title: Inhibition of nicotinamide phosphoribosyltransferase: cellular bioenergetics reveals a mitochondrial insensitive NAD pool.
abstract: The NAD rescue pathway consists of two enzymatic steps operated by nicotinamide phosphoribosyltransferase (Nampt) and nicotinamide mononucleotide adenylyltransferases. Recently, the potent Nampt inhibitor FK866 has been identified and evaluated in clinical trials against cancer. Yet, how Nampt inhibition affects NAD contents and bioenergetics is in part obscure. It is also unknown whether NAD rescue takes place in mitochondria, and FK866 alters NAD homeostasis within the organelle. Here, we show that FK866-dependent reduction of the NAD contents is paralleled by a concomitant increase of ATP in various cell types, in keeping with ATP utilization for NAD resynthesis. We also show that poly- and mono(ADP-ribose) transferases rather than Sirt-1 are responsible for NAD depletion in HeLa cells exposed to FK866. Mass spectrometry reveals that the drug distributes in the cytosolic and mitochondrial compartment. However, the cytoplasmic but not the mitochondrial NAD pool is reduced upon acute or chronic exposure to the drug. Accordingly, Nampt does not localize within the organelles and their bioenergetics is not affected by the drug. In the mouse, FK866-dependent reduction of NAD contents in various organs is prevented by inhibitors of poly(ADP-ribose) polymerases or the NAD precursor kynurenine. For the first time, our data indicate that mitochondria lack the canonical NAD rescue pathway, broadening current understanding of cellular bioenergetics.
authors: Department of Preclinical and Clinical Pharmacology, University of Florence, 50139 Florence, Italy.
authors: Maria
authors: M
authors: Pittelli
authors: Laura
authors: L
authors: Formentini
authors: Giuseppe
authors: G
authors: Faraco
authors: Andrea
authors: A
authors: Lapucci
authors: Elena
authors: E
authors: Rapizzi
authors: Francesca
authors: F
authors: Cialdai
authors: Giovanni
authors: G
authors: Romano
authors: Gloriano
authors: G
authors: Moneti
authors: Flavio
authors: F
authors: Moroni
authors: Alberto
authors: A
authors: Chiarugi
mesh_terms: D000178:Acrylamides / Q000494:pharmacology; D000255:Adenosine Triphosphate / Q000737:chemistry; D000818:Animals; D004791:Enzyme Inhibitors / Q000494:pharmacology*; D005347:Fibroblasts / Q000378:metabolism; D006367:HeLa Cells; D006801:Humans; D007737:Kynurenine / Q000737:chemistry; D008297:Male; D051379:Mice; D008928:Mitochondria / Q000378:metabolism*; D009243:NAD / Q000737:chemistry; D054409:Nicotinamide Phosphoribosyltransferase / Q000037:antagonists & inhibitors*; D010880:Piperidines / Q000494:pharmacology; D011065:Poly(ADP-ribose) Polymerases / Q000378:metabolism
chemical_list: D000178:Acrylamides; D004791:Enzyme Inhibitors; C480543:N-(4-(1-benzoylpiperidin-4-yl)butyl)-3-(pyridin-3-yl)acrylamide; D010880:Piperidines; D009243:NAD; D007737:Kynurenine; D000255:Adenosine Triphosphate; D054409:Nicotinamide Phosphoribosyltransferase; D011065:Poly(ADP-ribose) Polymerases