2007 - 11

Dig Liver Dis. 2007 Nov 6; [Epub ahead of print]

NSAIDs, coxibs, CINOD and H(2)S-releasing NSAIDs: What lies beyond the horizon.

Fiorucci S, Santucci L, Distrutti E. Dipartimento di Medicina Clinica e Sperimentale, Universita di Perugia, Via E. dal Pozzo, 06122 Perugia, Italy.

Nonsteroidal anti-inflammatory drugs are widely prescribed for treatment of pain and inflammation, despite their association with gastrointestinal complications, including bleeding and perforation. Inhibition of cyclo-oxygenases, is the main mechanism of action of aspirin and nonsteroidal anti-inflammatory drugs. Non-selective nonsteroidal anti-inflammatory drugs inhibit cyclo-oxygenase-1 and cyclo-oxygenase-2. Inhibition of cyclo-oxygenase-1 derived prostanoids in the stomach represent the underlying mechanism involved in development of gastric and duodenal ulcers in patients taking nonsteroidal anti-inflammatory drugs. Selective cyclo-oxygenases-2 inhibitor (coxibs) spare cyclo-oxygenase-1 show enhanced safety profile in the gastrointestinal tract, but increase the risk of heart attack and stroke. Spurred by these findings, two coxibs, rofecoxib and valdecoxib, were withdrawn from the market. In addition to prostanoids, two gaseous mediators, nitric oxide (NO) and hydrogen sulfide (H(2)S) exert protective effects in gastric mucosa. The inhibitory effects of NO on nonsteroidal anti-inflammatory drugs-induced leukocyte adherence have been exploited in the development of NO-releasing nonsteroidal anti-inflammatory drugs, also indicated as cyclo-oxygenase-inhibiting NO-donating drugs. Despite its non-selective profile versus cyclo-oxygenase isoenzymes, naprocyclo-oxygenase-inhibiting NO-donating drugs, the prototype of this class of anti-inflammatory agents, reduces systemic blood pressure and might have enhanced cardiovascular safety than coxibs, while causing less gastrointestinal damage than its parent drug, the naproxen. H(2)S-releasing nonsteroidal anti-inflammatory drugs derivatives have been recently developed, based on the observed ability of this gaseous mediator to cause vasodilation and to prevent leukocyte adherence. In pre-clinical settings, H(2)S-releasing nonsteroidal anti-inflammatory drugs produce less gastric damage as compared to the parent drugs. Cyclo-oxygenases-inhibiting NO-donating drugs and H(2)S-releasing nonsteroidal anti-inflammatory drugs represent examples of new anti-inflammatory drugs created through the exploitation of the beneficial effects of endogenous gaseous mediators in the gastrointestinal and cardiovascular systems.

2007 - 08

Drugs R D. 2007;8(4):255-8.

Naproxcinod: AZD 3582, HCT 3012, naproxen nitroxybutylester, nitronaproxen, NO-naproxen.

[No authors listed]

Naproxcinod is a derivative of naproxen with similar anti-inflammatory activity, but less gastrointestinal toxicity. It is the first of a new class of analgesic and anti-inflammatory drugs known as cyclo-oxygenase-(COX)-inhibiting nitric oxide donators (CINODs) under development with NicOx in several countries. Naproxcinod is in phase III clinical development in the US for the treatment of osteoarthritis.The improved gastrointestinal tolerability of naproxcinod appears to be due to its release of nitric oxide (NO) and the consequent maintenance of tissue perfusion and integrity.Naproxcinod is available for licensing.AstraZeneca had been a worldwide licensee for naproxcinod and other CINODs. However, the results of phase II clinical trials of naproxcinod did not fulfill AstraZeneca's strategic commercial criteria for further investment and NicOx reacquired rights following AstraZeneca's decision to discontinue its involvement in 2003. NicOx was surprised by AstraZeneca's decision, and remained fully convinced of the potential of naproxcinod. NicOx is seeking new partners for development of compounds of the CINOD class.The second phase III trial (302 study) for naproxcinod in patients with osteoarthritis of the knee was initiated in April 2007. The trial will enrol approximately 1020 patients from 120 clinical sites in the US. The study is designed to confirm naproxcinod's efficacy and provide additional blood pressure data. Efficacy results are expected in mid-2008. The third phase III trial (303 study) is scheduled to start in the first half of 2007 and will assess the efficacy and safety in patients with osteoarthritis (OA) of the hip. Following pooled analysis of all three phase III trials, NicOx hopes to file an NDA in the US during the first quarter of 2009.NicOx is in ongoing discussions with regulatory authorities worldwide regarding the safety of naproxcinod and its COX-inhibiting properties. The company submitted documentation to the US FDA in August 2006 that outlined the long-term safety assessments planned for naproxcinod and has requested scientific advice on naproxcinod from the EMEA in Europe. Pending successful outcomes of the three phase III trials in patients with OA, the company anticipates that regulatory submissions in the US and Europe could be made during Q1 2009. NicOx is planning to validate a proposed development plan for Japan with the Japanese authorities in Q1 2007.Another monitoring trial is being planned by NicOx: the 305 study is a clinical endoscopy study whose objective is to confirm previous clinical findings that treatment with naproxcinod results in less gastrointestinal damage than naproxen. The trial is expected to start in 2007.The STAR Multinational Study Group has conducted a phase II gastrointestinal safety and efficacy study of naproxcinod versus naproxen in 970 patients with osteoarthritis at 80 sites in Argentina, Brazil, Hungary, Mexico, Norway, Poland, South Africa and the UK. The study was completed in November 2002.Previously, AstraZeneca had conducted a randomised, phase II trial evaluating naproxcinod's efficacy and safety among 672 subjects with symptomatic knee osteoarthritis.

Inflammopharmacology. 2007 Jun;15(3):119-23.

The effects of AZD3582 [4-(nitroxy)butyl-(2S)-2-(6-methoxy-2-naphthyl) propanoate], and naproxen on key pathogenic steps in NSAID-enteropathy in the rat.

Walley M, Sigthorsson G, Hotz-Behofsits C, Simpson R, Bjarnason I. Guy's, King's and St Thomas' School of Medicine, Department of Gastroenterology, King's College Hospital Foundation Trust, Denmark Hill, London SE5 9PJ, London,UK. BACKGROUND: The pathogenesis of NSAID-induced enteropathy may involve dual inhibition of the cyclooxygenase (1 and 2) and a topical effect with sequential increased intestinal permeability, development of inflammation and ulcers. It has been suggested that nitric-oxide donating drugs cause significantly less gastrointestinal injury by counteracting for NSAID-induced reductions in blood flow. AIMS: To compare the effects of AZD3582 [4-(nitroxy)butyl-(2S)-2-(6-methoxy-2-naphthyl) propanoate], and naproxen on key pathogenic steps in NSAID-enteropathy in the rat. METHODS: Single doses of AZD3582, naproxen (dose range 10-300 micromol/kg) or vehicle were given to male Sprague Dawley rats. Intestinal permeability (51CrEDTA) and intestinal inflammation (granulocyte marker protein) was quantitated and ulcer counts made. RESULTS: Intestinal permeability (all doses) and inflammation (highest dose of the drugs) increased significantly from control levels following naproxen and AZD3582 and there was no significant difference between the drugs. Median ulcer counts were, however, significantly (p <>

2006 - 09

Clin Ther. 2006 Sep;28(9):1279-95.

Analgesic efficacy of the cyclooxygenase-inhibiting nitric oxide donor AZD3582 in postoperative dental pain: Comparison with naproxen and rofecoxib in two randomized, double-blind, placebo-controlled studies.

Michael Hill C, Sindet-Pederson S, Seymour RA, Hawkesford JE 2nd, Coulthard P, Lamey PJ, Gerry Cowan C, Wickens M, Jeppsson L, Dean AD, Svensson O. University Dental Hospital, Heath Park, Cardiff, United Kingdom.

OBJECTIVE: This study assessed the analgesic efficacy of single doses of 4-(nitrooxy)butyl-(2S)-2-(6-methoxy-2-naphthyl) propanoate (AZD3582) in acute postoperative dental pain after the removal of an impacted mandibular third molar (ie, wisdom tooth). METHODS: Two randomized, placebo-controlled, double-blind studies were performed. In a dose-finding study, 242 patients were randomized to AZD3582 375, 750, 1500, or 2250 mg (n = 41, 37, 42, and 41, respectively); naproxen 500 mg (n = 39); or placebo (n = 42). In a comparator study, 282 patients were randomized to AZD3582 500 mg (n = 78) or 750 mg (n = 83), rofecoxib 50 mg (n = 80), or placebo (n = 41). Primary outcomes included time to rescue medication, time to pain relief, and mean pain intensity difference (MPID), as well as safety profile. Pain was rated on a visual analog scale. RESULTS: In the dose-finding study, 52% (126/242) were women; the mean (SD) age was 25.1 (4) years, mean weight was 69.0 kg, and the mean (SD) body mass index (BMI) was 23.7 (3) kg/m2. In the comparator study, 58% (164/282) were women; the mean (SD) age was 27 (6.4) years, mean weight was 71 kg, and mean (SD) BMI was 24.2 (3) kg/m2. In the dose-finding study, the AZD3582 750-, 1500-, and 2250-mg groups were superior to placebo in the primary variables "time to rescue medication (0-8 hours)" (hazard ratios [HRs] [95% CIs], 0.17 [0.07-0.42], P < p =" NS," p =" 0.003," p =" 0.02)," p =" 0.002)." p =" NS"> or =750 mg. The proportions of patients with vertigo and decreased orthostatic blood pressure each group were as follows: AZD3582 500 mg, 6%; AZD3582 750 mg, 12%; rofecoxib, 3%; and placebo, 5%. CONCLUSIONS: AZD3582 750 mg had similar analgesic efficacy as equimolar doses of naproxen, but noninferiority to rofecoxib was not demonstrated.

