Zheng Bo-Xin, Long Wei, Zeng Yao-Xun, She Meng-Ting, Zheng Yingying, Zheng Wen-De, Wang Ya-Kun, Chan Ka-Hin, Leung Alan Siu-Lun, Chan Chun-Ming, Lu Yu-Jing, Wong Wing-Leung
Department of Applied Biology and Chemical Technology, The Hong Kong Polytechnic University, Hong Kong SAR, China.
The Hong Kong Polytechnic University Shenzhen Research Institute, Shenzhen, China.
Br J Pharmacol. 2025 Aug;182(16):3923-3951. doi: 10.1111/bph.70061. Epub 2025 May 9.
Regulation of mitochondrial calcium overload and ferroptosis with mitochondria-targeting ligands is an attractive anticancer strategy but it remains a challenge. The aim of the present study was to demonstrate that a mitochondria-targeting and mtDNA G-quadruplex-binding ligand, BYB, induced mitochondrial calcium overload and ferroptosis in HeLa cells and showed potent in vitro and in vivo anticancer activity.
Cellular functions and molecular mechanism were studied using cell viability assay, live-cell imaging, western blotting, immunofluorescence, cell uptake, cell cycle arrest and apoptosis analysis, mitochondrial metabolism analysis, Comet assay, and wound-healing analysis. Pharmacokinetic studies were conducted in rat. In vivo antitumor activity was studied in a cervical cancer HeLa cell xenograft mouse model.
Cellular results showed that BYB induced mitochondrial calcium overload, attributed to ligand-induced mitochondrial dysfunction via the mechanism of inhibiting mitochondrial DNA replication and transcription. The expression of respiratory chain complexes was markedly downregulated in BYB-treated HeLa cells. The respiratory chain function was also dysregulated. Mitophagy and mitochondrial calcium overload were induced in BYB-treated HeLa cells. Mitochondrial calcium overload markedly induced mtROS production. The induced mtDNA stress activated cGAS-STING pathway, leading to autophagy-dependent ferroptosis. The antitumour efficacy of BYB, evaluated in a HeLa tumour xenograft mouse model, achieved over 60% tumour weight reduction.
BYB, via targeting mitochondria and mtDNA G-quadruplexes, induced mitochondrial calcium overload and ferroptosis, exhibited high in vivo antitumour efficacy and low toxicity. It shows high potential to be a mitochondria-targeting lead compound for chemical biology and drug discovery.
利用线粒体靶向配体调节线粒体钙超载和铁死亡是一种颇具吸引力的抗癌策略，但仍面临挑战。本研究旨在证明一种线粒体靶向且能结合线粒体DNA G-四链体的配体BYB可诱导HeLa细胞发生线粒体钙超载和铁死亡，并展现出强大的体外和体内抗癌活性。
采用细胞活力测定、活细胞成像、蛋白质印迹法、免疫荧光、细胞摄取、细胞周期阻滞和凋亡分析、线粒体代谢分析、彗星试验和伤口愈合分析等方法研究细胞功能和分子机制。在大鼠中进行药代动力学研究。在宫颈癌HeLa细胞异种移植小鼠模型中研究体内抗肿瘤活性。
细胞实验结果表明，BYB诱导线粒体钙超载，这归因于配体通过抑制线粒体DNA复制和转录的机制诱导线粒体功能障碍。在经BYB处理的HeLa细胞中，呼吸链复合物的表达明显下调。呼吸链功能也失调。经BYB处理的HeLa细胞中诱导了线粒体自噬和线粒体钙超载。线粒体钙超载显著诱导线粒体活性氧的产生。诱导的线粒体DNA应激激活了cGAS-STING通路，导致自噬依赖性铁死亡。在HeLa肿瘤异种移植小鼠模型中评估的BYB抗肿瘤疗效使肿瘤重量减轻超过60%。
BYB通过靶向线粒体和线粒体DNA G-四链体，诱导线粒体钙超载和铁死亡，具有高体内抗肿瘤疗效和低毒性。它显示出作为化学生物学和药物发现中线粒体靶向先导化合物的巨大潜力。
