Combination of aspirin and metoclopramide produces a synergistic antithrombotic effect in a canine model of coronary artery thrombosis

Summary— We compared the antithrombotic properties of low doses of aspirin (0.03, 0.1 mg kg⁻¹ intravenously [iv]) and metoclopramide (0.1, 0.3 mg kg⁻¹ iv) alone or in combination. The animal model chosen for this study involved the generation of cyclic flow variations (CFV) in the circumflex coronary artery of anaesthetized dogs as a result of a critical coronary stenosis associated with a controlled arterial lesion at the site of stenosis. Subsequent regular CFV represent sequential thrombus formation and embolization in the damaged vessel. Neither aspirin nor metoclopramide alone demonstrated antithrombotic properties at the doses tested. However, the combination of aspirin 0.1 mg kg⁻¹ iv and metoclopramide 0.3 mg kg⁻¹ iv produced a significant antithrombotic effect, reducing the frequency of large CFV from 6.7 ± 0.5 to 0.8 ± 0.4 cycles h⁻¹ ( P < 0.01) and increasing minimum mean coronary blood flow from 5.0 ± 1.1 to 23.7 ± 2.6 mL min⁻¹ ( P < 0.01). This result apparently reflects an antithrombotic synergism between aspirin and metoclopramide since the effects of the combination were greater than the combined effects of the individual treatments. The antithrombotic influence of metoclopramide could be due to its 5HT₂-antagonist or α₂-antagonist properties, both of which would inhibit platelet aggregation. This demonstration of a synergistic antithrombotic action of the combination of aspirin and metoclopramide is of interest since these two agents are often combined in clinical use. Its therapeutic relevance, however, remains to be established.     

Antithrombotic efficacy of thrombin inhibitor L-374,087: intravenous activity in a primate model of venous thrombus extension and oral activity in a canine model of primary venous and coronary artery thrombosis

The small molecule direct thrombin inhibitor L-374,087 was characterized across species in an in vitro activated partial thromboplastin clotting time (aPTT) assay and in vivo in rhesus monkey and dog thrombosis models. In vitro in rhesus, dog, and human plasma, L-374,087 concentrations eliciting 2-fold increases in aPTT were 0.25, 1.9, and 0.28 microM, respectively. In anesthetized rhesus monkeys, 300 microgram/kg bolus plus 12 microgram/kg/min and 300 microgram/kg bolus plus 30 microgram/kg/min L-374,087 i.v. infusions significantly reduced jugular vein thrombus extension, with both regimens limiting venous thrombus extension to 2-fold that of baseline thrombus mass compared with a 5-fold extension observed in the vehicle control group. Antithrombotic efficacy in the rhesus with the lower-dose regimen was achieved with 2.3- to 2.4-fold increases in aPTT and prothrombin time. In a conscious instrumented dog model of electrolytic vessel injury, the oral administration of two 10 mg/kg L-374,087 doses 12 h apart significantly reduced jugular vein thrombus mass, reduced the incidence of and delayed time to occlusive coronary artery thrombosis, and significantly reduced coronary artery thrombus mass and ensuing posterolateral myocardial infarct size. Antithrombotic efficacy in the dog was achieved with 1.6- to 2.0-fold increases in aPTT at 1 to 6 h after oral dosing with L-374,087. These results indicate significant antithrombotic efficacy against both venous and coronary arterial thrombosis with L-374,087 with only moderate elevations in aPTT or prothrombin time. The oral efficacy of L-374,087 characterizes this compound as a prototype for the further development of orally active direct thrombin inhibitors.     

