CD8+ T-NK cell crosstalk establishes preemptive immunosurveillance to eliminate antigen-escape tumors.

Background and objective: Tumor antigen-escape variants undermine immunotherapy by subverting lymphocyte effector functions and reshaping tumor-immune dynamics. It is essential to delineate functional interplay within immune networks during tumor progression. We investigated whether homeostatic crosstalk between CD8+T cells and natural killer (NK) cells preempts tumor antigen-escape. Methods: Adoptive CD8+T cell transfers were administered before (D-7, homeostatic pre-priming) or after (D+1) tumor establishment in Rag1-/- and Rag1-/-γc-/- mice. Antigen presentation, immune activation, proliferation, cytotoxicity, and memory were quantified by flow cytometry, live bioluminescence and confocal imaging. Monoculture, co-culture, and a 3D silica nanofiber carpet mimicking basement-membrane-like topography modeled intercellular interactions. Signaling arrays and motion metrics (Speed-Distance Index, deceleration) were conducted. Human ligand-receptor pairs engaged in CD8+T-NK crosstalk were probed in silico. Results and discussion: Pre-tumor D-7 CD8+T cell transfer completely suppressed antigen-escape tumors with NK cells as major effectors showing elevated CD25, CD69, CD107a, and GzmB, marking activated and effector phenotype, and promoting central-memory CD62L⁺CD44⁺CD8⁺TCM precursors. By contrast, post-tumor D+1-transferred CD8+T cells allowed emergence of tumor variants resistant to antigen-specific cytolysis as assessed on day 25, despite those T cells retaining higher intrinsic cytotoxic capacity than the D-7 T cell cohort. Mechanistically, CD8+T and NK cells formed stable contacts through pseudopodial intercellular nanotubes enabling bidirectional membrane exchange and signaling via STAT, Akt, and mTOR pathways, augmenting NK effector function and promoting CD8+TCM differentiation. In silico analysis identified human ligand-receptor pairs engaged in CD8+T-NK adhesion, stimulatory and regulatory axes, including CD200-CD200R, PD-L1-PD-1, and CD18/CD11a-DNAM-1 (CD226). Together, data support a three-phase model of preemptive immunosurveillance initiated by early CD8⁺T-NK crosstalk. Conclusion: Homeostatic conditioning and effector cooperativity between CD8+T and NK cells protect against tumor immune escape. The findings uncover a mechanistic axis of preemptive immunosurveillance that lays the foundation for next-generation preventive immunotherapies to control antigen-escape tumors.