Registration Dossier

Administrative data

Endpoint:
in vivo mammalian somatic cell study: cytogenicity / erythrocyte micronucleus
Type of information:
experimental study planned
Study period:
The test will be conducted after a final decision on the requirement to carry out the proposed test has been taken and a deadline to submit the information required has been set by the Agency.
Justification for type of information:
TESTING PROPOSAL ON VERTEBRATE ANIMALS

NON-CONFIDENTIAL NAME OF SUBSTANCE:
- Name of the substance on which testing is proposed to be carried out: N-(2-(4-amino-N-ethyl-m-toluidino)ethyl)methanesulphonamide sesquisulphate (CAS 25646-71-3)

CONSIDERATIONS THAT THE GENERAL ADAPTATION POSSIBILITIES OF ANNEX XI OF THE REACH REGULATION ARE NOT ADEQUATE TO GENERATE THE NECESSARY INFORMATION:
- Available GLP studies: No adequate and reliable GLP studies addressing genetic toxicity in vivo are available with the test substance itself or similar substances defined according to the general rules for grouping of substances and read-across approach laid down in Annex XI, Item 1.5, of Regulation (EC) No. 1907/2006.
- Available non-GLP studies: No adequate and reliable non-GLP studies addressing genetic toxicity in vivo are available with the test substance itself or similar substances defined according to the general rules for grouping of substances and read-across approach laid down in Annex XI, Item 1.5, of Regulation (EC) No. 1907/2006.
- Historical human data: not available
- (Q)SAR methods: not applicable to assess the full scope of an in vivo genetic toxicity study
- In vitro methods:
An Ames test was conducted in 2018 with N-(2-(4-amino-N-ethyl-m toluidino)ethyl)methanesulphonamide sesquisulphate according to OECD Guideline 471 which was judged to be positive with and without metabolic activation (reference 7.6.1-1).
Briefly, the Salmonella typhimurium strains TA 1535, TA 1537, TA 98, TA 100 and the Escherichia coli WP2 uvrA were tested according to the direct plate and the pre-incubation method in the absence and presence of a metabolic activation system (Aroclor 1254-induced rat liver S9-mix). Test substance concentration, dissolved in water, from 1.7 to 5000 µg/plate were used for the dose-range finding experiment in Salmonella typhimurium strain TA100 and Escherichia coli strain WP2uvrA (direct plate assay). Test substance concentrations from 17 to 5000 µg/plate were used in the first experiment in Salmonella typhimurium strains TA 1535, TA 1537 and TA 98 (direct plate assay) and in the second experiment in Salmonella strains TA 1535, TA 1537, TA 98 and TA 100 and Escherichia coli strain WP2uvrA (pre-incubation assay). Cytotoxicity, evident by a decrease in the number of revertants and/or reduction of the bacterial background lawn, was observed in all tester strains from 1600 µg/plate onwards without metabolic activation and at 5000 µg/plate with metabolic activation, except for tester strain WP2 uvrA where no toxicity was observed. The test substance induced significant dose-related increases in the number of revertant colonies in the tester strain TA100 both in the absence and presence of S9-mix and in the WP2 uvrA strain only without metabolic activation. The included positive and negative controls showed the expected results. Under the conditions of this experiment, the test substance did show mutagenicity in one of the selected S. typhimurium strains in the presence and absence of metabolic activation and in the E. coli strain only in the absence of metabolic activation.
The clastogenic activity of the test substance dissolved in DMSO was investigated in an in vitro mammalian chromosome aberration test in human lymphocytes of fresh heparinised human whole blood cultures performed according to OECD Guideline 473 in 1986 (7.6.1-2). Test substance concentrations of 1.4 - 900 µg/mL (preliminary experiment), 7.5 - 30 µg/mL (main experiment: without metabolic activation) and 3.75 -15 µg/mL (main experiment: with metabolic activation) were applied for an exposure period of 24 hours. Cell division was arrested by the addition of colcemid, three hours before the cells were harvested. In the preliminary study marked depression of cell division was observed at concentrations of 36 µg/mL (65% reduction of mitotic index over control values without S-9 mix, 93% reduction with S-9 mix). In the main experiment statistically significant increase in % aberrant cells was observed at test substance concentration of 7.5 and 15 µg/mL with and 15 and 30 µg/mL without metabolic activation. In the absence of S-9 mix no reduction in mitotic activity were observed. In the presence of S-9 mix mean mitotic indices were 7.6, 7.3 and 6.1 for treatment levels of 3.75, 7.5 and 15 µg/mL, showing an approximately 26% reduction in mitotic activity at the highest test substance concentration tested compared to the control value of 8.2. The negative as well as the positive controls showed the expected results. Under the conditions of the performed in vitro chromosomal aberration study in peripheral human lymphocytes, the test substance showed clastogenic activity with and without metabolic activation.

- Weight of evidence: no weight of evidence data available

- Grouping and read-across: no read-across data available

Data source

Materials and methods

Test guideline
Qualifier:
according to
Guideline:
OECD Guideline 474 (Mammalian Erythrocyte Micronucleus Test)
GLP compliance:
yes

Results and discussion

Applicant's summary and conclusion