Integration of MUTZ-Langerhans cells into a 3D full-thickness skin equivalent and influences of serum reduction and undefined medium supplements on differentiation.

The MUTZ-3 cell line is a surrogate for Langerhans cells (LCs) employed in New Approach Methodologies for assessing the skin sensitizing potential of chemicals. However, MUTZ-3 cells must first be differentiated to achieve the LC-typical phenotype. As all protocols use high fetal calf serum (FCS) concentrations, we aimed at reducing, or even replacing FCS, while maintaining MUTZ-LC characteristics. Additionally, we assessed the impact of the poorly defined 5637-conditioned medium (5637CM) on MUTZ-LC differentiation. With reducing the FCS content by 75 %, the desired differentiation status was achieved after 7 instead of 14 days, identified by elevated CD207 and CD1a expression. Culture with Ultroser G, a synthetic surrogate for FCS, resulted in an insufficient number of MUTZ-LCs. 5 % FCS-differentiated MUTZ-LCs could be activated with DNCB, an extreme sensitizer, as demonstrated by increased CD83 expression. 5637CM did not affect MUTZ-LC differentiation and is therefore not needed as a supplement. For their intended role in an immunocompetent skin model to assess the sensitizing potential of chemicals, MUTZ-LCs were successfully integrated into the Phenion® Full-Thickness skin model, as demonstrated by CD1a expression. These results are important steps towards medium standardization and the generation of an immunocompetent skin model according to the 3R principles.

Deriving a Continuous Point of Departure for Skin Sensitization Risk Assessment Using a Bayesian Network Model.

Regulations of cosmetic ingredients and products have been the most advanced in embracing new approach methodologies (NAMs). Consequently, the cosmetic industry has assumed a forerunner role in the development and implementation of animal-free next-generation risk assessment (NGRA) that incorporates defined approaches (DAs) to assess the skin sensitization potency of ingredients. A Bayesian network DA predicting four potency categories (SkinSens-BN) was constructed against reference Local Lymph Node Assay data for a total of 297 substances, achieving a predictive performance similar to that of other DAs. With the aim of optimally informing risk assessment with a continuous point of departure (PoD), a weighted sum of the SkinSens-BN probabilities for four potency classes (non-, weak, moderate, and strong/extreme sensitizer) was calculated, using fixed weights based on associated LLNA EC3-values. The approach was promising, e.g., the derived PoDs for substances classified as non-sensitizers did not overlap with any others and 77% of PoDs were similar or more conservative than LLNA EC3. In addition, the predictions were assigned a level of confidence based on the probabilities to inform the evaluation of uncertainty in an NGRA context. In conclusion, the PoD derivation approach can substantially contribute to reliable skin sensitization NGRAs.

Limitations and Modifications of Skin Sensitization NAMs for Testing Inorganic Nanomaterials.

Since 2020, the REACh regulation requires toxicological data on nanoforms of materials, including the assessment of their skin-sensitizing properties. Small molecules' skin sensitization potential can be assessed by new approach methodologies (NAMs) addressing three key events (KE: protein interaction, activation of dendritic cells, and activation of keratinocytes) combined in a defined approach (DA) described in the OECD guideline 497. In the present study, the applicability of three NAMs (DPRA, LuSens, and h-CLAT) to nine materials (eight inorganic nanomaterials (NM) consisting of CeO2, BaSO4, TiO2 or SiO2, and quartz) was evaluated. The NAMs were technically applicable to NM using a specific sample preparation (NANOGENOTOX dispersion protocol) and method modifications to reduce interaction of NM with the photometric and flowcytometric read-outs. The results of the three assays were combined according to the defined approach described in the OECD guideline No. 497; two of the inorganic NM were identified as skin sensitizers. However, data from animal studies (for ZnO, also human data) indicate no skin sensitization potential. The remaining seven test substances were assessed as "inconclusive" because all inorganic NM were outside the domain of the DPRA, and the achievable test concentrations were not sufficiently high according to the current test guidelines of all three NAMs. The use of these NAMs for (inorganic) NM and the relevance of the results in general are challenged in three ways: (i) NAMs need modification to be applicable to insoluble, inorganic matter; (ii) current test guidelines lack adequate concentration metrics and top concentrations achievable for NM; and (iii) NM may not cause skin sensitization by the same molecular and cellular key events as small organic molecules do; in fact, T-cell-mediated hypersensitivity may not be the most relevant reaction of the immune system to NM. We conclude that the NAMs adopted by OECD test guidelines are currently not a good fit for testing inorganic NM.

