Transcriptome analysis reveals organ-specific effects of 2-deoxyglucose treatment in healthy mice.

OBJECTIVE: Glycolytic inhibition via 2-deoxy-D-glucose (2DG) has potential therapeutic benefits for a range of diseases, including cancer, epilepsy, systemic lupus erythematosus (SLE), and rheumatoid arthritis (RA), and COVID-19, but the systemic effects of 2DG on gene function across different tissues are unclear. METHODS: This study analyzed the transcriptional profiles of nine tissues from C57BL/6J mice treated with 2DG to understand how it modulates pathways systemically. Principal component analysis (PCA), weighted gene co-network analysis (WGCNA), analysis of variance, and pathway analysis were all performed to identify modules altered by 2DG treatment. RESULTS: PCA revealed that samples clustered predominantly by tissue, suggesting that 2DG affects each tissue uniquely. Unsupervised clustering and WGCNA revealed six distinct tissue-specific modules significantly affected by 2DG, each with unique key pathways and genes. 2DG predominantly affected mitochondrial metabolism in the heart, while in the small intestine, it affected immunological pathways. CONCLUSIONS: These findings suggest that 2DG has a systemic impact that varies across organs, potentially affecting multiple pathways and functions. The study provides insights into the potential therapeutic benefits of 2DG across different diseases and highlights the importance of understanding its systemic effects for future research and clinical applications.

Lesion of NPY Receptor-expressing Neurons in Perifornical Lateral Hypothalamus Attenuates Glucoprivic Feeding.

Glucoprivic feeding is one of several counterregulatory responses (CRRs) that facilitates restoration of euglycemia following acute glucose deficit (glucoprivation). Our previous work established that glucoprivic feeding requires ventrolateral medullary (VLM) catecholamine (CA) neurons that coexpress neuropeptide Y (NPY). However, the connections by which VLM CA/NPY neurons trigger increased feeding are uncertain. We have previously shown that glucoprivation, induced by an anti-glycolygic agent 2-deoxy-D-glucose (2DG), activates perifornical lateral hypothalamus (PeFLH) neurons and that expression of NPY in the VLM CA/NPY neurons is required for glucoprivic feeding. We therefore hypothesized that glucoprivic feeding and possibly other CRRs require NPY-sensitive PeFLH neurons. To test this, we used the ribosomal toxin conjugate NPY-saporin (NPY-SAP) to selectively lesion NPY receptor-expressing neurons in the PeFLH of male rats. We found that NPY-SAP destroyed a significant number of PeFLH neurons, including those expressing orexin, but not those expressing melanin-concentrating hormone. The PeFLH NPY-SAP lesions attenuated 2DG-induced feeding but did not affect 2DG-induced increase in locomotor activity, sympathoadrenal hyperglycemia, or corticosterone release. The 2DG-induced feeding response was also significantly attenuated in NPY-SAP-treated female rats. Interestingly, PeFLH NPY-SAP lesioned male rats had reduced body weights and decreased dark cycle feeding, but this effect was not seen in female rats. We conclude that a NPY projection to the PeFLH is necessary for glucoprivic feeding, but not locomotor activity, hyperglycemia, or corticosterone release, in both male and female rats.

2-Deoxy-D-glucose ameliorates inflammation and fibrosis in a silicosis mouse model by inhibiting hypoxia-inducible factor-1α in alveolar macrophages.

Inhaling silica causes the occupational illness silicosis, which mostly results in the gradual fibrosis of lung tissue. Previous research has demonstrated that hypoxia-inducible factor-1α (HIF-1α) and glycolysis-related genes are up-regulated in silicosis. The role of 2-deoxy-D-glucose (2-DG) as an inhibitor of glycolysis in silicosis mouse models and its molecular mechanisms remain unclear. Therefore, we used 2-DG to observe its effect on pulmonary inflammation and fibrosis in a silicosis mouse model. Furthermore, in vitro cell experiments were conducted to explore the specific mechanisms of HIF-1α. Our study found that 2-DG down-regulated HIF-1α levels in alveolar macrophages induced by silica exposure and reduced the interleukin-1ß (IL-1ß) level in pulmonary inflammation. Additionally, 2-DG reduced silica-induced pulmonary fibrosis. From these findings, we hypothesize that 2-DG reduced glucose transporter 1 (GLUT1) expression by inhibiting glycolysis, which inhibits the expression of HIF-1α and ultimately reduces transcription of the inflammatory cytokine, IL-1ß, thus alleviating lung damage. Therefore, we elucidated the important regulatory role of HIF-1α in an experimental silicosis model and the potential defense mechanisms of 2-DG. These results provide a possible effective strategy for 2-DG in the treatment of silicosis.

