P-1401

Skin sensitisation

Some information in this page has been claimed confidential.

Administrative data

Endpoint:

skin sensitisation: in vivo (LLNA)

Type of information:

experimental study

Adequacy of study:

key study

Reliability:

1 (reliable without restriction)

Rationale for reliability incl. deficiencies:

guideline study

Data source

Reference

Reference Type:

study report

Title:

Unnamed

Year:

2007

Report date:

2007

Materials and methods

Test guideline

Qualifier:

according to guideline

Guideline:

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

Deviations:

no

GLP compliance:

yes (incl. QA statement)

Type of study:

mouse local lymph node assay (LLNA)

Test material

Test material information

Constituent 1

Test material form:

solid

In vivo test system

Test animals

Species:

mouse

Strain:

other: CBA/CaOlaHsd

Sex:

female

Details on test animals and environmental conditions:

TEST ANIMALS
- Source: Harlan Netherlands, B.V. Postbus 6174, NL - 5960 AD Horst / The Netherlands
- Age at study initiation: 7 - 12 weeks (beginning of acclimatisation)
- Weight at study initiation: 20.1 ± 1.2 g (mean ± SD)
- Housing: Single; Makrolon Type I, with wire mesh top
- Diet (e.g. ad libitum): pelleted standard diet, ad libitum
- Water (e.g. ad libitum): tap water, ad libitum
- Acclimation period: Under test conditions after health examination. Only animals without any visible signs of illness were used for the study.

ENVIRONMENTAL CONDITIONS
- Temperature (°C): 22 ± 3ºC
- Humidity (%): 30 – 70 %
- Photoperiod (hrs dark / hrs light): artificial light 6.00 a.m. - 6.00 p.m.

Study design: in vivo (LLNA)

Vehicle:

dimethyl sulphoxide

Concentration:

2.5, 5 and 10% (w/w)

No. of animals per dose:

4 females (nulliparous and non-pregnant) per group

Details on study design:

RANGE FINDING TESTS:
In a pre-test in two mice, test item concentrations of 1.25, 2.5, 5, and 10 % were tested on one ear each. No irritation effects were observed at these
concentrations after a single application.

MAIN STUDY
- Criteria used to consider a positive response:
A test item is regarded as a sensitiser in the LLNA if the following criteria are fulfilled:
- First, that exposure to at least one concentration of the test item resulted in an incorporation of 3HTdR at least 3-fold or greater than that recorded in control mice, as indicated by the stimulation index.
- Second, that the data are compatible with a conventional dose response, although allowance must be made (especially at high topical concentrations) for either local toxicity or immunological suppression.
The decision to select a STIMULATION INDEX (S.I.) of 3 as an arbitrary indication of sensitizing activity was made on the basis of investigations performed with a wide range of chemicals.

TREATMENT PREPARATION AND ADMINISTRATION:
Each test group of mice was treated by topical (epidermal) application to the dorsal surface of each ear lobe (left and right) with different test item concentrations of 2.5, 5, and 10 % (w/v) in dimethylsulfoxide. The application volume, 25 μl, was spread over the entire dorsal surface (ø 8 mm) of each ear lobe once daily for three consecutive days. A further group of mice was treated with an equivalent volume of the relevant vehicle alone (control animals).

Five days after the first topical application, all mice were administered with 250 μl of 80.4 μCi/ml 3HTdR (corresponds to 20.1 μCi 3HTdR per mouse) by intravenous injection via a tail vein.

Positive control substance(s):

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

Statistics:

The mean values and standard deviations were calculated in the body weight tables.

Results and discussion

Positive control results:

Group 1: concentration 5%: Stimulation index: 2.04
Group 2: concentration 10%: Stimulation index: 6.31
Group 3: concentration 25%: Stimulation index: 12.45
EC3: 6.1% (w/v)<

In vivo (LLNA)

Results

Results 1

Key result

Parameter:

EC3

Remarks on result:

not determinable

Remarks:

In this study Stimulation indices of 2.55, 1.63 and 2.22 were determined with the test item at concentrations of 2.5, 5 and 10 % in dimethylsulfoxide. The EC3 value (concentration of test item required to produce a S.I of 3) could not be calculated, since none of the tested concentrations induced an S.I greater than 3.

Results 2

Parameter:

SI

Value:

2.55

Test group / Remarks:

2.5 %

Results 3

Parameter:

SI

Value:

1.63

Test group / Remarks:

5 %

Results 4

Parameter:

SI

Value:

2.22

Test group / Remarks:

10 %

Cellular proliferation data / Observations:

CELLULAR PROLIFERATION DATA
2.5%: DPM = 8589.9 (1073.7 DMP per lymph node)
5%: DPM = 5490.5 (686.3 DMP per lymph node)
10%: DPM = 7505.11 (7484.8 DMP per lymph node)

EC3 CALCULATION
The EC3 Value could not be calculated, since all SI´s are below 3.

CLINICAL OBSERVATIONS:
No symptoms of local toxicity at the ears of the animals and no systemic findings were observed during the study period.

BODY WEIGHTS
The body weight of the animals, recorded prior to the first application and prior to treatment with 3HTdR, was within the range commonly recorded for animals of this strain and age.

Applicant's summary and conclusion

Interpretation of results:

GHS criteria not met

Remarks:

CLP Implementation.

Conclusions:

The test substance was determined to be non-skin sensitizer in the Local Lymph Node Assay.

Executive summary:

A Local Lymph Node Assay was performed with the test substance P0310 for assessing its possible contact allergenic potential, according to OECD guideline 429 and following GLP. For this purpose a local lymph node assay was performed using test item concentrations of 2.5, 5 and 10% dissolved in dimethylsulfoxide. Four groups of four female CBA/CaOlaHsd mice received topically the three different doses and a positive control respectively. Five days after they received 20.1 μCi 3HTdR/mouse, by intravenous injection. The animals were examined for clinical signs, body weight and viability/mortality. The animals did not show any clinical signs during the course of the study and no cases of mortality were observed. In this study Stimulation Indices (S.I) of 2.55, 1.63 and 2.22 were determined with the test item at concentrations of 2.5, 5 and 10% dimethylsulfoxide, respectively. Based on these results, the test item was determined not to be a skin sensitizer.