Cell-Intrinsic MHC-I Expression Sustains NK Cell Effector Function and Its Loss Drives Exhaustion in Triple-Negative Breast Cancer

Abstract

Triple-negative breast cancer (TNBC), characterized by lack of targetable receptors and an immunosuppressive tumor microenvironment (TME), remains one of the most aggressive breast cancer subtypes with poor prognosis despite emerging immunotherapies. While adaptive immunity has been extensively studied, the role of innate immunity, particularly natural killer (NK) cells, is increasingly recognized. Although NK cells conventionally eliminate MHC-I-deficient tumor cells via "missing-self" recognition, their phenotype in TNBC TME is often compromised, with mechanisms remaining unclear. Here, we combined single-cell RNA sequencing analysis of human TNBC samples with functional in vitro assays to investigate tumor-infiltrating NK cells. Our analysis revealed that TNBC epithelial cells exhibit enhanced MHC-I expression compared to normal breast tissue. However, TNBC-infiltrating NK cells showed high expression of both effector molecules (GNLY, NKG7, PRF1) and inhibitory/exhaustion markers (HAVCR2, LAIR1, TGFB1), suggesting a chronically activated but dysfunctional state. To explore MHC-I function, we established an in vitro model knocking down B2M and HLA-A in NK-92 cells using siRNA. Results demonstrated that MHC-I downregulation does not affect NK cell proliferation but negatively regulates effector function, resulting in reduced degranulation (CD107a), cytokine production (IFN-γ, Granzyme B), and target cell killing. These findings reveal a previously unrecognized cell-autonomous role of MHC-I in sustaining NK cell functionality. Our study identifies NK cell-intrinsic MHC-I expression as a critical regulator of anti-tumor immunity and proposes MHC-I upregulation on NK cells as a potential next-generation immunotherapy strategy for immune checkpoint-resistant TNBC.