Case studies on the use of GARD®skin Medical Device for in vitro skin sensitization assessment
Presented at SOT and Eurotox 2024
Download a copy in PDF
Conclusion
In conclusion, GARDskin Medical Device can detect signals from diluted and complex extracts of solid devices, with a sensitivity superior to animal methods.
This may significantly reduce the need of animal studies, improve the safety of the final product, and avoid potential costly late-stage failures.
Abstract
The recent advancements in New Approach Methodologies enables the use of in vitro method for skin sensitization assessment as part of the biocompatibility testing for medical devices, which is conventionally tested in vivo. GARDskin OECD TG 442E is included in ISO 10993-10:2021 as the only OECD validated in vitro assay that is compatible with both polar and non-polar extraction vehicles, in line with ISO 10993-12:2021. GARDskin Medical Device is an adaptation of the GARDskin assay, including a pre-sample treatment procedure where solid devices are extracted using both polar and non-polar vehicles.
The aim of this study is to demonstrate the benefits of using GARD for early-stage screening of materials intended for use in medical devices for assessing their skin sensitization potential. Results from two case studies were summarized in which GARDskin Medical Device was used for skin sensitization assessment. The first case study describes the testing of an acrylic-based device with a coating consisting of a UV-cured lacquer, where chemical analysis indicated the potential for skin sensitization. The second case study describes the testing of a polymeric material consisting of Cellulose-Acetate Propionate (CAP) with a plasticizer (Triethylene glycol bis (2-ethylhexanoate), CAS# 94-28-0), with contradictive existing in vivo (negative) and in vitro (positive) data.
In the first case study, the acrylic-based device induced a positive response in both polar and non-polar vehicles in GARDskin Medical Device and was thus classified as a skin sensitizer. It was hypothesized that the positive results may be due to inadequate curing of the lacquer within cavitary structures of the devices, where UV light exposure was insufficient. To confirm the hypothesis, follow-up testing was performed on an identical device, but without cavities, which was classified as non-sensitizer. In vivo data confirmed the outcome of the in vitro assay. Consequently, a modification was made in the manufacturing process to prevent the presence of lacquer in cavitary structures of the device.
In the second case study, the CAP material was positive in the non-polar vehicle and was thus labelled as a skin sensitizer. The plasticizer was identified as a potential culprit, considering a borderline negative result in LLNA (SI=2.97) and reported positive clinical data. To support the hypothesis a follow-up study was conducted using another adaptation of the GARDskin protocol capable of providing continuous potency predictions. The results classified the plasticizer as a weak contact allergen. As a result, a different material was selected for the device.
Keywords: Biocompatiblity, ISO 10993-10, GARDskin Medical Device, Skin Sensitization, ISO 10993-12
Identify workplace hazards by testing skin and respiratory sensitizers
Redeye: CEO interview Q3 2024
Redeye interviewed the CEO of SenzaGen, Peter Nählstedt, where they discussed the company’s recently released Q3 2024 report.
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Penser Access by Carnegie: CEO interview (in Swedish)
Penser Access by Carnegie har intervjuat SenzaGens VD, Peter Nählstedt, i samband med bolagets Q3-rapport.
Intervjun behandlar utvecklingen under kvartalet, bolagets syn på det aktuella marknadsläget samt framtidsutsikter och strategiska prioriteringar framöver.
VD-intervju – Stark tillväxt i Q3
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SenzaGen’s Newsletter Oct 2024
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Joint poster with L’Oreal: Point of Departure for risk assessment
Improved Confidence of Quantitative Sensitizing Potency Assessment for Point of Departure Using GARD®skin Dose-Response
Presented at SOT and Eurotox 2024
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In conclusion, the readouts from GARDskin Dose-Response derive a quantitative continuous potency estimate of skin sensitizers that may be used directly as a PoD for a seamless integration into downstream NGRA.
