Gene therapy provides a promising approach for cancer treatment. Earlier studies suggested that poly-L-lysine-modified iron oxide nanoparticles (IONP-PLL) might be a promising gene delivery system that can transfect DNA efficiently in vitro and in vivo. In this study we used IONP-PLL as gene carriers to deliver the NM23-H1 gene, the first suppressor gene of cancer metastasis, to tumor cells in vivo. The intravenous injection of IONP-PLL carrying NM23-H1-GFP plasmid DNA significantly extended the survival time of an experimental pulmonary metastasis mouse model. In the IONP-PLL/NM23-H1-GFP-treated group, metastasis was clearly suppressed compared with the group treated with free NM23-H1-GFP plasmid. Furthermore, this gene therapy combined with cyclophosphamide treatment resulted in longer survival times and greater suppression of metastasis growth. In conclusion, treatment with IONP-PLL nanoparticles incorporating the NM23-H1gene is an efficient gene therapy method, and it is even more effective in combination with chemotherapy. This approach appears to be a promising strategy for treatment of metastatic tumors.