NATIONAL HEART, LUNG, AND BLOOD ADVISORY COUNCIL

MEETING MINUTES
February 7, 2002

I. CALL TO ORDER AND OPENING REMARKS – Dr. Claude Lenfant

Dr. Claude Lenfant opened the meeting and welcomed the Council members to the 205th meeting of the National Heart, Lung, and Blood Advisory Council (NHLBAC).  Dr. Lenfant reminded Council that February is Heart Month. Along with the American Heart Association, the American Red Cross, and the National Council of Aging, NHLBI has launched a campaign entitled "Act in Time to Heart Attack Signs".  The goal is to increase awareness and emphasize the importance of calling 911 at the first signs of heart attack symptoms.

Member Updates

Dr. Alcalay, Dr. Lipscomb, Dr. Ramirez, Dr. Whelton and Dr. Spragg were not present for the meeting.  Since the new Council members have not been approved yet, those Council members who had recently retired including Dr. Johnson, Dr. Martin, and Ms. Polite, were asked back.

Guests

Members from several Public Interest Organization (PIO) who had attended the Institute third annual Public Interest Organization Meeting on February 6, 2002, were in attendance.

Dr. Elizabeth Nabel, Director of Clinical Research, NHLBI, Dr. Charles Murry from the University of Washington, and Dr. Hal Broxmeyer from the University of Indiana were guest speakers at Council.

Personnel Announcements

Dr. Lenfant announced a number of new appointments: Dr. Barbara Alving is now the Deputy Director of the Institute; Dr. Charles Peterson is the Acting Director of the Division of Blood and Blood Diseases;                Dr. Deborah Beebe has been selected as the Director of the Division of Extramural Affairs; Dr. Robert Musson has been selected as the Deputy Director of the Division of Extramural Affairs; and Dr. Anne Clark has been selected as the Chief of the Review Branch.

New Publications

Dr. Lenfant noted three documents of interest to the Council namely the 2001 Facts Book, the public interest newsletter, and a document entitled "NHLBI Stem Cell Policy Update and NHLBI Review".  The latter document specifies the contact person in the Institute to answer questions on stem cell policy and initiatives and assure consistency.

II. REVIEW OF CONFIDENTIALITY & CONFLICT OF INTEREST – Dr.Claude Lenfant

The Council was reminded that according to Public Law 92-463, the Federal Advisory Committee Act, the meeting of the NHLBAC would be open to the public except during consideration of grant applications.  A notice of this meeting was published in the Federal Register indicating that it would start at 8:00 a.m. and remain open until approximately 11:00 a.m.  Dr. Lenfant also reminded the Council members that they are Special Government Employees and are subject to departmental conduct regulations. 

III.  REPORT OF THE DIRECTOR  

Dr. Lenfant presented an overview of the FY 2003 budget.  The budget has increased from $2.3 billion in FY 2001, to $2.6 billion in FY 2002.  In FY 2003, the budget will increase to nearly $2.8 billion.  A significant amount of money will be utilized for developing a program on bioterrorism as well as for continuing the war against cancer.  NHLBI will have an increase of over $200 million.  However, these figures may be modified after the Congressional hearings.  Dr. Lenfant also pointed out that the number of new and competing grants has also increased.  Dr. Lenfant noted that the funding policy of the NHLBI is to award grants at the fully recommended amount.

Dr. Lenfant announced that the Institute will issue the RFA for the Specialized Centers of Clinically Oriented Research (SCCOR) in Pediatric Heart Disease.  The original Specialized Centers of Research (SCOR) program has completed its first ten years and has been reviewed and evaluated.  A new program (SCCOR) has been developed and will emphasize clinical research.  An article describing the new program appeared in the January 28 volume of Circulation.  It is anticipated that most former SCOR programs will be renewed as SCCOR programs. 

The NIH Loan Repayment Program has been approved by the Office of Management and Budget and the applications are expected by the end of February.  They will be reviewed on April 19, 2002 and will be brought to May Council.  Dr. Lenfant pointed out that one issue concerns the definition of clinical research in terms of who is eligible.  Two recent editorials in the New England Journal of Medicine address this issue which has stirred considerable debate.  In addition, there is a question of limiting eligibility to those investigators who have been supported by NIH. These issues will be reexamined.

