KDM6A mutation as an epigenetic driver of multiple myeloma

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PROJECT SUMMARY Background: Next generation sequencing has identified recurrently mutated genes in MM. Together anomalies of epigenetic regulator genes are present in 25% MM patients [4]. Among these affected genes are those encoding the histone demethylase KDM6A, the histone methyltransferase KMT2C/D, the histone acetyltransferase CREBBP and the chromatin remodeling subunit ARID1A, which are known to interact together to activate enhancers. KMT2C mutations are more frequent in relapsed patients and this correlates with shorter therapy response duration. KDM6A gene copy loss occurs in about 50% of male and female patients, and mutations of this gene are associated with poor survival. These data emphasize the importance of enhancer deregulation in the biology of MM but yet no functional studies have explored this in detail. We have shown Loss of KDM6A stimulates growth, clonogenicity and adhesive properties of MM. Re-expression of KDM6A or a demethylase inactive form both suppresses cell growth. Hypothesis: KDM6A deletion causes loss of enhancer activity and gene expression favoring uncontrolled proliferation. The recruitment of activators and not demethylating activity of KDM6A is most critical for this effect. Specific Aims: 1a. Determine KDM6A's direct targets. KDM6A will be mapped by ChIP-Seq in MM cell lines CRISPR engineered to express V5-tagged and degradation-inducible KDM6A. 1b. Define the role of KDM6A demethylase activity in enhancer function. KMT2C/D, P300/CBP binding and H3K27me, H3K27Ac modifications will be mapped by ChIP-seq in engineered cells expressing endogenous KDM6A with no demethylase activity. 2. Explore biological effect of KDM6A loss in a mouse model of MM. MM will be generated in KDM6Aflox/floxCD19-Cre mice by transduction of bone marrow with activated IL6 receptor and the effects on tumor biology determined. The aims proposed in this LLS Special Fellowship are related to a grant R01CA180475 aiming to understand the importance of KMT2C and UTX/KDM6A in the control of chromatin structure and homeostasis of MM by identifying their genetic target and how they affect B-cell maturation. This fellowship is expanding the understanding of KDM6A as a tumor suppressor in multiple myeloma, and is facilitating the development of tool necessary to study the role of KMT2C in MM as well.