P242 Immune-cell specific biomarker of early intestinal inflammation: Neutrophil elastase degraded fragment of type III collagen is elevated in patients with Inflammatory Bowel Disease

Abstract

Abstract Background Inflammatory Bowel Disease (IBD) is characterized by epithelial barrier injury of the gastrointestinal (GI) tract. The disease course encompasses abnormal immune responses and excessive secretion of proteases from immune cells. Neutrophils are the first to migrate into the inflamed interstitial matrix, where type III collagen is significantly deposited. Detection of mucosal inflammation in early stages is crucial to prevent cumulative clinical damage, as a delayed diagnosis can hinder effective treatment. Thus, we aimed to develop a biomarker that reflects early intestinal inflammation prior to it becoming medically evident; allowing us to distinguish patients that would respond to an anti-inflammatory treatment. Methods A competitive enzyme-linked immunosorbent assay targeting a human neutrophil elastase derived fragment of type III collagen was developed (C3-HNE). The validation was conducted by assessing its dilution recovery, matrix spiking, interference, and inter- and intra-assay variation. The biological relevance was evaluated in commercial samples, one preclinical, and two clinical studies. In the preclinical study, C3-HNE was measured in serum samples from a dextran sodium sulfate (DSS)-induced colitis rat model. In the commercial samples and clinical cohorts, C3-HNE was measured in serum from patients with IBD and healthy donors (HD). To compare the biomarker levels between groups, two-way ANOVA with a Fisher's LSD, and Mann-Whitney U-tests or Kruskal-Wallis (Dunn’s corrected) were conducted. Results The novel biomarker C3-HNE was successfully validated, resulting in a technically robust assay with acceptable technical parameters (Table 1). In the preclinical study, the analysis revealed significant difference in the C3-HNE levels between the control and DSS groups on day 4 (p=0.01) (Figure 1A). Additionally, C3-HNE was released earlier (day 1) in the timeline than the elevation of the current disease activity (DAI) score (day 7) (Figure 1A-B). In both clinical cohorts and commercial samples, C3-HNE levels were significantly higher in patients with Crohn's disease (CD), ulcerative colitis (UC) and IBD than in HD (p<0.01, p<0.05 and p<0.05) (Figure 1C-E). Based on the ROC analysis, the biomarker can significantly discriminate between HD and CD, UC or IBD (Table 1). Conclusion C3-HNE is elevated in patients with CD, UC and IBD compared with HD. Furthermore, C3-HNE reflects early stages of clinically apparent mucosal damage during experimental colitis. This biomarker holds the potential to identify early mucosal damage or acute inflammation in the GI tract; nevertheless, additional studies are needed to evaluate its clinical validity.