Tag Archive for: in vitro

Next Generation Risk Assessment (NGRA) using NAMs for skin sensitization: Reproducibility and precision of the GARDskin Dose-Response assay for PoD determination of fragrance chemicals.

Presented at ASCCT 2022

Next Generation Risk Assessment (NGRA) using NAMs for skin sensitization: Reproducibility and precision of the GARDskin Dose-Response assay for PoD determination of fragrance chemicals.

Andy Forreryd1, Shashi Donthamsetty2, Paul Sterchele2, Xiao Huang2, Gregory Ladics2, Mihwa Na3, Isabelle Lee3, Anne Marie Api3, Robin Gradin1, Henrik Johansson1
1SenzaGen, Lund, Sweden , 2International Flavors & Fragrances, Hazlet, NJ, USA, 3Research Institute for Fragrance Materials. Woodcliff lake, NJ, USA

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Conclusion

  • GARD®skin Dose-Response can be used for continous predictions of skin sensitizing potency.
  • The continous readout from the assay is reproducible and the assay predicts LLNA EC3 and human NESIL values with high correlation to reference benchmark data.
  • The assay provides a nice tool for the fragrance industry to predict the NESIL value which can be used for conducting the quantitative risk assessment for generating the IFRA standard.

Abstract

New Approach Methods (NAMs) for assessment of skin sensitizers have been adopted as Test Guidelines (TGs) by OECD. When combined into Integrated Approaches to Testing and Assessment (IATA) or defined approaches (DA), they provide data supporting hazard classifications and GHS potency subcategorization. However, more granular potency information, preferably on a continuous scale, is needed to derive a point-of-departure (PoD) for Next Generation Risk Assessment (NGRA).

GARDskin was recently adopted into OECD TG 442E to support discrimination of skin sensitizers and non-sensitizers. Continous potency predictions are derived using a modified protocol that incorporates dose-response measurements. Linear regression models have further been developed to predict LLNA EC3 and human NOEL values. The aim of the following study, which represents a cross-sector collaboration was to evaluate precision and reproducibility of the potency predictions from GARDskin Dose-Response in blinded studies.

Preliminary results from estimate of precision (n=36 materials) indicated that GARDskin Dose-Response predicted LLNA EC3/ human NOEL values with median fold-misprediction factors < 3.0 and < 2.0, respectively. Interestingly, LLNA predicted human NOEL with a fold-change > 2 in the same dataset. For reproducibility assessment, test materials (n=11) were evaluated in separate experiments (n=3), which generated highly reproducible results, with an average median range of fold-changes between replicates of 2.5.

Results from this study demonstrate that continous potency predictions from GARDskin Dose-Response are reproducible. Together with performance data, this represents a major step towards establishment of the assay as a relevant source of information to derive a PoD for NGRA, avoiding generation of new animal data.

 

 

 

The use of the GARD®skin Dose-Response assay to assess skin sensitizing potency in developing novel fragrance ingredients

Presented at ASCCT 2022

The use of the GARD®skin Dose-Response assay to assess skin sensitizing potency in developing novel fragrance ingredients

Tim Lindberg1, Christopher Choi2, Ulrika Mattson1 and Satoshi Sasaki3
1SenzaGen, Lund, Sweden , 2Takasago International Corp, Rockleigh NJ, USA ,3Takasago International Corp, Hiratsuka city, Kanagawa, Japan

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Conclusion

The present study aimed at investigating the skin sensitizing potency of two novel fragrances, Fragrance 1 and 2, using three NAMs, the kDPRA, KeratinoSens and GARD®skin Dose-Response assays.

To move away from traditional safety testing, which includes animal studies, there is a paradigm shift towards the use of multiple NAMs in a weight-of-evidence approach when risk assessment of novel fragrance ingredients are conducted. However, the lack of established potency assays puts the alternative methods at a disadvantage as compared to the in vivo counterparts.

