35-1 (s / sensitive-03~e.35 :ARG0 (a / and~e.23 :op1 (s2 / slash~e.7 :op1 (g / gene :quant 3~e.1 :name (n / name :op1 "BRAF"~e.6)) :op2 (g4 / gene :name (n6 / name :op1 "NRAS"~e.8,20)) :ARG2-of (m / mutate-01~e.11,22 :mod (d / double~e.10)) :ARG1-of (i3 / include-91 :ARG3 (p / percentage-entity :value 100~e.3)) :mod (a2 / all~e.0)) :op2 (g2 / gene~e.11,22 :quant 11~e.13 :name (n2 / name :op1 "NRAS"~e.20) :ARG2-of~e.11,22 (m2 / mutate-01~e.11,22) :ARG1-of (i / include-91 :ARG2 (g3 / gene :quant 14~e.15 :name (n3 / name :op1 "NRAS"~e.8) :ARG2-of m2) :ARG3 (p2 / percentage-entity :value 78~e.17))) :op2 (m4 / medical-condition :quant 5~e.24 :name (n4 / name :op1 "melanoma"~e.33) :mod (w / wild-type~e.30,32) :ARG1-of (i2 / include-91 :ARG2 (m3 / medical-condition :quant 7~e.26 :name (n5 / name :op1 "melanoma"~e.33)) :ARG3 (p3 / percentage-entity :value 71~e.28))))) # ::id a_pmid_2514_2146.11 ::amr-annotator SDL-AMR-09 ::preferred # ::tok Among BRAF @ V600 @ mutants with in vitro acquired resistance to vemurafenib , those with MAPK pathway reactivation as the mechanism of resistance were sensitive to SCH772984 . # ::alignments 0-1.1.5 2-1.1.2.1 2-1.1.5.1.2.1 6-1.1.3.1 9-1.1 9-1.1.3 9-1.1.3.r 12-1.1.5.1.4.2.2 13-1.1.5.1.4.2.2 15-1.1.5.1.4.2 16-1.1.5.1 16-1.1.5.1.4 16-1.1.5.1.4.r 17-1.1.5.1.4.1.r 18-1.1.5.1.4.1.2.1 21-1.1.4.r 22-1.1.4.1.2.1 23-1.1.4.1 24-1.1.4 27-1.1.4.2.1 28-1.1.4.2.1.1.r 29-1.1.4.2.1.1 31-1 32-1.2.r 33-1.2.2.1 (s / sensitive-03~e.31 :ARG0 (g2 / gene~e.9 :wiki "BRAF_(gene)" :name (n3 / name :op1 "BRAF"~e.2) :ARG2-of~e.9 (m2 / mutate-01~e.9 :value "V600"~e.6) :ARG0-of~e.21 (r2 / reactivate-01~e.24 :ARG1 (p / pathway~e.23 :wiki "MAPK/ERK_pathway" :name (n4 / name :op1 "MAPK"~e.22)) :ARG1-of (m3 / mean-01 :ARG2 (m4 / mechanism~e.27 :mod~e.28 (r3 / resist-01~e.29)))) :ARG1-of (i2 / include-91~e.0 :ARG2 (g / gene~e.16 :wiki "BRAF_(gene)" :name (n / name :op1 "BRAF"~e.2) :ARG2-of m2 :ARG0-of~e.16 (r / resist-01~e.16 :ARG1~e.17 (s2 / small-molecule :wiki "Vemurafenib" :name (n2 / name :op1 "vemurafenib"~e.18)) :ARG1-of (a / acquire-01~e.15 :ARG0 g :manner (i / in-vitro~e.12,13)))))) :ARG1~e.32 (s3 / small-molecule :wiki - :name (n5 / name :op1 "SCH772984"~e.33))) # ::id a_pmid_2514_2146.12 ::amr-annotator SDL-AMR-09 ::preferred # ::tok SCH772984 caused G1 arrest and induced apoptosis . # ::alignments 0-1.1.1.1.1 1-1.1 2-1.1.2.1.1.1 3-1.1.2 4-1 5-1.2 6-1.2.2 (a / and~e.4 :op1 (c / cause-01~e.1 :ARG0 (s / small-molecule :name (n / name :op1 "SCH772984"~e.0)) :ARG1 (a2 / arrest-02~e.3 :ARG1 (t / thing :name (n2 / name :op1 "G1"~e.2)))) :op2 (i / induce-01~e.5 :ARG0 s :ARG2 (a3 / apoptosis~e.6))) # ::id a_pmid_2514_2146.54 ::amr-annotator SDL-AMR-09 ::preferred # ::tok BRAF @-@ mutant melanoma cell lines are sensitive to ERK inhibition # ::alignments 0-1.1.2.1.1 2-1.1.2 2-1.1.2.2 2-1.1.2.2.r 3-1.1.1.1.1 4-1.1 5-1.1 7-1 8-1.2.r 9-1.2.1.1.1 10-1.2 (s / sensitive-03~e.7 :ARG0 (c / cell-line~e.4,5 :mod (m / medical-condition :name (n3 / name :op1 "melanoma"~e.3)) :mod (e / enzyme~e.2 :name (n / name :op1 "BRAF"~e.0) :ARG2-of~e.2 (m2 / mutate-01~e.2))) :ARG1~e.8 (i / inhibit-01~e.10 :ARG1 (e2 / enzyme :name (n2 / name :op1 "ERK"~e.9)))) # ::id a_pmid_2514_2146.55 ::amr-annotator SDL-AMR-09 ::preferred # ::tok Twenty @-@ one melanoma cell lines containing mutations in the BRAF gene were evaluated to determine sensitivity to SCH772984 ( ERKi ) . # ::alignments 3-1.1.3.1.1 4-1.1 5-1.1 6-1.1.2 7-1.1.2.1 11-1.1.2.1.1.1.1 13-1.1.2.1.1 15-1 17-1.2 18-1.2.1 19-1.2.1.2.r 20-1.2.1.2.1.1 (e / evaluate-01~e.15 :ARG1 (c / cell-line~e.4,5 :quant 21 :ARG0-of (c2 / contain-01~e.6 :ARG1 (m2 / mutate-01~e.7 :ARG1 (g / gene~e.13 :name (n / name :op1 "BRAF"~e.11)))) :mod (m / medical-condition :name (n4 / name :op1 "melanoma"~e.3))) :purpose (d / determine-01~e.17 :ARG1 (s / sensitive-03~e.18 :ARG0 c :ARG1~e.19 (s2 / small-molecule :name (n2 / name :op1 "SCH772984"~e.20) :ARG0-of (i / inhibit-01 :ARG1 (p / protein-family :name (n3 / name :op1 "ERK"))))))) # ::id a_pmid_2514_2146.56 ::amr-annotator SDL-AMR-09 ::preferred # ::tok As a comparison , sensitivity to vemurafenib was also determined . # ::alignments 2-1.3 4-1.1 5-1.1.1.r 6-1.1.1.1.1 8-1.2 9-1 (d / determine-01~e.9 :ARG1 (s / sensitive-03~e.4 :ARG1~e.5 (s2 / small-molecule :name (n / name :op1 "vemurafenib"~e.6))) :mod (a / also~e.8) :ARG1-of (c / compare-01~e.2)) # ::id a_pmid_2514_2146.57 ::amr-annotator SDL-AMR-09 ::preferred # ::tok BRAF @ V600E @ was the most frequently observed BRAF mutation , present in 17 of 21 cell lines . # ::alignments 1-1.1.1.1 1-1.4.1.1 5-1.4.2.1 8-1.4.r 10-1.2.1.1 11-1.2.1 12-1.2 13-1.1.1.1 13-1.4.1.1 14-1 14-1.4 14-1.4.2 14-1.4.2.r 16-1.3 17-1.3.1.r 18-1.3.1.1 20-1.3.1.2.1.1 21-1.3.1 21-1.3.1.2.1 22-1.3.1.2.1 (m4 / mutate-01~e.14 :ARG2 (g3 / gene :name (n3 / name :op1 "BRAF"~e.1,13)) :ARG1-of (o / observe-01~e.12 :ARG1-of (f / frequent-02~e.11 :degree (m2 / most~e.10))) :ARG1-of (p / present-02~e.16 :ARG2~e.17 (c / cell-line~e.21 :quant 17~e.18 :ARG1-of (i2 / include-91 :ARG2 (c2 / cell-line~e.21,22 :quant 21~e.20)))) :domain~e.8 (g / gene~e.14 :name (n2 / name :op1 "BRAF"~e.1,13) :ARG2-of~e.14 (m / mutate-01~e.14 :value "V600E"~e.5))) # ::id a_pmid_2514_2146.58 ::amr-annotator SDL-AMR-09 ::preferred # ::tok M381 contains BRAF @ V600R @ substitution , M414 contains BRAF @ V600K @ , M417 contains BRAF @ G466E @ and M420 contains BRAF @ L597S @ mutation . # ::alignments 0-1.1.1.1.1 1-1.1 3-1.1.2.1.1.1 7-1.1.2.1.2.1 10-1.1.2 12-1.2.1.1.1 13-1.1 15-1.2.2.2 19-1.2.2.1 23-1.3.1.1.1 24-1.1 26-1.3.2.2 30-1.3.2.1 33-1 34-1.4.1.1.1 35-1.1 35-1.2 35-1.3 35-1.4 37-1.4.2.2 41-1.4.2.1 44-1.1.2.1 44-1.1.2.1.2 44-1.1.2.1.2.r 44-1.2.2 44-1.3.2 44-1.4.2 (a / and~e.33 :op1 (c / contain-01~e.1,13,24,35 :ARG0 (c2 / cell-line :name (n / name :op1 "M381"~e.0)) :ARG1 (s / substitute-01~e.10 :ARG2 (g / gene~e.44 :name (n2 / name :op1 "BRAF"~e.3) :ARG2-of~e.44 (m2 / mutate-01~e.44 :value "V600R"~e.7)))) :op2 (c3 / contain-01~e.35 :ARG0 (c4 / cell-line :name (n3 / name :op1 "M414"~e.12)) :ARG1 (m3 / mutate-01~e.44 :value "V600K"~e.19 :ARG1 g~e.15)) :op3 (c5 / contain-01~e.35 :ARG0 (c6 / cell-line :name (n4 / name :op1 "M417"~e.23)) :ARG1 (m4 / mutate-01~e.44 :value "G466E"~e.30 :ARG1 g~e.26)) :op4 (c7 / contain-01~e.35 :ARG0 (c8 / cell-line :name (n5 / name :op1 "M420"~e.34)) :ARG1 (m5 / mutate-01~e.44 :value "L597S"~e.41 :ARG1 g~e.37))) # ::id a_pmid_2514_2146.59 ::amr-annotator SDL-AMR-09 ::preferred # ::tok Among the 21 cell lines , sensitivity to vemurafenib or SCH @-@ 772984 fell into 3 groups : highly sensitive ( 50 % inhibitory concentration , IC @ 50 < 1 μM ) , intermediate sensitivity ( IC @ 50 1 @–@ 2 μM ) and resistant ( IC @ 50 > 2 μM ) . # ::alignments 2-1.1.1.1 3-1.1.1 4-1.1.1 6-1.1 7-1.1.2.r 8-1.1.2.1.1.1 9-1.1.2 10-1.1.2.2.1.1 12-1.1.2.2.1.1 13-1 14-1.2.r 15-1.2.1 16-1.2 18-1.2.2.1.1.1 19-1.2.2.1.1 21-1.2.2.1.2.2.1.1 23-1.2.2.1.2.2.1 24-1.2.2.1.1.2.1.2.1 24-1.2.2.1.2.2.1 24-1.2.2.1.2.2.1.2 24-1.2.2.1.2.2.1.2.r 24-1.2.2.1.3.1.1.2.1 26-1.2.2.1.1.2.1 26-1.2.2.1.2.2.1 28-1.2.2.1.2.2.1.1 30-1.2.2.1.1.2.1.2 31-1.2.2.1.1.2.1.2.1.1 32-1.2.2.1.1.2.1.2.1.2 35-1.2.2.1.2.1 36-1.2.2.1.2 38-1.2.2.1.1.2.1 40-1.2.2.1.1.2.1.1 42-1.2.2.1.2.2.1.2.1.1 44-1.2.2.1.2.2.1.2.1.2 44-1.2.2.1.3.1.1.2.1.1 45-1.2.2.1.1.2.1.2.1.2 47-1.2.2.1 47-1.2.2.1.2.2.1.2.1 48-1.2.2.1.3 50-1.2.2.1.3.1.1 52-1.2.2.1.3.1.1.1 54-1.2.2.1.3.1.1.2 55-1.2.2.1.2.2.1.2.1.2 56-1.2.2.1.2.2.1.2.2 (f / fall-04~e.13 :ARG1 (s / sensitive-03~e.6 :ARG0 (c2 / cell-line~e.3,4 :quant 21~e.2) :ARG1~e.7 (o / or~e.9 :op1 (s2 / small-molecule :name (n / name :op1 "vemurafenib"~e.8)) :op2 (s3 / small-molecule :name (n2 / name :op1 "SCH-772984"~e.10,12)))) :ARG2~e.14 (g / group~e.16 :quant 3~e.15 :ARG1-of (m6 / mean-01 :ARG2 (a / and~e.47 :op1 (s4 / sensitive-03~e.19 :ARG1-of (h / high-02~e.18) :ARG1-of (m / mean-01 :ARG2 (h2 / have-percentage-maximal-inhibitory-concentration-01~e.26,38 :ARG2 50~e.40 :ARG4 (l / less-than~e.30 :op1 (c / concentration-quantity~e.24 :quant 1~e.31 :unit (m2 / micromolar~e.32,45)))))) :op2 (s5 / sensitive-03~e.36 :mod (i2 / intermediate~e.35) :ARG1-of (m3 / mean-01 :ARG2 (h3 / have-percentage-maximal-inhibitory-concentration-01~e.23,24,26 :ARG2 50~e.21,28 :ARG4~e.24 (c3 / concentration-quantity~e.24 :quant (b / between~e.47 :op1 1~e.42 :op2 2~e.44,55) :unit m2~e.56)))) :op3 (r / resist-01~e.48 :ARG1-of (m4 / mean-01 :ARG2 (h4 / have-percentage-maximal-inhibitory-concentration-01~e.50 :ARG2 50~e.52 :ARG4 (m5 / more-than~e.54 :op1 (c4 / concentration-quantity~e.24 :quant 2~e.44 :unit m2))))))))) # ::id a_pmid_2514_2146.60 ::amr-annotator SDL-AMR-09 ::preferred # ::tok 15 cell lines were highly sensitive to SCH @-@ 772984 with IC @ 50 less than or equal to 1 μM . Of the 12 cell lines highly sensitive to vemurafenib , all contain BRAF @ V600E @ and were also sensitive to SCH @-@ 772984 . # ::alignments 0-1.1.1.1 1-1.1.1 2-1.1.1 4-1.1.3 5-1.1 6-1.1.2.r 7-1.1.2.1.1 9-1.1.2.1.1 11-1.1.2 11-1.1.2.2 11-1.1.2.2.r 13-1.1.2.2.1 19-1.1.2.2.2.r 20-1.1.2.2.2.1 21-1.1.2.2.2.2 25-1.2.1.1.1 26-1.2.1.1 27-1.2.1.1 28-1.2.1.1.3.2 29-1.2.1.1.3 30-1.2.1.1.3.1.r 31-1.2.1.1.3.1.1.1 33-1.2.1.1.2 34-1.2.1 36-1.2.1.2.1.1 40-1.2.1.2.2.1 43-1.2 45-1.2.2.3 46-1.2.2 47-1.2.2.2.r 48-1.2.2.2.1.1 50-1.2.2.2.1.1 (m / multi-sentence :snt1 (s / sensitive-03~e.5 :ARG0 (c / cell-line~e.1,2 :quant 15~e.0) :ARG1~e.6 (s2 / small-molecule~e.11 :name (n / name :op1 "SCH-772984"~e.7,9) :ARG1-of~e.11 (h3 / have-percentage-maximal-inhibitory-concentration-01~e.11 :ARG2 50~e.13 :ARG4~e.19 (c4 / concentration-quantity :quant 1~e.20 :unit (m2 / micromolar~e.21)))) :ARG1-of (h / high-02~e.4)) :snt2 (a / and~e.43 :op1 (c2 / contain-01~e.34 :ARG0 (c3 / cell-line~e.26,27 :quant 12~e.25 :mod (a2 / all~e.33) :ARG0-of (s3 / sensitive-03~e.29 :ARG1~e.30 (s4 / small-molecule :name (n2 / name :op1 "vemurafenib"~e.31)) :ARG1-of h~e.28)) :ARG1 (g / gene :name (n3 / name :op1 "BRAF"~e.36) :ARG2-of (m5 / mutate-01 :value "V600E"~e.40))) :op2 (s5 / sensitive-03~e.46 :ARG0 c3 :ARG1~e.47 (s6 / small-molecule :name (n4 / name :op1 "SCH-772984"~e.48,50)) :mod (a3 / also~e.45)))) # ::id a_pmid_2514_2146.61 ::amr-annotator SDL-AMR-09 ::preferred # ::tok Interestingly , M399 , M414 , M308 , and M409 were sensitive to SCH @-@ 772984 but only intermediately sensitive ( M399 , M409 and M414 ) or resistant ( M308 ) to vemurafenib . # ::alignments 0-1.4 2-1.1.1.1.1 4-1.1.2.1.1 6-1.1.3.1.1 8-1.1 9-1.1.4.1.1 11-1 12-1.2.r 13-1.2.1.1 15-1.2.1.1 16-1.3 17-1.3.1.1.3.1 18-1.3.1.1.3 19-1.3.1.1 21-1.1.1.1.1 23-1.1.4.1.1 25-1.1.2.1.1 27-1.3.1 28-1.3.1.2 30-1.3.1.2.1 32-1.3.1.1.2.r 33-1.3.1.1.2.1.1 (s / sensitive-03~e.11 :ARG0 (a / and~e.8 :op1 (c / cell-line :name (n / name :op1 "M399"~e.2,21)) :op2 (c2 / cell-line :name (n2 / name :op1 "M414"~e.4,25)) :op3 (c3 / cell-line :name (n3 / name :op1 "M308"~e.6)) :op4 (c5 / cell-line :name (n6 / name :op1 "M409"~e.9,23))) :ARG1~e.12 (s2 / small-molecule :name (n4 / name :op1 "SCH-772984"~e.13,15)) :ARG1-of (c4 / contrast-01~e.16 :ARG2 (o / or~e.27 :op1 (s3 / sensitive-03~e.19 :ARG0 (a3 / and :op1 c :op2 c5 :op3 c2) :ARG1~e.32 (s4 / small-molecule :name (n5 / name :op1 "vemurafenib"~e.33)) :mod (i2 / intermediate~e.18 :mod (o2 / only~e.17))) :op2 (r / resist-01~e.28 :ARG0 c3~e.30 :ARG1 s4))) :mod (i / interesting~e.0)) # ::id a_pmid_2514_2146.62 ::amr-annotator SDL-AMR-09 ::preferred # ::tok With the exception of M414 , all non @-@ V600E mutants were resistant to both vemurafenib and SCH772984 ( Figure 1 @ A ) . # ::alignments 2-1.1.4 3-1.1.4.1.r 4-1.1.4.1.1.1 6-1.1.3 7-1.1.2 7-1.1.2.r 9-1.1.1 10-1.1 12-1 15-1.2.1.1.1 16-1.2 17-1.2.2.1.1 19-1.3.1 (r / resist-01~e.12 :ARG0 (m / mutate-01~e.10 :value "V600E"~e.9 :polarity~e.7 -~e.7 :mod (a2 / all~e.6) :ARG2-of (e / except-01~e.2 :ARG1~e.3 (c / cell-line :name (n3 / name :op1 "M414"~e.4)))) :ARG1 (a / and~e.16 :op1 (s / small-molecule :name (n / name :op1 "vemurafenib"~e.15)) :op2 (s2 / small-molecule :name (n2 / name :op1 "SCH772984"~e.17))) :ARG1-of (d / describe-01 :ARG0 (f / figure~e.19 :mod "1A"))) # ::id a_pmid_2514_2146.63 ::amr-annotator SDL-AMR-09 ::preferred # ::tok As a comparison , sensitivity to the MEKi trametinib segregated all cell lines into three different groups : highly sensitive ( IC @ 50 < 2 nM ) , intermediately sensitive ( IC @ 50 2 @-@ 30 nM ) and resistant ( IC @ 50 > 30 nM ) ( Additional file 1 @ : Figure S1 ) . # ::alignments 2-1.5 4-1.1 8-1.1.1.1.1 9-1 10-1.2.1 11-1.2 12-1.2 13-1.3.r 14-1.3.1 15-1.3.2 16-1.3 18-1.3.3.1.1.1 19-1.3.3.1.1 21-1.3.3.1.1.2.1 23-1.3.3.1.1.2.1.1 25-1.3.3.1.1.2.1.2 26-1.3.3.1.1.2.1.2.1.1 27-1.3.3.1.1.2.1.2.1.2 30-1.3.3.1.2.1 30-1.3.3.1.2.1.r 31-1.3.3.1.2 33-1.3.3.1.1.2.1 33-1.3.3.1.2.2.1 35-1.3.3.1.1.2.1.1 35-1.3.3.1.2.2.1.1 37-1.3.3.1.1.2.1.2.1.1 37-1.3.3.1.2.2.1.2.1.1 39-1.3.3.1.2.2.1.2.1.2 40-1.3.3.1.2.2.1.2.2 42-1.3.3.1 42-1.3.3.1.2.2.1.2.1 43-1.3.3.1.3 45-1.3.3.1.3.1.1 47-1.3.3.1.3.1.1.1 49-1.3.3.1.3.1.1.2 50-1.3.3.1.2.2.1.2.1.2 50-1.3.3.1.3.1.1.2.1.1 51-1.3.3.1.1.2.1.2.1.2 55-1.4.1 57-1.4.1.1 60-1.4.1.3.1 61-1.4.1.3.1.1 (s / segregate-01~e.9 :ARG0 (s2 / sensitive-03~e.4 :ARG1 (s3 / small-molecule :name (n / name :op1 "trametinib"~e.8) :ARG0-of (i / inhibit-01 :ARG1 (e / enzyme :name (n2 / name :op1 "MEK"))))) :ARG1 (c2 / cell-line~e.11,12 :mod (a / all~e.10)) :ARG2~e.13 (g / group~e.16 :quant 3~e.14 :ARG1-of (d / differ-02~e.15) :ARG1-of (m5 / mean-01 :ARG2 (a2 / and~e.42 :op1 (s4 / sensitive-03~e.19 :ARG1-of (h / high-02~e.18) :ARG1-of (m / mean-01 :ARG2 (h2 / have-percentage-maximal-inhibitory-concentration-01~e.21,33 :ARG2 50~e.23,35 :ARG4 (l / less-than~e.25 :op1 (c3 / concentration-quantity :quant 2~e.26,37 :unit (n3 / nanomolar~e.27,51)))))) :op2 (s5 / sensitive-03~e.31 :manner~e.30 (i2 / intermediate~e.30) :ARG1-of (m2 / mean-01 :ARG2 (h3 / have-percentage-maximal-inhibitory-concentration-01~e.33 :ARG2 50~e.35 :ARG4 (c4 / concentration-quantity :quant (b / between~e.42 :op1 2~e.37 :op2 30~e.39,50) :unit n3~e.40)))) :op3 (r / resist-01~e.43 :ARG1-of (m3 / mean-01 :ARG2 (h4 / have-percentage-maximal-inhibitory-concentration-01~e.45 :ARG2 50~e.47 :ARG4 (m4 / more-than~e.49 :op1 (c5 / concentration-quantity :quant 30~e.50 :unit n3)))))))) :ARG1-of (d2 / describe-01 :ARG0 (f3 / file~e.55 :mod 1~e.57 :ARG1-of (a4 / add-02) :ARG1-of (m6 / mean-01 :ARG2 (f2 / figure~e.60 :mod "S1"~e.61)))) :ARG1-of (c / compare-01~e.2)) # ::id a_pmid_2514_2146.64 ::amr-annotator SDL-AMR-09 ::preferred # ::tok In general , cell lines sensitive to SCH772984 were also sensitive to trametinib . @ @

Effect of SCH @-@ 722984 on BRAF @-@ mutant melanoma cell lines . # ::alignments 1-1.1.3 3-1.1.1 4-1.1.1 5-1.1.1.1 6-1.1.1.1.1.r 7-1.1.1.1.1.1.1 9-1.1.4 10-1.1 11-1.1.2.r 12-1.1.2.1.1 16-1.2 17-1.2.1 22-1.2.2.1 23-1.2.2.1.1.r 24-1.2.2.1.1.1.1 26-1.2.2.1.1.1.1 27-1.2.2.1.2.r 28-1.2.2.1.2.2.1.1 30-1.2.2.1.2.2 30-1.2.2.1.2.2.2 30-1.2.2.1.2.2.2.r 31-1.2.2.1.2.1.1.1 32-1.2.2.1.2 33-1.2.2.1.2 (m / multi-sentence :snt1 (s / sensitive-03~e.10 :ARG0 (c / cell-line~e.3,4 :ARG0-of (s3 / sensitive-03~e.5 :ARG1~e.6 (s4 / small-molecule :name (n2 / name :op1 "SCH772984"~e.7)))) :ARG1~e.11 (s2 / small-molecule :name (n / name :op1 "trametinib"~e.12)) :ARG1-of (g / general-02~e.1) :mod (a / also~e.9)) :snt2 (f2 / figure~e.16 :mod 1~e.17 :ARG1-of (d2 / describe-01 :ARG2 (a2 / affect-01~e.22 :ARG0~e.23 (s5 / small-molecule :name (n3 / name :op1 "SCH-722984"~e.24,26)) :ARG1~e.27 (c2 / cell-line~e.32,33 :mod (m2 / medical-condition :name (n5 / name :op1 "melanoma"~e.31)) :mod (e / enzyme~e.30 :name (n4 / name :op1 "BRAF"~e.28) :ARG2-of~e.30 (m3 / mutate-01~e.30))))))) # ::id a_pmid_2514_2146.65 ::amr-annotator SDL-AMR-09 ::preferred # ::tok A @ . # ::alignments 0-1.1 (h / have-li-91 :ARG2 "A"~e.0) # ::id a_pmid_2514_2146.66 ::amr-annotator SDL-AMR-09 ::preferred # ::tok IC @ 50 ( nM ) . # ::alignments 0-1 2-1.1 5-1.2.1 (h / have-percentage-maximal-inhibitory-concentration-01~e.0 :ARG2 50~e.2 :ARG4 (c / concentration-quantity :unit (n / nanomolar~e.5))) # ::id a_pmid_2514_2146.67 ::amr-annotator SDL-AMR-09 ::preferred # ::tok 21 BRAF @-@ mutant melanoma cell lines were exposed to 0 @–@ 