Characterization of HCV vaccine induced-neutralizing antibody response in non-human primates

HCV 疫苗在非人灵长类动物中诱导中和抗体反应的特征

• 批准号: 10205734
• 负责人: Pamela J Bjorkman
• 金额: $ 39.23万
• 依托单位: JOHNS HOPKINS UNIVERSITY
• 依托单位国家: 美国
• 财政年份: 2021
• 资助国家: 美国
• 起止时间: 2021-05-01 至 2026-04-30
• 项目状态: 未结题
• 来源: https://reporter.nih.gov/project-details/10205734
• 关键词: Adenoviruses; Animals; Antibodies; Antibody Avidity; Antibody Response; Antigens; Antiviral Agents; B-Lymphocytes; Binding; Binding Sites; Biological Assay; Chronic; Chronic Hepatitis C; Complex; Cryo-electron tomography; Crystallization; Development; Environment; Enzyme-Linked Immunosorbent Assay; Epidemic; Epitopes; Genes; Genetic Variation; Hepatitis C; Hepatitis C Vaccine; Hepatitis C virus; Human; Immune response; Immunization; Immunize; In Vitro; Individual; Infection; Length; Liver Cirrhosis; Liver diseases; Macaca; Mediating; Membrane; Methods; Molecular Conformation; Mosaicism; Mus; Orthologous Gene; Pan Genus; Point Mutation; Preventive vaccine; Primary carcinoma of the liver cells; Production; Proteins; Research Project Grants; Resolution; Sampling; Serum; Structure; Study of serum; T cell response; T-Lymphocyte; Testing; Tomogram; Vaccinated; Vaccination; Vaccines; Variant; Virion; Virus; Virus Diseases; Virus-like particle; Work; antigen binding; base; cost; cross reactivity; design; env Gene Products; hepatitis C virus envelope 2 protein; immunogenic; immunogenicity; improved; nanoparticle; neutralizing antibody; nonhuman primate; novel; novel vaccines; preservation; receptor binding; response; three dimensional structure; vaccine candidate; vaccine development; vaccine trial

PROJECT SUMMARY Chronic HCV infection is a leading cause of liver disease and hepatocellular carcinoma, yet no vaccine has been developed against HCV due to its extremely high genetic diversity. Despite the high variability of the virus, 30% of HCV-infected individuals clear the viral infection by developing broadly neutralizing antibodies (bNAbs) that bind to conserved epitopes in the receptor binding site of HCV glycoprotein E2 and conformational epitopes spanning the E1E2 heterodimer. If we could stimulate the production of such HCV bNAbs along with protective T cell responses by vaccination, a rising epidemic of HCV might be stopped. We recently identified several E2 proteins from different HCV isolates that bind to germline precursors of human bNAbs, suggesting vaccine development using these variants. However, it is not yet clear which would make a more effective vaccine candidate: E2-based or E1E2-based immunogens. In this proposal, we aim to evaluate both sets of vaccine candidates along with different methods of immunogen delivery in non-human primates that include germline gene segments capable of targeting E1E2 neutralizing epitopes. The work is organized into three major Specific Aims. Aim #1: Develop immunization strategies that elicit bNAbs to conserved HCV E1E2 epitopes. Working together with Project 5, we will test different immunization platforms in mice, evaluating both, the development of neutralizing antibodies to E1E2 conformational epitopes (this Project), and multi-specific T cell responses (Project 1). Next, the best vaccine candidates with respect to both T cell and antibody induction will each be tested in non-human primates, which utilize an ortholog of an antibody gene segment frequently used by the most potent human HCV-specific bNAbs. Aim #2: Generate high-resolution structures of E2-specific bNAbs elicited after vaccination in non-human primates and after clearance of natural HCV infection in humans. In addition to analyzing serum responses from immunized animals in Aim 1, we will generate high-resolution crystal structures of vaccine-induced bNAbs or human antibodies isolated from individuals who cleared their infection (Project 2). We will sort and sequence antibody genes in E2-specific macaque B cells after immunization and express antibodies from these sequences. We will then determine the structures of vaccine-induced bNAbs in complex with immunogens and compare them to structures of human bNAbs bound to E2 proteins (Project 4) to elucidate the structural determinants of broad neutralization of HCV. Aim #3: Determine a structure of the full- length E1E2 heterodimer in a membrane environment using cryo-electron tomography (cryo-ET). To expand upon bNAb-E2 structures from Aim 2, we will use cryo-ET with sub-tomogram averaging to examine bNAb recognition of the E1E2 heterodimer in its intact membrane form. Together, the results of these aims will lead to the comprehensive characterization of novel vaccine candidates that elicit HCV-specific bNAbs and strong T- cell responses, facilitating the development of an effective HCV vaccine.

项目摘要 慢性HCV感染是肝病和肝细胞癌的主要原因，但没有疫苗是 由于其极高的遗传多样性而针对HCV开发。尽管病毒变异很高，但30％ HCV感染的个体通过开发广泛中和抗体（BNAB）来清除病毒感染 与HCV糖蛋白E2和构象表位的受体结合位点中的保守表位结合 跨越E1E2异二聚体。如果我们可以刺激此类HCV BNAB的产生以及保护性 T细胞反应通过疫苗接种，可能会停止HCV的流行。 我们最近从不同的HCV分离株中鉴定了几种与人类生殖前体结合的E2蛋白 BNAB，建议使用这些变体的疫苗开发。但是，尚不清楚哪个会成为 更有效的候选疫苗：基于E2或基于E1E2的免疫原。在此提案中，我们旨在评估 两组候选疫苗的候选物以及在非人类灵长类动物中的免疫传递方法 包括能够靶向E1E2中和表位的种系基因段。工作被组织成 三个主要的特定目标。目标＃1：制定免疫策略，以使BNABS为保守的HCV E1E2 表位。与项目5合作，我们将测试小鼠中不同的免疫平台，评估这两者， 对E1E2构象表位（该项目）和多特异性T的中和抗体的开发 细胞反应（项目1）。接下来，相对于T细胞和抗体诱导的最好的疫苗候选物 每个人都会在非人类灵长类动物中进行测试，该灵长类动物使用抗体基因段的直系同源 由最有效的人类HCV特异性BNAB使用。目标＃2：生成E2特异性的高分辨率结构 在非人类灵长类动物和人类自然HCV感染清除后，BNABS引起了疫苗接种后引起的。 除了分析AIM 1中免疫动物的血清反应外，我们还将产生高分辨率 疫苗诱导的BNABS或人类抗体的晶体结构从清除其的个体中分离出来 感染（项目2）。免疫后，我们将在E2特异性猕猴B细胞中对序列抗体基因进行排序和序列抗体基因 并从这些序列中表达抗体。然后，我们将确定疫苗诱导的BNAB的结构 与免疫原子复杂化，并将其与与E2蛋白结合的人类BNAB的结构进行比较（项目4） 阐明HCV广泛中和的结构决定因素。目标＃3：确定完整的结构 使用冷冻电子层析成像（Cryo-ET）在膜环境中的长度E1E2异二聚体。扩展 在AIM 2的BNAB-E2结构上，我们将使用与亚图构图平均的Cryo-ET检查BNAB 以其完整的膜形式识别E1E2异二聚体。这些目标的结果在一起将导致 引起HCV特异性BNAB和强t-的新型疫苗候选物的全面表征 细胞反应，促进有效的HCV疫苗的发展。