The efficacy of T cell–based immunotherapy to treat cancer patients remains a challenge partly because of the weak activity toward subdominant tumor antigens (TAg) and to tumors expressing suboptimal TAg levels. Recent reports indicate that Toll-like receptor (TLR) stimulation on T cells can lower the activation threshold. In this study, we examined the antitumor activity and survival of TLR2-MyD88–stimulated CD8 T cells derived from melanoma patients and T-cell receptor transgenic pmel mice. TLR2-stimulated pmel CD8 T cells, but not TLR2^−/−pmel or MyD88^−/−pmel T cells, responded to significantly lower TAg levels and resulted in increased production of effector molecules and cytotoxicity. Wild-type or MyD88^−/− mice treated with TLR2 ligand and pmel T cells, but not TLR2^−/−pmel or MyD88^−/−pmel T cells, showed tumor regression of an established melanoma tumor. Overexpressing TLR2 in TAg-specific T cells eradicated tumors; four times fewer cells were needed to generate antitumor responses. The enhanced antitumor activity of TLR2-MyD88–stimulated T cells was associated with increased effector function but perhaps more importantly with improved survival of T cells. Activating TLR-MyD88 signals in patient-derived T cells also reduced the activation threshold to several weakly immunogenic TAgs, resulting in increased cytokine production, expansion, and cytotoxicity. These data highlight a previously unappreciated role for activating TLR-MyD88 signals in tumor-reactive T lymphocytes. Cancer Res; 70(19); 7442–54. ©2010 AACR.