目的 基于数据非依赖型质谱采集（DIA）磷酸化蛋白质组学、网络药理学及分子对接等方法探讨非诺贝特抗肺腺癌的分子机制。方法 非诺贝特处理A549细胞后，进行蛋白质提取、酶解及磷酸化肽段富集，通过液相色谱–质谱进行磷酸化蛋白分析；用R语言limma包筛选激酶类差异表达蛋白，并进行基因本体论（GO）功能注释和京都基因和基因组百科全书（KEGG）通路富集分析；通过STRING数据库构建蛋白相互作用（PPI）网络，并运用Cytoscape可视化软件筛选核心靶点。运用分子对接技术评估非诺贝特与关键靶蛋白之间的结合稳定性。结果 共获得104个差异激酶蛋白，包括上调蛋白48个，下调蛋白56个；GO富集显示差异蛋白主要涉及蛋白质磷酸化、蛋白质修饰过程、肽基–丝氨酸磷酸化等87个生物学过程（P＜0.01)；KEGG通路富集则显示差异蛋白与丝裂原活化蛋白激酶（MAPK）和人表皮生长因子受体（ErbB）等10条信号通路相关。网络药理学分析预测到SRC、MAP3K2、PAK2、LATS1、BRD4、PRKD1、MAP2K2、MAP3K1为非诺贝特抗肺腺癌的8个关键靶蛋白，并且分子对接也证实了非诺贝特与上述靶点结合较为稳定。结论 非诺贝特主要通过抑制MAPK/ErbB通路磷酸化激活而发挥抗肺腺癌作用，而PAK2、LATS1、PRKD1等激酶可能是其抗肿瘤关键靶点，本研究为后续临床研究提供了理论基础。

Objective To investigate the molecular mechanism of fenofibrate against lung adenocarcinoma based on DIA phosphoproteomics, network pharmacology, and molecular docking. Methods After treating A549 cells with fenofibrate, proteins were extracted, enzymatically digested, and phosphorylated peptides were enriched. Phosphorylated proteins were analyzed via liquid chromatography-mass spectrometry (LC-MS). The limma package in R was used to screen differentially expressed kinase proteins, followed by GO functional annotation and KEGG pathway enrichment analysis. PPI network was constructed using the STRING database, and core targets were identified using Cytoscape visualization software. Molecular docking was employed to evaluate the binding stability between fenofibrate and key target proteins. Results A total of 104 differentially expressed kinase proteins were identified, including 48 upregulated and 56 downregulated proteins. GO enrichment revealed that these proteins were primarily involved in 87 biological processes (P < 0.01), such as protein phosphorylation, protein modification processes, and peptidyl-serine phosphorylation. KEGG pathway enrichment indicated associations with 10 signaling pathways, including MAPK and ErbB. Network pharmacology predicted eight key target proteins of fenofibrate against lung adenocarcinoma: SRC, MAP3K2, PAK2, LATS1, BRD4, PRKD1, MAP2K2, and MAP3K1. Molecular docking confirmed stable binding between fenofibrate and these targets. Conclusion Fenofibrate exerts its anti-lung adenocarcinoma effects mainly by inhibiting the phosphorylation activation of the MAPK/ErbB pathway. Kinases such as PAK2, LATS1, and PRKD1 may serve as critical anti-tumor targets, providing a theoretical foundation for subsequent clinical research.

R974;979.1

广东省普通高校特色创新类项目（2022KTSCX305，2024KTSCX292）