Dicoumaral Sensitizes Hepatocellular Carcinoma Cells to Ferroptosis Induced by Imidazole Ketone Erastin

Ferroptosis, an iron-dependent form of regulated cell death, is characterized by the lethal accumulation of lipid peroxides on cellular membranes. It not only inhibits tumor growth but also enhances immunotherapy responses and overcomes drug resistance in cancer therapy. The inhibition of the cystine-glutamate antiporter, system Xc–, induces ferroptosis. Imidazole ketone erastin (IKE), an inhibitor of the system Xc- functional subunit solute carrier family 7 member 11 (SLC7A11), is an effective and metabolically stable inducer of ferroptosis with potential in vivo applications. However, tumor cells exhibited differential sensitivity to IKE-induced ferroptosis. The intrinsic factors determining sensitivity to IKE-induced ferroptosis remain to be explored to improve its efficacy. Bulk RNA-sequencing data from hepatocellular carcinoma (HCC) and normal liver tissues were collected from The Cancer Genome Atlas (TCGA) and the Genotype-Tissue Expression (GTEx) databases. Differentially expressed genes were identified and intersected with the ferroptosis-related genes (FRGs) listed in the FerrDb database, yielding the identification of 13 distinct FRGs. A ferroptosis signature index model (Risk Score) was developed to predict HCC prognosis. And SLC7A11 and NAD(P)H quinone dehydrogenase 1 (NQO1) were identified as candidate FRGs indicating poorprognosisof HCC. Dicoumarol (DIC), an inhibitor of NQO1, was subsequently employed to assess its sensitizing effects on IKE in HCC treatment. In HCC cell lines and the subcutaneous xenograft model, the combined suppression of SLC7A11 and NQO1 significantly enhanced the inhibitory effect on tumor growth by inducing ferroptosis. In conclusion, our findings demonstrate that DIC sensitized HCC cells to IKE-induced ferroptosisin HCC. Moreover, the identification of potential drugs that enhance the susceptibility of HCC cells to ferroptosis could provide novel therapeutic strategies for the treatment of HCC.

铁死亡（Ferroptosis）是一种铁依赖型程序性细胞死亡形式，其特征为细胞膜上发生致死性脂质过氧化物蓄积。该死亡方式既可抑制肿瘤生长，又能增强免疫治疗应答，并克服癌症治疗中的药物耐药性。抑制胱氨酸-谷氨酸反向转运体Xc-系统（system Xc–）可诱导铁死亡。咪唑酮埃拉斯汀（Imidazole ketone erastin, IKE）是Xc-系统功能亚基溶质载体家族7成员11（solute carrier family 7 member 11, SLC7A11）的抑制剂，是一种高效且代谢稳定的铁死亡诱导剂，具备体内应用潜力。然而，肿瘤细胞对IKE诱导的铁死亡表现出差异化敏感性。决定肿瘤细胞对IKE诱导铁死亡敏感性的内在因素仍有待探索，以提升其治疗疗效。本研究从癌症基因组图谱（The Cancer Genome Atlas, TCGA）及基因型-组织表达（Genotype-Tissue Expression, GTEx）数据库中获取了肝细胞癌（hepatocellular carcinoma, HCC）与正常肝组织的批量RNA测序数据。通过鉴定差异表达基因，并与FerrDb数据库收录的铁死亡相关基因（ferroptosis-related genes, FRGs）取交集，最终筛选得到13个特征性铁死亡相关基因。研究人员构建了铁死亡特征指数模型（风险评分模型，Risk Score），用于预测肝细胞癌患者的预后，并鉴定出SLC7A11及烟酰胺腺嘌呤二核苷酸磷酸（NAD(P)H）醌脱氢酶1（NAD(P)H quinone dehydrogenase 1, NQO1）为提示肝细胞癌不良预后的候选铁死亡相关基因。香豆素（Dicoumarol, DIC）作为NQO1的抑制剂，后续被用于评估其在肝细胞癌治疗中对IKE的增敏作用。在肝细胞癌细胞系及皮下异种移植瘤模型中，联合抑制SLC7A11与NQO1可通过诱导铁死亡，显著增强对肿瘤生长的抑制效果。综上，本研究结果表明DIC可使肝细胞癌细胞对IKE诱导的铁死亡产生增敏作用。此外，筛选可增强肝细胞癌细胞铁死亡易感性的潜在药物，可为肝细胞癌的治疗提供全新的治疗策略。