Biotreatment blood serum, methods of production and application

FIELD: medicine.

SUBSTANCE: group of inventions concerns biology and medicine. There is disclosed method of biotreatment blood serum production involving electrostimulation of an animal, other than a human, blood drawing of specified animal, serum separation from specified blood and gamma irradiation in a dose 10 to 40 kGy. There is offered blood serum product and pharmaceutical composition containing said blood serum product, and also their application for treatment of various diseases.

EFFECT: invention provides production of biotreatment animal's blood serum applied for treatment of wide range of diseases, including epileptic attacks and apoplexy.

24 cl, 4 ex, 7 tbl, 11 dwg

 

The present invention relates to a method of obtaining a product of the blood serum, the product of the serum and pharmaceutical compositions containing the specified product serum, and also to their use for the treatment of various diseases and conditions, including seizures and apoplexy.

The level of technology

Ways to get active substances from the blood serum is known from the prior art. One of them involves taking blood from a human or animal, subsequent incubation and separation of the active substance and, finally, conservation of matter (see, for example, JP 2123287, EP 0542303, EN 2096041, EN 2120301). The method of the prior art relates to obtain serum, which increases the body's resistance to exogenous and endogenous factors, such as air pressure, air temperature, gravity, light, etc. as well as hunger, thirst, sleep and sex drive, etc. Serum collected from a donor who was previously given in a certain functional state and depending on the duration of application, functional status and type of the functional state, for example, insomnia, alcohol dependence, nicotine dependence, etc. can be obtained serum with different biological activity, which exhibits mitogenic, somnolence, ofta Imogene, audioactivity, thermoactivated affecting the diet that affect sexual activity, antihypoxic, anti-alcohol and antinicotine action.

A great way disclosed in EP 1283047 and for the processing of the blood serum of the animal gamma irradiation to improve the biological activity of the product of serum.

Currently, a large number of studies have focused on the mechanisms that regulate cell proliferation of various human tissues. In this respect investigated as stimulants and inhibitors of proliferation of normal and pathologically altered somatic cells, including nerve cells (see, for example, Aschmarin, R "Neurochemistry", that won, Publishers of the Biomedical Chemical Institute of the Russian Academy of Science, 1996).

It was noted that peptide growth factors in addition to the General trigger functions, such as stimulation of mitosis, cell differentiation and cell growth of different types of normal tissue, increase wound healing, can cause the formation and tumor growth (see, for example, Bouneres, P. (1993), Horm. Res, 40: 31; Robinson C., (1993) Ann. Med 25: 535; Dignez, and Casanueva F. (1995) Trends Endocrin. Metab. 6: 55, Menster D. et al. (1995) Clin. Exp. Metastasis 13: 67).

Such peptides, such as peptides of parathyroid gland, gastrin or bombezin contribute to the development of tumor cells, as well as the development of breast cancer, cancer of the bones and tools the big intestine (see, for example, Kitazawa S. and Maeda S. (1995) Clin. Orthop. 312: 45-50 and Kaji et al. (1995) Endocrinology 136: 842).

Despite the fact that some peptides contribute to the division of normal cells and are stimulants for humans and animals, there is a danger that their application will lead to the development of tumor cells and, ultimately, to the development of cancer.

Earlier experiments have shown that stimulation of animal electricity leads to increased levels of β-endorphin in the blood (see, for example, Litvinova S.V. et al. (1990) Biomed. Sci. 5: 471). In the reference work Udovitschenko W.L. presents numerous data regarding the results of stimulation or shock due to various reasons. It is shown that, for example, the shock leads to a noticeable increase in the concentration of β-endorphins, meta - and lancifolia in the blood (see Udowitschenko W.I. (1989) "Xenogenic Opioid System in Shock" Pathiological discrimination and Experimental Therapy" 6: 72-77).

The invention

The aim of the present invention is to develop a new method of obtaining biologically active blood serum from the blood of the animal. Unexpectedly it was discovered that the biologically active blood serum, obtained in accordance with the method of the present invention, demonstrated new therapeutic properties.

Therefore, one aspect of the present invention is a method of obtaining biologically active savored the blood, providing steps:

a) electrical stimulation of the animal, not a man,

b) taking blood from a specified animal,

c) allocation of serum from a specified blood, and

d) gamma irradiation specified whey.

In a preferred embodiment, the animal is not a human, selected from the group that includes mammals and birds, preferably poultry, such as chicken, duck, goose, ostrich and quail.

Despite the fact that electrical stimulation can be applied to any part of the body, it is preferable that the step (a) of the method according to the present invention was applied on the head, neck, body and/or one or more limbs of the animal. Of them, preferably, the electrical stimulation is subjected to the animal's head. In the context of the present invention, the term electrical stimulation and shock treatments are used interchangeably.

In a preferred embodiment of the method according to the present invention, electrical stimulation is performed during the interval of time between 1 and 60 seconds, preferably between 1 and 30 seconds and, more preferably, between 2 and 10 seconds. Also preferably, electrical stimulation is performed with a voltage in the range between 50 and 150, preferably in the range between 80 and 120, and more preferably in the range between 110 and 120 C. With the implementation of electrostim is acii preferred are some current values, and, preferably, the electrical stimulation is performed with amperage in the range between 0.01 A and 0.4 A, preferably, in the range between 0.02 A and 0.1 A, and, more preferably, in the range between 0.04 A and 0.06 A.

In a preferred embodiment of the method according to the present invention, electrical stimulation is performed with a frequency in the range between 10 and 200 Hz, preferably in the range between 20-100 Hz, and more preferably in the range of between 45-65 Hz.

In yet another preferred embodiment of the method according to the present invention, gamma irradiation is brought to the absorbed radiation dose between 10 and 40 kGy, preferably, 15 and 35 kGy, and more preferably, between 20 and 30 kGy. The gamma radiation source may be any source, however, the preferred source of gamma radiation selected from the group consisting of60Co.,137Cs67Cu67Ga111In192Ir,99mTc andl70Tm.

In a more preferred embodiment of the method according to the present invention the method further involves the step of incubating the specified blood before step c).

In a more preferred embodiment of the method according to the present invention the method further involves the step of freeze drying the specified serum before step d).

In a preferred embodiment, methods for the and the present invention blood is an arterial and/or venous blood.

Another aspect of the present invention is a biologically active blood serum, produced in accordance with the method of the present invention.

An additional aspect of the present invention is a pharmaceutical composition containing the serum of the present invention and one or more pharmaceutically acceptable diluents, carriers, excipients, including fillers, binders, lubricants, slip agents, dezintegriruetsja agents, adsorbents and/or preservative.

In a preferred embodiment, the pharmaceutical compositions of the present invention the composition is prepared in the form of a syrup, solution for infusion or injection, tablets, capsules, solid tablets in the form of capsules, tablets, liposomes, suppositories, plasters, bandage, capsule slow-release powder or drug slow release. Preferably the diluent is a water buffer, buffered saline, or saline, and the medium, preferably selected from the group comprising cocoa butter and vitebesole.

An additional aspect of the present invention is the application of the blood serum of the present invention or pharmaceutical compositions of the present invention to obtain drugs for the treatment of the Oia disease or condition, which may be caused by the increase of cyclic adenosinemonophosphate acid in the brain of the patient.

Another aspect of the present invention is the application of the blood serum of the present invention or pharmaceutical compositions of the present invention to obtain drugs to improve cognitive and/or academic skills, in particular, to improve long-term memory.

Another aspect of the present invention is the application of the blood serum of the present invention or pharmaceutical compositions of the present invention to obtain drugs for the treatment of seizures, particularly epileptic seizures.

An additional aspect of the present invention is the application of the blood serum of the present invention or pharmaceutical compositions of the present invention to obtain drugs for the treatment of proliferative diseases and apoplexy.

In a preferred embodiment, application of the present invention, the proliferative disease is selected from the group comprising maligna gastrointestinal or colorectal tract, liver, pancreas, kidney, bladder, thyroid, prostate, endometrium, cervix, ovary, uterus, testes, skin is, the oral cavity; melanoma; dysplastic oral mucosa; invasive cancer of the oral cavity; small cell and non-small cell lung carcinomas; breast tumors, in particular, hormonesensitive cancerous breast tumors and gormonalnozavisimyh cancer breast cancer; transitional cell and squamous cell cancers; neurological malignancies, including neuroblastoma, glioma, astrocytoma, osteosarcoma, meningiomas; soft tissue sarcoma; hemangiomas and tumors of the endocrine glands, in particular, pituitary adenomas, pheochromocytoma, paraganglioma, hematological malignancies, particularly lymphoma and leukemia.

In addition, in another preferred embodiment, application of the present invention proliferative disease contains cells that is similar to the line Jurkat lymphoma cells T-cells, lines Raji lymphoma cells In human cell lines Bro of human melanoma cell lines HeLa cancer cells human cervical, line MCF-7 adenocarcinoma cells human line Mg63 osteosarcoma cells, line HT1080 cells fibrosarcoma, line IMR-32 neuroblastoma cells and lines HepG2 cells hepatocarcinoma.

In an additional preferred embodiment, application of the present invention the drug is the first tool is administered to the patient in a quantity changing from 50 to 150 mg/kg body weight, preferably varying from 90 to 100 mg/kg body weight.

Detailed description of the invention

The authors of the present invention unexpectedly discovered that stimulation of the animals, particularly chickens, electric shock, and additional processing of the sera obtained from the blood, γ-radiation leads to a significant increase in biological activity of the product of serum. The resulting products can have a positive effect on various functions of the body, condition and disease of the patient.

Beneficial effect of serum treated with electroshock and γ-radiation, is unexpected for two reasons. First, in the prior art it is known that shocks cause severe violations of all vital functions and systems in the body, particularly the Central nervous system, blood and the circulatory system and the respiratory system (see, for example, Orlow, A.N. et al. (1977) Medicine). Secondly, it was also found that the blood and serum obtained from the blood that is sensitive to radiation and is unstable, and is inactivated when ionizing radiation (see, for example, Radiomedicine - M Atomisdat (1972) 123-125 and Gergely, 3. et al. (1967) Radiosterilization of Medical Products 115-124).

