Joint Effect of Arsenic Methylation Profile and 4-(methylnitrosamino)-1-(3-pyridyl)-1-butanone (NNK) Metabolites on Urothelial Carcinoma
Chia-Chang Wu, MD,1,2 Chien-Tien Su, MD,3 Hui-Ling Lee, PhD,4 Chi-Jung Chung, PhD,5,6
Chao-Yuan Huang, MD, PhD,7,8 Yeong-Shiau Pu, MD, PhD,8 Pinpin Lin, PhD,9 Yu-Mei Hsueh, PhD,10,1 1 School of Public Health, College of Public Health and Nutrition, Taipei Medical University,
Taipei, Taiwan.
2 Department of Urology, Taipei Medical Universtiy-Shuang Ho Hospital, Taipei, Taiwan. 3 Department of Family Medicine, Taipei Medical University Hospital, Taipei, Taiwan. 4 Department of Chemistry, Fu Jen Catholic University, Hsinchuang, Taipei County, Taiwan. 5Department of Health Risk Management, College of Public Health; China Medical University and
Hospital, Taichung, Taiwan.
6 Department of Medical Research, China Medical University Hospital, Taichung, Taiwan.
7 Graduate Institute of Clinical Medicine, College of Medicine, Taipei Medical University, Taipei,
Taiwan.
8Department of Urology, National Taiwan University Hospital, College of Medicine National
Taiwan University, Taipei, Taiwan.
9Division of Environmental Health and Occupational Medicine, National Health Research
Institutes, Miaoli County, Taiwan.
10Department of Public Health, School of Medicine, College of Medicine, Taipei Medical
University, Taipei, Taiwan.
Address correspondence to Yu-Mei Hsueh, PhD,
Department of Public Health, School of Medicine, College of Medicine, Taipei Medical University, No. 250 Wu-Hsing Street, Taipei 110, Taiwan.
E-mail: ymhsueh@tmu.edu.tw. TEL: 886-2-27361661 ext. 6513 FAX: 886-2-27384831
Running title: Arsenic, NNK and urothelial carcinoma
Key words: Arsenic, Arsenic Methylation Profile, 4-(methylnitrosamino)-1-(3-pyridyl)-1-butanone, Urothelial Carcinoma
Abstract
Purpose: Cigarette smoking interacts with the urinary arsenic profile to modify the urothelial carcinoma (UC) risk. (methylnitrosamino)-1-(3-pyridyl)-1-butanone (NNK) and its metabolite 4-(methylnitrosamino)-1-(3-pyridyl)-1-butanol (NNAL) are biomarkers for cigarette smoking
exposure. We explored the joint effects of urinary NNK metabolites and arsenic species on UC risk. Materials and Methods: We recruited 137 pairs of UC cases and matched healthy participants for a hospital-based case-control study. Participants completed questionnaires and provided 50 mL urine samples. Urine samples were analyzed for free NNAL and NNAL-glucuronides (NNAL-Gluc) using liquid chromatography-tandem mass spectrometry; samples were analyzed for arsenic species using high performance liquid chromatography-hydride generator-atomic absorption spectrometry. Results: Overall, subjects with high urinary total NNAL and high total arsenic had a higher UC risk than those with low total NNAL and low total arsenic. Subjects with a lower ratio of NNAL-Gluc/free NNAL and higher total arsenic had a higher UC risk than those with a higher NNAL-Gluc/free NNAL ratio and lower total arsenic.
Conclusions: This is the first study to observe a significant trend of progressively increased risk of UC in subjects who had none, one, or both of the following factors: urinary total arsenic and total NNAL or urinary total arsenic and the ratio of NNAL-Gluc/free NNAL.
