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United States Environmental Protection Agency
Office of Research and Development
National Center for Environmental Research and Quality Assurance
National Institute of Environmental Health Sciences

Interagency Announcement of Opportunity

1998 Chemical Mixtures in Environmental Health

RFA: ES-98-002

Opening Date: November 21, 1997
Letter of Intent Receipt Date: January 10, 1998
Application Receipt Date: February 10, 1998


    The National Institute of Environmental Health Sciences (NIEHS) and the U.S. Environmental Protection Agency (EPA) share the common goal of promoting research that will ultimately reduce the extent of adverse human health effects occurring as a consequence of exposure to hazardous environmental agents. Historically, research on individual environmental agents has been critical in providing the mechanistic information necessary to begin unraveling the relationships between human illness and dysfunction and environmental exposure, the timing of exposure and individual susceptibility. The protection of human health relies on setting standards for single substances that are based on the mechanistic knowledge gained from the integration of available data from all relevant research disciplines such as toxicology, molecular biology, biochemistry and epidemiology. Yet in our society, human exposure to chemicals, be it environmental or occupational, is rarely limited to a single chemical; rather, people are exposed to a myriad of chemicals throughout their lifetime. Unfortunately, not only is there a lack of knowledge concerning the characterization of real-life mixtures based on human exposure or human body burden but there are limited experimental strategies available that focus on understanding the mechanisms of action of chemical mixtures as it relates to human health. As a consequence, there is limited knowledge of the underlying biological and pathobiological processes associated with how living systems handle chemical mixtures and are affected by them. Moreover, we have limited abilities to predict how chemicals in a mixture interact with each other or biological systems. Accordingly, the objective of this Request For Applications (RFA) is to encourage innovative experimental approaches and/or computational, statistical or predictive strategies for chemical mixtures that focus on the mechanistic basis for chemical interactions and related health effects. It is anticipated that research projects generated as a result of this RFA will provide the scientific foundation necessary to develop biologically relevant risk assessment models for human exposure to chemical mixtures.


    The Public Health Service (PHS) is committed to achieving the health promotion and disease prevention objectives of "Healthy People 2000," a PHS-led national activity for setting priority areas. This RFA, "Chemical Mixtures" is related to the priority area of environmental health. Potential applicants may obtain a copy of "Healthy 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).


    Applications may be submitted by domestic and foreign, not-for-profit organizations, public and private, such as universities, colleges, hospitals, laboratories, and units of State or local governments. Minority individuals, persons with disabilities, and women are encouraged to apply as Principal Investigators.


    The funding mechanisms to be used to assist the scientific community in participating in this grant program will be those of: 1) the National Institutes of Health (NIH) Investigator Initiated Award (R01); or 2) the Environmental Protection Agency's Office of Research and Development, administered in accordance with 40 CFR Part 30 and 40. Policies that govern grant award programs of each agency will prevail for respective sources of support. Responsibility for the planning, direction, and execution of the proposed project will be solely that of the applicant. The requested direct costs for the applications submitted in response to this RFA may not exceed $250,000 per year.


    The total estimated funds available for the first year of support for this program (including direct and indirect costs) is expected to be up to $4.5 million in fiscal year 1998. NIEHS is designating $1.5 million in the first year and EPA is designating at least $3.0 million in total funds with grant support not to exceed 5 years. It is anticipated that most EPA awards will be for 3 years. The anticipated number of awards is eight to twelve, although the actual number may vary. The level of support is dependent on the receipt of sufficient number of applications of high scientific merit. Although this program is provided for within the financial plans of the NIEHS and EPA, awards pursuant to this RFA are contingent upon the availability of funds for this purpose.


    The issuance of this RFA is a joint announcement by NIEHS and EPA and reflects our common research interests. The program will be managed by both agencies to ensure complementary balance of the research portfolio.



    The U.S. EPA mission is to protect environmental quality and human health through effective regulations, policy decisions and outreach programs supported by sound science through research and development activities. Complementary to this mission is the goal of the NIEHS to support research that examines human health consequences of exposure to physical and chemical toxicants in the environment as well as the underlying mechanisms involved in environmentally related disease and dysfunction. Understanding what mixtures of chemicals the public is exposed to as well as the mechanisms involved in the interaction of these mixtures and subsequent health effects is a priority research area. Considering the approximate 80,000 chemicals in commerce, the task of testing these chemicals on an individual basis, let alone as a mixture, is not feasible.