These studies did not lead to the results of the present invention and one could assume that the returns the biological active blood serum by combining processing Electrochem and γ-radiation is impossible. Thus, the surprise was that from the arterial and/or venous blood taken from an animal, in particular the chicken treated with electroshock II-III level and gamma radiation, it is possible to obtain biologically active blood serum. The specific activity of the thus obtained blood serum are described in more detail below.

Accordingly, the first aspect of the present invention is a method of obtaining biologically active blood serum, providing steps:

a) electrical stimulation of the animal, not a man,

b) taking blood from a specified animal,

c) allocation of serum from a specified blood, and

d) gamma irradiation specified whey.

In the method of the present invention can be used in a variety of animals, however, preferably an animal, not a human, selected from the group that includes mammals and birds. Because of availability, in particular, it is preferable to use farm animals, such as poultry, such as chicken, duck, goose, ostrich and quail. Particularly preferred animals, which can be used in the method according to the present invention, is chicken. The type of mammal that can be used in the method according to the present invention is not specifically limited and includes, without limitation, grisone is, for example, a mouse, hamster and rat, cats, dogs, horses, donkeys, sheep, cows and goats.

It is assumed that the electrical stimulation leads to the release in the body of the animal certain compounds that cause and/or contribute to unexpected therapeutic effect of biological active blood serum of the present invention. Animal, not a person, can be stimulated in various areas of the body. Preferably, electrical stimulation is performed on the head, neck, body and/or on one or more limbs. You can stimulate the body only in one place or in several places at once. Especially preferred part of the body for stimulation is the head of the corresponding animal. When stimulation birds, particularly chickens preferably, the electrical stimulation expose the head.

Electrostimulation can be made known from the prior art methods, preferably with the use of metal electrodes or water baths, which, for example, use culling of cattle or killing poultry electric shock. Preferably, the electrical stimulation is performed during the interval of time between 1 and 60 seconds, preferably between 1 and 30 seconds, more preferably between 2 and 10 seconds and, most preferably, between 3 and 4 cut the DAMI. The interval time is longer in case of electrical stimulation is subjected to a large animal, and may be less if the stimulation is subjected to small animals. For example, to stimulate chickens, in particular, the preferred time interval of stimulation is between 2 and 10 seconds, and more preferably between 3 and 4 seconds. Other parameters that can be modified during the electrical stimulation of the animal, are the voltage, current and frequency, and the authors of the present invention has identified some preferred ranges for these parameters. Selected current setting depends partly on the size of the animal, as well as from the body of the animal, which stimulates. Essentially, larger animals and larger areas require a higher voltage and amperage. Thus, electrical stimulation is preferably performed with a voltage in the range between 50 volts and 150 volts, preferably from 80 volts to 120 volts and more preferably between 110 volts and 120 volts. The ranges for the current that can be applied are between 0.01 A and 0.4 A, preferably between 0.02 A and 0.1 A, more preferably between 0.04 and A 0,06 A and, most preferably, about 0.05 A. the Voltage, current, and time of application, preferably, selected to send the CN is rgii in the range between 1 and 1000 watts·, preferably, in the range from 10 to 200 watts·s and even more preferably in the range from 15 to 100 watts·C.

To stimulate your preferred animal, i.e. chicken, it is preferable to perform electrical stimulation with a voltage in the range between 80 volts and 120 volts, and more preferably between 110 volts and 120 volts. In addition, the current strength in the range between 0.04 and A 0,06 A, in particular, 0.05 A is preferred with respect to the stimulation of poultry, particularly chicken.

In a particularly preferred embodiment of the method according to the present invention electrostimulation poultry, particularly chicken, runs for 3-4 seconds at a voltage of between 80 and 120, in particular, 110 and 120 C. In this preferred embodiment, the current preferably is between 0.04 and A 0,06 A and, most preferably, about 0.05 A.

Apparently, the frequency of stimulation is not particularly critical, but preferably is in the range between 10 and 200 Hz, more preferably in the range between 45 and 65 Hz, and most preferably is about 50 Hz.

Gamma-irradiation of serum during phase d) of the method according to the present invention can be performed using any of a gamma source, including sources of x-rays and radionuclides. Preferably the source of gamma radiation is the FDS is th radionuclide specific nature of gamma radiation. Preferred sources of gamma radiation selected from the group including60Co.,137Cs67Cu67Ca111In192Ir,99mTc and170Tm. For the authors of the invention in the method of the present invention is particularly preferred sources of gamma radiation are60Co.,137Cs192Ir and170Tm and, most preferably, is60Co. The absorbed serum radiation dose is in the range between 10 and 40 kGy, preferably, in the range between 15 and 35 kGy, and more preferably in the range between 20 and 30 kGy, 25±5 kGy.

Taking the blood of the animal can be carried out by any method known in the art and includes the capture with a syringe, and the puncturing arteries or veins, or decapitation, in particular, in relation to the taking of blood from birds. You can take only part of the whole blood or the blood of an animal. The latter is preferable to use, if the animal was applied a lethal dose of electricity. Taken blood can be arterial and/or venous.

Serum can be isolated from the blood by any known method, including filtration, sedimentation and centrifugation. However, it is preferable incubation of blood within 4-72 hours at a low temperature, for example between 2 and 10°C, preferably between 4 and 8°C, to ensure the treatment of blood clotting, which leads to the release into the blood of additional factors. Thus, in a preferred embodiment of the method according to the present invention the method further involves the step of incubating the blood after taking blood from the animal and to highlight the serum from the blood, for example, within 4-72 hours at a low temperature, for example between 2 and 10°C, preferably between 4 and 8°C.

In an additional preferred embodiment of the method according to the present invention the method comprises the additional step of freeze-drying serum before irradiation (d). Lyophilization provides easier handling serum during irradiation and optimizes the absorption of radiation by components of serum.

An additional aspect of the present invention is directly biologically active blood serum, which is obtained by the method according to the present invention. It differs from serum prior art, upon receipt of which do not use the stages of the method according to the present invention, it proved a specific therapeutic effect, which does not show the products of the blood serum of the prior art.

Because it was unexpectedly discovered that the biologically active blood serum of the present invention provides ODA is divided therapeutic effect, for example antiproliferative activity or anti-epileptic activity, an additional aspect of the present invention is a pharmaceutical composition comprising a biologically active blood serum obtained in accordance with the method of the present invention. Such pharmaceutical composition may further comprise one or more pharmaceutically acceptable diluents; media; forming, including fillers, binders, lubricants, glidant, disintegrant and adsorbents; and/or preservatives.

The pharmaceutical composition of the present invention may be of various well known ways, including oral and parenteral administration, e.g. intravenous, intramuscular, intranasal, intradermal, subcutaneous and other routes of administration. Preferred are injecting and certain intravenous. Depending on the route of administration requires a variety of pharmaceutical drugs, some of which may require the application of drug of protective coatings to prevent splitting of biologically active serum, for example, in the digestive tract.

Thus, preferably, the pharmaceutical composition of the present invention receive the IDA syrup, solution for infusion solution for injection, tablets, capsules, solid tablets in the form of a capsule, solid tablets in the form of capsules, tablets, liposomes, suppositories, plasters, bandage, capsule slow-release powder or drug slow release.

Special preferred pharmaceutical forms are forms that are suitable for use in injection include sterile aqueous solutions or dispersions and sterile powders for the preparation of sterile solutions or dispersions for injection according to the individual recipe. In all cases, the final form of a solution or dispersion must be sterile and fluid. Typically, such a solution or dispersion includes a solvent or dispersion medium containing, for example, water-water buffer solutions, for example biologically compatible buffers, ethanol, polyhydric alcohol such as glycerin, propylene glycol, polyethylene glycol, suitable mixtures, surfactants or vegetable oil. Biologically active blood serum of the present invention can also be obtained in liposomes, in particular for parenteral administration. The advantage of liposomes is increased half-life from blood flow compared with the free drug and the even more prolonged release of the detainees in her medicines.

Sterilization of solutions for infusion or injection may be performed by any method adopted in this area, including, but not limited to, additives preservatives, such as antibacterial or antifungal agents such as parabens, chlorobutanol, phenol, sorbic acid or thimersal. Additionally, solutions for infusion or injection can be included isotonic agents such as sugars or salts, in particular sodium chloride.

Obtaining sterile solutions for injection that contains the biologically active blood serum is produced by combining the biologically active serum in a predetermined amount in an appropriate solvent, if necessary, with various ingredients enumerated above, followed by sterilization. If necessary to obtain a sterile powder of the above solutions are subjected to vacuum drying or freeze-drying. Preferred diluents of the present invention are water, physiologically acceptable buffers, a physiologically acceptable buffered saline solutions or salt solutions. The preferred media of the present invention are cocoa butter and vitebesole. Excipients that can be used in various pharmaceutical forms of the biologically active blood serum can be selected from abusage a non-limiting list:

a) binders, such as lactose, mannitol, crystalline sorbitol, dibasic phosphates, calcium phosphates, sugar, crystalline cellulose, carboxylmethylcellulose, hydroxyethylcellulose, polyvinyl pyrrolidone and the like;

b) lubricants such as magnesium stearate, talc, calcium stearate, zinc stearate, stearic acid, hydrogenated vegetable oil, leucine, glycerides and stearylamine sodium

c) dezintegriruetsja agents, such as starches, croscarmellose, nutrimetics, agar, bentonite, alginic acid, carboxylmethylcellulose, polyvinylpyrrolidone, etc.

Other suitable excipients can be found in the guidance Handbook of Pharmaceutical Excipients, published by American Pharmaceutical Association, which is included in this application by reference.

An additional aspect of the present invention is the use of serum or pharmaceutical compositions of the present invention to obtain drugs for the treatment of a disease or condition which may be caused by the increase of cyclic adenosinemonophosphate acid in the brain of the patient.

Another aspect of the present invention is the use of biologically active blood serum or pharmaceutical composition of the present is obreteniyu for the production of medicines to improve nootropic, cognitive and/or academic skills of the patient and, in particular, to improve long-term memory. The applicability of this evidence is based on the discovery that serum or pharmaceutical composition of the present invention enhance the learning abilities of the patients.

