The EPAA project platform (PP) is composed of EPAA partners and associates. It supervises and assesses the projects agreed upon by the EPAA, ensuring scientific quality and effectiveness.
The work of the project teams contributes to the development, validation, acceptance and implementation of 3R alternatives, in regulatory testing and decision making, in Europe and beyond.
The EPAA projects identify knowledge gaps and seek optimised approaches to testing and risk assessment strategies, in existing and upcoming legislation. They contribute to optimise implementation of 3Rs and to avoid redundant or unnecessary testing. Based on the work of its projects, the EPAA strives to share experience and build synergies across all 7 EPAA sectors.
The project platform is the link between the EPAA steering committee (SC) and the EPAA project teams. The key role of PP is to supervise the projects, by
- tracking progress against milestones, and where appropriate, defining further milestones
- proposing changes to milestones/ objectives
- proposing measures on the outcomes
- reporting to SC
- highlighting deviation from proposed timelines/ milestones
The Commission and industry partners jointly lead the project platform. Representatives of both parties were appointed as co-chairs, with Gianni Dal Negro (GSK) for industry partners and Raffaella Corvi (Directorate-General Joint Research Centre) for the European Commission. PP members meet quarterly, usually 15 days before the steering committee meetings, to discuss and report on respective projects and prepare SC meetings.
The EPAA partners promote better science and facilitate regulatory acceptance of alternatives. In doing so, they currently focus on several projects. These address different end-points and reach out to various sectors. The project platform manages all projects.
Comparison of 3D skin models for the assessment of ‘difficult to test substances’
The aim of this EPAA project is to evaluate the 4 most advanced 3D skin model-based methods for their reliability in predicting skin sensitisation.
Sensitisation of human skin to chemicals is a potential danger to human health and requires reliable hazard and risk assessments in order to provide appropriate risk management. The current legislation in Europe for the safety evaluation of chemicals (REACH: EU Regulation No.1907/2006) and cosmetics (EU Regulation No. 1223/2009) includes the requirement to assess the skin sensitisation potential of a substance or formulation. Assessment without the use of animals has been the focus of intensive previous work of many stakeholders and, as a result, a number of validated non-animal tests are accepted as OECD test guidelines (TGs). These and other approaches are being increasingly used as part of optimised testing strategies based on integrated approaches to testing and assessment (IATA) of safety. Furthermore, validated nonanimal test results are now the default information requirement for assessment of skin sensitisation potential of chemicals in REACH. 3 in vitro methods, 2 of which are based on cell culture in aqueous media (KeratinoSens®, human cell line activation test (h-CLAT)) and the in chemico direct peptide reactivity assay (DPRA) are now adopted OECD TGs. However, these methods have certain limitations, for example when testing substances with a very low water solubility (highly hydrophobic substances) or pH instability. Therefore, 3D skin tissue models were developed and relevant markers were identified to detect sensitisers and to differentiate from non-sensitisers. These 3D skin models are able to better mimic the skin structure and organisation, and offer other advantages such as the possibility to directly apply the substances to the model skin. However, the utility of these models for the evaluation of hydrophobic and other ‘difficult to test substances’ was unclear.
3Rs added value and EPAA actions
Establishing the complementarity of the 3D skin model-based methods with the existing methods is very important. The study addresses a critical challenge for industry, as testing of ‘unusual chemicals’ (unusual in the sense of physicochemical properties as compared to chemicals used in validation studies) is not simple. In addition, regulatory bodies need this type of information to understand the use of these models for regulatory classifications. The project should therefore help to better understand the applicability of the models, give guidance on which methods to further sponsor and promote at regulatory level, and to facilitate the acceptance, use, and expansion of the non-animal test methods available for sensitisation testing.
This project brings together stakeholders from European and national regulators and a wide range of industry stakeholders from various sectors (chemicals, cosmetics, fragrances, etc.). The EPAA has been collaborating with Cefic LRI and Cosmetics Europe to ensure optimal cross-fertilisation across sectors.
- The project started in Q1 2016
- A small set of reference chemicals (N=12) for 3D-assays was selected from the list of ‘difficult substances’ assessed at the time by Cosmetics Europe in a variety of in vitro test methods
- 2017-2018: Test data on these 12 blind-coded chemicals were generated by testing them in four 3D-assays
- The results were presented at the Eurotox 2018 conference in Brussels
- In 2019 the results were described in a manuscript that has been submitted for peer-reviewed publication
The project team may consider proposals for future testing of challenging substances and mixtures.
Novel in vitro methods to replace animal-based in-process controls
This pilot project on clostridium septicum antigen aims at validating in vitro assays for toxicity and antigenicity and at proposing their inclusion in the European Pharmacopoeia (Ph. Eur.) to replace the current animal-based tests.
Vaccines for protection against diseases caused by clostridial species in animals are widely used. Many of these vaccines currently require that the toxoid in the bulk preparation used to produce the final vaccine batches is evaluated by animal-based tests, such as the minimum lethal dose (MLD) for toxicity and the total combining power (TCP) for antigenicity. New in vitro methods to replace these animal-based tests are highly desirable and, because of higher sensitivity and accuracy, they offer the potential for improved assurance of quality and safety of the vaccines.
As the vast majority of veterinary clostridial vaccines are based on detoxified cytotoxic antigens, it is expected that these assays could be adapted to all cytotoxin-based clostridial antigens with the potential to greatly reduce the total animal usage in in-process control testing of veterinary vaccines.
3Rs added value and EPAA actions
The number of animals used for clostridial vaccines quality control tests, including the tests for toxicity and antigenicity, represents about 50% of all the animals used for quality control tests of veterinary vaccines. It is thus a high priority to identify alternative tests. Further to recommendations of the original EPAA vaccine consistency project, the EPAA has been supporting since 2013 the collaborative study coordinated by EDQM (Council of Europe) as part of the biological standardisation programme (BSP 130).
