U.S. Department of Health and Human Services

National Institute of Environmental Health Sciences 

National Institutes of Health

National Center for Toxicological Research

Food and Drug Administration

National Institute for Occupational Safety and Health

Centers for Disease Control and Prevention

September 2012

Department of Health and Human Services

National Toxicology Program

NIH Publication No. 12-5971

Letter from the NIEHS/NTP Director.....................................................................................................................................1

NTP Highlights ............................................................................................................................................................................... 2

Overview of the NTP .....................................................................................................................................................................3

Resources and Planning..............................................................................................................................................................7

Advisory Boards and Committees.........................................................................................................................................8

NIOSH/NTP ..................................................................................................................................................................................... 15

NCTR/NTP ..................................................................................................................................................................................... 19

NIEHS/NTP ..................................................................................................................................................................................... 26

Highlighted Activities ...................................................................................................................................................... 26

 Division of National Toxicology Program (DNTP) ........................................................................................... 26

 NIEHS Strategic Planning ......................................................................................................................................... 27

 Gulf Oil Spill Response ............................................................................................................................................... 27

 Workshop:Advancing Research on Mixtures:

 New Perspectives and Approaches for Predicting Adverse Human Health Effects ......................... 28

 Chemical Effects in Biological Systems ............................................................................................................... 29

 Use of NTP Products by Other Agencies ............................................................................................................ 30

Report on Carcinogens.................................................................................................................................................... 32

Office of Health Assessment and Translation...................................................................................................... 35

Interagency Center for the Evaluation of Alternative Toxicological Methods................................... 36

NTP Research and Testing Program .................................................................................................................................. 40

 Nomination, Selection, Evaluation, and Review ....................................................................................................... 40

Chronic Toxicity/Carcinogenicity Studies ................................................................................................................... 44

 General Toxicology Studies .............................................................................................................................................. 50

 Mutagenesis and Genetic Toxicity ................................................................................................................................. 54

Organ System Toxicity ........................................................................................................................................................ 55

Immunotoxicity ...................................................................................................................................................................... 57

 Disposition, Metabolism, and Toxicokinetic Studies .............................................................................................. 59

 Genetic and Alternative Test Systems .......................................................................................................................... 62

Cellular and Molecular Pathology ..................................................................................................................................... 63

NTP Archives .................................................................................................................................................................................. 66

NTP Laboratory ............................................................................................................................................................................ 66

NTP Postdoctoral Training Programs................................................................................................................................ 67

Interagency Agreements

NIEHS/NCTR Interagency Agreement .......................................................................................................................... 68

NIEHS/NIOSH Interagency Agreement ........................................................................................................................ 70

NIEHS/EPA Interagency Agreement ............................................................................................................................. 73

NTP/NHGRI/EPA/FDA Interagency Memorandum of Understanding on High Throughput

Screening, Toxicity Pathway Profiling, and Biological Interpretation of Findings .................................. 75

Appendix 1

 Agency Staff and Contact Information ......................................................................................................................... 82

Appendix 2

NTP FY 2011 Bibliography ................................................................................................................................................. 85
2M4N 2-methoxy-4-nitroaniline

3TC lamivudine

ACB allele-specific competitive blocker 

ACD allergic contact dermatitis

ADME absorption, distribution, metabolism, 

and excretion

Ag silver

ATSDR Agency for Toxic Substances and 

Disease Registry

AZT zidovudine, 3’-Azido-3’-

Deoxythymidine

BPA bisphenol A

BSC Board of Scientific Counselors

BALF bronchoalveolar lavage fluid

CASRN Chemical Abstracts Service Registry

Number

CDC Centers for Disease Control and 

Prevention

CDMA code division multiple access

CEBS Chemical Effects in Biological Systems 

CERHR Center for the Evaluation of Risks 

to Human Reproduction

CNS central nervous system

CNT carbon nanotube

CPSC U.S. Consumer Product 

Safey Commission

CYP cytochrome P450

DCA dichloroaniline

DEHP di(2-ethylhexyl) phthalate

DGE differential gene expression

DHHS Department of Health 

and Human Services

DILI drug-induced liver injury

DNMT DNA methyltransferases

DRPM direct reading, personal monitors

DTBBA dithiobisbenzanilide

DO diversity outbred

ECVAM European Centre for the Validation 

of Alternative Methods

EDTA ethylenediaminetetraacetate

EGEHE ethylene glycol 2-ethylhexyl ether

ENU N-ethyl-N’-nitrosourea

EPA U.S. Environmental Protection Agency

ESR electron spin resonance

FDA U.S. Food and Drug Administration

FFPE formalin fixed, paraffin embedded

FY fiscal year

GABA gamma-aminobutyric acid

GC/MS gas chromatography/mass

spectroscopy

GD gestational day

GEMM genetically engineered mouse model

GMM genetically modified model

GSM global system for mobile

communication

HHE Health Hazard Evaluations

HPLC high performance liquid

chromatography

HPRT hypoxanthine-guanine 

phosphoribosyltransferase

HTS high throughput screening

IAG interagency agreement

IARC International Agency for 

Research on Cancer

ICATM International Cooperation on

Alternative Test Methods

ICD irritant contact dermatitis

ICCVAM Interagency Coordinating 

Committee on the Validation 

of Alternative Methods

IgE immunoglobulin E

IRIS EPA’s Integrated Risk 

Information System

JaCVAM Japanese Center for the Validation 

of Alternative Methods

KoCVAM Korean Center for the Validation 

of Alternative Methods 

LC/MS/MS liquid chromatography/tandem 

mass spectrometry

LCM laser capture microdissection

LLNA Local Lymph Node Assay

LOH loss of heterozygosity

mAb monoclonal antibody
MiRNA microRNA

MLA Mouse Lymphoma Assay

Mn manganese

MOC Memorandum of Cooperation

MOU Memorandum of Understanding

MRI magnetic resonance imaging 

MS mass spectrometry

N/A not applicable

NASH nonalcoholic steatohepatitis

NCGC NIH Chemical Genomics Center

NCI National Cancer Institute

NCTR National Center for 

Toxicological Research

NICEATM NTP Interagency Center for the 

Evaluation of Alternative Toxicological 

Methods

NHGRI National Human Genome Research

Institute

NIEHS National Institute of Environmental 

Health Sciences

NIH National Institutes of Health

NIOSH National Institute for Occupational

Safety and Health

NMDA N-methyl-D-aspartic acid

NMR nuclear magnetic resonance

NNLA NTP Nonneoplastic Lesion Atlas

NTP National Toxicology Program

NVP Nevirapine

OECD Organisation for Economic 

Co-operation and Development

OEHHA Office of Environmental Health

Hazard Assessment 

OPA ortho-phthalaldehyde

ORA Office of Regulatory Affairs

OSHA Occupational Safety and Health

Administration

PAH polycyclic aromatic hydrocarbon

PBPK physiologically based 

pharmacokinetic

PBPK/PD PBPK/pharmacodynamic

PCB polychlorinated biphenyl

PCBTF chloro-4-(trifluoromethyl) benzene

PCR polymerase chain reaction

PET Positron Emission Tomography

PFAA perfluoralkyl acid

PFOA perfluorooctanoic acid

PIG-A phosphatidylinositol glycan – 

complementation group A 

PND postnatal day

PPAR peroxisome proliferator-activated

receptor

PWG pathology working group

qHTS quantitative high throughput screens

qNPA quantitative nuclease 

protection assay

qPCR quantitative polymerase 

chain reaction.

QSDAR quantitative spectrometric 

data-activity relationship

RACB reproductive assessment 

by continuous breeding

RBC/RET red blood cell/reticulocyte 

RFR radiofrequency radiation 

RoC Report on Carcinogens

ROS reactive oxygen species

RP retinyl palmitate

SACATM Scientific Advisory Committee on

Alternative Toxicological Methods

SD Sprague Dawley

SOT Society of Toxicology

SSL simulated solar light

STL Special Techniques Laboratory

STP Society of Toxicologic Pathologists

TCAB tetrachloroazobenzene

TiO2 titanium dioxide

TK toxicokinetics

TOX NTP Toxicity Report

TR NTP Technical Report

UDP uridine diphosphate

UGT UDP glucuronosyltransferases

UV ultraviolet

U.S. United States

VOC volatile organic compound

NTP Annual Report 2011 1

In 2011, Health and Human Services (HHS) Secretary Sebelius and Congress

approved reorganization of the National Institute of Environmental Health

Sciences (NIEHS), creating the Division of the National Toxicology Program (DNTP)

within NIEHS. This reorganization enhances NTP’s visibility and recognizes its 

unique mission and goals within the National Institutes of Health and the unique

capabilities and talents of its staff. The DNTP retained four previous branches 

and established the NTP Laboratory where staff will carry out important targeted 

in-house research. In addition, the NTP Center for the Evaluation of Risks to Human 

Reproduction (CERHR) was transformed into the Office of Health Assessment and 

Translation (OHAT), which will now expand the focus of its evaluations, grounded in reproduction and 

development assessments, to a broader range of human health effects.

The NTP carried out a number of significant, widely attended workshops this past year. OHAT held a timely

workshop on the Role of Environmental Chemicals in the Development of Diabetes and Obesity in January 

2011. Two workshops in the Series on Best Practices for Regulatory Safety Testing were held in January 2011, 

Assessing the Potential for Chemically Induced Eye Injuries and Assessing the Potential for Chemically Induced 

Allergic Contact Dermatitis. In September 2011, the NTP and NIEHS held a joint workshop on Advancing 

Research on Mixtures: New Perspectives and Approaches for Predicting Adverse Human Health Effects.

NTP’s research and testing program continually aimsto provide a safer tomorrow by studying substances

in our environment. Reports on studies of a number of dietary supplements, AIDS therapeutics, skin care

products, and widely used industrial chemicals were disseminated by NTP in FY 2011. These included

publication of nine NTP Technical Reports and two NTP Toxicity Reports, and drafts of 10 reports for 

which we held two public peer review meetings. We were pleased to provide important information 

on substances that pose a cancer risk through publication of the 12th Report on Carcinogens in June. This

edition added eight newly reviewed substances to the report for a total of 240 listings, including some

classes of related chemicals or agents. 

The NTP continued its efforts to address health concerns associated with the Deepwater Horizon oil spill in

the Gulf of Mexico. NTP research, together with that of other federal agencies and academic investigators,

has given us a better understanding of the chemical hazards created during this incident. A primary 

focus for the NTP is to increase our knowledge of the toxicity of components of oil that persist in the 

environment. We will continue to tackle lingering concerns regarding the safety of Gulf seafood, hazards 

to offshore and onshore cleanup workers, and potential long-term health impacts of residual oil in 

the environment.

All of these efforts, by teams of NTP scientists and through collaborations with other NIEHS divisions and 

Federal agency scientists, were successful in translating and communicating important scientific findings 

to advance human health protection. We will continue to proactively address complex new public health

issues as they emerge. 

Linda S. Birnbaum, Ph.D., D.A.B.T., A.T.S.
2 National Toxicology Program
NTP Annual Report 2011 3

Mission and Goals

Currently, the Toxic Substances Control Act Chemical Substance Inventory 

(http://www.epa.gov/oppt/newchems/pubs/invntory.htm), first published 

in 1979, lists more than 80,000 chemicals as being available for sale and 

use in the United States. Approximately 850 active pesticide ingredients 

are formulated into approximately 17,000 pesticide products. An estimated 

500 to 600 new industrial chemicals are introduced annually into U.S. 

commerce. The effects of many of these substances on human health 

are unknown, yet people and our environment may be exposed to them 

during their manufacture, distribution, use, and disposal or as pollutants in 

our air, water, or soil. While relatively few substances are thought to pose 

a significant risk to human health, safeguarding the public depends upon 

identifying the effects of these agents, and of certain naturally occurring 

chemicals, and determining the levels of exposure at which they may 

become potentially hazardous to humans.

The Department of Health, Education, and Welfare (now the Department of Health and Human Services, 

DHHS) established the National Toxicology Program (NTP) in 1978 as a focal point to coordinate toxicology

testing in the Federal government. In carrying out its mission, the NTP has several goals:

1. Coordinate toxicology testing programs within the federal government. 

2. Strengthen the science base in toxicology.

3. Develop and validate improved testing methods.

4. Provide information about potentially toxic chemicals to health, regulatory, and research agencies, 

 scientific and medical communities, and the public.

Organizational Structure and Oversight

Three agencies, the National Institute of Environmental Health Sciences (NIEHS) of the National Institutes of

Health, the National Institute for Occupational Safety and Health (NIOSH) of the Centers for Disease Control and 

Prevention, and the National Center for Toxicological Research (NCTR) of the Food and Drug Administration

(FDA), form the core for the NTP (Figure 1). The NTP is located at the NIEHS, and the Director of the NIEHS serves

as the NTP Director. Questions and inquiries about the NTP can be directed to the NTP Office of Liaison, Policy

and Review (919-541-7539) or CDM@niehs.nih.gov.
4 National Toxicology Program

NTP Management

Dr. Linda Birnbaum, Director of NIEHS and NTP

Agency Program Management

NCTR: Dr. Paul Howard, Associate Director, Office of Scientific Coordination

NIEHS: Dr. John Bucher, Associate Director, NTP 

NIOSH: Dr. Mark Toraason, Senior Fellow, Division of Applied Research and Technology and Dr. Gayle Debord, 

Chief, Biomonitoring and Health Assessment Branch

Staff of the agencies involved with the program and their contact information are provided in Appendix 1.

Addressing Scientific and Regulatory Needs

The NTP is flexible and innovative in its approach toward addressing public health concerns related to exposures 

to chemical and physical agents at home, in the workplace, and in the environment. Over the years since taking

over the National Cancer Institute’s (NCI’s) cancer bioassay program, the NTP has expanded its scope beyond

cancer to include examining the impact of substances on non-cancer outcomes, such as those affecting 

reproduction and development and the immune, respiratory, nervous, cardiovascular, and endocrine systems.

The NTP recognizes that initiatives addressing critical gaps in knowledge needed to evaluate environmental 

toxicants offer the best opportunities for preventing environmentally mediated diseases. Therefore, the NTP’s 

testing of substances continues to evolve to include more mechanism-based toxicology studies that focus on

understanding the modes of action of agents under study. In recent years, the NTP has placed a greater emphasis

on providing human relevance in interpreting and understanding toxicological information created from animal

or in vitro cell models. Thisis important if we are to be at the forefront in research efforts to improve methods to

assess risk that account for the entire sequence of events from initial chemical exposure to ultimate toxicity. 
NTP Annual Report 2011 5

Examples of activities it covers include:

• Increased use of both information on mechanisms of action and scientific judgment in deliberations 

for listings in the Report on Carcinogens

• Increased efforts to examine alternative testing methods that may provide better information than current 

models while using fewer animals or causing less pain or distress, and may provide better data for risk

assessments

• Increased efforts to collect information on (1) a broader variety of both environmental and occupational

exposures, (2) potentially toxic mixtures of compounds, and (3) susceptibilities based on life stages 

(e.g., neonatal, elderly)

Internationally, the NTP rodent bioassay is recognized as the standard for identifying carcinogenic agents; 

however, the NTP continues to work to reduce, refine, and/or replace the use of experimental animals and to

develop and validate alternative testing methods. This effort led to the creation of the NTP Interagency Center 

for the Evaluation of Alternative Toxicological Methods (NICEATM) in 1998. The NTP continues to work with the 

Interagency Coordinating Committee for the Validation of Alternative Methods (ICCVAM) through NICEATM 

in promoting the development, validation, and regulatory acceptance of new and revised alternative 

toxicological methods.

Strengthening existing partnerships and forging new ones are important to achieve NTP goals. Partnerships

with other sister Federal agencies are in place (interagency agreements [IAGs] are presented on page 67). The 

NTP continues to support an effort to evaluate the phototoxicity of various compounds through the NTP Center 

for Phototoxicology at the NCTR. The NTP is also contributing to toxicological assessments of emerging issues, 

such as nanotechnology, radiofrequency radiation emissions from cellular phones, herbal medicines/dietary 

supplements, water disinfection by-products, mold, and phthalates, and will provide this information to 

other agencies.

Regulatory agencies make decisions to protect public health based on scientific information from several 

sources (e.g., toxicology, human studies, and basic research). The NTP plays a critical role in providing needed

scientific data, interpretation, and guidance in the appropriate uses of these data to regulatory agencies as well

as other groups involved in health-related research. The NTP is committed to using the best science available in

setting priorities for future studies and in designing, conducting, and interpreting the findings of those studies.

The American people and government agencies at state and Federal levels rely on the NTP to provide a strong

scientific basis for making credible decisions that will protect public health. The NTP maintains an objective,

science-based approach in dealing with critical issues in toxicology and is recognized by many groups for its 

scientific rigor, objectivity, and open approach in the continuing dialogue on appropriately applying scientific

advances to toxicology research and testing.

Communication and Public Outreach

Maintaining open communications and ensuring dialogue with Federal and state agencies, industry, 

nongovernment groups, academia, and the public are goals of the NTP. NTP advisory groups (see page 8)

provide regular scientific and public peer review and input. NTP conferences and workshops remain a priority

and are designed to bring researchers, regulators, policy makers, and the public together to examine issues and 

achieve consensus on future directions in toxicology and risk assessment.

Distribution of NTP study results, program plans, initiatives, announcements, press advisories, and publications

is accomplished in several ways to communicate as much as possible with the public. Information is routinely
6 National Toxicology Program

distributed to interested parties through Federal Register announcements and the NTP website 

(http://ntp.niehs.nih.gov). The website offers access to information about the program that details and 

highlights ongoing and future initiatives, announcements, NTP centers, NTP publications, and study data. 

The public can subscribe to the NTP Listserv on the website to receive news and updates. Currently the 

Listserv has more than 4000 subscribers. The NTP publishes the quarterly newsletter, NTP Update, which 

can be downloaded from the NTP website. NTP actively participates in the annual Society of Toxicology 

(SOT) meeting. At the 2011 SOT meeting in Washington DC, NTP staff participated in approximately 

100 workshop/symposium/platform sessions, education/information sessions and posters.

The NIEHS/NTP Central Data Management Office oversees distribution (upon request) of specific chemical study 

information and printed NTP documents – NTP study status reports, final and draft copies of NTP Technical 

Report Series, and background documents for substances nominated to the NTP for study. On-line, searchable

access is available for the Report on Carcinogens(RoC) and the NTP Technical, Toxicity, and Genetically Modified 

Models series reports (http://ntp.niehs.nih.gov).

The NTP is interested in and welcomes stakeholder input into its programs and priorities. Nominations, inquiries,

and comments from the public and other interested parties are encouraged at any time. The NTP Office of

Liaison, Policy and Review at the NIEHS under the direction of Dr. Mary S. Wolfe serves as the focal point for 

receiving input to the program and for overseeing the distribution of information about programs, workshops,

initiatives, and other NTP projects. 

NTP Office of Liaison, Policy and Review Central Data Management

NIEHS/NIH NIEHS/NIH 

MD K2-03 MD K2-05 

111 TW Alexander Drive, RTP, NC 27709 111 TW Alexander Drive, RTP, NC 27709 

Phone: (919) 541-7539 Phone: (919) 541-3419

wolfe@niehs.nih.gov cdm@niehs.nih.gov
NTP Annual Report 2011 7

Current and Projected Research Capacity

The NTP relies on voluntary allocations from the program’s three core agencies (NIEHS, NCTR, and NIOSH)

to support its various programs and initiatives. These allocations are specified after yearly appropriations are

determined. As shown in Figure 2, the total NTP budget for FY 2011 was $126 million.

The NTP conducts its research studies mainly through contract laboratories or in-house at the core agencies, but 

also supports IAGs with other Federal agencies. Funds are used to sponsor workshops and conferences and to

produce and distribute printed programmatic materials. In FY 2011, the NIEHS funded 33 contracts and held four

workshops and three expert-panel meetings for the NTP. The NIEHS also funded IAGs with NIOSH, NCTR, EPA, 

and the NIH Chemical Genomics Center (NCGC) and held five scientific advisory meetings. 

The NTP maintains an objective, science-based approach in dealing with critical issues in toxicology and 

continually sets priorities to improve the nation’s ability to evaluate the human health effects caused by

environmental exposures.

In summary, the NTP is a comprehensive research program spanning several agencies committed to providing 

resources to support the NTP’s research and to communicating the knowledge learned to all stakeholders, 

public and private. The NTP’s efforts in testing, research, and assessing health hazards work to obtain the best

scientifically valid data for health regulatory and research agencies to use to make appropriate decisions about 

potential human risks from exposure to environmental toxicants. Toward that end, the NTP is continually evolving

to remain at the cutting edge of scientific research and development and application of technology.
8 National Toxicology Program

As shown in Figure 1 (page 4), the NTP relies on a number of external boards and committees for science and 

policy oversight and peer review. As needed, the program convenes Special Emphasis Panels and Working 

Groups to address specific topics.

NTP Board of Scientific Counselors

The NTP Board of Scientific Counselors (BSC), a federally chartered advisory group, provides scientific 

oversight to the NTP on the scientific merit of its programs and activities. The Secretary of DHHS appoints

members to the BSC. The BSC can consist of up to 35 scientists, primarily from the public and private sectors, 

with scientific expertise relevant to the NTP’s activities. The BSC charter and current roster are available at

http://ntp.niehs.nih.gov/go/164. Dr. Lori White, Designated Federal Officer, manages the BSC. A list of members 

during FY 2011 is provided in Table 1.

The BSC met twice in FY 2011. The first BSC meeting was held on November 30 – December 1, 2010. The BSC 

voted unanimously to approve the concept for a contract, NTP Sperm Count and Vaginal Cytology Evaluation. 

The BSC reviewed the activities of the Biomolecular Screening Branch and prepared a report (available at 

http://ntp.niehs.nih.gov/go/9741). The BSC provided input on four concepts for the NTP testing program:

(a) Exposure Characterization and Reproductive Health of Men Working with Bisphenol A in the United States, 

(b) Cholesterol and Lipid Modulating Agents: Toxicological Approaches to Assessing Complex Mixtures, 

(c) N-butylbenzenesulfonamide, and (d) Selected Flame Retardants – Update. The NIEHS/NTP Director and 

NTP Associate Director provided reports, updating the BSC on NTP progress since the June 2010 meeting.

The second BSC meeting was held

on April 13, 2011. The BSC voted 

unanimously to approve the concept for 

a contract, Potential for Environmental 

and Therapeutic Agents to Induce

Immunotoxicity. The BSC was provided

overviews of several NTP initiatives

including a Modified One-Generation

Reproduction Study Design and New 

Statistical Methods for Analyzing NTP’s 

2-year Cancer Bioassay Data. The BSC 

provided input on three concepts for the 

NTP testing program: (a) Nanomaterials 

Exposure Assessment, (b) Biospecimen 

Repository and Analysis Capabilities to

Support NTP Exposure Assessment Projects, 

and (c) the Center for the Evaluation of

Risks to Human Reproduction Folic Acid

Workshop. The NIEHS/NTP Director and NTP Associate Director provided reports, updating the BSC on NTP 

progress since the November/December 2010 meeting. Additional information about the BSC, including 

minutes from its meetings, is available on the NTP website (http://ntp.niehs.nih.gov/go/164) or from

Dr. White (whiteld@niehs.nih.gov).

BSC members and NTP staff at the December 2011 BSC meeting.
NTP Annual Report 2011 9

Name and Title Affiliation Term Ends

Tracie E. Bunton, DVM, PhD, DACVP 

Toxicology Consultant

Eicarte LLC 

Fairfield, PA

12/27/10

Edward W. Carney, PhD 

Technical Leader

Developmental, Reproductive and General Toxicology

The Dow Chemical Company 

Midland, MI

12/27/10

Russell C. Cattley, VMD, PhD 

Executive Director Pathology

Amgen 

Thousand Oaks, CA

12/27/10

David A. Eastmond, PhD 

(chair beginning 1/11) Professor and Chair 

Department of Cell Biology 

and Neuroscience

University of California

Riverside, CA

12/27/12

Janan T. Eppig, PhD 

Senior Staff Scientist

The Jackson Laboratory

Bar Harbor, ME

04/15/11

Elaine M. Faustman, PhD 

Professor and Director

Institute for Risk Analysis and Risk Communication 

Department of Environmental and Occupational Health Sciences

University of Washington

Seattle, WA

12/27/12

William P. Janzen, Professor 

Division of Medicinal Chemistry and Natural Products; 

Director, Assay Development and Compound Profiling

University of North Carolina

at Chapel Hill

Chapel Hill, NC

12/27/10

Dana Loomis, PhD 

Professor and Chair 

Department of Epidemiology

University of Nebraska 

Medical Center

Omaha, NE

12/27/12

Stephen W. Looney, PhD 

Professor, Department of Biostatistics 

Department of Oral Health and Diagnostic Science

Georgia Health Sciences University 

Augusta, GA

06/30/12

Melissa A. McDiarmid, MD, MPH 

Professor of Epidemiology and Preventive Medicine Director

Occupational Health Program

University of Maryland School of Medicine

Baltimore, MD

06/30/13

Lisa Minor, PhD 

In Vitro Strategies

In Vitro Strategies, LLC 

Flemington, NJ

06/30/13

Richard Miller, DVM, PhD 

Vice President, Safety Assessment

GlaxoSmithKline 

Research Triangle Park, NC 

06/30/13

Mitzi Nagarkatti, PhD 

Professor and Chair 

Department of Pathology, Microbiology and Immunology

University of South Carolina School of Medicine

Columbia, SC

12/27/11

Raymond F. Novak, PhD (chair 1/10 – 12/10)

Director, Institute of Environmental Health Sciences

Wayne State University

Detroit, MI

12/27/10

Ruthann A. Rudel, MS

Senior Scientist 

Toxicology and Environmental Health Risk Assessment

Silent Spring Institute 

Newton, MA

12/27/11

James L. Sherley, MD, PhD

Senior Scientist, Boston Biomedical Research Institute

Boston Biomedical Research Institute 

Watertown, MA

06/30/11

Gina M. Solomon, MD, MPH 

Senior Scientist

Natural Resources Defense Council 

San Francisco, CA

12/27/11

Justin G. Teeguarden, PhD 

Senior Scientist 

Fundamental and Computational Sciences Directorate

Pacific Northwest National Laboratory

Richland, WA

12/27/11

Judith Zelikoff, PhD 

Professor 

Environmental Medicine Director, Community Outreach

New York University School of Medicine 

Tuxedo, NY

12/27/12

Table 1. NTP Board of Scientific Counselors Membership Roster FY 2011
10 National Toxicology Program

The NTP convened two Technical Reports (TRs) Peer Review Panels in FY 2011 at the NIEHS. The meetings were

open to the public with time scheduled for oral public comment. The panels were charged to (1) peer review

the scientific and technical elements of the study and their presentation and (2) determine whether the study’s 

experimental design and conduct supported the NTP’s conclusions regarding the carcinogenic activity of the 

substance tested. The panels consisted of experts in general pathology, carcinogenesis, dermal toxicology, 

gastrointestinal pathology, and environmental toxicology. The panels reviewed the draft TRs for kava kava

extract, retinoic acid/retinyl palmitate, methyl trans-styryl ketone, styrene-acrylonitile trimer, and 

alpha,beta-thujone at the January 26, 2011 meeting and the draft TRs for senna, AIDS therapeutics, 

acrylamide, and a nondecolorized whole leaf extract of Aloe vera at the April 5, 2011 meeting. Dr. White 

served as Designated Federal Official for the January peer review meeting and Ms. Danica Andrews for the 

April peer review meeting.

Additional information about past TR review panelsis available at http://ntp.niehs.nih.gov/go/36144 and 

information about upcoming TR review panels is available at http://ntp.niehs.nih.gov/go/36051.

NTP Technical Reports Peer Review Panels
NTP Annual Report 2011 11

Scientific Advisory Committee on Alternative Toxicological Methods

The Scientific Advisory Committee on Alternative Toxicological Methods (SACATM) is a federally chartered 

advisory committee established on January 9, 2002, in response to the ICCVAM Authorization Act of 2000

(42 U.S.C. 285l-3(d)). SACATM advises ICCVAM, NICEATM, and the Director of the NIEHS regarding statutorily

mandated duties of ICCVAM and activities of NICEATM (see page 36). SACATM provides advice on priorities and 

activities related to the development, validation, scientific review, regulatory acceptance, implementation, and 

national and international harmonization of new, revised, and alternative toxicological test methods. The SACATM

charter and current roster are available at http://ntp.niehs.nih.gov/go/167. A list of members during

FY 2011 is provided in Table 2. SACATM typically meets once a year and members serve rotating terms of up 

to four years. Dr. Lori White, Designated Federal Officer, manages SACATM.

SACATM met once during FY 2011 at the Hilton Arlington, Arlington, Virginia, on June 16-17, 2011. At that meeting,

SACATM voted unanimously to give a high priority to validation efforts for botulinum in vitro assays and a high

priority to further discussions for an in vitro pyrogen test. SACATM also agreed with the conclusions of the Peer

Review Panel Report on the Evaluation of an In Vitro Estrogen Receptor Transcriptional Activation Test Method for 

Endocrine Disruptor Chemical Screening. ICCVAM agency representatives presented updates on the NIH-FDA 

Regulatory Science Initiative and Small Business Innovation Research grants, and reports on the workshops on

Best Practices for Regulatory Safety Testing and Alternative Methods to Reduce, Refine, and Replace the Use of Animals 

in Vaccine Potency Testing: State of the Science and Future Directions. Liaisons from the European Center for the 

Validation of Alternative Methods, the Korean Center for the Validation of Alternative Methods, the Japanese

Center or the Validation of Alternative Methods, and Health Canada presented updates on the activities of 

their groups. Additional information about SACATM, including minutes from its meetings, is available at 

http://ntp.niehs.nih.gov/go/167 or from Dr. Lori White (whiteld@niehs.nih.gov).

Members of SACATM and ICCVAM and NIEHS/NTP staff at the June 2011 SACATM meeting.
12 National Toxicology Program

Name and Title Affiliation Term Ends

Laura Andrews, PhD, DABT 

Vice President 

Pharmacology and Toxicology

Genzyme Corporation 

Framington, MA

06/30/12

Karen K. Brown, PhD 

President 

Pair O’ Docs Enterprises

Parkville, MO

06/30/11

Joy Cavagnaro, PhD, DABT, RAC, ATS, RAPS

President and Founder

Access BIO, L.C.

Boyce,VA

11/30/14

George B. Corcoran, PhD, ATS 

Professor and Chairman

Department of Pharmaceutical Sciences

Wayne State University 

Detroit, MI

06/30/11

Eugene L. Elmore, PhD 

Senior Project Scientist 

Department of Radiation Oncology

University of California

Irvine, CA

11/30/12

Steven R. Hansen, DVM, MS, MBA, DABT, ABVT

Senior Vice President, Animal Health Services

ASPCA Animal Poison Control Center

Urbana, IL

11/30/12

Gwendolyn Y. McCormick, DVM, MS, DACLAM Attending 

Veterinarian and Distinguished Research Fellow, Animal

Resources Department

Boehringer Ingelheim Pharmaceuticals, Inc

Ridgefield, CT

11/30/12

Sharon A. Meyer, PhD 

Associate Professor, Department of Toxicology

The University of Louisiana at Monroe

Monroe, LA

06/30/11

Steven M. Niemi, DVM 

Director, Center for Comparative Medicine

Massachusetts General Hospital 

Charlestown, MA

06/30/13

Ricardo Ochoa, DVM, PhD, ACVP 

President and Principal

Pre-Clinical Safety, Inc. 

Niantic, CT

11/30/14

Michael J. Olson, PhD, ATS 

Director, Occupational Toxicology Corporate Environment,

Health, Safety and Sustainability

GlaxoSmithKline 

Research Triangle Park, NC

06/30/13

Linda A. Toth, DVM, PhD 

Associate Dean for Research and Faculty Affairs Professor,

Department of Pharmacology

Southern Illinois University School of Medicine 

Springfield, IL

06/30/13

Daniel M. Wilson, PhD, DABT

Mammalian Toxicology Consultant

Toxicology and Environmental Research and Consulting

The Dow Chemical Company 

Midland, MI

11/30/14

Gary Wnorowski, MBA, LAT 

President

Eurofins/Product Safety Laboratories

Dayton, NJ

06/30/11

Table 2. Scientific Advisory Committie on Alternative Toxicological Methods Roster FY 2011
NTP Annual Report 2011 13

NTP Executive Committee

The NTP Executive Committee provides programmatic and policy oversight to the NTP Director. The Executive 

Committee meets once or twice a year in closed forum. In FY 2011 the Department of Defense joined the 

Executive Committee. Members of this committee include the heads (or their designees) from the following 

Federal agencies:

• U.S. Agency for Toxic Substances and Disease Registry/National Center for Environmental Health (ATSDR/NCEH)

• U.S. Consumer Product Safety Commission (CPSC)

• U.S. Department of Defense (DOD)

• U.S. Environmental Protection Agency (EPA)

• U.S. Food and Drug Administration (FDA)

• National Cancer Institute (NCI)

• National Institute of Environmental Health Sciences (NIEHS)

• National Institute for Occupational Safety and Health (NIOSH)

• Occupational Safety and Health Administration (OSHA)

To enhance agency interactions, the NTP uses agency Points of Contact (POCs) in lieu of formal committees to

streamline communication and better utilize agency staff. Agency POCs have a dedicated responsibility and time

commitment, are knowledgeable about the NTP mission and programs and their agency’s resources, and allow 

the most relevant agency expertise to be brought to bear on NTP issues.

Interagency Coordinating Committee on the Validation of Alternative Methods

ICCVAM is a permanent interagency committee of the NIEHS under NICEATM. The committee was formally 

established by the ICCVAM Authorization Act of 2000 (42 U.S.C. 285l-3). The purpose of ICCVAM isto establish,

wherever feasible, guidelines, recommendations, and regulations that promote the regulatory acceptance of 

new or revised scientifically valid toxicological tests that protect human and animal health and the environment 

while reducing, refining, or replacing animal tests and ensuring human safety and product effectiveness 

(see http://iccvam.niehs.nih.gov/about/process.htm). Members of this committee include representatives 

from the following Federal agencies:

• U.S. Consumer Product Safety Commission (CPSC)

• U.S. Department of Agriculture (USDA)

• U.S. Department of Defense (DOD)

• U.S. Department of Energy (DOE) 

• U.S. Department of Health and Human Services

 – Centers for Disease Control and Prevention

 • Agency for Toxic Substances and Disease Registry (ATSDR)

 • National Institute for Occupational Safety and Health (NIOSH)

 – Food and Drug Administration (FDA)

 – National Institutes of Health (NIH)

 • National Cancer Institute (NCI)

 • National Institute of Environmental Health Sciences (NIEHS)

 • National Library of Medicine (NLM)

 • Office of the Director

• U.S. Department of the Interior (DOI)

• U.S. Department of Labor

 – Occupational Safety and Health Administration (OSHA)

• U.S. Department of Transportation

• U.S. Environmental Protection Agency (EPA)
14 National Toxicology Program
NTP Annual Report 2011 15

The National Institute for Occupational Safety and 

Health (NIOSH) is the Federal agency responsible for 

conducting research and making recommendations

for the prevention of work-related injury and illness.

The mission of NIOSH is to generate new knowledge 

in the field of occupational safety and health and 

to transfer that knowledge into practice for the 

betterment of workers. To accomplish this mission,

NIOSH conducts scientific research, develops

guidance and authoritative recommendations, 

distributes information, and responds to requests

for workplace Health Hazard Evaluations (HHEs).

NIOSH’s participation in the NTP is consistent with

its mandate to protect workers’ health and safety

under the Occupational Safety and Health Act and 

the Federal Mine Safety and Health Act. Setting 

priorities in occupational toxicological research is

based upon several sources of information that

are developed and maintained by NIOSH. These 

include HHEs, industry-wide studies, gaps in

knowledge identified while developing criteria for 

recommended standards or Criteria Documents,

Current Intelligence Bulletins, hazard reviews or

alerts, other technical reports, and information

profiles on chemical hazards. Toxicological research

on important occupational chemicals is conducted 

in genetic toxicology, carcinogenesis, toxicological 

characterization, chemical disposition, biological

monitoring, reproductive and developmental 

toxicology, dermal toxicology, and exposure

assessment. NIOSH research projects are 

conducted by the:

• Division of Applied Research and 

Technology – Cincinnati, Ohio

• Division of Surveillance, Hazard Evaluations, 

and Field Studies – Cincinnati, Ohio

• Education and Information Division – 

Cincinnati, Ohio

• Health Effects Laboratory Division – 

Morgantown, West Virginia

NIOSH/NTP studies funded by NIOSH voluntary 

contributions are listed in Table 3.

