Natural Killer Cell-Derived Extracellular Vesicles Reprogram Human Immunity to Enhance Tumour Cytotoxicity

Despite evidence of natural killer cell–derived extracellular vesicles (NK-EVs) possessing intrinsic cytotoxicity, their broader immunomodulatory effects on human immune subsets remain poorly defined, representing a key knowledge gap that must be addressed to advance their clinical translation as cancer immunotherapeutics. In this study, the immunomodulatory effects of NK-EVs were evaluated on peripheral blood mononuclear cells (PBMCs), a mixed population of circulating immune cells, from healthy donors and patients with aggressive breast cancer. NK-EVs were produced through a clinically scalable production process and PBMCs exposed to NK-EVs were assessed using high-resolution single-cell transcriptomics and functional cytotoxicity assays. NK-EVs reprogrammed PBMC on a transcriptomic (gene regulation) and cellular level, resulting in functional improvement of the PBMCs' tumour cell killing capabilities. These effects were mediated through distinct PBMC subsets, including cytotoxic T cells, NK cells and myeloid cells. Interestingly, CD8+ T cells, CD56+ NK cells, and CD33+ myeloid cells appear to play a more prominent role in NK-EV-mediated immune reprogramming than CD4+ T cells, with genes involved in their activation being markedly affected upon EV stimulation. Notably, the responses were consistent across all biological PBMC donors (n=6), regardless of disease status. Collectively, these findings demonstrate that NK-EVs orchestrate coordinated immune activation, highlighting their promise as an immunotherapeutic for cancer and other immune-mediated diseases.