Expression of urokinase-type plasminogen activator (uPA) by human and rat brain tumor cells and their susceptibility to uPA-targeted rSeV/dMFct14(uPA2), namely, BioKnife (*P < 0.01). (a) Expression of uPA by human glioblastoma multiforme cells (U87, U138, U251, U373, and A172) and rat gliosarcoma (9L) in culture medium. Four days after changing to fresh medium, the culture supernatant was subjected to a colorimetric catalytic assay (n =6 wells/group). (b) Susceptibility of brain tumor cells to BioKnife-GFP or control vector (rSeV/dM-GFP) at an multiplicity of infection of 1.25. Four days after virus inoculation, cells were observed under a fluorescent microscope (panels) and the culture supernatant was subjected to cytotoxicity assay (graphs). In the case of U251, a group with PAI-1, a specific inhibitor of uPA, was included. Note that syncytium formation was observed in wells with U138, U251, U373, or A172 cells (panels), but not in wells with U87 or 9L cells. Significant cytotoxicity was observed against U251 and U373 cells, but not against A172 cells (left graph). Addition of PAI-1 almost completely prevented the BioKnife-dependent cytotoxicity (right graph; n = 6 wells/group).