Anticoagulants (thrombin inhibitors) and aspirin synergize with P2Y12 receptor antagonism in thrombosis.
BACKGROUND
This study was designed to determine whether (1) P2Y12 antagonism synergizes with other antithrombotics and (2) anticoagulants (thrombin inhibitors) affect the antithrombotic activity elicited by P2Y12 antagonism.


RESULTS
Thrombosis was achieved by perfusion of human and murine blood through type III collagen-coated capillaries at arterial shear rate. CT50547, a direct-acting P2Y12 antagonist, inhibited thrombosis in PPACK- but not heparin-anticoagulated human blood. In contrast, CT50547 inhibited thrombosis in aspirin-treated individuals independently of the anticoagulant. Thrombin and TXA2 also synergized with P2Y12 in the absence of anticoagulation, because combined treatment of aspirin or C921-78 (a factor Xa inhibitor) with CT50547 or 2-MeSAMP (a P2Y12 antagonist) inhibited the thrombotic process, whereas all treatments failed to inhibit thrombosis when used individually. Synergism was also observed ex vivo when P2Y12-deficient (P2Y12-/-) mice were administered aspirin or coagulation inhibitors (C921-78 and bivalirudin). Finally, using intravital microscopy, we found that both C921-78 and bivalirudin abrogated the thrombotic process in P2Y12+/- mice, whereas each showed only partial efficacy in P2Y12+/+ animals.


CONCLUSIONS
Our study indicates that (1) thrombin inhibitors and aspirin have a demonstrable synergy of antithrombotic activity with P2Y12 antagonism and (2) the in vitro analysis of the antithrombotic activity of P2Y12 antagonists is affected by the anticoagulant used for blood collection. This suggests that the antithrombotic potential of P2Y12 antagonists in vitro may be overestimated in anticoagulated samples of blood and best achieved in vivo by the inclusion of aspirin and/or a thrombin inhibitor.
André
Patrick
P
Millennium Pharmaceuticals, Inc, 256 E Grand Ave, South San Francisco, Calif 9408, USA.
LaRocca
Thomas
T
Delaney
Suzanne M
SM
Lin
Pei Hua
PH
Vincent
Diana
D
Sinha
Uma
U
Conley
Pamela B
PB
Phillips
David R
DR
D000590:Amino Acid Chloromethyl Ketones / Q000494:pharmacology; D000818:Animals; D000925:Anticoagulants / Q000494:pharmacology*; D000991:Antithrombins / Q000494:pharmacology*; D001241:Aspirin / Q000494:pharmacology*; D001777:Blood Coagulation / Q000187:drug effects; D024061:Collagen Type III / Q000378:metabolism; D015194:Diffusion Chambers, Culture; D004357:Drug Synergism; D000077542:Eptifibatide; D065427:Factor Xa Inhibitors; D006493:Heparin / Q000494:pharmacology; D006571:Heterocyclic Compounds / Q000494:pharmacology; D006801:Humans; D066298:In Vitro Techniques; D008565:Membrane Proteins; D051379:Mice; D018345:Mice, Knockout; D018715:Microscopy, Video; D010455:Peptides / Q000494:pharmacology; D010477:Perfusion; D010974:Platelet Aggregation / Q000187:drug effects; D058919:Purinergic P2 Receptor Antagonists; D018048:Receptors, Purinergic P2 / Q000172:deficiency / Q000235:genetics; D058925:Receptors, Purinergic P2Y12; D013314:Stress, Mechanical; D013844:Thiazoles / Q000494:pharmacology; D013917:Thrombin / Q000037:antagonists & inhibitors; D013927:Thrombosis / Q000517:prevention & control*
D000590:Amino Acid Chloromethyl Ketones; D000925:Anticoagulants; D000991:Antithrombins; C435072:CT 50547; D024061:Collagen Type III; D065427:Factor Xa Inhibitors; D006571:Heterocyclic Compounds; D008565:Membrane Proteins; C548576:P2RY12 protein, human; C548577:P2ry12 protein, mouse; D010455:Peptides; D058919:Purinergic P2 Receptor Antagonists; D018048:Receptors, Purinergic P2; D058925:Receptors, Purinergic P2Y12; D013844:Thiazoles; D006493:Heparin; D013917:Thrombin; C040308:phenylalanyl-prolyl-arginine-chloromethyl ketone; D000077542:Eptifibatide; D001241:Aspirin