The role of dendritic cells in the instruction of helper T cells in the allergic march.

Allergy is a complex array of diseases influenced by innate and adaptive immunity, genetic polymorphisms, and environmental triggers. Atopic dermatitis (AD) is a chronic inflammatory skin disease characterized by barrier defects and immune dysregulation, sometimes leading to asthma and food allergies because of the atopic march. During atopic skin inflammation, Langerhans cells and dendritic cells (DCs) in the skin capture and deliver allergen information to local lymph nodes. DCs are essential immune sensors coordinating immune reactions by capturing and presenting antigens to T cells. In the context of allergic responses, DCs play a crucial role in instructing two types of helper T cells - type 2 helper T (Th2) cells and follicular helper T (TFH) cells - in allergic responses and IgE antibody responses. In skin sensitization, the differentiation and function of Th2 cells and TFH cells are influenced by skin-derived factors, including epithelial cytokines, chemokines, and signaling pathways to modify the function of migratory DCs and conventional DCs. In this review, we aim to understand the specific mechanisms involving DCs in allergic responses to provide insights into the pathogenesis of allergic diseases and potential therapeutic strategies.

The dimer effect: A refinement approach towards skin sensitization assessment in-chemico using Amino acid Derivative Reactivity Assay.

Skin sensitization is a key endpoint for safety assessment, especially for cosmetics and personal care products. The adverse outcome pathway for skin sensitization and the chemical and biological events driving the induction of human skin sensitization are now well understood. Several non-animal test methods have been developed to predict sensitizer potential by measuring the impact of chemical sensitizers on these key events. In this work, we have focused on Key Event 1 (the molecular initiating step), which is based on formation of a covalent adduct between skin sensitizers and endogenous proteins and/or peptides in the skin. There exists three in-chemico assays approved by the Organization for Economic Co-operation and Development-(1) Direct Peptide Reactivity Assay (DPRA), (2) Amino Acid Derivative Reactivity Assay (ADRA), and (3) Kinetic Direct Peptide Reactivity Assay (kDPRA) to quantify peptide/amino acid derivative depletion after incubation with test chemicals. However, overestimated depletion of the cysteine-based peptide/amino acid derivatives is known in such assays because of the dimerization of the thiol group. In this present work, we report the synthesis and structural confirmation of the dimer of N-(2-[1-naphthyl]acetyl)-L-cysteine (NAC) from the ADRA assay to allow simultaneous determination of (a) peptide depletion by quantifying NAC monomer and (b) peptide dimerization by quantifying NAC dimer thereby eliminating the overestimation. We present a case study with three chemicals to demonstrate the importance of this approach. Thus, this simultaneous assay gives a more informed view of the peptide reactivity of chemicals to better identify skin sensitizers.

Low-cost quantum mechanical descriptors for data efficient skin sensitization QSAR models.

Quantitative Structure Activity Relationship modelling methodologies need to incorporate relevant mechanistic information to have high predictive performance and validity. Electrophilic reactivity is a common mechanistic feature of skin sensitization endpoints which could be concisely characterized with electronic descriptors which is key to enabling the modelling of small datasets in this domain. However, quantum mechanical methodologies have previously featured high computational costs which would exclude the use of large datasets. Consequently, we investigate the use of electronic descriptors calculated using the Hartree Fock with 3 corrections (Hf-3c) method, a low-cost ab initio methodology that has higher chemical accuracy than previous semiempirical methodologies for modelling in vitro skin sensitization assay outcomes. We also model the Ames assay as a surrogate for determining skin sensitization outcomes. The quantum chemical descriptors calculated using the Hf-3c method with conductor-like polarizable continuum model (CPCM) implicit solvation found improved QSAR model performance for the in vitro Ames (n = 6049, 0.770 AUC), KeratinoSens (n = 164, 0.763 AUC), and Direct Peptide Reactivity Assay (n = 122, 0.750 AUC) datasets, with their combination producing high predictive performance for unseen in vivo Local Lymph Node Assay (n = 86, 0.789 AUC) and Human Repeated Insult Patch Test (n = 86, 0.791 AUC) assay toxicant outcomes.

A rapid spectrophotometric test for assessing skin sensitization potential of chemicals by using N-acetyl-L-cysteine methyl ester in chemico.