Dr. Lenfant explained the review process for these applications.  Each application will be evaluated on the basis of six different criteria: a personal statement on career goals, a statement on the proposed research, the training program to which the applicant belongs, the research environment where the applicant will be performing the research, letters of recommendation, and an institutional statement of support.  Applications will not be reviewed on scientific merit since it is assumed that this has already been evaluated as part of their research support.

Dr. Lenfant described the review committee which has been constituted with very senior individuals who are well-known and respected for their clinical research.  The Committee will be chaired by Dr. Eugene Braunwald who has been twice a council member and who is deeply committed to clinical research training.  There are four distinguished cardiologists, pulmonologists, and hematologists on the committee.

In response to a question from Dr. Martin, Dr. Lenfant noted that the number of applications cannot be predicted at this point. Dr. Williams asked whether the Pediatric Loan Repayment program was included and Dr. Lenfant responded that it is and will be reviewed at the same time.  Those applications will not be assigned to a specific Institute, however, for the Clinical Research Loan Repayment Program, at least fifteen percent of the awards will be for clinical pediatric research.

IV.  SCIENTIFIC PRESENTATIONS ON STEM CELL BIOLOGY - Dr. Elizabeth Nabel

Dr. Elizabeth Nabel, who is the Director of Clinical Research in the intramural research program of the NHLBI, gave a presentation about the activities related to stem cells in the programs under her direction.  The NHLBI hematology program has had a longstanding interest in stem cell biology.  Initial efforts were aimed at improving stem cell transduction using retroviral vectors and, more recently, novel viral vectors including lentiviral ventors.  Studies include stem cell marking studies and examining integration by retroviral vectors into different hematopoietic lineages.  A very large primate program was established and is still being supported. Many of the pre-clinical studies have been important for developing a number of phase one gene therapy protocols particularly for SCID and ADA deficiencies.  There have been very promising results regarding SCID and ADA gene therapy within the past couple of years.  Current efforts are looking at lentiviral vectors to introduce recombinant genes into hematopoietic lineages.  These studies are being done in collaboration with a number of investigators across the country.

The NHLBI hematology group is also involved in studies of stem cell failure.  Dr. Neil Young has been very interested in myelodysplastic disorders as well as aplastic anemia.  The hematology group initiated a bone marrow transplant program in the intramural program in 1993.  That program is very vigorous and is the largest in-patient clinical program in the intramural program.  About 150 patients have received bone marrow transplants over the past 5 years.  One of the major focuses of this bone marrow transplant program is treating stem cell failure.

The other area that has had a lot of growth is the allogeneic transplant program.  The goals of the bone marrow transplant program is to try to reduce transplant related mortality, minimize the pre-treatment regimen, and optimize graft versus tumor effects.  Dr. Rick Childs has been very involved in these studies along with Drs. Cindy Dunbar and Neil Young, who have experience with the use of a nonablative regimen followed by transplant using peripheral derived stem cells.  The group has tried to focus on some novel applications in solid tumors and Rick Childs has pioneered the use of nonablative peripheral stem cell transplantation for renal cell cancer.  Dr. Childs reported the results of his initial cohort of 26 patients in a phase one study in the New England Journal several years ago, showing that he could achieve a graft versus tumor effect.  These were very exciting results and Dr. Childs is now carrying on subsequent protocols trying to titrate the immune suppression of the tumor while minimizing graft versus host disease.

The other major area has been graft engineering in the leukemias.  This is an area of interest to Dr. John Barrett.  His protocols have focused on total body irradiation, followed by T-cell-depleted transplantation, delayed add back of T-cells, and finally cyclosporin immunosuppression.  The goal is to titrate graft versus tumor and graft versus host disease and considerable success has been achieved in these efforts.  The group is now expanding into autologous transplantation and looking at novel applications in autoimmune diseases.

