B Cell Immunity to Hepatitis C Virus

丙型肝炎病毒（HCV）感染在大多数患者中持续多年，并导致慢性肝病， 强大的免疫反应。虽然没有明确建立的保护性免疫的体外相关性， 多种证据表明，CD4+和CD8 + T细胞反应，虽然对控制急性 感染，不足以预防长期持续。与此同时，新出现的证据表明， 支持病毒中和（Vn）抗体的重要性，以及B细胞应答改变Vn的能力。 感染的过程。因此，有效的疫苗将需要诱导稳健的T细胞以及B细胞 应答为了设计一种疫苗免疫原，能够引发广泛的反应性，Vn抗体对此 由于HCV是高度多样性的病毒，因此更好地定义HCV的免疫原性决定簇至关重要。我们 假设是这些Vn表位以离散簇（Vn结构域）组织，且特异性结构域 或者直接参与病毒辅助受体的附着或结合，或者参与构象变化 是病毒成熟或进入所必需的。我们还提出，某些重叠的Vn表位可能是 更保守，导致病毒逃逸的突变能力更低。这一提议汇集了两个 高效和互补的研究团队，努力测试这些假设，并开发 急需的关于HCV的Vn表位的结构的信息。多个人的表征 由Festival实验室分离的抗HCV单克隆抗体（HMAbs）揭示了 E2上的多个免疫原性结构域，其中至少三个通过阻断E2与CD81的结合介导Vn， HCV进入的必需辅助受体。柠檬实验室的补充工作导致了 允许在体外表征Vn和从Vn逃逸的新型细胞培养许可病毒 抗体的我们的近期目标是利用这些新的工具，并确定广泛保守， HCV包膜上的免疫原性结构域引发Vn抗体。我们将决定哪个信封 蛋白质结构域参与病毒进入的特定阶段，每个结构域内的表位 被广泛的Vn抗体识别，并且不太能够维持逃逸突变。这将是 通过进一步研究一组广泛的Vn HMAb，产生新的Vn HMAb， 使用细胞培养感染性HCV嵌合体体外选择和表征Vn逃逸突变体 与来自基因型1a H77c病毒的结构蛋白。这些新的逃逸突变体将被用作 抗原用于选择另外的HMAb。总的来说，这些研究将创造一个高分辨率，功能性 包括基因型特异性和广泛的主要结合位点的构象Vn表位图 HCV包膜上的Vn抗体。从这些努力中获得的信息将为以下方面提供基础： 合理的疫苗设计，并提供急需的抗体的分子特异性， 应该通过免疫引起或对免疫治疗有用

Hepatitis C virus (HCV) infection persists for years in most patients and leads to chronic liver disease despite robust immune responses. Although there are no clearly established in vitro correlates of protective immunity, multiple lines of evidence suggest that CD4+ and CD8+ T cell responses, while critical for controlling acute infection, are insufficient for prevention of long-term persistence. At the same time, emerging evidence supports the importance of virus neutralizing (Vn) antibodies, and the ability of B cell responses to modify the course of infection. Thus, an effective vaccine will need to induce both robust T cell as well as B cell responses. To design a vaccine immunogen capable of eliciting broadly reactive, Vn antibodies against this highly diverse virus, it is critically important to better define the immunogenic determinants of HCV. Our hypothesis is that these Vn epitopes are organized in discrete clusters (Vn domains) and that specific domains either participate directly in attachment or binding to viral co-receptors, or in the conformational changes required for viral maturation or entry. We also propose that certain overlapping Vn epitopes are likely to be more conserved and less capable of mutations leading to virus escape. This proposal brings together two highly productive and complementary research teams in an effort to test these hypotheses and develop critically needed information on the structure of the Vn epitopes of HCV. Characterization of multiple human monoclonal antibodies (HMAbs) to HCV isolated by the Foung laboratory has revealed the existence of multiple immunogenic domains on E2, at least three of which mediate Vn by blocking E2 binding to CD81, an essential co-receptor for HCV entry. Complementary work in the Lemon laboratory has led to the development of novel cell culture-permissive viruses allowing in vitro characterization of Vn and viral escape from Vn antibodies. Our immediate aims are to exploit these novel tools and to identify broadly conserved, immunogenic domains on the HCV envelope that elicit Vn antibodies. We will determine which envelope protein domains are involved in specific phases of virus entry, and which epitopes within each domain are recognized by broadly Vn antibodies and are less capable of sustaining escape mutations. This will be accomplished through further studies of an extensive panel of Vn HMAbs, generation of new Vn HMAbs, and the in vitro selection and characterization of Vn escape mutants using a cell culture-infectious HCV chimera with structural proteins derived from genotype 1a H77c virus. These novel escape mutants will be utilized as antigen for selection of additional HMAbs. Collectively, these studies will create a high-resolution, functional map of conformational Vn epitopes comprising the major binding sites of both genotype-specific and broadly Vn antibodies on the HCV envelope. The information gained from these efforts will provide the basis for rational vaccine design, and provide much needed insight into the molecular specificities of antibodies that should be elicited by immunization or that would be useful for immunotherapy