2006 - 07

Drugs R D. 2006;7(4):262-6.

Nitronaproxen: AZD 3582, HCT 3012, Naproxen Nitroxybutylester, NO-Naproxen.

[No authors listed]

Nitronaproxen [AZD 3582, HCT 3012, naproxen nitroxybutylester, NO-naproxen] is a naproxen derivative with similar anti-inflammatory activity to the parent compound, but with less gastrointestinal toxicity. It is the first of a new class of analgesic and anti-inflammatory drugs known as cyclo-oxygenase-inhibiting nitric oxide donators (CINODs), which are under development by NicOx. The better gastrointestinal tolerability of nitronaproxen appears to be due to its release of nitric oxide (NO) and the consequent maintenance of tissue perfusion and integrity.Nitronaproxen is in phase III clinical development for the treatment of osteoarthritis and is available for licensing.AstraZeneca had been a worldwide licensee for nitronaproxen and other CINODs. However, the results of phase II clinical trials of nitronaproxen did not fulfill AstraZeneca's strategic commercial criteria for further investment and NicOx reacquired rights following AstraZeneca's decision to discontinue its involvement in 2003. NicOx was surprised by AstraZeneca's decision, and remained fully convinced of the potential of nitronaproxen. NicOx is seeking new partners for development of compounds of the CINOD class.Nitronaproxen is in a phase III clinical trial for the treatment of osteoarthritis (OA) of the knee. The 13-week trial completed enrolment of 820 patients from 120 clinical sites in the US in May 2006. The study is designed to confirm that nitronaproxen is superior to placebo and is as effective as naproxen in relieving signs and symptoms of OA. The study will also seek to show that nitronaproxen has no adverse effect on blood pressure. An additional trial has begun that is employing ambulatory blood pressure monitoring to provide a description of the blood pressure effect of nitronaproxen over a 24-hour period in hypertensive subjects. This US trial will enrol approximately 120 volunteers with stable essential hypertension. The volunteers will not have osteoarthritis but will be between the ages of 50 and 75 years (representative of the osteoarthritis population). Results from both trials are expected in the fourth quarter of 2006.The phase II clinical programme for nitronaproxen, which included 2709 patients in five separate clinical studies, showed that the drug is a potent, safe anti-inflammatory agent, with potential for improved cardiovascular safety over NSAIDs and COX-2 selective NSAIDs. An independent advisory board recommended further development of nitronaproxen in the treatment of osteoarthritis in 2004 based on an evaluation of the full results of the phase II clinical programme.A clinical study had begun in September 2004 at the University of Pennsylvania in patients with mild essential hypertension, in which the effects of nitronaproxen and rofecoxib on arterial blood pressure would be compared. However, rofecoxib was withdrawn worldwide on 1 October 2004. It is unclear if the trial was completed. The STAR Multinational Study Group has conducted a phase II gastrointestinal safety and efficacy study of nitronaproxen versus naproxen in 970 patients with osteoarthritis at 80 sites in the following countries: Argentina, Brazil, Hungary, Mexico, Norway, Poland, South Africa and the UK. The study was completed in November 2002.AstraZeneca conducted a randomised, phase II trial evaluating the efficacy and safety of nitronaproxen among 672 subjects with symptomatic knee osteoarthritis. Results have been presented. Certain phase II trial data from 2003 had been somewhat disappointing. However, an underpowered trial and failures and deficiencies in a trial meant that it was not possible to draw conclusions from this data.

2006 - 04

Inflamm Allergy Drug Targets. 2006 Apr;5(2):121-31.

NO-NSAIDs: from inflammatory mediators to clinical readouts.

Fiorucci S, Antonelli E. University of Perugia, Italy.

Non-steroidal anti-inflammatory drugs (NSAIDs) and cyclo-oxygenase (COX)-2 selective inhibitors (COXIBs) are widely used drugs. However, their use is hampered by gastrointestinal, cardiovascular and renal side effects. Nitric oxide (NO)-releasing NSAIDs, NO-NSAID, are a new class of anti-inflammatory and analgesic drugs generated by adding a nitroxybutyl or a nitrosothiol moiety to the parent NSAID via a short-chain ester linkage. While efficacy of nitrosothiol-NSAIDs still awaits investigation, nitroxybutyl-NO-NSAIDs have been extensively studied in humans. The combination of balanced inhibition of the two main COX isoforms with release of NO confers to NO-NSAIDs reduced gastrointestinal and cardiorenal toxicity. It is suggested that the NO, which is released as the compounds are broken down, may counteract the consequences of the NSAID-induced decrease in gastric mucosal prostaglandins. Recent clinical trials with NO-NSAIDs have provided data consistent with pre-clinical observations.

Inflamm Allergy Drug Targets. 2006 Apr;5(2):115-9.

Nitric oxide and inflammation.

Cirino G, Distrutti E, Wallace JL. Department of Experimental Pharmacology, Università di Napoli-Federico II, Napoli, Italy.

There are several pre-clinical studies on the involvement of NO in inflammation. From this large amount of information it is clear that virtually every cell and many immunological parameters are modulated by NO. Thus, the final outcome is that NO cannot be rigidly classified as an anti-inflammatory or pro-inflammatory molecule. This peculiar aspect of the pathophysiology of NO has hampered the development of new drugs based on the concepts developed. Recent therapeutic approach are targeted to increase endogenous NO by activating the gene and some promising early data are available. At the present stage one of the most promising approach in the inflammation field is represented by a new class of NO-releasing compounds namely NO-NSAIDs that have recently enrolled in phase 2 clinical studies.

2006 - 03

Scand J Gastroenterol. 2006 Mar;41(3):264-73.

Dose-effect comparisons of the CINOD AZD3582 and naproxen on upper gastrointestinal tract mucosal injury in healthy subjects.

Wilder-Smith CH, Jonzon B, Fornstedt-Wallin B, Hedman A, Karlsson P. Gastroenterology Group Practice, Brain-Gut Research Group, Berne, Switzerland.

OBJECTIVE: The objective of this endoscopic study was to compare the effects on the gastroduodenal mucosa of healthy volunteers of different doses and dosing regimens of AZD3582, a cyclooxygenase-inhibiting nitric oxide donator (CINOD), with equimolar doses of naproxen. MATERIAL AND METHODS: Healthy volunteers were enrolled in a single-centre, randomized, double-blind, crossover trial consisting of two 12-day treatment periods and employing six sequences. The groups were: AZD3582 750 mg daily versus 375 mg twice daily (n=25), AZD3582 375 mg twice daily versus 750 mg twice daily (n=25) and naproxen 250 mg twice daily versus 500 mg twice daily (n=25). RESULTS: Gastroduodenal tract damage was similar with AZD3582 375 mg twice daily and 750 mg twice daily (mean number of erosions and ulcers+/-SD: 2.88+/-3.95 versus 3.08+/-2.80, respectively; p=0.824; 1 ulcer counted as 10 erosions). There was an indication of decreased gastroduodenal toxicity with AZD3582 750 mg daily compared with 375 mg twice daily (0.92+/-2.08 versus 2.71+/-4.75, respectively; p=0.068). Gastroduodenal toxicity was significantly lower with AZD3582 375 mg twice daily than with naproxen 250 mg twice daily (2.88+/-3.95 versus 6.16+/-9.36; p<0.05), and with AZD3582 750 mg twice daily versus naproxen 500 mg twice daily (3.08+/-2.80 versus 6.68+/-6.97; p<0.05). Equimolar twice-daily doses of AZD3582 and naproxen resulted in similar naproxen plasma levels and serum thromboxane B(2) inhibition. CONCLUSIONS: AZD3582 has an improved gastroduodenal safety profile compared with equimolar doses of naproxen. The gastroduodenal effects of AZD3582 375 mg and AZD3582 750 mg twice daily are similar. A once-daily regimen of AZD3582 might be less gastrotoxic than a twice-daily regimen.

2005 - 12

J Pharm Pharmacol. 2005 Dec;57(12):1539-54.

Clinical pharmacokinetics of the cyclooxygenase inhibiting nitric oxide donator (CINOD) AZD3582.

Fagerholm U, Bjarnsson MA. Clinical Pharmacology, AstraZeneca R&D Södertälje, S-151 85 Södertälje, Sweden.