Pharmacological profile of recombinant, human activated protein C (LY203638) in a canine model of coronary artery thrombosis

The antithrombotic activity of recombinant, human activated protein C (rh-APC, LY203638) was examined in a model of canine coronary artery thrombosis. Three doses of rh-APC (0.5, 1.0, and 2.0 mg/kg/h) were administered intravenously for 2 h. Whole blood clotting times (thrombin time, activated partial thromboplastin time), ex vivo platelet aggregation, and template bleeding times were determined. Activated partial thromboplastin time significantly increased 2- and 3.7-fold during the 2-h infusion of rh-APC (1.0 and 2.0 mg/kg/h, respectively); thrombin time did not change. Intravenous infusions of rh-APC (1.0 and 2.0 mg/kg/h) produced significant prolongations to occlusion, 186 +/- 21 and 190 +/- 22 min, respectively, compared with the vehicle and the 0.5 mg/kg/h group (86 +/- 12 and 93 +/- 17 min, respectively). Vessel patency was better at the end of the experiment in the intermediate- and high-dose groups (3 of 6 and 3 of 5 vessels, 1.0 and 2.0 mg/kg/h, respectively) compared with the vehicle and 0.5 mg/kg/h groups (0/5 and 0/6, respectively). Only the 1.0 mg/kg/h group was found to have significantly elevated template bleeding times, with peak increases seen 60 min into the drug infusion. All groups had returned to baseline values by the end of the study. There was no observed inhibition of platelet aggregation. These data demonstrate that recombinant, human activated protein C is an effective anticoagulant and antithrombotic agent in the dog.     

Potentiation of the antithrombotic effect of prostacyclin by simultaneous administration of aminophylline in a canine model of coronary artery thrombosis

The antithrombotic efficacy of prostacyclin (PGI2) when administered in conjunction with the phosphodiesterase inhibitor aminophylline was evaluated in a canine model in which coronary artery thrombosis was induced by electrical stimulation of the intimal surface of the left circumflex (LCX) coronary artery. Infusions of PGI2 (25 or 50 ng/kg/min) into the left atrial appendage and aminophylline (20 micrograms/kg/min) or ethylene diamine into the left jugular vein were initiated 10 min before the start of LCX coronary artery stimulation and continued for the 6-hr stimulation period. Every animal in the control (Tris buffer plus ethylene diamine, n = 7), PGI2 (25 ng/kg/min) only (n = 6) and aminophylline only (n = 7) groups developed completely occlusive coronary artery thrombi. In contrast, none of the animals receiving PGI2 (25 ng/kg/min) plus aminophylline or PGI2 (50 ng/kg/min) plus aminophylline underwent occlusive thrombus formation. The average thrombus mass developed in response to intimal injury of the LCX coronary artery was 57 +/- 14 mg (X +/- S.E.M.) in the control group. Aminophylline administration in conjunction with PGI2 infusion at doses of 25 and 50 ng/kg/min significantly reduced thrombus mass to 11 +/- 2 and 10 +/- 1 mg, respectively (P less than .05). PGI2 (25 ng/kg/min) plus aminophylline reduced mean arterial pressure by 12% from 116 +/- 5 to 102 +/- 4 mm Hg. These data demonstrate that the combined administration of aminophylline with low-dose PGI2 provides antithrombotic efficacy while minimizing the detrimental hemodynamic effects of large-dose PGI2 administration.     

Reduction in the incidence of thrombosis by the thromboxane synthetase inhibitor CGS 13080 in a canine model of coronary artery injury

The antithrombotic potential of the thromboxane (TX) synthetase inhibitor CGS 13080 (CGS) was studied in an anesthetized open-chest canine model of coronary artery intimal wall injury induced by the local application of a low amperage electrical current (100 microA for 6 hr). CGS was administered by i.v. infusion (1 mg/kg/min) beginning 30 min before applying the direct current to the intimal wall of the vessel. CGS did not alter basal values for heart rate, blood pressure or coronary blood flow. After 6 hr of current application to the vessel, 1 of 10 CGS-treated dogs exhibited complete thrombotic occlusion of the circumflex coronary artery compared to 8 of 10 nontreated control dogs (P less than .01). Thrombus masses within the injured left circumflex coronary artery were: Control, 25.9 +/- 4.5 mg (n = 10) and CGS, 11.0 +/- 2.8 mg (n = 10); P less than .01. The concentration of TXB2 determined ex vivo in serum from thrombin-activated whole blood was decreased by CGS administration: Control, 43.15 +/- 16.08 ng/ml (n = 9) vs. CGS, 1.72 +/- .69 ng/ml (n = 10); P less than .001. Ex vivo platelet aggregometry demonstrated that arachidonic acid (0.65 mM)-induced aggregation was reduced from a control value of 82.3 +/- 7.8% (n = 10) to 45.0 +/- 11.3% (n = 10) (P less than .05), whereas aggregation in response to ADP (5 micrograms/ml) or collagen (156 and 312 micrograms/ml) was unaffected. CGS was compared with two other TX synthetase inhibitors, U63557A and OKY1581, for the ability to divert cyclic endoperoxide metabolism to the synthesis of prostaglandin (PG) E2 and prostacyclin in response to stimulation of whole blood in vitro with collagen (25 micrograms/ml). CGS, U63557A and OKY 1581 were found to be equally effective with respect to PGE2 and 6-keto PGF1 alpha production in vitro. The data demonstrate that CGS is an effective antithrombotic agent in vivo and that it selectively inhibits arachidonic acid-induced platelet aggregation ex vivo and the formation of TXA2 in thrombin-activated whole blood.     

