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Description of key information

Data available for the structurally and functionally similar read across chemicals has been reviewed to determine the skin sensitization potential of the test chemical. Based on the summarized,it can be concluded that the testchemical is unable to cause skin sensitization and considered as not sensitizing. Comparing the above annotations with the criteria of CLP regulation, it can be classified under the category “Not Classified”.

Key value for chemical safety assessment

Skin sensitisation

Link to relevant study records
Reference
Endpoint:
skin sensitisation, other
Remarks:
(LLNA and Non -LLNA)
Type of information:
experimental study
Adequacy of study:
weight of evidence
Reliability:
2 (reliable with restrictions)
Rationale for reliability incl. deficiencies:
data from handbook or collection of data
Remarks:
experimental data of read across substances
Justification for type of information:
Data for the target chemical is summarized based on the structurally similar read across chemicals
Reason / purpose:
read-across source
Reason / purpose:
read-across source
Reason / purpose:
read-across source
Qualifier:
according to
Guideline:
other: as mentioned below
Principles of method if other than guideline:
WoE report is based on 3 skin sensitization studies as- WoE-2, WoE-3 and WoE-4.
Skin sensitization of test chemical was determined by performing patch tests on humans.

Skin sensitization of test chemical was determined by performing patch tests on humans.
GLP compliance:
not specified
Type of study:
other: 1, 2. mouse local lymphnode assay (LLNA) 3. Human maximization test
Justification for non-LLNA method:
Not specified
Species:
other: 1, 2. mouse 3.human
Strain:
other: 1,2.CBA 3.not applicable
Sex:
female
Details on test animals and environmental conditions:
1.Young adult (8–12 weeks old) CBA/Ca strain female mice (Harlan Seralab, Oxon, UK) were used throughout these studies. Animals were housed four per cage on flushing metal racks. Food (SDS PCD pelleted diet; Special Diets Services, Witham, Essex, UK) and water were available ad libitum.

2.TEST ANIMALS
Source: No data
Age at study initiation: 7-12 weeks

ENVIRONMENTAL CONDITIONS: No data

3.Not applicable
Route:
other: 3.epicutaneous, occlusive
Vehicle:
petrolatum
Concentration / amount:
10%
Day(s)/duration:
Not specified
Adequacy of induction:
not specified
No.:
#1
Route:
other: 3.epicutaneous, open
Vehicle:
petrolatum
Concentration / amount:
10%
Day(s)/duration:
not specified
Adequacy of challenge:
not specified
No. of animals per dose:
3.25
Details on study design:
3.No data available
Challenge controls:
3.No data available
Positive control substance(s):
not specified
Vehicle:
other: 1.dimethyl sulphoxide 2.acetone/olive oil (4:1 v/v)
Concentration:
1.0% ;25% ;50% ;75%
2.1%, 2.5%, 5%, 10% or 20% (25µl)
No. of animals per dose:
1.24
2.5 female mice/dose
Details on study design:
1. Groups of mice (n=4) were exposed topically on the dorsum of both ears to 25 l of various concentrations of the test chemicals, or to the same volume of the relevant vehicle alone, daily for 3 consecutive days. Five days following initiation of exposure all mice were injected via the tail vein with 250 l of phosphate buffered saline (PBS) containing 20 Ci of [3H] methyl thymidine. Five hours later the mice were sacrificed and the draining auricular lymph nodes excised and pooled for each experimental group. Single cell suspensions of LNC were prepared by mechanical disaggregation through 200-mesh stainless steel gauze. Pooled LNC were washed twice with PBS and precipitated in 5% trichloroacetic acid (TCA) at 4°C overnight. Pellets were then resuspended in 1 ml of TCA and transferred to 10 ml of scintillation fluid (Optiphase ‘Hisafe 3’, Wallac, Turku, Finland). The incorporation of 3H-TdR was measured by -scintillation counting as disintegrations per minute (dpm) per node for each experimental group. In each case a stimulation index (SI) relative to the concurrent vehicle-treated control was derived; an SI of 3 or greater being indicative of a chemical possessing the potential to cause contact sensitization.
2.Treatment was performed daily for 3 consecutive days. Five days after initiation of exposure all mice were injected via the tail vein with 250µl of PBS containing 20µCi of tritiatied thymidine. The mice were sacrificed 5 hours later, and draining the auricular lymph nodes were excised and pooled for each experimental group or each individual animal. The incorporation of tritiated thymidine measured by β-scintillation counting and was reported in disintegrations /minute. An SI was calculated for each chemical group as the ratio of disintegrations/minute of the treated group to the disintegrations/minute of the concurrent vehicle control group. A substance was classified skin sensitizer , if at one or more than one concentrations, it induced a three-fold or greater increase in local lymph node proliferative activity when treated with the concurrent vehicle treated controls (SI ≥3)


Statistics:
2.The approach to estimation of the relative skin sensitization potential is based on the mathematical estimation of the concentration of chemical necessary to obtain a threshold positive response (SI = 3); this is termed as the EC3 value

