Platelet thromboxane synthetase inhibitors with low doses of aspirin: possible resolution of the "aspirin dilemma".

Selective pharmacological inhibition of thromboxane A2 synthesis did not prevent arachidonate-induced aggregation of human platelets in vitro. Prevention was instead achieved by a combination of thromboxane A2 inhibitors with low concentrations of aspirin. The latter partially reduced the proaggregatory cyclooxygenase products that accumulated when thromboxane A2 synthesis was blocked. The aspirin concentrations did not affect per se either platelet aggregation or prostacyclin synthesis in cultured human endothelial cells. The combination of thromboxane synthetase inhibitors with low doses of aspirin may offer greater antithrombotic potential than either drug alone.

Provision of information on regulatory authorities' websites.

BACKGROUND: Several organizations have raised concerns about the excessive secrecy maintained by regulatory authorities around the world, in particular in the European Union, France, UK, Canada and Australia. However, limited research has assessed the provision of information by regulatory authorities. This study aimed to assess the type and availability of information provided on the regulatory authorities' websites. METHODS: Regulatory authorities' websites in six countries (USA, Canada, UK, France, Australia and New Zealand) and at the European level (European Medicines Evaluation Agency) were surveyed by two reviewers between October 2005 and March 2006. The survey instrument included 16 criteria organized in 3 domains: information on marketed drugs, information on assessment of drugs and information on drug safety. RESULTS: There was a great variability in the level of information provided. Several medicine agencies did not provide basic information on marketed drugs, such as the summary of products' characteristics. Information on registration dossiers was scant on most websites except that of the US Food and Drug Administration. The European Medicines Evaluation Agency, the French agency and the Canadian agency released public assessment reports that contained only summarized information of registration data. Only one country, Canada, provided full access to pharmacovigilance data. The periodic safety update reports that companies have to provide regularly to regulatory authorities were not available in any country. CONCLUSION: Information on which regulatory authorities base their decisions for licensing new drugs and the rationales behind these decisions were often not publicly available.

Heparin counteracts the antiaggregating effect of prostacyclin by potentiating platelet aggregation.

It has recently been reported that heparin neutralizes the inhibitory effect of prostacyclin (PGI2) on human platelet aggregation. The mechanism of this interaction has not yet been unequivocally established. We present here evidence that heparin (Liquemin Roche) does not react directly with PGI2 but counteracts its inhibitory effect by potentiating platelet aggregation. In the absence of heparin, PGI2 was a less effective inhibitor of platelet aggregation induced by the combination of ADP and serotonin than by ADP alone. Moreover, the inhibitory effect of PGI2 was similarly reduced when increasing the concentrations of ADP (in the absence of heparin). The lack of a specific interaction between heparin and PGI2 is supported by the observation that, in the presence of heparin, other prostaglandins such as PGD2 and PGE1 and a non-prostanoid compound such as adenosine also appeared to lose their inhibitory potency. It is concluded that heparin opposes platelet aggregation inhibitory effect of PGI2 by enhancement of platelet aggregation.

SQ 22536, an adenylate-cyclase inhibitor, prevents the antiplatelet effect of dazoxiben, a thromboxane-synthetase inhibitor.

This study shows that dazoxiben, a selective inhibitor of thromboxane A2-synthetase in human platelets, inhibited arachidonic acid-induced platelet aggregation in platelet-rich plasma samples from four out of 16 healthy volunteers. In these four "responder" samples, the anti-aggregating effect of dazoxiben was prevented by the compound SQ 22536, a 9-substituted adenine analogue, endowed with an inhibitory activity on adenylate-cyclase. The compound SQ 22536 also counteracted the antiaggregating effect of prostaglandin D2, a known activator of platelet adenylate-cyclase. When platelet thromboxane A2-synthetase was blocked by dazoxiben, a marked increase of prostaglandin D2 was concomitantly observed both in "responder" and "non responder" samples. The compound SQ 22536 blunted the increase in platelet cAMP caused by either dazoxiben and sodium arachidonate or prostaglandin D2. It is suggested that the antiaggregating effect of dazoxiben is mediated by newly synthesized prostaglandin D2. The latter acts by stimulating adenylate-cyclase and increasing cAMP levels. The compound SQ 22536 prevents both phenomena. In "non responder" samples some factors--still to be defined--might counteract similarly to the compound SQ 22536 the antiaggregating activity of PGD2.

The inhibitory effect of aspirin on fibrinolysis is reversed by iloprost, a prostacyclin analogue.

The reduced fibrinolytic response after aspirin intake may be due to prevention of prostacyclin production. The effect of iloprost (a stable prostacyclin analogue) was tested on the fibrinolytic activity (euglobulin lysis area on fibrin plate [E.L.A.], t-PA antigen, PAI activity and PAI-1 antigen) of plasma drawn after venous stasis test from six healthy male volunteers, who each received all the following treatments according to a single-blind randomized cross-over design: placebo, iloprost, aspirin + placebo, aspirin + iloprost. The mean E.L.A. value after venous occlusion was significantly higher than the basal level after every treatment but aspirin. Within each treatment group the t-PA antigen levels in response to venous stasis were significantly higher than the basal ones. PAI-1 antigen levels did not change significantly before and after venous stasis either within or among the treatment groups. These data are consistent with the hypothesis that the mechanism related to aspirin's effect on fibrinolysis is mediated by suppression of vessel wall prostacyclin production. Aspirin's inhibitory effect on fibrinolysis was in fact prevented by replacing endogenous prostacyclin with iloprost. Iloprost enhances fibrinolytic activity reduced by aspirin, but not by promoting t-PA release or by inhibiting release of the specific inhibitor, PAI-1.

