Understanding HIV latency reversal and clearance of infected cells in vivo

了解 HIV 潜伏期逆转和体内受感染细胞的清除

• 批准号: 9975686
• 负责人: Joseph K Wong
• 金额: $ 38.33万
• 依托单位: NORTHERN CALIFORNIA INSTITUTE/RES/EDU
• 依托单位国家: 美国
• 财政年份: 2017
• 资助国家: 美国
• 起止时间: 2017-07-21 至 2022-06-30
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/9975686
• 关键词: Affect; Aftercare; Agonist; Agreement; Antigens; Biological Assay; Blood; Blood specimen; CD4 Positive T Lymphocytes; Cell Line; Cell model; Cells; Cessation of life; Clinical Trials; DNA; Data; Development; Distal; Disulfiram; Genetic Transcription; HIV; HIV Infections; Histone Deacetylase Inhibitor; Human; Immune; Immune response; In Vitro; Inflammation; Knowledge; Laboratories; Lead; Measures; Mediating; Modeling; New Agents; Organ; Patients; Pharmaceutical Preparations; Plasma; Polyadenylation; Primates; Production; Proteins; RNA Splicing; Sampling; Series; System; TLR7 gene; Testing; Time; Tissue Sample; Toll-like receptors; Transcript; Transcription Elongation; Transcription Initiation; Viral; Virus; Vorinostat; antiretroviral therapy; base; cell killing; defense response; in vitro Model; in vivo; insight; latent infection; novel; novel drug combination; response; success

Project Summary/Abstract: Latently-infected CD4+ T cells are thought to be the main barrier to HIV eradication or functional cure, and viral reactivation from these cells likely contributes to the organ inflammation and damage observed on antiretroviral therapy. Major impediments to the development of more effective latency-reversing agents (LRAs) include the lack of knowledge about the mechanisms that govern latency and latency reversal in vivo, and the degree to which these are recapitulated by latency models in vitro. The lack of agreement between latency models and the incomplete success of human trials with LRAs suggest that it is critical to understand why different LRAs do or do not work in vivo. We have developed a new “transcription profiling” approach that can simultaneously measure the degree to which different mechanisms contribute to reversible inhibition of HIV transcription in vivo. By applying this approach to cells from ART-suppressed patients, we have generated preliminary data suggesting a new paradigm in which latency is not (as commonly assumed) due to a block to HIV transcriptional initiation, the block to proximal elongation is greater and more pervasive than previously realized, and the main reversible blocks to HIV transcription are a previously-unrecognized block to distal transcription/polyadenylation (completion) and a block to multiple-splicing. In addition, we have intriguing new data suggesting that LRAs may act selectively on the different mechanistic blocks to HIV transcription. This study will utilize samples from clinical trials of humans treated with LRAs (aims 1 and 2) to better understand how they reverse the mechanisms of latency in vivo and to identify the optimum model to test new agents in vitro (aim 3). In aim 1, we will apply our transcription profiling approach to samples from humans treated with disulfiram, vorinostat, panobinostat, and romidepsin. We hypothesize that these agents preferentially increase HIV transcriptional initiation and elongation but have less ability to overcome blocks to completion and splicing. In aim 2, we will apply our approach to blood and gut samples from clinical trials of humans treated with agonists of toll-like receptor (TLR) 7 and 9 to understand how these agents reverse latency and lead to death of infected cells in vivo. We hypothesize that TLR agonists can overcome later blocks to HIV transcription, increasing the completed transcripts (and HIV protein/antigen) that may facilitate clearance by intrinsic cell defenses or immune killing. In aim 3, we will compare in vitro models of latency based on the degree to which they recapitulate in vivo mechanisms of latency and responses to LRAs. We will then select the best model and test new combinations of agents for their ability to increase completed/spliced transcripts and lead to death of infected cells. The results from these 3 aims should provide critical new insights on the degree to which existing LRAs reverse the different mechanisms of latency in vivo (aims 1 and 2), the effects that correlate with clearance of reactivated cells (aims 2 and 3), the best system to study LRAs in the laboratory (aim 3), and new combinations that can lead to more effective latency reversal and/or killing of infected cells (aim 3).

项目摘要/摘要： 潜伏感染的CD4+T细胞被认为是根除艾滋病毒或治愈功能的主要障碍， 从这些细胞中重新激活病毒可能会导致观察到的器官炎症和损伤 抗逆转录病毒治疗。开发更有效的潜伏期反转剂的主要障碍 (LRA)包括缺乏关于体内潜伏期和潜伏期逆转的机制的知识， 以及这些数据在多大程度上被体外潜伏期模型概括。两国之间缺乏一致意见 潜伏期模型和LRA人体试验的不完全成功表明，理解 为什么不同的LRA在体内起作用或不起作用。我们已经开发出一种新的“转录分析”方法， 可以同时测量不同机制对可逆抑制的贡献程度 HIV在体内转录。通过将这种方法应用于ART抑制患者的细胞，我们已经产生了 初步数据表明了一种新的范式，其中延迟不是(通常假设的)由于阻塞到 HIV转录启动，对近端延伸的阻碍比以前更大、更普遍 HIV转录的主要可逆阻断是以前未被识别的远端阻断 转录/多聚腺苷化(完成)和阻止多重剪接。此外，我们还有耐人寻味的新产品 数据表明，LRA可能选择性地作用于HIV转录的不同机制障碍。这 这项研究将利用使用LRAS(目标1和2)治疗的人类临床试验的样本来更好地了解 他们如何逆转体内潜伏期的机制，并确定测试新药物的最佳模型 体外实验(目标3)。在目标1中，我们将应用我们的转录图谱方法来处理来自人类的样本 双硫仑、安非他明、帕诺比妥和罗米迪辛。我们假设这些药剂优先增加 HIV的转录起始和延长，但克服完成和剪接障碍的能力较弱。 在目标2中，我们将把我们的方法应用于从临床试验中采集的血液和肠道样本，这些样本来自于使用 Toll样受体(TLR)7和9的激动剂，以了解这些药物如何逆转潜伏期和导致死亡 体内感染细胞的数量。我们假设TLR激动剂可以克服后来对HIV转录的阻断， 增加完整的转录本(和HIV蛋白/抗原)，这可能有助于内在细胞的清除 防御或免疫杀戮。在目标3中，我们将基于以下程度比较潜伏期的体外模型 它们概括了体内潜伏期和对LRA的反应机制。然后我们将选出最好的型号 并测试新的药物组合增加完整/剪接转录本并导致死亡的能力 被感染的细胞。这三个目标的结果应该会为我们在多大程度上提供关键的新见解 现有的LRA逆转了体内潜伏期的不同机制(目标1和2)，这些影响与 清除重新激活的细胞(目标2和3)，实验室研究LRA的最佳系统(目标3)，以及新的 可导致更有效的潜伏期逆转和/或杀死受感染细胞的组合(目标3)。