Prodrugs of caspase inhibitors

Abstract

The present invention relates to an isoxazoline derivative having an ester portion and a pharmaceutical composition containing the isoxazoline derivative, wherein the isoxazoline derivative is a prodrug of a caspase inhibitor.

Description

Technical Field

[0001] The present invention relates to isocyanate having a lactone moiety as a prodrug of caspase inhibitors Azoline derivative, and a pharmaceutical composition comprising the same. Background Art

[0002] Caspase is an enzyme and a cysteine ​​protease that exists in the form of α2β2 tetramers. Caspase inhibitors interfere with the activity of these caspases, thereby regulating inflammation or apoptosis caused by the action of caspases. Diseases whose symptoms can be eliminated or alleviated by administering these compounds include osteoarthritis, rheumatoid arthritis, degenerative arthritis, destructive bone disease, liver disease caused by hepatitis viruses, acute hepatitis, cirrhosis, brain damage caused by hepatitis viruses, human fulminant liver failure, sepsis, organ transplant rejection, ischemic heart disease, dementia, stroke, brain damage caused by AIDS, diabetes, gastric ulcers, etc.

[0003] Among the various compounds known as caspase inhibitors, Oxazoline derivatives have been applied for as Korean Patent Application Nos. 10-2004-0066726, 10-2006-0013107 and 10-2008-0025123. In addition, International Publication No. WO2007 / 015931 (applicant: Vertex Pharmaceuticals Incorporated, USA) discloses isocyanate derivatives. Azoline derivatives are prodrugs of caspase inhibitors. Summary of the invention

[0004] Technical issues

[0005] The present invention aims to develop heterodimers having the structure of formula 2 as effective inhibitors of caspases. The prodrug of oxazoline derivatives is used to improve bioavailability. In addition, the caspase inhibitor of Formula 2 has high water solubility and high hydrophilicity, and thus may be advantageous for the development of oral preparations, but may be disadvantageous in the development of long-acting preparations. Therefore, the present invention aims to develop a prodrug form of the caspase inhibitor of Formula 2 having hydrophobicity to facilitate long-acting preparations.

[0006] Technical Solutions

[0007] In order to achieve the above object, the present invention provides a compound of the following formula 1, or a pharmaceutically acceptable salt or isomer thereof:

[0008] [Formula 1]

[0009]

[0010] in

[0011] R represents alkyl, haloalkyl, cycloalkyl, cycloalkylalkyl, aryl, arylalkyl, heteroaryl, heteroarylalkyl, alkoxy or alkoxyalkyl, wherein the heteroaryl contains one or more heteroatoms selected from N, O and S:;

[0012] wherein the alkyl, haloalkyl, cycloalkyl, cycloalkylalkyl, aryl, arylalkyl, heteroaryl, heteroarylalkyl or alkoxyalkyl is optionally substituted, and the substituent may be one or more selected from the group consisting of alkyl, halogen, haloalkyl, cycloalkyl, hydroxy, acyl, amino, alkoxy, alkoxycarbonyl, oxo, carboxyl, carboxylamino, cyano, nitro, thiol, aryloxy, sulfoxy and guanidino;

[0013] Provided that R is not tert-butyl.

[0014] The compound of formula 1 according to the present invention can form a pharmaceutically acceptable salt. Pharmaceutically acceptable salts may include acid addition salts formed by the following acids: inorganic acids such as hydrochloric acid, sulfuric acid, nitric acid, phosphoric acid, hydrobromic acid and hydroiodic acid; organic acids such as tartaric acid, formic acid, citric acid, acetic acid, trichloroacetic acid, trifluoroacetic acid, gluconic acid, benzoic acid, lactic acid, fumaric acid, maleic acid and salicylic acid; or sulfonic acids such as methanesulfonic acid, ethanesulfonic acid, benzenesulfonic acid and p-toluenesulfonic acid, the acid forming a non-toxic acid addition salt including a pharmaceutically acceptable anion. In addition, pharmaceutically acceptable carboxylates include: salts formed with alkali metals or alkaline earth metals such as lithium, sodium, potassium, calcium and magnesium; salts formed with amino acids such as lysine, arginine and guanidine; organic salts such as dicyclohexylamine, N-methyl-D-glucamine, tris(hydroxymethyl)methylamine, diethanolamine, choline and triethylamine. The compound of formula 1 according to the present invention can be converted into its salt by conventional methods.

[0015] Meanwhile, since the compounds of Formula 1 according to the present invention may have an asymmetric carbon center and an asymmetric axis or plane, they may exist as E- or Z-isomers, R- or S-isomers, racemic mixtures or diastereomeric mixtures as well as each diastereomer, all of which are within the scope of the present invention.

[0016] Herein, unless otherwise indicated, the term "compound of Formula 1" is used to refer to all compounds of Formula 1, including pharmaceutically acceptable salts and isomers thereof.

[0017] Here, the following concepts for defining substituents are used to define the compounds of formula 1.

[0018] The term "halogen" or "halo" refers to fluorine (F), chlorine (Cl), bromine (Br) or iodine (I).

[0019] The term "alkyl" refers to a straight or branched chain hydrocarbon, which may include single, double or triple bonds, and is preferably C1-C 18 Alkyl. Examples of alkyl include, but are not limited to, methyl, ethyl, n-propyl, isopropyl, n-butyl, isobutyl, tert-butyl, pentyl, hexyl, heptyl, octyl, decyl, dodecyl, pentadecyl, octadecyl, acetylene, vinyl, trifluoromethyl, and the like.

[0020] The term "cycloalkyl" refers to a partially or fully saturated monocyclic or condensed ring hydrocarbon, and is preferably C3-C 10 Cycloalkyl. Examples of cycloalkyl include, but are not limited to, cyclopropyl, cyclobutyl, cyclopentyl, cyclohexyl, cycloheptyl, and the like.

[0021] Unless otherwise defined, the term "alkoxy" refers to an alkoxy group having 1 to 10 carbon atoms.

[0022] The term "aryl" includes at least one ring having a conjugated π electron system, including, for example, a monocyclic or fused polycyclic (i.e., a ring sharing adjacent pairs of carbon atoms) group. For example, a fused polycyclic may include a C3-C8 cycloalkyl ring fused to an aryl. That is, unless otherwise defined herein, an aryl is an aromatic monocyclic or polycyclic group having 5 to 15 carbon atoms, preferably 6 to 10 carbon atoms, including phenyl, naphthyl, dihydroindene, etc. For example, an aryl may be a C5-C8 cycloalkyl ring. 12 Aryl, preferably C6-C 10 Aryl.

[0023] The term "heteroaryl" refers to a 3- to 12-membered, more preferably 5- to 10-membered aromatic hydrocarbon that forms a monocyclic or condensed ring, which may be fused with a benzene ring or a C3-C8 cycloalkyl group, and includes one or more heteroatoms selected from N, O and S as ring members. Examples of heteroaryl groups include, but are not limited to, pyridyl, pyrimidinyl, pyridazinyl, pyrazinyl, Oxazolyl, iso Oxazolyl, tetrazolyl, triazolyl, indolyl, indazolyl, iso Azolyl, oxazolyl, thiazolyl, isothiazolyl, furanyl, benzofuranyl, imidazolyl, thienyl, benzothiazole, benzimidazole, quinolyl, indolyl, 1,2,3,4-tetrahydroisoquinolyl, 3,4-dihydroisoquinolyl, thiazolopyridyl, 2,3-dihydrobenzofuran, 2,3-dihydrothiophene, 2,3-dihydroindole, benzo[1,3]dihydro Chromane, thiochromane, 1,2,3,4-tetrahydroquinoline, 4H-benzo[1,3]diol 2,3-Dihydrobenzo[1,4]-dihydro English: English, 6,7-dihydro-5H-cyclopenta[d]pyrimidine, etc.

[0024] Cycloalkyl-alkyl, aryl-alkyl, heteroaryl-alkyl and alkoxy-alkyl refer to groups formed by combinations of the above cycloalkyl, aryl, heteroaryl, alkoxy and / or alkyl groups. Examples include but are not limited to benzyl, thephenylmethyl, pyrimidinylmethyl, etc.

[0025] According to one embodiment of the present invention, R may represent C 1-20 Alkyl, C 3-10 Cycloalkyl, C 3-10 Cycloalkyl-C 1-6 Alkyl, C6-C 10 Aryl, C6-C 10 Aryl-C 1-6 alkyl, 3- to 10-membered heteroaryl, 3- to 10-membered heteroaryl-C 1-6 Alkyl, halo-C 1-6 Alkyl or C 1-6 Alkoxy-C 1-6 The heteroaryl group may contain 1 to 4 heteroatoms selected from N, O and S, but is not limited thereto.

[0026] According to one embodiment of the present invention, R may represent C 1-18 Alkyl, C 3-6 Cycloalkyl, C 3-6 Cycloalkyl-C 1-3 Alkyl, C6-C 10 Aryl, C6-C 10 Aryl-C 1-3 Alkyl, 4- to 6-membered heteroaryl-C 1-3 Alkyl, halo-C 1-3 Alkyl or C 1-3 Alkoxy-C 1-3 Alkyl, wherein the heteroaryl may contain 1 or 2 heteroatoms selected from N, O and S, and the substituent may be alkyl, halogen, alkoxy or oxo, but is not limited thereto.

[0027] Representative compounds of Formula 1 according to the present invention include, but are not limited to, the following compounds:

[0028] (S)-5-Fluoro-3-((R)-5-isopropyl-3-(isoquinolin-1-yl)-4,5-dihydroisoquinolin-1-yl) Methyl 4-oxopentanoate;

[0029] (S)-5-Fluoro-3-((R)-5-isopropyl-3-(isoquinolin-1-yl)-4,5-dihydroisoquinolin-1-yl) Ethyl 4-oxopentanoate;

[0030] (S)-5-Fluoro-3-((R)-5-isopropyl-3-(isoquinolin-1-yl)-4,5-dihydroisoquinolin-1-yl) Propyl 4-oxopentanoate;

[0031] (S)-5-Fluoro-3-((R)-5-isopropyl-3-(isoquinolin-1-yl)-4,5-dihydroisoquinolin-1-yl) 4-oxopentanoic acid butyl ester;

[0032] (S)-5-Fluoro-3-((R)-5-isopropyl-3-(isoquinolin-1-yl)-4,5-dihydroisoquinolin-1-yl) Isobutyl 4-oxopentanoate;

[0033] (S)-5-Fluoro-3-((R)-5-isopropyl-3-(isoquinolin-1-yl)-4,5-dihydroisoquinolin-1-yl) Isoamyl 4-oxopentanoate;

[0034] (S)-5-Fluoro-3-((R)-5-isopropyl-3-(isoquinolin-1-yl)-4,5-dihydroisoquinolin-1-yl) 4-oxopentanoic acid pentyl ester;

[0035] (S)-5-Fluoro-3-((R)-5-isopropyl-3-(isoquinolin-1-yl)-4,5-dihydroisoquinolin-1-yl) 4-oxopentanoic acid hexyl ester;

[0036] (S)-5-Fluoro-3-((R)-5-isopropyl-3-(isoquinolin-1-yl)-4,5-dihydroisoquinolin-1-yl) oxadiazole-5-carboxamido)-4-oxopentanoic acid heptyl ester;

[0037] (S)-5-Fluoro-3-((R)-5-isopropyl-3-(isoquinolin-1-yl)-4,5-dihydroisoquinolin-1-yl) 4-oxopentanoic acid octyl ester;

[0038] (S)-5-Fluoro-3-((R)-5-isopropyl-3-(isoquinolin-1-yl)-4,5-dihydroisoquinolin-1-yl) 4-oxopentanoic acid decyl ester;

[0039] (S)-5-Fluoro-3-((R)-5-isopropyl-3-(isoquinolin-1-yl)-4,5-dihydroisoquinolin-1-yl) oxadiazole-5-carboxamido)-4-oxopentanoic acid dodecyl ester;

[0040] (S)-5-Fluoro-3-((R)-5-isopropyl-3-(isoquinolin-1-yl)-4,5-dihydroisoquinolin-1-yl) Pentadecyl 4-oxopentanoate;

[0041] (S)-5-Fluoro-3-((R)-5-isopropyl-3-(isoquinolin-1-yl)-4,5-dihydroisoquinolin-1-yl) Octadecyl 4-oxopentanoate;

[0042] (S)-5-Fluoro-3-((R)-5-isopropyl-3-(isoquinolin-1-yl)-4,5-dihydroisoquinolin-1-yl) (9E,12E)-octadeca-9,12-dien-1-yl)-4-oxopentanoate;

[0043] (S)-5-Fluoro-3-((R)-5-isopropyl-3-(isoquinolin-1-yl)-4,5-dihydroisoquinolin-1-yl) 4-oxopentanoic acid cyclopropylmethyl ester;

[0044] (S)-5-Fluoro-3-((R)-5-isopropyl-3-(isoquinolin-1-yl)-4,5-dihydroisoquinolin-1-yl) 4-oxopentanoic acid cyclobutylmethyl ester;

[0045] (S)-5-Fluoro-3-((R)-5-isopropyl-3-(isoquinolin-1-yl)-4,5-dihydroisoquinolin-1-yl) 4-oxopentanoic acid cyclopentylmethyl ester;

[0046] (S)-5-Fluoro-3-((R)-5-isopropyl-3-(isoquinolin-1-yl)-4,5-dihydroisoquinolin-1-yl) Allyl 4-oxopentanoate;

[0047] (S)-5-Fluoro-3-((R)-5-isopropyl-3-(isoquinolin-1-yl)-4,5-dihydroisoquinolin-1-yl) Isopropyl 4-oxopentanoate;

[0048] (S)-5-Fluoro-3-((R)-5-isopropyl-3-(isoquinolin-1-yl)-4,5-dihydroisoquinolin-1-yl) 4-oxopentanoic acid pent-3-yl ester;

[0049] (S)-5-Fluoro-3-((R)-5-isopropyl-3-(isoquinolin-1-yl)-4,5-dihydroisoquinolin-1-yl) 4-oxopentanoic acid sec-butyl ester;

[0050] (S)-5-Fluoro-3-((R)-5-isopropyl-3-(isoquinolin-1-yl)-4,5-dihydroisoquinolin-1-yl) 4-oxopentanoic acid pent-2-yl ester;

[0051] (S)-5-Fluoro-3-((R)-5-isopropyl-3-(isoquinolin-1-yl)-4,5-dihydroisoquinolin-1-yl) oxadiazole-5-carboxamido)-4-oxopentanoic acid hept-2-yl ester;

[0052] (S)-5-Fluoro-3-((R)-5-isopropyl-3-(isoquinolin-1-yl)-4,5-dihydroisoquinolin-1-yl) 4-oxopentanoic acid cyclopentyl ester;

