Program of the 2002 Annual Meeting, Where the New Biology Meets Epidemiology: Impact on Radiation Risk Estimates
April 10-11, 2002
The New Biology
R.J. Michael Fry, Session Chair
The State of the Art in the 1990s: NCRP Report No. 136 on the Scientific Bases for Linearity
Arthur C. Upton, Rutgers University
To reassess the use of the linear-nonthreshold (LNT) dose-response model in the light of advancing knowledge, the NCRP charged Scientific Committee 1-6 to evaluate the weight of scientific evidence for and against the LNT model, without reference to any associated policy ramifications. To accomplish the task, the Committee reviewed the relevant theoretical, experimental and epidemiological data on those effects of ionizing radiation which are generally postulated to be stochastic in nature (i.e., genetic and carcinogenic effects). From its review of the data, the Committee concluded that the weight of evidence suggests that lesions which are precursors to cancer (i.e., mutations and chromosome aberrations), and certain types of cancer as well, may increase in frequency linearly with the dose in the low-dose domain. On this basis, the Committee concluded that no alternative dose-response model is more plausible than the LNT model but that other dose-response relationships cannot be excluded, especially in view of growing evidence that the dose-response relationship may be modified by adaptive responses, bystander effects, and other variables.
Radiation-Induced Genomic Instability In Vitro: Implications for Radiation Risk
William F. Morgan, University of Maryland
The short-term effects of cellular exposure to ionizing radiation are relatively well understood. The DNA lesions induced, and cellular responses to radiation including DNA repair, cell-cycle regulation, gene expression changes and even cell death have been the subject of intense investigation for many years. What is less clear however, are the long-term consequences of exposure to low dose/low dose rate radiation and the effects of radiation exposure on the progeny of surviving cells. This presentation will focus on the genomic effects observed in surviving cells multiple generations after the initial exposure to radiation. The genetic changes observed include delayed mutation, delayed reproductive cell death, delayed transformation and chromosomal instability and are grouped under the umbrella of radiation-induced genomic instability. Chromosomal changes are the best characterized delayed effect of radiation exposure and evidence for a recombination based process involving genomic rearrangements will be presented. We have exploited this recombination based process to develop a plasmid vector based assay for detecting recombination events at low frequencies, like those expected after low dose/low dose rate radiation. The frequency of radiation-induced genomic instability after acute exposures to high doses of radiation is extremely high indicating that induced instability is unlikely to be the result of a gene mutation. Instead this high frequency suggests changes in gene expression and epigenetic “bystander” effects could perpetuate the observed instability. Evidence for changes in gene expression and nontargeted bystander effects will also be presented. Finally we will attempt to relate those genomic changes occurring in cells that survive irradiation, with current concepts of radiation risk.
Genomic Instability, Susceptibility Genes, and Carcinogenesis
Robert L. Ullrich, Colorado State University
While it is generally accepted that the carcinogenic effects of radiation are somehow related to its clastogenic and mutagenic effects, details of the underlying mechanisms are not known. One model identifies specific radiationinduced gene or chromosomal mutations as the critical initial event while an alternative model suggests radiationinduced genomic instability as the earliest cellular event in the multi-step process of radiation carcinogenesis. From the genomic instability model it is predicted that this initial instability would put virtually all genes at a higher risk for mutagenesis but its major impact for carcinogenesis occurs when mutations arise involving certain critical genes. Considerable insight into mechanisms is now starting to emerge from genetic studies in humans and in animals. That individuals have differing heritable susceptibilities to cancer has been known for many years. During the last several years, molecular studies have identified a number of the genes that confer increased susceptibility. Because a number of these genes (e.g., ATM and NBS) appear to be involved in DNA damage response pathways of importance in the processing of damage of the type induced by ionizing radiation, it seems reasonable that individual differences in susceptibility to radiation-induced cancer also exist. Studies in humans have demonstrated the heritability of sensitivity to certain kinds of radiation-induced chromosome damage in cancer prone families. Recent studies with mouse models of radiation carcinogenesis provide, for the first time, evidence of associations between germ line polymorphic loci, post-irradiation chromosomal instability and tumor development. The association between susceptibility and specific genes and genes pathways also provide clues to underlying mechanisms. The current state of understanding of the link between radiation induced genomic instability, genetic susceptibility, and radiation-induced cancer will be presented.
The Bystander Effect
Eric J. Hall, Columbia University
The “bystander effect” refers to the induction of biological effects in cells that are not directly traversed by a charged particle, but are in close proximity to cells that are.
