Activity Of Cytochrome P450 Isoenzymes Research Articles

Characterisation of cytochrome P450 isoenzyme activity in sheep liver and placental microsomes

IntroductionDrug metabolism during pregnancy is a complex process that involves maternal, placental and fetal sites of metabolism. Indeed, there is a lack of clarity provided from drug metabolism in human pregnancy due to ethical limitations. Large animal models of human pregnancy provide an opportunity to quantify activity of phase 1 drug metabolism mediated by cytochrome P450 (CYP) enzymes in the maternal, placental, and fetal compartments. Herein, we have validated a comprehensive assay to quantify maternal, placental, and fetal CYP activity. MethodsIsolated microsomes from sheep maternal liver, placenta, and fetal liver (140d gestation, term = 150d) were incubated with CYP-specific probe drugs to quantify the activity of CYP1A2, CYP2B6, CYP2C8, CYP2C9, CYP2C19, CYP2D6, CYP2E1 and CYP3A. Inhibition studies were performed to validate specificity of probe drugs. The validated assay was developed using liquid chromatography-tandem mass spectrometry (LC-MS/MS). ResultsCYP1A2, CYP2B6, CYP2C8, CYP2C19, CYP2D6, CYP2E1 and CYP3A were active in maternal liver. In contrast, only CYP1A2, CYP2C8 and CYP2D6 were active in the placenta, whereas CYP2B6, CYP2C8 and CYP2D6 were active in the fetal liver. Of the placental-specific CYPs validated, CYP1A2 increased in type A compared with type D placentomes, whereas CYP2C8 activity increased in type B compared with type A and C. DiscussionThis study has established conditions for compartment-specific CYP activity in the sheep maternal-placental-fetal unit using a validated and standardised experimental workflow. Compartment- and placentome type-specific CYP activity are important considerations when examining drug metabolism in the maternal-placental-fetal unit and in determining the impact of pregnancy complications.

РАЗРАБОТКА СОСТАВА И ТЕХНОЛОГИИ КОКТЕЙЛЬНОЙ ТЕСТ-СИСТЕМЫ ДЛЯ РАЦИОНАЛЬНОЙ ФАРМАКОТЕРАПИИ

Cocktail test systems are being used for determination of individual activity of cytochrome P450 isoenzymes in a patient. The usage of test systems allows taking into account the phenotypic features of drug metabolism to prescribe the most effective drug therapy. Aim. Development of a test system in the form of a solid gelatin capsule containing microtablets of caffeine, efavirenz, flurbiprofen, metoprolol tartrate and evaluation of their technological properties. Development of a method for the determination of marker components of a "cocktail" formulation using the Proton NMR spectroscopy method. Materials and methods. Microtabletes containing caffeine, efavirenz, flurbiprofen (Molekula Limited, UK) and metoprolol tartrate (KONO Chem Co., LTD, China) were manufactured on a tablet press of the XSpress type (BOSCH, Germany), in one of the sockets of which a 12-punch press tool with a diameter of 2 mm matrix holes was inserted. The resulting microtablets passed pharmaceutical and technological tests for dosage forms in accordance with the Pharmacopoeia of the Russian Federation XIV edition. Finnipipette F1 mechanical pipettes were used to sample the solvent and analytic solution, and a Vortex 2 shaker, an ultrasonic water bath, a Minispin Plus centrifuge, benzoic acid, and DMSO-d6 were used for sample preparation. Results and discussion. A common solvent has been selected for all active substances of the test system. Conditions for the introduction of probe preparations into the filler substance of the sorbent have been selected. Tableting parameters have been worked out. Conclusion. As a result of the conducted research, the composition and production technology of a cocktail test system for rational pharmacotherapy have been developed. A technological scheme for obtaining microtablets of probe preparations has been developed. For a mixture of active substances, the conditions for quantitative analysis were selected using the Proton NMR spectroscopy method.

