Network-based Pharmacology and In vitro Validation Reveal that Galangin Induces Apoptosis in Bladder Cancer Cells by Promoting the P53 Signaling Pathway

Abstract

Background: Galangin is one of the flavonoids in Alpinia officinarum. It has various anti-tumor activities, but its anti-bladder cancer effect is unclear.

Objective: To investigate the mechanism of action of galangin against bladder cancer using a network pharmacology approach.

Methods: The TCM Systematic Pharmacology Database and Analysis Platform (TCMSP), SwissTargetPrediction database, and the Targetnet database were used to predict the targets of action of galangin. Bladder cancer-related targets were obtained through the GeneCards database. The intersection of the two was taken as the target of galangin's action against bladder cancer. The intersecting targets were screened for core targets using the STRING database and Cytoscape 3.9.0 software to build a protein-protein interaction (PPI) network of targets. The core targets were subjected to gene ontology (GO) functional annotation and Kyoto Encyclopedia of Genes and Genomes (KEGG) pathway enrichment analysis using the online annotation and visual integration analysis tool DAVIDBioinformaticsResources (2021Update). A drug-disease-target-pathway network was constructed using Cytoscape 3.9.0 software. The antibladder cancer effect of galangin was observed by cell proliferation, and plate cloning assay; apoptosis of bladder cancer cells induced by galangin was detected by Hoechst33342 staining and flow cytometry; protein immunoblotting (Western-blot) was used to detect the effect of galangin on apoptosis-related proteins Bax, Bcl-2, Cleaved-PARP, p53 signaling pathway p53 and cytc.

Results: A total of 115 genes were obtained from galangin against bladder cancer, and 16 core targets were screened. The kEGG pathway enrichment analysis included Pathways in cancer, PI3K-AKT signaling pathway, p53 signaling pathway, etc. In vitro experiments showed that galangin could inhibit bladder cancer cell proliferation, induce apoptosis, upregulate the expression of apoptosis-related proteins Bax and Cleaved-PARP and downregulate the expression of Bcl-2; meanwhile, galangin could promote the upregulation of the expression of p53 and cytc proteins by activating the p53 signaling pathway.

Conclusion: Galangin induced apoptosis in bladder cancer cells by activating the p53 signaling pathway.

Keywords: Bladder cancer; Sophora flavescens; apoptosis; galangin; network pharmacology; p53 signaling pathway.