United States
Environmental Protection
1=1 m m Agency
EPA/690/R-14/015F
Final
8-11-2014
Provisional Peer-Reviewed Toxicity Values for
Styrene-Acrylonitrile (SAN) Trimer
(Various CASRNs)
Superfund Health Risk Technical Support Center
National Center for Environmental Assessment
Office of Research and Development
U.S. Environmental Protection Agency
Cincinnati, OH 45268
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AUTHORS, CONTRIBUTORS, AND REVIEWERS
CHEMICAL MANAGERS
J. Phillip Kaiser, PhD, DABT
National Center for Environmental Assessment, Cincinnati, OH
Dan D. Petersen, PhD, DABT
National Center for Environmental Assessment, Cincinnati, OH
CONTRIBUTOR
Scott C. Wesselkamper, PhD
National Center for Environmental Assessment, Cincinnati, OH
PRIMARY INTERNAL REVIEWERS
Q. Jay Zhao, PhD, MPH, DABT
National Center for Environmental Assessment, Cincinnati, OH
Paul G. Reinhart, PhD, DABT
National Center for Environmental Assessment, Research Triangle Park, NC
This document was externally peer reviewed under contract to
Eastern Research Group, Inc.
110 Hartwell Avenue
Lexington, MA 02421-3136
Questions regarding the contents of this document may be directed to the U.S. EPA Office of
Research and Development's National Center for Environmental Assessment, Superfund Health
Risk Technical Support Center (513-569-7300).
l
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TABLE OF CONTENTS
COMMONLY USED ABBREVIATIONS AND ACRONYMS iii
BACKGROUND 1
DISCLAIMERS 1
QUESTIONS REGARDING PPRTVs 1
INTRODUCTION 2
REVIEW OF POTENTIALLY RELEVANT DATA (NONCANCER AND CANCER) 3
HUMAN STUDIES 17
Oral Exposure 17
Inhalation Exposures 23
ANIMAL STUDIES 23
Oral Exposures 23
Inhalation Exposure 33
OTHER DATA 33
Metabolism Studies 33
Genotoxicity 34
SYNTHESIS OF RESULTS FROM NONCANCER AND CANCER STUDIES 36
DERIVATION 01 PROVISIONAL VALUES 38
DERIVATION OF PROVISIONAL ORAL REFERENCE DOSES 40
Derivation of Subchronic Provisional RfD (Subchronic p-RfD) 40
Derivation of a Chronic Provisional RfD (Chronic p-RfD) 43
DERIVATION OF PROVISIONAL INHALATION REFERENCE CONCENTRATIONS . 47
CANCER WEIGII I -01 -EVIDENCE (WOE) DESCRIPTOR 47
DERIVATION OF PROVISIONAL CANCER POTENCY VALUES 49
Derivation of Provisional Oral Slope Factor (p-OSF) 49
Derivation of Provisional Inhalation Unit Risk (p-IUR) 49
APPENDIX A. FIGURES AND DATA TABLES 50
APPENDIX B. BENCHMARK DOSE CALCULATIONS FOR THE SUBCHRONIC p-RfD
AND CHRONIC p-RfD 73
APPENDIX C. DOSIMETRY CALCULATION EXAMPLES FOR FO DAMS AND F1 PUPS
IN THE 2-WEEK, 13-WEEK, AND 2-YEAR PERINATAL AND POSTNATAL FEED
STUDIES OF SAN TRIMER (NTP, 2012) 88
APPENDIX D. REFERENCES 93
li
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COMMONLY USED ABBREVIATIONS AND ACRONYMS
a2u-g
alpha 2u-globulin
MN
micronuclei
ACGIH
American Conference of Governmental
MNPCE
micronucleated polychromatic
Industrial Hygienists
erythrocyte
AIC
Akaike's information criterion
MOA
mode-of-action
ALD
approximate lethal dosage
MTD
maximum tolerated dose
ALT
alanine aminotransferase
NAG
N-acetyl-P-D-glucosaminidase
AST
aspartate aminotransferase
NCEA
National Center for Environmental
atm
atmosphere
Assessment
ATSDR
Agency for Toxic Substances and
NCI
National Cancer Institute
Disease Registry
NOAEL
no-observed-adverse-effect level
BMD
benchmark dose
NTP
National Toxicology Program
BMDL
benchmark dose lower confidence limit
NZW
New Zealand White (rabbit breed)
BMDS
Benchmark Dose Software
OCT
ornithine carbamoyl transferase
BMR
benchmark response
ORD
Office of Research and Development
BUN
blood urea nitrogen
PBPK
physiologically based pharmacokinetic
BW
body weight
PCNA
proliferating cell nuclear antigen
CA
chromosomal aberration
PND
postnatal day
CAS
Chemical Abstracts Service
POD
point of departure
CASRN
Chemical Abstracts Service Registry
POD[adj]
duration-adjusted POD
Number
QSAR
quantitative structure-activity
CBI
covalent binding index
relationship
CHO
Chinese hamster ovary (cell line cells)
RBC
red blood cell
CL
confidence limit
RDS
replicative DNA synthesis
CNS
central nervous system
RfC
inhalation reference concentration
CPN
chronic progressive nephropathy
RfD
oral reference dose
CYP450
cytochrome P450
RGDR
regional gas dose ratio
DAF
dosimetric adjustment factor
RNA
ribonucleic acid
DEN
diethylnitrosamine
SAR
structure activity relationship
DMSO
dimethylsulfoxide
SCE
sister chromatid exchange
DNA
deoxyribonucleic acid
SD
standard deviation
EPA
Environmental Protection Agency
SDH
sorbitol dehydrogenase
FDA
Food and Drug Administration
SE
standard error
FEV1
forced expiratory volume of 1 second
SGOT
glutamic oxaloacetic transaminase, also
GD
gestation day
known as AST
GDH
glutamate dehydrogenase
SGPT
glutamic pyruvic transaminase, also
GGT
y-glutamyl transferase
known as ALT
GSH
glutathione
SSD
systemic scleroderma
GST
glutathione -S -transferase
TCA
trichloroacetic acid
Hb/g-A
animal blood-gas partition coefficient
TCE
trichloroethylene
Hb/g-H
human blood-gas partition coefficient
TWA
time-weighted average
HEC
human equivalent concentration
UF
uncertainty factor
HED
human equivalent dose
UFa
interspecies uncertainty factor
i.p.
intraperitoneal
UFh
intraspecies uncertainty factor
IRIS
Integrated Risk Information System
UFS
subchronic-to-chronic uncertainty factor
IVF
in vitro fertilization
UFd
database uncertainty factor
LC50
median lethal concentration
U.S.
United States of America
LD50
median lethal dose
WBC
white blood cell
LOAEL
lowest-observed-adverse-effect level
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PROVISIONAL PEER-REVIEWED TOXICITY VALUES FOR
STYRENE ACRYLONITRILE TRIMER
BACKGROUND
A Provisional Peer-Reviewed Toxicity Value (PPRTV) is defined as a toxicity value
derived for use in the Superfund Program. PPRTVs are derived after a review of the relevant
scientific literature using established Agency guidance on human health toxicity value
derivations. All PPRTV assessments receive internal review by a standing panel of National
Center for Environment Assessment (NCEA) scientists and an independent external peer review
by five scientific experts.
The purpose of this document is to provide support for the hazard and dose-response
assessment pertaining to chronic and subchronic exposures to substances of concern, to present
the major conclusions reached in the hazard identification and derivation of the PPRTVs, and to
characterize the overall confidence in these conclusions and toxicity values. It is not intended to
be a comprehensive treatise on the chemical or toxicological nature of this substance.
The PPRTV review process provides needed toxicity values in a quick turnaround
timeframe while maintaining scientific quality. PPRTV assessments are updated approximately
on a 5-year cycle for new data or methodologies that might impact the toxicity values or
characterization of potential for adverse human health effects and are revised as appropriate. It is
important to utilize the PPRTV database (http://hhpprtv.ornl.gov) to obtain the current
information available. When a final Integrated Risk Information System (IRIS) assessment is
made publicly available on the Internet (www.epa.gov/iris). the respective PPRTVs are removed
from the database.
DISCLAIMERS
The PPRTV document provides toxicity values and information about the adverse effects
of the chemical and the evidence on which the value is based, including the strengths and
limitations of the data. All users are advised to review the information provided in this
document to ensure that the PPRTV used is appropriate for the types of exposures and
circumstances at the site in question and the risk management decision that would be supported
by the risk assessment.
Other U.S. Environmental Protection Agency (EPA) programs or external parties who
may choose to use PPRTVs are advised that Superfund resources will not generally be used to
respond to challenges, if any, of PPRTVs used in a context outside of the Superfund program.
QUESTIONS REGARDING PPRTVs
Questions regarding the contents and appropriate use of this PPRTV assessment should
be directed to the U.S. EPA Office of Research and Development's National Center for
Environmental Assessment, Superfund Health Risk Technical Support Center (513-569-7300).
1
SAN Trimer
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INTRODUCTION
Styrene-acrylonitrile Trimer (SAN Trimer) exists as a mixture of isomers of 4-cyano-
1,2,3,4-tetrahydro-a-methyl-l-naphthaleneacetonitrile (THNA; CASRN 57964-39-3) and
4-cyano-l,2,3,4-tetrahydro-l-naphthalene-propionitrile (THNP; CASRN 57964-40-6) and is
formed by the condensation of two moles of acrylonitrile and one mole of styrene. The
SAN Trimer mixture includes the THNA form, consisting of four stereoisomers (c/.s-R-THNA,
30%; c/s-S-THNA, 17%; //vms-R-THNA, 25%; and //vms-S-THNA, 14%), and the THNP form,
consisting of two stereoisomers (c/.v-THNP, 8% and /ram-THNP, 6%) CNTP. 2012) (see
Figure 1). SAN Trimer is a by-product of specific manufacturing processes for polymers of
styrene and acrylonitrile, but it is currently not considered commercially useful CNTP. 2012).
The mixture exists as a viscous, light-brown, opaque liquid (at room temperature). SAN Trimer
was selected for study by a United States Environmental Protection Agency (U.S. EPA)
workgroup that was formed due to reports by the New Jersey Department of Health and Senior
Services (NJ DHSS) regarding a potential link between childhood cancer incidence rates and
previously unknown semivolatile contaminants (now known as SAN Trimer) in groundwater
after dumping of spent process streams from styrene-acrylonitrile polymer manufacturing (NJ
DHSS. 2003a. b, 1997). A table of physicochemical properties for SAN Trimer is provided (see
Table 1).
THNA
(4 Isomers)
CM
THNP
(2 hornets)
Figure 1. Structures of the SAN Trimers (Gargas et al., 2008)
2
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Table 1. Physicochemical Properties of SAN Trimer (Various CASRNs)3
Property (unit)
Value
Boiling point (°C)
ND
Melting point (°C)
ND
Density (g/cm3 at 20°C)
1.101
Vapor pressure (mm Hg at 235°C)
Estimated at 2.5
pH (unitless)
ND
Solubility in water (mg/L)
84.9
Specific gravity (at 20°C)
1.103
Molecular weight (g/mol)
210
aNTP (20121.
ND = no data.
No toxicity values were identified for SAN Trimer.
Literature searches were conducted on sources published from 1900 through
February 2014 for studies relevant to the derivation of provisional toxicity values for
SAN Trimer (various CASRNs). The following databases were searched by chemical name,
synonyms, or CASRN: ACGM, ANEUPL, AT SDR, BIOSIS, Cal/EPA, CCRIS, CDAT,
ChemlDplus, CIS, CRISP, DART, EMIC, EPIDEM, ETICBACK, FEDRIP, GENE-TOX,
HAPAB, HERO, HMTC, HSDB, IARC, INCHEM IPCS, IP A, ITER, IUCLID, LactMed,
NIOSH, NTIS, NTP, OSHA, OPP/RED, PESTAB, PPBIB, PPRTV, PubMed (toxicology
subset), RISKLINE, RTECS, TOXLINE, TRI, U.S. EPA IRIS, U.S. EPA HEAST, U.S. EPA
HEEP, U.S. EPA OW, and U.S. EPA TSCATS/TSCATS2. The following databases were
searched for relevant health information: ACGIH, ATSDR, Cal/EPA, U.S. EPA IRIS, U.S. EPA
HEAST, U.S. EPA HEEP, U.S. EPA OW, U.S. EPA TSCATS/TSCATS2, NIOSH, NTP,
OSHA, and RTECS.
REVIEW OF POTENTIALLY RELEVANT DATA
(NONCANCER AND CANCER)
Tables 2 A and 2B provides an overview of the relevant database for SAN Trimer and
includes all potentially relevant repeat-dose short-term-, subchronic-, and chronic-duration
studies. Principal studies are identified. The phrase "statistical significance," used throughout
the document, indicates ap-value of <0.05 (i.e., the probability that differences in observed data
are not due to chance alone).
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Table 2A. Summary of Potentially Relevant Noncancer Data for SAN Trimer (Various CASRNs)
Category3
Number of
Male/Female,
Strain, Species,
Study Type, Study
Duration
Dosimetryb
Critical Effects
NOAEL
BMDL/
BMCL
LOAEL
Reference
Comments
Notes
Human
1. Oral
Acute0
ND
Short-termd
ND
Long-term6
ND
Chronicf
ND
2. Inhalation
Acute
ND
Short-term
ND
Long-term
ND
Chronic
ND
Animal
1. Oral
Acute
5/5, S-D, rat, gavage,
single administration
then observed for
14 days
0, 250, 500,
or
1,000 mg/kg
Mortality: (sex not specified),
20% at 250 mg/kg, 40% at
500 mg/kg, 90% at 1,000 mg/kg.
NA
NC
NA
Huntingdon
Life
Sciences
(1999a)
LD5o study
NPR
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Table 2A. Summary of Potentially Relevant Noncancer Data for SAN Trimer (Various CASRNs)
Category3
Number of
Male/Female,
Strain, Species,
Study Type, Study
Duration
Dosimetryb
Critical Effects
NOAEL
BMDL/
BMCL
LOAEL
Reference
Comments
Notes
Short-term
6/6, S-D, rat, gavage,
14 days
0, 30, 75, 150,
or 300
mg/kg-day
Increased absolute
(150 mg/kg-day) and relative
liver (>75 mg/kg-day) weights
in males; increased absolute and
relative liver weights at
>150 mg/kg-day in females;
increased absolute and relative
heart weights at >150 mg/kg-day
in females; mortality in both
males and females at
300 mg/kg-day.
30
NC
75
Huntingdon
NPR
Life
Sciences
(1999b)
0/7-8, F344/N, rat,
dietary,
GD 7-delivery
(F0 dams)
0, 18,35,67,
130, or 197
mg/kg-day
Decreased body weight on
GD 14.
130
NDr
197
NTP (2012)
Decreased
body weight
in dams could
be due to
decreased
food
consumption
PR;
gestational
component of
the 2-week
postweaning
NTP (2012)
study.
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Table 2A. Summary of Potentially Relevant Noncancer Data for SAN Trimer (Various CASRNs)
Category3
Number of
Male/Female,
Strain, Species,
Study Type, Study
Duration
Dosimetryb
Critical Effects
NOAEL
BMDL/
BMCL
LOAEL
Reference
Comments
Notes
Short-term
0/6-8, F344/N, rat,
dietary, PNDs 1-20
(F0 dams)
10/10 Flpups
0, 40, 85, 166,
325, or 634
mg/kg-day
Decreased body weight in
F0 dams on PNDs 1-20 and
F1 pups on PND 20 (males and
females).
F0 dams:
325
F1 pups
(males and
females):
166
NDr
634
325
NTP (2012)
Decreased
body weight
in dams could
be due to
decreased
food
consumption
PR;
lactational
component of
the 2-week
postweaning
NTP (2012)
study. Doses
forFl pups
were
calculated
assuming that
they received
100% of the
dose given to
dams.
10/10, F344/N, rat,
dietary, 2 weeks
postweaning
(F1 rats from dams
exposed
GD 7-PND 20)
0, 50, 90, 175,
270, or 410
mg/kg-day
(F1 males)
and 0, 45, 90,
185, 295, or
430
mg/kg-day
(F1 females)
Decreased absolute and relative
right testis and thymus weights
at >90 mg/kg-day in males;
increased relative liver weight at
>175 mg/kg-day in males and at
>185 mg/kg-day in females;
other relative organ weight
changes at doses
>270 mg/kg-day in males and
>295 mg/kg-day in females also
observed; decreased body
weight at >175 mg/kg-day in
males and >295 mg/kg-day in
females; increased brain
cellularity in females at
430 mg/kg-day.
50
NC
90
NTP (2012)
PR
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Table 2A. Summary of Potentially Relevant Noncancer Data for SAN Trimer (Various CASRNs)
Category3
Number of
Male/Female,
Strain, Species,
Study Type, Study
Duration
Dosimetryb
Critical Effects
NOAEL
BMDL/
BMCL
LOAEL
Reference
Comments
Notes
Subchronic
0/8-9, F344/N, rat,
dietary,
GD 7-delivery
(F0 dams)
0,6.9, 14, 28,
55, or 110
mg/kg-day
Decreased body weight on
GD 20.
55
NDr
110
NTP (2012)
PR;
gestational
component of
the 13-week
postweaning
NTP (2012)
study.
0/4-8, F344/N, rat,
dietary, PNDs 1-20
(F0 dams)
10/10 Flpups
0, 17,33,66,
132, or 264
mg/kg-day
No effects observed in F0 dams
orFl pups.
264
NDr
NDr
NTP (2012)
PR;
lactational
component of
the 13-week
postweaning
NTP (2012)
study.
10/10, F344/N, rat,
dietary, 13 weeks
postweaning
(F1 rats from dams
exposed
GD 7-PND 20)
0,10,20,40,
80, or 150
mg/kg-day
(F1 males
and females)
Increased absolute and
relative liver weights in males
at >40 mg/kg-day; increased
absolute and relative heart
weights in males at
>10 mg/kg-day and relative
heart weight in females at
>80 mg/kg-day; other absolute
and relative organ weight
changes at doses
>40 mg/kg-day in males and
females also observed;
decreased body weight at
150 mg/kg-day in males.
NDr
10
(based on
increased
absolute liver
weight in
males)
10
NTP
(2012)
PR, PS
7
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Table 2A. Summary of Potentially Relevant Noncancer Data for SAN Trimer (Various CASRNs)
Category3
Number of
Male/Female,
Strain, Species,
Study Type, Study
Duration
Dosimetryb
Critical Effects
NOAEL
BMDL/
BMCL
LOAEL
Reference
Comments
Notes
Chronic
0/41-42, F344/N, rat,
dietary,
GD 7-delivery
(F0 dams)
0, 28, 55, or
110
mg/kg-day
No effects observed.
110
NDr
NDr
NTP (2012)
PR;
gestational
component of
the 2-year
postweaning
NTP (2012)
study.
0/24-27, F344/N, rat,
dietary, PNDs 1-20
(F0 dams)
50/50 Flpups
0, 66, 132, or
264
mg/kg-day
No effects observed in F0 dams
orFl pups.
264
NDr
NDr
NTP (2012)
PR;
lactational
component of
the 2-year
postweaning
NTP (2012)
study.
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Table 2A. Summary of Potentially Relevant Noncancer Data for SAN Trimer (Various CASRNs)
Category3
Number of
Male/Female,
Strain, Species,
Study Type, Study
Duration
Dosimetryb
Critical Effects
NOAEL
BMDL/
BMCL
LOAEL
Reference
Comments
Notes
Chronic
50/50, F344/N, rat,
dietary, 2 years
postweaning
(F1 rats from dams
exposed
GD 7-PND 20)
0,20,40, or
75
mg/kg-day
(F1 males)
and 0,20,40,
or 85
mg/kg-day
(F1 females)
Increased incidences of mixed
cell foci in liver and
hyperplasia of the transitional
epithelium of the bladder at
85 mg/kg-day in females;
spinal root degeneration,
chronic active liver
inflammation, eosinophilic
angiectasis, and bone marrow
hyperplasia at 75 mg/kg-day in
males; sciatic nerve
degeneration, bone marrow
hyperplasia, and
granulomatosis inflammation
at >40 mg/kg-day in females;
mixed cell foci in liver at
>20 mg/kg-day in males.
NA
3.2
(based on
increased
incidence of
chronic active
liver
inflammation
in male rats)
20
NTP
(2012)
PR, PS
Developmental
ND
Reproductive
ND
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Table 2A. Summary of Potentially Relevant Noncancer Data for SAN Trimer (Various CASRNs)
Category3
Number of
Male/Female,
Strain, Species,
Study Type, Study
Duration
Dosimetryb
Critical Effects
NOAEL
BMDL/
BMCL
LOAEL
Reference
Comments
Notes
2. Inhalation
Subchronic
ND
Chronic
ND
Developmental
ND
Reproductive
ND
aThe duration classification for the studies from NTP (20121 is based on the time that F1 rats were directly treated with SAN Trimer in the diet.
bDosimetry: NOAEL, BMDL/BMCL, and LOAEL values are converted to an adjusted daily dose (ADD in mg/kg-day) for oral noncancer effects and a human equivalent
dose (HED in mg/kg-day) for oral carcinogenic effects. All long-term exposure values (4 weeks and longer) are converted from a discontinuous to a continuous exposure.
0 Acute = exposure for <24 hours (U.S. EPA. 2002).
dShort-term = repeated exposure for >24 hours <30 days (U.S. EPA. 2002).
"Long-term = repeated exposure for >30 days <10% lifespan (based on 70-year typical life span) (U.S. EPA. 2002).
fChronic = repeated exposure for >10% life span (U.S. EPA. 2002).
GD = Gestation Day; ND = no data; NA = not applicable; NDr = not determined; NC = not calculated; PR = peer-reviewed; NPR = non-peer-reviewed; PND = Postnatal
Day; PS = principal study; S-D = Sprague-Dawley.
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Table 2B. Summary of Potentially Relevant Cancer Data for SAN Trimer (Various CASRNs)
Category"
Number of
Male/Female,
Strain, Species,
Study Type, Study
Duration
Dosimetryb
Critical Effects
NOAEL
BMDL/
BMCL
LOAEL
Reference
Comments
Notes
Human
1. Oral
Acute0
ND
Short-termd
ND
Long-term6
ND
Chronicf
ND
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Table 2B. Summary of Potentially Relevant Cancer Data for SAN Trimer (Various CASRNs)
Category"
Number of
Male/Female,
Strain, Species,
Study Type, Study
Duration
Dosimetryb
Critical Effects
NOAEL
BMDL/
BMCL
LOAEL
Reference
Comments
Notes
Exposure
duration cannot
be determined
Evaluation of
childhood cancer
statistics from 1979
to 1995 for Ocean
County, Dover
Township, and the
Toms River section
of Dover
Township, NJ
NDr
Ocean County: Increased
incidences of sympathetic
nervous system cancers (i.e.,
neuroblastomas) particularly
in males <5 years old and
brain/CNS astrocytomas
(0-19 years of age and both
sexes).
Dover Township: Increased
incidences of total childhood
cancer (particularly in females
<5 years old), leukemia, and
acute lymphocytic leukemia
(both in females at all ages).
Toms River section of Dover
Township: Increased
incidences of total childhood
cancer particularly in females
<5 years old, brain/CNS
cancer (both sexes),
brain/CNS astrocytomas (both
sexes), leukemia (females),
and acute lymphocytic
leukemia (females) in children
<5 years old.
NDr
NC
NDr
NJ DHSS
(1997);
ATSDR
(1997)
Epidemiologic
study on small
population to
assess
standardized
incidence
ratios for
childhood
cancers.
NPR; the drinking
water supply was
contaminated with
SAN Trimer and
other chemicals
with known
carcinogenic
potential, including
trichloroethylene
and
tetrachloroethylene,
as discussed in NJ
DHSS (2003a) and
in a drinking water
quality analysis (NJ
DHSS. 2001).
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Table 2B. Summary of Potentially Relevant Cancer Data for SAN Trimer (Various CASRNs)
Category"
Number of
Male/Female,
Strain, Species,
Study Type, Study
Duration
Dosimetryb
Critical Effects
NOAEL
BMDL/
BMCL
LOAEL
Reference
Comments
Notes
Exposure
duration cannot
be determined
Case-control design
evaluating residents
from Dover
Township and the
Toms River section
of Dover
Township, NJ from
1979 to 1996:
interview
(40 incident
cases/159 controls);
birth records
(48 cases/480
controls)
NDr
Increased odds ratios (OR) for
prenatal exposure to
contaminated well water and
leukemia in females <19 years
old.
NDr
NC
NDr
NJ DHSS
(2003a)
Epidemiologic
study on small
population;
supports
increased
incidence of
leukemia in
children.
PR; the drinking
water supply was
contaminated with
SAN Trimer and
other chemicals
with known
carcinogenic
potential, including
trichloroethylene
and
tetrachloroethylene,
as discussed in NJ
DHSS C2003aN) and
in a drinking water
quality analysis (NJ
DHSS. 2001).
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Table 2B. Summary of Potentially Relevant Cancer Data for SAN Trimer (Various CASRNs)
Category"
Number of
Male/Female,
Strain, Species,
Study Type, Study
Duration
Dosimetryb
Critical Effects
NOAEL
BMDL/
BMCL
LOAEL
Reference
Comments
Notes
Exposure
duration cannot
be determined
Evaluation of
childhood cancer
statistics from 1996
to 2000 for Dover
Township and the
Toms River section
of Dover
Township, NJ
NDr
No statistically significant
increase in childhood cancer
was observed.
