Development of Novel Chagas Vaccines

描述（申请人提供）：克氏锥虫是一种重要的人类病原体，每年造成1 -5万人死亡，感染1200 - 2000万人。病媒控制措施取得了一些成功，但现有的药物疗法疗效低、毒性高，而且没有特定的疫苗进入人体试验。目前迫切需要一种安全有效的疫苗，以保护面临感染这种致命疾病风险的4000万至1亿人。在慢性感染期间，检测到对寄生虫裂解物和多种克氏锥虫蛋白的强大CD8+ T细胞反应。事实上，克氏锥虫特异性IFN-？产生CD8+ T细胞与减缓疾病进展相关。我们已经建立了动物模型来仔细分析与克氏锥虫感染相关的疫苗诱导的免疫反应和心脏病理。一些研究表明，CD4+ T细胞在保护性免疫的启动功能中是必需的，CD8+ T细胞在效应功能和寄生虫清除中是必不可少的。克氏t型病毒反式唾液酸酶（TS）抗原可诱导小鼠对正常致死性全身和亚致死性粘膜克氏t型病毒的有效保护；然而，TS疫苗尚未经过优化以供人类使用。此外，TS基因家族占所有克氏锥虫基因的5%，某些TS抗原的表达可能参与了寄生虫的免疫逃避。我们将通过比较编码功能性TS基因（编码12-15种酶活性蛋白）、非功能性TS基因（bbb700基因）和哺乳动物阶段非TS基因表位的疫苗来解决后一种可能性。我们建议利用最先进的免疫信息学技术，从这些预测能结合常见HLA等位基因的重要克氏t形虫蛋白中鉴定出关键的寄生虫表位（为所有人群的95%提供相关的选择覆盖率）。预测的表位序列将被合成，HLA结合试验将被用来验证MHC结合。我们将与加州大学洛杉矶分校的Meymandi博士合作，从感染克氏锥虫的个体（和未感染的对照）中获取血液样本，并用上述IFN-?？ELISPOT和流式细胞术检测。基于这些筛选试验的结果，我们将设计编码这些T细胞表位的新型疫苗，并在“人源化”HLA A2/DR1双转基因小鼠中测试其预防和治疗效果。我们的研究结果将提供进行第一阶段南美锥虫病疫苗试验所需的关键数据。

DESCRIPTION (provided by applicant): Trypanosoma cruzi is an important human pathogen responsible for 10-50,000 deaths/year infecting 12-20 million people. Vector control measures have met with some success but available drug therapies have low efficacy and high toxicity, and no specific vaccines have entered human trials. There is a desperate need for a safe and effective vaccine to protect the 40-100 million individuals at risk of acquiring this deadly diseas. Robust CD8+ T cell responses are detected against parasite lysate and a variety of T. cruzi proteins during chronic infection. In fact, increased frequencies of T. cruzi-specific IFN-? producing CD8+ T cells correlate with decreased disease progression. We have developed animal models to carefully analyze vaccine-induced immune responses and cardiac pathology associated with T. cruzi infection. Several studies have shown that CD4+ T cells are necessary during priming functions of protective immunity, and CD8+ T cells are essential for effector function and parasite clearance. T. cruzi trans-sialidase (TS) antigens can induce potent murine protection against normally lethal systemic and sub-lethal mucosal T. cruzi challenges; however, TS vaccines have not been optimized for human use. In addition, the large TS gene family represents >5% of all T. cruzi genes, and expression of some TS antigens may be involved in parasite immunoevasion. We will address this latter possibility by comparing vaccines encoding epitopes from functional TS genes (encoding the 12-15 enzymatically active proteins), non-functional TS genes (>700 genes), and mammalian stage non-TS genes. We propose to identify key parasite epitopes from these important T. cruzi protein sets predicted to bind common HLA alleles (providing selection coverage relevant for >95% of all human populations) utilizing state- of-the-art immunoinformatics. Predicted epitope sequences will be synthesized and HLA binding assays will be employed to validate MHC binding. In collaboration with Dr. Meymandi at UCLA, we will obtain blood samples from individuals infected with T. cruzi (and uninfected controls) and stimulate purified PBMC with peptides described above in IFN-?? ELISPOT and flow cytometric assays. Based on results from these screening assays, we will design novel vaccines encoding these T cell epitopes, and test their prophylactic and therapeutic efficacies in 'humanized' HLA A2/DR1 dual transgenic mice. The outcome of our studies will provide key data required for proceeding with phase I Chagas vaccine trials. PUBLIC HEALTH RELEVANCE: Approximately 12-20 million people are infected with T. cruzi, 10-50,000 people die each year from Chagas disease, people in the US are at risk of T. cruzi infection through blood transfusions and organ donations originating from infected persons, and there are no highly effective vaccines or chemotherapies for the prevention or treatment of T. cruzi infection and disease. The work proposed in this application focuses on the identification of T. cruzi T cell epitopes and will generate and test novel vaccines in 'humanized' HLA transgenic mice. Results of our studies will provide important new information relevant for the development of more effective T. cruzi vaccines, as well as the development of vaccines for other major human diseases (e.g.-AIDS, TB) caused by mucosally invasive, intracellular infections.