Uncertainty Distributions for Cancer Potency Factors: Combining Epidemiological Studies with Laboratory Bioassays—the Example of Acrylonitrile

For eight chemicals or chemical mixtures with clear positive epidemiological evidence of carcinogenicity by inhalation (acrylonitrile, arsenic, benzene, beryllium, cadmium, chromium VI, coke oven emissions, and nickel), the United States Environmental Protection Agency (USEPA) uses that evidence to obtain a single best estimate of cancer potency factor. The methods used have so far been ad hoc, because of the differences in published studies, although there are common factors. In every case, the uncertainties involved in the various stages of analysis are qualitatively acknowledged, and often quantitatively estimated, but no formal attempt has been made to propagate the uncertainties. I here provide a detailed case study for acrylonitrile that (a) incorporates all estimates of uncertainty mentioned by the US EPA in their analysis and propagates that uncertainty to produce an uncertainty distribution; (b) updates the USEPA analysis to incorporate more recent epidemiological data from the same study used in their analysis.

For most of the materials known to be carcinogenic to humans (through epidemiologic evidence), there are also available cancer bioassays performed on laboratory animals. If the procedures used for estimating human carcinogenic potencies from laboratory animal bioassays are to be believed in cases where there are no human epidemiological data, their evidence should also be used where there is epidemiological evidence. A consistent method of incorporating the results of both epidemiological studies and laboratory animal bioassays into a single probability distribution for a human cancer potency is here detailed, using acrylonitrile as an example for which there is positive epidemiological data. The methods are sufficiently general to allow the incorporation of any combinations of positive and negative bioassay and epidemiological data.     

Sister chromatid exchanges induced in human lymphocyte chromosomes by trifluralin, atrazine and simazine

Many epidemiological and experimental "in vivo" studies have proved in recent years the carcinogenic properties of herbicides. In order to evaluate the "in vitro" action on the human DNA of Trifluralin, Atrazine and Simazine (active principles of herbicides Treflan and Fogard S respectively) the authors have studied the rates of SCE in cultures of human lymphocytes exposed to different concentrations of a solution 1 ppm of the substances. Trifluralin and Simazine, but not Atrazine, increase SCE per cell, with statistical significance, in the cultures with the highest concentrations of these substances. (SCE per cell: Trifluralin 5.27 +/- 1.38, Simazine 5.09 +/- 1.19, Control 3.51 +/- 1.14).     

The oncogenic action of nonfibrous mineral dusts]

Nonfibrous mineral dusts antigorite, basalt, cement, zeolite-klinoptilolite and gamma-alumina were tested for carcinogenic activity in rat experiments. Intraperitoneal injections of zeolite-klinoptilolite and gamma-alumina led to development of peritoneal mesotheliomas, whereas antigorite and cement had no carcinogenic potential. There is no differences in physicochemical and chemical properties between carcinogenically active and inactive nonfibrous dusts. A new class of carcinogenic substances is defined including basalt, zeolite-klinoptilolite and quartz which belong to nonfibrous mineral dusts.     

Carcinogenicity of the aromatic amines: from structure-activity relationships to mechanisms of action and risk assessment

