Nanomaterial Toxicity Testing in the 21st Century: Use of a Predictive Toxicological Approach and High-Throughput Screening

Abstract

Conspectus The production of engineered nanomaterials (ENMs) is a scientific breakthrough in material design and the development of new consumer products. While the successful implementation of nanotechnology is important for the growth of the global economy, we also need to consider the possible environmental health and safety (EHS) impact as a result of the novel physicochemical properties that could generate hazardous biological outcomes. In order to assess ENM hazard, reliable and reproducible screening approaches are needed to test the basic materials as well as nano-enabled products. A platform is required to investigate the potentially endless number of bio-physicochemical interactions at the nano/bio interface, in response to which we have developed a predictive toxicological approach. We define a predictive toxicological approach as the use of mechanisms-based high throughput screening in vitro to make predictions about the physicochemical properties of ENMs that may lead to the generation of pathology or disease outcomes in vivo. The in vivo results are used to validate and improve the in vitro high throughput screening (HTS) and to establish structure-activity relationships (SARs) that allow hazard ranking and modeling by an appropriate combination of in vitro and in vivo testing. This notion is in agreement with the landmark 2007 report from the US National Academy of Sciences, “Toxicity Testing in the 21st Century: A Vision and a Strategy” (http://www.nap.edu/catalog.php?record_id=11970), which advocates increased efficiency of toxicity testing by transitioning from qualitative, descriptive animal testing to quantitative, mechanistic and pathway-based toxicity testing in human cells or cell lines using high throughput approaches. Accordingly, we have implemented HTS approaches to screen compositional and combinatorial ENM libraries to develop hazard ranking and structure-activity relationships that can be used for predicting in vivo injury outcomes. This predictive approach allows the bulk of the screening analysis and high volume data generation to be carried out in vitro, following which limited, but critical, validation studies are carried out in animals or whole organisms. Risk reduction in the exposed human or environmental populations can then focus on limiting or avoiding exposures that trigger these toxicological responses as well as implementing safer design of potentially hazardous ENMs. In this communication, we review the tools required for establishing predictive toxicology paradigms to assess inhalation and environmental toxicological scenarios through the use of compositional and combinatorial ENM libraries, mechanism-based HTS assays, hazard ranking and development of nano-SARs. We will discuss the major injury paradigms that have emerged based on specific ENM properties, as well as describing the safer design of ZnO nanoparticles based on characterization of dissolution chemistry as a major predictor of toxicity.

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

“…Biochemical assays with a fluorometric or spectrophotometric read-out, including enzymatic and (enzyme-linked) immunoassays, are used to the utmost extend due to the relatively short duration of most assays, the uncomplicated detection principle and straight forward data processing 83 . Furthermore the possibility of upscaling and automation of the execution, detection and data processing make these assays highly convenient with regard to a future high content screening approach for every newly synthetized NP 83,84 . Another popular approach is staining the entire cell or specific cellular components with cellular dyes, fluorescently labeled antibodies (AB) or molecular probes, which will interact with a specific biomolecule 40,78,85 .…”

“…Since it was observed that NP can cause multiple effects via different mechanisms, awareness has risen on the necessity to evaluate nanotoxicity in vitro by a multiparametric method 43,83 .…”

“…However, many obstacles still need to be overcome before the first QSAR can be implemented that allows the prediction of the adverse effects of any NP towards human health. Therefore, clear correlations between the NP's physicochemical properties and observed effect must be found, large-scale comparative studies should be performed and the QSARs must be validated, which requires the indentification of reference nanomaterials with well-known effects 40,83 .…”

High throughput toxicity screening and intracellular detection of nanomaterials

“…There is therefore an urgent need to clarify the toxic effects of NPs and NMs and to elucidate the mechanisms involved in their toxicity. In view of the large number of NMs currently in use, high throughput screening (HTS) techniques aimed at accurately predicting and assessing toxicity are clearly needed; given the availability of reliable toxicity metrics, the HTS approach will generate large and valuable data sets 3, 4…”

“…HTS is defined as the use of automated tools to facilitate rapid execution of a large number and variety of biological assays that may include several substances in each assay 4. HTS was introduced in the pharmaceutical and chemical industries as a rapid way of evaluating effects of many novel compounds.…”