Background: Sipuleucel-T (sip-T) is an autologous therapeutic vaccine produced using a tumor antigen-cytokine fusion protein, and the only FDA approved cellular immunotherapy for metastatic castration-resistant prostate cancer (mCRPC) patients. Sip-T significantly improves overall survival (OS), but has limited impact on PSA and radiographic responses. Here, we present the first high dimensional analysis of the sip-T product; we used a preclinical model of human prostate cancer to show the effects of IL-15 on anti-tumor efficacy, tumor infiltration of effector cells, and reversal of immunosuppressive mediators. Methods: We performed a comprehensive assessment of the sip-T product collected from prostate cancer patients using mass cytometry (CyTOF). Control and IL-15 stimulated sip-T were evaluated, and changes in leukocyte subsets as well as markers of activation and exhaustion were identified. Using a preclinical mouse model of human prostate cancer, we examined the effects of IL-15 treatment on sip-T. CyTOF, flow cytometry, RNAseq, and qPCR were used to evaluate the effects of IL-15 on sip-T phenotype, function, and in vitro and in vivo efficacy. Results: CyTOF analysis revealed that CD3+ T cells constituted the highest proportion of sip-T, followed by B-cells, natural killer (NK) cells, NKT, and monocytes, with only a small percentage of dendritic cells. Following sip-T stimulation with IL-15, a significant expansion and activation of CD8+ T-cell and NK cell populations was seen. Co-culture of sip-T with IL-15 and control or prostate-relevant antigens showed significant activation and expansion of CD8 T and NKT cells in an antigen-specific manner, with significantly increased tumor cytotoxicity and evidence of antigen spread. IL-15 treatment combined with sip-T in a preclinical mouse model showed significant suppression of tumor growth compared to sip-T alone, with increased tumor-infiltrating lymphocytes (TIL). A significant increase in interferon (IFN)-γ producing CD8+ T and NKT cells within the tumor microenvironment was seen in the IL-15 group. Tumor transcriptomic analyses revealed IL-15 treatment was able to reverse immunoresistance, with significant decreases in PD-L1, CTLA, TIGIT, B7-H3, and others compared to sip-T alone. Conclusions: This is the first study to evaluate the sip-T product using CyTOF, and the first pre-clinical in vivo prostate tumor modeling of sip-T. IL-15 treatment significantly enhances anti-tumor efficacy, effector immune cell activation and tumor infiltration, and reverses key mediators of immune suppression. Studies with modified IL-15 cytokines are ongoing and will be presented.