Publications by authors named "Natalie Jäger"

38 Publications

Integrated molecular and clinical analysis of low-grade gliomas in children with neurofibromatosis type 1 (NF1).

Acta Neuropathol 2021 Feb 14. Epub 2021 Feb 14.

Department of Neurology, Washington University School of Medicine, 660 South Euclid Avenue, Box 8111, St. Louis, MO, 63110, USA.

Low-grade gliomas (LGGs) are the most common childhood brain tumor in the general population and in individuals with the Neurofibromatosis type 1 (NF1) cancer predisposition syndrome. Surgical biopsy is rarely performed prior to treatment in the setting of NF1, resulting in a paucity of tumor genomic information. To define the molecular landscape of NF1-associated LGGs (NF1-LGG), we integrated clinical data, histological diagnoses, and multi-level genetic/genomic analyses on 70 individuals from 25 centers worldwide. Whereas, most tumors harbored bi-allelic NF1 inactivation as the only genetic abnormality, 11% had additional mutations. Moreover, tumors classified as non-pilocytic astrocytoma based on DNA methylation analysis were significantly more likely to harbor these additional mutations. The most common secondary alteration was FGFR1 mutation, which conferred an additional growth advantage in multiple complementary experimental murine Nf1 models. Taken together, this comprehensive characterization has important implications for the management of children with NF1-LGG, distinct from their sporadic counterparts.
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http://dx.doi.org/10.1007/s00401-021-02276-5DOI Listing
February 2021

H3.3-K27M drives neural stem cell-specific gliomagenesis in a human iPSC-derived model.

Cancer Cell 2021 Jan 29. Epub 2021 Jan 29.

Hopp Children's Cancer Center Heidelberg (KiTZ), 69120 Heidelberg, Germany; Division of Pediatric Neurooncology (B062), German Cancer Research Center (DKFZ) and German Cancer Consortium (DKTK), 69120 Heidelberg, Germany; Department of Pediatric Hematology and Oncology, Heidelberg University Hospital, 69120 Heidelberg, Germany. Electronic address:

Diffuse intrinsic pontine glioma (DIPG) is an aggressive childhood tumor of the brainstem with currently no curative treatment available. The vast majority of DIPGs carry a histone H3 mutation leading to a lysine 27-to-methionine exchange (H3K27M). We engineered human induced pluripotent stem cells (iPSCs) to carry an inducible H3.3-K27M allele in the endogenous locus and studied the effects of the mutation in different disease-relevant neural cell types. H3.3-K27M upregulated bivalent promoter-associated developmental genes, producing diverse outcomes in different cell types. While being fatal for iPSCs, H3.3-K27M increased proliferation in neural stem cells (NSCs) and to a lesser extent in oligodendrocyte progenitor cells (OPCs). Only NSCs gave rise to tumors upon induction of H3.3-K27M and TP53 inactivation in an orthotopic xenograft model recapitulating human DIPGs. In NSCs, H3.3-K27M leads to maintained expression of stemness and proliferative genes and a premature activation of OPC programs that together may cause tumor initiation.
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http://dx.doi.org/10.1016/j.ccell.2021.01.005DOI Listing
January 2021

Bioinformatics workflows for clinical applications in precision oncology.

Authors:
Natalie Jäger

Semin Cancer Biol 2021 Jan 18. Epub 2021 Jan 18.

Hopp Children's Cancer Center Heidelberg (KiTZ) & Division of Pediatric Neurooncology, German Cancer Research Center (DKFZ), Heidelberg, Germany. Electronic address:

High-throughput molecular profiling of tumors is a fundamental aspect of precision oncology, enabling the identification of genomic alterations that can be targeted therapeutically. In this context, a patient is matched to a specific drug or therapy based on the tumor's underlying genetic driver events rather than the histologic classification. This approach requires extensive bioinformatics methodology and workflows, including raw sequencing data processing and quality control, variant calling and annotation, integration of different molecular data types, visualization and finally reporting the data to physicians, cancer researchers and pharmacologists in a format that is readily interpretable for clinical decision making. This review comprises a broad overview of these bioinformatics aspects and discusses the multiple analytical, technical and interpretational challenges that remain to efficiently translate molecular findings into personalized treatment recommendations.
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http://dx.doi.org/10.1016/j.semcancer.2020.12.020DOI Listing
January 2021

An extracellular vesicle-related gene expression signature identifies high-risk patients in medulloblastoma.

Neuro Oncol 2020 Nov 11. Epub 2020 Nov 11.

Department of Pediatric Hematology and Oncology, University Children's Hospital Münster, Münster, Germany.

Background: Medulloblastoma (MB) is a malignant brain tumor in childhood. It comprises four subgroups with different clinical behaviors. The aim of this study was to characterize the transcriptomic landscape of MB, both at the level of individual tumors as well as in large patient cohorts.

Methods: We used a combination of single-cell transcriptomics, cell culture models and biophysical methods such as nanoparticle tracking analysis and electron microscopy, to investigate intercellular communication in the MB tumor niche.

Results: Tumor cells of the SHH-MB subgroup show a differentiation blockade. These cells undergo extensive metabolic reprogramming. The gene expression profiles of individual tumor cells show a partial convergence with those of tumor-associated glial and immune cells. One possible cause is the transfer of extracellular vesicles (EVs) between cells in the tumor niche. We were able to detect EVs in co-culture models of MB tumor cells and oligodendrocytes. We also identified a gene expression signature, EVS, which shows overlap with the proteome profile of large oncosomes from prostate cancer cells. This signature is also present in MB patient samples. A high EVS expression is one common characteristic of tumors that occur in high-risk patients from different MB subgroups or subtypes.

Conclusions: With EVS, our study uncovered a novel gene expression signature that has a high prognostic significance across MB subgroups.
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http://dx.doi.org/10.1093/neuonc/noaa254DOI Listing
November 2020

Reduced chromatin binding of MYC is a key effect of HDAC inhibition in MYC amplified medulloblastoma.

Neuro Oncol 2021 Feb;23(2):226-239

Hopp Children's Cancer Center Heidelberg (KiTZ), Heidelberg, Germany.

