Pharmaceutical composition for preventing or treating interstitial cystitis / bladder pain syndrome

Abstract

To provide a pharmaceutical composition for preventing or treating interstitial cystitis / bladder pain syndrome.SOLUTION: A pharmaceutical composition contains an inhibitor against at least one integrin selected from the group consisting of integrin αvβ3, integrin αvβ5, and integrin α5β1. The present invention also provides a method for evaluating the effect of a substance being tested on interstitial cystitis / bladder pain syndrome, the method including using, as an index, an inhibitory activity against at least one integrin selected from the group consisting of αvβ3, αvβ5, and α5β1.SELECTED DRAWING: Figure 1

Description

Technical Field

[0001] The present invention relates to a pharmaceutical composition for preventing or treating interstitial cystitis / bladder pain syndrome, which contains an RGD integrin inhibitor.

Background Art

[0002] Interstitial Cystitis / Bladder Pain Syndrome (IC / BPS) is defined as "a state in which there is chronic pelvic pain, pressure, or discomfort related to the bladder, accompanied by lower urinary tract symptoms such as urinary urgency and frequency, and there is no confounding disease" (Non-Patent Document 1). Among them, IC / BPS with ulcerative lesions (Hunner lesions) is called Hunner type interstitial cystitis. Hunner type interstitial cystitis has clear abnormal findings both endoscopically and pathologically, and is considered to be more severe symptomatically, and has been designated as a refractory disease as Specified Refractory Disease 226 by the Ministry of Health, Labour and Welfare.

[0003] There is still no established method for the treatment of IC / BPS, and the current situation is that it remains at symptomatic treatment. As surgical procedures, hydrodistension of the bladder using an endoscope is performed, and when Hunner lesions are recognized in the bladder, cauterization using electricity or a laser is also performed simultaneously. In addition, as pharmaceutical treatment, in addition to oral treatment with centrally acting drugs such as amitriptyline and immunosuppressive agents such as cyclosporine A, intravesical instillation of drugs such as heparin, DMSO, and steroids, or injection of botulinum toxin into the bladder wall is performed.

[0004] Currently, although symptoms are relieved in about half of the cases by hydrodistension of the bladder or cauterization of Hunner lesions, long-term remission is limited to some cases. In addition, pharmaceutical treatment has not led to the cure of IC / BPS. Thus, IC / BPS is a refractory QOL disease that requires long-term medical management by repeating relapses and remissions, and the development of effective therapeutic drugs is desired.

Prior Art Documents

Non-Patent Documents

[0005]

Non-Patent Document 1

Disclosure of the Invention

Problems to be Solved by the Invention

[0006] An object of the present invention is to provide a new treatment means for IC / BPS.

Means for Solving the Problems

[0007] The present inventors have found that RGD integrin inhibitors are useful for the treatment of IC / BPS, particularly for improving frequent urination and pain, and have completed the following invention.

[0008] Item 1. A pharmaceutical composition for preventing or treating interstitial cystitis / bladder pain syndrome, containing an inhibitor against at least one integrin selected from the group consisting of integrin αvβ3, integrin αvβ5, and integrin α5β1. Item 2. The pharmaceutical composition according to Item 1, containing an inhibitor against integrin αvβ3. Item 3. The pharmaceutical composition according to Item 1 or 2, wherein the inhibitor against integrin is an RGD mimic. Item 4. The pharmaceutical composition according to any one of Items 1 to 3, wherein the inhibitor against integrin is a peptide or a syringotide consisting of the amino acid sequence shown in SEQ ID NO: 1. Item 5. The pharmaceutical composition according to any one of Items 1 to 4, for improving frequent urination or pain in interstitial cystitis / bladder pain syndrome. Item 6. The pharmaceutical composition according to any one of Items 1 to 5, for intravesical instillation. Item 7. The pharmaceutical composition according to any one of Items 1 to 6, further containing another drug for preventing or treating interstitial cystitis / bladder pain syndrome. Item 8. The pharmaceutical composition according to Item 7, wherein the other drug is heparin, DMSO, or a steroid. A method for evaluating the effect of a test substance on interstitial cystitis / bladder pain syndrome, using the inhibitory activity against at least one integrin selected from the group consisting of ανβ3, ανβ5, and α5β1 as an index.

