COMPARISON OF THE ANTITHROMBOTIC, ANTICLOTTING AND ANTIPLATELET AGGREGATORY ACTIVITIES OFLOW MOLECULAR WEIGHT RD HEPARIN WITH HEPARIN

1987 ◽  
Author(s):  
R L Fenichel ◽  
W Carmint ◽  
B Small ◽  
J Willis
Keyword(s):  
Human Platelet ◽  
Ex Vivo ◽  
Platelet Rich Plasma ◽  
Subcutaneous Administration ◽  
Factor Xa ◽  
Duration Of Action ◽  
Antithrombotic Activity ◽  
Significant Difference ◽  
Initial Comparison

An initial comparison of in vitro plasma anti-factor Xa (anti Xa) and activated partial thromboplastin time (APTT) values of RD heparin with heparin based upon USP units shows increased anti Xa and decreased APTT activity of RD heparin. An ex vivo experiment in rabbits in which 100 USP units/kg of RD heparinand 200 USP units/kg of heparin, when given by the subcutaneous route, reflects the significantly increased anti Xa activity generated by RD heparin as wellas its longer duration of action. No significant difference in APTT activity was observed for the two heparins, but an increased anti Xa/APTT ratio (greaterthan two) was observed for the RD heparin. Heparin (10 yg/ml), but not RD heparin, potentiated adenosinediphosphate (ADP) induced platelet aggregation in human platelet rich plasma. Subcutaneous administration of RD heparin or heparin to rabbits over the dosage range of 0.75 to 1.75 mg/kg gives similar mean dose response lines for these heparins in the thrombosis-stasis model as measured by the extent of jugular vein clotting. Administration of these heparins to rabbits on a USP unitage basis shows a significantly greater antithrombotic effect for RD heparin at 120 and 160 USP units/kg. Moreover, this experiment indicates that if the optimum dosage of 120 USP units/kgfor RD heparin is exceeded then we begin to see an indication of loss of antithrombotic activity.

Effects Of Bay g 6575 On Platelets And On Vascular PGI2 Production

1981 ◽  
Author(s):  
D E MacIntyre ◽  
E W Salzman
Keyword(s):  
Vascular Tissue ◽  
Ex Vivo ◽  
Platelet Rich Plasma ◽  
Duration Of Action ◽  
Antithrombotic Activity ◽  
Vascular Rings ◽  
And Control ◽  
Induced Aggregation

Bay g 6575 (1-[2-(β-naphthyloxy) ethyl]-3-methy1-2-pyrazolin-5-one) exerts a protective effect in several animal models of thrombosis. To elucidate its mechanism of action, we examined the effects of Bay g 6575 on platelets and on vascular PGI2 production. In vitro addition of Bay g 6575 (200 μM) to human citrated platelet rich plasma (PRP) did not inhibit aggregation induced by ADP or U44069, or augment inhibition of ADP-induced aggregation by PGD2, PGE1, PGI2 or papaverine. When added to isolated human or rat vascular rings, Bay g 6575 (200 μM) did not stimulate production of PGI2 or 6-oxo-PGF1α. Ex vivo studies one hour after administration of Baya g 6575 to rabbits (10 mg/kg, i.a.) or rats (100 mg/kg, p.o.) revealed no inhibition of ADP-induced aggregation or enhancement of the level of “circulating” PGI2 as measured by bio-immunoassay. When production of anti-aggregating activity by vascular rings from Bay g 6575 treated (B) and Control (C) rats were compared, in 6 of 8 experiments B inhibited more than C and B produced more 6-oxo-PGF1α than C (mean increase in B ± s.d.=74.3 ± 35.7%, range 42-135%). Production of antiaggregatory activity by “exhausted” C rings was enhanced by B>C platelet free plasma. In all cases, the inhibitor of aggregation produced by B and C rings acted on both human and rat PRP, and its effects could be reversed by anti-PGI1 antibodies that neutralize PGI2>6-oxo-PGE1>PGD2. When exogenous PGI2 was incubated with (exhausted) aspirin treated vascular rings, the duration of action of PGI2 was longer in the presence of B rings than C rings.Bay g 6575 has no direct effects on platelets or on vascular tissue. Its antithrombotic activity appears to be caused by regulation of PGI2 synthesis and metabolism, an effect mediated by factors, possibly Bay g 6575 metabolites, present in plasma after in vivo administration.

scholarly journals Antithrombotic Activity of a Potent, New Agent, 6,7-Dichloro-1,2,3,5-Tetrahydroimidazo[2,1-B] Quinazolin-2-One Hydrochloride Monohydrate