  • Fragrance 1 showed similar results across the three NAMs, with the GARD®skin Dose-Response assay predicting the cDV0–value to 18.4 µM, which in turn was used to predict a NESIL-value of 659 µg/cm2.
  • For Fragrance 2, equivocal results were seen, where the kDPRA assay predicting the ingredient not to be a category 1A skin sensitizer while KeratinoSens predicted it as a non-skin sensitizer. GARD®skin Dose-Response predicted the cDV0–value to 296 µM, which was used to predict a NESIL-value of 16600 µg/cm2. Combining the results from all three NAM assays, a confirmatory HRIPT testing concentration was determined for both ingredients, 562.5 µg/cm2 and 15000 µg/cm2 for Fragrance 1 and Fragrance 2, respectively.

In conclusion the data presented here show how the use of the GARD®skin Dose-Response assay in combination with other NAMs can be used as a replacement of animal studies for quantitative risk assessment of novel fragrance materials.

Abstract

Skin sensitization is one of the required endpoints for the development and registration of novel fragrance ingredients. Traditionally, testing has been performed using a combination of in vitro and in vivo assays, but recent developments has shifted the paradigm towards the use of New Approach Methodologies (NAMs), without the need for in vivo methods. However, none of the proposed NAMs are currently validated for continuous potency predictions, which is required for quantitative risk assessments of novel fragrance ingredients.

The GARD®skin assay (OECD TG 442E) is a genomics-based assay for hazard identification of sensitizers. To meet the need for quantitative potency information, GARD®skin Dose-Response has been developed based on the validated protocols of GARD®skin and generates a dose-response curve to identify the lowest concentration of a test compound required to elicit a positive classification (cDV0 value). These values correlate significantly to LLNA EC3 and human NESIL values.

The aim of this study was to investigate the sensitizing potency of two novel fragrance ingredients and to identify predicted non-sensitizing levels. Testing was performed in GARD®skin Dose-Response, with predicted EC3 and NESIL values of 1.93% and 27.8%, and 659µg/cm2 and 16600µg/cm2, for fragrance ingredients 1 and 2, respectively. These results in combination with data from kDPRA, KeratinoSens and in silico read- across, established the concentrations for confirmatory HRIPT testing (562.5µg/cm2 and 15000µg/cm2).

In conclusion, this study demonstrates how GARD®skin Dose-Response combined with other NAMs can be used for risk assessments and to establish a concentration for confirmatory HRIPT testing of novel fragrance ingredients.

 

 

The GARDskin Assay: Investigation of the Applicability Domain of Indirectly Acting Haptens

Presented at the 2022 SOT

Tim Lindberg1, Andy Forreryd1, Robin Gradin1 and Henrik Johansson1
1SenzaGen, Lund, Sweden

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Conclusion

  • The GARD®skin assay can accurately predict indirectly acting haptens and has the capacity to assess both pre- and pro-haptens as skin sensitizers.
  • No increased risk of false negative classifications due to possible limitations in metabolic capacity of the cell system.

Abstract

Hypersensitivity reactions in the skin, clinically manifested as Allergic Contact Dermatitits (ACD), are caused by the ensuing immunological response to low-molecular weight compounds termed skin sensitizers. Such substances, often referred to as haptens, have the inherent property to react with skin proteins and form immune inducing complexes. However, indirectly acting haptens need to be transformed to protein-reactive intermediates either through biotic (pro-hapten) or abiotic (pre-hapten) conversion in order to elicit an immune response.

Conventionally, safety tests of skin sensitizers have been done using animal experiments, but New Approach Methodologies (NAMs) have been developed over the past decades to replace the use of animals in such testing. However, one potential problem faced with the in vitro and in chemico alternatives is the lack of metabolic and chemical activity as compared to an in vivo system, which in turn may lead to false predictions for pre- and pro-haptens.

The GARDskin assay is a next-generation NAM for hazard classification of skin sensitizers. The assay is based on a human dendritic -like cell line and combines genomics and machine learning to achieve a high predictive performance with a large applicability domain. Currently, the method is approaching regulatory acceptance as an OECD test guideline.

