Rodent Carcinogenicity Studies Research Articles

Mutagenic activity of carcinogens detected in transgenic rodent mutagenicity assays at dose levels used in chronic rodent cancer bioassays

Data on transgenic rodent mutagenicity of five human carcinogens were summarised and compared with the results from rodent carcinogenicity studies. Four out of five carcinogens showed mutagenic activity already at daily dose levels which induced cancer in long-term rodent bioassays in at least one target tissue of carcinogenesis. In several of these studies, even single dose applications were sufficient to significantly increase the mutation frequency in vivo. Other genotoxic carcinogens required application of multiple dosing at dose-levels used in rodent cancer bioassays to show their in vivo mutagenicity. A rodent respiratory tract carcinogen, 1,2-dibromoethane (DBE), following inhalation exposure, displayed no mutagenic activity, neither in lung nor in nasal mucosa, at a single 2-h exposure to 30 ppm, which is below the highest concentration used in a NTP cancer bioassay. In contrast, after multiple treatment for 10 days at the same daily doses, a significant increase of the mutation frequency in nasal mucosa was apparent. We conclude, that especially when studying new chemicals in these transgenic rodent mutation assays, a multiple dosing protocol should be preferred. For dose selection, the same criteria could be applied as for chronic rodent bioassays.

The National Toxicology Program evaluation of genetically altered mice as predictive models for identifying carcinogens.

National Institute of Environmental Health Sciences researchers are exploring the utility of genetically altered mice to study mechanisms of carcinogenesis. Two of these mouse models, the Tg.AC (carrier of an activated mouse H-ras oncogene) and the p53+/- (heterozygous for the wild-type tumor suppressor gene Trp53), have genetic alterations that appear to hasten their expression of chemically induced tumors. These 2 models have been proposed as a basis for new strategies for identifying chemical carcinogens and for assessing risk. The National Toxicology Program (NTP) is conducting a series of studies with these 2 genetically altered strains to further examine their strengths and weaknesses for identification of documented rodent and human carcinogens. In this first evaluation, candidates for study were drawn from the NTP historical database of 2-yr rodent carcinogenicity studies and the open literature (primarily for drugs). Results with this first set of 11 chemicals tested in genetically altered mice, compared with previous findings in the traditional 2-yr rodent assays and literature on human tumor findings, appear to support the premise advanced by Tennant et al that these models have the potential to serve as more rapid and less expensive test systems to identify carcinogens.

Spontaneous neoplasm incidences in Fischer 344 rats and B6C3F1 mice in two-year carcinogenicity studies: a National Toxicology Program update.

Spontaneous neoplasm rates were determined for control Fischer 344 (F344) rats and B6C3F1 mice from 2-yr rodent carcinogenicity studies carried out by the National Toxicology Program (NTP). The most frequently occurring neoplasms in untreated male F344 rats were testicular adenoma (89.1%), mononuclear cell leukemia (50.5%), adrenal gland pheochromocytoma (31.9%), and pituitary gland neoplasms (30.4%). For untreated female F344 rats, the most frequently occurring neoplasms were pituitary gland neoplasms (54.2%), mammary gland fibroadenoma (41.2%), and mononuclear cell leukemia (28.1%). The most frequently occurring neoplasms in untreated male B6C3F1 mice were liver adenoma/carcinoma (42.2%), lung adenoma/carcinoma (20.5%), and malignant lymphoma (8.3%). For untreated female B6C3F1 mice, the most frequently occurring neoplasms were liver adenoma/carcinoma (23.6%), malignant lymphoma (20.9%), and pituitary gland adenoma/carcinoma (14.8%). The tumor rates observed in feeding study (untreated) and inhalation study (chamber) control rats were generally similar. The major exceptions were pituitary gland tumors and testicular adenoma in male F344 rats. The overall incidence of testicular adenoma was much lower in chamber controls (69.4%) than in feeding study controls (89.1%), whereas pituitary gland neoplasm showed the opposite trend (60.7% vs 30.4%). The most likely explanation for this difference is related to the individual housing of chamber controls and the group housing of feeding study controls. Differences in diagnostic criteria may influence reported tumor rates. To ensure consistency and comparability of tumor diagnosis from study to study, the NTP uses rigorous histopathology quality assurance and peer review procedures. Biological factors such as body weight may also affect tumor incidence. For example, increased body weights are associated with increased incidences of certain site-specific neoplasms, especially pituitary gland and mammary gland neoplasms in rats and liver tumors in mice. The presence of Helicobacter hepaticus has been associated with an increased incidence of liver neoplasms in male B6C3F1 mice. Other factors that may produce differences in control tumor rates from study to study include diet, environmental factors, genetic drift, study duration, and survival differences. The NTP database provides historical control data that may be useful in the evaluation of possible chemically related changes in tumor incidence. However, it is essential that the study being evaluated be comparable to those in the NTP database with respect to those factors that are known to influence tumor occurrence.

