Pharmaceutical composition for parenteral administration for the treatment, prevention, or relief of osteoarthritis

Abstract

The present invention addresses the problem of providing a pharmaceutical composition for parenteral administration for treating, preventing or alleviating osteoarthritis.　A pharmaceutical composition for parenteral administration for treating, preventing or alleviating osteoarthritis, said pharmaceutical composition comprising, as an active ingredient, a conjugate of curcumin with a water-soluble substance.

Description

Technical Field

[0001] The present invention relates to a pharmaceutical composition for parenteral administration for the treatment, prevention or alleviation of osteoarthritis.

Background Art

[0002] Due to social aging and changes in lifestyle, the number of patients with arthritis tends to increase. Among arthritides, osteoarthritis (OA) is the most common degenerative joint disease with approximately 250 million patients worldwide (Non-Patent Document 1). The number of patients with knee osteoarthritis in Japan is estimated to be approximately 10 million in terms of the number of patients with subjective symptoms and approximately 30 million in terms of the number of potential patients (patients diagnosed by X-ray). The incidence increases with age, and in patients aged 50 or older, the female-to-male ratio is 1.5 to 2 times higher in women than in men. As treatment methods for knee osteoarthritis, a very large number are known, including physical therapy, hyaluronic acid injection, steroid injection, surgery, and prevention by supplements. However, the effects are not sufficient, and there is a desire to provide an effective medical treatment, and further, an effective medical treatment that does not have an adverse effect on articular cartilage and subchondral bone. Furthermore, preventive supplementation is desired.

[0003] In recent years, curcumin has been found to have pharmacological effects such as tumor formation inhibitory action, antioxidant action, anti-inflammatory action, cholesterol lowering action, anti-allergic action, brain disease preventive action, and heart disease preventive and therapeutic action, and its use in foods (for example, functional foods), pharmaceuticals, cosmetics, etc. has been studied. Regarding arthritis, curcumin is exemplified as one of the other active ingredients in a preventive agent for arthritis containing lutein and glucosamine as active ingredients, but no specific description of its effect is made (Patent Document 1). On the other hand, it has been reported that a parenteral pharmaceutical composition containing curcumin as a conjugate with a water-soluble substance (e.g., glucuronic acid, sulfate, glutathione, and amino acids) as the active ingredient improves the absorption of curcumin, which is poorly soluble in water, and as a result, the pharmacological effects of curcumin, such as its antitumor effect, can be fully obtained (Patent Document 2). [Prior art documents] [Patent Documents]

[0004] [Patent Document 1] Japanese Patent Publication No. 2013-95723 [Patent Document 2] International Publication No. 2018 / 003857 [Non-patent literature]

[0005] [Non-Patent Document 1] SHEN J., et al. Recent progress in osteoarthritis research. The Journal of the American Academy of Orthopedic Surgeons,2014;22(7):467-468. [Overview of the project] [Problems that the invention aims to solve]

[0006] The present invention aims to provide a parenterally administered pharmaceutical composition for the treatment, prevention, or alleviation of osteoarthritis, with reduced side effects. [Means for solving the problem]

[0007] Therefore, the inventors used an animal model of osteoarthritis and evaluated the degree of damage to articular cartilage, type II collagen and other arthritis markers in articular cartilage, and cysts and osteophytes in the subchondral bone after parenteral administration of various components. They found that water-soluble conjugates of curcumin improve or suppress these symptoms characteristic of osteoarthritis in both articular cartilage and subchondral bone, and furthermore, that systemic symptoms disappear with parenteral administration of water-soluble conjugates of curcumin, thus completing the present invention.

