Discovery of novel pan-genotypic HCV NS5A inhibitors containing a novel tetracyclic core

A series of novel tetracyclic core-containing HCV NS5A inhibitors has been discovered. Incorporation of tetrahydropyran-substituted amino acid moiety improved their potency and yielded HCV NS5A inhibitors with a minimum potency shift from the GT1a strain compared to other genotypes and mutants. Compounds 53 and 54 showed the best potency profile and had reasonable half-times in rat PK studies. However, further optimization of their oral bioavailability is still needed in order to advance them for further development. [BMCL ABSTRACT] ©2000 Elsevier Science Ltd. All rights reserved.     

PGE2 MEDIATES OENOCYTOID CELL LYSIS VIA A SODIUM‐POTASSIUM‐CHLORIDE COTRANSPORTER

Prostaglandin E₂ (PGE₂) mediates immune responses of the beet armyworm, Spodoptera exigua, including oenocytoid cell lysis (a class of lepidopteran hemocytes: OCL) via its specific membrane receptor to release inactive prophenoloxidase (PPO) into hemolymph. PPO is activated into phenoloxidase in the plasma to play crucial roles in the immune responses of S. exigua. The mechanism of OCL has not been elucidated, however we posed the hypothesis that a rapid accumulation of sodium ions within the oenocytoids allows a massive influx of water by the ion gradient, which leads to the cell lysis. It remains unclear which sodium channel is responsible for the OCL in response to PGE₂. This study identified a specific sodium channel called sodium‐potassium‐chloride cotransporter 1 (Se‐NKCC1) expressed in hemocytes of S. exigua and analyzed its function in the OCL in response to PGE₂. Se‐NKCC1 encodes a basic membrane protein (pI value = 8.445) of 1,066 amino acid residues, which contains 12 putative transmembrane domains. Se‐NKCC1 was expressed in all developmental stages and tissues. qPCR showed that bacterial challenge significantly induced its expression. A specific inhibitor of NKCC, bumetanide, prevented the OCL in a dose‐dependent manner. When RNA interference (RNAi) using double‐stranded RNA specific to Se‐NKCC1 suppressed its expression, the OCL and PPO activation were significantly inhibited in response to PGE₂. The RNAi treatment also reduced nodule formation to bacterial challenge. These results suggest that Se‐NKCC1 is associated with OCL by facilitating inward transport of ions in response to PGE₂.     

Spatial Variation in Army Ant Swarm Raiding and its Potential Effect on Biodiversity

Invertebrate communities of the tropical rain forest floor are highly diverse, characterized by patchy species distribution patterns and high variation in species density. Spatial variation in the foraging activity of swarm raiding army ants, prime invertebrate predators in tropical rain forests, is discussed as a mechanism contributing to these patterns, but highly resolved long‐term data on army ant raiding on the local and landscape scale are hitherto lacking. In this study, 196 positions in 11 study sites in a tropical rain forest in western Kenya were continuously monitored over ~4 mo for the occurrence of swarm raids of army ants. Using population simulation analyses, the consequences of army ant raiding for prey communities were assessed. We found an unexpectedly high variation in raid rates at the study site and landscape scale. The weekly chance of communities to become raided by army ants was on average 0.11, but ranged from 0 to 0.50 among the 196 positions. Simulating population developments of two Lotka–Volterra species—showing slight trade‐offs between competitive strength and resistance to army ant raids—in the real raiding landscapes showed that the observed spatial variation in raid rates may produce high prey diversity at larger spatial scales (due to high β‐diversity) and strong variation in species density. Our results indicate that high spatial variation in army ant swarm raiding is a mechanism capable of generating patchy species distribution patterns and maintaining the high biodiversity of invertebrate communities of the tropical rain forest floor.     

Improved phylogenetic analyses corroborate a plausible position of Martialis heureka in the ant tree of life

Martialinae are pale, eyeless and probably hypogaeic predatory ants. Morphological character sets suggest a close relationship to the ant subfamily Leptanillinae. Recent analyses based on molecular sequence data suggest that Martialinae are the sister group to all extant ants. However, by comparing molecular studies and different reconstruction methods, the position of Martialinae remains ambiguous. While this sister group relationship was well supported by Bayesian partitioned analyses, Maximum Likelihood approaches could not unequivocally resolve the position of Martialinae. By re-analysing a previous published molecular data set, we show that the Maximum Likelihood approach is highly appropriate to resolve deep ant relationships, especially between Leptanillinae, Martialinae and the remaining ant subfamilies. Based on improved alignments, alignment masking, and tree reconstructions with a sufficient number of bootstrap replicates, our results strongly reject a placement of Martialinae at the first split within the ant tree of life. Instead, we suggest that Leptanillinae are a sister group to all other extant ant subfamilies, whereas Martialinae branch off as a second lineage. This assumption is backed by approximately unbiased (AU) tests, additional Bayesian analyses and split networks. Our results demonstrate clear effects of improved alignment approaches, alignment masking and data partitioning. We hope that our study illustrates the importance of thorough, comprehensible phylogenetic analyses using the example of ant relationships.     

