Compound, pharmaceutical composition containing the compound, and these pharmaceutical uses for the treatment or prevention of vascular diseases

Abstract

The present invention provides compounds that exhibit effects similar to those of 2-Cys-peroxiredoxin (2-Cys-Prx) in the body, have excellent pharmacological effects, and reduced side effects such as cytotoxicity, and pharmaceutical uses thereof. The compounds of the present invention and pharmaceutically acceptable salts thereof are useful for the treatment or prevention of vascular diseases, particularly ischemic coronary artery disease, arteriosclerosis, vascular restenosis, or pulmonary hypertension. The compounds of the present invention and pharmaceutically acceptable salts thereof are particularly useful for the treatment or prevention of pulmonary hypertension. The present invention also provides a method for producing the compounds according to the present invention.

Description

Detailed Description of the Invention

[0001] 〔Technical Field〕 The present invention relates to a compound exhibiting 2-Cys-peroxiredoxin (2-Cys-Prx) peroxidase mimetic activity and a pharmaceutically acceptable salt thereof. The present invention also relates to a pharmaceutical composition for preventing or treating vascular diseases. That is, the present invention relates to the pharmaceutical use of the compound according to the present invention and a pharmaceutically acceptable salt thereof. Furthermore, the present invention relates to a method for producing a compound exhibiting a 2-Cys-Prx activity mimetic effect and a pharmaceutically acceptable salt thereof.

[0002] 〔Background Art〕 Arterial vascular diseases are conditions in which fatty substances (plaques) containing cholesterol, phospholipids, calcium, etc. accumulate on the vascular intima, causing inflammation, the arteries lose elasticity and become thinner, blood supply is inhibited or pressure increases, leading to blood vessel rupture or detachment. In particular, the resulting blockage or occlusion of arterial blood vessels reduces blood supply, causing a shortage of nutrients and oxygen, which is a major cause of vascular diseases. The vascular diseases generally include cardiovascular diseases such as arteriosclerosis, heart failure, hypertensive heart disease, arrhythmia, myocardial infarction, angina pectoris, and cerebrovascular diseases such as stroke and peripheral vascular diseases.

[0003] As methods for overcoming such vascular occlusion, there are an arterial transplantation surgical method, which is a surgical method, and percutaneous transluminal angioplasty, which is a method of expanding blood vessels using a balloon stent. Vascular restenosis is defined as a stenosis of the blood vessel diameter of 50% or more on follow-up angiography after angioplasty. Although the restenosis incidence has been decreasing with the improvement of the materials used for stents, vascular restenosis still occurs in about 30% of the patients who have undergone angioplasty (balloon dilation and stent insertion). Although the mechanism of such restenosis has not yet been clearly elucidated, growth factors and cytokines are locally secreted due to damage to vascular endothelial cells during the procedure, and it is known that these induce the proliferation and migration of vascular smooth muscle, resulting in a narrowing of the arterial lumen and leading to restenosis. Therefore, in recent years, hyperplasia of smooth muscle cells has been raised as a major clinical problem limiting the efficiency of angioplasty. Thus, improved stents that release drugs that suppress the proliferation of vascular smooth muscle cells have been developed and applied clinically. However, currently, the drugs used for such purposes prevent intimal hyperplasia through a cytotoxic mechanism that kills vascular smooth muscle cells, are highly toxic, and have limitations in killing not only smooth muscle cells but also endothelial cells due to their toxicity. Therefore, there is an urgent need to develop drugs with therapeutic potential that can selectively promote the recovery of the damaged endothelial cell layer while suppressing the growth of vascular smooth muscle cells.

[0004] On the one hand, the pulmonary artery is a blood vessel that transports blood from the right ventricle of the heart to the lungs to replenish oxygen. Pulmonary hypertension is defined as a case where the average pulmonary artery pressure at rest is 25 mmHg or more and the average pulmonary artery pressure during exercise is 30 mmHg. Pulmonary hypertension is divided into "idiopathic pulmonary hypertension" with no specific known cause and "associated pulmonary hypertension", a specific disease that occurs secondarily due to a disease with a specific cause. In the latter case, it may occur due to collagen vascular diseases (systemic sclerosis, systemic lupus erythematosus, etc.), portal hypertension, HIV infection, congenital heart disease, and drugs and toxins such as appetite suppressants and cocaine. These pulmonary hypertension associated with specific diseases do not differ significantly from idiopathic pulmonary hypertension in terms of natural course, histopathological findings, and response to treatment.

[0005] Currently, there is a method of using general vasodilators as drugs for the treatment of pulmonary hypertension. Most vasodilators are calcium channel blockers. However, calcium channel blockers often have no significant effect in pulmonary hypertension and have many side effects due to administration. Therefore, it is necessary to develop an effective therapeutic agent for pulmonary hypertension.

[0006] On the one hand, "PrxII (also referred to as peroxiredoxin II or Prx2)" is one of the 2-Cys-Prx peroxidases that reduce intracellular hydrogen peroxide (H2O2). PrxII removes hydrogen peroxide generated by PDGF (platelet-derived growth factor) from vascular smooth muscle cells, suppresses site-specific phosphorylation occurring at PDGFRβ and PLC (phospholipase C) γ1, and thereby acts to suppress signal transduction amplification. Through such a mechanism, the said PrxII has the activity of suppressing the proliferation and migration of smooth muscle cells and suppressing the thickening of the vascular intima in damaged blood vessels (M. H. Choi et al., Nature 2005 May 19;435(7040):347-53). However, in vascular endothelial cells, it has been reported that PrxII protects VEGFR2 from oxidative inactivation and activates VEGF-induced signal transduction (D. H. Kang et al., Mol Cell. 2011 Nov 18;44(4):545-58).

[0007] In International Publication WO2013-077709, it was confirmed that a natural compound with an episulfide dioxopiperazine structure can exhibit 2-Cys-Prx-like activity intracellularly. By mimicking the intracellular activity of 2-Cys-Prx, the natural compound suppresses PDGF-induced proliferation and migration in vascular smooth muscle cells and promotes VEGF-induced proliferation and migration in vascular endothelial cells. Furthermore, in an experimental animal model, it is implied that such a compound can be usefully used for the prevention or treatment of vascular diseases by inhibiting the thickening of the vascular intima due to excessive proliferation of vascular smooth muscle cells and promoting the recovery of the vascular endothelial layer.

