Peptide Inhibition of Catalytic and Noncatalytic Activities of Matrix Metalloproteinase-9 Blocks Tumor Cell Migration and Invasion

Migration of invasive cells appears to be dependent on matrix metalloproteinases (MMPs) anchored on the cell surface through integrins. We have previously demonstrated an interaction between the integrin α-subunit I domain and the catalytic domain of MMP-9. We now show that there is also an interaction between the integrin β subunit and MMP-9. Using phage display, we have developed MMP-9 inhibitors that bind either to the MMP-9 catalytic domain, the collagen binding domain, or the C-terminal hemopexin-like domain. The C-terminal domain-binding peptide mimics an activation epitope in the stalk of the integrin β chain and inhibits the association of MMP-9 C-terminal domain with αVβ5 integrin. Unlike other MMP-9 binding peptides, it does not directly inhibit catalytic activity of MMP-9, but still prevents proenzyme activation and cell migration in vitro and tumor xenograft growth in vivo. We also find an association between MMP-9 and urokinase-plasminogen activator receptor and find that urokinase-plasminogen activator receptor is cleaved by MMP-9. Collectively, we have defined molecular details for several interactions mediated by the different MMP-9 domains. Migration of invasive cells appears to be dependent on matrix metalloproteinases (MMPs) anchored on the cell surface through integrins. We have previously demonstrated an interaction between the integrin α-subunit I domain and the catalytic domain of MMP-9. We now show that there is also an interaction between the integrin β subunit and MMP-9. Using phage display, we have developed MMP-9 inhibitors that bind either to the MMP-9 catalytic domain, the collagen binding domain, or the C-terminal hemopexin-like domain. The C-terminal domain-binding peptide mimics an activation epitope in the stalk of the integrin β chain and inhibits the association of MMP-9 C-terminal domain with αVβ5 integrin. Unlike other MMP-9 binding peptides, it does not directly inhibit catalytic activity of MMP-9, but still prevents proenzyme activation and cell migration in vitro and tumor xenograft growth in vivo. We also find an association between MMP-9 and urokinase-plasminogen activator receptor and find that urokinase-plasminogen activator receptor is cleaved by MMP-9. Collectively, we have defined molecular details for several interactions mediated by the different MMP-9 domains. Matrix metalloproteinases 2 and 9 (MMP-2 and -9), 1The abbreviations used are: MMP, matrix metalloproteinase; CBD, collagen binding domain; C domain, C-terminal hemopexin-like domain; pro-MMP-9-ΔHC, pro-MMP-9 lacking the hinge region and the C domain; TIMP, tissue inhibitor of MMPs; uPA, urokinase-plasminogen activator; uPAR, uPA receptor; CTT, CTTHWGFTLC peptide; CRV, CRVYGPYLLC peptide; PPC, ADGACGYGRFSPPCGAAG peptide; DDGW, ADGACILWMDDGWCGAAG peptide, PDBu, phorbol ester; BSA, bovine serum albumin; PBS, phosphate-buffered saline; PBST, PBS plus 0.05% Tween 20; I-EGF, integrin-epidermal growth factor; GST, glutathione S-transferase. also known as gelatinases, play an important role in cell migration and tissue remodeling during development but also in pathological conditions such as inflammation and cancer (1Sternlicht M.D. Werb Z. Annu. Rev. Cell Dev. Biol. 2001; 17: 463-516Crossref PubMed Scopus (3242) Google Scholar). We have identified a highly selective peptide inhibitor of gelatinases, CTTHWGFTLC (CTT) by phage display (2Koivunen E. Arap W. Valtanen H. Rainisalo A. Medina O.P. Heikkila P. Kantor C. Gahmberg C.G. Salo T. Konttinen Y.T. Sorsa T. Ruoslahti E. Pasqualini R. Nat. Biotechnol. 