Dannis G. van Vuurden

Functionally defined therapeutic targets in diffuse intrinsic pontine glioma

Diffuse intrinsic pontine glioma (DIPG) is a fatal childhood cancer. We performed a chemical screen in patient-derived DIPG cultures along with RNA-seq analyses and integrated computational modeling to identify potentially effective therapeutic strategies. The multi–histone deacetylase inhibitor panobinostat demonstrated therapeutic efficacy both in vitro and in DIPG orthotopic xenograft models. Combination testing of panobinostat and the histone demethylase inhibitor GSK-J4 revealed that the two had synergistic effects. Together, these data suggest a promising therapeutic strategy for DIPG.

Survival prediction model of children with diffuse intrinsic pontine glioma based on clinical and radiological criteria

BACKGROUND Although diffuse intrinsic pontine glioma (DIPG) carries the worst prognosis of all pediatric brain tumors, studies on prognostic factors in DIPG are sparse. To control for confounding variables in DIPG studies, which generally include relatively small patient numbers, a survival prediction tool is needed. METHODS A multicenter retrospective cohort study was performed in the Netherlands, the UK, and Germany with central review of clinical data and MRI scans of children with DIPG. Cox proportional hazards with backward regression was used to select prognostic variables (P < .05) to predict the accumulated 12-month risk of death. These predictors were transformed into a practical risk score. The models performance was validated by bootstrapping techniques. RESULTS A total of 316 patients were included. The median overall survival was 10 months. Multivariate Cox analysis yielded 5 prognostic variables of which the coefficients were included in the risk score. Age ≤3 years, longer symptom duration at diagnosis, and use of oral and intravenous chemotherapy were favorable predictors, while ring enhancement on MRI at diagnosis was an unfavorable predictor. With increasing risk score categories, overall survival decreased significantly. The model can distinguish between patients with very short, average, and increased overall survival (medians of 7.0, 9.7, and 13.7 mo, respectively). The area under the receiver operating characteristic curve was 0.68. CONCLUSIONS We developed a DIPG survival prediction tool that can be used to predict the outcome of patients and for stratification in trials. Validation of the model is needed in a prospective cohort.

WEE1 Kinase Inhibition Enhances the Radiation Response of Diffuse Intrinsic Pontine Gliomas

Diffuse intrinsic pontine glioma (DIPG) is a fatal pediatric disease. Thus far, no therapeutic agent has proven beneficial in the treatment of this malignancy. Therefore, conventional DNA-damaging radiotherapy remains the standard treatment, providing transient neurologic improvement without improving the probability of overall survival. During radiotherapy, WEE1 kinase controls the G2 cell-cycle checkpoint, allowing for repair of irradiation (IR)-induced DNA damage. Here, we show that WEE1 kinase is one of the highest overexpressed kinases in primary DIPG tissues compared with matching non-neoplastic brain tissues. Inhibition of WEE1 by MK-1775 treatment of DIPG cells inhibited the IR-induced WEE1-mediated phosphorylation of CDC2, resulting in reduced G2–M arrest and decreased cell viability. Finally, we show that MK-1775 enhances the radiation response of E98-Fluc-mCherry DIPG mouse xenografts. Altogether, these results show that inhibition of WEE1 kinase in conjunction with radiotherapy holds potential as a therapeutic approach for the treatment of DIPG. Mol Cancer Ther; 12(2); 141–50. ©2012 AACR.

EZH2-regulated DAB2IP is a medulloblastoma tumor suppressor and a positive marker for survival

Purpose: Medulloblastoma is the most common malignant brain tumor in children. Despite recent improvements, the molecular mechanisms driving medulloblastoma are not fully understood and further elucidation could provide cues to improve outcome prediction and therapeutic approaches. Experimental Design: Here, we conducted a meta-analysis of mouse and human medulloblastoma gene expression data sets, to identify potential medulloblastoma tumor suppressor genes. Results: We identified DAB2IP, a member of the RAS-GTPase–activating protein family (RAS GAP), and showed that DAB2IP expression is repressed in medulloblastoma by EZH2-induced trimethylation. Moreover, we observed that reduced DAB2IP expression correlates significantly with a poor overall survival of patients with medulloblastoma, independent of metastatic stage. Finally, we showed that ectopic DAB2IP expression enhances stress-induced apoptosis in medulloblastoma cells and that reduced expression of DAB2IP in medulloblastoma cells conveys resistance to irradiation-induced cell death. Conclusion: These results suggest that repression of DAB2IP may at least partly protect medulloblastoma cells from apoptotic cell death. Moreover, DAB2IP may represent a molecular marker to distinguish patients with medulloblastoma at high risk from those with a longer survival prognosis. Clin Cancer Res; 18(15); 4048–58. ©2012 AACR.