Introduction
Urothelial carcinoma (UC) arises from the urothelium, a tissue lining the inner surface of hollow organs, and includes cancers of the renal pelvis, ureter and bladder. Bladder cancer is the second most common malignancy of the genitourinary tract and it is the eighth most common malignancy among men in Taiwan. Cigarette smoking is a major risk factor for bladder cancer and accounts for up to 50% of all incident cases of bladder cancer 1. Our previous study indicated that a
increased exposure to cigarette smoking, measured by daily exposure and cumulative pack-years, caused cigarette smokers to have a significantly higher odds ratio (OR) of UC than non-smokers in a dose-dependent manner 2. This finding is consistent with the results reported in a meta-analysis
involving 43 case-control and cohort studies 3. Cigarette smoke contains more than 60 carcinogenic
constituents that can induce tumorgenesis or increase proliferation of urothelial cells 4.
Environmental tobacco smoke (ETS) is thought to be a risk factor for bladder cancer 5. The
nicotine-derived tobacco-specific nitrosamine, 4-(methylnitrosamino)-1-(3-pyridyl)-1-butanone (NNK), has also been linked to tobacco-related human malignancies 6. NNK is a pro-carcinogen
that requires metabolic activation by cytochrome P450 enzymes to exhibit its carcinogenic effects 7.
NNK is metabolized by carbonyl reduction to 4-(methylnitrosamino)-1-(3-pyridyl)-1-butanol (NNAL). NNAL is a carcinogen and can be detoxified to either O- or N-glucuronide, known as NNAL-glucuronides (NNAL-Gluc); this detoxification is mediated by
UDP-glucuronosyltransferases (UGTs) 8. Total NNAL (free NNAL and its glucuronides) is only found in
tobacco-related products and is one of the most important carcinogenic biomarkers in urine for individuals with ETS exposure 9. In addition, the ratio of NNAL-Gluc/free NNAL is a useful
biomarker for NNK detoxification in smokers and a lower ratio is regarded as a risk factor for UC
10. Both NNK and NNAL can be activated by α-hydroxylation to form DNA adducts such as
7-methylguanine (N7MeG) 11. In bladder cancer, N7MeG levels are higher in bladder cancer tissue
than in adjacent normal bladder epithelia 12. Our previous study found that the urinary levels of
total NNAL, free NNAL and N7MeG were positively correlated with cigarette smoking 5.
associated with chronic arsenic exposure 13. Inorganic arsenic is bio-transformed in humans to
monomethylarsonic acid (MMAV) and dimethylarsinic acid (DMAV). The methylation of inorganic
arsenic has always been considered a detoxification process because MMAV and DMAV have
relatively low toxicity 14 and are quickly excreted in urine 15. However, recent studies have shown
that monomethylarsonous acid (MMAIII) and dimethylarsinous acid (DMAIII) are more toxic than
inorganic arsenite 16. In addition, the metabolizing ability of inorganic arsenic varies among
individuals. We previously reported that decreased arsenic methylation capability is associated with skin cancer and UC 2,17. Further, the synergism between cigarette smoking and arsenic exposure has
been reported in previous studies 2,18,19. Because arsenic is a constituent of tobacco smoke 20, ETS
may be another potential source of arsenic exposure. A study reported the relationship between the interaction of ETS and arsenic methylation capability on bladder cancer risk 18, but the joint effects
of tobacco metabolites and the arsenic methylation profile on UC remain unknown. Therefore, we conducted a case-control study to investigate the joint effect of the tobacco-specific nitrosamine NNK metabolites and urinary arsenic methylation profile on UC.
Materials and methods Study participants
A detailed description of the study design has been described previously 2. Briefly, this was a
hospital-based case-control study involving 137 UC cases that were recruited from the Department of Urology at the National Taiwan University Hospital between September 2007 and May 2009. UC cases were confirmed with histological and pathological examinations. The examinations were performed using routine urological practices, including endoscopic biopsy or surgical resection of urinary tract tumors, followed by histopathological examination by board-certified pathologists. A total of 137 control participants were recruited from those receiving health examinations at Taipei Medical University Hospital and Taipei Municipal Wan Fang Hospital. These cancer-free controls were matched to UC cases in terms of age ± 3 years and gender. The majority of the study participants ( > 80%) came from Taipei city and drank tap water. All participants provided
informed consent before the questionnaire interview and biospecimen collection. The Research Ethics Committee of the National Taiwan University Hospital approved this study, and the study complied with the World Medical Association Declaration of Helsinki.
