Phthalazine PDE4 inhibitors. Part 2: the synthesis and biological evaluation of 6-methoxy-1,4-disubstituted derivatives.

This communication describes the synthesis and in vitro evaluation of a novel and potent series of phosphodiesterase type IV (PDE4) inhibitors. The compounds described present substituents in position 4 of the phthalazine ring to replace the commonly observed cyclopentyloxy moiety of rolipram analogues. Preliminary evidences of reduced side effects compared to standards and improved pharmacokinetic properties for selected derivatives are also reported.

The synthesis and biological evaluation of a novel series of phthalazine PDE4 inhibitors I.

This communication describes the synthesis and in vitro evaluation of a novel and potent series of phosphodiesterase type IV (PDE4) inhibitors. The compounds described represent conformationally constrained analogues of RP 73401, Piclamilast. Preliminary evidences of reduced side effects of II compared to standards are also reported.

Pharmacological evidence for the presence of a peripheral postjunctional D2-like dopamine receptor in rabbit splenic artery.

1. This study was designed to investigate the involvement of postjunctional D2-like receptors in a rabbit vasculature model used to evaluate the D1-like agonist activity. Dopamine, epinine and (-)-DP-5,6-ADTN, three mixed D1/D2-like agonists, fenoldopam and SKF 82958, two selective D1-like agonists and SKF 89124, a selective D2-like agonist, were administered cumulatively in precontracted and alpha/beta-blocked rabbit splenic artery rings in order to evaluate their D1-like-mediated vasorelaxant activity before and after pretreatment with the selective D2-like antagonist YM 09151-2 (1 nM). 2. Dopamine (pD2=6.35+/-0.09), epinine (pD2=6.73+/-0.13), (-)-DP-5,6-ADTN (pD2=7.56+/-0.09) and SKF 82958 (pD2=8.55+/-0.10) reversed completely the U46619-induced contracture whereas SKF 89124 was inactive up to 10 microM and fenoldopam acted like a partial agonist (pD2=8.31+/-0.09, alpha=0.62). The selective D2-like dopamine receptor antagonist YM 09151-2 (1 nM) significantly (P<0.05) potentiated the vasorelaxant activity of dopamine (pD2=7.01+/-0.07), epinine (pD2=7.14+/-0.08), (-)-DP-5,6-ADTN (pD2=8.19+/-0.09) and SKF 89124 (40% relaxation at 10 microM), whereas it did not alter the effects of fenoldopam (pD2=8.40+/-0.09, alpha=0.68) and SKF 82958 (pD2=8.58+/-0.08). 3. The D2-like antagonist YM 09151-2 induced the same degree of effect with all the substances tested in both endothelium-denuded and endothelium-intact preparations. 4. The selective D2-like dopamine receptor agonist SKF 89124 did not produce any intrinsic effect on the splenic artery, but was able to produce a rightward shift of the forskolin-induced relaxation. 5. The results of these experiments support the existence of a non-endothelial postjunctional D2-like dopamine receptor counteracting the D1-like-mediated vasodilatation in rabbit splenic artery, probably by the inhibition of adenylate cyclase.

The effect duration of selective phosphodiesterase inhibitors in the guinea pig.

The beta-adrenoceptor agonists, isoprenaline, salbutamol and salmeterol, the non-selective phosphodiesterase (PDE) isoenzyme inhibitors, theophylline, trequinsin; the PDE3 isoenzyme inhibitor, milrinone; the PDE3/4 isoenzyme inhibitor, benzafentrine; and the PDE4 isoenzyme inhibitors, denbufylline, nitraquazone, RP 73401, Ro-20-1724, rolipram and tibenelast all induced concentration-dependent reversal of prostaglandin F2alpha-induced contraction of guinea-pig superfused trachea in vitro. The relaxant response of the non-selective PDE isoenzyme inhibitor trequinsin was slow in onset and demonstrated very slow recovery, similar to that observed with the long-acting beta2-adrenoceptor agonist, salmeterol and the PDE4 inhibitor, RP 73401. The relaxant agonists also significantly reversed bombesin-induced bronchospasm in anaesthetised guinea-pigs and there was a highly significant correlation between the ability of drugs to reverse PGF2alpha-induced contraction of guinea-pig isolated trachea in vitro and bombesin-induced bronchoconstriction in vivo. Furthermore, both salmeterol and trequinsin demonstrated long lasting bronchodilator responses consistent with the in vitro data. These results show that PDE isoenzyme inhibitors demonstrate different pharmacodynamic profiles that is not determined by PDE4 inhibitory potency and indicate that other factors may be important in this regard.

