Borderline Range Determined Using Data From Validation Study of Alternative Methods for Skin Sensitization: ADRA, IL-8 Luc Assay, and EpiSensA.

Most predictive models that use alternatives to animal experiments divide judgements into two classes with a cutoff value for each model. However, if the results of alternative methods are close to the cutoff values, the true result may be ambiguous because of variability in the data. Therefore, the OECD GL497 uses a judgement method that establishes a borderline range (BR) around a cutoff value using a statistical method.

Evaluation of the immunotoxicity potential of nanomaterials using THP-1 cells.

With the expansion of nanomaterials (NMs) usage, concerns about their toxicity are increasing, and the wide variety of NMs makes it difficult to assess their toxicity. Therefore, the development of a high-throughput, accurate, and certified method to evaluate the immunotoxicity of NMs is required. In this study, we assessed the immunotoxicity potential of various NMs, such as nanoparticles of silver, silica, and titanium dioxide, using the human Cell Line Activation Test (h-CLAT) at the cellular level.

The inter-laboratory validation study of EpiSensA for predicting skin sensitization potential.

The Epidermal Sensitization Assay (EpiSensA) is a reconstructed human epidermis (RhE)-based gene expression assay for predicting the skin sensitization potential of chemicals. Since the RhE model is covered by a stratified stratum corneum, various kinds of test chemicals, including lipophilic ones and pre-/pro-haptens, can be tested with a route of exposure akin to an in vivo assay and human exposure. This article presents the results of a formally managed validation study of the EpiSensA that was carried out by three participating laboratories.

Development of an Sensitisation Test Using a Coculture System of Human Bronchial Epithelium and Immune Cells.

Chemical respiratory sensitisation is a serious health problem. However, to date, there are no validated test methods available for identifying respiratory sensitisers. The aim of this study was to develop an sensitisation test by modifying the human cell line activation test (h-CLAT) to detect respiratory sensitisers and distinguish them from skin sensitisers.

Validation study for in vitro skin irritation test using reconstructed human skin equivalents constructed by layer-by-layer cell coating technology.

The aim of this study is to validate an in vitro skin irritation test (SIT) using three-dimensional reconstructed human epidermal (RhE) skin equivalents prepared by layer-by-layer (LbL) method (LbL-3D Skin) in a series of interlaboratory studies. The goal of these validation studies is to evaluate the ability of this in vitro test to reliably discriminate skin irritant from nonirritant chemicals, as defined by OECD and UN GHS. This me-too validation study is to assess the within- and between-laboratory reproducibility, as well as the predictive capacity, of the LbL-3D Skin SIT in accordance with performance standards for OECD TG 439.

[International trend regarding immunotoxicity evaluation with New Approach Methods (NAMs)].

We have ongoing projects that are developing New Approach Methods (NAMs) for systematic toxicology. One NAM is to develop the immunotoxicity evaluation with non-animal test methods for the Organisation for Economic Co-operation and Development (OECD)Test Guideline (TG). The development of this evaluation includes the following steps: 1) adverse outcome pathway (AOP), 2) detailed review paper, 3) test methods based on AOP, 4) validation study of test methods for developing TGs, and 5) integrated approaches to testing and assessment (IATA).

In chemico sequential testing strategy for assessing the photoallegic potential.

Several non-animal testing methods to assess photoallergic potential have been developed so far, while none of them have yet to be validated and regulatory accepted. Currently, some photoreactivity assays such as UV-VIS spectral analysis and ROS assay are generally used for initial photosafety assessments because of their high sensitivity. However, they have a low specificity, generating a high percentage of false positive results, and the development of a follow-up assessment method is desired.

Development of quantitative model of a local lymph node assay for evaluating skin sensitization potency applying machine learning CatBoost.

The estimated concentrations for a stimulation index of 3 (EC3) in murine local lymph node assay (LLNA) is an important quantitative value for determining the strength of skin sensitization to chemicals, including cosmetic ingredients. However, animal testing bans on cosmetics in Europe necessitate the development of alternative testing methods to LLNA. A machine learning-based prediction method can predict complex toxicity risks from multiple variables.

[Current Issues in Photosafety Evaluation].