Establishment of bone-targeted nano-platform and the study of its combination with 2-deoxy-d-glucose enhanced photodynamic therapy to inhibit bone metastasis.

At present, simple anti-tumor drugs are ineffective at targeting bone tissue and are not purposed to treat patients with bone metastasis. In this study, zoledronic acid (ZOL) demonstrated excellent bone-targeting properties as a bone-targeting ligand. The metal-organic framework (MOF) known as ZIF-90 was modified with ZOL to construct a bone-targeting-based drug delivery system. Chlorin e6 (Ce6) was loaded in the bone-targeted drug delivery system and combined with 2-deoxy-D-glucose (2-DG), which successfully treated bone tumors when enhanced photodynamic therapy was applied. The Ce6@ZIF-PEG-ZOL (Ce6@ZPZ) nanoparticles were observed to have uniform morphology, a particle size of approximately 210 nm, and a potential of approximately -30.4 mV. The results of the bone-targeting experiments showed that Ce6@ZPZ exhibited a superior bone-targeted effect when compared to Ce6@ZIF-90-PEG. The Ce6@ZPZ solution was subjected to 660 nm irradiation and the resulting production of reactive oxygen species increased over time, which could be further increased when Ce6@ZPZ was used in combination with 2-DG. Their combination had a stronger inhibitory capacity against tumor cells than either 2-DG or Ce6@ZPZ alone, increasing the rate of tumor cell apoptosis. The apoptosis rate caused by HGC-27 was 61.56% when 2-DG was combined with Ce6@ZPZ. In vivo results also showed that Ce6@ZPZ combined with 2-DG maximally inhibited tumor growth and prolonged mice survival compared to the other experimental groups. Therefore, the combination of PDT and glycolytic inhibitors serves as a potential option for the treatment of cancer.

2-Deoxy-glucose ameliorates the peritoneal mesothelial and endothelial barrier function perturbation occurring due to Peritoneal Dialysis fluids exposure.

Peritoneal dialysis (PD) and prolonged exposure to PD fluids (PDF) induce peritoneal membrane (PM) fibrosis and hypervascularity, leading to functional PM degeneration. 2-deoxy-glucose (2-DG) has shown potential as PM antifibrotic by inhibiting hyper-glycolysis induced mesothelial-to-mesenchymal transition (MMT). We investigated whether administration of 2-DG with several PDF affects the permeability of mesothelial and endothelial barrier of the PM. The antifibrotic effect of 2-DG was confirmed by the gel contraction assay with embedded mesothelial (MeT-5A) or endothelial (EA.hy926) cells cultured in Dianeal® 2.5 % (CPDF), BicaVera® 2.3 % (BPDF), Balance® 2.3 % (LPDF) with/without 2-DG addition (0.2 mM), and qPCR for αSMA, CDH2 genes. Moreover, 2-DG effect was tested on the permeability of monolayers of mesothelial and endothelial cells by monitoring the transmembrane resistance (RTM), FITC-dextran (10, 70 kDa) diffusion and mRNA expression levels of CLDN-1 to -5, ZO1, SGLT1, and SGLT2 genes. Contractility of MeT-5A cells in CPDF/2-DG was decreased, accompanied by αSMA (0.17 ± 0.03) and CDH2 (2.92 ± 0.29) gene expression fold changes. Changes in αSMA, CDH2 were found in EA.hy926 cells, though αSMA also decreased under LPDF/2-DG incubation (0.42 ± 0.02). Overall, 2-DG mitigated the PDF-induced alterations in mesothelial and endothelial barrier function as shown by RTM, dextran transport and expression levels of the CLDN-1 to -5, ZO1, and SGLT2. Thus, supplementation of PDF with 2-DG not only reduces MMT but also improves functional permeability characteristics of the PM mesothelial and endothelial barrier.

