Chemical Mutagenesis and Carcinogenesis

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Experimental carcinogenesis and human epidemiology studies have clearly identified specific chemicals that can act as human carcinogens. Certain chemicals have been associated with increased human cancer incidence in both occupational and environmental exposure settings.


It is now widely recognized that exposure to chemicals in the workplace and the environment can contribute to human cancer risk. This was first postulated in 1775 by Dr. Percival Pott, who recognized an association between occupational exposure to chemicals in soot and an increased incidence of scrotal cancer in London chimney sweeps. Other sporadic clinical observations of this type throughout the 19th century also suggested an association between certain occupational exposures to specific chemical agents and increased human cancer. However, it was not until the 20th century that science and medicine actively investigated this aspect of cancer etiology. Boveri first hypothesized that cancer was a genetic disease in 1921, prior to the discovery of the genetic material. In 1915, Yamagawa and co-workers demonstrated that application of coal tar could induce tumors in animals.      
In the 1930s Kenneway and co-workers demonstrated that pure chemicals isolated from coal tar could also induce animal tumors. Parallel discoveries in the 1950s of the structure of the DNA double helix and its establishment as the hereditary material on the one hand and mutagenic potential of ionizing radiation and certain chemical carcinogens in humans and experimental systems on the other set the stage for extensive investigations into the relationship between chemically induced mutations and human cancer. Public perceptions, and the regulatory policies resulting from them, have had a strong influence on the direction of chemical carcinogenesis studies and their interpretation. The infamous Delaney Clause of the 1950s led to an intensive focus on the potential carcinogenicity of food additives and other contaminants, and also established the paradigm of the animal tumor model as a test system for predicting carcinogenicity of chemicals in humans. Similarly, the environmental movement of the 1960s and 1970s, which began with the publication of Rachel Carson's Silent Spring and the subsequent discovery of Love Canal, led to the establishment of the Federal Environmental Protection Agency and "Superfund," clean air, and clean water legislation. This movement was accompanied by a growing public concern that the widespread use of pesticides and exposure to chemicals from toxic waste sites could cause cancer, and more generally to the belief that chemicals in the environment, particularly man-made chemicals, were responsible for a major fraction, perhaps the majority, of human cancers. The 1980s saw the elucidation of the first oncogenes, genes that appeared to be responsible for the initiation of cancer as first predicted by Boveri. This era also saw the development of the Ames Salmonella bacterial mutagenesis assay and hundreds of similar genetic toxicology assays. These developments firmly established the basic paradigm for the field of chemical carcinogenesis; that chemicals that can cause mutations are presumed to be carcinogens. By extension, it was predicted that any chemical or physical agent that could covalently damage DNA could also cause mutations through its DNAdamaging mechanism, and hence could also be a carcinogen.      
The data that followed over the next decade appeared to strongly support this central tenet, as the vast majority of chemicals that were initially tested for DNA damage or mutations were also shown experimentally to be animal carcinogens. However, most of the chemicals that were initially tested were either known or suspected carcinogens, were agents of concern, or were closely related structurally to these known and suspected carcinogens.      
Over the most recent decade, our understanding of the molecular basis of cancer has substantially improved. In addition, investigations into the molecular basis of chemical carcinogenesis, as well as more extensive human cancer epidemiology studies using modern molecular tools, have greatly expanded our knowledge in this area. This has led to a reevaluation of the overall role of chemicals in human cancer, as well as a modification of the basic paradigms that form the initial foundations of the field. In contrast to earlier predictions, estimates suggest that perhaps only 4-8% of human cancers can be directly attributed to specific chemical exposures. Moreover, these exposures are primarily occupationally related or involve specific environmental contaminants, and it is therefore unlikely that environmental exposures to most chemicals contribute significantly to the overall burden of human cancer. The reasons for this are several fold.      
First, while many chemicals have the potential to be genotoxic (DNA damaging), mutagenic, or carcinogenic in various test systems, the levels of most environmental human exposures are usually too low, or are too transient, to have an appreciable impact on cancer relative to other risks. Second, it is also now clear that humans have many protective mechanisms that can act as practical threshold barriers to most minor chemical exposures. These include physiological barriers to chemicals, xenobiotic metabolism and other pharmacokinetic mechanisms of detoxification, a highly efficient DNA repair system, various active apoptotic mechanisms, and various immune and other active surveillance mechanisms. These thresholds are often not present, or are exceeded or bypassed, in many test systems, including rodent bioassays. For example, some agents are only carcinogenic in animals when administered by large bolus intraperitoneal or intramuscular injections that bypass the normal dose, time course, and route of exposure that would otherwise have resulted in detoxification. Chemicals are also usually given to test animals at or near their maximum tolerated dose (MTD), whereas people are usually exposed to these same chemicals at levels that are thousands or even millions of times lower. About half of all chemicals that have been tested in rodents, whether natural or man-made, have been shown to increase tumor formation. However, the list of chemicals that have been tested to date is highly biased.       
Third, based on a growing database, it is now also clear that while there are considerable overlaps among groups of chemicals that are genotoxic, mutagenic, or carcinogenic, these properties are not equivalent. For example, not all chemicals that are genotoxic are mutagenic. This is likely to be due to both the ability of the cell to recognize certain kinds of damage with extremely high efficiency at low levels and the observation that not all types of damage are equally mutagenic to the cell during DNA replication. Similarly, there are many chemicals that are mutagenic but not genotoxic. For example, certain metal ions that do not cause DNA damage can interact with and directly alter the fidelity of DNA polymerase, at least in vitro, and thereby lead to mutations due to replication errors. Moreover, DNA-intercalating agents such as ethidium bromide, which can transiently stack between the base pairs of the DNA, can also cause mutations in the form of frameshifts (additions or losses of DNA base pairs), without any covalent modification of DNA.      
Most importantly, there is a large list of chemicals of concern that are carcinogenic, based on evidence in experimental animals or as demonstrated directly in human epidemiology studies, but that have not been found as overtly genotoxic or mutagenic in experimental test systems. Approximately one-third of chemical mutagens are negative as chemical carcinogens in animals, and vice versa. Examples of carcinogenic agents that are not genotoxic or mutagenic include the so-called solid-state carcinogens (e.g., smooth plastic implants) and the carcinogenic metals, arsenic and cadmium. In fact, in the case of cadmium and arsenic, it has been difficult to demonstrate that they are carcinogenic as single agents in animal models, despite strong epidemiological evidence that they are carcinogenic in humans.
Moreover, many chemicals that were initially thought to act by genotoxic and mutagenic mechanisms may, in fact, act primarily through other as yet unclear mechanisms that do not involve DNA damage or mutations per se. For example, although the human lung toxin and carcinogen chromium(VI) is moderately positive for DNA damage and mutations in certain experimental test systems, there is growing evidence that it may act as a human carcinogen primarily through cell signaling and tumor promoter-associated mechanisms rather than or in addition to its genotoxic or mutagenic effects. It is therefore important, in assessing the overall carcinogenic potential of a chemical, to consider whether it is a genotoxin, mutagen, or animal or human carcinogen separately and to also classify the type of mechanism involved. Thus, chemically induced DNA damage, chemical mutagenesis, and chemical carcinogenesis are considered separately.


