ABSTRACT
BACKGROUND: S-nitrosoglutathione (GSNO) serves as a reservoir for nitric oxide (NO) and thus is a key homeostatic regulator of airway smooth muscle tone and inflammation. Decreased levels of GSNO in the lungs of asthmatics have been attributed to increased GSNO catabolism via GSNO reductase (GSNOR) leading to loss of GSNO- and NO- mediated bronchodilatory and anti-inflammatory actions. GSNOR inhibition with the novel small molecule, N6022, was explored as a therapeutic approach in an experimental model of asthma. METHODS: Female BALB/c mice were sensitized and subsequently challenged with ovalbumin (OVA). Efficacy was determined by measuring both airway hyper-responsiveness (AHR) upon methacholine (MCh) challenge using whole body plethysmography and pulmonary eosinophilia by quantifying the numbers of these cells in the bronchoalveolar lavage fluid (BALF). Several other potential biomarkers of GSNOR inhibition were measured including levels of nitrite, cyclic guanosine monophosphate (cGMP), and inflammatory cytokines, as well as DNA binding activity of nuclear factor kappa B (NFκB). The dose response, onset of action, and duration of action of a single intravenous dose of N6022 given from 30 min to 48 h prior to MCh challenge were determined and compared to effects in mice not sensitized to OVA. The direct effect of N6022 on airway smooth muscle tone also was assessed in isolated rat tracheal rings. RESULTS: N6022 attenuated AHR (ED50 of 0.015 ± 0.002 mg/kg; Mean ± SEM) and eosinophilia. Effects were observed from 30 min to 48 h after treatment and were comparable to those achieved with three inhaled doses of ipratropium plus albuterol used as the positive control. N6022 increased BALF nitrite and plasma cGMP, while restoring BALF and plasma inflammatory markers toward baseline values. N6022 treatment also attenuated the OVA-induced increase in NFκB activation. In rat tracheal rings, N6022 decreased contractile responses to MCh. CONCLUSIONS: The significant bronchodilatory and anti-inflammatory actions of N6022 in the airways are consistent with restoration of GSNO levels through GSNOR inhibition. GSNOR inhibition may offer a therapeutic approach for the treatment of asthma and other inflammatory lung diseases. N6022 is currently being evaluated in clinical trials for the treatment of inflammatory lung disease.
Subject(s)
Aldehyde Oxidoreductases/antagonists & inhibitors , Asthma/drug therapy , Benzamides/pharmacology , Benzamides/therapeutic use , Bronchoconstriction/drug effects , Inflammation/prevention & control , Pyrroles/pharmacology , Pyrroles/therapeutic use , Animals , Asthma/immunology , Asthma/physiopathology , Female , Mice , Mice, Inbred BALB CABSTRACT
Nitric oxide (NO) exerts a wide range of cellular effects in the cardiovascular system. NO is short lived, but S-nitrosoglutathione (GSNO) functions as a stable intracellular bioavailable NO pool. Accordingly, increased levels can facilitate NO-mediated processes, and conversely, catabolism of GSNO by the regulatory enzyme GSNO reductase (GSNOR) can impair these processes. Because dysregulated GSNOR can interfere with processes relevant to cardiovascular health, it follows that inhibition of GSNOR may be beneficial. However, the effect of GSNOR inhibition on vascular activity is unknown. To study the effects of GSNOR inhibition on endothelial function, we treated rats with a small-molecule inhibitor of GSNOR (N6338) that has vasodilatory effects on isolated aortic rings and assessed effects on arterial flow-mediated dilation (FMD), an NO-dependent process. GSNOR inhibition with a single intravenous dose of N6338 preserved FMD (15.3 ± 5.4 vs. 14.2 ± 6.3%, P = nonsignificant) under partial NO synthase inhibition that normally reduces FMD by roughly 50% (14.1 ± 2.9 vs. 7.6 ± 4.4%, P < 0.05). In hypertensive rats, daily oral administration of N6338 for 14 days reduced blood pressure (170.0 ± 5.3/122.7 ± 6.4 vs. 203.8 ± 1.9/143.7 ± 7.5 mmHg for vehicle, P < 0.001) and vascular resistance index (1.5 ± 0.4 vs. 3.2 ± 1.0 mmHg · min · l(-1) for vehicle, P < 0.001), and restored FMD from an initially impaired state (7.4 ± 1.7%, day 0) to a level (13.0 ± 3.1%, day 14, P < 0.001) similar to that observed in normotensive rats. N6338 also reversed the pathological kidney changes exhibited by the hypertensive rats. GSNOR inhibition preserves FMD under conditions of impaired NO production and protects against both microvascular and conduit artery dysfunction in a model of hypertension.
