IMPROVING DNA, RNA AND PROTEIN AVAILABILITY IN FIXED TISSUE

Release Date:  March 23, 2000

PA NUMBER:  PAR-00-079

National Cancer Institute

Letter of Intent Receipt Dates:  June 14, 2000 and February 9, 2001
Application Receipt Dates:       July 19, 2000 and March 16, 2001

THIS PA USES THE "MODULAR GRANT" AND "JUST-IN-TIME" CONCEPTS. IT INCLUDES 
DETAILED MODIFICATIONS TO STANDARD APPLICATION INSTRUCTIONS THAT MUST BE USED 
WHEN PREPARING APPLICATIONS IN RESPONSE TO THIS PA.

PURPOSE 

The application of many molecular technologies currently requires access to 
fresh or frozen tissue specimens because of the limited ability to extract 
sufficient high quality RNA or protein from fixed tissue.  Progress toward 
clinical application of potentially useful molecular technologies is also 
hampered by fixation methods that fail to conserve the structure of nucleic 
acids and proteins in tissues.  This initiative would encourage research to 
develop (1) new fixation methods to better preserve macromolecules and (2) 
methods to reverse the effects of formalin fixation to make nucleic acids and 
proteins more readily accessible in archived specimens. 

A better understanding of the physical and chemical basis of tissue 
preservation is vital for the rational development of new fixation 
methodologies.  Knowledge of the mechanism of action of existing fixatives 
may provide insight that will allow simple modification of current techniques 
or development of new approaches that will meet the needs of both 
pathologists and researchers.  In addition, carefully designed systematic 
studies may produce empirical solutions that will have the same end result.  

HEALTHY PEOPLE 2010

The Public Health Service (PHS) is committed to achieving the health 
promotion and disease prevention objectives of "Healthy People 2010," a 
PHS-led national activity for setting priority areas. This RFA, Improving 
DNA, RNA and Protein Availability in Fixed Tissue, is related to the priority 
area of cancer.  Potential applicants may obtain a copy of "Healthy People 
2010" at http://www.health.gov/healthypeople/.
 
ELIGIBILITY REQUIREMENTS 

Applications may be submitted by foreign and domestic, for-profit and 
non-profit organizations, public and private, such as universities, colleges, 
hospitals, laboratories, units of State and local governments, and eligible 
agencies of the Federal government. Racial/ethnic minority individuals, 
women, and persons with disabilities are encouraged to apply as principal 
investigators. 

MECHANISM OF SUPPORT 

This PA will use the National Institutes of Health (NIH) exploratory/ 
developmental (R21) grant mechanism.  This mechanism is utilized for pilot or 
feasibility studies to support exploratory research that may lead to 
innovative advances in science.  Because the R21 grant mechanism is designed 
to support pilot studies, extensive preliminary data as evidence of 
feasibility are not required.  However, the applicant must develop a sound 
research plan based on a strong scientific rationale. The R21 grant mechanism 
provides limited funds for short term (up to two years) research projects.  
The overall size of the budget should be proportional to the scope of 
research proposed.  Budgets must be requested in $25,000 increments. The 
total direct costs requested may not exceed $100,000.  If a subcontract is
necessary the total direct costs requested, including the subcontract F&A
costs may be up to $125,000.

Responsibility for the planning, direction, and execution of the proposed 
project will be solely that of the applicant. The total project period for an 
application submitted in response to this PA may not exceed TWO years. These 
grants are non-renewable and continuation of projects developed under this PA 
will be through the traditional unsolicited investigator initiated grant 
program. 

Specific application instructions have been modified to reflect "MODULAR 
GRANT" and "JUST-IN-TIME" streamlining efforts being examined by the NIH. 
Complete and detailed instructions and information on Modular Grant 
applications can be found at 
http://grants.nih.gov/grants/funding/modular/modular.htm

RESEARCH OBJECTIVES 

Improved access to adequately preserved tissue is critical to facilitating 
studies to translate promising comprehensive molecular technologies into 
medical practice. Two major goals of this program announcement are: 
 
(1) to develop novel, improved methodologies for tissue preservation, and 

(2) to develop methods to reverse the effects of formalin fixation on RNA and 
proteins to allow the effective use of existing tissue archives in molecular 
studies.  

