RESUMEN
CYP2D6 and CYP3A4, two members of the cytochrome P450 superfamily of monooxygenases, mediate the biotransformation of a variety of xenobiotics. The two enzymes differ in substrate specificity and size and characteristics of the active site cavity. The aim of this study was to determine whether the catalytic properties of these isoforms, reflected by the differences observed from crystal structures and homology models, could be confirmed with experimental data. Detailed metabolite identification, reversible inhibition, and time-dependent inhibition were examined for levorphanol and levallorphan with CYP2D6 and CYP3A4. The studies were designed to provide a comparison of the orientations of substrates, the catalytic sites of the two enzymes, and the subsequent outcomes on metabolism and inhibition. The metabolite identification revealed that CYP3A4 catalyzed the formation of a variety of metabolites as a result of presenting different parts of the substrates to the heme. CYP2D6 was a poorer catalyst that led to a more limited number of metabolites that were interpreted in terms to two orientations of the substrates. The inhibition studies showed evidence for strong reversible inhibition of CYP2D6 but not for CYP3A4. Levallorphan acted as a time-dependent inhibitor on CYP3A4, indicating a productive binding mode with this enzyme not observed with CYP2D6 that presumably resulted from close interactions of the N-allyl moiety oriented toward the heme. All the results are in agreement with the large and flexible active site of CYP3A4 and the more restricted active site of CYP2D6.
Asunto(s)
Biocatálisis , Citocromo P-450 CYP2D6/metabolismo , Citocromo P-450 CYP3A/metabolismo , Levalorfano/metabolismo , Levorfanol/metabolismo , Dominio Catalítico , Cromatografía Líquida de Alta Presión , Citocromo P-450 CYP2D6/química , Inhibidores del Citocromo P-450 CYP2D6 , Citocromo P-450 CYP3A/química , Inhibidores Enzimáticos/química , Inhibidores Enzimáticos/metabolismo , Glutatión/metabolismo , Humanos , Interacciones Hidrofóbicas e Hidrofílicas , Cinética , Levalorfano/análogos & derivados , Levalorfano/análisis , Levalorfano/química , Levorfanol/análogos & derivados , Levorfanol/análisis , Levorfanol/química , Modelos Químicos , Modelos Moleculares , Estructura Molecular , Proteínas Recombinantes/química , Proteínas Recombinantes/metabolismo , Espectrometría de Masa por Ionización de Electrospray , Electricidad Estática , Espectrometría de Masas en Tándem , TermodinámicaRESUMEN
A simple, achiral monoamine sigma-1 (sigma1) receptor selective ligand (sigma2Ki/sigma1Ki>2000) is described, which could replace the chiral (+)-pentazocine or dextrallorphan as a sigma1 masking agent in sigma2 binding assays.
Asunto(s)
Analgésicos Opioides/metabolismo , Etilaminas/metabolismo , Fenilacetatos/metabolismo , Pirrolidinas/metabolismo , Receptores sigma/metabolismo , Unión Competitiva , Etilaminas/farmacología , Cinética , Levalorfano/metabolismo , Levalorfano/farmacología , Ligandos , Pentazocina/metabolismo , Pentazocina/farmacología , Fenilacetatos/síntesis química , Pirrolidinas/síntesis química , Pirrolidinas/farmacología , Receptores sigma/antagonistas & inhibidores , Relación Estructura-Actividad , Especificidad por Sustrato , Receptor Sigma-1RESUMEN
To gather more information on stereochemical factors in the hepatic disposition of organic cations, mass spectrometry coupled to liquid chromatography was used to determine the identity of the metabolites excreted in bile after isolated rat liver perfusions with the quaternary ammonium derivatives of the enantiomeric drugs dextrorphan and levorphanol. Ionspray mass spectrometry was chosen for its soft ionization and absence of thermal degradation of labile compounds. The drugs were labelled with a stable (2H) isotope and mixed with unlabelled drugs to create an artificial isotope pattern in the mass spectrum and facilitate the recognition of unknown metabolites. In mass spectra that were recorded under normal conditions, fragmentation was absent and metabolites of N-methyl dextrorphan and N-methyl levorphanol were visible as parent-ion 'doublets'. Collision-induced fragmentation studies were performed to support the identification of the metabolites. For N-methyl dextrorphan the glucuronide, the glutathione conjugate and the glucuronide of the N-demethylated metabolite were found in bile. For N-methyl levorphanol the glucuronide, the glutathione conjugate, the sulphate conjugate and the glucuronide of a hydroxylated N-methyl levorphanol were excreted in bile. Thus a remarkable stereoselectivity occurs in the metabolism of these quaternary ammonium compounds in the rat liver.
