P450 Isoforms Research Articles

Urinary bladder carcinogenic potential of 4,4'-methylenebis(2-chloroaniline) in humanized-liver mice.

Occupational exposure to 4,4'-methylenebis(2-chloroaniline) (MOCA) has been linked to an increased risk of bladder cancer among employees in Japanese plants, indicating its significance as a risk factor for urinary bladder cancer. To investigate the role of MOCA metabolism in bladder carcinogenesis, we administered MOCA to non-humanized (F1-TKm30 mice) and humanized-liver mice for 4 and 28 weeks. We compared MOCA-induced changes in metabolic enzyme expression, metabolite formation, and effects on the urinary bladder epithelium in the two models. At week 4, MOCA exposure induced simple hyperplasia, cell proliferation, and DNA damage in the urothelium of the humanized-liver mice, while in the non-humanized mice these effects were not observed. Notably, the concentration of 4-amino-4'-hydroxylamino-3,3'-dichlorodiphenylmethane (N-OH-MOCA) in the urine of humanized-liver mice was more than 10 times higher than that in non-humanized mice at the 4-week mark. Additionally, we observed distinct differences in the expression of cytochrome P450 isoforms between the two models. Although no bladder tumors were detected after 28 weeks of treatment in either group, these findings suggest that N-OH-MOCA significantly contributes to the carcinogenic potential of MOCA in humans.

O74 EPOXYEICOSATRIENOIC ACIDS ANALOGUE (EET-A) FOR THE TREATMENT OF CHEMOTHERAPY-INDUCED HEART FAILURE WITH NEPHROTIC SYNDROME

Background and Objective: Doxorubicin (DOXO) is still commonly used anthracycline for the treatment of many types of cancers, but its application is burdened with a high risk of cardiotoxicity. The exact mechanisms of DOXO cardiotoxicity are still under investigation. Currently it is believed that the formation of topoisomerase II beta complex with DOXO, together with increased levels of reactive oxygen species (ROS) leads to cardiotoxicity. However, many additional mechanisms can weaken the heart and contribute to HFrEF development. Few years ago, new hypothesis emerged linking the DOXO-cardiotoxicity with the diminished expression of CYP2J2, an isoform of cytochrome P450, which is responsible for epoxyeicosatrienoic acids (EETs) formation. EETs were shown to exhibit vasodilation, decrease inflammation and to promote sodium excretion. Cardiac CYP2J2 overexpression decreased the cardiac injury in DOXO-treated mice. Hence, we decided to test the effectiveness of the EETs analogue (EET-A) in a model of DOXO-induced HFrEF with nephrotic syndrome. Methods: DOXO was administered to male Ren-2 transgenic rats (TGR) via five intravenous injections (cumulative dose 10 mg/kg of body weight). The long-term survival (20 weeks) of rats treated with EET-A (10 mg/kg of body weight administered in drinking water) was compared to ACE inhibitor (ACEi). Cardiac function was evaluated by echocardiography, urine and blood were collected for analysis. Results: The treatment with EET-A did not improve survival and cardiac function measured by echocardiography, however there was a clear renoprotective effect observed. Albuminuria was significantly lower in the EET-A treated group, when compared to ACEi. Plasma albumin did not change after 4-week treatment with EET-A, whereas in both placebo and ACEi a significant decrease was recorded. Conclusions: Despite the lack of improvement of survival, the beneficial effect of EET-A on the kidneys supports its potential for nephrotic syndrome treatment. Supported by MH CZ - DRO (“Institute for Clinical and Experimental Medicine - IKEM, IN 00023001”)

Effect of Itraconazole, a CYP3A4 Inhibitor, and Rifampin, a CYP3A4 Inducer, on the Pharmacokinetics of Vatiquinone.

