IJC

International Journal of Cancer

Use of acetochlor and cancer incidence in the Agricultural

Health Study
Catherine C. Lerro1,2, Stella Koutros1, Gabriella Andreotti1, Cynthia J. Hines3, Aaron Blair1, Jay Lubin4, Xiaomei Ma5,
Yawei Zhang2 and Laura E. Beane Freeman1
1
Occupational and Environmental Epidemiology Branch, Division of Cancer Epidemiology and Genetics, National Cancer Institute, Bethesda, MD
2
Environmental Health Sciences Department, Yale University School of Public Health, New Haven, CT
3
Division of Surveillance, Hazard Evaluations and Field Studies, National Institute for Occupational Safety and Heath, Cincinnati, OH
4
Biostatistics Branch, Division of Cancer Epidemiology and Genetics, National Cancer Institute, Bethesda, MD
5
Chronic Disease Epidemiology Department, Yale University School of Public Health, New Haven, CT

Since its registration in 1994 acetochlor has become a commonly used herbicide in the US, yet no epidemiologic study has
evaluated its carcinogenicity in humans. We evaluated the use of acetochlor and cancer incidence among licensed pesticide
applicators in the Agricultural Health Study. In telephone interviews administered during 1999–2005, participants provided
information on acetochlor use, use of other pesticides and additional potential confounders. We used Poisson regression to
estimate relative risks (RR) and 95% confidence intervals (95% CI) for cancers that occurred from the time of interview
through 2011 in Iowa and 2010 in North Carolina. Among 33,484 men, there were 4,026 applicators who used acetochlor and
3,234 incident cancers, with 304 acetochlor-exposed cases. Increased risk of lung cancer was observed among acetochlor
users (RR 5 1.74; 95% CI: 1.07–2.84) compared to nonusers, and among individuals who reported using acetochlor/atrazine
product mixtures (RR 5 2.33; 95% CI: 1.30–4.17), compared to nonusers of acetochlor. Colorectal cancer risk was significantly
elevated among the highest category of acetochlor users (RR 5 1.75; 95% CI: 1.08–2.83) compared to never users. Addition-
ally, borderline significantly increased risk of melanoma (RR 5 1.61; 95% CI: 0.98–2.66) and pancreatic cancer (RR 5 2.36;
95% CI: 0.98–5.65) were observed among acetochlor users. The associations between acetochlor use and lung cancer, colo-
rectal cancer, melanoma and pancreatic cancer are suggestive, however the lack of exposure-response trends, small number
of exposed cases and relatively short time between acetochlor use and cancer development prohibit definitive conclusions.

Acetochlor is a broad leaf herbicide and a member of the chlor-
acetanilide class,1 which also includes alachlor, butachlor, prop-
achlor and metolachlor.1,2 In 1994, the US Environmental
Key words: acetochlor, Agricultural Health Study, pesticides, cancer
Protection Agency (EPA) approved acetochlor for use on corn,
with continued registration dependent upon a reduction in the
use of several other corn herbicides of known health concern,
including alachlor, metolachlor, atrazine, 2,4-dichlorophenoxy-
acetic acid (2,4-D), butylate and S-ethyl dipropylthiocarbamate
(EPTC).3 Re-registration was not contingent upon reduction of

Abbreviations: AHS: Agricultural Health Study; BMI: body mass index;
CYP450: cytochrome P450; 2,4-D: 2,4-dichlorophenoxyacetic acid;
DNA: deoxyribonucleic acid; EPA: Environmental Protection Agency;
EPTC: S-ethyl dipropylthiocarbamate; NHL: non-Hodgkin lymphoma
Published 2015. This article is a US Government work and, as such, is
in the public domain of the United States of America.
Grant sponsor: Intramural Research Program of the National
Institutes of Health; Grant sponsor: National Cancer Institute;
Grant number: Z01-CP010119; Grant sponsor: National Institute of
Environmental Health Sciences; Grant number: Z01-ES049030
DOI: 10.1002/ijc.29416
History: Received 5 Sep 2014; Accepted 17 Dec 2014; Online 5 Jan 2015
Correspondence to: Catherine Lerro, Occupational and
Environmental Epidemiology Branch, National Cancer Institute,
9609 Medical Center Drive, 6E620, MSC 7991, Bethesda, MD
20892-7991, USA, Tel.: 1[240-276-7813], Fax: 1[240-276-7835],
E-mail: lerrocc@mail.nih.gov
Disclaimer: The findings and conclusions in this report are those of
the authors and do not necessarily represent the views of the
National Institute for Occupational Safety and Health.
Epidemiology
use for any particular herbicide, but rather an overall reduction
in the use of these six herbicides.3 EPA re-affirmed acetochlor
registration for use on field corn and popcorn in 2006.4 Over
32 million pounds of acetochlor were applied during 1997–
2001 on approximately 25% of corn crops in the US5; similar
amounts continue to be applied in recent years.6
The EPA considers acetochlor to have “suggestive evidence
of carcinogenic potential” based on increased incidence of cer-
tain tumors in laboratory animals7; other agencies have not
reviewed the carcinogenicity of this chemical. Acetochlor is not
considered mutagenic or genotoxic at relevant doses,8 however,
evidence suggests potential for acetochlor-induced chromosomal
aberration at cytotoxic doses.1 In animal studies, lung tumors
and histiocytic sarcomas have been reported in mice, while nasal
olfactory tumors and thyroid follicular cell tumors have been
reported in Sprague-Dawley rats.1 Liver adenomas and carcino-
mas, renal adenomas and sarcomas and benign ovarian tumors
have been reported in rats and mice, however, generally only at
excessively toxic doses or demonstrated weak associations with