As early as the 1940s there were reports that the inactivation of biological entities may be brought about equally by ionizations produced within the entity or by the ionization of the surrounding medium. By 1947, Kotral and Gray showed that alpha particles which pass close to the chromatid thread, as well as those which pass through it, have a significant probability of producing chromatid and isochromatid breaks or chromatid exchanges.
In modern times, the bystander effect came to light when it was observed that, if cells are exposed to a low dose of alpha particles, more cells showed a biological effect than were “hit” by an alpha particle. The most elegant bystander studies have involved the use of single particle microbeams, which allow specific cells to be irradiated, and effects studied in their neighbors. An alternative strategy is to transfer medium from irradiated cells, and study effects in cells receiving the medium.
Endpoints for which a bystander effect has been unequivocally demonstrated include sister chromatid exchanges, micronuclei, cell lethality, mutation, oncogenic transformation and gene expression.
It is already clear that there are two bystander effects that may operate separately or in combination. One involves communication via gap junctions when cells are in close contact. This is by far the larger effect. The other involves the release into the medium of some factor that then affects distant cells. In neither case has the effecting molecule been identified.
The existence of a substantial bystander effect casts some doubt on the validity of a linear extrapolation of biological effects from high to low doses, or at least casts some doubt on the dose level below which linearity can be assumed since the target size is larger than the cell.
Monitoring Human Radiation Exposure by Gene Expression Profiling: Possibilities and Pitfalls
Sally A. Amundson, National Institutes of Health
Advances in high throughput analysis of mRNA expression have made it possible to establish gene expression profiles for different cells, tissues, diseases, and exposure states. For instance, recent studies have demonstrated the utility of such an approach to classify subtypes of cancer with more detail than was previously possible. In addition, gene expression studies of ionizing radiation exposure both in vitro and in vivo are affording insight into the molecular mechanisms of mammalian radiation response. We have demonstrated that radiation expression profiles are a good predictor of p53 function in cell lines, and such profiles also indicate a major role of p53-regulated genes in the in vivo radiation response. Gene expression can be a sensitive indicator of radiation response, as we have shown linear dose responses for induction of several genes as low as 2 cGy. As profiles are established from radiation studies, it is hoped that they may be useful for identifying individuals with specific exposures or predisposition to negative outcome of exposure. Although this technology holds great promise, some obstacles remain to be overcome before it can be successfully applied to population studies.
The Epidemiology
S. James Adelstein, Session Chair
Dose Response and Temporal Patterns of Radiation-Induced Cancer Risks
Dale L. Preston, Radiation Effects Research Foundation
Findings of the Life Span Study (LSS) cohort of atomic-bomb survivors are a primary source for quantitative risk estimates that underlie radiation protection. Because of the size and length of follow-up of LSS provides considerable information on both the nature of the dose response and on how the radiation-associated excess risks vary with age, age at exposure, gender, and other factors. Our current analyses extend the mortality follow-up by 7 y (through 1997) and add By (through 1995) to the incidence follow-up. During the follow-up periods there have been a total of about 9,300 solid cancer deaths and almost 12,200 incident cases. As outlined in this presentation, while discussing issues related to the shape of the dose response and low dose risks in some detail, the new reports consider temporal patterns in greater detail than has been done previously. As we have reported, the LSS solid cancer dose response is well described by simple linear dose response over the 0 to 2 Sv range (with some leveling off at higher estimated doses). This remains the case with the extended follow-up. Although LSS is often referred to as a high dose study, about 75 percent of the 50,000 cohort members with doses in excess of 5 mSv have dose estimates in a range of direct interest for radiation protection (0 to 200 mSv). Analyses of data limited to this low dose range provide direct evidence of a significant solid cancer dose response with a risk per unit dose that consistent with that seen for the full dose range. Previous LSS reports have focused on descriptions of the solid cancer excess risks in which the excess relative risk varies with age at exposure and gender. In addition to the age at exposure effects, our current analyses suggest excess relative risks also vary with age (at death or diagnosis). Excess relative risks are higher for those exposed earlier in life, with attained age-specific risks changing by about 30 percent per decade, but tend to decrease with increasing attained age, roughly in proportion to 1/attained-age, for any age at exposure. Despite the decreasing relative risk, excess rates have increased rapidly throughout the study period with some indication, especially for the incidence data, that attained-age-specific rates are higher for those exposed at younger ages. Simple comparisons of site-specific excess risks are used to illustrate how the interpretation of age-at-exposure effects on excess relative risks or excess rates is complicated by changes in baseline rates with birth cohort or time period.
