RESUMEN
PURPOSE: To compare rates of prescriber acceptance of interruptive and noninterruptive clinical decision support (CDS) alerts regarding potential diminished therapeutic effectiveness and safety risks associated with proton pump inhibitor (PPI) use in carriers of gene variants affecting cytochrome P450 (CYP) isozyme 2C19 metabolism. METHODS: A retrospective study was conducted at a large rural health system to examine different approaches to improving CDS alert acceptance while minimizing alert fatigue. Manual reviews were conducted to identify alerts regarding CYP2C19 metabolizer status displayed at the time of PPI ordering over 30-day periods before and after the transition from interruptive to noninterruptive CDS alert functionality. A chi-square test was conducted to analyze prescriber acceptance of CDS recommendations by alert modality and type of treatment modification. RESULTS: Overall, interruptive alerts had an acceptance rate of 18.6% (64/344), compared to 8.4% acceptance (30/357 alerts) for noninterruptive alerts (P ≤ 0.0001). Analysis of acceptance criteria -revealed the noninterruptive alert cohort had higher acceptance, as determined by documented medication dose adjustments, than the interruptive alert cohort (53.3% [16/30] and 4.7% [3/64], respectively). The difference in acceptance rates by CDS modality and treatment modification was statistically significant (P ≤ 0.00001). The predominant indication for PPI use was gastroesophageal reflux disease (GERD) in both cohorts. CONCLUSION: Interruptive alerts that actively influenced workflow had higher acceptance rates than noninterruptive alerts that served an informational purpose without a direct disruption of workflow. The study results suggest the utilization of noninterruptive alerts may be a beneficial tool for prompting clinicians to alter dosing regimens rather than transition to an alternative agent.
Asunto(s)
Sistemas de Apoyo a Decisiones Clínicas , Sistemas de Entrada de Órdenes Médicas , Humanos , Citocromo P-450 CYP2C19/química , Citocromo P-450 CYP2C19/efectos de los fármacos , Errores de Medicación , Inhibidores de la Bomba de Protones , Estudios RetrospectivosRESUMEN
Interindividual differences in cytochrome P450 (CYP) 2C19 activity may result in variations in the therapeutic response to drugs metabolized by this enzyme. Differences at gene level may translate into protein level with consequent impairment of the enzyme activity. As a result patients with such genetic differences might experience undesirable effects or no effect at all. The aim of the present study was to find out the prevalence of allelic and genotype frequencies of low activity variants of CYP2C19 genes in healthy individuals from six distinct ethnicities of Pakistan. Blood sample was taken from healthy volunteers following informed consent. Isolation of the DNA was followed by the PCR amplification and restriction fragment length polymorphism. Selected samples were sequenced by Sanger sequencing. The frequency of major alleles was 84.93% for CYP2C19*2 and 91.85% for CYP2C19*3, while minor allele was present at 15.06% for CYP2C19*2 and 8.14% for CYP2C19*3. For CYP2C19*2, the frequency of *1*1 genotype was 75.80%, *1*2 was 18.27%, and *2*2 was 5.92% whereas for CYP2C19*3, The frequency of *1*1 genotype was 84.19%, *1*3 was 15.30%, and *3*3 was 0.49% in the Pakistani population. A substantial variation in genotype and allelic frequencies was observed in various ethnicities. Our study demonstrates that a significant Pakistani population has at least one minor allele, which indicates a large number of patients potentially being affected by these variations. Especially, a significant genotype frequency of PM suggests implication for the treatment response and severity/frequency of adverse effects in patients receiving drugs metabolized by CYP2C19.
Asunto(s)
Citocromo P-450 CYP2C19/genética , Etnicidad , Polimorfismo Genético/genética , Pueblo Asiatico/genética , Pueblo Asiatico/estadística & datos numéricos , Citocromo P-450 CYP2C19/química , Etnicidad/genética , Etnicidad/estadística & datos numéricos , Genotipo , Humanos , Pakistán/epidemiología , FarmacogenéticaRESUMEN
Ethofumesate is a chiral herbicide that may display enantioselective behavior in humans. For this reason, the enantioselective potential of ethofumesate and its main metabolite ethofumesate-2-hydroxy to cause pesticide-drug interactions on cytochrome P450 forms (CYPs) has been evaluated by using human liver microsomes. Among the evaluated CYPs, CYP2C19 had its activity decreased by the ethofumesate racemic mixture (rac-ETO), (+)-ethofumesate ((+)-ETO), and (-)-ethofumesate ((-)-ETO). CYP2C19 inhibition was not time-dependent, but a strong inhibition potential was observed for rac-ETO (IC50 = 5 ± 1 µmol L-1), (+)-ETO (IC50 = 1.6 ± 0.4 µmol L-1), and (-)-ETO (IC50 = 1.8 ± 0.4 µmol L-1). The reversible inhibition mechanism was competitive, and the inhibition constant (Ki) values for rac-ETO (2.6 ± 0.4 µmol L-1), (+)-ETO (1.5 ± 0.2 µmol L-1), and (-)-ETO (0.7 ± 0.1 µmol L-1) were comparable to the Ki values of strong CYP2C19 inhibitors. Inhibition of CYP2C19 by ethofumesate was enantioselective, being almost twice higher for (-)-ETO than for (+)-ETO, which indicates that this enantiomer may be a more potent inhibitor of this CYP form. For an in vitro-in vivo correlation, the Food and Drug Administration's (FDA) guideline on the assessment of drug-drug interactions used in the early stages of drug development was used. The FDA's R1 values were estimated on the basis of the obtained ethofumesate Ki and distribution volume, metabolism, unbound plasma fraction, gastrointestinal and dermal absorption data available in the literature. The correlation revealed that ethofumesate probably inhibits CYP2C19 in vivo for both chronic (oral) and occupational (dermal) exposure scenarios.
