Targeting HLA-E-NKG2A/CD94 to reverse pyroptosis-induced immunosuppression in HCC enhances NK cell antitumor activity

Background: The efficacy of adoptive cell therapy in hepatocellular carcinoma is limited by the immunosuppressive tumor microenvironment, yet the underlying mechanisms remain incompletely understood. Methods: Clinical data from hepatocellular carcinoma patients receiving NK cell therapy were analyzed alongside tumor specimens and mouse xenograft models. Immune cell composition and spatial distribution were evaluated using immunohistochemistry and immunofluorescence. In vitro assays, including flow cytometry, qPCR, Western blot, ELISA, and real-time cytotoxicity analysis, were performed to investigate macrophage polarization and NK cell function. RNA sequencing was used to identify key regulatory molecules, followed by functional validation and pharmacological blockade in vitro and in vivo . Results: NK cell therapy was associated with improved patient outcomes. However, tumor progression may resume upon treatment discontinuation. Tumor infiltrating NK cells positively correlated with CD163⁺ M2 macrophages. Mechanistically, NK cell mediated tumor killing induced tumor cell pyroptosis and triggered the release of interleukin-10, which promoted macrophage polarization toward an M2 phenotype via a STAT3-dependent positive feedback loop. These M2 macrophages exhibited high expression of HLA-E and suppressed NK cell cytotoxicity and cytokine production through a contact-dependent HLA-E-NKG2A/CD94 axis. Blockade of this pathway effectively restored NK cell function and enhanced antitumor efficacy both in vitro and in vivo. Conclusions: This study identifies a negative feedback loop in which NK cell induced tumor pyroptosis promotes IL-10 mediated M2 macrophage polarization, leading to subsequent suppression of NK cell function. Targeting the HLA-E-NKG2A/CD94 axis represents a promising strategy to overcome immune suppression and improve the efficacy of NK cell-based immunotherapy in hepatocellular carcinoma.