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1.
Nature ; 566(7745): 548-552, 2019 02.
Artículo en Inglés | MEDLINE | ID: mdl-30760924

RESUMEN

Singlet molecular oxygen (1O2) has well-established roles in photosynthetic plants, bacteria and fungi1-3, but not in mammals. Chemically generated 1O2 oxidizes the amino acid tryptophan to precursors of a key metabolite called N-formylkynurenine4, whereas enzymatic oxidation of tryptophan to N-formylkynurenine is catalysed by a family of dioxygenases, including indoleamine 2,3-dioxygenase 15. Under inflammatory conditions, this haem-containing enzyme is expressed in arterial endothelial cells, where it contributes to the regulation of blood pressure6. However, whether indoleamine 2,3-dioxygenase 1 forms 1O2 and whether this contributes to blood pressure control have remained unknown. Here we show that arterial indoleamine 2,3-dioxygenase 1 regulates blood pressure via formation of 1O2. We observed that in the presence of hydrogen peroxide, the enzyme generates 1O2 and that this is associated with the stereoselective oxidation of L-tryptophan to a tricyclic hydroperoxide via a previously unrecognized oxidative activation of the dioxygenase activity. The tryptophan-derived hydroperoxide acts in vivo as a signalling molecule, inducing arterial relaxation and decreasing blood pressure; this activity is dependent on Cys42 of protein kinase G1α. Our findings demonstrate a pathophysiological role for 1O2 in mammals through formation of an amino acid-derived hydroperoxide that regulates vascular tone and blood pressure under inflammatory conditions.


Asunto(s)
Presión Sanguínea/fisiología , Inflamación/sangre , Inflamación/fisiopatología , Oxígeno Singlete/metabolismo , Vasodilatadores/metabolismo , Animales , Línea Celular , Proteína Quinasa Dependiente de GMP Cíclico Tipo I/antagonistas & inhibidores , Proteína Quinasa Dependiente de GMP Cíclico Tipo I/química , Proteína Quinasa Dependiente de GMP Cíclico Tipo I/metabolismo , Cisteína/metabolismo , Activación Enzimática/efectos de los fármacos , Femenino , Humanos , Peróxido de Hidrógeno/química , Peróxido de Hidrógeno/metabolismo , Peróxido de Hidrógeno/farmacología , Indolamina-Pirrol 2,3,-Dioxigenasa/química , Indolamina-Pirrol 2,3,-Dioxigenasa/metabolismo , Inflamación/enzimología , Masculino , Oxidación-Reducción/efectos de los fármacos , Ratas , Transducción de Señal , Oxígeno Singlete/química , Triptófano/química , Triptófano/metabolismo
2.
Respir Res ; 23(1): 272, 2022 Oct 01.
Artículo en Inglés | MEDLINE | ID: mdl-36183104

RESUMEN

BACKGROUND: Oxidative stress associated with severe cardiopulmonary diseases leads to impairment in the nitric oxide/soluble guanylate cyclase signaling pathway, shifting native soluble guanylate cyclase toward heme-free apo-soluble guanylate cyclase. Here we describe a new inhaled soluble guanylate cyclase activator to target apo-soluble guanylate cyclase and outline its therapeutic potential. METHODS: We aimed to generate a novel soluble guanylate cyclase activator, specifically designed for local inhaled application in the lung. We report the discovery and in vitro and in vivo characterization of the soluble guanylate cyclase activator mosliciguat (BAY 1237592). RESULTS: Mosliciguat specifically activates apo-soluble guanylate cyclase leading to improved cardiopulmonary circulation. Lung-selective effects, e.g., reduced pulmonary artery pressure without reduced systemic artery pressure, were seen after inhaled but not after intravenous administration in a thromboxane-induced pulmonary hypertension minipig model. These effects were observed over a broad dose range with a long duration of action and were further enhanced under experimental oxidative stress conditions. In a unilateral broncho-occlusion minipig model, inhaled mosliciguat decreased pulmonary arterial pressure without ventilation/perfusion mismatch. With respect to airway resistance, mosliciguat showed additional beneficial bronchodilatory effects in an acetylcholine-induced rat model. CONCLUSION: Inhaled mosliciguat may overcome treatment limitations in patients with pulmonary hypertension by improving pulmonary circulation and airway resistance without systemic exposure or ventilation/perfusion mismatch. Mosliciguat has the potential to become a new therapeutic paradigm, exhibiting a unique mode of action and route of application, and is currently under clinical development in phase Ib for pulmonary hypertension.


Asunto(s)
Hipertensión Pulmonar , Acetilcolina , Animales , Guanilato Ciclasa/metabolismo , Guanilato Ciclasa/uso terapéutico , Óxido Nítrico/metabolismo , Ratas , Guanilil Ciclasa Soluble/metabolismo , Guanilil Ciclasa Soluble/uso terapéutico , Porcinos , Porcinos Enanos/metabolismo , Tromboxanos/uso terapéutico , Vasodilatadores
3.
Handb Exp Pharmacol ; 264: 355-394, 2021.
Artículo en Inglés | MEDLINE | ID: mdl-30689085

