Evaluation of the genotoxic potential of apoptosis inducers with the γH2AX assay in human cells

Human risk assessment of genotoxic chemicals is an important area of research. However, the specificity of in vitro mammalian genotoxicity assays is sometime low, as they yield to misleading positive results that are not observe in in vivo studies. Apoptosis can be a confounding factor in the interpretation of the results. Recently, a new strategy for genotoxicity screening, based on the combined analysis of phosphorylated histones H2AX (gamma H2AX) and H3 (pH3), was proposed to discriminate efficiently aneugenic from clastogenic compounds. However, gamma H2AX biomarker could also be induce by apoptosis. The aim of the present study was to investigate the specificity of this genotoxic biomarker. For this purpose, we analyzed 26 compounds inducing apoptosis by different mechanism of action, with the gamma H2AX assay in three human cell lines after 24 h treatment. Most of the tested chemicals were negative in the assay, whatever the cell line tested. The few compounds that generated positive data have also been report positive in other genotoxicity assays. The data presented here demonstrate that the gamma H2AX assay is not vulnerable to the generation of misleading positive results by apoptosis inducers. Currently, no formal guidelines have been approve for the gamma H2AX assay for regular genotoxicity studies, but we suggest that this biomarker could be used as a new standard genotoxicity assay.