Dihydropyrimidine dehydrogenase activity in 150 healthy Japanese volunteers and identification of novel mutations.
OBJECTIVE
Dihydropyrimidine dehydrogenase (DPD) is the initial and rate-limiting enzyme catalyzing the metabolic degradation of the anticancer drug 5-fluorouracil (5-FU). Population studies of DPD activity in peripheral blood mononuclear cells (PBMC) were reported in healthy volunteers and cancer patients. Although these studies were done in mainly Caucasian and African American populations, only a little information is available for a Japanese population.


METHODS
One hundred fifty healthy Japanese volunteers were screened for a population distribution of PBMC-DPD activity. Genetic analysis of a volunteer with very low DPD activity was carried out by reverse transcriptase-PCR and genomic sequencing. Bacterially expressed recombinant mutant DPD proteins were purified and characterized.


RESULTS
Mean and median values of PBMC-DPD activity for 5-FU reduction in the study population were 0.173 and 0.166 nmol/min/mg protein, respectively. A 57-year-old female volunteer (proband in this study) had very low DPD activity (0.014 nmol/min/mg protein) with a very low level of expression of DPD protein. Two novel nucleotide substitutions, at nucleotide positions 1097 (1097G > C) and 2303 (2303C > A), resulting in amino acid substitutions at positions 366 (G366A) and 768 (T768K), respectively, were identified. The G366A mutation caused not only a marked decrease in the affinity of the enzyme to cofactor NADPH but also reduced Vmax for 5-FU-reducing activity to approximately 0.5. T768K mutant lost its activity much faster than did wild DPD.


CONCLUSIONS
We found one healthy volunteer (0.7% of the population) with very low PBMC-DPD activity due to heterozygosity for a mutant allele of the DPYD gene in a population of 150 Japanese.
Ogura
Kenichiro
K
Department of Drug Metabolism and Molecular Toxicology, School of Pharmacy, Tokyo University of Pharmacy and Life Science, Hachioji-shi, Tokyo, Japan.
Ohnuma
Tomokazu
T
Minamide
Yoshiyuki
Y
Mizuno
Atsuhiro
A
Nishiyama
Takahito
T
Nagashima
Satoru
S
Kanamaru
Mitsutaka
M
Hiratsuka
Akira
A
Watabe
Tadashi
T
Uematsu
Toshihiko
T
D000328:Adult; D000964:Antimetabolites, Antineoplastic / Q000378:metabolism; D004252:DNA Mutational Analysis; D042943:Dihydrouracil Dehydrogenase (NADP) / Q000235:genetics* / Q000378:metabolism*; D005260:Female; D005472:Fluorouracil / Q000378:metabolism; D005820:Genetic Testing; D006801:Humans; D007564:Japan; D008297:Male; D008875:Middle Aged
D000964:Antimetabolites, Antineoplastic; D042943:Dihydrouracil Dehydrogenase (NADP); D005472:Fluorouracil