Full Text HL-96-017
NIH GUIDE, Volume 25, Number 27, August 9, 1996
RFA:  HL-96-017
P.T. 34

  Cardiovascular Diseases 

National Heart, Lung and Blood Institute
Letter of Intent Receipt Date:  February 1, 1997
Application Receipt Date:  April 24, 1997
This solicitation is intended to foster fundamental research into the
mechanisms responsible for the cardiovascular dysfunction and disease
that has been seen in HIV+ patients.  Investigations may be conducted
on cells, tissues or whole animals, including those that have been
genetically altered.  The purpose is to develop understanding of the
role of virus, viral proteins, immune cells, cytokine production,
growth factor expression and co-infection with other pathogens in the
altered function and disease manifestations of the cardiovascular
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
Request for Applications (RFA), Etiology of Cardiovascular
Complications in HIV Infection, is related to the priority area of
HIV Infection. 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, 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.
All current policies and requirements that govern the research grant
programs of the NIH will apply to grants awarded under the RFA.
Awards under this RFA to foreign institutions will be made only for
research of very unusual merit, need, and promise, and in accordance
with Public Health Service policy governing such awards.
Among the disciplines and expertise that may be appropriate for this
research program are cell biology, molecular biology, biochemistry,
genetics, immunology, pathology, virology and physiology.
This RFA will use the NIH individual research project grant (R01)
mechanism of support.  However, specific application instructions
have been modified to reflect "MODULAR GRANT" and "JUST-IN-TIME"
streamlining efforts being examined by the NIH.  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, applicant institutions, reviewers and Institute staff.
For this RFA, funds must be requested in $25,000 direct cost modules
and a maximum of ten modules ($250,000 direct costs) per year may be
requested.  A feature of the modular grant concept is that no
escalation is provided for future years, and all anticipated expenses
for all years of the project must be included within the number of
modules being requested.  Only limited budget information will be
required and any budget adjustments made by the Initial Review Group
will be in modules of $25,000.  Instructions for completing the
Biographical Sketch have also been modified.  In addition, Other
Support information and the application Checklist page are not
required aspart of the initial application.  If there is a
possibility for an award, necessary Budget, Other Support and
Checklist information will be requested by NHLBI staff following the
initial review.  The APPLICATION PROCEDURES section of the RFA
provides specific details of modifications to the standard PHS 398
application kit instructions.  Responsibility for the planning,
direction, and execution of the proposed project will be solely that
of the applicant.
It is anticipated that for fiscal year 1997, approximately $1.5
million total costs will be available for the first year of support
for this initiative.  Award of grants pursuant to this RFA is
contingent upon receipt of such funds for this purpose.  It is
anticipated that approximately four to six new grants will be awarded
under this program.  Applicants may request up to five years of
support.  The specific number to be funded will, however, depend on
the merit and scope of the applications received and on the
availability of funds.  Direct costs will be awarded in modules of
$25,000 less any overlap or other necessary administrative
adjustments.  Indirect costs will be awarded based on the negotiated
The incidence of HIV-infected patients suffering cardiac
complications is unknown.  Estimates vary considerably, depending
upon geographical location, patient population and diagnostic
procedures.  Conservatively, it is estimated that about six percent
to seven percent of HIV-infected adult patients have clinically
significant heart disease, and a larger proportion, possibly as high
as 18 percent, have clinically silent abnormalities.  The two most
common forms of HIV-related symptomatic cardiac disease are
cardiomyopathy and pericardial effusion associated with cardiac
tamponade.  Left ventricular dysfunction occurs in all age groups
including children and appears to increase when circulating CD4 cell
counts are severely depressed. Left ventricular diastolic dysfunction
has been reported in asymptomatic and symptomatic patients infected
with HIV. HIV-infected intravenous drug users may develop severe
cardiomyopathy and congestive heart failure with less CD4 cell
depression than HIV patients who do not use drugs.
