Nonpeptide angiotensin II receptor antagonists: synthesis and biological activity of 1H-imidazo and 1H-[1,2,3]-triazolo fused derivatives.

A series of 1H-Imidazo and 1H-[1,2,3]-Triazolo fused derivatives have been obtained and tested as non peptide AII receptor antagonists. Modification of the classical biphenyl moiety to a 4-arylthienyl fragment afforded interesting activities.

Imidazo[1,5-a]pyrimidine and benzo[4,5]imidazo-[1,2-a]pyrimidine derivatives as calcium antagonists.

Several bicyclic dihydropyrimidines were synthesized and evaluated for their calcium antagonistic activities by comparison with the usual 1,4-dihydropyridine calcium antagonist reference compound nifedipine. The solid-state structure of the isopropyl 2-methyl-4-(3'-nitrophenyl)-1,4-dihydrobenzo[4,5]imidazo[1,2- a]pyrimido-3-carboxylate shows that these compounds can adopt the most important structural features of the 1,4-dihydropyridine and 1,4-dihydropyrimidine calcium channel blockers. The high-potassium depolarized rat aorta assay was used for testing the compounds as calcium channel blockers. Some compounds showed interesting vasorelaxant activity.

Compound PCA-4248 interferes with bronchopulmonary anaphylaxis and with in vitro hyperresponsiveness to platelet-activating factor.

The intravenous (i.v.) or oral administration of the platelet-activating factor (PAF) antagonist, PCA-4248, to guinea-pigs blocked selectively the bronchoconstriction induced by PAF, as well as the accompanying thrombocytopenia and leucopenia. In addition, PCA-4248 i.v. or intratracheal (i.t.) administration blocked the bronchoconstriction caused by the i.t. instillation of PAF. As in the case of other PAF antagonists, bronchoconstriction caused by the i.t. instillation of antigen was only inhibited by PCA-4248 in guinea-pigs that did not receive a booster injection of antigen during sensitization whereas the booster injection of antigen made anaphylactic bronchoconstriction resistant to the compound. In vitro, when lungs from non-sensitized guinea-pigs were perfused with Krebs-bovine serum albumin (BSA) solution supplemented with PCA-4248, bronchoconstriction and the formation of thromboxane A2 by PAF were blocked. In this in vitro model of perfused lungs, active sensitization with a booster injection of antigen leads to bronchopulmonary hyperresponsiveness to PAF and failure of other PAF antagonists to inhibit the effects of PAF itself. Surprisingly, in lungs isolated from actively sensitized and boosted guinea-pigs, PCA-4248 blocked the effects of PAF, indicating that this compound possesses additional original properties in this model.

Interaction of platelets with subendothelium in rats treated with PCA-4230, a new antithrombotic agent. Effect of PCA-4230 on experimental thrombosis.

The effects of a new antithrombotic compound, PCA-4230, versus ticlopidine were investigated using an experimental thrombosis and vascular endothelial injury model in rats. Both PCA-4230 and ticlopidine protected rat arteries from the formation of prominent thrombi and most of microthrombi without modifying the formation of a first platelet monolayer. Neither coagulation parameters nor fibrinolysis were modified by these antithrombotic drugs. Neither PCA-4230 nor ticlopidine affected thromboxane A2 production in rats, whereas unlike PCA-4230, ticlopidine inhibited ex vivo fibrinogen binding to the fibrinogen receptor found on the platelet membrane. In conclusion, PCA-4230 and ticlopidine inhibited thrombus formation in vivo by a platelet-dependent mechanism which may be different for one or the other drug in spite of the fact that the protective effect measured in this thrombosis model is quite similar for either PCA-4230 or ticlopidine. The above-mentioned results clearly show that PCA-4230 is a new potent agent with both antivascular-damaging and antiplatelet activities, and devoid of effects on coagulation and fibrinolytic systems.

4-Alkyl-1,4-dihydropyridines derivatives as specific PAF-acether antagonists.

A new series of 4-alkyl-1,4-dihydropyridines (1,4-DHP) were synthesized and evaluated for their ability to inhibit washed rabbit platelet aggregation induced by PAF-acether (1-O-hexadecyl/octadecyl-2-O-acetyl-sn-glycero-3-phosphorylcholine) and to reverse PAF-induced hypotension in anesthetized rats. Additionally, compounds were evaluated for their ability to inhibit the binding of radiolabeled PAF to its receptor on rabbit platelets. Among these compounds, 6I and 6L were the most potent and specific antagonists. At concentrations up to 100 microM, neither compound 6I nor compound 6L caused platelet aggregation nor did they inhibit platelet aggregation induced by collagen or adenosine diphosphate. Compound 6L did not show in vitro calcium channel blocker activity measured on vascular smooth muscle preparations of rabbit aorta and on [3H]nitrendipine binding assays. The compound did not show any cardiovascular effects in anesthetized rat at iv doses up to 1000 micrograms/kg, and the Ki value was 568.62 nmol. These results indicate that compound 6L is a potent and specific PAF antagonist with 1,4-dihydropyridine structure but devoid of a significant cardiovascular activity related to calcium-antagonist properties.