1999; 17: 768-774Crossref PubMed Scopus (514) Google Scholar), whereas others have developed gelatinase-selective small molecule inhibitors (3Tamura Y. Watanabe F. Nakatani T. Yasui K. Fuji M. Komurasaki T. Tsuzuki H. Maekawa R. Yoshioka T. Kawada K. Sugita K. Ohtani M. J. Med. Chem. 1998; 41: 640-649Crossref PubMed Scopus (276) Google Scholar) to specifically target these enzymes. The unique structural feature of the gelatinases is the collagen-binding domain (CBD) within the catalytic domain (4Overall C.M. Mol. Biotechnol. 2002; 22: 51-86Crossref PubMed Google Scholar). The CBD is composed of three fibronectin type II repeats and is an intriguing target to develop gelatinase-specific compounds. Like most MMPs, the gelatinases also contain a C-terminal hemopexin/vitronectin-like domain (C domain or PEX), which contains the binding site for tissue inhibitors for matrix metalloproteinases (TIMPs) and is responsible for the dimerization of MMP-9 (5Cha H. Kopetzki E. Huber R. Lanzendorfer M. Brandstetter H. J. Mol. Biol. 2002; 320: 1065-1079Crossref PubMed Scopus (105) Google Scholar). Although MMP-2 and MMP-9 are closely related enzymes, they do have differences in the regulation of expression, activation, and glycosylation and in substrate selectivity (1Sternlicht M.D. Werb Z. Annu. Rev. Cell Dev. Biol. 2001; 17: 463-516Crossref PubMed Scopus (3242) Google Scholar, 4Overall C.M. Mol. Biotechnol. 2002; 22: 51-86Crossref PubMed Google Scholar). Of these two enzymes, MMP-2 has been investigated in a more detail. For example, the activation of pro-MMP-2 has been thoroughly characterized and involves interactions of TIMP-2, MT1-MMP, and αVβ3 integrin on the cell surface (6Brooks P.C. Silletti S. von Schalscha T.L. Friedlander M. Cheresh D.A. Cell. 1998; 92: 391-400Abstract Full Text Full Text PDF PubMed Scopus (572) Google Scholar, 7Deryugina E.I. Ratnikov B. Monosov E. Postnova T.I. DiScipio R. Smith J.W. Strongin A.Y. Exp. Cell Res. 2001; 263: 209-223Crossref PubMed Scopus (333) Google Scholar). MMP-9 has not been found to be activated via the same mechanism, and several proteinases including the plasmin/MMP-3 cascade (8Ramos-DeSimone N. Hahn-Dantona E. Sipley J. Nagase H. French D.L. Quigley J.P. J. Biol. Chem. 1999; 274: 13066-13076Abstract Full Text Full Text PDF PubMed Scopus (493) Google Scholar) and trypsin-2 (9Sorsa T. Salo T. Koivunen E. Tyynela J. Konttinen Y.T. Bergmann U. Tuuttila A. Niemi E. Teronen O. Heikkila P. Tschesche H. Leinonen J. Osman S. Stenman U.H. J. Biol. Chem. 1997; 272: 21067-21074Abstract Full Text Full Text PDF PubMed Scopus (173) Google Scholar) can activate MMP-9 in vitro. Relatively little is known about the molecular details of the MMP-9 interactions on the cell surface and how these regulate cell migration. MMP-9 has been found to interact with the α5β1 integrin, the α2 chain of type IV collagen, and the hyaluronan receptor CD44 (10Wang X.Q. Sun P. Paller A.S. J. Biol. Chem. 2003; 30: 30Google Scholar, 11Fridman R. Toth M. Chvyrkova I. Meroueh S.O. Mobashery S. Cancer Metastasis Rev. 2003; 22: 153-166Crossref PubMed Scopus (142) Google Scholar). We have recently identified the leukocyte specific β2-integrins as a binding partner for pro-MMP-9. The phage display peptide ADGACILWMDDGWCGAAG (DDGW) competed with pro-MMP-9 binding to the ligand-binding I domain of αM integrin subunit and inhibited migration of leukocytes (12Stefanidakis M. Bjorklund M. Ihanus E. Gahmberg C.G. Koivunen E. J. Biol. Chem. 2003; 278: 34674-34684Abstract Full Text Full Text PDF PubMed Scopus (59) Google Scholar). Here we have isolated MMP-9 binding peptides, which inhibit either substrate binding or proenzyme