Genes encoding 2C T cell receptor (TCR) α, β chains from H-2b-re-stricted Ld-specific CD8+ cells were successfully transduced into polyclonally activated CD8+ cells by retroviral modification to generate antigen-specific cytotoxic T lymphocytes (CTL). Antigen-nonspecific CD8+ T cells polyclonally expanded in the presence of interleukin (IL)-2, Th1 cytokines (interferon (IFN)-γ and IL-12) and anti-IL-4 monoclonal antibody showed neither cytokine production nor cytotoxicity in response to Ld-expressing P815 tumor cells. However, 2C-TCR gene-modified CD8+ T cells exhibited both IFN-γ production and cytotoxicity in response to P815 tumor cells. The antitumor activity of TCR gene-modified Tc1 cells was also demonstrated in vivo by Winn's assay. Thus, we have developed an efficient method to induce TCR gene-modified antigen-specific Tc1 cells that exhibit antitumor activity both in vitro and in vivo. (Cancer Sci 2003; 94: 389–393)