pyrimidinones and Thrombosis

Based on in vitro and animal data, PI3Kβ is given an important role in platelet adhesion and aggregation but its role in insulin signaling is unclear.. To strengthen the PI3Kβ target validation using the novel, short-acting inhibitor AZD6482.. AZD6482 is a potent, selective and ATP competitive PI3Kβ inhibitor (IC(50) 0.01 μm). A maximal anti-platelet effect was achieved at 1 μm in the in vitro and ex vivo tests both in dog and in man. In dog, in vivo AZD6482 produced a complete anti-thrombotic effect without an increased bleeding time or blood loss. AZD6482 was well tolerated in healthy volunteers during a 3-h infusion. The ex vivo anti-platelet effect and minimal bleeding time prolongation in the dog model translated well to data obtained in healthy volunteers. AZD6482 inhibited insulin-induced human adipocyte glucose uptake in vitro (IC(50) of 4.4 μm). In the euglycemic hyperinsulinemic clamp model, in rats, glucose infusion rate was not affected at 2.3 μm but reduced by about 60% at a plasma exposure of 27 μm. In man, the homeostasis model analysis (HOMA) index increased by about 10-20% at the highest plasma concentration of 5.3 μm.. This is the first human target validation for PI3Kβ inhibition as anti-platelet therapy showing a mild and generalized antiplatelet effect attenuating but not completely inhibiting multiple signaling pathways with an impressive separation towards primary hemostasis. AZD6482 at 'supratherapeutic' plasma concentrations may attenuate insulin signaling, most likely through PI3Kα inhibition.

Topics: Adipocytes; Adolescent; Adult; Animals; Bleeding Time; Blood Platelets; Cell Line, Tumor; Class Ia Phosphatidylinositol 3-Kinase; Disease Models, Animal; Dogs; Dose-Response Relationship, Drug; Double-Blind Method; Fibrinolytic Agents; Glucose; Hemostasis; Hemostatics; Humans; Insulin Resistance; Male; ortho-Aminobenzoates; Phosphoinositide-3 Kinase Inhibitors; Phosphorylation; Platelet Aggregation; Platelet Aggregation Inhibitors; Platelet Function Tests; Protein Kinase Inhibitors; Proto-Oncogene Proteins c-akt; Pyrimidinones; Rats; Signal Transduction; Thrombosis; Time Factors; Young Adult

Arterial thrombosis is triggered by tissue factor, which is a transmembrane glycoprotein can be released into the blood circulation after plaque rupture. Animal models with reflecting ruptured plaque lesions will be useful to understand efficacy of anticoagulant. In this study, we sought to improve a common arteriovenous shunt model in rabbits, aiming for a model of thrombosis stimulated with tissue factor, and to investigate the anti-thrombotic effect of a direct factor Xa inhibitor TAK-442 in the model.. In the model where thrombus was stimulated with a thrombogenic silk thread soaked with recombinant human tissue factor, thrombus formation was significantly reduced by TAK-442 at more than 37.5 µg/kg, accompanied with prolonged plasma hemostatic parameters. Although efficacious doses of anti-coagulants in ordinary arteriovenous thrombosis models are widely reported to be higher than those in venous thrombosis models, TAK-442 showed its efficacy in the present arteriovenous shunt thrombosis model, with equivalent sensitivity in a previously reported venous model. TAK-442 might be effective under conditions thrombus formed is more influenced by tissue factor pathway.