The Combined Metabolically Oriented Effect of Fucoidan from the Brown Alga Saccharina cichorioides and Its Carboxymethylated Derivative with 2-Deoxy-D-Glucose on Human Melanoma Cells.

Melanoma is the most aggressive and treatment-resistant form of skin cancer. It is phenotypically characterized by aerobic glycolysis that provides higher proliferative rates and resistance to cell death. The glycolysis regulation in melanoma cells by means of effective metabolic modifiers represents a promising therapeutic opportunity. This work aimed to assess the metabolically oriented effect and mechanism of action of fucoidan from the brown alga Saccharina cichorioides (ScF) and its carboxymethylated derivative (ScFCM) in combination with 2-deoxy-D-glucose (2-DG) on the proliferation and colony formation of human melanoma cell lines SK-MEL-28, SK-MEL-5, and RPMI-7951. The metabolic profile of melanoma cells was determined by the glucose uptake and Lactate-GloTM assays. The effect of 2-DG, ScF, ScFCM, and their combination on the proliferation, colony formation, and activity of glycolytic enzymes was assessed by the MTS, soft agar, and Western blot methods, respectively. When applied separately, 2-DG (IC50 at 72 h = 8.7 mM), ScF (IC50 at 72 h > 800 µg/mL), and ScFCM (IC50 at 72 h = 573.9 µg/mL) inhibited the proliferation and colony formation of SK-MEL-28 cells to varying degrees. ScF or ScFCM enhanced the inhibiting effect of 2-DG at low, non-toxic concentrations via the downregulation of Glut 1, Hexokinase II, PKM2, LDHA, and pyruvate dehydrogenase activities. The obtained results emphasize the potential of the use of 2-DG in combination with algal fucoidan or its derivative as metabolic modifiers for inhibition of melanoma SK-MEL-28 cell proliferation.

Chemogenetic activation of ventral medullary astrocytes enhances feeding and corticosterone release in response to mild glucoprivation.

To investigate the role of glial cells in the regulation of glucoprivic responses in rats, a chemogenetic approach was used to activate astrocytes neighboring catecholamine (CA) neurons in the ventromedial medulla (VLM) where A1 and C1 CA cell groups overlap (A1/C1). Previous results indicate that activation of CA neurons in this region is necessary and sufficient for feeding and corticosterone release in response to glucoprivation. However, it is not known whether astrocyte neighbors of CA neurons contribute to glucoregulatory responses. Hence, we made nanoinjections of AAV5-GFAP-hM3D(Gq)-mCherry to selectively transfect astrocytes in the A1/C1 region with the excitatory designer receptor exclusively activated by designer drugs (DREADDs), hM3D(Gq). After allowing time for DREADD expression, we evaluated the rats for increased food intake and corticosterone release in response to low systemic doses of the antiglycolytic agent, 2-deoxy-d-glucose (2DG), alone and in combination with the hM3D(Gq) activator clozapine-n-oxide (CNO). We found that DREADD-transfected rats ate significantly more food when 2DG and CNO were coadministered than when either 2DG or CNO was injected alone. We also found that CNO significantly enhanced 2DG-induced FOS expression in the A1/C1 CA neurons, and that corticosterone release also was enhanced when CNO and 2DG were administered together. Importantly, CNO-induced activation of astrocytes in the absence of 2DG did not trigger food intake or corticosterone release. Our results indicate that during glucoprivation, activation of VLM astrocytes cells markedly increases the sensitivity or responsiveness of neighboring A1/C1 CA neurons to glucose deficit, suggesting a potentially important role for VLM astrocytes in glucoregulation.

Effect of 2-deoxyglucose-mediated inhibition of glycolysis on migration and invasion of HTR-8/SVneo trophoblast cells.