Autopsy studies reveal a high prevalence of lymphocytic infiltrates
in the hearts of AIDS patients, even though death may have been due
to causes unrelated to cardiac disease, and some of these patients
were not noted to have had symptomatic cardiac disease.  In a
meta-analysis of 402 cases, 46 percent were found to have
histological evidence of myocarditis, yet a pathogenic viral,
bacterial, or fungal agent was confirmed in only one out of five
cases.  There has been one report of significant coronary artery
lesions in eight young HIV-infected patients who were found dead in
unexplained circumstances.  None of these patients had a family
history of hypercholesterolemia, hypertension, or diabetes.
The percentages of cardiac abnormalities are much higher in children
infected with HIV.  In one study of 81 patients, 64 percent were
noted to have resting sinus tachycardia, and 11 percent sinus
bradycardia.  Dysrhythmias occurred in 35 percent, cardiorespiratory
arrest occurred in 9 percent and 10 percent had chronic heart
failure.  In 12 percent, death was associated with marked cardiac
Taking the Public Health Service estimate of 350,000 U.S. AIDS cases
by the year 2000 and assuming a 6 percent rate of development of
symptomatic heart disease in adults, a total of 21,000 HIV+ patients
will require specialized cardiac care.  The number will be very much
larger worldwide.  The susceptibility of cells within the heart to
infection from HIV is undefined, as are possible mechanisms of
indirect effects of HIV on the cardiovascular system.  Research
focused on the etiology and pathophysiology of cardiac disease in
these patients could lead to improved treatment strategies and
prevention of the onset of cardiovascular disease.
Investigations of the etiology of cardiac complications in
HIV-infected patients are confounded by a variety of factors,
including drug use, the large number of self-prescribed medications,
problems of nutrition, intercurrent illnesses, and of course,
medications prescribed by the physician.  For children with
vertically transmitted HIV, the aforementioned factors may have
affected the in utero environment.  In addition, questions remain
regarding:  the effects of in utero infection; the possibility of
increased susceptibility of immature and growing cells of the fetus
and infant to HIV infection; and abnormal expression of growth
factors and cytokines.  For these reasons, it is proposed to study
the effects of HIV on the cardiovascular system in animal models, in
terms of a direct role and of an indirect role via expression of
viral proteins, dysregulation of the immune system, cytokine
expression and the pathogenicity of other infectious agents.
In general, the role of viruses in the etiology of cardiac disease is
poorly understood.  Theories include injury as a result of viral
infection of cardiac cells, apoptosis, myocytolysis as a consequence
of a T-cell response to virus infected myocytes, autoimmune responses
which target normal and infected myocytes, and cytokine and growth
factor mediated dysfunction and injury.  Susceptibility to
inflammation may involve a genetic component.  Similar speculations
have been advanced for the occurrence of cardiac disease in
HIV-infected patients.  Thus, study of the role of HIV in cardiac
complications might well lead to an overall advance in understanding
the immunopathological mechanisms of cardiac injury resulting from
infection by cardiotropic viruses and provide the basis for rational
therapeutic strategies.  One approach to understanding some of the
mechanisms of dysfunction and disease of various organs and tissues
seen in AIDS patients is to examine the role of various gene products
by constructing transgenic mice, possibly with cardiotropic
promoters.  Several attempts have been made in organs and tissues
other than heart, using various strains of mice, including SCID mice.
This latter model may be useful in studying the role of
immunodeficiency in the development of cardiac dysfunction and as an
approach to cardiac disease in pediatric patients with HIV infection.
Transgenic mice containing the complete HIV coding sequences fused to
the mouse mammary tumor virus long terminal repeat have been
generated.  These mice produce gag and env HIV proteins in organs
such as mammary gland, spleen and liver. In some transgenic lines,
low levels of HIV proteins could also be detected in serum.  The
report states that animals sacrificed at 17 months of age were
indistinguishable from non-transgenic mice on macroscopic and
histologic examination.  Thus, it would appear that cells from
various tissues in the mouse are capable of replicating retroviruses
and producing HIV proteins without causing disease. However, the
heart was not among the tissues examined.
