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Endpoint:
additional toxicological information
Type of information:
migrated information: read-across from supporting substance (structural analogue or surrogate)
Adequacy of study:
weight of evidence
Reliability:
other: Independent expert evaluation of study assigned reliability 1
Rationale for reliability incl. deficiencies:
other: see 'Remark'
Remarks:
Reliability ratings are assigned to specific studies or generated data. This report is an expert evaluation based on all available data and, as such, cannot be assigned a reliability rating. The evaluation was of thyroid tumors reported in an NTP carcinogenicity study conducted with tertiary butyl alcohol in B6C3F1 mice. The NTP study itself was rated as reliability 1. Tertiary butyl alcohol is the primary in vivo metabolite of tertiary butyl acetate.

Data source

Reference
Reference Type:
other: Unpublished expert evaluation
Title:
Unnamed
Year:
2001
Report Date:
2001

Materials and methods

Type of study / information:
Review of thyroid gland changes which occurred in B6C3F1 mice following long-term exposure to tertiary butyl alcohol. Includes an assessment of thyroid tumors, a discussion of tertiary butyl alcohol metabolism, and possible modes of action for observed effects.
Test guideline
Qualifier:
no guideline required
Guideline:
other: Submission is an expert evaluation of a study conducted according to NTP guidelines.
Deviations:
not applicable
GLP compliance:
no
Remarks:
Submission is an expert evaluation of a study that was GLP compliant. Expert evaluation does not require GLP compliance.

Test material

Reference
Name:
Unnamed
Type:
Constituent
Type:
Constituent

Results and discussion

Any other information on results incl. tables

NTP DATA USED BY McCLAIN IN EVALUATION:

13 WEEK STUDIES IN RATS AND MICE:

-Relative liver weights were significantly increased in male mice at 20 mg/mL and in both sexes at 40 mg/mL. Relative liver weights were significantly increased in male rats at 5, 10 and 20 mg/mL and for female rats, both the absolute and relative liver weights were increased at all dose levels. At the 15-month interim sacrifice in the rat carcinogenicity study, relative liver weights were significantly increased in high-dose male and female rats but there were no microscopic effects that would correlate with the increased liver weights in the subchronic mouse and rat studies. Because of high mortality, interim sacrifices were not conducted in the mouse bioassay. 

-There were no microscopic changes in the thyroids of rats or mice following a 13-week exposure. 

-Thyroid glands were not weighed in either species in the subchronic studies

  

TWO-YEAR STUDIES IN RATS AND MICE:

 - Incidence of thyroid follicular cell adenomas was marginally but not statistically significantly increased in male mice in the mid-dose group (10 mg/mL) and was significantly increased in female mice in the high-dose group (20 mg/mL). 

- No thyroid tumors were observed in either sex of rat.

- There was a statistically significant increase in follicular cell hyperplasia in all groups of treated male mice and in the 10 and 20 mg/mL group females.

- Follicular cell hyperplasia in mice consisted of foci with increased numbers of closely packed follicular epithelial cells, sometimes with minimal papillary folds.

- Hyperplastic lesions in mice treated with tertiary butyl alcohol were similar in appearance to spontaneous thyroid follicular hyperplasia observed in control mice.

- Follicular cell thyroid gland adenomas were more complex than the hyperplastic foci with more atypical features.

- No other morphological changes were observed in the thyroid glands of mice and no treatment-related lesions were observed in the thyroid glands of either sex of rat.

  

MECHANISMS FOR INDUCTION OF THYROID TUMORS - genetic versus epigenetic:

Citing Contrera et al.(1997), McClain states “Thyroid follicular cell neoplasia is one of the most common chemical-induced endocrine tumors observed in rat carcinogenicity studies”. A review of the literature suggests two basic mechanisms by which chemicals cause thyroid gland neoplasia in rodents. One involves chemicals that exert a direct genotoxic carcinogenic effect on the thyroid gland and the other involves an epigenetic mechanism in which chemicals, through a variety of mechanisms, alter thyroid gland function and produce thyroid gland neoplasia secondary to hormone imbalance (Hill et al., 1989; Capen, 1994; McClain, 1995; Hard, 1998).