A. Types of Chemical Interactions with DNA
Two basic types of chemical interaction with DNA are noncovalent and covalent binding. Examples of the primary types of noncovalent interactions include (a) ionic interactions, such as when Mg 2 and other cations interact with the negative phosphate groups on the outside of the DNA helix; (b) minor groovebinding chemicals, such as the dye Hoechst 33258; and (c) intercalating agents, such as ethidium bromide and bleomycin, which are planar aromatic compounds capable of stacking between the parallel base pairs inside the DNA helix. These various noncovalent interactions can each have toxicological consequences. For example, such interactions can affect the structure of the DNA helix or disrupt DNA- protein interactions within chromatin, leading to changes in DNA replication and RNA expression.      
They can also alter the fidelity of DNA and RNA polymerases, in the former case increasing the probability of mutations. Noncovalent chemical-DNA interactions are not considered damage perse; however, the noncovalent interactions of some chemicals can subsequently lead to the generation of covalent DNA damage. For example, intercalation of bleomycin into DNA and subsequent binding of iron to the drug can lead to the generation of reactive oxygen species that can damage the DNA at sites adjacent to the drug intercalation site. At high levels, this damage will kill the cell, which is the basis for the use of bleomycin as an anticancer drug, but at lower levels this damage can also result in mutations. Covalent interactions occur when the chemical, or a portion of the chemical, is covalently adducted to the DNA helix or when it causes other types of covalent modifications, such as oxidative damage. Chemicals that can directly adduct to DNA are often referred to generically as "DNA-alkylating agents." The various types of DNA damage are discussed in further detail.

B. Types of DNA Damage
1. Spontaneous DNA Damage
In the context of this topic there are two basic classes of DNA damage to consider, i.e., so-called "spontaneous" or background damage and chemically induced damage. Spontaneous DNA damage includes deamination, loss of bases to form abasic sites, and oxidative damage. Deamination, i.e., chemical loss of amino groups on bases, occurs frequently and spontaneously. It has been estimated that approximately 10,000 deamination events occur per cell per day, on average, in humans and other mammals. Deamination usually leads to the formation of unusual bases, e.g., cytosine deamination forms uracil (which is the thymine analog in RNA but not a normal DNA base) and adenine deamination forms hypoxanthine.
These deaminated bases are potentially mutagenic. If left unrepaired, some abnormal bases can form an alternative base pair with the wrong partner during replication, leading to a mutation at that site. For example, uracil can base pair with adenine because it is a structural analog of thymine so that deamination of cytosine to uracil can result in a transition from a C-G base pair to a T-A base pair if undetected. However, the common deamination products are detected by specific enzymes called DNA glycosylases, each of which recognizes a specific inappropriate base in DNA (e.g., uracil glycosylase). Some deamination events do not have a mutagenic consequence, e.g., deamination of guanine leads to the formation of xanthine, which, if unrepaired, normally will still be base paired preferentially with cytosine during replication. From the standpoint of mutations, the most important deamination is of 5-methylcytosine, as this leads to formation of thymine, which is a normal DNA base. If this mismatch is left unresolved, it can lead to a transition mutation to a T-A base pair during replication.      
In fact, it has been observed that sites of 5- methylcytosine are typically "hot spots" for mutation, with a much higher mutation frequency than other sites. Repair of these mismatches requires a complex repair process that queries each strand to identify the parental and daughter strands of the helix. This allows it to determine whether the site should be methylated to determine which base is incorrect.      
Abasic sites occur when a glycosidic bond in DNA weakens and the helix releases the base at that site. This also occurs at a high spontaneous level, with approximately 20,000 purines and 1000 pyrimidines lost per cell per day on average. These abasic sites are also recognized and repaired by the cellular repair machinery. Background oxidative DNA damage and radiation-induced DNA damage are often classified as "spontaneous" damage because they are part of the normal background. A low level of reactive oxygen species (primarily hydroxyl radical and superoxide) is generated chemically or enzymatically in cells on a continuous basis, and some of these reactive byproducts will attack DNA to cause oxidative base damages such as 8-oxodeoxyguanosine. Similarly, background levels of ultraviolet (UV) and ionizing radiation are encountered by virtually all organisms. Low levels of background DNA damage (pyrimidine dimers and 6-4 photoproducts from UV; oxidized bases, and single and double strand breaks from ionizing radiation) can occur from this radiation. However, these lesions are also normally recognized and repaired by the cellular machinery.       
It is important to consider spontaneous or background levels of DNA damage in considering chemically induced DNA damage for several reasons. First, it is clear that all the cells in our bodies are continuously challenged by these background levels of DNA damage, which, if left unrepaired, are potentially mutagenic and therefore possibly carcinogenic. Thus, normal individuals will have low background levels of DNA damage and mutations even in the absence of chemical exposure, which must be taken into account when assessing the added impact of chemical exposures that might contribute to these levels. One obvious way that this might occur is that chemicals can directly damage DNA and thereby induce mutations.      
However, additional mechanisms might include altering important cellular processes, such as DNA damage recognition and repair, replication fidelity, cell cycle checkpoints, apoptotic mechanisms, or overall cell proliferation rates, each of which might strongly influence the mutation rate from these background forms of DNA damage. Alternatively, one must consider very low levels of chemically induced DNA damage in the context of the background damage and repair that is always present. This additional damage may be inconsequential with respect to increasing the overall rate of mutation until a specific threshold of damage is attained such that the probability of mutation is increased to a measurable level. Understanding the relationship between background and chemically induced DNA damage and mutations has important implications for risk assessment, particularly for determining the appropriate model for extrapolating from measurable effects at high-dose human or animal exposures to possible risks from very low-dose exposures.

