Effects of MDM2 and HMG-CoA Reductase Inhibition on Epithelial to Mesenchymal Transition in Prostate Cancer

Abstract

According to the American Cancer Society, prostate cancer is the second leading cause of cancer death among men. Aggressive growth and metastasis are the main reasons for high mortality among prostate cancer patients. The proto-oncogene MDM2 controls multiple signaling pathways that are involved in cell cycle regulation and survival. MDM2 also has a major role in imparting stemness and Epithelial to Mesenchymal Transition (EMT) abilities to prostate cancer cells that are inherently less aggressive. During cancer growth, lipid metabolism also plays a pivotal role in cellular reprogramming, which eventually leads to the induction of EMT and the acquisition of stemness. It has been well documented that the intermediates of mevalonate metabolism can significantly affect gene expressions in cancer and influence the tumor microenvironment to support tumor growth and survival. At present, there is a lack of comprehensive knowledge about the use of MDM2 inhibitor RG7388 in cancers with p53 mutation, since most studies report that it is most effective in cancers with wild-type p53. Therefore, our study was designed to investigate the effects of RG7388 in p53 mutant or p53 null prostate cancer cells. Our study also explored the effect of RG7388 in combination with simvastatin, which blocks the mevalonate pathway by inhibiting the 3-hydroxy-3-methylglutaryl-coenzyme A (HMG-CoA) reductase enzyme. Simvastatin has been reported to have anticancer effects in several cancers but, so far, there is no data available regarding its use in combination with RG7388. Therefore, our study aimed to analyze the effects of RG7388, individually or in combination with simvastatin, on EMT and cancer stemness. Our experiments were conducted using DU145 and PC3 prostate cancer cells which have p53 mutant and null status respectively. Our experimental results showed a decrease in the viability of PC3 and DU145 cells, in a time and concentration-dependent manner, after treatment with RG7388 or simvastatin. The mesenchymal and stem cell markers were also downregulated after RG7388 and simvastatin combination treatment. In addition, individual and combination treatments with the above-mentioned drugs induced the disintegration of spheroids and eventually a reduction in the total number of spheroids. Immunofluorescence staining of spheroids revealed a downregulation of cancer stem cell markers among all treated groups. Furthermore, the scratch assay showed inhibition of migration of the DU145 and PC3 cells, particularly after the combination treatment. Our results have confirmed that RG7388, simvastatin, and their combination treatments can reverse EMT, inhibit migration, and reduce cancer stemness in p53 mutant or null prostate cancer cells. Our study also suggests that RG7388 and simvastatin combination can be effective for treating prostate cancers that are aggressive due to MDM2 expression.