OECD/EFSA workshop on developmental neurotoxicity (DNT): The use of non-animal test methods for regulatory purposes

Fritsche, Ellen; Crofton, Kevin M.; Hernández, Antonio F.; Bennekou, Susanne Hougaard; Leist, Marcel; Bal‐Price, Anna; Reaves, Elissa; Wilks, Martin F.; Terron, Andrea; Solecki, Roland; Sachana, Magdalini; Gourmelon, Anne

doi:10.14573/altex.1701171

Cited by 84 publications

(59 citation statements)

References 19 publications

(20 reference statements)

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“…Variability, data reporting and analysis, limitations and expectations of DNT have been summarized recently (Smirnova et al, 2014; Li et al, 2017). In vivo DNT testing seems to be insufficient to study and characterize potential hazardous compounds, thus, in vitro testing batteries are required to generate additional data on the effects of chemicals on brain development (Fritsche et al, 2017) and for better identification, prioritization, and evaluation of chemicals with the potential to induce DNT (Bal-Price et al, 2012). Diverse workshops and meetings during the past 10 years have summarized the problems, the available assays and roadmaps for DNT in vitro assays and methods (Coecke et al, 2007; Lein et al, 2007; Crofton et al, 2011; Crofton et al, 2014; Bal-Price et al, 2015a; Bal-Price et al, 2015b).…”

Section: Introductionmentioning

confidence: 99%

Rotenone exerts developmental neurotoxicity in a human brain spheroid model

Pamies

Block

Lau

et al. 2018

Toxicology and Applied Pharmacology

118

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Growing concern suggests that some chemicals exert (developmental) neurotoxicity (DNT and NT) and are linked to the increase in incidence of autism, attention deficit and hyperactivity disorders. The high cost of routine tests for DNT and NT assessment make it difficult to test the high numbers of existing chemicals. Thus, more cost effective neurodevelopmental models are needed. The use of induced pluripotent stem cells (iPSC) in combination with the emerging human 3D tissue culture platforms, present a novel tool to predict and study human toxicity. By combining these technologies, we generated multicellular brain spheroids (BrainSpheres) from human iPSC. The model has previously shown to be reproducible and recapitulates several neurodevelopmental features. Our results indicate, rotenone’s toxic potency varies depending on the differentiation status of the cells, showing higher reactive oxygen species (ROS) and higher mitochondrial dysfunction during early than later differentiation stages. Immunofluorescence morphology analysis after rotenone exposure indicated dopaminergic-neuron selective toxicity at non-cytotoxic concentrations (1 μM), while astrocytes and other neuronal cell types were affected at (general) cytotoxic concentrations (25 μM). Omics analysis showed changes in key pathways necessary for brain development, indicating rotenone as a developmental neurotoxicant and show a possible link between previously shown effects on neurite outgrowth and presently observed effects on Ca2+ reabsorption, synaptogenesis and PPAR pathway disruption. In conclusion, our BrainSpheres model has shown to be a reproducible and novel tool to study neurotoxicity and developmental neurotoxicity. Results presented here support the idea that rotenone can potentially be a developmental neurotoxicant.

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Section: Introductionmentioning

confidence: 99%

Rotenone exerts developmental neurotoxicity in a human brain spheroid model

Pamies

Block

Lau

et al. 2018

Toxicology and Applied Pharmacology

118

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“…According to the Toxicity testing in the 21 st century paradigm, there is a call for reliable in vitro methods that can provide rapid, high-throughput screening of chemicals [ 5 ]. However, due to the various uncertainties and problems mentioned for the current guidelines for DNT testing, there is also a great need of DNT screening specific alternative methods [ 6 ].…”

Section: Introductionmentioning

confidence: 99%

“…include a battery of relevant endpoints and should give mechanistic insight that would provide information to discriminate between different neural subpopulations and different stages of neural differentiation. International stakeholders have proposed a DNT testing strategy based on compound testing across a battery of in vitro tests including the important factors of timing and processes of brain development [ 6 ]. The use of mRNA biomarkers is a good example of such an approach that has previously been reported for DNT test systems [ 21 , 25 , 26 ].…”