Peculiarities of carbamazepine pharmacogenetics and the most common adverse drug reactions during its use (literature review)

Carbamazepine is widely used for the treatment of epilepsy, trigeminal neuralgia and psychiatric disorders. However, therapy is often ineffective, and some patients experience adverse drug reactions, which negatively affect both the efficacy and safety of therapy.
 This review systematized and presented up-to-date information regarding the pharmacokinetics and pharmacogenetics of carbamazepine, as well as the most common adverse drug reactions arising from its use. Knowledge of the pharmacokinetics, pharmacodynamics and pharmacogenetics of carbamazepine is necessary to assess the effect of genetically determined activity of cytochrome P450 isoenzymes on the efficacy and safety of this drug. This will make it possible to develop approaches to personalizing the selection of an effective and safe dose of therapy in patients based on individual clinical and biological parameters.

Effects of multi-walled carbon nanotubes in soil on earthworm growth and reproduction, enzymatic activities, and metabolomics

Increased production and environmental release of multi-walled carbon nanotubes (MWCNTs) increase soil exposure and potential risk to earthworms. However, MWCNT toxicity to earthworms remains unclear, with some studies identifying negative effects and others negligible effects. In this study, to determine whether exposure to MWCNTs negatively affects earthworms and to elucidate possible mechanisms of toxicity, earthworms were exposed to sublethal soil concentrations of MWCNTs (10, 50, and 100 mg/kg) for 28 days. Earthworm growth and reproduction, activities of cytochrome P450 (CYP) isoforms (1A2, 2C9, and 3A4) and antioxidant enzymes (superoxide dismutase (SOD), catalase (CAT), and glutathione-s-transferase (GST)), and metabolomics were determined. Effects of MWCNTs on earthworms depended on exposure concentration. Exposure to MWCNTs did not significantly affect growth and reproduction of individual earthworms. Exposure to 50 mg/kg MWCNTs significantly increased activities of CYP2C9, CYP3A4, SOD, CAT, and GST but clearly reduced levels of L-aspartate, L-asparagine, and glutamine. With exposure to 100 mg/kg MWCNTs, toxic effects on earthworms were observed, with significant inhibition in activities of CYP isoenzymes and SOD, significant reductions in L-aspartate, L-asparagine, glutamine, and tryptophan, and simultaneous accumulations of citrate, isocitrate, fumarate, 2-oxoglutarate, pyruvate, D-galactose, carbamoyl phosphate, formyl anthranilate, hypoxanthine, and xanthine. Results suggest that toxicity of MWCNTs to earthworms is associated with reduced detoxification capacity, excessive oxidative stress, and disturbance of multiple metabolic pathways, including amino acids metabolism, the tricarboxylic acid cycle, pyruvate metabolism, D-galactose metabolism, and purine metabolism. The study provides new insights to better understand and predict the toxicity of MWCNTs in soil.

Heart failure as a risk factor of adverse drug reactions. Part 1: potential changes in pharmacokinetics

The use of many drugs is associated with the risk of adverse drug reactions (ADRs), including those that increase mortality and / or morbidity and / or seek medical help or hospitalization, so called «drug-induced diseases» (DID). There is a number of factors that increase the risk of DID (risk factors) including comorbid diseases (for example, chronic kidney disease, hepatic impairment, obesity etc.). These pathologic conditions induce changes in pharmacokinetics (PK) and pharmacodynamics of drugs, thereby increasing the risk of ADRs. One of these diseases is heart failure (HF). Most studies of PK changes were conducted among patients with LVEF from 40 to 45 %, and excluded patients with concomitant diseases that could affect the PK of drugs (for example, serious liver and / or kidney diseases), therefore in polymorbid patients, trial findings may not be applicable. HF may be associated with a decrease in bioavailability, a decrease in volume of distribution, a change in the activity of cytochrome P450 isoenzymes, etc. Individual dose and dosage regimen adjustment can significantly reduce risks, improve the quality of medical care and improve the prognosis in patients with heart failure.

A narrative review of the importance of pharmacokinetics and drug-drug interactions of preventive therapies in migraine management.

