Homo- and hetero-polyamino acid fullerene c60 derivatives, method for preparing them and based pharmaceutical compositions

FIELD: medicine, pharmaceutics.

SUBSTANCE: invention refers to homo- and hetero-polyaminoacid fullerene derivatives of formula C60(H)x{NH(CH2)nCOO-}x{NH3+(L)COOH}x, wherein n - 2-5, x - 3, L = -(CH2)m, wherein m = 1-5, or -CO(CH2)kCH(NH2)-, wherein k = 1-2, which show activity against herpes virus, hepatitis C virus, influenza viruses of various nature, HIV, as well as anticancer and antipsoriatic activity.

EFFECT: developing the method for preparing said homo- and hetero-polyaminoacid fullerene derivatives and based pharmaceutical compositions.

7 cl, 19 tbl, 8 ex

 

The invention relates to pharmaceutical industry and medicine, namely to new Homo - and hetero-poliomyelitis derivative of fullerene C60formula (I), and the method of their production and creation of the pharmaceutical compositions based on them.

where n=2-5, x=3, L=-(CH2)mor-CO(CH2)kCH(NH2)-, m=2-5, where k=1-2.

The use of fullerene derivatives in medicine is based on the lipophilic properties of fullerene core, allowing fullerene derivative to penetrate through cell membranes, and the ability of the fullerene with high quantum yield to generate singlet oxygen that breaks down DNA. These properties provide a functional derivative of fullerene manifestation of cytotoxic, antiviral and other properties (D. Bedrov, G.D. Smith, Davande H., "Passive transport of fullerenes through a lipid membrane." J. Phys. Chem., B, 2008, v.112., p.2078-84, Qiao, R., Roberts, A.E., "Translocation of fullerene and its derivatives across a lipid bilayer", Nano Lett., 2007, v.7, p.614-9. Nelsen G.D., and others, "In vivo biology and toxicology of fullerenes and their derivatives". Basic and Clinical Pharmacology and Toxicology, 2008, v.103, p.197-208, Pat. US 6204391, 2005, "Water soluble fullerenes with antiviral activity").

The main problem impeding biological studies of derivatives of fullerenes and creating therapeutic drugs based on them, is the insolubility of fullerenes in water, which makes their direct introduction into an organism of the man. One of the possibilities is x options for overcoming these challenges is the introduction of fullerene molecules in solubilizing the matrix. Known methods for producing water-soluble fullerene due to the formation of adduct with polyvinylpyrrolidone (Kiselev O.I. et al. // Mol. Materials. 1998. V.11. P.121; discovered on L.B. et al. // Ibidem. 2000. V.13. P.41). Shown its effectiveness against influenza a - and b-type.

There is also known a method of producing fullerenes, comprising mixing a pre-dissolved in an organic solvent fullerene polymer matrix in chloroform, evaporation of the mixture under vacuum to remove the solvent, dissolving the obtained complex in phosphate-buffered saline (pH 7.4 and 7.6) followed by treatment of the product by ultrasound (RU # 2162819, 02.10.01). At the same time as the water-soluble polymer matrix with the use of membrane Catalina. The products resulting from such modifications are unstable compositions with disabilities storage.

A promising method of obtaining water-soluble fullerene compositions is chemical modification of fullerene sphere by introducing hydrophilic solubilizing ligands. In the international application WO 2005/070827 presents a series of amino acid derivatives of fullerene obtained by the cycloaddition reaction of amino acid fragments to the fullerene, and the products of their implementation in biologically-active organic substrates. Presented in this technical solution, IU the odes of multistage synthesis and ethnologica. The compounds have a low solubility in water.

At the present time has received a large number of functionalized fullerenes containing hydrophilic fragments in the side chain attached to the fullerene ligands (detergent type complexes), and spherical type derivatives when there are polar groups, distributed fullerene sphere (this type includes fullerenol, eminiaddict).

The most promising for use are amino acid derivatives of fullerene.

Analogs of the present invention are compounds and methods for their preparation, are described in international application WO 2009/00203, as well as the patent of Russian Federation №2236852.

In the international application WO 2009/00203 described polyfunctional amino acid derivatives of fullerene formulawhere R=H, mono - or dihydroxyethyl, haloalkyl, mono - or dinitrocresol, maleinimide; N-Z is a fragment of α, β, γ, ω-amino acids of General formula-NH-CmH2m-COOM or C4H8N-COOM, where m=2-5 and M represents nitrocellulous group, alkyl group, or a salt of an alkali metal, or a dipeptide. These compounds obtained by the reaction of equimolar attach amino acids to the fullerene with the subsequent replacement of active hydrogen organic biologically active ligada education with the joining type. The compounds have inhibitory activity against tumor metastasis, increase antileykemichesky activity of cyclophosphamide, and may be suitable as a donor of nitrogen monoxide or as vasodilators quick steps to antigipertenzivnoe therapy.

The main disadvantage of the compounds of this application is that they are products of covalent joining contain a small amount of polar groups and have a low solubility in water.

The closest in technical essence and the achieved result is a means for inhibiting the reproduction enveloped viruses and method thereof (patent RF №2236852). As a result of interaction of fullerene with the salt of the amino acids in the medium of organic solvent in the presence of polyalkylated received fullerenelike anions of General formula

C60Hn[NH(CH2)mC(O)O-]n,

where C60- fullerene core, NH(CH2)mC(O)O-- aminocarbonyl anion; m is an integer from 1 to 5, p is an integer from 2 to 12.

To obtain these compounds to the solution of fullerene in o-dichlorobenzene (toluene or other organic solvent) contribute the amino acid in the form of salts (potassium or sodium), then add a solubilizer. The order of entry in actionnow Wednesday amino acids and a solubilizer is not important, you can make them as complex, pre-mixing. As a solubilizer use different polyalkyloxy: glycols mol. mass of from 150 to 400, and above 400 (e.g., PEG-1500), and dimethyl ether of polyethylene glycol (mol. mass 500. To increase the rate of reaction add any strong reducing agent (alkali metals).

The ratio of the fullerene and amino acids increased more than 50 times. The transformation into the desired pharmaceutically acceptable salt, especially sodium or potassium, was carried out by treating the acid with a suitable base or by adding a salt of the weak volatile acid. In particular, it is not soluble in water fullerenelike acid into the preferred pharmaceutically acceptable salts such as sodium salt, which is soluble in water. The salt of the weak volatile acid by treatment of a solution of a salt of an alkali metal and a weak volatile acid. When the concentration of the solution by evaporation or lyophilization of a weak acid is removed and fullerenelike acid are in the form of their alkali metal salts. Target product according to this invention is characterized by the constancy of the composition, the content of the target product of the basic substance is only of 87.8%. In the description there are no technological regulations, is knitted with the determination of the optimum amounts of the parent compounds, ratios of quantities of the used solvents and, especially, descriptions of methods for the isolation of the desired compounds.

