Comparative assessment of 24-hr primary skin irritation test and human patch test data with <i>in vitro</i> skin irritation tests according to OECD Test Guideline 439 (for quasi-drugs in Japan)

Abstract

The Organisation for Economic Cooperation and Development (OECD) Test Guideline (TG) 439 is an in vitro test method of reconstructed human epidermis (RhE), which was developed for hazard identification of irritating chemicals in accordance with a primary skin irritation test using rabbits with 4-hr exposure. A regulation for quasi-drugs in Japan requires data from primary skin irritation tests using rabbits to undergo 24-hr exposure, and this is used as an evidence for 24-hr closed patch tests in humans. In this study with the same chemicals, primary skin irritation test data using rabbits undergoing 24-hr exposure and a 24-hr occlusive human patch test data were analyzed by comparing the results obtained with four test methods adopted in OECD TG 439. The performances of in vitro test methods showed a positive predictive value of 72.7-85.7% to predict the results of 24-hr primary rabbit skin irritation test knowing that its positive predictive value was 57.1% against humans only. The prediction factors of in vitro test methods were higher for the human patch test data with a sensitivity reaching 60 to 80%. Three surfactants gave false negatives in some of the RhE methods evaluated with the human patch test, but in each case, they were correctly classified as positive when evaluated at double concentration. Therefore, the approach of setting the margin to 2 was effective in eliminating false negatives. This suggests that in vitro test methods are useful for assessing skin irritation potential without animal testing for the application of quasi-drugs in Japan.

Predicting the results of a 24-hr human patch test for surfactants: utility of margin-setting in a reconstructed human epidermis model

“…For the purpose of examining the reproducibility of cytotoxicity and the utility of the margin 2 method, the test chemicals (BC, CPC, SLS, LB, and LC) included in the previous report (Sugiyama et al, 2018) and additional surfactants (SL, OL, and LG) from which patch test data was available (Kanto et al, 2013) were used. Based on the concentration in the result of the patch test, the concentration was set by the common ratio of 2, mainly in the high concentration region, for the purpose of verifying the margin 2 method and calculating IC 50 .…”

“…Based on the concentration in the result of the patch test, the concentration was set by the common ratio of 2, mainly in the high concentration region, for the purpose of verifying the margin 2 method and calculating IC 50 . Table 1 shows the results of the previous study (Run 1; Sugiyama et al, 2018), and those of the reproducibility examination (Run 2, and Run 3).…”

“…4) The data were quoted from (Sugiyama et al, 2018). 5) Classified (C): Viability ≤ 50%, Not Classified (NC): Viability > 50 % 6) When classification of the in vitro test was (C), the patch test was predicted as (I).…”

“…When classification with the in vitro test was (NC), if classification with the in vitro test with double the concentration was (NC), the patch test was predicted as (NI); it would be predicted as (I) with a classification of (C). Data of the 24 hr human patch test of 1) were quoted from (Sugiyama et al, 2018) and 2) were from (Kanto et al, 2013). An IL-1α ELISA Kit (Quantikine, R&D Systems, Abingdon, UK) was used to measure IL-1α, we used, and followed the specified protocol.…”

“…Conversely, evaluation of correspondence with a 24 hr closed human patch test resulted in improved sensitivity and accuracy compared with a primary skin irri-tation test (rabbits, 24 hr exposure); an in vitro test using a RhE model was assumed to be effective for evaluating irritation for humans. However, false negative results for surfactants were noted, and we proposed a method that sets margin of two as a solution (Sugiyama et al, 2018). As reported in OECD TG 439, a survival rate of 50% or less in an in vitro test using the RhE model would be classified as an "irritant" in category 2 on the United Nations (UN) Globally Harmonized System of Classification and Labelling of Chemicals (GHS), and judged as Classified (C).…”