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
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
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
Full article
Article on line with open access
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
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.
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).
