Experimental Study on the Anti-prostate Cancer Cell Effects of AZIN Small Molecule Inhibitors

　　Objective: To design and identify a small-molecule AZIN inhibitor capable of disrupting the AZ–AZIN interaction, restoring antizyme (AZ) activity, reducing ornithine decarboxylase (ODC) levels, and suppressing polyamine-dependent tumor cell growth.
　　Methods: The crystal structure of AZIN (PDB: 3BTN) was used to identify a druggable pocket and perform structure-based virtual screening. Chemical assays and PC3 cell experiments were conducted, including proliferation assays, RT-PCR, and Western blot for ODC/AZ/AZIN expression, polyamine quantification, and flow-cytometry-based analyses of cell cycle and apoptosis.
　　Results: Virtual screening yielded a lead compound, AZIN-53. AZIN-53 increased AZ protein levels and decreased ODC protein abundance without altering AZIN or the mRNA expression of ODC/AZ/AZIN, indicating enhanced free AZ availability. AZIN-53 reduced putrescine, spermidine, and spermine levels, demonstrating its ability to suppress polyamine synthesis and uptake. Functionally, AZIN-53 inhibited PC3 cell proliferation, induced G1 cell-cycle arrest, and triggered apoptosis.
　　Conclusion: AZIN-53 disrupts the AZ–AZIN interaction, restores AZ-mediated ODC degradation, and effectively lowers intracellular polyamines, ultimately inhibiting tumor cell growth. These findings highlight AZIN-53 as a promising first-in-class small-molecule AZIN inhibitor and a potential therapeutic strategy targeting polyamine dysregulation in cancer.