Recommendations for whole genome sequencing in diagnostics for rare diseases

In 2016, guidelines for diagnostic Next Generation Sequencing (NGS) have been published by EuroGentest in order to assist laboratories in the implementation and accreditation of NGS in a diagnostic setting. These guidelines mainly focused on Whole Exome Sequencing (WES) and targeted (gene panels) sequencing detecting small germline variants (Single Nucleotide Variants (SNVs) and insertions/deletions (indels)). Since then, Whole Genome Sequencing (WGS) has been increasingly introduced in the diagnosis of rare diseases as WGS allows the simultaneous detection of SNVs, Structural Variants (SVs) and other types of variants such as repeat expansions. The use of WGS in diagnostics warrants the re-evaluation and update of previously published guidelines. This work was jointly initiated by EuroGentest and the Horizon2020 project Solve-RD. Statements from the 2016 guidelines have been reviewed in the context of WGS and updated where necessary. The aim of these recommendations is primarily to list the points to consider for clinical (laboratory) geneticists, bioinformaticians, and (non-)geneticists, to provide technical advice, aid clinical decision-making and the reporting of the results

Myocardial revascularization in octogenarians. Early and late events, survival and functional evaluation.

Item does not contain fulltex

Myocardial revascularization in octogenerians. Functional evaluation by the barthel index.

Item does not contain fulltex

HTS conductor characterization at 27 K

A Ne-liquefaction system and cryostat has been developed and built to characterize the electrical and mechanical properties of HTS conductors at elevated temperatures. The device makes use of a coil type condenser inside the experiment dewar, which is fed by LHe. Ne-gas entering the dewar at room temperature, liquefies on the coil windings and drips to the bottom of the dewar, where the experiment is located. He-gas and Ne-gas are recovered. Approximately 1.5 l of LNe was condensed into the dewar in about 1 h after precooling of the dewar. The device has been used successfully to measure zero-field and in-field transport properties of Bi-2223 and YBCO high-temperature superconductors at 27 K

c-Jun/c-Fos heterodimers regulate cellular genes via a newly identified class of methylated DNA sequence motifs.

CpG methylation in mammalian DNA is known to interfere with gene expression by inhibiting the binding of transactivators to their cognate sequence motifs or recruiting proteins involved in gene repression. An Epstein-Barr virus-encoded transcription factor, Zta, was the first example of a sequence-specific transcription factor that preferentially recognizes and selectively binds DNA sequence motifs with methylated CpG residues, reverses epigenetic silencing and activates gene transcription. The DNA binding domain of Zta is homologous to c-Fos, a member of the cellular AP-1 (activator protein 1) transcription factor family, which regulates cell proliferation and survival, apoptosis, transformation and oncogenesis. We have identified a novel AP-1 binding site termed meAP-1, which contains a CpG dinucleotide. If methylated, meAP-1 sites are preferentially bound by the AP-1 heterodimer c-Jun/c-Fos in vitro and in cellular chromatin in vivo. In activated human primary B cells, c-Jun/c-Fos locates to these methylated elements in promoter regions of transcriptionally activated genes. Reminiscent of the viral Zta protein, c-Jun/c-Fos is the first identified cellular member of the AP-1 family of transactivators that can induce expression of genes with methylated, hence repressed promoters, reversing epigenetic silencing

Identification of recurring tumor-specific somatic mutations in acute myeloid leukemia by transcriptome sequencing.

Genetic lesions are crucial for cancer initiation. Recently, whole genome sequencing, using next generation technology, was used as a systematic approach to identify mutations in genomes of various types of tumors including melanoma, lung and breast cancer, as well as acute myeloid leukemia (AML). Here, we identify tumor-specific somatic mutations by sequencing transcriptionally active genes. Mutations were detected by comparing the transcriptome sequence of an AML sample with the corresponding remission sample. Using this approach, we found five non-synonymous mutations specific to the tumor sample. They include a nonsense mutation affecting the RUNX1 gene, which is a known mutational target in AML, and a missense mutation in the putative tumor suppressor gene TLE4, which encodes a RUNX1 interacting protein. Another missense mutation was identified in SHKBP1, which acts downstream of FLT3, a receptor tyrosine kinase mutated in about 30% of AML cases. The frequency of mutations in TLE4 and SHKBP1 in 95 cytogenetically normal AML patients was 2%. Our study demonstrates that whole transcriptome sequencing leads to the rapid detection of recurring point mutations in the coding regions of genes relevant to malignant transformation

Autosomal recessive cortical myoclonic tremor and epilepsy: Association with a mutation in the potassium channel associated gene <em>CNTN2</em>.

We characterize a consanguineous Egyptian family with an autosomal recessively inherited familial cortical myoclonic tremor and epilepsy. We used multipoint linkage analysis to map the causative mutation to a 12.7 megabase interval within 1q31.3-q32.2 with a log of odds score of 3.6. For further investigation of the linked region in an efficient and unbiased manner, we performed exome sequencing. Within the suspected region we identified a homozygous single base pair deletion (c.503_503delG) leading to a frameshift in the coding region of the sixth exon of CNTN2 alias TAG-1 (p.Trp168fs), which segregated in the respective family. Many studies point towards an important role of the CNTN2 product contactin 2 in neuronal excitability. Contactin 2, a glycosylphosphatidylinositol-anchored neuronal membrane protein, and another transmembrane protein called contactin associated protein-like 2 (CNTNAP2 alias CASPR2) are together necessary to maintain voltage-gated potassium channels at the juxtaparanodal region. CNTN2 knockout mice were previously reported to suffer from spontaneous seizures and mutations in the CNTNAP2 gene have been described to cause epilepsy in humans. To further delineate the role of CNTN2 in patients with epilepsy, we sequenced the coding exons in 189 Caucasian patients with epilepsy. No recessive mutation was detected and heterozygote carriers of rare CNTN2 variants do not seem to be predisposed to epilepsy. Given the severity of the mutation and the proposed function of the gene, we consider this mutation as the most likely cause for cortical myoclonic tremor and epilepsy in this family