RESUMO
Neutrophil elastase (NE) is a crucial marker of neutrophilic inflammation. We aimed to compare different techniques to detect active NE in sputum samples of 50 Bronchiectasis (BE) and 50 Cystic Fibrosis (CF) patients. Three methods including a ProteaseTag® Active NE Immunoassay (ELISA) and two enzymatic digestion assays (chromogenic -CS- and fluorogenic -FS- substrate) were compared. Results of active NE were also correlated with clinical data. The three methods provided statistically different values for NE activity in the same sputum samples in both cohorts. In the BE cohort, the highest correlations between NE activity and Bronchiectasis Severity Index (rhoâ¯=â¯0.40, Pâ¯<â¯0.0001), sputum purulence (AUCâ¯=â¯0.79), and chronic infections due to any pathogen (AUCâ¯=â¯0.76) and P. aeruginosa (AUCâ¯=â¯0.80) were found when NE was measured through the activity-based immunoassay. In the CF cohort, the highest correlations between NE activity and sputum quantity (rhoâ¯=â¯0.71) and FEV1% (rhoâ¯=â¯0.42, Pâ¯=â¯0.03) were observed when the FS method was used, while similar correlations with chronic P. aeruginosa infection were identified with the FS and ELISA methods. NE activity in sputum correlates with clinical variables in both diseases. However, different methods to evaluate active NE in sputum lead to significantly different results, also in terms of correlation with clinical data.
Assuntos
Bronquiectasia/enzimologia , Fibrose Cística/enzimologia , Elastase de Leucócito/metabolismo , Escarro/enzimologia , Adulto , Idoso , Bronquiectasia/fisiopatologia , Estudos de Coortes , Fibrose Cística/fisiopatologia , Feminino , Volume Expiratório Forçado , Humanos , Masculino , Pessoa de Meia-Idade , Estudos Prospectivos , Infecções por Pseudomonas/epidemiologia , Infecções por Pseudomonas/microbiologia , Pseudomonas aeruginosa/isolamento & purificação , Índice de Gravidade de DoençaRESUMO
The role of neutrophil elastase (NE) is poorly understood in bronchiectasis because of the lack of preclinical data and so most of the assumptions made about NE inhibitor potential benefit is based on data from CF. In this context, NE seems to be a predictor of long-term clinical outcomes and a possible target of treatment. In order to better evaluate the role of NE in bronchiectasis, a systematic search of scientific evidence was performed.Two investigators independently performed the search on PubMed and included studies published up to May 15, 2017 according to predefined criteria. A final pool of 31 studies was included in the systematic review, with a total of 2679 patients. For each paper data of interest were extracted and reported in table.In this review sputum NE has proved useful as an inflammatory marker both in stable state bronchiectasis and during exacerbations and local or systemic antibiotic treatment. NE has also been associated with risk of exacerbation, time to next exacerbation and all-cause mortality. This study reviews also the role of NE as a specific target of treatment in bronchiectasis. Inhibition of NE is at a very early stage and future interventional studies should evaluate safety and efficacy for new molecules and formulations.
Assuntos
Bronquiectasia/diagnóstico , Bronquiectasia/enzimologia , Elastase de Leucócito/metabolismo , Biomarcadores/metabolismo , Estudos Transversais , Humanos , Escarro/metabolismoRESUMO
In Duchenne muscular dystrophy (DMD) patients and the mouse model of DMD, mdx, dystrophin deficiency causes a decrease and mislocalization of muscle-specific neuronal nitric oxide synthase (nNOSµ), leading to functional impairments. Previous studies have shown that nitric oxide (NO) donation associated with anti-inflammatory action has beneficial effects in dystrophic mouse models. In this study, we have systematically investigated the effects of naproxcinod, an NO-donating naproxen derivative, on the skeletal and cardiac disease phenotype in mdx mice. Four-week-old mdx and C57BL/10 mice were treated with four different concentrations (0, 10, 21 and 41 mg/kg) of naproxcinod and 0.9 mg/kg of prednisolone in their food for 9 months. All mice were subjected to twice-weekly treadmill sessions, and functional and behavioral parameters were measured at 3, 6 and 9 months of treatment. In addition, we evaluated in vitro force contraction, optical imaging of inflammation, echocardiography and blood pressure (BP) at the 9-month endpoint prior to sacrifice. We found that naproxcinod treatment at 21 mg/kg resulted in significant improvement in hindlimb grip strength and a 30% decrease in inflammation in the fore- and hindlimbs of mdx mice. Furthermore, we found significant improvement in heart function, as evidenced by improved fraction shortening, ejection fraction and systolic BP. In addition, the long-term detrimental effects of prednisolone typically seen in mdx skeletal and heart function were not observed at the effective dose of naproxcinod. In conclusion, our results indicate that naproxcinod has significant potential as a safe therapeutic option for the treatment of muscular dystrophies.
