The novel anti-inflammatory agent VA694, endowed with both NO-releasing and COX2-selective inhibiting properties, exhibits NO-mediated positive effects on blood pressure, coronary flow and endothelium in an experimental model of hypertension and endothelial dysfunction.

Selective cyclooxygenase 2 (COX2) inhibitors (COXIBs) are effective anti-inflammatory and analgesic drugs with improved gastrointestinal (GI) safety compared to nonselective nonsteroidal anti-inflammatory drugs known as traditional (tNSAIDs). However, their use is associated with a cardiovascular (CV) hazard (i.e. increased incidence of thrombotic events and hypertension) due to the inhibition of COX2-dependent vascular prostacyclin. Aiming to design COX2-selective inhibitors with improved CV safety, new NO-releasing COXIBs (NO-COXIBs) have been developed. In these hybrid drugs, the NO-mediated CV effects are expected to compensate for the COXIB-mediated inhibition of prostacyclin. This study evaluates the potential CV beneficial effects of VA694, a promising NO-COXIB, the anti-inflammatory effects of which have been previously characterized in several in vitro and in vivo experimental models. When incubated in hepatic homogenate, VA694 acted as a slow NO-donor. Moreover, it caused NO-mediated relaxant effects in the vascular smooth muscle. The chronic oral administration of VA694 to young spontaneously hypertensive rats (SHRs) significantly slowed down the age-related development of hypertension and was associated with increased plasma levels of nitrates, stable end-metabolites of NO. Furthermore, a significant improvement of coronary flow and a significant reduction of endothelial dysfunction were observed in SHRs submitted to chronic administration of VA694. In conclusion, VA694 is a promising COX2-inhibiting hybrid drug, showing NO releasing properties which may mitigate the CV deleterious effects associated with the COX2-inhibition.

Clinical pharmacology of cyclooxygenase inhibition and pharmacodynamic interaction with aspirin by floctafenine in Thai healthy subjects.

Floctafenine, a hydroxyquinoline derivative with analgesic properties, is widely used in Thailand and many other countries. The objectives of this study were to evaluate in Thai healthy volunteers: i) the inhibition of whole blood cyclooxygenase(COX)-2 and COX-1 activity by floctafenine and its metabolite floctafenic acid in vitro and ex vivo after dosing with floctafenine; ii) the possible interference of floctafenine administration with aspirin antiplatelet effects. We performed an open-label, cross-over, 3-period study, on 11 healthy Thai volunteers, who received consecutively floctafenine(200mg/TID), low-dose aspirin(81mg/daily) or their combination for 4 days, separated by washout periods. Floctafenine and floctafenic acid resulted potent inhibitors of COX-1 and COX-2 in vitro (floctafenic acid was more potent than floctafenine) showing a slight preference for COX-1. After dosing with floctafenine alone, whole blood COX-1 and COX-2 activities were inhibited ex vivo in a time-dependent fashion which paralleled floctafenic acid plasma concentrations. Aspirin alone inhibited profoundly and persistently platelet COX-1 activity and AA-induced platelet aggregation throughout 24-h dosing interval which was affected by the co-administration of floctafenine. At 24 h after dosing with aspirin and floctafenine, the inhibition of platelet thromboxane(TX)B2 generation and aggregation were significantly(P less than 0.05) lower than that caused by aspirin alone. Therapeutic dosing with floctafenine profoundly inhibited prostanoid biosynthesis through the rapid conversion to floctafenic acid. Floctafenine interfered with the antiplatelet effect of aspirin. Our results suggest that floctafenine should be avoided in patients with cardiovascular disease under treatment with low-dose aspirin.

Selective cumulative inhibition of platelet thromboxane production by low-dose aspirin in healthy subjects.

