PEROXISOME PROLIFERATORS AND MECHANISMS OF CARCINOGENESIS

Release Date:  February 23, 1998

RFA:  ES-98-003

P.T

National Institute of Environmental Health Sciences

Letter of Intent Receipt Date:  April 1, 1998
Application Receipt Date:  May 20, 1998

PURPOSE

Environmental health research carried out by the National Institute of
Environmental Health Sciences (NIEHS) provides a solid scientific foundation for
understanding interrelationships between the environment, genetics and temporal
factors as they relate to human disease and dysfunction.  The NIEHS Division of
Extramural Research and Training is responsible for developing and directing the
investigator-initiated hypothesis-driven mechanistically-based research related
to the NIEHS mission. Research conducted by the National Toxicology Program
(NTP), centered at the NIEHS, focuses on the evaluation of
environmental/industrial agents for their toxic effects using a broad array of
assays and test systems and generates data to strengthen the scientific
foundations for risk assessment.

The goal of this Request for Applications (RFA) is to provide scientifically-
based data to aid the NIEHS in understanding  the mechanisms of action of agents
under study by the NTP to further improve the risk assessment process, and,
thereby, better protect the public health.

This program, as outlined in this RFA, utilizes the R03 Small Grants Program to
encourage investigator-initiated applications that will utilize frozen tissues
from an NTP study involving exposure to peroxisome proliferators.  It is
anticipated that these investigator-initiated research projects will complement
the NTP study  by providing additional information on the mechanism(s) of action
of the agents tested.

HEALTHY PEOPLE 2000

The Public Health Service (PHS) is committed to achieving the health promotion
and disease prevention objectives of þHealthy People 2000,þ a PHS-led national
activity for setting priority areas.  This RFA, Peroxisome Proliferators and
Mechanisms of Carcinogenesis, is related to the priority area of environmental
health.  Potential applicants may obtain a copy of þHealthy People 2000" (Full
Report: Stock no. 017-001-00474-0 or Summary Report: Stock No. 017-001-00473-1)
through the Superintendent of Documents, Government Printing Office, Washington,
DC 20402-9325 (telephone: (202) 512-1800).

ELIGIBILITY REQUIREMENTS

Applications may be submitted by domestic, for-profit and non-profit
organizations, public and private, such as universities, colleges, hospitals,
laboratories, units of State or local governments and eligible agencies of the
Federal government.  Foreign institutions and organizations are not eligible. 
Applications from minority individuals and women are encouraged.  Submission of
an application precludes concurrent submission of a regular research grant
application (R01 or R29) containing essentially the same research proposal.  In
addition, small grant research support may not be used to supplement research
projects currently supported by Federal or non-Federal funds or to provide
interim support for projects under review by the Public Health Service.

MECHANISM OF SUPPORT

This RFA will use the NIH Small Grants Program (R03) awards.  Responsibility for
the planning, direction, and execution of the proposed project will be solely
that of the applicant.  The requested costs and project period will be a maximum
of $50,000 (direct cost) for a maximum of one year.  Small grants are not
renewable but may be extended for an additional year with no additional funds at
the discretion of the applicant organization.

FUNDS AVAILABLE

The total estimated funds available for this small grants program is $400,000,
which will support approximately four to six awards.  This level of support is
dependent on the receipt of a sufficient number of applications of high
scientific merit.  Although this program is provided for within the financial
plans of the NIEHS/NTP, awards pursuant to this RFA are contingent upon the
availability of funds for this purpose.

