Spatially restricted iron-metabolism-associated stress programs prime invasive mesenchymal glioblastoma states in human tissue-context models

Glioblastoma (GBM) recurrence is driven by invasive tumor cells that infiltrate surrounding brain tissue and evade surgical and therapeutic eradication. While dysregulated iron handling is a well-recognized feature of malignant cancers, its contribution to invasive GBM cell states in patient derived models remains poorly defined. We performed multi-regional single-nucleus RNA sequencing of human GBM specimens encompassing tumor core, tumor periphery, and infiltrated cortex to map iron-associated transcriptional programs validated by spatial transcriptomics. Correlative analyses were combined with functional validation using patient-derived GBM cell lines and human organotypic cortical slice cultures exposed to non-cytotoxic iron supplementation to model iron-rich tumor microenvironments. Malignant cells from the tumor core exhibited coordinated upregulation of iron uptake and storage pathways alongside invasion-associated gene programs. At single-cell resolution, iron metabolism and invasion signatures were correlated, defining a core-enriched malignant subpopulation with mesenchymal-like transcriptional identity, stress-adaptive features, and angiogenic signaling that aggregate in specific spatial niches. Functionally, iron exposure altered migration in a cell-state-dependent manner and increased tumor growth and invasion in human cortical slice cultures. Increased VIM, MMP9, and HIF1A expression was consistent with activation of mesenchymal-like and stress-response programs, although mechanistic dependence was not tested. Iron-handling and invasion-associated programs co-occur in a core-enriched mesenchymal-like GBM state. In patient-derived cell and human cortical slice models, non-cytotoxic particulate and soluble iron altered migration in a cell-state-dependent manner and increased tissue invasion. These findings support iron availability as one component of the microenvironment associated with invasive GBM phenotypes, while its necessity and underlying mechanism remain to be established.