Background: The sensitivity of myelocytomatosis oncogene (MYC) amplified medulloblastoma to class I histone deacetylase (HDAC) inhibition has been shown previously; however, understanding the underlying molecular mechanism is crucial for selection of effective HDAC inhibitors for clinical use. The aim of this study was to investigate the direct molecular interaction of MYC and class I HDAC2, and the impact of class I HDAC inhibition on MYC function.

Methods: Co-immunoprecipitation and mass spectrometry were used to determine the co-localization of MYC and HDAC2. Chromatin immunoprecipitation (ChIP) sequencing and gene expression profiling were used to analyze the co-localization of MYC and HDAC2 on DNA and the impact on transcriptional activity in primary tumors and a MYC amplified cell line treated with the class I HDAC inhibitor entinostat. The effect on MYC was investigated by quantitative real-time PCR, western blot, and immunofluorescence.

Results: HDAC2 is a cofactor of MYC in MYC amplified medulloblastoma. The MYC-HDAC2 complex is bound to genes defining the MYC-dependent transcriptional profile. Class I HDAC inhibition leads to stabilization and reduced DNA binding of MYC protein, inducing a downregulation of MYC activated genes (MAGs) and upregulation of MYC repressed genes (MRGs). MAGs and MRGs are characterized by opposing biological functions and by distinct enhancer-box distribution.

Conclusions: Our data elucidate the molecular interaction of MYC and HDAC2 and support a model in which inhibition of class I HDACs directly targets MYC's transactivating and transrepressing functions.
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http://dx.doi.org/10.1093/neuonc/noaa191DOI Listing
February 2021

INFORM2 NivEnt: The first trial of the INFORM2 biomarker driven phase I/II trial series: the combination of nivolumab and entinostat in children and adolescents with refractory high-risk malignancies.

BMC Cancer 2020 Jun 5;20(1):523. Epub 2020 Jun 5.

KiTZ Clinical Trial Unit, Hopp Children's Cancer Center Heidelberg (KiTZ), German Cancer Research Center (DKFZ) and Heidelberg University Hospital, Im Neuenheimer Feld 430, 69120, Heidelberg, Germany.

Background: Pediatric patients with relapsed or refractory disease represent a population with a desperate medical need. The aim of the INFORM (INdividualized Therapy FOr Relapsed Malignancies in Childhood) program is to translate next generation molecular diagnostics into a biomarker driven treatment strategy. The program consists of two major foundations: the INFORM registry providing a molecular screening platform and the INFORM2 series of biomarker driven phase I/II trials. The INFORM2 NivEnt trial aims to determine the recommended phase 2 dose (RP2D) of the combination treatment of nivolumab and entinostat (phase I) and to evaluate activity and safety (phase II).

Methods: This is an exploratory non-randomized, open-label, multinational and multicenter seamless phase I/II trial in children and adolescents with relapsed / refractory or progressive high-risk solid tumors and CNS tumors. The phase I is divided in 2 age cohorts: 12-21 years and 6-11 years and follows a 3 + 3 design with two dose levels for entinostat (2 mg/m and 4 mg/m once per week) and fixed dose nivolumab (3 mg/kg every 2 weeks). Patients entering the trial on RP2D can seamlessly enter phase II which consists of a biomarker defined four group basket trial: high mutational load (group A), high PD-L1 mRNA expression (group B), focal MYC(N) amplification (group C), low mutational load and low PD-L1 mRNA expression and no MYC(N) amplification (group D). A Bayesian adaptive design will be used to early stop cohorts that fail to show evidence of activity. The maximum number of patients is 128.

Discussion: This trial intends to exploit the immune enhancing effects of entinostat on nivolumab using an innovative biomarker driven approach in order to maximize the chance of detecting signs of activity. It prevents exposure to unnecessary risks by applying the Bayesian adaptive design for early stopping for futility. The adaptive biomarker driven design provides an innovative approach accelerating drug development and reducing exposure to investigational treatments in these vulnerable children at the same time.

Trial Registration: ClinicalTrials.gov, NCT03838042. Registered on 12 February 2019.
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http://dx.doi.org/10.1186/s12885-020-07008-8DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7275428PMC
June 2020

Germline Elongator mutations in Sonic Hedgehog medulloblastoma.

Nature 2020 04 1;580(7803):396-401. Epub 2020 Apr 1.

Hopp Children's Cancer Center Heidelberg (KiTZ), Heidelberg, Germany.

Cancer genomics has revealed many genes and core molecular processes that contribute to human malignancies, but the genetic and molecular bases of many rare cancers remains unclear. Genetic predisposition accounts for 5 to 10% of cancer diagnoses in children, and genetic events that cooperate with known somatic driver events are poorly understood. Pathogenic germline variants in established cancer predisposition genes have been recently identified in 5% of patients with the malignant brain tumour medulloblastoma. Here, by analysing all protein-coding genes, we identify and replicate rare germline loss-of-function variants across ELP1 in 14% of paediatric patients with the medulloblastoma subgroup Sonic Hedgehog (MB) ELP1 was the most common medulloblastoma predisposition gene and increased the prevalence of genetic predisposition to 40% among paediatric patients with MB. Parent-offspring and pedigree analyses identified two families with a history of paediatric medulloblastoma. ELP1-associated medulloblastomas were restricted to the molecular SHHα subtype and characterized by universal biallelic inactivation of ELP1 owing to somatic loss of chromosome arm 9q. Most ELP1-associated medulloblastomas also exhibited somatic alterations in PTCH1, which suggests that germline ELP1 loss-of-function variants predispose individuals to tumour development in combination with constitutive activation of SHH signalling. ELP1 is the largest subunit of the evolutionarily conserved Elongator complex, which catalyses translational elongation through tRNA modifications at the wobble (U) position. Tumours from patients with ELP1-associated MB were characterized by a destabilized Elongator complex, loss of Elongator-dependent tRNA modifications, codon-dependent translational reprogramming, and induction of the unfolded protein response, consistent with loss of protein homeostasis due to Elongator deficiency in model systems. Thus, genetic predisposition to proteome instability may be a determinant in the pathogenesis of paediatric brain cancers. These results support investigation of the role of protein homeostasis in other cancer types and potential for therapeutic interference.
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http://dx.doi.org/10.1038/s41586-020-2164-5DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7430762PMC
April 2020

Germline Mutations Predispose to Pediatric Medulloblastoma.

