JP2008195625A - G protein-coupled receptor inhibitor and medicine - Google Patents

Abstract

<P>PROBLEM TO BE SOLVED: To provide a substance and a medicine inhibiting the activation of a G protein-coupled receptor. <P>SOLUTION: The increase of intracellular Ca<SP>2+</SP>concentration induced by a GPR120-containing cell is remarkably suppressed to suppress (inhibit) the activation of the GPR120-containing cell by the addition of a thiazolidine compound such as rosiglitazone, troglitazone and ciglitazone. Taking advantage of the action, the invention provides a G protein-coupled receptor inhibitor containing a compound selected from thiazolidine compounds as an active component, and similarly provides a medicine containing a G protein-coupled receptor inhibitor as an active component. <P>COPYRIGHT: (C)2008,JPO&INPIT

Description

  The present invention relates to a G protein-coupled receptor inhibitor, and a medicament for treating digestive organs, psychiatric disorders, hypoglycemia, lung diseases and the like.

  Physiologically active substances such as hormones and neurotransmitters can perform their functions through specific receptor proteins present in cell membranes. In general, a receptor protein is used for signal transduction within a cell through activation of a conjugated guanidine triphosphate (GTP) -binding protein (GTP-bindig protein) (hereinafter sometimes abbreviated as G protein). Are collectively referred to as G protein-coupled receptors (proteins). The G protein receptor is also collectively referred to as a seven-transmembrane receptor protein because of its structure having seven transmembrane domains.

As the G protein-coupled receptor, for example, GPR120 existing in the intestinal tract, lungs, brain, fat cells, pituitary gland, etc. is known. Those that promote the activation of intracellular physiologically active substances such as G protein-coupled receptors are called agonists, and those that inhibit are called antagonists. For example, in Patent Document 1 and Non-Patent Document 1, when a ligand that activates GPR120 is allowed to act on cells possessing GPR120, intracellular Ca ²⁺ increases, and cholecysts from enterohormone-secreting cells having GPR120 exist. It is disclosed that peptide hormones such as kinin (CCK) and GLP-1 (glucagon-like polypeptide-1) are released.

In view of this, it has been proposed to use an agonist or an antagonist as a therapeutic agent for various diseases. For example, Patent Document 1 describes that administration of a substance that promotes the release of CCK can improve symptoms such as eating disorders and associated diseases. Non-Patent Document 1 describes that administration of a substance that increases intracellular Ca ²⁺ concentration can increase GLP-1 and insulin secretion and treat diabetes. Patent Document 1 proposes that a substance that promotes the release of CCK from GPR120-bearing cells is used as a cooperative promoter of digestive activity and a therapeutic agent for digestive activity disorder. Non-patent document 3 discloses that such a ligand is used as an anti-obesity therapeutic agent for appetite suppression and a therapeutic agent for bulimia. Non-patent document 4 discloses such a ligand, A preventive and therapeutic agent for diabetes by promoting differentiation and proliferation of pancreatic β cells. In particular, it has been proposed to be used as a therapeutic effect promoter at the time of β-cell or its precursor cell transplantation treatment. Non-Patent Document 5 proposes that such a ligand is used as a therapeutic agent for diseases caused by neuronal cell damage such as nerve cell plasticity, nerve transplantation by viability maintenance action, treatment accelerator at the time of nerve junction, or Alzheimer's disease. Has been. Non-Patent Document 6 discloses that such a ligand is used as a therapeutic agent for intestinal motility abnormality during enteritis due to normalization of intestinal motility. Non-Patent Document 7 discloses that such a ligand is used as a therapeutic agent for promoting the effect of an analgesic agent such as morphine due to a decrease in CCK concentration, or for a disorder caused by anxiety or stress. Non-Patent Document 8 discloses that such a ligand is used as a therapeutic agent for lung diseases such as COPD (chronic obstructive pulmonary disease) by promoting surfactant secretion in the lung.