During the key event 1 of skin sensitization defined as covalent binding or haptenization of sensitizer to either thiol or amino group of skin proteins, a sensitizer not only covalently binds with skin proteins but also interacts with nucleophilic small molecules such as glutathione (GSH). Although GSH would not be directly associated with skin sensitization, this interaction may be applied for developing an alternative test method simulating key event 1, haptenization. Thus, the aim of the present study was to examine whether N-acetyl-L-cysteine methyl ester (NACME), a thiol-containing compound, was selected as an electron donor to determine whether NACME reacted with sensitizers. Following a reaction of NACME with a sensitizer in a 96-well plate, the remaining NACME was measured spectrophotometrically using 5,5'-dithio-bis-(2-nitrobenzoic acid) (DTNB). Following the optimization of test conditions with two different vehicles, such as acetonitrile (ACN) and dimethyl sulfoxide (DMSO), 64 test chemicals were tested to determine the predictive capacity of current NACME test method. The results obtained showed, the predictive capacity of 94.6% sensitivity, 88.9% specificity, and 92.2% accuracy utilizing DMSO as a vehicle with a cutoff NACME depletion of 5.85%. The three parameters were also over 85% in case of ACN. These values were comparable to or better than other OECD-approved test methods. Data demonstrated that a simple thiol-containing compound NACME might constitute as a reliable candidate for identifying reactive skin sensitizers, and that this method be considered as practical method as a screening tool for assessing a chemical's tendency to initiate skin sensitization.

Health risks from exposure to chemicals in clothing - Non-regulated halogenated aromatic compounds.

The objective of the present study was to investigate some commonly detected halogenated textile pollutants for their bioavailability and hazardous properties. Release into artificial sweat and skin absorption in vitro were examined as well as mutagenic effects by Ames test, and skin-sensitizing properties from a peptide reactivity assay combined with a cell test. All investigated compounds were shown to migrate from the textile into sweat and be absorbed by the skin, although to a different extent. The experimental values for migration were found to be up to 390 times higher compared to literature values. Two of the studied compounds, 2,5-dinitrochlorobenzene and 3,5-dinitrobromobenzene, both exhibited mutagenic effects in the Ames test, while both 2,5-dinitrochlorobenzene and 2,6-dichlorobenzene-1,4-diamine were classified as skin sensitizers. The allergenic reactivity of the latter was found to be due to an oxidized transformation product. Risks for the induction of skin allergy and other non-carcinogenic effects from dermal exposure to the individual compounds were found low, even when considering clothing with the highest reported levels. However, the complex mixtures of chemicals often present in garments may still constitute a health risk, especially when considering the many hours of daily exposure. It is important to further study the toxicity of other frequently occurring chemicals as well as the synergistic effects of chemicals that co-occur in clothing.

Safety assessment of cocamidopropyl betaine, a cosmetic ingredient.

Cocamidopropyl betaine (CAPB) is a surfactant derived from coconut oil that is widely used in cosmetics and personal products for several purposes, such as a surfactant, foam booster, mildness, and viscosity control. Cocamidopropyl betaine is used at concentrations up to 30% in cosmetics. The acute toxicity, skin irritation, eye irritation, skin sensitization, repeated dose toxicity, genotoxicity, carcinogenicity, and phototoxicity of cocamidopropyl betaine were evaluated. Cocamidopropyl betaine was observed to induce mild skin irritation, eye irritation and skin sensitization. The NOAEL of cocamidopropyl betaine was determined to be 250 mg/kg/day based on the results of a 92-day repeated-dose oral toxicity study in rats. The systemic exposure dose of cocamidopropyl betaine was estimated to range from 0.00120 to 0.93195 mg/kg/day when used in cosmetic products. The margin of safety of cocamidopropyl betaine was calculated to be greater than 100 when used at a maximum concentration of 6% in leave-on products and 30% in rinse-off products, suggesting that its use in cosmetic products is safe under current usage conditions.

Deriving a point of departure for assessing the skin sensitization risk of wearable device constituents with in vitro methods.

