The Toxicologist - Society of Toxicology

invasiveness at concentrations representative of human exposure (10 -7 M) but had
no effect on resistance to anoikis. DBP had no effect on invasiveness or resistance to
anoikis while its major metabolite increased resistance to anoikis at concentrations
representative of human exposure (10 -8 M). Therefore, at concentrations relevant to
human exposure, endocrine toxicants may promote breast cancer progression and
metastasis by altering resistance to anoikis and invasiveness.
229 ENVIRONMENTAL CHEMICALS AND BREAST
CANCER—GEOGRAPHIC ASSOCIATION BETWEEN
LIVESTOCK WASTE, CONTAMINATED SURFACE
WATER, AND BREAST CANCER INCIDENCE IN THE
SAN FRANCISCO BAY AREA.
H. Larson 1 , M. Mun 1 , W. Onyenwe 1 and D. E. Johnson 1, 2 . 1 Nutritional Science
& Toxicology, University of California Berkeley, Berkeley, CA and 2 California Breast
Cancer & Chemicals Policy Project, San Francisco, CA.
The aim of this study was to introduce a novel approach to understanding the high
incidence of breast cancer in the San Francisco Bay area and, in particular, Marin
County, which has the highest incidence. Geographical and environmental pollution
data were compared with breast cancer statistics in order to produce a
“geomedical” image of the disease and possible association with environmental contaminants.
Technologies and information sources included ArcGIS9, ArcMap, NCI
State Cancer Profiles, Scorecard, and Census of Agriculture Profiles by county. A
strong association occurred between breast cancer incidence and animal waste pollution
from livestock, making this the primary comparative focus. Accordingly,
livestock waste pollution and breast cancer incidence rates were compared among
eight counties in the California Bay Area: Marin, Alameda, Contra Costa, Napa,
San Francisco, San Mateo, Santa Clara, Solano, and Sonoma. Two non-California
counties (King County, WA; Delaware County, OH) were also selected for analysis
based on population demographics, water sources, and potential surface contamination
from livestock waste. The results demonstrate a visible correlation between
high amounts of animal waste and high incidence rates of breast cancer. The definitive
causal link has yet to be identified, but one possible explanation is that improperly
stored waste from large, industrial livestock farms may be contaminating
surface water sources. Farm animals are typically treated with hormones and antibiotics
to promote growth, but these treatments also increase the amount of hormonal
waste excreted. In Marin County, surface water is the primary source of
drinking water; however, adequate well-controlled studies on hormone contamination
and persistence in the supply do not exist.
230 NOVEL P450 REDUCTASE KNOCKOUT MOUSE
MODELS FOR STUDYING CHEMICAL
CARCINOGENESIS IN THE MAMMARY GLAND.
Y. Yao 1 , S. Liu 1 , W. Yang 1 , B. Moorthy 2 , X. Ding 1 and J. Gu 1 . 1 Wadsworth
Center, New York State Department of Health, and School of Public Health, State
University of New York at Albany, Albany, NY and 2 Baylor College of Medicine,
Houston, TX.
Microsomal P450 (P450) enzymes play a critical role in the metabolic activation of
chemical procarcinogens. However, it has been difficult to determine whether, following
systemic exposure to a given procarcinogen (such as 7,12dimethylbenz[a]anthracene,
DMBA), the bulk of the activated carcinogen, or its
DNA adducts, in the breast tissue is derived from local metabolism or from the
liver. The aim of this study is to test the hypothesis that target-tissue P450-mediated
metabolic activation plays an important role in DMBA-induced formation of
DNA adducts in the mammary gland (MG). This hypothesis is tested in mouse
models, by modulating P450 activities in the liver, in all extrahepatic tissues, or in
the MG alone, through conditional deletion (or suppression) of the gene for P450
reductase (Cpr), an enzyme required for the activity of all microsomal P450 enzymes.
Initial studies on the liver-specific Cpr-null mouse indicated that hepatic
P450 enzymes do not contribute significantly to DMBA-induced DNA-adduct
formation in the MG, as determined by [32]P-postlabeling. For a more direct determination
of the role of MG P450 enzymes in DNA-adduct formation, we have
also been studying an extrahepatic-Cpr-low mouse, which has normal P450 activities
in the liver, but much suppressed P450 activities in all extrahepatic tissues. The
level of CPR protein in MG microsomes from the extrahepatic-Cpr-low mouse was
found to be ~10% of the levels found in wild-type mice. In addition, a MG-Cprnull
mouse model, expected to have tissue-specific suppression of P450 activities in
the MG, is also being prepared, by crossbreeding Cpr-lox mouse with the MMTV-
Cre mouse. The extrahepatic-Cpr-low mouse and the MG-Cpr-null mouse will be
studied for determinations of the contributions of target tissue metabolic activation
to DMBA-induced DNA adduct formation in the MG. (Supported in part by NIH
grant ES018884)
231 IDENTIFYING INHIBITION PATHWAYS OF MAMMARY
CANCER STEM CELLS BY CHEMICAL GENETICS.
D. Castro, J. Maurer and R. Oshima. Tumor Development Program, Sanford-
Burnham Medical Research Institute, La Jolla, CA.
