An Advanced Model to Analyze CD4T Cel Reactions to Antibody and BCR V Regions

分析 CD4T 细胞对抗体和 BCR V 区反应的高级模型

• 批准号: 8190100
• 负责人: Lawrence J Wysocki
• 金额: $ 7.93万
• 依托单位: NATIONAL JEWISH HEALTH
• 依托单位国家: 美国
• 财政年份: 2011
• 资助国家: 美国
• 起止时间: 2011-08-04 至 2013-07-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/8190100
• 关键词: 4-hydroxy-5-nitrophenyl acetic acid; A/J Mouse; Abbreviations; Ablation; Adoptive Transfer; Affect; Affinity; Animals; Antibodies; Antibody Formation; Antigen Presentation; Antigen Receptors; Antigen-Antibody Complex; Antigens; Autoantigens; Autoimmune Diseases; Autoimmune Process; Autoimmunity; B-Cell Development; B-Lymphocytes; BAC (bacterial artificial chromosome); Back; Binding; Bone Marrow; Breeding; C57BL/6 Mouse; CD4 Positive T Lymphocytes; Chromosomes, Human, 4-5; Clinical; Complementary DNA; Complex; Cross Reactions; Cyan Fluorescent Protein; Data; Deposition; Development; Dissection; Engineering; Epitopes; Event; Fc Receptor; Fluorescence; Future; Gene Targeting; Genes; Genetic; Genetic Recombination; Genomics; Green Fluorescent Proteins; Haptens; Health Hazards; Helper-Inducer T-Lymphocyte; Immune system; Immunity; Immunoglobulin G; Immunoglobulin M; Immunoglobulin Somatic Hypermutation; Immunoglobulins; Inflammation; Inflammatory; Knock-in Mouse; Laboratories; Lectin; Libraries; Life; Light; Link; Location; Lymphopoiesis; Mediating; Memory B-Lymphocyte; Modeling; Mouse Strains; Mus; Mutation; Nuclear; Nuclear Antigens; Organ; Ovalbumin; Ovum; Patients; Peptide antibodies; Peptides; Physiological; Play; Point Mutation; Polysaccharides; Process; Production; Publishing; Reaction; Reagent; Receptors, Antigen, B-Cell; Refractory; Reporter; Role; Self Tolerance; Site; Source; Specificity; Staining method; Stains; Structure of germinal center of lymph node; Surface; Systemic Lupus Erythematosus; T cell differentiation; T cell response; T-Lymphocyte; Testing; Therapeutic; Therapeutic Monoclonal Antibodies; Thymus Gland; Tissues; Transgenic Mice; Transgenic Organisms; Work; acquired immunity; adverse outcome; antigen processing; autoreactive B cell; base; chemokine receptor; genetic analysis; immunogenic; immunogenicity; immunoregulation; mutant; neutralizing antibody; novel; prevent; receptor; response; sound; systemic autoimmune disease; therapy design

DESCRIPTION (provided by applicant): An advanced model to analyze CD4 T cell reactions to antibody and BCR V regions Project Summary This is an R03 proposal to generate an antibody construct and a corresponding transgenic mouse for genetic analyses of T cell reactions to V region peptides derived from antibodies and B cell antigen receptors (BCR). CD4 T cells make a living by interacting with B cells that present antigenic peptides in MHC II. While some of these peptides come from foreign antigen, others are neoantigens generated in the BCR through the processes of V/D/J gene recombination in the bone marrow and somatic hypermutation in the periphery. Neoantigenic BCR peptides in MHC II open a potential avenue of T cell help to B cells, in violation of the principle of linked recognition. We refer to this as the receptor presentation avenue of help. Neoantigenic Ab peptides may also activate CD4 T cells at sites of Ab deposition where APC gather Ab through Fc receptors or surface lectins. At present, we have only a rudimentary understanding of how the immune system regulates potentially dangerous interactions between CD4 T cells and B cells or other APC that are mediated by BCR and Ab peptides. As an example, IgG is ostensibly tolerogenic; yet therapeutic mAb sometimes elicit an endogenous neutralizing antibody