Diffuse Pontine Glioma Research Articles

INNV-34. APPLICATION OF 200 KHZ TUMOR TREATING FIELDS (TTFIELDS) IN PATIENTS WITH INFRATENTORIAL HIGH-GRADE GLIOMA: CASE SERIES

Abstract BACKGROUND 200 kHz Tumor Treating Fields (TTFields) therapy is FDA-approved for supratentorial newly-diagnosed and recurrent glioblastoma. However, reports of its application in patients with infratentorial high-grade glioma (HGG) are rare. Based on computational modeling, the therapeutic field strength (~1 V/cm peak-to-peak) is thought to be achievable for infratentorial HGG. METHODS Two patients with infratentorial HGG received off-label 200 kHz TTFields therapy with the Optune® device (Novocure, Ltd.). To reduce impaired range of motion of the neck, the electrode transducer arrays were “skeletonized” by cutting a portion of the adhesive material surrounding them. Application of the electrode arrays was performed with the neck in flexion, to further promote neck range of motion. Cloth tape was used for reinforcement as necessary. Electrode arrays were changed every 2-3 days. We report the results of patient survival and adverse events. RESULTS Patient 1: 22-year-old man with high-grade diffuse pontine glioma with recurrences and prior treatments (including radiation, temozolomide, lomustine, and dopamine receptor D2 [DRD2] antagonist), tolerated TTFields combined with systemic treatment (initially with a bi-functional DNA alkylating agent; subsequently with temozolomide and bevacizumab) for 5 months with stable disease for 4 months. He experienced grade 1 rash of the neck. Patient 2: 58-year-old woman with diffuse glioma of the brainstem (H3K27M-mutated) with multiple recurrences and prior treatments (including concurrent radiation and temozolomide, and DRD2 antagonist), tolerated TTFields combined with systemic treatment (lomustine and bevacizumab). The TTFields infratentorial layout was iteratively modified with respect to placement of the electrode transducer arrays, for improved tolerability. With the TTFields+systemic combination therapy, the patient was stable for 8 months with no local TTFields-related side effects. CONCLUSIONS We provide initial post-marketing evidence that off-label use of 200 kHz TTFields in infratentorial HGG is safe and feasible. Prospective studies are necessary to validate the efficacy of this approach.

A review of sonodynamic therapy for brain tumors.

Sonodynamic therapy (SDT) is gaining attention as a promising new noninvasive brain tumor treatment that targets and selectively kills tumor cells, with limited side effects. This review examines the mechanisms of SDT and ongoing clinical trials looking at optimization of sonication parameters for potential treatment of glioblastoma (GBM) and diffuse intrinsic pontine glioma (DIPG). The results in the first patient with recurrent GBM treated at the Mayo Clinic are briefly discussed. The authors of this literature review used electronic databases including PubMed, EMBASE, and OVID. Articles reporting relevant preclinical and clinical trials were identified by searching for text words/phrases and MeSH terms, including the following: "sonodynamic therapy," "SDT," "focused ultrasound," "5-ALA," "ALA," "brain tumors," "diffuse pontine glioma," "glioblastoma," and "high grade glioma." Preclinical and clinical trials investigating the specific use of SDT in brain tumors were reviewed. In preclinical models of high-grade glioma and GBM, SDT has shown evidence of targeted tumor cell death via the production of reactive oxygen species. Emerging clinical trial results within recurrent GBM and DIPG show evidence of successful treatment response, with minimal side effects experienced by recruited patients. So far, SDT has been shown to be a promising noninvasive cancer treatment that is well tolerated by patients. The authors present pilot data suggesting good radiological response of GBM to a single SDT treatment, with unpublished observation of a lack of off-target effects even after multiple (monthly) sonication outpatient treatments. The scope of the clinical trials of SDT is to investigate whether it can be the means by which the fatal diagnosis of GBM or DIPG is converted into that of a chronic, treatable disease. SDT is safe, repeatable, and better tolerated than both chemotherapy and radiotherapy. It has been shown to have an effect in human cancer therapy, but more clinical trials are needed to establish standardized protocols for sonosensitizer delivery, treatment parameters, and combination therapies. The most appropriate timing of treatment also remains to be determined-whether to prevent recurrence in the postoperative period, or as a salvage option in patients with recurrent GBM for which redo surgery is inappropriate. It is hoped that SDT will also be developed for a wider spectrum of clinical indications, such as metastases, meningioma, and low-grade glioma. Further clinical trials are in preparation.

Brainstem tumors in children: a monocentric series in the light of genetic and bio-molecular progress in pediatric neuro-oncology.

