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Development of a whole-cell screening system for evaluation of the human CYP1A2-mediated metabolism

Authors

  • Chih-Chieh Chu,

    1. Department of Chemical Engineering, National Tsing-Hua University, Hsin-Chu, Taiwan, Republic of China
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  • Kao-Lu Pan,

    1. Department of Chemical Engineering, National Tsing-Hua University, Hsin-Chu, Taiwan, Republic of China
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  • Hsien-Tsung Yao,

    1. Department of Nutrition, China Medical University, Taichung, Taiwan, Republic of China
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  • John Tsu-An Hsu

    Corresponding author
    1. National Health Research Institutes, Institute of Biotechnology and Pharmaceutical Research, No. 35, Keyan Road, Zhunan Town, Miaoli County 350, Taiwan, Republic of China; telephone: +886-37-246-166 ext. 35717; fax: +886-37-586-456
    2. Department of Biological Science and Technology, National Chiao Tung University, Hsin-Chu, Taiwan, Republic of China
    • National Health Research Institutes, Institute of Biotechnology and Pharmaceutical Research, No. 35, Keyan Road, Zhunan Town, Miaoli County 350, Taiwan, Republic of China; telephone: +886-37-246-166 ext. 35717; fax: +886-37-586-456.
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Abstract

Cytochrome P450 1A2 (CYP1A2) is an important member of cytochrome P450 involved in drug metabolism. In this study, a cell line, Huh7-1A2-I-E, with high expression level of CYP1A2 is established based on Huh7 cells. To achieve this, we constructed a recombinant lentiviral vector, pLenti-1A2-I-E, containing a single promoter encoding CYP1A2 followed by an internal ribosome entry site (IRES) to permit the translation of enhanced green fluorescence protein (EGFP). Such a design has greatly facilitated the selection of stable cell lines because the translations of CYP1A2 and EGFP proteins would be based on a single bi-cistronic mRNA. The Huh7-1A2-I-E cells were evaluated as a cell-based model for identification of CYP1A2 inhibitors and for studies of cytotoxicity resulted from CYP-mediated drug metabolism. Treatment of Huh7-1A2-I-E cells and the Huh7-E control cells with aflatoxin B1 showed that cells with CYP1A2 expression are much more sensitive to aflatoxin B1 and the cellular toxicity of aflatoxin B1 in Huh7-1A2-I-E cells could be prevented by furafylline, a CYP1A2 inhibitor. A collection of approximately 200 drugs were screened using this system and results indicate that for most drugs the metabolism by CYP1A2 is unlikely to have made a major contribution to the in vitro cytotoxicity except for thimerosal and evoxine. Several previously unidentified CYP1A2 inhibitors such as evoxine and berberine were also identified in this study. Biotechnol. Bioeng. 2011;108: 2932–2940. © 2011 Wiley Periodicals, Inc.

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