Tag Archive for: GARDskin

Assessing the Utility of the Genomic Allergen Rapid Detection (GARDskin) Assay to Detect Dermal Sensitization Potential in UVCBs and Formulated Lubricant Products

Presented at Eurotox 2023

T. Lindberg2, A.Greminger1 , K. Goyak1, O. Larne2, R. Gradin2, and A. Forreryd2
1ExxonMobil Biomedical Sciences Inc., Annandale, NJ; and 2SenzaGen AB, Lund, Sweden

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Summary

  • GARD®skin is considered to provide useful information in an overall weight of evidence assessment for difficult to test materials (mixtures, UVCBs) with challenging physical chemical properties.
  • The accuracy for prediction of skin sensitization hazard ranged from 66% for formulated lubricants/greases to 100% for synthetic base oils compared to expected outcomes based on reference data.

Abstract

Advances in new approach methods and their combinations into defined approaches can provide clarity and confidence in concluding on skin sensitization potential. However, challenges remain in utilizing these approaches for difficult to test materials such as those with challenging physical chemical properties (low water solubility, hydrophobic substances) or complex compositions like Unknown or Variable Composition Complex reaction products or Biological Materials (UVCBs) and formulated mixtures. The previously developed available non-animal test methods for skin sensitization based on key-events of the adverse outcome pathway (AOP) have clearly defined requirements for test material properties that impact feasibility or confound reliance on negative results particularly for difficult to test materials and impedes the application of defined approaches to conclude on skin sensitization hazard. A set of difficult to test materials were evaluated in the recently validated GARDskin assay since it offered advantages such as a broader applicability domain, availability of additional validated test solvents for poorly soluble materials and provides mechanistically relevant information on key events from across the skin sensitization AOP. The aim of the study was to evaluate the accuracy of the GARDskin assay for a set of synthetic base oils (UVCBs), lubricant additives (UVCBs/poorly soluble substances) and fully formulated lubricants/greases (mixtures) as well as to provide additional information to assist in a weight of evidence determination given that several of the test materials had borderline or conflicting data from other key events within the skin sensitization AOP. All test items were adequately solubilized in one of the following solvents, Ethanol (0.1% final), DMSO (0.25% or 0.1% final), or Xylenes (0.1% final). SenzaCells were incubated in triplicate under standard conditions with the test items at a max concentration of 500uM for those with a known molecular weight or 100 ppm (w/v) for those without a known molecular weight. Following cell stimulations, RNA was isolated and endpoint measurements were performed using the GARDskin genomic profile signature. Based on the results of this study, the accuracy for prediction of skin sensitization hazard was 100% for synthetic base oils (n=4), 83% for lubricant additives (n=6), and 66% for formulated lubricants/greases (n=6) compared to expected outcomes based on available reference data. In some cases, the available reference data was borderline or considered to have low confidence due to confounding factors such as irritation, and nonmonotonic dose responses impacting the accuracy determination when compared one to one with either animal or human data. However, the GARDskin assay is considered to provide useful insight into the overall weight of evidence for difficult to test materials with conflicting datasets as it provides an additional profile of bioactivity across the skin sensitization adverse outcome pathway. 

 

Assessing the Utility of the Genomic Allergen Rapid Detection (GARDskin) Assay to Detect Dermal Sensitization Potential in UVCBs and Formulated Lubricant Products

Presented at SOT 2023

A. Greminger1, K. Goyak1, T. Lindberg2, O. Larne2 , R. Gradin2, and A. Forreryd2
1ExxonMobil Biomedical Sciences Inc., Annandale, NJ; and 2SenzaGen AB, Lund, Sweden

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Summary

  • GARD®skin is considered to provide useful information in an overall weight of evidence assessment for difficult to test materials (mixtures, UVCBs) with challenging physical chemical properties.
  • The accuracy for prediction of skin sensitization hazard ranged from 66% for formulated lubricants/greases to 100% for synthetic base oils compared to expected outcomes based on reference data.

Abstract

Advances in new approach methods and their combinations into defined approaches can provide clarity and confidence in concluding on skin sensitization potential. However, challenges remain in utilizing these approaches for difficult to test materials such as those with challenging physical chemical properties (low water solubility, hydrophobic substances) or complex compositions like Unknown or Variable Composition Complex reaction products or Biological Materials (UVCBs) and formulated mixtures. The previously developed available non-animal test methods for skin sensitization based on key-events of the adverse outcome pathway (AOP) have clearly defined requirements for test material properties that impact feasibility or confound reliance on negative results particularly for difficult to test materials and impedes the application of defined approaches to conclude on skin sensitization hazard. A set of difficult to test materials were evaluated in the recently validated GARDskin assay since it offered advantages such as a broader applicability domain, availability of additional validated test solvents for poorly soluble materials and provides mechanistically relevant information on key events from across the skin sensitization AOP. The aim of the study was to evaluate the accuracy of the GARDskin assay for a set of synthetic base oils (UVCBs), lubricant additives (UVCBs/poorly soluble substances) and fully formulated lubricants/greases (mixtures) as well as to provide additional information to assist in a weight of evidence determination given that several of the test materials had borderline or conflicting data from other key events within the skin sensitization AOP. All test items were adequately solubilized in one of the following solvents, Ethanol (0.1% final), DMSO (0.25% or 0.1% final), or Xylenes (0.1% final). SenzaCells were incubated in triplicate under standard conditions with the test items at a max concentration of 500uM for those with a known molecular weight or 100 ppm (w/v) for those without a known molecular weight. Following cell stimulations, RNA was isolated and endpoint measurements were performed using the GARDskin genomic profile signature. Based on the results of this study, the accuracy for prediction of skin sensitization hazard was 100% for synthetic base oils (n=4), 83% for lubricant additives (n=6), and 66% for formulated lubricants/greases (n=6) compared to expected outcomes based on available reference data. In some cases, the available reference data was borderline or considered to have low confidence due to confounding factors such as irritation, and nonmonotonic dose responses impacting the accuracy determination when compared one to one with either animal or human data. However, the GARDskin assay is considered to provide useful insight into the overall weight of evidence for difficult to test materials with conflicting datasets as it provides an additional profile of bioactivity across the skin sensitization adverse outcome pathway. 

 

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.

Webcast: Positive scientific ESAC opinion on GARD®skin – paving the way for OECD validation and opens new commercial opportunities

CEO Axel Sjöblad and Chief Scientist Henrik Johansson comment on the recently published ESAC opinion.

 

Tag Archive for: GARDskin

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