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Toxicological information

Genetic toxicity: in vitro

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Administrative data

Endpoint:
in vitro gene mutation study in mammalian cells
Remarks:
Type of genotoxicity: gene mutation
Type of information:
experimental study
Adequacy of study:
key study
Study period:
2013
Reliability:
1 (reliable without restriction)
Rationale for reliability incl. deficiencies:
guideline study

Data source

Reference
Reference Type:
study report
Title:
Unnamed
Year:
2013
Report Date:
2013

Materials and methods

Test guideline
Qualifier:
according to
Guideline:
other: OECD 487
GLP compliance:
yes (incl. certificate)
Type of assay:
mammalian cell gene mutation assay

Test material

Reference
Name:
Unnamed
Type:
Constituent
Test material form:
solid: particulate/powder
Remarks:
migrated information: powder
Details on test material:
- Name of test material (as cited in study report): BRUGGOLEN M12 / N, N’-hexane-1,6-diylbis(hexahydro-2-oxo-1H-azepine-1-carboxamide)
- Substance type: organic
- Physical state: solid
- Analytical purity: > 99.5%
- Impurities (identity and concentrations): not specified
- Lot/batch No.: 12030701
- Expiration date of the lot/batch: 30 November 2013
- Stability under test conditions: stable
- Storage condition of test material: room temperature

Method

Species / strain
Species / strain / cell type:
Chinese hamster Ovary (CHO)
Additional strain / cell type characteristics:
not specified
Metabolic activation:
with and without
Metabolic activation system:
S9
Test concentrations with justification for top dose:
1970, 1130, 643, 368, 210, 120, 68.6, 39.2, 22.4 and 12.8 μg/mL
Vehicle / solvent:
- Vehicle(s)/solvent(s) used: DMSO
Controls
Untreated negative controls:
yes
Negative solvent / vehicle controls:
yes
True negative controls:
no
Positive controls:
yes
Positive control substance:
cyclophosphamide
mitomycin C
other: colchicine
Evaluation criteria:
A test item will be considered as clearly positive in this assay, if the following criteria are met:
(i)
Significant increases in the proportion of micronucleated cells over the concurrent controls occur at one or more concentrations.
(ii)
The proportion of micronucleated cells at such data points exceeds the normal range. If the increases fall within the range of values, normally observed in the negative control cultures, the test item cannot be classified as positive. Any significant increases over the concurrent negative controls will therefore be compared with historical control values derived from recent studies.
(iii) There is a significant dose-relationship.
Results which only partially satisfy the above criteria will be dealt with on a case-by-case basis. Similarly, positive responses, seen only at high levels of cytotoxicity, will require careful interpretation when assessing their biological significance.
Statistics:
The numbers of binucleated cells with micronuclei in the control and treated cultures will be compared using an appropriate statistical method. The solvent controls will be used as the reference point for comparison in the statistical evaluation and the evaluation of the results.

Results and discussion

Test results
Species / strain:
Chinese hamster Ovary (CHO)
Metabolic activation:
with and without
Genotoxicity:
positive
Cytotoxicity / choice of top concentrations:
cytotoxicity
Remarks:
Cytotoxicity above 253 ug/mL in the absence and presence of S9 metabolism.
Vehicle controls validity:
valid
Untreated negative controls validity:
valid
Positive controls validity:
valid
Additional information on results:
TEST-SPECIFIC CONFOUNDING FACTORS
- Effects of pH: Following treatment with the test item, no remarkable variation of pH over the control values was observed in the absence or presence of S9 metabolism.
- Effects of osmolality: Following treatment with the test item, no remarkable variation of osmolality over the control values was observed in the absence or presence of S9 metabolism.
Remarks on result:
other: all strains/cell types tested
Remarks:
Migrated from field 'Test system'.

Any other information on results incl. tables

 

 

 

 

 

 

 

Presence of S9 metabolism

Absence of S9 metabolism

Treatment

Dose level

 

 

 

 

 

µg/mL

%Mn cells

Sig.

%Cytotoxicity

%Mn cells

Sig.

%Cytotoxicity

 

 

 

 

 

 

Untreated

0.0

1.00

 

15

0.60

 

3

 

 

 

 

 

 

Solvent 1%

0.0

1.05

 

0

0.60

 

0

 

 

 

 

 

 

 

 

Test item

176

-

 

-

0.65

NS

22

 

 

 

 

 

 

 

 

Test item

211

1.35

NS

18

1.10

NS

22

 

 

 

 

 

 

Test item

253

11.0

***

35

3.60

***

58

 

 

 

 

 

 

Test item

311

13.1

***

64

-

-

-

 

 

 

 

 

 

 

 

Mitomycin-C

0.300

-

-

-

5.70

***

19

 

 

 

 

 

 

Colchicine

2.50

-

-

-

8.90

***

§

 

 

 

 

 

 

Cyclophosphamide

10.0

5.80

***

52

-

-

Key:

 

%Mn cells                                                 : Percentage of cells bearing micronuclei

Sig.          : Statistical significance

NS           : Not significant

-              : Not tested or not selected for scoring

*            : Statistically significant at p<0.05

**           : Statistically significant at p<0.01

***         : Statistically significant at p<0.001

§             : not calculated - induction of multinucleation was attributable to multipolar division as a

                 consequence of induction of multipolar spindles

Applicant's summary and conclusion

Conclusions:
Interpretation of results (migrated information):
positive

annex VIII, 8.4
Executive summary:

On the basis of the results obtained, it is concluded that BRUGGOLEN M12 induces micronuclei in Chinese hamster V79 cells after in vitro treatment in the absence and presence of S9 metabolic activation, under the reported experimental conditions.

Therefore, according to ECHA's guidance R.7a "Endpoint specific guidance", no second in vitro testing has to be performed but a testing proposal for an in vivo testing (REACH Annex IX) has to be submitted (see section 7.6.2).