Cytotoxicity of ungeremine towards multi-factorial drug resistant cancer cells and induction of apoptosis, ferroptosis, necroptosis and autophagy

Abstract

BackgroundSuccessful cancer chemotherapy is hampered by resistance of cancer cells to established anticancer drugs. Numerous natural products reveal cytotoxicity towards tumor cells. PurposeThe present study was aimed to determine the cytotoxicity of a betaine-type alkaloid, ungeremine, towards 9 cancer cell lines including various sensitive and drug-resistant phenotypes. The mode of action of this compound was further investigated. MethodsThe cytotoxicity, ferroptotic and necroptotic cell death were determined by the resazurin reduction assay. Caspase activation was evaluated using the caspase-Glo assay. Flow cytometry was applied for the analysis of cell cycle analysis (PI staining), apoptosis (annexin V/PI staining), mitochondrial membrane potential (MMP) (JC-1) and reactive oxygen species (ROS) (H2DCFH-DA). Apoptotic, necroptotic and autophagic markers were determined by Western blotting. CCRF-CEM leukemia cells were used for all mechanistic studies. ResultsUngeremine displayed cytotoxic activity towards the 9 cancer cell lines tested, including drug-sensitive and MDR phenotypes. The IC50values obtained varied from 3.67 µM (in MDA-MB-231-BCRP breast carcinoma cells) to 75.24 µM (against in CEM/ADR5000 leukemia cells) for ungeremine and from 0.02 µM (against CCRF-CEM cells) to 122.96 µM (against CEM/ADR5000 cells) for doxorubicin (control drug). Ungeremine induced ferroptosis, necroptosis, autophagy as well as apoptosis mediated by caspase activation, MMP alteration and increase ROS production. ConclusionThe present investigation showed that ungeremine is a promising cytotoxic compoundthat could be further explored in the future to develop new anticancer drugs to fight sensitive and resistant phenotypes.