JP2008255008A - Synoviocyte proliferation inhibitor - Google Patents

Abstract

<P>PROBLEM TO BE SOLVED: To provide a synoviocyte proliferation inhibitor which can inhibit the proliferation of synoviocytes, without causing immunosuppression, to provide a medicinal composition for treating diseases in which the proliferation of synoviocytes participates, and to provide a therapeutic method therefor. <P>SOLUTION: This synoviocyte proliferation inhibitor is characterized by containing one of flavopiridol which is a compound represented by structural formula (1), and a pharmaceutically acceptable salt thereof as an active ingredient. The medicinal composition is characterized by comprising the synoviocyte proliferation inhibitor. <P>COPYRIGHT: (C)2009,JPO&INPIT

Description

  The present invention relates to a synovial cell proliferation inhibitor comprising as an active ingredient any one of flavopiridol and pharmaceutically acceptable salts thereof, a pharmaceutical composition comprising the synovial cell proliferation inhibitor, and The present invention relates to a treatment method using a synovial cell proliferation inhibitor.

The synovial membrane is originally composed of a single layer of synovial cells, but the synovial cells may proliferate excessively due to some inflammatory mechanism, resulting in synovial hyperplasia.
Synovial cells are activated in the increased synovium, and various inflammatory mediators and tissue destructive proteolytic enzymes are produced from the synovial cells. This series of inflammatory reactions is referred to as proliferative synovitis, and the proliferative synovitis results in serious dysfunction such as cartilage degeneration and bone tissue destruction. Examples of diseases in which such synovial cell proliferation is involved in the pathology include rheumatoid arthritis, osteoarthritis, psoriatic arthritis, and gout.

  Rheumatoid arthritis is an autoimmune disease, and inflammatory mediators (eg, IL-1, IL-6, TNF-α, etc.) from lymphocytes and the like are excessively present in the vicinity of joints, resulting in tissue inflammation. It is a disease where bone destruction progresses and pathology appears. One of the etiologies is the proliferation of the synovial cells, but it is thought that there are multiple etiologies, so a multifaceted treatment method is required. Development is desired.

Conventionally, as a treatment method for rheumatoid arthritis, symptomatic treatment by the administration of analgesics and anti-inflammatory agents has been carried out. However, symptomatic treatment can delay the onset, but stops the progression of symptoms. Moreover, it was impossible to cure the disease itself.
On the other hand, based on the fact that some of the drugs originally used as anticancer agents in recent years are also effective as lymphocyte proliferation inhibitors, by administering them, anti-TNF-α antibodies are further used in combination. It has been reported that the treatment cures inflammation, and in some cases, the progression of the disease stops and the improvement rate and cure rate of the disease increase.
However, the administration of these drugs has a high therapeutic effect, but also has a high immunosuppressive effect due to suppression of lymphocyte proliferation, so that the administered patient is always in an immunosuppressed state and is extremely vulnerable to infection. is there.

  In addition to the anticancer drug and anti-TNF-α antibody, there are a plurality of drugs for treating rheumatoid arthritis by suppressing inflammation. For example, even if it is very effective for a specific patient, it is effective for other patients. There are many drugs that are low, and a drug that is highly versatile and has a complete healing effect is not yet known.

On the other hand, as one of the therapeutic agents for rheumatoid arthritis, a drug (synovial cell growth inhibitor) that suppresses the proliferation of the synovial cells, which is one of its etiology, has been proposed.
Examples of the synovial cell growth inhibitor include diterpene compounds (see Patent Document 1) and FADD genes (see Patent Document 2) that suppress synovial cell proliferation by inducing apoptosis of synovial cells. , Fas ligand (see Patent Document 3), anti-Fas antibody (see Patent Document 4), 3,6-anhydrogalactose and its derivatives (see Patent Document 5), etc. As inhibitors for the proliferation of membrane cells, imidazole derivatives (see Patent Document 6), quinoline derivatives, quinolone derivatives (see Patent Document 7) and the like have been proposed. In addition to these, as the synovial cell proliferation inhibitor, an association product of hyaluronic acid and zinc (see Patent Document 8), a quinolonecarboxylic acid derivative (see Patent Document 9), and a substance in milk whey protein (Patent Document 10) Reference), bFGF antagonist (see patent document 11), IL-6 antagonist (see patent document 12), cAMP derivative (see patent document 13), vitamin D3 and its derivative (see patent document 14), c-erbB-2 activity Inhibitors (see Patent Document 15), γ-secretase-like protease inhibitors (Patent Document 16), and the like have been proposed.