Asunto(s)
Benzofuranos/química , Benzofuranos/farmacología , Inhibidores del Citocromo P-450 CYP2C19/química , Inhibidores del Citocromo P-450 CYP2C19/farmacología , Citocromo P-450 CYP2C19/metabolismo , Mesilatos/química , Mesilatos/farmacología , Plaguicidas/química , Plaguicidas/farmacología , Citocromo P-450 CYP2C19/química , Inhibidores del Citocromo P-450 CYP2C19/metabolismo , Relación Dosis-Respuesta a Droga , Descubrimiento de Drogas , Humanos , Simulación del Acoplamiento Molecular , Unión Proteica , Conformación Proteica , EstereoisomerismoRESUMEN
7,8-dihydroxyflavone (DHF) is a flavone aglycone which has beneficial effects in several central nervous system diseases. Most of the pharmacokinetic properties of DHF have been characterized, while only limited information is available regarding its interactions with serum albumin and biotransformation enzymes. In this study, the interactions of DHF with albumin was examined employing fluorescence spectroscopy and ultrafiltration. Furthermore, the inhibitory effects of DHF on cytochrome P450 (CYP2C9, CYP2C19, and CYP3A4) and xanthine oxidase (XO) enzymes were also tested using in vitro models. Our results demonstrate that DHF forms a stable complex with albumin (K = 4.9 × 105 L/mol) and that it is able to displace both Site I and Site II ligands. Moreover, DHF proved to be a potent inhibitor of each enzyme tested, showing similar or slightly weaker effects than the positive controls used. Considering the above-listed observations, the coadministration of DHF with drugs may interfere with the drug therapy due to the development of pharmacokinetic interactions.
Asunto(s)
Citocromo P-450 CYP2C19/química , Citocromo P-450 CYP2C9/química , Citocromo P-450 CYP3A/química , Flavonas/química , Albúmina Sérica Humana/química , Xantina Oxidasa/química , BiotransformaciónRESUMEN
The human cytochrome P450 (CYP) 2C9 and 2C19 enzymes are two highly similar isoforms with key roles in drug metabolism. They are anchored to the endoplasmic reticulum membrane by their N-terminal transmembrane helix and interactions of their cytoplasmic globular domain with the membrane. However, their crystal structures were determined after N-terminal truncation and mutating residues in the globular domain that contact the membrane. Therefore, the CYP-membrane interactions are not structurally well-characterized and their dynamics and the influence of membrane interactions on CYP function are not well understood. We describe herein the modeling and simulation of CYP 2C9 and CYP 2C19 in a phospholipid bilayer. The simulations revealed that, despite high sequence conservation, the small sequence and structural differences between the two isoforms altered the interactions and orientations of the CYPs in the membrane bilayer. We identified residues (including K72, P73, and I99 in CYP 2C9 and E72, R73, and H99 in CYP 2C19) at the protein-membrane interface that contribute not only to the differing orientations adopted by the two isoforms in the membrane, but also to their differing substrate specificities by affecting the substrate access tunnels. Our findings provide a mechanistic interpretation of experimentally observed effects of mutagenesis on substrate selectivity.
Asunto(s)
Citocromo P-450 CYP2C19/química , Citocromo P-450 CYP2C9/química , Fosfolípidos/metabolismo , Sitios de Unión , Citocromo P-450 CYP2C19/metabolismo , Citocromo P-450 CYP2C9/metabolismo , Humanos , Membranas Intracelulares/metabolismo , Simulación del Acoplamiento Molecular , Unión ProteicaRESUMEN
Helicobacter pylori (H. pylori) infection is common and can result in gastric and duodenal ulcers, and in some cases, gastric lymphoma and cancer. Omeprazole (OMP)-in combination with clarithromycin (CLR), amoxicillin (AMX), tinidazole (TND), or metronidazole (MET)-is used in double or triple combination therapy for eradication of H. pylori. However, the roles of the drugs other than OMP are not clearly understood. Therefore, in the present study, we aimed to investigate any effects of these drugs on OMP metabolism by wild-type CYP2C19 using spectroscopy and enzyme kinetics. The dissociation constants (Kd) for CYP2C19 with OMP, CLR, AMX, TND, and MET were 8.6, 126, 156, 174, and 249 µM, respectively. The intrinsic clearance of OMP was determined to be 355 mL/min/µmol of CYP2C19. Metabolism of OMP was significantly inhibited by 69, 66, 28, and 40% in the presence of CLR, TND, AMX, and MET, respectively. Moreover, the combination of CLR and TND resulted in 76% inhibition of OMP metabolism, while the combination of AMX and MET resulted in 48% inhibition of OMP metabolism. Both combinations of drugs not only have antibacterial effects, but also enhance the effect of OMP by inhibiting its metabolism by CYP2C19. These results indicate that drug-drug interactions of co-administered drugs can cause complex effects, providing a basis for OMP dose adjustment when used in combination therapy for H. pylori eradication.