[0008] In other words, the present invention provides the following inventions [1] to

[11] . [1] A pharmaceutical composition for parenteral administration for the treatment, prevention, or relief of osteoarthritis, comprising a water-soluble conjugate of curcumin as an active ingredient. [2] The parenteral pharmaceutical composition according to [1] above, wherein the curcumins are one or more selected from curcumin and its derivatives. [3] The parenteral pharmaceutical composition according to [2] above, wherein the curcumin derivative is one or more selected from bisdemethoxycurcumin, demethoxycurcumin, and tetrahydrocurcumin. [4] A parenteral pharmaceutical composition according to any one of [1] to [3] above, wherein the water-soluble substance is one or more selected from glucuronic acid, sulfate, glutathione, and amino acids. [5] A parenteral pharmaceutical composition according to any one of [1] to [3] above, wherein the water-soluble substance is one or more selected from glucuronic acid and sulfuric acid. [6] The parenteral pharmaceutical composition according to [1] above, wherein the water-soluble conjugate of curcumins is curcumin monoglucuronide. [7] A parenteral pharmaceutical composition according to any one of [1] to [6] above, wherein the composition is a therapeutic agent for the treatment, prevention, or relief of osteoarthritis. [8] The parenteral pharmaceutical composition according to any one of [1] to [6] above, wherein the osteoarthritis is osteoarthritis of the knee, osteoarthritis of the hip, or osteoarthritis of the elbow. [9] Use of water-soluble conjugates of curcumins for the manufacture of parenteral pharmaceutical compositions for the treatment, prevention or relief of osteoarthritis.

[10] Water-soluble conjugates of curcumin-like substances for parenteral administration for the treatment, prevention, or relief of osteoarthritis.

[11] A method for treating, preventing, or alleviating osteoarthritis, characterized by administering an effective amount of a water-soluble conjugate of curcumin derivatives parenterally. [Effects of the Invention]

[0009] The present invention provides a parenteral pharmaceutical composition containing a water-soluble curcumin conjugate as an active ingredient, which can improve or suppress various symptoms of articular cartilage and subchondral bone in osteoarthritis. [Brief explanation of the drawing]

[0010] [Figure 1] Figure 1 shows the results of Safranin O and Fast-green staining of rat tibias in Example 1. The top two rows are substitute photographs; the first row shows the CMG-administered group, and the second row shows the control group, with staining results at 7 days, 14 days, 6 weeks, and 10 weeks in both cases. The bottom two rows of substitute photographs show the OA score quantified based on the Safranin O and Fast-green staining results. The first row shows the OARSI score, and the second row shows the modified Mankin histologic score. [Figure 2] Figure 2 shows the results of immunohistochemical evaluation of rat tibia from Example 1. The top two rows are substitute photographs; the first row (upper and lower) shows the localization of type I collagen, and the second row (upper and lower) shows the localization of type II collagen. The bottom two rows of substitute photographs show the lowest brightness of type I collagen in the first row and the average brightness of type II collagen in the second row. [Figure 3]Figure 3 shows the immunohistological evaluation results of IL1-β and IL6 in the rat tibia of Example 1. The upper two rows are substitute photographs of the drawings showing the expression status of the inflammatory cytokines IL1-β and IL6, and the lower two rows show the positive cell ratios (%) of IL1-β and IL6. [Figure 4] Figure 4 shows the immunohistological evaluation results of the inflammatory cytokine TNF-α in the rat tibia of Example 1. The upper row is a substitute photograph of the drawing showing the expression status of TNF-α, and the lower row shows the TNF-α positive cell ratio (%). [Figure 5] Figure 5 shows the number of cysts, the maximum cyst diameter, and the amount of osteophytes in the subchondral bone (SB) of the rat tibia in Example 1. The upper row shows a substitute photograph of the micro-CT imaging results of the subchondral bone, and the lower row shows the number of cysts (Cysts) in the subchondral bone and their diameter (mm), as well as the amount of osteophytes (mm3). [Figure 6] Figure 6 shows the results of Safranin O and Fast-green staining and the OA score for the rat knee cartilage in Example 1. The upper two rows are substitute photographs of the drawings. The first row is the CMG administration group, and the second row is the control group. Both are the staining results at 7 days, 14 days, 6 weeks, and 10 weeks. The lower two rows below the substitute photographs show the results of the OA score quantified based on the Safranin O and Fast-green staining results. The first row shows the OARSI score, and the second row shows the modified Mankin histologic score. [Figure 7] Figure 7 shows the immunohistological evaluation results for the rat knee cartilage in Example 1. The upper two rows are substitute photographs of the drawings showing the localization status of type I and type II collagen. The lower two rows below the substitute photographs show the minimum luminance of type I collagen and the average luminance of type II collagen.

