David Bodine joined the Clinical Hematology Branch (CHB), NHLBI, in 1984 as a postdoc. He remained with the CHB until 1993, when he joined NCHGR as the Chief of the Hematopoiesis Section. Bodine did his graduate work at Jackson Laboratory in Bar Harbor, Maine, and received his Ph.D. from the University of Maine in 1984.

Recent research in my laboratory has focused on pluripotent hematopoietic stem cells (PHSCs). PHSCs are the ultimate progenitors of all circulating blood cells and have the ability to self-renew numerous times without losing the ability to differentiate into cells as diverse as red blood cells and T lymphocytes. PHSCs are found in the bone marrow, and due to the great proliferative capacity of their descendants, they are exceedingly rare, less than one per 100,000 bone marrow cells. We have recently concentrated our efforts on describing gene expression in PHSCs and devising methods to introduce genes into PHSCs via retrovirus-mediated gene transfer.

To study gene expression in PHSCs, these rare cells must be greatly enriched. Our work, using mice as a model system, has shown that murine PHSCs express high levels of c-kit, the receptor for the hematopoietic growth factor, on their surface. By combining this observation with techniques to subtract cells expressing markers of mature blood cells, we were able to use fluorescence-activated cell sorting to isolate a population of cells highly enriched in PHSCs. It only takes 100 of these cells to fully reconstitute the hematopoietic system of a mouse, at least a 1,000-fold enrichment over the concentration of PHSCs in the original bone marrow cell population. Examination of messenger RNA purified from those highly enriched PHSCs revealed that c-kit, stem cell factor (SCF), and the receptors for the hematopoietic growth factors interleukin-3 (IL-3) and interleukin-6 (IL-6) were all expressed at high levels. Other work showed that when bone marrow cells were cultured for six days in SCF, IL-3, and IL-6, the number of PHSCs in the cultures increased two to threefold.

Efficient retrovirus-mediated gene transfer requires that the target cell divide, thus allowing the virus to become integrated into the host-cell DNA. Our finding that PHSC numbers could be increased by culture in SCF, IL-3, and IL-6 suggested that treatment with these growth factors might significantly increase the frequency of retrovirus-mediated gene transfer into these cells. Our results showed that the frequency of gene transfer into mouse PHSCs cultured without growth factor or in just a single growth factor was approximately 5%. The frequency of gene-transfer into mouse PHSCs cultured in all three factors was as high as 75%. These observations were successfully extended into a rhesus monkey model, and the success of our monkey gene transfer experiments helped serve as preclinical justification for human gene-therapy experiments in which retroviruses containing the adenosine deaminase (ADA) gene were introduced into the bone marrow of patients with severe combined immunodeficiency syndrome. These human trials were performed in Europe and the United States in 1993.

In the future, we hope to isolate novel genes that control PHSC division and differentiation from c-DNA libraries generated from mRNA from highly enriched populations of PHSCs. In addition, we are continuing our efforts to further define the factors required for the growth and differentiation of PHSCs both in vitro and in vivo.

Mark Boguski received his M.D. and Ph.D. from Washington University in St. Louis in 1986, and in 1988, he joined the Mathematical Research Branch, NIDDK, as a medical staff fellow. He became one of the first staff members of the newly formed National Center for Biotechnology Information (NCBI) in 1989, where he remains as a researcher in the Computational Biology Branch.

My earlier work at NIH focused on sequence motifs and conserved domains in proteins involved in signal transduction, particularly those that interact with and regulate GTPases. Although I continue to study the interrelationships of sequence, structure, and function in proteins, I have also been working on information analysis and retrieval problems in genome research. Three years ago, NCBI's Carolyn Tolstoshev and I founded the database of expressed sequence tags (dbEST) which is a division of GenBank for cDNA sequence and mapping data. Now, dbEST contains more than 100,000 sequences, has been queried by researchers more than 100,000 times, and is currently used nearly 7,000 times per month by intramural and extramural scientists.