RESUMEN

When Furchgott, Murad, and Ignarro were honored with the Nobel prize for the identification of nitric oxide (NO) in 1998, the therapeutic implications of this discovery could not be fully anticipated. This was due to the fact that available therapeutics like NO donors did not allow a constant and long-lasting cyclic guanylyl monophosphate (cGMP) stimulation and had a narrow therapeutic window. Now, 20 years later, the stimulator of soluble guanylate cyclase (sGC), riociguat, is on the market and is the only drug approved for the treatment of two forms of pulmonary hypertension (PAH/CTEPH), and a variety of other sGC stimulators and sGC activators are in preclinical and clinical development for additional indications. The discovery of sGC stimulators and sGC activators is a milestone in the field of NO/sGC/cGMP pharmacology. The sGC stimulators and sGC activators bind directly to reduced, heme-containing and oxidized, heme-free sGC, respectively, which results in an increase in cGMP production. The action of sGC stimulators at the heme-containing enzyme is independent of NO but is enhanced in the presence of NO whereas the sGC activators interact with the heme-free form of sGC. These highly innovative pharmacological principles of sGC stimulation and activation seem to have a very broad therapeutic potential. Therefore, in both academia and industry, intensive research and development efforts have been undertaken to fully exploit the therapeutic benefit of these new compound classes. Here we summarize the discovery of sGC stimulators and sGC activators and the current developments in both compound classes, including the mode of action, the chemical structures, and the genesis of the terminology and nomenclature. In addition, preclinical studies exploring multiple aspects of their in vitro, ex vivo, and in vivo pharmacology are reviewed, providing an overview of multiple potential applications. Finally, the clinical developments, investigating the treatment potential of these compounds in various diseases like heart failure, diabetic kidney disease, fibrotic diseases, and hypertension, are reported. In summary, sGC stimulators and sGC activators have a unique mode of action with a broad treatment potential in cardiovascular diseases and beyond.


Asunto(s)
Insuficiencia Cardíaca , Hipertensión Pulmonar , GMP Cíclico , Guanilato Ciclasa , Humanos , Óxido Nítrico , Guanilil Ciclasa Soluble
4.
Proc Natl Acad Sci U S A ; 113(17): E2355-62, 2016 Apr 26.
Artículo en Inglés | MEDLINE | ID: mdl-27071111

RESUMEN

Asthma is defined by airway inflammation and hyperresponsiveness, and contributes to morbidity and mortality worldwide. Although bronchodilation is a cornerstone of treatment, current bronchodilators become ineffective with worsening asthma severity. We investigated an alternative pathway that involves activating the airway smooth muscle enzyme, soluble guanylate cyclase (sGC). Activating sGC by its natural stimulant nitric oxide (NO), or by pharmacologic sGC agonists BAY 41-2272 and BAY 60-2770, triggered bronchodilation in normal human lung slices and in mouse airways. Both BAY 41-2272 and BAY 60-2770 reversed airway hyperresponsiveness in mice with allergic asthma and restored normal lung function. The sGC from mouse asthmatic lungs displayed three hallmarks of oxidative damage that render it NO-insensitive, and identical changes to sGC occurred in human lung slices or in human airway smooth muscle cells when given chronic NO exposure to mimic the high NO in asthmatic lung. Our findings show how allergic inflammation in asthma may impede NO-based bronchodilation, and reveal that pharmacologic sGC agonists can achieve bronchodilation despite this loss.


Asunto(s)
Antiasmáticos/farmacología , Asma/tratamiento farmacológico , Benzoatos/farmacología , Compuestos de Bifenilo/farmacología , Broncodilatadores/farmacología , Guanilato Ciclasa/efectos de los fármacos , Hidrocarburos Fluorados/farmacología , Pirazoles/farmacología , Piridinas/farmacología , Animales , Antiasmáticos/uso terapéutico , Asma/enzimología , Asma/fisiopatología , Benzoatos/uso terapéutico , Compuestos de Bifenilo/uso terapéutico , Hiperreactividad Bronquial/tratamiento farmacológico , Hiperreactividad Bronquial/enzimología , Broncodilatadores/uso terapéutico , Técnicas de Cocultivo , GMP Cíclico/metabolismo , Evaluación Preclínica de Medicamentos , Activación Enzimática/efectos de los fármacos , Femenino , Humanos , Hidrocarburos Fluorados/uso terapéutico , Pulmón/enzimología , Ratones , Ratones Endogámicos BALB C , Músculo Liso/efectos de los fármacos , Músculo Liso/enzimología , Óxido Nítrico/farmacología , Pirazoles/uso terapéutico , Piridinas/uso terapéutico , Solubilidad , Tráquea/efectos de los fármacos
5.
Am J Respir Cell Mol Biol ; 58(5): 636-647, 2018 05.
Artículo en Inglés | MEDLINE | ID: mdl-29268036

RESUMEN

Sickle cell disease (SCD) is associated with intravascular hemolysis and oxidative inhibition of nitric oxide (NO) signaling. BAY 54-6544 is a small-molecule activator of oxidized soluble guanylate cyclase (sGC), which, unlike endogenous NO and the sGC stimulator, BAY 41-8543, preferentially binds and activates heme-free, NO-insensitive sGC to restore enzymatic cGMP production. We tested orally delivered sGC activator, BAY 54-6544 (17 mg/kg/d), sGC stimulator, BAY 41-8543, sildenafil, and placebo for 4-12 weeks in the Berkeley transgenic mouse model of SCD (BERK-SCD) and their hemizygous (Hemi) littermate controls (BERK-Hemi). Right ventricular (RV) maximum systolic pressure (RVmaxSP) was measured using micro right-heart catheterization. RV hypertrophy (RVH) was determined using Fulton's index and RV corrected weight (ratio of RV to tibia). Pulmonary artery vasoreactivity was tested for endothelium-dependent and -independent vessel relaxation. Right-heart catheterization revealed higher RVmaxSP and RVH in BERK-SCD versus BERK-Hemi, which worsened with age. Treatment with the sGC activator more effectively lowered RVmaxSP and RVH, with 90-day treatment delivering superior results, when compared with other treatments and placebo groups. In myography experiments, acetylcholine-induced (endothelium-dependent) and sodium-nitroprusside-induced (endothelium-independent NO donor) relaxation of the pulmonary artery harvested from placebo-treated BERK-SCD was impaired relative to BERK-Hemi but improved after therapy with sGC activator. By contrast, no significant effect for sGC stimulator or sildenafil was observed in BERK-SCD. These findings suggest that sGC is oxidized in the pulmonary arteries of transgenic SCD mice, leading to blunted responses to NO, and that the sGC activator, BAY 54-6544, may represent a novel therapy for SCD-associated pulmonary arterial hypertension and cardiac remodeling.


