Cytotoxic lymphocyte heterogeneity in glioblastoma: insights from single-cell and spatial multiomics toward precision immunotherapeutic reprogramming.

Glioblastoma (GBM) represents the most aggressive primary brain tumor in adults, characterized by a profoundly immunosuppressive tumor microenvironment (TME) that systematically disables cytotoxic lymphocyte function and renders conventional immunotherapy largely ineffective. While exhaustion of CD8+ T cells and natural killer (NK) cells within solid tumors has been extensively studied in other cancer types, the CNS-specific architectural, metabolic, and molecular constraints that shape cytotoxic lymphocyte heterogeneity in GBM remain insufficiently characterized. Recent advances in single-cell RNA sequencing (scRNA-seq) and spatial multiomics have begun to reveal a rich landscape of cytotoxic lymphocyte subpopulations in GBM. These include TCF-1+ progenitor-exhausted T cells (Tpex), terminally exhausted CD8+ T cells (Tex), and dysfunctional natural killer (NK) cell subsets, each distributed across anatomically distinct immune niches. This review synthesizes current knowledge across three interconnected areas: the single-cell atlas of GBM-infiltrating cytotoxic lymphocytes; the spatial organization of their dysfunction within perinecrotic, perivascular, and infiltrative-edge niches; and the epigenetic and transcriptional programs that underlie GBM-specific cytotoxic failure, including dysregulation of the TOX/TCF-1 axis and IDH-mutation-driven silencing of NKG2D ligands. Critically, we compare CD8+ T cell and NK cell exhaustion mechanisms, highlighting their mechanistic divergence and therapeutic implications. We further discuss how these multiomics insights can be translated into neurosurgically relevant strategies, including intraoperative tumor profiling, progenitor T cell expansion via epigenetic priming, NKG2A/TIGIT dual blockade, and intracavitary delivery of engineered NK cells. Together, this review proposes a spatially and cellularly resolved framework for understanding cytotoxic immune failure in GBM and outlines precision immunotherapy approaches tailored to the unique immunobiology of the CNS tumor microenvironment.