Angiostatin

The quiescent vascular system in the adult body represents the imbalanced net outcome of overproduction of endogenous angiogenesis inhibitors and reduced levels of angiogenic factors. While some endogenous inhibitors are expressed under physiological conditions, they can also be generated in association with tumor growth. Angiostatin is such a specific angiogenesis inhibitor produced by tumors. It inhibits primary and metastatic tumor growth by blocking tumor angiogenesis. Having demonstrated potent antitumor activity in animal studies, angiostatin is now in clinical trials for human cancer therapy. Angiostatin is not a novel protein molecule coded by novel DNA sequences. Instead, it is an internal proteolytic fragment of a known protein, plasminogen. Surprisingly, most kringle domains of plasminogen only inhibit angiogenesis when cleaved as fragments from their parent protein that lacks antiangiogenic activity. These findings suggest that they are cryptic fragments hidden in large protein molecules. Thus, proteolytic processing plays a critical role in down-regulation of angiogenesis. Despite proteolytic processing, the antiangiogenic mechanism of angiostatin remains an enigma. Without knowing the mechanisms, it is difficult to predict the ultimate outcome of ongoing clinical trials. In this article, we discuss what is known about angiostatin and how this molecule specifically inhibits angiogenesis. We hope that the information will be useful for further development of angiostatin and its related inhibitors as therapeutic agents.

KEYWORDS

Neovascularization - angiogenesis - tumor - metastasis

REFERENCES

• 1 O'Reilly M S, Holmgren L, Shing Y et al.. Angiostatin: a novel angiogenesis inhibitor that mediates the suppression of metastases by a Lewis lung carcinoma.  Cell. 1994;  79 315-328
• 2 Magnusson S, Sottrup-Jensen L, Petersen T E, Claeys H. The primary structure of prothrombin, the role of vitamin K in blood coagulation and thrombin catalyzed “negative feed-back” control mechanism for limiting the activation of prothrombin. Prothrombin and related coagulation factors. Leiden; Universitaire Press 1975: 25-46
• 3 Nakamura T, Nishizawa T, Hagiya M et al.. Molecular cloning and expression of human hepatocyte growth factor.  Nature. 1989;  342 440-443
• 4 Han S, Stuart L A, Degen S J. Characterization of the DNF15S2 locus on human chromosome 3: identification of a gene coding for four kringle domains with homology to hepatocyte growth factor.  Biochemistry. 1991;  30 9768-9780
• 5 Gunzler W A, Steffens G J, Otting F et al.. The primary structure of high molecular mass urokinase from human urine. The complete amino acid sequence of the A chain.  Hoppe Seylers Z Physiol Chem. 1982;  363 1155-1165
• 6 Pennica D, Holmes W E, Kohr W J et al.. Cloning and expression of human tissue-type plasminogen activator cDNA in E. coli .  Nature. 1983;  301 214-221
• 7 Miyazawa K, Shimomura T, Kitamura A et al.. Molecular cloning and sequence analysis of the cDNA for a human serine protease responsible for activation of hepatocyte growth factor. Structural similarity of the protease precursor to blood coagulation factor XII.  J Biol Chem. 1993;  268 10024-10028
• 8 Gschwend T P, Krueger S R, Kozlov S V et al.. Neurotrypsin, a novel multidomain serine protease expressed in the nervous system.  Mol Cell Neurosci. 1997;  9 207-219
• 9 Arnold J M, Kennet C, Degnan B M, Lavin M F. Transient expression of a novel serine protease in the ectoderm of the ascidian Hermandia momus during development.  Dev Genes Evol. 1997;  206 455-463