Brainstem tumors represent a challenge. Their management and prognosis vary according to anatomopathological findings and genetic and bio-molecular fingerprints. We present our experience with pediatric brainstem tumors. All patients admitted for a brainstem tumor at the Pediatric Neurosurgical Unit at Hôpital Femme Mère Enfant hospital between January 1997 and December 2019 were considered. Patients data were obtained through a retrospective review of the medical records; follow-up was from the last outpatient consultation. One hundred and twelve patients were included. Eighty-five patients (75.9%) had open surgery or stereotactic biopsy. Thirty-five patients were treated for hydrocephalus. Sixty-six received an adjuvant treatment. Several protocols were adopted according to the SFOP and SIOP during this time period. The overall survival rate was 45% with a median follow-up of five years (range 1-18 year). However, the survival rate was very different between the diffuse intrinsic pontine gliomas (DIPG) and the others tumor types. If we exclude the DIPG (59 patients), of which only 1 was alive at 3 years, the survival rate was 90.6% (only 5 deaths over 53 patients) with a median follow up of 5 years. Our series confirms that benign tumors of the brainstem have a good survival when treated with surgical removal ± adjuvant therapy. Diffuse pontine gliomas continue to have a dismal prognosis. Individualized treatment based on molecular fingerprints may help to select the best adjuvant therapy and hence potentially improve survival.

P20.15.A DESCRIPTIVE ANALYSIS OF A SERIES OF H3 K27-ALTERED DIFFUSE MIDLINE GLIOMAS IN CHILDREN AND ADULT POPULATION

Abstract BACKGROUND Molecular biomarkers have gained importance in providing diagnostic information for brain and spinal cord tumors. Diffuse midline gliomas (DMG) are now defined as H3 K27-altered. H3 K27M mutations were first described in pediatric diffuse pontine gliomas but can also be encountered in adult gliomas. The H3 K27M mutation often co-occurs with mutations in TP53, PDGFR, or ACVR1 and is mutually exclusive to IDH1/2 mutations and 1p/19q co-deletion. Surgical resection is often not feasible for its localization and the benefit of temozolomide (TMZ) is not clear as they are MGMT unmethylated; all of it conferring a poor prognosis. It has been suggested that pediatric patients (pts) have worse outcomes. Our aim was to describe pts with H3 K27-altered gliomas to better understand the characteristics of this population. MATERIAL AND METHODS We analyzed clinical, histological and molecular features of 6 adults and 10 pediatric pts diagnosed with H3 K27M-mutant gliomas between May 2016 and January 2021 in a tertiary hospital in Barcelona. RESULTS For adults, mean age at diagnosis was 34.6 years (range = 20 - 55 years). For children, mean age was 11.2 years (range = 4 - 17 years). Male-to-female ratio was 1:1. Tumors involved thalamus (adult = 3; pediatric = 3), pons (adult = 1; pediatric = 6), and pineal region (n = 1). Ki67 range was 2-60% (pediatric mean 32%, adult mean 20%). A molecular panel was analyzed in 8 pediatric pts and 1 adult. No EGFR amplifications were detected.Three of them had TP53 mutations and 2 had PDGFRA amplification. Only 2 pediatric pts had MGMT methylation. Regarding treatment, 9 pediatric and 4 adult pts underwent radiotherapy and TMZ as first-line, with a mean time to intervention of 5.6 months. One child went through palliative care. As a second-line treatment, re-irradiation was used in 3 pediatric pts and irinotecan-bevacizumab was used in 2 adults. One adult received up to 4 different treatments (TMZ chemoradiotherapy followed by adjuvant TMZ, irinotecan-bevacizumab, lomustine and metronomic TMZ).All 9 pediatric pts died, with a mean OS of 12.0 months. Among adults, 2 pts died, 10.5 and 31.2 months after diagnosis, 1 has progressed to first-line and is receiving PCV as second-line treatment, and 1 has completed Stupp regimen with stable disease as best response. CONCLUSION Although the sample size is limited, our cohort data resembles what has been previously reported in terms of clinical and molecular characteristics: diagnosed in young adults and pediatric pts, characterized by a midline location, a low rate of MGMT promoter methylation, frequent TP53 and PDGFR alterations, and poor prognosis. There is currently no consensus on optimal treatment sequence or best treatment for these pts. Collaborative effort and participation in clinical trials are needed to keep advancing in this poor prognosis disease.

P17.03.A MAGIC-G1: A PHASE 1 DOSE-ESCALATION TRIAL OF WEEKLY INTRATUMORAL ADMINISTRATION OF THE SOLUBILISED FORMULATION OF PANOBINOSTAT (MTX110) IN PATIENTS WITH RECURRENT GLIOBLASTOMA (RGBM)

