Figure 2.

Nelfinavir affects glycolysis by interfering with glucose phosphorylation mediated by HKII bound to VDACs. A, Glucose flux in AMO-1 cells was estimated by measuring the uptake of 2-NDBG upon nelfinavir treatment; 10 mmol/L 2-deoxyglucose (2-DG) served as a positive control of glucose flux inhibition. B, ECAR was assessed in AMO-1 cells after incubation with 20 μmol/L nelfinavir for 3 and 6 hours. C, Relative levels of intracellular glucose and glucose-6-phosphate (G6P) after the treatment with nelfinavir. For relative levels of glucose in the cell culture media 8 hours after the treatment, see also Supplementary Fig. S4. D, Relative levels of intracellular levels of lactate and pyruvate after the treatment with nelfinavir. The legend for C and D represents the fractional abundance of ¹³C isomers in the metabolites. E, A scheme illustrating change in level of metabolites from ¹³C glucose after 24 hours incubation with 20 μmol/L nelfinavir or DMSO only in AMO-1 cells. The color scale indicates log₂-fold change between the metabolites. For a detailed heat map illustrating the changes after 8 and 24 hours with 10 and 20 μmol/L nelfinavir, see also Supplementary Fig. S5. F, Live imaging of single-cell–derived colonies from the U-2 OS cells equipped with FL-HKII (left) and Tr-HKII (right) constructs. ER is visualized with mCherry-ER-3 vector and nuclei by Hoechst staining. G, Dose response curves of U-2 OS cells equipped with FL-HKII and Tr-HKII exposed to increasing concentrations of nelfinavir or 2-deoxyglucose. Data represent a mean ±SD from three replicates and statistically significant differences are marked. *, P < 0.05; **, P < 0.01; ***, P < 0.001.