Antifactor Xa activity and antithrombotic activity in rats of structural analogues of the minimum antithrombin III binding sequence: discovery of compounds with a longer duration of action than of the natural pentasaccharide.

The total chemical synthesis of a series of structural analogues of the so-called natural AT III binding pentasaccharide together with the natural pentasaccharide itself has been accomplished. The structural analogues all contain an extra 3-O-sulfate group on glucosamine residue H of the pentasaccharide, some carry additional 3 or 4-O-sulfate groups on glucosamine residue D. All these structural analogues elicit a higher specific anti-Factor Xa activity than the natural pentasaccharide (700 anti-Factor Xa U/kg). The structural analogue carrying only an additional 3-O-sulfate on glucosamine unit H (Org 31550) has the highest specific activity (1230 anti-Factor Xa U/kg). The increased specific activity is presumably attributed to the stronger binding to AT III. All structural analogues have a prolonged duration of action of the plasma anti-Factor Xa activity. (T1/2, approximately 9 hours) compared with that of the natural pentasaccharide (T1/2, approximately 5 hours) after single intravenous administration of 600 anti-Factor Xa U/kg. All structural analogues exert dose-dependent antithrombotic activity in a rat stasis thrombosis model after intravenous administration. On an anti-Factor Xa basis, the compounds have the same potency as the natural pentasaccharide (ED50s are 35 to 55 anti-factor Xa U/kg). Of two structural analogues (Org 31550 and Org 31706), the time course of antithrombotic activity was assessed in the same model after subcutaneous administration of 600 anti-Factor Xa U/kg. The duration of antithrombotic activity of these compounds was four to five times longer than that of the natural pentasaccharide.

Pharmacological profile of the chemically synthesized antithrombin III binding fragment of heparin (pentasaccharide) in rats.

The antithrombotic and haemostatic effects of a pentasaccharide, the chemically synthesized antithrombin III (AT-III) binding fragment of heparin (PENTA), were investigated in rats in comparison with heparin. PENTA showed a dose-dependent antithrombotic effect in three thrombosis models in which thrombus formation was induced by different triggers. PENTA was consistently less potent than heparin in these models if doses were expressed in anti-Xa U/kg but PENTA showed more or less the same potency as heparin if doses were expressed in mg/kg. The antithrombotic effect of PENTA was strongly related to its anti-Xa activity as judged from its antithrombotic potency in the various models and from the time courses of both activities. PENTA caused a dose-dependent increase in blood loss in a bleeding model but the dose response curve was rather flat; the effect of PENTA on blood loss was small compared to that of heparin. The duration of action of PENTA as measured by the plasma anti-Xa levels was long compared to that of heparin and the duration of the antithrombotic effect was that expected on the basis of the plasma anti-Xa levels. Finally, PENTA showed comparable antithrombotic activity after s.c. and i.v. administration, as expected because of the approximately 100% bioavailability of the anti-Xa activity after s.c. administration.

Safety and pharmacokinetics of the low molecular weight heparinoid Org 10172 administered to healthy elderly volunteers.

1. In a cross-over study a new low molecular weight heparinoid Org 10172 was administered to 12 elderly male and female volunteers. It was well tolerated and no adverse effects occurred. 2. The absolute bioavailability of Org 10172 as measured by plasma anti-Xa activity, glycosaminoglycuronans with no affinity to antithrombin III (NoA-GAG) and thrombin generation inhibiting activity approached 100% in both sexes. 3. The half-life of elimination of its anti-Xa activity (19.2 +/- 6.1 h) was similar to that found previously in young volunteers. Org 10172 was further characterised by a rapid disappearance from the circulation of its anti-thrombin activity (t1/2 1.8 +/- 0.6 h) and of the NoA-GAG (t1/2 3.5 +/- 2.1 h). 4. Its thrombin generation inhibiting activity was of intermediate duration (t1/2 elimination 6.2 +/- 4.0 h).

ORG 10172: a low molecular weight heparinoid anticoagulant with a long half-life in man.