Asunto(s)
Anemia de Células Falciformes/complicaciones , Activadores de Enzimas/farmacología , Ventrículos Cardíacos/efectos de los fármacos , Hipertensión Pulmonar/tratamiento farmacológico , Hipertrofia Ventricular Izquierda/prevención & control , Arteria Pulmonar/efectos de los fármacos , Guanilil Ciclasa Soluble/metabolismo , Disfunción Ventricular Derecha/tratamiento farmacológico , Función Ventricular Derecha/efectos de los fármacos , Remodelación Ventricular/efectos de los fármacos , Anemia de Células Falciformes/genética , Animales , Presión Arterial/efectos de los fármacos , Modelos Animales de Enfermedad , Activación Enzimática , Activadores de Enzimas/farmacocinética , Ventrículos Cardíacos/enzimología , Ventrículos Cardíacos/fisiopatología , Hipertensión Pulmonar/enzimología , Hipertensión Pulmonar/genética , Hipertensión Pulmonar/fisiopatología , Hipertrofia Ventricular Izquierda/enzimología , Hipertrofia Ventricular Izquierda/genética , Hipertrofia Ventricular Izquierda/fisiopatología , Ratones Transgénicos , Morfolinas/farmacología , Óxido Nítrico/metabolismo , Arteria Pulmonar/enzimología , Arteria Pulmonar/fisiopatología , Pirimidinas/farmacología , Citrato de Sildenafil/farmacología , Vasodilatación/efectos de los fármacos , Disfunción Ventricular Derecha/enzimología , Disfunción Ventricular Derecha/genética , Disfunción Ventricular Derecha/fisiopatología , Presión Ventricular/efectos de los fármacos
6.
Nitric Oxide ; 77: 88-95, 2018 07 01.
Artículo en Inglés | MEDLINE | ID: mdl-29738821

RESUMEN

The NO/sGC/cGMP signaling cascade plays a pivotal role in regulation of cardiovascular, cardiopulmonary and cardiorenal diseases and impairment of this cascade results in severe pathologies. Therefore, pharmacological interventions, targeting this pathway are promising strategies for treating a variety of diseases. Nitrates, supplementing NO and, PDE5 inhibitors preventing cGMP degradation, are used for angina pectoris treatment and the treatment of pulmonary arterial hypertension (PAH), respectively. More recently, a new class of drugs which directly stimulate the sGC enzyme and trigger NO-independent cGMP production was introduced and termed sGC stimulators. In 2013, the first sGC stimulator, riociguat, was approved for the treatment of PAH and chronic thromboembolic pulmonary hypertension (CTEPH). Since cGMP targets multiple intracellular downstream targets, sGC stimulators have shown - beyond the well characterized vasodilatation - anti-fibrotic, anti-inflammatory and anti-proliferative effects. These additional modes of action might extend the therapeutic potential of this drug class substantially. This review summarizes the NO/sGC/cGMP signaling cascades, the discovery and the mode of action of sGC stimulators. Furthermore, the preclinical evidence and development of riociguat for the treatment of PAH and CTEPH is reviewed. Finally, a summary of the antifibrotic effects of sGC stimulators, especially the most recent finding for skin fibrosis are included which may indicate efficacy in fibrotic diseases like Systemic Sclerosis (SSc).


Asunto(s)
Activadores de Enzimas/farmacología , Hipertensión Pulmonar/tratamiento farmacológico , Pirazoles/farmacología , Pirimidinas/farmacología , Enfermedades Raras/tratamiento farmacológico , Esclerodermia Sistémica/tratamiento farmacológico , Guanilil Ciclasa Soluble/metabolismo , GMP Cíclico/metabolismo , Humanos , Hipertensión Pulmonar/metabolismo , Óxido Nítrico/metabolismo , Enfermedades Raras/metabolismo , Esclerodermia Sistémica/metabolismo
7.
Handb Exp Pharmacol ; 232: 223-34, 2016.
Artículo en Inglés | MEDLINE | ID: mdl-26552402

RESUMEN

Cardiovascular diseases are still the first leading cause of death and morbidity in developed countries. Experimental cardiology research and preclinical drug development in cardiology call for appropriate and especially clinically relevant in vitro and in vivo studies. The use of animal models has contributed to expand our knowledge and our understanding of the underlying mechanisms and accordingly provided new approaches focused on the improvement of diagnostic and treatment strategies of various cardiac pathologies.Numerous animal models in different species as well as in small and large animals have been developed to address cardiovascular complications, including heart failure, pulmonary hypertension, and thrombotic diseases. However, a perfect model of heart failure or other indications that reproduces every aspect of the natural disease does not exist. The complexity and heterogeneity of cardiac diseases plus the influence of genetic and environmental factors limit to mirror a particular disease with a single experimental model.Thus, drug development in the field of cardiology is not only very challenging but also inspiring; therefore animal models should be selected that reflect as best as possible the disease being investigated. Given the wide range of animal models, reflecting critical features of the human pathophysiology available nowadays increases the likelihood of the translation to the patients. Furthermore, this knowledge and the increase of the predictive value of preclinical models help us to find more efficient and reliable solutions as well as better and innovative treatment strategies for cardiovascular diseases.