NIOSH/NTP: Division of Surveillance, Hazard Evaluations, 

and Field Studies

NIOSH/NTP: Division of Applied Research and Technology and 

Education and Information Division

NIOSH/NTP: Health Effects Laboratory Division
16 National Toxicology Program

Table 3. NIOSH/NTP Projects FY 2011

NIOSH/NTP Project 

Project Officer

Objective and/or Summary

Reproductive Health To evaluate reproductive health hazards using a health profile consisting of biomarkers for 

Assessment of Male Workers assessing male fecundity.Current efforts will focus on completing the Longitudinal Investigation of

[Schrader] Fertility and the Environment (LIFE) project, a collaborative effort between NIOSH and the NICHD/

NIH. This work includes development of new biomarkers to include in the male reproductive 

health profile.

Immunochemical Biological 

Monitoring for Occupational

Exposure and Disease 

[Striley]

To evaluate industrial chemicals with known acute and chronic toxicities that present a significant 

exposure risk for workers. Biological monitoring can assess exposure by analyzing acute and latent 

metabolites in various biological media. The goal of this project is to develop low-cost, rapid

immunochemical and analytical chemistry biomonitoring methods that will be used to identify

exposures and evaluate potential interventions. This project will identify and develop new multiplex 

immunochemical methods to evaluate biomarkers of occupational illness or subclinical signs of

occupational illness.

Orthophthalaldehyde (OPA) 

Hazard Assessment

[Toraason]

To conduct an assessment of occupational exposures to OPA and to determine if healthcare workers

are experiencing adverse effects associated with exposure. This study will also develop analytical

methods for environmental monitoring of OPA and determine the feasibility of an OPA biomarker.

Because of the absence of published toxicological data on OPA, testing will be conducted in

experimental animals. The toxicological testing will focus on dermal and respiratory irritation and

sensitization. Dose-response data will be obtained for hazard identification risk assessment, which,

along with health assessments, will serve as the basis for establishing exposure limits.

UV Native Fluorescence Based 

Monitor for Workplace Exposures 

[Snawder]

To develop and evaluate a readily adaptable, next generation, direct reading, personal monitors

(DRPMs) for use in measuring worker exposure to a wide variety of chemicals including

naphthalene and components of asphalt fume. The development of DRPMs for volatile and 

semi-volatile workplace chemicals will be helpful in rapidly assessing chemical exposure and 

will result in more realistic occupational exposure assessments and allow for rapid interventions

leading to reduced worker exposures and thus preventing occupational illness and disease.

Analytical Research and To provide for the administrative needs and analytical instrumentation repair and maintenance in 

Development Infrastructure support of Chemical Exposure and Monitoring Branch chemists conducting research on sampling

[Streicher] and analytical methods development for workplace chemicals. New methods needed to assess

chemicals being investigated as part of the NIOSH/NTP exposure assessment interagency 

agreement are developed in this project.

Diacetyl Exposure Assessment

[Streicher]

To develop and evaluate sampling and analytical methods for diacetyl and other flavoring

compounds to enable accurate exposure assessment and evaluation of the effectiveness of control 

technology. Two sampling and analytical methods are being investigated for measurement of

specific flavoring compounds, most notably diacetyl and 2,3-pentanedione in airborne particles 

and bulk powders. Broader gas chromatography-mass spectrometry method(s) will be developed

for a range of compounds present in flavorings.

Chemical Exposure Monitoring

with Indoor Positioning 

[Brown]

To investigate a direct reading exposure method that uses a personal photo ionization detector 

chemical monitor with telemetry and an indoor positioning system to provide remote monitoring

of a worker’s exposure to volatile organic chemicals (VOCs) with position and time. The personal

monitor continuously samples and analyzes the workers breathing zone air for VOCs while

recording their position and time of exposure. Indoor positioning is accomplished using a radio

transmitter attached to the personal monitor and receivers placed in the ceiling corners of the

room. The positioning receivers communicate with each other and a remote laptop using wireless

local area network technology. The remote laptop calculates and visualizes the worker position and

exposure level. Once developed, this technology will be applied to analyze workplace exposures 

to diacetyl.

Exposure Assessment for 

Toxicologically-Important 

Chemicals 

[Estill]

To characterize workplace exposures to (1) welding fumes with emphasis on manganese

(Mn), (2) indium and indium compounds, (3) diacetyl, (4) 2-methoxy-4-nitroaniline, (5)

2’,2’’’-dithiobisbenzanilide, and (6) bisphenol A (BPA). These chemicals have been nominated 

by various groups to the NTP. NIOSH will identify possible candidate industries, labor unions, 

workplaces, and uses and users; determine if there is relevance for occupational health; estimate

number of workers exposed; and perform limited workplace exposure sampling.
NTP Annual Report 2011 17

NIOSH/NTP Project 

Project Officer

Objective and/or Summary

Exposures and Engineering 

Controls in the Flavoring Industry

[Curwin]

To conduct a complete exposure assessment, evaluate potential engineering controls within the

flavoring industry, and create appropriate work practice advice to reduce occupational exposures 

in these industries. Although a dose response curve for various flavoring compounds and associated 

health effects has not been established, improved work practices and engineer control advice can 

minimize occupational exposures. Data from initial research efforts have been very useful 

in preliminary rulemaking efforts for both OSHA and Cal-OSHA.

Immunotoxicological Evaluation

of Occupational Chemicals 

[Anderson]

To identify occupational and environmental chemical hazards and evaluate immune function and

mechanism associated with exposure. The Immunotoxicology and Hazard Identification Lab will

achieve this goal through both individual projects and collaborations. This research will contribute 

to increased identification of immunological hazards encountered in the workplace.Further

evaluation of these compounds will allow for better risk assessment, which will ultimately 

establish occupational exposure limits.

Evaluation of Perfluoralkyl acids 

(PFAAs) Immunotoxicity

[Franko]

To investigate the immunotoxicity of perfluorooctanoic acid (PFOA), no longer used in

manufacturing, but still persistent in the environment, which has been shown in a murine model 

to be both immunosuppressive and to have a potential role in asthma and allergy. Due to the

potential health effects linked to PFOA exposure, replacement perfluoralkyl acids (PFAAs) are now 

being used in the manufacturing process but little is known about what effects these compounds

will have on immune function. This project will evaluate the immunotoxic effects associated with

individual PFAAs that are still in use, and investigate the mechanism mediating the identified

immunological alterations associated with PFOA exposure.

PFOA and PFOS-induced

Oxidative Stress

[Qian]

To investigate whether exposure to PFOA and perfluorooctane sulfonate (PFOS) induces an

oxidative stress response in a mouse model and in human cells. PFOA and PFOS are widely known 

man-made fluorocarbon-based acids, which have been used in various industrial processes in the

aircraft, automotive, chemical, building material, and personal care products industries. They are 

non-biodegradable and persistent in the human body and environment. Surveillance data suggests

that PFOA and PFOS may cause adverse health effects, but no consistent association between

exposure and health effects has been proven.

Airway Fungal Exposure and 

Allergic Sensitization in Mice

[Templeton]

To compare the immunological health effects of lung exposure to fungal spores or to hyphal

fragment preparations. Agriculture, as well as construction and remediation workers, are exposed

to elevated levels of fungi and can experience rhinitis, respiratory allergic symptoms and/or asthma

as a result of their exposure. This project will have two major areas of study (1) determination of the

health effects following aspiration of hyphal fragments from Stachybotrys chartarum and Alternaria

alternata in the absence of intact spores in mice, and (2) comparison of the ability of aspirated

spores or hyphal suspensions from Aspergillus spp, S. chartarum, and A. alternata to exacerbate

respiratory allergy to ovalbumin.

Immune and Inflammatory 

Aspects of Occupational Rhinitis

[Johnson]

To investigate occupational asthma in workers who often exhibit concurrent occupational rhinitis

with the likelihood that rhinitis developed first. Understanding the mechanisms of occupational

rhinitis is an important area of research in occupational safety, health and medicine. A combined

study design utilizing human and animal research will be employed to identify the 

orthologously-conserved pathways and gene networks that characterize the pathobiology

of occupational rhinitis induced by diisocyanates. The outcomes of this research will benefit

occupational safety and health through improved diagnosis and prevention of allergic airways

disease caused by diisocyanates.

Genetics in 

Occupational Diseases

[Yucesoy]

To investigate susceptibility gene variants that contribute to the development and severity of

occupational irritant contact dermatitis (ICD) and asthma using high-density and high throughput

genotyping platforms. Previous and on-going studies showed that cytokine polymorphisms have

a major influence on silicosis, dementia, accelerated decline in lung function and vaccine efficacy.

Understanding the genetic contribution to the development, progression and outcomes of

complex occupational diseases will help improve the accuracy of risk assessment and improve 

safe exposure levels for genetically susceptible groups in the workforce.
18 National Toxicology Program

NIOSH/NTP Project 

Project Officer

Objective and/or Summary

Genetic Fingerprint of 

Mouse Lung Cancer

[Reynolds]

To determine if there are different carcinogen-specific chromosomal (genetic) markers in

spontaneously-occurring and chemically-induced mouse lung adenocarcinomas using in vitro

and in vivo animal models. Mice were exposed by inhalation to vanadium pentoxide, nickel oxide, 

or cumene (a benzene derivative). Workers in the construction and manufacturing sectors are 

exposed to these compounds. NIOSH is also planning to analyze mouse lung tumors induced

by single-wall carbon nanotubes. If these experiments are successful the plan is to extend these

findings to tumors from occupationally-exposed human populations. Results from these studies 

will be used to establish biomarkers for early detection and therapeutic intervention of lung cancer

in worker populations.

Workplace Exposure, 

Inflammation, and 

Cardiovascular Toxicity

[Erdely]

To investigate the role of ultrafine/nanoparticle-induced cardiopulmonary inflammation and

to identify specific markers of lung inflammation that directly correlate to systemic effects. This

laboratory-based research will evaluate novel molecular mechanisms involved in the link between

occupational exposures to ultrafine/nanosize particulates and the development of cardiovascular

diseases. Planned studies will help to identify potential risk factors, biomarkers, and specific targets

for prevention and therapeutic intervention of occupational-related cardiovascular diseases.

Cutaneous Bioactivation of

Xenobiotics: Hapten vs. 

Pro-hapten

[Siegel]

To develop an in vivo model of allergic skin sensitization that can discriminate between chemicals

requiring metabolic activation for sensitization (pro-haptens) and those that can sensitize without

biological activation (pro-haptens). The model will involve the dermal application of various

pharmacological inhibitors of the cytochrome P450 (CYP) pathway prior to performing either the

local lymph node assay (LLNA) or/and mouse ear swelling test. Selective inhibition of the 

CYP pathway should distinguish between direct-acting haptens and metabolically-activated 

pro-haptens. Validation of the models will be done using known direct acting haptens and 

pro-haptens. Successful development of these models will produce data that strengthen 

in silico hazard predictive models and allows for substitution or modification of allergenic

chemicals and drugs.

Identification of 

Occupational Allergens

[Beezhold]

To identify exposures to substances that can cause inflammatory or immune reactions in certain

work environments. These exposures are important causes of occupational lung diseases such

as asthma and allergic alveolitis. This project is intended to address these concerns through the

development of improved techniques for the detection of such immune reactions before adverse

clinical outcomes occur, and through the development of improved techniques for the detection

and identification of inciting occupational agents. The project will involve the analyses of clinical

samples, environmental bulk samples, and environmental aerosol samples. Successful completion

of these investigations should lead to the development of effective prevention strategies for 

occupational allergies and asthma.

NIOSH project to collect personal breathing 

zone and dermal samples to assess worker

exposures to paving asphalt emissions

NIOSH researcher processing worker

blood and urine samples at a field site for 

biomonitoring studies

NIOSH researcher collecting a surface wipe

sample for possible antineoplastic drug

contamination 
NTP Annual Report 2011 19

The National Center for Toxicological Research (NCTR), the FDA’s research center, plays a critical role in carrying

out the agency’s mission. NCTR, in partnership with researchers from elsewhere in FDA, other government

agencies, academia, and industry, provides innovative technology, methods development, vital scientific training,

and technical expertise. The unique scientific expertise of NCTR is critical in supporting FDA product centers and 

their regulatory roles. NCTR conducts an array of studies that reflect the NTP mission statement. These NCTR/NTP

studies, funded by NCTR voluntary allocations, are listed in Table 4.

NCTR/NTP Project 

Project Officer

Objective and/or Project Summary

Physiologically-based 

Pharmacokinetic (PBPK) 

Models for Bisphenol A 

[Fisher]

(1) To create PBPK models for BPA in three species of adult, neonatal, and pregnant (mother and 

fetus) and lactating (mother and neonate) laboratory animals (mouse, rat, and rhesus monkey);

(2) to use PBPK models to calculate internal measures of dose for both aglycone (i.e., active) and 

conjugated (i.e., inactive) forms of BPA; (3) to create PBPK models for BPA exposure in humans

(adult, child, and pregnant mother and fetus, and lactating mother and infant); (4) to extrapolate 

the internal doses of BPA associated with toxicity in laboratory animals to humans using the human 

suite of PBPK models; and (5) to extrapolate dosimetry from regions of observation to low levels 

of exposure to BPA for which no experimental data exist.

Relationship between Liver 

Epigenetic Phenotype and 

Susceptibility to Nonalcoholic 

Steatohepatitis (NASH)-Induced

Hepatocarcinogenesis in Mice

[Pogribny]

(1) To determine the role of epigenetic dysregulation in the etiology and pathogenesis of dietary 

NASH-induced hepatocarcinogenesis in mice, (2) to determine whether or not interstrain-specific 

susceptibility of mice to NASH-induced hepatocarcinogenesis is associated with differences in

individual hepatic epigenetic phenotypes, (3) to determine the role of epigenetic dysregulation 

in the etiology and pathogenesis of NASH-induced hepatocarcinogenesis in mice induced by

tamoxifen administration, and (4) to determine whether or not aberrant epigenetic markers 

can be used as targets for prevention of NASH-induced hepatocarcinogenesis in mice.

Table 4. NCTR/NTP Projects in FY 2011

NCTR/NTP
20 National Toxicology Program

NCTR/NTP Project 

Project Officer

Objective and/or Project Summary

Development of a 

Physiologically-based 

Pharmacokinetic/

Pharmacodynamic (PBPK/PD) 

Model for Acrylamide 

[Fisher]

(1) To develop a PBPK/PD model for acrylamide and glycidamide, (2) to determine mutagenicity of

acrylamide and its metabolite glycidamide in Big Blue® rats, and (3) to determine the DNA adduct 

levels and the extent of mutagenicity of furan and its metabolite cis-2-buten-4-dial in neonatal

B6C3F1/Tk +/-

 mice.

Global and Locus-Specific

DNA Hypomethylation: A 

Common Mechanism Involved

in Genotoxic and Non-Genotoxic 

Rat Hepatocarcinogenesis 

[Pogribny]

(1) To determine if the temporal alterations in genomic methylation profile in preneoplastic liver 

tissue observed in the folate/methyl-deficient model of rat endogenous hepatocarcinogenesis 

also occur in other carcinogenesis model; (2) to identify genes that are consistently up-regulated

or down-regulated in target tissue during the promotion stage of carcinogenesis; and (3) to

evaluate whether or not the global and locus-specific DNA hypomethylation, along with aberrant

expression of related genes and changes in chromatin conformation is specific only to target

tissues and may be used for early detection of chemicals with carcinogenic potential.

Further Evaluation of the Types 

of Genetic Events Detected by

the Mouse Lymphoma Assay 

(MLA) and the Role of the Assay 

in Mechanistically Based 

Risk Assessment

[Moore]

(1) To determine if the L5178Y/TK+/- MLA adequately detects both aneuploidy and mitotic 

recombination, (2) to determine if the L5178Y mouse lymphoma cells have active recombinase 

functions which lead to a large proportion of mutants that result from recombinase-mediated

rearrangements, and (3) to determine the fundamental genetic mechanism(s) causing the small 

and large colony thymidine kinase mutant phenotypes.

Effect of p53 Genotype on

Gene-Expression Profiles in Mice

Exposed to the Model Mutagen,

N-Ethyl-N’-Nitrosourea (ENU) 

[Morris]

(1) To determine the effect of mutation in the p53 tumor suppressor gene on gene-expression

profiles in young and aged mice, and (2) Determine the effect of mutation in p53 tumor-suppressor 

gene on gene-expression profiles in young and aged mice exposed to the model mutagen ENU.

Development of Cancer-relevant

Biomarkers for Identification of

Potential Carcinogens: Research

to Understand the Normal

Background Frequencies in Rats

[McKinzie]

To understand the distribution and range of spontaneous oncogene mutant frequencies in the 

major organs of rats and mice.

Assessment of Interindividual 

Variability in Expression of DNA 

Methyltransferases (DNMTs),

DNMT3a, and DNMT3b, in

Liver and Identification of

Factors Influencing Expressions 

[Hammons]

(1) To determine levels of expression of DNMT3a and DNMT3b in liver samples from a pool of 

donors selected according to smoking status, gender, and age; (2) to determine the effect

of tobacco smoke on DNMT1, 3a, and 3b expression in liver cell systems; and (3) to assess the 

polymorphism frequency identified in DNMT3b in the sample pool included in the study and 

assess whether it is correlated with expression.

Assessment of Ketamine in the 

Developing Nonhuman Primate 

[Wang]

(1) To determine, using neurohistochemical approaches, if, and at what developmental 

stages, ketamine exposure increases neuronal apoptosis/proliferation; (2) to determine, 

using neurohistochemical approaches, the dose-response for ketamine to produce apoptosis at

the most sensitive developmental stage; (3) to determine the reversibility or permanence of the 

response using behavioral, imaging, and neurohistochemical approaches; and (4) to determine, 

at the most sensitive stage and dose, genomic and proteomic responses to ketamine treatment.

Phosphatidylinositol Glycan – 

Complementation Group 

A (PIG-A) Mutagenesis:

Development of Methods for 

the Identification and Molecular 

Characterization of Mutations 

in the PIG-A Gene in Human 

Lymphoblastoid Cells and 

C57Bl/6 Mice 

[Dobrovolsky]

(1) To develop flow-cytometric methods for the detection of cells with mutations in the PIG-A gene

using wild-type and mutant human lymphoblastoid cells, TK6, and WTK1, as a model, and (2) to

develop flow cytometric methods for the detection of hematopoietic cells with mutations in the 

PIG-A gene in C57Bl/6 mice.
NTP Annual Report 2011 21

NCTR/NTP Project 

Project Officer

Objective and/or Project Summary

PIG-A Mutagenesis:

Development of Methods for 

the Identification and Molecular 

Characterization of Mutations in

the PIG-A Gene of F344 Rats

[Heflich]

(1) To develop methods for measuring PIG-A mutant frequencies in lymphocytes cultured 

from rats; (2) to develop flow cytometric methods for measuring PIG-A mutant/variant

frequencies in both the lymphocytes and red blood cells of rats; (3) to treat rats with either ENU 

or 7,12-Dimethylbenz(a)anthracene and measure PIG-A mutant manifestation in lymphoctyes 

and compare with PIG-A variant manifestation in red blood cells and hypoxanthine-guanine 

phosphoribosyltransferase (HPRT) mutant manifestation in lymphocytes; and (4) to determine 

the spectra of the PIG-A and HPRT mutations induced in the mutant lymphocytes detected in

Objectives 1, 2, and 3 

Phytoestrogens and Aging: 

Dose, Timing, and Tissue 

[Doerge]

(1) To evaluate the potential benefits or detrimental effects of dietary phytoestrogens on breast

cancer progression, adipose tissue, and the brain, using well-established laboratory animal 

models; and (2) to determine bioavailability of soy isoflavones in neonatal and adult male 

non-human primates.

Interagency Collaboration on

Identification of In Vitro and 

Omics Biomarkers for Liver 

Toxicity

[Tong]

(1) To characterize uridine diphosphate (UDP) glucuronosyltransferases (UGT) mRNA expression

in normal and malignant human-breast tissues isolated from the same donor and from different 

donors; (2) to identify polymorphisms in those UGT genes that show significant inter-individual

differences in UGT mRNA expression in all breast tissues; (3) to determine the methylation profile

of those UGTs identified in objective 2 and correlate it to UGT expression; and (4) to determine 

the effects of polymorphisms in UGT genes on glucuronidation of estradiol, 4-hydroxyestrone, 

and 4-hydroxy-tamoxifen using glucuronidation-activity assay and tetrazolium salt 

(MTT)-cytotoxicity assays.

Evaluation of the Genetic Toxicity

and Behavioral Effects of Chronic 

Methylphenidate Exposure in

Juvenile Male Rhesus Monkeys 

(Macaca mulatta)

[Morris]

(1) To determine the baseline frequency of measures of genetic damage in a population of juvenile 

rhesus monkeys; (2) to determine the frequency of these measures of genetic damage in a 

population of juvenile rhesus monkeys at defined intervals during a chronic exposure to

methylphenidate; (3) to determine if chronic exposure to methylphenidate results in measurable

effects on the behavior of juvenile rhesus monkeys utilizing the NCTR Operant Test Battery; and 

(4) to determine the plasma concentration of methylphenidate and its major metabolite, ritalinic 

acid, during the chronic exposure of juvenile rhesus monkeys to the drug.

Molecular Mechanisms

Underlying Gender-associated 

Differences in the Adverse 

Reactions to the Antiretroviral

Agent, Zidovudine (AZT): Role of

Mitochondrial Toxicity

[Desai]

(1) To to elucidate molecular mechanisms of mitochondrial dysfunction that will address 

gender-based differences in adverse effects of antiretroviral drugs, such as AZT; and (2) to provide 

critical information to the FDA for the development of guidelines to plan new treatment strategies

to reduce the frequency and severity of antiretroviral-related toxic effects in women, particularly

in pregnant women.

Neurotoxicity Assessment of Mn

Nanoparticles in PC-12 Cells and 

in Mice

[Ali]

(1) To evaluate the neurotoxicity of different size Mn nanoparticles using PC-12 cultured cells; (2) to

determine if in vitro exposure to Mn nanoparticles selectively induces specific genomic changes 

in PC-12 cultured cells using oligonucleotide microarrays; (3) to determine if multiple doses of

Mn nanoparticles produce reactive oxygen species (ROS), alterations in lipid peroxidation and/

or changes in antioxidant enzymes(catalase, superoxide dismutase, glutathione peroxidase), and 

levels of glutathione in various regions of the mouse brain; (4) to determine if single or multiple

doses of Mn nanoparticles induce specific genomic changes in various regions of the mouse brain

using oligonucleotide microarrays; (5) to determine if single or multiple doses of Mn nanoparticles

produce significant changes in neurotransmitter concentrations in various regions of the mouse 

brain; and (6) to determine if single or multiple doses of Mn nanoparticles produce significant

changes in the formation of 3-nitrotyrosine, an in vivo biomarker for oxidative stress, in various 

regions of the mouse brain.

Methods Development for HighResolution Dedicated Positron

Emission Tomography (microPET)

to Rodent Neuroplasticity and 

Toxicity During Development 

[Wang]

To use the microPET to screen and evaluate in vitro and in vivo measurements from a broad range 

of pathophysiological or pharmacological parameters using specific tracers in the developing rat,

using three different age groups of developing rats (pregnant gestational 

day (GD)18 female rats; postnatal day (PND)7 rat pups; PND35 rats).
22 National Toxicology Program

NCTR/NTP Project 

Project Officer

Objective and/or Project Summary

Cancer Mutations as Biomarkers

of Cancer Risk: Human Studies 

with Implications for 

Personalized Medicine 

[Parsons]

(1) To develop the information necessary for the rational use of oncogene mutations as

quantitative biomarkers of cancer risk, specifically; (2) to determine normal and pathological 

levels of relevant oncogene mutations in multiple human tissues and tumors using allele-specific

competitive blocker polymerase chain reaction (ACB -PCR); (3) to compare the information

derived from human tissues with data generated in a parallel rodent protocol as an approach for 

incorporating carcinogenesis-relevant data into the rodent-to-human extrapolation necessary 

in cancer risk assessment; (4) to validate a streamlined ACB-PCR methodology and to develop 

the methodology necessary to measure oncogene mutation frequency in cell-free DNA isolated

from plasma; and (5) to convey to the regulatory risk assessment community, through a series of

publications, the regulatory significance of the data regarding tumor-associated mutations which 

have and will be generated.

Development of a New Safety

Evaluation Method Using 

MicroRNA (miRNA) Expression

Analysis as a Biomarker for 

Detecting Carcinogens 

[Chen]

(1) To determine miRNA expression profiles of the tumor target tissues of rats and mice treated

with genotoxic carcinogens aristolochic acid, riddelliine and comfrey, and non-genotoxic 

carcinogens, propiconazole, and triadimeforn, and non-carcinogen myclobutanil using microarray 

technologies; (2) to develop a PCR array containing the primers that are specifically used to amplify

carcinogensis-related miRNAs and use the PCR array to conduct time-course and dose-response 

studies for miRNA expression alterations in tissues of rats treated with carcinogens; and (3) to

define the miRNA biomarker genes that are associated with carcinogen exposure by prediction 

of their target genes and determination of their biological functions.

Neurotoxicity Assessment 

of Silver (Ag) Nanoparticles 

in PC-12 Cells and in Rats

[Ali]

(1) To evaluate the neurotoxicity of different sizes of Ag nanoparticles using cultured PC-12 cells;

(2) to determine if in vitro exposure to Ag nanoparticles selectively induces specific genomic

changes in cultured PC-12 cells using microarrays; (3) to determine if single or multiple doses of

Ag nanoparticles produce ROS, alterations in lipid peroxidation and/or changes in antioxidant 

enzymes(catalase, superoxide dismutase, glutathione peroxidase) and glutathione levels in the 

rat brain; (4) to determine if single or multiple doses of Ag nanoparticles induce specific genomic

changes in the rat brain as indicated with microarrays; (5) to determine if single or multiple doses 

of Ag nanoparticles produce significant changes in neurotransmitter concentrations in the brain in

rat; (6) to determine if single or multiple doses of Ag nanoparticles produce significant changes in

the formation of 3-nitrotyrosine, an in vivo biomarker for oxidative stress, in the rat brain; and (7) to

determine if multiple doses of Ag nanoparticles produce morphological alterations in blood-brain

barrier, brain, or other visceral organs of the rat.

Development of High

Throughput Methodology for 

Detection of In Vivo Mutation

in the Endogenous PIG-A Gene 

of Human Blood Cells Using 

Flow Cytometry 

[Dobrovolsky]

(1) To design high throughput methods for detecting PIG-A mutant human red and white blood

cells by flow cytometric detection of cells lacking cell surface protein markers anchored by

glycosyl phosphatidyl inositol (e.g., CD59, CD48); (2) to establish a normal range of PIG-A mutant

frequencies in red and white blood cells and compare these ranges with those of different groups

of human subjects hypothesized to have increased mutational loads (patients with the disease 

paroxysmal nocturnal hemoglobinurea, patients undergoing radiation treatment or chemotherapy

with DNA reactive drugs, and patients predisposed to cancer due to inherited deficiencies in

endogenous pathways); (3) to compare red blood cell PIG-A mutant frequencies determined in

objective 2 with PIG-A mutant frequencies in white blood cells from these samples determined by

limiting-dilution cloning, and (4) to determine the PIG-A DNA sequence changes responsible for 

the white blood cell mutants.

Evaluation of the Ability of Both

the Agar and Microwell Versions

of the MLA to Optimally Detect

the Mutagenic Potential and 

Potency of Complex 

Chemical Mixtures 

[Moore]

To develop science-based best practice standards and tools to incorporate translational and 

applied toxicological advancements into the regulatory science process to create a seamless

bench-to-bedside continuum.

Assessment of Gaseous 

Anesthetics in the Developing

Nonhuman Primate 

[Wang]

(1) To evaluate dose-response effects of gaseous anesthetics to determine if prolonged exposure

to nitrous oxide or isoflurane alone will result in an increase in neuronal cell death, and determine 

if combinations of nitrous oxide and isoflurane will prevent or enhance each other’s effects on the 

developing nonhuman primate; (2) to determine if a relative high dose or prolonged exposure

of the developing nonhuman primates to nitrous oxide or isoflurane alone, or their combination 

will induce long-term behavioral deficits, as well as long-lasting pathological changes; (3) to

determine, using noninvasive imaging techniques (microPET and MRI), if a high dose or prolonged

exposure of the developing nonhuman primates to nitrous oxide or isoflurane alone, or in

combination will induce long-lasting pathological changes; and (4) to identify potential underlying

mechanisms that could link alteration of mitochondrial function and elevation of ROS to gaseous 

anesthetic-induced neuronal cell death.
NTP Annual Report 2011 23

NCTR/NTP Project 

Project Officer

Objective and/or Project Summary

Development of Methods for 

Evaluating DNA Damage using

Single Cell Gel Electrophoresis

(Comet Assay) in Rodents

[Aidoo]

To evaluate and establish methods and conditions that enhance the sensitivity and reproducibility 

of the in vivo alkaline-comet assay for use in preclinical-hazard identification and genotoxicity

testing of food ingredients and chemicals for regulatory purposes.

Gene Expression Responses by

Avirulent Bacillus anthracis and 

Human Epithelial Cells During

Initial Host-Pathogen Contact 

[Khan]

(1) To compare the gene expression profiles of the bacteria and cell lines with and without 

co-culture, focusing on differences that relate to pathogenesis; and (2) to identify key signal

transduction pathways and immune system interaction genes involved in epithelial cell stimulation

by B. anthracis.

Development and Application 

of a Mitochondria-Specific

Gene Array (Mitochip) for the 

Investigation of Clinical and 

Non-Clinical Predictive

Biomarkers of Toxicity

[Desai]

(1) To develop MitoChips for various mammalian species, including rat, non-human primate, and 

human; (2) to do transcriptional profiling of mitochondria-related genes using mitochondria-specific

gene arrays to investigate the mechanisms of drug toxicities and degenerative diseases associated 

with mitochondrial dysfunction in different mammalian species; and (3) to characterize 

species-specific transcriptional profiles to predict risk of drug toxicity or disease onset in 

different mammalian species

Inactivation of UGT in Human 

Breast Tissue: Accessing Cancer

Risk, Tamoxifen Safety 

and Toxicity

[Starlard-Davenport]

(1) To characterize UGT mRNA expression in normal and malignant human breast tissues isolated

from the same donor and different donors; (2) to identify polymorphisms in the UGT genes that

show significant inter-individual differences in UGT mRNA expression in all breast tissues; (3) to

determine the methylation profile of the UGTs identified in objective 2 and correlate with UGT 

expression; and (4) to determine the effects of polymorphisms in UGT genes on glucuronidation of

ethinyl estradiol, hydroxy-ethinyl estradiol and 4-hydroxy tamoxifen.

Method Development for Study 

of Antioxidant Properties in

Dietary Supplement

[Fu]

(1) To determine whether or not the studied herbal dietary supplements can enhance or inhibit

free radical formation or lipid peroxidation, mediated by microsomal metabolism, in a dose

dependent manner; (2) to determine the toxic effects including mitochondrial dehydrogenase 

activity, intracellular ROS concentration, and mitochondrial membrane potential, and (3) to use 

electron spin resonance (ESR) oximetry technique to determine the inhibition/induction of lipid 

peroxidation, of the herbal dietary supplements in cells, including A549 human lung carcinoma

cells and rabbit brain rBCECs cells.

Use of ESR Spectroscopy to

Characterize the Interactions

Between Nanoscale Materials 

and Model Biological Systems 

[Fu]

(1) To determine whether nanomaterials can catalyze Fenton reaction to initiate hydroxyl radical 

formation in a nanoparticle size dependent manner, or reduced by natural reducing agents, such

as ascorbic acid and glutathione, leading to the formation of reactive oxygen species; (2) to 

determine whether nanomaterials enhance or inhibit free radical formation mediated by microsomal

metabolism or inhibit microsomal metabolism mediated by lipid peroxidation, in a nanoparticle 

size dependent manner; (3) to determine the toxic effects, including mitochondrial dehydrogenase 

activity, intracellular ROS concentration, and mitochondrial membrane potential, and (4) to use ESR 

oximetry technique to determine the inhibition/induction of lipid peroxidation of nanomaterials of

different particle size in cells (A549 human lung carcinoma cells and rabbit brain rBCECs cells).

Neurotoxicity Assessment of

Carbon Nanotubes (CNTs) and 

Gold Nanoparticles using a Brain 

Microvascular Endothelial Cell

System, PC-12 Cultured Cells, 

and a Whole Animal Model 

(Rats or Mice) 

[Ali]

(1) To evaluate the neurotoxicity of single and multiple wall and different sizes of CNTs in brain

microvascular endothelial cells system and PC-12 cultured cells; (2) to determine, using microarrays, 

if the in vitro exposure to these CNTs(single and multiple walls) selectively induce specific 

genomic/proteomic changes; (3) to determine if acute or chronic exposure of CNTs in the mouse 

brain produce (a) ROS, alterations in lipid peroxidation and/or changes in antioxidant enzymes

(catalase, superoxide dismutase, glutathione peroxidase) and glutathione levels, (b) specific 

genomic/proteomic changes indicated with microarrays, or (c) changes in neurotransmitter

concentrations, formation of 3-nitrotyrosine, an in vivo biomarker for oxidative stress; and 

(3) to determine if acute or chronic exposure of CNTs produce morphological alterations in 

the brain or visceral organs of the mice.
24 National Toxicology Program

NCTR/NTP Project 

Project Officer

Objective and/or Project Summary

Assessment of the 

Nephrotoxicity of a Seven-Day 

Combined Exposure to Melamine

and Cyanuric Acid 

[Gamboa da Costa]

To investigate the nephrotoxic effect of a seven-day co-exposure to melamine and cyanuric acid 

in Fischer 344 rats.

Studies Comparing the 

Neurotoxicology of

Amphetamine with

Methamphetamine and 

Methylphenidate (Ritalin)

[Levi]

(1) To determine the appropriate dose range and plasma levels of methylphenidate that produce 

hyperthermic profile similar to that produced by neurotoxic doses of amphetamine and 

methamphetamine and (2) to evaluate the hyperthermic profiles resulting from the selected dose.

Methylphenidate (Ritalin)

Exposure during Pregnancy: 

Assessment of Neurotoxicity 

in Offspring 

[Ferguson]

To quantify the neurobehavioral toxicity associated with pre- and early postnatal treatment with

methylphenidate in rats, measuring a wide range of behaviors at preweaning, adolescent, and 

adult ages.

Biomarkers of Liver Toxicity

[Salminen]

(1) To determine biomarkers of hepatotoxicity in preclinical studies that are more predictive of

adverse effects in humans and (2) to qualify biomarkers(e.g., via the FDA/EMEA qualification 

process) and assess their potential translation for clinical use.

Development of a FDA Resource

and Knowledge Base for Sex 

Difference in Drug-Induced Liver 

Injury (DILI) 

[Tong]

To develop a knowledge base for the sex differences in drug-induced liver injury (DILI) through 

analyzing and modeling the molecular data in public domain, specifically through: (a) collection

of the genomic data from public resources and through collaborations; (b) development of a 

standard data curation model for the sex-biased DILI in ArrayTrack to manage the collected data; 

(c) meta-analysis, text mining, and network analyses to develop a relationship between drugs,

molecular signatures, liver-specific biomarkers, genes/proteins functions, pathways and sex-biased

liver toxicity.

3D- and 4D- Quantitative

Spectrometric Data-activity 

Relationship (QSDAR) 

Modeling Applied to Various 

Toxicological Endpoints 

[Beger]

(1) To develop 3D- and 4D- QSDAR models for endocrine disruptors, lowest-observed-adverse-effects

level, and no observed-adverse-effects level, and other relevant toxicological endpoints; and 

(2) to determine how the technique used to predict 13C or 15N NMR spectra affects on 3D-QSDAR

modeling.

Methods Development for 

Toxicity Assays using the 

Zebrafish Embryo as a Model 

System: Whole Animal High

Throughput Assays for 

Chemical Testing 

[Kanungo]

(1) To use the established high throughput assay system with zebrafish embryos for morphological 

and behavioral endpoints of toxicity, and subtle organ-specific toxicities to study the effect of

methamphetamine on Zebra fish embryos, especially relating to sensory and motor neuron

development; (2) to determine if CNTs pass through the blood brain barrier in zebrafish embryos 

and have any toxic effects on early development, and if so, determine if these nanomaterials

generate ROS, cause the depletion of dopamine and its metabolites, dihydroxyphenylacetic acid

and homovanillic acid, and alter markers of oxidative stress;and (3) to study the effect of nicotine

on zebrafish embryos, especially relating to sensory and motor neuron development and the 

mechanism of action.

