Merkel cell carcinoma (MCC) is an often-lethal skin cancer with a reported incidence that has quadrupled in the past 20 years to ~1,600 cases/year in the US. In 2008, the Merkel cell polyomavirus (MCPyV) was discovered to be integrated in ~80% of MCC tumors. MCCs require persistent expression of the viral T antigen (T-Ag) oncoprotein for growth, providing a target for rational immune therapy. Although most MCC patients have no apparent immune defect, persons with T cell dysfunction due to HIV, chronic leukemia or immunosuppressive drugs have a 10- to 30-fold increased MCC risk. This proposal builds on our findings that humoral immune responses to T-Ag are linked to MCC progression while cellular immune responses appear to be protective. Similar to the other known human polyomaviruses, MCPyV infection is prevalent in the general population (53% of adults have anti-MCPyV capsid protein antibodies). In marked contrast, we have found that antibodies to MCPyV T-Ag oncoproteins are rare in population controls (1%) but are prevalent among newly diagnosed MCC patients (~50%). Moreover, anti-T-Ag antibody titers are dynamic in MCC patients and increase in parallel to tumor burden, allowing detection of disease progression in some cases before identification by the patient or physician. In Aim 1, we propose to further characterize anti-T-Ag antibodies and to test their clinical utility as a specific biomarker for MCC recurrence. We have also found that cell-mediated immunity (intratumoral CD8+ T cell infiltration) is associated with excellent MCC prognosis. Therefore, in collaboration with Dr. David Koelle, in Aim 2 we will comprehensively map T cell epitopes/MHC restriction for MCPyV oncoproteins, develop peptide-MHC reagents, and use them to characterize the frequency and function of MCPyV-specific T cells in the blood and tumors of MCC patients. In Aim 3, we will use human tumor samples and a mouse model of MCC to determine the potential efficacy of three immune stimulating therapies for possible future human MCC trials. These studies will be performed in close collaboration with Drs. Thomas Blankenstein and Gerald Willimsky using their existing, well-characterized mouse model of spontaneous polyomavirus-induced cancer that shares striking biologic, clinical, and immunologic similarities with MCC. This proposal combines the expertise of investigators who each have over ten years of documented productivity in three synergistic areas: clinical and translational research in MCC (Nghiem), comprehensive characterization of the cellular immune response to human skin-associated viruses (Koelle) and characterization of the humoral and cellular immune responses to polyomavirus-driven sporadic cancer in mouse models (Blankenstein/Willimsky). Because antigen-specific cellular and humoral responses to viral proteins expressed in MCC can be tracked, these studies will provide significant biological insight into the immune response to cancer more generally, with directly applicable prognostic and therapeutic implications for MCC patients.