The Role of Pparg in Urothelial Carinoma

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Bladder cancer is currently the 6th most common cancer in the United States, resulting in 17,000 deaths annually. Clinically, bladder cancers are mostly urothelial carcinoma, classified as either non-muscle invasive bladder cancer (NMIBC) or muscle-invasive bladder cancer (MIBC), with the latter having a 5-year survival rate of merely 50%. With recent advances in next-generation sequencing, several international consortia have elucidated molecular subtypes of MIBC. The two major subtypes of MIBCs are basal and luminal; the basal subtype frequently exhibits hallmarks of squamous differentiation and highly expressed basal markers (CD44, KRT14, KRT6A, KRT6B). Tumors of the luminal subtype have papillary morphology and highly express differentiation-associated luminal markers (e.g., KRT20, PPARG, UPKs, and FOXA1). Notably, the transcription factor Peroxisome Proliferator Activated Receptor Gamma (PPARG) gene is frequently amplified in luminal MIBC. And recurrent activating mutations have been reported for its obligatory functional partner Retinoic X Receptor (RXR). In addition, the basal subtype is immune-infiltrated and is postulated as more likely to respond to immunotherapies. In contrast, the luminal subtype is immune-cold. Despite these advances in recent years, the molecular driver of subtype determination, specifically in luminal MIBC, remains poorly understood. Furthermore, subtype-specific targeted therapy for MIBCs is still in its infancy. Our previous work determined that Pparg activation can drive luminal tumor formation. We generated a novel Krt5CreERT2; VP16;Pparg transgenic mouse model, where Pparg expression is constitutively active in basal urothelial cells upon tamoxifen induction. During homeostasis, constitutive Pparg promoted luminal differentiation and cell cycle exit in basal cells but did not produce tumors. However, increased Pparg signaling in activated basal cells following 1-month exposure to bladder-specific carcinogen BBN produced luminal tumors. These tumors are similar both in morphology and molecular markers to human luminal MIBCs. The resulting VP16;Pparg luminal tumors have reduced Nf-kb expression and are immune cold compared to basal tumors. These findings suggest that Pparg is a driver of luminal tumor formation and a suppressor of immune infiltration in bladder cancer. In Chapter 2 of the thesis, I focus on the therapeutic potential of activating Pparg in basal MIBC. We treated mice bearing BBN-induced, Pparg-negative basal tumors with synthetic Pparg ligand - Rosiglitazone (Rosi) and Mek1/2 inhibitor Trametinib (Tram), both of which have been shown to induce Pparg signaling in vitro and in vivo. The combined RosiTram treatment induced apoptosis and significantly reduced tumor burden. The post-treatment urothelium appeared similar in morphology to a healthy urothelium. RosiTram treatment also restored normal urothelial differentiation and generated resident cell types (e.g., superficial cells, intermediate cells, Keratin5+ (K5), basal cells, and Keratin14+ (K14) basal cells) that are normally seen in a healthy urothelium. In contrast, basal tumors are almost entirely composed of K14-Basal cells. Mechanistically, RosiTram treatment partially restores differentiation through retinoic acid signaling and Ezh2 inhibition. Together, our study established targeted transcriptionally and epigenetically reprogramming as a promising differentiation therapeutic approach for basal bladder tumors.