NKG2A-HLA-E axis contributes to an immunosuppressive checkpoint network in hepatocellular carcinoma

Background The immunosuppressive tumor microenvironment in hepatocellular carcinoma (HCC) limits the efficacy of current immunotherapies. The interplay between the emerging inhibitory receptor NKG2A and its activating counterpart NKG2C, both binding HLA-E, and its interaction with the PD-1-PD-L1 pathway in cancer remains unclear. This study aims to delineate the functional network formed by these pathways in shaping the immunosuppressive landscape of HCC. Methods We analyzed public databases and performed immunohistochemistry on HCC tissues to assess HLA-E expression and prognostic relevance. Tumor-infiltrating lymphocytes from patient samples were examined by flow cytometry to evaluate the functional impact of NKG2A, NKG2C, and PD-1 expression on T and NK cells. Single-cell RNA and T cell receptor (TCR) sequencing were employed to characterize immune subset heterogeneity. Functional assays and mouse models were used to evaluate the effects of NKG2A and PD-1 blockade. Results In the absence of CD8 + T cell infiltration, elevated HLA-E expression in HCC tissue correlated with poorer overall survival in non-immunotherapy-treated patients. Conversely, in the presence of CD8 + T cells, high HLA-E expression was linked to an inflammatory tumor microenvironment characterized by IFN-γ-producing T cells, and increased major histocompatibility complex I, which was associated with improved patient response to immunotherapy. Tumor-infiltrating CD8 + T and NK cells from HCC patients upregulated NKG2A, conferring tissue-resident memory and tumor-reactive characteristics while simultaneously impairing cytotoxic function. Single-cell RNA and TCR sequencing revealed differential expression patterns of NKG2A, NKG2C, and PD-1 in different T and NK cell subsets, with NKG2A + PD-1 + T cells relevant to antigen-specific clonal expansion. Mechanistically, the inhibitory effect mediated by the NKG2A-HLA-E axis contributed more prominently than the stimulatory role of the NKG2C-HLA-E axis in suppressing antitumor immunity. Furthermore, the NKG2A-HLA-E axis acted cooperatively with the PD-1-PD-L1 axis to promote CD8 + T cell dysfunction. Combined blockade of NKG2A and PD-1 enhanced the cytotoxic activity of tumor-infiltrating CD8 + T and NK cells, resulting in potent tumor eradication in vitro and in vivo, outperforming single-agent treatments. Conclusions Our findings identify the NKG2A-HLA-E axis as an important immunoregulatory pathway associated with immune suppression in HCC and provide a rationale for further evaluation of combined NKG2A and PD-1 blockade strategies.