RU2610438C2 - Method of treating pathological syndrome and drug of central and peripheral action for treating pathological syndrome - Google Patents

Abstract

FIELD: medicine.

SUBSTANCE: invention relates to medicine and represents method of treating obesity, nicotine addiction and reduction of alarm, while smoking cessation by administering drug central and peripheral action in body, which is based on activated potentiated form of antibodies. Disclosed method is characterized by that drug contains activated potentiated form of antibodies to human cannabinoid receptor in form of activated potentiated aqueous or aqueous-alcoholic solution, obtained during successive multiple dilution of matrix solution in water or aqueous-alcohol solvent and intermediate external mechanical action.

EFFECT: higher efficiency of treating obesity, nicotine addiction and reduction of alarm, while smoking cessation.

17 cl, 12 tbl, 7 ex

Description

The invention relates to medicine and can be used to increase the effectiveness of the treatment of various diseases, including obesity, mainly exogenous-constitutional, including in combination with lipid metabolism disorders, including metabolic syndrome, as well as for the treatment of drug and / or psychoactive addiction substances.

According to the International Obesity Task Force, more than 300 million people on our planet are obese, and another 800 million are overweight. While maintaining such high rates of growth in the incidence rate by 2025, a twofold increase in the number of obese patients is expected.

Overweight and obesity are one of the most common problems in developed countries, and increase the risk of developing diseases of the cardiovascular system, dyslipidemia, type 2 diabetes, cancer.

Among the modern drugs used to reduce body weight, three main groups can be distinguished - peripheral and central action drugs that enhance the “burning” of calories, drugs that reduce fat intake in the body, and central action drugs (appetite regulators) (Padwal RS, Majumdar SR, Drug treatments for obesity: orlistat, sibutramine, and rimonabant. Lancet 2007; 366 (9555): 71-7). Preparations of all groups have multiple side effects, and therefore their use in everyday practice is extremely limited. Thus, the development and implementation of new safe and effective drugs to reduce body weight are very relevant.

The prior art composition for the treatment of obesity and insulin resistance in the regulation of blood glucose, containing a compound isolated from a natural source and purified, selected from the group including anthocyanin, anthocyanidin and mixtures thereof (RU 2359689 C2, A61K 36/45, 09/10/2008). However, when using this composition, it is difficult to select a dose.

The use of a drug based on antibodies to lipase for the treatment of obesity in mammals is also known (WO / 1999/002187, A61K 38/00, 01/21/1999). However, this drug can cause severe side effects and, as a result, is used in a small number of cases.

A medicament is known in the art for the oral treatment of obesity, diabetes mellitus and other diseases accompanied by impaired glucose tolerance, containing antibodies to the beta subunit of the insulin receptor in activated form (WO 2007/149010 A1, A61K 39/395, 12/27/2007) . However, this drug is effective mainly in obesity associated with insulin resistance.

The invention is aimed at creating an effective method for treating a pathological syndrome, including treating obesity and related metabolic disorders, as well as treating addiction to narcotic and / or psychoactive substances without side effects, and a corresponding central and peripheral drug based on antibodies to the known cannabinoid receptor human (Ken Mackie. Cannabinoid receptors as therapeutic targets. The Annual Review of Pharmacology and Toxicology, 2006. 46: 101-122).

The solution to this problem is provided by the fact that in the method of treating a pathological syndrome by introducing into the body a drug of central and peripheral action based on the activated-potentiated form of antibodies, according to the invention, the drug contains an activated-potentiated form of antibodies to the human cannabinoid receptor in the form of activated-potentiated aqueous or aqueous-alcoholic solution obtained in the process of sequential multiple dilution of the matrix of solution in aqueous or aqueous-alcoholic solvent, and an intermediate external mechanical action.

It is possible to use an activated-potentiated form of antibodies to a whole molecule of a human type I cannabinoid receptor or a polypeptide fragment with a sequence selected, for example, from the following group:

In addition, an activated-potentiated form of antibodies to the brain-specific protein S-100 is additionally introduced in the form of an activated-potentiated aqueous or aqueous-alcoholic solution obtained by sequential multiple dilution of the matrix solution in an aqueous or aqueous-alcoholic solvent and an intermediate external mechanical action.

Preferably, the activated-potentiated form of antibodies to the human cannabinoid receptor or the activated-potentiated form of antibodies to the human cannabinoid receptor in combination with the activated-potentiated form of antibodies to the brain-specific protein S-100 is used to treat obesity and related metabolic disorders.

In this case, the activated-potentiated form of antibodies to the human cannabinoid receptor or the activated-potentiated form of antibodies to the human cannabinoid receptor in combination with the activated-potentiated form of antibodies to the brain-specific protein S-100 is used to treat addiction to narcotic and / or psychoactive substances.

In this case, the activated-potentiated form of antibodies to the human cannabinoid receptor or the activated-potentiated form of antibodies to the human cannabinoid receptor in combination with the activated-potentiated form of antibodies to the brain-specific protein S-100 is used to treat alcohol dependence.

In this case, the activated-potentiated form of antibodies to the human cannabinoid receptor or the activated-potentiated form of antibodies to the human cannabinoid receptor in combination with the activated-potentiated form of antibodies to the brain-specific protein S-100 is used to treat smoking addiction.

Moreover, the drug can be made in solid dosage form in the form of a tablet containing an effective amount of granules of a neutral carrier saturated with an aqueous or aqueous-alcoholic solution of an activated-potentiated form of antibodies to a human type I cannabinoid receptor or an activated-potentiated form of antibodies to a human cannabinoid receptor in in combination with an activated-potentiated form of antibodies to the brain-specific protein S-100, and pharmaceutically acceptable additives.

Preferably, the drug is administered 1 ÷ 2 tablets 2 ÷ 6 times a day.

Alternatively, the activated-potentiated form of antibodies to the human cannabinoid receptor or the activated-potentiated form of antibodies to the human cannabinoid receptor in combination with the activated-potentiated form of antibodies to the brain-specific protein S-100 is used to treat mild smoking dependence according to the Fagerstrom test.

Alternatively, the activated-potentiated form of antibodies to the human cannabinoid receptor or the activated-potentiated form of antibodies to the human cannabinoid receptor in combination with the activated-potentiated form of antibodies to the brain-specific protein S-100 is used to treat severe tobacco dependence depending on the results of the Fagerstrom test.

The solution to this problem is also ensured by the fact that the drug of central and peripheral action for the treatment of pathological syndrome contains an activated-potentiated form of antibodies to the human cannabinoid receptor of type I in the form of an activated-potentized aqueous or aqueous-alcoholic solution obtained in the process of serial multiple dilutions in aqueous or a water-alcohol solvent and an intermediate external mechanical effect.

In addition, the drug may additionally contain an activated-potentiated form of antibodies to the brain-specific protein S-100 in the form of an activated-potentiated aqueous or aqueous-alcoholic solution obtained in the course of successive multiple dilutions of the matrix solution in an aqueous or aqueous-alcoholic solvent and an intermediate external mechanical exposure.

In this case, a water-alcohol solution of the activated-potentiated form of antibodies was obtained by repeated serial dilution and intermediate external exposure from the matrix solution of the corresponding affinity purified antibodies with a concentration of 0.5 ÷ 5.0 mg / ml.

The drug may be in solid dosage form and contain an effective amount of a neutral carrier granule saturated with an aqueous or aqueous-alcoholic solution of the activated-potentiated form of the corresponding antibodies, and pharmaceutically acceptable additives.

In addition, pharmaceutically acceptable additives include lactose, microcrystalline cellulose and magnesium stearate.

Preferably, a neutral carrier and pharmaceutically acceptable additives are formulated as a tablet soaked in 6 ml of an aqueous or aqueous-alcoholic solution of an activated potentiated antibody form weighing 300 mg.

The claimed technical result may be due to the fact that the known type 1 cannabinoid receptors and endocannabinoids represent a single system in the hypothalamus, which, affecting mainly specific areas of the mesolimbic region of the brain, affects the feeling of fullness and regulates appetite and, as a result, consumption food, and the activated-potentiated form of antibodies to the human cannabinoid receptor, including in combination with the activated-potentiated form of antibodies to the brain-specific Ceska protein S-100, provides both the normalization of carbohydrate and lipid metabolism, as well as a significant reduction of dependence on drugs and / or psychoactive substances.

