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Thromb Haemost 2000; 84(06): 1095-1102
DOI: 10.1055/s-0037-1614176
Review Article
Schattauer GmbH

Differential Expression of a Ligand Induced Binding Site (LIBS) by GPIIb-IIIa Ligand Recognition Peptides and Parenteral Antagonists

Lisa K. Jennings
1   From Vascular Biology Program, The University of Tennessee, Memphis, TN, USA
2   The Department of Biomedical Engineering, The University of Memphis, Memphis, TN, USA
,
Jason H. Haga
2   The Department of Biomedical Engineering, The University of Memphis, Memphis, TN, USA
,
Steven M. Slack
2   The Department of Biomedical Engineering, The University of Memphis, Memphis, TN, USA
› Author AffiliationsThis study was supported in part through a Grant-in-Aid (TN95G25) from the American Heart Association, Tennessee Affiliate (SMS), an Established Investigator Grant from the American Heart Association (LKJ), and grants HL53514 and HL38171 from the National Institutes of Health (LKJ).
Further Information

Publication History

Received 09 February 2000

Accepted after revision 01 June 2000

Publication Date:
13 December 2017 (online)

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Summary

The glycoprotein (GP) IIb-IIIa complex is an attractive anti-platelet target for the prevention of thrombotic events associated with coronary artery disease. Although GPIIb-IIIa antagonists inhibit GPIIb-IIIa binding to its ligands, the interactions have not been fully clarified, particularly with respect to their ability to induce structural changes in the complex that lead to exposure of neoantigenic epitopes or ligand-induced binding sites (LIBS). In this study we used the anti-LIBS monoclonal antibody (mAb) D3 to further define the activation states of purified active and inactive GPIIb-IIIa. We also compared the data obtained in the purified system to that observed with intact human platelets. Active GPIIb-IIIa expressed significantly greater high-affinity D3 LIBS sites compared to the inactive form. In addition, the ligand recognition peptides RGDS and H12 caused increased expression of the D3 epitope, with RGDS eliciting a much more potent response. The response of the purified GPIIb-IIIa to these peptides paralleled that observed with human platelets. To explore whether the platelet antagonists abciximab, eptifibatide and tirofiban induced expression of the D3 LIBS site, a modified competitive ELISA was developed. Our data indicate that the use of purified GPIIb-IIIa with this ELISA system provides a reproducible approach for exploring the interactions between GPIIb-IIIa and its antagonists. Whereas abciximab caused no detectable increase in the expression of the D3 epitope on purified GPIIb-IIIa, eptifibatide, tirofiban, RGDS, and H12 induced differential expression of the high-affinity LIBS. Studies with intact platelets suggested that abciximab blocked the binding of the D3 and LIBS6 mAbs, and that the pre bound anti-LIBS D3 sterically hindered abciximab binding.