    Chemical mixtures vary significantly in their composition. Simple mixtures may contain two or three chemicals whereas complex mixtures, such as those found at some hazardous waste sites, contain hundreds of chemicals, representing a variety of organic and inorganic classes of chemicals, with varying degrees of toxicity and different modes of toxic action. Clearly, toxicological evaluation of these complex mixtures is difficult but important for hazard assessment and assessment of risk to human health. This assessment, however, is made more challenging in that the behavior of chemicals in mixtures cannot necessarily be inferred from the behavior ascribed to a single pure chemical. For example, several outcomes have been observed as a result of chemical interactions in a mixture: (1) chemicals act independently, and thus the chemicals in the mixture are qualitatively and quantitatively similar to their separate effects; (2) they demonstrate additive effects, simple summing of the toxicity of the chemicals in a mixture describes the total toxicity; (3) there are antagonistic effects, resulting in toxicity being reduced to a greater extent than would have been predicted; and (4) chemicals demonstrate synergism, resulting in toxicity that is greater than additive.

    From a public health perspective, our inability to predict whether agents act in an additive, synergistic, or antagonistic fashion at concentrations encountered in the environment creates real problems for human health risk assessment. Many examples exist where the interactions of chemicals with each other or other physical or biological agents affect health to a greater extent than would have been predicted based on toxicity of the individual components. A mixture of ozone and aldehydes, which are components of smog, causes more severe effects than expected from the additive effects. A much greater risk of developing lung cancer is observed in smokers who are also exposed to radon or asbestos. People infected with hepatitis B are at greater risk for developing aflatoxin-induced liver cancer. Experimental studies in hens suggest that some of the reported illnesses described for the veterans of the Persian Gulf War might have resulted from interactions between immunization shots, anti-nerve gas prophylaxis, insecticides and insect repellents. This concept has been recently endorsed by Haley and Kurt in a cross-sectional epidemiological study in Gulf War veterans on their neurotoxicological health effects.

    Some of the mechanisms involved in alterations in toxicity have been studied and have been shown to be a consequence of changes in the toxicokinetics and/or toxicodynamics of one chemical by another. Toxicokinetic changes affect the absorption, distribution, metabolism and/or excretion of a chemical and can have profound effects on dose response relationships. Changes at the toxicodynamic level might involve a competition between chemicals for binding to a target site, such as a receptor or alter the susceptibility of target cells to the effects of another agent. This could also include changes in signal transduction pathways and cell cycle control. Though some of these alterations have been described for simple mixtures of two components, unraveling the effects of complex mixtures with up to hundreds of chemicals has not been attempted to an appreciable extent.

    It has been estimated that to design a full factorial study to examine the interaction(s) of three chemicals at five different dose levels requires 125 treatment groups and 750 animals if six animals are included in each treatment group. The end result of such a large expenditure of time and funds would be information on one temporal sequence of exposures at one time point post-exposure. Clearly, the information gained would be of limited value. Therefore a rational approach to studying mixtures includes prioritizing and identifying chemical mixtures that are based on known human exposures, and applying innovative experimental and computational strategies to dissect the mechanistic basis for interactions of chemicals in a mixture.

Research Goals

    The goals and scope of this RFA are to encourage and support research on chemical mixtures that will take advantage of the latest advances in computational and information technologies and molecular biology techniques to focus on the mechanistic basis for chemical interactions in biological systems and related health effects and to develop better mathematical tools for risk assessment. In addition, since there is a general lack of knowledge concerning the characterization of real-life mixtures based on human exposure and body burden, research will be supported that focuses on exposure assessment, including environmental transport and fate.

    Understanding the mechanistic basis for interactions at a quantitative level is necessary before confidence can be gained to provide realistic risk assessments for chemical mixtures. For example, using physiologically-based pharmacokinetics/pharmacodynamic (PBPK/PD) modeling and innovative experimental approaches, the mechanism for non-additive interactions of multiple chemicals could be discerned and applied to environmentally relevant concentrations of chemicals. Research on the mechanistic basis for cellular and/or molecular perturbations and associated health effects by mixtures of chemicals is encouraged in this RFA. Research on chemical interactions that exacerbate both cancer and non-cancer effects is strongly encouraged. For example, the toxic effects of chemical mixtures on liver have been extensively studied, but it also has been observed that opposing effects for different organs can occur.

    "Mixtures" for the purposes of this RFA is defined as two or more agents in combination. This could include but not be limited to chemical-chemical, chemical-biological or chemical-physical agent interactions. Some examples of mixtures that could be considered for study include, air pollutants, solvents, volatile organics, pesticides, PAHs, metals or some combination of these.

    The following areas of research interest are not intended to be complete, and investigators may study these and other topics that meet the objectives of this RFA:

      Developing innovative experimental strategies to elucidate the mechanism for interactive effects of chemical mixtures using animal models, in vitro cell systems and gene expression systems. Chemical mixtures at environmentally relevant concentrations should be considered for these mechanistic studies. Dose response, temporal effects and multiple target endpoints should be taken into consideration.