Aromatic amines represent one of the most important classes of industrial and environmental chemicals: many of them have been reported to be powerful carcinogens and mutagens, and/or hemotoxicants. Their toxicity has been studied also with quantitative structure-activity relationship (QSAR) methods: these studies are potentially suitable for investigating mechanisms of action and for estimating the toxicity of compounds lacking experimental determinations. In this paper, we first summarized the QSAR models for the rodent carcinogenicity of the aromatic amines. The gradation of potency of the carcinogenic amines depended firstly on their hydrophobicity, and secondly on electronic (reactivity, propensity to be metabolically transformed) and steric properties. On the contrary, the difference between carcinogenic and non-carcinogenic aromatic amines depended mainly on electronic and steric properties. These QSARs can be used directly for estimating the carcinogenicity of aromatic amines. A two-step prediction is possible: (1) estimation of yes/no activity; (2) if the answer from step 1 is yes, then prediction of the degree of potency. The QSARs for rodent carcinogenicity were put in a wider context by comparing them with those for: (a) Salmonella mutagenicity; (b) general toxicity; (c) enzymatic reactions; (d) physical-chemical reactions. This comparative QSAR exercise generated a coherent global picture of the action mechanisms of the aromatic amines. The QSARs for carcinogenicity were similar to those for Salmonella mutagenicity, thus pointing to a similar mechanism of action. On the contrary, the general toxicity QSARs (both in vitro and in vivo systems) were mostly based on hydrophobicity, pointing to an aspecific mechanism of action much simpler than that for carcinogenicity and mutagenicity. The oxidation of the amines (first step in the main metabolic pathway leading to carcinogenic and mutagenic species) had identical QSARs in both enzymatic and physical-chemical systems, thus providing evidence for the link between simple chemical reactions and those in biological systems. The results show that it is possible to generate mechanistically and statistically sound QSAR models for rodent carcinogenicity, and indirectly that the rodent bioassay is a reliable source of good quality data.     

Mutagenicity, carcinogenicity and teratogenicity of aluminium

Aluminium and its salts, which are extensively used in the household and in industry, do not constitute a carcinogenic, mutagenic or teratogenic hazard, except, perhaps, in cases of extremely high exposure. The large majority of the experiments performed to assess the carcinogenicity of aluminium in laboratory animals gave negative results or even suggested some antitumor activity. Moreover, epidemiological studies have not provided clear evidence of a carcinogenic hazard of aluminium to man, and short-term tests made in vitro and in vivo to demonstrate mutagenic activity of A1 were negative except for some experiments in plants. The embryotoxic properties suggested by the studies on birds and mammals could result from the influence of A1 on phosphate and calcium metabolism or from interference with the polymerization of microtubules.     

Facilitative mechanisms of lead as a carcinogen

The carcinogenicity of lead compounds has received renewed attention because of continuing environmental and occupational sources of exposure in many countries. The epidemiological evidence for an association between lead exposures and human cancer risk has been strengthened by recent studies, and new data on mechanisms of action provide biological plausibility for assessing lead as a human carcinogen. Both epidemiological and mechanistic data are consistent with a facilitative role for lead in carcinogenesis, that is, lead by itself may not be both necessary and sufficient for the induction of cancer, but at a cellular and molecular level lead may permit or enhance carcinogenic events involved in DNA damage, DNA repair, and regulation of tumor suppressor and promoter genes. Some of these events may also be relevant to understanding mechanisms of lead-induced reproductive toxicity.     

Genotoxic potency of monofunctional alkylating agents in E. coli: comparison with carcinogenic potency in rodents

A quantitative correlation between carcinogenicity and genotoxicity was investigated by a comparison between the carcinogenic potency in rodents and the mutagenic (M), recombinogenic (R) and SOS-inducing (I) potencies in a bacterial test (E. coli multitest) for 9 monofunctional alkylating agents: N-nitroso-N-methylurethane, N-nitroso-N-ethylurea, epichlorohydrin, N-nitroso-N-methylurea, N-nitroso-N-methyl-N'-nitroguanidine, methyl methanesulfonate, diethylsulfate, dimethylsulfate, ethyl methanesulfonate. A significant positive correlation between the carcinogenic potency and the product of the mutagenic and recombinogenic potencies was found for all tested compounds. Thus, the E. coli multitest may be used as a simple test to search for correlations between carcinogenicity and genotoxicity of DNA-damaging agents.     

