Engineering of B cell targeted antigens and pathogens

B细胞靶向抗原和病原体的工程

• 批准号: 8038439
• 负责人: Andrea Janine Sant
• 金额: $ 7.62万
• 依托单位: UNIVERSITY OF ROCHESTER
• 依托单位国家: 美国
• 财政年份: 2010
• 资助国家: 美国
• 起止时间: 2010-03-05 至 2012-02-29
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/8038439
• 关键词: Address; Affinity; Antibody Affinity; Antibody Formation; Antibody-Producing Cells; Antigen Targeting; Antigens; B-Lymphocytes; Binding; CD4 Positive T Lymphocytes; Cell surface; Dendritic Cells; Derivation procedure; Elements; Engineering; Epitopes; Evaluation; Frequencies; Generations; Goals; Histocompatibility Antigens Class II; Immunoglobulins; Influenza A virus; Lead; Libraries; Life; Lymphocyte; Mediating; Molecular; Pattern; Peptide antibodies; Peptides; Phage Display; Physiological; Problem Solving; Production; Proteins; Reagent; Receptors, Antigen, B-Cell; Recruitment Activity; T-Lymphocyte; T-Lymphocyte Epitopes; Transgenic Mice; Transgenic Organisms; Vaccination; Viral Antigens; Virus; immunoglobulin receptor; improved; in vivo; influenzavirus; insight; pathogen; prevent; public health relevance; research study; tool; uptake; vaccine efficacy

DESCRIPTION (provided by applicant): To generate a long-lived, high-affinity antibody response, B-cells need to obtain cognate help from antigen-specific CD4+ T cells. The ability of B cells to recruit help depends on their ability to internalize the antigen or pathogen, degrade it, and present the derived peptides via their MHC class II proteins. At the core of this proposal is the question of whether B cells in need of CD4 T cell help display a distinct subset of antigen-derived peptides than do the dendritic cells (DC) that initially drive CD4 T cell expansion. We hypothesize that the repertoire of pathogen and antigen-derived peptides displayed by B cells and DC will be distinct and that B cell-presented peptides will be only a subset of that displayed by DC. This incomplete pattern of epitope presentation by B cells will lead to limitations on the participation of CD4 T cells to deliver help for production of high affinity antibodies. The main element that has limited our ability to address this important issue is that antigen-specific B cells for any given antigen are present in exceeding low frequencies in vivo. These exceptionally low frequencies prevent tracking of the B cells and evaluation of their peptide display under normal physiological conditions. We propose to solve this problem by taking advantage of B cell receptor (BcR) transgenic mice where a majority of the B cells express a known, defined immunoglobulin on their cell surface. The goal of the experiments in this proposal is to use molecular engineering to endow an antigenic protein or virus the ability to be specifically recognized by the immunoglobulin receptor of BcR transgenic mice. We will use phage display libraries to identify peptide mimics (mimetopes) that bind selectively to the defined monoclonal immunoglobulin molecule and then transfer these mimetopes to several selected antigens, including influenza A virus. This strategy will allow selective targeting of the antigen or virus to the antigen-specific immunoglobulin receptor transgenic B cells. Successful derivation of these unique reagents will allow us to assess CD4 T cell epitope generation after immunoglobulin-mediated uptake of virus or antigen by B cells. Most importantly, use of the derived antigens and the B cell receptor transgenic mice will greatly enhance our ability to understand the interaction between influenza virus and antigen-specific B cells and cognate interactions between B cells and CD4 T cells in vivo. PUBLIC HEALTH RELEVANCE: For vaccination to be successful in eliciting a protective antibody response to pathogen challenge, B lymphocytes need to be able to recruit "help" from another lineage of lymphocytes called T cells. Our experiments are aimed at generating several key experimental tools that will allow us to dissect and ultimately control the focus of T cells, to allow them to more efficiently provide help to antibody producing cells. Through these experiments, we will gain the insight needed to improve vaccine efficacy.

描述（由申请人提供）：为了产生长寿命、高亲和力的抗体反应，B 细胞需要从抗原特异性 CD4+ T 细胞获得同源帮助。 B 细胞招募帮助的能力取决于它们内化抗原或病原体、降解抗原或病原体以及通过 MHC II 类蛋白呈递衍生肽的能力。该提案的核心问题是，与最初驱动 CD4 T 细胞扩增的树突状细胞 (DC) 相比，需要 CD4 T 细胞的 B 细胞是否有助于展示抗原衍生肽的独特子集。我们假设 B 细胞和 DC 展示的病原体和抗原衍生肽的库将是不同的，并且 B 细胞呈现的肽将只是 DC 展示的肽的一个子集。 B 细胞这种不完整的表位呈递模式将导致 CD4 T 细胞参与高亲和力抗体生产的过程受到限制。限制我们解决这一重要问题的能力的主要因素是，任何给定抗原的抗原特异性 B 细胞在体内的存在频率都非常低。这些极低的频率阻碍了对 B 细胞的追踪以及在正常生理条件下对其肽展示的评估。我们建议通过利用 B 细胞受体 (BcR) 转基因小鼠来解决这个问题，其中大多数 B 细胞在其细胞表面表达已知的、确定的免疫球蛋白。本提案的实验目标是利用分子工程赋予抗原蛋白或病毒被BcR转基因小鼠的免疫球蛋白受体特异性识别的能力。我们将使用噬菌体展示文库来识别选择性结合特定单克隆免疫球蛋白分子的肽模拟物（模拟表位），然后将这些模拟表位转移到几种选定的抗原上，包括甲型流感病毒。该策略将允许选择性地将抗原或病毒靶向抗原特异性免疫球蛋白受体转基因 B 细胞。这些独特试剂的成功衍生将使我们能够评估 B 细胞免疫球蛋白介导的病毒或抗原摄取后 CD4 T 细胞表位的生成。最重要的是，使用衍生的抗原和B细胞受体转基因小鼠将大大增强我们理解流感病毒和抗原特异性B细胞之间的相互作用以及体内B细胞和CD4 T细胞之间的同源相互作用的能力。 公共卫生相关性：为了使疫苗接种成功地引发针对病原体攻击的保护性抗体反应，B 淋巴细胞需要能够从另一种称为 T 细胞的淋巴细胞谱系中招募“帮助”。我们的实验旨在生成几个关键的实验工具，使我们能够剖析并最终控制 T 细胞的焦点，使它们能够更有效地为抗体产生细胞提供帮助。通过这些实验，我们将获得提高疫苗功效所需的见解。