Screening of compounds useful in treatment and/or prophylaxis of obesity

FIELD: medicine, in particular compounds with de novo lipogenesis inhibitor activity useful in treatment and/or prophylaxis of obesity.

SUBSTANCE: claimed method includes methods (variants) for screening of compounds which are capable of inhibit at least one carbonic anhydrase activity in mammalian organism and having no anticonvulsant activity. Also is disclosed production of pharmaceuticals for treatment and/or prophylaxis of obesity containing said compounds.

EFFECT: new method for screening pharmaceuticals for treatment and/or prophylaxis of obesity on the base of their carbonic anhydrase inhibitor activity.

17 cl

 

The present invention relates to a method of identifying compounds suitable for the treatment and/or prevention of obesity. The invention relates also to the use of compounds capable of inhibiting in a mammal occurring de novo lipogenesis and which exert almost no effect on the Central nervous system (CNS), to obtain drugs intended for the treatment and/or prevention of obesity.

Currently continuously increases the severity of problems associated with poor health due to obesity (or obesity), mainly due to unbalanced diet and eating foods too high in fat, which is most important especially for industrialized countries. With the increase of the total population of a particular country in the percentage of people suffering from obesity, increases the number resulting from obesity complications, ranging from simple dissatisfaction with the man in their appearance and to cardiovascular diseases or even the development of some forms of diabetes. With this in mind, to date, have already been developed certain therapeutic methods aimed at the treatment or prevention of obesity. As an example, you can call the application inhibit the affected lipase compounds, the action is based on the decrease in the breakdown of fats in the intestine and reduce in this way the output energy in the digestive process. Thus, when such therapy dietary fats at least partially again excreted in not decayed. However, there is a need to develop new therapeutic approaches for the treatment and/or prophylaxis of obesity in addition to already known therapeutic methods.

With the invention it has been unexpectedly found that compounds, which are able to inhibit in mammals and especially in humans, occurring de novo lipogenesis, suitable for effective treatment and/or prevention of obesity. The best results are achieved with the use of these compounds over an extended period of time, for example within a few weeks.

In accordance with this object of the present invention is a method of identifying compounds that are useful for the treatment and/or prevention of obesity, namely, that select those compounds that are able to inhibit in mammals and especially in humans, occurring de novo lipogenesis. In addition, the object of the invention is the use of compounds which are able to inhibit in mammals and especially in man is ka occurring de novo lipogenesis and which exert almost no effect on the Central nervous system, for example anticonvulsant actions to obtain drugs intended for the treatment and/or prevention of obesity.

Under the concept of "originating de novo lipogenesis (below referred to as DN-L) is the synthesis of autologous fatty acids from carbohydrates occurring in mammals. This reaction synthesis occurs in the cytosol of somatic cells and is based on the so-called citric acid cycle or Krebs cycle-Martius'. In this cycle during autogenic biochemical reaction of two components, one of which is pyruvate, which are carbohydrates, and the other is hydrogen, through various intermediate compounds, which include, in particular, maleate, fumarate and α-Ketoglutarate, eventually forming citrate. While citrate, because it is synthesized in excess, can turn through an intermediate acetyl-coenzyme a in free fatty acid (lipid products), which are fats and which subsequently can accumulate in fat cells (adipocytes). Excess deposition formed from fatty acids of fats in somatic cells can generally lead to obesity. In the citric acid cycle involves various enzymes. Metabolism occurring in the citric acid cycle, to a significant extent the head of the Sith on the number of available for the reaction of bicarbonate. The number of such available for the reaction of bicarbonate in turn depends on the speed with which it may be formed from carbon dioxide. Catalyze such an equilibrium reaction which produces hydrogen, the so-called carbonic anhydrase. Among currently known of carbamides and their isozyme in the catalysis of reactions resulting in the formation of bicarbonate, is available to participate in the citric acid cycle, in mammals, mainly engaged in the isozyme of carbamides subtype II (CA II) and subtype V (SA V). The most important role for carbonic anhydrase subtype V, since they are present in the mitochondria. In these mitochondria also citric acid cycle.

All existing methods of inhibiting DN-L in mammalian cells aimed at reducing the intensity of metabolism occurring in the citric acid cycle. Thereby, it is possible to reduce the concentration produced in excess of citrate, available for fatty acid synthesis. According to the present invention to inhibit DN-L preferably through inhibition of carbamides catalyzing the reaction, which is formed participating in the citric acid cycle hydrogen. Thus according to the invention preferably inhibit ka is bangerz subtypes II and/or V, more preferably CA V.

