3β-hydroxyandrost-5-en-17-one acetate

Carcinogenicity

Description of key information

One reliable carcinogenicity study with the test substance is published (Rao et al., 1992). In the study, the test item was administered via diet to male F-344 rats at a concentration of 0.45% for up to 84 weeks. Peroxisome proliferation might be involved in the hepatocarcinogenicity of the test item. The carcinogenic potential of peroxisome proliferation to humans is still uncertain, due to known marked differences between rats (highly sensitive) and humans (insensitive). This study by itself is not sufficient for human cancer risk assessment. Therefore, the substance is considered not to be classified as carcinogenic according to CLP Regulation.

Key value for chemical safety assessment

Carcinogenicity: via oral route

Link to relevant study records

Reference

Endpoint:

carcinogenicity: oral

Type of information:

experimental study

Adequacy of study:

supporting study

Study period:

The dates of the study were not reported in the publication.

Reliability:

2 (reliable with restrictions)

Rationale for reliability incl. deficiencies:

study well documented, meets generally accepted scientific principles, acceptable for assessment

Remarks:

Study not performed according to GLP, but is sufficient to accept the data and is nevertheless well documented and scientifically acceptable.

Qualifier:

no guideline followed

Principles of method if other than guideline:

Sixteen 15-week-old male F-344 rats were administered Dehydroepiandrosterone acetate in the diet at a concentration of 0.45% for up to 84 weeks to examine the long term hepatic effects of Dehydroepiandrosterone (DHEA). Ten rats were maintained on a control diet without DHEA. All rats had free access to food and water. Three DHEA-treated rats were sacrificed between 70 and 75 weeks, and the remaining 13 rats were sacrificed at 84 weeks. Two control rats were sacrificed at 70 weeks and the remaining controls were sacrificed at 84 weeks. A complete necropsy was performed on all rats. Livers were excised and examined for gross lesions after serial sectioning. Selected portions of liver were fixed in 10% neutral buffered formalin and processed for routine histological examination. In addition, some sections of the liver were fixed in ethanol-acetic acid and processed for histochemical localization of gamma-glutamyl transferase (GGT). Sections from lungs, pancreas, and kidneys were also processed for light microscopy. Paraffin sections (4-5 um thick) were stained with hematoxylin and eosin, and those containing neoplastic lesions were stained for glutathione-S-transferase placental form (GST-P) by an immunoperoxidase method.

GLP compliance:

not specified

Specific details on test material used for the study:

Dehydroepiandrosterone acetate (Sigma Chemical Co., St. Louis, Missouri, USA).
Note that there remains unclearity on the test substance identification. In the materials and methods section of the paper, dehydropepiandrosterone acetate is mentioned as test substance, but throughout the paper DHEA is mentioned (dehydroepiandrosterone). Both substances are deemed sufficiently similar to consider the data in this dossier.

Species:

rat

Strain:

Fischer 344

Sex:

male

Details on test animals or test system and environmental conditions:

TEST ANIMALS
- Source: Male F-344 rats were purchased from Charles River Breeding Laboratories (Wilmington, Massachusetts, USA)
- Age when purchased: 15 weeks
- Weight when received by the laboratory: 250-275 g
- Housing: Rats were housed in groups of 3 to 4 in polycarbonate cages containing San-i-Cel bedding
- Diet: Ad libitum; AIN-76 semipurified diet (corn oil stripped of vitamin E; US Biochemical Corp., Cleveland, Ohio, USA)
- Water: Ad libitum

ENVIRONMENTAL CONDITIONS
- Temperature: Rats were housed in a temperature-controlled room, although temperature range was not specified in the publication.
- Humidity (%): Rats were housed in a humidity-controlled room, although humidity range was not specified in the publication.
- Photoperiod (hrs dark / hrs light): 12-hr dark-light cycle

Route of administration:

oral: feed

Vehicle:

not specified

Details on exposure:

- Treatment Group: 16 male rats were administered Dehydroepiandrosterone (DHEA) acetate in the diet at a concentration of 0.45% w/w.
- Control Group: 10 rats were maintained on a control diet without DHEA acetate.
- Diet: DHEA acetate was administered in AIN-76 semipurified diet (corn oil stripped of vitamin E; US Biochemical Corp., Cleveland, Ohio, USA). All rats had free access to food.

Analytical verification of doses or concentrations:

not specified

Duration of treatment / exposure:

- Treatment Group: 3 DHEA acetate-treated rats were treated up to 70 and 75 weeks, and the remaining 13 rats were treated 84 weeks.
- Control Group: 2 control rats received the control diet up to 70 weeks and the remaining 8 controls received the control diet up to 84 weeks.

Frequency of treatment:

The rats had free access to the 0.45% DHEA acetate-containing food (treatment group) or the control diet (control group).

Post exposure period:

No post-exposure period was described in the publication.

