MIL-100(Fe)-Based Composite Films for Food Packaging.

A biocompatible metal-organic framework MIL-100(Fe) loaded with the active compounds of tea tree essential oil was used to produce composite films based on κ-carrageenan and hydroxypropyl methylcellulose with the uniform distribution of the particles of this filler. The composite films featured great UV-blocking properties, good water vapor permeability, and modest antibacterial activity against both Gram-negative and Gram-positive bacteria. The use of metal-organic frameworks as containers of hydrophobic molecules of natural active compounds makes the composites made from naturally occurring hydrocolloids attractive materials for active packaging of food products.

Targeted Cytokine Delivery for Cancer Treatment: Engineering and Biological Effects.

Anti-tumor properties of several cytokines have already been investigated in multiple experiments and clinical trials. However, those studies evidenced substantial toxicities, even at low cytokine doses, and the lack of tumor specificity. These factors significantly limit clinical applications. Due to their high specificity and affinity, tumor-specific monoclonal antibodies or their antigen-binding fragments are capable of delivering fused cytokines to tumors and, therefore, of decreasing the number and severity of side effects, as well as of enhancing the therapeutic index. The present review surveys the actual antibody-cytokine fusion protein (immunocytokine) formats, their targets, mechanisms of action, and anti-tumor and other biological effects. Special attention is paid to the formats designed to prevent the off-target cytokine-receptor interactions, potentially inducing side effects. Here, we describe preclinical and clinical data and the efficacy of the antibody-mediated cytokine delivery approach, either as a single therapy or in combination with other agents.

Universal Character of Breaking of Wormlike Surfactant Micelles by Additives of Different Hydrophobicity.

Wormlike surfactant micelles are widely used in various applications including fracturing technology in oil industry, template synthesis of different nanoobjects, micellar copolymerization of hydrophilic and hydrophobic monomers, and so forth. Most of those applications suggest the solubilization of different additives in the micelles. The present paper is aimed at the comparative study of the effect of the solubilization of hydrophobic (n-decane and 1-phenylhexane) and hydrophilic (N-isopropylacrylamide and acrylamide) substances on the rheological properties and structure of the micelles using several complementary techniques including rheometry, small angle neutron scattering, dynamic light scattering, and diffusion ordered NMR spectroscopy. For these studies, mixed micelles of potassium oleate and n-octyltrimethylammonium bromide containing the excess of either anionic or cationic surfactants were used. It was shown that hydrophobic additives are completely solubilized inside the micelles being localized deep in the core (n-decane, 1-phenylhexane) or near the core/corona interface (1-phenylhexane). At the same time, only a small fraction of hydrophilic additives (14% of N-isopropylacrylamide and 4% of acrylamide) penetrate the micelles being localized at the corona area. Despite different localization of the additives inside the micelles, all of them induce the breaking of wormlike micelles with the formation of either ellipsoidal microemulsion droplets (in the case of hydrophobic additives) or ellipsoidal surfactant micelles (in the case of hydrophilic additives). The breaking of micelles results in the drop of viscosity of the solution up to water value. The main result of this paper consists in the observation of the fact that for all the additives under study, the dependences of the viscosity on the volume fraction of additive lie on the same master curve being shifted along the volume fraction axis by a certain factor depending on the hydrophobicity of the added species. Those data are quite useful for various applications of wormlike surfactant micelles suggesting the solubilization of different additives inside them.

What are the prospects for using complexes of copper(ii) and zinc(ii) to suppress the vital activity of Mycolicibacterium smegmatis?

