Artificial neural network analysis of data from multiple in vitro assays for prediction of skin sensitization potency of chemicals.

In order to develop in vitro risk assessment systems for skin sensitization, it is important to predict a threshold from the murine local lymph node assay (LLNA). We first confirmed that the combination of the human Cell Line Activation Test (h-CLAT) and the SH test improved the accuracy and sensitivity of prediction of LLNA data compared with each individual test. Next, we assessed the mutual correlations among maximum amount of change of cell-surface thiols (MAC value) in the SH test, CV75 value (concentration giving 75% cell viability) in a cytotoxicity assay, EC150 and EC200 values (thresholds concentrations of CD86 and CD54 expression, respectively) in h-CLAT and published LLNA thresholds of 64 chemicals. Based on the results, we selected MAC value and the minimum of CV75, EC150 (CD86) and EC200 (CD54) as descriptors for the input layer of an artificial neural network (ANN) system. The ANN-predicted values were well correlated with reported LLNA thresholds. We also found a correlation between the SH test and the peptide-binding assay used to evaluate hapten-protein complex formation. Thus, this model, which we designate as the "iSENS ver. 1", may be useful for risk assessment of skin sensitization potential of chemicals from in vitro test data.

Predictive performance for human skin sensitizing potential of the human cell line activation test (h-CLAT).

BACKGROUND: Recent changes in regulatory restrictions and social opposition to animal toxicology experiments have driven the need for reliable in vitro tests for predicting the skin sensitizing potentials of a wide variety of industrial chemicals. Previously, we developed the human cell line activation test (h-CLAT) as a cell-based assay to predict the skin sensitizing potential of chemicals, and showed the correspondence between the h-CLAT and the murine local lymph node assay results. OBJECTIVES: This study was conducted to investigate the predictive performance of the h-CLAT for human skin sensitizing potential. MATERIALS/METHODS: We selected a total of 66 test chemicals with known human sensitizing potential, and tested all chemicals with the h-CLAT. We then evaluated the performance of the h-CLAT in predicting human sensitizing potential. RESULTS AND CONCLUSION: Forty-five of 51 tested sensitizers were positive in the h-CLAT, indicating relatively high sensitivity. Also, 10 of 15 non-sensitizers were correctly detected as negative. The overall agreement between human data and h-CLAT outcome was 83%. Furthermore, the h-CLAT could accurately predict the human sensitizing potential of 23 tested chemicals that were amines, heterocyclic compounds, or sulfur compounds. Our data indicate the utility of the h-CLAT for predicting the human skin sensitizing potential of a variety of chemicals.

A comparative evaluation of in vitro skin sensitisation tests: the human cell-line activation test (h-CLAT) versus the local lymph node assay (LLNA).

We previously developed the human cell-line activation test (h-CLAT) in vitro skin sensitisation test, based on our reported finding that a 24-hour exposure of THP-1 cells (a human monocytic leukaemia cell line) to sensitisers is sufficient to induce the augmented expression of CD86 and CD54. The aim of this study is to confirm the predictive value of h-CLAT for skin sensitisation activity by employing a larger number of test chemicals. One hundred chemicals were selected, according to their categorisation in the local lymph node assay (LLNA), as being: extreme, strong, moderate and weak sensitisers, and non-sensitisers. The correlation of the h-CLAT results with the LLNA results was 84%. There were some false negatives (e.g. benzoyl peroxide, hexyl cinnamic aldehyde) and some false positives (e.g. 1-bromobutane, diethylphthalate). Eight out of the 9 false negatives (89%) were water-insoluble chemicals. The h-CLAT could positively predict not only extreme and strong sensitisers, but also moderate and weak sensitisers, though the detection rates of weak sensitisers and non-sensitisers were comparatively low. Some sensitisers enhanced both CD86 and CD54 levels, and some enhanced the level of only one of them. The use of the combination of CD86 and CD54 induction as a positive indicator, improved the accuracy of the test. In conclusion, the h-CLAT is expected to be a useful cell-based in vitro method for predicting skin sensitisation potential.

The relationship between CD86/CD54 expression and THP-1 cell viability in an in vitro skin sensitization test--human cell line activation test (h-CLAT).

Recent regulations for cosmetics in Europe prohibit animal testing for evaluating the sensitization potential of chemicals to improve animal welfare. Yet, there is not an acceptable Organization for Economic Co-operation and Development non-animal skin sensitization test method. Several in vitro skin sensitization methods that focus on the activation of Langerhans cells, including human cell lines, are being evaluated as possible alternatives. In our previous study, we optimized our human cell line activation test (h-CLAT) using THP-1 cells (monocytic leukemia cell line) and conducted an inter-laboratory study. We found that measuring CD86/CD54 expression may be useful for predicting skin sensitization. The aim of this study was to confirm the relationship between CD86/CD54 expression and THP-1 cell viability in the h-CLAT. In this study, 21 allergens (e.g., dinitrochlorobenzene, p-phenylenediamine, Ni) and 8 non-allergens (e.g., SLS, lactic acid) were evaluated. For each chemical, more than 10 concentrations that gave a predicted cell viability range of 20-95% were used. The data showed that expression patterns of CD86/CD54 differed depending on chemical. For most allergens, cytotoxicity (65-90% cell viability) was needed for enhancement of CD86/CD54 expression. The criteria of "CD86 > or = 150 or CD54 > or = 200" resulted in an accuracy of 93%, which confirms appropriate cut-off criteria for h-CLAT. Furthermore, a good correlation was observed between EC3 of local lymph node assay and EC150(CD86) or EC200(CD54) of h-CLAT (12 or 16 chemicals, respectively), which would provide a useful estimate of allergic potency. These findings suggest that h-CLAT would be a good robust in vitro skin sensitization test.