  

References cited:

Contrera et al., 1997. Carcinogenicity testing and the evaluation of regulatory requirements for pharmaceuticals. Reg. Toxicol. Pharmacol., 25:130-145.

  

Hill et al., 1989. Review: Thyroid follicular cell carcinogenesis. Fund. Appl. Toxicol., 112: 629-697.

  

Capen CC, 1994. Mechanisms of chemical injury of thyroid gland. Prog. Clin. Biol. Res., 387: 173-191.

  

McClain RM, 1995. Mechanistic considerations for the relevance of animal data on thyroid neoplasia to human risk assessment. Mutation Res., 333: 131-142.

  

Hard G, 1998. Recent developments in the investigation of thyroid regulation and thyroid carcinogenesis. Env Health Perspect, 106: 427-436.

  

The total weight of evidence available indicates that tertiary butyl alcohol is not genotoxic. The mutagenic/genotoxic potential of tertiary butyl alcohol has been characterized in a series of well conduced bacterial mutagenicity tests, in vitro mammalian mutagenicity tests, and an in vivo somatic cell mutagenicity test. There was no evidence of mutagenicity or clastogenicity. Therefore, it is unlikely that tumor response in tertiary butyl alcohol-treated mice occurred via a genotoxic mode of action.

  

Studies using anti-thyroid drugs to induce thyroid tumors in rodents suggest a more likely mechanism for effects observed in the NTP bioassay involves hormone imbalance (Furth, 1959; Furth, 1969). When anti-thyroid compounds are first administered to rodents, the resulting hormone imbalance causes an increase in secretion of pituitary thyroid stimulating hormone (TSH) to stimulate thyroid function. As more TSH continues to be produced, it causes a variety of structural and functional changes in the thyroid, including follicular cell hypertrophy, hyperplasia and with long-term stimulation, ultimately neoplasia. A number of investigators have demonstrated that excessive stimulation by TSH alone, in the absence of any chemical treatment, can cause thyroid gland neoplasia in rodents (Axelrad and Leblond, 1955; Bielschowsky, 1953; Isler et al., 1958; Leblond et al., 1957; Furth, 1954).

  

References cited:

Furth J, 1959. A meeting of ways in cancer research: Thoughts on the evolution and nature of neoplasms. Cancer Res, 19: 241-58.

  

Furth J, 1969. Pituitary cybernetics and neoplasia Harvey lectures. New York/London: Academic Press: 47-71.

  

Axelrad AA and Leblond CP, 1955. Induction of thyroid tumors in rats by a low iodine diet. Cancer, 8: 339-67.

  

Bielschowsky F, 1953. Chronic iodine deficiency as cause of neoplasia in thyroid and pituitary of aged rats. Br J Cancer, 7: 203-213.

  

Isler et al., 1958. Influence of age and of iodine intake on the production of thyroid tumors in the rat. J Natl Cancer Inst, 21: 1065-1081.

  

Leblond et al., 1957. Induction of thyroid tumors by a low iodine diet. Can Cancer Conf, 2: 248-66.

  

Furth J, 1954. Morphologic changes associated with thyrotropin-secreting pituitary tumors. Am J Pathol, 30: 421-63.

   

  

SPECIES DIFFERENCES IN RESPONSE TO THYROID TOXICANTS:

Another factor that should be considered in extrapolating observed thyroid effects in rodents to potential effects in humans is species differences in thyroid gland physiology. Unlike primates, rodents and some other species lack thyroid binding globulin (TBG), which is the predominant plasma protein that binds and transports thyroid hormone in the blood (Dohler et al., 1979). Humans have three plasma proteins that bind and transport thyroid hormone, with TBG having a binding affinity of 3 to 5 orders of magnitude more than the other two. In rodents, the absence of TBG contributes to species differences in thyroid gland function including a much shorter half-life of thyroxine (T4) in the rat (12 hours vs. 5-9 days in humans) and a much higher circulating level of TSH (Dohler et al., 1979).

