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Epitope specificity determines pathogenicity and detectability in ANCA-associated vasculitis
Roth, A., J. Hess, J. Ooi, M. van Timmeren, E. Berg, C. Jennette, M. Burkart, S. Hogan, Y. Hu, W. Winnik, P. Nachman, P. Heeringa, R. Kitching, S. Holdsworth, C. Jennette, G. Preston, R. Falk, AND C. Stegeman. Epitope specificity determines pathogenicity and detectability in ANCA-associated vasculitis. JOURNAL OF CLINICAL INVESTIGATION. American Society for Clinical Investigation, Ann Arbor, MI, 123(4):1773-83, (2013).
ABSTRACT BACKGROUND Anti-neutrophil cytoplasmic autoantibodies (ANCA) specific for myeloperoxidase (MPO) or proteinase 3 (PR3) are detectable in >90% of patients with ANCA-associated vasculitis (AAV). ANCA titers do not correlate well with disease activity. In vivo and in vitro studies demonstrate that ANCA are pathogenic. We report an analysis of MPO-ANCA epitopes that explains the poor correlation between disease activity and ANCA titer, and failure to detect ANCA in some AAV patients. METHODS Epitope mapping entailed binding MPO-ANCA to native MPO, which protects epitopes from enzymatic digestion. Epitope peptides were eluted, identified by matrix-assisted laser desorption mass spectrometry, and mapped to MPO crystal structure. Samples were from active or remission AAV patients, and healthy controls. Reactivity with linear epitope peptides was assayed by ELISA. Pathogenic potential of ANCA was tested in vitro and in vivo. RESULTS 25 anti-MPO epitopes were detected and placed in 3 groups. Pathogenic epitopes reacted only with ANCA from patients with active disease. Nonpathogenic epitopes reacted with ANCA from patients in remission. Natural epitopes reacted at low levels with immunoglobulin from healthy controls. Antibodies to one pathogenic epitope were detectable when an epitope-masking ceruloplasmin fragment was removed in 8/10 AAV patients negative by conventional assays. CONCLUSIONS ANCA titers in clinical assays reflect a combination of pathogenic and nonpathogenic ANCA. Pathogenic ANCA correlate better with disease activity. IgG from patients with ANCA-negative AAV reacts with a pathogenic MPO epitope that is masked in serum by a ceruloplasmin fragment. Adjacency or natural and pathogenic epitopes implicates epitope spread in disease induction.
This manuscript resulted from advising a UNC student Aleeza Roth by Witold Winnik. No laboratory work was performed in EPA. The interaction with the student resulted in a refinement of mass spectrometry methodology that might benefit future EPA projects performed within the RCU Proteomics Research Core.
Record Details:Record Type: DOCUMENT (JOURNAL/PEER REVIEWED JOURNAL)
Organization:U.S. ENVIRONMENTAL PROTECTION AGENCY
OFFICE OF RESEARCH AND DEVELOPMENT
NATIONAL HEALTH AND ENVIRONMENTAL EFFECTS RESEARCH LAB
ASSOCIATE DIRECTOR FOR HEALTH
RESEARCH CORES UNIT