title: Fingerprinting of neurotoxic compounds using a mouse embryonic stem cell dual luminescence reporter assay.
abstract: Identification of neurotoxic drugs and environmental chemicals is an important challenge. However, only few tools to address this topic are available. The aim of this study was to develop a neurotoxicity/developmental neurotoxicity (DNT) test system, using the pluripotent mouse embryonic stem cell line CGR8 (ESCs). The test system uses ESCs at two differentiation stages: undifferentiated ESCs and ESC-derived neurons. Under each condition, concentration-response curves were obtained for three parameters: activity of the tubulin alpha 1 promoter (typically activated in early neurons), activity of the elongation factor 1 alpha promoter (active in all cells), and total DNA content (proportional to the number of surviving cells). We tested 37 compounds from the ESNATS test battery, which includes polypeptide hormones, environmental pollutants (including methylmercury), and clinically used drugs (including valproic acid and tyrosine kinase inhibitors). Different classes of compounds showed distinct concentration-response profiles. Plotting of the lowest observed adverse effect concentrations (LOAEL) of the neuronal promoter activity against the general promoter activity or against cytotoxicity, allowed the differentiation between neurotoxic/DNT substances and non-neurotoxic controls. Reporter activity responses in neurons were more susceptible to neurotoxic compounds than the reporter activities in ESCs from which they were derived. To relate the effective/toxic concentrations found in our study to relevant in vivo concentrations, we used a reverse pharmacokinetic modeling approach for three exemplary compounds (teriflunomide, geldanamycin, abiraterone). The dual luminescence reporter assay described in this study allows high-throughput, and should be particularly useful for the prioritization of the neurotoxic potential of a large number of compounds.
authors.affiliation: Department of Pathology and Immunology, Medical School, University of Geneva, Geneva, Switzerland.
authors.forename: Marilena
authors.initials: M
authors.lastname: Colaianna
authors.affiliation: Department of Pathology and Immunology, Medical School, University of Geneva, Geneva, Switzerland.
authors.forename: Sten
authors.initials: S
authors.lastname: Ilmjärv
authors.affiliation: Quretec Ltd, Ülikooli 6a, Tartu, Estonia.
authors.forename: Hedi
authors.initials: H
authors.lastname: Peterson
authors.affiliation: Department of Pediatrics, Geneva University Hospital, Geneva, Switzerland.|Department of Genetic and Laboratory Medicine, Geneva University Hospital, Centre Medical Universitaire, Rue Michel-Servet, 1211, Geneva 4, Switzerland.
authors.forename: Ilse
authors.initials: I
authors.lastname: Kern
authors.affiliation: Department of Pathology and Immunology, Medical School, University of Geneva, Geneva, Switzerland.
authors.forename: Stephanie
authors.initials: S
authors.lastname: Julien
authors.affiliation: NEURIX SA, Geneva, Switzerland.
authors.forename: Mathurin
authors.initials: M
authors.lastname: Baquié
authors.affiliation: Doerenkamp-Zbinden Chair for In Vitro Toxicology and Biomedicine, University of Konstanz, Constance, Germany.
authors.forename: Giorgia
authors.initials: G
authors.lastname: Pallocca
authors.affiliation: TNO, Zeist, The Netherlands.|BioMarin Pharmaceutical Inc., Leiden, The Netherlands.
authors.forename: Sieto
authors.initials: S
authors.lastname: Bosgra
authors.affiliation: Institute of Neurophysiology and Center for Molecular Medicine Cologne (CMMC), University of Cologne, Cologne, Germany.
authors.forename: Agapios
authors.initials: A
authors.lastname: Sachinidis
authors.affiliation: Leibniz Research Centre for Working Environment and Human Factors (IfADo), Technical University of Dortmund, Dortmund, Germany.
authors.forename: Jan G
authors.initials: JG
authors.lastname: Hengstler
authors.affiliation: Doerenkamp-Zbinden Chair for In Vitro Toxicology and Biomedicine, University of Konstanz, Constance, Germany.
authors.forename: Marcel
authors.initials: M
authors.lastname: Leist
authors.affiliation: Department of Pathology and Immunology, Medical School, University of Geneva, Geneva, Switzerland. Karl-Heinz.Krause@unige.ch.|Department of Genetic and Laboratory Medicine, Geneva University Hospital, Centre Medical Universitaire, Rue Michel-Servet, 1211, Geneva 4, Switzerland. Karl-Heinz.Krause@unige.ch.
authors.forename: Karl-Heinz
authors.initials: KH
authors.lastname: Krause
mesh_terms: D000818:Animals; D015415:Biomarkers / Q000378:metabolism; D002450:Cell Communication / Q000187:drug effects*; D002454:Cell Differentiation; D002460:Cell Line; D002470:Cell Survival / Q000187:drug effects; D018920:Coculture Techniques; D004353:Drug Evaluation, Preclinical; D015507:Drugs, Investigational / Q000009:adverse effects; D004784:Environmental Monitoring; D004785:Environmental Pollutants / Q000633:toxicity; D017930:Genes, Reporter / Q000187:drug effects; D008164:Luminescent Proteins / Q000235:genetics / Q000378:metabolism; D051379:Mice; D000066450:Mouse Embryonic Stem Cells / Q000166:cytology / Q000187:drug effects* / Q000378:metabolism; D009474:Neurons / Q000166:cytology / Q000187:drug effects* / Q000378:metabolism; D009498:Neurotoxins / Q000633:toxicity*; D020648:Peptide Elongation Factor 1 / Q000235:genetics / Q000378:metabolism; D039904:Pluripotent Stem Cells / Q000166:cytology / Q000187:drug effects* / Q000378:metabolism; D011401:Promoter Regions, Genetic / Q000187:drug effects; D017154:Stromal Cells / Q000166:cytology / Q000187:drug effects* / Q000378:metabolism; D014404:Tubulin / Q000235:genetics / Q000378:metabolism
chemical_list: D015415:Biomarkers; D015507:Drugs, Investigational; D004785:Environmental Pollutants; D008164:Luminescent Proteins; D009498:Neurotoxins; D020648:Peptide Elongation Factor 1; C569741:Tuba1a protein, mouse; D014404:Tubulin
keywords: In vitro screening; Mouse embryonic stem cells; Neuroactivity; Neuronal differentiation; Neurotoxicity