Method for decreasing of nutrient availability and method for appetite suppression
SUBSTANCE: invention relates to method for decreasing of nutrient availability, method for appetite suppression or reducing of food consumption. Claimed method includes application of PYY substance (YY peptide) or PYY agonist in peripheral administration. Method provides decreasing of nutrient availability in mouse females hungered for one night, reducing stomach ejection, inhibition of acid secretion in stomach, preventing cholecyst ejection, and inhibition of amylase secretion in pancreas.
EFFECT: preparation for decreasing of nutrient availability of improved effect.
35 cl, 10 ex, 8 dwg, 1 tbl
CROSS-REFERENCE TO RELATED APPLICATION
This application claims priority of the provisional application for the grant of a US patent application No.60/256216, entitled "Peptide YY and peptide agonists YY for the treatment of obesity, diabetes and other metabolic disorders," filed December 15, 2000, the contents of which are incorporated in this description by reference.
The SCOPE of the INVENTION
This invention relates to methods and compositions for treating metabolic conditions or disorders, in particular those which can be alleviated by reducing the availability of high-calorie substances, for example for the treatment of diabetes, obesity, eating disorders, syndrome of insulin resistance (syndrome X), intolerance to glucose, dyslipidemia and cardiovascular diseases.
A number of related hormones are a family of pancreatic polypeptide (PP). Pancreatic polypeptide was opened in the form of impurities in extracts of insulin and was named first by the authority of its origin, and not according to the functional value (Kimmel, Pollock et al. Endocrinology 83: 1323-30, 1968). PP is a 36-amino acid peptide [SEQ ID NO: 1], containing the characteristic structural motifs. Later related peptide was opened in extracts of intestine and called peptide YY (PYY) due to the presence of N - and C-terminal tyrosines (Tatemoto. Proc. Natl. Acad. Sci. USA 79: 2514-8, 1982) [SEQ ID NO: 2]. POS is her brain extracts was detected third related peptide and called neuropeptide Y (NPY) (Tatemoto. Proc. Natl. Acad. Sci. USA 79: 5485-9, 1982; Tatemoto, Carlquist et al. Nature 296: 659-60, 1982) [SEQ ID NO: 4].
It was reported that three of these peptides cause different biological effects. The effects of PP include inhibition of secretion of pancreas and relaxation of the gallbladder. When the Central introduction PP is a moderate increase in food intake, which may be mediated by receptors localized in the hypothalamus and the brain stem (review (Gehlert. Proc. Soc. Exp. Biol. Med. 218: 7-22, 1998)).
The release of PYY [SEQ ID NO: 2] occurs after a meal. Alternative molecular form of PYY is PYY[3-36] [SEQ ID NO: 3] (Eberlein, Eysselein et al. Peptides 10: 797-803, 1989) (Grandt, Schimiczek et al. Regul. Pept. 51: 151-9, 1994). This fragment is about 40% of total PYY-like immunoreactivity in the intestinal extracts of human and dog, and about 36% of the total PYY immunoreactivity in the plasma in a state of hunger to just over 50% after a meal. Apparently, he is a product of the splitting of PYY by dipeptidyl peptidase-IV (DPP4). Reportedly PYY[3-36] is a selective ligand receptor Y2 and Y5, which, apparently, are pharmacologically unique preference relation shortened at the N-end (i.e. the C-terminal fragments) and analogs of NPY. Reportedly with the peripheral introduction PYY reduces the secretion of gastric acid, the motility of the stomach, Exo is rinnooy secretion of the pancreas (Yoshinaga, Mochizuki et al. Am. J. Physiol. 263: G695-701, 1992), (Guan, Maouyo et al. Endocrinology 128: 911-6, 1991), (Pappas, Debas et al. Gastroenterology 91: 1386-9, 1986), the contraction of the gall bladder and intestinal motility (Savage, Adrian et al. Gut 28: 166-70, 1987). The effect of Central injection of PYY on gastric emptying, motility of the stomach and the secretion of gastric acid, which is observed after direct injection into the hind brain/brain stem or around the specified location (Chen and Rogers. Am. J. Physiol. 269: R787-R792, 1995), (Chen, Rogers et al. Regul. Pept. 61: 95-98, 1996), (Yang and Tache. Am. J. Physiol. 268: G943-8, 1995), (Chen, Stephens et al. Neurogastroenterol. Motil. 9: 109-116, 1997), may be different from the effect observed in peripheral injection. For example, when the Central introduction of PYY has some impact, the opposite effects listed in this description and in the case of peripheral injection of PYY[3-36], which was not the inhibition and stimulation of secretion of gastric acid. The motility of the stomach was suppressed only with the TRH stimulation, but not suppressed under a separate introduction, and in fact was stimulated at high doses, presumably due to interaction with receptors PP. It is shown that PYY stimulates the consumption of food and water after Central administration (Morley, Levine et al. Brain Res. 341: 200-203, 1985), (Corp, Melville et al. Am. J. Physiol. 259: R 317-23, 1990).
In this way one of the Central effects of NPY [SEQ ID NO: 4] in the early message was increasing p the consumption of food, in particular the effect on the hypothalamus (Stanley, Daniel et al. Peptides 6: 1205-11, 1985). It is reported that PYY and PP simulate these effects and that PYY is more efficient or effective as well as NPY (Morley, J.E., Levine, A.S., Grace, M., and Kneip, J. Brain Res. 341: 200-203, 1985), (Kanatani, Mashiko et al. Endocrinology 141: 1011-6, 2000), (Nakajima, Inui et al. J. Pharmacol. Exp. Ther. 268: 1010-4, 1994). Several groups found that the value of the induced NPY in food intake is higher than consumption, induced any previously tested pharmacological tool, as well as extremely long. Induced NPY stimulation of food intake reproduced in a number of species. Of the three main nutrients macronutrients (fats, proteins and carbohydrates) is preferably stimulated absorption of carbohydrates. Was not observed tolerance towards vozbujdayuschego appetite effects of NPY, and in the case where the introduction of peptide was repeated for 10 days, there was a marked increase in the rate of weight gain. After fasting concentration of NPY in the hypothalamus VAT has increased over time and quickly returned to control levels after a meal.
Pharmacological studies and attempts at cloning has revealed a number of receptor peptides of the family of PP, with seven transmembrane domains, and these receptors are named from Y1 to Y6 (and estimated cocktail recipes. is, preferably interacting with PYY, or Y7). The usual responses by signaling of these receptors are similar to the responses of other receptors associated with Gi/Gonamely , the inhibition of adenylate cyclase. Even at a relatively low homology sequences among receptors, there is no doubt that there are similarities in the formation of clusters of amino acid sequence between receptors Y1, Y4 and Y6, while Y2 and Y5 have the characteristics of other families. Other binding sites were identified on the basis of the rank order of effectiveness of different peptides. Receptor, preferably interact with NPY, which has not been cloned, named Y3, and also shows that there is a receptor, preferably interacting with PYY (estimated Y7) (review in (Michel, Beck-Sickinger et al. Pharmacol. Rev. 50: 143-50, 1998), (Gehlert, D.R. Proc. Soc. Exp. Biol. Med. 218: 7-22, 1998)).
It was assumed that the receptor Y5 and Y1 are the primary mediators of the response, manifested in the form of food consumption (Marsh, Hollopeter et al. Nat. Med. 4: 718-21, 1998), (Kanatani, A., Mashiko, S., Murai, N., Sugimoto, N., Ito, J., Fukuroda, T., Fukami, T., Morin, N., MacNeil, D.J., Van der Ploeg, L.H., Saga, Y., Nishimura, S., and Ihara, M. Endocrinology 141: 1011-6, 2000). The prevailing ideas was the idea that endogenous NPY through these receptors enhances the behavior associated with the intake of food. We offer therapy for obesity has consistently been focused on antag the lowland of NPY receptors, while therapy in the case of treatment of anorexia was converted to agonists specified family of ligands (see, for example, U.S. patents No.5939462; 6013622 and 4891357). In General, it was reported that PYY and NPY are equivalent and equally effective in all conducted tests receptors Y1 and Y5 (and Y2) (Gehlert, D.R. Proc. Soc. Exp. Biol. Med. 218: 7-22, 1998).
The main characteristics of the assumed receptor Y3 is that they learn NPY, whereas PYY is at least an order of magnitude less effective. The Y3 receptor is a receptor with a single binding site, which shows a preference in relation to NPY.
