Unsaponifiable fraction of black Vitis vinifera seed oil attenuates liver cancer progression by targeting apoptosis and key tumor-associated genes: In vitro, in vivo, and in silico studies.

Cancer is the leading cause of death worldwide. This study evaluated the anti-liver cancer influence of the unsaponifiable fraction (UnSap) derived from black Vitis vinifera seed oil (BVVO) using the HepG-2 cell line and a HCC mouse model. Compared with 5-fluorouracil (5-FU), UnSap exhibited a strong antioxidant capacity, excellent safety (in PBMCs), and a higher selectivity index (SI) with a lower IC₅₀ against HepG-2 cells. It effectively induced apoptosis and modulated the expression of key genes involved in cell cycle progression. The docking results predicted that the abundant constituents of UnSap can interfere with the interaction between β-catenin and human T-cell factor-4 (HTCF-4) and may inhibit the activity of key regulatory proteins involved in HCC proliferation and stemness. In HCC-bearing mice, UnSap significantly improved the expression of serum liver function markers (0.4-1.3-fold), restored redox balance (> threefold), and preserved hepatic tissue architecture. It also modulated the expression of critical genes associated with apoptosis (> 20-fold), cell cycle regulation (> fourfold), inflammation (> fivefold), angiogenesis (threefold), and cancer stemness (> 45-fold). These therapeutic effects are likely attributed to its high phenolic and phytosterol content, as confirmed by HPLC analysis. Across all investigated parameters, UnSap demonstrated superior anticancer efficacy compared with 5-FU. These findings highlight the potential of UnSap as a promising natural therapeutic candidate and provide a foundation for future studies on its bioactive constituents for the development of novel anticancer agents.