[0008] Also, in International Publication WO2018-008984, it was confirmed that a drug that mimics the intracellular activity of 2-Cys-Prx peroxidase is useful for the treatment or prevention of pulmonary hypertension.

[0009] However, in the case of the compounds disclosed in International Publication WO2013-077709 and International Publication WO2018-008984, the above-mentioned toxicity problems have not been completely solved. Therefore, there is still a need for pharmaceutical ingredients that exhibit enhanced pharmacological effects while having improved cytotoxicity.

[0010] 〔Summary of the Invention〕 〔Problems to be Solved by the Invention〕 An object of the present invention is to provide compounds that exhibit effects similar to those of 2-Cys-peroxiredoxin (2-Cys-Prx) in the body, have excellent pharmacological effects, and have reduced side effects such as cytotoxicity, as well as pharmaceutical uses thereof.

[0011] Another object of the present invention is to provide a pharmaceutical composition for the treatment or prevention of vascular diseases containing a compound having excellent pharmacological effects and reduced side effects such as toxicity as an active ingredient. That is, an object of the present invention is to provide a method for treating or preventing vascular diseases, which includes the step of administering a therapeutically effective amount of a compound according to the present invention or a salt thereof to an individual in need of treatment or prevention of vascular diseases.

[0012] Yet another object of the present invention is to provide a method for producing a specific compound.

[0013] 〔Means for Solving the Problems〕 To achieve the above object, one aspect of the present invention provides a compound represented by the following Chemical Formula 1 or Chemical Formula 2, or a pharmaceutically acceptable salt thereof.

[0014]

Chemical Formula

[0015]

Chemical Formula

[0016] In Chemical Formula 1 and Chemical Formula 2, n is an integer from 1 to 3, R1 and R2 are, independently of each other, C 1-3 alkyl (preferably methyl, ethyl), C 1-3 alkoxy-C 1-3 alkyl, -(CH2) 1-3 -C(R’)(R”)OH, -(CH2) 1-3 -N(R’)(R”), -(CH2) 0-3 -alkenyl, -(CH2) 0-3 -alkynyl, -(CH2) 0-3 -C(R’)(R”)CO2H, -(CH2) 0-5 -heterocycloalkyl, -(CH2) 0-5 -cycloalkyl, -(CH2) 0-5 -aryl (preferably -CH2-phenyl), or -(CH2) 0-5 -heteroaryl (preferably -CH2-pyridyl, -CH2-quinolinyl, -CH2-pyrazolyl, -CH2-thiophen-2-yl, -CH2-benzo[d]thiazol-2-yl, -CH2-pyrimidyl, -CH2-1H-imidazol-4-yl), where the alkyl, heterocycloalkyl, cycloalkyl, aryl and heteroaryl are unsubstituted or substituted with one or more substituents selected from the group consisting of C 1-3 alkyl, -CF3, C 1-3 alkoxy, -OCF3, halogen (preferably F), CN, amino, -N(R’)(R”), -OH, -COOH, -COO-C 1-3 alkyl, and =O, where R’ and R” are, independently of each other, hydrogen or C 1-3 alkyl; R3 is C 1-3 alkyl (preferably methyl), -(CH2) 0-3 -aryl, or -(CH2) 0-3 -heteroaryl, where the aryl or heteroaryl is unsubstituted or substituted with one or more substituents selected from the group consisting of C 1-3 alkyl, -CF3, C 1-3 alkoxy, -OCF3, halogen, -CN, amino, -OH, and -COOH; or R2 and R3 are connected to each other to form any one of the following structures fused with the piperazinedione present in Chemical Formula 1:

[0017]

Chem.

[0018] Here, X is S, SO2, CH2, O or NR6, and R6 is hydrogen or C 1-3 alkyl, R4 is hydrogen or C 1-3 alkyl, R5 is hydrogen, C 1-3 alkyl, -(CH2)1-2-aryl, or -(CH2) 1-2 -heteroaryl.

[0019] In one aspect of the present invention, in the above Chemical Formula 1 or Chemical Formula 2, n is an integer from 1 to 3, R1 and R2 are independently of each other C 1-3 alkyl, -(CH2) 1-2 -heterocycloalkyl, -(CH2) 1-2 -aryl, or -(CH2) 1-2 -heteroaryl, where the alkyl, heterocycloalkyl, aryl, and heteroaryl are unsubstituted or substituted with one or more substituents selected from the group consisting of C 1-3 alkyl, -CF3, C 1-3 alkoxy, CN, halogen, -OH, -COOH, and -COO-C 1-3 alkyl; R3 is C 1-3 alkyl, -CH2-aryl, or -CH2-heteroaryl, where the aryl or heteroaryl is unsubstituted or substituted with one or more substituents selected from the group consisting of methyl, methoxy, halogen, -CN, amino, -OH, and -COOH; or R2 and R3 are connected to each other to form any one of the following structures fused with the piperazinedione present in Chemical Formula 1:

[0020] [Chemical formula]

[0021] Here, X is S, SO2, CH2, O or NR6, and R6 is hydrogen or C 1-3 alkyl, R4 is hydrogen or C 1-3 alkyl, R5 is hydrogen, C 1-3 alkyl, or -(CH2) 1-2 -aryl, There is provided a compound or a pharmaceutically acceptable salt thereof.

[0022] In the case of the bridged epidithiodioxopiperazine-based compound according to the present invention, unlike the ETP (EpidiThiodioxoPiperazine) derivative, which is a structure frequently found in natural products, it has a novel structure not found in natural products. As confirmed by the present inventors, the compound of the present invention having such a structure has very high chemical and biological stability, and has the advantages that its pharmacological activity is further improved and its toxicity is greatly improved as compared with the conventional ETP derivative compound disclosed in International Publication WO2013-077709.

[0023] Compared with the conventional ETP derivative compounds disclosed in International Publication WO2013-077709, the compound of the present invention has one more methylene group introduced (i.e., bridged ETP), which imparts stability equivalent to that of a disulfide structure. That is, when comparing the dihedral angles of disulfide bonds, the conventional ETP derivatives are about 10°, with considerable stress on the ring and high reactivity, while the compound of the present invention is about 50 - 60° as seen from the X-ray analysis results, with relatively less stress on the ring. Such relaxation of ring stress is expected to play a role in reducing toxicity by lowering the non-specific reactivity of the compound, and at the same time, it is expected to increase the redox reactivity with thioredoxin (thioredoxin or Trx) protein due to the increased flexibility. However, the present invention is not limited to such a theoretical mechanism.