Acute antithrombotic effects of ximelagatran, an oral direct thrombin inhibitor, and r-hirudin in a human ex vivo model of arterial thrombosis

Summary. Background: Thrombin plays a major role in thrombus formation through activation of platelets and conversion of fibrinogen to fibrin. Objectives: To investigate the antithrombotic effects of the oral direct thrombin inhibitor (DTI) ximelagatran and the parenteral DTI r‐hirudin in humans. Subjects and methods: Healthy male volunteers randomized into four parallel groups each with 15 subjects received either ximelagatran (20, 40 or 80 mg orally) or r‐hirudin (0.4 mg kg^?1 intravenous bolus + infusion of 0.15 mg kg^?1 h^?1 for 2 h and 0.075 mg kg^?1 h^?1 for 3 h). Antithrombotic effects were assessed as changes in total thrombus area (TTA) and total fibrin area (TFA) from baseline, using the Badimon perfusion chamber model at baseline and 2 h and 5 h after drug administration. Results: Two hours postdosing, ximelagatran showed antithrombotic effects at both high and low shear rates (TTA% of mean baseline value ± SEM was 76 ± 13% and 71 ± 17% [both P < 0.05] for the 20‐mg dose, 85 ± 11% [P > 0.05] and 62 ± 15% [P < 0.05] for the 40‐mg dose and 60 ± 11% and 26 ± 7% [both P < 0.05] for the 80‐mg dose, respectively). r‐Hirudin also showed a significant antithrombotic effect at high and low shear rates (76 ± 11% [P = 0.05] and 57 ± 17% [P < 0.05] of baseline values, 2 h postdosing, respectively). The inhibitory effects on TFA were similar to those on TTA. Conclusions: The oral DTI ximelagatran shows antithrombotic effects under both high and low shear conditions. The antithrombotic effect of 40–80 mg ximelagatran appeared comparable to that of parenterally administered r‐hirudin, which has been previously demonstrated to be clinically effective in acute coronary syndromes.     

Interaction of the synthetic antithrombotic peptide P10 with thrombin: a spectroscopy study

Thrombin is a critical serine protease in the coagulation system and is widely used as a target protein for antithrombotics. Spectroscopic analysis is a simple and effective method that is used to study the interaction between small molecules and proteins. In this study, the interactions of a potential antithrombotic peptide AGFAGDDAPR (P10) with thrombin were investigated by fluorescence spectroscopy, UV-vis spectroscopy, circular dichroism, Fourier-transform infrared spectroscopy and Raman spectroscopy, respectively. The results showed that the peptide P10 bonded to thrombin via hydrogen bonding and van der Waals forces, resulting in fluorescence quenching. And, the secondary structure of thrombin changed, the β-sheet decreased, and the random coil increased. The peptide P10 bonded to proline and lysine, and changed the space structure of thrombin, resulting in inhibition of thrombin activity. The results contributed to exploration of the mechanism of this potential antithrombotic drug interaction with thrombin in order to provide a preliminary understanding of the pharmacodynamic properties of P10.

Thrombin is a critical serine protease in the coagulation system and is widely used as a target protein for antithrombotics.     