ObjectiveTo review the pharmacokinetics of major classes of migraine preventives and the clinical implications of drug–drug interactions (DDIs) with the use of these therapies in migraine management.BackgroundPreventive treatments for migraine are recommended for a large proportion of patients with frequent migraine attacks. These patients often exhibit a number of comorbidities, which may lead to the introduction of multiple concomitant therapies. Potential DDIs must be considered when using polytherapy to avoid increased risk of adverse events (AEs) or inadequate treatment of comorbid conditions.MethodsA literature search was performed to identify pharmacokinetic properties and potential DDIs of beta‐blockers, antiepileptic drugs, antidepressants, calcium channel blockers, gepants, and monoclonal antibody therapies targeting the calcitonin gene‐related peptide pathway with medications that may be used for comorbid conditions.ResultsMost DDIs occur through alterations in cytochrome P450 isoenzyme activity and may be complicated by genetic polymorphism for metabolic enzymes. Additionally, drug metabolism may be altered by grapefruit juice ingestion and smoking. The use of migraine preventive therapies may exacerbate symptoms of comorbid conditions or increase the risk of AEs associated with comorbid conditions as a result of DDIs.ConclusionsDDIs are important to consider in patients with migraine who use multiple medications. The development of migraine‐specific evidence‐based preventive treatments allows for tailored clinical management that reduces the risk of DDIs and associated AEs in patients with comorbidities.

Commiphora myrrha (Nees) Engl. resin extracts induce phase-I cytochrome P450 2C8, 2C9, 2C19, and 3A4 isoenzyme expressions in human hepatocellular carcinoma (HepG2) cells

Commiphora myrrha (Nees) Engl. (C. myrrha) resin is the most Middle Eastern herbal medicine used against numerous diseases. After being decocted or macerated, this resin is widely consumed among Saudi Arabian patients who are already under prescribed medication. Despite its popularity, no studies have been reported on potential modulation effects of these resin extracts on drug metabolism. Therefore, we studied C. myrrha resin extracts on the expression of cytochrome P450 (CYP) drug-metabolizing isoenzyme in human hepatocellular carcinoma cell line HepG2. The C. myrrha extracts were prepared by sonication and boiling, resembling the most popular traditional preparations of maceration and decoction, respectively. Both boiled and sonicated aqueous extracts were fingerprinted using high-performance liquid chromatography equipped with ultra-violet detector (HPLC-UVD). The viability of HepG2 cells treated with these aqueous extracts was determined using CellTiter-Glo® assay in order to select the efficient and non-toxic resin extract concentrations for phase-I metabolic CYP isoenzyme expression analysis. The isoenzyme gene and protein expression levels of CYP 2C8, 2C9, 2C19, and 3A4 were assessed using reverse transcription-quantitative polymerase chain reaction and Western blot technologies. The HPLC-UVD fingerprinting revealed different chromatograms for C. myrrha boiled and sonicated aqueous extracts. Both aqueous extracts were toxic to HepG2 cells when tested at concentrations exceeding 150 µg/ml of the dry crude extract. The CYP 2C8, 2C9, and 2C19 mRNA expression levels increased up to 4.0-fold in HepG2 cells treated with either boiled or sonicated C. myrrha aqueous extracts tested between 1 and 30 µg/ml, as compared with the untreated cells. However, CYP3A4 mRNA expression level exceeded the 2.0-fold cutoff when the cells were exposed to 30 µg/ml of C. myrrha extracts. The up-regulation of CYP mRNA expression levels induced by both boiled and sonicated C. myrrha aqueous extracts was confirmed at the CYP protein expression levels. In conclusion, both sonicated and boiled C. myrrha aqueous extracts modulate CYP 2C8, 2C9, 2C19, and 3A4 gene expression at clinically-relevant concentrations regardless of preparation methods. Further in vitro and in vivo experiments are required for CYP isoenzyme activity assessment and the establishment of herb-drug interaction profile for these traditional medicinal resin extracts.

Risk management strategy for preventing the reduced treatment effectiveness from the position of drug interactions and polypharmacy in patients with coronary heart disease

