Significant variations in the frequency of single nucleotide polymorphisms (SNPs) of human leukocyte antigen among Taiwan's aboriginal tribes and high homogeneity within each tribe have been reported. To identify variations in genetic polymorphisms of drug-metabolizing enzyme in seven of Taiwan's ethnic groups, the SNPs of N-acetyl transferase 2, a conjugation enzyme in phase II metabolism, were investigated. Methods: Three SNPs of NAT2, NAT2*5, NAT2*6, and NAT2*7, were determined by the TaqMan® method and subsequently confirmed by polymerase chain reaction-restriction fragment length polymorphism (PCR-RFLP). Our results showed statistically significant variations in the frequency of NAT2 genotype categories according to the 3 SNPs of NAT2 within 7 Taiwanese ethnic groups (Chi-square = 32.08, p<0.05). The distribution of the frequency of the slow acetylator genotype ranged from 32.0% in the Hakka population to 75.5% in the Paiwan population. A lack of NAT2*5 SNP alleles was found in the Atayal population, while a NAT2*5 allele frequency of 31.6% was observed in the Paiwan group. The frequencies of allele NAT2*6 and NAT2*7 in the investigated ethnic groups ranged from 11.2-37.0% and 17.0-53%, respectively. Our results also showed the considerable variations in drug-metabolizing enzyme NAT2 genotypes among Taiwan's aboriginal and general population. The variation in the genetic polymorphisms of drug-metabolizing enzyme in Taiwan's groups is worthy of further study to understand different therapeutic and adverse drug responses in ethnic groups.

Estrogen is suggested to play a dual role in breast cancer development, both as a promoter, triggering cell proliferation, and as an initiator, causing the DNA damage essential for driving tumorigenesis. The initiator mechanism of estrogen is suggested to involve its metabolite, catechol estrogen, which has been shown to lead to oxidative DNA damage in vitro. To test this possibility in human breast cancer, a multigenic case-control study was conducted to determine whether the association between estrogen exposure and breast cancer risk might be modified by genotypic polymorphism of the genes involved in oxidative damage repair, including those involved in base excision repair (BER)(hOGG1, APE1, and XRCC1) and transcription-coupled repair (TCR)(XPD and BRCA1). One hundred and thirty-six breast cancer patients and 232 healthy controls were recruited and the single-nucleotide polymorphisms (SNPs) and genotypes of their oxidative damage repair genes determined by PCR-based RFLP assays. Women with low-risk genotypes of repair genes (hOGG1 codon326 Ser/Ser or Cys/Ser, APE1 codon148 Glu/Glu or Glu/Asp, XRCC1 codon399 Arg/Arg or Gln/Arg, XPD codon751 Lys/Lys, or BRCA1 codon871 Pro/Pro) showed no significant association between increased cancer risk and longer estrogen exposure (＞10 years) from menarche to first full-term pregnancy, whereas women with high-risk genotypes(hOGG1 codon326 Cys/Cys, APE1 codon148 Asp/Asp, XRCC1 codon399 Gln/Gln, XPD codon751 Gln/Lys or Gln/Gln, or BRCA1 codon871 Leu/Pro or Leu/Leu) showed a consistently stronger and significant association. Furthermore, a trend to an increased risk of developing breast cancer was found in women harboring higher numbers of high-risk genotypes of the BER and TCR genes (P=0.009), particularly in those with prolonged estrogen exposure. Interestingly, this study provided supporting evidence for an interaction, during breast tumorigenesis, between BER and TCR via BRCA1, by demonstrating both an extra independent risk (aOR, 2.1;95%CI, 1.1-4.0) of breast cancer in those women in whom both the BER and TCR pathways were defective and a more pronounced risk (aOR, 1.8; 95%CI, 1.1-3.1), associated with defective BER, in women with high-risk BRCA1 genotypes. On the basis of a comprehensive profile of DNA oxidative damage repair genes, this study has not only confirmed a carcinogenic mechanism by which estrogen causes DNA lesions, but has also shed light on a connection between TCR and BER in the prevention of oxidative damage and breast cancer development.

