Diffuse glioneuronal tumour with oligodendroglioma-like features and nuclear clusters (DGONC) - a molecularly defined glioneuronal CNS tumour class displaying recurrent monosomy 14.

AIMS: DNA methylation-based central nervous system (CNS) tumour classification has identified numerous molecularly distinct tumour types, and clinically relevant subgroups among known CNS tumour entities that were previously thought to represent homogeneous diseases. Our study aimed at characterizing a novel, molecularly defined variant of glioneuronal CNS tumour. PATIENTS AND METHODS: DNA methylation profiling was performed using the Infinium MethylationEPIC or 450 k BeadChip arrays (Illumina) and analysed using the 'conumee' package in R computing environment. Additional gene panel sequencing was also performed. Tumour samples were collected at the German Cancer Research Centre (DKFZ) and provided by multinational collaborators. Histological sections were also collected and independently reviewed. RESULTS: Genome-wide DNA methylation data from >25 000 CNS tumours were screened for clusters separated from established DNA methylation classes, revealing a novel group comprising 31 tumours, mainly found in paediatric patients. This DNA methylation-defined variant of low-grade CNS tumours with glioneuronal differentiation displays recurrent monosomy 14, nuclear clusters within a morphology that is otherwise reminiscent of oligodendroglioma and other established entities with clear cell histology, and a lack of genetic alterations commonly observed in other (paediatric) glioneuronal entities. CONCLUSIONS: DNA methylation-based tumour classification is an objective method of assessing tumour origins, which may aid in diagnosis, especially for atypical cases. With increasing sample size, methylation analysis allows for the identification of rare, putative new tumour entities, which are currently not recognized by the WHO classification. Our study revealed the existence of a DNA methylation-defined class of low-grade glioneuronal tumours with recurrent monosomy 14, oligodendroglioma-like features and nuclear clusters.

Epigenomic alterations define lethal CIMP-positive ependymomas of infancy.

Ependymomas are common childhood brain tumours that occur throughout the nervous system, but are most common in the paediatric hindbrain. Current standard therapy comprises surgery and radiation, but not cytotoxic chemotherapy as it does not further increase survival. Whole-genome and whole-exome sequencing of 47 hindbrain ependymomas reveals an extremely low mutation rate, and zero significant recurrent somatic single nucleotide variants. Although devoid of recurrent single nucleotide variants and focal copy number aberrations, poor-prognosis hindbrain ependymomas exhibit a CpG island methylator phenotype. Transcriptional silencing driven by CpG methylation converges exclusively on targets of the Polycomb repressive complex 2 which represses expression of differentiation genes through trimethylation of H3K27. CpG island methylator phenotype-positive hindbrain ependymomas are responsive to clinical drugs that target either DNA or H3K27 methylation both in vitro and in vivo. We conclude that epigenetic modifiers are the first rational therapeutic candidates for this deadly malignancy, which is epigenetically deregulated but genetically bland.

Integrated molecular characterization of IDH-mutant glioblastomas.

AIMS: Mutations of isocitrate dehydrogenase (IDH)1/2 affect almost all astrocytomas of WHO grade II and III. A subset of IDH-mutant astrocytic tumours progresses to IDH-mutant glioblastoma or presents with the histology of a glioblastoma at first presentation. We set out here to assess the molecular spectrum of IDH-mutant glioblastomas. METHODS: We performed an integrated molecular analysis of a mono-centric cohort (n = 97); assessed through genome-wide DNA methylation analysis, copy-number profiling and targeted next generation sequencing using a neurooncology-tailored gene panel. RESULTS: Of these 97 IDH-mutant glioblastomas, 68 had a glioblastoma at first presentation ('de novo' IDH-mutant glioblastoma) and 29 emerged from a prior low-grade lesion ('evolved' IDH-mutant glioblastoma). Unsupervised hierarchical clustering of DNA methylation data disclosed that IDH-mutant glioblastoma ('de novo' and 'evolved') formed a distinct group separate from other diffuse glioma subtypes. Homozygous deletions of CDKN2A/B were found to be associated with shorter survival. CONCLUSIONS: This study demonstrates DNA methylation patterns in IDH-mutant glioblastoma to be distinct from lower-grade astrocytic counterparts but homogeneous within de novo and evolved IDH-mutant glioblastomas, and identifies CDKN2A as a marker for possible genetic sub-stratification.

Bevacizumab plus hypofractionated radiotherapy versus radiotherapy alone in elderly patients with glioblastoma: the randomized, open-label, phase II ARTE trial.