The main disadvantages of vulernability derivatives obtained are presented in patent method is that this way we obtain a mixture tolerancerelated anions, as salt and acid forms. To get a individual connection with a method described in the patent, is not possible. The fullerene-polyaminoamide obtained by the patented process, in acid form, practically insoluble in water. To get a stable pharmaceutical composition with fulleropyrrolidine anions was not possible, because in the process of storage compounds precipitate.

Application in the synthesis of a large excess of potassium or sodium salts of amino acids and a large excess of solvent leads to environmental problems associated with disposal of waste production and increase the cost of the production process. The use of alkali metals to increase the rate of reaction is technologically impossible when using chlorinated aromatic solvents.

However, presented in the patent the unique biochemical properties of drugs based on fullerene compounds with amino acid fragments are aiming for the teaching of new fullerene derivatives, development of high-tech way of their industrial production, which is simple and efficient process, purity, eco-friendliness and availability of the raw materials.

To solve this problem is proposed group of inventions combined to form a single inventive concept, namely Homo - and hetero-polyamidation fullerene derivative, the method of obtaining fullerene derivatives and pharmaceutical compositions comprising Homo - and hetero-polyamidation fullerene derivative as an active ingredient. By varying the ratio of reactants and process conditions, the present method it is possible to obtain various derivatives of fullerene.

This problem is solved Homo - and hetero-polinenasyschennami fullerene derivatives of General formula (II):

In the case when m=n receive Homo-polyaminoamide derivatives of fullerene, when m≠n - hetero-polyamidation fullerene derivative.

This task is solved by a method of obtaining Homo - and hetero-polyamidation fullerene derivatives formed by the interaction of fullerene with a 10-fold molar excess of anhydrous potassium salts of amino acids in the environment of aromatic organic solvent when added to the resulting suspension interphase catalyst under stirring and is the agrevanie to a temperature not above 60°C until complete discoloration of the solution and formation of a solid residue, then exhale, dissolved in water, followed by treatment of aqueous solutions of potassium salts of fullerenealuminum 1N solution of an organic or mineral acid followed by the addition of amino acid solutions in polar solvents. In the method using freshly prepared anhydrous potassium salts of amino acids in fine condition, and the selection of sediment potassium salts fullerenealuminum carried out by filtration, washing with ethanol and drying. During experiments it was found that fullerenealuminum specified structure can be obtained only when using a freshly prepared anhydrous potassium salts of amino acids. As phase transfer catalyst is used, the methyl ethers of polyethylene oxides of molecular weight 200, 400, 500.

This task is solved by the creation of the pharmaceutical compositions containing as active substance Homo - and hetero-polyamidation fullerene derivative of the formula (II)having activity against herpes virus, hepatitis C virus, influenza viruses of different nature, HIV and antitumor and protivopsoriaticescoe activity. The pharmaceutical compositions can be in the form of tablets, capsules, ointments, emulsions, suppositories, solutions, sprays.

F the pharmaceutical compositions according to the proposed technical solution containing the compound of General formula (II), effective to achieve the desired result, and can be entered as standard medicinal forms (for example, in solid, semisolid, or liquid form), containing compounds of the proposed technical solution as an active ingredient in a mixture with a carrier or excipient suitable for intramuscular, intravenous, oral, sublingual, inhalation, local, nasal, rectal, and vaginal administration. The active ingredient can be included in a composition together with commonly used non-toxic pharmaceutically acceptable carriers suitable for the manufacture of solutions, tablets, pills, capsules, pills, suppositories, emulsions, suspensions, ointments, gels or any other medicines.

A particular level of dosage and frequency of medication for each individual patient will depend on a number of factors, including the activity of a specific derivative of fullerene, the metabolic stability and length of action, allocation rate, the patient's age, body weight, General health, sex, drug combination and the severity of the disease in the individual being treated.

For oral administration in the form of a suspension compositions prepared according to methods widely known in the field of pharmaceutical preparation of prescriptions is ur and they may contain microcrystalline cellulose or its derivatives to ensure mass, alginic acid or sodium alginate as a suspending agent, methylcellulose as an amplifier viscosity and sweetening agents and/or fragrances, known in this area. In the form of tablets such compositions may contain microcrystalline cellulose, calcium phosphate, starch, magnesium stearate and lactose and/or other excipients, binders, extenders, disintegrators, diluents and lubricants known in this field.

When applied as nasal aerosols or by inhalation, such compositions are prepared by methods well known in the field of pharmaceutical formulations, and they can be produced as solutions in saline, using benzoic acid or other suitable preservatives, promoters adsorption to enhance bioremediate, and/or other solubilizing or dispersing agents known in this field.

Solutions or suspensions for injection can be formulated according to known methods, using non-toxic, parenterally applicable diluents or solvents, such as mannitol, 1,3-butanediol, water, ringer's solution or isotonic sodium chloride solution or suitable dispersing or is macevoy and suspendida agents, such as sterile, soft, sustainable oils, including synthetic mono - or diglycerides or fatty acids, including oleic acid.

When rectal or vaginal application in the form of candles such compositions can be prepared by mixing the drug with such a non-irritating excipients as cocoa butter, synthetic glyceride esters or polyethylene glycols, which are solid at ordinary temperatures, but sigalda and/or dissolve in the body cavity with the release of the medication.

The local application in the form of ointments, gels, creams, liniments, etc. such compositions can be prepared by mixing the active ingredients with acceptable ointment base.

As ointment bases can be used fat, hydrocarbon or hydrophilic bases such as vaseline, vaseline oil, paraffin wax, lanolin, polyethylene glycol, etc.

As the basis for gels can be used methyl cellulose, sodium carboxymethyl cellulose, oxypropylation, polyethylene glycol or polyethylene oxide, carbopol, polyvinylpyrrolidone, polyvinyl alcohol, etc.

The technical result of the invention is that obtained new Homo - and hetero-polyamidation fullerene derivative of the formula:where n=2-5, x=3, L=-(CH2) where m=1-5, or-CO(CH2)kCH(NH2)-, where k=1-2, characterized in that the compounds contain covalently-linked amino acid groups and polar ionic form amino acids, has developed a new industrial method of obtaining amino acid derivatives of fullerene C60with different ratio of components using the reaction of nucleophilic attach to the fullerene amino acids with the formation of addition polymerisation products. The proposed pharmaceutical composition containing as active substance Homo - and hetero-polyamidation fullerene derivative of the formula (II)having activity against herpes virus, hepatitis C virus, influenza viruses of different nature, HIV and antitumor and protivopsoriaticescoe activity. The pharmaceutical composition is in the form of tablets, capsules, ointments, suppositories, solutions, sprays.

The compounds possess new properties:

- solubility in a mixture of dimethylsulfoxide - water(1:100, 1:200);

- high bioavailability;

- high efficiency effects on infected cells;

- low toxicity.

