March 2023: PhD defence: Paving the road towards the Next Generation Risk Assessment - Quantitative in vitro to in vivo extrapolations for the pre-diction of human toxicity based on alternative testing
The research for this PhD thesis was performed by Stella Fragki of the RIVM and Utrecht University, and was funded by the European human biomonitoring project HBM4EU. Her thesis describes the extrapolation of concentration-response relations observed in in vitro assays to dose-response relations in human. For example, she showed that including data from a cell culture system mimicking the placental barrier improves extrapolation significantly. This approach called quantitative in vitro to in vivo extrapolation (QIVIVE) is an important element in innovative hazard and risk assessment of chemicals and pharmaceuticals.
February 2023: 3Rs Working Party (3RsWP) plenary meeting - Public session on the 2023 work plan
Join the public session of the 3RsWP of the European Medicine Agency (EMA) on 28 February, 09:00–09:45. In this session the 3RsWP will present their work plan and priorities for 2023 and give an opportunity to stakeholders to comment and provide their views on the working party's activities. This public session is part of this newly established working party's two-day plenary meeting to showcase EMA's commitment to ensuring the implementation of 3Rs principles in the development and evaluation of human and veterinary medicinal products
3Rs Working Party (3RsWP) plenary meeting
February 2023: Innovating human chemical hazard and risk assessment through a holistic approach
Regulatory toxicology has relied on animal studies as the gold standard for over half a century. It has increasingly been realised that the detour of the animal study for human risk assessment comes with drawbacks, both scientifically as well as ethically. In this publication, RIVM researchers pose that innovative New Approach Methodologies provide molecular and mechanistic data that can be integrated in computational models to predict adverse outcome at the level of the intact human. It is expected that in due course, current significant advances in computational toxicology and machine learning already effective in clinical practice, will be able to advance into a human-focused, animal-free risk assessment strategy. This strategy will be fine-tuned to the human situation and will therefore lack the scientific and ethical concerns of current classical regulatory animal testing for human chemical risk assessment.
Current Opinion in Toxicology 2023; available online 10 February 2023, 100386
February 2023: Prolonged differentiation of neuron-astrocyte co-cultures results in emergence of dopaminergic neurons
Epidemiological and experimental data suggest that pesticides affect dopaminergic cells in the brain. An effect that has been related to human neurodegenerative diseases such as Parkinson’s disease. To allow for human-relevant assessment of (developmental) neurotoxicity related to dopamine disruption, researchers from RIVM have developed a human stem cell-based differentiation protocol for dopaminergic neurons. Further characterisation and chemical testing are needed to define the role of the current model in a testing strategy to test how pesticides or other chemicals might affect the dopaminergic system.
Int J Mol Sci 2023; 24(4): 3608
February 2023: The State of the Art and Challenges of In Vitro Methods for Human Hazard Assessment of Nanomaterials in the Context of Safe-by-Design
The Safe-by-Design (SbD) concept aims to facilitate the development of safer materials/products throughout their life cycle. Early hazard screening is therefore essential. Partners of the SAbyNA consortium reviewed commonly used in vitro tests for their suitability for SbD hazard testing of nanomaterials. Some simple in vitro assays are capable of accurately predicting hazard warnings and hazard rankings for some but not all toxicological endpoints. The authors recommend that joint efforts of innovators, scientists, and regulators are needed to improve hazard testing for SbD approaches.
Nanomaterials 2023; 13(3): 472
January 2023: Smoking-associated exposure of human primary bronchial epithelial cells to aldehydes: impact on molecular mechanisms controlling mitochondrial content and function
Respiratory toxic aldehydes are generated during smoking of tobacco. To study the role of aldehydes on the molecular regulation of mitochondrial content/function, Maastricht University and RIVM used a differentiated in vitro model of human primary bronchial epithelial cells exposed to cigarette smoke following a realistic exposure protocol. Cigarette smoke caused disruption of molecular mechanisms controlling mitochondrial content and quality. However, exposure to a mixture of three aldehydes only caused a dysregulation of markers associated with autophagy and mitophagy. This suggests that other compounds then aldehydes contribute to cigarette smoke-induced dysregulation of mitochondrial content/function.