Low Resolution Solution Structure of HAMLET and the Importance of Its Alpha-Domains in Tumoricidal Activity

HAMLET (Human Alpha-lactalbumin Made LEthal to Tumor cells) is the first member in a new family of protein-lipid complexes with broad tumoricidal activity. Elucidating the molecular structure and the domains crucial for HAMLET formation is fundamental for understanding its tumoricidal function. Here we present the low-resolution solution structure of the complex of oleic acid bound HAMLET, derived from small angle X-ray scattering data. HAMLET shows a two-domain conformation with a large globular domain and an extended part of about 2.22 nm in length and 1.29 nm width. The structure has been superimposed into the related crystallographic structure of human α-lactalbumin, revealing that the major part of α-lactalbumin accommodates well in the shape of HAMLET. However, the C-terminal residues from L105 to L123 of the crystal structure of the human α-lactalbumin do not fit well into the HAMLET structure, resulting in an extended conformation in HAMLET, proposed to be required to form the tumoricidal active HAMLET complex with oleic acid. Consistent with this low resolution structure, we identified biologically active peptide epitopes in the globular as well as the extended domains of HAMLET. Peptides covering the alpha1 and alpha2 domains of the protein triggered rapid ion fluxes in the presence of sodium oleate and were internalized by tumor cells, causing rapid and sustained changes in cell morphology. The alpha peptide-oleate bound forms also triggered tumor cell death with comparable efficiency as HAMLET. In addition, shorter peptides corresponding to those domains are biologically active. These findings provide novel insights into the structural prerequisites for the dramatic effects of HAMLET on tumor cells.     

High Defibrillation Threshold: The Science,Signs and Solutions

Defibrillation threshold (DFT) testing has traditionally been an integral part of implantable cardioverter defibrillator (ICD) implantation. With the increasing number of patients receiving ICDs, physicians are encountering high DFT more often than before. Tackling the problem of high DFT, warrants an in-depth understanding of the science of defibrillation including the key electrophysiological concepts and the underlying molecular mechanisms. Numerous factors have been implicated in the causation of high DFT. Due consideration to the past medical history, pharmacotherapy, laboratory data and cardiac imaging, help in assessing the pre-procedural risk for occurrence of high DFT. Drugs, procedural changes, type and location of ICD lead system are some of the key players in predicting DFT during implantation. In the event of encountering an unacceptably high DFT, we recommend to follow a step-wise algorithm. Ruling out procedural complications like pneumothorax and tamponade is imperative before embarking on a search for potentially reversible clinical or metabolic derangements. Finally, if these attempts fail, the electrophysiologist must choose from a wide range of options for device adjustment and system modification. Although this review article is meant to be a treatise on the science, signs and solutions for high DFT, it is bound by limitations of space and scope of the article.     

New records of ant species from Yunnan,China

As with many other regions of the world, significant collecting, curation, and taxonomic efforts will be needed to complete the inventory of China’s ant fauna. This is especially true for the highly diverse tropical regions in the south of the country, where moist tropical forests harbor high species richness typical of the Southeast Asian region. We inventoried ants in the Xingshuangbanna prefecture, Yunnan, in June 2013, using a variety of methods including Winkler extraction and hand collection to sample ant diversity. We identified 213 species/morphospecies of ants from 10 subfamilies and 61 genera. After identification of 148 valid species of the 213 total species collected, 40 species represent new records for Yunnan province and 17 species are newly recorded for China. This increases the total number of named ant species in Yunnan and China to 447 and 951 respectively. The most common species collected were Brachyponera luteipes and Vollenhovia emeryi. Only one confirmed exotic species Strumigenys membranifera, was collected, although several others were potentially introduced by humans. These results highlight the high biodiversity value of the region, but also underscore how much work remains to fully document the native myrmecofauna.     

Synthesis of 2-monoacylglycerols and structured triacylglycerols rich in polyunsaturated fatty acids by enzyme catalyzed reactions

This paper studies the synthesis of structured triacylglycerols (STAGs) by a four-step process: (i) obtaining 2-monoacylglycerols (2-MAGs) by alcoholysis of cod liver oil with several alcohols, catalyzed by lipases Novozym 435, from Candida antartica and DF, from Rhizopus oryzae, (ii) purification of 2-MAGs, (iii) formation of STAGs by esterification of 2-MAGs with caprylic acid catalyzed by lipase DF, from R. oryzae, and (iv) purification of these STAGs. For the alcoholysis of cod liver oil, absolute ethanol, ethanol 96% (v/v) and 1-butanol were compared; the conditions with ethanol 96% were then optimized and 2-MAG yields of around 54-57% were attained using Novozym 435. In these 2-MAGs, DHA accounted for 24-31% of total fatty acids. In the operational conditions this lipase maintained a stable level of activity over at least 11 uses. These results were compared with those obtained with lipase DF, which deactivated after only three uses. The alcoholysis of cod liver oil and ethanol 96% catalyzed by Novozym 435 was scaled up by multiplying the reactant amounts 100-fold and maintaining the intensity of treatment constant (IOT=3g lipase h/g oil). In these conditions, the 2-MAG yield attained was about 67%; these 2-MAGs contained 36.6% DHA. The synthesized 2-MAGs were separated and purified from the alcoholysis reaction products by solvent extraction using solvents of low toxicity (ethanol and hexane); 2-MAG recovery yield and purity of the target product were approximately 96.4% and 83.9%, respectively. These 2-MAGs were transformed to STAGs using the optimal conditions obtained in a previous work. After synthesis and purification, 93% pure STAGs were obtained, containing 38% DHA at sn-2 position and 60% caprylic acid (CA) at sn-1,3 positions (of total fatty acids at these positions), i.e. the major TAG is the STAG with the structure CA-DHA-CA.