Zhang Gong, Wang Yufeng, Fuchs Bryan C, Guo Wei, Drum David L, Erstad Derek J, Shi Baomin, DeLeo Albert B, Zheng Hui, Cai Lei, Zhang Liyuan, Tanabe Kenneth K, Wang Xinhui
Division of Gastrointestinal and Oncologic Surgery, Department of Surgery, Massachusetts General Hospital, Harvard Medical School, Boston, MA, United States.
Department of Hepatobiliary and Pancreatic Surgery, The First Affiliated Hospital of Zhengzhou University, Zhengzhou, Henan, China.
Front Oncol. 2022 Jul 14;12:913736. doi: 10.3389/fonc.2022.913736. eCollection 2022.
Sorafenib, a kinase inhibitor, is a standard treatment for advanced hepatocellular carcinoma (HCC) but provides only a limited survival benefit. Disulfiram (DSF), a drug for treating alcoholism and a chelator of copper (Cu), forms a complex with Cu (DSF/Cu). DSF/Cu is a potent inducer of autophagic apoptosis of cancer stem cells, which can demonstrate drug resistance. Thus, we hypothesized that DSF/Cu could increase the sensitivity of HCC cells to sorafenib by targeting hepatic cancer stem cells.
The synergistic effect of DSF/Cu and sorafenib on human HCC cell lines was assessed by cell viability MTT assay. Changes in stemness gene expression in HCC cells were investigated by assessing the presence of hepatic cancer stem cells (HCSCs) (defined as ALDH cells) using flow cytometry, sphere formation ability as an index of tumorigenicity, and expression of stemness gene-encoded proteins by western blot. Autophagic apoptosis and the ERK signaling pathway were also assessed by western blot. Most importantly, the anti-tumor efficacy of DSF/Cu and sorafenib was tested using orthotopic HCC xenografts in mice.
Compared with sorafenib alone, DSF/Cu + sorafenib synergistically inhibited proliferation of all HCC cell lines, decreased the stemness of HCC cells, and increased the autophagy and apoptosis of HCC cells. The mechanism by which DSF/Cu mediated these phenomena with sorafenib was sustained activation of the ERK pathway. The combination of DSF/Cu (formed with endogenous Cu) and sorafenib was significantly more effective than sorafenib alone in inhibiting the growth of orthotopic HCC xenografts in mice. This anti-tumor efficacy was associated with decreased stemness in treated HCC tumors.
DSF/Cu and sorafenib can synergistically and effectively treat HCC by targeting HCSCs and . Our data provide a foundation for clinical translation.
索拉非尼是一种激酶抑制剂，是晚期肝细胞癌（HCC）的标准治疗药物，但仅能提供有限的生存获益。双硫仑（DSF）是一种治疗酒精中毒的药物以及铜（Cu）螯合剂，可与Cu形成复合物（DSF/Cu）。DSF/Cu是癌症干细胞自噬性凋亡的强效诱导剂，癌症干细胞可表现出耐药性。因此，我们推测DSF/Cu可通过靶向肝癌干细胞增加HCC细胞对索拉非尼的敏感性。
通过细胞活力MTT试验评估DSF/Cu与索拉非尼对人HCC细胞系的协同作用。通过使用流式细胞术评估肝癌干细胞（定义为乙醛脱氢酶阳性细胞）的存在、以成球能力作为致瘤性指标以及通过蛋白质印迹法检测干性基因编码蛋白的表达，来研究HCC细胞中干性基因表达的变化。还通过蛋白质印迹法评估自噬性凋亡和ERK信号通路。最重要的是，使用原位HCC异种移植小鼠模型测试DSF/Cu与索拉非尼的抗肿瘤疗效。
与单独使用索拉非尼相比，DSF/Cu联合索拉非尼可协同抑制所有HCC细胞系的增殖，降低HCC细胞的干性，并增加HCC细胞的自噬和凋亡。DSF/Cu与索拉非尼介导这些现象的机制是ERK通路的持续激活。DSF/Cu（与内源性Cu形成）与索拉非尼联合在抑制小鼠原位HCC异种移植瘤生长方面明显比单独使用索拉非尼更有效。这种抗肿瘤疗效与治疗的HCC肿瘤中干性降低有关。
DSF/Cu与索拉非尼可通过靶向肝癌干细胞协同有效地治疗HCC。我们的数据为临床转化提供了基础。