Compounds that are capable of non-specific inhibition of carbonic anhydrase (i.e. non-specific or traditional CA-inhibitors), are known and long used in various therapeutic purposes, mainly as diuretics, or ophthalmology. Review the information about the applications of such traditional CA inhibitors can be found in ..Supuran, Expert Opinion on Therapeutic Patents, 10 (5) (2000), cc.575-600. In contrast, under "specific CA-inhibitors" refers to compounds that inhibit basically only one subtype of SA (e.g., SA (V) or a specific group SA subtypes. Up to this time there was no information about the use of traditional CA inhibitors for targeted treatment and/or prevention of obesity.

It is believed that the main cause of obesity in the vast majority of known cases is relatively high content of fat alien in human consumption of food. In highly developed countries such as the USA, the share of fat in total consumption of fat people food for up to 30%. In accordance with this current data leading to obesity fat is caused mainly by the excessive consumption of dietary fat that inhibition DN-L, accompanying the metabolism of carbohydrates, h is has no effect.

To date, the inhibition DN-L, though it can also reduce the accumulation of fatty acids in adipocytes, was considered due to the insignificant effect of DN-L on the accumulation of fatty acids in somatic cells as unpromising way of treatment and/or prophylaxis of obesity in humans. Summary information reflecting the opinion of most experts on this subject can be found, for example, M.K. Hellerstein, European Journal of Clinical Nutrition, 53 (1) (1999), cc.53-65. Such prevailing among experts opinion prevented hitherto targeted search or the development of new medicines for the treatment and/or prevention of obesity, the principle of which is based on the inhibition DN-HP

However, when developing the present invention it has been unexpectedly found that compounds capable of inhibiting DN-L in mammals and especially in humans, can be effectively used for the treatment and/or prevention of obesity and primarily with the introduction of these compounds to the appropriate patients for an extended period of time, for example within six weeks. In accordance with this prolonged inhibition DN-L can significantly reduce body weight in obese people, even though the actual process DN-L has only a minor effect is the accumulation of fat in adipocytes. It was a long inhibition DN-L, which in principle is accompanied by a slight one-time effect, renders the accumulating action in General and leads to a significant reduction of body weight.

The fact that proposed in the invention method really allows you to purposefully identify compounds suitable for the treatment and/or prevention of obesity, can be confirmed on the example of such compounds as topiramate. Topiramate is an anti-epileptic agent, known from application EP 0138441 A2. In addition, topiramate, known to possess wide pharmacological spectrum of action. Thus, in particular, topiramate can block depends on the voltage sodium channels and thereby to stabilize the membrane potential of the cell, it activates the receptor gamma-aminobutyric acid (GABA) and thereby enhances mediated GABA inhibition, it has the effect of inhibitor carbamides and in addition has the ability to inhibit the receptor kainate/AMPK (amino-3-hydroxy-5-methylisoxazole-4-propionic acid), which is a subtype of glutamate receptors, which is accompanied by inhibition induced AMPK flows. In accordance with this topiramate exerts pronounced effects on the Central nervous system. However, to date it remains unknown who Tim, does anticonvulsant properties of topiramate above factors have pharmacological value.

From the application WO 98/00130 it is known further that topiramate has certain pharmacological properties, by which he presumably suitable for the treatment of obesity. Similar properties of topiramate were discovered accidentally as a side-effect, discovered in the course of extensive studies on epilepsy patients. However, to date, failed to establish what exactly pharmacological properties of topiramate due to the weight loss observed in epilepsy patients. Topiramate belongs to a group possessing anticonvulsant activity of the compounds of General formula I, in which the application WO 98/00130 specified as suitable for the treatment of obesity.

Using the proposed invention the method failed to establish that topiramate is a high-level SA-inhibitor, especially a highly active inhibitor found in mammals of carbamides subtypes II and V, and that topiramate can effectively inhibit DN-L in mammalian cells. In accordance with these by using the proposed invention the method for the first time it was possible to confirm the fact that the pharmacological adverse action is s topiramate, which to date has not yielded to the explanation which comes to weight loss patients suffering from epilepsy, mainly due to its ability to effectively inhibit DN-L in mammals. With this in mind, it can be expected that proposed in the invention method in the long term will help to identify compounds suitable for the treatment and/or prevention of obesity. Thus, using the proposed in the invention of the method for the first time, you receive the ability to relatively quickly and easily identify pharmacologically active compounds, the action of which is based on the inhibition DN-HP the Present invention, thus, allows at least at first, the estimated stage of selection to identify without extensive and costly in vivo experiments, such as experiments with feeding experimental animals, the compounds whose action is based on the above principle and are suitable for the treatment and/or prevention of obesity.

Such results are unexpected given the fact that in earlier studies of the pharmacological properties of topiramate its SA-inhibiting efficiency has always been regarded as insignificant or even just as being too low and therefore not of interest from a therapeutic point of view (see, for example, ..Maryanoff al. Journal of Medicinal Chemistry, 30 (1987), cc.880-887; ..Maryanoff and others, Journal of Medicinal Chemistry, 41 (1998), cc.1315-1343; S.J.Dodgson and others, Epilepsia 41 (1), (2000), cc.35-39). In these earlier studies to assess SA-inhibiting activity of topiramate in each case used the experimental solutions containing CA various mammals, as well as the various components of the body, such as blood or tissue of the organ.

