9 research outputs found
High-throughput in Vitro Data To Inform Prioritization of Ambient Water Monitoring and Testing for Endocrine Active Chemicals
The presence of industrial chemicals,
consumer product chemicals,
and pharmaceuticals is well documented in waters in the U.S. and globally.
Most of these chemicals lack health-protective guidelines and many
have been shown to have endocrine bioactivity. There is currently
no systematic or national prioritization for monitoring waters for
chemicals with endocrine disrupting activity. We propose ambient water
bioactivity concentrations (AWBCs) generated from high throughput
data as a health-based screen for endocrine bioactivity of chemicals
in water. The U.S. EPA ToxCast program has screened over 1800 chemicals
for estrogen receptor (ER) and androgen receptor (AR) pathway bioactivity.
AWBCs are calculated for 110 ER and 212 AR bioactive chemicals using
high throughput ToxCast data from in vitro screening assays and predictive
pathway models, high-throughput toxicokinetic data, and data-driven
assumptions about consumption of water. Chemical-specific AWBCs are
compared with measured water concentrations in data sets from the
greater Denver area, Minnesota lakes, and Oregon waters, demonstrating
a framework for identifying endocrine bioactive chemicals. This approach
can be used to screen potential cumulative endocrine activity in drinking
water and to inform prioritization of future monitoring, chemical
testing and pollution prevention efforts
Profiling 976 ToxCast Chemicals across 331 Enzymatic and Receptor Signaling Assays
Understanding potential health risks
is a significant challenge
due to the large numbers of diverse chemicals with poorly characterized
exposures and mechanisms of toxicities. The present study analyzes
976 chemicals (including failed pharmaceuticals, alternative plasticizers,
food additives, and pesticides) in Phases I and II of the U.S. EPA’s
ToxCast project across 331 cell-free enzymatic and ligand-binding
high-throughput screening (HTS) assays. Half-maximal activity concentrations
(AC50) were identified for 729 chemicals in 256 assays (7,135 chemical–assay
pairs). Some of the most commonly affected assays were CYPs (CYP2C9
and CYP2C19), transporters (mitochondrial TSPO, norepinephrine, and
dopaminergic), and GPCRs (aminergic). Heavy metals, surfactants, and
dithiocarbamate fungicides showed promiscuous but distinctly different
patterns of activity, whereas many of the pharmaceutical compounds
showed promiscuous activity across GPCRs. Literature analysis confirmed
>50% of the activities for the most potent chemical–assay
pairs
(54) but also revealed 10 missed interactions. Twenty-two chemicals
with known estrogenic activity were correctly identified for the majority
(77%), missing only the weaker interactions. In many cases, novel
findings for previously unreported chemical–target combinations
clustered with known chemical–target interactions. Results
from this large inventory of chemical–biological interactions
can inform read-across methods as well as link potential targets to
molecular initiating events in adverse outcome pathways for diverse
toxicities
Profiling 976 ToxCast Chemicals across 331 Enzymatic and Receptor Signaling Assays
Understanding potential health risks
is a significant challenge
due to the large numbers of diverse chemicals with poorly characterized
exposures and mechanisms of toxicities. The present study analyzes
976 chemicals (including failed pharmaceuticals, alternative plasticizers,
food additives, and pesticides) in Phases I and II of the U.S. EPA’s
ToxCast project across 331 cell-free enzymatic and ligand-binding
high-throughput screening (HTS) assays. Half-maximal activity concentrations
(AC50) were identified for 729 chemicals in 256 assays (7,135 chemical–assay
pairs). Some of the most commonly affected assays were CYPs (CYP2C9
and CYP2C19), transporters (mitochondrial TSPO, norepinephrine, and
dopaminergic), and GPCRs (aminergic). Heavy metals, surfactants, and
dithiocarbamate fungicides showed promiscuous but distinctly different
patterns of activity, whereas many of the pharmaceutical compounds
showed promiscuous activity across GPCRs. Literature analysis confirmed
>50% of the activities for the most potent chemical–assay
pairs
(54) but also revealed 10 missed interactions. Twenty-two chemicals
with known estrogenic activity were correctly identified for the majority
(77%), missing only the weaker interactions. In many cases, novel
findings for previously unreported chemical–target combinations
clustered with known chemical–target interactions. Results
from this large inventory of chemical–biological interactions
can inform read-across methods as well as link potential targets to
molecular initiating events in adverse outcome pathways for diverse
toxicities
Predictive Endocrine Testing in the 21st Century Using <i>in Vitro</i> Assays of Estrogen Receptor Signaling Responses
Thousands
of environmental chemicals are subject to regulatory review for their
potential to be endocrine disruptors (ED). <i>In vitro</i> high-throughput screening (HTS) assays have emerged as a potential
tool for prioritizing chemicals for ED-related whole-animal tests.
