Urinary arsenic profile affects the risk of urothelial carcinoma even at low arsenic exposure.
ABSTRACT Arsenic exposure is associated with an increased risk of urothelial carcinoma (UC). To explore the association between individual risk and urinary arsenic profile in subjects without evident exposure, 177 UC cases and 313 age-matched controls were recruited between September 2002 and May 2004 for a case-control study. Urinary arsenic species including the following three categories, inorganic arsenic (As(III)+As(V)), monomethylarsonic acid (MMA(V)) and dimethylarsinic acid (DMA(V)), were determined with high-performance liquid chromatography-linked hydride generator and atomic absorption spectrometry. Arsenic methylation profile was assessed by percentages of various arsenic species in the sum of the three categories measured. The primary methylation index (PMI) was defined as the ratio between MMA(V) and inorganic arsenic. Secondary methylation index (SMI) was determined as the ratio between DMA(V) and MMA(V). Smoking is associated with a significant risk of UC in a dose-dependent manner. After multivariate adjustment, UC cases had a significantly higher sum of all the urinary species measured, higher percent MMA(V), lower percent DMA(V), higher PMI and lower SMI values compared with controls. Smoking interacts with the urinary arsenic profile in modifying the UC risk. Differential carcinogenic effects of the urinary arsenic profile, however, were seen more prominently in non-smokers than in smokers, suggesting that smoking is not the only major environmental source of arsenic contamination since the UC risk differs in non-smokers. Subjects who have an unfavorable urinary arsenic profile have an increased UC risk even at low exposure levels.
- SourceAvailable from: Nathalie Saint-Jacques[Show abstract] [Hide abstract]
ABSTRACT: Arsenic in drinking water is a public health issue affecting hundreds of millions of people worldwide. This review summarizes 30 years of epidemiological studies on arsenic exposure in drinking water and the risk of bladder or kidney cancer, quantifying these risks using a meta-analytical framework.Environmental health : a global access science source. 06/2014; 13(1):44.
- [Show abstract] [Hide abstract]
ABSTRACT: Scientific debate surrounds the regulatory approach for evaluating carcinogenic risk of arsenic compounds. The arsenic ambient water quality criteria (AWQC), based on the assumption of a linear mode of action for skin cancer risk, results in an allowable limit of 0.018 ppb in ambient waters; the drinking water Maximum Contaminant Level (MCL) was determined using a similar linear approach. Integration of results from recent studies investigating arsenic’s mode of action provide the basis for a change in the approach for conducting an arsenic cancer risk assessment. Results provide support for a concentration demonstrating a dose-dependent transition in response from those representing adaptive changes to those that may be key events in the development of cancer endpoints. While additional information is needed, integration of current research results provides insight for a new quantitative cancer risk assessment methodology as an alternative toxicologically-based dose response (BBDR) cancer modeling. Integration of the new experimental results, combined with epidemiological evidence, support a dose-dependent transition concentration of approximately 0.1 μM arsenic. Some uncertainties remain; additional information from chronic in vitro studies underway is needed. Results to date also provide initial insight into variability in population response at low arsenic exposures.Regulatory Toxicology and Pharmacology 01/2014; · 2.13 Impact Factor
- [Show abstract] [Hide abstract]