2. Chemically Induced DNA Damage
Chemically induced DNA damage includes oxidative damage, simple alkylation, bulky monoadducts, and DNA cross-links. Chemically induced oxidative DNA damage can result from the generation of hydrogen peroxide in the cell or from redox reactions generated by chemicals such as menadione, bleomycin, and certain metals inside the cell. Simple alkylation typically occurs when a reactive parent compound has a reactive methyl or ethyl group that can covalently interact a DNA base, leaving the alkyl group behind.
Examples of alkylation agents include direct-acting compounds, such as methylor ethyl-methanesulfonate, and indirect-acting agents, such as methyl- or ethyl-nitrosourea, which require activation to a reactive intermediate metabolite by the phase I cytochrome P450 system. Simple alkylation normally involves the addition of a methyl or ethyl group to the N2, N7, and O6 of guanine but also occurs at other nucleophilic sites of DNA. The O6-methylguanine adduct in particular is highly mutagenic if unrepaired. Many organic mutagen-carcinogens form bulky monoadducts with DNA, typically following metabolism of the parent compound to a reactive intermediate by the phase I cytochrome P450 system. Examples of chemical carcinogens that do this are mycotoxin, aflatoxin B1 (AFB1), and the polycyclic aromatic hydrocarbon, benzo[a]pyrene (BaP), each of which has been shown to induce bulky monoadducts that cause specific mutations at the site of adduction.      
With the advent of modern molecular biology techniques, it has been possible to determine the precise mutations that result from each type of chemical adduct and to compare this mutational pattern with the mutations found in oncogenes and tumor suppressor genes of tumors produced by that chemical in experimental animals. For example, BaP is metabolized to the reactive intermediate benzo[a]prene-7,8,- diol-9,10-epoxide (BPDE), which forms an adduct at the N2 of guanine and results in a G-C to T-A mutation. Use of site-specific adducts of BPDE in shuttle vectors has shown that this lesion almost exclusively results in G-C to T-A mutations at the site of the lesion.       
Cells in culture or tumors from animals treated with BaP or BPDE have certain activated oncogenes, and the mutations that occur in these oncogenes are G-C to T-A mutations. Oncogenes that have been synthesized to contain specific G-C to T-A mutations have been shown to be sufficient to transform cells to tumorigenic cells in culture, indicating that this type of mutation in and of itself can contribute to the carcinogenic process. Thus, these empirical observations, at least in the example of BaP, match the predictions of the basic paradigm linking chemical genotoxicity to mutagenesis and carcinogenesis, i.e., that certain chemicals can cause cancer by attacking and damaging DNA, generating DNA adducts that cause specific mutations in critical cancer genes that, in turn, can contribute to a cancer cell genotype and phenotype.      
DNA cross-links are another category of DNA damage induced by chemicals and some forms of radiation. The three basic types are interstrand (covalent linkage of two bases on opposite DNA strands), intrastrand (linkage of bases on the same strand), and DNA-protein cross-links. Different bifunctional agents can induce one, two, or all three types of crosslinks, in addition to other types of DNA lesions, with varying efficiencies. For example, the human lung carcinogen chromium(VI) induces both DNA interstrand and DNA-protein cross-links, in addition to the formation of Cr-DNA monoadducts and the generation of reactive oxygen species that can also damage DNA. Similarly, the cancer chemotherapy agents mitomycin C and cisplatin can each induce a spectrum of monoadducts, DNA interstrand cross-links, and DNA intrastrand cross-links. DNA interstrand cross-links, in particular, are very difficult for the cell to repair because both strands of DNA are involved, and if left unrepaired these lesions can be lethal to the cell. This is believed to be the principal basis for the ability of mitomycin, cisplatin, and similar crosslinking agents to kill cancer cells. However, at lower doses it is not clear whether DNA interstrand crosslinks are mutagenic. Similarly, DNA-protein crosslinks do not appear to be strongly associated with an increase in mutations. These lesions are generally well recognized and repaired by the cellular machinery.      
Most chemically induced DNA intrastrand cross-links are also easily recognized and repaired, as they usually cause severe distortions of the DNA helix. UV light can cause thymine dimers and 6-4 photoproducts, two specific forms of interstrand cross-links. Thymine dimers are also recognized and repaired in cells, whereas the 6-4 photoproduct appears to be poorly recognized and is very mutagenic if left unrepaired. Formation of this adduct is probably the primary basis for the increased risk of skin cancer in heavily sunexposed people and in xeroderma pigmentosum (XP) patients who are deficient in specific repair enzymes (discussed in further detail later). Ionizing radiation and some chemicals can cause single and double strand breaks in the DNA helix. Single strand breaks are usually repaired by the cellular machinery and are a normal intermediate in many DNA replication and maintenance processes. Double strand breaks can be repaired, although poorly and with slow kinetics, as this involves a more complex repair process, and so these are considered lethal to a dividing cell. This is one of the primary cellular lesions induced by therapeutic ionizing radiation that leads to tumor cell cytotoxicity.      
Assays for DNA damage are generally of two types: they are either very specific for a particular type of damage or they detect overall levels of DNA damage without information about the type of damage. For example, carcinogen-binding assays with radiolabeled compounds provide information about covalent adduction but not other types of DNA damage (e.g., oxidative damage or strand breaks). Oxidative damage can be assessed by HPLC analysis of altered bases or use of lesion-specific antibodies. DNA alkaline elution is a technique that can be used to measure DNA interstrand and DNA-protein cross-links, as well as frank strand breaks and so-called "alkali labile sites," i.e., regions of weakness in the DNA helix, presumably as a result of damage, that are elaborated in the presence of strongly alkaline conditions. Assays using 32P postlabeling for altered bases can be used to assess a spectrum of adducts from a given agent, but the technique must be customized to each agent and its DNA damage products, and many agents are not amenable to this analysis for technical reasons. Similarly, HPLC and antibody-based detection of adducts can be developed, but these methods are agent specific.      
Sister chromatid exchanges occur at a low level in all dividing cells but are increased by many different chemical and physical agents that damage DNA. These events can be detected by differential staining or antibody techniques. However, it is not clear whether chemically induced exchanges between identical chromatids represent DNA damage, successful or unsuccessful repair of damage, or whether there are mutations associated with these events (e.g., by unequal exchange of sequences). Light microscopy can be used to measure increases in chromosomal aberrations, and formation of micronuclei and "unscheduled DNA synthesis" (i.e., DNA synthesis not associated with replication, which is presumed to be DNA repair in response to induced DNA damage) in treated cells can also be used to assess the effects of genotoxic agents, but these techniques also do not distinguish individual lesions. It is therefore often useful to use several complementary assays when assessing DNA damage from a particular chemical.

