Identification of GAS1 as an epirubicin resistance-related gene in human gastric cancer cells with a partially randomized small interfering RNA library.
Epirubicin has been widely used for chemotherapeutic treatment of gastric cancer; however, intrinsic and acquired chemoresistance remains an obstacle to successful management. The mechanisms underlying epirubicin resistance are still not well defined. Here we report the construction and application of a partially randomized retrovirus library of 4 x 10(6) small interfering RNAs to identify novel genes whose suppression confers epirubicin resistance in gastric cancer cells SGC7901. From 12 resistant cell colonies, two small interfering RNAs targeting GAS1 (growth arrest-specific 1) and PTEN (phosphatase and tensin homolog), respectively, were identified and validated. We identified a previously unrecognized chemoresistance role for GAS1. GAS1 suppression resulted in significant epirubicin resistance and cross-resistance to 5-fluorouracil and cisplatin in various gastric cancer cell lines. GAS1 suppression promoted multidrug resistance through apoptosis inhibition, partially by up-regulating the Bcl-2/Bax ratio that was abolished by Bcl-2 inhibition. GAS1 suppression induced chemoresistance partially by increasing drug efflux in an ATP-binding cassette transporter and drug-dependent manner. P-glycoprotein (P-gp) and BCRP (breast cancer resistance protein) but not MRP-1 were up-regulated, and targeted knockdown of P-gp and BCRP could partially reverse GAS1 suppression-induced epirubicin resistance. Verapamil, a P-gp inhibitor, could reverse P-gp substrate (epirubicin) but not non-P-gp substrate (5-fluorouracil and cisplatin) resistance in GAS1-suppressed gastric cancer cells. BCRP down-regulation could partially reverse 5-fluorouracil but not cisplatin resistance induced by GAS1 suppression, suggesting 5-fluorouracil but not cisplatin was a BCRP substrate. These results suggest that GAS1 might be a target to overcome multidrug resistance and provide a novel approach to identifying candidate genes that suppress chemoresistance of gastric cancers.
Zhao
Lina
L
State Key Laboratory of Cancer Biology and Xijing Hospital of Digestive Diseases, Fourth Military Medical University, Xi'an 710032, China.
Pan
Yanglin
Y
Gang
Yi
Y
Wang
Honghong
H
Jin
Haifeng
H
Tie
Jun
J
Xia
Lin
L
Zhang
Yongguo
Y
He
Lijie
L
Yao
Liping
L
Qiao
Taidong
T
Li
Tingting
T
Liu
Zhiguo
Z
Fan
Daiming
D
D020168:ATP Binding Cassette Transporter, Subfamily B, Member 1 / Q000235:genetics / Q000378:metabolism; D000070997:ATP Binding Cassette Transporter, Subfamily G, Member 2; D018528:ATP-Binding Cassette Transporters; D000903:Antibiotics, Antineoplastic / Q000494:pharmacology; D017209:Apoptosis / Q000187:drug effects; D001483:Base Sequence; D015153:Blotting, Western; D018797:Cell Cycle Proteins / Q000235:genetics / Q000378:metabolism*; D045744:Cell Line, Tumor; D002470:Cell Survival / Q000187:drug effects; D002945:Cisplatin / Q000494:pharmacology; D018432:Drug Resistance, Multiple / Q000235:genetics; D019008:Drug Resistance, Neoplasm / Q000235:genetics*; D015251:Epirubicin / Q000494:pharmacology*; D005434:Flow Cytometry; D005472:Fluorouracil / Q000494:pharmacology; D058851:GPI-Linked Proteins; D015723:Gene Library; D006801:Humans; D008565:Membrane Proteins / Q000235:genetics / Q000378:metabolism*; D008969:Molecular Sequence Data; D009363:Neoplasm Proteins; D051059:PTEN Phosphohydrolase / Q000235:genetics / Q000378:metabolism; D019253:Proto-Oncogene Proteins c-bcl-2 / Q000378:metabolism; D034741:RNA, Small Interfering / Q000235:genetics*; D020133:Reverse Transcriptase Polymerase Chain Reaction; D013274:Stomach Neoplasms / Q000235:genetics / Q000378:metabolism / Q000473:pathology; D014162:Transfection; D051028:bcl-2-Associated X Protein / Q000378:metabolism
C116614:ABCG2 protein, human; D020168:ATP Binding Cassette Transporter, Subfamily B, Member 1; D000070997:ATP Binding Cassette Transporter, Subfamily G, Member 2; D018528:ATP-Binding Cassette Transporters; D000903:Antibiotics, Antineoplastic; C494826:BAX protein, human; D018797:Cell Cycle Proteins; C096192:GAS1 protein, human; D058851:GPI-Linked Proteins; D008565:Membrane Proteins; D009363:Neoplasm Proteins; D019253:Proto-Oncogene Proteins c-bcl-2; D034741:RNA, Small Interfering; D051028:bcl-2-Associated X Protein; D015251:Epirubicin; D051059:PTEN Phosphohydrolase; C494929:PTEN protein, human; D002945:Cisplatin; D005472:Fluorouracil