Vatiquinone is a small molecule inhibitor of 15-lipoxygenase in development for patients with Friedreich's ataxia. The objective of this analysis was to determine the effect of a cytochrome P450 isoform 3A4 (CYP3A4) inhibitor and inducer on vatiquinone pharmacokinetics (PKs). The coadministration of 400mg of vatiquinone with 200mg of itraconazole (a CYP3A4 inhibitor) resulted in increased maximum observed concentration (Cmax) of vatiquinone and systemic exposure (AUC0-inf) by approximately 3.5- and 2.9-fold, respectively. The coadministration of 400mg of vatiquinone with 600mg of rifampin (a CYP3A4 inducer) resulted in decreased vatiquinone Cmax and AUC0-inf by approximately 0.64- and 0.54-fold, respectively. The terminal half-life of vatiquinone was not affected by itraconazole or rifampin. These clinical study results confirm the in vitro reaction phenotyping data that shows that CYP3A4 plays an important role in vatiquinone metabolism. The result of this analysis together with phase 3 efficacy and safety data, population PK analysis, and the exposure-response relationship will determine if the extent of vatiquinone changes in the presence of CYP3A4 inhibitors and inducers are considered clinically relevant.

Overview of preclinical and clinical studies investigating pharmacokinetics and drug–drug interactions of padsevonil

ABSTRACT Background Padsevonil is an antiseizure medication candidate intended to benefit patients with drug-resistant epilepsy. Our investigations aimed at characterizing pharmacokinetics and drug–drug interaction (DDI) profile of padsevonil. Research design and methods An overview of preclinical and clinical pharmacology studies conducted during padsevonil development is provided. Results In preclinical studies, cytochrome (CYP) 3A4 was identified as the main P450 isoform involved in padsevonil metabolism, with potential minor contribution from CYP2C19. Padsevonil was shown to be a time-dependent CYP2C19-inhibitor, weak CYP3A4-inducer, weak inhibitor of P-gp/OCT1/MATE2-K, and potent OCT2-inhibitor. Initial clinical pharmacology studies in healthy participants showed that padsevonil had (i) good absorption, (ii) clearance mediated mainly by metabolism, and (iii) time-dependent kinetics. A study in genotyped participants confirmed the role of CYP2C19 in clearance and time-dependent kinetics; the major contribution of CYP3A4 was confirmed in DDI studies with CYP3A4-inducers (carbamazepine, oxcarbazepine) and -inhibitor (erythromycin). Padsevonil did not affect pharmacokinetics of valproate/lamotrigine/levetiracetam/oxcarbazepine or oral contraceptives. In a cocktail clinical study, padsevonil showed moderate CYP2C19 inhibition (omeprazole) and weak CYP3A4 induction (oral midazolam). No specific effects on CYP1A2 (caffeine), CYP2C9 (S-warfarin), and CYP2D6 (dextromethorphan) were observed. Conclusions The studies presented helped in understanding padsevonil disposition and risks of DDIs, which would inform dosing and prescribing. Clinical trial registration https://www.clinicaltrials.gov identifiers are NCT04131517, NCT03480243, NCT03695094, NCT04075409.

Identification of the human Cytochrome P450 enzymes (P450s) responsible for metabolizing infigratinib to its pharmacologically active Metabolites, BHS697, and CQM157, and assessment of their in vitro inhibition of P450s and UDP-glucuronosyltransferases (UGTs)

Infigratinib, an oral FGFR inhibitor for advanced cholangiocarcinoma, yielded two active metabolites, BHS697 and CQM157, with similar receptor affinity. Our study characterized P450s that are responsible for the metabolism of infigratinib to its two major active metabolites, BHS697 and CQM157. In vitro inhibition of P450s and UGTs by infigratinib, BHS697 or CQM157 was further investigated. The unbound apparent Km values for metabolism of infigratinib to BHS697 by HLM, human recombinant CYP2C8, CYP2C19, CYP2D6 and CYP3A4 enzymes are 4.47, 0.65, 2.50, 30.6 and 2.08 μM, while Vmax values are 90.0 pmol/min/mg protein, 0.13, 0.027, 0.81, and 0.56 pmol/min/pmol protein, respectively. The unbound apparent Km value for metabolism of infigratinib to CQM157 by HLM is 0.049 μM, while the Vmax value is 0.32 pmol/min/mg protein respectively. In HLM, infigratinib displayed moderate inhibition of CYP3A4 and CYP2C19 and weak or negligible inhibition of other P450 isoforms. BHS697 exhibited weak inhibition of CYP2B6, CYP2C9, CYP2C19 and CYP3A4, and no inhibition of CYP2C8 and CYP2D6. CQM157 moderately inhibited CYP2C9 and CYP3A4, and weakly or negligibly inhibited other P450 isoforms. Regarding UGTs, infigratinib moderately inhibited UGT1A4 and weakly inhibited UGT1A1, respectively. BHS697 weakly inhibited UGT1A1. In contrast, CQM157 moderately inhibited both UGT1A1 and UGT1A4. Our findings provide novel insights into the metabolism of and potential DDIs implicating infigratinib.

