β-endorphin primes Natural Killer cells and NK-derived Extracellular Vesicle to enhance anti-tumor cytotoxicity

Abstract Psychoneuroimmunology suggests that positive physiological states, including laughter, could affect anti-tumor immunity, but the underlying mechanisms remain unclear. Here, we investigated whether β-endorphin (BE), an endogenous opioid peptide associated with positive physiological stimuli, modulates Natural Killer (NK) cell cytotoxicity and the anti-tumor activity of NK-derived extracellular vesicles (EVs). Using NK-92 cells, we assessed cytotoxicity against JIMT1 breast cancer cells, CD107a mobilization, cytotoxic activity of conditioned medium (CM), and EV yield, cargo, and function. BE enhanced NK-92-mediated killing of JIMT1 cells without increasing CD107a mobilization, suggesting that improved cytotoxicity was not driven by classical degranulation. Consistently, CM from BE-treated NK cells retained contact-independent cytotoxicity. NK-EVs were enriched in granzyme B and perforin following BE treatment exhibiting enhanced cytotoxicity against JIMT1 and BW tumor cells. BE also increased the cytotoxic activity of primary human NK cells, and BE-conditioned NK-EVs primed naïve NK-92 cells for enhanced tumor killing. These findings indicate that BE enhances NK anti-tumor immunity by remodeling the cytotoxic secretome and generating EVs that act as both direct cytotoxic effectors and mediators of NK cell priming. Graphical abstract Proposed neuroendocrine–immune model linking positive physiological stimuli, NK cell-derived extracellular vesicles (EVs), and anti-tumor activity. Laughter is depicted as a conceptual upstream trigger of hypothalamic–pituitary signaling leading to β-endorphin (BE) release. BE conditioning enhanced NK-92 cytotoxicity and the anti-tumor activity of NK-derived EVs, consistent with granzyme B and perforin enrichment and supporting EV-mediated contact-independent cancer cell killing.