Nebivolol inhibits cell growth, inhibits autophagic flux, and enhances growth inhibition in combination with bicalutamide in prostate cancer cells

Introduction: Nebivolol, a beta1-adrenergic receptor blocker, is being explored as a repurposed therapeutic for breast cancer, showing inhibition of cell growth and autophagic flux. This study evaluated nebivolol's anti-tumor effects in prostate cancer (PCa) cells and determined whether it enhances the effect of the androgen receptor antagonist bicalutamide. Methods: PCa cell lines, LNCap and 22Rv1, were treated with vehicle or nebivolol (0.01-30μM) for 7 days to assess cell growth using the EnSight™ Multimode Plate Reader. Clonogenic growth was measured after 14-15 days of vehicle or nebivolol (0.1-10μM). Autophagy and lysosome formation were evaluated after 24-hour treatment with vehicle, nebivolol (1 or 10μM), or chloroquine (50μM) using Hoechst 33342, Cyto-ID® Green, and Lyso-ID® Red staining followed by confocal imaging. Combination studies with nebivolol and bicalutamide (0-30μM) were performed over 7-day using concentration-response curves to access growth inhibition and interaction effects. IC 50 values were calculated by non-linear regression curve analysis. LC3B protein expression was measured in 22Rv1 cells by immunoblotting following vehicle or nebivolol (10μM) treatment for 24, 48, and 72 hours. Statistical significance was determined using paired student's t-test or one-way ANOVA followed by Dunnett's multiple comparison test, as appropriate, using GraphPad Prism v.10. Drug synergism of nebivolol and bicalutamide was tested by CompuSyn software using the Chou-Talalay method. Results: Nebivolol inhibited cell growth (IC 50 : 37.1μM in LNCap; 9.8μM in 22Rv1) and suppressed clonogenic potential (IC 50 : 1.1μM in LNCap; 2.5μM in 22Rv1) in a concentration-dependent manner. Nebivolol (10μM) significantly increased autophagosome and lysosome accumulation compared with vehicle (p<0.05). Nebivolol significantly upregulated LC3B protein expression in 22Rv1 cells in a time-dependent manner (p<0.05), suggesting inhibition of autophagic flux. Chou-Talalay analysis indicated a potential synergism at 1 to 30μM bicalutamide in combination with nebivolol 10 to 30μM, with combination index<1. Conclusion: Nebivolol inhibited PCa cell growth and clonogenic potential, and inhibited autophagic flux with increased LC3B expression. Furthermore, nebivolol enhanced growth inhibition when combined with bicalutamide. These findings support further investigation of nebivolol, with/without bicalutamide, in PCa.