Toxicology and carcinogenesis studies of riddelliine (CAS No. 23246-96-0) in F344/N rats and B6C3F1 mice (gavage studies)

Riddelliine belongs to a class of toxic pyrrolizidine alkaloids and is isolated from plants of the genera Crotalaria, Amsinckia, and Senecio that grow in the western United States. Cattle, horses, and sheep that ingest these plants succumb to their toxic effects. Riddelliine residues have been found in meat, milk, and honey, and the plants may contaminate human food sources. Riddelliine was nominated for study by the Food and Drug Administration because of its potential for human exposure and its economic impact on the livestock industry and because the toxicity of other pyrrolizidine alkaloids suggests riddelliine may be carcinogenic. Male and female F344/N rats and B6C3F1 mice received riddelliine (approximately 92% pure) by gavage. Female rats and male and female mice were dosed for 2 years; due to high mortality, the study in male rats was terminated at week 72. In vitro genetic toxicology studies were conducted in Salmonella typhimurium and in cultured Chinese hamster ovary (CHO) cells. In addition, riddelliine was evaluated in vivo for induction of micronuclei in mouse bone marrow and peripheral blood erythrocytes and for induction of S-phase DNA synthesis and unscheduled DNA synthesis in the liver of rats and mice. Riddelliine-induced DNA adduct levels were determined in liver tissue obtained from female rats admininstered riddelliine for 3 or 6 months. 2-YEAR STUDY IN RATS: Groups of 50 male and 50 female rats were administered 0 or 1 mg riddelliine/kg body weight in sodium phosphate buffer by gavage 5 days per week; additional groups of 50 female rats received 0.01, 0.033, 0.1, or 0.33 mg/kg. A wide dose range was used in female rats to better characterize the dose-response curve. Females were dosed for 105 weeks; due to high mortality, male rats were terminated at week 72. All but three 1 mg/kg males died before week 70, and all 1 mg/kg females died before week 97. Mean body weights of 1 mg/kg males and females were less than those of the vehicle controls throughout most of the study. The only clinical finding related to riddelliine administration was a general debilitation of the animals prior to death. Hemangiosarcomas were present in the liver of 86% of males and 76% of females in the 1 mg/kg groups, and this neoplasm was considered the cause of the large number of early deaths in these groups. The incidences of hepatocellular adenoma and mononuclear cell leukemia in 1 mg/kg males and females were significantly increased. Nonneoplastic lesions related to riddelliine treatment occurred in the liver and kidney of males and females. Analyses of liver tissue from female rats treated with riddelliine for 3 or 6 months yielded eight DNA adducts; these were the same as DNA adducts formed in vitro by the metabolism of riddelliine by human liver microsomes in the presence of calf thymus DNA. 2-YEAR STUDY IN MICE: Groups of 50 male and 50 female mice were administered riddelliine in sodium phosphate buffer by gavage at doses of 0 or 3 mg/kg, 5 days per week, for 105 weeks; additional groups of 50 male mice received 0.1, 0.3, or 1 mg/kg for 105 weeks. A wide dose range was used in male mice to better characterize the dose-response curve. Survival of males and females administered 3 mg/kg was significantly less than that of the vehicle controls. Mean body weights of 3 mg/kg mice were less than those of the vehicle controls throughout most of the study. Hemangiosarcomas of the liver were present in 62% of males in the 3 mg/kg group. The incidences of hepatocellular neoplasms occurred with negative trends in male mice and were significantly decreased in 3 mg/kg females. The incidences of alveolar/bronchiolar neoplasms in 3 mg/kg females were significantly increased. Nonneoplastic lesions related to riddelliine administration occurred in the liver and kidney of males and females and in the lung and arteries (multiple tissues) of females. GENETIC TOXICOLOGY: Riddelliine was mutagenic in S. typhimurium strain TA100 with, but not without, S9 activation; no significant mutagenic activity was detected in strain TA98 or TA1535,ed in strain TA98 or TA1535, with or without S9. A small, dose-related increase in mutant colonies seen in strain TA97 with S9 was judged to be equivocal. Riddelliine induced sister chromatid exchanges in cultured CHO cells with and without S9. Chromosomal aberrations were induced in CHO cells only in the presence of S9. Following 4 or 13 weeks of daily gavage treatment with riddelliine, no increases in the frequency of micronucleated erythrocytes were noted in the peripheral blood of male or female B6C3F1 mice. Use of a single intraperitoneal injection protocol, however, produced a small but significant increase in the frequency of micronucleated eryth-rocytes in peripheral blood of male Swiss mice 48 hours after injection; bone marrow analysis 24 hours after injection demonstrated a small but insignificant increase in the frequency of micronuclei. Unscheduled DNA synthesis was detected in cultured hepatocytes from male and female rats and mice following 5 or 30 days of riddelliine treatment by gavage. In addition, an S-phase DNA synthesis was observed in cultured hepatocytes of male and female rats treated for either time period. CONCLUSIONS: Under the conditions of these studies, there was clear evidence of carcinogenic activity of riddelliine in male and female F344/N rats based primarily on increased incidences of hemangiosarcoma in the liver. The increased incidences of hepatocellular adenoma and mononuclear cell leukemia in male and female rats were also considered to be treatment related. There was clear evidence of carcinogenic activity of riddelliine in male B6C3F1 mice based on increased incidences of hemangiosarcoma in the liver. There was clear evidence of carcinogenic activity in female B6C3F1 mice based on increased incidences of alveolar/bronchiolar neoplasms. Administration of riddelliine by gavage resulted in nonneoplastic lesions in the liver and kidney of male and female rats; the liver and kidney of male and female mice; and the lung and arteries (multiple tissues) of female mice. Decreased incidences of hepatocellular neoplasms in male and female mice were related to riddelliine administration.     

Species and gender differences in the carcinogenic activity of trimethylolpropane triacrylate in rats and mice

Trimethylolpropane triacrylate (TMPTA) is a multifunctional monomer with industrial applications. To determine the carcinogenic potential, male and female F344/N rats and B6C3F1/N mice were administered TMPTA (0, 0.3, 1.0, or 3.0 mg/kg) in acetone dermally for 2 years. There were no differences in the body weights and survival in the treated animals compared to controls. Nonneoplastic skin lesions at the site of application included epidermal hyperplasia and hyperkeratosis in both rats and mice. There were no incidences of tumors at the site of application in rats and mice. Rare malignant liver neoplasms were observed in female mice that included hepatoblastoma in the 0.3 and 3.0 mg/kg groups, and hepatocholangiocarcinoma in the 1.0 and 3.0 mg/kg groups. The incidences of uterine stromal polyp and stromal polyp or stromal sarcoma (combined) in female mice occurred with positive trends and the incidences were significantly increased in the 3.0 mg/kg group. A marginal increase in the incidences of malignant mesothelioma in male rats may have been related to TMPTA treatment. In conclusion, our studies show that TMPTA is a dermal irritant in both rats and mice of either sex. Increased incidences of tumor formation were observed in female mice and male rats.     