[0053] (S)-5-Fluoro-3-((R)-5-isopropyl-3-(isoquinolin-1-yl)-4,5-dihydroisoquinolin-1-yl) 4-oxopentanoic acid cyclohexyl ester;

[0054] (S)-5-Fluoro-3-((R)-5-isopropyl-3-(isoquinolin-1-yl)-4,5-dihydroisoquinolin-1-yl) Benzyl 4-oxopentanoate;

[0055] (S)-5-Fluoro-3-((R)-5-isopropyl-3-(isoquinolin-1-yl)-4,5-dihydroisoquinolin-1-yl) (5-methyl-2-oxo-1,3-dioxol-4-yl)methyl 4-oxopentanoate;

[0056] (S)-5-Fluoro-3-((R)-5-isopropyl-3-(isoquinolin-1-yl)-4,5-dihydroisoquinolin-1-yl) 2-methoxyphenyl oxadiazole-5-carboxamido)-4-oxopentanoate;

[0057] (S)-5-Fluoro-3-((R)-5-isopropyl-3-(isoquinolin-1-yl)-4,5-dihydroisoquinolin-1-yl) 2,3-dihydro-1H-inden-5-yl 4-oxopentanoate;

[0058] (S)-5-Fluoro-3-((R)-5-isopropyl-3-(isoquinolin-1-yl)-4,5-dihydroisoquinolin-1-yl) oxadiazole-5-carboxamido)-naphthyl-1-4-oxopentanoate;

[0059] (S)-5-Fluoro-3-((R)-5-isopropyl-3-(isoquinolin-1-yl)-4,5-dihydroisoquinolin-1-yl) 4-oxopentanoic acid phenyl ester;

[0060] (S)-5-Fluoro-3-((R)-5-isopropyl-3-(isoquinolin-1-yl)-4,5-dihydroisoquinolin-1-yl) oxadiazole-5-carboxamido)-naphthyl-1-ylmethyl 4-oxopentanoate;

[0061] (S)-5-Fluoro-3-((R)-5-isopropyl-3-(isoquinolin-1-yl)-4,5-dihydroisoquinolin-1-yl) 2,2,2-trifluoroethyl oxadiazole-5-carboxamido)-4-oxopentanoate;

[0062] (S)-5-Fluoro-3-((R)-5-isopropyl-3-(isoquinolin-1-yl)-4,5-dihydroisoquinolin-1-yl) 2-methoxyethyl oxadiazole-5-carboxamido)-4-oxopentanoate;

[0063] (S)-5-Fluoro-3-((R)-5-isopropyl-3-(isoquinolin-1-yl)-4,5-dihydroisoquinolin-1-yl) 2-Fluoroethyl 4-oxopentanoate;

[0064] (S)-5-Fluoro-3-((R)-5-isopropyl-3-(isoquinolin-1-yl)-4,5-dihydroisoquinolin-1-yl) oxadiazole-5-carboxamido)-4-oxopentyl ester;

[0065] (S)-5-Fluoro-3-((R)-5-isopropyl-3-(isoquinolin-1-yl)-4,5-dihydroisoquinolin-1-yl) 4-oxopentanoic acid thiophen-2-ylmethyl ester;

[0066] (S)-5-Fluoro-3-((R)-5-isopropyl-3-(isoquinolin-1-yl)-4,5-dihydroisoquinolin-1-yl) 4-oxopentanoic acid thiophen-3-ylmethyl ester; and

[0067] (S)-5-Fluoro-3-((R)-5-isopropyl-3-(isoquinolin-1-yl)-4,5-dihydroisoquinolin-1-yl) oxadiazole-5-carboxamido)-4-oxopentanoic acid furan-3-ylmethyl ester.

[0068] Unless otherwise specified, the terms and abbreviations used herein retain their original meanings.

[0069] The present invention also provides a method for preparing a compound of formula 1. Hereinafter, the method for preparing a compound of formula 1 is explained based on an exemplary reaction to illustrate the present invention. However, those skilled in the art can prepare the compound of formula 1 by a variety of methods based on the structure of formula 1, and these methods should be interpreted as being within the scope of the present invention. That is, the compound of formula 1 can be prepared by the methods described herein or by combining various methods disclosed in the prior art, and the methods should be interpreted as being within the scope of the present invention. Therefore, the method for preparing a compound of formula 1 is not limited to the following method.

[0070] The compound of formula 1 of the present invention can be prepared from the compound of formula 2 according to the method of the following reaction scheme 1. The compound of formula 1 as a prodrug can be synthesized by using the compound of formula 2 and oxalyl chloride, dimethylformamide (DMF), alcohol and dichloromethane (DCM) solvent, or can be synthesized by using the compound of formula 2 and alkyl halide, potassium carbonate and dimethylformamide solvent, or can be synthesized by using the compound of formula 2 and EDC (3-ethyliminomethylenamino-N,N-dimethylpropane-1-amine) or EDCI (N-ethyl-N′-(3-dimethylaminopropyl) carbodiimide hydrochloride), HOBt (1-hydroxybenzotriazole), triethylamine (Et3N), alkyl alcohol and dichloromethane solvent.

[0071] [Formula 1]

[0072]

[0073] [Formula 2]

[0074]

[0075] [Reaction Scheme 1]

[0076]

[0077] Compounds not specifically described in the production methods of the present specification are known compounds or compounds that can be easily synthesized from known compounds by a known synthetic method or a method similar to that of the known compounds.

[0078] The compound of Formula 1 obtained by the above method can be separated or purified from the reaction product by a conventional method such as recrystallization, ion electrophoresis (ionospheresis), silica gel column chromatography or ion exchange chromatography.

[0079] As explained above, the compounds according to the present invention, the starting materials or intermediates used in their preparation can be prepared by a variety of methods, which should be construed as being within the scope of the present invention with respect to the preparation of the compounds of Formula 1.

[0080] The compound of formula 1 according to the present invention can be used as a prodrug of a caspase inhibitor. Therefore, the present invention provides a pharmaceutical composition for preventing or treating inflammation or apoptosis, the pharmaceutical composition comprising a compound of formula 1 or a pharmaceutically acceptable salt or isomer thereof as an active ingredient, and a pharmaceutically acceptable carrier.

[0081] Exemplary diseases that can be prevented or treated by the pharmaceutical composition according to the present invention include, but are not limited to, diseases selected from apoptosis-related diseases, inflammatory diseases, osteoarthritis, rheumatoid arthritis, degenerative arthritis, and destructive bone diseases.

[0082] In the present invention, "pharmaceutical composition" may include other components such as carriers, diluents, excipients, etc. in addition to the active ingredients of the present invention. Therefore, if necessary, the pharmaceutical composition may include a pharmaceutically acceptable carrier, diluent, excipient or a combination thereof. The pharmaceutical composition helps to administer the compound into the body. Various methods of administering the compound include, but are not limited to, oral, injection, aerosol, parenteral and topical administration.

[0083] As used herein, "carrier" refers to a compound that facilitates the addition of a compound to cells or tissues. For example, dimethyl sulfoxide (DMSO) is a common carrier that facilitates the administration of many organic compounds into living cells or tissues.

[0084] In this article, "diluent" refers to a compound that not only stabilizes the biologically active form but also dilutes it in a solvent that dissolves the compound. Dissolved salts in buffers are used as diluents in this field. A conventionally used buffer is phosphate-buffered saline, which simulates the salt form in body fluids. Because buffer solutions are able to control the pH of a solution at low concentrations, buffer diluents rarely change the biological activity of the compound.

[0085] As used herein, "pharmaceutically acceptable" refers to such properties that do not impair the biological activity and physical properties of the compound.

[0086] The compound according to the present invention can be formulated into various pharmaceutically administered dosage forms. In the preparation of the pharmaceutical composition of the present invention, the active ingredient (specifically, the compound of formula 1 or its pharmaceutically acceptable salt or isomer) is mixed with a pharmaceutically acceptable carrier selected in consideration of the dosage form to be prepared. For example, the pharmaceutical composition of the present invention can be formulated into injections, oral preparations, etc. as required.

[0087] The pharmaceutical composition of the present invention can be formulated into an oral dosage form, an injection form or a patch form, but is not limited thereto.

[0088] The compounds of the present invention can be prepared by conventional methods using known pharmaceutical carriers and excipients, and inserted into a unit or multi-unit container. The preparation can be a solution, suspension or emulsion in an oil or aqueous solvent, and includes conventional dispersants, suspending agents or stabilizers. In addition, the compound can be in the form of a dry powder, for example, dissolved in sterile pyrogen-free water before use. The compounds of the present invention can be formulated into suppositories using conventional suppository bases such as cocoa butter or other glycerides. Solid forms for oral administration include capsules, tablets, pills, powders and granules. Capsules and tablets are preferred. Tablets and pills are preferably enteric coated. The solid form is manufactured by mixing the compound of the present invention with at least one carrier selected from the following: an inert diluent such as sucrose, lactose or starch; a lubricant such as magnesium stearate; a disintegrant; a binder, etc.

[0089] In the case of parenteral preparations, sterile water is usually used and other components such as dissolution aids may also be included. Injection preparations, for example, sterile water-based or oil-based suspensions for injection can be prepared according to known techniques by using suitable dispersants, wetting agents or suspending agents. Solvents that can be used for this purpose include water, Ringer's solution and isotonic NaCl solution, and sterilized fixed oils are usually also used as solvents or suspension media. Any non-irritating fixed oil, including monoglycerides and diglycerides, can be used for this purpose, and fatty acids such as oleic acid can be used for injection preparations. In the case of transdermal preparations, penetration enhancers and / or suitable wetting agents can be used as carriers, optionally combined with suitable additives that are non-irritating to the skin. As this additive, it is possible to select an additive that helps to enhance the application by the skin and / or helps to prepare the desired composition. Transdermal preparations can be used in a variety of ways (for example, such as transdermal patches, drip treatments or ointments).

[0090] The compound according to the present invention or the pharmaceutical composition comprising the same may be administered in combination with other drugs (eg, other caspase inhibitors and / or caspase inhibitor prodrugs).

[0091] The dosage of the compound of formula 1 according to the present invention is determined by the physician's prescription considering the patient's weight, age and the specific condition and severity of the disease. When the compound of the present invention is used for clinical purposes, the total daily dose applied to the host in a single dose or divided doses is preferably in the range of about 5 to 500 mg / kg body weight, but the specific dosage level of a specific patient may change with the patient's weight, sex, health status, diet, drug administration time, method of administration, excretion rate, drug mixture, severity of disease, etc.

[0092] As used herein, the term "treat" or "treating" is used to mean preventing, delaying or ameliorating the progression of a disease in a subject exhibiting symptoms of the disease.

[0093] According to one embodiment of the present invention, the pharmaceutical composition may include microspheres comprising the compound of Formula 1 or a pharmaceutically acceptable salt or isomer and a biocompatible polymer, but is not limited thereto.

[0094] For example, the biocompatible polymer may be selected from: polylactide, polyglycolide, polylactide-co-glycolide, poly(lactide-co-glycolide) glucose, polycaprolactone, gelatin and hyaluronate, and preferably polyglycolide, polylactide or polylactide-co-glycolide (PLGA).

[0095] According to one embodiment of the present invention, the biocompatible polymer may be poly(lactide-co-glycolide) (PLGA) having a molar ratio of lactide to glycolide of 10:90 to 90:10, but is not limited thereto. For example, the molar ratio may preferably be 50:50 to 75:25.

[0096] According to one embodiment of the present invention, the weight ratio of the compound of formula 1 or its pharmaceutically acceptable salt or isomer to the biocompatible polymer in the microsphere may be 1:100 to 70:100, but is not limited thereto. Preferably, the weight ratio of the compound of formula 1 or its pharmaceutically acceptable salt or isomer to the biocompatible polymer may be 1:100 to 17:100. If the weight ratio is lower or higher than the above range, the drug may not be properly encapsulated into the microsphere or problems with microsphere aggregation may occur. For example, compared with the biocompatible polymer, the weight ratio of the compound of formula 1 or its pharmaceutically acceptable salt or isomer in the microsphere may be 5% or more and less than 30%, preferably 10% or more and less than 17%, more preferably about 16.7% by weight, but is not limited thereto.

[0097] For example, the molecular weight of the polylactide-co-glycolide may range from about 1 to 1000 kDa, preferably from about 30 to 150 kDa, and more preferably from about 38 to 54 kDa, but is not limited thereto.

[0098] For example, the terminal group of the polylactide-glycolide copolymer may be an ester or an acid, preferably an ester, but not limited thereto.

[0099] For example, the weight ratio of solid (drug and PLGA) to solvent used to prepare microspheres may be about 5% to 40%, preferably about 10% to 20%, more preferably about 10%, but is not limited thereto.

[0100] For example, the diameter of the microspheres may be about 1 to 250 μm, preferably about 20 to 100 μm, and more preferably about 30 to 70 μm, but is not limited thereto.

[0101] The solvent used to prepare the microspheres can be selected from the group consisting of dichloromethane, dimethyl sulfoxide, dimethylformamide, acetic acid, hydrochloric acid, methanol, ethanol, acetone, ethanol, chloroform, acetonitrile, N-methyl-2-pyrrolidone, tetrahydrofuran, methyl ethyl ketone, propyl acetate, ethyl acetate and methyl acetate.

[0102] In the microsphere preparation step of the present invention, the organic solvent may be removed by applying any conventional solvent removal method (eg, solvent extraction and stirring, heating, solvent evaporation such as nitrogen purge (N2 purge)).

[0103] Beneficial effects

[0104] The present invention relates to a novel compound having a structure of formula 1, which is an isocyanate having a structure of formula 2. Prodrugs of oxazoline derivatives (which are caspase inhibitors). That is, the compound of formula 1 is used as a prodrug of a caspase inhibitor. The prodrug compound having the structure of formula 1 is converted into an active form of a caspase inhibitor of formula 2 in vivo by an esterase isozyme. These prodrug compounds are superior to the caspase inhibitors of formula 2 in terms of pharmacokinetics. Specifically, the prodrug compound of formula 1 has increased drug durability compared to the caspase inhibitors of formula 2. In addition, because the prodrug compound of formula 1 can be converted into a caspase inhibitor of formula 2 by a degradative enzyme in the human body and is hydrophobic in itself, it may be suitable for long-acting preparations. BRIEF DESCRIPTION OF THE DRAWINGS

[0106] Figure 1 is a graph showing the conversion of caspase prodrugs to active forms of caspase inhibitors by hydrolases in rat plasma.

[0107] Figure 2 is a graph showing the mean drug concentration profile in the joints of dogs administered with caspase prodrug.

[0108] Figure 3 This is an image of the appearance of PLGA microspheres encapsulating caspase prodrug observed using a scanning electron microscope.