ABSTRACT: Oral exposure to inorganic arsenic (iAs) is associated with adverse health effects. Epidemiological studies suggest differences in susceptibility to these health effects, possibly due to genotypic variation. Genetic polymorphisms in iAs metabolism could lead to increased susceptibility by altering urinary iAs metabolite concentrations. To examine the impact of genotypic polymorphisms on iAs metabolism. We screened 360 publications from PubMed and Web of Science for data on urinary mono- and dimethylated arsenic (MMA and DMA) percentages and polymorphic genes encoding proteins that are hypothesized to play roles in arsenic metabolism. The genes we examined were arsenic (+3) methyltransferase (AS3MT), glutathione-s-transferase omega (GSTO), and purine nucleoside phosphorylase (PNP). Relevant data were pooled to determine which polymorphisms are associated across studies with changes in urinary metabolite concentration. In our review, AS3MT polymorphisms rs3740390, rs11191439, and rs11191453 were associated with statistically significant changes in percent urinary MMA. Studies of GSTO polymorphisms did not indicate statistically significant associations with methylation, and there are insufficient data on PNP polymorphisms to evaluate their impact on metabolism. Collectively, these data support the hypothesis that AS3MT polymorphisms alter in vivo metabolite concentrations. Preliminary evidence suggests that AS3MT genetic polymorphisms may impact disease susceptibility. GSTO polymorphisms were not associated with iAs-associated health outcomes. Additional data are needed to evaluate the association between PNP polymorphisms and iAs-associated health outcomes. Delineation of these relationships may inform iAs mode(s) of action and the approach for evaluating low-dose health effects for iAs. Genotype impacts urinary iAs metabolite concentrations and may be a potential mechanism for iAs-related disease susceptibility.Environmental Research 04/2014; 132C:156-167. · 3.24 Impact Factor
This article was originally published in a journal published by
Elsevier, and the attached copy is provided by Elsevier for the
author’s benefit and for the benefit of the author’s institution, for
non-commercial research and educational use including without
limitation use in instruction at your institution, sending it to specific
colleagues that you know, and providing a copy to your institution’s
All other uses, reproduction and distribution, including without
limitation commercial reprints, selling or licensing copies or access,
or posting on open internet sites, your personal or institution’s
website or repository, are prohibited. For exceptions, permission
may be sought for such use through Elsevier’s permissions site at:
Author's personal copy
Urinary arsenic profile affects the risk of urothelial carcinoma
even at low arsenic exposure☆
Yeong-Shiau Pua, Shu-Mei Yangb, Yung-Kai Huangc, Chi-Jung Chungb, Steven K. Huangc,d,
Allen Wen-Hsiang Chiud,e, Mo-Hsiung Yangf, Chien-Jen Cheng, Yu-Mei Hsuehh,⁎
aDepartment of Urology, National Taiwan University College of Medicine, Taipei, Taiwan
bGraduate Institute of Public Health, Taipei Medical University, Taipei, Taiwan
cGraduate Institute of Medical Sciences, Taipei Medical University, Taipei, Taiwan
dDepartment of Urology, Chi-Mei Medical Center, Tainan, Taiwan
eDepartment of Urology, Taipei City Hospital, Taipei, Taiwan
fDepartment of Nuclear Science, National Tsing-Hua University, Hsinchu, Taiwan
gGraduate Institute of Epidemiology, College of Public Health, National Taiwan University, Taipei, Taiwan
hDepartment of Public Health, School of Medicine, Taipei Medical University, Taipei, No. 250 Wu-Hsing Street, Taipei 110, Taiwan
Received 13 July 2006; revised 22 September 2006; accepted 23 September 2006
Available online 10 November 2006
Arsenic exposure is associated with an increased risk of urothelial carcinoma (UC). To explore the association between individual risk and
urinary arsenic profile in subjects without evident exposure, 177 UC cases and 313 age-matched controls were recruited between September 2002
and May 2004 for a case–control study. Urinary arsenic species including the following three categories, inorganic arsenic (AsIII+AsV),
monomethylarsonic acid (MMAV) and dimethylarsinic acid (DMAV), were determined with high-performance liquid chromatography-linked
hydride generator and atomic absorption spectrometry. Arsenic methylation profile was assessed by percentages of various arsenic species in the
sum of the three categories measured. The primary methylation index (PMI) was defined as the ratio between MMAVand inorganic arsenic.
