Brevilin A Inhibits STAT3 Signaling and Induces ROS-Dependent Apoptosis, Mitochondrial Stress and Endoplasmic Reticulum Stress in MCF-7 Breast Cancer Cells

Abstract

Muhammad Zubair Saleem, 1 Muhammad Azhar Nisar, 1 Mohammed Alshwmi, 2 Syed Riaz Ud Din, 1 Yaser Gamallat, 1 Muhammad Khan, 3 Tonghui Ma 1 1College of Basic Medical Sciences, Dalian Medical University, Dalian, Liaoning 116044, People’s Republic of China; 2Department of Clinical Laboratory, The First Affiliated Hospital of Dalian Medical University, Dalian Medical University, Dalian, Liaoning 116044, People’s Republic of China; 3Department of Zoology, University of the Punjab, Lahore, Punjab 54590, PakistanCorrespondence: Muhammad KhanDepartment of Zoology, University of the Punjab, Quaid-e-Azam Campus, Lahore, Punjab 54590, PakistanTel [phone]Email [email protected] MaCollege of Basic Medical Sciences, Dalian Medical University, 9 Western Section, Lvshun South Road, Dalian 116044, People’s Republic of ChinaTel [phone]Fax [phone]Email [email protected]: Breast cancer is the most common malignancy among women across the globe. Despite concerted efforts to improve the prevailing treatment modalities, the overall prognosis of breast cancer remains unsatisfactory. Recently, antiproliferative activity of Brevilin A (Brv-A), a sesquiterpene lactone compound of Centipeda minima, has been unveiled in various cancer types. Here, we have explored anticancer activity of Brv-A in MCF-7 breast carcinoma cells by targeting various pathways.Materials and Methods: Cell proliferation rate was determined by CCK-8 and clonogenic assay. Cellular morphological changes were observed under phase contrast microscope while calcein-AM and PI was used for live/dead assay. Cell cycle assay was performed by flow cytometry. Apoptotic cell percentage was determined by Hoechst 33258 staining and flow cytometric analysis. ROS generation and mitochondrial membrane potential were measured using commercially available kits while protein expression was measured by Western blotting.Results: In our study, Brv-A exerted antiproliferative effect through mitotic arrest at G 2/M phase of cell cycle and induced apoptosis in MCF-7 cells in a dose-dependent manner. Induction of apoptosis by Brv-A was found to be associated with ROS generation by targeting NOX2 and NOX3, mitochondrial dysfunction (MMP dissipation and Bcl-2 family proteins modulation), DNA fragmentation, JNK and p38 MAPK activation, endoplasmic reticulum (ER) stress by increasing Bip/GRP78, ATF4 and CHOP protein expressions and inhibition of STAT3 activation via decreased phosphorylation of JAK2 and SRC. Pretreatment of NAC, a ROS scavenger, partially reversed the aforesaid cellular events indicating ROS generation as the primary event to modulate cellular targets for induction of apoptosis. Besides, Brv-A has also been documented for inhibition of cell migration via decrease in COX-2 and MMP-2 expression.Conclusion: Taken together, Brv-A induces G 2/M phase arrest, ROS-dependent apoptosis, ER stress, mitochondrial dysfunction and inhibits STAT3 activation in MCF-7 cells signifying it to be one of the potential anticancer therapeutics in future.Keywords: breast cancer, Brevilin A, apoptosis, ROS, ER stress, STAT3, mitochondrial dysfunction

Keywords

• breast cancer
• brevilin a
• apoptosis
• ros
• er stress
• stat3
• mitochondrial dysfunction.