Prostate Cancer Cell Line Cultivation and Validation

The human PCa cell line PC3 was obtained from the American Type Culture Collection. C4-2 cells were provided by Prof. Thalmann (University of Berne, Switzerland) [33 (link)]. The enzalutamide-resistant LNCaP sub cell line MR49F cells were provided by Dr. Gleave [19 (link), 34 (link)]. Castration-resistance and enzalutamide-resistance was established by enzalutamide treatment of xenograft mouse models. To eliminate the possibility that the changes observed were due to the mouse microenvironment we choose the CRPC model C4-2 as enzalutamide sensitive model created by passaging twice through mice similar to the MR49F cells (S1A and S1B Fig). The cell lines LAPC4-CTRL, LAPC4-EnzaR, LNCaPabl-CTRL, LNCaPabl-EnzaR, DuCaP-CTRL, and DuCaP-EnzaR were provided by Prof. Culig (Medical University of Innsbruck, Austria) and enzalutamide resistance has been developed by the dose escalation method (S1C and S1D Fig) as described by Hoefer et al. [35 (link)]. Culture media used for the cell lines are listed in the S1 Table. All cells were maintained at 37°C in 5% CO₂. Enzalutamide-resistance has been confirmed by the increase of the IC₅₀ in cell viability (S1B and S1D Fig). Mycoplasma testing was performed regularly using the Mycoalert Detection Assay (Lonza). Cell line authentication was performed yearly by STR profiling.