Molecular Determinants of Cross-Reactive Antibody Response to Influenza in Humans

人类流感交叉反应抗体反应的分子决定因素

• 批准号: 8890770
• 负责人: James E Crowe
• 金额: $ 37.01万
• 依托单位: VANDERBILT UNIVERSITY
• 依托单位国家: 美国
• 财政年份: 2012
• 资助国家: 美国
• 起止时间: 2012-08-15 至 2016-04-29
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/8890770
• 关键词: Adult; Affect; Affinity; Animals; Antibodies; Antibody Formation; Antibody Response; Antigens; Area; B-Lymphocytes; Binding; Binding Sites; Biochemical; Biology; Complex; Coupled; Data; Dendritic Cells; Dengue Virus; Development; Disease; Elements; Epithelial Cells; Epitope Mapping; Epitopes; Escape Mutant; Exhibits; Generations; Genes; Genetic; Goals; Grant; Growth; HIV; Head; Human; Immune; Immunity; Immunoglobulin Somatic Hypermutation; Individual; Infection; Influenza; Influenza Hemagglutinin; Maps; Mediating; Mediator of activation protein; Molecular; Molecular Profiling; Monoclonal Antibodies; Mutagenesis; Mutation; Nature; Phylogenetic Analysis; Point Mutation; Polysaccharides; Reagent; Reporting; Scientist; Site; Somatic Mutation; Specific qualifier value; Specificity; Structure; Technology; Testing; Time; Treatment Efficacy; Vaccination; Vaccine Antigen; Vaccines; Viral; Virus; Work; alpha helix; base; cross reactivity; design; human monoclonal antibodies; human subject; immunogenic; in vivo; influenza virus vaccine; influenzavirus; interest; mutant; neutralizing monoclonal antibodies; novel; receptor; receptor binding; response; screening; sialic acid receptor; stem

Work in Project 2 will explore exciting recent developments in the study of cross-reactive immunity to influenza viruses. The goal of this project is to determine the molecular and structural basis for broadly cross-reactive human mAb responses to influenza HAs; As the epitope sequences and structures recognized by broadly cross-reactive Abs are defined, we will be better able to rationally design broadly protective immunogens. The work seeks to investigate the repertoire of human Abs recognizing three principal conserved regions of the HA molecule; a canonical stalk domain epitope, the long alpha-helix stalk domain, and the globular head domain. The hypothesis is that, unexpectedly, there are at least three immunogenic domains in influenza HA that retain highly conserved structural features and that most adult healthy subjects do possess circulating cross-reactive B cell clones to these conserved sites on HA. These domains are complex, however, and relatively inaccessible-for conventional Ab structures, we have developed robust technologies for isolating human mAbs that reveal the relatively frequent nature of cross-reactive B cells, and the genetic and structural basis for broad cross-reactivity of the secreted Abs for influenza viruses of diverse subtypes. We hypothesize that such cross-reactivity is driven by unusual features in the Ab repertoire, especially an exceptionally high level of somatic point mutations and somatic insertions. This highly interactive project will engage in collaborative work with Project 1 to determine mAb-HA co-crystal structures. Project 4 to study the biology of viral escape mutant viruses, Project 5 to examine the effect of mutants on DCs, and will use the Glycan Array core to study glycosyiation in escape sites and the Animal Core to determine in vivo potency of antibodies. Studies of such Abs and the sites recognized by them will inform the rational design of universal influenza vaccines based on the underlying principles of heterosubtypic immunity. The stalk domain is of interest because highly cross-reactive Ab have been identified that recognized a conserved region in this area. However, the rare Abs identified to date do not possess both group 1 and 2 specificities. In preliminary data we show that it is possible to isolate such Abs. Using these unusual reagents, we will define the basis for the group determinants or conserved elements: by mapping the interaction of large panels of new Abs to the: stalk region. We also will seek to isolate novel human Abs from human subjects that recognize the stalks of both major HA antigenic groups. The head domain is better studied, however there are only rare Abs that exhibit cross-reactivity for antigenically distinct HAs. By large-scale screening of B cells from subjects immunized with seasonal and experimental vaccines, we have identified cross-reactive Abs that recognize the head domain of HAs of multiple subtypes of influenza. By epitope mapping using biochemical arid biologic studies, coupled with studies to determine co-crystal structures, we will determine the structural basis for cross reactivity.

项目 2 的工作将探索流感交叉反应免疫研究中令人兴奋的最新进展 病毒。该项目的目标是确定广泛交叉反应的人单克隆抗体的分子和结构基础 对流感HA的反应；由于广泛交叉反应抗体识别的表位序列和结构是 定义后，我们将能够更好地合理设计具有广泛保护性的免疫原。这项工作旨在调查 识别 HA 分子的三个主要保守区域的人类抗体库；规范茎域 表位、长α螺旋柄结构域和球状头结构域。假设出乎意料的是，有 流感HA中至少有三个免疫原性结构域，它们保留了高度保守的结构特征，并且大多数 成年健康受试者确实拥有与 HA 上这些保守位点发生交叉反应的循环 B 细胞克隆。这些 然而，结构域很复杂，并且相对难以接近——对于传统的 Ab 结构，我们已经开发了稳健的 分离人类单克隆抗体的技术揭示了交叉反应 B 细胞相对频繁的性质，以及 不同亚型流感病毒分泌抗体广泛交叉反应的遗传和结构基础。我们 假设这种交叉反应性是由抗体库中的异常特征驱动的，特别是异常高水平的体细胞点突变和体细胞插入。这个高度互动的项目将与项目 1 合作确定 mAb-HA 共晶结构。项目 4 研究病毒逃逸突变病毒的生物学，项目 5 研究突变体对 DC 的影响，并将使用聚糖阵列核心研究逃逸位点的糖基化，并使用动物核心确定抗体的体内效力。对此类抗体及其识别位点的研究将为基于异亚型免疫基本原理的通用流感疫苗的合理设计提供信息。茎结构域之所以令人感兴趣，是因为已鉴定出高度交叉反应的抗体，该抗体识别该区域的保守区域。然而，迄今为止发现的稀有 Abs 并不同时具备第 1 组和第 2 组特异性。 在初步数据中，我们表明分离此类抗体是可能的。使用这些不寻常的试剂，我们将定义组决定因素或保守元素的基础：通过将大组新抗体的相互作用映射到： 茎区域。我们还将寻求从识别两种主要抗体茎的人类受试者中分离出新的人类抗体。 HA 抗原基团。头部结构域得到了更好的研究，但是只有极少数抗体表现出与抗原不同的 HA 的交叉反应性。通过对季节性和实验性免疫受试者的 B 细胞进行大规模筛选 疫苗中，我们已经鉴定出交叉反应抗体，可以识别多种亚型的 HA 的头域 流感。通过使用生化和生物学研究进行表位作图，并结合确定共晶的研究 结构，我们将确定交叉反应性的结构基础。