Transgenic mice expressing the entire HIV genome under the control of
the promoter for the human neurofilament NF-L gene exhibited
neuropathological changes 7 to 12 months after birth.  HIV was
expressed in the neurons of these mice in contrast to the findings in
humans where neurons are only rarely found to be infected with HIV.
However, there were parallels between the hypoactivity of these mice
and the psychomotor slowing and apathy seen in the early stages of
AIDS dementia.  These experiments suggest that disease may be caused
not by the replication of the virus but by other related factors.
Of more specific relevance is the transgenic mouse constructed to
investigate the pathogenesis of HIV-associated nephropathy which
occurs in about 10 percent of HIV-infected patients.  The mice were
made transgenic for a subgenomic proviral HIV construct,
pNL4-3:d1443, which lacks the gag and pol genes and is therefore
non-infectious. Abnormal renal histology was noted beginning at about
35 to 45 days of age.  About 20 percent of these animals developed
cardiac lesions at the base of the aortic valve, an adventitial
process with macrophage and neutrophil infiltrates.  Vasculitis
involving the coronary arteries was also observed.  The interesting
aspect of this proviral construct was that its expression in heart
tissue could not be detected by Northern analysis, although low level
expression would only be detected by more sensitive methods. However,
there was increased expression of atrial natriuretic factor, a
molecular marker of myocyte hypertrophy, in the ventricles of these
One line of these transgenic mice carrying a subgenomic HIV proviral
construct lacking the gag and pol genes was found to develop
proliferative epidermal lesions with some similarities to those seen
in HIV-infected patients.  Dermal lesions similar to Kaposi's sarcoma
were not seen in this model, in contrast to those seen in an HIV
LTR-tat transgenic mouse.  Thus, the proteins expressed by the genes
appear to play an important role in pathogenesis of skin disorders in
HIV-infected patients.
There have been a few reports of the detection of HIV transcripts in
endomyocardial biopsies of HIV-infected patients, but the
significance of these findings is unclear since the transcripts have
been detected in cardiac tissues of patients with and without known
cardiac dysfunction. This leads to the speculation that viral
proteins, immune mechanisms and cytokine and growth factor
dysregulation might be implicated in the pathogenesis of cardiac
dysfunction.  It is possible that host genetic susceptibility may
also play a role since it has been shown that, in mice infected with
CVB3, macrophage inflammatory protein (MIP-l-alpha) is an absolute
requirement for development of myocarditis.  It is also possible that
the constant, repeated cycles of viral replications lead to mutations
with differential pathogenic effects which result in neurologic,
kidney, or heart disease, or cancer.
While some studies may be performed on cells, organs or whole
animals, many will require the development and exploitation of
transgenic animal models which could contribute to knowledge of the
pathogenesis of cardiac complications in patients with HIV infection.
However, because the presence of virus or viral proteins in a tissue
does not necessarily induce dysfunction or disease and because
cardiac dysfunction can occur in the absence of gross abnormalities
it is important to evaluate these animals in terms of physiologic
function and pathology of the cardiac disorders.  Although not yet
applied to the study of HIV-associated cardiac disorders, it has been
demonstrated recently that microsurgical techniques can be used to
study systolic and diastolic abnormalities that occur in transgenic
mice.  The use of such techniques has provided valuable mechanistic
information regarding adrenergic signaling and calcium handling
mechanisms that regulate ventricular function in vivo.  These
experiments suggest that similar approaches could be used to study
relevant issues of ventricular dysfunction associated with HIV
Research Objectives and Scope
The following items are provided as examples only. Applicants are
urged to consider other projects within the scope of this
solicitation based on their knowledge of the field and their
expertise.  Development of animal models is encouraged but
preliminary data must be provided to indicate that the model will
have relevance to the research requested.
