Wang Yuan-Chun, Hui Jian-Rong, Xiao Gang, Ma Qiao-Lin
The First Clinical Medical College, Nanjing University of Chinese Medicine, Nanjing, 210023, China.
Affiliated Hospital of Shaanxi University of Chinese Medicine, Xianyang, Shaanxi Province, 712000, China.
Chin J Integr Med. 2023 Mar;29(3):244-252. doi: 10.1007/s11655-022-3681-x. Epub 2022 Aug 31.
To investigate the mechanism of the effect of Astragalus membranaceus (A. membranaceus) on lung adenocarcinoma at the molecular level to elucidate the specific targets according to the network pharmacology approach.
The active components of A. membranaceus and their potential targets were collected from the Traditional Chinese Medicine Systems Pharmacology Database. Lung adenocarcinoma-associated genes were acquired based on GeneCards, Online Mendelian Inheritance in Man (OMIM), PharmGKB, and Therapeutic Targets databases. The PI3K/AKT signaling pathway-related genes were obtained using Reactome portal. Networks of "ingredient-target" and "ingredient-target-pathway-disease" were constructed using the Cytoscape3.6.0 software. The relationships among targets were analyzed according protein-protein interaction (PPI) network. Finally, molecular docking was applied to construct the binding conformation between active ingredients and core targets. Cell counting kit 8 (CCK8) and Western blot assays were performed to determine the mechanism of the key ingredient of A. membranaceus.
A total of 20 active components and their 329 targets, and 7,501 lung adenocarcinoma-related genes and 130 PI3K/AKT signaling pathway-related genes were obtained. According to Venn diagram and PPI network analysis, 2 mainly active ingredients, including kaempferol and quercetin, and 6 core targets, including TP53, MAPK1, EGF, AKT1, ERBB2, and EGFR, were identified. The two important active ingredients of A. membranaceus, kaempferol and quercetin, exert the therapeutic effect in lung adenocarcinoma partly by acting on the 6 core targets (TP53, MAPK1, EGF, AKT1, ERBB2, and EGFR) of PI3K/AKT signaling pathway. Expressions of potential targets in lung adenocarcinoma and normal samples were analyzed by using UALCAN portal and found that ERBB2 was overexpressed in lung adenocarcinoma tissues and upregulation of it correlated with clinicopathological characteristics. Finally, quercetin repressed viabilities of lung adenocarcinoma cells by targeting ERBB2 on PI3K/AKT signaling confirmed by CCK8 and Western blot.
Our finding unraveled that an active ingredient of A. membranaceus, quercetin, significantly inhibited the lung adenocarcinoma cells proliferation by repressing ERBB2 level and inactivating the PI3K/AKT signaling pathway.
从分子水平探讨黄芪对肺腺癌作用的机制，依据网络药理学方法阐明其具体靶点。
从中药系统药理学数据库收集黄芪的活性成分及其潜在靶点。基于基因卡片、人类孟德尔遗传在线数据库（OMIM）、药物基因组知识库（PharmGKB）和治疗靶点数据库获取肺腺癌相关基因。使用Reactome数据库获取PI3K/AKT信号通路相关基因。利用Cytoscape3.6.0软件构建“成分-靶点”和“成分-靶点-通路-疾病”网络。根据蛋白质-蛋白质相互作用（PPI）网络分析靶点之间的关系。最后，应用分子对接构建活性成分与核心靶点之间的结合构象。采用细胞计数试剂盒8（CCK8）和蛋白质免疫印迹法检测黄芪关键成分的作用机制。
共获得20种活性成分及其329个靶点，以及7501个肺腺癌相关基因和130个PI3K/AKT信号通路相关基因。通过韦恩图和PPI网络分析，确定了2种主要活性成分，即山奈酚和槲皮素，以及6个核心靶点，包括TP53、MAPK1、表皮生长因子（EGF）、AKT1、人表皮生长因子受体2（ERBB2）和表皮生长因子受体（EGFR）。黄芪的两种重要活性成分山奈酚和槲皮素部分通过作用于PI3K/AKT信号通路的6个核心靶点（TP53、MAPK1、EGF、AKT1、ERBB2和EGFR）发挥对肺腺癌的治疗作用。利用UALCAN数据库分析肺腺癌和正常样本中潜在靶点的表达，发现ERBB2在肺腺癌组织中过表达，其上调与临床病理特征相关。最后，CCK8和蛋白质免疫印迹法证实槲皮素通过靶向PI3K/AKT信号通路中的ERBB2抑制肺腺癌细胞活力。
我们的研究发现黄芪活性成分槲皮素通过抑制ERBB2水平和使PI3K/AKT信号通路失活，显著抑制肺腺癌细胞增殖。
