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Toxicological information

Toxicity to reproduction: other studies

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Administrative data

Endpoint:
toxicity to reproduction: other studies
Type of information:
experimental study
Adequacy of study:
supporting study
Reliability:
1 (reliable without restriction)
Rationale for reliability incl. deficiencies:
guideline study

Data source

Reference
Reference Type:
study report
Title:
Unnamed
Year:
2003
Report Date:
2003

Materials and methods

Test guideline
Qualifier:
according to
Guideline:
other: OECD Protocol and Guidance for the Conduct of the Rodent Hershberger Assay; Phase 2 of the Validation of the Rodent Hershberger Assay
Deviations:
no
GLP compliance:
yes (incl. certificate)
Remarks:
BASF Aktiengesellschaft, Experimentelle Toxikologie und Ökologie, D-67056 Ludwigshafen
Type of method:
in vivo

Test material

Reference
Name:
Unnamed
Type:
Constituent
Test material form:
solid: particulate/powder
Remarks:
migrated information: powder

Test animals

Species:
rat
Strain:
Wistar
Remarks:
CrlGlxBrlHan:WI
Sex:
male
Details on test animals and environmental conditions:
TEST ANIMALS
- Source: supplied by Charles River, Sulzfeld, Germany
- Age at start of administration period: 56 +/- 1 days
- Weight at study initiation: The weight variation of the animals used did not exceed 20 % of the mean weight (mean weights: 199.4 g - 206.2 g).
- Housing: The rats were housed singly in type DK III strainless wire mesh cages supplied by Becker & CO., Castrop-Rauxel, Germany (floor area about 800 cm²). Underneath the cages, disposable waste trays were used. The animal room was completely disinfected using a disinfector ("AUTEX", fully automatic, formalin-ammonia-based terminal disinfector). The floor and the walls were cleaned once a week. The cleansing liquid used was water containing about 0.1 % Incidin (supplied by Henkel, Düsseldorf, Germany).
- Diet: The food used was ground Kliba maintenance diet rat-mouse, meal, supplied by Provimi Kliba SA, Kaiseraugst, Switzerland. Food was available ad libitum.
- Water: drinking water was available ad libitum.
- Reason for species selection: Rats were selected since this rodent species is recommended in the respective test guideline.

ENVIRONMENTAL CONDITIONS
- Temperature: 20-24 °C
- Humidity: 30-70 %
- Photoperiod: 12 hours (12 hours light from 06.00 a.m. - 06.00 p.m., 12 hours dark from 06.00 p.m. - 06.00 a.m.).

CASTRATION
The animals were anesthetized with Isoflo® (Essex GmbH, Munich, Germany). The scrotum as well as mesorchium was opened by a median incision and the testes were exteriorized. Blood vessels and seminal ducts were ligated by means of a Dexon thread. The testes and epididymides were removed. Finally, after confirming that no further bleeding occurred, the scrotum was closed with autoclips.
The castrated rats were treated/substituted by testosterone propionate at 0.4 mg/kg bw day (see PREPARATION OF DOSING SOLUTIONS).

Administration / exposure

Route of administration:
oral: gavage
Vehicle:
corn oil
Details on exposure:
PREPARATION OF DOSING SOLUTIONS:
The test substance was administered as a suspension in corn oil. To prepare the suspension, the appropriate amount of test substance was weighed, depending on the dose group, subsequently suspended in corn oil using a high speed homogenizer, filled up to the desired volume and mixed using a magnetic stirrer. During the administration the test substance preparations were kept homogenous with a magnetic stirrer. The test substance preparations were prepared at the beginning of the administration period and filled up in aliquots and kept cold in a refrigerator.
Testosterone propionate was administered as a solution in corn oil. To prepare the solution, the appropriate amount of testosterone propionate was weighed and filled up to the desired volume and mixed using a magnetic stirrer. Testosterone propionate preparations were prepared at the beginning of the administration period and filled up in aliquots and kept cold in a refrigerator.
Analytical verification of doses or concentrations:
yes
Details on analytical verification of doses or concentrations:
The analytical investigations of the test substance preparations were carried out at the Analytical Chemistry Laboratory of the Experimental Toxicology and Ecology of BASF Aktiengesellschaft, Ludwigshafen, Germany. The stability test of the test substance in corn oil was carried out from taking samples of the lowest concentration at the day of sample preparation and at the last day of using the test substance preparation. The stability of the test substance in corn oil over the application period was demonstrated.




Homogeneity analyses and concentration control analyses of the test substance preparations were carried out in samples of the highest and lowest concentration drawn from the bottom, middle and the top of the suspension at the start of the administration period. Additional concentration control analyses were performed with samples from the lowest concentration drawn from the middle of the suspension at the end of the administration period. The analyses demonstrated the correctness of the concentrations and the homogeneity of the test substance. The recovery rates were within an acceptable range (91.3 - 95.7 % of the target concentration).
Duration of treatment / exposure:
10 days
Frequency of treatment:
daily
Duration of test:
10 days
Doses / concentrationsopen allclose all
Dose / conc.:
1 000 mg/kg bw/day (nominal)
Dose / conc.:
300 mg/kg bw/day (nominal)
No. of animals per sex per dose:
6
Control animals:
yes
Details on study design:
The test substance was administered to groups of 6 castrated but testosterone propionate treated/substituted male Wistar rats (which were subdivided in three subsets) by gavage for 10 days at dose levels of 300 and 1000 mg/kg body weight/day. A vehicle control group was treated with corn oil by gavage. Another control group was administered subcutaneously 0.4 mg/kg bw/day testosterone propionate to test the androgenic efficacy. The used administration volume was 5 mL/kg bw/day by gavage and 0.5 mL/kg bw subcutaneously. All surviving animas were sacrified after a fasting period (wihdrawal of food) for at least 16-20 hours.
Statistics:
Mean and standard deviation of each test group were calculated for each parameter. The statistical analysis of the parameter hormones was performed using the Wilcoxon-test (two-sided) for the equal medians. For the parameters food consumption, body weight, body weight change and food efficiency, a comparison of each group with group 1 was performed using DUNNETT's test (two-sided) for the hypothesis of equal means.