Comparison of the thrombolytic activity of the novel plasminogen activator, LY210825, to anisoylated plasminogen-streptokinase activator complex in a canine model of coronary artery thrombolysis.

We have compared the thrombolytic efficacy of a novel single-chain, recombinant tissue-type plasminogen activator variant, LY210825, containing the second kringle and serine protease domains of native tissue-type plasminogen activator, with anisoylated plasminogen-streptokinase activator complex (APSAC). Male hounds (16-22 kg) were anesthetized, the left circumflex coronary artery was isolated and an electromagnetic flow probe was placed around the artery proximal to the first main branch for the measurement of coronary blood flow. An occlusive thrombus was formed after electrolytic injury of the intima of the coronary artery. After an occlusion period of 1 hr, either LY210825 (n = 8) or APSAC (n = 6) was administered as a single i.v. injection of 0.45 mg/kg. Blood was drawn (3.8% citrate) for determination of plasma fibrinogen, plasminogen and alpha-2 antiplasmin. Time to reperfusion was significantly faster with LY210825 than with APSAC, 20 +/- 2 vs. 54 +/- 8 min, respectively. The incidence of reocclusion was similar for both agents. APSAC produced significant depletion of alpha-2 antiplasmin, plasminogen and circulating fibrinogen, whereas LY210825 caused only slight consumption of plasminogen and only small decreases in fibrinogen. After a single injection of LY210825, thrombolytic concentrations of plasminogen activator were available immediately, whereas there was a significant delay in lytic concentrations of active streptokinase-plasmin complex. Consequently, LY210825 reperfused the coronary artery faster than did APSAC. In addition, LY210825 spared plasma fibrinogen, plasminogen and alpha-2 antiplasmin and therefore, could potentially minimize the risk of bleeding complications.     

Comparative spectrum and activity of NVP-PDF386 (VRC4887), a new peptide deformylase inhibitor

The antibacterial activity of NVP-PDF386 (VRC4887), a novel peptide deformylase (PDF) inhibitor, was tested against over 1000 recent clinical isolates collected during 2001 and 2002. The MIC(50/90) (mg/L) results for NVP-PDF386 (VRC4887) were: Staphylococcus aureus (SA) 0.5/1, coagulase-negative staphylococci (CoNS) 0.5/1, Streptococcus pneumoniae 0.25/0.5, other streptococci 0.25/0.5, enterococci 1/2, Moraxella catarrhalis 0.25/0.25, Haemophilus influenzae 8/32 and Enterobacteriaceae or non-fermentative Gram-negative bacilli >32/>32 mg/L. No differences in NVP-PDF386/(VRC4887) MIC distributions were observed between methicillin-resistant (MR) S. aureus and methicillin-susceptible (MS) S. aureus, MR-CoNS and MS-CoNS, penicillin-susceptible and non-susceptible streptococci, and macrolide-susceptible and -resistant strains. The potency of NVP-PDF386 (VRC4887) compared favourably with those of control compounds, including glycopeptides, oxazolidinones, a streptogramin combination and other agents with activity focused against Gram-positive cocci.     

Antithrombotic and anticoagulant effects of the direct thrombin inhibitor dabigatran, and its oral prodrug, dabigatran etexilate, in a rabbit model of venous thrombosis

BACKGROUND: Oral anticoagulant therapies targeted at thrombin are being developed to overcome limitations associated with current standard therapies. OBJECTIVES: This study was undertaken to assess and compare the antithrombotic and anticoagulant effects of the novel, selective and reversible, direct thrombin inhibitor (DTI), dabigatran, and its oral prodrug dabigatran etexilate, to that of unfractionated heparin (UFH), hirudin and melagatran using a rabbit model of venous thrombosis. METHODS: A rabbit model of venous thrombosis consisting of endothelial damage with blood flow reduction was used with minor modifications. RESULTS: All compounds demonstrated a dose-dependent reduction in thrombus formation following i.v. administration with complete or almost complete inhibition at the highest doses. Dabigatran (in the dose range 0.03-0.5 mg kg(-1)) had a 50% effective dose of 0.066 mg kg(-1). By comparison, UFH (5-50 U kg(-1)), hirudin (0.01-0.05 mg kg(-1)) and melagatran (0.01-0.3 mg kg(-1)) had a 50% effective dose of 9.8 U kg(-1), 0.016 mg kg(-1) and 0.058 mg kg(-1), respectively. Similarly, oral dabigatran etexilate (1-20 mg kg(-1)) inhibited thrombus formation in a dose-dependent manner. Maximum inhibition was achieved within 1 h of administration, suggesting a rapid onset of action. For both routes of administration, inhibition of thrombus formation directly correlated with prolongation of the activated partial thromboplastin time. CONCLUSIONS: These findings demonstrate the potent anticoagulant and antithrombotic activity of dabigatran as a selective thrombin inhibitor in a rabbit model of venous thrombosis. Notably, dose-dependent and long-lasting antithrombotic efficacy was observed after application of its oral form dabigatran etexilate, which is currently undergoing phase III clinical development.     

