A cationic lipid-based delivery system composed of N [(1-(2,3-dioleyloxy)propyl)]-N,N,N-trimethylammonium chloride (DOTMA) and cholesterol, at a 4:1 mole ratio, was developed to express anti-angiogenic gene products from normal and tumor vasculature upon intravenous administration. Plasmid biodistribution, expression, and efficacy were characterized in a syngeneic mouse tumor model using SCCVII (squamous cell carcinoma) cells. Transfection was demonstrated in both lung and primary tumors. Fluorescence microscopy showed that the plasmid-lipid complexes were passively targeted to the tumor vasculature and that the plasmid was localized to endothelial cells. Transgene expression levels were higher in larger tumors, correlating well with increased tumor vasculature as a function of tumor size. The levels of gene expression in subcutaneously implanted tumors peaked 8 to 18 hours after DNA administration, and expression was detectable up to 10 days after a single injection. A second administration, three days following the first, extended the duration of expression in the tumor, whereas no impact on expression was observed in the lungs. IL-12 plasmid produced protein levels in the lung, blood and tumor upon i.v. administration. A dose-response curve showed that IL-12 could be detected in the tumor with as little as 15 mg of plasmid. This dose, administered three times over a 14 day period, produced a significant inhibition of primary tumor growth. Co-expression of murine endostatin and murine IL-12 produced a synergistic inhibition of tumor growth. These results demonstrate the feasibility of systemically administered plasmid-based gene delivery systems for the treatment of angiogenesis-dependent tumors.