The inventive method allows to obtain various Homo - and hetero-polyamidation fullerene derivative based on the ratio of reactants and process conditions. The method is based on the use of the communicate in the stage of synthesis of the optimal ratio of the initial reagents (1:10), minimum quantities of organic solvent and phase transfer catalyst, followed by separation of the claimed compositions using concentrated solutions of organic and mineral acids, followed by the introduction of amino acids number of NH2LCOOH, where L=-(CH2)mor-CO(CH2)kCH(NH2), which leads to quantitative getting vulernability compositions of certain structure and possible applications of the proposed method for the industrial synthesis of different efficiency and environmental friendliness.

The invention is illustrated by the following examples.

Example 1. Getting fulleropyrrolidines acid formula

To a solution of 7.2 g (0,01 mol) of fullerene C60in 400 ml of o-dichlorobenzene add 17 g (0.1 mole) of their finely ground anhydrous potassium salt of ε-aminocaproic acid. To the resulting suspension with stirring and heated above 80°C is added during 2 hours to a mixture of o-dichlorbenzene and methyl ester of polyethylene glycol 400 in the ratio of 2.5:1. The reaction mixture is stirred at a temperature not exceeding 60°C for 2-3 hours until complete discoloration of the solution and formation of a solid precipitate. The mixture is then filtered, the filter cake was washed with several portions of ethyl alcohol iwishiwas in vacuum at a temperature not exceeding 60°C. Dedicated potassium salt of fullerenelike dissolved in 2 l of distilled water. To the solution slowly with stirring 1N solution of hydrochloric acid to a pH of 5.1. The mixture defend to complete planting of the product. Then the aqueous layer was decanted. To the precipitate, which is a fine suspension of solid product in water, is slowly added with stirring a solution of aminocaproic acid in a mixture of dimethyl sulfoxide with water (1:10). The mixture is stirred until complete dissolution. Then the solvents are removed by distillation in vacuum. The solid residue is dried at a temperature not above 60°C in a vacuum drying Cabinet.

The yield of the target product is quantitative with respect to the original fullerene. The connection is a dark-brown solid, soluble in a mixture of dimethylsulfoxide: water of 1:200, sparingly soluble in mixtures of CH3CN:H2O - 1:1 DMF - H2O.

Thermogravimetric analysis of the product indicates the presence of a complex of 3 moles of loosely coupled aminocaproic acid, useplease with decomposition at 200°C. thermal decomposition of fullerenealuminum takes place at a temperature of 345°C with the release of fullerene and its oxidation products. The amount of solid residue after decomposition of compounds, representing according to diffraction analysis nezam is placed fullerene, corresponds to the ratio of C60:amino acid fragment as 1:6.

Acid hydrolysis of compounds of 0.1 molar solution of HCl leads to the selection of aminocaproic acid hydrochloride in an amount of 3 moles per mole of the original substance.

Adsorption of the compound onto the surface of silica gel leads to the splitting of the ion relations and the allocation of free aminocaproic acid. Subsequent photometric analysis of the reaction products aminocaproic acid with ninhydrin defined number innovating amino acid groups. Their number corresponds to the composition of the claimed compounds.

Electronic absorption spectrum of the product does not contain absorption bands of free fullerene.

The IR spectrum of the product contains absorption bands, characteristic of N-substituted amino acids: a group-COOH - 1704 cm-1, 1658 cm-1group- 3100, 2550, 2000 cm-1, -N-H-stretching vibrations 3300 cm-1N-H deformation 1552 cm-1the absorption bands With60-NH-R - 1104 cm-1, 930 cm-1, 830 cm-1.

Elemental analysis of the product shows the following ratio of components:%=76,84; %H=4,80; %N=5,15, for gross formula C96H75N6O12(molecular weight 1503) calculated: %C=76,49, %N=4,90, %N=5,57.

Example 2. Obtaining N-fullerene-γ-aminobutyric acid with β-alanine formula:

To a solution of 7.2 g (0,01 mol) of fullerene C60in 400 ml of o-dichlorobenzene add 14 g (0.1 mole) of their anhydrous potassium salt of γ-aminobutyric acid. To the resulting suspension while stirring and heating above 60°C is added during 2 hours to a mixture of o-dichlorbenzene and methyl ester of polyethylene glycol 400 in a ratio of 3:1. The reaction mixture is stirred at a temperature not exceeding 60°C for 2-3 hours until complete discoloration of the solution and formation of a solid precipitate. The mixture is then filtered, the filter cake was washed with several portions of ethanol and dried in a vacuum at a temperature not exceeding 60°C. the isolated product was dissolved in distilled water. To the solution slowly with stirring 1N solution of hydrochloric acid to a pH of 5.1. The mixture defend to complete planting of the product. Then the aqueous layer was decanted. To the precipitate, which is a fine suspension of solid product in water, slowly with stirring, add alcohol solution of 2.7 g (0,03 mole) of β-alanine. The mixture is stirred for 2 hours to dissolve the precipitate. After removal of solvent selected 12 g of the product.

The connection is a dark-brown solid, soluble in a mixture of dimethylsulfoxide:water of 1:200, sparingly soluble in mixtures of CH3CN:H2O - 1:1 DMF - H2O.

Thermogravimetric analysis of the product indicates the presence of a complex of 3 moles of a loosely-coupled β-alanine, ottseplyayuschimsya at a temperature of 210°C. thermal decomposition of the whole complex takes place at a temperature of 335°C with the release of the fullerene in an amount corresponding to the value Of60:amino acid fragments as 1:6.

Acid hydrolysis of the compounds of 0.01 molar solution of HCl leads to the formation of the hydrochloride of β-alanine in an amount of 3 moles per mole of the original substance.

Spectra of the product in solution in dimethyl sulfoxide and 0.1 N aqueous NaOH solution contain absorption bands in the region 217, 260, 335 nm, characteristic of the fullerene derivatives and do not contain absorption bands of free fullerene.

The IR spectrum of the compound contains absorption bands, characteristic of N-substituted amino acids and cationic forms of the amino acid group-COOH-1717 cm-1, 1710 cm-1, 1658 cm-1group3100, 2550, 2000 cm-1N-H-stretching vibrations 3400 cm-1N-H-stretching vibrations - 3400 cm-1N-H-deformation - 1552 cm-1the absorption bands With60-NH-R - 1104 cm-1, 930 cm-1, 830 cm-1.

Elemental analysis of the product shows the following ratio of components:%=75,24; %H=3,80; %N=6,25, for gross formula C81H48N6O12(molecular weight 126) calculated: %C=75,00, %H=3,70, %N=6,48.

Example 3. Obtaining N-fullerene-ε-aminocaproic acid with glutamine formula:

The process is conducted as in example 1. Only at the stage sludge treatment after deposition of the acidic form the fullerene-aminocaproic acid add a solution of 4.5 g of glutamine in a mixture of dimethylsulfoxide - water. The solvents are removed by distillation in vacuum.

Elemental analysis of the solid product shows the following ratio of components:%=71,34; %H=4,80; %N=8,25 to gross formula C93H69N9O15(molecular weight 1551) calculated: %C=71,95, %N=4,50, %N=8,12.