We are now collaborating with researchers who are working on genetic and physical mapping to build a "transcript map" of the human genome. Only a small fraction of human DNA, probably less than 5%, consists of transcribed coding sequences. Our goal is to locate all of these transcribed coding sequences, starting with a comprehensive set of cDNAs, called expressed sequence tags, and map them back with high resolution onto the chromosomes. Such a map will help us to understand gene regulation, to pinpoint gene-rich regions for concerted genomic-sequencing efforts, and to greatly accelerate positional cloning of genes responsible for genetically complex diseases such as diabetes.

I am also collaborating with Phil Hieter's group at the Johns Hopkins University of School of Medicine in Baltimore on a project to identify and map all homologous genes in the yeast and human genomes. In so many instances, such as the recent studies of cystic fibrosis, neurofibromatosis and familial colon cancer, yeast biochemistry has shed tremendous light on human pathophysiology. However, these connections are usually made late in the research process, and much effort and expense could be saved if the relationships are identified earlier. Now, however, we're working toward making both the complete sequence of the Saccharomyces cervisiae genome and a comprehensive sampling of human coding sequences available within the next 18 months. This creates an unprecedented opportunity to construct a molecular cross-reference between yeast and humans and to populate the human-genome map with yeast-gene functions and phenotypes. This will facilitate the identification of candidate genes for human diseases and the development of assay systems for studying the functions of human gene products.

Seong-Jin Kim received his Ph.D. from the Tsukuba University in Japan in 1987. He came to NIH in 1987 and is currently a visiting scientist at the Laboratory of Chemoprevention, Division of Cancer Etiology, NCI.

Our laboratory has been studying the transcriptional and posttranscriptional regulation of the set of three homologous isoforms of transforming growth factor-ß (TGF-ß), TGF-ßs 1, 2, and 3. My research program has focused on the regulation of the TGF-ß gene by etiologic agents that are involved in disease: tumor-suppressor genes (retinoblastoma gene and Wilm's tumor gene), oncogenes (jun, fos, src, abl, and ras), and viruses (human T-lymphocyte virus type 1, human cytomegalovirus, and hepatitis B virus). Taken together, these studies of gene regulation have delineated the molecular basis for the observation that the type 1 isoform of TGF-ß is upregulated by cells in response to injury and pathological processes such as fibrogenesis and carcinogenesis. In contrast, the type 2 and 3 isoforms of TGF-ß are regulated principally by developmental cues and hormones.

We have also demonstrated that the protein encoded by the retinoblastoma susceptibility (Rb) gene can regulate expression of the TGF-ß1 and -ß2 genes through the Sp1 and activating transcription factor-2 (ATF-2) binding sites in the TGF-ß1 and TGF-ß1 promotors, respectively. In the latter case, ATF-2 can form a complex with the Rb protein with the help of an additional, and as-yet-unidentified, bridging protein. We are currently trying to clone the gene that encodes the bridging protein.

I have also been interested in posttranscriptional regulation of TGF-ß isoforms. It has been suggested that TGF-ß expression is regulated at the posttranscriptional level by members of the steroid-retinoid superfamily of nuclear receptors in an isoform-specific manner. Retinoic acid stabilizes TGF-ß2 mRNA, while the serum cholesterol-lowering drug lovastatin specifically downregulates the TGF-ß2 mRNA through a posttranscriptional mechanism. We are attempting to identify the factors controlling the stability TGF-ß2 mRNA.

Most recently, we have begun to explore mechanisms that regulate the expression of the TGF-ß receptors in human gastric cancer cell lines that are resistant to the growth-inhibitory effect of TGF-ß. We have found alterations in the gene that encodes the type II receptor for TGF-ß. One of our current goals is to characterize the mechanisms of transcriptional regulation of the TGF-ß type II receptor gene because several TGF-ß-resistant cell lines that displayed no alterations of the TGF-ß type II receptor gene expressed no detectable TGF-ß type II receptor mRNA.

Lois Travis received her M.D. from the University of Florida College of Medicine in Gainesville, Fla., in 1980. She received her Sc.D. in Epidemiology from the Harvard School of Public Health in Boston in 1994 and joined the Radiation Epidemiology Branch of the Epidemiology and Biostatistics Program, NCI, in 1989.

