Berberine sensitizes liver cancer cells to sorafenib by inducing SETDB1/NQO1/p53-dependent ferroptosis and genomic instability

Sorafenib, one of multi-kinase inhibitors, is a first-line clinical drug for hepatocellular carcinoma (HCC). However, it can only provide limited survival window and high recurrence of advanced HCC. So, to improve its therapeutical application and efficacy, it is urgent to develop a combination strategy. Berberine is a representative of bioactive compounds isolated from some medicinal herbs, and shows great anti-cancer capacity, including liver cancer. Importantly, berberine shows low toxicity to human beings, which makes it an ideal candidate. Therefore, the purpose of this study is to evaluate the possibility of berberine as a sensitizer to improve the efficacy of sorafenib against HCC and investigate the mechanisms involved. As demonstrated from our results, we found berberine greatly enhanced the inhibitory effects of sorafenib on cell proliferation synergistically against mouse and human liver cancer cell lines in vitro, Hepa1-6 xenograft model in C57/B6J mice in vivo and in the 3D culture system of tumor cells. Compared with sorafenib treatment alone, additional berberine treatment induced more severe mitochondrial dysfunction and ROS accumulation, following exacerbation of lipid peroxidation, which were abolished by ferroptosis inhibitors liproxstatin-1 and ferrostatin-1. In HepG2 cells, we validated ferroptosis induction in combined treatment is p53-dependent by RNA sequencing. Herein, we demonstrated p53 signaling activation is due to p53 protein accumulation without its upregulation transcriptionally. Mechanically, we found p53 protein accumulation is not due to ubiquitin-dependent proteasomal degradation but resulting from repression of NQO1-dependent ubiquitin-independent proteasomal degradation. Additionally, we demonstrated NQO1 activation induced by combined treatment was due to SETDB1 inactivation, following ERVs induction and genome instability, which may be another factor for lipid peroxidation and ferroptosis. Collectively, berberine has great potential to be used as a novel chemo-enhancer to improve the efficacy of sorafenib -based therapy.