Furthermore, Patent Document 2 shows that adipocytes and pituitary glands possess GPR120 and have a lipolysis inhibitory action and an ACTH (adrenocorticotropic hormone) secretion inhibitory action, respectively. However, a GPR120 antagonist can be a therapeutic agent for obesity by inhibiting fat accumulation in adipocytes or a therapeutic agent for ACTH secretion disorder and pituitary dysfunction.
JP 2005-15358 A WO 2004/065960 A1 Nature Medicine, 11 (1), 90-94, 2005 Nutrition 2001; 17 (3): 230-5 Physiol Behav. 2004; 83 (4): 617-21 Diobetology, 2005; 48 (9): 1700-13 Curr Drug Target CNS Neurol Disord 2002 Drug 2003; 63 (12): 1785-97 Neurosci Biobehav Rev. 2005: 29 (8): 1361-73 Endocrinology. 1998; 139 (5): 2363-8

    However, with regard to substances that suppress G protein-coupled receptors, that is, antagonists, their adverse effects have been pointed out, but there is little thought to actively use them.

  Thus, as a result of the present inventors conducting research on the usage, we have obtained knowledge about the use of antagonists that have not been established as therapeutic agents.

  An object of the present invention is to establish a technique for beneficially utilizing a substance that suppresses a G protein-coupled receptor.

  The G protein-coupled receptor inhibitor of the present invention is an antagonist of a G protein-coupled receptor (particularly GPR120) containing a thiazolidine compound as an active ingredient. Here, GPR120 is one of G protein-coupled receptors (hereinafter sometimes abbreviated as GPCR), and is a polypeptide represented by Sequence Listings 1-3.

  This makes it possible to use an antagonist of a G protein-coupled receptor, which has hitherto been taken up only with the harmful effects of lifestyle-related diseases, for various purposes. That is, a thiazolidine-based compound can be used to alleviate the action of a G protein-coupled receptor. In addition, the thiazolidine-based compound can be used as an administration agent for causing a cell or an organism to have a physiological state necessary for various experiments.

（式中、Ｒ１は置換されてもよい低級アルキル基を示す）で表される化合物である。 The thiazolidine compound of the present invention has the following general formula (1)

(Wherein R1 represents an optionally substituted lower alkyl group).

  Examples of the lower alkyl group of the optionally substituted lower alkyl group represented by R1 include a linear or branched C1-6 alkyl group, preferably a methyl group, an ethyl group, an n-propyl group, and the like. is there.

  Examples of the substituent on the lower alkyl group in R1 include an optionally substituted pyridyl group, an optionally substituted amino group, an optionally substituted cycloalkyl group, and an optionally substituted chromanyl group. Can be mentioned.

  Examples of the substituent on the pyridyl group of the substituted pyridyl group include a C1-3 alkyl group, a C1-3 alkoxy group, and a hydroxyl group.

  Examples of the substituent on the amino group of the substituted amino group include a C1 to C3 alkyl group and a pyridyl group.

  Examples of the substituent on the cycloalkyl group of the substituted cycloalkyl group include C1-3 alkyl groups.

  Examples of the substituent on the substituted chromanyl group include a C1 to C3 alkyl group and a hydroxyl group.

Preferable R1 is a group represented by the following formulas (2), (3), (4), (5).

  That is, preferred thiazolidine compounds include rosiglitazone, troglitazone, ciglitazone, pioglitazone, and the like shown in FIG.

  Conventionally, thiazolidine compounds such as rosiglitazone and troglitazone bind to peroxisome proliferator-activated receptor gamma (PPARγ), a nuclear receptor transcription factor of adipocytes. It is known to promote transcription of various molecules. As a result, it has been used to promote the differentiation of adipocytes, and as a result, improve insulin resistance and thus treat diabetes.

  However, this study has revealed that some thiazolidine compounds block (suppress) the function of G protein-coupled receptors such as GPR120. Specifically, thiazolidine-based compounds act on intestinal hormone-secreting cells and the like to suppress the release of peptide hormones such as cholecystokinin (CCK) and mediators such as chemokines, and to suppress diseases caused by these mediators. Based on the mechanism. That is, the thiazolidine-based compound exhibits a previously unknown medicinal effect on the basis of a novel mechanism that is completely different from the conventional mechanism for treating diabetes or the like.

The medicament of the present invention contains the above G protein-coupled receptor inhibitor as an active ingredient, which makes it possible to treat various diseases. For example, gastrointestinal diseases using the inhibitory action of the increase in intracellular Ca ²⁺ concentration that occurs in conjunction with the activation (activation state) of G protein-coupled receptors in the intestinal tract, lungs, brain, etc. where cells having GPR120 are present It can be used as a therapeutic drug, a psychiatric disorder drug, a hypoglycemia drug, and a pulmonary drug.