Phototoxicity is a toxic response elicited by topically applied or systemically administered photoreactive chemicals after exposure to light and can be broadly categorized into photoirritation, photoallergy, photogenotoxicity, and photocarcinogenicity. The need in the 21st century for accurate evaluation of photosafety has led to the publication of a number of guidelines from government agencies in Europe and the U.S.

[Challenge for Adverse Outcome Pathway (AOP)-based Chemical Safety Assessment].

The latest chemical management policies require toxicological evaluation of marketed but untested chemicals. Furthermore, in Europe, for animal welfare reasons sales of cosmetics and raw materials for which animal experiments were conducted were totally banned, in 2013. Responding to these regulatory trends, a strong demand exists to develop new in vitro test methods and to improve in silico prediction models for safety assessments.

Non-animal methods to predict skin sensitization (I): the Cosmetics Europe database.

Cosmetics Europe, the European Trade Association for the cosmetics and personal care industry, is conducting a multi-phase program to develop regulatory accepted, animal-free testing strategies enabling the cosmetics industry to conduct safety assessments. Based on a systematic evaluation of test methods for skin sensitization, five non-animal test methods (DPRA (Direct Peptide Reactivity Assay), KeratinoSens, h-CLAT (human cell line activation test), U-SENS, SENS-IS) were selected for inclusion in a comprehensive database of 128 substances. Existing data were compiled and completed with newly generated data, the latter amounting to one-third of all data.

Non-animal methods to predict skin sensitization (II): an assessment of defined approaches .

Skin sensitization is a toxicity endpoint of widespread concern, for which the mechanistic understanding and concurrent necessity for non-animal testing approaches have evolved to a critical juncture, with many available options for predicting sensitization without using animals. Cosmetics Europe and the National Toxicology Program Interagency Center for the Evaluation of Alternative Toxicological Methods collaborated to analyze the performance of multiple non-animal data integration approaches for the skin sensitization safety assessment of cosmetics ingredients. The Cosmetics Europe Skin Tolerance Task Force (STTF) collected and generated data on 128 substances in multiple in vitro and in chemico skin sensitization assays selected based on a systematic assessment by the STTF.

Development of an artificial neural network model for risk assessment of skin sensitization using human cell line activation test, direct peptide reactivity assay, KeratinoSens™ and in silico structure alert parameter.

It is important to predict the potential of cosmetic ingredients to cause skin sensitization, and in accordance with the European Union cosmetic directive for the replacement of animal tests, several in vitro tests based on the adverse outcome pathway have been developed for hazard identification, such as the direct peptide reactivity assay, KeratinoSens™ and the human cell line activation test. Here, we describe the development of an artificial neural network (ANN) prediction model for skin sensitization risk assessment based on the integrated testing strategy concept, using direct peptide reactivity assay, KeratinoSens™, human cell line activation test and an in silico or structure alert parameter. We first investigated the relationship between published murine local lymph node assay EC3 values, which represent skin sensitization potency, and in vitro test results using a panel of about 134 chemicals for which all the required data were available.

State-of-the-art and new options to assess T cell activation by skin sensitizers: Cosmetics Europe Workshop.

Significant progress has been made in the development and validation of non-animal test methods for skin sensitization assessment. At present, three of the four key events of the Adverse Outcome Pathway (AOP) are assessable by OECD-accepted in vitro methods. The fourth key event describes the immunological response in the draining lymph node where activated dendritic cells present major histocompatibility complex-bound chemically modified peptides to naive T cells, thereby priming the proliferation of antigen-specific T cells.

Chemical photoallergy: photobiochemical mechanisms, classification, and risk assessments.

Chemical photosensitivity can be elicited by exposure of the skin to various pharmaceutical substances, foods, cosmetics and other environmental chemicals, followed by exposure to sunlight. There are at least three types of chemical photosensitivity, i.e.

Comparative study on prediction performance of photosafety testing tools on photoallergens.

Several testing methods have been established to identify potential phototoxins. The present study was undertaken to clarify the predictive ability of in vitro photosafety assays for photoallergenicity. On the basis of animal and/or clinical photosafety information, 23 photoallergens and 7 non-phototoxic/non-photoallergenic chemicals were selected and subjected to UV/VIS spectral analysis, reactive oxygen species (ROS)/micellar ROS (mROS) assays, and 3T3 neutral red uptake phototoxicity testing (3T3 NRU PT).

