National Heart, Lung, and Blood Institute (NHLBI)

The National Heart, Lung, and Blood Institute provides leadership for a national program in the causes, diagnosis, treatment, and prevention of diseases of the heart, blood vessels, lungs, and blood, and sleep disorders; and in the uses of blood and the management of blood resources. It conducts and supports, through research in its own laboratories and through extramural research grants and contracts, an integrated and coordinated program that includes basic investigations, clinical trials, epidemiological studies, and demonstration and education projects.

While the major part of the research supported by the NHLBI addresses common conditions such as hypertension, coronary heart disease, and chronic obstructive pulmonary disease, a significant amount of research is devoted to rare diseases. In Fiscal Year 1998, the NHLBI supported research on sudden and unexplained death in sleep, various lipid disorders, congenital cardiovascular malformations, infectious myocarditis, aortic and cerebral aneurysms, cystic fibrosis, primary pulmonary hypertension, sarcoidosis, sickle cell disease, various anemias, leukemia, and hemophilia.

Recent Scientific Aadvances in Rare Diseases Research

Heart and Vascular Diseases Program

Abetalipoproteinemia is a recessive disorder characterized by the absence of very low density lipoproteins (VLDL) and low density lipoproteins (LDL) from plasma. Fat malabsorption is severe and triglyceride accumulation occurs. Acanthocytosis of erythrocytes is common. Spinocerebellar ataxia, peripheral neuropathy, retinopathy, and ceroid myopathy all appear to be secondary to defects of transport of tocopherol in blood. Genetic, biochemical, and nutritional approaches to study certain aspects of the disease are underway in an NHLBI program project grant. Genetically-manipulated animal models recently have been created to obtain an understanding of the etiology and genetics of the disease. Treatment involves reduction of dietary fat and supplementation with tocopherol.

Patients with antiphospholipid syndrome (APS) have circulating autoantibodies to certain phospholipids, chiefly cardiolipin and/or phospholipid, as well as lupus anticoagulant. APS is clinically manifested by recurrent venous and arterial thrombosis, history of fetal deaths, and autoimmune thrombocytopenia. Although many of the patients with APS have systematic lupus erythematosus (SLE), APS may also occur in the absence of lupus. A NHLBI-supported project is studying the molecular genetics of apolipoprotein H, which may affect the binding properties of antiphospholipid antibodies, in lupus patients. This research will further the understanding of the lupus risk and production of antiphospholipid antibodies. Additional research is underway to develop more standardized immunoassays to reliably detect individuals with moderate or high antiphospholipid antibodies, which could ultimately improve treatment of atherosclerosis.

Congenital heart disease occurs in approximately 1/100 live births. Of those, about one-third, or about 12,000 children born per year, have complex congenital heart disease, which involves many different malformations. Congenital heart disease is an important cause of infant mortality, pediatrics and adult morbidity, as well as shortened adult life expectancy. There is growing evidence that congenital heart disease, previously thought to be essentially random, has a genetic basis. NHLBI-supported researchers have identified two relevant genetic loci: 1) one on human chromosome 1, related to a complex congenital cardiac malformation known as atrioventricular canal defect; and 2) the other on human chromosome 18, that was previously associated only with immune modulation, but appears to be involved in the development of two of the heart's four valves. Other NHLBI grantees have made significant progress refining the genetic information about heart outflow defects (so-called conotruncal defects) and Di George syndrome, which affects development of the heart, as well as other organs. Other studies have produced important epidemiological data on late mortality in long-standing congenital heart defects. Additional innovative research currently underway may lead to fetal therapy of congenital heart defects in the coming years. Moreover, one researcher supported by NHLBI is developing a system to allow fetal electrocardiograms to be performed, which would be a significant clinical advance, since no direct way to evaluate abnormal fetal heart rhythms currently exists.

Dysbetalipoproteinemia is a disorder with a strong heritable component characterized by the presence of beta migrating VLDL. It leads to the formation of characteristic xanthomas and predisposes to early ischemic heart disease and peripheral vascular disease. Research into the genetics and the biochemical events to understand its etiology and pathophysiology are underway in one NHLBI grant. A mutant form of apo-E (E2) has been discovered to be the primary molecular defect. The role of apo-E variants in remnant retention currently is being studied at several locations.

Familial hypercholesterolemia (FH) is an inherited autosomal dominant trait characterized by elevated concentrations of LDL. Cholesterol derived from LDL is deposited in arteries causing heart attacks and in tendons and skin causing xanthomas. The defect in FH is a mutation in the gene specifying the receptor for plasma LDL, which facilitates LDL removal. When the receptors are deficient or absent, the rate of LDL removal declines and the level of LDL rises. The homozygous form is rare [one in a million], but persons with it are highly prone to premature coronary heart disease. Recent scientific advances include the generation of animal models and a better understanding of the regulation of LDL receptor activity. Apheresis approaches to remove excess LDL from plasma are progressing, and gene therapy attempts to generate LDL receptors in the liver are underway. In addition, the NHLBI supports several grants that study the biochemistry, genetics, and potential treatment of the disease. A major program project supports research on various aspects of regulating LDL receptor and cholesterol levels in the blood. NHLBI intramural scientists are also evaluating the extent of atherosclerosis in patients with FH. Both invasive and non-invasive techniques are being applied to determine which method will be most effective in the selection of treatment programs and prognosis of the disease.

Familial hypertrophic cardiomyopathy (FHC) is a disease that is transmitted in an autosomal dominant fashion. The disease is associated with myofibrillar disarray and leads to cardiac hypertrophy. While patients are asymptomatic, the disease may lead to shortness of breath, palpitations, heart failure or sudden death. Some individuals with the disease die during childhood whereas others survive to the 6th to 7th decades of their lives. In addition, the disease is associated with mutations in various proteins which suggest that FHC may not be a single disease, but rather may represent a heterogeneous group of disorders. During the past decade scientists have made significant progress in uncovering the genes associated with the disease. It is known, for instance, that FHC can be caused by many different mutations in contractile proteins that make up the heart wall. However, understanding who will die suddenly or whether certain factors, such as high blood pressure or extreme stress, will trigger sudden death remains elusive. Several groups of geneticists and cardiologists have now developed a mouse model of FHC that should greatly enhance genetic studies of the disease. The NHLBI supports research on the genetic basis and mechanisms involved through several investigator-initiated grants and in two Heart Failure Specialized Center of Research (SCOR) grants.

