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.

Gradin R, Tourneix F, Mattson U, Andersson J, Amaral F, Forreryd A, Alépée N, Johansson H. 
Toxics. 2024; 12(9):626. 
https://doi.org/10.3390/toxics12090626

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.

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Conclusion

  • This study suggests that GARDskin Dose-Response is a potent predictor of skin sensitizing potency on a continuous scale.
  • Predicted potency values are in µg/cm2 and can be used for risk assessment and to establish a maximum allowable concentration of a sensitizer in various consumer products.

​

Abstract

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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Joint publication with IFF and RIFM: GARDskin dose-response assay and its application in conducting Quantitative Risk Assessment (QRA) for fragrance materials using a Next Generation Risk Assessment (NGRA) framework

New joint publication with International Flavors & Fragrances Inc (IFF) and Research Institute for Fragrance Materials (RIFM).

SenzaGen scientists, alongside the scientific teams at International Flavors & Fragrances Inc and Research Institute for Fragrance Materials, have jointly published an article in Regulatory Toxicology and Pharmacology, presenting new peer-reviewed evidence on the performance of the GARD®skin Dose-Response assay for Quantitative Risk Assessment of fragrance materials.

The study results confirm the ability of GARD®skin Dose-Response to predict human NESIL values with good predictive performance, showing good concordance with published reference Human data and demonstrating good reproducibility.

Furthermore, the paper also presents a case study to illustrate how the predicted NESIL value from GARDskin Dose-Response can be used in practice within a NGRA framework to establish a maximum allowable concentration of a sensitizer in different consumer products.

The study represents a major step towards the establishment of the assay to derive NESIL values for conducting QRA evaluations for fragrance materials using an NGRA framework.

Shashikiran Donthamsetty, Andy Forreryd, Paul Sterchele, Xiao Huang, Robin Gradin, Henrik Johansson, Ulrika Mattson, Isabelle Lee, Anne Marie Api, Gregory Ladics,
Regulatory Toxicology and Pharmacology, Volume 149, 2024, 105597, ISSN 0273-2300,
https://doi.org/10.1016/j.yrtph.2024.105597

Keywords

QRA (Quantitative Risk Assesment); Dermal sensitization; Fragrance materials; Next Generation Risk Assesment (NGRA); GARD assay; No Expected Sensitization Induction Level (NESIL); New Approach Methodologies (NAMs); OECD 442E

Highlights

  • Developed a Next Generation Risk Assessment (NGRA) framework for conducting QRA2 for fragrance materials.
  • The GARDskin Dose Response (DR) assay is a reliable and reproducible method for predicting NESIL for fragrance materials.
  • NGRA for QRA2 was validated using isocyclocitral as a case study.


Abstract

Development of New Approach Methodologies (NAMs) capable of providing a No Expected Sensitization Induction Level (NESIL) value remains a high priority for the fragrance industry for conducting a Quantitative Risk Assesment (QRA) to evaluate dermal sensitization. The in vitro GARDskin assay was recently adopted by the OECD (TG 442E) for the hazard identification of skin sensitizers. Continuous potency predictions are derived using a modified protocol that incorporates dose-response measurements. Linear regression models have been developed to predict human NESIL values. The aim of the study was to evaluate the precision and reproducibility of the continuous potency predictions from the GARDskin Dose-Response (DR) assay and its application in conducting QRA for fragrance materials using a Next Generation Risk Assessment (NGRA) framework. Results indicated that the GARDskin Dose-Response model predicted human NESIL values with a good degree of concordance with published NESIL values, which were also reproducible in 3 separate experiments. Using Isocyclocitral as an example, a QRA was conducted to determine its safe use levels in different consumer product types using a NGRA framework. This study represents a major step towards the establishment of the assay to derive NESIL values for conducting QRA evaluations for fragrance materials using a NGRA framework.

Evaluation of the applicability of GARDskin to predict skin sensitizers in extracts from medical device materials

Peer-reviewed article in Frontiers in Toxicology.

The application of GARD®skin to predict potential skin sensitizers in extracts from Medical Device materials has recently been published in a peer-reviewed article in Frontiers in Toxicology.

The study results provide evidence recognizing the GARD®skin Medical Device assay as a scientifically sound and ethical alternative to conventional animal methods, compatible with both polar and non-polar extraction vehicles, in line with the ISO 10993-12:2021 standard.