[0024] In addition, the compound of the present invention has a structural feature that facilitates interaction with the thiol group of the cysteine residue of the C-X-X-C motif of thioredoxin, which is an electron donor for the reduction of hydrogen peroxide, and thus is expected to be more specific and exhibit high activity. For example, in the case of the conventional ETP derivatives disclosed in International Publication WO2013-077709, the pharmacological activity decreases in the case of bulky substituents, but in the chemical formula of the present invention, substituents with large structures such as aryl and heteroaryl also show good activity. However, the present invention is not limited to such theoretical speculation.

[0025] In the compound of the present invention, in the case of a trisulfide compound having three sulfurs or a tetrasulfide compound having four sulfurs, the connection between the sulfurs can be broken in the body and then connected as a disulfide compound. In this case, the trisulfide compound having three sulfurs or the tetrasulfide compound having four sulfurs is expected to act as a prodrug of the disulfide compound having two sulfurs. However, the present invention is not limited to such theoretical speculation.

[0026] As used herein, if a substituent is described as "optionally substituted" or "selectively substituted", the substituent may be (1) unsubstituted or (2) substituted with one or more of the defined substituents. If the position where substitution is possible is unsubstituted, the default substituent is hydrogen.

[0027] As used herein, "alkyl" means a saturated straight-chain or branched acyclic hydrocarbon having 1 to 10 carbon atoms. "Lower alkyl" means alkyl having 1 to 4 carbon atoms. Representative saturated straight-chain alkyls include -methyl, -ethyl, -1-propyl, -1-butyl, -1-pentyl, -1-hexyl, -1-heptyl, -1-octyl, -1-nonyl, and -1-decyl, etc., and branched alkyls include -isopropyl, -sec-butyl, -isobutyl, -tert-butyl, -isopentyl, 2-methylbutyl, 3-methylbutyl, 2-methylpentyl, 3-methylpentyl, 4-methylpentyl, 2-methylhexyl, 3-methylhexyl, 4-methylhexyl, 5-methylhexyl, 2,3-dimethylbutyl, 2,3-dimethylpentyl, 2,4-dimethylpentyl, 2,3-dimethylhexyl, 2,4-dimethylhexyl, 2,5-dimethylhexyl, 2,2-dimethylpentyl, 2,2-dimethylhexyl, 3,3-dimethylpentyl, 3,3-dimethylhexyl, 4,4-dimethylhexyl, 2-ethylpentyl, 3-ethylpentyl, 2-ethylhexyl, 3-ethylhexyl, 4-ethylhexyl, 2-methyl-2-ethylpentyl, 2-methyl-3-ethylpentyl, 2-methyl-4-ethylpentyl, 2-methyl-2-ethylhexyl, 2-methyl-3-ethylhexyl, 2-methyl-4-ethylhexyl, 2,2-diethylpentyl, 3,3-diethylhexyl, 2,2-diethylhexyl, 3,3-diethylhexyl, etc. In a preferred embodiment of the present invention, alkyl is methyl.

[0028] As used herein, "alkenyl" means a saturated straight-chain or branched acyclic hydrocarbon containing 2 to 10 carbon atoms and at least one carbon-carbon double bond. Representative straight-chain and branched (C2-C 10 ) alkenyls include -vinyl, -allyl, -1-butenyl, -2-butenyl, -isobutenylenyl, -1-pentenyl, -2-pentenyl, -3-methyl-1-butenyl, -2-methyl-2-butenyl, -2,3-dimethyl-2-butenyl, -1-hexenyl, -2-hexenyl, -3-hexenyl, -1-heptenyl, -2-heptenyl, -3-heptenyl, -1-octenyl, -2-octenyl, -3-octenyl, -1-nonenyl, -2-nonenyl, -3-nonenyl, -1-disenyl, -2-disenyl, and -3-disenyl. In one embodiment of the present invention, the alkenyl is vinyl or allyl.

[0029] As used herein, the term "alkynyl" means a straight-chain or branched acyclic hydrocarbon having 2 to 10 carbon atoms and containing at least one carbon-carbon triple bond. Representative straight-chain or branched (C2-C 10 ) alkynyls include -ethynyl, -propynyl, -1-butynyl, -2-butynyl, -1-pentynyl, -2-pentynyl, -3-methyl-1-butynyl, -4-pentynyl, -1-hexynyl, -2-hexynyl, -5-hexynyl, -1-heptynyl, -2-heptynyl, -6-heptynyl, -1-octynyl, -2-octynyl, -7-octynyl, -1-nonynyl, -2-nonynyl, -8-nonynyl, -1-decynyl, -2-decynyl, and -9-decynyl. In one embodiment of the present invention, the alkynyl is ethynyl or -propynyl.

[0030] As used herein, when described as "C 1-6 ", "C1-6", "C1-C6" or "C1-C6", this means that the number of carbon atoms is 1 to 6. For example, C 1-6 alkyl means an alkyl having 1 to 6 carbon atoms.

[0031] As used herein, "C 1-6 alkoxy" means -O-(alkyl), where alkyl is as defined above. For example, methoxy, ethoxy, n-propoxy, 1-methylethoxy (isopropoxy), butoxy, 1-methylpropoxy (sec-butoxy), 2-methylpropoxy (isobutoxy), 1,1-dimethylethoxy (tert-butoxy), pentoxy, 1-methylbutoxy, 2-methylbutoxy, 3-methylbutoxy, 1,1-dimethylpropoxy, 1,2-dimethylpropoxy, 2,2-dimethylpropoxy, 1-ethylpropoxy, hexoxy, 1-methylpentoxy, 2-methylpentoxy, 3-methylpentoxy, 4-methylpentoxy, 1,1-dimethylbutoxy, 1,2-dimethylbutoxy, 1,3-dimethylbutoxy, 2,2-dimethylbutoxy, 2,3-dimethylbutoxy, 3,3-dimethylbutoxy, 1-ethylbutoxy, 2-ethylbutoxy, 1,1,2-trimethylpropoxy, 1,2,2-trimethylpropoxy, 1-ethyl-1-methylpropoxy, or 1-ethyl-2-methylpropoxy may be mentioned.

[0032] As used herein, "halogen" is fluorine, chlorine, bromine, or iodine.

