ID #1093 Patient-derived diffuse midline glioma organoids model H3K27M-driven OPC stemness and guide personalized therapy

Abstract

H3K27 altered diffuse midline gliomas (DMGs) remain therapeutically intractable due to their anatomical inaccessibility, preserved blood-brain barrier and intrinsic treatment resistance. Progress in understanding disease mechanisms and developing effective therapies has been hindered by the low efficiency and limited fidelity of existing preclinical models. Here, we established a patient-derived diffuse midline glioma organoid system with high generation efficiency (54/59, 91.5%), encompassing all midline segments and representing the largest cohort reported to date. These organoids faithfully recapitulate the histopathological architecture, molecular landscapes, and microenvironmental features of their parental tumors. Integrated multi-omics analyses and functional assays revealed that H3K27M-mutant DMGs sustain oligodendrocyte precursor cell (OPC) stemness through dysregulation of ACTIVIN signaling. To overcome drug delivery barriers and therapeutically target stemness, we performed multi-level anti-stemness drug screening and identified a triple intrathecal therapy (TIT) regimen with selective efficacy against DMGs. TIT markedly suppressed OPC stemness and synergistically inhibited tumor growth in vitro and in vivo. Moreover, organoid-based avatar drug testing across distinct genetic backgrounds revealed differential genotype-phenotype associations between therapeutic responses. At equivalent concentrations, TIT demonstrated a stronger association with H3.3K27M mutation status and showed greater sensitivity in pontine DMG organoids, independent of TP53 mutation status. In contrast, ONC-201 exhibited higher efficacy in thalamic or midbrain DMG organoids and in tumors harboring concurrent ACVR1 mutations.In patients with DMG, organoid-guided personalized TIT or ONC-201 treatment achieved sustained clinical responses. Collectively, this study established a robust organoid platform for mechanistic interrogation and personalized therapy screening, and identified TIT as a promising translational strategy for H3K27M-mutant DMG.