Cynthia Hawkins

Medulloblastoma Comprises Four Distinct Molecular Variants

PURPOSE Recent genomic approaches have suggested the existence of multiple distinct subtypes of medulloblastoma. We studied a large cohort of medulloblastomas to determine how many subgroups of the disease exist, how they differ, and the extent of overlap between subgroups. METHODS We determined gene expression profiles and DNA copy number aberrations for 103 primary medulloblastomas. Bioinformatic tools were used for class discovery of medulloblastoma subgroups based on the most informative genes in the data set. Immunohistochemistry for subgroup-specific signature genes was used to determine subgroup affiliation for 294 nonoverlapping medulloblastomas on two independent tissue microarrays. RESULTS Multiple unsupervised analyses of transcriptional profiles identified the following four distinct, nonoverlapping molecular variants: WNT, SHH, group C, and group D. Supervised analysis of these four subgroups revealed significant subgroup-specific demographics, histology, metastatic status, and DNA copy number aberrations. Immunohistochemistry for DKK1 (WNT), SFRP1 (SHH), NPR3 (group C), and KCNA1 (group D) could reliably and uniquely classify formalin-fixed medulloblastomas in approximately 98% of patients. Group C patients (NPR3-positive tumors) exhibited a significantly diminished progression-free and overall survival irrespective of their metastatic status. CONCLUSION Our integrative genomics approach to a large cohort of medulloblastomas has identified four disparate subgroups with distinct demographics, clinical presentation, transcriptional profiles, genetic abnormalities, and clinical outcome. Medulloblastomas can be reliably assigned to subgroups through immunohistochemistry, thereby making medulloblastoma subclassification widely available. Future research on medulloblastoma and the development of clinical trials should take into consideration these four distinct types of medulloblastoma.

Central nervous system atypical teratoid rhabdoid tumours: The Canadian Paediatric Brain Tumour Consortium experience

BACKGROUND Atypical teratoid rhabdoid tumours (ATRT) are aggressive brain tumours mostly occurring in early childhood. Largest published series arise from registries and institutional experiences (1-4). The aim of this report is to provide population-based data to further characterise this rare entity and to delineate prognostic factors. PATIENTS AND METHODS A national retrospective study of children ⩽18years diagnosed with a central nervous system (CNS) ATRT between 1995 and 2007 was undertaken. All cases underwent central pathology review. RESULTS There were 50 patients (31 males; median age at diagnosis of 16.7months). Twelve patients were >36months. Infratentorial location accounted for 52% of all cases. Nineteen patients (38%) had metastatic disease. Fifteen (30%) underwent gross total resection (GTR). Ten patients (20%) underwent palliation. Among the 40 remaining patients, 22 received conventional chemotherapy and 18 received high dose chemotherapy regimens (HDC); nine received intrathecal chemotherapy and 15 received adjuvant radiation. Thirty of the 40 treated patients relapsed/progressed at a median time of 5.5months (0-32). The median survival time of the entire cohort was 13.5months (1-117.5months). Age, tumour location and metastatic status were not prognostic. Patients with GTR had a better survival (2years overall survival (OS): 60%±12.6 versus 21.7%±8.5, p=0.03). HDC conferred better outcome (2years OS 47.9%±12.1 versus 27.3%±9.5, p=0.036). Upfront radiation did not provide survival benefit. Six of the 12 survivors (50%) did not receive radiation. CONCLUSION The outcome of CNS ATRT remains poor. However, the use of HDC provides encouraging results. GTR is a significant prognostic factor. The role of adjuvant radiation remains unclear.

Medulloblastoma subgroups remain stable across primary and metastatic compartments

