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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
Type of information:
experimental study
Adequacy of study:
key study
Study period:
2015-11-02 to 2016-03-16
Reliability:
1 (reliable without restriction)
Rationale for reliability incl. deficiencies:
guideline study

Data source

Reference
Reference Type:
study report
Title:
Unnamed
Year:
2016
Report date:
2016

Materials and methods

Test guideline
Qualifier:
according to guideline
Guideline:
OECD Guideline 473 (In Vitro Mammalian Chromosome Aberration Test)
Version / remarks:
2014
Deviations:
no
GLP compliance:
yes
Type of assay:
in vitro mammalian chromosome aberration test

Test material

Constituent 1
Chemical structure
Reference substance name:
Ethane-1,2-diylbis[dichloromethylsilane]
EC Number:
222-123-0
EC Name:
Ethane-1,2-diylbis[dichloromethylsilane]
Cas Number:
3353-69-3
Molecular formula:
C4H10Cl4Si2
IUPAC Name:
dichloro({2-[dichloro(methyl)silyl]ethyl})methylsilane
Constituent 2
Reference substance name:
1,2-bis[dichloro(methyl)silyl]ethane
IUPAC Name:
1,2-bis[dichloro(methyl)silyl]ethane
Test material form:
other: liquid

Method

Species / strain
Species / strain / cell type:
Chinese hamster lung fibroblasts (V79)
Details on mammalian cell type (if applicable):
- Type and identity of media: minimum essential medium (MEM)
- Properly maintained: yes
- Periodically checked for Mycoplasma contamination: yes
Metabolic activation:
with and without
Metabolic activation system:
phenobarbital (80 mg/kg bw) and -naphthoflavone (100 mg/kg bw) induced rat liver S9
Test concentrations with justification for top dose:
Experiment I:
200, 400, 600 and 800 μg/mL, without metabolic activation
500, 1000 and 1500 μg/mL, with metabolic activation
Experiment II:
400, 600, 800 and 1000 μg/mL, without metabolic activation
Vehicle / solvent:
- Vehicle(s)/solvent(s) used: tetrahydrofuran (THF)
- Justification for choice of solvent/vehicle: The solvent was compatible with the survival of the cells and the S9 activity.
Controlsopen allclose all
Untreated negative controls:
yes
Remarks:
treatment medium
Negative solvent / vehicle controls:
yes
Remarks:
THF
True negative controls:
no
Positive controls:
yes
Positive control substance:
ethylmethanesulphonate
Remarks:
without metabolic activation
Untreated negative controls:
yes
Remarks:
treatment medium
Negative solvent / vehicle controls:
yes
Remarks:
THF
True negative controls:
no
Positive control substance:
cyclophosphamide
Remarks:
with metabolic activation
Details on test system and experimental conditions:
METHOD OF APPLICATION: in medium

ACTIVATION: The S9 supernatant was mixed with S9 co-factors to result in a final protein concentration of 0.75 mg/mL in the cultures. The added co-factors were: 8 mM MgCl2, 33 mM KCl, 5 mM Glucose-6-phosphate, 5 mM NADP in 100 mM sodium-phosphate-buffer pH 7.4.

DURATION

- Exposure duration:
Experiment I: 4 hours exposure, with and without metabolic activation
Experiment II: 21 hours exposure, without metabolic activation
- Expression time (cells in growth medium):
Experiment I: 16 +/- 2 hours
Experiment II: Preparation of the cells straight after the 21-hour exposure
- Fixation time (start of exposure up to fixation or harvest of cells): at 21 hours after the start of exposure

SPINDLE INHIBITOR (cytogenetic assays): Colcemid (0.2 μg/mL culture medium) ) added 2.5 hours before cell preparation
STAIN (for cytogenetic assays): Giemsa

NUMBER OF REPLICATIONS: Duplicate cultures

NUMBER OF CELLS EVALUATED: 150 metaphases per culture

DETERMINATION OF CYTOTOXICITY
- Method: mitotic index and RICC (%) (relative increase in cell count)

