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

项目摘要 HIV-1病毒宿主被认为含有不同的细胞，它们对宿主免疫清除的敏感性不同 系统。识别允许感染细胞避免清除的机制可以为治愈的发展提供信息 能够更有效地消除长寿水库中感染艾滋病毒的细胞的战略。持久性 在长期未经治疗的感染期间，当对艾滋病毒感染细胞的免疫监测最高时， 免疫环境是高度炎性的，可能会选择具有增强躲避能力的感染细胞 由宿主免疫系统进行检测和清除。这项研究将集中在两组储集层细胞上 携带完整的前病毒基因组，很可能在清除方面有所不同。第一类是“早期”的储集层细胞 在未经治疗的感染早期被感染(或被感染的细胞的克隆)。第二类是“晚期”储集层单元 在抗逆转录病毒治疗开始时被感染的(或被感染的细胞的克隆)。这些“晚”的储集层细胞 在水库中持续的时间较短，主要在抗逆转录病毒治疗期间持续，当艾滋病毒- 特定的免疫反应迟钝，感染艾滋病毒的细胞通常有很长的半衰期。相比之下， “早期”存储细胞在未经治疗的感染中持续很长时间，而大多数感染艾滋病毒的细胞都有较短的 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细胞产生抵抗力 刺激性强，不易被清除。在这项研究中，我们将识别完整的“早期”和“晚期”前驱语 从15名接受抗逆转录病毒治疗的参与者的血液中分离出CD4T细胞(目标1a)。然后我们将检查 表型预测会影响储集层清除。具体地说，我们将审查“早”和“晚” 前病毒对T细胞刺激的敏感性不同(目标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(目标1c)。为了检验“早期”前驱虫比 很难明确，我们将测量“早期”和“晚期”前病毒对自体CD8 T杀伤的易感性 细胞(目标2a)，并评估表观遗传调控因子的干扰是否改变了它们对 自体CD8 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 在对清除敏感性可能不同的人群中，与清除相关的机制， 然而，这项提案工作也将揭示除前驱年龄以外的其他特征是否预示着易感性 通行证。然后，我们将探索改进感染细胞中抗原提呈的策略(目标3a)，并评估 在人群中，改进的抗原提呈是否可以增强自体CD8 T细胞的杀伤作用 难以清除(目标3b)。该项目的总体目标是使用 增强了逃避通关的能力，识别与通关逃避相关的主机和前病毒特征，以及 测试更好的抗原呈递是否可以使感染艾滋病毒的细胞更容易被清除。成功 完成这些目标将把前病毒遗传学和表观遗传学与病毒重新激活和杀死联系起来，以告知 制定改进的清除战略。

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.