Studies have identified subpopulations of cells within tumors that drive tumorigenesis,
termed cancer stem cells (CSCs). CSCs have also displayed increased resistance
to traditional therapies. This persistent viability permits the regeneration of the
tumor and contributes to the failure of chemotherapeutics. We have isolated mammary
cancer stem cells from the mouse mammary tumor virus (MMTV)-Wnt-1
transgenic model of breast cancer. These tumorigenic progenitor cells express markers
of both luminal (keratin 8) and basal cells (keratin 14) as well as stem cell associated
markers for CD24, CD29, and CD49f. Cell cultures enriched for the bipotent
progenitor (K8 + ,K14 + ) have generated tumors with the differentiated
morphology of the tumor of origin from as few as a single cell. We employed a small
library of 243 annotated kinase inhibitors in an effort to establish a chemical sensitivity
profile towards the CSCs. Our initial screen with a dose of 1 μM identified
35 compounds with the ability to reduce cell survival by >50% after a 3 day incubation
period. A cluster of 7 annotated Protein kinase C (PKC) inhibitors exhibited
the most dramatic effects on cell survival. The hydroxylated form of staurosporine,
(UCN-01), a derived anti-cancer agent, suppressed growth at submicromolar concentrations
and produced an IC 50 of 47 nM. The concerted effectiveness of the
PKC inhibitors at low nanomolar concentrations indicate an inhibitory role for
PKC and its downstream targets. In order to demonstrate selective toxicity for our
CSCs, we are currently evaluating the effectiveness of validated targets on representative
mammary cancer cell lines. We also plan to implement this screening method
with a small library of the most current FDA-approved anti-cancer drugs. The
chemical sensitivity profile of our CSCs will eventually be applied to human cancer
cells to identify those that may be regulated in a similar way.
232 SEVERE URINARY BLADDER CYTOTOXICITY AND
REGENERATIVE HYPERPLASIA WITH DOSE
RESPONSE INDUCED BY ARSENITE IN ARSENIC (+3
OXIDATION STATE) METHYLTRANSFERASE
KNOCKOUT MICE.
M. Yokohira 2, 1 , L. L. Arnold 2 , K. L. Pennington 2 , S. Suzuki 2 , S. Kakiuchi-
Kiyota 2 , K. Herbin-Davis 3 , D. J. Thomas 3 , K. Imaida 1 and S. M. Cohen 2 .
1 Oncopathology, Kagawa University, Faculty of Medicine, Kita-gun, Kagawa, Japan,
2 Pathology and Microbiology, University of Nebraska Medical Center, Omaha, NE
and 3 Pharmacokinetics Branch, Integrated Systems Toxicology Division, National
Health and Environmental Effects Research Laboratory, U.S. EPA, Omaha, NE.
Arsenic (+3 oxidation state) methyltransferase (As3mt) catalyzes reactions converting
inorganic arsenic to methylated metabolites. We evaluated whether the As3mt
null genotype in mice modifies cytotoxic and proliferative effects in urinary bladders
of wild type mice after inorganic arsenic exposure. Previously we showed female
As3mt KO mice treated with diet containing 0, 100 or 150 ppm arsenic as arsenite
(AsIII) had systemic toxicity and significant effects on the urothelium. In the
present study, we determined if the cytotoxic and proliferative effects of AsIII on
the urothelium are dose dependent over 4 weeks of treatment. Female wild type
C57BL/6 mice and As3mt KO mice were divided into 5 groups each (n=7) with
free access to drinking water containing 0, 1, 10, 25 or 50 ppm arsenic as AsIII for
4 weeks. At sacrifice, urinary bladders of both As3mt KO and wild type mice
showed hyperplasia by light microscopy. In As3mt KO mice, numerous cytoplasmic
and nuclear granules were present in all urothelial layers and were more abundant
and larger than in wild type mice. In As3mt KO mice, livers showed mild
acute inflammation and kidneys showed hydronephrosis. Thus, diminished arsenic
methylation in As3mt KO mice exacerbates the effects of AsIII on the bladder epithelium,
showing that altered kinetic and dynamic behavior of arsenic can affect its
systemic and urothelial toxicity.
233 TUMORS AND PROLIFERATIVE LESIONS IN ADULT
OFFSPRING AFTER MATERNAL EXPOSURE TO
METHYLARSONOUS ACID DURING GESTATION IN
CD1 MICE.
M. Waalkes 2, 1 , B. A. Diwan 3 , E. J. Tokar 2, 1 and D. J. Thomas 4 . 1 NCI at NIEHS,
Research Triangle Park, NC, 2 NTP, NIEHS, Research Triangle Park, NC, 3 IRSP,
SAIC, NCI, Frederick, MD and 4 U.S. EPA, Research Triangle Park, NC.
Inorganic arsenic exposure is carcinogenic in humans and rodents. When pregnant
mice are exposed to inorganic arsenic in the drinking water their offspring, when
adults, develop tumors and proliferative lesions at several sites, such as lung, liver,
SOT 2011 ANNUAL MEETING 49