Méthodologie

Risk Assessment is increasingly generating information using new approach methodologies (NAMs) as alternatives to animal testing. Yet, chemical risk assessments are still based mainly on toxicological data using animal models and many efforts are ongoing at EFSA, and other European and International agencies to integrate such NAMs in next generation risk assessment. In addition, European Commission services will publish a road map for the phasing out of animal studies, requested by over 1.2 million citizens to the European parliament, to further support these efforts and provide practical short-, mid- and long-term solutions.

Objectives: Develop in silico models and an open access platform: TKPlate to support the integration of NAMs in next generation risk assessment of chemicals for human, animal health and environmental risk assessment with a focus on toxicokinetic (TK) and toxicodynamic (TD) models.

Methodology: The project focuses on the development of TK and TD models for human populations, test species and farm animals and their integration in the TKPlate platform. These include physiologically based kinetic models in humans, test species and farm animals, TD models for benchmark dose modelling, dynamic energy budget models for species in the environment and other more complex models investigating mixture toxicity and impact of multiple stressors on humans and animal species.

When setting health-based guidance values (HBGVs) uncertainty factors are applied to extrapolate animal data to humans and to consider inter-human variability. Yet, little is known about toxicodynamic variability in humans, and it is unclear if the uncertainty factors are sufficiently protective.

Objectives: Improve chemical risk assessment and refine HBGVs by addressing human variability in toxicodynamics (TD), more specifically understanding how differences in how individuals respond to toxic substances at cellular level, particularly in terms of cell stress and cell death pathways.

Methodology: The project addresses human variability in TD by generating cellular and transcriptional response data following the activation of specific stress and cell death pathways. An in vitro cell model of human peripheral blood lymphocytes is used as a source of freshly isolated cells from 150 individuals. Bayesian statistics is applied to model inter-individual differences.

Developmental neurotoxicity (DNT) is a serious public health concern and better knowledge of risk factors is crucial to minimise new cases of neurodevelopmental disorders and cognitive deficits.​ Several studies indicate that to explain current incidence of brain disabilities in humans, exposure to chemicals must be considered. ​Current DNT in vitro testing battery has gaps in coverage of neurodevelopment processes and cell types.

Objectives: Enhance the understanding of how chemical exposures affect brain health and development, focusing particularly on glial cells. The project develops and implements New Approach Methodologies (NAMs) to assess the toxicity of food toxicants on glial cell development and function, addressing existing gaps in developmental neurotoxicity testing.

Methodology: The project employs an integrative approach that combines transcriptomics, in vitro assays, and Adverse Outcome Pathway (AOP) development. It involves the creation of novel in vitro models to predict glial-specific toxicological events, the expansion of existing AOP networks to incorporate glial-related key events, and the use of transcriptomics to generate and analyse relevant datasets. The findings will contribute to refining risk assessment frameworks and regulatory guidelines on DNT.

The lack of specific data requirements for pesticides residues, i.e. metabolites, in crops and/or animal products, e.g. milk, limits the harmonisation of risk assessment which has to be carried out on a case-by-case basis.

Objectives: Integrate New Approach Methodologies (NAMs) into the risk assessment of pesticide metabolites. The project is developing semi-automated and standardised workflows to improve the efficiency, transparency and harmonisation of pesticide metabolite assessments, reducing reliance on traditional testing methods.

Methodology: The project involves the creation of a semi-automated workflow for mutagenicity assessment using the OECD QSAR Toolbox. Case-studies are conducted to demonstrate the practical implementation of these workflows, overall contributing to regulatory guidance on pesticide metabolite risk assessment.

The EU Green Deal aims at reducing the risk associated with pesticides use and reversing biodiversity loss. The development of risk assessment methodologies for non-target organisms could contribute to such ambitions.

Objectives: Advance Environmental Risk Assessment (ERA) of plant protection products (pesticides) by integrating landscape and ecological features into risk assessment methodologies. The project addresses knowledge gaps in terrestrial ecotoxicology and enhances the assessment of non-target organisms in agro-ecosystems.

Methodology: The project involves the collection and generation of data to improve exposure and sensitivity assessments of non-target organisms. It also focuses on the identification of tools, such as population models, to assess effects at higher biological levels. Additionally, the feasibility of developing an interoperable environment for risk assessment is explored. The findings will support the revision and development of guidance documents to enhance ERA methodologies.

Historical Control Data (HCD) are data from studies conducted under the same or similar conditions as the toxicity study under assessment. Their primary use is to support the interpretation of (possible) treatment-related effects in the toxicity study. However, there is no harmonised approach on the collection, evaluation and use of HCD.

