Abstract 1723: The BH4 domain of Bcl-2 is sufficient to induce RNA expression profile changes in human prostate cancer cell lines.

Abstract

Abstract Bcl-2 is the archetypal gene and first characterized member of the family bearing its name. Prostate cancer was the first solid tumor malignancy in which an elevation of cellular Bcl-2 levels was associated with advanced disease. It is thought that the anti-apoptotic activity of Bcl-2 contributes to the uniformly lethal nature of castration-resistant advanced prostate cancer. Bcl-2 is a validated target for cancer therapies. Current small molecule drug inhibitors mimic the BH3 domain of pro-apoptotic Bcl-2 family members, binding to a pocket composed mainly of Bcl-2 homology (BH) domains 1-3, and only partially BH4. Surprisingly, while these agents are effective inducers of apoptosis, they do not inhibit the oncogenic functions of the BH4 domain. The BH4 domain is essential for the antiapoptotic functions of Bcl-2, and appears to mediate interactions with proteins not part of the Bcl-2 family. This explains the small but significant body of evidence supporting an oncogenic signaling role for Bcl-2 driven by the BH4 domain. The full extent of the non-apoptotic roles of Bcl-2, including the contribution of the BH4 domain, has yet to be clarified. Our work aims to address this in prostate cancer cells, developing and utilizing a system that will implicate both the full length gene product and the BH4 domain. Eukaryotic expression constructs were designed to contain the full length Bcl-2 gene built to co-express with GFP, or a fusion gene composed of the Bcl-2 BH4 domain, GFP, and the Bcl-2 C-terminal transmembrane domain. These plasmids were then transfected into the 22Rv1, LNCaP, and LAPC4 human prostate cancer cell lines to generate stable expression clones. The clones were validated for protein expression by western blot prior to a determination of classical functionality by treatment with apoptosis-inducing agents. RNA was then isolated from the clones for expression array analysis. We successfully generated human prostate cancer cells stably overexpressing Bcl-2 and the Bcl-2 BH4 domain. These clones displayed enhanced resistance to apoptosis induction. Bcl-2 overexpression also induced expression profile changes in these clones. This was mimicked by expression of the BH4 domain alone. We have developed a useful system to help enhance our understanding of Bcl-2 by highlighting changes in RNA expression profiles induced by the gene's overexpression. We also show that the BH4 domain drives much of this function. The mechanisms responsible for these Bcl-2/BH4-induced changes warrant further investigation. Citation Format: Jennifer R. Cohen, PhD, Simon A. Williams, PhD. The BH4 domain of Bcl-2 is sufficient to induce RNA expression profile changes in human prostate cancer cell lines. [abstract]. In: Proceedings of the 104th Annual Meeting of the American Association for Cancer Research; 2013 Apr 6-10; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2013;73(8 Suppl):Abstract nr 1723. doi:10.1158/1538-7445.AM2013-1723