Fig. 4.

BIRC5 inhibits EGFRi-induced apoptosis in the context of CBX5 loss in EGFR-mutant LUAD. (A) Immunoblot analysis measuring levels of CASP3 in LUAD cell lines expressing an NS or CBX5 shRNA and treated with erlotinib (50 nM) or DMSO for 24 h. (B) The indicated LUAD cell lines expressing an NS or CBX5 shRNA were treated with erlotinib (50 nM) or DMSO for 24 h and then analyzed by fluorescence-activated cell sorting (FACS)-based annexin V-PE staining. (C) Immunoblot analysis measuring levels of cleaved CASP3 in PC9 cells expressing an NS or CBX5 shRNA alone, or in conjunction with an NS shRNA or BIRC5 shRNA, and treated with erlotinib (50 nM) or DMSO for 24 h. (D) PC9 cells expressing an NS or CBX5 shRNA alone, or in conjunction with an NS or BIRC5 shRNA, were treated with erlotinib (100 nM) or DMSO, and survival was measured in clonogenic assays. Representative wells for cells grown under the indicated conditions are shown. (E) PC9 cells expressing an NS or CBX5 shRNA alone, or in conjunction with an NS or BIRC5 shRNA, were injected subcutaneously into the flanks of NSG mice (n = 5). Mice were treated with either vehicle (0.5% methylcellulose) or erlotinib (25 mg/kg) 3 d per week until the end of the experiment. Average tumor volumes at the end of the experiment are shown. Data are presented as the mean ± SEM. ns = not significant; *P < 0.05, **P < 0.01, ***P < 0.001, and ****P < 0.0001.