• 10 Sottrup-Jensen L, Claeys H, Zajdel M et al.. The primary structure of human plasminogen: isolation of two lysine-binding fragments and one “mini-”plasminogen (MW, 38000) by elastase-catalyzed-specific limited proteolysis.  Progr Chem Fibrinolysis Thrombolysis. 1978;  3 191-208
• 11 Sottrup-Jensen L, Zajdel M, Claeys H et al.. Amino-acid sequence of activation cleavage site in plasminogen: homology with “pro” part of prothrombin.  Proc Natl Acad Sci USA. 1975;  72 2577-2581
• 12 McMullen B A, Fujikawa K. Amino acid sequence of the heavy chain of human alpha-factor XIIa (activated Hageman factor).  J Biol Chem. 1985;  260 5328-5341
• 13 Prodinger W M, Schwendinger M G, Schoch J et al.. Characterization of C3dg binding to a recess formed between short consensus repeats 1 and 2 of complement receptor type 2 (CR-2; CD21).  J Immunol. 1998;  161 4604-4610
• 14 van de Poel R H, Meijers J C, Bouma B N. Interaction between protein S and complement C4b-binding protein (C4BP). Affinity studies using chimeras containing c4bp beta-chain short consensus repeats.  J Biol Chem. 1999;  274 15144-15150
• 15 Ozhogina O A, Trexler M, Banyai L et al.. Origin of fibronectin type II (FN2) modules: structural analyses of distantly related members of the kringle family idey the kringle domain of neurotrypsin as a potential link between FN2 domains and kringles.  Protein Sci. 2001;  10 2114-2122
• 16 Cao Y. Endogenous angiogenesis inhibitors: angiostatin, endostatin, and other proteolytic fragments.  Prog Mol Subcell Biol. 1998;  20 161-176
• 17 Cao Y, Ji R W, Davidson D et al.. Kringle domains of human angiostatin. Characterization of the anti-proliferative activity on endothelial cells.  J Biol Chem. 1996;  271 29461-29467
• 18 Mulichak A M, Tulinsky A, Ravichandran K G. Crystal and molecular structure of human plasminogen kringle 4 refined at 1.9-A resolution.  Biochemistry. 1991;  30 10576-10588
• 19 Rejante M R, Byeon I J, Llinas M. Ligand specificity of human plasminogen kringle 4.  Biochemistry. 1991;  30 11081-11092
• 20 Cao Y, Chen A, An S S et al.. Kringle 5 of plasminogen is a novel inhibitor of endothelial cell growth.  J Biol Chem. 1997;  272 22924-22928
• 21 Cao R, Wu H L, Veitonmaki N et al.. Suppression of angiogenesis and tumor growth by the inhibitor K1-5 generated by plasmin-mediated proteolysis.  Proc Natl Acad Sci USA. 1999;  96 5728-5733
• 22 Li F, Yang J, Liu X et al.. Human glioma cell BT325 expresses a proteinase that converts human plasminogen to kringle 1-5-containing fragments.  Biochem Biophys Res Commun. 2000;  278 821-825
• 23 Lee T H, Rhim T, Kim S S. Prothrombin kringle-2 domain has a growth inhibitory activity against basic fibroblast growth factor-stimulated capillary endothelial cells.  J Biol Chem. 1998;  273 28805-28812
• 24 Rhim T Y, Park C S, Kim E, Kim S S. Human prothrombin fragment 1 and 2 inhibit bFGF-induced BCE cell growth.  Biochem Biophys Res Commun. 1998;  252 513-516
• 25 Schulter V, Koolwijk P, Peters E et al.. Impact of apolipoprotein(a) on in vitro angiogenesis.  Arterioscler Thromb Vasc Biol. 2001;  21 433-438
• 26 Trieu V N, Uckun F M. Apolipoprotein(a), a link between atherosclerosis and tumor angiogenesis.  Biochem Biophys Res Commun. 1999;  257 714-718
• 27 O'Reilly M S, Holmgren L, Chen C, Folkman J. Angiostatin induces and sustains dormancy of human primary tumors in mice.  Nat Med. 1996;  2 689-692
• 28 Holmgren L. Antiangiogenis restricted tumor dormancy.  Cancer Metastasis Rev. 1996;  15 241-245
• 29 Folkman J. Angiogenesis in cancer, vascular, rheumatoid and other disease.  Nat Med. 1995;  1 27-31
• 30 Folkman J. Seminars in Medicine of the Beth Israel Hospital, Boston. Clinical applications of research on angiogenesis.  N Engl J Med. 1995;  333 1757-1763
• 31 Available at: http://www.nci.nih.gov/clinical_trials.
