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We report a simple synthetic scheme for the preparation of several azido-derived analogues of N-acetylglucosamine (GlcNAc). The synthesis of GlcNAc analogues has been achieved through a straightforward approach starting from GlcNAc-OMe via an intermediate C6 azido derivative. Products reported in this work were then obtained respectively by azido-alkyne cycloaddition reactions and reductive derivatizations of the same azido-intermediate. This synthetic pathway presents different possibilities of functionalization that can be exploited for the preparation of novel GlcNAc-based drugs. [ABSTRACT FROM AUTHOR]

Viruses have undergone evolutionary adaptations to tune their utilization of carbon sources, enabling them to extract specific cellular substrates necessary for their replication. The lack of a reliable cell culture system and a small-animal model has hampered our understanding of the molecular mechanism of replication of hepatitis E virus (HEV) genotype 1. Our recent identification of a replicative ensemble of mutant HEV RNA libraries has allowed us to study the metabolic prerequisites for HEV replication. Initial assessments revealed increased glucose and glutamine utilization during HEV replication. Inhibition of glycolysis and glycolysis + glutaminolysis reduced the levels of HEV replication to similar levels. An integrated analysis of protein-metabolite pathways suggests that HEV replication markedly alters glycolysis, the TCA cycle, and glutamine-associated metabolic pathways. Cells supporting HEV replication showed a requirement for fructose-6-phosphate and glutamine utilization through the hexosamine biosynthetic pathway (HBP), stimulating HSP70 expression to facilitate virus replication. Observations of mannose utilization and glutamine dependence suggest a crucial role of the HBP in supporting HEV replication. Inhibition of glycolysis and HSP70 activity or knockdown of glutamine fructose-6-phosphate amidotransferase expression led to a substantial reduction in HEV RNA and ORF2 expression accompanied by a significant decrease in HSP70 levels. This study demonstrates that glucose and glutamine play critical roles in facilitating HEV replication. [ABSTRACT FROM AUTHOR]

O-GlcNAcylation, a nutritionally driven, post-translational modification of proteins, is gaining importance because of its health implications. Changes in O-GlcNAcylation are observed in various disease conditions. Changes in O-GlcNAcylation by diet that causes hypercholesterolemia are not critically looked into in the liver. To address it, both in vitro and in vivo approaches were employed. Hypercholesterolemia was induced individually by feeding cholesterol (H)/high-fat (HF) diet. Global O-GlcNAcylation levels and modulation of AMPK activation in both preventive and curative approaches were looked into. Diet-induced hypercholesterolemia resulted in decreased O-GlcNAcylation of liver proteins which was associated with decreased O-linked N-acetylglucosaminyltransferase (OGT) and Glutamine fructose-6-phosphate amidotransferase-1 (GFAT1). Activation of AMPK by metformin in preventive mode restored the O-GlcNAcylation levels; however, metformin treatment of HepG2 cells in curative mode restored O-GlcNAcylation levels in HF but failed to in H condition (at 24 h). Further, maternal faulty diet resulted in decreased O-GlcNAcylation in pup liver despite feeding normal diet till adulthood. A faulty diet modulates global O-GlcNAcylation of liver proteins which is accompanied by decreased AMPK activation which could exacerbate metabolic syndromes through fat accumulation in the liver. [ABSTRACT FROM AUTHOR]

A number of cancer types have shown an increased prevalence and a higher mortality rate in patients with hyperglycemic associated pathologies. Although the correlation between diabetes and cancer incidence has been increasingly reported, the underlying molecular mechanisms beyond this association are not yet fully understood. Recent studies have suggested that high glucose levels support tumor progression through multiple mechanisms that are hallmarks of cancer, including cell proliferation, resistance to apoptosis, increased cell migration and invasiveness, epigenetic regulation (hyperglycemic memory), resistance to chemotherapy and altered metabolism. Most of the above occur because hyperglycemia through hexosamine biosynthetic pathway leads to aberrant O-GlcNAcylation of many intracellular proteins that are involved in those mechanisms. Deregulated O-GlcNAcylation is emerging as a general feature of cancer. Despite strong evidence suggesting that aberrant O-GlcNAcylation is or may be involved in the acquisition of all cancer hallmarks, it remains out of the list of the next generation of emerging hallmarks. Here, we discuss some of the current understanding on how hyperglycemia affects cancer cell biology and how aberrant O-GlcNAcylation stands in this context. [ABSTRACT FROM AUTHOR]