NDr
NC
NDr
NJ DHSS
f2003b)
Epidemiologic
study on small
population to
assess
standardized
incidence
ratios for
childhood
cancers
NPR; the drinking
water supply was
contaminated with
SAN Trimer and
other chemicals
with known
carcinogenic
potential, including
trichloroethylene
and
tetrachloroethylene,
as discussed in NJ
DHSS C2003aN) and
in a drinking water
quality analysis (NJ
DHSS. 2001).
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Table 2B. Summary of Potentially Relevant Cancer Data for SAN Trimer (Various CASRNs)
Category"
Number of
Male/Female,
Strain, Species,
Study Type, Study
Duration
Dosimetryb
Critical Effects
NOAEL
BMDL/
BMCL
LOAEL
Reference
Comments
Notes
Exposure
duration cannot
be determined
Evaluation of
childhood cancer
statistics from 2001
to 2005 for Toms
River Township
(formerly Dover
Township) and the
sub-Township area
(formerly the Toms
River section of
Dover Township,
NJ)
NDr
Increased incidence in soft
tissue sarcomas in females
<19 years old (2004-2005) in
both the Toms River
Township and sub-Township
area.
NDr
NC
NDr
NJ DHSS
(2008)
Epidemiologic
study on small
population;
standardized
incidence
ratios (SIRs)
for childhood
cancers
previously
elevated at an
earlier time
interval
(1979-1995)
returned to
background
levels.
NPR; the drinking
water supply was
contaminated with
SAN Trimer and
other chemicals
with known
carcinogenic
potential, including
trichloroethylene
and
tetrachloroethylene,
as discussed in NJ
DHSS C2003aN) and
in a drinking water
quality analysis (NJ
DHSS. 2001).
2. Inhalation
ND
Animal
1. Oral
ND
Carcinogenic
50/50, F344/N, rat,
dietary, 2 year
postweaning
(F1 rats from dams
exposed
GD 7-PND 20)
HEDs: 0, 5.6,
11, or
20 mg/kg-day
(F1 males)
and 0, 5.0,
9.8, or 20
mg/kg-day
(F1 females)
No statistically significant
increases in tumor incidence
observed.
NA
NC
NA
NTP (2012)
PR
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Table 2B. Summary of Potentially Relevant Cancer Data for SAN Trimer (Various CASRNs)
Category"
Number of
Male/Female,
Strain, Species,
Study Type, Study
Duration
Dosimetryb
Critical Effects
NOAEL
BMDL/
BMCL
LOAEL
Reference
Comments
Notes
2. Inhalation
Carcinogenic
ND
aThe duration classification for the studies from NTP (20121 is based on the time that F1 rats were directly treated with SAN Trimer in the diet.
bDosimetry: NOAEL, BMDL/BMCL, and LOAEL values are converted to an adjusted daily dose (ADD in mg/kg-day) for oral noncancer effects and a human equivalent
dose (HED in mg/kg-day) for oral carcinogenic effects. All long-term exposure values (4 weeks and longer) are converted from a discontinuous to a continuous exposure.
0 Acute = exposure for <24 hours (U.S. EPA. 2002).
dShort-term = repeated exposure for >24 hours <30 days (U.S. EPA. 2002).
eLong-term = repeated exposure for >30 days <10% lifespan (based on 70-year typical life span) (U.S. EPA. 2002).
fChronic = repeated exposure for >10% life span (U.S. EPA. 2002).
GD = Gestation Day; ND = no data; NA = not applicable; NDr = not determined; NC = not calculated; PR = peer-reviewed; PND = Postnatal Day.
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HUMAN STUDIES
Oral Exposure
Four human studies on oral exposure to drinking water containing various chemicals,
including SAN Trimer, are publicly available; one was a case-control study and the other three
were ecological in nature because they examined town-level incidence and exposure data. These
studies evaluated childhood cancer incidence in Ocean County, Dover Township, and the Toms
River section of Dover Township, New Jersey, as discussed in NJ DHSS (2003 a) and in a
drinking water quality analysis (NJ DHSS. 2001). The three ecological studies were conducted
covering the years 1979-1995 (NJDHSS. 1997). and then extended to cover subsequent 5-year
intervals from 1996-2000 (NJDHSS. 2003b) and 2001-2005 (NJDHSS. 2008). The
case-control study analyzed the same population from 1979-1996 (NJ DHSS. 2003a). These
studies are summarized chronologically below. It should be noted that due to the existence of
other contaminants in drinking water from the well fields investigated, the conclusions that can
be drawn from the epidemiologic studies regarding the toxicity of SAN Trimer are relatively
limited. Some of the chemicals that were present in the drinking water (e.g., trichloroethylene
and tetrachloroethylene) are known to have carcinogenic potential. Finally, even if SAN Trimer
were determined to have carcinogenic potential, the dosimetry necessary to establish a
dose-response relationship is absent because SAN Trimer concentrations in the drinking water
were not measured as part of these studies.
NJDHSS (1997)
Due to residential concern about cancer levels in the Toms River section of Dover
Township, New Jersey, the NJ DHSS in collaboration with the ATSDR reviewed cancer
incidence data from the New Jersey State Cancer Registry for children living in Ocean County,
Dover Township, and the Toms River section of Dover Township in 1997 (see Figure A-l) in a
non-peer-reviewed study (NJDHSS. 1997). The purpose of the report was to verify results and
extend the analysis of the time period investigated in a previous cancer analysis done by the New
Jersey Department of Health (NJDOH) and the ATSDR (NJDOH. 1996) (not publicly available)
and to determine which cancer types were increased. Residents under the age of 20 who were
diagnosed with selected childhood cancer types (i.e., leukemia, brain and central nervous system,
lymphomas, soft tissue sarcomas, sympathetic nervous system, bone, and kidney cancer) during
a 17-year period (1979-1995) were included in the analysis. To quantitatively analyze
childhood cancer incidence, the study authors calculated standardized incidence ratios (SIR) for
each area by dividing the reported number of new cancers by the expected number using the
State of New Jersey as a comparison population. An SIR greater than one indicates that more
cancer cases than expected were observed. The study authors conducted statistical evaluations
using a 95% confidence interval (CI) to determine whether differences in an SIR were due to
chance alone. A 95% CI that includes the null value of 1 indicates results that are not
statistically significant at ap-walue of 0.05.
In Ocean County, there was no increase in total childhood cancer incidence compared to
the expected number. However, there was a statistically significant increase in sympathetic
nervous system cancers (i.e., neuroblastomas), particularly in males (SIR = 2.21, CI = 1.29-3.54)
under the age of 5 (see Table A-l) (NJ DHSS. 1997). There was also a statistically significant
increase in brain/CNS astrocytomas when all age, race, and sex groups were pooled (SIR = 1.46,
CI = 1.02-2.03). The study authors also reported a slight increase (SIR = 1.77, CI = 0.88-3.17)
in astrocytomas between the observed and expected incidences in both sexes for children under
age 5 (not statistically significant). Similar findings for astrocytomas were also found in Dover
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Township (SIR = 1.91, CI = 0.21-6.91) in both sexes for children under age 5. Additional
findings in Dover Township included: (1) a statistically significant increase for total childhood
cancer incidence, especially in females under the age of 5 (SIR = 1.90, CI = 1.09-3.09; see
Table A-2), (2) a statistically significant increase in the incidence of leukemia in females from
ages 0-19 (SIR = 1.99, CI = 1.06-3.40) and 0-4 (SIR = 2.65, CI = 1.06-5.45), and (3) a
statistically significant increase in the incidence of acute lymphocytic leukemia in females from
ages 0-19 (SIR = 2.59, CI = 1.34-4.53) and ages 0-4 (SIR = 3.27, CI = 1.31-6.73). In the
Toms River section of Dover Township, the total childhood cancer incidence was statistically
significantly elevated, most notably in females under the age of 5 (SIR = 6.17, CI = 2.95-11.34;
see Table A-3). Additionally, the incidence of brain and central nervous system cancer was
statistically significantly higher in children of both sexes (SIR = 7.04, CI = 1.89-18.03) and
females only (SIR = 11.6, CI = 2.33-33.88), both under the age of 5. Astrocytomas were
increased in children of both sexes (SIR = 9.47, CI = 1.06-34.19) below the age of 5, and
elevated leukemia incidence (SIR = 7.84, CI = 2.11-20.06) and acute lymphocytic leukemia
(SIR = 9.68, CI = 2.60-24.78) were both observed in females. A time-trend analysis for the
State of New Jersey, Ocean County, Dover Township, and the Toms River section of Dover
Township provided limited evidence of differences in total cancer rates between the State and
Ocean County throughout the period of analysis. During the mid to late 1980s, increased
incidences for all combined cancers, leukemia, and brain/central nervous system cancers were
reported for Dover Township. From 1988-1990, there was a notable increase in cancer
incidence for the Toms River section of Dover Township.
The study authors concluded that these data support the findings of increased incidence
of childhood brain cancer in the Dover Township and the Toms River section of Dover
Township previously reported by an earlier analysis (NJDOH. 1996) (not publicly available).
Most notably, statistically significantly increased incidences of leukemia and brain/central
nervous system cancer were observed, particularly in females under the age of 5.
NJDHSS (2003a)
In response to their previous findings (NJDHSS. 1997). the NJ DHSS (2003a) conducted
a case-control epidemiologic analysis of childhood cancers in Dover Township and the Toms
River section of Dover Township on individuals exposed during the period between 1979 and
1996. During the study, water distributions and air plumes originating from the Ciba-Geigy
Superfund site and the Reich Farm Superfund site were modeled. Individuals were potentially
exposed to contaminants through air and by drinking water provided to the community from a
well system. The United Water Toms River (UWTR) Parkway and the UWTR Holly Street well
fields served this community, and there was also potential contamination of private wells due to
releases from both Superfund sites. Early in the course of evaluating sources of contamination
attributable to the Reich Farm site, Richardson et al. (1999) performed a chromatographic
analysis of drinking water samples from the UWTR Parkway well field in 1996. The study
authors determined that three to four of the highest chromatographic peaks were unknown
compounds later determined to be isomers of SAN Trimer (ATSDR. 1998).
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There were two components to the case-control epidemiologic study. First, four controls
were identified for each case, matched by age, sex, and residence yielding 40 cases and
159 controls in the Interview Study. Spatial location and other information were gathered at
diagnosis ± 1 year. In the second part of the study, 10 controls were identified per case, yielding
48 cases and 480 controls in the Birth Records Study.
The relative risk (odds ratio) of childhood cancers was calculated using conditional
logistic regression to evaluate the degree to which exposure factors were associated with disease.
An odds ratio (OR) greater than one means that the exposure factor was more common in cases
than controls. ORs were calculated for two groups: children diagnosed before age 5 and children
diagnosed before age 20. For the Interview Study, ORs were calculated for leukemia and
nervous system cancers, leukemia alone, all nervous system cancers, brain and central nervous
systems cancers, and all other cancers. The Birth Records Study was meant to address people
who were exposed to contaminants in drinking water from the UWTR well fields but had since
moved away. Cancer incidences in children that moved out of the study area prior to diagnosis
were identified using cancer registries in ten states, with roughly 70% of these children moving
to states with cancer registries. A source of confounding in this study is that the groundwater
contained several contaminants in addition to SAN Trimer, including trichloroethylene and
tetrachloroethylene that are both known to demonstrate carcinogenic potential. Specifically,
trichloroethylene and tetrachloroethylene have been classified by IRIS as carcinogenic to
humans and likely to be carcinogenic in humans, respectively (IRIS. 2012. 2011). and both
chemicals have been classified by the International Agency for Research on Cancer (IARC) as
probably carcinogenic to humans (IARC. 1997). There were also additional sources of air
emissions and lifestyle factors that potentially confound the analysis; these factors were only
partly addressed through matching of controls. Furthermore, the results from the case-control
study should be interpreted with caution based on the study authors' inference that "Due to the
relatively small number of study subjects, the analyses are sensitive to random fluctuations in
numbers, which can result in substantial imprecision in the odds ratios (reflected by wide
confidence intervals)."
Three exposure categories (high, medium, and low) were created based on the average
percent of public water delivered from each of the aforementioned well fields. When prenatal,
time-specific, UWTR Parkway well field, high-level exposures were analyzed separately by sex,
the study authors found an association (OR = 5.0, 95% CI = 0.8-31) for prenatal exposures and
leukemia in females 0-19 years old. For females with leukemia, the prenatal, time-specific,
UWTR Parkway well field, high-exposure category was also significantly associated (OR = 6.0,
95%) CI = 1.1-32). Because of uncertainty as to when the contaminants reached the wells, the
exposure period was broken into several windows. The new windows were 1978-1996,
1979-1996, 1981-1996, 1983-1996, 1984-1996, 1985-1996, and 1986-1996. For the period
of 1984-1996, the OR for exposure to UWTR Parkway well water and the development of
leukemia in females was 15 (95% CI = 0.8-274). The Birth Records Study showed no pattern or
consistency of association with childhood cancers. Exposure to unadjusted UWTR Holly Street
well field water or various time-specific periods of exposure to Holly Street well field water did
not appear to be associated with any childhood cancers in either the Birth Records or the
Interview Studies. Very few children lived in residences with private groundwater wells in any
of the groundwater regions, and no association was seen between leukemia and brain and central
nervous system cancer groupings in either the Interview or the Birth Record Studies.
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There was also an association with Ciba-Geigy air emissions that appeared only with
leukemia in females from ages 0-4 in high- (75th percentile), medium- (50th percentile), or
low-exposure categories. The OR = 19 for the high-exposure category (95% CI = 0.9-397); for
the medium-exposure category the OR = 5.2 (95% CI = 0.5-57) relative to the lowest exposure
category. No associations were found in males for overall leukemia or central nervous system
cancers. There were also no associations with air emissions from the Oyster Creek Nuclear
plant. Associations were seen for proximity to the Ciba-Geigy pipeline and leukemia for both
sexes combined and females only during the prenatal and postnatal period (OR range = 2.3-14).
For the Birth Records Study, only the prenatal period was evaluated, and it was assumed that the
residence at birth was also the residence throughout pregnancy. As in the Interview Study,
elevated ORs were found in the Birth Records Study for the high- and medium-exposure
categories of Ciba-Geigy ambient air emissions for prenatal exposure in females diagnosed with
leukemia prior to age 5 (high-exposure category OR = 7.8, 95% CI = 0.8-77; medium-exposure
category OR = 2.0, 95% CI = 0.1-35).
Exposure to UWTR Parkway well field water containing SAN Trimer was associated
with a statistically significant increase in childhood leukemia. No association was seen with
other public water supply fields. There was also an association with Ciba-Geigy air emissions
that appeared only with leukemia in females. Because of the small number of cases, the NJ
DHSS (2003a) study lacks the power to detect statistically significant findings (i.e., most
95% CIs overlap 1.0).
A groundwater treatment system (i.e., air stripper) had been installed at the Parkway well
field in the mid-1980s to remove the known contaminants but not SAN Trimer; the treated water
was used for drinking. In November 1996, wells contaminated with SAN Trimer were removed
as sources of drinking water. Beginning in May 1997, filtration systems began to be installed on
drinking water wells to specifically remove SAN Trimer contamination.
NJDHSS (2003b)
In response to a public comment draft of the report titled Case-control Study of
Childhood Cancers in Dover Township (Ocean County), New Jersey (NJ DHSS. 2003a). the
NJ DHSS and ATSDR analyzed five further years (1996-2000) of childhood cancer incidence
data in Dover Township and the Toms River section of Dover Township to identify any
differences in incidence and time trends of childhood cancer (NJ DHSS. 2003b). In this
non-peer-reviewed report, no statistically significant increases in the incidences of childhood
cancers in either Dover Township or the Toms River section of Dover Township were observed
during the 5-year period. Furthermore, the study authors stated that SIRs for childhood cancer
incidences in Dover Township and the Toms River section of Dover Township appeared to be
decreasing. A time-trend analysis for the 22-year period (1979-2000) determined that the
incidence of all combined childhood cancers for the State of New Jersey increased slightly
through the period but then dropped in the late 1990s. The incidence rates for Dover Township
increased compared to those of the State from 1985-1995 and in the latter part of the 1990s. The
childhood cancer incidence rates for the Toms River section of Dover Township were the highest
and the most variable during the late 1980s. The study authors concluded that these results
should be interpreted with caution due to the small number of cases that were diagnosed during
the most recent 5-year period (1996-2000) (NJ DHSS. 2003b).
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NJDHSS (2008)
In an additional non-peer-reviewed analysis (NJ DHSS. 2008). the cancer registry data
from the (NJDHSS. 1997) study were updated for the period from 2001-2005, thus extending
the period of analysis for childhood cancers from 1979-2005. During the 5-year interval from
2001-2005, there were 26 additional cancers among children of Toms River Township (formerly
Dover Township) and five in the sub-Township (formerly the Toms River section of Dover
Township). The most frequently diagnosed cancers were brain/CNS, leukemias, and soft-tissue
sarcomas (which were statistically significantly increased from 2004-2005 in females in both the
Toms River Township [SIR = 5.5; 95% CI = 1.5-14] and the sub-Township area [SIR = 21;
95% CI = 4.2-62]). Compared to the expected incidences based on State levels, childhood
cancer incidence in females was not significantly increased (as measured by SIRs), while
leukemia incidence was lower than expected. The total cancer incidence in males was similar to
background levels. Trend analysis for the 1979-2005 period showed increased total childhood
cancer, brain/CNS cancer, and leukemia incidence from the mid 1980s to mid 1990s for female
children in Toms River Township.
In conclusion, the initial epidemiologic study reported statistically significant increases in
childhood cancer incidences from 1979-1995 (NJDHSS. 1997). In the case-control study (NJ
DHSS. 2003a). the study authors observed a statistically significant association for leukemia in
females (0-19 years of age) and exposure to drinking water from well fields containing multiple
chemicals, including SAN Trimer, some of which are known to demonstrate carcinogenic
potential (e.g., trichloroethylene and tetrachloroethylene). The results from the case-control
study should be interpreted with caution because of the small number of study subjects as
described above. Following cleanup and removal of SAN Trimer along with the other chemicals
(e.g., trichloroethylene and tetrachloroethylene) from the drinking water, total childhood cancer
and brain/CNS cancer incidence have returned to background levels, and fewer than expected
cases of leukemia have been observed (see Figures 2 and 3 and Tables A-2 and A-3). However,
due to the existence of other contaminants in drinking water from the well fields investigated, the
conclusions that can be drawn from the epidemiologic studies are relatively limited.
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1.9
1.7
1.5
1.3 —«
1.1
0.9
0.7 1
Dover/Toms River Township
SIRs
All Types, 0-19, M&F All Types, 0-19 F All Types, 0-4 F
1979-1995 ¦ 1996-2000 2001-2005
Figure 2. Standardized Incidence Ratios (SIRs) of Cancer (All Types) in Three Populations
during Three Time Intervals from 1979-2005
Toms River Section of Dover Township
SIRs
Cancer All Types
1979-1995 ¦ 1996-2000 2001-2005
Figure 3. Standardized Incidence Ratios (SIRs) of Cancer (All Types) in Males and
Females (0-19 years) during Three Time Intervals from 1979-2005
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Inhalation Exposures
No studies have been identified.
ANIMAL STUDIES
Oral Exposures
The effects of oral exposure of rats to SAN Trimer have been evaluated in one acute
study (Huntingdon Life Sciences. 1999a). two short-term-duration studies Huntingdon Life
Sciences (1999b); NTP (2012). one sub chronic-duration study (NTP. 2012). and one
chronic-duration study (NTP. 2012). No developmental or reproductive toxicity studies were
identified.
Acute Studies
Huntingdon Life Sciences (1999a)
In a non-peer-reviewed Good Laboratory Practice (GLP) study conducted by Huntingdon
Life Sciences (1999a) for the Union Carbide Corporation that was submitted to the U.S. EPA,
male and female S-D rats (5/sex/dose) were treated with SAN Trimer (98% purity) in corn oil by
a single gavage administration at doses of 0, 250, 500, or 1,000 mg/kg. In determination of the
LD50, there were 2/10 deaths at the 250 mg/kg dose, 4/10 deaths at the 500 mg/kg dose, and
9/10 deaths at the 1,000 mg/kg dose. All animals that died did so within 72 hours, and the
surviving rats were observed for 14 days. The study authors concluded that the LD50 was
440 mg/kg for males and 590 mg/kg for females at both 72 hours and 14 days.
Short-Term-Duration Studies
Huntingdon Life Sciences (1999b)
In a non-peer-reviewed report, Huntingdon Life Sciences (1999b) conducted a 14-day
study for the Union Carbide Corporation in which Sprague-Dawley (S-D) rats (6/sex/dose) were
treated daily with 0, 30, 75, 150, or 300 mg/kg-day SAN Trimer (purity 98%) by gavage in corn
oil. At 300 mg/kg-day all the males and 5/6 females died. Health effects evaluated included
standard hematology (hemoglobin concentration, hematocrit, erythrocyte count, platelet count,
mean corpuscular volume, mean corpuscular hemoglobin, total leukocyte count, differential
leukocyte count, absolute lymphocyte count, absolute segmented neutrophil count, prothrombin
time, activated partial thromboplastin time, and reticulocyte count), standard clinical chemistry
(aspartate aminotransferase, alanine aminotransferase, alkaline phosphatase, lactate
dehydrogenase, sorbitol dehydrogenase, blood urea nitrogen, creatinine, glucose, creatine kinase,
total protein, albumin, globulin, albumin/globulin ratio, total bilirubin, direct bilirubin, indirect
bilirubin, sodium, potassium, chloride, calcium, inorganic phosphorus, gamma-glutamyl
transferase), urinalysis (specific gravity, appearance, osmolality, protein, glucose, ketones, occult
blood, pH, bilirubin, urobilinogen, creatinine, and /V-acetyl-B-D-glucosaminidase), body
weights, food consumption, and organ weights (testes, ovaries, heart, liver, and kidney). In
addition, adrenal glands, aorta (thoracic), bone (sternum/femur with articular surface), bone
marrow (sternum/femur), brain (medulla/pons, cerebrum, and cerebellum), esophagus, eyes (with
optic nerve), heart, kidneys, large intestine (cecum, colon, rectum), lacrimal glands, liver, lungs
(with mainstem bronchi), lymph nodes (mesenteric), mammary glands, muscle (Biceps femoris),
nerve (sciatic/tibial/sural), ovaries, pancreas, pituitary, prostate, salivary gland (submandibular),
seminal vesicles, skin, small intestine (duodenum, ileum, and jejunum), spinal cord (cervical,
thoracic, lumbar), spleen, stomach, testes with epididymides, thymic region, thyroid (with
parathyroid), trachea urinary bladder, uterus (body/horns with cervix), and Zymbal's gland.
Macroscopic lesions were preserved and examined microscopically.
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Food consumption was statistically significantly decreased at 150 mg/kg-day in males
after one week of treatment. Among the hematologic endpoints there were statistically
significant (approximately 10%) decreases in hemoglobin, hematocrit, and red blood cells at
150 mg/kg-day in females, while in the one surviving female treated with 300 mg/kg-day, these
same parameters showed a decrease of 20-30%. There was a statistically significant decrease
(approximately 20%) in mean activated partial thromboplastin time (a measurement of the time it
takes for blood to clot) in the 150 mg/kg-day males compared to control values. The mean
activated partial thromboplastin time for the single surviving female treated with 300 mg/kg-day
was also approximately 20% lower than the control mean. Globulin was statistically
significantly decreased at 30, 75, and 150 mg/kg-day in females, but the study authors did not
consider this biologically significant due to the "absence of any evidence of systemic toxic
changes at the lower doses." No significant findings were observed in the urinalysis. Relative
liver weight was statistically significantly increased in males at 150 mg/kg-day. Absolute and
relative liver weights were statistically significantly increased at 150 mg/kg-day in females.
Mean absolute and relative heart weights were also statistically significantly increased at
150 mg/kg-day in females. Other organ weights were similar to those of the control rats. The
histopathological analysis showed that 4/6 males had vacuolation of periacinar hepatocytes and
3/6 females had periacinar hypertrophy of the liver in the 300 mg/kg-day-dose group. In
addition, 4/6 males in the 300 mg/kg-day-dose group had vacuolation of cortical tubular
epithelial cells in the kidney, and the study authors indicated there was no evidence of
alpha 2u-globulin involvement (although it is unclear if immunostaining was performed).
In summary, a number of effects of SAN Trimer treatment were observed in this
short-term-duration study. The study authors reported dose-related effects indicative of toxicity
including decreased hematocrit and erythrocyte counts, decreased activated partial
thromboplastin time, and elevated liver and heart weights. A LOAEL of 75 mg/kg-day with a
corresponding NOAEL of 30 mg/kg-day is identified for this study based on increased relative
liver weight (>10% change compared to control values) in male rats.
