Injectable preparation containing isoxazoline derivative and its manufacturing method

Abstract

The present invention relates to an injectable formulation comprising, as an active pharmaceutical ingredient (API), an isoxazoline derivative or a pharma- ceutically acceptable salt thereof, which is useful as a caspase inhibitor. The injectable formulation according to the present invention comprises a first agent comprising a high-dose API and a second agent comprising a reconstitution solution, and can be prepared by mixing the first agent and the second agent just before administration to a patient. Since the injectable formulation stably contains the active API, it has the advantage that effective efficacy can be expected when administered to a patient.

Description

[Technical field]

[0001] This application claims the benefit of priority based on Korean Patent Application No. 10-2022-0036549 filed on March 24, 2022, and all contents disclosed in the documents of that Korean patent application are incorporated as part of this specification.

[0002] The present invention relates to an injectable formulation containing an isoxazoline derivative or a pharma- ceutically acceptable salt thereof, which is useful as a caspase inhibitor, as an active pharmaceutical ingredient (API), and a method for producing the same. [Background technology]

[0003] Caspase is a cysteine ​​protease, and caspase inhibitors are compounds that can regulate inflammation and apoptosis caused by the action of caspases by inhibiting the action of caspases. Diseases that can be treated with these compounds to eliminate or alleviate symptoms include osteoarthritis, rheumatoid arthritis, degenerative arthritis, destructive bone disease, liver disease caused by viral hepatitis, acute hepatitis, liver sclerosis, brain damage caused by viral hepatitis, human sudden liver failure, sepsis, organ transplant rejection, ischemic heart disease, dementia, stroke, brain damage caused by AIDS, diabetes, and gastric ulcers.

[0004] As caspase inhibitors, various kinds of isoxazoline derivatives and prodrugs of isoxazoline derivatives are known. Among the isoxazoline derivatives, (R)-N-((2S,3S)-2-(fluoromethyl)-2-hydroxy-5-oxotetrahydrofuran-3-yl)-5-isopropyl-3-(isoquinolin-1-yl)-4,5-dihydroisoxazole-5-carboxamide (hereinafter referred to as "compound of formula 1") has already been proven to be effective in liver diseases caused by viral hepatitis, liver fibrosis, and nonalcoholic steatohepatitis (NASH). [ka]

[0005] Recently, there have been attempts to apply the compound of Formula 1 to diseases related to bones and joints, such as osteoarthritis (OA), rheumatoid arthritis, degenerative arthritis, and destructive bone disorders.

[0006] In the treatment of diseases related to bones and joints, the general method of drug administration is to administer the drug directly into the joint cavity, which is filled with synovial fluid, using a syringe. When administered orally, the drug has a systemic effect, but systemic side effects cannot be avoided. On the other hand, when administered directly into the joint cavity, local administration of the drug has the advantage of not only achieving the effect of local treatment where the drug is directly delivered, but also of being able to administer a high concentration of the drug and minimizing systemic side effects.

[0007] The compound of Chemical Formula 1 has been confirmed to have therapeutic effects in oral preparations for existing liver diseases, but since the compound of Chemical Formula 1 is poorly soluble in water, it is difficult to formulate it into an injectable preparation.

[0008] Therefore, there is a need to develop a high-concentration injectable formulation that can use the compound of Chemical Formula 1 as an injection. [Prior art documents] [Patent documents]

[0009] [Patent Document 1] Korean Patent Publication No. 0774999 (November 2, 2007) Isoxazoline derivatives and their manufacturing method [Patent Document 2] Korean Patent Application No. 2021-0102476 (August 4, 2021) Use of caspase inhibitors in alleviating or treating osteoarthritis [Non-patent literature]

[0010] [Non-Patent Document 1] Ratziu et al.,'A Phase2, Randomized, Double‐Blind, Placebo‐Controlled Study of GS‐9450in Subjects with Nonalcoholic Steatohepatitis' HEPATOLOGY, Vol.55,No.2,2012. Summary of the Invention [Problem to be solved by the invention]

[0011] As a result of extensive research conducted by the present inventors to solve the above problems, the present invention aims to provide an injectable formulation having a high content of the compound of Chemical Formula 1 as an active pharmaceutical ingredient (API).

[0012] In addition, the present invention aims to provide a freeze-dried injectable formulation that can be stored for a long period of time by increasing the stability of an API in an injectable formulation containing a high content of the compound of Chemical Formula 1 as an API. [Means for solving the problem]

[0013] The present invention provides a method for producing a first solution comprising the steps of: (S1) dissolving a compound represented by the following Formula 1 in a solvent containing a saccharide; [ka]

[0014] The present invention provides a method for producing an injectable formulation, comprising: (S2) freeze-drying the first solution to produce a first agent in powder form; and (S3) producing a second agent which is a reconstituted solution containing a phosphate buffer, a surfactant, a pH adjuster, or a combination thereof.

[0015] The present invention also provides a method for producing an injectable formulation, wherein the solvent has a pH of 1 or less.

[0016] The present invention also provides a method for producing an injectable formulation, wherein the solvent comprises hydrochloric acid, perchloric acid, nitric acid or sulfuric acid.

[0017] The present invention also provides a method for producing an injectable formulation, wherein the solvent comprises dimethyl sulfoxide or tert-butyl alcohol.

[0018] The present invention also provides a method for producing an injectable formulation, wherein the saccharide is at least one selected from the group consisting of mannitol, lactose, sucrose, trehalose, and sorbitol.

[0019] The present invention also provides a method for preparing an injectable formulation, wherein the content of the saccharide is 1 to 10% (w / w) based on the solvent.

