Nebivolol exhibits cell growth inhibition in drug-resistant HR+/HER2- breast cancer and anti-tumoral synergism with abemaciclib in vitro

Background: We are investigating nebivolol, a third-generation beta-blocker, as a drug-repurposing candidate for breast cancer (BC). While prior studies have focused on triple-negative BC, we evaluated its efficacy and potency in hormone receptor-positive, HER2-negative (HR+/HER2-) BC, which is the most prevalent subtype. Since HR+/HER2- BC is typically treated with endocrine therapy (ET) alone or in combination with CDK4/6 inhibitors (CDK4/6i), we also assessed the effects of nebivolol in ET- and CDK4/6i-resistant models, as well as in combination with ET and CDK4/6i. Methods: The efficacy and potency (IC50) of nebivolol were evaluated using MCF7 and T47D parental cells and their estrogen deprivation-resistant (EDR), palbociclib-resistant (PalboR), and EDR/PalboR derivatives. Cells were treated with vehicle or nebivolol (0.01 to 30 μM). Efficacy (% inhibition at 30 μM) and IC50 of nebivolol were determined using a three-parameter non-linear regression model using GraphPad Prism v.10. For evaluating drug combinations, concentration ranges of the drugs (nebivolol with palbociclib or abemaciclib) were selected based on their estimated IC50 values in the concentration-response matrices. Growth inhibition was quantified using the EnSight TM Multimode Plate Reader, and drug interaction was analyzed using CompuSyn (combination index, CI) and SynergyFinder (Loewe synergy score). A parallel analysis was performed to determine the statistical significance between combination and single-agent groups using One-way ANOVA, followed by Tukey multiple comparison test. Results: Nebivolol inhibited growth in parental and ET- and CDK4/6i-resistant MCF7 and T47D cells in a concentration-dependent manner. Efficacy (%) and potency (μM) of nebivolol were as follows: MCF7-P [84.1 and 7.9]; EDR [79.1 and 12.3]; PalboR [73.5 and 13]; EDR/PalboR [68.4 and 20.4]; T47D-P [96.1 and 5.8]; EDR [94.6 and 5.4]; PalboR [86.7 and 5.7]; EDR/PalboR [91.6 and 5.2]. Additionally, nebivolol demonstrated synergism with abemaciclib in Chou-Talalay and Loewe analysis. Effective combinations included 1 or 10 μM and 0.3-10 μM of nebivolol with 30-300 nM of abemaciclib (CI < 1 in Chou-Talalay method) in MCF7P and T47DP, respectively, suggesting synergy. Significant % inhibition in combination compared to single agent alone was observed at 3 μM of nebivolol with 100 or 300 μM abemaciclib in MCF7P and 0.3-3 μM of nebivolol with 30 nM of abemaciclib in T47DP cells. Loewe analysis also revealed nebivolol-abemaciclib synergism, with the most synergistic area score of >10 (12.51 in MCF7P and 41.14 in T47DP). Conclusion: Nebivolol inhibited the growth of HR+/HER2- BC cell lines sensitive or resistant to ET and/or CDK4/6i and synergized with abemaciclib. Ongoing studies are evaluating nebivolol and abemaciclib in other cell-based assays and combination with ET for enhanced therapeutic potential.