Secondary methylation index (SMI) was determined as the ratio between DMAVand MMAV. Smoking is associated with a significant risk of UC
in a dose-dependent manner. After multivariate adjustment, UC cases had a significantly higher sum of all the urinary species measured, higher
percent MMAV, lower percent DMAV, higher PMI and lower SMI values compared with controls. Smoking interacts with the urinary arsenic
profile in modifying the UC risk. Differential carcinogenic effects of the urinary arsenic profile, however, were seen more prominently in non-
smokers than in smokers, suggesting that smoking is not the only major environmental source of arsenic contamination since the UC risk differs in
non-smokers. Subjects who have an unfavorable urinary arsenic profile have an increased UC risk even at low exposure levels.
© 2006 Published by Elsevier Inc.
Keywords: Bladder neoplasms; Contamination; Carcinogenesis; Synergy; Tobacco
Urothelial carcinoma (UC) arises exclusively from the
urothelium including the renal pelvis, ureter, bladder and
urethra, with bladder cancer being the most common form. In
most developed countries, it is among the top 10 leading
cancers. The most well known risk factor for UC is cigarette
smoking (Brennan et al., 2000; Zeegers et al., 2000). Current
cigarette smokers have approximately a two- to three-fold risk
compared to non-smokers. The mechanism by which cigarette
smoking causes bladder cancer has yet to be determined. It
seems likely that the risk is related to a large number of
carcinogenic chemicals in cigarette smoke.
We have previously shown that chronic arsenic intoxication
due to contamination of artesian well water with inorganic
arsenic compounds is responsible for elevated mortality rates
Toxicology and Applied Pharmacology 218 (2007) 99–106
☆Grant support: NSC91-3112-B-038-0019, NSC92-3112-B-038-001,
NSC93-3112-B-038-001, NSC94-2314-B-038-023 and NSC95-2314-B-038-
007 from the National Science Council, Executive Yuan, ROC.
⁎Corresponding author. Fax: +886 2 27384831.
E-mail address: email@example.com (Y.-M. Hsueh).
0041-008X/$ - see front matter © 2006 Published by Elsevier Inc.
Author's personal copy
among individuals. We have previously shown that the
capability of arsenic methylation is related to the risks of skin
cancer (Hsueh et al., 1997; Chen et al., 2003a) and bladder
cancer in high arsenic-exposed subjects (Chen et al., 2003b).
Whether or not urinary arsenic profile affects UC risks among
subjects who had no evident arsenic exposure history is an
important issue. We hypothesized that, if an arsenic level of
50 μg/L in the water supply is still “unsafe”, the UC risk may
from cancers of the bladder, renal pelvis and/or ureter, lung and
other organs in Taiwan (Chen et al., 1985). More specifically,
we have demonstrated that bladder cancer mortality rates for
patients who consumed well water with arsenic levels of
600 μg/L or higher had a mortality rate of over 30 to 60 times
greater than the unexposed population (Chen et al., 1988).
Evidence from elsewhere in the world also suggests that
ingested inorganic arsenic very likely causes internal cancers
(Morales et al., 2000; Bates et al., 2004; Smith et al., 2006). It
has also been shown that combined cancer mortality rates are as
high as 1 out of 100 people from drinking water containing
50 μg/L of arsenic (Smith et al., 1992). Two reports from the
National Research Council (USA) have also affirmed that
cancer risks might be of the order of 1 in 100 at an arsenic level
of 50 μg/L (National Research Council, 1999, 2001). This
estimated cancer risk is more than 100 times greater than that for
any other contaminant in drinking water at the maximal
contaminant level (US EPA, 1998). Thus, within the U.S., the
maximum contaminant level for arsenic in public water supplies
will be lowered from 50 μg/L, a level that was established in
1942, to 10 μg/L in 2006 (Smith et al., 2002). The arsenic
concentration allowance in public water supplies in Taiwan was
50 μg/L; in 2000, a new standard of 10 μg/L was then
announced. According to the Taipei Water Department of Taipei
City Government, the average arsenic concentration of tap
water in Taipei is 0.7 μg/L ranging from non-detectable to
4.0 μg/L. Whether cancer risks are higher at 50 μg/L than at
10 μg/L is still debatable. Even without confounding factors,
reduction of cancer risks with the new standard of 10 μg/L will
not be seen for decades. However, should it be shown that
arsenic metabolic capability affects cancer risks in subjects
exposed 50 μg/L of arsenic, it might still be carcinogenic for
some genetically predisposed subjects.