Background: The addition of bevacizumab to temozolomide-based chemoradiotherapy (TMZ/RT â†’ TMZ) did not prolong overall survival (OS) in patients with newly diagnosed glioblastoma in phase III trials. Elderly and frail patients are underrepresented in clinical trials, but early reports suggested preferential benefit in this population. Patients and methods: ARTE was a 2 : 1 randomized, multi-center, open-label, non-comparative phase II trial of hypofractionated RT (40 Gy in 15 fractions) with bevacizumab (10 mg/kg×14 days) (arm A, N = 50) or without bevacizumab (arm B, N = 25) in patients with newly diagnosed glioblastoma aged ≥65 years. The primary objective was to obtain evidence for prolongation of median OS by the addition of bevacizumab to RT. Response was assessed by RANO criteria. Quality of life (QoL) was monitored by the EORTC QLQ-C30/BN20 modules. Exploratory studies included molecular subtyping by 450k whole methylome and gene expression analyses. Results: Median PFS was longer in arm A than in arm B (7.6 and 4.8 months, P = 0.003), but OS was similar (12.1 and 12.2 months, P = 0.77). Clinical deterioration was delayed and more patients came off steroids in arm A. Prolonged PFS in arm A was confined to tumors with the receptor tyrosine kinase (RTK) I methylation subtype (HR 0.25, P = 0.014) and proneural gene expression (HR 0.29, P = 0.025). In a Cox model of OS controlling for established prognostic factors, associations with more favorable outcome were identified for age <70 years (HR 0.52, P = 0.018) and Karnofsky performance score 90%-100% (HR 0.51, P = 0.026). Including molecular subtypes into that model identified an association of the RTK II gene methylation subtype with inferior OS (HR 1.73, P = 0.076). Conclusion: Efficacy outcomes and exploratory analyses of ARTE do not support the hypothesis that the addition of bevacizumab to RT generally prolongs survival in elderly glioblastoma patients. Molecular biomarkers may identify patients with preferential benefit from bevacizumab. Clinical trial registration number: NCT01443676.

A taxonomic index, with names of descriptive authorities of termite genera and species: an accompaniment to Biology of Termites: A Modern Synthesis (Bignell DE, Roisin Y, Lo N, Editors. 2011. Springer, Dordrecht. 576 pp.).

Biology of Termites: A Modern Synthesis (Bignell DE, Roisin Y, Lo N, (Editors), Springer, Dordrecht, 576pp, ISBN 978-90-481-3976-7, e-ISBN 978-90-481-3977-4, DOI 10.1007/978-90-481-3977-4) was published in 2011. With the agreement of the publishers, we give a taxonomic index of the book comprising 494 termite entries, 103 entries of other multicellular animal species mentioned as associates or predators of termites, with 9 fungal, 60 protist, and 64 prokaryote identities, which are listed as termite symbionts (sensu stricto). In addition, we add descriptive authorities for living (and some fossil) termite genera and species. Higher taxonomic groupings for termites are indicated by 25 code numbers. Microorganisms (prokaryotes, protists, and fungi) are listed separately, using broad modern taxonomic affiliations from the contemporary literature of bacteriology, protozoology, and mycology.

Protein annotation and modelling servers at University College London.

The UCL Bioinformatics Group web portal offers several high quality protein structure prediction and function annotation algorithms including PSIPRED, pGenTHREADER, pDomTHREADER, MEMSAT, MetSite, DISOPRED2, DomPred and FFPred for the prediction of secondary structure, protein fold, protein structural domain, transmembrane helix topology, metal binding sites, regions of protein disorder, protein domain boundaries and protein function, respectively. We also now offer a fully automated 3D modelling pipeline: BioSerf, which performed well in CASP8 and uses a fragment-assembly approach which placed it in the top five servers in the de novo modelling category. The servers are available via the group web site at http://bioinf.cs.ucl.ac.uk/.

Changes in Ventricular and Cortical Volumes following Shunt Placement in Patients with Idiopathic Normal Pressure Hydrocephalus.

