Host and viral genomic determinants of HIV latent reservoir size and characteristics in individuals with substance use disorders

艾滋病毒潜伏库大小和物质使用障碍个体特征的宿主和病毒基因组决定因素

• 批准号: 10619974
• 负责人: ROBERT F SILICIANO
• 金额: $ 84.39万
• 依托单位: JOHNS HOPKINS UNIVERSITY
• 依托单位国家: 美国
• 财政年份: 2018
• 资助国家: 美国
• 起止时间: 2018-08-15 至 2024-05-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/10619974
• 关键词: Affect; Antigens; Autologous; Biological Assay; Biology; CD4 Positive T Lymphocytes; CD8B1 gene; Cell physiology; Cells; Characteristics; Chronic; Clonality; Cocaine; Cocaine Users; Defect; Drug usage; Drug user; Effector Cell; Failure; Follow-Up Studies; Genomic approach; Grant; HIV; HIV-1; Heroin; Homeostasis; Immune; Individual; Infection; Inflammation; Inflammatory; Injecting drug user; Interruption; Measurement; Measures; Mediating; Memory; Molecular; Mutation; Participant; Pathway interactions; Pattern; Persons; Pharmaceutical Preparations; Phase; Plasma; Population; Property; Proviruses; Research Personnel; Resources; Rest; Risk Factors; Sampling; Sequence Analysis; Specificity; Substance Use Disorder; T-Cell Activation; T-Lymphocyte; T-Lymphocyte Epitopes; T-Lymphocyte Subsets; T-Lymphocyte and Natural Killer Cell; Testing; base; cocaine use; cohort; curative treatments; design; digital; experience; genome sequencing; heroin use; high throughput screening; injection drug use; integration site; latent HIV reservoir; molecular subtypes; novel; pathogen exposure; response; transcriptome sequencing; transcriptomics; viral genomics; viral rebound; whole genome; Affect; Antigens; Autologous; Biological Assay; Biology; CD4 Positive T Lymphocytes; CD8B1 gene; Cell physiology; Cells; Characteristics; Chronic; Clonality; Cocaine; Cocaine Users; Defect; Drug usage; Drug user; Effector Cell; Failure; Follow-Up Studies; Genomic approach; Grant; HIV; HIV-1; Heroin; Homeostasis; Immune; Individual; Infection; Inflammation; Inflammatory; Injecting drug user; Interruption; Measurement; Measures; Mediating; Memory; Molecular; Mutation; Participant; Pathway interactions; Pattern; Persons; Pharmaceutical Preparations; Phase; Plasma; Population; Property; Proviruses; Research Personnel; Resources; Rest; Risk Factors; Sampling; Sequence Analysis; Specificity; Substance Use Disorder; T-Cell Activation; T-Lymphocyte; T-Lymphocyte Epitopes; T-Lymphocyte Subsets; T-Lymphocyte and Natural Killer Cell; Testing; base; cocaine use; cohort; curative treatments; design; digital; experience; genome sequencing; heroin use; high throughput screening; injection drug use; integration site; latent HIV reservoir; molecular subtypes; novel; pathogen exposure; response; transcriptome sequencing; transcriptomics; viral genomics; viral rebound; whole genome