Advantages of the Invention

[0009] According to the present invention, it becomes possible to prevent or treat IC / BPS based on a new mechanism of action.

Brief Description of the Drawings

[0010]

Figure 1

Figure 2

Figure 3

Figure 4

Embodiments for Carrying Out the Invention

[0011] The description of the present invention shown below may be based on representative embodiments or specific examples, but the present invention is not limited to such embodiments or specific examples. Also, the upper and lower limit values of each numerical range shown in this specification can be arbitrarily combined. Further, a numerical range represented by "~" or "-" in this specification means a range including the numerical values at both ends thereof as the upper limit value and the lower limit value, unless otherwise specified.

[0012] The present invention provides a pharmaceutical composition for preventing or treating IC / BPS, which contains an inhibitor against at least one integrin selected from the group consisting of integrin αvβ3, integrin αvβ5, and integrin α5β1.

[0013] Integrin is a heterodimer receptor composed of two subunits, an α chain and a β chain. In mammals, at least 18 different α subunits and 8 β subunits have been identified, and it is known that more than 24 heterodimers are formed. Eight subtypes of integrin, integrin αvβ1 (hereinafter simply referred to as αvβ1; the same applies to other integrins), αvβ3, αvβ5, αvβ6, αvβ8, αIIbβ3, α8β1, and α5β1, recognize the amino acid sequence Arg-Gly-Asp (RGD motif) and bind to a peptide having this sequence. These integrins are called RGD integrins or RGD-binding integrins.

[0014] RGD integrins have various physiological functions such as tissue repair, angiogenesis, inflammation, thrombosis, and promotion of tumor growth and metastasis. Since the function of RGD integrins is controlled by inhibiting their binding to peptide ligands having an RGD motif, it is expected to be effective in the treatment of diseases such as cancer and autoimmune diseases. Under such expectations, many compounds, specific antibodies, and other inhibitors against RGD integrins in various forms, also called RGD mimetics or RGD mimetics that mimic the structure of RGD, have been found and reported. The present invention provides a new use of an inhibitor against at least one integrin selected from the group consisting of αvβ3, αvβ5, and α5β1.

[0015] The inhibitor against at least one integrin selected from the group consisting of αvβ3, αvβ5, and α5β1 used in the present invention (hereinafter referred to as an RGD integrin inhibitor) refers to a substance having an activity of inhibiting the binding between at least one integrin selected from the group consisting of αvβ3, αvβ5, and α5β1 and a peptide having an RGD motif as its ligand, such as vitronectin or fibronectin. The RGD integrin inhibitor in the present invention inhibits the binding between at least one integrin selected from the group consisting of αvβ3, αvβ5, and α5β1 and a peptide having an RGD motif with an IC 50 of 1000 nM or less, preferably 500 nM or less, more preferably 200 nM or less, and even more preferably 100 nM or less.

[0016] The RGD integrin inhibitor in the present invention can be a compound such as a peptide, protein, glycoprotein, nucleic acid, or low-molecular organic compound. Also, a substance that inhibits the binding between an integrin and its ligand is sometimes referred to as an integrin antagonist. As long as it has an activity of inhibiting the binding between at least one integrin selected from the group consisting of αvβ3, αvβ5, and α5β1 and its ligand, those integrin antagonists are included in the RGD integrin inhibitor in the present invention.

[0017] Examples of RGD peptides and RGD mimetics that can be used as RGD integrin inhibitors in the present invention include, but are not limited to, cilengitide (CAS No. 188968-51-6), GLPG0187 (CAS No. 1320346-97-1), JSM-6427 (CAS No.), GSK3008348 (CAS No. 1629249-33-7), MK-0429 (CAS No. 227963-15-7), ATN-161 (CAS No. 262438-43-7), CWHM 12 (CAS No. 1564286-55-0), SB-273005 (CAS No. 205678-31-5), SB-267268 (CAS No. 205678-26-8), SC-68448 (CAS No. 188804-07-1), Compound 11 (CAS No. 287961-15-3), Example 1 (CAS No. 1893398-07-6), echistatin, RGD, RGDS (peptide consisting of the amino acid sequence shown in SEQ ID NO: 1), GRGD (peptide consisting of the amino acid sequence shown in SEQ ID NO: 2), GRGDS (peptide consisting of the amino acid sequence shown in SEQ ID NO: 3), GRGDSP (peptide consisting of the amino acid sequence shown in SEQ ID NO: 4), GRGDSPK (peptide consisting of the amino acid sequence shown in SEQ ID NO: 5), GRGDNP (peptide consisting of the amino acid sequence shown in SEQ ID NO: 6), GRGDTP (peptide consisting of the amino acid sequence shown in SEQ ID NO: 7), c(RGDfV), c(RGDfK), c(RGDyK), c(RGDfC), sn243, RGD-4C, RGD10, NC100717, c(phgisoDGRk), 44b, Mol 11, F-Galacto-c(RGDfK), (Ga)NOPO-c(RGDfK), c(RGDfK)-Peg-MPA, Flucilatide, c(phgisoDRGk)-Peg-MPA.

