Cloning, characterization and in vivo testing of mucosally transmitted SIV

粘膜传播的 SIV 的克隆、表征和体内测试

• 批准号: 8625408
• 负责人: GEORGE M SHAW
• 金额: $ 45.12万
• 依托单位: UNIVERSITY OF PENNSYLVANIA
• 依托单位国家: 美国
• 财政年份: 2010
• 资助国家: 美国
• 起止时间: 2010-02-01 至 2016-02-28
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/8625408
• 关键词: AIDS Vaccines; AIDS/HIV problem; Address; Antibodies; Area; Award; Biological; CD4 Positive T Lymphocytes; Cell Communication; Cells; Cessation of life; Clinical; Cloning; DNA; Development; Dose; Evolution; Failure; Genome; HIV; HIV-1; Human; Immune response; In Vitro; Infection; Kinetics; Knowledge; Laboratories; Lead; Length; Life; Light; Lymphocyte Depletion; Macaca; Macaca mulatta; Methods; Modeling; Molecular; Molecular Cloning; Nature; Pathogenesis; Pathogenicity; Pathway interactions; Phase; Phased Innovation Awards; Phenotype; Plasma; Property; Reagent; Research Priority; SIV; Sequence Analysis; Sexual Transmission; T-Lymphocyte; Testing; Time; Tropism; United States National Institutes of Health; Vaccine Design; Vaccine Research; Vaccines; Viral Genome; Viremia; Virus; Virus Diseases; abstracting; base; efficacy trial; in vivo; innovation; new technology; nonhuman primate; novel; novel strategies; novel vaccines; pandemic disease; programs; transmission process; vaccine development; viral RNA

Project Summary/Abstract One of the most important knowledge gaps in HIV/SIV vaccine research relates to the molecular properties of viruses that are responsible for mucosal transmission and the initial virus-host cell interactions that lead to productive viral infection. The present proposal addresses this priority area by taking advantage of our laboratory's recent discovery of a novel strategy for identifying mucosally transmitted HIV-1 and SIV viral genomes (Keele, PNAS 2008; Keele, J Exp Med 2009). This strategy, based on single genome amplification, sequencing and analysis of plasma viral RNA within the context of a model of random virus evolution, identifies those viruses that are actually responsible for transmission and productive infection. This innovation, in turn, makes possible for the first time the molecular cloning and biological analysis of actual transmitted/founder SIV viruses. In this application, we propose to extend this new approach to the SIV-macaque infection model and to test the following hypothesis: Molecular clones of full-length SIVsmE660 and SIVmac251 viral genomes corresponding to transmitted/founder viruses can be identified, will be shown to be infectious and replication competent, and will recapitulate pathogenic infection in Indian rhesus macaques. Such clones represent novel molecular reagents with which to decipher the earliest virus-host cell interactions responsible for mucosal SIV transmission and can provide new molecularly-defined virus challenge strains for transmission, pathogenesis and vaccine research. Specific aims of the project are organized into R21 (Aims #1-3) and R33 (Aims #4-5) phases amenable to a go/no-go decision. Aims are: 1) To identify transmitted/founder SIVsmE660 and SIVmac251 viruses responsible for establishing productive clinical infection following low-dose mucosal (ir and ivag) inoculation; 2) To molecularly clone full-length transmitted/founder SIVsmE660 and SIVmac251 proviral genomes; 3) To biologically characterize molecular clone-derived SIVsmE660 and SIVmac251 virus strains compared with SIVsmE660 and SIVmac251 virus isolates in vitro with respect to replication efficiency, cell tropism, and envelope phenotype; 4) To determine infectivity, replication kinetics, and pathogenicity of cloned SIVsmE660 and SIVmac251 virus strains in Indian rhesus macaques following low-dose mucosal inoculation; 5) To characterize molecular pathways of virus diversification and adaptation between transmission and the establishment of set-point viremia as a prelude to homologous and heterologous vaccine-challenge studies. Results from these studies promise to shed new light on the molecular basis of mucosal transmission by SIVsmE660 and SIVmac251, identify new potential targets for protective vaccine-elicited immune responses, and provide much needed molecular clones of genetically-diverse, mucosally-transmitted SIV strains for transmission, pathogenesis and vaccine research.

项目摘要/摘要 HIV/SIV疫苗研究中最重要的知识空白之一涉及到 负责粘膜传播和最初的病毒-宿主细胞相互作用的病毒，导致 有效的病毒感染。本提案通过利用我们的 实验室最近发现了一种识别经粘膜传播的HIV-1和SIV病毒的新策略 基因组(Keele，PNAS 2008；Keele，J Exp Med，2009)。这一策略基于单基因组扩增， 在随机病毒进化模型的背景下对血浆病毒RNA进行测序和分析，确定 那些真正负责传播和生产性感染的病毒。这项创新，反过来， 首次使实际传播/方正SIV的分子克隆和生物学分析成为可能 病毒。在本应用中，我们建议将这一新方法扩展到SIV-猕猴感染模型，并 检验以下假设：SIVsmE660和SIVmac251病毒基因组全长的分子克隆 相应的传播/方正病毒可以被识别，将被显示为具有传染性和复制性 并将概述印度恒河猴的致病感染情况。这样的克隆代表了新奇的东西 用于破译引起粘膜SIV的早期病毒-宿主细胞相互作用的分子试剂 并可为传播、发病机制提供新的分子定义的病毒挑战株 和疫苗研究。项目的具体目标分为R21(目标#1-3)和R33(目标#4-5) 阶段服从于通过/不通过的决定。目标是：1)识别被传播的/创始人SIVsmE660和 SIVmac251病毒负责建立低剂量粘膜(IR和 IVAG)接种；2)分子克隆全长传播型/方正SIVsmE660和SIVmac251前病毒 基因组；3)分子克隆衍生的SIVsmE660和SIVmac251病毒株的生物学特性 与SIVsmE660和SIVmac251病毒分离株的体外复制效率比较，细胞 嗜性和包膜表型；4)确定克隆的感染性、复制动力学和致病性 低剂量粘膜接种后印度猕猴体内的SIVsmE660和SIVmac251病毒株； 5)描述病毒多样化的分子途径以及传播和适应之间的关系 建立设定点病毒血症作为同源和异源疫苗挑战研究的前奏。 这些研究的结果有望为粘膜传播的分子基础提供新的线索。 SIVsmE660和SIVmac251，为疫苗引发的保护性免疫反应识别新的潜在靶点，以及 提供急需的遗传多样性、经粘膜传播的SIV毒株的分子克隆 传播、发病机制和疫苗研究。