To replace a test animal, which is clearly a complex system with blood and organs, you need a more holistic approach to model the entire allergic response within the body. Our CEO Anki Malmborg Hager explains the benefits of using alternative testing methods to prove product safety in this week’s Manufacturing Chemist.
Int. J. Mol. Sci. 2019, 20(3), 666; https://doi.org/10.3390/ijms20030666
Gunilla Grundström and Carl A.K. Borrebaeck
There is an increasing demand for alternative in vitro methods to replace animal testing, and, to succeed, new methods are required to be at least as accurate as existing in vivo tests. However, skin sensitization is a complex process requiring coordinated and tightly regulated interactions between a variety of cells and molecules. Consequently, there is considerable difficulty in reproducing this level of biological complexity in vitro, and as a result the development of non-animal methods has posed a major challenge. However, with the use of a relevant biological system, the high information content of whole genome expression, and comprehensive bioinformatics, assays for most complex biological processes can be achieved. We propose that the Genomic Allergen Rapid Detection (GARD™) assay, developed to create a holistic data-driven in vitro model with high informational content, could be such an example. Based on the genomic expression of a mature human dendritic cell line and state-of-the-art machine learning techniques, GARD™ can today accurately predict skin sensitizers and correctly categorize skin sensitizing potency. Consequently, by utilizing advanced processing tools in combination with high information genomic or proteomic data, we can take the next step toward alternative methods with the same predictive accuracy as today’s in vivo methods—and beyond.
Don’t miss the interesting article in Chemical Risk Manager in Chemical Watch, written by Dr. Emma Davies, with the title; Skin sensitization integrated testing strategies need rethink, expert suggests.
Emma Davies is interviewing the UK academic Dr. David Roberts after his publication of an article in Regulatory Toxicology and Pharmacology in July where he is discussing the relevance of Integrated Testing Strategy (ITS) for skin sensitization testing as well as he is presenting data on the performance of the GARDskin test method compared to the OECD -tests.
In short, David Roberts from Liverpool John Moores University questions the science behind integrated testing strategies (ITS) for combining in vitro test methods to predict skin sensitisation, as required by REACH. Instead, Roberts suggests, ”… that the genomic allergen rapid detection (GARD) assay for skin sensitisation may outperform other OECD validated test methods, either alone or in combination.”
Dr. David Roberts will present his data and the research behind the article at the EUSAAT meeting 23-26 September in Linz, and at the ESTIV conference 15-18 October in Berlin
Read the article here (requires subscription to Chemical Watch):
SenzaGen is proud to be hosting the event Understanding Biocompatibility of Medical Devices on October 2, in Lund, in cooperation with the Medicon Valley Medtech Network.
The regulatory framework of the medical device industry is going through major changes and the complexity of the process is increasing due to new requirements and standards. All medical devices that come into physical contact with patients must be assessed for allergy. Therefore, manufacturers must conduct biological safety evaluations before market approval.
Learn more about biocompatibility, the legal framework and the new ISO 10993
– sign up for the event here
Date: 2nd of October, 2018
Time: 17.00- 20.00
Host: SenzaGen AB
Venue: Auditorium, Medicon Village, Lund
Moderator: Rose-Marie Jenvert, Project Manager, SenzaGen AB
|17.00-17.30||Registration, networking and a bite to eat
– meet the fellow participants
Petter Hartman, CEO, Medicon Valley Alliance
Kerstin Jakobsson, CEO, Medicon Village Innovation
|17.35-17.40||Sponsor of the event
Peter Borg Gaarde, Partner & European Patent Attorney, HØIBERG P/S
|17.40-18.15||Introduction to Biocompatibility and ISO 10993
Monica Grekula, Business Area and Team Manager Toxicologist, Symbioteq
|18.15-18.35||Chemical characterization according to ISO 10993-18
Carsten Senholt, Principal Toxicologist, SAXOCON
|18.35-18.55||Industry perspective on hemocompatibility testing according to ISO 10993-4
Barbara Musi, Principal Research Scientist Toxicology & Biocompatibility, Baxter
|18.55-19.25||Case study: Medical Device Biocompatibility Testing
Helge Gehrke, Head of in vitro Pharmacology and Toxicology, Eurofins Biopharma Product Testing Munich
|19.25-19.30||Presentation of Swiss Medtech Delegation
Gustav Henriksson, Head of Trade Section, trade Point Nordics, Embassy of Switzerland in Sweden
Rose-Marie Jenvert, Project Manager, SenzaGen AB
|19.30-20.00||Networking and a light snack|
Deadline for registration is 25h of September, 2018.