Inhalation toxicity and carcinogenicity studies of cobalt sulfate.

Cobalt sulfate is a water-soluble cobalt salt with a variety of industrial and agricultural uses. Several cobalt compounds have induced sarcomas at injection sites in animals, and reports have suggested that exposure to cobalt-containing materials may cause lung cancer in humans. The present studies were done because no adequate rodent carcinogenicity studies had been performed with a soluble cobalt salt using a route relevant to occupational exposures. Groups of 50 male and 50 female F344/N rats and B6C3F1 mice were exposed to aerosols containing 0, 0.3, 1.0, or 3.0 mg/m3 cobalt sulfate hexahydrate, 6 h/day, 5 days/week, for 104 weeks. Survival and body weights of exposed rats and mice were generally unaffected by the exposures. In rats, proteinosis, alveolar epithelial metaplasia, granulomatous alveolar inflammation, and interstitial fibrosis were observed in the lung in all exposed groups. Nonneoplastic lesions of the nose and larynx were also attributed to exposure to all concentrations of cobalt sulfate. In 3.0 mg/m3 male rats and in female rats exposed to 1.0 or 3.0 mg/m3, the incidences of alveolar/bronchiolar neoplasms were increased over those in the control groups. Lung tumors occurred with significant positive trends in both sexes. The incidences of adrenal pheochromocytoma in 1.0 mg/m3 male rats and in 3.0 mg/m3 female rats were increased. Nonneoplastic lesions of the respiratory tract were less severe in mice than in rats. In mice, alveolar/bronchiolar neoplasms in 3.0 mg/m3 males and females were greater than those in the controls, and lung tumors occurred with significantly positive trends. Male mice had liver lesions consistent with a Helicobacter hepaticus infection. Incidences of liver hemangiosarcomas were increased in exposed groups of male mice; however, because of the infection, no conclusion could be reached concerning an association between liver hemangiosarcomas and cobalt sulfate. In summary, exposure to cobalt sulfate by inhalation resulted in increased incidence of alveolar/bronchiolar neoplasms and a spectrum of inflammatory, fibrotic, and proliferative lesions in the respiratory tracts of male and female rats and mice. Adrenal pheochromocytomas were increased in female rats, and possibly in male rats.     

Effects of a polybrominated biphenyl mixture in the rat and mouse. II. Lifetime study

This study was undertaken to characterize the long-term toxic and carcinogenic potential of a polybrominated biphenyl (PBB) mixture in rats and mice of both sexes. Fischer 344 rats and B6C3F1 mice were given 125 po doses of PBB over a 6-month period at 0 (control), 0.1, 0.3, 1.0, 3.0, and 10.0 mg/kg body weight/day (5 days/week) and observed for an additional 23 months for rats and 24 months for mice (lifetime observation). The treatments (0.3 mg/kg or higher dosages) shortened the survival time in male rats whereas no such effect was observed in treated females. There was also evidence of shortened survival time in mice treated with 10.0 mg/kg PBB. As observed by uv light, hepatic porphyrin markedly increased at the 6-month observation, then tended to decrease, primarily in mice, following cessation of exposure. Significantly higher incidences of atypical hepatocellular foci, neoplastic nodules, hepatocellular carcinomas, and cholangiocarcinomas were observed in exposed rats. The incidence of hepatocellular carcinoma was also increased in both male (95%) and female (88%) mice (highest dose level) compared with control male (48%) and female (0%) mice. The incidence of hepatic neoplasms appeared to be dose dependent in both species. Liver tumors were observed primarily in those groups of animals to which PBB was given in doses sufficient to induce readily observable hepatic toxicity. Under the conditions of this experiment, polybrominated biphenyl mixture (Firemaster FF-1) was carcinogenic for Fischer 344 rats and B6C3F1 mice of both sexes. Lesions included neoplastic nodules, hepatocellular carcinomas, and cholangiocarcinomas in rats and hepatocellular carcinomas in mice. Other manifestations of toxicity included porphyrogenic effects and hepatotoxicity. A significantly higher incidence of chronic progressive nephropathy was observed in male rats of the 1.0, 3.0, and 10.0 mg/kg dosage groups when compared with control males. Gastric ulcers and hyperplastic gastropathy of the glandular portion of the stomach were observed more frequently in male rats, primarily in the high dosage groups.     

Toxicity and carcinogenicity studies of nalidixic acid in rodents

Toxicity and carcinogenicity studies of nalidixic acid, an antimicrobial agent used to treat bacterial infections of the urinary tract, were conducted in F344/N rats and B6C3F1 mice of each sex for 13 weeks or 2 years. In the 13-week studies, nalidixic acid was administered at dietary concentrations ranging from 1,000 to 16,000 ppm. Body weights of both rats and mice were reduced in the groups receiving diet containing 8,000 and 16,000 ppm, and feed consumption of rats in the highest treatment groups was approximately two-thirds that of controls. Degeneration of the germinal epithelium in the seminiferous tubules of the testis was observed in male rats that received 16,000 ppm; no other compound-related histopathologic effects were observed in either species. Two-year studies were conducted by feeding diets containing 0, 2,000, or 4,000 ppm nalidixic acid to groups of 50 rats and mice/sex/group. The average daily feed consumption was slightly reduced compared to control groups and resulted in approximate daily doses of 82 or 175 mg nalidixic acid/kg for low dose and high dose rats, and 220 or 475 mg/kg for low dose and high dose mice. Mean body weights of dosed rats and mice were lower than those of controls, except for groups of low dose female rats and male mice. The incidences of preputial gland neoplasms in dosed male rats and of clitoral gland neoplasms in dosed female rats were significantly increased compared to those in controls; responses in low dose groups were similar to those in high dose groups. There were decreased incidences of leukemia and mammary gland neoplasms in dosed female rats and of pituitary gland neoplasms in dosed male rats. Subcutaneous tissue fibrosarcomas were marginally increased in dosed male mice. There were no increased incidences of neoplasms in dosed female mice. Under the conditions of these studies, the dietary administration of nalidixic acid was carcinogenic for rats, causing preputial gland or clitoral gland neoplasms in males and females, respectively. The association of subcutaneous neoplasms with administration of nalidixic acid to male mice was equivocal.     

