AIM:Over 90% of drugs are metabolized by the cytochromeP-450 (CYP) family of liver isoenzymes.The most importantenzymes are CYP1A2,3A4,2C9/19,2D6 and 2E1.AlthoughCYP2D6 accounts for <2% of the total CYP liver enzymecontent,it mediates metabolism in almost 25% of drugs.Inorder to study its enzymatic activity for drug metabolism,itscDNA was cloned and a HepG2 cell line stably expressingCYP2D6 was established.METHODS:Human CYP2D6 cDNA was amplified withreverse transcription-polymerase chain reaction (RT-PCR)from total RNA extracted from human liver tissue and clonedinto pGEM-T vector,cDNA segment was identified by DNAsequencing and subcloned into a mammalian expressionvector pREP9.A cell line was established by transfectingthe recombinant plasmid of pREP9-CYP2D6 to hepatomaHepG2 cells.Expression of mRNA was validated by RT-PCR.Enzyme activity of catalyzing dextromethorphan O-demethylation in postmitochondrial supernant (S9) fractionof the cells was determined by high performance liquidchromatography (HPLC).RESULTS:The cloned cDNA had 4 base differences,e.g.100 C→T,336 T→C,408 C→G and 1 457 G→C,whichresulted in P34S,and S486T amino acid substitutions,andtwo samesense mutations were 112 F and 136 V comparedwith that reported by Kimura et al (GenBank accessionnumber:M33388).P34S and S486T amino acid substitutionswere the characteristics of CYP2D6*10 allele.The relativeactivity of S9 fraction of HepG2-CYP2D6*10 metabolizeddetromethorphan O-demethylation was found to be2.31±0.19 nmol·min~(-1)·mg~(-1) S9 protein (n=3),but wasundetectable in parental HepG2 cells.CONCLUSION:cDNA of human CYP2D6*10 can be suocessfullycloned.A cell line,HepG2-CYP2D6*10,expressing CYP2D6*10mRNA and having metabolic activity,has been established. AIM: Over 90% of drugs are metabolized by the cytochrome P-450 (CYP) family of liver isoenzymes. The most importantenzymes are CYP1A2, 3A4, 2C9 / 19, 2D6 and 2E1.Although CYP2D6 accounts for <2% of the total CYP liver enzyme content , it mediates metabolism in almost 25% of drugs.Inorder to study its enzymatic activity for drug metabolism, its cDNA was cloned and a HepG2 cell line stably expressing CYP2D6 was established. METHODS: Human CYP2D6 cDNA was amplified with reverse transcription-polymerase chain reaction (RT- PCR) from total RNA extracted from human liver tissue and clonedinto pGEM-T vector, cDNA segment was identified by DNA sequencing and subcloned into mammalian expression vector pREP9.A cell line was established by transfecting the recombinant plasmid of pREP9-CYP2D6 to hepatomaHepG2 cells. Expression of mRNA was validated by RT-PCR. Enzyme activity of catalyzing dextromethorphan O-demethylation in postmitochondrial supernant (S9) fractionof the cells was determined by high performance liquidchr Results: o cloning (HPLC) .RESULTS: The cloned cDNA had 4 base differences, eg 100 C → T, 336 T → C, 408 C → G and 1 457 G → C, which was comprised in P34S, and S486T amino acid substitutions, were 112 F and 136 V compared with that reported by Kimura et al (GenBank accession number: M33388). P34S and S486T amino acid substitutionswere the characteristics of CYP2D6 * 10 allele. the relative activity of S9 fraction of HepG2-CYP2D6 * 10 metabolizeddetromethorphan O-demethylation was found to be 2.31 ± 0.19 nmol · min -1 · mg -1 S9 protein (n = 3), but was undetectable in parental HepG2 cells.CONCLUSION: cDNA of human CYP2D6 * 10 can be suocessfully cloned.A cell line, HepG2-CYP2D6 * 10, expressing CYP2D6 * 10 mRNA and having metabolic activity, has been established.