P53-mediated apoptosis and topoisomerase IIα inhibition by an acridine derivative in hepatocellular carcinoma

Abstract Background Primary liver malignancy represents a significant worldwide health burden characterized by restricted therapeutic alternatives and elevated relapse frequencies. While acridine-based compounds have demonstrated promising bioactive properties including anticancer capabilities, challenges persist in enhancing their therapeutic efficacy and target specificity. Purpose This investigation evaluated the antineoplastic properties and underlying molecular mechanisms of a newly synthesized acridine hydrochloride compound, N-(3-(4-(3-aminopropyl)piperazin-1-yl)propyl)acridin-9-amine hydrochloride (9-BAPPA), targeting HepG2 hepatocellular carcinoma cells. Methods Cell viability in NHPL and HepG2 cells was determined using the MTT assay. Cell death was showed both morphologically with fluorescent double staining (AO/EB) and biochemically with flow cytometry analysis. The levels of p53, BCL-2, BAX, caspase-3, and PCNA proteins was assessed by immunocytochemistry, while mRNA expression of p53, caspase-3, and the BAX/BCL-2 ratio were measured using qRT PCR. In addition, molecular docking with topoisomerase IIα was performed to explore the compound’s predicted binding mode. Results In vitro assays showed that 9-BAPPA selectively inhibited HepG2 cells (IC₅₀ = 15.2 µg/mL) with much lower toxicity toward normal NHPL cells (IC₅₀ = 34.3 µg/mL). Treated cells exhibited apoptotic morphology and cell cycle arrest at sub-G₀/G₁ and G₂/M phases ( p < 0.0001). Upregulation of p53, caspase-3, and the BAX/BCL-2 ratio at both mRNA and protein levels ( p ≤ 0.01) confirmed the activation of the intrinsic p53-dependent mitochondrial apoptosis pathway. Computational docking analyses revealed robust binding interactions between 9-BAPPA and essential residues (MET 762, MET 766, Q778) within topoisomerase IIα, confirming its mechanism as a topoisomerase-interfering compound. In silico Pharmacokinetics showed good oral bioavailability and promising permeability of Blood Brain Barrier. Conclusion 9-BAPPA exhibits significant selective antineoplastic properties through P53-mediated apoptotic pathway activation and cell cycle disruption in HepG2 cells, warranting additional investigation as a potential therapeutic compound for hepatic malignancies.