title: Genetic and functional characterization of human pemphigus vulgaris monoclonal autoantibodies isolated by phage display.
abstract: Pemphigus is a life-threatening blistering disorder of the skin and mucous membranes caused by pathogenic autoantibodies to desmosomal adhesion proteins desmoglein 3 (Dsg3) and Dsg1. Mechanisms of antibody pathogenicity are difficult to characterize using polyclonal patient sera. Using antibody phage display, we have isolated repertoires of human anti-Dsg mAbs as single-chain variable-region fragments (scFvs) from a patient with active mucocutaneous pemphigus vulgaris. ScFv mAbs demonstrated binding to Dsg3 or Dsg1 alone, or both Dsg3 and Dsg1. Inhibition ELISA showed that the epitopes defined by these scFvs are blocked by autoantibodies from multiple pemphigus patients. Injection of scFvs into neonatal mice identified 2 pathogenic scFvs that caused blisters histologically similar to those observed in pemphigus patients. Similarly, these 2 scFvs, but not others, induced cell sheet dissociation of cultured human keratinocytes, indicating that both pathogenic and nonpathogenic antibodies were isolated. Genetic analysis of these mAbs showed restricted patterns of heavy and light chain gene usage, which were distinct for scFvs with different desmoglein-binding specificities. Detailed characterization of these pemphigus mAbs should lead to a better understanding of the immunopathogenesis of disease and to more specifically targeted therapeutic approaches.
authors.affiliation: Department of Dermatology, University of Pennsylvania, Philadelphia, Pennsylvania 19104, USA. aimee.payne@uphs.upenn.edu
authors.forename: Aimee S
authors.initials: AS
authors.lastname: Payne
authors.forename: Ken
authors.initials: K
authors.lastname: Ishii
authors.forename: Stephen
authors.initials: S
authors.lastname: Kacir
authors.forename: Chenyan
authors.initials: C
authors.lastname: Lin
authors.forename: Hong
authors.initials: H
authors.lastname: Li
authors.forename: Yasushi
authors.initials: Y
authors.lastname: Hanakawa
authors.forename: Kazuyuki
authors.initials: K
authors.lastname: Tsunoda
authors.forename: Masayuki
authors.initials: M
authors.lastname: Amagai
authors.forename: John R
authors.initials: JR
authors.lastname: Stanley
authors.forename: Don L
authors.initials: DL
authors.lastname: Siegel
mesh_terms: D000595:Amino Acid Sequence; D000818:Animals; D000831:Animals, Newborn; D000911:Antibodies, Monoclonal / Q000235:genetics* / Q000276:immunology*; D001323:Autoantibodies / Q000235:genetics* / Q000276:immunology*; D015820:Cadherins / Q000276:immunology; D002478:Cells, Cultured; D022801:Complementarity Determining Regions / Q000235:genetics; D051184:Desmoglein 3; D000078404:Epidermal Cells; D004817:Epidermis / Q000378:metabolism; D018604:Epitope Mapping; D006801:Humans; D007135:Immunoglobulin Variable Region / Q000235:genetics / Q000276:immunology; D015603:Keratinocytes / Q000166:cytology / Q000378:metabolism; D051379:Mice; D008969:Molecular Sequence Data; D010392:Pemphigus / Q000276:immunology*; D019151:Peptide Library; D011897:Random Allocation; D016415:Sequence Alignment
chemical_list: D000911:Antibodies, Monoclonal; D001323:Autoantibodies; D015820:Cadherins; D022801:Complementarity Determining Regions; C071894:DSG3 protein, human; D051184:Desmoglein 3; D007135:Immunoglobulin Variable Region; D019151:Peptide Library