RESEARCH OBJECTIVES

Background

Peroxisome proliferators (PPs) represent a diverse group of chemicals with a high
likelihood of clinical, occupational and environmental exposure to humans.  A
human heath concern exists because of the association between peroxisome
proliferation and cancer in laboratory rodents.  Chemicals considered as PPs
include fibrate hypolipidemic drugs, phthalate ester plasticizers, herbicides
such as 2,4-D and endogenous, long chain fatty acids.  Characteristic responses
of rodent hepatocytes to PPs include hepatomegaly, a marked proliferation of
peroxisomes in parenchymal cells, and an increase in peroxisomal þ-oxidation of
fatty acids.  These responses are thought to be mediated by the PP activation of
a steroid hormone-like receptor termed the peroxisome proliferator-activated
receptor (PPAR).  PPARs are ligand-activated transcription factors that control
gene expression by interacting with specific DNA response elements located
upstream of responsive genes.  Despite being regulated by PPAR, the effects of
PPs on the induction of responsive genes varies widely depending on the PP itself
as well as the species of test animal.  Hepatocarcinogenicity has been produced
in rats and mice by the protypic peroxisome proliferator WY-14,643 and
gemfibrozil and has been reported for di-(2-ethylhexyl)phthalate.  Hepatocellular
carcinomas have been produced in rats administered clofibrate.  This class of
chemicals including di(2-ethylhexyl)phthalate and gemfibrozil have been shown not
to be mutagenic.  Consequently, the mechanism of carcinogenicity is unclear,
although biochemical and physiological effects associated with hepatic peroxisome
proliferation have been implicated in the etiology of liver toxicity and
carcinogenicity.  Understanding the mechanism(s) of peroxisome proliferator-
induced carcinogenicity in rodents will allow evaluation and comparison to
possible mechanisms in humans.

Research Goals

To define the mechanism(s) whereby peroxisome proliferators induce cancer in
rodents the NTP designed and carried out 14-28- and 90-day studies with dibutyl
phthalate, WY-14,643, gemfibrozil and 2,4-(Dichlorophenoxy) acetic acid (2,4-D)
in male rats, mice and hamsters.

Numerous endpoints were measured in order to identify gene products activated by
PPs that are mechanistically linked to the carcinogenicity of this class of
compounds in rodents.  It is anticipated that these would serve as biomarkers to
ascertain if similar gene products were produced in humans exposed to PPs thus
indicating exposure presented a risk of cancer in humans.

A detailed toxicity study design including species studied, doses tested,
endpoints measured and tissues available for investigator-initiated studies under
this RFA may be obtained from the  NIEHS home page at the following address: 
http://www.niehs.nih.gov/dert/ppar.htm

To aid the NTP in developing the scientific data set necessary to define the
mechanism of tumor development by these agents, the NTP, in coordination with the
Division of Extramural Research and Training, proposes to add additional
investigator-initiated studies from this RFA to these studies.

Projects that will provide biochemical endpoints that are related to (or predict)
liver carcinogenicity of peroxisome proliferators are requested.  They include
but are not limited to:

o  Cellular effects/biochemical changes in liver related to peroxisome
proliferator-induced carcinogenicity (i.e., cell mitosis/apoptosis).

o  The relationship between DNA repair enzyme activity or oxidative DNA damage
and peroxisome proliferator-induced carcinogenicity.

o  Oxidative stress, reactive oxygen species, antioxidant levels and peroxisome
proliferator-induced carcinogenicity.

o  Characterization of protein changes with chemical, species, time and dose of
peroxisome proliferation and carcinogenicity.

o  Gene expression and peroxisome proliferator-induced carcinogenicity.

o  Induction of cytochrome P450 isozymes or other oxidative stress associated
enzymes.

o  PPAR distribution (5 subtypes) in relation to carcinogenicity.

Research applications should  be hypothesis driven, mechanistically-based and
must be justified as to how this NTP study is unique in providing tissues needed
for the studies proposed, how the studies can be accomplished within the budget
and personnel proposed, and how the data will aid in understanding the mechanism
of the carcinogenicity  of peroxisome proliferators.

Study Design Considerations

o  Applications must be directly related only to the liver carcinogenicity of the
test chemicals, other endpoints such as reproductive or immune are not considered
responsive to the RFA and will not be accepted.

o  Since the NTP study is already completed, investigator-initiated studies must
be limited to utilization of the available specific frozen or formalin fixed
tissues.

o  Tissues  will be shared by these successful applicants of this RFA. 
Applicants should contact the NTP (see inquiries section) for tissue availability
before finalizing their applications.

o  It is expected that all studies will use tissues from a rodent (mouse and/or
rat) plus, hamster tissues in order to compare a responsive and nonresponsive
species. Investigators must, in their applications, be specific and justify the
tissues needed, preparation of tissues needed, doses needed, shipping
requirements, animal species to be studied and dosing times to be studied.

o  Applicants need not propose to examine  all studies or all time points or all
dose levels.  Investigators should carefully examine the NTP study design and
design a proposal to specifically address a particular question using the
appropriate number of tissues, chemicals and necropsy times.

