Role of EZH2 as a Driver and Therapeutic Target of Hepatocellular Carcinoma

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PROJECT SUMMARY Hepatocellular carcinoma (HCC) accounts for over 30,000 deaths annually in the United States and current therapies provide negligible clinical benefit. Factors that epigenetically silence HCC tumor suppressor genes are expected to promote tumor development and thus may provide novel therapeutic targets. The histone methyltransferase EZH2 is the catalytic subunit of polycomb repressive complex 2 (PRC2), a master regulator of epigenetic silencing. EZH2 is mutated or overexpressed in a variety of malignancies and has been shown to function as an oncogene in some cancers. Efficacious small molecule inhibitors of EZH2 have been developed and have shown promising results in clinical trials for certain EZH2-driven cancers and recently received FDA approval for the treatment of epithelioid sarcoma and follicular lymphoma. Based upon analysis of HCC patient samples, and experiments involving cultured HCC cells and mouse models of HCC, we have found that EZH2 is an HCC oncogene that is an intrinsic driver of tumor development. This intrinsic oncogenic activity of EZH2 is due, at least in part, to epigenetic silencing of an ERK-specific phosphatase, DUSP6, resulting in activation of a mitogen-activated protein (MAP) kinase pathway that stimulates cellular proliferation. Furthermore, we have demonstrated, in two published studies from our group, that EZH2 suppresses the ability of natural killer (NK) cells to eradicate cultured HCC cells by epigenetically silencing the gene encoding ULBP1, a tumor cell-surface activating ligand for NK cells, and suppresses NK cell migration to HCC cells by epigenetically silencing the gene encoding the chemokine CXCL10. Thus, our published and preliminary results suggest that EZH2 drives HCC tumor development through both a cell intrinsic mechanism (activation of a MAP kinase pathway that stimulates cellular proliferation) and a cell extrinsic mechanism (inhibition of NK cell-mediated tumor eradication). In this application our objectives are to establish these two roles of EZH2 in HCC mouse models, determine the molecular pathways through which EZH2 promotes tumor growth, and evaluate EZH2 as an HCC drug target. In specific aim 1, we will establish that EZH2 promotes HCC tumor growth by suppressing NK cell-mediated anti- tumor immunity in humanized mice, and define the roles of CXCL10 and ULBP1 as critical EZH2 targets whose epigenetic silencing suppresses NK cell-mediated eradication of HCC. In specific aim 2, we will establish DUSP6 as a critical EZH2 target gene whose epigenetic silencing promotes HCC tumor growth through increased ERK1/2 phosphorylation. In specific aim 3, we will evaluate pharmacological inhibition of EZH2 alone and in combination with other anti-cancer agents as a potential HCC therapy. Collectively, the results of the experiments proposed in this application will elucidate the cell intrinsic and cell extrinsic pathways through which EZH2 promotes HCC tumor growth, and evaluate new therapeutic approaches for HCC.