CYP2A13 expressed in human bladder metabolically activates 4-aminobiphenyl.
Cigarette smoking is the predominant risk factor for bladder cancer. Aromatic amines such as 4-aminobiphenyl (ABP) is the major carcinogens found in tobacco smoke. Although it is generally accepted that ABP is metabolically activated via N-hydroxylation by CYP1A2 in human liver, previous studies using Cyp1a2-null mice indicated the involvement of other enzyme(s). Here we found that CYP2A13 can metabolically activate ABP to show genotoxicity by Umu assay. The K(m) and V(max) values for ABP N-hydroxylation by recombinant CYP2A13 in E. coli were 38.5 +/- 0.6 microM and 7.8 +/- 0.0 pmol/min/pmol CYP, respectively. The K(m) and V(max) values by recombinant CYP1A2 were 9.9 +/- 0.9 microM and 39.6 +/- 0.9 pmol/min/pmol CYP, respectively, showing 20-fold higher intrinsic clearance than CYP2A13. In human bladder, CYP2A13 mRNA, but not CYP1A2, is expressed at a relatively high level. Human bladder microsomes showed ABP N-hydroxylase activity (K(m) = 34.9 +/- 4.7 microM and V(max) = 57.5 +/- 1.9 pmol/min/mg protein), although the intrinsic clearance was 5-fold lower than that in human liver microsomes (K(m) = 33.2 +/- 2.0 microM and V(max) = 293.9 +/- 5.8 pmol/min/mg protein). The activity in human bladder microsomes was prominently inhibited by 8-methoxypsoralen, but not by fluvoxamine, anti-CYP1A2 or anti-CYP2A6 antibodies. CYP2S1, which is expressed in human bladder and has relatively high amino acid identities with CYP2As, did not show detectable ABP N-hydroxylase activity. In conclusion, although the enzyme responsible for ABP N-hydroxylation in human bladder microsomes could not be determined, we found that CYP2A13 metabolically activates ABP.
Nakajima
Miki
M
Drug Metabolism and Toxicology, Division of Pharmaceutical Sciences, Graduate School of Medical Science, Kanazawa University, Kakuma-machi, Kanazawa, Japan.
Itoh
Masahiro
M
Sakai
Haruko
H
Fukami
Tatsuki
T
Katoh
Miki
M
Yamazaki
Hiroshi
H
Kadlubar
Fred F
FF
Imaoka
Susumu
S
Funae
Yoshihiko
Y
Yokoi
Tsuyoshi
T
D000611:Aminobiphenyl Compounds / Q000493:pharmacokinetics*; D001189:Aryl Hydrocarbon Hydroxylases / Q000235:genetics / Q000378:metabolism*; D001483:Base Sequence; D001711:Biotransformation; D017931:DNA Primers; D006801:Humans; D008861:Microsomes / Q000201:enzymology; D006899:Mixed Function Oxygenases / Q000378:metabolism; D012333:RNA, Messenger / Q000235:genetics; D011994:Recombinant Proteins / Q000235:genetics / Q000378:metabolism; D020133:Reverse Transcriptase Polymerase Chain Reaction; D001743:Urinary Bladder / Q000201:enzymology*
D000611:Aminobiphenyl Compounds; D017931:DNA Primers; D012333:RNA, Messenger; D011994:Recombinant Proteins; C006757:4-biphenylamine; D006899:Mixed Function Oxygenases; C027685:4-aminobiphenyl N-hydroxylase; D001189:Aryl Hydrocarbon Hydroxylases; C414206:CYP2A13 protein, human