Int. J. Cancer: 137, 1167–1175 (2015) 2015 UICC

 1168
What’s new?
Acetochlor is a commonly used herbicide in the U.S., yet no epidemiologic research has evaluated its carcinogenicity in
humans. In this study, the authors examined the relationship between occupational exposure to acetochlor and human cancer
risk in a large prospective cohort. Acetochlor use was associated with an increased risk of lung and colorectal cancer, and
possibly pancreatic cancer and melanoma. Use of mixtures of acetochlor and atrazine, another widely used herbicide often
applied concurrently with acetochlor, was also associated with an increased risk of lung cancer.
acetochlor exposure.1 Bioassays of other chloroacetanilide herbi-
cides have overlapping but not identical tumor profiles.1
Epidemiologic research examining the relationship between
acetochlor exposure and non-cancer human health outcomes is
limited. Acetochlor levels in umbilical cord blood of full-term
infants were associated with risk of low birth weight in a pilot
study from a Chinese birth cohort.9 Conversely, acetochlor levels
in maternal urine during pregnancy were not associated with
any adverse birth outcomes in a French birth cohort.10 A study
of various pesticides found an inverse association between aceto-
chlor exposure and semen quality, although this relationship was
non-significant after excluding men with other risk factors for
reduced semen quality.11 Though there have been no epidemio-
logic studies of acetochlor and cancer risk to date, studies have
evaluated other chloroacetanilide chemicals. Previous findings
from the Agricultural Health Study (AHS) noted associations
between alachlor exposure and lymphohematopoietic cancers,12
and between metolachor exposure and lung cancer.13,14
Because of the limited information on cancer risk with
acetochlor exposure, we examined occupational exposure to
acetochlor and subsequent cancer outcomes among pesticide
applicators in the AHS.
Material and Methods
Study population
The AHS cohort has been described elsewhere in detail.15 Briefly,
the AHS is a prospective cohort that includes 57,310 licensed pes-
Epidemiology
ticide applicators enrolled during 1993–1997 in Iowa and North
Carolina. A follow-up interview was conducted during 1999–
2003 for private applicators and 2003–2005 for commercial appli-
cators, with 36,342 total respondents. Incident cancer cases were
obtained via linkage with Iowa and North Carolina state cancer
registries. Cancer site was classified according to the International
Classification of Diseases for Oncology (third revision).16 Due to
its recent introduction in 1994, we ascertained acetochlor use
only at follow-up interview. Thus, we analyzed first primary can-
cers diagnosed from the date of follow-up interview through last
date of study follow-up (December 31, 2011 for Iowa, December
31, 2010 for North Carolina). The study protocol was approved
by the institutional review boards of the National Institutes of
Health, University of Iowa, and all other contractors in compli-
ance with applicable requirements of the US.
Exposure assessment
Computer-assisted telephone interviews assessed the use of
acetochlor and other factors, the texts of which are available
Use of acetochlor and cancer incidence
at http://aghealth.nih.gov/background/questionnaires.html.
Participants were asked to list all pesticides used on each
crop during the last year they farmed. Additionally, they
were probed to specify if they personally mixed, handled or
applied each pesticide, and how many days per year they
applied the pesticide. Where trade names were reported, indi-
vidual active ingredients were abstracted from the product
trade names.
Whether or not an applicator reported any application of
acetochlor (yes/no) was examined in order to broadly classify
personal use. Total years of acetochlor use for each applicator
was assumed to be number of years between the year the
applicator last farmed (follow-up interview reference year)
and the enrollment year; we used 1994 (year of acetochlor
registration) for applicators enrolled prior to this date. The
number of lifetime days of acetochlor use was calculated by
multiplying the number of days per year using acetochlor by
total years of use. Intensity-weighted lifetime exposure days
were computed by multiplying an intensity weighting factor
by lifetime exposure days. The intensity weighting factor is a
function of pesticide handling practices and personal protec-
tive equipment used.17 Exposure days and intensity-weighted
exposure days were classified as low (less than the median
among all exposed cancer cases) or high (greater than or
equal to the median among all exposed cancer cases). Addi-
tionally, because acetochlor is frequently applied as a mixture
with atrazine (53% of acetochlor applicators reported the use
of such a mixture), we classified the use as (i) never applied
acetochlor or atrazine, (ii) applied acetochlor but not atra-
zine, (iii) applied atrazine but not acetochlor or (iv) applied
both acetochlor and atrazine. Application of both acetochlor
and atrazine were further separated into those who reported
the use of acetochlor/atrazine mixtures (based on trade
names of product mixtures including acetochlor and atrazine)
and those who reported use of acetochlor and atrazine
separately.
Statistical analysis
Analyses were limited to individuals who responded to the
follow-up interview (n 5 36,342), and we further restricted
to male pesticide applicators due to the small number of
female applicators (n 5 1,006) and few exposed cancer cases
(n 5 4). We excluded men with cancer diagnoses prior to
completion of the follow-up interview (n 5 1,675) and men
with missing or zero person-years of follow-up (n 5 177),
leaving 33,484 applicators available for analysis. Analyses
Int. J. Cancer: 137, 1167–1175 (2015) 2015 UICC
Lerro et al. 1169

