Registration Dossier

Diss Factsheets

Toxicological information

Skin sensitisation

Currently viewing:

Administrative data

Endpoint:
skin sensitisation: in vivo (LLNA)
Type of information:
experimental study
Adequacy of study:
key study
Study period:
From 2019-05-14 to 2019-06-04
Reliability:
1 (reliable without restriction)
Rationale for reliability incl. deficiencies:
guideline study

Data source

Reference
Reference Type:
study report
Title:
Unnamed
Year:
2019
Report date:
2019

Materials and methods

Test guideline
Qualifier:
according to guideline
Guideline:
OECD Guideline 429 (Skin Sensitisation: Local Lymph Node Assay)
Version / remarks:
2010
Deviations:
no
GLP compliance:
yes (incl. QA statement)
Type of study:
mouse local lymph node assay (LLNA)

Test material

Constituent 1
Chemical structure
Reference substance name:
Tetravinylsilane
EC Number:
214-192-0
EC Name:
Tetravinylsilane
Cas Number:
1112-55-6
Molecular formula:
C8H12Si
IUPAC Name:
tetravinylsilane
Test material form:
liquid
Specific details on test material used for the study:
Batch No.: 707001
Purity: 98.5%

In vivo test system

Test animals

Species:
mouse
Strain:
CBA:J
Sex:
female
Details on test animals and environmental conditions:
TEST ANIMALS
- Source: Janvier, Le Genest-Saint-Isle, France
- Females nulliparous and non-pregnant: yes
- Microbiological status of animals, when known: Inbred, SPF-Quality
- Age at study initiation: approximately 10 weeks old
- Weight at study initiation: 19.8 to 24.0 g.
- Housing: group housed (up to 5 animals of the same sex and same dosing group together) in polycarbonate cages (Makrolon MIII type; height 18 cm.) containing sterilized sawdust as bedding material (Lignocel S 8-15, JRS - J.Rettenmaier & Söhne GmbH + CO. KG, Rosenberg, Germany) equipped with water bottles. Animals were separated during designated procedures/activities
- Diet: Pelleted rodent diet, ad libitum
- Water: Municipal tap-water, ad libitum
- Acclimation period: at least 5 days


ENVIRONMENTAL CONDITIONS
- Temperature (°C): 22°C
- Humidity (%): 42 to 56%.
- Air changes (per hr): Ten or greater
- Photoperiod: 12-hour light/12-hour dark cycle

Study design: in vivo (LLNA)

Vehicle:
acetone/olive oil (4:1 v/v)
Concentration:
25, 50, 100 %w/w
No. of animals per dose:
5
Details on study design:
PRE-SCREEN TESTS:
At both a 50% and 100% test item concentration, no signs of systemic toxicity were noted and very slight erythema and/or scaliness were observed between Days 1 and 3. Therefore the 100% concentration was selected as highest concentration for the main study.

MAIN STUDY
Three groups of five animals were treated with one test item concentration per group. The highest test item concentration was selected from the pre-screen test. One group of five animals was treated with the vehicle.

ANIMAL ASSIGNMENT AND TREATMENT
- Name of test method: Animals were assigned to the study at the discretion of the coordinating bio technician according to body weights, with all animals within ± 20% of the sex mean. Animals in poor health or at extremes of body weight range were not assigned to the study.
- Criteria used to consider a positive response: SI ≥ 3

TREATMENT PREPARATION AND ADMINISTRATION:
- Preparation of Test Item:
Test item dosing formulations (w/w) were homogenized to visually acceptable levels at appropriate concentrations to meet dose level requirements. The dosing formulations were prepared daily and dosed within 4 hours after adding the vehicle to the test item.
- Induction - Days 1, 2 and 3:
The dorsal surface of both ears was topically treated (25 μL/ear) with the test item, at approximately the same time on each day. The control animals were treated in the same way as the experimental animals, except that the vehicle was administered instead of the test item.
- Excision of the Nodes - Day 6:
Each animal was injected via the tail vein with 0.25 mL of sterile phosphate buffered saline (PBS) containing 20 μCi of 3H-methyl thymidine.
After five hours, all animals were euthanized by intraperitoneal injection (0.2 mL/animal) of Euthasol® 20%. The draining (auricular) lymph node of each ear was excised. The relative size of the nodes (as compared to normal) was estimated by visual examination and abnormalities of the nodes and surrounding area were recorded. The nodes were pooled for each animal in PBS.
- Tissue Processing for Radioactivity - Day 6:
Following excision of the nodes, a single cell suspension of lymph node cells (LNC) was prepared in PBS by gentle separation through stainless steel gauze (maze size: 200 μm, diameter: ± 1.5 cm). LNC were washed twice with an excess of PBS by centrifugation at 200g for 10 minutes at 4ºC. To precipitate the DNA, the LNC were exposed to 5% trichloroacetic acid (TCA) and then stored in the refrigerator until the next day.
- Radioactivity Measurements - Day 7:
Precipitates were recovered by centrifugation, resuspended in 1 mL TCA and transferred to 10 mL of Ultima Gold cocktail as the scintillation fluid. Radioactivity measurements were performed using a Packard scintillation counter (2910TR). Counting time was to a statistical precision of ± 0.2% or a maximum of 5 minutes whichever came first. The scintillation counter was programmed to automatically subtract background and convert Counts Per Minute (CPM) to Disintegrations Per Minute (DPM)