The role of dopaminergic agonists in congestive heart failure.

Congestive Heart Failure is a clinical syndrome characterised by myocardial dysfunction and sympathetic activation. Plasma norepinephrine (NE) levels have been related to the poorest survival. Large-scale clinical trials have proved the clinical benefits of Angiotensin Converting Enzyme inhibitors in reducing the risk of death and hospitalisation. However, mortality remains high in the treated group underlining the need to explore new therapeutic approaches. Specific activation of peripheral dopamine receptors exerts profound hemodynamic effects and neurohormonal control such as peripheral and renal vasodilation, diuresis and natriuresis and inhibition of NE release. Z1046, a mixed D1/D2-like agonist, reduces peripheral and renal vascular resistance increasing renal blood flow. In anaesthetised dogs the compound strongly reduces plasma NE without increasing plasma renin activity and plasma aldosterone. The inhibition of NE could be the basis of Z1046 potent cardioprotective effect observed in a dog model of myocardial ischemia and reperfusion, in which the severity of ventricular arrhythmias was markedly reduced, resulting in higher survival. These findings suggest that chronic oral treatment with specific dopaminergic agonists is able to alleviate the hemodynamic burden on the myocardium, and to suppress the sympatho-adrenal activity.

Effects of dopamine and selective dopamine agonists upon platelet accumulation in the cerebral and pulmonary vasculature of the rabbit.

1. A selection of novel compounds were shown to exhibit dopaminergic activity in vitro. 2. 111Indium-labelled platelets were continuously monitored in the cerebral and pulmonary vasculature of anaesthetized rabbits. The effects of dopamine and selective dopamine receptor agonists on ADP and thrombin induced platelet accumulation were recorded. 3. Pretreatment with dopamine (2 mg kg(-1) min(-1), i.v.) significantly reduced ADP (20 microg kg(-1), i.v.) induced platelet accumulation in the pulmonary vasculature whereas lower doses had no effect. 4. Dopamine (100 microg kg(-1) min(-1) intra-carotid, i.c.) potentiated thrombin (90 u kg(-1), i.c.) induced platelet accumulation in the cerebral vasculature whereas higher doses (1-2 mg kg(-1) min[-1]) inhibited accumulation. 5. The selective dopamine receptor agonists tested did not significantly inhibit platelet accumulation induced by ADP or thrombin. Two of these selective agonists, at doses higher than the intended therapeutic doses, induced thrombocytopaenia and an associated increase in platelet accumulation in the lung in response to thrombin. 6. These results extend previous in vitro studies regarding the dual actions of dopamine upon platelets and show for the first time the effects of selective dopamine receptor agonists upon platelet aggregation in vivo.

Z1046: a new specific peripheral dopaminergic compound.

It is well established that peripheral dopamine receptors activation evokes vasodilation and neurohormonal modulation. Z1046 is a potent mixed dopaminergic agonist and is highly selective over adrenergic and serotoninergic (5-HT2) activities. In contrast, dopamine had agonist activities on all adrenergic receptors while it is known that dopexamine has a beta 2 agonist activity and is an inhibitor of the neuronal uptake. As far as fenoldopam is concerned, selective stimulation of D1-like receptors leads to an increase of renin release. In conclusion, Z1046 is a potent and specific drug for dopamine receptors. This profile makes Z1046 different from other dopaminergic agents.

Characterization of big endothelin-1-induced contraction in rabbit saphenous artery.

We have characterized the endothelin-converting enzyme (ECE)-like activity involved in big endothelin (ET)-1-induced contraction in rabbit saphenous artery (RSA). Big ET-1 30 nM caused a contraction that was independent of the vascular endothelium. Phosphoramidon and the neutral endopeptidase (NEP) inhibitors thiorphan and candoxatrilat blocked the vasoconstriction caused by big ET-1 in endothelium-denuded RSA. Candoxatrilat (IC50 17 nM) and thiorphan (IC50 2.5 nM), were 5- to 30-fold more potent than phosphoramidon (IC50 83 nM). Other protease inhibitors were inactive. In cultured endothelial cells the ET-1 release was inhibited only by phosphoramidon (IC50 16 microM) but at a concentration 200-fold that required an endothelium-denuded RSA. In conclusion, we can speculate that the big ET-1 contraction in RSA is mediated by an ECE, probably present on smooth muscle cells, which is susceptible to NEP inhibitors and is different from the ECE on endothelial cells.