Abstract BACKGROUND One of the biggest challenges in treating rGBM is achieving high local drug concentration. Convection enhanced delivery (CED) is a ‘direct-to-brain’ drug delivery method that previously demonstrated its usefulness in overcoming the blood-brain barrier, while also limiting systemic toxicity. MTX110 (Midatech Ltd), a water-soluble formulation of panobinostat, has demonstrated cytotoxic activity in GBM cell lines, as well as safety and efficacy in two pediatric diffuse pontine glioma trials. Delivery of MTX110 via an implanted pump and CED catheter system in patients with GBM appears to be an attractive treatment option, that allows for therapeutic doses of the drug to be delivered weekly directly to the tumor, thus avoiding systemic administration and therefore toxicity. MATERIAL AND METHODS Eligibility includes adult rGBM patients with a single enhancing lesion ≤3cm in maximal diameter; ≥4 weeks after chemotherapy, bevacizumab, or study drug; adequate organ function; and KPS ≥60%. MTX110 is continuously infused over 48hrs weekly at a rate of 0.5mL/h, via a CED catheter implanted into the enhancing disease recurrence connected to a fully internalized pump system. After safety of a single patient treated at 60μM has been established, 3-6 patients will be treated with MTX110 at 90μM as a single agent (Cohort 1). Patients in cohort 2 will receive MTX110 in combination with lomustine at a starting MTX110 dose of 60 μM and following the same dose-escalation approach. RESULTS As of March 30, 2023, the 1st patient was treated in monotherapy at 60μM and has received 12 weekly infusions of MTX110. No dose-limiting toxicities were observed. Observed study related adverse events (AEs) included: grade 1 cognitive disturbance, headache, and seizure; grade 2 candida wound infection, erythema of the scalp, bruising, and hemiparesis; and grade 3 abdominal seroma at pump insertion site. Enrolment is ongoing and the next patient will receive the 90 μM dose. CONCLUSION Weekly intratumoral administration of MTX110 via a fully internally implanted CED pump system is feasible. Updated results will be presented.

Systemic cellular viroimmunotherapy for canine high-grade gliomas

BackgroundOncolytic viruses constitute a growing field of interest, both in human and veterinary oncology, given that they are particularly helpful for treating non-surgical tumors and disseminated cancer, such as high-grade...

Conventional and Cross-Channel MR Radiomic Features do Not Predict Histone H3 Status in DIPG: Genomic and Clinical Evaluation of a Multi-Institutional Cohort

<h3>Purpose/Objective(s)</h3> Genomic profiling of diffuse pontine glioma (DIPG) has suggested distinct and clinically relevant molecular subgroups based on the presence and type of histone H3K27M mutation. Recent studies have demonstrated improved safety for biopsy, but sampling carries risks. We evaluated the impact of radiomic features on the molecular classification of 154 histologically confirmed and centrally reviewed DIPG. <h3>Materials/Methods</h3> Automatically segmented and manually reviewed tumor volumes were used to compute single & cross channel radiomic features from T1, T2, FLAIR, and T1+gadolinium MR sequences obtained from histone H3 WT (N=30) and H3K27M-mutant (N=124) DIPG. Radiomic feature stability was assessed when multiple pretreatment MRs were available using the concordance correlation coefficient. Random Forest, Logistic Regression, XGBoost, and Deep Learning classifiers were used to build predictive models of H3K27M status in a training dataset, after exclusion of correlated features. Model accuracy was assessed in the remaining validation dataset. <h3>Results</h3> Two hundred and twelve of 456 imaging features (46.4%) demonstrated stability (CCC, p<0.05) between pretreatment scans. Accuracy varied by method, but none surpassed the No Information Rate (82%). Model accuracy was not improved by efforts to remedy class imbalance (resampling and class weight adjustment). Several cross channel radiomic features were consistently selected by the models, indicating some predictive utility, although cross institutional scanning techniques were confounding. <h3>Conclusion</h3> While early efforts with single institutional data were encouraging, differences in MR technique and class imbalance seemed to limit the identification of stable radiomic features predictive of H3 status in this multi-institutional cohort. Biopsy remains key for determining trial eligibility and directing precision medicine.

Role of Dynamic Parameters of 18F-DOPA PET/CT in Pediatric Gliomas.

PET with 18F-DOPA can be used to evaluate grading and aggressiveness of pediatric cerebral gliomas. However, standard uptake parameters may underperform in circumscribed lesions and in diffuse pontine gliomas. In this study, we tested whether dynamic 18F-DOPA PET could overcome these limitations. Patients with available dynamic 18F-DOPA PET were included retrospectively. Static parameters (tumor/striatum ratio [T/S] and tumor/cortex ratio [T/N]) and dynamic ones, calculated on the tumor time activity curve (TAC), including time-to-peak (TTP), slope steepness, the ratio between tumor and striatum TAC steepness (dynamic slope ratio [DSR]), and TAC shape (accumulation vs plateau), were evaluated as predictors of high/low grading (HG and LG) and of progression-free survival and overall survival. Fifteen patients were included; T/S, T/N, TTP, TAC slope steepness, and DSR were not significantly different between HG and LG. The accumulation TAC shape was more prevalent in the LG than in the HG group (75% vs 27%). On progression-free survival univariate analysis, TAC accumulation shape predicted longer survival (P < 0.001), whereas T/N and DSR showed borderline significance; on multivariate analyses, only TAC shape was retained (P < 0.01, Harrell C index, 0.93-0.95). On overall survival univariate analysis, T/N (P < 0.05), DSR (P < 0.05), and TAC "accumulating" shape predicted survival (P < 0.001); once more, only this last parameter was retained in the multivariate models (P < 0.05, Harrell C index, 0.86-0.89). Dynamic 18F-DOPA PET analysis outperforms the static parameter evaluation in grading assessment and survival prediction. Evaluation of the curve shape is a simple-to-use parameter with strong predictive power.