There is an additional binding site/receptor, which shows preference towards PYY called PYY-preferring receptor, which in this description referred to as the receptor(s) Y7. There are reports of different ranking orders binding with the specified receptor or class of receptors, suggesting that this family can be more than one receptor. In most cases this is the name used to describe a receptor in the case when PYY was three to five times more effective than NPY. Recommendations of the International Union of pharmacology regarding the nomenclature of receptors for NPY, PYY and PP are that the term PYY-preferring receptor not used in the case if not there is a difference in effectiveness between PYY and NPY, which shall be at least twenty times. (Michel, M.C., Beck-Sickinger, A., Cox, H., Doods, H.N., Herzog, H., Larhammar, D., Quirion, R., Schwartz, T., and Westfall, T. Pharmacol. Rev. 50: 143-50, 1998). However, for the purposes of this description, reference to a Y7 receptor or pharmacology PYY-preferring receptor means, the receptor has a certain degree of preference for PYY compared to NPY.
Obesity and related disorders are common and represent a very serious public health problems in the United States and around the world. A high degree of obesity of the body is the most powerful known risk factor for diabetes mellitus type 2 is a high risk factor for cardiovascular disease. Obesity is a recognized risk factor in the case of hypertension, atherosclerosis, congestive heart failure, stroke, gallbladder disease, osteoarthritis, sleep apnea, reproductive disorders such as polycystic ovarian malignant tumors of the breast, prostate, and colon and increased incidence of complications of General anesthesia (see, for example, (Kopelman. Nature 404: 635-43, 2000)). Obesity reduces life expectancy and entails a serious risk of the combination of the above diseases, as well as such Zabol the requirements, as infections, varicose veins, acanthosomatidae, eczema, intolerance to exercise, insulin resistance, hypertension, hypercholesterolemia, gall-stone disease, injury to the musculoskeletal and thromboembolic disease (Rissanen, Heliovaara et al. BMJ 301: 835-7, 1990).
Obesity also is a risk factor for the state group called the syndrome of insulin resistance or syndrome X". Recent estimates of the cost of treatment of obesity and related disorders at $ 150 billion dollars worldwide. It is assumed that the pathogenesis of obesity is multifactorial, but the main problem is that in subjects with obesity is not achieved balance between nutrient availability and cost of energy as long as there is excess adipose tissue. Obesity is currently poorly treatable, chronic, essentially intractable metabolic disorder. Therapeutic drug, applicable for reducing the weight of obese people can have a powerful healing effect on their health.
All documents referred to in this description included in this application by reference as if fully stated herein.
In contrast to reports about the activity of the x, show representatives of the family of pancreatic polypeptide in the Central introduction, it was found that when the peripheral introduction agonists PYY and PYY reduces nutrient availability and applicable for the treatment of obesity and related disorders. This invention presents a composition of PYY and PYY agonists and their use to modulate the availability of nutrients in a patient for the treatment of metabolic disorders that can be cured by reducing the availability of nutrients. These methods will be applicable, for example, in the treatment of obesity, diabetes, including, but not limited specified, type 2 diabetes or non-insulin-dependent diabetes, eating disorders, syndrome of insulin resistance and cardiovascular disease.
The term "PYY" means a polypeptide peptide YY, obtain or produce of any kind. Thus, the term "PYY" includes both full-size 36-amino acid peptide person specified in SEQ ID NO: 2, and species variants PYY, including, for example, PYY mouse, hamster, chicken, bovine, rat and dog. "The PYY agonist" refers to any compound that causes the effect of PYY, reducing the availability of nutrients, such as compound (1)having activity in the analysis of food intake, the emptying of the stomach, pancreatic secretion is EleSy or weight loss, described in this description in examples 1, 2, 5, or 6, and (2)that specific contacts in the analysis of receptor Y (example 10) or in the analysis of competitive binding with labeled PYY or PYY[3-36] some fabrics, having an excessive amount of Y receptors, including, for example, area postrema (example 9), the PYY agonist is not pancreatic polypeptide. Preferably PYY agonists can bind in such assays with an affinity of greater than 1 μm, and more preferably with affinity greater than 1-5 nm.
Such agonists can include a polypeptide having a functional domain PYY, an active fragment of PYY or a chemical or a small molecule. The PYY agonist may be a peptide or ones connections and include "agonistic analogs of PYY," and this term refers to any compound that is similar in structure with PYY, which has activity usually by binding or other direct or indirect interaction with the PYY receptor or other receptor or receptors with which it can interact with itself PYY, causing a biological response. Such compounds include derivatives of PYY, elongated PYY molecules with more than 36 amino acids, shortened PYY molecules with less than 36 amino acids, and substituted PYY molecules with one or more other amino acids, or any combination of compounds, perejil is the R above. Such compounds can also be modified by this method, as amidation, glycosylation, acylation, sulfation, phosphorylation, acetylation and cyclization.
One such agonistic analogs of PYY is PYY[3-36], identified herein as SEQ ID NO: 3. Polypeptides with numbers in brackets refer to a shortened polypeptides having the sequence of the full-length peptide over the provisions of the amino acids indicated in parentheses. Thus, PYY[3-36] has a sequence identical to PYY for amino acids 3 to 36. Agonist PYY may contact the PYY receptor with higher and lower affinity, to be longer or shorter half-life in vivo or in vitro, or is more efficient or less efficient than native PYY.
The term "condition or disorder that can be alleviated by reducing the availability of high-calorie (or nutritional) substances"means any condition or disorder in a subject, which is either caused by, complicated by, or exacerbated by the relatively high nutrient availability, or which can be relieved by reducing the availability of nutrients, for example, by reducing food intake. Such conditions or disorders include, but are not ogran is obtained specified, obesity, diabetes, including type 2 diabetes, eating disorders and the syndrome of insulin resistance.
In one aspect the invention relates to a method of treating obesity in a subject with obesity or overweight by introducing a therapeutically effective amount of PYY or a PYY agonist. While "obesity" is generally defined as the condition when the body mass index exceeds 30, for purposes of this description, any subject, including subjects with body mass index less than 30, who needs or wants to reduce body weight, is included in the scope of the term "obesity". The benefit of this method can get actors who have insulin resistance, glucose intolerance, or any form of diabetes (such as diabetes type 1, 2 or gestational diabetes).
In other aspects the hallmark of the invention are methods of decreasing food intake, diabetes treatment and improving lipid profile (including reducing LDL cholesterol and triglyceride levels and/or changing levels of HDL-cholesterol), which includes an introduction to the subject a therapeutically effective amount of PYY or a PYY agonist. In a preferred embodiment, the methods according to the invention are used to treat conditions or disorders which can be alleviated by reducing the availability of nutrients we need in this is subject, and methods include the introduction of a specified subject a therapeutically effective amount of PYY or a PYY agonist. Such conditions and disorders include, without limitation, hypertension, dyslipidemia, cardiovascular disease, eating disorders, insulin resistance, obesity and diabetes mellitus of any kind.
In the methods according to the invention the preferred PYY agonists are agonists with efficiency in one of the analyses described in this description (preferably in the analysis of food intake, gastric emptying, pancreatic secretion gland or weight reduction), which exceeds the efficiency of NPY in the same analysis.
When all the symptoms in the preferred embodiments the preferred PYY agonist is PYY[3-36], and it is preferably introduced perifericheskie at a dose of approximately from 1 μg to 5 mg per day in single or divided doses, or about 0.01 μg/kg to 500 μg/kg per dose, more preferably from about 0.05 mg/kg to 250 mg/kg, most preferably below about 50 μg/kg Dosing within the specified limits will vary depending on the efficiency of each agonist, treatment and easily determined by the person skilled in the art.
In the methods according to this invention PYY and PYY agonists can be entered separately or together with one them or more other compounds and compositions which have a long or short action, reducing nutrient availability, including, but without limiting the above, other compounds and compositions that contain the Amylin or Amylin agonist, cholecystokinin (CCK) or CCK agonist, a leptin (OB protein) or leptin agonist, on the basis or agonist of the basis or GLP-1 or agonist of GLP-1. Suitable Amylin agonists include, for example, [25, 28, 29 Pro-]human Amylin (also known as "pramlintide" and described in U.S. patent No.5686511 and 5998367) and calcitonin salmon. Used CCK preferably is octapeptide CCK (CCK-8). Leptin is discussed, for example, in (Pelleymounter, Cullen et al. Science 269: 540-543, 1995), (Halaas, Gajiwala et al. Science 269: 543-6, 1995) and (Campfield, Smith et al. Science 269: 546-549, 1995). Suitable exendin include the basis 3 and on the basis of 4, and the connection agonists basis include, for example, compounds described in PCT applications WO 99/07404, WO 99/25727 and WO 99/25728.
BRIEF DESCRIPTION of DRAWINGS
Figure 1 is a graph of the activity of ligands of the receptor Y in the analysis of food intake in mice NIH/SW starving during the night.
Figure 2 is a graph of activity of various ligands of the receptor Y analysis emptying of the stomach in rats HSD.