Decreases in spontaneous tumors in rats and mice after treatment with amphetamine

Toxicology and carcinogenesis studies of dl-amphetamine sulfate, a drug used in the treatment of weight control, narcolepsy, and behavioral syndromes in children, were performed in F344/N rats and B6C3F1 mice. In these studies, amphetamine was administered for 2 years at doses of 0, 20, or 100 ppm in the feed to groups of 50 animals/dose/sex/species. The average amount of amphetamine consumed per day was estimated to be 1 or 5 mg/kg for low or high dose rats, 4 or 30 mg/kg for low or high dose male mice, and 3 or 19 mg/kg for low or high dose female mice. Survival was similar in dosed and control groups. The most notable effect of long-term treatment with this drug was the reduction of body weight in comparison to controls, and reduction in spontaneous tumors including pheochromocytomas of the adrenal gland in male rats, fibroadenomas of the mammary gland in female rats, adenomas of the anterior pituitary gland in male and female rats and female mice, endometrial stromal polyps of the uterus of female rats, adenomas or carcinomas of the liver in male and female mice, adenomas of the Harderian gland in male and female mice, and adenomas or carcinomas of the lung in male and female mice. Decreases in spontaneous tumors have previously been seen in 2-year rodent studies in groups of animals that have a reduced body weight in comparison to controls, but the spectrum of reduction in spontaneous neoplasms after treatment with amphetamine is broader than has previously been observed.     

Toxicity and Carcinogenicity Studies of Nalidixic Acid in Rodents

ABSTRACT

Toxicity and carcinogenicity studies of nalidixic acid, an antimicrobial agent used to treat bacterial infections of the urinary tract, were conducted in F344/N rats and B6C3F mice of each sex for 13 weeks or 2 years. In the 13–week studies, nalidixlc acid was administered at dietary concentrations ranging from 1,000 to 16,000 pp. Body weights of both rats and mice were reduced in the groups receiving diet containing 8,000 and 16,000 ppm, and feed consumption of rats in the highest treatment groups was approximately tw-thirds that of controls. Degeneration of the germinal epithelium in the seminiferous tubules of the testis was observed in male rats that received 16,000 ppm; no other compound-related histopathologic effects were observed in either species. TWo-year studies were conducted by feeding diets containing 0, 2,000, or 4,000 ppm nalidixic acid to groups of 50 rats and mice/sex/group. The average daily feed consumption was slightly reduced compared to control groups and resulted in approximate daily doses of 82 or 175 mg nalidixic acidfig for low dose and high dose rats, and 220 or 475 mg/kg for low dose and high dose mice. Mean body weights of dosed rats and mice were lower than those of controls, except for groups of low dose female rats and male mice. The incidences of preputial gland neoplasms in dosed male rats and of clitoral gland neoplasms in dosed female rats were significantly increased compared to those in controls; responses in low dose groups were similar to those in high dose groups. There were decreased incidences of leukemia and mammary gland neoplasms in dosed female rats and of pituitary gland neoplasms in dosed male rats. Subcutaneous tissue fibrosarcomas were marginally increased in dosed male mice. There were no increased incidences of neoplasms in dosed female mice. under the conditions of these studies, the dietary administration of nalidixic acid was carcinogenic for rats, causing preputial gland or clitoral gland neoplasms in males and females, respectively. The association of subcutaneous neoplasms with administration of nalidixic acid to male mice was equivocal.     

Toxicity and carcinogenicity of hydroquinone in F344/N rats and B6C3F1 mice.

Toxicology and carcinogenesis studies were conducted by administering hydroquinone (more than 99% pure) by gavage to groups of F344/N rats and B6C3F1 mice of each sex for 14 days, 13 wk or 2 yr. 14-day studies were conducted by administering hydroquinone in corn oil to rats at doses ranging from 63 to 1000 mg/kg body weight and to mice at doses ranging from 31 to 500 mg/kg, 5 days/wk. In the 13-wk studies, doses for rats and mice ranged from 25 to 400 mg/kg. At those doses showing some indication of toxicity in the 14-day and 13-wk studies, the central nervous system, forestomach and liver were identified as target organs in both species and renal toxicity was observed in rats. Based on these results, 2-yr studies were conducted by administering 0, 25 or 50 mg hydroquinone/kg in deionized water by gavage to groups of 65 rats of each sex, 5 days/wk. Groups of 65 mice of each sex were given 0, 50 or 100 mg/kg on the same schedule. 10 rats and 10 mice from each group were killed and evaluated after 15 months. Mean body weights of high-dose male rats and high-dose mice were approx. 5-14% lower than those of controls during the second half of the study. No differences in survival were observed between dosed and control groups of rats or mice. Nearly all male rats and most female rats in all vehicle control and exposed groups had nephropathy, which was judged to be more severe in high-dose male rats. Hyperplasia of the renal pelvic transitional epithelium and renal cortical cysts were increased in male rats. Tubular cell hyperplasia of the kidney was seen in two high-dose male rats, and renal tubular adenomas were seen in 4/55 low-dose and 8/55 high-dose male rats; none was seen in vehicle controls or in female rats. Mononuclear cell leukaemia in female rats occurred with increased incidences in the dosed groups (vehicle control, 9/55; low dose, 15/55; high dose, 22/55). Compound-related lesions observed in the liver of high-dose male mice included anisokaryosis, syncytial alteration and basophilic foci. The incidences of hepatocellular neoplasms, primarily adenomas, were increased in dosed female mice (3/55; 16/55; 13/55). Follicular cell hyperplasia of the thyroid gland was increased in dosed mice.(ABSTRACT TRUNCATED AT 400 WORDS)     

Carcinogenic potential of o-nitrotoluene and p-nitrotoluene

The potential of o-nitrotoluene and p-nitrotoluene to cause cancer in mammalian species was studied in male and female F344/N rats and B6C3F1 mice. These chemicals are on the EPA list of high production chemicals and there is potential for human exposure (High Production Volume Chemical List (2000) http://oaspub.cpa.gov/opptintr/chemrtk/volchall.htm.). o-Nitrotoluene, administered in the feed for up to 2 years, caused clear evidence for cancer at multiple sites in rats and mice. Male rats, receiving o-nitrotoluene in the feed ( approximately 0, 25, 50, or 90 mg/kg per day), developed treatment-related mesotheliomas, subcutaneous skin neoplasms, mammary gland fibroadenomas, and liver neoplasms. By 2 years, mesotheliomas, skin, liver, mammary gland and liver tumors also occurred in 'stop-study' male rats that received o-nitrotoluene at 125 or 315 mg/kg per day for only the first 3 months of study. These 'stop-studies' showed that the critical events leading to tumor formation occurred after 3 months of dosing, and these events were irreversible and eventually led to cancer at multiple sites. o-Nitrotoluene given in the feed to female rats (approximately 0, 30, 60, or 100 mg/kg per day) and to male and female mice (approximately 0, 150, 320, or 700 mg/kg per day) also caused a carcinogenic response. In female rats, treatment-related subcutaneous skin neoplasms and mammary gland fibroadenomas occurred. Hemangiosarcomas and carcinomas of the large intestine (cecum) were seen in treated male and female mice. In contrast to o-nitrotoluene, p-nitrotoluene given in the feed over approximately the same exposure levels caused only equivocal evidence of carcinogenic activity in male rats (subcutaneous skin neoplasms); some evidence of carcinogenic activity in female rats (clitoral gland neoplasms); equivocal evidence of carcinogenic activity in male mice (lung neoplasms); and no evidence of carcinogenic activity in female mice. Differences in the o-nitrotoluene and p-nitrotoluene carcinogenic activity may be due to differences in the metabolism of the parent compound to carcinogenic metabolites.     

