The Globally Harmonized System for Classification and Labeling of Chemicals (GHS) defines a skin sensitizer as "a substance that will induce an allergic response following skin contact" (UNECE, 2004, p. 151). A substance is classified as a skin sensitizer when human data show it can induce a sensitization response following skin contact "in a substantial number of persons" or when "there are positive results from an appropriate animal test" (UNECE, 2004, p. 152). Human experiments to evaluate the skin sensitization potential of a material are not generally conducted, but human testing might be done to confirm a negative animal test result. Human data could also be the result of an epidemiological study or documentation of allergic contact dermatitis from more than one dermatology clinic.
Skin sensitization is a skin response to a hapten (a foreign, low molecular weight substance) that acts like an allergen. In some individuals, certain haptens can induce a type IV (delayed, cell-mediated) hypersensitivity response of the skin. The hapten (allergen) must be able to penetrate the skin, combine with skin proteins, and then produce an immune response (hapten-specific T cells are primed in lymph nodes by dendritic (Langerhans) cells that emigrate from the skin). The initial exposure is called the sensitization phase and has no clinical symptoms. The delayed skin response from a later exposure to the allergen is called the elicitation phase. Clinical symptoms include erythema (redness), vesicles/bullae, papules, scaling, and pruritus (itching). Common examples of substances that can induce a skin sensitization reaction in certain individuals—also known as allergic contact dermatitis—include metals in jewelry and chemicals in cosmetics or in latex gloves. The article Allergies Caused by Consumer Products and Foods provides an excellent overview of the types and mechanisms of allergies.
A diagram of the mechanism of skin sensitization (Gildea, 2005)
The Animal Test(s)
Prior to review of the mouse Local Lymph Node Assay (LLNA) by ICCVAM and ECVAM in 1999, the traditional tests accepted by regulatory authorities for the assessment of a substance's skin sensitization potential were two guinea pig tests, the Guinea Pig Maximization Test (GPMT) and the Buehler Test (OECD, 1992).
Currently, the mouse LLNA is the preferred and accepted method for assessing skin sensitization of most substances (ICCVAM, 1999; ECVAM, 1999; ICCVAM, 2001 (the protocol); OECD, 2002; EPA, 2003). Compared to the GPMT, the LLNA reduces animal use as well as pain and suffering; it is also faster and provides dose-response information (ICCVAM, 2007a). The LLNA is appropriate for testing most types of substances; exceptions such as substances that do not adhere sufficiently to the mouse ear are described in agency documents (ICCVAM, 2007a; EPA, 2003). ECVAM recently validated the reduced Local Lymph Node Assay (rLLNA) for skin sensitization, which provides an additional reduction in numbers of animals used (ECVAM, 2007).
Regulatory Requirements & Test Guidelines
The OECD Test Guideline 429 (TG 429), Skin Sensitisation: Local Lymph Node Assay, was adopted in 2002 (OECD, 2002); the OECD TG 406 describes the traditional animal test methods (OECD, 1992). For animal data, the GHS recommends using the OECD Guidelines, but notes that "other methods may be used provided that they are well-validated and scientific justification is given" (UNECE, 2004, p. 153).
US agencies that may require sensitization testing include the Environmental Protection Agency (EPA), Consumer Product Safety Commission (CPSC), Food and Drug Administration (FDA), and Occupational Safety and Health Administration (OSHA). The EPA, FDA, and OSHA accepted the LLNA in 1999 as an alternative to the GPMT (ICCVAM, 2007a). The EPA's revised test guideline, OPPTS 870.2600 Skin Sensitization, "incorporates the LLNA for use as an alternative method for assessing skin sensitization under the appropriate circumstances," and has been harmonized with the OECD TG 429 (EPA, 2003).
The GHS guidelines and most countries do not differentiate strong from weak sensitizers. However, only substances considered to be strong sensitizers are regulated under the Federal Hazardous Substances Act (FHSA) in the US. The OECD Scientific Issue Paper on Strong vs. Weak Sensitizers describes current approaches of different countries and regions to potency determination, as well as harmonization issues needing further consideration for classification purposes (OECD, 2006).
Non-animal Alternative Methods
There are currently no validated in vitro or in silico methods to replace animal testing for the detection of skin sensitizers. However, many promising methods are in various stages of development and use. It is expected that a predictive method to totally replace animal testing will be a test battery composed of molecular, cell-based, and/or computational methods.
Cell-based assays: Promising cell-based assays for skin sensitization are being developed and refined. These methods use cultured cells to model the mechanism(s) of induction of contact allergy in the skin. Such mechanisms include epidermal cell and dendritic cell interactions, and alterations in dendritic cell phenotype and function following allergen exposure.
Peptide reactivity (or depletion) assay: Chemical allergens bind to skin proteins and/or peptides as part of their mechanism of inducing skin sensitization. Quantification of this peptide reactivity represents another potential non-animal screening assay for skin sensitization.
(Quantitative) structure activity relationship [(Q)SAR]: Structure activity systems are being used and refined and may become validated within the next five years. These models classify allergens and nonallergens on the basis of physiochemical data and the reactivity parameters of functional groups.
Predictive Test Battery: It is anticipated that (Q)SAR and molecular and/or cell-based assays will be used together to form predictive test batteries to identify human skin sensitizers and replace animal testing.
More about recent developments in non-animal test methods can be found in Emerging Research, Methods & Policies.
Validation and Acceptance of Non-animal Alternative Methods
Validated non-animal methods are not yet available for assessing skin sensitization. The only validated alternative methods are the mouse LLNA and the mouse rLLNA.
An ECVAM panel estimated that (Q)SAR systems will be validated by 2010-12, but that in vitro test methods could take longer (ECVAM, 2005, p. 9). The following diagram shows the panel's proposed timeline for validating non-animal alternatives for skin sensitization testing of cosmetics.