Begell House Inc.
Journal of Environmental Pathology, Toxicology and Oncology
JEP(T)
0731-8898
20
4
2001
Molecular Epidemiology of Human Cancer: Biomarkers of Genotoxic Exposure and Susceptibility
17
10.1615/JEnvironPatholToxicolOncol.v20.i4.10
Stephen G.
Grant
Department of Environmental and Occupational Health, University of Pittsburgh, the University of Pittsburgh Cancer Institute and Magee-Womens Research Institute, Pittsburgh, PA
The new field of molecular epidemiology investigates the link between toxic exposure and an associated health effect by defining presumptive intermediate stages in the development of the disease based on known mechanisms. In the development of malignancy, these steps may involve exposure to known mutagens and carcinogens, internalization and potential metabolism of a chemical agent, characterization of the interaction of the agent at its site of action (usually DNA), characterization of induced preneoplastic changes, and, in certain instances, early detection of the cancer itself. These processes can be monitored through biomarkers specific to each of the steps in the progression toward disease using any of the host of applicable techniques now available. An overview of such techniques is presented, with emphasis on techniques offering insight into the malignant process. Evidence is presented suggesting that although there are many potential contributing mechanisms to carcinogenesis, mutagenesis remains the dominant driving force behind the process. Several methods of monitoring mutation have shown promise as predictors of cancer incidence. These methods might also be used as monitors of agents designed to intervene in the process to prevent the development of overt disease.
Estimation of Cancer Risk Caused by Environmental Chemicals Based on In Vivo Dose Measurement
9
10.1615/JEnvironPatholToxicolOncol.v20.i4.20
Margareta
Tornqvist
Department of Environmental Chemistry, Stockholm University, S-106 91 Stockholm, Sweden
Lars
Ehrenberg
Department of Genetic and Cellular Toxicology, Stockholm University, S-106 91 Stockholm, Sweden
Methods for estimating the risks, that is, the probabilities of contracting a disease, are required as a basis of decision-making regarding the needs for protection and risk reduction. A mechanism-based model has been developed for estimating the cancer risks from genotoxic chemicals using adducts to macromolecules for determining the in vivo dose. On the assumption that cancer is caused by an increased frequency of mutations in tissues, interacting with inherited or acquired growth-promoting factors, a simplified model has been proposed for estimating cancer risks from chemically reactive, that is, mutagenic agents. According to a multiplicative model, the risk increment (DP) is DP = b ґ D ґ P0, proportional to the background incidence (P0) and linearly dependent on dose at low to intermediate doses (D); b is the risk coefficient, which is approximately the same for different tumor sites and, probably, different species. This model is already in use for ionizing radiation and has been validated for a few mutagenic and carcinogenic chemical compounds. Inherent in this method is the measurement of dose. Sensitive chemical methods have been developed for determining reactive compounds and intermediates in vivo through their stable reaction products (adducts) with proteins, particularly hemoglobin. In humans or animals, the doses of genotoxic agents can be derived from measured levels of adducts and rates of adduct formation. This approach has been applied to various exposures such as air pollutants in occupational settings, carcinogens in foods, and tobacco smoke. By such methods, exposures to previously unknown mutagens and carcinogens may be detected and assessed in humans. Examples of this are epoxides (from endogenously produced alkenes) and compounds (such as acrylamide) formed in cooking foodstuffs.
DNA Repair Studies: Experimental Evidence in Support of Chicken DT40 Cell Line as a Unique Model
11
10.1615/JEnvironPatholToxicolOncol.v20.i4.30
Pawan Kumar
Dhar
Department of Radiation Genetics, Faculty of Medicine, Kyoto University, Kyoto, Japan
Eiichiro
Sonoda
Department of Radiation Genetics, Faculty of Medicine, Kyoto University, Kyoto, Japan
Akira
Fujimori
Department of Radiation Genetics, Faculty of Medicine, Kyoto University, Kyoto, Japan
Yukiko M.
Yamashita
Department of Radiation Genetics, Faculty of Medicine, Kyoto University, Kyoto, Japan
Shunichi
Takeda
Department of Radiation Genetics, Faculty of Medicine, Kyoto University, Kyoto, Japan
DT40 is a chicken B lymphocyte cell line that exhibits a high ratio of targeted and random integration of transfected DNA constructs. Using the DT40 cell line makes it comparatively easy to disrupt multiple genes in a single cell and to generate conditional targeted mutants including tet-controlled cre-lox-mediated and temperature-sensitive mutants. The DT40 mutants show a strong phenotypic resemblance to murine mutants with respect to genes involved in DNA recombination and repair. Because of these characteristics, DT40 is an attractive model for the analysis of DNA recombination and repair studies in vertebrates despite obvious concerns associated with the use of a transformed cell line that may have certain cell-line-specific characteristics. We present experimental evidence to demonstrate the usefulness of the DT40 cell line as a unique model to study DNA damaging events and their associated repair pathways.