NTP Studies
In a study conducted by Batelle Columbus Operations (Columbus, OH) for the NTP
(2012). male and female F344/N rats were treated with SAN Trimer in feed in perinatal and
postnatal studies for 7 weeks, 18 weeks, or 2 years. The results of these studies are also
published in a peer-reviewed manuscript by Behl et al. (2013). SAN Trimer (Batch 3,
96%) purity) was used in all of the NTP studies. As indicated in the NTP (2012) study, "Animals
in the 7-week, 18-week, and 2-year studies were exposed to the test chemical for 2 weeks during
their in utero development, for 3 weeks through their mother's milk plus consumption of dosed
feed, and for 2 weeks, 13 weeks, and 2 years after weaning through dosed feed." The durations
of these studies are classified for this PPRTV assessment based on the time that F1 rats were
directly treated with SAN Trimer in the diet (i.e., 2 weeks, 13 weeks, and 2 years) as depicted in
Figure 4.
For the 2-week, 13-week, and 2-year studies, male and female F344/N rats (30 males and
60 females for the 2- and 13-week studies, 100 males and 200 females for the 2-year study) were
mated during a 7-day breeding period (9 days for the 2-year study). During breeding, Gestation
Day (GD) 1 was designated upon the presence of a vaginal plug or the identification of sperm in
vaginal lavage fluid. Beginning on GD 7, the study authors administered dietary doses of
SAN Trimer in NIH-07 feed to groups of pregnant females (F0) through delivery (dosimetry
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calculations for the gestational component are presented in Appendix C). Groups of females that
birthed litters were treated with the same concentrations in the diet from Postnatal Day (PND) 1
to PND 20 of the last litter delivered (dosimetry calculations for the lactational component are
presented in Appendix C). Water and food were provided ad libitum. Dams were monitored
twice daily and observed for labor beginning at GD 18 up to 24. PND 0 was designated as the
day that litters were delivered. The study authors reported body weights and clinical findings
observed in dams on GDs and PNDs 1, 7, 14, and 20 (GD 18 for 2-week study). Pups (Fl) were
evaluated (number, weight, and sex) on PNDs 1, 4 (except for the 2-year study), 7, 14, and 20.
Pups from mated pairs were culled to eight pups (maximum) for the 2- and 13-week studies and
10 pups for the 2-year study with equal numbers of males and females per pair; culled weanlings
were removed and not examined further. Remaining pups were reevaluated on PND 20, and
two males and females per litter were chosen to be treated for the remainder of the study (2- and
13-week studies); the dams of these selected pups were necropsied at this time. For the 2-year
study, three pups/sex/litter were randomly chosen for the core study (2-year treatment) and two
pups/sex/litter were selected for special-study groups for hematology, clinical chemistry, or
urinalysis evaluation (treated up to 78 weeks) on PND 18. Necropsy results of the dams were
not provided by the study authors. Weaning of the Fl rats occurred on PND 21. Fl rats were
treated with the same concentrations of SAN Trimer in the diet (NTP-2000) as their respective
dams beginning on PND 21 until study termination. It is assumed that pups received 100% of
the dose given to their respective dams during lactation.
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All S tudies1
F0
GD 7 to Delivery
PND 1 to PND 20*
2-Week S tudy
F1
GD 7 to Delivery
PND 1 to PND 20
PND 21+2 weeks
13-Week Study
f:
GD 7 to Delivery
PND 1 to PND 20
PND 21 + 13 weeks1
2-Year Study
f:
GD 7 to Delivery
PND 1 to PND 20
PND 21 + 104 weeks, core
study eroup
PND 21 +27, 52. and 7S
weeks; special study groups
Lactational
and Dietary
GD = Gestation Day, PND = Postnatal Day
aS tudies are labeled based on ttie time F1 rats were directly exposed to SAN Trimer in the diet.
*PND 20 of the last litter delivered.
Identified as 14 weeks in Figure 1 of the NIP (2012) studyreport. Hiirteen weeks is the correct designation as
described on page 67 of the NIP (2012) studyreport; "Animals in the 7-week, 13-we ek. and 2-year studies were
exposed to the test chemical for 2 weeks during their in itiero development, for 3 weeks through their mother's
milk plus consumption of dosed feed, and for 2 weeks. 13 weeks. and 2 years after weaning through dosed feed."
Figure 4. Study Design for the 2-Week, 13-Week, and 2-Year Perinatal and Postnatal Feed
Studies of SAN Trimer (NTP. 2012)
NTP (2012) 2-Week Study
In the 2-week study, groups of 7 or 8 pregnant female F344/N rats (F0) were fed diets
containing 0, 250, 500, 1,000, 2,000, or 4,000 ppm SAN Trimer (equivalent to 0, 18, 35, 67, 130,
or 197 mg/kg-day determined for this PPRTV assessment by calculating the time-weighted
average doses provided by the study authors) from GD 7 to delivery of pups. Of the dams with
litters, groups of 6-8 females were further treated with the same concentrations in the diet as
gestation from PND 1 to PND 20 (equivalent to 0, 40, 85, 166, 325, or 634 mg/kg-day as
calculated for this PPRTV assessment). Doses for F1 pups were calculated with the assumption
that they received 100% of the dose given to their respective dams. Of the 60 F0 female rats that
were mated during the 7-day breeding period, 46 were identified to be sperm positive and
44 gave birth to live pups on GD 23 or GD 24. The two dams that did not give birth were
determined to not be pregnant or had unsuccessful pregnancies. Of the 44 dams that gave birth,
95% produced litters of 3 or more pups. Animals were monitored and data collected as
described above. No apparent effects of SAN Trimer were observed on gestation length,
fertility, litter size, live birth/implantation ratio, or sex number. At the highest dose tested
(197 mg/kg-day during gestation and 634 mg/kg-day during lactation), dams at GD 14 and 18
and PNDs 1, 7, 14, and 20 showed statistically significant decreases in body weight (see
Table A-4). Food consumption was decreased in dams at the two highest doses (130 and
197 mg/kg-day) from GDs 7-14. From PNDs 8-15 and 15-20, food consumption was reduced
Dietary
Inutero
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(30% lower compared to controls) in dams at the high dose (634 mg/kg-day). Statistical
evaluation was not performed for food consumption data by the study authors. For F1 male and
female pups, body weight was statistically significantly reduced at the highest dose
(634 mg/kg-day) from PNDs 1 to 14 and at 325 mg/kg-day on PND 20 (see Table A-5). From
this study, a maternal LOAEL of 197 mg/kg-day is identified during gestation with a
corresponding NOAEL of 130 mg/kg-day for statistically significantly decreased body weight in
F0 dams on GD 14. Also, a maternal LOAEL of 634 mg/kg-day is established during lactation
with a NOAEL of 325 mg/kg-day based on statistically significantly decreased body weight in
dams. For F1 pups, a LOAEL of 325 mg/kg-day (NOAEL of 166 mg/kg-day) is identified for
statistically significantly decreased body weight as measured at PND 20.
Groups of F1 rats (10/sex/dose) from the treated dams were fed diets containing 0, 250,
500, 1,000, 2,000, or 4,000 ppm SAN Trimer (equivalent to 0, 50, 90, 175, 270, or
410 mg/kg-day in males and 0, 45, 90, 185, 295, or 430 mg/kg-day in females, calculated by the
study authors) for 2 weeks postweaning (PND 21 to PND 35). Food consumption was recorded
on Days 1, 4, 8, 11, and at the end of the study. The heart, kidney, liver, lung, spleen, thymus,
and uterus were weighed at study termination. No clinical chemistry or hematologic analyses
were conducted. Histopathology was conducted on brain, kidney, liver, spleen, epididymis,
prostate, seminal vesicle, testis, ovary, and uterus where applicable. Body weights were
statistically significantly reduced in 175-, 270-, and 410-mg/kg-day males and 295- and
430-mg/kg-day females at study termination. At PND 35, body weight reductions in females
exceeded 10% at both 295 and 430 mg/kg-day. However, food consumption was also reduced
(not statistically significant) in the aforementioned dose groups. There was a dose-related
decrease in food consumption that was less than half of the control at the high dose. Observed
organ weight differences (see Table A-6) included: (1) significant decreases in absolute heart
(>175 mg/kg-day [M]; >295 mg/kg-day [F]), right kidney (>270 mg/kg-day [M];
>295 mg/kg-day [F]), liver (>270 mg/kg-day [M]; >295 mg/kg-day [F]), lung (>270 mg/kg-day
[M]; >295 mg/kg-day [F]), and spleen (>410 mg/kg-day [M]; >430 mg/kg-day [F]);
(2) significant decreases in absolute and relative thymus weight in males (>90 mg/kg-day) and
(>295 mg/kg-day) females, right testis weight in males (>90 mg/kg-day), and uterus weight in
(>295 mg/kg-day) females; and (3) significant increases in relative heart (>410 mg/kg-day [M];
>430 mg/kg-day [F]), right kidney (>410 mg/kg-day [M]; >430 mg/kg-day [F]), liver
(>175 mg/kg-day [M]; >185 mg/kg-day [F]), lung (>410 mg/kg-day [M]; >430 mg/kg-day [F]),
and spleen (>270 mg/kg-day [M]; >295 mg/kg-day [F]). Nonneoplastic lesions were observed in
the brain, thymus, spleen, liver, kidney, and reproductive organs of both sexes, but the study
authors characterized these lesions as not toxicologically relevant and considered them
secondary to severe calorie reduction; the dose levels at which these lesions occurred were not
specifically identified by the study authors. There was, however, increased cellularity in the
cerebrum at the level of the septal nuclei (incidences of 0/10, 0/9, 0/6, 0/3, 0/5, and 4/6 in males
and 0/5, 0/3, 0/2, 0/3, 1/5, and 10/10 in females). The increased cellularity could be due to a
developmental delay caused by inadequate caloric intake, but other mechanisms cannot be ruled
out.
A LOAEL of 90 mg/kg-day is established for statistically significantly decreased relative
right testis and thymus weights in the male F1 offspring with a corresponding NOAEL of
50 mg/kg-day.
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Subchronic-Duration Studies
NTP (2012) 13-Week Study
Beginning on presumed GD 7, the study authors administered dietary (NIH-07) doses of
0, 100, 200, 400, 800, and 1,600 ppm (equivalent to 0, 6.9, 14, 28, 55, or 110 mg/kg-day as
calculated for this PPRTV assessment)1 of SAN Trimer to groups of 8 or 9 sperm-positive
females (F0). Groups of 4-8 females that birthed litters were treated with the same
concentrations (equivalent to 0, 17, 33, 66, 132, or 264 mg/kg-day as calculated for this PPRTV
assessment)1 up to PND 20 of the last litter delivered. Doses for F1 pups were calculated with
the assumption that they received 100% of the dose given to their respective dams. Of the
60 F0 female rats that were mated during the 7-day breeding period, 50 were sperm positive and
39 gave birth to live pups on GD 23 or GD 24. For the females that did not give birth, it was
unclear whether they were not impregnated or the lack of delivery was due to resorption. Of the
dams that produced litters, only one (treated with 110 mg/kg-day) delivered less than five pups.
One pup died in the control group prior to PND 4; no cause of death was provided by the study
authors. A litter of 8 pups in the low-dose group was sacrificed moribund after the dam died due
to chylothorax (leakage of lymphatic fluid of intestinal origin into the pleural space). Besides
culling that occurred on PND 4, all remaining pups survived through weaning. The study
authors reported an effect of SAN Trimer on fertility index, number of litters, and litter size in
dams treated with 110 mg/kg-day of the chemical. The data for fertility index was determined
not to be statistically significant based on Fisher's Exact test, and no variance information was
provided for number of litters and litter size to perform statistical analysis.
The study authors reported statistically significant decreased body weight on GD 14 in
dams treated with 28, 55, and 110 mg/kg-day of SAN Trimer (see Table A-7). Body weight was
statistically significantly reduced in dams in the high-dose group (110 mg/kg-day) on GD 20.
No body-weight changes were observed in male pups during lactation. Body weight in female
pups was statistically significantly increased on PND 20 at 17 mg/kg-day (see Table A-8); the
biological relevance of increased body weight is unknown. A maternal LOAEL of
110 mg/kg-day is identified with a corresponding NOAEL of 55 mg/kg-day for statistically
significantly decreased body weight in dams observed during gestation. A NOAEL of
264 mg/kg-day is identified during lactation based on lack of effects observed in dams or pups.
Because 264 mg/kg-day is the highest dose tested, a LOAEL for lactational effects cannot be
determined.
Postweaning, F1 rats (10/sex/dose) were treated for 13 weeks with the same dietary
(NTP-2000) concentrations of SAN Trimer (equivalent to 0, 10, 20, 40, 80, or 150 mg/kg-day as
calculated by the study authors) as the F0 generation. Water and food were provided ad libitum.
Rats were monitored twice daily. The study authors reported body weights and clinical findings
at the beginning of treatment, weekly, and at study termination. Food consumption was reported
biweekly until the end of the study. At the end of the study, urine samples were collected and
the following parameters were included in the urinalysis: creatinine, glucose, protein, alkaline
phosphatase, aspartate aminotransferase, /V-acetyl-P-D-glucosaminidase, volume, and specific
gravity. The study authors collected blood samples from the retro-orbital sinus for measurement
of the following clinical chemistry parameters: urea nitrogen, creatinine, glucose, sodium,
1 Dose = Feed Concentration x Food Consumption per Day x (l -f- Body Weight) x (Days Dosed Total Days),
where the food factor was calculated from the body weight and food consumption data reported in dams from the
2-week NTP (2012) study.
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potassium, chloride, calcium, phosphorus, total protein, albumin, total bilirubin, cholesterol,
triglycerides, alanine aminotransferase, alkaline phosphatase, aspartate aminotransferase,
creatine kinase, lactate dehydrogenase, sorbitol dehydrogenase, y-glutamyl transferase, and bile
acids. The following parameters were evaluated for hematology: hematocrit, hemoglobin
concentration, erythrocyte, reticulocyte, platelet counts, Howell-Jolly bodies, mean cell volume,
mean cell hemoglobin, mean cell hemoglobin concentration, leukocyte count and differentials,
and activated partial thromboplastin time. To assess sperm motility, sperm samples were taken
from male rats treated with 0, 40, 80, and 150 mg/kg-day of SAN Trimer. The following
parameters were measured to evaluate sperm motility: spermatid heads per testis and per gram
testis, spermatid counts, and epididymal spermatozoa motility and concentration. The left cauda,
epididymis, and testis were weighed. To evaluate vaginal cytology, samples were collected for
up to 12 days before study termination from females in the 0, 40, 80, and 150 mg/kg-day dose
groups. Estrous cycles were evaluated to determine the probability of extended estrus, diestrus,
and metestrus. After 13 weeks of direct exposure to SAN Trimer in the diet, F1 rats were
necropsied. The study authors collected and weighed the following organs: brain, heart, right
kidney, liver, lung, spleen, right testis, thymus, and uterus. Complete histopathological
examinations for the 0 and 150 mg/kg-day-dose groups were performed on the following organs:
adrenal gland, bone with marrow, brain, clitoral gland, esophagus, eye, Harderian gland, heart
and aorta, large intestine (cecum, colon, rectum), small intestine (duodenum, jejunum, ileum),
kidney, liver, lung, lymph node (mandibular and mesenteric), mammary gland, nose, ovary,
pancreas, parathyroid gland, pituitary gland, preputial gland, prostate gland, salivary gland, skin,
spleen, stomach (forestomach and glandular), testes with epididymis and seminal vesicle,
thymus, thyroid gland, trachea, urinary bladder, and uterus. Tissues were fixed and preserved,
embedded in paraffin, sectioned, and stained with hematoxylin and eosin.
Survival was 100% for F1 rats that were directly exposed to SAN Trimer in the diet for
13 weeks. Body and organ weight data are presented in Table A-9. Terminal body weight was
statistically significantly decreased in male rats at the high dose. The study authors reported
statistically significantly decreased body weight in female rats at >20 mg/kg-day. Increased
absolute and relative liver weights were statistically significant at all dose levels in male rats.
Increased relative liver weight was statistically significant at >40 mg/kg-day in female rats.
Absolute spleen weight in male rats was statistically significantly increased at the two highest
doses and at the high dose in females. Relative spleen weight was statistically significantly
increased at >40 mg/kg-day in male rats and at the two highest doses in females. Increased
relative brain weight was statistically significant at the high dose in males and at doses
>20 mg/kg-day in females. Statistically significantly increased relative right testis weight was
reported in males at the high dose. Absolute right testis and left testis weights were statistically
significantly decreased in males at the high dose. Absolute heart weight was statistically
significantly increased at 10, 40, and 80 mg/kg-day in male rats. Increased relative heart weight
in male rats was statistically significant at all doses and at the two highest doses in females.
Increased relative kidney weight in male and female rats was reported to be statistically
significant in the two highest dose groups. The study authors reported that no histopathologic
lesions due to SAN Trimer were observed in the organs examined for both male and female rats.
Exposure to SAN Trimer had no statistically significant effect on sperm parameters in males or
on the estrous cyclicity in female rats.
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The hematology, clinical chemistry, and urinalysis findings are summarized in
Table A-10. In high-dose male rats, erythrocytes and hemoglobin were statistically significantly
decreased (both 3-4% below control) while mean cell volume was statistically significantly
increased (2% above control). There were no statistically significant changes in hematology in
female rats at any dose level. Serum albumin was statistically significantly increased in males in
the two highest dose groups. Serum total protein was statistically significantly increased in
males in the 80 mg/kg-day-dose group, but not in males of the highest dose group. There was
statistically significantly reduced serum cholesterol in male rats at the high dose. Serum
triglycerides were statistically significantly decreased at 40, 80, and 150 mg/kg-day in male rats
and 80 and 150 mg/kg-day in females. In males, alanine aminotransferase was statistically
significantly decreased starting at 20 mg/kg-day. Aspartate aminotransferase was statistically
significantly reduced at all dose levels in males and at 80 and 150 mg/kg-day in females. There
was a statistically significant decrease in bile acids at 40 mg/kg-day in males and at 10 and
80 mg/kg-day in females. In female rats, the following statistically significant changes were also
observed in clinical chemistry parameters: increased creatinine (20-150 mg/kg-day) and elevated
urea nitrogen (150 mg/kg-day). Urinalysis revealed the following statistically significant
changes in male rats: increased glucose (80 mg/kg-day), elevated glucose/creatinine ratio (80 and
150 mg/kg-day), increased protein (80 and 150 mg/kg-day), increased protein/creatinine ratio
(150 mg/kg-day), aspartate aminotransferase (decreased at 80 mg/kg-day and increased at
150 mg/kg-day), and aspartate aminotransferase/creatinine ratio (decreased at 80 mg/kg-day and
elevated at 150 mg/kg-day). Urinalysis in female rats revealed that the alkaline
phosphatase/creatinine ratio was statistically significantly decreased at the high dose and
A'-acetyl-P-creatinine-D-glucosaminidase/creatinine ratio was statistically significantly decreased
in the two highest dose groups.
Based on statistically significantly increased absolute and relative heart weight in
F1 male rats, a LOAEL of 10 mg/kg-day is identified. Because 10 mg/kg-day is the lowest dose
tested, a NOAEL cannot be determined.
Chronic-Duration Studies
NTP (2012) 2-Year Study
In the chronic-duration study by NTP (2012). 41 or 42 pregnant females per dose group
were treated with 0, 400, 800, or 1,600 ppm (equivalent to 0, 28, 55, or 110 mg/kg-day as
calculated for this PPRTV assessment)1 via the diet from GD 7 until delivery. Of the female rats
that were mated during the 7-day breeding period, 83% were determined to be sperm positive
and 52% gave birth to at least one live pup; those pups were treated with the same dietary
concentrations (equivalent to 0, 66, 132, or 264 mg/kg-day as calculated for this PPRTV
assessment)1 through PND 20. Doses for F1 pups were calculated with the assumption that they
received 100% of the dose given to their respective dams. There were no effects of SAN Trimer
on the fertility index, gestational index, or number of litters during gestation and lactation. Four
dams, whose offspring either died or were fostered to other litters, were sacrificed during the
postnatal phase. The study authors reported that 45 pups died (contributions from all dose
groups) from PND 1 to PND 20. However, the survival rate was greater than 92% for pups of all
dose groups. The study authors provided no explanation as to why pups died during postnatal
exposure. The study authors reported no changes in body weight in dams or F1 pups exposed
from GD 7 to PND 20 (data not shown). Based on no effects observed in dams or pups,
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NOAELs of 110 mg/kg-day (gestation) and 264 mg/kg-day (lactation) are identified. Because
these are the highest doses tested, LOAELs for gestational and lactational effects cannot be
determined.
Groups of F1 rats (50/sex/dose) were directly treated with SAN Trimer in the diet at
doses of a 0, 20, 40, and 75/85 (males/females, respectively) mg/kg-day as calculated by the
study authors for 2 years beginning at PND 21; these rats were referred to as the core study
groups by the study authors. Groups of rats (20/sex/dose) were exposed to the same
concentrations in the feed as the core study groups but were sacrificed at various time points up
to 78 weeks in special study groups to identify effects on urinalysis, clinical chemistry,
hematology parameters, and plasma concentrations of SAN Trimer. Rats were monitored twice
daily for mortality and morbidity. The study authors recorded body weight and food
consumption weekly for the first 13 weeks then monthly afterward; body weight was also
documented at the conclusion of the study. The study authors also reported clinical observations
at PND 29 and monthly, thereafter. At 23, 48, and 74 weeks postweaning, 3 rats/sex/dose along
with 6 controls (both sexes) from the special study groups were switched to a SAN Trimer-free
diet and blood was collected periodically up to 360 minutes after switching to SAN Trimer-free
feed. During the Week 23 collection, male rats scheduled to be evaluated at 240 and
360 minutes were actually bled at 180 and 300 minutes, respectively. Plasma from these animals
was analyzed by gas chromatography/mass spectrometry using 4-cyano-l,2,3,4-tetrahydro-
a-methyl-l-naphthaleneacetonitrile (THNA) and 4-cyano-l,2,3,4-tetrahydro-l-naphthalene-
propionitrile (THNP) as markers to determine concentrations of SAN Trimer. After blood was
collected from these animals, they were further exposed to SAN Trimer in the diet. Urine was
collected from 10 rats/sex/dose in the special study at postweaning weeks 26, 51, and 77 weeks.
Urine samples were evaluated for the same parameters (except y-glutamyltransferase) as the
13-week study described above. One week after urine collection (27, 52, and 78 weeks
postweaning), the study authors collected blood from the retro-orbital sinus of 10 rats/sex/dose in
the special study. These blood samples were analyzed for similar hematology and clinical
chemistry parameters as described in the 13-week study. All core study rats and special study
rats bled at Week 78 underwent complete necropsies. Complete histopathological examinations
were performed on various organs for all F1 rats treated with SAN Trimer in the diet for 2 years.
Organ weights were not examined in the 2-year study.
None of the plasma samples that were analyzed by gas chromatography/mass
spectrometry contained SAN Trimer at concentrations above the limit of detection
(0.400 (j,g/mL). Further experiments with 55 samples from the low- and high-dose groups and
one control from the 74-week time point using limits of detection of 0.004 [j,g/mL for THNA
isomers and 0.01 [j,g/mL for the THNP isomers determined that 14 of the 55 samples exceeded
the limit for THNA. THNP was not above the limit of detection in any of the samples. Of the
samples that contained THNA, the levels were below 0.0500 [j,g/mL in 13 of 14 samples; 9 of the
14 samples were less than 0.0052 [j.g/mL. The study authors stated that no correlation was seen
between THNA concentration and exposure time.
After 2 years of treatment, there were no treatment-related effects on survival in F1 rats.
Statistical analysis of these body-weight data was not provided by the study authors and cannot
be performed due to the lack of variance information. There were no differences in food
consumption among the treated groups compared to controls. The study authors reported
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statistically significant changes in hematology, clinical chemistry, and urinalysis parameters (see
Table A-l 1). These changes were not consistent and were transient; therefore, the biological
significance of these effects is unclear.
In the histopathology evaluations, a number of nonneoplastic lesions and degenerative
changes were observed and are shown in Table A-12. There was a statistically significant
increase in the incidence of spinal nerve root degeneration in males in the high-dose group with
an accompanying dose-related increase in severity. In female rats, there was a statistically
significant increased incidence of sciatic nerve fiber degeneration at the two highest dose levels.
In the livers of males, there was a statistically significant increased incidence of angiectasis,
eosinic foci, and chronic active inflammation in the high-dose group. The incidence of mixed
cell foci in the liver was statistically significantly increased in males in the low- and high-dose
groups and in females at the high dose. There was a statistically significant increased incidence
of bone marrow hyperplasia in both sexes (at 75 mg/kg-day for males and at 40 and
85 mg/kg-day for females). In the urinary bladder, there was a statistically significant increased
incidence of hyperplasia of the transitional epithelium in females at the highest dose tested.
A LOAEL of 20 mg/kg-day is identified for this study based on a statistically significant
increased incidence of mixed cell foci in the livers of male rats. Because 20 mg/kg-day is the
lowest dose tested, a corresponding NOAEL cannot be identified.
Developmental Portion of the NTP (2012) Study
The NTP (2012) study did not examine any endpoints identified by the U.S. EPA's
Guidelines for Developmental Toxicity Risk Assessment (U.S. EPA. 1991) to assess
developmental toxicity.
