ORIGINAL ARTICLE Neutralization of Membrane TNF, but Not Soluble TNF, Is Crucial for the Treatment of Experimental Colitis Cl  ementine Perrier, PhD,* ,† Gert de Hertogh, MD, PhD, ‡ Jonathan Cremer,* Severine Vermeire, MD, PhD,* Paul Rutgeerts, MD, PhD,* Gert Van Assche, MD, PhD,* David E. Szymkowski, PhD, § and Jan L. Ceuppens, MD, PhD † Background: Agents neutralizing membrane tumor necrosis factor (mTNF) and soluble TNF (sTNF) are widely used for the treatment of inflammatory bowel disease (IBD). Neutralization of mTNF, however, is associated with increased susceptibility to infectious diseases. The aim of this study was to determine whether neutralization of sTNF exclusively, by the use of a dominant negative mutant of TNF (XENP1595), could reduce the severity of colitis in mice. Methods: Colitis was induced in immunodeficient mice by transfer of CD45RB hi CD25 À T-cells. Once the disease had developed, mice were treated twice a week with XENP1595, phosphate-buffered saline (PBS), anti-TNF monoclonal antibody (mAb), or isotype control. The anti-TNF mAb blocks both mTNF and sTNF. Weights, disease activity index, macroscopic inflammation of the colon, and histological sections were eval- uated. T-cell populations from the colon were analyzed by flow cytometry. Results: Treatment of mice with XENP1595 did not change the course of the disease, whereas mice treated with anti-TNF mAb recovered weight soon after the first treatment dose. Inflammation in the colon was reduced in mice treated with anti-TNF mAb compared to isotype con- trol-treated animals. Mice treated with XENP1595 had a similar degree of inflammation in the colon as PBS-treated animals. The number of effector and regulatory T-cells in the colon remained unaffected by all treatments. Conclusions: Neutralization of sTNF exclusively was unable to induce remission in T-cell-mediated colitis, suggesting that neutralization of mTNF is crucial for the treatment of IBD. (Inflamm Bowel Dis 2012;000:000–000) Key Words: TNF, colitis, XENP1595, anti-TNF treatment, inflammatory bowel disease C rohn’s disease (CD) and ulcerative colitis (UC) are the two main forms of inflammatory bowel disease (IBD). They are complex and polygenic diseases with typi- cal remitting and relapsing courses. Although the etiology of IBD remains unclear, it is thought to result from a deregulation of the immunologic response to commensal bacteria of the gut in genetically susceptible subjects. 1 Currently, there are no curative treatments for IBD, but the introduction of biological therapies targeting tumor necrosis factor (TNF) has dramatically improved the qual- ity of treatment. So far, three anti-TNF agents are approved for the treatment of IBD: infliximab (Remicade; Centocor, Johnson & Johnson, Warren, NJ), adalimumab (Humira; Abbott, Abbott Park, IL), and certolizumab pegol (Cimzia; UCB, Brussels, Belgium). 2 Although the overall safety pro- file of these agents is good, anti-TNF agents also increase the risk of mycobacterial infections (reactivation of latent or new infections) and opportunistic infections. 3,4 More recently, skin lesions have been shown to develop in patients with anti-TNF treatments. 5 TNF is synthesized and presented on the cell surface as a membrane protein (mTNF) that can be cleaved to release soluble TNF (sTNF). Both forms of TNF can induce a signal via TNF receptors 1 and 2 (TNFR1 and TFNR2). In addition, mTNF can also act as a receptor that transmits an ‘‘outside-to-inside’’ signal back to the produc- ing cells after binding to its receptors, a process known as reverse signaling. 6 Clinical evidence and experimental stud- ies in rodents suggest that signaling by sTNF is associated with chronic and excessive inflammation in autoimmune diseases, 7–9 whereas signaling by mTNF plays an essential Additional Supporting Information may be found in the online version of this article. Received for publication April 12, 2012; Accepted April 30, 2012. From the *Department of Gastroenterology, Translational Research Center for Gastrointestinal Disorders (TARGID), KU Leuven, Leuven, Belgium, † Laboratory of Clinical Immunology, Department of Microbiology and Immunology, KU Leuven, Leuven, Belgium, ‡ Department of Morphology and Molecular Pathology, University Hospitals, Leuven, Belgium, § Xencor, Inc., Monrovia, California, USA. Partly supported by a grant from the Swiss Science Research Foundation (PBLAP3-129427/1) (to C.P.). Reprints: Cl ementine Perrier, Department of Gastroenterology, Translational Research Center for Gastrointestinal Disorders (TARGID), KU Leuven, O&N 1, Herestraat 49, box 811, 3000 Leuven, Belgium (e-mail: clementine.perrier@med. kuleuven.be). Copyright V C 2012 Crohn’s & Colitis Foundation of America, Inc. DOI 10.1002/ibd.23023 Published online in Wiley Online Library (wileyonlinelibrary. com). Inflamm Bowel Dis 1