<p>Supplementary Figures S1-S4- Cytotoxicity data for four cell lines with all compounds used in the study. S5-S7 - C-trapping via western blotting with in three cell lines. S8-S9 - C-trapping via imaging on populations of cells with high and low magnifications. S10- Time dependence of c-trapping S11 - Comparison of drug accumulations in the nuclei of cells via autofluorescence. S12 - Effect of inhibition of MDR on cytotoxicity of the drugs. S13 - Comparison of the potency of c-trapping caused by different compounds in HCT116 cells S14. Quantitation of Histone H4 eviction. S15. Immunofluorescent staining of histone H3. S16. Comparison of patterns of c-trapping caused by high concentrations of the drugs. S17-S20 - Analyses of genome-wide distribution of FACT via ChIP-seq. S21. Assessment of DNA damaging properties of the compounds. S22. Assessment of correlations between different parameters measured. S</p>
ARTICLE ABSTRACT
Precisely how DNA-targeting chemotherapeutic drugs trigger cancer cell death remains unclear, as it is difficult to separate direct DNA damage from other effects in cells. Recent work on curaxins, a class of small-molecule drugs with broad anticancer activity, shows that they interfere with histone–DNA interactions and destabilize nucleosomes without causing detectable DNA damage. Chromatin damage caused by curaxins is sensed by the histone chaperone FACT, which binds unfolded nucleosomes becoming trapped in chromatin. In this study, we investigated whether classical DNA-targeting chemotherapeutic drugs also similarly disturbed chromatin to cause chromatin trapping of FACT (c-trapping). Drugs that directly bound DNA induced both chromatin damage and c-trapping. However, chromatin damage occurred irrespective of direct DNA damage and was dependent on how a drug bound DNA, specifically, in the way it bound chromatinized DNA in cells. FACT was sensitive to a plethora of nucleosome perturbations induced by DNA-binding small molecules, including displacement of the linker histone, eviction of core histones, and accumulation of negative supercoiling. Strikingly, we found that the cytotoxicity of DNA-binding small molecules correlated with their ability to cause chromatin damage, not DNA damage. Our results suggest implications for the development of chromatin-damaging agents as selective anticancer drugs.Significance: These provocative results suggest that the anticancer efficacy of traditional DNA-targeting chemotherapeutic drugs may be based in large part on chromatin damage rather than direct DNA damage. Cancer Res; 78(6); 1431–43. ©2018 AACR.