Topics: Animals; Arteriovenous Shunt, Surgical; Disease Models, Animal; Factor Xa Inhibitors; Fibrinolytic Agents; Male; Pyrimidinones; Rabbits; Sulfones; Thrombosis

Class IA phosphoinositide 3-kinase β (PI3Kβ) is considered a potential drug target in arterial thrombosis, which is a major cause of death worldwide. Here we show that a striking phenotype of mice with selective p110β deletion in the megakaryocyte lineage is thrombus instability at a high shear rate, which is an effect that is not detected in the absence of p110α in platelets. The high shear rate-dependent thrombus instability in the absence of p110β is observed both ex vivo and in vivo with the formation of platelet emboli. Moreover, PI3Kβ is required for the recruitment of new platelets to a growing thrombus when a pathological high shear is applied. Treatment of human blood with AZD6482, a selective PI3Kβ inhibitor, phenocopies p110β deletion in mouse platelets, which highlights the role of the kinase activity of p110β. Within the growing platelet thrombus, p110β inactivation impairs the activating phosphorylations of Akt and the inhibitory phosphorylation of GSK3. In accord with these data, pharmacologic inhibition of GSK3 restores thrombus stability. Thus, platelet PI3Kβ is not essential for thrombus growth and stability at normal arterial shear but has a specific and critical role in maintaining the integrity of the formed thrombus on elevation of shear rate, suggesting a potential risk of embolization on treatment with PI3Kβ inhibitors.

Topics: Animals; Blood Platelets; Class I Phosphatidylinositol 3-Kinases; Gene Expression Regulation; Glycogen Synthase Kinase 3; Humans; Mechanotransduction, Cellular; Mice; Mice, Transgenic; ortho-Aminobenzoates; Phosphorylation; Platelet Aggregation; Proto-Oncogene Proteins c-akt; Pyridines; Pyrimidines; Pyrimidinones; Stress, Mechanical; Thrombosis

Treatment of myocardial infarction within the first 1 to 2 hours with a thrombolytic agent, percutaneous coronary intervention, or an αIIbβ3 antagonist decreases mortality and the later development of heart failure. We previously reported on a novel small molecule αIIbβ3 antagonist, RUC-2, that has a unique mechanism of action. We have now developed a more potent and more soluble congener of RUC-2, RUC-4, designed to be easily administered intramuscularly by autoinjector to facilitate its use in the prehospital setting. Here, we report the properties of RUC-4 and the antiplatelet and antithrombotic effects of RUC-2 and RUC-4 in animal models.. RUC-4 was ≈ 20% more potent than RUC-2 in inhibiting human ADP-induced platelet aggregation and much more soluble in aqueous solutions (60-80 mg/mL). It shared RUC-2's specificity for αIIbβ3 versus αVβ3, did not prime the receptor to bind fibrinogen, or induce changes in β3 identified by a conformation-specific monoclonal antibody. Both RUC-2 and RUC-4 prevented FeCl3-induced thrombotic occlusion of the carotid artery in mice and decreased microvascular thrombi in response to laser injury produced by human platelets infused into transgenic mice containing a mutated von Willebrand factor that reacts with human but not mouse platelets. Intramuscular injection of RUC-4 in nonhuman primates at 1.9 and 3.85 mg/kg led to complete inhibition of platelet aggregation within 15 minutes, with dose-dependent return of platelet aggregation after 4.5 to 24 hours.. RUC-4 has favorable biochemical, pharmacokinetic, pharmacodynamic, antithrombotic, and solubility properties as a prehospital therapy of myocardial infarction, but the possibility of increased bleeding with therapeutic doses remains to be evaluated.

Topics: Animals; Binding Sites; Blood Platelets; Carotid Stenosis; Chlorides; Disease Models, Animal; Emergency Medical Services; Ferric Compounds; Fibrinolytic Agents; Humans; Macaca fascicularis; Male; Mice; Mice, Transgenic; Molecular Dynamics Simulation; Myocardial Infarction; Platelet Aggregation; Platelet Aggregation Inhibitors; Platelet Glycoprotein GPIIb-IIIa Complex; Protein Binding; Protein Conformation; Pyrimidinones; Solubility; Thiadiazoles; Thrombosis; von Willebrand Factor

