SEPA
EPA/690/R-20/005F | September 2020 | FINAL
United States
Environmental Protection
Agency
Provisional Peer-Reviewed Toxicity Values for
N,N,N/,N"-Tetramethylphosphoramide (TMPA)
(CASRN 16853-36-4)
U.S. EPA Office of Research and Development
Center for Public Health and Environmental Assessment
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JBI 1#%, United Slates
Environmental Protection
ImI M * Agency
EPA/690/R-20/005F
September 2020
https ://www. epa. gov/pprtv
Provisional Peer-Reviewed Toxicity Values for
N,N,N',N"-Tetramethylphosphoramide (TMPA)
(CASRN 16853-36-4)
Center for Public Health and Environmental Assessment
Office of Research and Development
U.S. Environmental Protection Agency
Cincinnati, OH 45268
ii
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AUTHORS, CONTRIBUTORS, AND REVIEWERS
CHEMICAL MANAGER
Jeffry L. Dean II, PhD
Center for Public Health and Environmental Assessment, Cincinnati, OH
DRAFT DOCUMENT PREPARED BY
SRC, Inc.
7502 Round Pond Road
North Syracuse, NY 13212
PRIMARY INTERNAL REVIEWERS
Jay Zhao, PhD
Center for Public Health and Environmental Assessment, Cincinnati, OH
Lucina Lizarraga, PhD
Center for Public Health and Environmental Assessment, Cincinnati, OH
This document was externally peer reviewed under contract to:
Eastern Research Group, Inc.
110 Hartwell Avenue
Lexington, MA 02421-3136
Questions regarding the content of this PPRTV assessment should be directed to the U.S. EPA
Office of Research and Development (ORD) CPHEA website at
https://www.epa.gov/pprtv/forms/contact-us-about-pprtvs.
in
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TABLE OF CONTENTS
COMMONLY USED ABBREVIATIONS AND ACRONYMS v
BACKGROUND 1
QUALITY ASSURANCE 1
DISCLAIMERS 2
QUESTIONS REGARDING PPRTVs 2
INTRODUCTION 3
REVIEW OF POTENTIALLY RELEVANT DATA (NONCANCER AND CANCER) 6
HUMAN STUDIES 8
ANIMAL STUDIES 8
OTHER DATA (SHORT-TERM TESTS, OTHER EXAMINATIONS) 8
Toxicokinetics 8
Genotoxicity 8
DERIVATION 01 PROVISIONAL VALUES 9
DERIVATION 01 ORAL REFERENCE DOSES 9
DERIVATION OF INHALATION REFERENCE CONCENTRATIONS 9
CANCER WEIGHT-OF-EVIDENCE DESCRIPTOR 9
DERIVATION OF PROVISIONAL CANCER RISK ESTIMATES 10
APPENDIX A. SCREENING NONCANCER PROVISIONAL VALUES 11
APPENDIX B. BACKGROUND AND METHODOLOGY FOR THE SCREENING
EVALUATION OF POTENTIAL CARCINOGENICITY 24
APPENDIX C. RESULTS OF THE SCREENING EVALUATION OF POTENTIAL
CARCINOGENICITY 32
APPENDIX D. REFERENCES 39
iv
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COMMONLY USED ABBREVIATIONS AND ACRONYMS
a2u-g
alpha 2u-globulin
IVF
in vitro fertilization
ACGIH
American Conference of Governmental
LC50
median lethal concentration
Industrial Hygienists
LD50
median lethal dose
AIC
Akaike's information criterion
LOAEL
lowest-observed-adverse-effect level
ALD
approximate lethal dosage
MN
micronuclei
ALT
alanine aminotransferase
MNPCE
micronucleated polychromatic
AR
androgen receptor
erythrocyte
AST
aspartate aminotransferase
MOA
mode of action
atm
atmosphere
MTD
maximum tolerated dose
ATSDR
Agency for Toxic Substances and
NAG
7V-acetyl-P-D-glucosaminidase
Disease Registry
NCI
National Cancer Institute
BMC
benchmark concentration
NOAEL
no-observed-adverse-effect level
BMCL
benchmark concentration lower
NTP
National Toxicology Program
confidence limit
NZW
New Zealand White (rabbit breed)
BMD
benchmark dose
OCT
ornithine carbamoyl transferase
BMDL
benchmark dose lower confidence limit
ORD
Office of Research and Development
BMDS
Benchmark Dose Software
PBPK
physiologically based pharmacokinetic
BMR
benchmark response
PCNA
proliferating cell nuclear antigen
BUN
blood urea nitrogen
PND
postnatal day
BW
body weight
POD
point of departure
CA
chromosomal aberration
PODadj
duration-adjusted POD
CAS
Chemical Abstracts Service
QSAR
quantitative structure-activity
CASRN
Chemical Abstracts Service registry
relationship
number
RBC
red blood cell
CBI
covalent binding index
RDS
replicative DNA synthesis
CHO
Chinese hamster ovary (cell line cells)
RfC
inhalation reference concentration
CL
confidence limit
RfD
oral reference dose
CNS
central nervous system
RGDR
regional gas dose ratio
CPHEA
Center for Public Health and
RNA
ribonucleic acid
Environmental Assessment
SAR
structure activity relationship
CPN
chronic progressive nephropathy
SCE
sister chromatid exchange
CYP450
cytochrome P450
SD
standard deviation
DAF
dosimetric adjustment factor
SDH
sorbitol dehydrogenase
DEN
diethylnitrosamine
SE
standard error
DMSO
dimethylsulfoxide
SGOT
serum glutamic oxaloacetic
DNA
deoxyribonucleic acid
transaminase, also known as AST
EPA
Environmental Protection Agency
SGPT
serum glutamic pyruvic transaminase,
ER
estrogen receptor
also known as ALT
FDA
Food and Drug Administration
SSD
systemic scleroderma
FEVi
forced expiratory volume of 1 second
TCA
trichloroacetic acid
GD
gestation day
TCE
trichloroethylene
GDH
glutamate dehydrogenase
TWA
time-weighted average
GGT
y-glutamyl transferase
UF
uncertainty factor
GSH
glutathione
UFa
interspecies uncertainty factor
GST
glutathione-S-transferase
UFC
composite uncertainty factor
Hb/g-A
animal blood-gas partition coefficient
UFd
database uncertainty factor
Hb/g-H
human blood-gas partition coefficient
UFh
intraspecies uncertainty factor
HEC
human equivalent concentration
UFl
LOAEL-to-NOAEL uncertainty factor
HED
human equivalent dose
UFS
subchronic-to-chronic uncertainty factor
i.p.
intraperitoneal
U.S.
United States of America
IRIS
Integrated Risk Information System
WBC
white blood cell
Abbreviations and acronyms not listed on this page are defined upon first use in the
PPRTV document.
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PROVISIONAL PEER-REVIEWED TOXICITY VALUES FOR
N,N,N ',N "-TETRAMETHYLPHOSPHORAMIDE (TMPA) (CASRN 16853-36-4)
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.
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.
Currently available PPRTV assessments can be accessed on the U.S. Environmental
Protection Agency's (EPA's) PPRTV website at https://www.epa.gov/pprtv. PPRTV
assessments are eligible to be updated on a 5-year cycle and revised as appropriate to incorporate
new data or methodologies that might impact the toxicity values or affect the characterization of
the chemical's potential for causing adverse human-health effects. Questions regarding
nomination of chemicals for update can be sent to the appropriate U.S. EPA Superfund and
Technology Liaison (https://www.epa.gov/research/fact-sheets-regional-science).
QUALITY ASSURANCE
This work was conducted under the U.S. EPA Quality Assurance (QA) program to ensure
data are of known and acceptable quality to support their intended use. Surveillance of the work
by the assessment managers and programmatic scientific leads ensured adherence to QA
processes and criteria, as well as quick and effective resolution of any problems. The QA
manager, assessment managers, and programmatic scientific leads have determined under the
QA program that this work meets all U.S. EPA quality requirements. This PPRTV was written
with guidance from the CPHEA Program Quality Assurance Project Plan (PQAPP), the QAPP
titled Program Quality Assurance Project Plan (PQAPP) for the Provisional Peer-Reviewed
Toxicity Values (PPRTVs) and Related Assessments/Documents (L-CPAD-0032718-QP), and the
PPRTV development contractor QAPP titled Quality Assurance Project Plan—Preparation of
Provisional Toxicity Value (PTV) Documents (L-CPAD-0031971-QP). As part of the QA
system, a quality product review is done prior to management clearance. A Technical Systems
Audit may be performed at the discretion of the QA staff.
All PPRTV assessments receive internal peer review by at least two Center for Public
Health and Environmental Assessment (CPHEA) scientists and an independent external peer
review by at least three scientific experts. The reviews focus on whether all studies have been
correctly selected, interpreted, and adequately described for the purposes of deriving a
provisional reference value. The reviews also cover quantitative and qualitative aspects of the
provisional value development and address whether uncertainties associated with the assessment
have been adequately characterized.
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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. 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.
This document has been reviewed in accordance with U.S. EPA policy and approved for
publication. Mention of trade names or commercial products does not constitute endorsement or
recommendation for use.
QUESTIONS REGARDING PPRTVS
Questions regarding the content of this PPRTV assessment should be directed to the
U.S. EPA Office of Research and Development (ORD) CPHEA website at
https://www.epa.gov/pprtv/forms/contact-us-about-pprtvs.
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INTRODUCTION
N,N,N',N"-Tetramethylphosphoramide (TMPA), CASRN 16853-36-4, belongs to the
class of compounds known as phosphoramides. It is used as a solvent (Yang et al.. 2011).
TMPA is not listed on U.S. EPA's Toxic Substances Control Act's public inventory (U.S. HP A.
2018b). nor is it registered with Europe's Registration, Evaluation, Authorisation and Restriction
of Chemicals (REACH) program (ECHA. 2018).
The empirical formula for TMPA is C4H14N3OP, and its structure is shown in Figure 1.
Table 1 summarizes the physicochemical properties of TMPA. TMPA is a liquid at room
temperature. TMPA's moderate vapor pressure and Henry's law constant indicate that it is not
expected to volatilize from either dry or moist surfaces. If TMPA does partition to the
atmosphere, the estimated vapor pressure indicates that it will exist there almost entirely as a
vapor. The estimated half-life of vapor-phase TMPA in air by reaction with photochemically
produced hydroxyl radicals is 1.5 hours. The estimated high water solubility, and low soil
adsorption coefficient for TMPA indicate that it may leach to groundwater or undergo runoff
after a rain event.
HjC
O
\
f
/
P
/
H
HX
5
Figure 1. N,N,N',N"-Tetramethylphosphoramide
(CASRN 16853-36-4) Structure
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Table 1. Physicochemical Properties of TMPA (CASRN 16853-36-4)a
Property (unit)
Value3
Physical state
Liquid
Boiling point (°C)
226 (predicted average)
Melting point (°C)
69.4 (predicted average)
Density (g/cm3)
1.12 (predicted average)
Vapor pressure (mm Hg at 25°C)
0.243 (predicted average)
pH (unitless)
NV
pKa (unitless)
NV
Water solubility (mol/L)
4.44 (predicted average)
Octanol-water partition coefficient (log P)
-1.14 (predicted average)
Henry's law constant (atm-m3/mol at 25°C)
8.75 x 10 x (predicted average)
Soil adsorption coefficient Koc (L/kg)
20.8 (predicted average)
Atmospheric OH rate constant (cm3/molecule-sec at 25°C)
4.67 x 10 11 (predicted average)
Atmospheric half-life (hours)
NV
Relative vapor density (air = 1)
NV
Molecular weight (g/mol)
151.15
Flash point (°C)
67.5 (predicted average)
aData were extracted from the U.S. EPA CompTox Chemicals Dashboard (N,N,N',N"-tetramethylphosphoramide
CASRN 16853-36-4; https://comptox.epa.gov/dashboard/dsstoxdb/results7searcliFDTXSIP80168576: accessed
April 20, 2020). All values are experimental averages unless otherwise specified.
NA = not applicable; NY = not available; TMPA = N,N,N',N"-tetramethylphosphoramide.
There are no available toxicity values for TMPA from U.S. EPA or other
agencies/organizations (see Table 2).
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Table 2. Summary of Available Toxicity Values for TMPA (CASRN 16853-36-4)
Source3
Value
Notes
Reference(s)b
Noncancer
IRIS
NV
NA
U.S. EPA (2018a)
HEAST
NV
NA
U.S. EPA (2011b)
DWSHA
NV
NA
U.S. EPA (2012a)
ATSDR
NV
NA
ATSDR (2017)
IPCS
NV
NA
IPCS (2018)
CalFPA
NV
NA
CalEPA (2016): CalEPA (2018a): CalEPA (2018b)
OSHA
NV
NA
OSHA (2017b): OSHA (2017a)
NIOSH
NV
NA
NIOSH (2016)
ACGIH
NV
NA
ACGIH (2017)
Cancer
IRIS
NV
NA
U.S. EPA (2018a)
HEAST
NV
NA
U.S. EPA (2011b)
DWSHA
NV
NA
U.S. EPA (2012a)
NTP
NV
NA
NTP (2016)
IARC
NV
NA
IARC (2018)
CalEPA
NV
NA
CalEPA (2011): CalEPA (2018a): CalEPA (2018b)
ACGIH
NV
NA
ACGIH (2017)
aSources: ACGIH = American Conference of Governmental Industrial Hygienists; ATSDR = Agency for Toxic
Substances and Disease Registry; CalEPA = California Environmental Protection Agency; DWSHA = Drinking
Water Standards and Health Advisories; HEAST = Health Effects Assessment Summary Tables;
IARC = International Agency for Research on Cancer; IPCS = International Programme on Chemical Safety;
IRIS = Integrated Risk Information System; NIOSH = National Institute for Occupational Safety and Health;
NTP = National Toxicology Program; OSHA = Occupational Safety and Health Administration.
bReference date is the publication date for the database and not the date the source was accessed.
NA = not applicable; NY = not available; TMPA = N,N,N',N"-tetramethylphosphoramide.