Hypoalphalipoproteinemia represents a group of rare inborn errors of lipid metabolism that affect high density lipoprotein (HDL) levels. The errors include apo-AI synthetic and structural defects, defects in lipoprotein lipase or its co-factor apo C-II, hepatic triglyceride lipase deficiency, lecithin:cholesterol acyltransferase deficiency, and deficient activity of cholesteryl ester transfer protein. Some, but not all, are associated with a marked increase in the risk for atherosclerosis. Research into the genetics and the biochemical events to better understand the various correlations among the interrelated metabolic pathways is underway in one NHLBI-supported grant. In addition, animal models are being created to generate knowledge and design strategies for treatment of the diseases. NHLBI intramural researchers are directing their efforts toward the identification and characterization of the genetic defect leading to low HDL levels and heart disease. This research has recently identified the cellular defect in familial hypoalphalipoproteinemia as a defect in the ability to remove excess intracellular cholesterol, which leads to an accumulation of cellular cholesterol and early heart disease.

Infectious myocarditis, which affects both children and adults, is characterized by inflammation of heart muscle, sometimes leading to progressive heart failure and the need for heart transplantation. The NHLBI supports grants that are investigating various aspects of myocardial inflammation, including virus-receptor interactions. One grantee has identified a novel receptor component that may be responsible for many of the clinical manifestations of myocarditis. In addition, a large clinical trial is under development to determine the safety and efficacy of intravenous gamma globulin treatment for children with myocarditis.

Lecithin cholesterol acyltransferase (LCAT) deficiency results in a cloudy cornea, a marked decrease in plasma HDL, and renal disease. The NHLBI intramural Molecular Disease Branch has succeeded in developing the first knockout mouse model for LCAT deficiency and has used it to discover that the renal lesions present in the model are similar to the lesions present in patients with the LCAT deficiency.

The long QT syndrome (LQTS) is identified by a prolonged QT segment on an electrocardiogram and is associated with syncope, ventricular arrhythmias, and, frequently, sudden cardiac death. Studies have found that LQTS is often inherited and is related to mutations in membrane ion channels. In some forms of the disease, afflicted individuals may inherit other abnormalities, such as deafness, and have varied clinical outcomes depending on their specific mutational patterns. About 70 percent of diagnosed cases are in women. Effects of age and gender, as well as genotype specific impact on clinical course for three major chromosomal subtypes, have recently been explored. Other recent scientific advances have provided emerging clues on the role of newly described channel accessory proteins in modulating the disease and on the efficacy of pharmaco-genetic therapies. The NHLBI is supporting research on the molecular genetic mechanisms of LQTS in more than six investigator-initiated grants and in one Sudden Cardiac Death Specialized Center of Research (SCOR) grant. One new award supports the first study on using genotype-specific pharmacologic therapy. In addition, the NHLBI supports a patient registry for this disease, which is continually updated by worldwide referrals. The LQTS registry is currently providing data for a multi-center collaborative study sponsored by the NHLBI to investigate the population and genetic heterogeneity of this heritable disease. In addition, a prospective longitudinal follow-up of LQTS families is designed to provide clinical data and samples from well-characterized LQTS families for statistical genetic and genetic linkage studies. This research could potentially improve presymptomatic diagnosis of LQTS.

The muscular dystrophies are a clinically and genetically heterogeneous group of skeletal muscle-wasting diseases that differ widely in their frequency and pattern of cardiac involvement. Myocardial disease manifesting predominantly as cardiomyopathy and congestive heart failure is characteristic of Duchenne and Becker muscular dystrophies and X-linked dilated cardiomyopathy, whereas conduction system abnormalities that cause heart block, arrhythmias, and sudden death are more commonly seen in limb-girdle type 1B, myotonic, and Emery-Dreifuss muscular dystrophies. Mutations in the gene encoding dystrophin, a membrane-associated protein in muscle and brain, yield a clinical spectrum that includes muscular dystrophy, cardiomyopathy, and mental retardation. This year a NHLBI-supported study concluded that amino acid changes in caveolin-3, a dystrophin-containing protein, are disease-causing mutations, rather than benign polymorphisms. This research could lead to an understanding of the role of caveolin-3 in the dystrophic process.

Smith-Lemli-Opitz (SLO) syndrome is an inherited autosomal recessive disorder caused by a defect in the enzyme that catalyzes the last reaction in cholesterol biosynthesis. As a result, deficient cholesterol is produced and the precursor 7-dehydrocholesterol and its derivatives accumulate. Newborns with SLO have a distinctive appearance with specific facial dysmorphism and suffer from multiple congenital anomalies including cleft palate, congenital heart disease, genitourinary abnormalities, and malformed limbs. Development of refined and simple biochemical diagnostic methods is in progress, and nutritional studies of a high cholesterol intake are encouraging. NHLBI-supported research is generating animal models and developing biochemical methods for diagnosis. This research is also investigating the feasibility of identifying possible genetic mutations.

Tangier disease is a genetic dyslipoproteinemia characterized by very low plasma HDL levels and increased risk of premature cardiovascular disease. Investigators supported by the NHLBI are studying the biochemistry, metabolism, and genetics of this disease. Recently, other studies have demonstrated defective cholesterol efflux from Tangier cells, which helps explain the cholesterol deposits in various tissues. In addition, the genetic locus has been identified and a number of researchers are trying to identify the gene.

Lung Diseases Program

Alpha-1-antitrypsin (AAT) deficiency is an inherited absence of a circulating proteinase inhibitor, alpha-1 antitrypsin, that is manufactured primarily in the liver. Deficiency states (circulating serum AAT levels below 0.6 mg/ml) are associated with emphysema, presumably from inadequate protection of enzymatic destruction by neutrophil elastase. Fifteen percent of the AAT-deficient population also develops liver disease. The NHLBI currently funds a variety of clinical and basic research on AAT deficiency, which includes the study of the molecular mechanisms that impair secretion of AAT; inherited traits or environmental factors that exaggerate the accumulation of AAT or response to it; methods of gene therapy delivery; personality, coping styles and quality of life of AAT patients; and how to increase availability of defective, but partially active AAT. In addition to the research that specifically focuses on AAT, the NHLBI supports a number of grants that study the function, synthesis, secretion, and interaction of similar enzymes. The Institute also supports a AAT patient registry with the primary objective of examining the decline in FEV1 and mortality in relation to augmentation therapy. The Registry was initiated in1988 and enrolled 1,129 patients who were followed 3.5 to 7 years. Data analyses was completed in 1998. The data showed the overall registry mortality was 11 percent at 3 years and 19 percent at 5 years. Overall, recipients of augmentation therapy demonstrated a lower mortality rate compared to those who did not receive augmentation therapy. The results are consistent with a possible beneficial effect of augmentation therapy; however, the results must be interpreted cautiously, since the Registry was not designed as a clinical trial with a control group. The primary results from the Registry were published in the American Journal of Respiratory and Critical Care Medicine.