Asunto(s)
Antibacterianos/farmacología , Citocromo P-450 CYP2C19/metabolismo , Infecciones por Helicobacter/tratamiento farmacológico , Helicobacter pylori/efectos de los fármacos , Omeprazol/farmacología , Amoxicilina/química , Amoxicilina/farmacología , Antibacterianos/química , Antibacterianos/metabolismo , Cromatografía Líquida de Alta Presión , Claritromicina/química , Claritromicina/farmacología , Citocromo P-450 CYP2C19/química , Combinación de Medicamentos , Humanos , Metronidazol/química , Metronidazol/farmacología , Pruebas de Sensibilidad Microbiana , Estructura Molecular , Omeprazol/antagonistas & inhibidores , Omeprazol/metabolismo , Tinidazol/química , Tinidazol/farmacologíaRESUMEN
Protein-ligand docking is a widely used method to generate solutions for the binding of a small molecule with its target in a short amount of time. However, these methods provide identification of physically sound protein-ligand complexes without a complete view of the binding process dynamics, which has been recognized to be a major discriminant in binding affinity and ligand selectivity. In this paper, a novel piece of open-source software to approach this problem is presented, called GPathFinder. It is built as an extension of the modular GaudiMM platform and is able to simulate ligand diffusion pathways at atomistic level. The method has been benchmarked on a set of 20 systems whose ligand-binding routes were studied by other computational tools or suggested from experimental "snapshots". In all of this set, GPathFinder identifies those channels that were already reported in the literature. Interestingly, the low-energy pathways in some cases indicate novel possible binding routes. To show the usefulness of GPathFinder, the analysis of three case systems is reported. We believe that GPathFinder is a software solution with a good balance between accuracy and computational cost, and represents a step forward in extending protein-ligand docking capacities, with implications in several fields such as drug or enzyme design.
Asunto(s)
Simulación del Acoplamiento Molecular/métodos , Programas Informáticos , Algoritmos , Acuaporinas/química , Acuaporinas/metabolismo , Proteínas Arqueales/química , Proteínas Arqueales/metabolismo , Sitios de Unión , Citocromo P-450 CYP2C19/química , Citocromo P-450 CYP2C19/metabolismo , Humanos , Indolamina-Pirrol 2,3,-Dioxigenasa/química , Indolamina-Pirrol 2,3,-Dioxigenasa/metabolismo , Ligandos , Unión ProteicaRESUMEN
Cytochrome P450 (CYP) is a class of heme-containing enzymes which mainly catalyze a monooxygenation reaction of various chemicals, and hence CYP plays a key role in the drug metabolism. Although CYP2C19 isoform is a minor hepatic CYP, it metabolizes clinically important drugs such as omeprazole and Smephenytoin. In this work, the interaction of purified CYP2C19 WT (CYP2C19) with seven drugs (phenytoin, Smephenytoin, omeprazole, lansoprazole, cimetidine, propranolol, and warfarin) was investigated using spectroscopic methods. The binding of each drug and the induced structural change in the heme distal environment were evaluated. Ferric form of CYP2C19 was revealed to contain a six-coordinate low-spin heme with a water molecule as a sixth ligand in a distal site, and the addition of each drug caused varied minor fraction of five-coordinate heme. It was suggested that the ligated water molecule was partly moved away from the heme distal environment and that the degree of water removal was dependent on the type of drugs. The effect on the coordination was varied with the studied drugs with wide variation in the dissociation constants from 2.6⯵M for lansoprazole to 5400⯵M for warfarin. Phenytoin and Smephenytoin showed that binding to CYP2C19 occurred in a stepwise manner and that the coordination of a water molecule was facilitated in the second binding step. In the ferrous CO-bound state, ν(FeCO) stretching mode was clearly observed at 471â¯cm-1 in the absence of drugs. The Raman line was greatly up-shifted by omeprazole (487â¯cm-1) and lansoprazole (477â¯cm-1) but was minimally affected by propranolol, phenytoin, and Smephenytoin. These results indicate that slight chemical modification of a drug greatly affects the heme distal environments upon binding.