Moreover, the proposed drug based on the activated-potentiated form of antibodies to the human cannabinoid receptor, including in combination with the activated-potentiated form of antibodies to the brain-specific protein S-100 (i.e., forms of antibodies to the human cannabinoid receptor type I and brain-specific protein S-100, prepared according to the homeopathic technology of potentiation by multiple sequential dilutions in combination with external exposure - vertical shaking of each dilution (see , for example, V. Schwabe "Homeopathic medicines", M., 1967, p.14-29), which are active in pharmacological models and / or clinical methods as normalization of carbohydrate and lipid metabolism, leading to weight loss without side effects, and treatment of dependence on narcotic and / or psychoactive substances, mainly alcoholism and smoking) provides an unexpected synergistic therapeutic effect, confirmed on adequate (valid) experimental models and clinics FIR studies. Moreover, the claimed drug, being a central and peripheral drug, not only normalizes carbohydrate and lipid metabolism, but also modulates the system of positive emotional support, thereby reducing the severity of craving for psychoactive substances, including alcoholism and nicotinism, normalizes the balance of neurotransmitters in central nervous system, improves neuroprotective activity, increases the resistance of the brain to toxic effects, improves integrative activity, elimination of cognitive impairment exists, enhances mental performance, normalizes somatovegetativnye manifestations of abstinence. As a result of this, irritability is eliminated, mood improves, internal discomfort is eliminated, thoughts of nicotine or alcohol are eliminated, working capacity is improved, sweating, tachycardia and general weakness are eliminated, as well as other affective, ideator and asthenovegetative disorders; quality of life improves. At the same time, the claimed neurotropic drug does not have a sedative, muscle relaxant effect, is not addictive, does not have drug dependence, does not have narcogenic potential and other side effects and has an extended therapeutic range that provides treatment for various diseases, including obesity, and dependence on narcotic and / or psychoactive substances.

In addition, the technical solution obtained in accordance with the invention extends the arsenal of drugs of central and peripheral action, intended for the effective treatment of various diseases, including as part of complex therapy.

A drug is prepared, for example, as follows.

To prepare a homeopathically activated-potentiated form of antibodies, monoclonal or, mainly, polyclonal antibodies are used, which can be obtained by known technology-methods described, for example, in the book: Immunological methods, ed. G. Fremel, M., "Medicine", 1987, S. 9-33; or, for example, in an article by Laffly E., Sodoyer R. Hum. Antibodies. Monoclonal and recombinant antibodies, 30 years after. - 2005 - Vol.14. - N 1-2. P.33-55.

Monoclonal antibodies are obtained, for example, using hybridoma technology. Moreover, the initial stage of the process includes immunization, based on the principles already developed in the preparation of polyclonal antisera. Further stages of the work include obtaining hybrid cells producing clones of antibodies of the same specificity. Their isolation in an individual form is carried out by the same methods as in the case of polyclonal antisera.

Polyclonal antibody antibodies to the human cannabinoid receptor can be obtained by active immunization of animals. To do this, according to a specially developed scheme, the animals are given a series of injections of a substance-antigen required by the invention: a human type I cannabinoid receptor. As a result of this procedure, a monospecific antiserum with a high antibody content is obtained, which is used to obtain the activated-potentiated form. If necessary, the antibodies present in the antiserum are purified, for example, by affinity chromatography, using fractionation by salt precipitation or ion exchange chromatography.

For immunization of rabbits, you can use as an immunogen (antigen) adjuvant and the whole molecule of the cannabinoid receptor type I person of the following sequence:

or, preferably, a polypeptide fragment of a human type I cannabinoid receptor with a sequence selected, for example, from the following group:

Before blood sampling for 1-3 days, 1-3 intravenous injections are performed to increase the level of antibodies. During immunization, small blood samples are taken from rabbits to evaluate the amount of antibody. The maximum level of the immune response to the administration of most soluble antigens is achieved 40-60 days after the first injection. After the end of the first cycle of immunization of rabbits for 30 days, they restore health and re-immunization, including 1-3 intravenous injections. To obtain antisera from immunized rabbits, blood is collected in a 50 ml centrifuge tube. Using a wooden spatula, the formed clots are removed from the walls of the tube and the wand is placed in the clot formed in the center of the tube. Blood is placed in a refrigerator (temperature 4 ° C) at night. The next day, the clot attached to the spatula is removed and the remaining liquid is centrifuged at 13000 g for 10 minutes. The supernatant (supernatant) is an antiserum. The resulting antiserum, which should be yellow, is stored until use in a frozen state at a temperature of -70 ° C. To isolate antibodies to the cannabinoid receptor type I from an antiserum of a person, type 1 absorption is performed on the solid phase in the following sequence:

1. 10 ml of rabbit antiserum is diluted 2 times with 0.15 M NaCl, 6.26 g of Na ₂ SO ₄ are added, stirred and incubated for 12-16 hours at 4 ° C;

2. The precipitate formed is removed by centrifugation, dissolved in 10 ml of phosphate buffer and then dialyzed against the same buffer overnight at room temperature;

3. after removal of the precipitate by centrifugation, the solution is applied to a column with DEAE-cellulose, balanced with phosphate buffer;

4. The antibody fraction is determined by measuring the optical density of the eluate at 280 nm.

The antibodies are then purified by affinity chromatography by attaching the obtained antibodies to the human type I cannabinoid receptor, which is located on an insoluble matrix, followed by elution with concentrated salt solutions.

Thus obtained buffer solution of polyclonal rabbit antibodies to the cannabinoid receptor of human type I, purified on antigen, with a concentration of 0.5 ÷ 5.0 mg / ml, preferably 2.0 ÷ 2.5 mg / ml, is used as a matrix (primary) solution for the subsequent preparation of the activated-potentiated form.

To obtain polyclonal antibodies to the brain-specific protein S-100, the brain-specific protein S-100 is used, the physicochemical properties of which are described in the article by M.V. Starostin, S. M. Sviridov, Neurospecific protein S-100, Successes in modern biology, 1977. , t.5, p. 170-178; book of MB Stark, Brain-specific proteins / antigens / and neuron functions, M., "Medicine", 1985; p.12-14, isolated from tissue of the brain of a bull by the following method:

- fabric frozen in liquid nitrogen is ground into powder in a special mill;

- the extraction of proteins is carried out in a ratio of 1: 3 (weight / volume) in the extraction buffer during homogenization;

- the homogenate is heated for 10 min at 60 ° C and cooled to 4 ° C in an ice bath;

- thermolabile proteins are removed by centrifugation;

- carry out stepwise fractionation with ammonium sulfate, followed by removal of precipitated impurity proteins;

- the S-100 protein-containing fraction is precipitated with 100% saturated ammonium sulfate when the pH is reduced to 4.0 and collected by centrifugation;

- the precipitate is dissolved in a minimum volume of a buffer containing EDTA and mercaptoethanol, dialyzed against deionized water and freeze-dried;

- fractionation of acidic proteins is continued by chromatography on ion-exchange media — DEAE cellulose DE-52 and then DEAE-Sephadex A-50;

- collected and dialyzed fractions containing S-100 protein are separated by molecular weight by gel filtration on Sephadex G-100;

- purified S-100 protein is dialyzed and freeze-dried.

The molecular weight of the purified brain-specific protein S-100 is 21,000 d.

Due to the high content of aspartic and glutamic acids, the brain-specific protein S-100 is strongly acidic and occupies the extreme anode position during electrophoresis in a discontinuous buffer system in a polyacrylamide gel, which facilitates its identification.

To obtain a brain-specific antiserum, a purified S-100 protein (antigen) is prepared in combination with methylated bovine serum albumin as a carrier with Freund's complete adjuvant, which is injected subcutaneously to a laboratory rabbit animal in the amount of 1- to a selected brain-specific protein S-100. 2 ml On the 8th, 15th day, re-immunization is carried out. Blood is taken (for example, from an ear vein) on the 26th and 28th day.

The resulting antiserum has a titer of 1: 500-1: 1000, forms a single precipitation band with extract from nervous tissue, but does not react with extracts of heterologous organs, and forms a single precipitation peak with both pure S-100 protein and nervous tissue extract, which indicates the monospecificity of the obtained antiserum.

Thus obtained buffer solution of polyclonal rabbit antibodies to brain-specific protein S-100, purified on antigen, with a concentration of 0.5 ÷ 5.0 mg / ml, preferably 2.0 ÷ 2.5 mg / ml, is used as a matrix (primary) solution for the subsequent preparation of the activated-potentiated form.

An activated-potentiated form of antibodies to the human type I cannabinoid receptor (or to the brain-specific protein S-100) in an ultra-low dose is prepared by uniformly reducing the concentration as a result of sequential dilution of 1 part of the matrix solution in 9 parts (for decimal dilution) or in 99 parts ( for a hundredth dilution C) or in 999 parts (for a thousandth dilution M) of a neutral aqueous or hydroalcoholic solvent with multiple vertical shaking-potentiation (or "dynamization") to each dilution obtained and the use of separate containers for each subsequent dilution to obtain the required dilution potency-dilution according to the homeopathic method (see, for example, V. Schwabe "Homeopathic Medicines", M., 1967, pp. 14-29).

For example, to prepare the 12th hundredth C12 dilution, one part of the above matrix solution of human type I cannabinoid receptor antibodies of 2.5 mg / ml is diluted in 99 parts of a neutral aqueous or aqueous-alcoholic solvent and many times (10 times or more) vertically shake-potentiate the obtained 1st hundredth C1 dilution. From the 1st hundredth C1 dilution prepare the 2nd hundredth dilution C2. This operation is repeated 11 times, obtaining the 12th hundredth dilution of C12. Thus, the 12th hundredth dilution of C12 is a solution obtained by diluting successively in different containers 12 times of the first part of the initial matrix solution of antibodies to the cannabinoid receptor of human type I with a concentration of 2.5 mg / ml in 99 parts of a neutral solvent , i.e. the solution obtained by super-dilution of the initial matrix solution in 100 to ¹² times. Similar operations with the appropriate dilution ratio are carried out to obtain a dilution of C30 and C200.