Cells 2022; 11(21): 3481
January 2023: Re-evaluating the need for chronic toxicity studies with therapeutic monoclonal antibodies, using a weight of evidence approach
For the registration of monoclonal antibodies (mAbs) for chronic indications, 6-month as well as 3-month toxicity studies in rodents are conducted. However, experience has shown that mAbs are generally well tolerated, thereby questioning the relevance to assess human safety in long term rodent toxicity studies. Researchers of the CBG-MEB studied whether a 3-month study is already sufficient to assess the long-term safety of mAbs. For the majority of the 111 assessed mAbs novel toxicity is rarely detected in animal studies of longer than 3 months. This may allow for a case-by-case approach to reduce animal use by conducting a 3- instead of 6-month study.
Regul Toxicol Pharmacol 2023; 138:105329
January 2023: The use of recovery animals in nonclinical safety assessment studies with monoclonal antibodies: further 3Rs opportunities remain
When performing nonclinical safety testing of a monoclonal antibody (mAb) in animals, it is sometimes required to include a recovery group. According to the guidelines, this is only needed in one of the nonclinical studies. A working group, including participants from CBG-MEB, reviewed 51 mAb toxicity studies and found that often recovery groups are included in multiple nonclinical studies over the course of testing a mAb. The working group concluded that by limiting inclusion of a recovery group to a single nonclinical toxicology study provides opportunities to decrease animal use for this drug class.
Regul Toxicol Pharmacol 2023; 138:105339
December 2022: Towards achieving a modern science-based paradigm for agrochemical carcinogenicity assessment
The rodent cancer bioassay has been the standard approach to test chemicals for human carcinogenic potential. However, there are known limitations to this assay, leading to a search for more human-relevant approaches providing mechanistic information on chemical carcinogenesis. In this paper the authors outlined the challenges and opportunities that authorities should consider when building a roadmap that should lead to global acceptance and incorporation of fit-for-purpose new approaches for carcinogenicity assessment of agrochemicals. New approaches include shorter-term tests, including in vitro and in silico approaches. To the publication.
December 2022: Application of AOPs to assist regulatory assessment of chemical risks – Case studies, needs and recommendations
The Adverse Outcome Pathway (AOP) framework is an important tool in moving towards next generation risk assessment (NGRA). A workshop organised within the European Union partnerships HBM4EU and PARC was held to address issues related to using AOPs in a regulatory context. AOPs have shown useful in e.g. hazard characterisation and in identification and validation of biomarkers of effect. However, e.g. lack of quantitative information on response-response relationships holds AOP application back. The paper discusses suggestions, initiatives and tools that may help to better implement AOPs in the NGRA. To the publication.
December 2022: Neurodegeneration in a Regulatory Context: The Need for Speed
There is increasing evidence that pesticides can induce or accelerate neurodegenerative diseases. Current regulatory guidelines for safety assessment cannot fully capture neurodegenerative effects of chemical substances. In this publication, researchers from RIVM propose a tiered approach to update the regulatory guidelines. In the short term, adaptations to current guideline studies can be made and in the long term transition of chemical safety assessment to a strategy based on human biology using in vitro and in silico methods should be adopted. To the publication.
December 2022: Applicability of generic PBK modelling in chemical hazard assessment: A case study with IndusChemFate
Various physiologically based kinetic (PBK) models exist that may all vary in their chemical applicability domain. In this study, the generic physiologically based kinetic model IndusChemFate was evaluated and compared to the PBK “TNO Model”. Both models were evaluated with a broad span of chemicals varying in physicochemical properties. Results showed that the simpler IndusChemFate model performed better as a first tier for simulating toxicokinetics based on QSARs and in vitro parameters. Findings were illustrated in HepaRG cells exposed to acetaminophen. To the publication.