activation, leading to an inhibition of cell migration and invasion. Using these peptides, we identify MMP-9 interaction sites in fibronectin, vitronectin, and αVβ5 integrin. Phage Display—Phage display selections were made peptide and E. J. M. Arap W. Pasqualini R. Mol. Biol. 1999; Google Scholar). pro-MMP-9 (9Sorsa T. Salo T. Koivunen E. Tyynela J. Konttinen Y.T. Bergmann U. Tuuttila A. Niemi E. Teronen O. Heikkila P. Tschesche H. Leinonen J. Osman S. Stenman U.H. J. Biol. Chem. 1997; 272: 21067-21074Abstract Full Text Full Text PDF PubMed Scopus (173) Google Scholar) or MMP-9 C domain on and the were with The phage were in three of the phage were E. A. S. A. M. Ihanus E. Gahmberg C.G. J. Cell Biol. 2001; PubMed Scopus Google Scholar). The phage binding with or the The phage were to bind in the or of or by with PBS plus 0.05% Tween The phage were with a phage were in a (12Stefanidakis M. Bjorklund M. Ihanus E. Gahmberg C.G. Koivunen E. J. Biol. Chem. 2003; 278: 34674-34684Abstract Full Text Full Text PDF PubMed Scopus (59) Google Scholar, M. Valtanen H. H. Koivunen E. Chem. 2003; PubMed Scopus Google Scholar). peptide and the and of the were by M. Valtanen H. H. Koivunen E. Chem. 2003; PubMed Scopus Google Scholar). The were in the CRVYGPYLLC and peptides, which were in a and PBS to the The and were on a The with in plus 0.05% Tween and with CBD and of the and MMP-9 with the and and The CBD in and For CBD with The C-terminal of MMP-2 and MMP-9 were as M. Koivunen E. Google Scholar). The with the and with and and the The by with in the of and with The integrin-epidermal growth domain 2 integrin and with and and The with an in E. and in a The and were by The of by of MMP-2 and MMP-9 as a substrate M. Valtanen H. H. Koivunen E. Chem. 2003; PubMed Scopus Google Scholar). and CBD CBD or fibronectin in in The were with in with or the or with an of and to bind for with CBD binding to fibronectin, the of fibronectin or T. M. H. M. Cell PubMed Scopus Google Scholar) CBD in in the or of of the MMP-9 C C domain or CBD a in PBS were on by with C domain with the for in and to the a the were with and with a Cell and cell cells were as (2Koivunen E. Arap W. Valtanen H. Rainisalo A. Medina O.P. Heikkila P. Kantor C. Gahmberg C.G. Salo T. Konttinen Y.T. Sorsa T. Ruoslahti E. Pasqualini R. Nat. Biotechnol. 1999; 17: 768-774Crossref PubMed Scopus (514) Google Scholar, E. A. S. A. M. Ihanus E. Gahmberg C.G. J. Cell Biol. 2001; PubMed Scopus Google Scholar, P. Heikkila P. Sorsa T. J. R. Stenman U.H. T. Salo T. J. Biol. Chem. 2003; 278: Full Text Full Text PDF PubMed Scopus Google Scholar). type and cells were by Cheresh the and as Cheresh D.A. J. Cell Google Scholar). Cell a Cell cells were to on or fibronectin in the or of or a integrin or The as E. A. S. A. M. Ihanus E. Gahmberg C.G. J. Cell Biol. 2001; PubMed Scopus Google Scholar). of cells to or in the or of or Cell Migration and cell migration migration in (2Koivunen E. Arap W. Valtanen H. Rainisalo A. Medina O.P. Heikkila P. Kantor C. Gahmberg C.G. Salo T. Konttinen Y.T. Sorsa T. Ruoslahti E. Pasqualini R. Nat. Biotechnol. 1999; 17: 768-774Crossref PubMed Scopus (514) Google Scholar, E. A. S. A. M. Ihanus E. Gahmberg C.G. J. Cell Biol. 2001; PubMed Scopus Google Scholar). the were on with or with the integrin peptide and with cells were with the for in The cells were to for and were with and E. A. S. A. M. Ihanus E. Gahmberg C.G. J. Cell Biol. 2001; PubMed Scopus Google Scholar). The as the that were were with a of fibronectin and by with in cells in of were in the of and the or the inhibitor a cells and the cells were as the of a of the a and E. and to αM binding to the αM I