Targeted Next-Generation Sequencing of Flat Urothelial Lesions Reveals Putative Pathobiological Pathways, Potential Biomarkers, and Rational Therapeutic Targets

Flat urothelial lesions are controversial diagnostic and prognostic urologic entities whose impor-tance relies mainly on their ability to progress to muscle-invasive tumors via urothelial carcinoma in situ (CIS). However, the carcinogenetic progression of preneoplastic flat urothelial lesions is not well established. Moreover, predictive biomarkers and therapeutic targets of the highly recurrent and aggressive urothelial CIS lesion are lacking. Using a targeted next-generation sequencing (NGS) panel of 17 genes directly involved in bladder cancer pathogenesis, we investigated alterations of genes and pathways with clinical and carcinogenic implications on 119 samples of flat urothelium, including normal urothelium (n = 7), reactive atypia (n = 10), atypia of unknown significance ( n = 34), dysplasia ( n = 23), and CIS (n = 45). The majority of the flat lesions were tumor-associated but grossly/microscopically or temporally separated from the main tumor. Mutations were compared across flat lesions and concerning the concomitant urothelial tumor. Associations between genomic mutations and recurrence after intravesical bacillus Calmette-Guerin treatment were estimated with Cox regression analysis. TERT promoter mutations were highly prevalent in intraurothelial lesions but not in the normal or reactive urothelium, suggesting that it is a critical driver mutation in urothelial tumorigenesis. We found that synchronous atypia of unknown significance-dysplasia-CIS lesions without concomitant papillary urothelial carcinomas had a similar genomic profile that differed from atypia of unknown significance-dysplasia lesions associated with papillary urothelial carcinomas, which harbored significantly more FGFR3, ARID1A, and PIK3CA mutations. KRAS G12C and ERBB2 S310F/Y mutations were exclusively detected in CIS and were associated with recurrence after bacillus Calmette-Guerin treatment (P = .0006 and P = .01, respectively). This targeted NGS study revealed critical mutations involved in the carcinogenetic progression of flat lesions with putative pathobiological pathways. Importantly, KRAS G12C and ERBB2 S310F/Y mutations were identified as potential prognostic and therapeutic biomarkers for urothelial carcinoma.(c) 2023 United States & Canadian Academy of Pathology. Published by Elsevier Inc. All rights reserved.