Arsenic is usually found in drinking water in the form of
arsenate (AsV) or arsenite (AsIII) (Andreae, 1977). Inorganic
arsenic is bio-transformed in humans to monomethylarsonic
acid (MMAV) and dimethylarsinic acid (DMAV). Previously,
methylation of inorganic arsenic has always been considered a
detoxification mechanism because pentavalent MMA (MMAV)
and pentavalent DMA (DMAV) have relatively low toxicity
(Yamauchi and Fowler, 1994) and are rapidly excreted in the
urine (Vahter, 2002). However, recent studies have confirmed
the existence of trivalent intermediates and products of
monomethylarsonous acid (MMAIII) and dimethylarsinous
acid (DMAIII) that are more toxic than inorganic arsenite
more accurate assessment of arsenic metabolism, it is therefore
necessary to determine specific arsenic species in urine.
The capability of metabolizing inorganic arsenic differs
significantly differ between people with favorable and unfavor-
able arsenic methylation profile. We therefore conducted a
hospital-based case–control study to investigate the association
of urinary arsenic profile with the risk of UC. The possible
interactions between urinary arsenic profile and cigarette
smoking were also explored.
Material and methods
Study subjects and questionnaire interview.
seven patients with pathologically proven UC (age range 24 to 93 years)
were recruited from the Department of Urology, National Taiwan University
Hospital, between September 2002 and May 2004. Pathological verification of
UC was done by routine urological practice including endoscopic biopsy or
surgical resection of urinary tract tumors followed by histopathological
examination by board-certified pathologists. Cytological evidence alone was
not accepted as an adequate diagnosis of UC. A total of 313 age-matched
control subjects with no evidence of UC or any other malignancy were accrued
from a hospital-based pool, including those receiving senior citizen health
examinations at Taipei Medical University Hospital and those receiving adult
health examinations at Taipei Municipal Wan Fang Hospital. All three hospitals
are located in Taipei approximately 200 to 300 km away from the arsenic-
contaminated areas in Taiwan. No case subjects or controls came from arsenic-
contaminated areas in southwestern (Chen et al., 2003b) or northeastern (Chiou
et al., 2001) Taiwan.
Well-trained personnel carried out standardized personal interviews based
on a structured questionnaire. Information collected included demographic and
socioeconomic characteristics, general potential risk factors for malignancies
such as lifestyle, alcohol consumption, cigarette smoking in quantified details,
exposure to potential occupational and environmental carcinogens such as hair
dyes and pesticides, chronic medication history, consumption of conventional
and alternative medicines and personal and family history of urological
diseases. Frequent alcohol drinkers referred to those who consumed alcohol
three or more days per week, continuing for at least 6 months. Those who
consumed less than this level were classified as occasional drinkers. Pesticide
users were farmers who used pesticides for agricultural purposes. The Research
Ethics Committee of National Taiwan University Hospital, Taipei, Taiwan,
approved the study. All patients provided informed consent forms before
sample and data collection. The study was consistent with the World Medical
Association Declaration of Helsinki. Urine samples were stored at −20°C until
further use for urinary arsenic speciation. Bladder cancer was staged into three
groups: superficial (Ta, T1 and Tis), locally advanced (T2–4N0M0) and
metastatic (N+ or M+). Tumor grading was based on the WHO (1999)
classification system (WHO, 1999).
