Registration Dossier

Administrative data

Endpoint:
endocrine system modulation
Type of information:
experimental study
Adequacy of study:
other information
Reliability:
2 (reliable with restrictions)
Rationale for reliability incl. deficiencies:
other: Well documented study which meets generally accepted scientific standards

Data source

Reference
Reference Type:
publication
Title:
Screening of some anti-androgenic endocrine disruptors using a recombinant cell-based in vitro bioassay
Author:
Roy P, Salminen H, Koskimies P, Simola J, Smeds A, Saukko P & Huhtaniemi IT
Year:
2004
Bibliographic source:
PMID: 15084347, J Steroid Biochem Mol Biol. 88(2):157-66

Materials and methods

Test guideline
Qualifier:
no guideline followed
Principles of method if other than guideline:
Cell-based androgen reporter assay using the Chinese hamster ovarian cell line (CHO K1) in the 96 -well format. The purpose of this assay is a high throughput screening for endocrine disrupting (ED) substances in environmental and biological samples.
CHO cells were cotransfected with plasmids encoding mouse mammary tumour virus-neomycin-luciferase and human androgen receptor (hAR). A stable cell line was established. After selection with neomycin, a highly active clone was obtained which stably expressed both the hAR and the androgen-responsive luciferase reporter.
GLP compliance:
no
Type of method:
in vitro

Test material

Reference
Name:
Unnamed
Type:
Constituent
Details on test material:
- Name of test material (as cited in study report): 4,4'-Dichlorodiphenyl sulphone

Administration / exposure

Analytical verification of doses or concentrations:
no
Details on analytical verification of doses or concentrations:
The concentrations of the stock solutions of all the test compounds were 1 mmol/L in ethanol. They were then further diluted in the medium, resulting in the final concentration of ethanol in the incubations of 0.01%. Thus the final concentration was 0.01/100 mmol/L = 0.0001 mmol/L = 0.1 µmol/L.
Doses / concentrations
Remarks:
Doses / Concentrations:
0.1 µmol/L = 0.287 ng/L for anti-androgenic effects up to 50 µmol/L
Basis:
nominal conc.

Examinations

Examinations:
stimulation of luciferase activity
Positive control:
R1881 (Metribolone CAS 965-93-5)

Results and discussion

Details on results:
Test result:
- DCDPS incubation to the in vitro asssay resulted in a negative response. Therefore no androgenic or anti-androgenic effects were identified.

Validity of the assay:
- the test system was idendified as specific and functioning, since stimulation of the cells with androgens for 24h resulted in about 15-fold stimulation of luciferase activity. Furthermore potent steroidal and non-steroidal anti-androgens significantly inhibited the androgen-induced transactivation.
- non-androgenic steroids showed weak activity at high concentrations.
- RT-PCR and western blot confirmed proper transcription and translation as well as stable expression of the AR gene in the cells.

Applicant's summary and conclusion