Therapeutic prospects of exon skipping for epidermolysis bullosa

Epidermolysis bullosa is a group of genetic skin conditions characterized by abnormal skin (and mucosal) fragility caused by pathogenic variants in various genes. The disease severity ranges from early childhood mortality in the most severe types to occasional acral blistering in the mildest types. The subtype and severity of EB is linked to the gene involved and the specific variants in that gene, which also determine its mode of inheritance. Current treatment is mainly focused on symptomatic relief such as wound care and blister prevention, because truly curative treatment options are still at the preclinical stage. Given the current level of understanding, the broad spectrum of genes and variants underlying EB makes it impossible to develop a single treatment strategy for all patients. It is likely that many different variant-specific treatment strategies will be needed to ultimately treat all patients. Antisense-oligonucleotide (ASO)-mediated exon skipping aims to counteract pathogenic sequence variants by restoring the open reading frame through the removal of the mutant exon from the pre-messenger RNA. This should lead to the restored production of the protein absent in the affected skin and, consequently, improvement of the phenotype. Several preclinical studies have demonstrated that exon skipping can restore protein production in vitro, in skin equivalents, and in skin grafts derived from EB-patient skin cells, indicating that ASO-mediated exon skipping could be a viable strategy as a topical or systemic treatment. The potential value of exon skipping for EB is supported by a study showing reduced phenotypic severity in patients who carry variants that result in natural exon skipping. In this article, we review the substantial progress made on exon skipping for EB in the past 15 years and highlight the opportunities and current challenges of this RNA-based therapy approach. In addition, we present a prioritization strategy for the development of exon skipping based on genomic information of all EB-involved genes

Rationale and design of the PRAETORIAN-COVID trial:A double-blind, placebo-controlled randomized clinical trial with valsartan for PRevention of Acute rEspiraTORy dIstress syndrome in hospitAlized patieNts with SARS-COV-2 Infection Disease

There is much debate on the use of angiotensin receptor blockers (ARBs) in severe acute respiratory syndrome–coronavirus-2 (SARS-CoV-2)–infected patients. Although it has been suggested that ARBs might lead to a higher susceptibility and severity of SARS-CoV-2 infection, experimental data suggest that ARBs may reduce acute lung injury via blocking angiotensin-II–mediated pulmonary permeability, inflammation, and fibrosis. However, despite these hypotheses, specific studies on ARBs in SARS-CoV-2 patients are lacking. Methods: The PRAETORIAN-COVID trial is a multicenter, double-blind, placebo-controlled 1:1 randomized clinical trial in adult hospitalized SARS-CoV-2–infected patients (n = 651). The primary aim is to investigate the effect of the ARB valsartan compared to placebo on the composite end point of admission to an intensive care unit, mechanical ventilation, or death within 14 days of randomization. The active-treatment arm will receive valsartan in a dosage titrated to blood pressure up to a maximum of 160 mg bid, and the placebo arm will receive matching placebo. Treatment duration will be 14 days, or until the occurrence of the primary end point or until hospital discharge, if either of these occurs within 14 days. The trial is registered at clinicaltrials.gov (NCT04335786, 2020). The PRAETORIAN-COVID trial is a double-blind, placebo-controlled 1:1 randomized trial to assess the effect of valsartan compared to placebo on the occurrence of ICU admission, mechanical ventilation, and death in hospitalized SARS-CoV-2–infected patients. The results of this study might impact the treatment of SARS-CoV-2 patients globally

MoVam7, a Conserved SNARE Involved in Vacuole Assembly, Is Required for Growth, Endocytosis, ROS Accumulation, and Pathogenesis of Magnaporthe oryzae

