Osteosarcoma (OS) is the most common primary bone malignancy characterized by deposition of an immature osteoid matrix. OS treatment has proven challenging because of the high risk of metastatic progression and recurrence after chemotherapy. Melittin (MLT) is recognized as a potential antitumor candidate to overcome chemotherapy resistance and provoke superior immunostimulatory effects. However, the application of MLT to OS is hampered by severe toxic side effects and a lack of tumor-targeting ability. Herein, a self-assembled nanopolymer named LC09-MLT@F127 was developed by binding MLT with F127 micelles and then modifying an aptamer (LC09) for targeted drug delivery during OS treatment. LC09-MLT@F127 exhibited significant OS-targeting ability in vitro and in vivo owing to the aptamer LC09 decoration. Moreover, LC09-MLT@F127 significantly reduced the hemolytic toxicity of MLT while maintaining its tumor-killing ability. In an orthotopic transplantation model of OS, LC09-MLT@F127 induced immunogenic cell death and facilitated the maturation of dendritic cells (DCs), thereby resulting in the activation of tumor-specific immune responses and the inhibition of OS deterioration. Taken together, these finding suggest that LC09-MLT@F127 may be an encouraging MLT-based immunotherapy option for OS.