[0020] The present invention also provides a method for producing an injectable formulation, wherein the surfactant is one or more selected from the group consisting of polysorbate 20, polysorbate 40, polysorbate 60, polysorbate 65 and polysorbate 80.

[0021] The present invention also provides a method for preparing an injectable formulation, wherein the pH adjuster contained in the second agent is NaOH.

[0022] The present invention provides an injectable formulation comprising a first agent in powder form comprising a compound of formula 1 and a saccharide; and a second agent which is a reconstituted solution comprising a phosphate buffer, a surfactant, a pH adjuster, or a combination thereof: [ka]

[0023] The present invention also provides an injectable preparation, wherein the saccharide is at least one selected from the group consisting of mannitol, lactose, sucrose, trehalose, and sorbitol.

[0024] The present invention also provides an injectable preparation, wherein the content of the saccharide is 1 to 10% (w / w) based on the solvent.

[0025] The present invention also provides an injectable preparation, wherein the surfactant is one or more selected from the group consisting of polysorbate 20, polysorbate 40, polysorbate 60, polysorbate 65 and polysorbate 80.

[0026] The present invention also provides an injectable formulation, wherein the pH adjuster contained in the second agent is NaOH.

[0027] The present invention also provides an injectable preparation having a compound of formula 1 in a content of 1 to 20 mg / ml based on a mixed solution of the first and second agents.

[0028] The present invention also provides an injectable preparation for treating or preventing a disease selected from the group consisting of osteoarthritis, rheumatoid arthritis, degenerative arthritis, destructive bone disorders, liver disease due to viral hepatitis, acute hepatitis, liver sclerosis, brain damage due to viral hepatitis, sudden human liver failure, sepsis, organ transplant rejection, ischemic heart disease, dementia, stroke, brain damage due to AIDS, diabetes, and gastric ulcer.

[0029] The present invention also provides an injectable preparation, wherein the disease is selected from the group consisting of osteoarthritis, rheumatoid arthritis, degenerative arthritis and destructive bone disorders.

[0030] The present invention also provides an injectable preparation that is used in a single dose in the form of a mixed solution of the first and second agents.

[0031] The present invention also provides an injectable preparation which is prepared by mixing the first and second agents and then administering the mixture into the articular cavity.

[0032] The present invention also provides an injectable preparation which is administered into the articular cavity within 30 minutes after mixing the first and second agents.

[0033] The present invention also provides an injectable formulation which is administered once after the first and second agents are mixed.

[0034] The present invention provides an injectable formulation comprising: a first agent in the form of a powder containing a compound of the following formula 1 and a saccharide; and a second agent which is a reconstituted solution containing a phosphate buffer, a surfactant, a pH adjuster, or a combination thereof; [ka]

[0035] When the contents of the compound of the following formula 2 and the compound of the following formula 3 are measured within 30 minutes after mixing the first and second agents, the content of the compound of the following formula 2 is 94% or more of the total content of the compound of the following formula 2 and the compound of the following formula 3 is provided. [ka] [ka] Effect of the Invention

[0036] The injectable formulation according to the present invention comprises a first agent in the form of a lyophilized formulation and a second agent containing a reconstitution solution, and the first and second agents are mixed together just before administration to a patient to prepare a drug, which is then administered to the patient within 30 minutes, thereby providing an injectable formulation with a high content of active API.

[0037] It is also possible to provide a freeze-dried injectable preparation that enhances the stability of the active pharmaceutical ingredient (API) and can be stored for a long period of time. [Brief description of the drawings]

[0038] [Figure 1] 1 shows the XRD pattern measurement results of the lyophilized API dosage forms prepared in Examples 9 to 12. [Diagram 2]1 shows the TGA measurement results of the lyophilized dosage forms and DMSO prepared in Examples 9 and 10. [Diagram 3] 13(a) shows the TGA measurement result of the lyophilized dosage form prepared in Example 14, and (b) shows the DSC measurement result of the lyophilized dosage form prepared in Example 14. [Figure 4] 1A shows the TGA measurement results of the lyophilized dosage forms prepared in Examples 17 to 20, and FIG. 1B shows the DSC measurement results of the lyophilized dosage forms prepared in Examples 17 to 20. BEST MODE FOR CARRYING OUT THEINVENTION

[0039] The present invention will now be described in more detail.

[0040] All technical terms used in the present invention are used in the same sense as commonly understood by those having ordinary skill in the art unless otherwise defined. In addition, although preferred methods and samples are described in this specification, similar or equivalent methods and samples are also included in the scope of the present invention. The contents of all publications cited as references in this specification are incorporated herein by reference in their entirety.

[0041] Since the compound of the present invention represented by Chemical Formula 1 is almost insoluble in water, the solubility in solvents of various compositions was analyzed in order to find a solvent capable of dissolving the compound of Chemical Formula 1 at a high content.

[0042] As a result, it was confirmed that the compound of Chemical Formula 1 is highly soluble in basic solvents with a pH of 9.0 or more, strong acidic solvents with a pH of 1 or less, or specific solvents that can be freeze-dried. It was also confirmed that the compound of Chemical Formula 1 has the property of decreasing the pH of the solution while dissolving in the solvent.

[0043] In addition, it was discovered that when the compound of formula 1 is dissolved in an aqueous solvent, the lactone ring is cleaved and the compound is converted to the compound of formula 2. At this time, the compound of formula 2 is in an active form, but the compound of formula 2 is quickly and irreversibly converted to the structural isomer of formula 3 in an aqueous solution, and the compound of formula 3 is in an inactive form that does not exhibit caspase inhibitory activity, unlike the compound of formula 2. [ka] [ka] [ka]

[0044] In addition, the conversion rate of the compound of Chemical Formula 2 to the compound of Chemical Formula 3 was examined depending on the pH of the solvent. As a result, it was confirmed that the compound of Chemical Formula 3 was hardly produced in an acidic solvent, but was rapidly produced in a neutral or more basic solvent.