The IR spectrum of the compound contains absorption bands, characteristic of N-substituted amino acids and cationic forms of the amino acid group-COOH - 1714 cm-1, 1707 cm-1- =O(NH-C60) 1658 cm-1group3000, 2550, 2000 cm-1N-H-stretching vibrations 3400 cm-1N-H deformation 1552 cm-1the absorption bands With60-NH-R - 1104 cm-1, 930 cm-1, 830 cm-1.

Acid hydrolysis of compounds leads to the separation of glutamine hydrochloride in an amount of 3 moles per mole of the original substance.

Was studied the antiviral activity of compounds against HIV, HSV, influenza virus, and anti-tumor activity. In the following examples, the preparation obtained by the method described in examples is e 1, named by the text preparation No. 1 (fulleropyrrolidines acid).

Example 4. The study of the activity of the drug fulleropyrrolidines acid against human immunodeficiency virus.

The studies were conducted in the laboratory of the human immunodeficiency virus with a test Centre for expert evaluation of antiviral and disinfectants (Institute of Virology. Dijanoveckog RAMS).

To the cells was added to the study drug and were infected with virus at a dose of 0.01 TCID50/cell. Incubated cell culture at 37°C in an atmosphere with 5% CO2and 98% humidity 4-5 days. Analysis was performed by staining cells with dye and light microscopy: study of the cytopathic effect of the virus (JRC) and virusinduced since-tyabrskaya (syncytium - a conglomerate of several cells from the total cell membrane, formed by the merger of their membranes).

The degree of cytodestructive was evaluated under a microscope by the common chetyrehrazovoe the system by the signs + or - respectively the number of dead cells in each of the four holes corresponding to one of the investigated parameter.

++++ - 100%thcell death in four holes used in the experience of one breeding

+++ - 75%thcell death in each of the four holes,

++ - 50%ththe death of cells is in each of the four holes,

+ - 25%thcell death in each of the four holes,

+- - beginning degeneration

- - no cytodestructive.

The results of the study are presented in tables 1-2.

The data obtained (table 1, 2) showed that the investigated drugs fulleropyrrolidines acid possessed antiviral activity against human immunodeficiency virus type 1. EC50(50%effective concentration) of the drug of 0.9 µg/ml of the Drug in concentrations of 0.5-10 µg/ml had no cytotoxic effect on the cells.

Example 5. The study of the antiherpetic activity of the drug fulleropyrrolidines acid in vitro experiments. The research was carried out Institute of Virology. Dijanoveckog RAMS, Moscow.

During the study examined the cytotoxic and anti-herpes drug activity in cell cultures Vero.

In the study were used: human cell culture of monkey kidney (VERO)derived from tissue culture collections of the Institute of Virology. Idivisor RAMS; herpes simple (Herpes simplex virus type 1, strain L2that is propagated in Vero cells. The cells were infected with virus at a dose of 100 TCID50/0.2 ml and 1000 TCID50/0,2 ml

For the study presents a sample of the substance in powder form dark color.

The original drug is dissolved in dimethyl sulfoxide (DMSO) at a ratio of 1:20, and then prepared working concentration in the medium NEEDLE MEM. Assessment of the activity of the tested samples was performed according to the degree of protection of cells from virusinduced cytotoxic effect of HSV microscopic method and the method of MTT optical density.

The results of the study. Investigated the toxicity of substances used in concentrations, and solvent (1.5 ml DMSO in 50 ml of water).

The drug in final concentrations of 50 to 1000 μg/ml was added to the monolayer of VERO cell culture, and incubated in an atmosphere of 5.0% CO2at 37°C for 24-48 hours. The monolayer cells were stained with 0.4% solution Trypanosoma blue and examined under a microscope.

Sample concentrations up to 100 µg/ml had no toxic effects on the culture of VERO cells, at a concentration of 200 μg/ml signs of toxicity - celebrated the death of 25% of the cells. The concentration of 500 μg/ml or more is already toxic to Vero cells (table 3).

The study of protective properties of a sample was carried out at one scheme introduction - an hour before infection, and two doses of virus - 100 TCID50and 1000 TCID50.

The results are presented in tables 3-7.

Studies have shown that the introduction of the 60 minute sample of the drug to the infection of the cell culture virus herpes simple in a dose of 100 TCID50fully protects cells is t cytodestructive steps of the virus at all tested concentrations from 5.0 μg/ml and above (see table 4 and 5).

When increasing the infective dose herpes virus to 1000 TCID50/0.2 ml (see table 6 and 7), the sample preparation ensures complete protection of sensitive cell culture from cytodestructive steps of the virus in this scheme test (infection of cells with virus after 60 minutes after introduction of the drug in the culture medium).

Example 6. Learning activity fulleropyrrolidines acid (product No. 1) in respect of the influenza virus A/IIV-Moscow/01/2009 (H1N1)swl.

The studies were conducted in the Institute of Virology. Dijanoveckog RAMS, Moscow. The objective of the research was to study the antiviral activity of these drugs in the culture of MDCK cells against influenza virus A/IIV-Moscow/01/2009 (H1N1)swl.

The drug was dissolved in DMSO (5 mg substance + 0.5 ml DMSO) followed by the addition of 4.5 ml of medium for cell cultures MEM, and thus, the runoff concentrations of 1.0 mg/ml In subsequent conducted breeding wastewater environment MEM to working concentrations of 6.5 μg/ml and 12.5-25,0-50,0-100 µg/ml

Determination of the antiviral activity of the substances was performed to reduce the reproduction of influenza virus in cell culture MDCK detected by ELISA.

With this purpose, cells MDCK were grown in 96-well tablets up to a full monolayer was washed from the growth medium and added substances in double concentration in 100 μl of medium Maminirina virus in the working dose of 100-1000 TCID 50conducted in two modes: 2 hours after introduction of substances and simultaneously. The plates were incubated in an incubator with CO2for 24 hours at 37°C. After incubation the medium was removed and cells were fixed with 80% acetone in PBS for 15 minutes, well dried and staged ELISA, by conducting successively the adsorption of specific reagents monoclonal antibody, conjugate and substrate (orthophenylphenol). The reaction was accounted by optical density at 492 nm on a spectrophotometer firm "Biocom". Each dilution of virus was investigated in 3 repetitions, for which the calculated average value of the optical density (OD). The percentage of inhibition was determined as the ratio between the difference OP OP experience and the cell control divided by the difference OP virus control and es cell control, multiplied by 100%. On the basis of the data obtained, the value of the minimum concentration of a substance that causes 50.0% inhibition of viral reproduction (MIC50).

Assessment of the suppression of the reproduction of the virus of influenza A(H1N1) was performed in 3 experiments at different multiplicity of infection. The results are presented in table 8 (protocols 3 experiments) and table (average values of the obtained results of 3 experiments).

As can be seen from table 9, there was clear dependence of the degree of reproduction and concentrations: from above the receiving concentration - reduced reproduction of the virus. In addition, significant differences in performance in different modes of infection (2 hours after making the drug or simultaneously) is not marked.

