Simultaneous exploitation of phaeanthine - a bisbenzylisoquinoline alkaloid for dual anti-metastatic and anti-angiogenic potential by disruption of Akt/NF-κB/MMP regulatory pathways.

Metastasis and angiogenesis are critical drivers of cancer progression, contributing to tumor aggressiveness and therapeutic resistance. Phaeanthine (PHA), a bisbenzylisoquinoline alkaloid isolated from Cyclea peltata, has previously demonstrated anticancer activity by inhibiting Akt signaling. The present study investigated the anti-metastatic and anti-angiogenic potential of PHA in triple-negative breast cancer (TNBC) and analysed the molecular changes associated with its activity using experimental and computational approaches. Anti-metastatic activity was evaluated in MDA-MB-231 cells using wound-healing, adhesion, and transwell invasion assays, followed by Western blot analysis of key regulatory proteins. Anti-angiogenic effects were assessed in EA.hy926 endothelial cells using tube formation assays and VEGFR2 immunofluorescence analysis. Molecular docking and molecular dynamics (MD) simulations were performed to investigate potential molecular interactions. PHA significantly inhibited the migration, adhesion, and invasion of MDA-MB-231 cells at sub-micromolar concentrations and induced apoptosis. It suppressed the expression of metastasis-associated proteins, including MMP-2, MMP-9, integrins, Akt, NF-κB, and HIF-1α, while reversing epithelial-mesenchymal transition, as evidenced by reduced N-cadherin and vimentin expression. In EA.hy926 cells, PHA markedly inhibited tube formation at 2 μM and downregulated VEGFR2 expression. In silico studies revealed favourable binding affinities (<-6.0 kcal mol-1) toward multiple metastatic and angiogenic targets. Subsequently, MD simulations provided computational support for potential interactions of PHA with MMP-2 and MMP-9, consistent with their pronounced downregulation observed experimentally. Collectively, these findings indicate the dual anti-metastatic and anti-angiogenic effects of PHA through coordinated modulation of Akt/NF-κB/HIF-1α/MMP-associated proteins, extracellular-matrix remodeling and angiogenesis. This study provides the first evidence of the activity of PHA against TNBC and its anti-angiogenic potential, supporting further investigation of PHA as a potential lead phytochemical against TNBC-associated metastatic and angiogenic processes.