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1.
Impact of Acute Inflammation on Cytochromes P450 Activity Assessed by the Geneva Cocktail.
Lenoir, C, Daali, Y, Rollason, V, Curtin, F, Gloor, Y, Bosilkovska, M, Walder, B, Gabay, C, Nissen, MJ, Desmeules, JA, et al
Clinical pharmacology and therapeutics. 2021;(6):1668-1676
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Abstract
Cytochromes P450 (CYP) are subject to important interindividual variability in their activity due to genetic and environmental factors and some diseases. Limited human data support the idea that inflammation downregulates CYP activities. Our study aimed to evaluate the impact of orthopedic surgery (acute inflammation model) on the activity of six human CYP. This prospective observational study was conducted in 30 patients who underwent elective hip surgery at the Geneva University Hospitals in Switzerland. The Geneva phenotyping cocktail containing caffeine, bupropion, flurbiprofen, omeprazole, dextromethorphan, and midazolam as probe drugs respectively assessing CYP1A2, 2B6, 2C9, 2C19, 2D6, and 3A activities was administered orally before surgery, day 1 (D1) and 3 (D3) postsurgery and at discharge. Capillary blood samples were collected 2 hours after cocktail intake to assess metabolic ratios (MRs). Serum inflammatory markers (CRP, IL-6, IL-1β, TNF-α, and IFN-γ) were also measured in blood. CYP1A2 MRs decreased by 53% (P < 0.0001) between baseline and the nadir at D1. CYP2C19 and CYP3A activities (MRs) decreased by 57% (P = 0.0002) and 61% (P < 0.0001), respectively, with the nadir at D3. CYP2B6 and CYP2C9 MRs increased by 120% (P < 0.0001) and 79% (P = 0.018), respectively, and peaked at D1. Surgery did not have a significant impact on CYP2D6 MR. Hip surgery was a good acute inflammation model as CRP, IL-6, and TNF-α peak levels were reached between D1 and day 2 (D2). Acute inflammation modulated CYP activity in an isoform-specific manner, with different magnitudes and kinetics. Acute inflammation may thus have a clinically relevant impact on the pharmacokinetics of these CYP substrates.
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Prediction of the Area under the Curve Using Limited-Point Blood Sampling in a Cocktail Study to Assess Multiple CYP Activities.
Miura, M, Tanaka, S, Uchida, S, Kamiya, C, Katayama, N, Hakamata, A, Odagiri, K, Inui, N, Kawakami, J, Watanabe, H, et al
Biological & pharmaceutical bulletin. 2021;(6):762-770
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Abstract
A cocktail study is an in vivo evaluation method to assess multiple CYP activities via a single trial and single administration of a cocktail drug that is a combination of multiple CYP substrates. However, multiple blood samples are required to evaluate the pharmacokinetics of a CYP probe drug. A limited-point sampling method is generally beneficial in clinical studies because of the simplified protocol and reduced participant burden. The aim of this study was to evaluate whether a limited-point plasma concentration analysis of CYP substrates in a cocktail drug could predict their area under the curve (AUC). We created prediction models of five CYP substrates (caffeine, losartan, omeprazole, dextromethorphan, and midazolam) using multiple linear regressions from the data of two cocktail studies, and then performed predictability analysis of these models using data derived from data in the co-administration with inducer (rifampicin) and inhibitors (fluvoxamine and cimetidine). For the administration of inhibitors, the AUC prediction accuracy (mean absolute error (MAE)) were <39.5% in Model 1 and <26.2% in Model 2 which were created using 1- and 4-point sampling data. MAE shows larger values in the administration of inducer in compared with the administration of inhibitors. The accuracy of the prediction in Model 2 could be acceptable for screening of inhibitions. MAE for caffeine, dextromethorphan, and midazolam were acceptable in the model that used 4 sampling points from all data. The use of this method could reduce the burden on the subject and make it possible to evaluate each AUC in a minimally invasive manner.
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Cytochrome P450-epoxygenated fatty acids inhibit Müller glial inflammation.
