Abstract 

 Neuroblastoma (NB) is a common solid tumor in children and infants, with its formation and regression closely linked to the tumor-host immune mechanism. STING agonists, particularly cyclic dinucleotide (CDN), have promising potential in NB therapy by generating innate and adaptive immune stimulation, leading to tumor control. In vivo CDN delivery is challenging due to its negative charge, hydrophilicity, and susceptibility to degradation by phosphodiesterase, hindering its effectiveness. Thus, our study proposed four methods to load CDN into liposomes, using 2',3'-cGAMP as the model drug. Lipid nanoparticles were prepared, followed by physicochemical characterization. Subsequently, cellular inhibition and immune stimulation were investigated. As a result, lipid calcium phosphate nanoparticles (LCP-NP) possessed the highest encapsulation efficiency among the four preparation methods, with a diameter of 82.57±3.72nm. LCP-NP maintained size stability under refrigeration conditions at 4°C within 48 hours. The surface of the liposome was positively charged. Compared with free cGAMP, LCP-NP resulted in a slower release, enhanced cytotoxicity against tumor cells, greater activation of the cGAS-STING pathway, and increased expression of the immune factors. These findings clearly demonstrated the effectiveness of the liposomal delivery system for cGAMP and provided a promising strategy for the treatment of NB.