Thus, the results obtained studying the activity of different dilution of the drug against influenza virus A/IIV-Moscow/01/2009 (H1N1)swl revealed high activity of suppressing its reproduction in the culture of MDCK cells. While the modes of making preparations for 2 hours prior to infection or simultaneously with infection had no effect on their activity in cell culture MDCK.

Example 7. Study of antiviral activity of fullerenol-aminocapronic acid (product No. 1) on the model of influenza pneumonia in mice.

The study was performed in the center of the chemistry of drugs (CHLS-UNIFI), Moscow.

In this work, we used the drug in the form of a dark-brown crystalline powder. For oral administration prepared the necessary dosage, dissolving the sample in a 1% solution of starch, boiled water. For vnutribruchinnogo and intramuscular administration sample preparation was dissolved in a 1.5% solution of DMSO.

The work was used influenza virus A/Aichi/2/69 (H3N2), adapted to mice. This virus is widely used to determine the effectiveness of antiviral drugs on the model of influenza pneumonia in mice and was received is from the Museum of viral strains and cell cultures Institute of Virology RAMS. For the preparation of infective material, the mice were infected intranasally allantoin virus, after developing signs of disease were scored and in sterile conditions received the homogenate of lung tissue. Next, the homogenate was used to infect 10-day-old chick embryos, from which received allantoine virus and after titration his mice was used to infect the animals.

Outbred mice (females) weighing 12-14 g were obtained from the kennel "Andreevka" (Moscow region) and were kept on a standard diet in regulated vivarium conditions.

Weighted mouse (female non-linear, the average weight of 12-14 g) were infected intranasally under light ether anesthesia influenza virus A/Aichi/2/69 (H3N2) (LD50100 µl). In the preliminary experience carried out a determination of LD50by titration allantoinase virus such as mice, which are then used in the main experience. Was used following treatment with investigational drug: 24 hours prior to infection, 1 hour before infection, after 24 hours, and then 1 time per day after 24 hours for 5 days. For oral administration used disposable insulin syringe with a special needle (lavage), each dose was administered in a volume of 100 μl. For intraperitoneal and intramuscular treatment also each dose was administered in the volume of 100 ál. The virus control group consisted of 10 mice infected with the virus, but not treated with drugs. Also in the experiment had two groups of 10 uninfected mice, which were injected intraperitoneal and intramuscular injection of 100 μl of 1.5% DMSO, which was used as a solvent preparations. In other groups also initially contained 10 animals. For treated and control animals was performed daily observation, in the first 5 days after infection mice were weighed every day, after day. Chemotherapeutic activity of the drug on the model of influenza pneumonia mice was assessed by three criteria: a measure of protection against lethal viral infection, increasing life expectancy and reducing weight loss in the groups of animals treated with the drug compared with the control group.

Treatment filerenameoperations acid was effective, reducing the mortality of mice against influenza pneumonia and loss their weight and increasing life expectancy compared to the virus control. The effectiveness of this treatment depended on the dose and method of treatment. The results are presented in tables 10-11.

The most effective in all three parameters, a measure of protection from mortality, life expectancy and weight loss) was the treatment fullerenelike ronojoy acid intramuscularly, which in doses of 100 and 200 mg/kg/day prevented the loss of 60-70% of infected animals and the loss of their weight, and increased their life expectancy by almost 2 times. Intraperitoneal treatment fulleropyrrolidines acid was effective only at doses of 50 and 100 mg/kg/day. The death of animals, a significant reduction in life expectancy and weight of mice after intraperitoneal treatment fulleropyrrolidines acid at a dose of 200 mg/kg/day suggest that this dose in this method of introduction is toxic to infected mice.

Example 8. The study of antitumor activity fulleropyrrolidines acid (product No. 1) in transplantable tumors leukemia L 1210, mammary adenocarcinoma CA-755 and carcinoma Lewis.

The study was performed at the Institute of toxicology, Saint-Petersburg, 2006, in accordance with the "guidelines for the study of antitumor activity of pharmacological substances" ("Guidance on experimental (preclinical) study of new pharmacological substances", the Ministry of health, Remedium. M, 2000, s-325).

Strains of tumor cell leukemia L1210, mammary adenocarcinoma CA-755, and the strain of tumor cell carcinoma Lewis (3LL) were obtained from the Institute of Oncology. Professor New MZ the Russian Federation (St. Petersburg).

In researching the AI used the following test systems:

Mouse strain DBA/2 with inoculated cell leukemia L1210. The age of the mice 6-8 weeks, weight 19-25,

Mouse male line From 57 BL/6j with inoculated cells CA-755. The age of the mice 6-8 weeks, weight 19-25,

Mouse male line From 57 BL/6j intertwined with 3LL cells. The age of the mice 6-8 weeks, weighing 18-24 g

Evaluation of antitumor action was based on:

- assessment of the accumulation of ascites registration weight gain of animals;

assessment of animal longevity;

assessment of tumor growth. A measurement was performed in a larger tumor size (length) and perpendicular to it smaller size (width). Expected volume of tumors and inhibition of tumor growth was calculated by the formula. The evaluation criterion was the day of the achievements of each individual tumor volume of 500 mm3.

The effects of the drug on the development of leukemia L1210 presented in table 12.

Table 13 shows the average life expectancy of animals with transplantable tumors leukemia L1210 and control groups, and these increase in life expectancy compared with the control group in %.

As can be seen from table 13 the average life expectancy of the control group mice with inoculated tumor cell leukemia L1210 was 6.0±0.21 days. The introduction of the drug significantly increased the lifespan of mice to 10.7±0.3 days and or 10.60±0.37 days, the percentage increase in life span compared to control was 78,33% 76,67% for doses of 100 mg/kg and 200 mg/kg, respectively, but the differences between the experimental groups was not statistically significant.

In the course of the experiment were conducted daily weighing of animals to assess the accumulation of ascites and study compared the drugs on this process.

The results of the evaluation of weight gain are presented in table 14.

As can be seen from table 14, the drug significantly reduced the accumulation of ascites in mice with inoculated tumors leukemia L1210. Gain body weight of mice treated with the study drug was significantly less than in the control group and animals treated with the study drug at a dose of 250 mg/kg, the average weight gain was significantly lower (1.5 times) compared with those observed for animals treated with the drug at a dose of 100 mg/kg

Thus, the obtained results allow to conclude that the drug is pronounced antitumor activity and inhibits growth of tumor cell leukemia L1210 in mice, which was reflected in a significant increase in life expectancy (78,33% 76,67%) for doses of 100 and 250 mg/kg and reliable braking accumulation of ascites experimental animals (78-43%). Signs of intoxication on the background of the introduction of a test prep the rata has not been registered. The analysis of the data revealed significant differences between the dose of 100 mg/kg and 250 mg/kg - increase in dose leads to a significant decrease in the accumulation of ascites in experimental animals.