Acetylation of platelet cyclooxygenase by oral aspirin is dose dependent and cumulative with repeated administration. However, no single dose of aspirin has been found to be completely selective of platelet thromboxane (TX) synthesis inhibition in man. We determined the dose dependence, cumulative nature and selectivity of aspirin effects on platelet TXB(2) and renal prostaglandin (PG) and prostacyclin (PGI(2)) production. We measured, by radioimmunoassay, serum TXB(2) levels after whole blood clotting and urinary excretion of PGE(2), PGF(2alpha), and 6-keto-PGF(1alpha), before and after single or repeated oral aspirin doses given to 46 healthy subjects. Single doses of 6-100 mg aspirin resulted in a linear (r = 0.92, P < 0.01) inhibition of platelet TXB(2) production, ranging from 12 to 95% after 24 h. A daily dose of 0.45 mg/kg given for 7 d produced a cumulative and virtually complete inhibition of platelet TXB(2) production, without significantly reducing the urinary excretion of PGE(2), PGF(2alpha), and 6-keto-PGF(1alpha) in both healthy men and women. The platelet inhibitory effect of this regimen was maintained unaltered throughout 1 mo of therapy, with no evidence of cumulative inhibition of renal PG-synthesis. Moreover, furosemide-induced renal PGI(2) synthesis and renin release were unaffected by chronic low-dose aspirin. Following cessation of aspirin therapy, platelet TXB(2) production returned toward control values at a similar rate as after a single higher dose. WE CONCLUDE THAT IN HEALTHY SUBJECTS: (a) aspirin causes a dose-dependent inhibition of platelet TXA(2) production, with no obvious sex-related difference; (b) the inhibitory effect of daily low-dose aspirin is cumulative on platelet TXA(2) but not on renal PG-synthesis; (c) during chronic low-dose aspirin therapy, renal PGI(2)-producing cells are readily activable by furosemide at a time of virtually complete suppression of platelet cyclooxygenase activity.

Evidence for a direct stimulatory effect of prostacyclin on renin release in man.

THE OBJECTIVES OF THIS INVESTIGATION WERE: (a) to characterize the time and dose dependence of the effects of prostacyclin (PGI(2)) on renin release in healthy men; (b) to define whether PGI(2)-induced renin release is secondary to hemodynamic changes; (c) to determine the plasma and urine concentrations of 6-keto-PGF(1alpha) (the stable breakdown product of PGI(2)) associated with renin release induced by exogenous or pharmacologically enhanced endogenous PGI(2). Intravenous PGI(2) or 6-keto-PGF(1alpha) infusions at nominal rates of 2.5, 5.0, 10.0, and 20.0 ng/kg per min were performed in each of six normal human subjects; in three of them, PGI(2) infusion was repeated after beta-adrenergic blockade and cyclooxygenase inhibition. PGI(2), but not 6-keto-PGF(1alpha), caused a time- and dose-dependent increase of plasma renin activity, which reached statistical significance at 5.0 ng/kg per min and was still significantly elevated 30 min after discontinuing the infusion. Although combined propranolol and indomethacin treatment significantly enhanced the hypotensive effects of infused PGI(2), it did not modify the dose-related pattern of PGI(2)-induced renin release. Plasma 6-keto-PGF(1alpha) levels rose from undetectable levels (<7.5 pg/ml) in a stepwise fashion during increasingly higher infusion rates of PGI(2) or 6-keto-PGF(1alpha). The threshold concentration of plasma 6-keto-PGF(1alpha) associated with a statistically significant stimulation of renin release was approximately 200 pg/ml. Upon discontinuing PGI(2) or 6-keto-PGF(1alpha) infusion, the disappearance of 6-keto-PGF(1alpha) from blood showed an identical biphasic behavior, the initial phase having an apparent t((1/2)) of 3.2 min. The intravenous infusion of furosemide, which is known to stimulate renin release via a cyclooxygenase-dependent mechanism, caused a three-to fourfold increase of urinary 6-keto-PGF(1alpha) excretion rate, concomitant with the elevation of plasma renin activity levels, in six healthy women. 6-Keto-PGF(1alpha) remained undetectable in peripheral venous plasma throughout the study. WE CONCLUDE THAT IN HUMAN SUBJECTS: (a) PGI(2)-induced renin release occurs with a dose and time dependence similar to its reported platelet effects; (b) PGI(2)-induced renin release is not mediated by adrenergic stimuli or cyclooxygenase-dependent mechanisms secondary to hemodynamic changes; (c) furosemide-induced renin release is associated with increased renal PGI(2) formation; and (d) PGI(2) appears to act as a local modulator rather than a circulating hormone in controlling juxtaglomerular function.

Reduced platelet thromboxane formation in uremia. Evidence for a functional cyclooxygenase defect.