Wearable devices are in contact with the skin for extended periods. As such, the device constituents should be evaluated for their skin sensitization potential, and a Point of Departure (PoD) should be derived to conduct a proper risk assessment. Without historical in vivo data, the PoD must be derived with New Approach Methods (NAMs). To accomplish this, regression models trained on LLNA data that use data inputs from OECD-validated in vitro tests were used to derive a predicted EC3 value, the LLNA value used to classify skin sensitization potency, for three adhesive monomers (Isobornyl acrylate (IBOA), N, N- Dimethylacrylamide (NNDMA), and Acryloylmorpholine (ACMO) and one dye (Solvent Orange 60 (SO60)). These chemicals can be used as constituents of wearable devices and have been associated with causing allergic contact dermatitis (ACD). Using kinetic DPRA and KeratinoSens™ data, the PoDs obtained with the regression model were 180, 215, 1535, and 8325 µg/cm2 for IBOA, SO60, ACMO, and NNDMA, respectively. The PoDs derived with the regression model using NAMs data will enable a proper skin sensitization risk assessment without using animals.

Allergens grouping names on packaging: Are the substances similar enough to ensure safety? The worst case scenarios question this.

OBJECTIVE: European Commission Regulation (EU) n°2023/1545 introduced the concept of grouping names in the cosmetics sector in July 2023. These groups bring together allergenic substances with the same level of skin sensitization. Their purpose is to lighten the list of ingredients on cosmetic packaging, by grouping together substances deemed to be similar under the same name. As this classification is based on a single toxic effect - skin sensitization - the present study aims to analyse the relevance of these groupings with regard to other toxic effects of substances in the same group. METHODS: This study was carried out by consulting an available database, various reports from 5 committees, 2 books and 5 articles in order to complete the toxicological profile of each substance. Then, in order to highlight any discrepancies within the classification, the worst cases were identified. For this purpose, the data for each substance in a group were compared, and in the event of greater criticality for a toxic effect, this was qualified as a worst case. In addition, similar toxic effects between several substances within the same group were also recorded. The aim of this additional research was to validate the definition of the grouping name and the similarities between substances in the same group. RESULTS: From the 17 grouping names, 5 presented worst cases. Two groups had 2 worst cases and the others only one. In total, from the 7 worst cases detected, 3 were due to the toxic effect "skin irritation". In most cases, the substances in the groupings shared the presence or absence of risk. Only the degree of risk criticality varied. CONCLUSION: Classification by grouping names appears justified regarding the similarities between substances, particularly in terms of skin sensitization. However, the presence of worst cases qualifies it and highlights the importance of being vigilant when assessing the risk of cosmetic products including these grouping names in their list of ingredients.

OBJECTIF: Le règlement (UE) n°2023/1545 de la Commission européenne a introduit la notion de « grouping names ¼ dans le domaine des cosmétiques en juillet 2023. Ces groupes rassemblent des substances allergènes ayant le même niveau de sensibilisation cutanée. Ils ont pour objectif d'alléger la liste des ingrédients figurant sur les emballages des produits cosmétiques, en regroupant sous un même nom des substances jugées similaires. Cette classification étant fondée sur un seul effet toxique ­ la sensibilisation cutanée ­ la présente étude vise à analyser la pertinence de ces regroupements au regard des autres effets toxiques des substances d'un même groupe. MÉTHODES: Cette étude a été réalisée en consultant une base de données disponible, différents rapports de 5 comités, 2 livres et 5 articles afin de compléter le profil toxicologique de chaque substance. Ensuite, afin de mettre en évidence les divergences au sein de la classification, les cas de criticité plus importante ont été identifiés. Pour ce faire, les données de chaque substance d'un groupe ont été comparées, et en cas de criticité supérieure d'un effet toxique, celui­ci a été qualifié de « worst case ¼. En outre, les effets toxiques similaires entre plusieurs substances d'un même groupe ont également été enregistrés. L'objectif de cette recherche complémentaire était de valider la définition du « grouping name ¼ et les similitudes entre les substances d'un même groupe. RÉSULTATS: Sur les 17 « grouping names ¼, 5 présentaient des « worst cases ¼. Deux groupes présentaient deux « worst cases ¼ et les autres un seul. Au total, sur les 7 « worst cases ¼ détectés, 3 étaient dus à l'effet toxique "irritation cutanée". Dans la plupart des cas, les substances des groupes partagent la présence ou l'absence de risque. Seul le degré de criticité du risque variait. CONCLUSION: La classification par « grouping names ¼ semble justifiée au regard des similitudes entre les substances, notamment en termes de sensibilisation cutanée. Cependant, la présence de « worst cases ¼ la nuance et souligne l'importance d'être vigilant lors de l'évaluation du risque des produits cosmétiques incluant ces « grouping names ¼ dans leur liste d'ingrédients.

