HSP27 mRNA Silencing and Radiation-Driven Endosomal-Lysosomal Escape of Lipid Nanomicelles Synergistically Radiosensitize Castration-Resistant Prostate Cancer Cells

Despite significant advances in antisense oligonucleotide (ASO) chemistry and nanocarrier design, therapeutic efficacy is often limited by inefficient cellular uptake and sequestration within endosomal–lysosomal compartments. Here, we show that ionizing radiation can be leveraged to overcome these barriers and enhance antisense nanomedicine delivery in castration-resistant prostate cancer (CRPC). CRPC cells (PC-3, C4-2) exhibited greater resistance to irradiation than castration-sensitive prostate cancer cells (LNCaP) and upregulated heat shock protein 27 (HSP27) in response to radiation, indicating an adaptive stress response. HSP27 downregulation by HSP27 mRNA -targeting ASO restored radiation-induced apoptotic signaling. To improve intracellular delivery, we developed lipid-modified ASOs (LASOs) targeting HSP27 mRNA that self-assemble into nanomicelles and enter cells via macropinocytosis. Although LASOs showed enhanced cellular uptake, they were predominantly sequestered within endosome–lysosome compartments, limiting their therapeutic activity. Strikingly, irradiation promoted endosomal-lysosomal escape of LASO nanomicelles, facilitating cytoplasmic delivery, robust HSP27 suppression, ultimately leading to increased anti-cancer effects, including an approximately 75% reduction in CRPC cell viability and delayed CRPC 3D spheroid growth. Collectively, these findings reveal a bidirectional therapeutic synergy in which HSP27 silencing resensitizes CRPC cells to radiation while irradiation enhances LASO intracellular trafficking. This study establishes irradiation as a trigger for intracellular drug release and provides a synergistic strategy integrating radiotherapy with antisense nanomedicine to overcome delivery barriers and restore therapeutic efficacy in CRPC.