Sponsored by the U.S. Department of Energy Human Genome Program
Human Genome News Archive Edition
Human Genome News, September 1994; 6(3):8
The DOE Second International Workshop on the Development and Application of Biomarkers was held on April 26-28 in Santa Fe, New Mexico. Over 100 U.S. and foreign scientists, occupational physicians, radiation biologists, and government officials attended the meeting, which was supported by the DOE offices of Energy Research; Environment, Safety, and Health; and Environmental Management. The stimulus for this workshop was Public Law 102-484 (1992), which mandates that DOE survey its past and present workers who may have been exposed to chemicals or radiation.
New technologies developed by the Human Genome Project, as well as societal issues raised by such a large-scale health monitoring program, were among the issues discussed at the DOE workshop. Selected presentation highlights follow.
Detecting Exposure and Its Effects
Anthony Carrano (Lawrence Livermore National Laboratory) reviewed the study of human biomarkers, characterizing them as both exciting and frustrating. He commented that a variety of approaches have been explored for detecting biological effects of exposure to external agents; these include somatic cell cytogenetics, sperm damage, gene mutation endpoints, DNA adducts, studies of mechanisms and population, animal models, radiation, and chemical damage measurements.
Mortimer Mendelsohn (Radiation Effects Research Foundation, Japan) reviewed his extensive biomarker research into the effects of radiation on atomic bomb survivors, in whom some 10,000 cancers have been documented. Using chromosome aberration studies, Mendelsohn has shown only a very slight correlation of aberrations with cancers, an indication that the studied lymphocytes may have "forgotten" about the initial radiation exposure and that compensatory mechanisms work remarkably well. Mendelsohn noted that about half of all carcinogens are not mutagens and that validation of novel biomarkers is likely to be a serious stumbling block for research.
Charles Cantor (Boston University) said the Human Genome Project should be a rich source for new methods, discoveries of relevant human genes, identification of DNA regions particularly sensitive to various kinds of damage, integration of major databases, and spinoff technologies in unexpected areas. Advantages to using DNA as a biomarker include its uniqueness and relative stability, usefulness as a label for other molecules, and the ready detectibility of single molecules. Biomarkers could be used in such applications as surveys of organisms at toxic sites, unique identifier tags for released organisms in environmental modification or waste cleanup, and indicators of external damaging agents.
Paul Schulte (National Institute for Occupational Safety and Health) pointed out that markers of exposure may not be equivalent to markers of effect. In a major goal of occupational-disease prevention-reducing exposure-biomarkers would have no prominent role but could be useful in reducing the effects of exposure through medical monitoring. Schulte agreed that, before any medical screening program is started, ethical safeguards should be established. Also, relevant diseases should be significant and treatable, and medical tests should not be inordinately invasive, painful, or costly. He also felt that good consultation and counseling should be available, and tests should be targeted to specific risks. Test validity should be demonstrated first, both at the laboratory and population levels, and the underlying prevalence of the condition established. Although genetic screening is controversial, it could be the most powerful tool in conducting such tests.
Larry Clevenger (Sandia National Laboratory) noted that exposure to an environmental agent does not necessarily imply an effect. This is due to individual and immunological variation, different decay rates for exposure effects, and a very wide continuum for health and disease among people-all of which factors are important in discussions of genetic effects. Thus any correlations between biomarker results and future diseases may be only partial at best and include an irreducible chance component.
Assessing Disease Risk
Richard Albertini (University of Vermont Cancer Center) discussed three types of biomarkers for individual disease risk (see Biomarkers). Markers in these categories include somatic mutations in reporter genes, chromosomal aberrations, gpa variant frequencies, hprt mutations (mutational spectra in placental blood), and hprt mutational spectra for "specificity." Albertini stated that the major difficulty in using these as biomarkers for an individual's risk is that many current risk estimates are based on population studies, from which an average is determined.
Paul Brandt-Rauf (Columbia University) discussed the usefulness of p21, a product of the K-ras-2 oncogene, as a biomarker. A human version of p21 is located on human chromosome 12p11.1. Alterations or mutations in K-ras, which appears to be involved in a signal transduction pathway, could disrupt the pathway and lead to carcinogenesis. Some 80% of patients with liver angiosarcoma, associated with the known carcinogen vinyl chloride, have the aspartic acid-associated GAC codon, which is present in none of the controls. In one patient, semiquantitation of serum p21 served as a crude biomarker, with some predictive value, until the patient died.
