Integrating Network Pharmacology and Bioinformatics to Explore the Effects of Dangshen (Codonopsis pilosula) Against Hepatocellular Carcinoma: Validation Based on the Active Compound Luteolin.

This study aimed to explore the pharmacological mechanism of Dangshen (Codonopsis pilosula) against hepatocellular carcinoma (HCC) based on network pharmacology and bioinformatics, and to verify the anticancer effect of luteolin, the active ingredient of Codonopsis pilosula, on HCC cells. The effective compounds and potential targets of Codonopsis pilosula were established using the Traditional Chinese Medicine Systems Pharmacology Database and Analysis Platform (TCMSP) database. The genes related to HCC were obtained through the GeneCards database. The interactive genes were imported into the Visualization and Integrated Discovery database for Gene Ontology (GO) annotation and Kyoto Encyclopedia of Genes and Genomes (KEGG) signal enrichment, and the hub genes were screened out. The Cancer Genome Atlas database was used to construct a prognosis model, and the prognosis and clinicopathological correlation were analyzed. In in vitro experiments, we verified the effects of luteolin, an active compound of Codonopsis pilosula, on the proliferation, cell cycle, apoptosis and migration of HCC cells. A total of 21 effective compounds of Codonopsis pilosula and 98 potential downstream target genes were screened through the TCMSP database, and 1406 HCC target genes were obtained through the GeneCards database. Finally, 53 interacting genes between the two databases were obtained, among which, the 10 key node genes were CASP3, TP53, MDM2, AKT1, ESR1, BCL2L1, MCL1, HSP90AA1, CASP9, and CCND1, involving 77 typical GO terms and 72 KEGG signals. The Kaplan-Meier survival curve of the model group showed that the overall survival of the low-risk group was significantly higher than that of the high-risk group. Luteolin significantly inhibited the proliferation and migration of HCC cells, induced apoptosis, and increased the G2/M phase ratio. Mechanistically, luteolin significantly inhibited the phosphorylation of MAPK-JNK and Akt (Thr308) and subsequently led to upregulation of ESR1. Pharmacological inhibition of ESR1 with fulvestrant enhanced cell viability and migration and attenuated apoptosis. Codonopsis pilosula has potential for clinical development due to its anti-HCC properties. Luteolin, the effective component of Codonopsis pilosula, plays anti-HCC role through AKT- or MAPK-JNK signaling mediated ESR1.

Yu Y ，Ding S ，Xu X ，Yan D ，Fan Y ，Ruan B ，Zhang X ，Zheng L ，Jie W ，Zheng S ... -  《Drug Design Development and Therapy》

Molecular mechanisms of Codonopsis pilosula in inhibiting hepatocellular carcinoma growth and metastasis.