A qualitative platelet abnormality and a bleeding tendency are frequently associated with renal failure and uremia. We demonstrated previously that uremic patients display an abnormal platelet aggregation to arachidonic acid and reduced malondialdehyde production in response to thrombin and arachidonic acid. The objectives of this investigation were: (a) to compare platelet prostaglandin (PG) and thromboxane (TX) production in whole blood and in platelet-rich plasma (PRP) of 21 uremic patients and 22 healthy subjects; (b) to evaluate the concentration and activity of platelet PG- and TX-forming enzymes; (c) to assess the functional responsiveness of the platelet TXA(2)/PGH(2) receptor; (d) to explore the hemostatic consequences of partially reduced TXA(2) production.Platelet immunoreactive TXB(2) production during whole blood clotting was significantly reduced, by approximately 60%, in uremic patients as compared to age- and sex-matched controls. Exogenous thrombin (5-30 IU/ml) failed to restore normal TXB(2) production in uremic platelets. Uremic PRP produced comparable or slightly higher amounts of TXB(2) than normal PRP at arachidonate concentrations 0.25-1 mM. However, when exposed to substrate concentrations >2 mM, uremic PRP produced significantly less TXB(2) than normal PRP. To discriminate between reduced arachidonic acid oxygenation and altered endoperoxide metabolism, the time course of immunoreactive TXB(2) and PGE(2) production was measured during whole blood clotting. The synthesis and release of both cyclooxygenase-derived products was slower and significantly reduced, at all time intervals considered. Furthermore, PGI(2) production in whole blood, as reflected by serum immunoreactive 6-keto-PGF(1alpha) concentrations, was significantly reduced in uremic patients as compared with healthy subjects. PGH synthase levels, as determined by an immunoradiometric assay, were not significantly different in platelets from uremic patients as compared to control platelets. A single 40-mg dose of aspirin given to five healthy volunteers reduced their serum TXB(2) to levels found in uremic patients. This was associated with a significant increase of threshold aggregating concentrations of ADP and arachidonic acid and prolongation of bleeding time. Substantially similar threshold concentrations of U46619, a TXA(2) agonist, induced aggregation of normal and uremic platelets. Prostacyclin induced a significant elevation of uremic platelet cyclic AMP, which was suppressed by U46619, further suggesting normal responsiveness of the TXA(2)/PGH(2) receptor. WE CONCLUDE THAT: (a) an abnormality of platelet arachidonic acid metabolism exists in uremia, leading to a reduced TXA(2) production; (b) the characteristics of this abnormality are consistent with a functional cyclooxygenase defect; (c) reduced TXA(2) production may partially explain the previously described abnormality of platelet function in uremia.

Low-Dose Aspirin Acetylates Cyclooxygenase-1 in Human Colorectal Mucosa: Implications for the Chemoprevention of Colorectal Cancer.

The mechanism of action of low-dose aspirin in the prevention of colorectal cancer (CRC) remains largely hypothetical. We aimed to compare the effects of low-dose aspirin (100 mg/day for 7 days) given to 40 individuals undergoing CRC screening on the extent of cyclooxygenase (COX)-1 acetylation at serine-529 (AceCOX-1), in blood platelets vs. colorectal mucosa, at 7 (group 1) and 24 h (group 2) after dosing. A significantly (P < 0.01) lower %AceCOX-1 was detected in colonic and rectal mucosa (average 64%) vs. platelets (average 75%) in both groups. This effect was associated with an average 46% (P < 0.01) and 35% (P < 0.05) reduction in prostaglandin (PG) E2 levels and phosphorylated S6 (p-S6) levels, respectively. Rectal mucosal levels of p-S6/S6 significantly (P < 0.01) correlated with PGE2 . These findings demonstrate that low-dose aspirin produces long-lasting acetylation of COX-1 and downregulation of p-S6 in human colorectal mucosa, an effect that may interfere with early colorectal carcinogenesis.

[Italian consensus on EULAR recommendations 2005 for the management of hip osteoarthritis]. / Consensus italiana sulle raccomandazioni EULAR 2005 per il trattamento dell'artrosi dell'anca.

The recommendations for the management of osteoarthritis (OA) of the hip were proposed by EULAR in 2005. Among the most important objectives of the expert charged to provide these recommendations were their wide dissemination and implementation. Thus, the information generated can be used by each individual country to produce their own set of management guidelines and algorithms for treatment in primary care. According with that previously executed for the EU-LAR recommendation 2003 for the knee, the Italian Society of Rheumatology (SIR) has organised a Consensus on the EULAR recommendations 2005 for the management of hip OA. To obtain an acceptability as large as possible, the group of experts was composed by many physicians interested in the management of hip OA, including Orthopaedics, Rheumatologists, Physiatrists, and General Practitioners. Main aim of the Consensus was to analyse the acceptability and applicability of the recommendations according to own experience and local situations in the Italy. The results of this Consensus have demonstrated that a large majority of the EULAR recommendations are endorsed by the Italian experts. Furthermore, the final document of the Italian Consensus clearly indicated the need that the specialists involved in the management of hip OA strongly encourage the dissemination of the EULAR 2005 recommendations also in Italy.

