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Title: Cloning and characterization of IA-2 specific autoantibodies in Type 1 diabetes
Author: Johnson, Carolyn Cromien
ISNI:       0000 0004 5991 1474
Awarding Body: King's College London
Current Institution: King's College London (University of London)
Date of Award: 2016
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The presence of one or more autoantibodies to characterized autoantigens (GAD, IA-2, (pro) insulin, Znt8) is commonly used to diagnose or predict Type 1 diabetes. 94% of patients express antibodies to at least one of the major islet autoantigens at the point of diagnosis: of these 70% have autoantibodies to IA-2, and 94% of patients can be identified by screening for multiple autoantibodies. Autoantibodies to IA-2 are known to appear in early or pre-diabetes, suggesting that the appearance of B cell reactivity to this antigen is an early event in the progression to disease. Recent studies have shown that Rituximab, a CD20 antibody, which depletes B cells, is able to preserve β-cell function in newly diagnosed patients, suggesting that B cell depletion may be an effective method of preventing β-cell destruction. Autoantigen specific B cell depletion in the NOD mouse, using a monoclonal anti-insulin antibody, restores immune tolerance to insulin. Similar strategies aimed at the specific depletion of islet autoantigen-reactive B cells may be effective in preventing diabetes progression in man. Development of such strategies requires knowledge of the nature of epitopes recognized by the B cell. The purpose of this study was to characterize specific targets of autoimmunity in type 1 diabetes using human monoclonal antibody fragments, developed from B cell lines and antigen specific B cells, alongside B cells within the inflammation in the type 1 diabetic pancreas. Antibody binding of sites on the IA-2 antigen was evaluated using NMR. Finally, site directed mutagenesis was used to determine specific residues important for antibody binding within known epitope regions. This study demonstrates the feasibility of generating monoclonal antibody fragments from monoclonal B cell lines, single B cells and archival material. This study also clarifies key regions required for autoantibody recognition of the juxtamembrane domain of the IA-2 protein, and provides a basis for future analysis of antibody binding of this antigen using NMR.
Supervisor: Christie, Michael Robert ; Conte, Maria Rosaria Sponsor: Not available
Qualification Name: Thesis (Ph.D.) Qualification Level: Doctoral
EThOS ID:  DOI: Not available