1977 ◽  
Author(s):  
J. S. Fleming ◽  
J. P. Buyniski
Keyword(s):  
Platelet Aggregation ◽  
Animal Models ◽  
Ex Vivo ◽  
Platelet Rich Plasma ◽  
Oral Dosage ◽  
Antithrombotic Agent ◽  
Duration Of Action ◽  
Antithrombotic Activity ◽  
Small Vessels ◽  
Hydrochloride Monohydrate

A potent, new anti-thrombotic agent, 6 ,7-dichloro-l,2,3,5-tetrahydroimidazo[2,1-b]quinazolin-2-one hydrochloride monohydrate (BL-4162A) has been evaluated for activity against induced platelet aggregation in a series of in vitro and ex vivo experiments using platelet rich plasma (PRP) from various species including man. In addition, the compound’s potential utility as an antithrombotic agent has been tested in various in vivo animal models including two models of induced thrombosis involving both large and small vessels. Aggregometry was employed to test the ability of BL-4162A to inhibit platelet aggregation induced by aggregating agents such as ADP, collagen, thrombin and antigen-antibody complexes. The compound’s antithrombotic activity was evaluated in small vessels using the biolaser-rabbit ear chamber technique while the effect of BL-4162A on large vessel thrombosis was assessed against electrically-induced carotid artery thrombosis in dogs. Results indicate that BL-4162A inhibits platelet aggregation in the range of 0.08 to 0.68 pg/ml. Platelet antiaggregating activity was also observed in ex vivo aggregometry studies in dogs and rats at oral doses in the range of 1 to 10 mg/kg. Of particular interest, is the fact that BL-4162A effectively inhibited thrombosis in both animal models employed over the same oral dosage range. Results of the above investigations as well as an appreciable duration of action suggest that BL-4162A may be an important candidate for clinical evaluation in thromboembolism.

scholarly journals Synthesis and Thrombin, Factor Xa and U46619 Inhibitory Effects of Non-amidino and Amidino N2-Thiophenecarbonyl- and N2-Tosylanthranilamides

2017 ◽  
Author(s):  
Soo Hyun Lee ◽  
Wonhwa Lee ◽  
Nguyen Thi Ha ◽  
Il Soo Um ◽  
Jong-Sup Bae ◽  
...  
Keyword(s):  
Platelet Aggregation ◽  
Bleeding Time ◽  
Ex Vivo ◽  
Factor Xa ◽  
Inhibitory Effects ◽  
Human Endothelial Cells ◽  
Antithrombotic Activity ◽  
Pharmacological Targets

Thrombin (factor IIa) and factor Xa (FXa) are key enzymes at the junction of the intrinsic and extrinsic coagulation pathways and are the most attractive pharmacological targets for the development of novel anticoagulants. Twenty non-amidino N2-thiophencarbonyl- and N2-tosyl anthranilamides 1-20 and six amidino N2-thiophencarbonyl- and N2-tosylanthranilamides 21-26 were synthesized and evaluated prothrombin time (PT) and activated partial thromboplastin time (aPTT) using human plasma at concentration 30 μg/mL in vitro. From these results, compounds 5, 9, and 21-23 were selected to study the further antithrombotic activity. The anticoagulant properties of 5, 9, and 21-23 significantly exhibited a concentration-dependent prolongation of in vitro PT and aPTT, in vivo bleeding time, and ex vivo clotting time. These compounds concentration-dependently inhibited the activities of thrombin and FXa and inhibited the generation of thrombin and FXa in human endothelial cells. In addition, data showed that 5, 9, and 21-23 significantly inhibited thrombin catalyzed fibrin polymerization and mouse platelet aggregation and inhibited platelet aggregation induced U46619 in vitro and ex vivo. N-(3'-Amidinophenyl)-2-((thiophen-2''-yl)carbonyl amino)benzamide (21) was most active.

The Effect of the Synthetic Pentasaccharide SR 90107/ORG 31540 on Thrombin Generation Ex Vivo Is Uniquely due to ATIII-Mediated Neutralization of Factor Xa

1995 ◽  
Vol 74 (06) ◽  
pp. 1474-1477 ◽  
Author(s):  
J C Lormeau ◽  
J P Herault
Keyword(s):  
Thrombin Generation ◽  
Graphical Representation ◽  
Ex Vivo ◽  
Subcutaneous Administration ◽  
Factor Xa ◽  
Significant Rise ◽  
Time Courses ◽  
The One