[0018] In addition, an antibody or its derivative that specifically binds to at least one integrin selected from the group consisting of αvβ3, αvβ5, and α5β1 and inhibits the binding to a ligand can also be used as an RGD integrin inhibitor in the present invention. A specific antibody against at least one integrin (target integrin) selected from the group consisting of αvβ3, αvβ5, and α5β1 can also be represented as an antibody that preferentially binds to the target integrin rather than to non-target proteins, or an antibody that has a high binding affinity for the target integrin. The specific antibody can be an antibody that binds to the target integrin with an affinity that is at least 5-fold stronger, preferably at least 10-fold stronger, more preferably at least 100-fold stronger, and most preferably at least 1000-fold stronger than the binding to non-target proteins. The specific antibody can also be an antibody that binds to the target integrin and has a dissociation constant of 10- 7 M, preferably 10- 8 M, more preferably 10- 9 M.

[0019] The specific antibody can be derived from any species including, for example, mouse, rat, shark, rabbit, pig, hamster, camel, llama, goat, or human. The specific antibody can also be of any class (e.g., IgG, IgE, IgM, IgD, or IgA) and subclass of immunoglobulin molecules, but is preferably IgG.

[0020] In the present invention, the specific antibody can be a polyclonal antibody or a monoclonal antibody, but is preferably a monoclonal antibody. The specific antibody can also be a chimeric antibody, a humanized antibody, or a human antibody.

[0021] Derivatives of specific antibodies are defined as antigen-binding fragments that are derived from specific antibodies and retain the ability to specifically bind to antigens, such as Fab (fragment of antigen binding), Fab', F(ab')2, single-chain antibodies (single chain Fv), disulfide-stabilized antibodies (disulfide stabilized Fv), and peptides containing CDRs, etc., but are not limited thereto.

[0022] Specific antibodies and their derivatives can be prepared using methods known to those skilled in the art. For example, based on the amino acid sequence of a human or the nucleotide sequence of DNA encoding the same, at least one integrin selected from the group consisting of αvβ3, αvβ5, and α5β1, or a partial peptide thereof having an RGD motif is prepared by genetic recombination techniques, and this is used as an antigen to immunize a suitable animal. Furthermore, it can be prepared by fusing B cells of the animal with myeloma cells to obtain a hybridoma. For the hybridoma method, see, for example, Meyaard et al. (1997) Immunity 7:283-290; Wright et al. (2000) Immunity 13:233-242; Kaithamana et al. (1999) J.Immunol. 163:5157-5164, etc. The specific antibody or its derivative may further be an antibody-drug conjugate bound to another drug.

[0023] Non-limiting examples of specific antibodies against at least one integrin selected from the group consisting of ανβ3, ανβ5 and α5β1 include Intetumumab (CNTO 95) (CAS No. 725735-28-4), Etaracizumab (MEDI-522, Abergrin) (CAS No. 89255-42-3), PF-04605412 (CAS No. 1677682-18-6), Volociximab (CAS No. 558480-40-3), EMD525797 (abituzumab, DI17E6) (CAS No. 1105038-73-0), Abciximab (ReoPro, c7E3) (CAS No. 143653-53-6), LM609 (MAB1976Z), P5H9 (MAB2528), P5D2 (MAB17781), 272-17E6 (MABT207).

[0024] The RGD integrin inhibitor in the present invention may have binding inhibitory activity against two or more RGD integrins selected from the group consisting of ανβ3, ανβ5 and α5β1, and may also have inhibitory activity against RGD integrins other than ανβ3, ανβ5 and α5β1.