[0045] Therefore, it was found that the compound of Chemical Formula 1 is highly soluble in a basic solvent with a high pH, ​​but the active form of the dissolved compound is rapidly converted to an inactive form in the basic solvent. This mechanism may reduce the purity of the active ingredient in an injectable dosage form in which the compound of Chemical Formula 1 is an API, thereby affecting its efficacy, so there is a problem in that it is difficult to select an appropriate solvent that maintains high purity of the API after dissolving the compound of Chemical Formula 1.

[0046] Therefore, the present inventors have studied various injectable formulations and have completed a freeze-dried injectable formulation containing API with high purity and having improved long-term storage stability by dissolving the compound of Chemical Formula 1 in a strong acidic solvent having a pH of 1 or less or a freeze-driable organic solvent, and then freeze-drying the solution.

[0047] Manufacturing method of injection preparation According to one embodiment of the present invention, a method for preparing an injectable formulation comprises the following steps: (S1) preparing a first solution by dissolving a compound of Formula 1 below in a solvent containing a saccharide; [ka] (S2) freeze-drying the first solution to prepare a first agent in powder form; and (S3) preparing a second agent which is a reconstitution solution comprising a phosphate buffer, a surfactant, a pH adjuster, or a combination thereof.

[0048] In one embodiment of the present invention, a method for preparing an injectable formulation may include the step of (S1) dissolving a compound of Formula 1 in a solvent containing a saccharide to prepare a first solution: [ka]

[0049] The solvent may include a solvent composition capable of dissolving the compound of Chemical Formula 1.

[0050] The solvent may have a pH of less than 1. For example, any one may be selected from the group consisting of hydrochloric acid, perchloric acid, nitric acid, and sulfuric acid, but is not limited to these examples.

[0051] The solvent may also be dimethyl sulfoxide or tert-butyl alcohol.

[0052] When the solvent is tert-butyl alcohol, the solvent may further include distilled water (DW). The tert-butyl alcohol and distilled water may be mixed in a ratio of 1:1 to 10:1. For example, the compound of Formula 1 may be dissolved in a solvent in which the tert-butyl alcohol and distilled water are mixed in a ratio of 1:1, 2:1, 3:1, 4:1, 5:1, 6:1, 7:1, 8:1, 9:1, or 10:1.

[0053] The solvent may include sugars, which may improve the dissolution stability of the compound of Formula 1 and may affect the formation of a stable and uniform cake during lyophilization.

[0054] Specifically, the saccharide may be any one selected from the group consisting of mannitol, lactose, sucrose, trehalose, and sorbitol, but is not limited to these examples.

[0055] The content of the sugars may be 1 to 10% (w / w) based on the solvent.

[0056] In one embodiment of the present invention, the method for preparing an injectable formulation may include (S2) freeze-drying the first solution to prepare a first agent in powder form.

[0057] As used herein, the term "lyophilization" refers to a process in which the material to be dried is first frozen and then the ice or frozen solvent is removed by sublimation in a vacuum environment.

[0058] The freeze-drying can be carried out by freezing at -80°C for 3 to 5 hours and then drying at 10 to 20 Pa for 10 to 50 hours.

[0059] The first agent may be in the form of a powder, a dry powder, or a lyophile. The first agent can be stored for a long period of time and is easy to formulate as an injection. In addition, since there are no additives that may cause side effects, it is completely harmless to the human body when used.

[0060] In one embodiment of the present invention, the method for preparing an injectable formulation may include preparing a second agent (S3) which is a reconstitution solution containing a phosphate buffer, a surfactant, a pH adjuster, or a combination thereof.

[0061] The term "reconstitution" as used herein means returning a solid state to a liquid state by adding water or a solution to a dry powder such as a lyophilized powder, and the reconstituted solution according to the present invention means a solution to be added to a first agent in powder form to produce a liquid state.

[0062] The second agent may include a reconstitution solution that includes a reconstitution medium such as distilled water, sterile purified water, water for injection, physiological saline, or a combination thereof, and further includes a phosphate buffer, a surfactant, a pH adjuster, or a combination thereof.

[0063] The injectable formulation of the present invention may include a buffer in the second agent. The buffer may be a common buffer that can be used in an injectable formulation, and specific examples include, but are not limited to, a phosphate buffer, a tris buffer, and an MES buffer. When a phosphate buffer is used, it may be selected from the group consisting of sodium phosphate dibasic, sodium phosphate monobasic, potassium phosphate dibasic, potassium phosphate monobasic, and mixtures thereof. When a tris buffer is used, a TRIZMA® buffer may be used.

[0064] The concentration of the buffer may be 50 to 150 mM based on the second agent, specifically, 50 to 150 mM, 50 to 145 mM, 50 to 140 mM, 50 to 135 mM, 50 to 130 mM, 50 to 125 mM, or 50 to 120 mM.

[0065] The injectable formulation of the present invention may include a surfactant in the second agent. The surfactant may be useful for dissolving the compound of Chemical Formula 1. The surfactant may be a common surfactant that can be used in an injection solvent, and may be a cyclodextrin-based or polysorbate-based surfactant. Specific examples of polysorbate-based surfactants include, but are not limited to, polysorbate 20, polysorbate 40, polysorbate 60, polysorbate 65, or polysorbate 80.

[0066] The concentration of the surfactant may be 0.1 to 1.0% based on the second agent, specifically, 0.1 to 1.0%, 0.2 to 1.0%, 0.3 to 1.0%, 0.4 to 1.0%, 0.1 to 0.9%, 0.2 to 0.9%, 0.3 to 0.9%, or 0.4 to 0.9%.