A Human Skin Explant Test as a Novel In Vitro Assay for the Detection of Skin Sensitization to Aggregated Monoclonal Antibodies.

Introduction: Monoclonal antibodies (mAbs) are important therapeutics. However, the enhanced potential for aggregation has become a critical quality parameter during the production of mAbs. Furthermore, mAb aggregation may also present a potential health risk in a clinical setting during the administration of mAb therapeutics to patients. While the extent of immunotoxicity in patient populations is uncertain, reports show it can lead to immune responses via cell activation and cytokine release. In this study, an autologous in vitro skin test designed to predict adverse immune events, including skin sensitization, was used as a novel assay for the assessment of immunotoxicity caused by mAb aggregation. Material and Methods: Aggregation of mAbs was induced by a heat stress protocol, followed by characterization of protein content by analytical ultra-centrifugation and transmission electron microscopy, revealing a 4% aggregation level of total protein content. Immunotoxicity and potential skin sensitization caused by the aggregates, were then tested in a skin explant assay. Results: Aggregated Herceptin and Rituximab caused skin sensitization, as shown by histopathological damage (grade II-III positive response) together with positive staining for Heat Shock Protein 70 (HSP70). Changes in T cell proliferation were not observed. Cytokine analysis revealed a significant increase of IL-10 for the most extreme condition of aggregation (65 °C at pH3) and a trend for an overall increase of IFN-γ, especially in response to Rituximab. Conclusions: The skin explant assay demonstrated that aggregated mAbs showed adverse immune reactions, as demonstrated as skin sensitization, with histopathological grades II-III. The assay may, therefore, be a novel tool for assessing immunotoxicity and skin sensitization caused by mAb aggregation.

Assessment of skin sensitization potential of zinc oxide, aluminum oxide, manganese oxide, and copper oxide nanoparticles through the local lymph node assay: 5-bromo-deoxyuridine flow cytometry method.

The advent of nanotechnology has significantly spurred the utilization of nanoparticles (NPs) across diverse sectors encompassing industry, agriculture, engineering, cosmetics, and medicine. Metallic oxides including zinc oxide (ZnO), copper oxide (CuO), manganese oxide (Mn2O3), and aluminum oxide (Al2O3), in their NP forms, have become prevalent in cosmetics and various dermal products. Despite the expanding consideration of these compounds for dermal applications, their potential for initiating skin sensitization (SS) has not been comprehensively examined. An in vivo assay, local lymph node assay: 5-bromo-2-deoxyuridine-flow cytometry method (LLNA: BrdU-FCM) recognized as an alternative testing method for screening SS potential was used to address these issues. Following the OECD TG 442B guidelines, NPs suspensions smaller than 50 nm size were prepared for ZnO and Al2O3 at concentrations of 10, 25, and 50%, and Mn2O3 and CuO at concentrations of 5, 10, and 25%, and applied to the dorsum of each ear of female BALB/c mice on a daily basis for 3 consecutive days. Regarding the prediction of test substance to skin sensitizer if sensitization index (SI)≥2.7, all 4 NPs were classified as non-sensitizing. The SI values were below 2.06, 1.33, 1.42, and 0.99 for ZnO, Al2O3, Mn2O3, and CuO, respectively, at all test concentrations. Although data presented were negative with respect to adverse SS potential for these 4 NPs, further confirmatory tests addressing other key events associated with SS adverse outcome pathway need to be carried out to arrive at an acceptable conclusion on the skin safety for both cosmetic and dermal applications.

Use of human predictive patch test (HPPT) data for the classification of skin sensitization hazard and potency.