Asunto(s)
Enfermedades Cardiovasculares/tratamiento farmacológico , Modelos Animales de Enfermedad , Descubrimiento de Drogas , Animales , Evaluación Preclínica de Medicamentos , Humanos , Investigación Biomédica Traslacional
9.
J Biol Chem ; 289(22): 15259-71, 2014 May 30.
Artículo en Inglés | MEDLINE | ID: mdl-24733395

RESUMEN

The chaperone heat shock protein 90 (hsp90) associates with signaling proteins in cells including soluble guanylate cyclase (sGC). hsp90 associates with the heme-free (apo) sGC-ß1 subunit and helps to drive heme insertion during maturation of sGC to its NO-responsive active form. Here, we found that NO caused apo-sGC-ß1 to rapidly and transiently dissociate from hsp90 and associate with sGC-α1 in cells. This NO response (i) required that hsp90 be active and that cellular heme be available and be capable of inserting into apo-sGC-ß1; (ii) was associated with an increase in sGC-ß1 heme content; (iii) could be mimicked by the heme-independent sGC activator BAY 60-2770; and (iv) was followed by desensitization of sGC toward NO, sGC-α1 disassociation, and reassociation with hsp90. Thus, NO promoted a rapid, transient, and hsp90-dependent heme insertion into the apo-sGC-ß1 subpopulation in cells, which enabled it to combine with the sGC-α1 subunit to form the mature enzyme. The driving mechanism likely involves conformational changes near the heme site in sGC-ß1 that can be mimicked by the pharmacologic sGC activator. Such dynamic interplay between hsp90, apo-sGC-ß1, and sGC-α1 in response to NO is unprecedented and represent new steps by which cells can modulate the heme content and activity of sGC for signaling cascades.


Asunto(s)
Guanilato Ciclasa/metabolismo , Proteínas HSP90 de Choque Térmico/metabolismo , Hemo/metabolismo , Óxido Nítrico/metabolismo , Receptores Citoplasmáticos y Nucleares/metabolismo , Transducción de Señal/fisiología , Animales , Aorta/citología , Benzoatos/farmacología , Compuestos de Bifenilo/farmacología , Células COS , Bovinos , Chlorocebus aethiops , Dimerización , Células Endoteliales/citología , Células Endoteliales/enzimología , Fibroblastos/citología , Fibroblastos/enzimología , Guanilato Ciclasa/química , Guanilato Ciclasa/genética , Proteínas HSP90 de Choque Térmico/agonistas , Humanos , Hidrocarburos Fluorados/farmacología , Pulmón/citología , Donantes de Óxido Nítrico/farmacología , Pirazoles/farmacología , Piridinas/farmacología , Ratas , Receptores Citoplasmáticos y Nucleares/química , Receptores Citoplasmáticos y Nucleares/genética , Transducción de Señal/efectos de los fármacos , Guanilil Ciclasa Soluble
10.
Cell Physiol Biochem ; 37(4): 1474-90, 2015.
Artículo en Inglés | MEDLINE | ID: mdl-26509263

RESUMEN

BACKGROUND/AIMS: To investigate the renal phenotype under conditions of an activated renal ET-1 system in the status of nitric oxide deficiency, we compared kidney function and morphology in wild-type, ET-1 transgenic (ET+/+), endothelial nitric oxide synthase knockout (eNOS-/-) and ET+/+eNOS-/- mice. METHODS: We assessed blood pressure, parameters of renal morphology, plasma cystatin C, urinary protein excretion, expression of genes associated with glomerular filtration barrier and tissue remodeling, and plasma metabolites using metabolomics. RESULTS: eNOS-/- and ET+/+eNOS-/- mice developed hypertension. Osteopontin, albumin and protein excretion were increased in eNOS-/- and restored in ET+/+eNOS-/- animals. All genetically modified mice developed renal interstitial fibrosis and glomerulosclerosis. Genes involved in tissue remodeling (serpine1, TIMP1, Col1a1, CCL2) were up-regulated in eNOS-/-, but not in ET+/+eNOS-/- mice. Plasma levels of free carnitine and acylcarnitines, amino acids, diacyl phosphatidylcholines, lysophosphatidylcholines and hexoses were descreased in eNOS-/- and were in the normal range in ET+/+eNOS-/- mice. CONCLUSION: eNOS-/- mice developed renal dysfunction, which was partially rescued by ET-1 overexpression in eNOS-/- mice. The metabolomics results suggest that ET-1 overexpression on top of eNOS knockout is associated with a functional recovery of mitochondria (rescue effect in ß-oxidation of fatty acids) and an increase in antioxidative properties (normalization of monounsaturated fatty acids levels).


Asunto(s)
Endotelina-1/metabolismo , Riñón/metabolismo , Óxido Nítrico Sintasa de Tipo III/genética , Albúminas/análisis , Animales , Presión Sanguínea , Carnitina/sangre , Colágeno Tipo I/genética , Colágeno Tipo I/metabolismo , Cadena alfa 1 del Colágeno Tipo I , Factor de Crecimiento del Tejido Conjuntivo/genética , Factor de Crecimiento del Tejido Conjuntivo/metabolismo , Cistatina C/sangre , Endotelina-1/genética , Proteína 3 de Unión a Ácidos Grasos , Proteínas de Unión a Ácidos Grasos/sangre , Proteínas de Unión a Ácidos Grasos/orina , Femenino , Humanos , Riñón/patología , Masculino , Ratones , Ratones Noqueados , Ratones Transgénicos , Mitocondrias/metabolismo , Óxido Nítrico/metabolismo , Óxido Nítrico Sintasa de Tipo III/deficiencia , Osteopontina/sangre , Osteopontina/orina , Inhibidor Tisular de Metaloproteinasa-1/genética , Inhibidor Tisular de Metaloproteinasa-1/metabolismo
11.
Am J Respir Crit Care Med ; 189(11): 1359-73, 2014 Jun 01.
Artículo en Inglés | MEDLINE | ID: mdl-24738736