Lack of micronuclei formation in bone marrow of rats after repeated oral exposure to nickel sulfate hexahydrate

Workplace exposures to mixtures of nickel compounds have been associated with excess respiratory cancer risk. Animal studies with individual nickel compounds indicate that not all nickel substances have the same potency or potential to induce tumors. The bioavailability of nickel ions at critical cellular sites seems to be important to determine the potential of a substance to induce tumors in animals, but much less is understood about the exact nature (genotoxic or non-genotoxic) of the nickel effects. Within many regulatory frameworks (e.g., European Union), substances are classified for mutagenicity based on the available data and this classification will often influence the mode of action assigned to carcinogenic substances and the way in which risk assessment will be conducted. The objective of this study was to evaluate the ability of nickel sulfate hexahydrate to induce micronuclei in polychromatic erythrocytes (PCEs) in rat bone marrow. This study was conducted according to OECD and EU protocol guidelines. In the dose range-finding assays, the maximum tolerated dose was estimated to be 500 mg/kg/day. The doses used in the micronucleus assay were 125, 250, and 500 mg/kg/day. At least 2000 PCEs per animal were analyzed for micronuclei in PCEs. Cytotoxicity was assessed by scoring a minimum of 500 consecutive total polychromatic (PCE) and normochromatic (NCE) erythrocytes (PCE/NCE ratio). Nickel sulfate hexahydrate did not induce statistically significant increases in micronucleated PCEs at any dose examined. The negative results in the present study contribute significantly to the weight of evidence evaluation of the mutagenicity (chromosomal level) of nickel substances. These results are consistent with a non-genotoxic mode of action for soluble nickel that could explain the enhancement of cancer risk seen among refinery workers with mixed exposures and its lack of carcinogenicity in animal studies with single exposures.     

Prevalence of genotoxic chemicals among animal and human carcinogens evaluated in the IARC Monograph series.

To determine whether genotoxic and non-genotoxic carcinogens contribute similarly to the cancer burden in humans, an analysis was performed on agents that were evaluated in Supplements 6 and 7 to the IARC Monographs for their carcinogenic effects in humans and animals and for the activity in short-term genotoxicity tests. The prevalence of genotoxic carcinogens on four groups of agents, consisting of established human carcinogens (group 1, n = 30), probable human carcinogens (group 2A, n = 37), possible human carcinogens (group 2B, n = 113) and on agents with limited evidence of carcinogenicity in animals (a subset of group 3, n = 149) was determined. A high prevalence in the order of 80 to 90% of genotoxic carcinogens was found in each of the groups 1, 2A and 2B, which were also shown to be multi-species/multi-tissues carcinogens. The distribution of carcinogenic potency in rodents did not reveal any specific characteristic of the human carcinogens in group 1 that would differentiate them from agents in groups 2A, 2B and 3. The results of this analysis indicate that (a) an agent with unknown carcinogenic potential showing sufficient evidence of activity in in vitro/in vivo genotoxicity assays (involving as endpoints DNA damage and chromosomal/mutational damage) may represent a hazard to humans; and b) an agent showing lack of activity in this spectrum of genotoxicity assays should undergo evaluation for carcinogenicity by rodent bioassay, in view of the present lack of validated short-term tests for non-genotoxic carcinogens. Overall, this analysis implies that genotoxic carcinogens add more to the cancer burden in man than non-genotoxic carcinogens. Thus, identification of such genotoxic carcinogens and subsequent lowering of exposure will remain the main goal for primary cancer prevention in man.     

Update on genotoxicity and carcinogenicity testing of 472 marketed pharmaceuticals

This survey is a compendium of genotoxicity and carcinogenicity information of 838 marketed drugs, whose expected clinical use is continuous for at least 6 months or intermittent over an extended period of time. Of these 838 drugs, 366 (43.7%) do not have retrievable genotoxicity or carcinogenicity data. The remaining 472 (56.3%) have at least one genotoxicity or carcinogenicity test result. Of the 449 drugs with at least one genotoxicity test result, 183 (40.8%) have at least one positive finding. Of the 338 drugs with at least one carcinogenicity test result, 160 (47.3%) have at least one positive result. Concerning the predictivity of genetic toxicology findings for long-term carcinogenesis assays, of the 315 drugs which have both genotoxicity and carcinogenicity data 116 (36.8%) are neither genotoxic nor carcinogenic, 50 (15.9%) are non-carcinogens which test positive in at least one genotoxicity assay, 75 (23.8%) are carcinogenic in at least one sex of mice or rats but test negative in genotoxicity assays, and 74 (23.5%) are both genotoxic and carcinogenic. Only 208 (24.8%) of the 838 drugs considered have all data required by current guidelines for testing of pharmaceuticals. However, it should be noted that a large fraction of the drugs considered were developed and marketed prior to the present regulatory climate. Although the laws do not require re-testing based on revised standards, in the absence of epidemiological studies excluding a carcinogenic risk to humans, a re-evalutation would be appropriate.     

