L-Leucine, 4-fluoro-N-[(1S)-2,2,2-trifluoro-1-[4'-(methylsulfonyl)[1,1'-biphenyl]-4-yl]ethyl]-, compd. with N-cyclohexylcyclohexanamine (1:1)

Skin sensitisation

Administrative data

Endpoint:

skin sensitisation: in vivo (LLNA)

Type of information:

experimental study

Adequacy of study:

key study

Study period:

28 September - 04 October, 2005

Reliability:

1 (reliable without restriction)

Rationale for reliability incl. deficiencies:

other: Done by OECD and GLP standards

Data source

Reference

Reference Type:

study report

Title:

Unnamed

Year:

2005

Report date:

2005

Materials and methods

Test guideline

Test guideline 1

Qualifier:

according to guideline

Guideline:

OECD Guideline 429 (Skin Sensitisation: Local Lymph Node Assay)

Test guideline 2

Qualifier:

according to guideline

Guideline:

other: Durand, G., G. de Burlet, M. Virat and B.D. Naumann. “Use of the local lymph node assay in the evaluation of the sensitizing potential of pharmaceutical process intermediates”. Contact Dermat. 49 (2003):148-154

GLP compliance:

yes

Remarks:

40 CFR Part 792

Type of study:

mouse local lymph node assay (LLNA)

Test material

Test material information

Constituent 1

Reference substance name:

N-cyclohexylcyclohexanaminium (2S)-4-fluoro-4-methyl-2-{[(1S)-2,2,2-trifluoro-1-{4'-methanesulfonyl-[1,1'-biphenyl]-4-yl}ethyl]amino}pentanoate

EC Number:

700-778-8

Cas Number:

1064076-86-3

Molecular formula:

C21H23F4NO4S.C12H23N

IUPAC Name:

N-cyclohexylcyclohexanaminium (2S)-4-fluoro-4-methyl-2-{[(1S)-2,2,2-trifluoro-1-{4'-methanesulfonyl-[1,1'-biphenyl]-4-yl}ethyl]amino}pentanoate

In vivo test system

Test animals

Species:

mouse

Strain:

other: CBA/J Mice (Mus musculus)

Sex:

female

Details on test animals and environmental conditions:

At the start of the study the mice were in the weight range of 17.1 to 22.8 grams and were 8 - 12 weeks old.
Animlas sourced from Harlan, Indianapolis, IN, USA.
Housing: Group housing (maximum 5 per cage of same sex)
Bedding: Hardwood chips, P.W.I. Industries, St-Hyacinthe,
Quebec, Canada (contact)
Acclimatization: Minimum 5 days under the same conditions as for the actual test
Animal Room Temperature: 68° ± 5 °F
Animal Room Relative Humidity: 30 - 70%
Air Exchanges per Hour: 10 to 15
Lights: 12-hour light/dark cycle, full spectrum fluorescent lights
The laboratory and animal rooms were maintained as limited-access facilities.
Animal Rations: TEK 7012 Rodent Diet, Harlan Taklad, Madison, WI, ad libitum
Water: Tap water, ad libitum
There were no known contaminants present in the feed, water, or bedding expected to nterfere with the test data.
Animal Selection: Selected from a larger pool of animals and examined to ensure lack of adverse clinical signs.
.

Study design: in vivo (LLNA)

Vehicle:

dimethyl sulphoxide

Concentration:

The test substance was evaluated at 3 different concentrations, 10%, 1.0%, and 0.1%

No. of animals per dose:

Animals Used in the Test:
5 animals per concentration group (15)
1 negative control group (5)
1 positive control group (5)
Total number of animals: 25

Details on study design:

On Day 1, 25 µl of the test substance or control substance were applied to the dorsum of both ears.
The same procedure performed on Day 1 was repeated on Days 2 and 3.
Animals were not treated on Days 4 and 5.
On Day 6, 5 hours prior to sacrifice, all animals were injected intravenously with 250 µL of Phosphate-Buffered Saline (PBS) containing 20 µCi methyl-3H-thymidine.
The animals were sacrificed by carbon dioxide inhalation and the draining auricular lymph nodes were excised from each animal. The lymph nodes from each individual animal were pooled.
A single cell suspension of the lymph node cells from each mouse was prepared by gentle mechanical desegregation.
The cell suspension was centrifuged, resuspended in cold 5% TCA, and allowed to precipitate at 4° ± 2 °C for 18 ± 1 hours. After precipitation, the cells were centrifuged and resuspended in fresh 5% TCA.
The level of radioactivity in the cells was measured using a scintillation counter.
The animals were observed for clinical signs daily after dosing. Observations included all clinical and toxicological signs including local irritation at the application site or systemic toxicity.

Positive control substance(s):

hexyl cinnamic aldehyde (CAS No 101-86-0)

Results and discussion

Positive control results:

α-Hexylcinnamaldehyde, was considered to be a sensitiser under the conditions of the test with an average net DPM of 501.46 and a Stimulation index of 6.95

In vivo (LLNA)

Results

Results 1

Parameter:

SI

Remarks on result:

other: see Remark

Remarks:

Concentration of test material in the vehicle at 10 % resulted in a stimulation index (SI) of 1.26 which is a negative result. Concentration of test material in the vehicle at 1 % resulted in a stimulation index (SI) of 1.34 which is a negative result. Concentration of test material in the vehicle at 0.1 % resulted in a stimulation index (SI) of 1.33 which is a negative result. A stimulation index of less than 3 was recorded for the three concentrations of the test material.

Results 2

Parameter:

other: disintegrations per minute (DPM)

Remarks on result:

other: see Remark

Remarks:

Concentration of test material in the vehicle at 10 % resulted in a mean net DPM of 90.64 which is a negative result. Concentration of test material in the vehicle at 1 % resulted in a mean net DPM of 96.86 which is a negative result. Concentration of test material in the vehicle at 0.1 % resulted in a mean net DPM of 96.05 which is a negative result.

Any other information on results incl. tables

Body Weights: All of the test and control animals increased in weight.

Applicant's summary and conclusion

Interpretation of results:

GHS criteria not met

Remarks:

Migrated information

Conclusions:

Based on the test protocol and evaluation criteria, the test substance at 10%, 1.0%, and 0.1% is not considered to be a dermal sensitizing agent.

Executive summary:

The 10%, 1.0%, and 0.1% (w/v) solutions of L-001054748-001X001 in Dimethyl Sulfoxide (DMSO) did not induce a Stimulation Index greater than 3.0 when compared to the vehicle controls. The positive control, 25% hexyl cinnamic aldehyde, did induce a Stimulation Index greater than 3.0, thus validating the test system. Based on the test protocol and evaluation criteria, the test substance at 10%, 1.0%, and 0.1% is not considered to be a dermal sensitizing agent.