In accordance with the first embodiment proposed in the invention method is suitable for treatment and/or prevention of obesity connection, identify, selecting as those suitable compounds that can inhibit the activity of at least one occurring in mammals carbonic anhydrase. So, for example, to this end, at least one of the investigated connection, you can enter in contact with at least one carbonic anhydrase and then known method to identify compounds that inhibit the activity of this at least one carbonic anhydrase. In this embodiment proposed in the invention method, it is preferable to apply the carbonic anhydrase found in mammals, for example humans or rodents, in particular rats, mice or Guinea pigs, especially carbonic anhydrase subtypes II and/or V is Most preferable to apply carbonic anhydrase found in the body of man. Suitable for use in these purposes of carbonic anhydrase, for example, to allocate from the body of these mammals and, if necessary, clean or you can get them, preferably using known chemical or biotechnological methods.

In the first embodiment proposed in the invention method, the change of activity of carbamides under the influence of the studied compounds preferably be estimated using the well-known experiment for the determination of enzymatic activity in vitro, using carbonic anhydrase in the form of a dedicated and containing no impurities enzymes. It is preferable to use carbonic anhydrase obtained by chemical or biotechnological methods, because these methods allow you to get in their purest form. Experiments on determination of activity carbamides in vitro are well known in this field. The experiments on determination of enzymatic activity in vitro, which in accordance with the present invention allows to determine changes in the activity of carbamides include, for example, a method based on measuring changes in the pH values under the influence of carbamides (see ..Wilbur, N.G.Andersen, Journal of Biological Chemistry, 176 (1948), cc.147-154; G.Sanyal, Tnimage, Journal of Biological Chemistry, 256 (1981), cc.608-612), the method is stopped jet (see R.G.Khalifah, Journal of Biological Chemistry, 246 (1971), cc2561-2573) or method based on the use of 4-nitrophenylacetate (see Y.Pocker, J.T.Stone, Journal of the American Chemical Society, 87 (1965), cc.5497-5498). In the last of these methods determine the rate of hydrolysis of 4-nitrophenylacetate under the influence study of carbonic anhydrase, using the property of carbamides to have the same impact as an esterase.

In this regard, to determine the SA-inhibiting properties of the compounds according to the invention, it is preferable to use a method of testing described in ..Supuran and others European Journal of Medicinal Chemistry, 33 (1998)), SS-594 (see especially s) or A.Scozzafava and others in the Journal of Medicinal Chemistry, 42 (1999), cc.3690-3700 (see especially s). These are described in Supuran, etc. and Scozzafava and other testing methods included in the present description by reference. Connection values inhibitory concentration IC50which when they checked one of the above standard methods to determine the activity in vitro, described by Supuran, etc. or Scozzafava, etc. constitute at least 10 mcmole/l or less (corresponding to higher efficiency)can be selected in accordance with the present invention as suitable compounds with CA-inhibiting action (i.e. as CA-inhibitors). If you want to determine the activity of certain subtypes of human SA may be appropriate to use ins the methods other than based on the use of 4-nitrophenylacetate method. Thus, in particular, may be more appropriate to use methods that enable relatively quick to carry out the necessary reactions and to control their flow.

The test results for the determination of enzyme activity based on described by Scozzafava and others (see above) method using 4-nitrophenylacetate (originally described in Y.Pocker and J.T.Stone, Biochemistry, 6 (1967), cc.668-678), according to the present invention, it was found that topiramate exhibits a pronounced inhibitory effect on the obtained one of biotechnological methods of human carbonic anhydrase subtype II (IC50=5 nmole/l) and that this inhibitory effect is much stronger compared to the inhibiting action showed acetazolamide or methazolamide, which are the traditional CA-inhibitors and are used in experiments as control substances. Human carbonic anhydrase subtype II was obtained by the method described in Scozzafava and other

