With the number of prognostic and predictive genetic markers in neuro-oncology steadily growing, the need for comprehensive molecular analysis of neuropathology samples has vastly increased. We ...therefore developed a customized enrichment/hybrid-capture-based next-generation sequencing (NGS) gene panel comprising the entire coding and selected intronic and promoter regions of 130 genes recurrently altered in brain tumors, allowing for the detection of single nucleotide variations, fusions, and copy number aberrations. Optimization of probe design, library generation and sequencing conditions on 150 samples resulted in a 5-workday routine workflow from the formalin-fixed paraffin-embedded sample to neuropathological report. This protocol was applied to 79 retrospective cases with established molecular aberrations for validation and 71 prospective cases for discovery of potential therapeutic targets. Concordance of NGS compared to established, single biomarker methods was 98.0 %, with discrepancies resulting from one case where a
TERT
promoter mutation was not called by NGS and three ATRX mutations not being detected by Sanger sequencing. Importantly, in samples with low tumor cell content, NGS was able to identify mutant alleles that were not detectable by traditional methods. Information derived from NGS data identified potential targets for experimental therapy in 37/47 (79 %) glioblastomas, 9/10 (90 %) pilocytic astrocytomas, and 5/14 (36 %) medulloblastomas in the prospective target discovery cohort. In conclusion, we present the settings for high-throughput, adaptive next-generation sequencing in routine neuropathology diagnostics. Such an approach will likely become highly valuable in the near future for treatment decision making, as more therapeutic targets emerge and genetic information enters the classification of brain tumors.
Challenges to curing primary brain tumours Aldape, Kenneth; Brindle, Kevin M; Chesler, Louis ...
Nature reviews. Clinical oncology,
08/2019, Letnik:
16, Številka:
8
Journal Article
Recenzirano
Odprti dostop
Despite decades of research, brain tumours remain among the deadliest of all forms of cancer. The ability of these tumours to resist almost all conventional and novel treatments relates, in part, to ...the unique cell-intrinsic and microenvironmental properties of neural tissues. In an attempt to encourage progress in our understanding and ability to successfully treat patients with brain tumours, Cancer Research UK convened an international panel of clinicians and laboratory-based scientists to identify challenges that must be overcome if we are to cure all patients with a brain tumour. The seven key challenges summarized in this Position Paper are intended to serve as foci for future research and investment.
Ependymoma with RELA fusion has been defined as a novel entity of the revised World Health Organization 2016 classification of tumors of the central nervous system (CNS), characterized by fusion ...transcripts of the RELA gene and consequent pathological activation of the NFkB pathway. These tumors represent the majority of supratentorial ependymomas in children. The validation of diagnostic tools to identify this clinically relevant ependymoma entity is essential. Here, we have used interphase fluorescent in situ hybridization (FISH) for C11orf95 and RELA, immunohistochemistry (IHC) for p65‐RelA and the recently developed DNA methylation‐based classification besides conventional histopathology, and compared the precision of the methods in 40 supratentorial pediatric brain tumors diagnosed as ependymomas in the past years. Reverse transcription PCR (RT‐PCR) and RNA sequencing were performed to explore discordant cases. Furthermore, we integrated imaging and clinical features as additional layers of information. The concordance between nuclear RelA expression by IHC and RELA FISH was 100%. Concordance between IHC and DNA methylation profiling, and between FISH and DNA methylation profiling was also high (96.4% and 95.2%, respectively). Thirty‐four out of 40 (85%) cases were confirmed by integrated diagnoses as ependymal tumors, including 22 RELA‐fused ependymomas (71% of ependymal tumors), two YAP1‐fused ependymomas (6%), six non‐RELA/non‐YAP1 ependymomas (18%) and four ependymal/subependymal mixed tumors (12%). Ependymal/subependymal mixed tumors had an excellent clinical outcome despite the presence of histopathological signs of malignancy, suggesting that these tumors should not be diagnosed as classic ependymomas. DNA methylation profiling helped in the differential diagnosis of RELA‐fused ependymomas. IHC and FISH, which are available in the majority of pathology laboratories, are valuable tools to identify RELA‐fused ependymomas.
Astrocytoma and oligodendroglioma are histologically and genetically well-defined entities. The majority of astrocytomas harbor concurrent
TP53
and
ATRX
mutations, while most oligodendrogliomas carry ...the 1p/19q co-deletion. Both entities share high frequencies of
IDH
mutations. In contrast, oligoastrocytomas (OA) appear less clearly defined and, therefore, there is an ongoing debate whether these tumors indeed constitute an entity or whether they represent a mixed bag containing both astrocytomas and oligodendrogliomas. We investigated 43 OA diagnosed in different institutions employing histology, immunohistochemistry and in situ hybridization addressing surrogates for the molecular genetic markers
IDH1
R132H,
TP53
,
ATRX
and 1p/19q loss. In all but one OA the combination of nuclear p53 accumulation and ATRX loss was mutually exclusive with 1p/19q co-deletion. In 31/43 OA, only alterations typical for oligodendroglioma were observed, while in 11/43 OA, only indicators for mutations typical for astrocytomas were detected. A single case exhibited a distinct pattern, nuclear expression of p53, ATRX loss,
IDH1
mutation and partial 1p/19q loss. However, this was the only patient undergoing radiotherapy prior to surgery, possibly contributing to the acquisition of this uncommon combination. In OA with oligodendroglioma typical alterations, the portions corresponding to astrocytic part were determined as reactive, while in OA with astrocytoma typical alterations the portions corresponding to oligodendroglial differentiation were neoplastic. These data provide strong evidence against the existence of an independent OA entity.