[0033] As used herein, "cycloalkyl", when the number of carbon atoms is not particularly limited, is C having 3 to 7 carbon atoms 3-7Cycloalkyl means a monocyclic or polycyclic saturated ring that is cycloalkyl, has carbon and hydrogen atoms, and has no carbon-carbon multiple bonds. For example, monocyclic rings include, but are not limited to, (C3-C7) cycloalkyl (e.g., cyclopropyl, cyclobutyl, cyclopentyl, cyclohexyl, and cycloheptyl). Examples of polycyclic rings include fused bicyclic rings such as octahydropentalene, decahydronaphthalene; spiro rings such as spiro[3.3]heptane, spiro[3.4]octane, spiro[3.5]nonane, spiro[4.4]nonane, spiro[4.5]decane, spiro[5.5]undecane; and bridged bicyclic rings such as bicyclo[2.1.1]hexane, bicyclo[2.2.1]heptane, bicyclo[2.2.2]octane, but are not limited to these. The cycloalkyl group may be optionally substituted. In one embodiment, the cycloalkyl group is a monocyclic ring.

[0034] As used herein, "heterocycle" or "heterocycloalkyl" means a saturated monocyclic 4- to 7-membered or bicyclic 7- to 12-membered ring containing 1 to 4 heteroatoms independently selected from nitrogen, oxygen, and sulfur, where the nitrogen and sulfur heteroatoms may be optionally oxidized and the nitrogen heteroatoms may be optionally quaternized. As used herein, "heterocycloalkyl" means "hetero(C" when the number of carbon atoms is not particularly limited. 3-7) means "cycloalkyl". Representative heterocycles include oxirane, oxetane, tetrahydrofuran, tetrahydropyran, 1,4-dioxane, aziridine, azetidine, pyrrolidine, piperidine, piperazine, pyrrolidinone, hydantoin, valerolactam, thiirane, thietane, tetrahydrothiophene, tetrahydrothiopyran, morpholine, tetrahydropyridine, tetrahydropyrimidine, etc. The heterocycle includes a bicyclic ring in which a part of the heterocycle is fused to a benzene or cyclopenta-1,3-diene ring. The heterocycle can be attached by a heteroatom or a carbon atom. The heterocycle also includes a fused bicyclic ring, a spiro ring, and a bridged bicyclic ring in which one or more carbon atoms of the above-mentioned polycyclic ring are substituted with a nitrogen, oxygen, or sulfur atom. Examples of such are, when the heteroatom is nitrogen, fused hetero-bicyclic rings such as octahydrocyclopenta[c]pyrrole, octahydropyrrolo[3,4-c]pyrrole, decahydroisoquinoline, decahydro-2,6-naphthyridine; spiro rings such as 2-azaspiro[3.3]heptane, 2,6-diazaspiro[3.3]heptane, 2-azaspiro[3.4]octane, 2,6-diazaspiro[3.4]octane, 2-azaspiro[3.5]nonane, 2,7-diazaspiro[3.5]nonane, 2-azaspiro[4.4]nonane, 2,7-diazaspiro[4.4]nonane, 8-azaspiro[4.5]decane, 2,8-diazaspiro[4.5]decane, 3-azaspiro[5.5]undecane, 3,9-diazaspiro[5.5]undecane; and bridged hetero-bicyclic rings such as 2-azabicyclo[2.1.1]hexane, 2-azabicyclo[2.2.1]heptane, 2,5-diazabicyclo[2.2.1]heptane, 2-azabicyclo[2.2.2]octane, 2,5-diazabicyclo[2.2.2]octane, but are not limited thereto. In one embodiment of the present invention, the heterocycle is piperidyl, morpholinyl, or piperazinyl.

[0035] As used herein, the term "aryl" means a carbocyclic aromatic group containing 5 to 10 ring atoms. Typically, phenyl, tolyl, xylyl, naphthyl, tetrahydronaphthyl, anthracenyl, fluorenyl, indenyl, azulenyl, etc. may be mentioned, but it is not limited thereto. The carbocyclic aromatic group may be optionally substituted. In one embodiment of the present invention, aryl is phenyl or naphthyl.

[0036] As used herein, "heteroaryl" is a 5- to 10-membered aromatic heterocyclic ring having at least one heteroatom selected from the group consisting of nitrogen, oxygen and sulfur and containing at least one carbon atom including monocyclic and bicyclic ring systems. Representative heteroaryls are furan, 4H-pyran, pyrrole, imidazole, pyrazole, triazole, tetrazole, pyridine, pyrimidine, pyridazine, pyrazine, triazine, thiophene, oxazole, isoxazole, thiazole, isothiazole, oxadiazole, benzofuran, benzothiophene, quinoline, indole, benzoxazole, benzimidazole, benzothiazole, cinnoline, phthalazine, quinazoline, 1H-azepine, etc. In one embodiment of the present invention, heteroaryl is pyridine, quinoline, pyrazole, thiophene, benzo[d]thiazole, pyrimidine, imidazole, thiazole, isoxazole, indole, quinazoline, or benzimidazole.

[0037] In a preferred embodiment of the present invention, in the above Chemical Formula 1 or Chemical Formula 2, n is 1, R1 and R2 are each independently C 1-3alkyl, -CH2-piperidyl, -CH2-morpholinyl, -CH2-piperazinyl, -CH2-phenyl, -CH2-naphthyl, -CH2-pyridyl, -CH2-quinolinyl, -CH2-pyrazolyl, -CH2-thiophen-2-yl, -CH2-benzo[d]thiazol-2-yl, -CH2-pyrimidyl, -CH2-1H-imidazol-4-yl, -CH2-1H-imidazol-2-yl, -CH2-thiazol-4-yl, -CH2-thiazol-5-yl, -CH2-isoxazolyl, -CH2-indol-2-yl, -CH2-indol-3-yl, -CH2-benzimidazol-5-yl, -CH2-quinolin-4-yl, -CH2-quinazolin-2-yl, or -CH2-quinazolin-4-yl, wherein the piperidyl, morpholinyl, piperazinyl, phenyl, naphthyl, pyridyl, quinolinyl, pyrazolyl, thiophene, benzo[d]thiazole, pyrimidyl, imidazole, thiazole, isoxazolyl, indole, benzimidazole, quinoline, and quinazoline are unsubstituted or are substituted with one or more substituents selected from the group consisting of C 1-3 alkyl, -CF3, C 1-3 alkoxy, CN, halogen, and -COO-C 1-3 alkyl; R3 is C 1-3 alkyl or -CH2-aryl, or R2 and R3 are linked to each other to form any one of the following structures fused to the piperazinedione present in Chemical Formula 1:

[0038]

Chemical formula

[0039] wherein X is O or NR6, R6 is methyl, R4 is hydrogen, R5 is hydrogen, There is provided a compound or a pharmaceutically acceptable salt thereof.

