Use of 3' Rapid Amplification of cDNA Ends (3' RACE)-Based Targeted RNA Sequencing for Profiling of Druggable Genetic Alterations in Urothelial Carcinomas.

Targeted treatment of advanced or metastatic urothelial carcinomas (UCs) requires the identification of druggable mutations. This study describes the development of a 3' Rapid Amplification of cDNA Ends (3' RACE)-based targeted RNA sequencing panel which accounts for the status of all genes relevant to UC treatment, namely, , , , , (), () and . -activating point mutations or fusions were found in 54/233 (23.2%) tumors. rearrangements were identified in 11 patients, with eight of them being undetectable by commonly used PCR kits. In addition, one tumor contained a high-copy gene amplification accompanied by strong overexpression of the gene. Mutations in genes were present in 30/233 (12.9%) UCs and were mutually exclusive with alterations affecting genes. On the contrary, activating events in the oncogene (point mutations and overexpression), as well as mutations, which were relatively common, occurred with similar frequencies in - or -positive vs. negative samples. High mRNA expression was associated with advanced disease stage and was not observed in tumors with increased mRNA expression or in UCs with evidence for activation. Three of the studied carcinomas had high-level microsatellite instability (MSI). Overall, more than half of the UCs had potentially druggable genetic alterations. The proposed NGS panel permits comprehensive and cost-efficient analysis of UC-specific molecular targets and may be considered in clinical routine.