Glut-1 as a therapeutic target: increased chemoresistance and HIF-1-independent link with cell turnover is revealed through COMPARE analysis and metabolomic studies

Abstract

The facilitative glucose transporter Glut-1 is overexpressed and confers poor prognosis in a wide range of solid tumours. The peri-necrotic pattern of expression often seen in human tumour samples is linked with its transcriptional control in hypoxic conditions by hypoxia-inducible factor HIF-1 or through a reduced rate of oxidative phosphorylation. Hypoxia-regulated genes offer promise as novel therapeutic targets as a means of preventing the proliferation and eventual metastatic spread of tissue originating from residual chemically and radio resistant hypoxic cells that have survived treatment. Inhibiting the expression or functionality of Glut-1 may be a way of specifically targeting hypoxic cells within the tumour that depend upon a high rate of glucose uptake for anaerobic glycolysis. We used an array of formalin-fixed, paraffin-embedded samples of the NCI-60 panel of cell lines to carry out immunohistochemical detection of Glut-1 and to select possible candidate lead compounds by COMPARE analysis with agents from the NCI diversity screen, which may work via inhibition of Glut-1 or Glut-1-dependent processes. "Positive" COMPARE hits were mostly conjugated Pseudomonas toxins binding the epidermal growth factor receptor (EGFR). However, correlations with standard anticancer agents were virtually all negative, indicating a link between Glut-1 and chemoresistance. MTT proliferation assays carried out using stable, Glut-1 overexpressing cell lines generated from the bladder EJ138, human fibrosarcoma HT 1080 and the hepatoma wild type Hepa and HIF-1B-deficient c4 tumour cell lines revealed a cell line-dependent increase in chemoresistance to dacarbazine, vincristine and the bioreductive agent EO9 in Glut-1 overexpressing EJ138 relative to WT and empty vector controls. Metabolomic analysis ((31)P-MRS and (1)H MRS) carried out using cell lysates and xenografts generated from Glut-1 overexpressing Hepa and c4 cell lines showed higher glucose levels in Glut-1 overxpressing c4 relative to parental tumour extracts occurred in the absence of an increase in lactate levels, which were in turn significantly higher in the Glut-1 overexpressing Hepa xenografts. This implies that Glut-1 over-expression without a co-ordinate increase in HIF-1-regulated glycolytic enzymes increases glucose uptake but not the rate of glycolysis. Glut-1 overexpressing xenografts also showed higher levels of phosphodiester (PDE), which relates to the metabolite turnover of phospholipids and is involved in membrane lipid degradation, indicating a mechanism by which Glut-1 may increase cell turnover.

GLUT1 Expression Is Increased in Hepatocellular Carcinoma and Promotes Tumorigenesis

“…For instance, the PET tracer FDG has recently been investigated in mouse models of breast cancer as a radiomolecular therapy, and doses up to 5 mCi proved to be nonradiotoxic to normal organs. 38 In summary, this study revealed increased GLUT1 expression in a subset of HCC and suggests that this increased GLUT1 expression functionally affected proliferation and invasiveness of HCC cells. Herewith, GLUT1 expression in HCC appears as a potential innovative therapeutic target for this highly malignant tumor.…”

“…There has been significant progress in the theoretical and experimental characterization of the crystal structure of Glut1, which may prove useful for the rational design of Glut1-inhibiting agents. 37,38 Furthermore, lessons learned from the treatment of patients with Glut1 deficiency may open a way for overcoming potential adverse effects of such agents. 38 Studies investigating the use of glucose analogues or glucose conjugates that are likely to be taken up into target cells through Glut1 offer compelling evidence that the difference of Glut1 expression between the brain and tumors is large enough to allow targeting of Glut1.…”

“…37,38 Furthermore, lessons learned from the treatment of patients with Glut1 deficiency may open a way for overcoming potential adverse effects of such agents. 38 Studies investigating the use of glucose analogues or glucose conjugates that are likely to be taken up into target cells through Glut1 offer compelling evidence that the difference of Glut1 expression between the brain and tumors is large enough to allow targeting of Glut1. For instance, the PET tracer FDG has recently been investigated in mouse models of breast cancer as a radiomolecular therapy, and doses up to 5 mCi proved to be nonradiotoxic to normal organs.…”

Increased drug resistance is associated with reduced glucose levels and an enhanced glycolysis phenotype

“…To investigate the involvement of glucose uptake in the observed LG-associated drug resistance, Western blot analysis was performed for GLUT1, a major isoform of GLUT reported to be overexpressed in cancer (Macheda et al, 2005;Airley and Mobasheri, 2007;Evans et al, 2008). GLUT1 was elevated in LG conditions, but only in the PIK3CA mutant AGS and HGC27 cell lines ( Figure 3A).…”

“…To compensate for lower extracellular glucose levels, many cells will enhance glucose uptake through the GLUT family of transporters (Evans et al, 2008). To investigate the involvement of glucose uptake in the observed LG-associated drug resistance, Western blot analysis was performed for GLUT1, a major isoform of GLUT reported to be overexpressed in cancer (Macheda et al, 2005;Airley and Mobasheri, 2007;Evans et al, 2008).…”