The NHLBI cardiology group is beginning to look at mesenchymal stem cells and their potential to differentiate into multiple cell types to repair damaged tissue specifically myocardium.  A stem cell interest group has been formed in the intramural program.  Hematologists and cardiologists are now collaborating and designing protocols together.  One of the major areas that they have begun to look at is the area of myocardial remodeling.  One study demonstrated that within the area of regenerating myocardium there were areas of new myositis and new vascular tissue.  Using either Y-chromosome or GFP tagging, the group could document forty to fifty percent of the regenerating tissue derived from the bone marrow stem cells.  This very exciting study contradicted a preexisting concept that myocytes were terminally differentiated.

The major question is will bone marrow derived stem cells be mobilized and involved in myocardial remodeling?  The underlying question is whether administration of G-CSF will lead to mobilization of these bone marrow derived stem cells and will these stem cells home to regions of ischemic injury in a larger animal model?  Subsequent questions that need to be addressed are: Can bone marrow derived stem cells be identified in the myocardium?  What are the optimal mobilization factors, and what is the correct timing and dose?  Could G-CSF or other mobilization factors be delivered at the time of the myocardial infarct or for unstable angina?

The cardiology group is collaborating with an industrial group Osiris, located in Baltimore.  This group has derived autologous human mesenchymal stem cells grown in culture which can be injected directly into tissue or given intravenously.  The objective of the Phase One study is to determine the safety of intravenous delivery of allogeneic human mesenchymal stem cells in patients after a recent acute MI. Patients will come to the clinical center for their stem cell protocol.  The goal is to determine whether mesenchymal stem cells will improve regional left ventricular wall thickening in patients who have non-viable myocardium because of late presentation compared with placebo.

Dr. Letterman in the cardiology program has begun to apply MR imaging technology to evaluate patients. NHLBI has recently renovated the catheter lab so that a conventional fluoroscopy table and an MR table are contiguous.  Patients can be simultaneously evaluated with traditional cardiac catheterization as well as MR imaging technology.  The other intent, although not related to this particular protocol, is to begin to develop therapeutic techniques that can be performed directly in the MR suite.

In summary stem cell biology is a major focus in the NHLBI clinical program.  The goal is to explore the biology of stem cells and their potential application to human disease.  The intent is to move quickly from the pre-clinical stage into phase one studies to begin to address some of these initial safety and toxicity questions.

V.  THE ROLE OF STEM CELLS IN REPAIR OF DAMAGED CARDIAC MUSCLE-                 Dr. Charles Murry      

The second speaker, Dr. Charles Murry, who is on the faculty of the Department of Pathology at the University of Washington, gave a presentation about the role of stem cells in the repair of damaged cardiac muscles.  After myocardial infarct, there is the negative process of geometric remodeling that takes place in the heart.  The idea is that it may be possible to prevent the dilation of the left ventricle and the wall thinning associated with myocardial infarction by a cell-based therapeutic approach.  In fact, there is very good data to show that this has been done using skeletal-muscle transplantation.  Thus, it seems very likely that cellular therapeutics could be directed against left ventricular remodeling.

A second goal is to use hemangioblast type cells as a strategy to increase angiogenesis in a chronically ischemic heart and to have a cell-based angiogenic therapy.  There are pre-clinical data in rat models suggesting that human endothelial progenitor cells can in fact engraft in an infarcted rat heart and significantly improve both remodeling and contractile function in that model.

The ultimate goal is to provide something that would actually beat and restore systolic wall motion to the infarcted region of the heart.  The candidate cell types are the differentiated or committed cardiomyocytes, skeletal muscle cells, endothelial cells or their progenitors, fibroblasts, and finally pluripotent cells.

When cyanogenic cardiomyocytes were taken from neonatal animals and grafted into infarcted rat hearts, the results showed new cardiomyocytes which were elongated, rod-shaped, and appeared to be normal myocardium.  They went through a normal hypertrophic process so that eight weeks after grafting they were almost as hypertrophied as normal adult myocardium.  This followed the normal developmental time course for cardiomyocyte hypertrophy in the rat.  The intercellular junctions showed that the grafted cells formed normal reticulated disks.  The cells were joined mechanically by junctions containing N-cadherin and they were coupled electrically by junctions containing Connexin 43.