Soluble NSF attachment protein receptor (SNARE) proteins play a central role in membrane fusion and vesicle transport of eukaryotic organisms including fungi. We previously identified MoSce22 as a homolog of Saccharomyces cerevisiae SNARE protein Sec22 to be involved in growth, stress resistance, and pathogenicity of Magnaporthe oryzae. Here, we provide evidences that MoVam7, an ortholog of S. cerevisiae SNARE protein Vam7, exerts conserved functions in vacuolar morphogenesis and functions in pathogenicity of M. oryzae. Staining with neutral red and FM4-64 revealed the presence of abnormal fragmented vacuoles and an absence of the Spitzenkörper body in the ΔMovam7 mutant. The ΔMovam7 mutant also exhibited reduced vegetative growth, poor conidiation, and failure to produce the infection structure appressorium. Additionally, treatments with cell wall perturbing agents indicated weakened cell walls and altered distributions of the cell wall component chitin. Furthermore, the ΔMovam7 mutant showed a reduced accumulation of reactive oxygen species (ROS) in the hyphal apex and failed to cause diseases on the rice plant. In summary, our studies indicate that MoVam7, like MoSec22, is a component of the SNARE complex whose functions in vacuole assembly also underlies the growth, conidiation, appressorium formation, and pathogenicity of M. oryzae. Further studies of MoVam7, MoSec22, and additional members of the SNARE complex are likely to reveal critical mechanisms in vacuole formation and membrane trafficking that is linked to fungal pathogenicity

Phase-transformation and precipitation kinetics in vanadium micro-alloyed steels by in-situ, simultaneous neutron diffraction and SANS

In-situ Neutron Diffraction and Small-Angle Neutron Scattering (SANS) are employed for the first time simultaneously in order to reveal the interaction between the austenite to ferrite phase transformation and the precipitation kinetics during isothermal annealing at 650 and at 700 °C in three steels with different vanadium (V) and carbon (C) concentrations. Austenite-to-ferrite phase transformation is observed in all three steels at both temperatures. The phase transformation is completed during a 10 h annealing treatment in all cases. The phase transformation is faster at 650 than at 700 °C for all alloys. Additions of vanadium and carbon to the steel composition cause a retardation of the phase transformation. The effect of each element is explained through its contribution to the Gibbs free energy dissipation. The austenite-to-ferrite phase transformation is found to initiate the vanadium carbide precipitation. Larger and fewer precipitates are detected at 700 than at 650 °C in all three steels, and a larger number density of precipitates is detected in the steel with higher concentrations of vanadium and carbon. After 10 h of annealing, the precipitated phase does not reach the equilibrium fraction as calculated by ThermoCalc. The external magnetic field applied during the experiments, necessary for the SANS measurements, causes a delay in the onset and time evolution of the austenite-to-ferrite phase transformation and consequently on the precipitation kinetics.Team Erik OffermanTeam Jilt SietsmaRST/Neutron and Positron Methods in MaterialsInstrumenten groe

Evolution of the precipitate composition during annealing of vanadium micro-alloyed steels by in-situ SANS

In-situ Small-Angle Neutron Scattering (SANS) is used to determine the time evolution of the chemical composition of precipitates at 650 °C and 700 °C in three micro-alloyed steels with different vanadium (V) and carbon (C) concentrations. Precipitates with a distribution of substoichiometric carbon-to-metal ratios are measured in all steels. The precipitates are initially metastable with a high iron (Fe) content, which is gradually being substituted by vanadium during isothermal annealing. Eventually a plateau in the composition of the precipitate phase is reached. Faster changes in the precipitate chemical composition are observed at the higher temperature in all steels because of the faster vanadium diffusion at 700 °C. At both temperatures, the addition of more vanadium and more carbon to the steel has an accelerating effect on the evolution of the precipitate composition as a result of a higher driving force for precipitation. Addition of vanadium to the nominal composition of the steel leads to more vanadium rich precipitates, with less iron and a smaller carbon-to-metal ratio. Atom Probe Tomography (APT) shows the presence of precipitates with a distribution of carbon-to-metal ratios, ranging from 0.75 to 1, after 10 h of annealing at 650 °C or 700 °C in all steels. These experimental results are coupled to ThermoCalc equilibrium calculations and literature findings to support the Small-Angle Neutron Scattering results.(OLD) MSE-1(OLD) MSE-3RST/Neutron and Positron Methods in MaterialsMaterials Science and EngineeringBedrijfsondersteunin

Interaction of precipitation with austenite-to-ferrite phase transformation in vanadium micro-alloyed steels