response with a potential clinical health hazard. We are pursuing adoptive transfer studies to reveal the consequences of T cell reactions to BCR- and Ab- derived peptides using a model based in mice of the strain A genetic background. Our most recent results indicate that the "receptor presentation" avenue of T help to B cells is subject to immunoregulation. T-B interactions in the GC that are mediated by BCR-derived peptides impede GC participation and memory B cell development and ultimately induce a refractory state in the T cells. Efforts to dissect this novel form of immunoregulation and to explore the issue of Ab immunogenicity versus tolerogenicity, would be greatly enhanced if we could utilize the available library of genetically altered C57BL/6 (B6) mice. Accordingly, we would like to develop a corresponding model in the B6 strain so that genetic alterations can be introduced into adoptively transferred B cells, T cells or recipient mice. A B6 model would also enable us to genetically analyze CD4 T cell reactions to an antibody V region peptide in adoptive recipients of T cells that are injected with mAb carrying the cognate peptide. To this end, we propose to engineer an Ig; light chain construct that contains a sequence encoding a peptide called 3K, which is immunogenic for CD4 T cells in the context of by I-Ab (Aim 1). This will enable production of hapten-specific (NP) mAb of various isotypes for planned future studies of T cell responses to 3K;Ig, immune complexes thereof and glycan modified forms of 3K;Ig. For these studies, we would use a currently available 12TCR Tg mouse as a source of 3K-specific CD4 T cells. In Aim 2, we propose to engineer a modified 3K; genomic construct with a GFP reporter and generate a corresponding B6 transgenic mouse. This mouse will be crossed with a knock-in mouse (B1-8) that carries a complementary targeted heavy chain from an Ig; NP-specific mAb to produce a mouse with NP-specific 3K; B cells. These B cells can then be used with 3K-specific T cells in future co-adoptive transfer genetic studies of receptor presentation. The products of this work will enable a genetic dissection of mechanisms that regulate T cell responses to Ig V region peptides in the BCR or secreted antibody. The anticipated results from this model will advance our understanding of immunoregulation in the face of neoantigens that frequently arise in BCR and Ab and are presented to CD4 T cells on a regular basis during immunity. Abbreviations ANA: Anti-nuclear antibody BAC: Bacterial artificial chromosome BCR: B cell antigen receptor CFP: Cyan fluorescent protein GFP: Green fluorescent protein GC: Germinal Center NP: (4-hydroxy-3-nitro-phenyl)acetyl SHM: Somatic hypermutation PUBLIC HEALTH RELEVANCE: T lymphocytes that react against peptides derived from antibodies are potential health hazards in a variety of situations. They may induce patient responses against therapeutic monoclonal antibodies; they may induce organ inflammation when directed against antibodies that deposit in the organ, and they may stimulate B cells to produce antibodies against self-tissues in systemic autoimmune diseases such as systemic lupus erythematosus (SLE). This project aims to develop a unique state-of-the-art-model that will enable us to apply the power of genetics to reveal how T cells react to antibody peptides and mechanisms that regulate potential adverse outcomes. The information obtained from this model will provide a sound basis for the rational design of therapies to control or prevent inflammatory and autoimmune diseases in which antibodies and B cells play a significant role.