Asbestosis is the interstitial pneumonitis and fibrosis caused by exposure to asbestos fibers. In response to the deposition of asbestosis fibers, macrophages and lymphocytes accumulate, type II alveolar epithelial cells and smooth muscle cells proliferate, fibrosis appears in the adjacent walls of respiratory airways, and septa thicken. NHLBI-supported research is investigating the molecular and cellular events that trigger the cellular accumulation and proliferation and regulate remodeling of lung tissue that results in the fibrotic lesions in response to asbestos. Inhaled asbestos fibers induce the formation of reactive nitrogen species which are capable of causing damage to lung tissue and are likely to play a role in tissue damage seen in asbestosis. In an investigator-initiated NHLBI-supported study, researchers are exploring the mechanisms governing mesothelial cell activation in asbestos-induced injury. Additionally, the NHLBI supports three centers that foster multidisciplinary basic and clinical research to enable basic science findings in fibrotic lung diseases, including asbestosis, to be more rapidly applied to clinical problems.

Approximately 30 percent of the very low birth weight infants who survive prematurity develop bronchopulmonary dysplasia (BPD), a chronic lung disease characterized by disordered lung growth, i.e., alterations in cell size and shape, as well as decreased numbers of alveoli available for gas exchange. The current view is that BPD is a developmental disorder in which lung immaturity, mechanical ventilation, and oxygen toxicity all appear to be involved, with the developmental stage of the lung at birth being the paramount determinant. The NHLBI supports investigations on a molecular characterization of BPD developed in a uniform, non-human primate model of the disease. Additional NHLBI-supported research has recently identified the accumulation of two markers, 3-nitro-tyrosine in plasma and bombesin in urine, as possible predictors of the development of BPD in very low birth weight premature infants. Further research recently published in the Journal of Clinical Investigation indicated that postnatal administration of blocking anti-BLP is protective against the development of clinical and pathological evidence of BPD. In addition, dilation of pulmonary vessels with inhaled low dose NO has been observed to reverse some of the deleterious lung remodeling characteristic of BPD. A current hypothesis is that NO can decrease the regional hypoxemia created by the inflammatory component of BPD. Nitrotyrosine levels, used to monitor the production of oxygen radical species, were consistent with the safety of short-term use of NO therapy.

Cystic fibrosis (CF) is a multi system disease characterized by defective transport of chloride and sodium across the cell membrane. It is the nation's number one genetic killer of children and young adults. Ninety percent of persons with CF die from pulmonary complications. More than 25,000 Americans have CF, with an incidence of about 1 in 3,300 among Caucasians. The responsible gene, the CF transmembrane conductance regulator (CFTR), was identified in 1989. More than 600 mutations and DNA sequence variations identified in the CFTR gene contribute to its highly variable presentation and course. The most common mutation on the CF gene causes the CFTR protein to be misprocessed so that it never reaches the cell surface. Strategies are now being developed to overcome these trafficking problems and allow movement of the CFTR protein to the cell surface, where it serves as an opening or channel that lets out chloride. A NHLBI-sponsored pilot clinical trial that is treating CF patients who have this common mutation with the experimental drug phenylbutyrate, a non-toxic drug currently approved for treatment of metabolic (urea cycle) disorders, recently revealed partial restoration of CFTR function. This finding extends earlier in vitro studies demonstrating correction of the CF defect and restoration of CFTR-mediated chloride transport. Because this therapeutic approach circumvents many of the problems associated with other strategies for restoration of CFTR function, namely gene delivery systems and long-term persistence, it marks an important clinical advance. A clinical trial for assessing dosage and safety is presently underway, as well as studies identifying the precise mechanisms by which this drug works. Other NHLBI-supported researchers are studying the interrelationship between epithelial ion transport, airway surface liquid (ASL) physiology, and airway defenses in the normal lung. This knowledge is crucial to gain insight into how these relationships are disrupted in CF, leading to the vicious cycle of infection and inflammation with the eventual destruction of the lung and for treating CF lung disease. In addition, a NHLBI-supported study developed a novel strategy to circumvent a major barrier to gene therapy for CF lung disease.

Idiopathic pulmonary fibrosis (IPF), a disease of excessive wound healing resulting from injury to the lungs, is commonly treated with corticosteroids, sometimes in combination with other potent drugs. Despite some success with this therapy, pulmonary fibrosis in most patients worsens over a relatively short time. Recent estimates put the U.S. prevalence at 185,000 and the incidence at 70,000. NHLBI-supported research focuses on the molecular and cellular events that trigger the inflammation of alveoli seen in the early stage of IPF and that influence progression to the irreversible, fibrotic, end stage of the disease. Some NHLBI-supported researchers have participated in a clinical trial of interferon-beta in idiopathic pulmonary fibrosis and have developed a scoring system that can be used to assess the status of the patient's disease and predict and assess response to treatment. Other research is exploring whether CD40, a cell surface molecule through which fibroblasts can be activated by C40-ligand, is a critical element in mediating pulmonary inflammation, dysplasia, and fibrosis.

Lymphangioleiomyomatosis (LAM) is a rare lung disease that affects women, usually during their reproductive years. Symptoms develop as the result of the proliferation of atypical, non-malignant smooth muscle cells in the lungs. Diagnosis is usually made by lung biopsy. Common symptoms include shortness of breath, coughing, and sometimes coughing up blood. Patients often develop spontaneous pneumothorax or chylous pleural effusion (collapse of the lung or collection of milky looking fluid around the lung). The clinical course of LAM is quite variable, but is usually slowly progressive, eventually resulting in death from respiratory failure. No treatment has proven effective. Some patients with Tuberous Sclerosis Complex (TSC), a genetically-transmitted disease, develop lung lesions identical to those seen in LAM. In some cases, the clinical distinction between TSC and LAM may be difficult.