Effect of the thromboxane synthetase inhibitor UK-37,248 (dazoxiben) upon platelet aggregation, coronary artery thrombosis and vascular reactivity

The effects of the thromboxane synthetase inhibitor dazoxiben on thrombus formation, platelet aggregation and vascular reactivity have been studied in open-chest anesthetized dogs. Dazoxiben at 4 mg/kg prolonged the time required for occlusive thrombi to form at sites of electrical injury to the left circumflex coronary artery by 3-fold and also decreased venous thromboxane B2 concentrations by 45% within 30 min. The progressive decline in flow observed during thrombogenesis was interrupted by rapid and spontaneous reactive hyperemic responses, the frequency and number of which were diminished by dazoxiben. In vitro platelet aggregation responses to ADP and collagen determined during the course of these experiments also were inhibited to a variable extent. The effect of dazoxiben on coronary vascular responsiveness was examined using an in situ constant pressure perfused coronary vascular bed. Although basal left circumflex coronary blood flow was unaffected by the drug, vasodilation induced by arachidonic acid, but not prostacyclin, was potentiated. The data suggest that dazoxiben possesses in vivo antithrombotic activity due to modification of platelet reactivity and that it can enhance coronary vasodilator responses to exogenously administered arachidonic acid.     

Active site-blocked factor IXa prevents intravascular thrombus formation in the coronary vasculature without inhibiting extravascular coagulation in a canine thrombosis model.

To assess the contribution of Factor IX/IXa, to intravascular thrombosis, a canine coronary thrombosis model was studied. Thrombus formation was initiated by applying current to a needle in the circumflex coronary artery. When 50% occlusion of the vessel developed, the current was stopped and animals received an intravenous bolus of either saline, bovine glutamyl-glycyl-arginyl-Factor IXa (IXai), a competitive inhibitor of Factor IXa assembly into the intrinsic Factor X activation complex, bovine Factor IX, or heparin. Animals receiving saline or Factor IX developed coronary occlusion due to a fibrin/platelet thrombus in 70 +/- 11 min. In contrast, infusion of IXai prevented thrombus formation completely (greater than 180 min) at doses of 460 and 300 micrograms/kg, and partially blocked thrombus formation at 150 micrograms/kg. IXai attenuated the accumulation of 125I-fibrinogen/fibrin at the site of the thrombus by approximately 67% (P less than 0.001) and resulted in approximately 26% decrease in serotonin release from platelets in coronary sinus (P less than 0.05). Hemostatic variables in animals receiving IXai, remained within normal limits. Animals given heparin in a concentration sufficient to prevent occlusive thrombosis had markedly increased bleeding, whereas heparin levels that maintained extravascular hemostasis did not prevent intracoronary thrombosis. This suggests that Factor IX/IXa can contribute to thrombus formation, and that inhibition of IXa participation in the clotting mechanism blocks intravascular thrombosis without impairing extravascular hemostasis.     

Comparative antithrombotic effects of heparin, recombinant hirudin and argatroban in a hamster femoral vein platelet-rich mural thrombosis model.