CYP3A4 is involved in the metabolism of numerous biologically active compounds, including testosterone. A genetic variant located in the P450NF (nifedipine) specific element (NFSE) has been identified that disrupts a transciptional regulatory element located in the 5' regulatory region of CYP3A4. The CYP3A4 variant (CYP3A4-V) is associated with the clinical presentation of prostate cancer. There are significant differences in CYP3A4 metabolism and rates of prostate cancer across ethnic groups that may be associated with CYP3A4 genotypes. Therefore, we estimated the frequency of the CYP3A4 variant in three ethnic groups with different prostate cancer incidence rates. The frequency (q) of CYP3A4-V was significantly different (p<0.0001) in African Americans (q=0.53), U.S. Caucasians (q=0.09), and Taiwanese (q=0.0). CYP3A4-V segregated in a Mendelian manner in one large African American family, and 7 of 16 (44%) biologically unrelated "marry-ins" carried a CYP3A4 variant allele. Reflecting population-specific prostate cancer incidence rates, our results suggest a high frequency of this variant in African Americans compared with U.S. Caucasians and Taiwanese.

The dapsone recovery ratio (DPRR), which is determined after single oral dose administration of dapsone by measuring the parent drug and hydroxylated metabolite, provides an in vivo measure of the efficiency of the drug metabolizing enzymes responsible for this metabolic route, putatively CYP3A4. This affords the potential to evaluate the hypothesis that this drug metabolizing enzyme system is involved in the pathogenesis of human bladder cancer. The present study is a matched case-control comparison of DPRR in patients with nonaggressive bladder cancer (grades I and II or Ta, T1 and T2, n = 43), patients with aggressive bladder cancer without invasion (grade III or Ta, T1 and T2, n = 32), patients with aggressive bladder cancer and invasion (grade III or T3 and T4, n = 32), and age- and gender-matched subjects with no urologic tumor on cystoscopy from an urban U.K. community (n = 85). Demographic variables associated with aggressive bladder cancer (Gill or T3, T4, Tis) included pack-years of smoking, alcohol intake, and occupational exposure; for nonaggressive bladder cancer variables included smoking and occupational exposure. DPRR exhibited an unimodal distribution in all subjects: activity was significantly reduced in both noninvasive and invasive aggressive bladder cancer, and was a significant risk factor for cancer after adjustment for other significant risk factors. Combining the two aggressive groups, the lowest tertile of DPRR activity was associated with a sixfold increase in risk (p < 0.02) compared with the upper tertile. We conclude that a low dapsone recovery ratio is an independent risk factor for aggressive bladder cancer irrespective of its stage of invasion and suggest that the enzymes involved in its metabolism are detoxifying enzymes for unknown environmental factors to which an urban community is exposed.

Purpose: To investigate the clonal origin of malignant cells in recurrent superficial bladder cancer.Materials and Methods: We compared the protein expression of p53 and retinoblastoma (Rb) by immunohistochemistry using antibody P1801 and PMG3-245, respectively, in 13 patients at the time of primary superficial bladder cancer resection (6 Ta and 7 Tl) and their 15 corresponding recurrences of disease, Mutations in p53 and Rb is ere inferred on the basis of immunoperoxidase staining.Results: At the time of initial tumor resection, a p53 mutation was observed in 5 patients (39%) and an Rb mutation was observed in 3 (23%). The p53/Rb mutation status of recurrent bladder cancers completely matched their corresponding primary bladder cancer.Conclusions: The chance that recurrent bladder cancer originated from independent clones in this study was extremely small (p < 10(-6)). This result strongly supports the monoclonal origin of recurrent superficial bladder cancer.