Dose / conc.:

0.45 other: % - concentration of DHEA acetate in the diet

Remarks:

DHEA acetate treatment group

Dose / conc.:

0 other: Control diet (AIN-76 semipurified diet [corn oil stripped of vitamin E])

Remarks:

Control group

No. of animals per sex per dose:

- 16 male rats were in the DHEA acetate treatment group that received 0.45% DHEA acetate in the diet
- 10 male rats were in the control group that received the control diet

Control animals:

yes, concurrent no treatment

Positive control:

No positive control was reported for this study.

Observations and examinations performed and frequency:

The "Materials and Methods" section of the publication did not provide methods information on any of the following observations / examinations that are routinely performed in OECD test guideline carcinogenicity studies:
- Cage side observations
- Detailed clinical observations
- Food consumption and compound intake
- Hematology
- Clinical chemistry


BODY WEIGHT: yes
- No details were provided on the time schedule for body weights.

Sacrifice and pathology:

GROSS PATHOLOGY
- A complete necropsy was performed on all rats.
- Livers were excised and examined for gross lesions after serial sectioning.
- Sections from lungs, pancreas, and kidneys were also processed for light microscopy.

HISTOPATHOLOGY
- Livers were examined for gross lesions after serial sectioning.
- Selected portions of liver were fixed in 10% neutral buffered formalin and processed for routine histological examination.
- Some sections of the liver were fixed in ethanol-acetic acid and processed for histochemical localization of gamma-glutamyl transferase (GGT).
- Sections from lungs, pancreas, and kidneys were processed for light microscopy. Paraffin sections (4-5 um thick) were stained with hematoxylin and eosin, and those containing neoplastic lesions were stained for glutathione-S-transferase placental form (GST-P) by an immunoperoxidase method.

ORGAN WEIGHTS
- Liver weights were performed.

Statistics:

The Student's t-test was used to evaluate the statistical significance of body weight changes, liver weight/100 g body weight and tumor incidence of the treatment group relative to the control group.

Clinical signs:

not specified

Dermal irritation (if dermal study):

not examined

Mortality:

not specified

Body weight and weight changes:

effects observed, treatment-related

Description (incidence and severity):

DHEA acetate treatment caused a marked reduction in body weights when compared to controls. Mean body weight of treated animals was 285.6 +/- 6 grams at study end, while the mean body weight of control animals was 428 +/- 10 grams at study end.

Food consumption and compound intake (if feeding study):

not specified

Food efficiency:

not specified

Water consumption and compound intake (if drinking water study):

not examined

Ophthalmological findings:

not examined

Haematological findings:

not examined

Clinical biochemistry findings:

not examined

Urinalysis findings:

not examined

Behaviour (functional findings):

not examined

Immunological findings:

not examined

Organ weight findings including organ / body weight ratios:

effects observed, treatment-related

Description (incidence and severity):

The liver weight: body weight ratios were significantly different between control and DHEA acetate-fed rats (3.4 g/100 g body weight versus 6.5 g/100 g body weight), although the differences in absolute liver weights were not as marked between the two groups. The observed increases in absolute liver weight in animals fed DHEA acetate are attributable, in part, to the presence of tumors.

Gross pathological findings:

effects observed, treatment-related

Description (incidence and severity):

Grossly, livers of 15 out of 16 rats treated with DHEA acetate for 70 to 84 weeks showed one or several gray-white lesions, some as large as 25 mm in diameter. The liver of one DHEA acetate fed rat that did not contain any gross tumors was infiltrated with leukemic cells. Livers from control animals did not show any grossly visible tumors.

Neuropathological findings:

not examined

Histopathological findings: non-neoplastic:

effects observed, treatment-related

Description (incidence and severity):

Livers of 15 out of 16 rats treated with DHEA acetate for 70 to 84 weeks contained altered areas. Cells in altered areas were larger than adjacent parenchymal cells and exhibited eosinophilic or basophilic cytoplasm with an increased nuclear cytoplasmic ratio and prominent nucleoli.

Description (incidence and severity):

- Livers of 15 out of 16 rats treated with DHEA acetate for 70 to 84 weeks showed hepatocellular carcinoma (HCC). HCC were well differentiated with trabecular, solid sheets or pseudoglandular patterns. Tumor cells displayed cytological and nuclear features of low grade hepatocellular carcinomas. Extension of tumors into vascular spaces or metastasis to lungs was not observed.

- Livers of 15 out of 16 rats treated with DHEA acetate for 70 to 84 weeks contained neoplastic nodules (NN). Cells in NN usually displayed morphological features similar to those of cells in altered areas. However, in NN the mitotic activity was very prominent, with cells arranged in 2-3-cell-thick plates and causing compression of adjacent parenchyma.

Details on results:

All types of liver lesions, i.e., altered areas, NN and HCC, that were examined for gamma-glutamyl transferase (GGT) and glutathione-S-transferase placental form (GST-P) lacked these 2 marker enzymes.

