Aberrant activation of m6A demethylase FTO renders HIF2α ^low/− clear cell renal cell carcinoma sensitive to BRD9 inhibitors

Abstract

Hypoxia-inducible factor 2α (HIF2α) antagonists are effective against clear cell renal cell carcinomas (ccRCCs) that highly express HIF2α. To identify potential drug targets in HIF2α^low/− ccRCC, we constructed an epigenetic-focused single-guide RNA library and performed an in vivo CRISPR-Cas9 knockout screen in BALB/c nude mice transplanted with 786-O (HIF2α^high) or Caki-2 (HIF2α^low/−) cells. We found that the m6A demethylase fat mass and obesity-associated (<i>FTO</i>) gene was indispensable to the growth of HIF2α^low/− but not HIF2α^high ccRCC. Activation of FTO in HIF2α^low/− ccRCC was caused by an increased intracellular α-ketoglutarate–to-succinate ratio and stabilized bromodomain-containing protein 9 (<i>BRD9</i>) messenger RNA via m6A demethylation. RNA sequencing and chromatin immunoprecipitation sequencing profiling further revealed that SRY-box transcription factor 17 (SOX17) recruited BRD9 to de novo super enhancers associated with genes that feature prominently in ccRCC pathogenesis, including <i>CCND1</i>, <i>VEGFR2</i>, <i>CDC20</i>, <i>SRC</i>, and <i>MAPK6</i>. <i>BRD9</i> knockdown or the BRD9-selective antagonist I-BRD9 suppressed the growth of HIF2α^low/− but not HIF2α^high ccRCC cells in vitro. In BALB/c nude mice bearing HIF2α^low/− ccRCC cell line–derived xenografts and patient-derived tumor xenografts, I-BRD9 administration effectively inhibited tumor growth and prolonged the survival of tumor-bearing mice with greater efficacy than sunitinib. Together, these findings indicate that BRD9 is a druggable target for treating HIF2α^low/− ccRCC.