Project Summary/Abstract Using the unique resources of the ALIVE cohort of persons who inject drugs (PWID) led by study co- investigator Gregory Kirk and the extensive experience in reservoir biology provided by co-investigators Rafick Sekaly, Joel Blankson, Greg Laird, and study PI Robert Siliciano, we will investigate the effects of heroin and cocaine use on the latent reservoir for HIV-1 in CD4+ T cells, which is the major barrier to cure. We hypothesize that by affecting inflammatory and immune pathways, active heroin and/or cocaine may alter T cell homeostasis and the distribution of HIV-1 proviruses among memory cells subsets, thereby affecting the size, turnover, and inducibility of the latent reservoir. In addition, we hypothesize that past injection drug use and the associated unstructured treatment interruptions will permanently affect the size and inducibility of the latent reservoir. Active drug use and legacy effects of past drug use may also affect that ability of host effector cells to clear infected cells during curative interventions. These hypotheses will be tested using samples from ALIVE cohort participants and experimental approaches informed by full genome proviral sequencing in order to achieve a better understanding of the reservoir in PWID and inform how curative strategies can be applied to this population. We will first determine the effects of active heroin/cocaine use on the latent reservoir by quantifying the size of the reservoir in resting CD4+ T cells and T cell subsets in active drug users with a novel high-throughput assay for intact proviruses. This assay is based on full genome sequence analysis by the Siliciano lab that led to the discovery that most proviruses (98%) are defective. Thus we will use a novel digital droplet PCR assay that selectively detects proviruses that are intact and have the potential to cause viral rebound. We will also measure the inducibility of latent proviruses in response to T cell activation and latency reversing agents using the TILDA assay developed by Dr. Sekaly and colleages. In addition, we will carry out full genome sequencing of HIV-1 proviruses to define additional properties that are relevant to HIV-1 cure including replication capacity, clonality, turnover rates, and escape mutations in cytolytic T lymphocyte (CTL) epitopes. These studies will be carried out in PWID who are actively using as well as those who are no longer using in order to distinguish active and legacy effects of drug use. We will also measure the ability of autologous CD8+ CTL and NK cells from active drug users to eliminate autologous infected cells following latency reversal and carry out similar studies in PWID who are no longer actively using to distinguish direct drug effects from legacy effects of drug past drug use on effector cell function. Together these studies of the size, inducibility, and composition of the reservoir, and the ability of host effector cells to clear infected cells, should provide critical information for the design of cure strategies for PWID and guide longitudinal and mechanistic studies in the second phase of the grant.

项目摘要/摘要 利用研究毒品（PWID）的活着队列的独特资源（PWID） 研究人员格雷戈里·柯克（Gregory Kirk）和共同投资者Rafick提供的水库生物学的丰富经验 Sekaly，Joel Blankson，Greg Laird和研究Pi Robert Siliciano，我们将研究海洛因和 可卡因用于在CD4+ T细胞中用于HIV-1的潜在储层，这是治愈的主要障碍。我们 假设通过影响炎症和免疫途径，主动海洛因和/或可卡因可能会改变T细胞 稳态和HIV-1病毒在记忆细胞子集中的分布，从而影响大小 失误和潜在储层的诱导性。此外，我们假设过去的注射药物使用和 相关的非结构化治疗中断将永久影响潜在的大小和诱导性 水库。过去药物使用的主动药物使用和遗产影响也可能影响宿主效应细胞的能力 在治疗干预期间清除感染细胞。这些假设将使用来自 活着的队列参与者和通过完整基因组病毒测序告知的实验方法 为了更好地了解PWID中的储层并告知如何应用治疗策略 对这个人群。我们将首先确定主动海洛因/可卡因使用对潜在储层的影响 用新颖的活性吸毒者中的静止CD4+ T细胞和T细胞子集量化储层的大小 完整预科病毒的高通量测定法。该测定基于完整的基因组序列分析 硅体实验室导致发现大多数病毒（98％）有缺陷。因此，我们将使用一个新颖的数字 液滴PCR测定法检测完整的原病毒，并具有引起病毒的潜力 反弹。我们还将测量潜在病毒对T细胞激活和延迟的诱导性 使用Sekaly博士和Loceages开发的Tilda分析的逆转剂。此外，我们将执行 HIV-1 Provires的完整基因组测序，以定义与HIV-1治疗相关的其他特性 包括复制能力，克隆性，周转率和溶质T淋巴细胞（CTL）中的逃生突变 表位。这些研究将在PWID中进行，他们正在积极使用以及不再使用的研究 为了区分吸毒的主动和遗产效应。我们还将衡量 活跃吸毒者的自体CD8+ CTL和NK细胞，以消除自体感染细胞 潜伏期逆转并在不再积极使用直接的PWID的PWID中进行类似的研究 药物对过去药物使用对效应细胞功能的遗产影响的药物影响。这些研究共同 储层的大小，诱导性和组成，以及宿主效应细胞清除感染细胞的能力， 应该为PWID的治疗策略设计和指导纵向的策略提供关键信息 机械研究在赠款的第二阶段。