New complexes of zinc(ii) and copper(ii) with 2-furoic acid (Hfur), acetic acids and N-donor ligands with the compositions [Zn2(fur)4] n (1), [Zn2(fur)4(NH2py)2] (2, NH2py = 3-aminopyridine), [Zn(fur)2(neoc)] (3, neoc = 2,9-dimethyl-1,10-phenantroline), [Zn(OAc)2(neoc)] (4, OAc = acetat-anion), and [Cu(fur)2(neoc)(H2O)] (5) were synthesized. The structures of the compounds were established by single crystal X-ray diffraction analysis. Complexes 1 and 2 are binuclear; whereas 3-5 are mononuclear. The stabilization of supramolecular architectures in crystals for compounds 1-5 occurs due to π-π-bonding between heterocycles and hydrogen interactions that provide good solubility in aqueous solutions. The stability of the complexes upon dissolution in 5% dextrose and 0.9% NaCl was confirmed by UV-vis spectroscopic and NMR (1H) data. The study of in vitro biological activity was carried out against the non-pathogenic strain of Mycolicibacterium smegmatis that is a model for M. tuberculosis. The synergistic effect of ligands is observed for complexes 3-5 and is characterized by an increase in the biological activity values. On passage from Zn2+ to Cu2+ complexes, the biological activity increases and the maximum effect is observed for compound [Cu(fur)2(phen)]. Analysis of the transcriptomic profiles of the M. smegmatis mc 2 155 strain under the pressure of the copper complex [Cu(fur)2(phen)] made it possible to isolate 185 genes, one quarter of which are associated with the compensation of iron deficiency in the bacterial strain. Genes associated with the transport and metabolism of heavy metals, biosynthesis of fatty and amino acids, biodegradation and transport of urea were also isolated.

Features, modulation and analysis of glycosylation patterns of therapeutic recombinant immunoglobulin A.

Increasing the production of recombinant antibodies while ensuring high and stable protein quality remains a challenge in mammalian cell culture. This review is devoted to advances in the field of obtaining stable and optimal glycosylation of therapeutic antibodies based on IgA, as well as the subsequent issues of glycosylation control of glycoproteins during their production. Current studies also demonstrate a general need for a more fundamental understanding of the use of CHO cell-based producer cell lines, through which the glycoprofile of therapeutic IgA antibodies is produced and the dependence of glycosylation on culture conditions could be controlled. Optimization of glycosylation improves the therapeutic efficacy and can expand the possibilities for the creation of highly effective glycoprotein therapeutic drugs. Current status and trends in glycan analysis of therapeutic IgA, dominantly based on mass spectrometry and lectin microarrays are herein summarised as well.

Detrimental Effects of IFN-γ on an Epidermolysis Bullosa Simplex Cell Model and Protection by a Humanized Anti-IFN-γ Monoclonal Antibody.

Epidermolysis bullosa is a group of severe skin blistering disorders, which currently have no cure. The pathology of epidermolysis bullosa is recognized as having an inflammatory component, but the role of inflammation in different epidermolysis bullosa disorders is unclear. Epidermolysis bullosa simplex (EBS) is primarily caused by sequence variants in keratin genes; its most severe form, EBS generalized severe, is characterized by aggregates of keratin proteins, and cell models of EBS generalized severe show constitutively elevated stress. IFN-γ is a major mediator of inflammation, and we show that the addition of IFN-γ alone to disease model keratinocytes promotes keratin aggregation, decreases cell-cell junctions, delays wound closure, and reduces cell proliferation. IFN-γ exposure weakens the intercellular cohesion of monolayers on mechanical stress, with IFN-γ-treated EBS monolayers more fragmented than IFN-γ-treated wild-type monolayers. A humanized monoclonal antibody to IFN-γ neutralized the detrimental effects on keratinocytes, restoring cell proliferation, increasing cell-cell adhesion, accelerating wound closure in the presence of IFN-γ, and reducing IFN-γ-mediated keratin aggregation in EBS cells. These suggest that treatment with IFN-γ blocking antibodies may constitute a promising new therapeutic strategy for patients with EBS and may also have ameliorating effects on other inflammatory skin diseases.

Bispecific Antibodies for IFN-ß Delivery to ErbB2+ Tumors.

