Characterization of <i>in vitro</i> metabolites of deoxypodophyllotoxin in human and rat liver microsomes using liquid chromatography/tandem mass spectrometry

Lee, Sang Kyu; Jun, In Hye; Yoo, Hye Hyun; Kim, Ju Hyun; Seo, Young Min; Kang, Mi Jeong; Lee, Seung Ho; Jeong, Tae Cheon; Kim, Dong Hyun

doi:10.1002/rcm.3325

Cited by 13 publications

(14 citation statements)

References 14 publications

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“…The released carbene is unstable and would react with water to ultimately produce catechol. 39, 40 An additional pathway leads to the formation of the inactive P420 ( k 6 ) from the enzyme-inhibitor complex (EI), which degrades to an unknown product ( k 7 ). The estimated parameters (with their respective standard deviations) are presented in Table 1 for both CYP3A4 baculosomes and HLM, whereas two representative fittings for CYP3A4 baculosomes and HLM are shown in Figures 4a and 4b, respectively.…”

Section: Resultsmentioning

confidence: 99%

Mechanism-Based Inhibition of CYP3A4 by Podophyllotoxin: Aging of an Intermediate Is Important for in Vitro/in Vivo Correlations

et al. 2016

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An in vitro observation of time-dependent-inhibition of metabolic enzymes often results in removing a potential drug from the drug-pipeline. However, the accepted method for predicting TDIs of the important drug metabolizing cytochrome P450 enzymes often overestimates the drug interaction potential. Better models that take into account the complexities of the cytochrome P450 enzyme system will lead to better predictions. Herein we report the use of our previously described models for complex kinetics of podophyllotoxin. Spectral characterization of the kinetics indicates that an intermediate MI-complex is formed, which slowly progresses to an essentially irreversible MI-complex. The intermediate MI-complex can release free enzyme during the time-course of a typical 30 minute TDI experiment. This slow rate of MI-complex conversion results in an over-prediction of the kinact value if this process is not included in the analysis of the activity versus time profile. In vitro kinetic experiments in rat liver microsomes predicted a lack of drug interaction between podophyllotoxin and midazolam. In vivo rat pharmacokinetic studies confirmed this lack of drug interaction.

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Section: Resultsmentioning

confidence: 99%

Mechanism-Based Inhibition of CYP3A4 by Podophyllotoxin: Aging of an Intermediate Is Important for in Vitro/in Vivo Correlations

et al. 2016

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“…Chemical and pharmacological research on J. rigida has led to the discovery of bioactive components, such as flavonoids and phenolic compounds that potentially contribute to its antioxidant activity (Robards et al, 1999; Woo et al, 2011; Taviano et al, 2013). Moreover, lignans and flavonoids from J. rigida , such as catechin, podophyllotoxin, and amentoflavone, exert strong biological effects, including anti-inflammatory, anticancer, and antiviral activities (Lee et al, 2008; Gordien et al, 2009; Ryu et al, 2010; Lesjak et al, 2011, 2013, 2014; Jeong et al, 2012). …”

Section: Introductionmentioning

confidence: 99%

Quality Evaluation of Juniperus rigida Sieb. et Zucc. Based on Phenolic Profiles, Bioactivity, and HPLC Fingerprint Combined with Chemometrics

Liu

Wang

et al. 2017

Front. Pharmacol.

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Juniperus rigida (J. rigida) which is endemic to East Asia, has traditionally been used as an ethnomedicinal plant in China. This study was undertaken to evaluate the quality of J. rigida samples derived from 11 primary regions in China. Ten phenolic compounds were simultaneously quantified using reversed-phase high-performance liquid chromatography (RP-HPLC), and chlorogenic acid, catechin, podophyllotoxin, and amentoflavone were found to be the main compounds in J. rigida needles, with the highest contents detected for catechin and podophyllotoxin. J. rigida from Jilin (S9, S10) and Liaoning (S11) exhibited the highest contents of phenolic profiles (total phenolics, total flavonoids and 10 phenolic compounds) and the strongest antioxidant and antibacterial activities, followed by Shaanxi (S2, S3). A similarity analysis (SA) demonstrated substantial similarities in fingerprint chromatograms, from which 14 common peaks were selected. The similarity values varied from 0.85 to 0.98. Chemometrics techniques, including hierarchical cluster analysis (HCA), principal component analysis (PCA), and discriminant analysis (DA), were further applied to facilitate accurate classification and quantification of the J. rigida samples derived from the 11 regions. The results supported HPLC data showing that all J. rigida samples exhibit considerable variations in phenolic profiles, and the samples were further clustered into three major groups coincident with their geographical regions of origin. In addition, two discriminant functions with a 100% discrimination ratio were constructed to further distinguish and classify samples with unknown membership on the basis of eigenvalues to allow optimal discrimination among the groups. Our comprehensive findings on matching phenolic profiles and bioactivities along with data from fingerprint chromatograms with chemometrics provide an effective tool for screening and quality evaluation of J. rigida and related medicinal preparations.