When using as a biologically active liquid component a mixture of various homeopathic, mainly hundred, dilutions of the active substance, each component of the composition (for example, C12, C30, C200) is prepared separately according to the technology described above to the penultimate dilution (respectively, to obtain C11, C29, C199) and then, in accordance with the composition of the mixture, one part of each component is added to one container and mixed with the required amount of solvent (respectively, with 97 parts for hundred-percent dilution). In this case, an activated-potentiated form of antibodies to the human type I cannabinoid receptor is obtained in an ultra-low dose obtained by super-diluting the initial matrix solution 100 ¹² , 100 ³⁰ , 100 ²⁰⁰ times, which is equivalent to a mixture of hundred-percent homeopathic dilutions C12, C30, C200.

You can use a mixture of other different homeopathic dilutions, for example, decimal and / or hundredths (D20, C30, C100 or C12, C30, C50, etc.), the effectiveness of which is determined experimentally.

When potentiating instead of shaking (vertical or horizontal) in the process of decreasing the concentration, it is also possible to carry out external action with ultrasound, electromagnetic or other physical action.

The claimed drug can be prepared in the form of a pharmaceutical composition, while aqueous or aqueous-alcoholic solutions of the active components (the activated-potentiated form of antibodies to the human cannabinoid receptor type I and the brain-specific protein S-100) are mixed, mainly, in a volume ratio of 1: 1 and used in liquid dosage form.

The claimed drug can be used in solid dosage form, which contains an effective amount of granules of a neutral carrier - lactose, saturated by impregnation of the activated-potentiated form of antibodies to the human cannabinoid receptor or the activated-potentiated form of antibodies to the cannabinoid receptor with saturated aqueous or alcoholic solution human in combination with an activated-potentiated form of antibodies to the brain-specific protein S-100, and pharmaceutically when acceptable additives, including mainly lactose, microcrystalline cellulose and magnesium stearate.

A solid dosage form is prepared by tabletting from a tablet mass, which contains 8 ÷ 10 masses. parts of lactose granules irrigated in a fluidized bed with an aqueous or aqueous-alcoholic solution of an activated-potentiated form of antibodies to the cannabinoid receptor of human type I, 2-7 mass. parts moistened with 70% alcohol lactose, impregnated to saturation with an aqueous or aqueous-alcoholic solution of the activated-potentiated form of antibodies to the human cannabinoid receptor type I, and pharmaceutically acceptable additives, including 75 ÷ 95 wt. parts of pure lactose, 8 ÷ 15 mass. parts of microcrystalline cellulose and 0.8 ÷ 1.2 mass. parts of magnesium stearate. The weight of the tablet may be 150 ÷ 500 mg. It is preferable to use tablets weighing 250–300 mg, which include 3.0–6.0 mg / tablet of an activated-potentiated form of water-alcohol dilution of the substance, the active substance of polyclonal rabbit antibodies to the human type I cannabinoid receptor or of the activated-potentiated form of anti-cannabinoid antibodies human receptor in combination with an activated-potentiated form of antibodies to the brain-specific protein S-100, purified on antigen, in an ultra-low dose obtained by super-dilution of the original matrix thief 100 ^12, 100 ^30, 100 ^{to 200} times equivalent mixture of centesimal homeopathic dilutions C12, C30, C200.

It is preferable for effective treatment to use the claimed drug in 1 ÷ 2 tablets (keep in the mouth until completely dissolved) 2 ÷ 6 times a day.

To conduct experimental studies, polyclonal antibodies were used, prepared by order of a specialized pharmaceutical company.

Example 1

For the treatment of obesity, attempts have been repeatedly made to use psychoactive substances (antidepressants, nootropics, various endorphins, enkephalins, endocannabinoids) that affect the system of positive emotional support and, above all, the functional activity of the “pleasure centers” in the lateral hypothalamus. However, due to the expressed side effects, the development of addiction (addiction), an effective and at the same time safe central-acting drug has not yet been created.

The claimed drug in the direction of action - type 1 cannabinoid receptor - is central, but due to the use of special potentiation technology, it affects the receptor, leading to its sensitization and, as a result, to increased sensitivity to endogenous cannabinoids.

This effect is confirmed by the following experiment: a study was made of the effect of the drug on the basis of an activated-potentiated form of polyclonal rabbit antibodies to the human type I cannabinoid receptor, purified on antigen, in an ultra-low dose obtained by super-diluting the initial matrix solution 100 ¹² , 100 ³⁰ , 100 ²⁰⁰ times, equivalent mixture of hundreds of homeopathic dilutions of C12, C30, C200 (ultra-low doses - SMD anti-KR) to the functional state of type 1 cannabinoid receptor (in vitro) in 2 modes: in agonist mode and in re IME antagonist.

Agonist Mode:

Prior to insertion into the wells of a microplate (96-well plate, 250 μl well volume), the cells were suspended in HBSS buffer (Invitrogen) containing 20 mM HEPES (pH = 7.4). Cells were incubated for 10 minutes at room temperature with the addition of 20 μl of anti-Raman SMD. After pre-incubation, NKH477 adenylate cyclase activator was added to the wells. Cells were incubated for 10 minutes at 37 ° C and lysed. A fluorescent acceptor (cAMP labeled with D2) and a fluorescent donor (antibodies to cAMP labeled with europium cryptate) were added to the wells.

As a basal control, instead of the SMD anti-KR preparation, the cell suspension was incubated in the presence of HBSS buffer (20 μl). As a stimulated control, instead of SMD anti-CR, the cell suspension was preincubated in the presence of the reference agonist CP 55940 (20 μl).

Functional activity (by cAMP concentration) was evaluated by time-resolved homogeneous fluorescence (HTRF). The fluorescence intensity in the basal control is taken as the background and its value is subtracted from the fluorescence intensities in the experiment (SMD anti-Raman) and control (CP 55940):

The measured specific response of the cell to the introduction of SMD anti-Raman is calculated by the formula: the fluorescence intensity in the experiment (SMD anti-Raman) is the fluorescence intensity in the basal control.

The measured specific response of the cell to the introduction of a reference agonist (CP 55940) is calculated by the formula: fluorescence intensity in the control (CP 55940) - fluorescence intensity in the basal control.

The results are expressed as a percentage of the specific response of the cell to the introduction of a reference agonist in stimulated control:

% response of the reference agonist = ((measured specific response / specific response in the control on the introduction of the reference agonist) × 100%).

Antagonist Mode:

Prior to insertion into the wells of a microplate (96-well plate, 250 μl well volume), the cells were suspended in HBSS buffer (Invitrogen) containing 20 mM HEPES (pH = 7.4). Cells were incubated for 5 minutes at room temperature with the addition of 20 μl of anti-Raman SMD. After adding the reference agonist of CP 55940 to the wells, the cells were incubated for 10 minutes at room temperature. Next, NKH477 adenylate cyclase activator was added to the wells. Cells were incubated for 10 minutes at 37 ° C and lysed. A fluorescent acceptor (cAMP labeled with D2) and a fluorescent donor (antibodies to cAMP labeled with europium cryptate) were added to the wells.

As a basal control, instead of SMD anti-Raman cell suspension, the cell suspension was preincubated in the presence of the reference antagonist AM 281 (20 μl). The reference agonist of CP 55940 was not added to the wells with the basal control. As a stimulated control, the cell suspension was preincubated in the presence of HBSS buffer containing 20 mM HEPES (pH = 7.4) and the reference agonist of CP 55940 (20 μl).

Functional activity (by cAMP concentration) was evaluated by time-resolved homogeneous fluorescence (HTRF). The fluorescence intensity in the basal control is taken as the background and its value is subtracted from the fluorescence intensities in the experiment (SMD anti-Raman) and control (CP 55940):

The measured specific response of the cell to the introduction of SMD anti-Raman is calculated by the formula: the fluorescence intensity in the experiment (SMD anti-Raman) is the fluorescence intensity in the basal control.

The measured specific response of the cell to the introduction of a reference agonist (CP 55940) is calculated by the formula: fluorescence intensity in the control (CP 55940) - fluorescence intensity in the basal control.

The results are expressed as percent inhibition of the specific response of the cell to the introduction of a reference agonist (CP 55940) in the control:

% inhibition of the response of the reference agonist = 100% - ((measured specific response / specific response in the control on the introduction of the reference agonist) × 100%)

As a result of studies, it was shown (see Table 1) that SMD anti-KR alters the functional activity of type 1 cannabinoid receptor, estimated by the concentration of intracellular cAMP.