Asunto(s)
Dextrorfano/análisis , Levalorfano/análogos & derivados , Hígado/metabolismo , Animales , Bilis/metabolismo , Cromatografía Liquida , Dextrorfano/análogos & derivados , Técnicas In Vitro , Levalorfano/análisis , Levalorfano/metabolismo , Hígado/química , Espectroscopía de Resonancia Magnética , Masculino , Espectrometría de Masas , Conformación Molecular , Perfusión , Ratas , Ratas Wistar , EstereoisomerismoRESUMEN
1. The effects of the quaternary opioid antagonist N-methyl levallorphan upon cardiovascular responses to non-opioid agonists has been studied in the urethane-anaesthetised rat. 2. N-methyl levallorphan showed an initial nicotinic agonist effect followed by pronounced ganglion blocking activity after intravenous administration. These effects are observed at doses only 2-fold higher than those required to block cardiovascular responses to [Met] enkephalin. 3. The narrow selectivity of N-methyl levallorphan makes this compound inappropriate for opioid antagonist studies where nicotinic receptors may be involved.
Asunto(s)
Bloqueadores Ganglionares , Levalorfano/análogos & derivados , Acetilcolina/farmacología , Anestesia , Animales , Yoduro de Dimetilfenilpiperazina/farmacología , Encefalina Metionina/antagonistas & inhibidores , Semivida , Hemodinámica/efectos de los fármacos , Compuestos de Hexametonio/farmacología , Levalorfano/metabolismo , Levalorfano/farmacocinética , Levalorfano/farmacología , Masculino , Nitroprusiato/farmacología , Norepinefrina/farmacología , Ratas , Ratas Endogámicas , Receptores Opioides/efectos de los fármacos , UretanoRESUMEN
The relative affinities for different subtypes of opioid receptors (mu, kappa and delta) of the peripheral narcotic antagonist N-methyl levallorphan (SR 58002) have been studied in two in vitro smooth muscle systems (guinea-pig ileum and rabbit vas deferens) and by binding studies (guinea-pig brain and cerebellum membranes) using selective tritiated ligands. All the evidence obtained indicates that SR 58002 is a pure antagonist with relative affinity for mu receptors vs kappa and delta superior to that of the parent tertiary compound, levallorphan.
Asunto(s)
Levalorfano/análogos & derivados , Receptores Opioides/metabolismo , Animales , Unión Competitiva , Cerebelo/metabolismo , Cobayas , Íleon/metabolismo , Técnicas In Vitro , Levalorfano/metabolismo , Masculino , Narcóticos/metabolismo , Conejos , Receptores Opioides delta , Receptores Opioides kappa , Receptores Opioides mu , Conducto Deferente/metabolismoRESUMEN
By selectively blocking cross-interferences from other types of binding sites, a binding site which likely represents kappa opioid binding sites was obtained in the guinea-pig brain suspension of the particulate fraction. Selective ligands for mu, sigma, delta and epsilon opioid binding sites were poor inhibitors for inhibiting [3H]ethylketocyclazocine binding to this site, whereas kappa opioids like oxilorphan, dynorphin(1-13), ethylketocyclazocine, butorphanol, cyclazocine, ketocyclazocine, tifluadom, nalorphine, pentazocine, U-50-488, nalbuphine and naloxone were potent ligands. Buprenorphine, generally believed to be a mu opiate, was the most potent inhibitor at the kappa site. Scatchard analysis of the saturation curve of [3H]ethylketocyclazocine binding revealed two subtypes of kappa binding sites: a high-affinity site and a low-affinity site with Kd = 0.7 and 78 nM and maximum binding = 22 and 101 fmol/mg of protein, respectively. Analysis of the inhibition curves suggested that tifluadom may be a selective ligand for the high-affinity site and that dynorphin(1-13) and U-50-488 may bind preferentially the high-affinity site but still possess appreciable affinity for the low-affinity site. This study demonstrates a selective assay for kappa opioid binding sites and indicates a possibility of the heterogeneity of kappa opioid binding sites in the brain.
