Functional analysis of UGT1A4 ^P24T and UGT1A4 ^L48V variant enzymes

Aim: To investigate the effects of two nonsynonymous SNPs, UGT1A4*2 (rs: 6755571, 70C>A, P24T) and UGT1A4* (rs: 2011425, 142T>G, L48V), on the function of UGT1A4 against dihydrotestosterone (DHT), transandrosterone (t-AND), lamotrigine (LTG) and tamoxifen (TAM). Materials & methods: Detailed kinetic experiments were conducted with recombinant UGT1A4 ^wild-type, UGT1A4 ^P24T and UGT1A4 ^L48V, which were overexpressed in HEK293 cell lines. The kinetic profiles and kinetic parameters (K _m, V _max and CL _int) obtained with either UGT1A4 ^P24T or UGT1A4 ^L48V were compared with those obtained with the wild-type enzyme. The interaction of TAM on UG1A4-catalyzed DHT glucuronidation was also investigated with the three UGT1A4 polymorphic enzymes. Results: UGT1A4 ^L48V had higher enzyme efficiency (CL _int) compared with wild-type UGT1A4 on DHT glucuronidation; UGT1A4 ^P24T and UGT1A4 ^L48V had lower CL _int than wild-type UGT1A4 for t-AND and LTG glucuronidation. The TAM CL _int with UGT1A4 ^P24T and UGT1A4 ^L48V glucuronidation and the UGT1A4 ^P24T-catalyzed DHT glucuronidation were, on the other hand, similar to those of the wild-type enzyme. With all three enzymes, TAM activated UGT1A4-catalyzed DHT glucuronidation in a concentration-dependent fashion. Conclusion: Decreased CL _int of UGT1A4 ^P24T and UGT1A4 ^L48V on LTG glucuronidation may lead to interindividual variations in LTG metabolism in vivo. However, it is less likely that these polymorphisms would have impact on DHT and t-AND metabolism in vivo because these compounds are glucuronidated by multiple enzymes.