Table 1. Selected characteristics of AHS pesticide applicators1 who completed follow-up questionnaire (n 5 33,017), stratified by lifetime
intensity-weighted days of acetochlor use (none, low, high)2
Acetochlor use
Total Never Low use High use p-value3
N 5 33,017 N 5 29,458 N 5 1,621 N 5 1,938
N (%) N (%) N (%) N (%)
State
Iowa 21,810 (66.06) 18,297 (62.11) 1,601 (98.77) 1,912 (98.66) p < 0.0001
North Carolina 11,207 (33.94) 11,161 (37.89) 20 (1.23) 26 (1.34)
Type of applicator
Commercial 2,527 (7.65) 1,797 (6.10) 141 (8.70) 589 (30.39) p < 0.0001
Private 30,490 (92.35) 27,661 (93.9) 1,480 (91.30) 1,349 (69.61)
Age at follow-up
Quartile 1: <43.4 8,319 (25.20) 7,058 (23.96) 543 (33.50) 718 (37.05) p < 0.0001
Quartile 2: 43.4–51.2 8,260 (25.02) 7,234 (24.56) 430 (26.53) 596 (30.75)
Quartile 3: 51.3–61.4 8,227 (24.92) 7,431 (25.23) 362 (22.33) 434 (22.39)
Quartile 4: 61.5 8,211 (24.87) 7,735 (26.26) 286 (17.64) 190 (9.80)
4
Educational attainment
<High school 2,624 (7.95) 2,532 (8.60) 46 (2.84) 46 (2.37) p < 0.0001
High school graduate 14,774 (44.75) 13,168 (44.70) 734 (45.28) 872 (44.99)
>High school 14,383 (43.56) 12,596 (42.76) 813 (50.15) 974 (50.26)
Other/missing 1,236 (3.74) 1,162 (3.94) 28 (1.73) 46 (2.37)
Race
White 31,685 (95.97) 28,166 (95.61) 1,602 (98.83) 1,917 (98.92) p < 0.0001
Non-White 728 (2.20) 718 (2.44) 7 (0.43) 3 (0.15)
Other/missing 604 (1.83) 574 (1.95) 12 (0.74) 18 (0.93)
Tobacco use4
Never smoker 17,302 (52.40) 15,069 (51.15) 1,015 (62.62) 1,218 (62.85) p < 0.0001
Former smoker, 10 years 4,896 (14.83) 4,380 (14.87) 255 (15.73) 261 (13.47)
Former smoker, >10 years 4,548 (13.77) 4,225 (14.34) 159 (9.81) 164 (8.46)
Current smoker, 10 years 917 (2.78) 813 (2.76) 41 (2.53) 63 (3.25)

Epidemiology
Current smoker, >10 years 3,885 (11.77) 3,574 (12.13) 114 (7.03) 197 (10.17)
Missing 1,469 (4.45) 1,397 (4.74) 37 (2.28) 35 (1.81)
Alcohol consumption
No 17,702 (53.61) 16,291 (55.30) 711 (43.86) 700 (36.12) p < 0.0001
Yes 15,025 (45.51) 12,886 (43.74) 905 (55.83) 1,234 (63.67)
Missing 290 (0.88) 281 (0.95) 5 (0.31) 4 (0.21)
Family history of cancer4
No 17,390 (52.67) 15,347 (52.10) 876 (54.04) 1,167 (60.22) p < 0.0001
Yes 12,971 (39.29) 11,579 (39.31) 678 (41.83) 714 (36.84)
Missing 2,656 (8.04) 2,532 (8.60) 67 (4.13) 57 (2.94)
Use of enclosed cab
No 20,063 (60.77) 19,048 (64.66) 434 (26.77) 581 (29.98) p < 0.0001
Yes 12,954 (39.23) 10,410 (35.34) 1,187 (73.23) 1,357 (70.02)
Atrazine use
No 22,579 (68.39) 21,932 (74.45) 329 (20.30) 318 (16.41) p < 0.0001
Yes 10,438 (31.61) 7,526 (25.55) 1,292 (79.7) 1,620 (83.59)
1
Reported on follow-up questionnaire.
2
Low: 72-879 intensity-weighted days, High: 880- 47,520 intensity-weighted days.
3
Chi-square test for homogeneity.
4
Reported on enrollment questionnaire.
 1170 Use of acetochlor and cancer incidence

Table 2. Relative risks (RR) and 95% confidence intervals (95% CI)1 pesticides most highly associated with acetochlor use (life-
for ever-use of acetochlor, relative to never-use and risk for various time use: imazethapyr, dicamba, atrazine, EPTC, cyanazine,
cancers among AHS applicators (n 5 33,484).
pendimethalin, trifluralin; follow-up only: flumetsulam, clo-
Nunexposed Nexposed RR (95% CI) pyralid, isoxaflutole), pesticides mentioned in the acetochlor
All sites 2,930 304 1.06 (0.94–1.21) EPA registration (alachlor, metolachlor, atrazine, EPTC, 2,4-
Bladder 167 15 1.08 (0.61–1.91) D, butylate) and pesticides previously found to be associated
Lymphohematopoietic 293 23 0.77 (0.49–1.20) with specific cancer outcomes in the AHS.19 Tests for trend
Colon 206 21 1.07 (0.66–1.73)
used the midpoint of each exposure category treated as a
continuous variable. Sensitivity analyses were conducted in
Colorectal 300 31 1.03 (0.69–1.53)
which the cohort was restricted to cases diagnosed more
Esophagus 42 6 1.28 (0.47–3.44) than two years after completion of follow-up interview. All
Kidney 98 13 1.14 (0.61–2.11) tests were two-sided with a 5 0.05. Data in this analysis are
Leukemia 93 9 0.89 (0.43–1.84) based on the AHS data releases P2REL201209.00 and
Lung 251 23 1.74 (1.07–2.84)** P1REL201209.00.
Melanoma 124 23 1.61 (0.98–2.66)*
Results
NHL 249 18 0.70 (0.43–1.16)
Table 1 displays selected demographic, behavioral, health
Pancreas 55 7 2.36 (0.98–5.65)* and farming characteristics of the cohort, stratified by ace-
Prostate 1,232 130 0.99 (0.82–1.20) tochlor use. Acetochlor was used almost exclusively by
Prostate, aggressive 644 69 0.98 (0.75–1.28) applicators in Iowa (99%). A larger proportion of commer-
Rectum 94 10 0.97 (0.48–1.95) cial applicators used acetochlor compared to private appli-
cators (28.9% and 9.3%, respectively). Applicators who used
*Significant at p < 0.1. acetochlor were more likely to be younger, more educated
**Significant at p < 0.05.
1
Adjusted for age, race, state, applicator type, smoking, family history and white compared to non-users. Applicators in Iowa sim-
of cancer, alcohol consumption, BMI, use of an enclosed cab, educa- ilarly tend to be younger, more educated and white com-
tion and correlated/associated pesticide use. pared to applicators in North Carolina, so these differences
may be regional and not directly related to acetochlor use.
examining lifetime days and intensity-weighted days of ace- Additionally, acetochlor users were more likely than non-
tochlor use further excluded those missing days of use users to be never-smokers and ever-drinkers in the last 12
(n 5 456) or intensity (n 5 11), leaving 33,028 and 33,017 months, have no family history of cancer, and report that
applicators, respectively. the tractor regularly used to apply pesticides had an
We report results for all cancer sites with at least 15 enclosed cab.
exposed cases for days and intensity-weighted days of use, We evaluated the use of acetochlor in association with a
and results for ever use for sites with more than five exposed variety of cancer sites, controlling for the covariates described
cases. Colon and rectal tumors were combined as were lym- above (Table 2). We observed an association with ever-use of
Epidemiology

