Cell Lines and Culture Conditions

RWPE1, LNCaP, PC3, C4-2B, DU145, CWR22V1, and HEK293 cells were obtained from the American Type Culture Collection (ATCC, Manassas, VA). C4-2B cells resistant to enzalutamide (C4-2B-MDVR) were generated by culturing C4-2B cells to increasing concentrations of enzalutamide (5–40 µM) in media for over 12 months and maintained in complete media supplemented with 20 µM enzalutamide as described previously [58 (link), 59 (link)]. HEK293 cells were cultured in complete DMEM medium. RWPE1, LNCaP, LNCaP-KDM8 [17 (link)], PC3, C4-2B, DU145, and CWR22RV1 cells were cultured in RPMI-1640 medium supplied with 10% heat-inactivated FBS, 100 U/ml penicillin, and 0.1 mg/ml streptomycin in a humidified incubator with 5% CO₂. For hormone induction experiments, the growth media were replaced by RPMI-1640 containing 10% charcoal-dextran-treated (CDT) FBS (CDT-FBS) for 2 days before treatment with 10 nM dihydrotestosterone (DHT).