H3F3A mutations are seen in ∼30% of pediatric glioblastoma (GBMs) and involve either the lysine residue at position 27 (K27M) or glycine at position 34 (G34R/V). Sixteen genes encode histone H3, each ...variant differing in only a few amino acids. Therefore, how mutations in a single H3 gene contribute to carcinogenesis is unknown. H3F3A K27M mutations are predicted to alter methylation of H3K27. H3K27me3 is a repressive mark critical to stem cell maintenance and is mediated by EZH2, a member of the polycomb‐group (PcG) family. We evaluated H3K27me3 and EZH2 expression using immunohistochemistry in 76 pediatric brain tumors. H3K27me3 was lowered/absent in tumor cells but preserved in endothelial cells and infiltrating lymphocytes in six out of 20 GBMs. H3K27me3 showed strong immunoreactivity in all other tumor subtypes. Sequencing of GBMs showed H3F3A K27M mutations in all six cases with lowered/absent H3K27me3. EZH2 expression was high in GBMs, but absent/focal in other tumors. However, no significant differences in EZH2 expression were observed between H3F3A K27M mutant and wild type GBMs, suggesting that EZH2 mediated trimethylation of H3K27 is inhibited in GBM harboring K27M mutations. Our results indicate that H3F3A K27M mutant GBMs show decreased H3K27me3 that may be of both diagnostic and biological relevance.
Glucose and glutamine are the two principal nutrients that cancer cells use to proliferate and survive. Many cancers show altered glucose metabolism, which constitutes the basis for in vivo positron ...emission tomography (PET) imaging with (18)F-fluorodeoxyglucose ((18)F-FDG). However, (18)F-FDG is ineffective in evaluating gliomas because of high background uptake in the brain. Glutamine metabolism is also altered in many cancers, and we demonstrate that PET imaging in vivo with the glutamine analog 4-(18)F-(2S,4R)-fluoroglutamine ((18)F-FGln) shows high uptake in gliomas but low background brain uptake, facilitating clear tumor delineation. Chemo/radiation therapy reduced (18)F-FGln tumor avidity, corresponding with decreased tumor burden. (18)F-FGln uptake was not observed in animals with a permeable blood-brain barrier or neuroinflammation. We translated these findings to human subjects, where (18)F-FGln showed high tumor/background ratios with minimal uptake in the surrounding brain in human glioma patients with progressive disease. These data suggest that (18)F-FGln is avidly taken up by gliomas, can be used to assess metabolic nutrient uptake in gliomas in vivo, and may serve as a valuable tool in the clinical management of gliomas.
Childhood posterior fossa (PF) ependymomas cause substantial morbidity and mortality. These tumors lack recurrent genetic mutations, but a subset of these ependymomas exhibits CpG island (CpGi) ...hypermethylation PF group A (PFA), implicating epigenetic alterations in their pathogenesis. Further, histological grade does not reliably predict prognosis, highlighting the importance of developing more robust prognostic markers. We discovered global H3K27me3 reduction in a subset of these tumors (PF-ve ependymomas) analogous to H3K27M mutant gliomas. PF-ve tumors exhibited many clinical and biological similarities with PFA ependymomas. Genomic H3K27me3 distribution showed an inverse relationship with CpGi methylation, suggesting that CpGi hypermethylation drives low H3K27me3 in PF-ve ependymomas. Despite CpGi hypermethylation and global H3K27me3 reduction, these tumors showed DNA hypomethylation in the rest of the genome and exhibited increased H3K27me3 genomic enrichment at limited genomic loci similar to H3K27M mutant gliomas. Combined integrative analysis of PF-ve ependymomas with H3K27M gliomas uncovered common epigenetic deregulation of select factors that control radial glial biology, and PF radial glia in early human development exhibited reduced H3K27me3. Finally, H3K27me3 immunostaining served as a biomarker of poor prognosis and delineated radiologically invasive tumors, suggesting that reduced H3K27me3 may be a prognostic indicator in PF ependymomas.
Cancer metabolism and epigenetics are two relatively new areas of cancer research. Recent years have seen an explosion of studies implicating either altered tumor metabolism or epigenetic mechanisms ...in the pathogenesis or maintenance of brain tumors. A new paradigm is emerging in cancer biology that represents a convergence of these themes, the metabolic regulation of epigenetics. We discuss this interrelationship in the context of two metabolic enzymes that can influence the pathogenesis of gliomas by altering the epigenetic state. The first of these enzymes is isocitrate dehydrogenase 1 (IDH1), which is mutated in secondary glioblastomas and ∼70% of grade II/III astrocytomas and oligodendrogliomas. Mutant IDH1 results in the production of a metabolite 2‐hydroxyglutarate (2‐HG) that can inhibit DNA and histone demethylating enzymes resulting in the glioma‐CpG island phenotype (G‐CIMP) and increased histone methylation marks. Pyruvate kinase M2 (PKM2), an enzyme that plays a critical role in the glycolytic pathway, is a second example of a metabolic enzyme that can affect histone modifications. In epidermal growth factor receptor (EGFR)‐driven glioblastoma, PKM2 translocates to the nucleus and phosphorylates histone 3 at threonine 11 (H3‐T11). This causes dissociation of HDAC3 from the CCND1 (Cyclin D1) and c‐MYC promoters and subsequent histone acetylation, leading to transcription of Cyclin‐D1 and c‐MYC, and subsequent cell proliferation. Modification of the epigenetic state by alterations in metabolic enzymes is a novel phenomenon that contributes to the pathogenesis of gliomas and may help in the identification of new therapeutic targets.
