WO2008016118A1 - Agent targeting dopamine receptor and screening method therefor - Google Patents

Abstract

It is intended to obtain a new finding related to a mechanism of inducing differentiation of Th1/Th2/Th17 through dendritic cells, and to provide a pharmaceutical effective in an immunity-related disease by utilizing the finding, a screening method for the pharmaceutical, and an effective agent for regulating a mechanism of inducing differentiation of Th1/Th2/Th17 through dendritic cells. The invention relates to a pharmaceutical for treating or preventing a disease caused by the imbalance of Th, characterized by containing a compound that modifies the activity of dopamine receptor as an active ingredient, a screening method for the pharmaceutical, and various agents characterized by containing a dopamine D1-like receptor antagonist or a dopamine D2-like receptor antagonist as an active ingredient.

Description

 Specification

 Drugs targeting dopamine receptors and screening methods thereof

 [0001] The present invention relates to a medicament for immune-related diseases such as multiple sclerosis, and a screening method for the medicament, utilizing new knowledge about the Thl / Th2 / Thl7 differentiation induction mechanism via dendritic cells. The present invention also relates to various drugs for controlling the Thl / Th2 / Thl 7 differentiation induction mechanism via dendritic cells.

 Background art

 [0002] The helper T cells, which play a central role in acquired immunity, are divided into Thl (type 1 helper T cells) that promote cellular immunity and Th2 ( Type 2 helper T cells). Dendritic cells (DC) are antigen-presenting cells that play an important role in the early stages of the immune response. Recently, they have promoted Thl induction (DC1) or Th2 induction (DC2)! It became clear that there was a subset with functional differences. Several attempts have been made to correct the biased Thl / Th2 balance by artificially controlling the induction of Thl and Th2 from naive T cells via DC, and reported successful cases. (Morita Y et al .; Dendritic ce lis genetically engineered to express IL—4 inhioit murine collagen-induced arthritis. J Clin Invest. 2001 May; 107 (10): 1275—84.).

[0003] Conventionally, Thl / Th2 balance abnormalities are thought to be involved in the development of various immune-related diseases. For example, balance bias to Thl cells is a chronic inflammatory disease such as rheumatoid arthritis and , Organ-specific autoimmune diseases (e.g., multiple sclerosis, type 1 diabetes, inflammatory bowel disease, glomerulonephritis, hepatitis, liver damage, autoimmune hemolytic anemia, leukopenia, thrombocytopenia, demyelination Disease, Hashimoto's thyroiditis, pernicious anemia, psoriasis) and the balance bias to Th2 cells is thought to be involved in allergic diseases and many systemic autoimmune diseases. On the other hand, several papers recently reported Thl 7, a new Th subfraction. These cells are involved in exacerbating autoimmune inflammation by producing tangle IL-17. In particular, the aforementioned multiple sclerosis and Rheumatoid arthritis is suspected to be due to this Thl 7 bias! /, But it is considered to be strong! /, (Batten, M et al .; Interleukin 27 limits autoimmune encephalomyelitis by s uppressing the development of interleukin 17 — Producing T cells. N at Immunol. 2006. 7: 929—936. Thl 7 interacts with Thl in a mutually ft manner and with Th2 in a mutually reinforcing manner.

 [0004] As a method for effectively treating or preventing various immune-related diseases as described above, many methods for adjusting an abnormal Thl / Th2 balance as desired have been proposed. Specifically, For example, a method for treating a Thl-mediated disease by administering a TCCR (T cell site force-in-receptor) polypeptide antagonist (JP 2003-512824); excess caused by autoimmune diseases such as multiple sclerosis Method for suppressing Thl cell-mediated immune response by use of xanthophyll (Japanese Patent Publication No. 2003-510353); a pharmaceutical composition comprising a specific benzhydryl derivative for treating or preventing organ-specific autoimmune diseases, etc. 2003-300881)) and the like have been proposed.

 [0005] However, various factors are involved in the acquired immune system in a complicated manner, and it is still speculated whether which immune-related disease is involved in Thl, Th2 or Thl 7 bias. Some aspects are not exceeded. Therefore, more accurate knowledge on the Thl / Th2 / Thl 7 differentiation-inducing mechanism, in particular, the Thl / Th2 / Thl 7 differentiation-inducing mechanism via dendritic cells, and more on immune-related diseases using the above knowledge. At present, the development of effective medicines is desired.

 Disclosure of the invention

 [0006] An object of the present invention is to solve the conventional problems and achieve the following objects. That is, the present invention obtains new knowledge regarding the Thl / Th2 / Thl 7 differentiation induction mechanism via dendritic cells, and uses the knowledge to screen an effective drug for immune-related diseases and screening of the drug. It is an object of the present invention to provide a method and an effective drug for controlling a Thl / Th2 / Th17 differentiation induction mechanism via dendritic cells.

[0007] In order to solve the above-mentioned problems, the present inventors diligently studied and as a result, obtained the following knowledge. That is, the first is the finding that the action of dopamine (DA) is greatly involved in the mechanism of inducing differentiation of Thl / Th2 / Thl 7 through dendritic cells. Second, the conventional Thl deflection Various organ-specific autoimmune diseases and chronic inflammatory diseases, including those that have been attributed to the treatment of TM7 and Th2 differentiation, or induction of Thl differentiation through modification of dopamine receptor activity This is a finding that it can be effectively treated or prevented.

[0008] With regard to the first finding, it has been conventionally known that various site force ins are involved in the process of induction of Thl / Th2 / Thl 7 differentiation by dendritic cells (DC). In particular, when IL-12 production from DC is dominant, Thl differentiation is promoted, and IL-4 PGE production

 2 It was known that Th2 differentiation was promoted when life was dominant, and 1¾17 differentiation was promoted when production of 1-6 ゃ 0? / 3 was dominant.

[0009] However, it has not been known so far that the action of dopamine (DA) is greatly involved in the DC-mediated Thl / Th2 / Thl 7 differentiation induction mechanism. Look.

 [0010] Regarding the second finding, conventionally, attempts to treat immune-related diseases by adjusting the abnormality of Thl / Th2 / Thl7 balance have been widely performed. In particular, many organ-specific autoimmune diseases are conventionally thought to be caused by a bias in the balance of Thl cells, and can be treated or prevented by inducing differentiation into Th2 cells. There have been widespread attempts to do it.

 [0011] However, at least some of these organ-specific autoimmune diseases are actually diseases caused by a balance bias toward Thl 7 cells or Th2 cells, and thus suppress differentiation into Thl 7 cells or Th2 cells. In addition, it has not been known at all that the disease can be effectively treated or prevented by inducing differentiation into Thl cells, which is a new finding of the present inventors.

 [0012] The present invention is based on the above findings by the present inventors, and more specifically includes the following invention.

 [l] A medicament for treating or preventing a disease caused by Th imbalance, comprising a compound that modifies the activity of a dopamine receptor as an active ingredient.