SummaryThe inhibition of thrombin generation (TG) was studied in plasma from human volunteers after single subcutaneous administrations of 4000, 8000 or 12,000 anti-Xa units (i.e., 6, 12 or 18 mg) of the synthetic pentasaccharide (SR 90107/ORG 31540) (SP).SP impaired TG in plasma for up to 18 h after injection, and the time-courses of TG and factor Xa inhibitions were similar.In untreated plasma supplemented in vitro with SP to obtain the same anti-Xa activity as in ex vivo samples, equivalent TG inhibitions were observed thus showing that no transformed SP molecules were involved in the TG inhibition ex vivo.Functional as well as immunological assay of TFPI indicated that subcutaneous injection of 12,000 anti-Xa units of SP did not induce any TFPI release, whereas under the same conditions, 13,000 IU of Fraxiparine® produced a significant rise of TFPI in plasma.The plotting of TG inhibition versus SP concentration could be fitted with a good correlation (r = 0,94) to the graphical representation linking [ATIII-SP] to [SP].These results demonstrate that following subcutaneous administration to man, SP inhibits TG ex vivo and likely in vivo exclusively through the same selective ATIII-mediated inhibition of factor Xa as the one elicited in vitro.

SR123781A, a Synthetic Heparin Mimetic

2001 ◽  
Vol 85 (05) ◽  
pp. 852-860 ◽  
Author(s):  
J. P. Hérault ◽  
A. Bernat ◽  
P. Savi ◽  
P. Schaeffer ◽  
P. A. Driguez ◽  
...  
Keyword(s):  
Ex Vivo ◽  
Thrombus Formation ◽  
Vena Cava ◽  
Subcutaneous Administration ◽  
Factor Xa ◽  
Inhibitory Effect ◽  
Binding Domain ◽  
Heparin Binding ◽  
Venous Shunt

SummarySR123781A, a synthetic hexadecasaccharide comprising an anti-thrombin (AT) binding domain, a thrombin binding domain, and a neutral methylated hexasaccharide sequence, was obtained from glucose through a convergent synthesis. SR123781A showed high affinity for human AT (Kd = 58 ± 22 nM) and was a potent catalyst of its inhibitory effect with regard to factor Xa (IC50 = 77 ± 5 ng/ml – 297 ± 13 U/mg) and thrombin (IC50 = 4.0 ± 0.5 ng/ml – 150 ± 30 U/mg). SR123781A which acted exclusively via AT (no effect via heparin cofactor II at a concentration of 6 g/ml) inhibited thrombin generation occurring via both the extrinsic and intrinsic pathways in vitro in human plasma. SR123781A did not compete for 3H-heparin binding to PF4 and did not activate platelets in the presence of plasma from patients with heparin-induced thrombocytopenia. After intravenous or subcutaneous administration to rats, rabbits or baboons, SR123781A displayed prolonged anti-factor Xa and anti-factor IIa activity ex vivo. After intravenous injection to baboons, decreases of the anti-factor Xa and anti-thrombin activities were parallel and disappeared with the same pharmacodynamics. Intravenous administrations of SR123781A strongly inhibited thrombus formation in an experimental model of thromboplastin-induced venous thrombosis in rats with an ED50 value of 18 ± 0.1 g/kg (vs 77 ± 3 g/kg for heparin). SR123781A inhibited arterial thrombus formation induced on a silk thread in an arterio-venous shunt and in the vena cava (ED50 values of 225 ± 10 and 27 ± 8 g/kg, respectively). Compared to standard and low molecular weight heparin and to presently used drugs, SR123781A exhibited a highly favourable anti-thrombotic/bleeding ratio therefore showing that it might be considered as a promising compound in the treatment and prevention of various thrombotic diseases.

Effect of SC 38249, a Novel Substituted Imidazole, on Platelet Aggregation In Vitro and In Vivo

1988 ◽  
Vol 59 (02) ◽  
pp. 164-170 ◽  
Author(s):  
N Lad ◽  
A C Honey ◽  
D O Lunt ◽  
R F G Booth ◽  
J Westwick ◽  
...  
Keyword(s):  
Inhibitory Activity ◽  
Light Transmission ◽  
Human Platelet ◽  
Ex Vivo ◽  
Platelet Rich Plasma ◽  
Thromboxane A2 ◽  
Induced Responses ◽  
Maximum Increase

