Cooperation of BRD4 and TAT Associated Proteins in HIV Transcription and Latency

BRD4 和 TAT 相关蛋白在 HIV 转录和潜伏期中的合作

• 批准号: 9319617
• 负责人: Jian Zhu
• 金额: $ 25.13万
• 依托单位: UNIVERSITY OF ROCHESTER
• 依托单位国家: 美国
• 财政年份: 2014
• 资助国家: 美国
• 起止时间: 2014-09-20 至 2018-03-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/9319617
• 关键词: AIDS/HIV problem; Acquired Immunodeficiency Syndrome; Binding; Biological Assay; Bromodomain; CD4 Positive T Lymphocytes; CD8-Positive T-Lymphocytes; CD8B1 gene; Cell Death; Cell model; Cells; Co-Immunoprecipitations; Combined Modality Therapy; Computer Analysis; Data; Data Set; Disease; Elements; Evaluation; Foundations; Gene Silencing; Gene Targeting; Generations; Genes; Genetic; Genetic Transcription; Goals; HIV; HIV Infections; HIV tat Protein; Highly Active Antiretroviral Therapy; Human immunodeficiency virus test; In Vitro; Infection; Integration Host Factors; Investigation; Kinetics; Latent Virus; Lead; Measures; Mediating; Patients; Peripheral Blood Mononuclear Cell; Persons; Pharmaceutical Preparations; Phase; Play; Positive Transcriptional Elongation Factor B; Process; Proteins; Proteomics; Proviruses; Quantitative Structure-Activity Relationship; RNA Interference; RNA interference screen; Recruitment Activity; Regimen; Regulation; Reporter; Reporting; Rest; Role; Safety; T-Lymphocyte; Testing; Thermodynamics; Toxic effect; Transcription Elongation; Viral; Viral load measurement; Virus; Virus Latency; Virus Replication; antiretroviral therapy; base; chromatin immunoprecipitation; computerized tools; cooperative study; efficacy study; experimental study; genome-wide; high throughput screening; improved; in vivo; inhibitor/antagonist; novel; novel therapeutic intervention; preclinical efficacy; promoter; protein complex; public health relevance; reactivation from latency; screening; small molecule; small molecule inhibitor; small molecule libraries; small molecule therapeutics; tat Protein

 DESCRIPTION: Although highly active antiretroviral therapy (HAART) is successful to block active replication of HIV in AIDS patients, it does not eradicate viruses. Presence of latent HIV reservoirs remains a major obstacle to the cure. Reactivated viral replication from latency induces cytopathic effects and leads to cell death of reservoir cells. There are several promising small compounds being currently tested for HIV reactivation from latency. However, they are either less potent in vivo or associate with severe toxicity. Recently, it was uncovered that the size of latent HIV reservoirs is much larger than previously estimated, suggesting that a combinatory regimen targeting multiple restrictive mechanisms of HIV latency may be required. Thus, identification of novel gene targets for anti-latency therapy is urgent. Our recent studies o host factors modulating HIV replication identified the bromodomain protein, BRD4, plays an important role in maintaining HIV latency. More importantly, we found that the BRD4 inhibitor, JQ1, is able to synergize with other HIV latency reversers to activate latent HIV in certain cases. These preliminary data suggest that BRD4 is a promising gene target for HIV anti-latency therapy, but the ideal gene to be targeted simultaneously with BRD4 for reaching a better effect remains to be identified. Our recent studies, using an integrated approach to combine proteomic study of host proteins physically associating with HIV TAT protein and genetic results of host proteins regulating HIV replication using RNAi-mediated gene silencing, identified a set of HIV TAT-associated inhibitory proteins (TIPs) that negatively regulate HIV transcription through interacting TAT. In this proposal, we will first verify the role of a small set of TIPs in HIV transcription and latency in primary CD4+ T cells. We will further study their cooperative interactions with BRD4. Genetic interactions of TIPs and BRD4 as well as the physical interactions of TIPs with TAT-LTR elements at HIV latency and reactivation are measured. These data are subjected to computational analyses of this small host network including TAT-LTR, BRD4-P-TEFb, and TIPs. We aim to identify the kinetically and thermodynamically significant TIPs in this network that can be targeted by small compounds. An in vitro TIP-TAT interaction assay will be developed for high-throughput small compound screens to identify inhibitors that potently disrupt such interaction. These compounds may release the suppressive effect of TIP on TAT activity to allow full activation of HIV transcription, and thus are useful to revert HIV latency. We will evaluate them in both primary cell model of HIV latency as well as the CD8+ T cells depleted peripheral blood mononuclear cells (PBMCs) isolated from HAART-treated AIDS patients. Beneficial effect of these compounds together with BRD4 inhibitor JQ1 will be determined as well to identify better drug pair(s) for HIV anti-latency therapy using JQ1. We believe that our study of novel TIPs will improve our understanding of host machineries controlling HIV transcription and latency, and further target those druggable ones for reverting HIV latency and elimination.

