Hypoxia-inducible factors drive miRNA-mediated downregulation of L2HGDH and HIF1A in clear cell renal cancer independent of 14q deletion

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 was found to portend a strikingly worse prognosis in a multivariate survival analysis of ccRCC patients (n=509; TCGA) highlighting its translational importance, the lower expression of HIF1A in ccRCC despite the upregulated HIF1 protein (secondary to VHL loss) is of significant scientific interest. Using a comprehensive array of in vitro assays with genetic manipulation, clinicogenomic bioinformatic analyses of large ccRCC datasets, and the Cdh16-Cre Vhl-fl mouse model with kidney specific Vhl-loss, we demonstrate that the downregulation of both L2HGDH and HIF1A in ccRCC is driven by the hypoxia-inducible factors via miR21-3p and miR155-5p, respectively, during 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 and HIF1 in these metabolic-epigenetic aberrations. Translational StatementThis study demonstrates that hypoxia-inducible factors (HIFs) drive early and sustained metabolic and epigenetic dysregulation in clear cell renal cell carcinoma (ccRCC). Notably, these aberrations begin early in carcinogenesis. We show that inhibition of HIFs can reverse these pathological changes - most significantly, restoring levels of 5-hydroxymethylcytosine, a key epigenetic marker lost in ccRCC due to metabolic inhibition of TET enzymes. Our findings support the integration of HIF2 inhibition earlier in the therapeutic approach for advanced ccRCC. Moreover, the data suggest that dual inhibition of HIF2 and HIF1 - particularly in tumors with high expression of both factors - may yield a more robust reversal of epigenetic abnormalities and merits further investigation. Graphical Abstract O_FIG O_LINKSMALLFIG WIDTH=189 HEIGHT=200 SRC="FIGDIR/small/625494v2_ufig1.gif" ALT="Figure 1"> View larger version (85K): org.highwire.dtl.DTLVardef@a75aborg.highwire.dtl.DTLVardef@b36704org.highwire.dtl.DTLVardef@3057c4org.highwire.dtl.DTLVardef@1d24074_HPS_FORMAT_FIGEXP M_FIG C_FIG