10 μM SCH @-@ 722984 ( black bars ) or vemurafenib ( grey bars ) and cell viability determined by ATP @-@ based bioluminescence assay . # ::alignments 0-1.1.1.1 1-1.1.1.3.1.1 3-1.1.1.3 3-1.1.1.3.2 3-1.1.1.3.2.r 4-1.1.1.2.1.1 5-1.1.1 6-1.1.1 8-1.1 9-1.1.2.r 10-1.1.2.1.2.1.1 12-1.1.2.1.2.1.2 13-1.1.2.1.2.2 14-1.1.2.1.1.1 16-1.1.2.1.1.1 18-1.1.2.1.3.1.1 19-1.1.2.1.3.1 21-1.1.2 22-1.1.2.2.1.1 24-1.1.2.2.3.1.1 25-1.1.2.2.3.1 28-1.1.1 29-1.2.1 30-1.2 31-1.2.2.r 32-1.2.2.2.1.1.1 34-1.2.2.2 35-1.2.2.1 36-1.2.2 (a / and :op1 (e / expose-01~e.8 :ARG1 (c / cell-line~e.5,6,28 :quant 21~e.0 :mod (m3 / medical-condition :name (n5 / name :op1 "melanoma"~e.4)) :mod (e2 / enzyme~e.3 :name (n / name :op1 "BRAF"~e.1) :ARG2-of~e.3 (m2 / mutate-01~e.3))) :ARG2~e.9 (o / or~e.21 :op1 (s / small-molecule :name (n2 / name :op1 "SCH-722984"~e.14,16) :quant (c2 / concentration-quantity :quant (b / between :op1 0~e.10 :op2 10~e.12) :unit (m / micromolar~e.13)) :ARG1-of (d / describe-01 :ARG0 (b2 / bar~e.19 :ARG1-of (b3 / black-04~e.18)))) :op2 (s2 / small-molecule :name (n3 / name :op1 "vemurafenib"~e.22) :quant c2 :ARG1-of (d4 / describe-01 :ARG0 (b4 / bar~e.25 :ARG1-of (g2 / gray-02~e.24)))))) :op2 (d2 / determine-01~e.30 :ARG1 (v / viability~e.29 :mod c) :instrument~e.31 (a2 / assay-01~e.36 :ARG1 (b5 / bioluminescence~e.35) :ARG1-of (b6 / base-02~e.34 :ARG2 (s3 / small-molecule :name (n4 / name :op1 "ATP"~e.32)))))) # ::id a_pmid_2514_2146.68 ::amr-annotator SDL-AMR-09 ::preferred # ::tok Results are the mean of three experiments , performed in duplicates ( n = 6 ) . # ::alignments 0-1 3-1.1 4-1.1.1.r 5-1.1.1.1 6-1.1.1 8-1.1.1.2 9-1.1.1.2.1.r 10-1.1.1.2.1 14-1.1.1.2.1.1 (r / result-01~e.0 :ARG2 (m / mean~e.3 :poss~e.4 (e / experiment-01~e.6 :quant 3~e.5 :ARG1-of (p / perform-01~e.8 :manner~e.9 (d / duplicate~e.10 :quant 6~e.14))))) # ::id a_pmid_2514_2146.69 ::amr-annotator SDL-AMR-09 ::preferred # ::tok Error bars are standard deviation . # ::alignments 0-1.2.1 1-1.2 2-1.2.r 3-1.1 4-1 (d / deviate-01~e.4 :ARG1-of (s / standard-02~e.3) :domain~e.2 (b / bar~e.1 :mod (e / error~e.0))) # ::id a_pmid_2514_2146.70 ::amr-annotator SDL-AMR-09 ::preferred # ::tok Non @-@ V600E substitutions are denoted in the bar graph for each corresponding cell line ( M420 , BRAF @ L597S ; M381 , BRAF @ V600R , M417 , BRAF @ G466E , M414 , BRAF @ V600K ) . # ::alignments 0-1.1.1.2 0-1.1.1.2.r 2-1.1.1.1 3-1.1 5-1 6-1.2.r 8-1.2.1 9-1.2 12-1.1.2 12-1.3.1.1 12-1.3.1.2 12-1.3.1.3 12-1.3.1.4 13-1.1.2.1 14-1.1.2.1 16-1.3.1.1.1.1.1 18-1.3.1.1.2.1.1 20-1.3.1.1.2.2.1 23-1.3.1.2.1.1.1 25-1.3.1.2.2.1.1 27-1.3.1.2.2.2.1 30-1.3.1.3.1.1.1 32-1.3.1.3.2.1.1 34-1.3.1.3.2.2.1 37-1.3.1.4.1.1.1 39-1.3.1.4.2.1.1 41-1.3.1.4.2.2.1 (d / denote-01~e.5 :ARG1 (s / substitute-01~e.3 :ARG2 (m / mutate-01 :value "V600E"~e.2 :polarity~e.0 -~e.0) :ARG1-of (c / correspond-02~e.12 :ARG2 (c2 / cell-line~e.13,14))) :location~e.6 (g / graph~e.9 :mod (b / bar~e.8)) :ARG1-of (m2 / mean-01 :ARG2 (a / and :op1 (c3 / correspond-02~e.12 :ARG1 (c4 / cell-line :name (n / name :op1 "M420"~e.16)) :ARG2 (e / enzyme :name (n5 / name :op1 "BRAF"~e.18) :ARG2-of (m3 / mutate-01 :value "L597S"~e.20))) :op2 (c5 / correspond-02~e.12 :ARG1 (c8 / cell-line :name (n2 / name :op1 "M381"~e.23)) :ARG2 (e2 / enzyme :name (n6 / name :op1 "BRAF"~e.25) :ARG2-of (m4 / mutate-01 :value "V600R"~e.27))) :op3 (c6 / correspond-02~e.12 :ARG1 (c9 / cell-line :name (n3 / name :op1 "M417"~e.30)) :ARG2 (e3 / enzyme :name (n7 / name :op1 "BRAF"~e.32) :ARG2-of (m5 / mutate-01 :value "G466E"~e.34))) :op4 (c7 / correspond-02~e.12 :ARG1 (c10 / cell-line :name (n4 / name :op1 "M414"~e.37)) :ARG2 (e4 / enzyme :name (n8 / name :op1 "BRAF"~e.39) :ARG2-of (m6 / mutate-01 :value "V600K"~e.41)))))) # ::id a_pmid_2514_2146.71 ::amr-annotator SDL-AMR-09 ::preferred # ::tok Bar at 1 μM denotes threshold between sensitive and intermediate . # ::alignments 0-1.1 1-1.1.1.r 2-1.1.1.1 3-1.1.1.2 4-1 5-1.2 6-1.2.1 7-1.2.1.1 8-1.2.1 9-1.2.1.2 (d / denote-01~e.4 :ARG0 (b / bar~e.0 :quant~e.1 (c / concentration-quantity :quant 1~e.2 :unit (m / micromolar~e.3))) :ARG1 (t / threshold~e.5 :topic (b2 / between~e.6,8 :op1 (s / sensitive-03~e.7) :op2 (i / intermediate~e.9)))) # ::id a_pmid_2514_2146.72 ::amr-annotator SDL-AMR-09 ::preferred # ::tok Resistant cell lines have IC @ 50 higher than 2 μM . # ::alignments 0-1.2.1 1-1.2 2-1.2 3-1 4-1 6-1.1 9-1.3 10-1.3.1.1 11-1.3.1.2 (h2 / have-percentage-maximal-inhibitory-concentration-01~e.3,4 :ARG2 50~e.6 :ARG1 (c / cell-line~e.1,2 :ARG0-of (r / resist-01~e.0)) :ARG4 (m / more-than~e.9 :op1 (c2 / concentration-quantity :quant 2~e.10 :unit (m2 / micromolar~e.11)))) # ::id a_pmid_2514_2146.73 ::amr-annotator SDL-AMR-09 ::preferred # ::tok B @ . # ::alignments 1-1.1 (h / have-li-91 :ARG2 "B"~e.1) # ::id a_pmid_2514_2146.74 ::amr-annotator SDL-AMR-09 ::preferred # ::tok Timecourse effects of SCH722984 on the MAPK signaling . # ::alignments 0-1.3 1-1 2-1.1.r 3-1.1.1.1 4-1.2.r 6-1.2.1.1.1 7-1.2 (a / affect-01~e.1 :ARG0~e.2 (s / small-molecule :name (n / name :op1 "SCH722984"~e.3)) :ARG1~e.4 (s2 / signal-07~e.7 :ARG0 (p / pathway :name (n2 / name :op1 "MAPK"~e.6))) :mod (t / timecourse~e.0)) # ::id a_pmid_2514_2146.75 ::amr-annotator SDL-AMR-09 ::preferred # ::tok SCH722984 @-@ sensitive M238 , SCH722984 @-@ resistant M233 , were treated in a timecourse manner with 500 nM SCH722984 at 1 , 2 , 6 , 12 , 24 and 48 hours compared to DMSO as solvent control ( C ) @ . # ::alignments 0-1.1.1.2.1.1.1 0-1.2.1.1 2-1.1.1 2-1.1.1.2 2-1.1.1.2.r 3-1.1.1.1.1 5-1.2.1.1 7-1.1.2 7-1.1.2.2 7-1.1.2.2.r 8-1.1.2.1.1 11-1 14-1.4 15-1.4.r 17-1.2.2.1 18-1.2.2.2 19-1.2.1.1 21-1.3.1.1.1 23-1.3.1.2.1 25-1.3.1.3.1 27-1.3.1.4.1 29-1.3.1.5.1 30-1.3.1 31-1.3.1.6.1 32-1.3.1.1.2 33-1.2.3 34-1.2.3.1.r 35-1.2.3.1.1.1 36-1.3.r 38-1.2.3.1 38-1.2.3.1.2 38-1.2.3.1.2.r (t / treat-04~e.11 :ARG1 (a / and :op1 (c / cell-line~e.2 :name (n / name :op1 "M238"~e.3) :ARG0-of~e.2 (s2 / sensitive-03~e.2 :ARG1 (s / small-molecule :name (n2 / name :op1 "SCH722984"~e.0)))) :op2 (c2 / cell-line~e.7 :name (n3 / name :op1 "M233"~e.8) :ARG0-of~e.7 (r / resist-01~e.7 :ARG1 s))) :ARG2 (s3 / small-molecule :name (n4 / name :op1 "SCH722984"~e.0,5,19) :quant (c5 / concentration-quantity :quant 500~e.17 :unit (n5 / nanomolar~e.18)) :ARG1-of (c3 / compare-01~e.33 :ARG2~e.34 (s5 / small-molecule~e.38 :name (n6 / name :op1 "DMSO"~e.35) :ARG0-of~e.38 (c4 / control-01~e.38 :ARG1 (d / dissolve-01))))) :time~e.36 (a2 / after :op1 (a3 / and~e.30 :op1 (t2 / temporal-quantity :quant 1~e.21 :unit (h / hour~e.32)) :op2 (t3 / temporal-quantity :quant 2~e.23 :unit h) :op3 (t4 / temporal-quantity :quant 6~e.25 :unit h) :op4 (t5 / temporal-quantity :quant 12~e.27 :unit h) :op5 (t6 / temporal-quantity :quant 24~e.29 :unit h) :op6 (t7 / temporal-quantity :quant 48~e.31 :unit h))) :manner~e.15 (t8 / timecourse~e.14)) # ::id a_pmid_2514_2146.76 ::amr-annotator SDL-AMR-09 ::preferred # ::tok Phosphorylated or total MEK , ERK1 @/@ 2 , RSK , AKT , or beta @-@ actin as loading control were determined by western blot analysis . # ::alignments 0-1.1.1.1.2 1-1.1 2-1.1.2.1.2 3-1.1.1.1.1.1 3-1.1.2.1.1.1 5-1.1.1.2.1.1 5-1.1.2.2.1.1 7-1.1.1.2.1.1 7-1.1.2.2.1.1 9-1.1.1.3.1.1 9-1.1.2.3.1.1 11-1.1.1.4.1.1 11-1.1.2.4.1.1 13-1.1 14-1.1.3.1.1 16-1.1.3.1.1 18-1.1.3.2.1 19-1.1.3 19-1.1.3.2 19-1.1.3.2.r 21-1 22-1.2.r 23-1.2.1 24-1.2.1 25-1.2 (d / determine-01~e.21 :ARG1 (o / or~e.1,13 :op1 (a / and :op1 (e / enzyme :name (n / name :op1 "MEK"~e.3) :ARG3-of (p / phosphorylate-01~e.0)) :op2 (e2 / enzyme :name (n2 / name :op1 "ERK1/2"~e.5,7) :ARG3-of p) :op3 (e3 / enzyme :name (n3 / name :op1 "RSK"~e.9) :ARG3-of p) :op4 (e4 / enzyme :name (n4 / name :op1 "AKT"~e.11) :ARG3-of p)) :op2 (a2 / and :op1 (e8 / enzyme :name (n8 / name :op1 "MEK"~e.3) :mod (t / total~e.2)) :op2 (e7 / enzyme :name (n7 / name :op1 "ERK1/2"~e.5,7) :mod t) :op3 (e6 / enzyme :name (n6 / name :op1 "RSK"~e.9) :mod t) :op4 (e5 / enzyme :name (n5 / name :op1 "AKT"~e.11) :mod t)) :op3 (p2 / protein~e.19 :name (n9 / name :op1 "beta-actin"~e.14,16) :ARG0-of~e.19 (c / control-01~e.19 :ARG1 (l / load-01~e.18)))) :manner~e.22 (a3 / analyze-01~e.25 :manner (i / immunoblot-01~e.23,24))) # ::id a_pmid_2514_2146.77 ::amr-annotator SDL-AMR-09 ::preferred # ::tok C @ . # ::alignments 1-1.1 (h / have-li-91 :ARG2 "C"~e.1) # ::id a_pmid_2514_2146.78 ::amr-annotator SDL-AMR-09 ::preferred # ::tok Effects of SCH722984 on the MAPK signaling at 24 hours . # ::alignments 0-1 1-1.1.r 2-1.1.1.1 3-1.2.r 5-1.2.1.1.1 6-1.2 8-1.3.1.1 9-1.3.1.2 (a2 / affect-01~e.0 :ARG0~e.1 (s / small-molecule :name (n / name :op1 "SCH722984"~e.2)) :ARG1~e.3 (s2 / signal-07~e.6 :ARG0 (p / pathway :name (n2 / name :op1 "MAPK"~e.5))) :time (a / after :quant (t / temporal-quantity :quant 24~e.8 :unit (h / hour~e.9)))) # ::id a_pmid_2514_2146.79 ::amr-annotator SDL-AMR-09 ::preferred # ::tok SCH722984 @-@ sensitive M262 , SCH722984 @-@ resistant M381 , SCH722984 @-@ intermediately sensitive M409 cells were treated for 24 h with DMSO as solvent control (-) or 500 nM SCH722984 (+) .