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Non-date-limited literature searches were conducted in March 2018 and updated in
January 2020 for studies relevant to the derivation of provisional toxicity values for TMPA
(CASRN 16853-36-4). The database searches for PubMed, TOXLINE (including TSCATS1),
and Web of Science were conducted by an information specialist and records stored in the
U.S. EPA's Health and Environmental Research Online (HERO) database. The following
additional databases were searched for health-related data: American Conference of
Governmental Industrial Hygienists (ACGIH), Agency for Toxic Substances and Disease
Registry (ATSDR), California Environmental Protection Agency (CalEPA), European Centre for
Ecotoxicology and Toxicology of Chemicals (ECETOC), European Chemicals Agency (ECHA),
U.S. EPA Integrated Risk Information System (IRIS), U.S. EPA Health Effects Assessment
Summary Tables (HEAST), U.S. EPA Office of Water (OW), U.S. EPA TSCATS2/TSCATS8e,
U.S. EPA High Production Volume Information System (HPVIS), International Agency for
Research on Cancer (IARC), International Programme on Chemical Safety (IPCS)/INCHEM,
Japan Existing Chemical Data Base (JECDB), National Institute for Occupational Safety and
Health (NIOSH), National Toxicology Program (NTP), Organisation for Economic Co-operation
and Development (OECD) Screening Information Data Sets (SIDS), International Uniform
Chemical Information Database (IUCLID), High Production Volume (HPV), Occupational
Safety and Health Administration (OSHA), and World Health Organization (WHO).
REVIEW OF POTENTIALLY RELEVANT DATA
(NONCANCER AND CANCER)
As shown in Tables 3 A and 3B, there are no potentially relevant short-term, subchronic,
or chronic studies or developmental or reproductive toxicity studies in humans or animals. The
phrase "statistical significance" or the term "significant," used throughout the document,
indicates ap-walue of < 0.05 unless otherwise noted.
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Table 3A. Summary of Potentially Relevant Noncancer Data for TMPA (CASRN 16853-36-4)
Category
Number of Male/Female, Strain, Species, Study
Type, Reported Doses, Study Duration
Dosimetry
Critical Effects
Reference
Notes
Human
1. Oral (mg/kg-d)
ND
2. Inhalation (mg/m3)
ND
Animal
1. Oral (mg/kg-d)
ND
2. Inhalation (mg/m3)
ND
ND = no data; TMPA = N,N,N',N"-tetramethylphosphoramide.
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Table 3B. Summary of Potentially Relevant Cancer Data for TMPA (CASRN 16853-36-4)
Category
Number of Male/Female, Strain, Species, Study Type,
Reported Doses, Study Duration
Dosimetry
Critical Effects
Reference
Notes
Human
1. Oral (mg/kg-d)
ND
2. Inhalation (mg/m3)
ND
Animal
1. Oral (mg/kg-d)
ND
2. Inhalation (mg/m3)
ND
ND = no data; TMPA = N,N,N',N"-tetramethylphosphoramide.
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HUMAN STUDIES
No human exposure studies have been identified.
ANIMAL STUDIES
No animal exposure studies have been identified.
OTHER DATA (SHORT-TERM TESTS, OTHER EXAMINATIONS)
Toxicokinetics
There are no toxicokinetic data for TMPA following oral or inhalation exposure.
However, some metabolic and excretion information can be inferred from studies of related
phosphoramides, including hexamethylphosphoramide (HMPA), and
pentamethylphosphoramide (PMPA). HMPA, the parent chemical to TMPA, is broken down
into lower methylphosphoramides via sequential oxidative demethylation by cytochrome P450s
(CYP450s), yielding PMPA, TMPA, and N,N',N"-trimethylphosphoramide (triMPA). At each
demethylation step, the reaction forms unstable methylol intermediates that break down into the
demethylated phosphoramides, releasing formaldehyde in the process (Jones, 1970; Jones and
Jackson. 1968). A minor metabolic pathway for HMPA and PMPA involves the formation of
A-formyl-PMPA and A-formyl-TMPA, respectively, from the unstable methylol intermediates
(Jones. 1970). The toxicological consequences of products generated through this minor
metabolic pathway is currently unclear. A schematic of the proposed metabolism of TMPA can
be found in Appendix A (see Figure A-l).
Genotoxicity
The genotoxicity data for TMPA are limited to a single study using an unknown isomer
of TMPA at doses up to 5 mM. The study reported no induction of mitotic recombination in the
Drosophila melanogaster white/white (w/w+) eye mosaic system (Vogel and Nivard. 1993).
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DERIVATION OF PROVISIONAL VALUES
DERIVATION OF ORAL REFERENCE DOSES
No studies have been located regarding toxicity of TMPA to humans or animals by oral
exposure; therefore, subchronic and chronic provisional reference doses (p-RfDs) were not
derived directly. Instead, screening subchronic and chronic p-RfDs are developed in
Appendix A using an alternative analogue surrogate approach. Based on the overall analogue
approach presented in Appendix A, HMPA was selected as the most appropriate analogue for
TMPA for deriving a screening subchronic and chronic p-RfD (see Table 4).
DERIVATION OF INHALATION REFERENCE CONCENTRATIONS
No studies have been located regarding toxicity of TMPA to humans or animals by
inhalation; therefore, subchronic and chronic provisional reference concentrations (p-RfCs) were
not derived directly. An alternative analogue approach was not attempted for the p-RfCs
because of limitations in the inhalation toxicity data for the candidate analogue
(see Appendix A).
Table 4. Summary of Noncancer Reference Values for TMPA (CASRN 16853-36-4)
Toxicity Type
(units)
Species/
Sex
Critical
Effect
p-Reference
Value
POD
Method
POD
(HED)
UFc
Principal
Study
Screening
subchronic p-RfD
(mg/kg-d)
Rats/M
Increased
incidence of
nasal lesions
1 x 1(T3
NO A F.I.
0.29 (based on
analogue POD)
300
Keller etal. (1997)
as cited in U.S.
EPA (2012b)
Screening
chronic p-RfD
(mg/kg-d)
Rats/M
Increased
incidence of
nasal lesions
i x i(r4
NO A F.I.
0.29 (based on
analogue POD)
3,000
Keller etal. (1997)
as cited in U.S.
EPA (2012b)
Subchronic
p-RfC (mg/m3)
NDr
Chronic p-RfC
(mg/m3)
NDr
HED = human equivalent dose; M = male(s); NDr = not determined; NOAEL = no-observed-adverse-effect level;
POD = point of departure; p-RfC = provisional reference concentration; p-RfD = provisional reference dose;
TMPA = N,N,N',N"-tetramethylphosphoramide; UFC = composite uncertainty factor.
CANCER WEIGHT-OF-EVIDENCE DESCRIPTOR
Under the U.S. EPA Cancer Guidelines (U.S. EPA. 2005). there is "Inadequate
Information to Assess the Carcinogenic Potential" of TMPA (see Table 5). No relevant studies
are available in humans or animals. Within the current U.S. EPA Cancer Guidelines (U.S. EPA.
2005). there is no standard methodology to support the identification of a weight-of-evidence
(WOE) descriptor and derivation of provisional cancer risk estimates for data-poor chemicals
using an analogue approach. In the absence of an established framework, a screening evaluation
of potential carcinogenicity is provided using the methodology described in Appendix B. A
screening evaluation of potential carcinogenicity is provided using methodology described in
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Appendix B. This evaluation determined that there was a qualitative level of concern for
potential carcinogenicity for TMPA (see Appendix C).
Table 5. Cancer WOE Descriptor for TMPA (CASRN 16853-36-4)
Possible WOE Descriptor
Designation
Route of Entry
(oral, inhalation, or both)
Comments
"Carcinogenic to Humans"
NS
NA
There are no human carcinogenicity
data identified to support this descriptor.
"Likely to Be Carcinogenic
to Humans "
NS
NA
There are no animal carcinogenicity
studies identified to support this
descriptor.
"Suggestive Evidence of
Carcinogenic Potential"
NS
NA
There are no animal carcinogenicity
studies identified to support this
descriptor.
"Inadequate Information to
Assess Carcinogenic
Potential"
Selected
Both
This descriptor is selected due to the
lack of adequate data in humans or
animals to evaluate the carcinogenic
potential of TMPA.
"Not Likely to Be
Carcinogenic to Humans"
NS
NA
No evidence of noncarcinogenicity is
available.
NA = not applicable; NS = not selected; TMPA = N,N,N',N"-tetramethylphosphoramide; WOE = weight of
evidence.
DERIVATION OF PROVISIONAL CANCER RISK ESTIMATES
The absence of suitable data precludes development of cancer risk estimates for TMPA
(see Table 6).
Table 6. Summary of Cancer Risk Estimates for TMPA (CASRN 16853-36-4)
Toxicity Type (units)
Species/Sex
Tumor Type
Cancer Risk Estimate
Principal Study
p-OSF (mg/kg-d) 1
NDr
p-IUR (lng/in3) 1
NDr
NDr = not determined; p-IUR = provisional inhalation unit risk; p-OSF = provisional oral slope factor;
TMPA = N,N,N',N"-tetramethylphosphoramide.
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APPENDIX A. SCREENING NONCANCER PROVISIONAL VALUES
For reasons noted in the main Provisional Peer-Reviewed Toxicity Value (PPRTV)
document, it is inappropriate to derive provisional toxicity values for
N,N,N',N"-tetramethylphosphoramide (TMPA). However, information is available for this
chemical, which although insufficient to support deriving a provisional toxicity value under
current guidelines, may be of use to risk assessors. In such cases, the Center for Public Health
and Environmental Assessment (CPHEA) summarizes available information in an appendix and
develops a "screening value." Appendices receive the same level of internal and external
scientific peer review as the provisional reference values to ensure their appropriateness within
the limitations detailed in the document. Users of screening toxicity values in an appendix to a
PPRTV assessment should understand that there could be more uncertainty associated with the
derivation of an appendix screening toxicity value than for a value presented in the body of the
assessment. Questions or concerns about the appropriate use of screening values should be
directed to the CPHEA.
APPLICATION OF AN ALTERNATIVE ANALOGUE APPROACH
The analogue approach allows for the use of data from related compounds to calculate
screening values when data for the compound of interest are limited or unavailable. Details
regarding searches and methods for analogue analysis are presented in Wang et al. (2012).
Three types of potential analogues (structural, metabolic, and toxicity-like) are identified to help
select the final analogue chemical. The analogue approach may or may not be route specific or
applicable to multiple routes of exposure. All information was considered together as part of the
final weight-of-evidence (WOE) approach to select the most suitable analogue both
toxicologically and chemically.
Structural Analogues
An initial analogue search focused on identifying structurally similar chemicals with
toxicity values from the Integrated Risk Information System (IRIS), PPRTV, Agency for Toxic
Substances and Disease Registry (ATSDR), or California Environmental Protection Agency
(CalEPA) databases to take advantage of the well-characterized chemical-class information.
Under Wang et al. (20121 structural similarity for analogues is typically evaluated using
U.S. EPA's DSSTox database and the National Library of Medicine's (NLM's) ChemlDplus
database (ChemlDplus. 2016). Additionally, the Organisation for Economic Co-operation and
Development (OECD) Toolbox was used to calculate structural similarity using the Tanimoto
method (a quantitative method shared with ChemlDplus) (OECD. 2018).
Upon expert review of the structural fragments contained within the target chemical
(TMPA), several phosphoramides were identified as potential structural analogues.
Hexamethylphosphoramide (HMPA), pentamethylphosphoramide (PMPA), and
N,N',N"-trimethylphosphoramide (triMPA) were identified as relevant toxicokinetic precursors
or metabolites of the target compound because they share sufficient structural similarity. Of
these, only HMPA was associated with a published toxicity value. As such, HMPA was the only
structural analogue to TMPA that was further considered (U.S. EPA. 2012b). Table A-1
summarizes the analogue's physicochemical properties and similarity scores. ChemlDplus and
OECD Toolbox similarity scores for HMPA were 77 and 29%, respectively. TMPA and HMPA
are phosphoric acid amide derivatives, differing by two methyl substitutions. Physicochemical
11 N,N,N',N"-Tetramethylphosphoramide (TMPA)
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properties (i.e., low octanol-water partition coefficient) for TMPA and HMPA suggest that both
of these compounds will be bioavailable following oral and inhalation exposure (see Table A-l).
Lastly, while not available for TMPA, a low pKa reported for HMPA suggests that HMPA may
be an irritant or corrosive (see Table A-l). This characteristic may promote portal-of-entry
effects. In total, HMPA is considered an appropriate structural analogue for TMPA based on
commonalities in structural and physicochemical properties.
Table A-l. Physicochemical Properties of TMPA (CASRN 16853-36-4) and its Candidate
Analogue3
Chemical
TMPA
HMPA
Structure
v\ li r
N P N
/ 1 H
H ch3
H3C 7 ch3
\ 1 /
N P N
/ II \
H3c y CH,
CASRN
16853-36-4
680-31-9
Molecular weight (g/mol)
151.15
179.204
OECD Toolbox similarity score (%)b
100
29
ChemlDplus similarity score (%)°
100
77
Melting point (°C)
69.4 (predicted average)
6.95
Boiling point (°C)
226 (predicted average)
233
Vapor pressure (mm Hg at 25°C)
0.243 (predicted average)
0.0460
Henry's law constant (atm-m3/mol at 25°C)
8.75 x 10 x (predicted average)
8.86 x io~4 (predicted average)
Water solubility (mol/L)
4.44 (predicted average)
1.87 (predicted average)
Octanol-water partition coefficient (log P)
-1.14 (predicted average)
0.195 (predicted average)
pKa
NA
<1.6b
aData were extracted from the U.S. EPA CompTox Chemicals Dashboard (NNN',N"-tetramethylphosphoramide,
CASRN 16853-36-4; https://comptox.epa.gov/dashboard/dsstoxdb/results?search=DTXSID80168576; and
hexamethylphosphoramide, CASRN 680-31-9;
https://comptox.epa.gov/dashboard/dsstoxdb/results?search=DTXSID6020694#properties: accessed April 20,
2020). All values are experimental averages unless otherwise specified.
bOECD (2018).