  In addition, in the treatment of various diseases using a ligand that promotes the secretion of a physiologically active substance such as CCK or GLP-1, an excessive increase in the concentration of the physiologically active substance can be suppressed by using the medicament of the present invention in combination. Therefore, it becomes possible to adjust the physiologically active substance in various cells of various organs to an appropriate concentration.

  The target disease and efficacy of the present invention are based on the suppression of CCK concentration in the blood or target organ and the suppression of inflammatory mediators such as chemokines in the blood or target organ.

Examples of treatment target diseases associated with a decrease in CCK concentration include:
1) Since CCK promotes pancreatitis, treatment of pancreatitis by lowering CCK concentration 2) Since CCK causes behavioral disturbance due to anxiety and stress in the center, treatment of behavioral disorder by lowering CCK concentration 3) CCK Treatment of gastric hyperacidity by reducing gastric acid secretion by reducing concentration 4) Enhancement of analgesic effect by lowering CCK concentration to antagonize analgesics such as morphine 5) By lowering CCK concentration The treatment is for diarrhea such as infectious diarrhea. The present invention is not limited to this description, and the entire treatment associated with an increase or decrease in CCK concentration is targeted.

  On the other hand, as a disease to be treated with suppression of release of inflammatory mediators such as chemokines via G protein-coupled receptors such as GPR120, for example, enteritis such as inflammatory bowel disease (IBD), irritable bowel syndrome (IBS), etc. There are disorders such as degenerative neurological diseases and immune neurological diseases or lung diseases such as chronic obstructive pulmonary disease (COPD). The present invention is not limited to this description, and the entire treatment of diseases associated with chemokine release via G protein-coupled receptors such as GPR120 is targeted.

  Naturally, the efficacy of a thiazolidine-based compound differs from the case where it acts directly on the target organ and the case where it suppresses the release of peptide hormones such as CCK and inflammatory mediators such as chemokines via G protein-coupled receptors such as GPR120. If the medicinal effects different from the conventional direct action are described in particular, the application expansion will be as follows.

1) Treatment for pancreatitis 2) Tranquilizer 3) Treatment for hyperacidity 4) Analgesic adjuvants such as morphine 5) Antidiarrhea 6) Treatment for enteritis 7) Treatment for lung diseases

  The protein-coupled receptor inhibitor of the present invention or a medicine containing this as an active ingredient makes it possible to effectively use antagonists of G protein-coupled receptors such as GPR120 for various purposes.

  Hereinafter, in the embodiment of the present invention, a GPCR inhibitor containing a thiazolidine-based compound as an active ingredient and a medicine containing a GPCR inhibitor as an active ingredient, particularly focusing on functioning as an antagonist to GPR120 and the like explain.

GPR120 and the like are present in the intestinal tract, lungs, brain, fat cells, and pituitary gland, and when GPR120 and the like are blocked by a thiazolidine compound, as shown in Examples described later, suppress an increase in intracellular Ca ²⁺ concentration. It was recognized that there was an effect. Then, by suppressing the increase in Ca ²⁺ concentration, for example, the release of peptide hormones such as cholecystokinin (CCK) and GLP-1 from intestinal hormone-secreting cells having GPR120 is suppressed, and for example, diarrhea is suppressed. Effect is obtained. In addition, when stress is applied, a thiazolidine-based compound that suppresses GPR120 action, particularly rosiglitazone and troglitazone, develops due to stress load because it exhibits a hypersensitive reaction to fatty acids and derivatives thereof from food and is involved in the progression of the disease. It is desirable as a therapeutic agent for enteritis that exacerbates symptoms, such as IBS (irritable enteritis) and IBD (ulcerative enteritis). Therefore, by using thiazolidine compounds as inhibitors such as GPR120, it can be used for treating diseases and improving constitution.

  G protein-coupled receptor inhibitors such as GPR120 are formulated to be compatible with therapeutically relevant routes of administration, including intravenous and oral administration. Specifically, administration is possible in the following manner.