NHLBI supports research on LAM through both its intramural and extramural programs. As part of the intramural program, the Institute recently established a LAM Section in the Pulmonary and Critical Care Medicine Branch, where investigators are studying LAM patients to learn more about the cause and progression of the disease at the clinical, cellular, and molecular levels.

After patients with LAM were found to have a high incidence (more than 80 percent) of osteopenia and osteoporosis, a treatment protocol was initiated to prevent and/or reverse the excessive bone loss. In addition, these researchers are attempting to determine the characteristics of the unusual smooth muscle cells that damage the lungs of LAM patients. An important aspect of the research is learning how growth is regulated in these cells. The NHLBI extramural program supports a national LAM patient registry that is coordinated by the Cleveland Clinic Foundation. Patients can be enrolled through six major participating Centers (including the NIH Clinical Center), their personal physicians, and through the LAM Foundation, a private patient advocacy organization that publicizes NHLBI and other LAM research activities and that provides information about the disease to LAM patients and their families. The LAM registry, which began enrolling patients in the summer of 1998, will help facilitate the collection of clinical data and will identify a cohort of LAM patients who might be contacted in the future, if opportunities for clinical studies arise. In addition, NHLBI is supporting extramural research on the genetic mechanisms leading to smooth muscle proliferation in LAM, genetic mutations found in TSC, and the relationship between LAM and TSC. In a recent scientific advance, researchers conducted an immunohistochemical analysis that suggested the production of proteins that inhibit cell death in LAM cells may be controlled by hormones. This may be a clue to understanding the imbalance that causes overgrowth of these cells.

Primary pulmonary hypertension (PPH) is a rare, inherited, progressive, and fatal disease characterized by proliferative occlusion of small pulmonary arteries. PPH usually leads to death within 3 years of diagnosis. The mode of inheritance and genetic features of PPH are not fully understood. Women, usually of childbearing age, are affected twice as frequently as men. The use of appetite suppressants has recently been associated with the development of PPH. Although the risk of PPH is low among users of these drugs (probably only 1 in 1,000), it is not possible to predict which users might develop the disease. Some progress was made in this area this year when NHLBI-supported researchers found an abnormal endothelial growth pattern in patients who develop PPH following exposure to appetite suppressant drugs. The NHLBI also supports a family registry for PPH that maintains records and blood samples from families and individuals for use in the study of familial PPH inheritance and pathogenesis. Data from this registry have revealed that approximately 6 percent of the cases of PPH are due to familial PPH. Other research is seeking to increase the understanding of the cause of the proliferative and obliterative changes of the vasculature that characterizes PPH. Other studies are examining the balance between agents, such as nitric oxide, that cause vasodilation or vasoconstriction. Additionally, researchers are seeking to improve drug delivery for PPH patients.

Pulmonary alveolar proteinosis (PAP) is a rare disorder characterized by an abnormal accumulation in the alveoli of the normal constituents of surfactant. PAP has an estimated prevalence of 1 per 100,000. Congenital PAP is a rare disorder affecting newborn infants. The congenital form of PAP is associated with an hereditary deficiency of surfactant protein B. Acquired PAP may occur in association with a variety of conditions including silicosis and certain malignancies. The NHLBI supports investigator-initiated grants that seek to define the molecular basis of PAP and to determine the incidence and pathogenesis of congenital surfactant protein B deficiency. Mutations that alter the folding, processing, or interaction with lipids of SP-B may be clinically significant in human infants and may play a role in the development of respiratory distress syndrome. Studies of gender and race differences and of familial inherited differences in risk and frequency of respiratory distress have demonstrated a genetic contribution. These data suggest further population based studies are needed to assess frequency of potentially important genetic markers. A rare (1/1,000,000) mutation in the gene for Sp-B, 121ins2, has been identified by examination of cord blood specimens archived by the Center for Blood Research in New York and the Missouri Department of Health as components of the birth-death certificate data base for infant deaths from PAP. Researchers are performing molecular analysis of other populations to determine the extent of the impact of ethnic composition on the frequency of 121ins2 mutation.

Persistent pulmonary hypertension of the newborn (PPHN) occurs in approximately one in 1,000 live term births. With this condition, the lung arteries of newborns fail to dilate to allow for normal blood flow to the lung, causing poor oxygenation. Affected infants require costly and prolonged medical care including intubation of the airway, inhalation of 100 percent oxygen, mechanical ventilation, and often heart/lung bypass (extracorporeal membrane oxygenation). Experimental therapies that offer promise far less invasive treatment include inhaled nitric oxide (NO). Recent studies point to a critical role for NO as a modulator of levels of endogenous vasoactive mediators that regulate pulmonary vascular tone and vessel wall structure. NO regulates both endothelin and vascular endothelial growth factor production, potentially leading to alterations in vascular tone and permeability. A greater understanding of the molecular mechanisms by which NO regulates gene expression under hypoxic conditions and the role it may play in modifying the vascular responses may have widespread implications for the treatment of PPHN.

Sarcoidosis is a systemic disease involving organ systems throughout the body, in which normal tissue is invaded by pockets of inflammatory cells known as granuloma. Most sarcoid patients have granuloma in their lungs. The disease can exist in a mild form that spontaneously disappears or a severe form that results in a life-long condition. Estimates of the number of Americans afflicted with the disease range from 13,000 to 134,000, and between 2,600 and 27,000 new cases appear each year. As many as 5 percent of individuals with pulmonary sarcoidosis die of causes directly related to the disease and yet current treatment with corticosteroids may be toxic and ineffective. The cause(s) of sarcoidosis are presently unknown, but disease development is thought to involve the victim's immune system. The NHLBI is supporting both laboratory-based research to investigate granuloma formation and clinical research to obtain a better understanding of the process of this extremely complex disease. The NHLBI is supporting 10 clinical sites and a coordinating center in an etiologic study that will use information obtained from study participants to identify exposures to environmental factors, family characteristics, and co-existing diseases that contribute to sarcoidosis. Other studies focus on the genetic epidemiology of the disease and the genetic makeups that predispose for the development of sarcoidosis and their interaction with environmental risk factors. Another study is examining the role of the immunologic and inflammatory processes mediating granulomatous inflammation in pulmonary sarcoidosis and has recently uncovered clues in potential ways to treat the disease. In other recent advances, the angiotensin converting enzyme genotype was reported to potentially play a more important role in sarcoidosis susceptibility and progression in African Americans than Caucasians.