Ontko, CD, Capozzi, ME, Kim, MJ, McCollum, GW, Penn, JS
Scientific reports. 2021;(1):9677
Abstract
Free fatty acid dysregulation in diabetics may elicit the release of inflammatory cytokines from Müller cells (MC), promoting the onset and progression of diabetic retinopathy (DR). Palmitic acid (PA) is elevated in the sera of diabetics and stimulates the production of the DR-relevant cytokines by MC, including IL-1β, which induces the production of itself and other inflammatory cytokines in the retina as well. In this study we propose that experimental elevation of cytochrome P450 epoxygenase (CYP)-derived epoxygenated fatty acids, epoxyeicosatrienoic acid (EET) and epoxydocosapentaenoic acid (EDP), will reduce PA- and IL-1β-induced MC inflammation. Broad-spectrum CYP inhibition by SKF-525a increased MC expression of inflammatory cytokines. Exogenous 11,12-EET and 19,20-EDP significantly decreased PA- and IL-1β-induced MC expression of IL-1β and IL-6. Both epoxygenated fatty acids significantly decreased IL-8 expression in IL-1β-induced MC and TNFα in PA-induced MC. Interestingly, 11,12-EET and 19,20-EDP significantly increased TNFα in IL-1β-treated MC. GSK2256294, a soluble epoxide hydrolase (sEH) inhibitor, significantly reduced PA- and IL-1β-stimulated MC cytokine expression. 11,12-EET and 19,20-EDP were also found to decrease PA- and IL-1β-induced NFκB-dependent transcriptional activity. These data suggest that experimental elevation of 11,12-EET and 19,20-EDP decreases MC inflammation in part by blocking NFκB-dependent transcription and may represent a viable therapeutic strategy for inhibition of early retinal inflammation in DR.
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Napabucasin Drug-Drug Interaction Potential, Safety, Tolerability, and Pharmacokinetics Following Oral Dosing in Healthy Adult Volunteers.
Dai, X, Karol, MD, Hitron, M, Hard, ML, Goulet, MT, McLaughlin, CF, Brantley, SJ
Clinical pharmacology in drug development. 2021;(8):824-839
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Abstract
Napabucasin is an orally administered reactive oxygen species generator that is bioactivated by the intracellular antioxidant nicotinamide adenine dinucleotide phosphate:quinone oxidoreductase 1. Napabucasin induces cell death in cancer cells, including cancer stem cells. This phase 1 study (NCT03411122) evaluated napabucasin drug-drug interaction potential for 7 cytochrome P450 (CYP) enzymes and the breast cancer resistance protein transporter/organic anion transporter 3. Healthy volunteers who tolerated napabucasin during period 1 received probe drugs during period 2, and in period 3 received napabucasin (240 mg twice daily; days 1-11) plus a phenotyping cocktail containing omeprazole (CYP2C19), caffeine (CYP1A2), flurbiprofen (CYP2C9), bupropion (CYP2B6), dextromethorphan (CYP2D6), midazolam (CYP3A) (all oral; day 6), intravenous midazolam (day 7), repaglinide (CYP2C8; day 8), and rosuvastatin (breast cancer resistance protein/organic anion transporter 3; day 9). Drug-drug interaction potential was evaluated in 17 of 30 enrolled volunteers. Napabucasin coadministration increased the area under the plasma concentration-time curve from time 0 extrapolated to infinity (geometric mean ratio [90% confidence interval]) of caffeine (124% [109.0%-141.4%]), intravenous midazolam (118% [94.4%-147.3%]), repaglinide (127% [104.7%-153.3%]), and rosuvastatin (213% [42.5%-1068.3%]) and decreased the area under the plasma concentration-time curve from time 0 extrapolated to infinity of dextromethorphan (71% [47.1%-108.3%]), bupropion (79% [64.6%-97.0%]), and hydroxybupropion (45% [15.7%-129.6%]). No serious adverse events/deaths were reported. Generally, napabucasin is not expected to induce/inhibit drug clearance to a clinically meaningful degree.
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Effects of Vitamin D Receptor, Cytochrome P450 3A, and Cytochrome P450 Oxidoreductase Genetic Polymorphisms on the Pharmacokinetics of Remimazolam in Healthy Chinese Volunteers.
Hu, K, Xiang, Q, Wang, Z, Sheng, X, Li, L, Liang, Y, Zhao, X, Ye, X, Cui, Y
Clinical pharmacology in drug development. 2021;(1):22-29
Abstract
Remimazolam is a new ultra-short-acting benzodiazepine used to induce and maintain anesthesia and procedural sedation. Its compound structure is similar to midazolam's. Midazolam metabolism might be affected by vitamin D receptor (VDR), cytochrome P450 3A, and cytochrome P450 oxidoreductase genetic polymorphisms. This study investigated the effects of VDR, cytochrome P450 3A, and cytochrome P450 oxidoreductase genetic polymorphisms on the pharmacokinetics of remimazolam in healthy Chinese volunteers after a single intravenous injection of remimazolam besylate. Blood samples were collected from subjects (n = 62) at scheduled time intervals before and after injection. High-performance liquid chromatography-tandem mass spectrometry was used to quantify plasma remimazolam and RF7054 (its inactive carboxylic acid metabolite) concentrations. The relationship between plasma remimazolam concentration, pharmacokinetic parameters, and polymorphic alleles was assessed for each subject. The rs4516035 allele affected the elimination half-life of RF7054 (P = .043), while the rs1544410 allele affected the dose-normalized maximum observed plasma concentration (Cmax /D) of remimazolam (P = .025) in 46 volunteers. Results showed that VDR genetic polymorphisms might affect the pharmacokinetics of remimazolam in the Chinese population.