December 2022: Dynamic regulation of gene expression and morphogenesis in the zebrafish embryo test after exposure to all-trans retinoic acid
This publication evaluated the Zebrafish Embryo Test (ZET) for the prediction of developmental toxicity. The effect of exposure duration of all-trans retinoic acid on the magnitude of gene expression changes was investigated. Results showed that gene expression regulation indicative of developmental toxicity could be observed already at early time points after exposure. Considering early time point measurements of gene expression changes could help optimise the sensitivity of the ZET in future studies. To the publication.
18 November 2022: Hugo van Poelgeest Award won by Niels Harlaar
The Hugo van Poelgeest Award is a scientific prize for excellent, animal-free research in the life sciences. Niels Harlaar (LUMC) won the award for his research on transformed human heart cells. Valentina Palacio-Castañeda (Radboudumc) won the audience award, and Victoria de Leeuw (RIVM) was nominated. The prize is awarded every other year by stichting Proefdiervrij and stichting Bouwstenen voor Dierenwelzijn. Read more (in Dutch).
November 2022: The Current Status and Work of Three Rs Centres and Platforms in Europe
Over recent years, an increasing number of 3R centres and platforms have been established in Europe. Together with representatives of European 3R initiatives, RIVM contributed to a publication that provides an overview of the current activities of the 3R centres: the financial and organisational structures of the, what their Three Rs focus and core activities are (dissemination, education, implementation, scientific quality/translatability, ethics), and their areas of responsibility and projects. See the publication.
22 October 2022: Analyses of Transcriptomics Cell Signalling for Pre-Screening Applications in the Integrated Approach for Testing and Assessment of Non-Genotoxic Carcinogens
Mechanistic understanding of biological processes that lead to carcinogenesis is expanding. Transcriptomics can inform on non-genotoxic mechanisms of cancer caused by chemicals. An OECD working group, in which RIVM participates, reviewed the application of transcriptomics for regulatory applications. Specifically, they evaluated the potential of gene expression analyses to identify the most relevant approaches that could be utilised as (pre-)screening tools. Challenges related to the integration of gene panels and transcriptomic assays into an Integrated Approach to the Testing and Assessment of non-genotoxic carcinogens is also discussed. To the publication.
11 October 2022: Renewal of the EURL ECVAM Scientific Advisory Committee (ESAC) and open call for application subgroups
The Joint Research Centre (JRC) formally appointed nine experts as the new member of the ESAC on 11 October 2022. Anne Kienhuis, senior scientist of the RIVM is one of the new members. ESAC provides EURL ECVAM with independent scientific advice on the adequacy and outcome of formal validation studies carried out to assess the reliability and relevance of NAMs in the context of regulatory safety assessment. Read more in this news item.
New videos on TPI.tv
Working towards animal-free research strategies is an effort crossing international and interdisciplinary borders. TPI.tv is designed to facilitate this transition through connecting people and ideas via video messages. On TPI.tv, short video messages (‘pitches’) provide the medium for researchers to display their findings and request input if needed. New videos on TPI.tv:
In3 | Helpathon #6 | Helpathon #7 | In vitro atrial fibrillation | In vitro neurodevelopmental toxicity assessment | Tumour-on-chip
October 2022: 3Rs concepts to improve the quality of biomedical science COST-Action IMPROVE
Improving the quality of biomedical science by using 3Rs concepts is the main aim of the new COST-Action IMPROVE. In this initiative, a network of researchers will be established to refine, harmonise and promote 3Rs concepts, data and documents, in order to improve the quality of biomedical science. Researchers interested in contributing can apply for working groups on the (1) Quality and Translatability of Science, (2) Implementation, (3) Dissemination, and (4) Education. The consortium is co-led by Utrecht University and RIVM and is funded by the Dutch National Research Agenda. Read more about IMPROVE.