Jenvert RM, Larne O, TorstensdotterMattssonU. I., Johansson H, SenzaGen, Lund, Sweden
The Genomic Allergen Rapid Detection (GARD) assay is a state of the art in vitro assay developed for the assessment of skin sensitizers. It is based on gene expression analysis of SenzaCells, a human myeloid cell line, after stimulation by the test item.
During the development of the GARD platform, two solvents were used; DMSO (0.1%) and Water. To increase the applicability domain of GARD® and the solubility of certain test items, for e.g. Medical Device extracts and UVCBs, we here show a broader range of solvents compatible with GARD.
Here, we show that the GARD® platform is compatible with the following solvents:
• Super refined olive oil
U. I. Torstensdotter Mattson, C. Humfrey, O. Larne, H. Johansson, L. Sweet
SenzaGen, Lund, Sweden, Lubrizol, Derbyshire, United Kingdom, Lubrizol, Ohio, United States of America
The GARD – Genomic Allergen Rapid Detection – platform is a state of the art in vitro assay for assessment of chemical sensitizers. The GARD®skin assay is a powerful tool for assessment of chemical sensitizers, with a predictive accuracy of 94%. In this study, four UVCB test items, provided by Lubrizol and selected based on existing in vivo data (internal Lubrizol data), were evaluated. Sensitizing hazard was assessed using the GARD®skin assay, and the GARD®potency assay further subcategorized the sensitizers into strong (1A) or weak (1B) sensitizers according to GHS/CLP classification. Here we show the importance of using appropriate vehicles in order to predict a correct classification of Test items.
A UVCB Test item with poor water and DMSO solubility was assessed using a mixture of vehicles with different polarity indexes (DMF and Glycerol 1:1). This experimental vehicle mixture classified the UVCB as a skin sensitizer, being consistent with the in vivo data. This case study demonstrates the broadening of the applicability domain of the GARD -assay when assessing UVCBs.
Sandberg P, Johansson A, Agemark M, Gradin R, Larne O, Appelgren H, Forreryd A, Jerre A, Edwards A, Hoepflinger V, Burleson F, Gehrke H, Roggen E, Johansson H
SenzaGen, Lund, Sweden, Burleson Research Technologies, Morrisville, US, Eurofins, Munich, Germany
The prevalence of allergic contact dermatitis (ACD) is estimated to >20% in the western world. Not only the individual is affected, but downstream socioeconomic effects are high. To minimize exposure, chemicals must be safety tested. Traditional testing strategies like the murine local lymph node assay (LLNA) comprise animals, but the regulatory authorities, public opinion and economic interests require animal-free models. The Genomic Allergen Rapid Detection skin (GARD®skin) is an in vitro assay addressing this need. Here, we present the results of the GARD®skin ring trial (OECD TGP 4.106) for validity of the assay. In addition, we show data for GARD®potency – a complementary assay developed to categorize identified senitizers as CLP 1A or 1B.
WLR: 82 – 89%BLR: 92% (92 – 100%)
– Accuracy: 94%
– Sensitivity: 93%
– Specificity: 96%
A blinded ring trial was performed to assess the functionality of the GARDskin assay. The data demonstrates that GARDskin is a powerful tool for assessment of chemical skin sensitizers, with a predictive accuracy of 94% and excellent reproducibility between laboratories.
In addition, we show that GARDpotency accurately assesses potency of identified sensitizers.
GARD – Genomic Allergen Rapid Detection – is a state of the art non-animal based technology platform for classification of skin sensitizing chemicals. The assay has proven to be reliable and highly accurate for identification of skin sensitizing chemicals, and consistently reports predictive performances > 90% across external test sets. The aim of the current project is to complement assessments of average model performance with an estimate of uncertainty involved in each individual prediction, thus allowing for classification of skin sensitizers with confidence.
An Internal validation procedure was initially performed on samples in the GARD training set (n=38) using the strategy described in Fig. 3A. Results from this exercise is summarized in Fig. 4A. Conformal prediction by definition allows the user to determine a reasonable and acceptable significance level to guarantee a maximum error rate in predictions. The significance level was set to 15%, i.e. the model was allowed to make a maximum of 15% errors. Performance of the conformal predictor was measured by validity and efficiency. A model was valid if the number of prediction errors did not exceed the significance level, while efficiency corresponded to the percentage of single class predictions. Internal validation of the training data resulted in a valid and highly effective model (92% single classifications, 1 empty, 2 both), indicating that the ambitious significance level was at a reasonable level for the GARD® assay. Following internal validation, samples in a large external test set (n =70) was classified within the CP framework as described in Fig. 3B, which resulted in generation of a valid and highly efficient model (99% single classifications, 0 empty, 1 both) (Fig.4B). Additional data on model performance is illustrated in Table 1.