[0025] In a preferred embodiment, the RGD integrin inhibitor in the present invention has at least inhibitory activity against ανβ3. For example, the IC 50 value can be 1000 nM or less, preferably 500 nM or less, more preferably 200 nM or less, and even more preferably 100 nM or less.

[0026] Examples of RGD integrin inhibitors having inhibitory activity against ανβ3 include, for RGD peptides and RGD mimetics, cilengitide (CAS No. 188968-51-6), GLPG0187 (CAS No. 1320346-97-1), GSK3008348 (CAS No. 1629249-33-7), MK-0429 (CAS No. 227963-15-7), CWHM 12 (CAS No. 1564286-55-0), SB-273005 (CAS No. 205678-31-5), SB-267268 (CAS No. 205678-26-8), SC-68448 (CAS No. 188804-07-1), Compound 11 (CAS No. 287961-15-3), Example 1 (CAS No. 1893398-07-6), exastatin, RGD, RGDS, GRGD, GRGDS, GRGDSP, GRGDSPK, GRGDNP, GRGDTP, c(RGDfV), c(RGDfK), c(RGDyK), c(RGDfC), sn243, RGD-4C, RGD10, NC100717, Mol 11, F-Galacto-c(RGDfK), (Ga)NOPO-c(RGDfK), c(RGDfK)-Peg-MPA, Flucilatide. Examples of specific antibodies against at least one integrin selected from the group consisting of ανβ3, ανβ5 and α5β1 include Intetumumab (CNTO 95) (CAS No. 725735-28-4), Etaracizumab (MEDI-522, Abergrin) (CAS No. 89255-42-3), EMD525797 (abituzumab, DI17E6) (CAS No. 1105038-73-0), Abciximab (ReoPro, c7E3) (CAS No. 143653-53-6), LM609 (MAB1976Z), P5H9 (MAB2528), P5D2 (MAB17781), 272-17E6 (MABT207).

[0027] The RGD integrin inhibitor in the present invention is particularly preferably an RGDS peptide or cilengitide.

[0028] In addition to the RGD integrin inhibitors exemplified above, substances determined to have binding inhibitory activity by a method known to those skilled in the art for evaluating inhibitory activity against αvβ3, αvβ5 or α5β1, for example, by comparing the binding of at least one integrin selected from the group consisting of αvβ3, αvβ5 and α5β1 and its ligand in the presence and absence of a test substance, respectively, can also be used as the RGD integrin inhibitors in the present invention. The measurement of binding inhibitory activity can be carried out within the scope of the ordinary implementation ability of those skilled in the art by appropriately using the amino acid sequences of the respective subunits of αvβ3, α5β1 and α5β1 and the nucleotide sequences of the genes encoding the same registered in public databases such as GenBank, recombinant cells capable of expressing each integrin, etc.

[0029] The pharmaceutical composition of the present invention is a pharmaceutical composition for the prevention or treatment of IC / BPS containing the above-mentioned RGD integrin inhibitor. The pharmaceutical composition of the present invention may contain one or more types of RGD integrin inhibitors in an arbitrary ratio. Further, the pharmaceutical composition of the present invention may contain an RGD integrin inhibitor in the form of a free form, an ester, a salt or a mixture thereof according to its physical properties, etc., and may contain a plurality of types of salts or esters, etc.

[0030] The pharmaceutical composition of the present invention is used for the prevention or treatment of IC / BPS. As used herein, the term "prevention" encompasses all medically acceptable types of preventive interventions aimed at preventing or suppressing the onset or development of a disease. The term "treatment" also encompasses all medically acceptable types of therapeutic interventions aimed at curing a disease or condition, providing temporary relief, etc. Therefore, the prevention or treatment of IC / BPS includes various medically acceptable interventions including improvement of symptoms such as frequent urination, increased urinary urgency, urgent urinary sensation, pain, etc. in IC / BPS, delay or cessation of progression, prevention of onset or prevention of recurrence. In particular, the pharmaceutical composition of the present invention is used for improving frequent urination or improving pain in IC / BPS.

