Down-regulation of Rad51 expression overcomes drug resistance to gemcitabine in human non-small-cell lung cancer cells.
Gemcitabine (2',2'-difluorodeoxycytidine), a deoxycytidine analog, and erlotinib, an epidermal growth factor receptor-tyrosine kinase inhibitor, are used clinically to treat patients with non-small-cell lung cancer (NSCLC). However, the molecular mechanisms for the drug resistance of gemcitabine in NSCLC cells are poorly understood. In this study, we used constructs containing human Rad51 cDNA or specific Rad51 small interfering RNA (siRNA) to examine the role of Rad51 in chemoresistance of gemcitabine in three different human NSCLC cell lines. Exposure of human NSCLC cell lines to gemcitabine increased the phosphorylation levels of mitogen-activated protein kinase kinase (MKK) 1/2-extracellular signal-regulated kinase (ERK) 1/2 and AKT in a time- and dose-dependent manner, which was accompanied by an induction of Rad51 mRNA and protein expression. Gemcitabine increased the expression of Rad51 by increasing its mRNA and protein stability. Blockage of ERK1/2 or AKT activation by 1,4-diamino-2,3-dicyano-1,4-bis(methylthio)butadiene (U0126; MKK1/2 inhibitor) or 2-(4-morpholinyl)-8-phenyl-4H-1-benzopyran-4-one (LY294002; phosphatidyl inositol 3-kinase inhibitor), respectively, decreased the gemcitabine-induced Rad51 expression. Gemcitabine-induced cytotoxicity was significantly increased using siRNA depletion of Rad51 or blockage of ERK1/2 and AKT activation. Erlotinib enhanced the gemcitabine-induced cytotoxicity via the inactivation of ERK1/2 and AKT and the down-regulation of Rad51. Enforced expression of constitutively active MKK1/2 or AKT recovered cell viability and Rad51 protein levels that were decreased by the combination of erlotinib and gemcitabine. Suppression of Rad51 expression or the inactivation of ERK1/2 or AKT signaling may be considered potential therapeutic modalities for gemcitabine-resistant lung cancer.
Tsai
Min-Shao
MS
Department of Biochemical Science and Technology, National Chiayi University, Chiayi 600, Taiwan.
Kuo
Ya-Hsun
YH
Chiu
Yu-Fan
YF
Su
Ying-Chen
YC
Lin
Yun-Wei
YW
D000964:Antimetabolites, Antineoplastic / Q000494:pharmacology; D002289:Carcinoma, Non-Small-Cell Lung / Q000188:drug therapy* / Q000378:metabolism; D045744:Cell Line, Tumor; D049109:Cell Proliferation / Q000187:drug effects; D002470:Cell Survival / Q000187:drug effects; D003841:Deoxycytidine / Q000031:analogs & derivatives* / Q000494:pharmacology; D015536:Down-Regulation / Q000235:genetics*; D019008:Drug Resistance, Neoplasm / Q000235:genetics*; D004357:Drug Synergism; D000069347:Erlotinib Hydrochloride; D015870:Gene Expression / Q000187:drug effects / Q000235:genetics*; D006801:Humans; D048369:MAP Kinase Kinase 1 / Q000037:antagonists & inhibitors / Q000235:genetics / Q000378:metabolism; D048370:MAP Kinase Kinase 2 / Q000037:antagonists & inhibitors / Q000235:genetics / Q000378:metabolism; D019950:Mitogen-Activated Protein Kinase 1 / Q000378:metabolism; D048052:Mitogen-Activated Protein Kinase 3 / Q000378:metabolism; D000081082:Phosphoinositide-3 Kinase Inhibitors; D010766:Phosphorylation / Q000187:drug effects; D046988:Proteasome Endopeptidase Complex / Q000378:metabolism; D047428:Protein Kinase Inhibitors / Q000494:pharmacology; D051057:Proto-Oncogene Proteins c-akt / Q000037:antagonists & inhibitors / Q000235:genetics / Q000378:metabolism; D011799:Quinazolines / Q000494:pharmacology; D034741:RNA, Small Interfering / Q000235:genetics; D051135:Rad51 Recombinase / Q000235:genetics* / Q000378:metabolism; D014162:Transfection; D025801:Ubiquitin / Q000378:metabolism; D000093542:Gemcitabine
D000964:Antimetabolites, Antineoplastic; D000081082:Phosphoinositide-3 Kinase Inhibitors; D047428:Protein Kinase Inhibitors; D011799:Quinazolines; D034741:RNA, Small Interfering; D025801:Ubiquitin; D003841:Deoxycytidine; D000069347:Erlotinib Hydrochloride; C482128:MAP2K2 protein, human; D051057:Proto-Oncogene Proteins c-akt; C535150:MAPK1 protein, human; D019950:Mitogen-Activated Protein Kinase 1; D048052:Mitogen-Activated Protein Kinase 3; D048369:MAP Kinase Kinase 1; D048370:MAP Kinase Kinase 2; C482125:MAP2K1 protein, human; C495153:RAD51 protein, human; D051135:Rad51 Recombinase; D046988:Proteasome Endopeptidase Complex; C061553:ATP dependent 26S protease; D000093542:Gemcitabine