Isolated calvarial aneurysmal bone cyst in a pediatric patient: illustrative case.

BACKGROUND: Aneurysmal bone cysts (ABCs) are benign, osteolytic lesions that can occur in long bones, vertebrae, or rarely, the skull. Here the authors present the case of a 15-year-old male with a primary ABC of the left frontoparietal skull along with a review of the literature to provide insight into the nature of this rare disease. OBSERVATIONS: An otherwise healthy 15-year-old male presented with a tense, painful lesion of the left frontoparietal scalp. He could not identify any inciting trauma, but first noted the lesion less than 2 weeks prior to presentation with progressive enlargement. Cranial imaging revealed a lytic skull lesion with fluid-fluid levels suggestive of ABC. Curative therapy was provided via wide excision of the lesion and calvarial reconstruction of the resultant skull defect. This was performed without complication, and histopathological evaluation confirmed the diagnosis of primary ABC. LESSONS: ABCs of the skull are rare entities and most often arise in the skull base versus the calvaria. Typically, these lesions are associated with an underlying bone pathology (secondary ABCs) but can be rarely seen as isolated lesions (primary ABCs). Clinical management consists of excision and adjuvant therapy for underlying pathology where appropriate.

H3 K27M-altered glioma and diffuse intrinsic pontine glioma: Semi-systematic review of treatment landscape and future directions.

H3 K27M-mutant diffuse glioma is a recently identified brain tumor associated with poor prognosis. As of 2016, it is classified by the World Health Organization as a distinct form of grade IV glioma. Despite recognition as an important prognostic and diagnostic feature in diffuse glioma, radiation remains the sole standard of care and no effective systemic therapies are available for H3K27M mutant tumors. This review will detail treatment interventions applied to diffuse midline glioma and diffuse intrinsic pontine glioma (DIPG) prior to the identification of the H3 K27M mutation, the current standard-of-care for H3 K27M-mutant diffuse glioma treatment, and ongoing clinical trials listed on www.clinicaltrials.gov evaluating novel therapeutics in this population. Current clinical trials were identified using clinicaltrials.gov, and studies qualifying for this analysis were active or ongoing interventional trials that evaluated a therapy in at least 1 treatment arm or cohort comprised exclusively of patients with DIPG and H3 K27M-mutant glioma. Forty-one studies met these criteria, including trials evaluating H3 K27M vaccination, chimeric antigen receptor T-cell therapy, and small molecule inhibitors. Ongoing evaluation of novel therapeutics is necessary to identify safe and effective interventions in this underserved patient population.

Liquid Biopsy in H3K27M Diffuse Midline Glioma.

Diffuse midline glioma (DMG) with H3K27M mutation is an aggressive and difficult to treat pediatric brain tumor. Recurrent gain of function mutations in H3.3 (H3.3A) and H3.1 (H3C2) at the 27th lysine to methionine (H3K27M) are seen in over 2/3 of DMGs, and are associated with a worse prognosis. Due to the anatomical location of DMG, traditional biopsy carries risk for neurologic injury as it requires penetration of vital midline structures. Further, radiographic (MRI) monitoring of DMG often shows non-specific changes, which makes therapeutic monitoring difficult. This indicates a critical need for more minimally invasive methods, such as liquid biopsy, to understand, diagnose, and monitor H3K27M DMG. Here we review the use of all modalities to date to detect biomarkers of H3K27M in CSF, blood, and urine, and compare their effectiveness in detection, diagnosis, and monitoring treatment response. We provide specific detail of recent efforts to monitor CSF and plasma H3K27M cell-free DNA in patients undergoing therapy with the imipridone ONC201. Lastly, we discuss the future of therapeutic monitoring of H3K27M-DMG, including biomarkers such as mitochondrial DNA, mutant and modified histones, and novel sequencing-based approaches for improved detection methods.

H3K27M Mutant Glioma: Disease Definition and Biological Underpinnings.