These synovial cell proliferation inhibitors are not only for rheumatoid arthritis but also for preventing the onset, reducing the symptoms, and increasing the severity of diseases involving the proliferation of synovial cells such as osteoarthritis, psoriatic arthritis, and gout. Expected to be a useful inhibitor.
However, without inhibiting lymphocyte proliferation and activation, that is, without causing immunosuppression, a synovial cell proliferation inhibitor that can inhibit synovial cell proliferation, without reducing resistance to infection At present, a medicinal composition and a treatment method for treating a disease involving synovial cell proliferation have not yet been found.

JP 2004-168713 A JP 2000-198737 A Japanese Patent Laid-Open No. 11-060501 JP-A-08-208515 WO99 / 064424 JP 2003-040888 A Japanese Patent Laid-Open No. 2002-371078 JP 2003-089647 A JP 2002-293745 A JP 2001-011094 A JP 2000-229883 A JP-A-08-208514 Japanese Unexamined Patent Publication No. 07-324035 Japanese Unexamined Patent Publication No. 07-145042 WO01 / 076630 WO01 / 073211 publication

  An object of the present invention is to solve the conventional problems in response to the above-mentioned demands and achieve the following object. That is, the present invention relates to a synovial cell proliferation inhibitor capable of suppressing the proliferation of synovial cells without inhibiting lymphocyte proliferation and activation, that is, without causing immunosuppression, and resistance to infection. An object of the present invention is to provide a medicinal composition and a treatment method for treating a disease associated with synovial cell proliferation without lowering the pH.

As a result of intensive studies by the present inventors in order to solve the above problems, the following knowledge has been obtained. That is, it is a finding that flavopiridol exhibits a strong inhibitory activity on synovial cell proliferation, but does not affect the activity of lymphocytes.
Flavopyridol is a novel flavonoid synthesized with hints of a component extracted from the bark of the mahogany family, Dysoxylum biniferiferum , as a flavonoid that induces apoptosis of cancer cells. Activity, smooth muscle growth inhibitory activity and the like are known.
However, it is not known at all that flavopiridol suppresses the proliferation of synovial cells without suppressing the activation and proliferation of lymphocytes, which is a new finding of the present inventors.

This invention is based on the said knowledge by the present inventors, and the means for solving the said subject are as follows. That is,
<1> Synovial cell proliferation inhibition characterized by containing as an active ingredient flavopiridol, which is a compound represented by the following structural formula (1), and a pharmaceutically acceptable salt thereof It is an agent.

<2> A medicinal composition comprising at least the synovial cell proliferation inhibitor according to <1>.
<3> The medicinal composition according to <2>, including a pharmaceutically acceptable carrier.
<4> The medicinal composition according to any one of <2> to <3>, wherein the composition is at least one of an anti-rheumatoid arthritis agent, an anti-osteoarthritis agent, an anti-psoriatic arthritis agent, and an anti-gout agent. .
<5> The medicinal composition according to any one of <2> to <3>, which is used for the treatment of at least one of rheumatoid arthritis, osteoarthritis, anti-psoriatic arthritis, and gout.

