WP6: Improving in vitro/in vivo correlations – Metabolism

 

WP6 Results pdf 89 kB, download the entire summary.

 

The objective of WP6 has been to evaluate if the metabolism may influence the toxicity of a compound, since it may actually be a metabolite (a substance produced by biotransformation of the parent chemical in the liver) that is responsible for the observed toxicity.

 

The work in WP6 has been divided into three parts:

1. Simple models for metabolism-dependent toxicity
2. New strategies to incorporate metabolic capabilities into cell lines
3. Computer-based prediction of metabolism and integration of metabolism data into toxicity screening

 

1. Simple models for metabolism-dependent toxicity

In order to evaluate if toxicity is dependent on metabolism, the cytotoxic effects of 21 ACuteTox reference chemicals have been compared between a metabolic component model (primary hepatocytes) and a non-metabolising cell type (HepG2) by the use of MTT assay.

 

For four out of eleven compounds at least two partners have reported a lower IC50 value in hepatocytes than in HepG2 cells, indicating that these compounds produce a higher toxicity in hepatocytes. For five out of twelve compounds the IC50 values were similar in hepatocytes and HepG2 cells, indicating no bioactivation.

 

To examine the robustness of the strategy, intra-assay, inter-assay as well as intra-laboratory variability was investigated for each cell system. A low variability (%CV<10%), both intra-plate and intra-assay, was obtained in all laboratories. The model will be further used for testing of the 41 additional ACuteTox reference chemicals.

 

2. New strategies to incorporate metabolic capabilities into cell lines

HepG2 cells have been transfected with recombinant-defective adenoviral vectors encoding for the major CYP genes involved in foreign compound metabolism. The results demonstrate the applicability of the developed in vitro model as a predictive screening tool, as well as an easy-to-use system to elucidate the mechanism for CYP bioactivation-mediated toxicity of xenobiotics.

 

3. Computer-based prediction of metabolism and integration of metabolism data into toxicity screening

Preliminary evaluation of computer-based prediction models for toxicity, combining in vitro data on toxicity and PBBK/TD (physiologically based biokinetic/toxicodynamic) modeling, including data on in vitro metabolism have been performed in collaboration with WP5. It is concluded that the combined use of DEREK and METEOR is likely to improve the possibility to predict the toxicity of an unknown substance and/or its major metabolites.