RESUMEN

RATIONALE: Chronic obstructive pulmonary disease (COPD) is a major cause of death worldwide. No therapy stopping progress of the disease is available. OBJECTIVES: To investigate the role of the soluble guanylate cyclase (sGC)-cGMP axis in development of lung emphysema and pulmonary hypertension (PH) and to test whether the sGC-cGMP axis is a treatment target for these conditions. METHODS: Investigations were performed in human lung tissue from patients with COPD, healthy donors, mice, and guinea pigs. Mice were exposed to cigarette smoke (CS) for 6 hours per day, 5 days per week for up to 6 months and treated with BAY 63-2521. Guinea pigs were exposed to CS from six cigarettes per day for 3 months, 5 days per week and treated with BAY 41-2272. Both BAY compounds are sGC stimulators. Gene and protein expression analysis were performed by quantitative real-time polymerase chain reaction and Western blotting. Lung compliance, hemodynamics, right ventricular heart mass alterations, and alveolar and vascular morphometry were performed, as well as inflammatory cell infiltrate assessment. In vitro assays of cell adhesion, proliferation, and apoptosis have been done. MEASUREMENTS AND MAIN RESULTS: The functionally essential sGC ß1-subunit was down-regulated in patients with COPD and in CS-exposed mice. sGC stimulators prevented the development of PH and emphysema in the two different CS-exposed animal models. sGC stimulation prevented peroxynitrite-induced apoptosis of alveolar and endothelial cells, reduced CS-induced inflammatory cell infiltrate in lung parenchyma, and inhibited adhesion of CS-stimulated neutrophils. CONCLUSIONS: The sGC-cGMP axis is perturbed by chronic exposure to CS. Treatment of COPD animal models with sGC stimulators can prevent CS-induced PH and emphysema.


Asunto(s)
Enfisema/prevención & control , Guanilato Ciclasa/metabolismo , Hipertensión Pulmonar/prevención & control , Enfermedad Pulmonar Obstructiva Crónica/prevención & control , Receptores Citoplasmáticos y Nucleares/metabolismo , Fumar/efectos adversos , Animales , Biomarcadores/metabolismo , Western Blotting , Modelos Animales de Enfermedad , Regulación hacia Abajo , Enfisema/enzimología , Cobayas , Humanos , Hipertensión Pulmonar/enzimología , Técnicas In Vitro , Ratones , Enfermedad Pulmonar Obstructiva Crónica/enzimología , Reacción en Cadena en Tiempo Real de la Polimerasa , Fumar/metabolismo , Guanilil Ciclasa Soluble
12.
Mol Pharmacol ; 85(4): 598-607, 2014 Apr.
Artículo en Inglés | MEDLINE | ID: mdl-24470063

RESUMEN

Soluble guanylyl cyclase (sGC) plays an important role in cardiovascular function and catalyzes formation of cGMP. sGC is activated by nitric oxide and allosteric stimulators and activators. However, despite its therapeutic relevance, the regulatory mechanisms of sGC are still incompletely understood. A major reason for this situation is that no crystal structures of active sGC have been resolved so far. An important step toward this goal is the identification of high-affinity ligands that stabilize an sGC conformation resembling the active, "fully closed" state. Therefore, we examined inhibition of rat sGCα1ß1 by 38 purine- and pyrimidine-nucleotides with 2,4,6,-trinitrophenyl and (N-methyl)anthraniloyl substitutions at the ribosyl moiety and compared the data with that for the structurally related membranous adenylyl cyclases (mACs) 1, 2, 5 and the purified mAC catalytic subunits VC1:IIC2. TNP-GTP [2',3'-O-(2,4,6-trinitrophenyl)-GTP] was the most potent sGCα1ß1 inhibitor (Ki, 10.7 nM), followed by 2'-MANT-3'-dATP [2'-O-(N-methylanthraniloyl)-3'-deoxy-ATP] (Ki, 16.7 nM). Docking studies on an sGCαcat/sGCßcat model derived from the inactive heterodimeric crystal structure of the catalytic domains point to similar interactions of (M)ANT- and TNP-nucleotides with sGCα1ß1 and mAC VC1:IIC2. Reasonable binding modes of 2'-MANT-3'-dATP and bis-(M)ANT-nucleotides at sGC α1ß1 require a 3'-endo ribosyl conformation (versus 3'-exo in 3'-MANT-2'-dATP). Overall, inhibitory potencies of nucleotides at sGCα1ß1 versus mACs 1, 2, 5 correlated poorly. Collectively, we identified highly potent sGCα1ß1 inhibitors that may be useful for future crystallographic and fluorescence spectroscopy studies. Moreover, it may become possible to develop mAC inhibitors with selectivity relative to sGC.


