NAD(P)H:Quinone Oxidoreductase Activity Is the Principal Determinant of β-Lapachone Cytotoxicity

Abstract

␤-Lapachone activates a novel apoptotic response in a number of cell lines. We demonstrate that the enzyme NAD(P)H:quinone oxidoreductase (NQO1) substantially enhances the toxicity of ␤-lapachone. NQO1 expression directly correlated with sensitivity to a 4-h pulse of ␤-lapachone in a panel of breast cancer cell lines, and the NQO1 inhibitor, dicoumarol, significantly protected NQO1-expressing cells from all aspects of ␤-lapachone toxicity. Stable transfection of the NQO1-deficient cell line, MDA-MB-468, with an NQO1 expression plasmid increased apoptotic responses and lethality after ␤-lapachone exposure. Dicoumarol blocked both the apoptotic responses and lethality. Biochemical studies suggest that reduction of ␤-lapachone by NQO1 leads to a futile cycling between the quinone and hydroquinone forms, with a concomitant loss of reduced NAD(P)H. In addition, the activation of a cysteine protease, which has characteristics consistent with the neutral calciumdependent protease, calpain, is observed after ␤-lapachone treatment. This is the first definitive elucidation of an intracellular target for ␤-lapachone in tumor cells. NQO1 could be exploited for gene therapy, radiotherapy, and/or chemopreventive interventions, since the enzyme is elevated in a number of tumor types (i.e. breast and lung) and during neoplastic transformation.

Beta-lapachone inhibits proliferation and induces apoptosis in retinoblastoma cell lines

“…[31][32][33] Other studies in prostate and breast cancer cells have reported that beta-lapachone-induced apoptosis can occur through a mechanism involving NQOI-mediated cycling of this agent. 27,34,35 NQOI is an enzyme that reduces quinone-containing agents such as beta-lapachone to more reactive forms. NQO1-deficient prostate cancer cells, and prostate and breast cancer cells treated with dicoumarol (an NQO1 inhibitor) are significantly more resistant to apoptosis than NQO1-expressing prostate cells after beta-lapachone exposure.…”

“…NQO1-deficient prostate cancer cells, and prostate and breast cancer cells treated with dicoumarol (an NQO1 inhibitor) are significantly more resistant to apoptosis than NQO1-expressing prostate cells after beta-lapachone exposure. 27,34 Furthermore, transfection of NQO1-deficient cells with an NQO1 expression plasmid markedly enhanced sensitivity to beta-lapachone. 27,34 However, blocking of NQO1 in various cancer cell types does not completely abrogate apoptosis, and although NQO1 plays a major role in beta-lapachoneinduced apoptosis, alternate pathways also appear to be involved.…”

“…27,34 Furthermore, transfection of NQO1-deficient cells with an NQO1 expression plasmid markedly enhanced sensitivity to beta-lapachone. 27,34 However, blocking of NQO1 in various cancer cell types does not completely abrogate apoptosis, and although NQO1 plays a major role in beta-lapachoneinduced apoptosis, alternate pathways also appear to be involved. 27,34,35 Although our experiments were limited to evaluating the cytotoxic effects of beta-lapachone alone in RB cells, a number of studies have found significant synergistic effects of beta-lapachone with other chemotherapeutic agents.…”

“…17,18,23 Beta-lapachone cytotoxicity may also be mediated by the activity of the NAD(P)H:quinone oxidoreductase enzyme NQO1. 27 This agent has also been shown to selectively induce apoptosis in breast and prostate cancer cells but not in untransformed cells through a mechanism that appears to involve selective induction of the transcription factor E2F1, a regulator of both S-phase progression and apoptosis. 24 To our knowledge, the effects of beta-lapachone in RB cell lines have not yet been investigated.…”

Nuclear ULK1 promotes cell death in response to oxidative stress through PARP1

“…The orthonapthoquinone β-lapachone inhibits the catalytic activity of DNA topoisomerase 1, thereby causing PARP1 hyperactivation. [43][44][45] The efficacy of β-lapachone as an anticancer agent is being evaluated in multiple Phase II trials (http://www.cancer.gov/clinicaltrials). The AMPK-resistant 4SA mutation sensitized cells to a sublethal dose of β-lapachone to the same extent as WT ULK1 did.…”