Virus and Antibody Gene Sequencing Core

病毒和抗体基因测序核心

• 批准号: 10117168
• 负责人: Beatrice H Hahn
• 金额: $ 39.84万
• 依托单位: UNIVERSITY OF PENNSYLVANIA
• 依托单位国家: 美国
• 财政年份: 2017
• 资助国家: 美国
• 起止时间: 2017-03-07 至 2022-05-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/10117168
• 关键词: AIDS vaccine development; Address; Antibodies; Antibody Response; Antibody Specificity; Antigens; Appearance; Autologous; B-Cell Antigen Receptor; B-Lymphocytes; Base Sequence; Binding; Bioinformatics; Blood; CMV promoter; Cell Culture Techniques; Collaborations; DNA Sequencing Facility; Data; Data Set; Development; Dideoxy Chain Termination DNA Sequencing; Epitopes; Escape Mutant; Evolution; Family; Genes; Genetic; Goals; HIV; HIV-1; Human; Humoral Immunities; Immune; Immunization; Immunize; Immunoglobulin Variable Region; Individual; Infection; Lead; Ligation; Light; Macaca; Macaca mulatta; Maps; Mediating; Memory B-Lymphocyte; Methods; Monitor; Monoclonal Antibodies; Mutation; Pathway interactions; Pattern; Phylogenetic Analysis; Plasma; Polysaccharides; Process; Production; Race; Research Personnel; Rhesus; Satellite Viruses; Sequence Alignment; Sequence Analysis; Services; Testing; Time; Transcript; Vaccine Design; Variant; Viral; Virus; Virus Diseases; arm; cross reactivity; genome sequencing; member; neutralizing antibody; next generation sequencing; pressure; response; simian human immunodeficiency virus; statistics; vaccine development

PROJECT SUMMARY The development of broadly cross-reactive neutralizing antibodies (bNAbs) in HIV-1 infected individuals requires iterative rounds of envelope (Env)-mediated B cell receptor stimulation, strain specific (autologous) neutralizing antibody (NAb) elicitation, and associated virus escape. In this virus/antibody “arms race”, particular Env escape mutants are generated which, among the myriads of other Env variants present in infected individuals, are able to select for antibody lineages that acquire neutralization breadth. Although the principle of virus/antibody co-evolution has been established, the number, identity, succession, and cooperation of the particular Env variants that initiate and sustain bNAb lineage maturation have not been determined. It is also unclear to what extent the pathways of bNAb induction observed in one individual can be generalized to others. To address these questions, this HIVRAD team will test the scientific premise that simian-human immunodeficiency viruses (SHIVs) bearing HIV-1 Envs that engage V1V2 and V3-glycan bNAb unmutated common ancestor (UCA) and intermediate ancestor (IA) B cell receptors (BCR) in humans will bind related BCRs in rhesus macaques, leading to a recapitulation of virus/antibody coevolution and the development of NAb breadth and potency. We have already overcome a critical hurdle toward these objectives by generating primary and transmitted founder (TF) Env containing SHIVs, which replicate to high titers in rhesus macaques and yield patterns of early env evolution and autologous NAb escape that are astonishingly similar to those of humans infected with HIV-1 strains encoding the same Envs. These data suggest that B cell responses to a common Env antigen may, at least for some Envs, be deterministic and that by comparing the patterns of virus/antibody coevolution in outbred macaques and humans infected with the same Env expressing virus, it will be possible to identify common pathways of virus/BCR engagement and bNAb development that can be applied to vaccine design. The Virus and Antibody Gene Sequencing Core (Core B) will support this project by providing state-of-the-art sequencing services that will allow this HIVRAD consortium to dissect the mechanisms of SHIV elicited protective humoral immunity. Working closely with the Bioinformatics and Statistics Core (Core C) as well as Projects #1, #2 and #3, we will (i) characterize the evolving env quasispecies in SHIV infected macaques over time, and (ii) use Sanger and nextgen sequencing (NGS) methods to sequence antibody heavy (VHDJH) and light (VLJL) chain variable regions from clonal B cell cultures and rhesus memory B cells before and after SHIV infection and/or immunization. The goal is to understand the dynamic interactions between the evolving Env quasispecies and members of NAb and bNAb lineages to identify key Env escape mutations that reproducibly initiate and drive bNAb maturation.

项目概要 HIV-1 感染者体内广泛交叉反应的中和抗体 (bNAb) 的发展 需要迭代轮次包膜 (Env) 介导的 B 细胞受体刺激，菌株特异性（自体） 中和抗体（NAb）引发，以及相关的病毒逃逸。在这场病毒/抗体“军备竞赛”中， 产生了特定的 Env 逃逸突变体，其中存在于无数其他 Env 变体中 感染个体能够选择获得中和广度的抗体谱系。虽然 病毒/抗体共同进化的原理已经建立，数量、身份、演替、 启动和维持 bNAb 谱系成熟的特定 Env 变体之间的合作尚未被证实 决定。目前还不清楚在一个个体中观察到的 bNAb 诱导途径可以在多大程度上发挥作用。 推广到其他人。为了解决这些问题，HIVRAD 团队将测试以下科学前提： 携带 HIV-1 包膜的猿猴人类免疫缺陷病毒 (SHIV) 与 V1V2 和 V3-聚糖 bNAb 结合 人类未突变的共同祖先 (UCA) 和中间祖先 (IA) B 细胞受体 (BCR) 会结合 恒河猴中相关的 BCR，导致病毒/抗体共同进化的重现和 NAb 广度和效力的发展。我们已经克服了实现这些目标的关键障碍 通过生成包含 SHIV 的初级和传播创始人 (TF) 环境来实现目标，这些环境会复制 恒河猴的高滴度以及早期 env 进化和自体 NAb 逃逸的产量模式 这与感染编码相同 Env 的 HIV-1 病毒株的人类惊人地相似。 这些数据表明，B 细胞对共同 Env 抗原的反应可能（至少对于某些 Env 而言）是 确定性的，并通过比较近交系猕猴和抗体的病毒/抗体共同进化模式 如果人类感染相同的 Env 表达病毒，将有可能识别出感染相同 Env 病毒的共同途径 病毒/BCR 结合和 bNAb 开发可应用于疫苗设计。病毒和 抗体基因测序核心（核心 B）将通过提供最先进的测序来支持该项目 这些服务将使 HIVRAD 联盟能够剖析 SHIV 引发保护性体液的机制 免疫。与生物信息学和统计核心（核心 C）以及项目#1、#2 和 #3，我们将 (i) 描述感染 SHIV 的猕猴中不断进化的环境准种，以及 (ii) 使用 Sanger 和下一代测序 (NGS) 方法对抗体重链 (VHDJH) 和轻链 (VLJL) 进行测序 SHIV 感染前后克隆 B 细胞培养物和恒河猴记忆 B 细胞的可变区和/或 免疫接种。目标是了解不断进化的 Env 准种和 NAb 和 bNAb 谱系的成员，以识别可重复启动和驱动的关键 Env 逃逸突变 bNAb 成熟。