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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
Remarks:
Type of genotoxicity: chromosome aberration
Type of information:
migrated information: read-across from supporting substance (structural analogue or surrogate)
Adequacy of study:
key study
Study period:
2008-05-15 to 2008-06-20
Reliability:
1 (reliable without restriction)
Rationale for reliability incl. deficiencies:
other: The study was conducted according to the appropriate OECD test guideline, and in compliance with GLP. Read-across to the registered substance is scientifically justified and reliable.

Data source

Reference
Reference Type:
study report
Title:
Unnamed
Year:
2008
Report date:
2008

Materials and methods

Test guidelineopen allclose all
Qualifier:
according to guideline
Guideline:
OECD Guideline 473 (In Vitro Mammalian Chromosome Aberration Test)
Version / remarks:
1997
Deviations:
no
Qualifier:
according to guideline
Guideline:
other: Commission Directive 2000/32/EC, L1362000, Annex 4A: ”Mutagenicity – In vitro Mammalian Chromosome Aberration Test“, dated May 19, 2000.
Deviations:
no
GLP compliance:
yes (incl. QA statement)
Type of assay:
mammalian cell gene mutation assay

Test material

Constituent 1
Reference substance name:
002996-92-1
Cas Number:
002996-92-1
IUPAC Name:
002996-92-1
Constituent 2
Reference substance name:
Trimethoxyphenylsilane
EC Number:
221-066-9
EC Name:
Trimethoxyphenylsilane
Cas Number:
2996-92-1
IUPAC Name:
trimethoxy(phenyl)silane

Method

Species / strain
Species / strain / cell type:
Chinese hamster lung fibroblasts (V79)
Metabolic activation:
with and without
Metabolic activation system:
Phenobarbital/beta-Naphthoflavone induced rat liver S9
Test concentrations with justification for top dose:
Without activation: 63.9-2045.0 µg/ml; with activation 31.9-1022.5 µg/ml
Vehicle / solvent:
- Vehicle(s)/solvent(s) used: ethanol
- Justification for choice of solvent/vehicle: The solvent was chosen for its solubility properties and its relative non-toxicity to the cell cultures.
Controlsopen allclose all
Untreated negative controls:
no
Negative solvent / vehicle controls:
yes
True negative controls:
no
Positive controls:
yes
Positive control substance:
ethylmethanesulphonate
Remarks:
without activation

Migrated to IUCLID6: final concentration 900 µg/ml
Untreated negative controls:
no
Negative solvent / vehicle controls:
yes
True negative controls:
no
Positive controls:
yes
Positive control substance:
cyclophosphamide
Remarks:
with activation

Migrated to IUCLID6: final concentration 1.4 µg/ml
Details on test system and experimental conditions:
METHOD OF APPLICATION: in medium in Quadriperm dishes

DURATION
- Preincubation period: none
- Exposure duration: 4 hours
- Expression time (cells in growth medium): 14 hours
- Fixation time (start of exposure up to fixation or harvest of cells): 18 hours

SPINDLE INHIBITOR (cytogenetic assays):
STAIN (for cytogenetic assays): Giemsa

NUMBER OF REPLICATIONS: duplicate cultures (2 chambers per test group)

NUMBER OF CELLS EVALUATED: at least 100 per culture

DETERMINATION OF CYTOTOXICITY
- Method: mitotic index; other: reduced cell mumbers

OTHER EXAMINATIONS:
- Determination of polyploidy: yes
- Determination of endoreplication: yes

OTHER: a preliminary test to evaluate cytotoxicity was carried out to determine suitable dose range for the assay.
Evaluation criteria:
A test item is classified as clastogenic if:
a) the number of induced structural chromosome aberrations is not in the range of the laboratory’s historical control data range;
and b) either a concentration-related or a significant increase of the number of structural chromosome aberrations is observed.
Statistics:
Fisher’s exact test