Need for dietary control by caloric restriction in rodent toxicology and carcinogenicity studies

The conditions under which laboratory animals are maintained can powerfully influence the results of toxicological studies utilized for risk assessment. Nutrition is of importance in toxicological bioassays and research, because diet composition and the conditions under which it is fed can affect the metabolism and activity of xenobiotic test substances and alter the results and reproducibility of long‐term studies. It is known that ad libitum (AL) overfed sedentary laboratory rodents suffer from an early onset of degenerative disease and diet‐related tumors that lead to poor survival in chronic bioassays. AL‐fed animals are not well‐controlled subjects for any experimental studies. Examination of study‐to‐study variability in food consumption, body weight, and survival in carcinogenicity studies for the same strain or stock of rodents shows tremendous laboratory‐to‐laboratory variability. However, a significant correlation between average food (calorie) consumption, adult body weight, and survival has been clearly established. The use of moderate dietary restriction (DR) results in a better controlled rodent model with a lower incidence or delayed onset of spontaneous diseases and tumors. Operationally simple, moderate DR significantly improves survival, controls adult body weight and obesity, reduces age‐related renal, endocrine, and cardiac diseases, increases exposure time, and increases the statistical sensitivity of these expensive, chronic bioassays to detect a true treatment effect. A moderate DR regimen of 70‐75% of the maximum unrestricted AL food intake is recommended as a nutritionally intelligent, well‐established method in conducting well‐controlled toxicology and carcinogenicity studies.

In vivo transgenic bioassays and assessment of the carcinogenic potential of pharmaceuticals.

There is general agreement in the scientific community on the need to improve carcinogenicity testing and the assessment of human carcinogenic risk and to incorporate more information on mechanisms and modes of action into the risk assessment process. Advances in molecular biology have identified a growing number of genes such as protooncogenes and tumor-suppressor genes that are highly conserved across species and are associated with a wide variety of human and animal cancers. In vivo transgenic rodent models incorporating such mechanisms are used to identify mechanisms involved in tumor formation and as selective tests for carcinogens. Transgenic methods can be considered an extension of genetic manipulation by selective breeding, which long has been employed in science and agriculture. The use of two rodent species in carcinogenicity testing is especially important for identifying transspecies carcinogens. The capacity of a substance to induce neoplasia across species suggests that the mechanism(s) involved in the induction of the neoplasia are conserved and therefore may have significance for humans. Based on available information there is sufficient experience with some in vivo transgenic rodent carcinogenicity models to support their application as complementary second species studies in conjunction with a single 2-year rodent carcinogenicity study. The optional substitution of a second 2-year rodent carcinogenicity study with an alternative study such as an in vivo transgenic carcinogenicity study is part of the International Conference on Harmonization guidance S1B: Testing for Carcinogenicity of Pharmaceuticals. This guidance is intended to be flexible enough to accommodate a wide range of possible carcinogenicity assessment models currently under consideration or models that may be developed in the future. The use of an in vivo transgenic mouse model in place of a second 2-year mouse study will improve the assessment of carcinogenic risk by contributing insights into the mechanisms of tumorigenesis and potential human relevance not available from a standard 2-year bioassay. It is envisioned that this will stimulate the further development of more efficient and relevant methods for identifying and assessing potential human carcinogenic risk, which will benefit public health.