Subject(s)
Aldehyde Oxidoreductases/antagonists & inhibitors , Antihypertensive Agents/pharmacology , Endothelium, Vascular/drug effects , Enzyme Inhibitors/pharmacology , Femoral Artery/drug effects , Hypertension/drug therapy , Vasodilation/drug effects , Vasodilator Agents/pharmacology , Administration, Oral , Aldehyde Oxidoreductases/metabolism , Animals , Antihypertensive Agents/administration & dosage , Blood Pressure/drug effects , Cells, Cultured , Disease Models, Animal , Dose-Response Relationship, Drug , Endothelium, Vascular/enzymology , Endothelium, Vascular/physiopathology , Enzyme Inhibitors/administration & dosage , Femoral Artery/enzymology , Femoral Artery/physiopathology , Humans , Hypertension/enzymology , Hypertension/etiology , Hypertension/pathology , Hypertension/physiopathology , Injections, Intravenous , Kidney/drug effects , Kidney/pathology , Mice , Nitric Oxide/metabolism , Nitric Oxide Synthase Type III/antagonists & inhibitors , Nitric Oxide Synthase Type III/metabolism , Rats , Rats, Inbred Dahl , Rats, Sprague-Dawley , Sodium Chloride, Dietary , Time Factors , Vasodilator Agents/administration & dosageABSTRACT
The enzyme S-nitrosoglutathione reductase (GSNOR) is a member of the alcohol dehydrogenase family (ADH) that regulates the levels of S-nitrosothiols (SNOs) through catabolism of S-nitrosoglutathione (GSNO). GSNO and SNOs are implicated in the pathogenesis of many diseases including those in respiratory, gastrointestinal, and cardiovascular systems. The pyrrole based N6022 was recently identified as a potent, selective, reversible, and efficacious GSNOR inhibitor which is currently in clinical development for acute asthma. We describe here the synthesis and structure-activity relationships (SAR) of novel pyrrole based analogs of N6022 focusing on carboxamide modifications on the pendant N-phenyl moiety. We have identified potent and novel GSNOR inhibitors that demonstrate efficacy in an ovalbumin (OVA) induced asthma model in mice.
Subject(s)
Aldehyde Oxidoreductases/antagonists & inhibitors , Anti-Asthmatic Agents/chemical synthesis , Asthma/drug therapy , Benzamides/chemical synthesis , Enzyme Inhibitors/chemical synthesis , Pyrroles/chemical synthesis , Acute Disease , Aldehyde Oxidoreductases/metabolism , Animals , Anti-Asthmatic Agents/administration & dosage , Anti-Asthmatic Agents/therapeutic use , Asthma/chemically induced , Asthma/enzymology , Benzamides/administration & dosage , Benzamides/therapeutic use , Enzyme Inhibitors/administration & dosage , Enzyme Inhibitors/therapeutic use , Humans , Male , Mice , Ovalbumin , Pyrroles/administration & dosage , Pyrroles/therapeutic use , S-Nitrosoglutathione/metabolism , S-Nitrosothiols/metabolism , Structure-Activity RelationshipABSTRACT
The pyrrole based N6022 was recently identified as a potent, selective, reversible, and efficacious S-nitrosoglutathione reductase (GSNOR) inhibitor and is currently undergoing clinical development for the treatment of acute asthma. GSNOR is a member of the alcohol dehydrogenase family (ADH) and regulates the levels of S-nitrosothiols (SNOs) through catabolism of S-nitrosoglutathione (GSNO). Reduced levels of GSNO, as well as other nitrosothiols (SNOs), have been implicated in the pathogenesis of many diseases including those of the respiratory, cardiovascular, and gastrointestinal systems. Preservation of endogenous SNOs through GSNOR inhibition presents a novel therapeutic approach with broad applicability. We describe here the synthesis and structure-activity relationships (SAR) of novel pyrrole based analogues of N6022 focusing on removal of cytochrome P450 inhibition activities. We identified potent and novel GSNOR inhibitors having reduced CYP inhibition activities and demonstrated efficacy in a mouse ovalbumin (OVA) model of asthma.
