Dr. Escher, could you give us an example of a project?
In the EU-ToxRisk project, we recently completed a case study assessing the toxicity of a branched carboxylic acid by using a combination of in-vitro and in-silico models. We used a read-across approach with ten structural analogues. In addition, in-vivo data from three structurally related analogues were available. A team of partners from 14 academic institutions was involved in the design of the testing strategy. Based on the available in-vivo data, we assumed that the target compound 2-ethylbutyric acid (2-EBA) is a liver toxicant with special concern for hepatic steatosis. The NAM data proved this read-across hypothesis wrong by showing a consistent trend with regard to toxikokinetics and toxikodynamics within the grouped compounds.
To characterize the toxicodynamics, published signaling pathways leading to steatosis were compiled from literature and described in an AOP network. Two high-throughput models were used to measure some of the described molecular initiating events, MIEs for short. Furthermore, three liver models were used to measure lipid accumulation, which is seen as a direct in-vitro surrogate for in-vivo steatosis. It was shown that the number of activated MIEs and induction of lipid accumulation increases with the side chain length of the tested carboxylic acid, whereas short-chain analogues like 2-EBA remained inactive. This finding is in very good agreement with the in-vivo data.
The next question that arises is how to derive a dose, based on in-vitro tests, below which there is no risk to human health. For this purpose, a human PBPK model was used in the EU-ToxRisk project. By means of this PBPK model, an equivalent human dose was calculated based on the minimal-effect concentrations from all available in-vitro assays, a process known as quantitative in vitro to in vivo extrapolation. The results of this successful case study are currently being reviewed by toxicologists from national and international regulatory authorities. They are planned to be published before the end of 2019.