CloudBooks - LOOP ein neues Autorentool

Im E-Learning-Bereich gewinnt die technische Anforderung, mit mobilen Endgeräten auf Lerninhalte zugreifen zu können, zunehmend an Bedeutung. Ein zentraler Online-Speicherort, von dem die Inhalte – optimiert für das jeweilige Endgerät – bereitgestellt und verändert werden können, ist zur Erfüllung dieser Anforderung elementar. (...

Optimizing the anti-tumor efficacy of protein-drug conjugates by engineering the molecular size and half-life

Despite some approvals of antibody-drug conjugates for cancer therapy, their clinical success rate is unsatisfactory because of very small therapeutic windows, influenced by on-target and off-target toxicities of conjugate and liberated toxin. Additional formats with systematically investigated molecular parameters must therefore be explored to increase their therapeutic window. Here we focused on the effective molecular weight. To generate conjugates with exactly defined drug loads and tunable pharmacokinetics, we used Designed Ankyrin Repeat Proteins (DARPins), fused to unstructured polypeptides of different lengths, to produce proteins with any desired half-life, to identify those with the best efficacy. We generated an EpCAM-targeting DARPin-MMAF conjugate, fused to PAS or XTEN of different lengths, and a matched series of controls of a non-binding DARPin to account for the enhanced permeability and retention (EPR) effect, covering half-lives of minutes to 20.6 h in mice. All conjugates were produced at high purity, and demonstrated high specificity and cytotoxicity in human tumor cell cultures, with IC50 values in the low nM range, independent of the polypeptide type and length. Due to their more facile purification, the PASylated conjugates were tested in nude mice bearing HT29 tumor xenografts. Independent of their size, all PASylated conjugates were very well tolerated after repeated systemic administration of 300 nmol/kg. We found that the conjugates with intermediate size and half-life showed the strongest anti-tumor effects, and deduced that this effect is a compromise of serum half-life and diffusion within the tumor, as on-rates and affinities are essentially identical, with extravasation playing only a very minor role

Targeted delivery and endosomal cellular uptake of DARPin-siRNA bioconjugates: Influence of linker stability on gene silencing

Specific cell targeting and efficient intracellular delivery are major hurdles for the widespread therapeutic use of nucleic acid technologies, particularly siRNA mediated gene silencing. To enable receptor-mediated cell-specific targeting, we designed a synthesis scheme that can be generically used to engineer Designed Ankyrin Repeat Protein (DARPin)-siRNA bioconjugates. Different linkers, including labile disulfide-, and more stable thiol-maleimide- and triazole- (click chemistry) tethers were employed. Crosslinkers were first attached to a 3’-terminal aminohexyl chain on the siRNA sense strands. On the protein side thiols of a C-terminal cysteine were used as anchoring site for disulfide- and thiol-maleimide conjugate formation, while strain-promoted azido-alkyne cycloadditions were carried out at a metabolically introduced N-terminal azidohomoalanine. After establishing efficient purification methods, highly pure products were obtained. Bioconjugates of EpCAM-targeted DARPins with siRNA directed at the luciferase gene were evaluated for cell-specific binding, uptake and gene silencing. As shown by flow cytometry and fluorescence microscopy, all constructs retained the highly specific and high-affinity antigen recognition properties of the native DARPin. As expected, internalization was observed only in EpCAM-positive cell lines, and predominantly endolysosomal localization was detected. Disulfide linked conjugates showed lower serum stability against cleavage at the linker and thus lower internalization into endosomes compared to thiol-maleimide- and triazole-linked conjugates, yet induced more pronounced gene silencing. This indicates that the siRNA payload needs to be liberated from the protein in the endosome. Our data confirm the promise of DARPin-siRNA bioconjugates for tumor targeting, but also identified endosomal retention and limited cytosolic escape of the siRNA as the rate-limiting step for more efficient gene silencing

Portal hypertension in common variable immunodeficiency disorders – a single center analysis on clinical and immunological parameter in 196 patients

BackgroundLiver manifestations and in particular portal hypertension (PH) contribute significantly to morbidity and mortality of patients with common variable immunodeficiency disorders (CVID). Screening strategies and early detection are limited due to the lack of specific diagnostic tools.MethodsWe evaluated clinical, immunological, histological, and imaging parameters in CVID patients with clinical manifestation of portal hypertension (CVID+PH).ResultsPortal hypertension was present in 5.6% of CVID patients and was associated with high clinical burden and increased mortality (18%). Longitudinal data on clinical and immunological parameters in patients before and during clinically manifest portal hypertension revealed a growing splenomegaly and increasing gamma-glutamyl transferase (GGT) and soluble interleukin 2 receptor (SIL-2R) levels with decreasing platelets over time. While ultrasound of the liver failed to detect signs of portal hypertension in most affected patients, transient elastography was elevated in all patients. All CVID+PH patients had reduced naïve CD45RA+CD4+ T-cells (mean of 6,2%). The frequency of severe B-lymphocytopenia (Euroclass B-) was higher in CVID+PH patients. The main histological findings included lymphocytic infiltration, nodular regenerative hyperplasia-like changes (NRH-LC), and porto(-septal) fibrosis.ConclusionCVID patients with lower naïve CD45RA+CD4+ T-cells or severely reduced B-cells might be at higher risk for portal hypertension. The combination of biochemical (increasing sIL-2R, GGT, and decreasing platelets) and imaging parameters (increasing splenomegaly) should raise suspicion of the beginning of portal hypertension

Metastandard für den internationalen Austausch von MOOCs – der MOOChub als erster Prototyp