The sample (ultra-low doses of antibodies to the cannabinoid receptor type I (a mixture of homeopathic dilutions C12 + C30 + C200)) exhibits the properties of an agonist of the cannabinoid receptor type I, the severity of the modifying effect of which is 21% of the effect of the standard agonist CP 55940 (the effect of the standard agonist is taken as 100% )

Table 1. Receptor A drug The content of SMD anti-Raman in the well (% by volume) (total volume of all compounds in the well is 200 μl) Agonist mode Agonist mode Explanation of the results % response to SMD anti-KR Standard agonist % inhibition of agonist response Standard antagonist CB1 Type 1 Cannabinoid Receptor SMD anti-KR 10% 21 CP 55940 0 AM 281 SMD anti-KR have an agoniet effect, the severity of which is 21% of the effect of a standard agonist (taken as 100%)

Example 2

To study the properties of the claimed drug was used an aqueous solution of an activated-potentiated form of polyclonal rabbit antibodies to cannabinoid receptors of type I humans, purified on antigen, in an ultra-low dose obtained by super-dilution of the initial matrix solution in 100 ¹² , 100 ³⁰ , 100 ²⁰⁰ times equivalent to a mixture of hundred homeopathic dilutions of C12, C30, C200 (ultra-low doses - SMD anti-KR).

The study used 40 male mice of the C57B1 line (weight at the beginning of the study 13.5-15.5 g). 10 mice received the usual standard food (standard diet), 30 mice received the standard food with high-calorie supplements (high-calorie diet) and at the same time either distilled water (control, 0.4 ml / kg) or sibutramine ¹ ( ¹ Meridia 10 mg capsule, Abbott GMBH, Germany) (10 mg / kg), or SMD anti-KR (0.4 ml / kg). All drugs were administered intragastrically 1 time per day for 5 months. Before the introduction of drugs (initially), as well as every week after the start of their administration, the consumption of feed by mice was measured. Feed intake was estimated as the average amount of food (g) consumed by the mouse for 1 and 2 months of the study, and as the average amount of feed per 10 g of mouse body weight.

Mice in the low-calorie diet group consumed, on average, 15% less feed over the entire observation period than mice on a high-calorie diet (Table 2). Both sibutramine and SMD anti-KR reduced feed intake, and the effect of sibutramine was slightly higher: the drug reduced feed consumption per week by 19.3% (p <0.05), while SMD anti-KR - only by 9 , 3% relative to control (p> 0.05). Table 2 presents the results of the influence of SMD of anti-KR and sibutramine on food intake by C57B1 mice (average values over 5 months of observation), M ± m.

Table 2. A drug Feed intake (g / mouse) per day Feed intake (g per 10 g body weight) per day Standard diet 2.95 ± 1.42 1.35 ± 0.05 High calorie diet + distilled water 3.41 ± 1.64 # 1.54 ± 0.05 # High calorie diet + sibutramine 2.75 ± 1.36 ** 1.28 ± 0.05 ** High calorie diet + diet 3.10 ± 2.02 1.44 ± 0.08 Note: ** - differences from the control are significant, p <0.01;
# - differences from the standard diet are significant, p <0.05

However, when analyzing the dynamics of feed intake, the following features were identified. At the twentieth week of the experiment, mice on a high-fat diet treated with anti-Raman SMD consumed more food than the first week of the experiment, as shown in Table 3.

Table 3 The effect of SMD anti-Raman and sibutramine on body weight gain of C57B1 mice and food consumption per 10 g of weight over the last 20 weeks of the experiment.

Note: ** - differences from the control are significant, p <0.01; *** - p <0.001

At the same time, in mice treated with anti-Raman SMD, a decrease in body weight gain of 51.2% was still observed relative to the control. Sibutramine in the last twentieth week of the experiment also reduced body weight gain by 51.5% compared with the control group.

Example 3

To study the properties of the claimed medicinal product, an aqueous solution of an activated-potentiated form of polyclonal rabbit antibodies to human cannabinoid receptor type I, purified on antigen, was used in an ultra-low dose, an ultra-low dose obtained by super-diluting the initial matrix solution 100 ¹² , 100 ³⁰ , 100 ²⁰⁰ times , the equivalent mixture of hundreds of homeopathic dilutions of C12, C30, C200 (ultra-low doses - SMD anti-KR).

The study used 33 male mice of the C57B1 line (weight at the beginning of the study 13.09 ± 0.738 g). The mice received a modified diet with a high (45%) fat content and at the same time either distilled water (control, 0.2 ml / kg), or sibutramine (10 mg / kg), or antibodies to the human type I cannabinoid receptor, purified on antigen, a mixture of hundreds of homeopathic dilutions of C12 + C30 + C200 (ultra-low doses - SMD anti-KR) (0.2 ml / kg). All SMD anti-KR drugs were administered intragastrically 1 time per day for 2 months. Prior to the introduction of the preparations (initially), as well as every week after the beginning of their administration, the weight of mice was measured on an Philips Cucina HR 239016 electronic balance (Hungary). The increase in body weight of mice was evaluated in% of the original.

Starting from the 6th week of administration, SMD anti-KR reduced the weight gain of mice on a high-fat diet. Table 3 presents the average weekly weight (g) of C57B16 mice fed a high-fat diet and SMD anti-KR (0.2 ml / mouse) or sibutramine 10 mg / kg) (M ± m). Table 4 shows the gain in body weight of mice.

Table 4. Group Body weight (g), weeks of study 0 one 2 3 four 5 6 7 8 Distil. water 12.18 ± 0.501 15.27 ± 0.982 15.27 ± 0.488 16.00 ± 0.972 18.18 ± 0.569 17.27 ± 0.557 20.18 ± 0.501 20.55 ± 0.608 21.45 ± 0.857 Δ to the original 25,4 25,4 31.3 49.3 41.8 65.7 68.7 76.1 Sibutramine 13.82 ± 1.094 13.45 ± 0.474 16.73 ± 0.407 * 19.82 ± 0.501 ** 20.18 ± 0.784 20.91 ± 0.563 ** 22.36 ± 0.927 23.64 ± 1.343 22.91 ± 1.091 Δ to the original -2.6 21.1 43,4 46.1 51.3 61.8 71.1 65.8 SMD anti-KR 13.27 ± 0.619 16.18 ± 0.182 17.45 ± 0.282 ** 19.64 ± 0.453 ** 20.18 ± 0.423 * 20.18 ± 0.423 ** 19.82 ± 0.501 21.64 ± 0.364 22.00 ± 0.467 Δ to the original 21.9 31,4 47.9 52.1 52.1 49.43 63.0 65.8 Note: * - p <0.05 with respect to the control group;
** - p <0.001 in relation to the control group;

Thus, the SMD anti-KR drug reduces the body weight gain of mice on a high-fat diet, reducing their food intake and is not inferior in effectiveness to the well-known drug sibutramine widely used to reduce body weight.

Example 4

To study the properties of the claimed drug, 300 mg tablets were used, impregnated with an aqueous-alcoholic solution (6 mg / tablet) of an activated-potentiated form of polyclonal rabbit antibodies to the human cannabinoid receptor type I, purified on antigen, in an ultra-low dose obtained by super-dilution of the original matrix solution in 100 ¹² , 100 ³⁰ , 100 ²⁰⁰ times, equivalent to a mixture of hundreds of homeopathic dilutions of C12, C30, C200 (ultra-low doses - SMD anti-KR).

The study involved 80 volunteers (20 men and 60 women) aged 20 to 69 years (mean age 40.2 ± 1.26 years), 68.7% of whom were overweight or obese (I-III degrees), which were taken on the 1st tablet 2 times a day.

Table 5 presents the demographic and anthropometric indicators of the patients included in the study. The safety analysis included data from all patients participating in the study (n = 80). During the entire period of observation of patients, good tolerability of the drug was noted. Adverse events were absent. All patients of the study groups completed the treatment within the time period established by the study protocol; there were no prematurely retired patients. When assessing the effect of SMD anti-KR on the change in body weight of the subjects, it was found that the use of the drug led to a decrease in body weight of 56 (70%) patients. In 24 (30%) patients, the weight remained unchanged, or increased slightly, however, it should be noted that among these patients, 14 (17.5%) initially had normal body weight (BMI <25 kg / m ² ).

Table 5. Indicator Value Age years M ± m 40.2 ± 1.26 Height, cm M ± m 167.1 ± 0.89 Weight, kg M ± m 80.3 ± 1.85 Floor men 20 people (25%) women 60 people (75%) BMI, kg / m ² M ± m 28.7 ± 0.6 BMI, kg / m ² less than 25 25 people (31.3%) Normal body weight 25-29.99 28 people (35%) Overweight (obesity) 30-34.99 20 people (25%) Obesity I degree 35-39,99 5 people (6.2%) Obesity II degree 40 and more 2 people (2.5%) III degree obesity

In the dynamics of observation in patients responding to therapy, there was a statistically significant decrease in body weight (p <0.001), which after 15 days of receiving SMD anti-KR was 1.1 kg (1.3%), and after 1 month reached 1 , 9 kg (2.2%) of the initial value, as shown in Table 6.