°ChemIDplus Advanced, similarity scores (ChemlDplus. 2018).
HMPA = hexamethylphosphoramide; NV = not available; OECD = Organisation for Economic Co-operation and
Development; TMPA = N,N,N',N"-tetramethylphosphoramide.
Metabolic Analogues
Table A-2 summarizes available toxicokinetic data for TMPA and its structurally similar
analogue, HMPA. As discussed in the "Toxicokinetics" section in the main document, there are
no toxicokinetic data following oral or inhalation exposure to TMPA; however, some metabolic
and excretion information can be inferred from studies of related phosphoramides, including its
parent compound and candidate analogue, HMPA. The induction of toxic responses with oral
12 N,N,N',N"-Tetramethylphosphoramide (TMPA)
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and inhalation administration of HMPA provides evidence for its absorption (U.S. EPA. 2012b).
and additional studies directly demonstrate that HMPA is well absorbed and distributed
throughout the body after both oral and inhalation administration. HMPA exhibits preferential
deposition in rat nasal tissue, a major target organ of toxicity for this chemical [Rickard and
Guilles (1982) as cited in Keller et al. (1997)1.
TMPA is a primary intermediate in the metabolism of HMPA and results from sequential
demethylation of the parent compound by cytochrome P450 (CYP450) (Dahl and Brezinski.
1985; Jones and Jackson. 1968). Formaldehyde is released from methylol intermediates at each
demethylation step (see Figure A-l) (U.S. EPA. 2012b; Jones and Jackson. 1968). These
unstable methylol intermediates of HMPA, PMPA, and TMPA can form minor metabolites,
including A-formyl-PMPA and A'-formyl-TMPA, respectively (Jones. 1970). TMPA was readily
detected in rat urine following gavage administration of HMPA or PMPA, and formation of its
demethylated product, triMPA, has been also demonstrated in animals (Jones and Jackson.
1968). Rapid elimination of the parent phosphoramide, HMPA (>90% elimination within
24 hours after inhalation exposure), has been reported in experimental animals [Rickard and
Quarles (1981) as cited in Keller et al. (1997)1.
Together, these findings provide support for the in vivo oxidative demethylation of
TMPA and suggest that TMPA could share a similar metabolic fate as HMPA in the body.
Furthermore, toxicokinetics data showed that HMPA is well absorbed and distributed throughout
the body after both oral and inhalation treatments, displaying preferential deposition in rat nasal
tissue, a major target organ of toxicity for this chemical [Rickard and Gillies (1982) as cited in
Keller et al. (1997)1. While it is obvious that inhalation exposure can promote nasal deposition
of inhaled chemicals, Rickard and Guilles (1982) used 14C-HMPA gavage exposure to
demonstrate that the naso-, ethmo-, and maxilloturbinate regions of rat nasal passages retained
3- to 7-fold more radioactivity (compared with 7-, 21-, and 25-fold increases after inhalation
exposure) than other tissues in the body.
Table A-2. ADME Data for TMPA (CASRN 16853-36-4) and Its Candidate Analogue
Type of Data
TMPA
HMPA
Structure
v\ n
N P N
/ | H
H ch3
HjC^
H,C ';J CH,
\ 1 /
N P N
/ II \
H3c y CHj
CASRN
16853-36-4
680-31-9
Absorption
Rate and extent of oral
absorption
ND
ND
Rate and extent of
inhalation absorption
ND
ND
13 N,N,N',N"-Tetramethylphosphoramide (TMPA)
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Table A-2. ADME Data for TMPA (CASRN 16853-36-4) and Its Candidate Analogue
Type of Data
TMPA
HMPA
Distribution
Extent of distribution
ND
Generally widespread distribution with evidence
of increased deposition of HMPA and its
metabolites (no information on specific
metabolites was provided) in nasal tissue
following oral or inhalation exposures [Rickard
and Gillies (1982) as cited in Keller et al. (1997)1
Metabolism
Pathways and enzymes
Oxidative demethylation by
CYPs; based on metabolic
pathways established following
oral administration of HMPA or
PMPA Clones. 1970; Jones and
Jackson. 1968)
Oxidative demethylation by CYP in liver, lung,
and nasal cavity; CYP2A homologues have been
shown to be active in metabolic activation of
HMPA in vitro (human CYP2A6, rat CYP2A3,
rabbit CYP2A10/11) (Thornton-Manning et al..
1997; Liu et al.. 1996; Gervasi et al.. 1991; Loneo
et al.. 1988; Dahl and Brezinski. 1985; Jones and
Jackson. 1968)
Metabolites
Unstable methylol intermediates
break down to form triMPA and
formaldehyde; based on
metabolic pathways established
following oral administration of
HMPA or PMPA
(see Fieure A-1) (TJ.S. EPA.
2012b: Jones. 1970; Jones and
Jackson. 1968)
Primary: Unstable methylol intermediates break
down to form demethylated phosphoramides
(PMPA, TMPA, triMPA); formaldehyde is
released at each demethylation step
(see Fieure A-l) (U.S. EPA. 2012b; Jones. 1970;
Jones and Jackson. 1968)
Minor: Unstable methylol intermediates break
down to form Y-forim 1-PMPA (Jones. 1970)
Excretion
Rate of excretion
ND
Excretion is rapid (i.e., 90% excreted within 24 hr
following inhalation exposure; "most" excreted
within 20 hr of i.p. injection) [Rickard and Gillies
(1982) as cited in Keller et al. (1997)1
Route of excretion
Metabolic excretion expected via
the urine, based on excretion of
TMPA and triMPA as metabolites
following oral administration of
HMPA or PMPA (Jones. 1970;
Jones and Jacksoa 1968)
Parent compound and metabolites excreted in
urine (>90% administered dose; relative amounts
of oarent/metabolites not auantified) (Jones. 1970;
Jones and Jackson. 1968)
ADME = absorption, distribution, metabolism, and excretion; CYP = cytochrome P;
HMPA = hexamethylphosphoramide; i.p. = intraperitoneal; ND = no data; PMPA = pentamethylphosphoramide;
TMPA = N,N,N',N"-tetramethylphosphoramide; triMPA = N,N',N"-trimethylphosphoramide.
14 N,N,N',N"-Tetramethylphosphoramide (TMPA)
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H,C.
V
/
CHj
mctaboic
CHj ondJtjcri
HjC CHj
HJB,
Ch^OH
|
HjC CH,
HCHO
4 H,C
/ ' \ II
jN P-
"V
CH.
/ V
HMPA
Proposed mettiylo) intermediate
PMPA
fc. HCHO
P. O
\ II /" H,C\
HjC ^CH, —p H,C CHj
N / u
/ \ * / \
H,C H HCHO HjC ^H
TriMPA TMPA
Figure A-l. Primary Pathway of HMPA (CASRN 680-31-9) Metabolism (U.S. EPA, 2012b)
The primary metabolic pathway by which HMPA and TMPA progressively break down
into "lower" sequentially demethylated methylphosphoramides (PMPA and triMPA), leading to
the production of formaldehyde, is of toxicological relevance, given that both HMPA and
formaldehyde are known nasal toxicants in the rat (U.S. EPA 2012b; Kerns et al.. 1983;
Swenberg et al., 1980). Based on the data available, it is assumed that the systemic distribution
of HMPA and localized (nasal) metabolism is likely to promote the production of formaldehyde
(in addition to lower phosphoramides) in nasal tissues following oral exposure to HMPA. Thus,
the nasal lesions observed following exposure to HMPA/PMPA/TMPA may be attributable to
the localized production of formaldehyde, consistent with the portal-of-entry nasal lesions
observed following inhalation exposure to formaldehyde. To this point, microsomal enzymes
isolated from the olfactory epithelium of dogs are capable of metabolizing HMPA more
efficiently than liver microsomal enzymes (Dahl et al.. 1982). Furthermore, the pattern and
severity of nasal lesions induced by HMPA after oral exposure coincides with the selective
deposition of HMPA and its metabolites (no information on specific metabolites was provided)
in the nasal epithelium (Keller et al.. 1997). Thus, metabolism is thought to play a role in the
respiratory tract toxicity of HMPA following both oral and inhalation exposure (Keller et al..
1997). Because HMPA and TMPA share a similar potential bioactivation pathway, HMPA is
considered an appropriate metabolic analogue for TMPA.
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Toxicity-Like Analogues
Table A-3 summarizes available oral toxicity values for TMPA and its potential
analogue, HMPA. There are no toxicity data for TMPA. Repeated-dose toxicity information for
HMPA, the candidate analogue, indicates that the primary targets of toxicity are the respiratory
system (particularly the nasal epithelium) and the testes (U.S. EPA. 2012b). The point of
departure (POD) for deriving provisional reference doses (p-RfDs) for HMPA is based on a
no-observed-adverse-effect level (NOAEL) of 1.2 mg/kg-day for increased incidence of nasal
lesions in Sprague-Dawley (S-D) rats (U.S. HP A. 2012b). The principal study used in the
HMPA assessment reported dose-related increases in nasal/respiratory tract lesions (epithelial
denudation, regeneration, or squamous metaplasia) at doses >15 mg/kg-day and testicular effects
(testicular atrophy and decreased testes weight) at a dose of 123 mg/kg-day after a 90-day
administration of HMPA via drinking water (Keller et al.. 1997). Nasal and pulmonary toxicity
were also found in other subchronic and chronic studies with gavage or dietary exposure to
HMPA at similar doses (see Table A-3) (Kimbrough and Gaines. 1966). Likewise, reproductive
studies provided further evidence for the male reproductive effects of HMPA occurring at doses
>40 mg/kg-day in rats and at a dose of 100 mg/kg-day in rabbits, indicating that these effects are
less sensitive than the upper respiratory tract toxicity of HMPA (see Table A-3) (Jackson and
Craig. 1966).
16 N,N,N',N"-Tetramethylphosphoramide (TMPA)
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Table A-3. Comparison of Available Toxicity Data for TMPA (CASRN 16853-36-4) and
Its Candidate Analogue
Type of Data
TMPA
HMPA
Structure
*\ n
N P N
/ 1 H
H ch3
H3C
h3c ^ ch3
\ /
N P N
/ II \
H3c y CHj
CASRN
16853-36-4
680-31-9
Repeated-dose toxicity—oral subchronic
Critical effects
NA
Increased incidence of nasal and tracheal lesions at 15 mg/kg-d in
males and 20 mg/kg-d in females [Keller et al. (1997) as cited in
U.S. EPA (2012b)l
Other effects
(in principal study)
NA
Testicular atrophy, reduced absolute and relative testes weight,
and reduced body weight in males at 123 mg/kg-d [Keller et al.
(1997) as cited in U.S. EPA (2012b)l
Species
NA
Rat
Duration
NA
90 d
Route
NA
Drinking water
Additional toxicity data
(from other studies)
ND
Additional data reported in the principal study, nasal and
pulmonary lesions have been reported in subchronic studies in
rats exposed to HMPA via gavage at >15 mg/kg-d or via diet at
-115 me/ke-d (only dose tested) (Keller et al.. 1997).
Another toxicity target following short-term- or
subchronic-HMPA exposure is the male reproductive system. In
rats, testicular atrophy has been observed at >40 mg/kg-d
(Kimbroueh and Gaines. 1966). decreased male fertility at
>50 mg/kg-d and decreased testicular weight at >100 mg/kg-d.
Decreased fertility and impaired spermatogenesis has been
reported in male rabbits following short-term exposure to
100 mg/kg-d. Fertility and sperm effects following short-term
exposure in rats and rabbits were reversible. No adverse
testicular effects were reported in mice following short-term
exposure to doses up to 500 me/ke-d (Jackson et al.. 1969).
No adverse reproductive or developmental effects were observed
in a 2-generation gavage study in rats at doses up to 10 mg/kg-d
(Shottetal.. 1971).
No subchronic inhalation data are available.
17 N,N,N',N"-Tetramethylphosphoramide (TMPA)
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Table A-3. Comparison of Available Toxicity Data for TMPA (CASRN 16853-36-4) and
Its Candidate Analogue
Type of Data
TMPA
HMPA
Repeated-dose toxicity—oral chronic
Additional toxicity data
(from other studies)
NA
A 2-yr dietary study in rats found increased incidence of lung
disease at doses >0.78 mg/kg-d. The study was inadequate for
quantitative risk assessment owing to limited data reporting and
lack of statistical analyses (Kimbroueh and Gaines. 1973).
In an inhalation cancer bioassay (limited by inadequate reporting
of study design and results), damage to the nasal tissues and nasal
tumors were observed at >0.37 mg/m3 (no other noncancer or
cancer effects were reported) [Lee and Trochimowicz (1984,
1982a. 1982b. 1982c) as cited in U.S. EPA (2012b)l.
HMPA = hexamethylphosphoramide; NA = not available; TMPA = N,N,N',N"-tetramethylphosphoramide.
HMPA is also a potent nasal toxicant via the inhalation route, yet provisional reference
concentrations (p-RfCs) were not derived because no subchronic inhalation studies were
available and the data from the lone chronic inhalation study (Lee and Trochimowicz. 1984.
1982a. b, c) were not sufficient to support a quantitative dose-response assessment. This was
due to significant uncertainties regarding: the experimental design, establishing clear
dose-response data (which requires a number of assumptions to be made), and data reporting.
For example, it is difficult to determine, based on the published results from this study, how
many rats were in each exposure group and at what point in time they were sacrificed. Various
tables in the published papers give conflicting accounts of the actual experimental design. The
number of animals exposed for each length of time is needed to derive a quantitative risk
assessment value for HMPA. These same uncertainties in experimental design and data
reporting in the inhalation bioassay for HMPA further prevented the data from being used to
derive provisional cancer potency values for this chemical (U.S. EPA, 2012b).