  A preparation suitable for injection is a sterile injectable solution or dispersion medium, a sterile aqueous solution (water soluble) or dispersion medium and a sterile powder (lyophilized protein) for preparation at the time of use. , Nucleic acids and the like). For intravenous administration, suitable carriers include physiological saline, bacteriostatic water, CREMOPHOR ELTM (BASF, Parsippany, NJ), or phosphate buffered saline (PBS). When used as an injection, the GPCR agonist must be sterilized and retain sufficient fluidity to be administered using a syringe. The carrier can be a solvent or dispersion medium containing, for example, water, ethanol, polyol (for example, glycerol, propylene glycol, and liquid polyethylene glycol, and the like), and suitable mixtures. For example, using a coating agent such as lectin, maintaining a required particle size in the dispersion medium, and maintaining a proper fluidity by using a surfactant. Various antibacterial and antifungal agents, such as parabens, chlorobutanol, phenol, ascorbic acid, and thimerosal can be used to prevent microbial contamination. In addition, polyalcohols such as sugar, mannitol, sorbitol, and agents that maintain isotonicity such as sodium chloride may be included in the composition. Compositions that can delay adsorption include agents such as aluminum monostearate and gelatin.

  Sterile injectable solutions are prepared by adding the required ingredients alone or in combination with other ingredients, then adding the required amount of active compound in a suitable solvent and sterilizing. Generally, a dispersion medium is prepared by incorporating the active compound into a sterile medium that contains a basic dispersion medium and the other necessary ingredients described above. Sterile powder preparation methods for the preparation of sterile injectable solutions include vacuum drying and lyophilization to prepare a powder containing the active ingredient and any desired ingredients derived from the sterile solution. It is.

  In the case of an oral preparation, an inert diluent or a carrier that does not cause harm even when taken into the body is included. Oral formulations are, for example, contained in gelatin capsules or compressed into tablets. For oral treatment, the active compound is incorporated with excipients and used in the form of tablets, troches, or capsules. Oral preparations can also be prepared using a flowable carrier. In addition, pharmaceutically compatible binding agents, and / or adjuvant materials may be included.

  Tablets, pills, capsules, lozenges and the like can contain any of the following ingredients or compounds with similar properties. Excipients such as microcrystalline cellulose, binders such as gum arabic, tragacanth or gelatin; swelling agents such as starch or lactose, alginic acid, PRIMOGEL or corn starch; lubricants such as magnesium stearate or STRROTES; colloidal silicon Lubricants such as dioxides; sweeteners such as sucrose or saccharin; or flavoring agents such as peppermint, methyl salicylic acid or orange flavor.

  When a preparation for systemic administration is used, it can be transmucosally or transdermally. For transmucosal or transdermal administration, penetrants that can penetrate the target barrier are selected. Transmucosal penetrants include surfactants, bile salts, and fusidic acid derivatives. Nasal sprays or suppositories can be used for transmucosal administration. For transmucosal administration, the active compounds are formulated in ointments, ointments, gels or creams.

  In addition, the medicament containing the thiazolidine compound of the present invention as an active ingredient is prepared in the form of a suppository (for example, with a base such as cocoa butter and other glycerides) or a retentive enema for rectal delivery. You can also

  When a medicine containing the thiazolidine compound of the present invention as an active ingredient is used as a controlled release preparation, it can be prepared using a carrier that can prevent immediate removal from the body. For example, biodegradable, biocompatible polymers such as ethylene vinyl acetate, polyanhydrides, polyglycolic acid, collagen, polyorthoesters, and polylactic acid can be used. Such materials are available from ALZA Corporation (Mountain View, CA) and NOVA Pharmaceuticals, Inc. (Lake Elsinore, CA) and can also be readily prepared by one of ordinary skill in the art. Liposomal suspensions can also be used as pharmaceutically acceptable carriers. Useful liposomes are prepared as a lipid composition comprising, but not limited to, phosphatidylcholine, cholesterol and PEG-derivatized phosphatidylethanol (PEG-PE) through a filter of appropriate pore size so as to be of a size suitable for use. Purified by evaporation. For example, Fab 'fragments of antibodies and the like may be bound to liposomes via a disulfide exchange reaction (Martin and Papahadjopoulos, 1982). For details of the preparation method, for example, the description in the literature (Eppstein et al., 1985; Hwang et al., 1980) can be referred to.

  The dose of the pharmaceutical agent containing the GPCR inhibitor of the present invention as an active ingredient depends on the condition of the patient (human) or animal to be administered, the administration method, etc. in the treatment or prevention of a specific disease. If so, it can be easily optimized.

  For example, in the case of injection administration, for example, it is preferable to administer about 0.1 μg / kg to 500 mg / kg of the patient's body weight per day, and it will generally be possible to administer in one or more divided doses. Preferably, the dosage level is about 0.1 μg / kg to about 250 mg / kg per day, more preferably about 0.5 to about 100 mg / kg per day.

