Strategies to activate and rescue exhausted tumor-specific T cells, including the use of monoclonal antibodies (mAb) that block the negative costimulatory receptors CTLA-4 and PD-1 are proving very effective, but TIM3 has been relatively neglected as a target. Here we report an extensive characterization of the therapeutic activity and mechanism of action of an anti-mouse TIM3 mAb against experimental and carcinogen-induced tumors. For the first time we specifically define the mechanism of antitumor action of anti-TIM3 requiring IFN-γ producing CD8⁺ T cells and CD4⁺ T cells, and a higher ratio of tumor infiltrating CD8⁺:CD4⁺ T cells correlating with therapeutic success. Interestingly, in some models, anti-TIM3 appeared to be effective sometime before the appearance and accumulation of significant TIM3⁺PD-1⁺ T cell populations in tumor bearing mice. Anti-TIM3 displayed modest prophylactic and therapeutic activity against a small fraction of carcinogen-induced sarcomas, but comparative and combination studies of anti-TIM3 with anti-CTLA-4 and anti–PD-1 against experimental and carcinogen-induced tumors suggested that these agents might be well-tolerated and very effective in combination. Cancer Res; 71(10); 3540–51. ©2011 AACR.