P094 Human neutrophil elastase shows increased elastinolytic activity in ulcerative colitis and contributes to loss of function of anti-TNF-alpha agents in ulcerative colitis
P. Giuffrida*1, K. Kok2, A. Di Sabatino1, P. Biancheri2, A. Vanoli3, G. Mazza4, R. Curciarello2, A. Pasini1, N. Aronico1, M. Pinzani4, G. R. Corazza1, T. T. MacDonald2
1Fondazione IRCCS Policlinico San Matteo, University of Pavia, First Department of Medicine, Pavia, Italy, 2Centre for Immunobiology, Blizard Institute, Barts and the London School of Medicine and Dentistry, London, United Kingdom, 3Fondazione IRCCS Policlinico San Matteo, University of Pavia, Department of Molecular Medicine, Pavia, Italy, 4Institute for Liver and Digestive Health, University College London, London, United Kingdom
Up to a third of ulcerative colitis (UC) patients are non-responders to anti-TNF-alpha agents, which act in the UC protease-rich inflamed mucosa. Human neutrophil elastase (HNE), whose main target in extracellular matrix is elastin, is a protease highly expressed in UC mucosa; however, the balance between HNE and its inhibitor elafin remains unclear. Our aims were to investigate the elastinolytic activity in UC, and determine if HNE degrades anti-TNF-alpha agents, thus rendering them ineffective.
Intestinal biopsies were collected from inflamed mucosa of UC and Crohn’s disease (CD) patients, and normal mucosa of controls. Mucosal elastin was assessed by ELISA, and mucosal elastinolytic activity was determined by elastase activity assay. Mucosal elafin was evaluated by immunofluorescence, immunoblotting, and ELISA. After co-incubation with HNE in the presence or in the absence of elafin, both integrity and TNF-alpha neutralising function of infliximab, adalimumab, and etanercept were studied by immunoblotting or using a nuclear factor-kappaB reporter cell line, respectively.
A significantly (p < 0.01) lower concentration of elastin was observed in UC (mean 17.3 ± 2.8 ng/mg) compared with CD (mean 37.6 ± 5.8 ng/mg) and controls (mean 44.4 ± 6.5 ng/mg). The elastinolytic activity was significantly (p < 0.05) increased in a time-dependent manner in UC mucosa compared with CD and control mucosa. Addition of elafin restored elastinolytic activity to normal levels. Mucosal elafin was significantly (p < 0.05) increased in UC (mean 2947 ± 754 pg/mg) in comparison with controls (mean 553 ± 754 ng/mg), with no difference between UC and CD (mean 1978 ± 774 pg/mg). HNE degraded infliximab and etanercept, but not adalimumab, in a dose-dependent manner, and these cleavages were inhibited by the addition of elafin. After HNE co-incubation, cleaved etanercept largely lost its ability to neutralise TNF-alpha, whereas infliximab and adalimumab functioned.
The increased activity of HNE in UC may affect therapeutic efficacy of anti-TNF-alpha agents in these patients, thus providing an explanation for the unresponsiveness of a certain number of UC patients to biologics.