BACKGROUND AND PURPOSE: While changes in ventricular and extraventricular CSF spaces have been studied following shunt placement in patients with idiopathic normal pressure hydrocephalus, regional changes in cortical volumes have not. These changes are important to better inform disease pathophysiology and evaluation for copathology. The purpose of this work is to investigate changes in ventricular and cortical volumes in patients with idiopathic normal pressure hydrocephalus following ventriculoperitoneal shunt placement. MATERIALS AND METHODS: This is a retrospective cohort study of patients with idiopathic normal pressure hydrocephalus who underwent 3D T1-weighted MR imaging before and after ventriculoperitoneal shunt placement. Images were analyzed using tensor-based morphometry with symmetric normalization to determine the percentage change in ventricular and regional cortical volumes. Ventricular volume changes were assessed using the Wilcoxon signed rank test, and cortical volume changes, using a linear mixed-effects model (P < .05). RESULTS: The study included 22 patients (5 women/17 men; mean age, 73 [SD, 6] years). Ventricular volume decreased after shunt placement with a mean change of -15.4% (P < .001). Measured cortical volume across all participants and cortical ROIs showed a mean percentage increase of 1.4% (P < .001). ROIs near the vertex showed the greatest percentage increase in volume after shunt placement, with smaller decreases in volume in the medial temporal lobes. CONCLUSIONS: Overall, cortical volumes mildly increased after shunt placement in patients with idiopathic normal pressure hydrocephalus with the greatest increases in regions near the vertex, indicating postshunt decompression of the cortex and sulci. Ventricular volumes showed an expected decrease after shunt placement.

FFPred: an integrated feature-based function prediction server for vertebrate proteomes.

One of the challenges of the post-genomic era is to provide accurate function annotations for large volumes of data resulting from genome sequencing projects. Most function prediction servers utilize methods that transfer existing database annotations between orthologous sequences. In contrast, there are few methods that are independent of homology and can annotate distant and orphan protein sequences. The FFPred server adopts a machine-learning approach to perform function prediction in protein feature space using feature characteristics predicted from amino acid sequence. The features are scanned against a library of support vector machines representing over 300 Gene Ontology (GO) classes and probabilistic confidence scores returned for each annotation term. The GO term library has been modelled on human protein annotations; however, benchmark performance testing showed robust performance across higher eukaryotes. FFPred offers important advantages over traditional function prediction servers in its ability to annotate distant homologues and orphan protein sequences, and achieves greater coverage and classification accuracy than other feature-based prediction servers. A user may upload an amino acid and receive annotation predictions via email. Feature information is provided as easy to interpret graphics displayed on the sequence of interest, allowing for back-interpretation of the associations between features and function classes.

Real-time quantitative polymerase chain reaction (qPCR) analysis with fluorescence resonance energy transfer (FRET) probes reveals differential expression of the four ERBB4 juxtamembrane region variants between medulloblastoma and pilocytic astrocytoma.

AIMS: We report a comparative study on the mRNA expression of ErbB receptor tyrosine kinases, and in particular ERBB4 transcript variants, in two common paediatric brain tumours: medulloblastoma (MB) and pilocytic astrocytoma (PA). METHODS: While the conventional real-time quantitative polymerase chain reaction was used to measure the expression of ERRBs and ErbB4-processing protease genes, the LightCycler fluorescence resonance energy transfer probes were specifically designed to investigate all of the known ERBB4 juxtamembrane (JM) and cytoplasmic transcript variants. RESULTS: The overall expression of ERBBs suggests that ErbB2/ErbB4 heterodimers and ErbB4 homodimers may be major functional units of the ErbBs in MB, while ErbB2/ErbB3 heterodimers may play a more prominent role in addition to ErbB4-containing dimers in PA. Different expression patterns of ERBB4 JM transcripts in MB, PA and normal brain were observed. The JM-d variant was only detected in MBs, while JM-c was present in MB and PA but was not identified in normal brain. The expression of cleavable ERBB4 transcript variants was elevated in PAs and MBs compared with normal brain, while mRNA levels of ErbB4-processing proteases were similar in both tumour types and normal brain. This suggests that proteolytic cleavage of ErbB4 may be more common in MB and PA, which leads to signalling events divergent from those in normal brain. CONCLUSION: Taken together, these results suggest that ErbB4 processing and function may be altered in brain tumours, such as MB and PA, via differential expression of JM transcript variants.

Golf: an olfactory neuron specific-G protein involved in odorant signal transduction.

Biochemical and electrophysiological studies suggest that odorants induce responses in olfactory sensory neurons via an adenylate cyclase cascade mediated by a G protein. An olfactory-specific guanosine triphosphate (GTP)-binding protein alpha subunit has now been characterized and evidence is presented suggesting that this G protein, termed Golf, mediates olfaction. Messenger RNA that encodes Golf alpha is expressed in olfactory neuroephithelium but not in six other tissues tested. Moreover, within the olfactory epithelium, Golf alpha appears to be expressed only by the sensory neurons. Specific antisera were used to localize Golf alpha protein to the sensory apparatus of the receptor neurons. Golf alpha shares extensive amino acid identity (88 percent) with the stimulatory G protein, Gs alpha. The expression of Golf alpha in S49 cyc- kin- cells, a line deficient in endogenous stimulatory G proteins, demonstrates its capacity to stimulate adenylate cyclase in a heterologous system.