In this study, 1814 chemicals including pesticide active and inert
ingredients, industrial chemicals, food additives, and pharmaceuticals
were evaluated in a panel of 13 <i>in vitro</i> HTS assays.
The panel of <i>in vitro</i> assays interrogated multiple
end points related to estrogen receptor (ER) signaling, namely binding,
agonist, antagonist, and cell growth responses. The results from the <i>in vitro</i> assays were used to create an ER Interaction Score.
For 36 reference chemicals, an ER Interaction Score >0 showed 100%
sensitivity and 87.5% specificity for classifying potential ER activity.
The magnitude of the ER Interaction Score was significantly related
to the potency classification of the reference chemicals (<i>p</i> < 0.0001). ERα/ERβ selectivity was also
evaluated, but relatively few chemicals showed significant selectivity
for a specific isoform. When applied to a broader set of chemicals
with <i>in vivo</i> uterotrophic data, the ER Interaction
Scores showed 91% sensitivity and 65% specificity. Overall, this study
provides a novel method for combining <i>in vitro</i> concentration
response data from multiple assays and, when applied to a large set
of ER data, accurately predicted estrogenic responses and demonstrated
its utility for chemical prioritization
Real-Time Growth Kinetics Measuring Hormone Mimicry for ToxCast Chemicals in T‑47D Human Ductal Carcinoma Cells
High-throughput
screening (HTS) assays capable of profiling thousands of environmentally
relevant chemicals for <i>in vitro</i> biological activity
provide useful information on the potential for disrupting endocrine
pathways. Disruption of the estrogen signaling pathway has been implicated
in a variety of adverse health effects including impaired development,
reproduction, and carcinogenesis. The estrogen-responsive human mammary
ductal carcinoma cell line T-47D was exposed to 1815 ToxCast chemicals
comprising pesticides, industrial chemicals, pharmaceuticals, personal
care products, cosmetics, food ingredients, and other chemicals with
known or suspected human exposure potential. Cell growth kinetics
were evaluated using real-time cell electronic sensing. T-47D cells
were exposed to eight concentrations (0.006–100 μM),
and measurements of cellular impedance were repeatedly recorded for
105 h. Chemical effects were evaluated based on potency (concentration
at which response occurs) and efficacy (extent of response). A linear
growth response was observed in response to prototypical estrogen
receptor agonists (17β-estradiol, genistein, bisphenol A, nonylphenol,
and 4-<i>tert</i>-octylphenol). Several compounds, including
bisphenol A and genistein, induced cell growth comparable in efficacy
to that of 17β-estradiol, but with decreased potency. Progestins,
androgens, and corticosteroids invoked a biphasic growth response
indicative of changes in cell number or cell morphology. Results from
this cell growth assay were compared with results from additional
estrogen receptor (ER) binding and transactivation assays. Chemicals
detected as active in both the cell growth and ER receptor binding
assays demonstrated potencies highly correlated with two ER transactivation
assays (<i>r</i> = 0.72; <i>r</i> = 0.70). While
ER binding assays detected chemicals that were highly potent or efficacious
in the T-47D cell growth and transactivation assays, the binding assays
lacked sensitivity in detecting weakly active compounds. In conclusion,
this cell-based assay rapidly detects chemical effects on T-47D growth
and shows potential, in combination with other HTS assays, to detect
environmentally relevant chemicals with potential estrogenic activity
Real-Time Growth Kinetics Measuring Hormone Mimicry for ToxCast Chemicals in T‑47D Human Ductal Carcinoma Cells
High-throughput
screening (HTS) assays capable of profiling thousands of environmentally
relevant chemicals for <i>in vitro</i> biological activity
provide useful information on the potential for disrupting endocrine
pathways. Disruption of the estrogen signaling pathway has been implicated
in a variety of adverse health effects including impaired development,
reproduction, and carcinogenesis. The estrogen-responsive human mammary
ductal carcinoma cell line T-47D was exposed to 1815 ToxCast chemicals
