250 - 3PO inhibits glycolysis but does not bind to 6-phosphofructo-2-kinase/fructose-2,6-bisphosphatase-3 (PFKFB3) (ID 1342)
Abstract
Background and Aims
Intra-plaque angiogenesis (IP), present in advanced human atherosclerotic plaques, promotes unstable vascular disease and contributes to increased risk of myocardial infarction and stroke. Because proliferating endothelial cells (ECs) rely on glycolysis for up to 85% of their energy demand, glycolysis inhibition may represent a novel strategy to prevent IP neovascularization, thereby promoting plaque stabilization. 6-phosphofructo-2-kinase/fructose-2,6-bisphosphatase isoform 3 (PFKFB3) is a key enzyme of the glycolysis pathway and plays an essential role in angiogenesis. 3PO [3-(3-pyridinyl)-1-(4-pyridinyl)-2-propen-1-one] is frequently used as a partial and transient glycolysis inhibitor and thought to inhibit PFKFB3. However, this latter effect of 3PO has never been studied in detail.
Methods
We showed that 3PO inhibits glycolysis in ECs, inhibits capillary tube formation, EC migration, and formation of aortic sprouts. In order to demonstrate binding of 3PO to PFKFB3, we used isothermal titration calorimetry (ITC). To provide a complete thermodynamic profile of the molecular interaction, we used up to 6 mM of 3PO.
Results
However, we found that 3PO did not bind to PFKFB3. In contrast, 25 µM of AZ PFKFB3 67, a potent and specific PFKFB3 inhibitor, was sufficient to determine the binding constant by using ITC. Subsequently, we showed that 3PO hampers lactate transportation in ECs via monocarboxylate transporters (MCTs), thereby changing intracellular pH and indirectly affecting pH-dependent key regulatory enzymes of the glycolysis pathway.
Conclusions
In conclusion, the anti-glycolytic activity of 3PO relies in the capacity of this compound to interfere with MCTs and intracellular milieu acidification, rather than direct binding to PFKFB3.
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