Abstract Alarmins are endogenous mediators that are elicited rapidly in response to danger signals, enhancing innate and adaptive immune responses by promoting the recruitment and maturation of antigen-presenting cells (APC). The nucleosome-binding protein HMGN1 is a potent alarmin that binds TLR4 and induces antigen-specific Th1 immune responses, but its contributions to antitumor immunity have not been explored. We found that ovalbumin (OVA)-expressing EG7 mouse thymoma cells grew much faster in Hmgn1-deficient mice than littermate-matched controls. Tumor-bearing Hmgn1−/− mice generated fewer OVA-specific CD8 cells in the spleen than EG7-bearing Hmgn1+/+ mice, suggesting that HMGN1 supported T cell–mediated antitumor immunity. In addition, EG7 tumors expressing HMGN1 grew more slowly than control EG7 tumors, suggesting greater resistance to HMGN1-expressing tumors. This resistance relied on T cell–mediated immunity because it was abolished by in vivo depletion of CD4+ and CD8+ T cells. Moreover, mice vaccinated with a DNA vector expressing an HMGN1–gp100 fusion protein manifested gp100-specific, Th1-polarized immune responses, acquiring resistance to challenge with mouse B16F1 melanoma. Overall, our findings show that HMGN1 contributes to antitumor immunity and it may offer an effective adjuvant to heighten responses to cancer vaccines. Cancer Res; 74(21); 5989–98. ©2014 AACR.