Results and discussion

Test resultsopen allclose all
Species / strain:
Chinese hamster lung fibroblasts (V79)
Metabolic activation:
with
Genotoxicity:
positive
Cytotoxicity / choice of top concentrations:
cytotoxicity
Remarks:
at 511.3 µg/ml and above with S9
Vehicle controls validity:
valid
Untreated negative controls validity:
not applicable
Positive controls validity:
valid
Species / strain:
Chinese hamster lung fibroblasts (V79)
Metabolic activation:
without
Genotoxicity:
negative
Cytotoxicity / choice of top concentrations:
no cytotoxicity nor precipitates, but tested up to recommended limit concentrations
Vehicle controls validity:
valid
Untreated negative controls validity:
not applicable
Positive controls validity:
valid
Additional information on results:
TEST-SPECIFIC CONFOUNDING FACTORS: none. A non-interfering precipitate was observed at 2055.0 µg/ml

RANGE-FINDING/SCREENING STUDIES: significant toxicity was observed in the preliminary cytotoxicity test at concentrations of 1022.5 and 2045.0 µg/ml

CYTOTOXICITY: cytotoxic concentrations did not produce sufficient cells with analysable chromosomes to be included in the analysis.


COMPARISON WITH HISTORICAL CONTROL DATA: control results fall within the range of historical controls. All values of cultures treated with test substance in the presence of metabolic activation showed increases in the number of cells with aberrations excluding gaps clearly exceeded the control values.
Remarks on result:
other: all strains/cell types tested
Remarks:
Migrated from field 'Test system'.

Any other information on results incl. tables

No polyploidy was observed in any culture

Table 1 Summary of results of chromosome aberration study

 Activation  Concentration µg/ml Cell numbers %   Mitotic index %  Aberrant cells inc gaps  Aberrant cells exc gaps Aberrant cells with exchanges 
Without activation Solvent control  100  100  2.5  1.0  0
Positive control  NT  115.4  15.5  12.5*  7.5
 127.8  103.1  138.5  1.5  1.5  0.5
 255.6  93.5  1.2.0  3.0  2.0  0
 511.3  89.3  102.4  0  0  0
With activation         Solvent control  100  100  2.5  2.5  0.5
 Positive control  NT  72.8  18.5  16.5*  7.5
 127.8  81.7  89.0  8.5  7.5*  5.5
 255.6  55.1  78.0  14.5  13.5*  6.5
 511.3  64.6  69.1  18.0  16.0*  5.0
NT not tested * Aberration frequency statistically significantly higher than corresponding control values (p < 0.05)                                                                                                                               

Table 2 Results of chromosome analysis without activation (total of 200 cells scored)


 Treatment    Solvent control  Positive control  Low dose 127.8 µg/ml Mid dose 255.6 µg/ml   High dose 511.3  µg/ml  
Cytotoxicity     no   no no  no 

no*

Chromatid aberrations 

 break

12 

 

fragment 

0  

 

deletion 

 

exchange 

21 

Chromosome aberrations 

break 

 0

 

fragment 

 

deletion 

 

exchange 

  0

mitotic index %    

100 

 115.4

138.5 

 102.0

102.4 

*cultures treated with toxic concentrations did not have sufficient analysable chromosomes to be included in the analysis

Table 3 Results of chromosome analysis with activation (total of 200 cells scored)

 Treatment  

Solvent control 

Positive control 

Low dose 127.8 µg/ml

Mid dose 255.6 µg/ml 

High dose 511.3 µg/ml 

Cytotoxicity    

no 

 no

no 

 no

 no

Chromatid aberrations 

 break

19 

16 

25 

 

fragment 

 

deletion 

 

exchange 

15

11 

15 

10 

Chromosome aberrations 

break 

 

fragment 

 

deletion 

 

exchange 

mitotic index  %  100 72.8   89.0  78.0  69.1

Applicant's summary and conclusion

Conclusions:
Interpretation of results (migrated information):
positive with metabolic activation
negative without metabolic activation

Trimethoxyphenylsilane CAS 2996-92-1 has been tested in a valid in vitro cytogenicity study according to OECD 473 and under GLP. A statistically significant, dose dependent increase in the number of cells with aberrations excluding gaps were detected in the evaluated concentration tested in the presence of activation. It is concluded by the authors of the study that this result is biologically significant and that trimethoxyphenylsilane is positive for the induction of structural chromosome aberrations (clastogenic) under the conditions of the test.