Photochemical Mutagenesis: Examples and Toxicological Relevance
8
10.1615/JEnvironPatholToxicolOncol.v20.i4.40
Elmar
Gocke
F. Hoffmann-La Roche Ltd., PRNS, Bldg. 73/215, CH 4020 Basel, Switzerland
Induction of DNA damage as a consequence of exposure to UV light has been established as the major cause of skin cancer. DNA molecules absorb photon energy directly for wavelengths <320 nm, and lead to well-characterized mutagenic DNA damage. Alternatively, endogenous or exogenous chemicals (sensitizers) may absorb light with the potential of subsequent energy or electron transfer, and lead indirectly to DNA damage. A few light-absorbing pharmaceuticals have long been known to cause photo (geno)toxic effects. Notably, psoralen and chlorpromazine derivatives have been established as photomutagens and the reaction mechanisms have been identified; the fluoroquinolone antibiotics have more recently been recognized as being photomutagenic. The type of DNA damage and the modulation by antioxidants indicate the involvement of reactive oxygen species (ROS), but other mechanisms are also reported for, at least, some derivatives. In routine genotoxicity studies, we observed the photomutagenic activity of a compound (Ro 19-8022) under development as an anxiolytic agent in the Ames tester strain TA102 under normal laboratory illumination conditions. Further investigations showed strong photogenotoxic activity in tests for gene mutations and chromosomal aberrations in mammalian cells. The finding led to the termination of drug development. Another example of a pharmaceutical for which photogenotoxic properties were observed during development is Ro 47-7737, a bisquinoline derivative of the antimalaria compound chloroquine. Also in this case, the photochemical reactivity contributed to the termination of the development process. The risk/benefit assessment for the described compounds has to take into account the human exposure situation, for example, the ability to avoid light exposure during treatment. Consideration of photochemical mutagenesis is specifically important for sunscreen ingredients. The active components of sunscreen lotions are efficient UV absorbers. Consequently, they reduce the genotoxicity of UV light and thus may be considered antimutagens. However, photodegradation to reactive molecules or energy transfer to DNA is possible, in principle, as has been reported for para-aminobenzoic acid (PABA).
Fluorescence In Situ Hybridization (FISH) in Genetic Toxicology
6
10.1615/JEnvironPatholToxicolOncol.v20.i4.50
A. T.
Natarajan
Department of Radiation Genetics and Chemical Mutagenesis, Leiden University Medical Center, Wassenaarseweg 72, 2333 AL, Leiden, The Netherlands
Structural and numerical chromosomal aberrations have been considered important biological end points in genotoxic studies. Conventional solid staining (such as Giemsa) has been employed to evaluate the frequencies of induced chromosomal aberrations following exposure to chemical or physical agents. Recently, molecular cytogenetic techniques that have become available, such as fluorescence in situ hybridization (FISH) using chromosome-specific or chromosomal regions-specific DNA libraries, have increased the resolution of detection of aberrations. The present paper reviews briefly the results obtained from basic and applied studies using the FISH technique.
Unanswered Questions in Arsenic Toxicology
11
10.1615/JEnvironPatholToxicolOncol.v20.i4.60
T. W.
Gebel
Medical Institute of General Hygiene and Environmental Health, University of Goettingen, Windausweg 2, D-37073 Goettingen, Germany
Arsenic (As) is one of the most important environmental global toxicants. In various countries and for decades people have been and currently are exposed to inorganic As through geogenically contaminated drinking water. An increased incidence of diseases mediated by this toxic element is the consequence of long-term exposure. Despite past extensive research on the toxicology of As, many questions remained unanswered, making risk assessment difficult. For instance, it is still not known how the carcinogenicity of As is mechanistically operative. Moreover, there is an increasing debate on whether the metabolic methylation of As has to be considered a detoxification process. Furthermore, it is historically documented that longterm intake of small amounts of As can lead to an acquired increased tolerance to its acute toxicity. It is not known whether this tolerance may be associated with a reduced chronic toxicity as well. In contrast to nonhuman cells, the selection of As-induced self-tolerance in human cells in vitro had been unsuccessful until now. However, we recently selected As-resistant human hepatoma HepG2 sublines that have low-level tolerance to As. Besides an approximately twofold elevated resistance to As cytotoxicity, this tolerance was associated with a significantly suppressed induction of As-mediated genotoxicity, which was evident in the cytokinesis-block micronucleus test. Additional questions arise when we consider several factors suspected to modulate the long-term toxicity of arsenic in vitro, variables that may either enhance or suppress the environmental genotoxicity and carcinogenicity of the metalloid. Besides malnutrition, these are single nutritional factors such as selenium and possible drinking water co-contaminants such as antimony. For instance, in the case of selenium, we could show that antimony (III) is able to suppress As genotoxicity. Taken together, research answers in many fields of As toxicology are needed in order to reduce the uncertainties in risk assessment of environmental As.