Assuntos
Testes de Função Cardíaca/efeitos dos fármacos , Músculo Esquelético/efeitos dos fármacos , Distrofia Muscular de Duchenne/tratamento farmacológico , Distrofia Muscular de Duchenne/fisiopatologia , Naproxeno/análogos & derivados , Doadores de Óxido Nítrico/administração & dosagem , Animais , Anti-Inflamatórios/administração & dosagem , Anti-Inflamatórios/uso terapêutico , Peso Corporal/efeitos dos fármacos , Modelos Animais de Doenças , Relação Dose-Resposta a Droga , Membro Posterior/efeitos dos fármacos , Camundongos , Camundongos Endogâmicos C57BL , Camundongos Endogâmicos mdx , Músculo Esquelético/fisiopatologia , Distrofia Muscular de Duchenne/patologia , Naproxeno/administração & dosagem , Naproxeno/uso terapêutico , Doadores de Óxido Nítrico/uso terapêutico , Prednisolona/administração & dosagem , Prednisolona/uso terapêuticoRESUMO
Clinical guidelines for COPD and asthma recommend inhaled ß-adrenergic agonists, muscarinic antagonists, and, for frequent exacerbators, inhaled corticosteroids, with the challenge of combining them into a single device. The MABA (muscarinic antagonist and ß2 agonist) concept has the potential to simplify this complexity while increasing the efficacy of both pharmacologies. In this article, we report the outcome of our solid-state driven back-up program that led to the discovery of the MABA compound CHF-6550. A soft drug approach was applied, aiming at high plasma protein binding and high hepatic clearance, concurrently with an early stage assessment of crystallinity through a dedicated experimental workflow. A new chemotype was identified, the diphenyl hydroxyacetic esters, able to generate crystalline material. Among this class, CHF-6550 demonstrated in vivo efficacy, suitability for dry powder inhaler development, favorable pharmacokinetics, and safety in preclinical settings and was selected as a back-up candidate, fulfilling the desired pharmacological and solid-state profile.
Assuntos
Agonistas de Receptores Adrenérgicos beta 2 , Antagonistas Muscarínicos , Antagonistas Muscarínicos/farmacocinética , Antagonistas Muscarínicos/farmacologia , Antagonistas Muscarínicos/química , Antagonistas Muscarínicos/síntese química , Antagonistas Muscarínicos/uso terapêutico , Antagonistas Muscarínicos/administração & dosagem , Animais , Humanos , Agonistas de Receptores Adrenérgicos beta 2/farmacocinética , Agonistas de Receptores Adrenérgicos beta 2/farmacologia , Agonistas de Receptores Adrenérgicos beta 2/química , Agonistas de Receptores Adrenérgicos beta 2/administração & dosagem , Administração por Inalação , Ratos , Descoberta de Drogas , Relação Estrutura-Atividade , Masculino , Doença Pulmonar Obstrutiva Crônica/tratamento farmacológicoRESUMO
INTRODUCTION: Myopathy is the most common side effect of statins. Because nitric oxide (NO) has a key role in regulating skeletal muscle function, we studied whether the NO-donating atorvastatin NCX 6560 could show a better profile on skeletal muscle function and structure compared with atorvastatin. METHODS: C57BL/6 mice received atorvastatin 40 mg/kg/day or an equivalent dose of NCX 6560 for 2 months. Muscle function assessed by treadmill test, serum creatine kinase (CK) activity, citrate synthase (CS) activity, and muscle histology were evaluated. RESULTS: Atorvastatin significantly (P < 0.001) reduced muscle endurance, increased serum CK by 6-fold, and induced muscle fiber atrophy. Conversely, NCX 6560 preserved muscle function, prevented CK increase and did not modify muscle structure. Interestingly, atorvastatin reduced CS activity, a marker for mitochondrial function, in gastrocnemius, diaphragm, and heart, whereas NCX 6560 prevented such decrease. CONCLUSIONS: These findings suggest that NO may prevent statin-induced myopathy.
