A cysteine-reactive alkyl hydroquinone modifies topoisomerase IIα, enhances DNA breakage, and induces apoptosis in cancer cells.

Supporting InformationABSTRACT: We previously reported that the anticancer activity of abotanical compound 10′(Z),13′(E),15′(E)-heptadecatrienylhydroquinone[HQ17(3)] was attributed to topoisomerase (Topo) IIα poisoning andthe induction of oxidative damage. HQ17(3) irreversibly inhibits Topo IIαactivity in vitro and is more cytotoxic in leukemia HL-60 cells than inTopo IIα-deficient variant HL-60/MX2 cells, which suggests that Topo IIαis a cellular target of HQ17(3). This study further characterizes themolecular mechanisms of the anticancer activity of HQ17(3). Proteomicanalyses indicated that HQ17(3) reacted with Cys-427, Cys-733, and Cys-997 of recombinant Topo IIα in vitro, whereas it reacted with Cys-427 ofcellular Topo IIα in Huh7 hepatoma cells. The modification of HQ17(3)inhibited Topo IIα catalytic activity, increased the Topo IIα-DNA cleavagecomplex, and caused the accumulation of DNA breakage. In Huh7 cells, HQ17(3) treatment caused prompt inhibition of DNAsynthesis and consequently induced the expression of DNA damage-related genes DDIT3, GADD45A, and GADD45G. Topo IIαinhibition, apoptosis, and oxidative stress were found to account for cytotoxicity caused by HQ17(3). Pretreatment of Huh7 cellswith N-acetylcysteine (NAC) partially attenuated mitochondrial membrane damage, DNA breakage, and caspase activation.However, NAC pretreatment did not diminish HQ17(3)-induced cell death. These results suggest that the anticancer activity ofHQ17(3) is attributed significantly to Topo IIα poisoning. The structural feature of HQ17(3) can be used as a model for thedesign of Topo IIα inhibitors and anticancer drugs.