Epigenetic HIV Silencing in Macrophages

巨噬细胞中的表观遗传 HIV 沉默

• 批准号: 10231275
• 负责人: Ronald G Collman
• 金额: $ 86.35万
• 依托单位: UNIVERSITY OF PENNSYLVANIA
• 依托单位国家: 美国
• 财政年份: 2017
• 资助国家: 美国
• 起止时间: 2017-08-01 至 2024-07-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/10231275
• 关键词: Address; Affect; Biological Assay; Biology; Brain; CD4 Positive T Lymphocytes; Cell Lineage; Cell model; Cells; Chemicals; Chromatin; DNA; Data; Development; Dioxygenases; Enhancers; Enzymes; Epigenetic Process; Future; Gene Silencing; Genetic; Genetic Transcription; Genome; HIV; HIV Genome; HIV Infections; HIV-1; HIV-associated neurocognitive disorder; Histones; Human; Immune Targeting; Industry Standard; Infection; Lead; Methods; Microglia; Modeling; Molecular; Molecular Target; Myeloid Cells; National Institute of Mental Health; Neurocognitive; Pathway interactions; Pharmaceutical Chemistry; Pharmaceutical Preparations; Pharmacodynamics; Pharmacology; Phase; Property; Proviruses; Regulation; Research; Residual state; Role; Site; Structure; T-Lymphocyte; Testing; Toxicology; Transcriptional Regulation; Validation; Viral; Viral reservoir; Virus; Virus Latency; Work; antiretroviral therapy; base; cell type; chemical genetics; chronic infection; demethylation; drug discovery; epigenetic regulation; epigenetic silencing; functional genomics; high throughput analysis; high throughput screening; histone demethylase; improved; inhibitor/antagonist; insight; interest; lead candidate; macrophage; neurocognitive disorder; novel; prevent; promoter; response; screening; small molecule; small molecule libraries

Project Summary/Abstract - Epigenetic HIV Silencing in Macrophages Antiretroviral therapy (ART) is highly effective in HIV infection but has substantial limitations, and strategies to achieve an aviremic state without ART (“functional cure”) are a focus of great interest. A potential approach to prevent persistent viral reservoirs from giving rise to replicating virus once ART is stopped is to silence HIV in these reservoirs so infection cannot “re-ignite”. Infected macrophage lineage cells serve as the main long-term reservoir for HIV in the CNS, which is believed to be a “sanctuary site” where virus can persist even if extra- CNS reservoirs are purged. Furthermore, long-lived infected macrophages may continue to produce low levels of virus, which contributes to HIV-associated neurocognitive disorders (HAND). Thus, silencing HIV in macrophages may both contribute to “functional cure”, and ameliorate HAND in the setting of ART suppression. HIV integrates into the host cell genome and is subject to positive and negative epigenetic regulation. Much work has been done on epigenetic control of HIV in T cells, but less is known in macrophages. Chromatin organization and transcriptional regulation is highly cell and context-specific. Our scientific premise is that macrophages in the CNS fail to epigenetically suppress HIV and serve as a long-term reservoir. We hypothesize that macrophage-specific enhancer-promoter interactions regulate persistent HIV transcription activity, and that small molecules can reprogram epigenetic regulation of HIV infected macrophages to establish long-term silencing of the integrated HIV genome. To this end, we have established a primary macrophage model for high-throughput screening of small molecule modulators of epigenetic enzymes, and provide preliminary evidence for involvement of dioxygenase enzymes involved in histone demethylation. In Phase 1, we will optimize primary & secondary assays for a high-throughput screening campaign to identify small molecule epigenetic regulators of HIV-1 expression in primary human macrophages; test the role of candidate epigenetic modulators, especially histone & DNA demethylases that may prevent HIV silencing; and further investigate molecular mechanisms that regulate persistent HIV transcription in macrophages. In Phase 2, we will perform a high-throughput screen to develop small molecules to silence HIV-1 in macrophages using industry-standard milestone-driven drug discovery pipeline to identify bioactive chemotypes; validate lead compounds with primary HIV isolates & myeloid cell types; advance drug-like properties of candidate hits through medicinal chemistry; and define molecular targets and conduct mechanism of action studies. At the end of Phase I we will have a robust high-throughput primary cell-based screen, orthogonal validation assays to confirm on-target hits, and further insight into macrophage-specific epigenetic factors regulating HIV. At the end of Phase 2, we will complete a high-throughput screen of several structurally diverse small molecule libraries, identify & validate hits, and select and advance lead candidates. We anticipate this work will lead to identification of novel small molecules as a basis for HIV silencing approaches focused on the CNS reservoir.

项目摘要/摘要-表观遗传HIV沉默在Macrophages 抗逆转录病毒疗法（ART）在HIV感染中非常有效，但有很大的局限性， 在没有ART的情况下达到无病毒血症状态（“功能性治愈”）是非常感兴趣的焦点。一种潜在的方法， 一旦ART停止，防止持久性病毒储库产生复制病毒的方法是使艾滋病毒沉默， 这样感染就不会“死灰复燃”。感染的巨噬细胞系细胞作为主要的长期 中枢神经系统中的HIV储存库，这被认为是一个“避难所”，即使额外的- 清除CNS储液器。此外，长寿命的感染巨噬细胞可能继续产生低水平的 病毒，这有助于艾滋病毒相关的神经认知障碍（手）。因此，沉默艾滋病毒在 巨噬细胞可能有助于“功能性治愈”，并在ART抑制的情况下改善HAND。 HIV整合到宿主细胞基因组中，并受到正和负表观遗传调控。多 已经对T细胞中HIV的表观遗传控制进行了研究，但对巨噬细胞中的HIV知之甚少。染色质 组织和转录调控是高度细胞特异性和环境特异性的。我们的科学前提是 CNS中的巨噬细胞不能表观遗传地抑制HIV并作为长期储存库。我们 假设巨噬细胞特异性增强子-启动子相互作用调节HIV持续转录 活性，并且小分子可以重新编程HIV感染的巨噬细胞的表观遗传调节， 建立整合的HIV基因组的长期沉默。为此，我们设立了一个初级 用于高通量筛选表观遗传酶的小分子调节剂的巨噬细胞模型，和 为双加氧酶参与组蛋白去甲基化提供了初步证据。 在第一阶段，我们将优化高通量筛选活动的初级和二级检测，以确定 原代人巨噬细胞中HIV-1表达的小分子表观遗传调节剂;测试的作用 候选表观遗传调节剂，特别是可以防止HIV沉默的组蛋白和DNA去甲基酶;以及 进一步研究调节巨噬细胞中HIV持续转录的分子机制。同相 2，我们将进行高通量筛选，开发小分子来沉默巨噬细胞中的HIV-1， 行业标准里程碑驱动的药物发现管道，以识别生物活性化学型;验证先导 与原代HIV分离株和骨髓细胞类型的化合物;候选命中物的药物样特性 通过药物化学;并确定分子靶点和进行作用机制研究。 在第一阶段结束时，我们将有一个强大的高通量的原代细胞为基础的筛选，正交验证 检测以确认靶向命中，并进一步深入了解调节HIV的巨噬细胞特异性表观遗传因子。 在第2阶段结束时，我们将完成几种结构不同的小分子的高通量筛选， 图书馆，识别和验证命中，并选择和推进领先的候选人。我们预计这项工作将导致 鉴定新的小分子作为HIV沉默方法的基础，重点是CNS储库。