Potential Biomarkers for Sjögren’s Syndrome Detected in Saliva Using High–Resolution Mass Spectrometry and Bioinformatics

Abstract

Purpose: : Sjögren’s syndrome (SS) is considered to be the most common and under–diagnosed autoimmune disease where a delay of 5 to 8 years after symptom onset before a diagnosis is made is not uncommon. This is largely due to the fact that the diagnostic approach to SS is rather complicated and must include multiple subjective and objective criteria. Thus, the purpose of the current studies was to determine if saliva contains biomarkers that can be used as diagnosis/prognosis tools for SS.

Methods: : A total of 27 SS (14 primary and 13 secondary) patients and 27 age–matched healthy controls were recruited for these studies. Unstimulated submandibular glands saliva was collected from the Wharton’s duct using a suction device. Two µl of salvia were diluted in 180 µl of 0.2% trifluoroacetic acid and processed for mass spectrometry analyses utilizing a Millipore ZipPlate. Mass spectra were acquired on a prOTOF 2000 matrix–assisted laser desorption/ionization orthogonal time of flight (MALDI–O–TOF) mass spectrometer in the molecular weight range of 750 – 12,000 Da. Raw data were exported and sent for analysis by Predictive Diagnostics Inc. (Vacaville, CA) who utilizes proprietary bioinformatics (Biomarker AMplification Filter, BAMF^TM) tools to identify biomarkers.

Results: : BAMF^TM analysis resulted in several classification models built and several biomarkers were identified. A model based on 7 putative biomarkers yielded a sensitivity of 97.5%, specificity of 97.8% and an accuracy of 97.6%. Biomarkers identified were detected at m/z values 902.48, 2,407.30, 2,912.56, 3,655.78, 3,803.38, 4,281.14, and 5,942.00. Another set of 7 putative biomarkers was identified in another classification model with a sensitivity of 85.0%, specificity of 91.3%, and an accuracy of 88.1%. Biomarkers identified in this model were detected at m/z values 902.48, 1,479.76, 2,536.16, 3,655.78, 3,803.38, 4,930.28, and 5,843.94. Three of the biomarkers (underlined) were common to both models.

Conclusions: : Biomarkers detected in saliva by high–resolution mass spectrometry offer the potential to serve as diagnostic/prognostic tools for SS. Further studies using a larger population of SS patients are warranted.

Keywords: cornea: tears/tear film/dry eye • lacrimal gland • cornea: basic science