Evaluation of the inhibition performance of pyrrole derivatives against CYP450 isoforms

Cytochrome P450 is a heme-containing superfamily accountable for the oxidation of various pharmacologically active drugs. The aim of this study was to conduct in silico and in vitro activity assessments of recently synthesized pyrrole-based compounds against three major CYP450 isoforms – CYP1A2, CYP2D6 and CYP3A4. The in vitro study contained specific cytochrome P450 isoform inhibitors and substrates (for CYP1A2, CYP2D6, and CYP3A4) to determine the inhibition performance of the tile compounds at 1 µM concentration. The in vitro data showed that none of the implemented molecules (MI_a-e and MII_a-e) are capable of inhibiting neither of the CYP isoforms. In addition, the potential interactions of the title compounds and the evaluated CYP isoforms were displayed after molecular docking with Glide (Schrödinger). Induced-fit simulations and binding free energy (MM/GBSA) calculations were applied to elucidate the accessibility in each CYP isoform. None of the pyrrole-based compounds exerted significant inhibition towards the applied isoenzymes. Two of the best scored compounds were visualized in the active site of CYP3A4 (PDB: 2V0M). Overall, good correlation between the in vitro results and the free binding MM/GBSA recalculations was observed.

Drug-Drug Interaction Potential of Remimazolam: CYP 450, Transporters, and Protein Binding.

The ultra-short-acting benzodiazepine, remimazolam, is a new treatment modality for procedural sedation and general anesthesia. Its activity is terminated by carboxylesterase 1 (CES1). The objective of this study was to determine the drug-drug interaction (DDI) potential of remimazolam through mechanisms unrelated to its metabolizing enzyme, CES1. Conventional in vitro co-exposure experiments were conducted to study possible interactions of remimazolam and its primary metabolite, CNS7054, mediated by competitive binding to plasma protein or reactions with cytochrome P450 isoforms or drug transporters. No relevant interactions of remimazolam or its metabolite with cytochrome P450 (CYP) isoforms at clinically relevant concentrations were identified. Likewise, standard experiments revealed no clinically relevant interactions with drug transporters and plasma proteins. The present data and analyses suggest a very low potential of remimazolam for pharmacokinetic DDIs mediated by CYP isoforms, drug transporters, and protein binding.

Differential effects of oxytetracycline on detoxification and antioxidant defense in the hepatopancreas and intestine of Chinese mitten crab under cadmium stress