Blood Diseases and Resources Programs

Aplastic anemia affects about one in 10,000 individuals and occurs at any age. The often-fatal disease results in bone marrow failure, which in turn causes infections from lack of white blood cells, anemia from lack of red blood cells, and bleeding from lack of platelets. The cause is largely unknown, but an immune mechanism appears to be responsible for suppressing bone marrow function in the majority of affected individuals. Research findings in the areas of bone marrow transplantation (BMT), stem cell transplantation, transplant-regimen-related toxicity, chronic graft versus-host-disease, and late effects of transplant have significantly enhanced the lives of aplastic anemia patients. The probability of survival after BMT from an HLA-identical sibling has improved significantly over the past few years and at present is about 80 percent in AA patients under 16 years old. Transplantation from alternative donors, however, faces unsolved immunological problems.

The use of antithymocyte globulin (ATG) immunosuppression to treat AA was pioneered in the NHLBI intramural Hematology Branch. Current clinical protocols compare the effect of ATG versus cyclophosphamide immunosuppression in achieving lasting remissions and prolonged survival in severe AA. Early results from a randomized study show that treatment with cyclophosphamide requires longer hospitalizations than treatments with ATG. No difference in response rate has been identified to date but longer follow-up time will be required. In another recent study, a mouse model for aplastic anemia illustrated the role of interferon-( (IFN-() in producing bone marrow damage. A commonly used hematopoietic growth factor, called granulocyte colony stimulating factor, was shown to change the quality of cytokine response, decreasing lymphocyte expression of IFN-( associated with bone marrow suppression. Other intramural hematology research is directed at identifying viral agents causing the disease and understanding interactions of cytokines with bone marrow stem cells. In addition, to coordinating and collaborating on an ongoing epidemiological study of AA in East Asia, the intramural hematology program has also established a Cooperative Research and Development Agreement (CRADA) with MedImmune to test the feasibility and efficacy of a vaccine for parvovirus, which causes anemia as a result of bone marrow failure. The vaccine could be an effective treatment for several diseases, including sickle cell and aplastic anemia.

Blackfan-Diamond anemia is a rare congenital disorder resulting in failure of red blood cell development with subsequent anemia. The etiology is unknown and the only current cure is matched-sibling stem cell transplantation. The NHLBI intramural program is currently enrolling patients in a clinical trial to assess the benefits and effects of antithymocyte globulin and cyclosporine in combination as a possible treatment. In the laboratory, researchers are using in vitro assays to try to determine the pathogenesis of the disease, as well as any prognostic indicators in terms of response to treatment and overall clinical course.

Cooley's anemia (also called beta-thalassemia, thalassemia major, or Mediterranean anemia) is a genetic blood disease that results in an inadequate production of hemoglobin. Affected individuals require frequent and lifelong blood transfusions to sustain life. Because there are no natural means for the body to eliminate iron, the iron contained in the transfused red blood cells builds up over many years and eventually becomes toxic to tissues and organ systems. NHLBI grantees have used a beta-globin gene/beta-locus control region retroviral vector to optimize gene transfer and expression in a mouse transplant model. The results demonstrate for the first time high-level, long-term somatic human beta-globin gene transfer into the hematopoietic stem cell of an animal and suggest the potential feasibility of a retroviral gene therapy approach to the beta thalassemias and to sickle cell disease. A number of compounds have been identified that can induce hemoglobin F production, which occurs inadequately in Cooley's anemia. Several new iron chelating drugs are also in various stages of development. The NHLBI is working to establish a Cooley's Anemia clinical research network, which will build an infrastructure to encourage high quality, coordinated multi-center efforts in clinical research and therapeutic development.

Fanconi anemia (FA) is an autosomal recessive bone marrow failure syndrome characterized by pancytopenia, developmental defects, DNA cross-linking agent hypersensitivity, and cancer susceptibility. Many FA patients can be identified at birth because of congenital anomalies, although approximately 25 percent have no birth defects. FA is a clinically heterogeneous disorder and until recently had been divided into at least five different complementation groups designated A through E. Two FA genes, A and C, which account for an estimated 75 percent of all FA patients world-wide, were cloned by investigators in the NHLBI's intramural hematology program. Eight distinct complementation groups have now been reported, suggesting that at least 8 genes are involved in the manifestations of Fanconi Anemia. A third gene, FAG, has recently been characterized. NHLBI-supported research is focusing on the interrelationship and functions of these FA genes, as well as the identification and cloning of the remaining FA genes. NHLBI intramural research this year provided clues about how FA genes may be involved in controlling how cells die. NHLBI intramural scientists are also pursuing a CRADA with Signal Pharmaceuticals to study the mechanisms of FA cell death induced by a molecule that attaches to the cell surface.

Graft-versus-host disease (GVHD) may occur after a bone marrow or blood stem cell transplant if the marrow donor's lymphocytes produce an immune reaction against the skin, alimentary tract, and liver of the recipient. Both the acute and chronic forms can be fatal. The risk of GVHD rises with increasing disparity of matching between the donor and the recipient, and GVHD currently limits safe transplantation of marrow cells to well-matched donor-recipient pairs. The NHLBI intramural Hematology Branch is currently investigating ways to prevent GVHD by removing lymphocytes from the transplant followed by delayed add-back of donor lymphocytes several months after the transplant to confer necessary immune responses against infections and the patient's leukemia. This involves developing new methods to process peripheral blood stem cell transplants and removing donor lymphocytes that cause GVHD. Thus far, this approach has resulted in a low incidence of acute GVHD (less than 20 percent) and no severe or fatal disease.

Hemophilia is a hereditary bleeding disorder that results from a deficiency in either blood coagulation factor VIII or factor IX. There are about 20,000 individuals in the United States with hemophilia. They are dependent on lifelong treatment to control periodic bleeding episodes. Gene therapy studies by NHLBI-supported scientists have demonstrated long-term correction of hemophilia B in dogs using recombinant adeno-associated virus (rAAV) vectors when administered either by intramuscular injection for muscle tissue expression or intrahepatic administration for liver expression. NHLBI scientists and Targeted Genetics Corporation have entered into a CRADA to study the potential of using rAAV vectors to develop gene therapies for hemophilia and other diseases.