The study presented here aimed to explore the applicability domain of the GARDskin assay, specifically the capability to predict indirectly acting haptens. Available data obtained from GARDskin testing of indirectly acting haptens were compiled, resulting in a set of 28 substances. Further subcategorization identified 5 pro-haptens and 11 pre-haptens, while 12 substances were unable to be unambiguously assigned as either exclusively a pro- or a pre-hapten, due to the dual nature of the protein-reactive activity. Skin sensitizing hazard sensitivity of indirectly acting haptens (n=28) was 89% (25/28) while pro-haptens (n=5) and pre-haptens (n=11) were 80% and 100%, respectively. These data support GARDskin applicability in the domain of indirectly acting haptens, demonstrating that the method has the capacity to accurately assess both pre- and pro-haptens.

Ability of the GARDskin assay to Predict Skin Sensitization Response in the Guinea Pig Maximization Test

Joint poster with Risk Science Consortium,
Presented at the 2022 SOT

Rose-Marie Jenvert1, Alexandra Zambriczki Lee2, Ronald P Brown2
1SenzaGen, Lund, Sweden, 2Risk Science Consortium, LLC, Arnold, MD USA

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Conclusion

  • The GARD®skin assay is able to predict skin sensitization potential in humans with a level of accuracy that is equal to or exceeds that of GPMT and the LLNA.
  • As a result, the GARDskin assay serves as a promising alternative to assess the skin sensitization potential of medical devices.

Abstract

The preclinical safety assessment of medical devices typically involves an evaluation of the skin sensitization potential of the device. The GARDskin assay is being proposed as an in vitro alternative to the animal-based tests, Local Lymph Node Assay (LLNA) and Guinea Pig Maximization Test (GPMT), that are typically used to assess the skin sensitization potential of medical devices. The ability of the GARDskin assay to replace LLNA for prediction of skin sensitization response has been evaluated (e.g., Johansson et al., 2019) but since GARDskin has also been proposed as an alternative to the GPMT, it is important to compare the concordance of the prediction of the GARDskin assay with the in vivo response obtained in both of the animal-based tests.  Based on the results of the GARDskin assay for 122 compounds, this in vitro assay shows a high concordance with the predicted results of the LLNA (87.5%); however, the concordance for results obtained in the GPMT is much lower (71.2%). The concordance of the GARDskin assay and the GPMT is impacted by the relatively high number of false positive results (15 out of 73) compared to the false positives seen in the comparison between GARDskin and LLNA (2 out of 80).   The high number of false positives found when comparing the results from GARDskin and the GPMT results from the inaccurate characterization of the human skin sensitization potential of these compounds by the GPMT. Therefore, the low concordance between the GARDskin assay and the GPMT is due largely to inaccurate predictions of human skin sensitization potential by the GPMT and not by shortcomings of the GARDskin assay. Notably, the GARDskin assay (88.7% accuracy) outperforms the GPMT (83.0% accuracy) in the ability to predict the human sensitization response of compounds in this dataset. The results of this project show that the GARDskin assay is able to predict skin sensitization potential with a level of accuracy that is equal to or exceeds that of the currently accepted animal-based tests, suggesting that the GARDskin assay can serve as a promising alternative to the GPMT and the LLNA, and provide a more human relevant result for assessment of the skin sensitization potential of medical devices.

In vitro method for quantitative potency assessment of skin sensitizers during development of novel materials for intended use in medical devices

Joint poster with Sonova,
Presented at the 2022 SOT

Andy Forreryd1, Ulrika T Mattson1, David Waeckerlin2, Karla Lienau2, Robin Gradin1, Rose-Marie Jenvert1
1SenzaGen, Lund, Sweden, 2Sonova AG, Staefa, Switzerland

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Conclusion

The GARD®skin Dose-Response assay can be used as a tool for:

  • quantitative potency information of chemicals that might leach out of materials or medical devices.
  • internal decision-making during development of new materials for use in medical devices.

