GENOTRACE

Among the short-term tests developed to assess the mutagenicity, genotoxicity or carcinogenicity potency of chemical compounds, the micronucleus (MN) assay is largely recognized as a reliable and precise method for detecting chromosome damage. The MN assay is indeed a multi-target genotoxic endpoint, allowing the assessment of clastogenic and aneugenic events. The assay is amenable for automation and allows good extrapolation for potential limits of exposure, with easy measurement in both in vitro and in vivo experimental systems. However, classical MN assay does not provide a dynamic assessment. We propose to bring innovation in the field of tests for hazard and risk assessment of potential mutagens/carcinogens by developing a new in vitro MN assay. The GENOTRACE public/private consortium aims at monitoring the production of chromosome damage in real time, recording in parallel the transcription signal of a genotoxic stress response reporter. To achieve this aim, the project takes advantage of biotracers newly developed by Partners 1 & 2, first to label the nucleus by expressing a chromobody directed against the H2A-H2B histone complex, allowing to visualize chromatin dynamics in real-time, second to follow the DNA Damage Response (DDR) activation by expression of a p21 regulatory dependent reporter. Thus, a positive MN assay associated with either a positive or negative DDR signal will give insights regarding the MN origin, induced by clastogenic or aneugenic mechanisms respectively. These biotracers will be stably expressed into HepaRG metabolically active cells, optimized for MN assay by Partner 3, and the developed in vitro MN assay will be adapted to a medium- to high throughput straightforward readable assay, thanks to the implementation of high content screening imaging protocols and the development of an image analysis and classification-based pipeline. Overall, as our project will allow to determine and classify the genotoxic potential of substances/contaminants in vitro, and in real time, it will bring new capacity to classic genotoxic assays and may lead to progress in the prevention and/or the diagnosis of exposure to genotoxicants.