H-CLAT

Advances in the understanding of the immunobiology of skin sensitization have led to the establishment of predictive in vivo tests which not only identify sensitizing hazards but also characterize their potency. Recently, appreciation of the underlying biology has also resulted in the development of mechanistically based in vitro alternatives which offer the prospect of the replacement of current in vivo methods. Assays under active validation include the Direct Peptide Reactivity Assay (DPRA), the human Cell Line Activation Test (h-CLAT), and KeratinoSens. None of the methods have a sufficient level of accuracy or freedom from applicability domain limitations to allow them to act as a standalone replacement. Consequently, it will be necessary to consider how to deploy these assays, perhaps in combination and/or in a structured assessment of skin sensitization hazard, to ensure at least the same level of predictive accuracy as the in vivo methods. However, a challenge remains the capacity of these methods to provide potency information on skin-sensitizing chemicals has yet to be assessed. This is an essential requirement for future risk assessment without use of animal models if we are to retain the same level of human health protection that is currently delivered. 

Extensive reviews concerning the opportunities for the development of in vitro sensitization methods already exist [41-44], These reviews show that essentially all of the methods address one or other of the key mechanistic steps in the induction of skin sensitization—and these are nicely represented in the OECD adverse outcome pathway description [45], From this large body of work, three methods have emerged whose initial promise has been substantiated by demonstration not only of their predictive merits but also by verification of their robustness in terms of inter-laboratory transferability and within and between laboratory reproducibility [46]. The three methods are the direct peptide reactivity assay (DPRA) [47, 48], KeratinoSens [49, 50], and the human Cell Line Activation Test (h-CLAT) [51-53]. The first of these, the DPRA, addresses the question of chemical reactivity, the second investigates an aspect of keratinocyte activation... 

Human Cell Line Activation Test (h-CLAT)... 

Regarding each of the above three methods, DPRA, Keratin oSens, and h-CLAT, all of them have well defined protocols and prediction models and each assay has been tested with >100 substances, achieving a predictive accuracy of approximately 80 %. It is for this reason that they have been able to enter the formal validation process [46, 59],... 

Ashikaga T, Yoshida Y, Hirota M, Yoneyama K, Itagaki H, Sakaguchi H, Miyazawa M, Ito Y, Suzuki H, Toyoda H (2006) Development of an in vitro skin sensitization test using human cell lines the human cell line activation test (h-CLAT). I. Optimization oftheh-CLAT protocol. Toxicol In Vitro 20 767-773... 

Ashikaga T, Sakaguchi H, Sono S, Kosaka N, Ishikawa M, Nukada Y, Miyazawa M, Ito Y, Nishiyama M, Itagaki H (2010) A comparative evaluation of in vitro skin sensitisation tests the human cell-line activation test (h-CLAT) versus the local lymph node assay (LLNA). ATLA 38 275-284... 

Nukada Y, Ashikaga T, Miyazawa M, Hirota M, Sakaguchi H, Sasa H, Nishiyama N (2012) Prediction of skin sensitization potency of chemicals by human cell line activation test (h-CLAT) and an attempt at classifying skin sensitization potency. Toxicol In Vitro 26(7) 1150-1160... 

ECVAM (2015a) https //eurl-ecvam.jrc.ec.europa.eu/eurl-ecvam-recommendations/eurl-ecvam-recommendation-on-the-human-cell-line-activation-test-h-clat-for-skin-sensitisation-testing (accessed on October 14, 2015). 

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