Elsevier

Gynecologic Oncology

Volume 107, Issue 3, December 2007, Pages 450-457
Gynecologic Oncology

Resveratrol inhibits glucose metabolism in human ovarian cancer cells

https://doi.org/10.1016/j.ygyno.2007.07.065Get rights and content

Abstract

Objectives.

Resveratrol is a phytoalexin found in grapes that inhibits the in vitro growth of multiple tumor cell types. We showed previously that resveratrol induces autophagic cell death in ovarian cancer cells. Because autophagy is typically an adaptive response to nutrient starvation, we hypothesized that autophagy would also be triggered when ovarian cancer cells are nutrient deprived and that resveratrol could in fact be acting by inducing a starvation-like signaling response.

Methods.

Ovarian cancer cells were incubated with normal media, media containing resveratrol, glucose free media, or media lacking amino acids. Growth inhibition was determined using the sulforhodamine assay. Cells were evaluated for autophagocytosis by analyzing cleavage of LC3. Glucose uptake, lactate production, and activation of glycolytic regulators pAkt and pmTOR were analyzed following resveratrol treatment.

Results.

We show here that epithelial ovarian cancer cells are highly sensitive to glucose-deprivation-induced cell death and like resveratrol, glucose deprivation induces caspase-independent cell death with hallmarks of autophagy. Consistent with the hypothesis that resveratrol treatment results in biochemical conditions that mirror a nutrient deprived state, we found that resveratrol dramatically reduces glucose uptake and lactate production. Moreover, resveratrol reduces the levels of phosphorylated Akt and mTOR, two signals that increase glucose uptake and the rate limiting steps in glycolysis.

Conclusions.

Our findings are consistent with the hypothesis that resveratrol-induced changes in glucose utilization comprise the mechanism that underlies resveratrol-induced autophagocytosis in ovarian cancer. Inhibition of glycolysis in ovarian cancer with resveratrol or other compounds may be effective therapy for ovarian cancer.

Introduction

In the United States, epithelial ovarian carcinoma is the leading cause of death among patients with gynecological cancers. It is the eighth most frequent cause of cancer death in women, and 15,000 women die of this disease annually [1]. Despite improved survival for many malignancies during the recent era in clinical oncology, the 5-year relative survival rate for patients with advanced stage ovarian cancer remains low (30%) [1].

Although the majority of these tumors initially respond to chemotherapy, most patients develop recurrent tumors that are chemotherapy-resistant. Hence, resistance of recurrent disease, which has been linked to the failure of cytotoxic drugs to cause apoptotic cell death [2], is the principal factor limiting long-term treatment success against ovarian cancer. Tumor cells often develop defects in apoptosis, which provides a survival advantage that promotes tumorigenesis and contributes towards chemotherapy resistance in ovarian cancer [3], [4], [5]. Recent evidence indicates that tumor cells with acquired defects in apoptosis can be diverted to another cell death pathway, providing an alternative therapeutic approach. Nccrotic cell death can occur in apoptosis deficient cells as the result of metabolic catastrophe [6].

Many types of cancer, including ovarian cancer, have increased glycolytic activity compared to non-malignant cells [7]. Glycolysis is the series of biochemical reactions that convert glucose into 3-carbon carbohydrates and results in net ATP production. Increased glycolysis and increased glucose uptake may be adaptive responses to a hypoxic tumor environment [8] or be upregulated to meet increased demands for carbon substrates used in macromolecular synthesis in rapidly growing cells [9]. The high rate of glycolysis exhibited in tumors appears to be required to support tumor growth [10]. Ovarian cancer appears to be a cancer that has increased glucose metabolism. Compared to normal ovarian epithelial cells, ovarian cancer cells over-express the Glut1 glucose tranporter which brings glucose into cells a [11], [12]. Moreover, even small volume ovarian tumors are visualized using (18) F-fluorodexoyglucose positron emission tomography (FDG-PET) [13], a clinical marker of increased glucose uptake and glycolytic activity relative to normal tissues.

