Fig. 5

ETV6-TWIST1 signaling is involved in the molecular mechanism of drug resistance. (a) Proliferation assay in three DU145 derived cells treated with a tyrosine kinase inhibitor (TKI: AG1478, 0.1~ 10 μM), n = 8. shLacZ, control; shETV6, ETV6-knockdown; shETV6 + siTWIST1, both ETV6- and TWIST1-knockdown. (b) Proliferation assay in three stable RasB1 derived cells treated with a TKI (CI1033, 0.1~ 10 nM), n = 8. EV, control vector; ETV6, ETV6-expressing vector; ETV6 + TWIST1, both ETV6- and TWIST1-expressing vectors. (c) Tumor growth analysis of stable RasB1 cell lines (EV vs. ETV6) subcutaneously inoculated in male nude mice followed by treatment with CI1033. Tumor sizes were monitored weekly (left, n = 5). At the end, tumor weights were also measured (right, n = 5). (d) Selected images of mice from panel C, containing tumors (arrows) derived from stable RasB1 cell lines (EV vs. ETV6). (e) Western blot analysis of human prostate cancer cells. RasB1 cells were introduced with exogenous ETV6 or under ETV6-knockdown in 22RV1 and LNCaP cells by an siRNA approach (siETV6). EV, empty vector; scr., siRNA control. (f) Working model, activation of the epidermal growth factor receptor (EGFR) promotes tumor progression and drug resistance through RAS signaling and suppression of ETV6, which lead to TWIST1-dependent malignant phenotypes. A mutual inhibitory circuit exists between EGFR-RAS signaling and ETV6