Medulloblastoma comprises four distinct molecular variants with distinct genetics, transcriptomes, and outcomes. Subgroup affiliation has been previously shown to remain stable at the time of recurrence, which likely reflects their distinct cells of origin. However, a therapeutically relevant question that remains unanswered is subgroup stability in the metastatic compartment. We assembled a cohort of 12-paired primary-metastatic tumors collected in the MAGIC consortium, and established their molecular subgroup affiliation by performing integrative gene expression and DNA methylation analysis. Frozen tissues were collected and profiled using Affymetrix gene expression arrays and Illumina methylation arrays. Class prediction and hierarchical clustering were performed using existing published datasets. Our molecular analysis, using consensus integrative genomic data, establishes the unequivocal maintenance of molecular subgroup affiliation in metastatic medulloblastoma. We further validated these findings by interrogating a non-overlapping cohort of 19 pairs of primary-metastatic tumors from the Burdenko Neurosurgical Institute using an orthogonal technique of immunohistochemical staining. This investigation represents the largest reported primary-metastatic paired cohort profiled to date and provides a unique opportunity to evaluate subgroup-specific molecular aberrations within the metastatic compartment. Our findings further support the hypothesis that medulloblastoma subgroups arise from distinct cells of origin, which are carried forward from ontogeny to oncology.

c-kit Expression and Mutational Analysis in Medulloblastoma

The proto-oncogene c-kit is a receptor tyrosine kinase recognized to initiate essential signal transduction pathways that transmit biological signals for cellular proliferation, differentiation, and metastasis. Aberrant expression or mutation of c-kit has been shown to be involved in the pathogenesis of many cancers. Studies using imatinib mesylate (STI 571, Gleevec, Novartis, East Hanover, NJ, USA), an inhibitor of the tyrosine kinases brc-abl, c-kit, and PDGFR, have shown significant response in patients with chronic myelogenous leukemia and gastrointestinal stromal tumor. With the aim of identifying additional groups of tumors that may use the stem cell factor/c-kit pathway and, secondarily, may be responsive to imatinib mesylate treatment, we looked at the expression of c-kit in medulloblastoma. Medulloblastoma, a highly invasive primitive neuroectodermal tumor of the cerebellum, is the most common, malignant central nervous system tumor of childhood. Histologic features of medulloblastoma have failed to provide an accurate prediction of the clinical-biological behavior of these tumors. Characterizing the genetic events that play a role in the biology of these tumors may allow for molecular sub-typing and could lead to the development of novel therapeutic strategies. This study evaluated c-kit expression and mutational status in 10 medulloblastoma tumor samples. All 10 medulloblastoma tumors expressed c-kit by reverse transcriptase-polymerase chain reaction and 9 by immunohistochemical analysis. All tumor samples were screened for mutations in exons 9, 11, and 13 of the c-kit gene by direct sequencing. No sequence abnormalities were detected in these exons. These experiments lead us to the conclusion that c-kit activation in medulloblastoma is independent of mutation.

The international diffuse intrinsic pontine glioma registry: an infrastructure to accelerate collaborative research for an orphan disease

Diffuse intrinsic pontine glioma (DIPG), a rare, often fatal childhood brain tumor, remains a major therapeutic challenge. In 2012, investigators, funded by the DIPG Collaborative (a philanthropic partnership among 29 private foundations), launched the International DIPG Registry (IDIPGR) to advance understanding of DIPG. Comprised of comprehensive deidentified but linked clinical, imaging, histopathological, and genomic repositories, the IDIPGR uses standardized case report forms for uniform data collection; serial imaging and histopathology are centrally reviewed by IDIPGR neuro-radiologists and neuro-pathologists, respectively. Tissue and genomic data, and cell cultures derived from autopsies coordinated by the IDIPGR are available to investigators for studies approved by the Scientific Advisory Committee. From April 2012 to December 2016, 670 patients diagnosed with DIPG have been enrolled from 55 participating institutions in the US, Canada, Australia and New Zealand. The radiology repository contains 3558 studies from 448 patients. The pathology repository contains tissue on 81 patients with another 98 samples available for submission. Fresh DIPG tissue from seven autopsies has been sent to investigators to develop primary cell cultures. The bioinformatics repository contains next-generation sequencing data on 66 tumors. Nine projects using data/tissue from the IDIPGR by 13 principle investigators from around the world are now underway. The IDIPGR, a successful alliance among philanthropic agencies and investigators, has developed and maintained a highly collaborative, hypothesis-driven research infrastructure for interdisciplinary and translational projects in DIPG to improve diagnosis, response assessment, treatment and outcome for patients.

