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UCSF Pediatric Bithalamic Glioma Genome Project

ABSTRACT: Brain tumors are the most common solid tumors of childhood, and the genetic drivers and optimal therapeutic strategies for many of the different subtypes remain unknown. We performed targeted next-generation sequencing of approximately 500 cancer-associated genes on a cohort of 13 pediatric bithalamic diffuse gliomas, a lethal brain tumor of childhood for which the genetic basis is largely unknown. We identified that bithalamic diffuse gliomas harbor frequent mutations in the EGFR oncogene in the absence of accompanying gene amplification and only rare histone H3 mutation. These EGFR mutations were either small in-frame insertions within exon 20 (intracellular tyrosine kinase domain) or missense mutations within exon 7 (extracellular ligand-binding domain). Accompanying alterations included frequent TP53 mutation, CDK6 amplification or CDKN2C mutation, and BCOR and BCORL1 mutation or deletion.

PROVIDER: EGAS00001004033 | EGA |

REPOSITORIES: EGA

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Project description:Purpose: To define copy number alterations and gene expression signatures underlying pediatric high-grade glioma (HGG). Patients and Methods: We conducted a high-resolution analysis of genomic imbalances in 78 de novo pediatric HGG, including 7 diffuse intrinsic pontine gliomas, and 10 HGG cases arising in children who received cranial irradiation for a previous cancer, using Affymetrix 500K GeneChips. Gene expression signatures for 53 tumors were analyzed with Affymetrix U133v2 arrays. Results were compared with publicly available data from adult tumors. Results: Pediatric and adult glioblastoma were clearly distinguished by frequent gain of chromosome 1q (30% vs 9%) and lower frequency of chromosome 7 gain (13% vs 74%), respectively. The most common focal amplifications also differed, with PDGFRA and EGFR predominant in childhood and adult populations respectively. These common alterations in pediatric HGG were detected at higher frequency in irradiation-induced tumors, suggesting that these are initiating events in childhood gliomagenesis. CDKN2A was the most common tumor suppressor gene targeted by homozygous deletion in pediatric HGG. No IDH1 hotspot mutations were found in pediatric tumors, highlighting molecular differences in pathogenesis between childhood HGG and adult secondary glioblastoma. Integrated copy number and gene expression data indicated that deregulated PDGFRA signaling plays a major role in pediatric HGG. Conclusions: Integrated molecular profiling showed substantial differences in the molecular features underlying pediatric and adult HGG, indicating that findings in adult tumors cannot be simply extrapolated to younger patients. PDGFRA may be a useful target for pediatric HGG including diffuse pontine gliomas. Keywords: disease state analysis 78 samples for SNP analysis, including 10 samples arising in children who received cranial irradiation for a previous cancer and 7 diffuse pontine gliomas; 53 of them with gene expression analysis; 2 tumor grades To have access to SNP CEL files, please contact Dr. Suzanne Baker (suzzane.baker@stjude.org).

Project description:The outcome for children with high-grade gliomas (HGG) remains dismal, with a two-year survival rate of only 10-30%. Approximately half of pediatric HGGs are diffuse intrinsic pontine glioma (DIPG), a brainstem tumor that arises almost exclusively in children. Genome-wide analyses of copy number imbalances previously showed that platelet derived growth factor receptor alpha (PDGFRA) is the most frequent target of focal amplification in pediatric HGGs. To determine whether the PDGFRA is also targeted by more subtle mutations not detected by copy number analysis, we sequenced all PDGFRA coding exons from a cohort of pediatric HGGs. Somatic activating mutations were identified in 14.4% (13/90) of non-brainstem pediatric HGGs and 4.7% (2/43) of DIPGs, including missense mutations and in-frame deletions and insertions not previously described. 40% of tumors with mutation showed concurrent amplification, while 60% carried heterozygous mutations. Six different mutations impacting different domains all resulted in ligand-independent receptor activation that was blocked by small molecule inhibitors of PDGFR. Expression of mutants in p53-null primary mouse astrocytes conferred a proliferative advantage in vitro, and generated HGGs in vivo with complete penetrance when implanted into brain. The gene expression signatures reflected the spectrum of human diffuse HGGs. PDGFRA intragenic deletion of exons 8 and 9 were previously shown in adult HGG, but were not detected in 83 non-brainstem pediatric HGG and 57 DIPGs. Thus, a distinct spectrum of mutations confers constitutive receptor activation and oncogenic activity to PDGFR in childhood HGG. To better understand the consequence of PDGFRα mutation in pediatric gliomagenesis, retroviral constructs expressing wild-type PDGFRα or six selected PDGFRα mutants that affect different regions of the receptor were generated for functional studies. p53-null primary mouse astrocyte (PMA) cultures were chosen as a relevant cellular background to assess PDGFRα function.

