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Diffuse intrinsic brain stem gliomas (DIPGs) with characteristic K27M mutation of H3.3 are lethal and poorly understood childhood cancers. In this issue of Cancer Cell, Larson et al. exploit a unique murine DIPG model with inducible, endogenous K27M expression to reveal insights into mechanisms of K27M-mediated transformation in DIPG. Diffuse intrinsic brain stem gliomas (DIPGs) with characteristic K27M mutation of H3.3 are lethal and poorly understood childhood cancers. In this issue of Cancer Cell, Larson et al. exploit a unique murine DIPG model with inducible, endogenous K27M expression to reveal insights into mechanisms of K27M-mediated transformation in DIPG. Diffuse Intrinsic Pontine Gliomas (DIPGs) are highly fatal neoplasm of the brain stem that characteristically arise in children 5–7 years of age. Landmark studies have shown that 70%–80% of DIPGs have recurrent somatic mutations in H3F3A, resulting in the replacement of lysine 27 (H3K27) in the H3.3 histone variant by methionine (Schwartzentruber et al., 2012Schwartzentruber J. Korshunov A. Liu X.Y. Jones D.T. Pfaff E. Jacob K. Sturm D. Fontebasso A.M. Quang D.A. Tönjes M. et al.Driver mutations in histone H3.3 and chromatin remodelling genes in paediatric glioblastoma.Nature. 2012; 482: 226-231Crossref PubMed Scopus (1721) Google Scholar, Wu et al., 2012Wu G. Broniscer A. McEachron T.A. Lu C. Paugh B.S. Becksfort J. Qu C. Ding L. Huether R. Parker M. et al.St. Jude Children’s Research Hospital–Washington University Pediatric Cancer Genome ProjectSomatic histone H3 alterations in pediatric diffuse intrinsic pontine gliomas and non-brainstem glioblastomas.Nat. Genet. 2012; 44: 251-253Crossref PubMed Scopus (1106) Google Scholar). These heterozygous H3.3K27M mutations, hereon referred to as K27M, are predicted to have dominant-negative effects and interfere with post-translational modification of the lysine tails by the Polycomb repressor 2 complex (PRC2) that have critical functions in normal development. Various studies have now shown K27M is insufficient for inducing tumor growth but does enhance tumorigenesis with mutant p53 and/or activated PDGFRα (Funato et al., 2014Funato K. Major T. Lewis P.W. Allis C.D. Tabar V. Use of human embryonic stem cells to model pediatric gliomas with H3.3K27M histone mutation.Science. 2014; 346: 1529-1533Crossref PubMed Scopus (249) Google Scholar, Misuraca et al., 2016Misuraca K.L. Hu G. Barton K.L. Chung A. Becher O.J. A novel mouse model of diffuse intrinsic pontine glioma initiated in Pax3-expressing cells.Neoplasia. 2016; 18: 60-70Abstract Full Text Full Text PDF PubMed Scopus (34) Google Scholar, Pathania et al., 2017Pathania M. De Jay N. Maestro N. Harutyunyan A.S. Nitarska J. Pahlavan P. Henderson S. Mikael L.G. Richard-Londt A. Zhang Y. et al.H3.3K27M cooperates with Trp53 loss and PDGFRA gain in mouse embryonic neural progenitor cells to induce invasive high-grade gliomas.Cancer Cell. 2017; 32: 684-700.e9Abstract Full Text Full Text PDF PubMed Scopus (135) Google Scholar) that are frequently found in K27M human tumors. Based on striking loss of H3K27me3 repressive marks with exogenous K27M expression, global epigenetic reprogramming has been invoked in K27M-mediated transformation (Lewis et al., 2013Lewis P.W. Müller M.M. Koletsky M.S. Cordero F. Lin S. Banaszynski L.A. Garcia B.A. Muir T.W. Becher O.J. Allis C.D. Inhibition of PRC2 activity by a gain-of-function H3 mutation found in pediatric glioblastoma.Science. 2013; 340: 857-861Crossref PubMed Scopus (851) Google Scholar). However, to date, deeper mechanistic understanding of how K27M mediates such a process in DIPG development has been hampered by lack of models where endogenous K27M expression can be assessed in a dynamic epigenetic and developmental context that mimics the human disease. In this issue of Cancer Cell, Larson et al. developed a murine model with Nestin-Cre-mediated conditional knockin of the K27M-associated mutation to the endogenous H3f3a locus and used a corresponding isogenic control strain with knockin of wild-type H3f3a to methodically examine the specific cellular and molecular consequences of K27M expression in the developing post-natal brain (Larson et al., 2018Larson J.D. Kasper L.H. Paugh B.S. Jin H. Wu G. Kwon C.