Evaluation of fecal mutagenicity and colorectal cancer risk

Colorectal cancer is one of the most common internal malignancies in Western society. The cause of this disease appears to be multifactorial and involves genetic as well as environmental aspects. The human colon is continuously exposed to a complex mixture of compounds, which is either of direct dietary origin or the result of digestive, microbial and excretory processes. In order to establish the mutagenic burden of the colorectal mucosa, analysis of specific compounds in feces is usually preferred. Alternatively, the mutagenic potency of fecal extracts has been determined, but the interpretation of these more integrative measurements is hampered by methodological shortcomings. In this review, we focus on exposure of the large bowel to five different classes of fecal mutagens that have previously been related to colorectal cancer risk. These include heterocyclic aromatic amines (HCA) and polycyclic aromatic hydrocarbons (PAH), two exogenous factors that are predominantly ingested as pyrolysis products present in food and (partially) excreted in the feces. Additionally, we discuss N-nitroso-compounds, fecapentaenes and bile acids, all fecal constituents (mainly) of endogenous origin. The mutagenic and carcinogenic potency of the above mentioned compounds as well as their presence in feces, proposed mode of action and potential role in the initiation and promotion of human colorectal cancer are discussed. The combined results from in vitro and in vivo research unequivocally demonstrate that these classes of compounds comprise potent mutagens that induce many different forms of genetic damage and that particularly bile acids and fecapentaenes may also affect the carcinogenic process by epigenetic mechanisms. Large inter-individual differences in levels of exposures have been reported, including those in a range where considerable genetic damage can be expected based on evidence from animal studies. Particularly, however, exposure profiles of PAH and N-nitroso compounds (NOC) have to be more accurately established to come to a risk evaluation. Moreover, lack of human studies and inconsistency between epidemiological data make it impossible to describe colorectal cancer risk as a result of specific exposures in quantitative terms, or even to indicate the relative importance of the mutagens discussed. Particularly, the polymorphisms of genes involved in the metabolism of heterocyclic amines are important determinants of carcinogenic risk. However, the present knowledge of gene-environment interactions with regard to colorectal cancer risk is rather limited. We expect that the introduction of DNA chip technology in colorectal cancer epidemiology will offer new opportunities to identify combinations of exposures and genetic polymorphisms that relate to increased cancer risk. This knowledge will enable us to improve epidemiological study design and statistical power in future research.     

Systematic review of the epidemiological evidence comparing lung cancer risk in smokers of mentholated and unmentholated cigarettes

Background

US mentholated cigarette sales have increased considerably over 50 years. Preference for mentholated cigarettes is markedly higher in Black people. While menthol itself is not genotoxic or carcinogenic, its acute respiratory effects might affect inhalation of cigarette smoke. This possibility seems consistent with the higher lung cancer risk in Black men, despite Black people smoking less and starting smoking later than White people. Despite experimental data suggesting similar carcinogenicity of mentholated and non-mentholated cigarettes, the lack of convincing evidence that mentholation increases puffing, inhalation or smoke uptake, and the similarity of lung cancer rates in Black and White females, a review of cigarette mentholation and lung cancer is timely given current regulatory interest in the topic.