Hereditary hemorrhagic telangiectasia (HHT) is a rare bleeding disorder due to weakness of the vascular support structures. The most common manifestations are red spots on the lips and nose bleeding. In an advanced stage of the disease, there are often arterio-venous malformations in the lung, brain, gut, and liver. In NHLBI-supported genetic studies on HHT, two gene defects have been identified. The first gene defect is in the protein, endoglin, which is a growth factor receptor on the endothelial cell surface. The second gene defect is related to activin receptor-like kinase 1. A third gene also appears to be involved in HHT but its identity remains unknown. A correlation may exist between the gene defect and the organ specificity of the disease. This information may be useful in the diagnosis and prognosis of HHT.

Antibodies produced against platelet surface antigens lead to thrombocytopenia (a decrease in blood platelets) and bleeding. Acute autoimmune idiopathic thrombocytopenic purpura (ITP) is a milder disorder but in its chronic adult form is a clinical challenge. This disease is about three times more prevalent in women than men. ITP has been considered a paradigm of autoimmune diseases whose understanding and methods may be applicable to similar disorders. Recombinant thrombopoietin has been produced and clinical trials are underway to determine its safety and efficacy in the treatment of thrombocytopenia. Researchers are applying new technology to understand the production of the antiplatelet antibodies, its genetic regulation, and design of possible therapeutics. Initial results from studies on blocking autoantibody production in ITP appear promising.

Leukemia is a disorder of the bone marrow which has a progressive and fatal outcome. Acute leukemias (Acute Lymphocytic and Acute Myeloblastic Leukemia) have a more rapid progression then chronic leukemias (Chronic Lymphocytic and Chronic Myelogenous). Leukemias can arise in lymphocytes or bone marrow cells, causing suppression of normal bone marrow production of red cells, neutrophils, and platelets that results in anemia, infections, and bleeding, respectively. Accumulation of leukemia cells in the bone marrow, blood, and other tissues gives rise to bone pains; wasting; fevers; enlargement of liver, spleen, and lymph glands and occasionally meningitis. Patients with leukemia who are incurable by chemotherapy are eligible for bone marrow transplantation using matched or mismatched donors. The experimental treatment protocols evaluate methods to reduce transplant related mortality and increase the anti-leukemic effect of the transplant. This year, the NHLBI intramural Hematology Branch safely and effectively performed bone marrow transplants of lower intensity ("mini-transplants") in patients up to the age of 68 years with minimal side effects. NHLBI intramural researchers are also collaborating with the NIH Clinical Center's Department of Transfusion Medicine to evaluate the "Nexcell 300i" cell processor, which permits the selection of large numbers of stem cells with depletion of lymphocytes to provide optimum donations of marrow stem cells to recipients.

Multiple myeloma is a malignancy of the B cell lineage, resulting in uncontrolled cloning growth of plasma cells. These cells cause bone destruction, resulting in pain and fractures, and secrete large amounts of an abnormal immunoglobulin, that can result in renal failure, neurologic complications, poor immune function, and other serious complications. Standard chemotherapy can improve symptoms, but is not curative. Recently, autologous stem cell transplantation has been shown to improve survival, but is also not curative. Allogeneic transplantation may be curative, but is highly toxic in this patient population. Researchers in the NHLBI intramural Hematology Branch are enrolling patients in clinical trials to study both of these transplantation approaches. Recent intramural laboratory studies are investigating the role of the herpes virus KSHV in myeloma, which has been reported by one group to be involved in the pathogenesis of myeloma. NHLBI intramural studies have found little evidence for the involvement of this virus, but instead suggest that a related virus may be activated in myeloma patients who have been previously infected.

Paroxysmal nocturnal hemoglobinuria (PNH) is an acquired stem cell disorder characterized by anemia resulting from destruction of red cells. Hemolysis results in the absence of a critical protein (CD59) on the red cell surface. Other complications of the disorder include life-threatening thromboses of peripheral and abdominal veins. PNH is increasingly being characterized in patients with aplastic anemia and who have successfully undergone treatment. The Hematology Branch pursues a program investigating the abnormal bone marrow cells in patients with and without a preceding history of aplastic anemia. Additional studies are aimed at correcting the CD59 protein defect by transfer of normal proteins to defective cells. This research has demonstrated that the missing proteins responsible for the normal cell's resistance to complement inhibitor can be transferred to abnormal cells in culture and correct the hemolytic propensity. In another intramural study of a large group of aplastic anemia patients receiving immunosuppressive therapy, abnormal PNH cells were present prior to therapy in most cases. Immunosuppressive therapy alone did not predispose patients to develop PNH. Other studies investigating the abnormally elevated prevalence of clotting abnormalities in these patients identified low levels of plasma heparin cofactor, as well as decreased or absent urokinase activator receptor on platelets. The role of these abnormalities in producing thrombosis is under investigation.

Sickle cell anemia is the most common genetic blood disorder in the U.S., affecting approximately 1 in 500 African-American and 1 in 1000 Hispanic newborns each year; with an estimated 72,000 affected individuals now alive in the U.S. It occurs when an infant inherits the gene for the sickle hemoglobin from both parents (sickle cell anemia) or the gene for sickle hemoglobin from one parent and the gene for another abnormal hemoglobin from the other parent (sickle cell disease types, e.g. Hb SC, Hb S-Beta thalassemia). In patients with the disease, the hemoglobin molecules in the red blood cells which carry oxygen throughout the body tend to damage the red cell walls because they are rigid and have a characteristic crescent or sickle shape, causing them to stick to blood vessel walls and block blood flow. This leads to sickle cell crises or painful episodes, which are the hallmark of the disease. Chronic end organ damage occurs to the vital organs because of blockage of blood flow to the brain, lungs, kidneys, spleen, and liver. Approximately 10 percent of children with sickle cell anemia experience strokes, which are among the most devastating complications of this disease. This year, the New England Journal of Medicine published the results of the NHLBI-supported Stroke Prevention Trial in Sickle Cell Anemia (STOP). This landmark study showed that administering regular blood transfusions, every three to four weeks, to children with sickle cell anemia who are at high risk for stroke reduced their rate of stroke by 90 percent. The data were so compelling that the study was stopped 16 months earlier than planned and an NHLBI Clinical Alert was issued to approximately 5,000 U.S. physicians who treat children with sickle cell anemia. The current sickle cell disease research portfolio includes research on the following topic areas: gene therapy for sickle cell disease, bone marrow transplantation as a possible cure, and clinical trials of drugs that increase the levels of fetal hemoglobin in sickle cell disease patients.