  For oral administration, it is preferably provided in the form of a tablet containing 1.0 to 1000 mg of active ingredient. Preferably, the dosage of the active ingredient for the patient (human) or animal to be treated is 0.01-100 mg / kg. The compounds are administered on a regimen of 1 to 4 times daily, preferably once or twice daily.

With the GPCR inhibitor agonist of the present invention, for example, the following diseases can be treated through an inhibitory action on the release of CCK, GLP-1, etc. by suppressing the concentration of intracellular Ca ²⁺ concentration.
1. Since CCK promotes pancreatitis, it is possible to treat pancreatitis by lowering the CCK concentration.
2. Since CCK promotes secretion of gastric acid, suppression of CCK concentration enables treatment of gastric hyperacidity.
3. Since CCK causes behavioral disorders due to anxiety and stress in the center, suppression of CCK concentration enables treatment of behavioral disorders and use as a tranquilizer.
4). Since CCK antagonizes analgesics such as morphine, the effect of the analgesic can be enhanced by lowering the CCK concentration.
5. Suppression of CCK concentration enables treatment of diarrhea such as infectious diarrhea.
6). Suppression of GLP-1 concentration enables treatment of hypoglycemia.
However, the present invention is not limited to the above diseases, and the secretion of various physiologically active substances other than CCK and GLP-1 by treatment of various diseases using a decrease in the concentration of CCK and GLP-1 and suppression of Ca ²⁺ concentration. It is possible to treat various diseases by utilizing the action of suppressing the above.

(Example 1)
In this example, constructs with a FLAG tag at the N-terminus of human GPR40 (see WO2004 / 065960) and human GPR120 (see Sequence Listing 1) were prepared as G protein-coupled receptors, respectively, and incorporated into an expression vector capable of drug induction. Stable expression cells of GPR40-carrying cells and GPR120-carrying cells were prepared. And by giving a predetermined treatment to the cells, the receptor was expressed, and the Ca ²⁺ concentration was measured. Instead of human GPR120, mouseGPR120 shown in Sequence Listing 2 may be used.

1. Measurement of intracellular Ca ²⁺ concentration
(1) A GPR40-carrying cell line and a GPR120-carrying cell line were used in this experiment.
(2) Each cell was seeded on a 96-well 1 clear bottom plate for cell culture whose surface was treated with collagen so that the cell density was 2 × 10 ⁵ cells / well / 50 μl to prepare a cell plate.
(3) FBS (-) Medium containing Doxycyclin at a concentration of 10 μg / ml was used for the seeding solution. Doxycyclin treatment and Starvation were allowed to proceed simultaneously by culturing with this solution for 21 hours.

2. Loading of Ca ²⁺ ion sensitive fluorescent dye into cells (1) In order to load calcium sensitive fluorescent dye into cells, Molecular Devices Calcium Assay Kit reagent was used.
(2) This reagent was dissolved in FLIPR buffer (solution prepared to pH 7.4 by adding 20 mM HEPES to Hanks' Balanced Salt Solution) at a double concentration described in the kit manual.
(3) 50 μl / well of the prepared Calcium Assay Kit reagent solution was added to the cell plate 21 hours after seeding. Since 50 μl / well of medium was used at the time of cell seeding, the concentration of the Calcium Assay Kit reagent in the well became the predetermined concentration described in the manual.
(4) The calcium ion sensitive fluorescent dye contained in the kit reagent was loaded into the cells by allowing the cells to which the Calcium Assay Kit reagent solution had been added to stand in the dark at room temperature for 1 hour.

3. Ligand Preparation (1) Thiazolidine compounds such as rosiglitazone, troglitazone, ciglitazone and the like shown in FIG. 2 were prepared as test compounds.
The above compounds were dissolved in DMSO and diluted with the FLIPR buffer so that the concentration of each compound was 1.5 × 10 −4 M and the DMSO concentration was 1.5%.

(2) DMSO at the same concentration as described above was used as a negative control, and α-LA, which is a known ligand of GPR40 and GPR120, was used as a positive control.

