Effect of PGV-1 on Apoptosis via Mitotic Arrest and Senescence in Polyploid Giant Cancer Cells of Hepatocellular Carcinoma JHH4

Objectives: Senescent cells release a senescence-associated secretory phenotype, promoting polyploid giant cancer cells (PGCCs) to emerge, fostering tumor heterogeneity and resistance. Pentagamavunone-1 (PGV-1) emerges as a promising agent inducing senescence and prometaphase arrest, resulting in permanent cytotoxicity. This study was aimed to investigate the effect of PGV-1 in dysregulating mitosis through the modulation of PGCCs and senescence in low MYCN-expressing hepatocellular carcinoma (HCC) cells JHH4. Materials and Methods: To assess the physiological effects of PGV-1, several in vitro tests were done including the MTT assay, cell cycle assay, May-Grünwald-Giemsa staining, senescence associated-β-galactosidase (SA-β-Gal) assay, and apoptosis assay. The protein levels of the apoptosis regulatory protein were evaluated using western blot analysis. The interaction of PGV-1 toward the protein that plays a role in PGCCs formation was simulated by molecular docking and molecular dynamics (MD). Results: The cytotoxic assay revealed that PGV-1 inhibited the proliferation of JHH4 liver cancer cells permanently. Inhibition of cell proliferation by PGV-1 was associated with the modulation of G2/M phase, particularly mitotic arrest and formation of PGCCs. The SA-β-Gal verified that PGV-1 induced senescence in cells (p<0.01), inducing PGCCs formation. Apoptotic mechanisms were validated via Annexin V staining, which showed the level of cleaved poly (ADP-ribose) polymerase (p<0.001). Molecular docking and MD simulations suggested that PGV-1 could interfere with the conformation of the chromosomal passenger complex (CPC) protein, particularly disrupting essential interactions within the inner centromere protein, Survivin, and Borealin. Conclusion: PGV-1 induced strong cytotoxicity in HCC cells by disrupting mitosis leading to PGCC formation, senescence, and subsequent apoptotic cell death.