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The use of transgenic cell lines for evaluating toxic metabolites of carbamazepine

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Abstract

Human lymphoblastoid cell lines transgenic for human CYP450s were evaluated for the identification of toxic metabolites of the anticonvulsant drug carbamazepine (CBZ). Human CYP450 isoforms expressed by these cell lines included 1A1, 1A2, 2E1, 2A6 and 3A4. A dose-dependent inhibition of population growth from 50–200 μg/ml CBZ was detected by measuring cell number and respiration. The inhibition increased with the growth rate of the various lines, which correlated inversely with the presence of CYP450s, and may have been caused by CBZ itself. Cytotoxicity was observed only at the highest dose and in the line lacking transfected CYP450s. Microsomal preparations from hCYP3A4/OR cells converted CBZ into its principal oxidative metabolite, carbamazepine-10,11-epoxide (CBZ-E), at a rate of 630 pmol/min per mg protein, confirming a major role of CYP3A4 in this reaction. However, no CBZ-E (or any metabolite) was recovered from any whole-cell incubation even though hCYP3A4 cells readily converted testosterone to 6ß-hydroxytestosterone. This suggests that differences exist between whole-cell and microsomal preparations of lymphoblastoid cells in their ability to metabolize CBZ.

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Abbreviations

BSTFA:

N,O-bis(trimethylsilyl)trifluoroacetamide

CBZ:

carbamazepine

CBZ-E:

carbamazepine-10, 11-epoxide

CYP450:

cytochrome P450

CYP3A4:

cytochrome P450, isoform 3A4

DMSO:

dimethyl sulfoxide

GC-MS:

gas chromatography-mass spectrometry

HPLC:

high-performance liquid chromatography

MTT:

(3-[4,5-dimethylthiazol-2-y1]-2,5-diphenyl)tetrazolium

SIM:

selected-ion monitoring

TMS:

trimethylsilyl

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Authors and Affiliations

  1. Division of Genetic Toxicology, Arkansas, USA

    C. R. Valentine & D. Casciano

  2. Dept. of Pediatrics, Section on Clinical Pharmacology, University of Arkansas for Medical Sciences and Arkansas Childrens Hospital, Little Rock, Arkansas, USA

    J. L. Valentine

  3. Division of Genetic Toxicology, National Center for Toxicological Research, 3900 NCTR Road, HFT-120, 72079-9501, Jefferson, AR, USA

    J. Leakey

Authors
  1. C. R. Valentine
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  2. J. L. Valentine
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  3. J. Leakey
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  4. D. Casciano
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Valentine, C.R., Valentine, J.L., Leakey, J. et al. The use of transgenic cell lines for evaluating toxic metabolites of carbamazepine. Cell Biol Toxicol 12, 155–165 (1996). https://doi.org/10.1007/BF00148169

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  • DOI: https://doi.org/10.1007/BF00148169

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