Abstract 4073: Efficient conditional reprogramming of differentiated prostate cancer cells back to a stem cell-like state with increased aggressive properties

Abstract

Abstract Androgen receptor- (AR-) expressing prostate cancer (PCa) cell lines can transdifferentiate to neuro-ectodermal cell lineages under androgen deprivation. We have proposed that this transdifferentiation is mediated by initial regression to an unstable cancer stem cell-like (CSCL) intermediate state and we have developed a serum- and androgen-free custom cell culture medium that now allows us to efficiently reprogram PCa cells to this CSCL state and expand them for characterization. Exposure of adherent cultures of parental LNCaP, VCaP, LAPC4 or CWR22rv1 cells to the reprogramming medium induced a rapid and efficient (>67%) morphological change of plated cells to smaller, rounded cells with large nuclei and prominent nucleoli that grow in 3-dimensions. Following transfer of the reprogrammed cells they, thereafter, grow as cell rosettes that will form larger spheroids. Analysis of changes in gene expression by microarray gene expression profiling or immunohistochemistry (IHC) showed a remarkable overexpression of stem cell genes in the reprogrammed cells and a change in cell surface CD protein expression that resembles neural stem cells. Reprogramming drastically reduced expression of AR RNA and protein in all cell lines. Once reprogrammed, the CSCLs have significantly higher tumor-initiating capacity based upon tumors formed from cell serial dilutions xenografted into intact or castrated male nude mice. In vitro, reprogrammed cells were significantly more resistant to the anti-androgen enzalutamide and to hypoxia compared to parentals. By placing CSCLs into specific differentiation mediums, we could differentiate them back into neural-, oligodendrocyte-, glia- or osteoblast-like cells based upon induced expression of lineage-specific RNAs/proteins or return them to prostate-like cells (that re-express AR) by supplementing them with serum and androgen. Reprogrammed cells have an EMT phenotype based on increased expression of EMT-related genes and significantly increased invasiveness in in vitro and in vivo models. Comparison of gene expression changes acquired by reprogrammed CSCLs amongst the different cell models revealed a shared set of 65 over-/under-expressed genes (≥1.8-fold) between them. Finally, we have found that growth of parental LNCaP cells in androgen-deprived or enzalutamide-supplemented (serum-containing) growth medium increased the population of cells bearing CSCL surface CD stem markers. In summary, our studies show that commonly used PCa cell lines have the ability to efficiently transition between a differentiated and a CSCL state in an appropriate microenvironment that is depleted of androgens. While in the CSCL state, these cells show increased invasiveness, increased resistance to hormonal therapeutics and a propensity to transdifferentiate to alternate cell lineages under appropriate stimulation. Citation Format: Josselin Caradec, Amy A. Lubik, Mannan Nouri, Na Li, Manuel Altimarano-Dimas, Jennifer Bishop, Mani Moniri, Down Cochrane, Martin Gleave, Ralph Buttyan. Efficient conditional reprogramming of differentiated prostate cancer cells back to a stem cell-like state with increased aggressive properties. [abstract]. In: Proceedings of the 106th Annual Meeting of the American Association for Cancer Research; 2015 Apr 18-22; Philadelphia, PA. Philadelphia (PA): AACR; Cancer Res 2015;75(15 Suppl):Abstract nr 4073. doi:10.1158/1538-7445.AM2015-4073