This study aims to evaluate the effects of oxytetracycline (OTC) on detoxification and oxidative defense in the hepatopancreas and intestine of Chinese mitten crab (Eriocheir sinensis) under cadmium (Cd) stress. The crab was exposed to 0.6 μM Cd, 0.6 μM OTC, and 0.6 μM Cd plus 0.6 μM OTC for 42 days. Our results showed that in the intestine, OTC alone enhanced protein carboxylation (PC) and malondialdehyde (MDA) contents, which was associated with the increased OTC accumulation. Compared to Cd alone, Cd plus OTC increased Cd and OTC contents, and reduced detoxification (i.e., glutathione (GSH) content, gene expressions of cytochrome P450 (CYP) isoforms, 7-ethoxyresorufin O-deethylase (EROD) activity, mRNA levels and activities of glutathione peroxidase (GPx), glutathione reductase (GR) and glutathione-S-transferase (GST)), and antioxidant defense (i.e., gene expressions and activities of catalase (CAT) and superoxide dismutase (SOD)) in the intestine, leading to the increased in PC and MDA contents, suggesting that OTC had a synergistic effect on Cd-induced oxidative damage. In the hepatopancreas, although OTC alone increased OTC accumulation, it did not affect PC and MDA contents. Compared to Cd alone, Cd plus OTC reduced MDA content, which was closely related to the improvement of detoxification (i.e., GSH content, mRNA levels of CYP isoforms, EROD activity, gene expressions and activities of GPx, GR and GST), and antioxidant defense (gene expressions and activities of CAT and SOD, metallothionein content). Aryl hydrocarbon receptor (AhR) and nuclear factor E2-related factor 2 (Nrf2) transcriptional expressions were positively correlated with most detoxification- and antioxidant-related gene expressions, respectively, indicating that AhR and Nrf2 were involved in the regulation of these gene expressions. Our results unambiguously demonstrated that OTC had tissue-specific effects on Cd-induced toxicological effect in E. sinensis, which contributed to accurately evaluating Cd toxicity modulated by TCs in crab.

Characterization of the metabolic contributions of cytochrome P450 isoforms to bicyclol using the relative activity factor method

Bicyclol is a hepatoprotective agent widely used for treating chronic hepatitis and drug-induced liver injuries in clinics. The purpose of the study was to elucidate the contribution of CYP450 enzymes to the metabolism of bicyclol using the relative activity factor approach. After incubation with human liver microsomes and recombinant human liver CYP450 enzymes, the calculated contribution of CYP3A4 and 2C19 to the metabolism of bicyclol was 85.6–90.3% and 9.2–9.7%, respectively. The metabolism was interrupted in the presence of CYP3A4 and 2C19 selective inhibitors. These findings help to predict or avoid metabolic drug–drug interactions or toxicity in clinical applications of bicyclol.

Human Leucocyte Antigen Genetics in Idiosyncratic Drug-Induced Liver Injury with Evidence Based on the Roussel Uclaf Causality Assessment Method.

The human leucocyte antigen (HLA) allele variability was studied in cohorts of patients with idiosyncratic drug-induced liver injury (iDILI). Some reports showed an association between HLA genetics and iDILI, proposing HLA alleles as a potential risk factor for the liver injury. However, the strength of such assumptions heavily depends on the quality of the iDILI diagnosis, calling for a thorough analysis. Using the PubMed database and Google Science, a total of 25 reports of case series or single cases were retrieved using the terms HLA genes and iDILI. It turned out that in 10/25 reports (40%), HLA genetics were determined in iDILI cases, for which no causality assessment method (CAM) was used or a non-validated tool was applied, meaning the findings were based on subjective opinion, providing disputable results and hence not scoring individual key elements. By contrast, in most iDILI reports (60%), the Roussel Uclaf Causality Assessment Method (RUCAM) was applied, which is the diagnostic algorithm preferred worldwide to assess causality in iDILI cases and represents a quantitative, objective tool that has been well validated by both internal and external DILI experts. The RUCAM provided evidence-based results concerning liver injury by 1 drug class (antituberculotics + antiretrovirals) and 19 different drugs, comprising 900 iDILI cases. Among the top-ranking drugs were amoxicillin-clavulanate (290 cases, HLA A*02:01 or HLA A*30:02), followed by flucloxacillin (255 cases, HLA B*57:01), trimethoprim-sulfamethoxazole (86 cases, HLA B*14:01 or HLA B*14:02), methimazole (40 cases, HLA C*03:02), carbamazepine (29 cases, HLA A*31:01), and nitrofurantoin (26 cases, HLA A*33:01). In conclusion, the HLA genetics in 900 idiosyncratic drug-induced liver injury cases with evidence based on the RUCAM are available for studying the mechanistic steps leading to the injury, including metabolic factors through cytochrome P450 isoforms and processes that activate the innate immune system to the adaptive immune system.

