Environmental Factors

Evolving Models for Chemical Carcinogenesis

Fig. 10.1 gives a linear representation of the traditional paradigm for progressive carcinogenesis. This model is useful in that it depicts what is thought to be the monoclonal nature of cancer (originating from a single cell) and the concept of clonal evolution and expansion. It evokes a chemical- or radiation-provoked initiating event, followed by promotion of clonal expansion to form a preneoplastic lesion. In this model, it is thought that a single cell of a preneoplastic lesion undergoes “conversion,” which theoretically equates to progeny cells acquiring enough cumulative genotypic and phenotypic alterations to express a malignant phenotype. Malignant cells, defined pathologically as “invasive,” can then undergo progression (volume expansion due to increased net cell proliferation) to a clinically relevant cancer with metastatic potential. This pictorial sequence is often punctuated with reference to mutagenic activation of oncogenes, inactivation of tumor suppressor genes, and, more recently, epigenetic alterations. Such a linear model is particularly illustrative of protective points of intervention beginning with avoidance of disease-consequential exposure and chemoprevention (primary prevention), early diagnosis and cancer interception (secondary prevention), and prevention of relapse and second cancers (tertiary prevention). It also helps give temporal perspective to the carcinogenic “lag” period, the presumptive period of time between an initiation event and clinical detection of a tumor.

A consequence of this sequential model is that it tends to narrowly relegate environmental carcinogens as mutagenic “initiating” agents, and certain other chemical agents as nonmutagenic “promoting” agents, and does not account for the increasing weight of evidence that the process of early carcinogenesis at the cellular level is highly nonlinear and better understood as a perturbation of molecular homeostasis resulting in accelerated evolution toward acquisition of any of the 10 hallmarks of cancer. As shown in Fig. 10.2 , these hallmarks are best graphically portrayed in circular fashion because they do not necessarily follow a temporal sequence. This model elaborates our understanding of oncogenesis as a disruption of highly interconnected and complex signaling networks that intersect cytostasis and differentiation circuitry, cell viability circuitry, proliferation circuitry, and motility circuitry. It necessitates broadening our understanding of mechanisms of action in environmental carcinogenesis to include the potential ability of chemical carcinogens to provoke genomic instability, sustained proliferative signaling, acquisition of death resistance, evasion of growth suppressors, enabling of replicative immortality, and/or activated cellular motility and invasiveness.

The “hallmarks of cancer” model also underscores the possible impact of environmental agents on deregulation of cellular energetics, avoidance of immune destruction by emerging tumor cells, promotion of tumor inflammation, and provocation of nonneoplastic additional cells in the tumor microenvironment, such as stromal cells, to contribute to the survival, development, and emergence of an early precancerous cell. One illustrative example of this complexity emerges from increased understanding of the role of cell death in cancer and carcinogenesis. Studies of genotoxin-treated cells in culture have drawn attention to the importance of an overlooked outcome in earlier attempts to measure mutagenesis as a surrogate marker for carcinogenic potential: the frequent emergence of death-resistant subclones of cells out of which an occasional cell carrying a mutated selectable gene could be detected as a low-frequency event. This carcinogen-provoked early acquisition of resistance to apoptosis has challenged the perspective that active cell death processes necessarily protect against cancer and carcinogenesis. To the contrary, the facile ability of cells to circumvent cell death when under stress actually renders the existence of such cell death signaling pathways as a risk factor for carcinogenesis. Indeed, it is now appreciated that additional modes of cell death, such as autophagy, can mediate either tumor cell survival or death; and even necrosis, often a consequence of toxic exposure, has been shown to be potentially proinflammatory and tumor promoting. In 2015 a series of review articles were published by an international consortium of scientists, collectively called the Halifax Project, which conducted detailed assessments of the potential of environmental chemicals to contribute causatively to the process of carcinogenesis by affecting each “hallmark” of cancer.

History of Environmental Carcinogenesis and Support for the Role of Environmental Agents in the Etiology of Human Cancers

The methods and concepts for identifying chemical agents potentially carcinogenic to humans have evolved over time and continue to evolve as both the general public and regulatory agencies grapple with the fact that humans live in a chemical-laden society, which generally contributes to improved global health and increased longevity. In addition to consumption of and exposure to tobacco smoke, itself containing more than 30 potentially carcinogenic chemicals ( Box 10.1 ) delivered directly by inhalation of a chronically injurious toxic fume, humans are awash in a sea of low-level exposure to both natural and anthropomorphic chemicals. For example, polycyclic aromatic hydrocarbons (PAHs), a broad class of chemicals including some with carcinogenic potential, are released into the environment by both anthropomorphic fuel combustion and natural volcanic eruptions and forest fires, as well as consumed through smoking and eating flame-cooked meat. Such exposure complexity necessitates improved differentiation of “any” exposure from “disease-consequential” exposures, which itself represents a next-generation evolution of carcinogen identification from historical observational studies, to occupational cohort epidemiology, to broad-scale screening and testing in experimental animals, to exposome approaches of integrating sophisticated individual exposure monitoring with highly sensitive “omics” approaches to detect perturbation of disease-relevant signaling pathways.