Effect of Dietary Optimization on Growth, Survival, Tumor Incidences and Clinical Pathology Parameters in CD Sprague-Dawley and Fischer-344 Rats: A 104-Week Study

ABSTRACTControversy regarding the use of ad libitum feeding in chronic rodent toxicity studies will soon result in issue of a FDA Points to Consider document. Caloric intakes are now recognized to be important uncontrolled variables in bioassays because rodents chronically fed ad libitum become obese, reproductively senile and have increased incidences of age-related diseases, higher tumor burdens and decreased survival. The available literature suggests that ad libitum feeding neither optimizes the health and well-being of rodents nor provides the best model for use in evaluation of pharmacological and toxicological profiles. Use of an optimized diet, restricted in terms of caloric intakes, has been proposed for chronic toxicity and carcinogenicity studies in rodents. It is suggested that limiting caloric intakes to 50-80% of ad libitum consumption would result in lower body weights, decreased tumor incidences and prolonged survival in the controls. To evaluate the influence of diet on chronic toxicity and carcinogenicity studies in rats, two 104-week studies were conducted. These studies consisted of 280 CD Sprague-Dawley and 280 Fischer-344 rats fed ad libitum, and 140 CD Sprague-Dawley and 140 Fischer-344 rats fed a diet that was optimized by limiting caloric intakes by 15-35%. Both diets consisted of certified commercial diet in meal form. The optimized diet reduced weight gain approximately 50% after 100 weeks. Clinical chemistry and hematology parameters showed negligible effects of reduced diet, with the exception that serum triglycerides were lower in males and females in both strains at weeks 52 and 104. The ad libitum-fed animals had a higher incidence of pseudopregnancy, aggressiveness, foot sores and abscesses than the animals fed an optimized diet. These effects were more pronounced in the CD Sprague-Dawley rats than in the Fischer-344 rats. At the completion of the 104-week study, survival in the ad libitum fed CD Sprague-Dawley rats was approximately one-half that of the animals fed an optimized diet (39% versus 76%). The difference in survival between Fischer-344 rats fed ad libitum and those fed an optimized diet was less pronounced (78% versus 89%). A reduced incidence of palpable tissue masses in the ad libitum-fed CD Sprague-Dawley rats versus the animals fed an optimized diet reflected inability to detect small masses in the obese ad libitum-fed animals. In contrast, the leaner Fischer-344 ad libitum-fed animals had an increased incidence of palpable tissue masses. After 52 weeks, 40 animals from each strain and feeding regimen were killed and subjected to complete necropsy and histopathological examination; the remainder of the survivors was examined at the completion of the study (104 weeks). Use of an optimized diet substantially reduced the incidences of endocrine-mediated tumors in both rat strains and delayed the onset of leukemia in Fischer-344 rats. These results indicate the need to further investigate the relationship of increased caloric intakes and endocrine-mediated or strain specific tumors and support FDA's and others' positions that use of diet optimization in chronic toxicity and carcinogenicity rodent bioassays has the potential to remarkably improve the scientific quality and relevance of these studies. It also identified that the small increases in cost associated with diet optimization are far exceeded by the advantages of increased survival of animals, reduced intercurrent disease and rumor burdens, and increased ease of histopathological processing and evaluation.

Calcium channel blockers and the risk of cancer: a preclinical assessment.

The preclinical evidence for a potential influence of calcium channel blockers (CCBs) on carcinogenesis is discussed in the light of a broad database from rodent carcinogenicity studies as well as literature data. In all bioassays performed in rats and mice on the dihydropyridine CCBs--nifedipine, nimodipine, nisoldipine, and nitrendipine--no evidence was found for a carcinogenic potential of these compounds. Calcium is an essential intracellular signal for cell proliferation and apoptosis. The crucial role of increased cell proliferation in all stages of carcinogenesis is well documented. Some indirect experimental evidence also points to a role of defective apoptosis in tumor promotion. CCBs uniformly inhibit cell proliferation, whereas the influence of CCBs on apoptosis is inconsistent, resulting in an inhibition or increase in apoptosis dependent on cell type. Accordingly, antitumorigenic effects of CCBs have been reported based on their antiproliferative action. A tumor-promoting effect of CCBs based on inhibition of apoptosis, however, remains purely speculative and, in fact, can be denied based on the results of in vivo bioassays. It is therefore concluded that there is no preclinical evidence that should give rise to concern over the carcinogenic potential of dihydropyridine-type CCBs.