EGFR
amplification (
EGFR
amp), the combination of gain of chromosome 7 and loss of chromosome 10 (7+/10−), and
TERT
promoter mutation (p
TERT
mut) are alterations frequently observed in adult
IDH
...-wild-type (
IDH
wt) glioblastoma (GBM). In the absence of endothelial proliferation and/or necrosis, these alterations currently are considered to serve as a surrogate for upgrading
IDH
wt diffuse or anaplastic astrocytoma to GBM. Here, we set out to determine the distribution of
EGFR
amp, 7+/10−, and p
TERT
mut by analyzing high-resolution copy-number profiles and next-generation sequencing data of primary brain tumors. In addition, we addressed the question whether combinations of partial gains on chromosome 7 and partial losses on chromosome 10 exhibited a diagnostic and prognostic value similar to that of complete 7+/10−. Several such combinations proved relevant and were combined as the 7/10 signature. Our results demonstrate that
EGFR
amp and the 7/10 signature are closely associated with
IDH
wt GBM. In contrast, p
TERT
mut is less specific for
IDH
wt GBM. We conclude that, in the absence of endothelial proliferation and/or necrosis, the detection of
EGFR
amp is a very strong surrogate marker for the diagnosis of GBM in
IDH
wt diffuse astrocytic tumors. The 7/10 signature is also a strong surrogate marker. However, care should be taken to exclude pleomorphic xanthoastrocytoma. p
TERT
mut is less restricted to this entity and needs companion analysis by other molecular markers to serve as a surrogate for diagnosing
IDH
wt GBM. A combination of any two of
EGFR
amp, the 7/10 signature and p
TERT
mut, is highly specific for
IDH
wt GBM and the combination of all three alterations is frequent and exclusively seen in
IDH
wt GBM.
Medulloblastoma Northcott, Paul A; Robinson, Giles W; Kratz, Christian P ...
Nature reviews. Disease primers,
02/2019, Letnik:
5, Številka:
1
Journal Article
Recenzirano
Medulloblastoma (MB) comprises a biologically heterogeneous group of embryonal tumours of the cerebellum. Four subgroups of MB have been described (WNT, sonic hedgehog (SHH), Group 3 and Group 4), ...each of which is associated with different genetic alterations, age at onset and prognosis. These subgroups have broadly been incorporated into the WHO classification of central nervous system tumours but still need to be accounted for to appropriately tailor disease risk to therapy intensity and to target therapy to disease biology. In this Primer, the epidemiology (including MB predisposition), molecular pathogenesis and integrative diagnosis taking histomorphology, molecular genetics and imaging into account are reviewed. In addition, management strategies, which encompass surgical resection of the tumour, cranio-spinal irradiation and chemotherapy, are discussed, together with the possibility of focusing more on disease biology and robust molecularly driven patient stratification in future clinical trials.
Abstract
Motivation
As non-coding driver mutations move more into the focus of cancer research, a comprehensive and easy-to-use software solution for regulatory variant analysis and data ...visualization is highly relevant. The interpretation of regulatory variants in large tumor genome cohorts requires specialized analysis and visualization of multiple layers of data, including for example breakpoints of structural variants, enhancer elements and additional available gene locus annotation, in the context of changes in gene expression.
Results
We introduce a user-friendly tool, Revana (REgulatory Variant ANAlysis), that can aggregate and visually represent regulatory variants from cancer genomes in a gene-centric manner. It requires whole-genome and RNA sequencing data of a cohort of tumor samples and creates interactive HTML reports summarizing the most important regulatory events.
Availability and implementation
Revana is implemented in R and JavaScript. It is available for download as an R package under <https://github.com/KiTZ-Heidelberg/revana>. Sample results can be viewed under <https://github.com/KiTZ-Heidelberg/revana-demo-report> and a short walkthrough is available under <https://github.com/KiTZ-Heidelberg/revana-demo-data>.
Supplementary information
Supplementary data are available at Bioinformatics online.
Summary Arguably, nowhere has there been a greater advance in our understanding of biological mechanisms and potential translational targets during the next-generation sequencing era than in ...paediatric brain tumours. The so-called omics revolution, enabled by high-throughput sequencing, has empowered large consortia and independent groups alike to make major genetic discoveries, from dominant-negative histone mutations and hijacking of distal enhancer elements, to new oncogenic gene fusions and aberrantly active gene expression. Epigenetic deregulation has also been revealed as a common theme across several tumour subtypes. This Review focuses on key findings that have been transforming the landscape of paediatric neuro-oncology research and how these results are opening new avenues towards potential therapeutic translation.
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.
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.
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.
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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