Asunto(s)
Citocromo P-450 CYP2C19/metabolismo , Compuestos Ferrosos/metabolismo , Hemo/metabolismo , Preparaciones Farmacéuticas/metabolismo , Espectrofotometría Ultravioleta/métodos , Espectrometría Raman/métodos , Sitios de Unión , Citocromo P-450 CYP2C19/química , Compuestos Ferrosos/química , Hemo/química , Humanos , Ligandos , Oxidación-Reducción , Preparaciones Farmacéuticas/química , Conformación ProteicaRESUMEN
Two chemotypes were examined in vitro with CYPs 3A4 and 2C19 by molecular docking, metabolic profiles, and intrinsic clearance deuterium isotope effects with specifically deuterated form to assess the potential for enhancement of pharmacokinetic parameters. The results show the complexity of deuteration as an approach for pharmacokinetic enhancement when CYP enzymes are involved in metabolic clearance. With CYP3A4 the rate limiting step was chemotype-dependent. With one chemotype no intrinsic clearance deuterium isotope effect was observed with any deuterated form, whereas with the other chemotype the rate limiting step was isotopically sensitive, and the magnitude of the intrinsic clearance isotope effect was dependent on the position(s) and extent of deuteration. Molecular docking and metabolic profiles aided in identifying sites for deuteration and predicted the possibility for metabolic switching. However, the potential for an isotope effect on the intrinsic clearance cannot be predicted and must be established by examining select deuterated versions of the chemotypes. The results show how in a deuteration strategy molecular docking, in-vitro metabolic profiles, and intrinsic clearance assessments with select deuterated versions of new chemical entities can be applied to determine the potential for pharmacokinetic enhancement in a discovery setting. They also help explain the substantial failures reported in the literature of deuterated versions of drugs to elicit a systemic enhancement on pharmacokinetic parameters.
Asunto(s)
Citocromo P-450 CYP2C19/química , Citocromo P-450 CYP3A/química , Deuterio/química , Farmacocinética , Citocromo P-450 CYP2C19/efectos de la radiación , Citocromo P-450 CYP3A/efectos de la radiación , Deuterio/farmacología , Hemo/química , Hemo/efectos de la radiación , Humanos , Inactivación Metabólica , Cinética , Microsomas/efectos de la radiación , Simulación del Acoplamiento Molecular , Oxidación-Reducción/efectos de la radiación , Especificidad por SustratoRESUMEN
Sauchinone, an active lignan isolated from the aerial parts of Saururus chinensis (Saururaceae), exhibits anti-inflammatory, anti-obesity, anti-hyperglycemic, and anti-hepatic steatosis effects. As herb-drug interaction (HDI) through cytochrome P450s (CYPs)-mediated metabolism limits clinical application of herbs and drugs in combination, this study sought to explore the enzyme kinetics of sauchinone towards CYP inhibition in in vitro human liver microsomes (HLMs) and in vivo mice studies and computational molecular docking analysis. In in vitro HLMs, sauchinone reversibly inhibited CYP2B6, 2C19, 2E1, and 3A4 activities in non-competitive modes, showing inhibition constant (Ki) values of 14.3, 16.8, 41.7, and 6.84 µM, respectively. Also, sauchinone time-dependently inhibited CYP2B6, 2E1 and 3A4 activities in vitro HLMs. Molecular docking study showed that sauchinone could be bound to a few key amino acid residues in the active site of CYP2B6, 2C19, 2E1, and 3A4. When sibutramine, clopidogrel, or chlorzoxazone was co-administered with sauchinone to mice, the systemic exposure of each drug was increased compared to that without sauchinone, because sauchinone reduced the metabolic clearance of each drug. In conclusion, when sauchinone was co-treated with drugs metabolized via CYP2B6, 2C19, 2E1, or 3A4, sauchinone-drug interactions occurred because sauchinone inhibited the CYP-mediated metabolic activities.
Asunto(s)
Benzopiranos/química , Citocromo P-450 CYP2B6/química , Citocromo P-450 CYP2C19/química , Citocromo P-450 CYP2E1/química , Citocromo P-450 CYP3A/química , Dioxoles/química , Interacciones de Hierba-Droga , Saururaceae/química , Animales , Antiinflamatorios/química , Antiinflamatorios/aislamiento & purificación , Antiinflamatorios/farmacología , Fármacos Antiobesidad/química , Fármacos Antiobesidad/aislamiento & purificación , Fármacos Antiobesidad/farmacología , Benzopiranos/aislamiento & purificación , Benzopiranos/farmacología , Sitios de Unión , Dominio Catalítico , Clorzoxazona/química , Clorzoxazona/farmacología , Clopidogrel , Ciclobutanos/química , Ciclobutanos/farmacología , Citocromo P-450 CYP2B6/metabolismo , Citocromo P-450 CYP2C19/metabolismo , Citocromo P-450 CYP2E1/metabolismo , Citocromo P-450 CYP3A/metabolismo , Inhibidores Enzimáticos del Citocromo P-450/química , Inhibidores Enzimáticos del Citocromo P-450/aislamiento & purificación , Inhibidores Enzimáticos del Citocromo P-450/farmacología , Dioxoles/aislamiento & purificación , Dioxoles/farmacología , Humanos , Hipoglucemiantes/química , Hipoglucemiantes/aislamiento & purificación , Hipoglucemiantes/farmacología , Cinética , Ratones , Microsomas Hepáticos/química , Microsomas Hepáticos/efectos de los fármacos , Microsomas Hepáticos/enzimología , Simulación del Acoplamiento Molecular , Componentes Aéreos de las Plantas/química , Extractos Vegetales/química , Extractos Vegetales/aislamiento & purificación , Extractos Vegetales/farmacología , Unión Proteica , Dominios y Motivos de Interacción de Proteínas , Estructura Secundaria de Proteína , Ticlopidina/análogos & derivados , Ticlopidina/química , Ticlopidina/farmacologíaRESUMEN
2,2',3,5',6-Pentachlorobiphenyl (PCB 95) and 2,2',3,4,4',5',6-heptachlorobiphenyl (PCB 183) possess axial chirality and form the aS and aR enantiomers. The enantiomers of these congeners have been reported to accumulate in the human body enantioselectively via unknown mechanisms. In this study, we determined the cytochrome P450 (CYP) monooxygenase responsible for the enantioselective oxidization of PCB 95 and PCB 183, using a recombinant human CYP monooxygenase. We evaluated 13 CYP monooxygenases, namely CYP1A1, CYP1A2, CYP1B1, CYP2A6, CYP2B6, CYP2C8, CYP2C19, CYP2E1, CYP2J2, CYP3A4, CYP3A5, CYP4F2, and aromatase (CYP19), and revealed that CYP2A6 preferably oxidizes aS-PCB 95 enantioselectively; however, it did not oxidize PCB 183. The enantiomer composition was elevated from 0.5 (racemate) to 0.54. In addition, following incubation with CYP2A6, the enantiomer fraction (EF) of PCB 95 demonstrated a time-dependent increase.
