Leveraging toxicogenomics in a weight of evidence approach to demonstrate a CAR-mediated mode of action for cyclobutrifluram-related mouse liver tumors

Abstract Toxicogenomics-based approaches are powerful tools for investigating the mode of action and human relevance of chemical-induced effects in animal toxicity studies, thus supporting human risk assessment and regulatory decisions. Here we incorporated transcriptomics and metabolomics into a mode of action assessment of male mouse liver tumors observed following 80-week dietary exposure to cyclobutrifluram, a novel complex II succinate dehydrogenase inhibitor (SDHI) agrochemical. The assessment was conducted using the framework developed by the International Programme on Chemical Safety (IPCS) and the International Life Sciences Institute (ILSI), based on activation of the nuclear constitutive androstane receptor (CAR) and subsequent downstream events that have been established as human non-relevant. Cyclobutrifluram was shown to activate rat, mouse and human CAR in in vitro transactivation assays. Dietary administration of cyclobutrifluram in male mice was associated with time and/or dose-dependent liver weight increases, centrilobular hepatocellular hypertrophy, induction of CAR-related liver enzyme activity, specifically CYP2B and CYP3A, and hepatocellular proliferation. Transcriptomics analysis of mouse liver identified cyclobutrifluram-induced gene expression profiles consistent with CAR activation, based on published signatures and similarity to the reference CAR inducer phenobarbital. Metabolomics analysis of mouse plasma and liver further indicated that cyclobutrifluram induced similar biochemical changes as phenobarbital, with no evidence of any additional activity. Overall, this work demonstrates how toxicogenomics can provide valuable weight of evidence to identify the mode of action for chemical-induced rodent liver tumors and to exclude alternative modes of action.