Questionnaire interview and biospecimen collection
Well-trained interviewers collected information from participants through a standardized personalized interview based on a structured questionnaire. Information was collected in numerous areas: demographic and socioeconomic characteristics; lifestyle factors such as cigarette smoking, ETS exposure, and alcohol, tea and coffee consumption; and personal and family histories of disease. A 50 mL sample of spot urine from each participant was collected at the time of recruitment and immediately transferred to a -20°C freezer for the evaluation of urinary arsenic species and tobacco carcinogen NNK metabolites within 6 months.
Determination of urinary arsenic species
The detailed laboratory procedures have been described previously 21. Briefly, a urine aliquot
of 200 μL was used for determining the arsenic species by high-performance liquid chromatography (HPLC) (Waters 501, Waters Associates, MA). Inorganic arsenic and its metabolites were quantified by hydride generator atomic absorption spectrometry (HG-AAS) 21. The concentrations
of four arsenic species of standard solution, sample and sample-spiked standard solution were determined by on-line HPLC-HG-AAS. Recovery rates for AsIII, DMAV, MMAV and AsV ranged
between 93.8% and 102.2%, with detection limits of 0.02, 0.06, 0.07 and 0.10 μg/L, respectively. The urinary concentration of the sum of inorganic arsenic (AsIII + AsV), MMAV and DMAV, named
“total arsenic”, was normalized against urinary creatinine levels (μg/g creatinine). The arsenic methylation index was calculated as the percentage of various urinary arsenic species as a portion of the total arsenic.
Analysis of NNK metabolites
Urinary NNK metabolites were evaluated using the liquid chromatography-tandem mass spectrometry method 5. Linearity was observed for NNAL and hydroxy acid (HA) compounds (R2 =
injected samples. Precision and accuracy were determined by analyzing urine collected from a non-smoker spiked with 1 ng/mL d3-NNAL (QC samples). The average intra-day and inter-day
variations were 6.2% and 5.5% (n = 5), respectively. Accuracy ranged from 88.0% to 107.1% (n = 5). QC samples and a blank were checked every 10 samples and the calibration standard solution was tested every 20 samples. Our data conformed to the acceptance criteria, showing good reproducibility for evaluating each urine sample.
Statistical analysis
Continuous variables were expressed as mean ± standard error (SE). Student's t-test was used to compare the differences in the continuous variables between UC cases and controls. Total NNAL concentration (μg/g creatinine) was the sum of urinary free NNAL and NNAL-Gluc. We performed a multivariate logistic regression to estimate the joint effect of the total NNAL (μg/g creatinine) or NNAL-Gluc/free NNAL ratio and the total arsenic (μg/g creatinine) on UC. For the dose-response analysis, the cutoff points of arsenic indices or NNK metabolites were the respective tertiles or quartiles of the controls. Significance tests for linear trend among ORs across exposure strata were calculated by categorizing exposure variables and treating scored variables as continuous. In addition, for the joint effect analysis, total arsenic (μg/g creatinine), total NNAL (μg/g creatinine) and the NNAL-Gluc/free NNAL ratio were the medians of the controls. All data were analyzed using the Statistical Analysis Software for Windows, version 9.1 (SAS Institute, Cary, NC). P-values of < 0.05 were considered statistically significant.
Results
Participants who had a higher educational level demonstrated a lower OR of UC in a dose-dependent manner. Ever-smokers had a higher OR of UC than non-smokers, but this association was statistically insignificant. Higher cigarette smoking indices, in terms of daily cigarette smoking amount and cumulative cigarette smoking in pack-years, were associated with a significantly higher OR of UC than in non-smokers in a dose-dependent manner after adjustment for age and gender.
The OR of UC was markedly higher (2-fold) in subjects with ETS exposure than in those without exposure (95% CI, 1.26 - 3.58). A significant trend of progressively increased OR of UC was observed in subjects who met none, one, or both of the following criteria: cigarette smoker and ETS exposure.