Section: Introductionmentioning

confidence: 99%

Whole genome microarray analysis of neural progenitor C17.2 cells during differentiation and validation of 30 neural mRNA biomarkers for estimation of developmental neurotoxicity

et al. 2017

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Despite its high relevance, developmental neurotoxicity (DNT) is one of the least studied forms of toxicity. Current guidelines for DNT testing are based on in vivo testing and they require extensive resources. Transcriptomic approaches using relevant in vitro models have been suggested as a useful tool for identifying possible DNT-generating compounds. In this study, we performed whole genome microarray analysis on the murine progenitor cell line C17.2 following 5 and 10 days of differentiation. We identified 30 genes that are strongly associated with neural differentiation. The C17.2 cell line can be differentiated into a co-culture of both neurons and neuroglial cells, giving a more relevant picture of the brain than using neuronal cells alone. Among the most highly upregulated genes were genes involved in neurogenesis (CHRDL1), axonal guidance (BMP4), neuronal connectivity (PLXDC2), axonogenesis (RTN4R) and astrocyte differentiation (S100B). The 30 biomarkers were further validated by exposure to non-cytotoxic concentrations of two DNT-inducing compounds (valproic acid and methylmercury) and one neurotoxic chemical possessing a possible DNT activity (acrylamide). Twenty-eight of the 30 biomarkers were altered by at least one of the neurotoxic substances, proving the importance of these biomarkers during differentiation. These results suggest that gene expression profiling using a predefined set of biomarkers could be used as a sensitive tool for initial DNT screening of chemicals. Using a predefined set of mRNA biomarkers, instead of the whole genome, makes this model affordable and high-throughput. The use of such models could help speed up the initial screening of substances, possibly indicating alerts that need to be further studied in more sophisticated models.

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“…The question is whether this study should be requested for the fourth compound? In such a situation, a read-across hypothesis supported on AOP-based testing for the effect could possibly answer the question and thus an in vivo study would be obsolete Consequently, consensus was reached between regulators, academic scientists and industry about the need for an alternative DNT testing strategy that is based on a standardised in vitro battery of tests in order to test more chemicals and inform more targeted in vivo studies (Fritsche et al, 2017). This activity was followed up by the OECD and there are ongoing efforts to develop a guidance document and generate data and case studies using a test battery and other NAMs in an IATA context (Sachana et al, 2019).…”

Section: Open Doorsmentioning

confidence: 99%

Moving towards a holistic approach for human health risk assessment – Is the current approach fit for purpose?

Bennekou

2019

EFS2

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It is recognised that new scientific improvements and their integration in risk assessment, as outlined in the National Academies of Sciences, Engineering and Medicine 2017 report, have the potential to improve human health risk assessments by enabling a mechanistic understanding of adverse effects and more accurate predictions of biological responses. Here, I discuss why such improvements are needed and can ease a paradigm shift in human health risk assessment. The current approach to human health risk assessment is limited by several elements: (1) the relevance of data is debatable, as they are largely based on in vivo animal models that are poorly predictive for complex endpoints, raise challenges with regard to interspecies extrapolations, and are seldom informative of the mechanism underlying the observed effects; (2) lack of flexibility in data requirements by regulators, which limits the uptake of new scientific developments in a timely manner; and (3) lack of data accessibility, which makes data integration difficult. However, mechanistic-based assessments are currently conducted for the identification of endocrine disruptors and are developed for addressing developmental neurotoxicity. Such assessments can serve as examples for changing the paradigm of risk assessment. There are several opportunities for improvement, such as: make regulatory standard requirements less prescriptive; enhance and use the opportunities for read-across; analyse and quantify uncertainties in order to benchmark new approach methods to the current system; better integrate screening methods early in regulatory assessments and decision-making; and develop more adverse outcome pathways in order to link new approach methods with the current approach and ultimately make it possible to base regulatory decisions on early key events of a toxicity pathway.

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OECD/EFSA workshop on developmental neurotoxicity (DNT): The use of non-animal test methods for regulatory purposes

Cited by 84 publications

References 19 publications

Rotenone exerts developmental neurotoxicity in a human brain spheroid model

Rotenone exerts developmental neurotoxicity in a human brain spheroid model

Whole genome microarray analysis of neural progenitor C17.2 cells during differentiation and validation of 30 neural mRNA biomarkers for estimation of developmental neurotoxicity

Moving towards a holistic approach for human health risk assessment – Is the current approach fit for purpose?

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