Table 6.

Note: *** - P from ref <0.001

Also, in this group of patients, 1 week after the start of SMD anti-KR, there was a statistically significant (p <0.001) decrease in the waist circumference and hip circumference, which reached 2.3% and 2.7% by the end of therapy, respectively. Table 7 presents the dynamics of changes in the waist circumference and hip circumference.

Table 7. Day 1 (ref) Day 7 Day 14 Day 15 Day 16 Day 22 Day 29 Day 30 waist circumference, cm M ± m, cm 96.3 ± 1.97 95.2 ± 2.07 ** 93.7 ± 1.82 *** 94.2 ± 1.96 *** 95.1 ± 2.16 *** 94.5 ± 2.16 *** 94.3 ± 2.10 *** 94.1 ± 2.07 *** And from ref,% -1.1% -2.7% -2.2% -1.2% -1.9% -2.1% -2.3% thigh circumference, cm M ± m, cm 110.8 ± 1.52 110.8 ± 1.66 *** 109.3 ± 1.37 ** 109.9 ± 1.50 *** 108.7 ± 1.60 *** 108.3 ± 1.56 *** 108.1 ± 1.58 *** 107.8 ± 1.69 *** Δ from ref,% 0,0% -1.4% -0.8% -1.9% -2.3% -2.4% -2.7% Note: ** - P from ref <0.01; *** - P from ref <0.001

When assessing the severity of hunger in patients on the Visual Analogue Scale (VAS), it was initially shown that the highest intensity of hunger was observed in patients in the evening. At the end of 1 month of taking SMD anti-KR, the level of hunger in the evening hours decreased significantly (p <0.001) from 49.4 ± 3.75 to 42.0 ± 4.32 points. In the morning and afternoon, there was also a tendency to a decrease in the severity of hunger (from 20.5 ± 3.23 to 13.6 ± 1.78 points in the morning, from 44.7 ± 3.45 to 27.3 ± 3, 72 points in the daytime), which, however, did not reach statistically significant values at the end of therapy, which is possibly due to an insufficient number of patients and moderate baseline values.

Thus, in a clinical study of SMD anti-KR, the drug was highly tolerated, there were no adverse effects when taking the drug.

Example 5

In three experimental studies, the effects of antibodies to type 1 cannabioid receptor affinity purified on antigen were studied in an ultra-low dose obtained by super-dilution of the initial matrix solution with a concentration of 2.5 mg / ml 100 ¹² , 100 ³⁰ , 100 ²⁰⁰ times (AT KP) in comparison with antibodies to protein S100 affinity purified on antigen in an ultra-low dose obtained by super-dilution of the initial matrix solution 100 ¹² , 100 ³⁰ , 100 ²⁰⁰ times (AT S100), and their combination (AT S100 + AT KP).

Effect on nicotine-induced increase in motor activity of mice.

The study used 40 white non-inbred male mice (22-25 g, 1.5 months). 10 mice were intact. The rest was administered nicotine subcutaneously at a dose of 0.3 mg / kg for 4 days, as well as distilled water (control, 0.4 ml / mouse), AT S100 (0.4 ml / mouse), AT KP (0.4 ml / mouse) and AT S100 + AT KP (0.4 ml / mouse) intragastrically 1 hour before nicotine administration. 30 minutes after the last administration of nicotine, an open field test was performed. Animals are placed in the center of the Tru Scan photosensor installation (Coulboum, USA), where vertical and horizontal activity indicators are automatically recorded for 2 minutes. This model allows us to evaluate the effect of the studied compounds on the locomotor activity of animals (Howard et al., 1997). Installation parameters: size 270 × 270 × 360 mm, divided into 64 squares 2.5 × 2.5, with 25 mm holes located in the platform floor, 150-watt lamp illumination.

Nicotine is an alkaloid found in plants of the nightshade family, mainly in tobacco, and with pronounced psychotropic activity. The effect of nicotine is mediated through peripheral and central n-cholinergic receptors. The effect of nicotine on human and animal behavior is very diverse. Depending on the dose, the introduction of this alkaloid into the body can cause the development of anxiety-depressive symptoms, euphoria, agitation, or vice versa. In addition, with prolonged use of nicotine addiction is formed. The drugs used to facilitate smoking cessation, including those aimed at eliminating pathological changes in the psycho-emotional sphere, which facilitates the disposal of patients from pathological addiction to tobacco.

In the present study, the introduction of nicotine led to an increase in the motor activity of mice: the activity time increased by 8.2% (p <0.05), the distance traveled - by 5.1%, the number of holes examined - by 78.2% (p <0 , 05), the time of the research reaction - by 76.9%, and the time of immobility, on the contrary, decreased by 13.5% (p <0.05) compared with intact animals.

AT S100 did not significantly affect the studied parameters compared with the control group. AT KP reduced the time of activity and the distance traveled to the level of intact mice, but did not affect the immobility time, the number of holes studied and the time of the research reaction. At the same time, the combined administration of AT S100 and AT KP reduced the time of activity and, accordingly, increased the immobility time to the level of intact animals, significantly reduced the distance traveled (by 29.2% compared with the control and 25.5% compared with intact ) and slightly reduced the research activity (the number of openings decreased by 15.3%, the time of the research reaction decreased by 17.4%).

Thus, the introduction of AT KP, as well as the combined use of AT S100 + AT KP, helps to eliminate changes in the behavior of animals caused by the introduction of nicotine. The use of the AT S100 + AT KP complex was more effective than the introduction of AT KP. The introduction of AT S100 in this model did not affect the behavior of animals.

Table 8 shows the effect of drugs on the motor activity of mice.

Table 8. The effect of drugs on the locomotor activity of mice (open field test), M ± m Activity Time, sec Distance of movement, cm Immobility time, sec Number of researches holes Time researched. reactions Intact 74.1 ± 1.9 393.6 ± 19.6 46.0 ± 2.0 5.5 ± 1.0 2.6 ± 0.6 The control 80.2 ± 1.3 * 413.7 ± 15.4 39.8 ± 1.3 * 9.8 ± 1.4 * 4.6 ± 0.9 # AT S100 76.5 ± 1.4 434.4 ± 23.5 43.5 ± 1.4 10.0 ± 1.1 ** 5.1 ± 1.0 # AT KR 72.4 ± 1.4 ## 366.3 ± 19.1 47.6 ± 1.4 ## 9.8 ± 0.8 5.6 ± 1.1 * # AT S100 + ATKR 74.5 ± 2.8 393.1 ± 28 45.5 ± 2.9 8.3 ± 1.5 3.8 ± 0.7 significant differences from intact: * - p <0.05
differences from control are significant: # - p <0.05;## - p <0.01

Example 6

To study the properties of the claimed drug for the treatment of patients of the active drug group, 300 mg tablets were used, impregnated with a pharmaceutical composition containing aqueous-alcoholic solutions (6 mg / tablet) of activated-potentiated forms of polyclonal rabbit antibodies to type I cannabinoid receptor (anti- CB1) and brain-specific proteins in S100 ultralow doses (SMD anti-S100), the received sverhrazvedeniem initial matrix solution at a concentration of 2.5 ml / 100 ml in ^12, 100 ^30, 100 ^{to 200} times equivalent mixture of centesimal homeopathic dilutions C12, C30, C200. To treat patients of the comparison group, 300 mg tablets were used, impregnated with an aqueous-alcoholic solution (6 mg / tablet) of an activated-potentiated form of polyclonal rabbit antibodies to human cannabinoid receptor type I, purified on antigen, in an ultra-low dose (SMD anti-CB1 ) obtained by over-dilution of the initial matrix solution 100 ¹² , 100 ³⁰ , 100 ²⁰⁰ times, equivalent to a mixture of hundred-percent homeopathic dilutions of C12, C30, C200 (reference drug).

A comparative double-blind, placebo-controlled study evaluating the efficacy of the anti-S100 and anti-CB1 SMD composition in the treatment of nicotine addiction included 59 patients with a desire to quit smoking, including 22 patients in the active drug group (anti-S100 + anti-CB1 SMD 1 tablet 3 times a day), 17 patients in the comparison drug group (SMD anti-CB1, 1 tablet 3 times a day) and 20 people in the placebo group (1 tablet containing granules of a neutral carrier (lactose), not saturated with water or water-alcohol solution ktivirovannoy-potentiated forms of antibodies, and a pharmaceutically acceptable additive, 300 mg 3 times a day). The duration of therapy was 12 weeks. All three groups of patients were comparable in terms of baseline demographic, anthropometric, and clinical laboratory parameters. All patients had mild nicotine dependence according to the results of the Fagerstrom test (less than 4 points), smoking experience of at least one year and 1 degree obesity (body mass index [BMI] = 30.0-34.9 kg / m ² ). All patients of the study groups completed the treatment within the time period established by the study protocol; there were no prematurely retired patients.