Assuntos
Ácidos Heptanoicos/farmacologia , Inibidores de Hidroximetilglutaril-CoA Redutases/farmacologia , Músculo Esquelético/efeitos dos fármacos , Doadores de Óxido Nítrico/farmacologia , Pirróis/farmacologia , Animais , Atorvastatina , Atrofia , Citrato (si)-Sintase/metabolismo , Creatina Quinase/metabolismo , L-Lactato Desidrogenase/metabolismo , Masculino , Camundongos , Camundongos Endogâmicos C57BL , Músculo Esquelético/metabolismo , Músculo Esquelético/patologia , Cadeias Pesadas de Miosina/metabolismo , Óxido Nítrico/metabolismoRESUMO
PURPOSE: Polymorphonuclear neutrophils, the first leukocytes to infiltrate the inflamed tissue, can make important contributions to vascular inflammatory processes driving the development of atherosclerosis. We herein investigated the effects of atorvastatin and NCX 6560 (a nitric oxide (NO)-donating atorvastatin derivative that has completed a successful phase 1b study) on neutrophilic inflammation in carotid arteries of normocholesterolemic rabbits subjected to perivascular collar placement. METHODS: Atorvastatin or NCX 6560 were administered orally (5 mg/kg/day or equimolar dose) to New Zealand White rabbits for 6 days, followed by collar implantation 1 h after the last dose. Twenty-four hours later carotids were harvested for neutrophil quantification by immunostaining. RESULTS: Treatment with NCX 6560 was associated with a lower neutrophil infiltration (-39.5 %), while atorvastatin did not affect neutrophil content. The result was independent of effects on plasma cholesterol or differences in atorvastatin bioavailability, which suggests an important role of NO-related mechanisms in mediating this effect. Consistent with these in vivo findings, in vitro studies showed that NCX 6560, as compared to atorvastatin, had greater inhibitory activity on processes involved in neutrophil recruitment, such as migration in response to IL-8 and IL-8 release by endothelial cells and by neutrophils themselves. Pretreatment with NCX 6560, but not with atorvastatin, reduced the ability of neutrophil supernatants to promote monocyte chemotaxis, a well-known pro-inflammatory activity of neutrophils. CONCLUSION: Experimental data suggest a potential role of NO-releasing statins in the control of the vascular inflammatory process mediated by polymorphonuclear neutrophils.