James Jett (Los Alamos National Laboratory) characterized flow cytometry (FCM) as rapid, capable of handling large numbers of samples, highly flexible, and very sensitive when compared to conventional microscopy. FCM can measure fluorescence; light-scattered, cell, or particle volume; and other optical properties. FCM probes exist for various targets including DNA, RNA, proteins, [Ca], pH, membrane fluidity, and surface and intracellular antibodies. Up to 32 measurements/cell can be realized at a flow rate of 100,000 cells/s, and various analysis levels and a number of applications could be useful for biomarker measurements. FCM's extensive capabilities could also include probes with wider fluorescent emissions and even FCM on a chip.
In reviewing molecular cytogenetics and biomarker development, Joe Gray (University of California, San Francisco) stated that an important approach is fluorescent in situ hybridization, called chromosome painting when used on entire chromosomes. With this technology, 48,000 to 50,000 metaphases can be scored each day, allowing chromosomal translocations to be directly visualized and counted. The greater the radiation dose, the fewer metaphases are required for detection of abnormalities. Fibroblasts are a convenient and successful target for studies to assess external agent damage. One approach in finding early lesions for cancer cell detection is whole genome subtraction, in which tumor DNA is "subtracted" from whole genomic DNA, identifying differences that may be associated with the tumor phenotype. DNA changes can be mapped to within 10 Mb. An expected outcome of the Human Genome Project is the identification of chromosomal changes associated with some cancers.
Chronic Beryllium Disease
A number of speakers discussed the health impact of beryllium exposure, which in susceptible individuals can lead to an autoimmune-like pulmonary disease. Beryllium is used industrially in the manufacture of light fixtures and certain ceramics. Milton Rossman (University of Pennsylvania) reviewed the immunology of chronic beryllium disease (CBD), first described in 1946 as a "delayed chemical pneumonitis." CBD also causes granulomas in the skin, liver, lymph nodes, and conjunctiva. Steroids are effective if CBD is diagnosed early, but if treatment is delayed, collagen deposits lead to scarring.
Cesare Saltini (University of Modena, Italy) reviewed his research showing that CBD appears to be mediated by beryllium-specific C4 positive T-cells, the same T-cells affected by the HIV virus in AIDS patients. The population frequency of the genetic marker associated with CBD susceptibility is about 30%, while in CDB patients this frequency is nearly 100%. However, CBD incidence in beryllium-exposed populations ranges from 0.4 to 4.9%. This pattern is typical of many genetically influenced (particularly autoimmune) diseases in that a large proportion of CBD patients have a given marker but few with the marker have the disease.
Lee Newman (National Jewish Center for Immunology and Respiratory Medicine, Denver) described CDB detection through the lymphocyte transformation test (LTT). In the presence of beryllium, lymphocytes from sensitized individuals can transform and incorporate an isotope, allowing the degree of sensitization to be measured. However, some people with clearly abnormal LTT blood results have no lung disease. Because sensitization is thought to precede disease, LTT might assist in accurate and early diagnosis.
A number of important issues emerged from this workshop. Researchers felt that a considerable gap exists between the demand for detailed and comprehensive data about medical implications of workplace hazards and the current ability to compile relevant biomarker-based information. Existing biomarkers are far from ideal, and their actual significance remains to be worked out. To make biomarkers more useful, attendees saw the urgent need for better population measurements; data on dose reconstruction and low doses, particularly the shape of dose-response curves; and studies of at least 755 little-known industrial chemicals.
More information is also needed on the distribution of risks in individuals other than the "average" working male and on what constitutes an "acceptable" risk. How are the risk-benefit analyses to be accomplished? What sort of communication and education campaign would support appropriate and reasonable research?
Many speakers recognized that ethical, legal, and social concerns are important in accomplishing any research agenda and that these sensitivities should be incorporated at the outset. Although new biomarkers offer many opportunities for increased accuracy, sensitivity, and predictability, the ability to measure will outrun the understanding of medical and health implications. Privacy of biomarker information is a general concern related to more-general issues of privacy of medical records and controls on their access.
Daniel Drell, DOE OHER
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Human Genome Program, U.S. Department of Energy, Human Genome News (v6n3).
The Human Genome Project (HGP) was an international 13-year effort, 1990 to 2003. Primary goals were to discover the complete set of human genes and make them accessible for further biological study, and determine the complete sequence of DNA bases in the human genome. See Timeline for more HGP history.
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