Molecular and structural characterization of broadly neutralizing anti-HCV antibodies

广泛中和抗 HCV 抗体的分子和结构表征

• 批准号: 9478874
• 负责人: Justin Richard Bailey
• 金额: $ 71.99万
• 依托单位: JOHNS HOPKINS UNIVERSITY
• 依托单位国家: 美国
• 财政年份: 2017
• 资助国家: 美国
• 起止时间: 2017-05-01 至 2022-04-30
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/9478874
• 关键词: Amino Acids; Animal Model; Antibodies; Antibody Binding Sites; Antiviral Therapy; Binding; Binding Sites; Biochemical; CD81 gene; Caring; Complex; Crystallization; Data; Development; Dimensions; Disease; Distant; Epidemiology; Epitopes; Future; Genetic; Genetic Polymorphism; Goals; HIV; Hepatitis C; Hepatitis C Antibodies; Hepatitis C Vaccine; Hepatitis C virus; Human; Immunologics; In Vitro; Individual; Infection; Infection prevention; Investigation; Measures; Modification; Molecular; Monoclonal Antibodies; Proteins; Reagent; Role; Sequence Analysis; Somatic Mutation; Structure; Therapeutic Agents; United States; Vaccine Antigen; Vaccine Design; Vaccines; Variant; Virus; Work; anti-hepatitis C; design; effective therapy; env Gene Products; glycosylation; high risk; neutralizing antibody; pandemic disease; protein folding; vaccine development; vaccine trial; virology; virus envelope

Project Summary A vaccine against Hepatitis C virus (HCV) is urgently needed. HCV infects over 170 million people worldwide and kills more people in the United States annually than HIV. While direct-acting antiviral (DAA) therapy has revolutionized HCV care, control of the HCV pandemic remains challenging due frequent reinfection in high-risk individuals and a high proportion of asymptomatic carriers who continue to infect others. Approximately 30% of individuals who become infected with HCV spontaneously clear the infection, and we have previously shown that this spontaneous clearance of HCV is associated with early development of broadly neutralizing antibodies (bNAbs) against the virus. BNAbs are also protective against HCV infection in multiple animal models. Unfortunately, to date, vaccines against HCV have not induced adequate titers of protective bNAbs. Our inability to induce potent bNAbs is in part due to our poor understanding of the molecular and structural interactions between bNAbs and HCV envelope proteins (E1 and E2). Our preliminary work indicates that envelope sequence polymorphisms distant from bNAb binding sites have a strong, unexpected influence on neutralization sensitivity. These data and rapidly emerging work in HIV indicate that these crucial bNAb-envelope interactions need to be understood in a three dimensional (structural) context. We hypothesize that molecular and structural analysis of bNAb-E2 interactions will allow us to rationally design stable HCV envelope proteins with optimized bNAb epitopes that are ideal for structural and vaccine studies as well as bNAbs with enhanced neutralizing potency and breadth, better defining the ideal antibodies that should be induced by a vaccine. We have characterized a diverse panel of unique HCV envelope proteins and isolated some of the most broadly neutralizing anti-HCV monoclonal antibodies described to date. In Aim 1, we will functionally and molecularly characterize interactions between this panel of diverse, naturally occurring HCV envelope variants and the panel of bNAbs, which will allow us to identify amino acid determinants of neutralization sensitivity of E2 as well as somatic mutations conferring neutralizing potency and breadth to bNAbs. In Aim 2, we will define biochemical and molecular factors influencing stability and native folding of HCV envelope proteins. We will clone more than 100 distinct natural HCV E2 variants and identify polymorphisms associated with stable in vitro E2 expression. In Aim 3, we will determine structural correlates of broad and potent neutralization of HCV. We will crystallize HCV E2 in complex with bNAbs of varying breadth and potency. We will use the data acquired through these three aims to design stable E2 variants with optimized bNAb epitopes that will be ideal reagents for future structure analyses and vaccine studies. In addition, we will design bNAbs with enhanced neutralizing potency and breadth that will define the ideal antibodies that could be induced by a vaccine and may also be useful therapeutic agents. Through these investigations, we will advance rational design of an HCV vaccine.

项目概要 迫切需要针对丙型肝炎病毒（HCV）的疫苗。 HCV 感染全球超过 1.7 亿人 在美国每年造成的死亡人数比艾滋病毒还要多。虽然直接作用抗病毒（DAA）疗法已 尽管彻底改变了 HCV 护理，但由于高危人群频繁再次感染，控制 HCV 大流行仍面临挑战 个人和很大比例的无症状携带者继续感染他人。大约 30% 感染 HCV 的个体会自发清除感染，我们之前已经证明 HCV 的这种自发清除与广泛中和抗体的早期发展有关 （bNAb）对抗病毒。在多种动物模型中，BNAb 还可预防 HCV 感染。 不幸的是，迄今为止，HCV 疫苗尚未诱导出足够滴度的保护性 bNAb。我们的无能 诱导有效的 bNAb 的部分原因是我们对分子和结构相互作用的了解不足 bNAb 和 HCV 包膜蛋白（E1 和 E2）之间的关系。我们的初步工作表明信封 远离 bNAb 结合位点的序列多态性对中和有强烈的、意想不到的影响 敏感性。这些数据和 HIV 领域迅速兴起的工作表明，这些关键的 bNAb-包膜相互作用 需要在三维（结构）背景下进行理解。我们假设分子和结构 bNAb-E2 相互作用的分析将使我们能够合理地设计具有优化的稳定的 HCV 包膜蛋白 非常适合结构和疫苗研究的 bNAb 表位以及具有增强中和作用的 bNAb 效力和广度，更好地定义疫苗应诱导的理想抗体。 我们对一系列独特的 HCV 包膜蛋白进行了表征，并分离出了一些最广泛的 迄今为止描述的中和抗HCV单克隆抗体。在目标 1 中，我们将从功能和分子角度 表征这组不同的、自然发生的 HCV 包膜变体与该组之间的相互作用 bNAb，这将使我们能够识别 E2 中和敏感性的氨基酸决定因素以及 体细胞突变赋予 bNAb 中和效力和广度。在目标 2 中，我们将定义生化 以及影响 HCV 包膜蛋白稳定性和天然折叠的分子因素。我们将克隆超过 100 种不同的天然 HCV E2 变体，并鉴定与稳定的体外 E2 表达相关的多态性。 在目标 3 中，我们将确定广泛且有效的 HCV 中和的结构相关性。我们将结晶 HCV E2 与不同宽度和效力的 bNAb 复合。我们将使用通过这些获得的数据 三个目标是设计具有优化 bNAb 表位的稳定 E2 变体，这将是未来的理想试剂 结构分析和疫苗研究。此外，我们将设计具有增强中和效力的 bNAb 和广度将定义可以由疫苗诱导并且也可能有用的理想抗体 治疗剂。通过这些研究，我们将推进HCV疫苗的合理设计。