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Inhibition of matrix metalloproteinase-14 in osteosarcoma cells by clodronate

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Author: Heikkilä, P. · Teronen, O. · Hirn, M.Y. · Sorsa, T. · Tervahartiala, T. · Salo, T. · Konttinen, Y.T. · Halttunen, T. · Moilanen, M. · Hanemaaijer, R. · Laitinen, M.
Source:Journal of Surgical Research, 1, 111, 45-52
Identifier: 237063
doi: doi10.1016/S0022-4804(03)00000-0
Keywords: Biology · Physiological Sciences · Bisphosphonate · Inhibition · MMP-2 · MT1-MMP · Osteosarcoma · clodronic acid · matrix metalloproteinase 14 · phorbol 13 acetate 12 myristate · article · cancer cell · cell activation · controlled study · cytotoxicity · down regulation · enzyme activation · enzyme activity · enzyme inhibition · human · human cell · immunohistochemistry · Northern blotting · osteosarcoma · priority journal · protein degradation · protein expression · Binding Sites · Blotting, Northern · Clodronic Acid · Collagenases · Culture Media, Conditioned · Enzyme Activation · Enzyme Inhibitors · Enzyme Precursors · Fluorescent Antibody Technique · Gelatinases · Humans · Matrix Metalloproteinase 13 · Matrix Metalloproteinase 2 · Matrix Metalloproteinases, Membrane-Associated · Metalloendopeptidases · Osteosarcoma · Recombinant Proteins · RNA, Messenger · Tetradecanoylphorbol Acetate · Tumor Cells, Cultured


Background. Bisphosphonates reduce the bone metastasis formation and angiogenesis but the exact molecular mechanisms involved are unclear. Progelatinase A (proMMP-2; 78 KDa) is activated up during the tumor spread and metastasis by a cell surface-associated matrix metalloproteinase (membrane-type matrix metalloproteinase [MT1-MMP] or MMP-14). Material and methods. We evaluated the effects of a bisphosphonate (clodronate) on MT1-MMP mRNA expression and protein production, catalytic activity and proteolytic activation of proMMP-2 by cultured human MG-63 osteosarcoma cells. Results. Clodronate, at therapeutically attainable noncytotoxic concentrations, dose-dependently inhibited phorbol myristic acetate (PMA)-induced proteolytic activation of proMMP-2 by human MG-63 osteosarcoma cells. Clodronate also downregulated the PMA-induced expression of MT1-MMP mRNA and protein production in human MG-63 osteosarcoma cells, as evidenced by Northern analysis and fluorescent immunohistochemistry. Furthermore, clodronate inhibited directly and dose-dependently MT1-MMP activity, and the MT1-MMP inhibition by clodronate was reduced in the presence of an increased (5 mM) Ca2+ concentrations when compared to physiological (1 mM) Ca2+ concentrations. Conclusion. We conclude that (1) the extracellular/cell-associated mechanism of bisphosphonate involves inhibition of MT1-MMP catalytic activity eventually by chelation, and that (2) intracellular mechanism involves downregulation of induced MT1-MMP mRNA and protein expression. The inhibition and downregulation of MT1-MMP by clodronate can be related to their ability to reduce MG-63 osteosarcoma cell invasion and spread. These findings may, at least in part, explain at molecular level the antitumor and antibone resorption activities of clodronate observed in clinical studies. © 2003 Elsevier Inc. All rights reserved.