Applicants are requested to set aside funds for a trip to NIEHS to discuss their
results and to be prepared to integrate their findings into an NTP final report. 
Inclusion of data in the NTP final report would not preclude independent
publication.  The investigator-initiated work may be published independently at
the discretion of the investigators.  Investigators may also have access to NTP
data on, for example, blood, tissue, adipose tissue levels of chemicals which may
be needed for analysis and interpretation of their own data derived from this
collaborative study.

LETTER OF INTENT

Prospective applicants are asked to submit, by April 1, 1998, a letter of intent
that includes a descriptive title of the proposed research, the name, address and
telephone number of the Principal Investigator, the identities of other key
personnel,  participating institutions, and the number and title of the RFA in
response to which the application may be submitted.  Although a letter of intent
is not required, is not binding, and does not enter into the review of subsequent
application, the information that it contains is helpful in planning for the
review of applications.  It allows NIEHS staff to estimate the potential review
work load and to avoid conflict of interest in the review.

The letter of intent is to be sent to:

Ethel B. Jackson, D.D.S.
Division of Extramural Research and Training
National Institute of Environmental Health Sciences
P.O. Box 12233, MD EC-24
111 T.W. Alexander Drive
Research Triangle Park, NC 27709
Telephone:  (919) 541-7826
FAX:  (919) 541-2503
Email:  jackson4@niehs.nih.gov

APPLICATION PROCEDURES

The research grant application form PHS 398 (rev. 5/95) is to be used in applying
for these grants.  These forms are available at most institutional offices of
sponsored research and may be obtained from the Division of Extramural Outreach
and Information Resources, National Institutes of Health, 6701 Rockledge Drive,
MSC 7910, Bethesda, MD 20892-7910, telephone 301/435-0714, Email:
ASKNIH@od.nih.gov.

THE FOLLOWING ARE SUPPLEMENTAL INSTRUCTIONS:

o  The application must detail the specific budget categories and percent efforts
that will be required.  This will be a $50,000 maximum award - direct cost.  The
budget must be justified. Equipment will be limited to $5,000.

o  The applicant must be explicit in describing the proposed interface between
the NTP study design and the proposed project.

o  Preliminary data are not required except to indicate the expertise of the
Principal Investigator to carry out the proposed studies.

o  The Research Plan (Specific Aims, Background and Significance, Preliminary
Studies, Research Design and Methods sections) is not to exceed Ten (10) pages. 
Tables and figures are included in the Ten-page limitation.  Applications that
exceed page limitations or PHS 398 requirements for font size (height or
letters), type density (characters per inch), and margins (see PHS 398
directions) will be returned to the investigator.

o  Do not use the appendix to circumvent the page limitation.

The RFA Label available in the PHS 398 (rev. 5/95) application form must be
affixed to the bottom of the face page of the application.  Failure to use this
label could result in delayed processing of the application such that it may not
reach the review committee in time for review.  In addition, the RFA title and
number must be typed on Line 2 of the face page of the application form and the
YES box must be marked.

Submit a signed, typewritten original of the application, including the
checklist, and three signed, clear, and single-sided photocopies in one package
to:

CENTER FOR SCIENTIFIC REVIEW
NATIONAL INSTITUTES OF HEALTH
6701 ROCKLEDGE DRIVE, ROOM 1040 - MSC 7710
BETHESDA, MD 20892-7710
BETHESDA, MD 20817 (for express/courier service)

At the time of submission, two additional copies of the application must be sent
to Dr. Ethel Jackson at the address listed under LETTER OF INTENT.  If these two
additional copies are not forwarded to Dr. Jackson, it will adversely affect the
review of the grant application.

Applications must be received by May 20, 1998.  If an application is received
after that date, it will be returned to the applicant without review.  The Center
for Scientific Review (CSR) will not accept any application in response to this
RFA that is essentially the same as one currently pending initial review, unless
the applicant withdraws the pending application.  The CSR will not accept any
application that is essentially the same as one already reviewed.  This does not
preclude the submission of substantial revisions of applications already
reviewed, but such applications must include an introduction addressing the
previous critique.