phohematopoietic cancers. Aggressive prostate cancer, as
defined by Koutros et al., was examined separately.18 Relative
risks (RR) and 95% confidence intervals (CI) were estimated
using Poisson regression in SAS version 9.3 (SAS Institute,
Cary, NC). Subjects contributed person-time from date of
follow-up interview through date of first cancer diagnosis,
date moved out of state, date of death or last follow-up,
whichever occurred first.
All models were adjusted for potential confounders
including age at follow-up interview (continuous), state of
residence (Iowa or North Carolina), applicator type (private
or commercial), duration of cigarette smoking as reported at
enrollment (never, <10 years, 10–19 years, 20–29 years, 301
years, missing), race (white, other, missing), alcohol use
(yes/no in preceding 12 months, missing), educational
attainment (less than a high school degree, high school
degree, more than a high school degree, missing), use of an
enclosed tractor cab (yes, no, missing) and family history of
cancer in first degree relatives (yes, no, missing; specific can-
cer site where available). We also adjusted for ever-use of
acetochlor and increased risk of lung cancer (RR 5 1.74; 95%
CI: 1.07–2.84), melanoma (RR 5 1.61; 95% CI: 0.98–2.66)
and pancreatic cancer (RR 5 2.36; 95% CI: 0.98–5.65),
although the associations for melanoma and pancreatic can-
cer were of borderline statistical significance.
Compared to never use of acetochlor, low use of aceto-
chlor, as measured by lifetime days used, was associated with
a decreased risk of colorectal cancer (RR 5 0.31; 95% CI:
0.11–0.83) while high use was associated with an increased
risk of colorectal cancer (RR 5 1.75; 95% CI: 1.08–2.83), with
a borderline significant exposure–response trend (p-
trend 5 0.07, Table 3). Results were similar for colorectal can-
cer risk and lifetime intensity-weighted days of acetochlor
use. Low use of acetochlor, as measured by lifetime days
used, was associated with an increased risk of lung cancer
(RR 5 2.64; 95% CI: 1.47–4.74) compared to never use; no
association was seen for high use and no exposure–response
trend was observed (p-trend 5 0.36). Similarly, low use of
acetochlor, as measured by lifetime intensity-weighted days
used, was associated with an increased risk of lung cancer

Int. J. Cancer: 137, 1167–1175 (2015) 2015 UICC

Table 3. Relative risks (RR) and 95% confidence intervals (95% CI)1 by days (n 5 33,028) and intensity-weighted days (n 5 33,017) of aceto-
chlor use, compared to never-use, for various cancers
Acetochlor intensity-weighted
Acetochlor days use2 days use3
N RR (95% CI) N RR (95% CI)
All Sites Never 2,930 1.00 (REF) 2,930 1.00 (REF)
Low 119 0.99 (0.82–1.20) 127 1.00 (0.83–1.20)
High 139 1.19 (1.00–1.43)* 131 1.20 (1.00–1.45)*
p-trend 0.07* 0.06*
Bladder Never 167 1.00 (REF) 167 1.00 (REF)
Low 7 1.11 (0.51–2.44) 6 0.91 (0.39–2.10)
High 4 0.74 (0.26–2.09) 5 1.01 (0.40–2.57)
p-trend 0.63 0.98
Lymphohematopoietic Never 293 1.00 (REF) 293 1.00 (REF)
Low 12 0.93 (0.51–1.68) 10 0.76 (0.40–1.44)
High 9 0.80 (0.40–1.59) 11 1.02 (0.55–1.91)
p-trend 0.51 0.91
Colon Never 206 1.00 (REF) 206 1.00 (REF)
Low 4 0.46 (0.17–1.27) 4 0.44 (0.16–1.20)
High 12 1.56 (0.83–2.91) 12 1.67 (0.90–3.11)
p-trend 0.28 0.20
Colorectal Never 300 1.00 (REF) 300 1.00 (REF)
Low 4 0.31 (0.11-0.83)** 7 0.51 (0.24–1.09)*
High 21 1.75 (1.08–2.83)** 18 1.59 (0.95–2.64)*
p-trend 0.07* 0.16
Kidney Never 98 1.00 (REF) 98 1.00 (REF)
Low 4 0.81 (0.29–2.25) 3 0.57 (0.18–1.86)
High 7 1.53 (0.67–3.47) 8 1.87 (0.86–4.03)
p-trend 0.36 0.16
Lung Never 251 1.00 (REF) 251 1.00 (REF)
Low 14 2.64 (1.47–4.74)** 13 2.26 (1.24–4.14)**
High 7 1.14 (0.51–2.57) 8 1.47 (0.68–3.16)
p-trend 0.36 0.16
Melanoma Never 124 1.00 (REF) 124 1.00 (REF)

Epidemiology
Low 8 1.41 (0.67–2.97) 11 1.79 (0.93–3.45)*
High 11 1.78 (0.90–3.52)* 8 1.38 (0.64–2.99)
p-trend 0.08* 0.29
NHL Never 249 1.00 (REF) 249 1.00 (REF)
Low 11 1.01 (0.54–1.87) 10 0.89 (0.47–1.71)
High 5 0.50 (0.20–1.25) 6 0.63 (0.28–1.46)
p-trend 0.16 0.27
Prostate Never 1,232 1.00 (REF) 1,232 1.00 (REF)
Low 52 0.95 (0.72–1.27) 57 0.99 (0.75–1.30)
High 59 1.10 (0.84–1.46) 54 1.08 (0.81–1.44)
p-trend 0.54 0.63
Prostate, Aggressive Never 644 1.00 (REF) 644 1.00 (REF)
Low 27 0.92 (0.62–1.36) 30 0.97 (0.67–1.42)
High 33 1.20 (0.83–1.74) 30 1.16 (0.78–1.70)
p-trend 0.40 0.50

*Significant at p < 0.1.