The proper invasion of trophoblasts is crucial for embryo implantation and placental development, which is helpful to establish a correct maternal-fetal relationship. Trophoblasts can produce a large amount of lactate through aerobic glycolysis during early pregnancy. Lactate creates a low pH microenvironment around the embryo to help uterine tissue decompose and promote the invasion of trophoblasts. The purpose of this study is to reveal the the potential mechanism of aerobic glycolysis regulating the invasiveness of trophoblasts by investigating the effect of 2-Deoxy-D-glucose (2-DG), a glycolysis inhibitor, on the biological function of HTR-8/SVneo trophoblast cells, the expressions of epithelial mesenchymal transformation (EMT) markers and invasion-related factors. 2-DG could inhibit the aerobic glycolysis of trophoblasts and decrease the activity of trophoblasts in a dose-dependent manner. Moreover, 2-DG inhibited the EMT of HTR-8/SVneo cells, down-regulated the expression of invasion-related factors matrix metalloproteinase 2/9 (MMP2/9) and up-regulated the expression of tissue inhibitor of matrix metalloproteinases 1/2 (TIMP1/2), thus inhibiting cell migration and invasion. This paper provides a foundation in the significance of aerobic glycolysis of trophoblasts in the process of invasion, and also provides ideas and insights for the promotion of embryo implantation.

2-Deoxyglucose drives plasticity via an adaptive ER stress-ATF4 pathway and elicits stroke recovery and Alzheimer's resilience.

Intermittent fasting (IF) is a diet with salutary effects on cognitive aging, Alzheimer's disease (AD), and stroke. IF restricts a number of nutrient components, including glucose. 2-deoxyglucose (2-DG), a glucose analog, can be used to mimic glucose restriction. 2-DG induced transcription of the pro-plasticity factor, Bdnf, in the brain without ketosis. Accordingly, 2-DG enhanced memory in an AD model (5xFAD) and functional recovery in an ischemic stroke model. 2-DG increased Bdnf transcription via reduced N-linked glycosylation, consequent ER stress, and activity of ATF4 at an enhancer of the Bdnf gene, as well as other regulatory regions of plasticity/regeneration (e.g., Creb5, Cdc42bpa, Ppp3cc, and Atf3) genes. These findings demonstrate an unrecognized role for N-linked glycosylation as an adaptive sensor to reduced glucose availability. They further demonstrate that ER stress induced by 2-DG can, in the absence of ketosis, lead to the transcription of genes involved in plasticity and cognitive resilience as well as proteostasis.

2-Deoxy-D-glucose administration after seizures has disease-modifying effects on kindling progression.

2-deoxy-D-glucose (2DG) is a glucose analog and reversible inhibitor of glycolysis with anticonvulsant and antiepileptic effects in multiple seizure models. 2DG at a dose of 250 mg/kg intraperitoneally (IP) delays progression of repeated seizures evoked by kindling in rats when administered 30 min prior to twice daily kindling stimulation. As toxicological studies have demonstrated that repeated daily oral administration of 2DG at doses of 60-375 mg/kg/day in rats induces dose-dependent, reversible cardiac myocyte vacuolation, it was of interest to determine if 2DG also slowed kindling progression when administered at or below doses causing cardiac toxicity and at various time points after evoked seizures. We found that: (1) 2DG slowed kindling progression nearly 2-fold when administered at a dose of 37.5 mg/kg given IP 30 min prior to kindling stimulation, and (2) 2DG 37.5 mg/kg IP also slowed kindling progression when given immediately after, and for as long as 10 min after evoked (kindled) seizures. These observations suggest potential clinical usefulness of post-seizure administration of 2DG to reduce seizure clusters and long-term consequences of repeated seizures at human equivalent doses that are likely to be safe and well tolerated in patients.

Induction of glioblastoma cell ferroptosis using combined treatment with chloramphenicol and 2-deoxy-D-glucose.

