Alpha-tocopheryl succinate (TOS), a vitamin E analog, is a promising anticancer agent due to its abilities to inhibit proliferation and to induce apoptosis in a variety of human malignant cell lines, while being relatively less active toward normal cells. However, the molecular mechanisms underlying the apoptotic effects of TOS are not precisely understood. Reports that TOS can generate reactive oxygen species (ROS) prompted us to investigate the role of ROS in TOS-induced apoptosis in cancer cells. We found that the human lung cancer A549 and H460 cell lines were much more sensitive to TOS-induced apoptosis than the human glioblastoma T98G and U87MG cell lines. Our data suggested that the differential TOS sensitivity was not caused by differences in the uptake and retention of TOS between TOS-sensitive and -resistant cancer cells. The differential ability of cancer cells to generate ROS in response to TOS appears to be an important factor in determining the susceptibility of cells to TOS-induced apoptosis. Our results further suggest that TOS-induced generation of ROS is involved in caspase-independent apoptosis. Taken together, our findings suggest an important role of ROS generation in TOS-induced, caspase-independent apoptosis of cancer cells.