Liver cancer, one of the most common types of cancer worldwide, accounts for millions of cases annually. With its multi-target and wide-ranging therapeutic effects, traditional Chinese medicine has emerged as a potential approach for treating various tumors. Codonopsis pilosula, a traditional herb, is known for its anti-inflammatory and antioxidant properties. In this study, we investigated the potential molecular mechanisms of Codonopsis pilosula in regulating the inhibition of CDK1 and the modulation of PDK1/β-catenin, which are involved in hepatocellular carcinoma growth and metastasis. Firstly, we screened the active chemical constituents of Codonopsis pilosula and identified their respective target proteins using the Herb database. Then, we applied the GeneCards database and transcriptome sequencing analysis to screen for critical genes associated with the occurrence and development of liver cancer. The intersection of the target proteins and disease-related genes was used to determine the potential targets of Codonopsis pilosula in hepatocellular carcinoma. Protein-protein interaction analysis and GO/KEGG analysis were subsequently performed to uncover the pathways through which Codonopsis pilosula acts on liver cancer. The Huh-7 cell line, exhibiting the highest sensitivity to Codonopsis pilosula polysaccharide solution (CPP) intervention, was chosen for subsequent studies. Cell viability was evaluated using the CCK-8 assay, colony formation assay was conducted to determine cell proliferation capacity, flow cytometry was used to analyze cell cycle, TUNEL staining was performed to assess cell apoptosis, scratch assay was carried out to evaluate cell migration ability, the expression of EMT-related proteins was detected and analyzed, and cell sphere formation assay was conducted to investigate cell stemness. Finally, a liver cancer animal model was established, and different doses of CPP were administered via gavage the next day. The expression levels of CDK1, PDK1, and β-catenin in mouse liver tissues were detected and analyzed, immunohistochemistry staining was performed to assess the expression of tumor cell proliferation-related proteins Ki67 and PCNA in mouse xenografts, and TUNEL staining was carried out to evaluate cell apoptosis in mouse liver tissues. After intervention with CDK1 expression, the expression levels of CDK1, PDK1, and β-catenin proteins and mRNA in each group of cells were detected using Western blot and RT-qPCR. Through network pharmacology analysis, transcriptome sequencing, and bioinformatics analysis, 35 target genes through which Codonopsis pilosula acts on liver cancer were identified. Among them, CDK1, with the highest degree in the PPI network, was considered an essential target protein for Codonopsis pilosula in treating liver cancer. In vitro cell experiments revealed that CPP could inhibit the expression of CDK1/PDK1/β-catenin signaling axis factors, suppress cell proliferation, decrease cell migration ability, influence the EMT process, and reduce cell stemness by inhibiting CDK1 and affecting the PDK1/β-catenin signaling axis. Similarly, in vivo experiments demonstrated that CPP could regulate the CDK1/PDK1/β-catenin signaling axis, inhibit tumor growth, and induce cell apoptosis. Codonopsis pilosula may inhibit hepatocellular carcinoma growth by suppressing CDK1 and affecting the PDK1/β-catenin signaling axis, limiting cell EMT and reducing cell stemness. These findings provide insights into the potential therapeutic role of Codonopsis pilosula in liver cancer.

Li N ，Yang C ，Xia J ，Wang W ，Xiong W ... -  《-》

Integration of network pharmacology and molecular docking technology reveals the mechanism of the herbal pairing of Codonopsis Pilosula (Franch.) Nannf and Astragalus Membranaceus (Fisch.) Bge on chronic heart failure.

Xu J ，Zhang Z ，Zhou K ，Li Y ，Wan J ，Mao T ，Ji X ，Liu J ，Lin Q ... -  《-》

Network Pharmacology Integrated with Transcriptomics Deciphered the Potential Mechanism of Codonopsis pilosula against Hepatocellular Carcinoma.

Hepatocellular carcinoma (HCC) is the fourth main reason of cancer-related death. Codonopsis pilosula is a commonly used traditional Chinese medicine (TCM) for patients with HCC. However, its potential mechanism for treatment of HCC remains unclear. Here, we used transcriptomics and network pharmacology to explore the potential molecular mechanisms of Codonopsis pilosula. In our study, twelve differentially expressed genes (DEGs) (5 upregulated and 7 downregulated) of Codonopsis pilosula treating HepG2 cells (a kind of HCC cell) were identified. Among the 12 DEGs, HMOX1 may play an essential role. Codonopsis pilosula mainly affects the mineral absorption pathway in HCC. We acquired 2957, 1877, and 255 targets from TCMID, SymMap, and TCMSP, respectively. Codonopsis pilosula could upregulate HMOX1 via luteolin, capsaicin, and sulforaphane. Our study provided new understanding of the potential pharmacological mechanisms of Codonopsis pilosula in treating HCC and pointed out a direction for further experimental research.

Liu Z ，Sun Y ，Zhen H ，Nie C ... -  《-》

Systems Network Pharmacology-Based Prediction and Analysis of Potential Targets and Pharmacological Mechanism of Actinidia chinensis Planch. Root Extract for Application in Hepatocellular Carcinoma.

