Shared molecular profiles of anti-PL12 autoantibodies in anti-synthetase syndrome

OBJECTIVES: Anti-PL12 autoantibodies are targeted against alanyl-tRNA synthetase, a cytosolic enzyme which plays a vital role in protein synthesis. Clinically, such antibodies are strongly associated with interstitial lung disease and confer a poor prognosis. To better understand their molecular composition, anti-PL12 immunoglobulin variable region subfamily expression and their mutational signatures were analysed by mass spectrometry (MS)-based proteomics to provide insights into personalized molecular diagnosis. METHODS: Patients with anti-PL12 autoantibodies were identified from routine myositis immunoblots. Serum anti-PL12 IgG autoantibodies were then purified from specific anti-PL12 precipitins using conventional counter-immunoelectrophoresis. Immunoglobulin (Ig) heavy and light-chains were separated by sodium dodecyl sulfate polyacrylamide gel electrophoresis (SDS-PAGE), digested by enzymes and profiled by MS sequencing for immunoglobulin variable region (IgV) subfamily usage and somatic hypermutation analysis. RESULTS: Ten patients were identified; seven females and three males with an average age of 65 years. All had lung disease, while two had concurrent myositis. MS sequencing of precipitating anti-PL12 IgGs revealed an oligoclonal repertoire with shared expression of IGHV3-23-encoded heavy chain and IGKV3-20- and IGKV1-27-encoded light chains. Furthermore, shared IgV somatic hypermutations were observed across unrelated patients in both heavy and light chain, in regions likely to be antigen interacting. CONCLUSION: Anti-PL12 autoantibodies among different patients show highly convergent sequences and variable region composition. The inter-individual similarity reveals the evolutionary process towards specific, presumably immuno-dominant, epitopes on PL12 during antigen-driven clonal selection. These unique anti-PL12 autoantibody molecular signatures have the potential to be valuable biomarkers when compared with the simple readout of conventional immunoassays.