documents from regulatory agencies and other organizations may be avail-able. However, it is important to consider the focus and goals of the authors for any particular document, because frequently the analysis is not aimed at providing for a causality assessment in an individual. In fact, usually such documents are intended to provide insights to protecting the public health.
As such, the authors provide assessments that will focus on safety, and utilize data that will maximize such.
exemplified by inheritance of a predisposition to retinoblastoma and=or osteosarcoma, where patients possess a homozygous loss of the Rb1 gene on chromosome 13 (1–3). In the familial form, loss of one allele is inherited and the other is lost through later mutation. Loss of suppressor and antime-tastasis genes can be an early or late event involving several steps, including angiogenesis and metastasis (4).
Another way of considering genes involved in the carcinogenic process is to classify them as caretaker and gatekeeper genes (5). This recognizes their respective roles in the maintenance of genomic integrity (e.g., DNA repair) and cellular proliferation, respectively. Landscaper genes are also considered that are responsible for maintaining the general environment around the cells (i.e., effects on stromal cells and signaling between cells).
Some examples of caretaker genes are those that are involved in DNA repair and carcinogen metabolism, while examples of gatekeeper genes are those involved in cell cycle control and DNA replication. Dysfunctional caretaker genes increase the probability of mutations in gatekeeper genes, which are necessary to initiate the molecular pathogenesis of cancer. It is interesting that the carcinogenic effects of caretaker and gatekeeper genes appear to be tissue-specific and lead to cancer only in those organs, even though these genes are expressed in many different organs.
Carcinogenic agents affect DNA in different ways. They can cova-lently bind to nucleotides and form adducts. These adducts may be promu-tagenic, and if present at the time of DNA synthesis, can cause mutations.
N-nitrosamines, for example, present in the diet and linked to esophageal and gastric carcinoma, result in nucleotide base substitutions due to mispairing at sites where adducts are formed. Some mutations may reflect specific carcinogen exposures or endogenous mechanisms, and frequently exhibit target organ specificity. For example, p53 mutations at codon 249 frequently observed in hepatocellular carcinoma from China (6) or southern Africa (7) are consistent with the type of damage caused by aflatoxin B1
exposure, a common dietary carcinogen linked to this tumor. In contrast, several types of p53 mutations have been observed in lung cancer (8), which is consistent with a multiple carcinogen exposure etiology from tobacco smoke. To date, however, the mutational spectra of p53 or other genes cannot be used to determine what caused a cancer in a person.
Independent of carcinogen exposure, human cells are continuously undergoing spontaneous mutations at a low rate. Oxidative damage, poly-merase infidelity, chromosomal rearrangement, recombinase infidelity, and telomere reduction are other sources of error. The process of cell and DNA replication can increase the rate of mutation (9). When one considers that the human body contains 1014 cells and that these cells undergo 1016 divisions over a person’s life span, it is quite possible that genomic instabil-ity plays an important role in carcinogenesis (10,11).
Balancing the ongoing exposure to carcinogens, cells have the ability to repair DNA damage, such as the removal of carcinogen DNA adducts through nucleotide excision repair or gross chromosomal damage through homologous recombination. When DNA is sufficiently damaged and cannot be repaired, then cells have the ability to trigger apoptosis (cell death), in which case it is no longer viable and cannot become a cancer cell.
In almost every step of the multistage process of carcinogenesis, person-to-person differences in cancer susceptibility can be found (12).
Interindividual differences for particular traits can be acquired, or inherited.
Inherited susceptibilities, manifest through evolutionary changes in nucleo-tide sequence, may augment human cancer pathogenesis that can vary from high penetrance with an attendant high likelihood of causing cancer to low penetrance genes with an attendant increased risk of causing cancer albeit less likely than high penetrance genes. Nevertheless, the range from low to high penetrance genes is a continuum, and studies in animal models indicate that the effects of high penetrance genes can be modified by other genes (13).
High penetrance genes that cause family cancer syndromes can have sub-stantial impact in affected families (e.g., BRCA1, hereditary nonpolyposis coli or Li–Fraumeni syndrome) (13), but they affect only a small percentage of the population. In contrast, the manifestations of cancer susceptibility genes with less penetrance contribute to common sporadic cancers, and thus affect a large segment of the population.
2.1. Methods to Assess Carcinogenesis
There are many tests that are used to increase our understanding of carcino-genesis and risk; these range from in vitro and experimental animal in vivo studies to human clinical and epidemiological studies. The usefulness of each method can be contrasted with its limitations (Table 1). Short-term assays for mutagenesis provide quick and inexpensive screens for potential carcinogens. Among the most widely used is the Ames’ test (14), which assesses mutagenic potential in Salmonella typhimurium bacteria. The Ames’
test has also been used as a biomonitor in humans, as exemplified by urine mutagenicity studies from cigarette smokers (15). While other assays exist, there are none with proven increased predictive value. Although short-term assays are useful in identifying potentially carcinogenic compounds, the same sensitivity makes the results difficult to extrapolate to humans. Posi-tive results might be unique to the strain, and factors such as metabolism, repair, and exposure cannot be assessed.