Metabolic transformation of HMPA to lower methylphosphoramides (including TMPA)
and formaldehyde is a proposed mechanism for the critical effects (nasal toxicity) of this
compound (U.S. EPA. 2012b; Keller et at.. 1997). Because both HMPA and TMPA share this
primary metabolic pathway (see "Metabolic Analogues" section and Table A-2 above), the
proposed mechanism of action for HMPA-induced nasal toxicity is plausible for TMPA, as well.
The testicular effects observed at higher HMPA doses may be related to the number of methyl
groups on the parent compound (and subsequent intracellular levels of formaldehyde produced as
a product of sequential demethylation), rather than metabolic products, as indicated by
reproductive studies in rodents and insects (Jones. 1970; Jackson et at.. 1969; Jones and Jackson.
1968). However, the absence of information specific to TMPA prevents an assessment of its
potential reproductive effects. While the lack of toxicity data for TMPA precludes the
determination of appropriate toxicity-like analogues, the putative bioactivation pathway for nasal
toxicity common to TMPA and HMPA indicates that TMPA is a potential respiratory tract
toxicant.
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Weight-of-Evidence Approach
A WOE approach is used to evaluate information from potential candidate analogues as
described by Wang et al. (2012). Commonalities in structural/physicochemical properties,
toxicokinetics, metabolism, toxicity, or mode of action (MOA) between potential analogues and
chemical(s) of concern are identified. Emphasis is given to toxicological and/or toxicokinetic
similarity over structural similarity. Analogue candidates are excluded if they do not have
commonality or demonstrate significantly different physicochemical properties and toxicokinetic
profiles that set them apart from the pool of potential analogues and/or chemical(s) of concern.
From the remaining potential analogues, the most biologically or toxicologically relevant
analogue chemical is selected based on MOA information, otherwise, the analogue with the
highest structural similarity and/or most conservative toxicity value is selected.
Oral Noncattcer
The only candidate compound identified, HMPA, is considered a structural analogue of
TMPA based on structural commonalities and physicochemical properties. HMPA is also a
parent chemical to TMPA and both compounds are metabolized similarly to lower
methylphosphoramides and formaldehyde via a potential bioactivation pathway for nasal
toxicity. Thus, HMPA is considered a metabolic analogue for TMPA. Nasal lesions (the critical
effect for HMPA) were attributed to systemic distribution of the metabolic product, or the parent
compound. Systemic distribution of the parent compound (or any of the expected methylol
intermediates) may also result in local metabolic production of formaldehyde. Given the
similarities in metabolic processing of HMPA and TMPA, nasal lesions are expected to be a
toxic effect of oral TMPA exposure, although direct evidence of such effect is currently lacking.
In total, HMPA is judged to be an appropriate analogue for TMPA for deriving screening
p-RfDs.
Inhalation Noncancer
No p-RfCs can be derived for TMPA, because p-RfCs were not derived in the PPRTV
assessment for HMPA (the selected analogue) because no subchronic inhalation studies were
available and the data from the lone chronic inhalation study were not sufficient to support a
quantitative dose-response assessment. This lone chronic inhalation study presented substantial
uncertainties regarding the experimental design, establishing clear dose-response data (which
requires a number of assumptions to be made), and data reporting (U.S. HP A. 2012b).
ORAL NONCANCER TOXICITY VALUES
Derivation of a Screening Subchronic Provisional Reference Dose
Based on the overall analogue approach presented in this PPRTV assessment, HMPA
was selected as the analogue for TMPA for deriving a screening subchronic p-RfD. A PPRTV
for HMPA is available. The study used to derive the PPRTV subchronic p-RfD for HMPA was a
90-day drinking water study in rats [Keller et al. (1997) as cited in U.S. EPA (2012bVI. The
PPRTV for HMPA describes this study as follows:
In a peer-reviewed study, Keller et al. (1997) evaluated the subchronic
nasal toxicity of HMPA administered to rats in drinking water and by gavage. It
was not stated whether the study was performed under GLP standards, but the
study appears scientifically sound. The study authors first conducted a drinking
water experiment in which four groups of 10 male and 10 female Charles
River-CD rats obtainedfrom Charles River Breeding Laboratories were
19 N,N,N',N"-Tetramethylphosphoramide (TMPA)
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administeredHMPA (99%pure) in drinking water at doses of0, 10, 100, 300, or
1,000 ppm (equivalent to approximately 0, 1.2, 15, 42, or 123 mg/kg-day in
males; 0, 2.3, 20, 63, or 229 mg/kg-day in females), 7 days/week, for 90 days.
The study authors state that the animals were caredfor in accordance with the
NIH Guide for Care and Use of Laboratory Animals and observed daily for
mortality and clinical signs of toxicity. Body weights, mean group food
consumptions, and mean group water consumption were determined weekly.
After 45 and 90 days of treatment, blood samples were collected for
10 rats/sex/group for hematology (erythrocyte, leukocyte, differential leukocyte,
platelet counts, hemoglobin, hematocrit, mean corpuscular hemoglobin, mean
corpuscular volume, and mean hemoglobin concentration) and clinical chemistry
(alkaline phosphatase, alanine aminotransferase, and aspartate aminotransferase
activities, and concentrations of blood urea nitrogen, total protein, potassium,
phosphate, and chloride). Urine was measuredfor volume, osmolality, pH,
glucose, protein, bilirubin, urobilinogen, ketone, and occult blood. At the end of
the treatment period, all surviving animals were sacrificed and necropsied.
Selected organs were weighed (liver, spleen, kidneys, heart, testes, and brain) and
histopathological examination was performed on comprehensive tissues: heart,
aorta (thoracic), trachea, lungs, nose, salivary glands, esophagus, stomach, liver,
pancreas, duodenum, jejunum, ileum, cecum, colon, rectum, femur, sternum, bone
marrow (sternum), mandibular lymph nodes, mesenteric lymph nodes, spleen,
thymus, kidneys, urinary bladder, testes, epididymides, prostate, seminal vesicles,
mammary glands, ovaries, uterus/cervix, vagina, uterine horn,
thyroid/parathyroid, pituitary, adrenals, brain, spinal cord, skeletal muscle,
sciatic nerve, skin, eyes, exorbital lacrimal glands, and harderian glands.
No treatment-related deaths or abnormal clinical signs were observed.
No effects on body weigh tor weight of other organs were reported in any female
treatment groups or males given 10, 100, or 300 ppm. The study authors reported
a significant reduction in the mean body weight of male rats administered
1,000 ppm on Days 15-92 (see Appendix A, Table A. 1 [in U. S. EPA, 2012d\). The
authors also reported a significant reduction in absolute and relative mean
testicular weights and a significant increase in relative kidney and brain weights
in male rats treated with 1,000-ppm HMPA (see Appendix A, Table A. 2 [in
U.S. EPA, 2012d\ for relative organ weights; the data for absolute organ weights
were not provided in the study). However, the increase in relative brain and
kidney weights was not correlated with any histopathological observations, and
the study authors considered these findings to be spurious.
Comprehensive histopathological examination of the controls and treated
groups identified the respiratory tract and the testis as the only tissues with
treatment-related lesions. The study authors reported a dose-related increase in
the lesion distribution and severity in the nasal passages. In the 10-, 100-, and
300-ppm groups (both males andfemales), nasal lesions (epithelial denudation,
regeneration, and squamous metaplasia) were limited mainly to the anterior
nasal passages, whereas the general architecture of the nasal cavity was occluded
by marked proliferation of the turbinate bone and myxoidfibrous tissue at
1,000 ppm. The study authors rated the severity of the respiratory tract lesions as
20 N,N,N',N"-Tetramethylphosphoramide (TMPA)
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normal, minimal, mild, marked, or severe. Table A. 3 [in U.S. EPA, 2012d]
presents a summary of the severity of the respiratory tract lesions in rats as
presented in the study; the study did not state whether the table is for males,
females, or both. The rats did not have any difficulty in breathing, despite the
severe distortion of the nasal passages at 1,000 ppm. Bilateral testicular atrophy
occurred at 1,000 ppm, and the epididymal tubules contained numerous exfoliated
germ cells with scanty spermatozoa. No further details were reported on the
testicular effects. The study authors identified a NOAEL in drinking water of
10 ppm (1.2 mg/kg-day in males and 2.3 mg/kg-day in females). Based on a
significant increase in severity of nasal lesions, 100 ppm (15 mg/kg-day in males
and 20 mg/kg-day in females) is considered a LOAEL. The other significant
effects noted in this study were reported in males and consisted of reduced body
weight and testes weights (absolute and relative), increased relative brain and
kidney weights, and testicular atrophy. These effects were all noted at a higher
dose (1,000 ppm, or 123 mg/kg-day in males).
The critical effect for the 90-day rat study of HMPA was increased severity of nasal
lesions at 100 ppm (15 mg/kg-day in males and 20 mg/kg-day in females). The male NOAEL of
1.2 mg/kg-day was used as the POD in deriving p-RfDs for HMPA (U.S. HP A. 2012b) and is
adopted herein as the analogue POD for TMPA.
In the current assessment, the NOAEL of 1.2 mg/kg-day was converted to a human
equivalent dose (HED) according to current U.S. EPA (2011c) guidance. In Recommended Use
of Body Weight3/4 as the Default Method in Derivation of the Oral Reference Dose (U.S. EPA.
2011c). the Agency endorses body-weight scaling to the 3/4 power (i.e., BW3'4) as a default to
extrapolate toxicologically equivalent doses of orally administered agents from all laboratory
animals to humans for the purpose of deriving an oral reference dose (RfD). This approach is
appropriate for the nasal lesions observed in the study by Keller et al. (1997) as cited in U.S.
EPA (2012b) because they occurred following drinking water exposure and are systemic in
origin (attributed to metabolism of HMPA in the liver and nasal tissues after absorption from the
gut and distribution throughout the body).
Following U.S. EPA (2011c) guidance, the POD for nasal lesions in rats is converted to
an HED through the application of a dosimetric adjustment factor (DAF) derived as follows:
DAF = (BWa1 4 - BWh1 4)
where
DAF = dosimetric adjustment factor
BWa = animal body weight
BWh = human body weight
Using an average strain-specific reference BWa of 0.235 kg for male rats and a reference
BWh of 70 kg for humans, the resulting DAF is 0.24 (U.S. EPA, 2011c). Applying this DAF to
the NOAEL of 1.2 mg/kg-day yields a POD (HED) as follows:
21 N,N,N',N"-Tetramethylphosphoramide (TMPA)
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POD (HED) = NOAEL (mg/kg-day) x DAF
= 1.2 mg/kg-day x 0.24
= 0.29 mg/kg-day
The subchronic p-RfD for TMPA is derived using an interspecies uncertainty factor
(UFa) of 3 because cross-species dosimetric adjustment to a POD (HED) was performed, as
described above. Additionally, for the derivation of the screening subchronic p-RfD for TMPA,
a database uncertainty factor (UFd) of 10 was applied to account for the absence of toxicity
information for TMPA, and an intraspecies uncertainty factor (UFh) of 10 was applied to account
for human-to-human variability in the absence of information to assess the toxicokinetics and
toxicodynamics of TMPA in humans. Thus, the screening subchronic p-RfD for TMPA is
derived using the analogue POD (HED) of 0.29 mg/kg-day and a composite uncertainty factor
(UFc) of 300 (reflecting a UFa of 3, a UFd of 10, and a UFh of 10):
Screening Subchronic p-RfD = Analogue POD (HED) UFc
= 0.29 mg/kg-day ^ 300
= 1 x 10"3 mg/kg-day
Table A-4 summarizes the uncertainty factors for the screening subchronic p-RfD for
TMPA.
Table A-4. Uncertainty Factors for the Screening Subchronic p-RfD for
TMPA (CASRN 16853-36-4)
UF
Value
Justification
UFa
3
A UFa of 3 (10°5) is applied to account for uncertainty in characterizing the toxicokinetic or
toxicodynamic differences between rats and humans following TMPA exposure. The toxicokinetic
uncertainty has been accounted for by calculating an HED through application of a 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 (U.S. EPA. 20110).
UFd
10
A UFd of 10 is applied to account for the absence of reliable toxicity studies evaluating potential
systemic, reproductive, and developmental effects of TMPA. The analogue HMPA was used to
identify a POD.
UFh
10
A UFh of 10 is applied to account for human-to-human variability in susceptibility in the absence of
quantitative information to assess the toxicokinetics and toxicodynamics of TMPA in humans.
UFl
1
A UFl of 1 is applied because the POD is a NOAEL.
UFS
1
A UFS of 1 is applied because a subchronic study was selected as the principal study for the
subchronic assessment.
UFC
300
Composite UF = UFA x UFD x UFH x UFL x UFS.
HED = human equivalent dose; LOAEL = lowest-observed-adverse-effect level;
NOAEL = no-observed-adverse-effect level; POD = point of departure; p-RfD = provisional reference dose;
TMPA = N,N,N',N"-tetramethylphosphoramide; UF = uncertainty factor; UFA = interspecies uncertainty factor;
UFc = composite uncertainty factor; UFD = database uncertainty factor; UFH = intraspecies uncertainty factor;
UFl = LOAEL-to-NOAEL uncertainty factor; UFS = subchronic-to-chronic uncertainty factor.
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Derivation of a Screening Chronic Provisional Reference Dose
The POD (HED) of 0.29 mg/kg-day identified for nasal lesions in the 90-day drinking
water study provided the most sensitive and reliable endpoint in the entire database and was,
therefore, selected for the derivation of chronic reference values for HMPA. This analogue POD
has been similarly adopted for deriving the screening chronic p-RfD for TMPA. The same
uncertainty factors used for the screening subchronic p-RfD (UFa of 3, UFd of 10, and UFh of
10) were applied with an additional subchronic-to-chronic uncertainty factor (UFs) of 10 to
account for extrapolation from a subchronic to a chronic duration. Thus, the screening chronic
p-RfD for TMPA was derived using a UFc of 3,000.
Screening Chronic p-RfD = Analogue POD (HED) UFc
= 0.29 mg/kg-day ^ 3,000
= 1 x 10"4 mg/kg-day
Table A-5 summarizes the uncertainty factors for the screening chronic p-RfD for
TMPA.