      Understanding the molecular and/or cellular perturbations associated with exposure to simple or complex mixtures. This could include cellular responses such as effects on receptor systems, changes in gene expression associated with xenobiotic metabolism, signal transduction, or cell cycle control. Such research could lead to eventual development of mechanistically based biomarkers.

      Investigating the effects of metal mixtures on molecular and cellular functions. Mixtures can include metal/metal, metal/metalloid, metal/organics. In addition speciation of metallic elements could also be considered.

      Developing and assessing novel methods for measuring and characterizing chemical mixtures in human populations and the environment, including the use of feral animals as models and/or sentinel species.

      Developing novel statistical and mathematical strategies to interpret results and provide better models for predictive toxicology. Some promising approaches include QSAR (quantitative structure activity relationships), PBPK/PD modeling and fractional factorial designs.


    Applicants should request funds for one trip/year for an annual meeting of grantees to be held either at NIEHS in Research Triangle Park, North Carolina or at the U.S. EPA in Washington D.C.


    The NIH Revitalization Act of 1993 (Section 492B of Public Law 103-43) requires that women and members of minority groups and their subpopulations must be included in all NIH-supported biomedical and behavioral research projects involving human subjects, unless a clear and compelling rationale and justification is provided that inclusion is inappropriate with respect to the health of the subjects or the purpose of the research. This new policy supersedes and strengthens the previous policies (Concerning the Inclusion of Women in Study Populations, and Concerning the Inclusion of Minorities in Study Populations), which have been in effect since 1990. The new policy contains some provisions that are substantially different from the 1990 policies. Grantees, regardless of funding source, will be expected to adhere to this policy.

    All investigators proposing research involving human subjects should read the "NIH Guidelines for Inclusion of Women and Minorities as Subjects in Clinical Research," which have been published in the Federal Register of March 28, 1994 (FR 59 14508-14513) and reprinted in the NIH Guide for Grants and Contracts, Volume 23, Number 11, March 18, 1994. Investigators also may obtain copies of the policy from the program staff listed under INQUIRIES. Program staff may also provide further discussion concerning the policy.


    EPA regulations as stated in 40 CFR 30.54 require the inclusion of a Quality Assurance Narrative Statement (QANS, OMB #2080-0033, approved 8/14/97) for any project application involving data collection or processing, environmental measurements, and/or modeling. The QANS provides information on how quality processes or products will be assured. While applications to NIEHS/NIH cannot be considered incomplete without this statement, EPA cannot fund any application without a reviewed QANS; therefore it is requested that the QANS be included with all applications. This statement should not exceed two consecutively numbered, 8.5x11-inch pages of single-spaced standard 12-point type with 1-inch margins. This statement is to be considered as additional information and is not counted against the 25 pages permitted for the Research Plan. The Quality Assurance Narrative Statement should, for each item listed below, either present the required information or provide a brief justification as to why the item does not apply to the proposed research. For awards that involve environmentally related measurements or data generation, a quality system that complies with the requirements of ANSI/ASQC E4, "Specifications and Guidelines for Quality Systems for Environmental Data Collection and Environmental Technology Programs," must be in place.

      1. The activities to be performed (reference may be made to the specific page and paragraph number in the application where this information may be found); criteria for determining the acceptability of data quality in terms of precision, accuracy, representativeness,completeness, comparability.

      2. The study design including sample type and location requirements and any statistical analyses that were used to estimate the types and numbers of samples required for physical samples or similar information for studies using survey and interview techniques.

      3. The procedures for the handling and custody of samples, including sample identification, preservation, transportation, and storage.

      4. The methods that will be used to analyze samples or data collected, including a description of the sampling and/or analytical instruments required.

      5. The procedures that will be used in the calibration and performance evaluation of the sampling and analytical methods used during the project.

      6. The procedures for data reduction and reporting, including a description of statistical analyses to be used and of any computer models to be designed or utilized associated with verification and validation techniques.

      7. The intended use of the data as they relate to the study objectives or hypotheses.

      8. The quantitative and or qualitative procedures that will be used to evaluate the success of the project.

      9. Any plans for peer or other reviews of the study design or analytical methods prior to data collection.

    ANSI/ASQC E4, "Specifications and Guidelines for Quality Systems for Environmental Data Collection and Environmental Technology Programs" is available for purchase from the American Society for Quality Control, phone 1-800-248-1946, item T55. Only in exceptional circumstances should it be necessary to consult this document.


    Prospective applicants are asked to submit, by January 10, 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 and consultants, the participating institutions, and the number and title of the RFA in response to which the application may be submitted. It is important to consider during the planning phase of the application that the requested direct costs for the applications submitted in response to this RFA may not exceed $250,000 per year.