**Significant at p < 0.05.
1
Adjusted for age, race, state, applicator type, smoking, family history of cancer, alcohol consumption, BMI, use of an enclosed cab, education, and
correlated/associated pesticide use.
2
Low: 2–19 days, High: 20–1,080 days.
3
Low: 72–879 intensity-weighted days, High: 880–47,520 intensity-weighted days.
 1172
(RR 5 2.26; 95% CI: 1.24–4.14, p-trend 5 0.16). High use of
acetochlor, as measured by lifetime days used, was associated
with borderline increased risk of melanoma (RR 5 1.78; 95%
CI: 0.90–3.52, p-trend 5 0.08) compared to never use. How-
ever, when measured using lifetime intensity-weighted days
used, low acetochlor use was associated with borderline
increased risk of melanoma (RR 5 1.79; 95% CI: 0.93–3.45,
p-trend 5 0.29).
Over 80% of acetochlor users also reported the use of
atrazine at follow-up interview (Table 1); 53% of applicators
reporting ever use of acetochlor report using it in a product
also containing atrazine. We found that applicators using
both atrazine and acetochlor were at increased risk of lung
cancer (RR 5 2.01; 95% CI: 1.17–3.46) compared to those
with no use of either (Table 4). This association was similarly
strong for persons using acetochlor/atrazine product mixtures
(RR 5 2.33; 95% CI: 1.30–4.17), compared to use of neither
atrazine nor acetochlor. Among those exposed to acetochlor
as a mixture only, we saw elevated risks among both low
(RR 5 2.71; 95% CI: 1.14–5.78) and high (RR 5 1.96; 95% CI:
0.75–5.11) categories of intensity-weighted days of exposure
(results not shown). We also observed a borderline significant
excess risk of pancreatic cancer among users of atrazine/ace-
tochlor product mixtures (RR 5 2.62; 95% CI: 0.95–7.20).
The findings for lung cancer, colorectal cancer, mela-
noma and pancreatic cancer were similar in magnitude and
direction after restricting the study population to those
diagnosed with cancer at least two years after completion
of the follow-up interview, though the risk estimates for
ever use of lung and pancreatic cancer no longer reached
statistical significance. We limited lung and pancreatic can-
cer analyses to never smokers to examine the results inde-
pendent of smoking status. The results were in a similar
direction and magnitude for low acetochlor exposure (days
of use) and risk of lung cancer (RR 5 3.30; 95% CI: 0.89–
Epidemiology
12.27), though the association was only borderline statisti-
cally significant. It is possible that the small number of
exposed cases (n 5 3) caused the estimates to be unstable.
We saw a stronger association for ever use of acetochlor
and risk of pancreatic cancer when restricting to never
smokers (RR 5 5.61; 95% CI: 1.58–19.97). We also
restricted our analyses to Iowa applicators, as the majority
of acetochlor was applied there (98.8%); the results were
overall very similar. Approximately, half of our study popu-
lation had complete information regarding potential con-
founders for colorectal cancer (physical activity, fruit and
vegetable consumption) and melanoma (sun sensitivity and
sun protection). When we limited our analyses to these
men, the relationship for high days of acetochlor use and
colorectal cancer was similar in magnitude and direction
(RR 5 1.71; 95% CI: 0.88–3.33) after controlling for physi-
cal activity and dietary factors, and the relationship
between ever use of acetochlor and melanoma risk was
strengthened (RR 5 2.55; 95% CI: 1.45–4.48) after control-
ling for sun sensitivity and exposure factors. We also exam-
Use of acetochlor and cancer incidence
ined days and intensity-weighted days of acetochlor use per
year, classified as high or low based on the median among
cancer cases, in order to determine whether number of
years between enrollment and follow-up was skewing our
results. The results were consistent with findings for days
and intensity-weighted days, and we additionally observed
a significant exposure-response trend for intensity-weighted
days per year of acetochlor use and lung cancer risk (p-
trend 5 0.05).
Discussion
In this analysis of reported use of acetochlor among AHS
applicators, we found elevated risk for several cancer sites.
To our knowledge, this is the first epidemiologic study
attempting to evaluate cancer risk from occupational use of
acetochlor. Specifically, we observed an increased risk of colo-
rectal cancer among pesticide applicators with high lifetime
use of acetochlor, and increased risk of lung cancer among
ever users and low lifetime users of acetochlor. Additionally,
ever use and increasing days of acetochlor use were associ-
ated with borderline significantly increased risk of melanoma.
Though the number of exposed cases was small, there was a
borderline significant increased risk of pancreatic cancer
among ever users of acetochlor.
The finding that the use of acetochlor is associated with
increased risk of lung cancer is complicated because it was
observed among those with below median use, but not
among those with higher exposure. However, this finding
was robust in sensitivity analyses restricting the cohort to
persons diagnosed two or five years after interview. This
suggests that reporting of acetochlor use was not likely to
be influenced by early disease. A previous AHS analysis
found that high use of metolachlor, another chloracetani-
lide herbicide, was associated with an increased risk of lung
cancer.14 The observed association in our study did not
appear to be related to previous metolachlor use, as there
was very little correlation between metolachlor and aceto-
chlor use (q 5 0.11). Concerned about the possibility of
residual confounding, we adjusted for smoking using sev-
eral metrics, including duration (years smoked), total ciga-
rette exposure (pack-years) and current/former/never
smoking status; all produced similar results and duration
was chosen based on model fit. We saw results of a similar
direction and magnitude for low acetochlor exposure and
risk of lung cancer when we restricted our sample to never
smokers. Because we did not see an excess of lung cancer
among the more highly exposed individuals and there was
no significant exposure-response trend, the association may
be spurious. Though we were underpowered to do so,
future studies should attempt to examine lung cancer risk
by histologic subtype; we noted slight differences in lung
cancer histologic subtype by acetochlor exposure in
descriptive analyses.
Colorectal cancer is not often associated with occupational
exposures20; however, we observed an association between
Int. J. Cancer: 137, 1167–1175 (2015) 2015 UICC
Lerro et al. 1173