In Vitro Assay to Predict 

Developmental Neurotoxicity 

of Pediatric Anesthetics 

[Wang]

(1) To use rodent in vitro organotypic and primary culture models to examine effects of propofol

(gamma-aminobutyric acid [GABA] A agonist), baclofen (GABA B agonist), diazepam (GABA 

A agonist), pentobarbital (GABA A agonist and 2-amino-3-(5-methyl-3-oxo-1, 2- oxazol-4-yl)

propanoic acid antagonist), etomidate (GABA A agonist), sevoflurane (N-methyl-D-aspartic acid

[NMDA] antagonist and GABA agonist), fentanyl (opiate agonist) and anesthetic combinations

commonly used in pediatric surgical procedures; (2) to determine the utility of in vitro culture

systems to predict in vivo outcomes in subsequent studies; (3) to determine the dose and 

time-course over which the potential neurotoxic effects of anesthetics are expressed in the 

developing brain; (4) to determine effective ways to protect against anesthetic-induced

developmental neurotoxicity that have potential clinical utility; (5) to identify mechanisms 

that link altered NMDA receptor function and/or elevation of ROS to anesthetic-induced

neuroapoptosis; and (6) to identify biomarkers such as genomic pathway signatures and 

determine their validity for predicting in vitro outcomes of pediatric anesthetic exposure.
NTP Annual Report 2011 25

NCTR/NTP Project 

Project Officer

Objective and/or Project Summary

PIG-A Mutagenesis; An

International Validation Study 

Comparing PIG-A Mutation in

Rats With Other Biomarkers of

Genetic Toxicity 

[Heflich]

(1) To generate data using a standardized protocol that, in combination with results from other 

investigators, will be used to determine the sensitivity, specificity, and portability of the rat red 

blood cell/reticulocyte (RBC/RET) PIG-A gene mutation assay; and (2) to perform the in vivo

Comet, micronucleus, and the PIG-A and HPRT lymphocyte gene mutation assays in conjunction 

with the RBC/RET PIG-A assay, to determine how the RBC/RET PIG-A assay compares in terms of

sensitivity and specificity with these other in vivo assays that have been used or considered for use 

as regulatory assays.

Effect of Soy-containing 

Diets on Ammonium 

Perchlorate-induced Thyroid 

Toxicity in Sprague-Dawley (SD)

Rats (II) 

[Doerge]

To determine the effect of dietary whole soy and purified genistein, the principal soy isoflavone, 

on the dose-response characteristics for perchlorate-induced thyroid toxicity in male SD rats.

Study of Drug Induced Liver 

Toxicity Using Primary Rat and 

Mouse Hepatocytes 

[Guo]

To obtain signature gene and protein expression patterns for mouse and rat primary hepatocytes 

following xenobiotic induced toxicity.

The Sequencing Quality 

Control Project 

[Shi]

(1) To assess the technical performance of the next-generation sequencing platforms that will be

used in toxicology studies, by using reference RNA samples analyzed at multiple times; (2) to use 

one next-generation sequencing platform to sequence RNA samples from a toxicogenomic study;

and (3) to use reference DNA samples to assess technical performance for exome sequencing.

Identification of New Mechanistic 

Biomarkers of Adverse Responses 

to Acetaminophen 

[Beger]

To provide pharmacokinetics data for acetaminophen and acetaminophen/N-acetylcysteine, 

in addition to metabolomics analyses of metabolites, involved in one carbon metabolism and 

energy metabolism such as those of the citric acid cycle in blood and urine samples from children 

and adolescents who have adverse reactions to acetaminophen.

Genotyping of Transporter 

Genes Associated with Gender 

Differences and Promoter

Methylation of UGT1A1 in 

Human Liver: A Means of

Assessing Safety and Toxicity 

of Chemotherapeutic Drugs 

[Lyn-Cook]

(1) To identify polymorphisms in drug transporter genes identified to be differentially expressed 

according to gender in human liver samples, (2) to correlate polymorphism frequencies in male

and female to gene expression, (3) to evaluate the methylation profile of UGT1A1 promoter in

human liver samples from male and female and correlate it to expression of UGT and its activity, 

and (4) to evaluate effects of polymorphisms in transporter genes on uptake and clearance of

chemotherapeutic drugs in a functional assay using the B-CLEAR human in vitro model.

Proteomic Approaches to

Elicidate 

Biodegradative Pathways 

[Wang]

To use a proteomic approach to isolate putative catabolic proteins that are over-expressed when 

microorganisms are grown in the presence of toxic polycyclic aromatic hydrocarbons.

NCTR/Office of Regulatory 

Affairs (ORA) Nanotechnology

Core Facility

[Howard]

(1) To support the needs of NCTR research projects to characterize nanoscale materials used in

toxicology tests and to detect these materials in biological samples, (2) to support the needs of

ORA to detect and characterize nanoscale materials in FDA regulated products, and (3) to develop 

methods and approaches to anticipate needs of the research community.

Analytical Assay for 

Photochemical Generation 

of Hydroxyl Radical 

[Howard]

(1) To provide support for analysis of the photoactivation of nanomaterials using the coumarin3-carboxylic acid oxidation assay, (2) to provide particle-size analysis for all nanomaterials being 

analyzed; and (3) to improve the assay using ultraviolet light diode laser as a replacement to the 

existing broad band ultraviolet light A source.
26 National Toxicology Program

Division of the National Toxicology Program

In FY 2011 the Division of the National Toxicology Program (DNTP) was established within the NIEHS. The 

NTP was formerly a program within DIR and the change was in recognition of the unique mission of NTP 

and the unique training requirements and capabilities of its staff. Secretary Sebelius signed the reorganization

documents following a review by the Office of Management and Budget. Congress was notified of the change 

in spring of 2011 and a Federal Register notice (http://ntp.niehs.nih.gov/go/frn) announcing the reorganization

was published on April 28, 2011. 

The new structure affords greater efficiency at NIEHS, by making a clear staffing and budgetary demarcation 

between the components of the NIEHS carrying out work dedicated solely to the pursuit of the goals and 

initiatives of the NTP, and the principal investigator-initiated efforts carried out by staff in the DIR. 

Dr. John Bucher serves as director of the DNTP. Information about the DNTP is available at 

http://www.niehs.nih.gov/research/atniehs/dntp/index.cfm.

NIEHS/NTP staff at the NIEHS building, Research Triangle Park, NC
NTP Annual Report 2011 27

NIEHS Strategic Planning

In 2011 NIEHS initiated a strategic planning process, which will guide the 

institute’s activities over the next five years. The process was designed with

three phases, the first of which was soliciting stakeholder input. A Stakeholder 

Community Workshop was held in July 2011, with more than 180 participants. 

It employed a Modified Open Space format, which ultimately yielded a slate 

of 13 “big topics” for further development. The strategic planning process will

continue in FY 2012 to develop the mission statement, vision statement, and 

supporting pillars. Information about the strategic plan is available at 

http://www.niehs.nih.gov/strategicplan.

Gulf Oil Spill Response

Immediately following the Deepwater Horizon explosion 

on April 20, 2010, and the ensuing massive oil spill in the 

Gulf of Mexico, NTP and NIEHS began coordinated efforts 

with other federal agencies (including FDA, OSHA, 

NHGRI, NIOSH, ATSDR, EPA, National Oceanographic 

and Atmospheric Administration, United States Fish

and Wildlife Service, U.S. Geological Survey, and U.S.

Coast Guard) to limit the adverse impacts of the oil and 

dispersants on human health, ecological health, 

and food sources.

Early efforts by NTP focused upon assembly and 

distribution of information on characteristics and toxicity

of oil and oil dispersants. The NTP is pursuing several 

lines of research to address concerns regarding the 

safety of Gulf seafood, hazards to offshore and onshore 

cleanup workers, and potential long-term health impacts 

of residual oil in the environment. NIEHS/NTP obtained

samples of source oil, tar balls, and the dispersant

COREXIT® EC9500A. The NTP is conducting analytical

chemistry studies to better understand the composition 

of the source oil and tar balls and oiled sediment collected 

along the shoreline. The focus of these analyses is on 

components that are more likely to persist in the 

environment and potentially lead to residual human 

exposures. These analytical chemistry efforts will help

understand exposures to response workers and inform 

exposure assessments in the Gulf Long-term Follow-up 

Study (GuLF STUDY, see below). The NTP will also use this

information to develop a toxicology research program 

on polycyclic aromatic hydrocarbons(PAHs). The aim of

this program isto further characterize the hazard of PAHs
28 National Toxicology Program

present in oil to determine if those PAHs routinely monitored in environmental media adequately capture the 

hazard of crude oil exposures, and to better understand whether there are combinations of individual PAHs that

require they be considered in aggregate when establishing safe exposure levels.

The NTP’s toxicology assessments will inform the ongoing GuLF study, which will assess the possible health

effects of the oil spill on up to 55,000 cleanup workers and volunteers in coastal towns across Louisiana, Mississippi,

Alabama, and Florida. This longitudinal cohort study, coordinated by the NIEHS, is the largest health study of

its kind ever conducted, and is an important component of the comprehensive federal response to the oil spill.

Information about the GuLF STUDY is available at http://nihgulfstudy.org/. NIEHS also led the creation of a trans-NIH

funded network of community and university partnerships that will investigate personal and community health

effects stemming from the Deepwater Horizon oil spill and enhance community resiliency to potential disasters.

Information about NIEHS oil spill response efforts is available at http://www.niehs.nih.gov/about/od/programs/gulfspill

and information about the overall federal oil spill response efforts is available at http://www.restorethegulf.gov/.

Contact Information: Dr. Scott Masten, masten@niehs.nih.gov

Workshop: Advancing Research on Mixtures: 

New Perspectives and Approaches for Predicting Adverse Human Health Effects

A collaborative effort between Division of

Extramural Research and Training and the NTP 

culminated in a workshop on September 26-27,

2011, in Chapel Hill, NC, that brought together

experts from diverse disciplines (exposure science,

epidemiology, toxicology, statistics, and risk

assessment) to focus on mixtures. The purpose 

of the workshop was to identify and address key 

issues presented in research on mixtures. The 

workshop goals were to (1) identify and prioritize 

the knowledge gaps and challenges in mixtures 

research in toxicology, epidemiology, exposure

science, risk assessment, and statistics; (2) obtain

advice on integrating multidisciplinary capabilities 

to address critical topics in mixtures research; 

(3) provide recommendations for research on 

key topics; (4) inform the development of a 

long-term NIEHS mixtures research agenda; 

and (5) foster collaborations between extramural 

and NIEHS scientists. The NIEHS will use the results 

from the workshop to inform the development 

of an intramural and extramural mixtures research

strategy and to provide input to the scientific

community for advancing mixtures research.

Contact Information: Dr. Cynthia Rider, cynthia.rider@nih.gov
NTP Annual Report 2011 29

Chemical Effects in Biological Systems (CEBS): 

A Publicly Accessible NTP Integrated Data Management System

CEBS is an NTP data management tool that

houses biological study data of interest

to toxicologists and environmental health

scientists. The CEBS database was designed

as a public reference repository and accepts 

studies from many sources. Governmental, 

academic, and pharmaceutical company 

laboratories contribute peer-reviewed studies 

to CEBS. 

An important feature of the database is its 

flexibility — CEBS can house “any” type of

biological measurements from “any” type of

study design. This flexibility in CEBS allows

data from different sources to be integrated

into one place, to permit data mining and 

analysis. In addition, any one study can have different types of data associated with it such as blood chemistry,

histopathology, and microarray — all viewed together under one study name. The investigator does not have to

access different databases, each housing one specific type of data. 

In 2011, the NTP added over 9000 studies to CEBS. Many of these additions were legacy NTP genetic toxicology, 

immunotoxicology, and carcinogenicity/toxicology studies. The legacy studies investigated the potential of

many different chemicals to cause genetic damage or mutations, adverse effects on the immune system, tumor 

formation, or toxicity in in vitro or in vivo models. The high throughput toxicological screening Phase I Tox21 

studies, which use in vitro biochemical and cell-based assays, were also added to CEBS.

The NTP is continuing to add legacy NTP data to CEBS and will be adding Phase II Tox21 data. When this project 

is completed, all the public NTP data will be accessible in CEBS for reference and for data mining.

CEBS consists of a database, a user interface, and tools for loading data and managing it for display. Testing has 

also begun on analytical tools and incorporating graphical displays of data. The user is able to download raw data

from a single study or batch files of many studies using the new File Transfer Protocol (FTP) site. Study conclusions 

are also shown and can be searched.

CEBS captures the details of the study design and execution plus the biological responses of subjects in a way 

that permits searching, filtering, and sub-setting of the data. The user can view the details of each study, search

for particular studies or study subjects of interest based on treatment, response, or other characteristics, and then

either analyze the data within CEBS or download for import into other tools. CEBS can be accessed at 

http://cebs.niehs.nih.gov.

Contact information: Dr. Jennifer Fostel, fostel@niehs.nih.gov
30 National Toxicology Program

Table 5. Use of NTP Study Data or Reccomendations by Federal and State Regulatory

Agencies in FY 2011 

Agency,Title, Additional Information Information Cited

California EPA Office of Environmental 

Health Hazard Assessment (OEHHA):

Chemical Listed as Known to the State of

California to Cause Reproductive Toxicity:

Acrylamide [79-06-1]

NTP-CERHR Monograph on the Potential Human Reproductive and Developmental 

Effects of Acrylamide (2005)

California EPA OEHHA:

Proposition 65 - Notice of Intent to List: 

Androstenedione [63-05-8]

NTP Toxicology and Carcinogenesis Studies of Androstenedione in F344/N Rats and 

B6C3F1 Mice (Gavage Studies). TR-560 (2010)

California EPA OEHHA:

Proposition 65 - Notice of Intent to List: 

Dibromoacetonitrile [3252-43-5]

NTP Toxicology and Carcinogenesis Studies of Dibromoacetonitrile in F344/N Rats and 

B6C3F1 Mice (Drinking Water Studies). TR-544 (2010)

California EPA OEHHA: 

Proposition 65 - Notice of Intent to List: 

Malonaldehyde, Sodium Salt [24382-04-5]

NTP Toxicology and Carcinogenesis Studies of Malonaldehyde, Sodium Salt 

(3-Hydroxy-2-propenal, Sodium Salt) in F344/N Rats and B6C3F1 Mice 

(Gavage Studies). TR-331 (1988)

Department of Defense: Defense Federal 

Acquisition Regulation Supplement; 

Minimizing the Use of Materials 

Containing Hexavalent Chromium 

(DFARS Case 2009–D004)

The RoC first listed hexavalent chromium compounds as known human carcinogens 

in 1980

Department of Transportation: Revision

to the List of Hazardous Substances

and Reportable Quantities – removing

saccharin [81-07-2] and its salts

RoC, Appendix B:saccharin and its salts do not pose a present or potential risk 

of causing carcinogenic effects on humans.

Department of Transportation: 

Greenhouse Gas Emissions Standards 

and Fuel Efficiency Standards for 

Medium- and Heavy-Duty Engines 

and Vehicles

RoC has listed: 

• benzene [71-43-2] and 1,3-butadiene [106-99-0] as known human carcinogens 

(1980 and 1989)

• acetaldehyde [75-07-0] as reasonably anticipated to be a human carcinogen (1991)

• naphthalene [91-20-3] as reasonably anticipated to be a human carcinogen (2004)

EPA: Testing of Certain High Production

Volume Chemicals; Second Group 

of Chemicals

NICEATM studies (2003): Test Method Protocol for Solubility Determination, 

In Vitro Cytotoxicity Validation Study — Phase III;

Test Method Protocol for the BALB/c 3T3 Neutral Red Uptake Cytotoxicity Test, a Test for 

Basal Cytotoxicity for an In Vitro Validation Study — Phase III ; Test Method Protocol for 

the NHK Neutral Red Uptake Cytotoxicity Test, a Test for Basal Cytotoxicity for an In Vitro

Validation Study — Phase III

EPA: (S,S)-Ethylenediamine Disuccinic Acid

Trisodium Salt [ 178949–82–1] Exemption 

From the Requirement of a Tolerance

Bioassay of Trisodium ethylenediaminetetraacetate trihydrate (EDTA) for Possible

Carcinogenicity in mice and rats showed no carcinogenic potential. TR-11 (1977). 

Since (S,S)-ethylenediamine disuccinic acid trisodium salt ((S,S)-EDDS) is similar to 

EDTA, (S,S)-EDDS is not likely to be carcinogenic to humans at low doses.

EPA: Sodium Ferric EDTA [178949–82–1];

Exemption From the Requirement 

of a Tolerance

Bioassay of Trisodium ethylenediaminetetraacetate trihydrate (EDTA) for Possible

Carcinogenicity in mice and rats showed no carcinogenic potential. TR-11 (1977).

Use of NTP Products by Other Agencies

Federal and state regulatory agencies use NTP study data and recommendations in considering the need to

regulate and test specific chemicals to protect human health. Table 5 lists NTP data and recommendations used 

in FY 2011.
NTP Annual Report 2011 31

Agency,Title, Additional Information Information Cited

EPA: National Emission Standards for 

Hazardous Air Pollutants From Coal- and 

Oil-Fired Electric Utility Steam Generating

Units and Standards of Performance for 

Fossil-Fuel-Fired Electric Utility, 

Industrial-Commercial-Institutional, and 

Small Industrial-Commercial-Institutional

Steam Generating Units

RoC has listed: 

• benzene [71-43-2] as known human carcinogens (1980)

• acetaldehyde [75-07-0] as reasonably anticipated to be a human carcinogen (1991)

EPA:Agency Information Collection

Activities; Submission to OMB for Review

and Approval; Proposed Collections; Toxic 

Chemical Release Reporting; Request 

for Comments on Proposed Renewal 

of Form R and Form A, Including Minor 

Form Revisions and the Ratio-Based 

Burden Methodology

RoC

EPA: Ethylene Glycol; Exemption from 

the Requirement of a Tolerance

Toxicology and Carcinogenesis Studies of Ethylene Glycol in B6C3F1 Mice (Feed Studies). 

TR-413 (1993)

EPA: Diethylene Glycol MonoEthyl 

Ether (DEGEE); Exemption From the 

Requirement of a Tolerance

Toxicology and Carcinogenesis Studies of Ethylene Glycol in B6C3F1 Mice (Feed Studies). 

TR-413 (1993)

EPA: Testing of BPA [80-05-7] NTP-CERHR Monograph on the Potential Human Reproductive and Developmental 

Effects of Bisphenol A (2008)

EPA: Carboxymethyl Guar Gum Sodium

Salt and Carboxymethyl-Hydroxypropyl 

Guar; Exemption From the Requirement 

of a Tolerance

Carcinogenesis Bioassay of Guar Gum in F344 Rats and B6C3F1 Mice (Feed Study) TR-229

(1982)

EPA: Harmful and Potentially Harmful 

Constituents in Tobacco Products and 

Tobacco Smoke; Request for Comments

RoC has identified constituents in tobacco products and tobacco smoke as human 

carcinogens or reasonably anticipated to be human carcinogens (2000)

EPA: Greenhouse Gas Emissions Standards 

and Fuel Efficiency Standards for 

Medium- and Heavy-Duty Engines 

and Vehicles

RoC has listed: 

• benzene [71-43-2] and 1,3-butadiene [106-99-0] as known human carcinogens (1980 

and 1989)

• acetaldehyde [75-07-0] as reasonably anticipated to be a human carcinogen (1991)

• naphthalene [91-20-3] as reasonably anticipated to be a human carcinogen (2004)

FDA: Labeling and Effectiveness Testing;

Sunscreen Drug Products for 

Over-the-Counter Human Use

RoC indicates that UVA and UVB radiation across the spectrum are known human 

carcinogens, but either UVA or UVB radiation alone is reasonably anticipated to be a 

human carcinogen. Cited in the National Toxicology Program’s Report on Carcinogens

(2002).

U.S. Forest Service: Information Collection; 

Qualified Products List for Long-Term 

Retardant for Wildland Firefighting

The Forest Service evaluates and approves commercial wildland firefighting chemicals. 

Products must not contain chemicals listed as a Chemical of Concern. 

To create their list, they include chemicals appearing in the RoC.

A complete listing of NTP studies used by Federal and state regulatory agencies is at: http://ntp.niehs.nih.gov/go/regact
32 National Toxicology Program

The Report on Carcinogens (RoC) is a congressionally mandated listing [Section 301(b)(4) of the Public Health

Services Act, 42 U.S.C. 241(b)(4)] of substances (1) that either are known to be human carcinogens or may 

reasonably be anticipated to be human carcinogens and (2) to which a significant number of persons residing

in the United States are exposed. 

Each substance listed in the RoC has a profile, which contains the listing status, a summary of the cancer studies

supporting the listing status, information on human exposure, and Federal regulations to reduce exposure.The 

RoC is a cumulative report and consists of substances newly reviewed in addition to those listed in previous

editions. The Secretary of Health and Human Services has delegated preparation of the RoC to the NTP, with

assistance from other Federal health and regulatory agencies. Preparation of the RoC is managed by the Office 

of the RoC (ORoC) under the direction of Dr. Ruth Lunn. Integrated Laboratory Systems, Inc. (ILS) provided

contract support for preparation of the RoC in FY 2011.

The 12th RoC, the latest edition, was published on June 10, 2011. The 12th RoC contains 240 listings, including

some classes of related chemicals or agents. There are six new listings and two revised listings in this edition (see

Table 6 or the website http://ntp.niehs.nih.gov/go/roc12).

Table 6. New or Revised Listing in the 12th Report on Carcinogens, published in FY 2011

Candidate Substance 

[CASRN]

Primary Uses/

Exposures

Listing Status

Aristolochic Acids (AA) A family of nitrophenanthrene carboxylic acids that occurs

naturally in plants in the Aristolochiaceae family, primarily of

the genera Aristolochia and Asarum. Botanical productsfrom

plants containing aristolochic acids are used in traditional folk

medicines, particularly in Chinese herbal medicine, and have

been used inadvertently as part of a weight-loss regimen.

Known to be human carcinogens

Captafol A broad-spectrum fungicide that has been widely used since Reasonably anticipated 

[2425-06-01] 1961 for the control of fungal diseases in fruits, vegetables, and

some other plants. Use of captafol in the United States was 

banned in 1999.

to be a human carcinogen

Cobalt–Tungsten Carbide: Composites of tungsten-carbides with a metallic cobalt, used Reasonably anticipated 

Powders and Hard Metals to make cutting and grinding tools, dies, and wear products for 

a broad spectrum of industries, including mining and oil and

gas drilling.

to be a human carcinogen

Formaldehyde

[50-00-0] 

(nominated formaldehyde 

for reclassification)

Primarily used to produce resins for the production of many 

different products, including plastics, adhesives and binders

for wood products, pulp and paper, and synthetic fibers, and in

textile finishing. It is also used as a disinfectant and preservative 

and as an intermediate for many industrial chemicals.

Known to be a human carcinogen

Certain Glass Wool Fibers Glass wool fibers, which are a type of synthetic vitreous fibers, Glass wool (respirable) was first

(inhalable) are an inorganic fibrous material manufactured primarily listed in the 7th RoC as reasonably 

(nominated glass wool from glass and processed inorganic oxides. The major uses of anticipated to be a human

(respirable size) for delisting) glass wool are in thermal, electrical, and acoustical insulation,

weatherproofing, and filtration media. Some special-purpose

glass wool fibers are used for high-efficiency air filtration media

and acid battery separators.

carcinogen, but the scope of the

listing changed and now certain

glass wool fibers (inhalable) are 

listed as reasonably anticipated

to be human carcinogens.

ortho-Nitrotoluene

[88-72-2]

A chemical intermediate used in the synthesis of azo dyes. 

It is also used (either directly or as an intermediate) in the

production of other dyes, agricultural chemicals, rubber

chemicals, pesticides, petrochemicals, pharmaceuticals, 

and explosives.

Reasonably anticipated 

to be a human carcinogen
Candidate Substance Primary Uses/ Listing Status 

[CASRN] Exposures 

Riddelliine Found in a class of plants growing in the western United States. Reasonably anticipated 

[2346-96-0] Some common names for Senecio plants are ragwort and to be a human carcinogen 

groundsel. These plants are not used for food in the U.S., 

but have been used in medicinal herb preparations. 

Styrene Used worldwide in the production of polymers, which are Reasonably anticipated 

[100-42-5] incorporated into products such as rubber, plastic, insulation, to be a human carcinogen 

fiberglass, pipes, automobile parts, food containers, and 

carpet backing. 

Candidate Substance 

[CASRN] 

Primary Uses/Exposures 

Alkenylbenzenes 

(selected dietary): 

estragole [140-67-0] 

myristicin [607-91-0] 

isosafrole [120-58-1] 

Naturally occurring organic chemicals used primarily as an additive, flavoring, or fragrance in food 

products, but may also be in other products such as cosmetic and cleaning agents. 

1- Bromopropane 

[106-94-5] 

Primarily used as a solvent for fats, waxes, or resins and in some spray adhesives and aerosol 

propellant applications. Also used as an intermediate in the synthesis of pharmaceuticals, 

insecticides, quarternary ammonium compounds, flavors and fragrances. 

Carbon black 

[1333-86-4] 

Family of products consisting of elemental carbon in the form of colloidal particles that rapidly form 

aggregates. The products are used primarily for rubber reinforcement, but are also used as black 

pigments and for electrical conductivity purposes. 

Cumene 

[98-82-8] 

An alkylated benzene used primarily to produce phenol and acetone. 

Diesel exhaust particulates 

(currently listed as reasonably 

anticipated to be a human 

carcinogen) 

A mixture of combustion products of diesel fuel; the exact composition of the mixture depends 

on the nature of the engine, operating conditions, lubricating oil, additives, emission control syste

and fuel composition. Occupational exposure occurs primarily among miners (who use diesel po

equipment), railroad workers and transportation workers such as truck drivers. 

m, 

wer 

Ethylbenzene 

[100-41-4] 

An alkylated benzene used primarily for the manufacture of styrene. 

Helicobacter pylori Bacteria responsible for most ulcers and many cases of stomach inflammation 

(chronic gastritis). 

Indium compounds
 Indium compounds are chemicals that include the metal indium. Indium phosphide is used 

primarily in the semiconductor industry; however, most of the semiconductor industry in the 

U.S. is silicon based. Other indium compounds (such as indium tin oxide) are primarily used in 

optoelectronics and flat panel display technology. 

Iron (excess) or iron overload
 

[7439-89-6] 


(elemental iron)
 

Iron is a transition metal that exists mainly in the ferrous (2+) and ferric (3+) oxidation states in 

biological systems. It is used for medical treatment, as well as for other consumer and industrial 

needs. It is also found naturally in food. Iron overload disorders can be hereditary, acquired 

or iatrogenic. 

During FY 2012, the NTP will make revisions to the multi-step process used to review nominations for the 

RoC. Public input on and BSC review of the proposed process will be sought and used to update the process. 

Information is available at http://ntp.niehs.nih.gov/go/rocprocess. 

Also during FY 2012, the 13th RoC will be under development. Substances nominated for possible review for future 

editions of the RoC are in Table 7 (http://ntp.niehs.nih.gov/go/rocnom). 

Table 7. Substances Nominated to the Report on Carcinogens at the end of FY 2011 

NTP Annual Report 2011 33 
34 National Toxicology Program

Candidate Substance 

[CASRN]

Primary Uses/Exposures

Pentachlorophenol 

[87-86-5]

General biocide that has been used extensively as a fungicide, bactericide, herbicide and insecticide

by agriculture and other industries. In 1987, over-the-counter use was banned and other uses

restricted. The only current use of pentachlorophenol in the United States is defined as a ‘heavy

duty’ wood preservative that is used primarily in the treatment of utility poles and cross arms.

Shiftwork involving light at night Shift work is synonymous with irregular, odd, flexible, variable, unusual, non-standard working hours. 

Shift work can be with or without night work.

ortho-Toluidine

[95-53-4] 

(currently listed as 

reasonably anticipated 

to be a human carcinogen) 

Arylamine used as an intermediate to manufacture herbicides, dyes, pigments and rubber

chemicals.

Trichloroethylene 

[79-01-6] 

(currently listed as 

reasonably anticipated 

to be a human carcinogen)

Halogenated alkene used mainly as an intermediate for hydrofluorocarbon production (67%) and 

as a degreaser for metal parts (30%).It is a major ingredient in many consumer products and has

been found in food and drinking water, and the environment (ambient air, ground and tap water).

Uranium (depleted) A byproduct of the production of enriched uranium for use in nuclear reactors. It has a lower 

content of the isotope U-235 and emits fewer alpha particles than natural uranium. Used by the

military for armor-piercing munitions and armor plates. Used as counterbalance weight in aircraft,

and radiation shielding in medical radiation therapy.

Viruses (selected): Kaposi’s

sarcoma – associated 

herpesvirus, Epstein-Barr virus,

human T-cell lymphotropic 

virus type 1, human 

immunodeficiency virus (HIV),

and Merkel cell polyomavirus

Epstein-Barr virus and Kaposi’s sarcoma – associated herpesvirus are members of the herpesvirus

family, which are double stranded DNA viruses with a capsid, and cause characteristic lesions. 

Epstein-Barr virus causes infectious mononucleosis. 

HIV is a retrovirus that causes acquired immunodeficiency syndrome.

Human T-cell lymphotropic virus type 1 is a retrovirus that causes multiple disorders including 

an inflammatory neurological disease. 

Merkel cell polyomavirus is a non-enveloped, double-stranded DNA virus.

Contact Information: Dr. Ruth Lunn, Director, ORoC, lunn@niehs.nih.gov. RoC website: http://ntp.niehs.nih.gov/go/roc
NTP Annual Report 2011 35

In FY 2011, NIEHS/NTP established the Office of Health Assessment and Translation (OHAT), formerly the Center 

for the Evaluation of Risks to Human Reproduction (CERHR), to serve as an environmental health resource to

the public and to regulatory and health agencies. This office conducts evaluations to assess the evidence that

environmental chemicals, physical substances, or mixtures (collectively referred to as “substances”) cause adverse

health effects and provides opinions on whether these substances may be of concern given what is known about 

current human exposure levels. 

NTP’s CERHR focused on effects on reproduction and development, which was appropriate at the time because 

of a strong interest in these health outcomes by the public, regulatory and health agencies, and the scientific

community. Using approaches developed for CERHR evaluations, OHAT extends them to other important heath 

outcomes, using a more flexible scientific analysis program that allows the exploration of linkages between 

toxicity pathways and disease outcomes (see Environmental Health Perspectives, 119(5) May 2011). OHAT accepts 

nominations of substances considered potential health hazards from individuals and from non-profit, commercial, 

governmental or other organizations. OHAT organizes workshops and state-of-the-science evaluations to address

issues of importance in environmental health sciences. OHAT assessments are published as NTP Monographs. 

NTP Workshop: Role of Environmental Chemicals 

in the Development of Diabetes and Obesity

To address the concept that environmental chemicals may be

contributing factors to the epidemics of diabetes and obesity, the NTP 

held a workshop in Raleigh, NC on January 11–13, 2011. The workshop

evaluated the science associating exposure to certain chemicals 

or chemical classes with the development of diabetes and obesity 

in humans. Goals of the workshop were to (1) evaluate strength/

weaknesses, consistency, and biological plausibility of findings 

reported in humans and experimental animals for certain 

environmental chemicals including arsenic and cadmium, 

persistent organic pollutants, pesticides, bisphenol A, phthalates, 

and organotins; (2) identify the most useful and relevant endpoints in

experimental animals and in vitro models; (3) identify relevant pathways

and biological targets for assays for the Toxicology Testing in the 

21st Century high throughput screening initiative (“Tox21”); and 

(4) identify data gaps and areas for future evaluation/research. 

Preliminary conclusions of the workshop included (1) plausibility of

“obesogen” hypothesis, (2) linkage of type 2 diabetes to certain chemical exposures (3) common mechanistic 

basis for certain chemical classes, (4) utilization of Tox21 approaches to identify substances of potential interest, 

and (5) refinement of endpoints examined using high throughput screening approaches. OHAT will publish an

NTP workshop report based on the discussions at the workshop. Complete information about the workshop is

found at http://ntp.niehs.nih.gov/go/36433.

During FY 2011 two important OHAT evaluations were in progress, Potential Developmental Effects of Cancer

Chemotherapy during Pregnancy (http://ntp.niehs.nih.gov/go/36495) and Heath Effects of Low-level Lead

(http://ntp.niehs.nih.gov/go/36443). Expert panel meetings are anticipated in FY 2012 for both of these evaluations.

Contact Information: Dr. Kristina Thayer, OHAT Director, thayer@niehs.nih.gov. OHAT website 

http://ntp.niehs.nih.gov/go/ohat
36 National Toxicology Program

NIEHS established ICCVAM in 1997 to advance the regulatory acceptance of

scientifically valid alternative testing methods. Alternative methods are those 

methods that reduce, refine (enhance animal well-being and lessen or avoid 

pain and distress), or replace the use of animals. NICEATM was established in

1998 to administer ICCVAM, provide scientific support for ICCVAM activities, 

and conduct independent validation studies on promising test methods.

Dr. William Stokes (Rear Admiral, U.S. Public Health Service) is the NICEATM 

Director and Executive Director of ICCVAM. Integrated Laboratory Systems,

Inc., provided contractsupport for NICEATM in FY 2011. Information about 

ICCVAM and NICEATM is available at http://iccvam.niehs.nih.gov). ICCVAM has 

contributed to the approval or endorsement of 44 alternative safety or potency 

testing methods by Federal regulatory agencies since its first test method

evaluation in 1998. 

NICEATM-ICCVAM organized two workshops in the ICCVAM Workshop Series on Best Practices for Regulatory

Safety Testing (http://iccvam.niehs.nih.gov/meetings/Implement-2011/ImplmtnWksp.htm) at the Natcher Conference 

Center in Bethesda, MD. The specific goals of these workshops were to (1) provide an overview of the available

methods in each area, including the applications, strengths and weaknesses of each method; (2) provide 

information on the procedures for conducting and interpreting data in accordance with regulatory testing 

requirements and guidelines; (3) give participants an opportunity to become familiar with data generated by

each test method; (4) provide a forum for scientists to share information on the appropriate use of results in

regulatory safety testing; (5) discuss challenges of incorporating alternative test methods into regulatory safety

testing guidelines; and (6) identify and discuss new methods in the development and validation pipeline for 

each safety testing area, and ways to increase the availability of high quality data necessary for validating new 

methods. The Assessing the Potential for Chemically Induced Eye Injuries workshop was held on January 19, 2011, 

and the Assessing the Potential for Chemically Induced Allergic Contact Dermatitis workshop was held on January 

20, 2011. Over seventy scientists from industry, academia, research and regulatory agencies, and animal welfare

organizations attended each workshop, which was also made available to remote participants via a live webcast.

On March 6 – 10, 2011, NICEATM-ICCVAM participated in the 50th annual meeting of the Society of Toxicology

(SOT) in Washington, DC. Dr. Stokes and former ICCVAM Chair Dr. Marilyn Wind co-chaired the platform session 

The International Cooperation on Alternative Test Methods (ICATM): Translating Science to Provide Improved Public

Health Safety Assessment Tools. NICEATM-ICCVAM presented posters in sessions titled Alternatives to Mammalian 

Models for Testing, Risk Assessment and Regulatory Policy Applications, Safety and Risk Assessment: Critical 

Characterizations for Chemicals and New Concerns, and Disease Prevention. 

During the SOT meeting, ICATM welcomed a new member, the Korean Center for the Validation of Alternative 

Methods (KoCVAM), at a signing ceremony on March 8 (see http://www.niehs.nih.gov/news/newsletter/2011/april/

science-agreement/index.cfm). ICATM is a voluntary international cooperation of national organizations including 

Canada, the European Union, Japan, South Korea, and the United States. 

Information about NICEATM-ICCVAM activities at the 2011 SOT Meeting can be found at http://iccvam.niehs.nih.

gov/meetings/SOT11/sotablst.htm; more information about ICATM can be found at http://iccvam.niehs.nih.gov/

about/icatm.htm.

On March 29-30, 2011, NICEATM and ICCVAM held a Peer Review Panel Meeting: Evaluation of an In Vitro Estrogen

Receptor Transcriptional Activation Test Method for Endocrine Disruptor Chemical Screening at the William 
H. Natcher Conference Center, Bethesda, MD. The panel was charged to review a draft background review 

document that evaluated the validation status of the B G1Luc estrogen receptor (ER) transcriptional activation (TA) 

test method (also known as the LUMI–CELL® ER test method) for identification of potential endocrine disruptor 

activity. The BG1Luc ER TA test method is an in vitro assay used to identify chemicals that can interact with 

human estrogen receptors. Meeting information is available at http://iccvam.niehs.nih.gov/methods/endocrine/ 

PeerPanel11.htm. 

