Evaluation of CYP3A4 inhibition and hepatotoxicity using DMSO-treated human hepatoma HuH-7 cells.

Therapeutic Approaches

Abstract

A human hepatoma cell line (HuH-7) was evaluated as a metabolically competent cell model to investigate cytochrome P450 3A4 (CYP3A4) inhibition, induction, and hepatotoxicity. First, CYP3A4 gene expression and activity were determined in HuH-7 cells under three culture conditions: 1-week culture, 3-week culture, or 1 % dimethyl sulfoxide (DMSO) treatment. HuH-7 cells treated with DMSO for 2 weeks after confluence expressed the highest CYP3A4 gene expression and activity compared to the other two culture conditions. Furthermore, CYP3A4 activity in DMSO-treated HuH-7 cells was compared to that in a human hepatoma cell line (HepG2/C3A) and human bipotent progenitor cell line (HepaRG), which yielded the following ranking: HepaRG > DMSO-treated HuH-7 > HepG2/C3A cells. The effects of three known CYP3A4 inhibitors were evaluated using DMSO-treated HuH-7 cells. CYP3A4 enzyme inhibition in HuH-7 cells was further compared to human recombinant CYP3A4, indicating similar potency for reversible inhibitors (IC 50 within 2.5-fold), but different potency for the irreversible inhibitor. Next, induction of CYP3A4 activity was compared between DMSO-treated HuH-7 and HepaRG cells using two known inducers. DMSO-treated HuH-7 cells yielded minimal CYP3A4 induction compared to that in the HepaRG cells after 48-h treatments. Finally, the cytotoxicity of five known hepatotoxicants was evaluated in DMSO-treated HuH-7, HepG2/C3A, and HepaRG cells, and significant differences in cytotoxic sensitivity were observed. Overall, DMSO-treated HuH-7 cells are a valuable model for medium- or high-throughput screening of chemicals for CYP3A4 inhibition and hepatotoxicity.

Authors

Liu, Yitong; Flynn, Thomas J; Xia, Menghang; Wiesenfeld, Paddy L; Ferguson, Martine S;

Keywords

  • Carcinoma, Hepatocellular/ enzymology
  • Cell Line, Tumor
  • Cytochrome P-450 CYP3A/ biosynthesis
  • Cytochrome P-450 CYP3A/ genetics
  • Cytochrome P-450 CYP3A/ metabolism
  • Cytochrome P-450 Enzyme Inhibitors/ pharmacology
  • Dimethyl Sulfoxide/ pharmacology
  • Gene Expression/ drug effects
  • Hep G2 Cells
  • Hepatocytes/ drug effects
  • Hepatocytes/ enzymology
  • Humans
  • Liver Neoplasms/ enzymology
  • Toxicity Tests/ methods

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