[0040] Among the compounds of the above Chemical Formula 1, in particular, 1,6,8-trimethyl-2,3-dithia-6,8-diazabicyclo[3.2.2]nonane-7,9-dione (Compound 1), 6-benzyl-1,8-dimethyl-2,3-dithia-6,8-diazabicyclo[3.2.2]nonane-7,9-dione (Compound 2), 1,8-dimethyl-6-(3,4,5-trimethoxybenzyl)-2,3-dithia-6,8-diazabicyclo[3.2.2]nonane-7,9-dione (Compound 3), 6-(3,5-difluorobenzyl)-1,8-dimethyl-2,3-dithia-6,8-diazabicyclo[3.2.2]nonane-7,9-dione (Compound 4), 1,8-dimethyl-6-(quinolin-2-ylmethyl)-2,3-dithia-6,8-diazabicyclo[3.2.2]nonane-7,9-dione (Compound 5), 1,8-dimethyl-6-(pyridin-2-ylmethyl)-2,3-dithia-6,8-diazabicyclo[3.2.2]nonane-7,9-dione (Compound 6), 11-benzyltetrahydro-5H,7H-4,9a-(epiminomethano)pyrrolo[2,1-c][1,2,4]dithiazepine-5,10-dione (Compound 7), 12-benzyltetrahydro-5H,10H-4,10a-(epiminomethano)[1,4]oxazino[3,4-c][1,2,4]dithiazepine-5,11-dione (Compound 8), 12-(pyridin-4-ylmethyl)tetrahydro-5H,10H-4,10a-(epiminomethano)[1,4]oxazino[3,4-c][1,2,4]dithiazepine-5,11-dione (Compound 9), 12-ethyltetrahydro-5H,10H-4,10a-(epiminomethano)[1,4]oxazino[3,4-c][1,2,4]dithiazepine-5,11-dione (Compound 10), 11-(pyridin-4-ylmethyl)tetrahydro-5H,7H-4,9a-(epiminomethano)pyrrolo[2,1-c][1,2,4]dithiazepine-5,10-dione (Compound 11), 1,8-dimethyl-6-((6-methylpyridin-2-yl)methyl)-2,3-dithia-6,8-diazabicyclo[3.2.2]nonane-7,9-dione (Compound 12), 1,8-dimethyl-6-((1-methyl-1H-pyrazol-4-yl)methyl)-2,3-dithia-6,8-diazabicyclo[3.2.2] Nonan-7,9-dione (Compound 13), 1,8-dimethyl-6-(thiophen-2-ylmethyl)-2,3-dithia-6,8-diazabicyclo[3.2.2]nonane-7,9-dione (Compound 14), 6-(benzo[d]thiazol-2-ylmethyl)-1,8-dimethyl-2,3-dithia-6,8-diazabicyclo[3.2.2]nonane-7,9-dione (Compound 15), 1,6-dimethyl-8-(pyrimidin-2-ylmethyl)-2,3-dithia-6,8-diazabicyclo[3.2.2]nonane-7,9-dione (Compound 16), 1,6-dimethyl-8-((1-methyl-1H-imidazol-4-yl)methyl)-2,3-dithia-6,8-diazabicyclo[3.2.2]nonane-7,9-dione (Compound 17), methyl 4-((1,6-dimethyl-7,9-dioxo-2,3-dithia-6,8-diazabicyclo[3.2.2]nonan-8-yl)methyl)benzoate (Compound 18), 1-benzyl-6,8-dimethyl-2,3-dithia-6,8-diazabicyclo[3.2.2]nonane-7,9-dione (Compound 19), 6,8-diethyl-1-methyl-2,3-dithia-6,8-diazabicyclo[3.2.2]nonane-7,9-dione (Compound 20), 12-benzyl-9-methylhexahydro-5H-4,10a-(epiminomethano)pyrazino[2,1-c][1,2,4]dithiazepine-5,11-dione (Compound 21), 12-benzyl-9-methylhexahydro-5H-4,10a-(epiminomethano)pyrazino[2,1-c][1,2,4]dithiazepine-5,11-dione (Compound 22), 12-benzyl-9-methylhexahydro-5H-4,10a-(epiminomethano)pyrazino[2,1-c][1,2,4]dithiazepine-5,11-dione (Compound 23), 6-benzyl-1,4,8-trimethyl-2,3-dithia-6,8-diazabicyclo[3.2.2]nonane-7,9-dione (Compound 24), and 6-benzyl-1,4,8-trimethyl-2,3-dithia-6,8-diazabicyclo[3.2.2]nonane-7,9-dione (Compound 25) are more preferable in various aspects such as 2-Cys-Prx mimetic activity and safety.

[0041] The compounds of Chemical Formula 1 or 2 of the present invention can be used in the form of pharmaceutically acceptable salts.

[0042] The term "pharmaceutically acceptable salt" as used in the present invention means all salts that possess the desired biological and / or physiological activity of the said compound and exhibit minimal undesirable toxicological effects. In the present invention, any salt can be used without limitation as long as the diketopiperazine ring containing an intramolecular disulfide bridge is maintained. Acid addition salts formed by pharmaceutically acceptable free acids are useful as salts. Acid addition salts are usually prepared by dissolving the compound in an excess of aqueous solution and precipitating the salt using a water-miscible organic solvent such as methanol, ethanol, acetone or acetonitrile. The same molar amounts of the compound and an acid or alcohol in water (e.g., glycol monomethyl ether) are heated, and then the mixture is evaporated to dryness or the precipitated salt is suction filtered. At this time, inorganic acids and organic acids can be used as the free acid. Examples of inorganic acids include hydrochloric acid, hydrobromic acid, phosphoric acid, nitric acid, sulfuric acid, tartaric acid, etc., and examples of organic acids include methanesulfonic acid, p-toluenesulfonic acid, acetic acid, trifluoroacetic acid, maleic acid, succinic acid, oxalic acid, benzoic acid, tartaric acid, fumaric acid, mandelic acid, propionic acid, citric acid, lactic acid, glycolic acid, gluconic acid, galacturonic acid, glutamic acid, glutaric acid, glucuronic acid, aspartic acid, ascorbic acid, carbonic acid, vanillic acid, hydroiodic acid, etc., but are not limited thereto.