[0109] Figure 4 are images of the properties of PLGA microspheres prepared by encapsulating caspase inhibitors observed using a scanning electron microscope.

[0110] Figure 5 These are images observing the properties of PLGA microspheres prepared by varying the weight ratio of caspase prodrug and polymer.

[0111] Figure 6 Figure 2 is the in vitro dissolution graph of PLGA microspheres encapsulating caspase prodrugs in PBS and synovial fluid.

[0112] Figure 7 is the in vitro dissolution profile of PLGA microspheres prepared by varying the molar ratio of lactide to glycolide.

[0113] Figure 8 The photographs are taken to observe the properties of the microspheres during the dissolution test of PLGA microspheres prepared by changing the molar ratio of lactide to glycolide.

[0114] Fig. 9 is a graph measuring the change in molecular weight over the course of a microsphere dissolution experiment.

[0115] Fig.10 is a graph showing intra-articular administration of PLGA microspheres prepared according to one embodiment of the present invention to measure the concentration of a caspase inhibitor in synovial fluid. DETAILED DESCRIPTION

[0116] Hereinafter, the present invention will be described in more detail by preparation examples and examples. However, these examples are only illustrative, and the scope of the present invention is not limited thereto.

[0117] Example 1: (S)-5-fluoro-3-((R)-5-isopropyl-3-(isoquinolin-1-yl)-4,5-dihydroisoquinolin-1-yl 5-Azole (acylamino)-4-oxopentanoic acid methyl ester

[0118]

[0119] The compound of formula 2 (5g, 12.0mmol) was dissolved in dichloromethane (50mL), and then oxalyl chloride (1.6mL, 18.0mmol, 1.5 equivalents) and dimethylformamide (0.04mL, 0.6mmol, 0.05 equivalents) were added thereto while the temperature was kept below 5°C. The reaction mixture was stirred at 25°C for about 1 hour and then distilled under reduced pressure. After being dissolved in dichloromethane (50mL), the temperature of the mixture was adjusted to 5°C, and anhydrous methanol (2.0mL, 48.0mmol, 4 equivalents) was added. After the reaction mixture was stirred at 25°C for 2 hours, 10% sodium bicarbonate aqueous solution (30mL) was added to terminate the reaction. The organic layer was separated and distilled under reduced pressure. The obtained mixture was recrystallized in isopropanol to obtain 2.4g (yield: 46%) of the title compound.

[0120] 1H NMR (400MHz, CDCl3) δ9.13(d,1H),8.56(d,1H),7.87(d,1H),7.75-7.67(m,4H),5.21(m,2H),4.95( m,1H),4.05(m,2H),4.01(d,1H),3.82(d,1H),3.59(s,3H),3.01(dd,2H),2.41(m,1H),1.12(dd,6H)

[0121] Example 2: (S)-5-fluoro-3-((R)-5-isopropyl-3-(isoquinolin-1-yl)-4,5-dihydroisoquinolin-1-yl)- 5-Azole (amido)-4-oxopentanoic acid ethyl ester

[0122]

[0123] The compound of formula 2 (5g, 12.0mmol) was dissolved in dichloromethane (50mL), and oxalyl chloride (1.6mL, 18.0mmol, 1.5 equivalents) and dimethylformamide (0.04mL, 0.6mmol, 0.05 equivalents) were then added thereto while the temperature was kept below 5°C. The reaction mixture was stirred at 25°C for about 1 hour and then distilled under reduced pressure. After being dissolved in dichloromethane (50mL), the temperature of the mixture was adjusted to 5°C, and anhydrous ethanol (2.8mL, 48.0mmol, 4 equivalents) was added. After the reaction mixture was stirred at 25°C for 2 hours, 10% sodium bicarbonate aqueous solution (30mL) was added to terminate the reaction. The organic layer was separated and distilled under reduced pressure. The obtained mixture was recrystallized in a 1:2 mixture of ethanol and hexane (EtOH: hexane = 1:2) to obtain 4.1g (yield: 76%) of the title compound.

[0124] 1 H NMR(400MHz, CDCl3)δ9.13(d,1H),8.56(d,1H),7.87(d,1H),7.75-7.67(m,4H),5.21(m,2H), 4.95(m,1H),4.07(m,2H),4.04(d,1H),3.82(d,1H),3.01(dd,2H),2.41(m,1H),1.12(dd,6H)

[0125] Example 3: (S)-5-fluoro-3-((R)-5-isopropyl-3-(isoquinolin-1-yl)-4,5-dihydroisoquinolin-1-yl 5-Azole Amide)-4-oxopentanoic acid propyl ester

[0126]

[0127] The compound of formula 2 (5g, 12.0mmol) was dissolved in dichloromethane (50mL), and oxalyl chloride (1.6mL, 18.0mmol, 1.5 equivalents) and dimethylformamide (0.04mL, 0.6mmol, 0.05 equivalents) were then added thereto while the temperature was kept below 5°C. The reaction mixture was stirred at 25°C for about 1 hour and then distilled under reduced pressure. After being dissolved in dichloromethane (50mL), the temperature of the mixture was adjusted to 5°C, and propanol (3.6mL, 48.0mmol, 4 equivalents) was added. After the reaction mixture was stirred at 25°C for 2 hours, 10% sodium bicarbonate aqueous solution (30mL) was added to terminate the reaction. The organic layer was separated and distilled under reduced pressure. The obtained mixture was recrystallized in a 1:4 mixture of ethanol and hexane (EtOH: hexane = 1:4) to obtain 4.2g (yield: 76%) of the title compound.

[0128] 1 H NMR(400MHz, CDCl3)δ9.14(d,1H),8.56(d,1H),7.87(d,1H),7.75-7.67(m,4H),5.21(m,2H),4.95(m,1H), 4.05(d,1H),3.94(m,2H),3.82(d,1H),3.01(dd,2H),2.41(m,1H),1.54(m,2H),1.11(dd,6H),0.80(t,3H)

[0129] Example 4: (S)-5-fluoro-3-((R)-5-isopropyl-3-(isoquinolin-1-yl)-4,5-dihydroisoquinolin-1-yl 5-Azole (Amino)-4-oxopentanoic acid butyl ester

[0130]

[0131] The compound of formula 2 (5g, 12.0mmol) was dissolved in dichloromethane (50mL), and oxalyl chloride (1.6mL, 18.0mmol, 1.5 equivalents) and dimethylformamide (0.04mL, 0.6mmol, 0.05 equivalents) were then added thereto while the temperature was kept below 5°C. The reaction mixture was stirred at 25°C for about 1 hour and then distilled under reduced pressure. After being dissolved in dichloromethane (50mL), the temperature of the mixture was adjusted to 5°C, and butanol (6.0mL, 48.0mmol, 4 equivalents) was added. After the reaction mixture was stirred at 25°C for 2 hours, 10% sodium bicarbonate aqueous solution (30mL) was added to terminate the reaction. The organic layer was separated and distilled under reduced pressure. The obtained mixture was recrystallized in a 1:4 mixture of ethanol and hexane (EtOH: hexane = 1:4) to obtain 4.1g (yield: 73%) of the title compound.

[0132] 1 H NMR(400MHz, CDCl3)δ9.15(d,1H),8.56(d,1H),7.88(d,1H),7.75-7.67(m,4H),5.21(m,2H),4.95(m,1H),4.04( d,1H),3.95(m,2H),3.82(d,1H),3.01(dd,2H),2.41(m,1H),1.51(m,2H),1.24(m,2H),1.11(dd,6H),0.81(t,3H)

[0133] Example 5: (S)-5-fluoro-3-((R)-5-isopropyl-3-(isoquinolin-1-yl)-4,5-dihydroisoquinolin-1-yl)- 5-Azole Amide)-4-oxopentanoic acid isobutyl ester

[0134]

[0135] The compound of formula 2 (5g, 12.0mmol) was dissolved in dichloromethane (50mL), and oxalyl chloride (1.6mL, 18.0mmol, 1.5 equivalents) and dimethylformamide (0.04mL, 0.6mmol, 0.05 equivalents) were then added thereto while the temperature was kept below 5°C. The reaction mixture was stirred at 25°C for about 1 hour and then distilled under reduced pressure. After being dissolved in dichloromethane (50mL), the temperature of the mixture was adjusted to 5°C, and isobutanol (3.7mL, 48.0mmol, 4 equivalents) was added. After the reaction mixture was stirred at 25°C for 2 hours, 10% sodium bicarbonate aqueous solution (30mL) was added to terminate the reaction. The organic layer was separated and distilled under reduced pressure. The obtained mixture was recrystallized in a 1:4 mixture of ethanol and hexane (EtOH: hexane = 1:4) to obtain 3.7g (yield: 66%) of the title compound.

[0136] 1 H NMR(400MHz, CDCl3)δ9.14(d,1H),8.55(d,1H),7.88(d,1H),7.75-7.67(m,4H),5.20(m,2H),4.95(m,1H), 4.03(d,1H),3.81(d,1H),3.77(m,2H),3.01(dd,2H),2.41(m,1H),1.78(m,2H),1.11(dd,6H),0.77(d,6H)

[0137] Example 6: (S)-5-fluoro-3-((R)-5-isopropyl-3-(isoquinolin-1-yl)-4,5-dihydroisoquinolin-1-yl 5-Azole Amide)-4-oxopentanoic acid isopentyl ester

[0138]

[0139] The compound of formula 2 (5g, 12.0mmol) was dissolved in dichloromethane (50mL), and oxalyl chloride (1.6mL, 18.0mmol, 1.5 equivalents) and dimethylformamide (0.04mL, 0.6mmol, 0.05 equivalents) were then added thereto while the temperature was kept below 5°C. The reaction mixture was stirred at 25°C for about 1 hour and then distilled under reduced pressure. After being dissolved in dichloromethane (50mL), the temperature of the mixture was adjusted to 5°C, and anhydrous isoamyl alcohol (5.3mL, 48.0mmol, 4 equivalents) was added. After the reaction mixture was stirred at 25°C for 2 hours, 10% sodium bicarbonate aqueous solution (30mL) was added to terminate the reaction. The organic layer was separated and distilled under reduced pressure. The obtained mixture was recrystallized in a 1:4 mixture of ethanol and hexane (EtOH: hexane = 1:4) to obtain 4.6g (yield: 72%) of the title compound.

[0140] 1 H NMR(400MHz, CDCl3)δ9.13(d,1H),8.55(d,1H),7.88(d,1H),7.75-7.67(m,4H),5.20(m,2H),4.95(m,1H),4.03( d,1H),3.99(m,2H),3.82(d,1H),3.01(dd,2H),2.41(m,1H),1.53(m,1H),1.39(m,2H),1.11(dd,6H),0.79(d,6H)

[0141] Example 7: (S)-5-fluoro-3-((R)-5-isopropyl-3-(isoquinolin-1-yl)-4,5-dihydroisoquinolin-1-yl)- 5-Azole Amide)-4-oxopentanoic acid pentyl ester

[0142]

[0143] The compound of formula 2 (5g, 12.0mmol) was dissolved in dichloromethane (50mL), and oxalyl chloride (1.6mL, 18.0mmol, 1.5 equivalents) and dimethylformamide (0.04mL, 0.6mmol, 0.05 equivalents) were then added thereto while the temperature was kept below 5°C. The reaction mixture was stirred at 25°C for about 1 hour and then distilled under reduced pressure. After being dissolved in dichloromethane (50mL), the temperature of the mixture was adjusted to 5°C, and amyl alcohol (2.6mL, 48.0mmol, 4 equivalents) was added. After the reaction mixture was stirred at 25°C for 2 hours, 10% sodium bicarbonate aqueous solution (30mL) was added to terminate the reaction. The organic layer was separated and distilled under reduced pressure. The obtained mixture was recrystallized in a 1:10 mixture of ethanol and hexane (EtOH: hexane = 1:10) to obtain 2.1g (yield: 36%) of the title compound.

[0144] 1 H NMR(400MHz, CDCl3)δ9.15(d,1H),8.56(d,1H),7.88(d,1H),7.75-7.66(m,4H),5.21(m,2H),4.95(m,1H),4.04( d,1H),3.97(m,2H),3.82(d,1H),3.01(dd,2H),2.41(m,1H),1.51(m,2H),1.23(m,4H),1.11(dd,6H),0.83(t,3H)

[0145] Example 8: (S)-5-fluoro-3-((R)-5-isopropyl-3-(isoquinolin-1-yl)-4,5-dihydroisoquinolin-1-yl)- 5-Azole Amide)-4-oxopentanoic acid hexyl ester

[0146]

[0147] The compound of formula 2 (5g, 12.0mmol) was dissolved in dichloromethane (50mL), and oxalyl chloride (1.6mL, 18.0mmol, 1.5 equivalents) and dimethylformamide (0.04mL, 0.6mmol, 0.05 equivalents) were then added thereto while the temperature was kept below 5°C. The reaction mixture was stirred at 25°C for about 1 hour and then distilled under reduced pressure. After being dissolved in dichloromethane (50mL), the temperature of the mixture was adjusted to 5°C, and hexanol (6.0mL, 48.0mmol, 4 equivalents) was added. After the reaction mixture was stirred at 25°C for 2 hours, 10% sodium bicarbonate aqueous solution (30mL) was added to terminate the reaction. The organic layer was separated and distilled under reduced pressure. The obtained mixture was recrystallized in a 1:4 mixture of ethanol and hexane (EtOH: hexane = 1:4) to obtain 4.4g (yield: 73%) of the title compound.

[0148] 1 H NMR(400MHz, CDCl3)δ9.15(d,1H),8.56(d,1H),7.88(d,1H),7.75-7.66(m,4H),5.21(m,2H),4.95(m,1H),4.04( d,1H),3.97(m,2H),3.82(d,1H),3.01(dd,2H),2.41(m,1H),1.50(m,2H),1.23(m,6H),1.11(dd,6H),0.84(t,3H)

[0149] Example 9: (S)-5-fluoro-3-((R)-5-isopropyl-3-(isoquinolin-1-yl)-4,5-dihydroisoquinolin-1-yl 5-Azole Amide)-4-oxopentanoic acid heptyl ester

[0150]

[0151] The compound of formula 2 (5g, 12.0mmol) was dissolved in dichloromethane (50mL), and oxalyl chloride (1.6mL, 18.0mmol, 1.5 equivalents) and dimethylformamide (0.04mL, 0.6mmol, 0.05 equivalents) were then added thereto while the temperature was kept below 5°C. The reaction mixture was stirred at 25°C for about 1 hour and then distilled under reduced pressure. After being dissolved in dichloromethane (50mL), the temperature of the mixture was adjusted to 5°C, and anhydrous heptanol (6.8mL, 48.0mmol, 4 equivalents) was added. After the reaction mixture was stirred at 25°C for 2 hours, 10% sodium bicarbonate aqueous solution (30mL) was added to terminate the reaction. The organic layer was separated and distilled under reduced pressure. The obtained mixture was recrystallized in a 1:4 mixture of ethanol and hexane (EtOH: hexane = 1:4) to obtain 4.9g (yield: 79%) of the title compound.

