Waldemar Radziszewski

Effects of a niacin receptor partial agonist, MK-0354, on plasma free fatty acids, lipids, and cutaneous flushing in humans

BACKGROUND Development of niacin-like agents that favorably affect lipids with an improved flushing profile would be beneficial. OBJECTIVE To evaluate a niacin receptor partial agonist, MK-0354, in Phase I and II studies. METHODS The pharmacokinetic/pharmacodynamic effects of single and multiple doses (7 days) of MK-0354 (300-4000 mg) were evaluated in two Phase I studies conducted in healthy men. A Phase II study assessed the effects of MK-0354 2.5 g once daily on lipids during 4 weeks in 66 dyslipidemic patients. RESULTS MK-0354 single doses up to 4000 mg and multiple doses (7 days) up to 3600 mg produced robust dose-related reductions in free fatty acid (FFA) over 5 hours. Single doses of MK-0354 300 mg and extended release-niacin (Niaspan) 1 g produced comparable reductions in FFA. Suppression of FFA following 7 daily doses of MK-0354 was similar to that after a single dose. In the Phase II study, MK-0354 2.5 g produced little flushing but no clinically meaningful effects on lipids (placebo-adjusted percent change: high-density lipoprotein cholesterol, 0.4%, 95% confidence interval -5.2 to 6.0; low-density lipoprotein cholesterol, -9.8%, 95% confidence interval -16.8 to -2.7; triglyceride, -5.8%, 95% confidence interval -22.6 to 11.9). CONCLUSION Treatment with MK-0354 for 7 days resulted in plasma FFA suppression with minimal cutaneous flushing. However, 4 weeks of treatment with MK-0354 failed to produce changes in high-density lipoprotein cholesterol, low-density lipoprotein cholesterol, or triglycerides.

Diphenylene iodonium, an inhibitor of free radical formation, inhibits platelet aggregation

Diphenylene iodonium is an inhibitor of the enzyme NADPH-oxidase and prevents the generation of oxygen-derived free radicals in neutrophils (Cross and Jones, 1986). Here we show that diphenylene iodonium (0.25-2 microM) inhibited, according to the dose, thrombin-induced platelet-aggregation in human washed platelets and ADP-induced platelet aggregation in platelet-rich plasma. At the concentrations which inhibited platelet aggregation diphenylene iodonium did not alter platelet concentrations of cAMP or cGMP but enhanced the anti-platelet activity of iloprost, sodium nitroprusside or cultured endothelial cells. These findings highlight the importance of free radicals as platelet pro-aggregatory agents.

Effects of extended release niacin/laropiprant, laropiprant, extended release niacin and placebo on platelet aggregation and bleeding time in healthy subjects

Laropiprant (LRPT) has been shown to reduce flushing symptoms induced by niacin and has been combined with niacin for treatment of dyslipidemia. LRPT, a potent PGD2 receptor (DP1) antagonist that also has modest activity at the thromboxane receptor (TP), may have the potential to alter platelet function either by enhancing platelet reactivity through DP1 antagonism or by inhibiting platelet aggregation through TP antagonism. Studies of platelet aggregation ex vivo and bleeding time have shown that LRPT, at therapeutic doses, does not produce clinically meaningful alterations in platelet function. The present study was conducted to assess platelet reactivity to LRPT using methods that increase the sensitivity to detect changes in platelet responsiveness to collagen and ADP. The responsiveness of platelets was quantified by determining the EC50 of collagen to induce platelet aggregation ex vivo. At 24 hours post-dose on Day 7, the responsiveness of platelets to collagen-induced aggregation was similar following daily treatment with extended-release niacin (ERN) 2 g/LRPT 40 mg or ERN 2 g. At 2 hours post-dose on Day 7, the EC50 for collagen-induced platelet aggregation was approximately two-fold higher in the presence of LRPT, consistent with a small, transient inhibition of platelet responsiveness to collagen. There was no clinical difference between treatments for bleeding time, suggesting that this small effect on collagen EC50 does not result in a clinically meaningful alteration of platelet function in vivo. The results of this highly sensitive method demonstrate that LRPT does not enhance platelet reactivity when given alone or with ERN.