Asunto(s)
Citocromo P-450 CYP1A1/química , Citocromo P-450 CYP1A2/química , Citocromo P-450 CYP2C19/química , Sistema Enzimático del Citocromo P-450/química , Bifenilos Policlorados/química , Catálisis , Citocromo P-450 CYP1A1/metabolismo , Citocromo P-450 CYP1A2/metabolismo , Citocromo P-450 CYP2C19/metabolismo , Sistema Enzimático del Citocromo P-450/metabolismo , Humanos , Oxidación-Reducción , EstereoisomerismoRESUMEN
Cytochromes P450 (CYP) are involved in numerous biochemical processes including metabolism of xenobiotics, biosynthesis of cholesterol, steroid hormones etc. Since some CYP catalyze indol oxidation to isatin, we have hypothesized that isatin can regulate protein-protein interactions (PPI) between components of the CYP system thus representing a (negative?) feedback mechanism. The aim of this study was to investigate a possible effect of isatin on interaction of human CYP with cytochrome b5 (CYB5A). Using the optical biosensor test system employing surface plasmon resonance (SPR) we have investigated interaction of immobilized CYB5A with various CYP in the absence and in the presence of isatin. The SPR-based experiments have shown that a high concentration of isatin (270 mM) increases Kd values for complexes CYB5A/CYP3Ð5 and CYB5A/CYP3A4 (twofold and threefold, respectively), but has no influence on complex formation between CYB5A and other CYP (including indol-metabolizing CYP2C19 and CYP2E1). Isatin injection to the optical biosensor chip with the preformed molecular complex CYB5A/CYP3A4 caused a 30%-increase in its dissociation rate. Molecular docking manipulations have shown that isatin can influence interaction of CYP3Ð5 or CYP3A4 with CYB5A acting at the contact region of CYB5A/CYP.
Asunto(s)
Citocromo P-450 CYP2C19/química , Citocromo P-450 CYP2E1/química , Citocromo P-450 CYP3A/química , Citocromos b5/química , Isatina/química , Sitios de Unión , Enzima de Desdoblamiento de la Cadena Lateral del Colesterol/química , Citocromo P-450 CYP2C9/química , Humanos , Cinética , Simulación del Acoplamiento Molecular , Unión Proteica , Conformación Proteica en Hélice alfa , Conformación Proteica en Lámina beta , Dominios y Motivos de Interacción de Proteínas , Proteínas Recombinantes/química , Soluciones , Esteroide 11-beta-Hidroxilasa/química , Esteroide 17-alfa-Hidroxilasa/química , Resonancia por Plasmón de SuperficieRESUMEN
Pharmacogenomics harnesses the utility of a patient's genome (n = 1) in decisions on which therapeutic drugs and in what amounts should be administered. Often, patients with shared ancestry present with comparable genetic profiles that predict drug response. However, populations are not static, thus, often, population mobility through migration, especially enmasse as is seen for refugees, changes the pharmacogenetic profiles of resultant populations and therefore observed responses to commonly used therapeutic drugs. For example, in the aftermath of the Syrian civil war since 2011, millions have fled their homes to neighboring countries in the Middle East. The growing permanence of refugees and mass migrations is a call to shift our focus in the life sciences community from old models of pharmaceutical innovation. These seismic social changes demand faster decisions for "population-to-population bridging," whereby novel drugs developed in or for particular regions/countries can meet with rational regulatory decisions/approval in world regions impacted by migrant/refugee populations whose profiles are dynamic, such as in the Eastern Mediterranean region at present. Thus, it is important to characterize and report on the prevalence of pharmacogenes that affect commonly used medications and predict if population changes may call for attention to particular differences that may impact health of patients. Thus, we report here on four single-nucleotide polymorphism (SNP) variations in CYP2C9 and CYP2C19 genes among Mersin-Turkish healthy volunteers in the Mersin Province in the Eastern Mediterranean region that is currently hosting a vast number of migrant populations from Syria. Both CYP2C9 and CYP2C19 are very important pharmacogene molecular targets. We compare and report here on the observed SNP genetic variation in our sample with data on 12 world populations from dbSNP and discuss the feasibility of forecasting the pharmacokinetics of drugs utilized by migrant communities in Mersin and the Eastern Mediterranean region. This study can serve as a catalyst to invest in research in Syrian populations currently living in the Eastern Mediterranean. The findings have salience for rapid and rational regulatory decision-making for worldwide precision medicine and, specifically, "pharmacogenovigilance-guided bridging of pharmacokinetics" across world populations in the current era of planetary scale migration.
