Towards an integrated in vitro strategy for repeated dose toxicity testing

Abstract

Currently, there is clear political incentive in the EU to minimize the number of animals involved in the chemical risk assessment process, particularly holding true for the cosmetics sector. Stringent requirements of full replacement of animals in safety testing are laid down in the EU Cosmetics Directive 76/768/EEC by the implementation of the 7th Amendment (2003/15/EC), setting a rigid time frame for the availability of valid 3R-alternative methods for the safety testing of cosmetic products and their ingredients (EU 2003). An important step forward to achieve this goal has recently been made through a joint initiative by DG Research and Colipa (The European Cosmetics Association). A cluster of calls was launched under the FP7 health programme specifically addressing the replacement of repeated dose systemic toxicity testing in human safety assessment. As such, 6 large-scale collaborative research projects started from January 2011 onwards, covering key themes related to human stem cell technology (SCREEN-TOX), organ-simulating devices (HeMiBio), human-relevant biomarker detection (DETECTIVE), computational modelling (COSMOS), systems biology (NOTOX) and integrated data analysis (TOXBANK). An umbrella coordination and support action project (COACH) is responsible for monitoring progress and potential gaps occurring throughout the 5-year running period. A major task, however, will be to streamline and combine the outcomes of the different projects in order to establish a realistic in vitro strategy to assess repeated dose toxicity.As in all projects the liver is involved as target organ, one might expect that hepatotoxicity could be used as a test case. This would be of particular interest since it is the major type of toxicity observed during in vivo repeated dose toxicity testing for cosmetic ingredients (Pauwels et al. 2009) and pharmaceutical compounds (Russmann et al. 2009). Figure 1 demonstrates how such an in vitro testing strategy could look, taking into consideration, the current approach of quantitative risk assessment for cosmetic ingredients using data derived from animal studies. Thus, a margin of safety (MoS) of at least 100 is applied for intraspecies and interspecies variation using the following equation:MoS ¼ NOAEL ðno observable adverse effect levelÞ systematic exposure dosage ! 100In the proposed strategy, a computed NOAEL value, preferably derived from human data (hNOAEL c ), could be used for the compound of interest. This estimated in vivo dosage could be obtained via processing of the compound concentration (hNOAEC) that does not cause any adverse effect using an in vitro model. For that purpose, physiologically based pharmacokinetic (PBPK) computational modelling can be applied taking into account an appropriate uncertainty factor for the in vitro/in vivo extrapolation. The hNOAEC value of the compound of interest could, for example,

A roadmap for the development of alternative (non-animal) methods for systemic toxicity testing

“…However, the implementation of decision trees and tiered approaches, i.e., integrated testing strategies (ITS), will contribute greatly to the reducing the use of animals (Grindon et al, 2008;Vanhaecke et al, 2011). This is something that can begin now and be modified as more and more alternative tests become available.…”

“…This is something that can begin now and be modified as more and more alternative tests become available. Approaches such as the decision tree proposed by Vanhaecke et al (2011) for predicting liver toxicity represent a very good start. The authors propose integrating computational and cell-based toxicity information, along with pre-existing data, to arrive at a theoretical NOAEL and assess an acceptable margin of safety.…”

“…Before implementing the ITS, a very careful evaluation of the assays is needed. For example, Vanhaecke et al (2011) suggests usof alternative methods and to estimate the time necessary to achieve a full replacement of animals in the cosmetic industry. An extensive and detailed document (Adler et al, 2011) summarized their conclusions.…”

Human Pluripotent Stem Cell Applications in Drug Discovery and Toxicology – An overview

“…Human Stringent cosmetics legislation amending directives especially within the European Union (EU) related to complete replacement of animal models in cosmetic industry safety testing by alternative methods has emphasized an urgent need for the development of reduction, refinement and replacement (3R) of the existing animal studies [25]. In order to fill gaps in non-animal alternative methods and to focus on complex RDT a research initiative called "Safety Evaluation Ultimately Replacing Animal Testing 1 (SEURAT-1)" composed of six complimentary research projects was launched in 2011 and jointly funded by the European Commission's FP7 HEALTH Programme and Cosmetics Europe (http://www.seurat-1.eu/).…”

“…Human ESC based in vitro reproductive toxicity, neurotoxicity, toxicogenomics, proteomics and kinetics were tested for their predictive value in the identification of toxicity endpoints (http://www.esnats.eu). The RDT delivers the No Observable Adverse Effect (NOAEL), which is used in calculation of the substance safety parameters [25]. PSC-derived models hold a great potential for refinement of current models of cardiotoxicity.…”