Coding region mutations in human UPD-glucuronosyltransferase 1A1 (UGT1A1) impair bilirubin and xenobiotic glucuronidation: PO-350

UGT1A1 is the sole enzyme involved in the glucuronidation ofbilirubin, and promoter and coding region polymorphisms in UGT1A1are associated with inherited disorders related to bilirubin elimination. In Asian populations, three coding regionpolymorphisms, UGT1A1*6 (G71R), UGT1A1*27 (P229Q) andUGT1A1*62 (F83L), have been implicated in Gilbert’s syndromewhile UGT1A1*7 (Y486D) has been linked to the Crigler Najjarsyndrome type II. However, the impact of these mutations onUGT1A1 glucuronidation kinetics and substrate selectivityremains unknown. Thus, studies were performed to investigate theeffects of these four coding region mutations on UGT1A1 activityand substrate selectivity. The UGT1A1 variants were generated by site-directed mutagenesis using the wild-type cDNA as template. Wild-type UGT1A1 and the UGT1A1 variants were stably expressed in a mammalian (HEK293) cell line, and activityof cell lysates was measured using 4-methylumbelliferone (4 MU), 1-naphthol (1NP), bilirubin and ethinylestradiol as the model substrates. 4 MU, bilirubin and ethinylestradiol glucuronidation byUGT1A1, UGT1A1*6 and UGT1A1*27 exhibited substrate inhibition kinetics. In contrast, 1NP glucuronidation by UGT1A1,UGT1A1*6 and UGT1A1*27 exhibited sigmoidal kinetics. Anapproximate 50% reduction in intrinsic clearance (Vmax/Km) wasobserved for the glucuronidation of all 4 substrates by UGT1A1*6and UGT1A1*27. UGT1A1*7 and UGT1A1*62 exhibited verylow activity towards 4 MU, 1NP bilirubin and ethinylestradiol. It is concluded that UGT1A1 coding region mutations associatedwith impaired bilirubin elimination also reduce xenobiotic glucuronidation to a similar extent.