Introduction: In modern clinical practice, various drug combinations are widely used, especially in cardiological pa­tients. The existing clinical recommendations necessitate using organ protective agents, especially with a patient having a comorbid pathology and with an ineffective monotherapy. In some cases, drug interaction decreases the effectiveness of pharmacotherapy and increases the risk of developing adverse events (AE).
 The purpose of the study was to analyze the modern pharmacotherapy of patients with coronary heart disease (CHD), identify polypharmacy of treatment, evaluate its significance for the treatment process, and determine ways to solve the problem of using a multi-component system of pharmacotherapy risk management.
 Materials and methods: The study involved 156 patients with CHD, among whom 39 received more than 8 drugs at a time.
 Results and discussion: In these patients, the evaluation of drug interactions revealed 580 variants (48 were danger­ous, 428 – significant, 104 – insignificant). The administration of a therapy to comorbid patients, taking into account possible changes in the activity of cytochrome P450 isoenzymes, is one of the promising ways to improve the safety of a combined pharmacotherapy.
 Conclusion: It was revealed that with a mutated cytochrome P450 most of processes of drug biotransformation occurs. And there is a greater risk of developing AE against the background of polypragmasia in polymorbid patients, which makes it possible to individually adjust the dose of beta-blockers, thus affecting the frequency of their development. The choice of management measures should be determined considering all the areas of personalized medicine, includ­ing pharmacogenetic predictors, pharmacoepidemiological data, pharmacoeconomic effectiveness, the development of adverse reactions, polypragmasia, and medical and social risk factors.

Evaluation of the individual detoxification potential in the firefighters and rescuers of emercom of Russia

Relevance. Professional activities of firefighters and rescuers are usually associated with aggressive chemical exposure with increased physical exertion and psychological stress. Therefore, for professional selection and monitoring the health status of firefighters and rescuers of EMERCOM of Russia it is necessary to evaluate the activity of cytochrome P450 isoenzymes that are directly involved in detoxification pathways in the liver.Intention. To evaluate the activity of the CYP3A4 enzyme by phenotyping methods and to establish the frequency of allelic variants of the gene of this enzyme (rs2740574 and rs4987161 polymorphisms) in firefighters and rescuers of EMERCOM of Russia for targeted treatment, rehabilitation and prevention.Methodology. Polymorphisms (rs2740574 and rs4987161) of the CYP3A4 gene and also CYP3A4 activity by the ratio of 6-β-hydroxycortisol / cortisol in the urine were determined during routine periodical medical examination of 64 rescuers and firefighters of rescue units of EMERCOM of Russia. The average age of examined persons was (29.8 ± 5.5) years; 30 of them are rescuers of the North-West Regional Search and Rescue Squad and 34 are firefighters of the territorial fire departments of St. Petersburg.Results and Discussion. According to the results of genotyping of rs2740574 polymorphism of the CYP3A4 gene, EMERCOM employees were divided into 2 groups depending on the presence or absence of a minor allele: “poor” and “rapid” metabolizers (9.4% and 80.6%, respectively). According to the results of genotyping of the rs4987161 polymorphism of the CYP3A4 gene, the examined were classified as “rapid” metabolizers, because patients with a minor allele were not identified. Depending on the ratio of 6-β-hydroxycortisol / cortisol in the urine, the activity level of CYP3A4 was “normal” in 67 %, “poor” in 13 % and “rapid” in 20 % of cases. The ratio of 6-β-hydroxycortisol / cortisol in the urine tended to increase with an increase in work experience and age. No relationship between the studied polymorphic variants of the CYP3A4 gene and the established enzyme activity was observed.Conclusion. Genotyping methods made it possible to identify allelic variants of the CYP3A4 gene that could affect the functionality of the enzyme; however, no association of the studied polymorphisms with enzyme activity was found. In such cases, in the absence of informative genetic markers, it is recommended to evaluate the enzyme activity by phenotyping methods.

The responses of the growth, cytochrome P450 isoenzymes activities and the metabolomics in earthworms to sublethal doses of dichlorvos in soil

In this paper, earthworms (Eisenia fetida) were exposed to sublethal doses of dichlorvos (spiked concentration of 0.1, 1.0, 10 mg/kg) in soil for 14 days, the metabolomics and activities of cytochrome P450 (CYP) isoenzymes (CYP1A2, CYP2C9 and CYP3A4) of earthworms were analyzed aiming to identify sensitive biomarkers and reveal possible mode of toxic action. The results showed that CYP1A2 and CYP2C9 activity appeared to be more sensitive than CYP3A4 activity in response to dichlorvos, and that metabolic responses based on the metabolomics depended on both of the length of exposure and exposure dose. Malate, ornithine, glucose, inosine, myo-inositol and some amino acids (glutamine, tryptophan, phenylalanine, tyrosine, leucine, histidine, glutamate, lysine) and CYP isozenzymes may be biomarkers to reveal the toxic effect of dichlorvos on earthworms. Compared to controls, when dichlorvos dose reached 1.0 and 10 mg/kg on day 14, glucose and ornithine increased significantly, malate and some amino acids (glutamine, tryptophan, phenylalanine, tyrosine, leucine) decreased significantly, and activities of CYP1A2 and CYP2C9 were inhibited significantly. The current results suggested that 1.0 and 10 mg/kg dichlorvos for 14 days of exposure blocked energy metabolism, disordered Krebs cycle, interfered amino acids metabolism and evoked toxic effects on earthworms.

