PROJECT SUMMARY Immunotherapy, particularly immune checkpoint inhibitors (ICI), has shown considerable success in the treatment of advanced renal cell carcinoma (RCC), leading to durable responses in a subset of patients. However, most patients do not derive long-term clinical benefit from existing immune therapies. Moving forward, our team has prioritized identifying the determinants of ICI response in RCC, including the antigenic targets, T cell phenotypes, and microenvironment features that ultimately dictate the effectiveness of tumor- specific immunity. Human endogenous retroviruses (ERVs) are often aberrantly expressed in numerous disease states, including malignancy, and can lead to activation of both innate immunity (through sensing by the RIG-I/MDA-5 and cGAS-STING pathways) and adaptive immunity (by providing antigenic targets for tumor-specific CD8+ T cells). The expression of ERVE-4 has been associated with response to ICI monotherapy (i.e. anti-PD-1) in an analysis of a phase III randomized controlled trial in RCC, and an ERV-derived peptide (from ERVE-4) was identified as a target epitope in a long-term RCC responder to allogeneic stem cell transplant. Beyond recognition of tumor antigens, effective anti-tumor immunity requires a tumor microenvironment that permits infiltrating CD8+ T cells to carry out their effector function. A high level of tumor infiltration by antigen- experienced, non-exhausted CD8+ T cells has been associated with improved response to ICI monotherapy in clinical trials of RCC. However, resistance to ICI monotherapy is still common, in part owing to the immunosuppressive effects of infiltrating Treg and myeloid cells, and consequently combination (and not single agent) therapies with PD-1 blockade as a backbone are now the standard-of-care treatment for advanced RCC. It is therefore critical to understand the role of ERV expression, T cell phenotype, and the immune microenvironment in the context of clinically relevant contemporary ICI-based combination therapies. We hypothesize that aberrant expression of ERVs is associated with improved response to ICI combination therapy with ipilimumab and nivolumab, and that characterization of T cell phenotypes and the composition and states of other infiltrating immune cells in the RCC microenvironment will improve our understanding of the determinants of ICI response. By leveraging our unique access to clinically relevant large- scale clinical trial specimens of single-agent ICI and ICI-based combination therapy in RCC, as well as our collaborative team of physicians, translational researchers, and experimental biologists, we aim to systemically assess the association of ERV expression and ICI response and to determine other immune microenvironment features that ultimately influence response to current RCC immunotherapies. Our multidisciplinary team brings together a collective expertise in RCC biology, pathology, and tumor immunoregulation who are dedicated to brin...