Figure 3 shows the inhibition of the secretion of gastric acid in rats with acute peripheral introduction of PYY[3-36]. Data are expressed as µmol secreted acid/10 minutes
Fig is a chart showing that acute peripheral introduction PYY[3-36] prevent emptying of the gall bladder in mice. The specified action may be reversible with the introduction of CCK-8.
Figure 5 shows an acute dose-dependent effect of subcutaneously injected PYY[3-36], inhibition stimulated CCK-8 exocrine secretion of the pancreas (based analysis of amylase activity) in rats.
Figure 6 shows the reduced weight gain in mice C57B1/6 obese (obesity induced by diet, or DIO) with a continuous peripheral infusion of PYY[3-36] in the period is four weeks.
7 shows the effect of continuous peripheral infusion of PYY[3-36] the degradation of the nutritious substances in mice C57B1/6 obese (obesity induced by diet or DIO) during the period of four weeks.
On Fig shows improved glycemic control, as measured by the change in HbA1c percent during the period of 28 days in diabetic rats with obesity (ZDF) in continuous peripheral infusion of PYY[3-36].
DETAILED description of the INVENTION
Was considered to be generally accepted that endogenous NPY (review in (Schwartz, Woods et al. Nature 404: 661-71, 2000)) and PYY (Morley, J.E., Levine, A.S., Grace, M., and Kneip, J. Brain Res. 341: 200-203, 1985)) through its receptors reinforce the behavior associated with the intake of food. In ways aimed at the treatment of obesity, deism the NGOs sought to exert antagonistic effects on receptors Y, while the claimed methods for the treatment of anorexia were directed to agonists of this family of ligands. However, as described and claimed in this invention, it has been unexpectedly discovered that the peripheral introduction PYY and its agonists have a strong effect, reducing nutrient availability, instead of increasing it, as evidenced by the messages in the patent and scientific literature (see, for example, U.S. patents No.5912227 and 6315203, which stated about the use of agonists of receptors PYY in order to increase the weight gain). Range of action for the inhibition of food intake, slowing of gastric emptying, inhibition of the secretion of gastric acid and inhibition of secretion of pancreatic enzymes useful, providing clinical benefits in metabolic diseases such as diabetes mellitus type 1, type 2 or diabetes mellitus, pregnancy, obesity, and other manifestations of the syndrome of insulin resistance (syndrome X), and any other application to reduce the availability of nutrients.
The use of PYY and PYY agonists to reduce food intake and availability of nutrients in the treatment of disorders such as diabetes and obesity, has not previously been declared. Indeed such use in diabetes impossible to predict due to the lack of sharp action the Oia on the level of glucose in the blood and on the basis of reports of inhibition of insulin secretion. However, this description shows that the group of agonistic ligands and ligands will be suitable for such conditions and related conditions.
Applicant data suggest that the effects perifericheskie entered PYY or PYY[3-36] in reducing food intake and slowing the emptying of the stomach are determined by the interaction with one or more unique classes receptor family of receptors with the Y-box or similar receptors. These data are best explained by interactions with the receptor or receptors, such PYY-preferring (or Y7) receptors, and worse can be explained by the interactions with other known receptors Y, such as Y1-Y6. In table 1 (below) shows the published data on the effectiveness of family ligands PP for known receptors, as well as some unpublished data of the applicant and the rank order of effectiveness of different ligands. The rank order of effectiveness described in this description of the examples does not correspond to any published pharmacological data on receptors and evidence of a new mechanism of action of PYY to reduce the availability of high-calorie substances.
In this invention can use any PYY is whether the PYY agonist. Preferred agonists include PYY peptide agonists, in particular agonistic analogs of PYY, such as PYY[3-36]. Analogues, for example, can be obtained by using conservative amino acid substitutions in the sequence of PYY or her part, and can be tested in the assays described in the examples, or other suitable assays that distinguish the actions of PYY from the action of NPY or PP. Also planned ones agonists.
Range of actions shown PYY, for example, inhibition of food intake, slowing the emptying of the stomach, inhibiting secretion of gastric acid and inhibition of secretion of pancreatic enzymes, etc. that is carried out consistently, limiting the digestion and thereby providing a clinical benefit in such metabolic diseases as diabetes, obesity, cardiovascular disease (atherosclerosis, hypertension, dyslipidemia, and so on), and manifestations of the syndrome of insulin resistance (e.g., syndrome X).
Sequence of peptides person in the family ligands PP, referred to in this description represent the following sequence (indicated using a conventional one-letter code of amino acids):
PP: APLEPVYPGDNATPEQMAQYAADLRRYINMLTRPRY (SEQ ID NO: 1)
PYY: YPIKPEAPGEDASPEELNRYYASLRHYLNLVTRQRY (SEQ ID NO: 2)
PYY[3-36]: IKPEAPGEDASPEELNRYYASLRHYLNLVTRQRY (SEQ ID NO: 3)
NPY: YPSKPDNPGEDAPAEDMARYYSALRHYINLITRQRY (SEQID NO: 4)
These peptides amydraula on-end at physiological expression, but do not require the amidation for the purposes of this invention. These peptides may also have other posttranslational modifications.
PYY and agonists on the basis of the PYY peptide presented in this description, can be obtained by standard methods of recombinant expression or chemical synthesis of peptides, known in this field, for example, by using automated or semi-automated peptide synthesizer.
Solid-phase peptide synthesis can be performed using an automatic peptide synthesizer (e.g., model 430A, Applied Biosystems Inc., Foster City, CA)using the NMP/HOBt (option 1) and the chemistry on the basis of the tBoc or Fmoc (see, Applied Biosystems User's Manual for the ABI 430A Peptide Synthesizer, Version 1.3B July 1, 1988, section 6, pp.49-70, Applied Biosystems, Inc., Foster City, CA) and kierowanie. The peptides can also be synthesized using a synthesizer, Advanced Chem Tech (model MPS 350, Louisville, Kentucky). The peptides can be cleaned RP-HPLC (preparative and analytical)using, for example, the system Waters Delta Prep 3000 preparative column C4, C8 or C18 (10 MK, 2,2×25 cm; Vydac, Hesperia, CA).
Peptide compounds applicable to the invention can also be obtained using recombinant DNA technology using methods known in this field. See, for example, Sambrook et al., Molecular Cloning: A Laboratory Manual, 2 Ed., Cold Spring Harbor (1989). Ones connection applicable in this invention, it is possible to get known in this field means. For example, you can get containing phosphate amino acids and peptides containing these amino acids using methods known in this field. See, for example, Bartlett and Landen, Biorg. Chem. 14: 356-377 (1986).
The above compounds are useful from the point of view of their pharmacological properties. In particular, the compounds according to the invention possess activity as agents that reduce nutrient availability, including a decrease in food intake.
The compositions or pharmaceutical compositions can be entered by any means, including intravenous, intraperitoneal, subcutaneous and intramuscular path, can be administered orally, topically, through the mucous membrane or by inhalation of the lungs. Composition applicable in the invention, it is convenient to put in the form of compositions suitable for parenteral (including intravenous, intramuscular and subcutaneous), nasal or oral administration. In some cases it is convenient to put the PYY or PYY agonist and another agent, reducing food intake, reducing the level of glucose in the blood or altering the lipid composition of the plasma, such as Amylin, an Amylin agonist, CCK or CCK agonist, or a leptin or leptin agonist, or on the basis or agonist of basis, one is notizie or solution for joint injection. In other cases, the preferred may be the introduction of an additional agent separately from the specified PYY or a PYY agonist.
A suitable form of introduction for each individual patient is best determined by the attending physician. Various pharmaceutically acceptable carriers and their compositions are described in standard textbooks on the preparation of the compositions, for example, Remington's Pharmaceutical Sciences, E.W.Martin. See also Wang, Y.J. and Hanson, M.A. "Parenteral Formulations of Proteins and Peptides: Stability and Stabilizers," Journal of Parenteral Science and Technology, Technical Report No.10, Supp. 42: 2S (1988).
Compounds applicable to the invention can be delivered in the form of parenteral compositions, for example, for injection or infusion. Preferably, they are suspended in an aqueous carrier, such as isotonic buffer solution at a pH of from about 3.0 to 8.0, preferably at a pH of about 3.5 to 7.4, 3.5 to 6.0, or 3.5 to about a 5.0. Suitable buffers include buffers: sodium citrate-citric acid and sodium phosphate-phosphoric acid and sodium acetate-acetic acid. Form of long-acting substance or drug slow release of the "depot" can be used to therapeutically effective amount of drug to be delivered into the bloodstream over many hours or days after transdermal injection or delivery.
Because the products according to the invention ablauts is amphoteric, they can be used as free bases, in the form of an acid additive salts or as salts of the metals. Of course the salts should be pharmaceutically acceptable, and they will include metal salts, in particular salts of alkali and alkaline earth metals, for example salts of potassium or sodium. Available with a wide variety of pharmaceutically acceptable acid additive salts. Such products are readily obtained by methods well known to specialists in this field.
