ANDROGEN RECEPTOR REGULATION IN PROSTATE STROMA

Abstract

Prostate cancer is the second leading cause of cancer-related deaths in men. The loss of stromal androgen receptor (AR) expression has been associated with an increasing Gleason grade and the development of aggressive prostate cancer. Stromal AR expression is a critical determinant of normal prostate gland differentiation, mediated through androgen-induced expression and subsequent secretion of stromal paracrine factors. However, the mechanisms leading to AR loss in the stroma are not well understood. The aim of this thesis was to test the hypothesis that tumor-secreted factor(s) represses AR expression in the tumor stroma through transcriptional repressive signal transduction pathways, leading to the loss of epithelial differentiation factors that normally maintain epithelial homeostasis. Using cytokine array profiling, TGFβ1 and TNFα were identified as common factors secreted by two different prostate cancer cell lines. I used pharmacological inhibitors and siRNA/shRNA knockdown approaches to identify the signaling pathway(s) involved in TNFα-induced down-regulation of stromal AR expression at the mRNA level. I found that shRNA mediated knock-down, as well as inhibitors of NF-κB blocked the ability of TNFα to downregulate AR protein and mRNA. Interestingly, I also discovered that p38α- and p38δ-MAPK, acting independently of TNFα, acts as intrinsic negative regulators of AR expression. Furthermore, I identified two regulatory regions within the AR promoter containing known and putative NF-κB and CREB/ATF binding elements potentially responsible for the suppression by NF-κB and p38-MAPK respectively. To determine the consequence of AR loss in the stroma, I identified Wnt16 and FGF10 as secreted epithelial cell differentiation factors whose expression is increased in stromal fibroblasts following androgen treatment. Treatment with TNFα caused loss of both Wnt16 and FGF10 expression, while blocking p38-MAPK enhanced Wtn16 and FGF10 expression, confirming the dependency on AR for their expression. I further demonstrated that Wnt16 is crucial for the survival of prostate basal epithelial cells through non-canonical Jnk signaling. Finally, I demonstrated that co-culturing stromal cells with tumor cells in a new prostate-on-chip model, causes the same downregulation of AR expression and that TNFα, TGFβ, and p38-MAPK similarly sup-press AR expression in primary stromal cells isolated from patients. These studies pave the way for determining if analysis of these pathways can be used as biomarkers to predict patient outcome.