Metabolism of a New Antiaggregant, Indolinone Derivative.

Cytochrome p450-mediated metabolism of GRS (indolinone antiaggregant) and its effects on activities of cytochrome p450 isoenzymes were studied. Inhibition of 6 isomers of cytochrome p450 in human liver microsomes was studied with the use of specific substrates. It was found that human liver cytochrome p450 enzymes could not induce degradation of GRS and that GRS was not an inductor or inhibitor of cytochrome p450 family members 1A2, 2C9, 2C19, 2D6, 2C8, and 3A4. Hence, clinical use of the prospective antiaggregant would not involve the risk of uncontrolled fluctuations in GRS concentrations in the organism because of interactions between the drugs.

Methods for Determination of Functional Activity of Cytochrome P450 Isoenzymes

The review deals with modern methods and technologies for determination of functional activity cytochrome P450 isoenzymes; these include absorbance and fluorescent spectroscopy, electron paramagnetic resonance (EPR), nuclear magnetic resonance (NMR), Raman, Mossbauer, and X-ray spectroscopy, surface plasmon resonance (SPR), atomic force microscopy (AFM). Methods of molecular genetic analysis have been considered in the context of personalized medicine. Using the methods of chromatography mass spectrometry it is possible to analyze reaction products formed in reactions catalyzed by cytochromes P450. Special attention is paid to modern electrochemical systems based on cytochrome P450 isoenzymes, their applicability for analysis of their catalytic activity, their use in practice and further development perspectives for experimental pharmacology, biotechnology and translational medicine.

The impact of porous silicon nanoparticles on human cytochrome P450 metabolism in human liver microsomes in vitro

Engineered nanoparticles are increasingly used as drug carriers in pharmaceutical formulations. This study focuses on the hitherto unaddressed impact of porous silicon (PSi) nanoparticles on human cytochrome P450 (CYP) metabolism, which is the major detoxification route of most pharmaceuticals and other xenobiotics. Three different surface chemistries, including thermally carbonized PSi (TCPSi), aminopropylsilane-modified TCPSi (APTES-TCPSi) and alkyne-terminated thermally hydrocarbonized PSi (Alkyne-THCPSi), were compared for their effects on the enzyme kinetics of the major CYP isoforms (CYP1A2, CYP2A6, CYP2D6, and CYP3A4) in human liver microsomes (HLM) in vitro. The enzyme kinetic parameters, Km and Vmax, and the intrinsic clearance (CLint) were determined using FDA-recommended, isoenzyme-specific model reactions with and without PSi nanoparticles. Data revealed statistically significant alterations of most isoenzyme activities in HLM in the presence of nanoparticles at 1mg/ml concentration, and polymorphic CYP2D6 was the most vulnerable to enzyme inhibition. However, the observed CYP2D6 inhibition was shown to be dose-dependent in case of TCPSi and Alkyne-THCPSi nanoparticles and attenuated at the concentrations below 1μg/ml. Adsorption of the probe substrates onto the hydrophobic Alkyne-THCPSi particles was also observed and taken into account in the determination of the kinetic parameters. Three polymer additives commonly used in pharmaceutical nanoformulations (Pluronics F68 and F127, and polyvinylalcohol) were also separately screened for their effects on CYP isoenzyme activities. These polymers had less effect on the enzyme kinetic parameters, and resulted in increased activity rather than enzyme inhibition, in contrast to the PSi nanoparticles. Although the chosen subcellular model (HLM) is not able to predict the cellular disposition of PSi nanoparticles in hepatocytes and thus provides limited information of probability of CYP interactions in vivo, the present study suggests that mechanistic interactions by the PSi nanoparticles or the polymer stabilizers may appear if these are effectively uptaken by the hepatocytes.