For use by physician compositions will be provided in a dosage form containing some amount of PYY or PYY agonist in the presence or without the other active ingredient, such as agent, reducing food intake, reducing the level of glucose in plasma or altering the lipid composition of the plasma. Therapeutically effective amounts of PYY or PYY agonist for use in reducing the availability of nutrients are the amounts that suppresses appetite to the desired level. As will be clear to experts in the field, an effective amount of therapeutic agent will vary depending on many factors, including the age and weight of the patient, the physical condition of the patient, the blood sugar level, the level of weight that you want to receive, and other factors.
Effective utochnyaetsya appetite dose of the compounds will typically be in the range from about 1-30 μg to about 5 mg/day, preferably from about 10-30 μg to 2 mg/day and more preferably from about 5-100 µg to 1 mg/day, most preferably from about 5 μg to 500 μg/day for patients weighing 50 kg in the introduction in the form of single or divided doses. Preferably the dose is from about 0.01 to 100 mg/kg/dose. The exact dose, which must be entered easily determined by the person skilled in the art, and it depends on the effectiveness of specific compounds, as well as the age, weight and condition of the individual. The introduction should start whenever you want suppression availability of nutrients, food consumption, body weight, lowering blood glucose or lipids in the plasma, for example, at the first sign of symptoms or shortly after diagnosis of obesity, diabetes or syndrome of insulin resistance. The introduction can be accomplished in any way, for example, by injection, preferably subcutaneous or intramuscular, oral, nazalnam, transdermal, etc. ways. Doses under certain routes of administration, such as oral administration, should be increased, taking into account the reduced bioavailability, for example, about 5-100 times.
The optimal dosage form and route of administration of the compounds according to this application, the patient is dependent on factors known in this about the region, such as a particular disease or disorder, the desired effect and the type of patient. Although, as a rule, the connection will be used for the treatment of humans, they can also be used to treat similar or identical diseases in other vertebrates, such as other primates, farm animals such as pig, cow and poultry, and sports animals and Pets such as horses, dogs and cats.
Screening additional PYY agonists
Other PYY agonists can be identified using assays of binding to the receptor, described below (for example, in examples 9 and 10) or known in this field, in combination with physiological screening studies described in the examples hereinafter. Potential PYY agonists can compare activity with PYY or PYY[3-36].
Alternatively, once characterized and cloned one or more PYY-preferring (Y7) receptors, it is possible to perform alternative analyses and high-performance screening, as discussed below or known in the field. A Y7 receptor is a receptor with affinity to PYY or PYY[3-36] is higher than their affinity for NPY. Methods of screening compounds that modulate the activity of PYY receptor include the implementation of contact of the test compounds with re what atrami PYY and analyzed for the presence of a complex between the compound and PYY receptors. In such tests, the test ligand is usually mark their territory. After suitable incubation, free ligand is separated from the ligand present in bound form, and the amount of free or not included in the complex label is a measure of the ability of a particular compound to bind to receptor sites PYY. Alternatively, it is possible to measure the bound labeled ligand (e.g., using downregulation associated with membrane receptors Y7).
In another embodiment of the invention applied high-throughput screening of compounds having suitable binding affinity of towards PYY receptors. For example, on a solid substrate to synthesize a large number of different small peptide test compounds. Shall contact the test peptide compounds with PYY receptor and washed. Then coupled receptor PYY register means, well known in this field. Cleaned tested compounds also can directly cover the tablets for use in the above methods of screening drugs. In addition, if the test compounds are proteins, it is possible to use antibody to capture the protein and mobilitat it on a solid medium by any means known in this field.
Other variants of the invention include ispolzovaniem competitive screening in which neutralizing antibodies capable of specific bind the polypeptide according to the invention, compete with a test compound for binding to the polypeptide. Thus, antibodies can be used to identify the presence of any peptide that has one or more antigenic determinants with a PYY agonist. The competitive binding studies with radioactive tagging described in A.H.Lin et al. Antimicrobial Agents and Chemotherapy, 1997, vol.41, no.10. pp.2127-2131, the description of which is included in this description by reference in full.
To facilitate understanding of the present invention in the description of the following examples are included. Of course, the experiments relating to this invention should not be construed as specifically limiting the invention, and believe that such variants of the invention, now known or later developed, which can be within the competence of the person skilled in the art, are included in the scope of the invention, which is represented in the following description and the following claims.
In the following experiments in the various analyses used the representatives of the family ligands PP. Unless otherwise noted, all test peptide compounds were dissolved in saline to a concentration of 1-5 mg/ml without pH. In all cases the courthouse square were transparent to the eye before the introduction.
Example 1: the Activity of ligands of the receptor Y affecting the consumption of food from starving during the night mice NIH/SW
Female mice NIH/Swiss (8-12 weeks of age) were kept in groups in terms of cycle light:dark 12:12 with the beginning of the light period in 06-00. Water and standard pelleted feed for mice was freely available, in addition to these cases. Animals were subjected to starvation and were kept separately, starting around 15-00 hours 1 day before the experiment. On the morning of the experiment (approximately 06-30), all animals were weighed and divided into experimental group, in order to obtain the most similar distribution of mass between groups. In a typical study, n=10 for control group and at least 5 for each group processing.
In point of time = 0 min, all animals were injected intraperitoneal injection of the carrier or compounds in an amount of 5 ml/kg and were immediately given a pre-weighed amount (10-15 g) standard feed. Were administered increasing doses of PYY[3-36] or PYY (from 0.1 μg/kg to 500 μg/kg) and NPY (100 and 500 mg/kg) and a single high dose of NPY[3-36] (100 µg/kg), acetylated at N-end Ac-PYY[22-36] (200 µg/kg) and PP (500 µg/kg), as shown in figure 1. Food was removed and weighed at the point 1 hour to determine the amount of feed consumed (Morley, Flood et al. Am. J. Physiol. 267: R178-R184, 1994).
Food consumption was calculated, visit the observed mass remaining food after one hour from the mass of food, which gave the original in the point = 0. The effect of treatment on food consumption were expressed as % change relative to control.
Significant effects processing was performed using ANOVA (p<0,05). In that case, if there were significant differences, the mean values in the test compared with the average value in the control, using the Dunnet test (Prism v2.01, GraphPad Software Inc., San Diego, CA).
As can be seen in figure 1, with the peripheral PYY administration (intraperitoneal injection) at doses of 10, 100 and 500 mg/kg significantly reduced food consumption, measured during 60 min in female mice NIH/SW starving during the night. These doses of PYY[3-36] had approximately equal efficiency. For PP and NPY tendency to belong to activity was observed at 500 mg/kg But NPY and NPY[3-36] [SEQ ID NO: 5] were inactive at 100 μg/kg of Ac-PYY [22-36] [SEQ ID NO: 6] at 200 mg/kg was inactive. The rank order of effectiveness was as follows: PYY[3-36]PYY≫NPY=NPY[3-36]=PP=Ac-PYY[22-36]. The rank order and, in particular, the lack of influence of NPY does not reflect pharmacology none of the known cloned receptors.
It was reported that PP with the peripheral introduction reduces food intake (Asakawa, Inui et al. Peptides 20: 1445-8, 1999). In addition, reportedly PP with peripheral administration to mice with obesity, decreased food consumption and weight gain(Malaisse-Lagae, Carpentier et al. Experientia 33: 915-7, 1977). It is reported that mouse ob/ob hypersensitive to multiple anoreksigena (Young and Bhavsar. Program and Abstracts, 10thInternational Congress of Endocrinology 419 (poster P2-58), 1996). It was reported that in mice, sverkhekspressiya PP, reduced body weight and food consumption (Ueno, Inui et al. Gastroenterology 117: 1427-32, 1999). The applicants were not able to reduce the consumption of food with PP in the test system shown in figure 1. Research Asakawa with co-workers was an acute study with a single injection, and were not presented data on changes in body weight. Although the study in transgenic mice PP (Ueno et al., Gastroenterology 117: 1427-32, 1999) stated that shown reduced body weight and reduced food consumption at sverkhekspressiya animals, half of the animals died in the perinatal period, which may be a signal of pathophysiology, except for direct explanation of the reduced consumption of milk, leading to starvation. In addition, the system gene expression is not specific to the pancreas, and the peptide is expressed in the brain, which creates interference to any interpretation of the data by overexpression. Ueno and co-authors on the basis of these data they concluded that PP may be involved in the regulation of food intake and body weight, to some extent, through the regulation of GE, but the data obtained in examples 1 and 2 below), show that PP has little or no effect on food consumption and essentially inactive in slowing gastric emptying. It is important that PP only 50% homologous PYY (or NPY)has a different primary localization in tissues (pancreas, but not L-cells of the intestine and neurons) and a clear preference towards the Y4 receptor as compared with Y1 and Y2. NPY, which is 70% homologous PYY, is a potent orexigen under the Central administration. He gives only a moderate decrease in food intake and completely inactive in the analysis of gastric emptying in example 2 (below) with the peripheral introduction.