comprising pesticides, industrial chemicals, pharmaceuticals, personal
care products, cosmetics, food ingredients, and other chemicals with
known or suspected human exposure potential. Cell growth kinetics
were evaluated using real-time cell electronic sensing. T-47D cells
were exposed to eight concentrations (0.006–100 μM),
and measurements of cellular impedance were repeatedly recorded for
105 h. Chemical effects were evaluated based on potency (concentration
at which response occurs) and efficacy (extent of response). A linear
growth response was observed in response to prototypical estrogen
receptor agonists (17β-estradiol, genistein, bisphenol A, nonylphenol,
and 4-<i>tert</i>-octylphenol). Several compounds, including
bisphenol A and genistein, induced cell growth comparable in efficacy
to that of 17β-estradiol, but with decreased potency. Progestins,
androgens, and corticosteroids invoked a biphasic growth response
indicative of changes in cell number or cell morphology. Results from
this cell growth assay were compared with results from additional
estrogen receptor (ER) binding and transactivation assays. Chemicals
detected as active in both the cell growth and ER receptor binding
assays demonstrated potencies highly correlated with two ER transactivation
assays (<i>r</i> = 0.72; <i>r</i> = 0.70). While
ER binding assays detected chemicals that were highly potent or efficacious
in the T-47D cell growth and transactivation assays, the binding assays
lacked sensitivity in detecting weakly active compounds. In conclusion,
this cell-based assay rapidly detects chemical effects on T-47D growth
and shows potential, in combination with other HTS assays, to detect
environmentally relevant chemicals with potential estrogenic activity
Real-Time Growth Kinetics Measuring Hormone Mimicry for ToxCast Chemicals in T‑47D Human Ductal Carcinoma Cells
High-throughput
screening (HTS) assays capable of profiling thousands of environmentally
relevant chemicals for <i>in vitro</i> biological activity
provide useful information on the potential for disrupting endocrine
pathways. Disruption of the estrogen signaling pathway has been implicated
in a variety of adverse health effects including impaired development,
reproduction, and carcinogenesis. The estrogen-responsive human mammary
ductal carcinoma cell line T-47D was exposed to 1815 ToxCast chemicals
comprising pesticides, industrial chemicals, pharmaceuticals, personal
care products, cosmetics, food ingredients, and other chemicals with
known or suspected human exposure potential. Cell growth kinetics
were evaluated using real-time cell electronic sensing. T-47D cells
were exposed to eight concentrations (0.006–100 μM),
and measurements of cellular impedance were repeatedly recorded for
105 h. Chemical effects were evaluated based on potency (concentration
at which response occurs) and efficacy (extent of response). A linear
growth response was observed in response to prototypical estrogen
receptor agonists (17β-estradiol, genistein, bisphenol A, nonylphenol,
and 4-<i>tert</i>-octylphenol). Several compounds, including
bisphenol A and genistein, induced cell growth comparable in efficacy
to that of 17β-estradiol, but with decreased potency. Progestins,
androgens, and corticosteroids invoked a biphasic growth response
indicative of changes in cell number or cell morphology. Results from
this cell growth assay were compared with results from additional
estrogen receptor (ER) binding and transactivation assays. Chemicals
detected as active in both the cell growth and ER receptor binding
assays demonstrated potencies highly correlated with two ER transactivation
assays (<i>r</i> = 0.72; <i>r</i> = 0.70). While
ER binding assays detected chemicals that were highly potent or efficacious
in the T-47D cell growth and transactivation assays, the binding assays
lacked sensitivity in detecting weakly active compounds. In conclusion,
this cell-based assay rapidly detects chemical effects on T-47D growth
and shows potential, in combination with other HTS assays, to detect
environmentally relevant chemicals with potential estrogenic activity
Development and Validation of a Computational Model for Androgen Receptor Activity
Testing thousands
of chemicals to identify potential androgen receptor
(AR) agonists or antagonists would cost millions of dollars and take