On August 21-25, 2011, NICEATM-ICCVAM participated in the 8th World Congress on Alternatives and Animal Use in 

the Life Sciences in Montreal, Canada. At the meeting NICEATM staff and ICCVAM members delivered a total of 

20 platform and poster presentations in the themes of Safety and Efficacy Testing of Chemicals, Pharmaceuticals 

and Biologicals; Policy/Law on Animal Use, Public Engagement and Ethics Review; Animal Welfare for Refinement 

and High Quality Science; and Replacement and Reduction in Basic Research. ICCVAM members also chaired or 

co-chaired eight platform presentation sessions. Information about NICEATM–ICCVAM activities at the 8th World 

Congress can be found at http://iccvam.niehs.nih.gov/meetings/8WC/8WCablst.htm. 

In FY 2012, NICEATM-ICCVAM has scheduled the International Workshop on Alternative Methods for Human and 

Veterinary Rabies Vaccine Testing: State of the Science and Planning the Way Forward (http://iccvam.niehs.nih.gov/ 

meetings/RabiesVaccWksp-2011/RabiesVaccWksp.htm) at the U.S. Department of Agriculture (USDA) Center for 

Veterinary Biologics, National Centers for Animal Health, Ames, Iowa. This October 2011 workshop will bring 

together international scientific experts from government, industry, and academia to (1) review the available 

methods and approaches that reduce, refine, and replace animals used in human and veterinary rabies vaccine 

potency testing and (2) develop an implementation strategy to achieve global acceptance and use of these 

alternatives. Workshop information is available at http://iccvam.niehs.nih.gov/meetings/RabiesVaccWksp-2011/ 

RabiesVaccWksp.htm. 

NICEATM–ICCVAM will present seven poster presentations at the 51st annual meeting of the Society of Toxicology 

in March 2012. The posters will report on evaluations of test methods to identify substances with the potential 

to affect endocrine function, cause allergic contact dermatitis, or produce systemic toxicity due to skin 

exposure. Information on NICEATM–ICCVAM presentations at the 2012 SOT meeting will be available at 

http://iccvam.niehs.nih.gov/meetings/SOT12/sotablst.htm. 

In FY 2012, NICEATM-ICCVAM will also convene an International Workshop on Alternative Methods for Leptospira 

Species Vaccine Potency Testing: State of the Science. This workshop will be held at the USDA Center for 

Veterinary Biologics in Ames, Iowa in September 2012. Workshop information will be available at 

http://iccvam.niehs.nih.gov/meetings/LeptoVaccWksp-2012/LeptoVaccWksp.htm 

Recent NICEATM-ICCVAM publications, test methods currently under review, and project status are presented 

in Tables 8, 9, and 10, respectively. 

Table 8. NICEA TM-ICCVAM Publications in FY 2011 

Date Title

October 23, 2010 Safety assessment of allergic contact dermatitis hazards: an analysis supporting reduced animal 

use for the murine local lymph node assay (published in Regulatory Toxicology and Pharmacology) 

February 20, 2011 Informational Brochure: “NICEATM-ICCVAM: Advancing Public Health and Animal Welfare” 

February 20, 2011 Informational Brochure: “Nominations and Submissions to IC CVAM: A Guide for Test Method 

Developers and Sponsors” 

May 18, 2011 Independent Scientific Peer Review Panel Report: Evaluation of the LUMI-CELL ER® (BG1Luc ER TA) 

Test Method. An evaluation of an in vitro estrogen receptor transcriptional activation test method 

for endocrine disruptor chemical screening 

June 30, 2011 ICCVAM Test Method Evaluation Report: Usefulness and Limitations of the Murine Local Lymph Node 

Assay for Potency Categorization of Chemicals Causing Allergic Contact Dermatitis in Humans (NIH 

Publication No. 11-7709) 

July 1, 2011 Using fewer animals to identify chemical eye hazards: revised criteria necessary to maintain 

equivalent hazard classification. (Published in Regulatory Toxicology and Pharmacology) 

NTP Annual Report 2011 37 
38 National Toxicology Program

A list of all NICEATM-ICCVAM publicationsis available at http://ntp-apps.niehs.nih.gov/iccvampb/searchDoc.cfm. 

A list of articles published by NICEATM-ICCVAM in scientific journals is available at 

http://iccvam.niehs.nih.gov/articles/publications.htm.

Table 9. Nominations or Submissions to NICEA TH-ICCVAM in FY 2011

Test Method Nomination or Submission Nominator or Sponsor/Activity Status

Submission of the BoTestTM, BoTestTM Matrix, and BoCellTM BioSentinel Pharmaceuticals/

Botulinum Neurotoxin Activity assays for interlaboratory Final ICCVAM priority: high, in conjunction with a review of other 

validation studies by ICCVAM and NICEATM botulinum toxin test methods

Nomination of an in vitro test method for assessing pyrogenicity Biotest AG/ 

of pharmaceuticals and other products for further evaluation Final ICCVAM priority: high, with a recommendation 

in order to expand its applicability domain to non-endotoxin of further discussion

pyrogens

Table 10. NICEA TH-ICCVAM Recomendations in FY 2011 

Test Method ICCVAM Recommendations/Agency Status

Routine Use of Topical Anesthetics, Systemic Analgesics, 

and Humane Endpoints for Ocular Safety Testing

Pain management procedures should always be used to avoid 

or minimize pain and distress when it is determined necessary 

to conduct the Draize rabbit eye test for regulatory safety

assessments. These procedures include the routine use of topical 

anesthetics, systemic analgesics, and humane endpoints. Agency 

responses received FY 2011. 

http://iccvam.niehs.nih.gov/methods/ocutox/Transmit-2010.htm

Cytosensor Microphysiometer (CM) Ocular Test Method CM can be used as a screening test to identify some types of

substances that may cause permanent or severe eye injuries. 

CM can be used to determine if a limited range of substances 

will not cause sufficient injury to require hazard labeling for 

eye irritation. Agency responses received FY 2011. 

http://iccvam.niehs.nih.gov/methods/ocutox/Transmit-2010.htm

Proposed in vitro testing strategy that uses three in vitro test

methods to assess the eye irritation potential of antimicrobial

cleaning products

Further studies recommended. Agency responses received FY 2011.

http://iccvam.niehs.nih.gov/methods/ocutox/Transmit-2010.htm

Proposed low volume rabbit eye test Proposed low volume rabbit eye test should not be used for 

regulatory testing due to performance issues when compared 

to the current standard rabbit eye test. Agency responses 

received FY 2011. 

http://iccvam.niehs.nih.gov/methods/ocutox/Transmit-2010.htm

Nonradioactive versions of the murine Local Lymph Node Assay 

(LLNA) and expanded applicability domain for the LLNA

Two nonradioactive LLNA methods can be used to identify

substances that may cause allergic skin reactions. The LLNA may 

be used to test most chemicals and products for their potential 

to cause allergic contact dermatitis. Agency responses received

FY 2011. 

http://iccvam.niehs.nih.gov/methods/immunotox/llna.htm

Use of the BG1Luc ER TA test method for endocrine disruptor 

chemical screening

Draft recommendations made available January 24, 2011; 

reviewed by independent peer review panel March 29-30, 2011. 

http://iccvam.niehs.nih.gov/methods/endocrine/end eval.

htm#agencyresponses

Using Fewer Animals to Identify Chemical Eye Hazards: Revised 

Criteria Necessary to Maintain Equivalent Hazard Classification

Draft recommendations made available August 3, 2011

http://iccvam.niehs.nih.gov/ methods/ocutox/reducenum.htm

Report and Recommendations on the Usefulness and Limitations 

of the Murine Local Lymph Node Assay for Potency Categorization

of Chemicals Causing Allergic Contact Dermatitis in Humans

ICCVAM test method recommendations to U.S. Federal agencies 

in June 2011 http://iccvam.niehs.nih.gov/methods/immunotox/

LLNApotency.htm

Further information on the status of nominations and submissions is available at 

http://iccvam.niehs.nih.gov/SuppDocs/submission.htm.
NTP Annual Report 2011 39

NICEATM and ICCVAM prepared, commented on, or otherwise contributed to the development of new test

guidelines, revisions of existing test guidelines, and guidance documents considered by the Organisation for 

Economic Co-operation and Development (OECD). An ICCVAM working group supported by NICEATM developed 

a guidance document for optimizing the value of data generated by in vitro ocular test methods, and an ICCVAM

working group provided comments on draft Test Guideline 488 for transgenic rodent in vivo gene mutation

assays. OECD adopted both of these in 2011. ICCVAM working groups developed or provided comments on test

guidelines or guidance documents currently being considered by OECD in the areas of ocular safety testing, acute 

systemic toxicity, dermal corrosivity and irritation, and endocrine disruptor testing. Dr. Warren Casey, Deputy

Director of NICEATM, is a member of the OECD Validation Management Group for Non-animal Testing (VMG-NA), 

and attended the VMG-NA meeting that took place in FY 2011.

Representatives from NICEATM and ICCVAM participated as liaison members at meetings of the scientific advisor y 

committee of the European Center for the Validation of Alternative Methods that took place in October 2010 and 

February 2011.

Ongoing collaborations on validation studies between NICEATM-ICCVAM and their ICATM partners are

summarized in Table 11.

Table 11 . NICEA TM Participation in International Validation Studies

Test Method Type of Test Lead NICEATM-ICCVAM 

Organization Involvement

BG1Luc estrogen receptor Endocrine disruption NICEATM NICEATM coordinated an international validation 

transcriptional activation study that included participating laboratories in 

test method the United States, European Union, and Japan.

CertiChem MCF-7 cell Endocrine disruption NICEATM NICEATM coordinated an international validation study 

proliferation test method that included participating laboratories in the 

United States, Korea, and Japan.

EpiOcularTM (MatTek) and Ocular irritation ECVAM NICEATM and ICCVAM liaison members served on the

SkinEthicTM (L’oreal) validation management team and provided comments

on study design, chemical selection, and test method 

performance criteria.

Human cryopreserved HepaRG Acute toxicity ECVAM NICEATM and ICCVAM liaison members served on the

and cryopreserved hepatocytes validation management team and provided comments

CYP induction on study design, chemical selection, test method 

test methods protocols, and study reports.

In vitro tests for assessing Allergic contact ECVAM NICEATM and ICCVAM liaison members served on the

skin sensitization potential of dermatitis validation management team and provided comments

chemicals on study design, chemical selection, and test method 

protocols.
40 National Toxicology Program

Nomination, Selection, Evaluation, and Review

Nominations for Study

The NTP maintains a balanced research and testing program that provides data addressing a wide variety of

issues important to public health. The NTP actively seeks to identify and select for study chemicals and other 

substances for which sufficient information is not available to adequately evaluate potential human health

hazards. The NTP accomplishes this goal through a formal open nomination and selection process. Substances

considered appropriate for study generally fall into two broad, yet overlapping, categories:

1. Substances judged to have high concern as a possible public health hazard based on the extent of human 

exposure and/or suspicion of toxicity.

2. Substances for which toxicological data gaps exist and additional studies would help assess potential 

human health risks, e.g., by extrapolating data across species or by evaluating dose-response relationships.

Input is also solicited regarding the nomination of studies that test hypotheses to enhance the predictive ability

of future NTP studies, address mechanisms of toxicity, or fill significant gaps in the knowledge of the toxicity of

classes of chemical, biological, or physical substances. Increased efforts continue to focus on:

• Improving the quality of the nominations of chemicals, environmental agents, or issues for study so that

public health and regulatory needs are addressed.

• Broadening the base and diversity of nominating organizations and individuals.

• Increasing nominations for studying non-cancer toxicological endpoints.

The nomination process is open to the public. The NTP routinely solicits nominations at conferences and 

workshops; through the NTP newsletter, Federal Register notices, NTP website (http://ntp.niehs.nih.gov); and from

interested individuals and groups. Also, NCI, FDA, NIOSH, and NIEHS routinely identify and forward nominations

to the NTP. The NTP also reviews environmental occurrence and human exposure databases and 

the scientific literature to identify substances of potential interest.

Contact Information: Office of Nomination and Selection, Dr. Scott Masten, masten@niehs.nih.gov.

Nomination website: http://ntp.niehs.nih.gov/go/nom

Review and Selection Process

Reviewing and selecting substances and issues nominated for study is a multi-step process (see figure 5 and 

http://ntp.niehs.nih.gov/go/156) that addresses a broad range of concerns from participating representatives 

from the NIEHS, other Federal agencies, NTP BSC (see page 8), NTP Executive Committees’ POCs (see page 12), 

and the public. This multi-step evaluative process provides the NTP with direction and guidance to ensure that 

its testing program addresses toxicological concerns in all areas of public health and that there is balance among

the types of substances selected for study (e.g., industrial chemicals, consumer products, therapeutic agents).

Figure 5 summarizes the study nomination review process, and Table 12 lists the nominations reviewed in 

FY 2011 by the NTP BSC.
Table 12. Research Concepts Reviewed by the BSC in FY2011

Substance [CASRN]

Nominator

Nomination Rationale Study Recommendations

N-Butylbenzene-sulfonamide 

[3622-84-2] 

NIEHS

– High production and use as

a plasticizer in industrial and 

consumer product

– Lack of adequate 

toxicological data

– Suspicion of toxicity based 

on limited toxicity studies 

in rodents and the presence 

of structural features which 

suggestive of potential hazard

– In vitro toxicity studies to assess potential for endocrine 

activity and neurotoxicity 

– ADME and toxicokinetics studies 

– Subchronic toxicity studies including assessments 

of reproductive, developmental, immune and 

neurologic toxicity

– Chronic toxicity and carcinogenicity studies

Drinking water disinfection by- – Co-occurrence in drinking – Mechanistic studies to explore individual and interactive 

products: HMG-CoA reductase water of therapeutic drugs, effects of prenatal exposure to agents that modulate

inhibition and developmental environmental contaminants cholesterol and lipid pathways 

toxicity; Interactive effects and disinfection by-products – A tiered program of mixture studies to include other drinking of antilipidemic agents and with known developmental water contaminants that affect lipid pathways and produce drinking water contaminants and/or reproductive toxicities developmental toxicities by similar but different modes 

in producing developmental – Inadequate data to understand of action

toxicity Private Individual potential hazard of 

mixed exposures

Hydroxyurea – Long term safety concern when – Multigenerational developmental and reproductive toxicity 

[127-07-1] used as therapy for sickle cell studies including neurotoxicity and immunotoxicity 

Private Individual anemia

– NTP CERHR Expert Panel

identified a critical data need for 

multi-generation experimental

animal studies to assess the

long-term effects of prenatal

and postnatal exposures

on postnatal development 

including developmental

neurotoxicity, reproductive 

function, and carcinogenicity

assessments

– Carcinogenicity studies

Isoflavones in soy infant formula 

NIEHS

– Widespread exposure and

concern for potential adverse

effects on human development

– Address certain critical data

gaps and research needs

identified in the NTP CERHR

Expert Panel Report on Soy 

Infant Formula

– Studies to determine the feasibility of direct oral

administration of soy infant formula to rodents

(rats and/or mice) during the period of lactation

– Reproductive development and fertility study in rodents

with soy infant formula and a mixture of isoflavones at the

ratio found in soy infant formula (oral administration to pups

during period of lactation)

– Uterotrophic assay to assess interactions among the

individual isoflavones in soy infant formula

– PK studies to assess metabolism of daidzin to equol during

the period of lactation

NTP Annual Report 2011 41
42 National Toxicology Program

Evaluation

In carrying out its mission, the NTP provides toxicological evaluations on substances of public health concern.

The NTP can initiate bioassays to characterize potential carcinogenicity of only a small fraction of the thousands 

of substances for which there is little or no information. Many more substances are also studied to assess a variety 

of health effects not related to cancer, for example, reproductive and developmental toxicities, immunotoxicity, 

neurotoxicity, and genotoxicity. Other biological parameters are often assessed to understand susceptibility

to toxic substances, such as the absorption, distribution, metabolism, and excretion (ADME) of substances

biochemical markers that indicate exposures, and genetic changes in enzymes that metabolize drugs.

An NTP project review committee evaluates a study’s project plan (e.g., design, methods, hypothesis, etc.). 

The toxicological evaluation for carcinogenicity generally involves repeated administration of a substance to

groups of laboratory animals for up to two years. Many short-term, subchronic studies are designed to provide 

dose-setting information for longer, chronic exposure studies and to address specific gaps in the toxicology

database. The adverse health effects from short- or long-term exposures to different dose levels of a substance 

are examined by observation, histopathology, and several toxicology endpoints, comparing them with control 

groups of animals not exposed to the substance. Many substances are also studied using protocols specifically

designed to address how the substance causes particular toxic effects. The NTP has specific requirements that

testing laboratories comply with the Animal Welfare Act of 1966 and adhere to the principles in the Guide for the 

Care and Use of Laboratory Animals (NRC, 2011). General information about the objectives and procedures of NTP 

study protocols is available on the NTP website (http://ntp.niehs.nih.gov/go/type). Current testing status can be

found at http://ntp.niehs.nih.gov:8080/index.html?col=010stat.

The NTP carries out toxicology and carcinogenesis testing through two primary mechanisms: laboratory studies 

conducted in contract laboratories (Table 13) and studies conducted via IAGs at agencies (see page 68). In

addition to toxicology research on compounds and exposures, the NTP supports developing new techniques

and methods to improve the ability to identify and assess potential environmental toxicants and to develop 

and validate novel and alternative testing methods that will reduce, replace, or refine animal use. The NTP also

supports developing improved statistical methods for designing and evaluating the results of toxicology studies.

Table 13. NTP Contracts That Supported NTP Testing Activities in FY 2011

Description Contractor

ADME Chemical Disposition Lovelace Biomedical

Archives and Specimen Repository Experimental Pathology Labs

Bioinformatics Methylation Project Murdock Research Institute

Cell Phone Radiation IIT Research Institute

Chemistry Support Midwest Research Institute

Chemistry Support Research Triangle Institute

Chemistry Support Battelle Memorial

Collaboration Between BPA Grantees and NTP Professional and Scientific

Collaborative Work Ramazzini Institute

Environmental and Therapeutic Agents Virginia Commonwealth University

Evaluation of Test Agents in Laboratory Animals Battelle Memorial

Evaluation of Test Agents via Inhalation Exposure Battelle Memorial

Evaluate Toxicological Potential of Test Agents Battelle Memorial

Evaluate Toxicological Potential of Test Agents Southern Research
NTP Annual Report 2011 43

Description Contractor

Genetic Toxicity in Bacteria and Rodents Integrated Laboratory Systems

Investigative Research Support Contract Integrated Laboratory Systems

NTP Computing and CEBS Support Vistronix, Inc.

NTP Information Systems Support SciMetrika

NTP Statistical and Computer Support SRA International

NTP Technical Reports Preparation Service Biotechnical Services

Pathology Support Charles River

Pathology Support Integrated Laboratory Systems

Provantis Software for NTP INSTEM LSS

Quality Assessment/Pathology Support Experimental Pathology Labs

Quality Assessment Reports for Audits and Inspections Dynamac

Reproductive Assessments by Continuous Breeding Research Triangle Institute

Sperm Count Vaginal Cytology Evaluation Research Triangle Institute

Support/Consultation for Provantis Software Labscience, Inc.

Production of B6C3F1 Mice Taconic Farms (2)

Review and Dissemination

The results of toxicology and carcinogenesis studies undergo rigorous peer review and are published in several 

NTP report series:

• Technical Reports (TR). This series presents the results of long-term, generally 2-year, toxicology and 

carcinogenicity studies, typically conducted in rats and mice. Results of genetic toxicology, ADME, and 

toxicokinetic studies are often included in the reports. The draft reports are reviewed by ad hoc expert

panels (http://ntp.niehs.nih.gov/go/36144).

• NTP Toxicity (TOX) Reports. TOX reports are prepared for studies where the substance exposure period 

is short term, generally up to 13 weeks. Draft reports are typically peer reviewed by letter review.

• Genetically Modified Models(GMM) Reports. NTP began the GMM report series in May 2003. 

This report series presents the results of substances evaluated by NTP in transgenic mouse strains 

(e.g., p53+/- heterozygous and Tg.AC mice).

Abstracts of the TR, TOX, and GMM series are posted on the NTP website, and PDF files of completed reports are

available at the NTP website (http://ntp.niehs.nih.gov/go/reports) and are also catalogued in PubMed. Pathology 

data from the NTP rodent studies included in these reports undergo several reviews. The final review is by a 

pathology working group (PWG), a panel of experts convened by the NTP to review the microscopic evaluations.

After the NTP PWG review, pathology tables and body weight and survival graphs for the completed studies 

are made publicly available (http://ntp.niehs.nih.gov/go/peerreview) until draft study reports are completed for 

peer review. When the draft reports are completed for peer-review, the abstracts are put on the NTP website 

with the list of reports. The pathology tables and the body weight and survival graphs are accessible at the end

of the abstract text. Following peer review, the NTP finalizes the report, posts it on its website (http://ntp.niehs.

nih.gov/go/reports) and provides electronic linksto the final pathology tables and the curves for body weight

and survival. Study summaries for other types of studies, such as immunotoxicity, developmental toxicity, and 

reproductive toxicity studies, are also available on the “NTP Study Reports” page on the website. New criteria 

for immunotoxicity, developmental, and reproductive studies were finalized in 2009 and NTP will publish 

these studies as technical reports following peer review. All types of NTP studies may also be published 

in peer-reviewed scientific journals.
44 National Toxicology Program

Chronic Toxicity/Carcinogenicity Studies

In the area of general toxicology assessments, the scope and types of studies performed are dictated mainly 

by the data needs for the specific substance being studied. General toxicology studies usually fall into two 

categories: subchronic or pre-chronic studies, and 2-year chronic toxicology and carcinogenicity studies. 

Two-year studies in rodents are a method by which chemicals or physical agents are identified as having 

the potential to be hazardous to humans.

The chronic toxicology and carcinogenicity studies in conventional rodent models generally use both male and 

female rats (Fischer 344/N, Harlan Sprague Dawley, or Wistar Han) and mice (B6C3F1 hybrid), with three exposure

levels plus untreated controls, in groups of 50 animals, for two years; other rodent models(e.g., genetically 

modified mice) are used as needed. If adequate data exist in the literature for one rodent species (rats or mice),

then typically only the remaining species is studied. The NTP interfaces its testing with regulatory agencies and 

the private sector to minimize duplication of effort. Studies ongoing, initiated, completed, and published in FY

2011 are listed in Tables 14, 15, 16, and 17, respectively. 

The NTP describes the results of individual experiments on a chemical agent and notes the strength of evidence 

for conclusions of each study. Negative results, in which the study animals do not have a greater incidence of

neoplasia than do control animals, do not necessarily mean that a chemical is not a carcinogen, because the 

experiments are conducted under a limited set of conditions. Positive results demonstrate that a chemical is

carcinogenic for laboratory animals under the conditions of the study and indicate that exposure to the 

chemical has the potential to be hazardous to humans.

Table 14. Chronic Toxicity/Carcinogenicity Studies Ongoing During FY 2011

Study CASRN Species/Strain Study Route Length Project Leader

Aging cohort study - 129/SvlmJ mouse Mice: 129S1/SvImJ N/A 2 years French

Aging cohort study - B6C3F1J mouse Mice: B6C3F1 (Jackson) N/A 2 years French

Aging cohort study - C3H/HeJ mouse Mice: C3H/HeJ N/A 2 years French

Aging cohort study - C57/BL/6J 

mouse

Mice: C57BL/6J (Jackson) N/A 2 years French

Aging cohort study - CAST/EiJ mouse Mice: CAST/EiJ 

(M. m. castaneus)

N/A 2 years French

Aging cohort study - 

NZO/HiLtJ mouse

Mice: NZO/HiLtJ N/A 2 years French

Aging Cohort Study - 

PWK/PhJ mouse

Mice: PWK/PhJ N/A 2 years French

Aging cohort study - 

WSB/EIJ mouse

Mice:WSB/EiJ 

(M. m. domesticus)

N/A 2 years French

Aging cohort study - A/J mouse Mice: A/J N/A 2 years French

Aging cohort study - NOD. 

B10Sn-H2(b)/J

Mice: NOD. B10Sn-H2(b)/J N/A 2 years French

Antimony trioxide 1309-64-4 Rats: Wistar Han 

Mice: B6C3F1/N

Inhalation 2 years Stout

3’-Azido-3’-deoxythymidine (AZT)* 30516-87-1 Mice: C3B6F1-+/

TRP53<TM1BRD> (NCTR)

Gavage 9 months Leakey

AZT/Drug Combinations

Transplacental/Neonatal Study

Mice: B6C3F1/NCTR BR

(C57BL/6N x C3H/HEN MTV-)

Gavage 2 years Beland

AZT/Drug Combinations

Transplacental Carcinogenesis Study

Mice: B6C3F1/NCTR BR

(C57BL/6N x C3H/HEN MTV-)

In Utero 2 years Beland

2,3-Butanedione (diacetyl) 431-03-8 Rats: Wistar Han 

Mice: B6C3F1/N

Inhalation 2 years Morgan
NTP Annual Report 2011 45

Study CASRN Species/Strain Study Route Length Project Leader

Cobalt 7440-48-4 Rats: F344/NTac 

Mice: B6C3F1/N

Inhalation 2 years Hooth

Dibutyl phthalate 84-74-2 Rats: Harlan SD

Mice: B6C3F1/N

Feed 2 years Blystone

N,N-Dimethyl-p-toluidine 99-97-8 Rats: F344/N

Mice: B6C3F1/N

Gavage 2 years Dunnick

Furan 110-00-9 Rats: F344 (NCTR) Gavage 2 years Beland

Ginkgo biloba extract 90045-36-6 Rats: F344/N

Mice: B6C3F1/N

Gavage 2 years Chan

Glycidamide 5694-00-8 Rats: F344 (NCTR)

Mice:B6C3F1/NCTR BR

(C57BL/6N x C3H/HEN MTV-)

Water 2 years Beland

Green tea extract Rats: Wistar Han 

Mice: B6C3F1/N

Gavage 2 years Chan

Indole-3-carbinol 700-06-1 Rats: Harlan SD

Mice: B6C3F1/N

Gavage 2 years Wyde

Metal working fluids (CIMSTAR 3800) Rats: Wistar Han 

Mice: B6C3F1/N

Inhalation 2 years Morgan

Metal working fluids (Trim VX) Rats: Wistar Han 

Mice: B6C3F1/N

Inhalation 2 years Morgan

Pentabromodiphenyl oxide 

(technical) (DE 71)

32534-81-9 Rats: Wistar Han 

Mice: B6C3F1/N

Gavage 2 years Dunnick

Perfluorooctanoic acid (PFOA) 335-67-1 Rats: Harlan SD Feed 2 years Blystone

beta-Picoline 108-99-6 Rats: F344/N

Mice: B6C3F1/N

Water 2 years Wyde

Pyrogallol 87-66-1 Rats: F344/N

Mice: B6C3F1/N

Topical 

Application

2 years MercadoFeliciano

Tetrabromobisphenol A 79-94-7 Rats: Wistar Han 

Mice B6C3F1/N

Gavage 2 years Dunnick

Trimethylolpropane triacrylate 15625-89-5 Rats: F344/N

Mice: B6C3F1/N

Topical 

Application

2 years Surh / Chhabra

Tripelennamine hydrochloride 154-69-8 Rats: F344/N

Mice: B6C3F1/N

Feed 11 months Jackson

Vinylidene chloride 75-35-4 Rats: F344/N

Mice: B6C3F1/N

Inhalation 2 years Wyde

Water disinfection byproducts

(bromodichloroacetic acid)

71133-14-7 Rats: F344/NTac 

Mice: B6C3F1/N

Water 2 years Hooth

Zinc carbonate, basic 5263-02-5 Rats: Harlan SD Feed 2 years Wyde

* Indicates study conducted using genetically-modified model

Table 15. Chronic Toxicity/Carcinogenicity Studies initiated during FY 2011

Study CASRN Species/Strain Study Route Length Project Leader

p-Chloro-a,a,a-trifluorotoluene 98-56-6 Rats: Harlan SD

Mice: B6C3F1/N

Inhalation 2 years Stout

Di(2-ethylhexyl) phthalate 117-81-7 Rats: Harlan SD Feed 2 years Foster

2-Hydroxy-4-methoxybenzophenone 131-57-7 Rats: Harlan SD 

Mice: B6C3F1/N

Feed 2 years Auerbach

Insertional mutagenesis – 

definitive vector study

Mice: C57BL/6 Intravenous 14 months Germolec
46 National Toxicology Program

Table 16. Technical Reports Completed in FY 2011

Chemical/Exposure – Study Type Technical

Report Number/

[CASRN]

Use Levels of Evidence of Carcinogenic Activity

Male Rats Female Rats Male Mice Female Mice

Acrylamide

Drinking water–

toxicology and carcinogenesis

TR-575

[79-06-1]

Polyacrylamides and dye 

intermediate; flocculants; 

waste and water treatment; 

paper and pulp industry. 

Associated with tobacco: 

reported either as a natural

component of tobacco, 

pyrolysis product (in tobacco

smoke), or additive for one 

or more types of tobacco

products.

 Clear 

Evidence

 Clear 

Evidence

 Clear 

Evidence

 Clear 

Evidence

Aloe vera whole leaf

extract (native)

Drinking watertoxicology and carcinogenesis

TR-577 Recognized as a therapeutic

dermatologic agent; extracts

incorporated in a variety of

topical health care products

and cosmetics.

 Clear 

Evidence

 Clear 

Evidence

 No 

Evidence

 No 

Evidence

AIDS Therapeutics:

3’-Azido-3’-Deoxythymidine

(AZT), Lamivudine (3T3),

Nevirapine (NVP), and 

Nelfinavir Mesylate (NFV)

Transplacental – 

toxicology and carcinogenesis

TR-569

[30516-87-1]

[134678-17-4]

[129618-40-2] 

[159989-65-8]

Used in the treatment of AIDS. N/A N/A No evidence of

carcinogenic activity of

AZT in male B6C3F1 mice

and equivocal evidence 

of carcinogenic activity in

female B6C3F1 mice

 No evidence of

carcinogenic activity of

mixtures of AZT and 3TC 

in male B6C3F1 mice and 

equivocal evidence of

carcinogenic activity in

female B6C3F1 mice

 Some evidence of

carcinogenic activity of

mixtures of AZT, 3TC, and 

NVP in male B6C3F1 mice 

and equivocal evidence 

of carcinogenic activity

in female B6C3F1 mice

 No evidence of

carcinogenic activity

of mixtures of AZT, 3TC,

and NFV in male or female 

B6C3F1 mice

Kava kava extract 

Gavage – 

toxicology and carcinogenesis

TR-571

[9000-38-8]

Herbal supplement 

alternative to anti-anxiety

drugs, used to help children 

with hyperactivity and as a 

skin-conditioning agent 

in cosmetics.

 Equivocal 

Evidence

 No 

Evidence

 Clear

Evidence

 Some

Evidence
NTP Annual Report 2011 47

Chemical/Exposure – Study Type Technical

Report Number/

[CASRN]

Use Levels of Evidence of Carcinogenic Activity

Male Rats Female Rats Male Mice Female Mice

Methyl trans-styryl ketone

Feed and dermal – 

toxicology and carcinogenesis

TR-572

[1896-62-4]

Reactive carbonyl 

compound used in organic 

synthetic reactions such

as intermediate in organic 

synthesis, electroplating 

chemical; pharmaceutical

intermediate, biochemical 

reagent, agricultural chemical

intermediate, proposed

sunscreen intermediate. 

Used as flavoring and 

fragrance additive. Associated

with tobacco: reported either

as a natural component of

tobacco, pyrolysis product 

(in tobacco smoke), or additive 

for one or more types of

tobacco products.

 No 

Evidence

 No 

Evidence

 No 

Evidence

 No 

Evidence

All-trans-retinyl palmitate

Simulated solar light and 

topical application study – 

photocarcinogenesis

TR-568

[79-81-2]

Retinol compound (Vitamin A) 

used in skin products.

Topical treatment of SKH-1 mice with the control cream 

resulted in earlier onsets of in-life skin lesions and higher

incidences and multiplicities of in-life skin lesions, when 

compared to untreated controls, in the absence and presence 

of simulated solar light (SSL). The topical treatment of SKH-1 

mice with control cream resulted in higher incidences and 

multiplicities of squamous cell neoplasms of the skin when 

compared to untreated controls in the absence and presence 

of SSL. Compared to the control cream, retinoic acid further 

enhanced the effects of SSL in SKH-1 mice based upon earlier 

onsets and increased multiplicities of in-life skin lesions. 

Compared to the control cream, retinyl palmitate further 

enhanced the effects of SSL in SKH-1 mice based upon earlier 

onsets and increased multiplicities of in-life skin lesions. 

Compared to the control cream, retinyl palmitate further 

enhanced the photocarcinogenic activity of SSL in SKH-1 

mice based upon increased incidences and multiplicities 

of squamous cell neoplasms of the skin.

*Senna (powdered)

Feed – 

toxicology and carcinogenesis

GMM-15

[8013-11-4]

Herbal product, extract of

senna fruit, used in laxative

preparations.

N/A N/A No 

Evidence

 No 

Evidence

alpha/beta Thujone mixture

Gavage – 

toxicology and carcinogenesis

TR-570

[76231-76-0]

The essential oils derived from

natural oil of cedar leaves

in which thujone occurs are

used in herbal medicine and 

as flavorings, fragrances, and 

rodent and mite repellents. 

Thujone is banned as a direct

food additive in the U.S.

Thujone-containing plant 

oils are used as flavoring

substances in the alcoholic 

drink industry.

 Some

Evidence

 No 

Evidence

 No 

Evidence

 No 

Evidence

*Indicates study conducted using genetically-modified model
48 National Toxicology Program

Table 17. Technical Reports Published in FY 2011

Chemical /Exposure 

Study Type

Technical

Report Number/

[CASRN]

Use Levels of Evidence of Carcinogenic Activity

Male Rats Female Rats Male Mice Female Mice

1-Bromopropane

Inhalation – 

toxicology and carcinogenesis

TR-564

[106-94-5]

Laboratory reagent;

replacement chemical for 

chlorinated solvents; metal 

cleaning and degreasing; and 

adhesives.

 Some

Evidence

 Clear 

Evidence

 No 

Evidence

 Clear 

Evidence

bis(2-Chloroethoxy)methane 

Topical – 

toxicology and carcinogenesis

TR-536

[111-91-1]

Used as a starting compound

to produce polysulfide

elastomers and as a solvent.

 No 

Evidence

 No 

Evidence

 No 

Evidence

 No 

Evidence

Ginseng

Gavage – 

toxicology and carcinogenesis

TR-567 

[50647-08-0]

Used as herbal remedy, dietary 

supplement, food additive,

and in cosmetics.

 No 

Evidence

 No 

Evidence

 No 

Evidence

 No 

Evidence

Milk thistle extract 

Feed – 

toxicology and carcinogenesis

TR-565

[84604-20-6]

Used as an herbal supplement

and medicinally as a 

hepatoprotectant. Used

for treatment of chronic 

inflammatory liver disorders.

 No 

Evidence

 No 

Evidence

 No

Evidence

 No

Evidence

-Myrcene

Gavage – 

toxicology and carcinogenesis

TR-557

[123-35-3]

Intermediate in the 

commercial production of

terpene alcohols that serve 

as intermediates for the 

production of large-volume

aroma and flavor chemicals. 

Used as scenting agents

in cosmetics, soaps, and 

detergents; as a peripheral

analgesic substance; and 

as the active ingredient in

lemongrass tea. Identified in

over 200 plants and detected

in emissions of plywood

veneer dryers.

 Clear 

Evidence

 Equivocal 

Evidence

 Clear 

Evidence

 Equivocal 

Evidence

2,3’,4,4’,5-

Pentachlorobiphenyl 

(PCB 118) 

Gavage – 

toxicology and carcinogenesis

TR-559

[31508-00-6]

PCBs and their mixtures 

including PCB 118 were

produced commercially before

1977 for the electric industry as

dielectric insulating fluids for 

transformers and capacitors. 

One of 2 dioxin-like chemicals 

being tested to determine if

the toxic equivalency factors 

(TEFs) for these chemicals 

would be predictive of rodent 

carcinogenicity.

