Introduction: Androgen deprivation therapy (ADT) is the prevalent first line therapy against advanced prostate cancer as an adjuvant for locally treating the high-risk disease. Malignant cells initially respond to ADT, but subsequently colonize and re-emerge as more aggressive phenotype known as castration resistant prostate cancer (CRPC). The second generation androgen receptor (AR) antagonist enzalutamide (ENZU) exhibits survival advantage in CRPC patients, however ~30% of patients develop resistance overtime activating AR in these tumors. These resistant tumors poorly respond to chemotherapy and acquire resistance partly due to enhanced aerobic glycolysis and biomass production known as the Warburg effect. Therefore, identification of an effective low-cost therapeutic alternative with fewer side effects may lead to increased survival and greatly benefit patient quality-of-life. Drug repurposing has emerged as a new option for overcoming resistance to chemotherapy. Previous studies from our laboratory (Mol Cancer Ther. 13:2288-302, 2014) has demonstrated that synergistic combination of simvastatin (SIM), a drug for the treatment of hypercholesterolemia and metformin (MET), a glucose lowering drug inhibits CRPC growth, invasiveness and migration potential with minimal effect on normal prostate epithelial cells. Here we investigate whether combination of SIM and MET could be effective in the treatment of ENZU-resistant prostate cancer cells.
Methods: Human CRPC cells C4-2B-ENZU (C4-2B enzalutamide resistant) were generated by growing C4-2B cells in progressive concentration of ENZU 5-20µM to develop resistance and maintained in 5µM ENZU in the culture medium for >20 generations. 22Rv1 cells are inherently resistant to ENZU. These cells were treated with SIM and MET individually and in combination, followed by assessment of cell viability, crystal violet assay, cell cycle analysis, migration, invasion and expression of various target genes by Western blotting.
Results: C4-2B-ENZU and 22Rv1 cells were treated individually and in a combination of SIM and MET at pharmacological dose range (500nM-4µM SIM and 250µM-2mM MET). Combination treatment with 4µM SIM and 2mM MET (SIM+MET) led to significant and synergistic inhibition of cell viability, migration, invasion and cell cycle blockade in both cell lines. The individual treatments of SIM and MET exhibited little or no effect on these cells. Furthermore, SIM+MET combination decreased the expression of AR, AR-V7, p-Akt (Ser473), p-AMPKa1/a2 (Ser-485/491) and simultaneously increased p-AMPKa1 (Thr-172) and AMPKa kinase activity in these cells.
Conclusions: Combined action of SIM and MET may be an effective regimen for treatment of ENZU-resistant tumors. This opens new therapeutic modality for castration-resistant prostate cancer patients. Source of
Funding: VA Merit Review 1I01BX002494; Department of Defense grant W81XWH-15-1-0558, W81XWH-18-1-0618 and W81XWH-19-1-0720 to SG.