Thrombotic thrombocytopenic purpura (TTP), a potentially fatal disease, is characterized by widespread platelet thrombi in arterioles and capillaries. Relapse is not uncommon in those who survive the acute phase. Despite treatment advances, TTP associated mortality has increased. Both endothelial cell damage and intravascular platelet aggregation have been suggested in the pathogenesis of TTP. Microscopic examination of the thrombi has revealed the presence of abundant von Willebrand factor (vWF) and NHLBI-supported research has shown that interactions between vWF and platelet surface glycoproteins are intimately involved in thrombus formation. In the normal blood circulation, vWF is cleaved to small units by a plasma enzyme. The presence of inhibitory antibodies to this enzyme in patients with acute TTP is being reported and offers a possible opportunity for therapeutic intervention.

Rare Diseases Research Initiatives

Initiatives Begun in 1998

Clinical Research on Cooley's Anemia: new Request for Applications to support clinically-related research focused on improving the clinical management of patients with Cooley's anemia. Areas of particular importance include studies on improved methods for the non-invasive measurement of tissue iron burden, alternative approaches to iron chelation therapy, and pharmacologic enhancement of fetal hemoglobin.

Comprehensive Sickle Cell Centers: renewal of Request for Applications to expedite the development and application of new knowledge for the diagnosis, prevention, and treatment of sickle cell disease by supporting basic, clinical, and behavioral research, as well as research training. In addition, the Centers provide programs in diagnosis, education, and counseling. Fifty-five awards have been made in seven competitions since 1972. The previous competition in 1993 included 10 awards. New components in the eighth competition include a Data Management Core, a Sickle Cell Research Scholars Program, and optional Pilot Projects for promising, but untested, innovative research.

Immunogenetics of Inhibitor Formation in Hemophilia: new Request for Applications to improve understanding of inhibitor formation and immune tolerance induction in hemophilia patients. The goals of this initiative are to be able to predict those patients most likely to develop inhibitors, to specifically and safely block inhibitor formation, and to induce tolerance or neutralize existing inhibitors, so that the risks and suffering caused by antibody inhibitors is reduced or eliminated.

Specialized Centers of Research (SCOR) in: Neurobiology of Sleep and Sleep Apnea: new Request for Applications to integrate molecular, cellular, and genetic approaches to sleep control with clinical investigations on the etiology and pathogenesis of sleep disorders, particularly sleep apnea. Airway Biology and Pathogenesis of Cystic Fibrosis (CF): new Request for Applications to use current knowledge of the CF transmembrane conductance regulator (CFTR) as a focus to promote advances in research on the pathogenesis of CF, the role of CFTR in airway biology, and the development of new treatment strategies.

Acute Lung Injury: new Request for Applications to determine the interaction of the different mediator sequences in the inflammatory/immune systems, examine the factors that promote fibrosis as opposed to healing after tissue injury, explore interactions between the lung and other organs in both animals and humans, and evaluate the genetic control of the response to tissue injury in humans.

Thrombocytopenia: Pathogenesis and Treatment: new Request for Applications to explore how a better understanding of the pathogenesis of idiopathic thrombocytopenic purpura may increase our understanding of thrombocytopenia in general and HIV-related thrombocytopenia in particular. The long term goal of the initiative is to develop better therapeutic approaches for all of these conditions.

Vascular and Hematopoietic Development and Disease: new Program Announcement to define the critical processes that direct differentiation and organization of the vascular system with specific emphasis on factors that specify the diverse phenotypes of endothelial, blood, and vascular smooth muscle cells. This program announcement encourages innovative approaches to identify and characterize precursor cells, studies to elucidate the regulatory mechanisms which determine and maintain the diverse phenotypes, and strategies to define how these developmental mechanisms might be involved in pathological conditions of the mature animal. The ultimate goal is to provide the foundation for new therapies to treat cardiovascular, lung, and blood diseases based on morphogenetic principles.

Initiatives Planned to Begin in FY 1999

Creutzfeldt-Jakob Disease (CJD) Assay Methods Development: new Request for Applications to develop assay methods for the detection of CJD, a disease that, along with other transmissible spongiform encephalopathies, has not been studied extensively due to the lack of a satisfactory assay system. The immediate goal of this program is the development of an assay to screen donated blood and donors of organs or tissues, although other uses of the technique are probable. Approaches that might be considered for mass screening for the hallmark abnormal isoform (PRP-res) or other potential markers for CJD are serologic, virologic, physical, and physico-chemical.

Developmental Processes in Differential Expression of Globin Genes: new Program Announcement to pursue basic investigations into the developmental processes involved in the differential expression of globin genes. This joint program announcement with the National Institute of Diabetes and Digestive and Kidney Diseases is designed to elucidate mutations in the globin gene cluster that cause some of the most common inherited diseases in humans, such as sickle cell anemia and thalassemia (Cooley's anemia). Current treatment for these disorders includes both pharmaceutical agents and gene therapy. However, not all patients respond to the pharmaceutical agents, their applicability to other disorders is not established, and the molecular and cellular basis for their mode of action remains unclear. In addition, studies of the developmental-and tissue-specific control of the globin gene cluster are important for optimizing gene transfer, via viral or chemical vectors, for gene replacement therapy. This program announcement promotes the further advances needed to broaden the range of possible therapeutic strategies for disorders of globin synthesis.

Specialized Centers of Research (SCOR) in Pediatric Cardiovascular Disease: renewal of Request for Applications to foster interdisciplinary studies of the etiology, pathophysiology, and diagnosis of congenital and acquired cardiovascular disease in children in a context that will lead to more effective methods of treatment and prevention. The goals of the program are to encourage: 1) exploitation of the latest techniques of developmental and molecular biology, as well as cellular and organ physiology, to elucidate the underlying mechanisms which are perturbed in pediatric cardiovascular disease states; and 2) application of fundamental knowledge and modern technology to improve diagnosis, treatment, and prevention of these diseases.

