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: 10614987
• 负责人: Pamela J Bjorkman
• 金额: $ 37.14万
• 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/10614987
• 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; 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; Mus; Orthologous Gene; Pan Genus; Persons; Point Mutation; Preventive vaccine; Primary carcinoma of the liver cells; Production; Proteins; Research Project Grants; Resolution; Sampling; Serum; Sorting; Structure; T cell response; T-Lymphocyte; Testing; Vaccinated; Vaccination; Vaccines; Variant; Virion; Virus; Virus Diseases; Virus-like particle; Work; antigen binding; cost; cross reactivity; design; env Gene Products; epidemic response; epidemic virus; hepatitis C virus envelope 2 protein; immunogenic; immunogenicity; improved; lonely individuals; mosaic; nanoparticle; neutralizing antibody; nonhuman primate; novel; novel vaccines; preservation; receptor binding; response; structural determinants; three dimensional structure; vaccination strategy; 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.

项目概要 慢性丙型肝炎病毒感染是肝病和肝细胞癌的主要原因，但尚未开发出疫苗 由于其极高的遗传多样性，针对丙型肝炎病毒（HCV）而开发。尽管病毒的变异性很高，但仍有 30% 的 HCV 感染者通过产生广泛中和抗体 (bNAb) 来清除病毒感染， 与 HCV 糖蛋白 E2 受体结合位点的保守表位和构象表位结合 跨越E1E2异二聚体。如果我们能够在保护性的同时刺激此类 HCV bNAb 的产生 通过疫苗接种产生 T 细胞反应，HCV 的流行可能会得到遏制。 我们最近从不同的 HCV 分离株中鉴定出几种 E2 蛋白，它们与人类种系前体结合。 bNAbs，表明使用这些变体开发疫苗。然而，目前尚不清楚哪一个会导致 更有效的候选疫苗：基于E2或E1E2的免疫原。在本提案中，我们旨在评估 两组候选疫苗以及在非人类灵长类动物中传递免疫原的不同方法 包括能够靶向 E1E2 中和表位的种系基因片段。该工作分为 三大具体目标。目标#1：制定免疫策略，引发 bNAb 来保护 HCV E1E2 表位。我们将与 Project 5 合作，在小鼠身上测试不同的免疫平台，并对两者进行评估， 针对 E1E2 构象表位的中和抗体的开发（本项目）和多特异性 T 细胞反应（项目 1）。接下来，关于 T 细胞和抗体诱导的最佳候选疫苗 每一个都将在非人类灵长类动物中进行测试，这些灵长类动物经常利用抗体基因片段的直系同源物 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中接种疫苗后以及在人类中清除自然 HCV 感染后引发 bNAb。 除了分析目标 1 中免疫动物的血清反应外，我们还将生成高分辨率 疫苗诱导的 bNAb 或从清除其抗体的个体中分离出的人抗体的晶体结构 感染（项目 2）。我们将在免疫后对E2特异性猕猴B细胞中的抗体基因进行排序和测序 并从这些序列表达抗体。然后我们将确定疫苗诱导的 bNAb 的结构 与免疫原形成复合物，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 使用冷冻电子断层扫描 (cryo-ET) 在膜环境中测量长度 E1E2 异二聚体。扩大 根据目标 2 中的 bNAb-E2 结构，我们将使用冷冻电子断层扫描和亚断层扫描平均来检查 bNAb 识别完整膜形式的 E1E2 异二聚体。这些目标的结果将共同导致 引发 HCV 特异性 bNAb 和强 T- 的新型候选疫苗的综合表征 细胞反应，促进有效 HCV 疫苗的开发。