[0152] 1 H NMR(400MHz, CDCl3)δ9.14(d,1H),8.56(d,1H),7.88(d,1H),7.75-7.67(m,4H),5.21(m,2H),4.95(m,1H),4.04( d,1H),3.98(m,2H),3.82(d,1H),3.01(dd,2H),2.41(m,1H),1.49(m,2H),1.20(m,8H),1.11(dd,6H),0.84(t,3H)

[0153] Example 10: (S)-5-fluoro-3-((R)-5-isopropyl-3-(isoquinolin-1-yl)-4,5-dihydroisoquinolin-1-yl)- 5-Azole Amide)-4-oxopentanoic acid octyl ester

[0154]

[0155] The compound of formula 2 (5g, 12.0mmol) was dissolved in dichloromethane (50mL), and oxalyl chloride (1.6mL, 18.0mmol, 1.5 equivalents) and dimethylformamide (0.04mL, 0.6mmol, 0.05 equivalents) were then added thereto while the temperature was kept below 5°C. The reaction mixture was stirred at 25°C for about 1 hour and then distilled under reduced pressure. After being dissolved in dichloromethane (50mL), the temperature of the mixture was adjusted to 5°C, and octanol (7.6mL, 48.0mmol, 4 equivalents) was added. After the reaction mixture was stirred at 25°C for 2 hours, 10% sodium bicarbonate aqueous solution (30mL) was added to terminate the reaction. The organic layer was separated and distilled under reduced pressure. The obtained mixture was recrystallized in a 1:4 mixture of ethanol and hexane (EtOH: hexane = 1:4) to obtain 4.2g (yield: 68%) of the title compound.

[0156] 1 H NMR(400MHz, CDCl3)δ9.15(d,1H),8.56(d,1H),7.88(d,1H),7.75-7.66(m,4H),5.21(m,2H),4.95(m,1H),4.05(d ,1H),3.98(m,2H),3.82(d,1H),3.01(dd,2H),2.41(m,1H),1.49(m,2H),1.25(m,10H),1.11(dd,6H),0.85(t,3H)

[0157] Example 11: (S)-5-fluoro-3-((R)-5-isopropyl-3-(isoquinolin-1-yl)-4,5-dihydroisoquinolin-1-yl)- 5-Azole Amide)-4-oxopentanoic acid decyl ester

[0158]

[0159] The compound of formula 2 (5g, 12.0mmol) was dissolved in dichloromethane (50mL), and oxalyl chloride (1.6mL, 18.0mmol, 1.5 equivalents) and dimethylformamide (0.04mL, 0.6mmol, 0.05 equivalents) were then added thereto while the temperature was kept below 5°C. The reaction mixture was stirred at 25°C for about 1 hour and then distilled under reduced pressure. After being dissolved in dichloromethane (50mL), the temperature of the mixture was adjusted to 5°C, and decanol (9.2mL, 48.0mmol, 4 equivalents) was added. After the reaction mixture was stirred at 25°C for 2 hours, 10% sodium bicarbonate aqueous solution (30mL) was added to terminate the reaction. The organic layer was separated and distilled under reduced pressure. The obtained mixture was recrystallized in a 1:4 mixture of ethanol and hexane (EtOH: hexane = 1:4) to obtain 1.71g (yield: 27%) of the title compound.

[0160] 1 H NMR(400MHz, CDCl3)δ9.14(d,1H),8.55(d,1H),7.88(d,1H),7.75-7.66(m,4H),5.20(m,2H),4.95(m,1H),4.04(d,1 H),3.98(m,2H),3.81(d,1H),3.00(dd,2H),2.41(m,1H),1.47(m,2H),1.28-1.17(m,14H),1.10(dd,6H),0.87(t,3H)

[0161] Example 12: (S)-5-fluoro-3-((R)-5-isopropyl-3-(isoquinolin-1-yl)-4,5-dihydroisoquinolin-1-yl)- 5-Azole Amide)-4-oxopentanoic acid dodecyl ester

[0162]

[0163] The compound of formula 2 (5g, 12.0mmol) was dissolved in dichloromethane (50mL), and oxalyl chloride (1.6mL, 18.0mmol, 1.5 equivalents) and dimethylformamide (0.04mL, 0.6mmol, 0.05 equivalents) were then added thereto while the temperature was kept below 5°C. The reaction mixture was stirred at 25°C for about 1 hour and then distilled under reduced pressure. After being dissolved in dichloromethane (50mL), the temperature of the mixture was adjusted to 5°C, and dodecanol (5.4mL, 24.0mmol, 2 equivalents) was added. After the reaction mixture was stirred at 25°C for 2 hours, 10% sodium bicarbonate aqueous solution (30mL) was added to terminate the reaction. The organic layer was separated and distilled under reduced pressure. The obtained mixture was recrystallized in a 1:4 mixture of ethanol and hexane (EtOH: hexane = 1:4) to obtain 2.4g (yield: 34%) of the title compound.

[0164] 1 H NMR(400MHz, CDCl3)δ9.15(d,1H),8.56(d,1H),7.88(d,1H),7.75-7.66(m,4H),5.21(m,2H),4.95(m,1H),4.05(d,1 H),3.97(m,2H),3.82(d,1H),3.00(dd,2H),2.41(m,1H),1.48(m,2H),1.30-1.18(m,18H),1.11(dd,6H),0.88(t,3H)

[0165] Example 13: (S)-5-fluoro-3-((R)-5-isopropyl-3-(isoquinolin-1-yl)-4,5-dihydroisoquinolin-1-yl)- 5-Azole Amide)-4-oxopentanoic acid pentadecyl ester

[0166]

[0167] Formula 2 compound (5g, 12.0mmol) is dissolved in dichloromethane (50mL), and then oxalyl chloride (1.6mL, 18.0mmol, 1.5 equivalents) and dimethylformamide (0.04mL, 0.6mmol, 0.05 equivalents) are added thereto while the temperature is kept below 5°C. The reaction mixture is stirred at 25°C for about 1 hour and then distilled under reduced pressure. After being dissolved in dichloromethane (50mL), the temperature of the mixture is adjusted to 5°C, and pentadecyl alcohol (11.0g, 48.0mmol, 4 equivalents) is added. After the reaction mixture is stirred at 25°C for 2 hours, 10% sodium bicarbonate aqueous solution (30mL) is added to terminate the reaction. The organic layer is separated and distilled under reduced pressure. The obtained mixture is subjected to column separation by using a 1:2 mixture of ethyl acetate and hexane (EtOAc: hexane = 1:2) to obtain 5.6g (yield: 75%) of the title compound.

[0168] 1 H NMR (400MHz, CDCl3) δ9.14(d,1H),8.55(d,1H),7.88(d,1H),7.75-7.66(m,4H),5.21(m,2H),4.95(m,1H),4.04(d,1 H),3.99(m,2H),3.81(d,1H),3.01(dd,2H),2.41(m,1H),1.48(m,2H),1.30-1.13(m,24H),1.11(dd,6H),0.89(t,3H)

[0169] Example 14: (S)-5-fluoro-3-((R)-5-isopropyl-3-(isoquinolin-1-yl)-4,5-dihydroisoquinolin-1-yl)- 5-Azole Amide)-4-oxopentanoic acid octadecyl ester

[0170]

[0171] Formula 2 compound (5g, 12.0mmol) is dissolved in dichloromethane (50mL), and then oxalyl chloride (1.6mL, 18.0mmol, 1.5 equivalents) and dimethylformamide (0.04mL, 0.6mmol, 0.05 equivalents) are added thereto while the temperature is kept below 5°C. The reaction mixture is stirred at 25°C for about 1 hour and then distilled under reduced pressure. After being dissolved in dichloromethane (50mL), the temperature of the mixture is adjusted to 5°C, and octadecyl alcohol (6.5g, 48.0mmol, 4 equivalents) is added. After the reaction mixture is stirred at 25°C for 2 hours, 10% sodium bicarbonate aqueous solution (30mL) is added to terminate the reaction. The organic layer is separated and distilled under reduced pressure. The obtained mixture is subjected to column separation by using a 1:2 mixture of ethyl acetate and hexane (EtOAc: hexane = 1:2) to obtain 2.1g (yield: 26%) of the title compound.

[0172] 1 H NMR(400MHz, CDCl3)δ9.15(d,1H),8.56(d,1H),7.88(d,1H),7.75-7.66(m,4H),5.21(m,2H),4.95(m,1H),4.06(d,1 H),3.97(m,2H),3.82(d,1H),3.00(dd,2H),2.41(m,1H),1.48(m,2H),1.30-1.18(m,30H),1.10(dd,6H),0.88(t,3H)

[0173] Example 15: (S)-5-fluoro-3-((R)-5-isopropyl-3-(isoquinolin-1-yl)-4,5-dihydroisoquinolin-1-yl 5-Azole (9E,12E)-octadec-9,12-dien-1-yl 4-oxopentanoate;

[0174]

[0175] Formula 2 compound (5g, 12.0mmol) is dissolved in dichloromethane (50mL), and then oxalyl chloride (1.6mL, 18.0mmol, 1.5 equivalents) and dimethylformamide (0.04mL, 0.6mmol, 0.05 equivalents) are added thereto while the temperature is maintained below 5°C. The reaction mixture is stirred at 25°C for about 1 hour and then distilled under reduced pressure. After being dissolved in dichloromethane (50mL), the temperature of the mixture is adjusted to 5°C, and linoleic alcohol (11.0g, 48.0mmol, 4 equivalents) is added. After the reaction mixture is stirred at 25°C for 2 hours, 10% sodium bicarbonate aqueous solution (30mL) is added to terminate the reaction. The organic layer is separated and distilled under reduced pressure. The obtained mixture is subjected to column separation by using a 1:2 mixture of ethyl acetate and hexane (EtOAc: hexane = 1:2) to obtain 0.1g of the title compound.

[0176] 1 H NMR(400MHz, CDCl3)δ9.14(d,1H),8.56(d,1H),7.88(d,1H),7.75-7.66(m,4H),5.35(m,4H),5.21(m,2H),4.95(m,1H),4.05(d,1H),3.9 8(m,2H),3.82(d,1H),3.00(dd,2H),2.78(t,2H),2.41(m,1H),2.04(m,4H),1.48(m,2H),1.33-1.18(m,16H),1.11(dd,6H),0.88(t,3H)

[0177] Example 16: (S)-5-fluoro-3-((R)-5-isopropyl-3-(isoquinolin-1-yl)-4,5-dihydroisoquinolin-1-yl)- 5-Azole Cyclopropyl methyl 4-oxopentanoate

[0178]

[0179] The compound of formula 2 (5g, 12.0mmol) was dissolved in dichloromethane (50mL), and oxalyl chloride (1.6mL, 18.0mmol, 1.5 equivalents) and dimethylformamide (0.04mL, 0.6mmol, 0.05 equivalents) were then added thereto while the temperature was kept below 5°C. The reaction mixture was stirred at 25°C for about 1 hour and then distilled under reduced pressure. After being dissolved in dichloromethane (50mL), the temperature of the mixture was adjusted to 5°C, and cyclopropylmethanol (3.9mL, 48.0mmol, 4 equivalents) was added. After the reaction mixture was stirred at 25°C for 2 hours, 10% sodium bicarbonate aqueous solution (30mL) was added to terminate the reaction. The organic layer was separated and distilled under reduced pressure. The obtained mixture was recrystallized in a 1:4 mixture of ethanol and hexane (EtOH: hexane = 1:4) to obtain 4.3g (yield: 76%) of the title compound.

[0180] 1 H NMR(400MHz, CDCl3)δ9.14(d,1H),8.56(d,1H),7.88(d,1H),7.76-7.66(m,4H),5.21(m,2H),4.95(m,1H),4.05( d,1H),3.84(m,2H),3.82(d,1H),3.01(dd,2H),2.41(m,1H),1.10(dd,6H),0.98(m,1H),0.45(d,2H),0.15(d,2H)

[0181] Example 17: (S)-5-fluoro-3-((R)-5-isopropyl-3-(isoquinolin-1-yl)-4,5-dihydroisoquinolin-1-yl)- 5-Azole Cyclobutylmethyl 4-oxopentanoate

[0182]

[0183] The compound of formula 2 (5g, 12.0mmol) was dissolved in dichloromethane (50mL), and oxalyl chloride (1.6mL, 18.0mmol, 1.5 equivalents) and dimethylformamide (0.04mL, 0.6mmol, 0.05 equivalents) were then added thereto while the temperature was kept below 5°C. The reaction mixture was stirred at 25°C for about 1 hour and then distilled under reduced pressure. After being dissolved in dichloromethane (50mL), the temperature of the mixture was adjusted to 5°C, and cyclobutylmethanol (5.1mL, 48.0mmol, 4 equivalents) was added. After the reaction mixture was stirred at 25°C for 2 hours, 10% sodium bicarbonate aqueous solution (30mL) was added to terminate the reaction. The organic layer was separated and distilled under reduced pressure. The obtained mixture was recrystallized in a 1:4 mixture of ethanol and hexane (EtOH: hexane = 1:4) to obtain 3.9g (yield: 67%) of the title compound.

[0184] 1 H NMR(400MHz, CDCl3)δ9.15(d,1H),8.56(d,1H),7.88(d,1H),7.75-7.66(m,4H),5.21(m,2H),4.95(m,1H),4.05( d,1H),3.96(m,2H),3.82(d,1H),3.01(dd,2H),2.42(m,2H),1.92(m,2H),1.75(m,2H),1.58(m,2H)1.10(dd,6H)

[0185] Example 18: (S)-5-fluoro-3-((R)-5-isopropyl-3-(isoquinolin-1-yl)-4,5-dihydroisoquinolin-1-yl)- 5-Azole Cyclopentylmethyl 4-oxopentanoate

[0186]

[0187] The compound of formula 2 (5g, 12.0mmol) was dissolved in dichloromethane (50mL), and oxalyl chloride (1.6mL, 18.0mmol, 1.5 equivalents) and dimethylformamide (0.04mL, 0.6mmol, 0.05 equivalents) were then added thereto while the temperature was kept below 5°C. The reaction mixture was stirred at 25°C for about 1 hour and then distilled under reduced pressure. After being dissolved in dichloromethane (50mL), the temperature of the mixture was adjusted to 5°C, and cyclopentylmethanol (5.2mL, 48.0mmol, 4 equivalents) was added. After the reaction mixture was stirred at 25°C for 2 hours, 10% sodium bicarbonate aqueous solution (30mL) was added to terminate the reaction. The organic layer was separated and distilled under reduced pressure. The obtained mixture was recrystallized in a 1:4 mixture of ethanol and hexane (EtOH: hexane = 1:4) to obtain 4.2g (yield: 69%) of the title compound.