<6> A method for treating rheumatoid arthritis, osteoarthritis, anti-psoriatic arthritis, and gout by administering at least the synovial cell proliferation inhibitor according to <1> above It is.
<7> The above <6>, wherein the synovial cell proliferation inhibitor according to <2> to <5> is administered to treat at least one of rheumatoid arthritis, osteoarthritis, anti-psoriatic arthritis, and gout. It is the treatment method as described in above.

  According to the present invention, conventional problems can be solved, and synovial cell proliferation that can inhibit synoviocyte proliferation without inhibiting lymphocyte proliferation or activation, that is, without causing immunosuppression. It is possible to provide an inhibitor and a medicinal composition and a treatment method for treating a disease involving synovial cell proliferation without reducing resistance to infection.

(Endotoxin activity inhibitor)
The synovial cell proliferation inhibitor of the present invention contains, as an active ingredient, flavopiridol, which is a compound represented by the following structural formula (1), and a pharmaceutically acceptable salt thereof.

  The salt is not particularly limited and may be appropriately selected depending on the intended purpose. Examples thereof include inorganic salts and organic salts. Specific examples include hydrochloride, hydrobromide, sulfuric acid, and the like. Salt, phosphate, acetate, oxalate, tartrate, citrate, maleate, fumarate and the like are preferable.

  There is no restriction | limiting in particular as a manufacturing method of flavopiridol, According to the objective, it can select suitably from a well-known method. As flavopiridol, a commercially available product may be used.

The synovial cell proliferation inhibitor has an action of suppressing the proliferation of synovial cells and suppressing proliferative synovitis without suppressing the proliferation and activation of lymphocytes, and as a result, is accompanied by immunosuppression. Without preventing or preventing cartilage degeneration or bone tissue destruction.
This ability to suppress synovial cell proliferation without immunosuppression can be achieved, for example, by transferring sera from mice that spontaneously develop arthritis (eg, KxB / N mice) to lymphocyte-deficient mice (eg, RAG gene-deficient mice). To evaluate the severity of inflammation between the group of mice administered with the synovial cell proliferation inhibitor of the present invention and the group of mice not administered with the model mice that have developed synovial hyperplasia. Can do.
In the evaluation, for example, since dose-dependent suppression of inflammation is observed in the group administered with the synovial cell proliferation inhibitor of the present invention, the action mechanism of the synovial cell proliferation inhibitor of the present invention is It can be seen that this is not due to suppression of lymphocyte proliferation.

The synovial cell proliferation inhibitor of the present invention has low cytotoxicity and high safety. In addition, since the molecular weight is small, for example, when administered to a living body, it is easy to move to a target site where synovial hyperplasia occurs, which is advantageous.
Moreover, the synovial cell proliferation inhibitor of the present invention is suitable as an active ingredient of the medicinal composition of the present invention described below, and is suitably used for the treatment method of the present invention described below.

(Medicinal composition)
The medicinal composition of the present invention contains at least the synovial cell proliferation inhibitor of the present invention and, if necessary, contains other components appropriately selected from known ones.
Examples of the other components include a pharmaceutically acceptable carrier, and the carrier is not particularly limited and may be appropriately selected depending on the dosage form of the medicinal composition.

  There is no restriction | limiting in particular as a dosage form of the said medicinal composition, According to an administration method, it can select suitably, For example, an oral solid agent (a tablet, a coated tablet, a granule, a powder, a capsule etc.), an oral liquid agent (Internal solutions, syrups, elixirs, etc.), injections (solutions, suspensions, solid preparations for erection, etc.), suppositories, ointments, patches, gels, creams, external powders, sprays, And inhaled powders.