[0043] Also, pharmaceutically acceptable metal salts can be produced using a base. Alkali metal or alkaline earth metal salts are obtained, for example, by dissolving the compound in a solution of an excess of alkali metal hydroxide or alkaline earth metal hydroxide, filtering the insoluble compound salt, and evaporating and drying the filtrate. At this time, it is pharmaceutically preferred to produce sodium, potassium or calcium salts as the metal salts, but is not limited thereto. Also, the corresponding silver salt is obtained by reacting an alkali metal or alkaline earth metal salt with a suitable silver salt (e.g., silver nitrate).

[0044] The pharmaceutically acceptable salts of the compounds according to the present invention include, unless otherwise specified, all salts of acidic or basic groups that may be present. For example, pharmaceutically acceptable salts may include sodium, calcium, and potassium salts of hydroxy groups, and other pharmaceutically acceptable salts of amino groups include hydrobromide, sulfate, bisulfate, phosphate, hydrogen phosphate, dihydrogen phosphate, acetate, succinate, citrate, tartrate, lactate, mandelate, methanesulfonate (mesylate), and p-toluenesulfonate (tosylate), etc., and can be prepared through methods for producing salts well-known in the art.

[0045] As used herein, the term "compound of the present invention" means not only each compound of Chemical Formula 1, but also includes their clathrates, hydrates, solvates, or polymorphs. Also, the term "compound of the present invention" includes pharmaceutically acceptable salts of the compounds of the present invention when their pharmaceutically acceptable salts are not mentioned. In one embodiment, the compound of the present invention may exist as a stereochemically pure compound (e.g., substantially free of other stereoisomers (e.g., 85% ee or more, 90% ee or more, 95% ee or more, 97% ee or more, or 99% ee or more)). That is, when the compound of Chemical Formula 1 according to the present invention or its salt is a tautomeric isomer and / or a stereoisomer (e.g., a geometrical isomer and conformational isomers), their separated isomers and mixtures are each also included in the scope of the compound of the present invention. When the compound of the present invention or its salt has an asymmetric carbon in its structure, their optically active compounds and racemic mixtures are also included in the scope of the compound of the present invention.

[0046] As used herein, the term "polymorph" means a solid crystalline form of a compound of the invention or a complex thereof. Different polymorphs of the same compound exhibit different physical, chemical, and / or spectral properties. Differences in physical properties include, but are not limited to, stability (e.g., thermal or light stability), compressibility and density (important for formulation and product manufacture), and dissolution rate (which can affect biological availability). Differences in stability can cause changes in chemical reactivity (e.g., differential oxidation that causes faster discoloration when composed of one polymorph compared to when composed of other polymorphs), or mechanical characteristics (e.g., a purified fragment stored as a kinetically preferred polymorph converts to a thermodynamically more stable polymorph), or both (tablets of one polymorph are more sensitive to further degradation at high humidity). The different physical properties of the polymorphs can affect their processing. For example, one crystalline polymorph may be more likely to form a solvate, e.g., due to its morphology or particle size distribution, compared to other crystalline polymorphs, or filtration or washing may be more difficult.

[0047] As used herein, the term "solvate" means a compound of the invention or a pharmaceutically acceptable salt thereof that contains a stoichiometric or non-stoichiometric amount of solvent bound by non-covalent intermolecular forces. Preferred solvents are volatile and non-toxic and can be administered to humans in very small amounts.

[0048] As used herein, the term "hydrate" means a compound of the invention or a pharmaceutically acceptable salt thereof that contains a stoichiometric or non-stoichiometric amount of water bound by non-covalent intermolecular forces.

[0049] As used herein, the term "clathrate" means a crystalline lattice compound of the invention or a salt thereof that contains a space (e.g., a channel) that entraps a guest molecule (e.g., a solvent or water).

[0050] As used herein, the term "purified" means that when separated, the separated entity is 90% or more pure, in one embodiment 95% or more pure, in other embodiments 99% or more pure, and in still other embodiments 99.9% or more pure.

[0051] "2-Cys-Prx" is a thiol-specific antioxidant enzyme that plays a role in protecting cells through peroxidase activity that reduces hydrogen peroxide, peroxynitrite, and other hydroperoxides intracellularly. Its functional unit is a homodimer and has an intramolecular redox-active disulfide center that plays an important role in the activity of the enzyme. Each cysteine of the dimer exists as a thiol group in the reduced state. On the other hand, during the reaction with peroxide, the peroxidized cysteine of 2-Cys-Prx is oxidized to a sulfenic acid intermediate and is distinguished from the cysteine of the other subunit of the dimer that maintains the thiol group, and the sulfenic acid group and the thiol group of the intermediate undergo dehydration condensation to form an intramolecular disulfide cross-link. For example, intracellular 2-Cys-Trx reduces intracellular hydrogen peroxide while being oxidized from the reduced form containing two thiol groups to the oxidized form in which the two thiol groups form an intramolecular disulfide bond. At this time, the oxidized form of 2-Cys-Prx containing an intramolecular disulfide bond can be coupled with a redox system including thioredoxin (Trx), thioredoxin reductase (TR); alkyl hydroperoxide reductase (AhpF); trypanothione reductase, trypanothione, and tryparedoxin or lipoamide dehydrogenase, SucB (dihydrolipoyltranssuccinylase), and AhpD, etc., and converted to the reduced form which is the active form having two thiol groups.

[0052] The compounds of the present invention mimic 2-Cys-Prx activity intracellularly (especially in coupling with the thioredoxin (Trx) and thioredoxin reductase (TR) systems), and exhibit excellent activity in suppressing the proliferation and migration of vascular smooth muscle cells induced by PDGF. Conversely, the compounds of the present invention promote the proliferation and migration of VEGF-induced vascular endothelial cells.

[0053] Accordingly, one aspect of the present invention provides a pharmaceutical composition for preventing or treating vascular diseases, comprising, as an active ingredient, a compound represented by the above chemical formula 1 or chemical formula 2 or a pharmaceutically acceptable salt thereof.

[0054] Another aspect of the present invention provides a method for treating or preventing vascular diseases, comprising the step of administering a therapeutically effective amount of a compound represented by the above chemical formula 1 or chemical formula 2 or a pharmaceutically acceptable salt thereof to an individual in need of prevention or treatment of vascular diseases or an individual suspected of having vascular diseases.

