Abstract The current study demonstrates that the combination of a DNA vaccine encoding the chemokine MIP-3α fused to melanoma antigen gp100 with immunomodulatory antibodies prolongs survival in a melanoma mouse model. The current studies utilize the B16F10 mouse spontaneous melanoma syngeneic transplantable mouse model system. Previous published work with this vaccine platform has indicated the MIP-3α component targets nascent peptides to immature dendritic cells leading to processing by class I and class II MHC pathways. It has further been shown in this tumor system that prophylactic vaccination with the DNA construct alone delayed tumor growth. Other studies have provided evidence that αPD-1 or αIL-10 neutralizing antibodies enhance immunological melanoma therapies by modulating the tolerogenic tumor microenvironment. First, this study expanded previous results by finding a significant decrease in tumor burden and increase in survival by prophylactic intramuscular electroporation [IM EP] of the DNA vaccine construct. Adding three therapeutic doses of αIL-10 given subcutaneously [subQ] at the tumor site to the treatment further increased mouse survival. The development of cell-mediated and humoral immune responses were documented for the first time in this system by Elispot and in-cell ELISA assays. Secondly, a novel therapeutic vaccination protocol was developed by initiating vaccination and antibody therapies after tumor challenge. With this therapeutic protocol, we demonstrated for the first time that vaccine significantly delays tumor burden and increases median survival from 17 to 21 days (p = 0.006). Addition of αIL-10 (subQ) and αPD-1 (intra-peritoneally) antibodies led to further highly significant decreases in tumor burden and increased median survival from 17 to 25 days (p = 0.0005). Efficient targeting of antigen to immature dendritic cells with a chemokine fusion vaccine offers a potential alternative approach to the ex vivo dendritic cell antigen loading protocols currently undergoing clinical investigation. Combining this approach with agents able to modulate the tolerogenic tumor microenvironment offers promise as a novel melanoma therapy. Citation Format: James Gordy, Richard B. Markham. Therapeutic dendritic cell targeting MIP3α-gp100 DNA vaccination with immunomodulatory αIL-10 and αPD-1 antibodies significantly enhances survival in a mouse melanoma model system. [abstract]. In: Proceedings of the 106th Annual Meeting of the American Association for Cancer Research; 2015 Apr 18-22; Philadelphia, PA. Philadelphia (PA): AACR; Cancer Res 2015;75(15 Suppl):Abstract nr 2511. doi:10.1158/1538-7445.AM2015-2511