The carcinogenic effects of a sizable number of environmental or industrial chemicals were first described observationally in humans. The influences of occupation and lifestyle in cancer occurrence trace back at least to the 16th century when Ramazzini, in 1700, noted that nuns showed a higher frequency of breast cancer than was observed among other women. Also in that century, Paracelsus and Agricola described Bergkrankheiten (lung cancer) in German miners, probably caused by uranium and its decay product radon. In 1761, Hill observationally associated the use of tobacco snuff with cancer in the nasal passage, and in 1775 Pott noted the occurrence of soot-related scrotal cancer among chimney sweeps. In 1895, Rehn published evidence that occupational exposure to aromatic amines was associated with bladder cancer, and Unna in 1894 associated sunlight exposure with skin cancer.

These observational associations gradually gave way to more proactive approaches to attempt to identify potential carcinogens both computationally (retrospective epidemiology, usually of occupationally exposed or catastrophe-exposed cohorts) and by animal experimentation. However, it was not until the early 20th century that animal models for chemical carcinogenesis were developed, beginning in 1915 when Yamagiwa and Ichikawa demonstrated the production of skin tumors after topical application of crude coal tar to the ears of rabbits. These early studies gradually led to the standardization of animal carcinogenesis bioassays, with the goal of testing chemical class prototypes for carcinogenic potential, ostensibly before human exposure.

Contrary to experiences in earlier centuries, with the advent of animal bioassay programs, evidence of carcinogenicity in experimental animals has sometimes preceded evidence obtained from epidemiologic studies or case reports. Although the term carcinogen means “giving rise to carcinomas,” loosely referring to malignancies in general, more operational definitions are used for carcinogens in animal bioassays. In this context, a carcinogen may be defined as an agent whose administration to previously untreated cells or animals leads to a statistically significant increased incidence of malignant neoplasms, compared with the incidence in appropriate untreated control cells or animals. Data from such studies, combined with epidemiologic and mechanistic studies, are employed by regulatory agencies to try to protect humans from known chemical, radiation, and infectious hazards. From a mechanism of action (MOA) perspective, the regulatory thrust has been to determine whether a carcinogenic agent was mutagenic or nonmutagenic; the latter involved recognition that chronic tissue toxicity itself, in the absence of DNA damage, was potentially carcinogenic. Synthetic and naturally occurring chemicals comprise the largest group of known human carcinogens. More than 100 chemicals, chemical mixtures, biologic agents, physical agents, or industrial processes have been classified as Class 1 human carcinogens ( Table 10.1 ) by the International Agency for Research on Cancer (IARC), and more than 300 chemicals have been designated as probable (Class 2A) or possible (Class 2B) human carcinogens. For perspective, these numbers of potential carcinogenic hazards derive from an environmental milieu of nearly 10 million chemicals.

Substantial growth has occurred in our understanding of how these extrinsic factors (chemicals, radiation, infectious agents) interact with intrinsic factors (e.g., genetics of inherited genes, “omics” profile, diet, body mass index (BMI), hormones, immune status, and microbiome) to determine overall susceptibility and risk. Indeed, a key concept underpinning the role of the environment in cancer risk relates to the impact of geography on lifestyle adoption and adaptation and cancer susceptibility, and derives from the following series of epidemiologic observations:

• Although the overall incidence of cancer development is reasonably constant among countries, incidences of specific cancer types can vary up to several hundredfold.

• Large differences in tumor incidence exist within populations of a single country.

• Migrant populations assume the cancer incidence of their new environment within one to two generations.

• Cancer rates within a population can change rapidly.

It should be noted, however, that a major aspect of geography-influenced carcinogenesis is related to exposure to unique dietary and lifestyle behaviors and infectious agents, and the possible additive or synergistic interactions between infectious agents and diet. For example, the incidence of cancer of the esophagus, the sixth most common cause of death from cancer worldwide but the third most common in developing countries after stomach and lung cancer, varies from less than 5 per 100,000 persons per year in Central America and Western Africa to more than 50 per 100,000 persons per year in parts of China and central Asia. Putative causes for such “hot spots” have been identified, such as consumption of hot yerba maté in Uruguay, Brazil, and northeast Argentina; however, the causative factors in other high-risk areas remain unknown. Other cancers demonstrating unique geographic distribution include biliary cancer (cholangiocarcinoma) associated with liver fluke infection in some Asian (especially Thailand) and African countries; gastric adenocarcinoma associated with virulent strains of Helicobacter pylori infection in South Korea, Japan, and parts of China; and hepatocellular carcinoma (HCC) associated with hepatitis B virus (HBV) and aflatoxin exposure in parts of western Africa and eastern China. Other connections between geography and lifestyle/environment are documented by studies of shifting cancer organ-site rates over generations following immigration, such as the decrease in stomach cancer rates and increase in prostate cancer rates in offspring of Japanese immigrants to North America.

