Stable expression of human cytochrome P450 3A4 in V79 cells and its application for metabolic profiling of ergot derivatives.
Expression of human cytochrome (CYP) in heterologous cells is a means of specifically studying the role of these enzymes in drug metabolism. The complete cDNA encoding CYP3A4 (PCN1) was inserted into an expression vector containing the strong myeloproliferative sarcoma virus promoter in combination with the enhancer of the cytomegalovirus and stably expressed in V79 Chinese hamster cells. The presence of genomically integrated CYP3A4 cDNA cell clones was confirmed by polymerase chain reaction analysis. Transcription was detected by reverse transcribed polymerase chain reaction analysis. Functional expression could be demonstrated by conversion of testosterone to the specific 6beta-hydroxylated product. In recombinant V79 cells expressing CYP3A4 about 6% of the substrate was converted to 6beta-hydroxytestosterone. The metabolism of two dopaminergic ergot derivatives was investigated in live recombinant V79 cells. Both lisuride and terguride were monodeethylated.
Rauschenbach
R
R
Research Laboratories, Schering AG, Berlin, Germany.
Gieschen
H
H
Husemann
M
M
Salomon
B
B
Hildebrand
M
M
D000818:Animals; D002460:Cell Line; D002478:Cells, Cultured; D003001:Cloning, Molecular; D006224:Cricetinae; D003412:Cricetulus; D051544:Cytochrome P-450 CYP3A; D003577:Cytochrome P-450 Enzyme System / Q000096:biosynthesis* / Q000235:genetics*; D018491:Dopamine Agonists / Q000378:metabolism*; D005347:Fibroblasts / Q000201:enzymology*; D015971:Gene Expression Regulation, Enzymologic; D005822:Genetic Vectors; D006801:Humans; D008090:Lisuride / Q000031:analogs & derivatives / Q000378:metabolism; D008099:Liver / Q000201:enzymology; D006899:Mixed Function Oxygenases / Q000096:biosynthesis* / Q000235:genetics*; D010957:Plasmids; D016133:Polymerase Chain Reaction; D013739:Testosterone / Q000378:metabolism; D014158:Transcription, Genetic; D014162:Transfection
D018491:Dopamine Agonists; C006208:dironyl; D013739:Testosterone; D003577:Cytochrome P-450 Enzyme System; D008090:Lisuride; D006899:Mixed Function Oxygenases; C104464:CYP3A protein, human; D051544:Cytochrome P-450 CYP3A; C510163:CYP3A4 protein, human