Characterization of enzyme activities of Cytochrome P450 enzymes, Flavin-dependent monooxygenases, N-acetyltransferases and UDP-glucuronyltransferases in human reconstructed epidermis and full-thickness skin models

Jäckh, Christine; Blatz, Veronika; Fabian, Eric; Guth, Katharina; Ravenzwaay, B. van; Reisinger, Kerstin; Landsiedel, Robert

doi:10.1016/j.tiv.2011.03.012

Cited by 43 publications

(24 citation statements)

References 22 publications

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“…Since previous skin metabolism studies used different experimental models (e.g., split-thickness explants, cells, or subcellular fractions), a wide range of test substrate concentrations [e.g., 500 mM of 4-methylumbelliferone in Jäckh et al (2011) versus 10 mM in the present study], or even different administration routes [topical triclosan in Moss et al (2000) versus media inclusion in the present study], direct comparison of the measured activity rates is challenging. If substrate depletion only is taken into account (i.e., conversion rate percentage), our results on triclosan glucuronidation and sulfation correlate well with the values published previously (Moss et al, 2000).…”

Section: Discussionmentioning

confidence: 99%

Phase II Metabolism in Human Skin: Skin Explants Show Full Coverage for Glucuronidation, Sulfation,N-Acetylation, Catechol Methylation, and Glutathione Conjugation

et al. 2014

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Although skin is the largest organ of the human body, cutaneous drug metabolism is often overlooked, and existing experimental models are insufficiently validated. This proof-of-concept study investigated phase II biotransformation of 11 test substrates in fresh full-thickness human skin explants, a model containing all skin cell types. Results show that skin explants have significant capacity for glucuronidation, sulfation, N-acetylation, catechol methylation, and glutathione conjugation. Novel skin metabolites were identified, including acyl glucuronides of indomethacin and diclofenac, glucuronides of 17b-estradiol, N-acetylprocainamide, and methoxy derivatives of 4-nitrocatechol and 2,3-dihydroxynaphthalene. Measured activities for 10 mM substrate incubations spanned a 1000-fold: from the highest 4.758 pmol·mg skin ·h -1 for 17b-estradiol 17-glucuronidation. Interindividual variability was 1.4-to 13.0-fold, the highest being 4-methylumbelliferone and diclofenac glucuronidation. Reaction rates were generally linear up to 4 hours, although 24-hour incubations enabled detection of metabolites in trace amounts. All reactions were unaffected by the inclusion of cosubstrates, and freezing of the fresh skin led to loss of glucuronidation activity. The predicted whole-skin intrinsic metabolic clearances were significantly lower compared with corresponding whole-liver intrinsic clearances, suggesting a relatively limited contribution of the skin to the body's total systemic phase II enzymemediated metabolic clearance. Nevertheless, the fresh full-thickness skin explants represent a suitable model to study cutaneous phase II metabolism not only in drug elimination but also in toxicity, as formation of acyl glucuronides and sulfate conjugates could play a role in skin adverse reactions.

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Section: Discussionmentioning

confidence: 99%

Phase II Metabolism in Human Skin: Skin Explants Show Full Coverage for Glucuronidation, Sulfation,N-Acetylation, Catechol Methylation, and Glutathione Conjugation

et al. 2014

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“…The K m of carbazeran 4-hydroxylation with partially purified human AO was 40 mM (Beedham et al, 1987), whereas Dalvie et al (2010) reported a K m of 3.4 mM for zoniporide 2-hydroxylation in pooled human liver cytosol. Data from existing literature suggest that cutaneous phase II metabolic enzymes, for example, UGTs, SULTs, and NATs (Luu-The et al, 2009;Bonifas et al, 2010a,b;Hu et al, 2010;Jäckh et al, 2011;Kushida et al, 2011;Götz et al, 2012b;van Eijl et al, 2012), have higher expression and activity levels compared with corresponding cutaneous phase I metabolic enzymes (Jäckh et al, 2011;Götz et al, 2012a). To compare the cutaneous activity of AO with the known cutaneous phase II reactions, this study tested the glucuronidation and sulfation of triclosan (Moss et al, 2000) and N-acetylation of p-toluidine (Götz et al, 2012b), together with hydroxylations of carbazeran and zoniporide.…”

Section: Discussionmentioning

confidence: 99%

“…Besides pharmacotherapy, daily life also exposes skin to numerous cosmetic ingredients and environmental xenobiotics, many of which share structural features with drugs and may cause drug interactions. Because biotransformation in human skin can lead to metabolic elimination of drugs, as well as to their activation or toxification (Sharma et al, 2013), understanding cutaneous drug metabolism is becoming increasingly important for drug discovery, safety, and development (Jäckh et al, 2011;Götz et al, 2012a,b;Gundert-Remy et al, 2014). Although AO was semiquantitatively detected in the human skin at the level of mRNA and protein (van Eijl et al, 2012), cutaneous enzyme activity of AO remains unknown.…”