Table 4. Relative risk (RR) and 95% confidence intervals (95% CI)1 for ever applying acetochlor and atrazine alone or as a mixture, compared
to never use, for selected cancers
N RR (95% CI)
All Sites Neither acetochlor or atrazine 2,282 1.00 (REF)
Acetochlor 59 0.99 (0.76–1.29)
Atrazine 648 1.07 (0.97–1.17)
Both acetochlor and atrazine 245 1.12 (0.97–1.29)
Applied separately 83 1.20 (0.95–1.50)
Applied as a mixture 162 1.08 (0.91–1.28)
Colorectal Neither acetochlor or atrazine 240 1.00 (REF)
Acetochlor 6 0.87 (0.38–1.99)
Atrazine 60 0.91 (0.67–1.23)
Both acetochlor and atrazine 25 1.03 (0.66–1.61)
Applied separately 8 1.01 (0.49–2.11)
Applied as a mixture 17 1.04 (0.62–1.75)
Lung Neither acetochlor or atrazine 204 1.00 (REF)
Acetochlor 4 1.96 (0.70–5.45)
Atrazine 47 1.32 (0.93–1.88)
Both acetochlor and atrazine 19 2.01 (1.17–3.46)**
Applied separately 4 1.29 (0.45–3.68)
Applied as a mixture 15 2.33 (1.30–4.17)**
Melanoma Neither acetochlor or atrazine 89 1.00 (REF)
Acetochlor 6 2.24 (0.94–5.33)*
Atrazine 35 1.39 (0.91–2.14)
Both acetochlor and atrazine 17 1.75 (0.97–3.14)*
Applied separately 6 1.87 (0.77–4.51)
Applied as a mixture 11 1.69 (0.86–3.34)
Pancreas Neither acetochlor or atrazine 49 1.00 (REF)
Acetochlor 2 2.62 (0.6–11.44)
Atrazine 6 0.63 (0.26–1.55)
Both acetochlor and atrazine 5 1.84 (0.67–5.08)

Epidemiology
Applied separately 0 –
Applied as a mixture 5 2.62 (0.95–7.20)*

*Significant at p <0.1.
**Significant at p < 0.05.
1
Adjusted for age, race, state, applicator type, smoking, family history of cancer, alcohol consumption, BMI, use of an enclosed cab, education and
correlated/associated pesticide use.

reported acetochlor use and risk of colorectal cancer, with
some indication of an exposure–response trend. In the AHS,
several pesticides have been associated with colon and rectal
cancers; aldicarb, dicamba, EPTC, imazethapyr and trifluralin
were associated with colon cancer, and chlordane, chlorpyri-
fos, pendimethalin and toxaphene were associated with rectal
cancer.19 A small study of manufacturing workers exposed to
alachlor, a chemical structurally related to acetochlor, found
elevated risks of colorectal cancer.21 A limitation in our study
is that while we could control for body mass index, an estab-
lished risk factor for colorectal cancer,22 we were not able to
control for other known risk factors such as dietary habits or
physical activity,20 which were only collected for a portion of
the cohort. When restricted to individuals who provided
information on leisure time physical activity and average fruit
and vegetable consumption, our results remained relatively
unchanged.
We observed an association between melanoma and ever
use of acetochlor, though the exposure–response relationship
was not consistent for days and intensity-weighted days of
use. The relationship between pesticides and melanoma is
complicated by other occupational exposures, particularly