1,4-Dihydropyridines, a new class of platelet-activating factor receptor antagonists: in vitro pharmacologic studies.

PCA 4233 [2-(phenylthio)ethyl-5-ethoxycarbonyl-2,4,6-trimethyl- 1,4-dihydropyridine-3-carboxylate] and PCA 4248 [2-(phenylthio) ethyl-5-methoxycarbonyl-2, 4, 6-trimethyl-1, 4-dihydropyridine-3-carboxylate], two compounds developed from a series of 1,4-dihydropyridines that lack pharmacologic effects on voltage-operated calcium channels, were found to block selectively rabbit operated calcium channels, were found to block selectively rabbit and human platelet aggregation and secretion, and binding of [3H]-labeled platelet-activating factor (PAF) to human platelet and polymorphonuclear PAF receptors. Rabbit platelet aggregation was tested with 1.9 nM PAF, i.e., a concentration producing maximal response, and was completely blocked with 10 microM PCA 4233 and 3 microM 4248 (IC50 values, 2.55 and 1.05 microM, respectively). Human platelet aggregation in platelet-rich plasma was studied with 1 microM PAF, a concentration that caused a response comparable with that of 1.9 nM PAF in rabbit platelets. The IC50 of PCA 4248 for ATP release under these conditions was 3.6 microM. PCA 4248 behaved as a competitive and selective antagonist in [3H]serotonin secretion studies on rabbit platelets, since it displaced rightwards log dose-response curves and lacked any effect on thrombin- and ionophore A23187-induced platelet secretion. A pA2 value of 7.5 was obtained from Schild plots on [3H]serotonin secretion studies. PCA 4248 also produced a dose-dependent inhibition of [3H]PAF binding to human platelets and to human polymorphonuclear leukocytes. These data indicate that PCA 4233 and PCA 4248 belong to a new class of selective PAF-receptor antagonists.

Pharmacological actions of PCA 4248, a new platelet-activating factor receptor antagonist: in vivo studies.

The ability of PCA 4248 [2-(phenylthio)ethyl-5-methoxycarbonyl- 2,4,6-trimethyl-1,4-dihydropyridine-3-carboxylate] to block PAF-induced systemic hypotension and protein-rich plasma extravasation in rats, and PAF-induced death in mice, was tested. These studies were complemented with experiments using soluble aggregates of immunoglobulin G (A-IgG), bacterial endotoxin and the cytokine tumor necrosis factor as putative inducers of the generation of endogenous PAF. Significant inhibition of PAF-induced systemic hypotension was observed with i.v. PCA 4248 at doses of 0.3 to 1 mg/kg (IC50 value, 0.45 mg/kg, with PAF 0.33 micrograms/kg). Reversal of the hypotension was rapidly observed when PCA 4248 was administered after PAF. The extravasation induced by 1 microgram/kg PAF was also blocked by PCA 4248 (IC50 value, 0.36 mg/kg). Inhibition of the extravasation induced by A-IgG and endotoxin was also provided by PCA 4248 at the dose of 1 mg/kg, and lasted for at least 1 hr in the experiments carried out with endotoxin, which caused extravasation with a temporal pattern more protracted than that of PAF and A-IgG. Intradermal extravasation induced by PAF reached a maximum at 30 min after injection, and was also inhibited by PCA 4248. In contrast, PCA 4248 caused a less remarkable, but statistically significant reduction of the intradermal extravasation caused by tumor necrosis factor. Pretreatment of mice with an oral dose of 30 mg/kg PCA 4248, 5 min before challenge with PAF (LD84 = 80 micrograms/kg PAF, i.v.) increased the survival rate from 16% to 68%. These data indicate that compounds containing a 1,4-dihydropyridine structure can antagonize PAF effects on experimental animals.(ABSTRACT TRUNCATED AT 250 WORDS)

Synthesis, platelet aggregation inhibitory activity, and in vivo antithrombotic activity of new 1,4-dihydropyridines.

