Up-regulation of miR-381 inhibits NAD+ salvage pathway and promotes apoptosis in breast cancer cells

Abstract

Nicotinamide phosphoribosyltransferase (NAMPT), a rate-limiting enzyme involved in nicotinamide adenine di- nucleotide (NAD) salvage pathway, is overexpressed in many human malignancies such as breast cancer. This enzyme plays a critical role in survival and growth of cancer cells. MicroRNAs (miRNAs) are among the most important regulators of gene expression, and serve as potential targets for diagnosis, prognosis, and therapy of breast cancer. Therefore, the aim of this study was to asse ss the effect of NAMPT inhibition by miR-381 on breast cancer cell survival. MCF-7 and MDA-MB-2 31 cancer cell lines were transfected with miR-381 mimic, inhibitor, and their corresponding negative controls (NCs). Subsequently, the level of NAMPT and NAD was assessed using real-time PCR, immuno-blotting, and enzymatic methods, resp ectively. In order to evalua te apoptosis, cells were labelled with Annexin V-FITC and propidium iodide and analyzed by flow cytometry. Bioinformatics analysis was performed to recognize whether NAMPT 3 ′ -untranslated region (UTR) is a direct target of miR-381 and the results were authenticated by the luciferase re porter assay using a vector containing the 3 ′ -UTR sequence of NAMPT. Our results revealed that the 3 ′ -UTR of NAMPT was a direct target of miR-381 and its up-regulation decreased NAMPT gene and protein expression, leading to a notable reduction in intracellular NAD and subse- quently cell survival and induction of apoptosis. It can be concluded that miR-381 has a vital role in tumor sup- pression by down-regulation of NAMPT, and it can be a promising candidate for breast cancer therapy.