基于网络药理学探讨槲皮素靶向NOX4抗非小细胞肺癌的作用机制OA

[Objective]To explore the potential targets of quercetin against non-small cell lung cancer(NSCLC)based on network pharmacology,and to validate its effects on the proliferation,migration and invasion of NSCLC cells.[Methods]The Swiss Target Prediction and Similarity Ensemble Approach(SEA)databases were used to screen potential targets of quercetin.The NSCLC-related targets were obtained from The Human Gene Database(GeneCards).Transcriptomic data of lung cancer and adjacent tissues from the Gene Expression Omnibus(GEO)database were integrated to identify common targets between quercetin and NSCLC,and construct Venn diagram.The enrichment analyses,including gene ontology(GO)and Kyoto encyclopedia of genes and genomes(KEGG)pathway analyses,were performed by using the Metascape and SangerBox platforms.Molecular docking was conducted by using AutoDock Vina 1.5.7 software,and molecular dynamics(MD)simulations were employed to assess the binding stability of quercetin with the target protein.The Tumor Immune Estimation Resource(TIMER)and Gene Expression Profiling Interactive Analysis 2(GEPIA2)databases were utilized to analyze the expression characteristics and prognostic relevance of key target proteins in lung cancer patients.Western blot and immunofluorescence were used to detect the effects of quercetin on target protein expression and the relevant signaling pathway.Cell counting kit-8(CCK-8)and colony formation assays were performed to evaluate the effect of quercetin on NSCLC cell proliferation,while scratch wound healing and Transwell assays were used to assess cell migration and invasion capabilities.[Results]Network pharmacology and bioinformatics analyses revealed 26 common potential targets associated with quercetin and lung cancer.Swiss Target Prediction database analysis indicated that NADPH oxidase 4(NOX4)was the most likely potential target of quercetin.The KEGG and GO enrichment analyses suggested that quercetin may regulate NSCLC through phosphatidylinositol 3-kinase-protein kinase B(PI3K-AKT)signaling pathway.Molecular docking and molecular dynamics simulations confirmed that quercetin stably binded to the NOX4 protein.Western blot demonstrated that NOX4 was highly expressed in lung cancer tissues and was significantly associated with poor prognosis in NSCLC patients,along with bioinformatic analysis.Cellular experiments further demonstrated that quercetin significantly downregulated NOX4 protein expression in H1975 and PC9 cells.Compared with control group,both low and high-dose quercetin treatments significantly inhibited the phosphorylation levels of key proteins PI3K and AKT in PI3K-AKT signaling pathway and effectively reduced cell proliferation,migration and invasion.[Conclusion]Quercetin suppresses the malignant biological behaviors of NSCLC cells by targeted downregulating NOX4 expression and inhibiting the activation of PI3K-AKT signaling pathway.