DETECTIVE : Detection of endpoints and biomarkers of repeated dose toxicity using in vitro systems.

As one of the building blocks of the SEURAT-1 Research Initiative (“Safety Evaluation Ultimately Replacing Animal Testing”), DETECTIVE focused on a key element on which in vitro toxicity testing relies: the development of robust and reliable, sensitive and specific in vitro biomarkers and surrogate endpoints that can be used for safety assessment of chronically acting toxicants relevant for humans.
Emphasis was given to systematic exploitation of a battery of complementary functional and “-omics” readouts, including high content and high throughput screening platforms to identify and investigate human biomarkers in cellular models for repeated dose in vitro testing. While functional parameters give more insights into the effects of toxicants on specific cell functions of interest, “-omics” techniques deliver data on the entire cellular situation at molecular level. Importantly, DETECTIVE has performed for the first time an in-depth investigation of repeated dose effects on epigenetics and microRNA (miRNA) expression, thus exploring whether such analyses deepen our understanding of toxic modes of action. In the last years, these two parameters have been identified as critical for cell behaviour and it has been a challenging task to determine whether long-term application of chemicals will affect cells at this level.
Upon combination and subsequent integration of the various readouts, biomarkers for predicting human long-term toxicity in vitro have been obtained. Based on integrative statistical analysis, systematic verification and correlation with in vivo data, relevant, specific, sensitive and predictive biomarkers have been selected.

DETECTIVE concentrated on hepatotoxic, cardiotoxic and nephrotoxic effects representing three target organs of repeated dose toxicity. Ultimately, developed concepts will also be applicable to other organs or organ systems affected by systemic toxicants such as the nervous system. Furthermore, DETECTIVE was able to define both generic and organ-specific human toxicity pathways.
DETECTIVE, as a multidisciplinary collaboration of European researchers, has achieved its key scientific objectives by
• Establishing a successful private-public partnership addressing an important societal need
• Developing new tools and technologies together with a smart integration approach represents a substantial step forward to animal free safety testing
• Creating a screening pipeline for identifying biomarkers and surrogate endpoints relevant for assessing human long term organ toxicity and the data derived from which has been regularly uploaded to the Quretec (64 GB of data) and TOXBANK data repository (98 GB of data), which made available to research community.
• Compiling a repository of
o verified, stable and easy-to-measure functional and “-omics” biomarkers of different organs, including GLP-compliant SOPs for selected, most relevant biomarkers
o human toxicity pathways relevant for different organs (in cooperation with TOXBANK and NOTOX)
o concepts for translating the knowledge gained to toxicity of the entire organism
o adverse outcome pathways (AOPs)
• Construct from Bile Salt Export Pump Inhibition to Cholestasis
• Cardiotoxicity
• Novel public toxicogenomics directory of chemically exposed human hepatocytes (146 compounds)
• New live cell imaging pathway-based toxicity reporters for identification of hepatotoxicant-induced cellular stress responses
• Human skin-derived precursors (hSKP) as a novel cell source for in vitro screening of compounds that induce liver steatosis
• Completion of 3 case studies at SEURAT-1 level