Neonatal Mouse Assay for Tumorigenicity: Alternative to the Chronic Rodent Bioassay

The chronic rodent bioassay for tumors has been utilized systematically for 25 years to identify chemicals with carcinogenic potential in man. In general, those chemicals exhibiting tumorigenicity at multiple sites in both mice and rats have been regarded as possessing strong carcinogenic potential in humans. In comparison, the value of data collected for those test chemicals exhibiting more sporadic tumorigenicity results (e.g., single species/single sex or dose-independent) has been questioned. As knowledge of the carcinogenic process has increased, several alternative test systems, usually faster and less expensive than the 2-year bioassay, have been suggested for identification of the strongly acting, transspecies carcinogens. The International Conference on Harmonization for Technical Requirements for the Registration of Pharmaceuticals for Human Use has proposed an international standard that allows for the use of one long-term rodent carcinogenicity study, plus one supplementary study to identify potential human pharmaceutical carcinogens. The neonatal mouse assay for tumorigenicity has been used since 1959; however, relative to other alternate tests, little has been written about this system. It is clear that this assay system successfully identifies transspecies carcinogens from numerous chemical classes, thus recommending itself as a strong candidate for a supplementary study to identify potential human carcinogens. In contrast, there are decidedly less data available from this assay in response to pharmaceuticals shown to exhibit weak and/or conflicting results in the 2-year bioassay, knowledge invaluable to the regulatory process. This paper reviews the historical development and our experience with the neonatal mouse assay and includes suggestions for a standardized protocol and strategies to document its response to “weak” and/or “nongenotoxic” carcinogens.

An update of the National Toxicology Program database on nasal carcinogens

Nearly 500 long-term rodent carcinogenicity studies carried out by the National Cancer Institute and the National Toxicology Program were examined, and 12 chemicals were identified that produced nasal tumors: allyl glycidol ether, p-cresidine, 1,2-dibromo-3-chloropropane, 1,2-dibromoethane, 2,3-dibromo-1-propanol, dimethylvinyl chloride, 1,4-dioxane, 1,2-epoxybutane, iodinated glycerol, procarbazine, propylene oxide, and 2,6-xylidine. All 12 of these chemicals produced nasal tumors in rats, and 5 also produced nasal tumors in mice. Most of the nasal carcinogens (1) produced tumor increases in both sexes, (2) produced tumors at other sites as well, (3) had significantly reduced survival at doses that were carcinogenic, and (4) were genotoxic. Only 5 of the 12 nasal carcinogens were administered by inhalation. A variety of different types of nasal cavity tumors were produced, and specific tumor rates are given for those chemicals causing multiple tumor types. Increased incidences of nasal neoplasms were often accompanied by suppurative/acute inflammation, epithelial/focal hyperplasia and squamous metaplasia. However, high incidences of these nonneoplastic nasal lesions were also frequently seen in inhalation studies showing no evidence of nasal carcinogenicity, suggesting that in general nasal carcinogenesis is not associated with the magnitude of chronic toxicity observed at this site.

Cytochrome P450 mediated bioactivation of methyleugenol to 1'- hydroxymethyleugenol in Fischer 344 rat and human liver microsomes