Carcinogenicity of dermally administered 1,2-dihydro-2,2,4-trimethylquinoline monomer in F344 rats and B6C3F1 mice

1,2-Dihydro-2,2,4-trimethylquinoline (TMQ) was evaluated in a 2-year study in which groups of 60 male or female F344 rats received 0, 36 or 60 mg kg(-1) (0, 0.022, or 0.037 mg cm(-2)) and groups of 60 male or female B6C3F1 mice received 0, 3.6 or 10 mg kg(-1) (0, 0.00136, 0.00435 mg cm(-2)) in acetone by topical administration. Survival of all treated groups was comparable to survival of controls. Mean body weights of female rats were lower than those of controls throughout the study but mean body weights of male rats and male and female mice were comparable to the mean body weights of controls. No neoplasms of the skin were observed in any group of rats or mice. Acanthosis at the site of application was increased in male and female rats that received 60 or 100 mg kg(-1) and hyperkeratosis was increased in female rats that received 60 mg kg(-1). The incidences of renal tubule adenoma and renal tubule adenoma or carcinoma were increased significantly in the 60 and 100 mg kg(-1) groups of male rats. There were no neoplastic or non-neoplastic lesions in mice associated with exposure to 1,2-dihydro-2,2,4-trimethylquinoline. In a 1-year initiation-promotion study, groups of 30 female SENCAR mice received an initiating dose of 50 mg kg(-1) 1,2-dihydro-2,2,4-trimethylquinoline followed by promotion with 12-O-tetradecanoylphorbol-13-acetate (TPA), or an initiating dose of 7,12-dimethylbenzanthracene (DMBA) followed by promotion with 5, 10 or 25 mg kg(-1) 1,2-dihydro-2,2,4-trimethylquinoline. Other groups served as initiator control, promoter control, vehicle control and positive control (DMBA initiation, TPA promotion). In this system, 1,2-dihydro-2,2,4-trimethylquinoline-initiated skin was not promoted by TPA, and DMBA-initiated skin was not promoted by 1,2-dihydro-2,2,4-trimethylquinoline.     

The hepatic endothelial carcinogen riddelliine induces endothelial apoptosis,mitosis, S phase,and p53 and hepatocytic vascular endothelial growth factor expression after short-term exposure

Riddelliineis a naturally occurring pyrrolizidine alkaloid found in certain poisonous rangeland plants of the western United States. In National Toxicology Program 2-year studies, riddelliine induced high incidences of hemangiosarcoma in the liver of F344/N rats (both sexes) and B6C3F1 mice (males). To understand this pathogenesis, we tested short-term effects of riddelliine. Three groups (control; 1.0 mg/kg/day, high dose used in the 2-year study; and 2.5 mg/kg/day) of seven male F344/N rats per group were terminated after 8 consecutive doses and 30 doses (6 weeks, excluding weekends). Serum vascular endothelial growth factor (VEGF), histological, immunohistochemical [factor VIII-related antigen/von Willebrand factor (fVIII-ra/vWf)], VEGF, VEGF receptor-2 (VEGFR2), glutathione S-transferase-pi, S-phase (BrdU), p53, apoptosis, and ultrastructural evaluations were performed on the liver. Following 8 doses of 1.0 and 2.5 mg/kg/day, increased numbers of apoptotic and S-phase nuclei appeared in hepatocytes and endothelial cells. Following 30 doses of 1.0 and 2.5 mg/kg/day, hepatocytes exhibited reduced mitosis, fewer S-phase nuclei, increased hypertrophy, and fatty degeneration, while endothelial cells showed karyomegaly, cytomegaly, decreased apoptosis, more S-phase nuclei, and p53 positivity. Hepatocytes of treated animals expressed higher VEGF immunopositivity. That altered endothelial cells were fVIII-ra/vWf and VEGFR2 positive confirmed their identity. These changes may have promoted hemangiosarcoma development upon long-term exposure through endothelial adduct formation, apoptosis, proliferation of endothelial cells having undamaged and/or damaged DNA, and mutation. Endothelial proliferation may also have been promoted through endothelial arrest at S phase, which was associated with endothelial karyo- and cytomegaly, resulting in hepatocytic hypoxia, triggering VEGF induction.     

Tumor induction in F344/N rats and B6C3F1 mice following inhalation exposure to ethylbenzene

Carcinogenesis studies of ethylbenzene were conducted because of its extensive use as a solvent and because it is structurally similar to the known carcinogen benzene. Groups of 50 male and 50 female Fischer rats and B6C3F1 mice were exposed to ethylbenzene by inhalation at 0, 75, 250, and 750 ppm 6 h per day, 5 days per week, for 2 years. The dose levels were selected based on the results of 13-week studies. In the 750 ppm group of male and female rats, body weights were slightly lower and incidences of renal hyperplasia and tubular neoplasms were significantly increased compared with controls. Incidence of testicular tumors was also significantly increased in male rats. Survival and body weights of the exposed groups of male and female mice and controls were comparable. Incidences of alveolar epithelium metaplasia, alveolar/bronchiolar adenoma, and hepatocyte hypertrophy and necrosis were significantly increased in the 750 ppm male mice and incidences of liver eosinophilic foci and hepatocellular neoplasms were significantly increased in the 750 ppm female mice compared with controls. Ethylbenzene is carcinogenic inducing neoplasms in kidneys and testes in Fischer rats and in lungs in male and liver in female B6C3F1 mice.     

