피부감작성 동물대체시험법인 ARE-Nrf2 루시퍼라아제 LuSens 시험법(OECD TG 442D)의 국내 확립 연구 = Establishment Study of ARE-Nrf2 Luciferase LuSens Skin Sensitization Test(OECD TG 442D) in Korea

다국어 초록 (Multilingual Abstract)

This study aimed to establish the LuSens test method for identification of skin sensitisers in our laboratory and to facilitate the domestic use of the method. We utilized 10 recommended proficiency substances in OECD TG 442D consisting of 6 skin sensitisers (UN GHS category 1A and 1B: Eugenol, Cinnamyl alcohol, 2-Mercaptobenzo-thiazole, 4-Methylaminophenol sulfate, Methyl dibromo glutaronitrile and 2,4-Dinitro-chlorobenzene) and 4 non-sensitisers (No category: Salicylic acid, Glycerol, Isopropanol and Sulfanilamide). We measured the activity of luciferase induced by the test substances based on the CV₇₅ that was determined by cytotoxicity dose-finding test. While the maximal luciferase fold induction values for each skin non-sensitisers ranged from 0.95 to 1.28, those for each skin sensitisers ranged from 1.96 to 5.66. We predicted sensitivity of the test substances on the basis of the luciferase fold induction values. Our results were within the range of acceptance criteria and accord with in vivo and in vitro references in OECD TG 442D. We found that the LuSens test method correctly identified proficiency substances into sensitisers and non-sensitisers. Therefore, the results obtained from the proficiency test demonstrated that we successfully introduced the LuSens test method in our laboratory. Furthermore, we have prepared a new in vitro skin sensitization test (ARE-Nrf2 luciferase LuSens) guideline for the safety evaluation of cosmetics and contributed to the dissemination of the method via technical transfer in Korea.

참고문헌 (Reference)

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2 Natsch A., "The Nrf2-Keap1-ARE Toxicity Pathway as a Cellular Sensor for Skin Sensitizers-Functional Relevance and Hypothesis on Innate Reactions to Skin Sensitizers" 113 (113): 284-292, 2010

3 OECD, "Series on Testing and Assessment No. 168. The Adverse Outcome Pathway for Skin Sensitisation Initiated by Covalent Binding to Proteins. Part 1:Scientific Evidence" Organisation for Economic Cooperation and Development

4 OECD, "OECD Guideline for testing of chemicals, Section 4: Health effects, Test No. 442D. IN VITRO SKIN SENSITISATION ASSAYS ADDRESSING THE AOP KEY EVENT ON KERATINOCYTE ACTIVATION"

5 Ramirez T., "LuSens : a keratinocyte based ARE reporter gene assay for use in integrated testing strategies for skin sensitization hazard identification" 28 : 1482-1497, 2014

6 "DB-ALM Protocol No. 184. LuSens Assay"