Other indicators contrast (contrast medium for groups that received different doses of the study drug) was at the level of error caused by natural variation of the data.

The effects of the drug on the development of mammary adenocarcinoma CA-755 presented in table 15.

Table 16 shows the average life expectancy of animals with transplantable tumors mammary adenocarcinoma CA-755 and the control group, and these increase in life expectancy compared with the control group in %.

As can be seen from table 16, the average life expectancy of the control group mice with inoculated tumor cells tumors of the mammary adenocarcinoma CA-755 amounted to 37.9±0.74 days. The introduction of the drug significantly increased the lifespan of mice to 71.9±2.58 days and 73.4±0.92 days, the percentage increase in life span compared to control was 89,71% and 93,67% for doses of 100 mg/kg and 200 mg/kg, respectively, but the differences between the experimental groups was not statistically significant.

Thus, the results obtained allow the transaction shall be the conclusion of the the drug is pronounced antitumor activity and inhibits growth of tumor cells mammary adenocarcinoma CA-755 in mice, which was reflected in a significant increase in life expectancy (89,71% and 93,67%) for doses of 100 and 250 mg/kg signs of intoxication on the background of the introduction of the investigational product is not registered.

Evaluation of the effect of the drug on the average volume of tumors carcinoma Lewis at different days after inoculation. In the control group tumors developed in 21 of the 26 mice (80,8%), the first tumor were recorded on the 7th day after inoculation and were recorded up to 40 days. The first tumor in the group with the influence of the drug at a dose of 100 mg/kg were recorded on the 7th day after inoculation and were recorded up to 60 days; in the group with the influence of the drug at a dose of 250 mg/kg were recorded on day 8 after inoculation and were recorded up to 60 days. The drug showed a pronounced antitumor effect on the development of carcinoma of the lung Lewis; the percentage of inhibition in a group of drug is 100 mg/kg compared with the control group on 10-17 days was significantly 71,77% and 58.5%, in the group of drug - 250 mg/kg compared with the control group on 10-17 days 84,37% 54,2%, respectively.

Table 17 presents the effect of the drug on the growth of carcinoma Lewis, were analyzed for achieving tumor size 00 mm 3; in the control group, this figure ranged from 12 to 20 days, and in the experimental groups from 17 to 28 days. As can be seen from this table, the drug significantly increased this ratio to 22-27% compared to control.

The death of the mice of the control group was caused by the progression of tumor growth of the primary tumor, as well as due to extensive metastatic lung lesions. Individual lifespan of mice in the control group ranged from 28 to 39 days in the group with drug from 51 to 60 days. Table 18 presents the effect of the drug on the average lifespan of mice after inoculation of the tumor. As can be seen from table 18, the drug significantly increased the average life span of about 70% compared with the control group, the difference between the experimental groups are statistically unreliable.

Individual lung weight and the number of lung metastases in mice of the control group and experimental groups with the influence of the compared drugs are presented in table 19.

As can be seen from table 19, the drug significantly reduced lung weight and the number of lung metastases in 1,5-2 times. The difference between the experimental groups are statistically unreliable.

Table 1
The study of the cytotoxicity of the drug on the model of lymphoblastoid cells
Conditions of experience, concentration, µg/mlCell viability, %The number of cells ×103/ml
Control cells98800
Preparation No. 10,595833
1,094833
5,098799
10,094767
10095600

td align="center"> The number of cells ×103/ml
Table 2
The antiviral activity of the drug on the model of human cells infected with HIV-1
Conditions of experienceConcentration, mg/mlCell viability, %The centre e/sincity (+)
Control cells0988000
The control virus020664,0
Preparation No. 10,518334,0
1,0264954,0
5,0986330
10986000

Table 3
Study of cytotoxic effect of sample fulleropyrrolidines acids in cell culture Vero
Concentration, mg/mlCytotoxic effect on the cells, %
500,0
1000,0
20025,0
500100,0
1000100,0
Note: DMSO concentration used to dissolve not have a cytotoxic effect on cells

Table 4
Antiherpetic activity of the sample fulleropyrrolidines acid in the culture of Vero cells with infective dose herpes virus - 100 TCID50/0.2 ml
Concentration, mg/mlProtection of cells from the cytopathic effect of herpes virus, %
5,0an 81.25±12,5
10,0100±0,0
50,0100±0,0
100,0100±0,0
Note: cells infected in an hour after you put the I samples.

Table 5
The protective effect of the drug from cytodestructive actions herpes virus, type 1 when the infective dose of the virus 100 TCID50
The drug concentration, µg/mlPreparation No. 1
The optical densityDifferences from control significant at p
00,403±0,02
5,01,110 land only±0,08<0,01
10,01,015±0,07<0,01
50,01,153±0,06<0,01
100,01,79±0,05<0,01
Control cells1,133±0,07<0,01

Table 6
Antiherpetic asset is ity of the sample fulleropyrrolidines acid in the culture of Vero cells with infective dose herpes virus - 1000 TCID50/0.2 ml
Concentration, mg/mlProtection of cells from the cytopathic effect of herpes virus, %
5,0100±0,0
10,0100±0,0
50,0100±0,0
100,0100±0,0
Note: the cells were infected through hours after injection sample

td align="center"> 50,0
Table 7
The protective effect of the drug from the cytopathic effect of herpes virus, type 1 when the infective dose of the virus 1000 TCID50
The drug concentration, µg/mlThe optical densityDifferences from control significant at P
0coefficient was 0.796±0,06
5,0of 1.027±0,04<0,01
10,01,021±0,04<0,01
1,033±0,03<0,01
100,01,083±0,01<0,01
Control cells1,133±0,07

Table 8
The results of the study activity of the drug No. 1 in respect of the influenza virus A/IIV-Moscow/01/2009 (H1N1)swl
Concentration is
tion drugs (µg/ml)
Making medicationReduction (%) reproduction of influenza virus in cell culture MDCK relative to the control in the presence of batches of product No. 1
6,252 hours before infection27,0-28,0-0
with simultaneous infection18,0-0-0
12,52 hours before infection47,0-74,0-9,5
with simultaneous infection39,0-13,0-0
25,02 hours on the infections 41,0-79,0-12
with simultaneous infection40,0-0
50,02 hours before infection28,0-72,0-31,0
with simultaneous infection33,0-10,0
100,02 hours before infection4,0-0-20,0
with simultaneous infection29,0-28,0

Differences in rates of 2 experiments was dependent on the multiplicity of infection of cell culture MDCK.