Project description:DNA methylation profiling of 26 individual isomorphic diffuse gliomas using Illumina HumanMethylation450 BeadChips or Illumina Infinium HumanMethylation850 BeadChips. Abstract: The “isomorphic subtype of diffuse astrocytoma” was identified histologically in 2004 as a supratentorial, highly differentiated glioma with low cellularity, low proliferation and focal diffuse brain infiltration. Patients typically had seizures since childhood and all were operated on as adults. To define the position of these lesions among brain tumours, we histologically, molecularly and clinically analysed 26 histologically prototypical isomorphic diffuse gliomas. Immunohistochemically, they were GFAP-positive, MAP2-, OLIG2- and CD34-negative, nuclear ATRX-expression was retained and proliferation was low. All 24 cases sequenced were IDH-wildtype. In cluster analyses of DNA-methylation data, isomorphic diffuse gliomas formed a group clearly distinct from other glial/glio-neuronal brain tumours and normal hemispheric tissue, most closely related to paediatric MYB/MYBL1-altered diffuse astrocytomas and angiocentric gliomas. Half of the isomorphic diffuse gliomas had copy number alterations of MYBL1 or MYB (13/25, 52%). Gene fusions of MYBL1 or MYB with various gene partners were identified in 11/22 (50%) and were associated with an increased RNA-expression of the respective MYB-family gene. Integrating copy number alterations and available RNA sequencing data, 20/26 (77%) of isomorphic diffuse gliomas demonstrated MYBL1 (54%) or MYB (23%) alterations. Clinically, 89% of patients were seizure-free after surgery and all had a good outcome. In summary, we here define a distinct benign tumour class belonging to the family of MYB/MYBL1-altered gliomas. Isomorphic diffuse glioma occurs both in children and adults, has a concise morphology, frequent MYBL1- and MYB-alterations and a specific DNA-methylation profile. As an exclusively histological diagnosis may be very challenging and as paediatric MYB/MYBL1-altered diffuse astrocytomas may have the same gene fusions, we consider DNA-methylation profiling very helpful for their identification.

Project description:We analyzed mutations in Epidermal Growth Factor Receptor (EGFR) Tyrosine kinase (TK) domain, EGFR expression and gene profiling in prostate carcinoma (PC) in order to find out molecular prognostic markers and supply a proof for EGFR targeted therapies. 100 glyofixx-fixed, paraffin-embedded PC specimens were recovered after radical prostatectomy from locally advanced PC patients. Exons from 18 to 21 of EGFR TK domain were amplified and sequenced. For the entire cohort, EGFR protein evaluation by immunohistochemistry was performed. Gene expression profile was analyzed on 51 out of 100 samples by whole genome microarray. Statistical tests were performed in order to detect any significant association between EGFR iperexpression and prognosis. None out of 100 specimens presented mutations in exon 18; 2 point mutations were identified in exon 19, 5 in exon 20 and 6 in exon 21. In addiction, 58 out of 100 patients had the same silent mutation, at codon 787 in exon 20. EGFR iper-expression was found in 36% of specimens and was significantly associated with biochemical relapse. Gene profiling analysis on mutated samples selected 29 modulated genes differentially expressed between mutated EGFR+ and mutated EGFR- samples; 4 down-regulated genes, EAF2, ABCC4, KLK3 and ANXA3 and one up-regulated gene, FOXC1, are involved in prostate cancer progression. Our findings suggest that a subgroup of PC patients could potentially benefit of EGFR targeted therapies. The EGFR protein evaluation could contribute to identify PC relapsers. Keywords: EGFR expression, TK mutations, target therapy, microarray data. In this work we studied mutation status and expression of Epidermal Growth Factor Receptor (EGFR) in a case series of 100 primary prostate cancer tissue specimens. Results indicate that 13% and 36% of PC patients presents EGFR tyrosine kinase domain mutations and iperexpression respectively, suggesting that this receptor could be a therapeutic target in progressive prostate cancer. The analysis of correlation between EGFR protein expression, mutations, clinical parameters and outcome allow us to identify EGFR as significantly associated with biochemical relapse and high Gleason score. Gene expression profiling in 51 of PC tissues led to the identification of a gene list which separated EGFR mutated patients according to EGFR protein expression. These results could give more information on clinical outcome and possible development of new targeted therapies.

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UCSF Pediatric Bithalamic Glioma Genome Project

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