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Carcinogenicity

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Description of key information

Carcinogenicity Study (OECD 451), mouse:

NOAEL (general) = 100 ppm (equivalent to 13.6 and 16.7 mg/kg bw/day in males and females, respectively)

NOAEL (carcinogenicity) = 4000 ppm (equivalent to 560 and 713 mg/kg bw/day in males and females, respectively)

Carcinogenicity Study (OECD 453), rat:

NOAEL (general) = 25 ppm (equivalent to 1.1 and 1.4 mg/kg bw/day in males and females, respectively)

NOAEL (carcinogenicity) = 2500 ppm (equivalent to 104 and 140 mg/kg bw/day in males and females, respectively)

Key value for chemical safety assessment

Carcinogenicity: via oral route

Endpoint conclusion
Endpoint conclusion:
adverse effect observed
Dose descriptor:
NOAEL
41 mg/kg bw/day
Study duration:
chronic
Species:
rat
Quality of whole database:
The available information comprises adequate and reliable studies (Klimisch score 1), and is thus sufficient to fulfil the standard information requirements set out in Annex X, 8.9.1, of Regulation (EC) No 1907/2006.

Carcinogenicity: via inhalation route

Endpoint conclusion
Endpoint conclusion:
no study available

Carcinogenicity: via dermal route

Endpoint conclusion
Endpoint conclusion:
no study available

Justification for classification or non-classification

The available data on the carcinogenicy potential of 5-hydroxy-1,3-dimethylpyrazol-4-yl)(α,α,α-trifluoro-2-mesyl-p-tolyl)methanone do

not meet the criteria for classification according to Regulation (EC) 1272/2008, and are therefore conclusive but not sufficient for classification.

Additional information

Reliable studies to assess the carcinogenetic potential of 5-hydroxy-1,3-dimethylpyrazol-4-yl)(α,α,α-trifluoro-2-mesyl-p-tolyl)methanone are available in mice and rats.

 

Carcinogenicity study in the C57BL/6J mouse by dietary administration

In this GLP-conform study conducted according to OECD 451, 5-hydroxy-1,3-dimethylpyrazol-4-yl)(α,α,α-trifluoro-2-mesyl-p-tolyl)methanone was incorporated into rodent diet and administered at concentrations of 0, 100, 1000, and 4000 ppm to groups of 60 male C57BL/6J mice (M-267521-01-2). Groups of 60 females received 0, 100, 1000, and 6000 ppm for the first 10 weeks of the study. However, mortality at 6000 ppm was considered to be excessive and this dose was decreased to 4000 ppm from week 11 onwards. These concentrations resulted in doses of 0, 13.6, 137, and 560 mg/kg bw/day for males and 0, 16.7, 168, and 713 mg/kg bw/day for females. Clinical signs and mortality were monitored twice daily on weekdays and once daily on weekends and holidays. Body weights were measured on the first day of treatment, weekly for the first 13 weeks of the study, and then at 4-week intervals through the end of the study. Ophthalmological examinations were conducted on all animals during the acclimatization phase and on all surviving animals at the end of the study. At 3, 6, 9, and 12 months, ophthalmoscopy was conducted on 24 animals per sex and treatment group. At the end of the one-year treatment period, blood was collected for hematology from the surviving animals in the one-year sacrifice group and from the first 10 surviving animals of the terminal sacrifice group. At the end of the study, blood was collected from the first 20 surviving animals of the terminal sacrifice group. At both time points, blood was collected after overnight fasting. At the 12-month and terminal sacrifices, a gross necropsy was conducted, selected organs were weighed, and organ and tissue samples were taken for histopathological examination (terminal sacrifice only).

In all sections, the female high dose group will be referred to as having received the test substance at a dietary concentration of 4000 ppm, although these animals received a dietary concentration of 6000 ppm for the first 10 weeks of the study.

There was a treatment-related increase in mortality in both males and females at 4000 ppm, but mortality at 100 and 1000 ppm were similar to observations in control animals. There were a number of treatment-related clinical signs, most related to the excretion of parent compound in the urine. These included hardness in the area of the urinary bladder, soiled fur, reduced motor activity, labored or rapid respiration, and red urine. Body weight and body weight gain were reduced at 4000 ppm in both males and females. Body weight was unaffected at 1000 ppm in males, however body weight gain was statistically significantly decreased compared to controls at a number of time points. As the males in the 1000 ppm treatment group were slightly heavier at the beginning of the study than their concurrent controls, decreased body weight gain over the course of the study resulted in similar terminal body weights. This effect was considered to be test substance-related. There was no biologically significant effect on body weight or body weight gain in females at 1000 ppm or on either males or females at 100 ppm. There was no effect of treatment on food consumption at any dose level.