Table 9
The results of the study activity of drugs against influenza virus A/IIV-Moscow/01/2009 (H1N1)swl, averages
Concentration is
tion drugs (µg/ml)
Making medicationReduction (%) reproduction of influenza virus in cell culture MDCK relative to the control in the presence of batches of product No. 1
6,252 hours before infection 18,0
with simultaneous infection6,0
12,52 hours before infection44,0
with simultaneous infection26
25,02 hours before infection44,0
with simultaneous infection20,0
50,02 hours before infection44,0
with simultaneous infection22,0
100,02 hours before infection8
with simultaneous infection29,0

Table 12
The effect of the drug on the lifespan of mice with transplantable tumor leukemia L1210
Name No. of the animal and its life expectancy in days
group12345678910
Control6776656656
Preparation No. 1 100 mg/kg9111311121011101010
Preparation No. 1 250 mg/kg10101091011 13101112

Table 13
The effect of the drug on the average life expectancy of animals with transplantable tumor leukemia L1210
Group nameMean life span in days, M±üUPI, %
Control6,0±0,21
The preparation step 1, 100 mg/kg10,7±0,37*78,33
Preparation No. 1, 250 mg/kg10,6010,37*76,67
Note: * - significant difference from control (p<0,05)

Table 14
The effect of the drug on the development of ascites in mice with transplantable tumor leukemia L 1210
DayThe control groupThe drug is 100 mg/kg The drug is 250 mg/kg
The average weight gain, M±mThe average weight gain, M±m% inhibition compared to controlThe average weight gain, M±M% inhibition compared to control
10,4±0,20,3±0,20,3±0,1
20,9±0,30,8±0,40,5±0,3
35,0±0,82,5±0,550,0%*1,1±0,478,0%*
411,4±1,14,2±1,063,2%*2,8±0,975,4%*
516,3±2,08,4±1,248,5%*4,7±1,271,2%*
621,2±2,1 12,1±2,142,9%*8,4±1,660,4%*
726,3±1,714,2±2,246,0%*10,0±1,362,0%*
Note: * - significant difference from control (p<0,05)

39
Table 15
The effect of the drug on the lifespan of mice with transplantable tumor leukemia, CA-755
Group nameN animal and its life expectancy in days
12345678910
Control363941343540384037
The preparation step 1, 100 mg/kg79817382806663606768
Preparation No. 1, 250 mg/kg77727078767173707275

Table 16
The effect of the drug on the average life expectancy of animals with transplantable tumor mammary adenocarcinoma CA-755
Group nameMean life span in days, M±mUPI, %
Control37,9±0,74
Preparation No. 1,100 mg/kg to 71.9±2,58*89,71
Preparation No. 1, 250 mg/kg73,40±0,92*93,67
Note: * - significant difference from control (p<0,05)

Table 17
The effect of the drug on the size of the carcinoma Lewis (achievement V=500 mm3)
Day achieve a tumor volume of 500 mm3
GroupM±mpercent increase statistical significance
Control16,7±0,65
The preparation step 1, 100 mg/kg21,3±1,2127,3% *
Preparation No. 1, 250 mg/kg20,5±1,2022,9%*
Note: * - significant difference from control (p<0,05)

The table is 18
The effect of the drug on the lifespan of mice after inoculation carcinoma Lewis
The life expectancy
GroupM±m% increase Statistical significance
Control32,81±0,86
The drug is 100 mg/kg55,92±0,9870,42%*
The drug is 250 mg/kg56,38±0,6871,85%*
Note: * - significant difference from control (p<0,05)

1486,7±64,08
Table 19
The effect of the drug on the metastasis of carcinoma Lewis in mice
Group # (number of animals)Lung weight (mg M±mThe number of lung metastases, M±MThe number of large (≥3 mm) metastases in the lungs, M±M
Control (n=21)70,5±5,5013,4±2,05
The preparation step 1, 100 mg/kg (n=12)491,9±63,72*53,5±11,23*5,8±1,91*
Preparation No. 1, 250 mg/kg (n=13)505,4±68,82*55,8±11,60*5,7±1,54*
Note: * - significant difference from control (p<0,05)

1. Homo - and hetero-polyamidation fullerene derivative of General formulawhere n=2-5, x=3, L=-(CH2)mwhere m=1-5, or-CO(CH2)kCH(NH2)-, where k=1-2, characterized in that the compounds contain covalently-linked amino acid groups and polar ionic forms of amino acids.

2. Derivatives of fullerene according to claim 1, characterized in that the amino acid groups use fragments of the amino acids of the aliphatic series of the General formula NH(CH2)nCOOH, where n=2-5.

3. Derivatives of fullerene according to claim 1, characterized in that as the polar ionic forms of amino acids using fragments amides of dicarboxylic amino acids of General formula NH2(CO)(CH2)kCH(NH2)COOH, where k=1-2.

4. The method of obtaining derivatives of fullerene according to claim 1, characterized is the action scene themes they are formed by the interaction of fullerene with a 10-fold molar excess of freshly prepared anhydrous potassium salts of amino acids of General formula NH2(CH2)nCOOK, where n=2-5, in the environment of aromatic organic solvent when added to the resulting suspension interphase catalyst under stirring and heated to a temperature not higher than 60-80°C until complete discoloration of the solution and formation of a solid precipitate, which is then allocated, followed by the treatment of aqueous solutions of potassium salts of fullerenealuminum 1N solution of an organic or mineral acid, followed by the introduction of a solution of amino acids of General formula NH2(L)COOH, where L=-(CH2)mwhere m=1-5, or-CO(CH2)kCH(NH2)-, where k=1-2 in polar solvents, mixing, removing the solvent, washing and drying the precipitate.

5. The method according to claim 4, characterized in that the use of anhydrous potassium salts of amino acids in fine condition, and the selection of sediment potassium salts fullerenealuminum carried out by filtration, washing with ethanol and drying.

6. The method according to any of claims 4 and 5, characterized in that as the interfacial catalyst using methyl esters of polyethylene oxides of molecular weight 200, 400, 500.

7. Pharmaceutical compositions the Oia, possessing activity against herpes virus, hepatitis C virus, influenza viruses of different nature, HIV and antitumor and protivopsoriaticescoe activity, containing as active substance fullerene derivative according to claim 1 in an effective amount.



 

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FIELD: organic chemistry, chemical technology.

SUBSTANCE: invention relates to novel 1,2,3-tris-{[aminopoly(ethyleneamino)ethylammonio]-methylcarbonyloxypoly(alkyleneoxy)]}propane trichlorides of the formula:

wherein at a + c + e (the general degree of oxypropylation) = 49; b + d + f (the general degree of ethylation) = 0, n = 1-6; at a + c + e = 55, b + d + f = 0, n = 1-6; at a + c + e = 49, b + d + f = 9, n = 1-6; at a + c + e = 10, b + d + f = 10, n = 1-6; at a + c + e = 66, b + d + f = 15, n = 1-6; at a + c + e = 76, b + d + f = 18, n = 1-6, and to a method for their synthesis. Method involves interaction of 1,2,3-tris-[hydroxypoly(alkyleneoxy)]propanes of the formula:

wherein a + c + e = 49-76, b + d + f = 0-18 with monochloroacetic acid in the presence of acidic catalysts, in boiling organic solvent medium, azeotropic removing water formed and the following treatment at heating the synthesized reaction product with polyethylenepolyamines of the formula: H2N(CH2CH2NH)nCH2CH2NH2 wherein n = 1-6, and in the following mole ratios of reagents - hydroxyl derivatives of propane : monochloroacetic acid : polyethylenepolyamines = 1:(3.0-3.2):(3.0-3.2), respectively. New compounds possess the fungicide activity, properties of emulsifiers of cationic bitumen emulsions, capacity to enhance adhesion of bitumen to mineral materials.

