Abstract
Glioblastoma invasion into brain parenchyma presents significant challenges for treatment but remains poorly understood. In this study, we combine single-cell RNA sequencing, spatial transcriptomics, and multiplexed imaging of orthotopic xenograft models to investigate glioblastoma invasion. We first screen 20 patient-derived gliomasphere models for their distal (i.e., extending to the contralateral hemisphere) and local invasive potential in mice. We show that models with distal invasion potential are enriched with oligodendrocyte progenitor-like cells, while models with only local invasion potential are enriched with mesenchymal-like cells. These patterns reflect predominantly peri-axonal vs peri-vascular invasion routes, respectively. Next, we analyze the transcriptomes of invading cells within models (compared to tumor core) and identify programs associated with distal and local invasion. Thus, we decouple transcriptional features associated with invasion potential from those associated with the process of invasion. We validate our findings by spatial transcriptomics and multiplexed imaging, further describing the spatial niche of invasive cells. Taken together, our results provide a blueprint for the invasive potential of glioblastoma cell states and of the programs associated with invasion across different scales.
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Data availability
The single-cell RNA sequencing and spatial transcriptomics data generated in this study have been deposited in the Gene Expression Omnibus (GEO) database under accession code GSE281796 (https://www.ncbi.nlm.nih.gov/geo/query/acc.cgi?acc=GSE281796) The published spatial transcriptomics data that was used in this study are available in Datadryad (https://doi.org/10.5061/dryad.wpzgmsbv6) The published bulk RNA-seq data of GBM PDXs that was used in this study are available in the NCBI BioProject database under accession code: PRJNA548556 (https://www.ncbi.nlm.nih.gov/bioproject/PRJNA548556) The source data are provided in the Source Data file. Source data are provided with this paper.
Code availability
The code used to create the figures in this manuscript was uploaded to (https://github.com/rchanoch/GBM_Invasion_script).
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Acknowledgements
This work was supported by Grant-in-Aid for JSPS Fellows from the Japan Society for the Promotion of Science (to T.H.), SENSHIN Medical Research Foundation (to T.H.), Kanae Foundation (to T.H.), Brain Research Foundation (to T.H.), American Brain Tumor Association (to T.H.), Seth M Boyer Foundation (to T.H.), N.I.H. R37CA245523 (to M.L.S.), N.I.H. R01CA258763 (to M.L.S.), Mark Foundation Emerging Leader Award (to M.L.S.), Sontag Foundation Distinguished Scientist Award (to M.L.S.), MGH Research Scholars (to M.L.S.), Broad Institute-Israel Science Foundation Collaborative Project Award (to I.T. and M.L.S.), European Research Council Consolidator Grant 101044318 (to I.T.), Zuckerman STEM Leadership Program (to I.T.), Mexican Friends New Generation (to I.T.),. I.T. is the incumbent of the Dr. Celia Zwillenberg-Fridman and Dr. Lutz Zwillenberg Career Development Chair. IMMediate Advanced Clinician Scientist-Program, Department of Medicine II, Medical Center–University of Freiburg and Faculty of Medicine, University of Freiburg, funded by the Bundesministerium für Bildung und Forschung (BMBF, Federal Ministry of Education and Research) − 01EO2103 (to R.H.).
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R.C.M., T.H., A.C.G., M.L.S., and I.T. conceived the project, designed the study, interpreted results, and wrote the manuscript. R.C.M. and A.C.G. performed computational analyses. T.H. performed GBM model experiments. A.C.G. and E.C.F. performed spatial transcriptomics. R.H. performed antibody-based multiplexed imaging (CODEX) experiments. L.B. and H.R.W. helped generate plate-based scRNA-seq data. E.N.B., J.G., Z.B., A.J., J.M.H.F., W.A., and S.C.P. helped perform animal and pathological studies. C.B., J.B., R.G., and H.W. helped GBM model development.
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I.T. is an advisory board member of Immunitas Therapeutics, and a scientific co-founder, equity holder and advisory board member of Cellyrix Therapeutics. M.L.S. is an equity holder, scientific co-founder, and advisory board member of Immunitas Therapeutics. The remaining authors declare no competing interests.
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Chanoch-Myers, R., Hara, T., Greenwald, A.C. et al. A blueprint for local and distal invasion programs in glioblastoma. Nat Commun (2026). https://doi.org/10.1038/s41467-026-70470-8
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DOI: https://doi.org/10.1038/s41467-026-70470-8