One of my major areas of interest is the study of multiple primary cancers, particularly the evaluation of cancer risk following exposure to ionizing radiation and/or chemotherapeutic drugs. As long-term-survival rates improve for many types of cancer patients, it becomes critical to identify the late consequences of therapy. One of the most serious side effects of cancer treatment is the induction of new malignancies. Characterization of therapy-related risks is crucial to enabling the clinician to make an informed decision regarding treatment, balancing efficacy against acute and chronic sequelae. In addition, quantification of the late effects of cytotoxic drugs and radiation therapy provides a unique opportunity for interdisciplinary studies of carcinogenesis because humans are deliberately treated with measured amounts of potentially cancer-inducing agents.

When I arrived at NCI, little work had been carried out in the area of secondary cancers following therapy for non-Hodgkin's lymphoma (NHL). Since then, in collaboration with investigators worldwide, our group has provided quantitative estimates of the risk of secondary malignancies among several populations of NHL patients. In one of my first projects, we identified a significant excess of solid tumors after NHL, noting that the pattern of risk increased with time, consistent with the late effects of treatment. In a subsequent study, we showed that the increased risk of secondary malignancies persisted for up to two decades after the initial NHL diagnosis. We also quantified the association between the risk of secondary leukemia and the dose of various cytotoxic drugs, such prednimustine, chlorambucil, and cyclophosphamide. We found a dose-response relationship between the cumulative amount of cyclophosphamide and bladder cancer, and we described the combined effect of cyclophosphamide and radiotherapy in the induction of bladder cancer. Now, with the Laboratory of Human Carcinogenesis at NCI, we are collaborating on molecular studies examining the mutational spectrum of the p53 tumor-suppressor gene in cyclophosphamide-related bladder cancer.

Our group also coordinated an autopsy evaluation of a woman who had been injected with radioactive Thorotrast (thorium-232) decades previously during angiography. Thorotrast, once used as a radiologic contrast agent, is not excreted from the body to any appreciable extent and has induced high rates of liver angiosarcoma and leukemia. I organized an international workshop that assembled the clinical and pathologic findings with dosimetric, radiochemical, autoradiographic, and molecular evaluations for this unique case. Our efforts enabled correlation of concentrations of radioactive-decay products in various organs with epidemiologic, histo-pathologic, and molecular observations.

We are now characterizing the risk of cancer after long-term exposure to radioactive Thorotrast among several large populations. This study may provide a special opportunity to evaluate the potential effects of low-level radon exposure. In our investigation, excesses of lung cancer has been noted among patients exposed to Thorotrast. Since thorium-232 decays into radon, which is continuously exhaled over the course of the patient's life , it is possible that this exposure caused the excess lung cancer. Detailed dosimetric studies are under way to quantify the dose of radon to pulmonary tissues and to relate dose to lung cancer risk. This study has public-health relevance because indoor radon is considered the single most important source of radiation exposure and risks of low-level exposure are poorly understood.

I am also interested in understanding the patterns and determinants of cancer risk following bone marrow transplantation, an increasingly common procedure used in the treatment of cancer, and in examining the possible interactions amoung immunosuppression, total-body radiotherapy, chemotherapy, viral cofactors, and graft-vs.-host disease. Our recent studies of more than 20,000 recipients of bone marrow transplants identified high rates of secondary lymphoma, and solid tumors are just now emerging as an important late consequence. With the Laboratory of Pathology at NCI, we are investigating the histologic and immunophenotypic characteristics of posttransplant lymphoproliferative disorders, Epstein-Barr virus status, and host-vs.-donor origin.

Whenever possible, my research in cancer epidemiology seeks to integrate new laboratory approaches in efforts to clarify mechanisms of carcinogenesis. Ultimately, one goal is to develop methods that might predict which cancer patients are at substantial risk for developing new malignancies and would thus benefit from targeted screening and preventive measures. Toward this end, we welcome further collaboration with clinical and laboratory colleagues at NIH.

Table of Contents