Exposure Biomarkers and Assessing Human Exposure

Increased understanding of the mechanistic basis of carcinogenesis provides opportunities for the identification of molecular biologic markers that reflect both disease-relevant exposure and/or events occurring between exposure and clinical disease. It is hoped that the use of biologic markers will help define the roles of environmental agents (particularly at low levels and in complex mixtures) in the etiology of human cancer. These molecular biologic markers can be classified into three major categories:

• 1.

  Markers of exposure reflecting a biologically effective dose of a carcinogen

• 2.

  Markers of effect indicating a disease-relevant biologic response to an exposure

• 3.

  Markers of susceptibility that characterize the inherent susceptibility of an individual to a carcinogenic agent

The detectable persistent interaction of a carcinogen with macromolecules was first demonstrated by Miller and Miller in 1947, who showed azo dye residues bound to liver proteins of treated rats. Later studies indicated the importance of carcinogen modification of DNA (either directly or after metabolic activation) in the cancer process. The measurement of carcinogen metabolites, carcinogen-DNA adducts, or carcinogen-protein adducts in human tissues or fluids provides the basis for development and application of individual exposure monitoring, molecular dosimetry research, and the rapidly expanding field of “molecular” cancer epidemiology. The potential advantage of this approach is that more accurate assessments of individual or group dose may be achieved than through estimates of carcinogen exposure in the environment or workplace.

Carcinogen-DNA and carcinogen-protein adducts have been detected in tissues from a variety of human populations with known or suspected carcinogen exposure. PAH-DNA adducts are elevated in white blood cells from persons occupationally exposed to airborne PAHs, including coke oven workers, foundry workers, and aluminum plant workers, and in lung tissue from heavy smokers. DNA isolated from exfoliated bladder epithelial cells of cigarette smokers has been shown to contain 4-aminobiphenyl adducts. Alkylation damage is detected in DNA from esophageal tissue of persons with documented dietary exposure to nitrosamines. Aflatoxin adducts are found in hepatic DNA after dietary exposure to this mycotoxin. Cisplatin-DNA intrastrand adducts have been measured in white blood cells of patients undergoing cancer chemotherapy.

Carcinogen-DNA adducts excreted in urine provide a potentially noninvasive means for quantifying DNA damage. Urinary concentration of aflatoxin-guanine adducts is strongly correlated with dietary aflatoxin intake. In addition to their use as indicators of previous carcinogen exposure, DNA adducts have the potential for use as direct indicators of future cancer risk. The first prospective test of this application appeared in 1992, when detectable levels of aflatoxin-guanine adducts and several other aflatoxin metabolites in urine were shown to be predictive of the development of liver cancer.

Carcinogen-protein adducts have also been studied as biomarkers of human exposure. Alkylation damage in hemoglobin has been used as an exposure index in risk assessment analyses of persons occupationally exposed to ethylene oxide. It has been found that 4-aminobiphenyl adducts in hemoglobin are highly specific (and sensitive) markers of exposure to cigarette smoke. The level of 4-aminobiphenyl hemoglobin adducts is related to the number of cigarettes smoked, the type of tobacco smoked, and the metabolic phenotype of the smoker. The levels of these adducts drop markedly after smoking cessation.

Unreacted carcinogen metabolites excreted in urine also are used to monitor human exposure to and uptake of carcinogens and related compounds. Concentrations of hydroxylated PAHs (e.g., 1-hydroxypyrene) in urine are elevated in smokers, in patients after topical treatment with coal tar, in road pavers, in coke oven workers, in aluminum plant workers, and in persons ingesting PAHs from food. In occupational settings, the concentration of urinary 1-hydroxypyrene is highly correlated with estimated or measured concentrations of airborne PAHs.

These noninvasive approaches to exposure assessment have potential applications as biomonitoring tools, but at present they have major limitations, and by themselves do not address the key question of parsing out disease-relevant exposure. In all of these studies, significant differences in adduct or metabolite levels are often observed between persons with similar carcinogen exposure, likely due to individual biologic variability, exposure misclassification, and confounding variables such as diet, physical activity, smoking, or personal environment. It is hoped that the exposome approach, at the intersection between high-sensitivity biomarker detection and perturbation of disease-relevant signaling pathways, may improve capabilities for detection of meaningful exposures as a function of individual susceptibility.