RNA sequencing of Metastatic urothelial carcinoma (UC)

Histological variation is a common feature of metastatic cancer but its clonal origins and impact on disease history remains poorly defined. In this study, we developed a first-in-kind metastatic bladder cancer (BLCA) rapid autopsy cohort enriched in rare histological subtypes to extensively profile individuals with terminal disease. By reconstructing the evolutionary history of patient tumors, we determined that metastasis-to-metastasis seeding was the dominant driver of cancer spread and that increased polyclonal migration contributed to poor prognosis. The burden, heterogeneity, and timing of acquired genomic alterations were markedly different between histological subtypes. Plasmacytoid and neuroendocrine variants developed early genomic driver alterations that associated with shorter survival. Mutational signature analysis further revealed that plasmacytoid, but not other subtypes, utilized the Fanconi Anemia pathway to shield from chemotherapy-induced genomic scarring. Using experimental models, we demonstrated that Fanconi Anemia pathway activity positively correlated with resistance to cisplatin chemotherapy, demonstrating a molecular basis for therapy resistance in plasmacytoid patients. We report the first single-nucleus profiling of mBLCA histological subtypes, revealing mixed cell states in plasmacytoid tumors and that increased overall transcriptional heterogeneity was associated with longer patient survival. Characterization of the tumor microenvironment uncovered distinct patterns of immune activation across histological subtypes, with plasmacytoid tumors exhibiting an immune-inflamed profile, while squamous subtype was dominated by immunosuppressive interactions. Lastly, we demonstrated that post-mortem cell-free DNA captured both genomic heterogeneity and transcriptional activity of the subtypes, providing a potential strategy to non-invasively assess tumor identity and aggressiveness in patients with mBLCA. Our results provide new principles into how tumor heterogeneity shapes the evolutionary history of cancer progression within subtypes of human bladder cancer