Table A-5. Uncertainty Factors for the Screening Chronic p-RfD for
TMPA (CASRN 16853-36-4)
UF
Value
Justification
UFa
3
A UFa of 3 (10°5) is applied to account for uncertainty in characterizing the toxicokinetic or
toxicodynamic differences between rats and humans following PMPA exposure. The toxicokinetic
uncertainty has been accounted for by calculating an HED through application of a D AF 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. 20110).
UFd
10
A UFd of 10 is applied to account for the absence of reliable toxicity studies evaluating potential
systemic, reproductive, and developmental effects of TMPA. The analogue HMPA was used to
identify a POD.
UFh
10
A UFh of 10 is applied to account for human-to-human variability in susceptibility in the absence of
quantitative information to assess the toxicokinetics and toxicodynamics of TMPA in humans.
UFl
1
A UFl of 1 is applied because the POD is a NOAEL.
UFS
10
A UFS of 10 is applied because a subchronic study was selected as the principal study for the chronic
assessment.
UFC
3,000
Composite UF = UFA x UFD x UFH x UFL x UFS.
HED = human equivalent dose; LOAEL = lowest-observed-adverse-effect level;
NOAEL = no-observed-adverse-effect level; POD = point of departure; p-RfD = provisional reference dose;
TMPA = N,N,N',N"-tetramethylphosphoramide; UF = uncertainty factor; UFA = interspecies uncertainty factor;
UFc = composite uncertainty factor; UFD = database uncertainty factor; UFH = intraspecies uncertainty factor;
UFl = LOAEL-to-NOAEL uncertainty factor; UFS = subchronic-to-chronic uncertainty factor.
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APPENDIX B. BACKGROUND AND METHODOLOGY FOR THE SCREENING
EVALUATION OF POTENTIAL CARCINOGENICITY
For reasons noted in the main Provisional Peer-Reviewed Toxicity Value (PPRTV)
document, there is inadequate information to assess the carcinogenic potential of
N,N,N',N"-tetramethylphosphoramide (TMPA). However, information is available for this
chemical which, although insufficient to support a weight-of-evidence (WOE) descriptor and
derivation of provisional cancer risk estimates under current guidelines, may be of use to risk
assessors. In such cases, the Center for Public Health and Environmental Assessment (CPHEA)
summarizes available information in an appendix and develops a "screening evaluation of
potential carcinogenicity." Appendices receive the same level of internal and external scientific
peer review as the provisional cancer assessments in PPRTVs to ensure their appropriateness
within the limitations detailed in the document. Users of the information regarding potential
carcinogenicity in this appendix should understand that there could be more uncertainty
associated with this evaluation than for the cancer WOE descriptors presented in the body of the
assessment. Questions or concerns about the appropriate use of the screening evaluation of
potential carcinogenicity should be directed to the CPHEA.
The screening evaluation of potential carcinogenicity includes the general steps shown in
Figure B-l. The methods for Steps 1-8 apply to any target chemical and they are described in
this Appendix. Chemical (TMPA)-specific data for all steps in this process are summarized in
Appendix C.
Use automated tools
to identify an initial
list of structural
analogues with
genotoxicity and/or
carcinogenicity data
Apply expert
judgment to refine
the list of analogues
(based on
physiochemical
properties, ADME,
and mechanisms of
toxicity)
Summarize cancer
data and MOA
information for
analogues.
Use computational
tools to identify
common structural
alerts and SAR
predictions for
genotoxicity and/or
carcinogenicity
Compare
experimental
genotoxicity data (if
any) for the target
and analogue
compounds
Summarize ADME
data from targeted
literature searches.
Identify metabolites
likely related to
genotoxic and/or
carcinogenic alerts
J
Integrate evidence
streams
Assign qualitative
level of concern for
carcinogenicity based
on evidence
integration (potential
concern or
inadequate
information)
Figure B-l. Steps Used in the Screening Evaluation of Potential Carcinogenicity
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STEP 1. USE OF AUTOMATED TOOLS TO IDENTIFY STRUCTURAL ANALOGUES
WITH CARCINOGENICITY AND/OR GENOTOXICITY DATA
ChemACE Clustering
The U.S. EPA's Chemical Assessment Clustering Engine [ChemACE; U.S. EPA
(2011a)1 is an automated tool that groups (or clusters) a user-defined list of chemicals based on
chemical structure fragments. The methodology used to develop ChemACE was derived from
U.S. EPA's Analog Identification Methodology (AIM) tool, which identifies structural analogues
for a chemical based on common structural fragments. ChemACE uses the AIM structural
fragment recognition approach for analogue identification and applies advanced queries and
user-defined rules to create the chemical clusters. The ChemACE cluster outputs are available in
several formats and layouts (i.e., Microsoft Excel, Adobe PDF) to allow rapid evaluation of
structures, properties, mechanisms, and other parameters which are customizable based on an
individual user's needs. ChemACE clustering has been successfully used with chemical
inventories for identifying trends within a series of structurally similar chemicals, demonstrating
structural diversity in a chemical inventory, and detecting structural analogues to fill data gaps
and/or perform read-across.
For this project, ChemACE is used to identify potential structural analogues of the target
compound that have available carcinogenicity assessments and/or carcinogenicity data. An
overview of the ChemACE process in shown in Figure B-2.
Create and curate an
inventory of chemicals
with carcinogenicity
assessments and/or
cancer data
*
Cluster the target
compound with the
chemical inventory
using ChemACE
*
Identify structural
analogues for the
target compound from
specific ChemACE
clusters
Figure B-2. Overview of ChemACE Process
The chemical inventory was populated with chemicals that have available carcinogenicity
assessments and/or carcinogenicity data from the following databases and lists:
• Carcinogenic Potency Database [CPDB; CPDB (2011)1
• Agents classified as Group 1 or 2 carcinogens by the International Agency for Research
on Cancer (IARC) monographs (I ARC. 2016)
• National Toxicology Program (NTP) Report on Carcinogens [ROC; NTP (2016)1
• NTP technical reports (NTP, 2017)
• Integrated Risk Information System (IRIS) carcinogens (U.S. EPA 2017)
• California EPA Prop 65 list (CalEPA. 2018a)
• European Chemicals Agency (ECHA) carcinogenicity data available in the Organisation
for Economic Co-operation and Development (OECD) Quantitative Structure-Activity
Relationship (QSAR) Toolbox (OECD. 2018)
• PPRTVs for Superfund (U.S. EPA. 2019)
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In total, 2,123 distinct substances were identified from the sources above. For ChemACE
clustering, each individual substance needed to meet the following criteria:
1) Substance is not a polymer, metal, inorganic, or complex salt because ChemACE is not
designed to accommodate these substances;
2) Substance has a CASRN or unambiguous chemical identification; and
3) Substance has a unique Simplified Molecular Input Line Entry System (SMILES)
notation (encoded molecular structure format used in ChemACE) that can be identified
from one of these sources:
a. SRC and DSSTox lists of known SMILES associated with unique CASRNs (the
combined lists contained >200,000 SMILES) or
b. ChemlDplus, U.S. EPA CompTox Chemicals Dashboard, or internet searches.
Of the initial list of 2,123 substances, 201 were removed because they did not meet one
of the first two criteria, and 155 were removed because they did not meet the third. The final
inventory of substances contained 1,767 unique compounds.
Two separate ChemACE approaches were compared for clustering of the chemical
inventory. The restrictive clustering approach, in which all compounds in a cluster contain the
same fragments and no different fragments, resulted in 208 clusters. The less restrictive
approach included the following rules for remapping the chemical inventory:
• treat adjacent halogens as equivalent, allowing fluorine (F) to be substituted for chlorine
(CI), CI for bromine (Br), Br for iodine (I);
• allow methyl, methylene, and methane to be equivalent;
• allow primary, secondary, and tertiary amines to be equivalent; and
• exclude aromatic thiols (removes thiols from consideration).
Clustering using the less restrictive approach (Pass 2) resulted in 284 clusters.
ChemACE results for clustering of the target chemical (TMPA) within the clusters of the
chemical inventory are described in Appendix C.
Analogue Searches in the OECD QSAR Toolbox (DICE Method)
The OECD QSAR Toolbox (Version 4.1) is used to search for additional structural
analogues of the target compound. There are several structural similarity score equations
available in the Toolbox (DICE, Tanimoto, Kulczynski-2, Ochiai/Cosine and Yule). DICE is
considered the default equation. The specific options that are selected for performing this search
include a comparison of molecular features (atom-centered fragments) and atom characteristics
(atom type, count hydrogens attached and hybridization). Chemicals identified in these
similarity searches are selected if their similarity scores exceed 50%.
The OECD QSAR Toolbox Profiler is used to identify those structural analogues from
the DICE search that have carcinogenicity and/or genotoxicity data. Nine databases in the
OECD QSAR Toolbox (Version 4.1) provide data for carcinogenicity or genotoxicity
(see Table B-l).
Analogue search results for the target chemical are described in Appendix C.
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Table B-l. Databases Providing Carcinogenicity and Genotoxicity Data in the OECD
QSAR Toolbox (Version 4.1)
Database Name
Toolbox Database Description3
CPDB
The CPDB provides access to bioassay literature with qualitative and quantitative analysis
of published experiments from the general literature (through 2001) and from the NTP
(through 2004). Reported results include bioassays in rats, mice, hamsters, dogs, and
nonhuman primates. A calculated carcinogenic potency (TD5o) is provided to standardize
quantitative measures for comparison across chemicals. The CPDB contains
1,531 chemicals and 3,501 data points.
ISSCAN
The ISSCAN database provides information on carcinogenicity bioassays in rats and mice
reported in sources including NTP, CPDB, CCRIS, and IARC. This database reports a
carcinogenicity TD5o. The ISSCAN database includes 1,149 chemicals and 4,518 data
points.
ECHA CHEM
The ECHA CHEM database provides information on chemicals manufactured or imported
in Europe from registration dossiers submitted by companies to ECHA to comply with the
REACH regulation framework. The ECHA database includes 9,229 chemicals with
almost 430,000 data points for a variety of endpoints including carcinogenicity and
genotoxicity. ECHA does not verify the information provided by the submitters.
ECVAM Genotoxicity
and Carcinogenicity
The ECVAM Genotoxicity and Carcinogenicity database provides genotoxicity and
carcinogenicity data for Ames positive chemicals in a harmonized format. ECVAM
contains in vitro and in vivo bacteria mutagenicity, carcinogenicity, CA, CA/aneuploidy,
DNA damage, DNA damage and repair, mammalian culture cell mutagenicity, and rodent
gene mutation data for 744 chemicals and 9,186 data points.
Cell Transformation
Assay ISSCTA
ISSCTA provides results of four types of in vitro cell transformation assays including
Syrian hamster embryo cells, mouse BALB/c 3T3, mouse C3H/10T1/2, and mouse Bhas
42 assays that inform nongenotoxic carcinogenicity. ISSCTA consists of 352 chemicals
and 760 data points.
Bacterial mutagenicity
ISSSTY
The ISSSTY database provides data on in vitro Salmonella typhimurium Ames test
mutagenicity (positive and negative) taken from the CCRIS database in TOXNET. The
ISSSTY database provides data for 7,367 chemicals and 41,634 data points.
Genotoxicity OASIS
The Genotoxicity OASIS database provides experimental results for mutagenicity from
"Ames tests (with and without metabolic activation), in vitro CAs and MN, and MLA
evaluated in vivo and in vitro, respectively." The Genotoxicity OASIS database consists
of 7,920 chemicals with 29,940 data points from 7 sources.
Micronucleus OASIS
The Micronucleus OASIS database provides experimental results for in vivo bone marrow
and peripheral blood MNT CA studies in blood erythrocytes, bone marrow cells, and
polychromatic erythrocytes of humans, mice, rabbits, and rats for 557 chemicals.
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Table B-l. Databases Providing Carcinogenicity and Genotoxicity Data in the OECD
QSAR Toolbox (Version 4.1)
Database Name
Toolbox Database Description3
Micronucleus ISSMIC
The ISSMIC database provides data on the results of in vivo MN mutagenicity assay to
detect CAs in bone marrow cells, peripheral blood cells, and splenocytes in mice and rats.
Sources include TOXNET, NTP, and the Leadscope FDA CRADA Toxicity Database.
The ISSMIC database includes data for 563 chemicals and 1,022 data points.
'Descriptions were obtained from the OECD QSAR Toolbox documentation (Version 4.1) (OECD. 20181.
CA = chromosomal aberration; CCRIS = Chemical Carcinogenesis Research Information System;
CPDB = Carcinogenic Potency Database; CRADA = Cooperative Research and Development Agreement;
DNA = deoxyribonucleic acid; ECHA = European Chemicals Agency; ECVAM = European Centre for the
Validation of Alternative Methods; FDA = Food and Drug Administration; IARC = International Agency for
Research on Cancer; ISSCAN = Istituto Superiore di Sanita Chemical Carcinogen Database; ISSCTA = Istituto
Superiore di Sanita Cell Transformation Assay Database; ISSMIC = Istituto Superiore di Sanita Micronucleus
Database; ISSSTY = Istituto Superiore di Sanita Salmonella Typhimurium Database; MLA = mouse lymphoma
gene mutation assay; MN = micronuclei; MNT = micronucleus test; NTP = National Toxicology Program;
OECD = Organisation for Economic Co-operation and Development; QSAR = Quantitative Structure-Activity
Relationship; REACH = Registration, Evaluation, Authorisation and Restriction of Chemicals; TD5o = median
toxic dose.
STEPS 2-5. ANALOGUE REFINEMENT AND SUMMARY OF EXPERIMENTAL
DATA FOR GENOTOXICITY, TOXICOKINETICS, CARCINOGENICITY, AND
MODE OF ACTION
The outcome of the Step 1 analogue identification process using ChemACE and the
OECD QSAR Toolbox is an initial list of structural analogues with genotoxicity and/or
carcinogenicity data. Expert judgment is applied in Step 2 to refine the list of analogues based
on physiochemical properties, absorption, distribution, metabolism, and excretion (ADME), and
mechanisms of toxicity. The analogue refinement process is chemical-specific and is described
in detail for TMPA in Appendix C. Steps 3, 4, and 5 (summary of experimental data for
genotoxicity, toxicokinetics, carcinogenicity, and mode of action [MOA]) are also chemical
specific (see Appendix C for further details).