Abstract

New innovative materials for use in medical devices based on acrylates can bring several advantages such as super-absorbency, transparency, flexibility, toughness and hardness.

The manufacturing of acrylates typically involves using a monomer of either acrylate or methacrylate that is polymerized into the final product. The polymerization or hardening of material makes the monomers inert, however several methodologies can be used for polymerization, and they differ in the degree to which they result in a fully polymerized final product. Some products therefore contain more residual monomers than others and human exposure to these well-known skin sensitizers may increase the risk of skin sensitization and allergic contact dermatitis. To assess the risks resulting from exposure to these chemicals, potentially found in medical device material, it is necessary to accurately identify and characterize their skin sensitizing potential.

The GARDskin (OECD TGP 4.106) assay was initially developed for hazard identification of skin sensitizers. To derive potency information, a modification of the standard GARDskin protocol based on dose-response measurements has been proposed. The readout of the assay is a cDV0 value, which corresponds to the lowest concentration required to exceed a binary classification threshold in GARDskin. This concentration correlates significantly with LLNA EC3 and human NOEL values and linear regression models have been established to exploit these relationships for potency predictions. In this study, we explore the potential to use this novel assay for quantitative potency assessment of two acrylate monomers.

The GARDskin Dose-response assay classified both acrylate monomers as skin sensitizers with predicted LLNA EC3 values and human NOEL values of 0.848% and 22.4%, and 230 µg/cm2and 12200 µg/cm2, resulting in final classifications as a strong to moderate skin sensitizer (HP 2) and a moderate to weak sensitizer (HP 5), respectively. The results agreed with information in the ECHA registration dossiers and gathered human data evidence for the respective monomers, illustrating that GARDskin Dose-Response has the potential to replace the in vivo LLNA method for quantitative potency assessment of potential skin sensitizers during development of novel materials for use in medical devices.

Skin Sensitization Potency Assessments of Fragrance Materials using GARDskin Dose-Response

Joint poster with Research Institute for Fragrance Materials (RIFM),
Presented at the 2021 RIFM annual meeting & 2021 ACT annual meeting 

Mihwa Na, Ulrika Mattson, Robin Gradin, Henrik Johansson, Andy Forreryd, Anne Marie Api, Research Institute for Fragrance Materials, Inc., 50 Tice Boulevard, Woodcliff Lake, NJ, 07677, USA., SenzaGen AB, Lund, Sweden

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Conclusion

  • GARDskin Dose-Response closely approximated the potency categories of 9/12 fragrance materials tested.
  • Based on results from this dataset, GARDskin Dose-Response appears useful for potency assessment for weak sensitizers and may constitute a promising strategy for deriving a point of departure for quantitative risk assessments.

Abstract

Several New Approach Methods for hazard identification of skin sensitizers have been developed and incorporated as OECD Test Guidelines. However, the methods for potency assessment are still lacking. GARDskin (OECD TGP 4.106) was initially developed to identify skin sensitizers by monitoring transcriptional patterns of a biomarker signature in a dendritic like cell line.

The predictive capacity of GARDskin has been demonstrated previously, with 95.8% accuracy, 91.7% positive predictive value, and 100.0% negative predictive value (1 false positive, n=24) (Johansson, Gradin et al. 2019). To derive potency information, a strategy based on dose-response measurements in GARDskin, referred to as the GARDskin Dose-Response assay, has recently been proposed. The readout of the assay corresponds to the lowest concentration required to exceed the binary classification threshold in GARDskin. This concentration correlates with local lymph node assay (LLNA) EC3 and human no observed effect level (NOEL) values and linear regression models have been established to exploit these relationships for potency predictions.