Modes for Carrying Out the Invention

[0011] The active ingredient of the pharmaceutical composition for parenteral administration for the treatment, prevention or alleviation of osteoarthritis of the present invention is a water-soluble substance aggregate of curcumins. Curcumin is the main component of curcuminoids contained in turmeric pigment and is a compound represented by the following structural formula (1).

[0012] [ka]

[0013] The curcumins of the present invention are one or more selected from curcumin and its derivatives. Examples of curcumin derivatives include bisdemethoxycurcumin, demethoxycurcumin, and tetrahydrocurcumin. In the present invention, curcumin may be chemically synthesized curcumin or commercially available curcumin pigment. Examples of curcumin pigment include turmeric powder obtained by powdering the dried rhizome of turmeric (Curcuma longa LINNE) of the Zingiberaceae family, crude curcumin or oleoresin (turmeric oleoresin) obtained by extracting the turmeric powder with a suitable solvent (e.g., ethanol, oils and fats, propylene glycol, hexane, acetone, etc.), and purified curcumin. Curcumin contains both tautomers, the keto and enol forms.

[0014] The water-soluble substance in the water-soluble substance conjugate of curcumin compounds of the present invention can be one or more selected from glucuronic acid, sulfate, glutathione, and amino acids. The amino acids can be amino acids present in living organisms, such as essential amino acids. Water-soluble conjugates of curcumin can maintain high blood concentrations of free curcumin. This allows for the full utilization of the pharmacological effects of curcumin, and because curcumin conjugates are endogenous metabolites of curcumin, they are highly safe and therefore preferable. In curcumin-based water-soluble substance conjugates, the molar ratio of curcumin to the water-soluble substance is preferably curcumin:water-soluble substance = 1:1 to 1:3, more preferably 1:1 to 1:2, and even more preferably 1:1.

[0015] The conjugation form (binding form) of curcumin and the water-soluble substance is, for example, the form shown in formula (2).

[0016] [ka]

[0017] (In the formula, R 1 and R 2 At least one of them is a residue of the water-soluble substance, and the remainder is a hydrogen atom.

[0018] In equation (2) above, R 1 and R 2 One or both of these are preferably glucuronic acid residues or sulfate residues, and the remainder is preferably a hydrogen atom. Of these, R 1 and R 2 It is more preferable that one or both of them are glucuronic acid residues and the remainder is a hydrogen atom. In particular, R 1 R is a glucuronic acid residue 2 Curcumin monoglucuronide, in which the hydrogen atom is present, is even more preferable because it exhibits superior therapeutic, preventive, and mitigating effects on osteoarthritis.

[0019] Water-soluble conjugates of curcumin can be produced by the method described in Patent Document 2.

[0020] As shown in the examples below, water-soluble curcumin conjugates, when administered parenterally, improve or suppress various symptoms characteristic of osteoarthritis in both articular cartilage and subchondral bone, and furthermore, systemic symptoms disappear with parenteral administration of water-soluble curcumin conjugates. Symptoms specific to osteoarthritis include damage to articular cartilage, type II collagen in articular cartilage, arthropathy markers such as inflammatory markers, and the formation of subchondral bone cysts and osteophytes. Water-soluble conjugates of curcumin significantly suppress these conditions, including damage to articular cartilage, type II collagen in articular cartilage, arthropathy markers such as inflammatory markers, and the formation of subchondral bone cysts and osteophytes. Examples of osteoarthritis targeted by the parenterally administered pharmaceutical composition of the present invention include osteoarthritis of the knee, osteoarthritis of the hip, osteoarthritis of the elbow, and among these, osteoarthritis of the knee is particularly preferred.

