Hypermethylation of FOXA1and allelic loss of PTENdrive squamous differentiation and promote heterogeneity in bladder cancer

Intratumoral heterogeneity in bladder cancer is a barrier to accurate molecular sub-classification and treatment efficacy. However, individual cellular and mechanistic contributions to tumor heterogeneity are controversial. We examined potential mechanisms of FOXA1and PTENinactivation in bladder cancer and their contribution to tumor heterogeneity. These analyses were complemented with inactivation of FOXA1and PTENin intermediate and luminal mouse urothelium. We show inactivation and reduced expression of FOXA1and PTENis prevalent in human disease, where PTENand FOXA1are downregulated by allelic loss and site-specific DNA hypermethylation, respectively. Conditional inactivation of both Foxa1and Ptenin intermediate/luminal cells in mice results in development of bladder cancer exhibiting squamous features as well as enhanced sensitivity to a bladder-specific carcinogen. In addition, FOXA1is hypermethylated in basal bladder cancer cell lines, and this is reversed by treatment with DNA methyltransferase inhibitors. By integrating human correlative and in vivo studies, we define a critical role for PTENloss and epigenetic silencing of FOXA1in heterogeneous human disease and show genetic targeting of luminal/intermediate cells in mice drives squamous differentiation.