Marta L. Capone

Clinical Pharmacology of Platelet, Monocyte, and Vascular Cyclooxygenase Inhibition by Naproxen and Low-Dose Aspirin in Healthy Subjects

Background—The current controversy on the potential cardioprotective effect of naproxen prompted us to evaluate the extent and duration of platelet, monocyte, and vascular cyclooxygenase (COX) inhibition by naproxen compared with low-dose aspirin. Methods and Results—We performed a crossover, open-label study of low-dose aspirin (100 mg/d) or naproxen (500 mg BID) administered to 9 healthy subjects for 6 days. The effects on thromboxane (TX) and prostacyclin biosynthesis were assessed up to 24 hours after oral dosing. Serum TXB2, plasma prostaglandin (PG) E2, and urinary 11-dehydro-TXB2 and 2,3-dinor-6-keto-PGF1&agr; were measured by previously validated radioimmunoassays. The administration of naproxen or aspirin caused a similar suppression of whole-blood TXB2 production, an index of platelet COX-1 activity ex vivo, by 94±3% and 99±0.3% (mean±SD), respectively, and of the urinary excretion of 11-dehydro-TXB2, an index of systemic biosynthesis of TXA2 in vivo, by 85±8% and 78±7%, respectively, that persisted throughout the dosing interval. Naproxen, in contrast to aspirin, significantly reduced systemic prostacyclin biosynthesis by 77±19%, consistent with differential inhibition of monocyte COX-2 activity measured ex vivo. Conclusions—The regular administration of naproxen 500 mg BID can mimic the antiplatelet COX-1 effect of low-dose aspirin. Naproxen, unlike aspirin, decreased prostacyclin biosynthesis in vivo.

Increased Oxidative Stress and Platelet Activation in Patients With Hypertension and Renovascular Disease

Background—Hypertensive patients with renovascular disease (RVD) may be exposed to increased oxidative stress, possibly related to activation of the renin-angiotensin system. Methods and Results—We measured the urinary excretion of 8-iso-prostaglandin (PG) F2&agr; and 11-dehydro-thromboxane (TX) B2 as indexes of in vivo lipid peroxidation and platelet activation, respectively, in 25 patients with RVD, 25 patients with essential hypertension, and 25 healthy subjects. Plasma renin activity in peripheral and renal veins, angiotensin II in renal veins, cholesterol, glucose, triglycerides, homocysteine, and antioxidant vitamins A, C, and E were also determined. Patients were also studied 6 months after a technically successful angioplasty of the stenotic renal arteries. Urinary 8-iso-PGF2&agr; was significantly higher in patients with RVD (median, 305 pg/mg creatinine; range, 124 to 1224 pg/mg creatinine) than in patients with essential hypertension (median, 176 pg/mg creatinine; range, 48 to 384 pg/mg creatinine) or in healthy subjects (median, 123 pg/mg creatinine; range, 58 to 385 pg/mg creatinine). Urinary 11-dehydro-TXB2 was also significantly higher in RVD patients compared with healthy subjects. In RVD patients , urinary 8-iso-PGF2&agr; correlated with 11-dehydro-TXB2 (rs=0.48;P <0.05) and renal vein renin (rs=0.67;P <0.005) and angiotensin II (rs=0.65;P =0.005) ratios. A reduction in 8-iso-PGF2&agr; after angioplasty was observed in RVD patients with high baseline levels of lipid peroxidation. Changes in 8-iso-PGF2&agr; were related to baseline lipid peroxidation (rs=−0.73;P <0.001), renal vein angiotensin II (rs=−0.70;P <0.01) and renin (rs=−0.63;P <0.05) ratios. Conclusions—Lipid peroxidation is markedly enhanced in hypertensive patients with RVD and is related to activation of the renin-angiotensin system. Moreover, persistent platelet activation triggered or amplified by bioactive isoprostanes may contribute to the progression of cardiovascular and renal damage in this setting.

Celecoxib, ibuprofen, and the antiplatelet effect of aspirin in patients with osteoarthritis and ischemic heart disease.