[2] The disease caused by Th imbalance is a disease caused by excessive reaction of TM7 or Th2, and the compound that modifies the activity of dopamine receptor is dopamine D1-like receptor antagonist. [1] The medicament according to 1. [3] The medicament according to [2], wherein the dopamine Dl-like receptor antagonist is SCH23390.

 [4] A screening method for a drug for the treatment or prevention of a disease caused by Th imbalance, wherein binding to a donomin receptor is used as an index.

 [5] The screening method according to [4], wherein the disease caused by Th imbalance is a disease caused by excessive reaction of TM7 or Th2, and the dopamine receptor is a dopamine D1-like receptor.

 [6] A screening method for a drug for the treatment or prevention of a disease caused by Th imbalance, wherein the screening method uses a modification of the activity of a donomin receptor as an index.

 [7] The disease caused by Th imbalance is a disease caused by excessive reaction of TM7 or Th2, and the modification of the activity of the dopamine receptor is inhibition of the activity of the dopamine D1-like receptor. Screening method.

 [8] A drug that inhibits the synthesis or storage of donomin in dendritic cells and contains a dopamine D1-like receptor antagonist as an active ingredient.

 [9] An agent that suppresses differentiation of naive T cells into Thl 7 or Th2, or promotes differentiation into Thl, and contains a dopamine D1-like receptor antagonist as an active ingredient.

 [10] The drug according to [8] or [9], wherein the dopamine D1-like receptor antagonist is SCH23390.

 [11] A drug that promotes the synthesis or storage of donomin in dendritic cells and contains a dopamine D2-like receptor antagonist as an active ingredient.

 [12] A drug that promotes differentiation of naive T cells into Th2, and contains a dopamine D2-like receptor antagonist as an active ingredient.

 [0013] According to the present invention, various problems in the prior art can be solved, and effective for immune-related diseases can be obtained by utilizing new knowledge on Th 1 / Th2 / Thl 7 differentiation induction mechanism via dendritic cells. And an effective drug for controlling the Thl / Th2 / Thl7 differentiation-inducing mechanism via dendritic cells.

 Brief Description of Drawings

[0014] [FIG. 1A] FIG. 1A shows that the addition of various drugs to dendritic cells affects the Th differentiation bias of T cells. It is the graph which showed the influence which drops.

 FIG. 1B is a graph showing the effect of addition of various concentrations of sulpiride on dendritic cells on the Th differentiation tendency of T cells.

2] Fig. 2 is a photograph of an electrophoretic image after RT-PCR that confirmed the expression of the dopamine receptor subtype in dendritic cells.

Fig. 3A is a graph showing the effect of the addition of dorminamine sulpiride on changes in dendritic cell Ca ²⁺ concentration.

Fig. 3B is a graph showing the effect of the addition of dopamine sulpiride on the change in dendritic cell cAMP concentration.

Fig. 4A is a graph showing the effect of the addition of various drugs to dendritic cells on the production of cytodynamic in (IL-12p70) by dendritic cells.

 FIG. 4B is a graph showing the effect of the addition of various drugs on dendritic cells on the surface antigen molecule expression of dendritic cells.

Fig. 5A is a photograph of a confocal microscope image showing that donomin is stored in granules in dendritic cells.

 [FIG. 5B] FIG. 5B is a photograph of a fluorescence microscopic image showing the effect of addition of various drugs on dendritic cells on the amount of dopamine stored in granules.

6A] Fig. 6A is a graph showing changes in cAMP concentration in T cells due to dopamine stimulation at various concentrations.

 FIG. 6B is a graph showing changes in cAMP concentration in T cells when SCH23390 (dopamine D1-like receptor inhibitor) is added in addition to dopamine stimulation.

Fig. 6C is a graph showing the effect of various concentrations of donomin stimulating cells on Th differentiation bias.

 [FIG. 7A] FIG. 7A is a photograph of a phase-contrast microscope image showing a state of dendritic cell uncondyles by co-culture (alio-MLR) of dendritic cells and T cells.

 FIG. 7B is a graph showing the effect of colchicine treatment on the dendritic cell degranulation effect on Th differentiation bias of T cells.

[Fig. 7C] Fig. 7C shows that α (α- methy has the function of inhibiting the process of dopamine synthesis. This graph shows the effect of 1-p-tyrosine) on Th differentiation bias in T cells.

[FIG. 8] FIG. 8 is a schematic diagram showing a Th differentiation induction mechanism through donomin release from dendritic cells (DCs) suggested in Examples;! -7.

 [FIG. 9A] FIG. 9A is a graph showing the prophylactic effect of EAE by prophylactic administration of various donomin receptor inhibitors.

 FIG. 9B is a graph showing the therapeutic effect of EAE by therapeutic administration of various dopamine receptor inhibitors.

 [FIG. 9C] FIG. 9C is a graph showing T cell Thl / Th2 / Thl 7 differentiation bias (IL-4, INF-γ, IL-17 production) by prophylactic administration of various dopamine receptor inhibitors It is.

[Fig. 9D] Fig. 9D shows Thl / Th2 / Thl 7 differentiation bias (IL-4, INF-γ, IL-17 production) of sputum cells by therapeutic administration of various donomin receptor inhibitors It is a graph.

[Fig. 10] Fig. 10 is a graph showing the preventive effect of CIA by prophylactic administration of various donomin receptor inhibitors.

 [FIG. 11] FIG. 11 is a graph showing the preventive effects of NOD by prophylactic administration of various donomin receptor inhibitors.

 [FIG. 12A] FIG. 12A is a photograph of a light micrograph (positive staining of hematoxylin eosin) of a positive control (PC) group showing the prevention effect of glomerulonephritis by prophylactic administration of various donomin receptor inhibitors. is there.

 [FIG. 12B] FIG. 12B is a photograph of an optical microscope image (stained with hematoxylin eosin) of the negative control (NC) group to show the prevention effect of glomerulonephritis by prophylactic administration of various dopamine receptor inhibitors. .

 [FIG. 12C] FIG. 12C is a photograph of an optical microscope image (hematoxylin eosin staining) of the SCH23390 administration group to show the effect of preventing glomerulonephritis by prophylactic administration of various donomin receptor inhibitors.

BEST MODE FOR CARRYING OUT THE INVENTION

 (Medicine)

The medicament of the present invention is used for treatment or prevention of diseases caused by Th imbalance. It is a pharmaceutical that can be obtained and contains a compound that modifies the activity of the dopamine receptor as an active ingredient, and further contains other ingredients as necessary. The disease caused by Th imbalance is preferably a disease caused by an excessive reaction of Thl 7 or Th2, and in this case, the compound that modifies the activity of the dopamine receptor is preferably a donomin D1-like receptor. Antago is a second striker.

 [0016] <Dopamine D1-like receptor antagonist>

 There are five subtypes of “dopamine receptors” from D1 to D5, and it is generally known that they have a function of transmitting signals into cells in combination with G proteins. D1 and D5 are known as “dopamine D1-like receptors” and are coupled to Gs proteins that increase adenyl cyclase activity and increase cAMP concentration. On the other hand, it is known that D2 to D4 are conjugated with Gi proteins that suppress adenyl cyclase activity as “dopamine D2-like receptors”. Therefore, as the “dopamine D1-like receptor antagonist”, a substance having a function of inhibiting the action of at least one of the subtypes D1 and D5 of the dopamine receptor can be used.