According to the results of the same test, it was found that topiramate has a pronounced inhibitory effect on the received biotechnological method of murine carbonic anhydrase subtype Va (mCA Va) (IC50=74 nmole/l). However, unlike described in cozzafava and other the Protocol of the experiment in this case, the enzyme mCA Va was used in a concentration of 120 nm. The enzyme mCA Va received a well-known way according to the method described in H.R.Heck and others in the Journal of Biological Chemistry, 269 (1994), cc.24742-24746. With this purpose used the bacterial strain Escherichia coli BL 21 (DE3), which was transfectional plasmid vector containing the coding mCA Va sequence under the control of the T7-promoter induced by isopropyl-β-thiogalactopyranoside (IPTG). Cultures of bacteria were made at 37°in a liquid environment, Luria-Bertani, supplemented with ampicillin (100 μg/ml), and the growth of the culture was monitored spectrophotometrically at a wavelength of 600 nm. Upon reaching the culture of bacteria exponential growth phase were added IPTG at a final concentration of 1 mmol/l After incubation for 3 h (at 37°and under stirring) of the bacteria culture was centrifuged for 15 min at 7000 g and the supernatant discarded. Obtained after centrifugation the residue (debris) was dissolved in a 0.1-fold volume of double-distilled water and was added lysozyme (100 µg/ml). Cells were literally sonification. With this purpose, aliquots of 10 ml of the resulting suspension of bacteria were placed in open-top glass vessel and each sample was treated with ultrasound for 4 times for 3 minutes After each ultrasonic pulse was determined by the absorption of radiation treatment who were at a wavelength of 600 nm, for this, 100 μl of each sample was diluted with 900 μl of double-distilled water. The process was completed at the time when the absorption of radiation by the sample at a wavelength of 600 nm was reduced to approximately 1/10 of its original level. After lysis of the cells was added CaCl2buffer for binding (Stratagene) and the resulting cell lysate was purified on a column of resin having affinity to calmoduline. This method of cleaning is based on the high affinity associated with resin domains of calmoduline to communicate with calmoduline peptide tag, which is present at the N end of the expressed protein mCA Va. Purification was performed by a known method (see reference Affinity LIC Cloning and Protein Purification Kit Manual" company Stratagene).

In accordance with the second option proposed in the invention method is suitable for treatment and/or prevention of obesity compounds identified by selecting those compounds that are able to reduce the amount of metabolites formed in the citric acid cycle, taking place in a dedicated living mammalian cells or lipid products generated in the citric acid cycle. Acceptable metabolites, which are formed in the citric acid cycle and the number of which is reduced under the influence of the studied compounds on measurable value, are soluble is in acid metabolites, such as citrate, maleate, fumarate and/or α-Ketoglutarate. Preferred while the citrate. Other acceptable metabolites are lipid products generated in the citric acid cycle, such as free fatty acids. In this second embodiment proposed in the invention method ultimately also determine the ability of these compounds to inhibit the activity of at least one occurring in mammals, carbonic anhydrase, however, applied in this case, the test model differs from the one used in the above-described first embodiment. The principle underlying the method of testing used in the second embodiment of the method according to the invention, is that a known way to determine the absorption of radiation emitted by the labeled isotope14With the substrates used in the citric acid cycle, a metabolic intermediate or final products produced in the citric acid cycle, taking place in a dedicated living mammalian cells, and to compare the obtained results with results obtained in other respects under the same conditions, but under the influence with SA-inhibiting action of the compounds. In this embodiment proposed in which sobienie method, it is preferable to use a dedicated living cells such rodents, as rats, mice or Guinea pigs, or humans. Preferred with human cells. As rodent cells are adipocytes and hepatocytes. Preferred in this rodent adipocytes. As human cells can also be used adipocytes and hepatocytes. Preferred in this human hepatocytes. Used in this embodiment of the method of mammalian cells can be obtained by conventional methods of cultivation and/or cloning. Use may be made of natural or modified by biotechnological methods mammalian cells. If you checked the connections above described method is required to determine the absorption of radiation soluble in acid intermediate products formed in the citrate cycle, as labeled isotope14With the substrate, it is preferable to use14With-hydrogen (i.e. NaH[14S]O3). If when testing compounds by the method described above is required to determine the absorption of radiation end products formed in the citrate cycle, as labeled isotope14With the substrate, it is preferable to use [U14C]glucose.

In this second embodiment proposed in the invention method, the ability of the compounds inhibit is to DN-L in mammals, it is preferable to determine according to the method, described in S.A.Hazen and others in the FASEB Journal, 10 (4) (1996), cc.481-490. This is described in Hazen and other testing method included in the present description by reference. Compounds that, when checked using the above method, described by Hazen and others, can greatly inhibit the absorption of radiation emitted by the labeled isotope14With substances which can serve as precursors in the citric acid cycle, for example the absorption of the emitted14C-bicarbonate radiation citrate, maleate, fumarate and/or α-Ketoglutarate, i.e. the connection ID50-the importance of which in accordance with this rate of at least 10 mcmole/l or less (corresponding to higher efficiency)can be selected in accordance with the present invention as suitable compounds that meet certain criteria.

According to the results of the test, which is described in Hazen and other method of testing and conducted in accordance with the present invention, it was found that topiramate exhibits a pronounced inhibitory effect on the formation of soluble acid metabolites in the citric acid cycle occurring in the received biotechnological method rat adipocytes line 3T3-F442A. Similar inhibitory effect of topiramate (IC =348 nmole/l) significantly exceeded the traditional CA-inhibitor, as ethoxzolamide, which was used as a control substance.

