C.I. Solvent Red 119

Skin sensitisation

Administrative data

Endpoint:

skin sensitisation: in vitro

Type of information:

experimental study

Adequacy of study:

key study

Study period:

May 2017- October 2017

Reliability:

1 (reliable without restriction)

Rationale for reliability incl. deficiencies:

guideline study

Data source

Reference

Reference Type:

study report

Title:

Unnamed

Year:

2017

Materials and methods

Test guideline

Test guideline 1

Qualifier:

according to guideline

Guideline:

OECD Guideline 442D (In Vitro Skin Sensitisation: ARE-Nrf2 Luciferase Test Method)

Version / remarks:

The LuSens test is an ARE Reporter Gene Assay that was developed by the BASF SE (Ludwigshafen, Germany) and is only based on the OECD 442D Guideline. The used cell line as well as the negative and the positive control is different in comparison to the OECD 442D guideline. Also the test performance differs in details, even if the general procedure is similar to the one described in the BASF SE protocol. However, since it was already announced that this study is performed in accordance with the BASF SE protocol and only based on the OECD 442D guideline, those deviations do not affect the result of this study.

Deviations:

yes

Remarks:

See remarks.

Test guideline 2

Qualifier:

according to guideline

Guideline:

other: LuSens Assay Protocol provided by BASF SE

Deviations:

no

GLP compliance:

yes (incl. QA statement)

Type of study:

other: ARE-Nrf2 Luciferase Test

Justification for non-LLNA method:

This in vitro study is performed to assess the potential of the test item Solvent Red 119 to activate the Nrf2 transcription factor by using the genetically modified keratinocyte cell-line “LuSens” (Bauch et al. 2012).

Test material

Test material information

Constituent 1

Reference substance name:

C.I. Solvent Red 119

EC Number:

602-676-8

Cas Number:

12237-27-3

Molecular formula:

C32H22N10O8Cr

IUPAC Name:

C.I. Solvent Red 119

impurity 1

Reference substance name:

Unknown impurities

Cas Number:

not available

IUPAC Name:

Unknown impurities

impurity 2

Reference substance name:

Water

EC Number:

231-791-2

EC Name:

Water

Cas Number:

7732-18-5

Molecular formula:

H2O

IUPAC Name:

water

Test material form:

solid: particulate/powder

Results and discussion

In vitro / in chemico

Results

Key result

Run / experiment:

other: LuSens Test

Parameter:

other: luciferase induction

Value:

1.5

Vehicle controls validity:

valid

Negative controls validity:

valid

Positive controls validity:

valid

Remarks on result:

no indication of skin sensitisation

Remarks:

Due to the low test item concentrations, the result is inconclusive in accordance to the OECD 442D.

Applicant's summary and conclusion

Interpretation of results:

GHS criteria not met

Conclusions:

This in vitro study was performed to investigate the potential of Solvent Red 119 to acti-vate the Nrf2 transcription factor, by using the LuSens cell line. For this purpose, two inde-pendent experiments were performed.
A detailed listing of all measured and calculated values of the assay is given in annex 2 (values of CRFT), annex 3 (values of experiment I), and annex 4 (values of experiment II). In addition, the final results of both experiments are summarized in table 8-a and 8-b and graphically illustrated in figure 8-a to 8-d.
The assay was performed in two independent experiments. 8 concentrations of the test item were evaluated. The exposure time was 48 h. The following nominal concentrations of the test item were investigated in experiment I and II: 0.85 μg/mL, 1.02 μg/mL, 1.22 μg/mL, 1.47 μg/mL, 1.76 μg/mL, 2.11 μg/mL, 2.53 μg/mL, 3.04 μg/mL, 3.65 μg/mL, 4.38 μg/mL, 5.25 μg/mL, 6.30 μg/mL
None of the real treatment concentrations in both experiments deviated more than 10 % from the nominal concentration. Precipitation of the test item was not visible up to the highest concentration.
EGDMA (120 μM) was used as positive control. The viability was above 70 % and a dis-tinct increase in luciferase induction above 2.5 fold in comparison to the solvent control was detected.
DL-lactic acid (5000 μM) was used as negative control. The viability was above 70 % and the induction of the luciferase was < 1.5 fold in comparison to the solvent control.
The induction of the luciferase of the growth control (Medium no. 3) was < 1.5 fold.
Since all acceptability criteria of the assay were met (see chapter 9.1) the study is valid.
In experiment I and II cytotoxic effects were observed at 3.65 μg/mL, 4.38 μg/mL, 5.25 μg/mL and 6.30 μg/mL. Those concentrations were excluded from the evaluation of the luciferase induction.
Finally the following test item concentrations showed a viability ≥ 70 % and could therefore be evaluated for luciferase induction in experiment I and II: 0.85 μg/mL, 1.02 μg/mL, 1.22 μg/mL, 1.47 μg/mL, 1.76 μg/mL, 2.11 μg/mL, 2.53 μg/mL and 3.04 μg/mL
In all tested concentrations of the test item no substantial and reproducible dose depend-ent increase of luciferase induction was measured.
In conclusion, it can be stated that under the experimental conditions reported, the test item did not induce the luciferase activity in the LuSens cells above the threshold.
The recorded data in this study declare the test item Solvent Red 119 does not has the potential to activate the Nrf2 transcription factor. According to the OECD 442 D, the result is inconclusive because of the low test item concentrations.

Executive summary:

This in vitro study evaluates the potential of the test item Solvent Red 119 to activate the Nrf2 transcription factor by using the LuSens cell line. This test is part of a tiered strategy for the evaluation of skin sensitization potential. Thus, data generated with the present Test Guideline should be used to support the discrimination between skin sensitizers and non-sensitizers in the context of an integrated approach to testing and assessment.

The assay included a cytotoxicity range finder test (CRFT) and two independent experi-ments (experiment I and II) with a treatment period of 48 h. The CRFT was performed to detect a potential cytotoxic effect of the test item. Based on the results of this test the con-centrations for the two experiments were determined.

In the main experiments, the highest nominal applied concentration (6.3 μg/mL) was cho-sen based on the results obtained in the CRFT. A geometric series (factor 1.21) of eleven dilutions thereof was prepared. Precipitation of the test item was not visible in any of the experiments.

DMSO (final concentration: 1 %) was used as solvent control and medium no. 3 as growth control. Lactic acid (5000 μM) was used as negative control and EGDMA2 (120 μM) as positive control.

The evaluated experimental points and the results are summarised in chapter 8.

Conclusion:

Under the experimental conditions of this study, the test item, Solvent Red 119, was nega-tive in the LuSens assay and is therefore considered not having the potential to activate the Nrf2 transcription factor. According to the OECD 442 D, the result is inconclusive be-cause of the low test item concentrations.

No substantial and reproducible dose dependent increase in luciferase induction above 1.5 fold was observed in both experiments up to the maximal non-cytotoxic concentration of the test item.