Registration Dossier

Administrative data

Endpoint:
skin sensitisation: in vivo (LLNA)
Type of information:
experimental study
Adequacy of study:
key study
Study period:
July 24, 2013 - August 12, 2013
Reliability:
1 (reliable without restriction)
Rationale for reliability incl. deficiencies:
other: This study has been performed according to OECD and/or EC guidelines and according to GLP principles.

Data source

Reference
Reference Type:
study report
Title:
Unnamed
Year:
2013
Report Date:
2013

Materials and methods

Test guidelineopen allclose all
Qualifier:
according to
Guideline:
OECD Guideline 429 (Skin Sensitisation: Local Lymph Node Assay)
Version / remarks:
(2010)
Deviations:
no
Qualifier:
according to
Guideline:
EU Method B.42 (Skin Sensitisation: Local Lymph Node Assay)
Version / remarks:
(2008)
Deviations:
no
Qualifier:
according to
Guideline:
EPA OPPTS 870.2600 (Skin Sensitisation)
Version / remarks:
(2003)
Deviations:
no
GLP compliance:
yes
Type of study:
mouse local lymph node assay (LLNA)

Test material

Reference
Name:
Unnamed
Type:
Constituent
Test material form:
other: liquid
Details on test material:
- Name of test material (as cited in study report): KF-1622
- Description: Clear colourless liquid

In vivo test system

Test animals

Species:
mouse
Strain:
other: CBA/J
Sex:
female
Details on test animals and environmental conditions:
TEST ANIMALS
- Source: Janvier, Le Genest-Saint-Isle, France
- Age at study initiation: Young adult animals (approx. 11 weeks old)
- Weight at study initiation: 21 - 23 g
- Housing: Animals were group housed in labeld makrolon cages.
- Diet: Free access to pelleted rodent diet (SM R/M-Z from SSNIFF® Spezialdiäten GmbH, Soest, Germany).
- Water: Free access to tap water.
- Acclimation period: At least 5 days

ENVIRONMENTAL CONDITIONS
- Temperature (°C): 18 – 24
- Humidity (%): 40 - 70
- Air changes (per hr): approx 15
- Photoperiod (hrs dark / hrs light): 12/12

Study design: in vivo (LLNA)

Vehicle:
acetone/olive oil (4:1 v/v)
Concentration:
0, 25, 50 and 100%
No. of animals per dose:
5
Details on study design:
RANGE FINDING TESTS:
Two test substance concentrations were tested; a 100% and 50% concentration.
The test system, procedures and techniques were identical to those used in the main study except that the assessment of lymph node proliferation and necropsy were not performed. Two young adult animals per concentration were selected (in the range of 8 to14 weeks old). Each animal was treated
with one concentration on three consecutive days. Animals were group housed in labeled Makrolon cages (MII type, height 14 cm). Ear thickness measurements were conducted using a digital thickness gauge prior to dosing on Days 1 and 3, and on Day 6.
Animals were sacrificed after the final observation.

MAIN STUDY
ANIMAL ASSIGNMENT AND TREATMENT
- Name of test method: Local Lymph Node Assay
- Criteria used to consider a positive response: DPM values are presented for each animal and for each dose group. A Stimulation Index (SI) is calculated for each group. The SI is the ratio of the DPM/group compared to DPM/vehicle control group. If the results indicate a SI ≥ 3, the test substance may be regarded as a skin sensitizer. Consideration was given to the EC3 value (the estimated test substance concentration that will give a SI =3).

ANIMAL ASSIGNMENT
Three groups of five animals were treated with one test substance concentration per group. One group of five animals was treated with vehicle.

TREATMENT PREPARATION AND ADMINISTRATION:
Test substance preparation: The test substance formulations (w/w) were prepared within 4 hours prior to each dosing. Homogeneity was obtained to visually acceptable levels.
Rationale for vehicle: The vehicle was selected based on trial formulations performed at WIL Research Europe and on test substance data supplied by the sponsor.

Induction - Days 1, 2 and 3; Excision of nodes - Day 6; Tissue processing for radioacitivity - Day 6; Radioactivity measurements - Day 7; Performed according to test guideline OECD 429.

Observations:
Mortality/Viability: Twice daily.
Body weights: On Day 1 (pre-dose) and Day 6 (prior to necropsy).
Clinical signs: Once daily on Days 1-6 (on Days 1-3 between 3 and 4 hours after dosing).
Irritation: Once daily on Days 1-6 (on Days 1 - 3 immediately after dosing) according to the numerical scoring system according to OECD 429. Furthermore, a description of all other (local) effects was recorded according to guidelines.

Necropsy: No necropsy was performed according to protocol.
Positive control substance(s):
hexyl cinnamic aldehyde (CAS No 101-86-0)
Statistics:
Not performed.

Results and discussion

Positive control results:
The six-month reliability check with Alpha-hexylcinnamicaldehyde indicates that the Local Lymph Node Assay as performed at WIL Research is an appropriate model for testing for contact hypersensitivity. See attached document 'Reliability check'.

In vivo (LLNA)

Resultsopen allclose all
Parameter:
SI
Remarks on result:
other: The SI values calculated for the substance concentrations 25, 50 and 100% were 1.1, 1.8 and 1.4, respectively.
Parameter:
other: disintegrations per minute (DPM)
Remarks on result:
other: Mean DPM/animal values for the experimental groups treated with test substance concentrations 25, 50 and 100% were 435, 672 and 517 DPM, respectively. The mean DPM/animal value for the vehicle control group was 381 DPM.

Any other information on results incl. tables

Results Pre-screen test:

No irritation and no signs of systemic toxicity were observed in any of the animals examined.

Based on these results, the highest test substance concentration selected for the main study was a 100% concentration.

Other results - main study:

No irritation of the ears was observed in any of the animals examined.

 

The majority of auricular lymph nodes were considered normal in size, except for the nodes of some animals dosed at 25% and all animals dosed at 50%, which were enlarged. No macroscopic abnormalities of the surrounding area were noted in any of the animals.

 

No mortality occurred and no clinical signs of systemic toxicity were observed in the animals of the main study. Body weights and body weight gain of experimental animals remained in the same range as controls over the study period.

Applicant's summary and conclusion

Interpretation of results:
not sensitising
Remarks:
Migrated information Criteria used for interpretation of results: EU
Conclusions:
In an LLNA skin sensitisation study, performed according to OECD 429 test guidelines and GLP principles, the substance was considered not to be a skin sensitiser, as the SI appeared not to be ≥ 3 when tested up to 100%.
Executive summary:

KF-1622 was assessed for contact hypersensitivity in a LLNA skin sensitisation study, performed according to OECD 429 test guideline and GLP principles up to and including 100%.

No irritation of the ears was observed in any of the animals examined. The majority of auricular lymph nodes were considered normal in size, except for the nodes of some animals dosed at 25% and all animals dosed at 50%, which were enlarged. No macroscopic abnormalities of the surrounding area were noted in any of the animals. No mortality occurred and no clinical signs of systemic toxicity were observed in the animals of the main study. Body weights and body weight gain of experimental animals remained in the same range as controls over the study period.

The SI values calculated for the substance concentrations 25, 50 and 100% were 1.1, 1.8 and 1.4, respectively. The substance was considered not to be a skin sensitiser, as the SI appeared not to be ≥ 3 when tested up to and including 100%.

Based on the results, KF-1622 does not have to be classified as skin sensitizer according to Regulation (EC) No 1272/2008.