The clinical pharmacokinetics of the COX-inhibiting nitric oxide donator (CINOD) AZD3582 and its metabolites, including naproxen, nitric oxide and nitrate, are summarized. AZD3582 has low aqueous solubility, moderate and passive intestinal permeability and is degraded by intestinal esterases. Its oral bioavailability (F) appears to be maximally a few per cent, and increases by several-fold after food intake. Ninety-four per cent or more of an AZD3582 dose is absorbed, of which at least 9-20% appears to be taken up as intact substance. AZD3582 has a predicted plasma protein binding degree of approximately 0.1%, a half-life (t1/2) of 3 to 10 h and does not accumulate after repeated once- and twice-daily dosing. In patients AZD3582 does not provide a significantly better gastrointestinal (GI) side-effect profile than the highly permeable and locally irritating naproxen. Possible reasons for this include considerable GI uptake as naproxen, limited duration and extent of nitric oxide donation in the GI mucosa and the circulation, tolerance development (involving auto-inhibition of nitric oxide catalysing enzymes) and mucosal damage caused by nitric oxide. Blood pressure data suggest that nitric oxide is mainly donated within 3 h. The uptake of naproxen is slightly slower and lower (> or = 94% relative GI uptake and 80-85% relative F) after AZD3582 administration compared with naproxen dosing. The naproxen t1/2 and trough steady-state concentrations after AZD3582 and naproxen dosing are similar. The average systemic nitrate exposure is approximately doubled after dosing of 375 to 750 mg AZD3582 twice daily.

Arthritis Rheum. 2005 Dec 15;53(6):827-37.

Comparison of the COX-inhibiting nitric oxide donator AZD3582 and rofecoxib in treating the signs and symptoms of Osteoarthritis of the knee.

Schnitzer TJ, Kivitz AJ, Lipetz RS, Sanders N, Hee A. Northwestern Center for Clinical Research, Chicago, Illinois 60611, USA.

OBJECTIVE: To compare the efficacy, safety, and tolerability of AZD3582 with that of rofecoxib, naproxen, and placebo in patients with osteoarthritis (OA) of the knee, and to define the dosage of AZD3582 (125 mg, 375 mg, and 750 mg twice a day) that is noninferior in efficacy to rofecoxib. METHODS: A double-blind study of 672 patients with OA of the knee was conducted. Patients who experienced increased pain on withdrawal of analgesia were randomized to receive AZD3582 125 mg, 375 mg, or 750 mg twice a day; rofecoxib 25 mg once a day; naproxen 500 mg twice a day; or placebo for 6 weeks. Efficacy, tolerability, and safety were monitored throughout the study. The primary variable was the change in Western Ontario and McMaster Universities Osteoarthritis Index pain subscale from baseline to the mean of weeks 4 and 6, comparing AZD3582 with placebo for superiority and with rofecoxib for noninferiority using a predefined margin of 10 mm. RESULTS: For the primary variable, AZD3582 375 mg and 750 mg were superior to placebo (least squares mean difference [95% confidence interval] -12 mm [-18, -6], P < 0.001 and -13 mm [-19, -7], P < 0.001, respectively) and were noninferior to rofecoxib (-2 mm [-8, 4], P < 0.001 and -3 mm [-9, 3], P < 0.001, respectively). AZD3582 125 mg was not significantly different from placebo for the primary variable. CONCLUSION: AZD3582 375 mg and 750 mg twice a day were superior to placebo and as effective as rofecoxib 25 mg/day in treating the signs and symptoms of OA of the knee. AZD3582 125 mg twice a day was not statistically different from placebo.

Inflammopharmacology. 2005;12(5-6):521-34.

NMI-1182, a gastro-protective cyclo-oxygenase-inhibiting nitric oxide donor.

Ellis JL, Augustyniak ME, Cochran ED, Earl RA, Garvey DS, Gordon LJ, Janero DR, Khanapure SP, Letts LG, Melim TL, Murty MG, Schwalb DJ, Shumway MJ, Selig WM, Trocha AM, Young DV, Zemtseva IS. NitroMed Inc., 125 Spring Street, Lexington, MA 02421-0781, USA.

Non-steroidal anti-inflammatory drugs (NSAIDs) are widely used to treat inflammation and to provide pain relief but suffer from a major liability concerning their propensity to cause gastric damage. As nitric oxide (NO) is known to be gastro-protective we have synthesized a NO-donating prodrug of naproxen named NMI-1182. We evaluated two cyclo-oxygenase (COX)-inhibiting nitric oxide donors (CINODs), NMI-1182 and AZD3582, for their ability to be gastro-protective compared to naproxen and for their anti-inflammatory activity. NMI-1182 and AZD3582 were found to produce similar inhibition of COX activity to that produced by naproxen. Both NMI-1182 and AZD3582 produced significantly less gastric lesions after oral administration than naproxen. All three compounds effectively inhibited paw swelling in the rat carrageenan paw edema model. In the carrageenan air pouch model all three compounds significantly reduced PGE2 levels in the pouch exudate but only NMI-1182 and naproxen inhibited leukocyte influx. These data demonstrate that NMI-1182 has comparable anti-inflammatory activity to naproxen but with a much reduced likelihood to cause gastric damage.

2005 - 11

Biochem Pharmacol. 2005 Nov 1;70(9):1343-51. Erratum in: Biochem Pharmacol. 2006 Feb 28;71(5):711.

A comparison of the cyclooxygenase inhibitor-NO donors (CINOD), NMI-1182 and AZD3582, using in vitro biochemical and pharmacological methods.

Young DV, Cochran ED, Dhawan V, Earl RA, Ellis JL, Garvey DS, Janero DR, Khanapure SP, Letts LG, Melim TL, Murty MG, Shumway MJ, Wey SJ, Zemtseva IS, Selig WM. Departments of Biology, 125 Spring St., Lexington, MA 02421, USA.

Cyclooxygenase (COX, EC inhibitor-nitric oxide (NO) donor (CINOD) hybrid compounds represent an attractive alternative to NSAID and coxib therapy. This report compares two CINODs, NMI-1182 (naproxen-glyceryl dinitrate) and AZD3582 (naproxen-n-butyl nitrate), for their ability to inhibit COX-1 and -2, deliver bioavailable nitric oxide, and release naproxen, using in vitro biochemical and pharmacological methods. In human whole blood, both CINODs showed inhibition, comparable to naproxen, of both COX isozymes and slowly released naproxen. Both CINODs donated bioavailable NO, as detected by cGMP induction in the pig kidney transformed cell line, LLC-PK1, but NMI-1182 was more potent by 30-100 times than AZD3582, GTN, GDN, and ISDN and considerably faster in inducing cGMP synthesis than AZD3582. The nitrate groups of GTN, NMI-1182, and AZD3582 appeared to be bioactivated via a common pathway, since each compound desensitized LLC-PK1 cells to subsequent challenge with the other compounds. Similar cGMP induction also occurred in normal, untransformed cells (human renal proximal tubule epithelial cells and hepatocytes from man, rat, and monkey); again, NMI-1182 was superior to AZD3582. NMI-1182 was also the more metabolically labile compound, releasing more absolute nitrate and nitrite (total NO(x)) in human stomach (in which NO is salutary) and liver S9 homogenates. Naproxen was also more rapidly freed from NMI-1182 than AZD3582 in human stomach, although liver S9 hydrolyzed both CINODs with similar rates. These in vitro tests revealed that NMI-1182 may be a better CINOD than AZD3582 because of its superior NO donating and naproxen liberating properties.

Expert Opin Investig Drugs. 2005 Nov;14(11):1347-58.

Nitric oxide-modulating agents for gastrointestinal disorders.

Whittle BJ. William Harvey Research Institute, Bart's and The London, Queen Mary's School of Medicine, Charterhouse Square, London, EC1M 6BQ, UK.

Almost 20 years after the identification of the biological role of nitric oxide (NO), the full therapeutic potential of novel agents that mimic the activity of NO or interfere with its synthesis has yet to be realised for utilities involving the gastrointestinal tract. New utilities for classical NO donors, which were used as vasodilators for decades, in the treatment of motility disorders have been explored and a product for treating anal fissure was recently launched. New classes of compounds incorporating a NO-donating moiety into standard non-steroidal anti-inflammatory drugs, the NO-non-steroidal anti-inflammatory drugs (NO-NSAIDs) or COX-inhibiting nitric oxide donors (CINODs) have also been developed. These have been shown to exhibit reduced gastrointestinal injury in experimental models, and reports on their efficacy and safety in Phase I and II studies are now available. Modulation of the inducible NO synthase isoform that generates excessive NO that can lead to subsequent cytotoxic moieties, such as peroxynitrite, may have therapeutic possibilities in a range of inflammatory diseases of the gut. Likewise, agents that promote the decomposition of peroxynitrite or removal of its other component, superoxide, may also prove to be of use. Further targets for pharmaceutical exploitation are likely to come from both genomic and molecular insights into the processes that regulate the NO system.

2005 - 07

Br J Pharmacol. 2005 Jul;145(5):679-87.

Direct gas measurements indicate that the novel cyclooxygenase inhibitor AZD3582 is an effective nitric oxide donor in vivo.

Adding LC, Agvald P, Andersson LI, Jonzon B, Hoogstraate J, Gustafsson LE. Department of Physiology and Pharmacology, Karolinska Institute, SE-171 77 Stockholm, Sweden.