NG-hydroxy-L-arginine and hydroxyguanidine potentiate the biological activity of endothelium-derived relaxing factor released from the rabbit aorta

We investigated the effects of NG-hydroxy-L-arginine (L-HOArg) and hydroxyguanidine (HOG) on the synthesis and vasorelaxant activity of endothelium-derived relaxing factor (NO) released from the rabbit aortic endothelium. Both L-HOArg (10 microM) and HOG (10 microM) equally potentiated the vasorelaxant activity of NO released by Ach (0.1 or 0.3 microM) from the luminally perfused rabbit aorta and bioassayed using the superfused strips of the endothelium-denuded rabbit aorta. This potentiation was caused by the generation of a more stable vasodilator during the chemical reaction of L-HOArg or HOG with NO and it was abolished by NG-nitro-L-arginine methyl ester (L-NO2Arg, 10 microM). In contrast, in organ baths, L-HOArg (10 microM) or HOG (10 microM) did not affect the relaxations of intact rabbit aortic rings induced by Ach (0.01-1 microM). At concentrations higher than 10 microM, both L-HOArg and HOG were endothelium-independent vasorelaxants. However, L-HOArg (100 microM) prevented the inhibition by L-NO2Arg (10 microM) of Ach-induced relaxations of bathed aortic rings which indicates that L-HOArg is still a substrate for the NO synthase in the endothelium of the rabbit aorta.

The effects of laropiprant, a selective prostaglandin D₂ receptor 1 antagonist, on the antiplatelet activity of clopidogrel or aspirin.

Laropiprant (LRPT) is being developed in combination with Mercks extended-release niacin (ERN) formulation for the treatment of dyslipidemia. LRPT, an antagonist of the prostaglandin PGD2 receptor DP1, reduces flushing symptoms associated with ERN. LRPT also has affinity for the thromboxane A2 receptor TP (approximately 190-fold less potent at TP compared with DP1). Aspirin and clopidogrel are two frequently used anti-clotting agents with different mechanisms of action. Since LRPT may potentially be co-administered with either one of these agents, these studies were conducted to assess the effects of steady-state LRPT on the antiplatelet activity of steady-state clopidogrel or aspirin. Bleeding time at 24 h post-dose (trough) was pre-specified as the primary pharmacodynamic endpoint in both studies. Two separate, double-blind, randomized, placebo-controlled, crossover studies evaluated the effects of multiple-dose LRPT on the pharmacodynamics of multiple-dose clopidogrel or aspirin. Healthy subjects were randomized to once-daily oral doses of LRPT 40 mg or placebo to LRTP co-administered with clopidogrel 75 mg or aspirin 81 mg for 7 days with at least a 21-day washout between treatments. In both studies, bleeding time and platelet aggregation were assessed 4 and 24 hours post-dose on Day 7. Comparability was declared if the 90% confidence interval for the estimated geometric mean ratio ([LRPT+clopidogrel]/clopidogrel alone or [LRPT+aspirin]/aspirin alone) for bleeding time at 24 hours post-dose on Day 7 was contained within (0.66, 1.50). Concomitant daily administration of LRPT 40 mg with clopidogrel 75 mg or aspirin 81 mg resulted in an approximate 4–5% increase in bleeding time at 24 hours after the last dose vs. bleeding time after treatment with clopidogrel or aspirin alone, demonstrating that the treatments had comparable effects on bleeding time. Percent inhibition of platelet aggregation was not significantly different between LRPT co-administered with clopidogrel or aspirin vs. clopidogrel or aspirin alone at 24 hours post-dose at steady state. At 4 hours after the last dose, co-administration of LRPT 40 mg resulted in 3% and 41% increase in bleeding time vs. bleeding time after treatment with aspirin or clopidogrel alone, respectively. Co-administration of LPRT with clopidogrel or aspirin was generally well tolerated in healthy subjects. Co-administration of multiple doses of LRPT 40 mg and clopidogrel 75 mg or aspirin 81 mg had no clinically important effects on bleeding time or platelet aggregation.

