EWS-FLI1 confers exquisite sensitivity to NAMPT inhibition in Ewing sarcoma cells

Abstract

Ewing sarcoma (EwS) is the second most common bone cancer in children and adolescents with a high metastatic potential. EwS development is driven by a specific chromosomal translocation resulting in the generation of a chimeric EWS-ETS transcription factor, most frequently EWS-FLI1.Nicotinamide adenine dinucleotide (NAD) is a key metabolite of energy metabolism involved in cellular redox reactions, DNA repair, and in the maintenance of genomic stability. This study describes targeting nicotinamide phosphoribosyltransferase (NAMPT), the rate-limiting enzyme of NAD synthesis, by FK866 in EwS cells. Here we report that blocking NAMPT leads to exhaustive NAD depletion in EwS cells, followed by a metabolic collapse and cell death. Using conditional EWS-FLI1 knockdown by doxycycline-inducible shRNA revealed that EWS-FLI1 depletion significantly reduces the sensitivity of EwS cells to NAMPT inhibition. Consistent with this finding, a comparison of 7 EwS cell lines of different genotypes with 5 Non-EwS cell lines and mesenchymal stem cells revealed significantly higher FK866 sensitivity of EWS-ETS positive EwS cells, with IC50 values mostly below 1nM.Taken together, our data reveal evidence of an important role of the NAMPT-mediated NAD salvage pathway in the energy homeostasis of EwS cells and suggest NAMPT inhibition as a potential new treatment approach for Ewing sarcoma.

EWS/FLI is a Master Regulator of Metabolic Reprogramming in Ewing Sarcoma

“…Consistent with a cancerous phenotype, Ewing sarcoma cells are hyperproliferative and require increased macromolecule synthesis to sustain accelerated growth and proliferation. Additionally, EF-driven alterations in nucleobase-containing metabolic processes such as (NAD salvage) could be sensitive to inhibition and therapeutic exploitation (42). The increased metabolic demands of rapid proliferation and growth could be exploited in similar ways.…”

Metabolic modulation of Ewing sarcoma cells inhibits tumor growth and stem cell properties

“…While preparing this manuscript, a study was published demonstrating the efficacy of metabolic inhibition of energy pathways in EWS cells through NAMPT, the rate limiting enzyme of NAD [ 70 ]. It is encouraging that multiple approaches to inducing energy stress through metabolic inhibition can indeed be a specific and effective therapeutic strategy.…”

Identifying the druggable interactome of EWS-FLI1 reveals MCL-1 dependent differential sensitivities of Ewing sarcoma cells to apoptosis inducers

“…Briefly, using the dox-inducible A673/TR/shEF cell line [ 41 ], transfections were performed with the Lipofectamine-Plus reagent (Invitrogen, Groningen, the Netherlands) according to the manufacturer’s recommendations. EWS-FLI1 knockdown and cell viability determinations were done as previously reported [ 42 ]. Transfections were carried out using Lipofectamine Plus reagent (Invitrogen, Groningen, the Netherlands).…”