Topics: Animals; Blood Platelets; Carotid Stenosis; Emergency Medical Services; Fibrinolytic Agents; Humans; Male; Myocardial Infarction; Platelet Aggregation; Platelet Aggregation Inhibitors; Platelet Glycoprotein GPIIb-IIIa Complex; Pyrimidinones; Thiadiazoles; Thrombosis

Revaprazan is the first acid pump antagonist with a function similar to that of proton pump inhibitors (PPIs). It has a dual action, active suppression of gastric acid secretion and gastric mucosa protection. While PPIs are known to enhance the prolongation of prothrombin time by warfarin, no research has been done on the drug interaction between revaprazan and warfarin. This study was conducted in order to verify the potential drug interaction between revaprazan and warfarin. Omeprazole, a representative PPI, was used as the control for revaprazan. We searched for patients who were given either revaprazan or omeprazole along with warfarin using the medical record database of Seoul National University Hospital between July 2007 and June 2010. Among the 15 patients who took revaprazan and warfarin together, 73.3% (11/15) showed more than 30% reduction of anticoagulation effect by warfarin after revaprazan was added. The revaprazan group showed a significant shortening of prothrombin time during revaprazan administration compared to pre- and post-revaprazan medication (P < 0.05) while the omeprazole group did not show such difference. Revaprazan seems to have cumulative dose-dependent anti-warfarin or anti-coagulation effect, as judged from the fact that the longer medication with revaprazan showed correlation with the shortening of prothrombin time (R = -0.632, P < 0.05). This study shows a possible interaction between revaprazan and warfarin and suggests that revaprazan can cause shortening of prothrombin time. Therefore, when revaprazan is prescribed to patients on warfarin therapy, prothrombin time should be frequently monitored.

Topics: Aged; Aged, 80 and over; Anti-Ulcer Agents; Anticoagulants; Drug Interactions; Female; Humans; Male; Middle Aged; Omeprazole; Prothrombin Time; Proton Pump Inhibitors; Pyrimidinones; Tetrahydroisoquinolines; Thrombosis; Warfarin

To evaluate the potential of a novel dihydropyrimidinone, ethyl 4-(4'-heptanoyloxyphenyl)-6-methyl-3,4-dihydropyrimidin-2-one-5-carboxylate (H-DHPM), as a calcium channel blocker, endowed with the ability to inhibit platelet aggregation effectively.. In-vitro and in-vivo studies were conducted for the determination of antiplatelet activity using adenosine diphosphate (ADP), collagen or thrombin as inducers. Calcium channel blocking activity and nitric oxide synthase (NOS) activity were monitored. Lipopolysaccharide (LPS)-mediated prothrombotic conditions were developed in rats to study the efficacy of H-DHPM to suitably modulate the inflammatory mediators such as inducible NOS (iNOS) and tissue factor. The cGMP level and endothelial NOS (eNOS) expression were checked in aortic homogenate of LPS-challenged rats pretreated with H-DHPM. The effect of H-DHPM on FeCl(3) -induced thrombus formation in rats was examined.. The concentrations of H-DHPM required to give 50% inhibition (IC50) of in-vitro platelet aggregation induced by ADP, collagen or thrombin were 98.2±2.1, 74.5±2.3 and 180.7±3.4µm, respectively. H-DHPM at a dose of 52.0±0.02mg/kg (133µmol/kg) was found to optimally inhibit ADP-induced platelet aggregation in-vivo. The level of nitric oxide was found to be up to 9±0.08-fold in H-DHPM-treated platelets in-vitro and 8.2±0.05-fold in H-DHPM-pretreated rat platelets in-vivo compared with control. OH-DHPM, the parent compound was found to be ineffective both in-vitro and in-vivo. H-DHPM-pretreated rats were able to resist significantly the prothrombotic changes caused by LPS by blunting the expression of iNOS, tissue factor and diminishing the increased level of cGMP to normal. H-DHPM enhanced the eNOS expression in aorta of rats treated with LPS. H-DHPM displayed synergy with antiplatelet activity of aspirin even at lower doses. H-DHPM was found to inhibit the LPS-induced platelet aggregation in younger as well as older rats. H-DHPM exhibited the ability to markedly decrease FeCl(3) -induced thrombus formation in rats.. H-DHPM has the attributes of a promising potent antiplatelet candidate molecule that should attract further study. H-DHPM displayed antiplatelet activity both in vivo and in vitro, which was due partially by lowering the intraplatelet calcium concentration.