Histological spectrum of cutaneous reactions to aspirin in chronic idiopathic urticaria

Background:  During a clinical trial, we obtained 16 biopsies of skin eruptions induced by aspirin in patients with chronic idiopathic urticaria (CIU). In this setting, aspirin triggers skin eruptions through a well‐established non‐immunological mechanism involving the inhibition of cyclooxygenase type I. This presented the rare opportunity to evaluate histological features of a series of skin eruptions induced by a drug acting through a defined mechanism in a controlled experimantal setting.

Plasma ANP and Cyclic GMP Levels Versus Left Ventricular Performance at Different AV Delays in AV Sequential Pacing

Eleven resting patients with an implanted DDD pacemaker were studied. After 30 minutes of AV sequentiai pacing at a rate of 80 beats/min with three consecutive atrioventricular delays (AVDs; 100, 150, and 200 msec) peripheral venous blood was drawn for further analyses by specific radioimmunoassays of atrial natriuretic peptide (ANP) and the ANP second messenger, cyclic guanosine monophosphate (cGMP). Relative changes in left ventricular (LV) stroke volume following alterations of AVD were assessed by means of pulsed‐Doppler echocardiography through measurement of LV outflow time‐velocity integrals (TVI). The optimal AVD (oA VD) was defined in individual patients as that which was associated with the greatest TVI and with improvement over both other AVDs of more than 4%. The oA VD was found in nine patients. For these nine patients no significant differences in either plasma ANP or cGMP between various AVDs were observed. However, we found such differences with respect to values measured at oAVD; both ANP and cGMP levels were lowest at oAVD. Pooling together the data obtained in 11 patients at three AVDs, a positive correlation between ANP and cGMP levels was found (r = 0.7, P < 0.0001. n = 33). Moreover, changes of plasma ANP and cGMP induced by every A VD increment of 50 msec were also correlated (r = 0.6, P < 0.01, n = 22). It is concluded that in AV sequential pacing at rest piasma ANP reaches minimal levels at the AVD, which provides the best LV performance. Although levels of cGMP changed in parallel with those of ANP, low relative values of cGMP differences may limit the usefulness of cGMP assays in optimization of the AVD.

Development and validation of a LC/MS/MS method for 6-keto PGF1α, a metabolite of prostacyclin (PGI2)

A sensitive liquid chromatography/tandem mass spectrometry (LC/MS/MS) method was developed for the quantitation of 6-keto PGF(1α) in human urine and plasma. Prostacyclin (PGI(2)) is a locally acting prostanoid, which mediates vasorelaxation and inhibition of platelet aggregation. 6-Keto PGF(1α) is the most-immediate metabolite of PGI(2). Samples were spiked with an internal standard (6-keto PGF(1α)-d(4)), purified by immuno-affinity chromatography and selected reaction monitoring (SRM) was performed. Analytical validation of the 6-keto PGF(1α) assay was performed in urine. This included an assessment of assay precision, recovery, stability, sensitivity and linearity. Urinary 6-keto PGF(1α) concentrations were also correlated to urinary 2,3-dinor-6-keto PGF(1α) (PGIM) concentrations using urine samples collected from 16 healthy volunteers. The mean concentration of 6-keto PGF(1α) in urine (mean ± SD) was 92 ± 51 pg/ml or 168 ± 91 pg/mg creatinine. Overall, there was a statistically significant correlation between urinary 6-keto PGF(1α) and PGIM (r(2)=0.55, p ≤ 0.001; slope=2.7; y-intercept=130). However, PGIM was approximately 3-fold more abundant than 6-keto PGF(1α) in urine. In addition, 6-keto PGF(1α) concentrations were measured in EDTA plasma samples obtained from 7 healthy donors. The mean concentration of 6-keto PGF(1α) in plasma was 1.9 ± 0.8 pg/ml (± SD).

