Lung cancer ranks first in both incidence and mortality and is a major health concern worldwide. Upon recognition of specific antigens on tumor cells, complement-dependent cytotoxicity (CDC) is activated, arresting cell growth or inducing apoptosis. However, by overexpressing CD59, a membrane complement regulatory protein (mCRP), lung cancer cells develop resistance to CDC. We previously showed that virus-like particles (VLPs) of human JC polyomavirus (JCPyV) could be used as a gene therapy vector to carry a suicide gene expression plasmid with a lung-specific promoter (SP-B (surfactant protein B)) for lung adenocarcinomas. Herein, we designed a CD59-specific short hairpin RNA (shRNA) expression plasmid driven by SP-B (pSPB-shCD59) to effectively and specifically inhibit CD59 overexpression in lung cancer cells. Treatment of lung cancer cells in vitro with JCPyV VLPs containing pSPB-shCD59 (pSPB-shCD59/VLPs) induces CDC and death of cancer cells. Mice that were subcutaneously injected with human lung cancer cells showed an 87% inhibition in tumor growth after tail vein injection of pSPB-shCD59/VLPs. Moreover, in a mouse model of lung cancer metastasis, a reduction in the lung weight by 39%, compared with the control group, was observed in mice treated with pSPB-shCD59/VLPs after tail vein injection of human lung cancer cells. Furthermore, tissue sectioning showed that the number and size of tumors produced was significantly reduced in the lungs of mice in the treatment group than those of the untreated group, indicating inhibition of metastasis by pSPB-shCD59/VLPs. Together, these results demonstrate the potential of pSPB-shCD59/VLPs as a therapeutic agent for CD59 overexpressed lung cancer.