Acylhydrazone inhibitors of HIV-1 ribonuclease H

HIV-1 核糖核酸酶 H 的酰腙抑制剂

• 批准号: 7620292
• 负责人: RIEKO ISHIMA
• 金额: $ 37.43万
• 依托单位: UNIVERSITY OF PITTSBURGH AT PITTSBURGH
• 依托单位国家: 美国
• 财政年份: 2009
• 资助国家: 美国
• 起止时间: 2009-06-01 至 2011-05-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/7620292
• 关键词: Anti-HIV Agents; Anti-Infective Agents; Anti-Retroviral Agents; Binding; Binding Sites; Biochemical; CCR5 gene; Cells; Complex; DNA; DNA-Directed DNA Polymerase; Data; Databases; Development; Drug Industry; Drug usage; Enzymes; Escherichia coli; Essential Drugs; Genomics; Goals; HIV; HIV Infections; HIV Integrase Inhibitors; HIV drug resistance; HIV-1; HIV-1 Reverse Transcriptase; Hydrazones; Individual; Libraries; Microscopic; Molecular Conformation; Multi-Drug Resistance; Mutation; Patients; Peptide Hydrolases; Pharmaceutical Preparations; Phenotype; Polymerase; Prevalence; Program Development; Property; RNA; RNA-Directed DNA Polymerase; Research; Resistance; Resistance development; Resistance profile; Ribonuclease H; Screening procedure; Site; Structure; Therapeutic; Validation; Variant; Viral; Virus; analog; base; design; drug discovery; drug resistant virus; enzyme activity; improved; inhibitor/antagonist; interest; member; novel; pathogen; prevent; public health relevance

DESCRIPTION (provided by applicant): The increasing prevalence of drug resistant HIV compromises the continued utility of current HIV drugs. Virtually all clinically used drugs are directed at only two HIV targets, protease and reverse transcriptase (RT), thus one approach to the problem of HIV drug resistance is to develop therapeutics directed at novel HIV targets, since therapeutics directed at novel HIV targets will almost certainly be active against current drug- resistant virus strains. One such target is RT-associated ribonuclease H (RNaseH), an essential enzyme activity for HIV replication and the only HIV enzyme not yet targeted by any clinically used or pipeline therapeutics. We have identified acylhydrazones as interesting compounds in that certain analogs are potent inhibitors of both HIV-1 RT DNA polymerase and RT-RNaseH activities. We hypothesize that this bifunctional inhibition is due to binding of the inhibitor to two distinct sites on RT, one of which is in or near the RNaseH domain. We recently obtained a crystal structure of acylhydrazones in the RT polymerase (pol) domain, so detailed understanding of the interaction of acylhydrazones with the RT RNaseH domain will be invaluable for design of both new RNaseH inhibitors (RNaseHI) and new bifunctional inhibitors. To this end, we propose four Specific Aims, (1) to determine the structure of HIV-RT RNaseH in complex with acylhydrazone inhibitors. Detailed NMR studies of the interaction of acylhydrazones with an active isolated HIV-1 RT RNaseH domain will provide precise structural information of the inhibitor binding pocket in the RNaseH domain, the microscopic conformation of RNaseH-inhibitor interaction sites, and influence of inhibitor binding on RT-RNaseH structure; (2) to validate the RNaseHI binding pocket determined in Aim 1. The RNaseHI binding pocket determined in Aim 1 will be validated in the context of intact RT and HIV-1 virus by evaluating the effect of mutation of selected residues interacting with RNaseHI, in the RNaseH fragment (biochemical and NMR structural analyses), in intact RT, and in HIV-1; (3) to optimize the inhibitory potency of acylhydrazone and analog inhibitors. Several approaches will be used to identify and develop potent monofunctional (RNaseH-specific) and bifunctional inhibitors, including screening of a library of 5000 hydrazone derivatives and synthesis of new analogs based on structures of acylhydrazones in the RNaseH and pol site binding pockets; (4) to conduct detailed biochemical and virologic characterizations of new RNaseHI. The inhibitory properties of potent RNaseHI and bifunctional acylhydrazones from Aim 3 will be characterized with purified RT and in cell-based HIV replication studies (including resistance development). We hypothesize that bifunctional inhibitors may be preferable in the context of resistance development. Selected RNaseHI will be further evaluated in NMR structural studies to better define the binding pocket for RNaseHI. This structural information along with the crystal data will provide a firm basis for rational design of new inhibitors. PUBLIC HEALTH RELEVANCE: There are more than 20 drugs for the treatment of HIV infection, yet the virus continues to spread in the US and worldwide. Furthermore, drug resistant HIV is increasing, limiting treatment options for individuals infected with these strains. New drugs are essential. This project will explore compounds directed at a novel HIV target, ribonuclease H, with the goal of developing new drugs that will be active against existing drug resistant HIV.