domain in the of as (12Stefanidakis M. Bjorklund M. Ihanus E. Gahmberg C.G. Koivunen E. J. Biol. Chem. 2003; 278: 34674-34684Abstract Full Text Full Text PDF PubMed Scopus (59) Google Scholar). binding to the integrin by the integrin (12Stefanidakis M. Bjorklund M. Ihanus E. Gahmberg C.G. Koivunen E. J. Biol. Chem. 2003; 278: 34674-34684Abstract Full Text Full Text PDF PubMed Scopus (59) Google Scholar) or as a in plus and by of the with in for 2 and the pro-MMP-9 to bind for with of cells or cells were for in the or of PDBu, and the a of the were by (9Sorsa T. Salo T. Koivunen E. Tyynela J. Konttinen Y.T. Bergmann U. Tuuttila A. Niemi E. Teronen O. Heikkila P. Tschesche H. Leinonen J. Osman S. Stenman U.H. J. Biol. Chem. 1997; 272: 21067-21074Abstract Full Text Full Text PDF PubMed Scopus (173) Google Scholar). MMP-9 activated in vitro with to in Tween for in the or of or the The were by and cells were activated with for in with PBS, and in of with of or or a were with 2 of domain A. W. M.D. F. Ruoslahti E. J. Cell Biol. PubMed Scopus Google Scholar). The were on an and with cells were to on in migration of the cells by the cells with in and of bovine serum with to the of the cells were with PBS, with and with the 2 or integrin R. J. S. J. Cell Google and MMP-9 The were with and of with of MMP-9 in with or The were for in and in a by with on the surface of cells or cells in a with or for in the or of or peptide, or The cells were three with PBS, with plus to cell urokinase-plasminogen activator and MMPs, and with plus were by N. M. E. K. PubMed Scopus Google Scholar), and or of on and for as were the with and uPA with H. I. A. Biol. Chem. 2001; PubMed Scopus Google Scholar). C domain, MMP-2 C domain, CBD, or were in to the which were with the for in and for with to MMP-9 C domain with to a specific activity of The domain of the as by the The cells were with in PBS and in a of cells in a on for with the C domain and for on The cells were to of for and M. J. Pasqualini R. Arap W. Nat. Med. 2001; PubMed Scopus Google Scholar). The of the the cells and with a in were by the of were by tumor cells in a in PBS in of the the of or the peptide or the via the in a were a and the tumor were were the tumor For the of the tumor tissue were with and with the or with in of to of to cell migration in we for by phage display of peptide The pro-MMP-9 and were used in the MMP-9 with a (2Koivunen E. Arap W. Valtanen H. Rainisalo A. Medina O.P. Heikkila P. Kantor C. Gahmberg C.G. Salo T. Konttinen Y.T. Sorsa T. Ruoslahti E. Pasqualini R. Nat. Biotechnol. 1999; 17: 768-774Crossref PubMed Scopus (514) Google Scholar). of pro-MMP-9 binding were found I a an show to found in the fibronectin and (4Overall C.M. Mol. Biotechnol. 2002; 22: 51-86Crossref PubMed Google Scholar). II a the CRVYGPYLLC peptide by with whereas the other were with a C-terminal domain. The and were for as of the two binding peptide C domain-binding in a identify the binding sites of these peptide we phage binding of phage to pro-MMP-9 inhibited by a ADGACGYGRFSPPCGAAG peptide and but not with or a peptide binding of the inhibited by and not by PPC, CTT, or binding sites for these of phage binding by that that to the CBD of MMP-9. phage with the MMP-2 CBD has also a peptide M. K. M. M. J. Biol. Chem. 2003; 278: Full Text Full Text PDF PubMed Scopus Google Scholar). The peptide, but not or CRV, inhibited binding to CBD in a but on binding to fibronectin not that is specific for the fibronectin type II repeats of a inhibited MMP-9 and MMP-2 activity the peptide The peptide not inhibit with the of to with The peptide a inhibitor CTT, which inhibits a in M. Valtanen H. H. Koivunen