The antithrombotic properties of bolus i.v. injections of heparin, of recombinant hirudin (r-hirudin) or of the synthetic competitive thrombin inhibitor Argatroban were investigated in a quantitative hamster femoral vein platelet-rich mural thrombosis model. Heparin at a dose of 100 U/kg prolonged the activated partial thromboplastin time from 26 +/- 15 to 177 +/- 45 sec (P = .001), but did not significantly inhibit platelet-rich thrombus formation (7 +/- 44% inhibition, P = NS vs. placebo). However, 400 U/kg of heparin produced total inhibition of thrombus formation (101 +/- 14+, P less than .06 vs. control). R-hirudin and argatroban inhibited thrombus formation in a dose-dependent manner: 50% inhibition was obtained with 1.4 mg/kg for r-hirudin and with 2.0 mg/kg for Argatroban. A linear correlation was observed between the percentage of inhibition of thrombus formation vs. Activated partial thromboplastin time (r = 0.57, P = .003 for r-hirudin and r = 0.66, P = .002 for Argatroban). These results suggest that thrombin plays a pivotal role in platelet-rich mural thrombus formation, that this small animal model may be useful for investigation of the pharmacodynamics of synthetic thrombin inhibitors and that platelet-rich thrombus formation is inhibited effectively by heparin, r-hirudin and Argatroban. However, r-hirudin and Argatroban cause less profound changes in the coagulant function at doses that inhibit platelet-rich thrombus formation than heparin.     

Triple antithrombotic therapy in patients with atrial fibrillation undergoing coronary artery stenting: hovering among bleeding risk, thromboembolic events, and stent thrombosis

Dual antiplatelet treatment with aspirin and clopidogrel is the antithrombotic treatment recommended after an acute coronary syndrome and/or coronary artery stenting. The evidence for optimal antiplatelet therapy for patients, in whom long-term treatment oral anticoagulation is mandatory, is however scarce. To evaluate the safety and efficacy of the various antithrombotic strategies adopted in this population, we reviewed the available evidence on the management of patients receiving oral anticoagulation, such as a vitamin-k-antagonists, referred for coronary artery stenting. Atrial fibrillation is the most frequent indication for oral anticoagulation. The need of starting antiplatelet therapy in this clinical scenario raises concerns about the combination to choose: triple therapy with warfarin, aspirin, and a thienopyridine being the most frequent and advised. The safety of this regimen appeared suboptimal because of an increased risk in hemorrhagic complications. On the other hand, the combination of oral anticoagulation and an antiplatelet agent is suboptimal in preventing thromboembolic events and stent thrombosis; dual antiplatelet therapy may be considered only when a high hemorrhagic risk and low thromboembolic risk are perceived. Indeed, the need for prolonged multiple-drug antithrombotic therapy increases the bleeding risks when drug eluting stents are used. Since current evidence derives mainly from small, single-center and retrospective studies, large-scale prospective multicenter studies are urgently needed.     

Anti-inflammatory and antithrombotic effects of statins in the management of coronary artery disease

Inhibitors of 3-hydroxy-3-methylglutaryl coenzyme A reductase (statins), a potent class of cholesterol-lowering drugs, exert a number of pleiotropic effects, including anti-inflammatory and antithrombotic properties. Evidence is now accumulating that these effects are not related to the reduction in lipid levels. In vitro studies, supported recently by in vivo data, indicate that treatment with statins results in a significant decrease in the levels of inflammation markers, such as C-reactive protein, interleukin 6, and tumor necrosis factor alpha, which appear to be predictors of acute coronary events and help stratify cardiovascular risk. Up to now, only high-sensitive C-reactive protein testing has the potential to become an adjunctive method to assess the risk of coronary events in low- and high-risk individuals. Statins can also inhibit tissue factor expression, leading to impaired activation of the blood coagulation cascade, as evidenced by a decrease in thrombin generation in vivo. Interrelated inhibition of inflammation and thrombosis induced by statins is believed to largely contribute to clinical benefits from statin therapy, regardless of cholesterol levels. Further studies will answer the question whether markers of inflammation, other than C-reactive protein and possibly indices of thrombin formation, might improve cardiovascular risk stratification.     