Assuntos
Aterosclerose/prevenção & controle , Colesterol/sangue , Ácidos Heptanoicos/uso terapêutico , Infiltração de Neutrófilos/efeitos dos fármacos , Doadores de Óxido Nítrico/uso terapêutico , Pirróis/uso terapêutico , Doença Aguda , Animais , Aterosclerose/sangue , Aterosclerose/imunologia , Aterosclerose/patologia , Atorvastatina , Artérias Carótidas/efeitos dos fármacos , Artérias Carótidas/patologia , Sobrevivência Celular/efeitos dos fármacos , Quimiotaxia de Leucócito/efeitos dos fármacos , Quimiotaxia de Leucócito/imunologia , Ácidos Heptanoicos/administração & dosagem , Ácidos Heptanoicos/farmacocinética , Ácidos Heptanoicos/farmacologia , Células Endoteliais da Veia Umbilical Humana , Humanos , Interleucina-8/imunologia , Masculino , Estrutura Molecular , Monócitos/citologia , Monócitos/efeitos dos fármacos , Infiltração de Neutrófilos/imunologia , Neutrófilos/citologia , Neutrófilos/efeitos dos fármacos , Doadores de Óxido Nítrico/administração & dosagem , Doadores de Óxido Nítrico/farmacocinética , Doadores de Óxido Nítrico/farmacologia , Pirróis/administração & dosagem , Pirróis/farmacocinética , Pirróis/farmacologia , CoelhosRESUMO
Background: Idiopathic pulmonary fibrosis (IPF) is a chronic, progressive, age-related interstitial lung disease (ILD) with limited therapeutic options. Despite the wide variety of different in vivo models for IPF, these preclinical models have shown limitations that may significantly impair their translational potential. Among the most relevant limitations are the methodologies used to assess the efficacy of anti-fibrotic treatments, that are not the ones used in humans. In this scenario, the goal of the work presented in this paper is to provide translational relevance to the bleomycin (BLM)-induced pulmonary fibrosis mouse model, introducing and validating novel readouts to evaluate the efficacy of treatments for IPF. Methods: The BLM model was optimized by introducing the use of functional assessments such as the Forced Vital Capacity (FVC) and the Diffusion Factor for Carbon Monoxide (DFCO), that are respectively the primary and secondary endpoints in clinical trials for IPF, comparing them to more common readouts such as lung histology, improved by the application of Artificial Intelligence (AI) to detect and quantify fibrotic tissue deposition, and metalloproitenase-7 (MMP-7), a clinical prognostic biomarker. Results: Lung function measurement and DFCO changes well correlated with Ashcroft score, the current gold-standard for the assessment of pulmonary fibrosis in mice. The relevance and robustness of these novel readouts in the BLM model was confirmed by the results obtained testing Nintedanib and Pirfenidone, the only drugs approved for the treatment of IPF patients: in fact, both drugs administered therapeutically, significantly affected the changes in these parameters induced by BLM treatment, with results that closely reflected the efficacy observed in the clinic. Changes in biomarkers such as MMP-7 were also evaluated, and well correlated with the modifications of FVC and DFCO. Conclusion: Novel functional readouts such as FVC and DFCO can be efficiently used to assess pathology progression in the BLM-induced pulmonary fibrosis mouse model as well as compound efficacy, substantially improving its translational and predictivity potential.
RESUMO
The development of molecules embedding two distinct pharmacophores acting as muscarinic antagonists and ß2 agonists (MABAs) promises to be an excellent opportunity to reduce formulation issues and boost efficacy through cross-talk and allosteric interactions. Herein, we report the results of our drug discovery campaign aimed at improving the therapeutic index of a previous MABA series by exploiting the super soft-drug concept. The incorporation of a metabolic liability, stable at the site of administration but undergoing rapid systemic metabolism, to generate poorly active and quickly eliminated fragments was pursued. Our SAR studies yielded MABA 29, which demonstrated a balanced in vivo profile up to 24 h, high instability in plasma and the liver, as well as sustained exposure in the lung. In vitro safety and non-GLP toxicity studies supported the nomination of 29 (CHF-6366) as a clinical candidate, attesting to the successful development of a novel super-soft MABA compound.