Erratum: Functionally defined therapeutic targets in diffuse intrinsic pontine glioma(Nature Medicine (2015) 21 (555-559) DOI: 10.1038/nm.3855)

Catherine S Grasso, Yujie Tang, Nathalene Truffaux, Noah E Berlow, Lining Liu, Marie-Anne Debily, Michael J Quist, Lara E Davis, Elaine C Huang, Pamelyn J Woo, Anitha Ponnuswami, Spenser Chen, Tessa B Johung, Wenchao Sun, Mari Kogiso, Yuchen Du, Lin Qi, Yulun Huang, Marianne Hütt-Cabezas, Katherine E Warren, Ludivine Le Dret, Paul S Meltzer, Hua Mao, Martha Quezado, Dannis G van Vuurden, Jinu Abraham, Maryam Fouladi, Matthew N Svalina, Nicholas Wang, Cynthia Hawkins, Javad Nazarian, Marta M Alonso, Eric H Raabe, Esther Hulleman, Paul T Spellman, Xiao-Nan Li, Charles Keller, Ranadip Pal, Jacques Grill & Michelle Monje Nat. Med. 21, 555–559 (2015); doi:10.1038/nm.3855; published online 4 May 2015; corrected after print 15 June 2015

Subventricular spread of diffuse intrinsic pontine glioma

Diffuse intrinsic pontine glioma (DIPG) is the second most common malignant pediatric brain tumor and the leading cause of brain tumor death in childhood [1]. 80 % of DIPG tumors exhibit a specific mutation (H3K27M) in the genes encoding histone 3.1 or 3.3 [2, 3]. standard therapy consisting of local radiotherapy to a dosage of 54–60 Gy extends median survival from 5 months to ∼9 months; 5-year survival remains less than 1 % [1]. The practice of focal radio-therapy to the brainstem is based in part on a 1982 autopsy study reporting DIPG to be relatively localized to the pons and adjacent structures [4]. In contrast, other neuroimaging and autopsy studies have identified widespread disease including supratentorial extension and leptomeningeal spread [5, 6]. Here, we report an autopsy series of 16 patients evaluated from 2009–2014 at stanford (n = 10) and VU (n = 6) University Medical Centers [7]. patient characteristics are listed in Table S1. Consistent with previous reports [5, 6], we found widespread dissemination of DIPG with extension to midbrain and medulla in 63 %, cerebellum in 56 %, thalamus in 56 %, frontal cortex in 25 % and supratentorial leptomeninges in 25 % (Fig. 1). The spinal cord was not consistently examined, but metastases were found in two of three cases examined; both had clinical evidence of spinal cord spread. Fig. 1 Extent of spread in DIPG. a Neuroanatomical sites and frequency of tumor invasion. Numbers indicate the percentage of cases that exhibit tumor invasion at the indicated anatomical location. The size of the circles marking each anatomical site (color key ... A previously under-recognized pattern of subventricular spread was noted in 10/16 cases, with infiltration of the subventricular zone (SVZ) and tumor nodules in the frontal horns of the lateral ventricles. In three cases lateral ventricular disease was noted on pre-mortem MRI (Fig. 2a), but subclinical tumor invasion in the SVZ of the lateral ventricles was found in six additional cases; subventricular spread was found in the third ventricle of one additional case (Fig. 2). The observed pattern of ventricular/subventricular involvement could be due to direct invasion along the SVZ corridor, intraventricular cerebrospinal fluid (CSF) seeding of the SVZ, or an as yet undescribed mechanism. The postnatal SVZ is a neural stem cell niche in the human brain [8] and DIPG cells express an immunophenotype reminiscent of neural precursor cells (Fig. S1 and [9]). Whether DIPG cells exhibit a particular tropism for this niche remains to be explored. Fig. 2 Invasion of the subventricular zone in DIPG (a). MRI images illustrating enhancing lesions (T1 post gadolinium, left image) at the frontal horns of the lateral ventricles with associated edema (FLAIR, right image) in case SU-DIPG-XIII. b H&E ( ... Following standard brainstem radiotherapy, disease progression typically occurs locally in the brainstem. However, in three of sixteen cases the subventricular frontal lobe disease contributed substantially to morbidity and mortality and preceded pontine recurrence in two cases. As therapies improve and patients survive longer in the natural history of their cancer, new patterns of regional relapse often appear (e.g. sanctuary disease in childhood leukemia). Our data show subventricular tumor spread in the majority of patients, typically later in the course of their disease. Thus as future therapies evolve to control local disease, strategies including extended or whole brain irradiation may become crucial. The patterns of widespread dissemination, including leptomeningeal, direct extension and subventricular spread, suggest that the extent of the optimal radiation field should be re-examined.

