Resistance to chemotherapy is associated with altered glucose metabolism in acute myeloid leukemia

Abstract

Altered glucose metabolism has been described as a cause of chemoresistance in multiple tumor types. The present study aimed to identify the expression profile of glucose metabolism in drug-resistant acute myeloid leukemia (AML) cells and provide potential strategies for the treatment of drug-resistant AML. Bone marrow and serum samples were obtained from patients with AML that were newly diagnosed or had relapsed. The messenger RNA expression of hypoxia inducible factor (HIF)-1α, glucose transporter (GLUT)1, and hexokinase-II was measured by quantitative polymerase chain reaction. The levels of LDH and β subunit of human F1-F0 adenosine triphosphate synthase (β-F1-ATPase) were detected by enzyme-linked immunosorbent and western blot assays. The HL-60 and HL-60/ADR cell lines were used to evaluate glycolytic activity and effect of glycolysis inhibition on cellular proliferation and apoptosis. Drug-resistant HL-60/ADR cells exhibited a significantly increased level of glycolysis compared with the drug-sensitive HL-60 cell line. The expression of HIF-1α, hexokinase-II, GLUT1 and LDH were increased in AML patients with no remission (NR), compared to healthy control individuals and patients with complete remission (CR) and partial remission. The expression of β-F1-ATPase in patients with NR was decreased compared with the expression in the CR group. Treatment of HL-60/ADR cells with 2-deoxy-D-glucose or 3-bromopyruvate increased in vitro sensitivity to Adriamycin (ADR), while treatment of HL-60 cells did not affect drug cytotoxicity. Subsequent to treatment for 24 h, apoptosis in these two cell lines showed no significant difference. However, glycolytic inhibitors in combination with ADR increased cellular necrosis. These findings indicate that increased glycolysis and low efficiency of oxidative phosphorylation may contribute to drug resistance. Targeting glycolysis is a viable strategy for modulating chemoresistance in AML.

Keywords: acute myeloid leukemia; chemoresistance; glycolysis; oxidative phosphorylation.

Figures

Figure 1.

Expression of HIF-1α, GLUT1 and HK-II mRNA in blasts from AML patients with various responses to chemotherapy. The mRNA expression of these genes was detected by reverse transcription-quantitative polymerase chain reaction. The upregulation of these three genes was associated with chemoresistance in AML patients (*P

Figure 2.

Expression of HIF-1α, GLUT1 and HK-II mRNA in AML ADR-resistant and sensitive cell lines. The expression levels of HIF-1α, GLUT1 and HK-II mRNA in the ADR-sensitive HL-60 and ADR-resistant HL-60/ADR cell lines were detected by reverse transcription-quantitative polymerase chain reaction. The upregulation of these three genes was associated with drug resistance in the HL-60/ADR cell line (**P

Figure 3.

Expression of β-F1-ATPase in AML ADR-resistant and sensitive cell lines. The β-F1-ATPase protein expression level in the ADR-sensitive HL-60 and ADR-resistant HL-60/ADR cell lines was detected by western blot analysis. Lane 1, HL-60 cells; Lane 2, HL-60/ADR cells. ADR, Adriamycin; AML, acute myeloid leukemia; β-F1-ATPase, β subunit of human F1-F0 adenosine triphosphate synthase.

Figure 4.

Effect of glycolysis inhibitors on glucose consumption and ADR-induced cytotoxicity in AML cell lines. Graphic representation of (A) relative glucose consumption or (B) in vitro ADR responsiveness in ADR-resistant and ADR-sensitive AML cell lines subsequent to 2-DG or 3BrPA treatment. Glucose consumption was calculated by measuring the conversion of glucose to 6-phosphogluconate, and glucose consumption in HL-60/ADR cells was set as 100%. The response to ADR was measured using the methyl thiazolyl tetrazolium assay, and cell survival in non-treated cells was set as 100%. Representative experiments were shown; data are expressed as the mean ± standard deviation (n=3). The combination of glycolysis inhibitor and ADR showed the synergistic effect in ADR-resistant HL-60/ADR cells (**P<0.01 vs. control group). ADR, Adriamycin; AML, acute myeloid leukemia; 2-DG, 2-deoxy-D-glucose; 3BrPA, 3-bromopyruvate.

Figure 5.

Effects of glycolysis inhibitiors on apopotosis in AML cell lines. ADR-induced apoptosis was measured in (A) HL-60 and (B) HL-60/ADR cells following treatment with 2-DG or 3BrPA using flow cytometry. The concentrations of ADR, 2-DG and 3BrPA used to treat the ADR-resistant HL-60/ADR cells were 0.5 µg/ml, 2 mM and 20 µM, respectively. ADR-sensitive HL-60 cells were incubated with 0.001 µg/ml ADR, 0.5 mM 2-DG or 20 µM 3BrPA. The data are expressed as the mean ± standard deviation (n=3). ADR, Adriamycin; AML, acute myeloid leukemia; 2-DG, 2-deoxy-D-glucose; 3BrPA, 3-bromopyruvate;