However, under low magnification, where the whole ventricle is viewed, the grafts are extremely small. Furthermore, no matter how many cells are added, the same-sized graft results.  The reason for this is that there is a tremendous amount of graft cell death after the cells are transplanted.  This indicates the limitation with using cardiomyocytes.  The cells don't divide after transfer similar to what happens to cells in an infarct.

The skeletal myoblast satellite cell is a committed stem cell that is essentially unipotent and has the capacity to form mature skeletal muscle.  If these cells are grafted into injured hearts, the grafts survive and are quite significant as demonstrated with skeletal muscle specific myosin staining.  In addition, the cells can be induced to twitch by electrically stimulating the graft in an organ chamber, ex vivo. At a higher frequency, the twitches start to superimpose and at higher frequency the graft goes into tetanus.  This is clearly indicative of physiological properties of skeletal muscle, not cardiac muscle, since cardiac muscle has a refractory period after each depolarization.  Sham-injected hearts show no contractile activity.  If skeletal muscle and cardiac muscle are co-cultured, they form synchronously contracting networks.  The interpretation is that the cardimyocytes or the pacemakers, when they capture the skeletal muscle, will induce synchronous contractions.  Unfortunately, when skeletal muscle is grafted into an injured heart, it downregulates the proteins that mediate this electromechanical coupling.  In vitro the skeletal muscle is capable of forming structures that are like an immature intercalated disk.  But once in vivo, it differentiates further and markedly down-regulates these proteins.  This is a major challenge to using skeletal muscle.

In another series of experiments cardiomyocyte transdifferentiation, by transgene activation, or by myosin expression, was not detected using lineage negative C-kit positive cells in multiple models that were examined. This was true whether there were co-cultures with embryonic stem cells or cardiomyocytes, and whether they were grafted in normal hearts, or whether they were grafted into injured hearts using multiple different injury models.  Thus, the data did not support the hypothesis that hematopoietic stem cells will differentiate to cardiomyocytes after grafting into the heart.

In some instances, men receive female hearts as an allograft.  In this situation, the Y-chromosome provides a marker for host cells rather than indigenous cells of the graft.  In five patients studied there was a very small, but a readily detectable, population of cardiomyocytes that contained the Y-chromosome.  Thus, a cardiomyosite that contained a Y-chromosome must not have come with the donor heart, but rather from the host.  A mean of about .02 percent of the total cardiomyocytes contained a Y-chromosome with a median value of .01 percent. This co-localized with areas of injury and areas of focal rejection.  In one patient in particular there were "hot spots" of up to 14 percent of the cardiomyocytes in a one millimeter square area that had Y-chromosomes.  It is concluded that human cardiografts contain a small, but a readily detectable, population of cardiomyocytes that come from somewhere outside the heart.

These co-localize with areas of rejection and this suggests that perhaps injury is required for mobilization or homing.

The critical questions that need to be addressed are: What is the cell and where is it coming from?  What are the factors that result in this stem cell mobilization?  What causes them to home?  What caused them to transdifferentiate?

Embryonic stem cells clearly have the best cardiogenic potential of any cell type that people have studied. Loren Fields' lab showed in 1996 that if you direct cardiomyocytes from embryonic stem cells, they will form grafts of perfectly normal looking cardiomyocytes in mouse hearts. However, the cardiomyocytes that are obtained from embryonic stem cells will probably be subject to all the limitations as primary cardiomyocytes, and will be susceptible to cell death.  Unless they are genetically modifed, they likely will not proliferate much after grafting.  Since there are immortalized cell lines, there are tremendous opportunities to genetically modify these cells in such a way so that survival, proliferation, and differentiation may be controlled.  There may be strategies to purify them from the unwanted cells.  However, human cells now exist and can be cultured and this is the direction for the future.  GERON has now optimized culture techniques to obtain 50 percent cardiomyocytes in an embryoid body that has been allowed to differentiate.  However, there is the possibility of tumorgenesis with embryonic cells.  If one does not have completely differentiated cells, there is a significant tumorogentic possibility that might create an intercardiac teratoma, an undesirable "side effect."  This would not be a therapeutic goal for patients with myocardium infarctions.