Der MOOChub ist eine Webseite, die weit über 700 Massive Open Online Courses (MOOCs) aus dem deutschsprachigen Raum von insgesamt neun unterschiedlichen Partner:innen listet. Damit eine solche Seite automatisiert aufgebaut werden kann, ist es notwendig, dass alle Partner:innen die Metadaten der Kurse in gleicher Weise beschreiben und verfügbar machen. Dieser Artikel beschreibt zunächst die Entstehung der Idee eines gemeinsamen Standards und wie dieser im Anschluss entwickelt worden ist. Das Ergebnis ist einerseits ein offen lizenzierter Quasi-Standard, der sich an üblichen Standards orientiert, und ein erster Prototyp, der sogenannte MOOChub, auf dem nun alle Kurse auffindbar und durchsuchbar sind. Abschließend wird über die nächsten möglichen und auch notwendigen Entwicklungen berichtet, die die Schnittstelle weiter optimieren sollen

A Distinct Urinary Biomarker Pattern Characteristic of Female Fabry Patients That Mirrors Response to Enzyme Replacement Therapy

Female patients affected by Fabry disease, an X-linked lysosomal storage disorder, exhibit a wide spectrum of symptoms, which renders diagnosis, and treatment decisions challenging. No diagnostic test, other than sequencing of the alpha-galactosidase A gene, is available and no biomarker has been proven useful to screen for the disease, predict disease course and monitor response to enzyme replacement therapy. Here, we used urine proteomic analysis based on capillary electrophoresis coupled to mass spectrometry and identified a biomarker profile in adult female Fabry patients. Urine samples were taken from 35 treatment-naive female Fabry patients and were compared to 89 age-matched healthy controls. We found a diagnostic biomarker pattern that exhibited 88.2% sensitivity and 97.8% specificity when tested in an independent validation cohort consisting of 17 treatment-naive Fabry patients and 45 controls. The model remained highly specific when applied to additional control patients with a variety of other renal, metabolic and cardiovascular diseases. Several of the 64 identified diagnostic biomarkers showed correlations with measures of disease severity. Notably, most biomarkers responded to enzyme replacement therapy, and 8 of 11 treated patients scored negative for Fabry disease in the diagnostic model. In conclusion, we defined a urinary biomarker model that seems to be of diagnostic use for Fabry disease in female patients and may be used to monitor response to enzyme replacement therapy

Explorative data analysis of MCL reveals gene expression networks implicated in survival and prognosis supported by explorative CGH analysis

<p>Abstract</p> <p>Background</p> <p>Mantle cell lymphoma (MCL) is an incurable B cell lymphoma and accounts for 6% of all non-Hodgkin's lymphomas. On the genetic level, MCL is characterized by the hallmark translocation t(11;14) that is present in most cases with few exceptions. Both gene expression and comparative genomic hybridization (CGH) data vary considerably between patients with implications for their prognosis.</p> <p>Methods</p> <p>We compare patients over and below the median of survival. Exploratory principal component analysis of gene expression data showed that the second principal component correlates well with patient survival. Explorative analysis of CGH data shows the same correlation.</p> <p>Results</p> <p>On chromosome 7 and 9 specific genes and bands are delineated which improve prognosis prediction independent of the previously described proliferation signature. We identify a compact survival predictor of seven genes for MCL patients. After extensive re-annotation using GEPAT, we established protein networks correlating with prognosis. Well known genes (CDC2, CCND1) and further proliferation markers (WEE1, CDC25, aurora kinases, BUB1, PCNA, E2F1) form a tight interaction network, but also non-proliferative genes (SOCS1, TUBA1B CEBPB) are shown to be associated with prognosis. Furthermore we show that aggressive MCL implicates a gene network shift to higher expressed genes in late cell cycle states and refine the set of non-proliferative genes implicated with bad prognosis in MCL.</p> <p>Conclusion</p> <p>The results from explorative data analysis of gene expression and CGH data are complementary to each other. Including further tests such as Wilcoxon rank test we point both to proliferative and non-proliferative gene networks implicated in inferior prognosis of MCL and identify suitable markers both in gene expression and CGH data.</p

The German National Registry of Primary Immunodeficiencies (2012-2017)

Introduction: The German PID-NET registry was founded in 2009, serving as the first national registry of patients with primary immunodeficiencies (PID) in Germany. It is part of the European Society for Immunodeficiencies (ESID) registry. The primary purpose of the registry is to gather data on the epidemiology, diagnostic delay, diagnosis, and treatment of PIDs. Methods: Clinical and laboratory data was collected from 2,453 patients from 36 German PID centres in an online registry. Data was analysed with the software Stata® and Excel. Results: The minimum prevalence of PID in Germany is 2.72 per 100,000 inhabitants. Among patients aged 1–25, there was a clear predominance of males. The median age of living patients ranged between 7 and 40 years, depending on the respective PID. Predominantly antibody disorders were the most prevalent group with 57% of all 2,453 PID patients (including 728 CVID patients). A gene defect was identified in 36% of patients. Familial cases were observed in 21% of patients. The age of onset for presenting symptoms ranged from birth to late adulthood (range 0–88 years). Presenting symptoms comprised infections (74%) and immune dysregulation (22%). Ninety-three patients were diagnosed without prior clinical symptoms. Regarding the general and clinical diagnostic delay, no PID had undergone a slight decrease within the last decade. However, both, SCID and hyper IgE- syndrome showed a substantial improvement in shortening the time between onset of symptoms and genetic diagnosis. Regarding treatment, 49% of all patients received immunoglobulin G (IgG) substitution (70%—subcutaneous; 29%—intravenous; 1%—unknown). Three-hundred patients underwent at least one hematopoietic stem cell transplantation (HSCT). Five patients had gene therapy. Conclusion: The German PID-NET registry is a precious tool for physicians, researchers, the pharmaceutical industry, politicians, and ultimately the patients, for whom the outcomes will eventually lead to a more timely diagnosis and better treatment