[0067] The injectable preparation of the present invention may contain a pH adjusting agent in the second agent. The pH adjusting agent may be NaOH.

[0068] The concentration of the NaOH may be 10 to 30 mM based on the second agent, specifically, 10 to 30 mM, 11 to 30 mM, 12 to 30 mM, 13 to 30 mM, 14 to 30 mM, 15 to 30 mM, 10 to 29 mM, 11 to 29 mM, 12 to 29 mM, 13 to 29 mM, 14 to 29 mM, 15 to 29 mM, 10 to 28 mM, 11 to 28 mM, 12 to 28 mM, 13 to 28 ... or 28 mM, 15 to 28 mM, 10 to 27 mM, 11 to 27 mM, 12 to 27 mM, 13 to 27 mM, 14 to 27 mM, 15 to 27 mM, 10 to 26 mM, 11 to 26 mM, 12 to 26 mM, 13 to 26 mM, 14 to 26 mM, 15 to 26 mM, 10 to 25 mM, 11 to 25 mM, 12 to 25 mM, 13 to 25 mM, 14 to 25 mM or 15 to 25 mM.

[0069] Injectable preparations According to one embodiment of the present invention, an injectable formulation may include a first agent in powder form including a compound of Formula 1 and a saccharide; and a second agent which is a reconstituted solution including a phosphate buffer, a surfactant, a pH adjuster, or a combination thereof: [ka]

[0070] The injectable formulation of the present invention may contain a saccharide in the first agent, which may improve the dissolution stability of the compound of Formula 1 and affect the formation of a stable and uniform cake during lyophilization.

[0071] Specifically, the saccharide may be any one selected from the group consisting of mannitol, lactose, sucrose, trehalose, and sorbitol, but is not limited to these examples.

[0072] The content of the sugar may be 1 to 10% (w / w) based on the solvent.

[0073] The first agent may be in the form of a powder, a dry powder, or a lyophile. The first agent can be stored for a long period of time and is easy to formulate into an injection. In addition, since there are no additives that cause side effects, the first agent is not harmful to the human body when used.

[0074] The second agent may include a reconstitution medium such as distilled water, sterile purified water, water for injection, saline, or a combination thereof, and the reconstitution medium may include a reconstitution solution containing a phosphate buffer, a surfactant, a pH adjuster, or a combination thereof.

[0075] The injectable formulation of the present invention may include a buffer in the second agent. The buffer may be a common buffer that can be used in an injectable formulation, and specific examples include, but are not limited to, a phosphate buffer, a tris buffer, and an MES buffer. When a phosphate buffer is used, it may be selected from the group consisting of sodium phosphate dibasic, sodium phosphate monobasic, potassium phosphate dibasic, potassium phosphate monobasic, and mixtures thereof. When a tris buffer is used, a TRIZMA® buffer may be used.

[0076] The concentration of the buffer may be 50 to 150 mM based on the second agent, specifically, 50 to 150 mM, 50 to 145 mM, 50 to 140 mM, 50 to 135 mM, 50 to 130 mM, 50 to 125 mM, or 50 to 120 mM.

[0077] The injectable formulation of the present invention may include a surfactant in the second agent. The surfactant may be useful for dissolving the compound of Chemical Formula 1. The surfactant may be a common surfactant that can be used in an injection solvent, and may be a cyclodextrin-based or polysorbate-based surfactant. Specific examples of polysorbate-based surfactants include, but are not limited to, polysorbate 20, polysorbate 40, polysorbate 60, polysorbate 65, or polysorbate 80.

[0078] The concentration of the surfactant may be 0.1 to 1.0% based on the second agent, specifically, 0.1 to 1.0%, 0.2 to 1.0%, 0.3 to 1.0%, 0.4 to 1.0%, 0.1 to 0.9%, 0.2 to 0.9%, 0.3 to 0.9%, or 0.4 to 0.9%.

[0079] The injectable preparation of the present invention may contain a pH adjusting agent in the second agent. The pH adjusting agent may be NaOH.

[0080] The concentration of the NaOH may be 10 to 30 mM based on the second agent, specifically, 10 to 30 mM, 11 to 30 mM, 12 to 30 mM, 13 to 30 mM, 14 to 30 mM, 15 to 30 mM, 10 to 29 mM, 11 to 29 mM, 12 to 29 mM, 13 to 29 mM, 14 to 29 mM, 15 to 29 mM, 10 to 28 mM, 11 to 28 mM, 12 to 28 mM, 13 to 28 ... or 28 mM, 15 to 28 mM, 10 to 27 mM, 11 to 27 mM, 12 to 27 mM, 13 to 27 mM, 14 to 27 mM, 15 to 27 mM, 10 to 26 mM, 11 to 26 mM, 12 to 26 mM, 13 to 26 mM, 14 to 26 mM, 15 to 26 mM, 10 to 25 mM, 11 to 25 mM, 12 to 25 mM, 13 to 25 mM, 14 to 25 mM or 15 to 25 mM.

[0081] According to an embodiment of the present invention, the pH of the reconstituted solution obtained by mixing the first and second agents may be 6.0 to 7.0. If the pH of the reconstituted solution is out of the range of 6.0 to 7.0, the patient may feel pain when administering the injectable preparation, and the reconstituted solution may not be suitable for use as an injectable preparation.

[0082] In an embodiment of the injectable formulation of the present invention, the osmotic pressure of the reconstituted solution obtained by mixing the first agent and the second agent may be 350 mOsm / kg to 500 mOsm / kg. If the osmotic pressure of the reconstituted solution is out of the range of 350 mOsm / kg to 500 mOsm / kg, the patient may feel pain when administering the injectable formulation, and the injectable formulation may not be suitable for use as an injectable formulation.