Objectives: Elucidate the requirements for the use of HCD to better interpret toxicity studies used for regulatory purposes; clarify how HCD should be compiled as part of the regulatory dossier; and assist the interpretation of HCD in the context of the evaluation of regulatory studies.

Methodology: Information from the scientific literature, publicly available databases and stakeholder experience (survey, workshop and technical hearings) have been used. The EFSA Panel on Plant Protection Products and their Residues (PPR) developed a quantitative approach for the collation, evaluation and use of HCDusing a decision scheme which includes seven distinct steps grouped in three overall clusters of activity: planning, evaluation of the HCD, and use of HCD. In addition, illustrative case studies are available to elucidate the proposed methodology and templates offered to improve harmonisation for data submission.

European law requires that no active substance can be authorised if it causes endocrine disruption i.e. has an adverse effect on endocrine activity. To support this requirement, it is necessary to better understand the endocrine disrupting mode of action of active substances.

Objectives: Develop Adverse Outcome Pathways (AOPs) for the identification of substances with endocrine-disrupting properties, addressing regulatory challenges during the assessment of endocrine activity, particularly within the estrogenic, androgenic, thyroidal, and steroidogenic modalities.

Methodology: A top-down approach is used to construct AOPs, identifying key molecular initiating events and key events linked to endocrine disruption. Collaboration with the European Chemicals Agency (ECHA) and the Joint Research Centre (JRC) ensures harmonisation with existing frameworks. The developed AOPs will be submitted to the OECD AOP Wiki for broader regulatory adoption.

Project results: The project delivered a comprehensive Adverse Outcome Pathway (AOP) framework for endocrine disruption assessment. The project outputs are now integrated into EFSA's regulatory assessments of pesticides and biocides, enhancing the scientific basis for endocrine disruptor identification and supporting reduced animal testing.

Some pesticides may have the potential to disrupt brain development. It is crucial to assess developmental neurotoxicity of chemical substances and to accelerate the development of in vitro methods to test the impact of chemicals on nervous system development.

Objectives: Improve the assessment of environmental neurotoxicants by developing an integrated in vitro neurotoxicity testing strategy, advancing hazard identification for developmental neurotoxicity (DNT) and other forms of neurotoxicity associated with chemical exposures, particularly pesticides.

Methodology: The project involves the development of a comprehensive DNT testing battery, incorporating in vitro assays and computational modelling to complement or replace in vivo testing. Data generated from these assays is analysed to refine assessment frameworks, with guidance documents developed to support regulatory implementation. Collaboration with the OECD, the Danish Environmental Protection Agency, and the US Environmental Protection Agency ensures alignment with international best practices.

New Approach Methodologies (NAMs) are considered promising for the assessment of nanomaterials, reducing the use of animal testing. ​Nanotechnology is a cross-cutting area impacting novel foods, feed additives, and food contact materials.

Objectives: Develop recommendations on the use of NAMs for nanomaterials and nanoparticles risk assessment in food and feed, and foster NAMs’ implementation within EFSA’s regulatory evaluations. The goal is to provide guidance and case studies to support EFSA Panels and applicants in adopting these innovative methodologies, minimising animal testing while moving towards mechanistic-based risk assessment. 

Methodology: The project identifies relevant data sources, establishes protocols, generates data and develops tools for integrating NAMs into risk assessments. The development of a qualification system for NAMs and NAM-based integrated approaches to testing and assessment case studies demonstrate the practical applications. Engagement with stakeholders and other scientific advisory bodies ensures alignment with international developments and regulatory relevance. Lastly, initial provisions for guidance on NAMs data integration and implementing hypothesis-driven Next Generation Risk Assessment is being developed, with specific focus on the area of food and feed nanotechnology.

There is resistance among some scientists towards the use of in vitro New Approach Methodology (NAM) test systems because of doubts about their representation of human physiology, absence of a consensus framework for interpreting toxicogenomics data, and the need for advanced bioinformatics tools for hazard characterisation.

Objectives: Advance chemical safety assessment by incorporating toxicogenomics and mechanism-based risk evaluation to develop a methodology for interpreting transcriptomics data, establish a quantitative framework and integrate bioinformatics for mapping Adverse Outcome Pathways.

Methodology: The project combines computational modelling, bioinformatics, and toxicogenomics to enhance risk assessment. It develops algorithms for transcriptomics interpretation, designs a prototype toxicogenomics tool, and aligns methodologies with international standards.