(3) Measurement of increase in intracellular Ca ²⁺ concentration 0.3% DMSO and 3 × 10 ⁻⁸ to 1 × 10 3 for GPR40-carrying cells and GPR120-carrying cells treated with no treatment (induction-) and expression induction (induction +) Various compounds of ⁻⁴ M (0.3% DMSO) were added, and the intracellular Ca ²⁺ concentration was measured with FLIPR (Molecular Devices). The peak value was detected and standardized (normalized) by the peak value when 1 × 10 ⁻⁴ M α-LA (cis type) was added.

FIGS. 1 (a) and 1 (b) show, in order, stimulation of α-LA (cis-type α-linolenic acid) in GPR40-carrying cells and GPR120-carrying cells treated with no treatment (induction-) and expression induction (induction +). It is data which shows the increase amount of Ca ^<2+ > density ^| concentration. 1A and 1B, the horizontal axis indicates the added concentration of α-LA, and the vertical axis indicates the amount of increase in Ca ²⁺ concentration. In any cell, an increase in intracellular Ca ²⁺ concentration was confirmed by the expression induction treatment (induction +), and it was confirmed that the expression induction treatment (induction +) was appropriately performed.

As already explained, the intracellular Ca ²⁺ concentration corresponds to the secreted amount of hormone, which is a physiologically active substance, from the cell, and it has been confirmed that it is an index representing the physiological activity state of the cell. Therefore, the following remarkable facts regarding the activation of each cell were confirmed from the data of FIGS. 3 to 6 described below.

-For GPR40-bearing cells-
FIG. 3 is a graph showing data on the increase in Ca ²⁺ concentration when thiazolidine-based compounds such as rosiglitazone, troglitazone and siglitazone are added to GPR40-bearing cells. The following can be seen from FIG.

The addition of troglitazone and ciglitazone hardly increases the intracellular Ca ²⁺ concentration, and therefore it is presumed that troglitazone and ciglitazone have almost no function as agonists for GPR40.

On the other hand, the addition of rosiglitazone significantly increased the intracellular Ca ²⁺ concentration, and it is estimated that rosiglitazone has a function as an agonist of GPR40.

-For GPR120-bearing cells-
FIG. 5 is a graph showing data on the increase in Ca ²⁺ concentration when thiazolidine-based compounds such as rosiglitazone, troglitazone, and ciglitazone are added to GPR120-bearing cells. The following can be understood from FIG.

With the addition of rosiglitazone, troglitazone, and ciglitazone, there is almost no increase in intracellular Ca ²⁺ concentration. Therefore, rosiglitazone, troglitazone, and ciglitazone are presumed not to have a function as agonists for GPR40.

In this Example, the amount of insulin secreted by GPR40-carrying cells and GLP-1 secreted by GPR120-carrying cells increased with the increase in intracellular Ca ²⁺ concentration. It has also been confirmed.

(4) Confirmation of cell activation inhibitory effect The activation inhibitory effect (antagonist activity) of the test compound is determined after pretreatment of the cells with the test compound at the same concentration as in the evaluation of the agonist activity for 10 minutes, that is, GPR40 Alternatively, after the treatment of activating cells by expressing GPR120, how much increase in intracellular Ca ²⁺ concentration is inhibited when cells in the activated state are stimulated with the same concentration of α-LA It was evaluated by what. Specifically, first, the test compound in the pretreatment plate is added to each well of the cell plate after dye loading using a multipipette, and then the cell plate and α-LA (cis type) are dispensed. The plate was set on the FLIPR, and the FLIPR was operated so that the addition of α-LA (cis type) was 10 minutes after the addition of the test compound. ) The time course change of intracellular Ca ²⁺ concentration before and after the addition stimulation was measured.

  In addition, after the measurement, the peak value of well fluorescence intensity increase corresponding to each test compound pretreatment was determined. This value was normalized with the value at the time of stimulation with α-LA when pretreated with DMSO alone, and the activation inhibitory effect (antagonist activity) of each test compound was evaluated.

-For GPR40-bearing cells-
FIG. 4 is data showing the effect of suppressing activation when a thiazolidine-based compound is added to GPR40-bearing cells. That is, this data shows the inhibitory activity of thiazolidine compounds such as rosiglitazone, troglitazone, and ciglitazone on GPR40-bearing cells activated by α-LA (cis type).

