Methanesulfonamide, N-[2-[(4,6-dimethoxy-1,3,5-triazin-2-yl)carbonyl]-6-fluorophenyl]-1,1-difluoro-N-methyl-

Endpoint summary

Key value for chemical safety assessment

Skin sensitisation

Link to relevant study records

Reference

Endpoint:

skin sensitisation: in vivo (LLNA)

Type of information:

experimental study

Adequacy of study:

key study

Study period:

09 July -04 Aug 2010

Reliability:

1 (reliable without restriction)

Rationale for reliability incl. deficiencies:

other: GLP guideline study

Qualifier:

according to guideline

Guideline:

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

Deviations:

no

Qualifier:

according to guideline

Guideline:

EPA OPPTS 870.2600 (Skin Sensitisation)

Deviations:

no

GLP compliance:

yes

Type of study:

mouse local lymph node assay (LLNA)

Species:

mouse

Strain:

other: CBA/J

Sex:

female

Details on test animals and environmental conditions:

TEST ANIMALS
- Source: R. Janvier, Le Genest St Isle, France
- Age at study initiation: at least 8 weeks old
- Weight at study initiation: 20.6-23.9 g
- Housing: Mice were housed individually in suspended, stainless steel, wire-mesh cages.
- Diet: certified rodent pellet diet and irradiated: A04C-10, S.A.F.E. (Scientific Animal Food and Engineering, Route de Saint Bris, Augy, France); ad libitum
- Water: tap water; ad libitum
- Acclimation period: at least 5 days

ENVIRONMENTAL CONDITIONS
- Temperature (°C): 20-24
- Humidity (%): 40-70
- Air changes (per hr): 10-15
- Photoperiod (hrs dark / hrs light): 12/12

IN-LIFE DATES: From: 2010-07-21 To: 2010-08-02

Vehicle:

dimethylformamide

Concentration:

25, 50 and 100% (w/v)

No. of animals per dose:

5

Details on study design:

RANGE FINDING TESTS:
- Irritation: Concentrations of test substance in dimethylformamide were chosen based on results of a screening test, in which no irritation (measured by ear weight) was observed at concentrations of 100%. Mean ear weight value of 12.6 mg was measured in the group treated at a concentration of 100%. This represented a variation of 5.9% in relation to the value obtained in control animals (11.9 mg). The values in treated and control groups were considered similar and the formulation was not considered to be irritant at 100%.

MAIN STUDY
ANIMAL ASSIGNMENT AND TREATMENT
- Name of test method: 3H-methyl thymidine incorporation determined by beta-scintillation
- Criteria used to consider a positive response: A test substance is regarded as a skin sensitizer if one concentration of the test substance results in a stimulation index of three or greater, when compared to control values in the absence of skin irritation and if there is a dose-related response.

TREATMENT PREPARATION AND ADMINISTRATION:
25 µL of test substance was applied to the dorsal surface of each ear of each mouse. The application was repeated on Days 2 and 3; local irritation reactions were assessed. On Day 6 an injection of 250 µL saline (0.9% NaCl) containing 20 µCi of 3H-methyl thymidine (3H-TdR) was made into the tail vein of each experimental mouse. Five hours later (± 30 min), the draining auricular lymph nodes from each mouse were placed in an individual tube containing physiological saline and were disaggregated by crushing with a plastic piston. A cell suspension was obtained, free of connective tissue. Cell suspensions were washed with 4 mL of 0.9% physiological saline, centrifuged for 20 min at 1800 rpm and the pellets obtained were resuspended in 2 mL of 5% trichloroacetic acid and stored overnight at 5±3°C. After a final centrifugation, the pellets were resuspended in 1 mL of saline, mixed and then placed for approximately 25 min in an Ultrasonic Bath to ensure a thoroughly dispersed suspension. Once prepared cell suspensions were added to numbered scintillation pots containing 19 mL of scintillation fluid and assayed in a beta-counter.

Positive control substance(s):

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

Positive control results:

The stimulation index (SI) value of the positive control alpha-hexylcinnamaldehyde was 4.5 (±1.1) at a treatment concentration of 30%, thus meeting the reliability criteria for the LLNA (SI >3: positive response).

Key result

Parameter:

SI

Remarks:

25%

Value:

0.6

Variability:

SD +/- 0.2

Test group / Remarks:

n = 5

Key result

Parameter:

SI

Remarks:

50%

Value:

0.5

Variability:

SD +/- 0.2

Test group / Remarks:

n = 5

Key result

Parameter:

SI

Remarks:

100%

Value:

0.7

Variability:

SD +/- 0.4

Test group / Remarks:

n = 5

Mortality was observed on two animals treated with the positive control. Before death these animals had soiled fur. No mortality or clinical signs were observed on the surviving animals of the positive control group, on the control animals or on animals treated with the test substance. No cutaneous reactions were observed in the vehicle, positive control or treated groups.

The disintegrations per minute (DPM) values of the test substance were 845 (± 430.7), 766 (± 297.3) and 987 (± 627.3) at treatment concentrations of 25, 50 and 100%, respectively. The DPM value of the control group was 1489 (± 430.7).

Interpretation of results:

not sensitising

Remarks:

Migrated information Criteria used for interpretation of results: EU

Conclusions:

CLP: not classified
DSD: not classified

Endpoint conclusion

Endpoint conclusion:

no adverse effect observed (not sensitising)

Additional information:

Triafamone was tested for skin sensitisation properties on mice using the local lymph node assay (LLNA) according to OECD 429 (2002). The study (M-390512-01-1) was conducted testing 25 female CBA/K mice, allocated to 5 groups containing 5 animals each. One control group received the vehicle Dimethylformamide (DMF). Three groups received the test substance at a concentration of 25%, 50% in vehicle, or 100% neat test substance. One positive control group received 30% alpha-Hexylcinnamaldehyde in vehicle. A preliminary screening test suggested that the undiluted test substance would not produce systemic toxicity or excessive local irritation. The mice were treated by topical application of the appropriate test concentrations to the dorsal surface of each ear for 3 consecutive days. 5 days following the first application of the test substance, all mice were injected with ³H-methyl thymidine (³HTdR) and sacrificed 5 h afterwards. The draining auricular lymph nodes from each mouse were excised. ³HTdR incorporation in lymphocytes was measured by beta-scintillation counting. Mortality was observed on 2 animals treated with the positive control (alpha-Hexylcinnamaldehyde). Before death these animals had soiled fur. No mortality or clinical signs were observed on the surviving animals of the positive control group, on the control animals or on animals treated with the test substance. No cutaneous reactions were observed in the vehicle, positive control or treated groups.

The mean stimulation (SI) index values of the test substance were 0.6, 0.5, and 0.7 at treatment concentrations of 25, 50 and 100%, respectively. The SI value of the positive control was 4.5 at treatment concentration of 30%, thus meeting the reliability criteria for the LLNA (SI >3: positive response). Under the test conditions, Triafamone was not considered to be a skin sensitiser, since all SI values were less than 3.

Migrated from Short description of key information:
skin sensitisation (OECD 429): not sensitising

Justification for selection of skin sensitisation endpoint:
The reliable GLP compliant OECD Guideline study was chosen.

Justification for classification or non-classification

The available data on skin sensitisation of the test substance do not meet the criteria for classification according to Regulation (EC) 1272/2008 or Directive 67/548/EEC, and are therefore conclusive but not sufficient for classification.