1. AZD3582 [4-(nitrooxy)butyl-(2S)-2-(6-methoxy-2-naphthyl)propanoate] is a COX-inhibiting nitric oxide donor that inhibits COX-1 and COX-2. It is as effective as naproxen in models of pain and inflammation, but causes less gastroduodenal damage. Nitric oxide (NO) is generated from AZD3582 in vitro, and this study sought to show that the drug donates NO in vivo. 2. In anaesthetised male New Zealand white rabbits, the endogenous NO concentration in exhaled air was reduced by N(G)-nitro-L-arginine methyl ester (L-NAME) (30 mg kg(- 1) i.v.) from 33.5+/-1.0 ppb (mean+/-s.e.m.; n=6 per group) to 3.0+/-1.0 ppb, while increasing blood pressure and reducing heart rate. AZD3582 (0.2, 0.6, 2.0 or 6.0 micromol kg(- 1) min(- 1)) given 30 min after L-NAME increased the concentration of NO in exhaled air (P<0.05), decreased blood pressure and increased heart rate in a dose-dependent manner versus L-NAME control values. The peak mean NO concentration obtained was 44+/-8.0 ppb. 3. In in situ-perfused rabbit lungs, L-NAME (185 micromol l(- 1)) reduced the NO concentration in exhaled air from 106+/-13 to 4.0+/-0.4 ppb (n=5). Addition of AZD3582 (6 micromol min(- 1)) to the perfusate produced an initial rapid increase in the NO concentration in exhaled air, followed by a sustained, but lower plateau. Infusion of L-NAME increased, and AZD3582 decreased, pulmonary arterial pressure. 4. In both anaesthetised rabbits and in the perfused lungs, brief periods of hypoxia increased NO concentrations generated by AZD3582. 5. We conclude that, in rabbits, AZD3582 donates NO in vivo with characteristics similar to those reported for nitroglycerin and isosorbide nitrates

2005 - 06

Eur J Pharm Sci. 2005 Jun;25(2-3):229-35. Epub 2005 Mar 29.

AZD3582 increases heme oxygenase-1 expression and antioxidant activity in vascular endothelial and gastric mucosal cells.

Berndt G, Grosser N, Hoogstraate J, Schröder H. Department of Pharmacology and Toxicology, School of Pharmacy, Martin Luther University, Halle-Wittenberg, 06099 Halle (Saale), Germany.

AZD3582 [4-(nitrooxy)-butyl-(2S)-2-(6-methoxy-2-naphthyl)-propanoate] is a COX-inhibiting nitric oxide donator (CINOD). Incubation of human endothelial cells (derived from umbilical cord) with AZD3582 (10-100muM) led to increased expression of heme oxygenase (HO)-1 mRNA and protein. Heme oxygenase-1 (HO-1) is a crucial mediator of antioxidant and tissue-protective actions. In contrast, naproxen (a non-selective NSAID) and rofecoxib (a selective inhibitor of COX-2), did not affect HO-1 expression. Pre-treating endothelial cells with AZD3582 at concentrations that were effective at inducing HO-1 also reduced NADPH-dependent production of oxygen radicals. Antioxidant activity in the endothelial cells persisted after AZD3582 had been washed out from the incubation medium. When added exogenously to the cells at low micromolar concentrations, the HO-1 metabolite, bilirubin, virtually abolished NADPH-dependent oxidative stress. AZD3582-induced blockade of free-radical formation was reversed in the presence of the HO-1 inhibitor, tin protoporphyrin-IX (SnPP). Similar results were obtained in human gastric mucosal cells (KATO-III). Our results demonstrate that HO-1 is a novel target of AZD3582.

2005 - 05

J Pharm Pharmacol. 2005 May;57(5):587-97.

Pre-clinical pharmacokinetics of the cyclooxygenase-inhibiting nitric oxide donor (CINOD) AZD3582.

Fagerholm U, Breuer O, Swedmark S, Hoogstraate J. Clinical Pharmacology, AstraZeneca R&D Södertälje, S-151 85 Södertälje, Sweden.

The pre-clinical pharmacokinetics of AZD3582 (4-(nitrooxy)butyl-(2S)-2-(6-methoxy-2-naphthyl) propanoate) and its primary metabolites (naproxen and nitrate) were evaluated. AZD3582 had intermediate and passive intestinal permeability (40 times lower than for naproxen), high systemic plasma clearance (CL), substantial gastrointestinal hydrolysis, intermediate volume of distribution (Vss; >or=3.4 L kg-1) and half-life (t1/2; 7 h), negligible plasma protein binding (approximately 0.1%), low/intermediate oral uptake (>or=13% as intact substance) and low and varying oral bioavailability (mean 1.4% in minipigs and 3.9% in dogs). Following administration of therapeutically relevant oral doses, plasma concentrations of AZD3582 were very low (40 h in rats, minipigs and dogs, respectively. The Vss and CL for naproxen were small. Plasma protein binding was extensive, and saturation was observed within the therapeutic dose and concentration range. Intake of food prolonged the systemic absorption of naproxen in the minipig. The pharmacokinetics of naproxen did not show apparent time- or gender-related dependency. Following oral dosing of [3H]-, [14C]- and [15N]-AZD3582, most [14C]- and [3H]-activity was excreted in urine and expired air, respectively. Seventeen per cent of [15N] was recovered in minipig urine as [15N]-nitrate. About 30% of [3H]-activity (naproxen and/or naproxen-related metabolites) was excreted in bile and re-absorbed. Concentrations of [14C]-activity (nitrooxy-butyl group and/or its metabolites) in milk were higher than in plasma and [3H]-activity in milk. [3H]- and [14C]-excretion data indicated that intact AZD3582 was not excreted in urine, bile or milk to a significant extent. There was no apparent consistency between tissue distribution of [14C]- and [3H]-activity in the rat, which suggests rapid and extensive metabolism of extravascularly distributed AZD3582. A substantial increase of plasma nitrate levels was found after single and repeated oral doses

Clin Pharmacol Ther. 2005 May;77(5):437-50.

Renal effects of the cyclooxygenase-inhibiting nitric oxide donator AZD3582 compared with rofecoxib and naproxen during normal and low sodium intake.

Huledal G, Jonzon B, Malmenas M, Hedman A, Andersson LI, Odlind B, Brater DC. Experimental Medicine, AstraZeneca R&D Södertälje, SE-151 85 Södertälje, Sweden.

BACKGROUND: Nonsteroidal anti-inflammatory drugs (NSAIDs) inhibit cyclooxygenase (COX) and can thereby reduce renal function, especially with respect to solute excretion and renal perfusion. AZD3582 [4-(nitrooxy)butyl-(2S)-2-(6-methoxy-2-naphthyl)propanoate] is a COX-inhibiting nitric oxide donator. Donation of nitric oxide by AZD3582 could preserve blood flow and thereby counteract the deleterious effects of COX inhibition in the gastrointestinal tract and possibly in other organ systems, including the kidney. The aim of this single-dose study was to assess the hypothesis that AZD3582 would not adversely affect renal function compared with NSAIDs. METHODS: In a parallel, randomized, double-blind fashion, a total of 60 healthy subjects (age range, 20-44 years) received 2 single doses of 750 mg AZD3582, 1500 mg AZD3582, 50 mg rofecoxib, 500 mg naproxen, or placebo (n = 12 per group). The first dose was given after a 5-day normal-sodium diet (150 mmol/d), and the second was given after a consecutive 3-day low-sodium diet (10 mmol/d). Urinary sodium excretion during normal sodium intake and glomerular filtration rate (GFR) (assessed by iohexol clearance) during sodium depletion were the primary variables measured. RESULTS: Urinary sodium excretion was reduced in all active treatment groups (maximal reduction of approximately 11 mmol/h during normal sodium intake, P < .05 versus placebo for all groups). GFR was also reduced in all active treatment groups. In sodium-depleted subjects, the mean (SD) maximal reduction in GFR during 0 to 6 hours for 750 mg AZD3582, 1500 mg AZD3582, 50 mg rofecoxib, and 500 mg naproxen was 28.1 mL/min (13.5 mL/min), 33.7 mL/min (23.3 mL/min), 25.2 mL/min (29.2 mL/min), and 41.7 mL/min (30.7 mL/min), respectively, with a statistically significant difference between 500 mg naproxen and placebo. Relative changes in sodium excretion and GFR were similar during normal sodium intake and sodium depletion during active treatment. CONCLUSION: The renal effects of 750 mg and 1500 mg AZD3582 were similar to those of 500 mg naproxen and 50 mg rofecoxib. Thus the potential for nitric oxide donation to create a renal-sparing agent was not found for AZD3582.

2005 - 03

Ann Rheum Dis. 2005 Mar;64(3):449-56. Epub 2004 Sep 2.

A randomised, placebo controlled, comparative trial of the gastrointestinal safety and efficacy of AZD3582 versus naproxen in osteoarthritis.

Lohmander LS, McKeith D, Svensson O, Malmenas M, Bolin L, Kalla A, Genti G, Szechinski J, Ramos-Remus C; STAR Multinational Study Group. Department of Orthopaedics, Lund University, SE-221 85 Lund, Sweden.

OBJECTIVE: To evaluate the gastrointestinal safety and efficacy of the COX inhibiting nitric oxide donator AZD3582 in patients with hip or knee osteoarthritis. METHODS: 970 patients were randomised (7:7:2) to AZD3582 750 mg twice daily, naproxen 500 mg twice daily, or placebo twice daily in a double blind study. The primary end point was the six week incidence of endoscopic gastroduodenal ulcers (diameter > or =3 mm). Overall damage measured on the Lanza scale was a secondary end point. Safety and tolerability assessments included endoscopic upper gastrointestinal erosions and the gastrointestinal symptom rating scale (GSRS). Efficacy was primarily assessed by WOMAC. RESULTS: The incidence of ulcers with AZD3582 was 9.7% and with naproxen 13.7% (p = 0.07, NS), v 0% on placebo. The incidence of Lanza scores >2 was higher with naproxen (43.7%) than with AZD3582 (32.2%) (p<0.001). Compared with baseline, significantly fewer ulcers and erosions developed in stomach and stomach/duodenum combined, and fewer erosions developed in stomach, duodenum, and both combined on AZD3582 than on naproxen. GSRS reflux and abdominal pain subscale scores were lower for AZD3582 than for naproxen but there was no difference for indigestion, constipation, and diarrhoea. AZD3582 was as effective as naproxen at improving WOMAC scores. Both agents were well tolerated, with no significant effects on blood pressure. CONCLUSIONS: At doses with similar efficacy in relieving osteoarthritis symptoms, the primary end point of six week endoscopic gastroduodenal ulcer incidence was not significantly different between AZD3582 and naproxen. Most secondary endoscopic gastrointestinal end points favoured AZD3582.