Int. J. Cancer: 137, 1167–1175 (2015) 2015 UICC

 1174
extended exposure to natural ultraviolet radiation,23 however
several pesticides have been associated with melanoma in
previous AHS analyses.24,25 Sun sensitivity and exposure
characteristics were only available for about half of our
cohort; however when we limited our analyses to men with
valid response and controlled for these factors, the relation-
ship between ever use of acetochlor and melanoma risk was
strengthened.
Occupationally, pancreatic cancer has been associated
with chlorinated hydrocarbon solvents,26 though findings for
chlorinated pesticides and pancreatic cancer have been less
clear.27–30 In the AHS, EPTC and pendimethalin were asso-
ciated with increased risk of pancreatic cancer, possibly via
the formation of N-nitroso-compounds.31 Smoking, an estab-
lished risk factor for pancreatic cancer, was controlled for in
our analyses and we observed a significantly increased risk
of pancreatic cancer among never smokers. While we did
not present the results due to small number of exposed
cases, we did see a significant exposure–response trend for
intensity-weighted days of acetochlor use and pancreatic
cancer (p-trend 5 0.01). Future studies should evaluate these
findings.
We evaluated atrazine and acetochlor product mixtures
because these active ingredients are often applied as a mix-
ture in our cohort. Previous studies in the AHS have not
been able to explicitly examine product mixtures because of
how pesticide use was assessed at enrollment.14,32,33 At
follow-up interview, participants self-reported specific pesti-
cide product names, and based on this information we deter-
mined whether acetochlor was being applied alone or as a
mixture with atrazine. These analyses did not account for
applicators who mixed individual ingredients or products on
their own (as opposed to using pre-mixed formulations); this
information was not collected. Little is known about the toxi-
cology and potential carcinogenicity of pesticide mixtures.
Epidemiology
Epidemiologic studies usually examine pesticides either inde-
pendently, or more often by chemical class. Findings here
suggest that future epidemiologic studies should consider the
effects of pesticide mixtures, particularly acetochlor and atra-
zine, and future toxicological studies should attempt to
understand whether exposure as a mixture influences geno-
toxicity and mutagenicity.
Acetochlor, like most pesticides licensed for use today,
does not appear to be mutagenic or genotoxic in laboratory
studies.1 Therefore, other mechanisms of carcinogenesis
should be considered. Occupational use of other chloracetani-
lide herbicides alachlor and metolachlor was significantly
associated with shortened telomere length in a recent AHS
study,34 acetochlor was not examined. Telomere length has
been implicated in cancer etiology.35 Another proposed
mechanism of carcinogenicity for chloracetanilide herbicides
involves bioactivation through several steps to DNA-reactive
metabolites (2-methyl-6-ethylbenzoquinone imine for aceto-
chlor and metolachlor and 2,6-diethylbenzoquinone imine for
Use of acetochlor and cancer incidence
alachlor).7 These chloracetanilide metabolites, 2-methyl-6-
ethylbenzoquinone imine and 2,6-diethylbenzoquinone imine,
may lead to the formation of DNA adducts or single-strand
DNA breaks.7,36,37 Studies suggest that triazine herbicides,
such as atrazine, stimulate cytochrome P450 (CYP450) sys-
tem activity.38 Acetochlor is metabolized and potentially bio-
activated by members of the CYP450 family of enzymes,
CYP2B6 and CYP34A, in the liver.7 Thus, when atrazine and
acetochlor are applied together, it is possible that there is
increased acetochlor bioactivation as a result of atrazine-
induced CYP450 activity.
Our analysis is the first epidemiologic study to examine
the effect of acetochlor on cancer outcomes in humans.
Strengths include longitudinal study design with regular
follow-up of participants for cancer and mortality outcomes,
as well as detailed and validated self-reported pesticide expo-
sure and intensity information.39 An important limitation for
our analysis was sample size; although AHS is a large cohort,
only about 12% (n 5 4,026) of our sample reported any ace-
tochlor use. Additionally, because acetochlor was introduced
after AHS enrollment and therefore assessed only in follow-
up interviews, the follow-up period is shorter than analyses
of other chemicals, and thus fewer cancer cases were accrued.
Because acetochlor was registered for use relatively recently,
the observed associations for acetochlor and solid tumors
may be due to earlier exposures and reflect long latency peri-
ods for these cancers. However, we restricted our cohort to
those diagnosed at least two and five years after the start date
of the study, and did not see marked differences in the mag-
nitude and direction of the effects for lung cancer, colorectal
cancer, pancreatic cancer and melanoma. Applicators in our
cohort were typically applying other pesticides prior to aceto-
chlor registration and concurrently at follow-up interview.
We controlled for use of pesticides that were highly corre-
lated with acetochlor, as well as herbicides that were specified
in the acetochlor EPA registration, in order to minimize
these possible sources of confounding. Due to limited num-
bers of cases, we were unable to evaluate rarer cancer sites,
as well as histologic subtypes of more common cancers. Due
to few exposed women, we were unable to evaluate cancer
sites more common in females such as breast or thyroid
cancer.
In conclusion, we observed associations between aceto-
chlor use and lung cancer, colorectal cancer, pancreatic can-
cer and melanoma. However, due to minimal evidence for
exposure–response relationships, the observed associations
could reflect chance findings. Findings regarding the carcino-
genicity of acetochlor need further evaluation in populations
with sufficient numbers of exposed subjects. Additionally,
future studies should attempt to evaluate potential mecha-
nisms of carcinogenesis for acetochlor and other chloroaceta-
nilide herbicides by examining genetic markers of
susceptibility, as well as markers of cellular and DNA
damage.
Int. J. Cancer: 137, 1167–1175 (2015) 2015 UICC
Lerro et al.
References
1. Evaluation of the carcinogenic potential of
acetochlor (fifth Assessment) (121601).
