Streamlining the design of promising clinical trials: in-vitro testing of antithrombotic regimens and multiple agonists of platelet activation

Abstract

Platelets are activated in vivo by multiple agonists; however, platelet function testing in vitro has been performed predominantly with only one or two agonists of platelet activation. Greater insight into anticipated effects of antithrombotic regimens should enhance the design of successful clinical trials. To test this concept, we assessed platelet activation induced by multiple agonists and two antithrombotic regimens, unfractionated heparin (UFH) and eptifibatide compared with bivalirudin and cangrelor. Blood samples from 10 patients with coronary artery disease were spiked with pharmacologic concentrations achieved in vivo of either UFH (1.2 U/ml) and eptifibatide (1.7 microg/ml), or with bivalirudin (8 microg/ml) and cangrelor (500 nmol/l). Platelet function was assessed with the use of flow cytometry. Agonists included thrombin (50 nmol/l), adenosine diphosphate (1 micromol/l), the collagen-mimetic convulxin (5 ng/ml), and platelet-activating factor (10 nmol/l). When platelet activation was identified by the surface expression of P-selectin in response to multiple agonists, the combination of bivalirudin and cangrelor suppressed activation more than UFH and eptifibatide. When platelet activation was identified by the activation of glycoprotein IIb-IIIa (PAC-1 binding), the combination of bivalirudin and cangrelor was more effective in suppressing activation in response to thrombin and adenosine diphosphate, whereas UFH and eptifibatide more effectively prevented binding of PAC-1 when platelets were activated with the collagen-mimetic convulxin. In conclusion, bivalirudin and cangrelor suppressed platelet activation in response to diverse agonists in vitro more than UFH and eptifibatide. These results and this approach to selection of promising interventions should be helpful in streamlining the design of clinical trials.