Asunto(s)
Citocromo P-450 CYP2C19/genética , Citocromo P-450 CYP2C9/genética , Efectos Colaterales y Reacciones Adversas Relacionados con Medicamentos/genética , Variantes Farmacogenómicas , Medicina de Precisión/métodos , Citocromo P-450 CYP2C19/química , Citocromo P-450 CYP2C9/química , Humanos , Región Mediterránea , Pruebas de Farmacogenómica , Farmacovigilancia , Polimorfismo de Nucleótido Simple , Siria/etnología , MigrantesRESUMEN
Racemic omeprazole has been used for clinically treating gastric acid-related diseases and also as a typical human cytochrome P450 (P450) 2C19 probe substrate in preclinical studies. S-Omeprazole has been developed as a single enantiomer medicine, which has been reported not to be associated with polymorphic human P450 2C19 phenotypes. In this study, 5-hydroxylation and sulfoxidation activities, with respect to stereoselective R- and S-omeprazole oxidations by liver microsomes from experimental animals including non-human primates and humans, were investigated in vitro. Liver microsomes from humans, cynomolgus monkeys, and mice preferentially mediated R-omeprazole 5-hydroxylations, however those from marmosets, minipigs, dogs, and rats preferentially mediated S-omeprazole 5-hydroxylations. High catalytic activities were observed for recombinant human P450 2C19 in R-omeprazole 5-hydroxlations, cynomolgus monkey P450 2C19 in both R- and S-omeprazole 5-hydroxlations, and marmoset P450 2C19 in S-omeprazole 5-hydroxlations. On the other hand, human, cynomolgus monkey, and marmoset P450 3A enzymes preferentially mediated S-omeprazole sulfoxidations. Correlation and kinetic analyses revealed a high affinity of polymorphic cynomolgus monkey and marmoset liver microsomal P450 2C19 enzymes with respect to R- and S-omeprazole 5-hydroxylations, respectively, and a high capacity of cynomolgus monkey and marmoset liver microsomal P450 3A4 for omeprazole 5-hydroxylations and sulfoxidations. R-and S-omeprazole 5-hydroxylation activities in cynomolgus monkey and marmoset liver microsomes were significantly different among wild-type, heterozygous, and homozygous animals genotyped for cynomolgus monkey P450 2C19 p.[(Phe100Asn; Ala103Val; Ile112Leu)] and for marmoset P450 2C19 p.[(Phe7Leu; Ser254Leu; Ile469Thr)], respectively. The results of this study demonstrate polymorphic cynomolgus monkey and marmoset P450 2C19-dependent omeprazole oxidation activities with individual variations similar to humans.
Asunto(s)
Antiulcerosos/metabolismo , Citocromo P-450 CYP2C19/metabolismo , Microsomas Hepáticos/enzimología , Omeprazol/metabolismo , Polimorfismo Genético , Inhibidores de la Bomba de Protones/metabolismo , Animales , Antiulcerosos/química , Biocatálisis , Biotransformación , Callithrix , Citocromo P-450 CYP2C19/química , Citocromo P-450 CYP2C19/genética , Citocromo P-450 CYP3A/química , Citocromo P-450 CYP3A/genética , Citocromo P-450 CYP3A/metabolismo , Perros , Humanos , Hidroxilación , Macaca fascicularis , Ratones , Microsomas Hepáticos/metabolismo , Simulación del Acoplamiento Molecular , Omeprazol/química , Oxidación-Reducción , Inhibidores de la Bomba de Protones/química , Ratas , Proteínas Recombinantes/química , Proteínas Recombinantes/metabolismo , Especificidad de la Especie , Estereoisomerismo , Porcinos , Porcinos EnanosRESUMEN
Cytochrome P450 2C19 (CYP2C19) is one of 57 drug metabolizing enzymes in humans and is responsible for the metabolism of â¼7-10% of drugs in clinical use. Recently omeprazole-based analogues were reported to be the potent inhibitors of CYP2C19 and have the potential to be used as the tool compounds for studying the substrate selectivity of CYP2C19. However, the binding modes of these compounds with CYP2C19 remain to be elucidated. In this study, a combination of molecular docking, molecular dynamics (MD), and MM/GBSA calculations was employed to systematically investigate the interactions between these compounds and CYP2C19. The binding modes of these analogues were analyzed in detail. The results indicated that the inclusion of explicit active site water molecules could improve binding energy prediction when the water molecules formed a hydrogen bonding network between the ligand and protein. We also found that the effect of active site water molecules on binding free energy prediction was dependent on the ligand binding modes. Our results unravel the interactions of these omeprazole-based analogues with CYP2C19 and might be helpful for the future design of potent CYP2C19 inhibitors with improved metabolic properties.
