With the growing numbers of nanomaterials (NMs), there is a great demand for rapid and reliable ways of testingNMsafety—preferably usingin vitroapproaches, to avoid the ethical dilemmas associated with animal research. Data are needed for developing intelligent testing strategies for risk assessment ofNMs, based on grouping and read‐across approaches. The adoption of high throughput screening (HTS) and high content analysis (HCA) forNMtoxicity testing allows the testing of numerous materials at different concentrations and on different types of cells, reduces the effect of inter‐experimental variation, and makes substantial savings in time and cost.HTS/HCAapproaches facilitate the classification of key biological indicators ofNM‐cell interactions. Validation ofin vitroHTStests is required, taking account of relevance toin vivoresults.HTS/HCAapproaches are needed to assess dose‐ and time‐dependent toxicity, allowing prediction ofin vivoadverse effects. SeveralHTS/HCAmethods are being validated and applied forNMtesting in theFP7projectNANoREG, including Label‐free cellular screening ofNMuptake, HCA, High throughput flow cytometry, Impedance‐based monitoring, Multiplex analysis of secreted products, and genotoxicity methods—namely High throughput comet assay, High throughputin vitromicronucleus assay, andγH2AXassay. There are several technical challenges withHTS/HCAforNMtesting, as toxicity screening needs to be coupled with characterization ofNMsin exposure medium prior to the test; possible interference ofNMswithHTS/HCAtechniques is another concern. Advantages and challenges ofHTS/HCAapproaches inNMsafety are discussed.WIREs Nanomed Nanobiotechnol2017, 9:e1413. doi: 10.1002/wnan.1413This article is categorized under: Therapeutic Approaches and Drug Discovery > Emerging Technologies Toxicology and Regulatory Issues in Nanomedicine > Regulatory and Policy Issues in Nanomedicine Toxicology and Regulatory Issues in Nanomedicine > Toxicology of Nanomaterials

In the current paper, a new strategy for risk assessment of nanomaterials is described, which builds upon previous project outcomes and is developed within the FP7 NANoREG project. NANoREG has the aim to develop, for the long term, new testing strategies adapted to a high number of nanomaterials where many factors can affect their environmental and health impact. In the proposed risk assessment strategy, approaches for (Quantitative) Structure Activity Relationships ((Q)SARs), grouping and read-across are integrated and expanded to guide the user how to prioritise those nanomaterial applications that may lead to high risks for human health. Furthermore, those aspects of exposure, kinetics and hazard assessment that are most likely to be influenced by the nanospecific properties of the material under assessment are identified. These aspects are summarised in six elements, which play a key role in the strategy: exposure potential, dissolution, nanomaterial transformation, accumulation, genotoxicity and immunotoxicity. With the current approach it is possible to identify those situations where the use of nanospecific grouping, read-across and (Q)SAR tools is likely to become feasible in the future, and to point towards the generation of the type of data that is needed for scientific justification, which may lead to regulatory acceptance of nanospecific applications of these tools.

The blood brain barrier (BBB) is the bottleneck of brain-targeted drug development. Due to their physico-chemical properties, nanoparticles (NP) can cross the BBB and accumulate in different areas of the central nervous system (CNS), thus are potential tools to carry drugs and treat brain disorders. In vitro systems and animal models have demonstrated that some NP types promote neurotoxic effects such as neuroinflammation and neurodegeneration in the CNS. Thus, risk assessment of the NP is required, but current 2D cell cultures fail to mimic complex in vivo cellular interactions, while animal models do not necessarily reflect human effects due to physiological and species differences. We evaluated the suitability of in vitro models that mimic the human CNS physiology, studying the effects of metallic gold NP (AuNP) functionalized with sodium citrate (Au-SC), or polyethylene glycol (Au-PEG), and polymeric polylactic acid NP (PLA-NP). Two different 3D neural models were used (i) human dopaminergic neurons differentiated from the LUHMES cell line (3D LUHMES) and (ii) human iPSC-derived brain spheroids (BrainSpheres). We evaluated NP uptake, mitochondrial membrane potential, viability, morphology, secretion of cytokines, chemokines and growth factors, and expression of genes related to ROS regulation after 24 and 72 h exposures. NP were efficiently taken up by spheroids, especially when PEGylated and in presence of glia. AuNP, especially PEGylated AuNP, effected mitochondria and anti-oxidative defense. PLA-NP were slightly cytotoxic to 3D LUHMES with no effects to BrainSpheres. 3D brain models, both monocellular and multicellular are useful in studying NP neurotoxicity and can help identify how specific cell types of CNS are affected by NP.