In addition, serum or pharmaceutical composition can be used for the treatment of seizures of any type, in particular for the treatment of epileptic seizures. In particular, in severe forms of epilepsy and for large epileptic seizures introduction of biologically active blood serum or pharmaceutical compositions of the present invention can prevent death, which in some cases is associated with severe attacks. In connection with the observation that the present invention can be used for the treatment of epileptic seizures, it was also found that the blood, serum or pharmaceutical composition according to this invention can be used for the treatment of nervous disorders, including but not limited to, bipolar disorder, depression, anxiety disorder, epilepsy, Alzheimer's disease, Parkinson's disease, peripheral neuropathy, amyloid angiopathy brain, neurodegenerative disorders and damage to the spinal cord.

An additional aspect of this image is to be placed is the use of serum or pharmaceutical compositions of the present invention to obtain drugs for the treatment of proliferative diseases and apoplexy. Particularly unexpected was the fact that the biologically active blood serum or pharmaceutical composition of the present invention indeed showed a marked antiproliferative effect when tested on different lines of tumor cells in vitro. In this regard, it seems that the biologically active blood serum or pharmaceutical composition of the present invention, in particular, is effective for the treatment of proliferative diseases, and preferred proliferative disease that can be treated using the present invention, selected from the group comprising malignoma gastrointestinal or colorectal tract, liver, pancreas, kidney, bladder, thyroid, prostate, endometrium, cervix, ovary, uterine, testicular, skin, oral cavity; melanoma; dysplastic oral mucosa; invasive cancer of the oral cavity; small cell and non-small cell lung carcinoma; tumors of the breast, in particular, hormonesensitive cancerous breast tumors and gormonalnozavisimyh cancer breast cancer; transitional cell and squamous cell cancer; neurological malignant tumors, including neuroblastoma, glioma, astrocytoma, osteosarcoma, Maine is nyomi; soft tissue sarcoma; hemangioma and tumors of the endocrine glands, in particular, pituitary adenomas, pheochromocytomas, paragangliomas, hematological malignancies, particularly lymphoma and leukemia.

As antiproliferative action of biologically active blood serum of the present invention or pharmaceutical compositions of the present invention has primarily been created for the different cell lines tumors, it is particularly suitable for the treatment of proliferative diseases, which contain cells and/or tumor tissue containing cells, such lines of tumor cells used in these experiments. Accordingly, the preferred proliferative diseases that are treated with the biologically active blood serum or pharmaceutical compositions containing biologically active serum that contain cells that is similar to the line Jurkat lymphoma cells T-cells, lines Raji lymphoma cells In human cell lines Bro of human melanoma cell lines HeLa cancer cells human cervical, line MCF-7 adenocarcinoma cells human line Mg63 osteosarcoma cells, line HT1080 cells fibrosarcoma, line IMR-32 neuroblastoma cells and lines HepG2 cells hepatocarcinoma. In this context, the term "similar cells" refers to cells that have the same p is kishorganj, for example, T-cell, b-cell, or series of generations of nerve cells and the corresponding cell line, and which carry a mutation in the same or a functionally equivalent gene, and where this mutation contributes to the proliferative activity of cells, for example a mutation in p53, pRb, cdc 2, cdk 4, cyclin A, cyclin B, p21ras, c-fos, c-jun, p107, p130, and the like; which are the same or functionally equivalent to the exogenous gene, such as human papilloma virus (HPV), E 6 and E 7, the insertion of hepatitis b virus in the promoter cyclina and the like; or which contain the same chromosomal rearrangement or violation, that is, a deletion, a plurality of chromosomes, etc.

On the basis of results obtained in animal and cell culture, preferred are a certain number of biologically active blood serum for the treatment of diseases and conditions which can be used the serum and pharmaceutical composition. However, it should be understood that for the detection of therapeutic action, depending on the state, as well as from the respective patient, i.e. depending on the severity of the disease or condition, General health of the patient, etc. needed by the various doses of the biologically active blood serum or pharmaceutical compositions. Determining the appropriate dose is on the discretion of the attending physician. It is expected that the dosage of the biologically active blood serum in therapeutic method of the invention should be in the range from about 0.1 mg to about 200 mg of whey per kg of body weight. However, in the preferred application of the present invention the biologically active blood serum is administered to the individual to whom it is required, in the range from 50 to 150 mg/kg body weight, preferably in the range from 90 to 100 mg/kg body weight. Duration of therapy biologically active blood serum varies depending on the severity of the disease, and status, and idiosyncrasy of each individual patient.

The following examples are given to demonstrate the preferred embodiment of the invention. Specialists in this field should take into account that the techniques disclosed in the following examples are methods for which the inventors have found that they function well in the practice of the invention and, therefore, they can be considered the preferred option of this practice. However, in light of the present description specialists in this field should take into account that in particular disclosed embodiments, the execution can be done multiple changes without derogating from the meaning and scope of the invention as set forth in the attached claims. All links on the us in this application as references.

Brief description of figures

Figure 1 - the cytotoxic effect of two pharmaceuticals in Jurkat cells.

Cytotoxic effect on the line Jurkat lymphoma cells T-cells displayed for two different pharmaceutical preparations containing different concentrations of biologically active serum in accordance with the invention. The viability of Jurkat cells displayed on the Y-axis in percent, while the number of biologically active serum is indicated on the X-axis in mg/ml

Figure 2 cytotoxic effect of two pharmaceutical drugs on Raji cells.

Cytotoxic effect on the line Raji lymphoma cells B-cells displayed for two different pharmaceutical preparations containing different concentrations of biologically active serum in accordance with the invention. The viability of Raji cells displayed on the Y-axis in percent, while the number of biologically active serum is indicated on the X-axis in mg/ml

Figure 3 cytotoxic effect of two pharmaceutical drugs on cells Bro B-19.

Cytotoxic effect on the line Bro B-19 cell lymphoma T-cells displayed for two different pharmaceutical preparations containing different concentrations of biologically active serum in accordance with the invention. The viability of cells is to Bro B-19 displayed on the Y-axis in percent, while a number of biologically active serum is indicated on the X-axis in mg/ml

4 is a cytotoxic action of two pharmaceuticals in HeLa cells.

Cytotoxic effect on the line HeLa cell lymphoma T-cells displayed for two different pharmaceutical preparations containing different concentrations of biologically active serum in accordance with the invention. The viability of HeLa cells displayed on the Y-axis in percent, while the number of biologically active serum is indicated on the X-axis in mg/ml

5 is a cytotoxic action of two pharmaceutical drugs on cells MCF-7.

Cytotoxic effect on the line MCF-7 cell lymphoma T-cells displayed for two different pharmaceutical preparations containing different concentrations of biologically active serum in accordance with the invention. The viability of MCF-7 cells displayed on the Y-axis in percent, while the number of biologically active serum is indicated on the X-axis in mg/ml

6 is a cytotoxic action of two pharmaceutical drugs on cells IMR-32.

Cytotoxic effect on the line IMR-32 cell lymphoma T-cells displayed for two different pharmaceutical preparations containing different concentrations of biologically active whey in with the accordance with the invention. The viability of cells IMR-32 displayed on the Y-axis in percent, while the number of biologically active serum is indicated on the X-axis in mg/ml

Fig.7 - cytotoxic effect of two pharmaceuticals in HT1080 cells.

Cytotoxic effect on the line HT1080 cell lymphoma T-cells displayed for two different pharmaceutical preparations containing different concentrations of biologically active serum in accordance with the invention. The viability of HT1080 cells displayed on the Y-axis in percent, while the number of biologically active serum is indicated on the X-axis in mg/ml

Fig - cytotoxic effect of two pharmaceuticals in HepG2 cells.

Cytotoxic effect on the line HepG2 cell lymphoma T-cells displayed for two different pharmaceutical preparations containing different concentrations of biologically active serum in accordance with the invention. The viability of HepG2 cells displayed on the Y-axis in percent, while the number of biologically active serum is indicated on the X-axis in mg/ml

Fig.9 - "bell-shaped" curve of the proliferative activity of cells treated with mitogens.

The curve of the proliferative activity of lymphocytes was analyzed based on the activity of DNA biosynthesis in relation to concentric and mitogen. The activity of DNA biosynthesis measured by the number of pulses per minute of incorporated radioactivity and insoluble acid.

Figure 10 - mitogenic activity of pharmaceutical compositions.

Displayed mitogenic effect of the pharmaceutical composition on the activity of DNA biosynthesis in the concentration range of the substance from 0.1 to 100.0 mg/ml the Activity of DNA synthesis was evaluated on the basis of the amount of radioactivity incorporated into DNA.

11 - effect of pharmacological compositions to cells MCF-7.

Displays the activity of DNA synthesis in line MCF-7 breast cancer cells human relative to the amount of biologically active serum that is incorporated into pharmaceutical compositions.

Examples

Example 1

The method of obtaining blood chicken, subjected to electroshock

To obtain serum chicken the chicken was treated with electroshock II-III level (voltage 80-120 In current of 0.05 A, frequency 50 Hz, the time of application with 3-4, on the head). Blood was taken from the carotid artery and additionally incubated at a temperature of 4-8°C for 18-24 hours in plastic flasks. After complete removal of blood clots flask was centrifuged at 3000 rpm for 20-30 minutes. Serum was separated from blood clots and liofilizirovanny under conditions known in the art. Flasks with liofilizirovannami processed in the instrument RZ-100-M 20-30 kGy, preferably, in the range of 25 kGy using as a source of gamma radiation60Co. The treated serum was stored at a temperature of between 4-8°C for later use.

The evidence of the stimulating action of the serum of chicken treated with electroshock

The following experiments were performed using male Wistar rats with an average body weight 280-300, Animals randomly divided into 4 groups of 10 rats each. The first group was injected with 1.0 ml of saline. The second group was injected dose serum 100±5.0 mg/kg body weight according to the present invention in 1.0 ml. Upon expiration of 30 minutes after the injection, rats were decapitated. Rats of the third group were injected with 1 ml saline, and rats of the fourth group received a biological active serum (100±5 mg/kg body weight) in solution in a volume of 1.0 ml after 30 minutes after injection the animals, which were attached cargo to the tail (10% of body weight in rats), were placed in a bowl of water (25°C). After the first symptoms of the suffering of the animals were removed from the water and was beheaded.