EFFECT: improved preparing method, valuable properties of compounds.

6 cl, 3 tbl, 6 ex

FIELD: organic chemistry, chemical technology.

SUBSTANCE: invention relates to novel 1,2,3-tris-[(ammonio)methylcarbonyloxypoly(alkyleneoxy)]-propane trichlorides of the general formula:

wherein at -X+ as -N+R1RR, R1 = R2 mean hydrogen atom (H), R3 means aliphatic hydrocarbon radical comprising 10-16 carbon atoms, a + c + e (the general degree of oxypropylation) = 49,b + d + f (the general degree of oxyethylation) = 0; at -X+ as -N+R1R2R3, R1 = R2 mean H, R3 means aliphatic hydrocarbon radical comprising 10-16 carbon atoms, a + c + e = 55, b + d + f = 0; at -X+ as -N+R1R2R3, R1 = R2 mean H, R3 means aliphatic hydrocarbon radical comprising 10-16 carbon atoms, a + c + e = 80, b + d + f = 24; at -X+ as -N+R1R2R3, R1 = R2 mean H, R3 means aliphatic hydrocarbon radical comprising 10-16 carbon atoms, a + c + e = 90, b + d + f = 27; at -X+ as -N+R1R2R3, R1 = R2 means H, R3 means phenyl, a + c + e = 80, b + d + f = 24; at -X+ as -N+R1R2R3, R1 = R2 mean H, R3 means phenyl, a + c + e = 90, b + d + f = 27; at -X+ as , a + c + e = 80, b + d + f = 24; at -X+ as , a + c + e = 90, b + d + f =27. Also, invention relates to a method for synthesis of these compounds. Method involves interaction of 1,2,3-tris-[hydroxypoly(alkyleneoxy)]-propane of the formula:

wherein a + c + e = 49-90, b + d + f = 0-27 with monochloroacetic acid in the presence of acidic catalyst, in boiling organic solvent medium with azeotropic removal of water formed and the following treatment of synthesized reaction product in polar solvent medium at heating with amino-compounds of the formula: NR1R2R3 wherein R1 = R2 mean H, R3 means aliphatic hydrocarbon radical comprising 10-16 carbon atoms, phenyl, or morpholine of the formula:

in the following mole ratios of reagents - propane hydroxyl derivative : monochloroacetic acid : amino-compound or morpholine = 1:(3.0-3.2):(3.0-3.2), respectively. New compounds show the bactericidal and fungicide activity and properties of demulsifying agents for petroleum emulsions.

EFFECT: improved method of synthesis, valuable properties of compounds.

7 cl, 3 tbl, 8 ex

FIELD: medicine.

SUBSTANCE: invention relates to medicine, oncology, and can be used in surgical treatment of tongue cancer. At the first stage under local anesthesia with novocaine solution into femoral vein from the side of injury guide is introduced and brought under radioscopy control into corresponding carotid artery, guide is installed at the orifice into corresponding lingual artery. Chemical embolisation of lingual artery is performed and guide is removed. At the second stage total biopsy of tumour ulceration is performed. At the third stage high-frequency hyperthermia of tongue is performed by thermal impact at temperatures 60-80 degrees Celsius during two minutes, with application of multiple introduction of apparatus antenna, which contains active electrode, into tongue tissue by punctures in area of lateral - free- edge of that half of tongue where tumour was located, from tongue tip to its root, to the depth not less than 1-2 cm.

EFFECT: such total and uniform impact ensures death of tumour cells, their "blocking" on tumour boundary, prevention of hematogenic and lymphatic cancer spread, intraoperation complications, in particular, bleeding; makes it possible to carry out complete histological analysis on tongue tissues which were not modified by hyperthermia.

1 dwg, 1 ex

FIELD: medicine.

SUBSTANCE: there are offered versions of: a SSMA monoclonal antibody and its antigen-binding fragment which are bound with a mark or a cytotoxic agent. There are described versions of a pharmaceutical composition and a diagnostic kit based on such antibodies. There are disclosed methods for identification in vitro of tumour cells, as well as for diagnostic identification of tumour cells based on such antibodies. What is described is an isolated polynucleotide for producing monoclonal antibodies.

EFFECT: using the invention provides the antibodies which can bind SSMA in its native form on the surface of tumour cells, are bound with LNCAP, but are not bound with cells with lost SSMA expression that can find further application in prostate cancer therapy.

15 cl, 21 dwg, 3 tbl, 18 ex

FIELD: medicine, pharmaceutics.

SUBSTANCE: invention refers to to new compounds of formula (1) or their pharmaceutically acceptable salts, optionally in the form of (1S)-isomers showing the properties of polo-like kinase (serine-threonine kinase) PLK1 inhibitor. In the compounds of formula (1) , R1 represents a halogen atom; a lower alkyl group having 1-2 carbon atoms, which can be substituted by 3 fluorine atoms; or a cyclopropyl group; R2 represents a hydrogen atom; one of R3 and R4 represents a hydrogen atom while the other one of R3 and R4 represents: a) a lower alkyl group substituted by NRaRb wherein each Ra and Rb, which can be identical or different, represent a lower alkyl group, or each Ra and Rb, which can be different, represent a hydrogen atom, a lower alkyl group or a cycloalkyl group having 3-6 carbon atoms wherein a cycloalkyl group can be substituted by one ore more substitutes which can be identical or different and specified in a group 1): a lower alkyl; b) a 4-6-member aliphatic heterocyclic group specified in an azetidinyl group, a pyrrolidinyl group and a piperidinyl group; c) a lower alkyl group substituted by a 4-6-member aliphatic heterocyclic group specified in an azetidinyl group, a pyrrolidinyl group and a piperidinyl group; d) a 6-member aromatic heterocyclic group specified in a pyridyl group wherein each of an aliphatic heterocyclic group and an aromatic heterocyclic group can be substituted by substitutes specified in a group 1) described above; R5 represents a hydrogen atom, a cyano group, a halogen atom or a lower alkyl group.

EFFECT: compounds can find application in treating oncological diseases.

10 cl, 4 dwg, 8 tbl, 42 ex

FIELD: medicine, pharmaceutics.

SUBSTANCE: invention refers to new hydrated N-fullerene-amino acids of general formula C60(H)3{NH(CH2)nCOOH}3·xH2O, wherein C60 represents fullerene, n = 5-7, x = 8-10, which possess herpes virus, influenza virus, HIV activity, as well as anticancer and antipsoriatic activity. The invention refers to a method for preparing said fullerene amino acids and based pharmaceutical compositions.

EFFECT: preparing new hydrated N-fullerene-amino acids.

5 cl, 28 tbl, 10 ex

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