Cosmetic products need to have a proven efficacy combined with a comprehensive toxicological assessment. Before the current Cosmetic regulation N°1223/2009, the 7th Amendment to the European Cosmetics Directive has banned animal testing for cosmetic products and for cosmetic ingredients in 2004 and 2009, respectively. An increasing number of alternatives to animal testing has been developed and validated for safety and efficacy testing of cosmetic products and cosmetic ingredients. For example, 2D cell culture models derived from human skin can be used to evaluate anti-inflammatory properties, or to predict skin sensitization potential; 3D human skin equivalent models are used to evaluate skin irritation potential; and excised human skin is used as the gold standard for the evaluation of dermal absorption. The aim of this manuscript is to give an overview of the main in vitro and ex vivo alternative models used in the safety testing of cosmetic products with a focus on regulatory requirements, genotoxicity potential, skin sensitization potential, skin and eye irritation, endocrine properties, and dermal absorption. Advantages and limitations of each model in safety testing of cosmetic products are discussed and novel technologies capable of addressing these limitations are presented.

Toxicological assessments of skin sensitizers have progressed towards a higher reliance on non-animal methods. Current technological trends aim to extend the utility of non-animal methods to accurately characterize skin-sensitizing potency. The GARDskin Dose–Response assay has previously been described; it was shown that its main readout, cDV₀ concentration, is associated with skin-sensitizing potency. The ability to predict potency from cDV₀ in the form of NESILs derived from LLNAs or human NOELs was evaluated. The assessment of a dataset of 30 chemicals showed that the cDV₀ values still correlated strongly and significantly with both LLNA EC3 and human NOEL values (ρ = 0.645–0.787 [p < 1 × 10⁻³]). A composite potency value that combined LLNA and human potency data was defined, which aided the performance of the proposed model for the prediction of NESILs. The potency model accurately predicted sensitizing potency, with cross-validation errors of 2.75 and 3.22 fold changes compared with NESILs from LLNAs and humans, respectively. In conclusion, the results suggest that the GARDskin Dose–Response assay may be used to derive an accurate quantitative continuous potency estimate of skin sensitizers.

New approach methods (NAMs) for hazard identification of skin sensitizing chemicals have been adopted as test guidelines by the OECD during the last decade as alternatives to animal models. These models align to individual key events (KE) in the adverse outcome pathway (AOP) for skin sensitization for which the molecular initiating event (MIE) is covalent binding to proteins. As it currently stands, the AOP does not include mechanistic events of sensitization by metals, and limited information is available on whether NAMs accurately the predict sensitization potential of such molecules, which have been proposed to act via alternative mechanisms to organic chemicals. Methods for assessing the sensitization potential of metals would comprise valuable tools to support risk management within e.g., occupational settings during production of new metal salts or within the medical device industry to evaluate leachables from metal alloys. This paper describes a systematic evaluation of the applicability domain of the GARD™skin assay for assessment of metals. Hazard classifications were supplemented with an extended analysis of gene expression profiles induced by metal sensitizers to compare the induction of toxicity pathways between metals and organic sensitizers. Based on the results of this study, the accuracy, sensitivity, and specificity of GARD™skin for prediction of skin sensitizing hazard were 92% (12/13), 100% (7/7) and 83% (5/6), respectively. Thus, the performance of GARD™skin for assessment of metals was found to be similar to what is observed on conventional organic substances, providing support for inclusion of metals within the applicability domain of the test method.