Registration Dossier

Administrative data

Endpoint:
skin sensitisation: in vivo (non-LLNA)
Type of information:
migrated information: read-across from supporting substance (structural analogue or surrogate)
Adequacy of study:
key study
Study period:
2008
Reliability:
1 (reliable without restriction)
Rationale for reliability incl. deficiencies:
other: Study conducted in compliance with agreed protocols, with no or minor deviations from standard test guidelines and/or minor methodological deficiencies, which do not affect the quality of the relevant results.

Data source

Reference
Reference Type:
study report
Title:
Unnamed
Year:
2008
Report Date:
2008

Materials and methods

Test guidelineopen allclose all
Qualifier:
according to
Guideline:
OECD Guideline 406 (Skin Sensitisation)
Deviations:
no
Qualifier:
according to
Guideline:
EPA OPPTS 870.2600 (Skin Sensitisation)
Deviations:
no
GLP compliance:
yes
Type of study:
Buehler test

Test material

Reference
Name:
Unnamed
Type:
Constituent
Details on test material:
Sponsor's identification: AS305BD
Description: dark brown, viscous liquid
Lot number: TS07002
Analytical purity: 100%
Storage conditions: room temperature in the dark

In vivo test system

Test animals

Species:
guinea pig
Strain:
Hartley
Sex:
male/female
Details on test animals and environmental conditions:
TEST ANIMALS
- Source: Charles River Laboratories, Saint Constant, Quebec
- Age at study initiation: approx 6 weeks (males); approx 7 weeks (females)
- Weight at study initiation: 379-449 g (males); 351-411 g (females)
- Housing: suspended stainless steel cages
- Diet (e.g. ad libitum): PMI Certified Guinea Pg Chow #5026 (PMI Nutrition International) ad libitum
- Water (e.g. ad libitum): municipal tap water ad libitum
- Acclimation period: minimum of five days

ENVIRONMENTAL CONDITIONS
- Temperature (°C): 21-22 ºC
- Humidity (%): 31-73%
- Air changes (per hr): 10-15 per hour
- Photoperiod (hrs dark / hrs light): 12 hrs light/12 hrs dark

Study design: in vivo (non-LLNA)

Inductionopen allclose all
Route:
epicutaneous, semiocclusive
Vehicle:
other: mineral oil, USP
Concentration / amount:
10% (Induction 1) and 5% (Inductions 2 and 3) w/w AS305BD in mineral oil, USP, once per week, for three consecutive weeks.
Challengeopen allclose all
Route:
epicutaneous, semiocclusive
Vehicle:
other: mineral oil, USP
Concentration / amount:
10% (Induction 1) and 5% (Inductions 2 and 3) w/w AS305BD in mineral oil, USP, once per week, for three consecutive weeks.
No. of animals per dose:
Ten male and ten female
Details on study design:
PRELIMINARY PROCEDURES
On the day prior to each dose administration, the guinea pigs had the hair removed with a small animal clipper with a No. 40 blade followed by further close clipping with a Braun Syncro Electric Razor (Model 7505). Care was taken to avoid abrading the skin.

DOSING
For the induction, challenge. and rechallenge phases, a dose of 0.3 mL was placed on a 25-mm Hilltop chamber backed by adhesive tape (occlusive patch). The chambers were applied to the clipped surface as quickly as possible. Following chamber application, the trunk of tile animal was wrapped with elastic wrap which was secured with adhesive tape to prevent removal of the chamber and the animal was returned to its cage.

Induction
On the day prior to the first induction dose administration (day -1), all test, challenge control, and rechallenge control animals were weighed and the hair was removed from the left side of the test animals. On the day following clipping (day 0), chambers were applied as follows:

Induction No. Concentration (%) Test site no. No. of animals (male) No. of animals (female)
1 10 1 10 10
2 5 3
3 5 5

The concentration was reduced for inductions 2 and 3 due to significant irritation at induction 1.

The induction procedure was repeated on study day 6 and on study day 13 so that a total of three consecutive induction exposures were made to the test animals.