REVIEW CONSIDERATIONS

Applications that are complete and responsive to the RFA will be evaluated for
scientific and technical merit by an appropriate peer review group convened by
the NIEHS in accordance with NIH peer review procedures.  As part of the initial
merit review, all applications will receive a written critique and undergo a
process in which only those applications deemed to have the highest scientific
merit, generally the top half of applications under review, will be discussed and
assigned a priority score.

Review Criteria

The goals of NIH-supported research are to advance our understanding of
biological systems, improve the control of disease, and enhance health.  In the
written review, comments on the following aspects of the application will be made
in order to judge the likelihood that the proposed research will have a
substantial impact on the pursuit of these goals.  Each of these criteria will
be addressed and considered in the assignment of the overall score.

(1) Significance: If the study is completed, how will scientific knowledge on the
mechanism of peroxisome proliferator-induced hepatocarcinogenesis be advanced?

(2) Approach: Are the designs, methods and analysis adequately developed and
appropriate to the aims of the project?  Does the applicant acknowledge potential
problems and consider alternative tactics?  Is there an explanation of how the
data obtained will be integrated with the NTP data to provide a more
comprehensive analysis of the mechanism of carcinogenicity of the test agents? 
Is the study focused on the relationship among exposure, endpoint, species and
hepatocarcinogeneses?

(3) Innovation: Is there justifications to how the tissues are unique to the
proposed study and how the study will make the best of the tissues?

(4) Investigator: Is the investigator appropriately trained and well suited to
carry out the proposed project?  Is the work proposed appropriate to the
experience level of the principal investigators and other researchers, (if any)?

(5) Environment: Does the scientific environment in which the work will be done
contribute to the probability of success?  Is there evidence of institutional
support?

NOTE: If the project can be completed without the aid of this NTP study, the
proposal will not qualify under this RFA.

AWARD CRITERIA

The anticipated date of award is September 1998 pending availability of funds. 
The following will be considered in making funding decisions:

o  quality of the proposed project as determined by peer review;
o  availability of funds; and
o  program balance among research areas of the RFA.

INQUIRIES

Written, telephone or e-mail inquiries concerning this RFA are encouraged.  The
opportunity to clarify any issues or questions from potential applicants is
welcome.

Direct inquiries regarding programmatic issues to:

Jerrold J. Heindel, Ph.D.
Division of Extramural Research and Training
National Institute of Environmental Health Sciences
P.O. Box 12233, MD EC-23
111 T.W. Alexander Drive
Research Triangle Park, NC  27709-2233
Telephone:  (919) 541-0781
FAX:  (919) 541-5064
Email:  heindelj@niehs.nih.gov

Direct inquiries regarding fiscal matters to:

Mr. David L. Mineo
Division of Extramural Research and Training
National Institute of Environmental Health Sciences
P.O. Box 12233, MD EC-22
111 T.W. Alexander Drive
Research Triangle Park, NC  27709-2233
Telephone:  (919) 541-7628
FAX:  (919) 541-2860
Email:  mineo@niehs.nih.gov

For clarification of NTP study design or for information on tissue availability
contact:

Michael L. Cunningham, Ph.D. DABT
Environmental Toxicology Program
National Institute of Environmental Health Sciences
P.O. Box 12233 MD B3-10
111 T.W. Alexander Drive
Research Triangle Park, NC  27709-2233
Telephone:  (919) 541-3799
FAX: (919) 541-4632
Email:  cunning1@-niehs.nih.gov

AUTHORITY AND REGULATIONS

This program is described in the Catalog of Federal Domestic Assistance No.
93.113 and 93.115.  Awards are made under authorization of the Public Health
Service Act, Title IV, Part A (Public Law 78-410, as amended by Public Law
99-158, 43 USC 241 and 285) and administered under PHS grants policies and
Federal Regulations 42 CFR 52 and 45 CFR Part 74.  This program is not subject
to the intergovernmental review requirements of Executive Order 12372 or Health
Systems Agency review.

The Public Health Service (PHS) strongly encourages all grant and contract
recipients to provide a smoke-free workplace and promote the non-use of all
tobacco products.  In addition, Public Law 103-227, the Pro Children Act of 1994,
prohibits smoking in certain facilities (or in some cases, any portion of a
facility) in which regular or routine education, library, day care, health care
or early childhood development services are provided to children.  This is
consistent with the PHS mission to protect and advance the physical and mental
health of the American people.

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