Synthetic Neoepitopes of the Transglutaminase–Deamidated Gliadin Complex as Biomarkers for Diagnosing and Monitoring Celiac Disease

Background & Aims: Celiac disease (CeD) has characteristics of an autoimmune disease, such as increased antibody levels to tissue transglutaminase (tTG). However, assays to measure these biomarkers in blood samples do not identify patients with sufficient accuracy for diagnosis or monitoring of CeD. We aimed to discover biomarkers of CeD derived from neoepitopes of deamidated gliadin peptides (DGP) and tTG fragments and to determine if immune reactivity against these epitopes can identify patients with CeD with mucosal healing. Methods: We analyzed serum samples from 90 patients with biopsy-proven CeD and 79 healthy individuals (controls) for immune reactivity against the tTG-DGP complex (discovery cohort). A fluorescent peptide microarray platform was used to estimate the antibody-binding intensity of each synthesized tTG-DGP epitope. We validated our findings in 82 patients with newly diagnosed CeD and 217 controls. We tested the ability of our peptide panel to identify patients with mucosal healing (based on the histologic analysis) using serum samples from patients with treated and healed CeD (n = 85), patients with treated but unhealed CeD (n = 81; villous atrophy despite a adhering a gluten-free diet), patients with untreated CeD (n = 82) and disease controls (n = 27), villous atrophy without CeD), and healthy controls (n = 217). Data were analyzed using principal component analysis followed by machine learning and support vector machine modeling. Results: We identified 172 immunogenic epitopes of the tTG-DGP complex. We found significantly increased immune reactivity against these epitopes vs controls. In the both cohort, the set of neoepitopes derived from the tTG-DGP complex identified patients with CeD with 99% sensitivity and 100% specificity. Serum samples from patients with untreated CeD had the greatest mean antibody-binding intensity against the tTG-DGP complex (32.5 ± 16.4). The average antibody-binding intensity was significantly higher in serum from patients with treated but unhealed CeD mucosa (15.1 ± 7.5) than in patients with treated and healed CeD mucosa (5.5 ± 3.4) (P <.001). The assay identified patients with mucosa healing status with 84% sensitivity and 95% specificity. Conclusions: We identified immunogenic epitopes of the tTG-DGP complex, and found that an assay to measure the immune response to epitopes accurately identified patients with CeD, as well as patients with mucosal healing. This biomarker assay might be used in detection and monitoring of patients with CeD.