Asunto(s)
Guanilato Ciclasa/antagonistas & inhibidores , Nitrocompuestos/química , Nucleótidos de Purina/química , Nucleótidos de Pirimidina/química , Receptores Citoplasmáticos y Nucleares/antagonistas & inhibidores , ortoaminobenzoatos/química , Inhibidores de Adenilato Ciclasa , Adenilil Ciclasas/química , Animales , Guanilato Ciclasa/química , Enlace de Hidrógeno , Interacciones Hidrofóbicas e Hidrofílicas , Simulación del Acoplamiento Molecular , Unión Proteica , Conformación Proteica , Subunidades de Proteína/antagonistas & inhibidores , Subunidades de Proteína/química , Ratas , Receptores Citoplasmáticos y Nucleares/química , Proteínas Recombinantes/química , Guanilil Ciclasa Soluble , Relación Estructura-Actividad
13.
Am J Physiol Renal Physiol ; 307(9): F1003-12, 2014 Nov 01.
Artículo en Inglés | MEDLINE | ID: mdl-25209860

RESUMEN

We studied the influence of soluble guanylate (sGC) on renal blood flow (RBF), glomerular filtration rate (GFR), and RBF autoregulation and its role in mediating the hemodynamic effects of endogenous nitric oxide (NO). Arterial pressure (AP), heart rate (HR), RBF, GFR, urine flow (UV), and the efficiency and mechanisms of RBF autoregulation were studied in anesthetized rats during intravenous infusion of sGC activator cinaciguat before and (except GFR) also after inhibition of NO synthase (NOS) by Nω-nitro-L-arginine methyl ester. Cinaciguat (0.1, 0.3, 1, 3, 10 µg·kg(-1)·min(-1), n=7) reduced AP and increased HR, but did not significantly alter RBF. In clearance experiments (FITC-sinistrin, n=7) GFR was not significantly altered by cinaciguat (0.1 and 1 µg·kg(-1)·min(-1)), but RBF slightly rose (+12%) and filtration fraction (FF) fell (-23%). RBF autoregulatory efficiency (67 vs. 104%) and myogenic response (33 vs. 44 units) were slightly depressed (n=9). NOS inhibition (n=7) increased AP (+38 mmHg), reduced RBF (-53%), and greatly augmented the myogenic response in RBF autoregulation (97 vs. 35 units), attenuating the other regulatory mechanisms. These changes were reversed by 77, 78, and 90% by 1 µg·kg(-1)·min(-1) cinaciguat. In vehicle controls (n=3), in which cinaciguat-induced hypotension was mimicked by aortic compression, the NOS inhibition-induced changes were not affected. We conclude that sGC activation leaves RBF and GFR well maintained despite hypotension and only slightly impairs autoregulation. The ability to largely normalize AP, RBF, RBF autoregulation, and renovascular myogenic response after NOS inhibition indicates that these hemodynamic effects of NO are predominantly mediated via sGC.


Asunto(s)
Guanilato Ciclasa/fisiología , Receptores Citoplasmáticos y Nucleares/fisiología , Circulación Renal/fisiología , Animales , Benzoatos/farmacología , Tasa de Filtración Glomerular/efectos de los fármacos , Masculino , Óxido Nítrico/fisiología , Ratas , Ratas Wistar , Circulación Renal/efectos de los fármacos , Guanilil Ciclasa Soluble
14.
Biochem Biophys Res Commun ; 443(4): 1195-9, 2014 Jan 24.
Artículo en Inglés | MEDLINE | ID: mdl-24380860

RESUMEN

Soluble guanylyl cyclase (sGC) is activated by nitric oxide (NO) and generates the second messenger cyclic GMP (cGMP). Recently, purified sGC α1ß1 has been shown to additionally generate the cyclic pyrimidine nucleotides cCMP and cUMP. However, since cyclic pyrimidine nucleotide formation occurred only the presence of Mn(2+) but not Mg(2+), the physiological relevance of these in vitro findings remained unclear. Therefore, we studied cyclic nucleotide formation in intact cells. We observed NO-dependent cCMP- and cUMP formation in intact HEK293 cells overexpressing sGC α1ß1 and in RFL-6 rat fibroblasts endogenously expressing sGC, using HPLC-tandem mass spectrometry. The identity of cCMP and cUMP was unambiguously confirmed by HPLC-time-of-flight mass spectrometry. Our data indicate that cCMP and cUMP play second messenger roles and that Mn(2+) is a physiological sGC cofactor.


Asunto(s)
Guanilato Ciclasa/metabolismo , Receptores Citoplasmáticos y Nucleares/metabolismo , Animales , Línea Celular , AMP Cíclico/biosíntesis , CMP Cíclico/biosíntesis , GMP Cíclico/biosíntesis , Guanilato Ciclasa/genética , Células HEK293 , Humanos , Manganeso/metabolismo , Óxido Nítrico/metabolismo , Donantes de Óxido Nítrico/farmacología , Nitroprusiato/farmacología , Nucleótidos Cíclicos/biosíntesis , Ratas , Receptores Citoplasmáticos y Nucleares/genética , Proteínas Recombinantes/genética , Proteínas Recombinantes/metabolismo , Sistemas de Mensajero Secundario , Guanilil Ciclasa Soluble , Transfección , Uridina Monofosfato/biosíntesis
15.
Eur Respir Rev ; 33(171)2024 Jan 31.
Artículo en Inglés | MEDLINE | ID: mdl-38508664

RESUMEN

The nitric oxide (NO)-soluble guanylate cyclase (sGC)-cyclic guanosine monophosphate (cGMP) pathway plays a key role in the pathogenesis of pulmonary hypertension (PH). Targeted treatments include phosphodiesterase type 5 inhibitors (PDE5i) and sGC stimulators. The sGC stimulator riociguat is approved for the treatment of pulmonary arterial hypertension (PAH) and chronic thromboembolic pulmonary hypertension (CTEPH). sGC stimulators have a dual mechanism of action, enhancing the sGC response to endogenous NO and directly stimulating sGC, independent of NO. This increase in cGMP production via a dual mechanism differs from PDE5i, which protects cGMP from degradation by PDE5, rather than increasing its production. sGC stimulators may therefore have the potential to increase cGMP levels under conditions of NO depletion that could limit the effectiveness of PDE5i. Such differences in mode of action between sGC stimulators and PDE5i could lead to differences in treatment efficacy between the classes. In addition to vascular effects, sGC stimulators have the potential to reduce inflammation, angiogenesis, fibrosis and right ventricular hypertrophy and remodelling. In this review we describe the evolution of treatments targeting the NO-sGC-cGMP pathway, with a focus on PH.