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“…The inhibitory effects of known P450 isoform-selective inhibitors and the metabolism of KRO-105714 were evaluated to determine the P450 isoforms involved in the metabolic pathway. Well characterized P450-selective inhibitors-i.e., a-naphthoflavone for CYP1A2 (1 and 5 mM), tranylcypromine for CYP2A6 (1 and 5 mM), quercetin for CYP2C8 (5 and 20 mM), fluconazole for CYP2C9 (5 and 10 mM), ticlopidine for CYP2C19 (2 and 10 mM), quinidine for CYP2D6 (10 and 50 mM), diethyldithiocarbamate for CYP2E1 (20 and 100 mM), and ketoconazole for CYP3A4 (2 and 10 mM)-were incubated with KRO-105714 (Newton et al, 1995;Ko et al, 2000;Lee et al, 2008;Khojasteh et al, 2011). Incubations were performed with P450-selective inhibitor, pooled HLMs (1 mg/ml) and KR-105714 (50 mM).…”

Section: Methodsmentioning

confidence: 99%

Identification of Metabolites of N-(5-Benzoyl-2-(4-(2-Methoxyphenyl)piperazin-1-yl)thiazol-4-yl)pivalamide Including CYP3A4-Mediated C-Demethylation in Human Liver Microsomes with High-Resolution/High-Accuracy Tandem Mass

et al. 2014

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KRO-105714 [N-(5-benzoyl-2-(4-(2-methoxyphenyl)piperazin-1-yl) thiazol-4-yl)pivalamide] is a 2,4,5-trisubstituted 1,3-thiazole derivative that exerts anti-atopic dermatitis activity via robust suppression of the sphingosylphosphorylcholine receptor. This study used high-resolution/high-accuracy tandem mass spectroscopy (HRMS) and recombinant cDNA-expressed cytochrome P450 (P450) isoforms to identify the metabolic pathway and metabolites of KRO-105714 in human liver microsomes (HLMs) as therapeutic agents for inflammation. The incubation of KRO-105714 with pooled HLMs in the presence of NADPH generated four metabolites (M1-M4). The metabolites were identified using HRMS and confirmed using synthetic standards for M2 and M4. M1 and M2 were identified as monohydroxylated metabolites, and M3 and M4 were identified as O-demethyl KRO-105714 and C-demethyl KRO-105714, respectively. In the inhibition study with selective CYP3A4 inhibitors and incubation in recombinant cDNA-expressed P450 enzymes, all the metabolites of KRO-105714 were formed by CYP3A4 in HLMs. The CYP3A4-mediated formation of M4 from M2 was confirmed via incubation of M2 in HLMs. These results showed that the unusual C-demethylated metabolite M4 was generated from monohydroxyl metabolite M2 via a CYP3A4-mediated enzymatic reaction in HLMs.

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Characterization of in vitro metabolites of deoxypodophyllotoxin in human and rat liver microsomes using liquid chromatography/tandem mass spectrometry

Cited by 13 publications

References 14 publications

Mechanism-Based Inhibition of CYP3A4 by Podophyllotoxin: Aging of an Intermediate Is Important for in Vitro/in Vivo Correlations

Mechanism-Based Inhibition of CYP3A4 by Podophyllotoxin: Aging of an Intermediate Is Important for in Vitro/in Vivo Correlations

Quality Evaluation of Juniperus rigida Sieb. et Zucc. Based on Phenolic Profiles, Bioactivity, and HPLC Fingerprint Combined with Chemometrics

Identification of Metabolites of N-(5-Benzoyl-2-(4-(2-Methoxyphenyl)piperazin-1-yl)thiazol-4-yl)pivalamide Including CYP3A4-Mediated C-Demethylation in Human Liver Microsomes with High-Resolution/High-Accuracy Tandem Mass

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