The Key Laboratory of Traditional Chinese Medicine Prescription Effect and Clinical Evaluation of State Administration of Traditional Chinese Medicine, School of Pharmacy, Binzhou Medical University, Yantai, Shandong 264003, PR China.
Department of Oncology, 970 Hospital of the PLA Joint Logistic Support Force, Yantai, Shandong 264002, PR China.
Biomed Pharmacother. 2024 Nov;180:117510. doi: 10.1016/j.biopha.2024.117510. Epub 2024 Sep 27.
Morusin (Mor), a prenylated flavonoid isolated from the root bark of Morus alba L., exhibits potent anti-tumour effects; however, the molecular target of Mor is still not entirely clear. This study aimed to elucidate the mechanism of Mor against hepatocellular carcinoma (HCC) and identify potential molecular targets.
Mitochondrial function was assessed by measuring the mitochondrial membrane potential, mitochondrial ultrastructure, oxygen consumption, and ATP levels. Mor-induced mitophagy was confirmed using western blotting, immunofluorescence, and fluorescent probes. Transcriptomics, flow cytometry, western blotting, qRT-PCR and biochemical assays were used to reveal the molecular mechanisms and targets of Mor against HCC. We further validated the interaction between Mor and the target proteins using molecular docking and biolayer interferometry (BLI). The inhibitory effect of Mor in vivo was evaluated using a Hep3B murine xenograft model.
Mor significantly reduced the ATP citrate lyase (ACLY) expression and inhibited ACLY activity in HCC cells. BLI analysis demonstrated a direct interaction between Mor and the ACLY active domain. Mor-induced ACLY inhibition led to ROS accumulation in HCC cells, which caused mitochondrial damage, triggered PINK1/Parkin-mediated mitophagy, and ultimately induced mitochondrial apoptosis. We further verified that ROS is crucial in the apoptotic action of Mor through experiments regarding an ROS scavenger. Mor also significantly inhibited tumour xenograft growth in vivo. In addition, analysis of human liver cancer clinical samples revealed elevated ACLY levels positively correlated with histologic grade.
Collectively, our findings highlight Mor as a potent bioactive inhibitor of ACLY and a promising candidate for HCC therapy.
桑辛素（Mor）是从桑白皮中分离得到的一种具有甲羟戊酸途径活性的类黄酮，具有很强的抗肿瘤作用；然而，Mor 的分子靶点仍不完全清楚。本研究旨在阐明 Mor 对肝癌（HCC）的作用机制，并确定潜在的分子靶点。
通过测量线粒体膜电位、线粒体超微结构、耗氧量和 ATP 水平来评估线粒体功能。使用 Western blot、免疫荧光和荧光探针证实 Mor 诱导的自噬。使用转录组学、流式细胞术、Western blot、qRT-PCR 和生化分析揭示 Mor 对 HCC 的分子机制和靶点。我们进一步使用分子对接和生物层干涉（BLI）验证了 Mor 与靶蛋白之间的相互作用。使用 Hep3B 鼠异种移植模型评估 Mor 在体内的抑制作用。
Mor 显著降低了肝癌细胞中 ATP 柠檬酸裂解酶（ACLY）的表达和 ACLY 活性。BLI 分析表明 Mor 与 ACLY 活性结构域直接相互作用。Mor 诱导的 ACLY 抑制导致 HCC 细胞中 ROS 积累，引起线粒体损伤，触发 PINK1/Parkin 介导的自噬，最终诱导线粒体凋亡。我们通过 ROS 清除剂实验进一步验证了 ROS 在 Mor 诱导的凋亡作用中的关键作用。Mor 还显著抑制了体内肿瘤异种移植的生长。此外，对人类肝癌临床样本的分析显示，ACLY 水平升高与组织学分级呈正相关。
总之，我们的研究结果强调了 Mor 作为 ACLY 有效生物抑制剂的潜力，是 HCC 治疗的有前途的候选药物。
