Coupling Epitranscriptomics to Molecular Disease Mechanisms and Nucleic Acid Therapeutics in Persistent Residual HIV Infection

表观转录组学与持续残留 HIV 感染的分子疾病机制和核酸治疗的耦合

• 批准号: 10907304
• 负责人: Keith Thomas Gagnon
• 金额: $ 24.88万
• 依托单位: WAKE FOREST UNIVERSITY HEALTH SCIENCES
• 依托单位国家: 美国
• 财政年份: 2022
• 资助国家: 美国
• 起止时间: 2022-05-01 至 2025-03-31
• 项目状态: 未结题
• 来源: https://reporter.nih.gov/project-details/10907304
• 关键词: Coupling; Epitranscriptomics; Molecular; Disease; Mechanisms

PROJECT SUMMARY Human immunodeficiency virus (HIV) is a highly manageable infection when treated with combination anti-retroviral therapy (cART). However, it is not yet curable and people living with HIV (PLWH) taking cART suffer from suboptimal immune health and reduced quality of life long-term. A leading cause of ongoing health concerns is persistent, residual infection. Persistently infected T-cells of PLWH that take cART still produce viral RNA and proteins, as well as defective proviruses, that can chronically activate the immune system despite undetectable viral loads. A better understanding of the biology and metabolism of HIV RNA and gene expression in PLWH taking cART could unlock additional therapeutic strategies to reduce viral gene products and improve quality of life. Cellular RNA and viral RNA can be subjected to a variety of post-transcriptional chemical modifications. These modifications modulate or fine-tune molecular interactions and thereby control gene expression and function through RNA processing, turnover, localization, or translation. Covalent base modification and its effects on RNA regulation at a broad level beyond the four canonical bases is often termed epitranscriptomics. The intimate relationship between RNA modification and RNA metabolism is becoming better appreciated. However, manipulating this process for therapeutic treatment of HIV and persistent infection in PLWH taking cART requires a deeper understanding of HIV epitranscriptomics. Here we propose systematic characterization and manipulation of HIV-1 RNA transcripts in model and patient T-cells. In the R61 phase, we will utilize three powerful sequencing methodologies (total RNA-seq, ribosome profiling, and nanopore direct RNA sequencing) to globally characterize the abundance, processing, translation, and modification status of both viral and host RNA in T-cells during cART. These experiments will include selection of persistently infected T-cells from PLWH taking cART. We will target several known modifications via enzyme knockouts and nanopore sequencing. These experiments will characterize HIV-1 RNA chemical modifications of potential significance and identify those with high a likelihood of impacting HIV RNA and protein loads in T-cells. In the R33 phase, we will use chemically modified oligonucleotides to alter modifications of high interest by blocking or directing site-specific modification. This will include oligonucleotides that sterically block modification or guide new modification, both therapeutic modalities that are maturing rapidly. Chemically modified oligonucleotides will provide a proof-of-concept for epiptranscriptome manipulation as a potential therapeutic approach while offering nucleic acid therapeutic candidates.

项目概要 当联合治疗时，人类免疫缺陷病毒 (HIV) 是一种高度可控的感染 抗逆转录病毒治疗（cART）。然而，该疾病尚无法治愈，艾滋病毒感染者 (PLWH) 需要服用 cART 会导致免疫健康状况不佳并导致长期生活质量下降。一个主要原因是 持续的健康问题是持续的、残留的感染。感染者持续感染的 T 细胞 cART 仍然产生病毒 RNA 和蛋白质，以及有缺陷的原病毒，可以长期激活 尽管病毒载量无法检测到，但免疫系统仍然受到影响。更好地了解生物学和 接受 cART 的 PLWH 中 HIV RNA 的代谢和基因表达可能会带来额外的治疗效果 减少病毒基因产物和改善生活质量的策略。 细胞RNA和病毒RNA可以受到多种转录后化学作用 修改。这些修饰调节或微调分子相互作用，从而控制基因 通过 RNA 加工、周转、定位或翻译来表达和功能。共价碱基 修饰及其对超越四个经典碱基的广泛水平的 RNA 调节的影响通常是 称为表观转录组学。 RNA修饰与RNA代谢之间的密切关系是 变得更好的欣赏。然而，操纵这一过程来治疗艾滋病毒和 接受 cART 治疗的感染者持续感染需要更深入地了解 HIV 表观转录组学。 在这里，我们提出了模型中 HIV-1 RNA 转录本的系统表征和操作，以及 患者 T 细胞。在 R61 阶段，我们将利用三种强大的测序方法（总 RNA-seq、 核糖体分析和纳米孔直接 RNA 测序）来全面表征丰度， cART 期间 T 细胞中病毒和宿主 RNA 的加工、翻译和修饰状态。这些 实验将包括从接受 cART 的 PLWH 中选择持续感染的 T 细胞。我们将瞄准 通过酶敲除和纳米孔测序进行了一些已知的修饰。这些实验将 表征具有潜在意义的 HIV-1 RNA 化学修饰并鉴定那些具有高 a 影响 T 细胞中 HIV RNA 和蛋白质负载的可能性。 在R33阶段，我们将使用化学修饰的寡核苷酸来改变高 通过阻止或指导特定位点的修改来引起兴趣。这将包括空间上的寡核苷酸 阻止修改或指导新的修改，这两种治疗方式正在迅速成熟。 化学修饰的寡核苷酸将为表观转录组操作提供概念验证 作为一种潜在的治疗方法，同时提供核酸治疗候选者。