Metabolism of Xenobiotic Compounds Increases Free Radical Production and Reduces Endogenous Antioxidant Mechanisms

Xenobiotic compounds are foreign substances that enter the body. These substances include drugs, insecticides, toluene, chloroform, formalin, and others that can enter by inhalation, through skin exposure or swallowed together with food. The use of xenobiotic compounds is increasing every year along with the development of the industrial sector, which impacts greater exposure for workers. Xenobiotic compounds that enter the body will cause harm to the body, and they will be detoxified in the liver. Xenobiotic compounds can bind to cytochrome p450 isoform 2E1, which will cause the formation of free radicals. A body exposed to xenobiotic compounds can respond by reducing free radicals using endogenous antioxidants. They are Superoxide Dismutase (SOD) and Glutathione Peroxidase (GPX). If the amount of free radicals exposure is too large, oxidative stress will occur. It triggers liver cell damage and release of alanine aminotransferase (ALT) and Aspartate Aminotransferase (AST) into the blood vessels, which causes an increase in these enzymes in the blood.

Pharmacokinetics of asundexian with combined CYP3A and P-gp inhibitors and an inducer: Target in vitro and in vivo studies.

Asundexian is an oral, direct and reversible inhibitor of activated factor XI (FXIa) in development for the treatment of thromboembolic events. This article summarizes results from preclinical and clinical studies, including identification of enzymes involved in asundexian pharmacokinetics, and evaluation of potential target drug-drug interactions. In vitro studies investigated the substrate characteristics of asundexian towards several cytochrome P450 (CYP) isoforms, hydrolytic enzymes and drug transporters. Inhibition of the amide hydrolysis of asundexian was investigated in vitro for several relevant drugs. Phase 1 studies in healthy male participants investigated the pharmacokinetics (PK) of asundexian upon co-administration of combined inhibitors or an inducer of P-gp and CYP3A4 (itraconazole, verapamil or carbamazepine, respectively, or the moderate CYP3A4 inhibitor fluconazole). The pharmacodynamic (PD) markers are activated partial thromboplastin time and FXIa inhibition. Asundexian was predominantly metabolized via carboxylesterase 1 and, to a lesser extent, via CYP3A4 and is a substrate of P-gp. The asundexian area under the plasma concentration-time curve (AUC) increased by 103% and 75.6% upon combined inhibition of P-gp and strong or moderate inhibition of CYP3A4, respectively, but was unaffected by moderate CYP3A4 inhibition. Combined P-gp and CYP3A4 induction by carbamazepine decreased asundexian AUC by 44.4%. PD is concentration-dependent, thus no differences in maximum responses and recovery commensurate with PK effect(s) were observed. Adverse events were mild and asundexian was well tolerated. The presented studies confirmed that CYP3A4 and P-gp contribute to asundexian metabolism and excretion. Observed effects were in line with data from a previous mass balance study.

Deciphering the Role of Fatty Acid-Metabolizing CYP4F11 in Lung Cancer and Its Potential As a Drug Target.

Lung cancer is the leading cause of cancer deaths worldwide. We found that the cytochrome P450 isoform CYP4F11 is significantly overexpressed in patients with lung squamous cell carcinoma. CYP4F11 is a fatty acid ω-hydroxylase and catalyzes the production of the lipid mediator 20-hydroxyeicosatetraenoic acid (20-HETE) from arachidonic acid. 20-HETE promotes cell proliferation and migration in cancer. Inhibition of 20-HETE-generating cytochrome P450 enzymes has been implicated as novel cancer therapy for more than a decade. However, the exact role of CYP4F11 and its potential as drug target for lung cancer therapy has not been established yet. Thus, we performed a transient knockdown of CYP4F11 in the lung cancer cell line NCI-H460. Knockdown of CYP4F11 significantly inhibits lung cancer cell proliferation and migration while the 20-HETE production is significantly reduced. For biochemical characterization of CYP4F11-inhibitor interactions, we generated recombinant human CYP4F11. Spectroscopic ligand binding assays were conducted to evaluate CYP4F11 binding to the unselective CYP4A/F inhibitor HET0016. HET0016 shows high affinity to recombinant CYP4F11 and inhibits CYP4F11-mediated 20-HETE production in vitro with a nanomolar IC 50 Cross evaluation of HET0016 in NCI-H460 cells shows that lung cancer cell proliferation is significantly reduced together with 20-HETE production. However, HET0016 also displays antiproliferative effects that are not 20-HETE mediated. Future studies aim to establish the role of CYP4F11 in lung cancer and the underlying mechanism and investigate the potential of CYP4F11 as a therapeutic target for lung cancer. SIGNIFICANCE STATEMENT: Lung cancer is a deadly cancer with limited treatment options. Cytochrome P450 4F11 (CYP4F11) is significantly upregulated in lung squamous cell carcinoma. Knockdown of CYP4F11 in a lung cancer cell line significantly attenuates cell proliferation and migration with reduced production of the lipid mediator 20-hydroxyeicosatetraenoic acid (20-HETE). Studies with the unselective inhibitor HET0016 show a high inhibitory potency of CYP4F11-mediated 20-HETE production using recombinant enzyme. Overall, our studies demonstrate the potential of targeting CYP4F11 for new transformative lung cancer treatment.

