Project Summary Currently no method exists that would allow physicians to rapidly and reliably establish T-cell immunity against tumor antigens. Bioinformatics tools can predict antigens on cancer cells that are recognized by T cells, but the vaccines based on them often fail because the immunized individuals have too few T cells with the appropriate receptors or lack them altogether. The overall goal of our research proposal is to resolve this problem by developing injectable nanoreagents that introduce into the peripheral T-cell repertoire engineered T-cell receptors (TCRs) that optimally bind the most prevalent vaccine epitope. Specifically, we hypothesize that a customized specificity can be programmed into T cell populations by combining anti-cancer vaccines with techniques that genetically enable endogenous CD8 T cells to express TCRs specific for the vaccines. We further hypothesize that we can use this platform to program CD4 T helper cells with defined MHC class-II- restricted TCRs, and thereby improve tumor-specific CD8 lymphocyte and B cell responses to tumor antigens compared to conventional immunization methods. Our multidisciplinary team of immunologists, bioengineers and geneticists has already established that intramuscularly injected nanoparticles can deliver engineered TCR genes into host T cells so they recognize cancer vaccine antigen. Following rapid vaccine-induced expansion, nanoparticle-programmed T cells ultimately differentiate into long-lived memory T cells. Our long-term goal is to develop a full suite of nanoparticles drugs that would allow physicians to rapidly establish anti-cancer immunity by introducing exogenous antigen-specific TCRs into the patient's T-cell pool. As essential steps toward achieving this goal, we propose the following Specific Aims: (1) To test the wider applicability and long- term safety of programming vaccine specificity into CD8+ T cells, (2) to quantify the degree to which host CD4+ T cells programmed with TCRs to cancer-vaccine antigens boost the immune response, and (3) to determine if providing optimized CD4 T-cell help and reversing tumor immune evasion mechanisms enables in situ programmed vaccine-specific T cells to eradicate disease. We believe that data, reagents, and technology systems generated by our research will provide a conceptual framework for the design of a broad repertoire of gene modification systems designed to generate selective immunity against any type of cancer. Using these in the clinic could make cancer vaccines not only more effective, but also reduce the likelihood of vaccine failure.