Molecular Assessment of Scutellaria barbata D. Don in the Treatment of Nasopharyngeal Carcinoma Based on Network Pharmacology and Experimental Verification

Abstract

Objective: To predict the molecular mechanisms behind the benefits of Scutellaria barbata D. Don (S. barbata) in nasopharyngeal carcinoma (NPC) by network pharmacology and experimental verification.

Methods: The active ingredients and targets of S. barbata were searched in the traditional Chinese medicine system pharmacology database and analysis platform, and the disease targets of NPC were obtained by searching the GeneCards database. A common target protein-protein interaction network was constructed by STRING, and then, an active ingredients-NPC-target interaction network map was constructed by Cytoscape 3.7.2 software. The functional enrichment analyses of Gene Ontology and KEGG pathway data were carried out by R software programming. Finally, cell proliferation was assessed by CCK8, apoptosis was detected by Annexin V-FITC/PI double fluorescence staining, and protein expression was analyzed by Western blotting.

Results: In this study, 29 active ingredients were found in S. barbata. Among these, the main targets for NPC were baicalein, wogonin, luteolin, and quercetin. The main molecular targets of S. barbata on NPC were EGFR, MYC, CASP3, CCND1, and ESR1. The main biological processes involved the binding of DNA-binding transcription factors, RNA polymerase II-specific DNA-binding transcription factors, ubiquitin-like protein ligases, and ubiquitin-protein ligases. S. barbata mainly affects NPC through the PI3K-Akt, p53, and MAPK signaling pathways. The experimental results showed that baicalein and wogonin could inhibit proliferation and induce apoptosis of NPC cells and downregulate the expression of PI3K, AKT, and p53, the key proteins of the PI3K/AKT and p53 signaling pathway in CNE2 cells.

Conclusion: Baicalein and wogonin, the main active ingredients of S. barbata, inhibited the proliferation and induced apoptosis of NPC cells through the PI3K/AKT and p53 signaling pathways.

Figure 1

Venn diagram of the action targets of active ingredients of S. barbata and the therapeutic targets of NPC.

Figure 2

The protein-protein interaction (PPI) network.

Figure 3

Frequency of common protein targets.

Figure 4

Active ingredients-NPC-target interaction network.

Figure 5

GO analysis bar chart.

Figure 6

GO analysis bubble chart. The node size represents the number of enrichment targets; the color of the node from red to dark blue indicates that the P value is small to large, so the larger the red node, the more significant the signaling pathway, indicating that the signaling pathway is more important. (1) DNA-binding transcription factor binding; (2) RNA polymerase II-specific DNA-binding transcription factor binding; (3) ubiquitin-like protein ligase binding; (4) ubiquitin protein ligase binding; (5) kinase regulator activity; (6) activating transcription factor binding; (7) nuclear hormone receptor binding; (8) hormone receptor binding; (9) cysteine-type endopeptidase activity involved in the apoptotic process; (10) estrogen receptor binding; (11) p53 binding; (12) repressing transcription factor binding; (13) steroid hormone receptor binding; (14) ATPase binding; (15) cysteine-type endopeptidase activity involved in the apoptotic signaling pathway; (16) histone kinase activity; (17) aromatase activity; (18) transcription coactivator binding; (19) NF-kappaB binding; (20) oxidoreductase activity.

Figure 7

Results of KEGG enrichment analysis.

Figure 8

CCK8 assessment of the effect of baicalein on the proliferation of CNE2 and 5-8F cells (vs. the control group: ^∗P < 0.05 and ^∗∗P < 0.01).

Figure 9

CCK8 assessment of the effect of wogonin on the proliferation of CNE2 and 5-8F cells (vs. the control group: ^∗P < 0.05 and ^∗∗P < 0.01).

Figure 10

Annexin V-FITC/PI double fluorescence staining used to detect the effect of baicalein and wogonin on apoptosis rate (vs. the control group: ^∗∗P < 0.01).

Figure 11

The effects of baicalein and wogonin on the expression of key proteins of the PI3K/AKT and p53 signaling pathway in CNE2 cells analyzed by Western blotting (vs. the control group: ^∗P < 0.05 and ^∗∗P < 0.01).