Abstract 4227: Pten and P53 control self-renewal ability and differentiation potential of prostate epithelial stem/progenitor cells

Pten and P53 tumor suppressors are among the most commonly inactivated genes in prostate cancer. Pten deletion in the mouse prostate epithelium results in the development of carcinoma in situ which eventually progresses to adenocarcinoma, while loss of P53 alone does not result in an obvious phenotype. The combination of PTEN and P53 loss leads to significantly more rapid and extensive development of prostate tumors and death due to urinary outflow obstruction. Pten and P53 together have been implicated as co-regulators of neural and glioma stem/progenitor cell renewal and differentiation. We hypothesize that Pten and P53 play a crucial role in regulating self-renewal and differentiation ability, two defining hallmarks of stem/progenitor cells. In order to test our hypothesis, cells were isolated from Pten−/−P53−/− adenocarcinomas and employed in a variety of assays. Clonogenic 2D colony and 3D sphere formation in minimal media measure progenitor activity. Colony and sphere formation activity was monitored over at least 4 generations of serial propagation in vitro. Protospheres generated from Pten−/−P53−/− single cell suspensions were approximately 3 times larger than wild type (wt) spheres and demonstrated significant heterogeneity in morphology, displaying less uniform cell shapes. Sphere forming units (SFU) in single cell suspensions from Pten−/−P53−/− prostates showed a 50%-80% increase when compared to wt. Pten−/−P53−/− SFU activity was maintained upon serial passage of single cell suspensions for 4 generations, while a reduction in SFU activity was observed over the same 4 generations in wt. Similar findings were obtained from analyzing the colony forming ability of cells taken from wt and Pten−/−P53−/− spheres. Pten and P53 deletion significantly affects the differentiation profile of colonies and spheres generated from knockout as compared to wt cells. Pten−/−P53−/− and wt protospheres (3D) and colonies (2D) were stained for markers of all prostate epithelial cell lineages using CK5, CK8, p63 and neuroendocrine markers. Interestingly, an increased number of luminal, transit-amplifying and neuroendocrine cells with a relative reduction in the number of basal cells were observed in the Pten−/−P53−/− spheres and colonies compared to wt. In addition to increased progenitor cell numbers and alterations in differentiation potential, Pten−/−P53−/− progenitor cells demonstrated dependence upon distinct signal transduction pathways for survival as revealed by their sensitivity to drugs against mTOR (Rapamycin), AKT (Triciribine) and androgen receptor (Nilutamide and Bicalutamide). In conclusion, our findings to date confirm the presence of an amplified stem/progenitor cell population in Pten−/−P53−/− prostates that generates morphologically abnormal protospheres displaying changes in the relative production of differentiated progeny. 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 4227.