We performed a placebo‐controlled, randomized study to address whether celecoxib or ibuprofen undermines the functional range of inhibition of platelet cyclooxygenase (COX)–1 activity by aspirin in patients with osteoarthritis and stable ischemic heart disease.

Determinants of Platelet Activation in Human Essential Hypertension

Abstract—Experimental data suggest that oxidative stress might be enhanced in hypertension and contribute to platelet activation. We hypothesized that both oxidative stress and platelet activation could be related to the clinical characteristics of hypertensive patients. The urinary excretion of 11-dehydrothromboxane (TX) B2, reflecting in vivo platelet activation, was measured in 75 patients with mild to severe essential hypertension and 75 pair-matched, healthy controls. The urinary excretion of 8-iso-prostaglandin (PG) F2&agr; was determined as an index of in vivo lipid peroxidation. Urinary 11-dehydro-TXB2 was significantly higher in essential hypertensives compared with controls. Although no statistically significant difference in urinary 8-iso-PGF2&agr; was observed between patients and controls, plasma vitamin C was lower and plasma homocysteine higher in hypertensive patients than in controls. Both urinary 11-dehydro-TXB2 and 8-iso-PGF2&agr; were higher in patients with advanced hypertensive retinopathy compared with patients without retinopathy. Multivariate linear regression analysis identified urinary 8-iso-PGF2&agr;, plasma fibrinogen, homocysteine, and vitamin E as the only variables independently correlated with urinary 11-dehydro-TXB2. Logistic regression analysis showed that high urinary 8-iso-PGF2&agr;, plasma fibrinogen, and homocysteine, as well as low plasma vitamin E, advanced retinopathy, elevated diastolic blood pressure, and the absence of antihypertensive treatment, were predictors of high urinary 11-dehydro-TXB2. We demonstrated increased oxidative stress and persistent platelet activation in essential hypertensives with advanced vascular lesions. These findings might help identify hypertensive patients who are at increased risk of cardiovascular events and who might benefit from long-term antiplatelet therapy.

The biochemical selectivity of novel COX-2 inhibitors in whole blood assays of COX-isozyme activity.

Summary We have evaluated the biochemical selectivity of novel cyclo-oxygenase (COX)-2 inhibitors, etoricoxib, valdecoxib, DFU and DFP, vs rofecoxib and celecoxib, using the human whole blood assays of COX-isozyme activity, in vitro. Compounds were incubated with human whole blood samples, allowed to clot for 1 h at 37°C, or stimulated with lipopolysaccharide (10|ig/ml) for 24 h at 37°C. Serum thromboxane (TX) B2 and plasma prostaglandin (PG) E2 levels were measured by specific radioimmunoassays as indices of platelet COX-1 and monocyte COX-2 activity, respectively. Valdecoxib, etoricoxib, DFU and DFP inhibited platelet COX-1 and monocyte COX-2 with the following COX-1/COX-2 IC50 ratios: 61.5, 344, 660 and 1918, respectively. The reference compounds, celecoxib and rofecoxib had corresponding values of 29.6 and 272. In conclusion, a second wave of COX-2 inhibitors with higher biochemical selectivity than the existing coxibs has been developed. Whether their administration will be associated with improved clinical efficacy and/or safety visà-vis celecoxib and rofecoxib remains to be established.

