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γ T ‐ S 195 A thrombin reduces the anticoagulant effects of dabigatran in vitro and in vivo
Author(s) -
Sheffield W. P.,
Lambourne M. D.,
EltringhamSmith L. J.,
Bhakta V.,
Arnold D. M.,
Crowther M. A.
Publication year - 2014
Publication title -
journal of thrombosis and haemostasis
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 1.947
H-Index - 178
eISSN - 1538-7836
pISSN - 1538-7933
DOI - 10.1111/jth.12601
Subject(s) - dabigatran , in vivo , thrombin , pharmacology , direct thrombin inhibitor , in vitro , antithrombins , discovery and development of direct thrombin inhibitors , medicine , anticoagulant , chemistry , warfarin , biochemistry , atrial fibrillation , platelet , biology , microbiology and biotechnology
Summary Background Dabigatran etexilate ( DE ) is an oral direct thrombin inhibitor used to prevent strokes in patients with atrial fibrillation. No licensed DE antidote is currently available. We hypothesized that active site‐mutated S 195 A thrombin ( S 195 A ‐ II a) and/or its trypsinized derivative (γ T ‐ S 195 A ‐ II a) would sequester dabigatran, the active form of DE , and reduce its anticoagulant effects. Objective To assess active site‐mutated S 195 A or γ T ‐ S 195 A ‐ II a as dabigatran reversal agents in vitro and in vivo . Methods Diluted t hrombin t ime (d TT ) assays were performed using human or murine plasma containing dabigatran, combined with S 195 A ‐ II a, γ T ‐ S 195 A ‐ II a or FPR ‐chloromethyl ketone‐treated thrombin ( FPR ‐ II a). Bleeding times were determined in anesthetized DE ‐treated mice also receiving γ T ‐ S 195 A ‐ II a or vehicle 15 min prior to tail transection. The time to occlusion of carotid arteries of DE ‐treated mice also receiving S 195 A ‐ II a, γ T ‐ S 195 A ‐ II a, prothrombin complex concentrate ( PCC ) or vehicle, 15 min prior to topical F e C l 3 , was determined using D oppler ultrasound. Results γ T ‐ S 195 A ‐ II a reduced d TT values of dabigatran‐containing human and murine plasma more effectively than S 195‐ II a; FPR ‐ II a had no effect. A dose of 13 mg kg −1 DE abrogated occlusive thrombus formation in the carotid arteries of F e C l 3 ‐treated mice; γ T ‐ S 195 A ‐ II a (6 mg kg −1 ) or PCC (14.3 IU kg −1 ), but not saline vehicle or S 195 A ‐ II a (6 mg kg −1 ), was equally effective in restoring thrombus formation. Bleeding times of mice treated with 60 mg kg −1 DE and γ T ‐ S 195 A ‐ II a (6 mg kg −1 ) or saline vehicle did not differ. Conclusions Our data suggest that γ T ‐ S 195 A ‐ II a decreases the anticoagulant effects of dabigatran in vitro and is partially effective at restoring hemostasis‐related thrombus formation in DE ‐treated mice in vivo .