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Administrative data

Endpoint:
additional ecotoxicological information
Type of information:
experimental study
Adequacy of study:
supporting study
Reliability:
2 (reliable with restrictions)
Rationale for reliability incl. deficiencies:
other: Well documented study which meets generally accepted scientific standards
Cross-reference
Reason / purpose:
reference to same study

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:
The measurement was made employing a Cell-based androgen reporter assay using the Chinese hamster ovarian cell line (CHO K1) in the 96 -well format. The purpose is high throughput screening for endocrine disrupting (ED) substances in environmental and biological samples. To this end, CHO cells were cotransfected with plasmids encoding mouse mammary tumour virus-neomycin-luciferase and human androgen receptor (hAR), and 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 study / information:
Endocrine effects

Test material

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

Results and discussion

Any other information on results incl. tables

Tested for androgenic or anti-androgenic effects, DCDPS gave in vitro a negative response.

None of the tested compounds appeared to have any androgenic activity in the cell line up to the concentration of 50 µmol/L, when tested alone.

Applicant's summary and conclusion

Conclusions:
Tested for anti-androgenic and androgenic effects, DCDPS gave in vitro a negative response at a concentration of 0.1 µmol/L = 0.287 ng/L.
Executive summary:

The development and optimization of a cell-based androgen reporter assay using the Chinese hamster ovarian cell line (CHO K1) in the 96 -well format is described. The intent is a use as screening system for identifying endocrine disrupting substances (ED) in a high throughput manner in environmental and biological samples. To this end, CHO cells were cotransfected with plasmids encoding mouse mammary tumour virus-neomycin-luciferase and human androgen receptor (hAR), and 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. Stimulation of the cells with androgens for 24 h resulted in about 15-fold stimulation of luciferase activity, with the minimum effective dose of testosterone being 0.1 nmol/L. Potent steroidal and non-steroidal anti-androgens, such as hydroxyflutamide and cyproterone acetate, significantly inhibited the androgen-induced transactivation. Non-androgenic steroids like estradiol, progesterone, dexamethasone and cortisol 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.

About 60 different substances (mostly pesticides or their metabolites, and common industrial chemicals including DCDPS) were screened with the cell line for their ability to stimulate luciferase activity (indicating androgenic activity) or inhibit that evoked by 0.1 nmol/L R1881 (indicating anti-androgenic activity), used as a positive androgenic control. About 10 highly potent anti-androgenic chemicals were identified. The most potent anti-androgenic compounds identified included bisphenol A, α-hexachlorocyclohexane, vinclozolin and 4,4-DDE (CAS 72-55-9), a structural analogue to DCDPS, which gave a negative response. These compounds had alone either no effect or were weak agonists (with cytotoxic effects at very high concentrations), but none showed any significant agonistic activity. None of the tested compounds appeared to have any androgenic activity in the cell line up to the concentration of 50 µmol/L, when tested alone.

In conclusion, it is demonstrated the bioassay based on this cell line provides a reliable test for detecting androgenic and anti-androgenic compounds. DCDPS was not active as androgen or anti-androgen endocrine disruptor.