Alternative in vitro assays in nanomaterial toxicology

Abstract

Nanomaterials are acclaimed for their novel properties, for which broad new uses are being discovered with increasing frequency. It is obvious that, as the properties change, unwanted properties (toxicity) are to be expected as well. Current toxicology, however, is already overwhelmed with the challenge of addressing new chemicals, not to mention the enormous number of old chemicals never properly assessed. Limitations of traditional approaches range from animal welfare issues, which were a strong driving force for alternative approaches (the 3Rs concept) over the last two decades, to aspects of throughput and accuracy of the predicted toxicities. The latter has prompted discussion about a revolutionary change in chemical safety assessment, now known as Toxicology for the 21st Century (Tox-21c). The multitude of possible formulations of nanomaterials to be assessed for novel toxic properties makes these alternative approaches especially attractive, given the well recognized limitations of traditional animal-based approaches--limitations that might be even more pronounced for nanomaterials, which have notably altered biokinetics.

Assessing nanoparticle toxicity in cell-based assays: influence of cell culture parameters and optimized models for bridging the in vitro–in vivo gap

“…Most studies, certainly preliminary studies, are conducted on rodents as costs are lower, animals easier to access and infrastructural requirements less elaborate 73 . Official instancies have determined the species and number of animals and that should be used for a specific tests, to meet the incentive to reduce the number of animals used as urged by the 3R concept by Russel and Burch, ethical criticism and the pressure to develop a more cost-effective toxicity assessment protocol 93,94 . The European Commission has bundled it's guidelines in the REACH (Registration, Evaluation and Administration of Chemicals) regulation.…”

“…The evaluation of NP toxicity using these common assays has however resulted in conflicting data as can be found in reviews listing data on the toxicity of a specific NP or the correlation between the NP's physicochemical properties and the toxic effects it evokes 29,[101][102][103] . This led to an increasing awareness that these methods, and especially the in vitro methods, are not as appropriate and well-suited for nanotoxicology purposes as was previously assumed 82,93 . Nel et al have first raised this thought in 2006, emphasizing on the necessity to optimize the classical in vitro toxicity assays as they show several shortcomings when they are being applied for NP toxicity evaluation 28 .…”

“…Nel et al have first raised this thought in 2006, emphasizing on the necessity to optimize the classical in vitro toxicity assays as they show several shortcomings when they are being applied for NP toxicity evaluation 28 . We believe that the major issues with the current in vitro methods are (i) the lack of a complete characterization of the particles, (ii ) the lack of standardisation and guidelines on how to perform a toxicological evaluation in vitro, (iii) the possibility of NP interfering with the assay and therefore lack of appropriate methods to evaluate nanotoxicity and (iv) the shortcomings inherent to the most used classical 2D monocultures 11,82,93,104,105 . It is therefore clear that further research on the optimization of methods is highly recommended in order to obtain reproducible data that would allow drawing firm conclusions regarding NP toxicity.…”

Evolution of toxicological science: the need for change

“…Nanoparticle (NP) toxicology is a rapidly growing concern (Hartung, 2010c;Hartung and Sabbioni, 2011), driven by the dramatic increase in industrial uses of NP and fuelled by public debate. Increasing funding and studies inevitably will result in reports of toxicological effects of NP -both publication bias for positive findings and the multiple testing fallacy (if 20 experiments or endpoints are analysed with p = 0.05 for significance, one should be false-positive) will come to bear here.…”

“…b the shift in pharma industry to new entities, especially biologicals (such as human proteins and antibodies) (Rovida et al, 2015) and medical countermeasures for biological and chemical warfare and terrorism (Hartung and Zurlo, 2012), but also nanoparticles (Hartung, 2010c;Hartung and Sabbioni, 2011), which inevitably require methodological changes c the crisis resulting from the fact that fewer and fewer therapeutic agents make it to the market, with concurrent demands for reviews to determine whether the candidate compounds were correctly selected for development (Hartung, 2013) d market forces have entered the field of alternatives and the first methods are already resulting in turnovers of tens or even hundreds of millions of euros, with lobbyists creating new pressures (Bottini and Hartung, 2009a) e legislation forestalling scientific developments -the most pertinent example being the 7th Amendment of the EU Cosmetics Directive, whereby animal tests are to be banned -even in the absence of alternatives -in order to create (via legislation) the necessary pressure on industry to develop the alternative approaches which are currently lacking (Hartung, 2008a) f globalisation (Bottini et al, 2007): global markets, global companies, the need for the global harmonisation of regulations, new information technologies....…”