Clinical profile and management of sixth nerve palsy in pediatric patients (0-15 years) in Southern India - A hospital-based study.

Purpose:This study was done to evaluate the clinical profile in pediatric patients (0–16 years) presenting with acute onset esotropia due to sixth nerve palsy and its management options in a tertiary care set up of Southern India.Methods:A total of 12 patients presenting to our OPD with acute onset esotropia due to sixth nerve palsies were included in this retrospective study. All patients were observed for 6 months and managed with prism and/or patching while waiting for spontaneous resolution and later managed surgically. Neuroimaging was done in all cases.Results:The mean deviation of esotropia at presentation was 30.17 ± 5.7 Prism Diopter (range 12–50 Prism Diopter 95% CI, SD 10.11). Mean age of the patients during presentation was 8.6 ± 2.4 years (range: 1–15 years, SD 4.27). Among the common causes of sixth nerve palsy in our study population were trauma and idiopathic intracranial hypertension followed by tumor and miscellaneous causes. Only three patients underwent surgical correction of residual deviation after a waiting period of 6 months for self-resolution. Spontaneous resolution was observed in 41.6% patients, and surgical correction (unilateral resection–recession) was done in 25% of the patients with good surgical outcome.Conclusion:At 1-year follow up, the motor outcome was satisfactory except for one patient who had diffuse pontine glioma and had worsening neurological symptoms on follow-up.

Endoscopic third ventriculostomy for the management of hydrocephalus secondary to posterior fossa tumors: A retrospective study.

Background: Endoscopic third ventriculostomy (ETV) is an effective alternative to ventriculoperitoneal shunting as well as external ventricular drainage for the urgent management of acute hydrocephalus. We performed this study to investigate the efficacy and safety of ETV before tumor resection in managing hydrocephalus in patients with posterior fossa brain tumors (PFBT) in our neurosurgery department.Methods: We conducted this retrospective observational study between February 2018 and February 2020 on all cases diagnosed with PFBT associated with triventricular obstructive hydrocephalus. We retrospectively reviewed the demographic characteristics, operative procedures, and radiological investigations of all cases. During the follow-up period, clinical, as well as radiological success were evaluated.Results: Twenty-two ETV procedures were performed in 22 cases of PFBT (mean age = 22.1 years, SD = 11.4). Of the 22 cases, 8 cases (36.4%) had ependymoma, 7 cases (31.8%) had cerebellar astrocytoma, and 5 cases (22.7 %) had medulloblastoma, while 2 cases (9.1%) had diffuse pontine gliomas. The median follow-up duration was 9 months (range 3–13 months). The most commonly reported clinical presentation was the significant intracranial pressure increase. All operations were performed successfully in all cases. Only two ETV post-tumor resection failures were documented during the follow-up period.Conclusion: Preoperative ETV has shown to be an effective long-term cerebrospinal fluid diversion procedure to manage PFBT-associated hydrocephalus, with a relatively low rate of complications. Further prospective studies are required to assess the regular use of ETV before complete tumor resection.

Pediatric brain tumor cell lines exhibit miRNA-depleted, Y RNA-enriched extracellular vesicles.

Medulloblastoma (MB) and diffuse infiltrative pontine glioma (DIPG) are malignant pediatric tumors. Extracellular vesicles (EVs) and their bioactive cargoes have been implicated in tumorigenesis. Most studies have focused on adult tumors, therefore the role of EVs and the noncoding RNA (ncRNA) landscape in pediatric brain tumors is not fully characterized. The overall aim of this pilot study was to isolate EVs from MB and DIPG patient-derived cell lines and to explore the small ncRNA transcriptome. EVs from 3 DIPG and 4MB patient-derived cell lines were analyzed. High-throughput next generation sequencing interrogated the short non-coding RNA (ncRNA) transcriptome. Known and novel miRNAs were quantified. Differential expression analysis, in silico target prediction, and functional gene enrichment were performed. EV secretomes from MB and DIPG patient-derived cell lines demonstrated discrete ncRNA biotypes. Notably, miRNAs were depleted and Y RNAs were enriched in EV samples. Hierarchical cluster analysis revealed high discrimination in miRNA expression between DIPG and MB cell lines and RNA-Seq identified novel miRNAs not previously implicated in MB or DIPG pathogenesis. Known and putative target genes of dysregulated miRNAs were identified. Functional annotation analysis of the target genes for differentially expressed EV-and parental-derived miRNAs revealed significant cancer-related pathway involvement. This hypothesis-generating study demonstrated that pediatric brain tumor-derived cell lines secrete EVs comprised of various ncRNA cargoes. Validation of these findings in patient samples may provide new insights into the pediatric brain tumor microenvironment and identification of novel therapeutic candidates.