-H. Fan Y. Shaw T.I. Silveira A.B. Qu C. et al.Histone H3.3 K27M accelerates spontaneous brainstem glioma and drives restricted changes in bivalent gene expression.Cancer Cell. 2018; 35 (this issue): 140-155Abstract Full Text Full Text PDF PubMed Scopus (135) Google Scholar). Consistent with prior reports, they showed post-natal expression of K27M alone did not induce spontaneous tumors. They next examined whether K27M altered tumor growth in constitutional Trp53 knockout mice predisposed to development of medulloblastoma and malignant gliomas, with or without activated PDGFRα. Interestingly, in addition to accelerated tumorigenesis, post-natal induction of K27M in Trp53 KO mice resulted in increased frequency of medulloblastoma tumors. Furthermore, while mice with Trp53 KO together with activated PDGFRα developed cerebral and brain stem malignant glial tumors at comparable frequency, 95% of malignant gliomas in isogenic strains combined with K27M expression arose in the brain stem indicating greatest transforming impact of K27M on the developing hind brain. Larson et al. used transcriptional and epigenomic mapping to investigate the nature of genes impacted by K27M in fore and hind brain derived NSCs, which exhibited only enhanced self-renewal and proliferation without immortalization. They observed expected substantial H3K27me3 downregulation in K27M NSCs but intriguingly saw only modest and selective changes in the transcriptome, which suggested K27M has very specific cell-context-dependent transcriptional impact. In addition to cell proliferation genes seen in NSCs, analyses of tumors from triple mutant (K27M together with Trp53 KO and activated PDGFRα) and corresponding H3f3a wild-type mice revealed K27M-dependent genes were most highly enriched for PRC1/PRC2 targets with functions in neurogenesis. Most notably, they uncovered a subset PRC1/PRC2 hind brain target genes with bi-valent H3K27me3 and H3K4me1 marks that were enriched in K27 wild-type tumors, indicating K27M-associated transcriptional changes may be partly mediated by epigenetic and transcriptional release of poised loci. Interestingly, K27M-dependent PRC1/2 target genes also exhibited a spectrum of seemingly paradoxical epigenetic and transcriptional states in tumors from the triple mutant mice, including enrichment of H3K27me3 marks at promoters of repressed loci (Figure 1). These varied but highly selective K27M-associated transcriptional effects is consistent with recent reports of dynamic, dosage-sensitive K27M:PRC2 interactions in human DIPG cells (Fang et al., 2018Fang D. Gan H. Cheng L. Lee J.H. Zhou H. Sarkaria J.N. Daniels D.J. Zhang Z. H3.3K27M mutant proteins reprogram epigenome by sequestering the PRC2 complex to poised enhancers.Elife. 2018; 7Crossref Scopus (48) Google Scholar, Stafford et al., 2018Stafford J.M. Lee C.H. Voigt P. Descostes N. Saldaña-Meyer R. Yu J.R. Leroy G. Oksuz O. Chapman J.R. Suarez F. et al.Multiple modes of PRC2 inhibition elicit global chromatin alterations in H3K27M pediatric glioma.Sci. Adv. 2018; 4: u5935Crossref Scopus (82) Google Scholar) and H3K27M synergy with other PRC2 components (Justin et al., 2016Justin N. Zhang Y. Tarricone C. Martin S.R. Chen S. Underwood E. De Marco V. Haire L.F. Walker P.A. Reinberg D. et al.Structural basis of oncogenic histone H3K27M inhibition of human polycomb repressive complex 2.Nat. Commun. 2016; 7: 11316Crossref PubMed Scopus (247) Google Scholar). These observations suggest much greater fluidity to K27M-driven transcriptional programs and epigenomes and underscore the biological complexity of this developmental cancer. The discovery of recurrent K27M mutations in DIPG represented a significant advance toward biological understanding of this enigmatic and challenging disease. Through development of a unique, endogenous H3K27 oncohistone-driven model of DIPG, Larson et al. have provided exciting new insights into the ontogeny of DIPG and how K27M acts via complex, as yet uncharacterized, mechanisms to alter transcriptional and epigenetic programs in DIPG. In addition to informing development of novel DIPG models and K27M targeted therapies, this model will serve as a critical tool to study genetic and epigenetic factors that contribute to the substantial molecular and clinical heterogeneity seen in DIPG, as well as other hind brain tumors. This work was supported by the Canadian Institute of Health Research Grant no. 327030. P.S.-C. is currently an employee of Regeneron Pharmaceuticals, Inc. Histone H3.3 K27M Accelerates Spontaneous Brainstem Glioma and Drives Restricted Changes in Bivalent Gene ExpressionLarson et al.Cancer CellDecember 27, 2018In BriefLarson et al. show that H3.3 K27M cooperates with active PDGFRα mutant and loss of p53 to induce brainstem gliomas molecularly resembling human DIPG in mice. These tumors show global H3K27 modification but restricted gene expression changes, including upregulation of genes associated with neural development. Full-Text PDF Open Archive

Tails of a Super Histone