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Structural basis for activity of and resistance to HIV integrase inhibitors

HIV整合酶抑制剂的活性和耐药性的结构基础

基本信息

批准号：
10238819
负责人：
Dmitry Lyumkis
金额：
$ 66.85万
依托单位：
SALK INSTITUTE FOR BIOLOGICAL STUDIES
依托单位国家：
美国
项目类别：
财政年份：
2017
资助国家：
美国
起止时间：
2017-09-25 至 2022-08-31
项目状态：
已结题

来源：
https://reporter.nih.gov/project-details/10238819
关键词：
Active Sites Address Adopted Affect Binding Biochemical Biological Biological Assay Biology Cells Chromatin Chromosomes Clinic Clinical Collaborations Communities Complement Complex Coupled Cryoelectron Microscopy DNA Data Development Disease Progression Drug Industry Drug Targeting Drug resistance Enzymes Future Generations Genes Guidelines HIV HIV Integrase HIV Integrase Inhibitors HIV-1 Histone H3 Hybrids Individual Infection Institutes Integrase Investigation Macromolecular Complexes Mediating Molecular Mutation Nucleoproteins Ohio Oligonucleotides Pathway interactions Patients Pattern Pharmaceutical Preparations Positioning Attribute Procedures Process Research Resistance Resistance development Resistance profile Resolution Role Scientist Spumavirus Structure Tail Techniques Translating United States National Institutes of Health Universities Variant Viral Virus Virus Integration Work antiretroviral therapy base clinically relevant design drug action drug development effective therapy experimental study improved inhibitor/antagonist insight interest lens epithelium-derived growth factor mutant novel particle profiles in patients prototype public health relevance response structural biology success tool transcriptional coactivator p75 viral DNA viral resistance virology

项目摘要

Abstract The Human Immunodeficiency Virus Type 1 (HIV-1, hereafter referred to as HIV) currently infects ~37 million people worldwide, and the number of infected individuals continues to rise. In the absence of a cure, antiretroviral therapy (ART) represents the primary treatment option, as it slows disease progression, and limits new infections. Integrase (IN) Strand Transfer Inhibitors (INSTIs) are a class of ART that block integration of viral DNA into host chromosomes, a process that is mediated by the viral IN enzyme, which assembles into oligomeric nucleoprotein complexes on the ends of viral DNA, termed “intasomes”. INSTIs selectively target intasomes and represent first-line therapies in the clinic. However, the emergence of IN variants resistant to INSTIs is a major clinical problem. Structural biology approaches can decipher the molecular mechanisms underlying drug action and resistance, providing useful information for rationally improving current therapies. In this proposal, approaches centered around revolutionary advances in cryo-electron microscopy for structural studies will be used to understand how INSTIs interact with their natural drug target, the HIV intasome, as well as mechanisms by which resistance to these drugs emerges. The proposed aims will address several major themes. Aim 1 will define the mechanisms of action of clinical and developmental INSTIs in the context of HIV intasomes. Procedures used to perform high-resolution structural studies of INSTI-bound complexes by cryo-EM will then be adopted to decipher novel, clinically relevant mechanisms of drug resistance that arise in response to INSTI treatment. This work will be complemented using biochemical and virology assays designed to dissect key interactions between HIV intasomes and INSTIs and to validate structural findings. Aim 2 will extend these findings to define INSTI mechanism of action in the presence of biologically relevant cellular factors, including methylated mononucleosomes (mMNs; the natural target for HIV integration) and lens epithelium-derived growth factor (LEDGF). Aim 2 will therefore define the biochemical and structural mechanisms by which INSTIs interact with and inhibit IN catalytic activity in the context of the intasome-LEDGF-mMN complex, thereby elucidating how INSTIs function in infected cells and their precise stage of activity. In addition to providing the first structural information for INSTI interactions with their natural drug target in the presence of relevant cellular factors, this work will: 1) elucidate how mutations within the IN active site disrupt drug binding, 2) define the precise stage and mechanism of action of this important class of drugs in a cellular context, and 3) provide blueprints for the rational improvement of future INSTIs. !

摘要人类免疫缺陷病毒1型（HIV-1，以下简称HIV）目前感染约3700万人，世界各地的人，受感染的人数继续上升。在没有治愈方法的情况下，抗逆转录病毒疗法（ART）代表了主要的治疗选择，因为它减缓了疾病的进展，并限制了新的感染。整合酶链转移抑制剂（Integrin ase Strand Transfer Inhibitors，INSTIs）是一类阻断病毒DNA整合到宿主细胞中的抗病毒药物染色体，这是一个由病毒IN酶介导的过程，它组装成寡聚核蛋白病毒DNA末端的复合物，称为“整合体”。INSTI选择性靶向整合体，临床一线治疗。然而，对INSTI耐药的IN变体的出现是INSTI的主要临床表现。问题.结构生物学方法可以破译药物作用的分子机制，耐药性，为合理改进目前的治疗方法提供了有用的信息。在这一建议中，方法围绕结构研究的冷冻电子显微镜的革命性进展，将用于了解INSTI如何与其天然药物靶点HIV整合体相互作用，以及对这些药物产生了抗药性。拟议的目标将涉及几个主要主题。目的1将定义临床的作用机制，和发育性INSTI在HIV融合体的背景下。用于执行高分辨率的程序通过冷冻电镜对INSTI结合的复合物进行结构研究，在响应于RNAi治疗中出现的耐药性的相关机制。这项工作将使用生物化学和病毒学分析来补充，旨在剖析艾滋病毒之间的关键相互作用，整合体和INSTI，并验证结构发现。目标2将扩展这些发现，以定义在生物相关细胞因子存在下的作用机制，包括甲基化单核小体（mMN; HIV整合的天然靶点）和透镜上皮衍生生长因子（LEDGF）。因此，目标2将定义INSTI与并在整合体-LEDGF-mMN复合物的情况下抑制IN催化活性，从而阐明如何 INSTI在受感染的细胞中发挥作用，并具有精确的活性阶段。除了提供第一个结构信息，用于与天然药物靶点相互作用，相关细胞因子的存在，这项工作将：1）阐明IN活性位点内的突变如何破坏药物结合，2）定义这类重要药物在细胞内的确切作用阶段和作用机制， 3）为未来INSTI的合理改进提供蓝图。 !