[0021] The content of water-soluble curcumin conjugates in the parenteral pharmaceutical composition of the present invention cannot be determined in general terms, as it is appropriately determined according to the type of osteoarthritis, the patient's sex, age, symptoms, etc. However, for example, 1 to 100% by mass is preferred, 5 to 100% by mass is more preferred, and 10 to 100% by mass is even more preferred. The composition of the present invention is not particularly limited in terms of dosage form when used as a therapeutic agent. For example, it can be used as a liquid formulation or as a formulation prepared immediately after use from a freeze-dried product. It may also be in a sustained-release or sustained-release form. Furthermore, it can be prepared using additives such as carriers and excipients commonly used in pharmaceutical formulations.

[0022] The pharmaceutical composition of the present invention may be administered parenterally, and depending on the disease, symptoms, etc., it may be administered systemically or locally. Specifically, it may be administered by injection (e.g., local injection, intravenous injection, intramuscular injection, etc.). In the case of injection, it can be administered by injection into the joint, subcutaneously, intradermally, or intramuscularly, in addition to the usual intravenous and intra-arterial administration.

[0023] Aqueous solvents for injectable preparations may be, for example, distilled water or physiological saline. Non-aqueous solvents for injectable preparations may be, for example, propylene glycol, polyethylene glycol, vegetable oils such as olive oil, alcohols such as ethanol, or polysorbate 80 (pharmacopoeia name). Such preparations may further contain isotonic agents (e.g., sodium chloride, glucose), preservatives, wetting agents, emulsifiers, dispersants, stabilizers, pH adjusters (e.g., sodium citrate, sodium acetate, sodium phosphate), buffers, local anesthetics (e.g., procaine hydrochloride, lidocaine hydrochloride), or solubilizers. These formulations can be sterilized, for example, by filtration using a bacterial retention filter, incorporation of a bactericide, or irradiation. Alternatively, compositions obtained by dissolving or suspending a sterile solid composition in sterile water or an injectable solvent before use can also be used as these formulations. These formulations can be manufactured by known methods commonly used in the formulation process.

[0024] When the pharmaceutical composition of the present invention is a therapeutic, preventative, or palliative agent for osteoarthritis of the knee, hip, or elbow, it is preferable to administer it as an injectable, and more preferably as an intra-articular injection. In this case, the composition of the present invention may be in the form of a powder filler that dissolves at the time of use, or as an aqueous solution. In addition to water-soluble curcumin conjugates, parenteral compositions may contain water, physiological saline, pH adjusters, sugars, acids, alkalis, buffers, isotonic agents, stabilizers, analgesics, preservatives, and the like. Examples of sugars include monosaccharides, disaccharides, trisaccharides, polysaccharides, and sugar alcohols. Examples of acids and alkalis include water-soluble inorganic acids, water-soluble inorganic salts, water-soluble organic acids, water-soluble organic salts, amino acids, and amino acid salts.

[0025] The pharmaceutical composition of the present invention can be further combined with a polymer base material such as hyaluronic acid.

[0026] In the case of osteoarthritis of the knee, hip, or elbow, it is used not only for treatment but also for the prevention or alleviation of the condition. [Examples]

[0027] The present invention will now be described in more detail with reference to examples, but the scope of the present invention is not limited to the specific embodiments described below.

[0028] Manufacturing Example 1 (Method for preparing a water-soluble conjugate of curcumin (curcumin monoglucuronide)) Curcumin monoglucuronide was synthesized according to the method described in publication WO2018 / 03857. Specifically, 1.0 g (2.52 mmol) of acetobromo-α-D-glucuronate methyl ester was reacted with 326.0 mg (2.15 mmol) of vanillin as a starting material to obtain compound (1) (β-D-glucopyranosideuronic acid, 4-formyl-2-methoxyphenyl, methyl ester, triacetate) 491.6 mg (yield 49%). On the other hand, 3.3 mL (32.07 mmol) of 2,4-pentadione was reacted with 2.2 g of vanillin as a starting material to obtain compound (2) (5-hydroxy-1-(4-hydroxy-3-methoxyphenyl)-1,4-hexadien-3-one). Compound (1) and compound (2) were reacted to obtain compound (3) (curcumin β-D-glucopyranosideuronic acid 2,3,4-tri-O-acetyl,methyl ester). Then, deacetylation and purification of compound (3) yielded curcumin monoglucuronide (253.1 mg).