OTHER EXAMINATIONS:
- Determination of polyploidy: yes
Evaluation criteria:
The result was considered positive when:
- at least one of the test concentrations exhibits a statistically significant increase compared with the concurrent negative control
- the increase is dose-related when evaluated with an appropriate trend test
- any of the results are outside the distribution of the historical negative control data
Statistics:
Fisher´s exact test - to verify the results in the experiment
Χ2 test for trend - to test whether there is a concentration-related increase in cells with chromosome aberrations

Results and discussion

Test results
Species / strain:
Chinese hamster lung fibroblasts (V79)
Metabolic activation:
with and without
Genotoxicity:
negative
Cytotoxicity / choice of top concentrations:
cytotoxicity
Remarks:
at 800 μg/mL without metabolic activation
Vehicle controls validity:
valid
Untreated negative controls validity:
valid
Positive controls validity:
valid
Additional information on results:
TEST-SPECIFIC CONFOUNDING FACTORS
- Precipitation: Precipitation of the test item was noted at 1000 μg/mL in experiment I with metabolic activation and experiment II without metabolic activation.

ADDITIONAL INFORMATION ON CYTOTOXICITY: In experiment I without metabolic activation at concentration of 800 μg/mL, only one of the two cultures was evaluated for mitotic index and aberrant cells.
Remarks on result:
other: No mutagenic potential

Any other information on results incl. tables

Table 1: Summary, Experiment I and II with and without metabolic activation

 

Dose

Group

Concentration

µg/mL

Relative Mitotic Index

 (%)

RICC

(%)

Mean % Aberrant Cells

Historical Laboratory Negative Control Range

Precipitation

Including

Gaps

Excluding Gaps

Experiment I and II, without metabolic activation

Experiment I

4 hour treatment, 21 hour preparation interval

C

0

102

114

2.3

0.7

0.0% - 4.0 %

aberrant cells

-

S

0

100

100

2.3

0.7

-

aC

0

97

97

2.0

1.7

-

3

200

92

85

5.0

2.0

-

4

400

98

75

3.3

2.3

-

5

600

92

72

3.0

2.3

-

6

800

39

7

2.7

2.0

-

EMS

600

94

78

10.0

6.0

-

 

Experiment II

21 hour treatment, 21 hour preparation interval

C

0

102

110

1.3

0.3

0.0 % - 4.0 %

aberrant cells

-

S

0

100

100

1.7

0.7

-

aC

0

74

114

3.3

1.7

-

6

400

89

88

1.7

1.0

-

7

600

65

87

2.3

1.7

-

8

800

8

25

1.2

0.8

-

9

1000

13

31

4.9

1.8

+

EMS

600

70

78

17.3

15.7

-

Experiment I, with metabolic activation

Experiment I

4 hour treatment, 21 hour preparation interval

C

0

106

103

2.0

1.3

0.0 % - 4.3 %

aberrant Cells

-

S

0

100

100

3.0

2.0

-

aC

0

113

114

3.3

1.3

-

5

500

117

91

5.0

2.7

-

6

1000

119

80

6.0

3.3

+

7

1500

116

88

3.3

1.7

+

CPA

1.5

115

78

10.0

5.7

-

C: Negative Control (Culture Medium)

S: Solvent Control (DMSO)

aC: Acidic Control

CPA: Cyclophosphamide

EMS: Ethylmethanesulfonate

+ : Precipitation

- : No precipitation

Applicant's summary and conclusion

Conclusions:
1,2-Bis[dichloro(methyl)silyl]ethane has been tested for ability to cause chromosome aberrations in Chinese hamster lung fibroblasts V79 according to OECD TG 473 and in compliance with GLP (Eurofins BioPharma, 2016). No increase in the number of cells with aberrations was observed either with or without metabolic activation up to cytotoxic or precipitating concentrations. Appropriate solvent (THF), negative (treatment medium) and positive controls were included and gave expected results. It is concluded that the test substance is negative for the induction of chromosome aberrations under the conditions of this study.

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