Defective fibrinolytic response in atherosclerotic patients--effect of iloprost and its possible mechanism of action.

Plasma fibrinolytic activity and tissue-type plasminogen activator (t-PA) were defective in response to venous stasis in five out of ten patients with peripheral occlusive artery disease. Discontinuous infusions of iloprost, a stable synthetic analogue of prostacyclin, restored a normal fibrinolytic response in all five patients but did not induce a parallel increase of plasma t-PA. These findings suggest that in addition to the possible benefits due to its vasodilatory and antiplatelet activity, iloprost may improve the fibrinolytic activity in patients with atherosclerotic disease, providing them with further antithrombotic protection. The profibrinolytic effect of iloprost seems not to depend on its ability to induce vascular t-PA release. Rather, it might be related to its inhibitory effect on PAI release from platelets, endothelial cells and/or hepatocytes. Venous occlusion test represents an easy diagnostic approach to fibrinolytic defects, even if related to arterial disease, and may help select patients who need therapeutic intervention.

Effects of any epoxymethano stable analogue of prostaglandin endoperoxides (U-46619) on human platelets.

U-46619 is a stable epoxymethano analogue of cyclic endoperoxide PGH2. We studied platelet aggregation, 14C-5HT release, LDH extrusion and prostaglandin and thromboxane production induced by this compound in platelet-rich plasma samples from 15 healthy volunteers. Each subject was tested both before and 90 min after aspirin (500 mg) ingestion. The threshold aggregating concentration (TAC) of U-44619 ranged between 0.18 and 0.90 micro M. Aggregation was maximal between 40 and 60 min after venipuncture and was concentration-dependent. At concentrations below the TAC, U-44619 induced primary reversible aggregation with minimal 14C-5HT release. At TAC or higher concentrations aggregation and release proceeded as parallel events. Neither prostaglandin or thromboxane production nor LDH loss could be detected in any of the situations tested. Aspirin ingestion did not modify the pattern of platelet responses. In unstirred, not aggregated platelet samples 14C-5HT release by U-46619 occurred to a similar extent as in stirred, aggregated platelet samples. Addition to citrated PRP of 0.3 mM Na2 EDTA blocked both aggregation and release induced by U-46619. This compound, however, aggregated washed platelets resuspended in Ca++-free-tyrode-albumin containing fibrinogen. The mechanism by which U-46619 activates platelets differs from that of all other common aggregating agents.

Prostaglandins and human platelet aggregation. Implications for the anti-aggregating activity of thromboxane-synthase inhibitors.

Selective pharmacological blockade of thromboxane-synthase in human platelets by dazoxiben resulted in the reorientation of cyclic-endoperoxides towards PGE2, PGD2 and PGF2 alpha. At concentrations which can be reached when thromboxane-synthase is inhibited, PGE2 (100-500 nM) exerted a marked, concentration-dependent pro-aggregatory effect. This required the formation of endogenous or the addition of exogenous endoperoxides and was prevented by PGD2 or 13-aza-prostanoic acid, a selective antagonist of PGH2/TxA2 receptors. The anti-aggregating effect of PGD2 was evident at concentrations lower than those obtained in dazoxiben-treated platelets. It is proposed that in the absence of TxA2 generation, a combination of endoperoxides and PGE2 may result in normal aggregation. The latter may be inhibited by PGD2. No interference of PGF2 alpha on platelet function could be shown.

Potentiation by dazoxiben, a thromboxane synthetase inhibitor, of platelet aggregation inhibitory activity of a thromboxane receptor antagonist and of prostacyclin.

Dazoxiben, a thromboxane (Tx)-synthetase inhibitor, completely prevented platelet TxB2 synthesis, but not the platelet aggregation induced by arachidonic acid (AA) or ADP. It was also ineffective against platelet aggregation brought about by a prostaglandin (PG) endoperoxide analogue, which does not trigger platelet TxA2 synthesis. The association of dazoxiben with a Tx receptor antagonist or with PGI2 resulted in marked potentiation of the latter compounds as inhibitors of platelet aggregation induced by AA or ADP (second wave), but not by U-46619 (a stable analogue of prostaglandin endoperoxides). It therefore appears that inhibition of Tx synthesis does not modify the platelet response to aggregating stimuli but renders platelets more susceptible to inhibition induced by other compounds.

Arachidonic acid induces human platelet-fibrin retraction: the role of platelet cyclic endoperoxides.

Arachidonic acid (0.2-0.8 mM) retracts clots formed in human citrated platelet-rich plasma by batroxobin. Extracellular calcium ions, but not the secretion of ADP by platelets, are required. AA-induced clot-retraction requires cyclo-oxygenase but not thromboxane synthetase activity since the retraction is inhibited by aspirin but not by selective inhibitors of thromboxane synthesis. The data indicate that endogenous cyclic endoperoxides mediate the retraction. Moreover, intact endoperoxide/thromboxane receptors also seem to be necessary because clot retraction is inhibited by thromboxane receptor antagonists.