Toxicity and Carcinogenicity of 2,3-Dibromo-1-propanol in F344/N Rats and B6C3F1 Mice

2,3-Dibromo-1-propanol is a metabolite of the flame retardanttris(2,3-dibromopropyl) phosphate, previously shown to be amutagen and carcinogen in experimental animals. Toxicology andcarcinogenesis studies of 2,3-dibromo-1-propanol were conductedby applying the chemical in 95% ethanol to the interscapularskin of male and female F344/N rats and B6C3F₁ mice 5 days aweek for 13 weeks in the prechronic study and 48–55 weeks(rats) or 36–42 weeks (mice) in the carcinogenicity study.In the 13-week study, 10 rats and 10 mice of each sex receiveddoses of 0, 44, 88, 177, 375, or 750 mg/kg. Deaths associatedwith chemical application occurred only in the high-dose (750mg/kg) male mice. Chemical-related lesions were seen in thekidney of male rats, liver of female rats, and liver and lungof both sexes of mice. Based on the toxicity observed in the13-week study, 50 rats of each sex received doses of 0, 188,or 375 mg/kg and 50 mice of each sex received 0, 88, or 177mg/kg in the carcinogenicity study. The planned 2-year studywas terminated early because of reduced survival of rats relatedto chemical-induced neoplasia and because of the appearanceof antibodies to lymphocytic choriomeningitis virus in sentinelmice. Nearly all dosed rats had malignant neoplasms at one ormore sites, while only one control male and one control femalehad malignant neoplasms. In rats, neoplasms induced by 2,3-dibromo-1-propanoloccurred in the skin, nasal mucosa, Zymbal's gland, oral mucosa,esophagus, forestomach, intestines, liver, kidney, mammary gland(females), clitoral gland (females), spleen (males), and mesothelium(males). In mice, chemical-induced neoplasms occurred in theskin, forestomach, liver (males), and lung (males).     

NTP technical report on the toxicology and carcinogenesis studies of Elmiron (Cas No. 37319-17-8) in F344/N rats and B6C3F1 mice (Gavage Studies)

[structure--see text] Elmiron, a white powder, is the sodium salt of pentosan polysulfate, a semisynthetic sulfated polyanion composed of beta-D-xylopyranose residues with biological properties similar to heparin. Elmiron is used in the United States for the relief of urinary bladder pain associated with interstitial cystitis. Because of its stimulating effect on fibrinolysis, Elmiron has been used clinically in the treatment and prevention of thrombotic disorders. The United States Food and Drug Administration nominated Elmiron for toxicology and carcinogenicity testing by the National Toxicology Program because of its orphan drug status. Male and female F344/N rats and B6C3F1 mice received Elmiron, which met product specifications provided by the manufacturer, in deionized water by gavage for 2 weeks, 3 months, or 2 years. Genetic toxicology studies were conducted in Salmonella typhimurium, rat and mouse bone marrow cells, and mouse peripheral blood erythrocytes. 2-WEEK STUDY IN RATS: Groups of five male and five female rats were administered 0, 33, 111, 333, 1,000, or 3,000 mg Elmiron/kg body weight in deionized water by gavage, 5 days per week, for 16 days. Elmiron administration had no effect on survival or body weight gain. Activated partial thromboplastin time was significantly increased in 3,000 mg/kg rats. Liver weights of 3,000 mg/kg rats were significantly greater than those of the vehicle controls. Hepatocellular cytoplasmic vacuolization occurred in all 3,000 mg/kg females. 2-WEEK STUDY IN MICE: Groups of five male and five female mice were administered Elmiron in deionized water by gavage at doses of 0, 33, 111, 333, 1,000, or 3,000 mg/kg, 5 days per week, for 16 days. All mice survived to the end of the study. Mean body weight gains of male mice administered 333 mg/kg or greater were significantly greater than that of the vehicle control group. Liver weights of 1,000 and 3,000 mg/kg males were significantly increased. 3-MONTH STUDY IN RATS: Groups of 10 male and 10 female rats were administered Elmiron in deionized water by gavage at doses of 0, 63, 125, 250, 500, or 1,000 mg/kg, 5 days per week, for 14 weeks. No deaths were attributed to administration of Elmiron. Mean body weights of 125 mg/kg males were less than those of vehicle controls and the mean body weights of all dosed groups of females were greater. Hematology results indicated that Elmiron, at the doses selected, induced a minimal erythron decrease and leukocyte and platelet count increases that may have been secondarily related to the inflammatory lesions observed in various tissues of rats. Liver and spleen weights of males administered 250 mg/kg or greater were significantly increased. Liver weights of all dosed groups of females, and kidney, lung, and spleen weights of 1,000 mg/kg females were significantly increased. Histiocytic cellular infiltration, chronic active inflammation, and ulcers of the rectum occurred in most 500 and 1,000 mg/kg rats. Administration of Elmiron was associated with the presence of vacuolated histiocytes in the mandibular and mesenteric lymph nodes, lung, kidney, and liver of male and female rats. Histochemical investigations of the vacuolated histiocytes indicated the presence of neutral and acidic mucins and lipid material within the vacuoles. Transmission electron microscopy identified these vacuoles as lysosomal structures that exhibited a variety of contents. 3-MONTH STUDY IN MICE: Groups of 10 male and 10 female mice were administered Elmiron in deionized water by gavage at doses of 0, 63, 125, 250, 500, or 1,000 mg/kg, 5 days per week, for 14 weeks. One 250 mg/kg female mouse was sacrificed moribund on day 84; all other mice survived to the end of the study. Mean body weights of dosed groups were similar to those of the vehicle control groups. Hematology results indicated that Elmiron, at the doses selected, induced a minimal erythron decrease and leukocyte and platelet count increases that may have been secondarily related to the inflammatory lesions observed in various tissues of mice. in various tissues of mice. Liver weights of 500 mg/kg males and 1,000 mg/kg males and females, and spleen weights of 1,000 mg/kg males were significantly increased. Histiocytic cellular infiltration and chronic active inflammation of the rectum occurred in most 1,000 mg/kg mice. Administration of Elmiron was associated with the presence of vacuolated histiocytes in the mandibular and mesenteric lymph nodes, liver, and spleen of males and females. Histochemical investigations of the vacuolated histiocytes indicated the presence of neutral and acidic mucins within the vacuoles. Transmission electron microscopy identified these vacuoles as lysosomal structures that exhibited a variety of contents. 2-YEAR STUDY IN RATS: Groups of 50 males and 50 females were administered Elmiron in deionized water by gavage at doses of 0, 14, 42, or 126 mg/kg to males and 0, 28, 84, or 252 mg/kg to females, 5 days per week, for 104 or 105 weeks. Survival of all dosed groups of rats was similar to that of the vehicle control groups. Mean body weights of all dosed groups were similar to those of the vehicle controls throughout the 2-year study. Microscopically, myxomatous changes were present in the rectum of 56% of 126 mg/kg males and 83% of 252 mg/kg females. The incidences of chronic active focal alveolar inflammation of the lung were increased in all dosed groups. The incidences of histiocytic cellular infiltration of the mesenteric lymph nodes were increased in 42 and 126 mg/kg males and in 84 and 252 mg/kg females, and lymphohistiocytic hyperplasia was present in the spleen of 126 mg/kg males and 252 mg/kg females. 2-YEAR STUDY IN MICE: Groups of 50 males and 50 females were administered Elmiron in deionized water by gavage at doses of 0, 56, 168, or 504 mg/kg, 5 days per week, for 104 or 105 weeks. Survival of all dosed groups of mice was similar to that of the vehicle control groups. Mean body weights of males were similar to those of vehicle controls. Mean body weights of 504 mg/kg females were progressively less than those of the vehicle controls during the second year of the study. Increased incidences of hemangiosarcomas of the liver and hepatocellular neoplasms were observed in male and female mice. The incidences of hemangiosarcomas in the 504 mg/kg groups exceeded the historical control ranges for males and females; both the trend and the incidence in the 504 mg/kg groups were significant for males. Hemangiosarcomas in males and females were attributed to Elmiron administration. The incidence of hepatocellular adenoma in 504 mg/kg females was significantly increased and exceeded the historical control range; the trends for hepatocellular adenoma and for hepatocellular adenoma or carcinoma (combined) were also significant in females and were attributed to Elmiron administration. There was also a marginal increase in the incidences of hepatocellular neoplasms in male mice, which may have been associated with Elmiron administration. Malignant lymphomas occurred with a positive trend in female mice; the incidence in the 504 mg/kg group was also significantly increased and matched the upper limit of the historical control range. These malignant lymphomas may have been associated with Elmiron administration. Nonneoplastic lesions related to the administration of Elmiron occurred in the liver, rectum, mesenteric lymph node, and spleen of 504 mg/kg mice and to a lesser extent in 168 mg/kg mice. These lesions were similar to those observed in the 3-month study. GENETIC TOXICOLOGY: Elmiron was not mutagenic in S. typhimurium strains TA97, TA98, TA100, or TA1535 with or without induced hamster or rat liver S9 enzymes. No increases in the frequency of micronucleated polychromatic erythrocytes were seen in bone marrow cells of rats or mice administered Elmiron by gavage three times at 24-hour intervals. No significant alterations in the frequency of micronucleated normochromatic erythrocytes were seen in peripheral blood samples from male or female mice administered Elmiron for 3 months by gavage. CONCLUSIONS: Under the conditions of these 2-year gavage studies, there was no evidence of carcinogenic activity of Elmiron in male F344/N rats administered 14, 42, or 126 mg/kg or in female F344/N rats administered 28, 84, or 252 mg/kg. There was some evidence of carcinogenic activity of Elmiron in male B6C3F1 mice based on increased incidences of liver hemangiosarcoma. The increased incidences of hepatocellular neoplasms in male mice may have been related to Elmiron administration. There was some evidence of carcinogenic activity of Elmiron in female B6C3F1 mice based on the increased incidences of liver hemangiosarcoma and hepatocellular neoplasms. The increased incidences of malignant lymphomas in female mice may have been related to Elmiron administration. Elmiron administration caused increased incidences of nonneoplastic lesions (presence of vacuolated histiocytes) of the rectum, lung, mesenteric lymph node, and spleen (males) in rats and of the liver, rectum, mesenteric lymph node, and spleen in mice.     