Effects of high-dose methylprednisolone on Na(+)-K+ ATPase and lipid peroxidation after experimental subarachnoid hemorrhage.

The production of oxygen-free radicals and their subsequent peroxidative action on membrane unsaturated fatty acids could be enhanced after subarachnoid hemorrhage. High-dose methylprednisolone (30 mg/Kg i.v.) treatment can antagonize acute SAH-induced brain hypoperfusion and protect the ultrastructural integrity of endothelial cell membranes. Experimental subarachnoid hemorrhage (SAH) was induced in anesthesized rats by slow injection of 0.3 ml of autologous arterial blood into cisterna magna. Tissue lipid peroxidation, quantified as thiobarbituric acid reactive material (TBAR) and Na(+)-K+ ATPase activity were assayed in three different rat brain areas (cerebral cortex, hippocampus and brain stem) of controls (without any surgical manipulation), sham-operated (0.3 ml. of mock CSF into cisterna magna) and after SAH induction, at 1 h, 6 h and 48 h. Na(+)-K+ ATPase activity decreased in the cerebral cortex at 1 h and 6 h and in brain stem at 1 h after SAH, while the same enzymatic activity was unchanged in the hippocampus. High-dose methyl-prednisolone treatment (started immediately after SAH induction) enhanced the Na(+)-K+ ATPase activity until control levels. There was no significant difference in lipid peroxide content between sham-operated and hemorrhagic animals; however, the injection itself induces a transient increase of TBAR (1 h after injection) and methylprednisolone treatment decreases the products of lipid peroxidation in all brain areas.

High-dose methylprednisolone and 'ex vivo' release of eicosanoids after experimental subarachnoid haemorrhage.

In the present study we have investigated the effects of high-dose methylprednisolone (MP) treatment on the 'ex vivo' release of four major eicosanoids in an experimental model of subarachnoid haemorrhage (SAH) with the aim of verifying: (a) the efficacy in reducing arachidonic acid metabolism enhancement; (b) whether high-dose methylprednisolone is effective on both the cyclooxygenase and lipoxygenase pathways; and (c) discussing the possible role of high-dose MP treatment in brain protection after SAH. Levels of prostaglandin D2 and E2, prostacyclin and also leukotriene C4 were determined by the radioimmunoassay technique after 1 h incubation of cerebral cortex samples of rats which had been subjected to experimental SAH procedure (injection of 0.3 ml of autologous arterial blood). The release of prostaglandin D2 at 48 h after SAH is significantly higher when compared to that of sham-operated animals (P less than 0.01); prostaglandin E2 release is significantly enhanced at 6 h after the SAH procedure (P less than 0.01); release of the lipoxygenase metabolite is significantly enhanced at 1, 6 and 48 h after SAH induction; MP significantly decreases the release of all eicosanoids, and values in treated animals do not differ from those of sham-operated animals. The results of the present study suggest that the global inhibitory effect of high-dose MP treatment on the 'ex vivo' release of eicosanoids after experimental SAH could be considered to be one of the neurochemical correlates for the reduced incidence and severity of arterial inflammatory response, which results in chronic vasospasm and supports the clinical evidence of MP efficacy in preventing or reducing the incidence of arterial vasospasm after aneurysmal rupture.

Experimental subarachnoid hemorrhage. Lipid peroxidation and Na+,K(+)-ATPase in different rat brain areas.

Subarachnoid hemorrhage (SAH) was produced in Sprague Dawley rats by injection of 0.30 mL of autologous arterial blood into the cisterna magna. Tissue lipid peroxide, quantified as thiobarbituric acid reactive material (TBAR), and Na+,K(+)-ATPase activity were assayed in three different rat brain areas (cerebral cortex, hippocampus, and brain stem) of sham-operated rats and in four hemorrhagic rat groups at 30 min, 1 h, 6 h, and 2 d after SAH. Na+,K(+)-ATPase activity decreased in the cerebral cortex at 30 min, 1 h, and 6 h and in the brain stem at 1 h after SAH induction, whereas enzymatic activity was unchanged in the hippocampus. There was no evident difference in lipid peroxide content between sham-operated animals and hemorrhagic animals. These results indicate that little modifications in lipid peroxidative process (as expressed in TBAR) are not responsible for changes in the ATPase activity.