Scoping review
Human-Based New Approach Methodologies for Assessment of Developmental Neurotoxicity: A Scoping Review on Regulatory Applicability, Limitations, and Opportunities
Rationale
Human neurodevelopment is a complex process characterized by extended developmental and maturational timelines, dynamic cellular processes, greater neural network complexity, and species-specific molecular functions that may differ substantially from those observed in small mammals. However, regulatory frameworks for neurotoxicity assessment continue to rely on rodent-based models to meet safety requirements for chemicals and pharmaceuticals, for both developmental neurotoxicity (DNT) and adult neurotoxicity in humans (Koch et al., 2022; Kuchovska et al., 2026; OECD, 2023; Popp & Ramos, 2026; Tal et al., 2024). The reliance on such resource-intensive models explains the low throughput of chemical neurotoxicity assessment and an associated key limitation: the small number of chemicals evaluated to date under DNT in vivo guidelines. Based on REACH registration estimates, only approximately 150 of the roughly 30,000 chemicals in the human exposome have so far been evaluated in regulatory DNT (Kuchovska et al., 2026).
Given the high costs, ethical concerns, and translational limitations associated with in vivo models, their use in developmental neurotoxicity testing has increasingly been questioned. To address the need for more affordable, high throughput, and human-relevant methods for DNT assessment, new approach methodologies (NAMs) are in increasing demand, aligning with the European Commission’s sustainability goals, the Green Deal, and its efforts to prioritize reducing animal testing in regulatory toxicology (Weber et al., 2025).
Human-based NAMs, such as human primary and induced pluripotent stem cell (hiPSC)-based models, 3D neurospheres/spheroids, organoids, and microphysiological systems/organ-on-a-chip based on human cells, are promising alternatives for non-animal DNT assessment. Some are already integrated into an in vitro battery (IVB) of assays currently under discussion at the regulatory level (OECD, 2023), which has demonstrated robust technical performance and accuracy for screening and prioritization (Blum et al., 2023). However, it does not fully capture the biological complexity of the developing nervous system, including incomplete coverage of glial biology, immune and neuroinflammatory responses, myelination, and physiological barriers such as the blood-brain and placental barriers (Sachana et al., 2026).
Despite the growing momentum and wealth of literature on non-animal methods for DNT assessment, there remains a need for a comprehensive scoping review mapping the current landscape of human-based NAM approaches for DNT, including their applications, documented limitations, and opportunities for evidence-based decision-making and wider regulatory use.
Objectives
The primary objective of this project is to conduct a scoping review on the current landscape of human-based NAMs for the assessment of developmental neurotoxicity (DNT). Specifically, the review aims to identify the DNT-relevant neurodevelopmental endpoints and modes of action currently covered by existing human-based NAMs and those that remain unaddressed, characterize the proposed regulatory use of the identified NAMs, and generate evidence to inform prioritization of future NAM development.
Methodology
This scoping review will be conducted and reported in accordance with the PRISMA-ScR (Preferred Reporting Items for Systematic Reviews and Meta-Analyses extension for Scoping Reviews) checklist (Tricco et al., 2018). A protocol has been developed a priori, in line with Peters et al. (2020), and will be made publicly available on a dedicated platform ahead of the review’s execution. An adapted PCC (Population, Concept, Context) framework guided the formulation of the research question and eligibility criteria, focusing on human cell-based in vitro models for DNT assessment within a regulatory context. Any amendments to the protocol will be clearly documented, justified, and reported in the resulting publication.
A preliminary literature search confirmed that a substantial and feasible body of literature exists to support the proposed review. The formal search strategy will draw on multiple biomedical databases, with records screened independently by two investigators against predefined eligibility criteria. Given that regulatory use is often reported in non-peer-reviewed regulatory documents, the review will also include a supplementary scan of key regulatory body publications (OECD, EFSA, ECHA, EURL ECVAM) to contextualize the peer-reviewed findings.
Status and outcomes
This project is ongoing. If successful, it will produce a structured scoping review characterizing the DNT-relevant endpoints covered by existing human-based NAMs, their validation and regulatory acceptance status, and documented gaps and limitations (e.g., immune-mediated modes of action, metabolic competence, glial biology, blood-brain barrier modeling). The work is intended to inform NAM developers on opportunities for further development and to provide the scientific community, including regulators, with greater transparency regarding the current capabilities of human-based NAMs. Findings will be submitted to a peer reviewed journal and shared through this platform and our LinkedIn page.
Open to collaborations: Yes. If you are interested in contributing to this project, get in touch.
