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Skin sensitisation

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Administrative data

Endpoint:
skin sensitisation: in vitro
Remarks:
“The ARE-Nrf2 Luciferase LuSens Test Method” according to OECD Guideline 442D “In Vitro Skin Sensitisation assays addressing the AOP Key Event on Keratinocyte activation”
Type of information:
experimental study
Adequacy of study:
key study
Study period:
Study plane dated 29. March 2019; Experimental Studing date: 1. April 2019; Experimental completion date: 26. April 2019
Reliability:
1 (reliable without restriction)
Rationale for reliability incl. deficiencies:
guideline study
Remarks:
OECD 442D

Data source

Reference
Reference Type:
study report
Title:
Unnamed
Year:
2019
Report date:
2019

Materials and methods

Test guideline
Qualifier:
equivalent or similar to guideline
Guideline:
OECD Guideline 442D (In Vitro Skin Sensitisation: ARE-Nrf2 Luciferase Test Method)
Version / remarks:
“The ARE-Nrf2 Luciferase LuSens Test Method” according to OECD Guideline 442D “In Vitro Skin Sensitisation assays addressing the AOP Key Event on Keratinocyte activation”
Deviations:
no
Principles of method if other than guideline:
In difference to the OECD 442D guideline, Ethylene glycol dimethylacrylate (EGDMA) was not used as positive control, but p-Phenylenediamine was. This is uncritical since the guide-line indicates that other suitable positive controls, preferentially providing EC 1.5 values in the mid-range, may be used if historical data are available to derive comparable run ac-ceptance criteria.
GLP compliance:
yes (incl. QA statement)
Type of study:
activation of keratinocytes

Test material

Constituent 1
Chemical structure
Reference substance name:
Sodium phenylphosphinate
EC Number:
224-305-5
EC Name:
Sodium phenylphosphinate
Cas Number:
4297-95-4
Molecular formula:
C6H7O2P.Na
IUPAC Name:
sodium phenylphosphinate
Test material form:
solid: particulate/powder
Specific details on test material used for the study:
Sodium phenylphosphinate
Batch no. 20190104
Appearance white powder
Composition approx. 98 % Sodium phenylphosphinate, approx. 1.5 % phenylphosphonic acid, approx. 0.5 % others
Purity 0,98
Homogeneity homogeneous
Expiry date 15. Jan. 2020
Storage Room temperature (20 ± 5°C)
The following information was provided by the sponsor as well:
CAS No. 4297-95-4
EC-No. 224-305-5
Molecular formula C6H6NaO2P
Molecular weight 194.07 g/mol
Vapour pressure not stated
Stability H2O: not stated; EtOH: not stated; acetone: not stated; CH3CN: not stated; DMSO: not stated
Solubility in solvents H2O: not stated; EtOH: not stated; acetone: not stated; CH3CN: not stated; DMSO: not stated
Production date 16. Jan. 2019

In vitro test system

Details on the study design:
Skin sensitisation (In vitro test system) - Details on study design:
This in vitro study evaluates the potential of the test item Sodium phenylphosphinate to ac-tivate the Nrf2 transcription factor (sensitizing potential) 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 be-tween 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 one experiment, consisting of three independent repetitions (repetition I, II and III) with a treatment period of 48 h. The CRFT was performed to detect a potential cytotoxic effect of the test item. Based on these results, the concentrations to be tested in the repetitions were determined.

Results and discussion

Positive control results:
p-Phenylenediamine (60 μM) was used as positive control. The viability was above 70 % and a distinct increase in luciferase induction above 2.5 fold in comparison to the solvent control was detected. In repetition III the luciferase induction is outside the historical data range of the positive control. Since the value is only marginal outside the range, and the repetition is valid regarding the acceptance criteria for the positive and negative control, there is no influence on the result of the study.

In vitro / in chemico

Results
Key result
Parameter:
other: Luciferase induction
Value:
1.5
Vehicle controls validity:
not examined
Negative controls validity:
valid
Positive controls validity:
valid
Remarks on result:
no indication of skin sensitisation

Any other information on results incl. tables

Table 9-a Acceptability of repetition I and III in attached Report - Study No.: 19022101G887

Applicant's summary and conclusion

Interpretation of results:
GHS criteria not met
Conclusions:
Under the experimental conditions of this study, the test item, Sodium phenylphosphinate, was negative in the LuSens assay and is therefore considered not to have the potential to activate the Nrf2 transcription factor (no sensitizing potential).
Executive summary:

This in vitro study evaluates the potential of the test item Sodium phenylphosphinate to ac-tivate the Nrf2 transcription factor (sensitizing potential) 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 be-tween skin sensitizers and non-sensitizers in the context of an integrated approach to testing and assessment.

The LuSens test is an ARE Reporter Gene Assay according to the OECD 442D Guideline with the title “In Vitro Skin Sensitisation assays addressing the AOP Key Event on Keratino-cyte activation”.

The assay included a cytotoxicity range finder test (CRFT) and one experiment, consisting of three independent repetitions (repetition I, II and III) with a treatment period of 48 h. The CRFT was performed to detect a potential cytotoxic effect of the test item. Based on these results, the concentrations to be tested in the repetitions were determined.

Since in repetition II the relative viability of the positive control showed a cytotoxic effect and abnormal high standard deviation values in all controls were visible, it was declared as in-valid. This repetition is not reported, all documentation is kept with the raw data and will be archived at the GLP test facility. To obtain a valid result, repetition III was performed.

In the experiment (repetition I and III), the highest nominal applied concentration (2000 μM) was chosen based on the results obtained in the CRFT. A geometric series (factor 1.2) of eleven dilutions thereof was prepared. Precipitation of the test item was not visible in any of the repetitions.

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 EGDMA (120 μM) as pos-itive control.

No substantial and reproducible dose-dependent increase in luciferase induction equal or above 1.5 fold was observed in both repetitions up to the maximal tested concentration of the test item.

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