Glut1 is the main glucose transporter for glucose uptake and is expressed in nearly all mammalian cells. Glut1 is encoded by the Slc2a1 gene, and consists of a sugar-binding pocket facing the outer cell in the outward open conformation. Binding of glucose causes a conformational change so that Glut1 opens into the cytoplasm and release glucose inside the cell. Glut1 exhibit a Michaelis–Menten constant (Km) about 1mM, that is less than the normal blood glucose level (5.5mM), resulting in the continuous transport of glucose inside cells at an essentially constant rate. Similar to the insulin-responsive glucose transporter Glut4, Glut1 cell surface localization is controlled by extrinsic signals. Up-regulation of GLUT1 and consequent increase in glucose uptake occur in some conditions that require extraordinary metabolic needs, such as in the case of tumors and in the case of T cell activation.
1. Novel drugs targeting Glut1
A number of natural or synthetic Glut inhibitors have been discovered over the years and a comprehensive review can be found elsewhere 
. These molecules have been extensively explored as therapeutic option to treat cancer and are recently gaining interest in metabolic targeting in T cells as well. In this section, we provide a summary of the latest progress in the field, focusing especially on new generation molecules that specifically target Glut1. Three small molecules gained interest in recent years (Table 1
STF-31 selectively inhibits the glucose transporter Glut1 and selectively impairs cell growth of kidney and other types of cancer cells that lack the von Hippel-Lindau (VHL) tumor suppressor protein 
. Inactivation of VHL increases the expression of the hypoxia-inducible factor transcription factor HIF, which in turn stimulates the transcription of genes involved in glucose metabolism, including the Glut1 gene. VHL-deficient cancer cells depend on Glut1 and aerobic glycolysis for ATP production. STF-31 binds directly to the Glut1 transporter, blocking glucose uptake, resulting in necrosis in VHL-deficient cancer cells. Normal kidney cells are not strictly dependent on glycolysis, use Glut2 for glucose uptake, and are therefore insensitive to STF-31 toxicity. When used in a mouse model, STF-31 efficiently blocked tumor growth without significant toxic effect on other organs. STF-31 was shown to efficiently inhibit Glut1 dependent glucose uptake and to suppress glycolysis in human T cells overexpressing Glut1 
. Moreover, STF-31 was shown to impair glucose responsiveness and insulin secretion in human beta cells expressing Glut1 
WZB117 is a small molecule that inhibits Glut1-mediated glucose transport by binding reversibly at the exofacial sugar-binding site of Glut1 
. WZB117 was shown to induce cell death in lung and breast cancer cells without affecting normal cells 
. As for STF-31, in vivo treatment of animal models with WZB117 affected tumors without causing significant adverse events in treated animals. We used WZB117 to inhibit Glut1 mediated glucose transport in T cells 
. WZB117 reduces T cell proliferation by 90% at a concentration of 3µM and differentiation of naïve T cells into memory T cells. Even though both STF-31 and WZB117 have proven effective both in vitro and in pre-clinical models, their Glut1 blocking activity in the µM range and the controversial selectivity for Glut1 
represent significant limitations for their translation into the clinic.
So far, the most promising molecule for translation into the clinical setting is BAY-876 
. It is characterized by an IC50 of 2 nM, and it is highly selective for Glut1 with a selectivity factor of >100 against Glut2, Glut3, and Glut4. Importantly, it is orally bioavailable, and showed a good metabolic stability in vitro. As for the two other compounds, it was successfully tested in preclinical models, showing a potent anti-cancer activity 
. At the moment, no data are available with respect to the effect of BAY-876 on cells of the immune system.
