Multiomics Dissection Reveals Natural Killer Cell–CD8+ T-cell Cooperation in Shaping an Immunostimulatory Tumor Microenvironment

Immune evasion remains a major barrier to effective immunotherapy in cancer. Coordinated activation of different immune cell types shapes immune responses within the tumor microenvironment (TME). In this study, we uncovered a spatiotemporal synergy between cytotoxic T lymphocytes (CTL) and natural killer (NK) cells in tumors with low T-cell infiltration. An integrative multiomics approach combining spatial transcriptomics, single-cell RNA sequencing, and high-dimensional flow cytometry demonstrated that vaccination, capable of activating both innate and adaptive immunity, rapidly reshapes the TME. Early infiltration of IRF8+KLRG1+ NK cells established a chemokine-rich niche together with myeloid cells that promoted the recruitment of antigen-primed CXCR3+ CTLs. The accompanying inflammatory monocyte recruitment and dendritic cell activation led to the coordinated structural and chemotactic remodeling of the TME. Functionally, NK-CTL clustering at the tumor margin was associated with enhanced cytotoxic activity and sustained immune engagement. These spatially organized immune interactions involved CCR5-CCL5 and CXCR3-CXCL9 signaling pathways that coordinate communication between innate and adaptive immune compartments. Together, these findings reveal a previously unrecognized NK-CTL cooperative program that promotes the transition of poorly infiltrated tumors toward an inflamed, immune-responsive state. More broadly, this study illustrates how spatially resolved multiomics approaches can uncover immune interactions and provides a conceptual framework for designing next-generation immunotherapies that mobilize coordinated innate and adaptive immunity. SIGNIFICANCE: Early infiltration of NK cells followed by T cell accumulation reshapes the tumor microenvironment to promote antitumor immunity following vaccination, providing a mechanistic rationale for the development of effective vaccine-based immunotherapy strategies.