• 32 Cao Y. Endogenous angiogenesis inhibitors and their therapeutic implications.  Int J Biochem Cell Biol. 2001;  33 357-369
• 33 O'Reilly M S, Boehm T, Shing Y et al.. Endostatin: an endogenous inhibitor of angiogenesis and tumor growth.  Cell. 1997;  88 277-285
• 34 O'Reilly M S, Pirie-Shepherd S, Lane W S, Folkman J. Antiangiogenic activity of the cleaved conformation of the serpin antithrombin.  Science. 1999;  285 1926-1928
• 35 Maeshima Y, Colorado P C, Torre A et al.. Distinct antitumor properties of a type IV collagen domain derived from basement membrane.  J Biol Chem. 2000;  275 21340-21348
• 36 Kamphaus G D, Colorado P C, Panka D J et al.. Canstatin, a novel matrix-derived inhibitor of angiogenesis and tumor growth.  J Biol Chem. 2000;  275 1209-1215
• 37 Pike S E, Yao L, Jones K D et al.. Vasostatin, a calreticulin fragment, inhibits angiogenesis and suppresses tumor growth.  J Exp Med. 1998;  188 2349-2356
• 38 Clapp C, Martial J A, Guzman R C et al.. The 16-kilodalton N-terminal fragment of human prolactin is a potent inhibitor of angiogenesis.  Endocrinology. 1993;  133 1292-1299
• 39 Ramchandran R, Dhanabal M, Volk R et al.. Antiangiogenic activity of restin, NC10 domain of human collagen XV: comparison to endostatin.  Biochem Biophys Res Commun. 1999;  255 735-739
• 40 Colorado P C, Torre A, Kamphaus G et al.. Anti-angiogenic cues from vascular basement membrane collagen.  Cancer Res. 2000;  60 2520-2526
• 41 Grant D S, Kleinman H K, Goldberg I D et al.. Scatter factor induces blood vessel formation in vivo.  Proc Natl Acad Sci USA. 1993;  90 1937-1941
• 42 Rosen E M, Goldberg I D. Scatter factor and angiogenesis.  Adv Cancer Res. 1995;  67 257-279
• 43 Xin X, Yang S, Ingle G et al.. Hepatocyte growth factor enhances vascular endothelial growth factor-induced angiogenesis in vitro and in vivo.  Am J Pathol. 2001;  158 1111-1120
• 44 Coussens L M, Tinkle C L, Hanahan D, Werb Z. MMP-9 supplied by bone marrow-derived cells contributes to skin carcinogenesis.  Cell. 2000;  103 481-490
• 45 Zhou Z, Apte S S, Soininen R et al.. Impaired endochondral ossification and angiogenesis in mice deficient in membrane-type matrix metalloproteinase I.  Proc Natl Acad Sci USA. 2000;  97 4052-4057
• 46 Kramer M D, Reinartz J, Brunner G, Schirrmacher V. Plasmin in pericellular proteolysis and cellular invasion.  Invasion Metastasis. 1994;  14 210-222
• 47 Andreasen P A, Egelund R, Petersen H H. The plasminogen activation system in tumor growth, invasion, and metastasis.  Cell Mol Life Sci. 2000;  57 25-40
• 48 Coussens L M, Fingleton B, Matrisian L M. Matrix metalloproteinase inhibitors and cancer: trials and tribulations.  Science. 2002;  295 2387-2392
• 49 Moser T L, Kenan D J, Ashley T A et al.. Endothelial cell surface F1-F0 ATP synthase is active in ATP synthesis and is inhibited by angiostatin.  Proc Natl Acad Sci USA. 2001;  98 6656-6661
• 50 Moser T L, Stack M S, Asplin I et al.. Angiostatin binds ATP synthase on the surface of human endothelial cells.  Proc Natl Acad Sci USA. 1999;  96 2811-2816
• 51 Troyanovsky B, Levchenko T, Mansson G et al.. Angiomotin: an angiostatin binding protein that regulates endothelial cell migration and tube formation.  J Cell Biol. 2001;  152 1247-1254
• 52 Griscelli F, Li H, Bennaceur-Griscelli A et al.. Angiostatin gene transfer: inhibition of tumor growth in vivo by blockage of endothelial cell proliferation associated with a mitosis arrest.  Proc Natl Acad Sci USA. 1998;  95 6367-6372
• 53 Lucas R, Holmgren L, Garcia I et al.. Multiple forms of angiostatin induce apoptosis in endothelial cells.  Blood. 1998;  92 4730-4741
• 54 Claesson-Welsh L, Welsh M, Ito N et al.. Angiostatin induces endothelial cell apoptosis and activation of focal adhesion kinase independently of the integrin-binding motif RGD.  Proc Natl Acad Sci USA. 1998;  95 5579-5583
• 55 Lu H, Dhanabal M, Volk R et al.. Kringle 5 causes cell cycle arrest and apoptosis of endothelial cells.  Biochem Biophys Res Commun. 1999;  258 668-673
• 56 Auerbach W, Auerbach R. Angiogenesis inhibition: a review.  Pharmacol Ther. 1994;  63 265-311
• 57 Hanahan D, Folkman J. Patterns and emerging mechanisms of the angiogenic switch during tumorigenesis.  Cell. 1996;  86 353-364
• 58 http://www.pbs.org/wgbh/nova/cancer/program.html.