J Clin Oncol 2020 01 14;38(1):43-50. Epub 2019 Oct 14.

RWTH Aachen University, Aachen, Germany.

Purpose: The identification of a heritable tumor predisposition often leads to changes in management and increased surveillance of individuals who are at risk; however, for many rare entities, our knowledge of heritable predisposition is incomplete.

Methods: Families with childhood medulloblastoma, one of the most prevalent childhood malignant brain tumors, were investigated to identify predisposing germline mutations. Initial findings were extended to genomes and epigenomes of 1,044 medulloblastoma cases from international multicenter cohorts, including retrospective and prospective clinical studies and patient series.

Results: We identified heterozygous germline mutations in the G protein-coupled receptor 161 () gene in six patients with infant-onset medulloblastoma (median age, 1.5 years). mutations were exclusively associated with the sonic hedgehog medulloblastoma (MB) subgroup and accounted for 5% of infant MB cases in our cohorts. Molecular tumor profiling revealed a loss of heterozygosity at in all affected MB tumors, atypical somatic copy number landscapes, and no additional somatic driver events. Analysis of 226 MB tumors revealed somatic copy-neutral loss of heterozygosity of chromosome 1q as the hallmark characteristic of deficiency and the primary mechanism for biallelic inactivation of in affected MB tumors.

Conclusion: Here, we describe a novel brain tumor predisposition syndrome that is caused by germline mutations and characterized by MB in infants. Additional studies are needed to identify a potential broader tumor spectrum associated with germline mutations.
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http://dx.doi.org/10.1200/JCO.19.00577DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6943973PMC
January 2020

Chromatin Remodeling in Response to BRCA2-Crisis.

Cell Rep 2019 08;28(8):2182-2193.e6

Department of Genetics, Stanford University, Stanford, CA 94305, USA. Electronic address:

Individuals with a single functional copy of the BRCA2 tumor suppressor have elevated risks for breast, ovarian, and other solid tumor malignancies. The exact mechanisms of carcinogenesis due to BRCA2 haploinsufficiency remain unclear, but one possibility is that at-risk cells are subject to acute periods of decreased BRCA2 availability and function ("BRCA2-crisis"), which may contribute to disease. Here, we establish an in vitro model for BRCA2-crisis that demonstrates chromatin remodeling and activation of an NF-κB survival pathway in response to transient BRCA2 depletion. Mechanistically, we identify BRCA2 chromatin binding, histone acetylation, and associated transcriptional activity as critical determinants of the epigenetic response to BRCA2-crisis. These chromatin alterations are reflected in transcriptional profiles of pre-malignant tissues from BRCA2 carriers and, therefore, may reflect natural steps in human disease. By modeling BRCA2-crisis in vitro, we have derived insights into pre-neoplastic molecular alterations that may enhance the development of preventative therapies.
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http://dx.doi.org/10.1016/j.celrep.2019.07.057DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6754178PMC
August 2019

RF_Purify: a novel tool for comprehensive analysis of tumor-purity in methylation array data based on random forest regression.

BMC Bioinformatics 2019 Aug 16;20(1):428. Epub 2019 Aug 16.

Division of Pediatric Neurooncology, German Cancer Research Center (DKFZ), Heidelberg, Germany.

Background: With the advent of array-based techniques to measure methylation levels in primary tumor samples, systematic investigations of methylomes have widely been performed on a large number of tumor entities. Most of these approaches are not based on measuring individual cell methylation but rather the bulk tumor sample DNA, which contains a mixture of tumor cells, infiltrating immune cells and other stromal components. This raises questions about the purity of a certain tumor sample, given the varying degrees of stromal infiltration in different entities. Previous methods to infer tumor purity require or are based on the use of matching control samples which are rarely available. Here we present a novel, reference free method to quantify tumor purity, based on two Random Forest classifiers, which were trained on ABSOLUTE as well as ESTIMATE purity values from TCGA tumor samples. We subsequently apply this method to a previously published, large dataset of brain tumors, proving that these models perform well in datasets that have not been characterized with respect to tumor purity .

Results: Using two gold standard methods to infer purity - the ABSOLUTE score based on whole genome sequencing data and the ESTIMATE score based on gene expression data- we have optimized Random Forest classifiers to predict tumor purity in entities that were contained in the TCGA project. We validated these classifiers using an independent test data set and cross-compared it to other methods which have been applied to the TCGA datasets (such as ESTIMATE and LUMP). Using Illumina methylation array data of brain tumor entities (as published in Capper et al. (Nature 555:469-474,2018)) we applied this model to estimate tumor purity and find that subgroups of brain tumors display substantial differences in tumor purity.

Conclusions: Random forest- based tumor purity prediction is a well suited tool to extrapolate gold standard measures of purity to novel methylation array datasets. In contrast to other available methylation based tumor purity estimation methods, our classifiers do not need a priori knowledge about the tumor entity or matching control tissue to predict tumor purity.
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http://dx.doi.org/10.1186/s12859-019-3014-zDOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6697926PMC
August 2019

Molecular characteristics and therapeutic vulnerabilities across paediatric solid tumours.

Nat Rev Cancer 2019 08 12;19(8):420-438. Epub 2019 Jul 12.

Hopp Children's Cancer Center Heidelberg (KiTZ), Heidelberg, Germany.