Highly sensitive LC-MS/MS methods for the determination of seven human CYP450 activities using small oral doses of probe-drugs in human.

Cocktails composed of several Cytochrome P450 (CYP450)-selective probe drugs have been shown of value to characterize in vivo drug-metabolism activities. Our objective was to develop and validate highly sensitive and selective LC-MS/MS assays allowing the determination of seven major human CYP450 isoenzyme activities following administration of low oral doses of a modified CYP450 probe-drug cocktail in patients. The seven-drug cocktail was composed of caffeine, bupropion, tolbutamide, omeprazole, dextromethorphan, midazolam (all administered concomitantly) and chlorzoxazone (administered separately) to phenotype for CYP1A2, 2B6, 2C9, 2C19, 2D6, 3A4/5 and 2E1, respectively. Serial plasma and urine samples were collected over an 8h period. The probe-drugs and their respective metabolites were measured in both human plasma and urine, except for omeprazole (plasma only) and chlorzoxazone (urine only). Samples were analyzed by high performance liquid chromatography with heated electrospray ionization tandem mass spectrometry (HPLC-HESI-MS/MS) using a Phenomenex Luna PFP (2) analytical column (3μm PFP(2) 150×3mm) for chromatographic separation. Optimal detection was achieved based on 3 different analytical methods; (1) isocratic elution with a mobile phase consisting of acetonitrile and water both fortified with 0.01% formic acid for the analysis of bupropion, tolbutamide, chlorzoxazone and their respective metabolites; (2) isocratic elution with a mobile phase composed of acetonitrile and ammonium formate (pH 3; 10mM) for omeprazole, dextromethorphan, midazolam and their metabolites; (3) for caffeine and paraxanthine, gradient elution using acetonitrile and 0.01% formic acid in water was used. All calibration functions were linear for all probe drugs and metabolites in both matrices over wide analytical ranges. The main advantages of our methods are the use of specific probe drugs available in most countries, the administration of small doses of probe drugs, small volume of plasma required for the analyses and simple and rapid extraction procedures. The methods met all requirements of specificity, sensitivity, linearity, precision and accuracy and stability generally accepted in bioanalytical chemistry. Determination of CYP450 phenotype in patients will permit characterization of their capacities to metabolize drugs through CYP450 under specific conditions at a definite time. This tool will be highly clinically relevant since wide intersubject variability observed in drug response is largely explained by variation in drug metabolism; it will be particularly useful in polymedicated patients with multiple comorbidities. So far, our CYP450 cocktail assays have been successfully applied to phenotype CYP450 activities in patients.

Imidazoacridinone antitumor agent C-1311 as a selective mechanism-based inactivator of human cytochrome P450 1A2 and 3A4 isoenzymes.

5-Diethylaminoethylamino-8-hydroxyimidazoacridinone (C-1311), a promising antitumor agent that is also active against autoimmune diseases, was determined to be a selective inhibitor of the cytochrome P450 (CYP) 1A2 and 3A4 isoenzymes. Therefore, C-1311 might modulate the effectiveness of other drugs used in multidrug therapy. The present work aimed to identify the mechanism of the observed C-1311-mediated inactivation of CYP1A2 and CYP3A4. The inactivation experiments were performed in vitro using the human recombinant CYP1A2 and CYP3A4 (Bactosomes). CYP isoenzyme activities were determined using the CYP-specific reactions, 7-ethoxycoumarin O-deethylation (CYP1A2) and testosterone 6β-hydroxylation (CYP3A4). The concentrations of CYP-specific substrates and their metabolites formed by CYP isoenzymes were measured by RP-HPLC with UV-Vis detection. The inhibition of CYPs by C-1311 was time-, concentration- and NADPH-dependent, which suggested a mechanism-based mode of action. Using a 10-fold dilution method and potassium ferricyanide we demonstrated the irreversible nature of the inhibition. In addition, the inhibition was attenuated by the presence of alternate substrates (alternative active site ligands) but not by a nucleophilic trapping agent (glutathione) or a reactive oxygen scavenger (catalase), which further supported a mechanism-based action. Substrate depletion partition ratios of 299 and 985 were calculated for the inactivation of CYP1A2 and CYP3A4, respectively. Our results indicated that C-1311 is a potent mechanism-based inactivator of CYP1A2 and CYP3A4. This finding provided new insights into the mechanism of C-1311 antitumor action, particularly in relation to potential pharmacokinetic drug-drug interactions between C-1311 and/or its derivatives and the substrates of CYP isoforms.