As samples for further analysis took the brain, heart, liver (bodies subjected to extensive power voltage in the processes of extreme adaptation), and skeletal muscle (as a body subjected to the most severe hcpa is setiu). Samples of tissue from each rat was weighed, cooled isotonic NaCl and quickly frozen in liquid nitrogen. Full time elapsed between the application of "stress" and the final processing of the samples was as a maximum in the range between 5-6 minutes.

Determined the amount of adenosine triphosphate, by acid and adenosinemonophosphate acid in skeletal muscle of rats. Nucleotides were separated by ion-exchange chromatography columns using Anionit Dowex 1. The determination of the number of adenosine triphosphate, by acid and adenosinemonophosphate acid was performed spectrophotometrically in the range of 256 nm (spectrophotometer Hitachi-557). The energy potential was determined according to the following formula

(ATP-0,ADP)/(ATP+ADP+AMP)

The determination of the number of cyclic adenosinemonophosphate acid in the brain, heart and liver was performed using known radioimmunological analysis device for detection Amersham (UK).

The definition of labeled adenosinemonophosphate acid was performed using methods known in the prior art, using a scintillation counter GS-8.

Results

The difference of the amounts of adenosine triphosphate, by acid and adenosinemonophosphate acid and withdrew the giving of the energy potential are obvious, compared with the third group or the control group or the second group (p<0,05) are obvious, as well as when comparing the fourth group or the control group or the second group (p<0.05) and, in addition, when comparing the third and fourth groups with each other (p<0,05) (see also table 1).

Table 1
The amount of adenosine triphosphate, by acid and adenosinemonophosphate acid and the value of the energy potential in the tissue of the skeletal muscles with the introduction of biologically active serum and without it
A group of ratsThe number of nucleotides (µmol/l/g tissue)Energy potential (relative ratio)
h M±Mh M±Mh M±M
I (saline control) n=107,55±0,19 (7,28-7,85)0,95±0,12 (0,79-1,18)0,25±0,08 (0,14-0,37)0,917±0,080
II (serum) n=107,89±0,12 (7,65-8,02)1,12+0,11 (0,93-1,27) 0,14±0,05 (0,08-0,25)0,923±0,125
III (saline + dipping)n=101,34±0,08 (1,19-1,49)3,56±0,17 (3,21-3,81)0,78±0,07 (0,65-0,93)0,549±0,014
IV (serum + dipping) n=104,78±0,17 (4,35-4,95)2,55±0,11 (2,36-2,72)0,33±0,09 (0,19-0,48)0,790±0,095

The number adenozintrifosfornoy acid defined during the active period and 30 minutes after moderate stress associated with injection (control group I), confirms that under the influence of serum in muscle tissue is the mixing amount of adenosine triphosphate and by acid when the values of the upper standards, while the number adenosinemonophosphate acid decreases.

These results can be interpreted so that the biologically active serum increases the energy potential in the muscle tissue. The calculation of the energy potential confirms this trend.

Under the influence of extreme stress dipping in the third group (saline + dipping) was determined to decrease adenosine fosforos acid content increased by and adenosinemonophosphate acid compared to the control group.

Under extreme stress rats of the fourth group (serum + dipping) General trends in quantities of ATP, diphosphorous and monophosphoric acids, observed in the third group remained similar, however, the amount of adenosine triphosphate remained significantly large, and when calculating the energy potential of increased energy capacity by 43% (p<0.05) as compared with the third group.

Comparative analysis of cyclic adenosinemonophosphate acid in the tissue of the heart and liver, and brain tissue of rats of the first and second groups showed that under the influence of serum in the brain tissue can be detected by increasing the number of cyclic monophosphoric acid (p<0,01), while in the tissue of the heart and liver of a significant difference compared with the control group was not identified (see table 2).

Table 2
The number of cyclic adenosinemonophosphate acid AMP in the tissue of the brain, heart and liver after administration of biologically active serum
A group of ratsThe number of cyclic adenosinemonophosphate acid (pmol/l/g raw fabric)
brain M±Mheart M±Mliver M±m
I (saline control) n=103,95±0,58 (3,32-4,55)3,07±0,11
(2,98-3,32)
2,75±0,18
(2,47-2,99)
II (serum) n=104,58±0,23 (4,26-4,85)3,26+0,16
(3,02-3,47)
2,98±0,99
(2,95-3,12)
III (saline + dipping) n=100,82±0,13 (0,66-1,05)0,49±0,17
(0,32-0,78)
1,01±0,07
(0,86-1,14)
IV (serum + dipping) n=101,58±0,13 (1,38-1,76)0,83±0,14 (0,56-0,96)1,27±0,08
(1,12-1,41)

Under the influence of extreme stress in all tissues of rats of the third group (saline + dipping) and fourth group (serum + dipping) there was a significant decrease (p<0,05) number of cyclic adenosinemonophosphate acid. However, this decrease has fallen under the influence of biologically active serum. Accordingly, the number adenosinemonophosphate acid in brain tissue was 92% higher than the corresponding number in the third is the group (p< 0.05), and 96% greater in the tissue of the heart and 25.7% higher in liver tissue.

Therefore, serum in accordance with the present invention, treated with electroshock and γ-radiation, increases the energy potential in skeletal muscle of rats, increases the amount of cyclic adenosinemonophosphate acid in brain tissue during phases of rest and during extreme physical stress, and contributes to the increase of cyclic adenosinemonophosphate acid in the tissue of the heart and liver after physical stress in rats.

Example 2

Long-term memory

In this experiment used a 150 male Wistar rats. All rats pre-tested using classical test "open field" and distributed in accordance with the activity into three groups: active, with an average active and passive.

To create reflex situation rats were placed in the cylinder, which was located in a regulated with a thermostat pool. The cylinder was placed on three legs, and he allowed the animals to dive under the bottom edge of the cylinder to reach the platform, located outside of the cylinder.

Determined the latent period for the first experiment, and also the latent period of the final solution to the problem of "release" from the closed room and out of the water.

When establishing reflex situation kr who were divided into three groups:

- "quickly emerging" - those rats who were able to find out within the first minute.

- "slowly emerging" - those rats, which only began to seek a way out after 2-3 minutes;

- rats who did not want to solve the problem within 10 minutes.

Biologically active serum was injected into the abdominal cavity for 30 minutes before the start of the experiment at a dose of 100 mg/kg body weight. Control animals were injected with 1 ml of physiological solution. The next day, tested the behavior of the reflex situation and subsequently, the rats were given a break in 40 days.

In accordance with this method have been "trained" 120 rats - 60 control rats and 60 rats receiving biologically active serum. Rats pre-divided into three groups, as indicated above, that is active, with an average active and passive (20 animals in each group), in addition, for data analyses they are divided into "fast" and "slow" emerging.

Results

Some rats did not show the reflex of surfacing" (table 3). In the control group six of the number of active rats and rats with medium activity and 12 animals passive group generally did not show a reflex of surfacing". On the second day, this number remained the same for the active rats and in rats with medium activity, however, decreased for passive animals up to 7. After 42 d is it the same 11 rats passive group refused to dive under the edge of the cylinder (1 rat died during the experiment).

Among the active rats receiving biologically active serum, there were no rats, which refused to solve the problem on the second day. Of all the rats with an average activity of three refused to solve the problem on the first day, but, nevertheless, decided on the second day and retain this ability even after 42 days. Among passive rats receiving serum, 4 rats did not show willingness to solve the problem, but on the second day, this number decreased to 2 and remained constant even after 42 days.

Table 3
The number of rats that did not show a reflex of surfacing"
Test animal1 day (24 hours)
1242
Active rats and rats with medium activityControl666
Serum300
Passive ratTo ntrol 12711∗
Serum422
∗ one rat died during the experiment

In the group "quickly emerging single injection of serum in the 40-day time interval of the experiment did not show significant changes in the maintenance of reflex situation.

For "slowly emerging" was significant, from the point of view of statistics, the ability to maintain the ability to quickly find a way to avoid extreme situations (table 4). Thus, the formation of the reflex on the second day decreased compared with the first day for the control group and the test group. In the control group these reflexes disappeared completely after 42 days, whereas animals that received an injection of biologically active whey, reflex fully preserved. In addition, there was a difference in time required for the reflex, which was 2-3 times more (p<0,01) in control rats.

Table 4
The influence of serum on the formation and maintenance of elexa situation slowly emerging" (C)
Test animal1 day (24 hours)
1242
LP-1 (latent period)LP-2LP-1LP-2LP-1LP-2
The group is activeControl125,7+25176,7+19,645,5+9,959,0+8,837,5+30,0181,5+15,5
Serum111,3+39,7164, 8mm+33,555,3+15,668,4+18,83,8+29,355,7+37,28
Group average activityControl144,8+21,8191,2+16,349,6+11,262+14,33,8+24,019,2+9,8
Serum118,2+71,6178,6+76,83,8+50,377+54,19,2+28,854,6+39,7
The group of passiveControl53,3+25,7189,6+12,653+15,066,6+8,0149,3+24,7196,0+11,1
serum18,2+71,6178,6+76,863,8+50,377+54,139,2+28,854,6+39,7

In the formation and long-term maintenance of information, a significant role is attributed to H-cholinergic mechanisms. 30 animals were trained for five days to dive under the "bell", subdivided into three groups.

Saline tsitizina (H-cholinergic antagonist of the brain) was administered in the dose of 1 mg/kg of body weight in the abdominal cavity for 30 minutes before testing the animals of the first group (ten rats).

Animals of the second group (ten rats) was administered biologically active serum in a dose of 100 mg/kg body weight and 30 minutes later were injected solution tsitizina dose of 1 mg/kg of body weight is eating.

The third group of animals (control - ten rats) received 1 ml injection of saline.

Complex behavior of the animals tested in accordance with the above method after 48 hours after the injection of the indicated substances. Upon the expiration of 48 hours after application tsitizina biological tests showed a marked slow reactions release" from the closed room, namely 3-fold slower compared to control animals.

Introduction serum injections before tsitizina not only reduced the effect of this antagonist, but also led to increased reaction "release" 20% compared with the control group, and the full action of biologically active serum exceeded the action of the antagonist in 5 times.

Table 5
Latent periods reactions of rats to the situation in connection with the introduction of tsitizina and biologically active serum (C)
TimeControl (the third group)Introduction tsitizina (first group)Introduction serum + tsitizin (the second group)
After 48 h13,33±0,5840,0±14 8,3±5,5
<0,05>0,05
<0,05

Based on the results, it was determined that N-kalinovye brain receptors play a role in the recovery "reactions release" in connection with the modeling of complex behavior of animals and that biologically active serum prevents deterioration of the ability to learn.

