Combination therapy with novel androgen receptor antagonists and statin for castration-resistant prostate cancer.BACKGROUND
 One of the growth mechanisms of castration-resistant prostate cancer (CRPC) is de novo androgen synthesis from intracellular cholesterol, and statins may be able to inhibit this mechanism.  In addition, statins have been reported to suppress the expression of androgen receptors (ARs) in prostate cancer cell lines.  In this study, we investigated a combination therapy of novel AR antagonists and statin, simvastatin, for CRPC.


 LNCaP, 22Rv1, and PC-3 human prostate cancer cell lines were used.  We developed androgen-independent LNCaP cells (LNCaP-LA).  Microarray analysis was performed, followed by pathway analysis, and mRNA and protein expression was evaluated by quantitative real-time polymerase chain reaction and Western blot analysis, respectively.  Cell viability was determined by MTS assay and cell counts.  All evaluations were performed on cells treated with simvastatin and with or without AR antagonists (enzalutamide, apalutamide, and darolutamide).


 The combination of darolutamide and simvastatin most significantly suppressed proliferation in LNCaP-LA and 22Rv1 cells.  In a 22Rv1-derived mouse xenograft model, the combination of darolutamide and simvastatin enhanced the inhibition of cell proliferation.  In LNCaP-LA cells, the combination of darolutamide and simvastatin led to reduction in the mRNA expression of the androgen-stimulated genes, KLK2 and PSA; however, this reduction in expression did not occur in 22Rv1 cells.  The microarray data and pathway analyses showed that the number of differentially expressed genes in the darolutamide and simvastatin-treated 22Rv1 cells was the highest in the pathway termed "role of cell cycle."  Consequently, we focused our efforts on the cell cycle regulator polo-like kinase 1 (PLK1), cyclin-dependent kinase 2 (CDK2), and cell cycle division 25C (CDC25C).  In 22Rv1 cells, the combination of darolutamide and simvastatin suppressed the mRNA and protein expression of these three genes.  In addition, in PC-3 cells (which lack AR expression), the combination of simvastatin and darolutamide enhanced the suppression of cell proliferation and expression of these genes.


 CONCLUSIONS
 Simvastatin alters the expression of many genes involved in the cell cycle in CRPC cells.  Thus, the combination of novel AR antagonists (darolutamide) and simvastatin can potentially affect CRPC growth through both androgen-dependent and androgen-independent mechanisms.
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