In this blinded study, 12 fragrance materials (10 very weak sensitizers and 2 weak sensitizers) were evaluated in GARDskin Dose-Response. Results were evaluated by comparing predicted values to the reference potency categories. Three of the very weak sensitizers were predicted as non-sensitizers by GARDskin Dose-Response. For the remaining nine materials which were predicted as sensitizers, the predicted EC3 and NOEL values closely approximated the reference data for most materials. Based on results from this dataset, GARDskin Dose-Response appears useful for potency assessment for weak sensitizers and may constitute a promising strategy for deriving a point of departure for quantitative risk assessments.

Assessment of the skin sensitizing potential of pandemic-associated medical devices using the GARDskin Medical Device assay

Joint poster with Essity Hygiene & Health AB,
Presented at the 2021 Eurotox annual meeting

P. Mohlin, A. Forreryd, O. Larne, R.-M. Jenvert, H. Johansson | Essity Hygiene & Health AB, Product Safety, Clinical & Regulatory Affairs, Mölndal, Sweden; SenzaGen AB, 22381 Lund, Sweden

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Conclusion

  • GARDskin is well adapted to risk assess the skin sensitizing potential of medical devices in accordance with ISO 10993.
  • GARDskin Medical Device classified the tested commercially available face masks as non-sensitizers and the nitrile glove as sensitizers.
  • New in vitro technologies, like GARD, is well suited as a routine tool to increase the speed of decision making in extraordinary situations as a pandemic.

Abstract

The current SARS-CoV-2 pandemic have led to an increased use of medical devices such as face masks and nitrile gloves, within occupational groups of medical care as well as within the general population. Consequently, the incidence reports of adverse effects associated with use of such medical devices have increased manyfold, including reports of allergic skin reactions.

The cause of allergic skin reactions, referred to as Allergic Contact Dermatitis (ACD) is the immunological process known as skin sensitization, which is induced by so-called chemical sensitizers. Assessment of skin sensitizing potential of leachables from medical devices is a part of biocompatibility testing of medical devices and is typically performed by use of the Guinea Pig Maximization Test (GPMT), in accordance with the ISO 10993 series of standards. However, recent developments of in vitro assays for assessment of chemical sensitizers motivates the exploration of such methods in the context of rapid pandemic-associated testing.

The GARDskin assay [1] is a next-generation in vitro assay for hazard assessment of skin sensitizers, currently progressing towards regulatory acceptance. The method evaluates the transcriptional patterns of a genomic biomarker signature in a human dendritic-like cell line following exposure, in order to provide hazard assessments of tested substances. The method has been adapted to testing of solid materials from e.g. medical device products, by application of extraction protocols using polar- and non-polar extraction vehicles, in accordance with ISO 10993-12.

Here, we present results from testing of a commonly used face mask and a nitrile glove in the GARDskin Medical Device assay. Results indicate that the face mask does not leach any compounds with skin sensitizing potential (among four different batches tested), while the nitrile glove was classified as a skin sensitizer. These results harmonize with preexisting experience of similar models of nitril gloves, which are known to be associated with adverse skin reactions, potentially induced by skin sensitization. Furthermore, these results may have implications on the continued use of similar medical devices throughout the pandemic and beyond, as the appropriate application and removal of face masks may indeed benefit from the discontinued simultaneous use of nitrile gloves.

Reliable and Truly Animal-Free Skin Sensitization Testing – Adaption of the In vitro GARD™skin to Animal-Free Conditions

Poster presented at the 2021 World Congress on Alternatives and Animal Use in the Life Sciences

Andy Forreryd, Anders Jerre, Fiona Jacobs, Carol Treasure, Henrik Johansson | SenzaGen AB, 22381 Lund, Sweden; XCellR8 Ltd, Techspace One, Sci-Tech Daresbury, Cheshire WA4 4AB, UK

 

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Conclusion

  • Senzacells adapted well to routine culture in human serum with comparable cell growth, morphology, and minimal impact on cellular phenotypes.
  • GARDskin based on HS demonstrated full concordance to protocols based on animal-derived FCS for hazard identification of skin sensitizers.
  • This work represents our ambition to gradually replace all animal derived components with human equivalents to enable completely animal-free skin sensitization testing.