Thus, the biologically active blood serum stimulates the formation of long term memory and especially shows this effect in animals with delayed reaction release from the closed room and out of the water. In addition, it appears that H-cholinergic mechanisms of the brain play an important role in maintaining long-term memory, and action of substances which may adversely affect H-cholinergic mechanisms of the brain, can resist biologically active serum of the present invention.

Example 3

Epilepsy

In the prior art it is known that the joints in toxic dosage leads to hyperactively motor areas of the Central nervous system, which in turn causes the development of tonic spasms. the connection camphor is used along with carazolol on model animals for inducing spasms. Depending on the dose injected camphor can cause all kinds of small and large epileptic seizure, including large seizures.

In experiments investigated the influence of biologically active serum epileptic activity induced by injection of camphor into the abdominal cavity of rats. In this experiment used 40 male Wistar rats weighing 190-210 g Solution camponovo oil (20%) was injected into the abdominal cavity in the amount of 0.25 and 0.5 ml (using 20 rats). Serum was administered at a dosage of 100±5.0 mg/kg body weight (using 20 rats).

Experts observe and assess attacks. Evaluated the following parameters. The latent period of reaction, the type of reaction spasm (toniceski-clonic, large and small seizures), duration epileptic seizure and the time intervals between them, the loss of normal mobility and the end result (the death of the animal, or restore from a pathological condition). The results are summarized in table 6.

Table 6
The action of biologically active serum on experimentally induced epilepsy
Doses of camphorEpileptic seizuresThe control group (Campo is a) The test group (camphor + serum at a dose of 100 mg/kg)
0,25The occurrence of tonic spasms7'20"±10"5'30"±15"
The occurrence of clonic spasms in the form of large seizures after12'30"±32"8'45""±28"
The average number of seizuresOne 6'-7'One 8'-9'
The duration of seizures70'-80'15"-20"
The condition of the animal after 48 hoursAll survivedAll survived
0.5 mlThe occurrence of tonic spasms5'40"±30"5'00"-32"
The occurrence of clonic spasms in the form of large seizures after8'30"±27"8'40"
The average number of seizuresOne in 5'The duration of seizures90"-120"25"-30"
The condition of the animal after 48 hours60% diedAll survived

Biologically active serum, injected at a dose of 100 mg/kg in the latent period of epileptic seizures caused by injection of camphor, reduced all symptoms activity of the attack; the latent period of attack was increased, was less severe clonic spasms and remained normal mobility and additionally serum treated with electroshock, prevented the death of the animal model of severe epilepsies (all animals survived the introduction of 0.5 ml of 20% solution of camphor).

Example 4

The action of the serum of the present invention on the proliferation of human cells

The serum of the present invention is a freeze-dried chicken blood, treated with electroshock II-III level and γ-radiation. Biologically active serum used in the two types of drugs: (i) serum, resuspendable in water at a concentration of 100 mg/ml (tested fraction 1), and (ii) supernatant suspension 100 mg/ml of biologically active whey in water after three minutes centrifuge holds the investments of the suspension at 10,000×g (test fraction 2).

Cell culture

Cell lines Jurkat and Raji, and lymphocytes in human peripheral blood were cultured in plastic plates for tissue culture (Nunc or Falcon) in the medium 1640-RPMI (Sigma)containing 10% fetal calf serum (FCS, Gibco), 100 units/ml penicillin and 100 μg/ml streptomycin at 37°C with 5% content of CO2and 95% humidity. Cell line Bro, HeLa, MCF-7, Mg63, HT1080, IMR-32 and HepG2 were cultured as described above, but using DMEM medium (Sigma) instead of medium 1640-RPMI.

The selection of mononuclear leukocytes (ML) according to the method of Boyum

Mononuclear leukocytes were isolated according to the method described by Boyum A. (Isolation of mononuclear cells and granulocytes from human blood, 1968, Cand. J. Lab. Clin, Invest, 120 (97): 9-18).

15 ml of a solution ficoll-Pak was placed in each of the two conical tubes (Falcon) and then ficoll was added 25 ml of blood, diluted two times in phosphate-buffered saline (PBS). Then the tubes were centrifuged at 400×g and 20°C for 30 minutes in a bucket-rotor. The upper phase containing the plasma is not used.

Mononuclear leukocytes (ML), concentrated at the interface between the plasma and the environment division, gently sucked up with a pipette and collected in a centrifuge tube. After that, the cells were twice washed in PBS by centrifugation of the cells at 250×g for 10 minutes and, in the end, the, of their suspension in the culture medium. This fraction contained 10-30% monocytes and 80-90% of lymphocytes, which are below called "lymphocytes". One part of the lymphocytes was used to study the mitogenic activity of biologically active serum according to the present invention and the other for the study of cell proliferation. With this purpose, the cell suspension was added phytohemaglutinin at a concentration of 20 mg/ml

Evaluation of the activity of the synthesis of deoxyribonucleic acid (DNA) in cells

To assess DNA synthesis cells were added3H-thymidine and insoluble fractions of the acid was evaluated by the number of pulses included radiation. Briefly, lymphocytes were incubated in 96-well-plates with 200 μl medium containing 200-800×103cells. To each well was added with different concentrations of biologically active serum of the present invention. Two hours before the end of the incubation were added3H-thymidine (1 µci/well, 40 MCI/mmol/l). For radiometric analysis cells were collected on filters using automated devices to collect cells. Soluble products of the acid was washed with 5% triperoxonane acid (H2On) and using a scintillation counter measured the radioactivity of the detained substances. The activity of DNA biosynthesis was measured in counts/min

Determination of the survival rate of cells after and is kupirovaniya with various substances, using MTT test

MTT test was performed as described by Mosmann T. Rapid colorimetric assay for cellular growth and survival: application to proliferation and cytotoxicity assays (1983) J. Immunol Meth. 65: 55 to 63).

To run the test cells were harvested in the logarithmic phase (adhesive cells were collected, when they were filled approximately half die for tissue culture). They were placed in a growing medium in the chamber Goryaeva, believed and then resuspendable in the medium at a concentration of 50-100×106/ml. Solutions, cells were placed in 96-hole dies after adding different concentrations of the tested fractions in a total volume of 100 μl. For counting viable cells in each of the different 96-well plates after incubation were added 50 μl of a solution of bromide 3-4,5-dimethylthiazol-2-yl)-2,5 of diphenyltetrazolium (MTT) in the culture medium. For solution 1 ml MTT MTT stock solution was mixed with 4 ml of culture medium. Preparation of stock solution of MTT was performed by dissolving MTT in PBS (PBS contains 0.01 mmol/l buffer phosphate, pH of 7.4, with 0.15 mmol/l NaCl) with a concentration of 5 mg MTT/ml, followed by filtration through a filter with pore size 0.45 µm. The mother solution was stored at +4°C up to one month. After adding MTT solution die for tissue culture additionally incubated for 4 hours in an incubator under identical conditions. After removal of the nutrient medium following the on stage by suction with a pump in each well was added 150 μl of dimethyl sulfoxide (DMSO) to dissolve the formed blue crystal formazan and record the optical density of the solution in each well, multichannel the spectrophotometer with the reader microplate at a wavelength of 540 nm (Labsystem). Cell viability relative to the concentration of added test fractions 1 and 2, respectively, indicated in percentage of survival of control cells. The data were processed using the software Origin.

Results and discussion

The results are shown in figure 1-8. From these results we can conclude the following: a high concentration of biologically active serum according to the present invention (2.5 to 20 mg/ml) had an inhibitory effect on all cell lines, however, the reaction of cells of different tissue types on the tested fractions 1 and 2 respectively different. Dose IC50(concentration of substance that causes 50% inhibition of cell) varies considerably between cell lines from 2.2 (Jurkat) to >20 mg/ml (Mg63) for test substance 1 and from 3.6 (peripheral blood lymphocytes) to >20 mg/ml (Mg63 and HeLa) for the soluble fractions of the blood serum of the present invention (test fraction 2). For all cell lines the toxicity of the starting substances (tested fraction 1) was higher toxicity of soluble substances (tested fraction 2). However, it should be noted that the toxicity observed for individual cell lines was almost identical for both of the tested fractions (Mg63, Rajii, lymphocytes PE fericelli blood) and also for other (Jurkat, MCG-7, IMR-32), moreover, was the difference IC502-3 times. The sensitivity of cells to the test groups compared with the usual cure for cancer, doxorubicin, displayed in table 7.

Table 7
The sensitivity of the cell lines tested with the use of doxorubicin
Cell lineIC50mmol/ml
Jurkat100
Raji20
Bro B-19-∗∗
HeLa400
MCF-7150
Mg63-∗∗
HT1080-∗∗
IMR-324
HepG2100
Peripheral blood lymphocytes40
Comments:
∗ - mmol/l = 0,58 mg/ml
∗∗ - sensitivity to doxorubicin has not been tested

In addition, what was lodales, in a concentration range of from 0.3 to 3 mg/ml depending on the cell line and the type of substance (i.e. the tested fraction 1 or test fraction 2) the tool had a stimulating effect on some cells. The stimulating action of the serum of the present invention was observed for cells Jurkat, Raji, Bro B-19, Mg63, HT1080 and HepG2. Stimulatory effect was negligible (10, 20, 40%), but was present in both types of test substance. Stimulation was not observed in cells of the cell line HeLa, MCF-7, JMR-32 and peripheral blood lymphocytes. The stimulating action of the test substance in the first form was observed at much lower concentrations than in the soluble fraction (tested fraction 2). Perhaps the reason this promotion is not stimulating proliferation as such, but rather an increase in the intensity of cell respiration, which could also be logged using the MTT method used to assess cell viability. The question was whether the observed stimulatory effect due to stimulation of proliferation or by increasing the intensity of breath, demanded some further investigation.

When studying the interaction between different substances with the immune competent cells, essentially, you must discuss at least two issues:

1. Does the test substance mitogenic activity, i.e. does it the ability to stimulate the proliferation of lymphocytes (in this case, the increase in the number of substances usually leads to increased biosynthesis of DNA of lymphocytes, which can be estimated by including3H-thymidine)?

