Inhibition of Nicotinamide Phosphoribosyltransferase Induces Apoptosis in Estrogen Receptor-Positive MCF-7 Breast Cancer Cells

Abstract

PurposeTumor cells have increased turnover of nicotinamide adenine dinucleotide (NAD+), the main coenzyme in processes including adenosine diphosphate-ribosylation, deacetylation, and calcium mobilization. NAD+ is predominantly synthesized in human cells via the salvage pathway, with the first component being nicotinamide. Nicotinamide phosphoribosyltransferase (NAMPT) is the key enzyme in this pathway, and its chemical inhibition by FK866 has elicited antitumor effects in several preclinical models of solid and hematologic cancers. However, its efficacy in estrogen receptor (ER)-positive and human epidermal growth factor receptor 2-positive breast cancer cells has not been previously investigated. In this study, we aimed to deplete the NAD+ content of MCF-7 cells, a model cell line for ER-positive breast cancer, by inhibiting NAMPT in order to evaluate downstream effects on p53 and its acetylation, p21 and Bcl-2-associated X protein (BAX) expression, and finally, apoptosis in MCF-7 breast cancer cells.MethodsMCF-7 cells were cultured and treated with FK866. NAD+ levels in cells were determined colorimetrically. Levels of p53 and its acetylated form were determined by Western blotting. Expression of p21 and BAX was determined by real-time polymerase chain reaction. Finally, levels of apoptosis were assessed by flow cytometry using markers for annexin V and propidium iodide.ResultsFK866 treatment was able to increase p53 levels and acetylation, upregulate BAX and p21 expression, and induce apoptosis in MCF-7 cells. Addition of exogenous NAD+ to cells reversed these effects, suggesting that FK866 exerted its effects by depleting NAD+ levels.ConclusionResults showed that FK866 could effectively inhibit NAD+ biosynthesis and induce programmed cell death in MCF-7 cells, suggesting that NAMPT inhibitors may be useful for the treatment of ER-positive breast cancers.

Down-regulation of NAMPT expression by miR-154 reduces the viability of breast cancer cells and increases their susceptibility to doxorubicin

“…Here we showed that inhibition of NAMPT and the further decline in NAD levels leads to diminished cell viability and a prominent induction of apoptosis. We had earlier reported that inhibition of NAMPT by its specific inhibitor could effectively reduce NAD levels and induce apoptosis in breast cancer cells (46). In line with our findings, inhibition of NAMPT by microRNAs has also been shown to be an appropriate method in modulating NAD content and cell viability (40)(41)(42).…”

Extracellular NAMPT/visfatin causes p53 deacetylation via NAD production and SIRT1 activation in breast cancer cells

“…Acetylation and phosphorylation are the regulatory mechanisms for p53 stability, and p53 acetylation also activates its sequence‐specific DNA‐binding activity . Inhibition of NAMPT and consequent decrease in cellular NAD levels have been shown to increase p53 acetylation in MCF‐7 breast cancer cells . Considering the importance of p53 as a substrate for SIRT1 and induction of SIRT1 by visfatin, in this study p53 acetylation status was investigated and found to be decreased by visfatin.…”

MicroRNA-494 induces breast cancer cell apoptosis and reduces cell viability by inhibition of nicotinamide phosphoribosyltransferase expression and activity

“…Previous studies have shown that NAMPT and miR-494 are involved in cell viability (Alaee et al, 2017[3], Liu et al, 2015[27], Chen et al, 2015[9], Zhang et al, 2013[54]). Viability of the cells was measured using WST-1.…”

“…It has been shown that inhibition of NAMPT leads to an induction of apoptosis and reduction of cell survival in cancer cell lines concomitant with the decline in NAD levels (Alaee et al, 2017[3]; Hasmann and Schemainda, 2003[18]; Yang et al, 2007[50]; Holen et al, 2008[21]) . Here we revealed that up-regulation of miR-494 reduced cell viability and induced apoptosis in BC cells.…”

“…The plate was incubated at RT in the dark room for 30 minutes following the addition of Cycling enzyme. Finally, the absorbance was measured at 450 nm by a plate reader and the concentration of NAD was determined (Alaee et al, 2017[3]).…”