The lack of information about fundamental molecular interactions during 
fixation and the need to make tissue specimens more available for molecular 
studies argue strongly for an effort to encourage groups of pathologists, 
chemists, materials scientists and/or molecular biologists to work together 
to develop new approaches to fixation.  These new methods should preserve 
macromolecules while at the same time maintaining appropriate morphology 
required for high quality diagnostic evaluation of specimens.  Research 
proposed in response to this PA must be clearly focused on understanding the 
molecular principles of fixation.  To achieve this goal, appropriate 
collaborations and interactions of chemists, biochemists and/or material 
scientists with molecular biologists and pathologists are strongly 
encouraged.  Some examples of the types of studies that could be supported by 
this PA include but are not limited to:

1)  Studies to better understand the basic mechanisms of fixation, with the 
ultimate goal of developing practical approaches to improve the availability 
of high quality DNA, RNA or protein from fixed tissue. 

2) Studies to understand the impact of tissue handling during fixation.  Such 
studies could evaluate the effects of heat, oxygen, diffusion rates and the 
penetration of fixatives into tissues and identify the major factors that 
affect the stability of macromolecules. 

3) Studies to determine the importance of cross-linking in the stable 
morphological preservation of formalin fixed tissue.  

4) Studies of methods to disrupt cross-links to free DNA, RNA and proteins.  
Approaches could include developing chemical reagents to remove cross-links 
in existing archival tissues or strategies to produce cross-links with 
cleavable groups to facilitate reversal.
 
5) Studies of the usefulness of enzyme inhibitors to stabilize nucleic acids 
and proteins in fixed tissues.

6) Studies to evaluate the impact of current embedding procedures on the 
integrity of macromolecules in fixed tissue. 

INCLUSION OF WOMEN AND MINORITIES IN RESEARCH INVOLVING HUMAN SUBJECTS 

It is the policy of the NIH 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 policy results from the NIH Revitalization Act of 1993 
(Section 492B of Public Law 103-43). 

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 20, 
1994 (FR 59 14508-14513) and in the NIH Guide for Grants and Contracts, 
Volume 23, Number 11, March 18, 1994 available on the web at the following 
URL address: http://grants.nih.gov/grants/guide/notice-files/not94-100.html 

Investigators also may obtain copies of the policy from the program staff 
listed under INQUIRIES.  Program staff may also provide additional relevant 
information concerning the policy. 

INCLUSION OF CHILDREN AS PARTICIPANTS IN RESEARCH INVOLVING HUMAN SUBJECTS

It is the policy of NIH that children (i.e., individuals under the age of 21) 
must be included in all human subjects research, conducted or supported by 
the NIH, unless there are clear and compelling scientific and ethical reasons 
not to include them. This policy applies to all initial (Type 1) applications 
submitted for receipt dates after October 1, 1998. 

All investigators proposing research involving human subjects should read the 
"NIH Policy and Guidelines on the Inclusion of Children as Participants in 
Research Involving Human Subjects" that was published in the NIH Guide for 
Grants and Contracts, March 6, 1998, and is available at the following URL 
address:  http://grants.nih.gov/grants/guide/notice-files/not98-024.html 

LETTER OF INTENT

Prospective applicants are asked to submit 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 participating institutions, and the number and title of the PA 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 
a subsequent application, the information that it contains allows NCI staff 
to estimate the potential review workload and plan the review.

The letter of intent is to be sent to Ms. Marianna J. Bledsoe, M.A., National 
Cancer Institute, at the address listed under INQUIRIES, by the letter of 
intent receipt date listed.
 
APPLICATION PROCEDURES 

The modular grant concept establishes specific modules in which direct costs 
may be requested as well as a maximum level for requested budgets. Only 
limited budgetary information is required under this approach. The just-in-
time concept allows applicants to submit certain information only when there 
is a possibility for an award. It is anticipated that these changes will 
reduce the administrative burden for the applicants, reviewers and Institute 
staff. The research grant application form PHS 398 (rev. 4/98) is to be used 
in applying for these grants, with the modifications noted below.  
Applications kits 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: 
grantsinfo@nih.gov.  The title and number of the program announcement must be 
typed on line 2 of the face page of the application form and the YES box must 
be marked.  For those applicants with internet access, the 398 kit may be 
found at: http://grants.nih.gov/grants/forms.htm.