This large EDQM project, to which EPAA is contributing, involves 14 vaccine manufacturers and official control laboratories in Europe, USA, Morocco and Mexico.
- A collaborative study organised by the European Directorate for the Quality of Medicines & Healthcare (EDQM, Council of Europe) co-sponsored by the EU started in 2013 and was successfully completed in 2015 with the support of EPAA.
- The results demonstrated that the proposed in vitro assays are relevant and reproducible and show excellent concordance with the animal-based tests currently widely used by the pharmaceutical industry.
- After a further optimisation, the validation of the in vitro assays through an extension of the collaborative study is close to completion.
The project’s results indicate that the non-animal, cell line-based assays for in-process toxicity and antigenicity testing of Cl. septicum vaccines will outperform the animal-based methods. It is expected that after successful completion of the project the assays will be incorporated into the relevant Ph. Eur. monographs. Proceedings from the 2015 EPAA workshop, where the results of the validation of the tissue culture seroneutralisation methods were discussed were published in Pharmeuropa Bio and 2 additional manuscripts are planned based on the final reports of the experimental work.
The project has stimulated considerable interest including the potential for application of the optimised protocol developed in this project to other, in vitro replacement, assay validation research work (e.g. Vac2Vac).
Replacement of animal-based potency tests
The aim of this project is to replace the current in vivo potency test for the release of human rabies vaccines (NIH, mice intracranial challenge test) with an in vitro antigen (G glycoprotein) quantification assay using ELISA technology.
The problematic NIH test involves the use of large numbers of animals, of which half develop distressful rabies symptoms. Furthermore, the variation of the NIH test is high and the test, therefore, may have to be repeated to meet regulatory requirements. Thus, its replacement will have a substantial impact on animal use.
3Rs added value and EPAA actions
EPAA’s contributions have been indispensable for this project by supporting previous workshops and facilitating manufacturers, regulatory and scientific bodies to elaborate a clear strategy for the replacement of the human rabies vaccine NIH method. Since 2012, the EPAA has enabled the creation of an international working group to coordinate a more harmonised approach of the alternative assay development through the acquisition and distribution of a common set of rabies vaccines. The EPAA supports the BSP 148 project which is required for full validation of the in vitro method. This activity requires input from various stakeholders (control authorities, manufacturers, and academia) and cannot be carried out by a single manufacturer. The 2 European manufacturers of human rabies vaccines (Sanofi Pasteur and GlaxoSmithKline) are members of EPAA and provide additional in-kind contribution by participating in the study. The project continues the work started within the EPAA Vaccines consistency approach project and if successful it will contribute to the promotion of 3Rs and overall harmonisation of vaccine quality control via dissemination workshops.
The results of the collaborative study have been presented under the name of EPAA in various world congresses and other international conferences.
In addition to EDQM that coordinates the BSP 148 collaborative study, a number of laboratories and international experts from control authorities, manufacturers, academia and EURL ECVAM contribute to the project.
- A pre-collaborative study involving 5 laboratories evaluating 3 ELISA methods was run in 2013 to identify a suitable replacement method. Compliant sub-potent samples of vaccines prepared from different virus strains were obtained from European and non-European sources with the support of WHO, EDQM and industry.
- Based on the results of this pre-collaborative study, a candidate method was selected.
- The detailed study report has been published in Vaccine (Morgeaux S., et al., 2017.) Replacement of in vivo human rabies vaccine potency testing by in vitro glycoprotein quantification using ELISA – Results of an international collaborative study. Vaccine 35, 966-971
- A major achievement in 2016 has been the endorsement of the proposal for a wide collaborative study to validate the transferability and robustness of the selected ELISA.
- This collaborative study is being run since 2017 under the biological standardisation programme of the Council of Europe and the EU Commission, coordinated by the EDQM (study coded BSP148) with the support of EPAA.
- First experiments confirmed that the selected ELISA is applicable to most virus strains used worldwide for human rabies vaccines.
- In 2018 the results were presented to the developing countries vaccine manufacturers network (DCVMN: India, May 2018), the European society for alternatives to animal testing (EUSAAT: Austria, September 2018), and the joint meeting of national toxicology program interagency center for evaluation of alternative toxicological methods (NICEATM).
It is expected that the data generated throughout the ongoing study will support the revision of the Ph. Eur. monograph on human rabies vaccines as well as global acceptance of the replacement method. The project has prompted considerable interest from international regulators and NGOs.
Identification of clinical signs predictive of mortality
This project aims to identify opportunities to waive the acute toxicity animal testing requirements completely or, where this is not possible, to refine the decision-making steps or assessment strategies so as to minimise the suffering of animals. The ultimate goal is to develop an animal-free decision framework for acute systemic toxicity testing.
Whilst acute toxicity testing is no longer needed in the pharmaceutical sector and in vivo acute toxicity testing is no longer possible in the cosmetics sector, evaluation of acute toxicity remains a requirement for chemicals and agrochemicals in order to establish their overall hazard profile and to meet classification, labelling and packaging (CLP) requirements that are relevant to human safety, for example, in emergency situations. The REACH standard information requirements for the endpoint of acute toxicity (REACH Annex VIII, point 8.5.3.) were revised in May 2016 allowing a waiving of acute toxicity testing via the dermal route under certain circumstances.
In previous years, the EPAA has provided scientific justification in support of such a ‘weight of evidence’ approach. Acute toxicity by the oral route is the most common testing requirement and therefore this route has been prioritised by EPAA.
3Rs added value and EPAA actions
Because the EPAA brings together a wide range of industry sectors, it can offer a unique overview of the regulatory and scientific issues in this field, and can recommend 3Rs approaches that could be adopted widely across different sectors.
The project is based on collaboration with industry partners, experts from EURL ECVAM, the European Commission, UK National Centre for the 3Rs (NC3Rs) and the UK Chemicals Regulation Directorate (CRD).