Cancer Risks from Medical Radiation
Elaine Ron, National Cancer Institute
About 15 percent of the ionizing radiation exposure to the general public comes from artificial sources, and almost all of this exposure is due to medical radiation, largely from diagnostic procedures. Of the approximately 3 mSv annual per caput effective dose estimated for the year 2000, 2.4 mSv is from natural background and 0.4 mSv from diagnostic medical exams. Diagnostic and therapeutic radiation was used in patients as early as 1896. Since then, continual improvements in diagnostic imaging and radiotherapy, as well as the aging of our population have led to greater use of medical radiation. Temporal trends indicate that worldwide population exposure from medical radiation is increasing. In the United States, there has been a steady rise in the use of diagnostic radiologic procedures, especially x rays. Radiotherapy also has increased so that today about 40 percent of cancer patients receive some treatment with radiation. Epidemiologic data on medically irradiated populations are an important complement to the atomic-bomb survivors’ studies.
Significant improvement in cancer treatment over the last few decades has resulted in longer survival and a growing number of radiation-related second cancers. Following high-dose radiotherapy for malignant diseases, elevated risks of a variety of radiation-related second cancers have been observed. Risks have been particularly high following treatment for childhood cancer. Radiation treatment for benign disease was relatively common from the 1940s to the 1 960s. While these treatments generally were effective, some resulted in enhanced cancer risks. As more was learned about radiation-associated cancer risks and new treatments became available, the use of radiotherapy for benign disease has declined. At moderate doses, such as those used to treat benign diseases, radiation-related cancers occur in or near the radiation field. Cancers of the thyroid, salivary gland, central nervous system, skin, and breast, as well as leukemia have been associated with radiotherapy for tinea capitis, enlarged tonsils or thymus gland, other benign conditions of the head and neck, or benign breast diseases. Because doses from diagnostic examinations typically are low, they are difficult to study using epidemiologic methods, unless multiple examinations are performed. An excess risk of breast cancer has been reported among women with tuberculosis who had multiple chest fluoroscopies, as well as among scoliosis patients who had frequent diagnostic x rays during late childhood and adolescence. Dental and medical diagnostic x rays performed many years ago, when doses were presumed to be high, also have been linked to increased cancer risks. The carcinogenic effects of diagnostic and therapeutic radionuclides are less well characterized. High risks of liver cancer and leukemia have been demonstrated following thorotrast injections, and patients treated with radium appear to have an elevated risk of bone sarcomas and possibly cancers of the breast, liver, kidney, thyroid and bladder.
Diagnostic X Rays, DNA Repair Genes, and Childhool Leukemia
Claire Infante-Rivard, McGill University, Montreal
The risk of childhood cancer associated with postnatal diagnostic irradiation has not been studied much. It is perhaps assumed that the radiation doses from common x rays are not such that they would increase the risk of cancer in children. However, a substantial number of children seem to have many x rays and on the other hand, from the published literature, doses in clinical settings seem to vary considerably. In the course of a case-control study of childhood acute lymphoblastic leukemia (ALL) we collected information on postnatal x rays. We also collected genetic material from a subgroup of cases and determined the presence of genetic polymorphisms in DNA repair genes, hypothesizing that these could modify susceptibility. We genotyped children for hMLH1 (exon 8), hMSH3 (exon 21), hMSH3 (exon 23), and XRCC1 (exon 6). We estimated the risk of postnatal irradiation on ALL and explored the interaction effects between exposure and genetic polymorphisms. We have shown that there is a moderate increase in ALL risk with number of reported x rays, and particularly among girls (odds ratio = 2.26; 95 percent confidence interval = 1.20 —4.23 for two x rays or more in comparison with none). We have also reported preliminary data suggesting that the studied polymorphisms either confer protection or potentialize risk. We are in the process of completing another phase of our epidemiologic study of ALL with additional cases and controls; in addition, more cases have been genotyped. I will report on risk associated with postnatal irradiation and on the potentially modifying effect of genetic polymorphisms in DNA repair genes from this expanded database. I will also address some issues related to the design of epidemiologic studies for complex diseases when evaluating genetic factors.
Genetic Effects of Radiotherapy for Childhood Cancer
John D. Boice, Jr., International Epidemilogy Institute
No radiation-induced genetic diseases have so far been demonstrated in humans and estimates of genetic risks have to be based on mouse experiments. The most comprehensive human studies have been of the Japanese survivors of the 1945 atomic bombings which show little evidence for inherited defects attributable to radiation. Studies of workers exposed to occupational radiation may not be sufficiently large to result in a detectable or interpretable genetic effect and maternal factors may not be available to adjust for in the analyses. Thus, there are theoretical and practical advantages of addressing genetic effects in persons exposed to higher dose medical radiation, such as survivors of cancers of childhood, adolescents, and young adulthood.