As shown in FIG. 4, when the Ca ²⁺ concentration increase in the activated state is 1 (normalized value), the addition of rosiglitazone decreases the Ca ²⁺ concentration almost to 0. It is greatly suppressed (inhibited) to a state where activation is hardly observed. Therefore, the activation suppression effect (normalized value) of GPR40-bearing cells by rosiglitazone is 1.0. In addition, the addition of troglitazone also decreased the Ca ²⁺ concentration to about 0.22 (normalized value), and the activation of GPR40-carrying cells was suppressed (inhibited). And the activation suppression effect (normalized value) of the GPR40 possession cell by troglitazone is 0.78.

  As can be seen from this data, rosiglitazone and troglitazone function as antagonists to GPR40. In particular, the antagonist function of rosiglitazone is very strong. According to the literature (The FFA recepter GPR40 links hyperinsulinemia, hepatic steatosis, and impaired glucose homeostasis in mouse (Cell Metab. 2005 Apr; 1 (4): 245-58)) Is disclosed to induce diabetes. Therefore, when rosiglitazone and troglitazone are administered, by suppressing the activity of GPR40, various diseases resulting from long-term stimulation of GPR40 such as diabetes induction, pancreatic β-cell exhaustion, and diseases caused based on these diseases such as high It is estimated to be effective for lipemia, obesity, metabolic syndrome, diabetic nephritis, etc.

On the other hand, the addition of siglitazone hardly reduces the Ca ²⁺ concentration, and the activation of GPR40-carrying cells is hardly suppressed (inhibited). Therefore, the activation inhibitory effect (normalized value) of GPR40-bearing cells by siglitazone is zero.

-For GPR120-bearing cells-
FIG. 6 is data showing the effect of inhibiting activation when a thiazolidine-based compound is added to GPR120-bearing cells. That is, it is data showing the inhibitory activity of thiazolidine compounds such as rosiglitazone, troglitazone, and ciglitazone on GPR120-bearing cells activated by α-LA (cis type).

As shown in FIG. 6, the addition of rosiglitazone decreased the Ca ²⁺ concentration to 0.01 (normalized value), and it was greatly suppressed (inhibited) to a state where almost no activation of GPR120-bearing cells was observed. ) Therefore, the activation suppression effect (normalized value) of GPR120-bearing cells by rosiglitazone is 0.99.

Further, the addition of troglitazone also greatly reduces the Ca ²⁺ concentration to about 0.02 (normalized value), and the activation of GPR120-carrying cells is greatly suppressed (inhibited). And the activation suppression effect (normalized value) of the GPR120 possession cell by troglitazone is 0.98.

On the other hand, the addition of siglitazone reduces the Ca ²⁺ concentration to about 0.42 (normalized value), and the activation of GPR-bearing cells GPR120 is considerably suppressed (inhibited). And the activation suppression effect (normalized value) of the GPR120 possession cell by siglitazone is 0.58.

  In other words, the experiment of this example demonstrated that thiazolidine compounds such as rosiglitazone, troglitazone and siglitazone can be used as antagonists of G protein receptors such as GPR40 and GPR120.

  In particular, it was confirmed that any compound has a function as an antagonist of GPR120. As for GPR40, it was confirmed that rosiglitazone and troglitazone have a function as an antagonist.

  As derived from the data shown in FIGS. 4 and 6, the activity can be quantitatively suppressed by adding a thiazolidine compound to cells in an activated state. The activated state is realized by adding an agonist to the G protein-coupled receptor. However, in a pathological condition in which the GPR120 response is hypersensitive, for example, when GPR120 is excessively expressed, an antagonist thiazolidine compound is added. Thus, the disease state can be treated. Therefore, the G protein-coupled receptor inhibitor containing the thiazolidine compound of the present invention as an active ingredient can be used as a medicine or a nutritional supplement for diseases caused by hypersensitivity of the GPR120 reaction.

  Conventionally, the function as an agonist or antagonist for a G protein-coupled receptor with a thiazolidine compound has been hardly known. However, from the results of this experiment, an antagonist of a thiazolidine compound against a G protein-coupled receptor, particularly GPR120 ( The function as an inhibitor was confirmed. And the effect as an above-mentioned therapeutic agent, a constitution improvement agent, etc. can be exhibited with the pharmaceutical which contains the inhibitor of GPR120 of this invention as an active ingredient.

  The function of thiazolidine compounds as agonists for GPR40-carrying cells and GPR120-carrying cells was confirmed only in rosiglitazone. However, by obtaining knowledge about other thiazolidine-based compounds in the future, G protein-coupled receptors It is expected that a substance that functions as an agonist of the above will be obtained.

