RT Journal Article
SR Electronic
T1 Functional characterization of 21 allelic variants of dihydropyrimidine dehydrogenase identified in 1,070 Japanese individuals
JF Drug Metabolism and Disposition
JO Drug Metab Dispos
FD American Society for Pharmacology and Experimental Therapeutics
SP dmd.118.081737
DO 10.1124/dmd.118.081737
A1 Eiji Hishinuma
A1 Yoko Narita
A1 Sakae Saito
A1 Masamitsu Maekawa
A1 Fumika Akai
A1 Yuya Nakanishi
A1 Jun Yasuda
A1 Masao Nagasaki
A1 Masayuki Yamamoto
A1 Hiroaki Yamaguchi
A1 Nariyasu Mano
A1 Noriyasu Hirasawa
A1 Masahiro Hiratsuka
YR 2018
UL http://dmd.aspetjournals.org/content/early/2018/05/16/dmd.118.081737.abstract
AB Dihydropyrimidine dehydrogenase (DPD, EC 1.3.1.2), encoded by the DPYD gene, is the rate-limiting enzyme in the degradation pathway of endogenous pyrimidine and fluoropyrimidine drugs such as 5-fluorouracil (5-FU). DPD catalyzes the reduction of uracil, thymine, and 5-FU. In Caucasians, DPYD mutations, including DPYD*2A, DPYD*13, c.2846A>T, and 1129-5923C>G/hapB3, are known to contribute to interindividual variations in the toxicity of 5-FU. However, none of these DPYD polymorphisms have been identified in the Asian population. Recently, 21 DPYD allelic variants, including some novel-single nucleotide variants (SNVs), were identified in 1,070 healthy Japanese individuals by analyzing their whole-genome sequences (WGS), but the functional alterations caused by these variants remain unknown. In this study, in vitro analysis was performed on 22 DPD allelic variants by transiently expressing wild-type DPD and 21 DPD variants in 293FT cells and characterizing their enzymatic activities, using 5-FU as a substrate. DPD expression levels and dimeric forms were determined using immunoblotting and blue native-PAGE, respectively. Additionally, the values of three kinetic parameters, the Michaelis constant (Km), maximum velocity (Vmax), and intrinsic clearance (CLint = Vmax/Km), were determined for the reduction of 5-FU. We found that 10 variants exhibited significantly decreased intrinsic clearance in comparison to wild-type DPD. Moreover, the band patterns observed in the immunoblots of blue native gels indicated that DPD dimerization is required for enzymatic activity in DPD. Thus, the detection of rare DPYD variants might facilitate severe adverse effect prediction of 5-FU-based chemotherapy in the Japanese population.