Cytochrome P450 mediated metabolism of methyleugenol to the proximate carcinogen 1'-hydroxymethyleugenol has been investigated in vitro. Kinetic studies undertaken in liver microsomes from control male Fischer 344 rats revealed that this reaction is catalyzed by high affinity (Km of 74.9 +/- 9.0 microM, Vmax of 1.42 +/- 0.17 nmol/min/nmol P450) and low affinity (apparent Km several mM) enzymic components. Studies undertaken at low substrate concentration (20 microM) with microsomes from livers of rats treated with the enzyme inducers phenobarbital, dexamethasone, isosafrole and isoniazid indicated that a number of cytochrome P450 isozymes can catalyze the high affinity component. In control rat liver microsomes, 1'-hydroxylation of methyleugenol (assayed at 20 microM substrate) was inhibited significantly (P < 0.05) by diallylsulfide (40%), p-nitrophenol (55%), tolbutamide (30%) and alpha-naphthoflavone (25%) but not by troleandomycin, furafylline, quinine or cimetidine. These results suggested that the reaction is catalyzed by CYP 2E1 and by another as yet unidentified isozyme(s) (most probably CYP 2C6), but not by CYP 3A, CYP 1A2, CYP 2D1 or CYP 2C11. Administration of methyleugenol (0-300 mg/kg/day for 5 days) to rats in vivo caused dose-dependent auto-induction of 1'-hydroxylation of methyleugenol in vitro which could be attributed to induction of various cytochrome P450 isozymes, including CYP 2B and CYP 1A2. Consequently, high dose rodent carcinogenicity studies are likely to over-estimate the risk to human health posed by methyleugenol. The rate of 1'-hydroxylation of methyleugenol in vitro in 13 human liver samples varied markedly (by 37-fold), with the highest activities being similar to the activity evident in control rat liver microsomes. This suggests that the risk posed by dietary ingestion of methyleugenol could vary markedly in the human population.

Body weight-tumor incidence correlations in long-term rodent carcinogenicity studies.

Associations between body weight and tumor incidence and among the incidences of selected site-specific tumors were examined for more than 4,000 male and female Fischer-344 rats and B6C3F1 mice in the National Toxicology Program historical control database. Incidences of certain site-specific tumors, most notably mammary gland and pituitary gland tumors in rats and liver tumors in mice, were shown to have a strong positive correlation with 52-wk body weight. Using individual animal data, logistic regression models were derived for predicting site-specific tumor incidence as a function of 52-wk body weight, age, and other factors. This association between body weight and tumor incidence can explain many of the decreased tumor incidences observed in National Toxicology Program carcinogenicity studies. Body weight differences between dosed and control groups can also mask carcinogenic effects for those sites sensitive to body weight changes. Thus, when designing long-term rodent carcinogenicity studies, measures should be taken to minimize potential body weight differences between dosed and control groups. There were a number of small but significant negative correlations among tumor incidences, reflecting primarily the lethality of the tumors in question. None of these correlations (nor the 2 small positive correlations found) are likely to have any impact on the interpretation of experimental results. However, the high negative correlation between pituitary gland and testis tumors in male Fischer-344 rats cannot be dismissed so easily, does not reflect tumor lethality, and is currently being studied further.

Carcinogenicity Testing and the Evaluation of Regulatory Requirements for Pharmaceuticals

The results of rat and mouse carcinogenicity studies for 282 human pharmaceuticals in the FDA database were analyzed and compared as part of an International Conference on Harmonization (ICH) evaluation of rodent carcinogenicity studies and their utility for carcinogenicity testing. A majority of the carcinogenicity studies in the FDA database were carried out in Sprague–Dawley-derived rats and Swiss–Webster-derived CD-1 mice in contrast to Fisher 344 rats and B6C3F1 mice employed in National Toxicology Program (NTP) studies. Despite the differences in rodent strains, the relative proportion of compounds with positive findings (44.3%) and the degree of overall concordance between rats and mice (74.1%) in the FDA database were similar to the NTP rodent carcinogenicity database. Carcinogenicity studies in two rodent species are necessary primarily to identify trans-species tumorigens, which are considered to pose a relatively greater potential risk to humans than single species positive compounds. Two-year carcinogenicity studies in both rats and mice may not be the only means of identifying trans-species tumorigens. Sufficient experience is now available for some alternativein vivocarcinogenicity models to support their application as complementary studies in combination with a single 2-year carcinogenicity study to identify trans-species tumorigens. Our analysis of the rodent carcinogenicity studies supports such an approach for assessing carcinogenic potential without compromising the public health.