 [0017] That is, the dopamine D1-like receptor antagonist can be appropriately selected according to the purpose without any particular limitation. For example, a known dopamine D1-like receptor antagonist (donomin D1-like receptor inhibitor) Or a substance evaluated to have the ability to bind to and / or inhibit the activity of the donomin D1-like receptor by the screening method of the present invention described later.

 [0018] Specific examples of the dopamine D1-like receptor antagonist include SCH23390, SKF83566, L-Stepholidine, LE300, and the like. Of these, SCH23390 is particularly preferable. The structural formulas of the respective dopamine D1-like receptor antagonists are shown below.

[0019] [Chemical 1]

ine)

[0022] [Chemical 4] (E300)

 [0023] The content of the dopamine Dl-like receptor antagonist in the medicament is not particularly limited and can be appropriately selected according to the purpose, and the medicament is the dopamine D1-like receptor antagonist itself. Moyore.

 [0024] <Other ingredients>

 The other components can be appropriately selected according to the purpose within a range that does not impair the effects of the present invention without particular limitations. Examples thereof include pharmaceutically acceptable carriers. The carrier is not particularly limited, and can be appropriately selected depending on, for example, the pharmaceutical dosage form described below. Further, the content of the other components in the medicine can be appropriately selected according to the purpose without any particular limitation.

 [0025] <Dosage form, production>

 The pharmaceutical dosage form is not particularly limited, and can be appropriately selected according to the desired administration method. For example, oral solid preparations (tablets, coated tablets, granules, powders, capsules, etc.), Oral solutions (internal solutions, syrups, elixirs, etc.), injections (solutions, suspensions, solids for erection, etc.), suppositories, ointments, patches, gels, creams, external powders, sprays And inhalable powders.

 [0026] Examples of the oral solid preparation include, for example, a compound that modifies the activity of the donomin receptor, an excipient, and optionally a binder, a disintegrant, a lubricant, a coloring agent, and a corrigent. * Adds additives such as flavoring agents, and can be manufactured using conventional methods.

[0027] Examples of the excipient include lactose, sucrose, sodium chloride, glucose, starch, calcium carbonate, kaolin, microcrystalline cellulose, and silicic acid. Examples of the binder include water, ethanol, propanol, simple syrup, glucose solution, starch solution, Propinorestarch, methinorescenellose, ethinoresenorelose, shellac, canolecus phosphate, polybylpyrrolidone and the like. Examples of the disintegrant include dry starch, sodium alginate, agar powder, sodium hydrogen carbonate, calcium carbonate, sodium laurinole sulfate, stearic acid monoglyceride, and lactose. Examples of the lubricant include purified talc, stearate, borax, and polyethylene glycol. Examples of the colorant include titanium oxide and iron oxide. Examples of the flavoring / flavoring agent include sucrose, orange peel, citrate, and tartaric acid.

[0028] The oral liquid preparation can be produced by a conventional method, for example, by adding additives such as a flavoring agent, a buffering agent, and a stabilizer to the compound that modifies the activity of the dopamine receptor. .

 [0029] Examples of the flavoring / flavoring agent include sucrose, orange peel, citrate, and tartaric acid. Examples of the buffer include sodium quenate. Examples of the stabilizer include tragacanth, gum arabic, and gelatin.

 [0030] As the injection, for example, a pH regulator, a buffer, a stabilizer, a tonicity agent, a local anesthetic, and the like are added to a compound that modifies the activity of the dopamine receptor, and a conventional method is used. Subcutaneous, intramuscular and intravenous injections can be manufactured.

 [0031] Examples of the pH adjusting agent and the buffering agent include sodium citrate, sodium acetate, sodium phosphate and the like. Examples of the stabilizer include sodium pyrosulfite, EDTA, thioglycolic acid, and thiolactic acid. Examples of the isotonic agent include sodium chloride and glucose. Examples of the local anesthetic include pro-power-in hydrochloride and lidocaine hydrochloride.

 As the suppository, for example, a compound that modifies the activity of the dopamine receptor, a carrier for a known suppository formulation such as polyethylene glycol, lanolin, cocoa butter, fatty acid triglyceride, and a tween if necessary After adding a surfactant such as (TWEEN: registered trademark), it can be manufactured in a conventional manner.

[0033] As the ointment, for example, a compound that modifies the activity of the dopamine receptor is mixed with a known base, stabilizer, wetting agent, preservative, etc., and mixed by a conventional method. Can do.

 [0034] Examples of the base include liquid paraffin, white petrolatum, white beeswax, octyl dodecyl alcohol, and norafine. Examples of the preservative include methyl paraoxybenzoate, ethyl parabenzoate, and propyl parabenzoate.

 [0035] As the patch, for example, it is possible to apply a cream, a glue, a paste or the like as the ointment to a known support by a conventional method. Examples of the support include cotton, suf, woven fabrics made of chemical fibers, non-woven fabrics, soft chlorinated chloride, polyethylene, polyurethane and other films, foam sheets, and the like.

 [0036] <Target disease>

 The target disease of the medicine is not particularly limited as long as it is a disease caused by Th imbalance, and can be appropriately selected according to the purpose, preferably a disease caused by Thl 7 or Th2 excess reaction. . In the present invention, it has been found that the disease is caused by at least a partial force S of a disease that has been conventionally attributed to an excessive reaction of Thl, and actually a balance bias toward Thl 7 or Th2. Therefore, in addition to allergic diseases and many systemic autoimmune diseases that have been conventionally known as diseases caused by excessive reaction of Thl 7 or Th2, as diseases caused by Thl 7 or Th2 excessive reactions, This includes diseases that have been attributed to Thl overreaction. The ability to determine whether a disease is caused by Thl 7 or Th2 hyperreactivity is assessed by, for example, inhibiting the activity of a dopamine D1-like receptor and determining whether the disease has a preventive or therapeutic effect. Is possible. Agents (antagonists) that suppress the activity of dopamine D1-like receptors are known as described above. The target diseases in the present invention are particularly suitable for organ-specific autoimmune diseases such as rheumatoid arthritis, which is a chronic inflammatory disease, multiple sclerosis, type 1 diabetes, glomerulonephritis, and the like.

[0037] <Administration>

 The administration target of the drug can be appropriately selected according to the purpose without any particular limitation, and examples thereof include humans, mice, rats, mice, pigs, monkeys and the like.

[0038] Further, the administration method of the medicine is not particularly limited, depending on the dosage form of the medicine. For example, oral administration, administration by injection and the like can be mentioned.

[0039] The dosage of the medicament can be appropriately selected according to the age, weight, sex, symptom, etc. of the patient who is not subject to any particular restriction. The amount of the compound that modifies the activity of the donomin receptor (for example, dopamine D1-like receptor antagonist) is considered to be preferably about 2 to about! OOmg. Further, the frequency of administration of the medicament can be appropriately selected according to the purpose without any particular limitation. For example, the daily dose may be administered once a day, Multiple doses may be given.