N/A Clear 

Evidence

N/A N/A
Chemical /Exposure 

Study Type

Technical

Report Number/

[CASRN]

Use Levels of Evidence of Carcinogenic Activity

Male Rats Female Rats Male Mice Female Mice

Pulegone

Gavage – 

toxicology and carcinogenesis

TR-563

[89-82-7]

Major constituent of 

oil of pennyroyal. Used for 

flavoring foods and drinks, 

as a fragrance agent, and 

as an herbal medicine.

 No 

Evidence

 Clear 

Evidence

 Clear 

Evidence

 Clear 

Evidence

3,3’,4,4’-

Tetrachloroazobenzene 

(TCAB)

Gavage – 

toxicology and carcinogenesis

TR-558

[14047-09-7]

Impurity in dichloroaniline 

(DCA) and herbicides derived

from DCA. Formed as an

unwanted byproduct in the 

manufacture of 3,4-DCA 

and its herbicidal derivatives

Propanil®, Linuron®, and 

Diuron®. Environmental 

contamination by 3,3’,4,4’-

TCAB occursfrom the 

degradation of chloranilide

herbicides and the photolysis 

and biolysis of 3,4-DCA.

 Clear 

Evidence

 Clear 

Evidence

 Clear 

Evidence

 Clear 

Evidence

Tetralin 

Inhalation – 

toxicology and carcinogenesis

TR-561

[119-64-2]

Solvents in paints, waxes,

and polishes derived from

naphthalene. Insecticide.

 Some

Evidence

 Some

Evidence

 No 

Evidence

 Equivocal 

Evidence

Only summaries of carcinogenic activity conclusions are included in the tables. Complete information is available in the full

study reports found at http://ntp.niehs.nih.gov/. The NTP anticipates that seven Technical Reports will undergo peer review

in FY 2012, as shown in Table 18.

Table 18. Technical Reports Expected to Undergo Peer Review in FY 2012

Chemical Technical Use

Report Number/

[CASRN]

Ginkgo biloba extract TR-578 Herbal supplement.

[90045-36-6]

-Picolene TR-580 Used as a solvent in the synthesis of pharmaceuticals, resins, dyes, and rubber accelerators and 

[108-99-6] as a lab reagent. Also used as an intermediate in the manufacture of insecticides, waterproofing 

agents, niacin and niacinamide.

Pyrogallol TR-574 Naturally occurring in food. Used as a modifier in oxidation dyes including hair dyes and 

[87-66-1] colors; as developer in photography; as a mordant for dying wool; as a reagent for antimony

and bismuth; as a reducer for gold, silver and mercury salts; for process engraving; for making

colloidal solutions of metals; and in the manufacture of pharmaceuticals and pesticides.

N,N-Dimethyl-p-toluidine TR-579 Used as a polymerization accelerator for the manufacture of bone cements and dental 

[99-97-8] materials. Found in industrial glues and artificial fingernail preparations. Intermediate in dye 

and pesticide synthesis.

AZT* GMM-14 Pyrimidine nucleoside analog with antiviral activity used in the treatment of AIDS.

[30516-87-1]

Mixtures AZT, 3TC, and NVP* GMM-14 Special combination to study AIDS therapeutics.

[30516-87-1]

[134678-17-4] 

[129618-40-2]

Trimethylolpropane triacrylate [15625-89-5] Used in the production of ultraviolet-curable inks, electron beam irradiation-curable coatings, 

and polymers and resins; as a component of photopolymer and flexographic printing plates

and photoresists; and as an ingredient in acrylic glues and anaerobic sealants. Also used in

paper and wood impregnates, wire and cable extrusion, polymer-impregnated concrete, 

and polymer concrete structural composites.

*Indicates study conducted using genetically-modified model

NTP Annual Report 2011 49
50 National Toxicology Program

General Toxicology Studies

The NTP performs prechronic toxicity studies to address specific deficiencies in toxicology databases for 

chemicals, such as an understanding of toxicity with repeated exposures; to identify target organs for more 

in depth systems toxicology evaluations and mechanistic studies; and to provide dose-setting information 

for possible chronic studies.

Although designs are flexible, prechronic studies usually involve exposures of rats and mice of both sexes 

to substances for periods of 14-90 days. Assessments almost always include tissue histopathology, clinical 

pathology, and sperm motility or measurements of estrous cycle length. The frequency of malformed red blood

cells (micronucleated erythrocytes, a measure of chromosomal damage) is determined as an in vivo measure

of genotoxic potential.

The study protocol may include more detailed or focused studies when findings published in the existing

scientific literature or identified in initial NTP studies suggest a target organ or system. The study protocol may 

include separate studies of reproductive, genetic, or immunological toxicity based on the outcome of the toxicity

screens and may use additional endpoints to improve our understanding of the mechanisms and modes of action

of a chemical.

In some cases, the NTP uses alternative models, including genetically modified mouse models and non-mammalian

models, for prechronic studies. Such studies are presented in the section “Genetic and Alternative Test Systems” 

(page 62). Tables 19-22 list the toxicity studies that were ongoing, initiated, completed, and published respectively, 

during FY 2011. Information is available at http: //ntp.niehs.nih.gov/go/reports. Table 23 lists toxicity studies 

planned for FY 2012.

Table 19. Ongoing Toxicity Studies during FY 2011

Study CASRN Species/Strain Study Route Length Project Leader

3’-Azido-3’-deoxythymidine (AZT)* 30516-87-1 Mice: P53 +/- (FVB/N) Gavage 9 months Leakey

BPA 80-05-7 Rats: SD (NCTR) Gavage 13 weeks Delclos

Cell phone radiation (CDMA) Rats: Harlan SD

Mice: B6C3F1

Whole-body 

exposure

5 days 

49 days 

28 day

Wyde

Cell phone radiation (GSM) Rats: Harlan SD

Mice: B6C3F1

Whole-body 

exposure

5 days 

49 days 

28 day

Wyde

bis(2-Chloroethoxy)methane 111-91-1 Mice: C57BL/6J (Jackson) 

Mice: C3H/HeJ 

Mice: B6C3F1 (Jackson)

Gavage 3 days, 

10 days

Dunnick

Diet evaluation study Mice: CD-1 Reg.[Crl:CD1(ICR)], Feed 13 weeks Delclos

Ephedrine + caffeine combination 299-42-3 

58-08-2

Mice: C57BL/6J (Jackson) 

Mice: C3H/HeJ

Mice: B6C3F1 (Jackson)

Gavage 3 days, 

10 days

Dunnick

Glucosamine hydrochloride + 

chondroitin sulfate

66-84-2 

9007-28-7

Rats: Zucker - Obese 

(HsdHlr: ZUCKER-Leprfa)

Rats: Zucker - Lean 

(HsdHlr: ZUCKER-Lepr+)

Gavage 13 weeks Leakey

1020 Long multiwalled CNTs Rats: Harlan SD

Mice: B6C3F1/N

Inhalation 30 days Walker

Melamine + cyanuric acid

combination

108-78-1

108-80-5

Rats: F344 Gavage 13 weeks Gamboa de

Costa

Nanoscale Ag 7440-22-4 Rats: SD Gavage 13 weeks Walker
NTP Annual Report 2011 51

Study CASRN Species/Strain Study Route Length Project Leader

Serotype 2 Adeno-associated Viral 

Vector hAQP1 (rAAV2hAQP1)

Mice: BALB/c Intraductal 

cannulation

13 weeks Germolec

Triclosan 3380-34-5 Mice: B6C3F1/N Dermal 13 weeks Fang

Usnea lichen Rats: F344

Mice: B6C3F1/NCTR BR

(C57BL/6N x C3H/HEN MTV-)

Feed 2 weeks 

13 weeks

Leakey

(+)-Usnic Acid 7562-61-0 Rats: F344

Mice: B6C3F1/NCTR BR

(C57BL/6N x C3H/HEN MTV-)

Feed 2 weeks 

13 weeks

Leakey

Vincamine 1617-90-9 Rats: Harlan SD

Mice: B6C3F1/N

Gavage 2 weeks Chan

* Indicates study conducted using genetically-modified model

Table 20. Toxicity Studies Initiated during FY 2011

Study CASRN Species/Strain Study Route Length Project Leader

2-Aminopyridine 504-29-0 Rats: F344/NTac 

Mice: B6C3F1

Gavage 14 days Dunnick

3-Aminopyridine 462-08-8 Rats: F344/NTac 

Mice: B6C3F1

Gavage 14 days Dunnick

4-Aminopyridine 504-24-5 Rats: F344/NTac 

Mice: B6C3F1

Gavage 14 days Dunnick

Comparison study of

aminopyridines/troponin levels

504-29-0

462-08-8

504-24-5

Rats: F344/NTac 

Mice: B6C3F1

Gavage 1 and 4 

hours

Dunnick

Cell phone radiation (CDMA) Rats: Harlan SD

Mice: B6C3F1

Whole-body 

exposure

5 days 

49 days 

28 day

Wyde

Cell phone radiation (GSM) Rats: Harlan SD

Mice: B6C3F1

Whole-body 

exposure

5 days 

49 days 

28 day

Wyde

Diet evaluation study Mice: CD-1 Reg.[Crl:CD1(ICR)] Feed 13 weeks Delclos

1020 Long multiwalled CNTs Rats: Harlan SD

Mice: B6C3F1/N

Inhalation 30 days Walker

Melamine + cyanuric acid

combination

108-78-1

108-80-5

Rats: F344 Gavage 13 weeks Gamboa de

Costa

Microcystin-LA (TGMX) 96180-79-9 Rats: Wistar Han Intravenous 1/2/6/24 

hours

Walker

Microcystin mixture (TGMX) 96180-79-9

101043-37-2

Rats: Wistar Han Intravenous 1/2/6/24 

hours

Walker

Nanoscale Ag 7440-22-4 Rats: SD Gavage 13 weeks Walker

Pyridine 110-86-1 Mice: B6C3F1 Gavage 14 days Dunnick

Serotype 2 adeno-associated viral 

vector hAQP1 (rAAV2hAQP1)

Mice: BALB/c Intraductal 

cannulation

13 weeks Germolec

Triclosan 3380-34-5 Mice: B6C3F1/N Dermal 13 weeks Fang 

Vincamine 1617-90-9 Rats: Harlan SD

Mice: B6C3F1

Gavage 2 weeks Chan
52 National Toxicology Program

Table 21. Toxicity Studies Completed during FY 2011

Study CASRN Species/Strain Study Route Length Project Leader

3-Aminopyridine 462-08-8 Rats: F344/NTac 

Mice: B6C3F1

Gavage 14 days Dunnick

2-Aminopyridine 504-29-0 Rats: F344/NTac 

Mice: B6C3F1

Gavage 14 days Dunnick

4-Aminopyridine 504-24-5 Rats: F344/NTac 

Mice: B6C3F1

Gavage 14 days Dunnick

Comparison study of

aminopyridines/troponin levels

462-08-8

504-29-0

504-24-5

Rats: F344/NTac 

Mice: B6C3F1

Gavage 1 and 4 

hours

Dunnick

Black cohosh 84776-26-1 Mice: B6C3F1 Gavage 13 weeks MercadoFeliciano

Insertional mutagenesis 

(radiation levels)

Mice: B6.SJL-Ptprc[a] 

Pepc[b]/BoyJ

Whole-body 

Exposure

8 weeks Germolec

Microcystin-LA (TGMX) 96180-79-9 Rats: Wistar Han Intravenous 1/2/6/24 

hours

Walker

Microcystin-LR (TGMX) 101043-37-2 Rats: Wistar Han Intravenous 1/2/6/24 

hours

Walker

Microcystin mixture (TGMX) 96180-79-9

101043-37-2

Rats: Wistar Han Intravenous 1/2/6/24 

hours

Walker

2,3-Pentanedione 600-14-6 Rats: Wistar Han 

Mice: B6C3F1/N

Inhalation 13 weeks Morgan

Pregnancy rate comparison study Rats: Harlan SD 

(Indianapolis Facility) 

Rats: Harlan SD 

(Dublin Facility)

N/A 16 weeks Vallant

Pyridine 110-86-1 Mice: B6C3F1 Gavage 14 days Dunnick

QT drugs (bepridil hydrochloride) 74764-40-2 Dog: Beagles Oral 

(capsule)

1 day Hooth

QT drugs (diltiazem hydrochloride) 33286-22-5 Dog: Beagles Oral 

(capsule)

1 day Hooth

QT drugs (Loratadine) 79794-75-5 Dog: Beagles Oral 

(capsule)

1 day Hooth

QT drugs (Lovastatin) 75330-75-5 Dog: Beagles Oral 

(capsule)

1 day Hooth

QT drugs (sotalol hydrochloride) 959-24-0 Dog: Beagles Oral 

(capsule)

1 day Hooth

QT drugs (terfenadine) 50679-08-8 Dog: Beagles Oral 

(capsule)

1 day Hooth

Sodium tungstate, dihydrate 10213-10-2 Rats: Harlan SD

Mice: B6C3F1/N

Water 13 weeks Hooth

Tris(2-chloroisopropyl)phosphate 13674-84-5 Rats: Harlan SD

Mice: B6C3F1/N

Feed 13 weeks Stout
Table 22. Toxicity Studies Published during FY 2011

Chemical/Exposure – Study Type Toxicity 

Report 

Number/

CASRN

Use Evidence of Carcinogenic Activity

Estragole 

Gavage – Toxicity Studies

TOX-82

140-67-0

Used in perfumes and as flavor 

in foods and liquors; as an

antimicrobial agent against 

acid-tolerant food microflora. 

Found naturally in tarragon,

basil, and fennel.

Under the conditions of these 3-month 

studies, estragole showed carcinogenic 

activity based on the occurrence of two 

cholangiocarcinomas and one hepatocellular

adenoma in the liver of three of 10 male

F344/N rats in the high dose group. Because 

rats and mice were exposed for only 

3 months, these studies do not access the 

full carcinogenic potential of estragole.

Nonneoplastic effects were observed in the 

liver, glandular stomach, nose, kidney, and 

salivary gland of male and female rats and in

the testes, epididymides, and pituitary gland 

of male rats. Nonneoplastic effects were also

observed in the liver and nose of male and 

female mice and in the stomach of 

female mice.

2,4-Decadienal 

Gavage – Toxicity Studies

TOX-76

25152-84-5

Synthetic flavoring and 

fragrance material; also

evaluated as a corrosion 

inhibitor for steel in oil field

operations. 

2,4-decadienal administration caused

decreased body weights and increased 

incidences of forestomach lesions in the 

3-month studies in rats and mice. In addition,

treatment-related lesions of the olfactory 

epithelium were observed in male rats and 

male and female mice. The no-observedadverse-effect level was determined to be

100 mg/kg in rats and mice. 2,4-Decadienal

was not mutagenic in vitro or in vivo.

Table 23. Toxicity Studies Planned for FY 2012*

Study CASRN Species/Strain Study Route Length Project Leader

(4-Chloro-6-(2,3-xylidino)-2-

pyrimidinylthio) acetic acid 

(WY-14643)

50892-23-4 Rats: Harlan SD Gavage 28 days Blystone

Diet evaluation study Mice: CD-1 Reg.[Crl:CD1(ICR)] Feed 13 weeks Delclos

Perfluorohexane-1-sulphonic acid - 

potassium salt

3871-99-6 Rats: Harlan SD Gavage 28 days Blystone

1-Perfluorobutanesulfonic acid 375-73-5 Rats: Harlan SD Gavage 28 days Blystone

Perfluorodecanoic acid (PFDA) 335-76-2 Rats: Harlan SD Gavage 28 days Blystone

Perfluorohexanoic acid (PFHXA) 307-24-4 Rats: Harlan SD Gavage 28 days Blystone

Perfluorononanoic acid (PFNA) 375-95-1 Rats: Harlan SD Gavage 28 days Blystone

Perfluorooctane Sulfonate, (PFOS) 1763-23-1 Rats: Harlan SD Gavage 28 days Blystone

Perfluorooctanoic acid (PFOA) 335-67-1 Rats: Harlan SD Gavage 28 days Blystone

* Known test articles as of 10/1/2011. Others may be scheduled as protocols are finalized.

NTP Annual Report 2011 53
54 National Toxicology Program

Mutagenesis and Genetic Toxicity

Genetic toxicity test results are used to help interpret toxicity, carcinogenicity, or other in vivo test results and 

to provide a database for use in structure-activity analyses. Analysis of the early, multi-test database showed 

that positive results for a chemical in the Salmonella gene mutation assay were sometimes correlated with 

carcinogenicity in several species/sexes of rodents and at several tissue sites. Subsequently, studies of the 

correlation between mutagenicity test data and rodent carcinogenicity showed a strong association between 

clearly positive results in long-term mouse peripheral blood micronucleus tests and rodent carcinogenicity. 

The importance of genetic toxicity test data in assessing exposure hazard for NTP chemicals is underscored by 

the fact that most organic chemicals (other than hormones) identified as human carcinogens by the International 

Agency for Research on Cancer (IARC) are genotoxic, and the vast majority of them are detected by both the 

Salmonella assay and rodent micronucleus tests. Additional assays may be conducted with certain chemicals 

to gain further insight into the types of DNA and chromosomal damage induced by a chemical. Gene mutations 

and DNA damage are examined in tumors from NTP studies on a case-by-case basis; cytogenetic effects, 

measured as the induction of micronuclei, are generally examined in bone marrow cells or in peripheral 

erythrocytes. Substances tested for genetic toxicity during FY 2011 are listed in Table 24. Information is 

available at http://ntp.niehs.nih.gov/go/reports.

Table 24. Ongoing and Completed Genetic Toxicity Studies during FY 2011

Chemical [CASRN] Testing Battery

3'-Azido-3'-deoxythymidine (AZT) 30516-87-1 Salmonella

Black cohosh 84776-26-1 Micronucleus

Chlorhexidine 55-56-1 Salmonella

N-Butylbenzenesulfonamide 3622-84-2 Salmonella

2,6-Diethylaniline (2,6-DEA) 579-66-8 Salmonella

N,N-Dimethyl-p-toluidine 99-97-8 Micronucleus

2',2'''-Dithiobisbenzanilide 135-57-9 Salmonella

2-Ethylaniline 578-54-1 Salmonella

3-Ethylaniline 587-02-0 Salmonella

4-Ethylaniline 589-16-2 Salmonella

2-metyl-6-ethylanline (2M6EA) 24549-06-2 Salmonella

Nelfinavir 159989-64-7 Salmonella

Nevirapine 129618-40-2 Salmonella

2-nitro-1-propanol 2902-96-7 Salmonella

2-Nitro-2-ethyl-1,3-propanediol 597-09-1 Salmonella

2-Nitro-2-methyl-1,3-propanediol 77-49-6 Salmonella

Styrene-acrylonitrile trimer Micronucleus

m-Toluidine 108-44-1 Salmonella

p-Toluidine 106-49-0 Salmonella

2,3-Pentanedione 600-14-6 Micronucleus

3TC (AIDS initiative) 134678-17-4 Salmonella

Valerian (various) Salmonella

Zinc carbonate, basic 5263-02-5 Micronucleus
Table 25. Ongoing and Completed Organ Systems Toxicity Studies During FY 2011 

Chemical CASRN Species/Strain Route Project 

Leader 

Testing Battery 

Acrylamide 79-06-1 Rats: F344 Gavage Beland Neurotox assessment 

Bitter orange Rats: SD Gavage Hansen Developmental 

Bitter orange with caffeine Rats: SD Gavage Hansen Developmental 

Black cohosh 84776-26-1 Rats: Harlan SD Gavage MercadoFeliciano RACB – range-finding 

Black cohosh 84776-26-1 Rats: Harlan SD Gavage MercadoFeliciano Continuous breeding 

tris(2-Chloroisopropyl)phosphate 13674-84-5 Rats: Harlan SD Gavage Stout Prenatal developmental toxicity 

Endocrine disruptor (Nonylphenol) 84852-15-3 Rats: SD Feed Newbold, 

Delclos 

Multigeneration 

2-Hydroxy-4­

methoxybenzophenone 

131-57-7 Rats: Harlan SD Feed Hansen Total reproductive capacity 

4-Methylimidazole 822-36-6 Rats: Harlan SD Feed Bishop RACB – range-finding 

p-Synephrine 94-07-5 Rats: SD Gavage Hansen Developmental 

p-Synephrine and caffeine 58-08-2 

94-07-5 

Rats: SD Gavage Hansen Developmental 

3,3',4,4'-Tetrachloroazobenzene 14047-09-7 Rats: Crl:CD SD Gavage Hooth Developmental – pre-implantation 

3,3',4,4'-Tetrachloroazobenzene 14047-09-7 Rats: Crl:CD SD Gavage Hooth Postnatal developmental toxicity 

Organ System Toxicity 

Nervous System, Developmental, and Reproductive Toxicity 

Behavioral and neurological alterations in response to deleterious environmental agents often represent the 

earliest evidence of toxicity. These testing batteries examine the sensory, motor, autonomic, and peripheral 

nervous systems. The Functional Observational Battery employs observational screening, while the NIEHS 

test battery uses automated test systems to evaluate the various nervous system components. 

As part of its charge to test chemicals of concern for potential toxicity, the NTP evaluates developmental and 

reproductive toxicity primarily by using teratology and Reproductive Assessment by Continuous Breeding (RACB) 

study designs (see http://ntp.niehs.nih.gov/go/33668). The RACB study design was developed by the NTP to 

identify potential hazards from toxic effects on male and/or female reproduction, to characterize that toxicity, 

and to define the dose-response relationships for each compound. The study design has evolved over the years: 

initially the studies mainly used mice as the test species; now, they use rats almost exclusively. As our knowledge 

has improved and use of sensitive endpoints has increased, these advances have been incorporated into revisions 

of the study design. Table 25 lists completed and ongoing nervous system, developmental, and reproductive 

studies during FY 2011. 

NTP Annual Report 2011 55 
56 National Toxicology Program

The NTP’s Modified One-generation Reproduction Study (see http://ntp.niehs.nih.gov/go/MG) design evolved 

from (1) the NTP changing its default exposure paradigm in rat cancer bioassays to include treatment exposure

during pregnancy and early life, (2) the increased knowledge of critical windows of exposure, which indicates the 

need for a larger focus on evaluating the potential for postnatal adverse effects, and (3) the updates to standard

study designs with more functional endpoints to assess how agents affect the reproductive and endocrine 

status of animals. The classical study used to evaluate reproductive toxicity is the multigenerational reproduction

experimental design. The NTP has modified this classical study design to better utilize the animals produced and 

to reduce animal use by improved experimental design and statistical power. Table 26 lists planned or ongoing

Modified One-generation studies. Table 27 lists RACB studies planned for FY 2012.

Chemical CASRN Species/Strain Study 

Route

Planned Cohorts Project Leader

Bisphenol AF 1478-61-1 Rats: Harlan SD Feed Dose range-finding

Maternal transfer

Developmental toxicity

Fertility assessment 

Neurotoxicity assessment

MercadoFelliciano

N-Butylbenznensulfonamide 3622-84-2 Rats: Harlan SD Feed Dose range-finding

Fertility assessment 

Developmental toxicity

Maternal transfer

Neurotoxicity assessment 

Subchronic toxicity

Rider

Dong quai (Angelica sinesis) root extract 299184-76-2 Rats: Harlan SD Gavage Dose range-finding

Fertility assessment 

Developmental toxicity

Neurotoxicity assessment

McIntyre

2-Ethylhexyl p-methoxycinnamate 5466-77-3 Rats: Harlan SD Feed Dose range-finding McIntyre

Hydroquinone 123-31-9 Rats: Harlan SD Gavage Dose range-finding

Fertility assessment 

Developmental toxicity

Neurotoxicity assessment

DeVito

2-Hydroxy-4-methoxybenzophenone 131-57-7 Rats: Harlan SD Feed Dose range-finding

Fertility assessment 

Developmental toxicity

Neurotoxicity assessment

McIntyre

Hydroxyurea 127-07-1 Rats: Harlan SD Gavage Dose range-finding

Fertility assessment 

Subchronic toxicity

Neurotoxicity assessment 

Developmental toxicity

McIntyre

Isopropylphenyl phosphate 68937-41-7 Rats: Harlan SD Feed Dose range-finding

Neurotoxicity assessment

Behl

Triphenyl phosphate 115-86-6 Rats: Harlan SD Feed Dose range-finding

Neurotoxicity assessment

Behl

Chemical CASRN Species/Strain Route Testing Battery

n-Butyl glycidyl ether 2426-08-6 Rats: Harlan SD TBD RACB – range-finding

n-Butyl glycidyl ether 2426-08-6 Rats: Harlan SD TBD RACB

p-Chloro- , , -trifluorotoluene 98-56-6 Rats: Harlan SD Inhalation RACB

Sodium tungstate, dihydrate 10213-10-2 Rats: Harlan SD TBD RACB

Table 26. Planned or Ongoing Modified One-generation Studies

Table 27. RACB Studies Planned for FY 2012 *

* Known test articles as of 10/1/2011. Others may be scheduled as protocols are finalized.
Table 28. Ongoing and Complete Immunotoxicity Studies During FY 2011 

Chemical CASRN Species/Strain Route Project 

Leader 

Testing Battery 

Abrasive blasting agents (blasting sand) Rats: Harlan SD Inhalation Roycroft Immunosuppression 

Abrasive, blasting agents 

(specular hematite) 

Rats: Harlan SD Inhalation Roycroft Immunosuppression 

Autumn sunset true color concentrate Mice: CBA/ Ca 

Jackson 

Subcutaneous 

injection 

Howard Hypersensitivity 

3'-Azido-3'-deoxythymidine (AZT) 30516-87-1 Mice: B6C3F1 Gavage Germolec Immunosuppression – 

range-finding 

Developmental 

Black cohosh 84776-26-1 Mice: B6C3F1 Gavage MercadoFeliciano Immunosuppression 

2,3-Butanedione (diacetyl) 431-03-8 Mice: BALB/c Topical 

application 

Morgan Hypersensitivity 

2,3-Butanedione (diacetyl) 431-03-8 Mice: BALB/c Inhalation Morgan Immunosuppressionrange-finding 

o-Cresol 95-48-7 Mice: BALB/c Topical 

application 

Chhabra Hypersensitivity 

Dibenz(a,h)anthracene 53-70-3 Mice: B6C3F1 Gavage Germolec Immunosuppressionrange-finding 

- full protocol 

Developmental 

2,3-Dibromo-7,8-dichlorodibenzo-p-dioxin 50585-40-5 Mice: B6C3F1 Gavage DeVito Immunosuppression 

1,3-Dichloropropene (Telone II) 542-75-6 Mice: B6C3F1 Water Germolec Immunosuppressionfull protocol 

Dimethylamine borane 74-94-2 Mice: BALB/c Topical 

application 

Germolec Hypersensitivity 

Double dark fudge true color concentrate Mice: CBA/ Ca 

Jackson 

Subcutaneous 

injection 

Howard Hypersensitivity 

Double fudge concentrate Mice: CBA/ Ca 

Jackson 

Subcutaneous 

injection 

Howard Hypersensitivity 

Immunotoxicity

NTP immunotoxicity studies address adverse effects on the immune system that may result from exposure 

to environmental chemicals, biological materials, or therapeutic agents. The identification of substances that 

have potential to cause injury to the immune system is of considerable public health significance as alterations 

in immune function can lead to increased incidence of hypersensitivity disorders, autoimmune or infectious 

disease, or neoplasia. Immunotoxicity caused by exposure to chemicals can be divided into two broad research 

areas: (1) studies of altered hematopoietic (blood cell development) or other immunologic events associated 

with exposure of humans and animals to chemicals and (2) studies of immune-mediated hypersensitivity (allergy 

and autoimmunity) resulting from exposure to environmental chemicals or therapeutics. In the former case, 

the immune system acts as a passive target (nonspecific) for the foreign substance, and the result may be an 

increased incidence or severity of infectious disease or neoplasia because of the inability to respond adequately 

to the invading agent. In hypersensitivity (i.e., allergy), the immune system responds to small molecular weight 

compounds that bind to host tissue, recognizing the complex as foreign antigen. This immune response to the 

chemical-host tissue complex may lead to diseases, such as respiratory tract allergies (e.g., asthma, rhinitis) or 

allergic contact (skin) dermatitis. Autoimmunity, another form of immune-mediated disease, is characterized by 

an immune response against constituents of the body’s own tissues (autoantigens). Table 28 lists ongoing and 

completed immunotoxicity studies during FY 2011. 

NTP Annual Report 2011 57 
58 National Toxicology Program

Chemical CASRN Species/Strain Route Project

Leader

Testing Battery

Echinacea purpurea, extract 90028-20-9 Mice: B6C3F1 Gavage Germolec Immunosuppression – 

full protocol

Elmiron (sodium pentosanpolysulfate) 37319-17-8 Mice: B6C3F1 Gavage Germolec Immunosuppressionrange-finding

– full protocol

Endocrine disruptor (Nonylphenol) 84852-15-3 Rats: SD Feed Newbold,

Delclos

Multigeneration

2-Ethylhexyl p-methoxycinnamate 5466-77-3 Mice: BALB/c Topical 

application

McIntyre Hypersensitivity

Genistein 446-72-0 Mice: NOD/

MrKTac

Gavage Germolec Autoimmunity

Gum guggul extract Mice: B6C3F1 Gavage Thakur Immunosuppression – 

range-finding

Ionic liquid 

(1-butyl-1-methylpyrrolidinium chloride)

479500-35-1 Mice: BALB/c Topical 

application

Hooth Hypersensitivity

Ionic liquid 

(n-butylpyridinium chloride)

1124-64-7 Mice: BALB/c Topical 

application

Hooth Hypersensitivity

Lovastatin 75330-75-5 Mice: B6C3F1 Gavage Germolec Immunosuppression – 

range-finding

2-Methoxy-4-nitroaniline 97-52-9 Mice: BALB/c Dermal Surh Hypersensitivity

4-Methylimidazole 822-36-6 Rats: Harlan SD Feed Bishop Range-finding

Monoclonal antibody 

protein therapeutics (CD-4)

Mice: B6C3F1 Intraperitoneal 

injection

Germolec Immunosuppression – 

full protocol

Monoclonal antibody 

protein therapeutics (CD-8)

Mice: B6C3F1 Intraperitoneal 

injection

Germolec Immunosuppression – 

full protocol

Nanoscale material 

(Fullerene-C60 1 micron)

99685-96-8 Rats: Wistar

Han

Inhalation Walker Immunosuppression – 

range-finding

Nanoscale material 

(Fullerene-C60 1 micron)

99685-96-8 Mice: B6C3F1 Inhalation Walker Immunosuppression – 

rang- finding

Nanoscale material 

(Fullerene-C60 50 nanometers)

99685-96-8 Rats: Wistar

Han

Inhalation Walker Immunosuppression – 

range-finding

Nanoscale material 

(Fullerene-C60 50 nanometers)

99685-96-8 Mice: B6C3F1 Inhalation Walker Immunosuppression – 

range-finding

1,5-Naphthalene diisocyanate 3173-72-6 Mice: BALB/c Topical 

application

Germolec Hypersensitivity

Nelfinavir mesylate 159989-

65-8

Mice: B6C3F1 Gavage Germolec Immunosuppression – 

full protocol

Developmental

Nevirapine 129618-40-2 Mice: B6C3F1 Gavage Germolec Immunosuppression – 

full protocol

– developmental

1,2,3,7,8-Pentabromodibenzofuran 107555-93-1 Mice: B6C3F1 Gavage DeVito Immunosuppression

2,3,4,7,8-Pentabromodibenzofuran 13116-92-2 Mice: B6C3F1 Gavage DeVito Immunosuppression

1,2,3,7,8-Pentachlorodibenzofuran (PCDF) 57117-41-6 Mice: B6C3F1 Gavage DeVito Immunosuppression

2,3,4,7,8-Pentachlorodibenzofuran 57117-31-4 Mice: B6C3F1 Gavage DeVito Immunosuppression

2,3-Pentanedione 600-14-6 Mice: BALB/c Topical 

application

Germolec Hypersensitivity

Perfluorodecanoic acid (PFDA) 335-76-2 Rats: Harlan SD Gavage Blystone Immunosuppression
Chemical CASRN Species/Strain Route Project 

Leader 

Testing Battery 

Phenol 108-95-2 Mice: B6C3F1 Water Germolec Immunosuppression – 

full protocol 

Resveratrol 501-36-0 Mice: B6C3F1 Gavage Germolec Immunosuppression – 

range finding 

Rosewood true color concentrate Mice: CBA/ Ca 

Jackson 

Subcutaneous 

injection 

Howard Hypersensitivity 

Sodium tungstate, dihydrate 10213-10-2 Mice: B6C3F1 Water Hooth Immunosuppression – 

full protocol 

2,3,7,8-Tetrabromodibenzofuran 67733-57-7 Mice: B6C3F1 Gavage DeVito Immunosuppression 

3,3',4,4'-Tetrachloroazobenzene 14047-09-7 Rats: SD Gavage Behl Immunosuppression – 

range finding 

– developmental 

2,3,7,8-Tetrachlorodibenzo-p-dioxin 1746-01-6 Mice: B6C3F1 Gavage DeVito Immunosuppression 

2,3,7,8-Tetrachlorodibenzofuran 51207-31-9 Mice: B6C3F1 Gavage DeVito Immunosuppression 

2,3,7-Tribromodibenzo-p-dioxin 51974-40-4 Mice: B6C3F1 Gavage DeVito Immunosuppression 

Table 29 . Ongoing and Complete Disposition, Metabolism, and Toxicokinetic Studies During FY 2011 

Chemical CASRN Species/Strain Route Project Leader 

Anatase (TiO2) 1317-70-0 Mice: Tg.AC (FVB/N) Topical application NCTR 

Mice: FVB/N 

Benzene 71-43-2 Mice: BALB/cByJ Gavage Cunningham 

Mice: WSB/EiJ (M. m. domesticus) 

Mice: CAST/EiJ (M. m. castaneus) 

Mice: C3H/HeJ 

Mice: DBA/2 Jackson 

Mice: BTBR T+ tf/J 2 

Mice:C57BL/6 

Mice: FVB/NJ 

Mice: KK/HIJ 

Mice: AKR/J 

Mice: NZW/LacJ 

Mice: A/J 

Mice: NOD/LtJ 

Mice: B6C3F1 

Mice: PWD/PhJ (M. m. musculus) 

Mice: MOLF/EiJ (M. m. molossinus) 

Mice: 129S1/SvImJ 

Disposition, Metabolism, and Toxicokinetic Studies 

Complete dosimetry of a chemical or physical agent describes its absorption, distribution, metabolism, and 

excretion (ADME) in the body at differing levels of exposure, over all ages, via several routes of exposure, 

and under varying genetic backgrounds in humans and test animals. Data from NTP chemical disposition 

and toxicokinetic studies are used in these studies. Substances evaluated during FY 2011 are listed in Table 29, 

and studies planned for FY 2012 are listed in Table 30. Most studies are conducted in intact laboratory animals; 

some require incubating human and rodent tissues (liver slices) with the chemical. This information provides 

dosimetric data that can be combined with other anatomical, biochemical, and physiological information to 

develop models based on biochemistry and physiologically based pharmacokinetics. Such models are used 

increasingly in risk assessment to extrapolate between species, across dose ranges, and across different routes 

of exposure. 