The Utility of Two Rodent Species in Carcinogenic Risk Assessment of Pharmaceuticals in Europe

For the past 20–30 years, lifespan carcinogenicity studies for pharmaceuticals have been required to be carried out in two rodent species. Due to scientific progress, the necessity/justification of lifespan studies in two species for the assessment of carcinogenic risk of pharmaceuticals is currently under discussion. A study in one species (either rat or mouse) might suffice. To appraise the need for a study in a second species, a database was compiled of all pharmaceuticals tested for carcinogenicity for which a marketing authorization was applied for in Germany and The Netherlands since 1980. The incidence of treatment-related tumor findings was determined in either rat or mouse or in both. Tumor findings occurred for nearly 50% of all compounds, with the rat being more sensitive than the mouse. Specific attention was given to the question whether tumor findings in mice ever caused the regulatory authorities to refuse registration, to restrict the proposed therapeutic indication of a pharmaceutical, or to apply a cautionary label. It was found that no tumor findings in mice alone ever led to such a regulatory action. In addition, whether mouse studies had been important in interpreting the results of rat studies was determined. A negative mouse study (no tumors found) was rarely used to declare the rat findings irrelevant to humans. A mechanistic explanation was used as a much more important argument in the assessment of tumor findings in rats. In case of transspecies findings, the target organs were the usual ones, such as lung and liver, or the tumors occurred as a result of an exaggerated pharmacodynamic action expected from the pharmacology of the compound. The results of the database thus question the need of maintaining the requirement of rodent carcinogenicity studies in two species.

Assessment of the genotoxic risk from laxative senna products.

Laxative senna products and several of their specific components have been submitted to a large number of genetic tests. While most studies gave negative responses, results from some of the studies suggest that components of senna products, particularly emodin and aloe-emodin, have genotoxic activity. Assessment of the genotoxicity profile of these substances, in light of other data from animal and human metabolism or kinetic studies, human clinical trials and rodent carcinogenicity studies do not support concerns that senna laxatives pose a genotoxic risk to humans when consumed under prescribed use conditions.

Calcium channel blockers and cancer: is there preclinical evidence for an association?

The preclinical evidence for a potential influence of calcium channel blockers (CCB) on carcinogenesis is discussed in the light of rodent carcinogenicity studies as well as mechanistic data. In the bioassays performed in rats and mice on the dihydropyridine CCB nifedipine, nimodipine, nisoldipine and nitrendipine, no evidence was found for a carcinogenic potential of these compounds. Moreover, the mechanistic knowledge on the influence of CCB on the fundamental processes of cell proliferation and apoptosis is not in favor of a tumor-promoting activity of these compounds. It is, therefore, concluded that there is no preclinical evidence that the therapeutic use of CCB of the dihydropyridine class is associated with an increased risk of cancer.

New Diet (NTP-2000) for Rats in the National Toxicology Program Toxicity and Carcinogenicity Studies

Composition of diet may influence growth, diseases, tumor rates, and responses to chemical treatment. Since 1980 the NIH-07 open formula nonpurified diet has been the selected diet for the National Toxicology Program (NTP) toxicity and carcinogenicity studies in rodents. Studies with nonpurified experimental diets with lower protein and higher fat and fiber than the NIH-07 diet indicated that the diet for Fischer-344 (F344) rats in long-term studies could be modified to decrease the severity of chronic diseases and to decrease/delay the development of spontaneous tumors. Based on the results of these studies a new open formula nonpurified diet designated as NTP-2000 was formulated to contain ∼14.5% protein, ∼8.5% fat, and ∼9.5% fiber. Corn, wheat, and wheat middlings contribute to about 60% of the ingredients; soybean meal, fish meal, and alfalfa meal are the additional sources of protein; purified cellulose, oat hulls, and alfalfa meal are the major sources of fiber; and soy oil and corn oil are the major sources of fat in the NTP-2000 diet. The Ca:P ratio and mineral and vitamin concentrations were reformulated based on AIN-93 and NRC-95 recommendations. The NIH-07 and the NTP-2000 diets were fed to groups of 6-week-old F344 rats for 13 weeks and evaluated for growth patterns, food and water consumptions, hematology and clinical chemistry parameters, and organ weights and pathological changes. Growth patterns and body weights were similar for both diets. Food consumptions were slightly higher and water consumptions were slightly lower for the groups fed NTP-2000 diet. There were no differences in hematological parameters between the groups fed the above diets. Serum levels of cholesterol, alkaline phosphatase, and 5′ nucleotidase were slightly higher in groups fed the NTP-2000 diet possibly due to higher fat content of this diet. However, the serum triglyceride levels were slightly lower in groups fed the NTP-2000 diet and it may be related to higher fiber content of the NTP-2000 diet. The liver and kidney weights of the groups fed NTP-2000 diet were significantly lower possibly due to lower protein content of this diet and lower protein consumption associated changes in Phase I and Phase II drug metabolizing enzyme systems. The adrenal weights were also lower in groups fed the new diet. The NTP-2000 diet prevented nephrocalcinosis and decreased the severity of nephropathy and cardiomyopathy, the common lesions of F344 rats in 13-week studies. These results indicate that the NTP-2000 diet is adequate for growth and maintenance of rats and appears to prevent or decrease the severity of diet-associated lesions.

