Salvicine triggers DNA double-strand breaks and apoptosis by GSH-depletion-driven H 2O 2 generation and topoisomerase II inhibition

Abstract

Glutathione (GSH), as the major small-molecule antioxidant in cells, has been implicated in the regulation of cell proliferation and apoptosis. Salvicine (SAL), a novel diterpenoid quinone compound, exhibits potent antitumor activities both in vitro and in vivo by poisoning topoisomerase II (Topo II) and has entered Phase II clinical trials for cancer therapy. Herein, we provide further evidence that SAL-induced DNA double-strand breaks (DSBs) and apoptosis by GSH depletion drives H 2O 2 generation and Topo II inhibition. Our data reveal that treatment with SAL results in a pronounced increase in intracellular H 2O 2 and is accompanied by the occurrence of DNA DSBs and apoptosis in epithelial HeLa cells. Furthermore, SAL was also noted to trigger a dramatic depletion of intracellular GSH via its direct reaction with GSH. Importantly, the introduction of GSH and overexpression of catalase antagonized SAL-mediated DNA DSBs and apoptosis, and the GSH synthesis inhibitor dl-buthionine-[ S,R]-sulfoximine reduced SAL-mediated H 2O 2 generation, indicating that SAL-mediated H 2O 2 generation is derived from intracellular GSH depletion. Notably, SAL-mediated Topo II inhibition was also concentration-dependently reversed by GSH. Furthermore, we found that Topo II-defective HL-60/MX2 cells were almost completely resistant to SAL-induced DNA DSBs, suggesting that, in addition to its direct inhibitory effect on Topo II, SAL-mediated H 2O 2 generation may also trigger DNA DSBs via poisoning of Topo II. All these findings together suggest that GSH-depletion-driven H 2O 2 generation and Topo II inhibition are both critical for SAL-induced DNA DSBs and apoptosis.