(S)-3-methyl-2-(2-oxotetrahydropyrimidine-1-yl)butyric acid

Genetic toxicity: in vitro

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:

April 5 - August 17, 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

Report date:

1999

Materials and methods

Test guideline

Test guideline 1

Qualifier:

according to guideline

Guideline:

other: Corning Hazleton protocol 449PCO, edition 1, modified for Abbott Labs

Test guideline 2

Qualifier:

equivalent or similar to guideline

Guideline:

OECD Guideline 473 (In Vitro Mammalian Chromosome Aberration Test)

GLP compliance:

yes

Type of assay:

in vitro mammalian chromosome aberration test

Test material

Test material information

Constituent 1

Reference substance name:

-

EC Number:

430-900-2

EC Name:

-

Cas Number:

192725-50-1

Molecular formula:

Hill formula: C9H16N2O3 CAS formula: C9H16N2O3

IUPAC Name:

3-methyl-2-(2-oxo-1,3-diazinan-1-yl)butanoic acid

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 2.5 mg/ml; dosing volume of 1 % (10 ul/ml). In DMSO.

Concentration range in the dose range : 4.93 ... 2500 µg/ml.
Reduction in mitotic index of 3% at 625 ug/ml dosed for 3 hours without metabolic activation as compared to vehicle controls.
Reduction in mitotic index of 29 % at 2500 ug/ml dosed for 19.1 hours without metabolic activation as compared to vehicle controls.
No reduction in mitotic index noted when dosed for 3 hours with metabolic activation as compared to vehicle controls.

Concentrations in the initial experiment dosed for 3 hrs, (without metabolic activation): 861-2500 µg/ml and harvested 22 hrs after treatment.
Concentrations in the initial experiment dosed for 3 hrs, (with metabolic activation): 861-2500 µg/ml and harvested 22 hrs after treatment.

Chromosomal aberrations were analyzed from the cultures treated with 861, 1230, 1750 and 2500 µg/ml with and without metabolic activation.

Concentrations in confirmatory test dosed for 19.5 hrs (without metabolic activation): 1130-2500 µg/ml and harvested 22.0 hrs after treatment.
Concentrations in confirmatory test dosed for 43.2 hrs (without metabolic activation): 1130-2500 µg/ml and harvested 46.1 hrs after treatment.

Concentrations in confirmatory test dosed for 3 hrs (with and without metabolic activation): 1130-2500 µg/mg and harvested 22.0 hrs and 46.1 hrs after treatment.

Chromosomal aberrations were analyzed from the cultures treated with 1130, 1500, 2000 and 2500 µg/ml with and without metabolic activation.

Vehicle / solvent:

vehicle : Dimethylsulfoxide

Controls

Controls 1

Untreated negative controls:

yes

Negative solvent / vehicle controls:

yes

Remarks:

DMSO

Positive controls:

yes

Positive control substance:

mitomycin C

Remarks:

without S9

Controls 2

Untreated negative controls:

yes

Positive controls:

yes

Positive control substance:

cyclophosphamide

Remarks:

with S9

Details on test system and experimental conditions:

initial experiment:
Exposure period (with metabolic activation): 3 hrs
Exposure period (without metabolic activation): 3 hrs

confirmatory experiment:
Exposure period (with metabolic activation): 3 hrs
Exposure period (without metabolic activation): 22.0 and 46.1 hr

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-Armitage 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 appreciable level of toxicity was observed as evident by decreased mitotic index. The test compound was tested to a concentration that required pH adjustment. Test concentrations analyzed for clastogenicity were the highest concentrations tested. Samples of test substance in DMSO were not analyzed for drug content in the test but an assay for stability after the study was completed and found to be stable.

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.