Irreversible Proviral Silencing in Myeloid Cells

骨髓细胞中不可逆的前病毒沉默

• 批准号: 10705469
• 负责人: Mario Stevenson
• 金额: $ 19.19万
• 依托单位: UNIVERSITY OF MIAMI SCHOOL OF MEDICINE
• 依托单位国家: 美国
• 财政年份: 2023
• 资助国家: 美国
• 起止时间: 2023-08-02 至 2025-07-31
• 项目状态: 未结题
• 来源: https://reporter.nih.gov/project-details/10705469
• 关键词: Alveolar Macrophages; Anatomy; Attention; Binding; CD4 Positive T Lymphocytes; CD8-Positive T-Lymphocytes; Cell Compartmentation; Cell physiology; Cells; Central Nervous System; Chemicals; Clinical; Competence; Development; Dominant-Negative Mutation; Epigenetic Process; Excision; Exhibits; Gene Expression; Genetic Transcription; HIV; HIV-1; HIV/AIDS; I-kappa B Proteins; Individual; Infection; Interruption; Kupffer Cells; Link; Liver; Lung; Macaca; Macrophage; Mediating; Methods; Methylation; Microglia; Modeling; Modification; Myelogenous; Myeloid Cells; NF-kappa B; Natural Killer Cells; Phagocytes; Phylogenetic Analysis; Physiological; Plasma; Population; Process; Production; Proviruses; Refractory; Research; Sampling; T-Lymphocyte; Therapeutic; Viral; Viral reservoir; Viremia; Virion; Virus; Virus Activation; Virus Latency; Virus Replication; antagonist; cancer therapy; cell killing; cytokine; cytotoxic CD8 T cells; epigenetic silencing; experimental analysis; experimental study; heterokaryon; in vivo; induced pluripotent stem cell; inhibitor; molecular clock; monocyte; novel; novel strategies; pharmacologic; resilience; simian human immunodeficiency virus

ABSTRACT: In the pursuit of strategies with which to cure HIV-1 infection, most of the attention has been on mechanisms that govern HIV-1 persistence in CD4+ T cell reservoirs and on approaches to limit persistence in those reservoirs. We recently provided evidence [1] that macrophages enable viral persistence in HIV-1-infected individuals on effective ART. We showed that rebound viremia from individuals undergoing an analytic treatment interruption (ATI) comprised highly macrophage-tropic viruses and we used a novel virion immunocapture method to demonstrate that those macrophage-tropic viruses in post-ATI plasma had a myeloid cell origin. Furthermore, Molecular Clock analysis indicated that macrophage-tropic viruses in plasma pre-dated the treatment interruption as opposed to having been created during the treatment interruption interval. Given these lines of evidence that myeloid cells can serve as viral reservoirs under effective ART, we set about establishing a latency model in primary macrophages. In the pursuit of this model of macrophage latency, we observed that NF-kB inhibition, originally intended to promote synchronous latency, led to irreversible loss of proviral competence- proviruses were refractory to latency reactivating agents (LRAs) even though host cell functions remained intact. On the basis of these observations, we hypothesize that the provirus is epigenetically modified under conditions where NF-kB is limiting and that sustained transcriptional activity is required to counteract epigenetic silencing processes of the host cell. In this proposal, we will assess the mechanisms governing loss of proviral competence. Aim 1: Assess correlates of proviral silencing in NF-kB-depleted macrophages. We will determine the stage in the viral replication cycle at which proviruses are rendered inactive and, through heterokaryon formation, whether dominant negative factors mediate irreversible proviral silencing by NF-kB inhibition. We will examine whether inactive proviruses exhibit evidence of epigenetic modification and methylation and whether inhibiting methylation can maintain proviral responsiveness to reactivation in the face of NF-kB inhibition. Aim 2: Examine whether the proviral silencing created under conditions of NF-kB inhibition can be extended to infected CD4+ T cells from ART-suppressed subjects and to other primary macrophage populations. We will assess whether NF-kB inhibition can irreversibly impede HIV-1 reactivation in CD4+ T cells from individuals on effective ART and whether this proviral silencing can also be achieved in macrophages that are physiologically representative of the major myeloid cell compartments including lung (alveolar macrophages), liver (Kupffer cells), and Central Nervous System (microglia). If the proposed studies are successful, they will provide the rationale for in vivo studies using established SHIV/macaque models of macrophage reservoirs and NF-kB antagonists currently in clinical use.

抽象的： 为了追求治愈HIV-1感染的策略，大多数关注一直存在 控制CD4+ T细胞库中HIV-1持久性的机制以及限制持久性的方法 那些水库。我们最近提供了证据[1]表明，巨噬细胞在HIV-1感染中启用病毒持久性 有效艺术的个人。我们表明，接受分析治疗的个体的反弹病毒血症 中断（ATI）包含高巨噬细胞 - 冰球病毒，我们使用了一种新型的病毒体免疫接触 证明ATI后血浆中那些巨噬细胞 - 循环病毒的方法具有髓样细胞的起源。 此外，分子时钟分析表明，血浆中的巨噬细胞 - 热带病毒预先预先 治疗中断而不是在治疗中断间隔中创建的中断。 鉴于以下证据表明髓样细胞可以在有效的艺术下用作病毒库，我们设定 关于在主要巨噬细胞中建立潜伏期模型。在追求这种巨噬细胞潜伏期的模型时， 我们观察到NF-KB抑制作用最初旨在促进同步潜伏期，导致不可逆转的损失 病毒疗法 - 预科病毒对延迟重新激活（LRA）是难治性的，即使宿主细胞 功能保持完整。根据这些观察，我们假设该病毒是表观遗传的 在NF-KB受到限制并且需要持续的转录活动的条件下进行修改 宿主细胞的表观遗传沉默过程。在此提案中，我们将评估机制 管理病毒能力的丧失。 AIM 1：评估NF-KB缺失巨噬细胞中前病毒沉默的相关性。我们将确定 病毒复制周期的阶段，在该阶段使病毒呈现不活跃，并通过异性疾病 形成，主要的负面因素是否通过NF-KB抑制介导不可逆的前病毒沉默。我们将 检查非活性病毒是否表现出表观遗传修饰和甲基化的证据 面对NF-KB抑制作用，抑制甲基化可以保持对重新激活的反应性。 目标2：检查在NF-KB抑制条件下产生的前病毒沉默是否可以 扩展到来自抑制Art抑制的受试者的感染CD4+ T细胞和其他原发性巨噬细胞 人群。我们将评估NF-KB抑制是否可以不可逆转地阻碍CD4+ T细胞中的HIV-1重新激活 从有效艺术的个人以及在巨噬细胞中是否也可以实现这种预防的沉默 在生理上代表了主要髓样细胞室，包括肺（肺泡巨噬细胞）， 肝（库普弗细胞）和中枢神经系统（小胶质细胞）。 如果拟议的研究成功，它们将使用已建立的体内研究为体内研究提供理由 巨噬细胞储层和NF-KB拮抗剂的湿夫/猕猴模型目前用于临床用途。