    Although a letter of intent is not required, is not binding, and does not enter into the review of the subsequent application, the information that it contains is helpful in planning for the review of applications. It allows review staff to estimate the potential review work load and to avoid conflicts 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
      111 T. W. Alexander Drive, EC-24
      Research Triangle Park, NC 27709
      Telephone: (919) 541-7826
      FAX: (191) 541-2503


    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 Office of Grants Information, Center for Scientific Review (CSR, formally known as Division of Research Grants, DRG), National Institute of Health, 6701 Rockledge Drive, Room 3034, MSC 7762, Bethesda, Maryland 20892,
    telephone 301/435-0714, Email:

    The RFA label available in the PHS 398 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:

      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 also 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
      111 T. W. Alexander Drive, EC-24
      Research Triangle Park, NC 27709

    Applications must be received by February 10, 1998. If an application is received after that date, the Center for Scientific Review (CRS)will contact the applicant to determine whether it will be returned to the applicant or be reviewed with unsolicited applications for the next regular receipt date. 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. In addition, CSR will not accept any application that is essentially the same as one already reviewed. This does not include the submission of substantial revisions of applications already reviewed, but such applications must include an introduction addressing the previous critique.


    Because the scope of the research proposed in response to this RFA encompasses the interest of several NIH institutes and centers, applications might receive dual assignments based on the established PHS referral guidelines. Applications will be administratively reviewed by NIH staff for completeness and responsiveness to this announcement. Applications found to be incomplete or non-responsive to this RFA will be inactivated and returned to the applicant without further consideration. Those applications which are complete and responsive to this RFA will be evaluated for scientific and technical merit by an appropriate peer review group convened by the NIEHS and EPA in accordance with NIH peer review procedures. As part of the initial merit review, all applications will receive a written critique. Those applications judged by the study section panel to be non-competitive will be administratively withdrawn from competition. Those applications judged to be competitive, deemed to have the highest scientific merit, generally the top half of the applications under review, will be discussed at the study section meeting, assigned a priority score, and receive a second level review by the EHS advisory council and EPA.


    The following criteria will be considered:

    (1) Significance: Does this study address an important problem? If the aims of the application are achieved, how will scientific knowledge be advanced? What will be the effect of these studies on the concepts or methods that drive this field?

    (2) Approach: Are the conceptual framework, design, methods, and analyses adequately developed, well-integrated, and appropriate to the aims of the project? Does the applicant acknowledge potential problem areas and consider alternative tactics?

    (3) Innovation: Does the project employ novel concepts, approaches or method? Are the aims original and innovative? Does the project challenge existing paradigms or develop new methodologies or technologies?

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

    (5) Environment: Does the scientific environment in which the work will be done contribute to the probability of success? Do the proposed experiments take advantage of unique features of the scientific environment or employ useful collaborative arrangements? Is there evidence of institutional support?


    The anticipated date of award is July 1, 1998. The following will be considered in making funding decisions:

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


    Written and telephone inquiries concerning this RFA are encouraged. The opportunity to clarify any issues or questions from potential applicants is welcomed.

    Direct inquiries regarding programmatic issues to:

      Claudia Thompson, Ph.D.
      Program Administrator
      Chemical Exposures and Molecular Biology Branch
      Division of Extramural Research and Training
      National Institute of Environmental Health Sciences
      P.O. Box 12233, 104 Alexander Drive, MD-3-04
      Research Triangle Park, NC 27709
      Telephone: (919) 541-4638
      Fax: (919) 541-4937


      Chris Saint, Ph.D.
      National Center for Environmental Research
      and Quality Assurance
      U.S. Environmental Protection Agency
      401 M Street SW
      Washington, D.C. 20460
      Telephone: (202) 564-6909
      FAX: (202) 565-2448


      Thomas Veirs
      National Center for Environmental Research
      and Quality Assurance
      U.S. Environmental Protection Agency
      401 M Street SW
      Washington, D.C. 20460
      Telephone: (202) 564-6831
      FAX: (202) 565-2447

      Direct inquiries regarding fiscal matters to:

      Mr. David L. Mineo
      Grants Management Officer
      Division of Extramural Research and Training
      P.O. Box 12233
      Research Triangle Park, NC 27709
      Telephone: (919) 541-7628
      Fax: (919) 541-2860

      Mr. Jack Puzak
      Deputy Director
      National Center for Environmental Research and Quality Assurance
      U.S. Environmental Protection Agency
      401 M Street, SW (8701R)
      Washington, D.C. 20460
      Telephone: (202) 564-6825
      Fax: (202) 565-2444


    This program is described in the Catalog of Federal Domestic Assistance No. 93.113, 93.114 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. EPA awards are made under authority of 40 CFR Part 30 and 40. This program is not subject to the intergovernmental review requirements of executive order 12372 or Health Systems Agency Review.

    The Public Health Service (PHS) and EPA 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.