Fig. 6.

Combined blockade of cholesterol and steroid biosynthesis with simvastatin and AKRi to inhibit PCa cells. Du/CAF (a) and LN/CAF (b) co-culture spheroids were established as described under methods. CAFs stably express GFP to distinguish them from tumor cells. Four days after seeding, spheroids were treated with 5 μM enzalutamide (enza), 5 μM simvastatin (sim), and 50 μM of an inhibitor against AKR1C3 (AKRi) in medium supplemented with 10% CS_FCS. Cell viability was assessed after 10 days using CellTiterGlo assay. (c, d, e) DuCaP EnzaR, CWR22Rv1, and LNCaPabl EnzaR were seeded into 96 well plates. After overnight incubation, drugs were added in medium with 10% CS_FCS as indicated. Representative images were taken after 6 days of treatment with 5 μM simvastatin and cropped to show typical rounded cells that were depicted with a black arrow. Cell viability was determined after 6 days via CellTiterGlo cell viability assay and expressed as mean relative luminescence units (RLU) from at least 3 independent experiments with SEM. f DuCaP EnzaR were seeded into 96 well plates. Treatment with 5 μM simvastatin was performed in RPMI + 10% FCS. Cell viability was determined after 6 days of treatment via CellTiterGlo cell viability assay and expressed as mean relative luminescence units (RLU) from at least three independent experiments with SEM. (* and ⁺ P < 0.05, ** and ⁺⁺ P < 0.01, *** and ⁺⁺⁺ P < 0.001, * related to mock control, ⁺ related to enzalutamide-treated cells)