o  Interaction of viral proteins with cardiovascular cells and the
effects on cardiac function in HIV infection
o  Examination of circulating antibodies for cross-reactivity with
cardiac tissue in HIV infection
o  Elucidation of the role of endothelium in the cardiovascular
complications of HIV infection
o  Role of the immune system and cytokine production in the etiology
of cardiac dysfunction in HIV infection
o  Evaluation of ventricular function and myocyte contractility in
transgenic mice that exhibit neuropathologic changes in HIV-related
o  Investigations in cell culture and whole heart of the mechanisms
whereby HIV infection perturbs the contractile function of the
o  Identification of the mechanisms whereby HIV mediates disorders of
impulse generation, conduction and EC coupling
o  Investigations of the alterations of gene expression of cardiac
contractile proteins, calcium regulatory proteins, and markers of
ventricular hypertrophy such as ANF in HIV-related complications
o  The role of coinfection with other pathogens, such as
cytomegalovirus, which have been implicated in HIV cardiomyopathy.
Upon initiation of the program, the NHLBI will sponsor periodic
meetings to encourage exchange of information among investigators who
participate in this program.  Travel funds for a one day meeting each
year, most likely to be held in Bethesda, Maryland should be included
in the modules. Applicants should also include a statement in their
applications indicating their willingness to participate in these
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 new policy results
from the NIH Revitalization Act of 1993 (Section 492B of Public Law
103-43) and 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.
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 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
additional relevant information concerning the policy.
Prospective applicants are asked to submit, by February 1, 1997, 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 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 a subsequent application, the information that it contains allows
NHLBI staff to estimate the potential review workload and avoid
conflict of interest in the review.
The letter of intent is to be sent to:
Dr. C. James Scheirer
Division of Extramural Affairs
National Heart, Lung, and Blood Institute
6701 Rockledge Drive, Room 7220, MSC 7924
Bethesda, MD  20892-7924
Telephone:  (301) 435-0266
FAX:  (301) 480-3541
Email:  James_Scheirer@NIH.GOV
The research grant application form PHS 398 (rev.5/95) is to be ujsed
in applying for these grants.  Applications kits are available at
most institutional offices of sponsored research and may be obtained
from the Grants Information Office, Office of Extramural Outreach and
Information Resources, National Institutes of Health, 6701 Rockledge
Drive, MSC 7910, Bethesda, MD 20892-7910, telephone 301/710-0267,
email:  ASKNIH@odrockm1.od.nih.gov.
The RFA label available in the application kit 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 and three
signed, 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 be sent to Dr. C. James Scheirer, at the same address given
above in the section on LETTER OF INTENT.
Sample budgets and justification page will be provided upon request
or following the submission of a letter of intent.
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:
Do not complete Form Page 4 of the PHS 398 (rev 5/95).  It is not
required nor will it be accepted at the time of application.
Do not complete the categorical budget tables on Form page 5 of the
PHS 398 (rev. 5/95).  Only the requested total direct costs line for
each year must be completed based on the number of $25,000 modules
being requested.  Applicants may not request a change in the amount
of each module.
A maximum of ten modules ($250,000 direct costs) per year may be
requested and each applicant may request up to five years of support
for this RFA.  Direct cost budgets will remain constant throughout
the life of the project (i.e., the same number of modules requested
for all budget periods).  Any necessary escalation should be
considered when determining the number of modules to be requested.
However, in the event that the number of modules requested must
change in any future year due to the nature of the research proposed,
appropriate justification must be provided.  Total Direct Costs for
the Entire Proposed Project Period should be shown in the box
Budget justifications should be provided under "Justifications" on
Form Page 5 of the PHS 398.
List the names, role on the project and proposed percent effort for
all project personnel (salaried or unsalaried) and provide a
narrative justification for each person based on his/her role on the
Identify all consultants by name and organizational affiliation and
describe the services to be performed.
Provide a general narrative justification for individual categories
(equipment, supplies, etc.) required to complete the work proposed.
More detailed justifications should be provided for high cost items.
