The Generation and Analysis of Tritium-substituted Methane

An unavoidable category of molecular species in large-scale tritium applications, such as nuclear fusion, are tritium-substituted hydrocarbons; these form by radiochemical reactions in the presence of (circulating) tritium and carbon (mainly from the steel of vessels and tubing). Tritiumsubstituted methane species, CQ$_4$ (with Q = H , D , T), are often the precursor for higher-order reaction chains, and thus are of particular interest. Here we describe the controlled production of CQ$_4$ carried out in the CAPER facility of the Tritium Laboratory Karlsruhe (TLK), exploiting catalytic reactions and species-enrichment via the CAPER-integral permeator. CQ4 was generated in substantial quantity (>1000 cm$^3$ at ~850 mbar, with CQ$_4$ - content of up to ~20 %). These samples were analyzed using laser Raman and mass spectrometry, to determine the relative isotopologue composition and to trace the generation of tritiated chain-hydrocarbons.Comment: 10 pages, 4 figures. This article has been accepted for publication in Fusion Science and Technology, published by Taylor & Franci

The Long Non-coding RNA Flatr Anticipates Foxp3 Expression in Regulatory T Cells

Mammalian genomes encode a plethora of long non-coding RNA (lncRNA). These transcripts are thought to regulate gene expression, influencing biological processes from development to pathology. Results from the few lncRNA that have been studied in the context of the immune system have highlighted potentially critical functions as network regulators. Here we explored the nature of the lncRNA transcriptome in regulatory T cells (Tregs), a subset of CD4+ T cells required to establish and maintain immunological self-tolerance. The identified Treg lncRNA transcriptome showed distinct differences from that of non-regulatory CD4+ T cells, with evidence of direct shaping of the lncRNA transcriptome by Foxp3, the master transcription factor driving the distinct mRNA profile of Tregs. Treg lncRNA changes were disproportionally reversed in the absence of Foxp3, with an enrichment for colocalisation with Foxp3 DNA binding sites, indicating a direct coordination of transcription by Foxp3 independent of the mRNA coordination function. We further identified a novel lncRNA Flatr, as a member of the core Treg lncRNA transcriptome. Flatr expression anticipates Foxp3 expression during in vitro Treg conversion, and Flatr-deficient mice show a mild delay in in vitro and peripheral Treg induction. These results implicate Flatr as part of the upstream cascade leading to Treg conversion, and may provide clues as to the nature of this process

First observation of tritium adsorption on graphene

In this work, we report on the first-ever studies of graphene exposed to tritium gas in a controlled environment. The single layer graphene on $\textrm{SiO}_2$/Si substrate was exposed to 400 mbar of $\textrm{T}_2$ for a total time of ~55 h. The resistivity of the graphene sample was measured in-situ during tritium exposure using the Van der Pauw method. After the exposure, the samples were scanned with a confocal Raman microscope to study the effect of tritium on the graphene structure as well as the homogeneity of spectral modifications. We found that the sheet resistance increases by three orders of magnitude during the exposure. By Raman microscopy, we demonstrate that the graphene film can withstand the bombardment from the beta-decay of tritium, and primary and secondary ions. Additionally, the Raman spectra after tritium exposure are comparable with previously observed results in hydrogen-loading experiments carried out by other groups. By thermal annealing we could demonstrate, using Raman spectral analysis, that the structural changes were partially reversible. Considering all observations, we conclude that the graphene film was at least partially tritiated during the tritium exposure.Comment: Submitted to Nanoscale Advances (RSC), 14 pages, 4 figure

A comprehensive inventory of TLX1 controlled long non-coding RNAs in T-cell acute lymphoblastic leukemia through polyA+ and total RNA sequencing