Applicants are strongly encouraged to call the program contacts listed under 
INQUIRIES with any questions regarding their proposed project and the goals 
of this PA.

BUDGET INSTRUCTIONS

Modular Grant applications submitted in response to this PA will request 
direct costs in $25,000 modules, up to a total direct cost request of 
$100,000 per year or up to a total direct cost of $125,000, including
subcontract F&A costs.The total direct costs must be requested in accordance
with the program guidelines and the modifications made to the standard PHS 398 
application instructions described below:

o  FACE PAGE:  Items 7a and 7b should be completed, indicating Direct Costs 
(in $25,000 increments) and Total Costs [Modular Total Direct plus Facilities 
and Administrative (F&A) costs] for the initial budget period.  Items 8a and 
8b should be completed indicating the Direct and Total Costs for the entire 
proposed period of support.

o  DETAILED BUDGET FOR THE INITIAL BUDGET PERIOD - Do not complete Form Page 
4 of the PHS 398.  It is not required and will not be accepted with the 
application.

o  BUDGET FOR THE ENTIRE PROPOSED PERIOD OF SUPPORT - Do not complete the
categorical budget table on Form Page 5 of the PHS 398.  It is not required 
and will not be accepted with the application.

o  NARRATIVE BUDGET JUSTIFICATION -  Prepare a Modular Grant Budget Narrative 
page (see http://grants.nih.gov/grants/funding/modular/modular.htm for sample 
pages). At the top of the page, enter the total direct costs requested for 
each year.  This is not a form page.

o  Under Personnel, list key project personnel, including their names, 
percent of effort, and roles on the project. No individual salary information 
should be provided. However, the applicant should use the NIH appropriation 
language salary cap and the NIH policy for graduate student compensation in 
developing the budget request.

For Consortium/Contractual costs, provide an estimate of total costs (direct 
plus facilities and administrative) for each year, each rounded to the 
nearest $1,000. List the individuals/organizations with whom consortium or 
contractual arrangements have been made, the percent effort of key personnel, 
and the role on the project. Indicate whether the collaborating institution 
is domestic or foreign.  The total cost for a consortium/contractual 
arrangement is included in the overall requested modular direct cost amount. 
Include the Letter of Intent to establish a consortium.

Provide an additional narrative budget justification for any variation in the 
number of modules requested.

o  BIOGRAPHICAL SKETCH - The Biographical Sketch provides information used by 
reviewers in the assessment of each individual's qualifications for a 
specific role in the proposed project, as well as to evaluate the overall 
qualifications of the research team.  A biographical sketch is required for 
all key personnel, following the instructions below.  No more than three 
pages may be used for each person.  A sample biographical sketch may be 
viewed at:
http://grants.nih.gov/grants/funding/modular/modular.htm

- Complete the educational block at the top of the form page;
- List position(s) and any honors;
- Provide information, including overall goals and responsibilities, on 
research projects ongoing or completed during the last three years;
- List selected peer-reviewed publications, with full citations.

o  CHECKLIST -  This page should be completed and submitted with the 
application. If the F&A rate agreement has been established, indicate the 
type of agreement and the date. All appropriate exclusions must be applied in 
the calculation of the F&A costs for the initial budget period and all future 
budget years.

The applicant should provide the name and phone number of the individual to 
contact concerning fiscal and administrative issues if additional information 
is necessary following the initial review.

Applications not conforming to these guidelines will be considered 
unresponsive to this PA and will be returned without further review.

Applications are to be submitted on the grant application form PHS 398 (rev. 
4/98). Application kits 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, 
E-mail: grantsinfo@nih.gov. The title and number of the announcement must be 
typed in Section 2 on the face page of the application and the Yes box must 
be marked. For those applicants with internet access, the 398 kit may be 
found at  http://grants.nih.gov/grants/forms.htm.

