网络药理学与实验验证探讨六味地黄汤在前列腺癌中的作用及潜在机制。
Network pharmacology and experimental validation to explore the role and potential mechanism of Liuwei Dihuang Decoction in prostate cancer.
机构信息
Urology Center, Shuguang Hospital Affiliated to Shanghai University of Traditional Chinese Medicine, Shanghai, 201203, China.
Surgical Institute of Integrative Medicine, Shuguang Hospital Affiliated to Shanghai University of Traditional Chinese Medicine, Shanghai, 201203, China.
出版信息
BMC Complement Med Ther. 2024 Jul 26;24(1):284. doi: 10.1186/s12906-024-04572-5.
OBJECTIVE
To evaluate the anti-tumor effector of Liuwei Dihuang Decoction (LWDHD) in prostate cancer (PCa) and explore the potential mechanism using experimental validation, network pharmacology, bioinformatics analysis, and molecular docking.
METHODS
CCK test, Clone formation assay and wound-healing assays were used to determine the effect of LWDHD on prostate cancer growth and metastasis. The active ingredients and targets of LWDHD were obtained from the TCMSP database, and the relevant targets were selected by GeneCards, OMIM and DisGeNET databases for PCa. The cross-targets of drugs and disease were imported into the STRING database to construct protein interactions. The network was also visualized using Cytoscape software and core targets are screened using the Network Analyzer plug-in. The Gene Ontology (GO) and Kyoto Encyclopedia of Genes and Genomes (KEGG) pathway enrichment were analyzed using R software. TCGA database was used to analyze the correlation of bioinformatics genes. AutoDock vina was used to predict the molecular docking and binding ability of active ingredients to key targets. Through WB and q-PCR experiments, the above gene targets were detected to verify the effect of LWDHD on PCa.
RESULTS
CCK and scratch tests confirmed that LWDHD could inhibit the proliferation, invasion and migration of prostate cancer cells. Clone formation experiments showed that LWDHD inhibited the long-term proliferative capacity of PC3 cells. LWDHD and PCa had a total of 99 common targets, establishing a "drug-ingredient-common target" network. Through GO and KEGG enrichment analysis, PI3K/AKT, MAPK, TP53 pathway, MYC, TNF pathway and other signaling pathways were found. Bioinformatics analysis showed that MYC gene was highly expressed and CCND1 and MAPK1 were low expressed in prostate cancer tissues. In addition, TP53, AKT1, MYC, TNF and CCND1 were positively correlated with MAPK1, among which AKT1 and CCND1 were most closely correlated with MAPK1. Molecular docking results showed that quercetin, kaempferol, β-sitosterol and other main active ingredients of LWDHD treatment for PCa were combined with core proteins MAPK1 and AKT1 well. WB and q-PCR results showed that LWDHD inhibited the expression of PI3K and AKT in PC3 cells.
CONCLUSION
The mechanism of LWDHD therapy for PCa is a multi-target and multi-pathway complex process, which may be related to the biological processes mediated by MAPK1 and AKT1 pathways, such as cell proliferation and inhibition of metastasis, and the regulation of signaling pathways. The PI3K/AKT signaling pathway may be a central pathway of LWDHD to inhibit prostate cancer proliferation.
目的
通过实验验证、网络药理学、生物信息学分析和分子对接，评估六味地黄汤（LWDHD）在前列腺癌（PCa）中的抗肿瘤作用，并探讨其潜在机制。
方法
用 CCK 试验、克隆形成试验和划痕试验检测 LWDHD 对前列腺癌生长和转移的影响。从 TCMSP 数据库中获得 LWDHD 的活性成分和靶点，通过 GeneCards、OMIM 和 DisGeNET 数据库选择与 PCa 相关的靶点。将药物和疾病的交叉靶点导入 STRING 数据库，构建蛋白质相互作用网络。使用 Cytoscape 软件可视化网络，并使用 Network Analyzer 插件筛选核心靶点。使用 R 软件对基因本体（GO）和京都基因与基因组百科全书（KEGG）通路进行富集分析。使用 TCGA 数据库分析生物信息学基因的相关性。使用 AutoDock vina 预测活性成分与关键靶标的分子对接和结合能力。通过 WB 和 q-PCR 实验检测上述基因靶标，验证 LWDHD 对 PCa 的作用。
结果
CCK 和划痕试验证实 LWDHD 可抑制前列腺癌细胞的增殖、侵袭和迁移。克隆形成实验表明 LWDHD 抑制了 PC3 细胞的长期增殖能力。LWDHD 和 PCa 共有 99 个共同靶点，建立了“药物-成分-共同靶点”网络。通过 GO 和 KEGG 富集分析，发现了 PI3K/AKT、MAPK、TP53 通路、MYC、TNF 通路等信号通路。生物信息学分析表明，前列腺癌组织中 MYC 基因高表达，CCND1 和 MAPK1 低表达。此外，TP53、AKT1、MYC、TNF 和 CCND1 与 MAPK1 呈正相关，其中 AKT1 和 CCND1 与 MAPK1 相关性最强。分子对接结果表明，LWDHD 治疗 PCa 的主要活性成分槲皮素、山柰酚、β-谷甾醇等与核心蛋白 MAPK1 和 AKT1 结合良好。WB 和 q-PCR 结果表明，LWDHD 抑制了 PC3 细胞中 PI3K 和 AKT 的表达。
结论
LWDHD 治疗 PCa 的机制是一个多靶点、多通路的复杂过程，可能与 MAPK1 和 AKT1 通路介导的细胞增殖和抑制转移等生物学过程以及信号通路的调节有关。PI3K/AKT 信号通路可能是 LWDHD 抑制前列腺癌增殖的核心通路。
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