Neither males nor females showed any treatment-related ophthalmological findings at any dose level. Red cell count, hemoglobin, hematocrit, and mean corpuscular hemoglobin concentration were decreased in females at 4000 ppm at 18 months, with decreases in most parameters also at 12 months. Mean corpuscular volume was slightly increased in females at 4000 ppm at 18 months. In males at 4000 ppm, similar hematological effects were generally only observed at 18 months. These findings are considered to be treatment-related.

There was no effect of treatment on terminal body weight at the 12-month sacrifice. After 18 months, however, there was a treatment-related decrease in terminal body weight at 4000 ppm in both males and females. At 12 months, males at 4000 ppm showed statistically significantly increased absolute and relative kidney weight compared to controls. At 18 months, absolute and relative liver weights at 1000 and 4000 ppm were increased in males with a slight effect in females, and absolute and relative kidney weights were increased at 4000 ppm in males only. There were no other treatment-related effects on organ weights.

Prior to the 12-month sacrifice, one male and three females out of the animals dedicated to the 12-month satellite group at 4000 ppm were found dead (male on day 240, females on days 60, 61, and 75). All of these animals had yellow urinary bladder stones. In the 12-month satellite animals sacrificed as scheduled, one male and four out of 7 females at 4000 ppm were found to have stone(s) in the renal pelvis. Both males and females at 4000 ppm showed enlarged or small kidneys, dilation of the renal pelvis, pale kidneys, or renal cyst(s). Stones or gritty content were also observed in the urinary bladders of nearly all animals sacrificed at 4000 ppm at 12 months, along with bladder distension in the majority of these animals. Gallbladder concretions were observed at 100 and 1000 ppm in males and 1000 and 4000 ppm in females, among animals sacrificed as scheduled at 12 months. Among the animals in the 18-month study group, the majority of those which died unscheduled at 4000 ppm were found to have died due to acute or chronic renal failure, due to urinary tract blockage or chronic kidney and/or urinary bladder inflammation, respectively. Stones were found in the kidney and/or urinary bladder of these animals; other findings at necropsy of unscheduled deaths were enlarged or small kidneys, renal pelvic dilation, pale kidneys, renal cyst(s), distension of the urinary bladder, and gallbladder concretions. Similar findings were observed in animals sacrificed on schedule at 18 months.

Microscopic examination was not conducted at 12 months. At 18 months, histopathologic findings included increased incidence in males and females at 1000 and 4000 ppm of centrilobular hepatocellular hypertrophy. In both males and females at all doses but in the absence of a dose relationship, there was a slight increase in the incidence of gall bladder epithelial hyperplasia. Treatment-related histopathological findings in the urinary tract were limited to 4000 ppm in bothmales and females and included epithelial hyperplasia, fibrosis and scarring, and inflammatory cell infiltration. These findings were related to the urinary tract calculi observed at this dose level in the majority of animals. Separate analysis of the gallstones revealed them to be composed primarily of cholesterol.

Treatment-related neoplastic findings at 4000 ppm were limited to the urinary tract of males and females, and were comprised of transitional cell carcinomas and papillomas of the urinary bladder and urethra. In all cases, these tumors were related to the presence of urinary tract stones. These tumors were considered to be the result of a non-genotoxic proliferative mechanism due to the concurrent presence of secondary inflammation and hyperplastic findings in the same tissues, induced by the urinary stones.

 

Conclusions

Administration of 5-hydroxy-1,3-dimethylpyrazol-4-yl)(α,α,α-trifluoro-2-mesyl-p-tolyl)methanone to male and female mice via dietary incorporation, at 4000 ppm, led to urinary tract stone formation, increased mortality generally due to either acute or chronic renal failure secondary to urinary tract stone development, decreased body weight and body weight gain, increased kidney and liver weights in males, renal pelvis dilation, and urothelial hyperplasia throughout the urinary tract. Liver weight and centrilobular hepatocellular hypertrophy were increased compared to controls in males at 1000 and 4000 ppm, and centrilobular hepatocellular hypertrophy was increased in females at 4000 ppm only. Gallbladder stones demonstrated to be cholesterol were observed in both males and females at all doses with no relationship to dose. Treatment-related neoplastic findings of the urinary tract were related to the presence of urinary tract stones and were only observed at 4000 ppm. These neoplasias were considered to be secondary to the administration of high doses of the test substance, as the urinary tract stones were shown to be composed of unmetabolized test substance, and were due to a non-genotoxic proliferative mechanism.