All applications and proposals for NIH funding must be self-contained within 
specified page limitations.  Unless otherwise specified in an NIH 
solicitation, internet addresses (URLs) should not be used to provide 
information necessary to the review because reviewers are under no obligation 
to view the Internet sites.  Reviewers are cautioned that their anonymity may 
be compromised when they directly access an Internet site (see 
http://grants.nih.gov/grants/guide/notice-files/NOT-OD-00-004.html).

Submit a signed, typewritten original of the application, including the 
checklist, and three signed, exact, 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)
 
Center for Scientific Review
National Institutes of Health
6701 Rockledge Drive
Room 1040 - MSC 7710
Bethesda, MD 20892-7710
(20817 for express service) 

At the time of submission, two additional copies of the application must be 
sent to: 

Ms. Toby Friedberg 
Referral Officer 
Division of Extramural Activities 
National Cancer Institute 
6116 Executive Blvd., Room 8062, MSC-8239 
Bethesda, MD 20892-8239
Rockville, MD 20852 (express courier)

The two initial receipt dates for this initiative are listed at the beginning 
of this PA.  It is anticipated that additional receipt dates will be 
announced at a later time. The Center for Scientific Review (CSR) will not 
accept any application in response to this PA 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 

Upon receipt, applications will be reviewed by the CSR for completeness and 
by the NCI program staff for adherence to the guidelines of this PA. 
Incomplete applications will be returned to the applicant without further 
consideration. Applications that are complete and adhere to the guidelines of 
this PA will be reviewed for scientific and technical merit by a committee 
convened by the NCI Division of Extramural Activities in accordance with the 
standard NIH peer review procedures. As part of the initial merit review, all 
applications will receive a written critique.  The applications may also 
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, assigned a priority score, and receive a second level review by 
the National Cancer Advisory Board.

Review Criteria 

The goals of NIH-supported research are to advance our understanding of 
biological systems, improve the control of disease, and enhance health. The 
reviewers will comment on the following aspects of the application in their 
written critiques 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 by the reviewers in assigning the 
overall score weighting them as appropriate for each application. Note that 
the application does not need to be strong in all categories to be judged 
likely to have a major scientific impact and thus deserve a high priority 
score. For example, an investigator may propose to carry out important work 
that by its nature is not innovative but is essential to move a field 
forward. 

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 initial review group will also examine: the appropriateness of proposed 
project budget and duration; the adequacy of plans to include both genders 
and minorities and their subgroups, and children as appropriate for the 
scientific goals of the research and plans for the recruitment and retention 
of subjects; the provisions for the protection of human and animal subjects; 
and the safety of the research environment. 

AWARD CRITERIA 

Applications will compete for available funds with all other recommended 
applications. The following will be considered in making funding decisions: 
Quality of the proposed project as determined by peer review, availability of 
funds, and program priority. 

INQUIRIES 

Inquiries are encouraged. The opportunity to clarify any issues or questions 
from potential applicants is welcome. 

Direct inquiries regarding programmatic issues to: 

Marianna Bledsoe, M.A.
Division of Cancer Treatment and Diagnosis
National Cancer Institute
Executive Plaza North, Room 700
6130 Executive Boulevard
Bethesda, MD  20892-7399
Rockville, MD  20852 (for express/courier service)
Telephone:  (301) 496-7147
FAX:  (301) 402-7819
Email: mb80s@nih.gov

Direct inquiries regarding fiscal matters to: 

Ms. Kathleen Shino
Grants Administration Branch
National Cancer Institute
Executive Plaza South, Suite 243
6120 Executive Boulevard
Bethesda, MD  20892-7150
Telephone:  (301) 496-8635
FAX:  (301) 496-8601
Email:  shinok@gab.nci.nih.gov

AUTHORITY AND REGULATIONS 

This program is described in the Catalog of Federal Domestic Assistance No. 
93.394, 93.395. Awards are made under authorization of  Sections 301 and 405 
of the Public Health Service Act as amended (42 USC 241 and 284) and 
administered under NIH grants policies and Federal Regulations 42 CFR 52 and 
45 CFR Parts 74 and 92. This program is not subject to the intergovernmental 
review requirements of Executive Order 12372 or Health Systems Agency
review. 

The 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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