Abstract
β-Lapachone is a natural product that can promote ROS generation and ultimately triggers tumor cells death by inducing DNA damage. Recent studies have indicated that the targeting of ferroptosis or iron metabolism is a feasible strategy for treating cancer. In this study, bulk RNA-seq analysis suggested that β-Lapachone might induce ferroptosis in CRC cells. We further tested this hypothesis using a xenograft model of human colorectal cancer as an animal model and in SW620 and DLD-1 of CRC cell lines. Western blot was used to determine the key proteins of ferroptosis (SLC7A11, GPX4), autophagy (LC3B, P62, ATG7), ferritinophagy (NCOA4, FTH1, TFRC), and JNK pathway (p-JNK, JNK, p-c-Jun, c-Jun). The levels of MDA, GSH/GSSG, lipid ROS, and intracellular ferrous iron were determined after β-Lapachone treatment, and inhibitors of various pathways, including NAC, Ferrostatin-1, DFO, 3-MA, and SP600125 were utilized to explore the molecular mechanism underlying β-Lapachone-mediated ferroptosis. As the result, we identified that β-Lapachone inhibited cell proliferation and induced apoptosis, autophagy, and ROS generation. In addition, β-Lapachone induced ferroptosis as demonstrated by intra-cellular iron overload, increased levels of lipid ROS and MDA. Mechanistically, JNK signaling pathway was involved in β-Lapachone-induced xCT/GPX4-mediated ferroptosis and NCOA4-mediated ferritinophagy in CRC cells. In vivo experiments in nude mice demonstrated that β-Lapachone significantly inhibited CRC growth and induced ferroptosis and NCOA4-mediated ferritinophagy. These findings not only identify a novel role for β-Lapachone in ferroptosis but also indicate that β-Lapachone may be a valuable candidate for the research and development of anti-cancer therapeutic agents.