[0031] The pharmaceutical composition of the present invention contains an effective amount of an RGD integrin inhibitor for the prevention or treatment of IC / BPS. The effective amount of the RGD integrin inhibitor can be appropriately determined according to the usage, age, sex, weight, severity, and other factors of the subject.

[0032] In addition to the RGD integrin inhibitor, the pharmaceutical composition of the present invention can contain pharmaceutically acceptable additives. Examples of pharmaceutically acceptable additives include buffers, stabilizers, preservatives, excipients, and the like. Pharmaceutically acceptable additives are well-known to those skilled in the art and can be appropriately selected and used within the scope of the ordinary implementation ability of those skilled in the art.

[0033] The pharmaceutical composition of the present invention can contain other drugs and the like for the prevention or treatment of IC / BPS. Examples of other drugs for the prevention or treatment of IC / BPS include the drugs described in Non-Patent Document 1 (Interstitial Cystitis / Bladder Pain Syndrome Treatment Guidelines). In particular, when the pharmaceutical composition of the present invention is intravesically instilled, drugs suitable for intravesical instillation, such as heparin, DMSO, or steroids, are preferred.

[0034] There is no particular limitation on the dosage form of the pharmaceutical composition of the present invention, but a dosage form suitable for intraperitoneal administration, intravenous administration, or transurethral or transvesical fistula intravesical instillation, such as a liquid preparation, is preferred. Such a pharmaceutical composition can be represented as a pharmaceutical composition for the prevention or treatment of IC / BPS for systemic administration or intravesical instillation, which contains an inhibitor of at least one integrin selected from the group consisting of αvβ3, αvβ5, and α5β1, i.e., an RGD integrin inhibitor.

[0035] The liquid preparation contains an effective amount of an RGD integrin inhibitor and a pharmaceutically acceptable medium. Examples of the pharmaceutically acceptable medium include aqueous media such as water, physiological saline, and phosphate-buffered saline (PBS). The unit dose of the liquid preparation can be determined according to the administration route, the effective amount of the RGD integrin inhibitor in the present invention, etc. For example, in the case of intravesical instillation, it is 30 to 50 mL, and this can be administered to the subject at appropriate intervals, such as once a week, once every other week, once a month, etc.

[0036] The pharmaceutical composition is administered to mammals including subjects at risk of developing or having developed IC / BPS, such as rodents including mice, rats, hamsters, guinea pigs, humans, primates including chimpanzees, macaques, livestock including pigs, cows, goats, horses, sheep, and pet animals including dogs, cats. A preferred subject is a human.

[0037] The present invention further provides a method for preventing or treating IC / BPS, which includes administering to a subject at risk of developing or having developed IC / BPS a pharmaceutical composition containing the above-mentioned RGD integrin inhibitor. The present invention also provides a method for improving frequent urination or pain in IC / BPS, which includes administering a pharmaceutical composition containing the above-mentioned RGD integrin inhibitor.

[0038] The present invention also provides, as another aspect, a method for evaluating the effect of a test substance on IC / BPS (hereinafter also referred to as this evaluation method), using the inhibitory activity against at least one integrin selected from the group consisting of αvβ3, αvβ5, and α5β1 as an index. A substance evaluated to be effective by this evaluation method is expected to be a substance that can be used for the prevention or treatment of IC / BPS through the inhibitory activity against at least one integrin selected from the group consisting of αvβ3, αvβ5, and α5β1.

[0039] One example of this evaluation method is a method known to those skilled in the art for evaluating the inhibitory activity against αvβ3, αvβ5 or α5β1. For example, by comparing the binding of at least one integrin selected from the group consisting of αvβ3, αvβ5 and α5β1 and its ligand in the presence and absence of a test substance, a test substance determined to have binding inhibitory activity is evaluated as a substance effective for the prevention or treatment of IC / BPS. The measurement of the binding inhibitory activity can be carried out within the scope of the normal implementation ability of those skilled in the art by appropriately using the amino acid sequences of the respective subunits of αvβ3, α5β1 and α5β1 registered in a public database such as GenBank and the nucleotide sequences of the genes encoding them, recombinant cells capable of expressing each integrin, etc.

[0040] The present invention will be described in more detail by the following examples, but the present invention is not limited thereto.