- The project was initiated in 2008 and reviewed the scientific and regulatory drivers for acute toxicity testing in order to promote the use of 3Rs approaches in this area.
- In 2010, a workshop was organised and a paper was published on ‘cross-sector review of drivers and available 3Rs approaches for acute systemic toxicity testing’ (T. Seidle et al, 2010, Toxicological Sciences). The workshop identified opportunities to waive acute animal testing requirements completely or, where this is not possible, to refine the decision-making steps or strategies to minimise suffering of test animals. Recommendations on a 3Rs-based classification & labelling decision framework were prepared including replacement of death as an endpoint.
- Since 2014, additional evidence in support of this decision framework is being developed through data mining of acute oral toxicity studies in collaboration with the NC3Rs and the UK Chemicals Regulation Directorate.
- Data from previously filed acute toxicity studies are collected and will be analysed to confirm that clinical signs (evident toxicity) are an appropriate alternative to death as an endpoint.
- The findings will be considered in the overall decision framework document.
- Due to delays in selection and accessibility of the data, the project has made slow progress in the last years. In 2016, more than 450 previously filed, acute toxicity studies were screened and in 2018 data from more than 70 studies were collected and evaluated by the NC3Rs.
Additional studies are required to provide a sufficient breadth of quality data to support a statistically robust analysis before the project can be completed. The results of the data mining will be summarized in a joint peer reviewed publication.
Deleting international regulatory requirements for in vivo safety tests
In the production of biological products, manufacturers are required to confirm potency and safety of each batch of product. This may involve the use of laboratory animals. Directive 2010/63/EU prohibits manufacturers in the EU from using an animal test method, if an alternative, non-animal method is recognised by the European pharmacopoeia.
However, if alternatives are not internationally harmonised and accepted in other regions, then excessive or unnecessary animal testing may be undertaken by manufacturers, in order to gain access to other, non-EU markets.
This project aims to achieve global harmonisation in batch testing requirements for human and veterinary vaccines, as well as other biologicals. It is hoped the project will lead to better incorporation of the 3Rs in potency and safety testing strategies. Project deliverables include mapping of regulatory bodies responsible for establishing potency and safety testing requirements, identifying key differences in QC testing requirements between pharmacopoeias or equivalents assessing test methods with related benefits and risks.
European Medicines Agency, European Commission, EURL ECVAM, pharmaceutical industry, animal health industry, Council of Europe - European Directorate for the Quality of Medicines and Healthcare (EDQM).
- 2013: Project launch
- 2013: Mapping of regulatory bodies responsible for potency and safety testing requirements
- 2015: EPAA workshop on international harmonisation of biologicals, Egmond aan Zee, the Netherlands. The mapping results, case studies and recommendations were discussed with regulatory authority representatives at this workshop, with a view to progressing to full international harmonisation.
- 2015: Publication of the September 2015 workshop report in a peer-reviewed journal
- 2016-2018: The project team submitted formal requests to WHO, OIE and Ph. Eur. to encourage deletion of specific tests (GST / ATT and TABST) from their recommendations or requirements.
- 2018: The Ph. Eur. Commission endorsed the complete suppression of the test for abnormal toxicity (ATT) from 49 monographs in the Ph. Eur. (implementation from 1 January 2019).
Further developments within this project are expected. An assessment is currently ongoing by the project team. 2 new project activities were identified and are being evaluated through consultation with users in EU countries and agencies. These activities are in the areas of (a) pyrogen testing in rabbits, for which Ph. Eur. monographs encourage replacement of in vivo pyrogenicity tests by suitable alternative methods but the rabbit test continues to be used widely, and (b) the use of animal studies in the non-clinical development of monoclonal antibodies including the potential to improve and reduce 6-month repeat-dose toxicity studies.
Waiving of 2-year carcinogenicity studies
The objectives of this project are to identify opportunities for improving the science supporting the regulatory testing of agrochemicals, and to achieve a reduction in the use of animals when assessing the potential for carcinogenicity.
The project was launched in 2017 as a follow-up of a previous, successful EPAA project on the prediction of carcinogenicity of pharmaceuticals which provided evidence that in many cases a 2-year carcinogenicity study in rats could be waived without compromising human safety. The waiver could be applied based on prior knowledge of the pharmacological properties of the compounds in question, integrated with histopathological findings from 3-6 month repeat dose toxicity studies and together with evidence for lack of genotoxic potential and lack of hormonal perturbation. The conclusions were based on data analysis of 289 pharmaceutical compounds and demonstrated a prediction rate of 92% and 98% for noncarcinogens and for carcinogens, respectively.
Two-year carcinogenicity studies are part of the regulatory requirements for pharmaceuticals, food additives, chemicals and agrochemicals. Such studies entail the use of large numbers of animals. Currently, to assess the potential for a non-genotoxic compound (i.e. not inducing DNA damage) to increase the risk of cancer in humans, 2-year carcinogenicity studies in rats or mice are performed. The relevance to the human safety of data from rodent carcinogenicity studies has often been questioned but still this type of study remains the default requirement. Regulatory requirements also include repeated dose toxicity studies of 3-6 months duration for compounds intended for long-term administration.
3Rs added value and EPAA actions
The expected impact to the 3Rs is substantial. Based on the results obtained in the previous EPAA-supported project for pharmaceuticals (Van der Laan et al. 2016), the number of agrochemicals requiring a carcinogenicity study may be reduced by 40-60%. Since the proposed approach makes use of sub-chronic (3-month) in vivo toxicity tests that are performed anyway, each agrochemical for which a carcinogenicity study may be waived would save a large number of animals from being tested for a long period of time. If successful, this project could be possibly extended to other types of chemicals in other industrial sectors.
The project is supported and coordinated by EPAA and is being conducted by RIVM (National Institute for Public Health and the Environment, The Netherlands). The project team includes members from industry and regulatory bodies, EURL ECVAM, as well as researchers from the previous pharmaceutical-focused project.