描述（由申请人提供）：一种用于分析抗体和BCR V区域的CD4 T细胞反应的高级模型，该项目摘要这是R03提案，用于生成抗体构建体和相应的转基因小鼠，用于源自抗体和B细胞抗体受体（BCR）的T细胞反应对V区域的T细胞反应。 CD4 T细胞通过与在MHC II中呈现抗原肽的B细胞相互作用来生存。尽管其中一些肽来自外来抗原，但其他肽是通过骨髓中的V/D/J基因重组的过程在BCR中产生的新抗原，而外围物质中的细胞性超含量。 MHC II中的新抗原BCR肽为B细胞开辟了潜在的T细胞途径，违反了连接识别的原则。我们将其称为帮助的受体表现途径。新抗原的AB肽还可以激活AB沉积部位的CD4 T细胞，其中APC通过FC受体或表面凝集素聚集AB。目前，我们对免疫系统如何调节CD4 T细胞与B细胞或其他由BCR和AB肽介导的其他APC之间的潜在危险相互作用有基本的了解。例如，IgG表面上是耐受性的。然而，治疗性mAB有时会引起内源性中和抗体反应，并具有潜在的临床健康危害。我们正在进行收养转移研究，以揭示使用基于菌株A遗传背景的小鼠中的模型对BCR和衍生肽的影响。我们最近的结果表明，T对B细胞的“受体表示”途径受到免疫调节的影响。由BCR衍生的肽介导的GC中的T-B相互作用阻碍了GC的参与和记忆B细胞的发育，并最终诱导T细胞中的难治状态。如果我们可以利用可用的遗传变化的C57BL/6（B6）小鼠，将大大增强这种新型免疫调节形式并探索AB免疫原性与耐受性问题的努力。因此，我们想在B6菌株中开发一个相应的模型，以便可以将遗传改变引入过继转移的B细胞，T细胞或受体小鼠中。 B6模型还将使我们能够在T细胞的收养受体中对CD4 T细胞反应进行抗体V区域肽的CD4 T细胞反应，这些受体注射了携带携带同源肽的mAb。为此，我们建议设计IG；轻链构造包含一个编码一个称为3K的肽的序列，该肽在I-AB的背景下对CD4 T细胞具有免疫原性（AIM 1）。这将使各种同种型的触觉特异性（NP）MAB能够产生针对3K细胞反应的未来研究； Ig，其免疫复合物以及3K； Ig的Glycan修饰形式。对于这些研究，我们将使用当前可用的12TCR TG小鼠作为3K特异性CD4 T细胞的来源。在AIM 2中，我们建议设计一个经过修改的3K；具有GFP报告基因的基因组构建体并生成相应的B6转基因小鼠。该小鼠将与敲入小鼠（B1-8）交叉，该小鼠从Ig中带有互补的靶向重链。 NP特异性mAb产生具有NP特异性3K的小鼠； B细胞。这些B细胞然后可以与3K特异性T细胞一起在受体表现的未来共同辅助转移遗传研究中使用。这项工作的产物将实现对BCR或分泌抗体中T细胞反应的机制的遗传解剖。该模型的预期结果将提高我们对BCR和AB中经常出现的新抗原的免疫调节的理解，并在免疫力期间定期出现给CD4 T细胞。缩写ANA：抗核抗体BAC：细菌人造染色体BCR：B细胞抗原受体CFP：氰荧光蛋白GFP：绿色荧光蛋白GC：生发蛋白GC：生发中心NP：（4-羟基-3-羟基 - 3-硝基高苯基）乙酰苯基乙酰基乙酰基乙酰基乙酰基乙酰基乙酰基乙酰基shm： 公共卫生相关性：针对抗体衍生的肽反应的T淋巴细胞在各种情况下都是潜在的健康危害。它们可能诱导患者对治疗性单克隆抗体的反应；当针对沉积在器官中的抗体时，它们可能会诱导器官炎症，并且可以刺激B细胞产生针对系统性自身免疫性疾病（例如全身性红斑狼疮）（SLE）等全身性自身免疫性疾病的抗体。该项目旨在开发独特的最先进的模型，使我们能够应用遗传学的力量，以揭示T细胞如何对调节潜在不良结果的抗体肽和机制反应。从该模型获得的信息将为疗法的合理设计提供可靠的基础，以控制或防止抗体和自身免疫性疾病，其中抗体和B细胞起着重要作用。