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Toxicological information

Genetic toxicity: in vivo

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Administrative data

Endpoint:
genetic toxicity in vivo, other
Remarks:
chromosomal aberration assay and micronucleus test
Type of information:
experimental study
Adequacy of study:
supporting study
Reliability:
2 (reliable with restrictions)
Rationale for reliability incl. deficiencies:
study well documented, meets generally accepted scientific principles, acceptable for assessment

Data source

Reference
Reference Type:
publication
Title:
Progestin (norethisterone)-induced genetic damage in mouse bone marrow
Author:
Shyama SK, Rahiman MA
Year:
1993
Bibliographic source:
Mutation Research, 300 (1993) 215-221

Materials and methods

Principles of method if other than guideline:
8-10-week-old healthy female Swiss mice (average body weight 25 g) were administered doses of 0.3, 1.5, 3.0, 6.0, 12.0, 18.0, 24.0 and 30.0 mg/kg body weight test item for 15 consecutive days by gavage.

Chromosome analysis
Mice were injected intraperitoneally with 0.2 ml of 0.025% colchicine, 1.5 h before they were killed by cervical dislocation. Bone marrow chromosomal preparations were made according to the method of Tjio and Whang (1962), using 0.56% potassium chloride as hypotonic solution. 100 metaphase spreads per animal were analysed. Breaks were identified as unstained regions present along the chromatids wider than the thickness of the chromatids that bear them. Transfer of the terminal portion of one chromosome to another leading to unequal length of arms was scored as a translocation. Improper spreading with stickiness and clumping of chromosomes was identified as stickiness. The extreme of stickiness represented by sticky degeneration of the chromosome was noted as pulverisation. Stickiness and pulverisations were scored together. The mitotic index (MI) was calculated by analysing 2000 cells per animal.

Micronucleus test
Experimental animals were killed at different time intervals. Bone marrow preparations were made according to the method of Schmid (1973), with a slight modification, i.e., the fetal calf serum was replaced by 5% bovine albumin solution in phosphate-buffered saline (PBS) (Seetharama Rao et al., 1983). Smears were stained with May-Grünwald Giemsa and analysed for the presence of MN in both polychromatic erythrocytes (PCEs) and normochromatic erythrocytes (NCEs). 2000 PCEs and the corresponding number of NCEs per animal were analysed for the presence of MN. The ratio of PCEs to NCEs (P/N) was calculated for all treated and control groups.
GLP compliance:
not specified
Type of assay:
other: chromosomal aberration assay and micronucleus test

Test material

Constituent 1
Chemical structure
Reference substance name:
Norethisterone
EC Number:
200-681-6
EC Name:
Norethisterone
Cas Number:
68-22-4
Molecular formula:
C20H26O2
IUPAC Name:
17-ethynyl-17-hydroxyestr-4-en-3-one

Test animals

Species:
mouse
Strain:
Swiss
Sex:
female

Administration / exposure

Route of administration:
oral: gavage
Vehicle:
water
Duration of treatment / exposure:
15 d
Frequency of treatment:
daily
Doses / concentrations
Remarks:
0.3, 1.5, 3.0, 6.0, 12.0, 18.0, 24.0 and 30.0 mg/kg body weight
No. of animals per sex per dose:
3
Control animals:
yes, concurrent vehicle
Positive control(s):
cyclophosphamide

Examinations

Tissues and cell types examined:
bone marrow

Results and discussion

Test resultsopen allclose all
Sex:
female
Genotoxicity:
negative
Remarks:
micronucleus assay
Vehicle controls validity:
valid
Positive controls validity:
valid
Sex:
female
Genotoxicity:
positive
Remarks:
chromosome aberration assay
Vehicle controls validity:
valid
Positive controls validity:
valid

Applicant's summary and conclusion

Conclusions:
equivocal
Executive summary:

Norethisterone was assessed for their in vivo genotoxic effect on the bone marrow cells of Swiss albino mice. The chromosomal aberration assay and the micronucleus test were employed for the study.

Dose-response analysis was carried out for the above doses 24 h after the final feeding (15 d; 0.3, 1.5, 3.0, 6.0, 12.0, 18.0, 24.0 and 30.0 mg/kg body weight). Time-response studies were done using mice fed 3.0 mg/kg/day, 6, 12, 24, 48, 96 h and 1, 2 and 3 weeks after the final feeding.

No treatment-related effect on the rate of micronuclei was observed after any of the doses. In addition, no increase of the incidence of chromosome breaks or chromosomal translocations which were designated as "classical" chromosomal aberrations were noted.

However, the incidence of so-called "non-classical" chromosomal aberrations such as "stickiness" and "pulverisations" was increased from 3.0 mg/kg/day onwards. Similar findings were said to occur in studies on the natural steroid progesterone. These "non-classical" findings weretransientwith a maximum at 12 and 24 hours after the final treatment and a sharp decline thereafter. A possible drug interaction with the nucleoproteins was discussed as one possible underlying mechanism which should account for the absence of a concomitant effect on the rate of micronuclei.