Glioblastoma, a malignant tumor, has no curative treatment. Recently, mitochondria have been considered a potential target for treating glioblastoma. Previously, we reported that agents initiating mitochondrial dysfunction were effective under glucose-starved conditions. Therefore, this study aimed to develop a mitochondria-targeted treatment to achieve normal glucose conditions. This study used U87MG (U87), U373, and patient-derived stem-like cells as well as chloramphenicol (CAP) and 2-deoxy-D-glucose (2-DG). We investigated whether CAP and 2-DG inhibited the growth of cells under normal and high glucose concentrations. In U87 cells, 2-DG and long-term CAP administration were more effective under normal glucose than high-glucose conditions. In addition, combined CAP and 2-DG treatment was significantly effective under normal glucose concentration in both normal oxygen and hypoxic conditions; this was validated in U373 and patient-derived stem-like cells. 2-DG and CAP acted by influencing iron dynamics; however, deferoxamine inhibited the efficacy of these agents. Thus, ferroptosis could be the underlying mechanism through which 2-DG and CAP act. In conclusion, combined treatment of CAP and 2-DG drastically inhibits cell growth of glioblastoma cell lines even under normal glucose conditions; therefore, this treatment could be effective for glioblastoma patients.

DNA Aptamer Beacon Probe (ABP) for Monitoring of Adenosine Triphosphate Level in SW480 Cancer Cells Treated with Glycolysis Inhibitor 2-Deoxyglucose.

Early cancer screening enables timely detection of carcinogenesis, and aids in prompt clinical intervention. Herein, we report on the development of a simple, sensitive, and rapid fluorometric assay based on the aptamer probe (aptamer beacon probe, ABP) for monitoring the energy-demand biomarker adenosine triphosphate (ATP), an essential energy source that is released into the tumor microenvironment. Its level plays a significant role in risk assessment of malignancies. The operation of the ABP for ATP was examined using solutions of ATP and other nucleotides (UTP, GTP, CTP), followed by monitoring of ATP production in SW480 cancer cells. Then, the effect of a glycolysis inhibitor, 2-deoxyglucose (2-DG), on SW480 cells was investigated. The stability of predominant ABP conformations in the temperature range of 23-91 °C and the effects of temperature on ABP interactions with ATP, UTP, GTP, and CTP were evaluated based on quenching efficiencies (QE) and Stern-Volmer constants (KSV). The optimized temperature for best selectivity of ABP toward ATP was 40 °C (KSV = 1093 M-1, QE = 42%). We have found that the inhibition of glycolysis in SW480 cancer cells by 2-deoxyglucose resulted in lowering of ATP production by 31.7%. Therefore, monitoring and modulation of ATP concentration may aid in future cancer treatment.

Multitranscript analysis reveals an effect of 2-deoxy-d-glucose on gene expression linked to unfolded protein response and integrated stress response in primary human monocytes and monocyte-derived macrophages.

BACKGROUND: Glycolytic inhibitor 2-deoxy-d-glucose (2-DG) binds to hexokinase in a non-competitive manner and phosphoglucose isomerase in a competitive manner, blocking the initial steps of the glycolytic pathway. Although 2-DG stimulates endoplasmic reticulum (ER) stress, activating the unfolded protein response to restore protein homeostasis, it is unclear which ER stress-related genes are modulated in response to 2-DG treatment in human primary cells. Here, we aimed to determine whether the treatment of monocytes and monocyte-derived macrophages (MDMs) with 2-DG leads to a transcriptional profile specific to ER stress. METHODS: We performed bioinformatics analysis to identify differentially expressed genes (DEGs) in previously reported RNA-seq datasets of 2-DG treated cells. RT-qPCR was performed to verify the sequencing data on cultured MDMs. RESULTS: A total of 95 common DEGs were found by transcriptional analysis of monocytes and MDMs treated with 2-DG. Among these, 74 were up-regulated and 21 were down-regulated. Multitranscript analysis showed that DEGs are linked to integrated stress response (GRP78/BiP, PERK, ATF4, CHOP, GADD34, IRE1α, XBP1, SESN2, ASNS, PHGDH), hexosamine biosynthetic pathway (GFAT1, GNA1, PGM3, UAP1), and mannose metabolism (GMPPA and GMPPB). CONCLUSIONS: Results reveal that 2-DG triggers a gene expression program that might be involved in restoring protein homeostasis in primary cells. GENERAL SIGNIFICANCE: 2-DG is known to inhibit glycolysis and induce ER stress; however, its effect on gene expression in primary cells is not well understood. This work shows that 2-DG is a stress inducer shifting the metabolic state of monocytes and macrophages.

