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.

Exploration of the GARDskin applicability domain: Indirectly acting haptens, hydrophobic substances and UVCBs

Joint publication with the Lubrizol Corporation

ALTEX – Alternatives to animal experimentation, published April 21, 2022, accepted manuscript, https://doi.org/10.14573/altex.2201281

Forreryd, A., Gradin, R., Humfrey, C., Sweet, L. and Johansson, H.

Abstract

Hazard assessments of skin sensitizers are increasingly being performed using new approach methodologies (NAMs), with several in chemico, in vitro and most recently also defined approaches (DAs) being accepted for regulatory use. However, keeping track of potential limitations of each method in order to define applicability domains remains a crucial component to ensure adequate predictivity as well as facilitating the appropriate selection of method(s) for each hazard assessment task. The objective of this report is to share test results generated with the GARD™skin assay on chemicals that have traditionally been considered as difficult to test in some of the conventional in vitro and in chemico OECD Test Guidelines for skin sensitization. Such compounds may include, for example, indirectly acting haptens, hydrophobic substances, and substances of unknown variable composition or biological substances (UVCBs). Based on the results of this study, the sensitivity for prediction of skin sensitizing hazard of indirectly acting haptenswas92.4%and 87.5%, when compared with LLNA(n=25)and human data(n=8), respectively. Similarly, the sensitivity for prediction of skin sensitizing hazard of hydrophobic substances was 85.1% and 100%, when compared with LLNA(n=24)and human data(n=9), respectively. Lastly, a case study involving the assessment of a set of hydrophobic UVCBs(n=7) resulted in a sensitivity of 100, as compared to available reference data. Thus, it was concluded that these data provide support for the inclusion of such chemistries in the GARD™skin applicability domain, without an increased risk of false negative classifications.

Key words: GARD, GARDskin, skin sensitization, applicability domain, difficult to test substances, Indirectly acting haptens, hydrophobic substances, UVCBs

 

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SenzaGen’s Newsletter April 2022

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SOT 2022 Recap: New GARD® data and expanded in vitro toxicology services

We had a fantastic time at this year’s SOT live meeting, reuniting with all our colleagues in the area of in vitro toxicology. Together with guest speakers from Takasago and Sonova, we presented new GARD data on agrochemical formulations, fragrance materials, medical devices and metals. We also introduced how in vitro methods can be used as a part of the biological evaluation of medical devices.  



Exhibitor Hosted Session: New GARD® data on agrochemical formulations, fragrance materials, medical devices and metals

Together with guest speakers from Takasago and Sonova, our Scientific Liaison Dr Andy Forreryd presented the latest user cases where the in vitro GARD assays have been used for skin sensitization testing with a focus on the quantitative potency assessment.

Posters by SenzaGen and jointly with Sonova, Risk Science Consortium

🎫 P460 | Applicability domain of GARDskin | Request a copy
The GARDskin assay: Investigation of the applicability domain of indirectly acting haptens

🎫 P850 | Joint poster with Sonova | Request a copy
In vitro method for quantitative potency assessment of skin sensitizers during development of novel materials for intended use in medical devices

🎫 P846 | Joint poster with Risk Science Consortium | Request a copy
Ability of the GARD assay to replace the GPMT and the LLNA for assessment of the skin sensitization potential of medical devices

Highlights at our booth

Many visited our booth and discussed with our team to learn more about GARD and our expanded in vitro toxicity testing services. Our Medical Device and ISO expert Dr Rose-Marie Jenvert also introduced how we can guide our customers through the comprehensive process of biological evaluation of medical devices.

 

SenzaGen’s Newsletter Dec 2021

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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.

A big step for non-animal methods in skin sensitization testing of medical devices

The new standard for skin sensitization ISO 10993-10 is freshly published and now includes in vitro methods – a big step for non-animal methods in skin sensitization testing of medical devices!

Our GARD assay is included in the standard, can be used with both saline and oil as extraction vehicles and provides human relevant results.

Learn how GARD can be used in your risk assessment

To learn how GARDskin Medical Device can be used in your risk assessment, contact our medical device and ISO expert Rose-Marie Jenvert, PhD on LinkedIn or mail: rose-marie.jenvert@senzagen.com.

Link to the standard: ISO 10993-10:2021: Biological evaluation of medical devices — Part 10: Tests for skin sensitization

SenzaGen and VitroScreen join forces

SenzaGen and VitroScreen have decided to join forces. The highly accurate and broadly applicable skin sensitization test platform GARD will be joined by a recognized pre-clinical CRO and leading in vitro research laboratory.

By joining forces, SenzaGen and VitroScreen will be able to combine their respective knowledge and expertise in novel and advanced biological systems, genomics, proteomics and machine learning to create innovative tools to help the industry transition to animal free testing with high performance human-based technology. SenzaGen and VitroScreen will provide a broadened in vitro service portfolio ranging from in vitro toxicology to preclinical efficacy testing.

About VitroScreen
For more than 20 years, VitroScreen has been leading innovation in pre-clinical testing based exclusively 3D human advanced tissue models offering their services to customers in the pharmaceutical, medical device, cosmetic, chemical, agrochemical and nutritional industries as more biologically relevant alternatives to animal testing. Based in Milan, Italy, VitroScreen provides pre-clinical research services organized in the following business units:

  • GLP certified facility for in vitro regulatory toxicology.
  • Pre-clinical in vitro efficacy testing platform on 3D advanced human tissue models.
  • In vitro Innovation Center with the VitroScreen ORA™ platform for the production of spheroids and organoids and microbiome research.
  • In vitro Consulting unit providing advice on regulatory in vitro toxicology strategies.

Learn more about VitroScreen at www.vitroscreen.com.

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.

Quantitative assessment of sensitizing potency using a dose-response adaptation of GARDskin

Nature Scientific Reports 11, 18904 (2021), https://doi.org/10.1038/s41598-021-98247-7

Robin Gradin, Andy Forreryd, Ulrika Mattson, Anders Jerre, Henrik Johansson

Abstract

Hundreds of chemicals have been identified as skin sensitizers. These are chemicals that possess the ability to induce hypersensitivity reactions in humans, giving rise to a condition termed allergic contact dermatitis. The capacity to limit hazardous exposure to such chemicals depends upon the ability to accurately identify and characterize their skin sensitizing potency. This has traditionally been accomplished using animal models, but their widespread use offers challenges from both an ethical and a scientific perspective. Comprehensive efforts have been made by the scientific community to develop new approach methodologies (NAMs) capable of replacing in vivo assays, which have successfully yielded several methods that can identify skin sensitizers. However, there is still a lack of new approaches that can effectively measure skin sensitizing potency. We present a novel methodology for quantitative assessment of skin sensitizing potency, which is founded on the already established protocols of the GARDskin assay. This approach analyses dose-response relationships in the GARDskin assay to identify chemical-specific concentrations that are sufficient to induce a positive response in the assay. We here compare results for 22 skin sensitizers analyzed using this method with both human and LLNA potency reference data and show that the results correlate strongly and significantly with both metrics (rLLNA = 0.81, p = 9.1 × 10–5; rHuman = 0.74, p = 1.5 × 10–3).

In conclusion, the results suggest that the proposed GARDskin dose-response methodology provides a novel non-animal approach for quantitative potency assessment, which could represent an important step towards reducing the need for in vivo experiments.

 

Key words: GARD, GARDskin, GARDskin Dose-Response, in vitro, sensitization, potency, chemical sensitizers, quantitative risk assessment

 

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