NTP Annual Report 2011 59 
60 National Toxicology Program

Chemical CASRN Species/Strain Route Project Leader

Bisphenol A 80-05-7 Rats: SD In-vitro NCTR

Bisphenol AF 1478-61-1 Mice: B6C3F1

Rats: Harlan SD

In-vitro Waidyanatha

2,3-Butanedione (diacetyl) 431-03-8 Mice: B6C3F1 

Rats: Harlan SD

Oropharyngeal Waidyanatha

2,3-Butanedione (diacetyl) 431-03-8 In-vitro Waidyanatha

2-Butene-1,4-diol 110-64-5 In-vitro NCTR

n-Butyl-p-hydroxybenzoate 94-26-8 Rats: Harlan SD Intravenous Blystone

Cumene 98-82-8 Rats: Fischer 344 

Mice: B6C3F1

Gavage Chan

1,3-Dichloro-2-propanol 96-23-1 Rats: Harlan SD

Mice: B6C3F1

Gavage Chan

Di(2-ethylhexyl) phthalate 117-81-7 Monkey: Rhesus Gavage Delclos

N,N-Dimethylacetoacetamide 2044-64-6 Rats: Fischer 344/N Charles River Gavage Waidyanatha

Dimethylamine borane 74-94-2 Human skin cells 

Rats: Harlan SD

In-vitro Germolec

Dimethylethanolamine 108-01-0 Rats: Wistar

Mice: B6C3F1

Gavage Waidyanatha

2,2’-Dimorpholinodiethyl 

ether

6425-39-4 Rats: Harlan SD

Mice: B6C3F1/N

Dermal Waidyanatha

2',2'''-Dithiobisbenzanilide 135-57-9 Rats; Harlan SD

Mice: B6C3F1

Gavage Waidyanatha

Ephedrine + caffeine

combination

58-08-2 

299-42-3

Rats: Fischer 344 Gavage Dunnick

2-Ethylhexyl 

p-methoxycinnamate

5466-77-3 Rats: Harlan SD

Mice: B6C3F1

Dermal McIntyre

2-Ethylhexyl 

p-methoxycinnamate

5466-77-3 Rats: SD Gavage Mercado-Feliciano

Furan 110-00-9 Rats: Fischer 344 Gavage Waidyanatha

Furan 110-00-9 Rats: Tg.Lac1/C57BL/6 (Big Blue) Gavage Waidyanatha

Gum guggul extract Human liver microsomes In-vitro Thakur

Isocyanuric acid 108-80-5 Rats: Fischer 344 Gavage NCTR

Isocyanuric acid 108-80-5 Rats: Fischer 344 Intravenous NCTR

2-Hydroxy-4-

methoxybenzophenone

131-57-7 Rats: Harlan SD

Mice: B6C3F1

Dermal Auerbach

Melamine 108-78-1 Rats: Fischer 344 Gavage NCTR

Melamine 108-78-1 Rats: Fischer 344 Intravenous NCTR
Chemical CASRN Species/Strain Route Project Leader 

Melamine cyanurate 37640-57-6 Rats: Fischer 344 Gavage NCTR 

Melamine + cyanuric acid 

combination 

108-78-1 

108-80-5 

Rats: Fischer 344 Feed NCTR 

Melamine + cyanuric acid 

combination 

108-78-1 

108-80-5 

Rats: Fischer 344 Intravenous NCTR 

2-Methoxy-4-nitroaniline 97-52-9 Rats: Harlan SD 

Mice: B6C3F1 

Gavage Auerbach 

L-beta-Methylaminoalanine 15920-93-1 Rats: Harlan SD 

Mice: B6C3F1 

Gavage Waidyanatha 

Nanoscale material 

(Fullerene C60) 

99685-96-8 Rats: Fischer 344/N Intratracheal Waidyanatha 

Nanoscale material 

(rutile titanium dioxide) 

1317-80-2 Mice: SKH-1 Hairless Topical application NCTR 

Nanoscale Ag 7440-22-4 Rats: SD Gavage Walker 

Nanoscale Ag 7440-22-4 Rats: SD Intravenous Walker 

Ag acetate 563-63-3 Rats: SD Intravenous NCTR 

Ag acetate 563-63-3 Rats: SD Gavage NCTR 

Triclosan 3380-34-5 Mice: B6C3F1 

Mice: B6C3F1/NCTR BR 

(C57BL/6N x C3H/HEN MTV-) 

Rats: Harlan SD 

Topical NCTR 

Tris(4-chlorophenyl)methane 27575-78-6 Rats: Harlan SD 

Mice: B6C3F1 

Gavage Surh 

Tris(4-chlorophenyl)methanol 3010-80-8 Rats: Harlan SD Gavage Surh 

Table 30: Disposition on, Metabolism and Toxicokinetic Studies Planned for FY 2012* 

Chemical CASRN Species/Strain Route 

Bisphenol AF 1478-61-1 Rats: Harlan SD 

Mice: B6C3F1 

Intravenous 

Tris(2-chlorisopropyl) phosphate 13674-84-5 Mice: B6C3F1 

Rats: Harlan SD 

Feed 

* Known test articles as of 10/1/2011. Others may be scheduled as protocols are finalized. 

NTP Annual Report 2011 61 
62 National Toxicology Program

Genetic and Alternative Test Systems

Biomolecular Screening

In FY 2008, the NTP established a high throughput screening (HTS) initiative, representing a new paradigm 

in toxicological testing. During FY 2011, the NTP continued using this HTS approach to screen for mechanistic 

targets active within cellular pathways critical to carcinogenicity, reproductive and developmental toxicity, 

genotoxicity, neurotoxicity, and immunotoxicity. The NTP’s HTS program is administered through the 

Biomolecular Screening Branch (BSB). The goals of the HTS Program are to:

• prioritize substances for further in-depth toxicological evaluation (to judiciously allocate efforts 

and resources to maximize public health impact)

• identify mechanisms of action for further investigation (e.g., disease-associated pathways)

• develop predictive models for biological responses in humans and animals (predictive toxicology)

Much of the research conducted in support of the HTS program is coordinated with the EPA, the National Human 

Genome Research Institute (NHGRI), and the FDA through a Memorandum of Understanding (see page 73).

Contact Information: Dr. Raymond Tice, Chief, BSB, tice@niehs.nih.gov. HTS website http://ntp.niehs.nih.gov/go/28213.
NTP Annual Report 2011 63

Comparative Pathology

At NTP emphasisis increasingly being placed upon the toxicologic pathology produced by acute,subchronic, 

and chronic chemical exposures in rodent bioassays for potential translation to human hazard identification.

Many of these studies produce data that can be utilized by regulatory agencies in setting exposure limits in the 

workplace (for example, the ongoing studies of chemical-induced bronchial fibrosis). Numerous animal models of

human disease, such as asthma, pneumonitis, nonalcoholic fatty liver disease, and bleomycin induced pulmonary 

fibrosis are regularly interpreted by the Cellular and Molecular Pathology Branch (CMPB) in collaboration with

NIEHS colleagues. Comparative pathology, both morphologic and molecular, is also of paramount importance

in assessing chemical-induced neoplasia, as demonstrated by the recent NIEHS and NCTR collaborative studies 

of chemical-induced intestinal tumors. Recent efforts by the CMPB have also focused upon the development 

of necropsy techniques for the rapid collection of rodent tissues for gene expression studies; these innovative 

techniques, combined with the use of laser capture microdissection, have resulted in the successful collection of

high quality RNA from the pulmonary airways and from the epithelium of the nasal cavity. Ongoing CMPB cellular

and molecular approaches are thus providing greater opportunities for comparing responses between animal

models and humans and for determining human relevance.

Molecular Mechanisms of Chemically Induced Neoplasia

NTP studies have identified several genetic alterations in chemically induced rodent neoplasms that have

potential mechanistic implications for human cancer. These studies provide convincing molecular data that serve 

to complement standard histopathology data. Taken together, these data aid the NTP in assessment of several 

compounds that are potentially hazardous. Epigenetic as well as genetic mechanisms play an important role in

the pathogenesis of cancer. Several independent and collaborative studies have been initiated to investigate the 

epigenetic changes occurring in chemically induced and spontaneous neoplasms, including global methylation 

profiling and pyrosequencing to better understand how these changes influence the process of tumorigenesis.

These studies further complement our investigations of genetic alterations in liver, lung, breast, mesothelioma, 

and colon tumors induced by compounds in NTP studies. Protocols to standardize frozen tissue collection from

NTP chronic studies for molecular biology analysis including gene expression and protein analysis have been

initiated. Using the high quality samples obtained from the NTP chronic studies in conjunction with 

high-throughput global gene expression profiling, important similarities were discovered between mouse 

and human tumors at the molecular level. Additionally, thistechnology provided important information on

chemical-specific events leading to tumorigenesis. It is becoming increasingly important in NTP studies to identify

pre-disease early genetic and epigenetic events in the process of toxicity and carcinogenicity in order to establish 

predictive as well as preventive measures to minimize human health hazards. In order to achieve that objective,

study design of new NTP studies have included early timepoints for molecular evaluation. Investigation of gene

and protein expression alterations in tissues during the pre-neoplastic stages will provide important data about 

the initiating events that will provide critical information on predictive events in the pathogenesis of chemically

induced neoplasms in NTP studies. NTP contributors were Drs. Mark Hoenerhoff, Sachin Bhusari, and Robert Sills,

Ms. Janice Harvey, Ms. Hue Hua Hong and Mr. Tai-Vu Ton.

NTP Nonneoplastic Lesion Atlas

Given the increased attention to nonneoplastic diseases in humans, and the potential to contribute to the body of

knowledge on nonneoplastic diseases, the NTP has been working to investigate the relevance of nonneoplastic

lesions seen in its toxicity and carcinogenicity studies to humans. A great deal of variation occurs among
64 National Toxicology Program

pathologists regarding the terminology and recording for nonneoplastic lesions, creating inconsistency in the 

NTP’s nonneoplastic lesion database. To resolve these issues, the NTP plans to implement a set of guidelines

for the diagnosis of nonneoplastic lesions, similar to the system currently used for neoplastic lesions. These 

guidelines will be presented in the NTP Nonneoplastic Lesion Atlas (NNLA), which will improve the organization

and diagnostic consistency of the NTP’s nonneoplastic lesion database. This will facilitate database searches and 

allow the generation of historical control data for nonneoplastic lesions.

The NNLA will be divided into 22 sections, each covering a particular organ or organ system. The sections will

consist of a series of documents and will include high-quality images of the lesions, a comments section, a 

recommendations section that presents the diagnostic guidelines, and a list of references. Each section will

be prepared by a group of 2 or 3 pathologists familiar with NTP studies, and will be reviewed by a non-NTP

pathologist (generally, a recognized expert on the particular organ or organ system) and by a group of NTP 

pathologists. Currently, two of the 22 sections have been completed, and an additional 10 sections are in review. 

The remaining sections will be in review by mid-2012 and the atlas is scheduled to be completed by late 2013. 

The NNLA will be published as a downloadable web document, will be updated periodically, and will be

available as an app for use on smart phones and tablets eventually. 

Additionally, NTP pathologists are in working groups for the Society of Toxicologic Pathology (STP) and 

International Harmonization of Nomenclature and Diagnostic Criteria (INHAND) committee, which has a goal of

a worldwide nomenclature standardization of rodent neoplastic and non-neoplastic lesions. The international

efforts are relying heavily on NTP expertise and opinion as well as using the NTP’s digital database of >60,000

pathology images for examples for the atlases. Some organ systems are complete. The NTP Pathology Group also

provided assistance and attended meetings to address pathology issues for the pending conversion of the NTP’s 

toxicology data management system to the new Provantis system. 

NTP Satellite Meeting

The 2011 annual NTP Satellite Symposium, titled Pathology Potpourri, was held in Denver, Colorado in

advance of the STP 30th Annual Meeting in June 2011. Some of the topics covered during the symposium 

include: proliferative lesions from various fish species including ameloblastoma, gas gland hyperplasia, nodular

regenerative hepatocellular hyperplasia, and malignant granulosa cell tumor; spontaneous cystic hyperplasia 

in the stomach of CD1 mice and histiocytic aggregates in the duodenal villous tips of treated mice; an olfactory 
NTP Annual Report 2011 65

neuroblastoma in a cynomolgus monkey; various rodent skin lesions including follicular parakeratotic 

hyperkeratosis, adnexal degeneration, and epithelial intracytoplasmic accumulations; oligodendroglioma 

and microgliomas in rats; a diagnostically challenging microcytic, hypochromic, responsive anemia in rats; 

a review of microcytes and microcytosis; nasal lesions associated with green tea extract and Ginkgo biloba

in rats; corneal dystrophy in Dutch belted rabbits; valvulopathy in rats; and lymphoproliferative disease in 

a cynomolgus monkey. Dr. Susan Elmore organized and chaired the Satellite Symposium.

Digital Imaging

The NTP Pathology Group has continued to foster the exploration and utilization of digital photo microscopy 

and slide scanning technologies for NTP and NIEHS studies. Such efforts have contributed towards abilities for 

publishing and presenting stellar microscopic images, providing web-based telepathology to conduct pathology 

peer reviews, and obtaining instantaneous opinions from pathologists at remote locations around the world. 

The NTP has completed the last of 12 studies designed to compare digital and light microscopy for diagnostic

accuracy in pathology peer review. 

Pathology Support Core Activities

Based on the recommendations of an onsite External Scientific Review of the 

CMPB Core Laboratories conducted in 2010, CMPB has undertaken several 

steps to modify the Core Labs Infrastructure: (1) the Immunohistochemistry 

and Electron Microscopy (EM) Cores have retained the services of onsite 

pathologists, (2) the Immunohistochemistry Core Laboratory has procured 

a new fully automated, robotic immunohistochemistry stainer, (3) the 

Immunohistochemistry Core Laboratory has added polymer-based technology

for protein detection to the IHC staining protocols, (4) a weeklong workshop

in immunohistochemistry was conducted at NIEHS to refresh and enhance

the knowledge base of the Immunohistochemistry lab staff and to provide 

them an opportunity to be familiar with the state of the art techniques in

immunohistochemistry, (5) the Immunohistochemistry Core Laboratory has 

worked to standardize the protocols for immunofluorescence microscopy and is

now able to provide more immunofluorescence service to the researchers in the coming year, (6) the EM Core 

has purchased a new state-of-the-art, 11-megapixel, fiber-optic coupled camera to replace an out dated camera

that is no longer being manufactured, (7) Core staff have worked with computer IT support to refine, improve, 

and update the computerized Immunhistochemistry/Necropsy/Histology Core Laboratory Tracking Applications, 

and (8) key Necropsy/Histology Core staff attended a five-day off-site retraining to reinforce and refine their skills

in histotechnology and to keep them up to date on new procedures and methods.

As part of the Pathology Services Core, the Special Techniques Laboratory (STL) provides services to DIR and 

DNTP in the areas of laser capture microdissection (LCM), imaging and mouse phenotyping with an emphasis on

embryonic lethality. In FY 2011, new services were offered to support investigators, which include extraction of

DNA and RNA from tissue, as well as, the amplification of these end products for final analysis by the investigators.

The imaging core acquired a microscopic slide scanner capable of digitizing fluorescence-stained slides. The 

other support service in STL, mouse phenotyping, places a focus on assisting investigators to identify embryonic 

lethality in genetically engineered mice. In 2011, multiple investigators were assisted with projects such as, 

evaluating CRE toxicity and development of special stainsto detect bone anomalies. In addition, studies were

designed and conducted to troubleshoot fixation and paraffin embedding problems with embryo samples.

Contact Information: Cellular and Molecular Pathology Branch, Dr. Robert Sills, Chief, sills@niehs.nih.gov.

Photo courtesy of EHP
66 National Toxicology Program

Established in 1984, the NTP Archives is a state-of-the-art facility, providing the primary public resource for 

toxicology research. The Archives houses an unmatched collection of research specimens and supporting data

from over 2,000 NTP studies. The facility consists of repositories for storage of histologic slides, paraffin blocks, 

formalin-fixed “wet” tissue, frozen tissues, and printed, microfiched, and electronic study records. Frozen samples 

include normal tissue, neoplastic lesions, tumor specimens, DNA, RNA, blood serum, BAL supernatant, urine, and 

sperm suspensions from treated and control rats and mice.

Contact Information: Dr. Ron Herbert, herbert1@niehs.nih.gov

Reorganization of NTP into a separate division yielded the new NTP Laboratory. The NTP Laboratory provides

high quality laboratory capabilities and support for the performance of agent-specific, targeted research directly

related to developing and applying tools of modern toxicology and molecular biology to the evaluation of

specific substances of concern to NTP, issues of central importance to NTP programs, or methods development 

to advance the NTP mission. Another key focus of the laboratory isto develop better methods to study the 

developmental origins of adult diseases.

Dr. Michael Waalkes is Laboratory Chief and Drs. Sue Fenton, Darlene Dixon, Jean Harry, and Daniel Morgan are

group leaders within the Laboratory, while other NTP scientists join projects on an as need basis. Several studies 

are being initiated or formulated as the Laboratory further develops its project repertoire. Mechanistic studies 

on formaldehyde and hematopoietic cancers, genomics of early life exposures and adulthood cancers in arsenic 

exposed humans, metal particle dissolution by macrophage, and effects of environmental estrogens are just

some of the first studies in design or process.

Contact Information: Dr. Michael Waalkes, waalkes@niehs.nih.gov and at

http://www.niehs.nih.gov/research/atniehs/labs/ntp/index.cfm
NTP Annual Report 2011 67

Laboratory Animal Medicine Training Program

This four-year training program, under the direction of Dr. Angela King-Herbert, includes clinical and surgical 

responsibilities, management of animal care facilities, participation in research projects, and laboratory animal

pathology. The training program is a collaborative effort between NIEHS and the University of North Carolina at

Chapel Hill. Fellows interact with laboratory animal veterinarians at NIEHS and at local area academic, industrial, 

and government facilities to receive didactic and hands-on training in laboratory animal medicine. Two 

postdoctoral fellows, Drs. Jacquelyn Tubbs and Dr. Coralie Zegre-Cannon, completed the program and 

passed the American College of Laboratory Animal Medicine certifying examination in June 2011. 

Contact Information: Dr. Angela King-Herbert, kingher1@niehs.nih.gov

Toxicological Pathology Training Program 

Since formalizing a training program in toxicological pathology in 2003, Dr. David Malarkey, training coordinator,

and other CMPB staff have mentored 11 post-doctoral fellows and over 30 veterinary student externs. In 2011, the 

training program included the training of IRTA fellows, multiple externs who come to CMPB for a few weeks, and 

other local pathology trainees. Three postdoctoral fellows and six veterinary students participated in the program 

during 2011. The program is designed to introduce students to the field and career opportunities in veterinary 

and toxicological pathology while also providing hands-on projects often leading to abstracts and publications. 

Postdoctoral fellows learn rodent and toxicological pathology, participate in NTP and other DIR research projects, 

work to achieve accuracy of NTP pathology data by assisting the NTP pathologist on NTP studies, and continue

education towards achievement of board certification by the American College of Veterinary Pathologists. CMPB

staff also served as committee members for three PhD graduate school students at North Carolina State University.

Contact Information: Dr. David Malarkey, malarkey@niehs.nih.gov

Toxicology and Carcinogenesis Training Program

Trainees in this program learn to perform all aspects of contracted toxicology studies for carcinogenic or 

non-carcinogenic endpoints (e.g., reproductive and developmental effects, immune system function). They learn 

about NTP efforts in molecular toxicology and HTS and receive training applicable to regulatory or industrial

toxicology. By serving as study scientists in non-laboratory positions, they evaluate the toxicity of substances 

of interest to the NTP. Trainees actively participate in designing, conducting, and evaluating studies and interact

extensively with chemistry, pathology, toxicokinetics, toxicogenomics, genetics, epidemiology, statistics, and 

molecular biology staff. The program currently has three postdoctoral fellows: Drs. Minerva Mercado-Feliciano,

In Ok Surh, and Sheetal Thakur. Two NTP trainees, Drs. Mamta Behl and Sang-Hyun Kim, successfully transitioned

to other full time appointments on completion of their training program.

Contact Information: Dr. Paul Foster, foster2@niehs.nih.gov
68 National Toxicology Program

NIEHS/NCTR Interagency Agreement

In 1992, FDA entered into an IAG with NIEHS. The IAG is an instrument that allows chemicals nominated to the 

NTP to be studied for toxicity using the unique resources and facilities at NCTR. The research conducted under 

the IAG allows the FDA to better assess study design input and initial data on the safety of FDA-regulated 

products. It has allowed continued collaborative toxicity testing on compounds of interest to the FDA and NTP 

and led to investigations of mechanisms of action and assessments of toxicity for many classes of chemicals 

including cosmetics, endocrine-disrupting compounds, food contaminants, food cooking by-products, dietary 

supplements, drugs, and anesthetics. The IAG supports the Phototoxicity Research and Testing Laboratory at the 

NTP Center for Phototoxicology and the Nanotechnology Core Facility at the NCTR/Office of Regulatory Affairs.

All toxicology studies conducted under the IAG are designed with input from FDA regulatory scientists, NCTR

and NIEHS scientists, scientists from other agencies, and invited subject matter experts. The IAG uses resources 

from public funds and exceptional scientific expertise to provide the best possible assessment of product safety

through toxicological studies. Table 31 lists projects completed or ongoing in FY 2011.

Study [CASRN] Principal Investigator Objective and/or Rationale

Assessment of Molecular Changes in Male and Female

SD Rats Orally Exposed to BPA from 

GD 6 through PND 90 [80-05-7] Camacho

To determine BPA-induced molecular changes in gene expression, protein levels, and 

epigenetic modifications in tissues collected from SD rats orally exposed to BPA from

GD 6 through PND 90.

Evaluation of Toxicity of BPA in Male and Female SD/

NCTR Rats Exposed Orally from GD 6 through PND 90 

[80-05-7] Delclos

(1) To assess the toxicity of BPA in rats dosed perinatally via gavage, and (2) to evaluate

estrogenic endpoints from the F1 generation.

Evaluation of Various Diets on Endpoints Critical to the 

Evaluation of BPA and other Endocrine Active Agents

Delclos

To evaluate reproductive and developmental endpoints in F0-F2 generation in CD-1

mice with various chows (e.g.,Purina Mills 5K96, NIH-41,Purina Mills 5001), some of

which have low isoflavone levels, in preparation of studies on BPA in CD-1 mice

The Role of Perinatal Development on

Pharmacokinetics of BPA 

[80-05-7] Doerge

To develop additional data from rat and monkey exposures that will provide data to

be used in the creation and validation of a PBPK model to predict internal exposures

to free BPA in the appropriate target tissues of fetuses and babies that are derived

from food contact and medical-device exposures.

13-week Studies to Determine the Pathogenesis of the 

Whole Leaf Extract of the Aloe vera Plant in the Cecum 

and Large Intestine of the F344 Rat 

[85507-69-3] Boudreau

To investigate the pathogenesis of Aloe vera extracts and gel, with and without

added Aloin A, in the cecum and large intestine of the F344 rat. Senna, having similar

components to those in Aloe vera, will also be studied to determine if it exerts

comparable effects when administered in the drinking water of F344 rats.

Bioassays in the F344 Rat and the B6C3F1 Mouse 

Administered Aloe vera Plant Constituents

in Drinking Water 

Boudreau

(1) To determine dose ranges for future toxicology studies, (2) to determine the toxic

effects and highest tolerated doses of the different Aloe vera components, including

the gel and whole leaf extracts [filtered and non-filtered], and (3) to determine the

chronic toxicity and carcinogenic potential of Aloe vera whole leaf extract in the rat 

and mouse by drinking water exposure.

Assessment of the Nephrotoxic Effect of a Combined

Exposure to Melamine and Cyanuric Acid 

[108-78-1, 108-90-5] Gamboa

To investigate the toxic effects noted in cats and small children who were exposed 

to melamine and cyanuric acid via adulterated food supplies. 

Assessment of the Nephrotoxicity from the 90-day

Combined Exposure to Melamine and Cyanuric Acid 

in F344 Rats 

[108-78-1, 108-90-5] Gamboa

To investigate the nephrotoxic effects noted in F344 rats exposed in NTP Study 

#C10119 to melamine, cyanuric acid and melamine cyanurate. The initial toxicity 

was reported in the kidneys of the pets and children exposed to adulterated foods.

13-week Study to Evaluate the Toxicology of Ag

Nanoparticles in SD Rats [744-22-4] Boudreau

To determine if exposure over a 13-week period to nanoscale (10, 70, and 107 nm) 

Ag particles induces toxicity.

Toxicological Evaluation of Nanoscale Ag Particles 

in Rodents [744-22-4] Boudreau

(1) To determine if reducing the size of Ag particles to nanoscale changes the

absorption, biodistribution, excretion, and toxicity, and (2) to determine the effects 

of reducing the particle size to nanoscale of a compound that has been regarded 

as nontoxic in micro and above sizes.

13-Week Dermal Toxicity of Triclosan in B6C3F1 Mice 

[3380-34-5] Fang

(1) To determine the toxicity of dermally applied triclosan with ethanol as the vehicle, 

and (2) to determine the possible toxicity and phototoxicity of triclosan under normal

use conditions.

Table 31 . NIEHS/NCTR Interagency Agreement Projects in FY 2011
NTP Annual Report 2011 69

Study [CASRN] Principal Investigator Objective and/or Rationale

Vehicle Selection for Triclosan Dermal Toxicity Studies 

in B6C3F1 Mice [3380-34-5] Fang

To determine ADME, toxicokinetics, and penetration of triclosan after an appropriate

solvent is determined for dermal application.

Dose-finding Study of Reproductive and 

Developmental Toxicity Study of Oxybenzone

[2-Hydroxy-4-methoxybenzophenone] 

[131-57-7] Hansen

(1) To determine doses for a full reproductive developmental toxicity study with

oxybenzone, a component of sunscreens, and (2) to assess the possible biological

activity of oxybenzone due to its high volume of use in individuals.

Neurotoxicity Assessment of Cell Phone Radio 

Frequency Radiation (RFR) using Rat and Bovine Brain 

Microvascular Endothelial Cells as Model Blood Brain 

Barrier Systems, PC-12 Cultured Cells, and Whole 

Animal Models Ali

(1) To determine whether power levels of RFR that are emitted from mobile phones

produces any changes in the CNS of mice and rats, and (2) to determine if there are 

disruptions in the blood brain barrier after being subjected to RFR radiation.

Mechanisms of Nevirapine Carcinogenicity 

[129618-40-2] Beland

To determine the mechanism by which nevirapine induces tumors in rats.

The Two-year Carcinogenicity Bioassay of Furan 

in Fischer 344 Rats[110-00-9] Beland

To determine the dose-response relationship for the carcinogenicity of furan in

F344/N/Nctr male rats in a 2-year bioassay.

Determination of Carcinogenic Mechanism for Furan 

in Male Fischer 344 Rats[110-00-9] Doerge

To determine the pharmacokinetic mechanisms, mutagenesis, and hepatotoxicity of

low doses of furan, similar to those ingested by humans from heat processed food.

Range-finding, Mechanistic and Toxicokinetic Studies of

Usnic Acid and Usnea barbata Herb in Fischer 344 Rats

and B6C3F1 Mice. 

[125-46-2, 84696-53-7] Leakey

To complete the standard NTP 14-day range-finding study with both usnic acid and

Usnea barbata administered in feed.

Subchronic Toxicity of Usnea Lichen in Male and Female

Fischer 344 Rats and B6C3F1 Mice 

[84696-53-7] Leakey

To evaluate the subchronic toxicity of Usnea lichen in 90-day toxicology study with

dose administered in chow. Acute toxicity in humans has been reported from the

consumption of the active agent (usnic acid) in the herbal product.

Subchronic Studies of Usnic Acid in Fischer 344 Rats

and B6C3F1 Mice 

[125-56-2] Leakey

To evaluate the subchronic toxicity of usnic acid in male and female Fischer 

344 rats and B6C3F1 mice using a 90-day toxicology studies. Usnic acid had been

shown to be hepatotoxic when consumed by humans in large doses for weight loss.

Toxicity Studies of Glucosamine and Glucosamine/

Chondroitin Sulfate Combination in Obese and 

Lean Zucker Rats[3416-24-8, 9007-28-7] Leakey

To investigate the potential toxicity of glucosamine and glucosamine/chondroitin

sulfate combinations, administered by oral gavage in male rats.

Perinatal Carcinogenicity of Drug Combinations Used

to Prevent Mother-to-Child Transmission of HIV 

[30616-87-1, 134678-17-4] Beland

To determine the carcinogenicity, genotoxicity and metabolism of antiretroviral drug

combinations administered to mice transplacentally and perinatally. The studies

include 14-day range-finding and 2-year chronic in pregnant C57BL6N females and in

B6C3F1 hybrid offspring.

Toxicity Studies of Combination of AIDS Drugs 

in p53(+/-) Transgenic Mice 

[30616-87-1, 134678-17-4] Leakey

(1) To evaluate the potential toxicity and carcinogenicity of perinatal and chronic

exposures to AIDS drugs, AZT and 3TC in C3B6F1trp53(+/-) haplodeficient F1

transgenic mice in a range-finding and a 6-9 month chronic phase, and (2) to 

evaluate the potential toxicity and carcinogenicity of AZT/3TC/NVP combinations 

in C3B6F1trp53(+/-) in a 9 month exposure in mice.

Developmental Neurotoxicity Assessment of

Acrylamide in Rats: Long-term [79-06-01] Paule

To determine the consequences of long-term exposure to acrylamide on a variety of

developmental milestones and measures of nervous system integrity throughout life.

Genotoxicity and Carcinogenicity of Acrylamide 

and its Metabolite Gycidamide in Rodents 

[79-06-1, 5694-00-8] Beland

To determine the genotoxicity and carcinogenicity of acrylamide and its metabolite 

gycidamide in male and female Fischer 344 rats and B6C3F1 mice using range-finding, 

subchronic, and 2-year chronic carcinogenicity studies.

Physiological Effects of Bitter Orange in Rats

[94-07-5] Hansen

To determine potential physiological effects of synthetic synephrine as well as an

extract from the botanical Citrus aurantium alone and in combination with caffeine

in rats, with and without exercise.

Potential Developmental Toxicity of Synthetic 

Synephrine and Citrus aurantium Extract in Rats

[94-07-5] Hansen

To determine the potential developmental toxicity of synthetic synephrine, 

Citrus aurantium and possible potentiation of the toxicity by caffeine.

Effect of Topically Applied Skin Creams Containing 

retinyl palmitate (RP) on the Photocarcinogenicity 

of Simulated Solar Light (SSL) in SKH-1 Mice 

[79-81-2] Boudreau

(1) To determine if RP applied to the skin of SKH-1 hairless mice alters the incidence 

of tumors produced by SSL and/or UV light, and (2) to determine the mechanisms of

tumor promotion by RP.

An Evaluation of the Effect of Vehicle Cream on the 

Photocarcinogenicity of RP in SKH-1 Mice 

[79-81-2, 6938-94-9] Boudreau

(1) To determine the stability and homogeneity of RP in the control cream 

used in the dermal Aloe vera study conducted at the NCTR, (2) to evaluate the

Photocarcinogenicity of RP, incorporated in the Aloe vera control cream when applied

to the skins of SKH-1 mice in the absence and presence of SSL, and (3) to determine

the Photocarcinogenicity of diisopropyl adipate which was used as 

the filler in the previous NTP RP dermal SKH-1 study.

Contact Information: Dr. Paul Howard, paul.howard@fda.hhs.gov and Dr. Nigel Walker, walker3@niehs.nih.gov.
70 National Toxicology Program

NIEHS/NIOSH Interagency Agreement — Comprehensive Assessment 

of Occupationally-Relevant Exposures

The NTP is coordinating an effort with NIOSH to better understand worker exposures, identify occupational

health research gaps, and educate workers. Current efforts listed in Table 32 address worker exposures to welding 

fumes, nano-sized materials, food flavorings, bisphenol A, indium compounds, and other industrial chemicals.

Study Principal Investigator Objective and/or Rationale

Administrative Support 

DeBord 

To enable NIOSH scientists to (1) participate in review and oversight of NTP activities, 

and (2) attend NTP-related meetings in Research Triangle Park, NC and Washington, DC.

Assess the Feasibility of an Occupational

Exposure Assessment of Welding Fume

with Emphasis on Mn Compounds 

Hanley

(1) To identify industries (e.g., construction, shipbuilding, railroad, and manufacturing),

companies, and/or unions involved in welding operations where the potential for substantial

manganese exposure exists, for exposure assessments; (2) to develop methods to identify

specific manganese compounds, different valence states, and potential solubility contained

within various welding fumes matrices; and (3) to characterize welding fume exposures

based on welding-associated jobs, tasks, and processes.

Exposure Assessment of Diacetyl and 

Other Flavorings in Food Production

Industries

Curwin

(1) To characterize workplace inhalation exposures to diacetyl in food production industries

that use food flavorings, (2) to document high-exposure activities and processes in the

flavored food production industries, (3) to identify work practices and procedures that 

affect exposure, (4) to document engineering controls, and (5) to field test novel techniques

for both gravimetric and volatile sampling.

Exposure Assessment of

Dithiobisbenzanilide (DTBBA) 

in a Manufacturing Setting

Wurzelbacher

(1) To identify worker populations at increased risk of inhalation and surface exposure 

to DTBBA during a manufacturing process; (2) to develop a NIOSH analytical method for 

quantitatively assessing DTBBA airborne particulate and surface exposures; and (3) to

characterize industry-wide occupational exposures, including total number of workers, 

and evaluate patterns of exposure to DTBBA.

A Pilot Exposure Assessment for 

2-methoxy-4-nitroaniline (2M4N) 

in a Manufacturing Setting 

Wurzelbacher

(1) To identify worker populations at increased risk of inhalation and surface exposure 

to 2M4N during a manufacturing process; (2) to develop a NIOSH analytical method 

for quantitatively assessing 2M4N airborne particulate and surface exposures; and (3) to

characterize industry-wide occupational exposures, including total number of workers, 

and evaluate patterns of exposure to 2M4N.

Assessment of Use of Indium and Indium

Compounds in the Workplace 

Hines

(1) To contact and visit companies to determine indium materials being used, jobs and

processes with potential indium exposure, exposure controls, and indium use trends, 

and (2) to conduct preliminary sampling for indium, if possible.

Exposure Assessment of Engineered

Nanoparticles

Geraci

(1) To identify workplaces engaged in the synthesis, manufacture, and use of engineered

nanomaterials, and (2) to characterize workplace exposure to selected engineered

nanoparticles.

Exposure Assessment of 1-Chloro-4-

(trifluoromethyl) benzene (PCBTF) 

Harper

(1) To identify worker populations at elevated risk of inhalation and surface exposure to

PCBTF during manufacturing processes; (2) to update a previously published analytical

method for quantitatively assessing PCBTF airborne vapors and surface exposures to

allow the use of capillary column chromatography; and (3) to characterize industry-wide

occupational exposures, including total number of workers, and evaluate patterns of

exposure to PCBTF.

A Pilot Exposure Assessment for Ethylene

Glycol 2-Ethylhexyl Ether (EGEHE) in a 

Manufacturing Setting 

Harper

(1) To identify worker populations at elevated risk of inhalation and surface exposure 

to EGEHE during manufacturing processes; (2) to develop an analytical method for 

quantitatively assessing EGEHE airborne aerosols, vapors, and surface contamination; 

and (3) to characterize industry-wide occupational exposures, including total number 

of workers, and evaluate patterns of exposure to EGEHE.

Durability of Nanoscale Cellulose Fibers 

in Artificial Human Lung Fluids 

Stefaniak

To investigate the in vitro durability of nanocellulose materials in artificial lung fluids. 

Data generated from this study will be used to inform larger and more costly in vivo

inhalation studies.

Table 32 . NIEHS/NIOSH Interagency Aggrement on Occupationally Relevant Exposures, FY 2011
NTP Annual Report 2011 71

Study Principal Investigator Objective and/or Rationale

Exposure Characterization and 

Reproductive Health of Men Working 

with BPA in the United States 

Hines

(1) To determine BPA usage in industry,e.g., which industries and jobs use BPA and

which tasks are associated with exposure; (2) to develop air and wipe sampling methods

for BPA using LC-MS (liquid chromatography mass spectrometry) and LC-UV (liquid

chromatography with UV detection); (3) to assess exposure to BPA among workers in

these industries through air, wipe, and urine sample collection. If worker exposures are 

confirmed; (4) to assess the reproductive health of men exposed to BPA in the workplace;

and (5) to determine if there is a relationship between occupational exposure to BPA and

reproductive health.

Contact Information: Dr. Dori Germolec, germolec@niehs.nih.gov and Dr. Elizabeth Whelan, eaw0@cdc.gov

NIEHS/NIOSH Interagency Agreement–Immunotoxicology

The goal of this IAG isto provide support of NTP hazard identification activities aimed at preventing diseases or

adverse effects caused by environmental exposure to chemical or physical agents. These cooperative studies 

continue to improve risk assessment by measuring what constitutes an adverse health effect on the immune

system in humans. The studies, listed in Table 33, evaluate unique cohorts of individuals from professions 

associated with immune-mediated occupational diseases, including asthma, contact dermatitis, allergy to mold

spores, chronic beryllium disease, and allergic rhinitis. These cohorts are being studied for a number of endpoints

including impact of genetic polymorphisms on development of inflammatory disease and clinical outcomes, and 

identification of unique immunological biomarkers for disease. The NIOSH Laboratory for Occupational Genomics 

serves as a resource for obtaining samples from individuals with occupational and occupationally related diseases.

Study Principal Investigator Objective and/or Rationale

Chronic Sinusitis and Mold Exposure 

Beezhold

(1) To investigate the role of fungi in chronic sinusitis, and (2) to determine if the prevalence 

of fungal sensitization is different to other seasonal allergens in chronic rhinosinusitis

patients. To date, the chronic rhinosinusitis and control patients have been recruited 

and tested. The dataset has been collected and statistically analyzed. 

