[Recombinant granulocyte-colony stimulating factor (filgrastim): optimization of conjugation with polyethylene glycol].

In order to create an active pharmaceutical substance of the drug with prolonged action the modification of recombinant human granulocyte colony-stimulating factor GCSF (filgrastim) with polyethylene glycol (PEG, M 21.5 kDa) was conducted. A method for preparation of PEG-filgrastim designed for the development and scaling-up of the technological process of production was described.

Ternary interpolyelectrolyte complexes insulin-poly(methylaminophosphazene)-dextran sulfate for oral delivery of insulin.

Ternary interpolyelectrolyte complexes of insulin with biodegradable synthetic cationic polymer, poly(methylaminophosphazene) hydrochloride (PMAP), and dextran sulfate (DS) were investigated by means of turbidimetry, dynamic light scattering, phase analysis, and high-sensitivity differential scanning calorimetry. Formation of ternary insoluble stoichiometric Insulin-PMAP-DS complexes was detected under conditions imitating the human gastric environment (pH 2, 0.15 M NaCl).

[DNA assay for genetically engineered pharmaceutical substances using the real-time PCR technique].

A real-time PCR procedure is proposed for assaying E. coli residual DNA in the pharmaceutical substance of human recombinant insulin. For the quantitative analysis of the DNA content, an amplification of fragments of the bla gene plasmid DNA and E.

[Optimization of the industrial production of the recombinant precursor of human insulin].

Conditions were found at the analytical level for the solubilization of a recombinant insulin precursor from inclusion bodies in different buffer systems at a wide pH range in the presence of different reducing (dithiothreitol, dithioerythritol) and chaotropic agents (urea, guanidine hydrochloride) and the subsequent renaturation with the use of redox pairs (cysteine-cystine, oxidized glutathione-reduced glutathione, and others). The scaling of the method for the production of the active substance of genetically engineered human insulin has been performed.

[Nanocomplexes of recombinant proteins and polysialic acid: preparation, characteristics, and biological activity].

A preparation of nanocomplexes containing recombinant proteins (interferons alpha2b and beta1b, insulin, and human granulocyte colony stimulating factor) and natural polysialic acid (PSA) has been described. The incorporation of protein into the complex changes its electrophoretic mobility. Atomic force microscopy reveals the average size of 23-kD insulin complexes with PSA of 10-20 nm and demonstrates that more than 60% of glycopolymer molecules carry a single protein molecule.

[Modification of recombinant proteins by covalent polysialation illustrated with the example of human insulin].

Methods of selective and nonselective covalent immobilization of genetically engineered proteins on molecules of natural polysialic acid are described by the example of human insulin. Such modification increases insulin lifetime in vivo.

[Validation of a method for monitoring of genetically engineered human insulin manufacture].

A method for monitoring the manufacture of genetically engineered human insulin by HPLC was developed. The method was validated by the estimation of its linearity, correctness, accuracy, specificity, and stability; the limits of detection and quantitative assessment were also determined. It was proven that HPLC analysis enables reliable and reproducible results to be obtained and can be used for monitoring insulin manufacture.

Displacement effect during HPLC preparative purification of human insulin.

HPLC plays a key role in the preparative purification of human insulin. A21-desamidoinsulin is one of the impurities that possesses the chromatographic behavior similar to that of insulin and hence separation from this by-product is rather difficult at the process scale. During the optimization of insulin reversed-phase HPLC purification, when a column was sufficiently overloaded, the effect of displacement of A21-desamidoinsulin molecules from active groups of sorbent by insulin ones was observed.

Russia through the prism of the world biopharmaceutical market.

Trends in the Russian pharmaceutical biotechnology and related fields representing the major sector of domestic biotech are reviewed through the prism of the world biopharmaceuticals market. A special emphasis is placed on biogenerics and follow-on biologics. The revival of national pharmbiotech is seen in close cooperation between private companies and the state, academia and industry.

[Prospects for designing Russian gene engineering agents for medicine. Rastan is the first Russian recombinant human growth hormone].

The development of modern pharmacology cannot be imagined without the use of genetic engineering methods (recombinant DNA technology). The success of medicine is increasingly based on the active use of protein preparations obtained using the technology of transferring hereditary information (genes) from one organism to another. The emergence of the ability to express foreign genes in the cells of various organisms (both eukaryotes and prokaryotes) has become one of the revolutionary events in the science of the last two decades of the 20th century and laid the foundations of modern biotechnology.

[A PCR-based semiquantitative assay of DNA impurities in recombinant protein preparations].

A semiquantitative assay of DNA impurities in preparations of human recombinant insulin is described. The assay is based on the detection of a fragment of the ampicillin-resistant gene within the producer strain DNA by PCR. The analysis of PCR products of the studied preparations and PCR products containing known amounts of E.

[The histamine-liberating action of polymyxin B and its analogs].

Histamine-releasing effect of polymyxin B1 and its deacylated analogues has been studied on purified rat mast cells. The structure-activity analysis showed that cyclic peptide fragment and acyl residue of molecule of polymyxin plays an important role in histamine-releasing activity. Histamine release, induced by polymyxin B1 and its analogue was blocked by metabolic inhibitor antimycin A.

[Structural and functional investigation of polymyxins. Structure and biological properties of polymyxin M analogs].

The effect of proteinases of plant and microbial origin on polymyxin M was studied. It was shown that this antibiotic was absolutely stable to the effect of papain and ficin. On hydrolysis with subtilisin there formed polymyxin decyclized analogs not described earlier.

[The action of combinations of the nonapeptide polymyxin B with antibiotics on gram-negative bacteria].

Activity of polymyxin B nonapeptide alone and in combination with other antibiotics against clinical strains of Pseudomonas and enteric bacteria was studied. It was shown that nonapeptide was highly active against Pseudomonas and moderately active against enteric bacteria. In combination with rifampicin, fusidic acid or erythromycin the nonapeptide had a potentiating effect on the tested strains.

[Structural and functional study of polymyxins. Isolation and properties of individual components of polymyxin M].

LPCC and HPLC revealed that polymyxin M was a mixture of five components of the polymyxin nature: PM1, PM2, PMx, PMy and PMz. The individual compounds PM1, PM2 and PMz were isolated. Their physicochemical properties and data on antimicrobial activity are presented.

[Structural and functional research on polymyxins. 1H NMR analysis of the conformation of polymyxin M in water].

Spatial structure of polypeptide antibiotic polymyxin M in water was studied by one-and two-dimensional (COSY, COSY-45, RELAY) H NMR spectroscopy. Analysis of the signal spectral parameters revealed two intramolecular hydrogen bonds in the cyclic part of the molecule which was analogous to the structure of polymyxin B. However, configuration of both the beta-turns in the polymyxin M structure differed from that of the detected earlier beta-turns in the structure of polymyxin B.

[Structural-functional research on polymyxins. 1H-NMR spectra of polymyxin B and its shortened analog].

Polymyxin B and its shortened analog were studied comparatively by 1H-NMR spectroscopy. Analysis of the signal chemical shifts, constants of spin-spin interaction of 3J HN-C alpha H and temperature coefficients of the NH signal chemical shifts revealed absolute structural identity of both molecules cyclic parts. This proved that there was no conformative interaction between the cyclic and linear parts of the polymyxin B molecule.