Fractional conversion of thromboxane A2 and B2 to urinary 2,3-dinor-thromboxane B2 and 11-dehydrothromboxane B2 in the cynomolgus monkey.

Following the intravenous administration of thromboxane (TX) B2, the stable hydration product of TXA2, to human and nonhuman primates the most abundant urinary metabolites are 2,3-dinor-TXB2 and 11-dehydro-TXB2. However, it is not known whether fractional conversion of TXB2 to its enzymatic metabolites is an accurate representation of TXA2 metabolism. Thus, we have compared the metabolic disposition of synthetic TXA2 and TXB2 via the beta-oxidation and 11-OH-dehydrogenase pathways in vivo in the monkey. TXA2 or TXB2 (20 ng/kg) was intravenously administered to four cynomolgus monkeys pretreated with aspirin in order to suppress endogenous TXA2 production. Urinary TXB2, 2,3-dinor-TXB2 and 11-dehydro-TXB2 were measured before, during and up to 24 h after thromboxane administration by means of reversed-phase high-performance liquid chromatography radioimmunoassay. Aspirin treatment suppressed urinary 2,3-dinor-TXB2 and 11-dehydro-TXB2 by approx. 75%. A similar fractional conversion of TXA2 and TXB2 into 2,3-dinor-TXB2 and 11-dehydro-TXB2 was found. These results suggest that TXA2 is hydrolyzed to TXB2 prior to enzymatic degradation and that metabolites of the latter represent reliable indices of TXA2 biosynthesis. Due to the variability in the conversion of thromboxanes into 2,3-dinor-TXB2 and 11-dehydro-TXB2, the measurement of both metabolites seems to represent a more reliable index of acute changes in TXA2 production.

Calcium mobilization and right-angle light scatter responses to 12-oxo-derivatives of arachidonic acid in neutrophils: evidence for the involvement of the leukotriene B4 receptor.

The biological activities of two carbonyl compounds derived from arachidonic acid, (5Z,8Z,10E,14Z)-12-keto-5,8,10,14-eicosatetraeno ic acid (12-OxoETE) and (5Z,8Z,10E)-12-oxo-5,8,10-dodecatrienoic acid (12-OxoDTrE) were investigated. The ability of these compounds to induce a mobilization of calcium and to trigger a right-angle scatter response in isolated peripheral blood human neutrophils was determined. The two compounds induced a rapid and dose-dependent increase in the concentration of cytoplasmic free calcium; these effects were clearly detectable at concentrations greater than or equal to 10(-8) M. Pre-exposure of neutrophils to leukotriene B4 completely abolished the calcium mobilization induced by 12-OxoDTre and 12-OxoETE, while pre-exposure of the cells to the carbonyl compounds only slightly reduced the response to subsequent stimulation of neutrophils by leukotriene B4. The carbonyl compounds also induced a decrease in right-angle light scatter and these effects were abolished by pretreatment of neutrophils with leukotriene B4. These data demonstrate that 12-OxoETE and 12-OxoDTrE show significant agonist activities towards human neutrophils and strongly suggest that their mechanisms of action involve the leukotriene B4 binding sites or a common activation sequence.

Oxidant stress and aspirin-insensitive thromboxane biosynthesis in severe unstable angina.

BACKGROUND: Unstable angina is associated with enhanced lipid peroxidation and reduced antioxidant defenses. We have previously reported aspirin failure in the suppression of enhanced thromboxane (TX) biosynthesis in a subset of episodes of platelet activation in this setting. We tested the hypothesis that the in vivo formation of the F(2)-isoprostane 8-iso-prostaglandin (PG)F(2alpha), a bioactive product of arachidonic acid peroxidation, is enhanced in unstable angina and contributes to aspirin-insensitive TX biosynthesis. METHODS AND RESULTS: Urine samples were obtained from patients with unstable angina (n=32), stable angina (n=32), or variant angina (n=4) and from 40 healthy subjects for the measurement of immunoreactive 8-iso-PGF(2alpha) and 11-dehydro-TXB(2). 8-Iso-PGF(2alpha) excretion was significantly higher in patients with unstable angina (339+/-122 pg/mg creatinine) than in matched patients with stable angina (236+/-83 pg/mg creatinine, P:=0.001) and control subjects (192+/-71 pg/mg creatinine, P:<0.0001). In patients with unstable angina, 8-iso-PGF(2alpha) was linearly correlated with 11-dehydro-TXB(2) excretion (rho=0.721, P:<0.0001) and inversely correlated with plasma vitamin E (rho=-0.710, P:=0. 004). Spontaneous myocardial ischemia in patients with variant angina or ischemia elicited by a stress test in patients with stable angina was not accompanied by any change in 8-iso-PGF(2alpha) excretion, thus excluding a role of ischemia per se in the induction of increased F(2)-isoprostane production. CONCLUSIONS: These findings establish a putative biochemical link between increased oxidant stress and aspirin-insensitive TX biosynthesis in patients with unstable angina and provide a rationale for dose-finding studies of antioxidants in this setting.