Toxicity and carcinogenicity studies of phenylephrine hydrochloride in F344/N rats and B6C3F1 mice

Phenylephrine HCl was incorporated into feed given to male and female F344/N rats and B6C3F1 mice in studies of 14 days, 12 weeks, and 2 years duration. In 12-week studies, body weight gains decreased with dose, and deaths of male rats and mice occurred at concentrations of 5,000 ppm and above; however, no organ-specific toxicity was evident. During 2-year studies, body weights of rats receiving diets at 620 and 1,250 ppm and mice at 1,250 and 2,500 ppm ranged up to 16% less than control. Survival of high dose male rats was substantially greater than controls. Survivals of other dose groups of rats and mice were similar to controls. Chronic focal inflammation of the liver, and inflammation of the prostate were increased in dosed rats. No increases in neoplasms were observed in rats or mice consuming diets containing phenylephrine HCl for 2 years. The incidences of mononuclear cell leukemia and pheochromocytomas of the adrenal gland were decreased in dosed male rats. Approximate time weighted average doses ranged up to 54 mg/kg/day for rats and 280 mg/kg/day for mice during the 2-year studies.     

Hyponeophagia and arousal in rats: Effects of diazepam, 5-methoxy-N,N-dimethyltryptamine,d-amphetamine and food deprivation

A modified hyponeophagia test is described as an animal model of anxiety. The effects of 0, 0.3, 1.0, 3.0 and 10 mg/kg diazepam, given both acutely and for 7 days pretest, were assessed in rats. Acutely, diazepam reduced hyponeophagia over the dose range 0.3–3.0 mg/kg but 10.0 mg/kg produced sedation and large variability. Chronically, the dose-response relationships were monotonic and the maximal effect was increased, suggesting that differential tolerance occurs to the sedative, but not to the anxiolytic, effects of this drug. Increased food deprivation did not mimic benzodiazepine effects on hyponeophagia, and actually prolonged eating latency in rats treated with 5-methoxy-N,N-dimethyltryptamine (2.5 mg/kg), which does not support an interpretation of diazepam effects in terms of appetitive actions. An arousal hypothesis of hyponeophagia was proposed and supported by the antagonism of the sedative effects of 10.0 mg/kg diazepam by d-amphetamine (0.5 mg/kg). Although both male and female rats were used throughout, sex differences were few in these studies.     

Toxicology and carcinogenesis studies of dibromoacetic acid (Cas No. 631-64-1) in F344/N rats and B6C3F1 mice (drinking water studies)