In embryonic stem cells, the big questions are: how do we guide their differentiation and how do we get them to become the cells that we are interested in, to the exclusion of many of the other lineages that these cells can take?  Also strategies for purifying the cardiomyocytes are important, particularly so we do not have tumorigenic cells that we are grafting into the heart.  With regard to adult stem cells, there is a cardiac progenitor in the adult and this appears to be a very rare event.  The goal would be to locate this cell anatomically and determine its phenotype as well as the cell type.  In summary, this approach may one day be therapeutic.  However, the emphasis should be on careful and deliberate studies.

VI.  HEMATOPOIETIC STEM CELL PLASTICITY -  Dr. Hal  Broxmeyer         

Dr. Hal Broxmeyer, who is the Chair of the Microbiology and Immunology Department, and a Professor of Medicine, at the Indiana University School of Medicine, gave a presentation about hematopoietic stem cell plasticity. During embryonic development, there are cells that give rise to endoderm, mesoderm, and ectoderm. Presumably there are embryonic stem cells that can give rise to all of these cells.  The question is are these cells found in adults and, if they are, in what frequency, and how can they be used?

There are data that suggest that neuro stem cells give rise to blood, muscle, and multiple embryonic tissues. There are other data which suggest that skeletal muscle gives rise to blood cells.  Most of the studies have used non-purified or at best semi-purified cell populations from muscle, and these have repopulated the hematopoiesis system.  There could be at least two interpretations: 1) there is a common cell that gives rise to both muscle and blood or; (2) there are separate cells that give rise to only muscle, or to only blood, but they happen to co-reside in the muscle.

Following reconstitution of irradiated mice with genetically marked marrow, the stem cells in the muscle that reconstituted hematopoiesis were all of donor origin.  This suggested the existence of marrow to muscle homing of hematopoiesis stem cells and the co-existence in muscle of distinct stem cell types.  The results did not, however, rule out the existence of a common muscle hematopoiesis stem cell.

The following questions are posed.  When the cells of the hematopoietic stem cell phenotype differentiate into cells of other tissues and lineages, do the differentiated cells of the new lineage maintain any markers or characteristics of the hematopoietic cells?  Alternatively, can one detect intermediate cells that share the markers' characteristics of the hematopoietic cells and the new lineage cells being formed?  What are the receptors on these stem cells that allow them to find their way to tissue other than marrow or lymphoid tissue if it is indeed a hematopoietic stem cell that is being plastic? Are the specific receptors, the homing receptors, different from those that would allow a hematopoietic stem cell to home to the marrow and all lymphoid tissue? CXCR4, a chemokine receptor, has been implicated in homing of hematopoietic stem cells.  Is this or other chemokine receptors involved in this homing?

Assuming that plasticity is rigorously proven, the regulation of the growth of these multi-tissue stem cells and their production of lineage specific cells both in vitro and in vivo in terms of these functional activities should be investigated.  The challenge is to direct them to the correct location in the body to produce the desired lineage tissue restrictive cell types in high enough quantity to be clinically useful cells.  Caution should be exercised due to the possibility that abnormal growth and/or differentiation of these cells might lead to malignant tumors.  We need to know if one stem cell type can differentiate into any tissue that derives from the three germ layers of the embryo: the ectoderm, the mesoderm, and the endoderm. Is there going to be a hierarchy of multi-tissue stem cells, where some will be more committed with limited capacity, and maybe they will only give rise to one or two, rather than three, of the germ cell layer types?  Are we going to have a stem cell that gives rise to all three, to combinations of the two, or to one, and is there going to be a parent-progeny relationship between these cells?  Will single adult stem cells have the same degree of plasticity that is currently being attributed to embryonic stem cells?  Is there a cell that comes before hematopoietic stem cells?  What is the impact of this if it turns out to be as good as all of us are hoping that it is going to be?  The frequency of the cells found may come from cells of another lineage that is crucial to the potential impact of plasticity for clinical usage.  If stem cells, hematopoietic stem cells, are going to be used to repair cells of other lineages, can enough of these new cells be produced to be useful?  Can the hematopoietic stem cell that has the added capability to form cells of other tissue/lineages be expanded?  These are all subject for further endeavors in stem cell biology.