Assuntos
Antagonistas Muscarínicos , Doença Pulmonar Obstrutiva Crônica , Administração por Inalação , Agonistas de Receptores Adrenérgicos beta 2/farmacologia , Agonistas de Receptores Adrenérgicos beta 2/uso terapêutico , Broncodilatadores/uso terapêutico , Descoberta de Drogas , Humanos , Pulmão , Antagonistas Muscarínicos/farmacologia , Antagonistas Muscarínicos/uso terapêutico , Doença Pulmonar Obstrutiva Crônica/tratamento farmacológicoRESUMO
A resolutive therapy for muscular dystrophies, a heterogeneous group of genetic diseases leading to muscular degeneration and in the severe forms to death, is still lacking. Since inflammation and defects in nitric oxide generation are recognized key pathogenic events in muscular dystrophy, we have analysed the effects of a derivative of ibuprofen, NCX 320, belonging to the class of cyclooxygenase inhibiting nitric oxide donator (CINOD), in the α-sarcoglycan null mice, a severe mouse model of dystrophy. NCX 320 was administered daily in the diet for 8months starting 1month from weaning. Muscle functional recovery was evaluated by free wheel and treadmill tests at 8months. Serum creatine kinase activity, as well as the number of diaphragm inflammatory infiltrates and necrotic fibres, was measured as indexes of skeletal muscle damage. Muscle regeneration was evaluated in diaphragm and tibialis anterior muscles, measuring the numbers of centronucleated fibres and of myogenic precursor cells. NCX 320 mitigated muscle damage, reducing significantly serum creatine kinase activity, the number of necrotic fibres and inflammatory infiltrates. Moreover, NCX 320 stimulated muscle regeneration increasing significantly the number of myogenic precursor cells and regenerating fibres. All these effects concurred in inducing a significant improvement of muscle function, as assessed by both free wheel and treadmill tests. These results describe the properties of a new compound incorporating nitric oxide donation together with anti-inflammatory properties, showing that it is effective in slowing muscle dystrophy progression long term. Of importance, this new compound deserves specific attention for its potential in the therapy of muscular dystrophy given that ibuprofen is well tolerated in paediatric patients and with a profile of safety that makes it suitable for chronic treatment such as the one required in muscular dystrophies.
Assuntos
Inibidores de Ciclo-Oxigenase/uso terapêutico , Ibuprofeno/análogos & derivados , Distrofia Muscular Animal/tratamento farmacológico , Óxido Nítrico/metabolismo , Animais , Linhagem Celular , Inibidores de Ciclo-Oxigenase/farmacologia , Deleção de Genes , Ibuprofeno/farmacologia , Ibuprofeno/uso terapêutico , Camundongos , Músculo Esquelético/efeitos dos fármacos , Músculo Esquelético/patologia , Distrofia Muscular Animal/patologia , Prostaglandina-Endoperóxido Sintases/metabolismo , Sarcoglicanas/genéticaRESUMO
Novel bi-functional compounds with a nitric oxide (NO)-releasing moiety bound to a dorzolamide scaffold were investigated. Several compounds were synthesized and their activity as selective carbonic anhydrase inhibitors (CAI) evaluated in vitro on recombinant hCA type I, II and IV enzyme isoforms where they showed different degrees of potency and selectivity to hCA II. A high resolution X-ray crystal structure for the CA II adduct with 8 confirmed the high affinity of this class of compounds for the enzyme. Compounds 4, 6, and 8 showed highly potent and efficacious NO-mediated properties as assessed by their vascular relaxant effect on methoxamine-precontracted rabbit aortic rings. Finally, compounds 4 and 6 exerted potent intraocular pressure (IOP) lowering effects in vivo in normotensive rabbits thereby anticipating their potential for the treatment of hypertensive glaucoma.
Assuntos
Anidrase Carbônica II/antagonistas & inibidores , Inibidores da Anidrase Carbônica/uso terapêutico , Glaucoma de Ângulo Aberto/tratamento farmacológico , Óxido Nítrico/química , Sulfonamidas/uso terapêutico , Tiofenos/uso terapêutico , Animais , Inibidores da Anidrase Carbônica/síntese química , Inibidores da Anidrase Carbônica/química , Cristalografia por Raios X , Descoberta de Drogas , Modelos Moleculares , Estrutura Molecular , Coelhos , Relação Estrutura-Atividade , Sulfonamidas/síntese química , Sulfonamidas/química , Tiofenos/síntese química , Tiofenos/químicaRESUMO
A novel class of timolol derivatives with nitric oxide (NO)-donating moieties achieved chemical stability yet under physiologically relevant conditions released timolol and NO. Hindered esters A were designed and synthesized, whose 'triggered' release relied on enzymatic hydrolysis of the nitrate ester in A to B, that in turn cyclized to liberate timolol.
