Registration Dossier

Data platform availability banner - registered substances factsheets

Please be aware that this old REACH registration data factsheet is no longer maintained; it remains frozen as of 19th May 2023.

The new ECHA CHEM database has been released by ECHA, and it now contains all REACH registration data. There are more details on the transition of ECHA's published data to ECHA CHEM here.

Diss Factsheets

Toxicological information

Skin sensitisation

Currently viewing:

Administrative data

Endpoint:
skin sensitisation: in vitro
Type of information:
experimental study
Adequacy of study:
key study
Study period:
From May 18th to July 09th, 2021
Reliability:
1 (reliable without restriction)
Rationale for reliability incl. deficiencies:
test procedure in accordance with generally accepted scientific standards and described in sufficient detail

Data source

Reference
Reference Type:
study report
Title:
Unnamed
Year:
2021
Report date:
2021

Materials and methods

Principles of method if other than guideline:
Method described in the following papers:
1. Genes specifically modulated in sensitized skins allow the detection of sensitizers in a reconstructed human skin model. Development of the SENS-IS assay. Cottrez F., Boitel E., Auriault C., Aeby P., Groux H. Toxicology in vitro 29: 787-802, 2015.
2. SENS-IS, a 3D reconstituted epidermis based model for quantifying chemical sensitization potency: reproductibility and predictivity results from an inter-laboratory study. Cottrez F., Boitel E., Ourlin J.C., Peiffer J.L., Fabre I., Henaoui I.S., Mari B., Vallauri A., Paquet A., Barbry P., Auriault C., Aeby P., Groux H. . Toxicology in Vitro 32: 248-260, 2016.
GLP compliance:
no
Remarks:
not fully validated method
Type of study:
other: activation of ARE and SENS-IS gene subset
Justification for non-LLNA method:
LLNA method was not carried out because the test substance is a complex metal

Test material

Constituent 1
Test material form:
solid

In vitro test system

Details on the study design:
Preliminary test: Solubility test
The solubility of the test item was assessed in phosphate buffered saline (PBS), olive oil (OO), and dimethylsulfoxide (DMSO) at a concentration of 10% and 50%, at room temperature and at 37°C.
The solubility of the test item was assessed by visual inspection of each preparation.
Main test: SENS-IS assay
The test item (30 µL) was deposited on the epidermis surface and gently spread on the entire surface [3]. After 15 minutes of exposure, the Episkin™ was rinsed with PBS and then incubated at 37°C for 6 hours.
After incubation, reconstructed epidermis was removed from the inserts with forceps and placed in a cryotube for freezing in liquid nitrogen. The epidermis was then transferred in a tube containing 1 mL of Qiazol reagent and 2 steel beads. Epidermis was homogenized using the TissueLyser II. After centrifugation, the supernatant was collected and stored at -20°C until RNA extraction.
After addition of bromochloropropane, total RNA were purified using the miRNeasy extraction Kit according to the manufacturer’s instructions (Qiagen, Courtaboeuf, France). RNA quality was assessed by measuring 260/280 absorbance ratio. The mRNA was then reversed as cDNA using SuperScript III Reverse Transcriptase kit and RNase inhibitor.
After reverse transcription, quantitative gene expression was measured by qRT-PCR using a SYBRGreen® buffer and specific primers defined for the SENS-IS test. The housekeeping genes (Glucuronidase ß, ß2 microglobuline, and Nono « non-POU domain containing octamer-binding ») were analyzed in parallel.
Vehicle / solvent control:
DMSO
Negative control:
other: DMSO
Positive control:
other: SLS, 10%= Positive control for irritation and negative control for sensitization; TNBS, 1M= Positive control for sensitization

Results and discussion

Positive control results:
SLS at 5% was classified as irritant (number of overexpressed irritant genes > 15) and non-sensitizer, the number of overexpressed genes in both the SENS-IS and ARE groups being below 7.
TNBS at 1M was classified as sensitizer since more than 6 genes are overexpressed in at least one of the two groups of genes (SENS-IS or ARE).

In vitro / in chemico

Resultsopen allclose all
Group:
test chemical
Run / experiment:
run/experiment 2
Parameter:
other: Number of overexpressed SENS-IS and ARE genes
Value:
2
Vehicle controls validity:
valid
Negative controls validity:
valid
Positive controls validity:
valid
Remarks on result:
other: Vehicle: 50% DMSO
Group:
test chemical
Run / experiment:
run/experiment 2
Parameter:
other: Number of overexpressed SENS-IS and ARE genes
Value:
1
Vehicle controls validity:
valid
Negative controls validity:
valid
Positive controls validity:
valid
Remarks on result:
other: Vehicle: 100% DMSO
Group:
test chemical
Run / experiment:
run/experiment 1
Parameter:
other: Number of overexpressed SENS-IS and ARE genes
Value:
2
Vehicle controls validity:
valid
Negative controls validity:
valid
Positive controls validity:
valid
Remarks on result:
other: Vehicle: 10% PBS
Group:
test chemical
Run / experiment:
run/experiment 1
Parameter:
other: Number of overexpressed SENS-IS and ARE genes
Value:
2
Vehicle controls validity:
valid
Negative controls validity:
valid
Positive controls validity:
valid
Remarks on result:
other: Vehicle: 1% DMSO
Group:
test chemical
Run / experiment:
run/experiment 1
Parameter:
other: Number of overexpressed SENS-IS and ARE genes
Value:
2
Vehicle controls validity:
valid
Negative controls validity:
valid
Positive controls validity:
valid
Remarks on result:
other: Vehicle: 10% DMSO
Group:
test chemical
Run / experiment:
run/experiment 1
Parameter:
other: Number of overexpressed SENS-IS and ARE genes
Value:
3
Vehicle controls validity:
valid
Negative controls validity:
valid
Positive controls validity:
valid
Remarks on result:
other: Vehicle: 50% DMSO

Any other information on results incl. tables

The test item was soluble at 10 and 50% (w/v) in PBS and in DMSO. It was not soluble at 10 and 50% (w/v) in olive oil.

Applicant's summary and conclusion

Interpretation of results:
other: No skin sensitising according to the classification criteria described in the SENS-IS assay
Conclusions:
not skin sensitising
Executive summary:

Method: the substance has been tested for its potential to induce skin sensitisation according to the SENS-IS assay (Cottrez F. et al, Toxicology in vitro 29 (2015) 787 -802).


The objective of this study was to evaluate the capacity of the test item to induce the expression of specific irritation and sensitization biomarkers in a 3D-reconstructed epidermis model.


The results obtained for the positive and negative controls were within acceptance criteria defined in the Study Plan.


Considering the number of over-expressed genes in the “SENS-IS” and “ARE” gene groups, the test item gave negative result (less than 7 genes induced) when it was tested at 1, 10, 50 (w/v) in DMSO and at 10% (w/v) in PBS. Moreover, negative results were obtained when the test item was incubated at 100% (w/v) in DMSO.


In conclusion, under the experimental conditions of this SENS-IS assay, the test item can be classified as a non-sensitizer.


It should be mentioned that the SENS-IS method correlate more than 92% with LLNA in potency class prediction and should demonstrated a higher performance for predicting sensitization when combined with other sensitization tests (in chemico, in vitro and in silico).