Discovery and Characterization of Novel Nonsubstrate and Substrate NAMPT Inhibitors

Abstract

Cancer cells are highly reliant on NAD-dependent processes, including glucose metabolism, calcium signaling, DNA repair, and regulation of gene expression. Nicotinamide phosphoribosyltransferase (NAMPT), the rate-limiting enzyme for NAD salvage from nicotinamide, has been investigated as a target for anticancer therapy. Known NAMPT inhibitors with potent cell activity are composed of a nitrogen-containing aromatic group, which is phosphoribosylated by the enzyme. Here, we identified two novel types of NAM-competitive NAMPT inhibitors, only one of which contains a modifiable, aromatic nitrogen that could be a phosphoribosyl acceptor. Both types of compound effectively deplete cellular NAD, and subsequently ATP, and produce cell death when NAMPT is inhibited in cultured cells for more than 48 hours. Careful characterization of the kinetics of NAMPT inhibition allowed us to optimize dosing to produce sufficient NAD depletion over time that resulted in efficacy in an HCT116 xenograft model. Our data demonstrate that direct phosphoribosylation of competitive inhibitors by the NAMPT enzyme is not required for potent cellular activity or antitumor efficacy. .

Recent Advances in NAMPT Inhibitors: A Novel Immunotherapic Strategy

“…Over the last years, this dogma was undermined by the identification of potent NAMPT inhibitors which did not contain a pyridine ring. For example, A-1293201 (28) ( Figure 11) is cytotoxic and able to deplete the NAD levels (Guo et al, 2017;Wilsbacher et al, 2017).…”

Utilization of18F-Fluorodeoxyglucose–Positron Emission Tomography To Understand the Mechanism of Nicotinamide Phosphoribosyltransferase Inhibitors In Vivo

“…Therefore, FDG uptake can give insights into glucose transport and hexokinase activity, which are likely to be impacted by ATP depletion resulting from upstream NAMPT inhibition. Previously published in vitro data also demonstrate that NADt levels are depleted within 24 hours, ATP levels are depleted within 48 to 72 hours, and cell viability is decreased at 72 hours, suggesting that cytotoxicity occurs after ATP depletion (Wilsbacher et al, 2017).…”

“…A-1293201 and A-1326133 are lead compounds optimized from this series that are potent inhibitors of NAMPT. Inhibition of NAMPT by these compounds for at least 48 hours resulting in .90% depletion of NADt was required to elicit NADt depletion-dependent cytotoxicity (Wilsbacher et al, 2017).…”

“…Inhibitors of NAMPT have been investigated as anticancer agents and elicit cytotoxicity via depletion of total cellular NAD 1 levels (Sampath et al, 2015). The mechanism of cytotoxicity is mediated by the sequential of depletion of total NAD levels, then ATP, and subsequently cell death (Wilsbacher et al, 2017). Given the retention of FDG in cells requires the ATP-dependent phosphorylation by hexokinase, the ability of FDG-PET to serve as a downstream pharmacodynamic marker of NAMPT inhibitors was investigated based on the following hypothesis: NAMPT inhibition will cause NAD 1 and ATP depletion, which are necessary for glycolysis.…”

“…Next, we performed FDG-PET experiments to understand the impact of the Preiss-Handler pathway on NAPRTdeficient and -proficient tumors by codosing NA with the NAMPT inhibitor. The NAMPT inhibitors used in these studies, A-1293201 and A-1326133, have previously been published (Wilsbacher et al, 2017).…”