Abstract Diffuse midline gliomas (DMG) are characterized by the histone mutation H3K27M and infiltrative tumors that occasionally migrate to distant CNS regions. We established a novel two-step pooled whole-genome CRISPR-Migration screen of metastatic H3K27M-DMG (n = 3) and glioblastoma (GBM, n = 2) stem cells by using serum-free conditions and laminin-dependent cell attachment. Genes involved in focal adhesion (ITGB1, CRKL, PARVA, PTK2, FERMT2) were identified as significantly restricting migration in all H3K27M-DMG lines. From those, only knockout of ITGB1 (ITGB1-KO) fully ablated in vitro migration in all models. ITGB1-KO in orthotopic UMPED83 cells led to significant extension of survival (p = 0.0003), and an unexpected reduction in proliferation (Ki67 and p-Rb). In human tumors, expression of ITGB1 correlated with higher glioma grade and worse survival specifically in H3K27M-DMG. Spatial transcriptomic analysis of ITGB1-KO orthotopic tumors revealed loss of precursor (OPC-like) K27M cells and increase in differentiated (OC-like) cells in invading border of tumor, suggesting a potential mechanism for reduced survival. PI3K/mTOR pathway was also downregulated in ITGB1-KO in vitro and in vivo, consistent with breast cancer data demonstrating that integrins β3 and β1 cooperate with insulin receptors to modulate the effects of mTOR on tumor initiation. Indeed, ITGB1-KO led to decreased TCA cycle activity (RNA and metabolite) and mitochondrial elongation by electron microscopy. Finally, pharmacologically targeting of ITGB1 (anti-ITGB1 antibody AIIB2, CNS delivered) led to significant extension of survival in one orthotopic model (UMPED83, p = 0.01), but less impact on another model (SVZ). SVZ tumors demonstrated upregulation of other integrins (ITGB3 and ITGB5). This resulted in susceptibility to the ITGB3/5 inhibitor Cilengitide (avg. survival = 84 days, control = 50 days) and combination treatment with AIIB2 and Cilengitide leading to long-term remission in 75% mice (148 days ongoing, p = 0.0055). In summary, targeting multiple integrins offers untapped therapeutic opportunities in H3K27M-DMG by impairing H3K27M-DMG- specific metabolic and differentiation programs.
Cartaxo et al. (Fri,) studied this question.