Example 2: Activity of peptide ligands Y associated with gastric emptying in rats HSD
Male rats HSD, 180-215 g contained in terms of cycle light:dark 12:12 and subjected to starvation for 20 h (overnight). In point of time = 0 min test peptide (PYY[3-36], PYY, Ac-PYY[22-36], NPY, NPY[3-36] or PP) or media in the form of saline were injected with (intraperitoneally) rats, in consciousness (n=6/group). At t=1 min rats, in consciousness, through the oropharyngeal tube-fed 1 ml of sterile water containing 5 mccoury3H-3-O-methylglucose. Blood samples (10 µl) were collected 40 min after feeding and analyzed for radioactivity in counts per minute (CPM) in the plasma. To numb PR is sampling from the tail vein, were injected with 2% lidocaine (0.1 ml) at a distance of 3-4 cm from the tip of the tail (Gedulin, Jodka et al. Gastroenterology 108: A604, 1995).
The effects of test compounds were expressed as changes relative to the control, which was calculated as 100*(1-(average value of the rats in the test/mean control).
The relative activity was defined as significant if p<0.05 using determination by using ANOVA. In that case, if there was a significant difference, the mean values in the test compared with the average value in the control, using the Dunnet test (Prism v2.01, GraphPad Software Inc., San Diego, CA).
As can be seen in figure 2, PYY[3-36] in peripheral administration (intraperitoneal injection) at doses greater than or equal to 10 µg/kg, and significantly dependent on the dose was reduced emptying of the stomach, measured at time point 40 minutes in rats HSD. PYY at 100 and 500 mg/kg was also effective. In contrast, NPY, NPY[3-36] or PP, injected at a dose of 500 mcg/kg, and Ac-PYY[22-36] at a dose of 200 µg/kg were inactive. The order of effectiveness of the test compounds is as follows: PYY[3-36]PYY≫NPY=NPY[3-36]=PP=Ac-PYY[22-36]. The specified performance profile similar to the profile observed in the case of food consumption (figure 1). The lack of effect of NPY does not reflect pharmacology none of the known cloned receptors. It is important that the Ac-PYY[22-36] was inactive in the Wallpaper the analyses, although Balasubramanian and co-authors, U.S. patent 5604203, reported that supperted is a ligand for both receptors PYY intestine, and for the Y2 receptor.
Example 3: Acute peripheral introduction PYY[3-36] inhibits the secretion of gastric acid in rats
Male rats Harlan Sprague Dawley contained in the conditions of cycle light:dark 12:12. All experiments were performed during the light phase of the cycle. Animals are fasted for approximately 20 hours before the experiment but had free access to water until the beginning of the experiment.
Rats (11-16 weeks of age with body mass 291-365 g) surgically installed fistula of stomach (Kato, Martinez et al. Peptides 16: 1257-1262, 1995). Rats, starving during the night, weighed and their gastric fistula was opened and attached to a flexible Tygon tube (3/8×1/16), in which was inserted a piece of tube PE205, which could take place inside the stomach. Through a narrower tube PE205 were injected with saline solution and flowing the fluid were collected from a Tygon tube. To ensure proper flow through the fistula and the emptying of the stomach, the stomach repeatedly washed ˜5 ml of physiological saline at room temperature until the flow became free, and flowing the fluid is clear. The secretion of gastric acid was measured with 10 minute intervals by injection of 5 ml of ideologicheskogo solution followed by injection of 3 ml of air and collecting the flowing fluid. 3 ml of each gastric aspirate was titrated to pH 7.0 with a 0.01 N sodium hydroxide using a pH meter. The required number of reasons, the adjusted relative to the total harvested volume used to calculate the moles of acid in each sample.
After collecting the sample source level and registering the extracted volume, the animals were injected subcutaneous injection of 125 μg/kg of pentagastrin to stimulate gastric secretion. Samples on secretion of gastric acid was collected every 10 minutes. Forty minutes after the injection of pentagastrin animals received subcutaneous injections of 100 µg/kg PYY[3-36] or saline solution, and sampling on the secretion of stomach continued every 10 minutes during 2 hours. Data are expressed in µmol acid, secreted within a 10-minute interval sampling (average ± SEM, n=4/group).
Figure 3 shows that PYY[3-36] in acute introduction by peripheral (intraperitoneal) injection (100 g/kg) inhibited stimulated pentagastrin secretion of gastric acid in rats. ED50for the above effect was ˜20 µg/kg
Example 4: Acute peripheral introduction PYY[3-36] prevent emptying of the gall bladder in mice, reversible CCK-8
Mice were kept in the room in terms of cycle light:dark 12:12 with free access to water and feed and up to the beginning of the experiment. At t=0, mice received subcutaneous injection of 1, 10, 100 or 1000 mg/kg PYY[3-36], 1 or 10 µg/kg CCK-8, both of the substance or saline (treatment and n/a group, as shown in figure 4). Thirty minutes later the animals were anestesiology and took them intact gall bladders and weighed.
Data are expressed as the body mass in mg Activity was defined as the change compared with the average value in the control group. Statistical significance was defined as p<0,05 using ANOVA and/or Dunnet test.
As can be seen in figure 4, PYY[3-36] in the introduction with acute peripheral injection at doses greater than or equal to 10 µg/kg, prevented the emptying of the gall bladder in mice. Specified inhibition emptying had ED50˜ 31 mg/kg and it can be canceled by using CCK-8, even at the highest tested doses of PYY[3-36].
Example 5: Acute peripheral introduction PYY[3-36] inhibits stimulated CCK-8 exocrine secretion of the pancreas (amylase) in rats
Male rats Harlan Sprague Dawley contained in the conditions of cycle light:dark 12:12. All experiments were performed during the light phase of the cycle. Animals are fasted for approximately 20 hours before the experiment, but had free access to water until the beginning of the experiment.
Rats were anestesiologi 5% halothane gas, in remakelyrics operations supported anesthesia with 2% halothane gas and then with 1% halothane gas. Performed the tracheotomy and the introduction of the cannula into the right femoral artery and the body temperature was controlled using a thermostat, which was switched on and off a heated operating table. System for intravascular infusion into the femoral artery is used for sampling blood was perfesional heparinised saline (2U/ml) and connected to the pressure sensor for registration of blood pressure. Through the incision in the midline in the United bile duct - pancreatic duct was inserted two polyethylene cannula at a point approximately 0.5 cm above the place where the duct enters the pancreas. The first cannula was inserted towards the liver to collect bile. The other end of this cannula was placed into the duodenum through a small incision in the duodenum. Thus, the bile flowed directly from the liver into the small intestine, while there was a full bypass of the pancreas. The second polyethylene cannula inserted into the joint bile duct - pancreatic duct next to the first, was sent to the pancreas to collect pancreatic juice. On the pancreatic duct has put a ligature upon its entry into the duodenum, forcing the secretory pancreatic juice to drain into the Kania is Yu for collection.
Pancreatic juice was collected in 15-minute intervals from t=-15 to +60 minutes, the Volume of pancreatic juice (measured by weight) and the amylase activity was determined in the aliquot taken every 15 minutes (Taniguchi, Yazaki et al. Eur. J. Pharmacol. 312: 227-33, 1996). Pancreatic juice before analysis were diluted 1:2000. The secretion of the enzyme was expressed in units per 15 min, obtained by multiplying the activity on collected amount (Taniguchi, H., Yazaki, N., Yomota, E., Shikano, T., Endo, T., and Nagasaki, M. Eur. J. Pharmacol. 312: 227-33, 1996).
Paired statistical analyses were performed using t-student test; multiple comparisons with a control, using the Dunnet test; overall effects were tested using one way ANOVA. The results are presented as average ± standard error of the mean. As the level of significance using P<0,05.
Figure 5 shows that PYY[3-36] in the introduction with acute peripheral (subcutaneous) injection at a dose of 30 mg/kg blocked stimulated CCK-8 secretion of the pancreas in rats, which was measured by the activity of amylase in pancreatic juice. In the absence of CCK-8, PYY[3-36] in a dose of 300 mg/kg had no effect on the initial amylase activity when compared with controls, which were injected with saline.
