Structure/function studies of immunogen recognition by anti-HIV antibody b12

抗HIV抗体b12免疫原识别的结构/功能研究

• 批准号: 8463117
• 负责人: Roland K Strong
• 金额: $ 57.18万
• 依托单位: SEATTLE BIOMEDICAL RESEARCH INSTITUTE
• 依托单位国家: 美国
• 财政年份: 2013
• 资助国家: 美国
• 起止时间: 2013-05-01 至 2016-04-30
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/8463117
• 关键词: AIDS Vaccines; Affect; Affinity; Antibodies; Antibody Binding Sites; Antibody Formation; Antigens; Applied Research; B-Cell Activation; B-Lymphocytes; Back; Basic Science; Binding; Binding Sites; Biological Assay; Characteristics; Complement; Complex; Crystallization; Crystallography; Data; Development; Engineering; Epitopes; Event; Generations; Goals; HIV; HIV Antibodies; HIV Envelope Protein gp120; HIV Infections; HIV vaccine; Highly Active Antiretroviral Therapy; Immune response; Immune system; Immunization; Immunodominant Epitopes; Immunoglobulin Somatic Hypermutation; Immunology; In Vitro; Infection; Kinetics; Laboratories; Lead; Membrane; Molecular; Molecular Conformation; Pharmaceutical Preparations; Plasma Cells; Process; Property; Proteins; Receptor Signaling; Receptors, Antigen, B-Cell; Research; Scaffolding Protein; Structure; Structure of germinal center of lymph node; Testing; Thermodynamics; Vaccines; Viral; Viral Vaccines; Virus; Virus Diseases; Virus Receptors; Washington; antigen binding; base; design; feeding; improved; in vivo; inhibiting antibody; neutralizing antibody; neutralizing monoclonal antibodies; novel vaccines; prevent; prophylactic; protein protein interaction; response; scaffold; vaccine candidate; vaccine development

Effective AIDS vaccines will likely need to elicit antibodies that can neutralize diverse strains of HIV, but vaccine immunogens based on soluble HIV Env constructs have, so far, failed to elicit such responses. Our first-generation attempts to design 'epitope-scaffold' antigens for bNAb 4E10 generate exquisite epitope-specific responses that, unfortunately, also fail to potently neutralize HIV. In order to improve on this approach (and embrace parallel strategies), we will now focus on the broadly neutralizing, CD4-binding site antibody b12. We propose to iteratively engineer b12-specific i) epitope-scaffolds and ii) minimized gp120 constructs, with binding optimized to both mature b12 and its germline precursor; the latter recognition event is essential for selectively activating the correct naive B cells that will lead to b12-equivalent responses. The biophysical parameters describing BCR/antigen interactions (affinities, kinetics, thermodynamics) have been shown to have profound effects on the course of humoral immune responses. We will test the hypothesis that i) b12 germline candidates will not completely recapitulate the binding, structural or neutralization properties of the mature antibody by some subset of the parameters that describe protein/protein interactions and function and ii) that, in order for designed immunogens to induce effective b12-like responses upon immunization, both immunogen/b12 and immunogen/germline interactions will need to be recapitulated. Since no single b12 germline precursor can confidently be identified, we propose to study the structures (using x-ray crystallography and small-angle x-ray scattering (SAXS)) and binding properties (using SPR and ITC) of an ensemble of the 12 likeliest candidates. Our proposed comprehensive biophysical studies of mature b12 and the complete ensemble of candidate b12 germline precursor antibodies will: i) support the rational immunogen redesign cycle in Project 1; ii) detail the molecular mechanisms of b12 maturation; and iii) provide the biophysical framework for interpreting the in vitro and in vivo B cell activation assays in Project 3. Our research goal is to understand the biophysical constraint/s on B cell activation thresholds for an epitope of one of the most potent and broadly anti-HlV-1 neutralizing MAbs known. We will then use these results to guide efforts to rationally design better immunogens to elicit anti-HIV NAbs while contributing significantly to the field of fundamental B cell immunology, combining basic and applied research goals.

有效的艾滋病疫苗可能需要引起可以中和可消除各种艾滋病毒菌株的抗体，但是迄今为止，基于可溶性HIV Env构建体的疫苗免疫原子未能引起此类反应。我们的第一代试图设计为BNAB 4E10设计的“表位 - 损坏”抗原产生了精美的表位特异性反应，不幸的是，这些反应也无法在中和HIV中有效中和。为了改善这种方法（并采用并行策略），我们现在将专注于广泛中和的CD4结合网站 抗体B12。我们向迭代的工程师提出了B12特异性I）表位型和II）最小化的GP120构建体，并与成熟的B12及其生殖线前体进行了优化；后一种识别事件对于选择性地激活将导致B12等效响应的正确的幼稚B细胞至关重要。描述BCR/抗原相互作用的生物物理参数（亲和力，动力学，热力学）已被证明对体液免疫反应的过程有深远的影响。我们将检验以下假设：i）B12种系候选者不会完全概括结合，结构或中和 成熟抗体的性质通过描述蛋白质/蛋白质相互作用和功能的参数的某个子集以及II），为了使设计的免疫原子在免疫后诱导有效的B12样反应，因此需要重新审查免疫原/B12和免疫原/生殖线相互作用。由于无法自信地识别单个B12生殖线前体，因此我们建议研究结构（使用X射线晶体学和小角度X射线散射（SAXS））和12个最有可能候选者的集合的结合特性（使用SPR和ITC）。我们提出的关于 成熟的B12和候选B12生殖线前体抗体的完整合奏将：i）支持项目1中的理性免疫重新设计周期； ii）详细说明B12成熟的分子机制； iii）提供了在项目3中解释体外和体内B细胞激活测定法的生物物理框架。我们的研究目标是了解B细胞激活阈值的生物物理约束，以了解最有效，最广泛的抗HLV-1中和中和中和的MAB的表位。然后，我们将使用这些结果来指导努力，以合理地设计更好的免疫原，以引起抗HIV NAB，同时贡献抗HIV NAB 重要的是基本的B细胞免疫学领域，结合了基本和应用研究目标。