Asunto(s)
Encéfalo/metabolismo , Receptores Opioides/metabolismo , 3,4-Dicloro-N-metil-N-(2-(1-pirrolidinil)-ciclohexil)-bencenacetamida, (trans)-Isómero , Animales , Benzodiazepinas/metabolismo , Sitios de Unión , Ciclazocina/análogos & derivados , Ciclazocina/metabolismo , Dinorfinas/metabolismo , Encefalina Leucina/análogos & derivados , Encefalina Leucina/metabolismo , Leucina Encefalina-2-Alanina , Etilcetociclazocina , Etorfina/metabolismo , Cobayas , Levalorfano/metabolismo , Fragmentos de Péptidos/metabolismo , Pirrolidinas/metabolismo , Receptores Opioides kappa , EstereoisomerismoAsunto(s)
Bivalvos/metabolismo , Endorfinas/metabolismo , Encefalina Metionina/análogos & derivados , Etorfina/metabolismo , Ganglios/metabolismo , Levalorfano/metabolismo , Morfinanos/metabolismo , Receptores Opioides/metabolismo , Animales , Sitios de Unión , Unión Competitiva , Membrana Celular/metabolismo , D-Ala(2),MePhe(4),Met(0)-ol-encefalina , Encefalina Metionina/metabolismo , CinéticaAsunto(s)
Cuerpo Estriado/metabolismo , Endorfinas/metabolismo , Encefalinas/metabolismo , Receptores Dopaminérgicos/metabolismo , Receptores de Droga/metabolismo , Tranquilizantes/metabolismo , Hormona Adrenocorticotrópica/metabolismo , Animales , Unión Competitiva , Técnicas In Vitro , Levalorfano/metabolismo , Masculino , Fragmentos de Péptidos/metabolismo , Ratas , Espiperona/metabolismo , Relación Estructura-ActividadRESUMEN
Opiate receptor sites in rat brain can be labeled in vivo by [3H]diprenorphine, a potent opiate antagoinst. Using techniques to minimize diffusion in fresh, frozen, unfixed brain, we have localized [3H]diprenorphine by autoradiography to visualize the distribution of opiate receptors. Silver grains indicative of the binding of labeled [3H]diprenorphine are discretely localized in numerous areas of the brain with very high densities in the locus coeruleus, the substantia gelatinosa of the spinal cord, and in clusters within the caudate-putamen, amygdala, and parts of the periventricular gray matter.
Asunto(s)
Encéfalo/metabolismo , Receptores Opioides/metabolismo , Animales , Cerebelo/metabolismo , Ventrículos Cerebrales/metabolismo , Cuerpo Estriado/metabolismo , Diprenorfina/metabolismo , Levalorfano/metabolismo , Masculino , Mesencéfalo/metabolismo , Ratas , Estereoisomerismo , Sustancia Gelatinosa/metabolismoAsunto(s)
Opio/metabolismo , Receptores de Droga , Animales , Autorradiografía , Encéfalo/metabolismo , Ciclopropanos/metabolismo , Tolerancia a Medicamentos , Etorfina/metabolismo , Humanos , Levalorfano/metabolismo , Ratones , Morfina/metabolismo , Naloxona/análogos & derivados , Naloxona/metabolismo , Dolor , Ratas , Estereoisomerismo , Relación Estructura-Actividad , Síndrome de Abstinencia a Sustancias , Trastornos Relacionados con Sustancias/metabolismoAsunto(s)
Encéfalo/metabolismo , Antagonistas de Narcóticos/metabolismo , Narcóticos/metabolismo , Receptores de Droga , Reactivos de Sulfhidrilo/farmacología , Animales , Sitios de Unión/efectos de los fármacos , Cloromercuribenzoatos/farmacología , Etilmaleimida/farmacología , Hidromorfona/metabolismo , Técnicas In Vitro , Yodoacetatos/farmacología , Levalorfano/metabolismo , Levorfanol/metabolismo , Naloxona/metabolismo , Oximorfona/metabolismo , Ratas , Relación Estructura-Actividad , TritioRESUMEN
Tritiated naloxone, a powerful opiate antagonist, specifically binds to an opiate receptor of mammalian brain and guinea pig intestine. Competition for the opiate receptor by various opiates and their antagonists closely parallels their pharmacological potency. The opiate receptor is confined to nervous tissue.