Since the 1940s, patch tests in healthy volunteers (Human Predictive Patch Tests, HPPTs) have been used to identify chemicals that cause skin sensitization in humans. Recently, we reported the results of a major curation effort to support the development of OECD Guideline 497 on Defined Approaches (DAs) for skin sensitization (OECD in Guideline No. 497: Defined Approaches on Skin Sensitisation, 2021a. https://doi.org/10.1787/b92879a4-en ). In the course of this work, we compiled and published a database of 2277 HPPT results for 1366 unique test substances (Strickland et al. in Arch Toxicol 97:2825-2837, 2023. https://doi.org/10.1007/s00204-023-03530-3 ). Here we report a detailed analysis of the value of HPPT data for classification of chemicals as skin sensitizers under the United Nations' Globally Harmonized System of Classification and Labelling of Chemicals (GHS). As a result, we propose the dose per skin area (DSA) used for classification by the GHS to be replaced by or complemented with a dose descriptor that may better reflect sensitization incidence [e.g., the DSA causing induction of sensitization in one individual (DSA1+) or the DSA leading to an incidence of induction in 5% of the tested individuals (DSA05)]. We also propose standardized concepts and workflows for assessing individual HPPT results, for integrating multiple HPPT results and for using them in concert with Local Lymph Node Assay (LLNA) data in a weight of evidence (WoE) assessment. Overall, our findings show that HPPT results are often not sufficient for deriving unambiguous classifications on their own. However, where they are, the resulting classifications are reliable and reproducible and can be integrated well with those from other skin sensitization data, such as the LLNA.

A convenient spectrophotometric test for screening skin-sensitizing chemicals using reactivity with glutathione in chemico.

To initiate skin sensitization, haptens react with endogenous proteins. During this process, skin sensitizers react with small endogenous molecules containing thiol or amino groups. In this study, a simple spectrophotometric method to identify skin sensitizers in chemico was developed using the reactivity of glutathione (GSH) with test chemicals in a 96-well plate. To quantitate the remaining GSH following the reaction with a skin sensitizer, 5,5'-dithiobis-(2-nitrobenzoic acid) (DTNB) was employed. The optimized experimental conditions included the pH- and time-dependent stability of GSH, stability of derivatized products of GSH with optimal concentration and incubation time of DTNB, incubation time of GSH with the test chemicals, and molar ratios of GSH to the test chemicals. With the optimized conditions with both acetonitrile and DMSO as vehicles and incubation of GSH with test chemicals in 1:10 and 1:15 ratios for 24 h at 4 °C, 23 skin sensitizers and 23 non-sensitizers, based on the local lymph node assay, were tested to determine the predictive capacity of individual conditions. The best result showed a predictive capacity of 95.2% sensitivity, 91.3% specificity, and 93.2% accuracy, with 93.2% consistency in three trials, when 5.8% depletion was used as a cut-off value in 1:10 of GSH:test chemicals in DMSO. It would be an economic and useful screening tool for determining the skin sensitization potential of small molecules, because the present method employs simple endogenous GSH as an electron donor for sensitizers with a spectrophotometric detection system in 96-well plates, and because the method requires neither experimental animals nor cell cultures. Supplementary Information: The online version contains supplementary material available at 10.1007/s43188-023-00218-9.

Use of in vitro methods combined with in silico analysis to identify potential skin sensitizers in the Tox21 10K compound library.

Introduction: Skin sensitization, which leads to allergic contact dermatitis, is a key toxicological endpoint with high occupational and consumer prevalence. This study optimized several in vitro assays listed in OECD skin sensitization test guidelines for use on a quantitative high-throughput screening (qHTS) platform and performed in silico model predictions to assess the skin sensitization potential of prioritized compounds from the Tox21 10K compound library. Methods: First, we screened the entire Tox21 10K compound library using a qHTS KeratinoSensTM (KS) assay and built a quantitative structure-activity relationship (QSAR) model based on the KS results. From the qHTS KS screening results, we prioritized 288 compounds to cover a wide range of structural chemotypes and tested them in the solid phase extraction-tandem mass spectrometry (SPE-MS/MS) direct peptide reactivity assay (DPRA), IL-8 homogeneous time-resolved fluorescence (HTRF) assay, CD86 and CD54 surface expression in THP1 cells, and predicted in silico sensitization potential using the OECD QSAR Toolbox (v4.5). Results: Interpreting tiered qHTS datasets using a defined approach showed the effectiveness and efficiency of in vitro methods. We selected structural chemotypes to present this diverse chemical collection and to explore previously unidentified structural contributions to sensitization potential. Discussion: Here, we provide a skin sensitization dataset of unprecedented size, along with associated tools, and analysis designed to support chemical assessments.

Evaluation of the applicability of GARDskin to predict skin sensitizers in extracts from medical device materials.