Example 6: Continuous peripheral infusion of PYY[3-36] reduce weight gain in mice C7B1/6 obese (DIO)
Male mice C57B1/6 (4 weeks of age at start of study) were fed food with high fat (HF; 58% kcal in food as fat) or low fat (LF; 11% kcal in food as fat). After 7 weeks of feeding each mouse was implanted osmotic pump (Alzet No. 2004), which was continuously delivered to the specified figure 6 dose of PYY[3-36] (30, 100, 300 or 1000 mg/kg/day) for 4 weeks. Weekly measured body weight and food consumption (Surwit, Feinglos et al. Metabolism-Clinical and Experimental 44: 645-651, 1995).
The effect of the test compounds were expressed as average ± sd change in grams of the initial mass, at least for the 14 mice per group treatment (p<0.05 in ANOVA, Dunnet test (Prism v2.01, GraphPad Software Inc., San Diego, CA).
Figure 6 shows that PYY[3-36] in the introduction through continuous peripheral infusion caused a dose-related reduction of weight gain in mice with obesity induced by diet (DIO). Effects were significant at the dose of 300 mg/kg/day during the first 3 weeks and all time points in the case of a dose of 1000 mg/kg/day.
Example 7: Continuous peripheral infusion of PYY[3-36] reduces the effective caloric content of food in mice C57B1/6 obese (DIO)
Male mice C57B1/6 (4 weeks of age at start of study) were fed food with high fat (HF; 58% kcal in food in VI is e fat) or low fat (LF; 11% kcal in food as fat). After 7 weeks of feeding each mouse was implanted osmotic pump (Alzet No. 2004), which was continuously delivered to the specified figure 6 dose of PYY[3-36] (30, 100, 300 or 1000 mg/kg/day) for 4 weeks. Weekly measured body weight and food consumption (Surwit, Feinglos et al. Metabolism-Clinical and Experimental 44: 645-651, 1995).
The effect of the test compounds were expressed as changes of body weight in grams of the initial mass relative to the consumed calories. Consumed kcal was calculated by multiplying the mass of food consumed (g) the density of calories (kcal/d), specified by the manufacturer. Please note that these data were obtained for animals used in example 3.
Activity was defined as the change in the average ± sd n average of at least 14 mice/group. Significance was defined as p<0,05 in ANOVA or Dunnet test.
7 shows that PYY[3-36] in the sub-chronic introduction through continuous peripheral infusion caused a dose-related decrease in the efficiency of caloric substances (measured as weight gain/kcal consumed) in mice with obesity induced by diet (DIO). Effects were significant at all time points in the case of a dose of 1000 mg/kg/day and at some time points the dose of 300 mg/kg/day.
Example 8: Continuous accessories the historic infusion of PYY[3-36] within 28 days improves glycemic control in diabetic rats with obesity (ZDF)
Diabetic Zucker rats (ZDF) (7-week old) were kept in the room in terms of cycle light:dark 12:12 and they had free access to feed for rodents with a high content of fat and water. After acclimatization for 1 week, took blood samples and animals, sorted by baseline HbA1c levels, to ensure similar within each group processing.
Animals implanted osmotic pumps, which continuously perifericheskie brought these on Fig doses of PYY[3-36] or saline for 28 days. HbA1c was measured at weekly intervals. Levels of HbA1c (%) was postponed on a graph against time (Brown, Henke et al. Diabetes 48: 1415-24, 1999).
As shown in Fig, PYY[3-36] in the continuous introduction by peripheral infusion diabetic rats Zucker obese (ZDF) led to a dependent dose-dependent improvement in long-term regulation of glucose levels in the blood, as measured by HbA1c. The degree of improvement in glycemic control was increased during the treatment period and was significant at all doses of PYY[3-36] on the 28th day.
The examples above are direct evidence that the PYY agonists applicable to reduce the availability of nutritious substances and can be used as therapeutic agents for treating conditions in which it is useful red eye reduction is the availability of nutritious substances, such as obesity and type 2 diabetes. In addition, the examples show that the effects of PYY agonists to reduce the availability of high-calorie substances can occur via several mechanisms and are the basis for identifying PYY agonist. As it was reported that PYY and NPY are equivalent and equally effective in all studies in the analysis of receptors Y1 and Y2, the data in examples 1 and 2 above indicate that the effects of PYY and agonists reduce food intake (figure 1) and the slow emptying of the stomach (figure 2) is not mediated by receptors Y1 or Y2. The obtained data show that the effects of PYY on food consumption and the emptying of the stomach cannot be compared with reported effects on receptors Y1 and Y2, as NPY showed negligible activity or showed no activity in these assays.
Figure 1 illustrates one aspect of the invention. It is known that the Central administration of the agonist PYY or NPY increases food intake (Clark, Kalra et al. Endocrinology 115: 427-9, 1984; Clark, Sahu et al. Regul. Pept. 17: 31-9, 1987). Unexpectedly, applicants have found that the peripheral introduction PYY or PYY[3-36] effectively reduces food intake. The applicants have documented that PYY[3-36] is reduced food intake in long-term studies in other models in rodents, including mice ob/ob and rat fa/fa, the results are not presented in the data is m description. When the peripheral introduction of other members of the family of PP have little influence or no influence on food consumption. The order of effectiveness and, in particular, the lack of influence of NPY does not reflect the pharmacological properties none of the known cloned receptors Y. the Unique pharmacology of PYY agonists further proved on the basis of their effective influence due to the slowing of gastric emptying, compared with other members of the family of PP, which is not active in this assay (see, for example, the data in figure 2).
Description PYY-agonist PYY[3-36] suggests additional mechanisms that can reduce the availability of high-calorie substances. Mechanisms include reduced secretion of gastric acid (figure 3), reduced exocrine secretion of the pancreas (figure 5) and slow emptying of the gall bladder (figure 4). Without binding the invention to any theory, the applicants put forward the hypothesis that the specified full range of influences on food consumption and the functioning of the gastrointestinal tract contributes to the applicability of PYY agonists to reduce the availability of nutritious substances. For example, reportedly PYY and PYY[3-36] inhibited vagal-stimulated secretion of gastric acid in rabbits (Lloyd, Grandt et al. Am. J. Physiol. 270: G123-G127, 1996). PYY also inhibi the oval stimulated pentagastrin secretion of gastric acid in humans (Adrian, Ferri et al. Gastroenterology 89: 1070-7, 1985) and rats (Greeley, Guo et al. Proc. Soc. Exp. Biol. Med. 189: 325-8, 1988) and induced CRF secretion of gastric acid in rats (Gue, Junien et al. Br. J. Pharmacol. 118: 237-42, 1996). It was also reported that PYY (Yoshinaga, Mochizuki et al. Am. J. Physiol. 263: G695-701, 1992), (Guan, Maouyo, et al. Endocrinology 128: 911-6, 1991), (Pappas, Debas et al. Am. J. Physiol. 248: G118-23, 1985) and PYY[3-36] (Deng, Guarita et al. Dig. Dis. Sci. 46: 156-65, 2001) inhibit the secretion of pancreatic enzymes. Recently it was reported that people in normal PPY decreases dependent on the brain, but CCK-dependent phase of the emptying of the gallbladder (Hoentjen, Hopman et al. Scand. J. Gastroenterol. 35: 166-71, 2000).
The applicants suggested that PYY agonist identified through this study of the mechanisms to reduce body weight. Applicants in several models of obesity in rodents have established that peripheral introduction PYY[3-36] causes a dose-dependent decrease in body mass and/or velocity of weight gain. In this description, the applicants have shown effect in the model in mice with obesity induced by diet (DIO) (6).
In addition, as summarized in figure 1, peripheral administration of PYY agonist reduces food intake. On the basis of the calculation of weight gain on kcal consumed in the study of DIO mice, it is clear that peripheral administration of PYY agonist reduces the efficiency with which calories are converted body weight (Fig.7). Accordingly, the p is emery confirm the effect of PYY agonists, expressed in reduced weight gain due to reduced availability of nutritious substances.
In particular, PYY and PYY agonists applicable for the treatment of diseases, which will be useful to reduce the availability of high-calorie substances, such as obesity, type 2 diabetes and cardiovascular disease. Applicants investigated the antidiabetic activity of PYY[3-36] in rodent diabetic and obese ZDF rats. Peripheral administration of PYY agonist causes a significant, strong, and a dose-related improvement in glycemic control, which was measured by the levels of hemoglobin A1C (Fig). Although it is not presented in the example, food consumption was also decreased with the introduction of PYY[3-36].
Example 9: analysis of the area postrema.
It was reported that PYY with the peripheral introduction activates neurons in the area postrema (Bonaz, Taylor et al. Neurosci. Lett. 163: 77-80, 1993). Evaluation of agonistic activity of PYY potential compounds according to the invention can be carried out using the analysis area postrema, as set forth below, in combination with analysis of the impact of PYY, such as analyses in examples 1 and 2.