Reproductive Portion of the NTP (2012) Study
The NTP (2012) study examined a limited number of reproductive parameters (e.g.,
fertility index) identified by the U.S. EPA's Guidelines for Reproductive Toxicity Risk
Assessment (U.S. EPA. 1996) to assess reproductive toxicity; therefore, this study is not
considered as a comprehensive reproductive study.
Carcinogenicity Studies
NTP (2012) 2-Year Cancer Results
The 2-year study by the NTP (2012) reported statistically significant changes in tumor
incidences in F1 male and female F344/N rats. Tumor data for male and female rats are listed in
Table A-13. In male rats, the incidences of total testicular interstitial cell adenomas (combined
unilateral and bilateral) and bilateral interstitial cell adenomas were statistically significantly
increased at the low (20 mg/kg-day) and high dose (75 mg/kg-day). An increase in combined
brain and spinal cord astrocytomas also occurred in male rats, but this increase was not
statistically significant at any dose level. The incidence of combined brain and spinal cord
granular cell tumors were also increased in male rats (not statistically significant). There was a
statistically significant decreased trend for pituitary gland adenomas, mononuclear cell leukemia
in all organs, and malignant neoplasms in all organs examined in male rats. Pituitary gland
adenomas were statistically significantly decreased at 75 mg/kg-day in male rats. There was a
statistically significant decrease in mononuclear cell leukemia in all organs at all dose groups.
Statistically significant decreased incidences of malignant neoplasms occurred at the mid
(40 mg/kg-day) and high dose (75 mg/kg-day) in male rats. For female rats, the following
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tumors occurred with a statistically significant negative trend: mammary gland tumors (classified
as fibroadenomas; fibroadenomas or adenoma; fibroadenomas, adenoma, or carcinoma), pituitary
gland tumors (i.e., adenomas and adenomas or carcinomas), and mononuclear cell leukemia in
all organs, and neoplasms (benign and benign or malignant). Mammary gland tumor incidences
were statistically significantly decreased at 40 and 85 mg/kg-day. Pituitary gland tumors in
female rats were statistically significantly decreased in the low- and high-dose groups (20 and
85 mg/kg-day, respectively). The incidence of mononuclear cell leukemia in all organs was
statistically significantly decreased at all dose levels. Malignant neoplasms (all organs) were
statistically significantly decreased at 20 and 40 mg/kg-day in female rats. Benign and benign or
malignant neoplasms incidences were statistically significantly decreased in female rats at the
high dose. Based on the results of this study, the NTP (2012) report concludes that there is no
evidence of carcinogenic activity for SAN Trimer.
Inhalation Exposure
No studies have been identified.
OTHER DATA
Metabolism Studies
Garsas et al. (2008)
Gargas et al. (2008) performed disposition studies to evaluate the pharmacokinetic
behavior of SAN Trimer in rats. Nonpregnant F344/N female rats were given a single dose of
[3H]SAN Trimer via intravenous injection (26 mg/kg) or gavage (25, 75, or 200 mg/kg in corn
oil), and pregnant and lactating rats were also administered a single gavage dose (200 mg/kg in
corn oil); the use of a control group was not reported by the study authors. For nonpregnant rats,
blood was taken at various time points ranging from 0.25 to 48 hours after SAN Trimer
administration, and urine and feces were collected between 24 and 168 hours after dosing. For
pregnant rats, the study authors sampled blood, placenta, fetus, and milk (collected from
lactating rats) at a single time point 2 hours after gavage. Liquid scintillation spectroscopy and
HPLC were performed to analyze the collected samples. The study authors determined that in
nonpregnant rats, the elimination half-lives for [3H]SAN Trimer following intravenous and
gavage dosing were approximately 1 hour and 3.5 hours, respectively. For nonpregnant rats,
Cmax was achieved from 0.25 to 2 hours and was dose dependent following gavage
administration of SAN Trimer, and was determined to be achieved 0.25 hours following an
intravenous dose. SAN Trimer was rapidly excreted in the urine and feces after intravenous and
gavage exposure. The study authors characterized the excreted radiolabel and determined that
less than 1% of the radioactivity in urine and less than 3% in feces was from unchanged
[3H]SAN Trimer. For pregnant rats 2 hours post gavage, concentrations of radioactivity and
SAN Trimer were highest in the blood, but the chemical was also detected in the placenta and
fetus. In lactating rats, there was marginally more radioactivity and SAN Trimer detected in
milk than in blood. The study authors concluded that SAN Trimer is rapidly absorbed,
metabolized, and excreted in nonpregnant female rats, and not likely to accumulate in blood or
tissues. They also concluded that fetal exposure to SAN Trimer can occur in utero through the
ability of the compound to pass the placental barrier during gestation. It was determined that
exposure can also occur via lactation.
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Genotoxicity
MA Bioservices (1998a). (1998c). (1998b). (1998d)
Table 3 summarizes the studies examining genotoxicity (e.g., clastogenicity,
mutagenicity) of SAN Trimer. One group of studies was conducted with SAN Trimer Batch 2,
which was made by distillation of the product to 98% purity. In MA Bioservices (1998a) a
bacterial reverse mutation assay was conducted in Salmonella typhimurium strains TA98,
TA100, TA1535, TA1537, and TA102 with and without S9 extracts. A positive response was
observed in strains TA98, TA100, and TA1537 in the absence of S9 activation. In a Chinese
hamster ovary (CHO) cell in vitro mutation assay by MA Bioservices (1998b). SAN trimer did
not induce mutations in the hgprt gene with and without S9 activation. MA Bioservices (1998c)
also conducted an in vitro mammalian chromosome aberration assay in CHO cells that was
concluded to be positive with a statistically significant increase in the percentage of cells with
chromosomal aberrations with and without S9 activation after a 4-hour treatment with
SAN Trimer, but not following 20 hours of treatment. In MA Bioservices (1998d). a mammalian
bone marrow chromosome aberration and micronucleus test was conducted in S-D rats in vivo.
There were no statistically significant increases in abnormal erythrocytes at 18 or 42 hours, and
the study authors concluded both tests to be negative.
Hobbs et al. (2012), NTP (2012), Vacek et al. (2005)
The following tests used SAN Trimer Batch 3 (96% purity), which is prepared by
extraction rather than distillation. It is postulated that distillation, which subjects the mixture to
high temperatures, produces a different mixture of compounds than Batch 2. Hobbs et al. (2012)
treated F344/N juvenile male and female rats (four to five rats per treatment group) with
SAN Trimer (0, 37.5, 75, 150, or 300 mg/kg-day) via gavage in corn oil, once daily for 4 days.
Using a comet assay, the study authors determined that SAN Trimer caused a statistically
significant increase in DNA damage in total brain, cerebrum, cerebellum (not statistically
significant in males), liver, and blood leukocytes (not statistically significant in females) in male
and female rats as determined by quantitative staining. Hobbs et al. (2012) also reported
statistically significant increased frequencies of micronucleated reticulocytes in both male
(300 mg/kg-day) and female (>150 mg/kg-day) rats (Hobbs et al.. 2012). In a study conducted
by SITEK Research Laboratories that was included in the NTP (2012) study, SAN Trimer was
not mutagenic in Salmonella typhimurium strains TA98 or TA100 or in Escherichia coli strain
WP2 wvr.4/pKM101 in tests conducted with and without S9 activation. Vacek et al. (2005)
compared the mutation frequencies of the hypoxanthine phosphoribosyl transferase (HPRT) gene
in nonexposed, healthy children to children living in Dover Township, New Jersey whose
siblings were included in a childhood cancer incidence study (NJDHSS. 1997). The study
authors observed no change in mutation frequency between the two groups (Vacek et al.. 2005).
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Table 3. Summary of Studies Evaluating Genotoxicity and Mutagenicity
Endpoint
Test System
Dose
Concentration
Results3
Comments
References
Without
Activation
With
Activation
Genotoxicity studies in prokaryotic organisms
Reverse
mutation
S. typhimurium
strains TA98,
TA100, TA1535,
TA1537, TA102
25-1,000 ng/plate
without S9
activation;
25-5,000 ng/plate
with S9 activation
+
(TA98,TA100,
TA1537)
(TA1535,
TA102)
SAN
Trimer
Batch 2
MA Bioservices
ri998a^
Reverse
mutation
S. typhimurium
strains TA98,
TA100
0-7,500 (ig/plate
SAN
Trimer
Batch 3
NTP (2012)
Reverse
mutation
Escherichia coli
strain WP2
uvrA pKM 101
0-10,000 (ig/plate
SAN
Trimer
Batch 3
NTP (2012)
Genotoxicity studies in mammals—in vitro
Gene mutation
CHO cellsIhgprt
50-400 iig/mL
without S9
activation;
150-600 ng/mL
with S9 activation
SAN
Trimer
Batch 2
MA Bioservices
(1998b)
Chromosome
aberration
CHO cells
50-450 |ig/mL
for 4 hours
25-400 ng/mL
for 20 hours
+
+
SAN
Trimer
Batch 2
Marginally
positive
results
obtained at
4 hours
(approx.
1.8% above
control at
440 ng/mL
without S9)
MA Bioservices
(1998c)
Genotoxicity studies in mammals—in vivo
Chromosome
aberration
S-D, rats, bone
marrow (single
dose gavage)
0, 125, 250, or
500 (males)and 0,
163, 325, or 650
(females) mg/kg
ND
SAN
Trimer
Batch 2
MA Bioservices
ri998d*)
Gene mutation
Human peripheral
lymphocyte s IHPR T
NDr
ND
SAN
Trimer
Batch 3
Vacek et al.
(2005)
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Table 3. Summary of Studies Evaluating Genotoxicity and Mutagenicity
Results3
Endpoint
Test System
Dose
Concentration
Without
Activation
With
Activation
Comments
References
DNA damage
F344/N juvenile
male and female
rats (gavage, four
to five rats per
treatment group,
4 days)
0, 37.5, 75, 150,
or 300 mg/kg-day
(+) Total brain
(+) Cerebrum
(+) Cerebellum
(females)
(+) Liver (at
p <0.05)
(+) Blood
Leukocytes
(males)
ND
SAN
Trimer
Batch 3
Hobbs et al.
(2012): also
rcDortcd in NTP
(2012)
Micronucleus
test
F344/N juvenile
male and female
rats (gavage,
4 days)
0, 37.5, 75, 150,
or 300 mg/kg-day
(+)
Reticulocytes
(-) Erythrocytes
ND
SAN
Trimer
Batch 3
Lowest
dose was
not
analyzed
Hobbs et al.
(2012): also
rcDortcd in NTP
(2012)
Micronucleus
test
S-D, rats, bone
marrow (single
dose gavage)
0, 125, 250, or
500 (males)and 0,
163, 325, or 650
(females) mg/kg
ND
SAN
Trimer
Batch 2
MA Bioservices
C1998dN)
a- = negative; NA = not applicable; ND = no data; NDr = not determined.
In summary, SAN Trimer was shown to be both positive and negative for mutagenicity in
S. typhimurium strains and was not mutagenic in E. coli strains. SAN Trimer was not shown to
induce hgprt mutations in CHO cells and HPRT mutations in human lymphocytes. In in vivo rat
studies, SAN Trimer caused micronuclei formation in reticulocytes but not in bone marrow
erythrocytes following gavage. SAN Trimer induced chromosomal aberrations in CHO cells in
vitro but was negative in bone marrow erythrocytes from treated rats. SAN Trimer also caused
DNA damage in brain, liver, and blood following gavage treatment in rats.
SYNTHESIS OF RESULTS FROM NONCANCER AND CANCER STUDIES
Four human studies on oral exposure to drinking water containing various chemicals,
including SAN Trimer, are publicly available; one was a case-control study and the other three
were ecological in nature because they examined town-level incidence and exposure data. The
experimental database for animals is limited and includes a single acute study, two short-term-
duration studies, a single sub chronic-duration study, and a single chronic-duration study, all
conducted in rats.
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During the period from 1979-1995, the following childhood cancers were determined to
be statistically significantly increased in children exposed to drinking water containing multiple
chemicals, including SAN Trimer, in Ocean County, Dover Township, and the Toms River
section of Dover Township, New Jersey: sympathetic nervous system, leukemia, brain/CNS
(including other tumor types and astrocytomas), acute lymphocytic leukemia, and brain/CNS
astrocytomas (NJDHSS. 1997). In the case-control study on individuals exposed during the
period between 1979 and 1996 in Dover Township and the Toms River section of Dover
Township (NJ DHSS. 2003a). the study authors observed a statistically significant association
for leukemia in females (0-19 years of age) and exposure to drinking water The results from the
case-control study should be interpreted with caution because of the small number of study
subjects. In addition, some of the other chemicals that were present in the drinking water,
including trichloroethylene and tetrachloroethylene, are known to demonstrate carcinogenic
potential. Following cleanup and removal of SAN Trimer along with the other chemicals from
the drinking water, total childhood cancer and brain/CNS cancer incidence have returned to
background levels, and fewer than expected cases of leukemia have been observed (see Figures 2
and 3 and Tables A-2 and A-3) (NJ DHSS 2008. 2003b). Due to the existence of other
contaminants in drinking water from the well fields investigated, the conclusions that can be
drawn from the epidemiologic studies are relatively limited.
In a 2-year cancer bioassay conducted by the NTP, the study authors reported increased
incidences of testicular tumors, brain and spinal cord astrocytomas, and granular cell tumors (see
Table 12) in male F344/N rats (Fl) treated perinatally and postnatally with SAN Trimer for
2 years (NTP. 2012). Although the testicular tumors were statistically significantly increased,
the high background incidence in the control group suggests that these tumors may not be related
to SAN Trimer treatment. The incidences of brain and spinal cord astrocytomas and granular
cell tumors in males were not statistically significantly increased. Noncancer nervous system
effects in rats consisting of sciatic nerve degeneration (at >40 mg/kg-day in female rats) and
spinal root degeneration (at 75 mg/kg-day in male rats) were reported in the 2-year NTP (2012)
study. There was also increased incidence of cellularity in the brain cerebrum in male and
female rats (statistically significant at 430 mg/kg-day in females) treated with SAN Trimer in the
2-week NTP study. These effects indicate the nervous system as a potential target organ for
SAN Trimer toxicity.
There were also consistent liver effects in rats treated with SAN Trimer at various time
points. At 2 weeks of treatment, SAN Trimer increased absolute (150 mg/kg-day) and relative
liver weights (>75 mg/kg-day) in males and females at >150 mg/kg-day as well as liver
histopathological lesions at 300 mg/kg-day, including vacuolation of periacinar hepatocytes in
males and periacinar hypertrophy in females (Huntingdon Life Sciences. 1999b). The NTP
studies reported increased relative liver weight at >175 mg/kg-day in male and female rats at
2 weeks and at >40 mg/kg-day in male rats at 13 weeks (NTP. 2012). Following 2 years of
treatment, the study authors reported the following histopathological noncancer changes in the
liver: chronic active inflammation, eosinophilic foci, angiectasis (all at 75 mg/kg-day in males),
and mixed cell foci at >20 mg/kg-day in males and at 85 mg/kg-day in females (NTP. 2012).
These observed effects identify the liver as a potential target organ for SAN Trimer toxicity.
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Finally, there were effects on reproductive parameters (e.g., testis weight, pup body
weight) and other organs including the heart, bone marrow, and bladder, but these effects were
not as consistently observed as the nervous system or hepatic effects. The potential points of
departure (PODs) for subchronic- and chronic-duration effects are shown in Tables 6 and 9,
respectively.
DERIVATION OF PROVISIONAL VALUES
Tables 4 and 5 present a summary of provisional noncancer reference values and cancer
values, respectively.
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Table 4. Summary of Provisional Noncancer Reference Values for SAN Trimer (Various CASRNs)
Toxicity Type
Species/Sex
Critical Effect
p-Reference Value
POD
Method
PODhed
UFc
Principal Study
Subchronic p-RfD
F344/N Rat/M
Increased absolute liver weight
8 x 1CT3 mg/kg-day
BMDLio
2.4 mg/kg-day
300
NTP (2012)
Chronic p-RfD
F344/N
Rat/Both
Increased incidences of chronic active
inflammation in the liver (males) and sciatic nerve
degeneration (females)
3 x 1CT3 mg/kg-day
BMDLio
0.77 mg/kg-day
300
NTP (2012)
Subchronic p-RfC
ND
Chronic p-RfC
ND
ND = not determined.
Table 5. Summary of Provisional Cancer Values for SAN Trimer (Various CASRNs)
Toxicity type
Species/Sex
Tumor Type
Cancer Value
Principal Study
p-OSF
ND
p-IUR
ND
ND = not determined.
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DERIVATION OF PROVISIONAL ORAL REFERENCE DOSES
Derivation of Subchronic Provisional RfD (Subchronic p-RfD)
The subchronic-duration portion of the NTP (2012) study in F344/N rats is selected
as the principal study for derivation of the subchronic p-RfD. In this study, groups of F1 rats
(10/sex/dose) from SAN Trimer-treated dams were fed diets containing SAN Trimer for
13 weeks postweaning. This study is a peer-reviewed technical report and was conducted
according to GLP standards. It is well conducted and meets the standard of study design and
performance, with numbers of animals, examination of potential toxicity endpoints, and
presentation of information (details are provided in the Review of Potentially Relevant Data
section), and is the only available study that reports effects following subchronic-duration
SAN Trimer treatment.
The study authors (NTP. 2012) reported statistically significant changes in body weight
and organ weights that were amenable in some cases to modeling using the U.S. EPA's
Benchmark Dose Software (BMDS version 2.2 as described in Appendix B) for consideration as
potential PODs (see Table 6). Increased absolute organ weight changes in female F1 rats
observed in the 13-week NTP (2012) study are not considered as potential critical effects
because there were concomitant statistically significant decreases in body weight at all doses
expect the lowest dose (10 mg/kg-day) that could have contributed to the increased absolute
organ weights. For absolute organ weight changes in male F1 rats, data were modeled without
the high dose because there was a statistically significant decrease in body weight at the highest
dose (150 mg/kg-day).
The most sensitive effects following subchronic-duration treatment with SAN Trimer
appear to be increased heart weight (absolute and relative) in male rats with a LOAEL of
10 mg/kg-day, and increased absolute liver weight in male rats with a BMDLio of 10 mg/kg-day
(BMR of 10% relative risk). However, there is more support for increased absolute liver weight
being selected as the critical effect based on the available data for SAN Trimer. When excluding
the high dose as described above, the data for increased absolute liver weight in male rats display
a clearer dose-response trend than the data for increased absolute and relative heart weights in
male rats (see Table A-9). Increased heart and liver weights were both observed in multiple
duration studies (NTP. 2012; Huntingdon Life Sciences. 1999b). However, increased incidences
of pathological indices of heart toxicity were not observed in F1 rats that received dietary
treatment of SAN Trimer for 2 weeks, 13 weeks, or 2 years (NTP. 2012). Heart lesions were
also not observed in S-D rats that received gavage treatment of SAN Trimer for 2 weeks. The
absence of pathological heart effects following SAN Trimer treatment suggests that increased
absolute and relative heart weights may not be biologically significant; therefore, data for
increased absolute and relative heart weights are not selected as a potential POD. The selection
of increased absolute liver weight as the critical effect is supported by the observation that the
liver appears to be a target organ of SAN Trimer toxicity. Liver effects (i.e., liver lesions and
increased absolute and relative liver weights) were reported in both sexes of two strains of rats
(F344/N and S-D) in short-term-duration studies (NTP. 2012; Huntingdon Life Sciences. 1999b).
and F344/N rats in a chronic-duration study of dietary SAN Trimer treatment (NTP. 2012).
Therefore, the BMDLio of 10 mg/kg-day based on increased absolute liver weight in male
F1 rats (NTP, 2012) is chosen as the POD to derive a subchronic p-RfD.
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Table 6. Candidate Subchronic PODs for SAN Trimer
Effect
Sex
NOAEL
(mg/kg-day)
LOAEL
(mg/kg-day)
BMD
BMDL
Comment
Decreased body weight in
dams (F0) during gestation
Females
55
110a
Not Run
Not Run
No data variability
available
Decreased body weight in
F1 ratsd
Males
80
150a
94b
141°
83b
127°
Increased relative heart weight
Males
NDr
10
NF
NF
Increased absolute heart
weight
Males
NDr
10
NF
NF
Increased relative right kidney
weightd
Males
80
150a
55b
lllc
44b
91°
Increased relative liver
weightd
Males
20
40a
12b
35°
8.3b
24°
Increased absolute liver
weightd
Males
20
40a
13b
25c
5b
10c
Increased relative spleen
weight
Males
20
40
46b
38b
Increased absolute spleen
weight
Males
40
80
82b
48b
Increased relative right testis
weight
Males
80
150
NF
NF
Decreased body weight in
F1 rats
Females
150
NDr
NF
NF
Increased relative heart weight
Females
40
80
46b
15b
Increased relative right kidney
weight
Females
150
NDr
NF
NF
Increased relative liver
weightd
Females
80
150a
24b
91°
13b
56°
Increased relative spleen
weight
Females
40
80
NF
NF
aChange was >10% compared to control values.
bBMR of 1 standard deviation relative risk.
°BMR of 10% relative risk.
dBased on language from U.S. EPA's Benchmark Dose Technical Guidance Document (U.S. EPA. 20121 that states,
"For consistency in reporting, the BMD corresponding to a one control SD shift in the control mean should always
be presented along with the BMDs and BMDL for whatever BMR is being used for the POD, " the BMD(L)s with
BMRs of 1SD and 10% are both shown.
NDr = not determined; NF = no fit.
Dosimetric Adjustments:
Because the NTP (2012) study is a continuous feed study, dosimetric adjustments to
convert to an adjusted daily dose were not needed.
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In the U.S. EP A's Recommended Use of Body Weight3/4 as the Default Method in
Derivation of the Oral Reference Dose (U.S. EPA. 2011). the U.S. EPA endorses a hierarchy of
approaches to derive human equivalent oral exposures from data from laboratory animal species,
with the preferred approach being physiologically based pharmacokinetic modeling. Other
approaches may include using some chemical-specific information without a complete
physiologically based pharmacokinetic model. In lieu of chemical-specific models or data to
inform the derivation of human equivalent oral exposures, the U.S. EPA endorses body-weight
scaling to the 3/4 power (i.e., BW3/4) as a standard method to extrapolate toxicologically
equivalent doses of orally administered agents from all laboratory animals to humans for the
purpose of deriving an RfD under certain exposure conditions. More specifically, the use of
BW3/4 scaling for deriving an RfD is recommended when the observed effects are associated
with the parent compound or a stable metabolite, but not for portal-of-entry effects or
developmental toxicity endpoints. A validated human physiologically based pharmacokinetic
model for SAN Trimer is not available for use in extrapolating doses from animals to humans.
The selected critical effect of increased absolute liver weight is associated with the parent
compound or a stable metabolite. Furthermore, this liver effect is not a portal-of-entry or
developmental toxicity effect. Therefore, scaling by BW3/4 is relevant for deriving human
equivalent doses (HEDs) for this effect.
Following U.S. EPA (2011) guidance, the POD for increased absolute liver weight in
adult animals is converted to a HED through application of a dosimetric adjustment factor
(DAF)2 derived as follows:
DAF = (BWa1/4 - BWh1/4)
where
DAF = dosimetric adjustment factor
BWa = animal body weight
BWh = human body weight
Using a BWa of 0.25 kg for rats and a BWh of 70 kg for humans (U.S. EPA. 1988). the
resulting DAF is 0.24. Applying this DAF to the BMDLio of 10 mg/kg-day identified for the
critical effect in rats yields a BMDLiohed as follows:
BMDLiohed = 10 mg/kg-day x DAF
= 10 mg/kg-day x 0.24
= 2.4 mg/kg-day
The subchronic p-RfD for SAN Trimer is derived as follows:
Subchronic p-RfD = BMDLiohed ^ UFc
= 2.4 mg/kg-day -^300
= 8 x 10"3 mg/kg-day
2As described in detail in Recommended Use of Body Weight3/4 as the Default Method in Derivation of the Oral
Reference Dose U.S. EPA (20111. rate-related processes scale across species in a manner related to both the direct
(BWm) and allometric scaling (BW3/4) aspects such that BW3/4 ^ BW1/1= BW ' I converted to a
DAF = BWa1'4 - BWh1'4.
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The composite uncertainty (UFc) for the subchronic p-RfD for SAN Trimer is 300, as
summarized in Table 7. Table 8 provides the confidence descriptors for the subchronic p-RfD.
Table 7. UFs for Subchronic p-RfD of SAN Trimer
UF
Value
Justification
UFa
3
A UFa of 3 (100 5) has been applied to account for uncertainty in characterizing the toxicokinetic
or toxicodynamic differences between rats and humans following oral SAN Trimer exposure.
The toxicokinetic uncertainty has been accounted for by calculation of a human equivalent dose
(HED) through application of a dosimetric adjustment factor (DAF) as outlined in the U.S.
EPA's Recommended Use of Body Weight3/4 as the Default Method in Derivation of the Oral
Reference Dose CU.S. EPA. 2011).
UFd
10
A UFd of 10 has been applied for deficiencies in the available database because there are no
acceptable two-generation reproductive or developmental toxicity studies for SAN Trimer via the
oral route. The NTP (2012) studv desisn did not satisfv the reauirements for either of these studv
types.