  

Reference cited:

Dohler et al., 1979. The rat as a model for the study of drug effects on thyroid function: Consideration of methodological problems. Pharmacol Ther, 5: 305-18.

  

McClain states “This indicates that the rat thyroid gland operates at a much higher functional level with respect to thyroid hormone turnover compared to the primate.” McClain also points out several structural differences between the rodent and primate thyroid gland. While the primate thyroid has follicles that are uniformly large with abundant colloid and are lined by relatively flattened follicular epithelial cells, the rodent thyroid has large follicles only at the periphery of the gland and most of the gland is made up of comparatively small follicles with small amounts of colloid surrounded by more cuboidal follicular epithelium. McClain concludes “both the physiologic parameters and the histologic appearance indicate that the rodent thyroid gland is markedly more active and operates at a considerably higher functional level with respect to thyroid hormone turnover as compared to the primate.”

  

MECHANISMS FOR ALTERED THYROID FUNCTION:

There are a variety of ways in which chemicals can interfere with thyroid gland function. Regardless of mechanism, the response to altered thyroid function in the rodent is similar. To compensate, the pituitary releases thyroid stimulating hormone (TSH) which in turn stimulates the thyroid gland to produce more hormone. Over time, this can lead to follicular cell hypertrophy, hyperplasia and with long-term stimulation, eventually neoplasia. McClain states “The effects of the chemical can involve either intra- or extrathyroidal mechanisms. Intrathyroidal mechanisms include interference with active iodine uptake, inhibition of the thyro-peroxidase (TPO) enzymes involved in hormone synthesis, or with hormone release. Extrathyroidal mechanisms include effects on the conversion of T4 to the active hormone, T3, by the 5’-monodeiodinase, and increased thyroid hormone metabolism and excretion, mainly through induction of UDP-glucuronyltransferase in the liver.” A variety of examples are provided in the evaluation to support these observations.

  

Rodents appear to be much more susceptible than humans to the development of thyroid gland neoplasia secondary to altered thyroid gland function. A number of relatively extensive epidemiologic studies have not shown a clear link between iodine deficiency in humans and the development of thyroid gland neoplasia (Doniach, 1970; Pendergrast et al., 1961; Saxen and Saxen, 1954) while rats on iodine deficient diets have a high incidence of thyroid gland neoplasia. The only known cause of thyroid cancer in humans is exposure to ionizing radiation. 

  

References cited:

Doniach I, 1970. Aetiological consideration of thyroid carcinoma. In: Smithers D, ed. Neoplastic diseases at various sites. Tumours of the thyroid gland. Edinburgh/London: E&S Livingstone.

  

Pendergrast et al., 1961. Thyroid cancer and thyrotoxicosis in the United States. Their relationship to endemic goiter. J Chronic Dis, 13: 2-38.

  

Saxen EA and Saxen LO, 1954. Mortality from thyroid diseases in an endemic goiter area. Studies in Finland. Docu Med Geograph Tropica, 6: 335-41.   

  

ENZYME INDUCTION:

A discussion of the enzyme inducing properties of tertiary butyl alcohol in rodents is also presented since one theory for increased thyroid tumor incidence in mice suggests that thyroid changes in male and female mice may be secondary to a compensatory increase in pituitary TSH due to microsomal enzyme induction which in turn causes an increase in metabolism and turnover of thyroid hormone. Induction of hepatic microsomal enzymes as a possible mode of action is also discussed by the NTP in the NTP study report. 

  

Tertiary butyl alcohol is metabolized by conjugation and oxidation. Conjugation with glucuronic acid results in the excretion of the glucuronide in the urine. Oxidation by microsomal enzymes involves reaction with hydroxyl radicals. The resulting major metabolites are 2-methyl-1,2-propanediol and 2-hydroxyisobutyrate.

