During the last funding period, we developed two Listeria monocytogenes based vaccines, Lm-E7 and LmLLO-E7, that induce immunity to the Human Papilloma Virus (HPV) oncogenic protein E7 but only Lm-LLO-E7 causes the regression of HPV immortalized tumors established in the mouse. We have discovered differences in the anti-E7 immunity induced by these vaccines that may explain their different efficacies in killing E7 expressing tumors. We request funding to explore these findings in an HPV-E7 system consisting of mouse tumors, naturally immortalized by HPV, transplanted into either syngeneic wild type mice or syngeneic mice transgenic for the E7 gene. We hypothesize that the efficacy of E7 based immunogens in inducing the regression of established cancer is related to the type of CD8+ and CD4+ T cell immunity induced. We have shown that both vaccines induce similar levels of CTL activity against E7 expressing tumor cells and high numbers of CD8+ T cells that stain with peptide/MHC class I tetramers. However, the cytokine profile of the CD8+ T cells, as determined by intra-cellular cytokine staining, is quite distinct. We have also shown that whereas CD4+ T cells induced by Lm-LLO-E7 are required for anti-tumor efficacy, this subset is counter productive to the ability of Lm-E7 to cure mice of E7 expressing tumors. In our first specific aim we will characterize the T cell responses induced by Lm-E7 and Lm-LLO-E7 in the spleen and those that home to the tumor. We also hypothesize that E7 is more immunogenic and will induce better anti-tumor immunity if it is delivered as a fusion protein with a listerial protein (LLO). We have already shown that both vaccinia virus and L. monocytogenes that expresses LLO-E7 are much more potent tumor immunotherapeutics than when they carry E7 alone. In our second specific aim we wish to determine whether LLO-E7 protein delivered by conventional adjuvants or pulsed dendritic cells can produce better anti-tumor immunotherapy than E7 alone. If the immunogenicity of a tumor associated antigen can be enhanced by fusion to this bacterial protein then we may be able to develop immunotherapeutic strategies that are safer than those that use live recombinant vectors to deliver the antigen. Finally, an important requirement for establishing the efficacy of any cancer therapeutic in mice is to show that it can induce immunity against endogenous antigens which may have induced immune tolerance in the host. In our third specific aim, we will test the most promising vaccine delivery systems in a transgenic mouse model for E7. The findings of these studies may be generally applicable to other cancer approaches and to the measurement of immune components that are indicative of good tumor immunotherapy.
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