The spectrum of tumours arising in childhood is fundamentally different from that seen in adults, and they are known to be divergent from adult malignancies in terms of cellular origins, epidemiology, genetic complexity, driver mutations and underlying mutational processes. Despite the immense knowledge generated through sequencing efforts and functional characterization of identified (epi-)genetic alterations over the past decade, the clinical implications of this knowledge have so far been limited. Novel preclinical platforms such as the European Innovative Therapies for Children with Cancer-Paediatric Preclinical Proof-of-Concept Platform and the US-based Pediatric Preclinical Testing Consortium are being developed to try to change this by aiming to recapitulate the extensive heterogeneity of paediatric tumours and thereby, hopefully, improve the ability to predict clinical benefit. Numerous studies have also been established worldwide to provide patients with access to real-time molecular profiling and the possibility of more precise mechanism-of-action-based treatments. In addition to tumour-intrinsic findings and mechanisms, ongoing studies are investigating features such as the immune microenvironment of paediatric tumours in comparison with adult cancers - currently of very timely clinical relevance. However, there is an ongoing need for rigorous preclinical biomarker and target validation to feed into the next generation of molecularly stratified clinical trials. This Review aims to provide a comprehensive state-of-the-art overview of the molecular landscape of paediatric solid tumours, including their underlying genomic alterations and interactions with the microenvironment, complemented with our current understanding of potential therapeutic vulnerabilities and how these can be preclinically tested using more accurate predictive methods. Finally, we provide an outlook on the challenges and opportunities associated with translating this overwhelming scientific progress into real clinical benefit.
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http://dx.doi.org/10.1038/s41568-019-0169-xDOI Listing
August 2019

High-Resolution Bisulfite-Sequencing of Peripheral Blood DNA Methylation in Early-Onset and Familial Risk Breast Cancer Patients.

Clin Cancer Res 2019 09 7;25(17):5301-5314. Epub 2019 Jun 7.

Department of Genetics, Stanford University, Stanford, California.

Purpose: Understanding and explaining hereditary predisposition to cancer has focused on the genetic etiology of the disease. However, mutations in known genes associated with breast cancer, such as and , account for less than 25% of familial cases of breast cancer. Recently, specific epigenetic modifications at have been shown to promote hereditary breast cancer, but the broader potential for epigenetic contribution to hereditary breast cancer is not yet well understood.

Experimental Design: We examined DNA methylation through deep bisulfite sequencing of CpG islands and known promoter or regulatory regions in peripheral blood DNA from 99 patients with familial or early-onset breast or ovarian cancer, 6 unaffected mutation carriers, and 49 unaffected controls.

Results: In 9% of patients, we observed altered methylation in the promoter regions of genes known to be involved in cancer, including hypermethylation at the tumor suppressor and hypomethylation at the proto-oncogene . These alterations occur in the form of allelic methylation that span up to hundreds of base pairs in length.

Conclusions: Our observations suggest a broader role for DNA methylation in early-onset, familial risk breast cancer. Further studies are warranted to clarify these mechanisms and the benefits of DNA methylation screening for early risk prediction of familial cancers.
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http://dx.doi.org/10.1158/1078-0432.CCR-18-2423DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6726519PMC
September 2019

Second-generation molecular subgrouping of medulloblastoma: an international meta-analysis of Group 3 and Group 4 subtypes.

Acta Neuropathol 2019 08 10;138(2):309-326. Epub 2019 May 10.

Wolfson Childhood Cancer Research Centre, Northern Institute for Cancer Research, Newcastle University, Newcastle upon Tyne, UK.

In 2012, an international consensus paper reported that medulloblastoma comprises four molecular subgroups (WNT, SHH, Group 3, and Group 4), each associated with distinct genomic features and clinical behavior. Independently, multiple recent reports have defined further intra-subgroup heterogeneity in the form of biologically and clinically relevant subtypes. However, owing to differences in patient cohorts and analytical methods, estimates of subtype number and definition have been inconsistent, especially within Group 3 and Group 4. Herein, we aimed to reconcile the definition of Group 3/Group 4 MB subtypes through the analysis of a series of 1501 medulloblastomas with DNA-methylation profiling data, including 852 with matched transcriptome data. Using multiple complementary bioinformatic approaches, we compared the concordance of subtype calls between published cohorts and analytical methods, including assessments of class-definition confidence and reproducibility. While the lowest complexity solutions continued to support the original consensus subgroups of Group 3 and Group 4, our analysis most strongly supported a definition comprising eight robust Group 3/Group 4 subtypes (types I-VIII). Subtype II was consistently identified across all component studies, while all others were supported by multiple class-definition methods. Regardless of analytical technique, increasing cohort size did not further increase the number of identified Group 3/Group 4 subtypes. Summarizing the molecular and clinico-pathological features of these eight subtypes indicated enrichment of specific driver gene alterations and cytogenetic events amongst subtypes, and identified highly disparate survival outcomes, further supporting their biological and clinical relevance. Collectively, this study provides continued support for consensus Groups 3 and 4 while enabling robust derivation of, and categorical accounting for, the extensive intertumoral heterogeneity within Groups 3 and 4, revealed by recent high-resolution subclassification approaches. Furthermore, these findings provide a basis for application of emerging methods (e.g., proteomics/single-cell approaches) which may additionally inform medulloblastoma subclassification. Outputs from this study will help shape definition of the next generation of medulloblastoma clinical protocols and facilitate the application of enhanced molecularly guided risk stratification to improve outcomes and quality of life for patients and their families.
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http://dx.doi.org/10.1007/s00401-019-02020-0DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6660496PMC
August 2019

DECIPHER pooled shRNA library screen identifies PP2A and FGFR signaling as potential therapeutic targets for diffuse intrinsic pontine gliomas.

Neuro Oncol 2019 07;21(7):867-877

Division of Molecular Genetics, German Cancer Research Center (DKFZ), Heidelberg, Germany.

Background: Diffuse intrinsic pontine gliomas (DIPGs) are highly aggressive pediatric brain tumors that are characterized by a recurrent mutation (K27M) within the histone H3 encoding genes H3F3A and HIST1H3A/B/C. These mutations have been shown to induce a global reduction in the repressive histone modification H3K27me3, which together with widespread changes in DNA methylation patterns results in an extensive transcriptional reprogramming hampering the identification of single therapeutic targets based on a molecular rationale.

Methods: We applied a large-scale gene knockdown approach using a pooled short hairpin (sh)RNA library in combination with next-generation sequencing in order to identify DIPG-specific vulnerabilities. The therapeutic potential of specific inhibitors of candidate targets was validated in a secondary drug screen.

Results: We identified fibroblast growth factor receptor (FGFR) signaling and the serine/threonine protein phosphatase 2A (PP2A) as top depleted hits in patient-derived DIPG cell cultures and validated their lethal potential by FGF ligand depletion and genetic knockdown of the PP2A structural subunit PPP2R1A. Further, pharmacological inhibition of FGFR and PP2A signaling through ponatinib and LB-100 treatment, respectively, exhibited strong tumor-specific anti-proliferative and apoptotic activity in cultured DIPG cells.