C. Repair of DNA Damage
Eukaryotic DNA repair is a highly efficient and coordinated process that normally protects cells and organisms from chemical or genetic alterations to the genetic material. This is a continuous process in the cell that counteracts the background levels of DNA damage that occur in each cell every day. That DNA repair is critical both for suppressing the cancer process and for maintaining life itself can be illustrated by two related observations. First, as mentioned earlier, it has been estimated that in the absence of any other chemical or physical agent, each cell of the body loses or experiences damage to several thousand DNA bases per day through spontaneous chemical degeneration of the DNA helix, which would be potentially mutagenic or lethal were it not repaired on an ongoing basis.      
These tens of thousands of potentially mutagenic lesions are occurring in each of the trillions of cells in our body on a daily basis. However, the average life span of U.S. citizens is now expected to extend well into the eighth decade, and cancer is not a significant risk for most of us until the sixth or seventh decade of life. Second, among the hereditary mutations described to date that are known to predispose humans to cancer, the majority involve defects in genes whose proteins mediate specific steps of DNA repair. For example, individual mutations in eight different genes all result in the clinical disease xeroderma pigmentosum. XP patients have a high rate of skin cancer from even low levels of sun exposure, and all of the XP mutations occur in genes involved in DNA excision repair.      
Studies of the different "complementation groups" of these XP mutations have given us major insights into the specific genes and proteins involved in DNA excision repair in mammals. Mutations in various other DNA repair genes are also associated with an increased risk of cancer and other diseases, including a gene involved in mismatch repair that predisposes to colorectal cancer (hereditary nonpolyposis colon cancer, HNPCC) and genes associated with Bloom's syndrome, Fanconi's anemia, and trichithiodystrophy.      
There are four basic types of repair, i.e., direct repair, base excision repair (BER), (poly)nucleotide excision repair (NER), and postreplication repair (recombination and mismatch repair). Sexual reproduction has been described as a fifth form of DNA repair, and the ultimate form of genomic DNA repair, as the pairing of alleles from two different individuals provides an opportunity for the genome of the offspring to contain at least one intact copy of each critical gene. Direct repair involves chemically restoring a damaged base to its original structure, e.g., by reversal of a UV light-induced lesion by the photoreactivation enzyme, photolyasem, or removal of a methyl or ethyl group from guanine by O6-methylguanine methyltransferase. BER involves removal of a damaged base from the DNA helix and subsequent replacement by the repair machinery. Different lesions are recognized by specific enzymes, e.g., uracil glycosylase. Nucleotide excision repair (more properly polynucleotide excision repair) involves recognition of a wide variety of DNA damages, excision of a patch of DNA surrounding the lesion by an enzyme complex, and repair of the gap by the replication machinery. Mismatch repair is a process that is linked to DNA replication, whereby mismatches created by DNA polymerase infidelity are corrected as part of the replication process. HNPCC genes are involved in mismatch repair. Why HNPCC mutations would specifically predispose people to colon cancer rather than a general increase in overall cancer risk is not clear. These examples of genetic predispositions to cancer illustrate how important DNA repair is for suppressing carcinogenesis from background and chemically induced DNA damage.


A. Types of Mutations
The three basic classes of genetic mutations are point mutations, clastogenic mutations, and aneuploidy events. Point mutations are arbitrarily defined as changes in the DNA sequence of 10 bp or less. These can include single base pair changes (purine-to-purine and pyrimidine-to-pyrimidine transitions and purine-pyrimidine transversions) and small insertions or deletions of 1-10 bp. Transitions and transversions can result in changes in the coding of an individual amino acid if it occurs in the first or second codon position, but can be silent mutations if in the "wobble" position of many amino acid codons. These mutations can also result in creation of stop codons that result in mRNA and protein truncation. These can also cause changes in other types of genetic information, such as alterations in binding sites for transcription factors and other regulatory proteins within promoter regions, methylation sites within promoter regions, and mRNA splice sites that define the final mRNA structure and sequence. Insertions or deletions that are not divisible by three and that occur within coding regions of base pairs will result in frameshifting, such that the remainder of the coding sequence will have a dramatically altered amino acid information. This often results in the creation of premature termination signals and truncated gene products.      
Clastogenic mutations include insertions or deletions of greater than 10 bp, inversion of a sequence of DNA within the same chromosome, duplication of DNA sequence (which can include entire gene segments or gene clusters), and gene amplification. Amplification events can result in homogeneously staining regions (HSRs) that are visible in chromosome karyotypic analysis, or extrachromosomal "minichromosomes" that contain high copy numbers of an amplified gene or genetic cluster. Amplification events are inducible even in normal cells, at least in cell culture, and also reversible, suggesting that they represent a survival strategy for upregulating certain important genes during times of severe stress.      
Translocation events can occur, where regions of different chromosomes swap locations. This is a common event in cancer and appears to be important in the etiology of certain cancers. The classic example of this is the "Philadelphia chromosome," which is a translocation of the short "p" arms of chromosomes 9 and 22 +(9;22) and a hallmark of chronic myelogenous leukemia. Many translocations in cancer result in the juxtapositioning of an oncogene with a strong promoter region, resulting in substantial upregulation of oncogene expression (e.g., BCL-2/IG). Other translocations result in either truncation of a gene product, which can be an activation event for certain oncogenes (e.g., MYB), or creation of a chimeric gene product (e.g., PML-RARα in acute promyelocytic leukemia or BRC-ABL in chronic myelogenous leukemia).      
Aneuploidy involves gain or loss of one or more entire chromosomes. Only two human aneuploidy events are compatible with life if they are germline or occur somatically in the early embryo, namely, trisomy 21 (Down's syndrome) and XO (Turner's syndrome). However, most malignant cancers arising from somatic mutations exhibit extensive genetic rearrangement and aneuploidy events, resulting in what is called "loss of heterozygosity" (LOH) or, more properly, loss of dizygosity as the genes that remain are haploid (single copy). This genetic instability and increasing haploidy is considered to provide a selective advantage to the tumor, but whether these events are primarily causal in development of a fully malignant phenotype, and/or are a result of processes that occur in a more advanced cancer phenotype, is still not fully understood.