 [0040] The administration timing of the medicament can also be appropriately selected according to the purpose for which there is no particular restriction. For example, it may be administered prophylactically before the onset of the disease, or the onset of the disease It may later be administered therapeutically.

 [0041] (Screening method)

 The screening method of the present invention is a method for screening a medicament for the treatment or prevention of a disease caused by the Th imbalance of the present invention, for example, a method using binding to a dopamine receptor as an index ( A first screening method) and a method using the modification of dopamine receptor activity as an index (second screening method).

 [0042] <First screening method (indicating binding)>

 The first screening method is not particularly limited as long as it is a method using the binding to the dopamine receptor as an index, and can be appropriately selected according to the purpose. For example, (a) Dopamine of a test substance And (b) selecting the test substance evaluated to have the ability to bind to the dopamine receptor in the step (a). Can be mentioned. The disease caused by Th imbalance targeted by this method is preferably a disease caused by excessive reaction of Thl 7 or Th2, and in this case, the dopamine receptor is preferably a donomin D1-like disease. It is a receptor. The test substance is not particularly limited. For example, the test substance can be appropriately selected according to the strength of the drug candidate substance and the purpose.

 [0043] (a) Evaluation process

In the evaluation step, the ability of the test substance to bind to the dopamine receptor The evaluation method can be appropriately selected according to the purpose for which there is no particular limitation, and examples thereof include a method based on a binding assay between a cell line expressing a donomin receptor protein and the test substance.

 [0044] In addition, when a result indicating that there is a difference in the degree of binding of the test substance to two types of cell lines that differ only in terms of dopamine receptor expression, the test substance is the dopamine receptor described above. It can be evaluated that it has binding ability to the body.

 [0045] One (b) One selection step

 In the selection step, the test substance evaluated as having the ability to bind to the dopamine receptor in the step (a) is selected.

 [0046] <Second screening method (indicating inhibition of activity)>

 The second screening method is not particularly limited as long as it is a method using the activity modification of the dopamine receptor as an index, and can be appropriately selected according to the purpose. For example, (a ′) dopamine of a test substance And (b ′) selecting the test substance evaluated to have the activity-modifying ability for the dopamine receptor in the step (a ′). Can be mentioned. The disease caused by Th imbalance to be the target of this method is preferably a disease caused by excessive reaction of TM7 or Th2, and in this case, activity modification of the donomin receptor used as a screening index Preferably, it is inhibition of dopamine D1-like receptor activity.

 [0047] One (a ') Evaluation process one

 In the evaluation step, the method for evaluating the activity-modifying ability of the test substance to the dopamine receptor can be appropriately selected according to the purpose without any particular limitation. For example, in the presence of the test substance, Examples include methods for examining changes in intracellular cAMP concentration, intracellular dopamine (DA) synthesis, intracellular donomin (DA) storage, and the like. The various changes can be examined using, for example, a conventionally known method.

[0048] In addition, screening was performed targeting dopamine D1-like receptors, and in the presence of the test substance, the intracellular cAMP concentration decreased and the intracellular DA synthesis amount decreased compared to the absence of the test substance. And / or if the result is that intracellular DA storage is reduced, the test substance is capable of inhibiting the activity of the dopamine D1-like receptor. Have the power to evaluate!

 [0049] One (b ') Selection process one

 In the selection step, the test substance evaluated as having an activity-modifying ability for the dopamine receptor in the step (a ′) is selected.

 [0050] As the screening method, only one of the first screening and the second screening may be performed, or both may be performed, but the drug can be efficiently selected. It is preferable to carry out both in terms of possible. In this case, the medicine can be selected more efficiently by performing the first screening and the second screening in this order.

 [0051] (drug)

 The drug of the present invention is a drug utilizing the new knowledge in the present invention that the action of dopamine is involved in the Thl / Th2 / Thl 7 differentiation induction mechanism mediated by dendritic cells. For example, the drug is dopamine D1-like Examples include drugs using receptor antagonists and drugs using donomin D2-like receptor antagonists.

 [0052] <Drugs using dopamine D1-like receptor antagonists>

 New findings by the present inventors indicate that a donomin D1-like receptor antagonist has a function of suppressing Thl 7 and Th2 differentiation of naive T cells or inducing Thl differentiation. It is also suggested that dopamine D1-like receptor antagonists may have a function of inhibiting dopamine synthesis or storage in dendritic cells (DC).

 [0053] Therefore, as the drug using the donomin D1-like receptor antagonist, for example, the drug of the present invention (first drug) that inhibits the synthesis or storage of dopamine in dendritic cells, which will be described later, and And the agent of the present invention (second agent) that suppresses the differentiation of naive T cells into Thl 7 or Th2 or promotes the differentiation into Thl.

 [0054] First drug, second drug

The first drug is a drug that inhibits the synthesis and storage of donomin in dendritic cells (DA synthesis or storage inhibitor in DC), and the second drug is directed to Thl 7 and Th2 of naive T cells. It is a drug that suppresses the differentiation of Thl or promotes the differentiation to Thl (Thl 7 or Th2 differentiation inhibitor or Thl differentiation promoter). The first drug and the second drug are Each of them contains the dopamine Dl-like receptor antagonist as an active ingredient and, if necessary, other ingredients.

[0055] The dominine D1-like receptor antagonist in the first drug and the second drug is not particularly limited, and can be appropriately selected according to the purpose. For example, the drug of the present invention Similarly, it can be appropriately selected.

[0056] The content of the donomin D1-like receptor antagonist in the first drug and the second drug is not particularly limited, and can be appropriately selected according to the purpose. The drug, the second drug, may be the donomin D1-like receptor antagonist itself.

 [0057] The type of the other component in the first drug and the second drug, the content of the other component, the dosage form, the production method, and the like are not particularly limited. For example, the present invention As with the above-mentioned pharmaceuticals, it can be appropriately selected.

 [0058] <Drugs using dopamine D2-like receptor antagonists>

 In addition, based on the new findings of the present inventors, donomin D2-like receptor antagonists inhibit the action of dopamine D2-like receptor in dendritic cells (DC), increase the cAMP concentration in DC, and dono in DC. It is shown to have a function of promoting the synthesis or storage of min. It is also shown that dopamine D2-like receptor antagonists have a function to induce Th2 differentiation of naive T cells.

 [0059] Accordingly, examples of the drug using the donomin D2-like receptor antagonist include, for example, the drug of the present invention (third drug) for promoting dopamine synthesis or storage in dendritic cells, which will be described later, and And the agent of the present invention (fourth agent) that promotes differentiation of naive T cells into Th2.