Human Pharmacology of Naproxen Sodium

We compared the variability in degree and recovery from steady-state inhibition of cyclooxygenase (COX)-1 and COX-2 ex vivo and in vivo and platelet aggregation by naproxen sodium at 220 versus 440 mg b.i.d. and low-dose aspirin in healthy subjects. Six healthy subjects received consecutively naproxen sodium (220 and 440 mg b.i.d.) and aspirin (100 mg daily) for 6 days, separated by washout periods of 2 weeks. COX-1 and COX-2 inhibition was determined using ex vivo and in vivo indices of enzymatic activity: 1) the measurement of serum thromboxane (TX)B2 levels and whole-blood lipopolysaccharide-stimulated prostaglandin (PG)E2 levels, markers of COX-1 in platelets and COX-2 in monocytes, respectively; 2) the measurement of urinary 11-dehydro-TXB2 and 2,3-dinor-6-keto-PGF1α levels, markers of systemic TXA2 biosynthesis (mostly COX-1-derived) and prostacyclin biosynthesis (mostly COX-2-derived), respectively. Arachidonic acid (AA)-induced platelet aggregation was also studied. The maximal inhibition of platelet COX-1 (95.9 ± 5.1 and 99.2 ± 0.4%) and AA-induced platelet aggregation (92 ± 3.5 and 93.7 ± 1.5%) obtained at 2 h after dosing with naproxen sodium at 220 and 440 mg b.i.d., respectively, was indistinguishable from aspirin, but at 12 and 24 h after dosing, we detected marked variability, which was higher with naproxen sodium at 220 mg than at 440 mg b.i.d. Assessment of the ratio of inhibition of urinary 11-dehydro-TXB2 versus 2,3-dinor-6-keto-PGF1α showed that the treatments caused a more profound inhibition of TXA2 than prostacyclin biosynthesis in vivo throughout dosing interval. In conclusion, neither of the two naproxen doses mimed the persistent and complete inhibition of platelet COX-1 activity obtained by aspirin, but marked heterogeneity was mitigated by the higher dose of the drug.

Low-Dose Naproxen Interferes With the Antiplatelet Effects of Aspirin in Healthy Subjects: Recommendations to Minimize the Functional Consequences

OBJECTIVE To investigate whether low-dose naproxen sodium (220 mg twice a day) interferes with aspirins antiplatelet effect in healthy subjects. METHODS We performed a crossover, open-label study in 9 healthy volunteers. They received for 6 days 3 different treatments separated by 14 days of washout: 1) naproxen 2 hours before aspirin, 2) aspirin 2 hours before naproxen, and 3) aspirin alone. The primary end point was the assessment of serum thromboxane B(2) (TXB(2)) 24 hours after the administration of naproxen 2 hours before aspirin on day 6 of treatment. In 5 volunteers, the rate of recovery of TXB(2) generation (up to 72 hours after drug discontinuation) was assessed in serum and in platelet-rich plasma stimulated with arachidonic acid (AA) or collagen. RESULTS Twenty-four hours after the last dosing on day 6 in volunteers receiving aspirin alone or aspirin before naproxen, serum TXB(2) was almost completely inhibited (median [range] 99.1% [97.4-99.4%] and 99.1% [98.0-99.7%], respectively). Naproxen given before aspirin caused a slightly lower inhibition of serum TXB(2) (median [range] 98.0% [90.6-99.4%]) than aspirin alone (P = 0.0007) or aspirin before naproxen (P = 0.0045). All treatments produced a maximal inhibition of AA-induced platelet aggregation. At 24 hours, compared with baseline, collagen-induced platelet aggregation was still inhibited by aspirin alone (P = 0.0003), but not by aspirin given 2 hours before or after naproxen. Compared with administration of aspirin alone, the sequential administration of naproxen and aspirin caused a significant parallel upward shift of the regression lines describing the recovery of platelet TXB(2). CONCLUSION Sequential administration of 220 mg naproxen twice a day and low-dose aspirin interferes with the irreversible inhibition of platelet cyclooxygenase 1 afforded by aspirin. The interaction was smaller when giving naproxen 2 hours after aspirin. The clinical consequences of these 2 schedules of administration of aspirin with naproxen remain to be studied in randomized clinical trials.

Clinical pharmacology of etoricoxib: a novel selective COX2 inhibitor.

