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.
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