Background:In fibrotic diseases, metabolic processes are altered with a tendency towards an anabolic state, which is partially reflected in serum. Circulating biomarkers for interstitial lung disease (ILD), the leading cause of death in systemic sclerosis (SSc), are still sparse and not established in routine care.Objectives:To assess the potential of serum metabolites as biomarkers for the presence and progression of SSc-ILD.Methods:Age and sex matched serum samples of SSc patients from the Zurich cohort and of healthy controls (HC) were analyzed. Progressive SSc-ILD was defined as either a relative decrease in forced vital capacity (FVC) >10%, a decrease in FVC of 5-9% and a concomitant decrease of carbon dioxide diffusion capacity >15%, or an increase of the extent of lung fibrosis on computed tomography from <20% to ≥20% compared to the last visit (mean follow-up interval = 14 months (range = 9-26)). Sera of HC, non-ILD SSc and stable vs. progressive SSc-ILD patients (n = 12 per group; total n = 48) were screened for 110 metabolites by targeted liquid chromatography coupled to tandem mass spectrometry (LC-MS/MS). Peak areas were analyzed with R 3.6. For univariate analysis, FDR-corrected one-way ANOVA was used. In multivariate group-wise partial least squares discriminant analysis (PLS-DA), variable importance in the projection (VIP) scores ≥2 were considered significant.Results:In total, 85 metabolites were detected. Univariate analysis of all groups were suggestive of changes for 1-methyladenosine, L-tryptophan, L-tyrosine, L-leucine and xanthosine (p = 0.077, 0.028, 0.077, 0.028 and 0.032, respectively). In PLS-DA, HCs and SSc patients differed in their levels of L-tyrosine and L-tryptophan, while levels of L-threonine, 3-aminoisobutyric acid, adenosine monophosphate and xanthosine were changed when comparing non-ILD and SSc-ILD patients. Receiver operating curve (ROC) analysis of significant metabolites from uni- and multivariate testing resulted in separation of SSc patients from HCs by L-tyrosine (area under the curve (AUC) = 0.81, 95% confidence interval (CI): 0.67-0.96), L-tryptophan (AUC = 0.86, CI: 0.75-0.97) and 1-methyladenosine (AUC = 0.82, CI: 0.71-0.94). Progressive SSc-ILD patients were separated from stable patients by their levels of L-isoleucine, L-leucine, adenosine monophosphate and xanthosine (AUC = 0.83, 0.85, 0.79 and 0.77; CI: 0.66-1.00, 0.70-1.00, 0.60-0.97 and 0.55-0.99, respectively). Validation of increased values of the branched-chain amino acids L-leucine and L-isoleucine in progressive SSc-ILD vs. stable ILD using an enzymatic assay resulted in similar results as LC-MS/MS analysis, with higher values detected in progressive vs. stable patients (mean = 286.5 and 235.5 nM, respectively; p = 0.005). In ROC analysis (AUC = 0.81, CI: 0.62-1.00), a cut-off value of 250.3 nM separated stable from progressive patients with a sensitivity of 72.7% and a specificity of 83.3%.Conclusion:This study in SSc(-ILD) patients suggested alterations in serum metabolite levels corresponding with their current state of disease, indicating the potential use of serum metabolites as discriminating biomarkers upon further confirmation in larger multicenter studies.Disclosure of Interests:Chantal Meier: None declared, Katrin Freiburghaus: None declared, Cédric Bovet: None declared, Janine Schniering: None declared, Oliver Distler Grant/research support from: Grants/Research support from Actelion, Bayer, Boehringer Ingelheim, Competitive Drug Development International Ltd. and Mitsubishi Tanabe; he also holds the issued Patent on mir-29 for the treatment of systemic sclerosis (US8247389, EP2331143)., Consultant of: Consultancy fees from Actelion, Acceleron Pharma, AnaMar, Bayer, Baecon Discovery, Blade Therapeutics, Boehringer, CSL Behring, Catenion, ChemomAb, Curzion Pharmaceuticals, Ergonex, Galapagos NV, GSK, Glenmark Pharmaceuticals, Inventiva, Italfarmaco, iQvia, medac, Medscape, Mitsubishi Tanabe Pharma, MSD, Roche, Sanofi and UCB, Speakers bureau: Speaker fees from Actelion, Bayer, Boehringer Ingelheim, Medscape, Pfizer and Roche, Christos Nakas: None declared, Britta Maurer Grant/research support from: AbbVie, Protagen, Novartis, congress support from Pfizer, Roche, Actelion, and MSD, Speakers bureau: Novartis
Serum metabolites as biomarkers in systemic sclerosis-associated interstitial lung disease