Abstract

A plethora of in vitro approaches for hazard assessment of skin sensitizers have recently been described and demonstrated to exhibit discriminatory properties competitive with those of accepted in vivo methods. However, the majority of these in vitro methods still use animal-derived components such as Foetal Calf Serum (FCS) within their standard protocols, imposing the question whether these methods should truly be considered as animal-free replacements.

Genomic Allergen Rapid Detection – GARD – is a genomics-based in vitro testing platform for assessment of numerous immunotoxicity endpoints. The endpoint-specific classification of skin sensitizers is a well-established application of the platform, referred to as GARDskin (OECD TGP 4.106). The assay is based on a human DC-like cell line (SenzaCells) and utilizes state-of-the-art machine learning to classify chemicals by monitoring the expression of 200 genes involved in cellular pathways associated with skin sensitization. GARDskin is progressing towards regulatory acceptance, and consistently reports accuracies > 90%.

Here, we present an adaption of the GARDskin standard protocol to enable for testing under animal-product-free conditions by replacing animal-based FCS with human derived serum. SenzaCells adapted well to routine culture in the human serum, showing comparable cell viability and growth rates to the animal-based FCS. A phenotypic analysis of common DC maturity markers showed minor changes in cell surface expression of the markers CD14 and CD1a, indicating that serum replacement did not significantly alter the phenotypic characteristics of the cells. Finally, a proficiency set of nine chemicals covering the full range from extreme sensitizers to non-sensitizers were evaluated. The protocol adapted to animal-free conditions showed full concordance to the conventional protocol, correctly classifying all chemicals.

In conclusion, this study demonstrates the potential to perform the GARDskin assay without the use of animal-derived components associated with animal welfare concerns, thus paving the way for truly animal-free and highly accurate hazard testing of skin sensitizers.

Hazard Assessment of Photoallergens Using GARD™skin

Poster presented at the 2021 World Congress on Alternatives and Animal Use in the Life Sciences

Andy Forreryd, Angelica Johansson, Gretchen Ritacco, Anne Marie Api, Henrik Johansson | SenzaGen AB, 22381 Lund, Sweden; Research Institute for Fragrance Materials, Inc., 50 Tice Boulevard, Woodcliff Lake, NJ, 07677, USA.

 

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Conclusion

  • The GARDskin assay for assessment of chemical skin sensitizers can be used to predict photoallergens.
  • By implementing a UV-light exposure step into the conventional GARDskin protocols, photoactivation was achieved. Non-irradiated photoallergens remain accurately classified as non-sensitizers.
  • UV-light exposure did not have an impact on classifications on true non-sensitizing and conventional sensitizing controls.

Abstract

Chemicals of different categories, such as cosmetics and drugs, have the potential to become photoactivated when exposed to UV light, giving rise to otherwise dormant adverse effects such as sensitization. Such chemicals, referred to as photoallergens, causes a Type IV delayed hypersensitivity, typically manifested as allergic contact dermatitis. While in vitro assays for prediction of a chemical’s potential to provoke phototoxicity (photoirritation) have been proposed, there is no recognized assay that specifically predicts photoallergens. Therefore, development of accurate in vitro assays that can detect photoallergens remains a high priority.

The Genomic Allergen Rapid Detection – GARD – platform constitutes a unique framework for classification of numerous immunotoxicity endpoints. The endpoint-specific classification of skin sensitizing chemicals is a well-established application of the platform, referred to as the GARDskin assay. GARDskin utilizes the readout of a genomic biomarker signature of 200 genes, which allows for machine learning-assisted classification of skin sensitizers. The assay is progressing towards regulatory acceptance, and demonstrates high predictive performance.

