CHARACTERIZATION OF THE CYTOCHROME-P450 ISOENZYMES INVOLVED IN THE IN-VITRO N-DEALKYLATION OF HALOPERIDOL

Aims The present study was carried out to identify the cytochrome P450 isoeneyme(s) involved in the N-dealkylation of haloperidol (HAL). Met hods In vitro studies were performed using human liver microsomes and c-DNA-expressed human P450 isoforms. N-dealkylation of HAL was assesse d by measuring the formation of 4-(4-chlorophenyl)-4-hydroxypiperidine (CPHP). Results There was a tenfold variation in the extent of CPHP f ormation amongst the nine human liver microsomal preparations. The CPH P formation rates as a function of substrate concentration, measured i n three Lives, followed monophasic enzyme kinetics. K-m and V-max valu es ranged respectively from 50 to 78 mu M and from 180 to 412 pmol mg( -1) min CPHP formation rates in the nine liver preparations were signi ficantly correlated with dextromethorphan N-demethylase activity (a ma rker of CYP3A4 activity), but not with the activity of dextromethorpha n O-demethylase (CYP2D6), phenacetin O-deethylase (CYP1A2) or tolbutam ide hydroxylase (CYP2C9). Ketoconazole, an inhibitor of CYP3A4, inhibi ted competitively CPHP formation (K-i=0.1 mu M), whereas sulphaphenazo le (CYP2C9), furafylline (CYP1A2) and quinidine (CYP2D6) gave only lit tle inhibition (IC50 > 100 mu M). CPHP formation was, moreover, enhanc ed by alpha-naphtoflavone, an effect common to CYP3A4 mediated reactio ns. Anti-CYP3A4 antibodies strongly inhibited CPHP formation, whereas no inhibition was observed in the presence of CYP2D6 antibodies. Among the recombinant human CYP isoforms tested, CYP3A4 exhibited the highe st activity with respect to CPMP formation rate, with no detectable ef fect of other CYP isoforms (CYP1A2, CYP2D6 and CYP2C9). HAL inhibited dextromethorphan O-demethylase (CYP2D6) with IC50 values between 2.7 a nd 8.5 mu M, but not (IC50 > 100 mu M) dextromethorphan N-demethylase (CYP3A4), phenacetin O-deethylase (CYP1A2) or tolbutamide hydroxylase (CYP2C9). Conclusions These results strongly suggest that the N-dealky lation of HAL in human liver microsomal preparations is mediated by CY P3A4.