Experimental animal studies provide a short-term ability to assess the effects of a carcinogen in mammals. However, the predictivity for human risk is poor, but few better methods to study possible cancer risk and carci-nogenesis exist. As used by the National Toxicology Program and others, rodent carcinogenicity studies are performed using lifetime exposures with
up to maximally tolerated doses (MTD), that is, those not producing clini-cally evident toxic effects. To infer that a carcinogenic effect is present in laboratory animals, dose–response relationships are examined, along with overall mortality rates and consistency with other species. The limitations in these experiments include the routine use of the MTD, thus potentially increasing cell replication with resultant increased endogenous mutations;
interspecies, and interstrain differences; use of rodents known to have high spontaneous rates of cancer; an inability to account for metabolic differ-ences between high- and low-dose exposure, and difficulty in interpreting data from doses that commonly exceed those received by humans (16–
18). Also, the tumors of experimental animals may not resemble human cancer, or may not be malignant.
Human investigations provide the most relevant data regarding human risk. Clinical studies might be done, where exposures are unavoid-able. For example, while it is conceivable to intentionally expose a person Table 1 Testing for Carcinogenicity
Method Advantages Disadvantages
In vitro testing Economical Uncertain in vitro to in vivo extrapolations
Rapid results Frequent false positives and negatives
Human cells can be used
Mutagenicity is not the same as carcinogenicity
Inter-laboratory variation Animal bioassay More predictive of
human experience than short-term tests Elucidates species
differences
Expensive
Doses are higher than those experienced by humans Uncertain animal-to-human
extrapolation Human clinical
studies
Direct measurement of human experience
Cancer is not an endpoint Biomarkers show
biologically effective dose and intermediate markers of cancer risk
Short-term results Epidemiology Direct measurement of
human experience
Insensitive
Does not prove causation Covariables examined
Dose–response data Unknown confounding variables Biomarkers can be used
Interindividual variation considered
who already smokes to a modified tobacco product intended to reduce exposure, it would not be advisable to expose a person to a new chemical that has not been considered to be safe through laboratory testing. Epide-miology measures the incidence or prevalence of disease in human popula-tions. One limitation is that epidemiology can inform us about cancer risk from prior exposures, the latency effect of most carcinogens is so long that we cannot wait to assess a cancer risk in the future, and these study repeat effects after too many people get cancer. Also, it must be realized that epi-demiologic methods, by themselves, do not demonstrate causation. The assessment of causation can be aided by Sir Austin Bradford–Hill’s (19) proposed criteria, summarized in Table 2.
A formal quantitative risk assessment using mathematical models is used by regulatory agencies to estimate a potential cancer risk to a popula-tion exposed to a particular carcinogen at a specific dose. Risk assessments serve public health interests as they attempt to predict the frequency of cancer in a population before epidemiologic investigations can be performed, that is, before significant exposure and adverse outcomes occur. Among the reasons that population risk assessment informs little understanding about individual cancer risk, or causality, is that the risk assessment process makes a variety of interpretations and assumptions in the public health interest, and many of these are open to debate. Examples include subjective evaluations of the literature and extrapolations from laboratory animals to humans, and the use of safety quotients to compensate for a lack of knowledge in some areas.
2.2. Gathering Risk Factor Information
The initial approach before considering the individual’s situation and pos-sible risk factors is to assess what risk factors are known, and the scientific basis that identifies these risk factors. In the case of a potential carcinogen exposure, the scientific data are considered at any level of exposure. This Table 2 Evaluating Cancer Etiology—Bradford–Hill Criteria
Criteria Explanation
Strength of association What is magnitude of risk?
Consistency Are there repeated observations
by multiple investigators in different populations?
Specificity Is the effect specific or are there other known causes?
Temporality Does exposure precede effect?
Biological gradient Is there a dose–response relation?
Biological plausibility Is the effect predictable?
Coherence Is the effect consistent with other scientific data?
Analogy Do other similar agents act similarly?
Source: From Ref. 19.
is done by reviewing scientific textbooks and articles identified by doing computerized literature searches.
For literature searches, public websites such as the National Library of Medicine PubMed website can be useful (http:==www.ncbi.nlm.nih.
gov=entrez=query.fcgi). Abstracts of publications are frequently available, some articles can be downloaded for free, or at a cost, and all can be ordered through a related service on that site called Loansome Doc. Search strate-gies should be broad in order to identify related articles that might not be categorized appropriately. PubMed also includes links to related articles, which are sometimes helpful.
The websites for governmental agencies and other organizations can provide information, including downloads of various monographs. For exam-ple, one can view the websites of the Agency for Toxic Substances and Disease Registry (http:==www.atsdr.cdc.gov)= and its parent website for the Centers for Disease Control (http:==www.cdc.gov)=, the American Conference of Governmental Hygienists (http:==www.acgih.org), the Environmental Pro-tection Agency (http:==www.epa.gov), Food and Drug Administration (http:==www.fda.gov), the International Agency for Research on Cancer of the World Health Organization (http:==w=niosh), National Institutes of Environmental Health Sciences (http:==www.niehs.nih.gov),and the Nuclear Regulatory Commission (http:==www.nrc.gov). Also, one might look to links by various nonprofit and advocacy organizations.
