Memory B Cell Isolation and Antibody Characterization

记忆 B 细胞分离和抗体表征

• 批准号: 8829136
• 负责人: Yuxing Li
• 金额: $ 40.31万
• 依托单位: SCRIPPS RESEARCH INSTITUTE, THE
• 依托单位国家: 美国
• 资助国家: 美国
• 起止时间: 至
• 项目状态: 未结题
• 来源: https://reporter.nih.gov/project-details/8829136
• 关键词: Affinity; Alanine; Animals; Antibodies; Antibody Response; Antibody Specificity; Antigens; Arts; Automobile Driving; B cell repertoire; B-Lymphocytes; Binding; Binding Sites; Bioinformatics; Biological Assay; CD4 Antigens; Cell Separation; Cells; Chronic; Cloning; Complement; Complex; Elements; Engineering; Epitopes; Evaluation; Evolution; Genetic; HIV; HIV vaccine; HIV-1; Human; Immune response; Immunization; Immunoglobulin Class Switching; Immunoglobulin G; Immunoglobulin M; Individual; Infection; Instruction; Kinetics; Knowledge; Maps; Memory; Memory B-Lymphocyte; Monoclonal Antibodies; Outcome Study; Pathway interactions; Play; Primates; Process; Property; Regimen; Resolution; Reverse Transcriptase Polymerase Chain Reaction; Role; Scanning; Serum; Sorting - Cell Movement; Specificity; Structure of germinal center of lymph node; Testing; Vaccinated; Vaccination; Vaccine Design; Vaccines; Virus; base; design; env Glycoproteins; improved; insight; neutralizing antibody; nonhuman primate; novel; response; vaccine candidate; vaccine development; vaccine efficacy

One of the major challenges of HlV-1 vaccine development is the elicitation of broadly neutralizing antibodies (bNAbs) against HIV Env. The recent isolation of several bNAbs from HIV-infected individuals demonstrates that the human B cell repertoire can generate bNAbs targeting the conserved Env region. However, there is still a tremendous knowledge gap regarding how the broad responses are elicited during chronic HIV-1 infection and, additionally, how these compare to the much more limited responses elicited by Env vaccination. To fill this information gap, we propose to define and improve the elicitation of neutralizing antibody responses toward the most functionally conserved and accessible element ofthe HIV-1 Env complex, including the receptor CD4 binding site (CD4bs). Previously, we developed a multicolor Env epitope-specific B cell sorting and RT-PCR strategy to analyze the memory B cell compartment of HIVinfected individuals, which led to the isolation of CD4bs potent and bNAb, VRCOl, which neutralizes >90% of circulating primary viruses. We adapted this epitope-specific memory B cell sorting strategy to extend our analyses to gain insight of Env B cell response during and following Env immunogen vaccination into nonhuman primates (NHPs). Accordingly, the aims of this Project 3 are summarized as follows. In Aim 1 we will develop envelope-specific, including CD4bs-specific memory B cell isolation and monoclonal antibody cloning from Env immunogen vaccinated NHP animals and design new and novel probes specific for bNAb isolation. In Aim 2, we will characterize the Env-specific monoclonal antibody specificities with state ofthe art binding and characterization assays. In Aim 3, we will investigate the role, composition, and evolution pathway ofthe Env-specific IgM memory B cell compartment, a subset of memory B cells, which recently has been shown to play an important role in the long term B cell response.

HLV-1疫苗开发的主要挑战之一是引起广泛中和抗体 （bnabs）反对艾滋病毒环境。最近，来自HIV感染者的几个BNAB的分离证明了 人类B细胞库可以产生针对保守的ENV区域的BNAB。但是，有 关于如何在慢性HIV-1期间引起广泛反应的巨大知识差距仍然存在巨大的知识差距 感染，此外，它们与Env引起的更有限的反应相比 疫苗接种。为了填补此信息空白，我们建议定义和改善中和的启发 抗体对HIV-1 Env的功能最保守和可访问元件的反应 复合物，包括受体CD4结合位点（CD4B）。以前，我们开发了一个多色env 表位特异性B细胞分选和RT-PCR策略，以分析Hivincted的记忆B细胞室 个体，导致CD4BS有效的隔离和BNAB，VRCOL，中和> 90％ 循环原发病毒。我们调整了这种表位特定的内存B细胞分类策略以扩展我们的 分析以洞察ENV B细胞反应期间和之后的INV免疫原生成疫苗接种到非人类 灵长类动物（NHP）。因此，该项目3的目的总结如下。在目标1中，我们将 开发特异性包膜，包括CD4BS特异性内存B细胞分离和单克隆抗体 从Env免疫原的NHP动物进行克隆，并设计针对BNAB的新的和新颖的探针 隔离。在AIM 2中，我们将表征ENV特定的单克隆抗体特异性的特异性 结合和表征分析。在AIM 3中，我们将研究角色，组成和进化 ENV特异性IgM内存B细胞室的途径，Memory B细胞的子集，最近 已显示在长期B细胞反应中起重要作用。