Asunto(s)
Citocromo P-450 CYP2C19/química , Simulación del Acoplamiento Molecular , Simulación de Dinámica Molecular , Omeprazol/análogos & derivados , Omeprazol/química , Algoritmos , Sitios de Unión , Dominio Catalítico , Humanos , Ligandos , Conformación Molecular , Estructura Molecular , Omeprazol/farmacología , Unión ProteicaRESUMEN
Structure-based prediction of sites of metabolism (SOMs) mediated by cytochrome P450s (CYPs) is of great interest in drug discovery and development. However, protein flexibility and active site water molecules remain a challenge for accurate SOM prediction. CYP2C19 is one of the major drug-metabolizing enzymes and has attracted considerable attention because of its polymorphism and capability of metabolizing â¼7% clinically used drugs. In this study, we systematically evaluated the effects of protein flexibility and active site water molecules on SOM prediction for CYP2C19 substrates. Multiple conformational sampling techniques including GOLD flexible residues sampling, molecular dynamics (MD) and tCONCOORD side-chain sampling were adopted for assessing the influence of protein flexibility on SOM prediction. The prediction accuracy could be significantly improved when protein flexibility was considered using the tCONCOORD sampling method, which indicated that the side-chain conformation was important for accurate prediction. However, the inclusion of the crystallographic or MD-derived water molecule(s) does not necessarily improve the prediction accuracy. Finally, a combination of docking results with SMARTCyp was found to be able to increase the SOM prediction accuracy.
Asunto(s)
Dominio Catalítico , Citocromo P-450 CYP2C19/química , Citocromo P-450 CYP2C19/metabolismo , Agua/química , Cristalografía por Rayos X , Bases de Datos de Proteínas , Simulación del Acoplamiento Molecular , Reproducibilidad de los Resultados , Especificidad por SustratoRESUMEN
The in vitro metabolism of (-)-cis- and (-)-trans-rose oxide was investigated using human liver microsomes and recombinant cytochrome P450 (P450 or CYP) enzymes for the first time. Both isomers of rose oxide were incubated with human liver microsomes, and the formation of the respective 9-oxidized metabolite were determined using gas chromatography-mass spectrometry (GC-MS). Of 11 different recombinant human P450 enzymes used, CYP2B6 and CYP2C19 were the primary enzymes catalysing the metabolism of (-)-cis- and (-)-trans-rose oxide. CYP1A2 also efficiently oxidized (-)-cis-rose oxide at the 9-position but not (-)-trans-rose oxide. α-Naphthoflavone (a selective CYP1A2 inhibitor), thioTEPA (a CYP2B6 inhibitor) and anti-CYP2B6 antibody inhibited (-)-cis-rose oxide 9-hydroxylation catalysed by human liver microsomes. On the other hand, the metabolism of (-)-trans-rose oxide was suppressed by thioTEPA and anti-CYP2B6 at a significant level in human liver microsomes. However, omeprazole (a CYP2C19 inhibitor) had no significant effects on the metabolism of both isomers of rose oxide. Using microsomal preparations from nine different human liver samples, (-)-9-hydroxy-cis- and (-)-9-hydroxy-trans-rose oxide formations correlated with (S)-mephenytoin N-demethylase activity (CYP2B6 marker activity). These results suggest that CYP2B6 plays important roles in the metabolism of (-)-cis- and (-)-trans-rose oxide in human liver microsomes.
Asunto(s)
Citocromo P-450 CYP2B6/metabolismo , Citocromo P-450 CYP2C19/metabolismo , Aditivos Alimentarios/metabolismo , Microsomas Hepáticos/enzimología , Monoterpenos/metabolismo , Perfumes/metabolismo , Monoterpenos Acíclicos , Animales , Anticuerpos Monoclonales/farmacología , Línea Celular , Citocromo P-450 CYP2B6/química , Citocromo P-450 CYP2B6/genética , Citocromo P-450 CYP2C19/química , Citocromo P-450 CYP2C19/genética , Inhibidores Enzimáticos del Citocromo P-450/farmacología , Aditivos Alimentarios/química , Cromatografía de Gases y Espectrometría de Masas , Humanos , Hidroxilación/efectos de los fármacos , Cinética , Microsomas Hepáticos/efectos de los fármacos , Estructura Molecular , Monoterpenos/química , Mariposas Nocturnas , Odorantes , Perfumes/química , Proteínas Recombinantes/química , Proteínas Recombinantes/metabolismo , Estereoisomerismo , Especificidad por SustratoRESUMEN
A rapid selective and sensitive liquid chromatography/tandem mass spectrometry (LC-MS/MS) method was developed for the quantitative determination of derivatised cytochrome P450-2C19 oxidation product (dansyl-4-OH mephenytoin) and its underivatised form (4-OH mephenytoin). Samples were anaysed on C18 column (Waters Xbridge, 50 mm×4.6 mm, 3.5 µm particle size) with the mobile phase consisting of 0.1% formic acid in water and 0.1% formic acid in acetonitrile. A gradient method with a short run time of 2.5 min and 3.5 min was developed for the analysis of dansyl-4-OH mephenytoin and 4-OH mephenytoin, respectively. The standard curve was linear (r(2)=0.9972 for 4-OH mephenytoin; r(2)=0.9946 for dansyl-4-OH mephenytoin) over the concentration range of 0.16 to 40 ng/mL for both derivatised and underivatised forms. The CV (%) and relative error (RE) for inter and intraassay at three QC levels for dansyl-4-OH mephenytoin was 0.97-5.85% and -9.80 to 2.51%, respectively. Whereas, for 4-OH mephenytoin the CV (%) and RE (%) at three QC levels was 0.82-3.47% and -6.69 to -0.01%, respectively. The developed method was validated for various parameters such as linearity, precision & accuracy, extraction recovery, matrix effect, autosampler stability and was proved to be consistent across three QC levels with overall CV (%) less than 15. Dansylation helped in increasing the sensitivity of hydroxy mephenytoin by 100-200 fold. Given the simplicity involved in derivatisation process, we believe that this novel methodology will change the current approaches used for the enhancing the detection sensitivity of 4-OH mephenytoin.