Examples

[0041] Example 1 A polyethylene catheter was inserted into and retained in the urethra of 10-week-old female F344 / NSIC344 rats (body weight approximately 140 - 170 g). Through the catheter, 250 μL of physiological saline (Sham group) or 250 μL of 0.1 M HCl (IC group) was injected into the bladder and retained for 1 minute, and then the bladder was washed twice with physiological saline. The Sham group and the IC group were each divided into two groups (n = 7 - 8 / group), and 200 μL of physiological saline or 200 μL of RGDS peptide solution (500 mg / L aqueous solution of physiological saline, MedChemExpress) was administered intraperitoneally. On the 7th day after administration, a polyethylene catheter was inserted into and retained at the vertex of the bladder, and a pressure transducer (MLT0699, ADInstruments Ltd, Dunedin, New Zealand) and PowerLab 4 / 26 (ADInstruments Ltd., Dunedin, New Zealand) were connected. A syringe pump (YSP-101, YMC Co., Ltd., Kyoto, Japan) was connected to the pressure transducer, and the intravesical pressure and urine volume were measured while continuously injecting 40 μL / min of physiological saline into the bladder. The pressure at the micturition peak on the intravesical pressure curve was defined as the maximum intravesical pressure, the interval between peaks was defined as the micturition interval, the pressure immediately after the peak was defined as the basal intravesical pressure, the pressure immediately before the peak was defined as the micturition threshold, and the urine volume at the peak was defined as the single micturition volume.

[0042] In the IC group injected with RGDS peptide (IC + R group), the micturition interval was significantly prolonged compared with the IC group injected with physiological saline (Figs. 1 and 2), and it was confirmed that frequent urination was improved. Also, the average value of the urine volume per micturition in the IC + R group was comparable to that of the Sham group. No differences were observed in the maximum intravesical pressure, basal intravesical pressure, and micturition threshold among all groups, indicating that RGDS peptide did not affect these parameters.

[0043] Example 2 In Example 1, the RGDS peptide was replaced with 400 μL of a sirencigitide solution (6.75 g / L aqueous physiological saline solution, Tokyo Chemical Industry), and the same test as in Example 1 was conducted. In the sirencigitide administration group, it was confirmed that the micturition interval was prolonged (Figure 3) and frequent urination was improved compared to the group injected with physiological saline.

[0044] Similar to Example 1, Sham group, IC group, and IC + R group were created. On the 5th day after administration, the rats were placed in a metabolic cage with a wire mesh bottom and acclimated for 10 minutes. Using an Electronic von Frey filament (BIO-EVF, Bioseb, Vitrolles, France), the pressure on the lower abdomen was gradually increased for stimulation, and the pressure at which an escape reaction was observed was measured as the pain threshold. The measurement was performed 3 times at 5-minute intervals, and the average value was calculated. In the IC + R group, the pain threshold increased compared to the IC group (Figure 4), and a tendency for pain improvement was confirmed.

Sequence Listing Free-Text

[0045] SEQ ID NO: 1 Amino acid sequence of a peptide that is an example of an RGD integrin inhibitor SEQ ID NO: 2 Amino acid sequence of a peptide that is an example of an RGD integrin inhibitor SEQ ID NO: 3 Amino acid sequence of a peptide that is an example of an RGD integrin inhibitor SEQ ID NO: 4 Amino acid sequence of a peptide that is an example of an RGD integrin inhibitor SEQ ID NO: 5 Amino acid sequence of a peptide that is an example of an RGD integrin inhibitor SEQ ID NO: 6 Amino acid sequence of a peptide that is an example of an RGD integrin inhibitor SEQ ID NO: 7 Amino acid sequence of a peptide that is an example of an RGD integrin inhibitor

Claims

1. A pharmaceutical composition for preventing or treating interstitial cystitis / bladder pain syndrome, containing a peptide or silengitide consisting of the amino acid sequence shown in SEQ ID NO:

1.

2. The pharmaceutical composition according to claim 1, for improving frequent urination or pain in interstitial cystitis / bladder pain syndrome.

3. The pharmaceutical composition according to claim 1 or 2, for intravesical instillation.

4. The pharmaceutical composition according to any one of claims 1 to 3, further comprising another agent for preventing or treating interstitial cystitis / bladder pain syndrome.

5. The pharmaceutical composition according to claim 4, wherein the other agent is heparin, DMSO or steroid.

Images