Alveolar rhabdomyosarcoma (ARMS) are aggressive soft tissue tumors harboring specific fusion transcripts, notably PAX3-FOXO1 (P3F). Current therapy concepts result in unsatisfactory survival rates making the search for innovative approaches necessary: targeting PAX3-FOXO1 could be a promising strategy. In this study, we developed integrin receptor-targeted Lipid–Protamine-siRNA (LPR) nanoparticles using the RGD peptide and validated target specificity as well as their post-silencing effects. We demonstrate that RGD-LPRs are specific to ARMS in vitro and in vivo. Loaded with siRNA directed against the breakpoint of P3F, these particles efficiently down regulated the fusion transcript and inhibited cell proliferation, but did not induce substantial apoptosis. In a xenograft ARMS model, LPR nanoparticles targeting P3F showed statistically significant tumor growth delay as well as inhibition of tumor initiation when injected in parallel with the tumor cells. These findings suggest that RGD-LPR targeting P3F are promising to be highly effective in the setting of minimal residual disease for ARMS.