EPCT-19. A PHASE I STUDY OF RIBOCICLIB AND EVEROLIMUS FOLLOWING RADIATION THERAPY IN CHILDREN WITH NEWLY DIAGNOSED NON-BIOPSIED DIFFUSE PONTINE GLIOMAS (DIPG) AND RB+ BIOPSIED DIPG AND HIGH GRADE GLIOMAS (HGG)

Genomic aberrations in the cell cycle and mTOR pathways have been reported in diffuse pontine gliomas (DIPG) and high-grade gliomas (HGG). Dual inhibition of CDK4/6 (ribociclib) and mTOR (everolimus) has strong biologic rationale, non-overlapping single-agent toxicities, and adult clinical experience. The maximum tolerated dose (MTD) and/or recommended phase two dose (RP2D) of ribociclib and everolimus administered during maintenance therapy following radiotherapy was determined in the phase I study as a rolling 6 design. Ribociclib and everolimus were administered once daily for 21 days and 28 days, respectively starting two-four weeks post completion of radiotherapy. All HGG patients and any DIPG patient who had undergone biopsy were screened for RB protein by immunohistochemistry. Eighteen eligible patients enrolled (median age 8 years; range: 2–18). Six patients enrolled at dose levels 1,2, and 3 without dose limiting toxicities (DLT). Currently, five patients are enrolled at dose level 3 expansion cohort. The median number of cycles are 4.5 (range: 1–20+). Among the expansion cohort, one dose limiting toxicity included a grade 3 infection and one patient required a dose reduction in course 3 due to grade 3 ALT and grade 4 hypokalemia. The most common grade 3/4 adverse events included neutropenia. Preliminary pharmacokinetic studies on 12 patients suggest an impact of ribociclib on everolimus pharmacokinetics. The MTD/RP2D of ribociclib and everolimus following radiotherapy in newly diagnosed DIPG and HGG is anticipated to be 170 mg/m2/day x 21 days and 1.5 mg/ m2/day every 28 days which is equivalent to the adult RP2D.

DIPG-23. SINGLE CASE REPORT OF LOW DOSE RADIOTHERAPY AND CHEMOTHERAPY IN THE TREATMENT OF DIPG

A 3 year-old girl, was noted to have progressive gait problem since Nov. 2017 and brought to hospital for checkup. Brain MRI on Jan. 2018 showed T2 hyperintensity infiltrative pontine lesion, favoring diffuse infiltrative pontine glioma and mild obstructive hydrocephalus, received VP-shunting on Jan. 15, 2018. Due to the refusal of surgical biopsy for tissue proof, we started the radiotherapy from Jan. 24, 2018, using Rapidarc technique with 6MV photon energy to treat the brain stem lesion, ended on Feb. 24, 2018 with total dose of 25.5Gy in 17 fractions as our usual practice. Following the completion of radiotherapy, we started the adjuvant chemotherapy using 1-week on, 1-week off regimen of temozolomide using dosage of 75 mg/sq-m/day, and this patient’s general condition returned back to nearly normal. Serial follow-up images of brain MRI on 04/30/2018, 08/01/2018, 11/30/2018, 02/26/2019, 05/30/2019, 08/28/2019 showed slow progression of the pontine lesion, without the development of contrast enhanced new lesion. She maintained the functional independent until Sep. 2019, she was noted to have symptoms of ataxic gait, esotropia and choking on drinking liquid. We started the retreatment of radiotherapy from Oct.7, 2019, using same technique, ended on Nov. 5, 2019 with total dose of 30Gy in 20 fractions. The symptoms improved partially after the treatment, with residual weakness over left extremity. We are still treating the patient with adjuvant temozolomide, and she has survived most of time functionally independent in these 2 years.

Diffuse infiltrative pontine glioma biopsy in children with neuronavigation, frameless procedure: A single center experience of 10 cases

IntroductionThis study presented pediatric DIPG ‘s biopsy with frameless Neuronavigation. Patients and methodsWe report our experience about 10 patients who had Diffuse Intrinsic Pontine Glioma between 2014 and 2018. All patients were biopsied with BrainLab Varioguide Neuronavigation®. We always used fusion between specific CT Scan and MRI to selected target, made planning and biopsies. All patients were included in BIOMEDE after scientific and ethic discussions. We always selected a trans-cerebellar trajectory and made same procedure (lot of biopsies at one level). All patients have MRI at J1 to verify site of biopsy and to eliminate complication. ResultsThe average age was 8.1 years. Symptoms were common with principally headaches and nystagmus. All biopsies were contributive for histopathological diagnosis and establish molecular profile for molecular study. We have no definitive morbidity and procedure duration was 93minutes in average. All MRI didn’t showed intracranial complication after procedure and showed great precision of biopsy compared with the selected target. DiscussionWe reviewed the literature and compare our results with series of DIPG biopsies using stereotactic frame or robotic assisted frameless. It was a safe, accuracy and easiness procedure. We always have histopathological and molecular result to proceed next step of treatment. This modality is an alternative possibility to biopsy very young patients with low morbidity.

