Abstract
Background: Approximately 80% of prostate cancer patients undergoing hormone therapy progress within 12-18 months to a hormone insensitive form of the disease known as hormone refractory prostate cancer. The median survival rate is less than 12 months and traditional therapeutic agents have shown little effect on the progression of the disease. The selective estrogen receptor modulator (SERM), raloxifene has been examined in a limited phase ІІ clinical trial and showed anti-tumor properties. The current study investigated the activity of raloxifene on expression of ERα, ERβ, AR, EGFR and caveolin-1. Also the study aimed to see whether synergistic cytotoxicity could be obtained administering raloxifene in combination with the curcumin analogue RL91.
Methods: Two different hormone refractory prostate cancer cell lines PC3 and DU-145 were used in the study. PC3 cells were specifically used to understand the role of raloxifene 10 and 15 μM on different receptors such as estrogen receptor α (ERα), estrogen receptor β (ERβ), androgen receptor (AR), epidermal growth factor receptor (EGFR) and caveolin-1. Since raloxifene is a SERM it should primarily target estrogen receptors. We wanted to determine the effect of raloxifene on different receptors. Immunocytochemistry and Western blotting was carried out to analyze the change in receptor localization and quantitative protein expression levels, respectively. To assess the synergistic potential of the combinational therapy, raloxifene 5, 10 μM and RL91 1.5 and 2 μM were measured individually and in combination on PC3 and DU-145 using sulforhodamine B assay.
Results: Raloxifene treatment affected the localization of ERβ, EGFR, AR and caveolin-1 in PC3 cells. The drug was shown to increase the translocation of ERβ and EGFR after 6 h of treatment with a maximum difference observed after 48 h. Also, a decreased expression of ERβ and EGFR was shown following 48 h of raloxifene treatment. It also decreased the co-localization of the EGFR and caveolin-1. Apart from this, it also induced vesicle formation and accumulation of cellular content inside the cells. The cytotoxicity assay showed more cytotoxic potential of RL91 compared to raloxifene, however the combination showed synergism with more than 80 % of cell death observed for both PC3 and DU-145 cells following raloxifene 10 μM and RL91 1.5 μM treatment.
Conclusions: In HRPC, ER and EGFR mediated signaling is important for the proliferation of the cells. Raloxifene’s modulation of ERβ, EGFR and AR develops a stressed condition for the cells which decreased the proliferation. By this mechanism raloxifene enhances the cytotoxicity elicited by RL91 when administered in combination.