- This 2-year project was launched in 2017 as a follow-up of a previous, successful EPAA project on the prediction of carcinogenicity of pharmaceuticals.
- To test the approach, case studies are being prepared and will be discussed in an expert Workshop planned for 12-13 June 2019, in Brussels. Regulators of the agrochemical sector, relevant industrial partners and stakeholders such as EURL ECVAM and OECD will be invited to the Workshop.
- The report will be prepared for publication in a peer-reviewed journal.
Next steps will be decided by the end of 2019 after the current project has been completed.
Training and knowledge-sharing
This project aims to promote training and knowledge-sharing regarding the application of non-animal strategies for assessing skin sensitisation.
A number of workshops on skin sensitisation have been organised by EPAA in previous years (2013, 2015) in collaboration with Cefic LRI, Cosmetics Europe and other stakeholders. In the last years, several tests addressing key events (KE) 1, 2 or 3 of the adverse outcome pathway (AOP) for skin sensitisation have appeared in the OECD test guidelines. Moreover, since 11 October 2016, the new REACH requirements for skin sensitisation entered into force making non-animal testing the default requirement. Therefore, a new knowledge-sharing cross-sector workshop was considered to be timely in 2019 and was organised in February targeting experts from industry and regulatory authorities. In order to facilitate participation of the latter, the workshop was hosted by ECHA in Helsinki.
3Rs added value and EPAA actions
The knowledge-sharing envisaged through this project is expected to improve communication about the scientific and regulatory realities. It will indicate to companies, regulators and validation authorities the advantages and potential limitations of the non-animal approaches that are currently the only possibility in the cosmetics sector, the default methods in REACH regulation and strongly encouraged in other pieces of EU legislation. This also has the potential to enhance coordination of the various research initiatives in this field, and make progress towards replacing animal testing for the sensitisation endpoint.
The 2019 workshop was co-organised by EPAA, Cefic LRI and IFRA Europe and was hosted in Helsinki by the European Chemicals Agency (ECHA). More than 60 experts participated at the event, including from ECHA and EU country regulatory agencies and industry sectors.
- This project started in 2018 and focuses on training and knowledge-sharing in the application of non-animal strategies for assessing skin sensitisation.
- A major activity was the workshop on 7-8 February 2019, at ECHA (Helsinki), with contributions from ECHA, OECD, EURL ECVAM and SCCS. See the flash report and documents and watch the video
- The workshop report is currently in preparation and will be published in a peer-reviewed journal.
To be defined later after publication of the workshop’s report.
The project aims to organize a multi-stakeholder Workshop that will generate new ideas for non-animal approaches to predict repeated-dose systemic toxicity.
The problem of predicting repeated-dose systemic toxicity with non-animal methods remains unsolved. Although there have been excellent results in predicting acute, site-of-contact effects, progress on repeated-dose, systemic toxicity has been limited. There are many reasons for this: the numerous modes of action that can cause toxicity, many of which are still undefined, the interplay among different cell types, temporal aspects of toxicity, including the relationship between repeated lesions and repair, and pharmacokinetic and metabolic considerations that are only imperfectly recapitulated in vitro. The complexity of this problem raises it to the level of one of the grand challenges of modern science. Recognising the complexity of this problem, and the urgency in solving it, in 2008 the EPAA convened a group of thought leaders from basic life sciences, chemistry and medicine to brainstorm possible ways forward. Most of what emerged from that ‘New perspectives on safety’ Workshop was a list of possible technologies that might be helpful in constructing predictive models. This became the framework for SEURAT-1, the large, multi-centre programme funded later by Cosmetics Europe and the European Commission. While SEURAT-1 created many useful tools, it has become clear that this programme was not sufficient to deliver alternatives for systemic toxicity.
3Rs added value and EPAA actions
The recommendations of the workshop and the publication reporting from it will be used to inform/advise future research strategies of the EPAA partners and other stakeholders, thereby increasing the chances to accelerate the development of future non-animal integrated approaches/strategies for assessing repeated dose toxicity.
The project team includes experts from different industry sectors, the E. Commission and EURL ECVAM. The workshop will be organised with international experts from different scientific disciplines coming from industry, academia and regulatory bodies.
- The workshop will take place in October 2019, in Brussels.
- The workshop summary with recommendations will be published in a peer-reviewed journal.
The workshop conclusions and recommendation will be presented at the 11th World Congress on Alternatives and Animal use in life sciences (WC11), in Maastricht, in 2020.
The objective of this project is to test the effectiveness of a computational algorithm developed to convert in vitro concentration-response data to in vivo dose-response data (known as QIVIVE) and its applicability to a range of chemical structures. More specifically, the project aims to
1. Increase confidence in the approach by demonstrating applicability with diverse chemical structures
2. Demonstrate with representative compounds a robust evidence of applicability on a broader landscape and across different industrial sectors
3. Provide evidence of a tool that may be incorporated within R-Vis, specifically designed for QIVIVE, which will be unique for modelling software platforms
4. Compare the predicted in vivo BMD to existing experimental BMD values used in chemical safety assessment by a regulatory agency
The overall context of this project is the development of a reliable non-animal in vitro bioassay-based testing strategy for human safety testing of chemicals. Specifically, the aim is to test the effectiveness of a computational algorithm that was developed earlier to convert in vitro concentration-response data to in vivo dose-response data (known as quantitative in vitro to in vivo extrapolation or QIVIVE) and its applicability to a range of chemical structures.
The algorithm could be incorporated as a tool in a future third phase of development of the earlier Cefic-LRI and EPAA funded user-friendly, freely available modelling platform called R-Vis. This algorithm was not a deliverable of the R-Vis project but was developed separately to address limitations in current approaches to QIVIVE which use the term ‘reverse dosimetry’. It would therefore expand the existing reverse dosimetry capability of R-Vis.