References
Blum, J., Masjosthusmann, S., Bartmann, K., Bendt, F., Dolde, X., Dönmez, A., Förster, N., Holzer, A. K., Hübenthal, U., Keßel, H. E., Kilic, S., Klose, J., Pahl, M., Stürzl, L. C., Mangas, I., Terron, A., Crofton, K. M., Scholze, M., Mosig, A., … Fritsche, E. (2023). Establishment of a human cell-based in vitro battery to assess developmental neurotoxicity hazard of chemicals. Chemosphere, 311. https://doi.org/10.1016/j.chemosphere.2022.137035
Koch, K., Bartmann, K., Hartmann, J., Kapr, J., Klose, J., Kuchovská, E., Pahl, M., Schlüppmann, K., Zühr, E., & Fritsche, E. (2022). Scientific Validation of Human Neurosphere Assays for Developmental Neurotoxicity Evaluation. Frontiers in Toxicology, 4. https://doi.org/10.3389/ftox.2022.816370
Kuchovska, E., Bartmann, K., Raad, G., Schade, M., Ladeira, L., Dönmez, A., Klose, J., Görts, N., Polozij, D., Saborowski, L. C., Bendt, F., Staumont, B., Geris, L., Koch, K., & Fritsche, E. (2026). Functional Mapping of Neurodevelopmental Disease Pathways to Key Neurodevelopmental Processes Represented in the Developmental Neurotoxicity In Vitro Testing Battery. Advanced Science. https://doi.org/10.1002/advs.202519889
OECD. (2023). Initial Recommendations on Evaluation of Data from the Developmental Neurotoxicity (DNT) In-Vitro Testing Battery Series on Testing and Assessment No. 377 JT03530742 OFDE.
Peters MDJ, Godfrey C, McInerney P, Munn Z, Tricco AC, Khalil, H. Scoping Reviews (2020). Aromataris E, Lockwood C, Porritt K, Pilla B, Jordan Z, editors. JBI Manual for Evidence Synthesis. JBI; 2024. Available from: https://synthesismanual.jbi.global. https://doi.org/10.46658/JBIMES-24-09
Popp, J. T., & Ramos, R. L. (2026). Best practices in neurotoxicology research must account for spontaneous brain malformations in rodent models. In NeuroToxicology (Vol. 114). Elsevier B.V. https://doi.org/10.1016/j.neuro.2026.103474
Sachana, M., Hogberg, H. T., & Mangas, I. (2026). From concept to regulation: OECD’s role in the development of the DNT in vitro battery. In Frontiers in Toxicology (Vol. 8). Frontiers Media SA. https://doi.org/10.3389/ftox.2026.1774469
Tal, T., Myhre, O., Fritsche, E., Rüegg, J., Craenen, K., Aiello-Holden, K., Agrillo, C., Babin, P. J., Escher, B. I., Dirven, H., Hellsten, K., Dolva, K., Hessel, E., Heusinkveld, H. J., Hadzhiev, Y., Hurem, S., Jagiello, K., Judzinska, B., Klüver, N., … Bartmann, K. (2024). New approach methods to assess developmental and adult neurotoxicity for regulatory use: a PARC work package 5 project. Frontiers in Toxicology, 6. https://doi.org/10.3389/ftox.2024.1359507
Tricco, A. C., Lillie, E., Zarin, W., O’Brien, K. K., Colquhoun, H., Levac, D., Moher, D., Peters, M. D. J., Horsley, T., Weeks, L., Hempel, S., Akl, E. A., Chang, C., McGowan, J., Stewart, L., Hartling, L., Aldcroft, A., Wilson, M. G., Garritty, C., Lewin, S., Godfrey, C. M., Macdonald, M. T., Langlois, E. V., Soares-Weiser, K., Moriarty, J., Clifford, T., Tunçalp, Ö., & Straus, S. E. (2018). PRISMA extension for scoping reviews (PRISMA-ScR): Checklist and explanation. Annals of Internal Medicine, 169(7), 467–473. https://doi.org/10.7326/M18-0850
Weber, P., Arnesdotter, E., Attar, N., Friedrichs, S., Seitz, C., Chowdhury, R. R., Koch, K., Buerki-Thurnherr, T., Brugger, B., Wick, P., Morikka, J., Serra, A., Capini, L., Greco, D., Lacconi, V., Marchese, A. S., Campagnolo, L., Exner, T. E., Brajnik, M., … Serchi, T. (2025). CHIASMA: Advancing chemicals and materials safety and sustainability assessments through innovative integration of in vitro and in silico (new approach) methodologies. Computational and Structural Biotechnology Journal, 29, 305–327. https://doi.org/10.1016/j.csbj.2025.11.032