title: Inhibition of clot-bound and free (fluid-phase thrombin) by a novel synthetic thrombin inhibitor (Ro 46-6240), recombinant hirudin and heparin in human plasma.
abstract: Clot-bound thrombin remains active and is less accessible to heparin-antithrombin III than fluid-phase thrombin. To determine whether clot-bound human thrombin is more susceptible to inactivation by direct thrombin inhibitors, the activity of a novel synthetic competitive thrombin inhibitor Ro 46-6240, recombinant hirudin and unfractionated heparin were compared with fluid-phase thrombin and clot-bound thrombin. Fibrinopeptide A generated in human plasma was used as an index of thrombin activity. Hirudin was the most potent inhibitor of fluid-phase and clot-bound thrombin. However, Ro 46-6240 inhibited clot-bound thrombin three times more potently than fluid-phase thrombin (IC50 19 vs 56 ng/ml) while hirudin was two times (IC50 8 vs 3 ng/ml) and heparin six times (IC50 1,205 vs 200 ng/ml) less active against clot-bound thrombin compared with fluid-phase thrombin. The relative selectivity for clot-bound thrombin is not a unique property of Ro 46-6240 since two other synthetic thrombin inhibitors tested inhibited clot-bound thrombin more effectively than fluid-phase thrombin and a third was equally active against both forms of thrombin. In contrast, the affinities of two chromogenic substrates were similar for both forms of thrombin. This study shows that direct thrombin inhibitors inhibit clot-bound thrombin more potently than heparin and suggests that an apparent selectivity for clot-bound thrombin can be achieved with some synthetic thrombin inhibitors. Further studies have to show whether the high potency of these direct thrombin inhibitor translates into antithrombotic efficacy in clinical situations with pre-existing clots.
authors.affiliation: Preclinical Research Department, F. Hoffmann-La Roche Ltd., Basel, Switzerland.
authors.forename: A
authors.initials: A
authors.lastname: Gast
authors.forename: T B
authors.initials: TB
authors.lastname: Tschopp
authors.forename: G
authors.initials: G
authors.lastname: Schmid
authors.forename: K
authors.initials: K
authors.lastname: Hilpert
authors.forename: J
authors.initials: J
authors.lastname: Ackermann
mesh_terms: D001667:Binding, Competitive; D001777:Blood Coagulation; D002863:Chromogenic Compounds / Q000378:metabolism; D005344:Fibrinopeptide A / Q000378:metabolism; D006493:Heparin / Q000494:pharmacology; D006629:Hirudins / Q000494:pharmacology*; D006801:Humans; D009281:Naphthalenes / Q000494:pharmacology*; D010880:Piperidines / Q000494:pharmacology*; D011994:Recombinant Proteins / Q000494:pharmacology; D013917:Thrombin / Q000037:antagonists & inhibitors* / Q000378:metabolism
chemical_list: D002863:Chromogenic Compounds; D006629:Hirudins; D009281:Naphthalenes; D010880:Piperidines; D011994:Recombinant Proteins; D005344:Fibrinopeptide A; D006493:Heparin; D013917:Thrombin; C087198:napsagatran