Conclusions: Our findings suggest FGFR and PP2A signaling as potential new therapeutic targets for the treatment of DIPGs.
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http://dx.doi.org/10.1093/neuonc/noz057DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6620639PMC
July 2019

The whole-genome landscape of medulloblastoma subtypes.

Nature 2017 07;547(7663):311-317

Division of Molecular Genetics, German Cancer Research Center (DKFZ), Heidelberg, Germany.

Current therapies for medulloblastoma, a highly malignant childhood brain tumour, impose debilitating effects on the developing child, and highlight the need for molecularly targeted treatments with reduced toxicity. Previous studies have been unable to identify the full spectrum of driver genes and molecular processes that operate in medulloblastoma subgroups. Here we analyse the somatic landscape across 491 sequenced medulloblastoma samples and the molecular heterogeneity among 1,256 epigenetically analysed cases, and identify subgroup-specific driver alterations that include previously undiscovered actionable targets. Driver mutations were confidently assigned to most patients belonging to Group 3 and Group 4 medulloblastoma subgroups, greatly enhancing previous knowledge. New molecular subtypes were differentially enriched for specific driver events, including hotspot in-frame insertions that target KBTBD4 and 'enhancer hijacking' events that activate PRDM6. Thus, the application of integrative genomics to an extensive cohort of clinical samples derived from a single childhood cancer entity revealed a series of cancer genes and biologically relevant subtype diversity that represent attractive therapeutic targets for the treatment of patients with medulloblastoma.
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http://dx.doi.org/10.1038/nature22973DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC5905700PMC
July 2017

Functional limitations in children and adolescents suffering from chronic pain: validation and psychometric properties of the German Functional Disability Inventory (FDI-G).

Rheumatol Int 2016 Oct 4;36(10):1439-48. Epub 2016 Jun 4.

German Center for Rheumatology in Children and Adolescents, Gehfeldstraße 24, 82467, Garmisch-Partenkirchen, Germany.

Our objective was to translate the Functional Disability Inventory (FDI) into German, to evaluate its validity and to assess functional limitation in a large cohort of children and adolescents with juvenile fibromyalgia syndrome (jFMS). We administered several questions (e.g., sociodemographics, school-related issues) and questionnaires to 329 patients and one parent. The questionnaires included, among others, a German version of the FDI, the CHAQ (parent report), KIDSCREEN, tender point score (TPS), Depression Inventory for Children and Adolescents (DIKJ) and others. Patients were asked about the severity of pain today (NRS = numerical rating scale) and other symptoms. Internal consistency was evaluated with Cronbach's alpha. Construct validity of the FDI was evaluated by correlating the FDI with the questionnaires as well as with the pain and other variables, e.g., days missed school. An exploratory factor analysis (EFA) was also performed. Mean age was 13.9 years (SD ±2.48). Means were for pain today 5.37 (±2.39) and for the TPS 39.71 (±21.56). Internal consistency was α = .90. Low-to-moderate correlations were obtained between the FDI and the CHAQ (ρ = .51**), KIDSCREEN (e.g., physical well-being ρ = -.62**; peers and social support ρ = -.28**) as well as the pain variables (NRS ρ = .24**; TPS ρ = .38**). Psychological variables were also correlated with the FDI (e.g., DIJK ρ = .28**). An EFA suggested a two-factor solution. The FDI is a valid instrument for measuring functional limitations in German children and adolescents with jFMS.
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http://dx.doi.org/10.1007/s00296-016-3504-5DOI Listing
October 2016

A comprehensive assessment of somatic mutation detection in cancer using whole-genome sequencing.

Nat Commun 2015 Dec 9;6:10001. Epub 2015 Dec 9.

CNAG-CRG, Centre for Genomic Regulation, Barcelona Institute of Science and Technology (BIST), Baldiri i Reixac 4, 08028 Barcelona, Spain.

As whole-genome sequencing for cancer genome analysis becomes a clinical tool, a full understanding of the variables affecting sequencing analysis output is required. Here using tumour-normal sample pairs from two different types of cancer, chronic lymphocytic leukaemia and medulloblastoma, we conduct a benchmarking exercise within the context of the International Cancer Genome Consortium. We compare sequencing methods, analysis pipelines and validation methods. We show that using PCR-free methods and increasing sequencing depth to ∼ 100 × shows benefits, as long as the tumour:control coverage ratio remains balanced. We observe widely varying mutation call rates and low concordance among analysis pipelines, reflecting the artefact-prone nature of the raw data and lack of standards for dealing with the artefacts. However, we show that, using the benchmark mutation set we have created, many issues are in fact easy to remedy and have an immediate positive impact on mutation detection accuracy.
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http://dx.doi.org/10.1038/ncomms10001DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC4682041PMC
December 2015

Enhancer hijacking activates GFI1 family oncogenes in medulloblastoma.

Nature 2014 Jul 22;511(7510):428-34. Epub 2014 Jun 22.

1] Division of Pediatric Neurooncology, German Cancer Research Center (DKFZ), Im Neuenheimer Feld 280, Heidelberg 69120, Germany [2] Department of Pediatric Oncology, Hematology & Immunology, Heidelberg University Hospital, Im Neuenheimer Feld 430, Heidelberg 69120, Germany.

Medulloblastoma is a highly malignant paediatric brain tumour currently treated with a combination of surgery, radiation and chemotherapy, posing a considerable burden of toxicity to the developing child. Genomics has illuminated the extensive intertumoral heterogeneity of medulloblastoma, identifying four distinct molecular subgroups. Group 3 and group 4 subgroup medulloblastomas account for most paediatric cases; yet, oncogenic drivers for these subtypes remain largely unidentified. Here we describe a series of prevalent, highly disparate genomic structural variants, restricted to groups 3 and 4, resulting in specific and mutually exclusive activation of the growth factor independent 1 family proto-oncogenes, GFI1 and GFI1B. Somatic structural variants juxtapose GFI1 or GFI1B coding sequences proximal to active enhancer elements, including super-enhancers, instigating oncogenic activity. Our results, supported by evidence from mouse models, identify GFI1 and GFI1B as prominent medulloblastoma oncogenes and implicate 'enhancer hijacking' as an efficient mechanism driving oncogene activation in a childhood cancer.
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http://dx.doi.org/10.1038/nature13379DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC4201514PMC
July 2014

Decoding the regulatory landscape of medulloblastoma using DNA methylation sequencing.

