Therapeutic Discovery Artesunate Induces Oxidative DNA Damage, Sustained DNA Double-Strand Breaks, and the ATM/ATR Damage Response in Cancer Cells

Artesunate, the active agent from Artemisia annua L. used in the traditional Chinese medicine, is being applied as afirst-line drug formalaria treatment, and trials are ongoing that include this drug in cancer therapy. Despite increasing interest in its therapeutic application, the mode of cell killing provoked by artesunate in human cells is unknown. Here, we show that artesunate is a powerful inducer of oxidative DNA damage, giving rise to formamidopyrimidineDNAglycosylase–sensitive sites and the formation of 8-oxoguanine and1, N6-ethenoadenine. Oxidative DNA damage was induced in LN-229 human glioblastoma cells dose depen-dently and was paralleled by cell death executed by apoptosis and necrosis, which could be attenuated by radical scavengers such as N-acetyl cysteine. Oxidative DNA damage resulted in DNA double-strand breaks (DSB) as determined by gH2AX foci that colocalized with 53BP1. Upon chronic treatment with artesunate, the level of DSB continuously increased over the treatment period up to a steady-state level, which is in contrast to ionizing radiation that induced a burst of DSB followed by a decline due to their repair. Knockdown of Rad51 by short interferingRNAand inactivation ofDNA-PKstrongly sensitizedglioma cells to artesunate. Thesedata indicate that both homologous recombination and nonhomologous end joining are involved in the repair of artesunate-inducedDSB.Artesunate provokedaDNAdamage response (DDR)withphosphorylation ofATM, ATR,Chk1, andChk2.Overall, thesedata revealed that artesunate induces oxidativeDNAlesions andDSB that continuously increase during the treatment period and accumulate until they trigger DDR and finally tumor cell death. Mol Cancer Ther; 10(12); 2224–33. 2011 AACR