Metabolic heterogeneity in TNBCs: A potential determinant of therapeutic efficacy of 2-deoxyglucose and metformin combinatory therapy.

Breast cancers (BCs) remain the leading cause of cancer-related deaths among women worldwide. Among the different types of BCs, treating the highly aggressive, invasive, and metastatic triple-negative BCs (TNBCs) that do not respond to hormonal/human epidermal growth factor receptor 2 (HER2) targeted interventions since they lack ER/PR/HER2 receptors remains challenging. While almost all BCs depend on glucose metabolism for their proliferation and survival, studies indicate that TNBCs are highly dependent on glucose metabolism compared to non-TNBC malignancies. Hence, limiting/inhibiting glucose metabolism in TNBCs should curb cell proliferation and tumor growth. Previous reports, including ours, have shown the efficacy of metformin, the most widely prescribed antidiabetic drug, in reducing cell proliferation and growth in MDA-MB-231 and MDA-MB-468 TNBC cells. In the current study, we investigated and compared the anticancer effects of either metformin (2 mM) in glucose-starved or 2-deoxyglucose (10 mM; glycolytic inhibitor; 2DG) exposed MDA-MB-231 and MDA-MB-468 TNBC cells. Assays for cell proliferation, rate of glycolysis, cell viability, and cell-cycle analysis were performed. The status of proteins of the mTOR pathway was assessed by Western blot analysis. Metformin treatment in glucose-starved and 2DG (10 mM) exposed TNBC cells inhibited the mTOR pathway compared to non-treated glucose-starved cells or 2DG/metformin alone treated controls. Cell proliferation is also significantly reduced under these combination treatment conditions. The results indicate that combining a glycolytic inhibitor and metformin could prove an efficient therapeutic approach for treating TNBCs, albeit the efficacy of the combination treatment may depend on metabolic heterogeneity across various subtypes of TNBCs.

Glycolytic stress deteriorates 229E virulence to improve host defense response.

Viral infection treatment is a difficult task due to its complex structure and metabolism. Additionally, viruses can alter the metabolism of host cells, mutate, and readily adjust to harsh environments. Coronavirus stimulates glycolysis, weakens mitochondrial activity, and impairs infected cells. In this study, we investigated the efficacy of 2-DG in inhibiting coronavirus-induced metabolic processes and antiviral host defense systems, which have not been explored so far. 2-Deoxy-d-glucose (2-DG), a molecule restricting substrate availability, has recently gained attention as a potential antiviral drug. The results revealed that 229E human coronavirus promoted glycolysis, producing a significant increase in the concentration of fluorescent 2-NBDG, a glucose analog, particularly in the infected host cells. The addition of 2-DG decreased its viral replication and suppressed infection-induced cell death and cytopathic effects, thereby improving the antiviral host defense response. It was also observed that administration of low doses of 2-DG inhibited glucose uptake, indicating that 2-DG consumption in virus-infected host cells was mediated by high-affinity glucose transporters, whose levels were amplified upon coronavirus infection. Our findings indicated that 2-DG could be a potential drug to improve the host defense system in coronavirus-infected cells.

2-Deoxy-D-glucose inhibits lymphocytic choriomeningitis virus propagation by targeting glycoprotein N-glycosylation.