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A Drug-Drug Interaction Study Evaluating the Effect of Givosiran, a Small Interfering Ribonucleic Acid, on Cytochrome P450 Activity in the Liver.
Vassiliou, D, Sardh, E, Harper, P, Simon, AR, Clausen, VA, Najafian, N, Robbie, GJ, Agarwal, S
Clinical pharmacology and therapeutics. 2021;(5):1250-1260
Abstract
Givosiran (trade name GIVLAARI) is a small interfering ribonucleic acid that targets hepatic delta-aminolevulinic acid synthase 1 (ALAS1) messenger RNA for degradation through RNA interference (RNAi) that has been approved for the treatment of acute hepatic porphyria (AHP). RNAi therapeutics, such as givosiran, have a low liability for drug-drug interactions (DDIs) because they are not metabolized by cytochrome 450 (CYP) enzymes, and do not directly inhibit or induce CYP enzymes in the liver. The pharmacodynamic effect of givosiran (lowering of hepatic ALAS1, the first and rate limiting enzyme in the heme biosynthesis pathway) presents a unique scenario where givosiran could potentially impact heme-dependent activities in the liver, such as CYP enzyme activity. This study assessed the impact of givosiran on the pharmacokinetics of substrates of 5 major CYP450 enzymes in subjects with acute intermittent porphyria (AIP), the most common type of AHP, by using the validated "Inje cocktail," comprised of caffeine (CYP1A2), losartan (CYP2C9), omeprazole (CYP2C19), dextromethorphan (CYP2D6), and midazolam (CYP3A4). We show that givosiran treatment had a differential inhibitory effect on CYP450 enzymes in the liver, resulting in a moderate reduction in activity of CYP1A2 and CYP2D6, a minor effect on CYP3A4 and CYP2C19, and a similar weak effect on CYP2C9. To date, this is the first study evaluating the DDI for an oligonucleotide therapeutic and highlights an atypical drug interaction due to the pharmacological effect of givosiran. The results of this study suggest that givosiran does not have a large effect on heme-dependent CYP enzyme activity in the liver.
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Plant cytochrome P450s: Role in stress tolerance and potential applications for human welfare.
Singh, A, Panwar, R, Mittal, P, Hassan, MI, Singh, IK
International journal of biological macromolecules. 2021;:874-886
Abstract
Cytochrome P450s (CYPs) are a versatile group of enzymes and one of the largest families of proteins, controlling various physiological processes via biosynthetic and detoxification pathways. CYPs perform multiple roles through a critical irreversible enzymatic reaction in which an oxygen atom is inserted within hydrophobic molecules, converting them into the reactive and hydro soluble components. During evolution, plants have acquired significantly more number of CYPs and represent about 1% of the encoded genes . CYPs are highly conserved proteins involved in growth, development and tolerance against biotic and abiotic stresses. Furthermore, CYPs reinforce plants' molecular and chemical defense mechanisms by regulating the biosynthesis of secondary metabolites, enhancing reactive oxygen species (ROS) scavenging and controlling biosynthesis and homeostasis of phytohormones, including abscisic acid (ABA) and jasmonates. Thus, they are the critical targets of metabolic engineering for enhancing plant defense against environmental stresses. Additionally, CYPs are also used as biocatalysts in the fields of pharmacology and phytoremediation. Herein, we highlight the role of CYPs in plant stress tolerance and their applications for human welfare.
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In Vitro Human Cell-Based Experimental Models for the Evaluation of Enteric Metabolism and Drug Interaction Potential of Drugs and Natural Products.