Evaluation of PAI-039 [{1-benzyl-5-[4-(trifluoromethoxy)phenyl]-1H-indol-3-yl}(oxo)acetic acid], a novel plasminogen activator inhibitor-1 inhibitor, in a canine model of coronary artery thrombosis

We tested a novel, orally active inhibitor of plasminogen activator inhibitor-1 (PAI-1) in a canine model of electrolytic injury. Dogs received by oral gavage either vehicle (control) or the PAI-1 inhibitor PAI-039 [{1-benzyl-5-[4-(trifluoromethoxy)phenyl]-1H-indol-3-yl}(oxo)acetic acid] (1, 3, and 10 mg/kg) and were subjected to electrolytic injury of the coronary artery. PAI-039 caused prolongation in time to coronary occlusion (control, 31.7 +/- 6.3 min; 3 mg/kg PAI-039, 66.0 +/- 6.4 min; 10 mg/kg, 56.7 +/- 7.4 min; n = 5-6; p < 0.05) and a reduced thrombus weight (control, 7.6 +/- 1.5 mg; 10 mg/kg PAI-039, 3.6 +/- 1.0 mg; p < 0.05). Although occlusive thrombosis was observed across all groups based upon the absence of measurable blood flow, a high incidence (>60%) of spontaneous reperfusion occurred only in those groups receiving PAI-039. Spontaneous reperfusion in the 10 mg/kg PAI-039 group accounted for total blood flow (area under the curve of coronary blood flow) of 99.6 +/- 11.7 ml after initial thrombotic occlusion (p < 0.05 compared with control). Plasma PAI-1 activity was reduced in all drug-treated groups (percentage of reduction in activity p < 0.05; 10 mg/kg PAI-039), whereas ADP-, 9,11-dideoxy-11alpha,9alpha-epoxymethanoprostaglandin F(2alpha) (U46619)-, and collagen-induced platelet aggregation, as well as template bleeding and prothrombin time, remained unaffected by PAI-039. Ex vivo clot lysis analysis revealed normal clot formation but accelerated clot lysis in PAI-039-treated groups. The pharmacokinetic profile of PAI-039 indicated an oral bioavailability of 43 +/- 15.3% and a plasma half-life of 6.2 +/- 1.3 h. In conclusion, PAI-039 is an orally active prothrombolytic drug that inhibits PAI-1 and accelerates fibrinolysis while maintaining normal coagulation in a model of coronary occlusion.     

Pharmacological interruption of acute thrombus formation with minimal hemorrhagic complications by a small molecule tissue factor/factor VIIa inhibitor: comparison to factor Xa and thrombin inhibition in a nonhuman primate thrombosis model

This study was designed to evaluate the antithrombotic efficacy and bleeding propensity of a selective, small-molecule inhibitor of tissue factor/factor VIIa (TF/VIIa) in comparison to small-molecule, selective inhibitors of factor Xa and thrombin in a nonhuman primate model of thrombosis. Acute, spontaneous thrombus formation was induced by electrolytic injury to the intimal surface of a femoral blood vessel, which results in thrombus propagation at the injured site. The TF/FVIIa inhibitor 3-amino-5-[1-[2-([4-[amino(imino)methyl]benzyl]amino)-2-oxoethyl]-3-chloro-5-(isopropylamino)-6-oxo-1,6-dihydropyrazin-2-yl]benzoic acid dihydrochloride (PHA-927F) was fully effective in prevention of thrombosis-induced vessel occlusion at a dose of 400 microg/kg/min, i.v., in the arterial vasculature (femoral artery). Neither the effective dose nor multiples up to 4.4-fold the effective arterial plasma concentration elicited any significant effect on bleeding time or blood loss from either the bleeding time site or the surgical (femoral isolation) site. Small-molecule inhibitors of factor Xa or thrombin were effective arterial antithrombotic agents; however, in contrast to the TF/FVIIa inhibitor, they both elicited substantial increases in bleeding propensity at the effective dose and at multiples of the effective plasma concentration. These data indicate that TF/VIIa inhibition effectively prevented arterial thrombosis with less impact on bleeding parameters than equivalent doses of factor Xa and thrombin inhibitors.     

Human thrombomodulin is not an efficient inhibitor of the procoagulant activity of thrombin.