The main aim of our work was to create a full-length bispecific antibody (BsAb) as a vehicle for the targeted delivery of interferon-beta (IFN-ß) to ErbB2+ tumor cells in the form of non-covalent complex of BsAb and IFN-ß. Such a construct is a CrossMab-type BsAb, consisting of an ErbB2-recognizing trastuzumab moiety, a part of chimeric antibody to IFN-ß, and human IgG1 Fc domain carrying knob-into-hole amino acid substitutions necessary for the proper assembly of bispecific molecules. The IFN-ß- recognizing arm of BsAb not only forms a complex with the cytokine but neutralizes its activity, thus providing a mechanism to avoid the side effects of the systemic action of IFN-ß by blocking IFN-ß Interaction with cell receptors in the process of cytokine delivery to tumor sites. Enzyme sandwich immunoassay confirmed the ability of BsAb to bind to human IFN-ß comparable to that of the parental chimeric mAb. The BsAb binds to the recombinant ErbB2 receptor, as well as to lysates of ErbB2+ tumor cell lines. The inhibition of the antiproliferative effect of IFN-ß by BsAb (IC50 = 49,3 µg/mL) was demonstrated on the HT29 cell line. It can be proposed that the BsAb obtained can serve as a component of the immunocytokine complex for the delivery of IFN-ß to ErbB2-associated tumor cells.

Photochemical synthesis, intercalation with DNA and antitumor evaluation in vitro of benzo[d]thiazolo[3,2-a]quinolin-10-ium derivatives.

A new anticancer benzo[d]thiazolo[3,2-a]quinolin-10-ium derivatives were synthesized and characterized. Anticancer evaluation in vitro against four cancer cell lines including adenocarcinomic human alveolar basal epithelial cells (A549), hepatocellular carcinoma (HepG2), prostate cancer (PC3) and breast cancer (MCF7) indicated that some of prepared compounds shows higher selectivity in comparison with doxorubicin. DNA interaction studies by optical, CD, NMR spectroscopies showed the high affinity of benzothiazole ligands towards the dsDNA. The ligand-DNA interaction occurs through the intercalation of benzo[d]thiazolo[3,2-a]quinolin-10-ium derivatives with nucleic acid. The investigation of formed ligand - DNA complexes by docking and molecular dynamic calculations was applied for analysis of the relationship between structure and anticancer activity. The results suggested that benzo[d]thiazolo[3,2-a]quinolin-10-ium derivatives might serve as a novel scaffold for the future development to new antitumor agents.

Synthesis, Structure and Electrochemical Properties of Acetamide- and Caprolactam-Containing Silicon Catecholates.

Hexacoordinated heteroligand silicon catecholates, although being prospective as easily soluble compounds with high hydrolytic stability and diverse redox properties, have been insufficiently studied. The transesterification of 1-(trimethoxysilylmethyl)-2-oxohexahydroaze or N-methyl-N-(trimethoxysilylmethyl)acetamide by two equivalents of catechol derivatives in the presence of dicyclohexylamine afforded a series of target compounds in good yield. The complexes were characterized using elemental analysis, FTIR, 1H, 13C and 29Si NMR spectra, X-ray crystallography and cyclic voltammetry. X-ray diffraction confirmed that the silicon atom possesses the octahedral geometry of the SiCO5 polyhedron that remains unchanged in solution as it follows from 29Si NMR data. The compounds demonstrated up to three oxidation waves; and the reduction profile strongly depended on the nature of the substituents on a catecholate anion.

Influence of media composition on recombinant monoclonal IgA1 glycosylation analysed by lectin-based protein microarray and MALDI-MS.