Global activation of oncogenic pathways underlies therapy resistance in diffuse midline glioma

Diffuse midline gliomas (DMGs) are aggressive pediatric brain tumors with dismal prognosis due to therapy-resistant tumor growth and invasion. We performed the first integrated histologic/genomic/proteomic analysis of 21 foci from three pontine DMG cases with supratentorial dissemination. Histone H3.3-K27M was the driver mutation, usually at high variant allele fraction due to recurrent chromosome 1q copy number gain, in combination with germline variants in ATM, FANCM and MYCN genes. Both previously reported and novel recurrent copy number variations and somatic pathogenic mutations in chromatin remodeling, DNA damage response and PI3K/MAPK growth pathways were variably detected, either in multiple or isolated foci. Proteomic analysis showed global upregulation of histone H3, lack of H3-K27 trimethylation, and further impairment of polycomb repressive complex 2 by ASXL1 downregulation. Activation of oncogenic pathways resulted from combined upregulation of N-MYC, SOX2, p65/p50 NF-κB and STAT3 transcription factors, EGFR, FGFR2, PDGFRα/β receptor tyrosine kinases, and downregulation of PHLPP1/2, PTEN and p16/INK4A tumor suppressors. Upregulation of SMAD4, PAI-1, CD44, and c-SRC in multiple foci most likely contributed to invasiveness. This integrated comprehensive analysis revealed a complex spatiotemporal evolution in diffuse intrisic pontine glioma, recommending pontine and cerebellar biopsies for accurate populational genetic characterization, and delineated common signaling pathways and potential therapeutic targets. It also revealed an unsuspected activation of a multitude of oncogenic pathways, including cancer cell reprogramming, explaining the resistance of DMG to current therapies.

Biopsy in diffuse pontine gliomas: expert neurosurgeon opinion-a survey from the SIOPE brain tumor group.

The prognosis of diffuse intrinsic pontine glioma (DIPG) is poor. The role of biopsy in DIPG remains controversial since the diagnosis may be established with imaging alone. Recent advances in understanding molecular biology and targeting of brain tumors have created a renewed interest in biopsy for DIPG. The Neurosurgery Working Group (NWG) of the SIOP-Europe Brain Tumor Group (BTG) undertook a survey among international pediatric neurosurgeons to define their current perceptions and practice regarding DIPG biopsy. The NWG developed a 20-question survey which was emailed to neurosurgeons in the International Society for Pediatric Neurosurgery (ISPN). The questionnaire included questions on diagnosis, indications, and techniques for biopsy, clinical trials, and healthcare infrastructure. The survey was sent to 202 neurosurgeons and 73 (36%) responded. Consensus of > 75% agreement was reached for 12/20 questions, which included (1) radiological diagnosis of DIPG is sufficient outside a trial, (2) clinical trial-based DIPG biopsy is justified if molecular targets are investigated and may be used for treatment, and (3) morbidity/mortality data must be collected to define the risk:benefit ratio. The remaining 8/20 questions proved controversial and failed to reach consensus. Routine DIPG biopsy continues to be debated. Most neurosurgeons agreed that DIPG biopsy within a clinical trial should be supported, with the aims of defining the procedure risks, improving understanding of tumor biology, and evaluating new treatment targets. Careful family counseling and consent remain important.

Magnetic resonance spectroscopy in pediatric brain tumors: how to make a more confident diagnosis

BackgroundNon-invasive diagnosis of pediatric brain tumors can be challenging due to diverse tumor pathologies and similar imaging appearances. Magnetic resonance spectroscopy (MRS), when combined with high spatial resolution anatomic imaging obtained with conventional magnetic resonance imaging (MRI), provides metabolic information within the lesion as well as the surrounding tissue. The differentiation of neoplastic from non-neoplastic lesions and low-grade from high-grade neoplasms is essential for determining the choice of treatment and the best treatment plan. We aimed to measure specific metabolic ratios and evaluate metabolic profiles of various lesions by MRS to assist in making a more confident diagnosis.ResultsThe choline/creatine (Cho/Cr), choline/N-acetylaspartate (Cho/NAA), and Cho/NAA+Cr ratios all had statistically significant values for the differentiation between neoplastic and non-neoplastic lesions at cutoffs 1.8, 2, and 0.8 respectively. The Cho/NAA, Cho/Cr, Cho/NAA+Cr, and myo-inositol/creatine (mI/Cr) ratios all had statistically significant values for the differentiation of high-grade from low-grade neoplasms at cutoffs 3.3, 3.5, 1.3, and 1.5 respectively. The presence of a lipid lactate peak was only significant for differentiating high-grade from low-grade neoplasms. Medulloblastomas, diffuse pontine gliomas, and choroid plexus carcinoma all showed characteristic metabolic profiles on MRS. Metastasis showed lower Cho/NAA and Cho/Cr ratios outside the tumor margin than high-grade neoplasms.ConclusionThe use of certain metabolite ratios with high sensitivity and specificity to distinguish neoplastic from non-neoplastic lesions and low-grade from high-grade neoplasms while assessing the metabolic profile of the lesion aids in the non-invasive diagnosis of pediatric brain tumors. MRS facilitates earlier treatment planning by determining tumor spatial extent and predicting tumor behavior with potential to solve sampling problems of inaccessible and heterogenous lesions as well as unnecessary sampling of benign lesions.

