This study addresses the treatment failure in glioblastoma (GBM) due to intratumoral heterogeneity and tumor evolution. The authors utilized 3D surgical neuronavigation to acquire spatially mapped samples from 10 patients with IDH-wildtype GBM, providing a comprehensive view of the tumor's spatial distribution and heterogeneity. Integrative tissue and single-cell analyses revealed genomic, epigenomic, and microenvironmental intratumoral heterogeneity, including oncogene amplifications, tumor suppressor deletions, and structural variants. The study identified distinct evolutionary trajectories, such as chromothripsis and genetic subclones, and their spatial patterns within the tumor. The authors also explored the transcriptomic and chromatin landscapes, uncovering neurodevelopmental programs that reflect GBM origins and contribute to heterogeneity. The findings highlight potential therapeutic targets and provide insights into GBM evolution and heterogeneity, offering a 3D whole-tumor perspective that can inform clinical care and outcomes. The 3D spatially resolved maps of GBM tumors are publicly available for further research.This study addresses the treatment failure in glioblastoma (GBM) due to intratumoral heterogeneity and tumor evolution. The authors utilized 3D surgical neuronavigation to acquire spatially mapped samples from 10 patients with IDH-wildtype GBM, providing a comprehensive view of the tumor's spatial distribution and heterogeneity. Integrative tissue and single-cell analyses revealed genomic, epigenomic, and microenvironmental intratumoral heterogeneity, including oncogene amplifications, tumor suppressor deletions, and structural variants. The study identified distinct evolutionary trajectories, such as chromothripsis and genetic subclones, and their spatial patterns within the tumor. The authors also explored the transcriptomic and chromatin landscapes, uncovering neurodevelopmental programs that reflect GBM origins and contribute to heterogeneity. The findings highlight potential therapeutic targets and provide insights into GBM evolution and heterogeneity, offering a 3D whole-tumor perspective that can inform clinical care and outcomes. The 3D spatially resolved maps of GBM tumors are publicly available for further research.