Topics: Adenosine Diphosphate; Animals; Aorta; Aspirin; Blood Platelets; Collagen; Cyclic GMP; Drug Synergism; Inflammation Mediators; Inhibitory Concentration 50; Lipopolysaccharides; Male; Nitric Oxide; Nitric Oxide Synthase Type II; Platelet Aggregation Inhibitors; Pyrimidinones; Rats; Rats, Sprague-Dawley; Thrombin; Thromboplastin; Thrombosis

Activated platelets facilitate blood coagulation by providing factor V and a procoagulant surface for prothrombinase. Here, we investigated the potential synergy of a potent factor Xa/prothrombinase inhibitor, TAK-442, plus aspirin or clopidogrel in preventing arterial thrombosis and whole blood coagulation.. Thrombus formation was initiated by FeCl(3)-induced rat carotid injury. Bleeding time was evaluated with the rat tail transection model. Whole blood coagulation was assessed by thromboelastographic examination (TEG) for which blood obtained from control, aspirin-, or clopidogrel-treated rats was transferred to a TEG analyzer containing, collagen or adenosine diphosphate (ADP), and TAK-442 or vehicle.. TAK-442 (3mg/kg, po), aspirin (100mg/kg, po) or clopidogrel (3mg/kg, po) alone had no significant effect on thrombus formation, whereas the combination of TAK-442 with aspirin and clopidogrel remarkably prolonged the time to thrombus formation without additional significant prolongation of bleeding time. TEG demonstrated that the onset of collagen-induced blood coagulation were slightly longer in aspirin-treated rats than control; however, when the blood from aspirin-treated rats was subsequently treated in vitro with 100 nM TAK-442, the onset of clotting was significantly prolonged. In contrast, only marginal prolongation was observed with TAK-442 treatment of blood from control animals. The onset time of ADP-induced blood coagulation was slightly longer in clopidogrel-treated rats compared with control, and it was further extended by TAK-442 treatment.. These results demonstrate that blood coagulation can be markedly delayed by the addition of TAK-442 to antiplatelets treatment which could contribute to synergistic antithrombotic efficacy in these settings.

Topics: Animals; Aspirin; Blood Coagulation; Blood Coagulation Tests; Clopidogrel; Drug Synergism; Factor Xa Inhibitors; Male; Platelet Aggregation; Platelet Aggregation Inhibitors; Pyrimidinones; Rats; Rats, Sprague-Dawley; Sulfones; Thrombosis; Ticlopidine

Previously, potent factor Xa inhibitors were described based on a pyrazole core. Modifications of the pyrazole core have provided additional novel, highly potent factor Xa inhibitors. This manuscript will describe the synthesis and biological activity of factor Xa inhibitors containing the 1H-pyrazolo[4,3-d]pyrimidin-7(6H)-one and related bicyclic cores. Many of these compounds are potent, selective, and orally bioavailable inhibitors of coagulation factor Xa.

Topics: Administration, Oral; Animals; Antithrombin III; Biological Availability; Crystallography, X-Ray; Dogs; Factor Xa Inhibitors; Fibrinolytic Agents; Pyrazoles; Pyrimidinones; Thrombosis

Topics: Humans; Morpholines; Platelet Aggregation Inhibitors; Pyrimidinones; Thrombosis

Topics: Adult; Cardiomyopathy, Dilated; Crystallization; Echocardiography, Transesophageal; Female; Heart Diseases; Humans; Infusions, Intravenous; Potassium Channel Blockers; Pyrimidinones; Thrombosis