According to the most preferred option proposed in the invention, a method of identifying compounds suitable for the treatment and/or prevention of obesity, select those compounds which in accordance with the above-described first embodiment of the method, primarily on the results of the above test for determination of enzyme activity in vitro, were selected as suitable for their ability to inhibit at least one occurring in mammals, carbonic anhydrase and additionally in accordance with the above-described second embodiment, the method has also been selected as suitable for their ability to reduce the amount of metabolites formed in the citric acid cycle, taking place in a dedicated live mammalian cells. The first variant of the method allows one to quickly and efficiently evaluate the ability of compounds to inhibit occurring in mammals carbonic anhydrase. The second variant implementation of the method allows to obtain, in particular, the primary data of the studied compounds the ability to penetrate into the mitochondria alive the x mammalian cells, in which localized SA V. For the selection of suitable compounds of the above-described first and second embodiments of the proposed invention the method can be performed in parallel or sequentially in any order.

According to the invention of compounds that can inhibit DN-L in mammals, suitable for the production of pharmaceuticals for the treatment and/or prevention of obesity. Thus select those compounds that are capable of inhibiting at least one carbonic anhydrase found in mammals. It is obvious that the selected compounds should be physiologically compatible and satisfy the General requirements for medicinal active substances, such as requirements regarding safety and tolerability. It is preferable to use compounds that can inhibit occurring in mammals carbonic anhydrase subtypes II and/or V. Most preferred in this connection that are able to specifically inhibit the CA II and/or SA V, first of all SA V.

Currently known connections, such as topiramate, which have a SA-inhibiting effect and the long-term use in patients a decrease in the weight of their body, also have a pronounced effect on the Central nervous system, in private the ti anticonvulsant action. However, the presence of side effects on the Central nervous system connections, intended for use over an extended period of time for treatment and/or prevention of obesity, is often undesirable. Therefore, compounds that are in accordance with the proposed invention method were selected as suitable for treatment and/or prevention of obesity, can also be checked to have any action on the Central nervous system. For example, such connection if they have anticonvulsant properties. For the study compounds have anticonvulsant properties can be used, for example, so-called "test with super-maximum electroshock" (SAS test, sometimes also called "test with maximum shock" or the abbreviation mash-test)described in G.Chen, etc. in the Proceedings of the Society for Experimental Biology and Medicine, 87 (1954), cc.334-339, and J.E.P.Toman and others in the Journal of Neurophysiology, 9 (1946), s. This is described in Chen et al and Toman and other testing method included in the present description by reference. Thus, in particular, compounds intended for administration to patients for a long period of time for treatment and/or prevention of obesity, almost should not show any activity in their study using the above SAS-test, described by Chen et al and Toman and others, and in their application and in high doses, with the element of at least 100 mg/kg when administered orally, preferably should not take actions according to the test criteria are considered significant (i.e. according to the G.Chen and other "protective dose" PD50should be ≥100 mg/kg when administered orally, when specified in the work G.Chen and other values PD50essentially correspond widely used to refer to dose the concept of "minimum effective dose", MED). Compounds that were selected as appropriate in accordance with the proposed invention is a method and that additionally in the MES test, described by Chen et al, showed almost no activity, the most suitable for the production of pharmaceuticals for the treatment and/or prevention of obesity. Above SAS - (respectively MES) tests are standard pharmacological tests that can be conducted by conventional methods providing similar services organizations and firms (for example, the firm "Panlabs").

Compounds that, in accordance with the proposed invention method were selected as suitable for treatment and/or prevention of obesity, it is possible by the conventional technology to recycle as active ingredients together with customary pharmaceutical auxiliary substances in glenavy compositions, such as tablets,capsules, suppositories or solutions. Such glenavy composition can be obtained by the known methods using conventional solid or liquid carriers, such as lactose, starch or talcum powder, or liquid paraffins and/or using conventional pharmaceutical auxiliary substances, such as leavening for tablets, hydrotropic of solubilization or preservatives. Pharmaceutical compositions suitable for use according to the invention, are known, for example, from application EP 0138441 A2, as well as from the publication WO 98/00130.

1. Method of identifying compounds suitable for the treatment and/or prevention of obesity, characterized in that as fit, select those compounds that can inhibit the activity of at least one carbonic anhydrase found in mammals.

2. The method according to claim 1, characterized in that at least one compound is administered in contact with at least one carbonic anhydrase and then identify those compounds that can inhibit the activity of this at least one carbonic anhydrase.

3. The method according to claim 1, characterized in that the use of carbonic anhydrase found in the body of rodents and humans.

