The apoptotic effects of 2-amino-4,4alpha-dihydro-4alpha, 7-dimethyl-3H-phenoxazine-3-one (Phx-1) and 2-aminophenoxazine-3-one (Phx-3) on human glioblastoma cell lines, A-172 and U-251 MG were studied. These phenoxazines extensively decreased the viability of A-172 and U-251 MG cells (IC50 of Phx-1: 60 microM, in both lines; IC50 of Phx-3: 10 and 3 microM, for A-172 and U-251 cells, respectively). Phx-1 and Phx-3 increased the population of annexin V and PI double-positive cells in A-172 and U-251 MG cells, resulting in cell death at late stage apoptosis/necrosis. The activities of caspase-3/7 were greatly increased in A-172 and U-251 MG cells treated with Phx-1 or Phx-3. However, a pan-caspase inhibitor, z-VAD-fmk, failed to reverse the antiproliferative and apoptotic effects of Phx-1 and Phx-3 in both cell lines. In conclusion, Phx-1 and Phx-3 exert significant anti-cancer effects against human glioblastoma cell lines, A-172 and U-251 MG, mediated by the caspase-independent apoptotic cell death pathway.