Department of Pediatric Surgery, First Affiliated Hospital of Gannan Medical University, Ganzhou, Jiangxi, China.
Jiangxi Provincial Clinical Research Center for Vascular Anomalies, The First Affiliated Hospital of GanNan Medical University, Ganzhou, Jiangxi, China.
Drug Dev Res. 2024 Feb;85(1):e22140. doi: 10.1002/ddr.22140.
Everolimus, a known inhibitor of the mammalian target of rapamycin (mTOR), has shown uncertain efficacy in treating hepatoblastoma. This study delves into the potential anti-hepatoblastoma properties of everolimus and its intricate relationship with autophagy and ferroptosis, both in vitro and in vivo. In vivo, tumor tissue from hepatoblastoma patient and human hepatoblastoma cell line HuH-6 were xenografted into nude mice to establish xenograft models for observing the effect of everolimus on tumor growth. In vitro, HuH-6 cells were cultured to evaluate the anti-hepatoblastoma activity of everolimus. Transmission electron microscopy and microtubule-associated proteins 1 light chain 3 (LC3), beclin 1, and p62 protein expressions were employed to investigate autophagy. Additionally, indicators of cell apoptosis, reactive oxygen species (ROS) and proteins associated with ferroptosis were measured to evaluate ferroptosis. The results demonstrate that everolimus treatment effectively induced the formation of autophagosomes in hepatoblastoma cells, upregulated the LC3II/I ratio and beclin 1 expression, and downregulated p62 expression, indicating an enhanced autophagy level both in vitro and in vivo. Furthermore, everolimus treatment induced cell apoptosis, increased ROS level, elevated concentrations of malondialdehyde, 4-hydroxynonenal, and iron content, while reducing the ratio of glutathione/oxidized glutathione, and downregulating the protein expression of glutathione peroxidase 4 and solute carrier family 7 member 11, suggesting its ability to induce ferroptosis in hepatoblastoma cells. Importantly, the induction of ferroptosis by everolimus was significantly reversed in the presence of autophinib, an autophagy inhibitor, indicating the autophagy-dependent of everolimus-induced ferroptosis. Taken together, these findings suggest that everolimus holds promise as an effective anti-hepatoblastoma drug, with its mechanism of action potentially involving the induction of autophagy-dependent ferroptosis in hepatoblastoma cells.
依维莫司是一种已知的哺乳动物雷帕霉素靶蛋白（mTOR）抑制剂，在治疗肝母细胞瘤方面疗效不确定。本研究深入探讨了依维莫司在体内和体外的潜在抗肝母细胞瘤特性及其与自噬和铁死亡的复杂关系。在体内，将肝母细胞瘤患者的肿瘤组织和人肝癌细胞系 HuH-6 异种移植到裸鼠中，建立异种移植模型，观察依维莫司对肿瘤生长的影响。在体外，培养 HuH-6 细胞评估依维莫司的抗肝母细胞瘤活性。采用透射电子显微镜和微管相关蛋白 1 轻链 3（LC3）、beclin 1 和 p62 蛋白表达来研究自噬。此外，还测量了细胞凋亡、活性氧（ROS）和与铁死亡相关的蛋白质的指标，以评估铁死亡。结果表明，依维莫司治疗有效诱导肝母细胞瘤细胞自噬体形成，上调 LC3II/I 比值和 beclin 1 表达，下调 p62 表达，表明在体外和体内均增强了自噬水平。此外，依维莫司治疗诱导细胞凋亡，增加 ROS 水平，提高丙二醛、4-羟基壬烯醛和铁含量，同时降低谷胱甘肽/氧化谷胱甘肽的比例，并下调谷胱甘肽过氧化物酶 4 和溶质载体家族 7 成员 11 的蛋白表达，提示其诱导肝母细胞瘤细胞铁死亡的能力。重要的是，在自噬抑制剂 autophinib 存在的情况下，依维莫司诱导的铁死亡明显逆转，表明依维莫司诱导的铁死亡依赖于自噬。综上所述，这些发现表明，依维莫司作为一种有效的抗肝母细胞瘤药物具有潜力，其作用机制可能涉及诱导肝母细胞瘤细胞自噬依赖性铁死亡。