SummarySC 38249 ((RS)-l-(2,3-bis-[(4-methoxyphenyl)methoxy] propyl)-lH-imidazole) caused dose-related inhibition of collagen- induced thromboxane A2 formation in human platelet rich plasma (IC50: 9.9 ± 1.0 μM) accompanied by a dose-dependent increase in plasma PGE2. Broad inhibitory activity was evident against human platelet aggregatory and secretory responses in vitro.IC50 values of 11.9 ± 1.9 μM (0.64 mM arachidonic acid), 18.3 ± 3.8 μM (0.5 μg ml−-1collagen) and 37.6 ± 6.1 μM (25 nM Paf-acether) were obtained against maximum increase in PRP light transmission achieved by each agonist. Although less potent, SC 38249 retained significant inhibitory activity against PRP responses induced by a higher (3.0 μg ml−-1) concentration of collagen (IC50: 272.5 ± 24.6 μM), and against Paf-acether-induced responses in PRP pre-treated with 10 μM indomethacin (I.C.50: 192.0 ± 16.1 μM).Experimental animal studies confirmed the in vitroanti-aggregatory efficacy of SC 38249, since significant inhibitory activity was observed against Paf-acether and ADP-induced responses in dog PRP ex vivo,anti-Forssman antibody-induced thrombocytopoenia in anaesthetized guinea pigs, and collagen-induced intravascular aggregation in anaesthetized rabbits. Thus, SC 38249 is a novel thromboxane synthase inhibitor which possesses interesting anti-aggregatory properties which cannot wholly be attributed to prevention of platelet thromboxane A2 formation.

Anticoagulant Efficacy and Immunogenicity of the Selective Factor Xa Inhibitor Antistasin Following Subcutaneous Administration in the Rhesus Monkey

1992 ◽  
Vol 67 (03) ◽  
pp. 371-376 ◽  
Author(s):  
Christopher T Dunwiddie ◽  
Elka M Nutt ◽  
George P Vlasuk ◽  
Peter K S Siegl ◽  
Linda W Schaffer
Keyword(s):  
Rhesus Monkey ◽  
Neutralizing Antibodies ◽  
Ex Vivo ◽  
Anticoagulant Activity ◽  
Subcutaneous Administration ◽  
Factor Xa ◽  
Specific Factor ◽  
Factor Xa Inhibitor ◽  
Duration Of Action ◽  
Monkey Model

SummaryThe antithrombotic efficacy and duration of action of a single subcutaneous administration of the selective factor Xa inhibitor recombinant antistasin (rATS) was evaluated in a rhesus monkey model of mild disseminated intravascular coagulation. rATS (1 mg/kg) was shown to be fully effective and comparable to standard heparin (1,000 U/kg) in the suppression of thromboplastin-induced fibrinopeptide A generation for at least 5 h following a single subcutaneous administration. The absorption rate of rATS, as measured by ex vivo activated partial thromboplastin times (aPTT), mirrored that of standard heparin exhibiting peak anticoagulant activity between 1 and 2 h post administration. The anticoagulant effects of a single rATS dose lasted for longer than 30 h maintaining an aPTT value at least 2-fold higher than baseline. Repeated subcutaneous administrations of rATS resulted in the generation of fully neutralizing antibodies. These results suggest that specific factor Xa inhibition may be as effective as standard heparin in the treatment of venous thrombosis. Due to its antigenicity however, rATS is probably not suitable for chronic subcutaneous anticoagulant therapy.

ANTITHROMBOTIC ACTIVITY OF A SYNTHETIC HEPARIN PENTASACCHARIDE IN A RABBIT STASIS THROMBOSIS MODEL USING DIFFERENT THROMBOGENIC CHALLENGES

1987 ◽  
Author(s):  
J M Walenga ◽  
J Fareed ◽  
M Petitou ◽  
J C Lormeau ◽  
M Samama ◽  
...  
Keyword(s):  
Prothrombin Complex Concentrate ◽  
Ex Vivo ◽  
Factor Xa ◽  
Antithrombotic Effect ◽  
Antithrombotic Activity ◽  
Thrombosis Model ◽  
Activated Prothrombin Complex Concentrate ◽  
Dose Dependent

The synthetic pentasccharide, representing the critical sequence required in heparin for binding to antithrombin III, provides a unique tool to study the question of whether an agent solely capable of inhibiting factor Xa but devoid of anti-factor Ila activity in vitro, has the capacity to produce an antithrombotic effect in vivo. We have previously demonstrated in a rabbit stasis thrombosis model using a human serum challenge, a significant antithrombotic effect of the pentasaccharide (Walenga et al., Thromb Res 43:243, 1986). To extend and confirm these studies, four modifications of the stasis thrombosis model were developed using more specified induction sites of thrombosis. The following thrombogenic challenges were selected: monkey brain thromboplastin, an activated prothrombin complex concentrate, a non-activated prothrombin complex concentrate administered simultaneously with Russell's viper venom, and factor Xa. Dose-dependent antithrombotic responses were obtained in all four systems with ED50 values between 25-43 ug/kg for pentasaccharide as compared to 16-47 ug/kg for heparin. Complete inhibition of induced thrombosis was obtained in all four systems for pentasaccharide. Ex vivo analysis revealed expected anti-factor Xa levels but no anti-factor IIa activity. It was also shown that pentasaccharide in the rabbit was capable of inhibiting the generation of thrombin without directly inhibiting formed thrombin. It is concluded that an oligosaccharide with high anti-factor Xa activity, devoid of anti-factor Ila activity, is capable of inhibiting thrombosis induced in rabbit stasis models, but that higher dosages than heparin are required for this effect-in terms of anti-factor Xa activity.