2.3. Assessing Causality in the Individual
The methodology for the determination of cancer causality is described below. It is important to assess different types of scientific data, relying on the best studies. Sometimes, a researcher might postulate causality (i.e., as might be done through a publication of a case report, or a case ser-ies), but this is different from concluding a causal relationship of exposure to outcome. Among the types of data that might be useful, human epidemio-logical data are substantially more helpful than nonhuman data. If there is sufficient reason to consider that the chemical has a potential to cause the type of cancer identified for the individual (target organ specificity is important), then an assessment is made to determine the doses reported in the literature that may be associated with an increased cancer risk, and in what settings. A mechanistic understanding of the carcinogenic process (known or hypothesized) is considered in the context of the alleged exposure and disease in the patient. Animal and in vitro studies can be helpful in these mechanistic assessments. A concurrent step for assessing causality in an individual is to confirm the diagnosis, as sometimes incorrect diagnoses are made, and so would not be appropriately related to the alleged exposure in the individual. Assuming that there is sufficient reason to believe that the chemical might increase cancer risk, then one would consider the individual’s
potential exposure or risk factor compared to those from the literature. For example, exposure level, route of exposure, other exposures or risk factors, or the type of population would be considered.
It is helpful to consider the Bradford–Hill guidelines (19) mentioned above and described in Table 2. While not all are criteria are required to be met, there are some criteria that if violated would exclude the likelihood of causation; while fulfilling some may not lead to a definitive conclusion of causation. Among the most important criteria is consistency in the litera-ture, that is, doing several well-designed and well-conducted epidemiology studies leading to similar findings in different populations, using different study designs. It should be noted that no single epidemiological study is defi-nitive. A determination of a biological gradient also is important, i.e., if there is a dose–response relationship identified in scientific studies, and if those doses occur in the human exposure scenario of interest. Another is the strength of association, which allows one to consider if the reported association in an epidemiological study is believable (i.e., not too high or too low). An evaluation of temporality considers if the exposure sufficiently preceded the cancer effect to allow for latency. Specificity considers if the cancer has other reported causes and if the effect occurs in the identified tar-get organ. Coherence refers to an evaluation and agreement of different types of scientific data (epidemiological, laboratory animal studies, cell cul-ture models, etc.) and they do provide similar findings that lead to a mechanistic understanding of how the chemical would cause cancer in humans. Analogy looks to see if similar chemicals are known to behave similarly and what is the available scientific data for those chemicals.
2.4. Assessing the Patient
A careful history and physical examination are critical to any medical assessment, and that is true for cancer risk assessment too. The history, detailed in Table 3, needs to be thorough. What is critical is to document all potential exposures. Parenthetically, medical records are often used in litigation over potential exposures, and so the health care provider needs to document potential exposures accurately and clearly. This is true for known cancer risk factors as well.
If a known or suspected carcinogen is identified for a patient, then an evaluation of the actual exposure can be undertaken. If there are validated biomarkers for such, then these can be relied upon. Some might reflect only recent exposure, however, and do not indicate what has possibly occurred over many years or a lifetime. There is considerable research into the devel-opment and validation of biomarkers. The validation has to include the reliability of the test itself as it reflects what it is supposed to be measuring, but also its validity as a risk factor. The latter can be more complicated, and the health care provider should use caution when considering the use of a
test that is still experimental. A major limitation is that recommendations based on results cannot be given in an informed manner.
Environmental monitoring might be taken, and although some of these are relatively inexpensive (i.e., radon), the cost of some monitoring and testing can be prohibitive. The use of biomarkers or environmental test-ing must be carefully considered, includtest-ing their validity. The choice of laboratory, and its competency and experience also must be considered.
Resources for environmental testing might include local industrial hygienists Table 3 Assessing a Patient’s History
Category Examples of questions
Medical History of present illness
History of medical disorders associated with secondary malignancies
Recent and distant medication use History of radiation exposure History of virus exposure
Family History of cancer in different generations, including immediate and next-to-immediate members
Assess passive smoke exposure (parents, current occupants) Occupational history of current and past household members Hereditary disorders associated with secondary malignancies Social Tobacco consumption (cigarettes and smokeless products)
Alcohol use
Risk factors for viral exposure Substance abuse
All recreations and hobbies
Diet and nutrition, including vitamin use, health fads, home gardens and locally grown food products
Foreign travel
Occupational All occupations, including summer and childhood work Parental occupation
Any jobs with known hazards
Any jobs where protective equipment was used Any jobs with cancer clusters
Any jobs with bad odors
Any jobs with chemicals, fumes, gases, or dusts Environmental All residences and types
Residential proximity to industry, waste sites, agriculture, or other areas with potential exposure
Source of water—well, community, and bottled Cancer clusters
Use of pesticides, herbicides, and termiticides House building materials and renovations
at the company where an exposure might be alleged, or a consulting firm.
Public health departments might also be helpful.