STEP 6. STRUCTURAL ALERTS AND STRUCTURE-ACTIVITY RELATIONSHIP
PREDICTIONS
Structural alerts and predictions for genotoxicity and carcinogenicity are identified using
six freely available structure-based tools (described in Table B-2).
28 N,N,N',N"-Tetramethylphosphoramide (TMPA)
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Table B-2. Tools Used to Identify Structural Alerts and Predict
Carcinogenicity and Genotoxicity
Name
Description3
OECD QSAR
Toolbox
(Version 4.1)
Seven OECD QSAR Toolbox profiling methods were used, including:
• Carcinogenicity (genotox and nongenotox) alerts by ISS (Version 2.3); updated version
of the module originally implemented in Toxtree. It is a decision tree for estimating
carcinogenicity, based on 55 structural alerts (35 from the Toxtree module and 20 newly
derived).
• DNA alerts for AMES by OASIS (Version 1.4); based on the Ames mutagenicity
TIMES model, uses 85 structural alerts responsible for interaction of chemicals with
DNA.
• DNA alerts for CA and MNT by OASIS (Version 1.1); based on the DNA reactivity of
the CAs TIMES model, uses 85 structural alerts for interaction of chemicals with DNA.
• In vitro mutagenicity (Ames test) alerts by ISS (Version 2.3); based on the Mutagenicity
module in Toxtree. It is a decision tree for estimating in vitro (Ames test) mutagenicity,
based on a list of 43 structural alerts relevant to the investigation of chemical
genotoxicity via DNA adduct formation.
• In vivo mutagenicity (MN) alerts by ISS (Version 2.3); based on the ToxMic rulebase in
Toxtree. The rulebase has 35 structural alerts for in vivo MN assay in rodents.
• OncoLogic™ Primary Classification (Version 4.0); "developed by LMC and OECD to
mimic the structural criteria of chemical classes of potential carcinogens covered by the
U.S. EPA's OncoLogic Cancer Expert System for Predicting the Carcinogenicity
Potential" for categorization purposes only, not for predicting carcinogenicity. It is
applicable to organic chemicals with at least one of the 48 alerts specified.
• Protein binding alerts for chromosomal aberrations by OASIS (Version 1.3); based on
33 structural alerts for interactions with specific proteins including topoisomerases,
cellular protein adducts, etc.
OncoLogic
(Version 7)
OncoLogic is a tool for predicting the potential carcinogenicity of chemicals based on the
application of rules for SAR analysis, developed by experts. Results may range from "Low"
to "High" concern level.
Tox Alerts
ToxAlerts is a platform for screening chemical compounds against structural alerts, developed
as an extension to the Online Chemical Monitorinu Environment (OCHEM: httos://ochem.eu)
system. Only "approved alerts" were selected, which corresponds to a moderator approving
the submitted data. A list of the ToxAlerts found for the chemicals screened in the
preliminary batch is below:
• Genotoxic carcinogenicity, mutagenicity
o Aliphatic halide (general)
o Aliphatic halide (specific)
o Aliphatic halogens
o Aromatic amine (general)
o Aromatic amine (specific)
o Aromatic amines
o Aromatic and aliphatic substituted primary alkyl halides
o Aromatic nitro (general)
o Aromatic nitro (specific)
o Aromatic nitro groups
o Nitroarenes
o Nitro-aromatic
o Primary and secondary aromatic amines
o Primary aromatic amine, hydroxyl amine, and its derived esters or amine generating
group
• Nongenotoxic carcinogenicity
o Aliphatic halogens
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Table B-2. Tools Used to Identify Structural Alerts and Predict
Carcinogenicity and Genotoxicity
Name
Description3
ToxRead
(Version 0.9)
ToxRead is a tool designed to assist in making read-across evaluations reproducible.
Structural alerts for mutagenicity are extracted from similar molecules with available
experimental data in its database. Five similar compounds were selected for this project. The
rule sets included:
• Benigni/Bossa as available in Toxtree (Version 1)
• SARpy rules extracted by Politecnico di Milano, with the automatic tool SARpy
• IRFMN rules extracted by human experts at Istituto di Ricerche Farmacologiche Mario
Negri
• CRS4 rules extracted by CRS4 Institute with automatic tools
Toxtree
(Version 2.6.13)
Toxtree estimates toxic hazard by applying a decision tree approach. Chemicals were queried
in Toxtree using the Benigni/Bossa rulebase for mutagenicity and carcinogenicity. If a
potential carcinogenic alert based on any QSAR model or if any structural alert for genotoxic
and nongenotoxic carcinogenicity was reported, then the prediction was recorded as a positive
carcinogenicity prediction for the test chemical. The output definitions from the tool manual
are listed below:
• SA for genotoxic carcinogenicity (recognizes the presence of one of more SAs and
specifies a genotoxic mechanism)
• SA for nongenotoxic carcinogenicity (recognizes the presence of one or more SAs, and
specifies a nongenotoxic mechanism)
• Potential Salmonella typhimurium TA100 mutagen based on QSAR
• Unlikely to be a S. typhimurium TA100 mutagen based on QSAR
• Potential carcinogen based on QSAR (assigned according to the output of QSAR8
aromatic amines)
• Unlikely to be a carcinogen based on QSAR (assigned according to the output of
QSAR8 aromatic amines)
• Negative for genotoxic carcinogenicity (no alert for genotoxic carcinogenicity)
• Negative for nongenotoxic carcinogenicity (no alert for nongenotoxic carcinogenicity)
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Table B-2. Tools Used to Identify Structural Alerts and Predict
Carcinogenicity and Genotoxicity
Name
Description3
VEGA
VEGA applies several QSARs to a given chemical, as described below:
• Mutagenicity (Ames test) CONSENSUS model: a consensus assessment is performed
based on predictions of the VEGA mutagenicity models (CAESAR, SARpy, ISS, and
&-NN)
• Mutagenicity (Ames test) model (CAESAR): integrates 2 models: 1 is a trained SVM
classifier, and the other is for FNs removal based on structural alerts matching
• Mutagenicity (Ames test) model (SARpy/IRFMN): rule-based approach with 112 rules
for mutagenicity and 93 for nonmutagenicity, extracted with SARpy software from the
original training set from the CAESAR model; includes rules for both mutagenicity and
non-mutagenicity
• Mutagenicity (Ames test) model (ISS): rule-based approach based on the work of
Benigni and Bossa (ISS) as implemented in the software Toxtree (Version 2.6)
• Mutagenicity (Ames test) model (A'-NN/read-across): performs a read-across and
provides a qualitative prediction of mutagenicity on S. typhimurium (Ames test)
• Carcinogenicity model (CAESAR): Counter Propagation Artificial neural network
developed using data for carcinogenicity in rats extracted from the CPDB database
• Carcinogenicity model (ISS): built implementing the same alerts Benigni and Bossa
(ISS) implemented in the software Toxtree (Version 2.6)
• Carcinogenicity model (IRFMN/ANTARES): a set of rules (127 structural alerts),
extracted with the SARpy software from a data set of 1,543 chemicals obtained from the
carcinogenicity database of EU-funded project ANT ARES
• Carcinogenicity model (IRFMN/ISSCAN-CGX): based on a set of rules (43 structural
alerts) extracted with the SARpy software from a data set of 986 compounds; the data set
of carcinogenicity of different species was provided bv Kirkland et al. (2005)
aThere is some overlap between the tools. For example, OncoLogic classification is provided by the QSAR
Toolbox but the prediction is available only through OncoLogic, and alerts or decision trees were used or adapted
in several models (e.g., Benigni and Bossa alerts and Toxtree decision tree) (OECD. 20181.
ANT ARES = Alternative Non-testing Methods Assessed for REACH Substances; CA = chromosomal aberration;
CAESAR = Computer Assisted Evaluation of Industrial Chemical Substances According to Regulations;
CPDB = Carcinogenic Potency Database; DNA = deoxyribonucleic acid; EU = European Union; FN = false
negative; IRFMN = Istituto di Ricerche Farmacologiche Mario Negri; ISS = Istituto Superiore di Sanita;
ISSCAN-CGX = Istituto Superiore di Sanita Chemical Carcinogen; &-NN = ^-nearest neighbor;
MN = micronucleus; MNT = micronucleus test; OCHEM = Online Chemical Monitoring Environment;
OECD = Organisation for Economic Co-operation and Development; QSAR = quantitative structure-activity
relationship; SA = structural alert; SAR = structure-activity relationship; SVM = support vector machine;
TIMES = The Integrated MARKEL-EFOM System.
The tool results for the target (TMPA) and analogue compounds are provided in
Appendix C.
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APPENDIX C. RESULTS OF THE SCREENING EVALUATION OF POTENTIAL
CARCINOGENICITY
STEP 1. USE OF AUTOMATED TOOLS TO IDENTIFY STRUCTURAL ANALOGUES
WITH CARCINOGENICITY AND/OR GENOTOXICITY DATA
ChemACE clustering was performed as described in Appendix B. The cluster containing
N,N,N',N"-tetramethylphosphoramide (TMPA; less restrictive approach; Cluster 16) also
contained pentamethylphosphoramide (PMPA), hexamethylphosphoramide (HMPA), and no
other compounds. All of the cluster members have chemical structures that contain: (1) a
phosphorus-oxygen double bond, (2) the phosphorus also has three single bonds to nitrogen, and
(3) the nitrogen is a secondary or tertiary amide (see Figure C-l).
The Organisation for Economic Co-operation and Development (OECD) Quantitative
Structure-Activity Relationship (QSAR) Toolbox Profiler was used to identify structural
analogues from the DICE analogue search that have carcinogenicity and/or genotoxicity data
(see Step 1 methods in Appendix B). The OECD QSAR Toolbox Profiler did not identify any
structural analogues of TMPA having carcinogenicity or genotoxicity data (including HMPA).
TMPA and HMPA have a similarity score of only 40% because HMPA has three -N(CH3)2
fragments and TMPA only has only one (see Figure C-2).
O Secondary amide
O Tertiary amide
O Nitrogen-phosphorus bond
O Phosphorus-oxygen double bond
Figure C-l. Illustration of Common Fragments in Cluster 16 for TMPA
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HMPA TMPA
Figure C-2. Illustration of Common Atom-Centered Fragments in the DICE Search
STEP 2. ANALOGUE REFINEMENT USING EXPERT JUDGMENT
Expert chemistry judgment (informed by toxicological expertise) was applied to evaluate
whether HMPA is a viable structural analogue for TMPA. TMPA is an TV-methylated
phosphoric acid triamide with four methyl groups located at the N, N, N', and N" positions.
Experimental toxicokinetic data for HMPA confirm that its predominant metabolic pathway is
oxidative demethylation, which results in sequential removal of one methyl group at each of the
amide positions. Thus, HMPA is converted to PMPA, then proceeds to TMPA, and finally to
N,N',N"-trimethylphosphoramide (triMPA) (see "Toxicokinetics" section and Appendix A for
further discussion). Each demethylation step produces formaldehyde, a known human
carcinogen (NTP. 2016). Demethylation by cytochrome P450 (CYP450) has been demonstrated
in liver, lung, and nasal cavity tissues, suggesting that carcinogenicity may occur following both
inhalation and oral exposure (see Appendix A). HMPA is an appropriate analogue for assessing
the carcinogenicity of TMPA because it is a metabolic precursor to the target chemical, and
because both compounds are expected to generate the same carcinogenic metabolite,
formaldehyde.
STEP 3. COMPARISON OF THE EXPERIMENTAL GENOTOXICITY DATA FOR
TMPA AND HMPA
TMPA (unknown isomer) did not induce mitotic recombination in the Drosophila
melanogaster white/white+ (w/w+) eye mosaic system |"Vogel and Nivard (1993); see "Other
Data" section in the main body of this PPRTV assessment]. No additional genotoxicity data
were available for TMPA.
U.S. EPA (2012b) reported that HMPA was generally negative in bacterial mutagenicity
studies using Salmonella typhimurium and Escherichia coli (with and without metabolic
activation), but did produce somatic mutation, mitotic recombination, and sex-linked recessive
lethal mutations in I), melanogaster and mitotic gene conversion in Saccharomyces cerevisiae
(with metabolic activation). The effects in Drosophila were suggested to fit a pattern seen with
crosslinking agents (Bogdanffy et al.. 1997). HMPA produced deoxyribonucleic acid
(DNA)-protein crosslinks in rat nasal epithelial cells following inhalation exposure (Kuvkendall
et al.. 1995).
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HMPA was mutagenic in mouse lymphoma P388F and L5178Y cells (with metabolic
activation), but was not mutagenic in Chinese hamster ovary (CHO) or V79 cells (with or
without metabolic activation) (U.S. EPA. 2012b). HMPA did not induce chromosome
aberrations (CAs) in human lymphocytes or rat liver RLi cells (not tested with metabolic
activation). Both negative and positive findings (with metabolic activation only) were reported
in sister chromatid exchange (SCE) assays in CHO cells. Micronucleus (MN) frequency was
increased in human HepG2 cells, but not in human lymphocytes tested without metabolic
activation (IARC. 1999).
Genotoxicity studies of HMPA in vivo showed positive findings of CAs in rat peripheral
lymphocytes (but not mouse bone marrow), increased MN frequency in rat and mouse bone
marrow, and SCE in mouse bone marrow (but not mouse liver) (U.S. EPA. 2012b; I ARC. 1999).
Mixed findings were reported in the mouse dominant lethal assay (one positive study,
one negative study).
STEP 4. TOXICOKINETICS OF TMPA AND HMPA
The toxicokinetics of TMPA and HMPA are described and compared in the "Metabolic
Analogues" section of Appendix A. Experimental data show that formaldehyde is released
during sequential oxidative demethylation of HMPA to form PMPA, which is further
metabolized to TMPA and triMPA.