Assuntos
Óxido Nítrico/metabolismo , Timolol/química , Antagonistas Adrenérgicos beta/metabolismo , Estabilidade de Medicamentos , Ésteres/química , Ésteres/metabolismo , Guanilato Ciclase/antagonistas & inibidores , Guanilato Ciclase/metabolismoRESUMO
NO mediates a variety of physiologic processes and is considered an important intracellular messenger in different cellular systems. Because of its complex regulation and multiple molecular and cellular targets, NO provides both stimulatory and suppressive properties in the immune system. Dendritic cells (DCs) are considered the most potent APCs, whose regulation has important implications in the induction of an effective immune response. In this study, we analyzed the effect of the compound NCX 2057, a new class of NO-releasing derivatives of ferulic acid, on activation and functional properties of DCs. NCX 2057 was able to modulate the inflammatory program, the cytokines production, and the cellular life cycle but not the maturation markers and the T cells stimulatory capacity of DCs in the presence or absence of LPS. The results indicate that NCX 2057 may modulate different aspects of the activation of DCs and suggest novel applications of NO donors in the contest of inflammatory response modulation through the life cycle regulation of DCs.
Assuntos
Anti-Inflamatórios/farmacologia , Células Dendríticas/citologia , Células Dendríticas/metabolismo , Inflamação/patologia , Doadores de Óxido Nítrico/farmacologia , Animais , Apresentação de Antígeno/efeitos dos fármacos , Apoptose/efeitos dos fármacos , Células da Medula Óssea/citologia , Butanos/farmacologia , Citocinas/biossíntese , Células Dendríticas/efeitos dos fármacos , Dinitrato de Isossorbida/análogos & derivados , Dinitrato de Isossorbida/farmacologia , Lipopolissacarídeos/farmacologia , Camundongos Endogâmicos C57BL , Inibidor de NF-kappaB alfa/metabolismo , Óxido Nítrico Sintase Tipo II/metabolismo , Nitritos/metabolismo , Nitrocompostos/farmacologia , Proteólise/efeitos dos fármacosRESUMO
AIMS: Cyclooxygenase (COX)-inhibiting nitric oxide donors (CINODs) are a new class of drugs that structurally combine a COX inhibitor with a nitric oxide (NO) donating moiety. This combination reduces potential toxicity of the non-steroidal anti-inflammatory drugs (NSAIDs) whilst maintaining the analgesic and anti-inflammatory effects. The present study was undertaken to investigate the anti-inflammatory effects of NCX 429, a naproxen-based CINOD, and to assess the additional properties of NO donation beyond those related to naproxen. MAIN METHODS: We evaluated the in vitro effects of NCX 429 on oxy-radical production, phagocytosis, cytokine release, MMP-9, PPARγ expression and NF-κB activation in human monocytes/MDM and compared to naproxen. Moreover, we compared the in vivo efficacy of NCX 429 and naproxen in a murine model of peritonitis. KEY FINDINGS: In all the experiments performed in vitro, NCX 429 reduced the inflammatory responses with equal or higher efficacy compared to naproxen. Moreover, in in vivo experiments, NCX 429, at the lowest dose tested, was able to significantly inhibit cell influx in response to IL-1ß administration although naproxen was found to be more potent than NCX 429 at reducing PGE2 in inflammatory exudates. SIGNIFICANCE: These results demonstrate that both in vitro and in vivo--in a murine model of peritonitis--NCX 429 elicits significant anti-inflammatory activity, beyond the simple COX inhibition or pure NO release. Therefore, NO donation along with COX inhibition may represent a strategy for investigating inflammatory diseases in which pain and function are not fully resolved by analgesics/anti-inflammatory drugs.