UFh
10
A UFh of 10 has been applied for inter-individual variability to account for human-to-human
variability in susceptibility in the absence of quantitative information to assess the toxicokinetics
and toxicodynamics of SAN Trimer in humans.
UFl
1
A UFl of 1 has been applied for LOAEL-to-NOAEL extrapolation because the POD is a
BMDLio.
UFS
1
A UFS of 1 has been applied because a subchronic-duration study was selected as the principal
study.
UFC
300
Composite Uncertainty Factor = UFA x UFD x UFH x UFL x UFS
Table 8. Confidence Descriptors for Subchronic p-RfD for SAN Trimer
Confidence Categories
Designation3
Discussion
Confidence in study
H
The studv bv the NTP (2012) is a well-conducted. Deer-reviewed.
GLP-compliant, and comprehensive study with a sufficient number of
animals that examined a variety of endpoints.
Confidence in database
M
The database for SAN Trimer includes comprehensive acute, short-term-,
subchronic-, and chronic-duration toxicity studies but is lacking
acceptable two-generation reproductive and developmental toxicity
studies.
Confidence in subchronic
p-RfDb
M
The overall confidence in the subchronic p-RfD is medium.
aL = low, M = medium, H = high.
bThe overall confidence cannot be greater than lowest entry in table.
Derivation of a Chronic Provisional RfD (Chronic p-RfD)
The chronic-duration portion of the NTP (2012) study in F344/N rats is selected as
the principal study for derivation of the chronic p-RfD. In this study, groups of F1 rats
(50/sex/dose) from SAN Trimer-treated dams were fed diets containing SAN Trimer for 2 years
(104 weeks) postweaning. This study is a peer-reviewed technical report and was conducted
according to GLP standards. It is well conducted and meets the standard of study design and
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performance with numbers of animals, examination of potential toxicity endpoints, and
presentation of information (details are provided in the Review of Potentially Relevant Data
section), and is the only available study that reports effects following chronic-duration
SAN Trimer treatment.
As discussed in the Review of Potentially Relevant Data section, pathological effects
were reported in the following organs in F1 rats following chronic-duration SAN Trimer
treatment (see Table A-12): liver (>20 mg/kg-day), bone marrow (>40 mg/kg-day), bladder
(85 mg/kg-day), and the peripheral nervous system (>40 mg/kg-day) (NTP. 2012). An attempt
was made to model these data using the U.S. EPA's BMDS version 2.2 (as described
Appendix B) for consideration as potential PODs (see Table 9). The most sensitive of these
effects is increased incidence of chronic active inflammation in the liver of males with a
BMDLio of 3.2 mg/kg-day (see Table 9). Although liver weight was not examined in the NTP
(2012) 2-year study, support for chronic active inflammation in the liver as the critical effect for
derivation of the chronic p-RfD is provided by the consistent observation of liver effects in male
and female F344/N rats in the 2-week, 13-week, and 104-week studies performed by the NTP
(NTP. 2012). as well as male and female S-D rats in the 2-week study by (NTP. 2012;
Huntingdon Life Sciences. 1999b).
There were also effects in the nervous system (i.e., spinal root degeneration in males and
sciatic nerve degeneration in females) that were statistically significantly increased in rats in the
2-year study NTP (2012). These effects were also modeled and the PODs are very similar to the
BMDLio of 3.2 mg/kg-day for increased incidence of chronic active hepatic inflammation in
male rats. As shown in Table 9, the BMDLio for spinal root degeneration in males is
4.9 mg/kg-day and the BMDLio for sciatic nerve degeneration in females is 3.5 mg/kg-day.
Therefore, sciatic nerve degeneration in female rats and chronic active inflammation in the liver
of male rats can be considered as co-critical effects due to the similarity in their BMDLs. In
addition, the majority recommendation of the external peer reviewers was that these effects
should be considered co-critical effects. Thus, the increased incidences of chronic active
inflammation in the liver of male rats and sciatic nerve degeneration in females are chosen
as co-critical effects, and the BMDLio of 3.2 mg/kg-day is chosen as the POD. The selection
of the BMDLio of 3.2 mg/kg-day as the POD will protect against both liver and nervous system
effects and also other chronic-duration effects that occurred in the bladder and bone marrow of
rats (see Table 9).
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Table 9. Candidate Chronic PODs: Effect Levels From Histopathological Endpoints in
F344/N Rats Treated with SAN Trimer by Diet for 2 Years3
Lesion
NOAEL
LOAEL
BMDio
BMDLio
Peripheral Nervous System
Sciatic nerve degeneration, females
20
40
4.7
3.5
Spinal root degeneration, males
40
75
67
4.9
Liver
Chronic active inflammation, males
40
75
8.0
3.2
Eosinophilic foci, males
40
75
17
13
Mixed cell foci, females
40
85
41
23
Mixed cell foci, males
None
20
8.5b
3.7b
Angiectasis, males
40
75
NF
NF
Bone Marrow
Bone marrow hyperplasia, females
20
40
13
3.6
Inflammation, granulomatous, females
20
40
40
34
Bone marrow hyperplasia, males
40
75
54
18
Bladder
Hyperplasia of transitional epithelium, females
40
85
80
62
aNTP (20121.
bData for incidence of mixed cell foci in male rats were modeled with BMDS and the mid- and high-dose groups
were removed from the analysis because of a plateau effect observed when the data were modeled with all doses.
NF = no fit (none of the BMD models adequately fit the data).
Dosimetric Adjustments:
Because the NTP (2012) study is a continuous feed study, dosimetric adjustments to
convert to an adjusted daily dose were not needed. As discussed in the subchronic p-RfD
derivation section above, the U.S. EPA endorses body-weight scaling to the 3/4 power
(i.e., BW3/4) as a standard methodology to extrapolate toxicologically equivalent doses of orally
administered agents from all laboratory animals to humans for the purpose of deriving an RfD
under certain exposure conditions (U.S. EPA. 2011). The POD for chronic active inflammation
in the liver of male rats is converted to a HED through application of a DAF derived as follows:
DAF = (BWa1/4 - BWh1/4)
where
DAF = dosimetric adjustment factor
BWa = animal body weight
BWh = human body weight
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Using a BWa of 0.25 kg for rats and a BWh of 70 kg for humans (U.S. EPA. 1988). the
resulting DAF is 0.24. Applying this DAF to the BMDLio of 3.2 mg/kg-day identified for the
co-critical effect of chronic active inflammation in the liver of male rats yields a BMDLiohed as
follows:
BMDLiohed = 3.2 mg/kg-day x DAF
= 3.2 mg/kg-day x 0.24
= 0.77 mg/kg-day
The chronic p-RfD for SAN Trimer is derived as follows:
Chronic p-RfD = BMDLiohed ^ UFC
= 0.77 mg/kg-day ^ 300
= 3 x 10"3 mg/kg-day
The UFc for the chronic p-RfD for SAN Trimer is 300, as summarized in Table 10.
Table 11 provides the confidence descriptors for the chronic p-RfD.
Table 10. UFs for Chronic p-RfD of SAN Trimer
UF
Value
Justification
UFa
3
A UFa of 3 (10°5) has been applied to account for uncertainty in characterizing the toxicokinetic or
toxicodynamic differences between rats and humans following oral SAN Trimer exposure. The
toxicokinetic uncertainty has been accounted for by calculation of a human equivalent dose (HED)
through application of a dosimetric adjustment factor (DAF) as outlined in the U.S. EPA's
Recommended Use of Body Weight3/4 as the Default Method in Derivation of the Oral Reference
Dose CU.S. EPA. 20111
UFd
10
A UFd of 10 has been applied for deficiencies in the available database because there are no
acceptable two-generation reproductive or developmental toxicity studies for SAN Trimer via the
oral route. The NTP (2012) studv desisn did not satisfv the reauirements for either of these studv
types.
UFh
10
A UFh of 10 has been applied for inter-individual variability to account for human-to-human
variability in susceptibility in the absence of quantitative information to assess the toxicokinetics
and toxicodynamics of SAN Trimer in humans.
UFl
1
A UFl of 1 has been applied for LOAEL-to-NOAEL extrapolation because the POD is a BMDLio.
UFS
1
A UFS of 1 has been applied because a chronic-duration study was selected as the principal study.
UFC
300
Composite Uncertainty Factor = UFA x UFD x UFH x UFL x UFS
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Table 11. Confidence Descriptors for Chronic p-RfD for SAN Trimer
Confidence Categories
Designation3
Discussion
Confidence in study
H
The studv bv the NTP (2012) is a well-conducted. Deer-reviewed.
GLP-compliant, and comprehensive study with a sufficient number of
animals that examined a variety of endpoints.
Confidence in database
M
The database for SAN Trimer includes comprehensive acute, short-term-,
subchronic-, and chronic-duration toxicity studies but is lacking
acceptable two-generation reproductive and developmental toxicity
studies.
Confidence in chronic
p-RfDb
M
The overall confidence in the chronic p-RfD is medium.
aL = low, M = medium, H = high.
bThe overall confidence cannot be greater than lowest entry in table.
DERIVATION OF PROVISIONAL INHALATION REFERENCE CONCENTRATIONS
No information that could be used to derive subchronic or chronic p-RfCs for
SAN Trimer was identified.
CANCER WEIGHT-OF-EVIDENCE (WOE) DESCRIPTOR
The Guidelines for Carcinogen Risk Assessment (Cancer Guidelines) (U.S. EPA. 2005)
emphasize the importance of applying a weight-of-evidence (WOE) approach in reaching
conclusions about the carcinogenic potential of chemicals in humans. Each cancer descriptor
may be applicable to a variety of potential data sets and represent points along a continuum of
evidence. The available carcinogenic evidence for SAN Trimer could be considered a borderline
case between two potential descriptors— suggestive evidence of carcinogenic potential and
inadequate information to assess carcinogenic potential. For example, oral treatment with
SAN Trimer caused a non-statistically significant increase in the incidences of brain and spinal
cord astrocytomas and granular cell tumors in male rats, which is consistent with one of the
examples provided in the Cancer Guidelines (U.S. EPA. 2005) for the descriptor suggestive
evidence of carcinogenic potential. The example states that supporting data for this descriptor
may include "a small, and possibly not statistically significant, increase in tumor incidence
observed in a single animal or human study that does not reach the weight of evidence for the
descriptor 'Likely to Be Carcinogenic to Humans.However, because there was not a
statistically significant increase in any relevant tumor type in rats treated with SAN Trimer for
2 years (NTP. 2012). some tumor types (e.g., pituitary gland adenomas, mammary gland tumors,
etc.) were statistically significantly decreased in male and/or female rats, and the carcinogenic
potential of SAN Trimer has only been evaluated in one animal species in one study (NTP.
2012). the descriptor indicating that there is "inadequate information to assess the carcinogenic
potential is also relevant. Finally, whereas children exposed to drinking water containing
SAN Trimer exhibited a statistically significant elevation in various tumor types (NJ DHSS.
1997). it is not possible to determine if SAN Trimer solely contributed to these cancer effects in
children because other chemicals that have been shown to demonstrate carcinogenic potential
(e.g., trichloroethylene, tetrachloroethylene) were also identified in the drinking water
(Richardson et al.. 1999).
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The evidence from the only available carcinogenicity study in laboratory animals shows
increased incidences of testicular tumors, brain and spinal cord astrocytomas and granular cell
tumors (see Table 12) in male F344/N rats (Fl) treated perinatally and postnatally with
SAN Trimer for 2 years (NTP. 2012). There was a statistically significant increase in the
incidence of total testicular tumors; however, the high background incidence in the control group
(41/50) indicates that these tumors may not be treatment-related. This tumor has been identified
as the most frequently observed spontaneous tumor in the male F344/N rat (Haseman et al..
1990; Takaki et al.. 1989). Therefore, the incidence of total testicular tumors is not considered in
the WOE for the carcinogenic potential of SAN Trimer. In addition, the incidences of brain and
spinal cord astrocytomas and granular cell tumors in male rats were not statistically significantly
increased at any dose level compared to concurrent controls. Furthermore, as pointed out by two
of the five external peer reviewers of this PPRTV assessment, data presented by Sills et al.
(1999) have shown brain astrocytoma rates in F334/N control rats fed the NIH-07 diet in several
NTP studies to be up to 4% in males, whereas brain granular cell tumor rates have been up to 2%
in males. In the NTP (2012) study, brain astrocytomas and granular cell tumors occurred in 2%
of male rats at the mid- and high-dose levels (see Table 12). These data indicate that, although
rare, the brain tumors observed in male rats in the NTP (2012) study are within the range of
historical controls as reported by Sills et al. (1999). Based on these results from the NTP (2012)
study, the NTP concluded that there was no evidence of carcinogenic activity for SAN Trimer.
In evaluating this borderline case, the U.S. EPA considered Section 2.5 of the Cancer
Guidelines which states that the descriptor suggestive evidence of carcinogenic potential is
appropriate when "the weight of evidence is suggestive of carcinogenicity, a concern for
potential carcinogenic effects is raised, but the data are not judged sufficient for a stronger
conclusion." The Cancer Guidelines further state that the descriptor inadequate information to
assess carcinogenic potential is appropriate when "available data are judged inadequate for
applying one of the other descriptors." Although either descriptor could be considered plausible,
this PPRTV assessment attaches greater weight to the fact there is only one cancer study in one
species (NTP. 2012) that reported mostly negative tumor findings and no statistically significant
increase in any relevant tumor type in male and female rats. Although there may be a similar
observation of the occurrence of brain/CNS astrocytomas between humans exposed to drinking
water containing SAN Trimer among many other chemicals (NJ DHSS. 1997) and male rats
following oral exposure to SAN Trimer (NTP. 2012). the incidence of brain and spinal cord
astrocytomas was not statistically significantly increased in male rats compared to concurrent
controls, and the observed incidences of these brain tumors are within the historical range of data
presented in a previous publication (Sills et al.. 1999). Thus under the U.S. EPA's Cancer
Guidelines (U.S. EPA. 2005) and in concurrence with the majority recommendation of the
external peer reviewers for this PPRTV assessment, there is inadequate information to assess
carcinogenic potential for SAN Trimer.
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Table 12. Incidences of Selected Cancer Endpoints in Male F344/N Rats Administered
SAN Trimer by Diet for 2 Years"
Exposure Group (Human Equivalency Dose, mg/kg-day)b
0
20
40
75
Parameter
(0)
(5.6)
(11)
(20)
Sample size
50
50
50
50
Brain
Astrocytomas0
0
0
1(2)
1(2)
Granular cell tumors0
0
0
1(2)
1(2)
Spinal Cord
Astrocytomas0
0
0
0
1(2)
Granular cell tumors0
0
1
0
0
Testes
Total, interstitial cell, adenoma (bilateral + unilateral,
41 (82)
49 (98)d
44 (88)
49 (98)d
combined)00
aNTP (20121.
bDoses were converted from adjusted daily doses to human equivalency doses using the following formula:
DoseHED = DoseADi x (Body Weight Animal ^ Body Weight Human)(0 25).
incidence, (corresponding percentage).
Significantly different from control (p < 0.05) using the Poly-3 test, as reported by the study authors.
"Statistically significant trend using Poly-3 test for dose-response relationship in male rats, as reported by the study
authors.
DERIVATION OF PROVISIONAL CANCER POTENCY VALUES
Derivation of Provisional Oral Slope Factor (p-OSF)
Because there are inadequate data to determine human carcinogenic potential, the
derivation of an oral slope factor is precluded. The decision to not develop a quantitative
estimate of cancer risk from oral exposure to SAN Trimer was supported by the majority of the
external reviewers.
Derivation of Provisional Inhalation Unit Risk (p-IUR)
No human or animal studies examining the carcinogenicity of SAN Trimer following
inhalation exposure have been identified. Therefore, derivation of a p-IUR is precluded.
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APPENDIX A. FIGURES AND DATA TABLES
Lakewood
.Point Pleasant
Toms'
Crestwood Village
Ocean County
Figure A-l. Map of Ocean County, Dover Township (currently Toms River Township) and
the Toms River section of Dover Township (currently the sub-Township area) New Jersey
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Table A-l. Summary of Statistically Significantly Elevated SIRs for Ocean Countya'b
1979-1995
Cancer Type
Age Group
Sex
Race
Number Observed
Number Expected
SIR
95% CI Lower-Upper
Brain/CNS astrocytoma
0-19
Both
All
35
23.9
1.46*
1.02-2.03
PNS neuroblastoma
0-19
Both
All
27
15.9
1.70*
1.12-2.47
PNS neuroblastoma
0-4
Both
All
24
13.3
1.81*
1.16-2.69
PNS neuroblastoma
0-19
Male
All
20
9.1
2.20*
1.34-3.40
PNS neuroblastoma
0-4
Male
All
17
7.7
2.21*
1.29-3.54
aNJ DHSS (19971.
incidence data for Ocean County were not reported in the later studies (NJ DHSS. 2008. 2003b).
* Statistically elevated, p < 0.05.
CNS = Central Nervous System; PNS = Peripheral Nervous System; SIR = Standardized Incidence Ratio.
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Table A-2. Summary of Statistically Significantly Elevated SIRs for Dover/Toms River Township
Cancer Type
Age Group
Sex
Race
Number Observed
Number Expected
SIR
95% CI Lower-Upper
1979-1995
All types3
0-19
Both
All
90
67.0
1.34* (1.3*)
1.08-1.65 (1.04-1.6)
All types3
0-19
Female
All
47
33.3
1.41* (1.4*)
1.04-1.88 (1.01-1.85)
All types3
0-4
Female
All
16
8.4
1.90* (1.8)
1.09-3.09 (0.99-3.04)
Acute lymphocytic leukemia3
0-19
Female
All
12
4.6
2.59* (2.6*)
1.34-4.53 (1.34-4.53)
Acute lymphocytic leukemia3
0-4
Female
All
7
2.2
3.27* (3.3*)
1.31-6.73 (1.30-6.70)
Leukemia
0-19
Female
All
13
6.5
1.99*
1.06-3.40
1996-2000
All types
0-19
Both
All
25
20.6
1.2
0.78-1.79
All types
0-19
Female
All
13
9.4
1.4
0.74-2.37
All types
0-4
Female
All
3
2.2
1.4
0.28
Acute lymphocytic leukemia
0-19
Female
All
3
1.5
2.0
0.40-5.83
Acute lymphocytic leukemia
0-4
Female
All
2
0.7
2.9
0.32-10.4
Leukemia
0-19
Female
All
4
2.0
2.0
0.53-5.06
2001-2005
All types
0-19
Both
All
26
22.8
1.1
0.75-1.7
All types
0-19
Female
All
14
10.4
1.3
0.73-2.3
All types
0-4
Female
All
2
2.6
0.8
0.09-2.8
Acute lymphocytic leukemia
0-19
Female
All
1
1.6
NC
NC
Acute lymphocytic leukemia
0-4
Female
All
0
0.6
NC
NDr
Leukemia
0-19
Female
All
1
1.6
NC
NC
aNumbers in parentheses are updated from NJDHSS (2003b).
* Statistically elevated, p < 0.05
NC= Not calculated by the study authors; NDr = Not determined.
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Table A-3. Summary of Statistically Significantly Elevated SIRs for Toms River Section/Sub-Township
Cancer Type
Age Group
Sex
Race
Number Observed
Number Expected
SIR
95% CI Lower-Upper
1979-1995
All types
0-19
Both
All
24 (24)
14.1 (144)
1.70* (1.7*)
1.09-2.53 (1.07-2.49)
All types
0-4
Both
All
13 (12)
3.5 (3.4)
3.73* (3.6*)
1.99-6.38 (1.84-6.22)
All types
0-4
Female
All
10(10)
1.6(1.5)
6.17* (6.5*)
2.95-11.34 (3.13-12.0)
Brain/CNS
0-4
Both
All
4(4)
0.6 (0.6)
7.04* (7.0*)
1.89-18.03 (1.87-17.8)
Brain/CNS
0-4
Female
All
3(3)
0.3 (0.3)
11.60* (11.3*)
2.33-33.88 (2.27-33.0)
Brain/CNS astrocytoma
0-4
Both
All
2
0.2 (0.2)
9.47* (8.9*)
1.06-34.19 (1.00-32.1)
Acute lymphocytic leukemia
0-4
Female
All
4
0.4 (0.4)
9.68* (9.4*)
2.60-24.78 (2.52-24.0)
Leukemia
0-4
Female
All
4
0.5
7.84*
2.11-20.06
1996-2000
All types
0-19
Both
All
5
4.2
1.2
0.39-2.81
All types
0-4
Both
All
0
1.1
NC
NDr
All types
0-4
Female
All
0
0.4
NC
NDr
Brain/CNS
0-4
Both
All
0
0.2
NC
NDr
Brain/CNS
0-4
Female
All
0
0.1
NC
NDr
Brain/CNS astrocytoma
0-4
Both
All
0
0.1
NDr
NDr
Acute lymphocytic leukemia
0-4
Female
All
0
0.1
NDr
NDr
Leukemia
0-4
Female
All
0
0.2
NDr
NDr
2001-2005
All types
0-19
Both
All
5
4.6
1.1
0.35-2.5
All types
0-4
Both
All
2
1.3
1.6
0.18-5.8
All types
0-4
Female
All
1
0.5
NC
NC
Brain/CNS
0-4
Both
All
1
0.2
NC
NC
Brain/CNS
0-4
Female
All
0
0.1
NC
NDr
Brain/CNS astrocytoma
0-4
Both
All
0
0.1
NC
NDr
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Table A-3. Summary of Statistically Significantly Elevated SIRs for Toms River Section/Sub-Township
Cancer Type
Age Group
Sex
Race
Number Observed
Number Expected
SIR
95% CI Lower-Upper
2001-2005
Acute lymphocytic leukemia
0-4
Female
All
0
0.1
NC
NDr
Leukemia
0-4
Female
All
0
0.2
NC
NDr
"Numbers in parentheses are updated from NJ DHSS (2003b').
* Statistically elevated,/) < 0.05.
CNS = Central Nervous System; NC= Not calculated by study authors; NDr= Not determined; SIR = Standardized Incidence Ratio.
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Table A-4. Mean Body Weights in Dams from 2-Week Toxicity Study of SAN Trimer"
Endpoint
Exposure Concentration in ppm (mg/kg-day)
0
250 (18)
500 (35)
1,000 (67)
2,000 (130)
4,000 (197)
Gestation
No. of animals'3
8
8
8
7
8
7
GD 1 body weight (g) (%)d
163
169 (|4)
161(41)
169 (|4)
166 (|2)
170 (|4)
GD 7 body weight (g) (%)d
180
185 (|3)
176 (|2)
187 (|4)
181 (|1)
187 (|4)
GD 14 body weight (g) (%)d
203
207 (|2)
195 (44)
207 (|2)
199 (42)
181** (411)
GD 18 body weight (g) (%)d
223
231 (|4)
217(43)
233 (|4)
226 (|1)
204* (48)
Endpoint
Exposure Concentration in ppm (mg/kg-day)
0
250 (40)
500 (85)
1,000 (166)
2,000 (325)
4,000 (634)
Postnatal
No. of animalsb
7
T
7
7
8
6
PND 1 body weight (g) (%)d
199
206 (|4)
193 (43)
202 (|2)
195 (42)
177** (411)
PND 7 body weight (g) (%)d
210
221 (|5)
206 (42)
219 (|4)
209 (0)
184** (412)
PND 14 body weight (g) (%)d
234
239 (|2)
225 (44)
236 (|1)
223 (45)
177** (424)
PND 20 body weight (g) (%)d
235
237 (|1)
227 (43)
241 (|3)
222 (46)
174** (426)
aNTP (20121.
bNo. of animals weighed on each day.
°Eight dams were weighed on PND 1.
dMean (% change from control).
*Dunnett's testp < 0.05.
**Dunnett's testp < 0.01.
GD = Gestation Day; PND = Postnatal Day.
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Table A-5. Mean Body Weight in F1 Pups from 2-Week Toxicity Study of SAN Trimer"
Endpoint
Exposure Concentration in ppm (mg/kg-day)
0
250 (40)
500 (85)
1,000 (166)
2,000 (325)
4,000 (634)
Male
No. of animals'3
39
30
33
31
38
27
PND 1 body weight (g) (%)
5.8
5.9 (|2)
6.0 (|3)
5.8 (0)
5.8 (0)
5.3** (49)
No. of animals0
10
10
10
10
10
10
PND 4 body weight (g) (%)d
8.8
9.0 (|2)
8.6 (|2)
84 (45)
8.8 (0)
7.5** (415)
PND 7 body weight (g) (%)d
13.5
13.6 (tl)
13.0 (|4)
13.0 (44)
12.9 (44)
10.4** (423)
PND 14 body weight (g) (%)d
25.7
26.2 (|2)
25.1 (42)
24.9 (43)
24.6 (44)
16.8** (435)
PND 20 body weight (g) (%)d
35.0
35.8 (|2)
34.7 (41)
34.5 (41)
31.6** (410)
19.8** (443)
Female
No. of animals'3
23
34
32
40
49
31
PND 1 body weight (g) (%)d
5.4
5.6 (|4)
5.4 (0)
5.5 (|2)
5.3 (42)
5.0** (47)
No. of animals0
10
10
10
10
10
10
PND 4 body weight (g) (%)d
8.2
8.5 (|4)
8.2 (0)
8.0 (42)
8.2 (0)
7.3* (411)
PND 7 body weight (g) (%)d
12.7
12.9 (|2)
12.7 (0)
12.4 (42)
12.4 (42)
9.9** (422)
PND 14 body weight (g) (%)d
24.7
24.7 (0)
25.0 (|1)
24.2 (42)
23.7 (44)
16.1** (435)
PND 20 body weight (g) (%)d
33.6
33.9 (|1)
33.6 (0)
32.7 (43)
30.3** (410)
18.8** (444)
aNTP (20121.
bNo. of animals weighed on PND 1.