2005 - 01

Eur J Pharmacol. 2005 Jan 31;508(1-3):7-13. Epub 2004 Dec 28.

NSAIDs increase GM-CSF release by human synoviocytes: comparison with nitric oxide-donating derivatives.

Zacharowski P, Breese E, Wood E, Del Soldato P, Warner T, Mitchell J. Cardiac, Vascular and Inflammation Research, The William Harvey Research Institute, Bart's and The London, Queen Mary School of Medicine and Dentistry, London, UK.

Non-steroidal anti-inflammatory drugs (NSAIDs) are used to treat the condition of rheumatoid arthritis, where levels of prostaglandin E2 (PGE2) and granulocyte macrophage-colony stimulating factor (GM-CSF) are elevated in the synovial fluid. NO-NSAIDs are a new class of cyclooxygenase (COX)-inhibitors developed by coupling a nitric oxide (NO)-donating moiety to conventional NSAIDs. We show that, in cytokine-treated synoviocytes (from non-rheumatic patients), NO-naproxen and NO-flurbiprofen like their parent compounds concentration-dependently reduce the levels of PGE2 (an index of COX-2 activity), with a corresponding rise in the release of GM-CSF. Unlike acetylsalicylic acid (ASA), NO-ASA reduces the levels of PGE2, without increasing GM-CSF release, although cell viability is reduced at the highest concentration (1 mM). The effects of NSAIDs and NO-NSAIDs on GM-CSF release were attributable to the PGE2 mediated cyclic (c) AMP pathway because PGE2 reversed the effects of COX blockade. Second, phosphodiesterase inhibitors 3-isobutyl-1-methylxanthine (IBMX) and Ro-201724 (both of which elevate camp levels) decreased GM-CSF release, in the presence of PGE2. Finally, neither sodium nitroprusside nor zaprinast (both of which elevate cGMP levels) affected GM-CSF or PGE2 release. Our findings demonstrate that GM-CSF is regulated by NSAIDs and NO-NSAIDs via inhibition of COX and appears to be mediated via the cAMP pathway. NO-ASA is the exception, because it does not increase GM-CSF release, although at millimolar concentrations cell viability is reduced.

2004 - 12

J Pharmacol Exp Ther. 2004 Dec;311(3):1264-71. Epub 2004 Aug 5.

Nitric oxide (NO)-releasing naproxen (HCT-3012 [(S)-6-methoxy-alpha-methyl-2-naphthaleneacetic Acid 4-(nitrooxy)butyl ester]) interactions with aspirin in gastric mucosa of arthritic rats reveal a role for aspirin-triggered lipoxin, prostaglandins, and NO in gastric protection.

Fiorucci S, Di Lorenzo A, Renga B, Farneti S, Morelli A, Cirino G. Clinica di Gastroenterologia ed Endoscopia Digestiva, Policlinico Monteluce, 06100 Perugia, Italy.

Administration of selective and nonselective cyclooxygenase (COX)-2 inhibitors to rheumatoid arthritis patients taking low doses of acetylsalicylic acid (ASA) for cardiovascular prevention associates with increased risk of gastrointestinal bleeding. The present study was undertaken to investigate whether administration of HCT-3012 [(S)-6-methoxy-alpha-methyl-2-naphthaleneacetic acid 4-(nitrooxy)butyl ester], a nitric oxide (NO)-releasing derivative of naproxen, exacerbates gastric mucosal injury in arthritic rats administered low doses of ASA. Our results demonstrated that while treating arthritic rats with a dose of 30 mg/kg/day ASA causes detectable mucosal injury, but had no effect on arthritis score and interleukin-6 plasma levels, coadministration of naproxen (10 mg/kg/day) and celecoxib (30 mg/kg/day), in combination with ASA from day 7 to day 21, attenuates arthritis development (P <0.01>0.001 versus arthritis) without exacerbating gastric mucosal injury caused by ASA. Arthritis development associates with gastric COX-2 induction, mRNA and protein, and enhanced gastric prostaglandin E2 (PGE2) synthesis (P <0.01 versus control rats). Although all treatments, including celecoxib, were effective in reducing gastric PGE2 synthesis, administering arthritic rats with ASA resulted in a significant increase in gastric content of aspirin-triggered lipoxin (ATL), a COX-2-derived lipid mediator that regulates proinflammatory responses at the neutrophils/endothelial interface. Administering arthritic rats with naproxen and celecoxib abrogates ATL formation induced by ASA although enhanced neutrophils accumulate into the gastric mucosa (P <0.01 versus ASA alone). In contrast, whereas HCT-3012 inhibited ATL formation, it did not increase neutrophil recruitment into the gastric microcirculation. Collectively, these data indicate that HCT-3012 derived from NO has the potential to compensate for inhibition of PGE2 and ATL and to protect the gastric mucosa by limiting the recruitment of neutrophils. These data suggest that HCT-3012 might be a safer alternative to nonsteroidal anti-inflammatory drugs and coxibs in rheumatic patients that take low doses of ASA.

2004 - 08

J Pharmacol Exp Ther. 2004 Aug;310(2):555-62. Epub 2004 Apr 13.

The nitric oxide-releasing naproxen derivative displays cardioprotection in perfused rabbit heart submitted to ischemia-reperfusion.

Rossoni G, Manfredi B, Del Soldato P, Berti F. Department of Pharmacological Sciences, University of Milan, Via Vanvitelli 32, 20129 Milan, Italy.

In this study, the pharmacological activity of HCT-3012 [(S)-6-methoxy-alpha-methyl-2-naphtaleneacetic acid 4-(nitrooxy)butyl ester], a nitric oxide (NO)-releasing derivative of naproxen, was compared with that of naproxen in a model of acute ischemia (40 min) and reperfusion (20 min) of the rabbit heart. HTC-3012 (3-100 microM), in spite of inhibition of 6-keto-prostaglandin F(1alpha) generation by the cardiac tissues, brought about a dose-dependent normalization of coronary perfusion pressure, associated with a reduction of ventricular contracture during ischemia with remarkable improvement of left ventricular developed pressure at reperfusion. These beneficial effects were accompanied by a substantial release of nitrite/nitrate in the heart perfusates, indicating that NO has been released by HCT-3012 and donated to the cardiac tissue. These events were paralleled by a significant reduction of creatine kinase activity in heart perfusates during reperfusion. Naproxen (10-100 microM) aggravated the myocardial damage in ischemic reperfused hearts, severely depressing the postischemic ventricular dysfunction. Perfusion of the heart with N(G)-monomethyl-l-arginine (10 microM) caused a marked aggravation of myocardial damage of the reperfused hearts, and this effect was dose dependently prevented by HCT-3012 but not by naproxen. The results of the present experiments clearly indicate that HCT-3012, by donating NO, displays a noticeable anti-ischemic effect in reperfused ischemic rabbit hearts. The safer gastrointestinal profile of HCT-3012 and its ability to control experimental hypertension, suggest that this compound may have therapeutical potential in cardiovascular disease, namely in the prevention of myocardial ischemic events, and may represent a better alternative to conventional nonsteroidal anti-inflammatory drugs.

2004 - 02

Eur J Pharm Sci. 2004 Feb;21(2-3):331-5.

A common pathway of nitric oxide release from AZD3582 and glyceryl trinitrate.

Berndt G, Grosser N, Hoogstraate J, Schröder H. Department of Pharmacology and Toxicology, School of Pharmacy, Martin Luther University, Wolfgang-Langenbeck-Street 4, 06099, Halle (Saale), Germany.

4-(Nitrooxy)-butyl-(S)-2-(6-methoxy-2-naphthyl)-propanoate (AZD3582) is a cyclooxygenase (COX)-inhibiting nitric oxide donator (CINOD). It donates nitric oxide (NO) in biological systems through as yet unidentified mechanisms. cGMP, a marker of intracellularly generated NO, was increased up to 27-fold over basal levels by AZD3582 (1-30microM) in LLC-PK1 kidney epithelial cells. A 5h pretreatment with glyceryl tinitrate (GTN, 0.1-1microM) attenuated the cGMP response to a subsequent challenge with AZD3582 or GTN. Similarly, AZD3582 (10-30microM) pretreatment reduced the increase in cGMP on subsequent incubation with AZD3582 or GTN. In contrast, cGMP stimulation by SIN-1, which releases NO independently of enzymatic catalysis, remained unimpaired in cells pretreated with GTN or AZD3582. Our results demonstrate that AZD3582 decreases the sensitivity of the guanylyl cyclase/cGMP system to GTN and vice versa. This suggests that bioactivation pathways for organic nitrates, which involve enzymatic catalysis, may be responsible for NO donation from AZD3582.

2003 - 11

Inflammopharmacology. 2003;11(4):437-44.

