Methods for the quantitative evaluation and prediction of CYP enzyme induction using human<i>in vitro</i>systems

Honma, Masashi; Kozawa, Masanari; Suzuki, Hiroshi

doi:10.1517/17460441003762717

Cited by 6 publications

(11 citation statements)

References 197 publications

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“…While there are views that compound turnover may impact the outcome, inclusion and application of the parameter are not completely understood (Honma et al. ).…”

Section: Resultsmentioning

confidence: 99%

“…To minimize this effect compound is replaced daily. The consistent assessment, inclusion, and interpretation of compound turnover in such studies are still a matter of investigation and debate (Honma et al 2010).…”

Section: Discussionmentioning

confidence: 99%

“…As detailed in the Materials and Methods section, test compounds were replaced daily to minimize compound turnover, hence it was not measured throughout the analysis. While there are views that compound turnover may impact the outcome, inclusion and application of the parameter are not completely understood (Honma et al 2010).…”

Section: Comparison Of Cyp Inductionmentioning

confidence: 99%

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Evaluation of a novel PXR‐knockout in HepaRG^™ cells

Williamson

Lorbeer

Mitchell³

et al. 2016

Pharmacology Res & Perspec

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The nuclear pregnane X receptor (PXR) regulates the expression of genes involved in the metabolism, hepatobiliary disposition, and toxicity of drugs and endogenous compounds. PXR is a promiscuous nuclear hormone receptor (NHR) with significant ligand and DNA‐binding crosstalk with the constitutive androstane receptor (CAR); hence, defining the precise role of PXR in gene regulation is challenging. Here, utilising a novel PXR‐knockout (KO) HepaRG cell line, real‐time PCR analysis was conducted to determine PXR involvement for a range of inducers. The selective PXR agonist rifampicin, a selective CAR activator, 6‐(4‐chlorophenyl)imidazo[2,1‐b][1,3]thiazole‐5‐carbaldehyde O‐(3,4‐dichlorobenzyl)oxime (CITCO), and dual activators of CAR and PXR including phenobarbital (PB) were analyzed. HepaRG control cells (5F clone) were responsive to prototypical inducers of CYP2B6 and CYP3A4. No response was observed in the PXR‐KO cells treated with rifampicin. Induction of CYP3A4 by PB, artemisinin, and phenytoin was also much reduced in PXR‐KO cells, while the response to CITCO was maintained. This finding is in agreement with the abolition of functional PXR expression. The apparent EC50 values for PB were in agreement between the cell lines; however, CITCO was ~threefold (0.3 μmol/L vs. 1 μmol/L) lower in the PXR‐KO cells compared with the 5F cells for CYP2B6 induction. Results presented support the application of the novel PXR‐KO cells in the definitive assignment of PXR‐mediated CYP2B6 and CYP3A4 induction. Utilization of such cell lines will allow advancement in composing structure activity relationships rather than relying predominantly on pharmacological manipulations and provide in‐depth mechanistic evaluation.

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“…While there are views that compound turnover may impact the outcome, inclusion and application of the parameter are not completely understood (Honma et al. ).…”

Section: Resultsmentioning

confidence: 99%

Section: Discussionmentioning

confidence: 99%

Section: Comparison Of Cyp Inductionmentioning

confidence: 99%

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Evaluation of a novel PXR‐knockout in HepaRG^™ cells

Williamson

Lorbeer

Mitchell³

et al. 2016

Pharmacology Res & Perspec

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“…HepG2 (ATCC HB-8065), the most widely used human hepatoma cell line in metabolic research, contains functional CYP, but expression levels of most isoforms (CYP2B6, 2C9, 3A4) are 2-3 orders of magnitude below that of primary hepatocytes (Rodríguez-Antona et al, 2002). Thus, changes in CYP are difficult to detect in these cells without reconstituting each component of the transcriptional network through transfection with a series of plasmid constructs and pairing them with specific reporter genes to monitor response to treatments over time (Honma et al, 2010). Although this system may be useful for our particular application, transfection with multiple plasmids is a stressful procedure that is often not compatible with virus infection.…”