2. Does the test substance toxic effect (this question is usually addressed by inhibition of proliferation of lymphocytes analyzed by using numbers include3H-thymidine, or by using the vital dye MTT-type lymphocytes, which are pre-stimulated with mitogens)?

In addition, it should be noted that non-activated cells do not divide in culture and that the degree of proliferation only increases with the number of mitogens, added to the culture medium. This is reflected in the increase of radioactively labeled DNA.

The effect of mitogens on lymphocytes in relation to the input doses can be described by the so-called "bell curve" (see Fig.9). The first part of the bell curve represents the range of concentrations of the mitogen, where the increase in mitogen leads to an increase in proliferation (as measured by DNA biosynthesis) and where, therefore, there is a direct correlation between the concentration of mitogen and degree of proliferation. The second is trezek curve (2) shows the effect of saturation, where further increase in the concentration of mitogen does not lead to further increase in the degree of proliferation, i.e. mitogen already showed maximum effect. Cytotoxic activity is not yet observed. Segment (3) shows the concentration range of mitogen where mitogen provides increasing cytotoxic effect on lymphocytes.

Study the mitogenic activity of the tested fractions 1 and 2 was performed in two experiments:

1. The subject of study of this experiment was to determine the validity of the tested fractions on nonactivated lymphocytes human peripheral blood. For this purpose, we determined the correlation between proliferative activity (i.e. activities) with the input doses.

2. The subject of study of this experiment was to determine the validity of the test substance on the activity of DNA synthesis of lymphocytes activated by phytohemagglutinin (20 μg/ml FHA) in the second phase.

In the study of substances mitogenic activity was not observed, i.e. in the range of concentrations of substances of 0.1-100 mg/ml were observed stimulation of biosynthesis of DNA of peripheral blood lymphocytes (figure 10).

Increasing the dose of the tested fractions, the number of peripheral blood lymphocytes was not significantly changed. When evaluating the effects of the tested fractions on lymphocytes, which subsequently, Aktivera the Ali with 20 mg/ml FHA, we observed inhibition of activated lymphocytes examinees factions.

In experiments it was determined that the subjects fraction inhibited the biosynthesis of DNA-stimulated lymphocytes at a concentration of 0.3 mg/ml (11). However, the mechanism of action remain to be determined. In this regard, an important additional experiments to explain the cytostatic and cytotoxic action.

In order to determine how the results obtained in cell culture, relate to the treatment of people, it is important to determine the concentration of a substance in various organs or tissues in experiments on animals. In this context it is important to investigate what different concentrations of biologically active serum according to the present invention can affect DNA synthesis in organs involved in lymphogenesis. Therefore, it is important to determine the concentration of biologically active serum, in particular, in the bone marrow, spleen and thymus of the mouse.

In addition to lymphocyte cytotoxic activity of the serum of the present invention also investigated using cells from the cell line MCF-7 breast cancer person in the monolayer. Due to contact inhibition of proliferation of cells was not observed. In this model it is possible to determine the toxic effect of a substance, it is possible to distinguish cytotoxic effect from the cytostat the economic actions. When using this model it was determined that the serum of the present invention exerted a cytotoxic effect at a concentration of 2.5 mg/ml irrespective of the fact whether the insoluble fraction of the serum of the present invention removed or not, that is, regardless of whether you have used the tested fraction 1 or 2.

It is noteworthy that the toxic effect is markedly decreased when the incubation of the serum according to the present invention with dividing cells, potentially due to the fact that the activity was measured after 24 hours after administration, but not after 72 hours after the injection, in the case of cell division (long-term incubation of cells in the monolayer can cause death control cells).

The above results demonstrate that biologically active serum according to the present invention has as cytostatic effect and cytotoxic activity. However, the cytotoxic activity was less pronounced, as combining cytotoxic and cytostatic activity, despite the fact that it was observed at the same concentration.

United blood serum from animals treated with electroshock II-III level, and subsequently prepared a biologically active substance lead to an increase in the degree of proliferation lines Jurkat, Raji, Bro B-1, Mg63, HT1080 and HepG2 human cells by 10-40% compared with the control group. In HeLa, MCF-7, JMR-32 and peripheral blood lymphocytes of biologically active serum according to the present invention showed a significant inhibition of proliferation of all tested lines cancerous human cells at higher doses (2.5 to 20 mg/ml). The sensitivity of cells to biological active blood serum of the present invention is much greater than the sensitivity to commonly used anti-cancer agent doxorubicin. In addition, when using DNA biosynthesis measured by including the label3H-thymidine, the substance of the present invention has as cytostatic and cytotoxic action.

1. The method of obtaining biologically active blood serum, involving the steps:
a) electrical stimulation of the animal, not a man,
b) taking blood from a specified animal,
c) allocation of serum from a specified blood and
d) gamma irradiation indicated serum, in which the absorbed radiation dose specified gamma radiation is from 10 to 40 kGy.

2. The method according to claim 1, in which an animal, not a human, selected from the group that includes mammals and birds.

3. The method according to claim 2, in which the bird is selected from the group including chicken, duck, goose, Stra the sa and quail.

4. The method according to any one of claims 1 to 3, in which step a) is subjected to electrostimulation (subject) head, neck, body, and/or one or more limbs of the animal, mainly the head.

5. The method according to claim 1, in which electrical stimulation is performed during the interval of time between 1 and 60, preferably between 1 and 30 and more preferably between 2 and 10 seconds

6. The method according to claim 1, in which electrical stimulation is performed with a voltage in the range between 50 and 150, preferably from 80 to 120 and more preferably between 110 and 120 C.

7. The method according to claim 1, in which electrical stimulation is performed with a current strength in the range between 0.01 And 0.4 And preferably between 0.02 and 0.1 a, and more preferably between 0.04 and 0.06 and A.

8. The method according to claim 1, in which electrical stimulation is performed with a frequency in the range between 10 and 200 Hz, preferably in the range of between 20-100 Hz and more preferably in the range of between 45-55 Hz.

9. The method according to claim 1, in which the absorbed dose of gamma-irradiation ranges from 15 to 35 kGy, preferably from 20 to 30 kGy.

10. The method according to claim 1, in which the gamma radiation source selected from the group including60Co.,137Cs67Cu67Ga111In192Ir,99mTc and170Tm.

11. The method according to claim 1, wherein the method further involves the step of incubating the specified blood before step C).

12. The method according to P1, in which the method further involves the step of freeze drying the specified serum before step d).

13. The method according to claim 1, in which the blood is an arterial and/or venous blood.

14. The serum obtained by the method according to any one of claims 1 to 13.

15. Pharmaceutical composition containing serum on 14 and one or more pharmaceutically acceptable diluents, carriers, excipients, including fillers, binders, lubricants, slip agents, dezintegriruetsja agents, adsorbents and/or preservatives.

16. The pharmaceutical composition according to item 15, in which the composition is prepared in the form of a syrup, solution for infusion or injection, tablets, capsules, solid tablets in the form of capsules, tablets, liposomes, suppositories, plasters, bandage, capsule slow-release powder or drug slow release.

17. The pharmaceutical composition according to item 15 or 16, in which the diluent is a water, buffer, buffer saline solution or saline solution.

18. The pharmaceutical composition according to item 15, in which the carrier is selected from the group comprising cocoa butter and vitebesole.

19. The use of serum for 14 or a pharmaceutical composition according to any one of p-18 to obtain drugs for the treatment of the disease or condition to the / establishment, which may be caused by the increase of cyclic adenosinemonophosphate acid in the brain of the patient.

20. The use of serum for 14 or a pharmaceutical composition according to any one of p-18 to obtain drugs to improve cognitive and/or academic skills, in particular to improve long-term memory.

21. The use of serum for 14 or a pharmaceutical composition according to any one of p-18 to obtain drugs for the treatment of epileptic seizures.

22. The use of serum for 14 or a pharmaceutical composition according to any one of p-18 to obtain drugs for the treatment of apoplexy and proliferative disease selected from the group consisting of malignoma liver, malignoma cervix, melanoma malignum of breast cancer, particularly hormone-dependent cancers of the breast and hormone-independent cancers breast cancer; neuroblastoma, soft tissue sarcoma and hematological malignancies, particularly lymphoma and leukemia.

23. The use of serum for 14 or a pharmaceutical composition according to any one of p-18 to obtain drugs for the treatment of proliferative diseases, in which proliferative disease contains cells that is similar to the line Jurkat lymphoma cells T-cells, lines Raji lymphoma cells In human cell lines Bro melanoma cells of man, line HeLa cancer cells human cervical, line MCF-7 adenocarcinoma cells human line Mg63 osteosarcoma cells, line HT1080 cells fibrosarcoma, line IMR-32 neuroblastoma cells and lines HepG2 cells hepatocarcinoma.

24. Use PP-23, in which the drug is administered to the patient in amounts varying from 50 to 150 mg/kg body weight, preferably varying from 90 to 100 mg/kg body weight.



 

Same patents:

FIELD: medicine.

SUBSTANCE: invention refers to medicine, namely to oncology and can be used for therapy of patients suffering from hepatic metastases of gastric carcinoma. The mode is implemented as follows. Celiac trunk is catheterised through femoral artery by Seldinger approach. That is followed with selective catheterisation of common hepatic artery wherein Cisplatin is injected within 24 hours, and Fluorouracil - within the next 72 hours. Therapeutic course includes 4-6 procedures every 4 weeks.

EFFECT: invention use allows downsizing the metastatic foci, extending life span and improving life quality of said patients.

1 ex

FIELD: medicine.

SUBSTANCE: invention concerns immunology area. Versions of the artificial fused protein consisting of an antibody (or its fragment) and cytokine, fused through a link peptide are offered. The antibody or its fragment is chosen from an antibody 225, 425, KS 1/4, 14.18, anti-CDx-antibody where x has the whole value 1-25. Each of versions of the fused protein has lowered quantity T-epitopes, at least, in the component of the fused protein presented by an antibody, and as consequence, possesses the lowered adjuvanticity, in comparison with an initial molecule. Identification of T-lymphocyte epitopes is performed by the automated calculation of sizes for the binding centres of class II MHC molecules with the subsequent experimental test of the obtained versions of protein for presence of the lowered adjuvanticity. The automated way of T-epitopes calculation is based on use of the Bjom's function modified in such manner that contribution of Van-der-vaals repulsion and lipophilic interaction in pairs between all lipophilic atoms of the chosen segments of the fused protein and a binding groove of a MHC P molecule is taken into account. Also a way of protein construction on the basis of the modified function Bjom's function with the subsequent experimental test of the received versions for presence of the lowered adjuvanticity is revealed, and also application of the fused protein for preparation of a pharmaceutical composition for tumour treatment is in addition considered.