E. Chem. 2003; PubMed Scopus Google Scholar). the of fibronectin and bind MMP-9, we the binding of CBD to these in a binding CBD to fibronectin and vitronectin, but not to the of fibronectin lacking the C-terminal domain and the site PPC, but not the peptide, inhibited the CBD were in a CBD to the fibronectin peptide but not to an peptide different were the phage a binding to and not to pro-MMP-2 or The peptide MMP-9 selectivity also with the MMP-9 and MMP-2 C domains. The phage the MMP-9 C domain in with the MMP-2 C domain not with the phage binding to the MMP-9 C domain or pro-MMP-9 The phage not bind to the CBD or a pro-MMP-9 lacking the hinge region and the C-terminal domain not We the of on the dimerization of MMP-9 C domain. C domain with or peptide and to with C domain. of the C domain inhibited by but not by the peptide Cell Migration and by the of the the role of the in cell migration and the CTT, PPC, and The binding site of to the catalytic domain, but not to CBD (12Stefanidakis M. Bjorklund M. Ihanus E. Gahmberg C.G. Koivunen E. J. Biol. Chem. 2003; 278: 34674-34684Abstract Full Text Full Text PDF PubMed Scopus (59) Google and E. and are for the CBD and the C domain, three inhibited a of or CTT, inhibition The peptide a to the same The were were with which on a E. A. S. A. M. Ihanus E. Gahmberg C.G. J. Cell Biol. 2001; PubMed Scopus Google Scholar) in a and PPC, CRV, and CTT, but not the peptides, an The inhibition of cell migration not to as there on cell the cells were for with the a inhibited as and PPC, as by a of the cells were for in the of on a The gelatinase-selective small molecule inhibitor also inhibited but the that not the inhibitors but also cell migration and We also that the and do not the interaction of MMP-9 with the leukocyte αM integrin I domain, which is by pro-MMP-9 binding to the I domain as by binding in the of binding to pro-MMP-9 in the of the I domain not by The that the MMP-9 bind CBD to a molecular between an integrin, MMP-9, and a directly pro-MMP-9 to bind to integrin, and binding of an MMP-9 to the integrin but not the integrin The integrin not binding MMP-9 with the investigated the of the on pro-MMP-9 activation in and The the cells in the of the by Of the three peptides, of pro-MMP-9 peptide inhibited pro-MMP-9 activation and the activation of pro-MMP-2 The of in pro-MMP-9 in and not activation pro-MMP-9 activation and and the activation by but not by the other pro-MMP-9 activation in the of or DDGW, and there were of MMP-9 as previously with (12Stefanidakis M. Bjorklund M. Ihanus E. Gahmberg C.G. Koivunen E. J. Biol. Chem. 2003; 278: 34674-34684Abstract Full Text Full Text PDF PubMed Scopus (59) Google Scholar). not inhibit the activation of pro-MMP-9 by in vitro the activation cascade is in pro-MMP-9 activation, we the that the receptor with MMP-9. cells that pro-MMP-9 with but not with the The association of and pro-MMP-9 found in cells and not by activation proteinases are to P. N. E. R. F. 2001; PubMed Scopus Google Scholar, A. M. F. N. 2002; PubMed Scopus Google we MMP-9 also does Using we that MMP-9 cleaved the domain as does The by MMP-9 in the of and inhibited by the inhibitor on the surface of cells P. N. E. R. F. 2001; PubMed Scopus Google Scholar). the of gelatinases in we cells with inhibitors and the by with to The gelatinase-selective inhibitor but not the inhibitors or inhibited The inhibition of with in the and on the cell the MMP-9 in whereas the for the cell The cell MMP-9 in the as previously M. R. Cancer Res. 1997; Google Scholar). the of cell uPA in the of on the cells inhibited by and but not by the peptide M. Valtanen H. H. Koivunen E. Chem. 