Phospholipase C from Clostridium Perfringens Induces Human Platelet Aggregation in Plasma

1988 ◽  
Vol 59 (02) ◽  
pp. 236-239 ◽  
Author(s):  
Giovanna Barzaghi ◽  
Chiara Cerletti ◽  
Giovanni de Gaetano
Keyword(s):  
Platelet Aggregation ◽  
Receptor Antagonist ◽  
Phospholipase C ◽  
Clostridium Perfringens ◽  
Human Platelet ◽  
Ex Vivo ◽  
Platelet Rich Plasma ◽  
Platelet Activating Factor ◽  
Inhibitory Effect ◽  
Thromboxane Receptor Antagonist

SummaryWe studied the aggregating effect of different concentrations of phospholipase C (PLC) (extracted from Clostridium perfringens) on human platelet-rich plasma (PRP). PRP was preincubated with PLC for 3 min at 37° C and the platelet aggregation was followed for 10 min. The threshold aggregating concentration (TAG) of PLC was 3-4 U/ml.We also studied the potentiation of PLC with other stimuli on platelet aggregation. Potentiating stimuli, such as arachidonic acid (AA), ADP. Platelet Activating Factor (PAF) and U-46619 (a stable analogue of cyclic endoperoxides) were all used at subthreshold concentrations. We also studied the possible inhibitory effect of aspirin, apyrase, TMQ, a prostaglandin endoper- oxide/thromboxane receptor antagonist and BN-52021, a PAF receptor antagonist. Only aspirin and apyrase were able to reduce aggregation induced by PLC alone and PLC + AA and PLC + ADP respectively. TMQ and BN-52021 were inactive. In ex vivo experiments oral aspirin (500 mg) partially inhibited platelet aggregation induced by PLC alone, PLC + AA and PLC + ADP 2 and 24 h after administration. Aspirin 20 mg for 7 days also reduced aggregation induced by PLC + AA.

Reaction of Acetaldehyde with Human Platelets

1992 ◽  
Vol 67 (01) ◽  
pp. 126-130 ◽  
Author(s):  
Olivier Spertini ◽  
Jacques Hauert ◽  
Fedor Bachmann
Keyword(s):  
Platelet Aggregation ◽  
Inhibitory Action ◽  
Ex Vivo ◽  
Platelet Rich Plasma ◽  
Shape Change ◽  
Reaction Medium ◽  
Human Platelets ◽  
Membrane Glycoproteins ◽  
Neutral Ph

SummaryPlatelet function defects observed in chronic alcoholics are not wholly explained by the inhibitory action of ethanol on platelet aggregation; they are not completely reproduced either in vivo by short-term ethanol perfusion into volunteers or in vitro by the addition of ethanol to platelet-rich plasma. As acetaldehyde (AcH) binds to many proteins and impairs cellular activities, we investigated the effect of this early degradation product of ethanol on platelets. AcH formed adducts with human platelets at neutral pH at 37° C which were stable to extensive washing, trichloracetic acid hydrolysis and heating at 100° C, and were not reduced by sodium borohydride. The amount of platelet adducts formed was a function of the incubation time and of the concentration of AcH in the reaction medium. At low AcH concentrations (<0.2 mM), platelet bound AcH was directly proportional to the concentration of AcH in the reaction medium. At higher concentrations (≥0.2 mM), AcH uptake by platelets tended to reach a plateau. The amount of adducts was also proportional to the number of exposures of platelets to pulses of 20 pM AcH.AcH adducts formation severely impaired platelet aggregation and shape change induced by ADP, collagen and thrombin. A positive correlation was established between platelet-bound AcH and inhibition of aggregation.SDS-PAGE analysis of AcH adducts at neutral pH demonstrated the binding of [14C]acetaldehyde to many platelet proteins. AcH adduct formation with membrane glycoproteins, cytoskeleton and enzymes might interfere with several steps of platelet activation and impair platelet aggregation.This in vitro study shows that AcH has a major inhibitory action on platelet aggregation and may account for the prolonged ex vivo inhibition of aggregation observed in chronic alcoholics even in the absence of alcoholemia.

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