To identify genes involved in local tumor cell invasion, we performed differential display using two RNA samples derived from glioblastoma multiforme (GBM) cells collected by laser capture microdissection. We harvested tumor cells (20,000 each) from the central core and the invasive rim of a human GBM specimen and identified neoplastic cells at the invasive edge of the tumor using morphological criteria (mainly nuclear atypia). We found differential expression of 50–60 complementary DNA bands between rim and core. Two sequences were markedly overexpressed in the tumor rim and showed strong homology to the 8-kD polypeptide P311 (GI: 1244509) and the 46-kD Death Associated Protein-3 (DAP3; GI: 10368653). Overexpression (2- to 20-fold) of P311 and DAP3 in invasive cells was confirmed by quantitative polymerase chain recation with reverse transcription (QRT-PCR) in six of seven and four of four additional GBM specimens, respectively. We found 2-fold and 5- to 80-fold overexpression of P311 and DAP3, respectively, by QRT-PCR and western blot analysis in GBM cell lines stimulated to migrate on cell-derived extracellular matrix (ECM) proteins. Immunostaining of glioblastoma specimens confirmed the presence of P311 and DAP3 at the invasive edge of the tumor. Treatment with antisense oligodeoxynucleotides directed against P311 and DAP3 messenger RNA caused a dose-dependent reduction in the migration rate of GBM cells on different substrates. The P311 and DAP3 proteins are overexpressed in invading human glioblastoma cells in vivo and in glioblastoma cell lines grown on cell-derived extracellular matrix in vitro. These proteins play an active role in migration on laminin- and cell-derived extracellular matrix, where the cells have a reduced ability to undergo apoptosis.