The CINOD, AZD3582, exhibits an improved gastrointestinal safety profile compared with naproxen in healthy volunteers.

Jonzon B, Bjarnason I, Hawkey C, Jones J, Goddard A, Fagerholm U, Karlsson P. Experimental Medicine, AstraZeneca R&D Södertälje, S-151 85 Södertälje, Sweden.

COX-inhibiting nitric oxide donators (CINODs) are a new class of drugs in development for the treatment of acute and chronic pain. They comprise a COX-inhibiting moiety linked to a nitric-oxide-donating component and are designed to provide an innovative mechanism of action of balanced COX inhibition and controlled nitric oxide donation. Through these pathways, CINODs should provide analgesic and anti-inflammatory efficacy, while offering gastrointestinal safety through the tissue-protective effects of nitric oxide donation. AZD3582 [4-(nitrooxy)butyl-(2S)-2-(6-methoxy-2-naphthyl)propanoate] is the first agent in the CINOD class to enter extensive clinical development. Pre-clinical studies demonstrate that AZD3582 has a superior gastrointestinal safety profile to naproxen, while demonstrating analgesic and anti-inflammatory efficacy. In healthy human volunteers, AZD3582 caused little gastrointestinal damage compared with equimolar doses of naproxen. Studies to evaluate the longer-term gastrointestinal safety of AZD3582, alongside its efficacy in alleviating chronic and acute pain, are ongoing.

Inflammopharmacology. 2003;11(4):423-8.

COX-inhibiting nitric oxide donators (CINODs) - a new paradigm in the treatment of pain and inflammation.

Hoogstraate J, Andersson LI, Berge OG, Jonzon B, Ojteg G. Research DMPK, AstraZeneca R&D Södertälje, S-151 85 Södertälje, Sweden.

The clinical utility of non-selective non-steroidal anti-inflammatory drugs (NSAIDs) for pain relief is tempered by their propensity to cause gastrointestinal toxicity. Cyclooxygenase (COX)-inhibiting nitric oxide donators (CINODs) are a new class of drugs designed to provide analgesic efficacy through COX inhibition and gastrointestinal safety through the protective effects of controlled nitric oxide donation. Pre-clinical studies assessing the pharmacology, efficacy and gastrointestinal safety of AZD3582 [4-(nitrooxy)butyl-(2S)-2-(6-methoxy-2-naphthyl)propanoate] support this concept. Based on these studies, AZD3582 was the first CINOD to enter clinical development for the treatment of acute and chronic pain. The potential clinical utility of this new class is illustrated by a study of AZD3582 in healthy volunteers in which it caused significantly less acute gastrointestinal toxicity than an equimolar dose of naproxen. The results of the animal studies and the initial clinical study warrant long-term tolerability studies of AZD3582 along with evaluation of its anti-inflammatory and analgesic effects in humans.

Gut. 2003 Nov;52(11):1537-42.

Gastrointestinal safety of AZD3582, a cyclooxygenase inhibiting nitric oxide donator: proof of concept study in humans.

Hawkey CJ, Jones JI, Atherton CT, Skelly MM, Bebb JR, Fagerholm U, Jonzon B, Karlsson P, Bjarnason IT. Division of Gastroenterology, University Hospital Nottingham, Nottingham, UK.

BACKGROUND: Cyclooxygenase inhibiting nitric oxide donators (CINODs) are a new class of anti-inflammatory and analgesic drugs that may minimise gastrointestinal toxicity compared with standard non-steroidal anti-inflammatory drugs (NSAIDs) by virtue of nitric oxide donation. METHODS: A proof of concept study of the gastrointestinal safety of AZD3582, the first CINOD available for human testing, was conducted. Thirty one subjects were randomised to receive placebo, naproxen 500 mg twice daily, or its nitroxybutyl derivative AZD3582 in an equimolar dose (750 mg twice daily) for 12 days in a double blind three period crossover volunteer study. At the start and end of each dosing period, gastroduodenal injury was assessed by endoscopy and small bowel permeability by differential urinary excretion of lactulose and L-rhamnose. Pharmacokinetic profiles were assessed at steady state. RESULTS: On naproxen, the mean total number of gastroduodenal erosions was 11.5 (and one subject developed an acute ulcer) versus 4.1 on AZD3582 (p<0.0001). More than half of the subjects had no erosions on AZD3582. Differences were seen for both the stomach and duodenum. Naproxen increased intestinal permeability (lactulose:L-rhamnose ratio 0.030 before v 0.040 after treatment) whereas AZD3582 (0.029 before, 0.029 after; p=0.006 v naproxen) and placebo (0.030 before, 0.028 after; p<0.001 v naproxen) did not. The steady state bioavailability of naproxen metabolised from AZD3582 was 95% (95% confidence interval 87-101%) of that after naproxen administration. CONCLUSIONS: This human study supports animal data showing reduced gastrointestinal toxicity with the CINOD AZD3582. The potential combination of effective pain relief and gastrointestinal protection offered by AZD3582 warrants further evaluation in human clinical studies.

2003 - 05

Dig Liver Dis. 2003 May;35 Suppl 2:S27-34.

NO-NSAIDs and cancer: promising novel agents.

Rigas B, Kalofonos H, Lebovics E, Vagenakis AG. American Health Foundation, 1 Dana Road, Valhalla, NY 10595, USA.

Three potential applications of NO-donating NSAIDs in human cancer include their use: as chemopreventive agents; against already developed cancers (chemotherapy); and for the control of cancer symptoms, notably cancer pain. The evidence to date of greater safety and enhanced efficacy of NO-donating NSAIDs underscores their potential to prevent colon cancer and overcome the limitations of traditional NSAIDs. NO-donating NSAIDs affect several pathways critical to colon carcinogenesis and this may explain in part their greater efficacy in colon cancer prevention as assessed in preclinical models.

2002 - 10

Br J Pharmacol. 2002 Oct;137(3):295-310.

Pharmacology and potential therapeutic applications of nitric oxide-releasing non-steroidal anti-inflammatory and related nitric oxide-donating drugs.

Keeble JE, Moore PK. Centre for Cardiovascular Biology and Medicine, King's College, University of London, Guy's Campus, London SE1 9RT.

This review examines the biological significance, therapeutic potential and mechanism(s) of action of a range of nitric oxide-releasing non-steroidal anti-inflammatory drugs (NO-NSAID) and related nitric oxide-releasing donating drugs (NODD). The slow release of nitric oxide (NO) from these compounds leads to subtle changes in the profile of pharmacological activity of the parent, non-steroidal anti-inflammatory drugs (NSAID). For example, compared with NSAID, NO-NSAID cause markedly diminished gastrointestinal toxicity and improved anti-inflammatory and anti-nociceptive efficacy. In addition, nitroparacetamol exhibits hepatoprotection as opposed to the hepatotoxic activity of paracetamol. The possibility that NO-NSAID or NODD may be of therapeutic benefit in a wide variety of disease states including pain and inflammation, thrombosis and restenosis, neurodegenerative diseases of the central nervous system, colitis, cancer, urinary incontinence, liver disease, impotence, bronchial asthma and osteoporosis is discussed.

2001 - 11

Drug Saf. 2001;24(11):801-11.

Nitric oxide-releasing NSAIDs: a review of their current status.

Fiorucci S, Antonelli E, Burgaud JL, Morelli A. Clinica di Gastroenterologia ed Epatologia Dipartimento di Medicina Clinica, e Sperimentale, Universita' degli Studi di Perugia, Italy.

Nonsteroidal anti-inflammatory drugs (NSAIDs) are among the most widely prescribed drugs worldwide owing to their anti-inflammatory, antipyretic and analgesic properties. However, their use is hampered by gastrointestinal (GI) toxicity, the most common drug-related serious adverse event in industrialised nations. Nitric oxide (NO)-releasing NSAIDs, a recently described class of drugs, are generated by adding a nitroxybutyl or a nitrosothiol moiety to the parent NSAID via a short-chain ester linkage. While efficacy of nitrosothiol-NO-NSAIDs still awaits investigation, nitroxybutyl-NO-NSAIDs have been extensively studied in animals, thus the abbreviation NO-NSAIDs used here refers to the latter group of NSAID derivatives. NO-NSAIDs retain the anti-inflammatory and antipyretic activity of original NSAIDs, although they exhibit markedly reduced gastrointestinal toxicity. NO-NSAIDs are nonselective cyclo-oxygenase (COX) inhibitors, and they also exert COX-independent activities that are NO-dependent. Indeed, NO-NSAIDs suppress production of the cytokines interleukin (IL)-1beta, IL-18 and interferon-gamma by causing the S-nitrosilation/inhibition of caspase-1. In acute and chronic animal models of inflammation, it has been demonstrated that NO-NSAIDs abrogated prostaglandin E2 as well as thromboxane B2 generation. In a murine model, NO-naproxen was approximately 10-fold more potent than naproxen in reducing animal writhing after intraperitoneal injection of acetic acid. Similar data have been obtained in chronic models of pain such as rat adjuvant arthritis. In vivo and in vitro studies suggest that NO-aspirin (acetylsalicylic acid) exerts more potent antithrombotic action than aspirin, probably by coupling the ability to inhibit COX-1 with the anti-adhesive effect of NO. Moreover, in a model of renal injury NO-flurbiprofen not only has been demonstrated to be devoid of nephrotoxicity but also to ameliorate renal function. Finally, in an animal model of chronic neurodegenerative disease, NO-flurbiprofen and NO-aspirin attenuated the brain inflammatory response. The GI toxicity of NO-flurbiprofen and NO-naproxen is currently being investigated in healthy individuals.