[0029] Example 1 (Effects of curcumin monoglucuronide (CMG) on an in vivo rat OA model) (I) Creation of an in vivo rat OA model and administration of CMG A rat osteoarthritis (OA) model was created by inducing OA in rats (Wistar, 12 weeks old, male) through medial meniscus destabilization surgery (DMM surgery), which involves cutting the tibial attachment of the anterior segment of the medial meniscus in the right knee (Glasson, SSet al, The surgical destabilization of the medial meniscus (DMM) model of osteoarthritis in the 29 / SvEv mouse. Osteoarthritis Cartilage 15, 1061-1069, (2007)). The left knee of the same rat was designated as the Sham surgery side, with only the skin and joint capsule incision being performed. Sixty-four OA model rats were divided into two groups: a treatment group that received intra-articular administration of CMG (camphor methylmethylglycolate) and a control group that received intra-articular administration of saline solution. The observation periods for each group were 7 days (7D), 14 days (14D), 6 weeks (6W), and 10 weeks (10W) postoperatively (n=8 rats for each group). Rats in the 7D postoperative observation period received intra-articular administration (CMG 30 mg / ml or saline solution, 50 μl) on 3 and 7 days postoperatively, respectively. Rats in the 14D postoperative observation period received intra-articular administration on 3, 7, 10, and 14 days postoperatively. Rats in the 6W and 10W postoperative observation periods received weekly intra-articular administration from 1 week postoperatively until immediately before slaughter.

[0030] Each animal was sacrificed one hour after intra-articular administration at the end of the observation period, and the knee joint was excised. After micro-CT scanning, the knee joint was fixed with PFA, decalcified, dehydrated, and then embedded in paraffin for histological examination. Thin sections were prepared and used for the following evaluations. (1) Histological evaluation Safranin O and Fast Green staining (staining and evaluating sections prepared in a sagittal section passing through the medial part of the tibial plateau and the center of the patellar cartilage). (2) OARSI score (quantified assessment of articular cartilage tissue using a histological scoring system by the Osteoarthritis Research Society International): Based on Safranin O and Fast Green staining, a score (0-24) is assigned from the following Grade and Stage, in accordance with Glasson, SSet al, The OARSI histopathology initiative-recommendations for histological assessments of osteoarthritis in the mouse, J. Osteoarthritis and Cartilage, 18, 2010, S17-S23. <grade> 0: Normal, no articular cartilage damage 1: There is no cartilage damage, but fibrosis and cell death are observed on the surface of the articular cartilage. 2: Damage and irregularity of the articular cartilage surface are observed, accompanied by decreased staining of the matrix. 3: There are vertical cracks in the articular cartilage, and the staining of the matrix is ​​also deeply reduced. 4: The superficial articular cartilage is detached. The cartilage has collapsed down to the middle layer. 5: The articular cartilage is deeply damaged, and the subchondral bone is exposed. 6: The articular cartilage is deeply damaged, the subchondral bone is exposed, and bone spurs have formed. <stage> 0: No signs of OA observed. 1: <10% 2:10-25% 3:25-50% 4:>50% (3) Modified Mankin (MM) score (assessment of the severity of articular cartilage damage): The score is calculated by summing the values ​​of the following items. <Pericellular Matrix Staining> 0: Normal 1: Brightness of a portion increases. 2: Brightness increases in cartilage with 40% or more of the cartilage. <Spatial Arrangement of Chondrocytes> 0: Normal 1: Increase in chondrocytes over a wide area 2: Decrease in some cartilage cells 3: Widespread reduction of chondrocytes <Interterritorial Matrix Staining> 0: Normal 1: Decreased stainability 2: Partial loss of staining 3: Staining disappears in more than 40% of articular cartilage.

