title: Knockdown of the Bcl-2 gene increases sensitivity to EGFR tyrosine kinase inhibitors in the H1975 lung cancer cell line harboring T790M mutation.
abstract: Epidermal growth factor receptor tyrosine kinase inhibitors (EGFR-TKIs) are being widely used as targeted therapy in non-small cell lung cancer (NSCLC), but most cases acquire drug-resistance in 9 months. However, the mechanisms of resistance are still not fully understood. Since it has been demonstrated that EGFR-TKI-mediated repression of downstream signaling cascades and apoptosis induction is a key mechanism through which EGFR-TKIs exert their cytotoxic effects, we reasoned that activation of downstream signaling pathways and changes in the expression of apoptosis-related proteins contribute to the acquired resistance to EGFR-TKIs. We analyzed the protein levels of p-Akt, Bcl-2, Bax between gefitinib-sensitive and gefitinib-resistant lung cancer cell lines and evaluated whether targeting the anti-apoptotic protein Bcl-2 induces cell apoptosis and further sensitizes resistant H1975 cells to gefitinib. The data showed that p-Akt was activated and accompanied by substantial Bcl-2 in the H1975 lung cancer cell line, whereas no evidence was observed in HCC827 cells. Using small interfering RNA (siRNA) to silence Bcl-2 in H1975 cells led to significant downregulation of Bcl-2 protein expression, decreased cell viability in vitro and induced intrinsic apoptosis confirmed by flow cytometry and PARP cleavage. In Bcl-2 siRNA-transfected cells, adding gefitinib further reduced the number of viable cells, induced apoptosis to a greater extent compared to either treatment alone. These preclinical data suggested that downregulation of Bcl-2 by RNAi in the gefitinib-resistant H1975 lung cancer cell line with T790M mutation enhanced the effects of gefitinib and may offer a novel therapeutic strategy for the treatment of NSCLC.
authors.affiliation: Department of Oncology, Tongji Hospital, Tongji Medical College, Huazhong University of Science and Technology, Wuhan 430030, P.R. China.
authors.forename: Man
authors.initials: M
authors.lastname: Zou
authors.forename: Shu
authors.initials: S
authors.lastname: Xia
authors.forename: Liang
authors.initials: L
authors.lastname: Zhuang
authors.forename: Na
authors.initials: N
authors.lastname: Han
authors.forename: Qian
authors.initials: Q
authors.lastname: Chu
authors.forename: Tengfei
authors.initials: T
authors.lastname: Chao
authors.forename: Ping
authors.initials: P
authors.lastname: Peng
authors.forename: Yu
authors.initials: Y
authors.lastname: Chen
authors.forename: Qi
authors.initials: Q
authors.lastname: Gui
authors.forename: Shiying
authors.initials: S
authors.lastname: Yu
mesh_terms: D000970:Antineoplastic Agents / Q000494:pharmacology; D017209:Apoptosis / Q000187:drug effects / Q000235:genetics; D002289:Carcinoma, Non-Small-Cell Lung / Q000188:drug therapy* / Q000235:genetics; D045744:Cell Line, Tumor; D019008:Drug Resistance, Neoplasm / Q000187:drug effects / Q000235:genetics; D066246:ErbB Receptors / Q000037:antagonists & inhibitors* / Q000235:genetics* / Q000378:metabolism; D000077156:Gefitinib; D055785:Gene Knockdown Techniques; D019254:Genes, bcl-2 / Q000187:drug effects*; D006801:Humans; D008175:Lung Neoplasms / Q000188:drug therapy* / Q000235:genetics* / Q000473:pathology; D009154:Mutation; D047428:Protein Kinase Inhibitors / Q000494:pharmacology*; D051057:Proto-Oncogene Proteins c-akt / Q000235:genetics / Q000378:metabolism; D019253:Proto-Oncogene Proteins c-bcl-2 / Q000235:genetics / Q000378:metabolism; D011799:Quinazolines / Q000494:pharmacology; D034622:RNA Interference; D034741:RNA, Small Interfering; D051028:bcl-2-Associated X Protein / Q000235:genetics / Q000378:metabolism
chemical_list: D000970:Antineoplastic Agents; C494826:BAX protein, human; D047428:Protein Kinase Inhibitors; D019253:Proto-Oncogene Proteins c-bcl-2; D011799:Quinazolines; D034741:RNA, Small Interfering; D051028:bcl-2-Associated X Protein; C512478:EGFR protein, human; D066246:ErbB Receptors; D051057:Proto-Oncogene Proteins c-akt; D000077156:Gefitinib