@ @ @ # ::alignments 0-1.2.2.1.1 2-1.1.3.2 3-1.1.1.1.1 5-1.1.2.2.1 7-1.1.2.2 8-1.1.2.1.1 10-1.1.3.2.1 12-1.1.3.2.2 13-1.1.1.2 13-1.1.3.2 14-1.1.3.1.1 15-1.1.1 15-1.1.2 15-1.1.3 17-1 18-1.3.r 19-1.3.1 20-1.3.2 21-1.2.r 22-1.2.1.1.1 25-1.2.1 25-1.2.1.2 25-1.2.1.2.r 27-1.2 28-1.2.2.2.1 29-1.2.2.2.2 30-1.2.2.1.1 (t / treat-04~e.17 :ARG1 (a / and :op1 (c / cell-line~e.15 :name (n / name :op1 "M262"~e.3) :ARG0-of (s / sensitive-03~e.13 :ARG1 (s2 / small-molecule :name (n2 / name :op1 "SCH722984")))) :op2 (c2 / cell-line~e.15 :name (n3 / name :op1 "M381"~e.8) :ARG0-of (r / resist-01~e.7 :ARG1 s2~e.5)) :op3 (c3 / cell-line~e.15 :name (n4 / name :op1 "M409"~e.14) :ARG0-of (s3 / sensitive-03~e.2,13 :ARG1 s2~e.10 :mod (i / intermediate~e.12)))) :ARG2~e.21 (o / or~e.27 :op1 (s4 / small-molecule~e.25 :name (n5 / name :op1 "DMSO"~e.22) :ARG0-of~e.25 (c4 / control-01~e.25 :ARG1 (d / dissolve-01))) :op2 (s5 / small-molecule :name (n6 / name :op1 "SCH722984"~e.0,30) :quant (c5 / concentration-quantity :quant 500~e.28 :unit (n7 / nanomolar~e.29)))) :duration~e.18 (t2 / temporal-quantity :quant 24~e.19 :unit (h / hour~e.20))) # ::id a_pmid_2514_2146.80 ::amr-annotator SDL-AMR-09 ::preferred # ::tok We next determined a time @-@ course of SCH772984 on MAPK and PI3K @/@ AKT pathway signaling for M238 , a SCH772984 @-@ sensitive BRAF @ V600E @ @ -@ mutant melanoma cell line and M233 , a SCH772984 @-@ resistant BRAF @ V600E @ @ -@ mutant melanoma cell line ( Figure 1 @ B ) . # ::alignments 0-1.1 1-1.3 2-1 4-1.3.r 8-1.2.1.1.1 10-1.2.1.2.1.1.1.1 11-1.2.1.2.1 12-1.2.1.2.1.2.1.1 14-1.2.1.2.1.2.1.1 15-1.2.1.2.1.1 15-1.2.1.2.1.2 16-1.2.1.2.1.1.2 18-1.2.2.1.1.1 21-1.2.2.1.2.1.1.1 23-1.2.2.1.2.1.1 25-1.2.2.1.2.1.2.1.1 29-1.2.2.1.2.1.2.2.1 33-1.2.2.1.2.1.2 33-1.2.2.1.2.1.2.2 33-1.2.2.1.2.1.2.2.r 34-1.2.2.1.2.1.3.1.1 35-1.2.2.1.2.1 36-1.2.2.1 36-1.2.2.2 37-1.2.2 38-1.2.2.2.1.1 41-1.2.2.2.2.1.1.1 43-1.2.2.2.2.1.1 45-1.2.2.2.2.1.3 46-1.2.2.2.2.1.3 47-1.2.2.2.2.1.3 48-1.2.2.2.2.1.3 49-1.2.2.2.2.1.3 50-1.2.2.2.2.1.3 51-1.2.2.2.2.1.3 52-1.2.2.2.2.1.3 53-1.2.2.2.2.1.3 54-1.2.2.1.2.1.3.1.1 55-1.2.2.2.2.1 56-1.2.2.2.2.1 58-1.4.1 (d / determine-01~e.2 :ARG0 (w / we~e.0) :ARG1 (t / timecourse :poss (s / small-molecule :name (n2 / name :op1 "SCH772984"~e.8) :ARG2-of (t2 / treat-04 :ARG1 (a / and~e.11 :op1 (p / pathway~e.15 :name (n4 / name :op1 "MAPK"~e.10) :ARG0-of (s2 / signal-07~e.16)) :op2 (p2 / pathway~e.15 :name (n5 / name :op1 "PI3K/AKT"~e.12,14) :ARG0-of s2)))) :beneficiary (a2 / and~e.37 :op1 (c2 / cell-line~e.36 :name (n6 / name :op1 "M238"~e.18) :ARG2-of (m / mean-01 :ARG1 (c3 / cell-line~e.35 :ARG0-of (s3 / sensitive-03~e.23 :ARG1 s~e.21) :mod (g / gene~e.33 :name (n7 / name :op1 "BRAF"~e.25) :ARG2-of~e.33 (m3 / mutate-01~e.33 :value "V600E"~e.29)) :mod (m2 / medical-condition :name (n3 / name :op1 "melanoma"~e.34,54))))) :op2 (c5 / cell-line~e.36 :name (n8 / name :op1 "M233"~e.38) :ARG1-of (m4 / mean-01 :ARG2 (c4 / cell-line~e.55,56 :ARG0-of (r / resist-01~e.43 :ARG1 s~e.41) :mod m2 :mod g~e.45,46,47,48,49,50,51,52,53))))) :time~e.4 (n / next~e.1) :ARG1-of (d3 / describe-01 :ARG0 (f / figure~e.58 :mod "1B"))) # ::id a_pmid_2514_2146.81 ::amr-annotator SDL-AMR-09 ::preferred # ::tok For both M233 and M238 , treatment with 500 nM SCH772984 inhibited pRSK , a known ERK1 @/@ 2 downstream target , as well as pERK1 @/@ 2 itself . # ::alignments 2-1.1.1.1.1.1 3-1.1.1 4-1.1.1.2.1.1 6-1.1 7-1.1.2.r 8-1.1.2.2.1 9-1.1.2.2.2 10-1.1.2.1.1 11-1 12-1.2.1.1.1 12-1.2.1.2 15-1.2.1.3.2 16-1.2.1.3.1.1.1 18-1.2.1.3.1.1.1 19-1.2.1.3.3 20-1.2.1 20-1.2.1.3 20-1.2.1.3.r 22-1.2 23-1.2 24-1.2 27-1.2.1.3.1.1.1 27-1.2.2.1.1 (i / inhibit-01~e.11 :ARG0 (t / treat-04~e.6 :ARG1 (a / and~e.3 :op1 (c / cell-line :name (n2 / name :op1 "M233"~e.2)) :op2 (c2 / cell-line :name (n3 / name :op1 "M238"~e.4))) :ARG2~e.7 (s / small-molecule :name (n / name :op1 "SCH772984"~e.10) :quant (c3 / concentration-quantity :quant 500~e.8 :unit (n4 / nanomolar~e.9)))) :ARG1 (a2 / and~e.22,23,24 :op1 (e / enzyme~e.20 :name (n5 / name :op1 "RSK"~e.12) :ARG3-of (p / phosphorylate-01~e.12) :ARG1-of~e.20 (t2 / target-01~e.20 :ARG0 (e3 / enzyme :name (n7 / name :op1 "ERK1/2"~e.16,18,27)) :ARG1-of (k / know-01~e.15) :location (d / downstream~e.19))) :op2 (e2 / enzyme :name (n6 / name :op1 "ERK1/2"~e.27) :ARG3-of p))) # ::id a_pmid_2514_2146.82 ::amr-annotator SDL-AMR-09 ::preferred # ::tok For the resistant M233 , the MAPK inhibition was strong as early as 1 hour post treatment , with decreased pERK and near @-@ complete disappearance of pRSK . # ::alignments 2-1.1.1.1.2 2-1.1.1.1.2.2 2-1.1.1.1.2.2.r 3-1.1.1.1.2.1.1 6-1.1.1.1.1.1 7-1.1.1 8-1.1.1.r 9-1.1 10-1.1.2.r 12-1.1.2.r 13-1.1.2.2.1 14-1.1.2.2.2 15-1.1.2 16-1.1.2.1 19-1.2 20-1.2.1.1.1 20-1.2.1.2 21-1 22-1.3.2.1 24-1.3.2 25-1.3 27-1.2.1.2 27-1.3.1.1.1 (a2 / and~e.21 :op1 (s / strong~e.9 :domain~e.8 (i / inhibit-01~e.7 :ARG1 (p / protein-family :name (n / name :op1 "MAPK"~e.6) :part-of (c / cell-line~e.2 :name (n2 / name :op1 "M233"~e.3) :ARG0-of~e.2 (r / resist-01~e.2)))) :time~e.10,12 (a / after~e.15 :op1 (t / treat-04~e.16) :quant (t2 / temporal-quantity :quant 1~e.13 :unit (h / hour~e.14)))) :op2 (d / decrease-01~e.19 :ARG1 (e / enzyme :name (n3 / name :op1 "ERK"~e.20) :ARG3-of (p2 / phosphorylate-01~e.20,27) :part-of c)) :op3 (d2 / disappear-01~e.25 :ARG1 (e2 / enzyme :name (n5 / name :op1 "RSK"~e.27) :ARG3-of p2 :part-of c) :ARG1-of (c2 / complete-01~e.24 :degree (n4 / near~e.22)))) # ::id a_pmid_2514_2146.83 ::amr-annotator SDL-AMR-09 ::preferred # ::tok However , between 12 and 24 hours we observe a rebound in the pathway with a return to baseline pERK1 @/@ 2 levels and an induction in pMEK above baseline levels by 24 hours . # ::alignments 0-1 2-1.1.3 3-1.1.3.1.1 4-1.1.3 5-1.1.3.2.1 6-1.1.3.1.2 7-1.1.1 8-1.1 10-1.1.2 11-1.1.2.1.r 13-1.1.2.1 16-1.1.2.2.1.1 18-1.1.2.2.1.1.1.2 21-1.1.2.2.1.1.1.1.1.1 22-1.1.2.2.1.1.1 23-1.1.2.2.1 25-1.1.2.2.1.2 28-1.1.2.2.1.2.2 29-1.1.2.2.1.2.2.1.2 30-1.1.2.2.1.1.1 30-1.1.2.2.1.2.2.1 32-1.1.2.2.1.3.1.1 32-1.1.3.2.1 33-1.1.2.2.1.3.1.2 (h3 / have-concession-91~e.0 :ARG2 (o / observe-01~e.8 :ARG0 (w / we~e.7) :ARG1 (r / rebound-01~e.10 :ARG1~e.11 (p / pathway~e.13) :ARG0-of (c2 / cause-01 :ARG1 (a / and~e.23 :op1 (r2 / return-01~e.16 :ARG1 (l / level~e.22,30 :quant-of (e / enzyme :name (n / name :op1 "ERK1/2"~e.21) :ARG3-of (p2 / phosphorylate-01)) :mod (b2 / baseline~e.18))) :op2 (i / induce-01~e.25 :ARG1 (e2 / enzyme :name (n2 / name :op1 "MEK") :ARG3-of p2) :ARG2 (a2 / above~e.28 :op1 (l2 / level~e.30 :quant-of e2 :mod b2~e.29))) :time (a3 / after :quant (t3 / temporal-quantity :quant 24~e.32 :unit (h2 / hour~e.33)))))) :time (b / between~e.2,4 :op1 (t / temporal-quantity :quant 12~e.3 :unit (h / hour~e.6)) :op2 (t2 / temporal-quantity :quant 24~e.5,32 :unit h)))) # ::id a_pmid_2514_2146.84 ::amr-annotator SDL-AMR-09 ::preferred # ::tok Little change in pAKT was seen at any timepoint up to 24 hours , though a mild induction was seen at 48 hours . # ::alignments 0-1.1.2 1-1.1 2-1.1.1.r 3-1.1.1.1.1 3-1.1.1.2 5-1 6-1.2.r 7-1.2.1 8-1.2 9-1.2.2 