°No. of animals weighed on PND 4, 7, 14, and 20.
dMean (% change from control).
*Dunnett's testp < 0.05.
**Dunnett's testp < 0.01.
PND = Postnatal Day.
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Table A-6. Absolute and Relative Organ Weights in Male and Female F1 Rats from 2-Week Toxicity Study of SAN Trimera
Endpoint
Exposure Concentration in ppm
0
250
500
1,000
2,000
4,000
Males
mg/kg-d
0
50
90
175
270
410
Number of animals
10
10
10
10
10
9
Necropsy body weight
(g) (%)b
118 ± 6
114 ±4 (|3)
109 ±4 (48)
106* ±2 (410)
74** ± 3 (437)
34** ±2 (471)
Absolute Heart (g) (%)b
0.52 ±0.02
0.51 ±0.02 (|2)
0.48 ±0.02 (48)
0.47* ±0.01 (410)
0.34** ± 0.01 (435)
0.34** ±0.02 (435)
Relative Heart Ratio
(%)b
4.395 ±0.062
4.478 ±0.061 (|2)
4.392 ±0.083 (0)
4.424 ±0.060 (|1)
4.564 ± 0.069 (|4)
10.297** ± 1.002 (f 134)
Absolute right kidney
(g)(%)b
0.60 ±0.03
0.58 ±0.02 (43)
0.56 ±0.02 (47)
0.55* ±0.01 (48)
0.39** ± 0.01 (435)
0.25** ±0.01 (458)
Relative right kidney
ratio (%)b
5.098 ±0.082
5.063 ±0.067 (41)
5.134 ±0.073 (|1)
5.158 ± 0.112 (|1)
5.346 ±0.092 (|5)
7.380** ±0.233 (|45)
Absolute liver (g) (%)b
5.91 ±0.34
6.21 ±0.35 (|5)
5.69 ±0.27 (44)
6.02 ±0.13 (|2)
3.89** ±0.13 (434)
2.34** ±0.10 (460)
Relative liver ratio (%)b
49.876 ± 0.756
54.119* ± 1.370 (|9)
52.179* ±0.592 (|5)
56.806** ± 1.009
(t 14)
52.935** ±0.825 (|6)
68.468** ± 1.307 (|37)
Absolute lung (g) (%)b
0.92 ±0.03
0.88 ±0.04 (44)
0.85 ±0.03 (48)
0.86 ±0.03 (47)
0.60** ±0.02 (435)
0.45** ±0.05 (451)
Relative lung ratio (%)b
7.896 ±0.289
7.762 ± 0.250 (42)
7.817 ±0.259 (41)
8.064 ± 0.245 (|2)
8.208 ±0.214(|4)
12.827** ±0.892 (|162)
Absolute spleen (g) (%)b
0.385 ±0.013
0.364 ±0.015 (45)
0.348 ±0.014 (410)
0.346 ±0.006 (410)
0.334 ±0.033 (413)
0.255** ±0.008 (434)
Relative spleen ratio
(%)b
3.281 ±0.069
3.186 ±0.065 (43)
3.211 ±0.070 (42)
3.269 ±0.067 (0)
4.461** ±0.327 (|36)
7.587** ±0.532 (|231)
Absolute right testis (g)
(%)b
0.561 ±0.037
0.535 ±0.043 (45)
0.451* ±0.037 (420)
0.440** ±0.019 (422)
0.237** ±0.017 (458)
0.069** ±0.003 (488)
Relative right testis ratio
(%)b
4.724 ±0.159
4.644 ± 0.260 (42)
4.121* ± 0.234 (413)
4.145* ±0.150 (412)
3.207** ±0.175 (432)
2.037** ±0.067 (457)
Absolute thymus (g)
0.378 ±0.018
0.348 ±0.019 (48)
0.306** ±0.011 (419)
0.298** ±0.009 (421)
0.174** ±0.007 (454)
0.045** ±0.004 (488)
57
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Table A-6. Absolute and Relative Organ Weights in Male and Female F1 Rats from 2-Week Toxicity Study of SAN Trimera
Endpoint
Exposure Concentration in ppm
0
250
500
1,000
2,000
4,000
Relative thymus ratio
(%)b
3.212 ±0.085
3.035 ±0.064 (|6)
2.832** ±0.092 (412)
2.810** ±0.063 (413)
2.371** ±0.060 (426)
1.304** ±0.086 (459)
Females
mg/kg-d
0
45
90
185
295
430
Number of animals
10
10
10
10
10
10
Necropsy body weight
(g) (%)b
106 ±5
103 ±3 (|3)
101 ±3 (45)
102 ± 2 (44)
72** ± 3 (432)
32** ± 1 (470)
Absolute heart (g) (%)b
0.46 ± 0.02
0.47 ±0.01 (|2)
0.45 ±0.01 (42)
0.46 ±0.01 (0)
0.32** ±0.01 (430)
0.28** ±0.01 (439)
Relative heart ratio (%)b
4.333 ±0.106
4.609 ±0.103 (|6)
4.465 ±0.093 (|3)
4.521 ±0.057 (|4)
4.493 ±0.052 (|4)
8.822** ±0.601 (|91)
Absolute right kidney
(g)(%)b
0.55 ±0.03
0.54 ±0.02 (42)
0.52 ±0.02 (45)
0.55 ±0.01 (0)
0.38** ±0.02 (431)
0.23** ±0.01 (458)
Relative right kidney
Ratio (%)b
5.171 ±0.086
5.232 ±0.060 (|1)
5.207 ±0.082 (|1)
5.449 ±0.073 (|5)
5.267 ± 0.052 (|2)
7.200** ±0.166 (|39)
Absolute liver (g) (%)b
5.12 ±0.28
5.38 ±0.20 (|5)
5.13 ±0.14(0)
5.84 ± 0.14 (|14)
3.83** ±0.20 (425)
2.07** ±0.10 (460)
Relative liver ratio (%)b
48.351 ±0.891
52.056* ±0.832 (|8)
50.916* ±0.478 (|5)
57.438** ±0.602
(t 19)
52.956** ± 1.362 (|10)
64.269** ± 1.124 (|33)
Absolute lung (g) (%)b
0.79 ±0.04
0.90 ±0.04 (f 14)
0.83 ±0.04 (|5)
0.87 ±0.04 (|10)
0.57** ±0.02 (428)
0.38** ±0.01 (452)
Relative lung ratio (%)b
7.534 ±0.279
8.801 ± 0.418 (|17)
8.208 ±0.285 (|9)
8.509 ±0.336 (|13)
7.971 ±0.303 (|6)
11.886** ±0.581 (|58)
Absolute spleen (g) (%)b
0.334 ±0.016
0.334 ±0.013 (0)
0.317 ±0.006 (45)
0.338 ±0.006 (|1)
0.300 ±0.028 (410)
0.223** ±0.007 (433)
Relative spleen ratio
(%)b
3.170 ±0.073
3.233 ±0.081 (|2)
3.154 ±0.064 (41)
3.331 ±0.031 (|5)
4.122 ±0.253** (|30)
7.072 ±0.386** (|123)
Absolute thymus (g)
(%)b
0.360 ±0.015
0.336 ±0.010 (47)
0.332 ±0.012 (48)
0.330 ±0.013 (48)
0.188** ±0.011 (448)
0.043** ±0.005 (488)
Relative thymus ratio
(%)b
3.438 ±0.117
3.270 ±0.081 (45)
3.300 ±0.093 (44)
3.236 ±0.085 (46)
2.627** ±0.150 (424)
1.310** ±0.100 (462)
58
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Table A-6. Absolute and Relative Organ Weights in Male and Female F1 Rats from 2-Week Toxicity Study of SAN Trimera
Endpoint
Exposure Concentration in ppm
0
250
500
1,000
2,000
4,000
Absolute uterus (g) (%)b
0.165 ±0.020
0.202 ± 0.029 (|22)
0.176 ±0.017(17)
0.161 ±0.022 (|2)
0.052** ±0.005 (|68)
0.023** ±0.002 (486)
Relative uterus ratio
(%)b
1.573 ±0.201
1.932 ±0.255 (|23)
1.746 ±0.162
(til)
1.600 ±0.226 (|2)
0.722** ±0.072 (454)
0.729** ±0.066 (454)
aNTP (20121.
bMean ± standard error (% change from control).
*Williams' testp < 0.05.
**Williams' testp < 0.01.
59
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Table A-7. Mean Body Weights in Dams from 13-Week Toxicity Study of SAN Trimer"
Endpoint
Dose Group ppm (mg/kg-day)
0
100 (6.9)
200 (14)
400 (28)
800 (55)
1,600 (110)
Gestation
No. of animals'3
9
9
8°
8
8
8
GD 1 body weight (g)
(%)d
187
178 (|5)
179 (44)
180 (44)
181(43)
183 (42)
GD 7 body weight (g)
(%)d
201
192 (|4)
194 (43)
191 (45)
193 (44)
195 (43)
GD 14 body weight (g)
(%)d
224
212 (45)
216 (44)
209* (47)
207* (48)
205* (48)
GD 20 body weight (g)
(%)d
268
254 (45)
259 (43)
245 (49)
243 (49)
230* (414)
Endpoint
Dose Group ppm (mg/kg-day)
0
100 (17)
200 (33)
400 (66)
800 (132)
1,600 (264)
Postnatal
No. of animalsb
8
8
7
6
6
4e
PND 1 body weight (g)
(%)d
217
209 (44)
211(43)
209 (44)
210(43)
209 (44)
PND 7 body weight (g)
(%)d
225
219(43)
224 (0)
221 (42)
218(43)
223 (41)
PND 14 body weight (g)
(%)d
252
243 (44)
247 (42)
240 (45)
243 (44)
243 (44)
PND 20 body weight (g)
(%)d
244
241 (41)
246 (|1)
239 (42)
245 (0)
240 (42)
aNTP (20121.
bNo. of animals weighed on each day.
°Nine dams were weighed on PND 20.
dMean (% change from control).
eThree dams were weighted on PND 20.
*Dunnett's testp < 0.05.
GD = Gestation Day; PND = Postnatal Day.
60
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Table A-8. Mean Body Weights of F1 Pups from 13-Week Toxicity Study of SAN Trimer"
Endpoint (n = 10)
Dose Group ppm (mg/kg-day)
0
100 (17)
200 (33)
400 (66)
800 (132)
1,600 (264)
Males
PND 4 BW (g) (%)b
8.4
8.8 (|5)
8.6 (|2)
8.8 (|5)
8.9 (|6)
8.5 (tl)
PND 7 BW (g) (%)b
12.8
12.7(41)
13.2 (|3)
13.3 (|4)
12.9 (|1)
12.7 (41)
PND 14 BW (g) (%)b
21.9
23.2 (1|6)
23.1 (|5)
23.0(t5)
22.5 (t3)
22.5 (t3)
PND 20 BW (g) (%)b
28.7
31.3 (|9)
30.7 (|7)
31.1 (|8)
30.0 (t5)
29.9 (t4)
Females
PND 4 BW (g) (%)b
8.2
8.5 (|4)
8.0 (|2)
8.3 (tl)
8.8 (t7)
8.3 (tl)
PND 7 BW (g) (%)b
12.4
12.4 (0)
12.1(42)
12.6 (|2)
12.8 (t3)
12.0 (43)
PND 14 BW (g) (%)b
21.3
22.7 (|7)
21.7 (|2)
21.9 (|3)
22.1 (t4)
21.4 (0)
PND 20 BW (g) (%)b
27.9
30.9* (fll)
28.9 (|4)
29.7 (|6)
29.4 (t5)
28.8 (t3)
aNTP (20121.
bMean (% change from control).
BW = Body Weight; PND = Postnatal Day.
61
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Table A-9. Absolute and Relative Organ Weights in Male and Female F1 Pups in the 13-Week Toxicity Study of SAN Trimer"
Endpoint
Dose Group ppm (mg/kg-day)
0
100 (10)
200 (20)
400 (40)
800 (80)
1,600 (150)
Males
Number of animals
10
10
10
10
10
10
Necropsy body weight (g)
(%)b
338 ±7
344 ± 3 (|2)
342 ±4 (t 1)
337 ± 4 (0)
335 ±6 (41)
302** ±5 (411)
Absolute brain (g) (%)b
1.951 ±0.017
1.964 ± 0.017 (|1)
1.990 ±0.020 (|2)
1.968 ±0.015 (|1)
1.988 ±0.019 (t2)
1.987 ± 0.010 (t2)
Relative brain ratio (%)b
5.785 ±0.097
5.715 ±0.083 (|1)
5.823 ±0.071 (|1)
5.846 ±0.037 (|1)
5.947 ±0.107 (t3)
6.588** ±0.098 (tl4)
Absolute heart (g) (%)b
0.91 ±0.02
1.00** ±0.01 (|10)
0.97 ± 0.02 (|7)
1.01** ±0.02 (fll)
0.98* ±0.02 (t8)
0.92 ±0.01 (tl)
Relative heart ratio (%)b
2.701 ±0.036
2.894** ±0.044 (|7)
2.830** ±0.042 (|5)
3.000** ±0.054 (fll)
2.935** ± 0.038 (t9)
3.049** ±0.034 (t 13)
Absolute R. kidney (g) (%)b
1.00 ±0.02
1.00 ±0.02 (0)
1.03 ±0.02 (|3)
1.04 ±0.02 (t4)
1.04 ±0.02 (t4)
1.01 ±0.03 (tl)
Relative R. kidney ratio
(%)b
2.956 ±0.041
2.897 ± 0.058 (|2)
3.004 ±0.033 (|2)
3.088 ±0.040 (t4)
3.099* ±0.052 (t5)
3.348** ±0.043 (tl3)
Absolute liver (g) (%)b
11.25 ±0.21
11.96* ± 0.17 (|6)
12.31** ±0.21 (|9)
12.53** ±0.25 (til)
13.13** ±0.34 (t 17)
12.68** ±0.24 (t 13)
Relative liver ratio (%)b
33.282 ±0.367
34.754* ±0.327 (|4)
35.998** ±0.446 (|8)
37.195** ±0.633 (tl2)
39.154** ±0.535 (tl8)
41.972** ±0.395
(T26)
Absolute lung (g) (%)b
1.99 ±0.07
1.92 ±0.07 (|4)
1.92 ±0.05 (44)
2.02 ±0.09 (t2)
2.11 ±0.09 (t6)
1.74 ±0.05 (413)
Relative lung ratio (%)b
5.879 ±0.173
5.597 ±0.215 (45)
5.631 ±0.175 (44)
6.006 ± 0.256 (t2)
6.281 ±0.213 (t7)
5.722 ±0.141 (43)
Absolute spleen (g) (%)b
0.733 ±0.014
0.780 ± 016 (|6)
0.764 ± 0.023 (|4)
0.786 ±0.015 (t7)
0.799* ±0.017 (t9)
0.799* ±0.013 (t9)
Relative spleen ratio (%)b
2.172 ±0.037
2.267 ± 0.042 (|4)
2.232 ±0.053 (|3)
2.333** ±0.041 (t7)
2.383** ±0.024 (tlO)
2.646** ± 0.042 (t22)
Absolute right testis (g)
(%)b
1.460 ±0.019
1.455 ±0.017(0)
1.448 ±0.030 (41)
1.463 ±0.014 (0)
1.457 ±0.026 (0)
1.381* ±0.014 (45)
Relative right testis ratio
(%)b
4.326 ±0.064
4.232 ±0.059 (42)
4.240 ±0.106 (42)
4.350 ±0.073 (tl)
4.350 ±0.030 (tl)
4.575* ±0.048 (t6)
Absolute left testis (g)b
1.5770 ±0.0200
Not Measured
Not Measured
1.5301 ±0.0167 (43)
1.5373 ±0.0220 (43)
1.4476 ±0.0141**
(|8)
62
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Table A-9. Absolute and Relative Organ Weights in Male and Female F1 Pups in the 13-Week Toxicity Study of SAN Trimer"
Endpoint
Dose Group ppm (mg/kg-day)
0
100 (10)
200 (20)
400 (40)
800 (80)
1,600 (150)
Absolute thymus (g) (%)b
0.316 ±0.009
0.337 ±0.014(17)
0.298 ±0.021 (46)
0.299 ±0.008 (45)
0.302 ±0.011 (44)
0.267 ±0.015 (416)
Relative thymus (%)b
0.938 ±0.034
0.980 ±0.038 (|4)
0.873 ± 0.060 (47)
0.888 ±0.025 (45)
0.900 ±0.030 (44)
0.882 ±0.041 (46)
Females
Number of animals
10
10
10
10
10
10
Necropsy body weight (g)
(%)b
203 ±4
202 ± 3 (0)
192* ±2 (45)
196* ±2 (43)
190** ± 1 (46)
184** ±2 (49)
Absolute brain (g) (%)b
1.823 ±0.013
1.813 ±0.017(|1)
1.798 ±0.018 (41)
1.845 ±0.015 (|1)
1.844 ±0.010 (|1)
1.841 ±0.007 (|1)
Relative brain ratio (%)b
9.022 ±0.179
9.000 ±0.136(0)
9.372* ±0.109 (|4)
9.410* ±0.070 (|4)
9.705** ±0.078 (|8)
10.016** ±0.109
(til)
Absolute heart (g) (%)b
0.64 ±0.01
0.64 ± 0.02 (0)
0.62 ±0.01 (43)
0.65 ±0.01 (|2)
0.64 ±0.01 (0)
0.63 ±0.01 (42)
Relative heart ratio (%)b
3.146 ±0.074
3.190 ±0.063 (|1)
3.238 ±0.045 (|3)
3.291 ±0.046 (|5)
3.363 ± 0.046** (|7)
3.412 ±0.046** (|8)
Absolute R. kidney (g) (%)b
0.68 ±0.01
0.67 ±0.01 (|1)
0.66 ±0.01 (43)
0.69 ±0.02 (|1)
0.67 ±0.01 (41)
0.66 ±0.01 (43)
Relative R. kidney ratio
(%)b
3.362 ±0.051
3.334 ±0.027 (|1)
3.417 ±0.031 (42)
3.492 ±0.073 (|4)
3.547 ±0.050** (|6)
3.609 ±0.019** (|7)
Absolute liver (g) (%)b
6.54 ±0.17
6.68 ±0.08 (|2)
6.40 ±0.12 (42)
6.74 ±0.09 (|3)
6.69 ±0.08 (|2)
6.74 ±0.09 (|3)
Relative liver ratio (%)b
32.250 ± 0.404
33.145 ±0.274 (|3)
33.359 ±0.554 (|3)
34.328** ±0.328 (|6)
35.213** ±0.395 (|9)
36.652** ±0.376
(t 14)
Absolute lung (g) (%)b
1.35 ±0.07
1.28 ±0.04 (|5)
1.26 ±0.03 (47)
1.24 ±0.04 (48)
1.28 ±0.06 (45)
1.20 ±0.05 (411)
Relative lung ratio (%)b
6.615 ±0.269
6.369 ±0.189 (41)
6.539 ±0.140 (41)
6.341 ±0.164 (44)
6.740 ±0.310 (|2)
6.542 ±0.291 (41)
63
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8-11-2014
Table A-9. Absolute and Relative Organ Weights in Male and Female F1 Pups in the 13-Week Toxicity Study of SAN Trimer"
Endpoint
Dose Group ppm (mg/kg-day)
0
100 (10)
200 (20)
400 (40)
800 (80)
1,600 (150)
Absolute spleen (g) (%)b
0.535 ±0.013
0.523 ±0.008 (|2)
0.528 ±0.021 (41)
0.533 ±0.012 (40)
0.550 ±0.010 (|3)
0.579* ±0.008 (|8)
Relative spleen ratio (%)b
2.639 ±0.032
2.596 ±0.038 (42)
2.748 ± 0.099 (|4)
2.712 ±0.042 (|3)
2.897** ±0.055 (|10)
3.147** ±0.051 (f 19)
Absolute thymus (g) (%)b
0.264 ± 0.007
0.249 ± 0.006 (46)
0.225** ±0.005 (415)
0.243 ± 0.008 (48)
0.242 ±0.012 (48)
0.244 ±0.011 (48)
Relative thymus (%)b
1.301 ±0.029
1.235 ±0.028 (45)
1.174 ±0.033 (410)
1.234 ±0.033 (45)
1.275 ±0.062 (42)
1.326 ±0.052 (|2)
Absolute uterus (g) (%)b
0.558 ±0.070
0.564 ±0.054 (|1)
0.513 ±0.066 (48)
0.655 ±0.067 (f 17)
0.463 ±0.065 (417)
0.643 ±0.101 (f 15)
Relative uterus ratio (%)b
2.797 ±0.395
2.795 ± 0.266 (0)
2.667 ±0.340 (45)
3.327 ±0.325 (|19)
2.432 ±0.335 (413)
3.497 ±0.548 (|25)
aNTP_£2012).
bMean ± standard error (% change from control).
*Williams' or Dunnett's testp < 0.05.
**Williams' or Dunnett's test p £ 0,01,
64
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Table A-10. Hematology, Clinical Chemistry, and Urinalysis Data for F1 Rats in the
13-Week Toxicity Study of SAN Trimera'b
Endpoint
Exposure Group (Adjusted Daily Dose, mg/kg-day)c
0
10
20
40
80
150
Males
Number of animals
10
10
10
10
10
10
Hematology
Hemoglobin (g/dL)
15.5 ±0.2
15.7 ±0.1
15.4 ±0.2
15.5 ±0.1
15.5 ±0.2
15.0 ±0.1*
Erythrocytes (10'7|iL)
9.25 ±0.08
9.44 ±0.08
9.32 ±0.09
9.27 ±0.10
9.31 ±0.10
8.88 ±
0.10*
Mean cell volume (fL)
54.3 ±0.2
54.3 ±0.1
54.0 ±0.2
54.4 ±0.2
54.3 ± 0/2
55.4 ±
0.3**
Clinical Chemistry
Urea nitrogen (mg/dL)
14.9 ±0.6
13.6 ±0.7
13.6 ±0.4
13.9 ±0.9
15.5 ±0.7
15.4 ±0.6
Creatinine (mg/dL)
0.55 ±0.02
0.53 ±0.02
0.51 ±0.01
0.58 ±0.02
0.57 ±0.02
0.60 ±0.01
Total protein (g/dL)
6.5 ±0.1
6.6 ±0.1
6.5 ±0.1
6.5 ±0.1
6.8 ± 0.1*
6.5 ±0.1
Albumin (g/dL)
4.5 ±0.1
4.5 ±0.1
4.5 ±0.0
4.5 ±0.0
4.7 ± 0.1**
4.6 ±0.0**
Cholesterol (mg/dL)
79 ±2
82 ±2
80 ±2
81 ±2
78 ± 1
69 ± 1**
Triglycerides (mg/dL)
255 ± 16
206 ± 15
210 ± 17
163 ±12**
199±16**
155 ±11**
Alanine aminotransferase (IU/L)
60 ±3
52 ±2
49 ± 2**
45 ± 1**
47 ± 2**
43 ± 1**
Aspartate aminotransferase (IU/L)
77 ±5
66 ±3*
63 ± 1**
62 ±2**
64 ± 4**
55 ±2**
Bile acids (|imol/L)
14.7 ±2.2
11.7 ± 1.1
11.2 ± 1.2
7.3 ±0.9**
11.0 ± 1.5
10.3 ± 1.4
Urinalysis
Glucose (mg/dL)
35 ±3
38 ±2
37 ±2
41 ±4
46 ±3*
37 ±2
Glucose/creatinine ratio
0.18 ±0.01
0.18 ±0.00
0.18 ±0.01
0.19 ±0.01
0.20 ±
0.01*
0.20 ±
0.01*
Protein (mg/dL)
164 ± 14
166 ±7
170 ± 13
170 ± 15
240±19**
227 ± 25*
Protein/creatinine ratio
0.88 ±0.71
0.79 ±0.03
0.84 ±0.04
0.81 ±0.05
1.05 ±0.06
1.20 ±
0.06**
Alkaline phosphatase/creatinine ratio
0.09 ±0.01
0.10±0.01
0.11±0.01
0.09 ±0.01
0.08 ±0.00
0.10 ±0.01
Aspartate aminotransferase (IU/L)
30 ± 14
18 ± 1
19 ±2
23 ±2
28 ±3**
35 ±6**
Aspartate aminotransferase/ creatinine
ratio
0.02 ±0.01
0.01 ±0.00
0.01 ±0.00
0.01 ±0.00
0.01 ±
0.00*
0.02 ±
0.00**
\-Acetyl-/;-D-
glucosaminidase/creatinine ratio
0.02 ±0.01
0.01 ±0.00
0.01 ±0.00
0.01 ±0.00
0.01 ±0.00
0.01 ±0.00
Females
Number of animals
10
10
10
10
10
10
Hematology
Hemoglobin (g/dL)
14.5 ±0.1
14.8 ±0.1
14.7 ±0.1
14.7 ±0.1
14.3 ±0.1
14.1 ±0.2
Erythrocytes (106/|iL)
8.23 ±0.07
8.43 ±0.08
8.39 ±0.06
8.38 ±0.07
8.16 ±0.06
8.02 ± 0.08
Mean cell volume (fL)
56.3 ±0.2
56.2 ±0.2
56.0 ±0.1
56.3 ±0.1
56.3 ±0.2
56.9 ±0.2
65
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8-11-2014
Table A-10. Hematology, Clinical Chemistry, and Urinalysis Data for F1 Rats in the
13-Week Toxicity Study of SAN Trimera'b
Endpoint
Exposure Group (Adjusted Daily Dose, mg/kg-day)c
0
10
20
40
80
150
Clinical Chemistry
Urea nitrogen (mg/dL)
15.5 ±0.6
15.8 ±0.4
17.6 ±0.6
15.7 ±0.5
16.6 ±0.5
18.2 ±
0.6**
Creatinine (mg/dL)
0.52 ±0.01
0.53 ±0.02
0.57 ±
0.02*
0.59 ±
0.01**
0.58 ±
0.01**
0.58 ±
0.01**
Total protein (g/dL)
6.4 ±0.1
6.7 ±0.1
6.6 ±0.1
6.7 ±0.1
6.6 ±0.1
6.6 ±0.1
Albumin (g/dL)
4.7 ±0.1
4.9 ±0.1
4.8 ±0.1
4.8 ±0.1
4.8 ±0.1
4.8 ±0.1
Cholesterol (mg/dL)
79 ±2
85 ± 1
83 ±2
83 ±2
77 ±2
74 ±2
Triglycerides (mg/dL)
85 ±9
87 ±8
72 ±7
72 ±7
65 ±9*
45 ±6**
Alanine aminotransferase (IU/L)
43 ±2
52 ±3
46 ±2
42 ±2
41 ± 2
39 ± 1
Aspartate aminotransferase (IU/L)
66 ± 1
67 ±3
66 ±2
62 ± 1
60 ± 1*
57 ±3**
Bile acids (|imol/L)
25.1 ± 1.8
15.4 ± 1.8*
17.0 ± 1.2
19.8 ±2.7
15.3 ±
3.1**
21.5 ±2.8
Urinalysis
Glucose (mg/dL)
26 ±3
29 ±3
26 ±3
27 ±3
24 ±4
29 ±3
Glucose/creatinine ratio
0.20 ±0.01
0.21 ±0.01
0.21 ±0.00
0.20 ±0.00
0.21 ±0.00
0.20 ± 0.00
Protein (mg/dL)
44 ±5
46 ±5
41 ±5
46 ±7
45 ± 10
73 ± 13
Protein/creatinine ratio
0.33 ±0.01
0.33 ±0.01
0.33 ±0.01
0.32 ±0.02
0.35 ±0.02
0.47 ± 0.04
Alkaline phosphatase/creatinine ratio
0.09 ±0.01
0.10 ±0.00
0.09 ±0.01
0.08 ±0.00
0.08 ±0.00
0.06 ±
0.00**
Aspartate aminotransferase (IU/L)
11 ± 2
10 ±2
9 ± 1
8 ± 2
8 ± 1
15 ±2
Aspartate aminotransferase/ creatinine
ratio
0.01 ±0.00
0.01 ±0.00
0.01 ±0.00
0.01 ±0.00
0.02 ± 0.00
0.01 ±0.00
\-Acetyl-/;-D-
glucosaminidase/creatinine ratio
0.01 ±0.00
0.01 ±0.00
0.01 ±0.00
0.01 ±0.00
0.01 ±
0.00**
0.01 ±
0.00**
aNTP (20121.
bData are presented as mean ± standard error. Statistical tests were performed on unrounded data.
