K voprosu o peredače prjamoj reči v chudožestvennom tekste (M.Ju.Lermontov)

On the representation of direct discourse in the artistic text (M. Iu. Lermontov) In her book Intonation in the Artistic Text N. D. Svetozarova presented an impressive quantity of material, as well as a refined categorization of this extremely interesting aspect of the artistic text — the occurrence and the specifi c forms of representation of features of intonation, tone, manner of speaking of literary characters. Thus her book has become an important contribution to contemporary narratology as well, especially with regard to the problem of the relationship between the narrator’s discourse and the discourse of literary characters. In her book the author also discusses the frequency and functions of this phenomenon in the works of different authors and in different literary periods. The present article aims to add to the findings of N. D. Svetozarova; a brief outline is given of the occurrence and roles of this phenomenon in the period 1790-1840. Special attention is given to Lermontov

Personalized medicine with biologics for severe type 2 asthma : current status and future prospects

Asthma affects more than 300 million people worldwide and poses a large socioeconomic burden, particularly in the 5% to 10% of severe asthmatics. So far, each entry of new biologics in clinical trials has led to high expectations for treating all severe asthma forms, but the outcome has only been successful if the biologic, as add-on treatment, targeted specific patient subgroups. Indeed, we now realize that asthma is a heterogeneous disease with multiple phenotypes, based on distinct pathophysiological mechanisms, called endotypes. Thus, asthma therapy is gradually moving to a personalized medicine approach, tailored to individual's asthma endotypes identified through biomarkers. Here, we review the clinical efficacy of antibody-related therapeutics undergoing clinical trials, or those already approved, for the treatment of severe type 2 asthma. Biologics targeting type 2 cytokines have shown consistent efficacy, especially in patients with evidence of type 2 inflammation, suggesting that the future of asthma biologics is promising

Lactobacillli expressing llama VHH fragments neutralise Lactococcus phages

<p>Abstract</p> <p>Background</p> <p>Bacteriophages infecting lactic acid bacteria (LAB) are widely acknowledged as the main cause of milk fermentation failures. In this study, we describe the surface-expression as well as the secretion of two functional llama heavy-chain antibody fragments, one binding to the major capsid protein (MCP) and the other to the receptor-binding proteins (RBP) of the lactococcal bacteriophage p2, by lactobacilli in order to neutralise lactococcal phages.</p> <p>Results</p> <p>The antibody fragment VHH5 that is directed against the RBP, was fused to a c-<it>myc </it>tag and expressed in a secreted form by a <it>Lactobacillus </it>strain. The fragment VHH2 that is binding to the MCP, was fused to an E-tag and anchored on the surface of the lactobacilli. Surface expression of VHH2 was confirmed by flow cytometry using an anti-E-tag antibody. Efficient binding of both the VHH2 and the secreted VHH5 fragment to the phage antigens was shown in ELISA. Scanning electron microscopy showed that lactobacilli expressing VHH2 anchored at their surface were able to bind lactococcal phages. A neutralisation assay also confirmed that the secreted VHH5 and the anchored VHH2 fragments prevented the adsorption of lactococcal phages to their host cells.</p> <p>Conclusion</p> <p>Lactobacilli were able to express functional VHH fragments in both a secreted and a cell surface form and reduced phage infection of lactococcal cells. Lactobacilli expressing llama heavy-chain antibody fragments represent a novel way to limit phage infection.</p

The in vivo phosphorylation sites of bovine αB-crystallin

AbstractPhosphate content determinations established that in αB-crystallin two phosphate groups can be present in vivo in bovine lenses. Comparison of tryptic digests of phosphorylated and unphosphorylated αB chains, revealed the location of the two phosphorylation sites in tryptic peptides T2 and T3. Thermolytic digestion and gas-phase sequencing demonstrated that Ser-19 and Ser-45 are the in vivo phosphorylation sites of bovine αB-crystallin. This pattern of phosphorylation differs from the previously reported in vitro obtained results

A bispecific antibody strategy to target multiple type 2 cytokines in asthma

Background: Asthma is a chronic inflammatory airway disease in which innate and adaptive immune cells act together to cause eosinophilic inflammation, goblet cell metaplasia (GCM), and bronchial hyperreactivity (BHR). In clinical trials using biologicals against IL-4 receptor (IL-4R) alpha or IL-5, only a subset of patients with moderate-to-severe asthma responded favorably, suggesting that distinct pathophysiologic mechanisms are at play in subgroups of patients called endotypes. However, the effect of multiple cytokine blockade using bispecific antibodies has not been tested. Objective: We sought to target simultaneously the IL-4, IL-13, and IL-5 signaling pathways with a novel IL-4R alpha/IL-5-bispecific antibody in a murine house dust mite (HDM) model of asthma. Methods: Two mAbs neutralizing IL-4R alpha and IL-5 were generated by using a llama-based antibody platform. Their heavy and light chains were then cotransfected in mammalian cells, resulting in a heterogeneous antibody mixture from which the bispecific antibody was isolated by using a dual anti-idiotypic purification process. C57BL/6J mice were finally sensitized and challenged to HDM extracts and treated during challenge with the antibodies. Results: We successfully generated and characterized the monospecific and bispecific antibodies targeting IL-4R alpha and IL-5. The monospecific antibodies could suppress eosinophilia, IgE synthesis, or both, whereas only the IL-4R alpha/IL-5-bispecific antibody and the combination of monospecific antibodies additionally inhibited GCM and BHR. Conclusion: Type 2 cytokines act synergistically to cause GCM and BHR in HDM-exposed mice. These preclinical results show the feasibility of generating bispecific antibodies that target multiple cytokine signaling pathways as superior inhibitors of asthma features, including the difficult-to-treat GCM

Dual anti-idiotypic purification of a novel, native-format biparatopic anti-MET antibody with improved in vitro and in vivo efficacy

Bispecific antibodies are of great interest due to their ability to simultaneously bind and engage different antigens or epitopes. Nevertheless, it remains a challenge to assemble, produce and/or purify them. Here we present an innovative dual anti-idiotypic purification process, which provides pure bispecific antibodies with native immunoglobulin format. Using this approach, a biparatopic IgG1 antibody targeting two distinct, HGF-competing, non-overlapping epitopes on the extracellular region of the MET receptor, was purified with camelid single-domain antibody fragments that bind specifically to the correct heavy chain/light chain pairings of each arm. The purity and functionality of the anti-MET biparatopic antibody was then confirmed by mass spectrometry and binding experiments, demonstrating its ability to simultaneously target the two epitopes recognized by the parental monoclonal antibodies. The improved MET-inhibitory activity of the biparatopic antibody compared to the parental monoclonal antibodies, was finally corroborated in cell-based assays and more importantly in a tumor xenograft mouse model. In conclusion, this approach is fast and specific, broadly applicable and results in the isolation of a pure, novel and native-format anti-MET biparatopic antibody that shows superior biological activity over the parental monospecific antibodies both in vitro and in vivo