  

Microsomal enzyme induction of tertiary butyl alcohol has been demonstrated in a number of studies in both rats and mice. When an aqueous solution of tertiary butyl alcohol was administered to Sprague-Dawley rats by the oral or intraperitoneal routes, microsomal enzyme activity was increased threefold (Bechtl and Cornish, 1972). The rate of elimination of tertiary butyl alcohol from the bloodstream was increased in mice previously exposed to tertiary butyl alcohol. The increased elimination was considered to reflect an induction of the smooth endoplasmic reticulum (McComb and Goldstein, 1979). When Sprague-Dawley rats were exposed to tertiary butyl alcohol for three (2000 ppm) or five (500 ppm) days by the inhalation route, the three day exposure induced hepatic cytochrome P450 and increased the metabolism of n-hexane (Aarstad et al., 1985). 

  

References cited:

Bechtel BH and Cornish HH, 1972. Effect of the butyl alcohols on liver microsomal enzymes. Toxicol. Appl. Pharmacol., 22: 298-299.

  

McComb JA and Goldstein DB, 1979. Quantitative comparison of physical dependence on tertiary butanol and ethanol in mice: correlation with lipid solubility. J. Pharmacol Exp. Ther., 208: 113-117.

  

Aarstad et al., 1985. Inhalation of butanols: changes in the cytochrome P450 enzyme system. Archives Toxicology, supplement 8: 418-421. 

  

Applicant's summary and conclusion

Conclusions:
The author concluded:
“The findings of thyroid follicular cell hyperplasia in male and female B6C3F1 mice and an increase in the incidence of follicular cell adenoma in female mice in a 2-year carcinogenicity study are compatible with a proliferative response secondary to hormone imbalance. The most likely hypothesis is altered thyroid hormone disposition as a result of microsomal enzyme induction in TBA treated mice. There is no evidence for a mutagenic or clastogenic effect of TBA, thus a genotoxic mode of action is unlikely. In the literature, there is substantial evidence that TBA is a microsomal enzyme inducer in both rats and mice, which is consistent with liver weight increases observed in rats and mice in the 13 week and 2 year studies.

Thyroid hormone imbalance is a commonly observed mode of action for the production of thyroid gland tumors in rodent carcinogenicity studies. There is a general scientific consensus that there is a marked species difference between rodents and humans in the susceptibility for the development of thyroid gland neoplasia secondary to altered thyroid gland function, with humans being far less susceptible than rats and mice (Hill et al., 1989; Capen, 1994; McClain, 1995; Hard, 1998; Hill et al., 1998).”

Reference cited:
Hill et al., 1998. Risk assessment of thyroid follicular cell tumors. Env Health Perspect, 106:447-457.

In vivo, tertiary butyl acetate is rapidly metabolized to tertiary butyl alcohol. Subchronic inhalation studies conducted in rats and mice with tertiary butyl acetate showed that exposure to high vapor concentrations caused increased liver weights in the absence of significant histopathologic effects. Similar liver effects were reported in repeat-exposure studies conducted with tertiary butyl alcohol. Inclusion of this expert opinion on tertiary butyl alcohol by McClain (2001) is relevant for the over-all evaluation of repeat-exposure toxicity/carcinogenicity of tertiary butyl acetate.
Executive summary:

In an expert evaluation conducted by R. Michael McClain, the author examines the thyroid follicular cell tumors observed in mice exposed ad libitum to tertiary butyl alcohol in the drinking water in an NTP carcinogenicity study and postulates species differences and possible modes of action for the observed effects at high doses. 

Based on a weight-of-the-evidence evaluation, tertiary butyl alcohol is unlikely to pose a significant risk for the development of thyroid tumors in humans exposed to low levels of this chemical and there is no evidence that the effect which occurred in mice at extremely high doses is relevant for human carcinogenicity classification.