Antitumor effect of combined NAMPT and CD73 inhibition in an ovarian cancer model.

Giovanna Sociali,Mirko Magnone,Aimable Nahimana,Nadia Raffaelli,Federica Zamporlini,Tiziana Vigliarolo,Santina Bruzzone,Alessio Nencioni,Laura Sturla,Lizzia Raffaghello,Laura Emionite,Giovanna Bianchi

doi:10.18632/oncotarget.6502

Abstract

Nicotinamide phosphoribosyltransferase (NAMPT) is a crucial enzyme in the biosynthesis of intracellular NAD+. NAMPT inhibitors have potent anticancer activity in several preclinical models by depleting NAD+ and ATP levels. Recently, we demonstrated that CD73 enables the utilization of extracellular NAD+/nicotinamide mononucleotide (NMN) by converting them to Nicotinamide riboside (NR), which can cross the plasmamembrane and fuel intracellular NAD+ biosynthesis in human cells. These processes are herein confirmed to also occur in a human ovarian carcinoma cell line (OVCAR-3), by means of CD73 or NRK1 specific silencing. Next, we investigated the anti-tumor activity of the simultaneous inhibition of NAMPT (with FK866) and CD73 (with α, β-methylene adenosine 5′-diphosphate, APCP), in an in vivo human ovarian carcinoma model. Interestingly, the combined therapy was found to significantly decrease intratumor NAD+, NMN and ATP levels, compared with single treatments. In addition, the concentration of these nucleotides in ascitic exudates was more remarkably reduced in animals treated with both FK866 and APCP compared with single treatments. Importantly, tumors treated with FK866 in combination with APCP contained a statistically significant lower proportion of Ki67 positive proliferating cells and a higher percentage of necrotic area. Finally, a slight but significant increase in animal survival in response to the combined therapy, compared to the single agents, could be demonstrated. Our results indicate that the pharmacological inhibition of CD73 enzymatic activity could be considered as a means to potentiate the anti-cancer effects of NAMPT inhibitors.

Highlights

Cells utilize nicotinamide adenine dinucleotide (NAD+) either as a coenzyme in redox reactions or as a substrate for NAD+-degrading enzymes, such as poly(ADP-ribose) polymerases (PARPs), ectocellular NAD+ases, including CD38 and sirtuins (SIRT1–7) [1]
To verify that OVCAR-3 cells represented an appropriate model for our study, we preemptively assessed i) the NAD+ biosynthetic pathways that are active in these cells, ii) their sensitivity to FK866, and iii) the expression of CD73
When OVCAR-3 cells were incubated for 24 h in the presence of FK866, NAM phosphoribosyltransferase (NAMPT) activity was almost completely inhibited, whereas the other enzymatic activities were not affected (Figure 1B): NAMPT inhibition with FK866 does not alter the enzymatic activity of the other enzymes involved in NAD+ synthesis

Summary

Introduction

Cells utilize nicotinamide adenine dinucleotide (NAD+) either as a coenzyme in redox reactions or as a substrate for NAD+-degrading enzymes, such as poly(ADP-ribose) polymerases (PARPs), ectocellular NAD+ases, including CD38 and sirtuins (SIRT1–7) [1]. NAD+ metabolism plays a crucial role in the fate of tumor cells and cellular NAD+ levels are the result of a fine balance between opposite processes of NAD+ synthesis and degradation. The mechanism by which NAMPT is fundamental in tumor progression certainly relies on the fact that NAD+ is required for the activity of NAD+-metabolizing enzymes, such as PARP1, SIRT1, SIRT6 and CD38, all having different cancerpromoting abilities, such as genomic stability, secretion of pro-inflammatory cytokines, angiogenesis, motility and invasion [8,9,10,11]

Methods

Results

Conclusion