Tumor Immunized Autologous Natural Killer Cell (NK) Therapy/Compassionate Use

As well known, diffuse infiltrative pontine glioma (DIPG)-despite all available treatment modalities- carries a dismal prognosis with a mean survival of around one year [1]. A four year old boy was referred to me with a radiological diagnosis of DIPG. He was irradiated and a ventriculoperitoneal shunt (VPs) was inserted in a health facility in Europe. On admission (seven months after the diagnosis), patient was tracheostomized, conscious, quariparetic and with bilateral abducens and lower cranial nerve paresis. I decided to operate on the patient in order to debulk the necrotic pontine mass and to verify the radiological diagnosis histopathologically. Additionally, I planned to co-culture the tumoral cells with the natural killer (NK) cells isolated from the peripheral venous blood to investigate the immunization and potentiation of autologous NK cells against glioma cells in vitro after the ethical committee approval was obtained from the Istanbul Aydin University. In January 2019, pontine mass was partially removed via telo-velar approach under neurophysiological monitorization with no additional post op deficits and the unilateral abducens nerve recovery. Tumor specimens were sent to the tissue culture labs and the cell culture procedures were carried out meticulously. Meanwhile, routine histopathological evaluation reported the pontine lesion as DIPG (Grade 4 astrocytoma) officially.

Abstract 2226: Validation of a liquid biopsy tool to identify point mutations in pediatric brain tumor patients

Abstract The use of liquid biopsies to diagnose and monitor tumors has been widely explored especially in adult cancers. In pediatric cancers, the feasibility of this method as a clinical tool is still to be established. Pediatric low grade gliomas (PLGG) are characterized by mutations in the MAPK pathway, and 20% harbour BRAFV600E mutation, which can also be found in paediatric patients with Langerhans cell histiocytoma (LCH), thyroid carcinoma and melanomas. Specific histone mutations have been described in pediatric high grade gliomas(HGG) and diffuse midline pontine gliomas(DIPG). Midline gliomas are not amenable to a gross total resection; however, a biopsy is needed for pathological and molecular diagnosis. In this scenario, liquid biopsy is of utmost importance and has the potential to spare the risk of morbidity with surgical procedures, determine diagnosis and prognosis, as well as serve as a tool to guide and monitor response to therapy with targeted agents. The objective of this study is to evaluate the use of droplet digital PCR (ddPCR) for identification of point mutations in cerebro-spinal fluid (CSF) or plasma of BRAF V600E or H3K27M positive pediatric patients. CSF was collected from a total of 51 patients, and plasma from 55 patients. ctDNA was extracted from 3 ml of plasma or 2 ml of CSF, and pre-amplified prior to ddPCR, which was conducted on the RainDance system. For BRAFV600E: Forty-five patients had CSF samples available for analysis, 8 had known positive brain tumors, 32 negative brain tumors, 5 normal controls; 5/8 positive cases had BRAFV600E mutation in ctDNA from CSF, one of the patients with negative CSF and positive tumor was on treatment with BRAF inhibitors. Plasma was analysed in 17 patients with known positive brain tumors and additionally 10 patients negative for BRAFV600E, all of the plasma samples were negative for the mutation. For H3K27M: Thirty patients had CSF samples for analysis 1 was a known positive tumor, 27 negative and 2 unknown status (radiological diagnosis of DIPG). Of the 3 DIPG samples 2 were positive, one with unknown status of the primary tumor. Plasma was also analysed for H3K27M mutation in 7 patients with known positive brain tumors, one DIPG with unknown status and 7 negative samples. All plasma samples tested negative for the mutation. Sensitivity and Specificity for this assay was respectively 62% and 97% for BRAF and 66% and 98 % for H3K27M in CSF. The sensitivity in plasma for brain tumors is poor, however seems better for non-CNS lesions. In summary, we show that liquid biopsy with analysis of CSF ctDNA is feasible with high specificity, important in the context of a diagnoses tool. Moving forward larger cohorts need to be validated towards the goal of implementing ddPCR as a clinical tool for diagnosis of specific point mutations in pediatric patients with CNS tumors. Citation Format: Liana Nobre, Michal Zapotocky, Monique Johnson, Jonathan Wasserman, Oussama Abla, Jim Whitlock, Uri Tabori, Cynthia Hawkins. Validation of a liquid biopsy tool to identify point mutations in pediatric brain tumor patients [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2019; 2019 Mar 29-Apr 3; Atlanta, GA. Philadelphia (PA): AACR; Cancer Res 2019;79(13 Suppl):Abstract nr 2226.