Effects of vitamin E supplementation on F(2)-isoprostane and thromboxane biosynthesis in healthy cigarette smokers.

BACKGROUND: Increased formation of 8-iso-prostaglandin (PG) F(2alpha) and thromboxane (TX) A(2), potent agonists of platelet and vascular thromboxane (TH)/PGH(2) receptors, has been detected in cigarette smokers. We performed a randomized, double-blind, placebo-controlled study of the effects of vitamin E (300, 600, and 1200 mg/d, each dose for 3 consecutive weeks) on 8-iso-PGF(2alpha) and TXA(2) biosynthesis in 46 moderate cigarette smokers. METHODS AND RESULTS: Urinary immunoreactive 8-iso-PGF(2alpha) and 11-dehydro-TXB(2), plasma vitamin E, and serum TXB(2) were measured by previously validated techniques. Baseline urinary 8-iso-PGF(2alpha) and 11-dehydro-TXB(2) excretion averaged 241+/-78 and 430+/-293 pg/mg creatinine, respectively. Urinary 8-iso-PGF(2alpha) was significantly correlated with 11-dehydro-TXB(2) (r=0.360, n=138, P<0.0001). Baseline plasma vitamin E levels averaged 20.6+/-4.9 micromol/L and were inversely correlated with urinary 11-dehydro-TXB(2) (r=-0.304, P=0.039) but not with 8-iso-PGF(2alpha) (r=-0.227, P=0.129). Vitamin E supplementation caused a dose-dependent increase in its plasma levels that reached a plateau at 600 mg (42.3+/-11.2 micromol/L, P<0. 001). This was not associated with any statistically significant change in urinary 8-iso-PGF(2alpha) or 11-dehydro-TXB(2) excretion. CONCLUSIONS: Supplementation with pharmacological doses of vitamin E has no detectable effects on lipid peroxidation and thromboxane biosynthesis in vivo in healthy subjects with a mild degree of oxidant stress. These findings are consistent with the hypothesis that the basal rate of lipid peroxidation is a major determinant of the response to vitamin E supplementation and have implications for the use of vitamin E in healthy subjects as well as for the design and interpretation of clinical trials of antioxidant intervention.

Platelet activation in patients with colorectal cancer.

Aspirin may reduce the risk of colorectal neoplasia at doses similar to those recommended for the prevention of cardiovascular disease. Thus, we aimed to address whether enhanced platelet activation, as assessed by the measurement of the urinary excretion of 11-dehydro-TXB(2) (a major enzymatic metabolite of TXB(2)), occurs in patients with colorectal cancer. In 10 patients with colorectal cancer, the urinary excretion of 11-dehydro-TXB(2) was significantly higher than in 10 controls, matched for sex, age and cardiovascular risk factors [1001(205-5571) versus 409(113-984) pg/mg creatinine, respectively, median (range), P<0.05]. The administration of aspirin 50 mg daily for 5 consecutive days to colorectal cancer patients caused a cumulative inhibition of platelet cyclooxygenase (COX)-1 activity either ex vivo, as assessed by the measurement of serum TXB(2) levels, or in vivo, as assessed by urinary 11-dehydro-TXB(2) excretion. In conclusion, enhanced platelet activation occurs in colorectal cancer patients. Permanent inactivation of platelet COX-1 by low-dose aspirin might restore anti-tumor reactivity.

Effects of nabumetone on prostanoid biosynthesis in humans.