UC cases had significantly higher levels of total NNAL (μg/g creatinine) and free NNAL (μg/g creatinine). However, the NNAL-Gluc/ free NNAL ratio was significantly lower in UC cases than in controls. UC cases had significantly higher total arsenic ( g/g creatinine), a higher
percentage of inorganic arsenic, a higher percentage of MMA , and V a lower percentage of
DMAV than controls. Based on trend analysis, urinary total arsenic, inorganic
arsenic percentage, and free NNAL were found to be significantly associated with UC. However, DMAV percentage and NNAL-Gluc/free NNAL were
significantly inversely associated with UC in a dose-dependent manner after the multivariate analysis.
Total NNAL was significantly positively correlated with cumulative cigarette smoking, total arsenic, and inorganic arsenic percentage, but total NNAL was significantly inversely correlated with DMAV percentage (Table 1).
Since total NNAL, NNAL-Gluc/free NNAL and urinary arsenic indices affect UC, further analyses were conducted to assess the joint effects of these risk factors. Although urinary total NNAL or NNAL-Gluc/free NNAL tended to interact additively with urinary total arsenic in modifying the OR of UC, the interactions were all statistically insignificant. A significant trend of progressively increased OR was observed in subjects who had none, one, or both of the two unfavorable indices (Table 2). The joint effect of total NNAL, NNAL-Gluc/free NNAL and the arsenic methylation profile indices (inorganic arsenic percentage, MMAV percentage, and DMAV percentage) had similar associations
with the OR of UC as urinary total arsenic (data not shown). Discussion
In this study, we observed that the higher the amount of daily cigarette smoking and cumulative cigarette smoking, the higher the OR of UC. Subjects with high total NNAL and high total arsenic had a higher OR of UC than those with low total NNAL and low total arsenic. Subjects with a lower ratio of NNAL-Gluc/free NNAL and higher total arsenic had a higher OR of UC than those with a higher NNAL-Gluc/free NNAL ratio and lower total arsenic. Based on the present data, NNK-related metabolites might interact with urinary total arsenic and affect UC.
A previous study reported a significant dose-response relationship between arsenic concentration in drinking water and UC in Taiwan 13. The results would have been more significant if the
percentages of urinary arsenic species were used as internal indicators of inorganic arsenic metabolism. Consistent with previous findings 2, we observed that UC cases had higher total
arsenic, inorganic arsenic and MMAV percentages and a lower DMAV percentage in the present
study. Generally, arsenic methylation is thought to be a detoxification process in which MMAV and
DMAV are considered non-toxic metabolites. In fact, inhibition of the secondary methylation
process, resulting in higher MMAV and lower DMAV percentages in urine, have weaker
detoxification capabilities that may lead to a higher risk of arsenic-related malignancies 2.
Recently, we also found that the levels of three urinary NNK metabolites, total NNAL, free NNAL and NNAL-Gluc, increased gradually with cumulative dosage of cigarette smoking 22. In
addition, cigarette smokers with a lower NNAL-Gluc/free NNAL ratio had a significantly higher OR of UC than non-smokers with a higher NNAL-Gluc/free NNAL ratio. Therefore, the ratio of NNAL-Gluc/free NNAL might be a useful biomarker to represent cigarette smoking exposure and to rule out potential error from a questionnaire interview 22. A previous study reported that the
half-life of total NNAL was 10 to 18 days, and defined urinary total NNAL as the specific biomarker to detect tobacco smoke exposure or second-hand smoke 23. In addition, female non-smokers who
were exposed to ETS and poorly metabolized the tobacco-specific carcinogen NNK demonstrated an increased lifetime risk of lung cancer 24. Although the carcinogenicity of NNK and its
metabolites in lung cancer have been reported, few studies have explored the carcinogenicity of these chemicals related to an increased bladder cancer risk.
