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: 8327358
• 负责人: GEORGE M SHAW
• 金额: $ 10.49万
• 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 2013-01-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/8327358
• 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; base; efficacy trial; in vivo; innovation; new technology; nonhuman primate; novel; novel strategies; novel vaccines; pandemic disease; programs; public health relevance; transmission process; vaccine development; viral RNA

DESCRIPTION (provided by applicant): 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. PUBLIC HEALTH RELEVANCE: One of the most important knowledge gaps in HIV/AIDS vaccine research relates to the molecular properties of HIV-1 that are responsible for sexual transmission and the initial virus-host cell interactions that lead to productive viral infection. The present proposal takes advantage of our laboratory's recent discovery of a novel method for identifying transmitted HIV-1 viruses and applies this new technology to the discovery and characterization of transmitted simian immunodeficiency viruses (SIV). These results will shed new light on the molecular basis of mucosal transmission by SIV in the rhesus macaque model and promise to aide in the identification of new vaccine targets and strategies.

描述(由申请人提供)：HIV/SIV疫苗研究中最重要的知识空白之一与负责粘膜传播的病毒的分子特性以及导致生产性病毒感染的初始病毒-宿主细胞相互作用有关。本提案利用我们实验室最近发现的一种新策略来识别经粘膜传播的HIV-1和SIV病毒基因组，从而解决了这一优先领域(Keele，PNAS，2008；Keele，J Exp Med，2009)。这一策略基于随机病毒进化模型中的血浆病毒RNA的单基因组扩增、测序和分析，确定了那些实际上负责传播和生产性感染的病毒。这项创新反过来又首次使实际传播/创始SIV病毒的分子克隆和生物学分析成为可能。在这一应用中，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：可以识别与传播/创始病毒相对应的全长SIVsmE660和SIVmac251病毒基因组的分子克隆，将被证明具有感染性和复制能力，并将概括印度猕猴的致病感染。这些克隆代表了新的分子试剂，可以用来破译导致粘膜SIV传播的最早的病毒-宿主细胞相互作用，并可以为传播、发病机制和疫苗研究提供新的分子定义的病毒挑战株。项目的具体目标被组织为R21(目标1-3)和R33(目标4-5)阶段，可根据通过/不通过决定。目的是：1)鉴定在低剂量黏膜(ir和ivag)接种后导致生产性临床感染的传播/创始人SIVsmE660和SIVmac251病毒；2)分子克隆全长传播/创始人SIVsmE660和SIVmac251前病毒基因组；3)比较分子克隆衍生的SIVsmE660和SIVmac251病毒分离株与SIVsmE660和SIVmac251病毒分离株的体外复制效率、细胞嗜性和包膜表型；4)确定克隆的SIVsmE660和SIVmac251病毒在低剂量黏膜接种后在印度猕猴体内的感染性、复制动力学和致病性；5)作为同源和异源疫苗挑战研究的前奏，表征病毒在传播和建立设定点病毒血症之间的多样性和适应性的分子途径。这些研究结果有望揭示SIVsmE660和SIVmac251黏膜传播的分子基础，识别新的保护性疫苗诱导免疫反应的潜在靶点，并为传播、发病机制和疫苗研究提供急需的遗传多样性、粘膜传播的SIV毒株的分子克隆。 公共卫生相关性：艾滋病毒/艾滋病疫苗研究中最重要的知识空白之一与艾滋病毒-1的分子特性有关，艾滋病毒-1的分子特性导致性传播，以及导致生产性病毒感染的最初病毒-宿主细胞相互作用。本建议利用我们实验室最近发现的一种识别传播的HIV-1病毒的新方法，并将这一新技术应用于传播的猿猴免疫缺陷病毒(SIV)的发现和表征。这些结果将为SIV在恒河猴模型中粘膜传播的分子基础提供新的线索，并有望帮助识别新的疫苗靶点和策略。