In this example, the measurement of intracellular Ca ²⁺ concentration was adopted as a method for quantitatively examining activation and activation inhibition, but other physiologically active substances such as CCK, cytocan, ERK, or signal transduction. Substance detection can be used.

And the treatment of various diseases is attained by the medicament of the present invention containing a G protein-coupled receptor inhibitor containing a thiazolidine compound as an active ingredient. That is, digestive tract diseases utilizing the inhibitory action on the increase in intracellular Ca ²⁺ concentration that occurs in conjunction with the activation (activation state) of G protein-coupled receptors in the intestinal tract, lungs, brain, etc. where cells having GPR120 are present It can be used as a therapeutic drug, a psychiatric disorder drug, a hypoglycemia drug, and a pulmonary drug.

  In addition, as can be seen from the data shown in FIGS. 4 and 6 and the literature “NATURE MEDICINE Vol.11 No.1 Jan.2005 p.90-94”, inhibition of G protein-coupled receptors containing thiazolidine compounds as active ingredients By administering the agent to the receptor-carrying cell, the activation of the receptor of the cell is inhibited. Therefore, a screening cell in which the function of a certain receptor is inhibited can be prepared. Then, by adding a test compound to the cells and measuring intestinal hormones in the blood, such as GLP-1, CCK, etc., a ligand effective in restoring the function of the receptor can be screened.

  Similarly, as can be seen from the data shown in FIGS. 4 and 6 and the document “Am. J. Clin. Nutr. 2001; 73: p.1019-1026”, a G protein-coupled type containing a thiazolidine compound as an active ingredient. By administering a receptor inhibitor to an animal, the activation of the receptor present in cells of each organ of the animal is inhibited, so that an animal for screening in which the function of a certain receptor is inhibited can be created. . Then, by adding a test compound to the animal and measuring an intestinal hormone in the blood, such as GLP-1, CCK, etc., a ligand effective for restoring the function of the receptor can be screened.

  The G protein-coupled receptor inhibitor of the present invention can be used not only as a medicine but also as a nutritional supplement.

(A), (b) is a graph which shows the increase data of Ca < ²⁺ > density | concentration by stimulation of (alpha) -LA (cis type | mold) in the GPR40 possession cell GPR120 possession cell which carried out the non-treatment and expression induction treatment in order. It is a figure which shows the chemical structure of thiazolidine type compounds, such as rosiglitazone (Rosiglitazone), troglitazone (Troglitazone), ciglitazone (Ciglitazone), pioglitazone (Pioglitazone). It is a graph which shows the increase amount data of Ca ^<2+ > density ^| concentration when thiazolidine type compounds, such as rosiglitazone, troglitazone, and siglitazone, are added to GPR40 possession cell. It is data which shows the activation inhibitory effect when a thiazolidine type compound is added to GPR40 possession cell. It is a graph which shows the increase amount data of Ca < ²⁺ > density | concentration when thiazolidine type compounds, such as rosiglitazone, troglitazone, and siglitazone, are added to GPR120 possession cell. It is data which shows the activation inhibitory effect when a thiazolidine type compound is added to GPR120 possession cell.

Claims (6)

  A G protein-coupled receptor inhibitor comprising a thiazolidine compound as an active ingredient and suppressing the function of a G protein-coupled receptor (GPCR). The G protein-coupled receptor inhibitor according to claim 1,
The thiazolidine compound has the following general formula (1):

A G protein-coupled receptor inhibitor, which is a compound represented by (R represents a lower alkyl group which may be substituted). The G protein-coupled receptor inhibitor according to claim 2,
A G protein-coupled receptor inhibitor, wherein the thiazolidine compound is at least one compound selected from the group consisting of rosiglitazone, troglitazone, and ciglitazone. In the G protein coupled receptor inhibitor according to any one of claims 1 to 3,
The G protein-coupled receptor inhibitor is GPR120, wherein the G protein-coupled receptor is GPR120.   A pharmaceutical comprising a thiazolidine compound as an active ingredient and a G protein-coupled receptor inhibitor as an active ingredient. The medicine according to claim 6,
A pharmaceutical agent that functions as at least one therapeutic agent selected from a therapeutic agent for digestive system diseases, a therapeutic agent for mental disorders, hypoglycemia, and pulmonary disease.

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