Preparation of membranes
In this analysis, the membrane area postrema were prepared from tissue obtained by dissection of the brain stem, pigs or cows. Preparation of membrane area postrema started short-term (4-10 seconds) by homogenization of tissues using a tissue homogenizer transmitter station (Brinkmn Instruments, NY) at temperatures obtained by cooling with ice in a buffer solution such as phosphate-saline buffer (138 mm NaCl, and 8.1 mm Na2PO4, 2.5 mm KCl, 1.2 mm KH2PO4, 0.9 mm CaCl2, 0.5 mm MgCl2, pH 7,4). After the destruction of the tissue removal of large particles and residues was performed by centrifugation (200×g, 5 minutes, 4°C) and the fraction nadeshiko kept on ice. Membranes were isolated from fraction nadeshiko high-speed centrifugation (at least 40000×g, at least for 10 minutes, 4°C). Typically, the membrane was washed at least twice by repeated homogenization in fresh buffer and re-centrifuged in order to remove endogenous impurities. Washed membranes suspended in buffer containing inhibitors of proteolytic enzymes, such as phenylmethylsulfonyl (PMSF) or bacitracin. The buffer can be added in amounts sufficient to bring the final concentration of tissue to a level suitable for the particular method used screening.
In one embodiment, the incubation mixture for a method of screening is prepared as follows. In a glass or plastic test tube add a small amount of buffer mixture ("HBBM"), consisting of a buffer, such as HEPES, containing a protease inhibitor, such as bacitracin or PMSF, not aderrasi proteases serum albumin (preferably fraction V BSA, not containing proteases) and optional Mg2+or Ca2+-salt and EDTA. To the buffer mixture is added a small volume of buffer containing unlabeled molecules that need to be tested against agonistic activity at concentrations from about 10-11up to 10-6M Control tube containing only one buffer. To the resulting mixture add the number of labeled ligand preparation area postrema (in this case PYY) in the buffer to obtain a final concentration of about 10 to 100 PM. Due to the achieved high specific activity and ease of chemical labelling for labelling ligands area postrema preferred125I. Ligands can be isolated from human tissue, animal tissue or to obtain chemical, synthetic or recombinant methods. Labeled ligands preparation area postrema dissolved in sterile water containing fraction V BSA without proteases, and stored frozen until use.
Reactions begin, for example, the addition of membranes in each incubation tube. The required number of membrane protein per tube varies so that the amount of labeled ligand bound by membranes in the analysis, was less than 10% of the total concentration of ligand in the assay (typically about 100 μg).
The reaction mixture is incubated for a period of time and when the temperature, sufficient to achieve stationary conditions over a period of time. It is implied that used in this description, the term "stationary state" includes the total of all reactions and processes that affect the net amount of bound hormone. The term may or may not be synonymous with "balance". Usually the tubes are incubated for approximately 60 minutes at room temperature.
In the case when the use of the membrane, they emit after binding to determine the amount of bound labeled ligand after competition between labeled and unlabeled ligands. Membrane convenient to collect by filtration using a vacuum collector cells Brandel (Brandel Instruments, Gaithersburg, Maryland, model M-24) through the filters of the optical fiber (e.g., GF/B, Whatman), which are pre-soaked with a reagent to reduce non-specific binding (NSB). It is preferable to soak the filters in 5 hours to about 0.3% of polyethylenimine. The specialist in this area other known device for collecting membranes, such as the filter unit, Millipore (model 1225) or filter block Sandbeck (Bennett, J.P., Neurotransmitter Receptor Binding, H.I. Yamura, et al.; Raven, New York 1978, Pages 57-90)collecting filters and reagents that reduce NSB, which can be used in the analysis of binding receptors. Not orestano before, and immediately after filtration, the filters are washed with large volumes (ml) of ice-cold buffer to remove contaminants, such as unbound labeled ligand. Filters extract and quantify associated with membranes labeled ligand. In the case when the label is125I, radioactivity can be estimated as the count of gamma-rays. In that case, when using chemiluminescent reporter molecule (e.g., AMPPD, Tropix, Inc., Bedford, MA), quantify formed by light radiation using a luminometer. You can also use an enzyme and a fluorescent label.
Instead of filtering membranes after incubation can be distinguished by centrifugation (for example, in a centrifuge Beckman-2-21-M with cooling at 21000 rpm or microcentrifuge Beckman 12 or Eppendorf), washed with ice-cold buffer, and then subjected account in this form or after solubilization of membranes by detergent or alkali.
Analysis of the saturation curve of Scatchard for results of the binding, which bound/free (B/F) labeled ligand is applied as a function of the associated quantity, perform standard ways. See, for example, (Scatchard. Ann. NY Acad. Sci. 51: 660, 1949).
Curves of competition on which the associated number (B) is applied as a function of log concentration of the ligand, can be analyzed using computer the EPA, for example, using the analysis using non-linear regression in accordance with the logistic equation, containing 4 parameters (Prism Program; GraphPAD Software, San Diego, California), or the program ALLFIT (version 2.7 (NIH, Bethesda, MD MD 20892 work)) (Munson and Rodbard. Anal. Biochem. 107: 220-39, 1980; de Lean, A., Munson, P.J. et al. 1988).
To determine the binding constants can be obtained curves saturation Scatchard and analyze according to modification of the method of Scatchard as described Bylund, D.B., et al., "Methods for Receptor Binding" in H.I.Yamamura et al., eds., Methods in Neurotransmitter Analysis, Raven Press, New York, 1990 pp.1-35.
In order to experimentally obtain specific values for binding, use the wide limits of the studied concentrations of labeled ligand (usually 1-150 PM)to get the total binding, and duplicating tubes analyze again in the presence of very high concentrations, such as 100 nm unlabeled ligand, to obtain a non-specific binding (NSM). The last value is subtracted from each value of the total binding, receiving specific binding at each concentration of labeled ligand.
Example 10: analysis of the binding of the receptor Y
Evaluation of agonistic activity towards PYY potential compounds according to the invention can be accomplished by examining their interaction with any of the known Y receptors, such as Y1-Y6, or one or more classes unique cocktail recipes. is s, similar to PYY-preferring receptors (such as Y7), expressed in cells, in combination with analysis of the impact of PYY, such as tests of examples 1 and 2. These cells may endogenously Express interest Y receptor (such as cells SK-N-MC, which Express receptors Y1, or cells SK-N-BE2, which Express receptors Y2) or they can be other cells (such as cells COS-7 or HEK293)that transferout clone of interest receptor Y. as an example, use binding to cells SK-N-BE2.
Cells SK-N-BE2 is grown, for example, on cups with a diameter of 150 mm in the medium for tissue culture with additives (modified Dulbecco Wednesday Needle with 10% fetal calf serum, 4 mm glutamine, 100 units/ml penicillin and 100 μg/ml streptomycin) at 37°C in a humid atmosphere with 5% CO2. The original Cup trypsinized and cells split 1:6 every 3-4 days.
Preparation of membranes
Cells scraped from the cups into a small volume of buffer solution such as phosphate-saline buffer (138 mm NaCl, and 8.1 mm Na2PO4, 2.5 mm KCl, 1.2 mm KH2PO4, 0.9 mm CaCl2, 0.5 mm MgCl2pH 7,4), or trypsinized, washed and resuspended in buffer solution. Preparation of membranes begin briefly (10 seconds) by homogenization of the cells, using, for is example, a tissue homogenizer transmitter station (Brinkman Instruments, NY), at temperatures obtained by cooling on ice. Next, the membrane is prepared by centrifugation as described above in example 9. Binding assays, recording and analysis of data are the same as described in example 9.
Also shown and described in this description of the modifications to a person skilled in this field will be apparent various modifications of the invention, which are included in the scope of the following claims.
1. The method of reducing the availability of nutrients, including peripheral introduction to the subject of the number of peptide YY (PYY or PYY agonist, therapeutically effective to reduce the availability of nutrients.
2. The method according to claim 1, wherein the subject is human.
3. The method according to claim 1, characterized in that the reduced availability of nutrients includes the reduction of the effective caloric content through the peripheral introduction to the subject of the amount of PYY or a PYY agonist, is effective to reduce the effective caloric content.
4. The method according to claim 1, characterized in that the reduced availability of nutrients includes suppression of appetite by peripheral injection amount of PYY or PYY agonist to suppress appetite for consumption of food with low-fat or simultaneously with high and low the fat content.
5. The method according to any one of claims 1 to 4, wherein the PYY agonist has a higher affinity for the Y2 receptor in SK-N-BE2 cells compared with the Y1 receptor in SK-N-MC cells and injected perifericheskie entity in the amount therapeutically effective to reduce the availability of nutrients.