OBSERVATIONS
- Mortality/Moribundity Checks: twice daily, in the morning and at the end of the working day.
- Clinical Observations: once daily on Days 1-6 (on Days 1-3 between 3 and 4 hours after dosing).
- Body Weights: weighed individually on Day 1 (predose) and 6 (prior to necropsy).
- Irritation: once daily on Days 1-6 (on Days 1-3 within 1 hour after dosing),


Positive control substance(s):
hexyl cinnamic aldehyde (CAS No 101-86-0)

Results and discussion

Positive control results:
An EC3 value of 16.3% was calculated using linear interpolation.
The calculated EC3 value was found to be in the acceptable range of 4.8 and 19.5%. The results of the 6 monthly HCA reliability checks of the recent years were 13.4, 14.1, 17.3, 9.8, 17.8 and 14.3%.

In vivo (LLNA)

Resultsopen allclose all
Parameter:
SI
Value:
0.7
Test group / Remarks:
25% w/w group
Parameter:
SI
Value:
1.1
Test group / Remarks:
50 % w/w group
Parameter:
SI
Value:
2
Test group / Remarks:
100 % w/w group
Cellular proliferation data / Observations:
- Skin Reactions / Irritation:
The very slight erythema and/or scaliness noted for the test item treated animals between Days 1 and 4 was considered not to have a toxicologically significant effect on the activity of the nodes.
- Systemic Toxicity:
No mortality occurred and no clinical signs of systemic toxicity were observed in the animals.
Body weights and body weight gain of experimental animals remained in the same range as controls over the study period.
- Macroscopic Examination of the Lymph Nodes and Surrounding Area:
The majority of auricular lymph nodes were considered normal in size, except for the nodes in two animals treated at 100% which were considered to be slightly enlarged.
No macroscopic abnormalities of the surrounding area were noted for any of the animals.
- Radioactivity Measurements and SI Values:
Mean DPM/animal values for the experimental groups treated with test item concentrations 25, 50 and 100% were 622, 937 and 1692 DPM, respectively. The mean DPM/animal value for the vehicle control group was 835 DPM.

Applicant's summary and conclusion

Conclusions:
The SI values calculated for the test item concentrations 25, 50 and 100% were 0.7, 1.1 and 2.0, respectively. Since there was no indication that the test item elicited a SI ≥ 3 when tested up to 100%, the test item was considered not to be a skin sensitizer.
Executive summary:

The objective of this study was to evaluate whether the test item induces skin sensitization in mice after three epidermal exposures of the animals based on OECD 429.

Three experimental groups of five female CBA/J mice were treated with test item concentrations of 25, 50 or 100% w/w on three consecutive days, by open application on the ears. Five vehicle control animals were similarly treated, but with the vehicle alone (Acetone/Olive oil (4:1 v/v) (AcOO)).

The majority of auricular lymph nodes were considered normal in size, except for the nodes in two animals treated at 100% which were considered to be slightly enlarged. No macroscopic abnormalities of the surrounding area were noted for any of the animals.

Mean DPM/animal values for the experimental groups treated with test item concentrations 25, 50 and 100% were 622, 937 and 1692 DPM, respectively. The mean DPM/animal value for the vehicle control group was 835 DPM. The SI values calculated for the test item concentrations 25, 50 and 100% were 0.7, 1.1 and 2.0, respectively.

Since there was no indication that the test item elicited a SI3 when tested up to 100%, the test item was considered not to be a skin sensitizer.