decades to complete using current validated methods. High-throughput
in vitro screening (HTS) and computational toxicology approaches can
more rapidly and inexpensively identify potential androgen-active
chemicals. We integrated 11 HTS ToxCast/Tox21 in vitro assays into
a computational network model to distinguish true AR pathway activity
from technology-specific assay interference. The in vitro HTS assays
probed perturbations of the AR pathway at multiple points (receptor
binding, coregulator recruitment, gene transcription, and protein
production) and multiple cell types. Confirmatory in vitro antagonist
assay data and cytotoxicity information were used as additional flags
for potential nonspecific activity. Validating such alternative testing
strategies requires high-quality reference data. We compiled 158 putative
androgen-active and -inactive chemicals from a combination of international
test method validation efforts and semiautomated systematic literature
reviews. Detailed in vitro assay information and results were compiled
into a single database using a standardized ontology. Reference chemical
concentrations that activated or inhibited AR pathway activity were
identified to establish a range of potencies with reproducible reference
chemical results. Comparison with existing Tier 1 AR binding data
from the U.S. EPA Endocrine Disruptor Screening Program revealed that
the model identified binders at relevant test concentrations (<100
ÎĽM) and was more sensitive to antagonist activity. The AR pathway
model based on the ToxCast/Tox21 assays had balanced accuracies of
95.2% for agonist (<i>n</i> = 29) and 97.5% for antagonist
(<i>n</i> = 28) reference chemicals. Out of 1855 chemicals
screened in the AR pathway model, 220 chemicals demonstrated AR agonist
or antagonist activity and an additional 174 chemicals were predicted
to have potential weak AR pathway activity
Real-Time Growth Kinetics Measuring Hormone Mimicry for ToxCast Chemicals in T‑47D Human Ductal Carcinoma Cells
High-throughput
screening (HTS) assays capable of profiling thousands of environmentally
relevant chemicals for <i>in vitro</i> biological activity
provide useful information on the potential for disrupting endocrine
pathways. Disruption of the estrogen signaling pathway has been implicated
in a variety of adverse health effects including impaired development,
reproduction, and carcinogenesis. The estrogen-responsive human mammary
ductal carcinoma cell line T-47D was exposed to 1815 ToxCast chemicals
comprising pesticides, industrial chemicals, pharmaceuticals, personal
care products, cosmetics, food ingredients, and other chemicals with
known or suspected human exposure potential. Cell growth kinetics
were evaluated using real-time cell electronic sensing. T-47D cells
were exposed to eight concentrations (0.006–100 μM),
and measurements of cellular impedance were repeatedly recorded for
105 h. Chemical effects were evaluated based on potency (concentration
at which response occurs) and efficacy (extent of response). A linear
growth response was observed in response to prototypical estrogen
receptor agonists (17β-estradiol, genistein, bisphenol A, nonylphenol,
and 4-<i>tert</i>-octylphenol). Several compounds, including
bisphenol A and genistein, induced cell growth comparable in efficacy
to that of 17β-estradiol, but with decreased potency. Progestins,
androgens, and corticosteroids invoked a biphasic growth response
indicative of changes in cell number or cell morphology. Results from
this cell growth assay were compared with results from additional
estrogen receptor (ER) binding and transactivation assays. Chemicals
detected as active in both the cell growth and ER receptor binding
assays demonstrated potencies highly correlated with two ER transactivation
assays (<i>r</i> = 0.72; <i>r</i> = 0.70). While
ER binding assays detected chemicals that were highly potent or efficacious
in the T-47D cell growth and transactivation assays, the binding assays
lacked sensitivity in detecting weakly active compounds. In conclusion,
this cell-based assay rapidly detects chemical effects on T-47D growth
and shows potential, in combination with other HTS assays, to detect
environmentally relevant chemicals with potential estrogenic activity