High-grade glioma (HGG) is the most common cause of cancer death in children, and the most common primary central nervous system (CNS) tumor in adults. While pediatric HGG was once thought to be biologically similar to the adult form of disease, research has shown these malignancies to be significantly molecularly distinct, necessitating distinct approaches to their clinical management. However, emerging data have shown shared molecular events in pediatric and adult HGG including the histone H3K27M mutation. This somatic missense mutation occurs in genes encoding one of two isoforms of the Histone H3 protein, H3F3A (H3.3) or HIST1H3B (H3.1), and is detected in up to 80% of pediatric diffuse midline gliomas and in up to 60% of adult diffuse gliomas. Importantly, the H3K27M mutation is associated with poorer overall survival and response to therapy compared to patients with H3 wild-type tumors. Here, we review the clinical features and biological underpinnings of pediatric and adult H3K27M mutant glioma, offering a groundwork for understanding current research and clinical approaches for the care of patients suffering with this challenging disease.

LIN28B and Let-7 in Diffuse Midline Glioma: A Review.

Diffuse midline glioma (DMG) is the most lethal of all childhood cancers. DMGs are driven by histone-tail-mutation-mediated epigenetic dysregulation and partner mutations in genes controlling proliferation and migration. One result of this epigenetic and genetic landscape is the overexpression of LIN28B RNA binding protein. In other systems, LIN28B has been shown to prevent let-7 microRNA biogenesis; however, let-7, when available, faithfully suppresses tumorigenic pathways and induces cellular maturation by preventing the translation of numerous oncogenes. Here, we review the current literature on LIN28A/B and the let-7 family and describe their role in gliomagenesis. Future research is then recommended, with a focus on the mechanisms of LIN28B overexpression and localization in DMG.

Patient and operative factors associated with unanticipated intensive care admission and outcomes following posterior fossa decompressions in children: A retrospective study.

INTRODUCTION: Posterior fossa decompression for Chiari I Malformation is a common pediatric neurosurgical procedure. We sought to identify the impact of anesthesia-related intraoperative complications on unanticipated admission to the intensive care unit and outcomes following posterior fossa decompression. METHODS: Medical records of all patients <18 years who underwent surgery for Chiari I malformation between 1/1/09 and 1/31/21 at the Ann & Robert H. Lurie Children's Hospital of Chicago were included. Records were reviewed for patient characteristics, anesthesia-related intraoperative complications, postoperative complications, and surgical outcomes. The primary outcome was the incidence of unanticipated admission to the intensive care unit, and the primary variable of interest was an anesthesia-related intraoperative complication. Patient, surgical characteristics, and year of surgery were also compared between patients with and without an unanticipated admission to the intensive care unit, and a multi-variable adjusted estimate of odds of unanticipated admission to the intensive care unit admission following an anesthesia-related intraoperative complication was performed. Secondary outcomes included anesthesia factors associated with an anesthesia-related intraoperative event, and postoperative complications and surgical outcomes between patients admitted to the intensive care unit and those who were not. RESULTS: Two hundred ninety-six patients with Chiari I Malformation were identified. Clinical characteristics associated with an unanticipated admission to the intensive care unit were younger age, American Society of Anesthesiologist (ASA) physical status >2 and an anesthesia-related intraoperative complication. 29 anesthesia-related intraoperative complications were observed in 25 patients (8.4%). Two of 25 patients (8%) with an anesthesia-related intraoperative complication compared with 3 of 271 (1%) patients without anesthesia-related intraoperative complication had an unanticipated admission to the intensive care unit, odds ratio 7.8 (95% CI 1.2-48.8, p = .010). When adjusted for age, sex, ASA physical status, presenting symptoms, concomitant syringomyelia, previous decompression surgery and year of surgery, the odds ratio for an unanticipated admission to the intensive care unit following an anesthesia-related intraoperative complication was 5.9 (95% CI 0.51-59.6, p = .149). There were no differences in surgical outcomes between patients with or without an unanticipated admission to the intensive care unit. CONCLUSION: Our study demonstrates that although anesthesia-related intraoperative complications during posterior fossa decompression are infrequent, they are associated with an increased risk of an unanticipated admission to the intensive care unit.

Gorham Stout disease of the temporal bone with cerebrospinal fluid leak.