10-1.2.2 11-1.2.2.1.1 12-1.2.2.1.2 14-1.3.r 16-1.3.1.2 17-1.3.1 19-1.3 21-1.3.2.1.1 22-1.3.2.1.2 (s / see-01~e.5 :ARG1 (c / change-01~e.1 :ARG1~e.2 (e / enzyme :name (n / name :op1 "AKT"~e.3) :ARG3-of (p / phosphorylate-01~e.3)) :mod (l / little~e.0)) :time~e.6 (t / timepoint~e.8 :mod (a / any~e.7) :quant (u / up-to~e.9,10 :op1 (t2 / temporal-quantity :quant 24~e.11 :unit (h / hour~e.12)))) :concession~e.14 (s2 / see-01~e.19 :ARG1 (i / induce-01~e.17 :ARG2 e :degree (m / mild~e.16)) :time (a2 / after :op1 (t3 / temporal-quantity :quant 48~e.21 :unit h~e.22)))) # ::id a_pmid_2514_2146.85 ::amr-annotator SDL-AMR-09 ::preferred # ::tok For the sensitive M238 , pRSK levels also decreased as early as 1 hour and levels continued to decrease thereafter . # ::alignments 2-1.1.1.1.3 2-1.1.1.1.3.2 2-1.1.1.1.3.2.r 3-1.1.1.1.3.1.1 5-1.1.1.1.1.1 5-1.1.1.1.2 6-1.1.1 7-1.1.2 8-1.1 9-1.1.3.r 11-1.1.3.r 12-1.1.3.1.1 13-1.1.3.1.2 15-1.1.1 16-1.2 18-1.2.1 19-1.2.2 (a / and :op1 (d / decrease-01~e.8 :ARG1 (l / level~e.6,15 :quant-of (e / enzyme :name (n / name :op1 "RSK"~e.5) :ARG3-of (p / phosphorylate-01~e.5) :part-of (c / cell-line~e.2 :name (n2 / name :op1 "M238"~e.3) :ARG0-of~e.2 (s / sensitive-03~e.2)))) :mod (a2 / also~e.7) :time~e.9,11 (a3 / after :op1 (t / temporal-quantity :quant 1~e.12 :unit (h / hour~e.13)))) :op2 (c2 / continue-01~e.16 :ARG1 (d2 / decrease-01~e.18 :ARG1 l) :time (t2 / thereafter~e.19))) # ::id a_pmid_2514_2146.86 ::amr-annotator SDL-AMR-09 ::preferred # ::tok Meanwhile , pERK1 @/@ 2 remained suppressed through 24 hours . # ::alignments 0-1.4 4-1.1.1.1 5-1 6-1.2 8-1.3.1 9-1.3.2 (r / remain-01~e.5 :ARG1 (e / enzyme :name (n / name :op1 "ERK1/2"~e.4) :ARG3-of (p / phosphorylate-01)) :ARG3 (s / suppress-01~e.6 :ARG1 e) :duration (t / temporal-quantity :quant 24~e.8 :unit (h / hour~e.9)) :time (m / meanwhile~e.0)) # ::id a_pmid_2514_2146.87 ::amr-annotator SDL-AMR-09 ::preferred # ::tok By 48 hours , pERK1 @/@ 2 levels increased , though at reduced levels compared to baseline . # ::alignments 1-1.2.1.1 2-1.2.1.2 6-1.1.1.1.1 7-1.1 7-1.3 8-1 10-1.3.r 12-1.3.1 13-1.3 14-1.3.1.1 15-1.3.1.1.1.r 16-1.3.1.1.1 (i / increase-01~e.8 :ARG1 (l / level~e.7 :quant-of (e / enzyme :name (n / name :op1 "ERK1/2"~e.6) :ARG3-of (p / phosphorylate-01))) :time (a / after :op1 (t / temporal-quantity :quant 48~e.1 :unit (h / hour~e.2))) :concession~e.10 (l2 / level~e.7,13 :ARG1-of (r / reduce-01~e.12 :ARG1-of (c / compare-01~e.14 :ARG2~e.15 (b / baseline~e.16))))) # ::id a_pmid_2514_2146.88 ::amr-annotator SDL-AMR-09 ::preferred # ::tok Concomitant with this , pMEK levels remained unchanged until 24 hours and increased further by 48 hours . # ::alignments 0-1.3 1-1.3.1.r 2-1.3.1 5-1.1.1 6-1.1 7-1.1.2 7-1.1.2.1 7-1.1.2.1.r 8-1.1.3 9-1.1.3.1.1 10-1.1.3.1.2 11-1 12-1.2 13-1.2.3 15-1.2.2.1.1 16-1.2.2.1.2 (a / and~e.11 :op1 (r / remain-01~e.6 :ARG1 (l / level~e.5 :quant-of (e / enzyme :name (n / name :op1 "MEK") :ARG3-of (p / phosphorylate-01))) :ARG3 (c / change-01~e.7 :polarity~e.7 -~e.7 :ARG1 l) :time (u / until~e.8 :op1 (t / temporal-quantity :quant 24~e.9 :unit (h / hour~e.10)))) :op2 (i / increase-01~e.12 :ARG1 l :time (a2 / after :op1 (t2 / temporal-quantity :quant 48~e.15 :unit h~e.16)) :degree (f / further~e.13)) :manner (c2 / concomitant~e.0 :prep-with~e.1 (t3 / this~e.2))) # ::id a_pmid_2514_2146.89 ::amr-annotator SDL-AMR-09 ::preferred # ::tok Regarding pAKT , an early induction at 1 hour occurred , followed by decreases thereafter though never becoming completely suppressed even at 48 hours . # ::alignments 1-1.1.1.1 1-1.1.2 4-1.2 5-1 7-1.3.1.1 8-1.3.1.2 11-1.3 11-1.4 11-1.4.1.3.3.2 12-1.4.1.r 13-1.4.1 14-1.4.1.2 15-1.4.1.3.r 16-1.4.1.3.1 16-1.4.1.3.1.r 16-1.4.1.3.4 17-1.4.1.3 18-1.4.1.3.3.4 19-1.4.1.3.3 20-1.4.1.3.3.3 21-1.4.1.3.3.2.1.r 22-1.4.1.3.3.2.1.1 23-1.4.1.3.3.2.1.2 (i / induce-01~e.5 :ARG2 (e2 / enzyme :name (n / name :op1 "AKT"~e.1) :ARG3-of (p / phosphorylate-01~e.1)) :time (e / early~e.4) :time (a / after~e.11 :op1 (t / temporal-quantity :quant 1~e.7 :unit (h / hour~e.8))) :ARG2-of (f / follow-01~e.11 :ARG1~e.12 (d / decrease-01~e.13 :ARG1 e :time (t2 / thereafter~e.14) :concession~e.15 (b / become-01~e.17 :polarity~e.16 -~e.16 :ARG1 e2 :ARG2 (s / suppress-01~e.19 :ARG1 e2 :time (a2 / after~e.11 :op1~e.21 (t3 / temporal-quantity :quant 48~e.22 :unit h~e.23)) :mod (e4 / even~e.20) :degree (c / complete~e.18)) :time (e3 / ever~e.16))))) # ::id a_pmid_2514_2146.90 ::amr-annotator SDL-AMR-09 ::preferred # ::tok In both cell lines , pRSK remained blocked at all timepoints , demonstrating ongoing , potent inhibition of ERK1 @/@ 2 activity by SCH772984 . # ::alignments 1-1.3.1 2-1.3 3-1.3 5-1.1.1.1 6-1 7-1.2 8-1.2.2.r 9-1.2.2.2.1 12-1.2.2 13-1.2.2.1.3 15-1.2.2.1.4 16-1.2.2.1 17-1.2.2.1.2.r 18-1.2.2.1.2.1.1.1 20-1.2.2.1.2.1.1.1 21-1.2.2.1.2 22-1.2.2.1.1.r 23-1.2.2.1.1.1.1 (r / remain-01~e.6 :ARG1 (p / protein :name (n / name :op1 "pRSK"~e.5)) :ARG3 (b / block-01~e.7 :ARG1 p :ARG0-of~e.8 (d / demonstrate-01~e.12 :ARG1 (i / inhibit-01~e.16 :ARG0~e.22 (s / small-molecule :name (n3 / name :op1 "SCH772984"~e.23)) :ARG1~e.17 (a2 / activity-06~e.21 :ARG0 (e / enzyme :name (n2 / name :op1 "ERK1/2"~e.18,20))) :ARG1-of (g / go-on-15~e.13) :mod (p2 / potent~e.15)) :time (t / timepoint :mod (a / all~e.9)))) :location (c2 / cell-line~e.2,3 :mod (b2 / both~e.1))) # ::id a_pmid_2514_2146.91 ::amr-annotator SDL-AMR-09 ::preferred # ::tok These data supports that the distinction between sensitive and resistant cell lines could be best made based on pERK recovery at 24 hours , as recovery of the feedback loop that restores MAPK activity occurred by 24 hours in the resistant cell line whereas the sensitive cell line required longer than 24 hours . # ::alignments 0-1.1.1 1-1.1 2-1 3-1.2.r 5-1.2.1.1 7-1.2.1.1.1.1 7-1.2.1.1.1.1.1 7-1.2.1.1.1.1.1.r 8-1.2.1.1.1 9-1.2.1.1.1.2.1 10-1.2.1.1.1.2 11-1.2.1.1.1.2 12-1.2 14-1.2.1.3 14-1.2.1.3.1 14-1.2.1.3.1.r 16-1.2.1 17-1.2.1.2.r 18-1.2.1.2.1.1.1 18-1.2.1.2.1.2 19-1.2.1.2 21-1.2.1.2.2.1.1 22-1.2.1.2.2.1.2 24-1.2.1.2.2.r 25-1.2.1.2.3.1 26-1.2.1.2.3.1.1.r 28-1.2.1.2.3.1.1.1 29-1.2.1.2.3.1.1 31-1.2.1.2.3.1.1.2 32-1.2.1.2.3.1.1.2.1.1.1.1 33-1.2.1.2.3.1.1.2.1 35-1.2.1.2.3.1.1.2.4.r 36-1.2.1.2.3.1.1.2.4 37-1.2.1.2.3.1.1.2.4 38-1.2.1.2.3.1.1.2.2.r 40-1.2.1.2.3.1.1.2.2 41-1.2.1.2.3.1.1.2.2 42-1.2.1.2.3.1.1.2.2 43-1.2.1.2.3.1.1.2.3 45-1.2.1.2.3.1.1.2.3.1.1 46-1.2.1.2.3.1.1.2.3.1.1 47-1.2.1.2.3.1.1.2.3.1.1 48-1.2.1.2.3.1.1.2.3.1 49-1.2.1.2.3.1.1.2.3.1.2 49-1.2.1.2.3.1.1.2.3.1.2.2 49-1.2.1.2.3.1.1.2.3.1.2.2.r 51-1.2.1.2.3.1.1.2.3.1.2.1 52-1.2.1.2.3.1.1.2.3.1.2.1 (s / support-01~e.2 :ARG0 (d2 / data~e.1 :mod (t2 / this~e.0)) :ARG1~e.3 (p / possible-01~e.12 :ARG1 (b / base-02~e.16 :ARG1 (d / distinguish-01~e.5 :ARG1 (a / and~e.8 :op1 (c / cell-line~e.7 :ARG0-of~e.7 (s2 / sensitive-03~e.7)) :op2 (c2 / cell-line~e.10,11 :ARG0-of (r / resist-01~e.9)))) :ARG2~e.17 (r2 / recover-02~e.19 :ARG0 (e / enzyme :name (n2 / name :op1 "ERK"~e.18) :ARG3-of (p3 / phosphorylate-01~e.18)) :time~e.24 (a3 / after :op1 (t / temporal-quantity :quant 24~e.21 :unit (h / hour~e.22))) :ARG1-of (m3 / mean-01 :ARG2 (r5 / recover-02~e.25 :ARG0~e.26 (l / loop~e.29 :mod (f / feedback~e.28) :ARG0-of (r3 / restore-01~e.31 :ARG1 (a2 / activity-06~e.33 :ARG0 (p2 / pathway :name (n / name :op1 "MAPK"~e.32))) :location~e.38 c2~e.40,41,42 :ARG1-of (c3 / contrast-01~e.43 :ARG2 (r4 / require-01~e.48 :ARG0 c~e.45,46,47 :ARG1 (l2 / long-03~e.49 :ARG2 t~e.51,52 :degree~e.49 (m2 / more~e.49)))) :time~e.35 a3~e.36,37))))) :ARG1-of (g / good-02~e.14 :degree~e.14 (m / most~e.14))))) # ::id a_pmid_2514_2146.92 ::amr-annotator SDL-AMR-09 ::preferred # ::tok Therefore , for subsequent analyses , we selected 24 hours as the optimal timepoint to compare signaling in our cell lines . # ::alignments 0-1 2-1.1.r 3-1.1.5.2 3-1.1.5.2.r 4-1.1.5 6-1.1.1 7-1.1 8-1.1.2.1 9-1.1.2.2 10-1.1.3.r 10-1.1.5.2.r 12-1.1.3.1 13-1.1.3 15-1.1.4 16-1.1.4.2 17-1.1.4.2.1.r 18-1.1.4.2.1.1 18-1.1.4.2.1.1.r 19-1.1.4.2.1 20-1.1.4.2.1 (c / cause-01~e.0 :ARG1~e.2 (s / select-01~e.7 :ARG0 (w / we~e.6) :ARG1 (t / temporal-quantity :quant 24~e.8 :unit (h / hour~e.9)) :ARG3~e.10 (t2 / timepoint~e.13 :mod (o / optimal~e.12)) :purpose (c2 / compare-01~e.15 :ARG0 w :ARG1 (s2 / signal-07~e.16 :location~e.17 (c3 / cell-line~e.19,20 :poss~e.18 w~e.18))) :purpose (a / analyze-01~e.4 :ARG0 w :time~e.3,10 (s3 / subsequent~e.3)))) # ::id a_pmid_2514_2146.93 ::amr-annotator SDL-AMR-09 ::preferred # ::tok As shown in Figure 1 @ C , three BRAF mutant cell lines representative of the different sensitivity groups to SCH772984 were profiled in terms of downstream signaling inhibition at 24 hours : a highly sensitive cell line ( M262 , BRAF @ V600E @ ) , an intermediately sensitive cell line ( M409 , BRAF @ V600E @ ) , and a resistant cell line ( M381 , BRAF @ V600R @ ) . # ::alignments 0-1.2.2.r 1-1.3 2-1.3.1.r 3-1.3.1 9-1.1.1 11-1.1.4.1.1 13-1.1.3.1.1.2.1.2 13-1.1.3.1.1.2.1.2.2 13-1.1.3.1.1.2.1.2.2.r 13-1.1.3.1.3.2.1.2 13-1.1.3.1.3.2.1.2.2 13-1.1.3.1.3.2.1.2.2.r 13-1.1.4 13-1.1.4.2 13-1.1.4.2.r 14-1.1 15-1.1 16-1.1.2 17-1.1.2.1.r 19-1.1.2.1.1 20-1.1.2.1.2 21-1.1.2.1 22-1.1.2.1.2.1.r 23-1.1.2.1.2.1.1.1 25-1 29-1.2.1.1 30-1.2.1 31-1.2 33-1.2.2.1.1 34-1.2.2.1.2 37-1.1.3.1.1.1.1 38-1.1.3.1.1.1 39-1.1.3.1.1 40-1.1.3.1.1 42-1.1.3.1.1.2.1.1.1 45-1.1.3.1.1.2.1.2.1.1 49-1.1.3.1.1.2.1.2.2.1 55-1.1.3.1.2.1.1 56-1.1.3.1.2.1 57-1.1.3.1.2 58-1.1.3.1.2 60-1.1.3.1.2.2.1.1.1 63-1.1.3.1.2.2.1.2 64-1.1.3.1.2.2.1.2 65-1.1.3.1.2.2.1.2 66-1.1.3.1.2.2.1.2 67-1.1.3.1.2.2.1.2 72-1.1.3.1 74-1.1.3.1.3.1 75-1.1.3.1.3 76-1.1.3.1.3 78-1.1.3.1.3.2.1.1.1 81-1.1.3.1.3.2.1.2.1.1 85-1.1.3.1.3.2.1.2.2.1 (p2 / profile-01~e.25 :ARG1 (c / cell-line~e.14,15 :quant 3~e.9 :ARG0-of (r2 / represent-01~e.16 :ARG1~e.17 (g / group~e.21 :ARG1-of (d2 / differ-02~e.19) :ARG0-of (s / sensitive-03~e.20 :ARG1~e.22 (s6 / small-molecule :name (n2 / name :op1 "SCH772984"~e.23))))) :ARG1-of (m7 / mean-01 :ARG2 (a / and~e.72 :op1 (c3 / cell-line~e.39,40 :ARG0-of (s3 / sensitive-03~e.38 :ARG1-of (h2 / high-02~e.37)) :ARG1-of (m2 / mean-01 :ARG2 (c4 / cell-line :name (n3 / name :op1 "M262"~e.42) :mod (g2 / gene~e.13 :name (n4 / name :op1 "BRAF"~e.45) :ARG2-of~e.13 (m3 / mutate-01~e.13 :value "V600E"~e.49))))) :op2 (c5 / cell-line~e.57,58 :ARG0-of (s4 / sensitive-03~e.56 :mod (i2 / intermediate~e.55)) :ARG1-of (m4 / mean-01 :ARG2 (c6 / cell-line :name (n5 / name :op1 "M409"~e.60) :mod g2~e.63,64,65,66,67))) :op3 (c7 / cell-line~e.75,76 :ARG0-of (r4 / resist-01~e.74) :ARG1-of (m5 / mean-01 :ARG2 (c8 / cell-line :name (n6 / name :op1 "M381"~e.78) :mod (g4 / gene~e.13 :name (n7 / name :op1 "BRAF"~e.81) :ARG2-of~e.13 (m6 / mutate-01~e.13 :value "V600R"~e.85))))))) :mod (g3 / gene~e.13 :name (n / name :op1 "BRAF"~e.11) :ARG2-of~e.13 (m / mutate-01~e.13))) :topic (i / inhibit-01~e.31 :ARG1 (s2 / signal-07~e.30 :location (d / downstream~e.29)) :time~e.0 (a2 / after :op1 (t / temporal-quantity :quant 24~e.33 :unit (h / hour~e.34)))) :ARG1-of (s5 / show-01~e.1 :ARG0~e.2 (f / figure~e.3 :mod "1C"))) # ::id a_pmid_2514_2146.94 ::amr-annotator SDL-AMR-09 ::preferred # ::tok For M262 , treatment with SCH772984 resulted in disappearance of pRSK , disappearance of pERK1 @/@ 2 , decrease in pAKT , and slight induction of pMEK at 24 hours . # ::alignments 1-1.1.1.1.1 3-1.1 4-1.1.2.r 5-1.1.2.1.1 6-1 7-1.2.r 8-1.2.1 10-1.2.1.1.1.1 10-1.2.1.1.2 12-1.2.1 12-1.2.2 16-1.2.2.1.1.1 18-1.2.3 20-1.2.1.1.2 20-1.2.3.1.1.1 22-1.2 23-1.2.4.2 24-1.2.4 28-1.2.5.1.1 29-1.2.5.1.2 (r / result-01~e.6 :ARG1 (t2 / treat-04~e.3 :ARG1 (c / cell-line :name (n / name :op1 "M262"~e.1)) :ARG2~e.4 (s / small-molecule :name (n2 / name :op1 "SCH772984"~e.5))) :ARG2~e.7 (a / and~e.22 :op1 (d / disappear-01~e.8,12 :ARG1 (e4 / enzyme :name (n3 / name :op1 "RSK"~e.10) :ARG3-of (p2 / phosphorylate-01~e.10,20))) :op2 (d2 / disappear-01~e.12 :ARG1 (e / enzyme :name (n4 / name :op1 "ERK1/2"~e.16) :ARG3-of p2)) :op3 (d3 / decrease-01~e.18 :ARG1 (e2 / enzyme :name (n5 / name :op1 "AKT"~e.20) :ARG3-of p2)) :op4 (i / induce-01~e.24 :ARG2 (e3 / enzyme :name (n6 / name :op1 "MEK") :ARG3-of p2) :degree (s2 / slight~e.23)) :time (a2 / after :op1 (t / temporal-quantity :quant 24~e.28 :unit (h / hour~e.29))))) # ::id a_pmid_2514_2146.95 ::amr-annotator SDL-AMR-09 ::preferred # ::tok M409 had a similar cell signaling profile as M262 , consistent with its modest sensitivity . # ::alignments 0-1.1.1.1 1-1 3-1.2.2 4-1.2.1.1 5-1.2.1 6-1.2 7-1.2.2.1.r 8-1.2.2.1.1.1 10-1.2.3 11-1.2.3.1.r 12-1.2.3.1.1 12-1.2.3.1.1.r 13-1.2.3.1.2 14-1.2.3.1 (h / have-03~e.1 :ARG0 (c / cell-line :name (n / name :op1 "M409"~e.0)) :ARG1 (p / profile-01~e.6 :ARG0-of (s / signal-07~e.5 :ARG1 (c2 / cell~e.4)) :ARG1-of (r / resemble-01~e.3 :ARG2~e.7 (c3 / cell-line :name (n2 / name :op1 "M262"~e.8))) :ARG1-of (c4 / consistent-01~e.10 :ARG2~e.11 (s2 / sensitive-03~e.14 :ARG0~e.12 p~e.12 :mod (m / modest~e.13))))) # ::id a_pmid_2514_2146.96 ::amr-annotator SDL-AMR-09 ::preferred # ::tok For M381 induction of pMEK and pERK1 @/@ 2 were seen with no change in pRSK at 24 h . # ::alignments 1-1.1.2.1.1 2-1.1 5-1.1.1 8-1.1.1.2.1.1 10-1 11-1.2.r 12-1.2.1 12-1.2.1.r 13-1.2 15-1.1.1.1.2 15-1.2.2.1.1 17-1.3.1.1 18-1.3.1.2 (s / see-01~e.10 :ARG1 (i / induce-01~e.2 :ARG2 (a2 / and~e.5 :op1 (e / enzyme :name (n / name :op1 "MEK") :ARG3-of (p / phosphorylate-01~e.15)) :op2 (e2 / enzyme :name (n2 / name :op1 "ERK1/2"~e.8) :ARG3-of p)) :location (c2 / cell-line :name (n4 / name :op1 "M381"~e.1))) :ARG2~e.11 (c / change-01~e.13 :polarity~e.12 -~e.12 :ARG1 (e3 / enzyme :name (n3 / name :op1 "RSK"~e.15) :ARG3-of p)) :time (a / after :op1 (t / temporal-quantity :quant 24~e.17 :unit (h / hour~e.18)))) # ::id a_pmid_2514_2146.97 ::amr-annotator SDL-AMR-09 ::preferred # ::tok To determine the effect of SCH772984 on the PI3K @/@ AKT pathway , we first evaluated the baseline pAKT levels for a group of cell lines ( Additional file 2 @ : Figure S2 ) . # ::alignments 1-1.3 3-1.3.2 4-1.3.2.1.r 5-1.3.2.1.1.1 6-1.3.2.2.r 8-1.3.2.2.1.1 10-1.3.2.2.1.1 11-1.3.2.2 13-1.1 14-1.5 15-1 17-1.2.2 18-1.2.1.1.1 18-1.2.1.2 19-1.2 22-1.2.1.3 23-1.2.1.3.1.r 24-1.2.1.3.1 25-1.2.1.3.1 28-1.4.1 30-1.4.1.1 33-1.4.1.2.1 34-1.4.1.2.1.1 (e / evaluate-01~e.15 :ARG0 (w / we~e.13) :ARG1 (l / level~e.19 :quant-of (e2 / enzyme :name (n2 / name :op1 "AKT"~e.18) :ARG3-of (p2 / phosphorylate-01~e.18) :part-of (g / group~e.22 :consist-of~e.23 (c / cell-line~e.24,25))) :mod (b / baseline~e.17)) :purpose (d / determine-01~e.1 :ARG0 w :ARG1 (a / affect-01~e.3 :ARG0~e.4 (s / small-molecule :name (n3 / name :op1 "SCH772984"~e.5)) :ARG1~e.6 (p / pathway~e.11 :name (n / name :op1 "PI3K/AKT"~e.8,10)))) :ARG1-of (d2 / describe-01 :ARG0 (f / file~e.28 :mod 2~e.30 :ARG1-of (m / mean-01 :ARG2 (f2 / figure~e.33 :mod "S2"~e.34)) :ARG1-of (a3 / add-02))) :time (f3 / first~e.14)) # ::id a_pmid_2514_2146.98 ::amr-annotator SDL-AMR-09 ::preferred # ::tok We found a weak correlation with the activity of the PI3K @/@ AKT pathway and sensitivity to SCH772984 for BRAF mutants . # ::alignments 0-1.1 1-1 3-1.2.1.3 4-1.2.1 5-1.2.1.2.r 7-1.2.1.2 8-1.2.1.2.1.r 10-1.2.1.2.1.1.1 12-1.2.1.2.1.1.1 13-1.2.1.2.1 14-1.2 15-1.2.2 16-1.2.2.2.r 17-1.2.2.2.1.1 18-1.2.2.1.r 19-1.2.2.1.1.1 20-1.2.2.1 20-1.2.2.1.2 20-1.2.2.1.2.r (f / find-01~e.1 :ARG0 (w / we~e.0) :ARG1 (a2 / and~e.14 :op1 (c / correlate-01~e.4 :ARG1 e :ARG2~e.5 (a / activity-06~e.7 :ARG0~e.8 (p / pathway~e.13 :name (n / name :op1 "PI3K/AKT"~e.10,12))) :ARG1-of (w2 / weak-02~e.3)) :op2 (s / sensitive-03~e.15 :ARG0~e.18 (e / enzyme~e.20 :name (n2 / name :op1 "BRAF"~e.19) :ARG2-of~e.20 (m / mutate-01~e.20)) :ARG1~e.16 (s2 / small-molecule :name (n3 / name :op1 "SCH772984"~e.17))))) # ::id a_pmid_2514_2146.99 ::amr-annotator SDL-AMR-09 ::preferred # ::tok For example , M238 and M409 are two clear examples of cell lines with low levels of pAKT related to sensitivity to SCH772984 . # ::alignments 0-1.3 1-1.3 1-1.5 3-1.1.1.1 4-1 5-1.2.1.1 7-1.3.1 8-1.3.3 9-1.3 10-1.3.2.r 11-1.3.2 12-1.3.2 14-1.3.2.1.1.2 15-1.3.2.1.1 16-1.3.2.1.1.1.r 17-1.3.2.1.1.1.1.1 17-1.3.2.1.1.1.2 18-1.4 19-1.4.1.r 20-1.4.1 21-1.4.1.1.r 