 [0060] Third drug, fourth drug

The third drug is a drug that promotes the synthesis and storage of donomin in dendritic cells (DA synthesis or storage promoter in DC), and the fourth drug is the differentiation of naive T cells into Th2. It is a drug that promotes (Th2 differentiation promoting agent). Each of the third drug and the fourth drug contains the dopamine D2-like receptor antagonist as an active ingredient, and other ingredients as required. [0061] The donomin D2-like receptor antagonist in the third drug and the fourth drug is not particularly limited, and can be appropriately selected according to the purpose. For example, the known dopamine D2-like receptor Antagonists may be used, and by using a screening method similar to the above-described screening method of the present invention, it is evaluated that it has the ability to bind to and / or inhibit the activity of the donomin D2-like receptor. You may also use other materials. Specific examples of the donomin D2-like receptor antagonist include sulpiride and nemonapride.

 [0062] The content of the dopamine D2-like receptor antagonist in the third drug and the fourth drug is not particularly limited and can be appropriately selected according to the purpose, and the third drug The fourth drug may be the donomin D2-like receptor antagonist itself.

 [0063] Further, the types of the other components in the third drug and the fourth drug, the content of the other components, the dosage form, the production method, and the like are not particularly limited. For example, the present invention As with the above-mentioned pharmaceuticals, it can be appropriately selected.

 [0064] <Application>

 The first to fourth agents are suitable as experimental reagents related to, for example, a Thl / Th2 / Thl7 differentiation induction mechanism via dendritic cells. In addition, the first to fourth drugs may be used for treatment or prevention of various immune-related diseases for which an improvement effect is expected by administration of the various drugs. For example, the fourth drug (a drug that promotes differentiation of naive T cells into Th2) can be used for the treatment or prevention of a disease caused by Thl hyperreactivity.

 Example

 Examples of the present invention are described below, but the present invention is not limited to these examples.

[0066] (Example 1: Effects of various drugs on Th balance of T cells mediated by dendritic cells)

In Example 1, the effect of the addition of various drugs, including dopamine D2-like receptor inhibitors (donomin D2-like receptor antagonists), on the Th balance in the process of dendritic cells inducing T cell differentiation. Examined. [0067] CD14-positive cells and CD45RA-positive (naïve) T cells were separated from human peripheral blood by MACS (magnetic cell separation method). The obtained CD14 positive cells were cultured in the presence of IL 4 and GM-CSF to induce differentiation into dendritic cells, and on the fifth day, stimulation with various drugs shown in FIGS. 1A to 1B was added, Collected on day 7. After washing, co-culture of the recovered dendritic cells and the naïve T cells was started, and restimulation with anti-CD3 and anti-CD28 antibodies on days 8 to 9 induced T cell differentiation did. Using the culture supernatant 16 hours after the restimulation, IFN-γ as a representative of Thl site force in and IL 5 as a representative of Th2 site force in were measured by ELISA. Th balance was confirmed by the production ratio of IFN-γ and IL-5. The results are shown in FIGS. 1A to 1B.

 [0068] FIG. 1A to FIG. 1B are graphs showing the values of IL 5 / IFN-γ when various drugs are added. A higher IL-5 / IFN-γ value indicates a Th2 differentiation bias, and a lower IL-5 / IFN-γ value indicates a Thl differentiation bias. Nemonapride and sulpiride are known dopamine D2-like receptor inhibitors. Forskolin is an activator of adenyl cyclase and is known to have the effect of increasing intracellular cAMP concentration. LPS is a lipopolysaccharide derived from Gram-negative bacteria and is known to have Thl differentiation-inducing action. Acetic acid was used as the vehicle. In addition, “none” in FIGS. 1A to 1B indicates a drug-free group (unless otherwise specified, the same applies to other figures).

 [0069] From the results shown in Fig. 1A, it was found that nemonapride and sulpiride, which are dopamine D2-like receptor inhibitors, and forskolin have Th2-inducing activity. Furthermore, the results in FIG. 1B revealed that sulpiride, a dopamine D2-like receptor inhibitor, has a concentration-dependent Th2-inducing activity.

 [0070] From the results of Example 1, the donomin D2-like receptor inhibitory power in dendritic cells (DC) was induced to differentiate into DC (DC2) having the ability to induce Th2 differentiation, and T cells were converted to Th2. Induction of differentiation, and DC itself has the ability to secrete donomin, and there is a mechanism that suppresses differentiation into DC2 by binding to autocrine to the donomin D2-like receptor. Was suggested.

[Example 2: Analysis of donomin receptor subtype in dendritic cells] In Example 2, expression of dopamine receptor subtypes in dendritic cells was analyzed by RT-PCR.

 [0072] There are five subtypes of dopamine receptors D1 to D5, which are known to couple with G proteins and send signals into cells. D1 and D5 are known as dopamine D1-like receptors, coupled to Gs proteins that increase adenyl cyclase activity and increase cAMP concentration, while D2-D4 are known as dopamine D2-like receptors. It is known to couple with Gi protein which suppresses adenyl cyclase activity.

 [0073] CD14-positive cells are cultured in the presence of IL-4 and GM-CSF for 5 days to induce differentiation into dendritic cells, RNA is recovered from the dendritic cells (DC), and RT-PCR is performed. went. For comparison, RNA was also collected from peripheral blood mononuclear cells (PMBC), naive T cells (naive T), and memory T cells (memory T), and RT-PCR was performed. RT-PCR used primers of various dopamine receptor subtypes such as Dl, D2, D3, D4, and D5. As a positive control, a primer for / 3-actin (beta-actin) was used. The result is shown in figure 2.

 [0074] From the results shown in Fig. 2, it was found that Dl, D4, and D5 receptors are expressed in DC. By comparing the results of Example 2 with the results of Example 1, it was suggested that dopamine D2-like receptor inhibitors may act on D4 in DC and cause Th2 bias in T cells. . In the previous report, naive CD4T cells express D3, memory CD4T cells express D2 and D3, and CD8 positive T cells express D3.

 [0075] (Example 3: Response of dendritic cells to dopamine)

In Example 3, the response of dendritic cells to dopamine stimulation (change in intracellular Ca ²⁺ concentration and change in intracellular cAMP concentration) was examined.

[0076] CD14-positive cells were cultured in the presence of IL4 and GM-CSF to induce differentiation into dendritic cells. The immature dendritic cells (iDC) on day 5 were treated with sulpiride or vehicle (acetic acid), and dopamine (DA) was added at various concentrations shown in FIGS. 3A to 3B. For each dendritic cell after dopamine stimulation, intracellular Ca ²⁺ concentration (FIG. 3A) and intracellular cAMP concentration (FIG. 3B) were measured.

[0077] From the results shown in FIGS. 3A and 3B, dopamine increased the Ca ²⁺ concentration in dendritic cells (DC). On the other hand, dopamine is D2-like (D4) in iDC because it reduces concentration of cAMP in a concentration-dependent manner and these are suppressed by inhibition of dopamine D2-like (D4) receptors by sulpiride. It was suggested that it binds predominantly to the receptor and sends a signal into the cell.

[0078] (Example 4: Effect of various drugs on site force-in production and surface antigen molecule expression in dendritic cells)

 In Example 4, the effects of various drugs on the production of cytodynamic force and surface antigen molecule expression in dendritic cells were examined.