4. The method according to claim 3, characterized in that the use of carbonic anhydrase found in the human body.

5. The method according to claim 1, otlichayushiesya, the use of carbonic anhydrase subtypes II and/or V, occurring in mammals, especially humans.

6. The method according to claim 5, characterized in that the use of carbonic anhydrase subtype V, occurring in mammals, especially humans.

7. The method according to claim 2, characterized in that the use of carbonic anhydrase, which are presented in the form of selected enzymes obtained by chemical or biotechnological method, with changes in the activity of carbamides under the influence of the studied compounds determined in the test for determination of enzyme activity in vitro.

8. The method according to claim 1, characterized in that as fit, select those compounds that are able to reduce the amount of metabolites formed in the citric acid cycle, taking place in a dedicated living mammalian cells.

9. The method of claim 8, wherein the determined reduction resulting in what is happening in the cells of the citric acid cycle citrate, maleate, fumarata, α-Ketoglutarate and/or reduce the formation of lipid end products of the citric acid cycle under the influence of the studied compounds.

10. The method of claim 8, wherein using the selected living cells of rodents or humans.

11. The method according to claim 10, characterized in that the use of adipocytes or hepatocytes.

12. With whom persons identifying compounds who is suitable for treatment and/or prevention of obesity, characterized in that select those compounds that are identified as suitable by the method according to claim 7 and additionally the method according to item 8.

13. Method of identifying compounds that are useful for the treatment and/or prevention of obesity, with select those compounds that are identified as suitable connection method according to claim 1 and which additionally on the results of a standard test for the determination of the anticonvulsant properties identified as practically does not exhibit activity.

14. The use of compounds able to inhibit at least one occurring in mammals, carbonic anhydrase, for obtaining a medicinal product intended for the treatment and/or prevention of obesity.

15. Application for 14 compounds able to inhibit occurring in mammals carbonic anhydrase subtypes II and/or V

16. Application for 14 compounds, which practically does not possess anticonvulsant properties.

17. The use of compounds that are first identified by the method according to claim 1 as suitable for treatment and/or prevention of obesity, for obtaining a medicinal product intended for the treatment and/or prevention of obesity.



 

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FIELD: medicine.

SUBSTANCE: method involves evaluating patient state using Hamilton scale, considering leukocyte formula of general clinical blood analysis for determining infrared spectroscopy absorption spectrum values within 30 s in the bandwidth of 3085-2832 cm-1 and 1543-1425 cm-1, respectively. Mean value of depression intensity being equal to 21.0±2.8 points, mean absorption values being equal to 32.3±1.8% and 35.0±2.2% in the bandwidth of 3085-2832 cm-1 and 1543-1425 cm-1, respectively and normal lymphocyte and monocyte content being observed, reactive depression is to be diagnosed. Mean absorption values being equal to 40.5±3.2% and 43.4±2.9% in the bandwidth of 3085-2832 cm-1 and 1543-1425 cm-1, respectively in combination with immunity characteristics like monocytosis>5% and lymphocytosis>25% leukocyte formula of general clinical blood analysis being observed, therapeutically resistant reactive depression cases are to be diagnosed.

EFFECT: high accuracy of diagnosis in differentiating cases showing positive changes as response to pharmacotherapy from therapeutically resistant cases.

1 tbl

FIELD: analytical methods in biology.

SUBSTANCE: invention relates to chemical analysis methods for biological materials and, in particular, concerns a method for determining acetylcholine in biological specimens consisting in incubation of a specimen, centrifugation, take-off of supernatant, preparation of alkaline hydroxylamine reagent by adding equal volume of 3.5 n NaOH to hydroxylamine chloride solution, mixing of thus prepared reagent with test specimen, and addition of iron chloride solution followed by measurement of optical density. Invention is characterized by that, before addition of 3.5 n NaOH, hydroxylamine chloride solution is mixed with 10-14% solution of Na2CO3, incubation of specimen is performed after the latter is mixed with alkaline hydroxylamine reagent at 1:2 ratio for 90-120 min at 33-37°C, centrifugation is preceded by adding aqueous hydrochloric acid solution, and 0.74 M iron chloride solution is added to taken off supernatant.

EFFECT: increased determination accuracy.

2 tbl, 3 ex

FIELD: veterinary medicine, biochemistry.

SUBSTANCE: the method deals with treating the smears with buffer mixture followed by washing with distilled water, drying, additional dyeing with 0.5%-methylene blue followed by washing, drying, detecting enzymatic activity by the quantity of dyed granules. The innovation enables to widen technological possibilities in the field of immunological investigations.

EFFECT: higher accuracy of detection.

5 dwg, 1 ex, 1 tbl

FIELD: food additives.