2001 - 08

Br J Pharmacol. 2001 Aug;133(7):1023-8.

Vasorelaxant effect of nitric oxide releasing steroidal and nonsteroidal anti-inflammatory drugs.

Keeble J, Al-Swayeh OA, Moore PK. Messengers and Signalling Group, School of Biomedical Sciences, King's College London, Hodgkin Building, Guy's Campus, London SE1 9RT, UK.

The effect of several nitric oxide releasing-non-steroidal anti-inflammatory drugs (NO-NSAID) and nitroprednisolone on blood vessel relaxation in vitro and in vivo was studied. Nitroflurbiprofen (NOF; EC(50), 688.8+/-93.8 microM), nitroaspirin (NOA; EC(50), 57.9+/-6.5 microM), nitroparacetamol (NOPARA; EC(50), 71.5+/-14.6 microM) and nitroprednisolone (EC(50), 15.1+/-1.4 microM) caused concentration-related relaxation of noradrenaline (NA)-contracted rat aortic rings. All NO releasing compounds tested were approximately three orders of magnitude less potent than sodium nitroprusside (SNP, EC(50), 35.7+/-3.5 nM). The vasorelaxant effect of NOF and NOPARA in the rat aorta was potentiated by zaprinast (5 microM) and reduced by ODQ (5 microM). Flurbiprofen and paracetamol (100 microM) caused minimal (<10%)>0.05) but increased by removal of the endothelium (EC(30), 164.3+/-26.3 microM cf. EC(50), 688.8+/-93.8 microM, P<0.05). NOF (0.1 - 50 microM) produced a small but not concentration-related vasodilation of the NA-preconstricted (i.e. "high tone") perfused rat mesentery preparation (cf. SNP, EC(30), 4.4+/-0.7 microM). In contrast, NOF (1 - 100 microM) produced concentration-related vasodilation of the "high tone" perfused rat kidney with an EC(50) of 33.1+/-4.4 microM. Neither NOF (74 mg kg(-1), i.p.) nor NOA (91.9 mg kg(-1), i.p.) nor equimolar doses of flurbiprofen (50 mg kg(-1), i.p.) or aspirin (50 mg kg(-1), i.p.) affected mean arterial blood pressure (MAP) or heart rate (HR) of pentobarbitone-anaesthetized rats over a 1 h period. NO-NSAID relax blood vessels in vitro by an NO-dependent mechanism. The absolute vasorelaxant effect of NO releasing drug varies greatly with the choice of compound and between blood vessel preparations.

2000 - 08

Am J Physiol Heart Circ Physiol. 2000 Aug;279(2):H528-35.

Antihypertensive properties of a nitric oxide-releasing naproxen derivative in two-kidney, one-clip rats.

Muscara MN, McKnight W, Lovren F, Triggle CR, Cirino G, Wallace JL. Department of Pharmacology and Therapeutics, University of Calgary, Calgary, Alberta, T2N 4N1, Canada.

Nonsteroidal anti-inflammatory drugs have been reported to exacerbate hypertension. In this study, we tested the hypothesis that a nitric oxide-releasing derivative of naproxen would ameliorate hypertension in the rat. Hypertension was induced by partially occluding one renal artery (the "2K,1C" model), and 2 wk later the rats started receiving naproxen, the nitric oxide-releasing derivative HCT-3012, or vehicle each day for 2 wk. Naproxen significantly exacerbated the hypertension. HCT-3012 significantly reduced blood pressure relative to both the naproxen- and vehicle-treated groups. Both naproxen and HCT-3012 markedly suppressed whole blood thromboxane B(2) synthesis. In studies of anesthetized rats, naproxen significantly enhanced the late hypertensive response to endothelin-1 and significantly blunted the early hypotensive response. In contrast, HCT-3102 did not affect either response to endothelin-1. In vitro, HCT-3012 significantly reduced the responsiveness of aortic rings to the contractile effects of phenylephrine. These studies suggest that HCT-3012 reduces blood pressure in hypertensive rats, not simply through the vasodilatory actions of the nitric oxide it releases, but through alterations in the responsiveness of the vasculature to endogenous pressor agents.

2000 - 07

Br J Pharmacol. 2000 Jul;130(6):1399-405.

NO-naproxen modulates inflammation, nociception and downregulates T cell response in rat Freund's adjuvant arthritis.

Cicala C, Ianaro A, Fiorucci S, Calignano A, Bucci M, Gerli R, Santucci L, Wallace JL, Cirino G. Dipartimento di Farmacologia Sperimentale, Università degli Studi di Napoli - Federico II, Napoli, Italy. 1.

Anti-inflammatory non steroidal drugs releasing NO (NO-NSAIDs) are a new class of anti-inflammatory drugs to which has been added an NO-releasing moiety. These compounds have been shown to retain the anti-inflammatory, analgesic and antipyretic activity of the parent compound but to be devoid of gastrointestinal (GI) toxicity. 2. Freund's adjuvant (FA) arthritis was induced in rats by a single intraplantar injection into the right hindpaw of 100 microl of mycobacterium butirricum (6 mg ml (-1)). The effect of equimolar doses of naproxen (1, 3 and 10 mg kg(-1)) and NO-naproxen (1.5, 4.5 and 16 mg kg(-1)) was evaluated using two dosage regimen protocols: (i) preventive, starting oral administration of the drugs at the time of induction of arthritis and for the following 21 days (day 1 - 21); (ii) therapeutic, starting oral administration of the drugs 7 days after adjuvant injection and for the following 14 days (day 7 - 21). 3. Hindpaw swelling (days 3, 7, 11, 14, 17, 21) and nociception (days 15 and 21) were measured. On day 22 rats were sacrificed, draining lymph nodes were removed and T cells isolated. In vitro proliferation of T cells following stimulation with concanavalin A (0.5 - 5 microg ml(-1)) was measured using a tritiated thymidine incorporation assay. IL-2 receptor expression on T cells was measured by FACS analysis. 4. Naproxen and NO-naproxen showed similar activity in reducing oedema formation in the non-injected (controlateral) hindpaw. Both drugs showed anti-nociceptive effect. NO-naproxen was anti-nociceptive at a dose of 4.5 mg kg(-1) while naproxen showed the same extent of inhibition only at a dose of 10 mg kg(-1). 5. T cells were isolated and characterized by FACS analysis. Stimulation of isolated T cells with concanavallin A in vitro caused a significant increase in thymidine uptake. NO-naproxen at a dose of 4.5 mg kg(-1) inhibited T cell proliferation to the same extent as 10 mg kg(-1) of naproxen. 6. Inhibition of T cell proliferation was well correlated with reduced IL-2 receptor expression on T cells. In addition, NO-naproxen reduced both IL-1beta and TNFalpha plasma levels whilst naproxen reduced IL-1beta levels only. 7. In conclusion, both naproxen and NO-naproxen reduce inflammation and nociception associated with arthritis. In addition NO-naproxen interferes to a larger extent with cellular mechanism involved in T cell activation in rat adjuvant arthritis indicating that introduction of the NO moiety in the naproxen structure increases the effect at the level of the immune system.

2000 - 02

Br J Pharmacol. 2000 Feb;129(4):681-6.

Wound collagen deposition in rats: effects of an NO-NSAID and a selective COX-2 inhibitor.

Muscara MN, McKnight W, Asfaha S, Wallace JL. Department of Pharmacology, University of Calgary, 3330 Hospital Drive NW, Calgary, AB, T2N 4N1, Canada.

Selective cyclo-oxygenase (COX)-2 inhibitors and nitric oxide-releasing nonsteroidal anti-inflammatory drugs (NSAIDs) exhibit reduced toxicity in the gastrointestinal tract, but may affect wound healing in other tissues. In this study, we have compared the effects of a selective COX-2 inhibitor (celecoxib), anitric-oxide releasing derivative of naproxen (HCT-3012) and naproxen in a model of wound collagen deposition in the rat. Polyvinyl alcohol sponges were implanted subcutaneously in rats. The rats were treated daily for 5 days with the test drugs at equieffective anti-inflammatory doses. Naproxen (10 mg kg(-1)) significantly decreased (45%) collagen deposition at the wound site relative to the vehicle-treated control group. In contrast, HCT-3012 (14.5 mg kg(-1)) significantly increased (62%) collagen deposition, while celecoxib (10 mg kg(-1)) had no effect. Naproxen and HCT-3012 suppressed prostaglandin (PG) E(2) levels at the wound site and whole blood thromboxane synthesis to similar degrees. Celecoxib had no significant effect on wound fluid PGE(2) levels, but slightly reduced whole blood thromboxane synthesis (by 17%). COX-1 mRNA and protein were expressed in the wound exudate, the skin surrounding the wound and in normal skin. In contrast, COX-2 mRNA, but not protein, was expressed in wound and normal skin. These results demonstrate that HCT-3012 can significantly enhance collagen deposition at a wound site, despite inhibiting prostaglandin synthesis to the same extent as the parent drug. Nitric oxide-releasing NSAIDs may represent a safer alternative to standard NSAIDs for use as anti-inflammatory and analgesic agents by post-surgery patients.

1999 - 03

Aliment Pharmacol Ther. 1999 Mar;13(3):421-35.

