Center Lab of Longhua Branch and Department of Infectious Disease, Shenzhen People's Hospital (The Second Clinical Medical College, Jinan University; The First Affiliated Hospital, Southern University of Science and Technology), Shenzhen, Guangdong, China.
Department of Biochemistry and Molecular Biology, Medical College of Jinan University, Guangzhou, Guangdong, China.
Autophagy. 2023 Apr;19(4):1221-1238. doi: 10.1080/15548627.2022.2119353. Epub 2022 Sep 13.
Isoginkgetin (ISO), a natural biflavonoid, exhibited cytotoxic activity against several types of cancer cells. However, its effects on hepatocellular carcinoma (HCC) cells and mechanism remain unclear. Here, we revealed that ISO effectively inhibited HCC cell proliferation and migration in vitro. LC3-II expression and autophagosomes were increased under ISO treatment. In addition, ISO-induced cell death was attenuated by treatment with chloroquine or knockdown of autophagy-related genes ( or ). ISO significantly suppressed (solute carrier family 2 member 1) expression and glucose uptake, leading to activation of the AMPK-ULK1 axis in HepG2 cells. Overexpression of SLC2A1/GLUT1 abrogated ISO-induced autophagy. Combining molecular docking with thermal shift analysis, we confirmed that ISO directly bound to the N terminus of CDK6 (cyclin-dependent kinase 6) and promoted its degradation. Overexpression of CDK6 abrogated ISO-induced inhibition of transcription and induction of autophagy. Furthermore, ISO treatment significantly decreased the H3K27ac, H4K8ac and H3K4me1 levels on the enhancer in HepG2 cells. Finally, ISO suppressed the hepatocarcinogenesis in the HepG2 xenograft mice and the diethylnitrosamine+carbon tetrachloride (DEN+CCl)-induced primary HCC mice and we confirmed and as promising oncogenes in HCC by analysis of TCGA data and human HCC tissues. Our results provide a new molecular mechanism by which ISO treatment or deletion promotes autophagy; that is, ISO targeting the N terminus of CDK6 for degradation inhibits the expression of by decreasing the enhancer activity of , resulting in decreased glucose levels and inducing the AMPK-ULK1 pathway.
异银杏素（ISO）是一种天然的双黄酮类化合物，对多种类型的癌细胞表现出细胞毒性作用。然而，其对肝细胞癌（HCC）细胞的作用及其机制尚不清楚。在这里，我们揭示了 ISO 能够有效抑制 HCC 细胞在体外的增殖和迁移。LC3-II 的表达和自噬体在 ISO 处理下增加。此外，用氯喹或敲低自噬相关基因（ 或 ）处理可减弱 ISO 诱导的细胞死亡。ISO 显著抑制 （溶质载体家族 2 成员 1）的表达和葡萄糖摄取，导致 HepG2 细胞中 AMPK-ULK1 轴的激活。SLC2A1/GLUT1 的过表达可消除 ISO 诱导的自噬。通过分子对接结合热移位分析，我们证实 ISO 直接与 CDK6（细胞周期蛋白依赖性激酶 6）的 N 端结合并促进其降解。CDK6 的过表达可消除 ISO 诱导的转录抑制和自噬诱导。此外，ISO 处理显著降低了 HepG2 细胞中 增强子上的 H3K27ac、H4K8ac 和 H3K4me1 水平。最后，ISO 抑制了 HepG2 异种移植小鼠和二乙基亚硝胺+四氯化碳（DEN+CCl）诱导的原发性 HCC 小鼠的肝癌发生，并通过 TCGA 数据和人 HCC 组织分析证实 和 是 HCC 中的潜在致癌基因。我们的结果提供了一个新的分子机制，即 ISO 处理或缺失通过靶向 CDK6 的 N 端进行降解来促进自噬；即 ISO 通过降低增强子的活性来抑制 的表达，从而降低葡萄糖水平并诱导 AMPK-ULK1 通路。