6. The method according to claim 1, characterized in that the reduced availability of nutrients includes the reduction of the effective caloric content or appetite suppression by peripheral introduction to the subject agonist PYY, PYY agonist has a higher affinity for the Y2 receptor in SK-N-BE2 cells compared with the Y1 receptor in SK-N-MC cells in the amount therapeutically effective to reduce caloric efficiency or suppress appetite.
7. The method according to claim 5 or 6, wherein the PYY agonist has a higher affinity to the Y5 receptor as compared with the Y1 receptor.
8. The method according to any one of claims 1 to 7, wherein the PYY or PYY agonist is administered in an amount of about 1 μg to 5 mg, or from about 5 to 100 μg per day in single or divided doses.
9. The method according to any one of claims 1 to 7, wherein the PYY or PYY agonist is administered in an amount of about from 0.1 to 10 µg/kg per day in single or divided doses.
10. The method according to claim 1, further comprising suppressing appetite by parenteral introduction the Oia subject number of PYY or a PYY agonist, effective for suppressing appetite, but such amount is from about 0.1 to 10 mg/kg per day in single or divided doses.
11. The method according to claim 1, further comprising suppressing appetite by injecting the subject of the amount of PYY or a PYY agonist, is effective to suppress appetite, and the specified amount is about 5 to 100 μg per day in single or divided doses.
12. The method according to any one of claims 1 to 11, wherein the PYY agonist is PYY [3-36].
13. The method according to any one of claims 1 to 12, wherein the PYY agonist, at least one of the tests of the consumption of food or empty stomach is more effective than neuropeptide Y (NPY).
14. The method according to any one of claims 1 to 13, further including the introduction of GLP-1, the basis, Amylin, their agonists or any combination thereof.
15. The method according to any one of claims 1 to 14, wherein the PYY or PYY agonist is administered parenterally, intravenously, administered intraperitoneally, intramuscularly, subcutaneously, topically, nasal, or by pulmonary inhalation.
16. The method according to claim 1, characterized in that the reduced availability of nutrients leads to weight reduction.
17. The method according to claim 1, characterized in that the reduced availability of nutrients leads to a decrease in weight gain.
18. SP is a way to reduce food intake or appetite suppressant, including peripheral introduction to the subject of the number of peptide YY (RUU) or PYY agonist, therapeutically effective to reduce food intake or appetite suppressant.
19. The method according to p, wherein the subject is human.
20. The method according to p, characterized in that the reduction in food intake or appetite suppressant includes the reduction of the effective caloric content or availability of nutrients through the peripheral introduction to the subject of the amount of PYY or a PYY agonist, is effective to reduce the effective caloric content or availability of nutrients.
21. The method according to p, characterized in that the reduced intake of food with low-fat or simultaneously with high and low fat content.
22. The method according to p, characterized in that it suppressed the appetite for food with low fat or simultaneously with high and low fat content.
23. The method according to any one of p-22, wherein the PYY agonist has a higher affinity for the Y2 receptor in SK-N-BE2 cells compared with the Y1 receptor in SK-N-MC cells and injected perifericheskie entity in the amount therapeutically effective to reduce food intake or appetite suppressant.
24. The method according to any of PP-22, wherein the PYY agonist has a higher affinity for the Y2 receptor in SK-N-BE2 is ledah compared with the Y1 receptor in SK-N-MC cells and is entered in the quantity therapeutically effective to reduce caloric efficiency or availability of nutrients.
25. The method according to item 23 or 24, wherein the PYY agonist has a higher affinity to the Y5 receptor as compared with the Y1 receptor.
26. The method according to any of PP-25, wherein the PYY or PYY agonist is administered in an amount of about 1 μg to 5 mg, or from about 5 to 100 μg per day in single or divided doses.
27. The method according to any of PP-25, wherein the PYY or PYY agonist is administered in an amount of about from 0.1 to 10 µg/kg per day in single or divided doses.
28. The method according to p, further comprising reducing caloric efficiency or availability of nutrients by injecting the subject of the amount of PYY or a PYY agonist, is effective for reducing caloric efficiency or availability of nutrients, with the specified amount is from about 0.1 to 10 mg/kg per day in single or divided doses.
29. The method according to p, further comprising reducing caloric efficiency or availability of nutrients by injecting the subject of the amount of PYY or a PYY agonist, is effective for reducing caloric efficiency or availability of nutrients, with the specified quantity is STV is about 5 to 100 μg per day in single or divided doses.
30. The method according to any of PP-29, wherein the PYY agonist is PYY [3-36].
31. The method according to any of PP-30, wherein the PYY agonist, at least one of the tests of the consumption of food or empty stomach is more effective than NPY.
32. The method according to any of p-31, further including the introduction of GLP-1, the basis, Amylin, their agonists or any combination thereof.
33. The method according to any of p-32, wherein the PYY or PYY agonist is administered parenterally, intravenously, administered intraperitoneally, intramuscularly, subcutaneously, topically, nasal, or by pulmonary inhalation.
34. The method according to p, characterized in that the reduction in food intake or suppress appetite leads to weight loss.
35. The method according to p, characterized in that the reduction in food intake or appetite suppressant decreases with increasing body mass.
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.
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
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 :
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 :
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  or  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  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
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.
FIELD: medicine, oncology.
SUBSTANCE: invention relates to a method for treatment of malignant tumors. Method involves administration in a patient the chemotherapeutically active dose of antitumor platinum compound, foe example, cisplatin or carboplatin and erythropoietin or erythropoietin-like substance wherein the latter is administrated before administration of platinum compound or simultaneously with its. This method provides attaining the synergistic antitumor effect.
EFFECT: improved and valuable medicinal effect.
8 cl, 2 tbl, 2 dwg, 2 ex
FIELD: medicine, endocrinology, biochemistry, peptides.
SUBSTANCE: invention represents new peptides that act in vivo as stimulators of insulin secretion by pancreas beta-cells in glucose-dependent regimen. Such peptides as enhancers of insulin secretion stimulate insulin secretion by insula cells in rats in vitro and in vivo. Proposed peptides represent a new way for treatment of patients with reduced secretion of endogenous insulin, in particular, for treatment of diabetes mellitus type 2. In particular, invention represents polypeptide taken among the specific group VIP/PACAP-related polypeptides or their functional equivalents. Also, invention claims method for preparing both recombinant and synthetic peptides. The advantage of invention involves new peptides that can be used as stimulators of insulin secretion.
EFFECT: improved and valuable medicinal properties of peptides.
47 cl, 4 tbl, 10 dwg, 18 ex
FIELD: medicine, therapy, gastroenterology, pharmacy.
SUBSTANCE: method involves oral intake of solid medicinal formulation at vertical position of patient and change of medicinal formulation position into stomach is carried out in each 5 min, not rare. Change of medicinal formulation position is carried out by pressing on epigastrium region by hand or by cyclic change of position of patient body from its vertical position to horizontal position and back. Method provides enhancing safety in enteral using a solid medicinal formulation due to diminishing its ulcerogenic effect. Invention can be used in enteral using solid medicinal formulations.
EFFECT: improved method for diminishing ulcerogenic effect.
FIELD: medicine, endocrinology, pharmacy.
SUBSTANCE: invention relates to medicinal agents, in particular, to the hormonal pharmaceutical composition. Invention proposes new pharmaceutical compositions and a method for preparing such compositions formed by the estrogen-gestagen combination with a single gestagen compound in mixture with one or some nontoxic, inert pharmaceutically acceptable carriers designated for oral administration. Also, invention relates to the estrogen-gestagen mixture wherein estrogenic component and gestagenic component are used by the combined method. Proposed composition is designated for treatment of estrogenic insufficiency, prophylaxis of osteoporosis and cardiovascular diseases in women in the menopause period. Invention provides the development of the new estrogen-gestagen combination showing activity in the oral route of applying and administrated by the combined method.
EFFECT: improved and valuable medicinal properties of composition.
12 cl, 8 tbl, 3 ex
FIELD: medicine, otorhinolaryngology.
SUBSTANCE: one should treat deformation in laryngeal and tracheal lumen due to excessive growth of granulation tissue in the sites of their lesions. One should introduce hormonal preparations, moreover, one should apply Diprosan as a hormonal preparation injected once intramucosally at 0.1 ml/sq. cm of granulation tissue, but not more than 0.3 ml.
EFFECT: higher efficiency of therapy.
3 ex, 1 tbl
FIELD: medicine, otorhinolaryngology.
SUBSTANCE: one should treat deformation in laryngeal and tracheal lumen due to excessive growth of granulation tissue in the sites of their lesions. One should introduce hormonal preparations, moreover, one should apply Diprosan as a hormonal preparation injected once intramucosally at 0.1 ml/sq. cm of granulation tissue, but not more than 0.3 ml.
EFFECT: higher efficiency of therapy.
3 ex, 1 tbl