Project Summary/Abstract Myocarditis, pathologic inflammation of the heart, is a serious cause of sudden cardiac death affecting patients of all age groups. Immune checkpoint inhibitors (ICIs) are monoclonal antibodies to cytotoxic T-cell antigen-4 (CTLA-4) or programmed death-1 (PD-1)/programmed death-1 ligand (PD-1L) used as novel cancer therapeutics to release intrinsic brakes on T-cell cytotoxicity against tumor cells. Although ICIs are now relied upon to treat many advanced cancers, fulminant myocarditis has been reported as a life-threatening side effect of these drugs, leading to severe arrhythmias, heart failure and death. Under histopathology, an acute lymphocytic infiltrate is found in the heart, and multiple lines of evidence point to a T-cell and antigen-mediated phenomenon. Although T-cell clonal analysis of patient heart tissues suggest the existence of a cardiac-specific antigen in ICI-induced myocarditis, the identity of such antigen(s) remains elusive. Understanding the culprit antigens in this disease may lead to novel insights on the mechanism of T-cell mediated myocardial damage. I hypothesize that ICI-induced myocarditis is an autoimmune disorder caused by cardiac-specific auto- antigens that trigger the activation and clonal expansion of T-cells, leading to myocardial inflammation. The goal of this study is to use an advanced immunophenotyping technique called CyTOF to characterize T-cell subsets, and then use single-cell T-cell receptor sequencing and a novel computational algorithm called GLIPH (Grouping Lymphocyte Interactions by Paratope Hotspots) to identify specific epitopes responsible for T-cell activation in ICI- induced myocardial damage in mice and humans. By completing this project, I will uncover key insights into the biological mechanism of ICI-induced myocarditis which will bridge a major knowledge gap regarding the role of T-cells in mediating myocardial damage. My results will answer the key question of which immune cell types play a dominant role in ICI-induced myocarditis, as well as identify disease-causing antigens causing cardiotoxicity. There is increasing evidence to suggest that immune-mediated cardiac damage may be at the heart of many major cardiovascular diseases, ranging from autoimmune myocarditis to heart failure to atherosclerosis. My long-term goal as a physician-scientist is to elucidate mechanisms of pathological inflammation in the heart to help cure cardiovascular disease and become a leader in the field of cardiac inflammation.