Abstract
Simple SummaryCancer is often characterized by profound changes in metabolism, some of which are appropriate targets for therapeutic intervention. Many cancers overexpress the two-electron reductase NQO1, which can bioactivate the drug β-lapachone, inducing a futile redox cycle that liberates large amounts of reactive oxygen species and results in subsequent cell death. However, β-lapachone has off-target toxicities in red blood cells, which makes minimal dosing for chemotherapeutic response desirable. Here, we show that magnetic resonance-based detection of [2H7]glucose metabolism provides a robust metric of NQO1 activation, as the redox perturbation causes downregulation of glycolytic flux that is detectable in the HDO and lactate signals. Imaging of either metabolic product could provide constraints for a continual reassessment model for controlling therapeutic dosing levels.Treatment of cancers with β-lapachone causes NAD(P)H: quinone oxidoreductase 1 (NQO1) to generate an unstable hydroquinone that regenerates itself in a futile cycle while producing reactive oxygen species (ROS) in the form of superoxide and subsequently hydrogen peroxide. Rapid accumulation of ROS damages DNA, hyperactivates poly-ADP-ribose polymerase-I, causes massive depletion of NAD+/ATP, and hampers glycolysis. Cells overexpressing NQO1 subsequently die rapidly through an NAD+-keresis mechanism. Assessing changes in glycolytic rates caused by NQO1 bioactivation would provide a means of assessing treatment efficacy, potentially lowering the chemotherapeutic dosage, and reducing off-target toxicities. NQO1-mediated changes in glycolytic flux were readily detected in A549 (lung), MiaPaCa2 (pancreatic), and HCT-116 (colon) cancer cell lines by 2H-NMR after administration of [2H7]glucose. The deuterated metabolic products 2H-lactate and HDO were quantified, and linear relationships with glucose consumption for both products were observed. The higher concentration of HDO compared to 2H-lactate allows for more sensitive measurement of the glycolytic flux in cancer. Gas chromatography-mass spectrometry analysis agreed with the NMR results and confirmed downregulated energy metabolism in NQO1+ cells after β-lapachone treatment. The demonstrated method is ideal for measuring glycolytic rates, the effects of chemotherapeutics that target glycolysis, and has the potential for in vivo translation.
Highlights
IntroductionMultiple cancers display an increased glucose uptake under normoxic conditions and favor lactate production as opposed to aerobic respiration (i.e., the Warburg effect) [3]
This process causes the rapid accumulation of reactive oxygen species (ROS), leading to DNA damage, followed by hyperactivation of poly(ADP-ribose) polymerase-1 (PARP1), leading to NAD+ and ATP loss, and cell death by a mechanism known as NAD+ -keresis [12]. β-lapachone-induced cytotoxicity directly involves the inhibition of the NAD+ -dependent glyceraldehyde 3-phosphate dehydrogenase (GAPDH), which suppresses overall glycolytic flux [6,12]
We report that HDO and 2 H-lactate production can be used as a sensitive marker to measure differences in glycolytic metabolism caused by β-lapachone drug action
Summary
Multiple cancers display an increased glucose uptake under normoxic conditions and favor lactate production as opposed to aerobic respiration (i.e., the Warburg effect) [3] These metabolic defects have been studied extensively as a way to differentiate cancer cells from healthy cells, both for diagnosis and treatment purposes [4,5]. Β-lapachone treatment takes advantage of the NQO1 catalyzed reaction through the formation of an unstable hydroquinone that regenerates itself while producing peroxides in a futile cycle [11]. This process causes the rapid accumulation of reactive oxygen species (ROS), leading to DNA damage, followed by hyperactivation of poly(ADP-ribose) polymerase-1 (PARP1), leading to NAD+ and ATP loss, and cell death by a mechanism known as NAD+ -keresis [12]. While β-lapachone has shown significant promise in Phase-1 clinical trials, off-target, dose-dependent toxicity in red blood cells due to methemoglobinemia has caused significant safety concerns [13]
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