One hundred and seventy-
Determination of urinary arsenic species.
arsenic species is stable for at least 6 months when preserved at −20°C (Chen et
al., 2002); thus, the urine assay was performed within 6 months post-collection.
Frozen urine samples were thawed at room temperature, dispersed by ultrasonic
wave, filtered through a Sep-Pak C18column (Mallinckrodt Baker In., NJ) and
levels for AsIII, AsV, MMAVand DMAVwere determined. A urine aliquot of
200 μL was used for the determination of arsenic species by high-performance
liquid chromatography (HPLC) (Waters 501, Waters Associates, MA) with
columns obtained from Phenomenex (Nucleosil, Torrance, CA). The contents of
inorganic arsenic and their metabolites were quantified by hydride generator-
atomic absorption spectrometry (HG-AAS) (Hsueh et al., 1998). The
concentration of four arsenic species in standard solution, sample and sample
spiked standard solution was determined by using on-line HPLC-HG-AAS
respectively. Recovery rates of the four arsenic species were calculated by
[(sample spiked standard solution concentration)−sample concentration]/
standard solution concentration×100 respectively. Recovery rates for AsIII,
DMAV, MMAVand AsVranged between 93.8% and 102.2% with detection
limits of 0.02, 0.06, 0.07 and 0.10 μg/L, respectively. Urinary concentration of
the sum of inorganic arsenic, MMA and DMA was normalized against urinary
creatinine levels (μg/g creatinine). The standard reference material, SRM 2670,
contains 480±100 μg/L inorganic arsenic and was obtained from the National
It has been shown that urinary
100Y.-S. Pu et al. / Toxicology and Applied Pharmacology 218 (2007) 99–106
Author's personal copy
Keuls multiple comparison correction was applied to compare urinary arsenic
profiles between varied exposure strata. Multiple logistic regression models
were used to estimate the multivariate adjusted odds ratio (OR) and the 95%
confidence interval (CI). Cutoff points for continuous variables were the
respective tertiles 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. For the joint effect analysis, the cutoff
points for the percentages of arsenic species, PMI and SMI were the respective
medians of the controls. The joint effects of urinary arsenic species and cigarette
Institute of Standards and Technology (NIST, Gaithersburg, MD). SRM 2670
was used as a quality standard and analyzed along with urine samples. The mean
value of SRM 2670 determined by our system was 507±17 (SD) μg/L (n=4).
Arsenic methylation index was assessed by percentages of various urinary
arsenic species in the sum of inorganic arsenic, MMA and DMA. The primary
methylation index (PMI) was defined as the ratio between MMAVand inorganic
arsenic (AsIII+AsV) levels and the secondary methylation index (SMI) was
defined as the ratio between DMAVand MMAV(Tseng et al., 2005).
error. Student's t-test was used to compare differences in the urinary arsenic
profile between case subjects and controls. ANOVA and Student–Newman–
Continuous variables are expressed as mean±standard
smoking on the UC risk were evaluated on both multiplicative and additive
scales. The binary interaction terms were calculated by multiplying the
indicators for two explored risk factors and were added to the main effect
models. Their significance was then tested by the likelihood ratio statistic based
on a multiplicative model. The synergy index proposed by Rothman was
computed to assess the empirical deviation from the additive interaction
relationship (Rothman, 1986). The synergy index is equal to the calculation of
[OR(AB)−1]/[(OR(Ab)−1)+(OR(aB)−1)], where A and B denote the
presence of the two risk factors and a and b are designated as the absence of
the risk factors, respectively. An observed synergy index value that departs
substantially from the expected additive null, i.e., synergy index not equal to 1,
suggests an additive interaction effect. The OR values and their variance
covariance matrix were then used to calculate values for synergy index and 95%
CIs (Hosmer and Lemeshow, 1992). SAS version 8.2 was used for all statistical
The sociodemographic characteristics of cases and controls
are shown in Table 1. Subjects who had higher educational
levels had a lower risk than those with lower educational levels.
