PROJECT 2:Structural studies of SOSIP trimers

项目2：SOSIP三聚体的结构研究

• 批准号: 10083182
• 负责人: IAN A WILSON
• 金额: $ 121.73万
• 依托单位: WEILL MEDICAL COLL OF CORNELL UNIV
• 依托单位国家: 美国
• 财政年份: 2015
• 资助国家: 美国
• 起止时间: 2015-06-01 至 2025-05-31
• 项目状态: 未结题
• 来源: https://reporter.nih.gov/project-details/10083182
• 关键词: Adjuvant; Affinity; Animals; Antibodies; Antibody Response; Antigen Presentation; Antigens; Autologous; B-Cell Antigen Receptor; B-Lymphocytes; Binding; Biochemical; Biological Assay; Biophysics; Calorimetry; Chemicals; Cleaved cell; Collaborations; Complex; Coronavirus; Cryoelectron Microscopy; Data; Differential Scanning Calorimetry; Ebola virus; Electron Microscopy; Engineering; Ensure; Epitopes; Exercise; Feedback; Ferritin; Formulation; Foundations; Funding; Generations; Glycoproteins; Goals; HIV-1; Hepatitis C virus; Human; Hydrogen; Immunization; Immunize; Immunologics; Infection; Letters; Macaca; Mass Spectrum Analysis; Methods; Molecular Sieve Chromatography; Negative Staining; Outcome; Particulate; Pathway interactions; Procedures; Property; Research Institute; Resolution; Roentgen Rays; Series; Sorting - Cell Movement; Stress Tests; Structural Biologist; Structure; Surface; Surface Plasmon Resonance; Techniques; Testing; Titrations; Vaccine Design; Vaccines; Validation; Viral; Viral Envelope Proteins; Work; X-Ray Crystallography; base; biophysical analysis; biophysical techniques; biophysical tools; crosslink; design; experience; immunogenicity; improved; in vivo; influenzavirus; iron oxide nanoparticle; light scattering; nanoparticle; neutralizing antibody; next generation; particle; preservation; response; simian human immunodeficiency virus; trafficking; vaccine trial; ward

Project 2 Abstract Project 2 will generate structural information to validate and improve the design of new generations of SOSIP trimers in close interaction with Project 1 and Core B. Project 2 will take a highly integrative approach involving cryo- and negative-stain-electron microscopy (NS-EM), X-ray crystallography and a range of other biophysical tools (SEC-MALS, ITC, DSC and SPR) to provide a comprehensive biophysical understanding of trimers and trimer-bnAb complexes. The data will be shared with Project 1 to improve the stability, antigenicity and immunogenicity of new SOSIP trimer designs. Dr. Ian A. Wilson will direct Project 2 at The Scripps Research Institute and Dr. Andrew B. Ward will be co-Leader. The Specific Aims are: Aim 1: To biophysically and structurally characterize new SOSIP trimers, including germline targeting SOSIP trimers. We will generate high-resolution structures by cryo-EM or X-ray crystallography to enable further engineering SOSIP trimers, and low-resolution structures by NS-EM to provide rapid feedback about the quality of SOSIP trimers. We will also use NS-EM to assess the quality of trimers after stress tests that mimic vaccine storage and formulation conditions such as the effect of adjuvant mixing. These assays are critical prior to animal immunization studies, to ensure only high-quality trimers are evaluated. Aim 2: To determine high-resolution structures of SHIV infection-elicited, vaccine-induced NAbs, or naïve precursor antibodies isolated with GT-SOSIP trimers. We will solve structures of neutralizing Abs (NAbs) isolated from trimer-immunized and SHIV-infected animals to assess their similarity to known bNAbs and their potential for developing breadth. The goals are to further optimize the presentation of bNAb epitopes on SOSIP trimers and facilitate the design of sets or series of trimers for steering responses toward bnAbs in prime-boost-boost strategies that mimic infection. We will solve structures of SOSIP trimer complexes with germline precursor antibodies to compare with affinity-matured bnAbs. The goal is to design SOSIP trimer boosts for guiding precursor antibodies toward breadth. Aim 3: To design, optimize and determine structures of SOSIP trimers displayed on nanoparticle platforms. We will incorporate SOSIP trimers into various proteinaceous nanoparticles (NPs), such as the two-component self-assembling NPs, via an iterative cycle of structure-based design and testing. The goal is to preserve the desirable antigenic properties of SOSIP trimers, while gaining the immunological benefits of particulate antigen presentation. Working with Project 1 and Core B, we will evaluate a range of designs and thereby generate an arsenal of SOSIP-NP immunogens that will be tested in animals. The outcomes of those studies will further guide the design of new SOSIP-NP constructs.

项目2摘要 项目2将生成结构信息，以验证和改善新一代SOSIP的设计 与项目1和CoreB密切互动的三聚体将采用高度集成的方法，涉及 冷冻污渍和负电源显微镜（NS-EM），X射线晶体学和一系列其他生物物理 工具（SEC-MALS，ITC，DSC和SPR），以提供对三聚体和 Trimer-BNAB复合物。数据将与项目1共享，以提高稳定性，抗原性和 新的SOSIP三聚体设计的免疫原性。 Ian A. Wilson博士将在Scripps Research指导项目2 研究所和安德鲁·B·沃德（Andrew B. Ward）博士将是共同领导。具体目的是： 目的1：以生物物理和结构表征新的SOSIP三聚体，包括种系 靶向SOSIP三聚体。我们将通过冷冻EM或X射线晶体学生成高分辨率结构 为了实现进一步的工程SOSIP三聚机和NS-EM的低分辨率结构以提供快速反馈 关于SOSIP三聚体的质量。我们还将使用NS-EM评估压力测试后的三聚体质量 模拟疫苗存储和制定条件，例如调整混合的效果。这些测定至关重要 在进行动物免疫抑制研究之前，以确保仅评估高质量的三聚体。 目标2：确定SHIV感染引诱的高分辨率结构，疫苗诱导的 用GT-SOSIP三聚体分离的NABS或天真的前体抗体。我们将解决 从三聚体免疫和湿透的动物中分离出的中和ABS（NABS），以评估其相似性 已知的bnabs及其发展广度的潜力。目标是进一步优化 sosip三聚体上的bnab表位，并促进了集合或一系列三聚体的设计，以转向响应 模仿感染的主要促进策略中的BNAB。我们将解决SOSIP三聚体配合物的结构 用生殖线前体抗体与亲和力成熟的bnab进行比较。目标是设计SOSIP Trimer 提升前体抗体的促进。 目标3：设计，优化和确定显示的SOSIP三聚机的结构 纳米颗粒平台。我们将将SOSIP三聚体纳入各种蛋白质纳米颗粒（NP）， 例如，通过基于结构的设计和测试的迭代循环，诸如两个组分的自组装NP。这 目标是保留SOSIP三聚体的理想抗原特性，同时获得免疫学益处 特殊的抗原表现。使用项目1和核心B，我们将评估一系列设计和 从而产生将在动物中测试的SOSIP-NP免疫原子库。这些结果 研究将进一步指导新的SOSIP-NP构建体的设计。