[0079] CD14-positive cells are cultured in the presence of IL4 and GM-CSF to induce differentiation into dendritic cells, and are shown in FIG. 4A to FIG. 4B on immature dendritic cells (iDC) on day 5. Stimulation with various drugs was added. After 48 hours, the amount of IL-12p70 in the culture supernatant was measured by ELISA (Fig. 4A), and the expression of surface antigen molecules of CD80, CD86, CD83, and HLA-DR was measured by FACS (Fig. 4B). did.

 [0080] From the results shown in FIGS. 4A to 4B, dopamine (DA) and dopamine D2-like receptor inhibitors such as sulpiride have little effect on IL 12p70 site force-in production and surface antigen molecule expression in dendritic cells. Turned out not to give.

 (Example 5: Dopamine storage in dendritic cells)

 In Example 5, the state of dopamine storage in dendritic cells and the effect of various drugs on dopamine storage were examined.

 [0082] CD14-positive cells were cultured in the presence of IL 4 and GM-CSF to induce differentiation into dendritic cells. For immature dendritic cells (iDC) on the 5th day, forskolin 10 M It was recovered 48 hours after stimulation. After fixation with 10% formalin, ethanol treatment was performed, and staining was performed with an anti-dopamine antibody and a FITC-labeled secondary antibody. Furthermore, double staining was performed using a Lamp-1 antibody and a Texa s-Red labeled secondary antibody! / And observed with a confocal microscope (orgin al X 60) (FIG. 5A).

 [0083] In addition, the state of dopamine storage of DC by each stimulation of forskolin and sulpiride was observed with a fluorescence microscope and measured with a FACScan to determine each mean fluorescence intensity (MFI) (Fig. 5B).

Yes [0084] From the results of FIG. 5A to FIG. 5B, it was clarified that dopamine accumulates in Lamp-1 positive granules in DC, and that there is a difference in the amount of stored donomin depending on the type of stimulation. . Moreover, by comparing the results of Example 5 with the results of Example 3, it is suggested that dopamine D2-like receptor inhibitors such as sulpiride cause an increase in the cAMP concentration increasing capacity of dopamine in iDC. It was.

 [0085] (Example 6: Response of naive T cells to dopamine)

 In Example 6, the response of naive T cells to dopamine stimulation (intracellular cAMP concentration change) was examined.

 [0086] CD4-positive naive T cells were treated with IBMX (3-isobuthyl-1-methyl xanthine) lm M for 10 minutes, stimulated with each concentration of dopamine shown in Fig. 6A, and intracellular cAMP 10 minutes later When the concentration was measured, an increase in intracellular cAMP concentration dependent on dopamine concentration was observed (FIG. 6A).

 [0087] From the results in Fig. 6A, it was suggested that in naive T cells, dopamine mainly binds to D1-like receptors predominantly and sends signals into the cells.

[0088] To confirm the suggestion, CD4 positive naive T cells were treated with IBMXlmM for 10 minutes, and SCH23390 (dopamine D1-like receptor inhibitor) 1 μ μ or vehicle (acetic acid) was added. After incubation for 10 minutes, stimulation with donomin (DA) was performed, and the intracellular cAMP concentration after 10 minutes was measured. Dopamine (10_ ⁷ M) cAMP concentration increase by stimulation was completely inhibited by pretreatment with SCH23390 is Dono Min D1-like receptor inhibitors (Figure 6B).

 [0089] From the results shown in Fig. 6B, it was confirmed that dopamine induces an increase in cAMP concentration in naïve T cells by a signal via the D1-like receptor.

 [0090] In naive T cells, forskolin was stimulated upon stimulation with anti-CD3 and anti-CD28 antibodies.

 It was suggested that Cas S and dopamine, which are known to induce Th2 differentiation by the action of cAMP-elevating drugs such as (forskolin), have similar effects.

[0091] In order to confirm the suggestion, when stimulating the isolated human CD45RA positive cells with anti-CD3 antibody and anti-CD28 antibody, culturing by adding each concentration of dopamine (Dopamine)! /, Stimulation After 8 days, restimulation was performed with anti-CD3 antibody and anti-CD28 antibody. After 16 hours, T cells were collected, and CCR4 and CXCR3 expression was measured using FACScan (Fig. 6C, top 1), IL-5, IFN— γ secretion was measured by ELISA (Fig. 6C, bottom 2), and Thl / Th2 differentiation was examined. In the dopamine concentration as physiological concentrations (10_ ⁹ M), high production of IFN- γ, CC R4 / CXCR3 reduced such that the force Thl differentiation is induced, further, a high concentration of dopamine, IL- 5 It was revealed that Th2 differentiation is induced, such as high production of CCR4 / CXCR3 (Fig. 6C).

 [0092] From the results of FIGS. 6A to 6C, the amount of dopamine released from the dendritic cells when the dendritic cells interact with the naive T cells determines the direction of Th differentiation! / Force S suggested. In other words, it was suggested that when the amount of dopamine released by dendritic cells is small, it is biased toward Thl, and when it is large, Th2 is biased.

 [Example 7: Effect of dendritic cell degranulation on Th differentiation]

 In Example 7, the presence or absence of degranulation of dendritic cells during alio-MLR and the direction of Th differentiation of T cells when the amount of dopamine released from the dendritic cells was changed were examined.

 [0094] CD14-positive cells were cultured in the presence of IL4 and GM-CSF to induce differentiation into dendritic cells, and forskolin 10 μΜ was added to immature dendritic cells (iDC) on day 5. It was collected 48 hours after stimulation. After washing, the cells were suspended on a 35 mm glass culture dish (Glass Base Dish), and alio human CD45RA positive cells were added. When observed with a phase-contrast microscope immediately after the start of co-culture (allo-MLR) and taken over time (time-lapse), a part of DC appears to degranulate mainly after contact with T cells. Confirmed (Figure 7A).

 Next, alio-MLR was performed in the same manner as described above using iD C treated with colchicine (0.5 ng / ml) having a tubulin polymerization inhibitory effect, and anti-CD3 antibody and anti-CD28 anti-CD28 antibody were administered on the 8th day. Re-stimulated with the body. After 16 hours, the culture supernatant was collected, and IL 5 and IFN-γ were measured by ELISA. DC treated with colchicine was confirmed to have decreased IL 5 / IFN-γ, suggesting that inhibition of DC degranulation induces a Thl reaction (Fig. 7 Β).

[0096] In addition, DOPA is synthesized from tyrosine by tyrosine hydroxylase (ΤΗ) during the dopamine synthesis process, and this ΤΗ is synthesized by a-methyl-p-tyrosine (ΑΜΡΤ). It is known to be inhibited. Therefore, alio-MLR was performed in the same manner as described above using this iDC treated with AMPT. For DC treated with AMPT, IL-5 / IFN-γ is AMP It was confirmed that the concentration decreased in a T-dependent manner, suggesting that when the amount of donomin synthesized in DC was small V, a Thl reaction was induced (Fig. 7C).

