Fig. 1: AR inhibitor-resistant prostate cancer cell model is established.

A 6*10³ cells were plated in 96-well plates and treated with various concentrations of ENZ and EPI for 48 h. The survival curve was detected by CCK8. B, C 1*10⁶ LNCaP cells were plated and either untreated (negative control; NC), treated with ENZ (10 µM) or EPI (8 µM) with normal FBS media change every 3 days for a total of 9 days (short-term treatment, ST) and for 33 days (long-term treatment, LT). Representative microscopic images (B) and cell proliferation curve (C) are shown. Data were expressed as mean ± std. of biological triplicates; Student’s t-test were performed; *p < 0.05. D Cells from NC, ST-EPI, ST-ENZ, LT-EPI, and LT-ENZ were analyzed for cell cycle distribution by FACS. E Levels of cell cycle markers are quantified in NC, ST, or LT cells by WB. β-actin protein is used as loading control. One-way ANOVA with Turkey test, p < 0.05. Data are expressed as mean ± std of biological triplicates. F Levels of AR, AR target (TMPRS2, PSA), AR-V567, AR-Vs target (UBE2C, CDC20) proteins in NC, ST-EPI, ST-ENZ, LT-EPI, and LT-ENZ were quantified by WB. GAPDH protein was used as loading control. One-way ANOVA with Turkey test, p < 0.05. Data are expressed as mean ± std of biological triplicates.