EFFECT: invention use allows obtaining the fused proteins with the lowered adjuvanticity and, basically, keeping identical biological activity in comparison with a parent molecule; it can be used in treatment of tumours.

4 cl, 6 dwg, 22 tbl, 19 ex

FIELD: chemistry.

SUBSTANCE: present invention refers to the aminopyridin compound of general formula (I) or its salt wherein X1, X2, X3, Z, Y1, Y2 are carbon or nitrogen atom, R, R1, R5, R6 are hydrogen atom, alkyl group, further see formula of the invention, and R7 is hydrogen or halogen atom, nitro or cyano group, -CpH2(p-1)(Ra1)(Ra2)-O-Ra3, -C(=O)-Rd1, 5-or 6-membered saturated heterocycle group, aromatic heterocycle group, -N(Rh1)(Rh2), further see formula of the invention. The invention refers also to the pharmaceutic composition thereof intended for treatment or prevention of allergic diseases, autoimmune diseases caused by malignant tumour, to the Syk inhibitor containing the compound of formula I and to the therapeutic and/or preventive agent.

EFFECT: compounds which not only possess high Syk inhibition activity but are selective Syk inhibitors are obtained and described.

24 cl, 24 ex, 2 tbl

FIELD: chemistry.

SUBSTANCE: present invention refers to the new naphtylene derivative having general formula (I-A) and to their pharmaceutically acceptable salts having the property of inhibition of the cytochrome ferment P450RAI (Cyp26) activity, to the pharmaceutic composition thereof and to the method of inhibition of cytochrome ferment P450RAI (Cyp26). , wherein X is selected from imidasolyl or triasolyl; R2 and R3, independently represent H, C1-10-alkyl; G1 is -NR72R82 or G1 and R3 taken together with attached carbon atom form 3-10-membered saturated ring or heterocyclic saturated ring containing N as heteroatom which is optionally substituted with substituting group R72, Z, R4b, R5b, Q1, R72, n2, n3 and n4 values are indicated in the formula of the invention.

EFFECT: present invention refers to the intermediates for compounds with general formula (I-A) and to their pharmaceutic salts thereof.

37 cl, 30 dwg, 7 tbl

FIELD: medicine.

SUBSTANCE: group of inventions concerns medicine, namely to the agents strengthening effect of processing by focused ultrasound of high intensity (HIFU). The agent, strengthening effect of HIFU processing, intended for HIFU-therapy, represents a substance which can reduce level of HIFU energy in a target site (EEF), containing the discrete phase consisting of a kernel, concluded in a capsule from a membrane-forming substance; and the continuous phase consisting of water medium, thus the discrete phase is homogeneously distributed in the continuous phase. The way of processing of a target site using the HIFU method consists in intravenous introduction of an effective dose of the specified strengthening agent by means of continuous and fast intravenous installation or bolus injection to the patient 0-168 hours prior to HIFU-therapy application. The way of an agent screening strengthening effect of HIFU processing, consists in HIFU-therapy application - to a concrete tissue, calculation of EEF(initial) energy, introduction of a strengthening agent-candidate in a biological tissue, measurement of EEF(experim.) energy, comparison of EEF(initial) and EEF(experim.), and in a choice of the strengthening agent-candidate concerning EEF(initial) to EEF(initial) it is more than 1. The given group of inventions allows reducing quantity of the acoustic energy necessary for damage of a target tissue, in unit of volume of a tissue in the course of HIFU processing, to reduce time of processing of target area and to improve therapeutic effects.

EFFECT: reduction of quantity of the acoustic energy necessary for damage of a target tissue, in unit of volume of a tissue in the course of HIFU processing, reduction of time of processing of target area and improvement of therapeutic effects.

19 cl, 24 ex, 10 tbl

FIELD: medicine.

SUBSTANCE: invention concerns medicine area, namely to treatment and cancer prevention. Application of nonradioactive strontium with an amino acid and an inorganic element for manufacturing of the pharmaceutical agent for treatment or prevention of cancer of prostate, carcinoma of kidneys or melanoma is offered. Application of nonradioactive strontium allows reducing cost of treatment and improving treatment acceptability by patients at the expense of absence of by-effects at application of the offered pharmaceutical agent. Also, application of the offered pharmaceutical agent allows reaching a nontoxic way of treatment and cancer prevention.

EFFECT: reduction of cost of treatment and improvement of treatment acceptability by patients at the expense of absence of by-effects at application of the offered pharmaceutical agent, and also achievement of a nontoxic way of treatment and cancer prevention.

8 cl, 3 ex, 4 tbl

FIELD: medicine.

SUBSTANCE: group of inventions relates to medicine, particularly to oncology and can be used for treating and preventing recurrence of malignant tumours in female reproductive system. According to the invention, the method involves introduction of 9-oxoacridine-10-acetic acid (CMA) and/or its pharmaceutically acceptable salt and/or its ester combined with aromatase inhibitor. The invention also pertains to use of CMA in making a medicinal agent and a set for use in the given methods.

EFFECT: provision for treatment and prevention of recurrence of malignant tumours in female reproductive system through potentiation of the effect of aromatase inhibitors and chemotherapeutic agents under the effect of CMA.

19 cl, 9 tbl, 15 ex

FIELD: medicine.

SUBSTANCE: invention relates to medicine, particularly to experimental oncology and prevention of carcinogenic effect of diethylnitrosamine in experimental animals. For this purpose a 100 mg/l portion of diethylnitrosamine is administered daily for 4 months together with an anticarcinogen, which is administered with food, 5 days before administering diethylnitrosamine. The anticarcinogen used is Essentiale N, in a dose of 75 mg/kg for nine months.

EFFECT: effective prevention of carcinogenesis in specific experimental conditions.

8 dwg, 2 ex, 2 tbl

FIELD: medicine.

SUBSTANCE: present invention relates to pharmaceutics and a tablet with high drug content, which contains an active component in form of a formula (I) compound or its pharmaceutically acceptable salt in amount of approximately 30-80 wt % of active component, in total mass of the tablet and at least one binding substance, which contains microcrystalline cellulose or hydroxypropylmethicellulose.

EFFECT: design of a tablet with high drug content.

16 cl, 1 dwg, 3 ex

FIELD: chemistry.

SUBSTANCE: invention relates to quinazolene derivatives of formula (I) and pharmaceutically acceptable salts of salt for slowing down cancer cells growth, as well as to method of obtaining compound and pharmaceutical composition based on them. Compounds can be applied for treatment of cancer diseases in which growth of cancer cells is induced by epithelial growth factor. In general formula (I) R1 represents hydrogen, C1-6alkyl, C3-7cycloalkyl, C2-6alkenyl, C2-6alkynyl, C1-6alkoxy, C1-6alkylamino, C1-6dialkylamino, C1-6alkyltio, 2-((Nmethyl-(2-hydroxyethyl)amino)mathyl)vinyl, phenoxy, phenyl, dimethylaminoethoxymethyl, methoxyethoxymethyl, 4-methylpiperazinylmethyl, ethoxycarbonyl, benzamino, cyclopentenyl, 2-(4-methylpiperazinylmethyl)vinyl, methanesulphonylethylaminomethyl, methoxyethylaminomethyl, 2-(methanesulphonylethylaminomethyl)vinyl or 5-member heteroaryl, containing 1-2 heteroatoms selected from N, O or S, which are optionally substituted with X; R2 represents hydrogen or C1-6alkyl; R3 represents C1-6alkyl or C1-6dialkylamino C1-6alkyl; Y represents -(CR11R12)-, phenyl or 5-member heteroaryl, containing one sulphur atom, R11 and R12, each independently represent hydrogen, halogen, hydroxy, carboxyC1-3alkyl, C1-6alkyl, C1-6alkoxy, C1-6alkyltio, C1-6alkylsulphinyl, C1-6alkylsulphonyl, C1-6alkanoyl, C1-6alkoxycarbonyl, phenyl; optionally are condensed with each other with formation of 3-member non-aromatic ring, or R11 or R12 can be condensed with R2 with formation together with carbon and nitrogen, to which they are bonded, of 4-6-member non-aromatic ring, optionally containing additional sulphur heteroatom; R4 represents hydrogen, halogen, C1-6alkoxy, ethylsulphanyl, cyclopropylmethoxy, cyclopentyloxy, or C1-6alkylamino, optionally substituted with R13, where R13 represents halogen, trifuoromethyl, C1-6alkoxy, 4-methylpiperidinylmethoxy, N,N-dimethylamino or 6-member heteroaryl, containing 1-2 heteroatoms selected from N and O; R5 represents hydrohen or C1-3alkyl; R6 represents R14, phenyl, 1-phenylethyl, 3-ethenylphenyl, 1-penta-2,4-dienyl-1N-indasol-5-yl, imidasol or 9-member heteroaryl, containing 1-2 nitrogen atoms, optionally substituted with substituent R15, where R14 and R15, each independently represent hydrogen, halogen, C1-6alkyl, or phenyl, benzyl, 10-member heteroarylC1-6alkyl, containing one nitrogen atom, benzyloxy, 6-member heteroarylC1-6alkoxy, containing one nitrogen atom, 6-member heteroaryloxy, containing one nitrogen atom, phenylcarbamoyl, optionally substituted with R16, where R16 represents halogen, nitro, C1-6alkyl, or C1-6alkoxy; n equals integer number within the range from 1 to 4; p equals 0 or 1; X represents halogen, nitro, cyano, trifluoromethyl, C1-6alkyl, C1-6alkoxy, C1-6dialkylamino, C1-6alkylsulphinyl, C1-6alkylsulphonyl, C1-6alkyloxycarbonyl, C1-6alkanoyl, phenyl or 5-6-member heteroaryl, containing 1-2 heteroatoms selected from N and O; where heteroaryl represents heteroaromatic or non-aromatic group.