Key words

GPIIb-IIIa - platelets - antagonists - ELISA - LIBS
 

  • References

  • 1 Sixma JJ, Wester J. The hemostatic plug. Semin Hematol 1977; 14: 265-99.
    PubMedSearch in Google Scholar
  • 2 Sakariassen KS, Nievelstein PFEM, Coller BS, Sixma JJ. The role of platelet membrane glycoproteins Ib and IIb-IIIa in platelet adherence to human artery subendothelium. Br J Haemotol 1986; 63: 681-91.
    CrossrefPubMedSearch in Google Scholar
  • 3 Sixma JJ, Pronk A, Nievelstein PN, Zwaginga JJ, Hindriks G, Tijburg P, Banga JD, De GP. Platelet adhesion to extracellular matrices of cultured cells. Ann N Y Acad Sci 1991; 614: 181-92.
    CrossrefPubMedSearch in Google Scholar
  • 4 Phillips DR, Charo IF, Scarborough RM. GPIIb-IIIa: The responsive integrin. Cell 1991; 65: 359-62.
    CrossrefPubMedSearch in Google Scholar
  • 5 Ruoslahti E, Pierschbacher MD. Arg-Gly-Asp: a versatile cell recognition signal. Cell 1986; 44: 517-8.
    CrossrefPubMedSearch in Google Scholar
  • 6 Ruoslahti E, Pierschbacher MD. New perspectives in cell adhesion: RGD and integrins. Science 1987; 238: 491-7.
    CrossrefPubMedSearch in Google Scholar
  • 7 Plow EF, Srouji AH, Meyer D, Marguerie G, Ginsberg MH. Evidence that three adhesive proteins interact with a common recognition site on activated platelets. J Biol Chem 1984; 259: 5388-91.
    PubMedSearch in Google Scholar
  • 8 Kloczewiak M, Timmons S, Lukas TJ, Hawiger J. Platelet receptor recognition site on human fibrinogen. Synthesis and structure-function relationship of peptides corresponding to the carboxy-terminal segment of the gamma chain. Biochemistry 1984; 23: 1767-74.
    CrossrefPubMedSearch in Google Scholar
  • 9 Lam SCT, Plow EF, Smith MA, Andrieux A, Ryckwaert JJ, Marguerie G, Ginsberg MH. Evidence that the arginyl-glycyl-aspartate peptides and fibrinogen γ chain peptides share a common binding site on platelets. J Biol Chem 1987; 262: 947-50.
    PubMedSearch in Google Scholar
  • 10 Santoro SA, Lawing WJ. Competition for related but nonidentical binding sites on the glycoprotein IIb-IIIa complex by peptides derived from platelet adhesive proteins. Cell 1987; 48: 867-73.
    CrossrefPubMedSearch in Google Scholar
  • 11 D’Souza SE, Ginsberg MH, Lam SC, Plow EF. Chemical cross-linking of arginyl-glycyl-aspartic acid peptides to an adhesion receptor on platelets. J Biol Chem 1988; 263: 3943-51.
    PubMedSearch in Google Scholar
  • 12 Gartner TK, Amrani DL, Derrick JM, Kirschbaum NE, Matsueda GR, Taylor DB. Characterization of adhesion of “resting” and stimulated platelets to fibrinogen and its fragments. Thromb Res 1993; 71: 47-60.
    CrossrefPubMedSearch in Google Scholar
  • 13 Holmback K, Danton M-J S, Daugherty CC, Degen JL. Impaired platelet aggregation and sustained bleeding in mice lacking the fibrinogen motif bound by integrin αIIbβ3 . EMBO 1996; 15: 5760-71.
    CrossrefPubMedSearch in Google Scholar
  • 14 Ruggeri ZM. von Willebrand Factor. J Clin Invest 1997; 99: 559-564.
    CrossrefPubMedSearch in Google Scholar
  • 15 Bennett JS, Vilaire G. Exposure of platelet fibrinogen receptors by ADP and epinephrine. J Clin Invest 1979; 64: 1393-401.
    CrossrefPubMedSearch in Google Scholar
  • 16 Marguerie GA, Edgington TS, Plow EF. Interaction of fibrinogen with its platelet receptor as part of a multi-step reaction in ADP-induced platelet aggregation. J Biol Chem 1980; 255: 154-61.
    PubMedSearch in Google Scholar
  • 17 Shadle PJ, Ginsberg MH, Plow EF, Barondes SH. Platelet-collagen adhesion: inhibition by a monoclonal antibody that binds glycoprotein IIb. J Cell Biol 1984; 99: 2056-60.
    CrossrefPubMedSearch in Google Scholar
  • 18 Frelinger AL, Cohen I, Plow EF, Smith MA, Roberts J, Lam SC, Ginsberg MH. Selective inhibition of integrin function by antibodies specific for ligand-occupied receptor conformers. J Biol Chem 1990; 265: 6346-52.
    PubMedSearch in Google Scholar
  • 19 Frelinger AL, Du XP, Plow EF, Ginsberg MH. Monoclonal antibodies to ligand-occupied conformers of integrin alpha IIb beta 3 (glycoprotein IIb-IIIa) alter receptor affinity, specificity, and function. J Biol Chem 1991; 266: 17106-11.
    PubMedSearch in Google Scholar
  • 20 Kouns WC, Wall CD, White MM, Fox CF, Jennings LK. A conformationdependent epitope of human platelet glycoprotein IIIa. J Biol Chem 1990; 265: 20594-601.
    PubMedSearch in Google Scholar
  • 21 Kouns WC, Jennings LK. Activation-independent exposure of the GPIIbIIIa fibrinogen receptor. Thromb Res 1991; 63: 343-54.
    CrossrefPubMedSearch in Google Scholar
  • 22 Steiner B, Cousot D, Trzeciak A, Gillessen D, Hadvery P. Ca2+-dependent binding of a synthetic Arg-Gly-Asp (RGD) peptide to a single site on purified platelet glycoprotein IIb-IIIa complex. J Biol Chem 1989; 264: 13102-8.
    PubMedSearch in Google Scholar
