title: Proteolytic cleavage and loss of function of biologic agents that neutralize tumor necrosis factor in the mucosa of patients with inflammatory bowel disease.
abstract: OBJECTIVE
  Many patients with inflammatory bowel disease (IBD) fail to respond to anti-tumor necrosis factor (TNF) agents such as infliximab and adalimumab, and etanercept is not effective for treatment of Crohn's disease. Activated matrix metalloproteinase 3 (MMP3) and MMP12, which are increased in inflamed mucosa of patients with IBD, have a wide range of substrates, including IgG1. TNF-neutralizing agents act in inflamed tissues; we investigated the effects of MMP3, MMP12, and mucosal proteins from IBD patients on these drugs.
  
  
  METHODS
  Biopsy specimens from inflamed colon of 8 patients with Crohn's disease and 8 patients with ulcerative colitis, and from normal colon of 8 healthy individuals (controls), were analyzed histologically, or homogenized and proteins were extracted. We also analyzed sera from 29 patients with active Crohn's disease and 33 patients with active ulcerative colitis who were candidates to receive infliximab treatment. Infliximab, adalimumab, and etanercept were incubated with mucosal homogenates from patients with IBD or activated recombinant human MMP3 or MMP12 and analyzed on immunoblots or in luciferase reporter assays designed to measure TNF activity. IgG cleaved by MMP3 or MMP12 and antihinge autoantibodies against neo-epitopes on cleaved IgG were measured in sera from IBD patients who subsequently responded (clinical remission and complete mucosal healing) or did not respond to infliximab.
  
  
  RESULTS
  MMP3 and MMP12 cleaved infliximab, adalimumab, and etanercept, releasing a 32-kilodalton Fc monomer. After MMP degradation, infliximab and adalimumab functioned as F(ab')2 fragments, whereas cleaved etanercept lost its ability to neutralize TNF. Proteins from the mucosa of patients with IBD reduced the integrity and function of infliximab, adalimumab, and etanercept. TNF-neutralizing function was restored after incubation of the drugs with MMP inhibitors. Serum levels of endogenous IgG cleaved by MMP3 and MMP12, and antihinge autoantibodies against neo-epitopes of cleaved IgG, were higher in patients who did not respond to treatment vs responders.
  
  
  CONCLUSIONS
  Proteolytic degradation may contribute to the nonresponsiveness of patients with IBD to anti-TNF agents.
authors.affiliation: Centre for Immunobiology, Blizard Institute, Barts and The London School of Medicine and Dentistry, London, United Kingdom; Department of Internal Medicine, S. Matteo Hospital, University of Pavia, Pavia, Italy.
authors.forename: Paolo
authors.initials: P
authors.lastname: Biancheri
authors.affiliation: Biologics Research, Janssen Research and Development, LLC, Spring House, Pennsylvania.
authors.forename: Randall J
authors.initials: RJ
authors.lastname: Brezski
authors.affiliation: Department of Internal Medicine, S. Matteo Hospital, University of Pavia, Pavia, Italy.
authors.forename: Antonio
authors.initials: A
authors.lastname: Di Sabatino
authors.affiliation: Biologics Research, Janssen Research and Development, LLC, Spring House, Pennsylvania.
authors.forename: Allison R
authors.initials: AR
authors.lastname: Greenplate
authors.affiliation: Biologics Research, Janssen Research and Development, LLC, Spring House, Pennsylvania.
authors.forename: Keri L
authors.initials: KL
authors.lastname: Soring
authors.affiliation: Department of Internal Medicine, S. Matteo Hospital, University of Pavia, Pavia, Italy.
authors.forename: Gino R
authors.initials: GR
authors.lastname: Corazza
authors.affiliation: Centre for Immunobiology, Blizard Institute, Barts and The London School of Medicine and Dentistry, London, United Kingdom.
authors.forename: Klaartje B
authors.initials: KB
authors.lastname: Kok
authors.affiliation: Department of Internal Medicine, S. Matteo Hospital, University of Pavia, Pavia, Italy.
authors.forename: Laura
authors.initials: L
authors.lastname: Rovedatti
authors.affiliation: Centre for Immunobiology, Blizard Institute, Barts and The London School of Medicine and Dentistry, London, United Kingdom.
authors.forename: Anna
authors.initials: A
authors.lastname: Vossenkämper
authors.affiliation: Centre for Immunobiology, Blizard Institute, Barts and The London School of Medicine and Dentistry, London, United Kingdom.
authors.forename: Nadja
authors.initials: N
authors.lastname: Ahmad
authors.affiliation: Centre for Immunobiology, Blizard Institute, Barts and The London School of Medicine and Dentistry, London, United Kingdom.
authors.forename: Susanne A
authors.initials: SA
authors.lastname: Snoek
authors.affiliation: Department of Gastroenterology, University Hospital Gasthuisberg, Leuven, Belgium.
authors.forename: Severine
authors.initials: S
authors.lastname: Vermeire
authors.affiliation: Department of Gastroenterology, University Hospital Gasthuisberg, Leuven, Belgium.
authors.forename: Paul
authors.initials: P
authors.lastname: Rutgeerts
authors.affiliation: Biologics Research, Janssen Research and Development, LLC, Spring House, Pennsylvania.
authors.forename: Robert E
authors.initials: RE
authors.lastname: Jordan
authors.affiliation: Centre for Immunobiology, Blizard Institute, Barts and The London School of Medicine and Dentistry, London, United Kingdom. Electronic address: t.t.macdonald@qmul.ac.uk.
authors.forename: Thomas T
authors.initials: TT
authors.lastname: MacDonald
mesh_terms: D000068879:Adalimumab / Q000378:metabolism; D061067:Antibodies, Monoclonal, Humanized / Q000378:metabolism; D001685:Biological Factors / Q000378:metabolism* / Q000494:pharmacology; D001706:Biopsy; D016022:Case-Control Studies; D003093:Colitis, Ulcerative / Q000188:drug therapy / Q000276:immunology / Q000378:metabolism; D003106:Colon / Q000276:immunology / Q000378:metabolism / Q000473:pathology; D003424:Crohn Disease / Q000188:drug therapy / Q000276:immunology / Q000378:metabolism; D000939:Epitopes / Q000378:metabolism; D000068800:Etanercept / Q000378:metabolism; D005260:Female; D006801:Humans; D015151:Immunoblotting / Q000379:methods; D007074:Immunoglobulin G / Q000378:metabolism*; D015212:Inflammatory Bowel Diseases / Q000188:drug therapy / Q000276:immunology / Q000378:metabolism*; D000069285:Infliximab / Q000378:metabolism; D007413:Intestinal Mucosa / Q000187:drug effects / Q000276:immunology / Q000378:metabolism*; D008297:Male; D053508:Matrix Metalloproteinase 12 / Q000378:metabolism; D019278:Matrix Metalloproteinase 3 / Q000378:metabolism; D008875:Middle Aged; D059748:Proteolysis / Q000187:drug effects*; D014409:Tumor Necrosis Factor-alpha / Q000378:metabolism*
chemical_list: D061067:Antibodies, Monoclonal, Humanized; D001685:Biological Factors; D000939:Epitopes; D007074:Immunoglobulin G; D014409:Tumor Necrosis Factor-alpha; D000069285:Infliximab; C511768:MMP3 protein, human; D019278:Matrix Metalloproteinase 3; C507122:MMP12 protein, human; D053508:Matrix Metalloproteinase 12; D000068879:Adalimumab; D000068800:Etanercept
keywords: CD; Inflammation; Therapy; UC