Gorham Stout disease (GSD) is a rare disease characterized by the proliferation of endothelial lined vessels and replacement of bone by fibrous tissue. The main imaging features are progressive osteolysis and cortical resorption. Temporal bone involvement is rare but presents as a destructive bone lesion that may be misinterpreted as more common lytic processes in the pediatric population, such as infection or Langerhans cell histiocytosis. GSD of the temporal bone is associated with cerebrospinal fluid (CSF) leaks, may present with otorrhea, and can mimic other causes of ear drainage. Here, we report the clinical course, imaging features, and outcomes of a 3-year-old girl with GSD of the temporal bone presenting with CSF leak initially attributed to infection.

Epigenomic landscape and 3D genome structure in pediatric high-grade glioma.

Pediatric high-grade gliomas (pHGGs), including glioblastoma multiforme (GBM) and diffuse intrinsic pontine glioma (DIPG), are morbid brain tumors. Even with treatment survival is poor, making pHGG the number one cause of cancer death in children. Up to 80% of DIPGs harbor a somatic missense mutation in genes encoding histone H3. To investigate whether H3K27M is associated with distinct chromatin structure that alters transcription regulation, we generated the first high-resolution Hi-C maps of pHGG cell lines and tumor tissue. By integrating transcriptome (RNA-seq), enhancer landscape (ChIP-seq), genome structure (Hi-C), and chromatin accessibility (ATAC-seq) datasets from H3K27M and wild-type specimens, we identified tumor-specific enhancers and regulatory networks for known oncogenes. We identified genomic structural variations that lead to potential enhancer hijacking and gene coamplification, including A2M, JAG2, and FLRT1 Together, our results imply three-dimensional genome alterations may play a critical role in the pHGG epigenetic landscape and contribute to tumorigenesis.

Standardization of the liquid biopsy for pediatric diffuse midline glioma using ddPCR.

Diffuse midline glioma (DMG) is a highly morbid pediatric brain tumor. Up to 80% of DMGs harbor mutations in histone H3-encoding genes, associated with poor prognosis. We previously showed the feasibility of detecting H3 mutations in circulating tumor DNA (ctDNA) in the liquid biome of children diagnosed with DMG. However, detection of low levels of ctDNA is highly dependent on platform sensitivity and sample type. To address this, we optimized ctDNA detection sensitivity and specificity across two commonly used digital droplet PCR (ddPCR) platforms (RainDance and BioRad), and validated methods for detecting H3F3A c.83A > T (H3.3K27M) mutations in DMG CSF, plasma, and primary tumor specimens across three different institutions. DNA was extracted from H3.3K27M mutant and H3 wildtype (H3WT) specimens, including H3.3K27M tumor tissue (n = 4), CSF (n = 6), plasma (n = 4), and human primary pediatric glioma cells (H3.3K27M, n = 2; H3WT, n = 1). ctDNA detection was enhanced via PCR pre-amplification and use of distinct custom primers and fluorescent LNA probes for c.83 A > T H3F3A mutation detection. Mutation allelic frequency (MAF) was determined and validated through parallel analysis of matched H3.3K27M tissue specimens (n = 3). We determined technical nuances between ddPCR instruments, and optimized sample preparation and sequencing protocols for H3.3K27M mutation detection and quantification. We observed 100% sensitivity and specificity for mutation detection in matched DMG tissue and CSF across assays, platforms and institutions. ctDNA is reliably and reproducibly detected in the liquid biome using ddPCR, representing a clinically feasible, reproducible, and minimally invasive approach for DMG diagnosis, molecular subtyping and therapeutic monitoring.

Therapeutic targeting of transcriptional elongation in diffuse intrinsic pontine glioma.