Predicting routes of phase I and II metabolism based on quantum mechanics and machine learning

Unexpected metabolism could lead to the failure of many late-stage drug candidates or even the withdrawal of approved drugs. Thus, it is critical to predict and study the dominant routes of metabolism in the early stages of research. We describe the development and validation of a ‘WhichEnzyme’ model that accurately predicts the enzyme families most likely to be responsible for a drug-like molecule’s metabolism. Furthermore, we combine this model with our previously published regioselectivity models for Cytochromes P450, Aldehyde Oxidases, Flavin-containing Monooxygenases, UDP-glucuronosyltransferases and Sulfotransferases – the most important Phase I and Phase II drug metabolising enzymes – and a ‘WhichP450’ model that predicts the Cytochrome P450 isoform(s) responsible for a compound’s metabolism. The regioselectivity models are based on a mechanistic understanding of these enzymes’ actions and use quantum mechanical simulations with machine learning methods to accurately predict sites of metabolism and the resulting metabolites. We train heuristics based on the outputs of the ‘WhichEnzyme’, ‘WhichP450’, and regioselectivity models to determine the most likely routes of metabolism and metabolites to be observed experimentally. Finally, we demonstrate that this combination delivers high sensitivity in identifying experimentally reported metabolites and higher precision than other methods for predicting in vivo metabolite profiles.

The discovery of aryl-2-nitroethyl triamino pyrimidines as anti-Trypanosoma brucei agents

Pteridine reductase 1 (PTR1) is a catalytic protein belonging to the folate metabolic pathway in Trypanosmatidic parasites. PTR1 is a known target for the medicinal chemistry development of antiparasitic agents against Trypanosomiasis and Leishmaniasis. In previous studies, new nitro derivatives were elaborated as PTR1 inhibitors. The compounds showing a diamino-pyrimidine core structure were previously developed but they showed limited efficacy. Therefore, a new class of phenyl-, heteroaryl- and benzyloxy-nitro derivatives based on the 2-nitroethyl-2,4,6-triaminopyrimidine scaffold were designed and tested. The compounds were assayed for their ability to inhibit T. brucei and L. major PTR1 enzymes and for their antiparasitic activity towards T. brucei and L. infantum parasites. To understand the structure-activity relationships of the compounds against TbPTR1, the X-ray crystallographic structure of the 2,4,6-triaminopyrimidine (TAP) was obtained and molecular modelling studies were performed. As a next step, only the most effective compounds against T. brucei were then tested against the amastigote cellular stage of T. cruzi, searching for a broad-spectrum antiprotozoal agent. An early ADME-Tox profile evaluation was performed. The early toxicity profile of this class of compounds was investigated by measuring their inhibition of hERG and five cytochrome P450 isoforms (CYP1A2, CYP2C9, CYP2C19, CYP2D6 and CYP3A4), cytotoxicity towards A549 cells and mitochondrial toxicity. Pharmacokinetic studies (SNAP-PK) were performed on selected compounds using hydroxypropyl-β-cyclodextrins (50 % w/v) to preliminarily study their plasma concentration when administered per os at a dose of 20 mg/kg. Compound 1p, showed the best pharmacodynamic and pharmacokinetic properties, can be considered a good candidate for further bioavailability and efficacy studies.