As the oral solid preparation, for example, excipients, if necessary, additives such as binders, disintegrants, lubricants, coloring agents, flavoring / flavoring agents, etc. are added to the synovial cell proliferation inhibitor, It can be manufactured by the method.
Examples of the excipient include lactose, sucrose, sodium chloride, glucose, starch, calcium carbonate, kaolin, microcrystalline cellulose, silicic acid and the like.
The additive is not particularly limited and may be appropriately selected depending on the intended purpose. Examples of the binder include water, ethanol, propanol, simple syrup, glucose solution, starch solution, gelatin solution, carboxy Examples include methyl cellulose, hydroxypropyl cellulose, hydroxypropyl starch, methyl cellulose, ethyl cellulose, shellac, calcium phosphate, polyvinyl pyrrolidone, etc. Examples of the disintegrant include dry starch, sodium alginate, agar powder, sodium bicarbonate, calcium carbonate, lauryl. Examples of the lubricant include sodium sulfate, stearic acid monoglyceride, and lactose. Examples of the lubricant include purified talc, stearate, borax, polyethylene glycol, and the like. , Titanium oxide, iron oxide and the like, as the flavoring agent include sucrose, bitter orange peel, citric acid, and tartaric acid.

The oral solution can be produced by a conventional method, for example, by adding additives such as a taste-masking / flavoring agent, a buffer, and a stabilizer to the synovial cell growth inhibitor.
The additive is not particularly limited and may be appropriately selected depending on the intended purpose. Examples of the flavoring / flavoring agent include sucrose, orange peel, citric acid, tartaric acid, and the like. Examples of the stabilizer include sodium citrate, and examples of the stabilizer include tragacanth, gum arabic, and gelatin.

As the injection, for example, a pH regulator, a buffer, a stabilizer, an isotonic agent, a local anesthetic, and the like are added to the synovial cell proliferation inhibitor, and subcutaneous, intramuscular and intravenous in a conventional manner. An internal injection can be produced.
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, thiolactic acid, and the like. Examples of the isotonic agent include sodium chloride and glucose. Examples of the local anesthetic include procaine hydrochloride and lidocaine hydrochloride.

  Examples of the suppository include the above-mentioned synovial cell growth inhibitor, a known carrier for suppository preparation, such as polyethylene glycol, lanolin, cocoa butter, fatty acid triglyceride and the like, and if necessary, Tween (registered trademark). Can be produced by a conventional method.

  The ointment can be produced by blending a known base, stabilizer, wetting agent, preservative and the like with the synovial cell proliferation inhibitor and mixing them by a conventional method. Examples of the base include liquid paraffin, white petrolatum, white beeswax, octyldodecyl alcohol, paraffin and the like. Examples of the preservative include methyl paraoxybenzoate, ethyl paraoxybenzoate, and propyl paraoxybenzoate.

  As the patch, for example, a cream, gel or paste as the ointment can be applied to a known support by a conventional method. Examples of the support include cotton, suf, woven fabric made of chemical fiber, non-woven fabric, soft vinyl chloride, polyethylene, polyurethane film, and foam sheet.

The content of the synovial cell proliferation inhibitor in the pharmaceutical composition can be appropriately selected depending on the dosage form and the degree of symptoms of the patient who is the subject of administration. About 1-1000 mg is preferable, about 0.1-500 mg is preferable for the injection, and about 5-1000 mg is preferable for the suppository.
The dosage of the pharmaceutical composition can be appropriately selected according to the patient's symptoms, body weight, age, sex, etc., but usually about 0.1 to 5000 mg per day for an adult is preferable, 1000 mg is more preferable.
As for the administration frequency of the pharmaceutical composition, it is preferable to administer the above-mentioned daily dose divided into 1 to about 2 to 4 times.

  In addition, the medicinal composition of the present invention is suitable as a therapeutic or prophylactic agent for diseases in which the proliferation of synovial cells is involved in its pathology, such as anti-rheumatoid arthritis agents, anti-osteoarthritis agents, anti-psoriatic properties Suitable as arthritis agent and anti-gout agent.

(Method of treatment)
The treatment method of the present invention is to treat at least the synovial cell proliferation inhibitor, treat at least one of rheumatoid arthritis, osteoarthritis, anti-psoriatic arthritis, and gout, and administer the pharmaceutical composition. It is preferable.