The effects of laropiprant on the antiplatelet activity of co-administered clopidogrel and aspirin

Abstract Laropiprant is an antagonist of the prostaglandin PGD2 receptor DP1. Laropiprant has a weak affinity for the thromboxane A2 receptor TP. Two double-blinded, randomized, placebo-controlled, crossover studies evaluated the effects of multiple-dose laropiprant at steady state on the antiplatelet effects of multiple-dose aspirin and clopidogrel. Study 1 had two treatment periods, in which each healthy subject received laropiprant 40 mg, clopidogrel 75 mg, and aspirin 80 mg (Treatment A), or placebo, clopidogrel 75 mg, and aspirin 80 mg (Treatment B) once daily for 7 days. Study 2 consisted of three treatment periods. In the first two, each patient with hypercholesterolemia or mixed dyslipidemia received laropiprant 40 mg, clopidogrel 75 mg, and aspirin 81 mg (Treatment A), or placebo, clopidogrel 75 mg, and aspirin 81 mg (Treatment B) once daily for 7 days. In period 3, patients received a single dose of two tablets of extended release nicotinic acid 1 g/laropiprant 20 mg (Treatment C). In both studies, pharmacodynamic endpoints included bleeding time at 24 (primary) and 4 hours (secondary) post-dose following 7 days of once-daily laropiprant in combination with clopidogrel and aspirin, and platelet aggregation in platelet-rich plasma at 4 and 24 hours post-dose on day 7 (secondary). After 7 days, increased bleeding time of 27% (Study 1) and 23% (Study 2) at 24 hours post-dose was observed with laropiprant compared to placebo (both combined with clopidogrel and aspirin), with corresponding upper bounds of the 90% CI marginally exceeding the prespecified upper comparability bound of 1.50 in both studies. The GMR and 90% CI for bleeding time of laropiprant compared to placebo (both combined with clopidogrel and aspirin) at 4 hours post-dose on day 7 was 0.92 (0.70, 1.21) in Study 1, and 1.46 (1.20, 1.78) in Study 2. Compared with placebo, laropiprant (both combined with clopidogrel and aspirin) increased the inhibition of collagen- and ADP-induced platelet aggregation, respectively, by ∼2.4% and ∼8.1% in Study 1 and by ∼4% and ∼5.4% in Study 2, at 24 hours post-dose on day 7. The inhibition of collagen- and ADP-induced platelet aggregation, respectively, was increased by ∼0.1% and ∼5.0% in Study 1, and by ∼5% and ∼12% in Study 2, at 4 hours post-dose on day 7. In conclusion, co-administration of multiple doses of laropiprant with aspirin and clopidogrel induced a prolongation of bleeding time and an inhibitory effect on platelet aggregation ex vivo in healthy subjects and patients with dyslipidemia.

Plasma ANP and cyclic GMP after physical exercise in patients with mitral valve disease and in healthy subjects

Plasma levels of both atrial natriuretic peptide (ANP) and cyclic GMP are elevated in patients with various heart diseases as compared to healthy subjects. In this study patients with advanced mitral valve disease (Group A) and healthy subjects (Group B) were exposed to symptom-limited upright stepwise physical exercise on a cycle ergometer. Concentrations of ANP and cyclic GMP were measured in plasma at rest (20 min in supine position) or 5 min after physical exercise by specific radioimmunoassays. Here we show that short dynamic exercise caused a significant increase in plasma levels of ANP and cyclic GMP, in both groups. In Group A strong correlation between plasma ANP and cyclic GMP was found at rest (r = 0.91, P < 0.001, n = 11) and after physical exercise (r = 0.85, P < 0.001, n = 11). In contrast, there was no correlation between plasma concentrations of ANP and cyclic GMP in Group B at rest (r = -0.16, P > 0.05, n = 10) or after exercise loading (r = 0.14, P > 0.05, n = 10). Absolute increases in circulating levels of both substances were not found to correlate in either group. These data suggest that exercise-induced elevations in plasma cyclic GMP may be due not only to ANP release but also to an as yet undetermined factor, possibly EDRF/NO.