Biocompatibility testing of medical devices is governed by the ISO 10993 series of standards and includes evaluation of skin sensitization potential of the final product. A majority of all medical devices are tested using in vivo methods, largely due to the lack of in vitro methods validated within the applicability domain of solid materials. The GARDskin method for assessment of chemical skin sensitizers is a validated method included in the OECD Test Guideline 442E, based on evaluation of transcriptional patterns of an endpoint-specific genomic biomarker signature in a dendritic cell-like cell, following test chemical exposure. The current study aimed to evaluate the applicability of GARDskin for the purpose of testing solid materials by incorporation of extraction procedures described in ISO 10993-12:2021, as well as to demonstrate the functionality of the proposed protocols, by testing of custom-made materials spiked with sensitizing agents. It was shown that GARDskin is compatible with both polar and non-polar extraction vehicles frequently used for the purpose of medical device biological testing. Further, exploring three different material types spiked with up to four different sensitizing agents, as well as three unspiked control materials and commercial reference products, it was shown that the method correctly classified all evaluated test materials. Taken together, the data presented suggest that GARDskin may constitute a valid alternative to in vivo experimentation for the purpose of skin sensitization assessment of medical devices.

Results of patch testing with five fragrance materials hitherto not tested: A dose-finding study in the clinical population.

BACKGROUND: Quantitative risk assessment (QRA) for skin sensitization is used to derive safe use levels of sensitising fragrance ingredients in products. Post-marketing surveillance of the prevalence of contact allergy to these ingredients provides relevant data to help evaluate the performance of these measures. OBJECTIVES: To determine a suitable patch test concentration for five fragrance materials that had hitherto not been tested on a regular basis. These concentrations are then to be used in a surveillance study with patch testing consecutive patients over an extended monitoring period. MATERIALS AND METHODS: Furaneol, CAS.3658-77-3; trans-2-hexenal, CAS.6728-26-3; 4,8-dimethyl-4,9-decadienal, CAS.71077-31-1; longifolene, CAS.475-20-7; benzaldehyde, CAS.10052-7, were patch tested with other fragrance allergens in four clinics. Patch testing was conducted in three rounds, starting with the lowest concentrations of the five ingredients. The doses were increased in the subsequent rounds if no late-appearing positive reactions and virtually no irritant reactions were reported. RESULTS: Overall, 373 patients were tested. No positive allergic reaction was reported to the five ingredients. Patch test results of other fragrance allergens are reported. CONCLUSIONS: The highest test concentrations are each considered safe for patch testing consecutive patients. Further surveillance based on these preparations will evaluate the hypothesis that QRA-driven consumer product levels of these fragrances can prevent sensitization.

Development of a hierarchical support vector regression-based in silico model for the prediction of the cysteine depletion in DPRA.

Topical and transdermal treatments have been dramatically growing recently and it is crucial to consider skin sensitization during the drug discovery and development process for these administration routes. Various tests, including animal and non-animal approaches, have been devised to assess the potential for skin sensitization. Furthermore, numerous in silico models have been created, providing swift and cost-effective alternatives to traditional methods such as in vivo, in vitro, and in chemico methods for categorizing compounds. In this study, a quantitative structure-activity relationship (QSAR) model was developed using the innovative hierarchical support vector regression (HSVR) scheme. The aim was to quantitatively predict the potential for skin sensitization by analyzing the percent of cysteine depletion in Direct Peptide Reactivity Assay (DPRA). The results demonstrated accurate, consistent, and robust predictions in the training set, test set, and outlier set. Consequently, this model can be employed to estimate skin sensitization potential of novel or virtual compounds.

The skin sensitization adverse outcome pathway: exploring the role of mechanistic understanding for higher tier risk assessment.

For over a decade, the skin sensitization Adverse Outcome Pathway (AOP) has served as a useful framework for development of novel in chemico and in vitro assays for use in skin sensitization hazard and risk assessment. Since its establishment, the AOP framework further fueled the existing efforts in new assay development and stimulated a plethora of activities with particular focus on validation, reproducibility and interpretation of individual assays and combination of assay outputs for use in hazard/risk assessment. In parallel, research efforts have also accelerated in pace, providing new molecular and dynamic insight into key events leading to sensitization. In light of novel hypotheses emerging from over a decade of focused research effort, mechanistic evidence relating to the key events in the skin sensitization AOP may complement the tools currently used in risk assessment. We reviewed the recent advances unraveling the complexity of molecular events in sensitization and signpost the most promising avenues for further exploration and development of useful assays.