BACKGROUND: The active metabolite of the anti-inflammatory drug nabumetone has been characterized as a selective inhibitor of the inducible prostaglandin H synthase (PGHS). The aim of this study was to investigate the rate of eicosanoid biosynthesis after oral dosing with nabumetone in nine healthy subjects. METHODS: We measured the urinary excretion of products of platelet (11-dehydro-thromboxane B2 [TXB2]) and renal (prostaglandin IF2 alpha [PGF2 alpha]) arachidonate metabolism as in vivo indexes of the constitutive PGHS-1 pathway. Moreover, the production of TXB2 during whole blood clotting was assessed as an index of the cyclooxygenase activity of platelet PGHS-1 ex vivo. RESULTS: At steady state, nabumetone (500 and 1000 mg daily for 7 days) was associated with statistically significant dose-dependent reduction in the urinary excretion of 11-dehydro-TXB2 and serum TXB2 levels by approximately 50% to 70%. However, the drug did not significantly affect the urinary excretion of PGF2 alpha. After discontinuation of nabumetone, urinary 11-dehydro-TXB2 excretion and whole blood TXB2 production returned to predrug levels with a similar timecourse that was consistent with the elimination half-life of its active metabolite. The daily administration of low-dose aspirin (40 mg), a selective inhibitor of platelet PGHS-1, caused a cumulative inhibition of urinary 11-dehydro-TXB2 and whole blood TXB2 production that recovered with a timecourse consistent with platelet turnover. CONCLUSIONS: Nabumetone does dose-dependently inhibit the cyclooxygenase activity of platelet PGHS-1 of healthy subjects both in vivo and ex vivo. Thus it is unlikely that its safety profile in patients may be related to selective inhibition of the inducible PGHS-2.

The human pharmacology of monocyte cyclooxygenase 2 inhibition by cortisol and synthetic glucocorticoids.

BACKGROUND: We studied the concentration dependence of the inhibitory effects of cortisol, 6-methylprednisolone, and dexamethasone on cyclooxygenase-2 (COX-2) expression and activity in human monocytes in response to lipopolysaccharide (LPS) in vitro. Moreover, we characterized the time and dose dependence of the inhibitory effects of 6-methylprednisolone, administered to healthy subjects, on LPS-inducible prostaglandin E2 (PGE2) biosynthesis in whole blood ex vivo. METHODS: Heparinized whole-blood samples obtained from healthy subjects and patients with rheumatoid arthritis were incubated with LPS (10 microg/ml) for 24 hours at 37 degrees C, and PGE2 was measured in plasma as an index of monocyte COX-2 activity. Comparative experiments were performed in LPS-stimulated isolated monocytes. The levels of COX-2-like immunoreactivity in monocyte lysates were measured by a specific Western blot technique. PGE2 was evaluated by radioimmunoassay. RESULTS: Nanomolar concentrations of cortisol, 6-methylprednisolone, and dexamethasone suppressed LPS-induced PGE2 biosynthesis both in whole blood and in isolated monocytes in vitro with relative potencies similar to those reported for their anti-inflammatory effects in vivo. The administration of single oral doses (4, 8, or 16 mg) of 6-methylprednisolone caused a dose- and time-dependent inhibition of whole-blood COX-2 activity. Whole-blood samples obtained from patients with rheumatoid arthritis treated with comparable maintenance doses of glucocorticoids produced significantly lower levels of LPS-inducible PGE2 than were found in untreated patients. CONCLUSIONS: Therapeutic plasma levels of synthetic glucocorticoids down-regulate inducible prostanoid biosynthesis in circulating monocytes. This effect may represent a readily measurable surrogate marker of their clinical efficacy for dose-finding studies.

Effects of sulindac on renal and extrarenal eicosanoid synthesis.

We measured the renal and extrarenal synthesis of prostacyclin and thromboxane A2, as reflected by the urinary excretion of the stable hydration products 6-keto-prostaglandin F 1 alpha and thromboxane B2 and the corresponding 2,3-dinor-derivatives, during chronic administration of sulindac (200, 400, 600, and 800 mg/day, each dose given for 7 days in successive weeks) in seven healthy subjects. Urinary eicosanoids were measured by negative ion, chemical ionization-GC/MS-validated RIA techniques. Both 2,3-dinor-thromboxane B2 and 2,3-dinor-6-keto-prostaglandin F 1 alpha showed a dose-dependent reduction, ranging between 45% and 85%. In contrast, the urinary excretion of 6-keto-prostaglandin F1 alpha and thromboxane B2 did not change significantly throughout the study. These results extend previous observations of a selective sparing of renal cyclooxygenase activity by sulindac in humans and demonstrate that this selectivity is not related to an overall weaker enzyme inhibition.

Effects of nimesulide on constitutive and inducible prostanoid biosynthesis in human beings.