Assuntos
Anti-Inflamatórios não Esteroides , Naproxeno/análogos & derivados , Nitratos , Doadores de Óxido Nítrico , Óxido Nítrico/farmacocinética , Peritonite , Animais , Anti-Inflamatórios não Esteroides/farmacocinética , Anti-Inflamatórios não Esteroides/farmacologia , Dinoprostona/metabolismo , Modelos Animais de Doenças , Avaliação Pré-Clínica de Medicamentos , Humanos , Interleucina-1beta/metabolismo , Metaloproteinase 9 da Matriz/metabolismo , Camundongos , NF-kappa B/metabolismo , Naproxeno/farmacocinética , Naproxeno/farmacologia , Nitratos/farmacocinética , Nitratos/farmacologia , Doadores de Óxido Nítrico/farmacocinética , Doadores de Óxido Nítrico/farmacologia , Peritonite/tratamento farmacológico , Peritonite/metabolismo , Peritonite/patologia , Fagocitose/efeitos dos fármacosRESUMO
BACKGROUND: In dystrophin-deficient muscles of Duchenne Muscular Dystrophy (DMD) patients and the mdx mouse model, nitric oxide (NO) signalling is impaired. Previous studies have shown that NO-donating drugs are beneficial in dystrophic mouse models. Recently, a long-term treatment (9 months) of mdx mice with naproxcinod, an NO-donating naproxen, has shown a significant improvement of the dystrophic phenotype with beneficial effects present throughout the disease progression. It remains however to be clearly dissected out which specific effects are due to the NO component compared with the anti-inflammatory activity associated with naproxen. Understanding the contribution of NO vs the anti-inflammatory effect is important, in view of the potential therapeutic perspective, and this is the final aim of this study. METHODS: Five-week-old mdx mice received either naproxcinod (30 mg/kg) or the equimolar dose of naproxen (20 mg/kg) in the diet for 6 months. Control mdx mice were used as reference. Treatments (or vehicle for control groups) were administered daily in the diet. For the first 3 months the study was performed in sedentary animals, then all mice were subjected to exercise until the sixth month. Skeletal muscle force was assessed by measuring whole body tension in sedentary animals as well as in exercised mice and resistance to fatigue was measured after 3 months of running exercise. At the end of 6 months of treatment, animals were sacrificed for histological analysis and measurement of naproxen levels in blood and skeletal muscle. RESULTS: Naproxcinod significantly ameliorated skeletal muscle force and resistance to fatigue in sedentary as well as in exercised mice, reduced inflammatory infiltrates and fibrosis deposition in both cardiac and diaphragm muscles. Conversely, the equimolar dose of naproxen showed no effects on fibrosis and improved muscle function only in sedentary mice, while the beneficial effects in exercised mice were lost demonstrating a limited and short-term effect. CONCLUSION: In conclusion, this study shows that NO donation may have an important role, in addition to anti-inflammatory activity, in slowing down the progression of the disease in the mdx mouse model therefore positioning naproxcinod as a promising candidate for treatment of DMD.
Assuntos
Distrofia Muscular de Duchenne/tratamento farmacológico , Naproxeno/análogos & derivados , Naproxeno/uso terapêutico , Doadores de Óxido Nítrico/uso terapêutico , Animais , Disponibilidade Biológica , Modelos Animais de Doenças , Masculino , Camundongos , Camundongos Endogâmicos C57BL , Camundongos Endogâmicos mdx , Distrofia Muscular de Duchenne/fisiopatologia , Naproxeno/farmacocinética , Doadores de Óxido Nítrico/farmacocinéticaRESUMO