• 59 Kisker O, Becker C M, Prox D et al.. Continuous administration of endostatin by intraperitoneally implanted osmotic pump improves the efficacy and potency of therapy in a mouse xenograft tumor model.  Cancer Res. 2001;  61 7669-7674
• 60 Drixler T A, Rinkes I H, Ritchie E D et al.. Continuous administration of angiostatin inhibits accelerated growth of colorectal liver metastases after partial hepatectomy.  Cancer Res. 2000;  60 1761-1765
• 61 Cao Y, O'Reilly M S, Marshall B et al.. Expression of angiostatin cDNA in a murine fibrosarcoma suppresses primary tumor growth and produces long-term dormancy of metastases.  J Clin Invest. 1998;  101 1055-1063
• 62 Cao Y. Antiangiogenic gene therapy.  Gene Therapy Regulation. 2000;  1 123-139
• 63 Boehm T, Folkman J, Browder T, O'Reilly M S. Antiangiogenic therapy of experimental cancer does not induce acquired drug resistance.  Nature. 1997;  390 404-407
• 64 Maeshima Y, Sudhakar A, Lively J C et al.. Tumstatin, an endothelial cell-specific inhibitor of protein synthesis.  Science. 2002;  295 140-143
• 65 Folkman J. Looking for a good endothelial address.  Cancer Cell. 2002;  2 113-155
• 66 Cool D E, Edgell C J, Louie G V, Zoller M J, Brayer G D, MacGillivray R T. Characterization of human blood coagulation factor XII cDNA. Prediction of the primary structure of factor XII and the tertiary structure of beta-factor XIIa.  J Biol Chem. 1985;  260 13666-13676
• 67 McLean J W, Tomlinson J E, Kuang W J et al.. cDNA sequence of human apolipoprotein(a) is homologous to plasminogen.  Nature. 1987;  330 132-137
• 68 Eaton D L, Fless G M, Kohr W J et al.. Partial amino acid sequence of apolipoprotein(a) shows that it is homologous to plasminogen.  Proc Natl Acad Sci USA. 1987;  84 3224-3228
• 69 Nakamura T, Aoki S, Kitajima K, Takahashi T, Matsumoto K. Molecular cloning and characterization of Kremen, a novel kringle-containing transmembrane protein.  Biochim Biophys Acta. 2001;  1518 63-72
• 70 Yamamura Y, Yamashiro K, Tsuruoka N, Nakazato H, Tsujimura A, Yamaguchi N. Molecular cloning of a novel brain-specific serine protease with a kringle-like structure and three scavenger receptor cysteine-rich motifs.  Biochem Biophys Res Commun. 1997;  239 386-392
• 71 Choi-Miura N H, Tobe T, Sumiya J et al.. Purification and characterization of a novel hyaluronan-binding protein (PHBP) from human plasma: it has three EGF, a kringle and a serine protease domain, similar to hepatocyte growth factor activator.  J Biochem (Tokyo). 1996;  119 1157-1165
• 72 Masiakowski P, Carroll R D. A novel family of cell surface receptors with tyrosine kinase-like domain.  J Biol Chem. 1992;  267 26181-26190
• 73 Oishi I, Sugiyama S, Liu Z J, Yamamura H, Nishida Y, Minami Y. A novel Drosophila receptor tyrosine kinase expressed specifically in the nervous system. Unique structural features and implication in developmental signaling.  J Biol Chem. 1997;  272 11916-11923
• 74 Wilson C, Goberdhan D C, Steller H. Dror, a potential neurotrophic receptor gene, encodes a Drosophila homolog of the vertebrate Ror family of Trk-related receptor tyrosine kinases.  Proc Natl Acad Sci USA. 1993;  90 7109-7113
• 75 Forrester W C, Dell M, Perens E, Garriga G. A C. elegans Ror receptor tyrosine kinase regulates cell motility and asymmetric cell division.  Nature. 1999;  400 881-885
• 76 Jennings C G, Dyer S M, Burden S J. Muscle-specific trk-related receptor with a kringle domain defines a distinct class of receptor tyrosine kinases.  Proc Natl Acad Sci USA. 1993;  90 2895-2899