Glycosylation of therapeutic glycoproteins significantly affects their physico-chemical properties, bioactivity and immunogenicity. The determination of glycan composition is highly important regarding their development and production. Therefore, there is a demand for analytical techniques enabling rapid and reliable glycoprofiling of therapeutic proteins. For the investigation of changes in glycan structures, we have employed two platforms: lectin-based protein microarray, and MALDI-MS. In lectin-based microarray analysis, the samples of IgA were printed on the microarray slide, incubated with the set of lectins with various specificity and evaluation of changes in glycosylation was based on differences in reactivity of samples with lectins. MALDI-MS was used for N-glycan analysis of IgA1 samples. IgAs are effective as therapeutic agents in defense against viruses that use sialic acid as a receptor. Dimeric IgA1 antibodies were produced by stable cell line IgA1/2G9 on the basal medium at different conditions (different supplementation and feeding) and we also evaluated the effect of different conditions on lactate production, which correlates with IgA productivity. Decrease of lactate levels was observed during supplementation with succinic acid, asparagine, or with mannose feeding. We found by lectin-based microarray analysis that the metabolic shift from glutamine to asparagine or feeding with glucose caused increase of high mannose type glycans what was confirmed also by MALDI-MS. Among other changes in IgA glycosylation determined by lectin-based protein microarray were, for example, reduced galactosylation after supplementation with succinic acid and increase of both sialylation and galactosylation after supplementation with glutamine and feeding with mannose. The elucidation of mechanism of determined changes requires further investigation, but the described analytical approach represent effective platform for determination, screening and evaluation of glycosylation of therapeutic proteins.

Effects of Succinic Acid Supplementation on Stable Cell Line Growth, Aggregation, and IgG and IgA Production.

BACKGROUND: Immunoglobulin (Ig) G is the most commonly used therapeutic antibodies. Recently, the interest in IgA antibodies to treat respiratory infectious diseases has been increasing. The reason for the inefficient use of IgA is recombinant antibody aggregation in cell culture, affecting the longevity and productivity of cell lines. Lactate is an important metabolite that affects the cultivation of stable cell lines producing monoclonal antibodies. METHODS: In the present study, we investigated whether different combinations of succinic acid and micro-additives affect lactate production, which correlates with productivity. The effect of succinic acid substitution on productivity of cells producing IgG/IgA was analyzed using the static culture method in a six-well plate. Lactate was measured in supernatant of cell culture indirectly by using the activity of Lactate Dehydrogenase (LDH).A low lactate level was observed in cultivation medium supplemented with succinic acid or asparagine combined with some inorganic salts. RESULTS: The results also demonstrated the effect of component supplementation on homogeneity, longevity, and productivity of cell culture. Supplementation of succinic acid eliminated cell aggregation and improved homogeneity of stable cell lines producing IgG and, especially, IgA. CONCLUSION: Overall, succinic acid supplementation to the culture medium has potential biotechnological applications in the production IgG and IgA.

Production of anti TNF-α antibodies in eukaryotic cells using different combinations of vectors carrying heavy and light chains.

Tumor necrosis factor-α (TNF-α) plays a key role in rheumatoid arthritis and some other autoimmune diseases. Therapy with anti-TNF-α recombinant antibodies (Ab) appears to be highly effective. Production of new hyper-producing eukaryotic cell lines can decrease the treatment cost, which currently is very high. However, due to the complexity of protein transcription, translation, processing, and secretion in mammalian cells, the stages at which antibody expression is affected are still poorly determined. The aim of this work was to compare the productivity of two cell lines developed in CHO DG44 cells, deficient in dihydrofolate reductase, transfected with vectors carrying either heavy (H) or light (L) chains of chimeric antibody under different combinations of selective elements. Both H and L chains were cloned either in pOptiVEC or pcDNA3.3 vectors and different combinations were used to produce HL and LH cell lines. We have shown that Ab production has been low and comparable between HL and LH cells until selection on methotrexate (MTX) when LH but not HL cells have responded with 3.5 times increased productivity. Flow cytometry analysis has demonstrated that intracellular concentration of full size Abs in LH cells was 5.6 times higher than in HL ones due to higher amount of H chain synthesis. No differences in viability between HL and LH cells have been found. We have concluded that the expression of H chain in the pOptiVEC vector, which is responsible for MTX resistance, has led to the suppression of H chain synthesis and limitation in full Ab assembly.