Restoration of BAP1 activity via base editing suppresses anchorage-independent survival in kidney cancer.

BAP1, a deubiquitinase with a ubiquitin C-terminal hydrolase domain, functions as a tumor suppressor involved in diverse cellular processes, including DNA repair, genome stability, and apoptosis. Inactivating mutations in BAP1-particularly missense and deletion variants-are recurrent across multiple cancers, with a high prevalence in clear cell renal cell carcinoma (ccRCC), mesothelioma, and uveal melanoma. Among these, the Glu31Lys mutation in ccRCC impairs BAP1's enzymatic activity, protein stability, and DNA repair functions. Here, we investigated the physiological impact of this recurrent mutation using isogenic KMRC-20 ccRCC cell clones in which the Glu31Lys substitution was precisely corrected to wild-type glutamate via CRISPR-Cas9-mediated adenine base editing. BAP1 reactivation restored anchorage-dependent growth in KMRC-20 cells-a hallmark of non-transformed epithelial cells-and increased apoptosis under non-adherent conditions, indicating reinstated sensitivity to anoikis. Mechanistically, this phenotypic switch was accompanied by post-transcriptional downregulation of N-cadherin and β-catenin under anchorage-free conditions, implicating BAP1 in the regulation of adhesion- and Wnt-related survival pathways. Furthermore, transcriptomic profiling revealed broad gene expression changes upon BAP1 restoration, suggesting that these combined alterations contribute to the re-establishment of anchorage-dependent growth in KMRC-20 cells. These findings uncover a previously unrecognized role for BAP1 in suppressing anchorage-independent survival, providing new insights into BAP1-driven tumorigenesis and underscoring the therapeutic potential of precise gene editing to restore tumor suppressor function in ccRCC.