Assessing dose metrics in in vitro cell assays to improve in vitro in vivo dose extrapolations

Abstract

High requirements and challenges formed by legislations like REACH and the 7th amendment of cosmetics have accelerated the development of new alternative toxicology testing methods. In vitro cell systems combined with in silico methods have been deemed good alternatives for toxicology testing, aiming to reduce or even replace conventional animal toxicity experiments. Unfortunately these methods are not ready to replace animal based toxicity assays yet for several reasons. One of the issues as mainly discussed in this thesis, is the lack of a standardised dose metric for use in dose-response relationships that are to be extrapolated f om in vitro cell systems to in vivo. Often total or nominal concentrations are used to express in vitro derived toxicity while these do not account for possible reductions in bioavailability, thus reducing the biologically effective dose through evaporation, binding to well plastic, serum, cell membranes etc. Differences in exposure media between in vitro assays and between in vitro and in vivo, may cause differences in the biologically effective dose even though nominal concentrations are equal. This can subsequently lead to different predictions of in vivo responses. Therefore this thesis discusses numerous alternative dose metrics available that may be used to improve the in vivo predictions. It is recommended to first choose the right external dose metric, either nominal or freely dissolved based on the physiological parameters of the test compound. Additionally, a choice can be made for an internal concentration, closer to the target site to improve the extrapolation of in vitro effect concentrations to equivalent in vivo doses. Finally the chosen metric can be integrated (AUC) or weighted (TWA) in the case of prolonged exposure and depending on the mechanisms of action. Further research needs to focus on whether internal concentrations are truly worth measuring and what cut-off value in bioavailability reduction should be used to choose between either free or nominal concentrations.