SUBSTANCE: it has been developed both the dietetic composition and method in which one should apply the foundation as food for mammalians and an active component being an estrogen, androgen or their mixture at the quantity to be sufficient for preventing body weight gain usually happened in mammalians after delayed maturation of reproductive organs, castration, ovariectomy or hysterectomy, or during post-climacteric period. Preferably, active component is being a phytoestrogen, phytoandrogen or their mixture, at the quantity 0.001-10 weight% against the weight of composition. Application of compositions efficiently prevents body weight gain in mammalians.

EFFECT: higher efficiency.

4 ex, 3 tbl

FIELD: organic chemistry, medicine, pharmacy.

SUBSTANCE: invention relates to derivatives of imidazole of the formula (I):

or its pharmaceutically acceptable salts wherein X represents -CH2-(CH2)p-, -O-; R1 represents phenyl, naphthyl, 1,2,3,4-tetrahydronaphthyl, (C3-C7)-cycloalkyl wherein indicated phenyl, naphthyl, 1,2,3,4-tetrahydronaphthyl, (C3-C7)-cycloalkyl are substituted optionally with 1-3 substitutes taken independently among halogen atom, -OH, halogen-(C1-C6)-alkyl, (C1-C6)-alkyl, (C1-C6)-alkoxy group and OH-(C1-C6)-alkyl; R2 represents hydrogen atom (H) or (C1-C6)-alkyl; R3 represents H or (C1-C6)-alkyl; R4 represents H or (C1-C6)-alkyl; R5 represents H, or R5 and R7 form in common a bond; each R6 represents independently halogen atom, -OH, halogen-(C1-C6)-alkyl, (C1-C6)-alkyl, (C1-C6)-alkoxy group or OH-(C1-C6)-alkyl; R7 represents H, or R7 and R5 form in common a bond; each R8 represents independently -OH, (C1-C6)-alkyl, halogen-(C1-C6)-alkyl or (C1-C6)-alkoxy group; m = 0, 1, 2 or 3; n = 0 or 1; p = 0 or 1; r = 0 or 1; t = 0. Also, invention relates to a method for preparing compounds of the formula (I) and to a pharmaceutical composition showing affinity to alpha-2-adrenoceptors based on these compounds. Invention provides preparing new compounds and pharmaceutical composition based on thereof used in aims for treatment of neurological disturbances, psychiatric disorders or disturbances in cognitive ability, diabetes mellitus, lipolytic diseases, orthostatic hypotension or sexual dysfunction.

EFFECT: improved preparing method, valuable medicinal properties of compounds and compositions.

25 cl, 1 tbl, 14 ex

Antagonist npy y5 // 2264810

FIELD: medicine, pharmacology.

SUBSTANCE: the present innovation deals with applying pharmaceutical composition as an antagonist of NPY Y5 receptor that contains the compound of formula I

, moreover, it deals with compounds of formula I and method for treating obesity and suppressing food intake, as well.

EFFECT: higher efficiency of therapy.

18 cl, 13 ex, 6 tbl

FIELD: organic chemistry, medicine, pharmacy.

SUBSTANCE: invention relates to substituted 8,8a-dihydro-3aH-indeno[1,2-d]thiazoles and to their physiologically acceptable salts and physiologically functional derivatives also. Invention describes compounds of the formula (I): wherein R1 and R1' mean independently of one another H, F, Cl, Br, J; R2 and R3 means H; R4 means phenyl hat can be replaced with hydroxyl group (OH); R5 means hydrogen atom (H); R6 means OH. Also, invention describes a method for preparing these compounds. Compounds can be used as anorexic agents for prophylaxis and treatment of obesity.

EFFECT: improved preparing method, valuable medicinal properties of compounds.

5 cl, 2 tbl, 1 ex

FIELD: organic chemistry, chemical technology, medicine.

SUBSTANCE: invention relates to polycyclic dihydrothiazole and to their physiologically acceptable salts and physiologically functional derivatives. Invention describes compounds of the formula (I): wherein R1 and R1' mean independently of one another atoms of hydrogen (H), fluorine (F), chlorine (Cl), bromine (Br) and iodine (J); R2 and R3 mean hydrogen atom (H); R4 means (CH2)n-R5 wherein n can be = 0-6; R5 means phenyl that can be substituted with NH-SO2-(C1-C6)-alkyl, NH-SO2-phenyl being phenyl ring up to twice-fold can be substituted with chlorine atom (Cl), (CH2)m-SO2-NH2, (CH2)-SO2-NH-(C1-C6)-alkyl, (CH2)m-SO2-N-[(C1-C6)-alkyl]2 or (CH2)m-SO2-N-[=CH-N(CH3)2] wherein m can be = 0-6, and a method for their preparing. Compounds are useful, for example, as anorexic agents used in prophylaxis or treatment of obesity.

EFFECT: improved preparing method, valuable medicinal properties of compounds.

5 cl, 12 tbl, 2 ex

FIELD: medicine, experimental medicine.

