Mechanisms of HIV persistence in distinct memory CD4+ T cell subsets in blood and

描述(申请人提供)：现在很明显，单靠抗逆转录病毒疗法(ART)并不能根除艾滋病毒：即使经过15年以上的密集和持续治疗，病毒的传播在停止ART后的几周内都会恢复，但在特殊情况下除外。一小群潜伏感染的细胞，通常被称为艾滋病毒感染的“蓄水池”，提供了反弹病毒血症的长期来源。由于缺乏关于储存病毒的特定亚群的信息以及与保持潜伏期相关的机制，艾滋病毒的根除一直受到阻碍。我们先前已经证明，HIV持续存在于三个记忆T细胞亚群中，它们显示出不同的功能和生存能力，即病毒抑制受试者血液中的中枢(中医)、过渡性(TTM)和效应性(TEM)记忆CD4T细胞。有趣的是，我们的初步数据表明，这些细胞亚群也确保了HIV在组织储存库中的持久性，如淋巴结、直肠、结肠和回肠末端。因此，迫切需要更好地确定这三个细胞艾滋病毒储存库的特征，以实现根除它们。该项目的目标是确定在病毒抑制的受试者中能够导致潜伏感染细胞重新激活和随后消除的作用机制。我们假设导致HIV潜伏期的机制在中药、TTM和TTM中是不同的，并且这些储存库应该是不同类别的抗潜伏剂的靶点。我们将首先确定导致HIV在中医、TTM和TEM细胞中潜伏的分子机制(特定目标1)。我们将集中于DNA的表观遗传修饰，通过甲基化和组蛋白去乙酰化，关键转录因子的有限可获得性和新生小瓶转录本的低效延长。我们还将使用RNA测序来识别新的细胞通路，这些通路与24名病毒抑制受试者CD4T细胞亚群中潜伏库的大小相关。然后，我们将选择最有效的化合物组合，这些化合物将干扰这些机制，并允许消除所有细胞储藏(特定目标2)。我们选择了在以前的研究中显示出有希望的效果和/或目前在临床试验中测试的分子。由于针对单个细胞储备物或涉及病毒潜伏期的单一机制的靶向极有可能不成功，我们将测试来自每一类先导化合物的组合破坏来自病毒抑制受试者的潜伏期感染的CD4T细胞的能力。使用最近在佛罗里达VGTI开发的一种新的共培养试验，我们将评估这种抗潜伏期药物的重新激活是否可以导致自体HIV特异性CTL消除存储细胞。这项研究项目的成果是3-5种临床可耐受药物的组合，这些药物可以逆转来自病毒抑制受试者的真实潜伏感染细胞的潜伏期。这些研究的结果将构成设计一项旨在根除艾滋病毒的临床试验的基础。

DESCRIPTION (provided by applicant): It is now clear that antiretroviral therapy (ART) alone does not eradicate HIV: Even after more than 15 years of intensive and continuous therapy, the spread of the virus resumes within a few weeks upon cessation of ART in all but exceptional cases. A small pool of latently infected cells, usually referred as the "reservoir" of HIV infectio, provides a long-lived source of rebound viremia. Eradication of HIV has been hampered by the lack of information regarding the specific subsets harboring the viral reservoir and the mechanisms associated with the maintenance of latency. We have previously demonstrated that HIV persists in three memory T cell subsets displaying distinct functional and survival capacities, namely central (TCM) transitional (TTM) and effector (TEM) memory CD4+ T cells in the blood of virally suppressed subjects. Interestingly, our preliminary data indicate that these cell subsets also ensure HIV persistence in tissue reservoirs such as the lymph nodes, rectum, colon and terminal ileum. Therefore, there is an urgent need to better characterize these three cellular HIV reservoirs to achieve their eradication. The objective of this project is to identify mechanisms of action that can lead to the reactivation and subsequent elimination of latently infected cells in virally suppressed subjects. We hypothesize that the mechanisms responsible for HIV latency in TCM, TTM and TEM are different, and that these reservoirs should be targeted by different classes of anti-latency agents. We will first identify the molecular mechanisms responsible for HIV latency in TCM, TTM and TEM cells (Specific Aim 1). We will focus on epigenetic modification of the DNA through methylation and histone deacetylation, limited availability of critical transcription factors and inefficient elongation of the nascent vial transcripts. We will also use RNA sequencing to identify novel cellular pathways that correlate with the magnitude of the latent reservoir in the CD4+ T cell subsets from 24 virally suppressed subjects. We will then select the most efficient combination of compounds that will interfere with these mechanisms and allow elimination of all cellular reservoirs (Specific Aim 2). We have selected molecules that have shown promising effects in previous studies and/or that are currently tested in clinical trials. As it is highly likely that targeting a single cellular reservir or a single mechanism involved in viral latency will not be successful, we will test the ability of combinations of lead compounds from each class to disrupt latency in latently infected CD4+ T cells from virally suppressed subjects. Using a novel co-culture assay recently developed at VGTI Florida, we will assess if reactivation with this combination of anti-latency drugs can lead to the elimination of reservoir cells by autologous HIV specific CTL. The deliverable of this research project is a combination of 3-5 clinically tolerable agents that can reverse latency in authentic latently infected cells from virally suppressed subjects. The results from these studies will constitute the basis for the design of a clinical trial aimed at eradicating HIV.