HD1, a thrombin- and prothrombin-binding DNA aptamer, inhibits thrombin generation by attenuating prothrombin activation and thrombin feedback reactions.
HD1, a DNA aptamer, binds exosite 1 on thrombin and blocks its clotting activity. Because HD1 also binds prothrombin and inhibits its activation by prothrombinase, we hypothesised that HD1 would be a more potent inhibitor of coagulation than other exosite 1-directed ligands, such as Hir(54-65)(SO(3)(-)). Supporting this concept, the effect of HD1 on the prothrombin time and activated partial thromboplastin time was two-fold greater than that of Hir(54-65)(SO(3)(-)) even though both agents inhibited thrombin-mediated factor (F) V and FVIII activation to a similar extent. In thrombin generation assays, HD1 (a) delayed the lag time, (b) reduced peak thrombin concentration, and (c) decreased endogenous thrombin potential to a greater extent than Hir54-65(SO(3)(-)). To eliminate thrombin feedback, studies were repeated in FV- and/or FVIII-deficient plasma supplemented with FVa and/or FVIIIa. Only HD1 prolonged the lag time in FV- and FVIII-deficient plasma supplemented with FVa and FVIIIa. In contrast, HD1 and Hir54-65(SO(3)(-)) inhibited the lag time in FVIII-deficient plasma supplemented with FVIIIa and in normal plasma. The more potent anticoagulant properties of HD1, therefore, reflect its capacity to attenuate FV activation by thrombin and inhibit prothrombinase assembly. These findings identify prothrombin as a potential target for new anticoagulants.
Kretz
Colin A
CA
Department of Medicine, McMaster University and Henderson Research Centre, Hamilton, Ontario, Canada.
Cuddy
Karl K
KK
Stafford
Alan R
AR
Fredenburgh
James C
JC
Roberts
Robin
R
Weitz
Jeffrey I
JI
D000925:Anticoagulants / Q000378:metabolism / Q000494:pharmacology*; D000991:Antithrombins / Q000378:metabolism; D052157:Aptamers, Nucleotide / Q000378:metabolism / Q000494:pharmacology*; D001665:Binding Sites; D001777:Blood Coagulation / Q000187:drug effects*; D004305:Dose-Response Relationship, Drug; D015536:Down-Regulation; D015944:Factor VIIIa / Q000378:metabolism; D015943:Factor Va / Q000378:metabolism; D025461:Feedback, Physiological; D015844:Heparin Cofactor II / Q000378:metabolism; D006629:Hirudins / Q000378:metabolism / Q000494:pharmacology*; D006801:Humans; D007700:Kinetics; D009842:Oligopeptides / Q000378:metabolism / Q000494:pharmacology*; D010314:Partial Thromboplastin Time; D011516:Prothrombin / Q000378:metabolism*; D011517:Prothrombin Time; D013431:Sulfates / Q000378:metabolism / Q000494:pharmacology*; D013917:Thrombin / Q000378:metabolism*; D013925:Thromboplastin / Q000378:metabolism
D000925:Anticoagulants; D000991:Antithrombins; D052157:Aptamers, Nucleotide; D006629:Hirudins; D009842:Oligopeptides; C047932:SERPIND1 protein, human; D013431:Sulfates; C081676:thrombin aptamer; D015943:Factor Va; D015944:Factor VIIIa; D015844:Heparin Cofactor II; D011516:Prothrombin; D013925:Thromboplastin; D013917:Thrombin