Objective To explore the potential mechanism of action of fraxinellone in the treatment of pulmonary arterial hypertension (PAH) using network pharmacology and molecular docking, and to validate the results by real-time quantitative PCR (RT-qPCR).

Methods The PubChem database was used to obtain the SMILES structure of fraxinellone. The SwissTargetPrediction platform was utilized to predict potential targets of fraxinellone in humans. PH-related disease targets were screened using the GeneCards, OMIM, and TTD databases, and the common drug-disease targets were obtained after taking the intersection. Furthermore, a protein-protein interaction (PPI) network was constructed using the STRING database, and key gene targets were screened through analysis combined with Cytoscape software. DAVID platform was utilized to conduct Gene Ontology (GO) functional enrichment analysis and Kyoto Encyclopedia of Genes and Genomes (KEGG) pathway enrichment analysis on the targets of fraxinellone in the treatment of PH. Molecular docking between fraxinellone and the core targets was performed using AutoDock Vina, and the docking results were visualized with PyMOL. RT-qPCR was employed to validate the changes in the expression levels of key targets following fraxinellone treatment.

Results A total of 101 fraxinellone-related targets and 1,482 PAH-related disease targets were obtained, yielding 27 common drug-disease targets. GO functional enrichment analysis revealed that these targets were primarily involved in biological processes such as inflammatory response regulation and oxidative stress. KEGG pathway enrichment analysis identified pathways directly associated with PAH, including the PI3K-Akt signaling pathway and the calcium signaling pathway. The molecular docking results showed that fraxinellone exhibited good binding affinity with the five screened key targets. The binding energies of fraxinellone with epidermal growth factor receptor (EGFR), myeloperoxidase (MPO), prostaglandin-endoperoxide synthase 2 (PTGS2), C-C chemokine receptor 2 (CCR2), and C-X-C chemokine receptor 2 (CXCR2) were -8.5, -7.3, -7, -6.1, -7.2 kcal/mol, respectively, indicating that fraxinellone had good binding activity with these five key targets. RT-qPCR revealed that after treatment with 10 μmol/L fraxinellone for 24 h, the levels of EGFR, CCR2, and CXCR2 all showed a decreasing trend, with the most significant decrease observed in EGFR.

Conclusion Fraxinellone may exert its therapeutic effect on PAH by regulating key targets such as EGFR, CCR2, and CXCR2, and participating in inflammation- and oxidative stress-related pathways. This study provides a theoretical basis for subsequent experimental validation and drug development.

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