BACKGROUND: Diffuse intrinsic pontine glioma (DIPG) is associated with transcriptional dysregulation driven by H3K27 mutation. The super elongation complex (SEC) is required for transcriptional elongation through release of RNA polymerase II (Pol II). Inhibition of transcription elongation by SEC disruption can be an effective therapeutic strategy of H3K27M-mutant DIPG. Here, we tested the effect of pharmacological disruption of the SEC in H3K27M-mutant DIPG to advance understanding of the molecular mechanism and as a new therapeutic strategy for DIPG. METHODS: Short hairpin RNAs (shRNAs) were used to suppress the expression of AF4/FMR2 4 (AFF4), a central SEC component, in H3K27M-mutant DIPG cells. A peptidomimetic lead compound KL-1 was used to disrupt a functional component of SEC. Cell viability assay, colony formation assay, and apoptosis assay were utilized to analyze the effects of KL-1 treatment. RNA- and ChIP-sequencing were used to determine the effects of KL-1 on gene expression and chromatin occupancy. We treated mice bearing H3K27M-mutant DIPG patient-derived xenografts (PDXs) with KL-1. Intracranial tumor growth was monitored by bioluminescence image and therapeutic response was evaluated by animal survival. RESULTS: Depletion of AFF4 significantly reduced the cell growth of H3K27M-mutant DIPG. KL-1 increased genome-wide Pol II occupancy and suppressed transcription involving multiple cellular processes that promote cell proliferation and differentiation of DIPG. KL-1 treatment suppressed DIPG cell growth, increased apoptosis, and prolonged animal survival with H3K27M-mutant DIPG PDXs. CONCLUSIONS: SEC disruption by KL-1 increased therapeutic benefit in vitro and in vivo, supporting a potential therapeutic activity of KL-1 in H3K27M-mutant DIPG.

Safety of Ventricular Reservoir Sampling in Pediatric Posthemorrhagic Hydrocephalus Patients: Institutional Experience and Review of the Literature.

ABSTRACT: INTRODUCTION: Posthemorrhagic hydrocephalus (PHH) is a common disease process encountered in neonates. Management often includes cerebrospinal fluid (CSF) aspiration through ventricular access devices (VADs). However, a common concern surrounding serial access of implanted subcutaneous reservoirs includes introduction of infection. In addition, there is great variability in aseptic technique. Therefore, the authors sought to evaluate the incidence of VAD access-associated infections in the literature and compare them with the rate of infection found at our institution. We also highlight the use of a preassembled VAD access kit and standardized access protocol, as well as the role of provider education, in maintaining safety and sterility during serial VAD access. METHODS: A single-institution retrospective review was performed for PHH patients younger than 1 year old undergoing serial CSF aspirations via implanted VADs (2009-2019). Patients were excluded if they had a ventriculoperitoneal shunt placed as primary intervention. MEDLINE search for reports of serial VAD access in PHH was also performed. Reports were excluded if they did not include full-text articles in the English literature. RESULTS: At our institution, subcutaneous reservoirs were placed in 37 neonates with PHH for serial CSF aspiration. No infections occurred after a total of 630 taps (average, 17 taps per reservoir; range, 0-83) and 10 420 collective reservoir days (average, 282 per patient; range, 6-3700). Only 2 reservoirs required revision for malfunction. Serial VAD taps for PHH were described in 14 articles in the medical literature, with 7.9% (n = 47/592) of patients reported with tap-related infectious complications. CONCLUSION: A standardized VAD access kit, along with stringent adherence to access protocol, can significantly minimize risk of infection associated with serial VAD access. These principles can be generalized to percutaneous aspiration of CSF from subcutaneous reservoirs placed for other indications to promote safety and sterility of this common procedure.

Mesenchymal Stem Cells Successfully Deliver Oncolytic Virotherapy to Diffuse Intrinsic Pontine Glioma.

PURPOSE: Diffuse intrinsic pontine glioma (DIPG) is among the deadliest of pediatric brain tumors. Radiotherapy is the standard-of-care treatment for DIPG, but offers only transient relief of symptoms for patients with DIPG without providing significant survival benefit. Oncolytic virotherapy is an anticancer treatment that has been investigated for treating various types of brain tumors. EXPERIMENTAL DESIGN: Here, we have explored the use of mesenchymal stem cells (MSC) for oncolytic virus (OV) delivery and evaluated treatment efficacy using preclinical models of DIPG. The survivin promoter drives the conditional replication of OV used in our studies. The efficiency of OV entry into the cells is mediated by fiber modification with seven lysine residues (CRAd.S.pK7). Patients' samples and cell lines were analyzed for the expression of viral entry proteins and survivin. The ability of MSCs to deliver OV to DIPG was studied in the context of a low dose of irradiation. RESULTS: Our results show that DIPG cells and tumors exhibit robust expression of cell surface proteins and survivin that enable efficient OV entry and replication in DIPG cells. MSCs loaded with OV disseminate within a tumor and release OV throughout the DIPG brainstem xenografts in mice. Administration of OV-loaded MSCs with radiotherapy to mice bearing brainstem DIPG xenografts results in more prolonged survival relative to that conferred by either therapy alone (P < 0.01). CONCLUSIONS: Our study supports OV, CRAd.S.pK7, encapsulated within MSCs as a therapeutic strategy that merits further investigation and potential translation for DIPG treatment.