3Rs added value and EPAA actions
An automated computational algorithm that accepts in vitro concentration response data as an input and returns in vivo dose response data as an output would represent a significant milestone in the development of tools that could contribute to the 3Rs.
The outputs from this work, which include characterising specific uncertainties associated with the computational tools and in vitro data used, will be presented according to the OECD PBK template currently under development. The suitability, utility and concordance of in vitro assay systems could be readily and reasonably examined. This would further encourage and facilitate uptake of alternative to animal methods.
The project is supported by EPAA and is conducted by HSE (Health Safety Executive, UK). In addition, experts from the European Commission, the Joint Research Centre, EURL ECVAM, EFSA and interested companies contribute to the project.
• This 2-year project started in 2019
• Build PBPK model, perform sensitivity analysis, and identify most sensitive parameters
• Translate in vitro concentration responses to in vivo
• Determine in vivo BMD values
• Submit manuscript for publication in the peer-reviewed literature
To be defined after completion of the project.
The project aims to re-evaluate regulatory practices from a non-clinical perspective focusing on monoclonal antibodies, building on previous research experience at the Dutch Medicines Evaluation Board (MEB) with support from EPAA, several pharmaceutical companies and the NC3Rs.
The results will demonstrate ongoing commitment of both regulatory agencies and industry to strive towards initiating fewer animal studies without compromising the ability to assess benefit and risk.
In order to evaluate the safety and efficacy of new drugs or indications, it is often necessary to conduct animal studies. These animal studies have evolved over time and are embedded in (inter-) national guidance and legislation. However, the translational and predictive value of animal studies is increasingly being debated and questioned in the public, scientific and regulatory community. With technological advances being made, new opportunities are emerging to further implement 3R principles (refinement, reduction and replacement) in drug development. In addition, re-evaluation of regulatory guidance can further provide opportunities to restrict the use of animals in safety and efficacy studies to those which provide meaningful information that is relevant to humans.
3Rs added value and EPAA actions
This project provides a scientific basis to further reduce non-clinical studies for specific biotechnology products (monoclonal antibodies) on a case-by-case basis. While the gains could be seen as modest at face value, they are an important basis to demonstrate the ongoing commitment of both regulatory agencies and industry to strive towards initiating fewer animal studies without compromising the ability to assess benefit and risk.
In addition to the Medicines Evaluation Board (MEB), experts from the European Commission, pharmaceutical companies and the NC3Rs contribute to the project.
- Establish criteria on the basis of which decisions can be made on the need and duration of non-clinical safety studies for monoclonal antibodies based on approved and non-approved drug development programmes
- Submit a publication to a scientific journal detailing the evidence that supports the implementation of shorter safety studies.
- Organise a workshop to discuss and disseminate the results, offering a potential starting point for formal discussions between agencies and companies at international level.
The results of the project could provide a scientific basis to further reduce non-clinical studies for specific biotechnology products (monoclonal antibodies) on a case-by-case basis.
This project aims to review the progress in PBK, assess the chemical space coverage of existing PBK models and determine methods to identify ‘similar’ chemicals. More specific project aims are to
- undertake and publish a complete systematic review and collation of existing, published PBK models in rats and humans in order to provide a readily updatable resource for PBK model developers and users.
- assess the chemical space coverage of existing PBK models in relation to the chemical space of food additives, drugs, cosmetics, pesticides and industrial chemicals. This will help to identify where gaps exist in current knowledge and which regions of chemical space are associated with higher confidence in potential read-across.
- investigate a range of similarity assessment metrics (e.g. chemical fingerprints) to determine which individual or consensus method leads to most appropriate selection of source chemicals to predict behaviour/properties of target chemicals and serve as templates in PBK model development.
- develop a freely available software tool to assist the identification of appropriate chemicals to use as templates via an automated workflow.
Increasing confidence in the use of non-animal alternatives for safety assessment is an important goal for many sectors. Read-across is increasingly being used as an alternative, whereby data from one or more source chemicals is used to predict the effect of a target chemical of interest. For any chemical (food additive, drug, cosmetic, pesticide etc) to have an effect, the chemical (or its transformation product) must not only possess intrinsic activity, but must also reach the relevant site of action at sufficient concentration. Hence, for more reliable risk assessment consideration must be given to both intrinsic activity and internal exposure. physiologically-based kinetic (PBK) models are used to predict the overall time-concentration curves for chemicals in blood/organs; they are increasingly being used by industry, academia and regulators. The models can be used in conjunction with pharmacological or toxicological information in order to determine the true potential of a chemical to elicit an effect – desirable or undesirable.
Also, the 2017 EPAA partners forum on ‘toxicokinetics and read across’ reiterated the importance of in silico toxicokinetics in this context.
3Rs added value and EPAA actions
A systematic review of existing PBK models that may serve as templates for the development and evaluation of new PBK models will be published, providing a valuable data resource for academia, industry and regulators. Using existing models and data as templates will help to reduce the number of new animal tests for PBK model development. Detailed methodology for the review would also be provided ensuring that the review could be readily updated as more information becomes available, ensuring sustainable impact.
Read-across is increasingly recognised as an important non-animal alternative in safety assessment; however, barriers to its acceptance and uptake remain due to lack of confidence in the predictions. Incorporating information from PBK models into the read-across prediction provides additional information and support of the prediction; this may help to promote the acceptability of read-across as an alternative to animal testing.
The project is supported by EPAA and is conducted by the Liverpool John Moores University (LJMU). Furthermore, experts from EURL ECVAM, the US Environment Protection Agency, HSE and industry contribute to the project.