The roles of CYP2C19 and CYP3A4 in the invitro metabolism of β-eudesmol in human liver: Reaction phenotyping and enzyme kinetics.

β-eudesmol is a major bioactive component of Atractylodes lancea (AL). AL has been developed as the capsule formulation of standardized AL extract for treating cholangiocarcinoma (CCA). However, the complex constituents of herbal products increase the risk of adverse drug interactions. β-eudesmol has demonstrated inhibitory effects on rCYP2C19 and rCYP3A4 in the previous research. This study aimed to identify the cytochrome P450 (CYP) isoforms responsible for the metabolism of β-eudesmol and determine the enzyme kinetic parameters and the metabolic stability of β-eudesmol metabolism in the microsomal system. Reaction phenotyping using human recombinant CYPs (rCYPs) and selective chemical inhibitors of CYP1A2, CYP2C9, CYP2C19, CYP2D6, and CYP3A4 was performed, and enzyme kinetics and metabolic stability were investigated using human liver microsome (HLM). The results suggest that CYP2C19 and CYP3A4 play significant roles in β-eudesmol metabolism. The disappearance half-life (t1/2 ) and intrinsic clearance (CLint ) of β-eudesmol were 17.09 min and 0.20 mL/min·mg protein, respectively. Enzyme kinetic analysis revealed the Michaelis-Menten constant (Km ) and maximum velocity (Vmax ) of 16.76 μM and 3.35 nmol/min·mg protein, respectively. As a component of AL, β-eudesmol, as a substrate and inhibitor of CYP2C19 and CYP3A4, has a high potential for drug-drug interactions when AL is co-administered with other herbs or conventional medicines.

Panax ginseng C.A. Mey. as a potential raw material in the treatment of negative effects of alcohol consumption

An alcohol hangover is a complex set of adverse symptoms that occur after excessive consumption of alcoholic beverages. Hangovers are not solely caused by ethanol, but mainly by its toxic metabolites, such as acetaldehyde and acetic acid. Most of the negative symptoms occur when the concentration of alcohol in the blood decreases after drinking. Given that a large percentage of people consume alcohol and experience negative effects from its consumption, potential supplementation to help eliminate these adverse effects has been proposed. One promising ingredient is ginseng, which has been used in Eastern culture and traditional Chinese medicine for over 2,000 years. The aim of this study was to present the potential of Panax ginseng C.A. Mey. as an aid for the adverse effects of alcohol consumption and its protective effect on liver function. The literature analysis was based on data indexed in medical databases PubMed, Scopus, and Medline since 2010, using keywords such as "Panax ginseng," "hangover," "alcohol consumption," and "metabolism," connected by the logical conjuction “and.” The analysis confirmed the effectiveness of P. ginseng in the treatment and prevention of hangovers. Preparations containing P. ginseng can support ethanol metabolism by inducing enzymes such as alcohol dehydrogenase, aldehyde dehydrogenase, and the cytochrome P450 isoform 2E1 system. An additional advantage of using P. ginseng may be its hepatoprotective effect.

Updates on the Interactions of Herbs Constituents with Cytochrome P450 Drug Metabolizing Enzymes

Abstract: Cytochrome P450 enzymes are the main drug-metabolizing enzyme, and their activity is influenced by numerous factors, including several plants’ constituents. Herbal products mediated induction or inhibition of cytochrome P450 leads to a change in drug metabolism and consequently, drug bioavailability, safety, therapeutic efficacy, and herb-drug interaction. Since herbal preparations are commonly used either alone or concomitantly with conventional drugs, understanding the interactions of plant constituents with cytochrome P450 is crucial. Therefore, this review highlighted plants with significant inhibition, induction, or modulatory /contradictory effects on human and animal cytochrome P450 enzymes. An extensive literature search was conducted on PubMed, ScienceDirect, and Google Scholar databases to identify relevant literature. About sixty medicinal plants with significant interaction with cytochrome P450 isoforms were summarized with their common uses and affected cytochrome P450 enzyme isoforms. Besides, representative examples of plant constituents and other factors contributing to plant-induced cytochrome P450 modulation and possible herb-drug interactions were also discussed in this review.

