Assessing Oxidative Stress in Tumors by Measuring the Rate of Hyperpolarized [1-13C]Dehydroascorbic Acid Reduction Using 13C Magnetic Resonance Spectroscopy

Abstract

Rapid cancer cell proliferation promotes the production of reducing equivalents, which counteract the effects of relatively high levels of reactive oxygen species. Reactive oxygen species levels increase in response to chemotherapy and cell death, whereas an increase in antioxidant capacity can confer resistance to chemotherapy and is associated with an aggressive tumor phenotype. The pentose phosphate pathway is a major site of NADPH production in the cell, which is used to maintain the main intracellular antioxidant, glutathione, in its reduced state. Previous studies have shown that the rate of hyperpolarized [1-13C]dehydroascorbic acid (DHA) reduction, which can be measured in vivo using non-invasive 13C magnetic resonance spectroscopic imaging, is increased in tumors and that this is correlated with the levels of reduced glutathione. We show here that the rate of hyperpolarized [1-13C]DHA reduction is increased in tumors that have been oxidatively prestressed by depleting the glutathione pool by buthionine sulfoximine treatment. This increase was associated with a corresponding increase in pentose phosphate pathway flux, assessed using 13C-labeled glucose, and an increase in glutaredoxin activity, which catalyzes the glutathione-dependent reduction of DHA. These results show that the rate of DHA reduction depends not only on the level of reduced glutathione, but also on the rate of NADPH production, contradicting the conclusions of some previous studies. Hyperpolarized [1-13C]DHA can be used, therefore, to assess the capacity of tumor cells to resist oxidative stress in vivo. However, DHA administration resulted in transient respiratory arrest and cardiac depression, which may prevent translation to the clinic.

Hyperpolarized 13C magnetic resonance evaluation of renal ischemia reperfusion injury in a murine model

“…GSH is a cofactor of DHA reductase, which recycles DHA back to reduced ascorbic acid (VitC), therefore linking the redox couple between GSH and VitC. A recent study in a tumor model also showed that the rate of HP [1− 13 C]DHA reduction to VitC depends both on the GSH level and rate of NADPH production from the pentose phosphate pathway, and reflects the capacity of the tumor to resist oxidative stress . It should also be noted that DHA reduction can be affected by redox enzymes, such as glutaredoxin and glutathione‐ S ‐transferase omega, which show DHA reductase activity .…”

“…A recent study in a tumor model also showed that the rate of HP [1− 13 C]DHA reduction to VitC depends both on the GSH level and rate of NADPH production from the pentose phosphate pathway, and reflects the capacity of the tumor to resist oxidative stress. 20 It should also be noted that DHA reduction can be affected by redox enzymes, such as glutaredoxin and glutathione-S-transferase omega, which show DHA reductase activity. 33,34 Therefore, the rate of DHA reduction to VitC can be affected by multiple components of the complex redox machinery.…”

“…DHA is an oxidized form of vitamin C (VitC), and is reduced to VitC via a glutathione‐dependent mechanism, with coupled reactions to NADPH. Previous studies in several preclinical disease models have shown that the rate of HP [1− 13 C]DHA reduction to [1− 13 C]VitC is related to the levels of reduced glutathione (GSH) and to the rate of NADPH production from the pentose phosphate pathway . As GSH and NADPH are key antioxidants that counteract reactive oxygen species (ROS), HP [1− 13 C]DHA reduction to [1− 13 C]VitC should reflect tissue redox capacity and may serve as an indicator for cellular oxidative stress.…”

“…Previous studies in several preclinical disease models have shown that the rate of HP [1− 13 C]DHA reduction to [1− 13 C]VitC is related to the levels of reduced glutathione (GSH) and to the rate of NADPH production from the pentose phosphate pathway. [18][19][20] As GSH and NADPH are key antioxidants that counteract reactive oxygen species (ROS), HP [1− 13 C]DHA reduction to [1− 13 C]VitC should reflect tissue redox capacity and may serve as an indicator for cellular oxidative stress. Pyruvate (Pyr), a critical substrate for energy metabolism, is metabolized to downstream products, including lactate (Lac) and alanine (Ala), in the cytosol, and is also shuttled into the mitochondria and converted to acetyl-CoA via the enzyme PDH in the first step of oxidative phosphorylation.…”

Assessing the effect of hypoxia on cardiac metabolism using hyperpolarized 13C magnetic resonance spectroscopy

“…Hypoxia is also one of many pathological factors of tumor development, fluctuating over time and in regions of the tumor, and as such Iversen et al used HPC MRS in a mouse tumor model, showing that inspiration of a hypoxic atmosphere caused increased lactate production in tumors . Oxidative stress has been investigated in a few noncardiac studies, using HP dehydroascorbate, but the toxicity of this compound may limit translation to clinical studies . The challenges and future of hyperpolarized probes for assessing renal and cardiac oxygen metabolism have been discussed in a review by Schroeder and Laustsen; thus far, no studies have investigated the use of HP 13 C MRS to assess the effect of hypoxia on pyruvate metabolism in the in vivo heart.…”