Malignant gliomas are characterized by invasive and infiltrative behavior that generally involves the destruction of normal brain tissue. Strategies to treat infiltrating gliomas, such as chemotherapy and gene therapy, have remained largely unsuccessful. The infiltrative nature of gliomas can be attributed largely to proteases, which include serine, metallo- and cysteine- proteases. Our previous work and that of others strongly suggest a relationship between the expression of uPAR, MMP-9, and MMP-2; this relationship is generally indicative of the infiltrative phenotype of gliomas. In the present study, we have demonstrated that the RNAi-mediated downregulation of MMP-2 induces apoptosis in the 4910 human glioma xenograft cell line. Using Western blot analysis, we observed that caspase-8 levels increased in MMP-2-downregulated cells whereas TRADD and TRAF-2 levels decreased. Further, NIK levels increased in MMP-2-downregulated cells. To determine the nuclear localization of AIF and IkappaBalpha, we analyzed the levels of AIF, IkappaBalpha and pIkappaBalpha in the cytosolic and nuclear fractions of MMP-2-downregulated cells. Western blot analysis revealed that MMP-2 downregulation resulted in the translocation of AIF to the nucleus and also inhibited the nuclear localization of pIkappaBalpha. To confirm the involvement of AIF, we performed FACS analysis to determine the integrity of the mitochondrial membrane using the MitoPT method. FACS analysis showed that the downregulation of MMP-2 caused a collapse in the mitochondrial cell membrane. Immunolocalization of AIF revealed that in MMP-2-downregulated cells, AIF translocates to the nucleus, thereby enabling the induction of apoptosis. RT-PCR analysis revealed that caspase-8 was overexpressed 57-fold, whereas p73 was downregulated 28-fold. Evidence of apoptosis was determined by TUNEL assay and visualization of nuclear fragmentation by DAPI staining. In summary, it is evident from our results that MMP-2 downregulation induces caspase-8 and AIF-mediated apoptosis and, as such, shows potential for glioma therapy.