[0083] The content of the compound of Formula 1 in the injectable preparation according to one embodiment of the present invention may vary depending on the condition of the patient to be administered and the degree of the desired treatment.

[0084] The injectable formulation may contain the compound of Chemical Formula 1 in an amount of 1 to 20 mg / ml based on the reconstituted solution obtained by mixing the first agent and the second agent. Specifically, the compound of Chemical Formula 1 may have an amount of 1 to 20 mg / ml, 5 to 20 mg / ml, 10 to 20 mg / ml, 1 to 15 mg / ml, 5 to 15 mg / ml, 10 to 15 mg / ml, 1 mg / ml, 5 mg / ml, 10 mg / ml, 15 mg / ml, or 20 mg / ml based on the reconstituted solution, which is preferred for one dose.

[0085] When the content of the compound of Chemical Formula 1 is a low concentration of less than 1 mg / ml, a large amount of injection solution is required to achieve a sufficient therapeutic effect, which may cause difficulties when administered to the affected area of ​​the patient. When the content of the compound of Chemical Formula 1 is a high concentration of more than 20 mg / ml, the API may precipitate when dissolved in a solvent, or may cause pain when administered to the affected area of ​​the patient.

[0086] The injectable formulation according to one embodiment of the present invention may be in a transparent state in which the first agent containing the compound of Chemical Formula 1 is mixed with the second agent and the compound of Chemical Formula 1 is completely dissolved.

[0087] The injectable formulation according to one embodiment of the present invention can be used by reconstituting a first agent containing the compound of Chemical Formula 1 with a second agent containing the reconstituted solution just before administration to a patient, and administering the reconstituted solution to the affected area of ​​the patient.

[0088] In an embodiment of the injectable formulation of the present invention, the first and second agents are mixed together immediately before administration to a patient, and therefore may not require the inclusion of additional excipients for preservation, such as antioxidants and preservatives. However, in the case of the second agent containing a reconstitution solution, it may contain preservatives to prevent secondary contamination.

[0089] The present invention provides a method for treating or preventing a caspase-related disease using the injectable formulation of the present invention.

[0090] The term "treatment" means to halt or delay the progression of a disease when used on a subject who shows symptoms of the disease, and the term "prevention" means to halt or delay the onset of a disease when used on a subject who does not show symptoms of the disease but is at high risk of the disease.

[0091] The "caspase-related disease" of the present invention refers to a disease that can be treated or prevented by inhibiting caspase, and examples thereof include osteoarthritis, rheumatoid arthritis, degenerative arthritis, destructive bone disorders, liver disease due to viral hepatitis, acute hepatitis, hepatosclerosis, brain damage due to viral hepatitis, sudden human liver failure, sepsis, organ transplant rejection, ischemic heart disease, dementia, stroke, brain damage due to AIDS, diabetes, and gastric ulcer, but are not limited to the above diseases.

[0092] The injectable preparation of the present invention can be administered to an individual in need of treatment or prevention to treat or prevent a caspase-related disease in the individual.

[0093] The term "administration" or "administering" as used herein refers to a method of administering a dose of the injectable formulation of the present invention to a vertebrate or invertebrate animal, including a mammal, a bird, a fish, or an amphibian, by injection. A preferred administration method can be selected from common injection methods such as intravascular injection, subcutaneous injection, transdermal injection, intramuscular injection, spinal injection, and intradermal injection, depending on the type of disease to be treated, the site of the disease, and the severity of the disease. In particular, in the case of osteoarthritis, rheumatoid arthritis, degenerative arthritis, or destructive bone disorders, which are diseases related to bone and joints, intra-articular injection can be selected.

[0094] When the injectable preparation of the present invention is injected into a joint cavity, it can be guided by using an imaging method such as ultrasound, or a local anesthetic can be administered before or in combination with the injection.

[0095] The term "individual" as used herein refers to a human or a non-human mammal, such as a dog, cat, mouse, rat, cow, sheep, pig, goat, non-human ape, or bird. In some embodiments, the individual is a human.

[0096] The injectable formulation of the present invention is administered to an individual within 30 minutes after the first agent and the second agent are mixed to prepare a mixed solution. Thus, the injectable formulation of the present invention is for single administration for injection or administration to a single patient.

[0097] In addition, the injectable preparation of the present invention can be administered in combination with drugs such as painkillers, anti-inflammatory drugs, steroids, etc. Specifically, it can be administered in combination with drugs such as ibuprofen, naproxen, aspirin, acetaminophen, indomethacin, diclofenac, meloxicam, celecoxib, piroxicam, etodolac, nabumetone, lumiracoxib, valdecoxib, etoricoxib, parecoxib, fenoprofen, oxaprozin, mefenamic acid, diflunisal, flurbiprofen, ketoprofen, and other nonsteroidal anti-inflammatory drugs (NSAIDs); corticoid drugs such as prednisone and methylprednisolone, narcotic painkillers such as codeine, pentanyl, morphine, and meperidine, or injections of hyaluronic acid derivatives.

[0098] The present invention provides a kit including the injectable formulation of the present invention. In one embodiment of the present invention, the kit may include an injectable formulation including a first agent in powder form including the compound of Formula 1 of the present invention and saccharides, and a second agent which is a reconstituted solution; an administration system such as one or more syringes for administering the formulation, a method for mixing the first agent and the second agent, and instructions for using the kit including instructions for treating a patient.