DNA methylation, transcriptome and genetic copy number signatures of diffuse cerebral WHO grade II/III gliomas resolve cancer heterogeneity and development.

BACKGROUND: Diffuse lower WHO grade II and III gliomas (LGG) are slowly progressing brain tumors, many of which eventually transform into a more aggressive type. LGG is characterized by widespread genetic and transcriptional heterogeneity, yet little is known about the heterogeneity of the DNA methylome, its function in tumor biology, coupling with the transcriptome and tumor microenvironment and its possible impact for tumor development. METHODS: We here present novel DNA methylation data of an LGG-cohort collected in the German Glioma Network containing about 85% isocitrate dehydrogenase (IDH) mutated tumors and performed a combined bioinformatics analysis using patient-matched genome and transcriptome data. RESULTS: Stratification of LGG based on gene expression and DNA-methylation provided four consensus subtypes. We characterized them in terms of genetic alterations, functional context, cellular composition, tumor microenvironment and their possible impact for treatment resistance and prognosis. Glioma with astrocytoma-resembling phenotypes constitute the largest fraction of nearly 60%. They revealed largest diversity and were divided into four expression and three methylation groups which only partly match each other thus reflecting largely decoupled expression and methylation patterns. We identified a novel G-protein coupled receptor and a cancer-related 'keratinization' methylation signature in in addition to the glioma-CpG island methylator phenotype (G-CIMP) signature. These different signatures overlap and combine in various ways giving rise to diverse methylation and expression patterns that shape the glioma phenotypes. The decrease of global methylation in astrocytoma-like LGG associates with higher WHO grade, age at diagnosis and inferior prognosis. We found analogies between astrocytoma-like LGG with grade IV IDH-wild type tumors regarding possible worsening of treatment resistance along a proneural-to-mesenchymal axis. Using gene signature-based inference we elucidated the impact of cellular composition of the tumors including immune cell bystanders such as macrophages. CONCLUSIONS: Genomic, epigenomic and transcriptomic factors act in concert but partly also in a decoupled fashion what underpins the need for integrative, multidimensional stratification of LGG by combining these data on gene and cellular levels to delineate mechanisms of gene (de-)regulation and to enable better patient stratification and individualization of treatment.

Pseudomonas aeruginosa transmission is infrequent in New Zealand cystic fibrosis clinics.

Pseudomonas aeruginosa is an important pathogen in cystic fibrosis (CF). Although most patients harbour unique P. aeruginosa isolates, some clinics report patients sharing common strains. The overall importance of person-to-person transmission in P. aeruginosa acquisition and whether routine patient segregation is necessary remains uncertain. The present authors therefore investigated the extent of P. aeruginosa transmission in New Zealand CF clinics. New Zealand's seven major CF centres were assessed, combining epidemiological data with computer-assisted SalI DNA fingerprinting of 496 isolates from 102 patients. One cluster of related isolates was significantly more prevalent in the largest clinic than expected by chance. The seven patients with isolates belonging to this cluster had more contact with each other than the remaining patients attending this centre. No other convincing evidence of transmission was found in any of the other smaller clinics. Three P. aeruginosa strains believed to be transmissible between patients in Australian and British CF clinics are present in New Zealand, but there was no definite evidence they had spread. Pseudomonas aeruginosa transmission is currently infrequent in New Zealand cystic fibrosis clinics. This situation could change rapidly and ongoing surveillance is required. The current results confirm that computer-assisted SalI DNA fingerprinting is ideally suited for such surveillance.

Benchmarking template selection and model quality assessment for high-resolution comparative modeling.

Comparative modeling is presently the most accurate method of protein structure prediction. Previous experiments have shown the selection of the correct template to be of paramount importance to the quality of the final model. We have derived a set of 732 targets for which a choice of ten or more templates exist with 30-80% sequence identity and used this set to compare a number of possible methods for template selection: BLAST, PSI-BLAST, profile-profile alignment, HHpred HMM-HMM comparison, global sequence alignment, and the use of a model quality assessment program (MQAP). In addition, we have investigated the question of whether any structurally defined subset of the sequence could be used to predict template quality better than overall sequence similarity. We find that template selection by BLAST is sufficient in 75% of cases but that there are examples in which improvement (global RMSD 0.5 A or more) could be made. No significant improvement is found for any of the more sophisticated sequence-based methods of template selection at high sequence identities. A subset of 118 targets extending to the lowest levels of sequence similarity was examined and the HHpred and MQAP methods were found to improve ranking when available templates had 35-40% maximum sequence identity. Structurally defined subsets in general are found to be less discriminative than overall sequence similarity, with the coil residue subset performing equivalently to sequence similarity. Finally, we demonstrate that if models are built and model quality is assessed in combination with the sequence-template sequence similarity that a extra 7% of "best" models can be found.