Dibromoacetic acid is a water disinfection by-product. Dibromoacetic acid was nominated to the National Toxicology Program by the United States Environmental Protection Agency for toxicity and carcinogenicity studies in rats and mice because of widespread human exposure and because a related dihaloacetate, dichloroacetate, was found to be carcinogenic to the liver of rats and mice. Drinking water was selected as the route of exposure to mimic human exposure to this chemical. Male and female F344/N rats and B6C3F1 mice were exposed to dibromoacetic acid (greater than 99% pure) in drinking water for 2 weeks, 3 months, or 2 years. Genetic toxicology studies were conducted in Salmonella typhimurium and peripheral blood erythrocytes of exposed mice. 2-WEEK STUDY IN RATS: Groups of five male and five female rats were exposed to 0, 125, 250, 500, 1,000, or 2,000 mg/L dibromoacetic acid in drinking water for 2 weeks, equivalent to average daily doses of approximately 17, 32, 67, 134, 270 (males), or 257 (females) mg dibromoacetic acid/kg body weight. All rats survived to the end of the study. Mean body weight gains of 1,000 mg/L males and of 500 mg/L females were significantly greater than those of the controls. Water consumption by exposed and control groups was similar. Liver weights of exposed males and females were significantly increased. Right testis weights of males exposed to 500 mg/L or greater were significantly decreased. The incidences of hepatocytic cytoplasmic alteration were significantly increased in males exposed to 500 mg/L or greater and in 2,000 mg/L females. Testicular lesions, characterized by a delay in spermiation and retained spermatids, were noted in males exposed to 500 mg/L or higher concentrations. 2-WEEK STUDY IN MICE: Groups of five male and five female mice were exposed to 0, 125, 250, 500, 1,000, or 2,000 mg/L dibromoacetic acid (equivalent to average daily doses of approximately 24, 47, 95, 178, or 370 mg/kg to males and 22, 53, 88, 166, or 309 mg/kg to females) in drinking water for 2 weeks. All mice survived to the end of the study. Mean body weight gains of 250 and 500 mg/L males were significantly greater than those of the controls. Water consumption by exposed and control groups was similar. Liver weights of males and females in the 1,000 and 2,000 mg/L groups were significantly increased. Thymus weights of males and females in the 1,000 and 2,000 mg/L groups were significantly less than those of controls. The incidences of thymus atrophy were significantly increased in 1,000 and 2,000 mg/L males and 2,000 mg/L females. The incidences of morphological changes to the germinal epithelium of the testes were increased in males exposed to 1,000 or 2,000 mg/L. 3-MONTH STUDY IN RATS: Groups of 10 male and 10 female rats were exposed to 0, 125, 250, 500, 1,000, or 2,000 mg/L dibromoacetic acid (equivalent to average daily doses of approximately 10, 20, 40, 90, and 166 mg/kg to males and 12, 23, 48, 93, and 181 mg/kg to females) in drinking water for 3 months. All rats survived to the end of the study. Mean body weights of male and female rats in the 2,000 mg/L group were significantly less than those of controls. Water consumption by the 2,000 mg/L males at weeks 1 and 13 and by females at week 13 was less than that by controls. Small decreases in the erythron and platelet counts occurred in rats exposed to 2,000 mg/L; minimally impaired erythropoiesis was also seen in 1,000 mg/L rats. Liver weights of all exposed groups of males and females were significantly increased. Male rats in the 2,000 mg/L group had significantly decreased testis weights. Testicular atrophy was noted in the 2,000 mg/L group, and retained spermatids were observed in the 500 and 1,000 mg/L groups. In the pituitary gland of male rats exposed to 2,000 mg/L, the incidence of cellular hypertrophy was significantly increased. The incidences of hepatocellular vacuolization were significantly increased in males exposed to 500 mg/L or greater and in females exposed to 2,000 mg/L. Hematopoietic cell proliferation was noted in females in the 2,000 mg/L group. 3-MONTH STUDY IN MICE: Groups of 10 male and 10 female mice were exposed to 0, 125, 250, 500, 1,000, or 2,000 mg/L dibromoacetic acid (equivalent to average daily doses of approximately 16, 30, 56, 115, and 230 mg/kg to males and 17, 34, 67, 132, and 260 mg/kg to females) in drinking water for 3 months. All mice survived to the end of the study. Mean body weights and body weight gains of female mice in the 2,000 mg/L group and the mean body weight gain of 2,000 mg/L males were significantly less than those of controls. Water consumption by males in the 2,000 mg/L group was decreased at weeks 1 and 13 relative to controls. Small decreases in mean cell hemoglobin and platelet counts occurred in 2,000 mg/L male mice. Liver weights of males and females exposed to 500 mg/L or greater were significantly increased. Hepatocellular cytoplasmic vacuolization was present in most mice and the severity was increased in 1,000 and 2,000 mg/L males and females. The incidences of abnormal testicular morphology were significantly increased in 1,000 and 2,000 mg/L males. 2-YEAR STUDY IN RATS: Groups of 50 male and 50 female rats were exposed to drinking water containing 0, 50, 500, and 1,000 mg/L dibromoacetic acid for 2 years (equivalent to average daily doses of approximately 2, 20, and 40 mg/kg to males and 2, 25, and 45 mg/kg to females). Survival of exposed rats was similar to that of the control groups. Mean body weights of 1,000 mg/L males and females were less than those of the controls after weeks 29 and 53, respectively, and those of 500 mg/L males and females were less after weeks 57 and 85, respectively. Water consumption by males and females exposed to 1,000 mg/L was less than that by controls during year 2 of the study. The incidence of malignant mesothelioma was significantly increased in 1,000 mg/L male rats. A positive trend in the incidence of mononuclear cell leukemia occurred in female rats, and the incidence in 1,000 mg/L females was significantly increased. The incidences of mononuclear cell leukemia were increased in 50 and 500 mg/L males. The incidences of cystic degeneration of the liver were significantly increased in all exposed groups of male rats. The incidences of alveolar epithelial hyperplasia were significantly increased in 500 and 1,000 mg/L females, and the incidences of nephropathy were significantly increased in all exposed groups of females. 2-YEAR STUDY IN MICE: Groups of 50 male and 50 female mice were exposed to drinking water containing 0, 50, 500, and 1,000 mg/L dibromoacetic acid for 2 years (equivalent to average daily doses of approximately 4, 45, and 87 mg/kg to males and 4, 35, and 65 mg/kg to females). Survival of exposed mice was similar to that of the controls. Mean body weights of 50 and 500 mg/L male mice were greater than those of the controls after week 85. Water consumption by exposed mice was generally similar to that by controls throughout the study. The incidences of liver neoplasms occurred with positive trends in male and female mice. The incidences of multiple hepatocellular adenoma and hepatocellular adenoma or carcinoma (combined) were significantly increased in all exposed groups of males and in 500 and 1,000 mg/L females. The incidences of hepatoblastoma were significantly increased in 500 and 1,000 mg/L males, and the incidences of hepatocellular carcinoma were significantly increased in 1,000 mg/L males and 500 mg/L females. The incidences of alveolar/bronchiolar adenoma occurred with positive trends in males and females, and the incidence in 500 mg/L male mice was significantly greater than that in controls. GENETIC TOXICOLOGY: Dibromoacetic acid was mutagenic in Salmonella typhimurium strain TA100 with and without rat or hamster liver metabolic activation enzymes (S9); no activity was detected in strain TA98, with or without S9. Increased frequencies of micronucleated normochromatic erythrocytes were observed in peripheral blood samples from male, but not female, mice administered dibromoacetic acid in drinking water for 3 months. CONCLUSIONS: Under the conditions of these studies, there was some evidence of carcinogenic activity of dibromoacetic acid in male rats based on an increased incidence of malignant mesothelioma. The increased incidences of mononuclear cell leukemia in male rats may have been related to dibromoacetic acid exposure. There was some evidence of carcinogenic activity of dibromoacetic acid in female rats based on an increased incidence and positive trend of mononuclear cell leukemia. There was clear evidence of carcinogenic activity of dibromoacetic acid in male and female mice based on increased incidences of hepatocellular neoplasms and hepatoblastoma (males only). Increased incidences of lung neoplasms in male mice were also considered to be exposure related. The slight increased incidence of lung neoplasms in female mice may have been related to dibromoacetic acid exposure. Exposure to dibromoacetic acid for 2 years caused increased incidences of cystic degeneration of the liver in male rats, increased incidences of alveolar epithelial hyperplasia and nephropathy in female rats, and increased incidences of splenic hematopoiesis in male mice.     