[0099] In one embodiment of the present invention, the kit may include a label indicating that the formulation and contents described herein are administered to a patient suffering from a bone and joint disease, such as osteoarthritis, rheumatoid arthritis, degenerative arthritis, or destructive bone disease, whereby selection of an appropriate dosage for a patient is within the knowledge of a person of ordinary skill in the art, taking into account the patient's symptoms, age, etc.

[0100] Unless otherwise indicated, all numbers used in the present specification and claims, whether stated or not, should be understood to be modified in all cases by the term "about". Also, the precise values ​​used in the present specification and claims should be understood to form additional embodiments of the present disclosure. Efforts have been made to ensure the accuracy of the numerical values ​​disclosed in the examples. However, all measured numerical values ​​inherently contain certain error values ​​generated from the standard deviations observed with the respective measuring techniques. DETAILED DESCRIPTION OF THE PREFERRED EMBODIMENTS

[0101] Manufacturing Example In the present invention, the compound of formula 1, (R)-N-((2S,3S)-2-(fluoromethyl)-2-hydroxy-5-oxotetrahydrofuran-3-yl)-5-isopropyl-3-(isoquinoline-1-yl)-4,5-dihydroisooxazol-5-carboxamide) (hereinafter referred to as "API") used as an active pharmaceutical ingredient (API) was prepared according to KR10-0774999.

[0102] Test Example 1: Preparation of API lyophilized dosage forms (Examples 1 to 8) The API was dissolved in the solvent composition shown in Table 1 below to prepare an API mixed solution with a concentration of 0.5 mg / ml, and then freeze-dried. The freeze-drying conditions were freezing at -80°C for 6 hours, followed by drying at 10 to 20 Pa for 18 hours. After freeze-drying, purified water was used to reconstitute the well-formed cake into a dosage form, and the pH, osmotic pressure, and API purity of the reconstituted solution were measured, and the results are shown in Table 2 below. [Table 1] [Table 2]

[0103] From the results of Tables 1 and 2, it was confirmed that in Examples 3, 4, 7, and 8, in which mannitol was contained in the solvent, a white cake was formed after freeze-drying, and freeze-drying was successful. On the other hand, in Examples 1, 2, 5, and 6, in which mannitol was not contained in the solvent, no cake was formed after freeze-drying, and they were not suitable for use as freeze-dried dosage forms. In addition, the osmotic pressure of Example 3 was 303 mOsm / kg, while that of Example 4 was 434 mOsm / kg, and it was found that the osmotic pressure of Example 4 was increased by phosphate-buffer saline (PBS).

[0104] The purity of the API in the reconstituted solutions in Examples 3 and 4 was 100%, confirming that the API was highly stable in strong acids.

[0105] It was confirmed that Examples 7 and 8, which contained DMSO, an organic solvent that can be lyophilized, are suitable for use as lyophilized formulations.

[0106] The pH of the reconstituted solution in Example 7 was 3.87, whereas the pH of the reconstituted solution in Example 8 was 5.79. It was confirmed that the pH of Example 8 was close to neutral due to the PBS.

[0107] In addition, the osmotic pressure of Example 7 was 346 mOsm / kg, whereas the osmotic pressure of Example 8 was 495 mOsm / kg, indicating that the osmotic pressure of Example 8 was increased by PBS.

[0108] It is presumed that the purity of Examples 7 and 8 decreased during lyophilization due to the inclusion of moisture due to the hygroscopicity of DMSO. In particular, the purity of Example 8 decreased more rapidly due to the inclusion of 50% PBS.

[0109] Test Example 2: Preparation of API freeze-dried dosage forms (Examples 9 to 12) The API was dissolved in the solvent composition shown in Table 3 below to prepare API mixed solutions with a concentration of 0.5 mg / ml or 5.0 mg / ml, and then freeze-dried. The freeze-drying conditions were freezing at -80°C for 4 hours, and drying at 10 to 20 Pa for 18 hours. [Table 3]

[0110] The lyophilized dosage forms prepared in Examples 9 to 12 were reconstituted, and the purity of API after reconstitution, pH and osmolality of the reconstituted solution were measured, and the results are shown in Table 4 below. The solutions used for reconstituting the lyophilized dosage forms prepared in Examples 9 to 12 contained 50 mM sodium phosphate buffer (PB) or 0.1 M sodium hydroxide (NaOH). The sodium phosphate buffer was prepared at pH 7.4 by mixing sodium phosphate dibasic heptahydrate and sodium phosphate monobasic monohydrate. [Table 4]

[0111] From the results of Tables 3 and 4, the freeze-dried dosage forms prepared in Examples 9, 10, and 12, which have a mannitol content of 2.5%, formed less cake and did not have consistent properties compared to the freeze-dried dosage form prepared in Example 11, which has a mannitol content of 5.0%. The freeze-dried dosage forms prepared in Examples 9 and 10 were reconstituted using a phosphate buffer and completely dissolved within 1 minute, and the pH of the reconstituted solution was neutral at about 7.0. The purity of API in the reconstituted solution of Example 9 was measured to be 94.7% 30 minutes after reconstitution, which is slightly higher than that of Example 10 because the pH of the reconstituted solution was slightly lower.

[0112] The lyophilized dosage forms prepared in Examples 11 and 12 were observed to have API purity of less than 50% in the reconstituted solution because the pH of the reconstituted solution was higher than about 12. The reconstituted solution in Example 11, which contains 5.0% mannitol, had an osmotic pressure of 410 mOsm / kg, whereas the reconstituted solution in Example 12, which contains 2.5% mannitol, had an osmotic pressure of 288 mOsm / kg. This confirmed that the mannitol content is a factor that affects the osmotic pressure of the reconstituted solution.