• 77 Fu A K, Smith F D, Zhou H et al.. Xenopus muscle-specific kinase: molecular cloning and prominent expression in neural tissues during early embryonic development.  Eur J Neurosci. 1999;  11 373-382
• 78 O'Reilly M S, Wiederschain D, Stetler-Stevenson W G, Folkman J, Moses M A. Regulation of angiostatin production by matrix metalloproteinase-2 in a model of concomitant resistance.  J Biol Chem. 1999;  274 29568-29571
• 79 Lijnen H R, Ugwu F, Bini A, Collen D. Generation of an angiostatin-like fragment from plasminogen by stromelysin-1 (MMP-3).  Biochemistry. 1998;  37 4699-4702
• 80 Patterson B C, Sang Q A. Angiostatin-converting enzyme activities of human matrilysin (MMP-7) and gelatinase B/type IV collagenase (MMP-9).  J Biol Chem. 1997;  272 28823-28825
• 81 Dong Z, Kumar R, Yang X, Fidler I J. Macrophage-derived metalloelastase is responsible for the generation of angiostatin in Lewis lung carcinoma.  Cell. 1997;  88 801-810
• 82 Takada A, Sugawara Y, Takada Y. Degradation of Glu- and Lys-plasminogen by elastase in the presence or absence of tranexamic acid.  Thromb Res. 1988;  50 285-294
• 83 Novokhatny V V, Kudinov S A, Privalov P L. Domains in human plasminogen.  J Mol Biol. 1984;  179 215-232
• 84 Lerch P G, Rickli E E, Lergier W, Gillessen D. Localization of individual lysine-binding regions in human plasminogen and investigations on their complex-forming properties.  Eur J Biochem. 1980;  107 7-13
• 85 Motta A, Laursen R A, Rajan N, Llinas M. Proton magnetic resonance study of kringle 1 from human plasminogen. Insights into the domain structure.  J Biol Chem. 1986;  261 13684-13692
• 86 Vali Z, Patthy L. The fibrin-binding site of human plasminogen. Arginines 32 and 34 are essential for fibrin affinity of the kringle 1 domain.  J Biol Chem. 1984;  259 13690-13694
• 87 Kwon M, Yoon C S, Fitzpatrick S et al.. p22 is a novel plasminogen fragment with antiangiogenic activity.  Biochemistry. 2001;  40 13246-13253
• 88 Falcone D J, Khan K M, Layne T, Fernandes L. Macrophage formation of angiostatin during inflammation. A byproduct of the activation of plasminogen.  J Biol Chem. 1998;  273 31480-31485
• 89 Stathakis P, Lay A J, Fitzgerald M, Schlieker C, Matthias L J, Hogg P J. Angiostatin formation involves disulfide bond reduction and proteolysis in kringle 5 of plasmin.  J Biol Chem. 1999;  274 8910-8916
• 90 Kassam G, Kwon M, Yoon C S et al.. Purification and characterization of A61. An angiostatin-like plasminogen fragment produced by plasmin autodigestion in the absence of sulfhydryl donors.  J Biol Chem. 2001;  276 8924-8933
• 91 Gately S, Twardowski P, Stack M S et al.. The mechanism of cancer-mediated conversion of plasminogen to the angiogenesis inhibitor angiostatin.  Proc Natl Acad Sci USA. 1997;  94 10868-10872
• 92 Lijnen H R, Van Hoef B, Ugwu F, Collen D, Roelants I. Specific proteolysis of human plasminogen by a 24 kDa endopeptidase from a novel Chryseobacterium Sp.  Biochemistry. 2000;  39 479-488
• 93 Morikawa W, Yamamoto K, Ishikawa S et al.. Angiostatin generation by cathepsin D secreted by human prostate carcinoma cells.  J Biol Chem. 2000;  275 38912-38920
• 94 Heidtmann H H, Nettelbeck D M, Mingels A, Jager R, Welker H G, Kontermann R E. Generation of angiostatin-like fragments from plasminogen by prostate-specific antigen.  Br J Cancer. 1999;  81 1269-1273