Regulation of glucose uptake in lymphoma cell lines by c-MYC- and PI3K-dependent signaling pathways and impact of glycolytic pathways on cell viability

Martina Broecker-Preuss, Nina Becher-Boveleth, Andreas Bockisch, Ulrich Dührsen, Stefan Müller

Changes in glucose and energy metabolism contribute to the altered phenotype of cancer cells and are the basis for positron emission tomography with (18)F-fluoro-2-deoxy-D-glucose (FDG) to visualize tumors in vivo. The molecular background of the enhanced glucose uptake and its regulation in lymphoma cells is not fully clarified and may provide new possibilities to reverse the altered metabolism. Thus in this study we investigated regulation of glucose uptake by different signaling pathways. Furthermore, the effect of the glucose analog 2-deoxy-D-glucose (2-DG) alone and in combination with other inhibitors on cell survival was studied. An FDG uptake assay was established and uptake of FDG by lymphoma cells was determined after incubation with inhibitors of the c-MYC and the PI3K signalling pathways that are known to be activated in lymphoma cells and able to regulate glucose metabolism. Inhibitors of MAPK signalling pathways whose role in altered metabolism is still unclear were also investigated. Expression of mRNAs of the glucose transporter 1 (GLUT1), hexokinase 2 (HK2), glucose-6-phosphatase (G6Pase) and lactate dehydrogenase A (LDHA) and of the glucose metabolism-regulating micro RNAs (miRNA) miR21, -23a, -133a, -133b, -138-1 and -143 was determined by RT-PCR. Cell viability was analysed by MTT assay. Treatment with the c-MYC inhibitor 10058-F4 and inhibitors of the PI3K/mTOR pathway diminished uptake of FDG in all three cell lines, while inhibition of MAPK pathways had no effect on glucose uptake. Expression of glycolysis-related genes and miRNAs were diminished, although to a variable degree in the three cell lines. The c-MYC inhibitor, the PI3K inhibitor LY294002, the mTOR inhibitor Rapamycin and 2-DG all diminished the number of viable cells. Interestingly, in combination with 2-DG, the c-MYC inhibitor, LY294002 and the p38 MAPK inhibitor SB203580 had synergistic effects on cell viability in all three cell lines. c-MYC- and PI3K/mTOR-inhibitors decreased viability of the lymphoma cells and led to decreased glucose uptake, expression of glycolysis-associated genes, and glucose metabolism-regulating miRNAs. Inhibition of HK by 2-DG reduced cell numbers as a single agent and synergistically with inhibitors of other intracellular pathways. Thus, targeted inhibition of the pathways investigated here could be a strategy to suppress the glycolytic phenotype of lymphoma cells and reduce proliferation.