B. Chemically Induced Mutations
As described earlier, certain chemicals are genotoxic, i.e., they can directly or indirectly cause covalent DNA damage. Certain forms of damage potentially result in mutations if unrepaired. The biochemical mechanism for this, at least for the handful of chemicals that have been investigated at this level of mechanistic detail, is primarily by causing DNA polymerase to misread a template base (usually containing, or immediately adjacent to a site of covalent DNA damage) and causing polymerase to insert the incorrect base into the daughter strand when forming a base pair. Lesions that are most mutagenic appear to not only cause DNA polymerase to make this mistake, but also form an alternative base pair that is poorly recognized as being incorrect by the DNA replication machinery, thereby eluding postreplication mismatch repair as well. For example, the BPDE lesion bound to the N2 of guanine can cause the damaged base to rotate about its glycosidic bond, flipping the base over.      
This results in polymerase adding an adenine at the site to form a base pair with the "Hoogsteen" face of the flipped guanine, ultimately resulting in the G-C to T-A transversion that is characteristic of BPDE mutations. Similarly, the O6-methylguanine lesion, which is caused by many simple alkylating agents, can continue to base pair with a protonated cytosine, such that the site appears normal to the machinery, but O6-methylguanine can also be inappropriately base paired with thymine, resulting in a G-C to A-T transition. Moreover, the cellular machinery appears to poorly recognize the alternative thymine-containing base pair as well. Thus, the O6-methylguanine adduct can persist without detection, and this lesion can cause mutations during replication that are poorly de- tected. The prevalence of this adduct in background levels of oxidative DNA damage and its high mutagenic potential are probably why evolution has developed a specific repair pathway (O6-methylguanine methyltransferase) that recognizes and removes this lesion from DNA.     
As mentioned previously, chemicals can influence mutations in the absence of inducing overt DNA damage. One class of agents that can do this are chemicals that noncovalently interact with DNA. DNAintercalating agents such as ethidium bromide, acridine orange, bleomycin, actinomycin, and the various anthracyclines (e.g., daunorubicin, doxorubicin) can insert between the base pairs of DNA, forming stable interactions by virtue of their planar multiring structure and the formation of favorable electronic interactions with the ring systems of the purine and pyrimidine DNA bases. However, this intercalation has the effect of "stretching" the DNA helix along its long axis, and also altering DNA-protein interactions along the face of the helix. Intercalating agents can cause frameshift mutations, especially in sequential runs of the same base, e.g., a run of four or more adenines. Adjacent bases in these runs can share electronic pairing of the opposite bases, such that if DNA polymerase adds or omits a base on the opposite strand, the error can be difficult to detect. Intercalating agents appear to be able to increase the probability of these frameshift events occurring, as one typically sees insertion or deletion of single bases in these contiguous runs. Minor groove-binding drugs can also influence the rate of mutation, although the mechanism for this is not clear. Other agents, especially certain metal ions, can influence the fidelity of DNA polymerase itself, at least in in vitro and cell culture systems. Agents that block or suppress specific steps in DNA repair can increase mutations, presumably resulting from background and spontaneous DNA damage events.      
Another class of nongenotoxic chemicals that can increase mutagenesis in the presence of other agents are called "comutagens." An example of this is 2,3,7,8- dibenzo-p-dioxin (TCDD, dioxin) and similar polyhalogenated hydrocarbons, which are strong inducers of specific isozymes of cytochrome P450. This can lead to increased activation of other agents such as BaP, resulting in much greater mutagenesis following a low-level exposure than would otherwise occur. Another example is agents that can influence DNA damage recognition and/or repair of DNA lesions. Arsenic, which is not genotoxic or mutagenic per se, is a strong comutagen when present with other genotoxic agents such as BaP or AFB1. The precise mechanism for this is not clear, but arsenic has been shown to influence the expression of DNA repair genes and to affect the fidelity of DNA replication and repair. Thus, one might predict that arsenic is most carcinogenic in combination with other agents, such as cigarette smoke or UV irradiation, and animal data and human epidemiology studies support this prediction.       
As mentioned earlier, although the vast majority of mutations seen in human tumors are large insertions, deletions, rearrangements, and aneuploidy events, the vast majority of tests for genotoxicity and mutagenesis, and therefore the vast majority of our information about chemical carcinogens, have focused on their ability to cause point mutations, especially single base pair transitions and transversions. The development of the Salmonella reversion assay by Bruce Ames and colleagues in the early 1980s (the "Ames test") led to an ability to rapidly screen chemicals for their potential to cause simple point mutations in this bacterial system. While this provided a high throughput and rapid and inexpensive screening system, there are two apparent drawbacks of this assay. First, it is a prokaryotic system, which differs in important respects from human cells in its uptake, metabolism, and DNA machinery.      
Second, it focuses on reversion mutations (i.e., reverting from a mutant to a wild-type sequence) under strong selective pressure for specific phenotypes. Thus, there is a strong bias to only observing certain mutations in this system. Similar assays have also been developed in yeast and other lower eukaryotes, utilizing both reverse and forward (wild-type to mutant) mutation screening, and in mammalian cells with screens for both forward and reverse mutations. Use of mammalian forward mutation systems addresses some concerns about the bacterial and yeast systems, but these systems still have limitations with respect to metabolism of most promutagens. This latter issue has been partially addressed by the development of cell lines that have been transfected with specific P450 isozymes to provide key metabolic activation steps.      
Most of these assay systems also use immortalized cell lines, either derived from tumors or transformed to an immortalized phenotype by a viral or other genetic change. These cellular systems, being simple, uniform monolayers, also lack the multicellular, three dimensional aspects of whole animal tissues, and also lack other pharmacokinetic and pharmacodynamic properties of intact animals. Nonetheless, these various assays have provided a great deal of information about the mutagenic potential of hundreds of chemicals to establish a growing database of information.      
Recent development of transgenic mice and rats (e.g., the "Big Blue" mice and rats) has allowed the screening of mutations in shuttle vectors following in vivo exposure, which largely alleviates the problems of cell culture systems. However, these systems still focus predominantly on single point mutations and selectable markers. In vivo systems that can look at other mutational events in vivo are limited but include dominant lethal, heritable translocation, and mouse spot test and specific locus assays. However, these systems are time-consuming, expensive, and have low throughput.      
The use of diverse genetic toxicology assays over the past two decades has provided a database of several hundred chemicals that are positive for DNA damage and/or point mutations. However, we still have little mechanistic insight into how chemicals might cause or influence the larger mutations that are the hallmark of human clinical tumors, and this remains an important goal. Vogelstein and co-workers have described a genetic model for colon cancer that is likely to be a relevant genetic paradigm for most cancers. According to this model, it requires four to seven distinct genetic changes in separate genes to progress from a normal colonic epithelium to a malignant and metastatic colon cancer. Although there appears to be a favored order to this process, these events do not strictly require a specific order of occurrence, but rather it is an accumulation of these genetic changes that results in cancer. Interestingly, the majority of these genetic events, and some of the key steps in the process, involve the inactivation of tumor suppressor genes rather than the activation of oncogenes.      
The precise mechanism for this inactivation is not well understood but is clearly critical to elucidate. It will be important in future studies to examine the genetic mechanisms underlying this process, e.g., focusing on how genotoxic and nongenotoxic chemicals can lead to genetic and phenotypic changes in cells that promote the carcinogenic process in addition to or in the absence of point mutations. In particular, a critical need remains for the development of moderate to high throughput molecular in vivo assays that can provide information on large mutations and other genetic events important in human cancers.