Nature 2014 Jun 18;510(7506):537-41. Epub 2014 May 18.

1] Division of Molecular Genetics, German Cancer Research Center (DKFZ), Im Neuenheimer Feld 280, Heidelberg 69120, Germany [2] Heidelberg Center for Personalised Oncology (DKFZ-HIPO), Im Neuenheimer Feld 280, Heidelberg 69120, Germany.

Epigenetic alterations, that is, disruption of DNA methylation and chromatin architecture, are now acknowledged as a universal feature of tumorigenesis. Medulloblastoma, a clinically challenging, malignant childhood brain tumour, is no exception. Despite much progress from recent genomics studies, with recurrent changes identified in each of the four distinct tumour subgroups (WNT-pathway-activated, SHH-pathway-activated, and the less-well-characterized Group 3 and Group 4), many cases still lack an obvious genetic driver. Here we present whole-genome bisulphite-sequencing data from thirty-four human and five murine tumours plus eight human and three murine normal controls, augmented with matched whole-genome, RNA and chromatin immunoprecipitation sequencing data. This comprehensive data set allowed us to decipher several features underlying the interplay between the genome, epigenome and transcriptome, and its effects on medulloblastoma pathophysiology. Most notable were highly prevalent regions of hypomethylation correlating with increased gene expression, extending tens of kilobases downstream of transcription start sites. Focal regions of low methylation linked to transcription-factor-binding sites shed light on differential transcriptional networks between subgroups, whereas increased methylation due to re-normalization of repressed chromatin in DNA methylation valleys was positively correlated with gene expression. Large, partially methylated domains affecting up to one-third of the genome showed increased mutation rates and gene silencing in a subgroup-specific fashion. Epigenetic alterations also affected novel medulloblastoma candidate genes (for example, LIN28B), resulting in alternative promoter usage and/or differential messenger RNA/microRNA expression. Analysis of mouse medulloblastoma and precursor-cell methylation demonstrated a somatic origin for many alterations. Our data provide insights into the epigenetic regulation of transcription and genome organization in medulloblastoma pathogenesis, which are probably also of importance in a wider developmental and disease context.
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http://dx.doi.org/10.1038/nature13268DOI Listing
June 2014

Genome sequencing of SHH medulloblastoma predicts genotype-related response to smoothened inhibition.

Cancer Cell 2014 Mar;25(3):393-405

Division of Pediatric Neurooncology, German Cancer Research Center (DKFZ), 69121 Heidelberg, Germany; Department of Pediatric Oncology, Hematology and Immunology, University Hospital Heidelberg, 69120 Heidelberg, Germany.

Smoothened (SMO) inhibitors recently entered clinical trials for sonic-hedgehog-driven medulloblastoma (SHH-MB). Clinical response is highly variable. To understand the mechanism(s) of primary resistance and identify pathways cooperating with aberrant SHH signaling, we sequenced and profiled a large cohort of SHH-MBs (n = 133). SHH pathway mutations involved PTCH1 (across all age groups), SUFU (infants, including germline), and SMO (adults). Children >3 years old harbored an excess of downstream MYCN and GLI2 amplifications and frequent TP53 mutations, often in the germline, all of which were rare in infants and adults. Functional assays in different SHH-MB xenograft models demonstrated that SHH-MBs harboring a PTCH1 mutation were responsive to SMO inhibition, whereas tumors harboring an SUFU mutation or MYCN amplification were primarily resistant.
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http://dx.doi.org/10.1016/j.ccr.2014.02.004DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC4493053PMC
March 2014

Hypermutation of the inactive X chromosome is a frequent event in cancer.

Cell 2013 Oct 17;155(3):567-81. Epub 2013 Oct 17.

Division of Theoretical Bioinformatics, German Cancer Research Center (DKFZ), Im Neuenheimer Feld 280, 69120 Heidelberg, Germany.

Mutation is a fundamental process in tumorigenesis. However, the degree to which the rate of somatic mutation varies across the human genome and the mechanistic basis underlying this variation remain to be fully elucidated. Here, we performed a cross-cancer comparison of 402 whole genomes comprising a diverse set of childhood and adult tumors, including both solid and hematopoietic malignancies. Surprisingly, we found that the inactive X chromosome of many female cancer genomes accumulates on average twice and up to four times as many somatic mutations per megabase, as compared to the individual autosomes. Whole-genome sequencing of clonally expanded hematopoietic stem/progenitor cells (HSPCs) from healthy individuals and a premalignant myelodysplastic syndrome (MDS) sample revealed no X chromosome hypermutation. Our data suggest that hypermutation of the inactive X chromosome is an early and frequent feature of tumorigenesis resulting from DNA replication stress in aberrantly proliferating cells.
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http://dx.doi.org/10.1016/j.cell.2013.09.042DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC3898475PMC
October 2013

Signatures of mutational processes in human cancer.

Nature 2013 Aug 14;500(7463):415-21. Epub 2013 Aug 14.

Cancer Genome Project, Wellcome Trust Sanger Institute, Wellcome Trust Genome Campus, Hinxton, Cambridgeshire CB10 1SA, UK.

All cancers are caused by somatic mutations; however, understanding of the biological processes generating these mutations is limited. The catalogue of somatic mutations from a cancer genome bears the signatures of the mutational processes that have been operative. Here we analysed 4,938,362 mutations from 7,042 cancers and extracted more than 20 distinct mutational signatures. Some are present in many cancer types, notably a signature attributed to the APOBEC family of cytidine deaminases, whereas others are confined to a single cancer class. Certain signatures are associated with age of the patient at cancer diagnosis, known mutagenic exposures or defects in DNA maintenance, but many are of cryptic origin. In addition to these genome-wide mutational signatures, hypermutation localized to small genomic regions, 'kataegis', is found in many cancer types. The results reveal the diversity of mutational processes underlying the development of cancer, with potential implications for understanding of cancer aetiology, prevention and therapy.
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http://dx.doi.org/10.1038/nature12477DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC3776390PMC
August 2013

Recurrent somatic alterations of FGFR1 and NTRK2 in pilocytic astrocytoma.