STEP 5. CARCINOGENICITY OF HMPA AND MODE-OF-ACTION DISCUSSION
Under the Guidelines for Carcinogen Risk Assessment (U.S. EPA. 2005). there is
"Suggestive Evidence of Carcinogenic Potential" for HMPA by the inhalation route of exposure
(U.S. EPA, 2012b). Increased incidences of squamous cell carcinomas of the rat nasal cavity
were observed in both sexes (nasal tumors were observed at 400 ppb after 7 months and at
50 ppb after 12 months) (Lee and Trochimowicz. 1982b). The data from this study are not
sufficient to support a quantitative cancer dose-response assessment because of uncertainties in
the experimental design, in establishing clear dose-response data (which requires a number of
assumptions to be made), and in data reporting (U.S. EPA. 2012b).
The HMPA mode of action (MOA) for cancer is not fully established. However, studies
indicate a role for metabolism of HMPA likely through CYP450-mediated A'-demethylation to
formaldehyde, which produces tumors in experimental animals following both inhalation and
oral exposure (U.S. EPA. 2012b). Whereas, a direct contribution of parent (HMPA) and the
subsequently demethylated phosphoramide products (PMPA and TMPA) to HMPA's
carcinogenic MOA cannot be ruled out due to a lack of relevant information, the
metabolically-induced intracellular release of formaldehyde plays a significant role in HMPA's
carcinogenic and mutagenic effects (IARC. 1999; Bogdanffv et at.. 1997). This role is supported
by studies describing a requirement for metabolism in the production of HMPA-induced
DNA-protein crosslinks (Kuvkendall et at.. 1995). Additionally, data concerning the
genotoxicity of PMPA (an expected demethylated methylphosphoramide product) also supports
the proposed role for formaldehyde in promoting genotoxicity, because the use of a
formaldehyde trapping agent abrogated the mutagenic potential of PMPA after metabolic
activation in S. typhimurium (Sarrit* et at.. 1997). The putative MOAs described above would
also be relevant for TMPA.
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STEP 6. STRUCTURAL ALERTS AND SAR PREDICTIONS FOR TMPA AND HMPA
Structural alerts (SAs) and predictions for genotoxicity and carcinogenicity were
identified using computational tools as described in Appendix B. The tool results for TMPA and
the analogue compound HMPA are shown in Table C-l.
Table C-l. Heat Map Illustrating the Structural Alert and SAR Prediction Results for
TMPA (CASRN 16853-36-4) and HMPA (CASRN 680-31-9)a
TMPA
HMPA
Tool
Model
VEGA
Mutagenicity (Ames test) CONSENSUS model—assessment
Mutagenicity (Ames test) model (CAESAR)—assessment
Mutagenicity (Ames test) model (SARpy/IRFMN)—assessment
Mutagenicity (Ames test) model (ISS)—assessment
Mutagenicity (Ames test) model (&-NN/read-across)—assessment
Carcinogenicity model (CAESAR)—assessment
Carcinogenicity model (ISS)—assessment
Carcinogenicity model (IRFMN/ANT ARES)—assessment
Carcinogenicity model (IRFMN/ISSCAN-CGX)—assessment
Toxtree
Negative for genotoxic carcinogenicity
Negative for nongenotoxic carcinogenicity
OncoLogic
OncoLogic (prediction of the carcinogenic potential of the chemical)
Model results or alerts indicating no concern for carcinogenicity/mutagenicity.
Model results outside the applicability domain for carcinogenicity/mutagenicity.
Model results or alerts indicating concern for carcinogenicity/mutagenicity.
All iools> and models described in Appendix B were used. Models with results are presented in the heat map
(models without results were omitted).
ANT ARES = Alternative Nontesting Methods Assessed for REACH Substances; CAESAR = Computer Assisted
Evaluation of Industrial Chemical Substances According to Regulations; CONSENSUS = Consensus Assessment
based on multiple models (CAESAR, SARpy, ISS, and /i-NN): DNA = deoxyribonucleic acid;
HMPA = hexamethylphosphoramide; IRFMN = Istituto di Ricerche Farmacologiche Mario Negri; ISS = Istituto
Superiore di Sanita; ISSCAN-CGX = Istituto Superiore di Sanita Chemical Carcinogen; &-NN = ^-nearest
neighbor; SAR = structure-activity relationship; TMPA = N,N,N',N"-tetramethylphosphoramide;
QSAR = quantitative structure-analysis relationship.
The QSAR models in the VEGA tool indicate no concern for mutagenicity of HMPA in
the Ames test, which is consistent with experimental data (see Step 3 above). Similarly, VEGA
models also indicated no concern for Ames test mutagenicity for TMPA (no experimental data).
While the results of QSAR-driven mutagenicity analysis suggest differing mutagenic capacity of
HMPA and TMPA, the production of mutagenic metabolic products (e.g., formaldehyde) cannot
be ruled out and may still result in mutagenic activity associated with TMPA/HMPA exposure.
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Carcinogenicity models were inconsistent for different tools (i.e., positive in VEGA for HMPA
and TMPA, negative for genotoxic and nongenotoxic carcinogenicity for both compounds in
Toxtree). However, prediction results for HMPA and TMPA were generally consistent when
data were available for both compounds. The OncoLogic results for HMPA indicate a structural
alert for "Organophosphorous Type Compounds"; however, the designated level of concern from
this tool is based on experimental evidence of HMPA carcinogenicity following inhalation
exposure. The OncoLogic output for HMPA states: "The final level of carcinogenicity concern
for this compound is 'high-moderate,' if the exposure is by inhalation; otherwise the level of
concern is 'marginal.'" OncoLogic did not produce a result for TMPA.
STEP 7. EVIDENCE INTEGRATION FOR SCREENING EVALUATION OF TMPA
CARCINOGENICITY
Table C-2 presents the data for multiple lines of evidence pertinent to the screening
evaluation of the carcinogenic potential of TMPA. HMPA and TMPA are structurally similar
compounds (i.e., both are methylated phosphoric acid triamides) and HMPA is a metabolic
precursor to TMPA. HMPA induced mitotic recombination in Drosophila, and these results fit a
pattern seen with crosslinking agents (Bogdanffv et al.. 1997). HMPA produced DNA-protein
crosslinks in rat nasal epithelial cells following inhalation exposure (Kuvkendall et al.. 1995).
which is consistent with the release of formaldehyde. TMPA metabolism is also expected to
generate formaldehyde, which is the proposed toxic moiety for HMPA carcinogenicity (Jones
and Jackson. 1968). Data from the parent phosphoramide (PMPA) detail that use of a
formaldehyde trapping agent has been demonstrated to inhibit PMPA-driven mutagenicity, and
further supports the requirement for metabolic processing in the ability of PMPA to promote
genotoxic activity (Sarrit*et al.. 1997). TMPA (unknown isomer) did not induce mitotic
recombination in Drosophila; however, TMPA is anticipated to release less formaldehyde than
HMPA during metabolism, which may account for the dissimilar results in this assay. No
additional genotoxicity data were available for TMPA. Bacterial mutagenicity findings were
negative for HMPA. HMPA produced both positive and negative results in the dominant lethal
test and assays evaluating mammalian cell mutagenicity, SCE, MN frequency, and CAs
(see Step 3 for details). Computational tools indicated no concern for Ames test mutagenicity for
TMPA or HMPA. The results from carcinogenicity models were consistent for both HMPA and
TMPA, but were inconsistent for different tools (i.e., positive in VEGA for HMPA and TMPA,
negative for genotoxic and nongenotoxic carcinogenicity for both compounds in Toxtree).
OncoLogic results indicate a structural alert for HMPA based on experimental evidence of
HMPA carcinogenicity following inhalation exposure.
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Table C-2. Integration of Evidence for TMPA (CASRN 16853-36-4) and
HMPA (CASRN 680-31-9)
Evidence Streams
TMPA
HMPA
Analogue selection and
evaluation
(see Steps 1 and 2)
NA
Appropriate analogue; both compounds are
TV-methylated phosphoric acid triamides;
HMPA is metabolic precursor to TMPA
Experimental
genotoxicity data
(see Step 3)
TMPA (unknown isomer) did not induce
mitotic recombination in the Drosophilcf,
no additional studies
Somatic mutation, mitotic recombination, and
sex-linked recessive lethal mutations in
Drosophilcf, mitotic gene conversion in yeast;
DNA-protein crosslinks in rat nasal epithelial
cells following inhalation; increased in vivo
MN frequency; generally negative in bacterial
mutagenicity studies; mixed findings (positive
and negative) for in vitro mammalian cell
mutagenicity, in vitro MN frequency, SCE (in
vitro and in vivo), in vivo CAs, and the mouse
dominant lethal assay
ADME evaluation
(see Step 4)
Formaldehyde released during sequential
oxidative demethylation; HMPA and
PMPA are metabolic precursors; triMPA
is a metabolite
Formaldehyde released during sequential
oxidative demethylation; PMPA, TMPA, and
triMPA are metabolites
Cancer data and MOA
(see Step 5)
ND
Nasal tumors in male and female rats after
chronic inhalation exposure; MOA is not fully
known, but intracellular release of
formaldehyde may be responsible;
DNA-protein crosslink formation following
HMPA inhalation provides support for this
MOA
Common structural
alerts and SAR
predictions
(see Step 6)
No structural alerts; no concern for
mutagenicity (Ames test);
carcinogenicity models were inconsistent
(i.e., positive in VEGA, negative for
genotoxic and nongenotoxic
carcinogenicity in Toxtree)
Structural alert for organophosphorus type
compounds (based on cancer bioassay data); no
concern for mutagenicity (Ames test);
carcinogenicity models were inconsistent
(i.e., positive in VEGA, negative for genotoxic
and nongenotoxic carcinogenicity in Toxtree)
ADME = absorption, distribution, metabolism, and excretion; CA = chromosomal aberration;
DNA = deoxyribonucleic acid; HMPA = hexamethylphosphoramide; MN = micronuclei; MOA = mode of action;
NA = not applicable; ND = no data; PMPA = pentamethylphosphoramide; SAR = structure-activity relationship;
SCE = sister chromatid exchange; TMPA = N,N,N',N"-tetramethylphosphoramide;
triMPA = N,N',N"-trimethylphosphoramide.
STEP 8. QUALITATIVE LEVEL OF CONCERN FOR TMPA POTENTIAL
CARCINOGENICITY
Table C-3 identifies the qualitative level of concern for potential carcinogenicity of
TMPA based on the multiple lines of evidence described above.
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Table C-3. Qualitative Level of Concern for Carcinogenicity of TMPA
(CASRN 16853-36-4)
Level of Concern
Designation
Comments
Concern for potential
carcinogenicity
Selected
HMPA is an appropriate analogue for assessing the
carcinogenicity of TMPA because it is a metabolic
precursor to the target chemical, and because both
compounds generate the same carcinogenic
metabolite, formaldehyde. There is "Suggestive
Evidence of Carcinogenic Potential" for HMPA by the
inhalation route of exposure based on increased
incidences of squamous cell carcinomas of the rat
nasal cavity in both sexes. The intracellular release
of formaldehyde, which has been suggested to be
responsible for HMPA's carcinogenic effects, would
be expected to occur for TMPA as well.
Inadequate information for assigning
qualitative level of concern
NS
NA
HMPA = hexamethylphosphoramide; NA = not applicable; NS = not selected;
TMPA = N,N,N',N"-tetramethylphosphoramide.
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APPENDIX D. REFERENCES
ACGIH (American Conference of Governmental Industrial Hygienists). (2017). 2017 TLVs and
BEIs: Based on the documentation of the threshold limit values for chemical substances
and physical agents & biological exposure indices. Cincinnati, OH.
http://www.acgih.org/forms/store/ProductFormPublic/2017-tlvs-and-beis
AT SDR (Agency for Toxic Substances and Disease Registry). (2017). Minimal risk levels
(MRLs). June 2017. Atlanta, GA: Agency for Toxic Substances and Disease Registry
(ATSDR). Retrieved from http://www.atsdr.cdc.gov/mrls/index.asp
Bogdanffv. MS; Mathison. BH; Kuvkendall. JR; Harman. AE. (1997). Critical factors in
assessing risk from exposure to nasal carcinogens [Review], Mutat Res-Fundam Mol
Mech Mutagen 380: 125-141. http://dx.doi.org/10.1016/S0Q27-5107(97)00131-0
CalEPA (California Environmental Protection Agency). (201 1). Hot spots unit risk and cancer
potency values. Appendix A. Sacramento, CA: Office of Environmental Health Hazard
Assessment.
http://standards.nsf.org/apps/group public download.php'Mocument id 19121
CalEPA (California Environmental Protection Agency). (2016). OEHHA toxicity criteria
database. Sacramento, CA: Office of Environmental Health Hazard Assessment.
Retrieved from http://www.ochha.ca.gov tcdb index.asp
CalEPA (California Environmental Protection Agency). (2018a). Chemicals known to the state
to cause cancer or reproductive toxicity May 25, 2018. (Proposition 65 list). Sacramento,
CA: Office of Enironmental Health Hazard Assessment, http://oehha.ca.gov/proposition-
65 proposi tion-65-list
CalEPA (California Environmental Protection Agency). (2018b). OEHHA chemical database.
Sacramento, CA: Office of Environmental Health Hazard Assessment. Retrieved from
https://oehha.ca.gov/chemicals
ChemlDplus. (2016). N,N,N',N" Tetramethylphosphoramide. CASRN 16853-36-4 [Database],
Bethesda, MD: U.S. Library Library of Medicine. Retrieved from
http://chem.nlm.nih.gov/chemidplus/rn/3732-86-3
ChemlDplus. (2018). ChemlDplus - a TOXNET database: National Institutes of Health, U.S.
