A Potent Mimetic of the Siglec-8 Ligand 6’-Sulfo-Sialyl Lewisx

Siglecs are members of the immunoglobulin gene family containing sialic acid binding N-terminal domains. Among them, Siglec-8 is expressed on various cell types of the immune system such as eosinophils, mast cells and weakly on basophils. Cross-linking of Siglec-8 with monoclonal antibodies triggers apoptosis in eosinophils and inhibits degranulation of mast cells, making Siglec-8 a promising target for the treatment of eosinophil- and mast cell-associated diseases such as asthma. The tetrasaccharide 6’-sulfo-sialyl Lewis^x has been identified as a specific Siglec-8 ligand in glycan array screening. Here, we describe an extended study enlightening the pharmacophores of 6’-sulfo-sialyl Lewis^x and the successful development of a high-affinity mimetic. Retaining the neuraminic acid core, the introduction of a carbocyclic mimetic of the Gal moiety and a sulfonamide substituent in the 9-position gave a 20-fold improved binding affinity. Finally, the residence time, which usually is the Achilles tendon of carbohydrate/lectin interactions, could be improved.     

Supramolecular Assembly of Aminoethylene‐Lipopeptide PMO Conjugates into RNA Splice‐Switching Nanomicelles

Phosphorodiamidate morpholino oligomers (PMOs) are oligonucleotide analogs that can be used for therapeutic modulation of pre‐mRNA splicing. Similar to other classes of nucleic acid‐based therapeutics, PMOs require delivery systems for efficient transport to the intracellular target sites. Here, artificial peptides based on the oligo(ethylenamino) acid succinyl‐tetraethylenpentamine (Stp), hydrophobic modifications, and an azide group are presented, which are used for strain‐promoted azide‐alkyne cycloaddition conjugation with splice‐switching PMOs. By systematically varying the lead structure and formulation, it is determined that the type of contained fatty acid and supramolecular assembly have a critical impact on the delivery efficacy. A compound containing linolenic acid with three cis double bonds exhibits the highest splice‐switching activity and significantly increases functional protein expression in pLuc/705 reporter cells in vitro and after local administration in vivo. Structural and mechanistic studies reveal that the lipopeptide PMO conjugates form nanoparticles, which accelerate cellular uptake and that the content of unsaturated fatty acids enhances endosomal escape. In an in vitro Duchenne muscular dystrophy exon skipping model using H2K‐mdx52 dystrophic skeletal myotubes, the highly potent PMO conjugates mediate significant splice‐switching at very low nanomolar concentrations. The presented aminoethylene‐lipopeptides are thus a promising platform for the generation of PMO‐therapeutics with a favorable activity/toxicity profile.     

Improvement of Aglycone π-Stacking Yields Nanomolar to Sub-nanomolar FimH Antagonists

Antimicrobial resistance has become a serious concern for the treatment of urinary tract infections. In this context, an anti-adhesive approach targeting FimH, a bacterial lectin enabling the attachment of E. coli to host cells, has attracted considerable interest. FimH can adopt a low/medium-affinity state in the absence and a high-affinity state in the presence of shear forces. Until recently, mostly the high-affinity state has been investigated, despite the fact that a therapeutic antagonist should bind predominantly to the low-affinity state. In this communication, we demonstrate that fluorination of biphenyl α-d -mannosides leads to compounds with perfect π–π stacking interactions with the tyrosine gate of FimH, yielding low nanomolar to sub-nanomolar K_D values for the low- and high-affinity states, respectively. The face-to-face alignment of the perfluorinated biphenyl group of FimH ligands and Tyr48 was confirmed by crystal structures as well as ¹H,¹⁵N-HSQC NMR analysis. Finally, fluorination improves pharmacokinetic parameters predictive for oral availability.     