When analyzing the effectiveness of therapy, it was found that the number of patients who quit smoking increased in the dynamics of observation among patients receiving anti-S100 + anti-CB1 and anti-CB1 (Table 9).

Table 9 The dynamics of the main indicators, depending on the type of therapy Period Anti-S100 + anti-CB1 (M + SE) Anti-CB1 (M ± SE) Placebo (M ± SE) Fagerstrom test, points (n - number of smokers) Originally 2.64 ± 0.10 (n = 22) 2.65 ± 0.12 (n = 17) 2.65 ± 0.11 (n = 20) Original *** 2.67 ± 0.14 (n = 12) 2.58 ± 0.14 (n = 12) 2.66 ± 0.11 (n = 18) 12 weeks 1.33 ± 0.14 * # (n = 12) 1.25 ± 0.13 * (n = 12) 2.39 ± 0.26 (n = 18) The proportion of patients who quit smoking,% 4 weeks 23% (n = 5) 12% (n = 2) 5% (n = 1) 8 weeks 36% (n = 8) 24% (n = 4) 5% (n = 1) 12 weeks 41% (n = 10) 29% (n = 5) 10% (n = 2) MPSS withdrawal symptoms scale, points (n is the number of people who quit smoking) 4 weeks 33.4 ± 1.25 (n = 5) 33.00 ± 2.00 (n = 2) 36 (n = 1) 8 weeks 28.12 ± 1.02 (n = 8) 25.50 ± 2.22 (n = 4) 34 (n = 1) 12 weeks 14.85 ± 2.41 * ** (n = 10) 20.80 ± 1.62 * (n = 5) 32.50 ± 0.50 (n = 2) Hospital scale of anxiety and depression HADS, scores (n - number of patients examined) Originally 11.73 ± 0.36 (n = 22) 11.41 ± 0.50 (n = 17) 11.4 ± 0.44 (n = 20) 4 weeks 10.32 ± 0.32 (n = 22) 10.47 ± 0.30 (n = 17) 11.95 ± 0.45 (n = 20) 8 weeks 8.59 ± 0.33 (n = 22) 9.88 ± 0.32 (n = 17) 12.55 ± 0.35 (n = 20) 12 weeks 7.68 ± 0.46 * # (n = 22) 9.41 ± 0.35 (n = 17) 12.05 ± 0.18 (n = 20) Note: * significance of differences with the placebo group, p <0.05;
** significance of differences between the Anti-S100 + anti-CB1 and anti-CB1 groups, p <0.05;# significance of differences with baseline
*** a subgroup of patients who failed to quit smoking.

In the active drug group, the proportion of quitters after 4 weeks of treatment was 23%, after 8 weeks - 36%, and by the end of 12 weeks it reached 41%; in the comparison drug group, the corresponding indicators were 12%, 24% and 29% (versus 5%, 5% and 10%, respectively, in the placebo group). The difference in treatment efficacy in the main indicator compared with placebo therapy was 31% (for the anti-S100 + anti-CB1 group) and 19% (for the anti-CB1 group) and was significant. The active drug in effectiveness significantly exceeded not only placebo, but the comparison drug.

Assessment of the effect of the composition of SMD anti-S100 and anti-CB1 on the severity of nicotine dependence showed a significant decrease both in the group as a whole and in the subgroup of patients who were not able to quit smoking. If the initial average score for the Fagerstrom test was 2.67 ± 0.14, then after 12 weeks of treatment it decreased to 1.33 ± 0.14, and the decrease was significant not only when compared with the baseline, but also with placebo group values. Patients who received a monocomponent anti-CB1 comparison drug and did not quit smoking also showed positive dynamics in terms of the severity of their nicotine addiction, which was significantly lower by the end of 3 months of treatment compared with the initial and with the placebo-therapy results.

The severity of withdrawal symptoms according to the Mood and Physical Symptoms Scale (MPSS) data among smoking cessation patients gradually decreased during the course of observation, reaching minimum values by the 12th week of treatment with both two- and one-component drugs (Table 9). The lowest scores were recorded in patients of the active drug group, indicating that the SMD anti-S100 and anti-CB1 compositions made it possible to more easily and painlessly tolerate smoking cessation, including when compared with a monocomponent drug.

Given the possible role of activation of the endocannabinoid system in the development of obesity, an analysis was made of the effect of the anti-S100 + anti-CB1 and anti-CB1 compositions on body weight during drug administration. The initial BMI calculation allowed the inclusion of patients with grade 1 obesity in all the studied groups. Table 10 presents the results of monitoring the body weight of patients of the three groups, which indicate a significant decrease in the groups of the active drug and the comparison drug, starting from the 8th week of therapy.

Table 10 Average differences between the initial and current weight (in kg) at each visit Period Anti-S100 + anti-CB1 (n = 22; M ± SE) Anti-CB1 (n = 17; M ± SE) Placebo (n = 20; M ± SE) 0 0 ± 0,0 0 ± 0,0 0 ± 0,0 2 weeks -1.2 ± 0.5 -0.9 ± 0.3 -0.4 ± 0.2 4 weeks -1.7 ± 0.4 -1.5 ± 0.5 -0.5 ± 0.4 6 weeks -2.5 ± 0.4 -2.2 ± 0.6 -1.3 ± 0.4 8 weeks -3.1 ± 0.7 * -2.8 ± 0.7 * -1.3 ± 0.4 10 weeks -3.8 ± 0.6 * -3.4 ± 0.8 * -1.8 ± 0.5 * 12 weeks -4.2 ± 0.9 * -3.8 ± 1.0 * -1.8 ± 0.6 * Note: To calculate the assessment of the reliability of the reduction indicators, the Student's bilateral criterion in the Dunnett modification was used to perform weight comparisons in the control visit (visit 0) with all subsequent visits. Significant differences (p <0.05) are marked with asterisks.

As a result of a 12-week course of treatment, body weight in both groups significantly decreased compared with the placebo group. In general, over 3 months of therapy, about half of the patients (52% and 47%) who received treatment reduced their weight by 5% or more compared to the initial state (versus 15% of patients in the placebo group; p <0.05) (Table eleven).

Table 11. The proportion of patients with weight loss of 5 percent or more

Note: * Comparison of the proportions of patients in the three groups was carried out using Fisher's one-sided exact criterion for each of the visits.

The safety assessment of therapy, based on the registration of adverse events during treatment and re-examination of laboratory parameters, indicated good tolerance to the drugs. The safety analysis included data from all patients participating in the study (n = 59). There was no negative effect of treatment on the central nervous system, any mental disorders were absent. This was confirmed by monitoring indicators using the Hospital Anxiety Scale (see Appendix 2) and Depression (HADS) (see Appendix 3) (Table 9). Moreover, the active drug showed its positive effect, exerting a leveling effect on the severity of symptoms of anxiety and depression, which significantly decreased by the end of therapy when comparing the values with baseline and placebo results. Adverse events associated with taking the drug from other organs and systems were absent. The study of laboratory parameters, including general and biochemical blood tests, clinical analysis of urine, did not record any significant deviations from normal values.

Thus, the study demonstrated the efficacy and safety of the SMD composition of anti-S100 and anti-CB1 in the treatment of mild nicotine dependence. The treatment effect consisted in a high percentage of patients who quit smoking, leveling the symptoms of withdrawal symptoms during therapy, and a decrease in the severity of nicotine addiction in those patients who could not quit smoking. All the noted effects had significant differences compared with the placebo group and, in addition, exceeded the positive results given by therapy with a monocomponent drug, which includes SMD anti-CB1. The safety profile was equally high for both drugs. Both the SMD anti-S100 + anti-CB1 composition and the monocomponent anti-CB1 SMD preparation did not lead to the appearance of clinically expressed anxiety / depression. As an additional criterion for the effectiveness of SMD anti-S100 + anti-CB1 and anti-CB1, a decrease in body weight was demonstrated in patients with grade 1 obesity.

Example 7

To study the properties of the claimed drug for the treatment of patients of the active drug group, 300 mg tablets were used, impregnated with an aqueous-alcoholic solution (6 mg / tablet) of activated-potentiated forms of polyclonal rabbit antibodies to cannabinoid type I receptor (anti-CB1) and brain-specific protein S-100 (anti-S100), in ultra-low doses (SMD), obtained by super-dilution of the initial matrix solution with a concentration of 2.5 mg / ml in 100 ¹² , 100 ³⁰ , 100 ²⁰⁰ times, equivalent to a mixture of hundreds of homeopathic dilutions C12, C30, C200. To treat patients of the comparison group, 300 mg tablets were used, impregnated with an aqueous-alcoholic solution (6 mg / tablet) of an activated-potentiated form of polyclonal rabbit antibodies to human cannabinoid receptor type I, purified on antigen, in an ultra-low dose (SMD anti-CB1 ) obtained by over-dilution of the initial matrix solution 100 ¹² , 100 ³⁰ , 100 ²⁰⁰ times, equivalent to a mixture of hundred-percent homeopathic dilutions of C12, C30, C200 (reference drug).