描述（由申请人提供）：抗药性HIV的患病率的增加损害了当前HIV药物的持续效用。几乎所有临床使用的药物仅针对两个HIV靶标，蛋白酶和逆转录酶（RT），因此，解决HIV耐药性问题的一种方法是开发针对新型HIV靶标的治疗剂，因为针对新型HIV靶标的治疗剂几乎可以肯定会肯定会对当前的药物抗药性病毒菌株有效。这样的靶标是与RT相关的核糖核酸酶H（RNASEH），这是HIV复制的必不可少的酶活性，也是任何临床使用或管道疗法尚未针对的HIV酶。我们已经确定酰基氢气是有趣的化合物，因为某些类似物是HIV-1 RT DNA聚合酶和RT-RNASEH活性的有效抑制剂。我们假设这种双功能抑制是由于抑制剂与RT上两个不同位点的结合，其中一个位于RNASEH结构域中或附近。我们最近在RT聚合酶（POL）结构域中获得了酰基氢唑的晶体结构，因此对于设计新的RNaseh抑制剂（RNASEHI）和新的双功能抑制剂的设计，对酰基羟基酮与RT RNASEH结构域相互作用的详细了解将是无价的。为此，我们提出了四个特定目标，（1）确定与酰基氢氮酮抑制剂复合物中HIV-RT RNASEH的结构。详细的NMR研究酰基氢唑与活性分离的HIV-1 RT RNASEH结构域的相互作用将提供RNASEH结构域中抑制剂结合袋的精确结构信息，RNASEH抑制剂相互作用位点的显微镜构象，以及抑制剂对RT-RNaseH结构的影响； (2) to validate the RNaseHI binding pocket determined in Aim 1. The RNaseHI binding pocket determined in Aim 1 will be validated in the context of intact RT and HIV-1 virus by evaluating the effect of mutation of selected residues interacting with RNaseHI, in the RNaseH fragment (biochemical and NMR structural analyses), in intact RT, and in HIV-1; （3）优化酰基氢氮酮和类似物抑制剂的抑制效力。几种方法将用于识别和开发有效的单功能（RNASEH特异性）和双功能抑制剂，包括筛选基于RNAseH和POL位点结合袋的酰基氢氮酮结构的5000个悬挂式衍生物的库以及新的类似物的合成； （4）进行新RNasehi的详细生化和病毒学特征。 AIM 3的有效RNasehi和双功能酰基氢化唑的抑制特性将以纯化的RT和基于细胞的HIV复制研究（包括抗性发展）来表征。我们假设在抵抗发展的背景下，双功能抑制剂可能是可取的。在NMR结构研究中将进一步评估选定的RNasehi，以更好地定义RNasehi的结合口袋。这些结构信息以及晶体数据将为新抑制剂的合理设计提供牢固的基础。公共卫生相关性：有20多种用于治疗艾滋病毒感染的药物，但该病毒在美国和全球范围内仍在传播。此外，抗药性HIV正在增加，限制了感染这些菌株的个体的治疗选择。新药至关重要。该项目将探索针对一种新型HIV靶标核糖核酸酶H的化合物，其目的是开发新药，这些药物将活跃于现有的抗药性HIV中。