Identification of skin sensitization hazard and potency characterization are central aspects of risk assessment of chemicals. Current legislation advocates a transition from hazard assessment using in vivo methods to UN GHS potency subclassification and quantitative risk assessment by use of New Approach Methodologies (NAM:s) as well as Defined Approaches (DA). However, the ability of NAM assays to quantitatively estimate sensitizing potency and thereby establish a point of departure (POD) for next-generation risk assessment (NGRA) strategies is currently lacking.
To this end, the GARDskin Dose-Response (DR) method, adapted from the OECD TG 442E method GARDskin, was recently introduced. The GARDskin DR method evaluates test chemicals in a titrated range of concentrations, in order to investigate the dose-response relationship between the output from the GARDskin prediction algorithm (Decision Values; DV:s) and test chemical concentration. The combined information can be used to derive a quantitative estimation of sensitizing potency, defined as the cDV0-value, i.e, the least required dose required to elicit a positive response by the prediction model.
The current work focuses on optimizing the ability of GARDskin DR to derive a quantitative POD based on conversion to a composite Potency Value (PV; µg/cm2), taking into account both human and in vivo reference data sources. A total of 25 chemicals were used to construct predictive regression models fitted to reference PV:s. Results show that the updated models fitted to reference PV:s produced more accurate potency predictions compared with models fitted with, and aiming to predict, only LLNA EC3 and NOEL, respectively. Mean fold-change errors ranged between 2.8 and 3.2, with predicted POD:s being within or close to the range of the variation of the historical in vivo data. In addition, uncertainty in predictions was reduced, as estimated by a minimum 2-fold reduction of 95%-confidence intervals, when comparing models fitted to reference PV:s with models fitter with only LLNA EC3 and human NOEL, respectively.
In conclusion, these improvements constitute a major step forward for the ability of NAM:s to assess quantitative sensitizing potency. It demonstrates how GARDskin Dose-Response can accurately estimate a POD and be incorporated into downstream strategies for quantitative risk assessment (QRA), to ultimately contribute to the assessment of safe use levels of chemicals.
Keywords: NAM, GARDskin Dose-Response, Sensitizing potency, Quantitative risk assessment, Point of departure
Joint poster with Cargill: Case study on Hydrophobic Esters
The applicability of GARD®skin for assessing skin sensitization potential of hydrophobic esters during product development
Presented at Eurotox 2024
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In conclusion, the inclusion of GARD®skin in the OECD test guidelines has expanded the applicability of NAMs for skin sensitization assessment.
This study demonstrates the applicability of the GARDskin assay to assess skin-sensitizing hazard of hydrophobic esters, which provides an ethical alternative to animal methods for safety assessment during product development.
The field of skin sensitization assessment is rapidly evolving and the recent advancements in New Approach Methodologies (NAMs) has made it possible for the industry to perform in vitro skin ssensitization testing with good predictivity across a large chemical space. However, challenges remain for “difficult-to-test” chemicals, those with challenging physical/chemical properties or of Unknown or Variable composition, Complex reaction products or Biological materials (UVCBs), which are often outside the applicability domain of conventional cell-based assays. GARDskin (OECD TG 442E) is a genomic-based assay with demonstrated applicability to “difficult-to-test” substances.
The aim of this study is to assess the skin sensitization potential of two ester substances of biological origin, substance A and B, using the GARDskin assay. These substances are very hydrophobic and fall outside of the applicability domain of the conventional in vitro assays.
Both substances were successfully solubilised in cell media by utilizing a combination of heating, sonication, and selection of appropriate solvent vehicles (ethanol or acetone). No cytotoxicity was observed for either substance, thus 500 μM was chosen as the input concentration for cellular stimulations. GARDskin combines a genomic readout with machine learning to predict skin sensitizing hazard, where values above the threshold (DV=0) is predicted as skin sensitizing and below as non-sensitizing. Both substances resulted in negative mean Decisions Values and thus were classified as non-sensitizers in GARDskin (A: -1.54, B: -0.339).