[0113] Test Example 3: XRD Pattern Analysis of API Lyophilized Dosage Forms (Examples 9 to 12) The XRD patterns of the lyophilized dosage forms prepared in Examples 9 to 12 were measured under the following measurement conditions: - Voltage: 45 kV, current: 40 mA, scan range: 4° to 40° (2θ). FIG. 1 shows the XRD patterns of the lyophilized API formulations prepared in Examples 9 to 12.

[0114] In the API freeze-dried formulations of Examples 9 to 12, the mannitol peak was observed, but the API powder peak was not observed (Figure 1). This indicates that the API was well dissolved in the solution and was well freeze-dried, turning it into an amorphous API.

[0115] In Examples 11 and 12, some of the mannitol peaks were absent or had very low intensity, suggesting that some of the mannitol may have been converted to amorphous form.

[0116] Test Example 4: Confirmation of residual DMSO in API lyophilized dosage forms (Examples 9 to 12) The lyophilized dosage forms prepared in Examples 9 and 10 were subjected to TGA measurement at a temperature range of 30 to 350° C. and a heating rate of 10° C. / min.

[0117] FIG. 2 shows the TGA measurement results of the lyophilized formulations prepared in Examples 9 and 10 and DMSO.

[0118] In FIG. 2, the DMSO graph shows that the weight decreases from about 40 to 145°C, which is the range in which DMSO evaporates. The graphs of Examples 9 and 10 also show weight loss in the temperature range of 40 to 145°C. Example 9 shows a weight loss of about 11%, and Example 10 shows a weight loss of about 7%, confirming that a large amount of residual solvent remains in both Examples 9 and 10. To reduce the residual DMSO, the freeze-drying time must be increased.

[0119] Test Example 5: Preparation of API lyophilized dosage forms (Examples 13 to 16) The API was dissolved in the solvent composition shown in Table 5 below to prepare API mixed solutions with a concentration of 1.0 mg / ml to 10.0 mg / ml, which were then freeze-dried. The freeze-drying conditions were freezing at -80°C for 5 hours, followed by drying at 10 to 20 Pa for 40 hours. [Table 5]

[0120] The lyophilized dosage forms prepared in Examples 13 to 16 were reconstituted, and the purity of API after reconstitution, pH and osmotic pressure of the reconstituted solution were measured, and the results are shown in Table 6 below. The solution used for reconstituting the lyophilized dosage forms prepared in Examples 13 to 16 included 50 mM sodium phosphate buffer (PB). The sodium phosphate buffer was prepared at pH 7.4 by mixing sodium phosphate dibasic heptahydrate and sodium phosphate monobasic monohydrate. [Table 6]

[0121] From the results of Tables 5 and 6, it was difficult to dissolve 2.5 mg / ml of API in Example 15, which is a solvent containing HCl. This is expected to result from mixing different compositions of the reconstituted solution and then increasing the pH to dissolve it transparently. The higher the pH of the reconstituted solution, the greater the decrease in purity 30 minutes after reconstitution compared to the purity immediately after reconstitution.

[0122] Test Example 6: Thermal Analysis of API Lyophilized Dosage Form (Example 14) The lyophilized formulation prepared in Example 14 was subjected to TGA and DSC measurements at a temperature range of 30 to 350° C. and a heating rate of 10° C. / min.

[0123] FIG. 3 (a) shows the TGA measurement result of the lyophilized dosage form prepared in Example 14, and (b) shows the DSC measurement result of the lyophilized dosage form prepared in Example 14.

[0124] In Figure 3 (a), the TGA measurement graph shows a weight loss of about 1.4% in the temperature range of about 40 to 145°C where DMSO evaporates. This confirms that the residual DMSO was significantly reduced compared to Examples 9 and 10 in Figure 2. In other words, it was confirmed that the residual DMSO was reduced by increasing the freeze-drying time.

[0125] In FIG. 3 (b), the DSC measurement graph shows an endothermic peak at about 164° C., which can be regarded as the melting point of mannitol (165° C.). Therefore, this can be interpreted as meaning that the mannitol in the freeze-dried dosage form prepared in Example 14 is present in a crystalline form.

[0126] Test Example 7: Preparation of API freeze-dried dosage forms (Examples 17 to 20) The API was dissolved in the solvent composition shown in Table 7 below to prepare an API mixed solution with a concentration of 15.0 mg / ml, which was then freeze-dried. The freeze-drying conditions were freezing at -80°C for 3 hours, followed by drying at 6 Pa for 18 hours.

[0127] Mannitol contents were varied by varying the ratio of tert-butyl alcohol (TBA) and distilled water (DW).

[0128] Since the freezing point of TBA is 26°C, the solvent was heated to 40°C to prepare the API mixed solution. [Table 7]

[0129] The properties of the lyophilized dosage forms of API prepared in Examples 17 to 20 were confirmed, and the purity and pH of the solutions reconstituted from the lyophilized dosage forms were measured, and the results are shown in Table 8. The solvent used for reconstitution included 10 mM sodium phosphate buffer or 0.5% polysorbate 80 (Tween 80). [Table 8]

[0130] The freeze-dried API formulations prepared in Examples 17 to 20 all successfully formed freeze-dried cakes. In relation to the freeze-dried formulations, the freeze-dried cake in Example 17 was weak like cotton candy, and a yellow, sticky mass was formed upon reconstitution. It was confirmed that the yellow mass contained a large amount of API, which was due to the API being almost insoluble in the solution after reconstitution, and the pH was also observed to be about 7.0.

[0131] In the above Examples 18 to 20, the lyophilized cake was not dissolved in the solution after reconstitution, and the solution was slightly cloudy and had large lumps.

[0132] In Examples 18 to 20, which contained mannitol, the freeze-dried cakes were firm, whereas in Example 17, which did not contain mannitol, the freeze-dried cake was not firm. This indicates that the addition of a bulking agent such as mannitol results in the formation of a freeze-dried dosage form.