Section: Introductionmentioning

confidence: 99%

Aldehyde Oxidase Activity in Fresh Human Skin

et al. 2014

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Human aldehyde oxidase (AO) is a molybdoflavoenzyme that commonly oxidizes azaheterocycles in therapeutic drugs. Although high metabolic clearance by AO resulted in several drug failures, existing in vitro-in vivo correlations are often poor and the extrahepatic role of AO practically unknown. This study investigated enzymatic activity of AO in fresh human skin, the largest organ of the body, frequently exposed to therapeutic drugs and xenobiotics. Fresh, full-thickness human skin was obtained from 13 individual donors and assayed with two specific AO substrates: carbazeran and zoniporide. Human skin explants from all donors metabolized carbazeran to 4-hydroxycarbazeran and zoniporide to 2-oxo-zoniporide. Average rates of carbazeran and zoniporide hydroxylations were 1.301 and 0.164 pmol×mg skin, resulting in 13 and 2% substrate turnover, respectively, after 24 hours of incubation with 10 mM substrate. Hydroxylation activities for the two substrates were significantly correlated (r 2 = 0.769), with interindividual variability ranging from 3-fold (zoniporide) to 6-fold (carbazeran). Inclusion of hydralazine, an irreversible inhibitor of AO, resulted in concentration-dependent decrease of hydroxylation activities, exceeding 90% inhibition of carbazeran 4-hydroxylation at 100 mM inhibitor. Reaction rates were linear up to 4 hours and well described by MichaelisMenten enzyme kinetics. Comparison of carbazeran and zoniporide hydroxylation with rates of triclosan glucuronidation and sulfation and p-toluidine N-acetylation showed that cutaneous AO activity is comparable to tested phase II metabolic reactions, indicating a significant role of AO in cutaneous drug metabolism. To our best knowledge, this is the first report of AO enzymatic activity in human skin.

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“…The constitutive levels of enzymes can be below the limits of detection, so exposure to an exogenous inducer is sometimes needed. 51,52 Some studies reported below and in Table 1 have measured mRNA expression. Even if this is not the most accurate method (compared with protein-level expression and activity), some authors claim that a correlation between mRNA levels and the physiological functions of corresponding genes is generally observed.…”

Section: Skin Versus Livermentioning

confidence: 99%

Review of the Availability ofIn VitroandIn SilicoMethods for Assessing Dermal Bioavailability

DumontCoralie

PrietoPilar

AsturiolDavid

et al. 2015

Applied In Vitro Toxicology

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The exposure of the skin to consumer products, drugs, and environmental chemicals can result in their penetrating the skin barrier and entering systemic circulation, potentially resulting in adverse effects in the skin and other organs. The assessment of dermal penetration and bioavailability (including penetration, metabolism, and entry into the systemic circulation) is therefore an important consideration in the risk assessment of chemicals. The skin is a heterogeneous organ with a multilayer structure. Based on its architecture and physiology, substances can penetrate through three major ways but can also be blocked in the different skin layers and in the skin appendages, which act as reservoir. In addition to that, as the skin is a metabolically competent organ, substances can undergo metabolism. After a brief description of the skin architecture, this review will focus on the skin penetration mechanisms and skin metabolic capacities. The skin absorption has traditionally been tested in vivo on animals. However, with the new legislation (i.e., Registration, Evaluation, Authorisation, and Restriction of Chemicals Regulation or Cosmetics Regulation), alternatives to animal testing have to be implemented. In a second part, this review will provide a description of the main in vitro and in silico or computational models available to study skin absorption and skin metabolism (i.e., ex vivo skin models, artificial membrane barriers, primary cells and cell lines, Quantitative Structure-Activity Relationship [QSAR], simulators for the prediction of skin metabolism).

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Characterization of enzyme activities of Cytochrome P450 enzymes, Flavin-dependent monooxygenases, N-acetyltransferases and UDP-glucuronyltransferases in human reconstructed epidermis and full-thickness skin models

Cited by 43 publications

References 22 publications

Phase II Metabolism in Human Skin: Skin Explants Show Full Coverage for Glucuronidation, Sulfation,N-Acetylation, Catechol Methylation, and Glutathione Conjugation

Phase II Metabolism in Human Skin: Skin Explants Show Full Coverage for Glucuronidation, Sulfation,N-Acetylation, Catechol Methylation, and Glutathione Conjugation

Aldehyde Oxidase Activity in Fresh Human Skin

Review of the Availability ofIn VitroandIn SilicoMethods for Assessing Dermal Bioavailability

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