22-1.4.1.1.1.1 (a2 / and~e.4 :op1 (c / cell-line :name (n / name :op1 "M238"~e.3)) :op2 (c2 / cell-line :name (n2 / name :op1 "M409"~e.5)) :ARG0-of (e2 / exemplify-01~e.0,1,9 :quant 2~e.7 :ARG1~e.10 (c3 / cell-line~e.11,12 :ARG0-of (h / have-03 :ARG1 (l / level~e.15 :quant-of~e.16 (e3 / enzyme :name (n3 / name :op1 "AKT"~e.17) :ARG3-of (p / phosphorylate-01~e.17)) :ARG1-of (l2 / low-04~e.14)))) :ARG1-of (c4 / clear-06~e.8)) :ARG1-of (r / relate-01~e.18 :ARG2~e.19 (s / sensitive-03~e.20 :ARG1~e.21 (s2 / small-molecule :name (n4 / name :op1 "SCH772984"~e.22)))) :ARG0-of (e / exemplify-01~e.1)) # ::id a_pmid_2514_2146.100 ::amr-annotator SDL-AMR-09 ::preferred # ::tok For both of them , ERK inhibition with SCH772984 was accompanied by an upregulation of pAKT levels even at 24 hours treatment ( Figure 1 @ B ) . # ::alignments 1-1.4.1 3-1.4 5-1.2.2.1.1 6-1.2 7-1.2.1.r 8-1.2.1.1.1 10-1 11-1.1.r 13-1.1 14-1.1.1.r 15-1.1.1.1.1.1 15-1.1.1.1.2 16-1.1.1 17-1.3.2 19-1.3.3.1 20-1.3.3.2 21-1.3.1 23-1.5.1 (a / accompany-01~e.10 :ARG0~e.11 (u / upregulate-01~e.13 :ARG1~e.14 (l / level~e.16 :quant-of (e3 / enzyme :name (n4 / name :op1 "AKT"~e.15) :ARG3-of (p / phosphorylate-01~e.15)))) :ARG1 (i / inhibit-01~e.6 :ARG0~e.7 (s / small-molecule :name (n3 / name :op1 "SCH772984"~e.8)) :ARG1 (e2 / enzyme :name (n2 / name :op1 "ERK"~e.5))) :time (a2 / after :op1 (t2 / treat-04~e.21) :mod (e / even~e.17) :quant (t / temporal-quantity :quant 24~e.19 :unit (h / hour~e.20))) :beneficiary (t3 / they~e.3 :mod (b / both~e.1)) :ARG1-of (d / describe-01 :ARG0 (f / figure~e.23 :mod "1B"))) # ::id a_pmid_2514_2146.101 ::amr-annotator SDL-AMR-09 ::preferred # ::tok M233 was among the resistant BRAF @ V600E @ melanoma cell lines , which appeared to have increased pAKT at baseline compared to other BRAF mutant cell lines . # ::alignments 0-1.1.1 2-1.3 4-1.4 6-1.3.1.1.1.1 10-1.3.1.1.2.1 13-1.3.1.2.1.1 14-1.3.1 15-1 18-1.2.2 20-1.2 21-1.2.1.3 22-1.2.1.1.1 22-1.2.1.2 23-1.2.1.3.1.r 24-1.2.1.3.1 25-1.2.1.3.2.r 27-1.2.1.3.2.2 29-1.2.1.3.2.1.1.1 31-1.2.1.3.2.1 31-1.2.1.3.2.1.2 31-1.2.1.3.2.1.2.r 31-1.3.1.1 31-1.3.1.1.2 31-1.3.1.1.2.r 32-1.2.1.3.2 33-1.2.1.3.2 (c4 / cell-line~e.15 :name (n5 / name :op1 "M233"~e.0) :ARG0-of (h / have-03~e.20 :ARG1 (e / enzyme :name (n3 / name :op1 "AKT"~e.22) :ARG3-of (p / phosphorylate-01~e.22) :ARG1-of (i2 / increase-01~e.21 :location~e.23 (b / baseline~e.24) :compared-to~e.25 (c3 / cell-line~e.32,33 :mod (g2 / gene~e.31 :name (n4 / name :op1 "BRAF"~e.29) :ARG2-of~e.31 (m3 / mutate-01~e.31)) :mod (o / other~e.27)))) :ARG1-of (a / appear-01~e.18)) :ARG1-of (i / include-91~e.2 :ARG2 (c2 / cell-line~e.14 :mod (g / gene~e.31 :name (n2 / name :op1 "BRAF"~e.6) :ARG2-of~e.31 (m2 / mutate-01~e.31 :value "V600E"~e.10)) :mod (m4 / medical-condition :name (n / name :op1 "melanoma"~e.13)))) :ARG0-of (r / resist-01~e.4)) # ::id a_pmid_2514_2146.102 ::amr-annotator SDL-AMR-09 ::preferred # ::tok Consistent with this , M233 is a PTEN null cell line and has a concomitant AKT1 amplification [ @ 31 @ ] . # ::alignments 0-1.3 1-1.3.1.r 2-1.3.1 4-1.1.3.1.1 5-1.1.3.r 7-1.1.2.1.1 8-1.1.1 9-1.1 10-1.1 11-1 12-1.2 14-1.2.2.2 16-1.2.2.1.1.1 18-1.2.2 21-1.4.1.1.1 (a / and~e.11 :op1 (c2 / cell-line~e.9,10 :mod (n2 / null~e.8) :mod (p / protein :name (n3 / name :op1 "PTEN"~e.7)) :domain~e.5 (c / cell-line :name (n / name :op1 "M233"~e.4))) :op2 (h / have-03~e.12 :ARG0 c :ARG1 (a2 / amplify-01~e.18 :ARG1 (g / gene :name (n4 / name :op1 "AKT1"~e.16)) :mod (c3 / concomitant~e.14))) :ARG1-of (c4 / consistent-01~e.0 :ARG2~e.1 (t / this~e.2)) :ARG1-of (d / describe-01 :ARG0 (p2 / publication :ARG1-of (c5 / cite-01 :ARG2 31~e.21)))) # ::id a_pmid_2514_2146.103 ::amr-annotator SDL-AMR-09 ::preferred # ::tok After treatment with SCH772984 ( Figure 1 @ B ) , these levels stay constant , indicating that dual inhibition with SCH772984 and AKT @/@ mTOR inhibitors may be a useful strategy . # ::alignments 0-1.4 1-1.4.1 2-1.4.1.1.r 3-1.4.1.1 5-1.4.2.1 12-1.1.1 13-1.1 14-1 15-1.2 17-1.3 18-1.3.1.r 19-1.3.1.1.1.2 20-1.3.1.1.1 22-1.3.1.1.1.1.1.1 23-1.3.1.1 24-1.3.1.1.2.1.1.1.1 26-1.3.1.1.2.1.1.1.1 27-1.3.1.1.1 27-1.3.1.1.2 27-1.3.1.1.2.1 27-1.3.1.1.2.1.r 28-1.3.1.3 29-1.3.1.1.r 31-1.3.1.2 32-1.3.1 (s / stay-01~e.14 :ARG1 (l / level~e.13 :mod (t2 / this~e.12)) :ARG3 (c / constant~e.15) :ARG0-of (i2 / indicate-01~e.17 :ARG1~e.18 (s2 / strategy~e.32 :domain~e.29 (a / and~e.23 :op1 (i3 / inhibit-01~e.20,27 :ARG0 (s3 / small-molecule :name (n / name :op1 "SCH772984"~e.22)) :mod (d2 / dual~e.19)) :op2 (m / molecular-physical-entity~e.27 :ARG0-of~e.27 (i / inhibit-01~e.27 :ARG1 (p2 / pathway :name (n2 / name :op1 "AKT/mTOR"~e.24,26))))) :ARG1-of (u / useful-05~e.31) :ARG1-of (p / possible-01~e.28))) :time (a2 / after~e.0 :op1 (t3 / treat-04~e.1 :ARG2~e.2 s3~e.3) :ARG1-of (d3 / describe-01 :ARG0 (f / figure~e.5 :mod "1B")))) # ::id a_pmid_2514_2146.104 ::amr-annotator SDL-AMR-09 ::preferred # ::tok In contrast , M262 is an AKT1 amplified cell line [ @ 31 @ ] with high sensitivity to SCH772984 and vemurafenib . # ::alignments 1-1 3-1.1.2.1.1 4-1.1.2.r 7-1.1.1.1.1.1 9-1.1.1 10-1.1 11-1.1 14-1.1.3.1.1.1 18-1.1.2.2.2 19-1.1.2 19-1.1.2.2 19-1.1.2.2.r 20-1.1.2.2.1.r 21-1.1.2.2.1.1.1.1 22-1.1.2.2.1 23-1.1.2.2.1.2.1.1 (c / contrast-01~e.1 :ARG2 (c2 / cell-line~e.10,11 :ARG1-of (a / amplify-01~e.9 :ARG0 (g / gene :name (n / name :op1 "AKT1"~e.7))) :domain~e.4 (c3 / cell-line~e.19 :name (n2 / name :op1 "M262"~e.3) :ARG0-of~e.19 (s / sensitive-03~e.19 :ARG1~e.20 (a2 / and~e.22 :op1 (s2 / small-molecule :name (n3 / name :op1 "SCH772984"~e.21)) :op2 (s3 / small-molecule :name (n4 / name :op1 "vemurafenib"~e.23))) :ARG1-of (h / high-02~e.18))) :ARG1-of (d / describe-01 :ARG0 (p / publication :ARG1-of (c4 / cite-01 :ARG2 31~e.14))))) # ::id a_pmid_2514_2146.105 ::amr-annotator SDL-AMR-09 ::preferred # ::tok Treatment of M262 with SCH772984 reduced both pERK1 @/@ 2 and pAKT levels , indicating blockade of the MAPK pathway and PI3K @/@ AKT pathway at the same time ( Figure 1 @ C ) . # ::alignments 0-1.1 1-1.1.1.r 2-1.1.1.1.1 3-1.1.2.r 4-1.1.2.1.1 5-1 9-1.2.1.1.1.1 10-1.2 11-1.2.1.1.2 11-1.2.2.1.1.1 12-1.2.1 12-1.2.2 14-1.3 15-1.3.1 16-1.3.1.1.r 18-1.3.1.1.1.1.1 19-1.3.1.1.1 19-1.3.1.1.2 20-1.3.1.1 21-1.3.1.1.2.1.1 23-1.3.1.1.2.1.1 24-1.3.1.1.1 25-1.3.1.2.r 27-1.3.1.2.1 28-1.3.1.2 30-1.4.1 (r / reduce-01~e.5 :ARG0 (t2 / treat-04~e.0 :ARG1~e.1 (c / cell-line :name (n3 / name :op1 "M262"~e.2)) :ARG2~e.3 (s2 / small-molecule :name (n4 / name :op1 "SCH772984"~e.4))) :ARG1 (a / and~e.10 :op1 (l / level~e.12 :quant-of (e / enzyme :name (n5 / name :op1 "ERK1/2"~e.9) :ARG3-of (p3 / phosphorylate-01~e.11))) :op2 (l2 / level~e.12 :quant-of (e2 / enzyme :name (n6 / name :op1 "AKT"~e.11) :ARG3-of p3))) :ARG0-of (i / indicate-01~e.14 :ARG1 (b / blockade-01~e.15 :ARG1~e.16 (a2 / and~e.20 :op1 (p / pathway~e.19,24 :name (n / name :op1 "MAPK"~e.18)) :op2 (p2 / pathway~e.19 :name (n2 / name :op1 "PI3K/AKT"~e.21,23))) :time~e.25 (t3 / time~e.28 :ARG1-of (s3 / same-01~e.27)))) :ARG1-of (d / describe-01 :ARG0 (f / figure~e.30 :mod "1C"))) # ::id a_pmid_2514_2146.106 ::amr-annotator SDL-AMR-09 ::preferred # ::tok In general , the presence of AKT1 or AKT2 amplification did not preclude sensitivity to SCH772984 , as three of five such cell lines were highly sensitive to SCH772984 ( M229 , M249 , and M262 ) , one was intermediately sensitive ( M255 ) , and M233 and M308 were resistant ( Figure 1 @ A ) . # ::alignments 1-1.6 7-1.2.1.1.1.1 11-1.2.1.2.1.1 13-1.2 15-1.1 15-1.1.r 16-1 17-1.3 18-1.3.1.r 19-1.3.1.1.1 21-1.4.r 22-1.4.1.1.1.1 24-1.4.1.1.1.2.1.1 26-1.4.1.1.1.2.1 27-1.4.1.1.1 27-1.4.1.2 29-1.4.1.1.3 30-1.4.1.1 31-1.4 32-1.3.1.1.1 34-1.4.1.1.1.3.1.1.1.1 36-1.4.1.1.1.3.1.2.1.1 38-1.4.1.1.1.3.1 39-1.4.1.1.1.3.1.3.1.1 42-1.4.1.2.1 44-1.4.1.2.2.2 45-1.4.1.2.2 47-1.4.1.2.3.1.1.1 50-1.4.1 50-1.4.1.3 50-1.4.1.3.r 51-1.4.1.3.1.1.1 52-1.4.1.3 53-1.4.1.3.2.1.1 55-1.4.1.3.3 57-1.5.1 59-1.4.1.2.1 (p / preclude-01~e.16 :polarity~e.15 -~e.15 :ARG0 (a / amplify-01~e.13 :ARG1 (a2 / and :op1 (g2 / gene :name (n / name :op1 "AKT1"~e.7)) :op2 (g3 / gene :name (n2 / name :op1 "AKT2"~e.11)))) :ARG1 (s / sensitive-03~e.17 :ARG1~e.18 (s2 / small-molecule :name (n3 / name :op1 "SCH772984"~e.19,32))) :ARG1-of~e.21 (c / cause-01~e.31 :ARG0 (a3 / and~e.50 :op1 (s3 / sensitive-03~e.30 :ARG0 (c2 / cell-line~e.27 :quant 3~e.22 :ARG1-of (i / include-91 :ARG2 (c3 / cell-line~e.26 :quant 5~e.24)) :ARG1-of (m / mean-01 :ARG2 (a4 / and~e.38 :op1 (c4 / cell-line :name (n5 / name :op1 "M229"~e.34)) :op2 (c5 / cell-line :name (n6 / name :op1 "M249"~e.36)) :op3 (c6 / cell-line :name (n7 / name :op1 "M262"~e.39))))) :ARG1 s2 :ARG1-of (h / high-02~e.29)) :op2 (c7 / cell-line~e.27 :quant 1~e.42,59 :ARG0-of (s5 / sensitive-03~e.45 :ARG1 s2 :mod (i2 / intermediate~e.44)) :ARG1-of (m2 / mean-01 :ARG2 (c8 / cell-line :name (n8 / name :op1 "M255"~e.47)))) :op3~e.50 (a5 / and~e.50,52 :op1 (c9 / cell-line :name (n9 / name :op1 "M233"~e.51)) :op2 (c10 / cell-line :name (n10 / name :op1 "M308"~e.53)) :ARG0-of (r / resist-01~e.55 :ARG1 s2)))) :ARG1-of (d / describe-01 :ARG0 (f / figure~e.57 :mod "1A")) :ARG1-of (g / general-02~e.1)) # ::id a_pmid_2514_2146.107 ::amr-annotator SDL-AMR-09 ::preferred # ::tok Given high baseline pAKT levels were seen in some cells resistant to ERK inhibition ( Additional file 2 @ : Figure S2 ) and the persistence of pAKT activity with SCH722984 treatment , we evaluated the effect of SCH772984 in combination with the AKT inhibitor MK @-@ 2206 or the mTOR inhibitor MK @-@ 8669 . # ::alignments 1-1.1.1.1.3 2-1.1.1.1.2 3-1.1.1.1.1.1.1 3-1.1.1.1.1.2 3-1.1.2.1.1.2 4-1.1.1.1 6-1.1.1 7-1.1.1.2.r 8-1.1.1.2.1 9-1.1.1.2 10-1.1.1.2.2 11-1.1.1.2.2.1.r 12-1.1.1.2.2.1.1.1.1 13-1.1.1.2.2.1 16-1.1.1.3.1 18-1.1.1.3.1.1 21-1.1.1.3.1.2.1 22-1.1.1.3.1.2.1.1 24-1.1 26-1.1.2 28-1.2.2.1.2.1.1.2.1.1.1 29-1.1.2.1 30-1.1.2.1.1.r 31-1.1.2.1.1.3.1.1.1 32-1.1.2.1.1 32-1.1.2.1.1.3 32-1.1.2.1.1.3.r 34-1.2.1 35-1.2 37-1.2.2 39-1.2.2.1.1.1 41-1.2.2.1 41-1.2.2.1.2 41-1.2.2.1.2.r 44-1.1.2.1.1.1.1 45-1.2.2.1.2.1.2 45-1.2.2.1.2.1.2.2 45-1.2.2.1.2.1.2.2.r 46-1.2.2.1.2.1.1.1.1 48-1.2.2.1.2.1.1.1.1 49-1.2.2.1.2.1 51-1.2.2.1.2.1.2.2.1.1.1 52-1.2.2.1.2.1.1 52-1.2.2.1.2.1.1.2 52-1.2.2.1.2.1.1.2.r 52-1.2.2.1.2.1.2 52-1.2.2.1.2.1.2.2 52-1.2.2.1.2.1.2.2.r 53-1.2.2.1.2.1.2.1.1 55-1.2.2.1.2.1.2.1.1 (c / cause-01 :ARG0 (a / and~e.24 :op1 (s / see-01~e.6 :ARG1 (l / level~e.4 :quant-of (e2 / enzyme :name (n2 / name :op1 "AKT"~e.3) :ARG3-of (p / phosphorylate-01~e.3)) :mod (b / baseline~e.2) :ARG1-of (h / high-02~e.1)) :location~e.7 (c2 / cell~e.9 :quant (s2 / some~e.8) :ARG0-of (r / resist-01~e.10 :ARG1~e.11 (i2 / inhibit-01~e.13 :ARG1 (e / enzyme :name (n3 / name :op1 "ERK"~e.12))))) :ARG1-of (d / describe-01 :ARG0 (f / file~e.16 :mod 2~e.18 :ARG1-of (m / mean-01 :ARG2 (f2 / figure~e.21 :mod "S2"~e.22)) :ARG1-of (a5 / add-02)))) :op2 (p2 / persist-01~e.26 :ARG1 (a2 / activity-06~e.29 :ARG0~e.30 (e4 / enzyme~e.32 :name (n4 / name :op1 "AKT"~e.44) :ARG3-of (p3 / phosphorylate-01~e.3) :ARG1-of~e.32 (t2 / treat-04~e.32 :ARG2 (s3 / small-molecule :name (n5 / name :op1 "SCH722984"~e.31))))))) :ARG1 (e5 / evaluate-01~e.35 :ARG0 (w / we~e.34) :ARG1 (a4 / affect-01~e.37 :ARG0 (s4 / small-molecule~e.41 :name (n6 / name :op1 "SCH772984"~e.39) :ARG1-of~e.41 (c3 / combine-01~e.41 :ARG2 (o / or~e.49 :op1 (s6 / small-molecule~e.52 :name (n7 / name :op1 "MK-2206"~e.46,48) :ARG0-of~e.52 (i3 / inhibit-01~e.52 :ARG1 (e6 / enzyme :name (n8 / name :op1 "AKT"~e.28)))) :op2 (s5 / small-molecule~e.45,52 :name (n9 / name :op1 "MK-8669"~e.53,55) :ARG0-of~e.45,52 (i4 / inhibit-01~e.45,52 :ARG1 (p4 / protein :name (n10 / name :op1 "mTOR"~e.51)))))))))) # ::id a_pmid_2514_2146.108 ::amr-annotator SDL-AMR-09 ::preferred # ::tok The addition of either the AKTi or mTORi always resulted in more potent cell growth inhibition compared to ERKi alone ( Additional file 3 @ : Figures S3A and 3B ) . # ::alignments 1-1.1 1-1.4.1.3 6-1.1.1 8-1.3 9-1 10-1.2.r 11-1.2.2.1 12-1.2.2 13-1.2.1.1 14-1.2.1 15-1.1.1.1 15-1.1.1.1.1 15-1.1.1.1.1.r 15-1.1.1.2 15-1.1.1.2.1 15-1.1.1.2.1.r 15-1.2 15-1.2.3 15-1.2.3.1 15-1.2.3.1.r 16-1.2.3.r 19-1.2.3.1.1.2 22-1.4.1 24-1.4.1.1 27-1.4.1.2.1.1 27-1.4.1.2.1.2 28-1.4.1.2.1.1.1 29-1.4.1.2.1 (r / result-01~e.9 :ARG1 (a / add-02~e.1 :ARG1 (o / or~e.6 :op1 (m3 / molecular-physical-entity~e.15 :ARG0-of~e.15 (i / inhibit-01~e.15 :ARG1 (p3 / protein-family :name (n / name :op1 "AKT")))) :op2 (m4 / molecular-physical-entity~e.15 :ARG0-of~e.15 (i2 / inhibit-01~e.15 :ARG1 (p / protein :name (n2 / name :op1 "mTOR")))))) :ARG2~e.10 (i3 / inhibit-01~e.15 :ARG1 (g / grow-01~e.14 :ARG1 (c / cell~e.13)) :mod (p2 / potent~e.12 :mod (m / more~e.11)) :compared-to~e.16 (m5 / molecular-physical-entity~e.15 :ARG0-of~e.15 (i4 / inhibit-01~e.15 :ARG1 (e2 / enzyme :name (n3 / name :op1 "ERK") :mod (a3 / alone~e.19))))) :time (a2 / always~e.8) :ARG1-of (d / describe-01 :ARG0 (f / file~e.22 :mod 3~e.24 :ARG1-of (m2 / mean-01 :ARG2 (a5 / and~e.29 :op1 (f2 / figure~e.27 :mod "S3A"~e.28) :op2 (f3 / figure~e.27 :mod "S3B"))) :ARG1-of (a6 / add-02~e.1)))) # ::id a_pmid_2514_2146.109 ::amr-annotator SDL-AMR-09 ::preferred # ::tok Combining SCH772984 with the mTOR inhibitor MK @-@ 8669 was particularly synergistic . # ::alignments 0-1.1 1-1.1.1.1.1 2-1.1.2.r 4-1.1.2.2.1.1.1 5-1.1.2 5-1.1.2.2 5-1.1.2.2.r 6-1.1.2.1.1 8-1.1.2.1.1 10-1.2 11-1 (s2 / synergize-01~e.11 :ARG0 (c / combine-01~e.0 :ARG1 (s / small-molecule :name (n / name :op1 "SCH772984"~e.1)) :ARG2~e.2 (s3 / small-molecule~e.5 :name (n2 / name :op1 "MK-8669"~e.6,8) :ARG0-of~e.5 (i2 / inhibit-01~e.5 :ARG1 (p / protein :name (n3 / name :op1 "mTOR"~e.4))))) :mod (p2 / particular~e.10)) # ::id a_pmid_2514_2146.110 ::amr-annotator SDL-AMR-09 ::preferred # ::tok For BRAF @-@ mutant cell line M233 , both combinations resulted in more complete decrease in pERK compared to treatment with SCH772984 alone . # ::alignments 1-1.3.2.1.1 3-1.3.2 3-1.3.2.2 3-1.3.2.2.r 4-1.3 5-1.3 6-1.3.1.1 8-1.1.1 9-1.1 10-1 11-1.2.r 12-1.2.2.1 13-1.2.2 14-1.2 15-1.2.1.r 16-1.2.1.1.1 16-1.2.1.2 17-1.2.3.r 19-1.2.3 20-1.2.3.1.r 21-1.2.3.1.1.1 22-1.2.3.1.2 (r / result-01~e.10 :ARG1 (c / combine-01~e.9 :mod (b / both~e.8)) :ARG2~e.11 (d / decrease-01~e.14 :ARG1~e.15 (e / enzyme :name (n / name :op1 "ERK"~e.16) :ARG3-of (p / phosphorylate-01~e.16)) :degree (c2 / complete~e.13 :mod (m / more~e.12)) :compared-to~e.17 (t / treat-04~e.19 :ARG2~e.20 (s / small-molecule :name (n2 / name :op1 "SCH772984"~e.21) :mod (a / alone~e.22)))) :beneficiary (c3 / cell-line~e.4,5 :name (n3 / name :op1 "M233"~e.6) :mod (e2 / enzyme~e.3 :name (n4 / name :op1 "BRAF"~e.1) :ARG2-of~e.3 (m2 / mutate-01~e.3)))) # ::id a_pmid_2514_2146.111 ::amr-annotator SDL-AMR-09 ::preferred # ::tok Despite the improved inhibition of the MAPK pathway , the levels of pAKT were largely unaffected by the addition of MK @-@ 2206 or MK @-@ 8669 ( Additional file 3 @ : Figure S3C ) . # ::alignments 0-1 2-1.2.2 3-1.2 4-1.2.1.r 6-1.2.1.1.1 7-1.2.1 10-1.1.3 11-1.1.3.1.r 12-1.1.3.1.1.1 12-1.1.3.1.2 14-1.1.4 15-1.1 15-1.1.1 15-1.1.1.r 18-1.1.2 18-1.3.1.3 19-1.1.2.1.r 20-1.1.2.1.1.1.1 20-1.1.2.1.2.1.1 22-1.1.2.1.1.1.1 23-1.1.2.1 24-1.1.2.1.1.1.1 24-1.1.2.1.2.1.1 26-1.1.2.1.2.1.1 29-1.3.1 31-1.3.1.1 34-1.3.1.2.1 35-1.3.1.2.1.1 (h / have-concession-91~e.0 :ARG1 (a / affect-01~e.15 :polarity~e.15 -~e.15 :ARG0 (a2 / add-02~e.18 :ARG1~e.19 (o / or~e.23 :op1 (s / small-molecule :name (n3 / name :op1 "MK-2206"~e.20,22,24)) :op2 (s2 / small-molecule :name (n4 / name :op1 "MK-8669"~e.20,24,26)))) :ARG1 (l / level~e.10 :quant-of~e.11 (e / enzyme :name (n2 / name :op1 "AKT"~e.12) :ARG3-of (p2 / phosphorylate-01~e.12))) :degree (l2 / large~e.14)) :ARG2 (i / inhibit-01~e.3 :ARG1~e.4 (p / pathway~e.7 :name (n / name :op1 "MAPK"~e.6)) :ARG1-of (i2 / improve-01~e.2)) :ARG1-of (d / describe-01 :ARG0 (f / file~e.29 :mod 3~e.31 :ARG1-of (m3 / mean-01 :ARG2 (f2 / figure~e.34 :mod "S3C"~e.35)) :ARG1-of (a4 / add-02~e.18)))) # ::id a_pmid_2514_2146.112 ::amr-annotator SDL-AMR-09 ::preferred # ::tok Potent SCH772984 @-@ mediated ERK inhibition in BRAF @ -@ wild type melanoma cell lines