Nat Genet 2013 Aug 30;45(8):927-32. Epub 2013 Jun 30.

Division of Pediatric Neurooncology, German Cancer Research Center (DKFZ), Heidelberg, Germany.

Pilocytic astrocytoma, the most common childhood brain tumor, is typically associated with mitogen-activated protein kinase (MAPK) pathway alterations. Surgically inaccessible midline tumors are therapeutically challenging, showing sustained tendency for progression and often becoming a chronic disease with substantial morbidities. Here we describe whole-genome sequencing of 96 pilocytic astrocytomas, with matched RNA sequencing (n = 73), conducted by the International Cancer Genome Consortium (ICGC) PedBrain Tumor Project. We identified recurrent activating mutations in FGFR1 and PTPN11 and new NTRK2 fusion genes in non-cerebellar tumors. New BRAF-activating changes were also observed. MAPK pathway alterations affected all tumors analyzed, with no other significant mutations identified, indicating that pilocytic astrocytoma is predominantly a single-pathway disease. Notably, we identified the same FGFR1 mutations in a subset of H3F3A-mutated pediatric glioblastoma with additional alterations in the NF1 gene. Our findings thus identify new potential therapeutic targets in distinct subsets of pilocytic astrocytoma and childhood glioblastoma.
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http://dx.doi.org/10.1038/ng.2682DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC3951336PMC
August 2013

Coverage bias and sensitivity of variant calling for four whole-genome sequencing technologies.

PLoS One 2013 11;8(6):e66621. Epub 2013 Jun 11.

Division of Theoretical Bioinformatics, German Cancer Research Center (DKFZ), Heidelberg, Germany.

The emergence of high-throughput, next-generation sequencing technologies has dramatically altered the way we assess genomes in population genetics and in cancer genomics. Currently, there are four commonly used whole-genome sequencing platforms on the market: Illumina's HiSeq2000, Life Technologies' SOLiD 4 and its completely redesigned 5500xl SOLiD, and Complete Genomics' technology. A number of earlier studies have compared a subset of those sequencing platforms or compared those platforms with Sanger sequencing, which is prohibitively expensive for whole genome studies. Here we present a detailed comparison of the performance of all currently available whole genome sequencing platforms, especially regarding their ability to call SNVs and to evenly cover the genome and specific genomic regions. Unlike earlier studies, we base our comparison on four different samples, allowing us to assess the between-sample variation of the platforms. We find a pronounced GC bias in GC-rich regions for Life Technologies' platforms, with Complete Genomics performing best here, while we see the least bias in GC-poor regions for HiSeq2000 and 5500xl. HiSeq2000 gives the most uniform coverage and displays the least sample-to-sample variation. In contrast, Complete Genomics exhibits by far the smallest fraction of bases not covered, while the SOLiD platforms reveal remarkable shortcomings, especially in covering CpG islands. When comparing the performance of the four platforms for calling SNPs, HiSeq2000 and Complete Genomics achieve the highest sensitivity, while the SOLiD platforms show the lowest false positive rate. Finally, we find that integrating sequencing data from different platforms offers the potential to combine the strengths of different technologies. In summary, our results detail the strengths and weaknesses of all four whole-genome sequencing platforms. It indicates application areas that call for a specific sequencing platform and disallow other platforms. This helps to identify the proper sequencing platform for whole genome studies with different application scopes.
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http://journals.plos.org/plosone/article?id=10.1371/journal.pone.0066621PLOS
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC3679043PMC
January 2014

Error detection and error memory in spatial navigation as reflected by electrodermal activity.

Cogn Process 2013 Nov 23;14(4):377-89. Epub 2013 May 23.

Biomedical Optics Research Laboratory (BORL), Division of Neonatology, University Hospital Zurich, Frauenklinikstrasse 10, 8091, Zurich, Switzerland,

The study investigated spatial navigation by means of electrodermal activity (EDA). Two groups of healthy subjects (group 1, age <38; group 2, age ≥ 38) were recorded during navigation through two 3-D virtual mazes differing in difficulty, that is, Maze Simple (MazeS) and Maze Complex (MazeC). Our results show (1) an effect of difficulty, that is, larger skin conductance responses (SCRs) and slower velocity profiles while navigating through MazeC as compared to MazeS. (2) An effect of age, that is, larger SCRs and faster velocity profiles in younger subjects (group 1) compared to older subjects (group 2). (3) An effect of maze region, that is, SCRs increased when subjects entered dead ends with group 1 (young group) decreasing in velocity, whereas group 2 (old group) increased in velocity. (4) And an error memory effect, that is, subjects who remembered an error at a given decision point (crossroads preceding dead ends in MazeC) from previous trials, and then if they did not repeat that error, elicited decreased SCRs as compared to subjects who did not remember and subsequently repeated an error. The latter aspect is the most impactful as it shows that EDA is able to reflect error detection and memory during spatial navigation. Our data designate EDA as suitable monitoring tool for identification and differentiation of the affective correlates underlying spatial navigation, which has recently attracted researchers' attention due to its increased use in 3-D virtual environments.
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http://dx.doi.org/10.1007/s10339-013-0567-zDOI Listing
November 2013

Integrative genomic analyses reveal an androgen-driven somatic alteration landscape in early-onset prostate cancer.

Cancer Cell 2013 Feb;23(2):159-70

Genome Biology Unit, European Molecular Biology Laboratory (EMBL), Meyerhofstr. 1, 69117 Heidelberg, Germany.