Refer Table 1 and 2 for more information
Parameter:
other: 1.EC3
Value:
> 75
Remarks on result:
other: The test chemical failed at all test concentrations (maximum concentrations of 75%) to induce a positive response in the local lymph node assay with relatively low levels of thymidine incorporation being recorded at all concentrations tested.
Parameter:
other: 2.EC3
Value:
> 20
Remarks on result:
other: The calculated EC3 value for test chemical was >20.0%
Cellular proliferation data / Observations:
1.The test chemical failed at all test concentrations (maximum concentrations of 75%) to induce a positive response in the local lymph node assay with relatively low levels of thymidine incorporation being recorded at all concentrations tested.

2.The calculated EC3 value for test chemical was >20.0%

2.

Table 1:Classification of relative skin sensitization potency by local lymphnode assay (LLNA) EC3 values

EC3 values (%)

Potency classification

≥10 to ≤100

Weak

≥1 to <10

Moderate

≥0.1 to <1

Strong

<0.1

Extreme

CAS

Vehicle

LLNA%

LLNA%

LLNA%

LLNA%

LLNA%

LLNA SI

LLNA SI

LLNA SI

LLNA SI

LLNA SI

LLNA EC3

Relative Potency

test chemical

AOO

1.0

2.5

5.0

10.0

20.0

1.0

1.1

1.6

1.4

0.9

NC

Nonsensitizer

Where AOO – Acetic acid in olive oil ; SI - Stimulation Index; NC-Not Calculated

 

Interpretation of results:
other: not sensitizing
Conclusions:
The test chemical was considered to be not sensitizing to the skin on the basis of summarized studies.
Executive summary:

Data available for the structurally and functionally similar read across chemicals has been reviewed to determine the skin sensitization potential of the test chemical. The studies are as mentioned below:

The skin sensitizing potential of the test chemical was assessed by using the Mouse Local Lymphnode Assay. In this study, groups of mice (n=4) were exposed topically on the dorsum of both ears to 25 l of various concentrations of the test chemicals, or to the same volume of the relevant vehicle alone, daily for 3 consecutive days. Five days following initiation of exposure all mice were injected via the tail vein with 250 l of phosphate buffered saline (PBS) containing 20 Ci of [3H] methyl thymidine. Five hours later the mice were sacrificed and the draining auricular lymph nodes excised and pooled for each experimental group. Single cell suspensions of LNC were prepared by mechanical disaggregation through 200-mesh stainless steel gauze. Pooled LNC were washed twice with PBS and precipitated in 5% trichloroacetic acid (TCA) at 4°C overnight. Pellets were then resuspended in 1 ml of TCA and transferred to 10 ml of scintillation fluid (Optiphase ‘Hisafe 3’, Wallac, Turku, Finland). The incorporation of 3H-TdR was measured by -scintillation counting as disintegrations per minute (dpm) per node for each experimental group. In each case a stimulation index (SI) relative to the concurrent vehicle-treated control was derived; an SI of 3 or greater being indicative of a chemical possessing the potential to cause contact sensitization. The stimulation indices (SI) at concentration 0, 25, 50 and 75% was observed to be 1, 1.31, 1.50 and 1.46 respectively. The test chemical failed at all test concentrations (maximum concentrations of 75%) to induce a positive response in the local lymph node assay with relatively low levels of thymidine incorporation being recorded at all concentrations tested. Therefore the test chemical was considered to be not sensitizing in LLNA study.

Another Mouse Local Lymphnode Assay was conducted to assess the skin sensitization potential of similar read across chemical. The LLNA was conducted on groups of five female CBA mice (7-12 weeks of age) by mean of topical application of chemical on the dorsum of both ears at a dose of 25µl of 1%, 2.5%, 5%, 10% or 20% in acetone/olive oil (4:1). Treatment was performed daily for 3 consecutive days. Five days after initiation of exposure all mice were injected via the tail vein with 250µl of PBS containing 20µCi of tritiatied thymidine. The mice were sacrificed 5 hours later, and draining the auricular lymph nodes were excised and pooled for each experimental group or each individual animal. The incorporation of tritiated thymidine measured by β-scintillation counting and was reported in disintegrations /minute. An SI was calculated for each chemical group as the ratio of disintegrations/minute of the treated group to the disintegrations/minute of the concurrent vehicle control group. A substance was classified skin sensitizer, if at one or more than one concentrations, it induced a three-fold or greater increase in local lymph node proliferative activity when treated with the concurrent vehicle treated controls (SI ≥3).The approach to estimation of the relative skin sensitization potential is based on the mathematical estimation of the concentration of chemical necessary to obtain a threshold positive response (SI = 3); this is termed as the EC3 value. For each concentration of test chemical, a stimulation index (SI) relative to the concurrent vehicle-treated control was calculated. The calculated EC3 value was >20.0%. Thus based on the relative potency index the test chemical was considered to be not sensitizing in the Mouse Local Lymph node Assay.

