State Key Laboratory of Antiviral Drugs, School of Pharmacy, Henan University, Kaifeng 475004, China.
State Key Laboratory for Quality Ensurance and Sustainable Use of Dao-di Herbs, Artemisinin Research Center, and Institute of Chinese Materia Medica, China Academy of Chinese Medical Sciences, Beijing 100700, China.
Phytomedicine. 2024 Jul;129:155657. doi: 10.1016/j.phymed.2024.155657. Epub 2024 Apr 20.
The pentose phosphate pathway (PPP) plays a crucial role in the material and energy metabolism in cancer cells. Targeting 6-phosphogluconate dehydrogenase (6PGD), the rate-limiting enzyme in the PPP metabolic process, to inhibit cellular metabolism is an effective anticancer strategy. In our previous study, we have preliminarily demonstrated that gambogic acid (GA) induced cancer cell death by inhibiting 6PGD and suppressing PPP at the cellular level. However, it is unclear whether GA could suppress cancer cell growth by inhibiting PPP pathway in mouse model.
This study aimed to confirm that GA as a covalent inhibitor of 6PGD protein and to validate that GA suppresses cancer cell growth by inhibiting the PPP pathway in a mouse model.
Cell viability was detected by CCK-8 assays as well as flow cytometry. The protein targets of GA were identified using a chemical probe and activity-based protein profiling (ABPP) technology. The target validation was performed by in-gel fluorescence assay, the Cellular Thermal Shift Assay (CETSA). A lung cancer mouse model was constructed to test the anticancer activity of GA. RNA sequencing was performed to analyze the global effect of GA on gene expression.
The chemical probe of GA exhibited high biological activity in vitro. 6PGD was identified as one of the binding proteins of GA by ABPP. Our findings revealed a direct interaction between GA and 6PGD. We also found that the anti-cancer activity of GA depended on reactive oxygen species (ROS), as evidenced by experiments on cells with 6PGD knocked down. More importantly, GA could effectively reduce the production of the two major metabolites of the PPP in lung tissue and inhibit cancer cell growth in the mouse model. Finally, RNA sequencing data suggested that GA treatment significantly regulated apoptosis and hypoxia-related physiological processes.
These results demonstrated that GA was a covalent inhibitor of 6PGD protein. GA effectively suppressed cancer cell growth by inhibiting the PPP pathway without causing significant side effects in the mouse model. Our study provides in vivo evidence that elucidates the anticancer mechanism of GA, which involves the inhibition of 6PGD and modulation of cellular metabolic processes.
戊糖磷酸途径（PPP）在癌细胞的物质和能量代谢中起着至关重要的作用。靶向 PPP 代谢过程中的限速酶 6-磷酸葡萄糖酸脱氢酶（6PGD）以抑制细胞代谢是一种有效的抗癌策略。在我们之前的研究中，我们已经初步证明，藤黄酸（GA）通过抑制 6PGD 和在细胞水平上抑制 PPP 诱导癌细胞死亡。然而，GA 是否能通过抑制 PPP 途径在小鼠模型中抑制癌细胞生长尚不清楚。
本研究旨在证实 GA 作为 6PGD 蛋白的共价抑制剂，并验证 GA 通过抑制 PPP 途径在小鼠模型中抑制癌细胞生长。
用 CCK-8 法和流式细胞术检测细胞活力。用化学探针和基于活性的蛋白质谱（ABPP）技术鉴定 GA 的蛋白靶标。通过凝胶荧光分析、细胞热转移分析（CETSA）进行靶标验证。构建肺癌小鼠模型以测试 GA 的抗癌活性。进行 RNA 测序以分析 GA 对基因表达的全局影响。
GA 的化学探针在体外具有很高的生物活性。ABPP 鉴定出 6PGD 是 GA 的结合蛋白之一。我们的研究结果表明 GA 与 6PGD 之间存在直接相互作用。我们还发现，GA 的抗癌活性依赖于活性氧（ROS），这可以通过敲低 6PGD 的细胞实验证明。更重要的是，GA 可以有效地降低肺组织中 PPP 的两种主要代谢物的产生，并抑制小鼠模型中的癌细胞生长。最后，RNA 测序数据表明，GA 处理显著调节了细胞凋亡和缺氧相关的生理过程。
这些结果表明，GA 是 6PGD 蛋白的共价抑制剂。GA 通过抑制 PPP 途径有效抑制癌细胞生长，而在小鼠模型中没有引起明显的副作用。我们的研究提供了体内证据，阐明了 GA 的抗癌机制，涉及到抑制 6PGD 和调节细胞代谢过程。
