Inhibition of the early stages of the thrombin generation reaction by various glycosaminoglycans.
Inhibition of thrombin generation was studied in strongly diluted human plasma by continuously measuring the splitting of the thrombin-specific chromogenic substrate S2288 as a function of the inhibitor concentration. To avoid the activation of clotting cascade proenzymes other than prothrombin, the thrombin generation reaction was initiated by a mixture of calcium, phospholipids and (bovine) factor Xa. Using this assay we have estimated the relative potency of the following glycosaminoglycans: heparin; a fraction of heparin with high affinity for antithrombin III (high affinity heparin); the low molecular weight heparin Fragmin; the low molecular weight heparinoid Org 10172; a fraction of Org 10172 with high affinity for antithrombin III (high affinity Org 10172) and the O-methyl derivative of the pentasaccharide, representing the minimal structure required for binding to antithrombin III. Based on concentrations expressed in amidolytic anti-Xa units, the descending order of potency observed is: Heparin approximately High Affinity Heparin greater than Fragmin greater than Org 10172 greater than High Affinity Org 10172 greater than O-methyl pentasaccharide. The more potent the glycosaminoglycan the stronger the concentration dependence of its inhibitory effect. These findings could be due to the different, additional anti-thrombin activities of these glycosaminoglycans and/or to their different anti-prothrombinase activities. With the pentasaccharide a striking saturation of the inhibition is observed, due to saturation of the antithrombin III.
Visser
A
A
Scientific Development Group, Organon International B.V., Oss, The Netherlands.
Meuleman
D G
DG
D000991:Antithrombins / Q000494:pharmacology; D002809:Chondroitin Sulfates; D002863:Chromogenic Compounds; D003871:Dermatan Sulfate; D006025:Glycosaminoglycans / Q000494:pharmacology*; D006493:Heparin / Q000494:pharmacology; D006495:Heparin, Low-Molecular-Weight / Q000494:pharmacology; D006497:Heparan Sulfate; D006801:Humans; D066298:In Vitro Techniques; D007700:Kinetics; D009842:Oligopeptides; D009844:Oligosaccharides / Q000494:pharmacology; D013917:Thrombin / Q000096:biosynthesis*
D000991:Antithrombins; D002809:Chondroitin Sulfates; D002863:Chromogenic Compounds; D003871:Dermatan Sulfate; D006025:Glycosaminoglycans; D006493:Heparin; D006495:Heparin, Low-Molecular-Weight; D006497:Heparan Sulfate; D009842:Oligopeptides; D009844:Oligosaccharides; D013917:Thrombin; C041319:isoleucyl-prolyl-arginine-4-nitroanilide; C049714:IC 831423; C035838:danaparoid