Abstract
Endogenous tissue inhibitors of metalloproteinases (TIMPs) have key roles in regulating physiological and pathological cellular processes1,2,3. Imitating the inhibitory molecular mechanisms of TIMPs while increasing selectivity has been a challenging but desired approach for antibody-based therapy4. TIMPs use hybrid protein-protein interactions to form an energetic bond with the catalytic metal ion, as well as with enzyme surface residues5,6,7. We used an innovative immunization strategy that exploits aspects of molecular mimicry to produce inhibitory antibodies that show TIMP-like binding mechanisms toward the activated forms of gelatinases (matrix metalloproteinases 2 and 9). Specifically, we immunized mice with a synthetic molecule that mimics the conserved structure of the metalloenzyme catalytic zinc-histidine complex residing within the enzyme active site. This immunization procedure yielded selective function-blocking monoclonal antibodies directed against the catalytic zinc-protein complex and enzyme surface conformational epitopes of endogenous gelatinases. The therapeutic potential of these antibodies has been demonstrated with relevant mouse models of inflammatory bowel disease8. Here we propose a general experimental strategy for generating inhibitory antibodies that effectively target the in vivo activity of dysregulated metalloproteinases by mimicking the mechanism employed by TIMPs.
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References
Gomis-Rüth, F.X. et al. Mechanism of inhibition of the human matrix metalloproteinase stromelysin-1 by TIMP-1. Nature 389, 77–81 (1997).
Cuniasse, P. et al. Future challenges facing the development of specific active-site-directed synthetic inhibitors of MMPs. Biochimie 87, 393–402 (2005).
Page-McCaw, A., Ewald, A.J. & Werb, Z. Matrix metalloproteinases and the regulation of tissue remodelling. Nat. Rev. Mol. Cell Biol. 8, 221–233 (2007).
Devy, L. et al. Selective inhibition of matrix metalloproteinase-14 blocks tumor growth, invasion and angiogenesis. Cancer Res. 69, 1517–1526 (2009).
Fernandez-Catalan, C. et al. Crystal structure of the complex formed by the membrane type 1-matrix metalloproteinase with the tissue inhibitor of metalloproteinases-2, the soluble progelatinase A receptor. EMBO J. 17, 5238–5248 (1998).
Grossman, M. et al. The intrinsic protein flexibility of endogenous protease inhibitor TIMP-1 controls its binding interface and affects its function. Biochemistry 49, 6184–6192 (2010).
Nagase, H., Visse, R. & Murphy, G. Structure and function of matrix metalloproteinases and TIMPs. Cardiovasc. Res. 69, 562–573 (2006).
Garg, P. et al. Matrix metalloproteinase-9–mediated tissue injury overrides the protective effect of matrix metalloproteinase-2 during colitis. Am. J. Physiol. Gastrointest. Liver Physiol. 296, G175–G184 (2009).
Gomis-Rüth, F.X. Catalytic domain architecture of metzincin metalloproteases. J. Biol. Chem. 284, 15353–15357 (2009).
Rowsell, S. et al. Crystal structure of human MMP9 in complex with a reverse hydroxamate inhibitor. J. Mol. Biol. 319, 173–181 (2002).
Bertini, I. et al. Conformational variability of matrix metalloproteinases: beyond a single 3D structure. Proc. Natl. Acad. Sci. USA 102, 5334–5339 (2005).
MacCallum, R.M., Martin, A.C.R. & Thornton, J.M. Antibody-antigen interactions: contact analysis and binding site topography. J. Mol. Biol. 262, 732–745 (1996).
Berchanski, A., Shapira, B. & Eisenstein, M. Hydrophobic complementarity in protein-protein docking. Proteins 56, 130–142 (2004).
Abraham, C. & Cho, J.H. Inflammatory bowel disease. N. Engl. J. Med. 361, 2066–2078 (2009).
Gao, Q. et al. Expression of matrix metalloproteinases-2 and -9 in intestinal tissue of patients with inflammatory bowel diseases. Dig. Liver Dis. 37, 584–592 (2005).
Kirkegaard, T., Hansen, A., Bruun, E. & Brynskov, J. Expression and localisation of matrix metalloproteinases and their natural inhibitors in fistulae of patients with Crohn's disease. Gut 53, 701–709 (2004).
Baugh, M.D. et al. Matrix metalloproteinase levels are elevated in inflammatory bowel disease. Gastroenterology 117, 814–822 (1999).
Santana, A. et al. Attenuation of dextran sodium sulphate induced colitis in matrix metalloproteinase-9 deficient mice. World J. Gastroenterol. 12, 6464–6472 (2006).
Castaneda, F.E. et al. Targeted deletion of metalloproteinase 9 attenuates experimental colitis in mice: central role of epithelial-derived MMP. Gastroenterology 129, 1991–2008 (2005).
Monteleone, G., Fina, D., Caruso, R. & Pallone, F. New mediators of immunity and inflammation in inflammatory bowel disease. Curr. Opin. Gastroenterol. 22, 361–364 (2006).