 

The above results were further supported by the skin sensitization study performed for another similar read across chemical in 25 volunteers via maximization test. About 10% in petrolatum of test sample was administrated on different panels of human subject. No known cutaneous reaction was observed in 25 volunteers. Therefore, the test chemical can be considered as not sensitizing on human skin.

 

Based on the above summarized studies for target chemical and its structurally and functionally similar read across substances,it can be concluded that the testchemical is unable to cause skin sensitization and considered as non-skin sensitizer. Comparing the above annotations with the criteria of CLP regulation, it can be classified under the category “Not Classified”.

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

Data available for the structurally and functionally similar read across chemicals has been reviewed to determine the skin sensitization potential of the test chemical. The studies are as mentioned below:

The skin sensitizing potential of the test chemical was assessed by using the Mouse Local Lymphnode Assay. In this study, groups of mice (n=4) were exposed topically on the dorsum of both ears to 25 l of various concentrations of the test chemicals, or to the same volume of the relevant vehicle alone, daily for 3 consecutive days. Five days following initiation of exposure all mice were injected via the tail vein with 250 l of phosphate buffered saline (PBS) containing 20 Ci of [3H] methyl thymidine. Five hours later the mice were sacrificed and the draining auricular lymph nodes excised and pooled for each experimental group. Single cell suspensions of LNC were prepared by mechanical disaggregation through 200-mesh stainless steel gauze. Pooled LNC were washed twice with PBS and precipitated in 5% trichloroacetic acid (TCA) at 4°C overnight. Pellets were then resuspended in 1 ml of TCA and transferred to 10 ml of scintillation fluid (Optiphase ‘Hisafe 3’, Wallac, Turku, Finland). The incorporation of 3H-TdR was measured by -scintillation counting as disintegrations per minute (dpm) per node for each experimental group. In each case a stimulation index (SI) relative to the concurrent vehicle-treated control was derived; an SI of 3 or greater being indicative of a chemical possessing the potential to cause contact sensitization. The stimulation indices (SI) at concentration 0, 25, 50 and 75% was observed to be 1, 1.31, 1.50 and 1.46 respectively. The test chemical failed at all test concentrations (maximum concentrations of 75%) to induce a positive response in the local lymph node assay with relatively low levels of thymidine incorporation being recorded at all concentrations tested. Therefore the test chemical was considered to be not sensitizing in LLNA study.

Another Mouse Local Lymphnode Assay was conducted to assess the skin sensitization potential of similar read across chemical. The LLNA was conducted on groups of five female CBA mice (7-12 weeks of age) by mean of topical application of chemical on the dorsum of both ears at a dose of 25µl of 1%, 2.5%, 5%, 10% or 20% in acetone/olive oil (4:1). Treatment was performed daily for 3 consecutive days. Five days after initiation of exposure all mice were injected via the tail vein with 250µl of PBS containing 20µCi of tritiatied thymidine. The mice were sacrificed 5 hours later, and draining the auricular lymph nodes were excised and pooled for each experimental group or each individual animal. The incorporation of tritiated thymidine measured by β-scintillation counting and was reported in disintegrations /minute. An SI was calculated for each chemical group as the ratio of disintegrations/minute of the treated group to the disintegrations/minute of the concurrent vehicle control group. A substance was classified skin sensitizer, if at one or more than one concentrations, it induced a three-fold or greater increase in local lymph node proliferative activity when treated with the concurrent vehicle treated controls (SI ≥3).The approach to estimation of the relative skin sensitization potential is based on the mathematical estimation of the concentration of chemical necessary to obtain a threshold positive response (SI = 3); this is termed as the EC3 value. For each concentration of test chemical, a stimulation index (SI) relative to the concurrent vehicle-treated control was calculated. The calculated EC3 value was >20.0%. Thus based on the relative potency index the test chemical was considered to be not sensitizing in the Mouse Local Lymph node Assay.

 

The above results were further supported by the skin sensitization study performed for another similar read across chemical in 25 volunteers via maximization test. About 10% in petrolatum of test sample was administrated on different panels of human subject. No known cutaneous reaction was observed in 25 volunteers. Therefore, the test chemical can be considered as not sensitizing on human skin.

 

Based on the above summarized studies for target chemical and its structurally and functionally similar read across substances,it can be concluded that the testchemical is unable to cause skin sensitization and considered as non-skin sensitizer. Comparing the above annotations with the criteria of CLP regulation, it can be classified under the category “Not Classified”.

Respiratory sensitisation

Endpoint conclusion
Endpoint conclusion:
no study available

Justification for classification or non-classification

The skin sensitization potential of test substance andits structurally and functionally similar read across substanceswere observed in various studies. From the results obtained from these studies it is concluded that the chemical is not likely to cause skin sensitization and hence can be classified as non-skin sensitizer.