The development of COX2 inhibitors with improved biochemical selectivity (such as etoricoxib and valdecoxib) over that of commercially available coxibs has been driven by the potential advantage of safety using higher coxib doses for increased efficacy. Etoricoxib has been approved in the UK as a once-daily medicine for symptomatic relief in the treatment of osteoarthritis (OA), rheumatoid arthritis (RA) and acute gouty arthritis. It is currently approved with additional indications (i.e., for relief of acute pain associated with dental surgery, for primary dysmenorrhoea and for chronic musculo-skeletal pain, including chronic lower-back pain) in Mexico, Brazil and Peru. Etoricoxib has an in vitro COX1/COX2 IC50 ratio of 344, the highest of any coxib. The administration of therapeutic doses of etoricoxib to healthy subjects does not affect COX1 activity in circulating platelets and gastric biopsies. The profound inhibition of monocyte COX2 activity at 24 h after dosing, as predicted by a pharmacological half-life of ~ 22 h, supports a once-daily dosing regimen of etoricoxib. In randomised, well-controlled clinical trials, etoricoxib has been shown to have a comparable clinical efficacy with traditional NSAIDs. Combined analysis of efficacy trials with etoricoxib versus non-selective NSAIDs has shown that the drug halves both investigator-reported upper gastrointestinal perforation, ulcers and bleeds (PUBs) and confirmed PUBs, and reduces the need for gastroprotective agents and gastrointestinal comedications by ~ 40%. The risk of lower extremity oedema and hypertension adverse experiences with etoricoxib was low and generally similar to comparator NSAIDs in a combined analysis of eight Phase III studies in OA, RA, chronic low-back pain and surveillance endoscopy. Large, randomised clinical trials have been planned to confirm the renal, gastrointestinal and cardiovascular safety of etoricoxib.

Glucose and collagen regulate human platelet activity through aldose reductase induction of thromboxane

Diabetes mellitus is associated with platelet hyperactivity, which leads to increased morbidity and mortality from cardiovascular disease. This is coupled with enhanced levels of thromboxane (TX), an eicosanoid that facilitates platelet aggregation. Although intensely studied, the mechanism underlying the relationship among hyperglycemia, TX generation, and platelet hyperactivity remains unclear. We sought to identify key signaling components that connect high levels of glucose to TX generation and to examine their clinical relevance. In human platelets, aldose reductase synergistically modulated platelet response to both hyperglycemia and collagen exposure through a pathway involving ROS/PLCγ2/PKC/p38α MAPK. In clinical patients with platelet activation (deep vein thrombosis; saphenous vein graft occlusion after coronary bypass surgery), and particularly those with diabetes, urinary levels of a major enzymatic metabolite of TX (11-dehydro-TXB2 [TX-M]) were substantially increased. Elevated TX-M persisted in diabetic patients taking low-dose aspirin (acetylsalicylic acid, ASA), suggesting that such patients may have underlying endothelial damage, collagen exposure, and thrombovascular disease. Thus, our study has identified multiple potential signaling targets for designing combination chemotherapies that could inhibit the synergistic activation of platelets by hyperglycemia and collagen exposure.

NSAIDs and cardiovascular disease: transducing human pharmacology results into clinical read-outs in the general population

Traditional (t) non-steroidal anti-inflammatory drugs (NSAIDs) and selective cyclooxygenase (COX)-2 inhibitors (coxibs) are important and efficacious drugs for the management of musculoskeletal symptoms. These drugs have both beneficial and adverse effects due to the inhibition of prostanoids. Although the tNSAID and coxib inhibition of COX-2-dependent prostaglandin (PG)E(2) production is effective in ameliorating symptoms of inflammation and pain, a small but consistent increased risk of myocardial infarction has been detected in association with their use. Convincing evidence suggests that cardiovascular toxicity associated with the administration of these compounds occurs through a common mechanism involving inhibition of COX-2-dependent prostacyclin. The development of biomarkers that predict the impact of NSAIDs on COX-1 and COX-2 activities in vitro, ex vivo and in vivo has been essential to read-out the clinical consequences of the varying degrees of inhibition of the two COX-isozymes in humans. Whole blood assays for COX-1 and COX-2 might be candidates as surrogate end-points of toxicity and efficacy of NSAIDs. Using a biomarker strategy, we have shown that the degree of inhibition of COX-2 and the functional selectivity with which it is achieved are relevant to the level of cardiovascular hazard from NSAIDs and relate to drug potency (exposure). We propose that the assessment of COX-2 in whole blood ex vivo, either alone or in combination with urinary levels of 2,3-dinor-6-keto-PGF(1 alpha) a biomarker of prostacyclin biosynthesis in vivo, may represent a valid surrogate end-point to predict cardiovascular risk for functionally selective COX-2 inhibitors.