The Epigenetic Enzyme CARM1 as a Negative Regulator of Dendritic cells and CD8 T cells in Melanoma

Abstract Protective immunity against melanoma requires two immune cell populations that form the melanoma – immunity cycle: Dendritic cells (DCs) act as sentinels that transport melanoma antigens to draining lymph nodes where they prime T cells, and CD8 T cells act as effector cells that identify and kill melanoma cells. The major hypothesis of this application is that therapeutic targeting of CARM1 will enhance the function of both cDC1 and CD8 T cells in the melanoma immunity cycle. CARM1 is an epigenetic enzyme that acts as a co- transcriptional activator by di-methylation of specific arginine residues in histone H3 and other chromatin- associated proteins. We identified CARM1 as a negative regulator of melanoma-infiltrating CD8 T cells using an in vivo CRISPR/Cas9 screen. Inactivation of Carm1 in CD8 T cells enhances their proliferation and effector function in the tumor microenvironment. We also discovered that inactivation of Carm1 in cDC1 enhances their ability to cross-present melanoma antigens to CD8 T cells. In Aim 1, we will investigate the molecular mechanisms by which therapeutic targeting of CARM1 inhibits T cell exhaustion. We will evaluate the available high-affinity CARM1 inhibitor in multiple melanoma models, with an emphasis on immunotherapy resistant models and metastatic disease. We will also use conditional knockout mice to selectively inactivate the Carm1 gene in CD8 T cells to test our hypothesis that Carm1 inactivation inhibits T cell exhaustion. We will also investigate the relevance of CARM1 in human melanoma using co-cultures of tumor-infiltrating T cells and autologous melanoma cells. In Aim 2, we will investigate the molecular mechanisms by which CARM1 inhibition enhances cross-presentation of melanoma antigens by cDC1 to CD8 T cells. Preliminary data show that conditional knockout of Carm1 in cDC1 enhances melanoma immunity and sensitizes resistant melanomas to PD-1 blockade. We will use these conditional knockout mice to study antigen cross-presentation in vitro and in vivo. Finally, we will identify the direct target genes regulated by CARM1 by CUT&TAG, ATAC-seq and RNA- seq. These studies will develop a novel strategy to enhance the anti-tumor activity of both DCs and T cells in the melanoma – immunity cycle.