Cancer Immunotherapy with a Live Recombinant Vaccine

描述（由申请人提供）：在上一个资助期间，我们 开发了两种基于单核细胞增生李斯特菌的疫苗，Lm-E7和LmLLO-E7， 诱导对人乳头瘤病毒（HPV）致癌蛋白E7的免疫， 只有Lm-LL0-E7导致在人宫颈癌中建立的HPV永生化肿瘤消退， 鼠标我们已经发现，在抗E7免疫诱导的差异， 这些疫苗可以解释它们在杀死E7方面的不同功效 表达肿瘤。我们请求资助在HPV-E7中探索这些发现。 由小鼠肿瘤组成的系统，通过HPV自然永生化，移植 导入同基因野生型小鼠或E7转基因同基因小鼠 基因我们假设基于E7的免疫原在诱导免疫应答中的功效是不明显的。 已建立的癌症的消退与CD8+和CD4 + T细胞的类型有关 免疫诱导我们已经证明，两种疫苗诱导的CTL水平相似， 对表达E7的肿瘤细胞和大量CD8 + T细胞的活性 用肽/MHC I类四聚体染色。然而， 如通过细胞内细胞因子染色所确定的，CD8 + T细胞相当 与众不同我们还表明，尽管由Lm-LL0-E7诱导的CD4 + T细胞是 需要的抗肿瘤功效，这一子集是适得其反， Lm-E7治愈表达E7的肿瘤的小鼠的能力。在我们的第一个具体的 目的我们将表征由Lm-E7和Lm-LLO-E7诱导的T细胞应答， 脾脏和肿瘤所在的区域我们还假设E7是 免疫原性更强，如果将其递送， 作为与胰蛋白酶的融合蛋白（LLO）。我们已经表明， 牛痘病毒和L.表达LLO-E7的单核细胞增多症 比单独携带E7时更有效的肿瘤免疫治疗剂。在我们的第二 具体目的，我们希望确定是否LLO-E7蛋白通过 常规佐剂或脉冲树突状细胞可以产生更好的抗肿瘤作用， 免疫疗法优于单独的E7。如果肿瘤的免疫原性 抗原可以通过融合到这种细菌蛋白质中来增强， 开发比使用活疫苗更安全的免疫策略， 重组载体来递送抗原。最后，一个重要的要求 在小鼠中建立任何癌症治疗剂的功效是为了表明， 它可以诱导针对内源性抗原的免疫力， 宿主的免疫耐受性在我们的第三个具体目标中，我们将测试最 在E7转基因小鼠模型中有希望的疫苗递送系统。的 这些研究的结果可能普遍适用于其他癌症。 方法和测量免疫成分， 良好的肿瘤免疫治疗。

DESCRIPTION (provided by applicant): 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.