Dual regulation of CXCR6(+)CD8(+) T cells modulates cytotoxic and exhaustion-associated programs during prostate cancer progression.

BACKGROUND: Prostate cancer (PCa) is widely recognized as an immunologically "cold" tumor, characterized by a paucity of effector T cells and a limited response to immune checkpoint blockade therapy. Although the chemokine receptor CXCR6 has been identified as a marker of highly cytotoxic CD8(+) T cells in other malignancies, its identity, regulatory mechanisms, and clinical significance in PCa remain poorly understood. METHODS: We integrated newly generated and publicly available single-cell RNA sequencing data encompassing 90 651 cells from nine patients. Multicolor immunofluorescence and flow cytometry were performed on human PCa tissue specimens stratified by Gleason Score (GS). In parallel, syngeneic mouse models, bone marrow chimeras, and in vitro T-cell functional assays were employed to investigate the role, recruitment, and transcriptional regulation of CXCR6(+)CD8(+) T cells. Mechanistic investigations included bulk RNA-seq, chromatin immunoprecipitation-quantitative real-time PCR, pharmacologic modulation of FOXO1, and detailed analysis of the IL-10-STAT3 signaling pathway. RESULTS: The CXCR6(+)CD8(+) T cells represent a transcriptionally distinct subset with high expression of cytotoxic markers (GZMA, GZMB, and PRF1), and their frequency declines significantly with increasing GS. Intratumoral maintenance of these cells is sustained by CXCL16 secreted by IL1B(+) M1-like macrophages; the loss of this macrophage population in advanced tumors leads to depletion of CXCR6(+)CD8(+) T cells. Genetic ablation of CXCR6 or depletion of CD8(+) T cells accelerates tumor growth, demonstrating the essential role of CXCR6(+)CD8(+) T cells in antitumor immunity. At the molecular level, the FOXO1-KLF2 axis transcriptionally represses CXCR6 expression, and IL-10-mediated activation of STAT3 upregulates FOXO1 and KLF2, thereby suppressing CXCR6 expression and impairing cytotoxic function. Pharmacologic inhibition of FOXO1 enhances the expansion of CXCR6(+)CD8(+) T cells and acts synergistically with anti-PD-1 therapy to inhibit tumor progression. CONCLUSIONS: The CXCR6(+)CD8(+) T cells are critical yet progressively diminished effectors in PCa. Their persistence within the tumor microenvironment depends on CXCL16 derived from M1-like macrophages and is counteracted by IL-10-driven FOXO1-KLF2 signaling. Targeting this regulatory axis-through inhibition of FOXO1 or IL-10, for example-represents a rational therapeutic strategy to restore CXCR6(+)CD8(+) T-cell-mediated immunity and enhance the efficacy of immunotherapy in PCa.