BACKGROUND: The aim of this study was to test the hypothesis that nimesulide, a nonsteroidal antiinflammatory drug, or its principal metabolite 4-hydroxynimesulide, is a selective inhibitor of prostaglandin H synthase-2 in human beings. METHODS: Heparinized whole blood samples obtained from healthy subjects were incubated with lipopolysaccharide (10 micrograms/ml) for 24 hours at 37 degrees C and prostaglandin E2 was measured in plasma as an index of monocyte prostaglandin H synthase-2 activity. The production of thromboxane B2 in whole blood allowed to clot at 37 degrees C for 60 minutes was assessed as an index of platelet prostaglandin H synthase-1 activity. We also measured the urinary excretion of 11-dehydrothromboxane B2, prostaglandin E2, 6-ketoprostaglandin F1 alpha, and thromboxane B2 as in vivo indexes of cyclooxygenase activity. All prostanoids were measured by previously validated radioimmunoassay techniques. RESULTS: In the whole blood assays in vitro, nimesulide was twentyfold more potent than 4-hydroxynimesulide toward the two isozymes and both compounds displayed a twentyfold preference for prostaglandin H synthase-2 versus prostaglandin H synthase-1. The administration of a single oral dose of 100 mg nimesulide to six healthy subjects significantly (p < 0.01) reduced monocyte prostaglandin H synthase-2 and prostaglandin H synthase-1 activity ex vivo by more than 90% and 50%, respectively, up to 6 hours. At 24 hours, prostaglandin H synthase-2 but not prostaglandin H synthase-1 activity was significantly reduced by 49% (p < 0.05). Nimesulide significantly (p < 0.05) reduced the urinary excretion of 11-dehydrothromboxane B2 and 6-ketoprostaglandin F1 alpha by approximately 30% and 25%, respectively, while not affecting that of prostaglandin E2 and thromboxane B2. CONCLUSIONS: Nimesulide is a potent inhibitor of human monocyte prostaglandin H synthase-2. However, despite a twentyfold selectivity ratio, therapeutic plasma levels of nimesulide are sufficiently high to cause detectable inhibition of platelet prostaglandin H synthase-1.

Clinical pharmacology of novel selective COX-2 inhibitors.

Novel coxibs (i.e. etoricoxib, valdecoxib, parecoxib and lumiracoxib) with enhanced biochemical cyclooxygenase (COX)-2 selectivity over that of rofecoxib and celecoxib have been recently developed. They have the potential advantage to spare COX-1 activity, thus reducing gastrointestinal toxicity, even when administered at high doses to improve efficacy. They are characterized by different pharmacodynamic and pharmacokinetics features. The higher biochemical selectivity of valdecoxib than celecoxib, evidenced in vitro, may be clinically relevant leading to an improved gastrointestinal safety. Interestingly, parecoxib, a pro-drug of valdecoxib, is the only injectable coxib. Etoricoxib shows only a slightly improved COX-2 selectivity than rofecoxib, a highly selective COX-2 inhibitor that has been reported to halve the incidence of serious gastrointestinal toxicity compared to nonselective nonsteroidal antiinflammatory drugs (NSAIDs). Lumiracoxib, the most selective COX-2 inhibitor in vitro, is the only acidic coxib. The hypothesis that this chemical property may lead to an increased and persistent drug accumulation in inflammatory sites and consequently to an improved clinical efficacy, however, remains to be verified. Several randomized clinical studies suggest that the novel coxibs have comparable efficacy to nonselective NSAIDs in the treatment of osteoarthritis, rheumatoid arthritis and acute pain, but they share similar renal side-effects. The apparent dose-dependence of renal toxicity may limit the use of higher doses of the novel coxibs for improved efficacy. Large-size randomized clinical trials are ongoing to define the gastrointestinal and cardiovascular safety of the novel coxibs.

Equal antiplatelet effects of aspirin 50 or 324 mg/day in patients after acute myocardial infarction.

This study explores the effects on some hematological parameters of a low-dose aspirin regimen (50 mg/day) versus a conventional aspirin treatment with reported antithrombotic efficacy (324 mg/day), in patients with acute myocardial infarction. Fifteen patients were randomized into 3 equal groups receiving 50 mg or 324 mg aspirin or placebo, daily for 21 days. Compared with placebo, bleeding time was significantly and similarly prolonged with both aspirin doses (+ 71 +/- 22% and + 69 +/- 20%, mean +/- S.D.). Aspirin 50 mg/day suppressed arachidonate-induced platelet aggregation and secondary phase aggregation after ADP and adrenaline. Collagen aggregation was inhibited by 44 +/- 15%. In no case were differences in the antiplatelet effects of the two doses observed. The effects of 50 mg/day persisted without attenuation during the observation period. Platelet thromboxane B2 generation during arachidonate-induced aggregation was inhibited by 95 +/- 2 and 99 +/- 1% compared to placebo group after 50 and 324 mg/day, respectively (P between doses less than 0.05). No change was observed with any treatment in coagulation time, prothrombin time or plasma thromboplastin time. Thus, in patients with acute myocardial infarction, the antiplatelet effects of aspirin 50 mg/day are stable over time and superimposable on those of 324 mg/day. The antithrombotic efficacy of aspirin 50 mg/day remains to be tested clinically.

