Abstract
Introduction: Glioblastoma (GBM) is the most devastating and incurable brain tumor. Although the identification of cells with tumor initiating mutations or their location can provide the fundamental basis for understanding disease progression, the origin of GBM remains controversial due to the lack of direct evidence in human GBM patients. Materials and Methods: Here, we performed ultra-deep sequencing of triple-matched tissues of i) radiologically and pathologically normal subventricular zone (SVZ), which is distant from tumor, ii) GBM tumor, and iii) blood (or normal cortical tissues) from patients with GBM (IDH-wildtype), compared to those with other type of brain tumors such as GBM (IDH-mutant), meningioma, oligodendrgolioma, metastatic brain tumor, and GBM (IDH-wildtype) with SVZ-invasion. Results: Surprisingly, we found that in 60% of GBM, IDH-wildtype patients (6 of 10), normal appearing and distant SVZ already contained the low level of GBM mutations such as TP53, PTEN, EGFR, PDGRF or TERT variations observed in the matched tumor. Single cell sequencing of GBM tumors and laser capture microdissection analysis of the SVZ show that mutations are enriched in GFAP-positive cells of the astrocyte ribbon area, which clonally evolved from the SVZ to the distant GBM tumor. Furthermore, using CRISPR-Cas9 system in the postnatal mouse brain, we showed that neural stem cells with TP53, PTEN, EGFR mutations migrated away from the mutated SVZ site and then formed the malignant glioma in the distant cortical region. Conclusions: Taken together, this study provides the direct evidence that human glioblastoma arises from the distant SVZ that is normal-appearing but harboring tumor-initiating mutations.