Any large one-time purchases, such as large equipment requests, must
be accommodated within these limits.
o  CONSORTIUM/CONTRACTUAL COSTS - If collaborations or subcontracts
are involved that require transfer of funds from the grantee to other
institutions, it is necessary to establish formal subcontract
agreements with each collaborating institution.  A letter of intent
from each collaborating institution should be submitted with the
application.  Only the percentage of the consortium/contractual TOTAL
COSTS (direct and indirect) relative to the total DIRECT COSTS of the
overall project needs to be stated at this time.  The following
example should be used to indicate the percentage cost of the
consortium, "The consortium agreement represents 27% of overall
$175,000 direct costs requested in the first year.". A budget
justification for the consortium should be provided as described in
the "Budget Justification" section above (no Form Page 5 required for
the consortium).  Please indicate whether the consortium will be in
place for the entire project period and identify any future year
changes in the percentage relative to the parent grant.
If there is a possibility for an award, the applicant will be
requested to identify actual direct and indirect costs for all years
of the consortium.  Please note that total subcontract costs need not
be calculated in $25,000 modules. However, when subcontract funds are
added to the parent grant budget, the total direct cost amount must
be included in the number of $25,000 modules requested.
o  BIOGRAPHICAL SKETCH - A biographical sketch is required for all
key personnel, following the modified instructions below.  Do not
exceed the two-page limit for each person.
Complete the educational block at the top of the form page;
List current position(s) and those previous positions directly
relevant to the application;
List selected peer-reviewed publications directly relevant to the
proposed project, with full citation;
The applicant has the option to provide information on research
projects completed and/or research grants participated in during the
last five years that are relevant to the proposed project.
o  OTHER SUPPORT - Do not complete the "Other Support" pages (Form
Page 7).  Selected other support information relevant to the proposed
research may be included in the Biographical Sketch as indicated
above.  Complete Other Support information will be requested by NHLBI
staff if there is a possibility for an award.
o  CHECKLIST - No "Checklist" page is required as part of the initial
application.  A completed Checklist will be requested by NHLBI staff
if there is a possibility for an award.
o  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 RFA and will be returned without further review.
Applications must be received by April 24, 1997.  If an application
is received after that date, it will be returned to the applicant
without review.  The Division of Research Grants (DRG) 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 DRG 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.
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 NHLBI 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, assigned a priority score, and receive a second level
review by the National Heart, Lung, and Blood Advisory Council.
The personnel category will be reviewed for appropriate staffing
based on the requested percent effort.  The direct costs budget
request will be reviewed for consistency with the proposed methods
and specific aims.  Any budgetary adjustments recommended by the
reviewers will be in $25,000 modules.  The duration of support will
be reviewed to determine if it is appropriate to ensure successful
completion of the requested scope of the project.  Other review
criteria will include:
o  scientific, technical or medical significance and originality of
proposed research
o  appropriateness and adequacy of the experimental approach and
methodology proposed to carry out the research
o  qualifications and research experience of the Principal
Investigator and staff, particularly, but not exclusively, in the
area of the proposed research
o  availability of the resources necessary to perform the research.
The initial review group will also examine the provisions for the
protection of human and animal subjects and the safety of the
research environment.
Applications will compete for available funds with all other approved
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 concerning this RFA are encouraged.  The opportunity to
clarify any issues or questions from potential applicants is welcome.
Direct inquiries regarding programmatic issues to:
Dr. Constance Weinstein
Division of Heart and Vascular Diseases
National Heart, Lung, and Blood Institute
6701 Rockledge Drive, Suite 9044, MSC 7940
Bethesda, MD  20892-7940
Telephone:  (301) 435-0510
FAX:  (301) 480-1335
Direct inquiries regarding fiscal matters to:
Mr. William Darby
Grants Operations Branch
National Heart, Lung, and Blood Institute
6701 Rockledge Drive, Suite 7128, MSC 7926
Bethesda, MD  20892-7926
Telephone:  (301) 435-0177
FAX:  (301) 480-3310
This program is described in the Catalog of Federal Domestic
Assistance No. 93.837.  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, 42 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
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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