Small activating RNA AW1-51 (CEBPA-51) elicits targeted DNA demethylation to promote gene activation [RNA-seq]

Small activating RNAs (saRNAs), are short double-stranded RNAs designed to upregulate transcription of target genes. By this virtue, they can be used to restore expression of genes frequently silenced in cancer. AW1-51 (also referred to as CEBPA-51), the first saRNA therapeutic to enter clinical evaluation, has demonstrated biological activity and safety in Phase II trials for hepatocellular carcinoma, both as monotherapy and in combination with sorafenib, and in Phase 1a/1b in combination with pembrolizumab for patients with advanced solid tumors. It targets the master regulator CCAAT enhancer-binding protein alpha (CEBPA), abnormally silenced by DNA methylation in a wide range of hematological and not hematological malignancies. However, the molecular events enabling this mechanism are only partially elucidated. In this study, we uncovered the molecular basis for AW1-51-induced transcriptional reactivation of CEBPA demonstrating that by directly promoting DNA demethylation of CEBPA promoter restores CEBPA expression, protein synthesis, and consequently cell differentiation. These findings unveil AW1-51 as a prototype for RNA-based precision medicine enabling conditional expression of CEBPA in diseases characterized by aberrant gene silencing and extending its potential therapeutic impact beyond cancer.