UC is a multi-factorial malignancy in the urinary tract system. Environmental risk factors, including arsenic exposure through drinking well water, cigarette smoking and ETS, are considered major risk factors for UC 25. Further, exposure to certain carcinogens contained in tobacco smoke
may lead to DNA damage through the generation of DNA adducts 26. The formation of reactive
oxygen species, which can induce DNA adducts, also occurs in the process of inorganic arsenic metabolism 27. Therefore, the joint effect of arsenic exposure and cigarette smoking in UC may be
due to shared mechanisms of carcinogenesis.
To our knowledge, this is the first study to investigate the joint effects of urinary arsenic species and NNK-related metabolites, including total NNAL and the ratio of NNAL-Gluc/free NNAL, on UC. In a recent study, we did not find a significant interaction between urinary total arsenic and total NNAL on UC; however, we observed that those who demonstrated high urinary total NNAL and high total arsenic simultaneously had a significantly greater OR of UC than individuals with lower urinary total NNAL and a lower total arsenic level. On the other hand, we found that total NNAL was significantly positively associated with total arsenic and inorganic arsenic percentage and inversely associated with DMA percentage. These findings indicated that cigarette smoking might alter arsenic methylation, leading to an amplification of arsenic carcinogenesis28. In contrast,
arsenic may also influence the metabolism of cigarette smoke components, such as bezopyrene or NNK, and promote cigarette-smoking related carcinogenesis29. In addition, we found that
individuals with a low NNAL-Gluc/free NNAL ratio and high total arsenic simultaneously had a significantly greater OR of UC compared to individuals with a high NNAL-Gluc/free NNAL ratio and low total arsenic. The NNAL-Gluc/free NNAL ratio was a better biomarker than total NNAL to evaluate NNK-related metabolites that can contribute to the carcinogenesis of UC. Therefore, it may suggest a mechanism by which the presence of arsenic and NNK metabolites could induce greater DNA adduct formation in hepatic tissues, resulting in a higher carcinogenetic potential30.
The present study has some limitations. First, the evaluation of a single spot of urinary arsenic species and urinary tobacco carcinogen NNK metabolites may be limited; however, the values were considered reliable if subjects did not have a recent change in their lifestyle. Second, U C
prevalence cases were recruited in this study. We cannot exclude the possibility that the association between urinary NNAL or arsenic levels and U C in the present study might have resulted from and not be the cause of U C. Finally, owing to a small sample size, statistical significance should be interpreted with caution.
In conclusion, to our knowledge, this is the first study to investigate the joint effect of the metabolic products of NNK and arsenic species in urine on UC. The findings are especially consequential for individuals with a lower NNAL-Gluc/free NNAL ratio and high total arsenic. Moreover, the ratio of NNAL-Gluc/free NNAL and total arsenic can be used as a biomarker for estimating the OR of UC.
Conflict of interest statement
The authors declare that there are no conflicts of interest.
Acknowledgment
This study was supported by grants from the National Science Council of the ROC (NSC 86-2314-B-038-038, NSC 87-2314-B-038-029, NSC-88-2314-B-038-112, N2314-B038-049, SC-89-2320-B038-013, NSC-90-2320-B-038-021, NSC-91-3112-B-038-0019, NSC-92-3112-B-038-001, NSC-93-3112-B-038-001, NSC-94-2314-B-038-023, NSC-95-2314-B-038-007, NSC-96-2314-B038-003, 2314-B-038-015-MY3 (1-3), 2314-B-038-015-MY3 (2-3), NSC-97-2314-B-038-015-MY3 (3-3)), and NSC 100-2314-B-038 -026. References
1. Strope, S. A. and Montie, J. E.: The causal role of cigarette smoking in bladder cancer
initiation and progression, and the role of urologists in smoking cessation. J Urol, 180: 31, 2008.
2. Pu, Y. S., Yang, S. M., Huang, Y. K., Chung, C. J., Huang, S. K., Chiu, A. W. et al.: Urinary arsenic profile affects the risk of urothelial carcinoma even at low arsenic exposure. Toxicol Appl Pharmacol, 218: 99, 2007.
