Anticancer Potential of Colebrookea oppositifolia Flower Extract Against Breast and Prostate Cancer: An Integrated LC-MS, Network Pharmacology, Molecular Dynamics, PCA, and MM-GBSA Approach

Colebrookea oppositifolia is traditionally used in Nepal for the treatment of cancer. This study investigated the anticancer mechanisms of C. oppositifolia flower extract against breast and prostate cancers using an integrated approach combining network pharmacology, molecular docking, molecular dynamics (MD) simulations, and experimental validation. Phytoconstituents from a 70% methanolic extract of C. oppositifolia flowers were identified via LC-MS analysis and screened for drug-likeness and ADME properties using the TCMSP and MolSoft platforms. The intersection of target profiles from Swiss Target Prediction and GeneCards via Venny 2.1.0 revealed overlapping cancer-related genes. Network pharmacology and enrichment analyses conducted using STRING, Cytoscape, and ShinyGO delineated protein–protein interaction (PPI) networks, Gene Ontology (GO) terms, and KEGG pathways. Molecular docking via PyRx 0.8 predicted high binding affinities of active flavonoids against AKT1, EGFR, and SRC. Subsequent 200-ns molecular dynamics (MD) simulations, principal component analysis (PCA), and MM-GBSA calculations confirmed the structural stability and favorable binding energetics of the top complexes. Finally, MTT assays validated the anticancer efficacy of the extract against breast and prostate cancer cell lines. LC-MS identified nine phytoconstituents, including three bioactive flavonoids: apigenin, diosmetin, and apigetrin. PPI network analysis revealed AKT1, EGFR, and SRC as core therapeutic targets. GO and KEGG pathway enrichment analyses revealed that these flavonoids primarily modulate cellular responses to UV-A, apoptosis, oxidative stress, and nitric oxide synthase regulation. Furthermore, the anti-cancer mechanisms were strongly associated with endocrine resistance, EGFR tyrosine kinase inhibitor resistance, ErbB signaling, and estrogen signaling pathways. Molecular docking demonstrated that apigetrin possesses high binding affinity for EGFR (− 9.9 kcal/mol) via robust hydrogen bonding, supported by stable MD trajectories. MM-GBSA calculations confirmed a highly favorable binding free energy (− 41.03 ± 1.17 kcal/mol) driven predominantly by van der Waals and electrostatic forces. Complementary MTT assays demonstrated the significant, dose-dependent cytotoxicity of the C. oppositifolia extract against both breast and prostate cancer cell lines. These findings establish apigetrin as a promising lead anticancer scaffold from the C. oppositifolia flower, warranting further targeted isolation of its bioactive constituents, as well as subsequent mechanistic in vitro and in vivo preclinical evaluations.