Microglia in diffuse midline glioma contribute to extracellular matrix remodelling and cancer cell invasion

Background: Diffuse midline glioma, H3K27-altered (DMG) are aggressive and universally fatal paediatric tumours that arise in midline structures of the brain. DMG tumours are characterized by a substantial infiltration of tissue-resident microglia. The aim of this study was to determine whether in response to stimulation by DMG cells, microglia could acquire a reactive state that contributes to the progression of DMG. Methods: The microglial transcriptomic response to DMG cells was analysed by RNA-sequencing of microglia exposed to DMG, H3K27M cells and of tumour-associated myeloid cells isolated from DMG tumour biopsies. Meta-analysis of human paediatric glioma cohorts (Kids First, PNOC, and CBTTC) was used to confirm upregulation of expression for selective genes in DMG patients. Immunofluorescence analysis performed on archival DMG biobank tissues was used to confirm the findings at protein level. Invasion assay’s using a panel of DMG, H3K27M cells was used to evaluate the contribution of microglia-derived fibronectin,to promote DMG cell invasion. Results: Microglia are enriched in human DMG, H3K27M tumours. Upon stimulation by DMG, H3K27M cells, microglia contribute to the expression of extracellular matrix (ECM) components. Microglia-derived fibronectin promotes tumour invasiveness, and its inhibition reduces DMG cell migration. Across patient cohorts, DMG, H3K27M tumours exhibit elevated expression of myeloid cell-derived ECM components, with fibronectin expression showing prognostic value. Conclusions: Microglia appear to actively contribute to the diffuse invasive nature of DMG by producing ECM components, particularly fibronectin, which enhances tumour invasiveness. These findings identify microglia as potential therapeutic targets in DMG.