The effect of human thrombomodulin isolated from placenta on the procoagulant activity of thrombin was studied and compared to that of rabbit thrombomodulin. The isolated protein was proved to be thrombomodulin because a rabbit antibody against the isolated protein blocked protein C activation by thrombomodulin in solution and also blocked the protein-C-activating cofactor activity of human umbilical vein endothelial cells. The affinity of human thrombomodulin for human thrombin in the presence of fibrinogen is 30 times less than that of rabbit thrombomodulin. This value is based on the measurements of the clotting time of human fibrinogen and thrombin in the presence of increasing amounts of thrombomodulin. Human thrombomodulin was also much less effective compared with rabbit thrombomodulin in inhibiting thrombin-induced human coagulation factor V activation. The ability to inhibit release of [3H]serotonin from washed human platelets was at least 10 times less using human thrombomodulin compared with rabbit thrombomodulin. A partially purified preparation of human lung thrombomodulin was also relatively ineffective in inhibiting thrombin-induced serotonin release from platelets, indicating that the difference between rabbit and human thrombomodulin is one of species rather than of tissue. Thus, while human thrombomodulin is a potent cofactor in protein C activation, it is not an efficient inhibitor of the procoagulant actions of thrombin.     

N端脑钠肽前体、脂蛋白(a)与冠心病患者冠状动脉病变的相关性

目的探讨冠心病患者冠状动脉病变和血清N端脑钠肽前体(NT-pro BNP)、脂蛋白(a)[LP(a)]的关系。方法选取经冠脉造影检查的患者108例,其中非冠心病28例作为对照组,冠心病80例作为研究组。研究组根据冠脉病变支数又分为单支组(n=22)、双支组(n=27)、三支组(n=31)。比较各组患者血清NT-pro BNP与LP(a)水平,分析血清NT-pro BNP,LP(a)与冠脉病变的关系。结果研究组患者血清NT-pro BNP与LP(a)均显著高于对照组(P0.05);研究组患者随着病变支数增多,血清NT-pro BNP与LP(a)也逐渐升高,三支组显著高于单支组(P0.05);血清NT-pro BNP,LP(a)与Gensini评分有显著正相关性(P0.05)。结论冠心病患者冠脉病变与血清NT-pro BNP,LP(a)有相关性。     

The role of thrombin activatable fibrinolysis inhibitor in arterial thrombosis at a young age: the ATTAC study1

Summary. Background and objectives: Thrombin activatable fibrinolysis inhibitor (TAFI) attenuates fibrinolysis and may therefore contribute to the pathophysiology of arterial thrombosis. The aim of the present study was to elucidate the pathogenetic role of TAFI levels and genotypes in young patients with arterial thrombosis. Patients and methods: In a case–control study, 327 young patients with a recent first‐ever event of coronary heart disease (CHD subgroup) or cerebrovascular disease (ischemic stroke subgroup) and 332 healthy young controls were included. TAFI levels [intact TAFI, activation peptide (TAFI‐AP) and (in)activated TAFI (TAFIa(i)] and TAFI activity were measured and genetic variations in the TAFI gene (?438G/A, 505G/A and 1040C/T) were determined. Results: In the total group of patients, TAFIa(i) levels were higher (145.1 ± 37.5%) than in controls (137.5 ± 31.3%, P = 0.02). Plasma levels of intact TAFI, TAFI‐AP and TAFI activity were similar in patients and controls. In the CHD subgroup (n = 218), intact TAFI levels were higher (109.4 ± 23.0%) than in controls (102.8 ± 20.7%, P = 0.02). In 325Ile/Ile homozygotes, lower TAFI levels and a decreased risk of arterial thrombosis were observed (OR 0.58, 95% CI 0.34–0.99) compared with patients with the common 325Thr/Thr genotype. This association was most evident in CHD patients (OR 0.48, 95% CI 0.26–0.90). Haplotype analyses supported a role for the Thr325Ile polymorphism. Conclusions: TAFIa(i) levels were higher in patients with cardiovascular disease. Furthermore, the TAFI 325Thr/Ile polymorphism was associated with lower TAFI levels and with the risk of cardiovascular disease in young patients, especially in CHD.