In order to evaluate the effectiveness of the composition of SMD anti-S100 and anti-CB1 in the treatment of severe nicotine addiction, a double-blind, placebo-controlled comparative study was conducted in 61 patients who were randomized into three groups, including 18 patients in the active drug group (SMD anti -S100 + anti-CB1, 1 tablet 4 times a day), 22 patients in the comparison drug group (SMD anti-CB1, 1 tablet 4 times a day) and 21 patients in the placebo group (1 tablet containing neutral carrier granules ( lactose) not saturated in dnym or aqueous-alcoholic solution of the activated-potentiated form of antibodies, and a pharmaceutically acceptable additive, 300 mg 4 times a day). The duration of therapy was 12 weeks. Patients of the three groups were comparable in baseline, including demographic, physical and laboratory data; according to the results of the initial assessment of the Fagerstrom test, all study participants had severe severe nicotine dependence (≥7 points) and smoking experience of more than three years. During monthly visits, patients underwent physical and laboratory examinations, testing (Fagerstrom test (see Appendix 1), Hospital anxiety scale (see Appendix 2) and depression (HADS) (see Appendix 3)), in patients who quit smoking , withdrawal symptoms were noted (Mood and Physical Symptoms Scale (MPSS) scale). All patients of the study groups completed the treatment within the time period established by the study protocol; there were no prematurely retired patients.

Table 12 shows the dynamics of the main indicators, depending on the type of therapy.

Analysis of the effectiveness of therapy showed that the number of patients who quit smoking increased in the dynamics of observation among patients receiving anti-S100 + anti-CB1 and anti-CB1 (see Table 12).

Table 12 The dynamics of the main indicators, depending on the type of therapy Period Anti-S100 + anti-CB1 (M ± SE) Anti-CB1 (M ± SE) Placebo (M ± SE) Fagerstrom test, points (n - number of smokers) Originally 8.78 ± 0.26 (n = 18) 8.55 ± 0.26 (n = 22) 8.52 ± 0.24 (n = 21) Original *** 8.92 ± 0.34 (n = 12) 8.47 ± 0.24 (n = 17) 8.47 ± 0.25 (n = 20) 12 weeks 4.50 ± 0.26 * ** # (n = 12) 6.11 ± 0.26 * # (n = 17) 8.73 ± 0.23 (n = 19) The proportion of patients who quit smoking,% 4 weeks 11% (n = 2) 9% (n = 2) 5% (n = 1) 8 weeks 22% (n = 4) 14% (n = 3) 5% (n = 1) 12 weeks 30% (n = 6) 23% (n = 5) 10% (n = 2) MPSS withdrawal symptoms scale, points (n is the number of people who quit smoking) 4 weeks 54.33 ± 1.2 (n = 2) 54.50 ± 0.50 (n = 2) 54 (n = 1) 8 weeks 46.75 ± 1.89 (n = 4) 47.33 ± 0.67 (n = 3) 50 (n = 1) 12 weeks 38.67 ± 0.88 * ** (n = 6) 45.25 ± 1.11 * (n = 5) 53.00 ± 1.00 (n = 2) Hospital scale of anxiety and depression HADS, scores (n - number of patients examined) Originally 12.61 ± 0.36 (n = 18) 12.50 ± 0.29 (n = 22) 11.81 ± 0.38 (n = 21) 4 weeks 11.17 ± 0.31 (n = 18) 11.95 ± 0.25 (n = 22) 13.05 ± 0.47 (n = 21) 8 weeks 9.56 ± 0.30 (n = 18) 10.86 ± 0.22 (n = 22) 13.24 ± 0.39 (n = 21) 12 weeks 7.72 ± 0.32 * # (n = 18) 9.50 ± 0.19 * # (n = 22) 12.67 ± 0.23 (n = 21) Note: * significance of differences with the placebo group, p <0.05;
** significance of differences between the Anti-S100 + anti-CB1 and anti-CB1 groups, p <0.05;
*** a subgroup of patients who failed to quit smoking.
* significance of differences with the initial indicator, p <0.05.

The proportion of quitters in the active drug group after 11 weeks of treatment was 11%, after 8 weeks - 22%, and by the end of 12 weeks it reached 30%; in the comparison drug group, the corresponding indicators were 9%, 14% and 23% (versus 5%, 5% and 10%, respectively, in the placebo group). According to the main indicator, the effectiveness of treatment with the anti-S100 + anti-CB1 composition exceeded the placebo therapy by 20% (p <0.05), the monocomponent anti-CB1 therapy by 13%.

The severity of nicotine dependence under the influence of the SMD composition anti-S100 and anti-CB1 significantly decreased both in the group as a whole and in the subgroup of patients who were not able to quit smoking (see Table 12). The initial average score for the Fagerstrom test was 8.92 ± 0.34, and after 12 weeks of treatment it decreased almost by half - to 4.50 ± 0.26, and the decrease was significant not only when compared with the baseline, but and with the values of the patient comparison drug groups (anti-CB1) and placebo.

The effectiveness of the anti-S100 + anti-CB1 composition was also manifested in lower, compared with anti-CB1 (p <0.05) and placebo (p <0.005), nicotine withdrawal symptoms, which, according to the MPSS scale, decreased during treatment and reached their minimum values by 12 weeks of treatment.

The safety assessment of therapy, the analysis of which included data from all patients participating in the study (n = 61), was carried out on the basis of registration of adverse events during treatment and re-examination of laboratory parameters. The results of the study testified not only to the good tolerability of both drugs, but also to the absence of any undesirable effects associated with the administration of drugs. In particular, there was no negative effect of the drugs on the central nervous system, including any mental disorders, which was confirmed by the monthly symptom score using HADS (see Table 12). On the contrary, the active drug had a leveling effect on the severity of symptoms of anxiety and depression, which significantly decreased by the end of therapy when comparing the values with baseline values and placebo group results. A study of laboratory parameters, which included general and biochemical blood tests, a clinical analysis of urine, did not register any significant deviations from normal values.

Thus, the results of the study showed the effectiveness and safety of the SMD anti-S100 + anti-CB1 composition in the treatment of severe nicotine addiction. Under the influence of a 12-week course of treatment, almost a third of smokers were able to get rid of nicotine addiction; this indicator significantly exceeded the results of using placebo. The composition of SMD anti-S100 + anti-CB1 made it possible to more easily and painlessly tolerate smoking cessation than the monocomponent anti-CB1 drug. Among those patients who were unable to quit smoking during the observation, the severity of nicotine dependence significantly decreased when using the anti-S100 + anti-CB1 composition and was significantly lower only when compared with placebo efficacy, but also when compared with the results of treatment with one component anti-CB1.

Both drugs had a high safety profile, including the absence of negative effects on the central nervous system.

Annex 1

Fagerstrom's Nicotine Test

Source: Heatherton T.F., Kozlowski L.T., Frecker R.C., Fagerström K.O. The Fagerström test for nicotine dependence: a revision of the Fagerström Tolerance Questionnaire. Br J Addict 1991; 86: 1119-27.

Description: The patient answers all questions. The final score gives an estimate of the degree of dependence on nicotine.

The degree of nicotine addiction is estimated by the sum of the points:

<4 - weak dependence

4-6 - medium dependence

7-10 - heavy addiction

1. How long after waking up do you light your first cigarette? More than 60 minutes 0 31-60 minutes one 6-30 minutes 2 less than 5 minutes 3 2. Is it difficult for you to stop smoking in places where smoking is prohibited, for example, in a meeting, on the plane, in the movies, etc.? No 0 Yes one 3. Which cigarette are you most difficult to refuse? from the first in the morning one from any other 0 4. How many cigarettes do you smoke per day? 10 or less 0 11-20 one 21-30 2 31 or more 3 5. Do you smoke more often in the first morning hours than at other times of the day? No 0 Yes one 6. Do you smoke even if you are sick and forced to lie in bed most of the day? No 0 Yes one

Appendix 2

Withdrawal Symptoms Scale (Moods and Physical Symptoms Scale, MPSS)

Source: West R, Hajek P. Evaluation of the mood and physical symptoms scale (MPSS) to assess cigarette withdrawal. Psychopharmacology, 2004; Volume 177, Numbers 1-2, 195-199

Description:

The patient who quit smoking answers 12 points (the score varies from 1 to 5 points for each point), assessing his well-being over the past 24 hours (points 1-7), craving for smoking (points 8-9) and severity physical symptoms (paragraphs 10-12). The patient circles only one number at each point. Results can be calculated in three separate domains (M - points 1-7, C - points 8-9 and P - points 10-12) or by the total amount of points, which can range from the minimum (12 points) to the maximum (60 points ), reflecting the severity of symptoms of nicotine withdrawal.

