Matrix metalloproteinases (MMP) are a group of proteinases that have normal physiologic roles degrading and remodeling the extracellular matrix. They also have multiple roles in different stages of tumor progression. Elevated levels of MMPs have been observed in many tumors; these increases have a strong association with the invasive phenotype. MMP-2 and MMP-9 are particularly involved in cancer invasion and metastasis. MMP inhibitors are currently being tested as therapeutic agents for a number of cancers in both preclinical models and in clinical trials. To date, clinical trials using this strategy have had limited efficacy. A major concern is the lack of specificity of commercially available MMP inhibitors. An adenoviral vector expressing small interfering RNA against the MMP-2 gene (Ad-MMP-2) was constructed to specifically inhibit MMP-2 expression. The effect of Ad-MMP-2 on invasion, angiogenesis, tumor growth, and metastasis of A549 lung cancer cell was evaluated. Ad-MMP-2 infection of lung cancer cells showed specific down-regulation of MMP-2 protein, activity, and transcription as determined by Western blotting, gelatin zymography, and reverse transcription-PCR. Ad-MMP-2 inhibition also mitigated lung cancer invasion and migration, and reduced tumor cell-induced angiogenesis in vitro. In an experimental metastatic lung tumor model, treatment of established tumors by Ad-MMP-2 inhibited s.c. tumor growth and formation of lung nodules in mice. Adenoviral-mediated RNA interference against MMP-2 has significant therapeutic potential for lung cancer and exerts some of this effect by inhibiting angiogenesis.