 [0097] The results shown in FIGS. 7A to 7C suggested that the direction of Th differentiation of donomin released from DC during the interaction of DC naive T cells was determined.

 [0098] (Th differentiation induction mechanism through dendritic cells)

 The results of Examples 1 to 7 suggested a mechanism for inducing Th differentiation via dendritic cells, as schematically shown in FIG.

 [0099] The recognition of antigenic peptide 'MHC molecule by T cells involves adhesion as a secondary signal' costimulatory molecule, and SMAC (supermolecular consisting of receptor / ligand pair containing TCR / antigen peptide MHC complex) It is known that local interaction regions called activation clusters or immune synapses are formed.

 [0100] On the other hand, in the case of synapses in nerves, neurotransmitters such as dopamine are released from presynaptic neurons and activate not only post-neurons but also negative feedback via self-receptors expressed in pre-neurons. Is known to have the ability to regulate dopamine synthesis!

[0101] In the terminal part of presynaptic neurons, dopamine is oxidized to dihydroxy xyphenylalanine (DOPA) by tyrosine hydroxylase (TH), and further decarboxylates aromatic L amino acids. It is decarboxylated by an enzyme (aromatic L-amino acid decarboxylase; AAD) and stored in vesicles. This dopamine production and extracellular release process is regulated mainly by cAMP and Ca ²⁺ by stimulation via G protein. Stimulation from Gs protein leads to an increase in cAMP due to activation of adenyl cyclase (AC), which increases conversion to DOPA and increases DA storage, whereas stimulation from Gi protein causes intracellular Ca ²⁺ As a result of this increase, the secretion of DA storage vesicles is caused, and at the same time, the amount of DA stored in the cells is suppressed by suppressing AC activity. This regulation of dopamine levels from presynaptic neurons changes the response of postsynaptic receptor neurons.

[0102] From the results of Examples 1 to 7, such a mechanistic force at the neural synapse is also present in the interaction between dendritic cells (DC) and naive T cells, and in Th balance. It was suggested that this would have an impact (Figure 8).

 [0103] An immune synapse is formed between dendritic cells and T cells in the same way as the power and nerve synapses. Dendritic cells are considered to correspond to presynaptic neurons and T cells to postsynaptic receptor neurons. From the results of the above Examples;! To 7, in iDC, the adenyl cyclase (AC) activity via the Gs protein D1-like (D1, D5) receptor Gi protein, the D2-like (D4) receptor It was suggested that the amount of dopamine stored was (self) regulated by being suppressed through the body. Moreover, it was suggested that inhibition of D2-like (D4) receptors by D2-like receptor inhibitors such as sulpiride leads to a state of predominant AC activity in iDC, leading to an increase in the amount of dopamine stored in DC. On the contrary, it was suggested that inhibition of D1-like (Dl, D5) receptors by D1-like receptor inhibitors such as SCH23390 suppresses AC activity in iDC, leading to a decrease in dopamine storage in DC.

 [0104] Thus, when DCs with dopamine (DA) in the cell come into contact with naive T cells by antigen-specific interaction, degranulation of dopamine from DC is induced, and this dopamine is mainly received as a D1-like receptor. Naive T cells received in the body are expected to induce differentiation and activation biased to either T hi or Th2 depending on the amount of dopamine.

 [Example 8: Effect of dopamine receptor inhibitor in EAE model mice]

 Using SJL mice, EAE induction was started with PLP139- 151 + CFA + PTx (dayO), and as a prophylactic administration, during day3 to dayl8, L750667 (D2-like (D4) receptor inhibitor), SCH23390 (D1-like) (Dl, D5) receptor inhibitors) were orally administered every other day and evaluated by EAE clinical score. Each drug was dissolved in phosphate buffered saline (PBS) and administered at an amount of 0.3 mg / kg / day. In addition, only PBS was administered to the control group (control). The test was conducted with 2 animals in each group.

 [0106] As a result, both L750667 (D2-like (D4) receptor inhibitor) administration groups died on dayl4. On the other hand, in the SCH23390 (D1-like (D1, D5) receptor inhibitor) administration group, no onset was observed in any of the two animals (FIG. 9A).

 [0107] In addition, ELP induction was started with PLP139-151 + CFA + PTx (dayO), and all patients developed

Two days later (day 28), L750667 (D2-like (D4) receptor inhibitor) and S CH23390 (D1-like (D1, D5) receptor inhibitor) were orally administered every other day for therapeutic administration. EAE Evaluation was made with the clinical score (EAE clinical score).

[0108] [EAE clinical score]

 1; Tension decreased

 2; hindlimb paraparesis

 3; hindlimb paraplegia

 4; limb paralysis

 5; Drowning or death.

 [0109] Each drug was dissolved in phosphate buffered saline (PBS) and administered at an amount of 0.3 mg / kg / day. Moreover, only PBS was administered to the control group (control). The test was conducted with one animal in each group.

 [0110] Mice treated with L750667 (D2-like (D4) receptor inhibitor) deteriorated significantly from day 32 and died on day 34. On the other hand, the SCH23390 (D1-like (Dl, D5) receptor inhibitor) -administered mice improved their symptoms from day 38 compared to the two controls and reached remission on day 44 (Fig. 9B).

 [0111] Furthermore, in order to examine whether antigen-specific helper T cell responses were shifted to Thl, Th2, or Thl 7 by administration of each dopamine receptor inhibitor, the spleen cells of each mouse were examined. PLP139-151 was added again at the in vitro port, and IFN-γ (IFN-gamma) in the culture supernatant 48 hours later, IL-4 in the culture supernatant 24 hours later, 48 hours later IL-17 in the culture supernatant was measured by ELISA (FIGS. 9C to 9D: FIG. 9C is a mouse to which the drug was administered prophylactically, and FIG. 9D was a mouse to which the drug was therapeutically administered).

 [0112] As a result of Figure 9C, in the SCH23390 (D1-like (D1, D5) receptor inhibitor) administration group, IFN—

For IL-4, which is significantly higher than the control group, there was no significant difference from the control group, and because IL-17 was significantly lower than the control group, Thl differentiation was induced. On the contrary, it was confirmed that Thl 7 differentiation was suppressed. On the other hand, in the L750667 (D2-like (D4) receptor inhibitor) administration group, IL-4 was significantly higher than that in the control group, and IFN-γ was significantly lower than that in the control group. Since no significant difference was observed, it was confirmed that Th2 differentiation was induced and, on the contrary, Thl differentiation was suppressed. In addition, from the result of Fig. 9D, SCH23390 (D1-like (Dl, D5) receptor It was confirmed that Thl differentiation was induced and Thl 7 and Th2 differentiation was suppressed in the inhibitor group. In FIG. 9D, immunization (+) indicates a mouse in which EAE induction is performed, and im munizaition () indicates a mouse in which EAE induction is not performed.