[0188] 1 H NMR(400MHz, CDCl3)δ9.15(d,1H),8.56(d,1H),7.88(d,1H),7.75-7.66(m,4H),5.21(m,2H),4.95(m,1H),4.04( d,1H),3.86(m,2H),3.82(d,1H),3.01(dd,2H),2.42(m,1H),2.03(m,1H),1.59(m,4H),1.43(m,4H)1.10(dd,6H)

[0189] Example 19: (S)-5-fluoro-3-((R)-5-isopropyl-3-(isoquinolin-1-yl)-4,5-dihydroisoquinolin-1-yl 5-Azole Amide)-4-oxopentanoic acid allyl ester

[0190]

[0191] The compound of formula 2 (5g, 12.0mmol) was dissolved in dichloromethane (50mL), and oxalyl chloride (1.6mL, 18.0mmol, 1.5 equivalents) and dimethylformamide (0.04mL, 0.6mmol, 0.05 equivalents) were then added thereto while the temperature was kept below 5°C. The reaction mixture was stirred at 25°C for about 1 hour and then distilled under reduced pressure. After being dissolved in dichloromethane (50mL), the temperature of the mixture was adjusted to 5°C, and allyl alcohol (2.0mL, 48.0mmol, 4 equivalents) was added. After the reaction mixture was stirred at 25°C for 2 hours, 10% sodium bicarbonate aqueous solution (30mL) was added to terminate the reaction. The organic layer was separated and distilled under reduced pressure. The obtained mixture was recrystallized in a 1:4 mixture of ethanol and hexane (EtOH: hexane = 1:4) to obtain 4.3g (yield: 78%) of the title compound.

[0192] 1 H NMR (400MHz, CDCl3) δ9.14(d,1H),8.56(d,1H),7.88(d,1H),7.75-7.66(m,4H),5.75(m,1H),5.21( m,4H),4.95(m,1H),4.50(m,2H),4.04(d,1H),3.82(d,1H),3.01(dd,2H),2.42(m,1H),1.10(dd,6H)

[0193] Example 20: (S)-5-fluoro-3-((R)-5-isopropyl-3-(isoquinolin-1-yl)-4,5-dihydroisoquinolin-1-yl)- 5-Azole Amide)-4-oxopentanoic acid isopropyl ester

[0194]

[0195] Formula 2 compound (0.5g, 1.2mmol) is dissolved in dimethylformamide (5mL), and isopropyl bromide (0.18mL, 1.8mmol, 1.5 equivalents) and potassium carbonate (0.2g, 1.8mmol, 1.2 equivalents) are then added thereto. The reaction mixture is stirred at 25°C for about 18 hours, diluted in ethyl acetate (EtOAc, 30mL), and 10% sodium bicarbonate aqueous solution (30mL) is added and reacted under stirring. After adding water (30ml) and stirring, the organic layer is separated and distilled under reduced pressure. The obtained mixture is subjected to column separation by using a 1:2 mixture of ethyl acetate and hexane (EtOAc: hexane = 1:2) to obtain 3.6g (yield: 27%) of the title compound.

[0196] 1 H NMR(400MHz, CDCl3)δ9.14(d,1H),8.56(d,1H),7.88(d,1H),7.75-7.66(m,4H),5.15(m,3H) ,4.95(m,1H),4.03(d,1H),3.82(d,1H),3.01(dd,2H),2.40(m,1H),1.23(m,6H)1.11(dd,6H)

[0197] Example 21: (S)-5-fluoro-3-((R)-5-isopropyl-3-(isoquinolin-1-yl)-4,5-dihydroisoquinolin-1-yl)- 5-Azole Amide)-4-oxopentanoic acid pent-3-yl ester

[0198]

[0199] Formula 2 compound (0.5g, 1.2mmol) is dissolved in dimethylformamide (5mL), and 3-bromopentane (0.22mL, 1.8mmol, 1.5 equivalents) and potassium carbonate (0.2g, 1.8mmol, 1.2 equivalents) are then added thereto. The reaction mixture is stirred at 25 ° C for about 18 hours, diluted in ethyl acetate (EtOAc, 30mL), and 10% sodium bicarbonate aqueous solution (30mL) is added and reacted under stirring. After adding water (30ml) and stirring, the organic layer is separated and distilled under reduced pressure. The obtained mixture is subjected to column separation by using a 1:2 mixture of ethyl acetate and hexane (EtOAc: hexane=1:2) to obtain 0.05g (yield: 9%) of the title compound.

[0200] 1 H NMR(400MHz, CDCl3)δ9.14(d,1H),8.56(d,1H),7.88(d,1H),7.75-7.66(m,4H),5.20(m,2H),4.95(m,1H), 4.76(m,1H),4.03(d,1H),3.82(d,1H),3.01(dd,2H),2.40(m,1H),1.54(m,4H),1.10(dd,6H),0.86(t,6H)

[0201] Example 22: (S)-5-fluoro-3-((R)-5-isopropyl-3-(isoquinolin-1-yl)-4,5-dihydroisoquinolin-1-yl)- 5-Azole 4-Oxopentanoic acid sec-butyl ester

[0202]

[0203] Formula 2 compound (0.5g, 1.2mmol) is dissolved in dimethylformamide (5mL), and 2-bromobutane (0.18mL, 1.8mmol, 1.5 equivalents) and potassium carbonate (0.2g, 1.8mmol, 1.2 equivalents) are then added thereto. The reaction mixture is stirred at 25°C for about 18 hours, diluted in ethyl acetate (EtOAc, 30mL), and 10% sodium bicarbonate aqueous solution (30mL) is added and reacted under stirring. After adding water (30ml) and stirring, the organic layer is separated and distilled under reduced pressure. The obtained mixture is subjected to column separation by using a 1:2 mixture of ethyl acetate and hexane (EtOAc: hexane=1:2) to obtain 0.5g (yield: 88%) of the title compound.

[0204] 1 H NMR(400MHz, CDCl3)δ9.15(d,1H),8.56(d,1H),7.88(d,1H),7.75-7.66(m,4H),5.20(m,2H),4.96(m,1H),4.86( m,1H),4.03(d,1H),3.82(d,1H),3.01(dd,2H),2.40(m,1H),1.54(m,2H),1.21(m,3H),1.10(dd,6H),0.89(t,3H)

[0205] Example 23: (S)-5-fluoro-3-((R)-5-isopropyl-3-(isoquinolin-1-yl)-4,5-dihydroisoquinolin-1-yl)- 5-Azole Amide)-4-oxopentanoic acid pent-2-yl ester

[0206]

[0207] Formula 2 compound (0.5g, 1.2mmol) is dissolved in dimethylformamide (5mL), and 1-bromopentane (0.19mL, 1.8mmol, 1.5 equivalents) and potassium carbonate (0.2g, 1.8mmol, 1.2 equivalents) are then added thereto. The reaction mixture is stirred at 25 ° C for about 18 hours, diluted in ethyl acetate (EtOAc, 30mL), and 10% sodium bicarbonate aqueous solution (30mL) is added and reacted under stirring. After adding water (30ml) and stirring, the organic layer is separated and distilled under reduced pressure. The obtained mixture is subjected to column separation by using a 1:2 mixture of ethyl acetate and hexane (EtOAc: hexane=1:2) to obtain 0.14g (yield: 24%) of the title compound.

[0208] 1H NMR(400MHz, CDCl3)δ9.15(d,1H),8.56(d,1H),7.88(d,1H),7.77-7.66(m,4H),5.20(m,2H),4.96(m,1H),4.86(m,1H), 4.04(d,1H),3.82(d,1H),3.01(dd,2H),2.40(m,1H),1.51(m,2H),1.40(m,2H),1.23(m,3H),1.10(dd,6H),0.89(t,3H)

[0209] Example 24: (S)-5-fluoro-3-((R)-5-isopropyl-3-(isoquinolin-1-yl)-4,5-dihydroisoquinolin-1-yl 5-Azole Amide)-4-oxopentanoic acid hept-2-yl ester

[0210]

[0211] Formula 2 compound (0.5g, 1.2mmol) is dissolved in dimethylformamide (5mL), and 3-bromoheptane (0.18mL, 1.8mmol, 1.5 equivalents) and potassium carbonate (0.2g, 1.8mmol, 1.2 equivalents) are then added thereto. The reaction mixture is stirred for about 18 hours at 25°C, diluted in ethyl acetate (EtOAc, 30mL), and 10% sodium bicarbonate aqueous solution (30mL) is added and reacted under stirring. After adding water (30ml) and stirring, the organic layer is separated and distilled under reduced pressure. The obtained mixture is subjected to column separation by using a 1:2 mixture of ethyl acetate and hexane (EtOAc: hexane=1:2) to obtain the title compound of 0.12g (yield: 20%).

[0212] 1 H NMR(400MHz, CDCl3)δ9.15(d,1H),8.56(d,1H),7.88(d,1H),7.77-7.66(m,4H),5.21(m,2H),4.96(m,1H),4.84(m,1H), 4.05(d,1H),3.82(d,1H),3.01(dd,2H),2.41(m,1H),1.49(m,2H),1.30(m,4H),1.22(m,3H),1.11(dd,6H),0.89(t,3H)

[0213] Example 25: (S)-5-fluoro-3-((R)-5-isopropyl-3-(isoquinolin-1-yl)-4,5-dihydroisoquinolin-1-yl)- 5-Azole Amide)-4-oxopentanoic acid cyclopentyl ester

[0214]

[0215] Formula 2 compound (0.5g, 1.2mmol) is dissolved in dimethylformamide (5mL), and 3-cyclopentyl bromide (0.19mL, 1.8mmol, 1.5 equivalents) and potassium carbonate (0.2g, 1.8mmol, 1.2 equivalents) are then added thereto. The reaction mixture is stirred at 25°C for about 18 hours, diluted in ethyl acetate (EtOAc, 30mL), and 10% sodium bicarbonate aqueous solution (30mL) is added and reacted under stirring. After adding water (30ml) and stirring, the organic layer is separated and distilled under reduced pressure. The obtained mixture is subjected to column separation by using a 1:2 mixture of ethyl acetate and hexane (EtOAc: hexane=1:2) to obtain 0.20g (yield: 35%) of the title compound.

[0216] 1 H NMR(400MHz, CDCl3)δ9.15(d,1H),8.56(d,1H),7.88(d,1H),7.77-7.66(m,4H),5.21(m,2H),4.96(m,1 H),4.84(m,1H),4.05(d,1H),3.84(d,1H),3.06(dd,2H),2.41(m,1H),1.72-1.23(m,8H),1.11(dd,6H)

[0217] Example 26: (S)-5-fluoro-3-((R)-5-isopropyl-3-(isoquinolin-1-yl)-4,5-dihydroisoquinolin-1-yl)- 5-Azole Amide)-4-oxopentanoic acid cyclohexyl ester

[0218]

[0219] Formula 2 compound (0.5g, 1.2mmol) is dissolved in dimethylformamide (5mL), and 3-cyclohexyl iodide (0.23mL, 1.8mmol, 1.5 equivalents) and potassium carbonate (0.2g, 1.8mmol, 1.2 equivalents) are then added thereto. The reaction mixture is stirred at 25 ° C for about 18 hours, diluted in ethyl acetate (EtOAc, 30mL), and 10% sodium bicarbonate aqueous solution (30mL) is added and reacted under stirring. After adding water (30ml) and stirring, the organic layer is separated and distilled under reduced pressure. The obtained mixture is subjected to column separation by using a 1:2 mixture of ethyl acetate and hexane (EtOAc: hexane=1:2) to obtain 0.06g (yield: 10%) of the title compound.

[0220] 1H NMR(400MHz, CDCl3)δ9.15(d,1H),8.56(d,1H),7.88(d,1H),7.77-7.66(m,4H),5.21(m,2H),4.96(m,1 H),4.84(m,1H),4.05(d,1H),3.83(d,1H),3.07(dd,2H),2.41(m,1H),1.73-1.22(m,10H),1.11(dd,6H)

[0221] Example 27: (S)-5-fluoro-3-((R)-5-isopropyl-3-(isoquinolin-1-yl)-4,5-dihydroisoquinolin-1-yl 5-Azole Benzyl 4-oxopentanoate

[0222]

[0223] Formula 2 compound (0.5g, 1.2mmol) is dissolved in dimethylformamide (5mL), and benzyl bromide (0.18mL, 1.8mmol, 1.5 equivalents) and potassium carbonate (0.2g, 1.8mmol, 1.2 equivalents) are then added thereto. The reaction mixture is stirred at 25°C for about 18 hours, diluted in ethyl acetate (EtOAc, 30mL), and 10% sodium bicarbonate aqueous solution (30mL) is added and reacted under stirring. After adding water (30ml) and stirring, the organic layer is separated and distilled under reduced pressure. The obtained mixture is subjected to column separation by using a 1:2 mixture of ethyl acetate and hexane (EtOAc: hexane=1:2) to obtain 0.5g (yield: 78%) of the title compound.

[0224] 1 H NMR(400MHz, CDCl3)δ9.14(d,1H),8.56(d,1H),7.88(d,1H),7.75-7.66(m,4H),7.36(m,3H),7.22(m,2H), 5.17(m,2H),5.05(m,2H),4.95(m,1H),4.05(d,1H),3.82(d,1H),3.01(dd,2H),2.40(m,1H),1.11(dd,6H)

[0225] Example 28: (S)-5-fluoro-3-((R)-5-isopropyl-3-(isoquinolin-1-yl)-4,5-dihydroisoquinolin-1-yl)- 5-Azole (5-methyl-2-oxo-1,3-dioxol-4-yl)-4-oxopentanoic acid methyl ester

[0226]

[0227] The compound of formula 2 (0.5g, 1.2mmol) was dissolved in dimethylformamide (5mL), and 4-chloromethyl-5-methyl-1,3-dioxol-2-one (0.19g, 1.8mmol, 1.5 equivalents) and potassium carbonate (0.2g, 1.8mmol, 1.2 equivalents) were then added thereto. The reaction mixture was stirred at 25°C for about 18 hours, diluted in ethyl acetate (EtOAc, 30mL), and 10% sodium bicarbonate aqueous solution (30mL) was added and reacted under stirring. After adding water (30ml) and stirring, the organic layer was separated and distilled under reduced pressure. The obtained mixture was subjected to column separation by using a 1:2 mixture of ethyl acetate and hexane (EtOAc: hexane = 1:2) to obtain 0.21g (yield: 33%) of the title compound.