2. Potential off-Target and Side Effects of Pharmacological Glut1 Blockade
The majority of the cells in the body express Glut1, but the effect of pharmacological Glut1 blockade largely depends on the co-expression of other Gluts, and on the metabolic activity of the cell, which determine the need for glucose. Two cell types that are strictly Glut1 dependent are erythrocytes and the endothelium of the blood brain barrier (BBB). Of all cell lineages, the human erythrocyte expresses the highest level of the Glut1 with a surface density of approximately 200.000 Glut1 molecules per cell 
. In addition to glucose, Glut1 also transports L-dehydroascorbic acid (DHA, vitamin C), and in human erythrocytes there is a preferential uptake of DHA instead of glucose 
. Glut1 expression in erythrocytes is a specific trait of vitamin C–deficient mammalian species, comprising only higher primates, guinea pigs, and fruit bats. In DHA synthetizing mammals, Glut1 expression is down-regulated after the neonatal period. These data suggest that the high expression of Glut1 in human erythrocytes in not related to a specific need for glucose when cells have a modest metabolic activity. Accordingly, treatment of human erythrocytes with the Glut1 inhibitor STF-31 significantly impaired glucose uptake but did not cause hemolysis or other effects 
The capillary endothelium of the brain, which makes up the BBB express Glut1, with a fundamental role in the transport of glucose from the blood to the central nervous system (CNS). The Glut1 deficiency syndrome (Glut1-DS) is the most relevant clinical setting showing the non-redundant role of Glut1 function in the BBB. The Glut1-DS is a rare genetic disease characterized by de novo or inherited mutations (approximately 100 identified) of the SLC2A1 gene. Mutations affect assembly, three-dimensional folding, trafficking to the cell membrane, or activation of the encoded protein Glut1 
. The extent of Glut1 dysfunction can be measured as in vitro uptake of glucose in erythrocytes showing that glucose transport is reduced by 30% to 70% as compared to healthy subjects 
. The disease has typical neurological symptoms including ataxia, lethargy, total body paralysis, movement disorders, and epilepsy. Neurological symptoms similar to those reported in patients with the Glut1-DS have to be considered as potential side effects when patients undergo to pharmacological Glut1 blockade. However, in the Glut1-DS most of the neurological symptoms are developed during childhood and in the adolescence, and later in adulthood most of the symptoms stabilize or even attenuate 
. The impact of genetic Glut1 deficiency on neurological damage appear to be related to the relative glucose shortage in the developing CNS, while once completely developed the impact of reduced glucose availability is limited. In terms of translation of a pharmacological blockade strategy in patients with T1D, the effects on the CNS have to be considered as the most relevant off-target effect, with important complications. To avoid neurological side effects pharmacological Glut1 blockade should be restricted to adolescent or adult patients and for a limited time period. Supporting this, clinical trials using 2DG in adult patients for several weeks, reported only mild and reversible neurological side effects, mainly dizziness 
A diabetes-specific side effect of pharmacological Glut blockade that has to be considered is the potential inhibition of insulin production by beta cells. Human, but not rat and mouse insulin producing beta cells express Glut1 
. Glut1 is used as glucose sensor for optimal insulin secretion and inhibition of Glut1 with STF31 resulted in impaired insulin secretion 
. Even though the effect of Glut1 inhibitors is reversible this suggests that pharmacological Glut1 blockade can only apply for a limited time period in order to restore proper insulin production. Moreover, nicotinamide phosphoribosyltransferase (Nampt), a rate-limiting enzyme in the mammalian NAD+ biosynthesis, has been suggested as an alternative target for STF-31 
. While staining for Nampt was found both in the exocrine and endocrine tissue of fetal pancreas, in adulthood, Nampt expression was localized predominantly in beta cells 
. These data suggest that beta cells are the main target for Glut1 blockade in human islets. Other potential target cells within the islet of Langherhans are alpha cells that were shown to express Glut1 
, but so far the effect of Glut1 blocking compounds on alpha cells has not been investigated.
Potential off-target and side effects of a pharmacological Glut1 blockade have not been addressed in humans, and animal models not always resemble humans for Glut1 expression and usage. An ideal model to study immune and endocrine effects of a reduced glucose transport through Glut1 is the Glut1-DS. This model has been extensively characterized for the neurological symptoms but so far no efforts have been made to address potential impairments in other cells and organs.