[0055] Still another aspect of the present invention provides the use of a compound represented by chemical formula 1 or 2 or a pharmaceutically acceptable salt thereof for the manufacture of a medicament for the treatment or prevention of vascular diseases.

[0056] In the pharmaceutical composition, method or use, the vascular disease may be hypertension (particularly, pulmonary hypertension), ischemic coronary artery disease (e.g., angina pectoris, myocardial infarction, unstable angina), cerebral artery occlusion (e.g., stroke), arteriosclerosis (e.g., cardiac arteriosclerosis, carotid arteriosclerosis), peripheral arterial occlusion disease (e.g., Buerger's disease), thromboembolism, diabetic foot lesion, venous ulcer, deep vein thrombosis, vasoconstriction, arteritis and vascular restenosis induced by hyperplasia of vascular smooth muscle and / or loss of vascular endothelium. That is, the pharmaceutical composition according to the present invention can be used for the treatment or prevention of any of the above vascular diseases. In one embodiment of the present invention, the vascular disease is ischemic coronary artery disease, arteriosclerosis, vascular restenosis, or pulmonary hypertension. In another embodiment of the present invention, the vascular disease is pulmonary hypertension.

[0057] In one embodiment of the present invention, the vascular restenosis may be vascular restenosis induced by vascular transplantation, vascular dissection, arteriosclerosis, fat accumulation in blood vessels, hypertension, vascular inflammation or angioplasty. The reason for the occurrence of vascular restenosis has not been clarified, but after vascular injury due to various reasons or damage to the vascular endothelium by the device inserted during angioplasty, it is known that vascular smooth muscle cells abnormally migrate and proliferate due to growth factors and / or cytokines secreted from surrounding cells as a mechanism for recovering it, causing intimal hyperplasia of blood vessels. The blood vessels include, but are not limited to, aorta, carotid artery, coronary artery, peripheral artery, renal artery, etc.

[0058] In one embodiment of the present invention, the pulmonary hypertension is a type of hypertension that affects the pulmonary arteries and the right side of the heart, and is the case where the mean pulmonary artery pressure at rest is 25 mmHg or more and the mean pulmonary artery pressure during exercise is 30 mmHg or more. In one form of pulmonary hypertension, the small arteries and capillaries in the lungs called pulmonary arteries become narrow, clogged, or damaged, which makes it difficult for blood to flow through the lungs and increases the pressure in the pulmonary arteries. As a result, due to the applied pressure, the lower part of the right atrium (right ventricle) has difficulty pumping blood through the lungs, and ultimately there is a risk of inducing right ventricular hypertrophy and losing heart function.

[0059] The composition according to the present invention may further contain suitable carriers, excipients, and diluents commonly used in the manufacture of pharmaceutical compositions. The composition is sterilized or aseptic and may be water, a buffer, an isotonic agent, etc., and the solution is sterilized or aseptic. Further, when the composition is applied to animals or humans, it may contain other components known to those with ordinary knowledge in the art that do not cause allergic or other harmful reactions.

[0060] The term "pharmaceutically acceptable carrier" used in the present invention includes any solvent, dispersion medium, coating agent, antibacterial agent, antifungal agent, isotonic agent, etc. It is known in the art to use the medium and formulation as a pharmaceutical carrier. Their use in therapeutic compositions is conceivable other than in normal media or formulations that are immiscible with the active ingredient. Further, supplementary active ingredients may be incorporated into the composition.

[0061] The composition may be manufactured in dosage forms such as solutions, emulsions, suspensions, or creams and may be used parenterally. The dosage of the composition may be used at the normal dosage for the prevention or treatment of vascular diseases, and it is preferably applied at different dosages according to the patient's age, sex, health condition, absorption degree of the active ingredient in the body, inactivation rate, and drugs used in combination.

[0062] In the present invention, "prevention" means any act of suppressing or delaying the onset of vascular diseases by administration of the pharmaceutical composition according to the present invention, and "treatment" means any act of improving or favorably changing the symptoms caused by vascular diseases by administration of the pharmaceutical composition.

[0063] In the present invention, "individual" means all animals including humans in whom vascular diseases have occurred or may occur, and by administering the pharmaceutical composition of the present invention to an individual, the vascular diseases can be effectively prevented or treated. Further, the pharmaceutical composition of the present invention may be administered in parallel with known therapeutic agents for vascular diseases.

[0064] The compound of the present invention or a pharmaceutically acceptable salt thereof is administered in a therapeutically effective amount. The "therapeutically effective amount" means an amount sufficient to treat a disease with a reasonable benefit / risk ratio applicable to medical treatment and that does not cause side effects, and the effective dosage level can be easily determined by those skilled in the art depending on factors including the sex, age, weight, health status, severity of the disease, activity of the drug, sensitivity to the drug, administration method, administration time, administration route and excretion rate, treatment period, ingredients including drugs formulated or used simultaneously, and other factors known in the medical field.

[0065] For example, the compound of the present invention or a pharmaceutically acceptable salt thereof can be administered by any appropriate route, in the form of a pharmaceutical composition appropriate for such route, in an effective dosage for the intended treatment. The effective dosage is generally about 0.0001 to about 100 mg / kg of body weight per day, preferably about 0.001 to about 50 mg / kg / day, either as a single or divided dose. More preferably, it is about 0.005 to about 20 mg / kg / day. Depending on age, species, and the disease or condition being treated, dosage levels below the lower limit of this range may be appropriate. In other cases, even larger dosages may be used without harmful side effects. Larger dosages can be divided into several smaller dosages for administration in a day.

[0066] In the present invention, "administration" means introducing a predetermined substance into a patient by an appropriate method, and the administration route of the composition can be any general route as long as it can reach the target tissue. As the administration method, oral administration, intravenous administration, subcutaneous administration, intraperitoneal administration, etc. can be used, but it is not limited thereto. In a preferred embodiment of the present invention, the compound of the present invention is administered orally. In other embodiments of the present invention, the compound of the present invention can be locally administered to the lesion. In particular, the compound of the present invention can be administered using a drug-eluting stent. That is, the compound of the present invention can be directly administered to the stenosis site by coating it in or on the stent. A double balloon catheter, dispatch, microporous balloon, etc. can be used for local administration of the drug, and in particular, a stent or sustained-release microparticles can be used for transmitting the drug for a long period of time.

[0067] Further, one aspect of the present invention provides a drug delivery device for local administration comprising a pharmaceutical composition for preventing or treating vascular diseases according to the present invention. The drug delivery device for local administration may include, but is not limited to, a double balloon catheter, dispatch, microporous balloon, stent, etc., and may preferably be a stent.

