Immune pressure drives HLA-E-defined metastatic evolution in hepatocellular carcinoma

Hepatocellular carcinoma (HCC) almost invariably develops in chronic hepatitis and cirrhotic livers, where sustained cytotoxic immune activity imposes persistent selective pressure on malignant hepatocytes. While acute anti-tumor immunity restrains tumor growth, whether chronic cytotoxic immune pressure actively programs metastatic initiation in HCC remains unresolved. We integrated multi-regional single-cell RNA sequencing profiles from non-tumor liver tissues, primary tumors, portal vein tumor thrombi, and distant metastases across three cohorts. Malignant hepatocytes were inferred by copy number profiling, and metastatic trajectories were reconstructed by Monocle pseudotime, followed by gene set scoring to localize metastasis-initiating programs in situ . We stimulated HCC cell lines with interferon-gamma (IFN-γ) and quantified human leukocyte antigen E (HLA-E) by quantitative PCR, immunoblotting, and flow cytometry, with Janus kinase–signal transducer and activator of transcription (JAK–STAT) perturbation using ruxolitinib and STAT3 knockdown. STAT3-dependent transcriptional control was assessed by CUT&RUN (cleavage under targets and release using nuclease) and dual-luciferase reporter assays. Metastatic function was tested in orthotopic Hepa1-6 liver implantation with IFN-γ administration and lung hematoxylin and eosin (H&E) quantification. An immune-associated malignant hepatocyte subpopulation carried the highest metastasis-initiating signature and represented the dominant source of metastatic seed cells. It was selectively marked by HLA-E, a non-classical major histocompatibility complex (MHC) class Ib molecule, whose expression was associated with immune pressure and tightly correlated with metastatic competence. Silencing of HLA-E/H2-T23 suppressed lung metastasis and prolonged survival in vivo and abrogated IFN-γ-driven metastatic acceleration. Mechanistically, IFN-γ induced HLA-E/H2-T23 through non-canonical JAK–STAT3 activation. Our findings demonstrate that chronic cytotoxic immune pressure actively instructs metastatic evolution in HCC by driving an HLA-E-defined immune-adaptive malignant hepatocyte state.