The results of this study demonstrate that DHEA acetate is a hepatocarcinogen in male F-344 rats. 94% of rats developed neoplastic lesions, including hepatocellular carcinoma in 88%. No liver tumors were observed in control rats that were fed the same diet without DHEA acetate. The morphological and phenotypic properties of DHEA acetate-induced liver lesions are similar to those induced by classical chemical peroxisome proliferators. Lack of GGT and GST-P is unique to peroxisome proliferator-induced liver tumors in contrast to genotoxic carcinogen-induced liver tumors in which these markers are consistently expressed.

Relevance of carcinogenic effects / potential:

In this study with male F-344 rats, DHEA acetate acts as a complete hepatocarcinogen. The authors provide evidence that peroxisome proliferation is involved in the hepatocarcinogenicity of DHEA acetate in male F-344 rats. However, the relevance of this study's results to humans is not clear based on the information provided in the publication.

Dose descriptor:

conc. level:

Effect level:

ca. 0.45 other: %

Based on:

test mat.

Sex:

male

Basis for effect level:

histopathology: neoplastic

histopathology: non-neoplastic

Critical effects observed:

yes

Lowest effective dose / conc.:

0.45 other: %

System:

hepatobiliary

Organ:

liver

Treatment related:

yes

Dose response relationship:

not specified

Relevant for humans:

not specified

A supporting study evaluated the effect of DHEA acetate on liver cell proliferation. A single dose of [3H]thymidine (1 uCi/g body weight) was injected 1 hour prior to sacrifice to control rats and rats fed DHEA acetate for 4, 7, or 14 days. Another group of rats fed DHEA for 24 weeks and age-matched controls were also used for [3H]thymidine labeling. In these studies, a decrease in thymidine uptake and the number of labeled nuclei was evident in DHEA acetate-treated rats. These results demonstrate that liver cell proliferation in vivo is inhibited by DHEA acetate. Thus, the peroxisome proliferating effects of DHEA acetate were hypothesized to occur through mechanisms other than liver cell proliferation.

Conclusions:

Dehydropeiandrosterone was administered in the diet at a concentration of 0.45% to F-344 rats for up to 84 weeks. At the termination of the experiment, 14 of 16 rats developed hepatocellular carcinomas. Liver tumors induced by dehydroepiandrosterone lacked gamma-glutamyl transpeptidase and glutathione S-transferase (placental form); these phenotypic properties are identical to the features exhibited by liver tumors induced by other peroxisome proliferators. Dehydroepiandrosterone was also shown to markedly inhibit liver cell [3H]thymidine labelling indices, suggesting that cell proliferation is not a critical feature in liver tumor development with this agent. These results show that although dehydroepiandrosterone exerts anticarcinogenic effecs in a variety of tissues, the peroxisome-proliferative property makes it a hepatocarcinogen.

Executive summary:

In this study with male F-344 rats, DHEA acetate acts as a complete hepatocarcinogen.  The authors provide evidence that peroxisome proliferation is involved in the hepatocarcinogenicity of DHEA acetate in male F-344 rats.   Although there are known marked differences between rats (highly sensitive) and humans (insensitive) to peroxisome proliferator substances, experts are still uncertain about the carcinogenic potential of peroxisome proliferators to  humans.  Overall, the relevance of this study's results to humans is not clear based on the information provided in the publication.  This study, by itself, is not sufficient for human cancer risk assessment purposes.  

Endpoint conclusion

Endpoint conclusion:

adverse effect observed

Study duration:

chronic

Species:

rat

Quality of whole database:

Single study in rats available, well described but with limited information on the test item used.

System:

hepatobiliary

Organ:

liver

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

Based on the results described above, the limited dataset and the uncertainty on human carcinogenic potential, the substance is considered not to be classified as carcinogenic according to CLP Regulation.

Additional information

One reliable study was published, describing experimental data in male rats.

Dehydroepiandrosterone was administered in the diet at a concentration of 0.45% to F-344 rats for up to 84 weeks (Rao et al., 1992). At the termination of the experiment, 14 of 16 rats developed hepatocellular carcinomas. Liver tumours induced by dehydroepiandrosterone lacked gamma-glutamyl transpeptidase and glutathione S-transferase (placental form); these phenotypic properties are identical to the features exhibited by liver tumors induced by peroxisome proliferators. Dehydroepiandrosterone was also shown to markedly inhibit liver cell [3H]thymidine labelling indices, suggesting that cell proliferation is not a critical feature in liver tumor development with this agent. These results show that although dehydroepiandrosterone exerts anticarcinogenic effects in a variety of tissues, the peroxisome-proliferative property makes it a hepatocercinogen in this particular study.

As there are known, marked differences between rats (highly sensitive) and humans (insensitive) to peroxisome proliferator substances, experts are still uncertain about the carcinogenic potential of peroxisome proliferators to humans. Overall , the relevance of this study's results to humans is not clear. This study, by itself, is not sufficient for human cancer risk assessment purposes.

It also should be noted that there remains unclarity on the test substance identification. In the materials and methods section of the paper, dehydropepiandrosterone acetate is mentioned as test substance, but throughout the paper DHEA is mentioned (dehydroepiandrosterone). Both substances are deemed sufficiently similar to consider the data in the current registration dossier.