Methods

Epidemiological studies comparing lung cancer risk in mentholated and non-mentholated cigarette smokers were identified from MedLine and other sources. Study details were extracted and strengths and weaknesses assessed. Relative risk estimates were extracted, or derived, for ever mentholated use and for long-term use, overall and by gender, race, and current/ever smoking, and meta-analyses conducted.

Results

Eight generally good quality studies were identified, with valid cases and controls, and appropriate adjustment for age, gender, race and smoking. The studies afforded good power to detect possible effects. However, only one study presented results by histological type, none adjusted for occupation or diet, and some provided no results by length of mentholated cigarette use. The data do not suggest any effect of mentholation on lung cancer risk. Adjusted relative risk estimates for ever use vary from 0.81 to 1.12, giving a combined estimate of 0.93 (95% confidence interval 0.84-1.02, n = 8), with no increase in males (1.01, 0.84-1.22, n = 5), females (0.80, 0.67-0.95, n = 5), White people (0.87, 0.75-1.03, n = 4) or Black people (0.90, 0.73-1.10, n = 4). Estimates for current and ever smokers are similar. The combined estimate for long-term use (0.95, 0.80-1.13, n = 4) again suggests no effect of mentholation.

Conclusion

Higher lung cancer rates in Black males cannot be due to their greater preference for mentholated cigarettes. While some study weaknesses exist, the epidemiological evidence is consistent with mentholation having no effect on the lung carcinogenicity of cigarettes.     

Mutagenic nitroarenes,diesel emissions,particulate-induced mutations and cancer: an essay on cancer-causation by a moving target

Initial analyses of the lung tumors seen in rats exposed for their lifetime to elevated levels of the emissions of diesel engines suggested that they were due to powerful mutagens and carcinogens (PAHs, nitro PAHS) adsorbed onto the diesel particles. However, further studies showed that carcinogenicity occurred only under conditions that resulted in impaired lung clearance (‘overloading’) leading to inflammatory reactions and other pathologic sequelae. These observations together with the findings that carbon black, a model for diesel particles devoid of organic mutagens and carcinogens, also induced lung cancers under conditions of overloading led to the suggestion that the cancers resulted from a non-genotoxic mechanism. However, the further finding that inert particulate carcinogens devoid of organics, induced mutations has led to a re-evaluation of the role of mutations in lung carcinogenesis caused by particles and the relevance of the rat model to humans. This is especially timely as epidemiological studies suggest that humans may develop lung cancers following occupational exposure to diesel emissions by a mechanism unlikely to involve lung overloading. Finally, the recent recognition that environmental PM-10 (respiratory size particles) may be responsible for a significant portion of human morbidity and mortality, ensures that the health effect of diesel emissions will continue to receive scrutiny as they contribute to the PM-10 load.     

International Commission for Protection against Environmental Mutagens and Carcinogens. ICPEMC working paper No. 5. Genotoxicity tests as predictors of carcinogens: an analysis

Differences between the results of numerical validation studies comparing in vitro and in vivo genotoxicity tests with the rodent cancer bioassay are leading to the perception that short-term tests predict carcinogenicity only with uncertainty. Consideration of factors such as the pharmacokinetic distribution of chemicals, the systems available for metabolic activation and detoxification, the ability of the active metabolite to move from the site of production to the target DNA, and the potential for expression of the induced lesions, strongly suggests that the disparate sensitivity of the different test systems is a major reason why numerical validation is not more successful. Furthermore, genotoxicity tests should be expected to detect only a subset of carcinogens, namely genotoxic carcinogens, rather than those carcinogens that appear to act by non-genetic mechanisms. Instead of relying primarily on short-term in vitro genotoxicity tests to predict carcinogenic activity, these tests should be used in a manner that emphasizes the accurate determination of mutagenicity or clastogenicity. It must then be determined whether the mutagenic activity is further expressed as carcinogenicity in the appropriate studies using test animals. The prospects for quantitative extrapolation of in vitro or in vivo genotoxicity test results to carcinogenicity requires a much more precise understanding of the critical molecular events in both processes.     