 产品说明：尽管高效抗逆转录病毒疗法（HAART）成功地阻断了艾滋病患者体内HIV的活跃复制，但它并不能根除病毒。潜伏的HIV宿主的存在仍然是治愈的主要障碍。从潜伏期重新激活的病毒复制诱导细胞病变效应并导致储库细胞的细胞死亡。目前有几种有希望的小化合物正在测试HIV从潜伏期重新激活。然而，它们要么在体内效力较低，要么与严重毒性相关。最近，它被发现，潜伏的HIV水库的大小比以前估计的要大得多，这表明可能需要一种针对HIV潜伏期的多种限制性机制的组合方案。因此，鉴定新的基因靶点用于抗潜伏期治疗是迫切的。我们最近对调节HIV复制的宿主因素的研究确定了溴结构域蛋白BRD 4在维持HIV潜伏期中起重要作用。更重要的是，我们发现BRD 4抑制剂JQ 1能够与其他HIV潜伏逆转剂协同作用，在某些情况下激活潜伏的HIV。 这些初步数据表明，BRD 4是HIV抗潜伏期治疗的一个有前途的基因靶点，但与BRD 4同时靶向以达到更好效果的理想基因仍有待确定。我们最近的研究，使用一个综合的方法，结合联合收割机的蛋白质组学研究的主机蛋白质与HIV达特蛋白的物理相关和主机蛋白质的遗传结果调节HIV复制使用RNAi介导的基因沉默，确定了一组HIV TAT相关抑制蛋白（TIP），负调控HIV转录通过相互作用达特。在这项提案中，我们将首先验证一小部分TIPs在HIV转录和原代CD 4 + T细胞潜伏期中的作用。我们将进一步研究它们与BRD 4的合作互动。TIPs和BRD 4的遗传相互作用以及TIPs与TAT-LTR元件在HIV潜伏期和再活化时的物理相互作用被测量。这些数据进行计算分析，这个小的主机网络，包括TAT-LTR，BRD 4-P-TEFb，和TIP。我们的目标是确定在这个网络中的动力学和药物学显着的TIP，可以通过小化合物的目标。将开发一种体外TIP-TAT相互作用试验，用于高通量小化合物筛选，以鉴定有效破坏这种相互作用的抑制剂。这些化合物可以释放TIP对达特活性的抑制作用，以允许HIV转录的完全激活，因此可用于治疗HIV感染。 逆转艾滋病毒潜伏期我们将在HIV潜伏期的原代细胞模型以及从HAART治疗的AIDS患者中分离的CD 8 + T细胞耗尽的外周血单核细胞（PBMC）中评估它们。还将确定这些化合物与BRD 4抑制剂JQ 1一起的有益效果，以确定使用JQ 1进行HIV抗潜伏期治疗的更好药物对。我们相信，我们对新型TIPs的研究将提高我们对控制HIV转录和潜伏期的宿主机制的理解，并进一步靶向那些可药物化的TIPs，以逆转HIV的潜伏期和消除。