In-house assessment of a modified in vitro cytotoxicity assay for higher throughput estimation of acute toxicity

The main objective of this study was to assess an in-house 3T3 NRU cytotoxicity assay for compatibility with a prediction model for acute rodent oral toxicity endorsed by an NIEHS–ICCVAM workshop. The aim is to use the NRU assay as one test component of HTS strategies for both acute oral toxicity and acute skin irritation, enabling the rejection of the most toxic materials and prioritisation of other materials for further testing. Groups of model cytotoxins and irritants were tested using the NRU assay and their EC₅₀ values obtained from dose–response curves. These values were compared with those estimated from a limited (three)-dose protocol, deemed more suitable for HTS. A good correlation was observed between the EC₅₀ values from both dose–response curves (R²=0.94). The relationships between EC₅₀ values and acute rodent oral toxicity were compared by application of the prediction model to the model cytotoxins. The results from both full and limited dose–responses fitted within the acceptance limits of the prediction model, with regression lines similar to that of the model. Results indicated that the performance of the currently used 3T3 NRU cytotoxicity assay was similar to that of the assays used to generate the data employed in developing the prediction model. This prediction model can be applied with both the standard and HT assays to estimate acute rodent oral toxicity.