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Background Memory NKG2C + natural killer (NK) cells play a critical role in controlling human cytomegalovirus (HCMV) infection and are increasingly recognized for their potent antitumor activity. These long-lived NK cells exhibit enhanced cytotoxicity and cytokine production, contributing to immune defense against both viral infections and malignancies. Head and neck squamous cell carcinomas (HNSCC) rank among the most common cancers worldwide, with a poor 5-year survival rate of 10–50% in advanced stages. Despite improvements in therapy, treatment options for recurrent or metastatic disease remain limited. NK cells are essential for tumor surveillance; however, in HNSCC, they are highly susceptible to tumor-driven immune evasion, leading to impaired function and promoting both local and systemic disease progression. The transcriptional and functional characteristics of Memory NKG2C+ NK cells within the immunosuppressive HNSCC tumor microenvironment remain poorly defined. In our current study, we identify a significant enrichment of Memory NKG2C+ NK cells with enhanced effector function in regional lymph nodes of HNSCC patients.Methods Lymph node samples were collected from two patients undergoing resection for HNSCC, including both metastatic and non-metastatic nodes. To serve as comparative controls, four additional lymph nodes were obtained from cancer-free individuals: two HCMV + and two HCMV-. All samples were processed into single-cell suspensions for single-cell RNA sequencing (scRNA-seq). cDNA libraries were prepared, and sequenced, and raw reads were aligned to the human reference genome using Cell Ranger. After alignment, cells were filtered for quality and clustered based on uniquely expressed genes. Module scores of each subset differentially expressed genes were utilized to characterize the distinct subsets of NK cells.Results Unbiased clustering of NK cells from all lymph node samples revealed six distinct subsets, which were visualized using a Uniform Manifold Approximation and Projection plot. Regional metastatic lymph nodes contained a significantly higher proportion of functionally immature NK cells characterized by the expression of inhibitory and suppressor genes such as KLRC1, TIGIT, and LAG3. In contrast, non-metastatic regional lymph nodes demonstrated a significant enrichment of Memory NKG2C + NK cells, marked by elevated expression of activating receptors including CD16, NKG2C, IL7R and CD3e. Additionally, we identified a distinct transcriptional program in NK cells from metastatic lymph nodes, including a unique set of upregulated transcription factors associated with NK cell dysfunction.Conclusions These findings suggest a potential protective role of Memory NKG2C + NK cells in limiting metastatic spread and highlight their relevance as targets for therapeutic intervention in HNSCC.