Here, we present an adaptation of GARD protocols, allowing for assessment of chemical photoallergens. By incorporating UVA exposure during sample preparation, photoactivation of latent photoallergens has been demonstrated. In a first step, protocols were optimized using the photoallergens 6-methylcoumarin and Ketoprofen, exposed to UVA light, both prior to and in association with cellular exposure, along with appropriate radiated/non-radiated controls. Photoallergenicity was accurately predicted in both test chemicals exposed to UVA light, while non-radiated counterparts were accurately classified as non-sensitizers.

In summary, our initial data demonstrates a potential of GARDskin to assess photoallergenicity of chemicals. Further evaluation and optimization of the method are currently in progress, in which an extended panel of fragrances are being studied in a collaboration with the Research Institute for Fragrance Materials (RIFM).

The GARD™potency assay for potency-associated subclassification of chemical skin sensitizers – Rationale, method development and ring trial results of predictive performance and reproducibility

Poster presented at the 2021 World Congress on Alternatives and Animal Use in the Life Sciences

Robin Gradin, Angelica Johansson, Andy Forreryd, Amber Edwards, Veronika Hoepflinge, Florence Burleson, Helge Gehrke, Erwin Roggen, Henrik Johansson| SenzaGen AB, 22381 Lund, Sweden; Burleson Research Technologies, Morrisville, NC 27560, USA; Eurofins BioPharma Product Testing Munich Gmbh, 82152 Planegg, Germany; 43RsMC Aps, 2800 Kongens Lyngby, Denmark

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Conclusion

  • GARDpotency is an assay for sub-categorization of strong sensitizers (CLP sub-category 1A), allowing for discrimination from weak sensitizers (CLP sub-category 1B) and non-sensitizers. The method is based on the GARD platform, combining human immune cells, a genomic biomarker readout and machine learning-assisted classifications.
  • Sequential combination of GARDskin and GARDpotency forms the GARD Defined Approach, for complete hazard and risk assessment of skin sensitizers into three categories (CLP 1A, CLP 1B, non-sensitizers).
  • A blinded ring trial, comprising 28 chemicals, demonstrated that GARDpotency is functional and reproducible, with an accumulated predictive accuracy of 91% across three laboratories. In the same dataset, the GARD Defined Approach classifies chemicals into three categories with 86% accuracy.

Abstract

The advancement of non-animal approaches for hazard assessment of skin sensitizers have generated a variety of alternative assays with discriminatory properties comparable with those of accepted in vivo methods. However, hazard identification is rarely sufficient and information permitting the relative ranking of chemicals’ skin sensitization potency is desired. For example, the globally harmonized system of classification and labelling of chemicals (GHS/CLP) extends the binary hazard assessment with a qualitative subcategorization to distinguish between weak and strong skin sensitizers.

Though substantial efforts have been made towards developing alternative methods for potency assessment, none have gained regulatory acceptance, emphasizing that continued development of improved alternative assays remains a high priority.

The genomic allergen rapid detection (GARD) is an in vitro testing platform for assessment and characterization of chemical sensitizers, based on evaluation of transcriptional patterns of endpoint-specific genomic biomarker signatures in a human dendritic-like cell line following chemical exposure, in order to provide machine learning-assisted classifications of tested substances. The GARDskin assay was recently subjected to a formal validation procedure (OECD TGP 4.106) and reported a reproducibility between laboratories of 92%, as well as a predictive accuracy of 94%, for sensitization hazard assessment.

Here, we present the implementation of the related GARD application GARDpotency, for potency-associated subcategorization of chemical sensitizers. Following prediction model establishment, the functionality of the assay was validated in a blinded ring-trial, in accordance with OECD-guidance documents, by assessing predictive performance and reproducibility. It was found that the assay is functional and predictive, with an estimated cumulative accuracy of 88% across three laboratories and nine independent experiments. The within-laboratory reproducibility measures ranged between 63-89%, and the between-laboratory reproducibility was estimated to 61%. In conclusion, the in vitro GARDpotency assay constitute a standardized, functional assay, which could be a valuable tool for hazard characterization of skin sensitizer potency.

Tag Archive for: in vitro

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