Section: Discussionmentioning

confidence: 99%

Evaluation of the HC-04 Cell Line as an In Vitro Model for Mechanistic Assessment of Changes in Hepatic Cytochrome P450 3A during Adenovirus Infection

et al. 2014

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HC-04 cells were evaluated as an in vitro model for mechanistic study of changes in the function of hepatic CYP3A during virus infection. Similar to in vivo observations, infection with a first generation recombinant adenovirus significantly inhibited CYP3A4 catalytic activity in an isoform-specific manner. Virus (MOI 100) significantly reduced expression of the retinoid X receptor (RXR) by 30% 96 hours after infection. Cytoplasmic concentrations of the pregnane X receptor (PXR) were reduced by 50%, whereas the amount of the constitutive androstane receptor (CAR) in the nuclear fraction doubled with respect to uninfected controls. Hepatocyte nuclear factor 4a (HNF-4a) and peroxisome proliferator-activated receptor g coactivator 1a (PGC-1a) were also reduced by ∼70% during infection. Virus suppressed CYP3A4 activity in the presence of the PXR agonist rifampicin and did not affect CYP3A4 activity in the presence of the CAR agonist CITCO [6-(4-chlorophenyl) imidazosuggesting that virus-induced modification of PXR may be responsible for observed changes in hepatic CYP3A4. The HC-04 cell line is easy to maintain, and CYP3A4 in these cells was responsive to known inducers and suppressors. Dexamethasone (200 mM) and phenobarbital (500 mM) increased activity by 230 and 124%, whereas ketoconazole (10 mM) and lipopolysaccharide (LPS) (10 mg/ml) reduced activity by 90 and 92%, respectively. This suggests that HC-04 cells can be a valuable tool for mechanistic study of drug metabolism during infection and for routine toxicological screening of novel compounds prior to use in the clinic.

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“…The function of drug-metabolizing enzymes and transporters is regulated by their expression at the transcriptional and posttranscriptional levels 22,23. The primary hepatocyte culture system contains nuclear receptors such as aromatic hydrocarbon receptor, pregnane X-receptor, and constitutive androstane receptor 20,24. Quantification of CYP messenger ribonucleic acid (mRNA) and protein levels and enzyme activities is recommended to evaluate drug interactions 25–27.…”

Section: Introductionmentioning

confidence: 99%

Effect of honokiol on the induction of drug-metabolizing enzymes in human hepatocytes

Cho¹,

Jeong²,

Kim³

et al. 2014

DDDT

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Honokiol, 2-(4-hydroxy-3-prop-2-enyl-phenyl)-4-prop-2-enyl-phenol, an active component of Magnolia officinalis and Magnolia grandiflora, exerts various pharmacological activities such as antitumorigenic, antioxidative, anti-inflammatory, neurotrophic, and antithrombotic effects. To investigate whether honokiol acts as a perpetrator in drug interactions, messenger ribonucleic acid (mRNA) levels of phase I and II drug-metabolizing enzymes, including cytochrome P450 (CYP), UDP-glucuronosyltransferase (UGT), and sulfotransferase 2A1 (SULT2A1), were analyzed by real-time reverse transcription polymerase chain reaction following 48-hour honokiol exposure in three independent cryopreserved human hepatocyte cultures. Honokiol treatment at the highest concentration tested (50 μM) increased the CYP2B6 mRNA level and CYP2B6-catalyzed bupropion hydroxylase activity more than two-fold in three different hepatocyte cultures, indicating that honokiol induces CYP2B6 at higher concentrations. However, honokiol treatment (0.5–50 μM) did not significantly alter the mRNA levels of phase I enzymes (CYP1A2, CYP3A4, CYP2C8, CYP2C9, and CYP2C19) or phase II enzymes (UGT1A1, UGT1A4, UGT1A9, UGT2B7, and SULT2A1) in cryopreserved human hepatocyte cultures. CYP1A2-catalyzed phenacetin O-deethylase and CYP3A4-catalyzed midazolam 1′-hydroxylase activities were not affected by 48-hour honokiol treatment in cryopreserved human hepatocytes. These results indicate that honokiol is a weak CYP2B6 inducer and is unlikely to increase the metabolism of concomitant CYP2B6 substrates and cause pharmacokinetic-based drug interactions in humans.

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Methods for the quantitative evaluation and prediction of CYP enzyme induction using humanin vitrosystems

Cited by 6 publications

References 197 publications

Evaluation of a novel PXR‐knockout in HepaRG^™ cells

Evaluation of a novel PXR‐knockout in HepaRG^™ cells

Evaluation of the HC-04 Cell Line as an In Vitro Model for Mechanistic Assessment of Changes in Hepatic Cytochrome P450 3A during Adenovirus Infection

Effect of honokiol on the induction of drug-metabolizing enzymes in human hepatocytes

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Methods for the quantitative evaluation and prediction of CYP enzyme induction using humanin vitrosystems

Cited by 6 publications

References 197 publications

Evaluation of a novel PXR‐knockout in HepaRG™ cells

Evaluation of a novel PXR‐knockout in HepaRG™ cells

Evaluation of the HC-04 Cell Line as an In Vitro Model for Mechanistic Assessment of Changes in Hepatic Cytochrome P450 3A during Adenovirus Infection

Effect of honokiol on the induction of drug-metabolizing enzymes in human hepatocytes

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Evaluation of a novel PXR‐knockout in HepaRG^™ cells

Evaluation of a novel PXR‐knockout in HepaRG^™ cells