Modern aspects of the safe use of extension QT interval medicines

Prolongation of QTinterval is apredictor offatal cardiac arrhythmias and sudden death. In clinical practice, medicines with possible and conditional risk of QTprolongation are combine. The danger of such interactions could be enhanced if medi- cines interact themselves at metabolic rate. The interaction of drugs with possible and conditional risk of QTprolongation and interaction of these drugs with drugs that can influence the metabolic activity of cytochrome P450 isoenzymes require specific attention from physicians. Predictability of QTprolongation by drug-drug interactions at metabolic rate in drug administration will increase the safety of pharmacotherapy of drugs with possible and conditional risk of QTprolongation.

Inhibition of human cytochrome P450 isoenzymes by a phenothiazine neuroleptic levomepromazine: An in vitro study.

Inhibition of cytochrome P450 (CYP) isoenzymes is the most common cause of harmful drug-drug interactions. The present study was aimed at examining the inhibitory effect of the phenothiazine neuroleptic levomepromazine on main CYP isoenzymes in human liver. The experiment was performed in vitro using the human cDNA-expressed CYP1A2, CYP2C9, CYP2C19, CYP2D6 and CYP3A4 (Supersomes). CYP isoenzyme activities were determined using the CYP-specific reactions: caffeine 3-N-demethylation (CYP1A2), diclofenac 4'-hydroxylation (CYP2C9), perazine N-demethylation (CYP2C19), bufuralol 1'-hydroxylation (CYP2D6) and testosterone 6β-hydroxylation (CYP3A4). The rates of the CYP-specific reactions were assessed in the absence and presence of levomepromazine (1-50 μM). The concentrations of CYP-specific substrates and their metabolites formed by CYP isoenzymes were measured by HPLC with UV or fluorimetric detection. Levomepromazine potently inhibited CYP2D6 (K(i) = 6 μM) in a competitive manner. Moreover, the neuroleptic moderately diminished the activity of CYP1A2 (K(i) = 47 μM) and CYP3A4 (K(i) = 34 μM) via a mixed mechanism. On the other hand, levomepromazine did not affect the activities of CYP2C9 and CYP2C19. The inhibition of CYP1A2, CYP2D6 and CYP3A4 by levomepromazine, demonstrated in vitro in the present study, should also be observed in vivo (especially the CYP2D6 inhibition by levomepromazine), since the calculated K(i) values are below or close to the presumed concentration range for levomepromazine in the liver in vivo. Therefore pharmacokinetic interactions involving levomepromazine and CYP2D6, CYP1A2 or CYP3A4 substrates are likely to occur in patients during co-administration of the above-mentioned substrates/drugs.

Pathophysiology and principles of management of the many faces of the acute vaso-occlusive crisis in patients with sickle cell disease.

Effective management of sickle cell pain entails a thorough understanding of its pathophysiology and the pharmacogenomics of the opioids used to manage it. In recent years, there has been significant progress along these two lines. At the pathophysiologic level, there is evidence that the severity and frequency of painful stimuli modulate their transmission at the level of the dorsal horn of the spinal cord. This modulation is achieved via two channels: the α-Amino-3-hydroxy-5-methyl-4-isoxazolepropionic acid (AMPA) and NMDA receptors. Initially, the AMPA channel controls the transmission of stimuli of mild-moderate severity. Once the AMPA channel reaches its limit of membrane depolarization, the NMDA channel is activated and facilitates the transmission of painful stimuli in a progressive fashion leading to central sensitization and glial activation. At the level of pharmacogenomics, the metabolism of each opioid is patient-specific. Glucuronidation is unique for the metabolism of morphine, hydromorphone, and oxymorphone. The metabolism of all other opioids requires specific Cytochrome P450 (CYP) isoenzymes. The activity of each isoenzyme and the activity of the metabolites of each opioid vary among patients depending on their genetic makeup and coexistent environmental factors such as the use of other medications that may enhance or inhibit the CYP isoenzyme activity.

