Nicotinamide N-methyltransferase inhibits autophagy induced by oxidative stress through suppressing the AMPK pathway in breast cancer cells

Background: Nicotinamide N-methyltransferase (NNMT) is highly expressed in several cancers and can regulate cell epigenetic status and various cell metabolism pathways, such as ATP synthesis and cellular stress response. We reported in our previous papers that NNMT overexpression inhibits the apoptosis and enhances the chemotherapy resistance of breast cancer cells. This study aims to investigate the effect of NNMT on autophagy induced by oxidative stress in breast cancer cells, which might provide a novel therapeutic strategy for breast cancer treatment.

Methods: NNMT and LC3B II protein levels in the two cell models (SK-BR-3 and MDA-MB-231) with NNMT overexpression or knockdown were detected by Western blotting and correlated with each other. Changes in cellular viability, intracellular reactive oxygen species (ROS) and ATP levels were assessed after H₂O₂ treatment. Then, autophagosomes were imaged by transmission electron microscopy, and LC3 puncta were examined by confocal microscopy and flow cytometry. The LC3B II level and AMPK-ULK1 pathway activity were both detected by Western blotting to determine the role of NNMT in the H₂O₂-induced autophagy.

Results: NNMT expression was negatively correlated with LC3B II expression in both cell models (SK-BR-3 and MDA-MB-231). Then, NNMT overexpression attenuated the autophagy induced by H₂O₂ in SK-BR-3 cells, whereas knockdown promoted autophagy induced by H₂O₂ in MDA-MB-231 cells. Furthermore, mechanistic studies showed that NNMT suppressed the ROS increase, ATP decrease and AMPK-ULK1 pathway activation, resulting in the inhibition of H₂O₂-induced autophagy in breast cancer cells.

Conclusions: We conclude that NNMT inhibits the autophagy induced by oxidative stress through the ROS-mediated AMPK-ULK1 pathway in breast cancer cells and may protect breast cancer cells against oxidative stress through autophagy suppression.

Keywords: AMPK; Autophagy; Breast cancer; Nicotinamide N-methyltransferase; Oxidative stress; ULK1.