Effects of H3.3G34V mutation on genomic H3K36 and H3K27 methylation patterns in isogenic pediatric glioma cells.

Histone H3.3 mutation (H3F3A) occurs in 50% of cortical pediatric high-grade gliomas. This mutation replaces glycine 34 with arginine or valine (G34R/V), impairing SETD2 activity (H3K36-specific trimethyltransferase). Consequently, reduced H3K36me3 is observed on H3.3G34V nucleosomes relative to wild-type, contributing to genomic instability and driving a distinct gene expression signature associated with tumorigenesis. However, it is not known if this differential H3K36me3 enrichment is due to H3.3G34V mutant protein alone. Therefore, we set to elucidate the effect of H3.3G34V mutant protein in pediatric glioma on H3K36me3, H3K27me3 and H3.3 enrichment in vitro. We found that the doxycycline-inducible shRNA knockdown of mutant H3F3A encoding the H3.3G34V protein resulted in loss of H3.3G34V enrichment and increased H3K36me3 enrichment throughout the genome. After knockdown, H3.3G34V enrichment was preserved at loci observed to have the greatest H3.3G34V and H3K36me3 enrichment prior to knockdown. Induced expression of mutant H3.3G34V protein in vitro was insufficient to induce genomic H3K36me3 enrichment patterns observed in H3.3G34V mutant glioma cells. We also observed strong co-enrichment of H3.3G34V and wild-type H3.3 protein, as well as greater H3K27me3 enrichment, in cells expressing H3.3G34V. Taken together, our study demonstrates the effects of H3.3G34V mutant protein on genomic H3K36me3, H3K27me3 and H3.3 enrichment patterns in isogenic cell lines.

Ventricular Cerebrospinal Fluid Sampling in Pediatric Diffuse Midline Glioma Patients: Institutional Experience and Review of the Literature.

Purpose: Increasing evidence suggests that circulating biomarkers may serve diagnostic and longitudinal monitoring purposes in pediatric neuro-oncology. Mutant tumor DNA is detectable in the cerebrospinal fluid (CSF) of pediatric diffuse midline glioma (DMG) patients and quantity can reflect disease burden. CSF sampling ("liquid biopsy") via a CSF access device could therefore play a role in DMG management. Therefore, we set to evaluate the incidence of hydrocephalus (HCP) in DMG patients, and to characterize ventricular reservoir placement and access practices. Methods: A single institution retrospective review of DMG patients ≤21-years-old was performed (1984-2019). MEDLINE searches for reports of ventricular reservoir or shunt placement in DMG, and reservoir access for intraventricular chemotherapy (IVC) were performed. Results: At our institution, 62.6% of DMG patients (67/108) required intervention for HCP: 19.4% provided transient CSF access (ETV alone n = 3, EVD n = 8, unspecified n = 2), and 80.6% permanent CSF access (ETV + reservoir n = 13, shunt n = 41). Further, 22/34 patients with initially transient CSF devices required conversion to a permanent device. Five devices were revised for malfunction, one for infection. Seventeen articles cited HCP in 22 to 100% of DMG patients. IVC administration was described in 632 patients (seven articles), with 42 infectious and 63 non-infectious complications. Conclusions: Management of HCP is often necessary in children with DMG. Given the low rate of clinical risk associated with VAD placement and access, and the potential utility of longitudinal disease monitoring via CSF analysis, VAD placement could be considered in future clinical trials to guide DMG treatment.

Histone tail analysis reveals H3K36me2 and H4K16ac as epigenetic signatures of diffuse intrinsic pontine glioma.

