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Description of key information

Key value for chemical safety assessment

Skin sensitisation

Link to relevant study records
Reference
Endpoint:
skin sensitisation: in vivo (LLNA)
Type of information:
experimental study
Adequacy of study:
key study
Study period:
2010-01-19 to 2010-04-16
Reliability:
1 (reliable without restriction)
Qualifier:
according to
Guideline:
OECD Guideline 429 (Skin Sensitisation: Local Lymph Node Assay)
Deviations:
no
Qualifier:
according to
Guideline:
EU Method B.42 (Skin Sensitisation: Local Lymph Node Assay)
Deviations:
no
GLP compliance:
yes (incl. certificate)
Type of study:
mouse local lymph node assay (LLNA)
Species:
mouse
Strain:
CBA
Sex:
female
Details on test animals and environmental conditions:
TEST ANIMALS
- Source: Breeder Janvier
- Age at study initiation: 9 weeks
- Weight at study initiation: 20.4 ± 1.1 g
- Housing: individual, crystal polystyrene cages
- Diet (e.g. ad libitum): "SSNIFF R/M-H pelleted maintenance diet", ad lib
- Water (e.g. ad libitum): drinking water filtered using a 0.22 micron meter filter, ad lib
- Acclimation period: 5 days at least


ENVIRONMENTAL CONDITIONS
- Temperature (°C): 19-25
- Humidity (%): 30-70
- Air changes (per hr): 15
- Photoperiod (hrs dark / hrs light): 12/12

IN-LIFE DATES: From: 4 March 2009 To: 02 April 2009
Vehicle:
acetone/olive oil (4:1 v/v)
Concentration:
25, 10, 5, 2.5, 1 and 0 % (v/v)
No. of animals per dose:
4 (at all doses)
Details on study design:
RANGE FINDING TESTS:
- Compound solubility: appropriate solution in AOO vehicle
- Irritation: a range-finding test performed on mice at the concentrations of 10, 25, 50 and 100%

MAIN STUDIES
ANIMAL ASSIGNMENT AND TREATMENT
- Criteria used to consider a positive response: SI ≥ 3

TREATMENT PREPARATION AND ADMINISTRATION:
in the vehicle, at 25 µL on both ears
Positive control substance(s):
hexyl cinnamic aldehyde (CAS No 101-86-0)
Positive control results:
The positive control results were valid: the conditions of the test allow the identification of positive dermal sensitisers.
Parameter:
SI
Remarks on result:
other: see Table 1
Parameter:
other: disintegrations per minute (DPM)
Remarks on result:
other: see Table 1

Table 1:

Concentration
(% v/v in vehicle) 
dpm per node SI  Increase in ear thickness (% between day 1 and day 6)  Interpretation of irritancy 
Vehicle AOO  45.83 - 3.13 -
Test item 1%  45.06 0.98 4.04 Non irritant 
Test item 2.5%  42.75 0.93 6.38 Non irritant
Test item 5%  36.00 0.79 4.12 Non irritant
Test item 10% 74.26 1.62 9.38 Non irritant
Test item 25% 475.96 10.39 20.41 Slightly irritant
HCA 25% 317.92 - - -
Interpretation of results:
not sensitising
Remarks:
Migrated information See in Attached background material "Statement CAE JFN 24th July 2011.pdf" Criteria used for interpretation of results: expert judgment
Conclusions:
Preliminary experiments showed that CAE concentrations > 25% induced a skin irritation at day 6. Thus, the 25% concentration was retained for highest tested concentration for the LLNA. Conclusion was that CAE should be considered as a weak sensitizer since the EC3 value was equal to 12%.
Of note, the 25% CAE concentration was not totally devoid of irritant properties as reported by study director:
i) clinical observation showed dryness of the 25% C CAE-treated ears in all animals (4/4) and erythema in ¼ animals (page 7 and 20/33);
ii) a 20 to 30% ear thickness increase was noted at day 6 in all 25% CAE-treated animals (page 17/33-preliminary exp. and page 22/33 pivotal exp.). Therefore, we interpret the results of the LLNA as false positives because of the use of a “border-line” irritant concentration of CAE.
If the LLNA had been conducted with the non-irritant 10% concentration, it would have been negative.
Executive summary:

According to the study director, the test item should be considered as a weak sensitizer based on the EC3 of 12%.

According to the expert judgment, there are 4 types of arguments for not taking into account the positive CAE LLNA:

i) LLNA is not validated for UVCB;

ii) positive LLNA is not synonymous of skin sensitization properties;

iii) there are strong arguments to consider the positive LLNA as false-positive through an irritant effect of CAE and the vehicle;

iv) there is no bibliographic and epidemiological data on CAE-induced allergy.

Please see the document "Statement CAE JFN 24th July 2011.pdf" for more details.

Based on these arguments, CAE is non-classified as skin sensitiser.

Endpoint conclusion
Endpoint conclusion:
no adverse effect observed (not sensitising)
Additional information:

Weight of evidence determination using expert judgment:

There are 4 types of arguments for not taking into account the positive CAE LLNAs:

i) LLNA is not validated for UVCB;

ii) positive LLNA is not synonymous of skin sensitization properties;

iii) there are strong arguments to consider the positive LLNA as false-positive through an irritant effect of CAE and the vehicle;

iv) there is no bibliographic and epidemiological data on CAE-induced allergy.

Please see attached document below for the Assessment of contact hypersensitivity to CAE in the mouse Local Lymph Node Assay signed by Professor NICOLAS: "Statement CAE JFN July 24th 2011.pdf"

Migrated from Short description of key information:

Skin sensitisation.001: According to the study director, the test item should be classified as a skin sensitizer based on the EC1.4 of 10% v/v. Significant irritancy was observed from 50% v/v only.

Skin sensitisation.002: According to the study director, the test item should be considered as a weak sensitizer based on the EC3 of 12%.

Respiratory sensitisation

Endpoint conclusion
Endpoint conclusion:
no study available

Justification for classification or non-classification

According to Article 9.3 then specifies that "where the criteria cannot be applied directly to available identified information, manufacturer, importers and downstream users shall carry out an evaluation by applying a weight of evidence determination using expert judgment in accordance with section 1.1.1 of Annex 1 to this Regulation weighing all available information having a bearing on the determination of the hazards of the substance of the mixture in accordance with section 1.2 of Annex XI to Regulation (EC) N°1907/2006. 

CLP classification Criteria related to skin sensitization Paragraph 3.4.2.2 of Annex 1 of the CLP provides the rules applicable to the evaluation of information for the determination of the classification of skin sensitizers. Essentially, it provides that a substance shall be classified as skin sensitizer (Category 1) in accordance with the following criteria:

·If there evidence in humans that the substance can lead to sensitization by skin contact in a substantial number of persons, or

·If there are positive results from an appropriate animal test. 

As the false positive LLNAs results cannot be taken into account for classification purpose and there is no evidence of skin sensitization in humans.

 

There is no definitive argument to consider CAE as a skin sensitizer. LLNA is not validated for UVCB. Most of the data obtained with CAE in the LLNA and in the ocular and skin tolerance assays show that CAE is a mild irritant. Furthermore, no human relevant cases of sensitization have been reported.

Therefore, the positive LLNA can be attributed to the irritant properties of CAE yielding to false positive LLNA.

Consequently, CAE is non-classified as skin sensitiser.