In conclusion, with the inclusion of GARDskin into the OECD test guidelines, the range to where NAMs are appropriate has been increased. This study demonstrates the applicability of the GARDskin assay to assess skin sensitizing hazard of hydrophobic ethyl esters, which provides an ethical alternative to animal methods for safety assessment during product development.
Keywords: NAM, skin sensitization, difficult-to-test, UVCB, Applicability domain
Joint poster with Sonova: Integrating NAMs into early-stage screening of novel materials
Case studies on the use of GARD®skin Medical Device for in vitro skin sensitization assessment
Presented at SOT and Eurotox 2024
Download a copy in PDF
In conclusion, GARDskin Medical Device can detect signals from diluted and complex extracts of solid devices, with a sensitivity superior to animal methods.
This may significantly reduce the need of animal studies, improve the safety of the final product, and avoid potential costly late-stage failures.
The recent advancements in New Approach Methodologies enables the use of in vitro method for skin sensitization assessment as part of the biocompatibility testing for medical devices, which is conventionally tested in vivo. GARDskin OECD TG 442E is included in ISO 10993-10:2021 as the only OECD validated in vitro assay that is compatible with both polar and non-polar extraction vehicles, in line with ISO 10993-12:2021. GARDskin Medical Device is an adaptation of the GARDskin assay, including a pre-sample treatment procedure where solid devices are extracted using both polar and non-polar vehicles.
The aim of this study is to demonstrate the benefits of using GARD for early-stage screening of materials intended for use in medical devices for assessing their skin sensitization potential. Results from two case studies were summarized in which GARDskin Medical Device was used for skin sensitization assessment. The first case study describes the testing of an acrylic-based device with a coating consisting of a UV-cured lacquer, where chemical analysis indicated the potential for skin sensitization. The second case study describes the testing of a polymeric material consisting of Cellulose-Acetate Propionate (CAP) with a plasticizer (Triethylene glycol bis (2-ethylhexanoate), CAS# 94-28-0), with contradictive existing in vivo (negative) and in vitro (positive) data.
In the first case study, the acrylic-based device induced a positive response in both polar and non-polar vehicles in GARDskin Medical Device and was thus classified as a skin sensitizer. It was hypothesized that the positive results may be due to inadequate curing of the lacquer within cavitary structures of the devices, where UV light exposure was insufficient. To confirm the hypothesis, follow-up testing was performed on an identical device, but without cavities, which was classified as non-sensitizer. In vivo data confirmed the outcome of the in vitro assay. Consequently, a modification was made in the manufacturing process to prevent the presence of lacquer in cavitary structures of the device.
In the second case study, the CAP material was positive in the non-polar vehicle and was thus labelled as a skin sensitizer. The plasticizer was identified as a potential culprit, considering a borderline negative result in LLNA (SI=2.97) and reported positive clinical data. To support the hypothesis a follow-up study was conducted using another adaptation of the GARDskin protocol capable of providing continuous potency predictions. The results classified the plasticizer as a weak contact allergen. As a result, a different material was selected for the device.
Keywords: Biocompatiblity, ISO 10993-10, GARDskin Medical Device, Skin Sensitization, ISO 10993-12
Joint publication with L’Oréal: In vitro prediction of skin sensitizing potency using the GARDskin Dose-Response assay: A simple regression approach
New joint publication with L’Oréal.
We are excited to announce the recent publication of a collaborative scientific paper by the expert teams at L’Oréal and SenzaGen, in Toxics MDPI. This peer-reviewed article presents new evidence on the performance of the GARD®skin Dose-Response for quantitative potency assessment of skin sensitizers.
With an extended set of 30 chemicals and a composite potency model for the prediction of sensitizing potency, the study demonstrates the ability of GARD®skin Dose-Response to predict a Point-of-Departure (PoD) for potential skin sensitizers, showing concordance with NESIL values derived from LLNA and Human data.
