PROJECT 2:Structural studies of SOSIP trimers

项目2：SOSIP三聚体的结构研究

• 批准号: 10427133
• 负责人: IAN A WILSON
• 金额: $ 119.94万
• 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/10427133
• 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; 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; rational design; 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 项目2将生成结构信息，以验证和改进新一代SOSIP的设计 三聚体与项目1和核心B密切相互作用。项目2将采取高度综合的方法， 低温和负染电子显微镜（NS-EM），X射线晶体学和一系列其他生物物理学 工具（SEC-MALS，ITC，DSC和SPR），以提供对三聚体的全面生物物理理解， 三聚体-bnAb复合物。数据将与项目1共享，以提高稳定性、抗原性和 新SOSIP三聚体设计的免疫原性。Ian A.博士威尔逊将在斯克里普斯研究中心指导项目2 研究所和安德鲁博士B。沃德将担任联合领导人。具体目标是： 目的1：对新的SOSIP三聚体进行生物药理学和结构表征， 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 通过NS-EM进一步设计SOSIP三聚体和低分辨率结构，以提供快速反馈 关于SOSIP三聚体的质量。我们还将使用NS-EM来评估压力测试后三聚体的质量， 模拟疫苗储存和配制条件，例如佐剂混合的效果。这些化验是至关重要的 在动物免疫研究之前，以确保仅评价高质量的三聚体。 目的2：确定SHIV感染引起的、疫苗诱导的、 NAb或用GT-SOSIP三聚体分离的幼稚前体抗体。我们将解决的结构 从三聚体免疫和SHIV感染的动物中分离的中和抗体（NAb），以评估它们与 已知的bNAb及其开发广度的潜力。目标是进一步优化 bNAb表位，并促进设计一组或一系列的三聚体，以引导对SOSIP三聚体的应答。 模拟感染的初免-加强-加强策略中的bnAb。我们将解决SOSIP三聚体复合物的结构 用种系前体抗体与亲和力成熟的bnAb进行比较。目标是设计SOSIP三聚体 加强引导前体抗体向广度。 目的3：设计、优化和确定SOSIP三聚体的结构， 纳米粒子平台。我们将SOSIP三聚体掺入各种蛋白质纳米颗粒（NP）中， 例如双组分自组装纳米粒子，通过基于结构的设计和测试的迭代循环。的 我们的目标是保留SOSIP三聚体的理想抗原特性，同时获得免疫学益处 微粒抗原递呈与项目1和核心B合作，我们将评估一系列设计， 从而产生将在动物中测试的SOSIP-NP免疫原的武库。这些成果 研究将进一步指导新SOSIP-NP构建体的设计。