Doses calculated by study authors.
* Significantly different (p < 0.05) from the control group by Dunn's or Shirley's test
**p < 0.01
66
SAN Trimer
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FINAL
8-11-2014
Table A-ll. Hematology, Clinical, and Urine Parameters in the F344/N Rats Treated with
SAN Trimer for up to 78 Weeksa'b
Parameter
Exposure Group, mg/kg-day
0
20
40
75 (males)/
85 (females)
Hematology
Male Rat—27 Weeks
Hemoglobin (g/dL)
16.2 ±0.1
15.6 ±0.2
15.8 ±0.2
15.5 ± 0.1*
Reticulocytes (107|iL
194.5 ±9.9
199.4 ±7.5
197.4 ±6.0
237.8 ± 10.2*
Mean cell hemoglobin (pg)
16.9 ±0.1
16.6 ±0.1*
16.7 ±0.1
16.8 ±0.1
Mean cell hemoglobin concentration
(g/dL)
31.6 ±0.2
31.2 ±0.2
31.2 ± 0.1
31.0 ± 0.1**
Segmented neutrophils (1057|iL)
1.51±0.08
1.51 ±0.08
1.34 ±0.06
1.28 ±0.04*
Male Rat—52 Weeks
Hemoglobin (g/dL)
16.1 ±0.2
15.8 ±0.2
15.7 ±0.1
15.6 ±0.2
Reticulocytes (107|iL
194.1 ±4.9
209.0 ±9.5
215.7 ±8.8
221.5± 9.7
Mean cell hemoglobin (pg)
17.2 ±0.1
17.0± 0.1
17.0 ±0.1
17.0 ±0.1
Mean cell hemoglobin concentration
(g/dL)
33.0 ±0.2
32.5 ±0.1
32.5 ±0.1
32.6± 0.2
Segmented neutrophils (105V|iL)
1.76±0.17
1.56± 0.13
1.64± 0.08
1.74 ±0.16
Male Rat—78 Weeks
Hemoglobin (g/dL)
15.7 ±0.2
15.9 ±0.2
14.7 ± 1.2
15.6 ±0.2
Reticulocytes (10:7|iL
224.7 ± 13.9
241.0 ± 15.2
314.6 ±83.3
222.5± 7.9
Mean cell hemoglobin (pg)
17.2 ±0.1
17.3± 0.2
16.7 ±0.3
17.1 ± 0.1
Mean cell hemoglobin concentration
(g/dL)
32.9 ±0.1
33.1 ± 0.3
32.6 ±0.3
32.7± 0.1
Segmented neutrophils (10:7|iL)
1.51±0.13
1.29± 0.11
1.57± 0.18
1.64 ±0.27
Urinalysis/Clinical Chemistry
Male Rat—26 Weeks (Urinalysis)/ 27 Weeks (Clinical Chemistry)
Creatinine (mg/dL)
0.63 ±0.02
0.64 ±0.02
0.65 ±0.02
0.69 ±0.01**
Glucose (mg/dL)
128 ±3
133 ±2
125 ±2
145 ±9*
Sodium (mEq/L)
148 ±0
149 ±0
149 ±0
151 ±0**
Chloride (mEq/L)
97 ±0
98 ±0
97 ±0
100 ±0**
Albumin (g/dL)
4.6 ±0.0
4.8 ±0.0*
4.9 ± 0.1**
4.9 ± 0.1**
Triglycerides (mg/dL)
315 ± 27
249 ± 13
220 ±14**
200 ±12**
Alanine aminotransferase (IU/L)
144 ±8
80 ±3**
73 ± 2**
65 ±2**
Alkaline phosphatase (IU/L)
169 ±4
154 ±3*
156 ±4*
158 ±4
Aspartate aminotransferase (IU/L)
122 ±8
69 ±3**
69 ±3**
64 ± 3**
Lactate dehydrogenase (IU/L)
187 ± 19
133 ±32
235 ±30
111±11*
Sorbitol dehydrogenase (IU/L)
29 ± 1
17 ± 1**
19 ± 1**
17 ± 1**
67
SAN Trimer
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FINAL
8-11-2014
Table A-ll. Hematology, Clinical, and Urine Parameters in the F344/N Rats Treated with
SAN Trimer for up to 78 Weeksa'b
Parameter
Exposure Group, mg/kg-day
0
20
40
75 (males)/
85 (females)
Male Rat—52 Weeks
Creatinine (mg/dL)
195.2 ±8.3
179.1 ±8.1
168.7 ± 10.2
194.4 ±9.9
Glucose (mg/dL)
39 ±2
36 ±2
35 ±3
39 ±2
Sodium (mEq/L)
149 ±0
150 ±0
149 ±0
149 ±0
Chloride (mEq/L)
98 ±0
98 ±0
98 ±0
99 ±0
Albumin (g/dL)
4.7 ±0.0
4.8 ±0.1
4.9 ±0.0**
4.9 ± 0.1*
Triglycerides (mg/dL)
251 ± 15
255 ± 17
225 ± 13
224 ± 12
Alanine aminotransferase (IU/L)
123 ±4
100 ±8**
83 ±4**
75 ±3**
Alkaline phosphatase (IU/L)
152 ±5
153 ±4
147 ±3
152 ±4
Aspartate aminotransferase (IU/L)
122 ±8
89 ± 4**
73 ±4**
69 ±2**
Lactate dehydrogenase (IU/L)
220 ± 35
203 ± 20
133 ±25*
150 ±52**
Sorbitol dehydrogenase (IU/L)
31 ± 1
27 ±2*
22 ± 2**
19 ± 1**
Male Rat—78 Weeks
Creatinine (mg/dL)
0.58 ±0.01
0.63 ±0.03
0.61 ±0.01
0.64 ±0.02*
Glucose (mg/dL)
141 ±4
141 ±3
133 ±3
131 ±3
Sodium (mEq/L)
151 ±0
151 ±0
151 ± 1
151 ±0
Chloride (mEq/L)
101 ±0
101 ± 1
101 ± 1
101 ±0
Albumin (g/dL)
4.4 ±0.0
4.3 ±0.1
4.3 ±0.1
4.4 ±0.1
Triglycerides (mg/dL)
187 ±9
194 ± 16
182 ±8
155 ± 12
Alanine aminotransferase (IU/L)
151 ± 12
111 ±9*
77 ±7*
73 ±5**
Alkaline phosphatase (IU/L)
151 ±3
144 ± 10
144 ±6
143 ±6
Aspartate aminotransferase (IU/L)
143 ± 12
158 ±45
89 ± 4**
88 ±4**
Lactate dehydrogenase (IU/L)
210 ±23
404 ±186
154 ± 13
177 ± 18
Sorbitol dehydrogenase (IU/L)
37 ±2
40 ± 11*
23 ± 2**
26 ± 1**
Parameter
Exposure Group, mg/kg-day
0
20
40
85
Hematology
Female Rat—27 Weeks
Hematocrit
47.1 ±0.6
47.2 ±0.5
46.7 ±0.7
44.7 ±0.7*
Hemoglobin (g/dL)
15.4 ±0.2
15.3 ±0.1
15.1 ±0.2
14.5 ±0.2**
Erythrocytes b (xl06/MM3)
8.52 ±0.10
8.35 ±0.08
8.31 ± 0.11
7.99 ± 0.11**
Mean cell hemoglobin (g/dl)
32.1 ± 0.1
32.3 ±0.1
31.9 ± 0.2
31.9 ± 0.2
Platelets (107|il)
615.9± 15.4
627.6 ±31.1
579.9 ±23.6
632.8 ± 19.3
68
SAN Trimer
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FINAL
8-11-2014
Table A-ll. Hematology, Clinical, and Urine Parameters in the F344/N Rats Treated with
SAN Trimer for up to 78 Weeksa'b
Parameter
Exposure Group, mg/kg-day
0
20
40
75 (males)/
85 (females)
Female Rat—52 Weeks
Hematocrit
47.9 ±0.5
46.9 ±0.6
47.0 ±0.5
45.3 ±0.4**
Hemoglobin (g/dL)
16.0 ±0.2
15.7 ±0.2
15.6 ±0.2
14.9 ±0.2**
Erythrocytes b (xl06/MM3)
8.69 ±0.09
8.53 ±0.09
8.54 ±0.08
8.16 ±0.10**
Mean cell hemoglobin (g/dl)
33.4 ±0.2
33.4 ±0.2
33.1 ± 0.1
32.9 ±0.1*
Platelets (103/jxl)
625.9 ± 12.8
624.1 ± 18.2
648.8 ± 13.1
697.4 ±3.9**
Female Rat—78 Weeks
Hematocrit
49.0 ±0.8
49.5 ±0.7
47.0 ± 1.1
47.7 ±0.7
Hemoglobin (g/dL)
15.8 ±0.2
15.7 ±0.2
15.0 ±0.4
15.2 ±0.2
Erythrocytes b (xl06/MM3)
8.45 ±0.11
8.38 ± 0.11
8.07 ±0.27
8.24 ±0.15
Mean cell hemoglobin (g/dl)
33.3 ±0.2
33.3 ±0.1
33.2 ±0.4
32.9 ±0.2
Platelets (103/jxl)
518.2 ± 15.7
507.3 ±21.2
583.7 ±47.7
514.1 ±28.0
Urinalysis/Clinical Chemistry
Female Rat—26 Weeks (Urinalysis)/ 27 Weeks (Clinical Chemistry)
Urea nitrogen (mg/dL)
16.1 ±0.6
16.0 ±0.5
15.2 ±0.3
16.6 ±0.4
Creatinine (mg/dL)
0.67 ±0.02
0.68 ±0.01
0.70 ±0.00
0.72 ±0.02
Potassium (mEq/L)
5.2 ±0.1
5.1 ±0.2
5.1 ± 0.1
5.1 ± 0.1
Chloride (mEq/L)
100 ± 1
98 ± 1
99 ± 1
101 ± 1
Calcium (mg/dL)
11.6 ± 0.1
11.5 ± 0.1
11.7 ± 0.1
11.1 ± 0.1**
Phosphorus (mg/dL)
7.2 ±0.2
6.8 ±0.2
6.7 ±0.3
5.9 ±0.3**
Total protein (g/dL)
6.8 ±0.1
7.1 ± 0.1
7.1 ± 0.1
6.8 ±0.1
Albumin (g/dL)
4.8 ±0.1
5.0 ±0.1
5.0 ±0.1
4.8 ±0.0
Cholesterol (mg/dL)
115 ± 3
109 ±3
111 ±2
97 ± 2**
Triglycerides (mg/dL)
165 ± 17
121 ±9
128 ±11
72 ± 4**
Alanine aminotransferase (IU/L)
90 ±8
55 ±2**
56 ±4**
54 ±2**
Aspartate aminotransferase (IU/L)
91 ±8
69 ±3*
68 ±4**
68 ±2*
Sorbitol dehydrogenase (IU/L)
21 ± 1
17 ± 1*
18 ± 1*
15 ± 1**
Glucose/creatinine ratio
0.20 ±0.01
0.19 ±0.01
0.19 ±0.00
0.19 ±0.01
Protein (mg/dL)
36 ±3
41 ±3
51 ±4**
53 ±6*
Protein/creatinine ratio
0.28 ±0.02
0.31 ±0.01
0.35 ±0.01**
0.40 ±0.02**
Aspartate aminotransferase (IU/L)(Urine)
8 ± 1
8 ± 2
9 ± 1
11 ±2
Female Rats—52 Weeks
Urea nitrogen (mg/dL)
14.7 ±0.4
15.1 ± 0.3
15.5 ±0.5
16.5 ±0.4**
Creatinine (mg/dL)
0.62 ±0.01
0.69 ±0.01**
0.68 ±0.01**
0.73 ±0.02**
Potassium (mEq/L)
5.5 ±0.1
5.5 ±0.1
5.6 ±0.1
5.6 ±0.1
69
SAN Trimer
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FINAL
8-11-2014
Table A-ll. Hematology, Clinical, and Urine Parameters in the F344/N Rats Treated with
SAN Trimer for up to 78 Weeksa'b
Parameter
Exposure Group, mg/kg-day
0
20
40
75 (males)/
85 (females)
Chloride (mEq/L)
99 ±0
99 ±0
98 ±0
99 ±0
Calcium (mg/dL)
12.3 ±0.1
12.0 ±0.1
12.2 ±0.1
11.9 ± 0.1*
Phosphorus (mg/dL)
6.0 ±0.2
5.1 ±0.2*
5.3 ±0.2*
5.1 ±0.2**
Total protein (g/dL)
8.0 ±0.1
8.0 ±0.1
8.1 ± 0.1
7.8 ±0.1
Albumin (g/dL)
5.6 ±0.1
5.6 ±0.1
5.7 ±0.1
5.5 ±0.1
Cholesterol (mg/dL)
139 ±3
139 ±2
137 ±2
128 ±3*
Triglycerides (mg/dL)
212 ±11
208 ± 17
212 ±30
178 ± 18
Alanine aminotransferase (IU/L)
69 ±3
56 ±2**
54 ±2**
48 ± 1**
Aspartate aminotransferase (IU/L)
70 ±6
56 ±2
58 ±3
55 ±2*
Sorbitol dehydrogenase (IU/L)
18 ± 1
16 ± 1
15 ± 1*
15 ± 1**
Glucose/creatinine ratio
0.21 ±0.01
0.23 ±0.01
0.20 ±0.01
0.20 ±0.01
Protein (mg/dL)
66 ±5
80 ±20
71 ±6
79 ±9
Protein/creatinine ratio
0.42 ±0.01
0.60 ±0.12
0.49 ±0.04
0.55 ±0.04**
Female Rats—78 Weeks
Urea nitrogen (mg/dL)
14.0 ±0.6
14.6 ±0.5
14.2 ±0.3
14.9 ±0.5
Creatinine (mg/dL)
0.58 ±0.01
0.62 ±0.01
0.62 ±0.01
0.67 ±0.02**
Potassium (mEq/L)
5.4 ±0.1
4.9 ±0.2*
5.2 ±0.1
4.8 ± 0.1*
Chloride (mEq/L)
100 ±0
100 ±0
100 ±0
99 ±0*
Calcium (mg/dL)
12.0 ±0.1
12.2 ±0.1
12.0 ±0.1
12.2 ±0.1
Phosphorus (mg/dL)
6.4 ±0.2
6.0 ±0.1
5.7 ±0.3
6.0 ±0.3
Total protein (g/dL)
7.0 ±0.1
7.3 ± 0.1*
7.5 ±0.2**
7.4 ±0.1**
Albumin (g/dL)
4.8 ±0.0
5.0 ± 0.1*
5.2 ± 0.1**
5.1 ± 0.1**
Cholesterol (mg/dL)
123 ±4
134 ±4
127 ±4
125 ±5
Triglycerides (mg/dL)
162 ± 15
177 ± 13
178 ±21
138 ± 15
Alanine aminotransferase (IU/L)
71 ±8
62 ±9
49 ±3*
60 ±7
Aspartate aminotransferase (IU/L)
92 ± 10
79 ± 11
67 ±5*
73 ±5
Sorbitol dehydrogenase (IU/L)
23 ±2
20 ±2
18 ± 1*
22 ±2
Glucose/creatinine ratio
0.24 ±0.00
0.24 ±0.01
0.23 ±0.01
0.22 ±0.01**
Protein (mg/dL)
83 ± 12
75 ± 14
93 ± 12
86 ±8
Protein/creatinine ratio
0.58 ± 0.11
0.62 ±0.11
0.66 ±0.06
0.65 ±0.06
aNTP (20121.
bData are presented as mean ± standard error. Statistical tests were performed on unrounded data.
* Significantly different (p < 0.05) from the control group by Dunn's or Shirley's test
**p < 0.01
70
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FINAL
8-11-2014
Table A-12. Incidence of Histopathological Endpoints in F344/N Rats Exposed to SAN
Trimer by Diet for 2 Years3
Parameterb
Exposure Group
Males
Females
0
20
40
75
0
20
40
85
Brain and Spinal Cord
Spinal root degeneration
(severity score)
34/47
(1.0)
37/48
(1.1)
37/50
(1.2)
43*/50
(1.3)
43/49
(1.2)
40/50
(1.2)
42/50
(1.3)
45/49
(1.5)
Sciatic nerve
degeneration
(severity score)
37/50
(1.1)
40/50
(1.2)
41/50
(1.3)
43/50
(1.3)
28/49
(1.0)
35/49
(1.1)
43**/49
(1.1)
40*/50
(1.1)
Liver
Angiectasis
(severity score)
1/50
(2.0)
5/50
(1.0)
1/50
(1.0)
9*/50
(1.1)
5/50
(NR)
5/50
(NR)
3/50
(NR)
5/50
(NR)
Eosinophilic foci
(severity score)
17/50
(NR)
19/50
(NR)
22/50
(NR)
33*/50
(NR)
23/50
(NR)
31/50
(NR)
30/50
(NR)
29/50
(NR)
Active chronic
inflammation
(severity score)
34/50
(1.0)
40/50
(1.1)
38/50
(1.1)
43*/50
(1.0)
41/50
(NR)
45/50
(NR)
43/50
(NR)
47/50
(NR)
Mixed cell foci
(severity score)
6/50
(NR)
19**/50
(NR)
12/50
(NR)
20**/50
(NR)
4/50
(NR)
8/50
(NR)
7/50
(NR)
13*/50
(NR)
Bone Marrow
Bone marrow
hyperplasia
(severity score)
24/50
(1.9)
24/50
(1.8)
24/50
(1.8)
37**/50
(1.6)
16/50
(1.8)
25/50
(1.8)
25*/50
(2.0)
38**/50
(1.5)
Inflammation,
granulomatous
(severity score)
0/50
(NR)
0/50
(NR)
0/50
(NR)
3/50
(2.0)
0/50
(NR)
0/50
(NR)
6*/50
(1.2)
2/50
(1.5)
Bladder
Hyperplasia of
transitional epithelium
(severity score)
0
(NR)
1
(NR)
0
(NR)
0
(NR)
1
(1.0)
0
(NR)
0
(NR)
12**
(2.3)
aNTP (20121.
bIncidence, reported by the study authors.
Severity Score, average severity grade of lesions in affected animals: 1 = minimal, 2 = mild, 3 = moderate,
4 = marked.
* Significantly different from control (p < 0.05) using the Poly-3 test, as reported by the study authors.
**Significantly different from control (p < 0.01) using the Poly-3 test, as reported by the study authors.
NR = Not reported.
71
SAN Trimer
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FINAL
8-11-2014
Table A-13. Tumor Incidence in F344/N Rats Administered SAN Trimer by Diet for
2 Years"
Parameter
Exposure Group, mg/kg-d (Human Equivalency Dose, mg/kg-day)b
Males
Females
0
(0)
20
(5.6)
40
(11)
75
(20)
0
(0)
20
(5)
40
(9.8)
85
(20)
Sample size
50
50
50
50
50
50
50
50
Brain and Spinal Cord
Astrocytomas0
0
0
1(2)
2(4)
NR
NR
NR
NR
Granular cell tumors0
0
1(2)
1(2)
1(2)
0
1(2)
1(2)
0
Glioma, Mixed Cell
NR
NR
NR
NR
1(2)
1(2)
1(2)
0
Granular cell tumor or meningioma
NR
NR
NR
NR
0
1(2)
1(2)
0
Pituitary gland
Adenoma00'
16 (32)
10 (20)
13 (26)
4 (8)d
22 (44)
12 (24)d
19 (38)
9 (18)d
Adenoma or carcinoma0 '
NR
NR
NR
NR
22 (44)
12 (24)d
20 (40)
9 (18)d
Testes
Bilateral, interstitial cell, adenoma0 8
33 (66)
45 (90)h
39 (78)
49 (98)h
NA
NA
NA
NA
Unilateral, interstitial cell, adenoma0
8(16)
4(8)
5(10)
0(0)
NA
NA
NA
NA
Overall, interstitial cell, adenoma
(bilateral + unilateral, combined)0 0
41 (82)
49 (98)d
44 (88)
49 (98)d
NA
NA
NA
NA
Mammary gland
Fibroadenoma0^
1(2)
2(4)
1(20)
3(6)
36 (72)
31 (62)
26 (52)d
20 (40)d
Fibroadenoma or adenomao f
NR
NR
NR
NR
36 (72)
31 (62)
27 (54)d
20 (40)d
Fibroadenoma, adenoma, or
carcinomaof
NR
NR
NR
NR
36 (72)
31 (62)
27 (54)d
21 (42)d
All organs
Mononuclear cell leukemiaoof
15 (30)
7 (14)d
5 (10)d
3 (6)d
13 (26)
2 (4)d
3 (6)d
2 (4)d
Benign neoplasms0
46 (92)
49 (98)
47 (94)
49 (98)
42 (84)
37 (74)
39 (78)
34 (68)d
Malignant neoplasms0 0
22 (44)
17 (34)
11 (22)d
8 (16)d
19 (38)
5 (10)d
6 (12)d
12 (24)
Benign or malignant neoplasmso f
49 (98)
50 (100)
49 (98)
49 (98)
44 (88)
41 (82)
41 (82)
35 (70)d
"NTP (2012).
bDoses were converted from adjusted daily doses to human equivalency doses using the following formula:
DoseHED = DoseADi x (body weight animal ^ body weight human)(0 25).
Incidence, (corresponding percentage).
Significantly different from control (p < 0.05) using the Poly-3 test, as reported by the study authors.
"Statistically significant trend using Poly-3 test for dose-response relationship in male rats, as reported by the
study authors.
Statistically significant trend using Poly-3 test for dose-response relationship in female rats, as reported by the
study authors.