Not withstanding the current uncertainties about the basis for an increased glucose requirement by tumor cells, limiting glucose uptake or glucose metabolism is a promising experimental therapeutic strategy for cancer [6], [14]. Restricting caloric intake has long been recognized as an approach to reduce tumor occurrence and growth in animal models [15]. 2-Deoxyglucose (2-DG) is a glucose analog that inhibits glycolysis allowing it to mimic certain aspects of a glucose deprived, calorie restricted state. 2-DG is effective in several mouse tumor models, particularly when applied in combination with other agents, demonstrating that targeting carbohydrate metabolism may be an effective therapeutic approach [14].

Resveratrol (3,5,4′-trihydroxystilbene) has been shown to prevent or slow the progression of cancer, as well as extend the lifespans of various organisms ranging from yeast to vertebrates [16]. Resveratrol activates sirtuin deacetylases, and is considered a caloric restriction mimetic in lower organisms [17]. In mammals, resveratrol was found to downregulate glycolysis, a well known marker of caloric restriction, and overall, opposed the effects of high caloric intake in a mouse model [18]. Thus, it is possible that resveratrol may utilize the same pathways activated by caloric restriction in lower organisms and mammals [17].

In our previous studies, we investigated the effects of resveratrol on ovarian cancer cells. We found that it killed ovarian cancer cells through autophagocytosis. Autophagocytosis is cellular process that is typically associated with nutrient deprivation including conditions in which glucose is limited or completely withdrawn. If the nutrient supply is not restored, cells will undergo necrotic cell death. Having established that resveratrol treatment results in autophagy in ovarian cancer cells, we hypothesized depriving these cells of glucose would also trigger autophagy, and that resveratrol-induced autophagy might in fact be mediated by resveratrol-induced changes in glucose uptake and utilization pathways. We present results that are consistent with both hypotheses and discuss these results in relation to proposed therapeutic strategies based on blocking glucose metabolism.

Section snippets

Chemicals

Resveratrol was generously provided by Royalmount Pharma (Montreal, Quebec) and was dissolved in aqueous DMSO at a final concentration of 0.1% (v/v). Phloretin and cytochalasin B was obtained from Sigma-Aldrich Chemical Co. (St. Louis, MO). z-VAD-fmk was obtained from Enyzme Systems Products (Livermore, CA). 3[H]-2-DG was obtained from Perkin Elmer Life Sciences (Boston, MA).

Cell lines

WI-38 human fibroblasts, SKOV3 and CaOV3 cells were obtained from the American Type Culture collection (Manassas, VA).

Glucose deprivation has selective effects against ovarian cancer cell growth

Because ovarian cancer cells over-express the Glut1 glucose transporter [11], [12], and are detectable using FDG-PET [13], a clinical marker of increased glucose uptake, we hypothesized that ovarian cancer cells have particular glucose requirements that would make these tumor cells highly sensitive to glucose withdrawal. To demonstrate the glucose dependence of ovarian cancer cells, we measured the growth in culture of 5 different ovarian cancer cell lines (A2780, OVCA432, OVCA429, CaOV3, and

Discussion

Warburg was the first to report that compared to normal cells, tumor cells exhibit elevated glycolytic activity to support metabolic requirements. In contrast with normal cells, tumor cells rely on glycolysis rather than oxidative phosphorylation for ATP production, even under aerobic conditions (Warburg effect) [27]. The biochemical and molecular mechanisms that contribute towards increased aerobic glycolysis in cancer cells is complex. Several factors that are thought to alter energy

Acknowledgments

This work was supported by the Ovarian Cancer Research Fund, the Gynecologic Cancer Foundation, and the Susan G. Komen Breast Cancer Foundation (JRL), and received the Donald F. Richardson Memorial Prize Paper Award, 2005 (AK).

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