A Comparative GC/MS Analysis of Citrus Essential Oils: Unveiling the Potential Benefits of Herb-Drug Interactions in Preventing Paracetamol-Induced Hepatotoxicity.

Our study aimed to test the potential of Citrus oils in protecting against paracetamol (PAR)-induced hepatotoxicity. The essential oils of Pineapple sweet orange (OO), Murcott mandarin (MO), Red grapefruit (GO), and Oval kumquat (KO) were investigated using gas chromatography coupled with mass spectrometry (GC/MS). Twenty-seven compounds were identified, with monoterpene hydrocarbons being abundant class. d-Limonene had the highest percentage (92.98 %, 92.82 %, 89.75 %, and 94.46 % in OO, MO, GO, and KO, respectively). Hierarchical cluster analysis (HCA) and principal components analysis (PCA) revealed that octanal, linalool, germacrene D, and d-limonene were the principal discriminatory metabolites that segregated the samples into three distinct clusters. In vitro antioxidant capacities were ranged from 1.2-12.27, 1.79-5.91, and 235.05-585.28 μM Trolox eq/mg oil for 2,2'-azinobis(3-ethylbenzothiazoline-6-sulfonic (ABTS), ferric-reducing antioxidant power (FRAP), and oxygen radical absorbance capacity (ORAC), respectively. In vivo, citrus oils exhibited a significant reduction in alanine transaminase (ALT), aspartate transaminase (AST), alkaline phosphatase (ALP), and nitric oxide (NO). Additionally, there was an increase in glutathione reductase (GSH), and the liver architecture was nearly normal. Molecular docking revealed that d-limonene exhibited a good inhibitory interaction with cytochrome P450 (CYP450) isoforms 1A2, 3A4, and 2E1, with binding energies of -6.17, -4.51, and -5.61 kcal/mol, respectively.

Unraveling a Combined Inactivation Mechanism of Cytochrome P450s by Genipin, the Major Reactive Aglycone Derived from Gardeniae Fructus.

Genipin (GP) is the reactive aglycone of geniposide, the main component of traditional Chinese medicine Gardeniae Fructus (GF). The covalent binding of GP to cellular proteins is suspected to be responsible for GF-induced hepatotoxicity and inhibits drug-metabolizing enzyme activity, although the mechanisms remain to be clarified. In this study, the mechanisms of GP-induced human hepatic P450 inactivation were systemically investigated. Results showed that GP inhibited all tested P450 isoforms via distinct mechanisms. CYP2C19 was directly and irreversibly inactivated without time dependency. CYP1A2, CYP2C9, CYP2D6, and CYP3A4 T (testosterone as substrate) showed time-dependent and mixed-type inactivation, while CYP2B6, CYP2C8, and CYP3A4 M (midazolam as substrate) showed time-dependent and irreversible inactivation. For CYP3A4 inactivation, the kinact/KI values in the presence or absence of NADPH were 0.26 or 0.16 min-1 mM-1 for the M site and 0.62 or 0.27 min-1 mM-1 for the T site. Ketoconazole and glutathione (GSH) both attenuated CYP3A4 inactivation, suggesting an active site occupation- and reactive metabolite-mediated inactivation mechanism. Moreover, the in vitro and in vivo formation of a P450-dependent GP-S-GSH conjugate indicated the involvement of metabolic activation and thiol residues binding in GP-induced enzyme inactivation. Lastly, molecular docking analysis simulated potential binding sites and modes of GP association with CYP2C19 and CYP3A4. We propose that direct covalent binding and metabolic activation mediate GP-induced P450 inactivation and alert readers to potential risk factors for GP-related clinical drug-drug interactions.