Cloning and genetics of human pemphigus autoantibodies

人天疱疮自身抗体的克隆和遗传学

• 批准号: 7069216
• 负责人: John R Stanley
• 金额: $ 34.87万
• 依托单位: UNIVERSITY OF PENNSYLVANIA
• 依托单位国家: 美国
• 财政年份: 2006
• 资助国家: 美国
• 起止时间: 2006-05-15 至 2011-04-30
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/7069216
• 关键词: antibody; autoantibody; clinical research; genetics; human; library; lighting; monoclonal antibody; pemphigus; peptides; phage display; skin

DESCRIPTION (provided by applicant): Pemphigus vulgaris and foliaceus are severe, sometimes life-threatening, autoimmune blistering skin diseases in which autoantibodies against desmoglein (Dsg) 3 and 1, desmosomal adhesion molecules, cause loss of keratinocyte cell adhesion. Ultimately, more specific treatments and diagnostic methods will require more detailed characterization of disease-causing autoantibodies. By determining the genetic properties and fine specificities of individual, i.e. monoclonal, antibodies (mAbs) from pemphigus patients, questions regarding common immunoglobulin variable segment gene usage, and shared pathogenic idiotypes and epitopes among patients can be addressed. A powerful molecular technology known as phage display will be used to clone and characterize human anti-Dsg autoantibodies. This technique will allow testing of the following hypotheses: a) mAbs against Dsg1 and Dsg3 from pemphigus patients are both pathogenic and non-pathogenic and both types can be isolated; b) There is genetically restricted heavy and light variable chain usage in anti-Dsg antibodies; c) Pathogenic mAbs bind a restricted set of epitopes at the amino terminus of Dsg3 and Dsg1; d) Pathogenic autoantibody idiotypes are shared among different patients; and e) Pathogenic idiotypes can be blocked by peptides mimicking the Dsg3 or Dsg1 pathogenic epitopes. Antibodies will be cloned from phage display libraries made from patients by selection on Dsg3 and Dsg1. The resulting human mAbs will be characterized by enzyme-linked immunosorbent assay, immunoflourescence and Western blotting. The mAbs will be tested for pathogenicity and then the genetics of the pathogenic and non-pathogenic antibodies will be compared. Phage peptide libraries will be screened with pathogenic mAbs. The respective epitopes on Dsgs will be mapped, and cross-reactive idiotypes (within and among patients) will be defined. The results of these experiments will greatly increase our knowledge of human autoantibodies in pemphigus. The project will: define how different human antibodies contribute to the pathology of this disease, develop valuable human monoclonal antibody reagents for other investigators studying these and other skin diseases, and will point to new ways to address targeted therapy and diagnosis to pathogenic antibodies in these potentially life-threatening diseases.

描述（由申请人提供）：pemphigus ufgaris和foliaus严重，有时威胁生命，自身免疫性的皮肤疾病，其中自身抗体对Desmoglein（DSG）3和1，Demosomal粘附分子，导致毛刺细胞细胞粘附的损失。最终，更具体的治疗方法和诊断方法将需要对引起疾病自身抗体的更详细的表征。通过确定个体的遗传特性和精细特异性，即来自Pemphigus患者的单克隆，抗体（MAB），有关常见的免疫球蛋白可变段基因使用的问题以及共享的致病性白痴型和患者的表现。一种称为噬菌体显示的强大分子技术将用于克隆和表征人类抗DSG自身抗体。该技术将允许对以下假设进行测试：a）来自Pemphigus患者的DSG1和DSG3的mABS既是致病性和非致病性，并且可以分离出两种类型； b）抗DSG抗体中有基因限制的重链和光变链使用； c）致病mAb在DSG3和DSG1的氨基末端结合了一组受限的表位； d）病原自身抗体白痴在不同的患者中共享； E）可以通过模仿DSG3或DSG1致病表位的肽来阻止致病性白痴。通过选择DSG3和DSG1的选择，将从患者制成的噬菌体展示文库中克隆抗体。由此产生的人mab将以酶联免疫吸附测定，免疫荧光和蛋白质印迹的特征。将对MAB进行致病性测试，然后比较致病性和非致病性抗体的遗传学。噬菌体肽库将用致病性mAB筛选。 DSG上的各自的表位将被映射，并将定义交叉反应性愚蠢（内部和患者之间）。这些实验的结果将大大增加我们对Pemphigus人类自身抗体的了解。该项目将：定义不同的人类抗体如何促进该疾病的病理学，为研究这些和其他皮肤疾病的其他研究人员开发有价值的人类单克隆抗体试剂，并将指出这些潜在的生命疾病中的致病性治疗和诊断的新方法。