Based on the increased mortality at 4000 ppm in both males and females, the top dose of 4000 ppm was considered to have exceeded the Maximum Tolerated Dose. The NOAEL was established at 100 ppm (13.6 mg/kg bw/day in males, 16.7 mg/kg bw/day in females), as despite gallbladder concretions at all doses there was no increase at 100 ppm in epithelial hyperplasia in the gallbladder. The LOAEL was 1000 ppm. Treatment-related neoplastic findings of the urinary tract were observed at 4000 ppm but were considered to be secondary to the administration of high doses of the test substance which precipitated in the urinary tract, causing continuous local irritation and to be due to a non-genotoxic proliferative mechanism. They were considered to have no relevance to humans. Accordingly, up to and including the highest administered dose, there was no indication of neoplastic findings which are relevant to man. Accordingly, the NOAEL for carcinogenicity is considered to be 4000 ppm (560 mg/kg bw/day in males, 713 mg/kg bw/day in females).

 

Carcinogenicity study in the Wistar rat by dietary administration

In this GLP-conform study conducted according to OECD 453, 5-hydroxy-1,3-dimethylpyrazol-4-yl)(α,α,α-trifluoro-2-mesyl-p-tolyl)methanone was incorporated into rodent diet and administered at concentrations of 0, 25, 250, 1000, and 2500 ppm to groups of 75 male and female Wistar rats (M-267037-01-2). In addition to the carcinogenicity phase, the study also included a 6-month and 1-year repeated-dose toxicity phase. Only the results of the carcinogenicity phase are summarized here. The repeated-dose results are summarized in the respective endpoint summary.

The administered concentrations provided final doses of 0, 1.0, 10, 41, and 105 mg/kg bw/day for males and 0, 1.4, 14, 57, and 141 mg/kg bw/day for females. Body weight and food consumption were measured weekly for the first 13 weeks of the study, then every 4 weeks through the remainder of the study until necropsy. Ophthalmological examinations were conducted on all animals during acclimatization and at months 3, 6, 12, 18, and 24 of treatment. Blood for hematological and clinical chemical examination was collected after 6, 12, and 24 months from overnight-fasted rats. Urine was collected overnight at 3, 6, 12, 18, and 24 months from animals fasted overnight of food and water.

A treatment-related increase in mortality was observed in males at 2500 ppm. There was no treatment-related effect on mortality in other male treatment groups or in females at any dose level. Treatment-related clinical signs of toxicity included white area on the eye and soiled fur in one or more areas. An increased incidence of these findings was observed at 250, 1000, and 2500 ppm in both males and females. Body weight and body weight gain were reduced in males and females at the higher doses, with a greater effect observed in males than in females. There was no effect on food consumption at any dose in either males or females.

Treatment-related findings (corneal opacity, neovascularization of the cornea, oedema of the cornea, and "snow flake" corneal opacities) were seen during ophthalmoscopy in both males and females. At 6 and 12 months, increased incidence of these findings was seen only at 250, 1000, and 2500 ppm. At 24 months, these findings were increased compared to controls at all doses in males. However, at 25 ppm there was only a slight increase in corneal opacity, neovascularization of the cornea, and oedema of the cornea in males in neovascularization of the cornea in females. The findings are related to tyrosinemia resulting from inhibition of 4-hydroxyphenylpyruvate dioxygenase (HPPDase), a key enzyme in the tyrosine catabolic pathway. As humans are capable of metabolizing and excreting excess tyrosine while rats are much less capable of this metabolism, the corneal effects of HPPDase inhibitors are considered to be relevant for rats but not for man (for a detailed justification see endpoint summary for repeated dose toxicity).

There was no treatment-related effect on hematology at any dose or time point in either male or female rats. Plasma cholesterol was biologically and statistically significantly increased at 250 ppm and above at months 7 and 12, and was biologically and/or statistically significantly increased at 1000 and 2500 ppm at 18 and 24 months. This effect on plasma cholesterol was considered to be the only treatment-related clinical chemistry finding. Although plasma cholesterol was statistically significantly increased in males at 25 ppm at the 7-month time point, this value was within historical control data and cholesterol concentrations in this dose group returned to control levels by the 12- month time point; therefore, this increase was considered not to be treatment-related.

During urinalysis, higher ketone concentrations were noted in both males and females at 1000 ppm and 2500 ppm at all collection periods, and in males at 250 ppm at months 19 and 24 only. These were due to the triketone structure of the molecule, which is largely excreted unchanged in the urine. Urine pH was decreased at 250 ppm and above in males at all time points, while in females at 250 ppm and above urine pH was decreased only at the 3-month time point. Urinary protein was increased in males at all doses from month 7 onwards.

Liver and kidney weight was statistically significantly increased in males only at the higher doses in this study. There were no biologically significant effects on organ weights in females at any group. At necropsy, treatment-related findings were observed in the eyes, liver, and kidney. Eye opacities seen in both males and females were related to increased tyrosinemia and are considered not to be relevant for human risk assessment. An increased incidence of pale kidneys or irregular surface of the kidney was noted in males at 1000 and 2500 ppm.