Please indicate for each item how you felt during the last 24 hours (Circle only one number for each item)

Did not have Mild Pretty strong Very much Too much 1. Depression one 2 3 four 5 2. Anxiety one 2 3 four 5 3. Irritability one 2 3 four 5 4. Anxiety one 2 3 four 5 5. Hunger one 2 3 four 5 6. Impaired attention span one 2 3 four 5 7. Sleep Disorder one 2 3 four 5

8. For how long have you wanted to smoke in the last 24 hours? (Circle only one number)

Never Very short time Less of the day Most of the day Almost all the time Constantly 0 one 2 3 four 5

9. How strong was the desire to smoke? (Circle only one number)

Did not have Lung Moderate Strong Very strong Overly strong 0 one 2 3 four 5

Have you had any manifestations in the last 24 hours? (Circle only one number)

No Lungs Moderate Heavy Very heavy 10. Pain in the mouth one 2 3 four 5 11. Constipation one 2 3 four 5 12. Cough / sore throat one 2 3 four 5

Appendix 3

Hospital Anxiety and Depression Scale (HADS)

Source: Zigmond A.S., Snaith R.P. The Hospital Anxiety and Depression scale // Acta Psychiatr. Scand. - 1983 .-- Vol.67. - P.361-370.

HADS refers to subjective scales and is intended for screening the detection of anxiety and depression in somatic patients and outpatients.

Methodology of use: the scale form is issued for self-completion by the patient and is accompanied by instructions as follows:

“Scientists are convinced that emotions play an important role in the occurrence of most diseases. If your doctor finds out more about your experiences, he will be able to better help you. This questionnaire is designed to help your doctor understand how you feel. Do not pay attention to the numbers and letters placed on the left side of the questionnaire. Read each statement carefully and in the blank column on the left mark with a cross the answer that most closely matches how you felt last week.

Do not think too much about each statement. Your first reaction will always be more true. ”

The scale is made up of 14 statements serving 2 subscales: “anxiety” (odd points - 1, 3, 5, 7, 9, 11, 13) and “depression” (even points - 2, 4, 6, 8, 10, 12 , fourteen). Each statement corresponds to 4 response options that reflect gradations of the severity of the symptom and are encoded by the increase in the severity of the symptom from 0 (absence) to 3 (maximum severity).

Assessment of the severity of the condition on the Hospital scale of anxiety and depression

When interpreting the results, the total indicator for each subscale is taken into account, while 3 areas of its values are highlighted: 0-7 - “normal” (absence of reliably expressed symptoms of anxiety and depression); 8-10 - "subclinically expressed anxiety / depression"; 11 and above - "clinically expressed anxiety / depression."

one I feel tense, I feel uneasy - all the time 3 - often 2 - from time to time, sometimes one - not at all 0 2 That which brought me great pleasure and now makes me feel the same - definitely it is 0 - probably it is one - only to a very small extent 2 - it's not like that at all 3 3 I'm in fear, it seems like something terrible is about to happen - definitely it is, and the fear is very strong 3 - yes, it is, but fear is not very strong 2 - sometimes, but it doesn’t bother me one - not at all 0 four I am able / able to laugh and see something funny in this or that event - definitely it is 0 - probably it is one - only to a very small extent 2 - not at all / capable 3 5 Restless thoughts spin in my head - constantly 3 - most part of time 2 - occasionally one - only sometimes 0 6 I am awake - not at all 3 - rarely 2 - sometimes one - almost all the time 0 7 I can easily sit and relax - definitely it is 0 - probably it is one - only occasionally 2 - I can’t do it at all 3 8 It seems to me that I began to do everything very slowly - almost all the time 3 - often 2 - sometimes one - not at all 0 9 I'm experiencing internal stress or trembling - not at all 0 - sometimes one - often 2 - Often 3 10 I do not follow my appearance - definitely it is 3 - I do not devote as much time to this as necessary 2 - maybe I began to pay less attention to this one - I follow myself as before 0 eleven I feel restless, like I’m constantly need to move - definitely it is 3 - probably it is 2 - only to some extent it is so one - not at all 0 12
I believe that my affairs (activities, hobbies) can bring me a feeling of satisfaction - exactly the same as usual 0 - yes, but not to the extent that before one - significantly less than usual 2 - I don’t think so at all 3 13
I have a sudden feeling of panic - Often 3 - often 2 - not so often one - it doesn’t happen at all 0 fourteen
I can enjoy a good book, radio or television program - often 0 - sometimes one - seldom 2 - rarely 3

Claims (17)

1. A method of treating obesity, nicotine addiction and reducing anxiety during smoking cessation by introducing a central and peripheral drug into the body based on an activated-potentiated form of antibodies, characterized in that the drug contains an activated-potentiated form of antibodies to the human cannabinoid receptor in the form activated potentiated aqueous or aqueous-alcoholic solution obtained in the process of sequential multiple dilution of matrix solution in aqueous or aqueous-alcoholic solvent, and an intermediate external mechanical action. 2. The method according to p. 1, characterized in that use the activated-potentiated form of antibodies to the whole molecule or polypeptide fragment of the cannabinoid receptor type I person. 3. The method according to p. 1, characterized in that it further introduces an activated-potentiated form of antibodies to the brain-specific protein S-100 in the form of an activated-potentiated aqueous or aqueous-alcoholic solution obtained by sequential multiple dilution of the matrix solution in aqueous or aqueous alcohol solvent and intermediate external mechanical stress. 4. The method according to p. 1 or 3, characterized in that the activated-potentiated form of antibodies is used to treat obesity and related metabolic disorders. 5. The method according to p. 1 or 3, characterized in that the activated-potentiated form of antibodies to the human cannabinoid receptor or the activated-potentiated form of antibodies to the cannabinoid receptor in combination with the activated-potentiated form of antibodies to the brain-specific protein S-100 is used to treat addiction to narcotic and / or psychoactive substances. 6. The method according to p. 5, characterized in that the activated-potentiated form of antibodies to the human cannabinoid receptor or the activated-potentiated form of antibodies to the cannabinoid receptor in combination with the activated-potentiated form of antibodies to the brain-specific protein S-100 is used to treat alcohol dependence . 7. The method according to p. 5, characterized in that the activated-potentiated form of antibodies to the human cannabinoid receptor or the activated-potentiated form of antibodies to the human cannabinoid receptor in combination with the activated-potentiated form of antibodies to the brain-specific protein S-100 is used to treat dependence on smoking 8. The method according to p. 1 or 3, characterized in that the use of a drug made in solid dosage form in the form of a tablet containing an effective amount of granules of a neutral carrier saturated with an aqueous or aqueous-alcoholic solution of an activated-potentiated form of antibodies to cannabinoid receptor I human type or an activated-potentiated form of antibodies to the human cannabinoid receptor in combination with an activated-potentiated form of antibodies to the brain-specific protein S-100, and a pharmacist cally acceptable additives. 9. The method according to p. 8, characterized in that during the treatment process, 1 ÷ 2 tablets of the drug are administered 2 ÷ 6 times a day. 10. The method according to p. 7, characterized in that the activated-potentiated form of antibodies to the human cannabinoid receptor or the activated-potentiated form of antibodies to the human cannabinoid receptor in combination with the activated-potentiated form of antibodies to the brain-specific protein S-100 is used to treat dependence on mild smoking according to the Fagerstrom test. 11. The method according to p. 7, characterized in that the activated-potentiated form of antibodies to the human cannabinoid receptor or the activated-potentiated form of antibodies to the cannabinoid receptor in combination with the activated-potentiated form of antibodies to the brain-specific protein S-100 is used to treat dependence on heavy smoking according to the results of the Fagerstrom test. 12. A central and peripheral drug for the treatment of obesity, nicotine addiction and anxiety reduction during smoking cancellation by way of claim 1, characterized in that it contains an activated-potentiated form of antibodies to the human type I cannabinoid receptor in the form of an activated-potentiated water or water -alcohol solution obtained in the process of successive multiple dilutions in an aqueous or aqueous-alcoholic solvent and an intermediate external mechanical effect. 13. The drug according to p. 12, characterized in that it further comprises an activated-potentiated form of antibodies to the brain-specific protein S-100 in the form of an activated-potentiated aqueous or aqueous-alcoholic solution obtained by sequential multiple dilution of the matrix solution in aqueous or aqueous -alcohol solvent and intermediate external mechanical stress. 14. The drug according to p. 12 or 13, characterized in that the aqueous or aqueous-alcoholic solution of the activated potentiated form of antibodies is obtained by repeated serial dilution and intermediate external exposure from the matrix solution of the corresponding affinity-purified antibodies with a concentration of 0.5 ÷ 5.0 mg / ml 15. The drug according to p. 12 or 13, characterized in that it is made in solid dosage form and contains an effective amount of granules of a neutral carrier saturated with an aqueous or aqueous-alcoholic solution of an activated-potentiated form of the corresponding antibodies, and pharmaceutically acceptable additives. 16. The drug of claim 15, wherein the pharmaceutically acceptable additives include lactose, microcrystalline cellulose and magnesium stearate. 17. The drug according to claim 16, characterized in that the neutral carrier and pharmaceutically acceptable additives are formed in the form of a tablet soaked in 6 ml of an aqueous or aqueous-alcoholic solution of an activated-potentiated antibody form, weighing 300 mg.