BACKGROUND: Increased glucose uptake and utilization via aerobic glycolysis are among the most prominent hallmarks of tumor cell metabolism. Accumulating evidence suggests that similar metabolic changes are also triggered in many virus-infected cells. Viral propagation, like highly proliferative tumor cells, increases the demand for energy and macromolecular synthesis, leading to high bioenergetic and biosynthetic requirements. Although significant progress has been made in understanding the metabolic changes induced by viruses, the interaction between host cell metabolism and arenavirus infection remains unclear. Our study sheds light on these processes during lymphocytic choriomeningitis virus (LCMV) infection, a model representative of the Arenaviridae family. METHODS: The impact of LCMV on glucose metabolism in MRC-5 cells was studied using reverse transcription-quantitative PCR and biochemical assays. A focus-forming assay and western blot analysis were used to determine the effects of glucose deficiency and glycolysis inhibition on the production of infectious LCMV particles. RESULTS: Despite changes in the expression of glucose transporters and glycolytic enzymes, LCMV infection did not result in increased glucose uptake or lactate excretion. Accordingly, depriving LCMV-infected cells of extracellular glucose or inhibiting lactate production had no impact on viral propagation. However, treatment with the commonly used glycolytic inhibitor 2-deoxy-D-glucose (2-DG) profoundly reduced the production of infectious LCMV particles. This effect of 2-DG was further shown to be the result of suppressed N-linked glycosylation of the viral glycoprotein. CONCLUSIONS: Although our results showed that the LCMV life cycle is not dependent on glucose supply or utilization, they did confirm the importance of N-glycosylation of LCMV GP-C. 2-DG potently reduces LCMV propagation not by disrupting glycolytic flux but by inhibiting N-linked protein glycosylation. These findings highlight the potential for developing new, targeted antiviral therapies that could be relevant to a wider range of arenaviruses.

Antiseizure effect of 2-deoxyglucose is not dependent on the presynaptic vacuole ATP pump or the somatic ATP-sensitive K+ channel.

Inhibition of glycolysis with 2-deoxyglucose (2-DG) produces antiseizure effects in brain slices and animal models, yet the mechanisms remain elusive. Here, we examined two glycolysis-derived ATP-associated mechanisms: vacuole ATP pump (V-ATPase) and ATP-sensitive K+ channel (KATP). Epileptiform bursts were generated in the CA3 area of hippocampal slices by 0 Mg2+ and 4-aminopyridine. 2-DG consistently abolished epileptiform bursts in the presence of pyruvate (to sustain tricarboxylic acid cycle for oxidative ATP production) at 30-33°C but not at room temperature (22°C). Under physiological conditions, 2-DG did not reduce the amplitude of evoked excitatory postsynaptic currents (EPSCs) or the paired-pulse ratio in CA3 neurons. During repetitive high-frequency (20 Hz, 20-50 pulses) stimulation, 2-DG did not accelerate the decline of EPSCs (i.e., depletion of transmitter release), even when preincubated with 8 mM K+ to enhance activity-dependent uptake of 2-DG. In addition, in 2-DG tetanic stimulation (200 Hz, 1 s) dramatically increased rather than diminished the occurrence of spontaneous EPSCs immediately after stimulation (i.e., no transmitter depletion). Moreover, a V-ATPase blocker (concanamycin) failed to block epileptiform bursts that were subsequently abolished by 2-DG. Furthermore, 2-DG did not induce detectable KATP current in hippocampal neurons. Finally, epileptiform bursts were not affected by either a KATP opener (diazoxide) or a KATP blocker (glibenclamide) but were blocked by 2-DG in the same slices. Altogether, these data suggest that 2-DG's antiseizure action is temperature dependent and achieved exclusively by inhibition of glycolysis and is not likely to be mediated by the two membrane-bound ATP-associated machinery mechanisms, V-ATPase and KATP.NEW & NOTEWORTHY Inhibition of glycolysis with 2-deoxyglucose (2-DG) represents a novel metabolic antiseizure approach, yet the mechanisms remain elusive. Here, we show that 2-DG's antiseizure action is both glycolysis and temperature dependent but not mediated by the vacuole ATP pump (V-ATPase) or ATP-sensitive K+ channel (KATP). Our data provide new insights to understand 2-DG's cellular mechanisms of action and, more broadly, neuronal metabolism and excitability.

2DG and glycolysis as therapeutic targets for status epilepticus.

2-deoxy-D-glucose (2DG) is a glucose analog differing from glucose only by removal of an oxygen atom at the 2 position, which prevents the isomerization of glucose-6-phosphate to fructose-6-phosphate, and thereby reversibly inhibits glycolysis. PET studies of regional brain glucose utilization positron-emitting 18F-2DG demonstrate that brain regions generating seizures have diminished glucose utilization during interictal conditions, but rapidly transition to markedly increased glucose delivery and utilization during seizures, particularly in status epilepticus (SE). 2-deoxy-D-glucose has acute antiseizure actions in multiple in vivo and in vitro seizure models, including models of SE induced by the chemo convulsants pilocarpine and kainic acid, suggesting that focal enhanced delivery of 2DG to ictal brain circuits is a potential novel anticonvulsant intervention for the treatment of SE.