[0228] 1 H NMR(400MHz, CDCl3)δ9.14(d,1H),8.57(d,1H),7.89(d,1H),7.75-7.66(m,4H),5.17(m,2H), 4.87(m,1H),4.42(s,2H),4.06(d,1H),3.00(dd,2H),2.39(m,1H),2.18(s,3H),1.11(dd,6H)

[0229] Example 29: (S)-5-fluoro-3-((R)-5-isopropyl-3-(isoquinolin-1-yl)-4,5-dihydroisoquinolin-1-yl 5-Azole Amide)-4-oxopentanoic acid 2-methoxyphenyl ester

[0230]

[0231] The compound of formula 2 (10 g, 24.0 mmol) was dissolved in dichloromethane (100 mL), and then 2-methoxyphenol (11.9 g, 96.0 mmol, 4 equivalents), hydroxybenzotriazole (HOBt, 0.64 g, 0.48 mmol, 0.2 equivalents) and triethylamine (0.6 mL, 0.48 mmol, 0.2 equivalents) were added, and N-ethyl-N'-(3-dimethylaminopropyl) carbodiimide hydrochloride (EDCI, 5.5 g, 28.8 mmol, 1.2 equivalents) was added thereto while keeping the temperature below 5 ° C. The reaction mixture was stirred at 25 ° C for about 6 hours, and 10% sodium bicarbonate aqueous solution (50 mL) was added to terminate the reaction. After adding water (50 mL) and stirring, the organic layer was separated and distilled under reduced pressure. The resulting mixture was subjected to column separation by using a 1:2 mixture of ethyl acetate and hexane (EtOAc:hexane=1:2) to obtain 3.6 g (yield: 27%) of the title compound.

[0232] 1H NMR(400MHz, CDCl3)δ9.15(d,1H),8.57(d,1H),7.88(d,1H),7.77-7.66(m,4H),7.19(m,2H),6.94(m,2H), 5.16(m,1H),4.58(m,2H),4.10(d,1H),3.91(s,3H),3.84(d,1H),3.01(dd,2H),2.44(m,1H),1.11(dd,6H)

[0233] Example 30: (S)-5-fluoro-3-((R)-5-isopropyl-3-(isoquinolin-1-yl)-4,5-dihydroisoquinolin-1-yl)- 5-Azole Amide)-4-oxopentanoic acid 2,3-dihydro-1H-inden-5-yl ester

[0234]

[0235] Formula 2 compound (10g, 24.0mmol) is dissolved in dichloromethane (100mL), then 5-indazole (12.8g, 96.0mmol, 4 equivalents), hydroxybenzotriazole (0.64g, 0.48mmol, 0.2 equivalents) and triethylamine (0.6mL, 0.48mmol, 0.2 equivalents), and EDCI (5.5g, 28.8mmol, 1.2 equivalents) is added thereto while the temperature is kept below 5°C. The reaction mixture is stirred at 25°C for about 6 hours, and 10% sodium bicarbonate aqueous solution (50mL) is added to terminate the reaction. After adding water (50mL) and stirring, the organic layer is separated and distilled under reduced pressure. The obtained mixture is subjected to column separation by using a 1:2 mixture of ethyl acetate and hexane (EtOAc: hexane = 1:2) to obtain 3.4g (yield: 28%) of the title compound.

[0236] 1 H NMR(400MHz, CDCl3)δ9.15(d,1H),8.57(d,1H),7.88(d,1H),7.77-7.66(m,4H),7.18(d,1H),7.03(s,1H),6.93(d,1H), 5.17(m,1H),4.63(m,2H),4.15(d,1H),3.85(d,1H),3.04(dd,2H),2.90(m,4H),2.45(m,1H),2.13(m,2H),1.11(dd,6H)

[0237] Example 31: (S)-5-fluoro-3-((R)-5-isopropyl-3-(isoquinolin-1-yl)-4,5-dihydroisoquinolin-1-yl)- 5-Azole Amide)-1-naphthyl 4-oxopentanoate

[0238]

[0239] Formula 2 compound (0.5g, 1.2mmol) is dissolved in dichloromethane (10mL), then 1-naphthol (0.69g, 4.8mmol, 4 equivalents), hydroxybenzotriazole (0.64g, 0.24mmol, 0.2 equivalents) and triethylamine (0.6mL, 0.24mmol, 0.2 equivalents), and EDCI (0.28g, 1.4mmol, 1.2 equivalents) is added thereto while the temperature is kept below 5°C. The reaction mixture is stirred at 25°C for about 6 hours, and 10% sodium bicarbonate aqueous solution (10mL) is added to terminate the reaction. After adding water (10mL) and stirring, the organic layer is separated and distilled under reduced pressure. The obtained mixture is subjected to column separation by using a 1:2 mixture of ethyl acetate and hexane (EtOAc: hexane = 1:2) to obtain 0.07g (yield: 10%) of the title compound.

[0240] 1 H NMR(400MHz, CDCl3)δ9.15(d,1H),8.57(d,1H),8.25(d,1H),7.95(d,1H),7.88(d,1H),7.74-7.57(m,7H), 7.47(m,2H),5.32(m,1H),4.66(m,2H),4.13(d,1H),3.88(d,1H),3.10(dd,2H),2.54(m,1H),1.11(dd,6H)

[0241] Example 32: (S)-5-fluoro-3-((R)-5-isopropyl-3-(isoquinolin-1-yl)-4,5-dihydroisoquinolin-1-yl)- 5-Azole Amide)-4-oxopentanoic acid phenyl ester

[0242]

[0243] Formula 2 compound (10g, 24.0mmol) is dissolved in dichloromethane (100mL), then phenol (9.0g, 96.0mmol, 4 equivalents), hydroxybenzotriazole (0.64g, 0.48mmol, 0.2 equivalents) and triethylamine (0.6mL, 0.48mmol, 0.2 equivalents) are added, and EDCI (5.5g, 28.8mmol, 1.2 equivalents) is added thereto while the temperature is kept below 5°C. The reaction mixture is stirred at 25°C for about 6 hours, and 10% sodium bicarbonate aqueous solution (50mL) is added to terminate the reaction. After adding water (50mL) and stirring, the organic layer is separated and distilled under reduced pressure. The obtained mixture is recrystallized in a 1:5 mixture of ethanol and hexane (EtOH: hexane = 1:5) to obtain 2.9g (yield: 25%) of the title compound.

[0244] 1 H NMR(400MHz, CDCl3)δ9.15(d,1H),8.57(d,1H),7.88(d,1H),7.77-7.66(m,4H),7.38(m,2H),7.20( m,3H),5.19(m,1H),4.62(m,2H),4.11(d,1H),3.82(d,1H),3.03(dd,2H),2.45(m,1H),1.11(dd,6H)

[0245] Example 33: (S)-5-fluoro-3-((R)-5-isopropyl-3-(isoquinolin-1-yl)-4,5-dihydroisoquinolin-1-yl)- 5-Azole (amido)-4-oxopentanoic acid naphth-1-ylmethyl ester

[0246]

[0247] Formula 2 compound (0.5g, 1.2mmol) is dissolved in dichloromethane (10mL), then 1-naphthalene methanol (0.38g, 4.8mmol, 4 equivalents), 4-dimethylaminopyridine (DMAP, 0.03g, 0.24mmol, 0.2 equivalents) and triethylamine (0.6mL, 0.24mmol, 0.2 equivalents), and EDCI (0.28g, 1.4mmol, 1.2 equivalents) is added thereto while the temperature is kept below 5°C. The reaction mixture is stirred at 25°C for about 6 hours, and 10% sodium bicarbonate aqueous solution (10mL) is added to terminate the reaction. After adding water (10mL) and stirring, the organic layer is separated and distilled under reduced pressure. The obtained mixture is subjected to column separation by using a 1:2 mixture of ethyl acetate and hexane (EtOAc: hexane = 1:2) to obtain 0.2g (yield: 33%) of the title compound.

[0248] 1 H NMR(400MHz, CDCl3)δ9.16(d,1H),8.57(d,1H),7.88(d,1H),7.81-7.55(m,7H),7.52(m,2H),7.43(m,2H), 5.58(m,2H),5.17(m,1H),5.02(m,2H),4.08(d,1H),3.85(d,1H),3.03(dd,2H),2.43(m,1H),1.11(dd,6H)

[0249] Example 34: (S)-5-fluoro-3-((R)-5-isopropyl-3-(isoquinolin-1-yl)-4,5-dihydroisoquinolin-1-yl)- 5-Azole Amide)-4-oxopentanoic acid 2,2,2-trifluoroethyl ester

[0250]

[0251] Formula 2 compound (0.5g, 1.2mmol) is dissolved in dichloromethane (10mL), then trifluoroethanol (0.35mL, 4.8mmol, 4 equivalents), hydroxybenzotriazole (0.64g, 0.24mmol, 0.2 equivalents) and triethylamine (0.6mL, 0.24mmol, 0.2 equivalents), and EDCI (0.28g, 1.4mmol, 1.2 equivalents) is added thereto while the temperature is kept below 5°C. The reaction mixture is stirred at 25°C for about 6 hours, and 10% sodium bicarbonate aqueous solution (10mL) is added to terminate the reaction. After adding water (10mL) and stirring, the organic layer is separated and distilled under reduced pressure. The obtained mixture is subjected to column separation by using a 1:2 mixture of ethyl acetate and hexane (EtOAc: hexane = 1:2) to obtain 0.1g (yield: 17%) of the title compound.

[0252] 1 H NMR (400MHz, CDCl3) δ9.14(d,1H),8.56(d,1H),7.88(d,1H),7.76-7.65(m,4H),4.88(m,1H), 4.70(m,2H),4.12(m,2H),4.06(d,1H),3.82(d,1H),3.06(dd,2H),2.40(m,1H),1.11(dd,6H)

[0253] Example 35: (S)-5-fluoro-3-((R)-5-isopropyl-3-(isoquinolin-1-yl)-4,5-dihydroisoquinolin-1-yl)- 5-Azole Amide)-4-oxopentanoic acid 2-methoxyethyl ester

[0254]

[0255] Formula 2 compound (0.5g, 1.2mmol) is dissolved in dichloromethane (10mL), then methoxyethanol (0.37mL, 4.8mmol, 4 equivalents), hydroxybenzotriazole (0.64g, 0.24mmol, 0.2 equivalents) and triethylamine (0.6mL, 0.24mmol, 0.2 equivalents), and EDCI (0.28g, 1.4mmol, 1.2 equivalents) is added thereto while the temperature is kept below 5°C. The reaction mixture is stirred at 25°C for about 6 hours, and 10% sodium bicarbonate aqueous solution (10mL) is added to terminate the reaction. After adding water (10mL) and stirring, the organic layer is separated and distilled under reduced pressure. The obtained mixture is subjected to column separation by using a 1:2 mixture of ethyl acetate and hexane (EtOAc: hexane = 1:2) to obtain 0.14g (yield: 25%) of the title compound.

[0256] 1H NMR(400MHz, CDCl3)δ9.15(d,1H),8.56(d,1H),7.88(d,1H),7.75-7.65(m,4H),5.21(m,2H),4.92(m,1H), 4.17(m,2H),4.05(d,1H),3.82(d,1H),3.47(m,2H),3.30(S,3H),3.05(dd,2H),2.42(m,1H),1.11(dd,6H)

[0257] Example 36: (S)-5-fluoro-3-((R)-5-isopropyl-3-(isoquinolin-1-yl)-4,5-dihydroisoquinolin-1-yl)- 5-Azole Amide)-4-oxopentanoic acid 2-fluoroethyl ester

[0258]

[0259] Formula 2 compound (100mg, 0.24mmol) and EDCI (69mg, 0.36mmol), DMAP (3mg, 0.02mmol) and 2-fluoroethanol (0.14mL, 2.41mmol) were reacted in dichloromethane (4mL) for 2 hours at room temperature. After adding water, reaction mixture was extracted with EtOAc. Organic layer was dried over sodium sulfate, concentrated and purified using medium pressure liquid chromatography (MPLC), to obtain title compound (76mg, 68%).

[0260] 1 H NMR(CDCl3)δ9.12(t,1H),8.55(d,1H),7.86~7.66(m,5H),5.28~4.88(m,3H),4.64~4.3 0(m,4H),4.01(dd,1H),3.79(dd,1H),3.16~2.94(m,2H),2.41(p,1H),1.08~1.07(m,6H)

[0261] Example 37: (S)-5-fluoro-3-((R)-5-isopropyl-3-(isoquinolin-1-yl)-4,5-dihydroisoquinolin-1-yl)- 5-Azole Amide)-4-oxopentyl ester

[0262]

[0263] Formula 2 compound (2.0 g, 4.81 mmol) was reacted with EDCI (1.4 g, 7.22 mmol), DMAP (118 mg, 0.96 mmol) and 2,2-dimethylpropane-1-ol (5.19 mL, 48.1 mmol) in dichloromethane (80 mL) for 18 hours at room temperature. After adding water, the reaction mixture was extracted with EtOAc. The organic layer was dried over sodium sulfate, concentrated and purified using MPLC to obtain the title compound (0.94 g, 40%).

[0264] 1 H NMR(CDCl3)δ9.13(t,1H),8.54(d,1H),7.86~7.66(m,5H),5.28~4.91(m,3H),4.01(dd,1H),3.82~3. 79(m,2H),3.67(q,1H),3.11~2.90(m,2H),2.37(p,1H),1.10~1.04(m,6H),0.91(s,5H),0.79(s,4H)

[0265] Example 38: (S)-5-fluoro-3-((R)-5-isopropyl-3-(isoquinolin-1-yl)-4,5-dihydroisoquinolin-1-yl)- 5-Azole Amino)-4-oxopentanoic acid thiophen-2-ylmethyl ester

[0266]

[0267] Formula 2 compound (500mg, 1.20mmol) and EDCI (277mg, 1.44mmol), DMAP (37mg, 0.24mmol) and thiophene-2-ylmethanol (550mg, 4.81mmol) were reacted in dichloromethane (4mL) for 12 hours at room temperature. After adding water, the reaction mixture was extracted with EtOAc. The organic layer was dried over sodium sulfate, concentrated and purified using MPLC to obtain the title compound (306mg, 50%).

[0268] 1 H NMR(CDCl3)δ9.15(t,1H),8.56(d,1H),7.88~7.67(m,4H),7.38~6.99(m,4H),5.32~4.90(m ,5H),4.01(dd,1H),3.80(dd,1H),3.13~2.88(m,2H),2.42~2.32(m,1H),1.10~1.03(m,6H)

[0269] Example 39: (S)-5-fluoro-3-((R)-5-isopropyl-3-(isoquinolin-1-yl)-4,5-dihydroisoquinolin-1-yl)- 5-Azole Amide)-4-oxopentanoic acid thiophen-3-ylmethyl ester

[0270]

[0271] Formula 2 compound (500mg, 1.20mmol) and EDCI (277mg, 1.44mmol), DMAP (37mg, 0.24mmol) and thiophene-3-ylmethanol (550mg, 4.81mmol) were reacted in dichloromethane (4mL) for 12 hours at room temperature. After adding water, the reaction mixture was extracted with EtOAc. The organic layer was dried over sodium sulfate, concentrated and purified using MPLC to obtain the title compound (243mg, 40%).