3. Zeegers, M. P., Tan, F. E., Dorant, E., and van Den Brandt, P. A.: The impact of
characteristics of cigarette smoking on urinary tract cancer risk: a meta-analysis of epidemiologic studies. Cancer, 89: 630, 2000.
4. Johansson, S. L. and Cohen, S. M.: Epidemiology and etiology of bladder cancer. Semin Surg Oncol, 13: 291, 1997.
5. Lee, H. L., Hsueh, Y. M., Chung, C. J., Pu, Y. S., Chang, L. W., Hsieh, D. P. et al.: Correlation between the urine profile of 4-(methylnitrosamino)-1-(3-pyridyl)-1-butanone metabolites and N7-methylguanine in urothelial carcinoma patients. Cancer Epidemiol Biomarkers Prev, 17: 3390, 2008.
6. Hecht, S. S.: Biochemistry, biology, and carcinogenicity of tobacco-specific N-nitrosamines. Chem Res Toxicol, 11: 559, 1998.
7. Richter, E., Engl, J., Friesenegger, S., and Tricker, A. R.: Biotransformation of
4-(methylnitrosamino)-1-(3-pyridyl)-1-butanone in lung tissue from mouse, rat, hamster, and man. Chem Res Toxicol, 22: 1008, 2009.
8. Muscat, J. E., Djordjevic, M. V., Colosimo, S., Stellman, S. D., and Richie, J. P., Jr.: Racial differences in exposure and glucuronidation of the tobacco-specific carcinogen 4-(methylnitrosamino)-1-(3-pyridyl)-1-butanone (NNK). Cancer, 103: 1420, 2005. 9. Tulunay, O. E., Hecht, S. S., Carmella, S. G., Zhang, Y., Lemmonds, C., Murphy, S. et al.:
Urinary metabolites of a tobacco-specific lung carcinogen in nonsmoking hospitality workers. Cancer Epidemiol Biomarkers Prev, 14: 1283, 2005.
10. Chung, C. J., Lee, H. L., Yang, H. Y., Lin, P., Pu, Y. S., Shiue, H. S. et al.: Low ratio of 4-(methylnitrosamino)-1-(3-pyridyl)-1-butanol-glucuronides (NNAL-Gluc)/free NNAL increases urothelial carcinoma risk. Sci Total Environ, 409: 1638, 2011.
11. Hecht, S. S.: Tobacco smoke carcinogens and lung cancer. J Natl Cancer Inst, 91: 1194, 1999. 12. Saad, A. A., O'Connor, P. J., Mostafa, M. H., Metwalli, N. E., Cooper, D. P., Margison, G. P. et al.: Bladder tumor contains higher N7-methylguanine levels in DNA than adjacent normal bladder epithelium. Cancer Epidemiol Biomarkers Prev, 15: 740, 2006.
13. Chiou, H. Y., Chiou, S. T., Hsu, Y. H., Chou, Y. L., Tseng, C. H., Wei, M. L. et al.: Incidence of transitional cell carcinoma and arsenic in drinking water: a follow-up study of 8,102 residents in an arseniasis-endemic area in northeastern Taiwan. Am J Epidemiol, 153: 411, 2001.
14. Yamauchi, H. and Fowler, B. A. Toxicity and metabolism of inorganic and methylated
arsenicals. Nriagu, J. O. 33-43. 1994. New York, John Wiley and Sons. Arsenic in the Environment, part 2:Human Health and Ecosystem Effects. Ref Type: Serial (Book,Monograph)
15. Vahter, M.: Mechanisms of arsenic biotransformation. Toxicology, 181-182: 211, 2002. 16. Styblo, M., Drobna, Z., Jaspers, I., Lin, S., and Thomas, D. J.: The role of biomethylation in
toxicity and carcinogenicity of arsenic: a research update. Environ Health Perspect, 110 Suppl 5: 767, 2002.
17. Hsueh, Y. M., Chiou, H. Y., Huang, Y. L., Wu, W. L., Huang, C. C., Yang, M. H. et al.: Serum beta-carotene level, arsenic methylation capability, and incidence of skin cancer. Cancer Epidemiol Biomarkers Prev, 6: 589, 1997.