Graft-versus-host disease (GvHD) assessment has been shown to be a challenge for healthcare professionals, leading to the development of the eGVHD App (www.uzleuven.be/egvhd). In this study, we formally evaluated the accuracy of using the App compared to traditional assessment methods to assess GvHD. Our national multicenter randomized controlled trial involved seven Belgian transplantation centers and 78 healthcare professionals selected using a 2-stage convenience sampling approach between January and April 2017. Using a 1:1 randomization stratified by profession, healthcare professionals were assigned to use either the App ("APP") or their usual GvHD assessment aids ("No APP") to assess the diagnosis and severity score of 10 expert-validated clinical vignettes. Our main outcome measure was the difference in accuracy for GvHD severity scoring between both groups. The odds of being correct were 6.14 (95% CI: 2.83-13.34) and 6.29 (95% CI: 4.32-9.15) times higher in favor of the "APP" group for diagnosis and scoring, respectively (P<0.001). Appassisted GvHD severity scoring was significantly superior for both acute and chronic GvHD, with an Odds Ratio of 17.89 and 4.34 respectively (P<0.001) and showed a significantly increased inter-observer agreement compared to standard practice. Despite a mean increase of 24 minutes (95% CI: 20.45-26.97) in the time needed to score the whole GvHD test package in the "APP" group (P<0.001), usability feedback was positive. The eGVHD App shows superior GvHD assessment accuracy compared to standard practice and has the potential to improve the quality of outcome data registration in allogeneic stem cell transplantation

Patient-specific cancer genes contribute to recurrently perturbed pathways and establish therapeutic vulnerabilities in esophageal adenocarcinoma

Abstract: The identification of cancer-promoting genetic alterations is challenging particularly in highly unstable and heterogeneous cancers, such as esophageal adenocarcinoma (EAC). Here we describe a machine learning algorithm to identify cancer genes in individual patients considering all types of damaging alterations simultaneously. Analysing 261 EACs from the OCCAMS Consortium, we discover helper genes that, alongside well-known drivers, promote cancer. We confirm the robustness of our approach in 107 additional EACs. Unlike recurrent alterations of known drivers, these cancer helper genes are rare or patient-specific. However, they converge towards perturbations of well-known cancer processes. Recurrence of the same process perturbations, rather than individual genes, divides EACs into six clusters differing in their molecular and clinical features. Experimentally mimicking the alterations of predicted helper genes in cancer and pre-cancer cells validates their contribution to disease progression, while reverting their alterations reveals EAC acquired dependencies that can be exploited in therapy

Mutational signatures in esophageal adenocarcinoma define etiologically distinct subgroups with therapeutic relevance.

Esophageal adenocarcinoma (EAC) has a poor outcome, and targeted therapy trials have thus far been disappointing owing to a lack of robust stratification methods. Whole-genome sequencing (WGS) analysis of 129 cases demonstrated that this is a heterogeneous cancer dominated by copy number alterations with frequent large-scale rearrangements. Co-amplification of receptor tyrosine kinases (RTKs) and/or downstream mitogenic activation is almost ubiquitous; thus tailored combination RTK inhibitor (RTKi) therapy might be required, as we demonstrate in vitro. However, mutational signatures showed three distinct molecular subtypes with potential therapeutic relevance, which we verified in an independent cohort (n = 87): (i) enrichment for BRCA signature with prevalent defects in the homologous recombination pathway; (ii) dominant T>G mutational pattern associated with a high mutational load and neoantigen burden; and (iii) C>A/T mutational pattern with evidence of an aging imprint. These subtypes could be ascertained using a clinically applicable sequencing strategy (low coverage) as a basis for therapy selection.Whole-genome sequencing of esophageal adenocarcinoma samples was performed as part of the International Cancer Genome Consortium (ICGC) through the oEsophageal Cancer Clinical and Molecular Stratification (OCCAMS) Consortium and was funded by Cancer Research UK. We thank the ICGC members for their input on verification standards as part of the benchmarking exercise. We thank the Human Research Tissue Bank, which is supported by the National Institute for Health Research (NIHR) Cambridge Biomedical Research Centre, from Addenbrooke’s Hospital and UCL. Also the University Hospital of Southampton Trust and the Southampton, Birmingham, Edinburgh and UCL Experimental Cancer Medicine Centres and the QEHB charities. This study was partly funded by a project grant from Cancer Research UK. R.C.F. is funded by an NIHR Professorship and receives core funding from the Medical Research Council and infrastructure support from the Biomedical Research Centre and the Experimental Cancer Medicine Centre. We acknowledge the support of The University of Cambridge, Cancer Research UK (C14303/A17197) and Hutchison Whampoa Limited. We would like to thank Dr. Peter Van Loo for providing the NGS version of ASCAT for copy number calling. We are grateful to all the patients who provided written consent for participation in this study and the staff at all participating centres. Some of the work was undertaken at UCLH/UCL who received a proportion of funding from the Department of Health’s NIHR Biomedical Research Centres funding scheme. The work at UCLH/UCL was also supported by the CRUK UCL Early Cancer Medicine Centre.This is the author accepted manuscript. The final version is available from Nature Publishing Group via http://dx.doi.org/10.1038/ng.365