Alpha-Particle Carcinogenesis in Thorotrast® Patients: Epidemiology, Dosimetry, Pathology, and Molecular Analysis
5
10.1615/JEnvironPatholToxicolOncol.v20.i4.70
Yuichi
Ishikawa
Department of Pathology, The Cancer Institute, Japanese Foundation for Cancer Research. Toshima-ku, Tokyo, Japan
Ikuo
Wada
Department of Pathology, The Cancer Institute, Japanese Foundation for Cancer Research. Toshima-ku, Tokyo, Japan
Manabu
Fukumoto
Department of Pathology, Institute for Development, Aging and Cancer, Tohoku University, Sendai, Japan
We studied the alpha-radiation risks in patients who received injections of Thorotrast®, an X-ray contrast medium used in Europe, Japan, and the United States from 1930 to 1955. Thorotrast was composed of thorium dioxide (ThO2) and Th-232, a naturally occurring radionuclide. Because the physical half-life of ThO2 is 14 billion years and Thorotrast is hardly eliminated from the body, tissues in which it was deposited are irradiated by alpha-radiation for the entire lifetime of the subject. The dosimetry of Thorotrast patients is very complicated, but currently its reliability is quite high compared with other irradiated populations. The major causes of the death of Thorotrast patients are liver cancer, liver cirrhosis, leukemia, and other cancers. Three histologies of liver cancer are found: cholangiocarcinoma, hepatocellular carcinoma, and angiosarcoma. Although cholangiocarcinoma is the most frequent, angiosarcoma is characteristic of alpha-radiation. Among blood neoplasms with a higher incidence of increase than the general population, erythroleukemia and myelodysplastic syndrome were remarkable. Thorotrast patients exhaled a high concentration of radon (Rn-220), a progeny of Th-232, but no excesses of lung cancer in the patients of Japan, Germany, and Denmark were reported. Mutation analyses of p53 genes and loss of heterozygosity (LOH) studies at 17p locus were performed to characterize the genetic changes in Thorotrast-induced liver tumors. Interestingly, LOH, supposedly corresponding to large deletions, was not frequent; most mutations were transitions, also seen in tumors of the general population, suggesting that genetic changes of Thorotrast-induced cancers are mainly delayed mutations, and not the result of the direct effects of radiation.
DNA Damage in Birds After the Mining Waste Spill in Southwestern Spain: A Comet Assay Evaluation
8
10.1615/JEnvironPatholToxicolOncol.v20.i4.80
Nuria
Pastor
Department of Cell Biology, Faculty of Biology, University of Seville, Spain
Miguel
Lopez-Lazaro
Department of Pharmacology, Faculty of Pharmacy, University of Seville, Spain
Jose Luis
Tella
Department of Applied Biology, Estacion Biologica de Donana, Consejo Superior de Investigaciones Cientificas (CSIC), Seville, Spain
Raquel
Baos
Department of Applied Biology, Estacion Biologica de Donana, Consejo Superior de Investigaciones Cientificas (CSIC), Seville, Spain
Manuela G.
Forrero
Department of Applied Biology, Estacion Biologica de Donana, Consejo Superior de Investigaciones Cientificas (CSIC), Seville, Spain
Fernando
Hiraldo
Department of Applied Biology, Estacion Biologica de Donana, Consejo Superior de Investigaciones Cientificas (CSIC), Seville, Spain
Felipe
Cortes
Department of Cell Biology, Faculty of Biology, University of Seville, Spain
In April 1998, an ecological disaster resulting from a massive toxic spill of mining acid waste rich in heavy metals posed a serious threat to the Donana National Park in southwestern Spain. This especially important protected area is the nesting and breeding site for many endangered bird species; white storks (Ciconia ciconia) and black kites (Milvus migrans) are considered the more representative. The suitability of the Comet assay as a biomarker for genotoxic analysis in environmental biomonitoring has been recently validated in studies using different sentinel organisms such as fish, amphibians, rodents, or mollusks. Birds preying on a variety of invertebrate and vertebrate species in the marshlands are appropriate for evaluating the potential deleterious effects of the toxic spill on wildlife of the Donana area. Our study on wetland birds high on the aquatic trophic chain sampled within a few months after the toxic spill in the area around Donana National Park has shown the accumulation of heavy metals. Fourteen months after the mine waste spill, blood samples from white storks and kites collected in the neighborhood of the park and from control birds at reference areas for comparison were examined by fluorescence image analysis after lymphocyte isolation, and by subsequent alkaline single-cell gel (SCG) electrophoresis, known as the Comet assay. Our results indicate that the exposed birds had a significantly increased level of genotoxic damage compared with control animals from noncontaminated locations.
