HSV-2 specificity and phenotyping of tissue-based T cells in genital skin biopsies of HSV-2 reactivation

项目摘要/摘要 提案摘要：该提案支持一项为期5年的辅导式职业发展奖，调查 通过晶状体的组织免疫反应最近的一期免疫治疗疫苗试验 为开发有效的单纯疱疹病毒2型(HSV-2)疫苗和 改善全球健康状况。这项试验包括三种顺序剂量的免疫治疗性疫苗HSV529， 在有症状的HSV-2感染的个人中。T细胞受体(TCR)谱系分析T细胞 HSV-2再活化区小活检组织中的细胞与非受累皮肤组织中T细胞的比较 并以HSV-2反应性CD4T细胞从外周血中提取两条主线进行研究。第一个是 随着时间的推移，生殖器皮肤在HSV-2重新激活部位的TCR谱系、手臂和 HSV-2特异性CD4T细胞随时间的变化；在生殖器组织中检测到的T细胞在血液中是否可检测到 标准抽样技术。第二个问题是这些关系和动态在 免疫治疗性疫苗接种。初步发现，一些T细胞在治疗后数量增加 每个人都接种疫苗，皮肤和血液之间几乎没有重叠。中心假设是 对针对HSV-2的疫苗有反应的T细胞可能是HSV-2特异性的，而且在组织中看到的T细胞 在拷贝数量较高的较长时间内很可能会有纸巾滞留。这项提议的目的是 就是更多地了解这些常驻T细胞，否则很难研究它们。这样做的理由是 到目前为止，HSV-2疫苗的开发还没有成功，这是因为重叠太少 在皮肤和血液之间，直接观察组织可能有助于识别病毒抗原或一种 以皮肤为基础的T细胞为靶点以提高疫苗效力。研究计划：目标1a将确定T的特异性 其中最常见的细胞出现在HSV-2重新激活的区域，集中在T细胞在 在免疫治疗疫苗接种后的数量，因为这些是最有可能是疱疹特异性的、组织驻留的 细胞。目标1b侧重于在小切片活检中确定这些细胞的空间位置 荧光分子标签，以可视化它们在皮肤中的排列。目标2侧重于了解更多关于这些T 通过使用转录图谱，这是一种在活检中分离每个T细胞并查看 一个接一个地揭示定义每个细胞活动的细胞过程。我们将把 来自TCR谱系分析的信息，以更有效地解释转录特征分析的结果 准确地说。这种方法以最全面的方式使用小皮肤样本来描述 HSV-2和人类免疫系统之间的斗争。通过与一位经验丰富的 以及在FHCRC和UW丰富的教育环境中独一无二的熟练导师团队，福特博士将 成为一名高效高效的首席调查员。

PROJECT SUMMARY / ABSTRACT Summary of Proposal: This proposal supports a 5-year mentored career development award investigating the tissue-based immune response through the lens of a recent phase 1 immunotherapeutic vaccine trial with the long-term goal of contributing to the development of an effective herpes simplex virus-2 (HSV-2) vaccine and improving global health.. This trial involved three sequential doses of an immunotherapeutic vaccine, HSV529, in individuals with symptomatic HSV-2 infection. T-cell receptor (TCR) repertoire analysis was performed of T cells in small biopsies of the area of HSV-2 reactivation in comparison to T cells from biopsies of uninvolved skin and HSV-2 reactive CD4+ T cells from peripheral blood to research two main lines of inquiry. The first is the interrelatedness, over time, of the TCR repertoire of the genital skin at HSV-2 reactivation sites, the arm, and HSV-2 specific CD4+ T cells over time; whether T cells detected in genital tissue are detectable in blood with standard sampling techniques. The second is how these relationships and dynamics change over the course of an immunotherapeutic vaccination. Preliminary findings suggest that some T cells increased in number after vaccination in everyone, with very little overlap between the skin and the blood. The central hypothesis is that T cells that respond to a vaccine targeting HSV-2 are likely to be HSV-2-specific, and that T cells seen in tissue at high copy number over a long period of time are likely to be tissue residents. The objective of this proposal is to learn more about these resident T cells, which are otherwise difficult to study. The rationale for this is that vaccine development for HSV-2 has thus far not been successful, and that because there is so little overlap between the skin and the blood, that looking directly at the tissue may help identify a viral antigen or a way to target skin-based T cells to improve vaccine efficacy. Research plan: AIM 1a will identify the specificity of T cells of that were seen most frequently in the region of HSV-2 reactivation, focusing on T cells that expanded in number after immunotherapeutic vaccination, as these are the most likely to be herpes-specific, tissue-resident cells. AIM 1b focuses on establishing the spatial location of these cells in small sections of biopsies using fluorescent molecular tags to visualize their arrangement in skin. AIM 2 focuses on learning more about these T cells by using transcriptional profiling, a technique that separates each of the T cells in a biopsy and looks at them one by one to uncover the cellular processes that define the activity of each cell. We will incorporate information from TCR repertoire analysis to interpret the results of transcriptional profiling more effectively and accurately. This approach uses small skin samples in the most comprehensive manner available to describe the fight between HSV-2 and the human immune system. By conducting this research with an extremely experienced and uniquely skilled group of mentors in the rich educational environment of the FHCRC and UW, Dr. Ford will become an effective and productive lead investigator.