BACKGROUND: Diffuse intrinsic pontine glioma (DIPG) is an aggressive pediatric brainstem tumor. Most DIPGs harbor a histone H3 mutation, which alters histone post-translational modification (PTM) states and transcription. Here, we employed quantitative proteomic analysis to elucidate the impact of the H3.3K27M mutation, as well as radiation and bromodomain inhibition (BRDi) with JQ1, on DIPG PTM profiles. METHODS: We performed targeted mass spectrometry on H3.3K27M mutant and wild-type tissues (n = 12) and cell lines (n = 7). RESULTS: We found 29.2 and 26.4% of total H3.3K27 peptides were H3.3K27M in mutant DIPG tumor cell lines and tissue specimens, respectively. Significant differences in modification states were observed in H3.3K27M specimens, including at H3K27, H3K36, and H4K16. In addition, H3.3K27me1 and H4K16ac were the most significantly distinct modifications in H3.3K27M mutant tumors, relative to wild-type. Further, H3.3K36me2 was the most abundant co-occurring modification on the H3.3K27M mutant peptide in DIPG tissue, while H4K16ac was the most acetylated residue. Radiation treatment caused changes in PTM abundance in vitro, including increased H3K9me3. JQ1 treatment resulted in increased mono- and di-methylation of H3.1K27, H3.3K27, H3.3K36 and H4K20 in vitro. CONCLUSION: Taken together, our findings provide insight into the effects of the H3K27M mutation on histone modification states and response to treatment, and suggest that H3K36me2 and H4K16ac may represent unique tumor epigenetic signatures for targeted DIPG therapy.

Correction to: Comparative multidimensional molecular analyses of pediatric diffuse intrinsic pontine glioma reveals distinct molecular subtypes.

The original version of this article unfortunately contained a mistake.

Correction: Detection of histone H3 K27M mutation and post-translational modifications in pediatric diffuse midline glioma via tissue immunohistochemistry informs diagnosis and clinical outcomes.

[This corrects the article DOI: 10.18632/oncotarget.26430.].

Choroidal artery embolization in the management of cerebrospinal fluid overproduction: case report and review of the literature.

Diffuse villous hyperplasia of the choroid plexus (DVHCP) is a rare cause of communicating hydrocephalus. DVHCP may be diagnosed radiographically and through histological evaluation. It may be associated with genetic abnormalities, particularly involving chromosome 9. Due to CSF overproduction, patients with DVHCP often fail management with shunting alone and may require adjuvant interventions. The authors present the case of a child with partial trisomy 9p and delayed diagnosis of hydrocephalus with radiographic evidence of DVHCP who was successfully managed with ventriculoperitoneal shunt (VPS) placement, adjuvant bilateral endoscopic choroid plexus coagulation (CPC), and the novel application of anterior choroidal artery embolization. In addition, a systematic MEDLINE search was conducted using the keywords "diffuse villous hyperplasia," "choroid plexus hypertrophy," and "idiopathic cerebrospinal fluid overproduction." Clinicopathological characteristics and outcomes of the present case were reviewed and compared to those in the literature.A 14-month-old girl with partial trisomy 9p presented with macrocephaly and radiographic evidence of communicating hydrocephalus and DVHCP. Ventriculoperitoneal shunting resulted in distal failure due to inadequate CSF absorption, and ventriculoatrial shunt (VAS) placement was not possible due to multiple cardiac anomalies. Daily CSF production was reduced via endoscopic third ventriculostomy and bilateral CPC, followed by distal choroidal artery embolization, enabling VPS re-internalization. The embolization was complicated by radiographic evidence of an iatrogenic cerebral infarct, but this was clinically occult. Thirty-two additional cases of communicating hydrocephalus due to DVHCP are reported in the literature: 27 pediatric, 3 adult, and 2 postmortem. Genetic abnormalities were noted in 14, with 7 (50%) involving chromosome 9. Twelve patients underwent plexectomy (9 bilateral, 2 unilateral, 1 partial), and 10 underwent CPC (4 bilateral, 3 unilateral, and 3 unspecified), with or without shunting. Eight patients were successfully managed with shunting alone (6 VASs, 2 VPSs), and none underwent arterial embolization.DVHCP is a rare cause of communicating hydrocephalus that may be associated with genetic abnormalities. A thorough review of the literature highlights diagnostic criteria and interventional options involved in managing this cause of CSF overproduction. The present case demonstrates that angiographic confirmation of prominent choroidal arteries may contribute to the diagnosis DVHCP. In addition, embolization of the distal choroidal arteries may be considered as a potential adjuvant treatment in patients for whom conventional treatments have failed or are not feasible.