Inhibiton of cytochrome P450 isoenzymes and P-gp activity by multiple extracts of Huang-Lian-Jie-Du decoction

Ethnopharmacological relevanceHuang-Lian-Jie-Du-Decotion (HLJDD), an important traditional Chinese medicine formula, has been used for various diseases in clinical practice, and thus has high potential to induce cytochrome P450 (CYP) isoenzymes/P-glycoprotein (P-gp) mediated herb–drug interactions (HDIs) with other co-administered drugs. The purpose of this study was to investigate the in vitro effects of multiple extracts including aqueous extracts, total flavonoids, iridoids, alkaloids from HLJDD on the activities of CYPs in rats (CYP1A2, CYP2C6, CYP2D2, CYP2E1 and CYP3A1) and P-gp, and then to predict potential interactions with co-administered drugs. Materials and methodsThe effects of the four extracts from HLJDD on the CYPs activity were evaluated in rat liver microsomes incubation system, and then determined by LC-MS/MS-based CYPs probe substrate assay. Caco-2 cell monolayer was used to investigate the effect of the four extracts on the efflux of Rhodamine 123 to evaluate their influences on P-gp activity. ResultsThe results show that total flavonoids and alkaloids exibited strong inhibition on rat CYP isoenzymes activities. Total flavonoids exhibited different inhibitory effects on CYPs activities with an order of CYP3A1>CYP2C6>CYP2E1>CYP1A2>CYP2D2, and the values of IC50 were 4.24, 8.16, 17.56, 19.03, 29.51μg/mL, respectively. Total alkaloids possessed similar inhibition on CYPs and could strongly inhibit the activity of CYP2D2 (IC50=2.38μg/mL), CYP3A1 (IC50=2.61μg/mL), CYP2E1 (IC50=22.35μg/mL), CYP1A2 (IC50=23.2μg/mL) and CYP2C6 (IC50=43.09μg/mL). Moderate degree of inhibition on CYPs activities was observed in aqueous and total iridoids extracts. Results from transport assay revealed that total flavonoids and alkaloids exhibited significant inhibitory effect on P-gp activity as evidenced by strong inhibition on the efflux of Rhodamine-123 with IC50 of 104.6 and 82.6μg/mL. But aqueous extract showed weak and iridoids had negligible effect on P-gp activity. ConclusionsThis study clearly demonstrated that total flavonoids and alkaloids from HLJDD can significantly inhibit the activities of CYPs and P-gp, which should be taken into consideration to predict any potential HDIs when HLJDD and its bioactive components are co-administered with other therapeutic drugs metabolized by CYPs or transported by P-gp.

Implications of cytochrome 450 isoenzymes, aryl-esterase and oxonase activity in the inhibition of the acetylcholinesterase of Chirostoma jordani treated with phosphorothionate pesticides

Organophosphate pesticides must be metabolized by cytochrome-P450 isoenzymes such CYP 2C19 as CYP 3A4 to induce neurotoxicity, but damage apparently depends on the activity of aryl esterases of the oxonase type that are involved in detoxication of these compounds. However, information on this subject is not available in fish. Chirostoma jordani has sustained significant population reductions, probably due to changes in land-use as well as pesticide impact; nevertheless, no specific studies demonstrating this are available. This study shows for the first time that the activity of cytochrome-P450 isoenzymes (CYP 2B6, CYP 2C19, CYP 3A4) in C. jordani is involved in diazinon and chlorpyrifos bioactivation. However, higher toxicity of chlorpyrifos cannot be explained solely because its bioactivation. Differences in toxicity between both pesticides are due to the activity of aryl esterases and oxonases that are responsible for oxon detoxication. Both hepatic enzymes metabolize diazoxon more efficiently than chlorpyrifos oxon. At lethal concentrations, detoxication is particularly important since mortality was lower with diazinon (LC50=1.5μg/L) than with chlorpyrifos (LC50=0.17μg/L). At sublethal levels, maximum acetylcholinesterase inhibition took place at 4h in both brain and muscle and was of lower magnitude in diazinon-treated fish. This is due to the higher affinity of both aryl esterases for diazoxon, which allows higher detoxication rates and therefore greater recovery of acetylcholinesterase activity.