基于细胞实验及网络药理学分析桂枝改善非小细胞肺癌耐药的分子机制  

Molecular Mechanism Analysis of Cinnamomi Ramulus in Improving Drug Resistance of Non-small Cell Lung Cancer Using Cell Experiments and Network Pharmacology

作　　者：徐佳禛 吴加元 王涛 张慧 姜宁华 XU Jiazhen;WU Jiayuan;WANG Tao;ZHANG Hui;JIANG Ninghua(Department of Pharmacy,The Second Affiliated Hospital of Jiaxing University,Jiaxing 314000,China;Key Laboratory of Clinical in Vitro Diagnostic Techniques of Zhejiang Province,Hangzhou 310030,China;Dian Diagnostics Group Co.,Ltd.,Hangzhou 310030,China)

机构地区：[1]嘉兴市第二医院药学部,浙江嘉兴314000 [2]浙江省数字医学诊断技术重点实验室,杭州310030 [3]杭州迪安医学检验中心有限公司,杭州310030

出　　处：《中国现代应用药学》2025年第1期34-42,共9页Chinese Journal of Modern Applied Pharmacy

基　　金：浙江省中医药科技计划项目(2021ZB295)。

摘　　要：目的研究中药桂枝对非小细胞肺癌(non-small cell lung cancer,NSCLC)耐药细胞株H1650增殖、转移、凋亡的影响,揭示桂枝改善H1650吉非替尼耐药性的功能及潜在分子机制。方法在H1650中,使用桂枝、吉非替尼单药以及桂枝与吉非替尼联合用药,利用MTT法、细胞划痕法、流式细胞术分别检测H1650细胞的增殖、迁移、凋亡情况。随后,利用网络药理学预测桂枝的有效活性成分及其影响NSCLC细胞功能的可能分子靶点,构建“桂枝-活性成分-靶点-疾病”的关系网络,并对桂枝潜在作用分子靶点进行GO功能分析和KEGG信号通路富集分析,预测与桂枝活性成分密切相关的核心分子及信号通路。最后,利用Western blotting检测预测得到的核心分子蛋白质表达及蛋白质磷酸化修饰水平在桂枝、吉非替尼作用后的变化情况。结果桂枝单药可抑制H1650细胞增殖、迁移,促进细胞凋亡;与吉非替尼单药组相比,桂枝联合吉非替尼可降低吉非替尼抑制H1650细胞增殖的IC_(50)。网络药理学预测得到7个桂枝活性成分、252个NSCLC疾病相关分子靶点及多条与肿瘤发生发展密切相关的信号通路。其中,Src、PI3K、MAPK1及其相关信号通路可能是桂枝影响NCSLC细胞功能及耐药性的核心分子靶点。分子水平上,桂枝降低了H1650细胞Src蛋白质表达水平以及Src信号通路下游分子——Akt和mTOR的磷酸化水平。结论桂枝可通过抑制Src/Akt/mTOR信号通路,改善H1650细胞对吉非替尼的耐药现象。OBJECTIVE To investigate the effect of traditional Chinese medicine Cinnamomi Ramulus on the proliferation,metastasis,and apoptosis of drug-resistant cell line H1650 in non-small cell lung cancer(NSCLC),and reveal the function and potential molecular mechanism of Cinnamomi Ramulus in improving the resistance of H1650 to gefitinib.METHODS H1650 cells were treated with Cinnamomi Ramulus extract,gefitinib,or two in combination.Using MTT,wound healing assay,and flow cytometry to study H1650 cells’effect on cell viability,migration,and apoptosis.Then,network pharmacology was used to predict the effective active components of Cinnamomi Ramulus extract and the possible molecular targets relating to NSCLC,and to conduct the relationship network of"Cinnamomi Ramulus-active ingredient-targets-disease".GO and KEGG analysis were subsequently performed on the potential molecular targets of Cinnamomi Ramulus,and the core targets and signaling pathways that are closely related to the active components of Cinnamomi Ramulus are further predicted.Finally,Western blotting was used to detect the changes in protein expression and protein phosphorylation level of the predicted core molecules after treatment with Cinnamomi Ramulus extract and gefitinib.RESULTS In H1650,Cinnamomi Ramulus extract inhibited cell viability and migration and prompted cell apoptosis.Compared with the single-drug group with gefitinib,those two drugs in combination reduced the IC_(50) of gefitinib inhibiting cell proliferation.Network pharmacology predicted 7 active components,252 molecular targets related to NSCLC disease,and a number of signaling pathways closely related to the initiation and development of tumors.In particular,Src,PI3K,MAPK1,and their related signaling pathways might be the core molecular targets affecting NCSLC cell function and drug resistance.At the molecular level,Cinnamomi Ramulus extract significantly reduced the protein expression of Src and the downstream molecules in the Src signaling pathway(phosphorylation level of Akt and mTOR

关 键 词：桂枝 非小细胞肺癌 网络药理学 表皮生长因子受体-酪氨酸激酶抑制剂 SRC 

分 类 号：R966[医药卫生—药理学]