Pediatric sinonasal undifferentiated carcinoma: case report and literature review.

aggressive malignancy arising from the nasal cavity and paranasal sinuses. Sinonasal undifferentiated carcinoma was first reported in 1986 by Frierson et al, and since then less than 100 cases have been reported in the literature.1 One previous pediatric case has been described in a 12-year-old boy in India.2 Sinonasal undifferentiated carcinoma is histologically classified as part of a spectrum of neuroendocrine carcinomas originating from the Schneiderian epithelium or the nasal ectoderm in the paranasal sinuses.1 Here, we report only the second case of SNUC to arise in childhood involving a teenager with disease arising from the nasal cavity and extending through the paranasal air sinuses into the anterior cranial fossa and brain.

Detecting Stem Cell Marker Expression Using the NanoString nCounter System

The use of embryonic stem cells (ESCs) and induced pluripotent stem cells (iPSCs) has become commonplace in the study of neuronal development, physiology, disease modelling, and therapy development. Due to the transient nature of working with these cells, it is important to regularly confirm the cell status as a naive stem cell versus a more defined neural progenitor cell (NPC). Classically, this has been done using a panel of specific antibodies to test for the expression of transcription factors known to be observed in ESCs, but not NPCs. However, this method is both time consuming and expensive. Here, we describe the use of the NanoString nCounter system for determining the levels of expression of key transcription factors that will effectively aid in determining the state of your stem cell cultures.

Feasibility and efficacy of second-line chemotherapy at recurrence for pediatric low-grade gliomas: A comparative population-based study

2069 Background: Chemotherapy is widely accepted as the first-line therapy for pediatric low-grade gliomas (PLGG). Treatment modalities for further progression are controversial. Radiation therapy is still widely used, but the role for repeated chemotherapy remains unclear. The aim of the study was to determine treatment outcome for PLGG treated by chemotherapy at recurrence. Methods: The study group consisted of patients who received chemotherapy due to progression after initial chemotherapy protocol. These were compared to patients treated with chemotherapy at diagnosis, patients who received chemotherapy at third progression and to patients who received radiotherapy at recurrence. Results: From 1990–2007, 80 patients received chemotherapy as first-line treatment for PLGG. 36 received chemotherapy as second-line and 11 at further recurrence. 31 patients were treated with radiation at recurrence. There was no gender preference. Median follow-up time was 6.85 yrs. Histology revealed grade 1 astrocytoma in...

Microvessel density predicts behavior in pediatric optic pathway/hypothalamic gliomas

1556 Background: Optic pathway/hypothalamic gliomas (OPHG) are predominately low-grade tumors. However, OPHGs show an unpredictable behavior and there is no recognized histologic or molecular marker anticipating this behavior. This study was performed to investigate angiogenic features as possible independent prognostic factors. METHODS We searched the databases of the Hospital for Sick Children for patients with pathologically diagnosed OPHGs between 1985 and 2002. Tumor specimens were reviewed and reconfirmed as low-grade gliomas. Those with sufficient tissue for staining were included in the study. Sections were immunostained with factor VIII (F8) and counted for microvessel density (MD). A ratio of alpha-smooth muscle actin to F8 immunostaining was calculated to give a vascular maturity index. Vascular endothelial growth factor (VEGF) and VEGF-receptor immunostaining was performed to assess angiogenic features and MIB-1 labeling index (LI) to assess proliferation. These factors were evaluated with respect to progression-free survival (PFS) and outcome of treatment. RESULTS 60 patients were identified and 41 had sufficient material for further analysis. All patients underwent surgery. 30 required additional treatment (16 chemotherapy, 8 radiation, 6 chemotherapy + radiation). 8 patients had NF1. 38 are alive. MD showed a wide range of variation between tumors (4.8-73.6, median: 28 vessels/1.2 mm2). The absolute number of F8 stained vessels was significantly higher in infants (p < 0.01). A high MD (> 20 vessels/1.2mm2) was associated with a significantly shorter PFS compared to tumors with a low MD (<20 vessels/1.2 mm2)(p=0.025). MIB-1 values ranged from 0-10 (median 2.5). Intensity and distribution of VEGF and VEGFR staining and MIB-1 LI were not significantly associated with outcome. CONCLUSIONS Our findings suggest that angiogenesis is important in low-grade glioma and MD rather than MIB-1 has a prognostic value in OPHGs. [Table: see text].