2-Deoxy-D-glucose simultaneously targets glycolysis and Wnt/ß-catenin signaling to inhibit cervical cancer progression.

Cervical cancer is one of the most common female malignant tumors, with typical cancer metabolism characteristics of increased glycolysis flux and lactate accumulation. 2-Deoxy-D-glucose (2-DG) is a glycolysis inhibitor that acts on hexokinase, the first rate-limiting enzyme in the glycolysis pathway. In this research, we demonstrated that 2-DG effectively reduced glycolysis and impaired mitochondrial function in cervical cancer cell lines HeLa and SiHa. Cell function experiments revealed that 2-DG significantly inhibited cell growth, migration, and invasion, and induced G0/G1 phase arrest at non-cytotoxic concentrations. In addition, we found that 2-DG down-regulated Wingless-type (Wnt)/ß-catenin signaling. Mechanistically, 2-DG accelerated the degradation of ß-catenin protein, which resulted in the decrease of ß-catenin expression in both nucleus and cytoplasm. The Wnt agonist lithium chloride and ß-catenin overexpression vector could partially reverse the inhibition of malignant phenotype by 2-DG. These data suggested that 2-DG exerted its anti-cancer effects on cervical cancer by co-targeting glycolysis and Wnt/ß-catenin signaling. As expected, the combination of 2-DG and Wnt inhibitor synergistically inhibited cell growth. It is noteworthy that, down-regulation of Wnt/ß-catenin signaling also inhibited glycolysis, indicating a similar positive feedback regulation between glycolysis and Wnt/ß-catenin signaling. In conclusion, we investigated the molecular mechanism by which 2-DG inhibits the progression of cervical cancer in vitro, elucidated the interregulation between glycolysis and Wnt/ß-catenin signaling, and preliminarily explored the effect of combined targeting of glycolysis and Wnt/ß-catenin signaling on cell proliferation, which provides more possibilities for the formulation of subsequent clinical treatment strategies.

The Glycolysis Inhibitor 2-Deoxy-D-Glucose Exerts Different Neuronal Effects at Circuit and Cellular Levels, Partially Reverses Behavioral Alterations and does not Prevent NADPH Diaphorase Activity Reduction in the Intrahippocampal Kainic Acid Model of Temporal Lobe Epilepsy.

Temporal lobe epilepsy is the most drug-resistant type with the highest incidence among the other focal epilepsies. Metabolic manipulations are of great interest among others, glycolysis inhibitors like 2-deoxy D-glucose (2-DG) being the most promising intervention. Here, we sought to investigate the effects of 2-DG treatment on cellular and circuit level electrophysiological properties using patch-clamp and local field potentials recordings and behavioral alterations such as depression and anxiety behaviors, and changes in nitric oxide signaling in the intrahippocampal kainic acid model. We found that epileptic animals were less anxious, more depressed, with more locomotion activity. Interestingly, by masking the effect of increased locomotor activity on the parameters of the zero-maze test, no altered anxiety behavior was noted in epileptic animals. However, 2-DG could partially reverse the behavioral changes induced by kainic acid. The findings also showed that 2-DG treatment partially suppresses cellular level alterations while failing to reverse circuit-level changes resulting from kainic acid injection. Analysis of NADPH-diaphorase positive neurons in the CA1 area of the hippocampus revealed that the number of positive neurons was significantly reduced in dorsal CA1 of the epileptic animals and 2-DG treatment did not affect the diminishing effect of kainic acid on NADPH-d+ neurons in the CA1 area. In the control group receiving 2-DG, however, an augmented NADPH-d+ cell number was noted. These data suggest that 2-DG cannot suppress epileptiform activity at the circuit-level in this model of epilepsy and therefore, may fail to control the seizures in temporal lobe epilepsy cases.