Modulation of c-FLIP_L expression in prostate cancer cells by the synthetic androgen, Mibolerone, and antiandrogen, Casodex Free

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Prostate cancer is the most frequently diagnosed and the second leading cause of cancer deaths among men in North America. Tumors initially respond to androgen-ablation therapy, which inhibits the transcriptional activity of the androgen receptor (AR) to slow cell proliferation and promote apoptosis. However, most cancers eventually relapse to an androgen-depletion-independent phenotype that is often more aggressive than the original hormone-dependent tumor. Despite independence from androgen, these advanced tumor cells still rely upon the activity of the AR for proliferation and survival. Interactions of the AR with other cellular pathways, such as those that regulate apoptosis, are important in the progression to the advanced disease state. The cellular Fas/FasL-associated death domain protein-like inhibitory protein (c-FLIP) inhibits the activation of procaspase-8, thus preventing pro-apoptotic death receptor mediated signaling at the cell surface. We used two prostate cancer cell lines, LNCaP and C4-2, to examine c-FLIP mRNA and protein expression via Real-Time PCR and Western blotting, respectively, following treatment with the synthetic androgen, Mibolerone, or the anti-androgen Bicalutamide (Casodex). Basal message and protein expression levels of c-FLIP were higher in the androgen-independent (AI) C4-2 cell line than the androgen-dependent (AD) LNCaP cells. Androgenic regulation of c-FLIP message and protein was evident in LNCaP cells in both a dose- and time-dependent manner, however no such regulation was observed in C4-2 cells. Treatment of cells with Casodex alone was followed by a downregulation of c-FLIP protein that was not accompanied by discernible changes in c-FLIP message. These initial studies suggest that regulation of c-FLIP is a complex process in prostate cancer cells. In AD cells, mibolerone treatment was correlated with increases in c-FLIP protein and moderate changes in message, while AI cells did. Casodex treatment, however, was followed by a decrease of c-FLIP protein in both cell types. These data point toward an independent and novel activity of Casodex that may not be associated with its anti-androgenic activity. Future work aims to examine the mechanism of Casodex activity in c-FLIP protein downregulation and to address possibilities of exploiting this response as a potential target in prostate cancer therapy.

This research was supported in part by grants from the NIH (DK65236, CA12177, CA091956) and the T.J. Martell Foundation.