NKTR-214 is a first-in-class, CD122-preferential IL-2 pathway agonist that provides sustained activation of the IL-2 pathway via IL-2R beta chain-biased signaling, selectively stimulating CD8+ T cells over regulatory T cells (Tregs), which require binding to the IL-2R alpha chain. NKTR-214 has proven safe and effective in increasing CD8+ T cells in the circulation and in tumor tissue in patients with a variety of cancer, including melanoma, renal, lung, bladder and breast cancer. Interestingly, NKTR-214 dramatically decreased Treg levels in tumors, but not in the periphery. To better understand this phenomenon, we used a model of gp100 melanoma antigen-specific vaccination and gp100-specific pmel-1 T cells to study the impact of NKTR-214 treatment on CD8+ Teff and CD4+Foxp3+ Tregs. We found that NKTR-214 induced CD8+ T cell accumulation in tumor tissue as well as Treg depletion in tumor tissue but not in the circulation. Mechanistically, NKTR-214 treatment favored proliferation and survival of effector CD8+ T cells while reducing proliferation and promoting apoptosis of intratumoral Tregs. In vivo cytokine neutralization experiments and in vitro assays together with proteomic analysis of Tregs revealed that NKTR-214 plus vaccine treatment induced potent release of CD8+ T cell-associated cytokines IFN-γ and TNF-α, which synergized to induce Treg proliferative block and apoptosis by inhibiting IL-2 and TCR-downstream signaling. These results shed light on the mechanism of action of NKTR-214 and point to a mechanism of increased CD8+ T cell influx and temporal, localized Treg depletion at sites of strong effector T cell activity.