Heading off Environmental Asthma 

in Louisiana

Beezhold

(1) To evaluate the effectiveness of a novel asthma case management intervention

among children with asthma after Hurricane Katrina in New Orleans, and (2) to assess

total immunoglobulin E (IgE) and mold specific IgE in asthmatic children before and after 

intervention. This project is completed. All sera have been screened and the dataset has

been statistically analyzed.

NIEHS Agricultural Pesticide Study 

Beezhold

To evaluate allergic sensitization in a cohort of 677 farmers with or without pesticide

exposures. To date, 677 farmers have been recruited and the serum total IgE and specific

IgE quantified using Phadia ImmunoCap. Serum concentrations of the fungal mycotoxin,

deoxynivalenol, have been additionally quantified. The second phase of the study has

begun and includes the evaluation of a cohort of >1,500 farmers located in Iowa and 

North Carolina for allergic sensitization.Currently, all farmer sera have been screened for 

total IgE and specific IgE measurements will be in 2012.

A Marker for Aspergillus terreus Exposure

Beezhold

To develop new and improved methods for detecting fungal exposure. In this project 

the emerging opportunistic fungal pathogen,A. terreus, has been used as a model

fungal species. Terrelysin, a cytolysin and potential biomarker of fungal infection, was 

characterized and a recombinant protein produced. The recombinant terrelysin was then

used to immunize mice to produce terrelysin-specific mAbs. Seven specific mAbs were

produced and used to characterize the production of terrelysin. mAb binding sites are 

currently being determined in epitope mapping experiments. The development of a 

sensitive immunoassay that can be used in the serological detection of this biomarker 

is anticipated.

Table 33. NIEHS/NIOSH Interagency Agreement on Immunotoxicology Studies, FY 2011
72 National Toxicology Program

Study Principal Investigator Objective and/or Rationale

Animal Model for Airway Exposure to 

Dry Fungal Aerosols 

Beezhold

To develop a murine model of dry fungal exposure to better mimic natural human

exposures to fungi. An acoustical generation system has been developed. In preliminary 

experiments, histopathology and BALF analysis demonstrated pulmonary deposition of

conidia following dry fungal bioaerosol exposure.Currently, the system is being optimized

for the generation of fungal and hyphal fragments. Once the system has been fully

characterized, experiments are planned to further characterize the immune responses

associated with various fungal species that frequently colonize water-damaged buildings

or are occupationally relevant.

Characterization of Fungal Diversity in the 

Indoor Built Environment

Beezhold

To investigate and characterize the diversity of fungal bioaerosols in the indoor built

environment using large-scale ribosomal RNA (rRNA) sequencing in collaboration with

the Kansas City Safe and Healthy Home Partnership Project. In preliminary studies, various

methods of extraction were compared and tested on occupational dust samples. Currently, 

bulk dust samples derived from the project are being sequenced. Air samples collected 

with NIOSH two stage samplers will also be processed in 2012.

The Role of Genetic Variation in

Environmental and Occupational Diseases

– Irritant Contact Dermatitis (ICD) 

Yucesoy

(1) To investigate whether the 24-hour irritant patch test is predictive of occupational

hand dermatitis caused by high exposure to hand washing in health care workers, and

(2) to investigate an association between genetic variations in specific candidate genes

(with emphasis on variants of cytokines, MHC region, antioxidant enzyme genes and

genes related to skin barrier integrity) and irritation threshold levels of the subjects with

development of ICD. This study is in collaboration with Case Western Reserve and West

Virginia Universities. Subject recruitment and genotyping have been completed. Data

analyses have been initiated.

The Role of Genetic Variation in

Environmental and Occupational Diseases

– Allergic Contact Dermatitis (ACD) 

Yucesoy

(1) To investigate genetic factors in individuals predisposed to develop ACD, specifically

induced by nickel, and (2) to investigate genetic factors involved in the development of

ACD in individuals sensitized to weak allergens, individuals sensitized to allergens that 

require metabolism in the skin, and individuals who react to more than 3 allergens of

the standard screening series. This study is in collaboration with Case Western Reserve

University and Dartmouth-Hitchcock Medical Center. Subject enrollment and sample

processing are currently underway.

The Role of Genetic Variation in

Environmental and Occupational Diseases

– Occupational Asthma (OA)Yucesoy

(1) To investigate whether genetic variations in specific candidate genes (e.g., cytokine, 

MHC region, antioxidant enzyme genes) are associated with asthma induced by

diisocyanates, and (2) to investigate potential associations between genetic variations in

candidate genes and occupational asthma caused by low molecular weight agents. This

project is in collaboration with the Universities of Montreal and Cincinnati. Genotyping 

of antioxidant enzyme and MHC region variations have been completed and the dataset

has been analyzed.

The Role of Genetic Variation in

Environmental and Occupational Diseases

– Chronic Beryllium Disease 

Yucesoy

To investigate the contribution of genetic variations in the MHC region to the development 

of beryllium sensitization and chronic beryllium disease. This study is in collaboration with

National Jewish Medical and Research Center. Genotyping efforts are currently underway.

Investigations into Health Effects Caused

by Exposure to Indoor Air Reaction

Products(supportive animal studies) 

Wells, Anderson

(1) To identify and measure the reaction products of gas-phase compounds present in the

indoor environment, especially oxygenated organics; (2) to further develop and validate a 

novel in vitro exposure method utilizing realistic indoor chemistry scenarios to expose cells

and tissues to these indoor air reaction products; (3) to complete both in vitro and in vivo

assays to assess adverse health effects caused by indoor air reaction products; and (4) to

further investigate the role of structurally similar indoor air chemicals present in mixtures 

in the indoor environment.

Immunological Mechanisms of

Occupational Rhinitis Induced by

Diisocyanate Exposure (supportive

animal studies and field studies) 

Johnson

(1) To use specific inhalation challenge with suspected diisocyanates to diagnose allergic

rhinitis in workers from Canada and Spain who are exposed to diisocyanates and clinically

phenotyped for upper and lower airway disease; (2) to collect nasal mucosal samples from

worker with diisocyanate rhinitis and use whole genome approaches to study mechanisms

of disease; (3) to identify molecular targets in the human nasal mucosa for biomarker 

development and validation using the NIOSH toluene diisocyanate rhinitis mouse

model; and (4) to develop the mucosal sampling technique and identified biomarker(s)

into a simple and non-invasive tool for worker health surveillance and early diagnosis of

sensitization/disease.
NTP Annual Report 2011 73

Study Principal Investigator Objective and/or Rationale

Diisocyanate Monoclonal Antibody

Production and Characterization

(improved methods) 

Siegel

(1) To develop mAbs that recognize methylene diphenyldiisocyanate conjugated protein

as a potential biomonitoring and research tool; (2) to test the specificity of the mAbs; and

(3) to evaluate the mAbs for potential use in immunoassay-based biomonitoring.

Alternative Methods for Chemical Allergen

Identification and Assessment

Siegel

(1) To develop an amine-based probe for kinetic assessment of chemical binding

(haptenation) that will complement the thiol-based probe previously reported by NIOSH;

(2) to assess potential for inclusion of a metabolic activation step in our kinetic-based

assays for identification of prohaptens; and (3) to expand the compilation of chemical

allergens assessed by kinetic electrophilic reactivity analyses for comparison to reported

allergenic potencies (murine local lymph node assay EC3 values).

Analysis of Mycotoxins in Dust Samples 

from a Water-damaged Building

Park

To develop the in-house laboratory protocol for analysis of macrocyclic trichothecene

mycotoxins using GC tandem MS and to analyze existing dust samples from a water-damaged

building. By modifying an existing method, a protocol that substantially improved

extraction efficiency of mycotoxins in samples was developed. With the developed

protocol, samples harvested from culture media with 7 different strains of Stachybotrys

chartarum, building material samples inoculated with these fungi, and floor dust samples

from water-damaged buildings for the mycotoxins were analyzed. In the course of

the study, a potential matrix effect in the GC/MSMS analysis was detected which can 

substantially affect accurate quantification of the mycotoxins in samples. Current 

efforts involve further investigation of the potential matrix effect and development 

of a 13C isotopically-labeled satratoxin internal standard to further improve the protocol.

Contact Information: Dr. Dori Germolec, germolec@niehs.nih.gov and Dr. Don Beezhold, zec1@cdc.gov

NIEHS/EPA Interagency Agreement – The Phthalate Initiative

Di(2-ethyl)hexyl phthalate (DEHP) and other phthalates have been nominated to the NTP for testing. To address

these nominations, the NIEHS and EPA signed The Phthalate Initiative IAG in June 2008, which was renewed in

July 2009 and 2010, and is now in the last year of funding. Many aspects of this IAG would fall under nominations

previously approved by the BSC for the study of peroxisome proliferators (begun in the 1990s), the nomination of

DEHP by the FDA in 2004, and the critical data need highlighted in the NTP Monograph on DEHP issued in 2006. 

These studies will clarify how the peroxisome proliferator-activated receptor alpha (PPAR ) develops in the rat 

and its relationship to DEHP-related cancer and other developmental toxicities. The studies will also provide 

critical data for future mixture studies using various models to inform on potential risks for toxicity over time 

and during development. Recent data have indicated that because phthalate esters have similar modes of 

action in utero, they show dose-addition when administered in combination. Therefore, it would be appropriate 

to consider cumulative risk for the class, since human subjects (including fetuses) are typically exposed to 

multiple phthalates.

The initiative has two major specific aims: 

(1) Undertake an ontogeny study of PPAR in the rat to determine when the receptor is first expressed in target

tissues. This study will test the hypothesis that PPAR is developmentally regulated in the rat and unlikely to

contribute to toxicity initiated in utero after exposure to DEHP. 

(2) Undertake perinatal phthalate mixture studies. These studies will test the hypothesis that exposures to

mixtures of phthalates, based on their individual potencies, would result in dose-addition for cancer (and

potentially other) outcomes.
74 National Toxicology Program

Based on data generated in 2010 and 2011, using a specific short-term in utero screen developed in the initiative to

evaluate individual phthalates and mixtures in the Harlan SD rat to alter fetal testicular testosterone production, 

phthalates induce changes in specific fetal testicular genes (especially related to steroidogenesis and testicular

descent), and the induction of specific male reproductive tract malformations that have been termed the 

“Phthalate Syndrome.”

Thus far, data indicating a positive response have been obtained from butylbenzyl, di-n-butyl, di-iso-butyl, 

di-ethylhexyl, di-n-propyl, di-pentyl, di-n-hexyl, di-iso-heptyl, di-n-heptyl, di-heptylpropyl, dimethoxyethyl, 

di-cyclohexyl, and 3 different preparations of di-iso-nonyl phthalate (a complex mixture). Brominated DEHP, 

dimethyl, diethyl, and di-isodecyl phthalate, and di-octyl terephthalate were negative in this fetal phthalate 

screen. Specific potency factors have been developed based on this screening information. Thus a five-day screen

using minimal numbers of rats has been developed to discriminate positives from negatives. The concordance

with long-term multigenerational and pubertal effects is 100%. In the mixture studies the IAG also examined the 

potential effect of a negative phthalate ester (diethyl phthalate) on the activity of a positive ester,since they use 

the same esteras activity to produce monoesters for excretion. Diethyl phthalate had no effect either on its own 

or in the mixture.

These data would guide selection of individual studies to investigate potential short-term biomarkers for toxicity, 

which in turn would support future perinatal bioassays and phthalate mixture work. The NTP has now designed

both perinatal and standard bioassays to evaluate developmental and cancer potential from different exposure

paradigms for DEHP, which commenced in-life evaluation in early 2011. This information will be valuable in

assessing potential cumulative risk of phthalates, as has been advocated by a recent National Academy of

Sciences committee report and efforts by EPA’s IRIS program.

Work has also progressed on the ontogeny study and the development of a genomic signature for phthalates

that induce reproductive toxicity in utero, where the IAG is evaluating the potential role of activation of PPAR

pathways in the disruption of male sexual differentiation and testis development using custom-designed RT-PCR

plates. These plates allow determination of relative mRNA expression of PPAR , PPAR , PPAR , and several genes 

activated by PPAR including CYP4a1 on a single sample simultaneously. For six positive phthalates examined 

so far, none of the PPAR genes, CYP4a1, or any other genes “downstream” of PPAR are activated by any of

the phthalates studied to date at a single high dosage level (750 mg/kg/d GD 14-18) or at any dose in the dose

response studies. PPARs and mRNA were expressed in GD18 fetal testes whereas PPAR was not detected. 

These results do not support the hypothesis that PPAR is involved in induction of the Phthalate Syndrome in

rats. These results also are supported by the observation that administration of a potent PPAR agonist (Wyeth

14643) did not reduce anogenital distance or testicular testosterone levels in GD21 male rats, whereas the positive

control di-iso-butyl phthalate did reduce the measures. Furthermore, maternal liver PPAR genes were induced in

the Wyeth treated animals indicating a lifestage specific effect. These studies will be further extended looking at

potential PPAR induction in the fetal liver.

The IAG has identified approximately 10 genes that are part of the genomic signature for phthalates that are

consistently altered on the PCR array. The lack of response in the detection of PPAR is of note since thisis in direct

contrast with the claim that PPAR is part of the HTS “signature” for reproductive toxicity described by ToxCast,

which included phthalate esters. In addition, the IAG has initiated a project in which they are interrogating the 

effects of phthalates on additional pathways using standard RT-PCR arrays to expand the genomic signature

upstream. The IAG has had about 1000 additional genes identified so far from about 10 different pathways 

using seven different phthalate esters. During 2011, the IAG has produced four peer reviewed publications 

and four abstracts.

Contact Information: Dr. Paul Foster, foster2@niehs.nih.gov
NTP Annual Report 2011 75

Much of the research conducted in support of the NTP’s high throughput screening (HTS) program is coordinated 

through a Memorandum of Understanding (MOU), High Throughput Screening, Toxicity Pathway Profiling and 

Biological Interpretation of Findings, first signed on February 14, 2008. Information about the MOU is available

at http://ntp.niehs.nih.gov/go/28213. Through this MOU, NTP formally entered into a partnership with the 

National Human Genome Research Institute’s (NHGRI) NCGC and the U.S. EPA’s National Center for Computational

Toxicology located within the Office of Research and Development. This interagency partnership is the basis for 

the U.S. Tox21 Program, developed in response to the National Research Council’s 2007 report Toxicity Testing 

in the 21st Century: A Vision and a Strategy. A central component of this MOU is exploration of quantitative high

throughput screens (qHTS) and tests using phylogenetically lower animal species (e.g., fish, worms), as well as

high throughput whole-genome analytical methods to evaluate mechanisms of toxicity. The ultimate goal of

Tox21 isto provide the data generated by these new tools to risk assessors for use in their mission to protect 

human health and the environment. The MOU was amended in 2010, adding the FDA, which bringsto the 

Tox21 partnership its experience in human diseases, animal models of human disease, and expertise in toxicity

pathway analysis and computational toxicology. FDA’s active participation is in recognition of its commitment 

to developing new methods to evaluate the toxicity of the substances it regulates.

The goals of Tox21 and the HTS program are the same (i.e., prioritization for further in-depth toxicological 

evaluation, identify mechanisms of action, predictive toxicology). The results of this collaborative effort should

yield test methods for toxicity determination that are more scientifically and economically efficient, and models

for risk assessment that are more biologically and mechanistically based. This approach should ultimately reduce

or replace animals in regulatory testing and is anticipated to occur in parallel with an enhancement of the ability

to evaluate the large numbers of chemicals that currently lack adequate toxicological evaluation. This interagency 

partnership makes it possible to pool resources to overcome the resource limitations of a single agency, build on

existing expertise, and avoid the need to create a new administrative and support structure.

In Tox21 Phase I (FY 2006 through FY 2010), a library of approximately 2,800 compounds provided by the NTP 

and EPA to the NCGC was tested in approximately 100 qHTSs. These assays broadly evaluated the ability of

compounds (1) to induce cytotoxicity, apoptosis, DNA damage, changes in methylation status, mitochondrial

toxicity, and up-regulation of various stress response pathways (e.g., antioxidant, hypoxia, heat shock) in a variety 

of cell types or (2) to act as an agonist or antagonist for 12 different nuclear receptors including the estrogen

and androgen receptors. Also evaluated in qHTS was the extent of differential sensitivity exhibited in terms 

of induction of cytotoxicity and/or apoptosis by 87 densely sequenced human lymphoblastoid cell lines (the

HapMap project CEPH panel, which refers to the collection of cell lines established from Utah residents with

ancestry from northern and western Europe) exposed to 240 toxic chemicals.

Results from Phase I were carefully evaluated during FY 2011 and used to determine the approach for Tox21 

Phase II at the NCGC. Major emphasis was placed on acquiring the most robust assays (both full length and partial 

nuclear receptor assays) for evaluating the ability of environmental chemicals to interact with the estrogen and/or

androgen receptors. In addition, other nuclear receptor assays, assays measuring the ability of chemicals to induce

various stress response pathways (e.g., antioxidant, DNA damage, heat shock, hypoxia), and assays measuring 

specific toxicity endpoints such as mitochondrial membrane changes, caspase activation, and NFkB upregulation

were included in the queue for Phase II screening during FY 2012. This screening isto be conducted in a new,

high-speed robotics screening facility dedicated at the NCGC in March 2011 and built with funds provided by 

the NIEHS/NTP. This robot facility was designed to screen a 10,000 compound library in qHTS. 
76 National Toxicology Program

Phase II screening is scheduled to begin early in FY 2012, after the completion of the Tox21 10,000 compound

library. EPA, NTP, and NCGC will each supply approximately one-third of the compounds for this library. The 

compounds cover a wide variety of classifications and include consumer products, food additives, chemicals 

found in industrial processes, and human and veterinary drugs. A complete list of the compounds is publicly

available at http://www.epa.gov/ncct/dsstox/. Chemical analyses by the Program Operations Branch within DNTP

to determine the identity, purity, and stability of all compounds in this library were initiated in FY 2011 and 

will continue through FY 2012. Testing was started on a subset of these compounds (~700) in Phase II of EPA’s 

ToxCastTM Program (http://www.epa.gov/ncct/toxcast/). The data from all these assays, along with full chemical

characterization and assay protocol details, are being deposited into publicly accessible, relational databases such

as the National Library of Medicine’s PubChem (http://pubchem.ncbi.nlm.nih.gov/), EPA’s ACToR 

(http://www.epa.gov/ACToR/) and NTP’s CEBS (http://www.niehs.nih.gov/research/resources/databases/cebs/).

In FY 2011, the differential cytotoxicity response of cells from humans was evaluated by screening ~1100 densely

sequenced lymphoblastoid cell lines against 180 toxic chemicals. This study was designed to measure inter-individual

differences in sensitivity to environmental toxicants, based on genotype. The data generated by this study will be

evaluated during FY 2012. 

Non-mammalian models – Caenorhabditis elegans

The NTP WormTox Facility develops toxicological assays using the nematode C. elegans and evaluates their utility

as medium throughput screening tools. The use of C. elegans is consistent with NTP’s strategy to reduce the 

number of mammals used in testing. Several toxicology assays and statistical analysis tools have been developed 

to monitor C. elegans feeding, growth, reproduction, and movement after chemical exposures. In FY 2011, 

screening began of the U.S. EPA’s ToxCast Phase II chemical library, which consists of 700 compounds, using the 

C. elegans growth assay. The studies will continue through FY 2012. Also during FY 2011, development of a new 

assay to monitor in vivo gene expression changes in transgenic C. elegans after exposure to various classes of

environmental toxicants was initiated. Table 34 lists ongoing and completed C. elegans studies.

Chemical CASRN

Hexafluorosilicic acid 16961-83-4

Sodium fluoride 7681-49-4

Sodium hexafluorosilicate 16893-85-9

EPA ToxCastTM Phase 1 320 compounds List of chemicals available at 

http://www.epa.gov/ncct/toxcast/chemicals.html EPA ToxCastTM Phase II 700 compounds

The NCGC BioPlanet

No one comprehensive and uniform resource covers all known annotations of cellular pathways and no single

platform allows integrated browsing, retrieval, and analysis of information from the many existing individual

pathway resources; therefore, the NCGC (with support from the NTP) built an integrated pathway resource that

hosts information on ~1100 human pathways from manually curated and publicly available resources. The NCGC

BioPlanet (http://www.ncgc.nih.gov/pub/bioplanet/) complements this pathway warehouse by allowing easy

browsing, visualization, and analysis of the universe of pathways. 

Table 34. Completed and Ongoing C. elegans Studies During FY2011
NTP Annual Report 2011 77

Toxicogenomics Studies

The NTP is working to bring the latest toxicogenomics technology into its testing program to help revolutionize 

the way NTP conducts its studies. Toxicogenomics examines how the entire genetic structure, or genome, is

involved in an organism’s response to environmental toxicants. It applies gene and protein technologies to

environmental medicine by studying the effect of toxicants on gene activity and specific proteins produced by

genes. This information could be useful to identify biomarkers of disease and exposure to toxic substances and 

for understanding individual genetic susceptibilities. 

Preliminary toxicogenomic studies suggest that gene expression often is predictive for phenotypic alterations.

The NTP is interested in determining if differential gene expression (DGE) analysis can provide indicators of

toxicity at earlier time points and at lower doses than possible with traditional toxicology parameters. DGE 

may provide more than a genotypic link to a morphology, because it is expected to provide insights into the 

pathogenesis of the disease and how different rodent models respond to toxicants.

Perhaps the most exciting potential of toxicogenomics is the possibility to identify biomarkers of exposure or

biomarkers of effect. Changes that can be found in easily obtainable samples (blood and urine) could then be

monitored in clinical studies. When the technology is validated, it will allow repeated sampling during long-term 

NTP studies to determine whether chemical exposures can be detected or whether developing cancers will

provide a genetic signature. 

The NTP is currently evaluating study conditions that may contribute to gene expression (e.g., animal and tissue

variability), best methods of tissue sampling, and establishing standards for conducting toxicogenomic studies 

under laboratory conditions. A long-term goal of the NTP isto identify more accurate methods of predicting

carcinogenicity, because current NTP carcinogenicity studies take four to five years to complete and are costly. 

Planned or ongoing NTP toxicogenomic studies are listed in Table 35.

Chemical CASRN Species/Strain Study Route Study Length Platform

1,2,3,7,8-Pentabromodibenzofuran

1,2,3,7,8-Pentachlorodibenzofuran

2,3-Dibromo-7,8-dichlorodibenzo-p-dioxin

2,3,4,7,8-Pentabromodibenzofuran

2,3,4,7,8-Pentachlorodibenzofuran

2,3,7,8-Tetrabromodibenzofuran

107555-93-1

57117-41-6

50585-40-5

13116-92-2

57117-31-4

67733-57-7

Mice: B6C3F1 Gavage 3/11 days Microarray 

qPCR

Acetochlor 

Bisphenol A 

Carbaryl 

N,N-Dimethyl-p-toluidine

Flusilazole

Perfluorooctane sulfonic acid

2,5-Pyridinedicarboxylic acid, dipropyl ester 

Simazine 

Triclosan

34256-82-1

80-05-7

63-25-2

99-97-8

85509-19-9

335-67-1

136-45-8

122-34-9

3380-34-5

Rats: Harlan

Sprague–Dawley

Gavage 4 days Microarray

Aflatoxin B1 1162-65-8 Rats: F344/N Feed 90 days NextGen 

Sequence,

qPCR

Aloe vera extract NA Rats: F344/N Oral 2 years qPCR

Antimony trioxide 1309-64-4 Mice: B6C3F1 Oral 2 years Microarray

Bromodichloroacetic acid (BDCA) 71133-14-7 Rats: F344 Oral 2 years qPCR

Effect on loss of ROR- and TAK1 in

knockout mice on metabolomics profile – 

no chemical

NA Mouse: C57/BL6 NA NA NMR, LCMS/MS

Table 35. Toxicogenomics studies (Ongoi ng, Completed, Initiated)
78 National Toxicology Program

Mining the NTP Tissue Archives for Gene Signatures

The NTP archives contain stained histopathology slides, paraffin tissue blocks, formalin-fixed tissues and 

organs, and selected frozen tissue from over 2,000 studies including toxicity, carcinogenicity, immunotoxicity, 

reproductive, and developmental studies. In FY 2010, NTP initiated pilot studies to evaluate the extent to which 

gene expression signatures can be reliably derived from the molecular analysis of tissue samples collected from

the laboratory animals used in NTP’s toxicological studies and stored as formalin-fixed, paraffin-embedded

(FFPE) tissues in the NTP archives. Signature expression profiles are critical sets of altered transcripts or proteins

that distinguish toxicity and disease from a comparable normal state. Two sets of studies were conducted during

FY 2011. In one study, RNA was extracted from a limited number of four-year old paraffin blocks of liver and a 

selected set of genes evaluated for expression using RT-PCR. The results compared favorably with those obtained

for the same genes using RNA from fresh samples, indicating that this technology could be applied to FFPE 

samples. In a second study, a different transcript profiling technique–qNPA–was used to compare transcripts 

from fresh or FFPE brain and liver. Results again demonstrated the usefulness of archival tissue blocks for gene

profiling. Future studies will make a wider and more systematic query of NTP archival materials of different organs

and storage times for their value in transcript profiling.

Chemical CASRN Species/Strain Study Route Study Length Platform

Effect of diet on liver transcriptome – 

no chemical

NA Rats: Wistar Han NA 90 days Microarray

Ginkgo biloba extract NA Mice: BC3F1 Oral 2 years Microarray

Oxybenzone 131-57-7 Rats: Harlan SD Gavage 90 days Microarray

BDE 47

BDE 71

BDE 99 

DE 153 

BDE Mixture

5436-43-1

32534-81-9

60348-60-9

68631-49-2

Mixtures

Rats: Wistar Han

Mice: B6C3F1/N

Gavage 14/90 days Microarray

Pentabromodiphenyl oxide (technical) 

(DE 71)

32534-81-9 Rats: Wistar Han Gavage GD 6 – PD 21 Microarray

2,3,-Pentanedione 600-14-6 Rats: F344 Inhalation 3 weeks Microarray

Sodium arsenite 7784-46-5 Human/RWPE-1 

prostate cells

In vitro 30 weeks NextGen 

Sequence

Spontaneous Liver and Lung tumors NA Mice: B6C3F1 NA NA Microarray

Trimethyltin hydroxide 56-24-6 Mice: CD-1 Intra-peritoneal

injection

2 days qNPA

Vinylidene chloride 75-35-4 Rats: F344/N

Mice: B6C3F1

Oral 2 years Microarray
NTP Annual Report 2011 79

DrugMatrix®

Related to the goal of developing analysistools and approaches to allow an integrated

assessment of HTS endpoints and associations with findings from traditional toxicology

and cancer models, the NTP acquired DrugMatrix,® a toxicogenomics reference 

database, the accompanying extensive frozen tissue archives, and the informatics

system. This resource will expand NTP’s ability to develop predictive models for 

toxicological effects based on gene signatures, provide additional tools for linking

in vitro data to in vivo gene signatures and disease outcomes, and provide additional

tissue samples for NextGen-based investigations. In late FY 2011, DrugMatrix® 

and its companion, automated analysistool, ToxFX®, were made accessible to the 

international scientific community in beta release format at https://ntp.niehs.nih.

gov/drugmatrix and https://ntp.niehs.nih.gov/toxfx/, respectively. To date, over 100 

researchers have registered to use the DrugMatrix® database. In addition, the data and 

biological samples from DrugMatrix® are a focal point in a number of collaborations

between DNTP scientists and research groups from the FDA’s National Center for 

Toxicological Research, the City of Hope, Stanford University, Boston University, 

U.S. EPA, Health Canada, Abbott Laboratories, Eli Lilly and Company, SAS, Maastrich University, GeneData, 

University of Massachusetts, and University of North Dakota. The goal of these collaborationsisto use the 

DrugMatrix® data to better understand the molecular underpinnings of disease and toxicological pathology.

Host Susceptibility Program

Host Susceptibility Aging Cohort

Genetic and epigenetic differences between individuals within the human population are believed to be the 

basis for individual susceptibility to environmental stressors, including idiosyncratic drug toxicities. However,

at present, environmental and drug safety assessments are conducted with a small number of commonly used

animal models with limited genetic diversity that is insufficient to evaluate the influence of individual genetic 

differences on chemical and drug toxicity and is of limited value in extrapolating to human toxicity and disease.

To improve the extrapolation of results from rodent modelsto human hazard identification and risk assessment,

an aging study to produce a benchmark reference data set was started in FY 2009 using 10 inbred strains of mice

(including the 8 used in the Collaborative Cross; see Diversity Outbred mice below) to provide data on survival

rates, spontaneous disease incidence, histopathology, hematology, clinical chemistry, quantitative trait loci, and 

behavioral observations, as well as for collecting tissue/fluids for future genetic/genomic/metabolomic analysis. 

The strains selected include classical and wild-derived inbred strains that represent the greatest degree of genetic 

diversity in the mouse genome that is known at the present (based upon the analysis of the NTP-Perlegen

genotyping initiative data). This study is expected be completed in FY 2012 and study results will continue to be

analyzed during FY 2013; the results will aid scientists in selecting the most appropriate mouse strains for study 

and aid in the extrapolation of response to toxicant exposure and prediction between species.

Diversity Outbred (DO) Mice and Population Based Models for Toxicology and Disease

Population-based models are required in order to quantify the impact of genetic and epigenetic diversity 

(based upon sequence variation) on toxicity and associated disease. From ADME studies initiated in FY2009 using 

a panel of 18 inbred strains (including the 8 used in the Collaborative Cross), we had identified significant inbred

strain variation (10-36X kinetic differences) in both area under the curve (AUC) and clearance from blood and 

bone marrow using [14C]-benzene following a single oral dose. In particular, the difference in clearance of 

[14C]-benzene, including metabolites, from the bone marrow suggests that significant population based 
80 National Toxicology Program

differences might be observed for benzene-induced genotoxicity and hematotoxicity. Using the data from 

the NTP-Perlegen sequencing effort, researchers at The Jackson Laboratory and UNC-Chapel Hill have performed 

a “collaborative cross” using an 8-way parental outcross that incorporated 90% of all the known genetic 

diversity observed in all inbred laboratory and wild-derived strains. From this complex outcross, more than 

300 advanced intercross lines are being produced. From a large sample of male and female mice from the 

4th and 5th generation, a large randomized breeding program was created to produce the diversity outbred 

mouse population with low inbreeding coefficient. Due to the large number of meiotic recombination events, 

the genomes of the 8 parental lines (included in the aforementioned ADME studies panel) were segregated into 

small segments resulting in minor allele variants of no less than 10% across the genomes. This feature significantly 

improves the power to detect genotype-phenotype differences in this genetically diverse random breeding 

population. Research was initiated on this outbred stock for utility in the NTP research and testing program in

late FY2011 using 28-day old DO male mice exposed to inhaled benzene (0, 1, 10, or 100 ppm; 75 mice/group) 

for 5 days/week for 4 weeks in two independent studies. Benzene is a known mouse and human toxicant and 

carcinogen with a significant published literature that aids data correlation between mouse and human toxicity. 

Pre-exposure differences in blood cell and red blood cell micronuclei values in samples from 300 male DO male

mice were significant. A repeat study will occur in FY 2012. The data from two independent studies of DO mice

will be evaluated using computation toxicology and quantitative genotype-phenotype analysis.

Host Susceptibility to Ionizing Radiation

Genetically engineered mouse models(GEMM) have been effectively used as surrogates for studying human 

cancers and for determining the mode/mechanistic basis of action for carcinogens. In FY 2009, six different 

p53 haploinsufficient F1 mouse strains were created by outcross of females from selected strains (A/J, BALB/C, 

BTBR.T/J, C3H/HeJ, DBA2/J, or 129S1.SvlmJ), based upon their genetic diversity in DNA double strand break 

repair, to B6.129-Trp53tm1Brd N12 homozygous null males. At 8 weeks of age, the F1 progeny were exposed to

0, 3, or 6 Gy of ionizing radiation. A dominant feature of carcinogen-induced tumors in this mouse model is the 

loss of the Trp53 wildtype allele and genome wide loss of heterozygosity (LOH) often associated with loss of

tumor suppressor gene function in human cancer. The pattern of LOH suggests significant changes in gene 

copy number variation (gene deletions and gene duplications). This project is focused on the further 

development of GEMM of tumor suppressor gene deficient mice for predicting carcinogenicity, determining 

the mode of action, and the presumptive risk of environmental exposures to humans. An additional aim is the 

identification of causally related genes and the specific functional allelic and copy number variants within inbred

strains of mice that modify carcinogen potency. Preliminary results indicate significant differences across the six 

different p53 haploinsufficient F1 strains employed in terms of survival, median time to tumor, tumor phenotype

spectrum, and tumor prevalence. The results implicate dysregulation of DNA strand break repair. In FY 2012, a 

tiered approach will be used to define system level differences in toxicogenomics associated with differences in

strain susceptibilities and to better define a short-term cancer bioassay for research and testing. Determination 

of the allelic variants of genes causally related to DNA damage and repair with altered function may be critical 

to understand the differences in risk due to exposure to environmental mutagens.
NTP Annual Report 2011 81

NIH Mouse Methylome Project

An individual’s response to exposure-related toxicity and concomitant disease is influenced at the genomic 

level by genetic, epigenetic, gene-gene interactions (intrinsic factors), and interaction with the environment 

(extrinsic factors). Individual DNA sequence variation does not account for all of the heritability for susceptibility

to toxicity or diseases such as asthma, cancer, and diabetes. An intrinsic factor that quantitative and molecular 

geneticists believe is the basis for the observed “missing heritability” is the methylome, an individual’s 

genome-wide methylated CpG sequence pattern. The methylome (a component of the epigenome) may 

be the major epigenetic modifier of the susceptibility to cancer and other chemical exposure-related diseases. 