[0133] Test Example 8: Thermal Analysis of API Lyophilized Dosage Forms (Examples 17 to 20) The lyophilized dosage forms prepared in Examples 17 to 20 were subjected to TGA and DSC measurements at a temperature range of 30 to 350° C. and a heating rate of 10° C. / min.

[0134] FIG. 4(a) shows the TGA measurement results of the lyophilized dosage forms prepared in Examples 17 to 20, and (b) shows the DSC measurement results of the lyophilized dosage forms prepared in Examples 17 to 20.

[0135] In FIG. 4(a), the TGA measurement graph shows that there was a weight loss due to residual solvent in the 82° C. range, which is the boiling point of TBA.

[0136] In FIG. 4 (b), a DSC measurement graph shows an endothermic peak at 165° C. in Examples 18 to 20, which coincides with the peak of the melting point of mannitol, indicating that mannitol crystals are present in the freeze-dried cake.

[0137] Test Example 9: Analysis of API lyophilized dosage forms (Example 19) The pH, API recovery rate, and purity change for 60 minutes after reconstitution of the dosage form prepared in Example 19 were measured, and the results are shown in Table 9 below. [Table 9]

[0138] As can be seen from the results in Table 9, if the pH of the reconstituted solution is too low, the API does not dissolve. Therefore, by adding NaOH, the pH that decreases as the API dissolution proceeds is offset, allowing all of the API to dissolve.

[0139] It was confirmed that the lyophilized dosage form prepared in Example 19 maintained an API purity of 94% or more for 60 minutes after reconstitution and was stable for 60 minutes after reconstitution.

Claims

1. (S1) preparing a first solution by dissolving a compound represented by the following Formula 1 in a solvent containing a saccharide; 【Chemistry 1】 (S2) freeze-drying the first solution to prepare a first agent in powder form; and (S3) preparing a second agent which is a reconstitution solution containing a phosphate buffer, a surfactant, a pH adjuster, or a combination thereof.

2. The method for producing an injectable formulation according to claim 1 , wherein the solvent has a pH of 1 or less.

3. The method for producing an injectable preparation according to claim 2 , wherein the solvent contains hydrochloric acid, perchloric acid, nitric acid or sulfuric acid.

4. The method for producing an injectable formulation according to claim 1, wherein the solvent comprises dimethyl sulfoxide or tert-butyl alcohol.

5. The method for producing an injectable preparation according to claim 1, wherein the saccharide is at least one selected from the group consisting of mannitol, lactose, sucrose, trehalose and sorbitol.

6. The method for producing an injectable formulation according to claim 1, wherein the content of the sugar is 1 to 10% (w / w) based on the solvent.

7. The method for producing an injectable formulation according to claim 1, wherein the surfactant is at least one selected from the group consisting of polysorbate 20, polysorbate 40, polysorbate 60, polysorbate 65 and polysorbate 80.

8. The method for producing an injectable preparation according to claim 1 , wherein the pH adjuster contained in the second agent is NaOH.

9. A first agent in powder form containing a compound of the following formula 1 and sugars; and a second agent which is a reconstitution solution comprising a phosphate buffer, a surfactant, a pH adjuster, or a combination thereof; 【Chemistry 2】

10. The injectable formulation according to claim 9, wherein the saccharide is at least one selected from the group consisting of mannitol, lactose, sucrose, trehalose and sorbitol.

11. The injectable preparation according to claim 9, wherein the content of the sugar is 1 to 10% (w / w) based on the solvent.

12. The injectable preparation according to claim 9, wherein the surfactant is any one selected from the group consisting of polysorbate 20, polysorbate 40, polysorbate 60, polysorbate 65 and polysorbate 80.

13. The injectable preparation according to claim 9, wherein the pH adjusting agent contained in the second agent is NaOH.

14. The injectable preparation according to claim 9, wherein the content of the compound of Formula 1 is 1 to 20 mg / ml based on a mixed solution of the first agent and the second agent.

15. The injectable formulation according to claim 9, which is for treating or preventing a disease selected from the group consisting of osteoarthritis, rheumatoid arthritis, degenerative arthritis, destructive bone disorders, liver disease due to viral hepatitis, acute hepatitis, liver sclerosis, brain damage due to viral hepatitis, sudden human liver failure, sepsis, organ transplant rejection, ischemic heart disease, dementia, stroke, brain damage due to AIDS, diabetes and gastric ulcer.

16. The injectable formulation according to claim 9, wherein the disease is selected from the group consisting of osteoarthritis, rheumatoid arthritis, degenerative arthritis and destructive bone disorders.

17. The injectable preparation according to claim 9 , which is used in a single dose in the form of a solution in which the first agent and the second agent are mixed.

18. The injectable preparation according to claim 9 , wherein the injectable preparation is administered into an articular cavity after the first agent and the second agent are mixed.

19. The injectable preparation according to claim 9 , which is administered into the articular cavity within 30 minutes after mixing the first agent and the second agent.

20. The injectable preparation according to claim 9 , wherein the injectable preparation is administered once after the first agent and the second agent are mixed.

21. A first agent in powder form containing a compound of the following formula 1 and sugars; and a second agent which is a reconstitution solution comprising a phosphate buffer, a surfactant, a pH adjuster, or a combination thereof; 【Chemistry 3】 The injectable preparation is such that, when the contents of the compound of the following formula 2 and the compound of the following formula 3 are measured within 30 minutes after mixing the first and second agents, the content of the compound of the following formula 2 is 94% or more of the total content of the compound of the following formula 2 and the compound of the following formula 3: 【Chemistry 4】 【Chemistry 5】

Images