[0272] 1 H NMR(CDCl3)δ9.14(t,1H),8.54(d,1H),7.87~7.64(m,4H),7.37~7.17(m,4H),5.15~4.90(m ,4H),4.01(dd,1H),3.78(dd,1H),3.14~2.87(m,2H),2.40~2.33(m,1H),1.09~1.02(m,6H)

[0273] Example 40: (S)-5-fluoro-3-((R)-5-isopropyl-3-(isoquinolin-1-yl)-4,5-dihydroisoquinolin-1-yl)- 5-Azole Amide)-4-oxopentanoic acid furan-3-yl methyl ester

[0274]

[0275] Formula 2 compound (500mg, 1.20mmol) and EDCI (277mg, 1.44mmol), DMAP (37mg, 0.24mmol) and furan-3-ylmethanol (471mg, 4.81mmol) were reacted in dichloromethane (4mL) for 12 hours at room temperature. After adding water, the reaction mixture was extracted with EtOAc. The organic layer was dried over sodium sulfate, concentrated and purified using MPLC to obtain the title compound (269mg, 44%).

[0276] 1 H-NMR(CDCl3)δ9.13(t,1H),8.54(d,1H),7.87~7.67(m,4H),7.40~7.35(m,3H),6.54(m,1H),5.32 ~4.56(m,5H),4.01(dd,1H),3.77(dd,1H),3.11~2.88(m,2H),2.40~2.33(m,1H),1.10~1.02(m,6H)

[0277] Experimental Example 1: Hydrolysis test of the prodrug compound of Formula 1 in plasma

[0278] Collect whole blood of 7-week-old male SD rats and centrifuge to obtain fresh plasma. Select the compound of Example 2 as the test prodrug, and use its 5mg / mL DMSO storage solution as the working solution. The solution is diluted to a concentration of 0.5mg / mL in acetonitrile by 1 / 10, and then the fresh plasma obtained above is added in a ratio of 1 / 100 so that the final concentration in plasma is 1μg / mL. It is set to the starting concentration for measuring drug stability in plasma. Thereafter, 50μl plasma is collected at 10 seconds, 5 minutes, 10 minutes, 20 minutes and 30 minutes, respectively, deproteinized and centrifuged with acetonitrile containing internal standard (IS), and the supernatant is injected into LC-MS / MS for analysis. The peak area of ​​the prodrug obtained by correcting the peak area with the peak area of ​​IS is used to obtain the peak response of each sample collection time, and converted into a residual rate (remaining %) compared with the initial value.

[0279] Furthermore, by correcting the obtained peak area of ​​the compound of Formula 2 with the peak area of ​​IS, the peak response at the time of each sample collection was obtained, and the pattern of generation and disappearance over time was confirmed.

[0280] The analysis results are shown in Figure 1 In. According to Figure 1 , when the prodrug of Formula 1 was mixed with fresh plasma of rats, most of the prodrug was lost within about 5 minutes, and the result was analyzed, this was because the prodrug of Formula 1 was hydrolyzed and converted into the compound of Formula 2 due to esterase present in plasma.

[0281] Experimental Example 2: Pharmacokinetics test in mice

[0282] For the subcutaneous injection (SC) PK test of the prodrug compound of Formula 1 (compounds of Examples 2, 16 and 32), C57BL6 mice of about 7 weeks of age were prepared, and group separation was performed by allocating 3 animals for each administered compound. Because it was a subcutaneous injection situation, fasting was not performed. On the day of administration, a drug solution with a concentration of 5 mg / mL was prepared by using 0.5% methylcellulose (MC) as a vehicle, and the drug solution was subcutaneously injected with 10 mL per kg of body weight per individual so that the final dose was 50 mg / kg. At 1, 2, 4, 6, 8, 24 and 48 hours after administration, blood was collected through the orbital vein, and the blood of each group of three animals was collected in a heparin tube. The obtained blood sample was centrifuged at 15,000 rpm for 2 minutes to separate plasma, and 50 μl was taken and stored frozen at -20 ° C. On the day of analysis, the sample was thawed at room temperature and deproteinized with 200 μl of acetonitrile (a total of 4 times the storage volume). At this time, acetonitrile contains internal standard and 5% formic acid (FA). To make a calibration curve, acetonitrile solutions (containing IS and 5% FA) with known concentrations of 0.1, 0.5, 5, 50 and 500 ng / mL were prepared respectively, and the blank serum was deproteinized with 4 volumes of acetonitrile as described above to prepare a final calibration curve of 0.4 to 2000 ng / mL. After 0.5 μl of the supernatant obtained after deproteinization was injected into LC-MS / MS, the peak area of ​​the compound of formula 2 was corrected relative to the peak area of ​​IS to obtain the peak response at each sample collection point, and the concentration was converted by the calibration curve. For the blood concentration values ​​of each dosing group that changed over time, the pharmacokinetic parameters (C max , T max , AUC last ,t 1 / 2 The pharmacokinetic characteristics of each compound were compared by comparing the changes in exposure and half-life among the dosing groups.

[0283] In the case of subcutaneous injection of the prodrug of the compound of formula 2 as the parent drug, the peak blood concentration value of the compound of formula 2 was reduced. In addition, the elimination half-life (t 1 / 2 ) tends to increase with the substance. The characteristics observed in the case of administering the prodrug are advantageous in terms of the manifestation of side effects caused by high blood concentrations, and are expected to have advantages in terms of maintaining the effective concentration required for efficacy expression and achieving the desired efficacy compared to direct administration of the parent drug.

[0284] The pharmacokinetic parameters of the parent drug, which is the metabolite measured in mouse plasma after subcutaneous administration of the compounds of Examples 2, 16 and 32 (prodrug compounds of Formula 1), are shown in Table 1 below.

[0285] [Table 1]

[0286]

[0287] Figure 2 Figure 2 shows the mean plasma concentration curve of the compound of Formula 2 obtained after subcutaneous injection of the prodrugs of the compounds of Examples 2, 16 and 32 into C57B0L / 6 mice over time.

[0288] Example 41: Preparation of sustained-release microspheres for intra-articular administration using caspase inhibitor prodrugs

[0289] According to the composition shown in Table 2 and Table 3 below, 16 sustained-release test microspheres for intra-articular administration encapsulated with caspase inhibitor prodrugs were prepared. First, the prodrug and PLGA (L / G ratio = 50:50, molecular weight 38,000 to 54,000) were weighed in a weight ratio of 1:5, the organic solvent dichloromethane was added, and stirred to prepare the dispersed phase. For the continuous phase, 150mL2% polyvinyl alcohol (molecular weight 31000 to 50000, degree of hydrolysis 8%7 to 89%) was used, and an emulsion was prepared by membrane emulsification. The prepared emulsion was stirred at room temperature overnight to remove the solvent (solvent evaporation), washed with sterile purified water, and then lyophilized to obtain microspheres.

[0290] [Table 2]

[0291]

[0292] [Table 3]

[0293]

[0294] Example 42: Preparation of sustained-release microspheres for intra-articular administration using caspase inhibitors

[0295] According to the composition shown in Table 4 below, two kinds of sustained-release comparison microspheres for intra-articular administration encapsulated with caspase inhibitors were prepared. The dispersed phase was prepared by weighing the caspase inhibitor and PLGA in a weight ratio of 1:5, adding the organic solvent dichloromethane and stirring. At this time, the L / G ratio of the PLGA used was 50:50 (molecular weight 38,000 to 54,000) and 75:25 (molecular weight 76,000 to 115,000). For the continuous phase, 150mL of 2% polyvinyl alcohol (molecular weight 31,000 to 50,000, degree of hydrolysis 87 to 89%) was used, and an emulsion was prepared by membrane emulsification. The prepared emulsion was stirred overnight at room temperature to remove the solvent, washed repeatedly with sterile purified water, and then freeze-dried to prepare microspheres.

[0296] [Table 4]

[0297] Compare the number of microspheres 1 2 Amount of Nivocasan (g) 0.4 0.4 Amount of PLGA (g) 2.0 2.0 L / G ratio of PLGA 50:50 75:25 Molecular weight of PLGA (kDa) 38-54 76-115 Amount of solvent (g) 20.0 20.0 PVA concentration (%, weight / volume) 2 2 Amount of PVA solution (mL) 150 150

[0298] Experimental Example 3: Analysis of microsphere properties and drug encapsulation efficiency

[0299] The properties of the microspheres prepared in Example 41 (test microspheres 1 to 16) and Example 42 (comparative microspheres 1 and 2) were characterized by the precipitation of the drug during the manufacturing process, the morphology of the lyophilized microspheres, and the floating in the aqueous phase when redispersed. During the manufacturing process, whether the drug was precipitated was confirmed by optical microscopy, and the morphology of the lyophilized microspheres was observed by scanning electron microscopy. Whether the microspheres floated in the aqueous phase was confirmed by redispersing the lyophilized microspheres in water. For the amount of drug encapsulated in the microspheres, 30 mg of microspheres were dissolved in 50 mL of acetonitrile, and the supernatant obtained by ultracentrifugation was analyzed by HPLC (high performance liquid chromatography). In addition, the occurrence of a large amount of caspase inhibitor precipitation during the preparation of comparative microspheres 1 and comparative microspheres 2 was confirmed by using an optical microscope.

[0300] The observation or measurement results of whether there is drug precipitation in the test microspheres 1, 2, 12 to 14 and 16, the morphology of the microspheres, whether the microspheres float, and the drug encapsulation rate (%, weight / weight) are shown in the following Table 5. In addition, the morphology of these test microspheres was observed by scanning electron microscopy and is shown in Table 5. Figure 3 The diameter of the microspheres is about 50 μm, and all the microspheres have a good surface. Figure 3 In the experimental microspheres 2, the needle-shaped substances still remained in the form of drug crystals and could be removed by washing.

[0301] [Table 5]

[0302]

[0303] The observation or measurement results of drug precipitation in comparative microspheres 1 and 2, microsphere morphology, whether the microspheres float, and drug encapsulation efficiency (%, weight / weight) are shown in Table 6 below. The comparative microspheres containing the caspase inhibitor had a good morphology and no microsphere floating phenomenon, but a large amount of drug was precipitated during the manufacturing process. The drug encapsulation efficiency was observed to be about 8% in both comparative microspheres 1 and 2, and was confirmed to be about half of the drug encapsulation efficiency of the test microspheres measured in Table 5 above. In addition, according to Figure 4 It was confirmed that a large amount of caspase inhibitor crystals were precipitated in Comparative Microspheres 1 and 2 during the solidification step.

[0304] [Table 6]

[0305] Compare the number of microspheres 1 2 Drug precipitation Large Large Microsphere morphology good good Microsphere floating none none Drug encapsulation efficiency (%, weight / weight) 8.2 7.8

[0306] Therefore, it was confirmed that the drug encapsulation efficiency was not high when the microspheres were prepared using the caspase inhibitor, but was greatly improved when the microspheres were prepared using the caspase inhibitor prodrug.

[0307] Example 43: Preparation of sustained-release microparticles for intra-articular administration using the caspase inhibitor prodrug of Example 16 ball

[0308] According to the composition shown in Table 7 below, test microspheres 17 to 24 encapsulating the caspase inhibitor prodrug of Example 16 were prepared for drug sustained-release microspheres for intra-articular administration. As shown in Table 7 below, the weight ratio of the prodrug compound and PLGA was 10, 13, 16 or 20% (weight / weight), and an organic solvent, dichloromethane, was added thereto and stirred to prepare a dispersed phase. The L / G ratio of the PLGA used had two different forms: 50:50 (molecular weight 38,000 to 54,000) and 75:25 (molecular weight 76,000 to 115,000). For the continuous phase, 150 mL of 2% polyvinyl alcohol (molecular weight 31,000 to 50,000, degree of hydrolysis 87 to 89%) was used, and an emulsion was prepared by membrane emulsification. The prepared emulsion was stirred overnight at room temperature to remove the solvent, washed repeatedly with sterile purified water, and then freeze-dried to prepare microspheres.

[0309] [Table 7]

[0310]

[0311]

[0312] Experimental Example 4: Analysis of physical stability and drug encapsulation efficiency of microspheres

[0313] The physical stability of test microspheres 17 to 24 was judged by whether aggregation occurred after shaking in a buffer solution PBS (phosphate buffered saline, 37°C) for one day. Table 8 shows the analysis results of the physical stability and drug encapsulation efficiency of test microspheres 17 to 24. In test microspheres 17 to 19, aggregation of the microspheres in PBS was not observed within one day, but aggregation was found in test microsphere 20. In addition, in test microspheres 21 to 23, no microsphere aggregation phenomenon was found within one day, but aggregation phenomenon was found in test microsphere 24. Because it has been confirmed that most of the loading amount of the prodrug of Example 16 was encapsulated during the preparation of the microspheres, the encapsulation efficiency was not measured separately in this experiment.

[0314] [Table 8]

[0315]

[0316] The state of the test microspheres 17 to 24 dispersed in PBS for one day is shown in FIG. Figure 5 In. According to Figure 5 a. It can be confirmed that the microspheres encapsulating 10, 13 and 16% (theoretical encapsulation rate) of the prodrug of Example 16 were well dispersed in PBS and no particles were observed, but the microspheres encapsulating 20% ​​(theoretical encapsulation rate) of the prodrug of Example 16 were aggregated and agglomerated. Figure 5b. It has been confirmed that the microspheres encapsulating 10, 13 and 16% (theoretical encapsulation rate) of the prodrug of Example 16, respectively, are well dispersed in PBS and no particles are seen, but the microspheres encapsulating 20% ​​(theoretical encapsulation rate) of the prodrug of Example 16 aggregate and agglomerate.

[0317] Therefore, it was confirmed that when microspheres containing 20% ​​or more of the prodrug of Example 16 were prepared, the microspheres aggregated within one day in PBS.

[0318] Experimental Example 5: In vitro dissolution test of microspheres containing the prodrug of Example 2

[0319] An in vitro dissolution test was performed on the microspheres prepared by encapsulating the prodrug of Example 2 according to Test Microspheres 2. More specifically, the microspheres were shaken in PBS buffer (37° C.), the eluate was collected and filtered at a specific time, and the drug release amount was analyzed by HPLC. Because the prodrug is converted into the parent drug, i.e., the caspase inhibitor, by hydrolysis in an aqueous solution, the drug release amount was determined by the amount of the caspase inhibitor measured by HPLC.

