Editor's Highlight: Analysis of the Effects of Cell Stress and Cytotoxicity onIn VitroAssay Activity Across a Diverse Chemical and Assay Space

Abstract

Chemical toxicity can arise from disruption of specific biomolecular functions or through more generalized cell stress and cytotoxicity-mediated processes. Here, responses of 1060 chemicals including pharmaceuticals, natural products, pesticidals, consumer, and industrial chemicals across a battery of 815 in vitro assay endpoints from 7 high-throughput assay technology platforms were analyzed in order to distinguish between these types of activities. Both cell-based and cell-free assays showed a rapid increase in the frequency of responses at concentrations where cell stress/cytotoxicity responses were observed in cell-based assays. Chemicals that were positive on at least 2 viability/cytotoxicity assays within the concentration range tested (typically up to 100 μM) activated a median of 12% of assay endpoints whereas those that were not cytotoxic in this concentration range activated 1.3% of the assays endpoints. The results suggest that activity can be broadly divided into: (1) specific biomolecular interactions against one or more targets (eg, receptors or enzymes) at concentrations below which overt cytotoxicity-associated activity is observed; and (2) activity associated with cell stress or cytotoxicity, which may result from triggering specific cell stress pathways, chemical reactivity, physico-chemical disruption of proteins or membranes, or broad low-affinity non-covalent interactions. Chemicals showing a greater number of specific biomolecular interactions are generally designed to be bioactive (pharmaceuticals or pesticidal active ingredients), whereas intentional food-use chemicals tended to show the fewest specific interactions. The analyses presented here provide context for use of these data in ongoing studies to predict in vivo toxicity from chemicals lacking extensive hazard assessment.

Use of high-throughput in vitro toxicity screening data in cancer hazard evaluations by IARC Monograph Working Groups

“…As data grow and analysis methods mature, updates to activity calls and AC 50 values in the database will hopefully present a more precise portrait of underlying biology. For example, data obtained for the applications described here were obtained prior to publication of a strategy to post-filter results at concentrations exceeding a putative cytotoxic interference threshold (Judson et al, 2016). The appropriateness of accounting for cytotoxicity across molecular targets linked to cell cycle and cellular stress can also be argued.…”

“…The authors note that methodological refinements introduce periodic updates to activity calls and AC 50 values in the database, although previous versions are maintained at the public download site to allow for exact replication of particular applications. Also, the data used for these analyses were not subject to further evaluations of the effects of cell stress and cytotoxicity on in vitro assay activity that were implemented into ToxCast data dashboards more recently (Judson et al, 2016). …”

“…However, both active and inactive assay endpoints were included across in all analyses. These were not subject to further evaluations of the effects of cell stress and cytotoxicity on in vitro assay activity that were implemented into ToxCast data dashboards more recently (Judson et al, 2016). …”

Development and Validation of a Computational Model for Androgen Receptor Activity

“…32 We flagged potential nonselective assay hits attributed to cell stress using the distance between the logAC 50 (assay) and the median logAC 50 (cytotox) with respect to the global cytotoxicity median of the median absolute deviation (MAD) of the logAC 50 (cytotox) distributions across all chemicals. Details are given in ref (32).…”

“…32 We flagged potential nonselective assay hits attributed to cell stress using the distance between the logAC 50 (assay) and the median logAC 50 (cytotox) with respect to the global cytotoxicity median of the median absolute deviation (MAD) of the logAC 50 (cytotox) distributions across all chemicals. Details are given in ref (32). Briefly, for chemicals with two or more positive responses in assays measuring cytotoxicity or inhibition of proliferation, a “ Z -score” was calculated for each AR pathway assay hit asA large Z -score indicates an in vitro assay logAC 50 at concentrations significantly below those causing cytotoxicity or inhibiting proliferation.…”