Network Pharmacology and Experimental Evidence Reveal Dioscin Suppresses Proliferation, Invasion, and EMT via AKT/GSK3b/mTOR Signaling in Lung Adenocarcinoma

Abstract

Purpose

Dioscin, a natural glycoside derived from many plants, has been proved to exert anti-cancer activity. Several studies have found that it reverses TGF-β1-induced epithelial–mesenchymal transition (EMT). Whether dioscin can reverse EMT by pathways other than TGF-β is still unknown.

Methods

We used network-based pharmacological methods to systematically explore the potential mechanisms by which dioscin acts on lung cancer. Cell Counting Kit-8 assay, scratch healing, Transwell assay, Matrigel invasion assay, immunofluorescence assay, and Western blotting were employed to confirm the prediction of key targets and the effects of dioscin on EMT.

Results

Here, using network-based pharmacological methods, we found 42 possible lung cancer-related targets of dioscin, which were assigned to 98 KEGG pathways. Among the 20 with the lowest p-values, the PI3K-AKT signaling pathway is involved and significantly related to EMT. AKT1 and mTOR, with high degrees (reflecting higher connectivity) in the compound-target analysis, participate in the PI3K-AKT signaling pathway. Molecular docking indicated the occurrence of dioscin-AKT1 and dioscin-mTOR binding. Functional experiments demonstrated that dioscin suppressed the proliferation, migration, invasion, and EMT of human lung adenocarcinoma cells in a dose-dependent manner, without TGF-β stimulation. Furthermore, we determined that dioscin downregulated p-AKT, p-mTOR and p-GSK3β in human lung adenocarcinoma cells without affecting their total protein levels. The PI3K inhibitor LY294002 augmented these changes.

Conclusion

Dioscin suppressed proliferation, invasion and EMT of lung adenocarcinoma cells via the inactivation of AKT/mTOR/GSK3β signaling, probably by binding to AKT and mTOR, and inhibiting their phosphorylation.

Keywords:

• dioscin
• lung adenocarcinoma
• epithelial–mesenchymal transition
• network-based pharmacology

Acknowledgments

This work was supported by the National Natural Science Foundation of China (Grant No. 81873154) and the 2017-annual project of construction of high-level university (Grant No. A1-AFD018171Z11003). We thank Professor Guoan Chen from South University of Science and Technology for his revision of the manuscript.

Abbreviations

EMT, epithelial–mesenchymal transition; DMSO, dimethyl sulfoxide; OD, optical density; MMP, matrix metalloproteinases; PVDF, polyvinylidene fluoride; DAPI, 4’,6-diamidino-2-phenylindole; ANOVA, one-way analysis of variance; PPI, protein-protein interaction; GO, Gene Ontology; KEGG, Kyoto Encyclopedia of Genes and Genomes; CCK8, Cell Counting Kit-8 assay; Dio, dioscin; ZEB1, E-box binding homeobox 1; SLUG, snail family transcriptional repressor 2; SNAIL, snail family transcriptional repressor 1; ZO-1, zona occludens 1; LY, LY294002; SEA, Similarity Ensemble Approach; NSCLC, non-small-cell lung cancer; p-AKT, phosphorylated AKT; p-mTOR, phosphorylated mTOR; p-GSK3β, phosphorylated GSK3β; TGF-β, transforming growth factor beta; FCS, fetal calf serum.

Disclosure

The authors report no conflicts of interest in this work.