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Droplet digital PCR minimum mutant allele frequency detection. (PPTX 38Â kb)

Droplet digital PCR H3K27M detection validation as compared to WES and Sanger Sequencing. (DOCX 12Â kb)

Droplet digital PCR annealing temperature comparison. (PPTX 173Â kb)

Clinical characteristics of H3K27M and H3WT paediatric thalamic glioma Canadian cohort. (PPTX 84Â kb)

Clinical characteristics of paediatric thalamic glioma Canadian cohort. (DOCX 12Â kb)

Droplet digital PCR minimum concentration detection. (PPTX 55Â kb)

Droplet digital PCR pre-PCR digestion versus no digestion comparison. (PPTX 102Â kb)

Droplet digital PCR pre-PCR digestion versus no digestion comparison. (PPTX 102Â kb)

Droplet digital PCR minimum mutant allele frequency detection. (PPTX 38Â kb)

Droplet digital PCR annealing temperature comparison. (PPTX 173Â kb)

Droplet digital PCR minimum concentration detection. (PPTX 55Â kb)

Tumors of the central nervous system constitute the most common type of solid pediatric tumors and account for the majority of mortality and morbidity in pediatric oncology. Brainstem glioma (BSG) is a type of brain tumor that arises in the medulla, pons or midbrain and accounts for approximately 10-20% of pediatric brain tumors. Diffuse gliomas of the pontine region, diffuse intrinsic pontine glioma (DIPG) are the most common type of brainstem tumor, accounting for about 58-78% of BSG and the number one cause of brain tumor related death in children with a 1-2 year survival. Despite this very little is understood about the biology of these tumors. To try to address this lack of knowledge we have undertaken genomic analysis of a series of DIPGs and compared their copy number changes to those seen in pediatric supratentorial high grade astrocytomas as well as low grade astrocytomas. Post mortem tumor and matched normal brain samples (n= 9) and surgical samples (n= 4) were collected for a series of DIPG patients. We performed high-resolution genetic analysis using whole-genome single-nucleotide polymorphism (SNP) arrays (Affymetrix 500K and 6.0). Data analysis was conducted using Partek Genome Suite and Copy Number Analysis Tool (Affymetrix). Analysis of copy number alterations returned hits in several cancer related pathways including MAPK, Wnt, prostate cancer and gliomagenesis. DIPG copy number was also compared to that of pediatric high grade (n= 20) and low grade (n= 30) astrocytomas. Unsupervised Pearson's Dissimilarity clustering resulted in 3 distinct groups. One contained 6 DIPGs; the second contained mostly LGA with a few HGA, while the third group contained mostly HGA with the remaining DIPGs clustering with this group. Although there were some similarities, DIPG are genetically distinct from supratentorial high grade and low grade astrocytomas. A better understanding of DIPG biology is needed in order to develop agents targeted more specifically towards this disease particularly given the dismal prognosis and unresponsive nature of these tumors to conventional treatment. Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 101st Annual Meeting of the American Association for Cancer Research; 2010 Apr 17-21; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2010;70(8 Suppl):Abstract nr 2129.

Histology Kaplan-Meier survival analysis within the A) SickKids cohort and B) Canadian cohort. (DOCX 12Â kb)

Clinical characteristics of H3K27M and H3WT paediatric thalamic glioma in the Canadian cohort. (DOCX 12Â kb)

Histology Kaplan-Meier survival analysis within the A) SickKids cohort and B) Canadian cohort. (DOCX 12Â kb)

Clinical characteristics of H3K27M and H3WT paediatric thalamic glioma in the Canadian cohort. (DOCX 12Â kb)

Paediatric brain tumours arising in the thalamus present significant diagnostic and therapeutic challenges to physicians due to their sensitive midline location. As such, genetic analysis for biomarkers to aid in the diagnosis, prognosis and treatment of these tumours is needed. Here, we identified 64 thalamic gliomas with clinical follow-up and characterized targeted genomic alterations using newly optimized droplet digital and NanoString-based assays. The median age at diagnosis was 9.25 years (range, 0.63–17.55) and median survival was 6.43 (range, 0.01–27.63) years. Our cohort contained 42 and 22 tumours reviewed as low and high grade gliomas, respectively. Five (12 %) low grade and 11 (50 %) high grade gliomas were positive for the H3F3A/HIST1H3B K27M (H3K27M) mutation. Kaplan-Meier survival analysis revealed significantly worse overall survival for patients harbouring the H3K27M mutation versus H3F3A/HIST1H3B wild type (H3WT) samples (log-rank p