Abstract: Exhaustion mechanisms in Merkel cell polyomavirus-specific T cells Merkel cell carcinoma (MCC) is a rare and deadly cancer that is rapidly rising in incidence. Approximately 80% of MCC cases are caused by the Merkel cell polyomavirus (MCPyV) with the remaining 20% of cases caused by UV mutations. Immunotherapies targeting the PD-1 pathway have been highly successful in patients with MCC. Anti-PD-1 therapy works by reinvigorating T cells which have entered a dysfunctional state known as exhaustion. However, these therapies are still insufficient for long-term disease control in half of patients with advanced MCC and must be further studied. In most other cancers T cells recognize unique tumor mutations that vary from patient to patient. Because T cell targets are unique for each patient, studies of cancer-specific T cells are largely infeasible in mutationally driven cancers. However, in the case of virus- positive MCC, T cells recognize viral proteins driving the cancer which are shared across patients. Over the past decade our lab has developed tools to study T cells that recognize these viral oncoproteins. These tools put us in a unique position to study cancer-specific T cells from our extensive repository of more than 1,500 MCC patients. These reagents will be key to understanding why some patients do or do not respond to anti- PD-1 therapy in MCC as well as in other cancers where studies of tumor-specific cells would be infeasible. Aim 1 of this proposal will use newly developed techniques to extensively characterize circulating cancer- specific T cells in MCC patients over the course of anti-PD-1 therapy. Specifically, high dimensional flow cytometry, single cell RNA sequencing and T cell receptor sequencing will be used to characterize clonal, MCPyV-specific T cells over the course of anti-PD-1 therapy. T cells from patients that respond or do not respond to therapy will be compared to identify deficiencies in the T cell response which could be augmented in future clinical trials. Aim 2 will study a putative state of dysfunction that we identified in MCC-specific CD4 helper T cells. CD4 T cells are highly understudied but are known to be important for control of chronic infections and cancers. This aim will investigate the cytokine secretion profile and cytotoxic capacities of these cells to determine if they exert an immune suppressive phenotype. We hypothesize this dysfunction could be helping cancers evade the immune response and believe these studies could inform future clinical trials. This project is ideal for training young physician scientists due the highly translational nature of the research, the extensive background of Dr. Paul Nghiem in training previous MD-PhD students, and the quality research and medical education at the University of Washington. The activities detailed in this proposal will provide a strong background for a future career as a physician scientist.