Tear Protein Biomarkers for Ocular Mucous Membrane Pemphigoid Uncovered Using Targeted LC-MS/MS

Abstract

Mucous membrane pemphigoid (MMP) is a multisystemic rare autoimmune disease affecting the skin and mucous membranes. Ocular involvement is characterized by chronic conjunctival inflammation causing scar formation, leading to corneal opacification and vision loss. Conjunctival biopsies are currently used to confirm diagnosis, and the associated immunosuppression treatments prescribed can have serious consequences on patients. To address these challenges, a noninvasive approach to collect patient tears using untargeted and targeted liquid chromatography-tandem mass spectrometry (LC-MS/MS) analyses has been developed to identify a list of potential biomarkers of ocular MMP. Samples were collected on Schirmer strips and subjected to tryptic digestion and LC-MS/MS analysis. Three cohorts of patients were studied, and targeted LC-scheduled multiple reaction monitoring (LC-sMRM) methods were developed for the verification of putative biomarkers. The comparison of three groups of patients, those diagnosed with ocular MMP, non-ocular MMP, and lichen planus, another ocular cicatricial conjunctivitis disorder, yielded 56 biomarkers of interest. Proteins distinguishing MMP patients with and without ocular involvement were linked to the extracellular matrix, metabolism, and neutrophil degranulation. The comparison between MMP and lichen planus patients highlighted lower levels of metabolic enzymes in the latter. This study highlights the use of multiple patient cohorts and tailored targeted quantitative proteomics methods for the discovery of biomarkers for diagnostic and prognostic purposes.