Micronucleus Test in Fish from a Pampasic Pond (Argentina): An Estimation of the Presence of Genotoxic Compounds
7
10.1615/JEnvironPatholToxicolOncol.v20.i4.90
Marcela A.
Campana
Facultad de Ciencias Exactas y Naturales, Universidad Nacional de Mar del Plata, Mar del Plata, Argentina
Ana M.
Panzeri
Facultad de Ciencias Exactas y Naturales, Universidad Nacional de Mar del Plata, Mar del Plata, Argentina
Alicia H.
Escalante
Facultad de Ciencias Exactas y Naturales, Universidad Nacional de Mar del Plata, Mar del Plata, Argentina
Victor J.
Moreno
Facultad de Ciencias Exactas y Naturales, Universidad Nacional de Mar del Plata, Mar del Plata, Argentina
Fernando N.
Dulout
Centro de Investigaciones en Genetica Basica y Aplicada (CIGEBA), Facultad de Ciencias Veterinarias, Universidad Nacional de La Plata, La Plata, Argentina
The Los Padres pond is one of the commonly shallow, polymictic lakes from the so-called depressed Pampa (Argentina). Its watershed includes one affluent, named Los Padres creek, which flows through horticultural lands wherein great amounts of pesticides are applied. Opposite to this stream, the pond drains into La Tapera creek that is the effluent running toward the sea. Many studies have confirmed the capacity of various pesticides to induce genetic damage. The use of micronucleus (MN) tests in fish has enabled us to detect the presence of contaminants in the lake water and to evaluate their genotoxic effects. For this purpose, water samples were collected during April, August, and December 1999 from both creeks characterized by different environmental conditions. In the laboratory, specimens of tetras Cheirodon interruptus (Pisces, Characidae) were reared in water samples from the two creeks. Control fish were kept in drinking water. Fifteen individuals from each experimental group were sacrificed after 24-, 48-, and 72-hour exposure intervals. Micronucleus frequency in fish erythrocytes was determined, and the Kruskal–Wallis test for statistic analysis was used. We made the following observations: (1) Highly significant differences occurred in MN frequency between the control group and the samples from both creeks. (2) An increase in MN frequency was evident in specimens sampled from the affluent input during the month of December. These results allowed us to conclude that the increase in MN frequency observed in fish belonging to both sampling sites would indicate the existence of genotoxic compounds in the Los Padres pond. The high MN frequency in fish collected near Los Padres creek inlet might be related to the polluted load transported by the affluent and discharged into the lake’s surface waters. Future work would allow us to develop efficient methods for predicting the presence of genotoxic contaminants. It would be possible then to propose strategies for regulating and decreasing the sources of pollution that affect human health.
Inhibition of Cell Proliferation and Antitumor Activity of a Novel Organotin Compound
10
10.1615/JEnvironPatholToxicolOncol.v20.i4.100
Christine
Syng-ai
Genetics Laboratory, Department of Zoology, North Eastern Hill University, Bijni Complex, Shillong, India
Tushar S. Basu
Baul
Chemical Laboratory, Regional Sophisticated Instrumentation, North Eastern Hill University, Bijni Complex, Shillong, India
Anupam
Chatterjee
Genetics Laboratory, Department of Zoology, North Eastern Hill University, Bijni Complex, Shillong, India
Organotin compounds showed more antineoplastic effect against P388 leukemia in mice than any other class of compounds. However, they have not received as much attention as the platinum compounds. The present compound, Et2SnCl2.L [L = N-(2-pyridylmethylene)-4-toluidine] (OTC), showed an Sn-N bond length of 2.46 A which, because it is bigger than 2.39 A, is expected to achieve better formation of tin-DNA complexes. We previously reported on the synthesis and biological activity of this compound and on its ability to cause a delay in cell proliferation and sister chromatid exchanges in mouse bone marrow cells. In this study, we carried out further investigations on the antiproliferative and antitumor activity of OTC in relation to the cellular glutathione (GSH) level, which plays an important role in the cellular defense mechanism. OTC induced significant delay in the cell cycle in mouse bone marrow cells and, when the GSH level was low, the extent of the delay was reduced. The antitumor activity was determined in accordance with the US National Cancer Institute (NCI) standard protocol for primary screening in Dalton’s lymphoma (DL) that was maintained by serial intraperitoneal transplantation. The T/C (treated/control) value was 146% when organotin was treated after transplantation; this improved significantly when buthionine sulfoximine (BSO), a GSH-depleting agent, was added 24 hours before the tin treatment. Our data suggest that the present tin compound has antiproliferative ability and can increase the survival of mice bearing DL. The endogenous GSH level influences the effect of the tin compound.