Mutation Research/Reviews in Genetic Toxicology最新文献

Data supporting the use of transgenic lines to identify carcinogens and noncarcinogens are thus far based on a limited number of chemicals for which there are also long-term bioassay results in rats and/or mice. Six chemicals have been tested in the heterozygous p53-deficient mice and 13 in the Tg · AC line. The results show that the p53^def responds rapidly to mutagenic carcinogens and the Tg · AC responds rapidly to both mutagenic and nonmutagenic carcinogens. Neither transgenic line responded to the noncarcinogens that were tested. The p53^def line failed to respond to two nonmutagenic carcinogens (N-methyloacrylamide and reserpine), the Tg · AC line failed to respond to ethyl acrylate, a nonmutagenic chemical that induced tumors of the forestomach when administered by gavage, and to triethanolamine that caused an increase in hepatocellular tumors in B6C3F1 mice via skin painting. Both of the latter chemicals are examples of highly specific responses related to either route of administration or to strain susceptibility. Further efforts to evaluate the range of chemicals to which these transgenic lines respond are currently in progress.

Recent years have seen the rapid expansion of scientific understanding of the underlying biologic bases of toxic reactions to chemicals. Use of this information in health risk assessment is expanding, but it has yet to reach its full potential. This article considers what has successfully been done, what approaches are now being developed, and what impediments and difficulties have been encountered in attempts to bring case-specific, mechanistic toxicological information to bear on risk estimation. In hazard identification, mechanistic information can help explain the bearing of various empirical experimental results for inferring human hazard, can increase the sensitivity of detection, and can be considered in attempts to replace 2-year animal bioassays with hazard identification methods that rest on identifying key biological properties underlying carcinogenicity rather than relying only on the experimental observation of tumors. In carcinogen potency estimation, mechanistic information can potentially extend relevant observation to lower dose levels, provide the basis for choosing among empirically based dose-response models, lead to potency estimates through relationships with quantitative measures of short-term test outcomes, and can be considered as a basis for providing direct observation of the biological parameters in biologically based dose-response modeling.

DNA replication is not an error-free process; therefore induction of cell proliferation with the requisite increase in DNA replication may be an important mechanism by which carcinogenesis can be induced by chemicals.

Data presented in this overview indicate a positive association between increased cell proliferation and carcinogenesis, and illustrate the value of performing mechanistic studies such as cell proliferation assays in conjunction with short-term tests to further investigate the results of cancer bioassays. Whereas chemically-induced cell proliferation per se may not be sufficient to induce carcinogenesis, it creates a favorable environment for tumor development. There are two types of chemically-induced cell proliferation, mitogenic and cytotoxic, and they have different consequences regarding the mechanism of carcinogenesis of a chemical. Mitogenic chemical such as phenobarbital, oxazepam, and the peroxisome proliferating agents exert a short-term cell proliferative response that may exert its primary effect in carcinogenesis at the promotion stages. It is not clear at what stage(s) cytotoxic agents such as methapyrilene, α_2u-globulin inducers or saccharin exert their effects in carcinogenesis. A confounding factor in evaluation of cell proliferation in risk assessments is the production of chemical specific pleiotropic effects that may contribute to the carcinogenicity of a chemical. It is clear that mechanistic studies performed to understand the relationship of sex, species and dose in rodent carcinogenicity assays of chemicals is critical for the extrapolation of such data for human health assessments.

This paper describes the four cytogenetic endpoints most frequently used in hazard identification assays as the first step in the risk assessment process. These are structural chromosome aberrations, micronuclei, aneuploidy, and sister chromatid exchanges. The biological mechanisms involved in the formation of the alterations observed in each assay are briefly discussed. Variations in and recent improvements to each assay are described, with an emphasis on the use of molecular techniques to improve the sensitivity of the assay, and to allow for detection of specific alterations that are, or could be, associated with cancer induction. This, in turn, will make the data obtained in the cytogenetic assays more useful in cancer and genetic risk assessment. Thus, the aim of this paper is to encourage cytogeneticists to design their experiments in such a way that the data obtained will be of maximum possible benefit for characterizing and quantifying adverse human health effects, particularly cancer.

Short-term genetic toxicology tests were developed for the purpose of identifying chemical carcinogens in the environment. After two decades of development and validation, the tests are well-established in routine testing schemes, but our views of their utility for safety evaluation have undergone re-assessment. The correlation between identified mutagens and identified carcinogens has turned out to be significantly less than one. Processes or mechanisms that are not directly genotoxic appear to play a role in carcinogenesis. While short term test data are still components of the assessment of carcinogenic risk, genetic damage also has been recognized as important in its own right, in relation to heritable genetic risk and other health-related effects, such as aging, reproductive failure and developmental toxicity. The revolution in molecular biology and genetic analysis occurring over the past 20 years has contributed to the wealth of new information on the complexities of cell regulation, differentiation, and the carcinogenic process. These technologies have provided new experimental approaches to genetic toxicology assessments, including transgenic cell and animal models, human monitoring, and analysis of macromolecular interactions at environmentally relevant exposures. The potential exists for the development of more efficient and more relevant genetic toxicology testing schemes for use assessing human safety. A delineation of contemporary needs, a modern view of the elements of cancer induction, and an examination of new assays and technologies may provide a framework for integrating new approaches into current schemes for evaluating the potential genetic and carcinogenic risk of environmental chemicals.