A. Chemical Carcinogenesis in Animals
Kenneway first demonstrated in the 1930s that a pure chemical could cause cancer in animals. Experimental evidence of tumorigenicity in animals has remained the gold standard for determining whether something is considered a potential human chemical carcinogen, particularly in the absence of strong human epidemiology data. The mouse or rat 2-year tumorigenicity study is currently the primary assay for experimentally assessing chemical carcinogenesis in animals for the purposes of human risk assessment. Generally, chemicals are administered at the maximum tolerated dose (MTD), and one-half the MTD for the lifetime of the animals, typically by gavage or in the diet, and tumor incidence is assessed in comparison to control animals. It is important to note that the background incidence of tumors in the control animals of these studies can be substantial. For example, in the two most widely used strains, B6C3F1 mice and F344 rats, the overall incidence of tumors can be as high as 50-60% and individual tissues can have a tumor incidence of 10-50%. Thus, a chemical must increase the tumor burden above this spontaneous background. Agents are considered positive that increase overall incidence, shift the overall tissue distribution or specific sites of tumors, or decrease the time to tumors.      
It is important to note that these long-term rodent tumor assays do not usually provide information about involved mechanisms. However, other short- and long-term experimental animal tumor assays can provide such information. The mouse two-stage skin cancer assay and similar assays developed in mouse lung and rat liver have provided a paradigm for assessing the basic mode of action of different chemicals. In the mouse two-stage skin cancer assay, four basic phases of carcinogenesis have been defined, initiation, early promotion (promotion I), late promotion (promotion II), and progression. Different chemicals can act during one or more of these phases and can be operationally defined by their mode of action in this model. For example, most genotoxic mutagens act as classic "initiating agents" in this assay, as a single application of one of these chemicals will usually result in increased skin tumors in the absence of any other treatment.      
A classic "tumor promoter," such as the phorbol ester, tetradecanoyl phorbol acetate (TPA), will usually not cause skin tumors by itself, but will substantially increase the tumor incidence of another agent if applied daily for several weeks or months following application of an initiating agent. Tumor promotion can experimentally be divided into an early and late phase in this system, the first of which appears to be reversible (tumor incidence decreases if chemical promotion is terminated) and the second of which is usually irreversible. The final phase of tumor progression is a period of time in which existing tumors progress from a nonmalignant to a fully malignant and metastatic phenotype. "Progressors" are chemicals that enhance this last phase of the process.      
"Complete carcinogens" are initiating agents that cause a substantial increase in fully malignant tumors in virtually all treatment animals, and therefore appear to be able to push cells through the entire process, without any additional help during the promotion or progression phases. A "cocarcinogen" in this assay is one that is not an initiating agent by itself, but enhances tumorigenesis when applied simultaneously with or prior to an initiating agent. Conversely, an "anticarcinogen" is one that can suppress the potency of another initiating chemical when given in combination. Likewise, other chemicals can be identified in this system that act as "chemopreventive agents" by being able to suppress the promotion and/or progression phases following initiation by another chemical. Much of our experimental evidence for the mechanism of action of chemicals as carcinogens is derived from these multistage tumor models.      
Another mechanism of action that has been described for some chemicals that induce tumors in long-term rodent assays appears to involve the induction of cell proliferation. Many agents that are only tumorigenic in animals at their MTD may be directly inducing cell proliferation or causing tissue damage leading to compensatory cell proliferation. In the case of chemicals that only act by this mode of action, it seems unlikely that this will also occur at the lower doses that humans are typically exposed to (often at thousands or millions of times lower than the MTD).       
Thus, there is some concern about determining the carcinogenic potential of some chemicals only at or near their MTD in these rodent assays. It is well known that cell proliferation is a strongly cocarcinogenic process. For example, in the rat liver, tumor induction by an initiator such as AFB1 can be substantially increased by giving the AFB1 in conjunction with a partial hepatectomy in which a large portion of the liver is surgically removed, leading to rapid proliferation of the liver to its fully restored size over the subsequent 72 h. Similarly, hepatitis B is strongly synergistic with AFB1 as a risk factor in human liver cancer, which is likely due to the continual proliferative response induced by this chronic viral infection.       
Conversely, it has been shown that mutagenesis from chemical and physical genotoxins can be strongly suppressed in mammalian cell culture by inhibiting cell division, presumably by providing an opportunity for cells to repair DNA damage before DNA replication, whereas the same treatment in cells that are rapidly dividing induces high levels of mutagenesis. Thus proliferation alone can be a strongly cocarcinogenic process, and agents or conditions that involve continual cell proliferation, such as chronic viral infections, other inflammatory responses, or chronic tissue injury, may strongly influence both background and chemically induced carcinogenesis. An additional mechanism of carcinogenesis that can be influenced by chemicals may involve disruption of cell-cell communication, which may be most important during the later phases of carcinogenesis when cells are acquiring a malignant and metastatic phenotype.      
Because of the large expense of long-term rodent tumor bioassays, many of the chemicals that have been screened to date were already strongly suspected of being carcinogenic in humans, based on human epidemiology studies or short-term assays such as those described in the preceding sections. The majority of these chemicals are genotoxic mutagen-carcinogens, as this class of agents is of considerable concern. However, more than a third of the chemicals that have been shown to be positive in long-term tumor assays are not overtly genotoxic or mutagenic. These nongenotoxic chemicals likely are acting through other epigenetic mechanisms as described earlier. There is concern about both false-positive and false-negative results in these assays. For example, there are agents that are positive in these long-term rodent assays but do not appear to be carcinogens in humans who are exposed to lower doses and different exposure conditions.      
Conversely, many chemicals that are known to be human carcinogens based on strong epidemiology evidence have been negative or equivocal in longterm rodent tumor assays (e.g., the carcinogenic metals arsenic and cadmium) or require nonphysiological exposures (e.g., intraperitoneal, intratracheal or implantation) to be positive (e.g., nickel and chromium). Thus, while animal tumorigenicity data are useful in assessing the carcinogenic potential and mechanism of action of many chemicals, there are limitations to these systems both for screening and for human risk assessment that are important to consider when evaluating individual agents.      
One of the more controversial uses of animal tumorigenicity data is in quantitative human risk assessment. This involves a mathematical extrapolation from the tumor incidence that is observed at high doses in rodents (often at only one or two doses at or near the MTD) to the potential cancer risk that might occur at the very low doses encountered by humans. This risk extrapolation from high to low doses may be required to extend over a dose range of three to six orders of magnitude. The conservative assumptions used by the U.S. Environmental Protection Agency and most other state and federal regulatory agencies that perform these risk assessments is that the dose- response will be linear over this range. This linear, low-dose extrapolation model was initially based on the so-called "one-hit" hypothesis, i.e., that one molecule of a chemical mutagen-carcinogen can theoretically interact with one critical target (i.e., DNA base) within a cell, thereby causing a change (mutation) that produces a cancer cell that can give rise to a tumor. We have little or no empirical data to support or reject this or other alternative models regarding the shape of the dose-response curve. However, as discussed earlier, it is likely that there are practical thresholds to the carcinogenic response to chemicals in vivo, and our current model of the multihit, multistage nature of the cancer process in humans suggests that the one-hit model is not a valid risk assessment model. Thus, it is likely that these risk assessments substantially overestimate risk at low doses. There has been a move toward the use of mechanistic information, when available, to do risk assessments of specific chemicals. For example, it appears that virtually all of the biological effects of TCDD can be attributed to its interaction with a cellular receptor, the Ah receptor, leading to activation of the receptor as a transcription factor. The Kd of TCDD for the Ah receptor in humans is known, and we would predict, based on basic receptor pharmacology principles, that there would be little or no biological effect of TCDD in humans at doses well below those that would be required to bind half or more of the available receptors. As additional mechanistic information such as this becomes available for individual chemicals, it will be increasingly possible to do meaningful, mechanistically based risk assessments that more accurately predict actual cancer risk to the human population in occupational and environmental exposure settings.

