Identification of targets of CD4 and CD8 T cell reactivity conserved across Sarbecoviruses and recognized across different animal species

项目总结 项目2的首要目标是发现和设计具有广泛交叉能力的候选疫苗抗原。 对具有大流行潜力的多种冠状病毒(CoV)物种的保护性T细胞免疫。的交叉反应性 在SARS-CoV-2、其他肉瘤病毒和其他病毒之间已经明确地证明了T细胞水平 在我们以前的研究中有远缘关系的冠状病毒。通过采用明确地诱导细胞免疫的策略 转化为项目3产生的疫苗，针对未来从冠状病毒宿主中出现的大流行的交叉保护 也许是可能的。特定目标1将定义广泛存在的CD4+和CD8+T细胞反应性的靶点 在不同的肉芽病毒、汞病毒和其他关注的贝塔冠状病毒中保守 通过(I)确定整个冠状病毒蛋白质组的保守区，包括所有成熟的非结构蛋白 (2)对每一个分类群进行元分析 免疫原性数据，对这些抗原的已知T细胞表位进行分类，以确定哪些抗原亚区 在不同冠状病毒物种自然感染后被识别，以及(Iii)整合这些数据以选择一组 候选保留了三个分类组中每一个组的T细胞表位区域，用于实验评估。 这种生物信息学分析将分配特定的定量分数，包括序列保守分数， 人类免疫原性评分，以及人类T细胞对不同蛋白质亚区的交叉反应评分。 特定目标2将针对来自目标1的表位测试人类T细胞的识别能力 不同肉毒、分枝病毒和其他贝塔冠状病毒的同源序列 通过(I)从原型病毒合成目标1中定义的保守表位区域和 来自不同分类组的同源多肽在目标1中进行评估，(Ii)诱导T细胞系 来自新冠肺炎恢复期人类供者和人类受试者的NL63和OC43反应性记忆T细胞 使用SARS-CoV-2、NL63-和OC43衍生的表位，(Iii)检测这些表位特异性T细胞系的交叉 使用一种方法与来自三个分类组中的每一个的相应同源多肽的反应性 我们实验室开发的用于评估冠状病毒交叉反应的方法，以及(Iv)执行生物信息学分析以预测 我们的啮齿动物模型的免疫原性。在这项试验性工作的基础上，我们将优先考虑潜在的候选人， 并将候选名单提交给核心A的总体计划负责人，以纳入两个疫苗平台 在项目3中，并在核心B中测试免疫原性和交叉保护。该项目将产生第一个 全面评价冠状病毒全蛋白质组的T细胞免疫原性 使用生物信息学预测和紧密集成的具有大流行潜力的CoV部分 为疫苗设计创造抗原以引发广泛的交叉保护细胞的实验验证 免疫是对通过项目1引起的体液免疫的补充。

PROJECT SUMMARY The overarching goal of Project 2 is to discover and design vaccine antigen candidates that confer broad cross- protective T cell immunity to multiple coronavirus (CoV) species with pandemic potential. Cross-reactivity at the T cell level has been unequivocally demonstrated between SARS-CoV-2, other Sarbecoviruses, and other more distantly related CoV in our previous studies. By incorporating strategies that explicitly elicit cellular immunity into vaccines generated by Project 3, cross-protection against future pandemics emerging from CoV reservoirs may be possible. Specific Aim 1 will define targets of CD4+ and CD8+ T cell reactivity that are broadly conserved across different Sarbecoviruses, Merbecoviruses, and other betacoronaviruses of concern by (i) identifying conserved regions across the entire CoV proteome, including all mature non-structural proteins (nsp) and open reading frames (ORFs) for each of these taxonomic groups, (ii) performing a meta-analysis of immunogenicity data to catalog known T cell epitopes for these antigens to define which antigen subregions are recognized following natural infection by different CoV species, and (iii) integrating these data to select a set of candidate conserved T cell epitope regions from each of the three taxonomic groups for experimental evaluation. This bioinformatic analysis will assign specific quantitative scores, including sequence conservation scores, human immunogenicity scores, and cross-reactivity for human T cells scores to various proteins subregions. Specific Aim 2 will test human T cells directed against epitopes from Aim 1 for their capacity to recognize homologous sequences from different Sarbecoviruses, Merbecoviruses, and other betacoronaviruses of concern by (i) synthesizing the conserved epitope regions defined in Aim 1 from prototype viruses and homologous peptides from each of the different taxonomic groups evaluated in Aim 1, (ii) eliciting T cell lines from COVID-19 convalescent human donors and human subjects with NL63 and OC43-reactive memory T cells using SARS-CoV-2-, NL63-, and OC43-derived epitopes, (iii) testing these epitope-specific T cell lines for cross- reactivity with corresponding homologous peptides from each of the three taxonomic groups using an approach developed in our laboratory to evaluate CoV cross-reactivity, and (iv) performing bioinformatic analyses to predict immunogenicity in our rodent models. Based on this experimental work, we will prioritize potential candidates, and deliver lists of candidates to the overall Program lead in Core A for incorporation into two vaccine platforms in Project 3 and testing for immunogenicity and cross-protection in Core B. This project will produce the first comprehensive evaluation of T cell immunogenicity of the entire CoV proteome across a broad cross- section of CoVs with pandemic potential using a tight integration of bioinformatics prediction and experimental validation to create antigens for vaccine design that elicit broad cross-protective cellular immunity to complement the humoral immunity elicited through Project 1.