Acetylated Nanoformulation of N‐(4‐Hydroxyphenyl)‐4‐Oxoretinamide Inhibits EZH2‐Mediated Epigenetic Repression in Neuroblastoma

ABSTRACT Neuroblastoma (NB) is a highly malignant pediatric extracranial tumor with genetic and epigenetic factors influencing its pathogenesis. Key genetic contributors include germline mutations in ALK and PHOX2B, as well as somatic amplification of MYCN. The epigenetic regulator EZH2, a histone methyltransferase of the PRC2 complex, is overexpressed in NB and linked to poor prognosis in advanced cases. Targeting EZH2 offers a promising therapeutic approach due to its role in tumor proliferation, survival, and maintenance of undifferentiated states. This study explores the use of acetylated human serum albumin nanoparticles (HSANPs) loaded with 4O4HPR for epigenetic therapy in NB, tested in vitro and in vivo using nude mice xenograft models. The nanoparticles enter cells via clathrin‐mediated endocytosis and induce G2‐M cell cycle arrest, mitochondrial depolarization, and the production of reactive oxygen species, which activate Caspase 3 and trigger apoptosis. Mechanistic studies show increased p53 acetylation, stabilizing p53 and inhibiting EZH2's epigenetic silencing. Chromatin Immunoprecipitation reveals the disruption of EZH2 binding to the E‐Cadherin promoter, which suppresses cell migration and EMT‐like transition. Western blotting confirms upregulation of epithelial and downregulation of mesenchymal markers, with inhibited wound closure in SH‐SY5Y cells. This nanotherapy disrupts EZH2‐E‐Cadherin interaction, offering novel translational potential for NB treatment.