Figure 5. Metabolic effects of OPi.

(A) [U-¹³C]-glucose labeling of Glycolysis and TCA cycle intermediates in SKMEL5 cells treated with the indicated inhibitors (shown adjacent to 3-PG). The metabolites in which the incorporated ¹³C carbons were measured by GC-MS, are indicated in red. These are 3-PG (3-phospho glycerate), PYR (pyruvate), LAC (lactate), CA (citrate), MA (malate), and aspartate (Asp). The color intensities of the ¹³C-glucose-derived carbon atoms in each of these metabolites represent the percentage ranges of incorporation of these carbons, as shown in the box labeled “¹³C incorporation (%)”. (B) Percentage-range incorporation of ¹³C-glucose-derived carbons into lactate in A375, A375-R1 and SKMEL5 cells treated with the indicated inhibitors. (C) [U-¹³C]-glutamine labeling of TCA cycle intermediates in SKMEL5 cells treated with the indicated inhibitors (shown adjacent to 3-PG). Results are average of triplicates. (D) Percentage-range incorporation of ¹³C-glutamine-derived carbons from glutamine to malate in A375, A375-R1 and SKMEL5 cells treated with the indicated inhibitors. (E) Metabolites were extracted from SKMEL5 cells treated with the indicated inhibitors or mock for 12 h, and targeted LC-MS analysis was performed. Significant treatment-induced alterations (fold changes compared to mock) in nucleotide building blocks were plotted as average of triplicates. For all the above data, asterisks (*) indicate p<0.05, hashes (#) indicate p<0.01, and plus (+) indicate p<0.001 by T tests.