  In the treatment method, the administration method of the synovial cell proliferation inhibitor or the pharmaceutical composition is not particularly limited and may be appropriately selected depending on the dosage form and the like.

  In addition, the treatment methods include rheumatoid arthritis, osteoarthritis, psoriatic arthritis, and gout, and other known anti-rheumatoid arthritis agents, anti-osteoarthritis agents, anti-psoriatic arthritis agents and It can also be used in combination with gout agents.

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

Example 1
<Inhibition of mouse collagen-induced arthritis>
Collagen-induced arthritis in mice is a disease in which synovitis with proliferation of synovial cells occurs as a result of autoimmunity against type II collagen, and the joint is destroyed like rheumatoid arthritis. A good model. The arthritis model mouse was prepared by the following method, and the suppression of arthritis by the synovial cell proliferation inhibitor of the present invention was evaluated.

Arthritis was induced by subcutaneously injecting 100 μL of the following adjuvant into the tail of a 7-week-old DBA / 1J mouse (male), and booster immunization was performed 21 days later to obtain an arthritis model mouse.
The adjuvant was prepared by mixing equal amounts of 4 mg / mL collagen solution prepared by dissolving bovine type II collagen (manufactured by Collagen Technology Training Society) in 0.05N acetic acid aqueous solution and complete adjuvant Freund (manufactured by Difco).

  The arthritis model mouse was intraperitoneally administered with flavopiridol dissolved in DMSO as the synovial cell proliferation inhibitor for 10 consecutive days from 4 days after booster immunization. The administration group was divided into two groups, and the dose of flavopiridol was 1 mg / kg or 2.5 mg / kg, respectively. In addition, as a comparison target group, DMSO alone was administered in an equivalent amount peritoneally.

The degree of arthritis onset in the arthritis model mice was evaluated by the arthritic index. Evaluation was performed according to the following criteria, with 4 points or less per limb and 16 points or less per individual. The results are shown in FIG.
[Evaluation criteria]
1.0: Swelling is observed at one location of the joint.
2.0: Weak swelling is observed at two or more locations of the joint.
3.0: Severe swelling is observed on the back of limbs, but not all fingers.
4.0: Severe swelling is observed on the back and fingers of the limbs.

(Example 2)
The arthritis model mice prepared in the same manner as in Example 1 were subjected to 3 days after booster immunization (24 days after the first immunization), twice a week for 5 weeks (28, 34, 38 from the first immunization, 42, 45, 48, 52, 55, 59, and 62 days later) As the synovial cell proliferation inhibitor, flavopiridol dissolved in DMSO was administered intraperitoneally. The administration group was divided into 3 groups, and the doses of flavopiridol were 1 mg / kg, 2.5 mg / kg, and 5.0 mg / kg, respectively. In addition, as a comparison target group, DMSO alone was administered in an equivalent amount peritoneally. The results of evaluating the degree of arthritis development in the same manner as above are shown in FIG.

  From the results of FIG. 1 and FIG. 2, the degree of arthritis is improved in the mouse administered with the synovial cell proliferation inhibitor of the present invention, compared to the mouse administered only with DMSO not containing the synovial cell proliferation inhibitor. The degree of improvement was found to be dependent on the dose of flavopiridol.

(Example 3)
<Inhibition of arthritis in lymphocyte-deficient mice>
Mice lacking the RAG gene have no immune response due to lymphocytes because they lack lymphocytes, but they receive the serum (KxB / N serum) of a model mouse (KxB / N) that spontaneously develops arthritis very similar to rheumatoid arthritis Then, it is known that the proliferation of synovial cells is promoted and arthritis is caused. Using this model mouse, inhibition of arthritis by the synovial cell proliferation inhibitor of the present invention was evaluated.

