Abstract
Commonly used antitumor agents, such as DNA topoisomerase I/II poisons, kill cancer cells by creating nonrepairable DNA double-strand breaks (DSBs). To repair DSBs, error-free homologous recombination (HR), and/or error-prone nonhomologous end joining (NHEJ) are activated. These processes involve the phosphatidylinositol 3'-kinase-related kinase family of serine/threonine enzymes: ataxia telangiectasia mutated (ATM), ATM- and Rad3-related for HR, and DNA-dependent protein kinase catalytic subunit (DNA-PKcs) for NHEJ. Alterations in these repair processes can cause drug/radiation resistance and increased genomic instability. beta-Lapachone (beta-lap; also known as ARQ 501), currently in phase II clinical trials for the treatment of pancreatic cancer, causes a novel caspase- and p53-independent cell death in cancer cells overexpressing NAD(P)H:quinone oxidoreductase-1 (NQO1). NQO1 catalyzes a futile oxidoreduction of beta-lap leading to reactive oxygen species generation, DNA breaks, gamma-H2AX foci formation, and hyperactivation of poly(ADP-ribose) polymerase-1, which is required for cell death. Here, we report that beta-lap exposure results in NQO1-dependent activation of the MRE11-Rad50-Nbs-1 complex. In addition, ATM serine 1981, DNA-PKcs threonine 2609, and Chk1 serine 345 phosphorylation were noted; indicative of simultaneous HR and NHEJ activation. However, inhibition of NHEJ, but not HR, by genetic or chemical means potentiated beta-lap lethality. These studies give insight into the mechanism by which beta-lap radiosensitizes cancer cells and suggest that NHEJ is a potent target for enhancing the therapeutic efficacy of beta-lap alone or in combination with other agents in cancer cells that express elevated NQO1 levels.
Highlights
Many cancer chemotherapeutic agents, such as ionizing radiation (IR), and DNA-damaging chemotherapeutic compounds cause cell death by creating DNA double-strand breaks (DSBs; refs. 1, 2)
We recently showed that h-lap–induced cell death was dependent on poly(ADP-ribose) polymerase-1 (PARP-1) hyperactivation, but not on typical apoptotic mediators, such as p53, Bax/Bak or caspases [17]
We recently showed that h-lap–induced cell death was initiated by the NQO1dependent generation of reactive oxygen species (ROS), subsequent formation of DNA damage, and calcium-dependent PARP-1 hyperactivation
Summary
Many cancer chemotherapeutic agents, such as ionizing radiation (IR), and DNA-damaging chemotherapeutic compounds cause cell death by creating DNA double-strand breaks (DSBs; refs. 1, 2). Homologous recombination (HR) and nonhomologous end joining (NHEJ) are two distinct, yet complementary, mechanisms for mammalian DSB repair that can interact simultaneously at DSB sites [5,6,7]. Essential to both HR and NHEJ is the activation of one or all three related phosphatidylinositol 3¶-kinase–like kinases (PI3K) in response to DNA damage [7]. After DSB detection and PI3K activation, H2AX becomes phosphorylated on serine 139 (g-H2AX) in a 2-Mb region surrounding the break. In HR, the complex acts as an exonuclease to produce 3¶ single-strand overhangs bound by Rad52 [12]
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