The project will start in the autumn of 2019 and will last for 3 years. The following tasks and milestones are foreseen
- Development of systematic review protocol and pilot study
- Completion and publication of systematic review of existing PBK models (output 1)
- Analysis of chemical space of PBK models; comparison to food additives, drugs, cosmetics, pesticides and industrial chemicals
- Assessment of relevant similarity metrics to be used for selection of template chemicals
- Case studies part 1 – Development of new PBK models using selected template
- Case studies part 2 – Sensitivity analysis (e.g. RVis); effect of chemical selection on model reliability
- Optimised PBK model for target chemical based on model for appropriate template chemical
- Development of a software tool for rational chemical selection using PBK models (output 2)
The systematic review of PBK models will be published in the scientific literature and registered. This allows for the review to be updated in the future when more models become available providing a sustainable resource. The open source software tool developed under the project will be made freely available for all users.
The project’s dissemination aspect is a key pathway to ensure further impact. Several targeted activities will be used to promote widespread uptake of the methodology, e.g. publications, conference presentations, web-based tools/webinars and promotion via existing networks of collaborators in academia, industry and regulators, in Europe, Canada and the US, who strive to increase the acceptance of non-test methods in PBK modelling.
The aim of this project is to identify opportunities for improving the science behind the regulatory testing of medicines and chemicals through the application of the 3Rs. Among the seven sectors involved primarily in this project are those concerned with the development of human medicines, veterinary medicines, and crop protection products.
- The main goal is to examine how each sector approaches the issue of regulatory toxicology and to identify opportunities for cross-sector alignment on best practice and on the introduction of new methodologies advancing the 3Rs.
- Alignment has 2 aspects: industry alignment on testing methodologies, and regulatory harmonisation across sectors and global regions.
- Thus the project aims not only at identifying possibilities but also how these might be translated into regulatory practice
- Pharmaceuticals, animal health, crop protection
Development and priorities
The project team initially conducted a survey of regulatory requirements in the various sectors and then sent a questionnaire to the relevant EPAA member associations (EFPIA, IFAH-Europe and ECPA).
Respondents were asked for more detailed information on
- the scope for variation in study design within the existing guidelines
- regional variations in regulatory requirements
- individual company practice
- the development of alternatives that could advance the 3Rs
The questionnaire was distributed to the member companies of the aforementioned organisations and the results were collated and analysed by the project team.
It was clear that there is a considerable divergence in practice within sectors, between sectors and between geographical areas, despite the existence of international harmonisation bodies such as ICH and VICH.
Moreover, the differing practice was as likely to be a result of tradition than the application of science.
Following these discussions, the Project Team selected the area of carcinogenicity testing as one that offered great potential for the project. Therefore, all sectors need to consider the carcinogenic potential of their products. Sector practice is quite divergent, the scientific value of some study designs is currently being questioned and the introduction of in vitro methods offers the possibility of a more targeted and progressive approach to animal testing.
- 28 February 2013: EPAA workshop on carcinogenicity testing: scope for harmonisation and advancing the 3rs flash report (87 kB)
- Ongoing discussions on MTD and human effects database
Since late 2008, the EPAA has been exploring the opportunities that stem cells could play in developing novel approaches for the potential hazard characterisation of chemicals and drugs
- Much research on stem cells is currently ongoing within the EU and beyond, and this is considered valuable in assessing how such work may fit into an overall strategy for evaluating safety without animal testing
- This EPAA project aims to direct the stem cells research towards the industry need for safety assessment
- In the opinion of the EPAA platform on science, the current research on the use of stem cells lacks clear guidance to develop novel testing strategies for safety assessment
- The identification of gaps in testing strategies is relevant in order to select the cell types of interest, as well as the most appropriate readout to identify modes of action. This is important for multiple sectors and therefore a good opportunity for the EPAA to coordinate this activity.
- Workgroup 1 on fundamental research on stem cells
- Workgroup 2 on communication: stem cells forum
Project added value on 3Rs and priorities
- establishing a permanent stem cell network, which brings together experts from both sides (those working on alternatives and those working on basic research)
- setting up a central unit monitoring the quality of the stem cells
- focusing on inducible pluripotent stem cells (iPSC)
- focusing on pathways, instead of organs or cell types is preferred
- noting that first steps in analysing the signal cascades of human iPSC should be performed with well-known compounds that address a well-defined pathway
- 28-29 April 2008: EPAA new perspectives on safety workshop, flash report (480 KB)
- October 2009: Expert meeting on the future directions of stem cells research for safety testing
- 4-5 October 2011: Stem cells and their derivatives in toxicological research programmes and as a possible regulatory tool – A gap analysis, flash report (710 KB)
- 25 April 2013: EPAA stem cells Forum kickoff meeting (WG3), flash report (360 KB)
- 28-29 August 2013: Stem cell-derived organ-like models for analysis mid-term and long-term dosing dynamics, flash report (135 KB)
- August 2014: Joint EPAA-Liverpool University CDSS stem cells forum international meeting
Project aim and rationale
The risk assessment of chemicals and pharmaceuticals continues to rely on the use of in vivo assessments, although there is an international drive towards the replacement of animal testing with mechanistic, in vitro systems.
This new approach includes the in vitro measurement of concentration-response relationships to identify where prolonged or excessive perturbations of biochemical pathways are likely to cause adverse health effects.
- This approach requires a framework, into which a wide range of in vitro, in silico and in chemico generated data, may be integrated and used to predict the consequences of exposure in humans.
- One of the major drawbacks of most in vitro methods compared to intact organisms is indeed their inability or low competence to metabolise xenobiotics. To address this issue, the EPAA launched the absorption, distribution, excretion & metabolism (ADME) project with the following goal: to develop a new, innovative bioinformatics tool to predict the ADME or human exposure based on inputs from in vitro data (and vice versa).
The tool will be web based on a Peters’ model and freely available for downloading and independent use on desktops PC to ensure confidentiality of data.
A prototype was already developed in partnership with the British Health and Safety Laboratory (HSL). This project aims to develop a tool that is more tailored to industry needs. The first tasks to be undertaken included the building of the model and interface. These were completed as planned, within the allocate 6-month period.
