Alpinumisoflavone against cancer pro-angiogenic targets: In silico, In vitro, and In ovo evaluation.

Background: Breast cancer is currently the world's most predominant malignancy. In cancer progression, angiogenesis is a requirement for tumor growth and metastasis.Alpinumisoflavone (AIF), a bioactive isoflavonoid, exhibited good binding affinity with the angiogenesis pathway's druggable target through molecular docking.

Objectives: To confirm AIF's angiogenesis inhibitory activity, cytotoxic potential toward breast cancer cells, and druggability.

Methods: Antiangiogenic activity was evaluated in six pro-angiogenic proteins in vitro, duck chorioallantoic membrane (CAM) in ovo, molecular docking and druggability in silico.

Results: Findings showed that AIF significantly inhibited (p =  < 0.001) the HER2(IC₅₀ = 2.96 µM), VEGFR-2(IC₅₀ = 4.80 µM), MMP-9(IC₅₀ = 23.00 µM), FGFR4(IC₅₀ = 57.65 µM), EGFR(IC₅₀ = 92.06 µM) and RET(IC₅₀ =  > 200 µM) activity in vitro.AIF at 25 µM-200 µM significantly inhibited (p =  < 0.001) the total number of branch points (IC₅₀ = 14.25 μM) and mean length of tubule complexes (IC₅₀ = 3.52 μM) of duck CAM comparable (p =  > 0.001) with the positive control 200 µM celecoxib on both parameters.AIF inhibited the growth of the estrogen-receptor-positive (ER +) human breast cancer cells (MCF-7) by 44.92 ± 1.79% at 100 µM while presenting less toxicity to human dermal fibroblast neonatal (HDFn) normal cells.The positive control 100 µM doxorubicin showed 86.66 ± 0.93% and 92.97 ± 1.27% inhibition with MCF-7 (IC₅₀ = 3.62 μM) and HDFn, (IC₅₀ = 27.16 μM) respectively.In docking, AIF has the greatest in silico binding affinity on HER2 (-10.9 kcal/mol) among the key angiogenic molecules tested. In silico rat oral LD₅₀ calculation indicates that AIF is moderate to slightly toxic at 146.4 mg/kg with 1.1 g/kg and 20.1 mg/kg upper and lower 95% confidence limits. Lastly, it sufficiently complies with Lipinski's, Veber's, Egan's, Ghose's, and Muegge's Rule, supporting its oral drug-like property.

Conclusion: This study revealed that AIF possesses characteristics of a phytoestrogen compound with significant binding affinity, inhibitory activity against pro-angiogenic proteins, and cytotoxic potential against ER + breast cancer cells.The acceptable and considerable safety and drug-likeness profiles of AIF are worthy of further confirmation in vivo and advanced pre-clinical studies so that AIF can be elevated as a promising molecule for breast cancer therapy.