Asunto(s)
Citocromo P-450 CYP2C19/química , Compuestos de Dansilo/química , Pruebas de Enzimas/métodos , Inhibidores Enzimáticos/química , Mefenitoína/química , Cinética , Oxidación-ReducciónRESUMEN
OBJECTIVES: This study aimed to investigate the effects of five tanshinones, the lipophilic components from Danshen (Salvia miltiorrhiza), on CYP2C19 activity in pooled human liver microsomes (HLMs). METHODS: The effects of tanshinones on CYP2C19 activity were compared by enzyme inhibition study using omeprazole 5-hydroxylation in pooled HLMs. The inhibition constant (Ki) values and inhibition modes of effective tanshinones were evaluated by enzyme kinetic study. Molecular docking analysis was used to simulate the binding conformations of tanshinones to the active cavity of human CYP2C19. RESULTS: Dihydrotanshinone and miltirone showed potent inhibitory effects on CYP2C19 activity in a concentration-dependent manner. Tanshinone I showed weaker inhibitory effect, whereas tanshinone IIA and cryptotanshinone had no inhibitory effect. Further enzyme kinetic study showed that the inhibition by dihydrotanshinone and miltirone was a mixed type. The effects of tanshinones were also confirmed by a molecular docking study. Besides, the ethanol extract of Danshen also showed a mixed type of inhibition, whereas the water extract had no inhibitory effect. CONCLUSIONS: The current findings demonstrate the inhibition of CYP2C19 activity by the ethanol extract of Danshen and its components tanshinones, implicating the potential herb-drug interactions between Danshen and therapeutic agents metabolized by CYP2C19 in clinical practice.
Asunto(s)
Abietanos/farmacología , Inhibidores del Citocromo P-450 CYP2C19/farmacología , Citocromo P-450 CYP2C19/metabolismo , Microsomas Hepáticos/efectos de los fármacos , Salvia miltiorrhiza/química , Dominio Catalítico , Simulación por Computador , Citocromo P-450 CYP2C19/química , Inhibidores del Citocromo P-450 CYP2C19/química , Inhibidores del Citocromo P-450 CYP2C19/metabolismo , Medicamentos Herbarios Chinos/química , Medicamentos Herbarios Chinos/farmacología , Etanol , Interacciones de Hierba-Droga , Humanos , Hidroxilación , Cinética , Microsomas Hepáticos/enzimología , Microsomas Hepáticos/metabolismo , Simulación del Acoplamiento Molecular , Fenantrenos/química , Fenantrenos/metabolismo , Fenantrenos/farmacologíaRESUMEN
The binding and cytochrome P45051 (CYP51) inhibition properties of a novel antifungal compound, VT-1161, against purified recombinant Candida albicans CYP51 (ERG11) and Homo sapiens CYP51 were compared with those of clotrimazole, fluconazole, itraconazole, and voriconazole. VT-1161 produced a type II binding spectrum with Candida albicans CYP51, characteristic of heme iron coordination. The binding affinity of VT-1161 for Candida albicans CYP51 was high (dissociation constant [Kd], ≤ 39 nM) and similar to that of the pharmaceutical azole antifungals (Kd, ≤ 50 nM). In stark contrast, VT-1161 at concentrations up to 86 µM did not perturb the spectrum of recombinant human CYP51, whereas all the pharmaceutical azoles bound to human CYP51. In reconstitution assays, VT-1161 inhibited Candida albicans CYP51 activity in a tight-binding fashion with a potency similar to that of the pharmaceutical azoles but failed to inhibit the human enzyme at the highest concentration tested (50 µM). In addition, VT-1161 (MIC = 0.002 µg ml(-1)) had a more pronounced fungal sterol disruption profile (increased levels of methylated sterols and decreased levels of ergosterol) than the known CYP51 inhibitor voriconazole (MIC = 0.004 µg ml(-1)). Furthermore, VT-1161 weakly inhibited human CYP2C9, CYP2C19, and CYP3A4, suggesting a low drug-drug interaction potential. In summary, VT-1161 potently inhibited Candida albicans CYP51 and culture growth but did not inhibit human CYP51, demonstrating a >2,000-fold selectivity. This degree of potency and selectivity strongly supports the potential utility of VT-1161 in the treatment of Candida infections.