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.