We are proud to provide the only OECD-validated in vitro assay for sensitization that is fully compatible with testing requirements specified in ISO 10993-12.

Jenvert Rose-Marie, Larne Olivia, Johansson Angelica, Berglin Mattias, Pedersen Emma, Johansson Henrik
Frontiers in Toxicology, Volume 6, 2024,ISSN 2673-3080
DOI=10.3389/ftox.2024.1320367

Keywords

GARD™; In vitro; Skin sensitisation; NAMs; Medical Device, ISO 10993, Biocompatibility

Abstract

Biocompatibility testing of medical devices is governed by the ISO 10993 series of standards and includes evaluation of skin sensitization potential of the final product. A majority of all medical devices are tested using in vivo methods, largely due to the lack of in vitro methods validated within the applicability domain of solid materials. The GARDskin method for assessment of chemical skin sensitizers is a validated method included in the OECD Test Guideline 442E, based on evaluation of transcriptional patterns of an endpoint-specific genomic biomarker signature in a dendritic cell-like cell, following test chemical exposure. The current study aimed to evaluate the applicability of GARDskin for the purpose of testing solid materials by incorporation of extraction procedures described in ISO 10993-12:2021, as well as to demonstrate the functionality of the proposed protocols, by testing of custom-made materials spiked with sensitizing agents. It was shown that GARDskin is compatible with both polar and non-polar extraction vehicles frequently used for the purpose of medical device biological testing. Further, exploring three different material types spiked with up to four different sensitizing agents, as well as three unspiked control materials and commercial reference products, it was shown that the method correctly classified all evaluated test materials. Taken together, the data presented suggest that GARDskin may constitute a valid alternative to in vivo experimentation for the purpose of skin sensitization assessment of medical devices.

 

Joint publication with Corteva Agriscience: GARD™skin and GARD™potency: A proof-of-concept study investigating applicability domain for agrochemical formulations

New joint publication with Corteva Agriscience.

SenzaGen scientists, alongside the toxicology team at Corteva Agriscience, have recently published a joint study in Regulatory Toxicology and Pharmacology, presenting new peer-reviewed evidence on the applicability of GARD® for agrochemical formulations.

The study demonstrates a satisfactory performance of GARD®skin and GARD®potency for skin sensitization hazard and GHS potency categorization of tested agrochemical formulations.

Marco Corvaro, Joseph Henriquez, Raja Settivari, Ulrika Mattson, Andy Forreryd, Robin Gradin, Henrik Johansson, Sean Gehen,
Regulatory Toxicology and Pharmacology, Volume 148, 2024, 105595, ISSN 0273-2300,
https://doi.org/10.1016/j.yrtph.2024.105595.

Keywords

GARD™; In vitro; Skin sensitisation; NAMs; Agrochemical formulations

Highlights

  • Tested 42 agrochemical formulations to expand applicability domain of GARD.

  • GARDskin showed good accuracy (76.2%), sensitivity (85.0%) and specificity (68.2%).
  • GARDpotency correctly subcategorized 14/17, correctly predicted sensitisers.
  • GARD satisfactory for Key Event 3 characterisation of agrochemical formulations.


Abstract

Several New Approach Methodologies (NAMs) for hazard assessment of skin sensitisers have been formally validated. However, data regarding their applicability on certain product classes are limited. The purpose of this project was to provide initial evidence on the applicability domain of GARD™skin and GARD™potency for the product class of agrochemical formulations.

For this proof of concept, 30 liquid and 12 solid agrochemical formulations were tested in GARDskin for hazard predictions. Formulations predicted as sensitisers were further evaluated in the GARDpotency assay to determine GHS skin sensitisation category. The selected formulations were of product types, efficacy groups and sensitisation hazard classes representative of the industry’s products.

The performance of GARDskin was estimated by comparing results to existing in vivo animal data. The overall accuracy, sensitivity, and specificity were 76.2% (32/42), 85.0% (17/20), and 68.2% (15/22), respectively, with the predictivity for liquid formulations being slightly higher compared to the solid formulations. GARDpotency correctly subcategorized 14 out of the 17 correctly predicted sensitisers. Lack of concordance was justifiable by compositional or borderline response analysis. In conclusion, GARDskin and GARDpotency showed satisfactory performance in this initial proof-of-concept study, which supports consideration of agrochemical formulations being within the applicability domain of the test methods.