2003; PubMed Scopus Google Scholar) or the of PDBu, the cells in a of The a of an β is to with and Y. H. Z. S. J. Biol. Chem. Full Text Full Text PDF PubMed Scopus Google Scholar, W. I. Cancer Res. 1997; Google Scholar, S. M. P. A. M. Cancer Res. 1999; Google Scholar, K. J. Cell Biol. 1999; PubMed Scopus Google Scholar). We investigated which interact with MMP-9 in were with and integrins. with the and that α5β1 and αVβ5 are the in pro-MMP-9 binding in cells on a tissue and can interact with through C-terminal (6Brooks P.C. Silletti S. von Schalscha T.L. Friedlander M. Cheresh D.A. Cell. 1998; 92: 391-400Abstract Full Text Full Text PDF PubMed Scopus (572) Google Scholar, Nagase H. J. Biol. Chem. 2001; Full Text Full Text PDF PubMed Scopus Google Scholar). a that the peptide a to found in the stalk of the integrin β in the of the a or a in the are in the domain 2 and in the activated as by the of activation C. M. J. J. 2001; PubMed Scopus Google Scholar, J. Biol. Chem. Full Text Full Text PDF PubMed Scopus Google Scholar). the epitope to the in the integrin chain C. M. J. J. 2001; PubMed Scopus Google Scholar). MMP-9 to integrin activation we the of the MMP-9 C domain on cell to and the C domain the or the peptide inhibited cell to or fibronectin to in a as demonstrated by inhibition with the αVβ5 We not specific of cells to the C domain that the interaction site of the MMP-9 C domain in α5β1 and αVβ5 is not the ligand-binding site or a cell to the 2 and N. J. Nat. Biol. 2002; PubMed Scopus Google Scholar) the integrin. specifically to the MMP-9 C domain in a The not bind to MMP-9 CBD, vitronectin, or or the C domain of MMP-2 The binding and be inhibited with We the and of the to to the of the in a of the and for the binding of to the MMP-9 C domain The also the of to inhibit through MMP-9 C domain and MMP-2 C domain inhibited with a whereas find for the integrin we the binding of MMP-9 C domain to αVβ5 MMP-9 C domain a specific binding to but not to the cells The binding competed with MMP-9 C domain, the and to a the with the MMP-2 C domain or the the MMP-9 C domain is to bind to the site of the integrin, we cell binding to the cells to the but not to an The C domain inhibited the cell binding by whereas CBD not The of the binding is by with but not by a The C domain on binding to of cells on a for uPAR, MMP-9 and integrin. MMP-9 on the leading of the the with integrin and are with of with MMP-9 also found on the surface of cells a of of the peptide, we on tumor growth in vivo. cell were with the peptide the were in an and not CRV, the peptide, or PBS a inhibition of tumor growth by in with the peptide or PBS the of the and 2 were whereas the the peptide or PBS been to The of be for by inhibition of The a developed tumor as by of the We have developed peptide to the gelatinases and molecular interactions important for these enzymes. of the inhibited cell that the three of MMP-9 catalytic domain, CBD, and the C play a We have previously that in pro-MMP-9 with the αM and integrin I through the catalytic domain (12Stefanidakis M. Bjorklund M. Ihanus E. Gahmberg C.G. Koivunen E. J. Biol. Chem. 2003; 278: 34674-34684Abstract Full Text Full Text PDF PubMed Scopus (59) Google Scholar). Here we have found integrin interaction for MMP-9, the C domain of MMP-9 to the integrin β to the I domain which in the of and pro-MMP-9 the C subunit interaction activated and appears to play a role in MMP-9 activation and Of the MMP-9 binding identified in the peptide as an inhibitor of MMP-2 and binding and but on the MMP-9 interactions with integrins. We identified a in the domain of fibronectin as a CBD a but binding site for not bind to the