Methylation-dependent Tissue Factor Suppression Contributes to the Reduced Malignancy of IDH1-mutant Gliomas.

PURPOSE: Gliomas with isocitrate dehydrogenase 1 mutations (IDH1mut) are less aggressive than IDH1 wild-type (IDH1wt) gliomas and have global genomic hypermethylation. Yet it is unclear how specific hypermethylation events contribute to the IDH1mut phenotype. Previously, we showed that the gene encoding the procoagulant tissue factor (TF), F3, is among the most hypermethylated and downregulated genes in IDH1mut gliomas, correlating with greatly reduced thrombosis in patients with IDH1mut glioma. Because TF also increases the aggressiveness of many cancers, the current study explored the contribution of TF suppression to the reduced malignancy of IDH1mut gliomas.Experimental Design: TF expression was manipulated in patient-derived IDH1mut and IDH1wt glioma cells, followed by evaluation of in vitro and in vivo behavior and analyses of cell signaling pathways. RESULTS: A demethylating agent, decitabine, increased F3 transcription and TF-dependent coagulative activity in IDH1mut cells, but not in IDH1wt cells. TF induction enhanced the proliferation, invasion, and colony formation of IDH1mut cells, and increased the intracranial engraftment of IDH1mut GBM164 from 0% to 100% (P = 0.0001). Conversely, TF knockdown doubled the median survival of mice engrafted with IDH1wt/EGFRvIIIamp GBM6, and caused complete regression of IDH1wt/EGFRamp GBM12 (P = 0.001). In vitro and in vivo effects were linked to activation of receptor tyrosine kinases (RTK) by TF through a Src-dependent intracellular pathway, even when extracellular RTK stimulation was blocked. TF stimulated invasion predominately through upregulation of ß-catenin. CONCLUSIONS: These data show that TF suppression is a component of IDH1mut glioma behavior, and that it may therefore be an attractive target against IDH1wt gliomas.

Detection of histone H3 K27M mutation and post-translational modifications in pediatric diffuse midline glioma via tissue immunohistochemistry informs diagnosis and clinical outcomes.

Pediatric diffuse midline glioma is a highly morbid glial neoplasm that may arise in the thalamus or brainstem (also known as diffuse intrinsic pontine glioma or DIPG). Because tumor anatomic location precludes surgical resection, diagnosis and treatment is based on MR imaging and analysis of biopsy specimens. Up to 80% of pediatric diffuse midline gliomas harbor a histone H3 mutation resulting in the replacement of lysine 27 with methionine (K27M) in genes encoding histone H3 variant H3.3 (H3F3A) or H3.1 (HIST1H3B). H3K27M mutant glioma responds more poorly to treatment and is associated with worse clinical outcome than wild-type tumors, so mutation detection is now diagnostic for a new clinical entity, diffuse midline glioma H3K27M mutant, as defined in the most recent WHO classification system. We previously reported patterns of histone H3 trimethylation (H3K27me3) and acetylation (H3K27Ac) associated with H3K27M mutation that impact transcription regulation and contribute to tumorigenesis. Given the clinical implications of the H3K27M mutation and these associated H3 post-translational modifications (PTMs), we set to determine whether they can be characterized via immunohistochemistry (IHC) in a cohort of pediatric glioma (n = 69) and normal brain tissue (n = 4) specimens. We observed 100% concordance between tissue IHC and molecular sequencing for detecting H3K27M mutation. In turn, H3K37M and H3K27me3 results, but not H3K27Ac staining patterns, were predictive of clinical outcomes. Our results demonstrate H3K27M and H3K27me3 staining of pediatric glioma tissue may be useful for diagnosis, stratification to epigenetic targeted therapies, and longitudinal monitoring of treatment response.