Challenge
On the day prior to challenge dose administration, the test and challenge control animals were weighed and the hair was removed from the right side of the animals. On the day following clipping (day 26), chambers were applied as follows:

Group Concentration (%) Test site no. No. of animals (male) No. of animals (female)
Test 2.5 2 10 10
Challenge control 2.5 2 5 5

Rechallenge
On the day prior to rechallenge dose administration, the test and rechallenge control animals were weighed and the hair was removed from the right side of the animals. On the day following clipping (day 40), chambers were applied as follows:

Group Concentration (%) Test site no. No. of animals (male) No. of animals (female)
Test 2.5 6 10 10
Test 1 4 10 10
Rechallenge control 2.5 6 5 5
Rechallenge control 1 4 5 5

TEST ARTICLE REMOVAL
Approximately six hours after chamber application, the binding materials were removed. The test sites were wiped with gauze moistened in mineral oil, USP, followed by dry gauze followed by gauze moistened in deionized water, followed by dry gauze, to remove test article residue. The animals were then returned to their cages.

DERMAL OBSERVATIONS
The test sites were graded for irritation at approximately 24 and 48 hours following chamber application (Induction) or chamber removal (Challenge and Rechallenge) using the Dermal Grading System presented in Appendix 3.

CLINICAL OBSERVATIONS
Any unusual observations and mortality were recordcd. The animals were observed for general health/mortality twice daily, once in the morning and once in the afternoon.

BODY WEIGHTS
Individual body weights were obtained for all test, challenge control, and rechallenge control animals on the day prior to the first induction (day -1), and for the appropriate test and challenge control animals on the day prior to challenge dosing. The appropriate test and rechallenge control animals wcre weighed on the day prior to rechallenge dosing.

SCHEDULED EUTHANASIA
All sensitization study animals were euthanized by carbon dioxide inhalation following each animal's final scoring interval. Gross necropsy examinations were not required for these animals.
Challenge controls:
See above.
Positive control substance(s):
yes
Remarks:
α-hexylcinnamaldehyde

Study design: in vivo (LLNA)

Concentration:
Not applicable
No. of animals per dose:
Not applicable
Details on study design:
Not applicable
Statistics:
Criteria for determining if a skin reaction in a guinea pig was indicative of skin sensitization under conditions where there was no confounding irritation in the concurrent irritation control animals were as follows:

-The skin reaction score was greater than a ± in the induced animals.
-The skin reaction scores in the concurrent irritation control animals were not greater than a ± (if skin reaction scores in the concurrent irritation control animals were greater than a ±, then a rechallenge at a lower concentration was required and different criteria may apply).
-Special consideration was given to the skin reaction score for individual animals that persist at both the 24·hour and 48-hour observations in the induced animals.
-Skin reactions indicative of skin sensitization in individual animals were present at both challenge and rechallenge.

Criteria for determining if a test material is classifiable as a skin sensitizer:
-15% of the induced animals had skin reactions indicative of skin sensitization at challenge and rechallenge, and there was no confounding irritation present in the concurrent irritation control animals.

Results and discussion

Positive control results:
The results of the HCA historical control study demonstrated that a valid test was performed and indicated that the test design would detect
potential contact sensitizers.

In vivo (non-LLNA)

Resultsopen allclose all
Reading:
1st reading
Hours after challenge:
24
Group:
test group
Dose level:
2.5% w/w
No. with + reactions:
19
Total no. in group:
20
Remarks on result:
other: Reading: 1st reading. . Hours after challenge: 24.0. Group: test group. Dose level: 2.5% w/w. No with. + reactions: 19.0. Total no. in groups: 20.0.
Reading:
1st reading
Hours after challenge:
48
Group:
test group
Dose level:
2.5% w/w
No. with + reactions:
17
Total no. in group:
20
Remarks on result:
other: Reading: 1st reading. . Hours after challenge: 48.0. Group: test group. Dose level: 2.5% w/w. No with. + reactions: 17.0. Total no. in groups: 20.0.
Reading:
rechallenge
Hours after challenge:
24
Group:
test group
Dose level:
2.5% w/w
No. with + reactions:
13
Total no. in group:
20
Remarks on result:
other: Reading: rechallenge. . Hours after challenge: 24.0. Group: test group. Dose level: 2.5% w/w. No with. + reactions: 13.0. Total no. in groups: 20.0.
Reading:
rechallenge
Hours after challenge:
48
Group:
test group
Dose level:
2.5% w/w
No. with + reactions:
9
Total no. in group:
20
Remarks on result:
other: Reading: rechallenge. . Hours after challenge: 48.0. Group: test group. Dose level: 2.5% w/w. No with. + reactions: 9.0. Total no. in groups: 20.0.
Reading:
rechallenge
Hours after challenge:
24
Group:
test group
Dose level:
1.0% w/w
No. with + reactions:
14
Total no. in group:
20
Remarks on result:
other: Reading: rechallenge. . Hours after challenge: 24.0. Group: test group. Dose level: 1.0% w/w. No with. + reactions: 14.0. Total no. in groups: 20.0.
Reading:
rechallenge
Hours after challenge:
48
Group:
test group
Dose level:
1.0% w/w
No. with + reactions:
12
Total no. in group:
20
Remarks on result:
other: Reading: rechallenge. . Hours after challenge: 48.0. Group: test group. Dose level: 1.0% w/w. No with. + reactions: 12.0. Total no. in groups: 20.0.