UK Health and Safety Laboratory, GlaxoSmithKline, European Council of the Chemical Industry - Long-range initiative (CEFIC LRI), European Union Reference Laboratory - European Centre for Validation of Alternative Methods (EURL ECVAM), European Centre for Ecotoxicology and Toxicology of Chemicals (ECETC)
- November 2008: EPAA In vitro absorption, distribution, excretion & metabolism (ADME) in safety testing workshop to evaluate jointly with regulators, what information was concerning metabolism is needed to permit safety assessment of substances.
- October 2011: EPAA –ECVAM joint ADME workshop on the potential for further integration of PBTK modelling into the prediction of in vivo dose-response curves without animal experiments
- June 2014: Launch of the latest project workstream on user-friendly, web-based tools for toxicokinetics in exposure assessments
- Bessems et al., Reg. Tox. Pharmacol., 68 (2014) 119-139): 'PBTK modelling platforms and parameter estimation tools to enable animal-free risk assessment. Recommendations from a joint EPAA – EURL ECVAM ADME workshop'
- Peters, S.A., 'Evaluation of a generic physiologically based pharmacokinetic model for lineshape analysis.' Clin. Pharmacokinet., 2008. 47(4): p. 261-75.
This project was supported by EPAA during 2006-2012.
Project aims and background
Transferring successful approaches from one sector to another is a deliberate strategy of EPAA. In 2006-2008, the EPAA devoted attention to whether a valuable alternative test developed for agrochemicals by the ILSI/HESI project on agricultural chemical safety assessment (ACSA) could also be applied to other sectors such as industrial chemicals. The feasibility of the ACSA extended one-generation study protocol was evaluated.
The ACSA testing proposal was originally designed as an intelligent testing strategy for the evaluation of agrochemicals. It aimed to address inefficient development of data, much of which is not used in the final risk assessment. This provided a scientifically robust approach to reduce and refine the studies required for registration of agrochemicals. One of the core elements was the extended one-generation reproductive toxicity study proposed to replace the classical two-generation study (OECD 416).
Potentially saving a substantial number of animals, the EOGRTS design was considered as an alternative to the two-generation test (OECD TG 416) with potential applicability and impact on chemical safety assessment across many industry sectors. The OECD states that the EOGRTS can provide all the information regulators need to make a proper safety assessment of a substance without automatically requiring the use of a second generation of animals, thus potentially saving substantial numbers of animals per test.
Experts from companies (chemical, crop protection, soaps & detergents), EURL ECVAM, ECHA, national and international regulators
A 2006 EPAA workshop concluded that the extended one-generation study could, in principle, be applicable to safety testing under REACH. However, it was also agreed that the complex ACSA protocol would have to be modified to meet the requirements for industrial chemical safety testing. The EOGRTS addresses far more endpoints than currently required for chemical risk assessment. Therefore, the ACSA test protocol for use under REACH needed modification to design reliable triggering and/or waiving criteria for the components of the protocol as modules.
In 2008, a task force of the European Centre for Ecotoxicology and Toxicology of Chemicals developed these criteria (ECETOC Doc. No. 45, 2008). A subsequent April 2008 workshop by the ECETOC and EURL ECVAM discussed the criteria, their relevance, and possible validation needs with invited representatives of industry, academia and the regulatory community.
The OECD held a number of expert discussions in 2010 to advance the approach.
Studies on model compounds, based on the ACSA protocol, were conducted by industry partners of EPAA (BASF, Bayer CropScience, Dow Agroscience, and Syngenta). The results from these feasibility studies have been published in a peer reviewed journal1.
[1Fegert et al. Feasibility of the extended one-generation reproductive toxicity study (OECD 443). Reprod Toxicol. 2012 Nov; 34(3):331-9]
In July 2011 the OECD endorsed the extended one-generation as test guideline 443.
Following the adoption of the test method by the OECD, an expert sub-group under CARACAL (competent authorities for REACH and CLP) was tasked with considering all the options to apply it and to explore how it could be used in the context of REACH. Through the EPAA, industry was invited to contribute to this discussion with information on costs and practical issues related to applying the test guideline. Cefic coordinated the industry response.
Practical questions related to the use of EOGRTS were discussed during further CARACAL meetings. The Commission included EOGRTS in the test method regulation and has modified the corresponding REACH annexes. The design of EOGRTS under REACH regulation was modified further by a set of triggers for the use of F2 generation and DIT, DNT cohorts to underline further the 3Rs aspects.
In 2018, the European Commission published the communication 'Towards a comprehensive European Union framework on endocrine disruptors in the different legislative frameworks to improve identification of endocrine disruptors'. It highlights a number of initiatives which are also focused on the EOGRTS test. These can provide useful information about ED properties, as far as the study is considered in its full scope, (i.e. inclusion of the use of F2 generation and DIT, DNT cohorts). This will most probably have an impact on the current chemical legislation.
The overall goal of this platform is to synchronise initiatives for 3R methods in safety and potency testing of vaccines in Europe.
This project aims to create a technical platform for human and veterinary vaccines. The platform
- includes general strategies and policies to introduce the consistency approach in routine release activities
- prioritises the replacement of compendial in vivo tests with validated alternative in vitro tests
- defines minimal acceptance criteria for alternative in vitro tests
- reviews work plans and reports provided by human and veterinary vaccine task forces or groups working on test development and validation
Vaccines require batch-specific quality control to ensure their quality, including safety and efficacy. Part of quality control, particularly for the final product, is based on animal tests required by legislation. Out of about 100 million animals that are used each year in laboratories throughout the world, 10 to 15 million animals are still being used for vaccine batch testing.
The vaccines consistency approach (VCA) for batch release is based on a thorough characterisation of the vaccine during manufacture, including formulation, using non-animal testing. The quality of subsequent batches is guaranteed by the strict application of quality systems and of consistent production of batches that are comparable to reference lots of known potency and safety.