Folgen des Globalen Wandels für das Grundwasser in Süddeutschland – Teil 2: Sozioökonomische Aspekte

In order to account for complex interactions between humans climate and the water cycle, the research consortium GLOWA-Danube (www.glowa-danube.de) has developed the simulation system DANUBIA which consists of 17 coupled models. DANUBIA was applied to investigate various impacts of global-change between 2011 and 2060 in the Upper Danube Catchment. This article describes part 2 of an article series with investigations of socio-economic aspects, while part 1 (Barthel et al. in Grundwasser 16(4), doi:10.1007/s007-011-01794, 2011) deals with natural-spatial aspects. The principles of socio-economic actor-modeling and interactions between socio-economic and natural science model components are described here. We present selected simulations that show impacts on groundwater from changes in agriculture, tourism, economy, domestic water users and water supply. Despite decreases in water consumption, the scenario simulations show significant decreases in groundwater quantity. On the other hand, groundwater quality will likely be influenced more severely by land use changes compared to direct climatic causes. However, overall changes will not be dramatic.     

Evaluation of buffer zone effectiveness in mitigating the risks associated with agricultural runoff in Prince Edward Island

To minimize the risk posed by runoff from row crops, Prince Edward Island introduced buffer legislation in 2000. The legislation mandates 10-m and 20-m buffers, respectively, for moderate sloped (i.e. < 5%) and steep sloped (i.e. > 5%) agricultural fields that border streams. Since 2001, Environment Canada has been evaluating the effectiveness of various buffer widths on operational farms in reducing toxicity and contaminant concentrations in runoff. Sample collectors, placed in 44 fields at the field edge (0 m), 10 m and at distances out to 30 m, collected overland flow following rainfall-induced runoff events. Samples were collected within 24 hours of an event and analysed for seven pesticides (endosulfan, chlorothalonil, carbofuran, linuron, metribuzin, metalaxyl, mancozeb), water quality parameters and Daphnia magna toxicity. The 10-m buffer required for moderate sloped fields was effective at reducing contaminant concentrations but not always to less than lethal concentrations to Daphnia magna. Limited data beyond 10 m for fields of both slope types precluded making recommendations on a suitable buffer width for shallow sloped fields and evaluating the effectiveness of 20-m buffers for steep sloped fields. When paired data were combined and statistically tested for all fields, the studied pesticides underwent a 52-98% and 68-100% reduction in aqueous and particulate concentrations within 10 m and 30 m, respectively. In addition, by 10 m, soluble phosphorus, nitrate-nitrogen and total suspended solids were reduced by 34%, 38% and 64%, respectively. Results suggest buffer zones on operational farms are capable of achieving contaminant reductions comparable to those reported for controlled experiments. Inconsistent siting of sample collectors beyond 10 m limited the evaluation of the effects of field slope and buffer width on buffer effectiveness on working farms. Future studies on buffer efficiency on operational farms should focus on building the data set beyond 10 m and evaluating load reductions.     

Impact of coagulation and adsorption on DOC fractions of secondary effluent and resulting fouling behaviour in ultrafiltration

Membrane fouling by macromolecular dissolved organic compounds is still a fundamental drawback in low-pressure membrane filtration of secondary effluent. In this study, pre-treatment of secondary effluent by coagulation and/or adsorption was investigated in terms of removal of different dissolved organic carbon (DOC) fractions, especially macromolecular substances. DOC fractionation has been characterised by size exclusion chromatography. Adsorption tests using four commercially available activated carbons yielded a removal of small as well as larger organic compounds, revealing differences in the affinity towards macromolecules depending on the type of applied activated carbon. By contrast, coagulation removed predominantly larger molecules, i.e., biopolymers and humic substances. In terms of DOC reduction, the coagulant ferric chloride was superior to aluminium chloride. A combination of coagulation and adsorption resulted in the addition of individual removal efficiencies, suggesting that different fractions of organic compounds were involved in each of the processes. After removal of macromolecular organic compounds either by coagulation or by adsorption, a significant reduction of membrane fouling was observed in tests using two different types of ultrafiltration flat-sheet membranes in 20-h cross-flow filtration tests.     