Toxicity and carcinogenicity of Elmiron in F344/N rats and B6C3F<Subscript>1</Subscript> mice following 2 years of gavage administration

Elmiron (sodium pentosan polysulfate) is used for the relief of urinary bladder pain associated with interstitial cystitis. The National Toxicology Program (NTP) tested this compound because of its orphan drug status and lack of information about its chronic toxicity and carcinogenicity. Groups of 50 male and 50 female F344/N rats were given Elmiron in de-ionized water by gavage at doses of 0, 14, 42, or 126 mg/kg to males and 0, 28, 84, or 252 mg/kg to females once daily, 5 days per week, for up to 2 years. The same numbers of male and female B6C3F₁ mice were dosed similarly with 0, 56, 168, or 504 mg/kg. Elmiron administration produced no effect on the body weight of rats, male mice, or low- and mid-dose groups of female mice. The body weights of the high-dose female mice were significantly decreased relative to those of controls. Pairwise comparison showed that survival of all dosed groups of rats and mice was similar to that of the controls. Elmiron was not carcinogenic in F344/N rats. An increased incidence of liver hemangiosarcoma provided evidence of some carcinogenic activity for Elmiron in male B6C3F₁ mice. Increased incidences of liver hemangiosarcoma, hepatocellular neoplasms (predominantly adenomas), and malignant lymphomas revealed carcinogenic activity of Elmiron in female B6C3F₁ mice. Elmiron administration produced elevated occurrences of nonneoplastic lesions, such as vacuolated histiocytes in the rectum, lung, spleen (males only), and mesenteric lymph node in rats and liver, rectum, mesenteric lymph node, and spleen in mice. Myxomatous change, chronic inflammation, and squamous metaplasia (mice only) were observed in the large intestine, and lymphohistiocytic hyperplasia was found to be increased in the spleen of rats of both sexes treated with the highest dose. In the latter lesion, the histiocytes contained pale, finely granular cytoplasm and were not considered to represent the same change as the vacuolated histiocytes seen in the mesenteric lymph node and rectum. Under the conditions of these 2-year studies, Elmiron was carcinogenic to mice but not rats.The studies were conducted at Battelle, Columbus, OH, USA.     

Evaluation of the carcinogenic potential of clofibrate in the rasH2 mouse

The purpose of the study was to support of the International Life Sciences Institute (ILSI) alternative carcinogenicity models initiative to evaluate the carcinogenic potential of the nongenotoxic carcinogen, clofibrate, a peroxisome proliferator-activated receptor (PPAR) alpha agonist, following oral administration to rasH2 mice. Peroxisome proliferators are one of the most widely studied of the nongenotoxic carcinogens and have diverse industrial and therapeutic uses (Gonzalez et al. J. Nat. Cancer Inst. 90: 1702-1709, 1998); however, the nongenotoxic mechanism of carcinogenicity is currently unknown. Male mice were administered doses of clofibrate at 50, 100, or 200 mg/kg/day and female mice were administered doses of 50, 150, or 250 mg/kg/day by oral gavage at 10 ml/kg for 27 weeks. In addition, rasH2 male and female mice were treated with N-nitroso-N-methylurea (NMU). Nontransgenic male and female mice were treated with 200 and 250 mg/kg/day, respectively, of clofibrate. The NMU-treated mice were given a single intraperitoneal dose of 75 mg/kg, which was followed by a 90-day observation period; all others were sacrificed after 6 months of daily dosing. Hepatocellular neoplasms were observed in clofibrate-treated rasH2 male mice after 6 months of treatment but not in nontransgenic males or females. Clofibrate treatment (250 mg/kg/day) of female rasH2 mice was associated with a slight increase in the incidence of various neoplasms (harderian gland, lungs, skin, spleen, tail, thymus, and uterus) compared with untreated transgenic mice and with similarly treated nontransgenic mice. Non-neoplastic changes were found in the liver of transgenic and nontransgenic mice of both sexes and in the kidneys of male mice. NMU produced findings are consistent with previous studies. The data suggest that the rasH2 mice are a good model for testing epigenetic carcinogens in a shorter timeframe than conventional mouse carcinogenicity bioassays.     

Repeated dose 28-day oral toxicity study in Wistar rats with a mixture of five pesticides often found as residues in food: alphacypermethrin, bromopropylate, carbendazim, chlorpyrifos and mancozeb.

Six dose groups of 8 male and female rats respectively received a daily dose equivalent to 0, 0.15, 0.006, 0.03, 0.15 or 0.3 mg/kg b.w./day chlorpyrifos (groups 1-6) and the last four dose groups (groups 3-6) received in addition daily doses equivalent to 18 mg/kg b.w./day alphacypermethrin, 30 mg/kg b.w./day bromopropylate, 45 mg/kg b.w./day carbendazim and 12.5 mg/kg b.w./day mancozeb for 28 days. Plasma acetylcholinesterase was significantly decreased in the groups 2, 5 and 6 males. Total white blood cell count was significantly lower in females of group 6. Total red blood cell count, haematocrite and haemoglobin concentration was significantly reduced in both male and female rats of groups 5 and 6. Relative liver weight was significantly increased in groups 3-6 male and female rats. Absolute thyroid gland weight was significantly increased in groups 3, 5 and 6 male rats and of groups 3-6 female rats, and relative thyroid gland weight was significantly increased in groups 2-6 male rats and of groups 3-6 female rats. Absolute thymus weight of groups 3-6 male and female rats and relative thymus weight of groups 3-6 male rats and groups 3 and 4 female rats was significantly decreased. A mild degree of centrilobular cell hypertrophy of the liver was seen in all male rats and of three female rats of group 6. In the thyroid gland follicular cell hypertrophy was present in one female in the control group and in six females and seven males of group 6. It was concluded that inhibition of acetylcholinesterase activity in plasma and brain by chlorpyrifos was not enhanced by co-administration of the other four pesticides. Effects were seen in liver, thyroid, thymus and blood in the combination groups. However, identification of the pesticide(s) responsible for these changes would require further studies of the individually pesticides as well as various combinations of the pesticides.