The precipitation kinetics of vanadium carbides and its interaction with the austenite-to-ferrite phase transformation is studied in two micro-alloyed steels that differ in vanadium and carbon concentrations by a factor of two, but have the same vanadium-to-carbon atomic ratio of 1:1. Dilatometry is used for heat-treating the specimens and studying the phase transformation kinetics during annealing at isothermal holding temperatures of 900, 750 and 650 °C for up to 10 h. Small-Angle Neutron Scattering (SANS) and Atom Probe Tomography (APT) measurements are performed to study the vanadium carbide precipitation kinetics. Vanadium carbide precipitation is not observed after annealing for 10 h at 900 and 750 °C, which is contrary to predictions from thermodynamic equilibrium calculations. Vanadium carbide precipitation is only observed during or after the austenite-to-ferrite phase transformation at 650 °C. The precipitate volume fraction and mean radius continuously increase as holding time increases, while the precipitate number density starts to decrease after 20 min, which corresponds to the time at which the austenite-to-ferrite phase transformation is finished. This indicates that nucleation and growth are dominant during the first 20 min, while later precipitate growth with soft impingement (overlapping diffusion fields) and coarsening take place. APT shows gradual changes in the precipitate chemical composition during annealing at 650 °C, which finally reaches a 1:1 atomic ratio of vanadium-to-carbon in the core of the precipitates after 10 h.Green Open Access added to TU Delft Institutional Repository ‘You share, we take care!’ – Taverne project https://www.openaccess.nl/en/you-share-we-take-care Otherwise as indicated in the copyright section: the publisher is the copyright holder of this work and the author uses the Dutch legislation to make this work public.(OLD) MSE-1RST/Neutron and Positron Methods in MaterialsMaterials Science and EngineeringBedrijfsondersteunin

A polygenic two-hit hypothesis for prostate cancer

Prostate cancer is one of the most heritable cancers. Hundreds of germline polymorphisms have been linked to prostate cancer diagnosis and prognosis. Polygenic risk scores can predict genetic risk of a prostate cancer diagnosis. Although these scores inform the probability of developing a tumor, it remains unknown how germline risk influences the tumor molecular evolution. We cultivated a cohort of 1250 localized European-descent patients with germline and somatic DNA profiling. Men of European descent with higher genetic risk were diagnosed earlier and had less genomic instability and fewer driver genes mutated. Higher genetic risk was associated with better outcome. These data imply a polygenic "two-hit" model where germline risk reduces the number of somatic alterations required for tumorigenesis. These findings support further clinical studies of polygenic risk scores as inexpensive and minimally invasive adjuncts to standard risk stratification. Further studies are required to interrogate generalizability to more ancestrally and clinically diverse populations

A polygenic two-hit hypothesis for prostate cancer

Prostate cancer is one of the most heritable cancers. Hundreds of germline polymorphisms have been linked to prostate cancer diagnosis and prognosis. Polygenic risk scores can predict genetic risk of a prostate cancer diagnosis. Although these scores inform the probability of developing a tumor, it remains unknown how germline risk influences the tumor molecular evolution. We cultivated a cohort of 1250 localized European-descent patients with germline and somatic DNA profiling. Men of European descent with higher genetic risk were diagnosed earlier and had less genomic instability and fewer driver genes mutated. Higher genetic risk was associated with better outcome. These data imply a polygenic "two-hit" model where germline risk reduces the number of somatic alterations required for tumorigenesis. These findings support further clinical studies of polygenic risk scores as inexpensive and minimally invasive adjuncts to standard risk stratification. Further studies are required to interrogate generalizability to more ancestrally and clinically diverse populations.</p

Integrative genome analyses identify key somatic driver mutations of small-cell lung cancer

Small-cell lung cancer (SCLC) is an aggressive lung tumor subtype with poor prognosis. We sequenced 29 SCLC exomes, 2 genomes and 15 transcriptomes and found an extremely high mutation rate of 7.4 ± 1 protein-changing mutations per million base pairs. Therefore, we conducted integrated analyses of the various data sets to identify pathogenetically relevant mutated genes. In all cases, we found evidence for inactivation of TP53 and RB1 and identified recurrent mutations in the CREBBP, EP300 and MLL genes that encode histone modifiers. Furthermore, we observed mutations in PTEN, SLIT2 and EPHA7, as well as focal amplifications of the FGFR1 tyrosine kinase gene. Finally, we detected many of the alterations found in humans in SCLC tumors from Tp53 and Rb1 double knockout mice. Our study implicates histone modification as a major feature of SCLC, reveals potentially therapeutically tractable genomic alterations and provides a generalizable framework for the identification of biologically relevant genes in the context of high mutational background