An arthritis model was prepared by intraperitoneally administering 300 μL of KxB / N mouse serum to RAG gene-deficient mice (9-10 weeks old, RAG2 / B6 KO, C57BL / 6 genetic background).
One day after the transfer of serum, 10 days after continuous administration, flavopiridol dissolved in DMSO was administered intraperitoneally as the synoviocyte growth inhibitor. The administration group was divided into two groups, and the dose of flavopiridol was 1 mg / kg or 2.5 mg / kg, respectively. In addition, as a comparison target group, DMSO alone was administered in an equivalent amount peritoneally.
The degree of arthritis onset in the arthritis model mice was evaluated in the same manner as in Example 1. The results are shown in FIG.

From the results shown in FIG. 3, the degree of arthritis was improved in the mice administered with the synovial cell proliferation inhibitor of the present invention compared to the mice administered with DMSO alone, which did not contain the synovial cell proliferation inhibitor. It was found that the degree of the flavopiridol improved in a dose-dependent manner.
In addition, this revealed that suppression of proliferative arthritis was not due to suppression of immune reactions such as suppression of lymphocyte activation and proliferation suppression of the synovial cell proliferation inhibitor of the present invention. .

Example 4
<Antibody response in collagen-induced arthritis in mice>
The model mouse in which arthritis was induced in the same manner as in Example 1 was administered with the above synovial cell proliferation inhibitor in the same manner as in Example 1, and the administration start date (0 day, immediately before administration), 11 days after the administration. On the 39th day after administration, the antibody responses of the mouse subclasses (IgG1, IgG2a, IgG2b) were measured by the following method. The results are shown in FIGS.

  The antibody response was measured by reacting a 96-well plate coated with bovine type II collagen with a diluted solution of serum collected from a mouse, washing the 96-well plate, and then peroxidase-conjugated anti-mouse IgG1, IgG2a or IgG2b antibody (manufactured by Zymed) was added and reacted, and these were colored with a TMBZ (3,3 ′, 5,5′-tetramethylbenzidine) substrate, and the absorbance at 450 nm wavelength was measured.

  From the results of FIGS. 4 to 6, no significant difference was observed in the antibody response of the subclass. From this result, it was found that the synovial cell proliferation inhibitor of the present invention can strongly suppress proliferative synovitis without suppressing the antibody response by lymphocytes.

  The synovial cell proliferation inhibitor comprising flavopiridol as an active ingredient and the medicinal composition comprising the synovial cell proliferation inhibitor as an active ingredient of the present invention are associated with the pathological condition of synovial cell proliferation by administration. Since the disease can be prevented or treated without suppressing immunity, it is suitable as an anti-rheumatoid arthritis agent, anti-osteoarthritis agent, anti-psoriatic arthritis agent and anti-gout agent. In addition, by administering the synovial cell proliferation inhibitor and the medicinal composition, it is possible to prevent or treat a disease in which synovial cell proliferation is involved in a disease state without suppressing immunity.

FIG. 1 is a graph showing the results of Example 1. FIG. 2 is a graph showing the results of Example 2. FIG. 3 is a graph showing the results of Example 3. FIG. 4 is a graph showing the antibody response results of the IgG1 subclass of Example 4. FIG. 5 is a graph showing the antibody response results of the IgG2a subclass of Example 4. FIG. 6 is a graph showing the results of antibody responses of the IgG2b subclass of Example 4.

Claims (5)

A synovial cell proliferation inhibitor comprising as an active ingredient any one of flavopiridol, which is a compound represented by the following structural formula (1), and a pharmaceutically acceptable salt thereof.

  A medicinal composition comprising at least the synovial cell proliferation inhibitor according to claim 1.   The medicinal composition according to claim 2, which is at least one of an anti-rheumatoid arthritis agent, an anti-osteoarthritis agent, an anti-psoriatic arthritis agent, and an anti-gout agent.   The medicinal composition according to claim 2, which is used for treating at least one of rheumatoid arthritis, osteoarthritis, anti-psoriatic arthritis, and gout. A method for treating rheumatoid arthritis, osteoarthritis, anti-psoriatic arthritis, and gout by administering at least the synovial cell proliferation inhibitor according to claim 1.

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