Abstract Purpose: Therapeutic blockade of immune checkpoints has revolutionized cancer treatment. Durable responses, however, occur in less than half of those treated, and efforts to improve treatment efficacy are confounded by a lack of understanding of the characteristics of the cells that initiate antitumor immune response. Patients and Methods: We performed multiparameter flow cytometry and quantitative multiplex immunofluorescence staining on tumor specimens from immunotherapy-naïve melanoma patients and longitudinal biopsy specimen obtained from patients undergoing anti–PD-1 therapy. Results: Increased numbers of CD69+CD103+ tumor-resident CD8+ T cells were associated with improved melanoma-specific survival in immunotherapy-naïve melanoma patients. Local IL15 expression levels strongly correlated with these tumor-resident T-cell numbers. The expression of several immune checkpoints including PD-1 and LAG3 was highly enriched in this subset, and these cells significantly expanded early during anti–PD-1 immunotherapy. Conclusions: Tumor-resident CD8+ T-cell numbers are more prognostic than total CD8+ T cells in metastatic melanoma. In addition, they are likely to initiate response to anti–PD-1 and anti–LAG-3 treatments. We propose that the immune profile of these cells prior to treatment could inform strategies for immune checkpoint blockade. Clin Cancer Res; 24(13); 3036–45. ©2018 AACR.