  • 23 Parise LV, Helgerson SL, Steiner B, Nannizzi L, Phillips DR. Synthetic peptides derived from fibrinogen and fibronectin change the conformation of purified platelet GPIIb-IIIa. J Biol Chem 1987; 262: 12597-602.
    PubMedSearch in Google Scholar
  • 24 Kouns WC, Hadvary P, Haering P, Steiner B. Conformational modulation of purified glycoprotein (GP) IIb-IIIa allows proteolytic generation of active fragments from either active or inactive GP IIb-IIIa. J Biol Chem 1992; 267: 18844-51.
    PubMedSearch in Google Scholar
  • 25 Peterson JA, Visentin GP, Newman PJ, Aster RH. A recombinant soluble form of the integrin αIIbβ3 (GPIIb-IIIa) assumes an active, ligand-binding conformation and is recognized by GPIIb-IIIa-specific monoclonal, allo-, auto-, and drug-dependent platelet antibodies. Blood 1998; 92: 2053-63.
    PubMedSearch in Google Scholar
  • 26 Coller BS. Blockade of platelet GPIIb/IIIa as an antithrombotic strategy. Circulation 1995; 92: 2373-80.
    CrossrefPubMedSearch in Google Scholar
  • 27 The EPIC Investigators. Use of a monoclonal antibody directed against the platelet glycoprotein IIb/IIIa receptor in high-risk coronary angioplasty. N Engl J Med 1994; 330: 956-61.
    CrossrefPubMedSearch in Google Scholar
  • 28 CAPTURE Investigators. Randomized placebo-controlled trial of abciximab before and during coronary intervention in refractory unstable angina: the CAPTURE study. Lancet 1997; 349: 1429-35.
    CrossrefPubMedSearch in Google Scholar
  • 29 Goa KL, Noble S. Eptifibatide. Drugs 1999; 57 (03) 439-62.
    CrossrefPubMedSearch in Google Scholar
  • 30 Platelet Glycoprotein IIb/IIIa in Unstable Angina: Receptor Suppression Using Integrilin Therapy (PURSUIT) Trial Investigators. Inhibition of platelet glycoprotein IIb/IIIa with eptifibatide in patients with acute coronary syndromes without persistent ST-segment elevation. N Eng Med 1998; 339: 436-43.
    CrossrefPubMedSearch in Google Scholar
  • 31 The IMPACT II Investigators. Effects of competitive platelet glycoprotein IIb/IIIa inhibition with integrelin in reducing complications of percutaneous coronary intervention. Lancet 1997; 349: 1422-28.
    CrossrefPubMedSearch in Google Scholar
  • 32 McClellan KJ, Goa KL. Tirofiban. Drugs 1998; 56 (06) 1067-80.
    CrossrefPubMedSearch in Google Scholar
  • 33 The RESTORE Investigators. Effects of platelet glycoprotein IIb/IIIa blockade with tirofiban on adverse cardiac events in patients with unstable angina. N Engl J Med 1998; 1498-1505.
    PubMedSearch in Google Scholar
  • 34 Platelet Receptor Inhibition in Ischemic Syndrome Management in Patients Limited by Unstable Signs and Symptoms (PRISM-PLUS) Study Investigators. Inhibition of the platelet glycoprotein IIb/IIIa receptor with tirofiban in unstable angina and non-Q-wave myocardial infarction. N Eng J Med 1998; 338: 1488-97.
    CrossrefPubMedSearch in Google Scholar
  • 35 Coller BS. Potential non-glycoprotein IIb/IIIa effects of abciximab. Am Heart J 1999; 138: S1-S5.
    CrossrefPubMedSearch in Google Scholar
  • 36 Simon DI, Xu H, Ortlepp S, Rogers C, Rao NK. 7E3 monoclonal antibody directed against the platelet glycoprotein IIb/IIIa cross-reacts with the leukocyte integrin Mac-1 and blocks adhesion to fibrinogen and ICAM. Arterioscler Thromb Vasc Biol 1997; 17: 528-35.
    CrossrefPubMedSearch in Google Scholar
  • 37 Frelinger III AL, Du X, Plow EF, Ginsberg MH. Monoclonal antibodies to ligand-occupied conformers of integrin αIIbβ3 (glycoprotein IIb-IIIa) alter receptor affinity, specificity, and function. J Biol Chem 1991; 266: 6346-52.
    PubMedSearch in Google Scholar
  • 38 Fitzgerald LA, Leung B, Phillips DR. A method for purifying the platelet membrane glycoprotein IIb-IIIa complex. Anal Biochem 1985; 151: 169-77.
    CrossrefPubMedSearch in Google Scholar
  • 39 Daniel WW. Biostatistics. A Foundation for Analysis in the Health Sciences. 7th Edition.. New York: Wiley; 1995
    Search in Google Scholar
  • 40 Coller BS, Peerschke EI, Scudder LE, Sullivan CA. A murine monoclonal antibody that completely blocks the binding of fibrinogen to platelets produces a thrombasthenic-like state in normal platelets and binds to glycoproteins IIb and/or IIIa. J Clin Invest 1983; 72: 325-38.
    CrossrefPubMedSearch in Google Scholar
  • 41 Wittig K, Rothe G, Schmitz G. Inhibition of fibrinogen binding and surface recruitment of GPIIb/IIIa as dose-dependent effects of the RGD-mimetic MK-852. Thromb Haemost 1998; 79: 625-30.
    Thieme ConnectPubMedSearch in Google Scholar
  • 42 Peter K, Schwarz M, Ylanne J, Kohler B, Moser M, Nordt T, Salbach P, Kubler W, Bode C. Induction of fibrinogen binding and platelet aggregation as a potential intrinsic property of various glycoprotein IIb/IIIa (αIIbβ3) inhibitors. Blood 1998; 92: 3240-9.
    PubMedSearch in Google Scholar
  • 43 Mondoro TH, Wall CD, White MM, Jennings LK. Selective induction of a GPIIIa LIBS by fibrinogen and von Willebrand Factor. Blood 1996; 88: 3824-30.
    PubMedSearch in Google Scholar