Stem Cell Transplantation to Establish Allochimerism: new Request for Applications to develop improved and novel preparative regimens that will permit incompatible hematopoietic stem cell transplantation in immunized recipients with hemoglobinopathies, such as sickle cell disease and Cooley's anemia. This initiative will explore the possibility of performing successful stem cell transplantation for hemoglobinopathies where complete destruction of the bone marrow (myeloablation) is not desirable and partial replacement of defective marrow may be sufficient for clinical benefit. The overall goal of the initiative is to enable successful stem transplantation for hemoglobinopathies and minimize recipient morbidity and mortality. Proposed approaches must possess the potential to evolve into human clinical studies.

Strategies to Augment Alveolization: new Request for Applications to define the critical processes that direct the formation and functional differentiation of lung alveoli and determine whether similar principles apply to the induction of alveolar regeneration. Recent progress in developmental biology has opened new opportunities to expand the study of morphogenesis to the molecular level. The information generated by a full exploration of the genetic, cellular, and molecular mechanisms regulating alveolar formation should ultimately lead to the development of rational interventions for aberrant development of the lungs, for lung injury, and for diseases such as emphysema, diffuse interstitial fibrosis, and bronchopulmonary dysplasia.

Rare Diseases-Related Program Activities

Workshops, Symposia, and Meetings

Workshop on Molecular Embryology of the Lung: Then, Now, and in the Future (June 1998). Purpose: Inventory progress since the original Molecular Embryology of the Lung Workshop, held in June 1992, that generated interest in performing these types of studies. Results: The 1998 workshop stimulated interest in performing additional studies on the molecular embryology of the lung, which has ramifications for bronchopulmonary dysplasia and other rare lung disease. Participants identified those areas where lack of basic information about lung development remains a barrier to the development of a rational therapeutic approach. Discussion also stressed the need for high throughput screening technology and databases for gene expression comparison.

Workshop on Pharmacological Therapy for Idiopathic Pulmonary Fibrosis (IPF): Past, Present, and Future (September 1998). Jointly supported by the NHLBI and the Office of Rare Diseases. Participants identified new promising approaches to treating IPF, which would benefit from a clinical trial, and what elements are important to consider when designing such trials. A summary report has been submitted for publication.

Specialized Center of Research (SCOR) in Pathobiology of Fibrotic Lung Disease Program Investigators Meeting (September 1998). Purpose: to examine current and future research and areas of potential cooperation in fibrotic lung diseases, such as asbestosis, idiopathic pulmonary fibrosis, and persistent pulmonary hypertension of the newborn. Results: At the meeting, discussions addressed the need for multi-center clinical trials; collaboration on assessing surrogate markers for disease status and response to therapy; presence of unique fibroblast phenotypes in pulmonary fibrosis lesions; potential candidates for use in clinical trials; and validation of a scoring system for determining disease status and predicting disease progression.

Annual Sickle Cell Disease Clinical Research Meetings (August 1998). Purpose: Convene the steering committees of the epidemiologic and clinical trials that the NHLBI supports in sickle cell disease. Results: Researchers from the Pediatric Hydroyurea Study Group, MSH Patient's Follow-up, Cooperative Study of Sickle Cell Disease; and Stroke Prevention Trial in Sickle Cell Anemia participated and discussed operation of clinical studies, clinical research findings, and upcoming publications.

Problem Areas Related to Rare Diseases

Due to the complexity of congenital heart disease, much work is still needed to understand abnormal heart development. In addition, because even relatively large congenital heart disease centers do not see large numbers of children with particular defects, there is a pressing need for congenital heart disease clinical trials networks to answer important therapeutic questions, such as the role of medicines successful in adults for children with similar conditions, and the optimal therapy for complex malformations.

Individuals enrolled in the Long QT Syndrome Registry are referred by physicians familiar with this program. Because this is a rare syndrome and not a reportable disease, specific minority demographic information may not be complete. NHLBI investigators are trying to increase awareness of this registry to the African-American community by becoming involved with the Association of Black Cardiologists.

There is a shortage of product for patients with alpha-1-antitrypsin deficiency needing augmentation therapy. The only product that has been approved by the Food and Drug Administration (FDA) was endorsed without evidence of clinical benefit. However, new products are in various phases of evaluation and there is much debate whether a clinical trial is needed to assess the clinical efficacy of these new products before they are approved. The NHLBI has participated in several FDA-convened meetings to discuss the design of clinical trials to determine dosage, route of administration, and clinical efficacy of the new products, as well as the one approved product that is currently in clinical use. The FDA and the NHLBI have also discussed a potential NHLBI role in clinical trial(s) of augmentation therapy.

There is a large variability existing in the incidence of bronchopulmonary dysplasia (BPD) among hospital centers that is likely associated with differences in ventilatory strategies and other support practices. This issue needs further exploration.

Scarcity of data and tissue has hindered learning about the etiology of lymphangioleiomyomatosis. The small number of patients makes it difficult to learn about the prevalence, prognosis, clinical course, and effect of various treatments. The NHLBI-sponsored national LAM patient registry will facilitate collection of clinical data and identify a cohort of LAM patients who might be contacted in the future, if opportunities for clinical studies arise.

The pharmaceutical industry has demonstrated little or no effort to support research on Fanconi anemia, perhaps because of a perceived lack of marketability. Consequently, it has been extremely difficult to initiate clinical trials.

Approximately 20 percent of severe hemophilia patients develop antibody inhibitors, which specifically neutralize the activity of the replacement factor and complicate treatment. An initiative in this area has resulted in six new grant awards that support research to improve understanding of the mechanism of inhibitor formation so that risk and suffering is reduced for individuals with inhibitors.

Diagnosis of patients with hereditary hemorrhagic telangiectasia, particularly in younger individuals, is often difficult due to an overlapping constellation of clinical symptoms that becomes more pronounced with age. Establishment of a genetic linkage may allow earlier diagnosis and better treatment.

The bone marrow stem cell transplant program, used for treating leukemia patients who are incurable by chemotherpay, has expanded rapidly in 1998 to an annual transplant total of 43 patients. The number is predicted to double in 1999. This increase has created a waiting list, decreased bed availability, and put additional demands on the NIH Clinical Center Transfusion Medicine Department. The Clinical Center is actively working to accommodate this increased clinical activity.