A series of 1,4-dihydropyridines (DHP) bound to 1,2-benzisothiazol-3-ones were synthesized and evaluated for their ability to inhibit platelet aggregation induced by collagen in human platelet-rich plasma (PRP) and to protect mice against experimental thrombosis. The results showed that the compounds were in vitro inhibitors of collagen-induced platelet aggregation. Most of them were also effective in reducing mortality in the mouse antithrombotic assay. 2-(1,1,3-Trioxo-2,3-dihydro-1,2-benzisothiazol-2-yl)ethyl 2,6-dimethyl-5-(ethoxycarbonyl)-4-methyl-1,4-dihydropyridinecarboxyla te (4A) is the most promising compound. This compound did not show any cardiovascular effects either in the anesthetized cat or in the anesthetized rat at iv doses up to 750 or 500 micrograms/kg, respectively. Likewise, antiplatelet and cardiovascular effects of compound 4A were simultaneously studied in anesthetized rats and compared with those of nitrendipine.

The antithrombogenic in vivo effects of calcium channel blockers in experimental thrombosis in mice.

A new model of thrombotic challenge, well suited for screening agents and which acts primarily against platelet thromboembolism, has been used to test the in vivo anti-platelet effects of four calcium channel blockers (CCB). An i.v. injection of a mixture of collagen plus epinephrine (15 micrograms and 1.8 micrograms/mouse, respectively) was given to male mice. 94% control mice died or remained paralyzed for more than 15 minutes. The dihydropyridine agents, CRE-223 and Nifedipine, were highly protective against experimental thrombosis, whereas Verapamil had a weaker and much shorter effect and, on the other hand, Diltiazem had no protective effect over a range of doses. The activity on both dihydropyridines lasted for seven hours or even longer.

Effect of etersalate on human platelet responsiveness. A study in healthy volunteers.

Ex vivo antiplatelet properties of 2-(p-acetamido-phenoxy)ethyl-o-acetoxybenzoate (etersalate, Daital) and its effects on serum thromboxane A2 (TXA2) levels and prostaglandin I2 (PGI2) generation were studied in human volunteers at two levels of oral dosing. Etersalate inhibited at the lower dosage platelet function and decreased TXA2 levels, but PGI2 generation from rat aortic rings was stimulated when incubated with plasma from etersalate-treated donors. Blood coagulation parameters remained within normal values. It is suggested that etersalate administration could act on platelet arachidonate metabolism at a different level than that of the cyclooxygenase pathway.

The metabolism of the anti-inflammatory drug eterylate in rat, dog and man.

Oral doses of 14C-eterylate were well absorbed by rat and man and excreted mainly in the urine (94% dose by rat in three days and 91% by man in five days). Oral doses to dogs were excreted in similar proportions in both the urine and faeces, although faecal 14C was probably derived in part, from biliary-excreted material. Peak plasma 14C and drug concn. were generally reached between one and three hours after oral doses. In humans, only two metabolites, salicylic acid and 4-acetamido-phenoxyacetic acid, were detected in plasma. The latter was cleared more rapidly than the former and hence plasma salicyclate concn. reached a peak (10.9 and 19.8 micrograms/ml in Subjects 1 and 2, respectively) and initially declined with a half-life of about two-three hours. Plasma 4-acetamidophenoxyacetic acid concn. reached a peak (4.3, 10.0 micrograms/ml, respectively) and declined with a half-life of about one hour. Tissue concn. of 14C were generally greater in dogs than in rats. Highest conc. occurred at three hours in dogs and at one hour in rats. Apart from those in the liver and kidneys, tissue concn. were lower than those in the corresponding plasma. Unchanged drug was not detected in urine or plasma of any species and was rapidly metabolized in human plasma. The major 14C components in human urine were identified as salicyluric acid and 4-acetamidophenoxyacetic acid; minor metabolites were salicylic acid, gentisic acid and paracetamol. These metabolites were also detected in rat urine albeit in different proportions to those in human urine. Dog urine contained less of these metabolites and a major proportion of the 14C was associated with relatively non-polar components. Although salicylic acid and 4-acetamidophenoxyacetic acid were the only major circulating metabolites in man and rat, dog plasma also contained the non-polar urine metabolites.

Effect of clofibrate on the phospholipid biosynthesis in rat liver.

The effect of clofibrate on rat liver phospholipid biosynthesis was studied using 32P as a precursor. Phospholipid classes, levels and specific radioactivity were evaluated. Significant increases in levels of phosphatidylethanolamine and phosphatidylcholine were found and could account for the observed increase in total phospholipids. Specific activity of phosphatidylserine increased and that of phosphatidylethanolamine decreased. This fact suggests that clofibrate seems to alter the systems engaged in the transformation occurring within the different classes of phospholipids but not the de novo biosynthesis.