The project addresses the increasing complexity of novel food/feed products containing protein and the need to modernise protein safety assessment which is currently based on Codex Alimentarius guidelines published in 2003-2009. 

Objective: Clarify and advance protein safety assessment making use of experience gained and new scientific advances achieved over the last 20 years. Through the review of trends and the exploration of new strategies, it aims to develop new approaches for protein safety assessments. 

Methodology: The main activities are outsourcing and a workshop to communicate findings and gather feedback. This involves developing new approaches for allergenicity assessment and in silico protein toxicity prediction software. The project is engaging with Codex Alimentarius and aims to revise current risk assessment strategies for open reading frames analysis.

Project results: The project delivered a comprehensive protein safety assessment framework, including novel approaches for allergenicity and toxicological assessment, effects of processing and adverse outcome pathways for celiac disease. These outputs are now integrated into EFSA's risk assessment processes, enhancing scientific rigour and supporting reduced animal testing.

Metagenomics approaches are techniques that enable the analysis of microbial communities, providing a more comprehensive evaluation of complex microbial products used in food and feed.

Objectives: Develop fit-for-purpose metagenomics methodologies for assessing complex microbial mixtures in regulated products and provide recommendations for integrating metagenomics data into EFSA risk assessment workflows.

Methodology: development and validation of standardised protocols for metagenomics data generation and analysis, including laboratory procedures, bioinformatics pipelines for microbial community characterisation and detection of safety-relevant genomic features such as antimicrobial resistance genes and virulence factors. The project will also develop quality control standards, acceptance criteria, for risk assessment.

Emerging evidence demonstrates that dietary xenobiotics can affect the gut microbiome and host health through multiple pathways. Traditional risk assessment approaches do not adequately address these microbiome-mediated effects. EFSA is advancing its risk assessment methodologies to incorporate this scientific understanding and provide a more comprehensive evaluation of dietary safety. 

Objectives: Establish methodological foundations to develop a risk assessment framework for evaluating adverse effects of dietary xenobiotics on the gut microbiome and host health. The project aims to systematically evaluate and prioritise microbiome-relevant biomarkers, assess the suitability of existing in vitro models, identify robust biomarker–model pairings, and provide recommendations for the feasibility of future experimental validation.

Methodology: The project will employ a systematic appraisal and evidence synthesis approach to identify and prioritise microbiome-relevant biomarkers and in vitro models related to key health endpoints. It will also develop quality control standards and data requirements for regulatory science, assess complementary assessment methods for validation, and evaluate the technical and resource requirements for potential future experimental testing.

Allergenicity is a cross-cutting issue in the risk assessment (RA) of proteins, particularly for regulated products. However, current strategies for allergenicity RA are based on 2003 Codex Alimentarius guidelines, which do not necessarily reflect the latest scientific advances.

Objectives: Develop a novel strategy for fit-for-purpose allergenicity RA of proteins in food and feed, building on recent scientific advances, gained in-house experience and ongoing dialogue with key stakeholders. This novel strategy will enhance the reliability and speed of allergenicity RA of proteins in food and feed and support process simplification and harmonisation, ultimately enabling better- informed risk management decision-making.

Methodology: For both clinically-relevant cross-reactivity and de novo sensitisation, the project involves the development of open source in silico prediction/appraisal tools to be validated with experimental in vitro and in vivo data; assessment of future prospects for in silico tools; estimation of thresholds of allergological concern; and the development of a systematic tiered approach supported by a decision-making framework with criteria triggering each of the tiers. 

Human biomonitoring (HBM) data measures chemical exposure in human populations and provides valuable information for chemical risk assessment. Recent European regulatory developments require a comprehensive strategy for HBM data integration in chemical risk assessments.

Objectives: Develop a comprehensive strategy for enhancing the reliability of EFSA’s chemicals risk assessment through the integration of HBM data.

Methodology: Assess the availability of existing HBM data, define assessment questions relevant to EFSA for chemicals and identify methodological challenges. Subsequently, two pilot case studies will verify the applicability of HBM data in risk assessment and final recommendations will be provided.

Chemical risk assessment typically evaluates exposure from individual sources separately. However, simultaneous exposure from multiple sources and routes is a reality. A unified approach to assess aggregate exposure across different regulatory frameworks and bodies is needed to improve the consistency and efficiency of chemical risk assessments in the EU.

Objectives: Develop a unified EU methodological framework for assessing aggregate exposure to chemicals from multiple sources and routes of exposure. The framework, once developed, will support different regulatory contexts.

Methodology: Existing tools, methods, and data from other relevant initiatives are integrated to develop a draft framework, to be refined through public consultation and pilot case studies.