[0031] (4) Immunohistological evaluation (evaluation of type I and type II collagen, IL1-β, IL6, and TNF-α) Type I collagen: After staining, the brightness of the articular cartilage in the medial part of the tibial plateau and the patellar cartilage was measured using Image-J, with the lowest (darkest) brightness being measured. Type II collagen: After staining, the brightness of articular cartilage in the medial part of the tibial plateau and patellar cartilage was measured using Image-J (NIH) to obtain the average brightness. After staining with IL1-β, IL6, and TNF-α, the percentage of positive cells among the chondrocytes in the medial part of the tibial plateau was calculated. (5) Evaluation of subchondral bone cysts and osteophytes by CT scan Subchondral bone and osteophytes were evaluated from micro-CT images. The number and diameter of the largest bone cyst were measured. The amount of osteophytes on the medial side of the tibial plateau was calculated using Amira (Thermo Fisher Scientific).

[0032] (II) Evaluation Results First, at 7 days, 14 days, and 6 weeks post-surgery, a fluorescent substance presumed to be curcumin was detected in the articular cartilage.

[0033] (Histological evaluation of the medial portion of the tibial plateau) Safranin O and Fast-green stained images and OA scores are shown in Figure 1. In the stained images, the CMG administration group (top panel) showed little effect in suppressing OA during the early postoperative period of high inflammation. However, at 10 weeks, both the OARSI score (top panel) and the MMH score (bottom panel) were significantly lower in the CMG administration group, indicating that CMG administration was able to suppress the progression of OA in the chronic phase. (Immunohistological evaluation of the medial portion of the tibial plateau) Furthermore, the immunohistochemical evaluation results shown in Figures 2, 3, and 4 indicate that type II collagen at 10W was significantly (p<0.05) darker and more intensely stained in the CMG-administered group (Figure 2), suggesting that OA-induced degradation of type II collagen was suppressed. The percentage of cells expressing positive for inflammatory cytokines IL1-β and IL6 (Figure 3), and TNF-α (Figure 4) was significantly lower (p<0.05) in the CMG-administered group at 6W, indicating that inflammatory cytokine expression was suppressed.

[0034] (Evaluation of subchondral bone in the medial part of the tibial plateau) Micro-CT imaging results (Figure 5) show that at 10 weeks, the number of cysts and the maximum cyst diameter were significantly smaller in the CMG administration group (p<0.01), and the amount of osteophytes at 10 weeks was also significantly smaller in the CMG administration group (p<0.01), indicating that the progression of OA was suppressed. (Evaluation of patellar cartilage) Similar to the evaluation of the articular cartilage in the medial tibial plateau, the patellar cartilage was also evaluated. Safranin O and Fast-green stained images and OA scores are shown in Figure 6, and immunohistochemical evaluation results (type I and type II collagen) are shown in Figure 7. The OARSI score at 10W (upper panel of Figure 6) was significantly lower in the CMG-administered group (p<0.01), and the average brightness of type II collagen at 10W was significantly lower in the CMG-administered group (p<0.05), indicating that OA was suppressed.

[0035] From the above, it was confirmed that OA symptoms improved in both articular cartilage and subchondral bone between weeks 6 and 10. Furthermore, no significant systemic toxicity symptoms were observed in any of the CMG administration groups, confirming that it did not adversely affect cartilage or subchondral bone.< / stage> < / grade>

Claims

1. A pharmaceutical composition for intra-articular administration for the treatment, prevention, or relief of osteoarthritis, comprising curcumin monoglucuronide as the active ingredient.

2. The intra-articular pharmaceutical composition according to claim 1, wherein the osteoarthritis is osteoarthritis of the knee, osteoarthritis of the hip, or osteoarthritis of the elbow.

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