 [0113] From the results of FIGS. 9A to 9D, L750667, a dopamine D2-like (D4) receptor inhibitor having Th2 inducing ability, worsened EAE. On the other hand, SCH23390, a D1-like (Dl, D5) receptor inhibitor with Thl-inducing ability (inhibition of Thl 7 and Th2 differentiation), is prophylactic and therapeutic! However, it was extremely high!

 [Example 9: Effect of donomin receptor inhibitor in collagen-induced arthritis (CIA) model mouse]

 Induction of CIA was started (dayO) in 7-week-old male mice (DB A / 1J) using sushi-derived type 2 collagen (200 Hg / animal) + CFA and boosted on day21 . On the other hand, as a prophylactic administration, SCH23390 (D1-like (D1, D5) receptor inhibitor) was orally administered three times a week between dayl8 and day34, and mouse spleen and serum were collected on day36. For comparison, L750667 (D2-like (D4) receptor inhibitor) was also examined. The CIA clinical score was evaluated for each limb according to the following criteria, and one mouse (total of 4 limbs) was evaluated with a maximum of 16 points.

[0115] [CIA Clinical Score]

 0; Normal

 1; swelling in one place

 2; Swelling of more than 2 powers and not severe

 3; Severe swelling of two or more power points

 4; Swelling in 2 or more places including fingertips, intense!

[0116] Each drug was dissolved in phosphate buffered saline (PBS) and administered in an amount to give 0.05 mg / kg / day. Moreover, only PBS was administered to the control group (control). The test was conducted with 8 animals in each group.

As a result, in the L750667 (D2-like (D4) receptor inhibitor) administration group, the onset of CIA was not suppressed, but in the SCH23390 (D1-like (D1, D5) receptor inhibitor) administration group, CIA This was suppressed (Fig. 10). [0118] (Example 10: Effect of a dopamine receptor inhibitor in a spontaneous mouse model of type 1 diabetes)

 As a prophylactic administration, NOD mice, a spontaneous model of type 1 diabetes, were orally administered SCH23390 twice a week from 4 weeks of age. For comparison, L750 667 (D2-like (D4) receptor inhibitor) was similarly examined. The drug was dissolved in phosphate buffered saline (PBS) and administered in an amount of 0.3 mg / kg / day. Thereafter, urinary glucose was monitored from the age of 14 weeks, and the incidence of diabetes in each group (n = 16) was compared. As a result, the onset of diabetes was suppressed in the SCH23390 administration group (FIG. 11).

 (Example 11: Effect of a donomin receptor inhibitor in glomerulonephritis model mice) As a pre-immunization, 5 &#12316; 6-week-old SJL male mice (12 mice) were used as a CFA suspension. Hid IgGO. 5 mg / 20 g body weight was injected subcutaneously. As a prophylactic administration, SCH23390 (D1-like (D1, D5) receptor inhibitor) was orally administered three times a week with the day of preimmunization as the first day. The drug was dissolved in phosphate buffered saline (PBS) and administered at an amount of 0.3 mg / kg / day. Only the PBS was administered to the positive control (PC) group and the negative control (NC) group. The test was conducted with 4 animals in each group.

[0120] Next, an anti-basement membrane antibody was administered to the SCH23390 administration group and the PC group. Pre-immune mouse glomerular basement membrane antiserum (Hedge) 15

Body weight was intravenously administered via the tail vein for 3 days. PBS was intravenously injected into the NC group of mice. Thereafter, the patient was decapitated on the 14th day, and blood, spleen and kidney were collected.

 [0121] As a result, in the PC group, all-round cellular meniscus was formed in 80% of the glomeruli, and advanced necrosis of glomerular capillaries and cell infiltration into the periglomerular stroma were advanced. Inflammatory changes were observed (Figure 12A). These inflammatory changes were not observed at all in the NC group (FIG. 12B). In the SCH23390 administration group, 20% of the glomeruli showed capillary necrosis and meniscus formation, but both were minor and advanced such as diffuse changes occupying the entire glomerulus and cell infiltration into the surrounding stroma. Inflammatory findings were not observed (Figure 12C).

[0122] As described above, the results of Examples 8 to 11 include organ-specific autoimmunity such as multiple sclerosis and glomerulonephritis, including diseases conventionally considered to be diseases caused by Thl excess reaction. Rheumatoid arthritis S, which is a disease and chronic inflammatory disease. Can be effectively treated or prevented by actively inducing Thl (or suppressing Thl 7 or Th2 differentiation) by administering a Dl-like (Dl, D5) receptor inhibitor It indicates that it is a disease. In this example, the details of the mechanism by which SCH23390 (D1-like (Dl, D5) receptor inhibitor) showed an extremely high effect are unknown, but a decrease in IL 17 or an increase in IFN γ It is highly probable that they acted defensively.

Industrial applicability

 The medicament of the present invention is useful as a therapeutic or prophylactic agent for diseases caused by Thl 7 or Th2 excessive reaction, and the screening method of the present invention is useful as a method for screening the medicament. Further, the various drugs of the present invention are useful as experimental reagents related to the Thl / Th2 / Thl 7 differentiation induction mechanism mediated by dendritic cells, and an improvement effect is expected by administration of the various drugs. It is also useful as a therapeutic or prophylactic agent for various diseases

Claims

The scope of the claims

 [I] A medicament for treating or preventing a disease caused by Th imbalance, comprising a compound that modifies the activity of a dopamine receptor as an active ingredient.

 [2] The disease caused by Th imbalance is caused by an excessive reaction of Thl 7 or Th2.

The medicament according to claim 1, wherein the compound that modifies the activity of the dopamine receptor is a dopamine D1-like receptor antagonist.

[3] The medicament according to claim 2, wherein the dopamine D1-like receptor antagonist is SCH23390.

[4] A screening method for a drug for the treatment or prevention of a disease caused by Th imbalance, wherein the screening method uses binding to a donomin receptor as an index.

[5] The disease caused by Th imbalance is caused by an excessive reaction of Thl 7 or Th2.

5. The screening method according to claim 4, wherein the dopamine receptor is a dopamine D1-like receptor.

 [6] A screening method for a drug for treating or preventing a disease caused by Th imbalance, wherein the screening method uses modification of activity of a donomin receptor as an index.

[7] The disease caused by Th imbalance is caused by an excessive reaction of Thl 7 or Th2.

7. The screening method according to claim 6, wherein the modification of the activity of the dopamine receptor is inhibition of the activity of the dopamine D1-like receptor.

[8] A drug that inhibits the synthesis or storage of donomin in dendritic cells, which is dopamine D

A drug containing a 1-like receptor antagonist as an active ingredient.

[9] A drug that suppresses differentiation of naive T cells into Thl 7 or Th2, or promotes differentiation into Thl, and contains a donomin D1-like receptor antagonist as an active ingredient.

 [10] The agent according to claim 8 or 9, wherein the dopamine D1-like receptor antagonist is SCH23390.

 [I I] A drug that promotes the synthesis or storage of donomin in dendritic cells and contains a dopamine D2-like receptor antagonist as an active ingredient.

 [12] A drug that promotes differentiation of naive T cells into Th2, and contains a dopamine D2-like receptor antagonist as an active ingredient.