Traditional Chinese medicine (TCM) sometimes plays a crucial role in advanced cancer treatment. Despite the significant therapeutic efficacy in hepatocellular carcinoma (HCC) that Actinidia chinensis Planch root extract (acRoots) has proven, its complex composition and underlying mechanism have not been fully elucidated. Therefore, this study analyzed the multiple chemical compounds in acRoots and their targets via network pharmacology and bioinformatics analysis, with the overarching goal of revealing the potential mechanisms of the anti-HCC effect. The main ingredients contained in acRoots were initially screened from the traditional Chinese Medicine Systems Pharmacology Database and Analysis Platform (TCMSP), and the candidate bioactive ingredient targets were identified using DrugBank and the UniProt public databases. Second, the biological processes of the targets of active molecules filtered from the ingredients of acRoots were evaluated using gene ontology (GO) enrichment and Kyoto Encyclopedia of Genes and Genomes (KEGG) pathway analyses. Third, weighted gene coexpression network analysis (WGCNA) was performed to identify gene coexpression modules associated with HCC. The hub genes of acRoots in HCC were defined via contrasting the above module eigengenes with candidate target genes of acRoots. Furthermore, the target-pathway network was analyzed to explore the mechanism for anti-HCC effect of hub genes. Kaplan-Meier plotter database analysis was performed to validate the hub genes of acRoots correlation with prognostic values in HCC. In order to verify the results of the network pharmacological analysis, we performed a molecular docking approach on the active ingredients and key targets using the Discovery Studio software. The viability of SMMC-7721 and HL-7702 cells was determined by Cell counting kit-8 (CCK-8) after being treated with different concentrations of (+)-catechin (0, 50, 100, 150, 200, and 250 g/ml) for 24, 48, and 72 hours, respectively. Finally, qRT-PCR and Western blot involving human hepatocarcinoma cells were utilized to verify the impact of (+)-catechin on the hub genes associated with prognosis. 6 out of 26 active ingredients extracted from TCMSP were deemed as the core ingredients of acRoots. 175 bioactive-ingredient targets of acRoots were obtained and a bioactive-ingredient targets network was established correspondingly. The biological processes (BP) of target genes mainly involved processes, such as toxic substance and wounding. The results of KEGG pathways indicated that the target genes were mainly enriched in pathways in cancer, AGE-RAGE signaling pathway in diabetic complications, IL-17 signaling pathway, and other pathways. Also, the two hub genes (i.e., ESR1 and CAT) were closely associated with the prognosis of HCC patients. As a consequence, we predicated a series of signaling pathways, including estrogen signaling pathway and longevity regulation pathway, through which acRoots could facilitate the treatment for HCC. The molecular docking experiment ascertained that ESR1 and CAT had an effective binding force with (+)-catechin, one of the core ingredients of acRoots. Furthermore, (+)-catechin inhibited SMMC-7721 cell growth in a dose-dependent manner and a time-dependent manner. Finally, we suggest that the expression level of ESR1 and CAT is positively related to the (+)-catechin concentrations in in-vitro experiments. The bioactive ingredients of acRoots, including quercetin, (+)-catechin, beta-sitosterol, and aloe-emodin, have synergistic interactions in reinforcing the anticancer effect in HCC. Evidently, acRoots took effect by regulating multitargets and multipathways through its active ingredients. Further, (+)-catechin, the possible paramount anti-HCC active ingredient in acRoots, helped improve the prognosis of HCC patients by increasing the expression of ESR1 and CAT. Additionally, the findings yielded provide a conceptual guidance for the clinical treatment of HCC and the methods adopted are potentially applicable in the future comprehensive analysis of the underlying mechanisms of TCMs.

Hu Y ，Yang L ，Lu Y ，Wang Y ，Jiang J ，Liu Y ，Cao Q ... -  《-》