Identification of transcriptome signatures and biomarkers specific for potential developmental toxicants inhibiting human neural crest cell migration

Abstract

The in vitro test battery of the European research consortium ESNATS (‘novel stem cell-based test systems’) has been used to screen for potential human developmental toxicants. As part of this effort, the migration of neural crest (MINC) assay has been used to evaluate chemical effects on neural crest function. It identified some drug-like compounds in addition to known environmental toxicants. The hits included the HSP90 inhibitor geldanamycin, the chemotherapeutic arsenic trioxide, the flame-retardant PBDE-99, the pesticide triadimefon and the histone deacetylase inhibitors valproic acid and trichostatin A. Transcriptome changes triggered by these substances in human neural crest cells were recorded and analysed here to answer three questions: (1) can toxicants be individually identified based on their transcript profile; (2) how can the toxicity pattern reflected by transcript changes be compacted/dimensionality-reduced for practical regulatory use; (3) how can a reduced set of biomarkers be selected for large-scale follow-up? Transcript profiling allowed clear separation of different toxicants and the identification of toxicant types in a blinded test study. We also developed a diagrammatic system to visualize and compare toxicity patterns of a group of chemicals by giving a quantitative overview of altered superordinate biological processes (e.g. activation of KEGG pathways or overrepresentation of gene ontology terms). The transcript data were mined for potential markers of toxicity, and 39 transcripts were selected to either indicate general developmental toxicity or distinguish compounds with different modes-of-action in read-across. In summary, we found inclusion of transcriptome data to largely increase the information from the MINC phenotypic test.Electronic supplementary materialThe online version of this article (doi:10.1007/s00204-015-1658-7) contains supplementary material, which is available to authorized users.

Reference compounds for alternative test methods to indicate developmental neurotoxicity (DNT) potential of chemicals: example lists and criteria for their selection and use

“…This precludes an evaluation of test predictivity based solely on the correlation of its results with in vivo findings (Leist et al, 2012). One way forward would involve the three following activities: (a) obtaining more knowledge on modes of action of DNT chemicals by profiling them in a broad set of well characterized and robust in vitro test systems (Behl et al, 2015; Zimmer et al, 2014; Daneshian et al, 2016; Hirsch et al, 2016; Pallocca et al, 2016); (b) optimizing in vitro test systems, by using endpoint specific controls and already well-characterized DNT compounds; (c) using steps (a) and (b) in an iterative fashion to optimize test systems and test methods.…”

“…In addition, most screens have up to now been performed at fixed compound concentrations that are not related to the pharmacological potency of the compounds screened. A change of this practice has been suggested for the ESNATS test battery (Pallocca et al, 2015; Zimmer et al, 2014), for which initial concentrations for testing have been based on a biological/mechanistic rationale. In addition, for many omics studies the chosen concentration is anchored to a biological effect (e.g.…”

Reference compounds for alternative test methods to indicate developmental neurotoxicity (DNT) potential of chemicals: Example lists and criteria for their selection and use_suppl

“…Based on comparing data generated across multiple diverse test systems, the most sensitive endpoints and the most reliable test them in a broad set of well characterized and robust in vitro test systems Zimmer et al, 2014;Daneshian et al, 2016;Hirsch et al, 2016;Pallocca et al, 2016); (b) optimizing in vitro test systems by using endpoint specific controls and already well-characterized DNT compounds; (c) using steps (a) and (b) in an iterative fashion to optimize test systems and test methods.…”

“…In addition, most screens have up to now been performed at fixed compound concentrations that are not related to the pharmacological potency of the compounds screened. A change of this practice has been suggested for the ESNATS test battery (Pallocca et al, 2016;Zimmer et al, 2014), for which initial concentrations used for testing have been based on a biological/mechanistic rationale. In addition, for many omics studies the chosen concentration is anchored to a biological effect (e.g., maximum non-cytotoxic concentration).…”

Stem cells in toxicological research

“…Chemicals are added during the differentiation period to test whether this process is disturbed (Pallocca et al 2016;Rempel et al 2015). Such approaches led to the establishment of the developmental toxicity index, which quantifies the fraction of genes involved in normal differentiation whose expression is compromised by the test chemicals (Shinde et al 2017).…”