B. Evidence for Chemically Induced Cancers in Humans
There is substantial evidence from human epidemiology studies that chemicals can increase cancer in humans. Cigarette smoking alone is estimated to be responsible for approximately 30% of all cancer deaths, and this is clearly related to the hundreds of toxic chemicals in cigarette smoke, which collectively and individually have been shown to cause cancer in animals.      
Many of these chemicals are genotoxic mutagens, such as BaP, and others act as classic tumor promoters, comutagens, and cocarcinogens in the assays described earlier. Other major causes of cancer deaths include diet (35%), infection (10%), reproductive and sexual behavior (7%), and alcohol (3%) and, therefore, like tobacco use, are presumed to be largely preventable by identifying and instituting appropriate lifestyle changes. Human exposure to chemicals has been associated with cancer risk in environmental and occupational settings, but these exposures overall are estimated to contribute to only a small fraction of total cancer deaths. Approximately 4% of all cancer deaths have been attributed to occupational exposures, presumably involving exposures to individual chemicals and chemical mixtures in most cases. Similarly, exposure to pollution, industrial products, medicines or medical procedures, and food additives has been estimated to only account for an additional 2-4% of total cancer deaths. This is evident by examining data for death rates from major individual cancers over the past century. If occupational and environmental chemical exposures were responsible for a large and growing number of cancers, one might predict that cancer death rates should have increased from the 1950s onward, when there was a concomitant increase in manufacturing, use, and environmental release of industrial chemicals. However, with the exception of increases in male and female lung cancer from cigarette smoking, and a few other exceptions, most individual cancer death rates have been relevantly constant or have declined over the past century. Thus, the vast majority of cancers appear to be attributable to other causes. In particular, it is unlikely that exposure to trace levels of chemicals in the environment--in air, water, soil, as pesticide residues on food, and similar background exposures--contribute significantly to individual cancer risk, despite a public perception that this is the case.      
However, certain chemicals have been linked to human cancer in specific occupational or environmental settings. Hill first described an association between tobacco snuff and nasal polyps in 1761. Pott was the first to describe an occupational exposure of chimney sweeps to soot and their increased incidence of scrotal cancer. In the 1920s, the Germans described a strong association between occupational exposure to chromium dusts and an increase in lung and other respiratory tract cancers. This led to the development of industrial hygiene practices to reduce exposure as well as alterations in the manufacturing processes.      
Workers who began their employment in the chromium ore industries after the institution of these practices have demonstrated cancer risks similar to the general population. Occupational exposure to several other metals, particularly nickel, arsenic, and cadmium, has also been strongly associated with increased cancer risk, especially respiratory tract cancers. However, with the exception of arsenic (discussed later), there does not appear to be an increased cancer risk from exposure to the much lower environmental levels of these metals, particularly via noninhalation routes of exposure.      
Epidemiological studies of past occupational exposures have provided the best evidence for a link between individual chemicals and human cancer. Groups of workers in the late 19th and early 20th century who were exposed to 2-naphthylamine during its purification demonstrated a very high incidence of bladder cancer, some up to 100%. Similarly, occupational benzene exposure has been strongly associated with an increased incidence of acute myelogenous leukemia. Occupational exposure to vinyl chloride, an important chemical used in the manufacture of plastics, has been associated with an increased risk of liver angiosarcoma. This increased risk is fairly low even in exposed workers; however, the relative rarity of this particular cancer allowed its increased incidence to be detected. The relationship between increased human cancer risk and occupational exposure to other chemicals has been more controversial. This includes agents such as phenoxy herbicides and their contamination products (including TCDD), formaldehyde, and several organic solvents. The various conflicting studies published to date for some of these agents suggest that if there is an increased risk, it may be relatively low even in occupational settings.      
Occupational exposure to asbestos has been clearly associated with an increased risk of lung carcinoma and mesothelioma. There appears to be a strong synergy between asbestos and cigarette smoking for the former neoplasm. In fact, asbestos alone appears to be a weak or nonsignificant risk factor for this cancer in the absence of smoking, whereas asbestos exposure alone is clearly associated with mesothelioma risk and smoking appears to play a smaller or inconsequential role in this disease. This illustrates a common issue in both occupational and environmental exposures, namely, that humans are exposed to many agents simultaneously.      
Moreover, these agents can act additively, synergistically, or even antagonistically in contributing to overall cancer risk. Wood dust exposure, especially in the furniture manufacturing industries, is associated with an increased risk of lung cancer and other lung diseases and exhibits an interaction with smoking. The mechanistic basis for increased cancer risk from wood dust is not known, but may involve both a physical component (small, respirable fibers) and one or more specific chemical components.      
Use of certain drugs in medical therapies has also been associated with an increased risk of cancers. The most well-known example of this is the use of various DNA-damaging agents in cancer chemotherapy. By the nature of their mechanism of action, it is predicted that the majority of these drugs will have carcinogenic potential. The therapeutic goal when using these agents is to saturate the more rapidly dividing target tumor cells with sufficient DNA damage to arrest cell division and induce cell death. However, other normal dividing tissues are also receiving considerable DNA damage during systemic treatment with these agents, and surviving nontarget cells would be expected to have increased mutations that may contribute to carcinogenesis in these tissues, giving rise to so-called "second site" neoplasms. Ironically, because of the latency period of these tumors, this is usually only observed in long-term, disease-free survivors of the initial treatment. For example, in children with cancer, approximately 3-12 percent of survivors will develop second site cancers by 25 years after treatment. In adults, the risk of second site cancers is highly dependent on initial tumor type and mode of treatment. The greatest risk is for survivors of Hodgkin's disease, in which there is a 10-15% risk of second site tumors at 15 years after therapy. Not surprisingly, the risk of second site tumors is highest for those cancer patients receiving both radiation therapy and chemotherapy. A few other drugs in noncancer therapeutic treatments have also been shown to increase cancer risk. Use of diethylstilbestrol (DES) is a well-known but unusual example of hormonally induced cancer, where the adult female offspring of DES-treated mothers have an increased risk of clear cell vaginal adenocarcinoma. Other hormone therapies have also been associated with increased cancer risk, especially liver cancer risk associated with the use of estrogens in premenopausal women. Phenacatin use has been associated with an increased risk of renal carcinoma, and use of the immunosuppressive drug azothioprine has been associated with an increased risk of lymphoma, skin cancer, and Kaposi's sarcoma.      
Environmental exposure to certain chemicals is associated with an increased cancer risk. Exposure of certain populations to AFB1 through contaminated diet is clearly associated with an increased risk of liver cancer, although as mentioned earlier, the risk is synergistically greater with simultaneous infection by hepatitis B virus. AFB1 exposure is endemic in tropical areas of Africa, China, and South and Central America. Regulation of AFB1 levels in the food supply in the United States and the Western world has largely eliminated this risk to those populations. Environmental exposure to arsenic in drinking water in various regions throughout the world is also an important factor in the increased incidence of several cancers, as well as an increased risk of vascular disease and type 2 diabetes. Exposure to arsenic in drinking water occurs primarily as a result of leaching of arsenic from natural geological sources into well water. Arsenic is odorless, tasteless, and colorless and is therefore usually undetectable without chemical analysis. Certain areas of South America, southeast Asia, Europe, Asia, and North America contain appreciable arsenic levels in groundwater. Long-term exposure to arsenic through drinking water has been associated with elevated incidences of skin, lung, bladder, liver, kidney, and other cancers and can approach 10-fold above control levels in some areas.      
Chemical carcinogens that are associated with lifestyle include those in tobacco products, Betel nut chewing, and consumption of alcoholic beverages. As mentioned previously, dietary factors have been associated with up to 35% of all cancers. Many of these "dietary factors" may involve specific chemicals in the diet, apart from contaminants such as AFB1 or arsenic. These may include certain components of fats or fatty acids; various specific chemical inducers or inhibitors of cytochrome P450s, such as some of the natural plant chemicals found in dark green vegetables; heterocyclic amines and other food mutagens, especially those produced by cooking; polycyclic aromatic hydrocarbons produced by charring of meat and other foods; and nitrosamines found in certain foods.      
Conversely, certain plant chemicals may act as chemopreventive agents, such as some of those identified in certain fruits, vegetables, and teas. However, while it may be possible to optimize our diets to achieve the best balance between factors that enhance and sup- press carcinogenesis, we may be limited in the extent to which this is possible because virtually all foods appear to contain both types of agents. However, specific chemicals are being identified from the diet that may be useful pharmacologically in pure form as chemopreventive agents.