Are lifespan rodent carcinogenicity studies defensible for pharmaceutical agents?

For most pharmaceuticals, the assessment of carcinogenic risk to humans can be made from information available from a) genotoxicity studies in vivo, b) 3-6 month toxicology studies in two or more animal species and c) clinical investigations in Phase I and II studies aimed at assessing the existence of risk factors (genotoxicity, immune suppression, hormonal activity and chronic irritation/inflammation) associated with cancer in humans caused by pharmaceuticals. The rodent carcinogenicity bioassay is redundant for compounds with such properties. In considering the utility of a bioassay, one must recognize that the outcome of the bioassay has been shown to be predictable for about half of a random selection of chemicals in the US National Toxicology Program (NTP). The predictability of bioassay outcomes for many pharmaceuticals should be even better, given the availability of extensive knowledge on genotoxic potential in vivo and of pharmacological mechanisms, this adding to the redundancy of the bioassay. Furthermore, the value of the bioassay is itself questionable. The inconsistencies in tumor responses between rodent species and strains, the simultaneous tumor increase and decreases within a study and the susceptibility to tumorigenicity from non-genotoxic chemicals by mechanisms now shown to be of no relevance to humans, together make the use of rodents highly misleading as predictors of human cancer risk. For pharmaceuticals with a novel or poorly-understood pharmacodynamic mechanism, useful information on long-term adverse effects that might presage a carcinogenic hazard to humans may be obtained from a 12 month study, usually in rats, conducted at clinically relevant dose levels.

Analysis of National Toxicology Program rodent bioassay data for anticarcinogenic effects

We reanalyzed data from 218 two-year rodent carcinogenicity studies carried out by the National Toxicology Program (NTP). These data were originally collected for the purpose of identifying potential human carcinogens. However, the objective of our analysis was to investigate the frequency of possible anticarcinogenic effects in these data, since recurring cases of chemical-associated tumor reductions have been noted in the course of these studies over time. Our analysis reveals that most (> 90%) NTP-tested chemicals show at least one statistically significant ( p < 0.05) decrease in site-specific tumor incidence. Because of the large number of statistical comparisons made in a long-term bioassay, random variability can account for many of these tumor decreases. However, we found that certain tumors (predominantly those with a high spontaneous incidence) show chemically related decreases far more frequently than chance expectation. Many of these decreases, particularly those for pituitary and mammary gland tumors, adrenal pheochromocytoma and uterine polyps in rats and liver and lung tumors in mice, are associated with the reduced body weights frequently observed in the dosed groups. The chemically related decreased incidences of leukemia in rats appear to be related to spleen damage, i.e., chemically related splenic toxicity is evident for most chemicals showing decreased incidences of leukemia. While random variability, associations with body weight and splenic toxicity can account for most of the decreased tumor incidences observed in NTP studies, there are other tumor decreases that could not be totally explained by these factors. Further investigations of possible mechanisms of action are underway. These data are relevant to the concept of chemoprevention as well as to the task of using long-term laboratory animal studies to predict enhanced human environmental-cancer risk for regulatory purposes.

