CYP3A4 Activity in the Presence of Organic Cosolvents, Ionic Liquids, or Water‐Immiscible Organic Solvents

Chefson, Amandine; Auclair, Karine

doi:10.1002/cbic.200700128

Cited by 38 publications

(39 citation statements)

References 29 publications

(26 reference statements)

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“…Our results on the effects of organic solvents on LIPA metabolism, that DMSO had a higher inhibitory potential than acetonitrile and methanol, are consistent with previously published observations with CYP3A4 in cDNA-expressed human microsomes (Busby et al, 1999), human liver microsomes (Chauret et al, 1998;Chefson and Auclair, 2007), and our previous findings using testosterone as sub- *** *** *** *** *** strate in human hepatocytes (Easterbrook et al, 2001). The results suggest that acetonitrile and methanol are preferred solvents to be used at concentrations up to 1% when LIPA is chosen as a substrate for the quantification of CYP3A4 activity.…”

Section: Discussionsupporting

confidence: 93%

“…There are numerous reports on the inhibitory effects of organic solvents on P450-dependent metabolism (Chauret et al, 1998;Busby et al, 1999;Easterbrook et al, 2001;Chefson and Auclair, 2007). Our results on the effects of organic solvents on LIPA metabolism, that DMSO had a higher inhibitory potential than acetonitrile and methanol, are consistent with previously published observations with CYP3A4 in cDNA-expressed human microsomes (Busby et al, 1999), human liver microsomes (Chauret et al, 1998;Chefson and Auclair, 2007), and our previous findings using testosterone as sub- *** *** *** *** *** strate in human hepatocytes (Easterbrook et al, 2001).…”

Section: Discussionmentioning

confidence: 99%

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Evaluation of Luciferin-Isopropyl Acetal as a CYP3A4 Substrate for Human Hepatocytes: Effects of Organic Solvents, Cytochrome P450 (P450) Inhibitors, and P450 Inducers

Li¹

2009

Drug Metab Dispos

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ABSTRACT:This study represents the first report on the characterization of luciferin-isopropyl acetal (LIPA) as a CYP3A4 substrate in human hepatocytes. LIPA metabolism by human hepatocytes was found to be linear with time up to 120 min and followed Michaelis-Menten kinetics, with apparent K m value of 15 M and V max of 41 pmol/ min/million hepatocytes for the hepatocytes used in the study. The nonspecific cytochrome P450 (P450) inhibitor 1-aminobenzotriazole (ABT) and the CYP3A4-selective inhibitor ketoconazole (KTZ) caused concentration-dependent inhibition of LIPA metabolism, with more than 50% inhibition observed at the lowest concentrations evaluated of 7.8 M (ABT) and 0.78 M (KTZ), and near 100% inhibition observed at higher concentrations. Substantially lower inhibitory effects were observed for the non-CYP3A4 inhibitors diethyldithiocarbamate, furafylline, omeprazole, orphenadrine, sulfaphenazole, and quinidine. The commonly used organic solvents-acetonitrile, dimethyl sulfoxide (DMSO), and methanolwere found to inhibit LIPA metabolism, with approximately 50% inhibition at concentrations of 5, 1.25, and 5% (by volume), respectively. The comparatively higher inhibitory effects of DMSO relative to that for acetonitrile and methanol on LIPA metabolism were consistent with its known CYP3A4 inhibitory effects reported by others. LIPA metabolism in human hepatocytes was found to be induced by the treatment of human hepatocytes with the prototypical CYP3A4 inducers rifampin, carbamazepine, omeprazole, phenobarbital, and phenytoin but not by the CYP1A2 inducer 3-methylcholanthrene. Although the selectivity toward CYP3A4 needs to be definitively evaluated using cDNA-expressed P450 isoforms, the results suggest that LIPA is a suitable substrate to be used with human hepatocytes for the evaluation of CYP3A4 activities.Cytochrome P450 (P450) is an important family of enzymes responsible for xenobiotic metabolism. Of the multiple isoforms, P450 isoform 3A4 (CYP3A4) is the most abundant of the isoforms in the human liver. CYP3A4 has been found to be responsible for the metabolism of a large variety of exogenous and endogenous substrates (Guengerich, 2006;Zhou, 2008). In drug development, there is a need to evaluate the inhibitory and inductive potential of drug candidates toward CYP3A4 to estimate their drug-drug interaction potential with the myriad drugs that are substrates of this important P450 isoform (Lin and Lu, 1998;Li, 2001).Quantification of CYP3A4 activity of an enzyme system (e.g., liver microsomes, hepatocytes) is performed by incubation with a CYP3A4-selective substrate, followed by quantification of the metabolites formed. The most commonly used substrates are testosterone and midazolam, with 6␤-hydroxytestosterone (Chauret et al., 1998;Easterbrook et al., 2001) and 1Ј-hydroxymidazolam (Emoto andIwasaki, 2007), respectively, as the CYP3A4-mediated metabolites (Guengerich, 2006). Quantification of these metabolites requires the use of high-performance liquid chromatography (HPLC), often facilitated by ma...

