Antigenic landscape of the human helminth IgE antibody response

人类蠕虫 IgE 抗体反应的抗原图谱

基本信息

批准号：
9897458
负责人：
Scott Alan Smith
金额：
$ 35.78万
依托单位：
VANDERBILT UNIVERSITY MEDICAL CENTER
依托单位国家：
美国
项目类别：
财政年份：
2017
资助国家：
美国
起止时间：
2017-04-06 至 2022-03-31
项目状态：
已结题

来源：
https://reporter.nih.gov/project-details/9897458
关键词：
Adaptive Immune System Affect Affinity Allergens Allergic Allergic Disease Antibodies Antibody Repertoire Antibody Response Antigen Targeting B-Lymphocytes Basophils Binding Biological Assay Blood Blood Circulation Cell Culture Techniques Cell physiology Cells Clinical Clone Cells Column Chromatography Complex Developed Countries Developing Countries Development Diagnostic Disease Dose Effector Cell Enzyme-Linked Immunosorbent Assay Epitopes Filaria bancrofti Filariasis Frequencies Funding Generations Goals Helminth Proteins Helminths Homologous Gene Homologous Protein Human Hybridomas IgE IgG4 Immune Immune response Immune system Immunity Immunoglobulin Class Switching Immunoglobulin Somatic Hypermutation In Vitro Knowledge Length Maps Mass Spectrum Analysis Mediating Mediator of activation protein Memory B-Lymphocyte Methods Molecular Monoclonal Antibodies Mononuclear Parasites Patients Play Population Preparation Process Protein Family Proteins Recombinants Role Sampling Sequence Analysis Serum Signal Transduction Site Specificity Strongyloides stercoralis Strongyloidiasis Techniques Technology Testing Time Vaccines Variant Western Blotting Work base cross reactivity design falls group competition helminth infection human monoclonal antibodies immunoaffinity chromatography improved insight mast cell murine monoclonal antibody novel novel vaccines pathogen prevent restraint screening vaccine candidate

项目摘要

Antigenic landscape of the human anti-helminth IgE antibody response Scott A. Smith ABSTRACT Despite its perceived central role in helminth immunity, very little is known about the naturally occurring human IgE antibody response, the dominant helminth target proteins or even the size and complexity of the functional antibody repertoire. Most of our knowledge of the IgE antibodies targeting helminth infection and/or allergens has come from studies using polyclonal sera or inferred from murine monoclonal antibodies (mAbs). We hypothesize that the human anti-helminth IgE antibody response is complex but is comprised of B cell clones which target helminth proteins that are homologues of known allergen proteins and are capable of mediating key effector cell functions in vitro. Here we employ a human B cell hybridoma method newly created in my lab, immortalizing growing memory B cells to generate for the first time ever, naturally occurring full-length human IgE mAbs. Patient clinical information is used to select samples that contain cells directed toward important helminth pathogens. In the initial studies described in this proposal, we selected three helminth infected subjects, one having strongyloidiasis and two with filariasis. The frequencies of IgE encoding B cells was found to be approximately three per million mononuclear cells with great variability seen in their reactivity to helminth lysate. These B cell frequencies and our technical efficiencies are sufficient to greatly expand this work and make hundreds of IgE mAbs. Due to funding restraints our initial characterization studies were performed on a small panel of six Wuchereria bancrofti IgE and three Strongyloides stercoralis IgE mAbs. Purified IgE mAbs were also found to have variable reactivity to helminth lysate in both ELISA and Western blot. Specific helminth protein targets were identified by immunoaffinity chromatography/mass spectrometry and fell into known allergen protein families. Crude allergen protein cross-reactivity studies were performed using Phadia diagnostic technologies and found in some cases to be positive, suggesting that allergens can resemble helminth proteins. IgE sequence analysis demonstrates significant degrees of somatic hypermutation. Initial functional studies show that each IgE helminth-specific mAb is capable of dose dependent mast cell mediator release in the presence of lysate - functional potency studies are underway. Ultimately helminth- specific IgE mAb panels will be assembled to reflect a hierarchy of immune dominant helminth protein targets and effector cell functionality. In addition to improving our basic understanding of this poorly studied branch of human immunity and allergic sensitization, the information obtained through studies outlined in this proposal will allow for the design and development of new helminth vaccines.

人类抗螺旋IgE抗体反应的抗原景观Scott A. Smith 抽象的尽管它在蠕虫免疫中具有中心作用，但对自然的知识知之甚少发生人IgE抗体反应，主要的蠕虫靶蛋白，甚至大小和功能抗体库的复杂性。我们大多数对IgE抗体靶向的知识蠕虫感染和/或过敏原来自使用多克隆血清或从中推断出的研究鼠单克隆抗体（mAb）。我们假设人类抗螺旋IgE抗体反应很复杂，但由B细胞克隆组成，靶向蠕虫蛋白已知过敏原蛋白的同源物，能够介导关键效应细胞体外功能。在这里，我们采用了在我的实验室中新创建的人类B细胞杂交瘤方法，使生长不朽记忆B细胞首次生成，自然发生全长的人IgE mAb。患者临床信息用于选择包含针对重要细胞的细胞的样品蠕虫病原体。在本提案中描述的最初研究中，我们选择了三个舵手感染的受试者，一名患有较强的山地病，两个患有丝虫病。 IGE编码的频率发现B细胞约为每百万个单核细胞大约三个，在它们对蠕虫裂解物的反应性。这些B细胞频率和我们的技术效率足够为了大大扩展这项工作并制造数百个Ige mabs。由于资金限制了我们的最初对六个wucheria bancrofti ige和三个小组进行了表征研究强肌链蛋白酶刺mabs。还发现纯化的IgE mab具有可变的反应性 Elisa和Western印迹中的蠕虫裂解物。特定的蠕虫蛋白靶标通过免疫亲和力色谱/质谱法，并落入已知的过敏原蛋白家族。粗制过敏原蛋白交叉反应性研究使用Phadia诊断技术进行并在某些情况下发现阳性，这表明过敏原可能类似于蠕虫蛋白。 IgE序列分析表明，体细胞过度的显着程度。初始功能研究表明，每个IgE蠕虫特异性MAB都能依赖剂量的肥大细胞介质在存在裂解物 - 功能效力研究的情况下正在释放。最终蠕虫 - 特定的IgE mAb面板将组装以反映免疫主导蠕虫蛋白的层次结构靶标和效应细胞功能。除了提高我们对这一差的基本理解之外研究了人类免疫力和过敏敏化的分支，通过研究获得的信息该提案中概述的将允许设计和开发新的蠕虫疫苗。