Novel quinazolinone MJ-33 induces AKT/mTOR-mediated autophagy-associated apoptosis in 5FU-resistant colorectal cancer cells

Novel quinazolinone compounds have been studied in the field of drug discovery for a long time. Among their broad range of pharmacological effects, certain compounds effectively inhibit cancer cell proliferation. MJ-33 is a quinazolinone derivative with proposed anticancer activities that was synthesized in our laboratory. The present study aimed to evaluate the anticancer activity of MJ-33 in fluorouracil (5FU)-resistant colorectal cancer cells (HT-29/5FUR) and to investigate the underlying molecular mechanisms. The cell viability assay results indicated that HT-29/5FUR cell viability was inhibited by MJ-33 treatment in a concentration-dependent manner compared with the control group. The cellular morphological alterations observed following MJ-33 treatment indicated the occurrence of apoptosis and autophagy, as well as inhibition of cell proliferation in a time-dependent manner compared with the control group. The acridine orange, LysoTracker Red and LC3-green fluorescent protein staining results indicated that MJ-33 treatment significantly induced autophagy compared with the control group. The DAPI/TUNEL dual staining results demonstrated increased nuclear fragmentation and condensation following MJ-33 treatment compared with the control group. The Annexin V apoptosis assay and image cytometry analysis results demonstrated a significant increase in apoptotic cells following MJ-33 treatment compared with the control group. The western blotting results demonstrated markedly decreased Bcl-2, phosphorylated (p)-BAD, pro-caspase-9 and pro-caspase-3 expression levels, and notably increased cytochrome c and apoptotic peptidase activating factor 1 expression levels following MJ-33 treatment compared with the control group. Moreover, the expression levels of autophagy-related proteins, including autophagy related (ATG)-5, ATG-7, ATG-12, ATG-16, p62 and LC3-II, were increased following MJ-33 treatment compared with the control group. Furthermore, MJ-33-treated HT-29/5FUR cells displayed decreased expression levels of p-AKT and p-mTOR compared with control cells. The results suggested that MJ-33-induced apoptosis was mediated by AKT signaling, and subsequently modulated via the mitochondria-dependent signaling pathway. Therefore, the results suggested that suppression of AKT/mTOR activity triggered autophagy in the HT-29/5FUR cell line. In summary, the results indicated that MJ-33 inhibited HT-29/5FUR cell viability, and induced apoptosis and autophagy via the AKT/mTOR signaling pathway. The present study may provide novel insight into the anticancer effects and mechanisms underlying MJ-33 in 5FU-resistant colorectal cancer cells.