The myocardial response to pressure overload involves coordination of multiple transcriptional, posttranscriptional, and metabolic cues. The previous studies show that one such metabolic cue, O-GlcNAc, is elevated in the pressure-overloaded heart, and the increase in O-GlcNAcylation is required for cardiomyocyte hypertrophy in vitro. Yet, it is not clear whether and how O-GlcNAcylation participates in the hypertrophic response in vivo. Here, we addressed this question using patient samples and a preclinical model of heart failure. Protein O-GlcNAcylation levels were increased in myocardial tissue from heart failure patients compared with normal patients. To test the role of OGT in the heart, we subjected cardiomyocyte-specific, inducibly deficient Ogt (i-cm Ogt ) mice and Ogt competent littermate wild-type (WT) mice to transverse aortic constriction. Deletion of cardiomyocyte Ogt significantly decreased O-GlcNAcylation and exacerbated ventricular dysfunction, without producing widespread changes in metabolic transcripts. Although some changes in hypertrophic and fibrotic signaling were noted, there were no histological differences in hypertrophy or fibrosis. We next determined whether significant differences were present in i-cm Ogt cardiomyocytes from surgically naïve mice. Interestingly, markers of cardiomyocyte dedifferentiation were elevated in Ogt-deficient cardiomyocytes. Although no significant differences in cardiac dysfunction were apparent after recombination, it is possible that such changes in dedifferentiation markers could reflect a larger phenotypic shift within the Ogt-deficient cardiomyocytes. We conclude that cardiomyocyte Ogt is not required for cardiomyocyte hypertrophy in vivo; however, loss of Ogt may exert subtle phenotypic differences in cardiomyocytes that sensitize the heart to pressure overload-induced ventricular dysfunction. [ABSTRACT FROM AUTHOR]

Purpose: The severity of untreated or refractory diabetes mellitus has been functionally linked to elevated concentrations of free plasma glucose, clinically defined as hyperglycemia. Operationally, the pathophysiological presentations of prolonged hyperglycemia may be categorized within insulin-dependent and insulin-independent, type 1 and type 2 diabetic phenotypes, respectively. Accordingly, major areas of empirical biomedical research have focused on the elucidation of underlying mechanisms driving key cellular signaling systems that are significantly altered in patients presenting with diabetes-associated chronic hyperglycemia. Methods: Presently, we provide a translationally oriented review of key studies evaluating the aberrant effects of hyperglycemia on two major signaling pathways linked to debilitating cellular and systemic effects via targeted disruption of mitochondrial bioenergetics: (1) advanced glycation end-products (AGEs)/and their cognate receptor for advanced glycation end-products (RAGEs), and (2) the hexosamine biosynthetic pathway (HBP). Results: In preclinical models, cultured vascular endothelial cells exposed to hyperglycemic glucose concentrations were observed to produce enhanced levels of reactive oxygen species (ROS) functionally linked to increased formation of AGEs and expression of their cognate RAGEs. Importantly, inhibitors of AGEs formation, mitochondrial complex II, or un-couplers of oxidative phosphorylation, were observed to significantly reduce the effects of hyperglycemia on ROS production and cellular damage, thereby establishing a critical linkage to multiple levels of mitochondrial functioning. Hyperglycemia-mediated enhancement of mitochondrial ROS/superoxide production in vascular endothelial cells has been functionally linked to the shunting of glucose into the HBP with resultant long-term activation of pro-inflammatory signaling processes. Additionally, exposure of cultured cells to hyperglycemic conditions resulted in enhanced HBP-mediated inhibition of protein subunits of mitochondrial respiratory complexes I, III, and IV, intimately associated with normative cellular bioenergetics and ATP production. Conclusions: Convergent lines of evidence link chronic hyperglycemic conditions to aberrant expression of AGEs/RAGEs and HBP signaling pathways in relation to the pathophysiological formation of ROS and pro-inflammatory processes on the functional dysregulation of mitochondrial bioenergetics. [ABSTRACT FROM AUTHOR]

Glycometabolism is a distinctive aspect of energy metabolism in breast cancer, and key glycometabolism enzymes/pathways (glycolysis, hexosamine biosynthetic pathway, and pentose phosphate pathway) may directly or indirectly affect the clinical features. In this study, we analyzed the particular correlation between the altered glycometabolism and clinical features of breast cancer to instruct research and clinical treatment. Tissue microarrays containing 189 hollow needle aspiration samples and 295 triple-negative breast cancer tissues were used to test the expression of M2 isoform of pyruvate kinase (PKM2), glutamine-fructose-6-phosphate transaminase 1 (GFPT1), glucose-6-phosphate dehydrogenase (G6PD), and p53 by immunohistochemistry and the intensity of these glycometabolism-related protein was evaluated. Chi-square test, Kaplan-Meier estimates, and Cox proportional hazards model were used to analyze the relationship between the expression of these factors and major clinical features. PKM2, GFPT1, and G6PD affect the pathologic complete response rate of neoadjuvant chemotherapy patients in different ways; pyruvate kinase muscle isozyme 2 (PKM2) and G6PD are closely associated with the molecular subtypes, whereas GFPT1 is correlated with cancer size. All these three factors as well as p53 have impacts on the progression-free survival and overall survival of triple-negative breast cancer patients. Cancer size shows significant association with PKM2 and GFPT1 expression, while the pN stage and grade are associated with PKM2 and G6PD expression. Our study support that clinical characteristics are reflections of specific glycometabolism pathways, so their relationships may shed light on the orientation of research or clinical treatment. The expression of PKM2, GFPT1, and G6PD are hazardous factors for prognosis: high expression of these proteins predict worse progression-free survival and overall survival in triple-negative breast cancer, as well as worse pathologic complete response rate in neoadjuvant chemotherapy breast cancer. However, p53 appears as a protective factor only in the patients receiving neoadjuvant chemotherapy. All the four proteins, PKM2, GFPT1, G6PD and p53, are prognostic markers of breast cancer. The correlation among them suggests that there may be crosstalk of the four proteins in breast cancer. [ABSTRACT FROM AUTHOR]