This research represents a significant advancement in deriving PoD values for chemicals which can be used directly in improving downstream risk assessment strategies.
Keywords
NAM; GARDskin Dose-Response; Sensitizing potency; Quantitative risk assessment; Point of departure
Abstract
Toxicological assessments of skin sensitizers have progressed towards a higher reliance on non-animal methods. Current technological trends aim to extend the utility of non-animal methods to accurately characterize skin sensitizer potency.
The GARDskin Dose-Response assay was previously described where it was shown that its main readout, the cDV0 concentration, was associated with skin sensitizing potency. The ability to predict potency in the form of NESILs derived from LLNA or human NOEL, from cDV0, was evaluated. The assessment of a dataset of 30 chemicals showed that the cDV0 values still correlated strongly and significantly with both LLNA EC3 and human NOEL values (ρ = 0.645-0.787 [p < 1×10-3]).
A composite potency value that combined LLNA and human potency data was defined, which aided the performance of the proposed model for the prediction of NESIL. The potency model accurately predicted sensitizing potency, with cross-validation errors of 2.75 and 3.22 fold changes compared with NESILs from LLNA and human, respectively.
In conclusion, the results suggest that the GARDskin Dose-Response assay may be used to derive an accurate quantitative continuous potency estimate of skin sensitizers.
CEO comments on successful financing round to accelerate portfolio expansion
We are excited to announce that SenzaGen has secured financing to accelerate portfolio expansion within non-animal testing.With this, we are poised for an even brighter growth path.
The funding enables us to better meet customer needs within in vitro testing, fostering innovation and advancing non-animal testing solutions.
CEO Peter Nählstedt comments: “This achievement underscores our commitment to non-animal testing, innovation, and excellence. We are proud of the progress we have made and look forward to continuing to drive the transition from animal testing to methods better suited to reflect human biology. This will contribute to creating safer and more efficacious products for society.”
CEO interview by Redeye analyst Gustaf Meyer (ENG)
Joint poster with RIFM and IFF: In vitro Skin Sensitization Potency Prediction Performance on the RCPL list
GARD®skin Dose-Response for skin sensitizing potency assessment: Performance on the Reference Chemical Potency List (RCPL)
Presented at BTS 2024.
Download a copy in PDF
New Approach methods (NAMs) for assessment of skin sensitizing hazard and GHS potency have been adopted as OECD Test Guidelines. However, NAMs able to estimate skin sensitization potency on a continous scale, and thereby generating data for use as a point of departure (PoD) in next-generation risk assessment (NGRA) strategies, are currently lacking.
GARDskin Dose-Response (DR) is based on the validated protocols of GARDskin (OECD TG 442E). The readout of the assay is a continous potency prediction, or a No Expected Sensitization Induction Level (NESIL) value (ug/cm2).
The aim of this study was to evaluate the performance of the potency predictions from GARDskin DR using the chemicals in the Reference Chemical Potency List (RCPL) as a reference. The RCPL contains curated Potency Values (PVs) and was developed to serve as a resource for evaluating potency predictions from NAM-based approaches.
GARDskin data was generated (n=27 chemicals) and potency predictions were compared to PV values from the RCPL list. The continous potency predictions from GARDskin DR correlated well with PVs (p: 0.74), with an average misprediction of 3.7-fold from reference values. Furthermore, GARDskin DR provided a very similar potency ranking compared to the ranking based on PV values in the RCPL list (ρ: 0.69).
In conclusion, based on the results in this study, GARDskin DR was demonstrated to be a potent predictor of skin sensitizing potency, increasing the confidence in using the assay to conduct NGRA on new chemical entities, ultimately reducing the need for confirmatory human and/or animal studies.
Keywords: NAM, GARDskin Dose-Response, Sensitizing potency, Quantitative risk assessment, Point of departure
SenzaGen’s Newsletter April 2024
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