SUBSTANCE: one should introduce tripeptide Pro-Gly-Pro for laboratory animals as injections at the quantity of 0.09-1.0 mg/kg body weight, and, also, gelatin as fodder additive. The method suggested enables to suppress appetite, decrease the quantity of fodder intake that leads to decreased body weight as a result.

EFFECT: higher efficiency.

2 cl, 5 dwg, 5 ex

FIELD: organic chemistry, medicine.

SUBSTANCE: invention relates to polycyclic dihydrothiazoles and their physiologically acceptable salts and physiologically functional derivatives. Invention describes compounds of the formula (I): wherein r1 and R1' mean independently of one another atoms H, F, Cl, Br and J; R2 means hydrogen atom (H); R3 means chlorine (Cl), bromine (Br) atom; R4 means phenyl, and a method for their preparing. Compounds can be used, for example, as anorectics for prophylaxis or obesity treatment.

EFFECT: improved preparing method, valuable medicinal properties of compounds.

5 cl, 2 tbl, 1 ex

FIELD: organic chemistry, medicine, pharmacy.

SUBSTANCE: invention relates to a prophylactic or therapeutic agent used against hyperlipidemia and comprising as an active component the heterocyclic compound of the formula [1]:

or its pharmaceutically acceptable salt wherein R1 represents aryl optionally substituted with similar or different one-three groups taken among alkyl, halogenalkyl, trihalogen alkyl, alkoxy-group and halogen atom; Het represents bivalent aromatic heterocyclic group of the formula [5]:

wherein X represents oxygen, sulfur atom or NR6 wherein R6 represents hydrogen atom or alkyl; R2 represents hydrogen atom, alkyl or trihalogenalkyl; D represents alkylene and alkenylene; E represents group of the formulae [3] or [4] wherein Y represents oxygen or sulfur atom; R3 and R4 are similar or different and each represents hydrogen atom or alkyl; p = 1; Z represents carboxy-group, alkoxycarbonyl, cyano-group or 1H-5-tetrazolyl. Also, invention relates to new compounds belonging to group of above enumerated heterocyclic compounds of the formula [1] that show effect reducing blood triglycerides level, low density lipoprotein cholesterol, glucose and insulin or effect enhancing high density lipoprotein cholesterol and effect reducing the atherogenic effect. Therefore, these compounds can be used in prophylaxis or treatment of hyperlipidemia, arteriosclerosis, heart ischemic disease, brain infarction, rheocclusion after percutaneous intraluminal coronary angioplasty, diabetes mellitus and obesity.

EFFECT: valuable medicinal properties of compounds and pharmaceutical composition.

29 cl, 1 tbl, 170 ex

FIELD: endocrinology.

SUBSTANCE: abnormally corpulent persons are treated by reduced-caloricity diet with limited content of carbohydrate-containing components and fats. In particular, caloricity of meal is reduced to 1200 kcal, including carbohydrate-containing components with glycemic index below 40. When initial weight is reduced by 5% and the weight is stabilized for 3 months, caloricity is increased to a specified value, calculated in terms of formula for daily caloricity recommended by World Health Organization taking into account sex, age, weight, and physical activity, glycemic index of carbohydrate-containing components ranging from 40 to 69 until the weight is lowered to desired level.

EFFECT: achieved stable and long-term reduction of weight owing to lowered insulin resistance of organism.

3 tbl

FIELD: cosmetics, pharmaceutical chemistry.

SUBSTANCE: invention relates to cosmetic composition used for loss weight comprising at least one compound that induces producing IL-6 by adipocytes in form of mixture with NPY antagonist and/or α2 antagonist and with an excipient for the cosmetic variant. Invention provides effectiveness of fatty acids efflux from adipocytes and inhibitory effect on fatty acids incorporation into cells that results to the loss weight effect. The composition shows good stability in storage being without reducing its activity. The composition has no adverse effects.

EFFECT: valuable properties of composition.

11 cl, 3 dwg, 5 ex

FIELD: medicine, obstetrics, gynecology.

SUBSTANCE: one should intravenously inject Reamberin 1.5% up to 400 ml solution daily, percutaneously affect with magneto-infrared laser, wave length being 0.89 mcm, impulse power of infrared radiation corresponds to 4 W. One should affect uterine and adnexal area alternatively, at exposure being about 2-3 min for every region, radiation frequency being 5 Hz and 50 Hz altered every other day. Since the second day of therapy one should additionally supply laser light flow directly towards uterine area through vaginal arch for 1 min at frequency of 1000 Hz. Total course of therapy corresponds to about 5-7 d. The innovation enables to increase immunological body strength, decrease relapse frequency and decrease side effects of curative preparations applied.

EFFECT: higher efficiency of therapy.

2 ex, 1 tbl

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