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Section: Discussionsupporting

confidence: 93%

Section: Discussionmentioning

confidence: 99%

Evaluation of Luciferin-Isopropyl Acetal as a CYP3A4 Substrate for Human Hepatocytes: Effects of Organic Solvents, Cytochrome P450 (P450) Inhibitors, and P450 Inducers

Li¹

2009

Drug Metab Dispos

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“…Our group has recently reported a detailed survey of the activity of wild type human CYP3A4 in the presence of organic solvents and ionic liquids (ILs) (Chefson and Auclair, 2007a). The CYP3A4 enzyme activity was found to decrease significantly with the addition of solvents or ILs, except with biphasic solvent systems, which allowed retention of $85% of the activity (Chefson and Auclair, 2007a). We have also demonstrated that human CYP2D6 can retain most of its activity in selected biphasic solvent systems (Zhao et al, 2007).…”

Section: Introductionmentioning

confidence: 92%

“…A third mutant created by combination of the first two variants was > 2-fold more tolerant to DMSO (Kumar et al, 2006). Our group has recently reported a detailed survey of the activity of wild type human CYP3A4 in the presence of organic solvents and ionic liquids (ILs) (Chefson and Auclair, 2007a). The CYP3A4 enzyme activity was found to decrease significantly with the addition of solvents or ILs, except with biphasic solvent systems, which allowed retention of $85% of the activity (Chefson and Auclair, 2007a).…”

Section: Introductionmentioning

confidence: 96%

The activity of human CYP2D6 in low water organic solvents

Zhao

Auclair

2008

Biotech & Bioengineering

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P450 enzymes are of high interest for synthetic applications due to their ability to catalyze hydroxylation reactions at inactivated C-H bonds. The low solubility of many substrates in buffer, however, is limiting the applications of P450s. Our recent demonstration that the P450 enzymes CYP2D6 and CYP3A4 can function very well in biphasic solvent systems is one step towards overcoming this drawback, but is not practical when substrates or products are unstable in water, or with water-soluble products. An alternative strategy, which also facilitates enzyme recycling, is to directly resuspend lyophilized enzyme into nearly anhydrous organic solvents. Interestingly, we report here that CYP2D6 colyophilized with trehalose and suspended in n-decane shows higher activity than in aqueous buffer. This study demonstrates the unexpected high tolerance of CYP2D6 to some low water organic solvents and provides an alternative strategy to facilitate the use of this enzyme in synthesis.

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“…As enzyme deactivation would depend on its structure and hydrophobicity of the media, stability against the media [39,40] and enzyme conformation [40,41] are examined. Some extent of stability enhancement would be expected by immobilization, so that some biosensors can be prepared by using native enzymes [42,43].…”

Section: Stability Against Organic Mediamentioning

confidence: 99%

How to Lengthen the Long-Term Stability of Enzyme Membranes: Trends and Strategies

Yabuki

2017

Catalysts

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Abstract:In this review, factors that contribute to enhancing the stability of immobilized enzyme membranes have been indicated, and the solutions to each factor, based on examples, are discussed. The factors are divided into two categories: one is dependent on the improvement of enzyme properties, and the other, on the development of supporting materials. Improvement of an enzyme itself would effectively improve its properties. However, some novel materials or novel preparation methods are required for improving the properties of supporting materials. Examples have been provided principally aimed at improvements in membrane stability.

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CYP3A4 Activity in the Presence of Organic Cosolvents, Ionic Liquids, or Water‐Immiscible Organic Solvents

Cited by 38 publications

References 29 publications

Evaluation of Luciferin-Isopropyl Acetal as a CYP3A4 Substrate for Human Hepatocytes: Effects of Organic Solvents, Cytochrome P450 (P450) Inhibitors, and P450 Inducers

Evaluation of Luciferin-Isopropyl Acetal as a CYP3A4 Substrate for Human Hepatocytes: Effects of Organic Solvents, Cytochrome P450 (P450) Inhibitors, and P450 Inducers

The activity of human CYP2D6 in low water organic solvents

How to Lengthen the Long-Term Stability of Enzyme Membranes: Trends and Strategies

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