Early-onset prostate cancer (EO-PCA) represents the earliest clinical manifestation of prostate cancer. To compare the genomic alteration landscapes of EO-PCA with "classical" (elderly-onset) PCA, we performed deep sequencing-based genomics analyses in 11 tumors diagnosed at young age, and pursued comparative assessments with seven elderly-onset PCA genomes. Remarkable age-related differences in structural rearrangement (SR) formation became evident, suggesting distinct disease pathomechanisms. Whereas EO-PCAs harbored a prevalence of balanced SRs, with a specific abundance of androgen-regulated ETS gene fusions including TMPRSS2:ERG, elderly-onset PCAs displayed primarily non-androgen-associated SRs. Data from a validation cohort of > 10,000 patients showed age-dependent androgen receptor levels and a prevalence of SRs affecting androgen-regulated genes, further substantiating the activity of a characteristic "androgen-type" pathomechanism in EO-PCA.
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http://dx.doi.org/10.1016/j.ccr.2013.01.002DOI Listing
February 2013

Secretory meningiomas are defined by combined KLF4 K409Q and TRAF7 mutations.

Acta Neuropathol 2013 Mar 12;125(3):351-8. Epub 2013 Feb 12.

Department of Neuropathology, Institute of Pathology, Ruprecht-Karls-University Heidelberg, Im Neuenheimer Feld 224, 69120, Heidelberg, Germany.

Meningiomas are among the most frequent intracranial tumors. The secretory variant of meningioma is characterized by glandular differentiation, formation of intracellular lumina and pseudopsammoma bodies, expression of a distinct pattern of cytokeratins and clinically by pronounced perifocal brain edema. Here we describe whole-exome sequencing analysis of DNA from 16 secretory meningiomas and corresponding constitutional tissues. All secretory meningiomas invariably harbored a mutation in both KLF4 and TRAF7. Validation in an independent cohort of 14 secretory meningiomas by Sanger sequencing or derived cleaved amplified polymorphic sequence (dCAPS) assay detected the same pattern, with KLF4 mutations observed in a total of 30/30 and TRAF7 mutations in 29/30 of these tumors. All KLF4 mutations were identical, affected codon 409 and resulted in a lysine to glutamine exchange (K409Q). KLF4 mutations were not found in 89 non-secretory meningiomas, 267 other intracranial tumors including gliomas, glioneuronal tumors, pituitary adenomas and metastases, 59 peripheral nerve sheath tumors and 52 pancreatic tumors. TRAF7 mutations were restricted to the WD40 domains. While KLF4 mutations were exclusively seen in secretory meningiomas, TRAF7 mutations were also observed in 7/89 (8 %) of non-secretory meningiomas. KLF4 and TRAF7 mutations were mutually exclusive with NF2 mutations. In conclusion, our findings suggest an essential contribution of combined KLF4 K409Q and TRAF7 mutations in the genesis of secretory meningioma and demonstrate a role for TRAF7 alterations in other non-NF2 meningiomas.
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http://dx.doi.org/10.1007/s00401-013-1093-xDOI Listing
March 2013

Dissecting the genomic complexity underlying medulloblastoma.

Nature 2012 Aug;488(7409):100-5

Division of Pediatric Neurooncology, German Cancer Research Center (DKFZ), Im Neuenheimer Feld 280, Heidelberg 69120, Germany.

Medulloblastoma is an aggressively growing tumour, arising in the cerebellum or medulla/brain stem. It is the most common malignant brain tumour in children, and shows tremendous biological and clinical heterogeneity. Despite recent treatment advances, approximately 40% of children experience tumour recurrence, and 30% will die from their disease. Those who survive often have a significantly reduced quality of life. Four tumour subgroups with distinct clinical, biological and genetic profiles are currently identified. WNT tumours, showing activated wingless pathway signalling, carry a favourable prognosis under current treatment regimens. SHH tumours show hedgehog pathway activation, and have an intermediate prognosis. Group 3 and 4 tumours are molecularly less well characterized, and also present the greatest clinical challenges. The full repertoire of genetic events driving this distinction, however, remains unclear. Here we describe an integrative deep-sequencing analysis of 125 tumour-normal pairs, conducted as part of the International Cancer Genome Consortium (ICGC) PedBrain Tumor Project. Tetraploidy was identified as a frequent early event in Group 3 and 4 tumours, and a positive correlation between patient age and mutation rate was observed. Several recurrent mutations were identified, both in known medulloblastoma-related genes (CTNNB1, PTCH1, MLL2, SMARCA4) and in genes not previously linked to this tumour (DDX3X, CTDNEP1, KDM6A, TBR1), often in subgroup-specific patterns. RNA sequencing confirmed these alterations, and revealed the expression of what are, to our knowledge, the first medulloblastoma fusion genes identified. Chromatin modifiers were frequently altered across all subgroups. These findings enhance our understanding of the genomic complexity and heterogeneity underlying medulloblastoma, and provide several potential targets for new therapeutics, especially for Group 3 and 4 patients.
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http://dx.doi.org/10.1038/nature11284DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC3662966PMC
August 2012

Medulloblastoma exome sequencing uncovers subtype-specific somatic mutations.

Nature 2012 Aug;488(7409):106-10

Broad Institute of MIT and Harvard, Cambridge, Massachusetts 02142, USA.

Medulloblastomas are the most common malignant brain tumours in children. Identifying and understanding the genetic events that drive these tumours is critical for the development of more effective diagnostic, prognostic and therapeutic strategies. Recently, our group and others described distinct molecular subtypes of medulloblastoma on the basis of transcriptional and copy number profiles. Here we use whole-exome hybrid capture and deep sequencing to identify somatic mutations across the coding regions of 92 primary medulloblastoma/normal pairs. Overall, medulloblastomas have low mutation rates consistent with other paediatric tumours, with a median of 0.35 non-silent mutations per megabase. We identified twelve genes mutated at statistically significant frequencies, including previously known mutated genes in medulloblastoma such as CTNNB1, PTCH1, MLL2, SMARCA4 and TP53. Recurrent somatic mutations were newly identified in an RNA helicase gene, DDX3X, often concurrent with CTNNB1 mutations, and in the nuclear co-repressor (N-CoR) complex genes GPS2, BCOR and LDB1. We show that mutant DDX3X potentiates transactivation of a TCF promoter and enhances cell viability in combination with mutant, but not wild-type, β-catenin. Together, our study reveals the alteration of WNT, hedgehog, histone methyltransferase and now N-CoR pathways across medulloblastomas and within specific subtypes of this disease, and nominates the RNA helicase DDX3X as a component of pathogenic β-catenin signalling in medulloblastoma.
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http://dx.doi.org/10.1038/nature11329DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC3413789PMC
August 2012