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Thyroid cancer is one of the malignancies with better clinical outcomes. However, a minority of patients develops an aggressive anaplastic thyroid carcinoma. Development of innovative and multimodal therapeutic strategies is urgently needed. Here, we investigated the role of CXCR5+ CD8 T cells in the peripheral blood, tumor-involved lymph nodes (TILN), and tumor mass of thyroid cancer patients. In peripheral blood mononuclear cells, CXCR5+ cells represented 1.4% ± 0.84% (mean ± s.d.) of total CD8 T cells, while in TILN and in tumor, the frequencies of CXCR5+ CD8 T cells were significantly higher at 27.7% ± 7.8% and 15.5% ± 2.9%, respectively. Compared to CXCR5− CD8 T cells, CXCR5+ CD8 T cells presented significantly higher PD-1 expression and lower or comparable TIM-3 and CTLA-4 expression. To compare and contrast the functional characteristics of CXCR5+ CD8 T cells and CXCR5− CD8 T cells, these cells were separated from TILNs and were TCR-stimulated via anti-CD3/CD28. Upon stimulation, CXCR5+ CD8 T cells presented stronger downregulation of CD27, higher expression of proinflammatory cytokines IL-2, IFN-γ, and TNF-α, and higher proliferation capacity than CXCR5− CD8 T cells. Moreover, CXCR5+ CD8 T cells presented higher expression of cytotoxic molecules Gzm-A, Gzm-B, and perforin. Overall, these results demonstrated that in thyroid cancer patients CXCR5+ CD8 T cells infiltrated the TILNs and the tumors, and were functionally more potent compared to their CXCR5− counterpart.Frequencies of CXCR5+ CD8 T cells were significantly increased in TILN and tumors.CXCR5+ CD8 T cells presented significantly higher PD-1 expression than counterparts.CXCR5+ CD8 T cells presented high expression of IL-2, IFN-γ, and TNF-α.CXCR5+ CD8 T cells presented higher proliferation capacity than CXCR5− CD8 T cells.CXCR5+ CD8 T cells presented high levels of cytotoxic molecules and perforin.