Mainland Chinese had a significantly lower UC risk than the
Fukien Taiwanese. Age, ABO blood type, marital status or use
of hair dye did not affect the UC risk. Occasional alcohol
drinkers had a significantly lower UC risk than non-drinkers
and frequent drinkers. Pesticide users had a significantly higher
UC risk than non-users.
Comparing cases and controls in terms of smoking duration,
daily smoking amount and cumulative smoking in pack-years,
Sociodemographic characteristics of 177 urothelial carcinoma cases and 313
age-matched non-cancer controls
Age and gender
Marital status 1.0
College or above
83 (26.7)73 (41.3)
aAge-adjusted odds ratio by logistic regression.
bGender-adjusted odds ratio by logistic regression.
Comparison of detailed smoking behavior between cases and controlsa
VariablesControls n (%)UC cases
Duration of cigarette smoking (years)
0 196 (65.1)
Amount of cigarette smoking (pack/day)
Cumulative cigarette smoking (pack-years)c
0 196 (65.1)
>22 56 (18.6)
aDetailed quantified smoking history was unavailable in twelve (3.8%)
controls and ten (5.6%) case subjects.
bMultivariate adjusted ORs: adjusted for age, gender, education, paternal and
maternal ethnicity, alcohol drinking and pesticide usage.
cCumulative smoking=(amount in pack/day)×(duration in years).
101Y.-S. Pu et al. / Toxicology and Applied Pharmacology 218 (2007) 99–106
Author's personal copy
UC cases and non-cancer controls
Controls 313 25.0±1.0 5.5±0.4
Cases 177 38.7±3.16.7±0.5
cigarette smokers had a significantly higher UC risk than non-
smokers in a dose-dependent manner (Table 2). Heavy smokers
who smoked for more than 33 years, 0.75 packs per day and 22
pack-years had a 2.4-, 2.2- and 2.5-fold risk, respectively,
compared to non-smokers by multivariate adjusted logistic
regression. Modest dose smokers, however, did not have a
significantly increased risk compared to non-smokers.
Table 3 compares the urinary arsenic profile between
varied exposure strata. Among non-cancer controls, male
subjects had a lower sum of inorganic arsenic, MMA and
DMA level, higher MMA percentage and lower DMA
percentage than females. Current smokers had a higher
MMA percentage and lower DMA percentage than non-
smokers. No significant differences were found in urinary
arsenic profiles between subjects consuming varied amount of
alcohol. Pesticide users had an insignificantly higher MMA
percentage than non-users.
UC cases had a significantly higher sum of inorganic arsenic,
MMA and DMA levels, higher MMA percentages, lower DMA
percentages and higher PMI levels than controls (Table 3).
Inorganic arsenic percentage was marginally higher in cases
than in controls (P=0.052). To examine if various cancer stages
affect urinary arsenic profile, we performed an analysis showed
that urinary arsenic profile did not differ between patients of
various tumor stages or grades in case subjects (Table 3). With
trend analysis on exposure strata in tertiles, all urinary arsenic
parameters, with the exception of inorganic arsenic percentage,
were found to be significantly associated with the UC risk after
the multivariate analysis (Table 4).
Since both smoking and urinary arsenic indices affect the UC
risk, further analyses were carried out to assess joint effects of
the two risk factors (Table 5). Trend analysis revealed
progressively increased risks through exposure to no risk factor,
either one of the factors, or both of the two risk factors.
Comparison of urinary arsenic species between varied exposure strata
Variablesn Sum of inorganic arsenic,
MMA and DMA (μg/g creatinine)
MMA percentage DMA percentage Primary
‡,§P<0.05 by Student–Newman–Keuls multiple comparisons.
¶Information of tumor staging and grading was not available in 16 and 9 UC patients, respectively.
102Y.-S. Pu et al. / Toxicology and Applied Pharmacology 218 (2007) 99–106