Renato Ivan de Ávila, Tim Lindbergh, Malin Lindstedt and Marize Campos Valadares
Lab. of Education and Research in Pharmacology and Cellular Toxicology, Faculty of Pharmacy, Federal University of Goiás, Goiânia, Goiás, Brazil.
Department of Immunotechnology, Medicon Village, Lund University, Lund, Sweden.
Genomic fingerprints in dendritic cells after chemical exposure is a recent strategy in in vitro techniques for skin sensitization hazard. Within this perspective, Genomic Allergen Rapid Detection (GARDskinTM), an assay based on a support vector machine (SVM) model, was developed for identifying contact allergens using a myeloid cell line as a surrogate for dendritic cells. Predictive system behind the GARDskin™ consists on the transcriptional quantitative analysis of 200 genes, referred as the GARDskin™ prediction signature. Mechanistically, GARDskin™ is linked to key event 3 “Activation of DCs”, as defined by the Adverse Outcome Pathways for skin sensitization published in 2012 by OECD (https://read.oecd-ilibrary.org/environment/the-adverse-outcome-pathway-for-skin-sensitisation-initiated-by-covalent-binding-to-proteins_9789264221444-en#page1)
Information declared on the label and lawsone and PDD levels found in ten commercial henna-based hair coloring cosmetics are show in Table 1. Since all products analyzed were declared as henna cosmetics by the manufactures, the presence of LAW, the main active phytochemical of henna, was then expected in all samples. However, HPLC analysis showed no LAW level in the product nº 2, suggesting falsification. Furthermore, the presence of PPD was declared on the products nº 2 and 8 only by the manufactures. However, this substance was detected in all products, suggesting undisclosed adulteration.
There are more and more countries banning animal testing of ingredients and final products within the cosmetics industry. Also other industries are facing a growing demand for accurate animal-free test methods. The primary forces driving this development are stricter regulatory demands and an urge to reduce the use of animals in experimental testing.
A new scientific study confirms that SenzaGen´s test method GARDskin™ is more accurate than the other animal free methods (DPRA, KeratinoSens (ARE-Nrf2 luciferase) and h-CLAT) on the market today, for skin sensitization testing. Due to the relatively low accuracy of these methods, regulatory authorities recommend a combination of them, in order to better predict the skin sensitization activity of a substance. However, not even a combination of these tests show the same high level of accuracy as displayed by GARDskin™.
”This recently published scientific study shows that GARDskin™ can give producers and contract laboratories a possibility to use one single test to evaluate their substances with even better accuracy than a combination of several test methods. GARDskin™ has the potential to make testing of chemicals more efficient with respect to time, resources and quality”, says SenzaGen CEO Anki Malmborg Hager.
The scientific study was carried out by Dr David W Roberts at the School of Pharmacy and Biomolecular Sciences, Liverpool John Moores University in the UK. The results are published in the scientific journal Regulatory Toxicology and Pharmacology and can be accessed via this link : “Is a combination of assays really needed for non-animal prediction of skin sensitization potential? Performance of the GARD™ (Genomic Allergen Rapid Detection) assay in comparison with OECD guideline assays alone and in combination”
For more information:
Anki Malmborg Hager, CEO, SenzaGen AB
Phone: +46 768 284822
GARD™ is a group of tests for assessing chemical skin sensitizers. The tests make use of genetic biomarkers for more than 200 genes which cover the entire immune reaction and are relevant to predicting the risk of hypersensitivity. The tests have over 90 percent reliability. This compares with the current predominant test method, experiments on mice, which has an accuracy of 70-75 percent. SenzaGen’s tests are also capable of measuring the potency of a substance’s allergenic properties. Consequently, GARD tests provide a much more comprehensive basis for determining whether a substance should be classified as an allergen than current testing methods.
SenzaGen makes it possible to replace animal experiments with in vitro genetic testing to determine the allergenicity of the chemicals we come into contact with in our daily lives, such as for example in cosmetics, pharmaceuticals, food products and dyes. The company’s patented tests are the most reliable on the market and provide more information than traditional evaluation methods. We ourselves sell the tests in Sweden and the USA, and we sell through partners in several other countries. Over the next few years the company will expand geographically, make alliances with more distribution partners and launch further unique tests. SenzaGen has its headquarters in Lund in Sweden and a subsidiary in San Francisco, USA. For more information visit http://www.senzagen.com.
The information was submitted for publication, through the contact person set out above on the 31 July 2018 at 08.50.
SenzaGen AB is listed on Nasdaq First North in Stockholm and FNCA is the company’s Certified Adviser. For more information, please visit http://www.senzagen.com.