NAD is mainly biosynthesized by the enzymatic action of nicotinamide phosphoribosyltransferase (NAMPT) through the salvage pathway. NAD is indispensable for the proper function and metabolism of all living cells, including cancer cells. Our previous researches revealed that inhibition of NAMPT by miRNA (miR) could suppress NAD levels and thereby hinder the growth and promotion of breast cancer (BC). Therefore, the current study was undertaken to investigate the inhibitory effects of miR-613 on NAMPT and BC cells' survival. Bioinformatics analysis and luciferase reporter assay confirmed that NAMPT 3′-untranslated region is a direct target for miR-613. The expression of miR-613 was noticed to be significantly decreased in both clinical tissue samples and BC cells by real-time PCR. Following transfection with miR-613 mimic, the expression of miR-613 was elevated in the BC cells leading to inhibition of NAMPT expression at both mRNA and protein level as measured by real-time PCR and western blotting, respectively. Inhibition of NAMPT led to a remarkable reduction in the concentration of NAD in the BC cells. The transfection also declined cell viability roughly 40% in MD Anderson-Metastatic Breast-231 (MDA-MB-231) cells. Consistently, the apoptosis rate was remarkably increased, around 65% in these cells as assayed by labeling the cells with Annexin V-fluorescein isothiocyanate (FITC) and Propidium Iodide. Targeting the NAMPT-mediated NAD salvage pathway by miR-613 is a novel approach for managing BC, which is worth further investigation.

中文翻译：

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MiR-613 通过下调烟酰胺磷酸核糖基转移酶和减少 NAD 促进乳腺癌细胞死亡

NAD 主要由烟酰胺磷酸核糖基转移酶 (NAMPT) 通过补救途径的酶促作用生物合成。NAD对于包括癌细胞在内的所有活细胞的正常功能和新陈代谢是必不可少的。我们之前的研究表明，miRNA (miR)对NAMPT的抑制可以抑制 NAD 水平，从而阻碍乳腺癌 (BC) 的生长和促进。因此，本研究旨在研究 miR-613 对NAMPT和 BC 细胞存活的抑制作用。生物信息学分析和荧光素酶报告基因检测证实NAMPT3'-非翻译区是 miR-613 的直接靶标。通过实时 PCR 观察到 miR-613 的表达在临床组织样本和 BC 细胞中均显着降低。用 miR-613 模拟物转染后，BC 细胞中 miR-613 的表达升高，导致分别通过实时 PCR 和蛋白质印迹法测量的 mRNA 和蛋白质水平的NAMPT表达受到抑制。抑制NAMPT导致 BC 细胞中 NAD 浓度的显着降低。在 MD Anderson-Metastatic Breast-231 (MDA-MB-231) 细胞中，转染也使细胞活力下降了大约 40%。一致地，通过用膜联蛋白 V-异硫氰酸荧光素 (FITC) 和碘化丙啶标记细胞进行测定，这些细胞的凋亡率显着增加，约为 65%。通过 miR-613靶向NAMPT介导的 NAD 补救途径是一种管理 BC 的新方法，值得进一步研究。