Conventional methods for intraoperative histopathologic diagnosis are labour- and time-intensive, and may delay decision-making during brain-tumour surgery. Stimulated Raman scattering (SRS) ...microscopy, a label-free optical process, has been shown to rapidly detect brain-tumour infiltration in fresh, unprocessed human tissues. Here, we demonstrate the first application of SRS microscopy in the operating room by using a portable fibre-laser-based microscope and unprocessed specimens from 101 neurosurgical patients. We also introduce an image-processing method - stimulated Raman histology (SRH) - which leverages SRS images to create virtual haematoxylin-and-eosin-stained slides, revealing essential diagnostic features. In a simulation of intraoperative pathologic consultation in 30 patients, we found a remarkable concordance of SRH and conventional histology for predicting diagnosis (Cohen's kappa, κ > 0.89), with accuracy exceeding 92%. We also built and validated a multilayer perceptron based on quantified SRH image attributes that predicts brain-tumour subtype with 90% accuracy. Our findings provide insight into how SRH can now be used to improve the surgical care of brain tumour patients.
Pediatric glioblastomas (GBM) are highly aggressive and lethal tumors. Recent sequencing studies have shown that ~30 % of pediatric GBM and ~80 % of diffuse intrinsic pontine gliomas show K27M ...mutations in the
H3F3A
gene, a variant encoding histone H3.3.
H3F3A
K27M mutations lead to global reduction in H3K27me3. Our goal was to develop biomarkers for the histopathologic detection of these tumors. Therefore, we evaluated the utility of measuring H3K27me3 global reduction as a histopathologic and prognostic biomarker and tested an antibody directed specifically against the H3.3 K27M mutation in 290 samples. The study cohort included 203 pediatric (including 38 pediatric high-grade astrocytomas) and 38 adult brain tumors of various subtypes and grades and 49 non-neoplastic reactive brain tissues. Detection of H3.3 K27M by immunohistochemistry showed 100 % sensitivity and specificity and was superior to global reduction in H3K27me3 as a biomarker in diagnosing
H3F3A
K27M mutations. Moreover, cases that stained positive for H3.3 K27M showed a significantly poor prognosis compared to corresponding negative tumors. These results suggest that immunohistochemical detection of H3.3 K27M is a sensitive and specific surrogate for the
H3F3A
K27M mutation and defines a prognostically poor subset of pediatric GBM.
Inactivating mutations in SMARCA4 (BRG1), a key SWI/SNF chromatin remodelling gene, underlie small cell carcinoma of the ovary, hypercalcemic type (SCCOHT). To reveal its druggable vulnerabilities, ...we perform kinase-focused RNAi screens and uncover that SMARCA4-deficient SCCOHT cells are highly sensitive to the inhibition of cyclin-dependent kinase 4/6 (CDK4/6). SMARCA4 loss causes profound downregulation of cyclin D1, which limits CDK4/6 kinase activity in SCCOHT cells and leads to in vitro and in vivo susceptibility to CDK4/6 inhibitors. SCCOHT patient tumors are deficient in cyclin D1 yet retain the retinoblastoma-proficient/p16
-deficient profile associated with positive responses to CDK4/6 inhibitors. Thus, our findings indicate that CDK4/6 inhibitors, approved for a breast cancer subtype addicted to CDK4/6 activation, could be repurposed to treat SCCOHT. Moreover, our study suggests a novel paradigm whereby critically low oncogene levels, caused by loss of a driver tumor suppressor, may also be exploited therapeutically.
More than 50% of patients with chondrosarcomas exhibit gain-of-function mutations in either isocitrate dehydrogenase 1 (IDH1) or IDH2. In this study, we performed genome-wide CpG methylation ...sequencing of chondrosarcoma biopsies and found that IDH mutations were associated with DNA hypermethylation at CpG islands but not other genomic regions. Regions of CpG island hypermethylation were enriched for genes implicated in stem cell maintenance/differentiation and lineage specification. In murine 10T1/2 mesenchymal progenitor cells, expression of mutant IDH2 led to DNA hypermethylation and an impairment in differentiation that could be reversed by treatment with DNA-hypomethylating agents. Introduction of mutant IDH2 also induced loss of contact inhibition and generated undifferentiated sarcomas in vivo. The oncogenic potential of mutant IDH2 correlated with the ability to produce 2-hydroxyglutarate. Together, these data demonstrate that neomorphic IDH2 mutations can be oncogenic in mesenchymal cells.
Common pathways and mechanisms can be found in both cancers and inborn errors of metabolism. 2‐Hydroxyglutarate (2‐HG) acidurias and isocitrate dehydrogenase (IDH) 1/2 mutant tumors are examples of ...this phenomenon. 2‐HG can exist in two chiral forms, D(R)‐2‐HG and L(S)‐2‐HG, which are elevated in D‐ and L‐acidurias, respectively. D‐2‐HG was subsequently discovered to be synthesized in IDH 1/2 mutant tumors including ∼70% of intermediate‐grade gliomas and secondary glioblastomas (GBM). Recent studies have revealed that L‐2‐HG is generated in hypoxia in IDH wild‐type tumors. Both 2‐HG enantiomers have similar structures as α‐ketoglutarate (α‐KG) and can competitively inhibit α‐KG‐dependent enzymes. This inhibition modulates numerous cellular processes, including histone and DNA methylation, and can ultimately impact oncogenesis. D‐2‐HG can be detected in vivo in glioma patients and animal models using advanced imaging modalities. Finally, pharmacologic inhibitors of mutant IDH 1/2 attenuate the production of D‐2‐HG and show great promise as therapeutic agents.
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