fibronectin type II repeats of fibronectin, it as a for the development of highly specific The C-terminal domain-binding peptide not the activity of MMP-9 but inhibited dimerization of the MMP-9 C domain, activation of the pro-MMP-9 via and that is a of the activation epitope in the integrin β the The C domain of MMP-9 inhibited leukocyte to the which the site in the integrin. The integrin specifically to the C domain in a and the of the and in the The but not the the C domain of MMP-9. pro-MMP-9 with to and the and the C domain inhibited of cell MMP-9 and to the leading of the we the that the peptide inhibits also other C We not association of MMP-9 with αVβ3 in a between MMP-9 and the αVβ3 integrin has been found M. E. J. A. B. U. S. A. 2003; Scholar). the that cells the αVβ5 integrin for MMP-9 binding to αVβ5 be more αVβ5 and MMP-9 are regulation (8Ramos-DeSimone N. Hahn-Dantona E. Sipley J. Nagase H. French D.L. Quigley J.P. J. Biol. Chem. 1999; 274: 13066-13076Abstract Full Text Full Text PDF PubMed Scopus (493) Google Scholar, M. P.C. C.M. Cheresh D.A. PubMed Scopus Google Scholar). αVβ3 and MMP-2 to be W. W. R. Y. K. R. 1999; PubMed Scopus Google Scholar). the peptide inhibited pro-MMP-2 activation, and the C of MMP-2 and MMP-9 not with other in binding The that mimics an integrin activation epitope an for the of in pro-MMP-9 activation M. E. J. A. B. U. S. A. 2003; Scholar, C. A. E. C. M. Exp. Cell Res. 2002; PubMed Scopus Google Scholar). We also that uPAR, which is for MMP-9 activation, with pro-MMP-9 in and a substrate for MMP-9 in and the of by MMP-9 in the of the domain of uPAR, which has also been with other such as A. M. F. N. 2002; PubMed Scopus Google Scholar). of uPA binding and the of and N. S. P. J. Biol. Chem. 2002; Full Text Full Text PDF PubMed Scopus Google Scholar). MMP-9 not activation but also of MMP-9 and has been to be for the of tumor cells J. W. K. Cell. 1998; Full Text Full Text PDF PubMed Scopus Google Scholar). and gelatinases in in cells A. S. Nagase H. P. J. Biol. Chem. 1997; 272: Full Text Full Text PDF PubMed Scopus Google Scholar, T.L. T. T. S. PubMed Scopus Google Scholar). of tumor growth by an important for the in tumor growth tumor growth inhibition is in the and integrin Nat. Med. 2002; PubMed Scopus Google Scholar) and also in with of MMP-9 A. 2002; PubMed Scopus Google Scholar). The of MMP-9 to or Y. M. H. Y. C. Werb Z. A. R. Cancer Cell. 2003; Full Text Full Text PDF PubMed Scopus Google Scholar) these and MMP-9 is also used for the of by MMP-9 also inhibit tumor tumor inhibition of activity have not been the to inhibit be more (6Brooks P.C. Silletti S. von Schalscha T.L. Friedlander M. Cheresh D.A. Cell. 1998; 92: 391-400Abstract Full Text Full Text PDF PubMed Scopus (572) Google Scholar). is that phage specifically with different interactions either the catalytic or the C-terminal domain that specific can be developed that but not the also the peptide, which the is in in an inflammation in M. T. N. Gahmberg C. Koivunen E. J. PubMed Scopus Google Scholar). of the MMP-9 interactions with is on a with phage display and that pro-MMP-9 can interact with in two the interaction between the integrin I domain and the MMP-9 catalytic domain is and pro-MMP-9 in an do not the that and β interactions the same binding the integrin and the activation epitope in the β which can as a site for the C domain of MMP-9. MMP-9 be activated by or by binding to a substrate J. Biol. Chem. 2002; Full Text Full Text PDF PubMed Scopus Google Scholar). that an I domain in the the MMP-9 C interaction be the We for MMP-9 Cheresh for Gahmberg for integrin and Ruoslahti for other for of for and for peptide