Abstract
Abstract In the current report we provide a novel finding about the regulation of Bcl-2 expression in prostate cancer (CaP) cells. Bcl-2 is known to be regulated by Sonic Hedgehog (SHH) signaling. Here, we provide evidence that human CaP cells exhibit SHH-independent Bcl-2 activation which is regulated by Bmi-1-induced Wnt signaling. Bmi-1 is a member of polycomb group gene family and is reported to be involved in stemness and self-renewal capability of cancer stem cells. We investigated the role of Bmi-1 in human CaP development and delineate its mechanism of action. On the basis of immunohistochemical analysis of prostatic specimens of 125 human CaP patients, we show that Bmi-1 protein levels are highly elevated in patients with advanced disease. To understand the mechanism of action of Bmi-1, we employed two-prong strategy. Firstly, Bmi-1 was knocked down in CaP cells (LNCaP, DU145 and PC-3) by employing siRNA technique. Bmi-1-silenced CaP cells exhibited decreased proliferative and clonogenic potential. Secondly, Bmi-1 was over-expressed in CaP cells by transfecting Bmi-1 overexpressing plasmid (pbabe-Bmi-1) in CaP cells. Bmi-1-overexpressing cells exhibited increased clonogenicity and rate of proliferation. We performed a micro-array analysis of Bmi-1-silenced cells and found that silencing of Bmi-1 caused a decrease in the Cyclin D1 (Wnt signaling target) and Bcl-2 (SHH signaling target), however an increase in p16 was observed. These studies were validated by performing immunoblot analysis. We generated a hypothesis that the Bmi-1 regulates the expression of Cyclin D1 and Bcl-2 by interacting with Wnt /SHH signaling in CaP cells. We show that despite blocking SHH signaling (by treating CaP cells with cyclopamine, a SHH signaling inhibitor), Bmi-1 induces the Bcl-2 expression in CaP cells suggesting the involvement of SHH-independent pathway in the regulation of Bcl-2 activation. We identified Wnt signaling as the pathway that regulates Bcl-2 independent of SHH signaling in CaP cells. We provide evidence that Bcl-2 promoter region of Bcl-2 gene has binding sites for Tcf transcriptional factor (Wnt signaling component). We found that Bmi-1 induces Wnt signaling in CaP cells. Taken together, these studies provide deep-insight into the mechanism of CaP cell proliferation (even after chemotherapy such as cyclopamine treatment). We suggest that Bmi-1 could be developed as a novel molecular and therapeutic target for human CaP treatment. Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 101st Annual Meeting of the American Association for Cancer Research; 2010 Apr 17-21; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2010;70(8 Suppl):Abstract nr LB-234.
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