A comparative analysis of whole genome sequencing of esophageal adenocarcinoma pre- and post-chemotherapy

The scientific community has avoided using tissue samples from patients that have been exposed to systemic chemotherapy to infer the genomic landscape of a given cancer. Esophageal adenocarcinoma is a heterogeneous, chemoresistant tumor for which the availability and size of pretreatment endoscopic samples are limiting. This study compares whole-genome sequencing data obtained from chemo-naive and chemo-treated samples. The quality of whole-genomic sequencing data is comparable across all samples regardless of chemotherapy status. Inclusion of samples collected post-chemotherapy increased the proportion of late-stage tumors. When comparing matched pre- and post-chemotherapy samples from 10 cases, the mutational signatures, copy number, and SNV mutational profiles reflect the expected heterogeneity in this disease. Analysis of SNVs in relation to allele-specific copy-number changes pinpoints the common ancestor to a point prior to chemotherapy. For cases in which pre- and post-chemotherapy samples do show substantial differences, the timing of the divergence is near-synchronous with endoreduplication. Comparison across a large prospective cohort (62 treatment-naive, 58 chemotherapy-treated samples) reveals no significant differences in the overall mutation rate, mutation signatures, specific recurrent point mutations, or copy-number events in respect to chemotherapy status. In conclusion, whole-genome sequencing of samples obtained following neoadjuvant chemotherapy is representative of the genomic landscape of esophageal adenocarcinoma. Excluding these samples reduces the material available for cataloging and introduces a bias toward the earlier stages of cancer.This study was partly funded by a project grant from Cancer Research UK. R.C.F. is funded by an NIHR Professorship and receives core funding from the Medical Research Council and infrastructure support from the Biomedical Research Centre and the Experimental Cancer Medicine Centre. We acknowledge the support of The University of Cambridge, Cancer Research UK (C14303/A17197) and Hutchison Whampoa Limited

Oncogenic cooperation in T-cell acute lymphoblastic leukemia

T-cell acute lymphoblastic leukemia (T-ALL) is an aggressive hematologic malignancy, characterized by high white blood cell counts and infiltration of immature T cells into the bone marrow and other tissues. The management of T-ALL involves chemotherapy treatment, which is not specifically targeting leukemic cells. Thus, current treatments are associated with severe side effects for T-ALL patients. Gaining more insight into the mechanism of T-ALL development will be critical for the development of new targeted therapeutic agents, tailored to specific mutations present in the leukemia cells. Recent clinical sequencing studies showed that T-ALL is caused by the accumulation of multiple mutations combined with the ectopic expression of transcription factors in developing T cells. However, dissecting the molecular mechanism of cooperation between transcription factor expression and additional oncogenic mutations in driving T-ALL has been difficult to assess due to limited robust T cell model systems. Here we utilize a new ex vivo pro-T cell model to study oncogenic cooperation. Using a systems biological approach we first dissect the pro-T cell signaling network driven by interleukin-7 (IL-7), stem cell factor (Scf) and Notch1 and identify key downstream Akt, Stat, E2f and Myc genetic signaling networks. Next, this pro-T cell system was used to demonstrate that ectopic expression of the TAL1 transcription factor and Pten deletion are bona-fide cooperating events resulting in an increased stem-cell signature, upregulation of a specific E2f signaling network and metabolic reprogramming with higher influx of glucose carbons into the TCA cycle. As a next step, we aimed to assess the potential of IL7R mutations to transform pro-T cells to IL-7 independent growth. We demonstrated that IL7R mutations are weak activators of the Stat5 signaling pathway and only cysteine inserting mutations in the IL7R lead to cell transformation and IL-7 independency. However, aberrant expression of TLX1, which frequently co-occurs with IL7R mutations in T-ALL patients, lowered the threshold requirement for phospho-Stat5. TLX1 bound directly to Stat5 target genes and induced a similar geneset as Stat5. This ex vivo pro-T cell system thereby provides a powerful new model system to investigate how normal T cell signaling networks are perturbed and/or hijacked by different oncogenic events found in T-ALL.nrpages: 164status: publishe