Project Summary Genetic and gene expression studies have revolutionized our understanding of the plasma cell malignancy multiple myeloma. As a result, myeloma molecular subtypes have been defined, and genetic markers of high- risk disease now prioritize patients for risk-adapted therapies. Yet current molecular markers fail to identify over 40% of high-risk patients who relapse or die within two years of diagnosis. High-risk myeloma is exemplified by a proliferation gene expression subtype that contains genetic alterations normally found in standard-risk subtypes. Therefore, this high-risk myeloma is not explained by underlying genetic events suggesting an epigenetic component. Epigenetic modifications such as DNA methylation (DNAm) regulate gene expression and provide mechanistic insight into tumor suppressor and oncogene activity. Importantly, DNAm is maintained through cell division and can propagate cancerous phenotypes. Our studies in mice have shown a dramatic remodeling of DNAm during plasma cell differentiation, and that disruption of DNA methyltransferase activity promotes plasma cell proliferation. Yet there is limited data on the role of DNAm in high-risk myeloma and the proliferation subtype. We hypothesize that dysregulation of DNAm contributes to uncontrolled proliferation in high-risk myeloma. This will be investigated through computational analyses of somatic genetic alterations from whole genome sequencing, gene expression from RNA-seq, and DNAm data derived from whole genome bisulfite sequencing on 392 specimens from the CoMMpass trial (NCT01454297). In aim 1, we will determine the DNAm program of myeloma subtypes defined by both somatic alterations and gene expression. Here, a focus will be to elucidate the epigenetic program of high-risk proliferative disease. In aim 2, we will identify the DNAm program prognostic of poor outcome and contrast that with changes observed in relapsed specimens. DNAm loci associated with the proliferative subtype, prognostic of outcome, and reprogrammed in relapse myeloma will be prioritized for interrogation using CRISPR-based site-specific epigenetic editing. Results from these aims will help resolve the functional consequences of dysregulated DNAm in aggressive myeloma and provide a molecular map of multiple myeloma in the context of patient outcomes. Insights from this work will prioritize avenues for follow-up translational and mechanistic studies to target high-risk multiple myeloma.