September 2022: Prioritisation of chemicals in food for risk assessment by integrating exposure estimates and new approach methodologies: A next generation risk assessment case study
Next generation risk assessment (NGRA) is a knowledge-driven system that allows for cost-efficient, animal-free chemical risk assessment. In this case study, researchers at RIVM showed how NGRA can facilitate prioritisation of chemical substances in food. They explored how exposure data combined with the Threshold of Toxicological Concern concept could be used to prioritise chemicals for toxicity testing. A selected set of compounds from this analysis was screened for toxicological potential using high-throughput screening. This study is a proof-of-principle that this approach can be a supportive element in NGRA. To the publication.
September 2022: Development of a cell line-based in vitro assay for assessment of Diphtheria, Tetanus and acellular Pertussis (DTaP)-induced inflammasome activation
Safety and potency assessment for batch-release testing of established vaccines still partly relies on animal tests. As part of the VAC2VAC project, researchers at RIVM developed an in vitro testing method for adjuvant biological activity that can contribute to animal-free potency testing of vaccines. Specifically, they developed an NLRP3 inflammasome activation test for aluminium-adjuvanted vaccines (the DTaP vaccine). The results indicated that inflammasome activation may be used to measure adjuvant biological activity as an important quality attribute for control or characterisation of the adjuvant. More information in the publication Development of a cell line-based in vitro assay for assessment of Diphtheria, Tetanus and acellular Pertussis (DTaP)-induced inflammasome activation.
September 2022: Realistic in vitro exposure of neuronal cells to e-cigarette vapour
August 2022: Biomarkers for quality control of Weil’s disease vaccines
Potency testing of Leptospira (Weil's disease) vaccine batches relies partially on laboratory animals. Cell-based assays could replace in vivo tests for vaccine quality control if biomarkers indicative of Leptospira vaccine potency are identified. RIVM contributed to a study that used a canine monocyte/macrophage cell line to investigate innate immune responsiveness by Leptospira species as well as Leptospira vaccines from two different manufacturers. Gene and protein expression analysis led to the identification of four proteins for the development of biomarker-based assays. Read more Transcriptome and proteome analysis of innate immune responses to inactivated Leptospira and bivalent Leptospira vaccines in canine 030-D cells.
August 2022: Animal-free assays for endocrine disruption – from science to regulatory acceptance
The zebrafish embryo is a promising alternative non-rodent model in toxicology. Scientists at RIVM and Leiden University exposed zebrafish embryos to nine hepatotoxicants (targeted at induction of steatosis, cholestasis and necrosis). Next, they determined protein expression changes using LC-MS/MS and identified differentially expressed proteins and pathways. Overlap between these responses resulted in a set of ten common protein markers for the detection of adverse liver responses. This may allow for a reduction in rodent use for hepatotoxicity assessment. Read more Identification of proteome markers for drug-induced liver injury in zebrafish embryos.
June 2022: Roadmap for Action on New Approach Methodologies in Risk Assessment
The European Food Safety Authority set out a call to develop a roadmap for the implementation of New Approach Methodologies (NAMs) in food risk assessment. A team of experts, from Fraunhofer ITEM, RIVM, Vrije Universiteit Amsterdam, Technical University of Denmark, and EURA AG defined key barriers and opportunities as well as activity proposals for implementation of NAMs in food risk assessment in seven areas: development of additional adverse outcome pathways (AOPs)/AOP networks, advanced cell culture models including organ-on-chip, toxicokinetic assessment with a focus on physiological based kinetic modelling, use of exposome data, integration of human susceptibility, data integration and new concepts in human risk assessment for NAM implementation. Furthermore, a Forum is proposed to facilitate interaction between stakeholders for the implementation of NAMs in risk assessment. Read more.
May 2022: Update Landscape NAMs
We have published an update of the Landscape NAMs for the safety assessment of chemical substances. The overview provides information on each step in the implementation curve. With this Landscape, we facilitate the uptake of NAMs in regulatory safety testing. Go to the Landscape NAMs.