Inhibition of Nicotinamide Phosphoribosyltransferase (NAMPT), an Enzyme Essential for NAD+ Biosynthesis, Leads to Altered Carbohydrate Metabolism in Cancer Cells

Abstract

Background: NAMPT inhibition leads to attenuation of glycolysis in cancer cells. Results: NAMPT inhibition also perturbs other carbohydrate metabolism, resulting in elevated fructose 1-phosphate, sedoheptulose 1-phosphate, glyceraldehyde, and erythrose levels. Conclusion: Condensation of dihydroxyacetone phosphate/glyceraldehyde and dihydroxyacetone phosphate/erythrose by aldolase leads to increased fructose 1-phosphate and sedoheptulose 1-phosphate, respectively. Significance: NAMPT plays a key role in regulating glycolysis-related carbohydrate metabolism in cancer cells.

Discovery of a Highly Selective NAMPT Inhibitor That Demonstrates Robust Efficacy and Improved Retinal Toxicity with Nicotinic Acid Coadministration

“…One of the major consequences of NAMPT inhibition is the attenuation of glycolysis at the GAPDH step because this enzyme requires NAD þ for activity (37,38). As a result, metabolites such as fructose 1, 6-bisphosphate (FBP) and dihydroxyacetone phosphate (DHAP) before or at the GAPDH step are increased.…”

“…NAD þ levels were determined by measuring fluorescence at 360 (excitation) and 420 nm (emission; CytoFluor reader). NAD þ levels were also measured by LC-MS as described previously (37,38). For NA rescue time course studies, cells were grown and treated with LSN3154567 (100 nmol/L) AE NA (10 mmol/L) for different periods of times prior to measuring NAD þ levels.…”

“…Cancer cells were cultured as described before (37,38). Cells were seeded into a 96-well culture plate at a density of 8 Â 10 4 cells per well and incubated at 37 C in 5% CO 2 for 4 hours, and then treated with LSN3154567 at various concentrations (0.03-1,000 nmol/L) for 24 hours.…”

“…Tumor samples were collected 7 hours after the second dose and processed as described before (37). NAD þ levels were quantified as described before (38).…”

“…Analysis of dihydroxyacetone phosphate and fructose 1,6,-bisphosphate Cellular and tumor extracts were prepared and injected into an LC-triple quadrupole mass spectrometer (API 4000, AB Sciex), coupled with a pair of Shimadzu 10ADVP pumps, and a Shimadzu SIL-20A autosampler (Shimadzu) as described before (38).…”