 

SenzaGen’s Newsletter Year-end 2023

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Joint poster with Takasago: Determining safe use level for novel fragrance ingredients

Practical application of the GARD®skin Dose-Response assay to derive a No Expected Sensitization Induction Level (NESIL) value for confirmatory human patch studies to determine safe use level for novel fragrance ingredients


Presented at ACT 2023

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Summary

  • Combining the results from the GARD®skin Dose-Response assay with other NAMs enables weight-of-evidence based approaches to determine safe use levels of novel fragrance ingredients.
  • Based on the results from the weight-of-evidence approach, confirmatory human patch test and HRIPT are conducted. HRIPT is performed at the top concentration of 11250ug/cm2. Both results are negative, confirming the predicted NESIL-value from GARD®.

 

Abstract

Skin sensitization is one of the required endpoints for the development and registration of novel fragrance ingredients. Traditionally, assessment has been performed combining in vitro and in vivo assays, but recent developments has shifted to the use of New Approach Methodologies (NAMs), without 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.

This study presents the use of the GARDskin Dose-Response assay to determine safe human user levels for one novel fragrance ingredient. The experimentally predicted NESIL value was determined to 37800µg/cm2. Using a weight-of- evidence approach mainly guided by the quantitative data from the GARDskin Dose-Response, confirmatory Human Repeated Insult Patch Testing (HRIPT) studies were conducted and at the tested concentrations, no sensitization reactions were observed. In conclusion, this study expands the toxicologist´s toolbox and illustrates the potential to use the GARDskin Dose-Response assay to derive NESIL values that are protective of human health, without having to rely on the Dermal Sensitization Threshold (DST) approach or reverting to traditional animal testing approaches.

New scientific publication by ExxonMobil: Challenges integrating skin sensitization data for assessment of difficult to test substances

New scientific publication by ExxonMobil.

Check out this newly published article by ExxonMobil focusing on the challenges of determining skin sensitization hazard in the case of difficult-to-test substances with conflicting or low-confidence data, where GARDskin data on UVCBs and hydrophobic substances provides valuable input for the integrated hazard assessment.

The article provides new peer-reviewed evidence for the applicability domain of GARDskin on UVCBs, hydrophobic and highly complex substances.

Greminger A, Frasca J, Goyak K, North C. 
Challenges integrating skin sensitization data for assessment of difficult to test substances. 
ALTEX - Alternatives to animal experimentation, published Oct 12, 2023
doi: 10.14573/altex.2201122. Epub ahead of print. PMID: 37843019.

Keywords

LLNA; ToxPi; new approach methodologies; skin sensitization; weight of evidence.


Abstract

Difficult to test substances including poorly soluble, mildly irritating, or Unknown or Variable Composition Complex reaction products or Biological Materials (UVCBs), producing weak or borderline in vivo results, face additional challenges in in vitro assays that often necessitates data integration in a weight of evidence (WOE) approach to inform skin sensitization potential. Here we present several case studies on difficult to test substances and highlight the utility of Toxicological Prioritization Index (ToxPi) as a data visualization tool to compare skin sensitization biological activity. The case study test substances represent two poorly soluble substances, tetrakis (2-ethylbutyl) orthosilicate and decyl palmitate, and two UVCB substances, alkylated anisole and hydrazinecarboximidamide, 2-[(2-hydroxyphenyl)methylene]-, reaction products with 2 undecanone. Data from key events within the skin sensitization adverse outcome pathway were gathered from publicly available sources or specifically generated. Incorporating the data for these case study test substances as well as on chemicals of a known sensitization class (sensitizer, irritating non-sensitizer, and non-sensitizer) into ToxPi produced biological activity profiles which were grouped using unsupervised hierarchical clustering. Three of the case study test substances concluded to lack skin sensitization potential by traditional WOE produced biological activity profiles most consistent with non-sensitizing substances, whereas the prediction was less definitive for a substance considered positive by traditional WOE. Visualizing the data using bioactivity profiles can provide further support for WOE conclusions in certain circumstances but is unlikely to replace WOE as a stand-alone prediction due to limitations of the method including the impact of missing data points.