Regulation of FOXO3a/β-Catenin/GSK-3β Signaling by 3,3′-Diindolylmethane Contributes to Inhibition of Cell Proliferation and Induction of Apoptosis in Prostate Cancer Cells
Yiwei Li, Zhiwei Wang, Dejuan Kong, Shalini Murthy, Q. Ping Dou, Shijie Sheng, G. Prem Veer Reddy, Fazlul H. Sarkar
Journal of Biological Chemistry
Previous studies from our laboratory have shown anti-proliferative and pro-apoptotic effects of 3,3-diindolylmethane (DIM) through regulation of Akt and androgen receptor (AR) in prostate cancer cells. However, the mechanism by which DIM regulates Akt and AR signaling pathways has not been fully investigated. It has been known that FOXO3a and glycogen synthase kinase-3␤ (GSK-3␤), two targets of activated Akt, interact with ␤-catenin, regulating cell proliferation and apoptotic cell death. More
... mportantly, FOXO3a, GSK-3␤, and ␤-catenin are all AR coregulators and regulate the activity of AR, mediating the development and progression of prostate cancers. Here, we investigated the molecular effects of B-DIM, a formulated DIM with higher bioavailability, on Akt/FOXO3a/GSK-3␤/␤-catenin/AR signaling in hormone-sensitive LNCaP and hormoneinsensitive C4-2B prostate cancer cells. We found that B-DIM significantly inhibited the phosphorylation of Akt and FOXO3a and increased the phosphorylation of ␤-catenin, leading to the inhibition of cell growth and induction of apoptosis. We also found that B-DIM significantly inhibited ␤-catenin nuclear translocation. By electrophoretic mobility shift and chromatin immunoprecipitation assays, we found that B-DIM inhibited FOXO3a binding to the promoter of AR and promoted FOXO3a binding to the p27 KIP1 promoter, resulting in the alteration of AR and p27 KIP1 expression, the inhibition of cell proliferation, and the induction of apoptosis in both androgen-sensitive and -insensitive prostate cancer cells. These results suggest that B-DIM-induced cell growth inhibition and apoptosis induction are partly mediated through the regulation of Akt/FOXO3a/GSK-3␤/␤-catenin/AR signaling. Therefore, B-DIM could be a promising non-toxic agent for possible treatment of hormone-sensitive but most importantly hormone-refractory prostate cancers. Prostate cancer has become a significant health problem in the United States because of its high incidence and mortality (1). Despite an initial efficacy of androgen-deprivation therapy, most patients with prostate cancer progress from androgen-dependent status to hormone-refractory prostate cancer (HRPC) 2 for which there is no curative therapy. It is still unclear how the prostate cancer cells progress to androgen independence. However, more evidence indicates that androgen receptor (AR) and Akt pathways participate in the development of HRPC (2, 3). It has been known that during the progression of prostate cancers from androgen-sensitive status to an androgen-independent stage, the majority of prostate cancer cells still expresses AR, suggesting that AR signaling plays a critical role in the development and progression of prostate cancer (4). In prostate epithelial cells, ligand-free AR is sequestered in the cytoplasm bound to heat shock proteins. Binding of androgen to the AR induces a AR conformational change, which allows dissociation of heat shock proteins. The AR then forms a homodimer and is phosphorylated. This phosphorylation stabilizes the ligand-AR complex and the complex translocates to the nucleus (5). The activated AR then initiates gene transcription by binding to specific androgen-response elements in the promoter regions of target genes. Evidence is emerging showing that phosphorylation of the AR by other molecules in cell signaling pathways can also influence AR transactivation (6). It has been found that Akt can phosphorylate the AR at Ser-210/ 213 and Ser-790/791 and transactivate the activity of AR (7, 8). The phosphorylation by Akt also sensitizes the AR to low circulating levels of androgen, such as those present during maximum androgen blockade (9). This sensitization allows low levels of androgen to induce phosphorylation at specific sites required for translocation of the AR to the nucleus. Therefore, Akt is another activator of AR required for androgen-independent survival and growth of prostate cancer cells. Akt can be activated by the lipid kinase phosphatidylinositol 3-kinase (10). The activated Akt can inhibit apoptosis by direct Authors are urged to introduce these corrections into any reprints they distribute. Secondary (abstract) services are urged to carry notice of these corrections as prominently as they carried the original abstracts.