Genomic systems approach to measure HSV-1-specific T-cell dominance in humans

描述(申请人提供)：单纯疱疹病毒1型(HSV-1)是一种具有重要医学意义的病原体。60%的美国公民被感染，发展中国家的感染率更高。严重的并发症，如成人或新生儿脑炎造成的永久性脑损伤，以及导致失明的角膜炎症，即使在角膜移植后也会出现长期效果不佳的情况，人均发生率较低。由于感染如此普遍，这些并发症的累积医学影响是相当可观的。目前还没有获得许可的HSV-1疫苗。HSV-2是一种氨基酸同一性为50%的病原体，在最近的III期试验中，一种疫苗没有临床活性。这种HSV-2疫苗针对抗体和CD4T细胞反应，而不是CD8T细胞反应。这项研究的长期目标是合理设计HSV-1预防性和治疗性疫苗。CD8T细胞是HSV-1免疫应答的重要组成部分，定位于感染的皮肤、角膜、视网膜和三叉神经节。从人类HSV-2和小鼠HSV-1数据可以看出，CD8细胞持续存在于愈合的皮肤感染部位，为重新激活提供早期监测。对它们在人类身上的细微特异性几乎一无所知。我们开发了新技术，使我们能够以前所未有的详细程度研究CD8和CD4T细胞对HSV-1的反应，包括研究从未患过疱疹皮损的人的外周血，即免疫成功的人。目的1采用一系列新的方法：高效交叉呈现、基于CD137的排序和克隆的HSV-1 ORF文库，以高通量和清晰度解码CD8应答HSV。我们将比较有症状和无症状HSV-1感染者的免疫反应，并确定既有高人群流行率又与无症状感染相关的抗原。HSV-1特异性CD4定位于人类感染的眼睛、甘油三酯和皮肤，以及小鼠感染的组织，可能具有重要的直接抗病毒和辅助功能。目的2使用完整的ORFeome高通量定义HSV-1特异性CD4应答的广度、特异性和受试者内真实的免疫优势等级。该工作流程使用基于CD137的分选和高度抗原性的HSV-1或Feome蛋白质集。显性分析是两个维度的组合(应答细胞寡克隆和基质ORF池)，并提供每个受试者约100个HSV-1特异性CD4T细胞的良好特异性，以定义CD4显性结构。该项目的总体目标是确定HSV-1 ORF疫苗，这种疫苗能刺激与轻度感染相关的不同人类白细胞抗原类型的人的主要和流行的CD8和CD4表位。

DESCRIPTION (provided by applicant): Herpes simplex virus type 1 (HSV-1) is a medically significant pathogen. 60% of US citizens are infected and rates are higher in developing countries. Serious complications, such as permanent brain damage from encephalitis in adults or neonates, and corneal inflammation leading to blindness and poor long- term outcomes even after corneal transplantation, occur at a low per-person rate. Because infection is so prevalent, the cumulative medical impact of these complications is quite considerable. There is no licensed vaccine for HSV-1. A vaccine for HSV-2, a pathogen with overall 50% amino acid identity, had no clinical activity in a recent phase III trial. This HSV-2 vaccine targeted antibody and CD4 T-cell responses, but not CD8 T-cell responses. The long-term goal of this research is the rational design of preventative and therapeutic vaccines for HSV-1. CD8 T-cells are a critical component of the immune response to HSV-1 and localize to infected skin, cornea, retina, and trigeminal ganglia. By implication from human HSV-2 and murine HSV-1 data, CD8 cells persist at sites of healed skin infection to provide early surveillance for reactivation. Almost nothing is known about their fine specificity in humans. We have developed novel technology what allows us to study the CD8 and CD4 T-cell response to HSV-1 in unprecedented detail, including the study of peripheral blood from person who never have herpetic lesions, that is, person with immunologic success. Aim 1 uses a sequence of novel methods: efficient cross-presentation, CD137-based sorting, and a library of cloned HSV-1 ORFs, to decode the CD8 response to HSV with high throughput and definition. We will compare immune responses in persons with symptomatic and asymptomatic HSV-1 infection and identify antigens that have both high population prevalence and correlate with asymptomatic infection. HSV-1-specific CD4s localize to infected eyes, TG, and skin in humans, and to infected tissues in mice, and likely provide important direct antiviral and helper functions. Aim 2 defines with high throughput, using the complete ORFeome, the breadth, specificity, and within-subject true immunodominance hierarchy of HSV-1-specific CD4 responses. The workflow uses CD137-based sorting to and a highly antigenic HSV-1 ORFeome protein set. Dominance assays are combinatorial in two dimensions (responder cell oligoclonality and matrix ORF pooling) and deliver the fine specificity of ~100 individual HSV-1-specific CD4 T-cells per subject to define the CD4 dominance architecture. The overall goal of the Project is to identify HSV-1 ORFs for vaccines that stimulate dominant and prevalent CD8 and CD4 epitopes to persons of diverse HLA type that are correlated with mild infection.