Identifying Roadblocks to Antigen Expression and Enhancing Killing of HIV-Infected Cells That Are Refractory to Clearance

项目摘要 HIV-1储存库被认为含有对宿主免疫清除敏感性不同的细胞， 系统识别允许受感染细胞避免清除的机制可以为治愈的发展提供信息 这些策略能够更有效地消除长寿水库中的HIV感染细胞。持久性 在长期未经治疗的感染期间，当对HIV感染细胞的免疫监视最高时， 免疫环境是高度炎症性的，可以选择具有增强的逃避能力的感染细胞， 被宿主免疫系统检测和清除。这项研究将集中在两组水库细胞， 携带完整的前病毒基因组，可能在清除率方面不同。第一种是“早期”储库细胞， 在未经治疗的感染早期被感染（或者是被感染细胞的克隆）。二是“晚期”储层细胞 在ART开始时被感染（或者是感染细胞的克隆）。这些“晚期”储库细胞 在储存库中持续时间较短，主要在抗逆转录病毒治疗期间持续存在， 特异性免疫反应减弱，HIV感染的细胞通常具有很长的半衰期。与此相反， “早期”储库细胞在未经治疗的感染中持续了很长时间，而大多数HIV感染细胞在感染后的短时间内都没有存活。 半衰期我们假设早期的前病毒具有表观遗传特征，使它们对T细胞免疫具有抗性。 刺激，不易清除。在这项研究中，我们将鉴定完整的“早期”和“晚期”前病毒， 从接受抑制性ART的15名参与者的血液中分离的CD 4 + T细胞（目的1a）。然后我们将检查 预测影响储层清除的表型。具体来说，我们将研究是否“早”和“晚” 前病毒对T细胞刺激的敏感性不同（Aim 1b）和/或具有不同的遗传（前病毒序列 和整合位点）或表观遗传特征（Aim 1c）。为了验证“早期”前病毒比其他病毒更容易感染的假设， 由于难以明确，我们将测量“早期”和“晚期”前病毒对自体CD 8 T细胞杀伤的敏感性， 细胞（目标2a），并评估表观遗传调节因子的破坏是否改变了它们对通过 自体CD 8 T细胞（Aim 2b）。我们对“早期”和“晚期”病毒的关注使我们能够探索潜在的 与清除相关的机制可能在对清除的敏感性方面存在差异的人群中， 然而，这项提案的工作也将揭示除了前病毒年龄以外的其他特征是否可以预测对 间隙然后，我们将探索改善感染细胞中抗原呈递的策略（Aim 3a），并评估 在通常情况下，改善的抗原呈递是否可以增强群体中自体CD 8 T细胞的杀伤 清除难治性（目标3b）。这个项目的首要目标是定义一个细胞群， 提高逃避清除的能力，识别与逃避清除相关的宿主和前病毒特征， 测试是否更好的抗原呈递可以使HIV感染细胞更容易被清除。成功 这些目标的完成将把前病毒遗传学和表观遗传学与病毒的再活化和杀死联系起来， 制定更好的排雷战略。

PROJECT ABSTRACT The HIV-1 reservoir is thought to contain cells that vary in their susceptibility to clearance by the host immune system. Identifying mechanisms that allow infected cells to avoid clearance could inform the development of cure strategies that are able to more effectively eliminate HIV-infected cells in the long-lived reservoir. Persistence during long periods of untreated infection, when immune surveillance for HIV-infected cells is highest and the immune environment is highly inflammatory, may select for infected cells with an enhanced ability to evade detection and clearance by the host immune system. This study will focus on two groups of reservoir cells that carry intact proviral genomes and likely differ in susceptibly to clearance. The first are "early" reservoir cells that were infected (or are a clone of a cell infected) early in untreated infection. The second are "late" reservoir cells that were infected (or are a clone of a cell infected) near the time of ART initiation. These "late" reservoir cells have persisted in the reservoir for less time and have primarily persisted during antiretroviral therapy when HIV- specific immune responses are blunted and HIV-infected cells typically have very long half-lives. In contrast, "early" reservoir cells persisted for long periods of untreated infection when most HIV-infected cells have a short half-life. We hypothesize that early proviruses will have epigenetic features that make them resistant to T cell stimulation and less susceptible to clearance. In this study we will identify intact "early" and "late" proviruses in CD4+ T cells isolated from the blood of 15 participants on suppressive ART (Aim 1a). We will then examine phenotypes predicted to impact reservoir clearance. Specifically, we will examine whether "early" and "late" proviruses differ in their sensitivity to T cell stimulation (Aim 1b) and/or have different genetic (proviral sequence and integration site) or epigenetic features (Aim 1c). To test the hypothesis that "early" proviruses are more difficult to clear, we will measure the susceptibility of "early" and "late" proviruses to killing by autologous CD8 T cells (Aim 2a) and assess whether disruption of epigenetic regulators alters their susceptibility to killing by autologous CD8 T cells (Aim 2b). Our focus on "early" and "late" viruses allows us to explore potential mechanisms associated with clearance in populations that likely differ in their susceptibility to clearance, however, the proposal work will also reveal if features other than proviral age are predictive of susceptibility to clearance. We will then explore strategies to improve antigen presentation in infected cells (Aim 3a) and assess whether improved antigen presentation can enhance killing by autologous CD8 T cells in populations typically refractory to clearance (Aim 3b). The overarching goal of this project is to define a population of cells with an enhanced ability to evade clearance, identify host and proviral features associated with clearance evasion and test whether better antigen presentation can make HIV-infected cells more susceptible to clearance. Successful completion of these aims will connect proviral genetics and epigenetics to virus reactivation and killing to inform the development of improved clearance strategies.