Recurring structural motifs in proteins with different functions.

BACKGROUND: Structures that have diverged from a common ancestor often retain functional and sequence similarity, although the latter may be very reduced. Even so, the overall fold of the structure is generally highly conserved. Now however, several have been identified of proteins that have been identified that have different functions but which have converged to a similar fold. These proteins will also have low sequence identities. RESULTS: By comparing the complete structure databank against itself, using sequence and structure alignment techniques, we have been able to identify six new examples of structurally related folds that have no apparent sequence or functional similarity. These related proteins include a family of crambin-like folds and a family of ferredoxin II folds. We found that all the similarities between structures are present in small proteins and occur as motifs within the core of a larger protein. CONCLUSION: The low sequence similarity and the lack of any obvious functional relationship between proteins with similar structures suggest that the proteins have diverged from independent ancestors. The similarities may therefore be of interest for understanding the various stereochemical and physical criteria that operate to generate a favourable fold.

Progress in protein structure prediction.

If protein structure prediction methods are to make any impact on the impending onerous task of analyzing the large numbers of unknown protein sequences generated by the ongoing genome-sequencing projects, it is vital that they make the difficult transition from computational 'gedankenexperiments' to practical software tools. This has already happened in the field of comparative modelling and is currently happening in the threading field. Unfortunately, there is little evidence of this transition happening in the field of ab initio tertiary-structure prediction.

Protein structure prediction in the postgenomic era.

As the number of completely sequenced genomes rapidly increases, the postgenomic problem of gene function identification becomes ever more pressing. Predicting the structures of proteins encoded by genes of interest is one possible means to glean subtle clues as to the functions of these proteins. There are limitations to this approach to gene identification and a survey of the expected reliability of different protein structure prediction techniques has been undertaken.

Potential energy functions for threading.

Despite little progress in ab initio solutions to the problem of predicting a protein's tertiary structure, over the past four years or so the development of fold-recognition methods for tertiary structure prediction has been the source of some encouragement in this difficult field. Despite promising initial results, these methods are clearly not yet fully mature and many groups are now working on different aspects of the methods involved in the hope of increasing the reliability and sensitivity of these tools.

Beta propellers: structural rigidity and functional diversity.

Recently solved structures and proposed models have helped to reveal the structural characteristics of the beta-propeller fold, as well as the features that contribute to its high rigidity and stability. Possible strategies for identifying beta-propeller proteins in newly characterised sequences are helping to overcome the problems of predicting the beta-propeller fold from amino acid sequences.

The sesquiterpene lactone parthenolide induces selective apoptosis of B-chronic lymphocytic leukemia cells in vitro.

We have studied the in vitro actions of the sesquiterpene lactone parthenolide (PTL) on cells isolated from patients with chronic lymphocytic leukemia (CLL). Dye reduction viability assays showed that the median LD(50) for PTL was 6.2 muM (n=78). Fifteen of these isolates were relatively resistant to the conventional agent chlorambucil but retained sensitivity to PTL. Brief exposures to PTL (1-3 h) were sufficient to induce caspase activation and commitment to cell death. Chronic lymphocytic leukemia cells were more sensitive towards PTL than were normal T lymphocytes or CD34(+) haematopoietic progenitor cells. The mechanism of cell killing was via PTL-induced generation of reactive oxygen species, resulting in turn in a proapoptotic Bax conformational change, release of mitochondrial cytochrome c and caspase activation. Parthenolide also decreased nuclear levels of the antiapoptotic transcription factor nuclear factor-kappa B and diminished phosphorylation of its negative regulator IkappaB. Killing of CLL cells by PTL was apparently independent of p53 induction. This is the first report showing the relative selectivity of PTL towards CLL cells. The data here warrant further investigation of this class of natural product as potential therapeutic agents for CLL.

A compact high-energy neutron spectrometer.

A compact liquid organic neutron spectrometer based on a single NE213 liquid scintillator (5 cm diameter x 5 cm) is described. The spectrometer is designed to measure neutron fluence spectra over the energy range 2-200 MeV and is suitable for use in neutron fields having any type of time structure. Neutron fluence spectra are obtained from measurements of two-parameter distributions (counts versus pulse-height and pulse shape) using the Bayesian unfolding code MAXED. Calibration and test measurements made using a pulsed neutron beam with a continuous energy spectrum are described and the application of the spectrometer to radiation dose measurements is discussed.