Wei Tingting, Cheng Jiajie, Ji Yonggan, Cao Xue, Ding Shuqin, Liu Quanxia, Wang Zhisheng
Department of Oncology, General Hospital of Ningxia Medical University, Ningxia Hui Autonomous Region, Yinchuan, 750001, P.R. China.
School of Pharmacy, Ningxia Medical University, Ningxia Hui Autonomous Region, Yinchuan, 750004, P. R. China.
Open Life Sci. 2024 Jul 29;19(1):20220914. doi: 10.1515/biol-2022-0914. eCollection 2024.
Hepatocellular carcinoma (HCC) is a highly vascularized carcinoma, and targeting its neovascularization represents an effective therapeutic approach. Our previous study demonstrated that the baculovirus-mediated endostatin and angiostatin fusion protein (BDS-hEA) effectively inhibits the angiogenesis of vascular endothelial cells and the growth of HCC tumors. However, the mechanism underlying its anti-angiogenic effect remains unclear. Increasing evidence suggests that autophagy has a significant impact on the function of vascular endothelial cells and response to cancer therapy. Hence, the objective of this research was to investigate the correlation between BDS-hEA-induced angiogenesis inhibition and autophagy, along with potential regulatory mechanisms. Our results demonstrated that BDS-hEA induced autophagy in EA.hy926 cells, as evidenced by the increasing number of autophagosomes and reactive oxygen species, accompanied by an upregulation of Beclin-1, LC3-II/LC3-I, and p62 protein expression. Suppression of autophagy using 3-methyladenine attenuated the functions of BDS-hEA-induced EA.hy926 cells, including the viability, proliferation, invasion, migration, and angiogenesis. Moreover, BDS-hEA induced autophagy by downregulating the expression of CD31, VEGF, and VEGFR2, as well as phosphorylated protein kinase B (p-AKT) and phosphorylated mammalian target of rapamycin (p-mTOR), while concurrently upregulating phosphorylated AMP-activated protein kinase (p-AMPK). The results further indicated that inhibition of autophagy by chloroquine significantly impeded the ability of BDS-hEA to suppress HCC tumor growth in mice. Mechanistically, BDS-hEA prominently facilitated autophagic apoptosis in tumor tissues and decreased the levels of ki67, CD31, VEGF, MMP-9, p-AKT, and p-mTOR while simultaneously enhancing the p-AMPK expression. In conclusion, our findings suggest that BDS-hEA induces autophagy as a cytotoxic response by modulating the AMPK/AKT/mTOR signaling pathway, thereby exerting anti-angiogenic effects against HCC.
肝细胞癌（HCC）是一种血管高度丰富的癌症，针对其新生血管形成进行靶向治疗是一种有效的治疗方法。我们之前的研究表明，杆状病毒介导的内皮抑素和血管抑素融合蛋白（BDS-hEA）可有效抑制血管内皮细胞的血管生成以及HCC肿瘤的生长。然而，其抗血管生成作用的潜在机制仍不清楚。越来越多的证据表明，自噬对血管内皮细胞的功能和癌症治疗反应具有重大影响。因此，本研究的目的是探讨BDS-hEA诱导的血管生成抑制与自噬之间的相关性以及潜在的调控机制。我们的结果表明，BDS-hEA在EA.hy926细胞中诱导自噬，自噬体和活性氧数量增加证明了这一点，同时伴随着Beclin-1、LC3-II/LC3-I和p62蛋白表达的上调。使用3-甲基腺嘌呤抑制自噬减弱了BDS-hEA诱导的EA.hy926细胞的功能，包括活力、增殖、侵袭、迁移和血管生成。此外，BDS-hEA通过下调CD31、VEGF和VEGFR2的表达，以及磷酸化蛋白激酶B（p-AKT）和磷酸化雷帕霉素哺乳动物靶蛋白（p-mTOR），同时上调磷酸化AMP激活蛋白激酶（p-AMPK）来诱导自噬。结果进一步表明，氯喹抑制自噬显著阻碍了BDS-hEA抑制小鼠HCC肿瘤生长的能力。从机制上讲，BDS-hEA显著促进肿瘤组织中的自噬性凋亡，并降低ki67、CD31、VEGF、MMP-9、p-AKT和p-mTOR的水平，同时增强p-AMPK的表达。总之，我们的研究结果表明，BDS-hEA通过调节AMPK/AKT/mTOR信号通路诱导自噬作为一种细胞毒性反应，从而对HCC发挥抗血管生成作用。
