Simultaneous Targeting Of Endoplasmic Reticulum Stress And Akt Pathways As A Novel Chemosensitization Approach Against Castration Resistant Prostate Cancer
Docetaxel (DTX)-based regimen is the mainstay treatment against castration resistant prostate cancer (CRPC). However, significant side-effects of DTX mandate that strategies to chemosensitize CRPC cells be utilized. We investigated whether physiologically achievable concentrations of nelfinavir (NFR) and curcumin (CUR), known to target the endoplasmic reticulum (ER) stress and AKT pathways, can increase DTX cytotoxicity. A significant reduction (~70%) in survival of a CRPC cell line, C4-2B, was evident within 24 hrs post-exposure to a combination of DTX (10 nM), NFR (5 Ã‚ÂµM) and CUR (5 Ã‚ÂµM), as compared to DTX alone (~34%). This rapid cytotoxicity was not seen in non-tumorigenic RWPE-1 cells as well as in primary prostate epithelial cells (PrEC) and bone-marrow mesenchymal stem cells (BM-MSC). A significant increase in apoptosis was seen in C4-2B cells but not RWPE-1 cells, as indicated by DNA-fragmentation, caspase-3 assay, and PARP cleavage. A significant reduction in C4-2B-derived colony forming units (CFU) was observed following exposure to DTX-NFR-CUR combination (92%), as compared to DTX alone (34%). In C4-2B cells, immunodetection and real-time PCR studies showed that exposure to 3-drug combination drastically reduced AKT activation, increased unfolded protein response (UPR) markers, such as XBP-1 mRNA and phosphorylated eIF-2ÃŽ±, and increased ER-stress induced pro-apoptotic markers such as CHOP, ATF4 and TRIB3. In RWPE-1 cells, upregulation of CHOP was observed with DTX-NFR-CUR combination, but no increase in ATF-4 and TRIB3 were observed. In vivo studies using C4-2B tumor xenografts showed a significant reduction in tumor volume following 4 week exposure to the 3-drug combination, as compared to DTX alone. Immunohistochemistry (IHC) of tumor sections revealed decreased Ki-67 staining indicating reduced cell proliferation and increased TUNEL staining indicating apoptosis, in DTX-NFR-CUR treated mice as compared to DTX alone. Therefore, our studies show that NFR and CUR can provide a promising approach as an adjuvant therapy to chemosensitize CRPC to DTX therapy.