Subgroup-specific localization of human medulloblastoma based on pre-operative MRI

cochlear nucleus [2, 5], while WNT-MBs originate from dorsal brainstem precursors [1]. In order to unravel a possible relation between human MB biology and localization, and to provide novel hints regarding the cellular origin of MB, we characterized localization patterns of 71 human MBs using pre-operative MrI. Molecular MB subgrouping was established via gene expression profiling as described [4] and MrIs were analyzed blinded. Cohort details are Medulloblastoma (MB) is a group of malignant pediatric brain tumors arising in the posterior fossa. These tumors are classified into four molecular subgroups, namely WNT-, sHH-, Group 3and Group 4-MB [6]. In mice, sHH-MBs arise from granule cell precursors of the cerebellum and the

EphB2 activity plays a pivotal role in pediatric medulloblastoma cell adhesion and invasion

Eph/ephrin signaling has been implicated in various types of key cancer-enhancing processes, like migration, proliferation, and angiogenesis. In medulloblastoma, invading tumor cells characteristically lead to early recurrence and a decreased prognosis. Based on kinase-activity profiling data published recently, we hypothesized a key role for the Eph/ephrin signaling system in medulloblastoma invasion. In primary medulloblastoma samples, a significantly higher expression of EphB2 and the ligand ephrin-B1 was observed compared with normal cerebellum. Furthermore, medulloblastoma cell lines showed high expression of EphA2, EphB2, and EphB4. Stimulation of medulloblastoma cells with ephrin-B1 resulted in a marked decrease in in vitro cell adhesion and an increase in the invasion capacity of cells expressing high levels of EphB2. The cell lines that showed an ephrin-B1-induced phenotype possessed increased levels of phosphorylated EphB2 and, to a lesser extent, EphB4 after stimulation. Knockdown of EphB2 expression by short hairpin RNA completely abolished ephrin ligand-induced effects on adhesion and migration. Analysis of signal transduction identified p38, Erk, and mTOR as downstream signaling mediators potentially inducing the ephrin-B1 phenotype. In conclusion, the observed deregulation of Eph/ephrin expression in medulloblastoma enhances the invasive phenotype, suggesting a potential role in local tumor cell invasion and the formation of metastases.

Attenuated AMPA Receptor Expression Allows Glioblastoma Cell Survival in Glutamate-Rich Environment

Background Glioblastoma multiforme (GBM) cells secrete large amounts of glutamate that can trigger AMPA-type glutamate receptors (AMPARs). This commonly results in Na+ and Ca2+-permeability and thereby in excitotoxic cell death of the surrounding neurons. Here we investigated how the GBM cells themselves survive in a glutamate-rich environment. Methods and Findings In silico analysis of published reports shows down-regulation of all ionotropic glutamate receptors in GBM as compared to normal brain. In vitro, in all GBM samples tested, mRNA expression of AMPAR subunit GluR1, 2 and 4 was relatively low compared to adult and fetal total brain mRNA and adult cerebellum mRNA. These findings were in line with primary GBM samples, in which protein expression patterns were down-regulated as compared to the normal tissue. Furthermore, mislocalized expression of these receptors was found. Sequence analysis of GluR2 RNA in primary and established GBM cell lines showed that the GluR2 subunit was found to be partly unedited. Conclusions Together with the lack of functional effect of AMPAR inhibition by NBQX our results suggest that down-regulation and afunctionality of AMPARs, enable GBM cells to survive in a high glutamate environment without going into excitotoxic cell death themselves. It can be speculated that specific AMPA receptor inhibitors may protect normal neurons against the high glutamate microenvironment of GBM tumors.

A twenty-year review of diagnosing and treating children with diffuse intrinsic pontine glioma in The Netherlands

Introduction: Children with diffuse intrinsic pontine glioma (DIPG) face a dismal prognosis, with a median overall survival of 9 months. Our aims are to determine the incidence of DIPG in the Netherlands and to identify points for improvement in clinical research, a prerequisite for increasing the chance to find a cure. Methods: We performed a population-based retrospective cohort study by evaluating all children diagnosed with DIPG in the Netherlands between 1990 and 2010. Results: The incidence of DIPG in the Netherlands corresponds with international literature. Between 1990 and 2010, a large heterogeneity of treatment schedules was applied and only a minority of patients was included in clinical trials. Discussion: Given the rarity of DIPG, we emphasize the need for (inter-)national trials to facilitate the identification of potentially effective therapeutics in the future. This can be supported by the recent development of a European DIPG registry enabling international study collaborations.