Project Summary Multiple myeloma is a cancer of plasma cells resulting in over 12,000 U.S. deaths each year. Genetic alterations in myeloma include trisomy of most odd-numbered chromosomes, translocations that result in ectopic expression of oncogenes as well as structural variants and mutations in oncogenes and tumor suppressor genes. These distinct genetic alterations manifest as unique molecular subtypes. Significant improvements in outcomes have been made using therapies including proteasome inhibitors, thalidomide-analogs collectively known as immunomodulatory imide drugs (IMiDs), and immunotherapies. Unfortunately, most patients still develop disease that is refractory to treatment and succumb to myeloma. These frontline therapies are used regardless of myeloma subtype and whether they are equally effective in all myelomas is not fully understood. This is underscored by our recent study identifying Immunoglobulin Light Chain Lambda (IGL) translocations as predictive of poor IMiD responses. IMiDs mediate the degradation of the lymphocyte transcription factors IKAROS and AIOLOS. Interestingly, the IGL enhancer was bound by some of the highest levels of IKAROS in the myeloma epigenome. This suggests that location and levels of IKAROS and AIOLOS activity in the genome dictate IMiD responses. To test the hypothesis that the genomic elements bound by IKAROS and AIOLOS determine IMiD response and that these regions vary between myeloma genetic subtypes the following aims are proposed. 1) IKAROS and AIOLOS genomic binding sites will be determined and compared between IMiD- sensitive and -resistant myeloma cells to see if these factors localize to distinct regions of the genome in IMiD- resistant myeloma. In conjunction, the epigenetic program of IKAROS- and AIOLOS-bound regions will be characterized under baseline and IMiD-treated conditions to identify how IKAROS and AIOLOS depletion affects these elements as well as the resultant impact on gene expression. 2) IKAROS and AIOLOS will each be inhibited using CRISPR interference and the resultant phenotypic and molecular impact will be assessed to determine the contribution of each factor to IMiD responses. 3) IKAROS- and AIOLOS-bound genomic elements will be systematically disrupted to test the function of these regulatory elements in mediating IMiD resistance. The long-term goals of the candidate are to run an independent research program investigating the etiology of cancer and translate findings into better therapeutic targeting and patient outcomes. The above aims will provide a framework for establishing an independent research program. The above aims will also apply innovative new approaches to discover the mechanism by which IMiDs exert their therapeutic effects and why some myelomas become IMiD resistant, a major cause of multiple myeloma mortality.