Man-made mineral (vitreous) fibres: evaluations of cancer hazards by the IARC Monographs Programme

Man-made vitreous (glass-like) fibres are non-crystalline, fibrous inorganic substances (silicates) made primarily from rock, slag, glass or other processed minerals. These materials, also called man-made mineral fibres, include glass fibres (used in glass wool and continuous glass filament), rock or stone wool, slag wool and refractory ceramic fibres. They are widely used for thermal and acoustical insulation and to a lesser extent for other purposes. These products are potentially hazardous to human health because they release airborne respirable fibres during their production, use and removal. Man-made mineral fibres and man-made vitreous fibres have been the subject of reviews by IARC Monographs Working Groups in 1987 and 2001, respectively, which resulted in evaluations of the carcinogenic hazard to humans from exposure to these materials. These reviews and evaluations have been published as Volumes 43 and 81 of the IARC Monographs series [IARC Monographs on the Evaluation of Carcinogenic Risks to Humans, vol. 43, Man-made Mineral Fibres and Radon (1988); IARC Monographs on the Evaluation of Carcinogenic Risks to Humans, vol. 81, Man-made Vitreous Fibres (2002)]. The re-evaluation in 2001 was undertaken because there have been substantial improvements in the quality of the epidemiological information available on the carcinogenicity to humans of glass fibres, continuous glass filament and rock/slag wool. The new evaluations have addressed the limitations of earlier cohort studies, particularly concerning the lack of adjustment with respect to concomitant risk factors such as smoking and other sources of occupational exposure. In addition, the evaluation of the evidence for carcinogenicity of glass fibres to experimental animals has been refined, by making a distinction between insulation glass wool and special-purpose glass fibres. The results of the evaluations in 1987 and 2001 are thus different in several aspects. In this paper, the reviews and evaluations of the carcinogenic hazards of exposure to man-made mineral fibres (MMMF, Monograph volume 43, [1]) and man-made vitreous fibres (MMVF, Monograph volume 81, [2]) are summarised, and the differences explained. In particular, the considerations of the respective IARC Monographs Working Groups (1987, 2001) in reaching their conclusions are discussed in some detail.     

Mycotoxins as human carcinogens—the <Emphasis Type="Italic">IARC Monographs</Emphasis> classification

Humans are constantly exposed to mycotoxins (e.g. aflatoxins, ochratoxins), mainly via food intake of plant and animal origin. The health risks stemming from mycotoxins may result from their toxicity, in particular their carcinogenicity. In order to prevent these risks, the International Agency for Research on Cancer (IARC) in Lyon (France)—through its IARC Monographs programme—has performed the carcinogenic hazard assessment of some mycotoxins in humans, on the basis of epidemiological data, studies of cancer in experimental animals and mechanistic studies. The present article summarizes the carcinogenic hazard assessments of those mycotoxins, especially aflatoxins (aflatoxin B₁, B₂, G₁, G₂ and M₁), fumonisins (fumonisin B₁ and B₂) and ochratoxin A (OTA). New information regarding the genotoxicity of OTA (formation of OTA-DNA adducts), the role of OTA in oxidative stress and the identification of epigenetic factors involved in OTA carcinogenesis–should they indeed provide strong evidence that OTA carcinogenicity is mediated by a mechanism that also operates in humans–could lead to the reclassification of OTA.     

Choosing Priority-Setting Criteria for Carcinogens

For priority-setting purposes, simple criteria are needed to estimate, provisionally, the inherent properties of potential carcinogens for which adequate data are not available. Expected utility analysis is used to evaluate three such criteria from a decision-theoretic point of view: (1) the species criterion, which assigns lower priority to experimental than to epidemiological carcinogens, (2) the genotoxicity criterion, which prioritizes substances known to be genotoxic, and (3) the potency criterion, which apportions priorities according to carcinogenic potencies that are derived from animal experiments. The outcome of this analysis is favorable to the potency criterion. It is concluded that considerations of potency should have a much more prominent role than what they have in current regulatory practice.