Subject(s)
Aldehyde Oxidoreductases/antagonists & inhibitors , Benzamides/pharmacology , Cytochrome P-450 Enzyme System/metabolism , Enzyme Inhibitors/chemical synthesis , Enzyme Inhibitors/pharmacology , Imidazoles/chemical synthesis , Imidazoles/pharmacology , Pyrroles/pharmacology , Animals , Asthma/drug therapy , Asthma/enzymology , Benzamides/chemistry , Benzamides/toxicity , Cytochrome P-450 Enzyme Inhibitors , Disease Models, Animal , Dose-Response Relationship, Drug , Drug Administration Schedule , Drug Design , Drug Evaluation, Preclinical , Enzyme Inhibitors/pharmacokinetics , Enzyme Inhibitors/toxicity , Humans , Imidazoles/pharmacokinetics , Imidazoles/toxicity , Lung/pathology , Lung/physiopathology , Mice , Molecular Structure , Molecular Targeted Therapy , No-Observed-Adverse-Effect Level , Pyrroles/chemistry , Pyrroles/toxicity , Receptors, Opioid, delta/metabolism , S-Nitrosoglutathione/metabolism , Structure-Activity RelationshipABSTRACT
S-Nitrosoglutathione reductase (GSNOR) is a member of the alcohol dehydrogenase family (ADH) that regulates the levels of S-nitrosothiols (SNOs) through catabolism of S-nitrosoglutathione (GSNO). GSNO and SNOs are implicated in the pathogenesis of many diseases including those in respiratory, cardiovascular, and gastrointestinal systems. The pyrrole based N6022 was recently identified as a potent, selective, reversible, and efficacious GSNOR inhibitor which is currently undergoing clinical development. We describe here the synthesis and structure-activity relationships (SAR) of novel pyrrole based analogues of N6022 focusing on scaffold modification and propionic acid replacement. We identified equally potent and novel GSNOR inhibitors having pyrrole regioisomers as scaffolds using a structure based approach.
Subject(s)
Aldehyde Oxidoreductases/antagonists & inhibitors , Benzamides/chemistry , Benzamides/pharmacology , Enzyme Inhibitors/chemistry , Enzyme Inhibitors/pharmacology , Propionates/chemistry , Propionates/pharmacology , Pyrroles/chemistry , Pyrroles/pharmacology , Benzamides/chemical synthesis , Enzyme Activation/drug effects , Enzyme Inhibitors/chemical synthesis , Inhibitory Concentration 50 , Molecular Structure , Propionates/chemical synthesis , Pyrroles/chemical synthesis , Stereoisomerism , Structure-Activity RelationshipABSTRACT
S-Nitrosoglutathione reductase (GSNOR) regulates S-nitrosothiols (SNOs) and nitric oxide (NO) in vivo through catabolism of S-nitrosoglutathione (GSNO). GSNOR and the anti-inflammatory and smooth muscle relaxant activities of SNOs, GSNO, and NO play significant roles in pulmonary, cardiovascular, and gastrointestinal function. In GSNOR knockout mice, basal airway tone is reduced and the response to challenge with bronchoconstrictors or airway allergens is attenuated. Consequently, GSNOR has emerged as an attractive therapeutic target for several clinically important human diseases. As such, small molecule inhibitors of GSNOR were developed. These GSNOR inhibitors were potent, selective, and efficacious in animal models of inflammatory disease characterized by reduced levels of GSNO and bioavailable NO. N6022, a potent and reversible GSNOR inhibitor, reduced bronchoconstriction and pulmonary inflammation in a mouse model of asthma and demonstrated an acceptable safety profile. N6022 is currently in clinical development as a potential agent for the treatment of acute asthma.