Cancer survivors have detailed medical records that permit accurate estimates of gonadal doses and the assessment of potentially confounding variables, such as intercurrent illness, personal and family medical histories, lifestyle characteristics such as tobacco and drug use, as well as circumstances at delivery. To address potential genetic effects in this population, an international study is nearing completion of over 25,000 survivors of childhood cancer in the United States and Denmark who gave birth or fathered over 6,000 children. Dose to gonads is being reconstructed from the radiotherapy records indicating to date that 46 percent received over 100 mSv and 11 percent over 1,000 mSv. Pregnancy outcomes being evaluated include congenital malformations, stillbirths, neonatal deaths, total deaths, leukemia and childhood cancer incidence, and altered sex ratio. Siblings constitute one comparison group. The main analyses will be internal based on dose-response evaluations. Blood studies of trios (cancer survivor, spouse and offspring) have been initiated to evaluate evidence for the transmission of any genetic damage that could be detected in blood cells, such as minisatellite mutations.
In the United States series to date, 4,214 survivors reported that 157 of their children had a genetic disease (3.7 percent) in contrast to 95 (4.1 percent) among 2,339 sibling controls. Somewhat similar observations are seen in the initial analyses of the Danish series. Coupled with prior studies, these preliminary findings, if sustained by ongoing dose-response analyses, provide reassurance that cancer treatments including radiotherapy do not carry a large risk for genetic disease in offspring conceived after exposure. Survivors of childhood cancer form the largest group of people exposed to high doses of ionizing radiation before reproduction. Thus, the study results will provide another source of human data on the possible level of heritable genetic effects following radiation exposure.
Tracking the Errant Cell After the Atomic Bombings: What Went Wrong
Kei Iwamoto, University of California
Epidemiological data collected after the atomic-bomb blasts of Hiroshima and Nagasaki have established a link between radiation exposure and human cancer development and are the major source of information for current radiation-induced cancer risk assessment. To determine the mechanistic basis for radiation carcinogenesis, retrospective molecular analyses of archival hepatocellular carcinoma (HCC) tissues from the atomic-bomb survivors were conducted. The tumor suppressor genesps3 and M6P//GF2r were examined. HCC cases had either p53 mutations of M6P//GF2r mutations, but rarely both. Moreover, the frequency of cases with M6P//GF2r mutations actually decreased with dose, while those for p53 increased. This implies two independent selection processes leading to liver cancer and that in radiation-induced HCC tumors the spectrum of molecular changes is different from that in “background” tumors.
Twenty-Sixth Lauriston S. Taylor Lecture on Radiation Protection and Measurements
Introduction of the Lecturer, R.J. Michael Fry
Developing Mechanistic Data for Incorporation into Cancer Risk Assessment: Old Problems and New Approaches, R. Julian Preston
Where Do We Go From Here?
Marco A Zaider, Session Chair
How Radiation-Induced Phenotypes Contribute to Neoplastic Progression
Mary Helen Barcellos-Hoff, Lawrence Berkeley National Laboratory
Ionizing radiation elicits both rapid and persistent changes in extracellular signaling, which can affect cell behavior, communication and multicellularorganization. Some of these effects are exhibited by daughters of irradiated cells, suggesting heritable alterations in signaling, communication and adhesion. We postulate that such phenotypes contribute to the carcinogenic potential of radiation in that aberrant extracellular signaling might predispose cells to genomic instability and neoplastic progression.
Developing a Scientific Basis for Radiation Risk Estimates: The Goal of the DOE Low Dose Research Program
Antone L. Brooks, Washington State University-Tricities
The U.S. Department of Energy’s Low Dose Radiation Research Program is a 10 y activity currently funded at $21 million per year. It focuses on biological responses to low doses (<0.1 Gy) of low-LET ionizing radiation. The overall goal of this program is to provide a sound scientific basis for the radiation protection standards. The program supports basic research that combines modern genomic, molecular and cellular techniques with recent advances in scientific instrumentation. These combinations make it possible to detect responses and test paradigms associated with the mechanisms of low dose radiation action not previously measurable or testable. Research to date is briefly reviewed and suggests the need for some major paradigm shifts. Exposure of the extra-cellular matrix can modify both the pattern of gene expression and the phenotype of the cells which result in cell transformation without direct mutation. Low dose radiation exposure results in a range of dose-response relationships for changes in the number, types and patterns of gene expression. Such studies suggest an increased role for gene expression relative to single mutations for radiation induced cancer. Low dose research using microbeams demonstrated that cells do not require a direct “hit” to result in significant biological alterations. These “bystander effects” demonstrate that “non-hit” cells respond with changes in gene expression, DNA repair, chromosome aberrations, mutations, and cell killing. Research to link genomic instability with cancer is also being conducted and will be discussed. Detection of radiosensitivity genes as markers of genetic susceptibility in individuals and populations and can be used in epidemiological studies to determine how molecular changes may impact risk. It is not possible to determine how this research will influence current radiation standards. However, the Low Dose Research Program will help ensure that radiation standards are set using the best scientific data available, and that they are adequate and appropriate for the protection of workers and the public.
