Update on the development of a regulatory framework for developmental neurotoxicity testing with specific focus on non-animal testing approaches

Reports of developmental neurotoxicity (DNT) incidences have become increasing over the last 2-3 decades, triggering the need for higher focus on this endpoint and an efficient, tiered testing approach to allow identifying developmental neurotoxicants. Unfortunately, the current progress of non-animal alternative methods for testing DNT did not reach yet the stage of regulatory acceptance.

Over the last 50 years there has been significant development in our understanding of the developmental neurotoxicity hazard. Published research during 1960-1980s helped to identify and characterise the hazard of in utero exposure of chemicals to the developing young animals. Subsequent research work helped develop the in vivo developmental neurotoxicity (DNT) testing guidelines by US EPA and OECD. Subsequently, a number of workshops and inter-laboratory studies have helped to identify the validity, reliability and sensitivity of the in vivo tests including other aspects such as human relevance, statistical analyses, positive controls, or sensitivity of behavioural measures.

So far, systematic testing for DNT is not a mandatory regulatory requirement in Europe for chemical safety assessment purposes. The testing is performed only as higher tiered tests if evidence of neurotoxicity in standard, systemic, adult developmental or reproductive toxicity studies. However, with the growing number of incidences for neurodevelopmental disorders in children (e.g., autism, dyslexia, obsessive compulsive disorder to name a few), increasing number of potential chemicals (e.g., lead, polychlorinated biphenyls) and inherent limitations of the in vivo tests (e.g., use of high number of animals, time or cost) as well as animal welfare considerations, the development of high-throughput, efficient and validated in vitro methods have become essential.

Since early 2000, a series of workshops have been held specifically to promote the development and use of in vitro DNT studies. Ready-to-go assays have been developed for a number of neurodevelopmental processes such as differentiation into neural precursor cells (NPC), NPC apoptosis, astrocyte and oligodendrocyte differentiation, myelin formation, synaptogenesis, neural network formation etc. Additionally, over the last few years, multiple reviews have been undertaken to generate lists of reference chemicals. Open databases have been created to share and compare methods and results.

Lately, few structured literature reviews and workshops have been conducted to understand the state-of-the-art knowledge base on the alternative models for DNT and progress towards regulatory decision-making framework. An EFSA-contracted literature review in 2015 indicated the need for standardisation across laboratories and in vitro-in vivo validation. More recently, the workshop co-organised by OECD and EFSA on “Developmental Neurotoxicity (DNT): the use of non-animal test methods for regulatory purposes” in October 2016 in Brussels, Belgium, focused on opportunities and challenges related to alternative methods for testing and assessment. Among others, generation of data for more chemicals, need for data sharing in order to compare work across laboratories and across assays, co-ordination between research and regulators have been identified as the key factors in establishing tiered testing approaches and decision-making frameworks for this endpoint.

In a recent consensus statement from representatives of industry, academia and regulatory authorities, the need for non-animal, reliable and human-focused mechanistic hazard assessment for DNT has been expressed. We concur that development of such a framework is critical which should only be possible through development of new OECD guidance considering the in vitro and in silico methods, generation of data for more number of chemicals to enrich the framework, implementation of the appropriate alternative methods already available and through continuous research to complement the existing knowledge.

Suggested reading:

Crofton KM, Mundy WR, Shafer TJ, 2012. Developmental neurotoxicity testing: A path forward. Yuji Nakajima (ed.), Congenital Anomalies. Wiley-Blackwell, Hoboken, NJ, 52(3):140-146.

Fritsche E, Alm H, Baumann J, Geerts L, Håkansson H, Masjosthusmann S, Witters H, 2015. Literature review on in vitro and alternative developmental neurotoxicity (DNT) testing methods. EFSA Supporting Publication; 12(4):EN‐778, 186 pp.

Joint Research Centre, 2016. EURL ECVAM status report on the development, validation and regulatory acceptance of alternative methods and approaches. JRC103522. EUR 28156.

Fritsche E, Crofton K, Hernandez A, Hougaard Bennekou S, Leist M, Bal-Price A, Reaves E, Wilks M, Terron A, Sachana M, Gourmelon A, 2017. OECD/EFSA workshop on developmental neurotoxicity (DNT): The use of non-animal test methods for regulatory purposes. ALTEX – Alternatives to animal experimentation. 34(2):  311-315.

Fritsche E et al., 2018. Consensus statement on the need for innovation, transition and implementation of developmental neurotoxicity (DNT) testing for regulatory purposes. Toxicol Appl Pharmacol. 354:3-6.