Under normoxia, 2-deoxy-d-glucose elicits cell death in select tumor types not by inhibition of glycolysis but by interfering with N-linked glycosylation

Abstract

In tumor cells growing under hypoxia, inhibiting glycolysis with 2-deoxy-D-glucose (2-DG) leads to cell death, whereas under normoxic conditions cells similarly treated survive. Surprisingly, here we find that 2-DG is toxic in select tumor cell lines growing under normal oxygen tension. In contrast, a more potent glycolytic inhibitor, 2-fluorodeoxy-D-glucose, shows little or no toxicity in these cell types, indicating that a mechanism other than inhibition of glycolysis is responsible for their sensitivity to 2-DG under normoxia. A clue to this other mechanism comes from previous studies in which it was shown that 2-DG interferes with viral N-linked glycosylation and is reversible by exogenous addition of mannose. Similarly, we find that 2-DG interferes with N-linked glycosylation more potently in the tumor cell types that are sensitive to 2-DG under normoxia, which can be reversed by exogenous mannose. Additionally, 2-DG induces an unfolded protein response, including up-regulation of GADD153 (C/ EBP-homologous protein), an unfolded protein responsespecific mediator of apoptosis, more effectively in 2-DGsensitive cells. We conclude that 2-DG seems to be toxic in select tumor cell types growing under normoxia by inhibition of N-linked glycosylation and not by glycolysis.

Inhibition of Akt Potentiates 2-DG–Induced Apoptosis via Downregulation of UPR in Acute Lymphoblastic Leukemia

“…It has been reported that in cancer phenotypes in which low-dose treatment with 2-DG (<4 mmol/L) induces cell death under normoxia, 2-DG acts through inhibition of Nlinked glycosylation (12,14). D-Mannose has been shown to reverse both the growth inhibitory and cytotoxic effects associated with inhibition of N-linked glycosylation but cannot reverse the cytotoxic effects triggered by inhibition of glycolysis (25,26).…”

“…2-DG can induce cell growth arrest and cell death by inhibiting 2 key glycolytic enzymes, hexokinase and phosphoglucose isomerase (PGI; ref. 12). Following facilitated diffusion into cells through glucose transporters, 2-DG is converted to 2-DG-6-phosphate by hexokinase, but unlike glucose-6-phosphate, 2-DG-6-phosphate cannot be further metabolized by PGI, and its accumulation inhibits the glycolytic pathway (13).…”

“…Inhibition of glycolysis has been shown to be a main mechanism of 2-DG's cytotoxicity in hypoxic cells of solid tumors (14). However, because of its structural similarity with mannose, 2-DG has also been shown to be a potent inhibitor of N-linked glycosylation (12,15). Indeed, 2-DG can be incorporated in place of mannose into lipid-linked oligosaccharide (LLO) chains (15), but unlike mannose, 2-DG cannot accommodate the required complex branching structure of these molecules, leading to premature termination of LLO synthesis, and accumulation of misfolded, nascent ER-synthesized proteins (16).…”

“…The 2-DG-induced accumulation of misfolded glycoproteins in the ER lumen leads to ER stress and induction of the unfolded protein response (UPR), a protective physiologic response aimed at reducing the presence of unfolded proteins and consequently ER stress (11,12). The UPR is mediated via 3 ER transmembrane receptors: protein kinase dsRNAlike ER kinase (PERK), activating transcription factor 6 (ATF6), and inositol-requiring enzyme 1a (IRE1a), which are activated upon dissociation from the main ER chaperone protein GRP78 to fully engage the UPR (17).…”

Sugar-free approaches to cancer cell killing

“…When proteins cannot be glycosylated, misfolded proteins accumulate, and the unfolded protein response is engaged. Indeed, ER stress, rather than energetic stress, may be the leading cause of cell death on treatment of some tumor cells with antiglycolytics such as 2-DG (Kurtoglu et al, 2007).…”