 

Non-neoplastic microscopic findings: Treatment-related findings were observed in the eyes, liver, pancreas, thyroid gland, and kidneys. Findings observed in other organs were considered to be of little to no toxicological relevance or not to be treatment-related.

In the eyes, the incidence of corneal inflammation was increased in males at 250 ppm and above, and in females at 1000 and 2500 ppm. At 24 months, there was a very slight increase in corneal inflammation in males at 25 ppm and in females at 250 ppm. Regenerative hyperplasia of the cornea was increased in males at 250 ppm and above, and in females at 1000 and 2500 ppm, only. Neovascularization of the cornea was increased in males 250 ppm and above, and in females at 250 ppm and above. There was an increase in males in the incidence of mucous metaplasia of the cornea at 250 ppm and above. Corneal atrophy was increased in males at 250 ppm and above and in females at 1000 and 2500 ppm, and peripheral retinal atrophy was increased in both males and females at 250 ppm and above.

Centrilobular hepatocellular hypertrophy was increased in males from 250 ppm. In females centrilobular hepatocellular hypertrophy was only reported in one animal at 2500 ppm. In the pancreas, diffuse acinar degeneration / atrophy was reported in both males and females; the incidence of diffuse acinar degeneration / atrophy was increased in males and females at 1000 and 2500 ppm. The incidence of focal acinar degeneration / atrophy was increased in females at 2500 ppm.

In the thyroid, the incidence of altered colloid was increased in males and females from 250 ppm and above. Pigment deposition in the follicular cells was increased in both males and females in all dietary groups. The incidence of focal follicular cell hyperplasia was slightly increased in males from 250 ppm. In females, this finding was only at a very low incidence. No clear relationship of follicular cell hyperplasia to dose was evident. Diffuse follicular cell hypertrophy was noted in a few males and females from 250 ppm, again in the absence of a clear dose-relationship. The observed thyroid findings are considered a rat-specific phenomenon of HPPDase inhibitors without relevance for humans (for a detailed justification see endpoint summary for repeated dose toxicity).

There was no substantive dose-related increase in the incidence of chronic progressive nephropathy in males at 24 months, and the incidence in control animals at 24 months of 80% indicates that this is a very common finding in the aging male rat. Chronic progressive nephropathy was observed in females, with no dose relationship. Hyperplasia of the collecting ducts was increased in males at 1000 and 2500 ppm.

 

The only treatment-related neoplastic finding was that of squamous cell tumors (one papilloma, one carcinoma) observed in two males in the 2500-ppm dose group. These tumors were considered to have resulted from the corneal inflammation and regenerative hyperplasia resulting from tyrosinemia, and are not relevant to human risk assessment. Other tumors which were observed showed no relationship to dose or were considered to be consistent with those tumors found in ageing rats, and were not evaluated as related to treatment.

 

Conclusions

Based on increased mortality at 2500 ppm in males, this dose is considered to have exceeded the Maximum Tolerated Dose. Treatment-related findings were seen in the eyes, liver, kidney, thyroid, and pancreas. Findings in the eye were related to the biochemical mechanism of the test substance as an HPPDase inhibitor leading to increased tyrosinemia in rats, and were considered not to be relevant for humans. The observations in the liver, including centrilobular hepatocellular hypertrophy, were evaluated as being adaptive responses to treatment rather than adverse effects. The finding of chronic progressive nephropathy is considered to be a rodent-specific finding not relevant to man. Similarly, the findings of colloid alteration and pigment deposition were considered to be specific to the rat and not indicative of an adverse finding, but to be normal findings in ageing rats. The increased incidence of acinar degeneration / atrophy in the pancreas was only observed at 1000 and 2500 ppm, and may indicate a tyrosine-linked effect as well, as these findings were not observed in the mouse or dog, which are not as sensitive as the rat to the effects of tyrosinemia. The only treatment-related neoplastic findings, squamous cell tumors of the cornea, were evaluated as being due to a nongenotoxic proliferative mechanism and to be secondary to rat-specific increased plasma tyrosine concentrations not relevant to man.

The NOAEL in this study was therefore considered to be 25 ppm (1.0 mg/kg bw/day in males and 1.4 mg/kg bw/day in females), while the LOAEL was 250 ppm (10 mg/kg bw/day in males and 14 mg/kg bw/day in females). Treatment-related neoplastic findings of the cornea were observed at 2500 ppm but were considered to be secondary to continuous corneal inflammation and ulceration. These cornea effects are a rat specific phenomeon and to be due to a non-genotoxic proliferative mechanism. They were considered to have no relevance to humans. Accordingly, up to and including the highest administered dose, there was no indication of neoplastic findings which are relevant to man. Accordingly, the NOAEL for carcinogenicity is considered to be 2500 ppm (104 mg/kg bw/day in males, 140 mg/kg bw/day in females).