AIMS: The aim of the present study was to assess whether the addition of a nitric oxide (NO)-donating moiety to atorvastatin enhances anti-inflammatory and anti-atherogenic effects in an animal model of endothelial dysfunction, systemic peroxidation and inflammation, and accelerated atherosclerosis. METHODS AND RESULTS: Low-density lipoprotein receptor (LDLR)(-/-) mice kept on a high-fat diet (HFD) for 16 weeks underwent photochemical injury to the femoral artery with the local production of oxygen radicals. HFD markedly enhanced cholesterol, inflammatory biomarkers in plasma and in the femoral arterial wall, and atherosclerotic lesions in the aortic arch; inflammation and atherosclerosis were further increased by photochemically generated oxygen radicals. Treatment with the NO-donating atorvastatin NCX 6560 (11.7 mg/kg) was significantly more effective than atorvastatin (10 mg/kg) in reducing the following parameters: lipid-rich lesions in the aortic arch (surface covered: atorvastatin = 24 ± 5%; NCX 6560 = 14.7 ± 3.9%; P< 0.05); the production of radical oxygen species in the aorta (dichlorofluorescein fluorescence intensity per milligram of protein: atorvastatin = 2419 ± 136.7; NCX 6560 = 1766 ± 161.2; P< 0.05); femoral artery intima/media thickness (atorvastatin = 1.2 ± 0.11; NCX 6560 = 0.3 ± 0.14; P< 0.05); circulating interleukin-6 (atorvastatin = 34.3 ± 6.8 pg/mL; NCX 6560 = 17.7 ± 14.4 pg/mL; P< 0.05); and matrix metalloproteinase 2 in the arterial wall (atorvastatin = 55.2 ± 1.9 ng/µg of proteins; NCX 6560 = 45.8 ± 2.6 ng/µg of proteins; P < 0.05). CONCLUSION: In conditions of severe endothelial dysfunction, systemic peroxidation and inflammation, and accelerated atherosclerosis, atorvastatin, even at high doses, displays suboptimal anti-atherogenic and anti-inflammatory effects, while the addition of a NO-donating property confers enhanced anti-atherogenic and anti-inflammatory effects.
Assuntos
Anti-Inflamatórios/farmacologia , Aterosclerose/metabolismo , Ácidos Heptanoicos/farmacologia , Óxido Nítrico/metabolismo , Pirróis/farmacologia , Animais , Anticolesterolemiantes/farmacologia , Aterosclerose/tratamento farmacológico , Atorvastatina , Modelos Animais de Doenças , Inflamação/tratamento farmacológico , Inflamação/metabolismo , Camundongos , Camundongos Endogâmicos C57BL , Camundongos Knockout , Receptores de LDL/deficiênciaRESUMO
PURPOSE: Hypoxia of the retina and optic nerve head (ONH) is believed to be pivotal in the development of ocular vascular disorders, including diabetic macular edema (DME). Glucocorticoids are among the most effective agents for the treatment of back of the eye diseases. However, this class of compounds is highly liable to increase intraocular pressure (IOP) and does not improve ocular perfusion or tissue oxygenation. Nitric oxide (NO) has vasodilating properties and lowers IOP in experimental models and humans, suggesting that its properties might complement those of glucocorticoids. NCX 434 is an NO-donating triamcinolone acetonide (TA) that is less likely to increase IOP while targeting both the vascular and inflammatory components of DME. METHODS: NCX 434 was studied in vitro with respect to its NO-releasing properties in isolated methoxamine-precontracted rabbit aortic rings and glucocorticoid-like activity in recombinant human glucocorticoid receptors. IOP and oxygen saturation in the ONH and overlaying arteries and veins were studied in the anesthetized cynomolgus monkey. Measurements were taken using, respectively, an applanation tonometer and a hyperspectral imaging system before and 7, 14, 21, 31 and 41 days after the intravitreal injection of NCX 434 (5.8 mg/eye) or TA equimolar doses (4.0 mg/eye). RESULTS: NCX 434 inhibited (3)H-dexamethasone-specific binding (IC(50)=34±5 nM) on human glucocorticoid receptors and elicited NO-dependent aortic ring relaxation (EC(50) of 0.5±0.1 µM, E(max) 98.9%). In monkey eyes, NCX 434 enhanced, whereas TA did not, oxygen saturation in various ONH areas (*P<0.05 vs. basal), decreased it in veins, and did not affect it in the overlaying arteries. Neither NCX 434 nor TA altered IOP significantly at all time points. However, at 31 days post-treatment TA appeared to start increasing IOP (Δ(IOP)=+3.31±0.51 mmHg, 30.8%, over baseline, NS). CONCLUSIONS: NCX 434 enhances ocular tissue oxygenation. This feature appears to depend on its NO-donating properties; thus, the compound deserves to be further investigated for the treatment of DME and other ocular disorders with impaired ocular perfusion.