H3.3-K27M drives neural stem cell-specific gliomagenesis in a human iPSC-derived model

•iPSCs with inducible H3.3-K27M mutation targeted to the endogenous histone locus•The induces specific effects in different stem and progenitor cell types•H3.3-K27M drives tumorigenesis from neural cells but not glial progenitors•K27M variant H3.3 particularly disrupts bivalent chromatin domains Diffuse intrinsic pontine glioma (DIPG) is an aggressive childhood tumor of brainstem currently no curative treatment available. The vast majority DIPGs carry a H3 leading lysine 27-to-methionine exchange (H3K27M). We engineered human induced pluripotent (iPSCs) allele locus studied disease-relevant types. upregulated promoter-associated developmental genes, producing diverse outcomes While being fatal for iPSCs, increased proliferation (NSCs) lesser extent oligodendrocyte (OPCs). Only NSCs gave rise tumors upon induction TP53 inactivation orthotopic xenograft model recapitulating DIPGs. 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Comparable H3.1-K27M corresponding confirmed staining flag-tagged transgenes immunoblotting (Figure S1B). Intriguingly, small molecule inhibition (EPZ005687) despite considerable reduction applied dose range, whereas strongly 1C 1D). observed increase G1 fraction out four S1C) consistent previous reports Wiese 2016Wiese Schill Pfister Hulleman Johnsen S.A. No significant cytotoxic effect inhibitor tazemetostat (EPZ-6438) wildtype 3.3.Klin Padiatr. 228: 113-117Crossref Targeted disruption CRISPR/Cas9 S1D S1E). Concordantly, non-responsive line also little overexpression, indicating capacity become independent resetting, likely through acquisition additional S1F). confirm general vulnerability altering balance pathologic landscape imply K27M-mutant secondary disease context. 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Rapid robust long-term self-renewing ability generate mature astroglia.Sci. 5: 16321Crossref Differentiated iNSCs Nestin, SOX2, PAX6, iOPCs OLIG2-positive, iAstroglia robustly GFAP S2B). arrest media switch demonstrating identity commitment S2B S2C). (K27M), infected lentivirus expressing Cre-GFP. As control, used Flp-GFP lentivirus, merely looped initial selection cassette left frame. Upon induction, mild 2C). iOPCs, led pronounced iNSCs. 2C S2D). then whether iNSC iOPC reflects advantage stages blocking self-organizing cerebral organoids derived (Lancaster Knoblich, 2014Lancaster Knoblich J.A. Generation 2329-2340Crossref (604) Induction first steps embryoid body lead difference organoid size nor outgrowth Cre-GFP after weeks differentiation. contained slightly SOX2+ Nestin+ change OLIG2+ suggesting tendency retain self-renewal 2D S2E). enriched Ki-67. Thus, confer prior NSC specification. H3K27M-dependent transformation Funato Larson Henderso