p53 and Rb gene mutations are intermediate biomarkers useful for the prediction of neoplastic progression in bladder cancers. Previously, we have shown that low CYP3A activity, measured by dapsone N-hydroxylation, and high CYP2D6 activity, assessed by debrisoquine 4-hydroxylation, were significant susceptibility risk factors in developing aggressive bladder cancer. However, no information is available about the relationship between drug/xenobiotic metabolizing enzyme activities and p53/Rb mutations that may suggest mechanisms of bladder carcinogenesis. We evaluated in vivo CYP3A activity by the dapsone recovery ratio (DPRR), CYP2D6 activity by the debrisoquine recovery ratio (DBRR), CYP2C19 activity by the mephenytoin R/S ratio (RSR), N-acetyltransferase activity by the monoacetyl dapsone to dapsone ratio and glutathione-S-transferase M1 (GSTM1) genotype by PCR. In immunohistochemical studies of bladder tumor tissue, over expression of p53 protein was detected with antibody pAb1801 and loss of Rb protein expression was evaluated with antibody PMG3-245 in patients with transitional cell carcinoma of the bladder. Low CYP3A activity was significantly associated with over expression of or mutated p53 protein (P < 0.05). High CYP2D6 activity (within the extensive metabolizer group) was significantly associated with loss of expression of or mutated Rb protein (P < 0.05). Positive p53 staining also predicted aggressive bladder cancer histopathology (P < 0.05, odds ratio 2.9), and the lowest tertile of DPRR predicted p53 positivity (P < 0.01, odds ratio 3.9 comparing means of lower tertile versus upper tertile of DPRR). These selective associations are consistent with the hypothesis that an environmental pro-carcinogen fails to be detoxified by CYP3A which may preferentially induce p53 mutations, whereas, an alternative pro-carcinogen that may be activated by CYP2D6, may selectively induce Rb mutations.

Bladder cancer provides the most definitive example for an association between environmental agents and cancer. However, in the absence of industrial occupational exposure, the primary carcinogen is rarely identified, and the mechanisms involved in cancer formation are poorly understood. The environmental procarcinogen hypothesis of tumour pathogenesis proposes that many carcinogens require metabolic activation by drug metabolizing enzymes to form the proximate carcinogen. A balance of exposure to the carcinogen, the activity of the enzymes involved in either formation of proximate carcinogen, or production of non-toxic metabolites, will determine tumour risk. We have used mephenytoin, debrisoquine and dapsone as selective probes for the phenotypic measures of activity of CYP2C19, CYP2D6, and CYP3A4, respectively. Within subject reproducibility of phenotypic measures, and the lack of cross-inhibition when the three drugs are given in a concurrent cocktail, have been confirmed. We have applied the cocktail drug approach in two, non-overlapping series of cases with bladder cancer and matched controls. In both series, patients with aggressive bladder cancer (GIII histopathology) had a history of excess alcohol intake, an under-representation of poor metabolizers of debrisoquine, a significant mean reduction in dapsone recovery ratio, but no difference in mephenytoin phenotype. Collectively, these observations involving multiple routes of drug metabolism support the procarcinogen environmental hypothesis for bladder cancer and suggest that measurement of activity of selected individual drug metabolizing enzymes involved in the pathogenesis of this tumour can be used to identify subjects at high risk of developing bladder cancer.

Debrisoquine hydroxylase activity has been attributed to CYP2D6 and poor metabolizers of debrisoquine have a reduced relative risk of developing aggressive bladder cancer. Production of a proximate carcinogen could occur in liver or bladder mucosa. However, it is not known if CYP2D6 is expressed in human bladder mucosa. In vivo whole body debrisoquine hydroxylase activity was measured as the debrisoquine recovery ratio (DBRR) following single dose oral administration of debrisoquine (10 mg) in 10 normal subjects and 20 patients with bladder cancer prior to diagnostic cystoscopy. Semi-quantitative PCR was used to measure mRNA for CYP2D6 in bladder tissue obtained at cystoscopy. Of the 30 subjects, three were phenotypically and genotypically poor metabolizers. Among the extensive metabolizers, there were extensive intersubject variations in DBRR. A 10-fold variation in CYP2D6 mRNA levels was observed in bladder tissue. There was a highly significant association between DBRR and CYP2D6 mRNA expression (r2 = 0.702, P < 0.001). These results demonstrate the presence of CYP2D6 mRNA in bladder mucosa. Furthermore, they are consistent with debrisoquine hydroxylation being mediated by CYP2D6 and suggest that differences in mRNA concentration are rate limiting for enzyme activity and that bladder mucosal regulation reflects total body regulation for this enzyme. The expression of CYP2D6 in bladder mucosa suggests that this enzyme could be involved in the local production of a proximate carcinogen in this tissue and contribute to the pathogenesis of bladder cancer in man.