The ToxTracker reporter system is a mechanistic genotoxicity platform that discriminates clastogenic from aneugenic compounds.Giel Hendriks1, Remco Derr1, Branislav Misovic2, Bruno Morolli2, Fabienne Calléja2 and Harry Vrieling2.
2 Department of Human Genetics, Leiden University Medical Center, Leiden, the Netherlands.
With the increasing production of new chemicals for a wide range of applications in health care, food and as cosmetics, the demand for rapid and reliable toxicity assessment is growing. Novel innovative in vitro systems should ideally not only identify genotoxic properties of chemicals, but also provide insight into the type of cellular damage inflicted in order to more reliably predict human health hazard of novel compounds.
The ToxTracker assay is a mammalian stem cell-based assay that detects activation of specific cellular signalling pathways as the result of biological reactivity of the tested compounds. ToxTracker discriminates between induction of DNA damage, oxidative stress and protein damage by detection of DNA replication stress, NF-kB associated DNA damage signalling, various cellular anti-oxidant responses and activation of the unfolded protein response. In addition, ToxTracker could discriminate clastogenic genotoxins from eugenic compounds based on the differential induction and kinetics to the different genotoxicity reporters.
For data analysis, we have developed the software tool ToxPlot. Automated data analysis and graphical representation of the test results allow clear and rapid assessment of the reactive properties of compounds.
The integrative approach of the ToxTracker assay provides a powerful tool for in vitro carcinogenic hazard identification of chemicals by unveiling activation of specific cellular signalling pathways upon exposure and deliver insight into the underlying mechanism of toxicity.
Mechanistic toxicity profiling of chemicals and nanomaterials by combining the ToxTracker genotoxicity assay and MiToxView mitochondrial toxicity platform.Giel Hendriks1, Remco Derr1, Mathieu Porceddu2, Nelly Buron2, Annie Borgne-Sanchez 2
2 Mitologics S.A.S., Hôpital Robert Debré, 48 Boulevard Sérurier, F-75019, Paris, France. aborgne.sanchez@mitologics.com
The ever-increasing number of chemical compounds as well as nanomaterials that are developed by industry poses a potential threat for human health. These compounds and materials may react with various biomolecules thereby disrupting organelle function and cellular homeostasis.
ToxTracker is a mammalian stem cell-based GFP reporter assay that discriminates between induction of DNA damage, oxidative stress, protein damage and general cytotoxicity. The MiToxView platform on mouse liver mitochondria allows identification of compounds inducing direct mitochondrial damages by assessing global membrane permeabilisation (swelling), transmembrane potential (ΔΨm) loss, cytochrome c release and oxygen consumption through respiratory chain complex I and complex II.
We evaluated the improved in vitro toxicity screening by combining the mechanistic ToxTracker and MiToxView platforms. For this we tested a selection of 20 primarily DILI compounds and various metal or metal-oxide nanoparticles. ToxTracker reliably identified the genotoxic properties of the tested chemicals and nanoparticles. These substances generally also affected mitochondrial function. Various tested DILI compounds that induced mitochondrial toxicity in MiToxView activated the oxidative stress or unfolded protein response in the ToxTracker platform. However, there were several established DILI compounds that did not activate the ToxTracker reporters but clearly affected mitochondrial integrity.
Together, our data suggest that integration of a cellular signalling reporter assay with a functional mitochondrial toxicity system could be a valuable platform for reliable identification of the hazardous properties and the mechanism of action of new compounds and nanomaterials.
