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Construction and application of double-transfected cells expressing the human transporter P-glycoprotein and cytochrome P450 3A4.
Hu, H H; Su, C; Jiang, Y; Yu, L S; Liu, Y; Tian, Y; Xu, S Y; Zhou, H; He, X; Jiang, H D; Zeng, S.
Afiliação
  • Hu HH; College of Pharmaceutical Sciences, Zhejiang University, Zhejiang, China.
Pharmazie ; 68(10): 816-20, 2013 Oct.
Article em En | MEDLINE | ID: mdl-24273886
Intestinal P-glycoprotein (P-gp) and cytochrome P450 (CYP) enzymes are known to influence oral bioavailabilities of drugs. Recombinant plasmids pcDNA3.1/Hypgro/CYP3A4 were transfected into MDCK and MDCK-MDR1 cells to construct the single-transfected cell line MDCK-CYP3A4 and double-transfected cell line MDCK-MDR1/CYP3A4. The expression of CYP3A4 in the double-transfected cell line was determined by Western blot and its activity was detected by the metabolism assays of three substrates of CYP3A4, which were 7-benzyloxy-4-trifluoro-methylcoumarin (BFC), testosterone and midazolam. In addition, the selection of monoclones with high CYP3A4 activities in the single-tranfected cell line was performed by the P450 Glo CYP3A4 assay. Through MTT assay, the interaction between P-gp and CYP3A4 was preliminarily determined based on the changes of IC50 values. The results showed that paclitaxel detoxified in the single-transfected MDCK-MDR1 cell because of P-gp efflux. And it was also less toxic in the single-transfected CYP3A4 cell line due to the metabolism by CYP3A4. In the double-transfected MDCK-MDR1/CYP3A4 cell line, the toxicity decreased dramatically because of the interplay between P-gp and CYP3A4. Therefore, the cell model could be applied to study the toxicity and detoxification of chemicals due to the metabolism by CYP3A4 and the efflux through P-gp.
Assuntos
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Coleções: 01-internacional Base de dados: MEDLINE Assunto principal: Membro 1 da Subfamília B de Cassetes de Ligação de ATP / Citocromo P-450 CYP3A Limite: Animals / Humans Idioma: En Ano de publicação: 2013 Tipo de documento: Article
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Coleções: 01-internacional Base de dados: MEDLINE Assunto principal: Membro 1 da Subfamília B de Cassetes de Ligação de ATP / Citocromo P-450 CYP3A Limite: Animals / Humans Idioma: En Ano de publicação: 2013 Tipo de documento: Article