Nuclear and cytosolic proteins are glycosylated on serine or threonine residues by O-linked β- N-acetylglucosamine ( O-GlcNAc). O-GlcNAc modification is one of various posttranslational modifications and seems to be involved in the modulation of transcription and signal transduction. Accumulating data suggest a role for O-GlcNAc-modified proteins in diabetes, acting as a glucose sensor. It has been suggested that the hexosamine biosynthetic pathway is involved in the mechanism causing insulin resistance and diabetic complications. Excess glucose entering into the hexosamine biosynthetic pathway might cause elevated O-GlcNAc modification of various proteins. In this article, we review the current data regarding the relationship between O-GlcNAc modification and diabetes. [ABSTRACT FROM AUTHOR]

The ability to regulate energy balance at both the cellular and whole body level is an essential process of life. As western society has shifted to a higher caloric diet and more sedentary lifestyle, the incidence of type 2 diabetes (non-insulin-dependent diabetes mellitus) has increased to epidemic proportions. Thus, type 2 diabetes has been described as a disease of 'chronic overnutrition'. There are abundant data to support the relationship between nutrient availability and insulin action. However, there have been multiple hypotheses and debates as to the mechanism by which nutrient availability modulates insulin signaling and how excess nutrients lead to insulin resistance. One well-established pathway for nutrient sensing is the hexosamine biosynthetic pathway (HSP), which produces the acetylated aminosugar nucleotide uridine 5′-diphospho-N-acetylglucosamine (UDP-GlcNAc) as its end product. Since UDP-GlcNAc is the donor substrate for modification of nucleocytoplasmic proteins at serine and threonine residues with N-acetylglucosamine (O-GlcNAc), the possibility of this posttranslational modification serving as the nutrient sensor has been proposed. We have recently directly tested this model in adipocytes by examining the effect of elevated levels of O-GlcNAc on insulin-stimulated glucose uptake. In this review, we summarize the existing work that implicates the HSP and O-GlcNAc modification as nutrient sensors and regulators of insulin signaling. [ABSTRACT FROM AUTHOR]

Several post-translational modifications figure prominently in ventricular remodeling. The beta-O-linkage of N-acetylglucosamine (O-GlcNAc) to proteins has emerged as an important signal in the cardiovascular system. Although there are limited insights about the regulation of the biosynthetic pathway that gives rise to the O-GlcNAc post-translational modification, much remains to be elucidated regarding the enzymes, such as O-GlcNAc transferase (OGT) and O-GlcNAcase (OGA), which regulate the presence/absence of O-GlcNAcylation. Recently, we showed that the transcription factor, E2F1, could negatively regulate OGT and OGA expression in vitro. The present study sought to determine whether E2f1 deletion would improve post-infarct ventricular function by de-repressing expression of OGT and OGA. Male and female mice were subjected to non-reperfused myocardial infarction (MI) and followed for 1 or 4 week. MI significantly increased E2F1 expression. Deletion of E2f1 alone was not sufficient to alter OGT or OGA expression in a naïve setting. Cardiac dysfunction was significantly attenuated at 1-week post-MI in E2f1-ablated mice. During chronic heart failure, E2f1 deletion also attenuated cardiac dysfunction. Despite the improvement in function, OGT and OGA expression was not normalized and protein O-GlcNAcyltion was not changed at 1-week post-MI. OGA expression was significantly upregulated at 4-week post-MI but overall protein O-GlcNAcylation was not changed. As an alternative explanation, we also performed guided transcriptional profiling of predicted targets of E2F1, which indicated potential differences in cardiac metabolism, angiogenesis, and apoptosis. E2f1 ablation increased heart size and preserved remote zone capillary density at 1-week post-MI. During chronic heart failure, cardiomyocytes in the remote zone of E2f1-deleted hearts were larger than wildtype. These data indicate that, overall, E2f1 exerts a deleterious effect on ventricular remodeling. Thus, E2f1 deletion improves ventricular remodeling with limited impact on enzymes regulating O-GlcNAcylation. [ABSTRACT FROM AUTHOR]