Environmental Protection Agency, Cancer
Assessment Review Committee, Health Effects
Division, Office of Pesticide Programs,
Washington, DC, 2007. 1–66.
2. Green T, Lee R, Moore RB, et al. Acetochlor-
induced rat nasal tumors: further studies on the
mode of action and relevance to humans. Regul
Toxicol Pharmacol 2000;32:127–33.
3. Acetochlor (EPA-HQ-OPP-2007-0725-0002).
Environmental Protection Agency, Washington,
DC, 2007.
4. Acetochlor; Tolerance reassessment decision
(EPA-HQ-OPP-2005-0227), vol. 71.
Environmental Protection Agency, Washington,
DC, 2006. 15730–1.
5. Usage Report in Support of Registration for the
Herbicide Acetochlor (121601). Washington, DC:
Environmental Protection Agency, Office of
Prevention, Pesticides, and Toxic Substances,
2004.
6. Pesticide industry sales and usage: 2006 and 2007
market estimates office of pesticide programs.
Washington, DC: US Environmental Protection
Agency, 2011.
7. Coleman S, Linderman R, Hodgson E, et al.
Comparative metabolism of chloroacetamide her-
bicides and selected metabolites in human and
rat liver microsomes. Environ Health Perspect
2000;108:1151–7.
8. Ashby J, Kier L, Wilson AG, et al. Evaluation of
the potential carcinogenicity and genetic toxicity
to humans of the herbicide acetochlor. Hum Exp
Toxicol 1996;15:702–35.
9. Wickerham EL, Lozoff B, Shao J, et al. Reduced
birth weight in relation to pesticide mixtures
detected in cord blood of full-term infants. Envi-
ron Int 2012;47:80–5.
10. Chevrier C, Limon G, Monfort C, Rouget F,
Garlantezec R, Petit C, Durand G, Cordier S. Uri-
nary biomarkers of prenatal atrazine exposure
and adverse birth outcomes in the PELAGIE
birth cohort. Environ Health Perspect 2011;119:
1034–41.
11. Swan SH, Kruse RL, Liu F, et al. Semen
quality in relation to biomarkers of pesticide
exposure. Environ Health Perspect 2003;111:1478–
84.
12. Lee WJ, Hoppin JA, Blair A, et al. Cancer inci-
dence among pesticide applicators exposed to ala-
Int. J. Cancer: 137, 1167–1175 (2015) 2015 UICC
chlor in the Agricultural Health Study. Am J
Epidemiol 2004;159:373–80.
13. Rusiecki JA, Hou L, Lee WJ, et al. Cancer inci-
dence among pesticide applicators exposed to
metolachlor in the Agricultural Health Study. Int
J Cancer 2006;118:3118–23.
14. Alavanja MC, Dosemeci M, Samanic C, et al.
Pesticides and lung cancer risk in the agricultural
health study cohort. Am J Epidemiol 2004;160:
876–85.
15. Alavanja MC, Sandler DP, McMaster SB, et al.
The Agricultural Health Study. Environ Health
Perspect 1996;104:362–9.
16. Fritz AG. International Classification of Diseases
for Oncology: ICD-Oed. World Health Organiza-
tion, Geneva, Switzerland, 2000.
17. Coble J, Thomas KW, Hines CJ, et al. An
updated algorithm for estimation of pesticide
exposure intensity in the agricultural health
study. Int J Environ Res Public Health 2011;8:
4608–22.
18. Koutros S, Beane Freeman LE, Lubin JH, et al.
Risk of total and aggressive prostate cancer and
pesticide use in the Agricultural Health Study.
Am J Epidemiol 2013;177:59–74.
19. Weichenthal S, Moase C, Chan P. A review of
pesticide exposure and cancer incidence in the
Agricultural Health Study cohort. Environ Health
Perspect 2010;118:1117–25.
20. Schottenfeld D, Fraumeni JF. Cancer epidemiol-
ogy and preventioned. New York, NY, USA:
Oxford University Press, 2006.
21. Acquavella JF, Delzell E, Cheng H, et al. Mortal-
ity and cancer incidence among alachlor manu-
facturing workers 1968–99. Occup Environ Med
2004;61:680–5.
22. Siegel R, Ma J, Zou Z, et al. Cancer statistics,
2014. CA Cancer J Clin 2014;64:9–29.
23. MacKie RM, Hauschild A, Eggermont AM. Epi-
demiology of invasive cutaneous melanoma. Ann
Oncol 2009;20 (Suppl 6):vi1–7.
24. Dennis LK, Lynch CF, Sandler DP, et al. Pesticide
use and cutaneous melanoma in pesticide appli-
cators in the agricultural heath study. Environ
Health Perspect 2010;118:812–7.
25. Purdue MP, Hoppin JA, Blair A, et al. Occupa-
tional exposure to organochlorine insecticides
and cancer incidence in the Agricultural Health
Study. Int J Cancer 2007;120:642–9.
26. Ojajarvi A, Partanen T, Ahlbom A, et al. Risk of
pancreatic cancer in workers exposed to chlori-
1175
nated hydrocarbon solvents and related com-
pounds: a meta-analysis. Am J Epidemiol 2001;
153:841–50.
27. Ji BT, Silverman DT, Stewart PA, et al. Occupa-
tional exposure to pesticides and pancreatic can-
cer. Am J Ind Med 2001;39:92–9.
28. Fryzek JP, Garabrant DH, Harlow SD, et al. A
case–control study of self-reported exposures to
pesticides and pancreas cancer in southeastern
Michigan. Int J Cancer 1997;72:62–7.
29. Garabrant DH, Held J, Langholz B, et al. DDT
and related compounds and risk of pancreatic
cancer. J Natl Cancer Inst 1992;84:764–71.
30. Hoppin JA, Tolbert PE, Holly EA, et al. Pancre-
atic cancer and serum organochlorine levels. Can-
cer Epidemiol Biomarkers Prev 2000;9:199–205.
31. Andreotti G, Freeman LE, Hou L, et al. Agricul-
tural pesticide use and pancreatic cancer risk in
the Agricultural Health Study Cohort. Int J Can-
cer 2009;124:2495–500.
32. Freeman LE, Rusiecki JA, Hoppin JA, et al. Atra-
zine and cancer incidence among pesticide appli-
cators in the agricultural health study (1994-
2007). Environ Health Perspect 2011;119:1253–9.
33. Rusiecki JA, De Roos A, Lee WJ, et al. Cancer
incidence among pesticide applicators exposed to
atrazine in the Agricultural Health Study. J Natl
Cancer Inst 2004;96:1375–82.
34. Hou L, Andreotti G, Baccarelli AA, et al. Lifetime
pesticide use and telomere shortening among
male pesticide applicators in the Agricultural
Health Study. Environ Health Perspect 2013;121:
919–24.
35. Ma H, Zhou Z, Wei S, et al. Shortened telomere
length is associated with increased risk of cancer:
a meta-analysis. PLoS One 2011;6:e20466.
36. Bonfanti M, Taverna P, Chiappetta L, et al. DNA
damage induced by alachlor after in vitro activa-
tion by rat hepatocytes. Toxicology 1992;72:207–
19.
37. Brown MA, Kimmel EC, Casida JE. DNA adduct
formation by alachlor metabolites. Life Sci 1988;
43:2087–94.
38. Hernandez AF, Parron T, Tsatsakis AM, et al.
Toxic effects of pesticide mixtures at a molecular
Epidemiology
level: their relevance to human health. Toxicology
2013;307:136–45.
39. Thomas KW, Dosemeci M, Coble JB, et al.
Assessment of a pesticide exposure intensity algo-
rithm in the agricultural health study. J Expo Sci
Environ Epidemiol 2010;20:559–69.