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DOP049 Inflammatory dyskinesis: defects of NF-κB dynamic behaviour as a new potential biomarker for personalized medicine in inflammatory bowel disease

Sellge G.*1, Papoutsopoulou S.2, Verdier J.1, Trautwein C.1, Bergey F.3, Burkitt M.4, Burkitt M.4, Sheibani R.5, Pierik M.6, Jonkers D.6, White M.2, Paszek P.2, Martins dos Santos V.3, Rosenstiel P.5, Müller W.2, Probert C.4 SysmedIBD Consortium2

1University Hospital RWTH Aachen, Department of Internal Medicine III, Aachen, Germany 2University of Manchester, Faculty of Life Sciences, Manchester, United Kingdom 3LifeGlimmer GmbH, Berlin, Germany 4University of Liverpool, Cellular and Molecular Physiology, Liverpool, United Kingdom 5Kiel University, Institute of Clinical Molecular Biology, Kiel, Germany 6Maastricht University Medical Centre, Division Gastroenterology-Hepatology, Maastricht, Netherlands

Background

To improve stratification and optimize treatment strategies for individuals with inflammatory bowel diseases (IBD) new precision medicine approaches based on innovative biomarkers are needed. Most current biomarkers are collected at fixed time points, while biological processes are intrinsically dynamic. The NF-κB family of transcription factors regulate immune responses to multiple signals. In this study, two methods were combined to monitor dynamic NF-κB activation in response to cytokines and LPS in immune cells of control and IBD patients.

Methods

Peripheral blood mononuclear cells (PBMC) were donated by controls (n=40), patients with Crohn's disease (CD, n=50) or with ulcerative colitis (UC, n=30). NF-κB activation (phosphorylation of p65 and degradation of IκBα) and transcriptional activity of NF-κB were assessed by flow cytometry in PBMC-derived immune cells stimulated by TNF, LPS and MDP. Independently, PBMC-derived macrophages were transduced with a lentiviral construct encoding an NF-κB luciferase reporter. Luciferase activity in response to LPS was quantified. NF-κB dynamic responses were analysed using the area under the curve (AUC), the peak intensity and time of the response.

Results

NF-κB activation and activity respectively peak 15mn and 2h following stimulation on average. UC patients displayed significantly lower delayed NF-κB activation as compared to controls in CD4, CD8 and NK cells while AUC for luciferase activity was significantly lower in both UC and CD patients as compared to controls in PBMC-derived macrophages. Interestingly, 10/26 CD patients exhibited either hyperactive or suppressed NF-κB activities. compared with control and CD patients. Similar results were obtained with PBMCs collected in two different clinical centres, ruling out a batch effect.

Figure 1. NF-κB responses in immune cells of IBD patients and controls.

A. Methods B. Hypothetical NF-κB response profiles C. Luciferase activity in LPS-stimulated macrophages D. Area under the curve of log2 luciferase activity E. NF-κB response curves of TNF-stimulated CD4+ cells obtained by flow cytometry. F. Peak intensity and time of NF-κB responses in different cell types upon TNF stimulation. (* p<0.05)

Conclusion

These early findings suggest that NF-κB dynamic response may provide a new method for stratification of IBD patients. It remains to be seen whether this stratification correlates with other clinically relevant phenotypes