Prostate Cancer Radiosensitization through Poly(ADP-Ribose) Polymerase-1 Hyperactivation

Abstract

The clinical experimental agent, β-lapachone (β-lap; Arq 501), can act as a potent radiosensitizer in vitro through an unknown mechanism. In this study, we analyzed the mechanism to determine whether β-lap may warrant clinical evaluation as a radiosensitizer. β-Lap killed prostate cancer cells by NAD(P)H:quinone oxidoreductase 1 (NQO1) metabolic bioactivation, triggering a massive induction of reactive oxygen species, irreversible DNA single-strand breaks (SSB), poly(ADP-ribose) polymerase-1 (PARP-1) hyperactivation, NAD + /ATP depletion, and μ-calpain-induced programmed necrosis. In combination with ionizing radiation (IR), β-lap radiosensitized NQO1 + prostate cancer cells under conditions where nontoxic doses of either agent alone achieved threshold levels of SSBs required for hyperactivation of PARP-1. Combination therapy significantly elevated SSB level, γ-H2AX foci formation, and poly(ADP-ribosylation) of PARP-1, which were associated with ATP loss and induction of μ-calpain-induced programmed cell death. Radiosensitization by β-lap was blocked by the NQO1 inhibitor dicoumarol or the PARP-1 inhibitor DPQ. In a mouse xenograft model of prostate cancer, β-lap synergized with IR to promote antitumor efficacy. NQO1 levels were elevated in ∼60% of human prostate tumors evaluated relative to adjacent normal tissue, where β-lap might be efficacious alone or in combination with radiation. Our findings offer a rationale for the clinical utilization of β-lap (Arq 501) as a radiosensitizer in prostate cancers that overexpress NQO1, offering a potentially synergistic targeting strategy to exploit PARP-1 hyperactivation. Cancer Res; 70(20); 8088-96. ©2010 AACR.

PARP1-mediated necrosis is dependent on parallel JNK and Ca2+/calpain pathways

“…In contrast JNK2 knockdown considerably attenuated cell death in response to both compounds suggesting that it is this isoform that plays a causative role in PARP1-mediated necrotic death. -dependent protease calpain as key proximal signals in b-Lapachone-and MNNG-induced necrosis (Tagliarino et al, 2003;Moubarak et al, 2007;Dong et al, 2010). Consistent with this, co-treatment with the Ca 2+ chelating agent BAPTA-AM significantly attenuated the degree of necrotic cell death in response to b-Lapachone and MNNG (Fig.…”

“…It was originally thought that necrosis induced by PARP1 hyperactivation was simply due to metabolic catastrophe, where the overactive PARP1 used up the supply of NAD + in the cell, and subsequently ATP (van Wijk and Hageman, 2005). However, it is now appreciated that PARP1-induced necrosis follows more specific molecular pathways (Xu et al, 2006;Moubarak et al, 2007;Artus et al, 2010;Dong et al, 2010;Chiu et al, 2011;Park et al, 2011).…”

-Lapachone Induces NAD(P)H:Quinone Oxidoreductase-1- and Oxidative Stress-Dependent Heat Shock Protein 90 Cleavage and Inhibits Tumor Growth and Angiogenesis

“…We observed no significant body weight loss or other gross toxicity in the cohorts receiving b-Lap is bioactivated by NQO1, creating a "futile redox" cycle. By generating high levels of superoxide, b-lap causes programmed necrosis (called "necroptosis") in cancer cells (Bey et al, 2007;Dong et al, 2010). In this study, we showed for the first time that b-lap provokes the cleavage of Hsp90, leading to the degradation of various Hsp90 client oncoproteins in vitro and in vivo.…”

“…Previous studies demonstrated that b-lap causes NQO1-and Ca 21 -dependent m-calpain activation (Tagliarino et al, 2003;Dong et al, 2010). Calpain inhibitors were also reported to abolish the cleavage of Hsp90 (Bellocq et al, 1999).…”

“…First, the stimulation of b-lap is a rapid process that leads to more potent anticancer activity than the conventional Hsp90 inhibitors. In a heterogeneous panel of prostate cancer (Dong et al, 2010) and lung cancer cells that express high levels of endogenous NQO1, exposure to b-lap (6 mM) for 2 hours was sufficient to achieve maximal cytotoxicity. By contrast, a 24-hour treatment of 17-AAG (10 mM) was required to reduce cell viability by 50%-60% in various tumor cells (Plescia et al, 2005).…”