Most advanced solid tumors metastasize to different organs. However, no gene therapy effective for multiple tumors has yet been developed. Since a unique characteristic of bone marrow-derived mesenchymal stem cells (MSCs) is that they migrate to tumor tissues, we wanted to determine whether MSCs could serve as a vehicle of gene therapy for targeting multiple tumors. First, we confirmed that mouse MSCs preferentially migrate to multiple tumors of the lung in the Colon-26 (C-26) lung metastasis model. Next, MSCs were efficiently transduced with NK4, an antagonist of hepatocyte growth factor (HGF), by an adenoviral vector with an RGD motif. MSCs expressing NK4 (NK4-MSCs) strongly inhibited development of lung metastases in the C-26 lung metastasis model after systemic administration via a tail vein. Treatment with NK4-MSCs significantly prolonged survival of the C-26-tumor-bearing mice by inhibiting tumor-associated angiogenesis and lymphangiogenesis and inducing apoptosis of the tumor cells. MSC-based gene therapy did not induce the severe adverse effects induced by conventional adenoviral vectors. These results indicate that MSCs can serve as a vehicle of gene therapy for targeting multiple lung metastatic tumors.