In vivo (LLNA)

Resultsopen allclose all
Parameter:
SI
Remarks on result:
other: Not applicable.
Parameter:
other: disintegrations per minute (DPM)
Remarks on result:
other: Not applicable.

Any other information on results incl. tables

TOPICAL RANGE-FINDING STUDY

The results of the range-finding study indicated that a test material concentration of 10% w/w in mineral oil, USP was considered appropriate for induction as this concentration produced a mild to moderate response. Due to significant irritation observed at Induction 1, the test article concentration was reduced to 5% w/w in mineral oil, USP for Inductions 2 and 3. A concentration of 2.5% w/w in mineral oil, USP was selected for challenge as this concentration was the highest nonirritating concentration. For the rechallenge phase,

concentrations of 2.5% and 1.0% w/w in mineral oil, USP were selected to evaluate the response observed at challenge.

CLINICAL OBSERVATIONS/BODY WEIGHTS

The sensitization study animals gained weight during the test period and generally appeared in good health.

Applicant's summary and conclusion

Interpretation of results:
sensitising
Remarks:
Migrated information
Conclusions:
Under the conditions of this test, AS305BD does meet the criteria to be considered a contact sensitizer under the conditions tested.
Executive summary:

The dermal sensitization potential of AS305BD was evaluated in guinea pigs in a study following OCED Guideline 406. Ten male and ten female guinea pigs were topically treated with 10% (Induction 1) and 5% (Inductions 2 and 3) w/w AS305BD once per week, for three consecutive weeks. Following an approximate 2-week rest period, a challenge was performed whereby the 20 test and 10 previously untreated (naive) challenge control guinea pigs were topically treated with 2.5% w/w AS305BD. Challenge responses in the test animals were compared with those of the challenge control animals. Following a 2-week rest period, a rechallenge was preformed whereby the 20 test and 10 previollsly untreated (naive) rechallenge control guinea pigs were topically treated with 2.5% and 1% w/w AS30SBD. Rechallenge responses in the test animals were compared with those of the rechallenge control animals.

Following challenge with 2.5% w/w AS305BD, dermal scores of 1 were noted in 19/20 test animals at the 24-hour scoring interval and in 17/20 test animals at the 48-hour scoring interval. Dermal reactions in the remaining test and challenge control animals were limited to scores of 0 or ±. Group mean dermal scores were higher in the test animals than in the challenge control animals. Following rechallenge with 2.5% w/w AS305BD, dermal scores of 1 or 2 were noted in 13/20 test animals at the 24-hour scoring interval and dermal scores of 1 were noted in 9/20 test animals at the 48-hour scoring interval. Dermal reactions in the remaining test and rechallenge control animals were limited to scores of 0 or ±. Group mean dermal scores were higher in the test animals than in the rechallenge control animals. Following rechallenge with 1.0% w/w AS305BD, dermal scores of 1 or 2 were noted in 14/20 test animals at the 24-hour scoring interval and dermal scores of 1 were noted in 12/20 test animals at the 48 -hour scoring interval. Dermal reactions in the remaining test and rechallenge control animals were limited to scores of 0 or ±. Group mean dermal scores were higher in the test animals than in the rechallenge control animals.

Under the conditions of this test, AS305BD meets the criteria to be considered a contact sensitizer.