Rosuvastatin Enhances the Catabolism of LDL apoB‐100 in Subjects with Combined Hyperlipidemia in a Dose Dependent Manner

Dose‐associated effects of rosuvastatin on the metabolism of apolipoprotein (apo) B‐100 in triacylglycerol rich lipoprotein (TRL, d < 1.019 g/ml) and low density lipoprotein (LDL) and of apoA‐I in high density lipoprotein (HDL) were assessed in subjects with combined hyperlipidemia. Our primary hypothesis was that maximal dose rosuvastatin would decrease the apoB‐100 production rate (PR), as well as increase apoB‐100 fractional catabolic rate (FCR). Eight subjects received placebo, rosuvastatin 5 mg/day, and rosuvastatin 40 mg/day for 8 weeks each in sequential order. The kinetics of apoB‐100 in TRL and LDL and apoA‐I in HDL were determined at the end of each phase using stable isotope methodology, gas chromatography‐mass spectrometry, and multicompartmental modeling. Rosuvastatin at 5 and 40 mg/day decreased LDL cholesterol by 44 and 54 % (both P < 0.0001), triacylglycerol by 14 % (ns) and 35 % (P < 0.01), apoB by 30 and 36 % (both P < 0.0001), respectively, and had no significant effects on HDL cholesterol or apoA‐I levels. Significant decreases in plasma markers of cholesterol synthesis and increases in cholesterol absorption markers were observed. Rosuvastatin 5 and 40 mg/day increased TRL apoB‐100 FCR by 36 and 46 % (both ns) and LDL apoB‐100 by 63 and 102 % (both P < 0.05), respectively. HDL apoA‐I PR increased with low dose rosuvastatin (12 %, P < 0.05) but not with maximal dose rosuvastatin. Neither rosuvastatin dose altered apoB‐100 PR or HDL apoA‐I FCR. Our data indicate that maximal dose rosuvastatin treatment in subjects with combined hyperlipidemia resulted in significant increases in the catabolism of LDL apoB‐100, with no significant effects on apoB‐100 production or HDL apoA‐I kinetics.     

The Tyrosine Gate of the Bacterial Lectin FimH: A Conformational Analysis by NMR Spectroscopy and X‐ray Crystallography

Urinary tract infections caused by uropathogenic E. coli are among the most prevalent infectious diseases. The mannose‐specific lectin FimH mediates the adhesion of the bacteria to the urothelium, thus enabling host cell invasion and recurrent infections. An attractive alternative to antibiotic treatment is the development of FimH antagonists that mimic the physiological ligand. A large variety of candidate drugs have been developed and characterized by means of in vitro studies and animal models. Here we present the X‐ray co‐crystal structures of FimH with members of four antagonist classes. In three of these cases no structural data had previously been available. We used NMR spectroscopy to characterize FimH–antagonist interactions further by chemical shift perturbation. The analysis allowed a clear determination of the conformation of the tyrosine gate motif that is crucial for the interaction with aglycone moieties and was not obvious from X‐ray structural data alone. Finally, ITC experiments provided insight into the thermodynamics of antagonist binding. In conjunction with the structural information from X‐ray and NMR experiments the results provide a mechanism for the often‐observed enthalpy–entropy compensation of FimH antagonists that plays a role in fine‐tuning of the interaction.     

Helium droplet assisted synthesis of plasmonic Ag@ZnO core@shell nanoparticles

Plasmonic Ag@ZnO core@shell nanoparticles are formed by synthesis inside helium droplets with subsequent deposition and controlled oxidation. The particle size and shape can be controlled from spherical sub-10 nm particles to larger elongated structures. An advantage of the method is the complete absence of solvents, precursors, and other chemical agents. The obtained particle morphology and elemental composition have been analyzed by scanning transmission electron microscopy (STEM) and energy dispersive X-ray spectroscopy (EDS). The results reveal that the produced particles form a closed and homogeneous ZnO layer around a 2–3 nm Ag core with a uniform thickness of (1.33 ± 0.15) nm and (1.63 ± 0.31) nm for spherical and wire-like particles, respectively. The results are supported by ultraviolet photoelectron spectroscopy (UPS), which indicates a fully oxidized shell layer for the particles studied by STEM. The plasmonic properties of the produced spherical Ag@ZnO core@shell particles are investigated by two-photon photoelectron (2PPE) spectroscopy. Upon excitation of the localized surface plasmon resonance in Ag at around 3 eV, plasmonic enhancement leads to the liberation of electrons with high kinetic energy. This is observed for both Ag and Ag@ZnO particles, showing that even if a Ag cluster is covered by the ZnO layer, a plasmonic enhancement can be observed by photoelectron spectroscopy.