Network pharmacological analysis of corosolic acid reveals P4HA2 inhibits hepatocellular carcinoma progression.

BACKGROUND: Corosolic acid is a pentacyclic triterpene acid with hypoglycemic, anti-inflammatory, and anti-cancer effects. However, its potential targets in hepatocellular carcinoma (HCC) are unknown, hindering clinical utilization. METHODS: Differentially expressed proteins of the Bel-7404 cell line were identified with tandem mass tag analysis and differentially expressed genes (DEGs) of an HCC TCGA dataset using bioinformatics. Gene functions and pathways were inferred using the DAVID database. Online databases were used to establish P4HA2 expression in HCC (GEPIA2) and its relationship with patient survival (UALCAN and The Human Protein Atlas), the association between P4HA2 expression and immune cell infiltration (TIMER2), and DNA methylation of the P4HA2 gene (MethSurv). Cell proliferation, cell cycle, and cell death were assessed with PI and SYTOX-Green staining, CCK-8, and colony formation assays. Protein expression levels were detected by Western blotting. RESULTS: A total of 44 differentially expressed proteins and 4498 DEGs were identified. Four genes whose proteins were also found in the differential protein profile but with opposing expressions were selected as candidate targets. The candidate gene prolyl 4-hydroxylase subunit alpha 2 (P4HA2) was recognized as the only potential target due to its high expression in public datasets, association with poor patient survival, and relation to immune cell infiltration in HCC tissues. Moreover, the DNA methylation status in 4 CpG islands of the P4HA2 gene correlated with a poor prognosis. Furthermore, corosolic acid treatment inhibited the proliferation of HCC cell lines Bel-7404 and HepG2 in a dose-dependent manner, caused G2/M phase cell cycle arrest, and promoted cell death. In addition, the treatment reduced P4HA2 protein levels. CONCLUSION: Our results indicate that P4HA2 is a potential target of corosolic acid. Thus, they contribute to understanding molecular changes in HCC after corosolic acid treatment and facilitate finding new treatment regimens.

The Tian-Men-Dong decoction suppresses the tumour-infiltrating G-MDSCs via IL-1ß-mediated signalling in lung cancer.

ETHNOPHARMACOLOGICAL RELEVANCE: The traditional Chinese medicine (TCM) Tian-Men-Dong decoction (TD) has been able to effectively treat lung cancer in China for thousands of years. TD improves the quality of life in lung cancer patients by promoting nourishment of yin and reducing dryness, clearing the lung and removing toxins. Pharmacological studies show that TD contains active antitumour ingredients, but its underlying mechanism remains unknown. AIM OF THE STUDY: This study aims at exploring potential mechanisms of TD in the treatment of lung cancer by regulating granulocytic-myeloid-derived suppressor cells (G-MDSCs). MATERIALS AND METHODS: An orthotopic lung cancer mouse model was generated by intrapulmonary injection with LLC-luciferase cells in immunocompetent C57BL/6 mice or immunodeficient nude mice. TD/saline was orally administered once to the model mice daily for 4 weeks. Live imaging was conducted to monitor tumour growth. Immune profiles were detected by flow cytometry. H&E and ELISA were applied to test the cytotoxicity of the TD treatment. RT-qPCR and western blotting were performed to detect apoptosis-related proteins in G-MDSCs. A neutralizing antibody (anti-Ly6G) was utilized to exhaust the G-MDSCs via intraperitoneal injection. G-MDSCs were adoptively transferred from wild-type tumour-bearing mice. Immunofluorescence, TUNEL and Annexin V/PI staining were conducted to analyse apoptosis-related markers. A coculture assay of purified MDSCs and T cells labelled with CFSE was performed to test the immunosuppressive activity of MDSCs. The presence of TD/IL-1ß/TD + IL-1ß in purified G-MDSCs cocultured with the LLC system was used for ex vivo experiments to detect IL-1ß-mediated apoptosis of G-MDSCs. RESULTS: TD prolonged the survival of immune competent C57BL/6 mice in an orthotopic lung cancer model, but did not have the same effect in immunodeficient nude mice, indicating that its antitumour properties of TD are exerted by regulating immunity. TD induced G-MDSC apoptosis via the IL-1ß-mediated NF-κB signalling cascade leading to effectively weaken the immunosuppressive activity of G-MDSCs and promote CD8+ T-cell infiltration, which was supported by both the depletion and adoptive transfer of G-MDSCs assays. In addition, TD also showed minimal cytotoxicity both in vivo and in vitro. CONCLUSION: This study reveals for the first time that TD, a classic TCM prescription, is able to regulate G-MDSC activity and trigger its apoptosis via the IL-1ß-mediated NF-κB signalling pathway, reshaping the tumour microenvironment and demonstrating antitumour effects. These findings provide a scientific foundation the clinical treatment of lung cancer with TD.