Comparative N-glucuronidation kinetics of ketotifen and amitriptyline by expressed human UDP-glucuronosyltransferases and liver microsomes.

Like other basic amphiphilic drugs, the ( S )-enantiomer of the antiallergic drug ketotifen exhibited biphasic kinetics when it was converted to two isomeric quaternary ammonium-linked glucuronides in human liver microsomes. For ( R )-ketotifen this applied when incubations were carried out in the absence of a detergent. Two UDP-glucuronosyltransferases (UGTs) present in human liver, UGT1A4 and UGT1A3, were previously shown to catalyze tertiary amine N- glucuronidation when expressed in HK293 cells. Therefore, the conjugation kinetics of ( R )- and ( S )-ketotifen were investigated with the two expressed proteins. When homogenates of HK293 cells expressing UGT1A4 were incubated without detergent, N- glucuronidation kinetics were monophasic with K M values of 59 ± 5 μM for ( R )- and 86 ± 26 μM for ( S )-ketotifen. In experiments with membranes containing expressed UGT1A3, somewhat higher K M values were obtained. These values correspond to the high rather than to the low K M components of ketotifen glucuronidation in liver microsomes, the latter exhibiting K M values around 2 and 1 μM, respectively, with ( R )- and ( S )-ketotifen. With amitriptyline as the substrate, N- glucuronidation kinetics in the absence of detergent were biphasic in human liver microsomes and monophasic with a high K M value in cell homogenates containing UGT1A4. The results suggest that UGT1A4 and UGT1A3 catalyze high- K M N- glucuronidation of tertiary amine drugs, whereas the low- K M reaction requires either an alternative enzyme or a special conformation of UGT1A4 or UGT1A3 that can be attained in liver microsomes, but not in HK293 cell membranes.