Objective Identify autoantibodies in anti-Ro/SS-A negative primary Sjögren’s syndrome (SS).
Methods This is a proof-of-concept, case-control study of SS, healthy (HC) and other disease (OD) controls. A discovery dataset of plasma samples (n=30 SS, n=15 HC) was tested on human proteome arrays containing 19 500 proteins. A validation dataset of plasma and stimulated parotid saliva from additional SS cases (n=46 anti-Ro+, n=50 anti-Ro–), HC (n=42) and OD (n=54) was tested on custom arrays containing 74 proteins. For each protein, the mean+3 SD of the HC value defined the positivity threshold. Differences from HC were determined by Fisher’s exact test and random forest machine learning using 2/3 of the validation dataset for training and 1/3 for testing. Applicability of the results was explored in an independent rheumatology practice cohort (n=38 Ro+, n=36 Ro–, n=10 HC). Relationships among antigens were explored using Search Tool for the Retrieval of Interacting Genes/Proteins (STRING) interactome analysis.
Results Ro+ SS parotid saliva contained autoantibodies binding to Ro60, Ro52, La/SS-B and muscarinic receptor 5. SS plasma contained 12 novel autoantibody specificities, 11 of which were detected in both the discovery and validation datasets. Binding to ≥1 of the novel antigens identified 54% of Ro– SS and 37% of Ro+ SS cases, with 100% specificity in both groups. Machine learning identified 30 novel specificities showing receiver operating characteristic area under the curve of 0.79 (95% CI 0.64 to 0.93) for identifying Ro– SS. Sera from Ro– cases of an independent cohort bound 17 of the non-canonical antigens. Antigenic targets in both Ro+ and Ro– SS were part of leukaemia cell, ubiquitin conjugation and antiviral defence pathways.
Conclusion We identified antigenic targets of the autoantibody response in SS that may be useful for identifying up to half of Ro seronegative SS cases.
- Sjogren's syndrome
- autoimmune diseases
Data availability statement
Data are available on reasonable request.
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Handling editor Josef S Smolen
Deceased KAK deceased on 02 May 2022
Contributors SL and ADF conceived the study. ADF is the guarantor of the study. SL, CL-D, CL and KAK (deceased in May 2022) optimised procedures and collected the data. SL, ADF, BK, CG, JDW and CL analysed the data. CP-F, CL-D and LR collected and processed the samples. AR, LR, RHAS and GP performed clinical evaluations. RCA and GP provided MS patient samples. JMG and JJ recruited healthy controls. ANB, SAR and ED contributed samples and data. SL and ADF wrote the manuscript. All authors (except KAK) reviewed and edited the manuscript. ADF is the guarantor of the study.
Funding Research reported in this publication was supported by National Institutes of Health grants R01AR074310 (ADF), T32AI007633 (SL), R01AR073855 (CJL), R01AR065953 (CJL), P30AR073750 (JAJ, JMG), U54GM104938 (JAJ, JDW), P50AR060804 (Oklahoma Sjögren’s Syndrome Center of Research Translation), UM1AI144292 (JMG), R01DE012354 (ANB) and P30AR053503 (Johns Hopkins Rheumatic Disease Research Core Center). Other supporting grants were from the Rheumatology Research Foundation Innovative Grant Award (ADF), Jerome L. Greene Foundation, Presbyterian Health Foundation and Sjögren’s Foundation.
Disclaimer The content is solely the responsibility of the authors and does not necessarily reflect the official views of the National Institutes of Health.
Competing interests ADF and CJL have received grant support from Janssen Research and Development. AND has received consulting fees from Bristol-Myers Squibb.
Patient and public involvement Patients and/or the public were not involved in the design, or conduct, or reporting, or dissemination plans of this research.
Provenance and peer review Not commissioned; externally peer reviewed.
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