The VCA is already used for recently registered vaccines, whereas many vaccines developed several decades ago, continue to rely on animal tests for confirming the quality of each batch.
Due to the potential of the VCA to significantly reduce the number of animal tests used in vaccines quality control, the EPAA initiated this project with the aim to provide a framework for resolving remaining scientific and technical issues and for fostering the regulatory adoption of VCA as a non-animal approach for quality control of established (conventional) vaccines.
The European Commission, industry (European vaccines manufacturers), European Medicines Agency (EMA), national regulators, OMCLs (Official Medicines Control Laboratory), EDQM (European Directorate for the Quality of Medicines and HealthCare), international regulators (observers from the US, Canada and India) and academia.
The project activities were organised by a project committee (PC) chaired by the coordinator (Ian Ragan, consultant to EPAA) in consultation with a technical committee (TC) composed of experts from vaccine manufacturers, EDQM, EURL ECVAM, OMCLs and regulatory authorities for both human and veterinary vaccines. The TC was chaired by Coenraad Hendriksen, academic member of the PC.
At the technical committee meeting on 30 September 2011, 4 priority vaccines were identified and progressed further by expert working groups: DTaP, human rabies, veterinary rabies, and clostridial vaccines.
January 2010: Workshop jointly organised by EPAA and EURL ECVAM in Brussels discussed the VCA and its potential to reduce the number of animal tests used in quality control of human and veterinary vaccines (De Mattia et al 2011, The consistency approach for quality control of vaccines: A strategy to improve quality control and implement 3Rs, Biologicals 39, p 59-65). As a follow-up, in late 2010 the EPAA agreed to initiate a VCA project.
7 April 2011: Kick-off meeting - Application of the 3Rs and the consistency approach for improved vaccine quality control, flash report (265 KB)
Milestones of the specific working groups
- Workshop, 30-31 August 2012,flash report (409 KB)
- No activities started under EPAA umbrella since several activities were ongoing elsewhere (to avoid duplication)
- In 2016: activities supported by IMI 2 funding programme (VAC2VAC – IMI 2 Call 3)
2. Human rabies
This workstream aims at replacing the current in vivo immunisation challenge test for batch release (the NIH test) by in vitro tests based on antigen quantification (ELISA).
- Workshop, 8-9 October 2012,flash report (264 KB)
- Workshop, 9-11 May 2015 Joint veterinary & human rabies vaccines workshop (report in preparation, to be published soon). Agreed that in vitro ELISA is acceptable to replace current challenge/serology tests
- Collaborative study to identify most suitable ELISA was finalised in 2015
- Global agreement reached regarding the way forward
- In 2016: in cooperation with EDQM (Council of Europe), the preparation of a formal validation study supported by EPAA
3. Veterinary rabies
- Workshop, 5-6 November 2012,flash report (403 KB)
- Activities ongoing at manufacturer level
- Cross-fertilisation from the learnings of the Human rabies project
- In 2016: supported by the IMI-2 funding programme (VAC2VAC – IMI 2 Call 3)
4. Clostridial vaccines
This work stream aims at replacing in-process controls which use testing on animals, with cell-culture-based assays that have been developed by MSD animal health with support by the NC3Rs. The application of the consistency approach is feasible for clostridial vaccines but in vitro tests remain to be developed for some important strains.
- Workshop, 19 March 2013 (co-organised with EDQM, Council of Europe), flash report (138 KB)
- Workshop, 11 September 2013 (co-organised with EDQM, Council of Europe), flash report (426 KB). This workshop helped prepare the clostridium septicum vaccine collaborative study (BSP130). The EDQM BSP130 study was successfully completed in 2015 and demonstrated concordance of in vivo and in vitro methods
- Workshop, 15-16 September 2015 (co-organised with EDQM, Council of Europe, in Egmond aan Zee, the Netherlands). This workshop discussed the results of the BSP130 study as a proof of concept and start of validation for clostridium septicum vaccine in process control methods
- In 2016: a phase III study (formal in vitro test full validation) will be progressed as part of BSP130 and with the support of EPAA
The final aim is to introduce alternative methods into the Ph. Eur. relevant monograph(s).
- One peer-reviewed publication: De Mattia et al 2015, The vaccines consistency approach project: an EPAA initiative, Pharmeuropa Bio& SN, May 2015, p 30-56
The technical committee, where the Commission provided visibility and all parties committed to alternatives shared scientific know-how, has
- raised awareness in all stakeholders (industry, regulators, civil society) on the vaccines consistency approach
- facilitated cross-sharing between human and veterinary vaccines helping to promote new testing paradigms
- offered coordination, identified scientific gaps and enabled early collaboration helping to address them
- prepared the ground for the IMI-2 Vaccines consistency project to work towards ultimately replacing animal testing in the batch release of vaccines. This project has been accepted and will start in March 2016. With funding of approximately €8 million, this project will help develop new methods, work towards their validation and ensure regulatory engagement with an ultimate aim to lead to acceptance by regulators, implementation and use by industry
The EPAA has
- provided funding and organisational support for the regular expert meetings, 7 workshops and 2 peer-reviewed publications (De Mattia et al. 2011 and 2015).
- provided a neutral, scientifically based platform for regular discussions and exchanges between the various stakeholders facilitating the development, validation and acceptance of the approach
- supported specific collaborative studies (e.g. by funding shipment of testing materials for the clostridials collaborative study BSP130, the European Commission (via EURL ECVAM) funded data analysis within the human rabies vaccines project)
- promoted the VCA through presentations at EPAA annual conferences and other events (e.g. briefings to MEPs / intergroup for animal welfare)
- December 2015: Activities under the general VCA project are completed.
- 2016: Human rabies and Clostridial vaccines continue as stand-alone EPAA projects (see above).
The EPAA partners would like to thank the members of the technical committee and project committee for their valuable contributions to the project.