Targeting MYC-inducing enhancer-associated noncoding (MYC-IEANC) RNAs inhibits the proliferation of HCC cells

Abstract Background MYC, a critical oncogene, encodes the c-MYC transcription factor (TF) and plays an essential role in hepatocellular carcinoma (HCC) development. Recent studies have identified numerous tissue-specific enhancers of MYC in various cancers, but an HCC-specific enhancer of MYC remains elusive. Methods We analyzed enhancer markers, including H3K27ac enrichment and enhancer RNA (eRNA) expression, to determine putative enhancer regions of MYC in HCC cells. Enhancer activity was detected using a dual-luciferase reporter assay. We used the CRISPR-Cas9 system to edit the enhancer regions and performed antisense oligonucleotide (ASO) to inhibit eRNA. The functions of enhancers and eRNAs on HCC cells were confirmed by cell proliferation assay and sphere formation assay. Results We choose two active enhancers R2 and R3, with high activity among six putative enhancer regions by analyzing enhancer markers. Enhancer R2 and R3 are present approximately 800 kb downstream from the MYC gene. We confirmed eRNA activities in the enhancer regions. Depletion of these enhancer regions inhibited eRNAs significantly reduced MYC expression. In addition, MYC enhancers and eRNAs regulated HCC cell proliferation and progression. Conclusion In this study, we present MYC enhancers in HCC and elucidate the molecular functions of MYC-inducing enhancer-associated noncoding (MYC-IEANC) RNAs in the proliferation of HCC cells. Furthermore, our results suggest that MYC-IEANC RNAs, which play an oncogenic role in HCC cells, can be a target for HCC treatment.