title: AlphaIIbbeta3 priming and clustering by orally active and intravenous integrin antagonists.
abstract: BACKGROUND
  Drugs that block platelet-platelet and platelet-fibrin interactions via the alpha(IIb)beta(3) (glycoprotein IIb/IIIa) receptor are used daily in patients undergoing percutaneous coronary interventions. Along with expected increases in spontaneous bleeding, clinical trials have revealed a surprising increase in thrombosis when these drugs are used without other anticoagulants. A better understanding of their mechanisms can minimize these risks.
  
  
  OBJECTIVE
  This study tested the hypothesis that interventions designed to block fibrinogen binding inevitably leave the alpha(IIb)beta(3) receptor in an activated state. It compared the effects on platelet function and alpha(IIb)beta(3) conformation of the orally active compounds orbofiban and roxifiban, the i.v. agents eptifibatide and tirofiban, and echistatin, an arginine-glycine-aspartate (RGD) disintegrin.
  
  
  METHODS
  The integrin antagonist concentrations required to saturate platelets and to block platelet-platelet and platelet-fibrin interactions were determined by flow cytometry, aggregometry, and clot-based adhesion assays, respectively. Analytical ultracentrifugation measured each antagonist's effects on the solution structure of alpha(IIb)beta(3). Fluorescence anisotropy provided equilibrium and kinetic data for integrin:antagonist interactions.
  
  
  RESULTS
  Both orally active drugs bound more tightly and inhibited platelet aggregation and adhesion to fibrin more effectively than echistatin. Analytical ultracentrifugation yielded this order for perturbing alpha(IIb)beta(3) conformation (priming) and promoting oligomerization (clustering): echistatin > eptifibatide > orbofiban > tirofiban > roxifiban. Roxifiban was also most effective at disrupting the rapidly forming/slowly dissociating alpha(IIb)beta(3):echistatin complex.
  
  
  CONCLUSIONS
  Our results suggest that the same molecular mechanisms that enable glycoprotein IIb/IIIa inhibitors to bind tightly to the alpha(IIb)beta(3) receptor and block fibrinogen binding contribute to their ability to perturb the resting integrin's conformation, thus limiting the safety and efficacy of both oral and i.v. integrin antagonists.
authors.affiliation: Department of Biochemistry, Wake Forest University School of Medicine, Winston-Salem, NC 27157-1019, USA. hantgan@wfubmc.edu
authors.forename: R R
authors.initials: RR
authors.lastname: Hantgan
authors.forename: M C
authors.initials: MC
authors.lastname: Stahle
authors.forename: J H
authors.initials: JH
authors.lastname: Connor
authors.forename: R F
authors.initials: RF
authors.lastname: Connor
authors.forename: S A
authors.initials: SA
authors.lastname: Mousa
mesh_terms: D000284:Administration, Oral; D000409:Alanine / Q000494:pharmacology; D000578:Amidines / Q000494:pharmacology; D001667:Binding, Competitive; D001792:Blood Platelets / Q000187:drug effects* / Q000378:metabolism; D019281:Dimerization; D004305:Dose-Response Relationship, Drug; D000077542:Eptifibatide; D005260:Female; D005340:Fibrinogen / Q000378:metabolism; D006801:Humans; D066298:In Vitro Techniques; D007275:Injections, Intravenous; D036341:Intercellular Signaling Peptides and Proteins; D007555:Isoxazoles / Q000494:pharmacology; D007700:Kinetics; D008297:Male; D008958:Models, Molecular; D010455:Peptides / Q000494:pharmacology; D015539:Platelet Activation / Q000187:drug effects*; D010975:Platelet Aggregation Inhibitors / Q000008:administration & dosage / Q000378:metabolism / Q000494:pharmacology*; D010979:Platelet Function Tests; D019039:Platelet Glycoprotein GPIIb-IIIa Complex / Q000737:chemistry / Q000187:drug effects* / Q000378:metabolism; D011485:Protein Binding; D011487:Protein Conformation / Q000187:drug effects; D011759:Pyrrolidines / Q000494:pharmacology; D012016:Reference Values; D013329:Structure-Activity Relationship; D000077466:Tirofiban; D014443:Tyrosine / Q000031:analogs & derivatives / Q000494:pharmacology
chemical_list: D000578:Amidines; D036341:Intercellular Signaling Peptides and Proteins; D007555:Isoxazoles; D010455:Peptides; D010975:Platelet Aggregation Inhibitors; D019039:Platelet Glycoprotein GPIIb-IIIa Complex; D011759:Pyrrolidines; C057986:echistatin; D014443:Tyrosine; D005340:Fibrinogen; C120234:orbofiban; D000077466:Tirofiban; D000077542:Eptifibatide; D000409:Alanine; C118235:roxifiban