Abstract Background Diffuse midline glioma (DMG), including tumors originating in the brainstem known as diffuse intrinsic pontine glioma (DIPG), are childhood and adolescent brain tumors that remain uniformly fatal despite extensive genomic and epigenomic characterization. Their unique anatomy, genetic heterogeneity, cell states, immunosuppressive microenvironment, and variable drug penetration within the midline structures create a disease that is biologically complex and difficult to treat with current therapeutic strategies. Methods Spatial multi-omics technologies have emerged as powerful tools for addressing these challenges, particularly given the limited amount of tumor tissue available from biopsy. These methods require minimal material while preserving anatomical context, enabling simultaneous profiling of thousands of transcripts, proteins, and metabolites within a single tissue section. Such approaches can profile tumor cell states, immune interactions, and pharmacological gradients within their native tissue architecture. Results Building on discussions from the 2025 ChadTough Defeat DIPG Research Workshop, we outline a practical roadmap for implementing spatial transcriptomics, spatial proteomics, imaging-based mass spectrometry, and related spatial approaches across the DMG disease course, from diagnostic biopsy through autopsy. Key considerations are discussed, including tissue triage, spatial platform selection, quality control standards, analytical workflows, and data governance frameworks needed to support collaborative analysis. Conclusions Spatial multi-omics offers an opportunity to link tumor biology, microenvironmental features, and therapeutic exposure at cellular or near-cellular resolution. Establishing shared standards and coordinated international frameworks will be essential to translate these technologies from discovery research into clinically meaningful applications for patients with DMG and DIPG. Importance of the study For lethal tumors such as DMG, including DIPG, genetic heterogeneity, anatomical location, diverse malignant cell states, and a profoundly immunosuppressive tumor microenvironment means that no single therapy is likely to achieve durable survival. Understanding how these factors interact within the spatial architecture of the tumor is therefore essential for developing more effective therapeutic strategies. Spatial multi-omics technologies enable high-resolution mapping of tumor biology directly within intact tissue, allowing researchers to identify cellular states, immune interactions, and regional differences in drug exposure that are not captured by bulk molecular profiling approaches. These approaches are particularly valuable in DMG, where biopsy tissue is scarce. By outlining practical considerations and a coordinated framework for implementing spatial multi-omics throughout the DMG disease course, this review highlights how these technologies can accelerate biological discovery and support the development of rational combination therapies for patients with these devastating tumors.
Vo et al. (Fri,) studied this question.