Carcinogenicity of Lipid-Lowering Drugs

To review the findings and implications of studies of rodent carcinogenicity of lipid-lowering drugs. Summaries of carcinogenicity studies published in the 1992 and 1994 Physicians' Desk Reference (PDR), additional information obtained from the US Food and Drug Administration, and published articles identified by computer searching, bibliographies, and consultation with experts. We tabulated rodent carcinogenicity data from the 1994 PDR for all drugs listed as "hypolipidemics." For comparison, we selected a stratified random sample of antihypertensive drugs. We also reviewed methods and interpretation of carcinogenicity studies in rodents and results of clinical trials in humans. All members of the two most popular classes of lipid-lowering drugs (the fibrates and the statins) cause cancer in rodents, in some cases at levels of animal exposure close to those prescribed to humans. In contrast, few of the antihypertensive drugs have been found to be carcinogenic in rodents. Evidence of carcinogenicity of lipid-lowering drugs from clinical trials in humans is inconclusive because of inconsistent results and insufficient duration of follow-up. Extrapolation of this evidence of carcinogenesis from rodents to humans is an uncertain process. Longer-term clinical trials and careful postmarketing surveillance during the next several decades are needed to determine whether cholesterol-lowering drugs cause cancer in humans. In the meantime, the results of experiments in animals and humans suggest that lipid-lowering drug treatment, especially with the fibrates and statins, should be avoided except in patients at high short-term risk of coronary heart disease.

New Diet (NTP-2000) for Rats in the National Toxicology Program Toxicity and Carcinogenicity Studies

Composition of diet may influence growth, diseases, tumor rates, and responses to chemical treatment. Since 1980 the NIH-07 open formula nonpurified diet has been the selected diet for the National Toxicology Program (NTP) toxicity and carcinogenicity studies in rodents. Studies with nonpurified experimental diets with lower protein and higher fat and fiber than the NIH-07 diet indicated that the diet for Fischer-344 (F344) rats in long-term studies could be modified to decrease the severity of chronic diseases and to decrease/delay the development of spontaneous tumors. Based on the results of these studies a new open formula nonpurified diet designated as NTP-2000 was formulated to contain approximately 14.5% protein, approximately 8.5% fat, and approximately 9.5% fiber. Corn, wheat, and wheat middlings contribute to about 60% of the ingredients; soybean meal, fish meal, and alfalfa meal are the additional sources of protein; purified cellulose, oat hulls, and alfalfa meal are the major sources of fiber; and soy oil and corn oil are the major sources of fat in the NTP-2000 diet. The Ca:P ratio and mineral and vitamin concentrations were reformulated based on AIN-93 and NRC-95 recommendations. The NIH-07 and the NTP-2000 diets were fed to groups of 6-week-old F344 rats for 13 weeks and evaluated for growth patterns, food and water consumptions, hematology and clinical chemistry parameters, and organ weights and pathological changes. Growth patterns and body weights were similar for both diets. Food consumptions were slightly higher and water consumptions were slightly lower for the groups fed NTP-2000 diet. There were no differences in hematological parameters between the groups fed the above diets. Serum levels of cholesterol, alkaline phosphatase, and 5' nucleotidase were slightly higher in groups fed the NTP-2000 diet possibly due to higher fat content of this diet. However, the serum triglyceride levels were slightly lower in groups fed the NTP-2000 diet and it may be related to higher fiber content of the NTP-2000 diet. The liver and kidney weights of the groups fed NTP-2000 diet were significantly lower possibly due to lower protein content of this diet and lower protein consumption associated changes in Phase I and Phase II drug metabolizing enzyme systems. The adrenal weights were also lower in groups fed the new diet. The NTP-2000 diet prevented nephrocalcinosis and decreased the severity of nephropathy and cardiomyopathy, the common lesions of F344 rats in 13-week studies. These results indicate that the NTP-2000 diet is adequate for growth and maintenance of rats and appears to prevent or decrease the severity of diet-associated lesions.