Asunto(s)
Hipertensión Pulmonar , Hipertensión Arterial Pulmonar , Humanos , Guanilil Ciclasa Soluble/metabolismo , Hipertensión Pulmonar/etiología , Óxido Nítrico/metabolismo , Transducción de Señal , GMP Cíclico/metabolismo , Guanilato Ciclasa/metabolismo
16.
Biochemistry ; 52(20): 3601-8, 2013 May 21.
Artículo en Inglés | MEDLINE | ID: mdl-23614626

RESUMEN

The soluble guanylyl cyclase (sGC) is an important receptor for nitric oxide (NO). Nitric oxide activates sGC several hundred fold to generate cGMP from GTP. Because of sGC's salutary roles in cardiovascular physiology, it has received substantial attention as a drug target. The heme domain of sGC is key to its regulation as it not only contains the NO activation site but also harbors sites for NO-independent sGC activators as well an S-nitrosylation site (ß1 C122) involved in desensitization. Here we report the crystal structure of the activator BAY 60-2770 bound to the Nostoc H-NOX domain that is homologous to sGC. The structure reveals that BAY 60-2770 has displaced the heme and acts as a heme mimetic via carboxylate-mediated interactions with the conserved YxSxR motif as well as hydrophobic interactions. Comparisons with the previously determined BAY 58-2667 bound structure reveal that BAY 60-2770 is more ordered in its hydrophobic tail region. sGC activity assays demonstrate that BAY 60-2770 has about 10% higher fold maximal stimulation compared to BAY 58-2667. S-Nitrosylation of the BAY 60-2770 substituted Nostoc H-NOX domain causes subtle changes in the vicinity of the S-nitrosylated C122 residue. These shifts could impact the adjacent YxSxR motif and αF helix and as such potentially inhibit either heme incorporation or NO-activation of sGC and thus provide a structural basis for desensitization.


Asunto(s)
Proteínas Bacterianas/química , Benzoatos/química , Compuestos de Bifenilo/química , Guanilato Ciclasa/química , Hidrocarburos Fluorados/química , Nostoc/metabolismo , Receptores Citoplasmáticos y Nucleares/química , Proteínas Bacterianas/metabolismo , Benzoatos/metabolismo , Compuestos de Bifenilo/metabolismo , Cristalografía por Rayos X , Guanilato Ciclasa/metabolismo , Hidrocarburos Fluorados/metabolismo , Modelos Moleculares , Óxido Nítrico/metabolismo , Estructura Terciaria de Proteína , Receptores Citoplasmáticos y Nucleares/metabolismo , Guanilil Ciclasa Soluble
17.
Heart Fail Rev ; 18(2): 123-34, 2013 Mar.
Artículo en Inglés | MEDLINE | ID: mdl-22622468

RESUMEN

The number of annual hospitalizations for heart failure (HF) and the mortality rates among patients hospitalized for HF remains unacceptably high. The search continues for safe and effective agents that improve outcomes when added to standard therapy. The nitric oxide (NO)-soluble guanylate cyclase (sGC)-cyclic guanosine monophosphate (cGMP) pathway serves an important physiologic role in both vascular and non-vascular tissues, including regulation of myocardial and renal function, and is disrupted in the setting of HF, leading to decreased protection against myocardial injury, ventricular remodeling, and the cardio-renal syndrome. The impaired NO-sGC-cGMP pathway signaling in HF is secondary to reduced NO bioavailability and an alteration in the redox state of sGC, making it unresponsive to NO. Accordingly, increasing directly the activity of sGC is an attractive pharmacologic strategy. With the development of two novel classes of drugs, sGC stimulators and sGC activators, the hypothesis that restoration of NO-sGC-cGMP signaling is beneficial in HF patients can now be tested. Characterization of these agents in pre-clinical and clinical studies has begun with investigations suggesting both hemodynamic effects and organ-protective properties independent of hemodynamic changes. The latter could prove valuable in long-term low-dose therapy in HF patients. This review will explain the role of the NO-sGC-cGMP pathway in HF pathophysiology and outcomes, data obtained with sGC stimulators and sGC activators in pre-clinical and clinical studies, and a plan for the further clinical development to study these agents as HF therapy.


Asunto(s)
GMP Cíclico/uso terapéutico , Guanilato Ciclasa/efectos de los fármacos , Insuficiencia Cardíaca/tratamiento farmacológico , Hemodinámica/efectos de los fármacos , Óxido Nítrico/uso terapéutico , Receptores Citoplasmáticos y Nucleares/metabolismo , Transducción de Señal/efectos de los fármacos , GMP Cíclico/metabolismo , Guanilato Ciclasa/biosíntesis , Guanilato Ciclasa/metabolismo , Insuficiencia Cardíaca/fisiopatología , Humanos , Óxido Nítrico/metabolismo , Receptores Citoplasmáticos y Nucleares/agonistas , Receptores Citoplasmáticos y Nucleares/biosíntesis , Guanilil Ciclasa Soluble , Resultado del Tratamiento
18.
J Sex Med ; 10(3): 704-18, 2013 Mar.
Artículo en Inglés | MEDLINE | ID: mdl-22989320