18. Chen, Y. C., Su, H. J., Guo, Y. L., Houseman, E. A., and Christiani, D. C.: Interaction between environmental tobacco smoke and arsenic methylation ability on the risk of bladder cancer. Cancer Causes Control, 16: 75, 2005.
19. Melkonian, S., Argos, M., Pierce, B. L., Chen, Y., Islam, T., Ahmed, A. et al.: A prospective study of the synergistic effects of arsenic exposure and smoking, sun exposure, fertilizer use, and pesticide use on risk of premalignant skin lesions in Bangladeshi men. Am J Epidemiol, 173: 183, 2011.
20. Landsberger, S. and Wu, D.: The impact of heavy metals from environmental tobacco smoke on indoor air quality as determined by Compton suppression neutron activation analysis. Sci Total Environ, 173-174: 323, 1995.
21. Hsueh, Y. M., Huang, Y. L., Huang, C. C., Wu, W. L., Chen, H. M., Yang, M. H. et al.: Urinary levels of inorganic and organic arsenic metabolites among residents in an arseniasis-hyperendemic area in Taiwan. J Toxicol Environ Health A, 54: 431, 1998. 22. Chung, C. J., Lee, H. L., Yang, H. Y., Lin, P., Pu, Y. S., Shiue, H. S. et al.: Low ratio of
4-(methylnitrosamino)-1-(3-pyridyl)-1-butanol-glucuronides (NNAL-Gluc)/free NNAL increases urothelial carcinoma risk. Sci Total Environ, 2011.
23. Goniewicz, M. L., Havel, C. M., Peng, M. W., Jacob, P., III, Dempsey, D., Yu, L. et al.: Elimination kinetics of the tobacco-specific biomarker and lung carcinogen
4-(methylnitrosamino)-1-(3-pyridyl)-1-butanol. Cancer Epidemiol Biomarkers Prev, 18: 3421, 2009.
24. Anderson, K. E., Carmella, S. G., Ye, M., Bliss, R. L., Le, C., Murphy, L. et al.: Metabolites of a tobacco-specific lung carcinogen in nonsmoking women exposed to
environmental tobacco smoke. J Natl Cancer Inst, 93: 378, 2001.
25. Pelucchi, C., Bosetti, C., Negri, E., Malvezzi, M., and La, V. C.: Mechanisms of disease: The epidemiology of bladder cancer. Nat Clin Pract Urol, 3: 327, 2006.
26. Vineis, P., Talaska, G., Malaveille, C., Bartsch, H., Martone, T., Sithisarankul, P. et al.: DNA adducts in urothelial cells: relationship with biomarkers of exposure to arylamines and polycyclic aromatic hydrocarbons from tobacco smoke. Int J Cancer, 65: 314, 1996. 27. Yamanaka, K., Kato, K., Mizoi, M., An, Y., Nakanao, M., Hoshino, M. et al.: Dimethylarsine
likely acts as a mouse-pulmonary tumor initiator via the production of dimethylarsine radical and/or its peroxy radical. Life Sci, 84: 627, 2009.
28. Mead, M. N.: Arsenic: in search of an antidote to a global poison. Environ Health Perspect, 113: A378, 2005.
29. Fischer, J. M., Robbins, S. B., Al Zoughool, M., Kannamkumarath, S. S., Stringer, S. L., Larson, J. S. et al.: Co-mutagenic activity of arsenic and benzo[a]pyrene in mouse skin. Mutat Res, 588: 35, 2005.
30. Lee, H. L., Chang, L. W., Wu, J. P., Ueng, Y. F., Tsai, M. H., Hsieh, D. P. et al.:
Enhancements of 4-(methylnitrosamino)-1-(3-pyridyl)-1-butanone (NNK) metabolism and carcinogenic risk via NNK/arsenic interaction. Toxicol Appl Pharmacol, 227: 108, 2008.
