Abstract A major mechanism by which human regulatory T cells (Treg) have been shown to suppress and kill autologous immune cells is through the granzyme-perforin pathway. However, it is unknown whether Tregs also possess the capacity to kill tumor cells using similar mechanisms. Bispecific antibodies (bscAb) have emerged as a promising class of therapeutics that activate T cells against tumor antigens without the need for classical MHC-restricted T-cell receptor (TCR) recognition. Here, we show that a bscAb targeting the tumor-specific mutation of the EGF receptor, EGFRvIII, redirects human CD4+CD25+FoxP3+ Tregs to kill glioblastoma cells. This activity was significantly abrogated by inhibitors of the granzyme-perforin pathway. Notably, analyses of human primary glioblastoma also displayed diffused infiltration of granzyme-expressing FoxP3+ T cells. Together, these data suggest that despite their known suppressive functions, tumor-infiltrating Tregs possess potent cytotoxic mechanisms that can be co-opted for efficient tumor cell lysis. Cancer Immunol Res; 1(3); 163–7. ©2013 AACR.