Statistically significant trend using Cochran-Armitage test for dose-response relationship in male rats, as
performed for this PPRTV assessment.
Significantly different from control (p < 0.05) using the Fisher's Exact test, as performed for this PPRTV
assessment.
NA = Not applicable, NR = Not reported.
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APPENDIX B. BENCHMARK DOSE CALCULATIONS FOR THE
SUBCHRONIC p-RfD AND CHRONIC p-RfD
MODELING PROCEDURE FOR DICHOTOMOUS DATA
The BMD modeling of dichotomous data was conducted with the U.S. EPA's BMDS
(version 2.2.2). For these data, all of the dichotomous models (i.e., Gamma, Multistage,
Logistic, Log-logistic, Probit, Log-probit, and Weibull models) available within the software
were fit using a default BMR of 10% extra risk based on the U.S. EPA's Benchmark Dose
Technical Guidance Document (U.S. EPA. 2012). Adequacy of model fit was judged based on
the x2 goodness-of-fit p-walue (p> 0.1), magnitude of scaled residuals in the vicinity of the
BMR, and visual inspection of the model fit. Among all models providing adequate fit, the
lowest BMDL was selected if the BMDLs estimated from different models varied greater than
3-fold; otherwise, the BMDL from the model with the lowest AIC was selected as a potential
POD from which to derive a p-RfD.
In addition, data from exposures much higher than the study LOAEL do not provide
reliable information regarding the shape of the response curve at low doses. However, such
exposures can have a strong effect on the shape of the fitted model in the low-dose region of the
dose-response curve in some cases. Thus, if lack of fit is due to characteristics associated with
dose-response data for high doses, then the U.S. EPA's Benchmark Dose Technical Guidance
Document allows for data to be adjusted by eliminating high-dose groups (U.S. EPA. 2012).
MODELING PROCEDURE FOR CONTINUOUS DATA
The BMD modeling of continuous data from the NTP (2012) study was conducted with
the U.S. EPA's BMDS (version 2.2.2). For these data, all continuous models available within
the software were fit using a default BMR of 1 SD relative risk. For changes in liver, body, and
kidney weights, a BMR of 10% change relative to the control mean was also used. An adequate
fit was judged based on the goodness-of-fit p-w alue (p> 0.1), magnitude of the scaled residuals
in the vicinity of the BMR, and visual inspection of the model fit. In addition to these three
criteria forjudging adequacy of model fit, a determination was made as to whether the variance
across dose groups was constant. If a constant variance model was deemed appropriate based on
the statistical test provided in BMDS (i.e., Test 2), the final BMD results were estimated from a
constant variance model. If the test for homogeneity of variance was rejected (p< 0.1), the
model was run again while modeling the variance as a power function of the mean to account for
this nonconstant variance. If this nonconstant variance model did not adequately fit the variance
data (i.e., Test 3;/?<0. 1), the data set was considered unsuitable for BMD modeling. Among all
models providing adequate fit, the lowest BMDL was selected if the BMDLs estimated from
different models varied greater than 3-fold; otherwise, the BMDL from the model with the
lowest AIC was selected as a potential POD from which to derive a p-RfD.
INCREASED ABSOLUTE LIVER WEIGHT IN MALE F344/N RATS TREATED WITH
SAN TRIMER FOR 13 WEEKS (NTP. 2012)
All available continuous models in BMDS (version 2.2.2) were fit to the increased
absolute liver weight data from male F344/N rats exposed to SAN Trimer for 13 weeks (NTP.
2012) (see Table A-9). For increased absolute liver weight, a BMR of a 10% change relative to
the control mean was used. In addition, a BMR of 1 SD relative risk was also estimated for
comparison purposes based on the U.S. EPA's BMD guidance (U.S. EPA. 2012). For increased
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absolute liver weight in male F1 rats, data were modeled without the highest dose of
150 mg/kg-day because there was a statistically significant decrease in body weight at that dose
that confounds the interpretation of organ weight changes. Therefore, only the BMD modeling
results based on data without the highest dose group are summarized in Table B-l and
Figure B-l. As assessed by the %2 goodness-of-fit statistic, AIC score, and visual inspection,
after excluding the highest dose group, the Hill model provided the best model fit based on the
lowest AIC and BMDL (see Table B-l and Figure B-l). Test 2 indicated that using a constant
variance model was appropriate for modeling these data. Estimated doses associated with a
10% BMR and the 95% lower confidence limit on these doses (BMDio and BMDLio values,
respectively) were 25 and 10 mg/kg-day.
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Table B-l. Model Predictions for Absolute Liver Weight in Male Rats3
Modelb
BMDio
BMDLio
BMDisd
BMDLisd
/7-Value
Test 2b
/7-Value
Test 3b
Goodness-of-
Fit
/>-Valucb
AIC
Conclusion
Exponential (M2)
59
45
39
30
0.215
0.215
0.174
29.38
Exponential (M3)
59
45
39
30
0.215
0.215
0.174
29.38
Exponential (M4)
27
12
15
6.8
0.215
0.215
0.594
27.45
Exponential (M5)
27
12
15
6.8
0.215
0.215
0.594
27.45
Hill
25
10
13
5.0
0.215
0.215
0.721
27.06
Lowest AIC
Power
57
43
37
28
0.215
0.215
0.200
29.05
Polynomial
57
43
37
28
0.215
0.215
0.200
29.05
Linear
57
43
37
28
0.215
0.215
0.200
29.05
aNTP (20121 13-week study.
bValues <0.10 fail to meet conventional goodness-of-fit criteria.
AIC = Akaike's Information Criteria; BMD = Benchmark dose; BMDL = Lower confidence limit (95%) on the benchmark dose.
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Hill Model, with BMR of 0.1 Rel. Dev. for the BMD and 0.95 Lower Confidence Limit for the BMDL
Hill
14
13.5
13
12.5
12
1.5
11
BMDL
BMD
10.5
0
10
20
30
40
50
60
70
80
dose
07:49 04/24 2014
Figure B-l. Hill BMD Model for Increased Absolute Liver Weight Data in Male Rats at
13 Weeks
(NTP. 2012)
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Text Output for Hill BMD Model for Increased Absolute Liver Weight in Male Rats
Treated with SAN Trimer for 13 weeks (NTP, 2012)
Hill Model. (Version: 2.17; Date: 01/28/2013)
Input Data File: C:/Users/JKaiser/Desktop/modeling
results/hil_aliver_sant_mfl_nhd_Hil-ConstantVariance-BMRlO-Restrict.(d)
Gnuplot Plotting File: C:/Users/JKaiser/Desktop/modeling
results/hil_aliver_sant_mfl_nhd_Hil-ConstantVariance-BMRlO-Restrict.pit
Wed Apr 30 14:10:33 2014
BMDS Model Run
The form of the response function is:
Y[dose] = intercept + v*dose^n/(k^n + dose^n)
Dependent variable = mean
Independent variable = dose
rho is set to 0
Power parameter restricted to be greater than 1
A constant variance model is fit
Total number of dose groups = 5
Total number of records with missing values = 0
Maximum number of iterations = 5 00
Relative Function Convergence has been set to: le-008
Parameter Convergence has been set to: le-008
Default Initial Parameter Values
alpha = 0.59066
rho = 0 Specified
intercept = 11.25
v = 1.88
n = 0.667711
k = 23.4286
Asymptotic Correlation Matrix of Parameter Estimates
( *** The model parameter(s) -rho -n
have been estimated at a boundary point, or have been specified by
and do not appear in the correlation matrix )
alpha intercept v k
alpha 1 -2.6e-007 -1.9e-007 -3.8e-007
intercept -2.6e-007 1 0.033 0.57
v -1.9e-007 0.033 1 0.79
k -3.8e-007 0.57 0.79 1
the user,
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Interval
Variable
Limit
alpha
0.749729
intercept
11.7356
v
3.74462
n
k
75.7046
Parameter Estimates
Estimate Std. Err.
0.538601 0.10772
11.2787 0.233129
2.42814 0.671689
1 NA
29.2305 23.7117
NA - Indicates that this parameter has hit a bound
implied by some inequality constraint and thus
has no standard error.
Table of Data and Estimated Values of Interest
95.0% Wald Confidence
Lower Conf. Limit Upper Conf.
0.327473
10.8217
1.11165
-17.2436
Dose
Obs Mean
Est Mean Obs Std Dev Est Std Dev Scaled Res.
0
10
20
40
80
10
10
10
10
10
11.3
12
12.3
12.5
13.1
11.3
11.9
12 .3
12 .7
13.1
0.66
0.54
0.66
0.79
1.08
0.734
0.734
0.734
0.734
0.734
-0.124
0.269
0.193
-0.653
0.314
Model Descriptions for likelihoods calculated
Model A1: Yij = Mu(i) + e(ij)
Var{e(ij)} = Sigma^2
Model A2: Yij = Mu(i) + e(ij)
Var{e(ij)} = Sigma(i)^2
Model A3: Yij = Mu(i) + e(ij)
Var{e(ij)} = Sigma^2
Model A3 uses any fixed variance parameters that
were specified by the user
Model R: Yi = Mu + e(i)
Var{e(i)} = Sigma^2
Likelihoods of Interest
Model
A1
A2
A3
fitted
R
Log(likelihood)
-9.203119
-6.306204
-9.203119
-9.530512
-22.901815
# Param's
6
10
6
4
2
AIC
30.406238
32.612408
30.406238
27.061024
49.803629
Explanation of Tests
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Test 1: Do responses and/or variances differ among Dose levels?
(A2 vs. R)
Test 2: Are Variances Homogeneous? (A1 vs A2)
Test 3: Are variances adeguately modeled? (A2 vs. A3)
Test 4: Does the Model for the Mean Fit? (A3 vs. fitted)
(Note: When rho=0 the results of Test 3 and Test 2 will be the same.)
Tests of Interest
Test -2*log(Likelihood Ratio) Test df p-value
Test 1 33.1912 8 <.0001
Test 2 5.79383 4 0.2151
Test 3 5.79383 4 0.2151
Test 4 0.654786 2 0.7208
The p-value for Test 1 is less than .05. There appears to be a
difference between response and/or variances among the dose levels
It seems appropriate to model the data
The p-value for Test 2 is greater than .1. A homogeneous variance
model appears to be appropriate here
The p-value for Test 3 is greater than .1. The modeled variance appears
to be appropriate here
The p-value for Test 4 is greater than .1. The model chosen seems
to adeguately describe the data
Benchmark Dose Computation
Specified effect = 0.1
Risk Type = Relative deviation
Confidence level = 0.95
BMD = 25.3548
BMDL = 10.0451
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INCREASED INCIDENCE OF CHRONIC ACTIVE HEPATIC INFLAMMATION IN
MALE F334/N RATS TREATED WITH SAN TRIMER FOR 2 YEARS (NTP. 2012)
All available dichotomous models in BMDS (version 2.2.2) were fit to the chronic active
hepatic inflammation data from male F344/N rats treated with SAN Trimer for 2 years (NTP.
2012) (see Table A-12). A BMR of 10% extra risk was used in all model runs. As assessed by
the x2 goodness-of-fit statistic, AIC score, and visual inspection, the Log-Logistic model
provided the best model fit (see Table B-2 and Figure B-2). Estimated doses associated with
10% extra risk and the 95% lower confidence limit on these doses (BMDio and BMDLio values,
respectively) were 8.0 and 3.2 mg/kg-day.
Table B-2. Model Predictions for Chronic Active Hepatic Inflammation in Males3
Model
BMDio
BMDLio
/; /J-Valuc
AIC
Conclusion
Gamma
11
5.7
0.575
213.45
Logistic
12
6.8
0.571
213.47
Log-Logisticb
8.0
3.2
0.579
213.42
Lowest AIC
LogProbit
21
11
0.480
213.84
Multistage
11
5.7
0.575
213.45
Probit
12
7.3
0.570
213.48
Weibull
11
5.7
0.575
213.45
"NTP (20121 2-year study.
bThe Log-Logistic model was selected as a best-fitting model based on lowest AIC.
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Log-Logistic Model with 0.95 Confidence Level
Log-Logistic
BMD Lower Bound
0.95
0.9
0.85
0.8
0.75
0.7
0.65
0.6
0.55
BMDL
BMD
0.5
0
10
20
30
40
50
60
70
dose
11:20 07/30 2012
Figure B-2. Log-Logistic Model Fit for Chronic Active Hepatic Inflammation in Males
(NTP. 2012)
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Text Output for Log-Logistic BMD Model for Increased Incidence of Chronic Active
Hepatic Inflammation in Male Rats Treated with SAN Trimer for 2 Years (NTP, 2012)
Logistic Model. (Version: 2.13; Date: 10/28/2009)
Input Data File:
C:/USEPA/BMDS22 0/BMDS22 0/Data/SessionFiles/lnl_santa_cai_m_Lnl-BMR10-Restrict.(d)
Gnuplot Plotting File:
C:/USEPA/BMDS22 0/BMDS22 0/Data/SessionFiles/lnl_santa_cai_m_Lnl-BMR10-Restrict.pit
Mon Jul 30 11:20:23 2012
BMDS Model Run
The form of the probability function is:
P[response] = background+(1-background)/[1+EXP(-intercept-siope*Log(dose))]
Dependent variable = Response
Independent variable = Dose
Slope parameter is restricted as slope >= 1
Total number of observations = 4
Total number of records with missing values = 0
Maximum number of iterations = 250
Relative Function Convergence has been set to: le-008
Parameter Convergence has been set to: le-008
User has chosen the log transformed model
Default Initial Parameter Values
background = 0.68
intercept = -4.10081
slope = 1
Asymptotic Correlation Matrix of Parameter Estimates
( *** The model parameter(s) -slope
have been estimated at a boundary point, or have been specified by
the user,
and do not appear in the correlation matrix )
background intercept
background 1 -0.75
intercept -0.75 1
Parameter Estimates
95.0% Wald Confidence
Interval
Variable Estimate Std. Err. Lower Conf. Limit Upper Conf.
Limit
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background
intercept
slope
0.690985
-4.27239
1
Indicates that this value is not calculated.
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-11-2014
Model
Full model
Fitted model
Reduced model
AIC:
Analysis of Deviance Table
Log(likelihood)
-104.166
-104.71
-106.633
213.421
# Param's Deviance Test d.f.
4
2 1.08917 2
1 4.93401 3
P-value
0.5801
0.1767
Dose
Goodness of Fit
Est. Prob.
Expected
Observed
Size
Scaled
Residual
0.0000
20.0000
40.0000
75.0000
Chi^2 = 1.09
0.6910
0.7584
0.8017
0.8490
d.f. = 2
34.549 34.000 50 -0.168
37.919 40.000 50 0.687
40.083 38.000 50 -0.739
42.449 43.000 50 0.218
P-value = 0.5788
Benchmark Dose Computation
Specified effect
Risk Type
Confidence level
BMD
BMDL
0.1
Extra risk
0. 95
7.96586
3.16409
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INCREASED INCIDENCE OF SCIATIC NERVE DEGENERATION IN FEMALE
F334/N RATS TREATED WITH SAN TRIMER FOR 2 YEARS (MP. 2012)
All available dichotomous models in BMDS (version 2.2.2) were fit to the sciatic nerve
degeneration data from female F344/N rats treated with SAN Trimer for 2 years CNTP. 2012)
(see Table A-12). A BMR of 10% extra risk was used in all model runs. The initial modeling of
these data including all dose groups failed to provide an adequate fit to the data, as assessed by
the x2 goodness-of-fit test. Therefore, only the BMD modeling results based on data without the
high-dose group included are summarized in Table B-3 and Figure B-3. As assessed by the
X2 goodness-of-fit statistic, AIC score, and visual inspection, after excluding the high-dose group
(85 mg/kg-day) in order to improve fit, the Probit model provided the best model fit (see
Table B-3 and Figure B-3). Estimated doses associated with 10% extra risk and the 95% lower
confidence limit on these doses (BMDio and BMDLio values, respectively) were 4.7 and
3.5 mg/kg-day.
Table B-3. Model Predictions for Sciatic Nerve Degeneration in Females"
Model
BMDio
BMDLio
X2/7-Value
AIC
Conclusion
Gamma
9.6
2.5
NDr
167.99
Logistic
4.5
3.2
0.605
166.26
Log-Logistic
10
1.6
NDr
167.99
LogProbit
11
4.7
NDr
167.99
Multistage
7.8
2.5
NDr
167.99
Probitb
4.7
3.5
0.659
166.18
Lowest AIC
Weibull
8.7
2.5
NDr
167.99
"NTP (20121 2-year study.
bThe Probit model was selected as a best-fitting model based on lowest AIC.
NDr = Not determined.
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Probit Model with 0.95 Confidence Level
Probit
BMD Lower Bound
0.9
0.7
0.6
0.5
0.4
BMDL
BMD
0
5
10
15
20
25
30
35
40
dose
13:14 07/12 2012
Figure B-3. Probit Model Fit for Sciatic Nerve Degeneration in Females (N I P. 2012)
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Text Output for Probit BMD Model for Increased Incidence of Sciatic Nerve Degeneration
in Female Rats Treated with SAN Trimer for 2 Years (NTP, 2012)
Probit Model. (Version: 3.2; Date: 10/28/2009)
Input Data File: C:/Documents and Settings/JKaiser/Desktop/modeling
results/pro_santa_srd_f_Pro-BMR10.(d)
Gnuplot Plotting File: C:/Documents and Settings/JKaiser/Desktop/modeling
results/pro_santa_srd_f_Pro-BMR10.pit
Thu Jul 12 13:14:24 2012
BMDS Model Run
The form of the probability function is:
P[response] = CumNorm(Intercept+Slope*Dose) ,
where CumNormf .) is the cumulative normal distribution function
Dependent variable = Response
Independent variable = Dose
Slope parameter is not restricted
Total number of observations = 3
Total number of records with missing values = 0
Maximum number of iterations = 250
Relative Function Convergence has been set to: le-008
Parameter Convergence has been set to: le-008
Default Initial (and Specified) Parameter Values
background = 0 Specified
intercept = 0.144466
slope = 0.0241571
Asymptotic Correlation Matrix of Parameter Estimates
( *** The model parameter(s) -background
have been estimated at a boundary point, or have been specified by
the user,
and do not appear in the correlation matrix )
intercept slope
intercept 1 -0.73
slope -0.73 1
Parameter Estimates
95.0% Wald Confidence
Interval
Variable Estimate Std. Err. Lower Conf. Limit Upper Conf.
Limit
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intercept
0.477264
slope
0.0381782
0.150347
0.0241112
0.166797
0.00717716
-0.176569
0.0100442
Analysis of Deviance Table
Model
Full model
Fitted model
Reduced model
AIC:
Log(likelihood)
-80.9947
-81.0914
-87.0126
166.183
# Param's Deviance Test d.f. P-value
3
2 0.193293 1 0.6602
1 12.0357 2 0.002435
Dose
Goodness of Fit
Est._Prob. Expected Observed Size
Scaled
Residual
0.0000
20.0000
40.0000
Chi^2 = 0.19
0.5598
0.7365
0.8675
d.f. = 1
27.428 28.000 49
36.088 35.000 49
42.509 43.000 49
P-value = 0.65 93
0.165
-0.353
0.207
Benchmark Dose Computation
Specified effect
Risk Type
Confidence level
BMD
BMDL
0.1
Extra risk
0. 95
4 . 67809
3.45186
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APPENDIX C. DOSIMETRY CALCULATION EXAMPLES FOR F0 DAMS AND F1
PUPS IN THE 2-WEEK, 13-WEEK, AND 2-YEAR PERINATAL AND POSTNATAL
FEED STUDIES OF SAN TRIMER (NTP. 2012)
NTP (2012) 2-WEEK STUDY
For this study, the study authors reported doses from GD 7 to 14, GD 15 to 18, PND 1 to
7, PND 8 to 14, and PND 15 to 20. A time-weighted average (TWA) dose was calculated for
both the gestational and lactational components of the study.
Gestational Component:
TWA Dose = (Di x TO + (D? x T?)
T1 + T2
Where
D1 = Dose received from GD 7 to 14 with T1 = 8 days
D2 = Dose received from GD 15 to 18 with T2 = 4 days.
Example for 250 ppm:
D1 = 17 mg/kg-day as calculated by the study authors, T1 = 8 days; D2 = 20 mg/kg-day as
calculated by the study authors, T2 = 4 days.
TWA Dose = (17 mg/kg-day x 8 days) + (20 mg/kg-day x 4 days)
12 days
TWA Dose = 18 mg/kg-day
Lactational Component:
TWA Dose
(Di x Ti) + (D2 x T2) + (D3 x TO
T1 + T2 + T3
Where
Di = Dose received from PND 1 to 7 with Ti = 7 days
D2 = Dose received from PND 8 to 14 with T2 = 7 days
D3 = Dose received from PND 15 to 20 with T3 = 6 days.
Example for 250 ppm:
Di = 30 mg/kg-day as calculated by the study authors, Ti = 7 days
D2 = 43 mg/kg-day as calculated by the study authors, T2 = 7 days
D3 = 49 mg/kg-day as calculated by the study authors, T2= 6 days.
TWA Dose = (30 mg/kg-day x 7 days) + (43 mg/kg-day x 7 days) + (49 mg/kg-day x 6 days)
20 days
TWA Dose = 40 mg/kg-day
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NTP (2012) 13-WEEK STUDY
For the 13-week and 2-year studies (NTP. 2012). the study authors did not present food
consumption data during gestation and lactation so it was not possible to calculate food factors
for these studies. Therefore, the food factors were calculated from the body weight and food
consumption data reported in dams from the 2-week NTP (2012) study and used to calculate
gestational and lactational doses for the 13-week and 2-year NTP (2012) studies.
Dose calculation examples for the gestational component of the 13-week NTP (2012)
study:
Table C-l. Average Food Factor Calculations for the Gestational Component of the
2-Week NTP (2012) Studv
GD 7-14
PPM
Food Consumption/Day3
Body Weight3
Food Factorb
0
13
191
0.068
250
14
196
0.071
500
12
185
0.065
1,000
12
197
0.061
2,000
11
190
0.058
Average food factor for GD 7-14 in the 2-week NTP study
0.065
GD 15-18
PPM
Food Consumption/Day"
Body Weight3
Food Factorb
0
16
213
0.075
250
17
219
0.078
500
16
206
0.078
1,000
17
220
0.077
2,000
16
213
0.075
Average food factor for GD 15-18 in the 2-week NTP study
0.077
"Data were reported by the study authors.
bFood factor = Food consumption per day x (1 -f- Body Weight).
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Table C-2. Dose Calculations for the Gestational Component of the 13-Week NTP (2012)
Study
PPM
Average Food Factor
(GD 7-14)a
Dose
(GD 7-14)b
Average Food Factor
(GD 15-18)a
Dose
(GD 15-18)b
TWA
Dosec
0
0.065
0
0.077
0
0
100
0.065
6.5
0.077
7.7
6.9
200
0.065
13
0.077
15
14
400
0.065
26
0.077
31
28
800
0.065
52
0.077
62
55
1600
0.065
104
0.077
123
110
11 As presented in Table C-l.
bDose = Feed concentration x food factor x (days dosed total days).
°TWA doses were calculated as described above.
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Dose calculation examples for the lactational component of the 13-week NTP (2012)
study:
Table C-3. Average Food Factor Calculations for the Lactational Component of the
2-Week NTP (2012) Studv
PND 1-7
PPM
Food Consumption/Day"
Body Weight"
Food Factorb
0
24
204
0.12
250
26
214
0.12
500
25
199
0.13
1,000
26
210
0.12
2,000
23
202
0.11
Average food factor for PND 1-7 in the 2-week NTP study
0.12
PND 8-14
PPM
Food Consumption/Day3
Body Weight3
Food Factorb
0
41
222
0.18
250
39
230
0.17
500
39
215
0.18
1,000
40
227
0.18
2,000
40
216
0.19
Average food factor for PND 8-14 in the 2-week NTP study
0.18
PND 15-20
PPM
Food Consumption/Day3
Body Weight3
Food Factorb
0
47
234
0.20
250
46
238
0.19
500
47
226
0.21
1,000
48
238
0.20
2,000
43
222
0.19
Average food factor for PND 15-20 in the 2-week NTP study
0.20
aData were reported by the study authors.
bFood factor = Food consumption per day x (1 -f- Body Weight).
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Table C-4. Dose Calculations for the Lactational Component of the 13-Week NTP (2012)
Study
PPM
Average Food
Factor
(PND l-7)a
Dose
(PND l-7)b
Average Food
Factor
(PND 8-14)a
Dose
(PND 8-14)b
Average Food
Factor
(PND 15-20)a
Dose
(PND 15-20)b
TWA
Dosec
0
0.12
0
0.18
0
0.20
20
0
100
0.12
12
0.18
18
0.20
40
17
200
0.12
24
0.18
36
0.20
80
33
400
0.12
48
0.18
72
0.20
160
66
800
0.12
96
0.18
144
0.20
320
132
1600
0.12
192
0.18
288
0.20
20
264
aAs presented in Table C-3.
bDose = Feed concentration x food factor x (days dosed total days).
°TWA doses were calculated as described above.
NTP (2012) 2-YEAR STUDY
The same methodology used for the 13-week study was also used to perform dosimetry
calculations for the gestational and lactational components of the 2-year NTP (2012) study and
are not shown here.
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