Characterization of ficus hispida fruit extract and assessment of Gallic acid using in Silico and in vitro studies against breast cancer

This study aimed to characterize the phytochemicals in Ficus hispida fruits and evaluate the anticancer potential of gallic acid against breast cancer using in-silico and in-vitro approaches. The objectives were to identify bioactive compounds using LC-MS, assess gallic acid's binding affinity with breast cancer-related targets via molecular docking and dynamics, and validate its cytotoxicity and apoptotic effects in breast cancer (MDA-MB-231) cells. Ethyl acetate extract of Ficus hispida fruits was prepared via Soxhlet extraction and analyzed using LC-MS/MS. Molecular docking studies were performed to evaluate gallic acid's interactions with breast cancer proteins (2VCJ-Heat shock protein HSP90-alpha, 1P7K-Anti-ssDNA antibody Fab fragment, 5OTE-Serine/threonine-protein kinase MRCK beta), followed by molecular dynamics (MD) simulations for stability analysis. In-vitro cytotoxicity (MTT assay) and apoptosis detection (AO/EB staining, DAPI staining) were conducted on MDA-MB-231 cells. LC-MS analysis identified key phytochemicals, including gallic acid (m/z 169.14), quercetin (m/z 302.33), and β-sitosterol (m/z 414.58). Molecular docking revealed strong binding of gallic acid to Heat shock protein HSP90-alpha (-6.2 kcal/mol), stabilized by hydrogen bonds and hydrophobic interactions. MD simulations confirmed stable protein-ligand complexes with an RMSD of 2.1 Å for the protein and 3.2 Å for the ligand. In vitro studies showed the dose-dependent cytotoxicity of gallic acid against MDA-MB-231 cells, with an IC₅₀ of 15-20 µg/mL and 34.80% viability at 20 µg/mL. Apoptosis was confirmed through nuclear fragmentation and chromatin condensation. Ficus hispida fruits are rich in bioactive compounds, particularly gallic acid, demonstratinga strong binding affinity to breast cancer targets and potent cytotoxic effects against MDA-MB-231 cells. These findings highlight its potential as a therapeutic candidate for breast cancer treatment, warranting further clinical exploration.