[0320] Furthermore, the same experiment was repeated in PBS containing 1% hyaluronic acid and 2.5% bovine serum albumin simulating synovial fluid, and it was examined whether the dissolution pattern was altered in simulated synovial fluid (sSF).

[0321] The in vitro dissolution curve of PLGA microspheres encapsulating the prodrug of Example 2 is shown in Figure 6 The experimental results confirmed that the dissolution of the caspase inhibitor lasted for about 12 weeks, and no difference in the dissolution pattern was found between the two dissolution test solutions, PBS and sSF. In summary, since no difference was found in the dissolution pattern between PBS and sSF, it was confirmed that sSF was not required in subsequent dissolution tests.

[0322] Experimental Example 6: In vitro dissolution test of microspheres containing the prodrug of Example 16

[0323] The microspheres prepared by encapsulating the prodrug of Example 16 according to the test microspheres 18 and 22 were subjected to an in vitro dissolution test. More specifically, the microspheres were shaken in PBS buffer (37° C.), the eluate was collected and filtered at a specific time, and the drug release was analyzed by HPLC. Because the prodrug is converted into the parent drug, i.e., a caspase inhibitor, by hydrolysis in an aqueous solution, the drug release is determined by the amount of the caspase inhibitor measured by HPLC. In addition, it was confirmed by scanning electron microscopy that the properties of the microspheres change with the hydrolysis process. The molecular weight of PLGA was measured using GPC (gel permeation chromatography), which changes as the hydrolysis proceeds.

[0324] The in vitro dissolution curves of PLGA microspheres according to test microspheres 18 and 22 are shown in Figure 7The dissolution of the test microsphere 18 having an L / G ratio of PLGA of 50:50 lasted for about 6 weeks, and the dissolution of the test microsphere 22 having an L / G ratio of 75:25 lasted for about 14 weeks. Both types of microspheres showed a pattern in which dissolution continued at a slow slope after an initial peak, and the dissolution amount increased rapidly in the middle.

[0325] The photographs of the microsphere shapes observed during the dissolution test are shown in Figure 8 Around week 4, test microsphere 18 (5050 PLGA) had more pores, while at week 8, the microspheres swelled and grew larger with larger pores. In test microsphere 22 (7525 PLGA), pore formation was gradually observed at week 8, while at week 12, PLGA swelled and the sphere shape collapsed.

[0326] The change in molecular weight of the test microspheres 18 (5050 PLGA) during the dissolution test is shown in FIG. Fig. 9 With the start of the dissolution test, the molecular weight decreased rapidly, and in the 4th week the molecular weight decreased to about 1 / 5 of the original molecular weight, and thereafter maintained a similar level.

[0327] As a result of the dissolution test, when PLGA is decomposed by hydrolysis from the start of dissolution, the drug on the surface of the microsphere is rapidly eluted, and analysis shows that when PLGA is further decomposed, pores are formed and the drug is actively released and diffused at a specific time point. In addition, it has been confirmed that a more delayed release pattern is observed when PLGA (7525PLGA) with an L / G ratio of 75:25 is used compared to PLGA (5050PLGA) with an L / G ratio of 50:50, and the morphology is changed and the molecular weight is reduced due to hydrolysis.

[0328] Experimental Example 7: Pharmacokinetic (PK) test of microspheres containing the prodrug of Example 2

[0329] A PK study was conducted in dogs using microspheres prepared by encapsulating the prodrug of Example 2 according to Test Microspheres 2. Microspheres at a concentration of 300 mg / ml were administered to the joint cavity, and synovial fluid was collected at specific time points to measure the concentration of the caspase inhibitor, and the results are shown in Fig.10 middle.

[0330] As a result of measuring the caspase inhibitor in the joint cavity, a significant initial increase occurred after 24 hours, after which the concentration gradually decreased. The duration was expected to be up to 2 weeks, but it was confirmed that the test microsphere 2 (5050 PLGA) encapsulating the prodrug of Example 2 continuously released the caspase inhibitor for about 4 weeks, because the concentration increased slightly when measured after 4 weeks. In short, it can be confirmed that in the test microsphere 2 encapsulating the prodrug of Example 2, the caspase inhibitor was continuously released for about 4 weeks.

Claims

1. A compound of the following formula 1, or a pharmaceutically acceptable salt thereof: [Formula 1] in R stands for C 1-7 Alkyl, halogenated C 1-7 Alkyl, C 3-6 Cycloalkyl, C 3-6 Cycloalkyl-C 1-3 Alkyl, C6-C 10 Aryl or C6-C 10 Aryl-C 1-3 Alkyl; and Provided that R is not tert-butyl.

2. The compound according to claim 1 or a pharmaceutically acceptable salt thereof, wherein the compound is selected from: (S)-5-Fluoro-3-((R)-5-isopropyl-3-(isoquinolin-1-yl)-4,5-dihydroisoquinolin-1-yl) Methyl 4-oxopentanoate; (S)-5-Fluoro-3-((R)-5-isopropyl-3-(isoquinolin-1-yl)-4,5-dihydroisoquinolin-1-yl) Ethyl 4-oxopentanoate; (S)-5-Fluoro-3-((R)-5-isopropyl-3-(isoquinolin-1-yl)-4,5-dihydroisoquinolin-1-yl) Propyl 4-oxopentanoate; (S)-5-Fluoro-3-((R)-5-isopropyl-3-(isoquinolin-1-yl)-4,5-dihydroisoquinolin-1-yl) 4-oxopentanoic acid butyl ester; (S)-5-Fluoro-3-((R)-5-isopropyl-3-(isoquinolin-1-yl)-4,5-dihydroisoquinolin-1-yl) Isobutyl 4-oxopentanoate; (S)-5-Fluoro-3-((R)-5-isopropyl-3-(isoquinolin-1-yl)-4,5-dihydroisoquinolin-1-yl) Isoamyl 4-oxopentanoate; (S)-5-Fluoro-3-((R)-5-isopropyl-3-(isoquinolin-1-yl)-4,5-dihydroisoquinolin-1-yl) 4-oxopentanoic acid pentyl ester; (S)-5-Fluoro-3-((R)-5-isopropyl-3-(isoquinolin-1-yl)-4,5-dihydroisoquinolin-1-yl) 4-oxopentanoic acid hexyl ester; (S)-5-Fluoro-3-((R)-5-isopropyl-3-(isoquinolin-1-yl)-4,5-dihydroisoquinolin-1-yl) oxadiazole-5-carboxamido)-4-oxopentanoic acid heptyl ester; (S)-5-Fluoro-3-((R)-5-isopropyl-3-(isoquinolin-1-yl)-4,5-dihydroisoquinolin-1-yl) 4-oxopentanoic acid octyl ester; (S)-5-Fluoro-3-((R)-5-isopropyl-3-(isoquinolin-1-yl)-4,5-dihydroisoquinolin-1-yl) 4-oxopentanoic acid decyl ester; (S)-5-Fluoro-3-((R)-5-isopropyl-3-(isoquinolin-1-yl)-4,5-dihydroisoquinolin-1-yl) oxadiazole-5-carboxamido)-4-oxopentanoic acid dodecyl ester; (S)-5-Fluoro-3-((R)-5-isopropyl-3-(isoquinolin-1-yl)-4,5-dihydroisoquinolin-1-yl) Pentadecyl 4-oxopentanoate; (S)-5-Fluoro-3-((R)-5-isopropyl-3-(isoquinolin-1-yl)-4,5-dihydroisoquinolin-1-yl) Octadecyl 4-oxopentanoate; (S)-5-Fluoro-3-((R)-5-isopropyl-3-(isoquinolin-1-yl)-4,5-dihydroisoquinolin-1-yl) (9E,12E)-octadeca-9,12-dien-1-yl)-4-oxopentanoate; (S)-5-Fluoro-3-((R)-5-isopropyl-3-(isoquinolin-1-yl)-4,5-dihydroisoquinolin-1-yl) 4-oxopentanoic acid cyclopropylmethyl ester; (S)-5-Fluoro-3-((R)-5-isopropyl-3-(isoquinolin-1-yl)-4,5-dihydroisoquinolin-1-yl) 4-oxopentanoic acid cyclobutylmethyl ester; (S)-5-Fluoro-3-((R)-5-isopropyl-3-(isoquinolin-1-yl)-4,5-dihydroisoquinolin-1-yl) 4-oxopentanoic acid cyclopentylmethyl ester; (S)-5-Fluoro-3-((R)-5-isopropyl-3-(isoquinolin-1-yl)-4,5-dihydroisoquinolin-1-yl) Allyl 4-oxopentanoate; (S)-5-Fluoro-3-((R)-5-isopropyl-3-(isoquinolin-1-yl)-4,5-dihydroisoquinolin-1-yl) Isopropyl 4-oxopentanoate; (S)-5-Fluoro-3-((R)-5-isopropyl-3-(isoquinolin-1-yl)-4,5-dihydroisoquinolin-1-yl) 4-oxopentanoic acid pent-3-yl ester; (S)-5-Fluoro-3-((R)-5-isopropyl-3-(isoquinolin-1-yl)-4,5-dihydroisoquinolin-1-yl) 4-oxopentanoic acid sec-butyl ester; (S)-5-Fluoro-3-((R)-5-isopropyl-3-(isoquinolin-1-yl)-4,5-dihydroisoquinolin-1-yl) 4-oxopentanoic acid pent-2-yl ester; (S)-5-Fluoro-3-((R)-5-isopropyl-3-(isoquinolin-1-yl)-4,5-dihydroisoquinolin-1-yl) oxadiazole-5-carboxamido)-4-oxopentanoic acid hept-2-yl ester; (S)-5-Fluoro-3-((R)-5-isopropyl-3-(isoquinolin-1-yl)-4,5-dihydroisoquinolin-1-yl) 4-oxopentanoic acid cyclopentyl ester; (S)-5-Fluoro-3-((R)-5-isopropyl-3-(isoquinolin-1-yl)-4,5-dihydroisoquinolin-1-yl) 4-oxopentanoic acid cyclohexyl ester; (S)-5-Fluoro-3-((R)-5-isopropyl-3-(isoquinolin-1-yl)-4,5-dihydroisoquinolin-1-yl) Benzyl 4-oxopentanoate; (S)-5-Fluoro-3-((R)-5-isopropyl-3-(isoquinolin-1-yl)-4,5-dihydroisoquinolin-1-yl) (5-methyl-2-oxo-1,3-dioxol-4-yl)methyl 4-oxopentanoate; (S)-5-Fluoro-3-((R)-5-isopropyl-3-(isoquinolin-1-yl)-4,5-dihydroisoquinolin-1-yl) 2-methoxyphenyl oxadiazole-5-carboxamido)-4-oxopentanoate; (S)-5-Fluoro-3-((R)-5-isopropyl-3-(isoquinolin-1-yl)-4,5-dihydroisoquinolin-1-yl) 2,3-dihydro-1H-inden-5-yl 4-oxopentanoate; (S)-5-Fluoro-3-((R)-5-isopropyl-3-(isoquinolin-1-yl)-4,5-dihydroisoquinolin-1-yl) oxadiazole-5-carboxamido)-naphthyl-1-4-oxopentanoate; (S)-5-Fluoro-3-((R)-5-isopropyl-3-(isoquinolin-1-yl)-4,5-dihydroisoquinolin-1-yl) 4-oxopentanoic acid phenyl ester; (S)-5-Fluoro-3-((R)-5-isopropyl-3-(isoquinolin-1-yl)-4,5-dihydroisoquinolin-1-yl) 4-oxopentanoic acid naphthalen-1-ylmethyl ester; (S)-5-Fluoro-3-((R)-5-isopropyl-3-(isoquinolin-1-yl)-4,5-dihydroisoquinolin-1-yl) 2,2,2-trifluoroethyl oxadiazole-5-carboxamido)-4-oxopentanoate; (S)-5-Fluoro-3-((R)-5-isopropyl-3-(isoquinolin-1-yl)-4,5-dihydroisoquinolin-1-yl) 2-methoxyethyl oxadiazole-5-carboxamido)-4-oxopentanoate; (S)-5-Fluoro-3-((R)-5-isopropyl-3-(isoquinolin-1-yl)-4,5-dihydroisoquinolin-1-yl) 2-Fluoroethyl 4-oxopentanoate; (S)-5-Fluoro-3-((R)-5-isopropyl-3-(isoquinolin-1-yl)-4,5-dihydroisoquinolin-1-yl) oxadiazole-5-carboxamido)-4-oxopentyl ester; (S)-5-Fluoro-3-((R)-5-isopropyl-3-(isoquinolin-1-yl)-4,5-dihydroisoquinolin-1-yl) 4-oxopentanoic acid thiophen-2-ylmethyl ester; (S)-5-Fluoro-3-((R)-5-isopropyl-3-(isoquinolin-1-yl)-4,5-dihydroisoquinolin-1-yl) 4-oxopentanoic acid thiophen-3-ylmethyl ester; and (S)-5-Fluoro-3-((R)-5-isopropyl-3-(isoquinolin-1-yl)-4,5-dihydroisoquinolin-1-yl) oxadiazole-5-carboxamido)-4-oxopentanoic acid furan-3-ylmethyl ester.

3. The compound according to claim 1 or a pharmaceutically acceptable salt thereof, wherein the compound is a prodrug of a caspase inhibitor.

4. A pharmaceutical composition for preventing or treating inflammation or apoptosis, comprising as an active ingredient a compound of formula 1 as defined in any one of claims 1 to 3 or a pharmaceutically acceptable salt thereof, and a pharmaceutically acceptable carrier.

5. The pharmaceutical composition according to claim 4, which is formulated into an oral dosage form, an injection form or a patch form.

6. The pharmaceutical composition according to claim 4, comprising microspheres, wherein the microspheres comprise the compound of formula 1 or a pharmaceutically acceptable salt thereof and a biocompatible polymer.

7. The pharmaceutical composition according to claim 6, wherein the biocompatible polymer is a poly(lactide-co-glycolide) having a molar ratio of lactide to glycolide of 10:90 to 90:

10.

8. The pharmaceutical composition according to claim 6, wherein the weight ratio of the compound of Formula 1 or a pharmaceutically acceptable salt thereof to the biocompatible polymer in the microspheres is 1:100 to 70:

100.

9. A pharmaceutical composition for preventing or treating a disease selected from apoptosis-related diseases and inflammatory diseases, the pharmaceutical composition comprising as an active ingredient a compound of formula 1 or a pharmaceutically acceptable salt thereof according to any one of claims 1 to 3, and a pharmaceutically acceptable carrier.

10. The pharmaceutical composition according to claim 9, wherein the disease is selected from osteoarthritis, rheumatoid arthritis and degenerative arthritis.

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