EFFECT: obtaining quinazoline derivatives and pharmaceutically acceptable salts of salts for slowing down growth of cancer cells.

11 cl, 7 dwg, 5 tbl, 225 ex

FIELD: chemistry, biochemistry.

SUBSTANCE: claimed invention relates to immunology and biotechnology. Claimed are versions of neutralising antibodies or their functional analogues against human NOGO-A, each of which is characterised by set of specific CDR. Described is pharmaceutical composition for human NOGO-A neutralisation. Described are versions of application of antibodies or their functional analogues in production of medication for treatment and/or prevention of neurological diseases/ disorders. Described is method of axon growth stimulation using antibodies of invention.

EFFECT: ensuring neutralising antibodies with high affinity to human NOGO-A, with KD of 10-9 M, which can be further applied in medicine for activation of neurite growth.

17 cl, 45 dwg, 14 tbl, 18 ex

FIELD: medicine; pharmaceutics.

SUBSTANCE: invention concerns application of (2R)-2-{[1 - ({[(3S)-1 (carboxymethyl)-2-oxo-2,3,4,5-tetrahydro-1H-1-benzazepine-3-il]amino}carbonyl)cyclopentyl]methyl}-4-of phenylbutane acid or (2R)-2-{[1-({[(3S)-1-(carboxymethyl)-2-oxo-2,3,4,5-tetrahydro-1H-1-benzazepine-3-il]amino}carbonyl)cyclopentyl]methyl}-4-(1-naphthyl) of butane acid for reception of pharmaceutical compositions for prevention and treatment of craniocerebral trauma and spinal cord damage.

EFFECT: high efficiency of treatment.

2 cl, 2 tbl, 1 ex

FIELD: chemistry.

SUBSTANCE: invention relates to new compounds with general formula (I) , where R1 and R2 are independently chosen from hydrogen, halogen, nitro, alkyl, alkylaryl and XYR5; X and Y are independently chosen from O and (CR6R7)n; R3 represents hydrogen, alkyl or M; M represents an ion, chosen from aluminium, calcium, lithium, magnesium, potassium, sodium, zinc or their mixture; Z represents CR4; R4 is chosen from hydrogen, halogen, alkyl, alkylaryl and XYR5; R5 is chosen from aryl, substituted aryl, heteroaryl and substituted heteroaryl; R6 and R7 are independently chosen from hydrogen and alkyl; n is an integer from 1 to 6; at least one of R1 and R2 represents XYR5, and at least one of X and Y represents (CR6R7)n. The invention also pertains to the method of increasing concentration of D-serine and/or reducing concentration of toxic products of D-serine oxidation under the effect of DAAO in mammals, involving introduction into a subject of a therapeutically effective amount of a formula I compound, to the method of treating schizophrenia, treating or preventing loss of memory and/or cognitive ability, to the method of improving learning ability, method of treating neuropathic pain, as well as to a pharmaceutical composition, with DAAO inhibitory activity, based on these compounds.

EFFECT: obtained are new compounds and a pharmaceutical composition based on these compounds.

27 cl, 4 tbl, 72 ex

FIELD: medicine.

SUBSTANCE: invention concerns medicine and pharmacy area, in particular, to the medical products used for treatment of epilepsy and neuropathic pain. The agent contains carbamazepine and thyotriazoline in a mass parity 1: (0.25-2.5) in quality of an active base according to the invention. The agent possesses anticonvulsant, neuroprotective, nootropic, analgetic, antidepressive, actoprotective and antioxidatic action. Use of the combined agent under the invention allows to raise efficiency of treatment of the specified diseases in comparison with use of carbamazepine and thyotriazoline separately and to lower implication of toxic action of an agent at treatment.

EFFECT: rising of efficiency of treatment of the specified diseases in comparison with use of carbamazepine and thyotriazoline separately and depression of implication of toxic action of an agent at treatment.

9 tbl, 8 ex

FIELD: chemistry, medicine.

SUBSTANCE: in the general formula (I): X is oxygen atom; R1 is C1-10-alkyl , substituted if necessary by phenyl or thienyl group; or R1 is C3-7-cycloalkyl, thienyl, pyridinyl; the thienyl groups can be substituted if necessary by 1-2 C1-3-alkyl groups; phenyl can be substituted if necessary by 1-2 halogen atoms; R2 is C1-6-alkyl; or R2 is C3-7-cycloalkyl, phenyl or pyridinyl; phenyl if necessary can be substituted by one or more halogen atoms or by the CN, C1-3-alkyl, C1-3-alkoxyl, C1-3-fluoroalkyl groups; R3 is C1-6-alkyl; R4 is hydrogen atom or C1-6-alkyl; R5 and R5' are independently of each other the hydrogen atom, hydroxyl; or R5 and R5' form together the oxo-group; n is integer value in the range from 0 to 3; R6 is independently of each other hydrogen atom, halogen atom, C1-3-alkyl, C1-3-alkoxyl.

EFFECT: compounds of present invention can find application as pharmaceutical for pathology treatment where the inhibitor of β-amiloyd peptide β-A4 is useful.

8 cl, 1 tbl, 7 ex

FIELD: chemistry; medicine.

SUBSTANCE: compounds of claimed invention possess properties of positive allosteric modulator mGluR5. In general formula I , W represents 6-member heterocycloalkyl ring with 1-2 heteroatoms, selected from N, O; R1 and R2 independently represent hydrogen, C1-C6-alkyl; P and Q each independently is selected from: , R3, R4, R5, R6 and R7 independently represent hydrogen; halogen; -CN; nitro; C1-C6-alkyl; C3-C6-cycloalkyl; halogen-C1-C6-alkyl; 5-6-member heteroaryl with 1-2 atoms N as heteroatoms; 6-member heterocycle with 2 heteroatoms representing N, O; phenyl, optionally substituted with halogen; naphtyl; -OR8; where optionally two substituents together with located between them atoms form 9-10-member bicyclic aryl or heteroaryl ring with 1-2 heteroatoms, selected from N, S; R8 represents hydrogen, C1-C6-alkyl; D, E, F, G and H independently represent -C(R3)=, -O-, -N=, -N(R3)- or -S-; A represents ethinyl, -C(=O)NR8- or group of formula . B represents -C(=O)-C0-C2-alkyl-, -C(=O)-C2-C6-alkenyl-. Invention also relates to pharmaceutical composition based on invention compounds.

EFFECT: novel compounds possess useful biological proprties.

20 cl, 3 dwg, 75 ex

FIELD: chemistry, medicine.

SUBSTANCE: invention refers to the method for modulation of the CRTh2-receptor activity with usage of the compounds of formula (I) or their pharmaceutically acceptable salts where: W is O, S(O)n (where n is equal 0, 1 or 2), NR15, CR1OR2 or CR1R2; X is hydrogen, halogen or C1-6 alkyl which can be substituted with one or more halogen atom; Y is hydrogen, halogen; Z is phenyl, pyridyl, pyrimidyl or quinolyl possibly substituted with one or more substituting group independently selected from following groups: halogen, CN, nitro, SO2R9, SO2NR10R11, CONR10R11, NHSO2R9 or C1-3 alkyl substituted with one or more halogen atom; R1 and R2 are independently hydrogen atom or C1-6 alkyl; R9 is C1-6 alkyl; R10 and R11 are independently hydrogen atom or C1-6 alkyl; R15 is hydrogen atom or C1-6 alkyl.

EFFECT: improvement of the method.

19 cl, 68 ex

FIELD: medicine.

SUBSTANCE: invention refers to medicine and covers therapy of patients suffering from pancreatic diabetes combined with central nervous system damage. The therapy involves intravenous infusion of Mexidol 5% 5.0 ml. In 30-35 minutes, the patient is exposed to transcerebral sinusoidal current of frequency 5000 Hz, modulated low-frequency oscillation with using fronto-occipital technique. The half-period duration is 1:1.5 in variable mode, III operation mode, modulation depth 75%, frequency 30 Hz. Current strength within 0.8 to 2.0 mA is dosed to sensation of easy, painless vibration. Duration of procedures is 15 minutes in therapeutic course 8-10 daily procedures.

EFFECT: method allows reducing frequency and intensity of headaches, degree of manifestation of depression and anxiety, improving state of health, memory, attention, cerebral circulation and putting the patient in good spirit, normalising sleep and blood lipids.

2 ex

FIELD: chemistry.

SUBSTANCE: invention is related to new compound - 1-((3-pyridylcarbonyl)aminomethyl)-1-cyclohexane acetic acid of formula I and its pharmaceutically acceptable salts, which display nootropic, anticonvulsive, anxiolytic properties and may find application as neurotropic drug. Compound I is produced by condensation of alkyl ether of 3-pyridine-carbonic acid with salt 1-(aminomethyl)cyclohexane acetic acid, normally, in organic dissolvent at dissolvent boiling temperature with further treatment with mineral or organic acid, if required.

EFFECT: in order to produce salt of compound I, it is treated usually with oxide hydrate or oxide of alkaline or alkali-earth metal.

5 cl, 8 tbl, 5 ex

FIELD: medicine, pharmaceutics.

SUBSTANCE: present invention concerns area of medical products, in particular to application of complex bonds of iron (III) with carbohydrates or their derivatives for reception of a medical product for improvement of immune protection and-or activity of a brain at patients without an anaemia caused by deficiency of iron or deficiency of iron in which complex bond of iron (III) is iron (III)-polymaltose complex bond or complex bond of iron (III) with a product of oxidation one or several maltodextrins.

EFFECT: development of preparations for improvement of immune protection and-or activity of a brain at patients without an anaemia caused by deficiency of iron or deficiency of iron.

7 cl, 1 tbl, 1 ex

FIELD: chemistry.

SUBSTANCE: in derivatives of colchicine of general formula (I) X is oxygen or sulphur, particularly - to 3-O-β-D- xylopyronozyl-3-O-demethyltiocolchicine and 3-O-β-L- xylopyrano-zyl-3-O-demethyltiocolchicine.

EFFECT: efficient for preparation of miorelaxant medications, medications for treatment of inflammatory conditions, which influence muscular system, and antigout medications.

11 cl, 2 tbl, 6 ex

Up!