Nitric oxide-releasing NSAIDs inhibit interleukin-1beta converting enzyme-like cysteine proteases and protect endothelial cells from apoptosis induced by TNFalpha.

Fiorucci S, Santucci L, Federici B, Antonelli E, Distrutti E, Morelli O, Renzo GD, Coata G, Cirino G, Soldato PD, Morelli A. Sezione di Gastroenterologia ed Epatologia, Dipartimento di Medicina Clinica, e Sperimentale, Universita' degli Studi di Perugia, Perugia, Italy.

BACKGROUND: Nitric oxide (NO)-releasing NSAIDs are a new class of NSAID derivatives with markedly reduced gastrointestinal toxicity. Although it has been demonstrated that NO-NSAIDs spare gastric mucosal blood flow, molecular determinants involved in this effect are unknown. AIM: To investigate the effect of aspirin, naproxen and flurbiprofen, and their NO-derivatives, on gastric apoptosis and endothelial cell damage induced by tumour necrosis factor-alpha (TNFalpha). In other systems, TNFalpha-induced apoptosis is mediated by caspases, a growing family of cysteine proteases similar to the IL-1beta converting enzyme (ICE), and so we have investigated whether NO-NSAIDs modulate ICE-like endopeptidases. METHODS: Rats were treated orally with aspirin, naproxen and flurbiprofen, or their NO-releasing derivatives in equimolar doses, and were killed 3 h later to assess mucosal damage and caspase activity. Endothelial cells (HUVECs) were obtained from human umbilical cord by enzymatic digestion. Caspase 1 and 3 activities were measured by a fluorimetric assay using selective peptides as substrates and inhibitors. Apoptosis was quantified by ELISA specific for histone-associated DNA fragments and by the terminal transferase nick-end translation method (TUNEL). RESULTS : In vivo NSAID administration caused a time-dependent increase in gastric mucosal damage and caspase activity. NCX-4016, NO-naproxen and NO-flurbiprofen did not cause any mucosal damage and prevented cysteine protease activation. NSAIDs and NO-NSAIDs stimulated TNFalpha release. Exposure to TNFalpha resulted in a time- and concentration-dependent HUVEC apoptosis, an effect that was prevented by pretreating the cells with NCX-4016, NO-naproxen, NO-flurbiprofen, SNP or Z-VAD.FMK, a pan-caspase inhibitor. The activation of ICE-like cysteine proteases was required to mediate TNFalpha-induced apoptosis of HUVECs. Exogenous NO donors inhibited TNFalpha-induced cysteine protease activation. Inhibition of caspase activity was due to S-nitrosylation of ICE/CPP32-like proteases. NO-NSAIDs prevented IL-1beta release from endotoxin-stimulated macrophages. CONCLUSIONS: NO-releasing NSAIDs are a new class of non-peptide caspase inhibitors. Inhibition of ICE-like cysteine proteases prevents endothelial cell damage induced by pro-inflammatory agents and might contribute to the gastro-protective effects of NO-NSAIDs.

1998 - 01

Life Sci. 1998;62(15):PL235-40.

Effect of a nitric oxide-releasing naproxen derivative on hypertension and gastric damage induced by chronic nitric oxide inhibition in the rat.

Muscara MN, McKnight W, Del Soldato P, Wallace JL. Department of Pharmacology and Therapeutics, University of Calgary, Alberta, Canada.

NSAIDs can elevate blood pressure through mechanisms such as renal vasoconstriction and sodium retention. These effects are particularly evident in hypertensive individuals. Nitric oxide-releasing NSAID derivatives have been shown to have greatly reduced toxicity in the gastrointestinal tract and kidney. We therefore evaluated the effects of a 4 week treatment with either naproxen or its nitric oxide-releasing derivative (NO-naproxen) on systemic arterial blood pressure and gastric damage in rats in which hypertension was induced by L-NAME. Rats received either L-NAME dissolved in the drinking water (400 mg/L) or tap water (control). Vehicle, naproxen (10 mg/kg) or an equimolar dose of NO-naproxen (14.5 mg/kg) were administered orally each day. After 4 weeks, blood pressure was measured, blood samples were taken for measurement of thromboxane synthesis, and gastric damage was evaluated by blind, macroscopic scoring. Both naproxen and NO-naproxen inhibited systemic cyclooxygenase activity by >90%. NO-naproxen-treated rats exhibited no significant gastric damage. The gastric damage produced by L-NAME alone was potentiated by naproxen but prevented by NO-naproxen. L-NAME treatment significantly increased blood pressure. In the absence of L-NAME, the naproxen group had significantly higher blood pressure than both the control and NO-naproxen groups. In rats receiving L-NAME, the same conclusions apply, but the concomitant administration of NO-naproxen was able to significantly reduce the blood pressure compared to L-NAME alone. Based on these results, we conclude that NO-naproxen may represent a safer alternative to standard NSAIDs in the treatment of inflammatory conditions in hypertensive patients.

IDrugs. 1998 Nov;1(7):807-12.

Nitroflurbiprofen (NicOx SA)

Wang X. Cornell University Medical College, 450 East 63rd Street, Rm 7G, New York, NY10021, USA.
NicOx is developing nitroflurbiprofen (HCT-1026) as a non-steroidal anti-inflammatory drug (NSAID) which has the ability to release nitric oxide. It has completed phase I clinical trials as a potential treatment for inflammation and rheumatoid arthritis [198694]. In the trial, which took place at Queens Medical Center, Nottingham, UK, the drug showed excellent tolerability, as well as potent and long lasting serum thromboxane inhibition in healthy volunteers after single oral doses of 50 and 100 mg [243679]. A repeated dose endoscopic study showed that nitroflurbiprofen causes less gastrointestinal damage in healthy volunteers than flurbiprofen [265025,295029]. Although phase II studies in patients with musculoskeletal disorders were scheduled for 1997 [243679], it seems they have not yet commenced.The compound is as potent as conventional flurbiprofen, but is better tolerated in rats, dogs and rabbits when given orally or parenterally following either single or repeated doses [198694]. Unlike conventional NSAIDs, nitro-flurbiprofen is able to release NO and increase cGMP in endothelial cells, and to inhibit the expression of inducible nitric oxide synthase and endotoxin in the gastrointestinal tract [190759].

1997 - 02

Aliment Pharmacol Ther. 1997 Feb;11(1):69-79.

NO-naproxen vs. naproxen: ulcerogenic, analgesic and anti-inflammatory effects.

Davies NM, Roseth AG, Appleyard CB, McKnight W, Del Soldato P, Calignano A, Cirino G, Wallace JL. Intestinal Disease Research Unit, Faculty of Medicine, University of Calgary, Alberta, Canada.

BACKGROUND: A novel class of nitric oxide-releasing nonsteroidal anti-inflammatory drug (NO-NSAID) derivatives has recently been described which exert anti-inflammatory activities but produce significantly less gastrointestinal injury than the parent NSAID from which they are derived. The present studies were performed to determine if a nitroxybutylester derivative of naproxen was less ulcerogenic to the gastrointestinal tract than its parent NSAID, and if it exerted comparable analgesic and anti-inflammatory properties to the parent NSAID. METHODS: The two drugs were compared in an acute gastric injury model, an antral ulcer model and after twice-daily administration for 18 days (small intestinal damage model). Anti-inflammatory activity was examined in the carrageenan-induced paw oedema model, while analgesia was examined in the acetic acid-induced writhing model. The pharmacokinetic profiles of naproxen vs. NO-naproxen were compared by HPLC analysis. RESULTS: NO-naproxen was found to produce significantly less gastric damage despite inducing similar increases in plasma TNF alpha to naproxen. With chronic administration, small intestinal damage was markedly less with NO-naproxen than with the parent NSAID. However, NO-naproxen exerted superior analgesic and comparable anti-inflammatory effects to naproxen. NO-naproxen was not completely converted to naproxen, but the reduced plasma levels of the latter was not the underlying reason for reduced gastrointestinal toxicity of NO-naproxen. CONCLUSION: NO-naproxen represents a novel, gastrointestinal-sparing NSAID derivative with superior analgesic and comparable anti-inflammatory properties to naproxen.

1995 - 01

Pharmacol Res. 1995 Jan;31(1):61-5.

Anti-inflammatory potency and gastrointestinal toxicity of a new compound, nitronaproxen.

Cuzzolin L, Conforti A, Adami A, Lussignoli S, Menestrina F, Del Soldato P, Benoni G. Institute of Pharmacology, University of Verona, Italy.

Naproxen and its derivative nitronaproxen at the doses of 5 and 10 mg kg-1 were compared for their acute anti-inflammatory efficacy in a carrageenan oedema model and gastrointestinal toxicity in rats. Moreover, the effects of the two drugs were evaluated in the adjuvant arthritis, after chronic doses of 4 and 8 mg kg-1 administered orally for 18 days. The oedema reduction was maintained much longer (until 5 h) with nitronaproxen; the inhibition of arthritis was 50% or more with both doses of the examined drugs. From the histological examination of the stomachs, an extensive mucosal vasocongestion and haemorrhagic lesions have been observed in some rats treated with naproxen. The percentages of animals with ulcers were 50, 100 and 10 with naproxen 6 and 18 mg kg-1 and nitronaproxen 54 mg kg-1 respectively. A better gastrointestinal tolerability has been observed in arthritic and oedemic rats treated with nitronaproxen compared to naproxen: this could be due to the presence of nitric oxide that acts in maintaining the tissue perfusion and integrity.