The Impact of the New Biology on Radiation Risks in Space
John F. Dicello, Johns Hopkins Oncology Center
Radiation is considered to be one of three or four major hazards for personnel in space and has emerged as the most critical issue to be resolved for long-term missions, both orbital and interplanetary. Space habitats are stressful and dangerous environments. Health and medical consequences arising from microgravity, stress, and trauma include weakened immune systems, increased viral activity, and loss of bone mass. The greatest risks from radiation are generally assumed to be cancers and possibly damage to the central nervous system. Synergistic effects arising from the other environmental hazards along with abscopal and exogenic factors are likely.
Space programs represent an exceptional opportunity for examining the biological consequences of low-dose exposures of humans to radiation at every level of progression. Although astronauts are a relatively small population, they are healthy, physically active volunteers who undergo extensive testing and medical examinations before, during, and after protracted exposures with periodic follow-up examinations. The radiation environments along with other hazards are likewise monitored and documented. Extensive international research programs are in progress, including those at the Brookhaven National Laboratory (BNL) in the United States, and the National Institute of Radiological Sciences in Chiba, Japan. The NASA Specialized Center for Research and Training for space radiation health has been in existence for almost a decade. About 5 y ago, the National Space Biomedical Research Institute began through a cooperative agreement with the U. S. National Aeronautics and Space Administration in order to address radiation issues through a concerted, programmatic effort. A comprehensive Booster Applications Facility, dedicated to studying effects produced by energetic ions, is nearing completion at BNL.
Advanced technologies are rapidly being incorporated into these programs to determine the significance of new biological data and to evaluate the interplay among the different medical hazards. Programmatic in vivo and in vitro studies of the processes leading to carcinogenesis are in progress. Drugs and dietary supplements are being examined at the cellular and in vivo levels to assess their potential as dose-modifying agents. This presentation will review the major medical hazards in space, new programs, recent accomplishments, and future directions.
Do Low Dose-Rate Bystander Effects Influence Domestic Radon Risks?
David J. Brenner, Columbia University
Radon risks derive from exposure of bronchio-epithelial cells to high-LET alpha particles. Alpha-particle exposure can result in bystander effects, where irradiated cells emit signals resulting in damage to nearby unirradiated bystander cells. This can result in nonlinear dose-response relations, and inverse dose-rate effects. Domestic radon risk estimates are currently extrapolated from miner data which are at both higher doses and higher dose rates, so bystander effects on unhit cells could play a large role in the extrapolation of risks from mines to homes. We therefore discuss a quantitative mechanistic model of bystander effects to include protracted exposure, with the aim of quantifying the significance of the bystander effect for very prolonged exposures.
A model of high-LET bystander effects developed to analyze oncogenic transformation is extended to low dose rates. The model considers radiation response as a superposition of bystander and linear direct effects. It attributes bystander effects to a small subpopulation of hypersensitive cells, with the bystander contribution dominating the direct contribution at very low acute doses but saturating as the dose increases. Inverse dose-rate effects are attributed to replenishment of the hypersensitive subpopulation during prolonged irradiation.
Parameter estimates based on applying the model to dose and dose-rate dependent miner data suggest that one directly-hit target bronchio-epithelial cell can send bystander signals to about 50 neighboring target cells. The model suggests that a naïve linear extrapolation of radon miner data to low doses, without accounting for exposure time, would result in an underestimation of domestic radon risks by about a factor of four, a value comparable to the empirical estimate applied in the recent BEIR VI report on radon risk estimation.
Bystander effects represent a plausible quantitative and mechanistic explanation of inverse dose-rate effects by high-LET radiation, resulting in nonlinear dose-response relations, and a complex interplay between dose and exposure time. The model discussed here provides a potential mechanistic underpinning for the empirical exposure-time correction factors applied in the recent BEIR VI report.
The Program Committee
Eric J. Hall,Chair
Antone L. Brooks
John F. Dicello
Dale L. Preston
R. Julian Preston
Elaine Ron
Robert L. Ullrich
Marco A. Zaider
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