Seal, S.; Luchtel, R. A.; Akram, T.; Isaila, B.; Perry, K. T.; Procissi, D.; Hafizi, S.; Shenoy, N. K.

Clear cell renal cell carcinoma (ccRCC) is characterized by pseudohypoxic signaling as well as metabolic and epigenetic aberrations of increasing severity during its pathogenesis and progression. Upon identifying that the markedly lower expression of L2HGDH and HIF1A in ccRCC tumors is not explained only by 14q deletion (40% of ccRCC), we sought to elucidate the mechanisms underscoring their down-regulation independent of 14q deletion, given the marked translational and scientific importance. While lower L2HGDH expression was found to portend a strikingly worse prognosis in a multivariate survival analysis of ccRCC patients (n=509; TCGA) highlighting its translational importance, the paradoxical downregulation of HIF1A mRNA in ccRCC despite the upregulation of HIF1 protein (secondary to VHL loss) is of significant scientific interest. Using a comprehensive array of in vitro assays, clinicogenomic bioinformatic analyses of large ccRCC datasets, and the Cdh16-Cre Vhl-fl mouse model with kidney specific Vhl-loss, we show that the hypoxia-inducible factors upregulate miR21-3p and miR155-5p, which subsequently target L2HGDH and HIF1A respectively, during ccRCC pathogenesis and progression. In doing so, we unveil a conserved inhibitory feedback loop for HIF1A in ccRCC (HIFs[->]miR155[->]HIF1A), providing a mechanistic explanation for HIF1A downregulation despite elevated HIF1 protein. Furthermore- and most importantly- we show that inhibition of hypoxia-inducible factors reverses the adverse loss of L2HGDH and hydroxymethylcytosine in ccRCC, implicating both HIF2 (primarily) and HIF1 in these metabolic-epigenetic aberrations.