RESUMEN

INTRODUCTION: Soluble guanylate cyclase (sGC) is the receptor for nitric oxide (NO) and in pathophysiologic conditions where NO formation or bioavailability is impaired, erectile dysfunction (ED) occurs. AIM: The aim of this study was to investigate erectile responses to the sGC stimulator BAY 41-8543 in physiologic and pathophysiologic conditions. METHODS: Increases in intracavernosal pressure (ICP) in response to intracavernosal (ic) injections of BAY 41-8543 were investigated in the anesthetized rat. MAIN OUTCOME MEASURES: Increases in ICP/MAP in response to ic injections of BAY 41-8543 and the interaction of BAY 41-8543 with exogenous and endogenously released NO were investigated and the effect of the sGC stimulator on cavernosal nerve injury was assessed. The mechanism of the increase in ICP/MAP in response to ic injection of acetylcholine was investigated. RESULTS: The ic injections of BAY 41-8543 increased ICP/MAP and the duration of the response. BAY 41-8543 was less potent than sodium nitroprusside (SNP) and ic injections of BAY 41-8543 and SNP produced a larger response than the algebraic sum of responses to either agent alone. Simultaneous ic injection of BAY 41-8543 and cavernosal nerve stimulation produced a greater response than either intervention alone. Atropine and cavernosal nerve crush injury decreased the response to nerve stimulation and ic injection of BAY 41-8543 restored the response. CONCLUSION: These data show that BAY 41-8543 has significant erectile activity and can synergize with exogenous and endogenously released NO. This study shows that atropine and nerve crush attenuate the response to cavernosal nerve stimulation and that BAY 41-8543 can restore the response. The results with atropine, L-NAME and hexamethonium indicate that the response to ic injection of acetylcholine is mediated by muscarinic receptors and the release of NO with no significant role for nicotinic receptors. These results suggest that BAY 41-8543 would be useful in the treatment of ED.


Asunto(s)
Estimulación Eléctrica , Morfolinas/farmacología , Erección Peniana/efectos de los fármacos , Pene/inervación , Pirimidinas/farmacología , Acetilcolina/farmacología , Animales , Atropina/farmacología , Presión Sanguínea/efectos de los fármacos , Agonistas Colinérgicos/farmacología , Inhibidores Enzimáticos/farmacología , Frecuencia Cardíaca/efectos de los fármacos , Hexametonio/farmacología , Inyecciones , Masculino , NG-Nitroarginina Metil Éster/farmacología , Antagonistas Nicotínicos/farmacología , Donantes de Óxido Nítrico/farmacología , Nitroprusiato/farmacología , Parasimpatolíticos/farmacología , Traumatismos de los Nervios Periféricos , Ratas , Ratas Sprague-Dawley
20.
Handb Exp Pharmacol ; 218: 279-313, 2013.
Artículo en Inglés | MEDLINE | ID: mdl-24092345

RESUMEN

Soluble guanylate cyclase (sGC) is a key enzyme in the nitric oxide (NO) signalling pathway. On binding of NO to its prosthetic haem group, sGC catalyses the synthesis of the second messenger cyclic guanosine monophosphate (cGMP), which promotes vasodilation and inhibits smooth muscle proliferation, leukocyte recruitment, platelet aggregation and vascular remodelling through a number of downstream mechanisms. The central role of the NO-sGC-cGMP pathway in regulating pulmonary vascular tone is demonstrated by the dysregulation of NO production, sGC activity and cGMP degradation in pulmonary hypertension (PH). The sGC stimulators are novel pharmacological agents that directly stimulate sGC, both independently of NO and in synergy with NO. Optimisation of the first sGC stimulator, YC-1, led to the development of the more potent and more specific sGC stimulators, BAY 41-2272, BAY 41-8543 and riociguat (BAY 63-2521). Other sGC stimulators include CFM-1571, BAY 60-4552, vericiguat (BAY 1021189), the acrylamide analogue A-350619 and the aminopyrimidine analogues. BAY 41-2272, BAY 41-8543 and riociguat induced marked dose-dependent reductions in mean pulmonary arterial pressure and vascular resistance with a concomitant increase in cardiac output, and they also reversed vascular remodelling and right heart hypertrophy in several experimental models of PH. Riociguat is the first sGC stimulator that has entered clinical development. Clinical trials have shown that it significantly improves pulmonary vascular haemodynamics and increases exercise ability in patients with pulmonary arterial hypertension (PAH), chronic thromboembolic PH and PH associated with interstitial lung disease. Furthermore, riociguat reduces mean pulmonary arterial pressure in patients with PH associated with chronic obstructive pulmonary disease and improves cardiac index and pulmonary vascular resistance in patients with PH associated with left ventricular systolic dysfunction. These promising results suggest that sGC stimulators may constitute a valuable new therapy for PH. Other trials of riociguat are in progress, including a study of the haemodynamic effects and safety of riociguat in patients with PH associated with left ventricular diastolic dysfunction, and long-term extensions of the phase 3 trials investigating the efficacy and safety of riociguat in patients with PAH and chronic thromboembolic PH. Finally, sGC stimulators may also have potential therapeutic applications in other diseases, including heart failure, lung fibrosis, scleroderma and sickle cell disease.


Asunto(s)
Guanilato Ciclasa/fisiología , Compuestos Heterocíclicos con 2 Anillos/uso terapéutico , Hipertensión Pulmonar/tratamiento farmacológico , Pirazoles/uso terapéutico , Pirimidinas/uso terapéutico , Receptores Citoplasmáticos y Nucleares/fisiología , Animales , Ensayos Clínicos como Asunto , GMP Cíclico/fisiología , Humanos , Morfolinas/uso terapéutico , Óxido Nítrico/fisiología , Piridinas/uso terapéutico , Guanilil Ciclasa Soluble
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