An extremely large database describes genotypes associated with the human cancer phenotype and genotypes of human populations with genetic predisposition to cancer. Aspects of this database are examined from the perspective of risk analysis, and the following conclusions and hypotheses are proposed: (1) The genotypes of human cancer cells are characterized by multiple mutated genes. Each type of cancer is characterized by a set of mutated genes, a subset from a total of more than 80 genes, that varies between tissue types and between different tumors from the same tissue. No single cancer-associated gene nor carcinogenic pathway appears suitable as an overall indicator whose induction serves as a quantitative marker for risk analysis. (2) Genetic defects that predispose human populations to cancer are numerous and diverse, and provide a model for associating cancer rates with induced genetic changes. As these syndromes contribute significantly to the overall cancer rate, risk analysis should include an estimation of the effect of putative carcinogens on individuals with genetic predisposition. (3) Gene activation and inactivation events are observed in the cancer genotype at different frequencies, and the potency of carcinogens to induce these events varies significantly. There is a paradox between the observed frequency for induction of single mutational events in test systems and the frequency of multiple events in a single cancer cell, suggesting events are not independent. Quantitative prediction of cancer risk will depend on identifying rate-limiting events in carcinogenesis. Hyperproliferation and hypermutation may be such events. (4) Four sets of data suggest that hypermutation may be an important carcinogenic process. Current mechanisms of risk analysis do not properly evaluate the potency of putative carcinogens to induce the hypermutable state or to increase mutation in hypermutable cells. (5) High-dose exposure to carcinogens in model systems changes patterns of gene expression and may induce protective effects through delay in cell progression and other processes that affect mutagenesis and toxicity. Paradigms in risk analysis that require extrapolation over wide ranges of exposure levels may be flawed mechanistically and may underestimate carcinogenic effects of test agents at environmental levels. Characteristics of the human cancer genotype suggest that approaches to risk analysis must be broadened to consider the multiplicity of carcinogenic pathways and the relative roles of hyperproliferation and hypermutation. Further, estimation of risk to general human populations must consider effects on hypersusceptible individuals. The extrapolation of effects over wide exposure levels is an imprecise process.

DNA adducts have been investigated extensively during the past decade. This research has been advanced, in part, by the development of ultrasensitive analytical methods, such as ³²P-postlabeling and mass spectrometry, that enable detection of DNA adducts at concentrations as low as one adduct per 10⁹ to 10¹⁰ normal nucleotides. Studies of mutations in activated oncogenes such as ras, inactivated tumor suppressor genes such as p53, and surrogate genes such as hprt provide linkage between DNA adducts and carcinogenesis. The measurement of DNA adducts, or molecular dosimetry, has important applications for cancer risk assessment. Cancer risk assessment currently involves estimating the probable effects of carcinogens in humans based on results of animal bioassays. Estimates of risk are then derived from mathematical models that fit data of tumor incidence at the high animal exposures and extrapolate to probable human exposures that may be orders of magnitude lower. Molecular dosimetry could extend the observable range of mechanistic data several orders of magnitude lower than can be achieved in carcinogenesis bioassays. This measurement also compensates automatically for individual and species differences in toxicokinetic factors, as well as any nonlinearities that affect the quantitative relationships between exposure and molecular dose. As a result, molecular dosimetry can provide a basis for conducting high- to low-dose, route-to-route, and interspecies extrapolations. The incorporation of such data into risk assessment promises to reduce uncertainties and produce more accurate estimates of risk compared to current methods.

The ability to quantify somatic mutations in vivo provides a new source of toxicological information that is relevant to the assessment of cancer risk. The major experimental factors that influence the mutant frequency are age, time after treatment, treatment protocol, and tissue analyzed. In untreated mice, the mutant frequency increases very rapidly with age from conception to birth, more slowly from birth to adulthood, and very slowly thereafter. All somatic tissues studied so far in adults have similar mutant frequencies. The time after treatment (expression time) is the most important experimental variable. The minimum time for expression varies from one tissue to another. To be valid, comparisons between tissues and treatments must be made after complete expression of the mutations. Unfortunately, the minimum expression time has not been characterized in most tissues. Since carcinogens are tissue specific, and many chemicals are distributed in the body in complex patterns, it is to be expected that there will be differences in the frequency of mutation induced in different tissues. As yet this has not been extensively studied. Since the mutations detected by the transgenic assays are neutral, the mutants should accumulate as the integral of the mutation rate. Hence chronic treatment protocols should be more effective than acute and subacute protocols whenever they permit substantially larger doses to be delivered. Such protocols are more relevant to human exposure and are preferable for dose extrapolations. The importance of transcription in determining mutation rates is not yet known, but it is noteworthy that the transgenes are not transcribed whereas the loci involved in carcinogenesis are. The mutation spectrum is important for quantitative risk estimation. Risk estimation must also take into account the spectrum of mutations that are involved in the carcinogenic process in the tissue and the spectrum of mutations that are detectable by the assay. New assays are being used to quantify mutations in vivo in order to understand the carcinogenic process, to search for the environmental factors involved in human cancer, and to evaluate the carcinogenic hazard qualitatively.

Genotoxicity test batteries have become a standard tool for identifying chemicals that may have potential carcinogenic risk to humans. It is now apparent, however, that the use of genotoxicity batteries for assessing carcinogenic potential has limitations including an overall low specificity and a limited ability to detect carcinogens acting via ‘nongenotoxic’ mechanisms. In vitro cell transformation models, because they measure a chemical's ability to induce preneoplastic or neoplastic endpoints regardless of mechanism, may fulfil the current need for an in vitro biologically relevant model with increased predictiveness for determining carcinogenic potential. This review will focus on data demonstrating the similarities of chemically induced cell transformation in vitro to carcinogenesis in vivo. Furthermore, a growing database demonstrating a high overall correlation between cell transformation results with those of the rodent bioassay will also be discussed. Finally, the inclusion of cell transformation approaches for assessing the carcinogenic potential of chemicals relative to currently used genotoxicity batteries will be presented.