Abstract PR08: Overexpression and mutations of CXorf67 in “infant-type” posterior fossa type-A ependymomas

Abstract Our study aims to establish whether CXorf67 has a role in the pathogenesis of posterior fossa (PF) ependymoma. Among molecular groups of ependymoma defined by DNA methylation profiling, PF type-A (PFA) is the commonest. PFA ependymomas present mainly in infants and are difficult to treat, having a 10-year overall survival of approximately 40%. Published genomic studies have not identified a recurrent driver mutation in PFA ependymomas, but they do show widespread epigenetic alterations, including global loss of histone H3 K27-trimethlyation (H3K27-me3). Another childhood PF tumor, the diffuse pontine glioma (DPG), also shows loss of H3K27-me3. In DPGs, loss of H3K27-me3 is associated with H3 K27M mutation, but the mechanism in PFA ependymomas is not yet established. We reexamined published PF ependymoma sequencing data and discovered recurrent mutations in a novel gene, CXorf67. Targeted sequencing in a series of PFA ependymomas revealed CXorf67 mutations in 22/234 (9.4%). CXorf67 is a single-exon gene of unknown function. Its protein product is predicted to be “disordered,” apart from one region towards the N terminus. Mutations in PFA ependymomas are missense, and there is a mutation hotspot in the “ordered” region. CXorf67 mutations are not present in other molecular groups of ependymoma and are rare in other cancers. Analyzing the DNA methylation profiles of 675 PFA ependymomas, we have discovered two subgroups, PFA-1 and PFA-2, and nine subtypes among these two subgroups. All PFA subtypes harbor CXorf67 mutations with the exception of PFA-1f and PFA-2c. Targeted sequencing in our tumor series also revealed H3 K27M mutations in PFA ependymomas at a frequency of 3.9%. Two thirds of mutations in HIST1H3B, HIST1H3C, and H3F3A were found in PFA-1f ependymomas, among which H3 mutations were present at a frequency of 35%. Mutations in H3 genes and CXorf67 were mutually exclusive across our series of PFA ependymomas. We used Affymetrix u133v2 arrays to establish that CXorf67 is expressed at high levels in PFA ependymomas, in contrast to low levels in other ependymomas from the PF and supratentorial compartments. A mechanism for CXorf67 overexpression was revealed in a similar comparative analysis of CpG island methylation profiles, which showed that the promoter region of CXorf67 is hypomethylated in PFA tumors, but not in other ependymomas. Using immunohistochemical preparations, we detected expression of CXorf67 at the protein level in the nuclei of PFA ependymomas; PFB and supratentorial tumors were immunonegative. CXorf67 overexpression is found in all PFA molecular subtypes, except PFA-1f, which shows similar levels to those in supratentorial and PFB ependymomas. CXorf67 expression is unrelated to mutation status. Elevated CXorf67 expression is found in the Daoy and U2-OS cancer cell lines. We used immunoprecipitation (IP)/mass spectrometry (MS) to study proteins bound to CXorf67 in Daoy and U2-OS. Analysis of enriched peptides following immunoprecipitation of CXorf67 indicated that it binds EZH2, SUZ12, and EED, three core components of the PRC2 complex. Complementary immunoprecipitation of SUZ12 detected CXorf67. Detecting CXorf67 mutations in almost 10% of PFA ependymomas led to the discovery of overexpression related to promoter region hypomethylation in these tumors. Overexpression of CXorf67 was detected in all PFA ependymoma subtypes except PFA-1f, 35% of which harbor an H3 K27M mutation. The protein product of CXorf67 is found in tumor cell nuclei, where our IP/MS data suggest it is bound to PRC2. Our findings suggest that global loss of H3K27-me3 in PFA ependymomas could be related to overexpression of CXorf67 and its interaction with PRC2, except in tumors where alterations in histone H3 genes are responsible. However, the selective advantage of CXorf67 mutation is yet to be explained. Citation Format: Ji Wen, Jens-Martin Hübner, Wilda Orisme, Gang Wu, Bo Tang, Sujuan Jia, John Easton, Kelly Haupfear, Brian D. Freibaum, Hong Joo Kim, Anthony High, Junmin Peng, Ruth G. Tatevossian, J. Paul Taylor, Stefan M. Pfister, Jinghui Zhang, Kristian W. Pajtler, Marcel Kool, David W. Ellison. Overexpression and mutations of CXorf67 in “infant-type” posterior fossa type-A ependymomas [abstract]. In: Proceedings of the AACR Special Conference: Pediatric Cancer Research: From Basic Science to the Clinic; 2017 Dec 3-6; Atlanta, Georgia. Philadelphia (PA): AACR; Cancer Res 2018;78(19 Suppl):Abstract nr PR08.