ORG 10172 is a heparinoid with mean molecular weight 6500 daltons. Intravenous bolus injections of ORG 10172 were compared with placebo and heparin injections in 91 separate studies in 83 healthy male subjects. 6400 units ORG 10172 produced a mean maximum change of 14.7 s in kaolin cephalin time (c.f. greater than 120 s for 5000 units heparin). Changes in prothrombin time were minimal (1.6 s for 6400 units ORG 10172 and 4.5 s after 5000 units heparin). A dose-related increase in bleeding time occurred after ORG 10172 and at high doses (greater than 3200 units) some secondary bleeding, which was never serious, occurred at between 1 and 4 h after incision. A dose-dependent reduction in ex vivo platelet adhesiveness was found at 10 min after ORG 10172 injection. ORG 10172 promoted a much smaller release of lipoprotein lipase as compared with heparin. The effect of ORG 10172 on plasma factor Xa activity (one measure of its action) was described by a biexponential decay with a mean distribution half-life of 2.34 (s.e. mean 0.16) h and mean disposition half-life of 17.6 (s.e. mean 1.1) h. It thus has a much longer duration of effect than heparin. There was a linear relationship of plasma anti-Xa response to increasing dose although there was some variability only partly explained by differences in body weight or surface area. ORG 10172 administration by bolus intravenous injection was well tolerated and there was no evidence of adverse effects on clinical chemistry or haematology tests.

The effects of antithrombin III, heparin and a novel heparinoid infusions on the bleeding tendency using an experimental model in the rat.

The effects of selective increase of plasma AT-III concentration (to 2 U/ml and 8 U/ml) in the presence or absence of low dose commercial heparin and a novel natural heparinoid (Org 10172) on APTT, clotting factors and bleeding were investigated in an experimental model in rats. Slight increases in the APTT were observed with: a. Org 10172, b. both AT-III doses and c. combinations hereoff. However, synergism was observed in the prolongations of the APTT when the AT-III concentrate (70 U/kg and 500 U/kg) was combined with both heparin dosages. AT-III concentrates (70 U/kg and 500 U/kg) or Org 10172 (2,5 mg/kg) separately or in combination showed no effect on bleeding. Heparin (0, 125 and 0,25 mg/kg i.v.) also showed no effect on bleeding neither if they were combined with the lower dose AT-III. However, heparin (both dosages) combined with 500 U/kg of AT-III concentrates significant increased blood loss. These observations suggest that infusion of AT-III concentrate additionally to low dose heparin therapy did not increase bleeding in the rat model used provided that extreme high AT-III plasma levels (8 U/ml) were avoided. The novel natural heparinoid Org 10172 alone or combined with either AT-III dosage induced no increased bleeding.

A novel anti-thrombotic heparinoid (Org 10172) devoid of bleeding inducing capacity: a survey of its pharmacological properties in experimental animal models.

The pharmacological profile of Org 10172, a mixture of sulphated glycosaminoglycorunans derived from hog intestinal mucosa, has been assessed in experimental thrombosis and bleeding models in rats and compared with heparin USP. Org 10172 inhibited thrombus formation in arterio-venous shunts dose dependently, the dose required for 50% inhibition (ID50) of thrombus formation was 40 anti-Xa units/kg i.v. The ID50 for heparin USP was 70 anti-Xa units/kg i.v. Org 10172 hardly increased bleeding in doses upto 1600 anti-Xa units/kg i.v., whereas heparin USP dose dependently increased bleeding from 90 anti-Xa units/kg i.v. onwards. The benefit (anti-thrombotic)/risk (bleeding) ratio of Org 10172 was therefore considerably better than that of heparin USP. The improved profile of Org 10172 towards bleeding might be caused by differences in the interaction with blood platelets in comparison with heparin USP. Org 10172 had less effect on the platelet content in thrombi than heparin USP. Org 10172 did not inhibit collagen induced release of serotonin in contrast to heparin USP. Org 10172 inhibited factor Xa induced aggregation of rabbit platelets but only at anti-Xa levels which were fifteen times higher than for heparin USP. In contrast to heparin USP Org 10172 had only a very weak effect on the activated partial thromboplastin time (APTT) ex vivo.