INTERNATIONAL PREVALIDATION STUDIES OF THE EPIDERM™ 3D HUMAN RECONSTRUCTED SKIN MICRONUCLEUS (RSMN) ASSAY: TRANSFERABILITY AND REPRODUCIBILITY.

Aardema1, M.J., Barnett1, B.C., Khambatta1, Z., Reisinger2, K., Ouedraogo-Arras3, G., Faquet3, B., Ginestet3, A.-C., Mun4, G.C., Dahl4, E.L., Hewitt5, N.J., Corvi6, R., Curren, R.D. 1Procter & Gamble, Miami Valley Laboratories, 11810 East Miami River Road, Cincinnati, OH 45253, USA. 2Henkel AG & Co. KGaA, Henkelstraße 67, 40589 Düsseldorf, Germany. 3L’Oreal Life Sciences Research, 1, Av Eugene Schueller 93600 Aulnay sous Bois, France. 4Institute for In Vitro Sciences (IIVS), Inc., 30 West Watkins Mill Road, Suite 100, Gaithersburg, MD 20878, USA. 5Scientific Writing Services, Wingertstrasse 25, 64390 Erzhausen, Germany. 6European Centre for the Validation of Alternative Methods (ECVAM), Institute for Health and Consumer Protection (IHCP), European Commission Joint Research Centre (JRC), 21020 Ispra, Va, Italy.

Abstract

Recently, a novel in vitro reconstructed skin micronucleus (RSMN) assay incorporating the EpiDermTM 3D human skin model (Curren et al., Mutat. Res. 607 (2006) 192–204; Mun et al., Mutat. Res. 673 (2009) 92–99) has been shown to produce comparable data when utilized in three different laboratories in the United States (Hu et al., Mutat. Res. 673 (2009) 100–108). As part of a project sponsored by the European cosmetics companies trade association (COLIPA), with a contribution from the European Center for the Validation of Alternative Methods (ECVAM), international prevalidation studies of the RSMN assay have been initiated. The assay was transferred and optimized in two laboratories in Europe, where dose-dependent, reproducibly positive results for mitomycin C and vinblastine sulfate were obtained. Further intra- and inter-laboratory reproducibility of the RSMN assay was established by testing three coded chemicals, N-ethyl-N-nitrosourea, cyclohexanone, and mitomycin C. All chemicals were correctly identified by all laboratories as either positive or negative. These results support the international interlaboratory and inter-experimental reproducibility of the assay and reinforce the conclusion that the RSMN assay in the EpiDermTM 3D human skin model is a valuable in vitro method for assessment of genotoxicity of dermally applied chemicals.

Keywords

7th Amendment to the Cosmetics Directive, Assay reproducibility, Assay transferability, COLIPA, ECVAM, EpiDermTM , EpiDermTM EPI-200-MNA, Inter-experimental reproducibility, Inter-laboratory reproducibility, Micronucleus assay, REACH (Registration, Evaluation, Authorisation and Restriction of Chemical substances), Reconstructed skin, Reconstructed skin micronucleus (RSMN) assay, Validation

Materials Tested

Acetone, Cyclohexanone, Ethanol, Mitomycin C, N-Ethyl-N-nitrosourea, Vinblastine sulfate

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