Development of an in vitro photosensitization test based on changes of cell-surface thiols and amines as biomarkers: the photo-SH/NH₂ test.

As a part of our studies to develop a cell-based in vitro photosensitization assay, we examined whether changes of cell-surface thiols and amines on human monocytic cell line THP-1 could be used to predict photosensitizing potential of chemicals. First, we identified a suitable ultraviolet A (UV-A) irradiation dose to be 5.0 J/cm(2) by investigating the effect of UV-A on the levels of cell-surface thiols and amines in ketoprofen (KP; a representative photoallergen)-treated THP-1 cells.

Bayesian integrated testing strategy (ITS) for skin sensitization potency assessment: a decision support system for quantitative weight of evidence and adaptive testing strategy.

The presented Bayesian network Integrated Testing Strategy (ITS-3) for skin sensitization potency assessment is a decision support system for a risk assessor that provides quantitative weight of evidence, leading to a mechanistically interpretable potency hypothesis, and formulates adaptive testing strategy for a chemical. The system was constructed with an aim to improve precision and accuracy for predicting LLNA potency beyond ITS-2 (Jaworska et al., J Appl Toxicol 33(11):1353-1364, 2013) by improving representation of chemistry and biology.

Development of novel in vitro photosafety assays focused on the Keap1-Nrf2-ARE pathway.

Although photoallergens require UV energy for antigen formation, the subsequent immune response is considered to be the same as in ordinary skin sensitization. Therefore, in vitro tests for skin sensitization should also be applicable for photoallergy testing. In this study, we examined whether activation of the Keap1 (Kelch-like ECH-associated protein 1)-Nrf2 (nuclear factor-erythroid 2-related factor 2)-ARE (antioxidant response element) pathway could be used to assess the photoallergenic potential of chemicals, using the reporter cell line AREc32 or KeratinoSens(TM) .

Alternatives for skin sensitisation: Hazard identification and potency categorisation: Report from an EPAA/CEFIC LRI/Cosmetics Europe cross sector workshop, ECHA Helsinki, April 23rd and 24th 2015.

In the two years since the last workshop report, the environment surrounding the prediction of skin sensitisation hazards has experienced major change. Validated non-animal tests are now OECD Test Guidelines. Accordingly, the recent cross sector workshop focused on how to use in vitro data for regulatory decision-making.

Evaluation of combinations of in vitro sensitization test descriptors for the artificial neural network-based risk assessment model of skin sensitization.

The skin sensitization potential of chemicals has been determined with the use of the murine local lymph node assay (LLNA). However, in recent years public concern about animal welfare has led to a requirement for non-animal risk assessment systems for the prediction of skin sensitization potential, to replace LLNA. Selection of an appropriate in vitro test or in silico model descriptors is critical to obtain good predictive performance.

Test battery with the human cell line activation test, direct peptide reactivity assay and DEREK based on a 139 chemical data set for predicting skin sensitizing potential and potency of chemicals.

To develop a testing strategy incorporating the human cell line activation test (h-CLAT), direct peptide reactivity assay (DPRA) and DEREK, we created an expanded data set of 139 chemicals (102 sensitizers and 37 non-sensitizers) by combining the existing data set of 101 chemicals through the collaborative projects of Japan Cosmetic Industry Association. Of the additional 38 chemicals, 15 chemicals with relatively low water solubility (log Kow > 3.5) were selected to clarify the limitation of testing strategies regarding the lipophilic chemicals.

In silico risk assessment for skin sensitization using artificial neural network analysis.

The sensitizing potential of chemicals is usually identified and characterized using in vivo methods such as the murine local lymph node assay (LLNA). Due to regulatory constraints and ethical concerns, alternatives to animal testing are needed to predict the skin sensitization potential of chemicals. For this purpose, an integrated evaluation system employing multiple in vitro and in silico parameters that reflect different aspects of the sensitization process seems promising.

Assessing skin sensitization hazard in mice and men using non-animal test methods.

Sensitization, the prerequisite event in the development of allergic contact dermatitis, is a key parameter in both hazard and risk assessments. The pathways involved have recently been formally described in the OECD adverse outcome pathway (AOP) for skin sensitization. One single non-animal test method will not be sufficient to fully address this AOP and in many cases the use of a battery of tests will be necessary.