Garg, P. et al. Selective ablation of matrix metalloproteinase-2 exacerbates experimental colitis: contrasting role of gelatinases in the pathogenesis of colitis. J. Immunol. 177, 4103–4112 (2006).
Korzenik, J.R. & Podolsky, D.K. Evolving knowledge and therapy of inflammatory bowel disease. Nat. Rev. Drug Discov. 5, 197–209 (2006).
Aychek, T., Vandoorne, K., Brenner, O., Jung, S. & Neeman, M. Quantitative analysis of intravenously administered contrast media reveals changes in vascular barrier functions in a murine colitis model. Magn. Reson. Med. 66, 235–243 (2011).
Overall, C.M. & Kleifeld, O. Tumour microenvironment—opinion: validating matrix metalloproteinases as drug targets and anti-targets for cancer therapy. Nat. Rev. Cancer 6, 227–239 (2006).
Coussens, L.M., Fingleton, B. & Matrisian, L.M. Matrix metalloproteinase inhibitors and cancer: trials and tribulations. Science 295, 2387–2392 (2002).
Ikejiri, M. et al. Potent mechanism-based inhibitors for matrix metalloproteinases. J. Biol. Chem. 280, 33992–34002 (2005).
Johnson, J.L. et al. A selective matrix metalloproteinase-12 inhibitor retards atherosclerotic plaque development in apolipoprotein E-knockout mice. Arterioscler. Thromb. Vasc. Biol. 31, 528–535 (2011).
Sela-Passwell, N., Trahtenherts, A., Krüger, A. & Sagi, I. New opportunities in drug design of metalloproteinase inhibitors: combination between structure-function experimental approaches and systems biology. Expert Opin. Drug Discov. 6, 527–542 (2011).
Devel, L. et al. Third generation of matrix metalloprotease inhibitors: gain in selectivity by targeting the depth of the S1′ cavity. Biochimie 92, 1501–1508 (2010).
Morrison, C.J., Butler, G.S., Rodriguez, D. & Overall, C.M. Matrix metalloproteinase proteomics: substrates, targets, and therapy. Curr. Opin. Cell Biol. 21, 645–653 (2009).
Krüger, A., Kates, R.E. & Edwards, D.R. Avoiding spam in the proteolytic internet: future strategies for anti-metastatic MMP inhibition. Biochim. Biophys. Acta 1803, 95–102 (2010).
Morgunova, E. et al. Structure of human pro-matrix metalloproteinase-2: activation mechanism revealed. Science 284, 1667–1670 (1999).
Elkins, P.A. et al. Structure of the C-terminally truncated human ProMMP9, a gelatin-binding matrix metalloproteinase. Acta Crystallogr. D Biol. Crystallogr. 58, 1182–1192 (2002).
Harlow, E.D.P.L. Antibodies: A Laboratory Manual. Ch. 6 (Cold Spring Harbor Laboratory Press, 1998).
Knight, C.G., Willenbrock, F. & Murphy, G. A novel coumarin-labelled peptide for sensitive continuous assays of the matrix metalloproteinases. FEBS Lett. 296, 263–266 (1992).
Solomon, A. et al. Pronounced diversity in electronic and chemical properties between the catalytic zinc sites of tumor necrosis factor-α–converting enzyme and matrix metalloproteinases despite their high structural similarity. J. Biol. Chem. 279, 31646–31654 (2004).
Wirtz, S., Neufert, C., Weigmann, B. & Neurath, M.F. Chemically induced mouse models of intestinal inflammation. Nat. Protoc. 2, 541–546 (2007).
Acknowledgements
We would like to thank D. Tawfik and I. Shachar for critical reading of the manuscript, K. Vyacheslav for assistance in optical imaging data collection and analysis, and C. Power, O. Leger and O. Lightner for technical support in antibody production. This work was supported by the Israel Science Foundation, the Minerva Foundation, the Helmsley Foundation, the Kimmelman Center at the Weizmann Institute and funds from C. Berall and L. Berall, M. Steinberg and C. Adelson. I.S. is the Incumbent of the Maurizio Pontecorvo Professorial Chair.
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N.S.-P. designed, performed and analyzed the antibody biophysical analyses, animal experiments and wrote the manuscript. R.K., R.A.-Y. and A.S. designed, synthesized and analyzed the Zn-tripod antigen. R.M. performed the animal experiments. O.D. and H.R. collected X-ray diffraction data, O.D. supervised crystallization experiments and determined the crystal structure. M.E. performed the computational docking analysis. O.B. performed pathological analysis of tissue samples. T.S. assisted with figures and experimental design. T.D. assisted and provided technical support with antibody generation. I.S. designed the experiments, analyzed the data and wrote the manuscript.
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Sela-Passwell, N., Kikkeri, R., Dym, O. et al. Antibodies targeting the catalytic zinc complex of activated matrix metalloproteinases show therapeutic potential. Nat Med 18, 143–147 (2012). https://doi.org/10.1038/nm.2582
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DOI: https://doi.org/10.1038/nm.2582
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