Presently, there is no mouse reference database for the methylome. The absence of a methylome reference 

database for the mouse significantly handicaps understanding of the mouse model in toxicology and 

environmentally related diseases and in designing and conducting hypothesis based genetic and epigenetic

research studies to understand the associated mechanisms. To produce a methylome reference database based 

for 5-methylcytosine genome wide patterns in mice relevant to the NTP studies, we outcrossed C56BL/6N

females to C3H/HeN males and vice versa to produce F1 siblings under NTP dietary and environmental 

specifications. Five tissues (brain, liver, cardiac and skeletal muscle, brown and white fat, epididymal sperm) 

were collected at the average age mice would start an NTP subchronic toxicity study and flash-frozen for DNA/

RNA isolation and sequencing of liver (starting in FY 2011 and continuing through FY 2012) from each parental

line and outcross and gender are being used to (1) determine genome wide methylated CpG sites by deep

sequencing of bisulfite treated genomic DNA to determine sequence context and cytosine methylation 

variation (BIS-Seq) and RNA (RNA-Seq) and (2) fractionate DNA sequences using differential restriction and/

or affinity capture (MMDE-seq) to enrich for methylated DNA sequences. Together, these tools allow targeted

interrogation of CpG regions of interest using bioinformatic data mining tools and a determination of within 

and between strain variations as well as parent of origin differences. These data will allow creation of a definitive 

map of the mouse liver methylome from these parental strains (C57BL/6N and C3H/HeN) and their F1 hybrid

offspring that exhibit dramatically different rates of interstrain and sex dependent spontaneous liver cancer 

that often confound NTP 2-year toxicology and carcinogenesis studies.
Agency Staff and Contact Information

NTP staff are listed alphabetically by agency

NCTR/NTP Paredes, Angel (870) 543-7949 angel.paredes@fda.hhs.gov

Babb, Amy (870) 543-7697 amy .babb@fda.hhs.gov Paule, Merle (870) 543-7147 merle.paule@fda.hhs.gov

Beland, Frederick (870) 543-7205 frederick.beland@fda.hhs.gov Pogribna, Marta (870) 543-7681 marta.pogribna@fda.hhs.gov

Boudreau, Mary (870) 543-7526 mary .boudreau@fda.hhs.gov Pogribny, Igor (870) 543-7096 igor.pogribny@fda.hhs.gov

Brakebill, Nancy (870) 543-7785 nancy .brakebill@fda.hhs.gov Sepehr, Estatira (870) 543-7902 estatira.sepehr@fda.hhs.gov

Chen, Xinping (870) 543-7702 xinping.chen@fda.hhs.gov Siitonen, Paul (870) 543-7656 paul.siitonen@fda.hhs.gov

Chidambaram, Mani (870) 543-7368 mani.chidambaram@fda.hhs.gov Shaddock, Joseph (870) 543-7280 joseph.shaddock@fda.hhs.gov

Churchwell, Mona (870) 543-7698 mona.churchwell@fda.hhs.gov Stingley, Robin (870) 543-7350 robin.stingley@fda.hhs.gov

Corlett, Angela (870) 543-7403 angela.corlett@fda.hhs.gov Surratt, Gordon (870) 543-7608 gordon.surratt@fda.hhs.gov

Couch, Letha (870) 543-7137 letha.couch@fda.hhs.gov Thorn, Brett (870) 543-7338 brett.thorn@fda.hhs.gov

Cozart, Christy (870) 543-7602 christy.cozart@fda.hhs.gov Trbojevich, Raul (870) 543-7490 raul.trbojevich@fda.hhs.gov

Cunningham, Candice (870) 543-7886 candice.cunningham@fda.hhs Twaddle, Nathan (870) 543-7178 nathan.twaddle@fda.hhs.gov

Delclos, Barry (870) 543-7372 barry .delclos@fda.hhs.gov Vanlandingham, Michelle (870) 543-7458 michelle.vanlandingham@fda.hhs.gov

Doerge, Daniel (870) 543-7943 daniel.doerge@fda.hhs.gov VonTungeln, Linda (870) 543-7620 linda.vontungeln@fda.hhs.gov

Duffy, Peter (870) 543-7054 peter.duffy@fda.hhs.gov Wang, Cheng (870) 543-7259 cheng.wang@fda.hhs.gov

Fang, Jia-Long (870) 543-7612 jia-long.fang@fda.hhs.gov Weis, Connie (870) 543-7378 connie.weis@fda.hhs.gov

Felton, Robert (870) 543-7504 robert.felton@fda.hhs.gov Willingham, Evelyn (870) 543-7938 evelyn.willingham@fda.hhs.gov

Ferguson, Sherry (870) 543-7589 sherry .ferguson@fda.hhs.gov Woodling, Kellie (870) 543-7491 lkellie.woodling@fda.hhs.gov

Fisher, Jeffrey (870) 543-7703 jeffrey.fisher@fda.hhs.gov

Gamboa, Goncalo (870) 543-7400 goncalo.gamboa@fda.hhs.gov NIEHS/NTP 

Guo, Lei (870) 543-7048 lei.guo@fda.hhs.gov Alden, Charles (919) 541-5722 alden@niehs.nih.gov

Hansen, Deborah (870) 543-7480 deborah.hansen@fda.hhs.gov Allen, Bonnie (919) 541-3449 allenb@niehs.nih.gov

Heflich, Robert (870) 543-7493 robert.heflich@fda.hhs.gov Allen, David (919) 316-4664 allen7@niehs.nih.gov

Heinze, Thomas (870) 543-7468 thomas.henize@fda.hhs.gov Alper, Scott (919) 541-4377 alpers@niehs.nih.gov

Howard, Paul C. (870) 543-7672 paul.howard@fda.hhs.gov Andrews, Danica (919) 541-2595 andrewsda@niehs.nih.gov

Imam, Mohammed Sadiq (870) 543-7127 mohammed.imam@fda.hhs.gov Auerbach, Scott (919) 541-4505 auerbachs@niehs.nih.gov

Keasling, Adam (870) 543-7441 adam.keasling@fda.hhs.gov Bhusari, Sachin (919) 541-3069 bhusaris@niehs.nih.gov

Koturbash, Igor (870) 543-7724 igor.koturbash@fda.hhs.gov Birnbaum, Linda (919) 541-3201 birnbaumls@niehs.nih.gov

Law, Charles (870) 543-7013 charles.law@fda.hhs.gov Bishop, Jack (919) 541-1876 bishop@niehs.nih.gov

Leakey, Julian (870) 543-7916 julian.leakey@fda.hhs.gov Blystone, Chad (919) 541-2741 blystonecr@niehs.nih.gov

Lewis, Sherry (870) 543-7627 sherrym.lewis@fda.hhs.gov Borde, Aditi (919) 541-0461 (special volunteer to Dr. Fenton)

Loukotkova, Lucie (870) 543-7406 lucie.loukotkova@fda.hhs.gov Borde, Pretish (919) 541-4141 (special volunteer to Dr. Fenton)

McDaniel, Lea (870) 543-7213 lea.mcdaniel@fda.hhs.gov Bowden, Beth (919) 541-3355 bowden1@niehs.nih.gov

Mendoza, Maria (870) 543-7844 maria.mendoza@fda.hhs.gov Boyd, Windy (919) 541-9810 boydw@niehs.nih.gov

Mittelstaedt, Roberta (870) 543-7190 roberta.mittelstaedt@fda.hhs.gov Boyle, Michael (919) 541-1676 boylemc@niehs.nih.gov

Moon, Steven (870) 543-7124 steven.moon@fda.hhs.gov Boyles, Abee (919) 541-7886 boylesa@niehs.nih.gov

Nichols, Jasyl (870) 543-7210 jasyl.nichols@fda.hhs.gov Brown, Quashana (919) 541-4069 brownqd@niehs.nih.gov

Nysia, George (870) 543-7047 nysia.george@fda.hhs.gov Bucher, John (919) 541-4532 bucher@niehs.nih.gov

Pack, Lindsay (870) 543-7681 lindsay.pack@fda.hhs.gov Casey, Warren (919) 316-4729 warren.casey@nih.gov

82 National Toxicology Program
Caspary, William (919) 541-2150 caspary@niehs.nih.gov Harvey, Janice (919) 541-1745 janice.harvey@nih.gov

Castro, Lysandra (919) 541-3373 castro@niehs.nih.gov Herbert, Ronald (919) 541-4613 herbert1@niehs.nih.gov

Cesta, Mark (919) 541-3229 cesta@niehs.nih.gov Hoenerhoff, Mark (919) 541-3440 hoenerhm@niehs.nih.gov

Chan, Po-Chuen (919) 541-7561 chanp@niehs.nih.gov Hong, Hue-Hua (919) 541-2141 hong5@niehs.nih.gov

Chan, Melissa (919) 541-3857 chanp2@niehs.nih.gov Hooth, Michelle (919) 316-4643 hooth@niehs.nih.gov

Chang, Xiaoging (919) 541-2373 changx@niehs.nih.gov Howdeshell, Kembra (919) 316-4708 howdeshellkl@niehs.nih.gov

Chen, Vivian (919) 541-3474 (special volunteer w/S Elmore) Hsieh, Jui-Hua (919) 316-4879 hsiehj2@niehs.nih.gov

Chhabra, Rajendra (919) 541-3386 chhabrar@niehs.nih.gov Irwin, Richard (919) 541-3340 irwin@niehs.nih.gov

Clayton, Natasha (919) 541-7843 clayton2@niehs.nih.gov Jahnke, Gloria (919) 541-3376 jahnke@niehs.nih.gov

Collins, Bradley (919) 541-4666 collin10@niehs.nih.gov Jameson, CW (Bill) (919) 541-4096 (special volunteer - Dr. Bucher)

Cora, Michelle (1919) 316-4717 coramc@niehs.nih.gov Jayaram, Beby (919) 541-1480 jayaram1@niehs.nih.gov

Corniffe, Glenda (919) 541-4880 corniffe@niehs.nih.gov Jensen, Heather (919) 541-4075 jensen1@niehs.nih.gov

Cunningham, Michael (919) 541-3799 cunning1@niehs.nih.gov Jeter, Shawn (919) 541-3476 jeter@niehs.nih.gov

Cunny, Helen (919) 541-5717 cunnyh@niehs.nih.gov Johnson, Frank (919) 541-3503 johnso54@niehs.nih.gov

de Serres, Frederick (919) 541-0718 deserres@niehs.nih.gov Jones, Tina (919) 541-2836 jones30@niehs.nih.gov

DeVito, Michael (919) 541-4142 devitom@niehs.nih.gov Kim, Sang-Hyun (919) 541-9468 sang-hyum.kim@nih.gov

Di, Xudong (919) 316-4611 dix3@niehs.nih.gokamel King, Debra (919) 541-0698 kingdh@niehs.nih.gov

Dixon, Darlene (919) 541-3814 dixon@niehs.nih.gov King-Herbert, Angela (919) 541-3464 kingher1@niehs.nih.gov

Dunlap, Paul (919) 541-2679 dunlapp@niehs.nih.gov Kissling, Grace (919) 541-1756 kissling@niehs.nih.gov

Dunnick, June (919) 541-4811 dunnickj@niehs.nih.gov Koivisto, Christopher (919) 541-3241 koivist@niehs.nih.gov

Elmore, Susan (919) 541-3474 elmore@niehs.nih.gov Lahousse, Stephanie (919) 541-3069 lahousses@niehs.nih.gov

Faustman, Elaine (919) 541-7539 faustman@niehs.nih.gov Lasko, Denise (919) 541-0255 lasko@niehs.nih.gov

Fenton, Suzanne (919) 541-4141 fentonse@niehs.nih.gov Liu, Ailin (919) 541-2679 Liua2@niehs.nih.gov

Fields, Sally (919) 541-3971 fields3@niehs.nih.gov Litton, Dan (919) 541-2100 litton@niehs.nih.gov

Filgo, Adam (919) 541-4672 adam.filgo@nih.gov Lunn, Ruth (919) 316-4637 lunn@niehs.nih.gov

Flagler, Norris (919) 541-2397 flagler@niehs.nih.gov Macon, Madisa (919) 541-4703 madisa.macon@nih.gov

Flake, Gordon (919) 541-0037 flake@niehs.nih.gov Malarkey, David (919) 541-1745 malarkey@niehs.nih.gov

Foley, Julie (919) 541-3772 foley1@niehs.nih.gov Malling, Heinrich (919) 541-3378 malling@niehs.nih.gov

Fostel, Jennifer (919) 541-5055 fostel@niehs.nih.gov Masinde, Tiwanda (919) 541-4069 marsh2@niehs.nih.gov

Foster, Paul (919) 541-2513 foster2@niehs.nih.gov Masten, Scott (919) 541-5710 masten@niehs.nih.gov

Fraller, Deirdre (919) 541-0927 frallerdb@niehs.nih.gov Maull, Elizabeth (919) 316-4668 maull@niehs.nih.gov

Frawley, Rachel (919) 541-2151 frawleyr@niehs.nih.gov McCarley, Deborah (919) 541-2384 mccarley@niehs.nih.gov

Freedman, Jonathan (919) 541-7899 freedma1@niehs.nih.gov McIntyre, Barry (919) 316-4747 mcintyrebs@niehs.nih.gov

French, Jef (919) 541-2569 french@niehs.nih.gov McPherson, Chris (919) 541-0167 Mcphers1@niehs.nih.gov

Fuhrman, Laura (919) 541-3415 Laura.Fuhrman@nih.gov Melnick, Ronald (919) 541-4142 melnickr@niehs.nih.gov

Gao, Xiaohua (919) 316-4611 gaox3@niehs.nih.gov Mercado-Feliciano, Minerva (919) 541-2756 mercadofelicim@niehs.nih.gov

Germolec, Dori (919) 541-3230 germolec@niehs.nih.gov Merrick, Alex (919) 541-1531 merrick1@niehs.nih.gov

Goldsmith, Rachel (919) 541-2522 Rachel.goldsmith@nih.gov Moore, Alicia (919) 541-7914 moore5@niehs.nih.gov

Guy, Robbin (919) 541-4363 guyr2@niehs.nih.gov Morgan, Daniel (919) 541-2264 morgan3@niehs.nih.gov

Gwinn, William (919) 541-0598 gwinnwm@niehs.nih.gov Newbold, Retha (919) 541-0738 newbold1@niehs.nih.gov

Hall, Carolyn (919) 541-4995 hall7@niehs.nih.gov Ney, Elizabeth (919) 541-5182 ney@niehs.nih.gov

Hall, Julie (919) 541-2679 Hall2@niehs.nih.gov Ngalame, Ntube (919) 541-5436 ngalamenn@niehs.nih.gov

Hall, Laura (919) 541-7931 hall@niehs.nih.gov Nichols, Sharon (919) 541-7992 nicholss2@niehs.nih.gov

Harry, Jean (919) 541-0927 harry@niehs.nih.gov Orihuela Garcia, Ruben (919) 541-0540 ruben.orihuela@nih.gov

NTP Annual Report 2011 83
Ovwigho, Pamela (919) 541-3278 ovwighope@niehs.nih.gov White, Lori (919) 541-9834 whiteld@niehs.nih.gov

Parham, Frederick (919) 541-0760 parham@niehs.nih.gov Wilson, Ralph (919) 541-4140 wilson23@niehs.nih.gov

Person, Rachel 919) 541-0685 rachel.person@nih.gov Withers, Sheila (919) 541-5066 withersg@niehs.nih.gov

Qu, Wei (919) 541-0540 qu@niehs.nih.gov Witt, Kristine (919) 541-2761 witt@niehs.nih.gov

Quist, Erin (919) 541-3036 erin.quist@nih.gov Wolfe, Mary (919) 541-7539 wolfe@niehs.nih.gov

Rebolloso, Yvette (919) 541-4209 rebollo1@niehs.nih.gov Wyde, Michael (919) 316-4640 wyde@niehs.nih.gov

Rice, Julie (919) 541-2533 ricej2@niehs.nih.gov Xu, Yuanyuan (919) 541-3024 Xuy5@mail.nih.gov

Rider, Cynthia (919) 541-7638 ridercv@niehs.nih.gov Yu, Xianhong (919) 316-4599 yu1@niehs.nih.gov

Roach, Donna (919) 541-3023 roachd@od.nih.gov

Robinson, Deidre (919) 541-4141 deidre.robinson@nih.gov NIOSH/NTP 

Robinson, Veronica (919) 541-2238 godfrey@niehs.nih.gov Anderson, Stacy (304) 285-6174-x2 dbx7@cdc.gov

Rooney, Andrew (919) 316-4704 rooneyaa@niehs.nih.gov Beezhold, Don (304) 285-5963 zec1@cdc.gov

Rowley, Michael (919) 541-3436 rowley@niehs.nih.gov Brown, Kenneth (513) 841-4364 krb1@cdc.gov

Roycroft, Joseph (919) 541-3627 roycroft@niehs.nih.gov Chipinda, Itai (304) 285-5721 clz0@cdc.gov

Sanders, Michael (919) 541-1872 sander10@niehs.nih.gov Curwin, Brian (513) 841-4432 bic4@cdc.gov

Shelby, Michael (919) 541-3455 shelby@niehs.nih.gov DeBord, D. Gayle (513) 533-8212 ded4@cdc.gov

Shines, Cassandra (919) 541-0959 shines1@niehs.nih.gov Erdely, Aaron (304) 285-5903 efi4@cdc.gov

Shockley, Keith (919) 541-3033 shockleykr@niehs.nih.gov Estill, Cherie (513) 841-4476 clf4@cdc.gov

Sills, Robert (919) 541-0180 sills@niehs.nih.gov Franko, Jennifer (304) 285-6174-x8 hfy0@cdc.gov

Smith, Cynthia (919) 541-3473 smith19@niehs.nih.gov Geraci, Chuck (513) 533-8339 ciu9@cdc.gov

Soward, Sharon (919) 541-5132 soward@niehs.nih.gov Green, Brett (304) 285-5884 dox6@cdc.gov

Spencer, Diane (919) 541-2759 spencer2@niehs.nih.gov Hanley, Kevin (513) 841-4113 kwh0@cdc.gov

Stanko, Jason (919) 541-0461 stankojp@niehs.nih.gov Harper, Martin (304) 285-6034 zzg7@cdc.gov

Stockton, Patricia (919) 541-4471 stockton@niehs.nih.gov Hodson, Laura (513) 533-8526 gey7@cdc.gov

Stokes, William (919) 541-7997 stokes@niehs.nih.gov Johnson, Victor (304) 285-6249 vcj6@cdc.gov

Stout, Matthew (919) 541-3489 stoutm@niehs.nih.gov Luster, Michael (304) 285-5940 myl6@cdc.gov

Sun, Yang (919) 541-9812 suny2@mail.nih.gov Lynch, Dennis (513) 533-8213 dwl1@cdc.gov

Surh, In Ok (919) 541-3862 surhi@niehs.nih.gov Methner, Mark (513) 841-4325 mmm5@cdc.gov

Sutton, Deloris (919) 541-5112 sutton2@niehs.nih.gov Munson, Albert E. (304) 285-6121 akm5@cdc.gov

Taylor, Kyla (919) 316-4707 taylorkw@niehs.nih.gov Qian, Yong (304) 285-6286 yaq2@cdc.gov

Teng, Christina (919) 541-0344 teng1@niehs.nih.gov Reynolds, Steve (304) 285-5806 sir6@cdc.gov

Thakur, Sheetal (919) 541-9468 thakursa@niehs.nih.gov Schubauer-Berigan, Mary (513) 841-4251 zcg3@cdc.gov

Thayer, Kristina (919) 541-5021 thayer@niehs.nih.gov Siegel, Paul (304) 285-5855 pds3@cdc.gov

Tice, Raymond (919) 541-4482 tice@niehs.nih.gov Snawder, John (513) 533-8496 jts5@cdc.gov

Tokar, Erik (919) 541-3951 tokare@niehs.nih.gov Stefaniak, Aleksandr (304) 285-6302 boq9@cdc.gov

Ton, Thai-Vu (919) 541-2118 ton@niehs.nih.gov Streicher, Robert (513) 841-4296 rps3@cdc.gov

Travlos, Gregory (919) 541-0653 travlos@niehs.nih.gov Striley, Cynthia (513) 533-8123 chs3@cdc.gov

Tubbs, Jacquelyn (919) 541-2919 tubbsj@niehs.nih.gov Virji, Mohammed (304) 285-5797 fnb8@cdc.gov

Vallant, Molly (919) 541-5234 vallant@niehs.nih.gov Wells, Ray (304) 285-6341 ozw0@cdc.gov

Waalkes, Michael (919) 541-2328 waalkes@niehs.nih.gov Whelan, Elizabeth (513) 841-4437 eaw0@cdc.gov

Waidyanatha, Suramya (919) 541-2144 waidyanathas@niehs.nih.gov Wurzelbacher, Steve (513) 841-4322 srw3@cdc.gov

Walker, Nigel (919) 541-4893 walker3@niehs.nih.gov Yucesoy, Berran (304) 285-5993 yab7@cdc.gov

Walker, Vickie (919) 541-4514 ray3@niehs.nih.gov

84 National Toxicology Program
NTP Annual Report 2011 85

NTP FY 2011 Bibliography

1 Funded by the NIEHS/NIOSH Interagency Agreement 2 Funded by NIOSH voluntary allocations to the NTP 3 Funded by the NIEHS/NCTR Interagency Agreement 4 Funded by NCTR voluntary allocations to the NTP 5 Funded by the NTP/NHGRI/EPA/FDA Interagency Memorandum of Understanding 6

 Funded by the NIEHS/EPA Interagency Agreement 7 Funded by NIEHS voluntary allocations to the NTP

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7 Akgul Y, Derk RC, Meighan T, Rao KMK, Murono EP. 2011. The methoxychlor metabolite, HPTE, inhibits rat luteal

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3 Ali AA, Lewis SM, Badgley HL, Allaben WT, Leakey JE. 2011. Oral glucosamine increases expression of

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4 Ali R, Mittelstaedt RA, Shaddock JG, Ding W, Bhalli JA, Khan QM, Heflich RH. 2011. Comparative analysis of

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3 Antunes AMM, Godinho ALA, Martins IL, Oliveira MC, Gomes RA, Coelho AV, Beland FA, Marques MM. 2010. 

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2 Archer Hartmann SA, Sargent LM, Lowry DT, Holland LA. 2011. Microscale exoglycosidase processing and lectin

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7 Asemu G, Fishbein K, Lao QZ, Ravindran A, Herbert R, Canuto HC, Spencer RG, Soldatov NM. 2011. Cardiac 

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Channels 5(2):138-147.
86 National Toxicology Program

7 Auerbach SS, Thomas R, Shah R, Xu H, Vallant MK, Nyska A, Dunnick JK. 2010. Comparative phenotypic 

assessment of cardiac pathology, physiology, and gene expression in C3H/HeJ, C57BL/6J, and B6C3F1/J mice. 

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1 Beezhold DH. 2011. Monoclonal antibodies 3C3, 6D4, 7D11, 9G6, 24D11, 27C10, 27E2, and 29E5. 

Hybridoma 30(1):103-103.

2 Beezhold K, Liu J, Kan H, Meighan T, Castranova V, Shi X, Chen F. 2011. miR-190-mediated downregulation

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7 Bhave VS, Donthamsetty S, Latendresse JR, Cunningham ML, Mehendale HM. 2011. Secretory phospholipase 

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2 B’Hymer C. 2011. Validation of an HPLC-MS-MS method for the determination of urinary S-benzylmercapturic

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4 Binienda ZK, Beaudoin MA, Thorn BT, Ali SF. 2011. Analysis of electrical brain waves in neurotoxicology:

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2 Blair A, Thomas K, Coble J, Sandler DP, Hines CJ, Lynch CF, Knott C, Purdue MP, Zahm SH, Alavanja MCR,

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2 Bledsoe ML, Pinkerton LE, Silver S, Deddens JA, Biagini RE. 2011. Thyroxine and free thyroxine levels in workers 

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7 Blystone CR, Elmore SA, Witt KL, Malarkey DE, Foster PM. 2011. Toxicity and carcinogenicity of androstenedione 

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4 Boctor SY, Ferguson SA. 2010. Altered adult locomotor activity in rats from phencyclidine treatment on postnatal 

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7 Bucher JR, Thayer K, Birnbaum LS. 2011. The Office of Health Assessment and Translation: A problem-solving 

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1 Funded by the NIEHS/NIOSH Interagency Agreement 2 Funded by NIOSH voluntary allocations to the NTP 3 Funded by the NIEHS/NCTR Interagency Agreement 4 Funded by NCTR voluntary allocations to the NTP 5 Funded by the NTP/NHGRI/EPA/FDA Interagency Memorandum of Understanding 6

 Funded by the NIEHS/EPA Interagency Agreement 7 Funded by NIEHS voluntary allocations to the NTP
NTP Annual Report 2011 87

2 Buck Louis GM, Schisterman EF, Sweeney AM, Wilcosky TC, Gore Langton RE, Lynch CD, Barr DB, Schrader SM, 

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1 Buskirk AD, Hettick JM, Chipinda I, Law BF, Siegel PD, Slaven JE, Green BJ, Beezhold DH. 2011. Fungal pigments

inhibit the matrix-assisted laser desorption/ionization time-of-flight mass spectrometry analysis of darkly 

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2 Calvert GM, Ruder AM, Petersen MR. 2011. Mortality and end-stage renal disease incidence among dry cleaning 

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7 Cannon CZ, Kissling GE, Hoenerhoff MJ, King-Herbert AP, Blankenship-Paris T. 2010. Evaluation of dosages 

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7 Cannon CZ, Kissling GE, Goulding DR, King-Herbert AP and Blankenship-Paris T. 2011. Analgesic effects of

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7 Carney EW, Ellis AL, Tyl RW, Foster PM, Scialli AR, Thompson K, Kim J. 2011. Critical evaluation of current 

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2 Carreon T, Hein MJ, Viet SM, Hanley KW, Ruder AM, Ward EM. 2010. Increased bladder cancer risk among workers 

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7 Casey W, Schmitt M, McFarland R, Isbrucker R, Levis R, Arciniega JL, Descamps J, Finn T, Hendriksen C, Horiuchi

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2 Castranova V. 2011. Overview of current toxicological knowledge of engineered nanoparticles. J Occ Environ Med

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7 Chan PC, Ramot Y, Malarkey DE, Blackshear P, Kissling GE, Travlos G, Nyska A. 2010. Fourteen-week toxicity study 

of green tea extract in rats and mice. Toxicol Path 38(7):1070-1084.

7 Chan PC, Peckham JC, Malarkey DE, Kissling GE, Travlos GS, Fu PP. 2011. Two-year toxicity and carcinogenicity

studies of panax ginseng in Fischer 344 rats and B6C3F1 mice. Amer J Chinese Med 39(4):779-788.

7 Chan PC. 2011. NTP technical report on the toxicity studies of 2,4-decadienal (CAS No. 25152-84-5) administered

by gavage to F344/N rats and B6C3F1 mice. Toxicity Report Series 76:1-94.

4 Chang CW, Beland FA, Hines WM, Fuscoe JC, Han T, Chen JJ. 2011. Identification and categorization of liver 

toxicity markers induced by a related pair of drugs. Int J Mol Sci 12(7):4609-4624.

1 Funded by the NIEHS/NIOSH Interagency Agreement 2 Funded by NIOSH voluntary allocations to the NTP 3 Funded by the NIEHS/NCTR Interagency Agreement 4 Funded by NCTR voluntary allocations to the NTP 5 Funded by the NTP/NHGRI/EPA/FDA Interagency Memorandum of Understanding 6

 Funded by the NIEHS/EPA Interagency Agreement 7 Funded by NIEHS voluntary allocations to the NTP
88 National Toxicology Program

4 Chang HC, Doerge DR, Hsieh CH, Lin YJ, Tsai FJ. 2011. The covalent binding of genistein to the non-prostheticheme-moiety of bovine lactoperoxidase leads to enzymatic inactivation. Biomed Environ Sci 24(3):284-290.

4 ChelonisJ, Kubacak B, Osborn S, Edwards M, Baldwin R, Paule M. 2011. The effects of stimulant medication on

performance of an incremental repeated acquisition task in children with ADHD. Neurotoxicol Teratol 33(4):507-507.

4 ChelonisJJ, Gravelin CR, Paule MG. 2011. Assessing motivation in children using a progressive ratio task. 

Behav Proc 87(2):203-209.

4 ChelonisJJ, Johnson TA, Ferguson SA, Berry KJ, Kubacak B, Edwards MC, Paule MG. 2011. Effect of

methylphenidate on motivation in children with Attention-Deficit/Hyperactivity Disorder. 

Exp Clin Psychopharmacol 19(2):145-153.

2 Chen BT, Afshari A, Stone S, Jackson M, Schwegler-Berry D, Frazer DG, Castranova V, Thomas TA. 2010. 

Nanoparticles – containing spray can aerosol: characterization, exposure assessment, and generator design. 

Inhal Toxicol 22(13):1072-1082.

7 Chen LJ, Wegerski CJ, Kramer DJ, Thomas LA, McDonald JD, Dix KJ, Sanders JM. 2011. Disposition and 

metabolism of cumene in F344 rats and B6C3F1 mice. Drug Met Disp 39(3):498-509.

4 Chen T, Heflich RH, Moore MM, Mei N. 2010. Differential mutagenicity of aflatoxin B-1 in the liver of neonatal and 

adult mice. Environ Mol Mut 51(2):156-163.

7 Cheng Y, Wright SH, Kuester RK, Hooth MJ, Sipes IG. 2011. Characterization of the inhibitory effects of

N-butylpyridinium chloride and structurally related ionic liquids on kidney organic cation transporters. 

Drug Metabolism and Disposition 39(9): 1755-61.

7 Chernoff N, Rogers EH, Zehr RD, Gage MI, Malarkey DE, Bradfield CA, Liu Y, Schmid JE, Jaskot RH, Richards JH, 

Wood CR, Rosen MB. 2011. Toxicity and recovery in the pregnant mouse after gestational exposure to the 

cyanobacterial toxin, cylindrospermopsin. J Appl Toxicol 31(3):242-254.

1 Chipinda I, Blachere FM, Anderson SE, Siegel PD. 2011. Discrimination of haptens from prohaptens using the 

metabolically deficient Cpr(low/low) mouse. Toxicol Appl Pharmacol 252(3):268-272.

1 Chipinda I, Hettick JM, Siegel PD. 2011. Haptenation: chemical reactivity and protein binding.J Allergy (Cairo) EPub. 

1 Chipinda I, Ruwona TB, Templeton SP, Siegel PD. 2011. Use of the human monocytic leukemia THP-1 cell line and 

co-incubation with microsomes to identify and differentiate hapten and prohapten sensitizers. Toxicol 280(3):135-

143.

2 Chirila MM, Lee T, Flemmer MM, Slaven JE, Harper M. 2011. Quantitative mid-infrared diffuse reflection of

occupational wood dust exposures. Appl Spectroscopy 65(3):243-249.

7 Chu IM, Michalowski AM, Hoenerhoff M, Szauter KM, Luger D, Sato M, Flanders K, Oshima A, Csiszar K, Green 

JE. 2011. GATA3 inhibits lysyl oxidase-mediated metastases of human basal triple-negative breast cancer cells.

Oncogene doi: 10.1038/onc.2011.382.

1 Funded by the NIEHS/NIOSH Interagency Agreement 2 Funded by NIOSH voluntary allocations to the NTP 3 Funded by the NIEHS/NCTR Interagency Agreement 4 Funded by NCTR voluntary allocations to the NTP 5 Funded by the NTP/NHGRI/EPA/FDA Interagency Memorandum of Understanding 6

 Funded by the NIEHS/EPA Interagency Agreement 7 Funded by NIEHS voluntary allocations to the NTP
NTP Annual Report 2011 89

7 Collins BJ, Stout MD, Levine KE, Kissling GE, Melnick RL, Fennell TR, Walden R, Abdo K, Pritchard JB, Fernando RA, 

Burka LT, Hooth MJ. 2010. Exposure to hexavalent chromium resulted in significantly higher tissue chromium

burden compared with trivalent chromium following similar oral doses to male F344/N rats and female B6C3F1

mice. Toxicol Sci 118(2):368-379.

7 Collins BJ, Stout MD, Levine KE, Kissling GE, Melnick RL, Fennell TR, Pritchard JB, Walden R, Abdo K, Fernando RA, 

Burka LT, Hooth MJ. 2011. Hexavalent chromium in drinking water, reply. Toxicol Sci 119(2):425-425.

2 Connor TH, DeBord G, Pretty JR, Oliver MS, Roth TS, Lees PSJ, Krieg EF, Rogers B, Escalante CP, Toennis CA, Clark 

JC, Johnson BC, McDiarmid MA. 2010. Evaluation of antineoplastic drug exposure of health care workers at three

university-based U.S. cancer centers. J Occ Environ Med 52(10):1019-1027.

7 Corsini E, Avogadro A, Galbiati V, dell’Agli M, Marinovich M, Galli CL, Germolec DR. 2011. In vitro evaluation of

the immunotoxic potential of perfluorinated compounds (PFCs). Toxicol Appl Pharmacol 250(2):108-116.

2 Costa C, Silva S, Neves J, Coelho P, Costa S, Laffon B, Snawder J, Teixeira JP. 2011. Micronucleus frequencies 

in lymphocytes and reticulocytes in a pesticide-exposed population in Portugal. J Toxicol Environ Health A

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7 Cowin PA, Gold E, Aleksova J, O’Bryan MK, Foster PMD, Scott HS, Risbridger GP. 2010. Vinclozolin exposure

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7 Crawford LW, Foley JF, Elmore SA. 2010. Histology atlas of the developing mouse hepatobiliary system with

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4 Cuevas E, Lantz SM, Tobon-Velasco JC, Newport GD, Wu Q, Virmani A, Ali SF, Santamaria A. 2011. On the in vivo

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1 Cummings KJ, Gaughan DM, Kullman GJ, Beezhold DH, Green BJ, Blachere FM, Bledsoe T, Kreiss K, Cox-Ganser J.

2010. Adverse respiratory outcomes associated with occupational exposures at a soy processing plant. 

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7 Cunningham ML, Collins B, Hejtmancik M, Herbert R, Travlos G, Vallant M, Stout M. 2010. Effects of the PPAR

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1 Dahm MM, Yencken MS, Schubauer-Berigan MK. 2011. Exposure control strategies in the carbonaceous

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1 Day G, LeBouf R, Grote A, Pendergrass S, Cummings K, Kreiss K, Kullman G. 2011. Identification and measurement

of diacetyl substitutes in dry bakery mix production. J Occ Environ Hyg 8(2):93-103.

7 Dearfield KL, Thybaud V, Cimino MC, Custer L, Czich A, Harvey JS, Hester S, Kim JH, Kirkland D, Levy DD, Lorge E,

Moore MM, Ouedraogo-Arras G, Schuler M, Suter W, Sweder K, Tarlo K, van Benthem J, van Goethem F, Witt KL. 

2011. Follow-up actions from positive results of in vitro genetic toxicity testing. Environ Mol Mut 52(3):177-204.

1 Funded by the NIEHS/NIOSH Interagency Agreement 2 Funded by NIOSH voluntary allocations to the NTP 3 Funded by the NIEHS/NCTR Interagency Agreement 4 Funded by NCTR voluntary allocations to the NTP 5 Funded by the NTP/NHGRI/EPA/FDA Interagency Memorandum of Understanding 6

 Funded by the NIEHS/EPA Interagency Agreement 7 Funded by NIEHS voluntary allocations to the NTP
90 National Toxicology Program

7 Deng Y, Edin ML, Theken KN, Schuck RN, Flake GP, Kannon MA, DeGraff LM, Lih FB, Foley J, Bradbury JA, Graves

JP, Tomer KB, Falck JR, Zeldin DC, Lee CR. 2011. Endothelial CYP epoxygenase overexpression and soluble 

epoxide hydrolase disruption attenuate acute vascular inflammatory responses in mice. J Fed Amer Soc Exp Biol

25(2):703-713.

2 DiNapoli VA, Benkovic SA, Li X, Kelly KA, Miller DB, Rosen CL, Huber JD, O’Callaghan JP. 2010. Age exaggerates 

proinflammatory cytokine signaling and truncates signal transducers and activators of transcription 3 signaling 

following ischemic stroke in the rat. Neurosci 170(2):633-644.

4 Dobrovolsky VN, Miura D, Heflich RH, Dertinger SD. 2010. The in vivo pig-a gene mutation assay, a potential tool

for regulatory safety assessment. Environ Mol Mut 51(8-9):825-835.

2 Dodrill MW, Fedan. 2010. Lipopolysaccharide hyperpolarizes guinea pig airway epithelium by increasing

the activities of the epithelial Na+ channel and the Na+-K+ pump. Amer J Physiol-Lung Cell Mol Physiol

299(4):L550-L558.

2 Dodrill MW, Beezhold DH, Meighan T, Kashon ML, Fedan JS. 2011. Lipopolysaccharide increases Na+,K+-pump, 

but not ENaC, expression in guinea-pig airway epithelium. Eur J Pharmacol 651(1-3):176-186.

4 Doerge DR, Twaddle NC, Churchwell MI, Newbold RR, Delclos KB. 2010. Lactational transfer of the soy isoflavone,

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3 Doerge DR, Twaddle NC, Woodling KA, Fisher JW. 2010. Pharmacokinetics of bisphenol A in neonatal and adult 

rhesus monkeys. Toxicol Appl Pharmacol 248(1):1-11.

3 Doerge DR, Vanlandingham M, Twaddle NC, Delclos, KB. 2010. Lactational transfer of bisphenol A in

Sprague-Dawley rats. Toxicol Lett 199(3):372-376. 

4 Doerge DR. 2011. Bioavailability of soy isoflavones through placental/lactational transfer and soy food. 

Toxico Appl Pharmacol 254(2):145-147. 

4 Dorne JL, Doerge D, Boobis A. 2011. Mechanistic basis for the harmonisation of ecological and human risk

assessment. Toxicol Lett 205(S9).

7 Dunnick JK, Singh B, Nyska A, Peckham J, Kissling GE, Sanders JM. 2011. Investigating the potential for toxicity

from long-term use of the herbal products, goldenseal and milk thistle. Toxicol Path 39(2):398-409.

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1 Funded by the NIEHS/NIOSH Interagency Agreement 2 Funded by NIOSH voluntary allocations to the NTP 3 Funded by the NIEHS/NCTR Interagency Agreement 4 Funded by NCTR voluntary allocations to the NTP 5 Funded by the NTP/NHGRI/EPA/FDA Interagency Memorandum of Understanding 6

 Funded by the NIEHS/EPA Interagency Agreement 7 Funded by NIEHS voluntary allocations to the NTP
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1 Funded by the NIEHS/NIOSH Interagency Agreement 2 Funded by NIOSH voluntary allocations to the NTP 3 Funded by the NIEHS/NCTR Interagency Agreement 4 Funded by NCTR voluntary allocations to the NTP 5 Funded by the NTP/NHGRI/EPA/FDA Interagency Memorandum of Understanding 6

 Funded by the NIEHS/EPA Interagency Agreement 7 Funded by NIEHS voluntary allocations to the NTP

U.S. Department of Health and Human Services

National Toxicology Program 

PO Box 12233 

111 TW Alexander Drive 

Research Triangle Park, NC 27709 

www.ntp.niehs.nih.gov

NIH Publication No. 12-5971