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

项目概要/摘要 提案摘要：该提案支持一项为期5年的职业发展指导奖， 通过最近的一期免疫接种试验的透镜， 长期目标是促进开发有效的单纯疱疹病毒2型（HSV-2）疫苗， 改善全球健康该试验涉及三个连续剂量的免疫接种疫苗，HSV 529， 在有症状的HSV-2感染的个体中。T细胞受体（TCR）库分析进行了T HSV-2再活化区域的小活检组织中的T细胞与来自未受累皮肤活检组织的T细胞相比 和外周血HSV-2反应性CD 4 + T细胞的研究两条主线。首先是 随着时间的推移，生殖器皮肤在HSV-2再活化位点、手臂和 HSV-2特异性CD 4 + T细胞随时间的变化;生殖器组织中检测到的T细胞是否在血液中可检测到， 标准抽样技术。第二个问题是这些关系和动态在整个过程中是如何变化的。 免疫接种初步的研究结果表明，一些T细胞的数量增加后， 疫苗接种在每个人身上，皮肤和血液之间的重叠很少。核心假设是， 对靶向HSV-2的疫苗产生反应的T细胞可能是HSV-2特异性的，并且在组织中观察到的T细胞可能是HSV-2特异性的。 在高拷贝数在很长一段时间很可能是组织居民。本提案的目的 就是更多地了解这些常驻T细胞，否则很难研究。其理由是， HSV-2的疫苗开发迄今为止还没有成功，这是因为重叠太少， 直接观察组织可能有助于识别病毒抗原或一种方法， 靶向基于皮肤的T细胞以提高疫苗效力。研究计划：AIM 1a将确定T 其中的T细胞最常见于HSV-2再活化区域，主要集中在HSV-2再活化区域中扩增的T细胞。 免疫接种后的数量，因为这些是最有可能是疱疹特异性，组织驻留 细胞AIM 1b侧重于建立这些细胞在小切片活检中的空间位置， 荧光分子标签，以可视化它们在皮肤中的排列。AIM 2侧重于了解更多关于这些T 通过使用转录谱分析，一种在活检中分离每个T细胞并观察 他们一个接一个地揭示细胞过程，定义每个细胞的活动。我们将合并 - 来自TCR库分析的信息，以更有效地解释转录谱分析的结果， 准确地这种方法以最全面的方式使用小的皮肤样本来描述 HSV-2与人体免疫系统的对抗通过与一位经验丰富的 在FHCRC和华盛顿大学丰富的教育环境中，福特博士将 成为一名高效的首席调查员。

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.