测试编号442C：化学皮肤致敏：针对与蛋白质共价结合的不良结果途径关键事件的检测

OECD/OCDEYou are free to use this material subject to the terms and conditions available athttp://www.oecd.org/termsandconditions/OECD GUIDELINE FORTHETESTING OF CHEMICALSIn chemicoskin sensitisation assays addressing the Adverse Outcome PathwayKey Eventon Covalent Binding to ProteinsCovalent binding to proteins Key Event based TestGuideline.A skin sensitiser refers to a substance that will lead to an allergic response following repeated skincontact as defined by the United Nations Globally Harmonized System of Classification andLabelling of Chemicals (UN GHS) (1). There is general agreement on the key biological eventsunderlying skin sensitisation. The current knowledge of the chemical and biological mechanismsassociated with skin sensitisation has been summarised as an Adverse Outcome Pathway (AOP)(2) starting with a molecularinitiating event through intermediate events to the adverse effect,namely allergic contact dermatitis. This AOP focuses on chemicals that react with amino-acidresidues (i.e. cysteine or lysine) such as organic chemicals. In this instance, the molecularinitiatingevent (i.e. the first key event), is the covalent binding of electrophilic substances to nucleophiliccentres in skin proteins. The second key event in this AOP takes place in the keratinocytes andincludes inflammatory responses as well as changes in gene expression associated with specificcell signalling pathways such as the antioxidant/electrophile response element (ARE)-dependentpathways. The third key event is the activation of dendritic cells, typically assessed by expressionof specificcell surface markers, chemokines and cytokines. The fourth key event is T-cellThe assessment of skin sensitisation has typically involved the use of laboratory animals. Theclassical methods that use guinea-pigs, the Guinea Pig MaximisationTest (GPMT) of Magnussonand Kligman and the Buehler Test (OECD TG 406) (11) assess both the induction and elicitationphases of skin sensitisation. The murine tests, such as the LLNA (OECD TG 429) (12) and its threenon-radioactive modifications—LLNA:DA (OECD TG 442A) (13), LLNA:BrdU-ELISA, and BrdU-FCM (OECD TG 442B) (14)—all assess the induction response exclusively and have gainedacceptance, since they provide an advantage over the guinea pig tests in terms of animal welfaretogether with an objective measurement of the induction phase of skin sensitisation.Mechanistically-basedin chemicoandin vitrotest methods addressing the first three key eventsof the skin sensitisation AOP have been adopted for contributing to the evaluation of the skinsensitisation hazard potential of chemicals: the present Test Guideline assesses covalent bindingto proteins, addressing the first key event; the OECD TG 442D assesses keratinocyte activation(15), the second key event and the OECD TG 442E addresses the activation of dendritic cells (16),the third key event of the skin sensitisation AOP. Finally, the fourth key event representing T-cellproliferation is indirectly assessed in the murine Local Lymph Node Assay (LLNA) (12). 442CAdopted:20 June 2022Corrected: 25 June 2024 ©OECD, (2024)INTRODUCTION1.proliferation.2.3. OECD/OCDE2Background and principles of the test methods included in the Key Event based4.This Test Guideline (TG) describesin chemicoassays that address mechanisms described underthe first key event of the AOP for skin sensitisation, namely covalent binding to proteins (2). Thetest methods currently included in this Test Guideline are:•The Direct Peptide Reactivity Assay (DPRA) (Appendix I),•The Amino Acid Derivative Reactivity Assay (ADRA) (Appendix II), and•The kinetic Direct Peptide Reactivity Assay (kDPRA) (Appendix III).5.The test methods are based onin chemicocovalent binding to proteins and are considered to bescientifically valid. The DPRA has been evaluated in a European Union Reference Laboratory forAlternatives to Animal Testing (EURL ECVAM)-lead validation study and subsequent independentpeer review by the EURL ECVAM Scientific Advisory Committee (ESAC) (3) (4) (5). The ADRAunderwent a validation study coordinated by the Japanese Center for the Validation of AlternativeMethods (JaCVAM) (6) (7) (8) (9) followed by an independent peer-review (10). The kDPRAunderwent an industry-coordinated validation study followed by an independent peer-review (17).6.The correlation of protein reactivity with skin sensitisation potential is well established (18) (19)(20). Nevertheless, since protein reactivity represents only one key event of the skin sensitisationAOP (2) (21), information generated with test methods developed to address this specific keyevent may not be sufficient as stand-alonemethods to conclude on the presence or absence ofskin sensitisation potentialof chemicals.Therefore,data generated with the test methodsdescribed in this Test Guideline are proposed to be used within Integrated Approaches to Testingand Assessment (IATA), together with other relevant complementary information fromin vitroassay