Library of Medicine. Retrieved from http s: //chem. nl m. nih. gov/chemi dpi us/
CPDB (Carcinogenic Potency Database). (201 1). The carcinogenic potency project: The
carcinogenic potency database [Database]: Department of Energy; National Cancer
Institute; Environmental Protection Agency; National Institute of Environmental Health
Sciences; National Toxicology Program; University of California, Berkeley. Retrieved
from https://toxnet.nlm.nih.gov/cpdb/
Dahl. AR; B re/in ski. DA. (1985). Inhibition of rabbit nasal and hepatic cytochrome P-450-
dependent hexamethylphosphoramide (HMPA) N-demethylase by methylenedioxyphenyl
compounds. Biochem Pharmacol 34: 63 1-636. http://dx.doi.org/10.1016/00Q6-
2952(85)90256-4
Dahl. AR; Hadlev. WM; Hahn. FF; Benson. JM; McClellan. RO. (1982). Cytochrome P-450-
dependent monooxygenases in olfactory epithelium of dogs: possible role in
tumorigenicity. Science 216: 57-59.
ECHA (European Chemicals Agency). (2018). Registered Substances: Helsinki, Finland.
Retrieved from http://echa.europa.eu/web/guest/information-on-chemicals/registered-
substances
39 N,N,N',N"-Tetramethylphosphoramide (TMPA)
-------
FINAL
September 2020
Gervasi- PG; Longo, V; Naldi. F; Panattoni, G; Ursino. F. (1991). Xenobiotic-metabolizing
enzymes in human respiratory nasal mucosa. Biochem Pharmacol 41: 177-184.
http://dx.doi.org/10.1016/0006-2952(91 )90474-J
IARC (International Agency for Research on Cancer). (1999). I ARC monographs on the
evaluation of carcinogenic risks to humans - Hexamethylphosphoramide [Review], In
Re-evaluation of some organic chemicals, hydrazine and hydrogen peroxide (pp. 1465-
1481). (ISSN 1017-1606). Lyon, France.
http://monographs.iarc.fr/ENG/Monographs/vol71/mono71-101.pdf
IARC (International Agency for Research on Cancer). (2016). I ARC Monographs on the
evaluation of carcinogenic risk to humans. Geneva, Switzerland: International Agency for
Research on Cancer, WHO. Retrieved from
http://monographs.iarc.fr/ENG/Monographs/PDFs/index.php
IARC (International Agency for Research on Cancer). (2018). IARC monographs on the
evaluation of carcinogenic risk to humans.
http://monographs.iarc.fr/ENG/Monographs/PDFs/index.php
IPCS (International Programme on Chemical Safety). (2018). INCHEM: Chemical safety
information from intergovernmental organizations. Geneva, Switzerland: World Health
Organization, Canadian Centre for Occupational Health and Safety. Inter-Organization
Programme for the Sound Management of Chemicals. Retrieved from
http://www.inchem.org/
Jackson. H; Craig. AW. (1966). Antifertility action and metabolism of
hexamethylphosphoramide. Nature 212: 86-87. http://dx.doi.org/10.1038/212086a0
Jackson. H; Jones. AR; Cooper. ERA. (1969). Effects of hexamethylphosphoramide on rat
spermatogenesis and fertility. J Reprod Fertil 20: 263-269.
http://dx.doi.org/10.1530/irf0.0200263
Jones. AR. (1970). Further metabolites of hexamethylphosphoramide. Biochem Pharmacol 19:
603-604. http://dx.doi.org/10.1016/0006-2952(70)90215-7
Jones, AR; Jackson, H. (1968). The metabolism of hexamethylphosphoramide and related
compounds. Biochem Pharmacol 17: 2247-2252. http://dx.doi.org/10.1016/00Q6-
2952(68)90031-2
Keller. DA; Marshall. CE; Lee. KP. (1997). Subchronic nasal toxicity of
hexamethylphosphoramide administered to rats orally for 90 days. Fundam Appl Toxicol
40: 15-29. http://dx.doi.org/10.1006/faat.1997.2375
Kerns, WD; Pavkov, KL; Donofrio, DJ; Grail a. EJ; Swenberg, JA. (1983). Carcinogenicity of
formaldehyde in rats and mice after long-term inhalation exposure. Cancer Res 43: 4382-
4392.
Kimbrough, R; Gaines, TB. (1966). Toxicity of hexamethylphosphoramide in rats. Nature 211:
146-147. http://dx.doi.org/10.1038/211146a0
Kimbrough, R; Gaines. TB. (1973). The chronic toxicity of hexamethylphosphoramide in rats.
Bull Environ Contain Toxicol 10: 225-226. http://dx.doi.org/10.1007/BF01684548
Kirkland. D; Aardema, M; Henderson, L; Miiller, L. (2005). Evaluation of the ability of a battery
of three in vitro genotoxicity tests to discriminate rodent carcinogens and non-
carcinogens: I. Sensitivity, specificity and relative predictivity. Mutat Res 584: 1-256.
http: //dx. doi. or g/10.1016/i. mr gentox .2005.02.004
Kuvkendatt, JR; Trela, BA; Bogdanffv, MS. (1995). DNA-protein crosslink formation in rat
nasal epithelial cells by hexamethylphosphoramide and its correlation with formaldehyde
production. Mutat Res 343: 209-218. http://dx.doi.org/10.1016/0165-1218(95)90016-0
40 N,N,N',N"-Tetramethylphosphoramide (TMPA)
-------
FINAL
September 2020
Lee. KP; Trochimowicz. HI. (1982a). Induction of nasal tumors in rats exposed to
hexamethylphosphoramide by inhalation. J Natl Cancer Inst 68: 157-171.
Lee. KP; Trochimowicz. HI. (1982b). Metaplastic changes of nasal respiratory epithelium in rats
exposed to hexamethylphosphoramide (HMPA) by inhalation. Am J Pathol 106: 8-19.
Lee. KP; Trochimowicz. HI. (1982c). Pulmonary response to inhaled hexamethylphosphoramide
in rats. Toxicol Appl Pharmacol 62: 90-103. http://dx.doi.org/10.1016/0Q41-
008X782)90105-3
Lee. KP; Trochimowicz. HI. (1984). Morphogenesis of nasal tumors in rats exposed to
hexamethylphosphoramide by inhalation. Environ Res 33: 106-118.
http://dx.doi.org/10.1016/0013-9351 (84)90012-4
Liu. ('; Zhuo. X; Gonzalez. FJ; Ding. X. (1996). Baculovirus-mediated expression and
characterization of rat CYP2A3 and human CYP2a6: Role in metabolic activation of
nasal toxicants. Mol Pharmacol 50: 781-788.
I .ongo. V; Citti- L; Gervasi. P. (1988). Biotransformation enzymes in nasal mucosa and liver of
Sprague-Dawley rats. Toxicol Lett 44: 289-297. http://dx.doi.org/10.1016/Q378-
4274(88)90168-3
NIOSH (National Institute for Occupational Safety and Health). (2016). NIOSH pocket guide to
chemical hazards. Index of chemical abstracts service registry numbers (CAS No.).
Atlanta, GA: Center for Disease Control and Prevention, U.S. Department of Health,
Education and Welfare, http://www.cdc.gov/niosh/npg/npgdcas.html
N I P (National Toxicology Program). (2016). 14th Report on carcinogens. Research Triangle
Park, NC: U.S. Department of Health and Human Services, Public Health Service.
https://ntp.niehs.nih.gov/pubhealth/roc/index-l.html
NTP (National Toxicology Program). (2017). NTP Technical reports index. Available online at
https://ntp.niehs.nih.gov/results/summaries/chronicstudies/index.html
OECD (Organisation for Economic Co-operation and Development). (2018). The OECD QSAR
toolbox for grouping chemicals into categories. Retrieved from
https://www.qsartoolbox.org/
OSHA (Occupational Safety & Health Administration). (2017a). Air contaminants: Occupational
safety and health standards for shipyard employment, subpart Z, toxic and hazardous
substances. (OSHA Standard 1915.1000). Washington, DC: U.S. Department of Labor.
https://www.osha.gov/pls/oshaweb/owadisp.show document?p table STANDARDS&p
id10286
OSHA (Occupational Safety & Health Administration). (2017b). Table Z-l: Limits for air
contaminants. Occupational safety and health standards, subpart Z, toxic and hazardous
substances. (OSHA standard 1910.1000, 29 CFR). Washington, DC: U.S. Department of
Labor.
http://www.osha.gov/pls/oshaweb/owadisp.show document?p table STANDARDS&p
id=9992
Sarrit'. AM; Krahn. DF; Donovan. SM; O'Neil. RM. (1997). Evaluation of
hexamethylphosphoramide for gene mutations in Salmonella typhimurium using plate
incorporation, preincubation, and suspension assays. Mutat Res 380: 167-177.
http://dx.doi.org/10.1016/S0027-5107(97)00134-6
Shott. LP; Borkovec. AB; Knapp. WA. Jr. (1971). Toxicology of hexamethylphosphoric
triamide in rats and rabbits. Toxicol Appl Pharmacol 18: 499-506.
http://dx.doi.org/10.1016/S0041 -008 X( 71)80002-9
41 N,N,N',N"-Tetramethylphosphoramide (TMPA)
-------
FINAL
September 2020
Swenberg. JA; Kerns. WD; Mitchell RI; Grail a. EJ; Pavkov. KL. (1980). Induction of squamous
cell carcinomas of the rat nasal cavity by inhalation exposure to formaldehyde vapor.
Cancer Res 40: 3398-3402.
Thornton-Manning. JR; Nikula. KJ; Hotchkiss. JA; Avila. KJ; Rohrbacher. KD; Ding. X; Dahl.
AR. (1997). Nasal cytochrome P450 2A: identification, regional localization, and
metabolic activity toward hexamethylphosphoramide, a known nasal carcinogen. Toxicol
Appl Pharmacol 142: 22-30. http://dx.doi.org/10.1006/taap.1996.7975
U.S. EPA (U.S. Environmental Protection Agency). (2005). Guidelines for carcinogen risk
assessment [EPA Report], (EPA/630/P-03/001B). Washington, DC: U.S. Environmental
Protection Agency, Risk Assessment Forum.
https://www.epa.gov/sites/production/files/2013-
09/documents cancer guidelines final 3-25-05.pdf
U.S. EPA (U.S. Environmental Protection Agency). (201 la). Chemical Assessment Clustering
Engine (ChemACE) [Database], Retrieved from https://www.epa.gov/tsca-screening-
tools/chemical-assessment-clustering-engine-chemace
U.S. EPA (U.S. Environmental Protection Agency). (201 lb). Health effects assessment summary
tables for superfund (HEAST). Available online at https://epa-heast.ornl.gov/heast.php
(accessed December 13, 2017).
U.S. EPA (U.S. Environmental Protection Agency). (201 lc). Recommended use of body weight
3/4 as the default method in derivation of the oral reference dose (pp. 1-50). (EPA/100/R-
11/0001). Washington, DC: U.S. Environmental Protection Agency, Risk Assessment
Forum, Office of the Science Advisor, https://www.epa.gov/sites/production/files/2013-
09/documents recommended-use-of-bw34.pdf
U.S. EPA (U.S. Environmental Protection Agency). (2012a). 2012 Edition of the drinking water
standards and health advisories [EPA Report], (EPA/822/S-12/001). Washington, DC:
U.S. Environmental Protection Agency, Office of Water.
https://rais.ornl. gov/documents/2012 drinking water.pdf
U.S. HP A (U.S. Environmental Protection Agency). (2012b). Provisional Peer Reviewed
Toxicity Values for hexamethylphosphoramide (CASRN 680-31-9) [EPA Report],
Cincinnati, OH. https://hhpprtv.ornl.gov/issue papers/Hexamethvlphosphoramide.pdf
U.S. HP A (U.S. Environmental Protection Agency). (2017). IRIS Carcinogens: 69502.2(a)( 1 )(E).
Chemicals that are identified as "carcinogenic to humans," "likely to be carcinogenic to
humans," or Group A, Bl, or B2 carcinogens in the United States Environmental
Protection Agencys Integrated Risk Information System. Available online at
https://www.dtsc.ca.gov/SCP/SourceLists.cfm
U.S. HP A (U.S. Environmental Protection Agency). (2018a). Integrated risk information system
(IRIS) [Database], Washington, DC: U.S. Environmental Protection Agency, Integrated
Risk Information System. Retrieved from http://www.epa.gov/iris/
U.S. HP A (U.S. Environmental Protection Agency). (2018b). The Toxic Substances Control
Act's public inventory (TSCA inventory)- Retrieved from https://www.epa.gov/tsca-
inventorv/how-access-tsca-inventorv#download
U.S. HP A (U.S. Environmental Protection Agency). (2019). Provisional peer-re vie wed toxicity
values for superfund (PPRTV): Derivation support documents. Washington, DC.
Retrieved from https://www.epa.gov/pprtv
Vogel. EW; Nivard. MJ. (1993). Performance of 181 chemicals in a drosophila assay
predominantly monitoring interchromosomal mitotic recombination. Mutagenesis 8: 57-
81. http://dx.doi.org/10.1093/mutage/8.1.57
42 N,N,N',N"-Tetramethylphosphoramide (TMPA)
-------
FINAL
September 2020
Wang. NC; Zhao. QJ; Wesselkamper. SC; Lambert. JC; Petersen. D; Hess-Wilson. JK. (2012).
Application of computational toxicological approaches in human health risk assessment.
I. A tiered surrogate approach. Regul Toxicol Pharmacol 63: 10-19.
http://dx.doi.Org/10.1016/i.vrtph.2012.02.006
Yang. YS; Jung. MS; Cho. CK; Suh. SH; Gu. BH. (2011). Composition for producing printed
circuit board and printed circuit board using the same. (Patent Application #12/328,458).
Yang, YS; Jung, MS; Cho, CK; Suh, SH; Gu, BH. http://patft.uspto.gov/netacgi/nph-
Parser?d=PALL&p=l&u=%2Fnetahtml%2FPTO%2Fsrchnum.htm&r=l&f=G&l=5Q&sl
=7943 856.PN.&OS=PN/7943856&RS=PN/7943 856
43 N,N,N',N"-Tetramethylphosphoramide (TMPA)
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