Role of prostaglandins in the renal handling of a salt load in essential hypertension.

Renal function and systemic hemodynamics were assessed in 10 hypertensive patients and in 10 age-matched normotensive subjects during control conditions (80 mEq of sodium/day) and after a salt load, either alone (480 mEq/day) or combined with indomethacin or sulindac. Indomethacin was used to induce ubiquitous inhibition of prostaglandin synthesis and sulindac to inhibit prostaglandin synthesis in all tissues except the kidney. Under control conditions there was no significant difference between the 2 groups in any measurement except blood pressure and total peripheral resistance. Also, the changes induced by salt load in the 2 groups were comparable. However, after indomethacin administration, only hypertensive patients showed a significant reduction in the 24-hour sodium excretion (from 417 +/- 61 to 317 +/- 49 mEq, p less than 0.05), so that the difference between this value and the corresponding value of normotensive subjects (453 +/- 79 mEq) became significant (p less than 0.05). The changes in sodium excretion in hypertensive patients were significantly correlated with the changes in renal plasma flow (r = 0.803, p less than 0.01). However, cardiac output and renal blood flow showed a similar pattern in normal and hypertensive persons. Finally, after the addition of sulindac to salt load, the differences in the 24-hour sodium excretion vanished. These results were also confirmed in an ancillary study performed, using the same protocol, in 10 other hypertensive patients using ibuprofen rather than indomethacin. Our data suggest that renal prostaglandins participate in renal disposal of chronic salt load in hypertensive patients but not in normal persons.

Induction of prostaglandin endoperoxide synthase-2 in human monocytes associated with cyclo-oxygenase-dependent F2-isoprostane formation.

1. The isoprostane 8-epi-prostaglandin (PG)F2 alpha is produced by free radical-catalyzed peroxidation of arachidonic acid. It may also be formed as a minor product of the cyclo-oxygenase activity of platelet PGH synthase (PGHS)-1. We investigated 8-epi-PGF2 alpha production associated with induction of the human monocyte PGHS-2 and its pharmacological modulation. 2. Heparinized whole blood samples were drawn from healthy volunteers, 48 h following oral dosing with aspirin 300 mg to suppress platelet cyclo-oxygenase activity. One ml aliquots were incubated with lipopolysaccharide (LPS: 0.1-50 micrograms ml-1) for 0-24 h at 37 degrees C. PGE2 and 8-epi-PGF2 alpha were measured in separated plasma by radioimmunoassay and enzyme immunoassay techniques. 3. Levels of both eicosanoids were undetectable (i.e. < 60 pg ml-1) at time 0. LPS induced the formation of PGE2 and 8-epi-PGF2 alpha in a time- and concentration-dependent fashion, coincident with the induction of PGHS-2 detected by Western blot analysis of monocyte lysates. After 24 h at 10 micrograms ml-1 LPS, immunoreactive PGE2 and 8-epi-PGF2 alpha averaged 10,480 +/- 4,643 and 295 +/- 140 pg ml-1 (mean +/- s.d., n = 6), respectively. 4. Dexamethasone and 5-methanesulphonamido-6-(2,4-difluorothiophenyl)-1-indano ne (L-745,337), a selective inhibitor of the cyclo-oxygenase activity of PGHS-2, reduced PGE2 and 8-epi-PGF2 alpha production in response to LPS. 5. Isolated monocytes produced PGE2 and 8-epi-PGE2 alpha in response to LPS (10 micrograms ml-1) in a time-dependent fashion. Monocyte PGE2 and 8-epi-PGF2 alpha production was largely prevented by dexamethasone (2 microM) and cycloheximide (10 micrograms ml-1) in association with suppression of PGHS-2 but not of PGHS-1 expression. 6. We conclude that the induction of PGHS-2 in human monocytes is associated with cyclo-oxygenase-dependent generation of the vasoconstrictor and platelet-agonist 8-epi-PGF2 alpha.