Results and discussion

Any other information on results incl. tables

CLINICAL EXAMINATIONS

Mortality: 

No animals died prematurely. Clinical examinations: One animal (No. 34) showed piloerection, slight reduced general condition and respiration sounds on day 3, 4, 5. Most probably these findings were caused by an aspiration of the test-compound preparation during gavage at the beginning of the study. Therefore, these single occurrences were clearly no adverse systemic substance-related findings. Moreover, these clinical findings were only observed until day 5, towards the end of the study no symptoms were observed, anymore. Food consumption: Food consumption was reduced statistically significantly on day 5 (-33 .4%) in high dose animals. Animal No. 34 caused this finding, because only this rat showed significantly reduced food consumption on day 5 as shown by single data. Therefore, the impaired food consumption was assessed as not substance-related but was clearly related to the findings observed during clinical examinations in animal No. 34, only.

 

Water consumption: 

There were no overt deviations in volume between treated groups and controls. Body weight data: Body weight was reduced statistically significantly in high dose animals on day 2 (-8 .3%), whereas body weight change was statistically significantly impaired throughout the whole study, up to -359.1 % on day 1 of the study. These findings were considered to be not specific substance-related but were also caused by animal No. 34, only - see also clinical examinations as well as food consumption.

 

Food efficiency: 

No substance-related findings were obtained.

 

CLINICAL PATHOLOGY 

Hormones: No treatment-related effects on mean testosterone (4.21 vs. 3.72 mmol in TP ctrl.), dihydrotestosterone (326.9 vs 325.13 pmol/l in TP ctrl.) concentrations were found in the serum of castrated males treated with 1000 mg/kg bw/day of the test substance + 0.4 mg/kg bw/day testosterone propionate for 10 days when compared with the hormone levels of animals given testosterone propionate, only. The 300 mg/kg bw/d dose group was not investigated. The significant decrease in luteinizing hormone (0.62 vs. 2.41 µg/l in TP ctrl.) in the serum of rats, that received the test substance is considered to be an incidental finding.

In the serum of castrated male rats testosterone levels were significantly decreased (0.53 vs 3.72 mmol in TP ctrl.) and luteinizing hormone concentrations significantly increased (14.33 vs. 2.41 µg/l in TP ctrl.) compared to the values of castrated animals received 0.4 mg/kg bw/day of testosterone propionate as reference androgen. No significant difference in serum dihydrotestosterone concentrations was seen between both test groups (361.38 vs 325.13 pmol/l in TP ctrl.).

PATHOLOGY 

Weight parameters  

Absolute weights: 

When compared with the TP control group , the mean absolute weights of following organs were significantly increased or decreased:

 

Male animals

Group

1000 mg/kg bw/d

Terminal body weight

-7.1%

Prostate ventral fixed

+21.2%

All other mean absolute weight parameters did not show significant differences when compared with the TP control. The reduced terminal body weight and the increase of the prostate weight in 1000 mg/kg bw/d dose group are considered incidental.
Control group without testosterone (without TP):
When compared with TP control group, the mean absolute weights of following organs were significantly increased or decreased:

 

Male animals

Group

without TP

Kidneys

-15.3%

Seminal vesicle

-93.1%

Bulbo-urethral gland

-82.5%

Prostate ventral fresh

-87.8%

Prostate ventral fixed

89.3%

Glans penis

48.9%

Musc. Levator ani with Musc. bulbocavernosus

-62.9%

All other mean absolute weight parameters did not show significant differences when compared with the TP control group. The reduced kidney weight is considered incidental.
 
Relative Organ Weights:
When compared with TP control group, the mean relative weights of following organs were significantly increased or decreased:

 

Male animals

Group

1000 mg/kg bw/d

Bulbo-urethral gland

+36.4%

Prostate ventral fresh

+33.3%

Prostate ventral fixed

+31.4%

All other mean relative weight parameters did not show significant differences when compared with TP control group. The increased weights of bulbo-urethral gland and prostate are considered incidental.
Control group without testosterone (without TP):
When compared with TP control group, the mean relative weights of following organs were significantly increased or decreased:

 

Male animals

Group

without TP

Seminal vesicle

-92.7%

Bulbo-urethral gland

-81.8%

Prostate ventral fresh

-86.7%

Prostate ventral fixed

-88.2%

Glans penis

-44.8%

Adrenal glands

+21.4%

Musc. Levator ani with Musc. bulbocavernosus

-61.0%

All other mean relative weight parameters did not show significant differences when compared with TP control group. The increased weight of adrenal glands is considered incidental.
 
Gross lesions:
There were no gross lesions.
 
Histopathology:
In the treatment group (1000 mg/kg bw/d dose group) the histology of prostate, seminal vesicle and the bulbo-urethral gland was comparable to the TP dose group. There were no histopathological findings.

The accessory sex organs (ventral prostate, seminal vesicle and bulbo-urethral gland) were immature in males of control (without TP) dose group . The administration of testosterone propionate led to the maturation of the accessory sex organs.

Applicant's summary and conclusion