Li, AP
Drug metabolism and disposition: the biological fate of chemicals. 2020;(10):980-992
Abstract
Elements of key enteric drug metabolism and disposition pathways are reviewed to aid the assessment of the applicability of current cell-based enteric experimental systems for the evaluation of enteric metabolism and drug interaction potential. Enteric nuclear receptors include vitamin D receptor, constitutive androstane receptor, pregnane X receptor, farnesoid X receptor, liver X receptor, aryl hydrocarbon receptor, and peroxisome proliferator-activated receptor. Enteric drug metabolizing enzyme pathways include both cytochrome P450 (P450) and non-P450 drug metabolizing enzymes based on gene expression, proteomics, and activity. Both uptake and efflux transporters are present in the small intestine, with P-glycoprotein found to be responsible for most drug-drug and food-drug interactions. The cell-based in vitro enteric systems reviewed are 1) immortalized cell line model: the human colon adenocarcinoma (Caco-2) cells; 2) human stem cell-derived enterocyte models: stem cell enteric systems, either from intestinal crypt cells or induced pluripotent stem cells; and 3) primary cell models: human intestinal slices, cryopreserved human enterocytes, permeabilized cofactor-supplemented (MetMax) cryopreserved human enterocytes, and cryopreserved human intestinal mucosa. The major deficiency with both immortalized cell lines and stem cell-derived enterocytes is that drug metabolizing enzyme activities, although they are detectable, are substantially lower than those for the intestinal mucosa in vivo. Human intestine slices, cryopreserved human enterocytes, MetMax cryopreserved human enterocytes, and cryopreserved human intestinal mucosa retain robust enteric drug metabolizing enzyme activity and represent appropriate models for the evaluation of metabolism and metabolism-dependent drug interaction potential of orally administered xenobiotics including drugs, botanical products, and dietary supplements. SIGNIFICANCE STATEMENT Enteric drug metabolism plays an important role in the bioavailability and metabolic fate of orally administered drugs as well as in enteric drug-drug and food-drug interactions. The current status of key enteric drug metabolism and disposition pathways and in vitro human cell-based enteric experimental systems for the evaluation of the metabolism and drug interaction potential of orally administered substances is reviewed.
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In vitro and in vivo human metabolism and pharmacokinetics of S- and R-praziquantel.
Kapungu, NN, Li, X, Nhachi, C, Masimirembwa, C, Thelingwani, RS
Pharmacology research & perspectives. 2020;(4):e00618
Abstract
Racemic praziquantel (PZQ) is the drug of choice for the treatment of schistosomiasis. R-Praziquantel (R-PZQ) has been shown as the therapeutic form, whereas S-PZQ is less efficacious and responsible for the bitter taste of the tablet. This study aimed at investigating the metabolism of R- and S-PZQ as this could have implications on efficacy and safety of racemate and R-PZQ specific formulations under development. In vitro CYP reaction phenotyping assay using 10 recombinant CYP (rCYP) isoenzymes showed hepatic CYP1A2, 2C19, 2D6, 3A4, and 3A5 were the major enzymes involved in metabolism of PZQ. Enzyme kinetic studies were performed by substrate depletion and metabolite formation methods, by incubating PZQ and its R- or S-enantiomers in human liver microsomes (HLM) and the rCYP enzymes. The effect of selective CYP inhibitors on PZQ metabolism was assessed in HLM. CYP1A2, 2C19, and 3A4 exhibited different catalytic activity toward PZQ, R- and S-enantiomers. Metabolism of R-PZQ was mainly catalyzed by CYP1A2 and CYP2C19, whereas metabolism of S-PZQ was mainly by CYP2C19 and CYP3A4. Based on metabolic CLint obtained through formation of hydroxylated metabolites, CYP3A4 was estimated to contribute 89.88% to metabolism of S-PZQ using SIMCYP® IVIVE prediction. Reanalysis of samples from a human PZQ-ketoconazole (KTZ) drug-drug interaction pharmacokinetic study confirmed these findings in that KTZ, a potent inhibitor of CYP3A, selectively increased area under the curve of S-PZQ by 68% and that of R-PZQ by just 9%. Knowledge of enantioselective metabolism will enable better understanding of variable efficacy of PZQ in patients and the R-PZQ formulation under development.
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Amino acid levels determine metabolism and CYP450 function of hepatocytes and hepatoma cell lines.
Boon, R, Kumar, M, Tricot, T, Elia, I, Ordovas, L, Jacobs, F, One, J, De Smedt, J, Eelen, G, Bird, M, et al
Nature communications. 2020;(1):1393
Abstract
Predicting drug-induced liver injury in a preclinical setting remains challenging, as cultured primary human hepatocytes (PHHs), pluripotent stem cell-derived hepatocyte-like cells (HLCs), and hepatoma cells exhibit poor drug biotransformation capacity. We here demonstrate that hepatic functionality depends more on cellular metabolism and extracellular nutrients than on developmental regulators. Specifically, we demonstrate that increasing extracellular amino acids beyond the nutritional need of HLCs and HepG2 cells induces glucose independence, mitochondrial function, and the acquisition of a transcriptional profile that is closer to PHHs. Moreover, we show that these high levels of amino acids are sufficient to drive HLC and HepG2 drug biotransformation and liver-toxin sensitivity to levels similar to those in PHHs. In conclusion, we provide data indicating that extracellular nutrient levels represent a major determinant of cellular maturity and can be utilized to guide stem cell differentiation to the hepatic lineage.