Metastasis is the major cause of cancer mortality. Effective therapies are urgently needed for patients with metastatic diseases. Increasing evidence suggests that interactions between cancer cells and tumor immune microenvironment drive metastatic progression. Although prior studies established the vital roles of immunosuppressive TME in metastatic progression, the epigenetic determinant in shaping the metastatic niche remains understudied. Absent, small or homeotic 1-like (ASH1L) is a histone lysine methyltransferase that induces methylations at H3K36 and . Although ASH1L was found to drive leukemogenesis, little is known about its biological function in solid tumors and metastatic disease. Our preliminary studies showed that ASH1L is genetically amplified and overexpressed in metastatic tumors and contributes to metastasis and immunosuppression. The goal of this application is to determine the role and mechanisms of action of ASH1L in the metastatic niche. The central hypothesis in this application is that histone H3K4 and activates gene transcription methyltransferase ASH1L contributes to the immunosuppressive metastatic niche and is a potential therapeutic target in metastatic cancers. In the proposed studies, we will 1) determine ASH1L’s role in reshaping the metastatic niche by combining a newly developed genetically engineered mouse model and cutting-edge single- cell transcriptomic technologies; 2) elucidate the mechanism by which ASH1L induces immunosuppressive metastatic niche by performing epigenetic profiling, proteomic approaches, and functional studies; 3) determine the anti-metastatic effects of genetically depleting or pharmacologically inhibiting ASH1L in combination with checkpoint immunotherapy in preclinical models, followed by profiling their impact on immune components in the metastatic niche. We expect to identify ASH1L as a key epigenetic determinant in priming immunosuppressive metastatic niche and develop effective targeted therapy and combinatorial immunotherapy for metastatic cancers. These studies are expected to have significant positive impacts, including bridging the knowledge gap on the role and mechanisms of action of an understudied epigenetic factor ASH1L in metastatic cancers, advancing our understanding of the crosstalk between invading cancer cells and immune components in the metastatic niche, and offering implications in regard to the biomarkers, therapeutic targets, and rational combinations for metastatic malignancies. 1