Experimental carcinogenesis and human epidemiology studies have clearly identified specific chemicals that can act as human carcinogens. Certain chemicals have been associated with increased human cancer incidence in both occupational and environmental exposure settings. Experimental cell and animal systems have been useful in helping identify potential human carcinogens, as well as in determining their basic mechanisms of action. Many of the chemicals that are known carcinogens act by causing covalent DNA damage that can lead to mutations in critical oncogenes and tumor suppressor genes, which in turn can contribute to the carcinogenic process. However, many other chemicals initiate or promote carcinogenesis through non-genotoxic mechanisms that may also be important, including cell proliferation and disruption of cell-cell communication. Moreover, chemicals that are identified in experimental systems as being positive for a particular end point such as DNA damage, mutations, cell transformation, and animal tumor formation, while having the potential to be carcinogenic, may not represent a major cancer risk in humans due to differences between these end points and the actual cancer process or limitations in the assay systems that do not allow direct comparisons with human exposures. In this regard, it is difficult to do meaningful human risk assessment when one must extrapolate from high-dose animal exposures to the more typical low human exposures using theoretical models that are based on unproven hypotheses and assumptions. There is a clear need to understand better the mechanisms of action of nonovertly genotoxic carcinogens. It will be important to determine the mechanistic basis for the induction of large clastogenic mutations and aneuploidy events, which are a hallmark of human cancers, and to develop effective screening assays for assessing the potential of chemicals to induce these types of genetic events.

Joshua W. Hamilton
Dartmouth Medical School

See Also

carcinogen Any chemical or physical agent that increases cancer burden by increasing the incidence, altering the tissue distribution, increasing the malignant or metastatic potential, or decreasing the latency period of cancers in an individual or a population.

genotoxin Any chemical or physical agent that directly or indirectly causes DNA damage, i.e., a covalent chemical modification to a DNA molecule.

mutagen Any chemical or physical agent that directly or indirectly leads to a heritable alteration in the genetic sequence of bases in DNA.

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