Genomic alterations and treatment define spatiotemporal patterns of glioblastoma recurrence

The surgical margin is the primary site of glioblastoma recurrence, yet its molecular features remain poorly characterized. We performed histology-guided sampling and single-cell spatial transcriptomics of surgical margin and adjacent recurrent tumor regions, with matched primary tumors from 40 patients. The surgical margin constituted a distinct wound-healing microenvironment in the post-treatment setting. We also identified malignant states with pronounced mesenchymal transcriptomic and histological features that were enriched at the surgical margin and defined a "stromal" subtype. Patients who transitioned to this subtype at recurrence were characterized by early TP53 mutation, sustained proliferation during adjuvant temozolomide treatment, and shorter survival. Conversely, patients with TP53-wild-type tumors recurring during temozolomide treatment transitioned toward the mesenchymal subtype, with increased p53 pathway activity and reduced proliferation. Functional validation showed TP53 loss-of-function increased temozolomide resistance in a patient-derived glioblastoma model. Our findings identify distinct glioblastoma recurrence trajectories linked to genomic context, treatment exposure, and clinical outcomes.