Council Discussion

Council asked whether the regulations that are being debated in Congress pose a barrier to these studies?     Dr. Murry stated that the embryonic stem cells approved for use are probably sufficient.  These may be able to be used as an allograft for clinical purposes.  Council questioned whether clinical use is being studied.  Dr. Murry replied that there are three clinical trials for skeletal muscle transplantation for cardiac repair.  Dr. Nabel commented that there are cases to justify proceeding with Phase One studies. Council questioned the relevance of the mouse heart model and Dr. Murry stated that the use of the model is justified.  Dr. Broxmeyer agreed that clinical research should proceed alongside basic research. Council also questioned how much proprietary information being gathered by biotech companies will be made available and Dr. Broxmeyer commented that this information is not as forthcoming and that much more is learned from work being done in the public sector.

Dr. Lenfant commented that it is extremely important to be cautious with moving the basic research in to the clinical arena since the gene therapy studies have had problems despite the much anticipated results.  Council concurred that well thought-out developmental biology studies should be simultaneously conducted for maximum benefit.

VII.  PUBLIC INTEREST ORGANIZATION (PIO) MEETING   

Many public interest organizations, each representing different diseases, attended the Institute’S third annual PIO meeting on February 6, 2002.  This is an opportunity for exchange between these groups and the Institute. The goal is to involve the public in determining priority areas of health research.  The Office of the Director, NIH has convened a group called the Council of Public Representatives which meets regularly during the year.

Four members of Council attended the PIO meeting and were asked to comment.  They noted that it was a very successful meeting and that there were excellent presentations and opportunities for learning about the NHLBI.  One improvement would be to allow time for sharing information, ie., networking among the different groups.  Dr. Hedrick from the National Emphysema/COPD association expressed his gratitude for the opportunities provided by this meeting as well as for the sincere effort which was made to listen to their concerns.  Dr. Lenfant remarked on the devotion of the different groups, large and small, to achieve results. The NHLBI is revising its website to include better access to information. Ms. Fulton from the National Sarcoidosis Society emphasized the importance of interacting with the medical research community on Sarcoidosis.  Ms. Connor from the Society of Women’s Health research also thanked NHLBI staff for the meeting and described the research they are interested in.

Dr. Lenfant concluded the discussion by reiterating the commitment of the Institute to this endeavor.

VIII.  TOPICS FOR MAY COUNCIL 

Dr. Lenfant apprized Council that the next meeting would include the review of the Loan Repayment Program applications.  Council will be involved in review of the Contracts to ensure the transparency of the review process.  In addition, Council will review the Board of Extramural Advisors initiatives.

GENERAL RECOMMENDATIONS

Dr. Lenfant asked Council to approve the Council-delegated authorities.  These have not changed since last year and are very important in terms of expediting awards.  Council recommended approval.

CLOSED PORTION

This portion of the meeting was closed to the public in accordance with the determination that it was concerned with matters exempt from mandatory disclosure under Sections 552b(c)(4) and 552b(c)(6), Title 5, U.S. Code and Section 10(d) of the Federal Advisory Committee Act, as amended (5 U.S.C. appendix 2).

There was a discussion of procedures and policies regarding voting and confidentiality of application materials, committee discussions and recommendations.  Members absented themselves from the meeting during discussion of and voting on applications from their own institutions, or other applications in which there was a potential conflict of interest, real or apparent.  Members were asked to sign a statement to this effect.

IX.  REVIEW OF APPLICATIONS

The Council considered 774 applications requesting $911,306,527 in total direct costs.  The Council recommended 757 applications with total direct costs of $897,516.422.  A summary of applications by activity code may be found in Attachment B.

ADJOURNMENT

The meeting was adjourned at 3:00 p.m. on February 7, 2002.

CERTIFICATION

I hereby certify that the foregoing minutes are accurate and complete.

Claude Lenfant, M.D.

Chairperson

National Heart, Lung and Blood Advisory Council

on 04/09/2002