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Toxicological information

Genetic toxicity: in vitro

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Administrative data

Endpoint:
in vitro cytogenicity / chromosome aberration study in mammalian cells
Remarks:
Type of genotoxicity: chromosome aberration
Type of information:
experimental study
Adequacy of study:
key study
Study period:
24 March - 11 Jun, 1999
Reliability:
1 (reliable without restriction)
Rationale for reliability incl. deficiencies:
other: Done under GLP and OECD Methods

Data source

Reference
Reference Type:
study report
Title:
Unnamed
Year:
1999

Materials and methods

Test guidelineopen allclose all
Qualifier:
according to
Guideline:
other: Corning Hazleton protocol 449PCO
Qualifier:
equivalent or similar to
Guideline:
OECD Guideline 473 (In Vitro Mammalian Chromosome Aberration Test)
GLP compliance:
yes
Type of assay:
in vitro mammalian chromosome aberration test

Test material

Reference
Name:
Unnamed
Type:
Constituent

Method

Species / strain
Species / strain / cell type:
mammalian cell line, other: Human lymphocytes
Details on mammalian cell type (if applicable):
Human venous blood
Metabolic activation:
with and without
Metabolic activation system:
Exogenous metabolic activation
Test concentrations with justification for top dose:
Target dose 2.00 mg/ml; dosing volume of 1 % (10 ul/ml). In DMSO.

Concentration range in the dose range : 3.98 ... 2010 µg/ml.
Reduction in mitotic index < 11%, 34% and 37% at 127, 1010 and 2010 dosed for 3 hours without metabolic activation as compared to vehicle controls.
Reduction in mitotic index of 19%, 19% , 33 % and 81% at 15.9, 253, 1010 and 2010 dosed for 19.1 hours without metabolic activation as compared to vehicle controls.
Reduction in mitotic index 3% and 52% at 505 and 2010 dosed for 3 hours with metabolic activation as compared to vehicle controls.

Concentrations in the initial experiment dosed for 3 hrs, (with and without metabolic activation): 235, 336, 480, 686, 980, 1400 and 2000 µg/ml and harvested 22 hrs after treatment. Chromosomal aberrations were analyzed from the cultures treated with 480, 686, 980, and 2000 µg/ml without metabolic activation.
Chromosomal aberrations were analyzed from the cultures treated with 686, 980, 1400 and 2000 µg/ml with metabolic activation.
Chromosomal aberrations were analyzed from the cultures treated with 480, 686, 980, and 2000 µg/ml without metabolic activation.
Chromosomal aberrations were analyzed from the cultures treated with 686, 980, 1400 and 2000 µg/ml with metabolic activation.

Concentrations in confirmatory test dosed for 19.5 hrs (without metabolic activation): 159, 318, 636, 848, 1130, 1500 and 2000 µg/ml and harvested 22.2 hrs and 46 hrs after treatment.
Concentrations in confirmatory test dosed for 43.5 hrs (without metabolic activation): 79.5, 159, 318, 636, 848, 1130, 1500 and 2000 µg/ml and harvested 22.2 hrs and 46 hrs after treatment.

Concentrations in confirmatory test dosed for 3 hrs (with metabolic activation): 318, 636, 848, 1130, 1500 and 2000 µg/ml and harvested 22.2 hrs and 46 hrs after treatment.
Vehicle / solvent:
vehicle : Dimethylsulfoxide
Controlsopen allclose all
Untreated negative controls:
yes
Negative solvent / vehicle controls:
yes
Remarks:
DMSO
Positive controls:
yes
Positive control substance:
mitomycin C
Remarks:
without S9
Untreated negative controls:
yes
Positive controls:
yes
Positive control substance:
cyclophosphamide
Remarks:
with S9
Details on test system and experimental conditions:
first experiment:
Exposure period (with metabolic activation): 3 hrs
Exposure period (without metabolic activation): 3 hrs

second experiment:
Exposure period (with metabolic activation): 3 hrs
Exposure period (without metabolic activation): 22.2 and 46hr

at 2 hours before harvest each of the cultures were treated with colcemid solution ( 0.1µg/ml) to block cells at the metaphase stage of mitosis.

cultured medium-for each culture heparinsed whole blood was added to culture medium containing RPMI 1640 supplemented with 15% fetal bovine serum, 1% photohaemogglutinin, 1% penicillin streptomycin and 1% L-glutamine. Total culture volume: 10ml. The culture was incubated on a slope at 37°C in a humidified atmosphere at 5% CO2/95% air for 48 hours.
Evaluation criteria:
100 cells from each replicate culture at at least 3 concentrations and the negative and vehicle (solvent) control cultures were analyzed for the different types of chromosomal aberration. At least 25 cells were analyzed for chromosomal aberrations from the positive control cultures.
Mitotic index was assessed by analyzing the number of mitotic cells in 1,000 cells and the ratio was expressed as a ratio of the mitotic cells.
Statistics:
Cochran-Armitgae test for linear trend and Fisher's Exact test comparing treated to veicle controls at p<0.01 significance.

Results and discussion

Test results
Species / strain:
human lymphoblastoid cells (TK6)
Metabolic activation:
with and without
Genotoxicity:
negative
Cytotoxicity / choice of top concentrations:
cytotoxicity
Vehicle controls validity:
valid
Positive controls validity:
valid
Additional information on results:
No significant increase in cells with chromosomal aberrations was observed at the concentrations analyzed with and without metabolic activation.

vehicle and positive controls were acceptable and valid.
Remarks on result:
other: all strains/cell types tested
Remarks:
Migrated from field 'Test system'.

Applicant's summary and conclusion

Conclusions:
Interpretation of results (migrated information):
negative

No significant increase in cells with chromosomal aberrations was observed at the concentrations analyzed with and without metabolic activation.Test substance was considered negative for inducing chromosomal aberrations and results were verified in independently conducted confirmatory assay with multiple harvests.