[0068] In the present invention, "stent" means a general device for endoluminal application, for example, for application into a blood vessel as described above. In particular, when a disease occurs at a site where the blood flow must be smooth and an obstacle occurs in the flow, the stent is inserted into a narrowed or blocked blood vessel site under fluoroscopy instead of performing a surgical laparotomy to normalize the blood flow, and means a cylindrical medical material. In a preferred embodiment of the present invention, the stent is a sustained-release drug-releasing stent.

[0069] As a method for coating the stent with the pharmaceutical composition of the present invention, ordinary coating methods known to those with ordinary knowledge in the technical field of the present invention can be applied. For example, there are dip coating methods and polymer coating methods. The dip coating method is the simplest coating method, and since only the pharmaceutical composition is coated, it is easy to observe the biological effects of only the drug, but it is not limited thereto. Preferably, the stent of the present invention can be produced by mixing and coating the composition with a polymer substance on a drug-releasing stent so that the composition of the present invention is gradually released. Polymer substances that can be used for drug-releasing stents are well-known in the art. For example, polyurethane, polyethylene terephthalate, PLLA-polyglycolic acid copolymer (PLGA), polycaprolactone, poly(hydroxybutyrate / hydroxyvalerate) copolymer, polyvinyl pyrrolidone, polytetrafluoroethylene, poly(2-hydroxyethyl methacrylate), poly(ether urethane urea), silicone, polyacrylic acid, polyepoxide, polyester, urethane, parylene, polyphosphazene polymer, fluoropolymer, polyamide, polyolefin, and mixtures thereof, etc., but are not limited thereto.

[0070] The stent can be composed of or coated with one or more substances selected from polysaccharides, heparin, gelatin, collagen, alginate, hyaluronic acid, alginic acid, carrageenan, chondroitin, pectin, chitosan and their derivatives and copolymers, and can be further coated to form a layer containing these or an antithrombotic agent. These substances can be appropriately integrated into a biocompatible top coat as disclosed in US Patent Publication US2006 / 0083772.

[0071] Another aspect of the present invention further provides a method for producing a compound represented by Chemical Formula 1', which has improved intracellular permeability and is reduced intracellularly to mimic 2-Cys-Prx activity, including the step of forming an intramolecular disulfide cross-linkage from a 3-mercapto-6-methylmercaptopyrazinedione derivative compound represented by the following Chemical Formula 2 by an oxidation reaction.

[0072] As described above, unless otherwise specified, when stereoisomers are possible for the compounds represented by the chemical formulas of the present invention, such stereoisomers are also included within the scope of the compounds and production methods according to the present invention.

[0073]

Chemical formula

[0074]

Chemical formula

[0075] (In Chemical Formulas 1' and 2, R1, R2, R3, R4, and R5 are the same as those in Chemical Formula 1 described above.) For example, the derivative of the present invention in which R5 is hydrogen in Chemical Formula 1' can be produced by oxidizing 3-mercapto-6-methylmercaptopyrazinedione represented by Chemical Formula 2. At this time, the oxidation reaction can be carried out using reactions known in the art without limitation. Preferably, iodine (I2) or DDQ (2,3-dichloro-5,6-dicyano-1,4-benzoquinone) can be used, but it is not limited thereto.

[0076] Derivatives in which R5 is not hydrogen in Chemical Formula 1' can be produced by treating a compound of Chemical Formula 1' in which R5 is hydrogen with an appropriate electrophile.

[0077] Another aspect of the present invention provides a method for producing a 3-mercapto-6-methylmercaptopyperazinedione derivative represented by Chemical Formula 2, characterized by reducing an intermediate having a ring structure containing 3 to 4 sulfur atoms represented by the following Chemical Formula 3 to form two thiol groups of Chemical Formula 2.

[0078]

Chem.

[0079] (In Chemical Formula 3, R1, R2, R3, and R4 are the same as those in Chemical Formula 1 described above, R5 is hydrogen, and n is 2 or 3.) At this time, the reduction reaction can be carried out without limitation using reactions known in the art. Preferably, it can be carried out using a hydride-based reducing agent such as sodium borohydride or lithium borohydride. More preferably, it can be carried out using sodium borohydride, but it is not limited thereto.

[0080] Another aspect of the present invention provides a method for producing a compound represented by Chemical Formula 3, characterized by reacting a 6-(1-hydroxyalkyl)piperazine-2,5-dione derivative represented by the following Chemical Formula 4 (where R5 = hydrogen) with (a) sulfur (S8) and (b) LiHMDS (lithium bis(trimethylsilyl)amide) or NaHMDS (sodium bis(trimethylsilyl)amide).

[0081]

Chem.

[0082] (In Chemical Formula 4, R1, R2, R3, and R4 are the same as those in Chemical Formula 1 described above, R5 is hydrogen, and R is a protecting group.) In Chemical Formula 4, R is a protecting group, and protecting groups for hydroxy groups known in the art can be used without limitation. Preferably, silicon-based protecting groups such as TBDMS (t-butyldimethylsilyl) and TMS (trimethylsilyl); ether-based protecting groups such as t-butyl; ester-based protecting groups such as acetyl; protecting groups such as dihydropyran can be used, but are not limited thereto.

[0083] In the case of a general 3,6-substituted-piperazinedione, when reacted with sulfur under strong alkali conditions, a ring structure in which sulfur atoms are directly bonded to the 3,6-positions is formed. On the other hand, in the case of Chemical Formula 4, after the hydroxymethyl group is first removed under strong alkali conditions, an intermediate of α,β-unsaturated ketone with very strong reactivity is generated, and this intermediate immediately reacts with sulfur to form a specific ring structure having a methylene bridge as in Chemical Formula 3. At this time, the number of sulfur atoms forming the ring structure is three or four, which varies depending on the structure of the starting material and reaction conditions (see the following mechanism).

[0084]

Chemical Formula

[0085] Another aspect of the present invention is (S1) reacting a compound represented by the following Chemical Formula 4 with (a) sulfur (S8) and (b) LiHMDS (lithium bis(trimethylsilyl)amide) or NaHMDS (sodium bis(trimethylsilyl)amide) to produce a compound represented by the following Chemical Formula 3, (S2) reducing the compound of Chemical Formula 3 below to produce a compound represented by the following Chemical Formula 2; and (S3) A method for producing a compound represented by the following Chemical Formula 1' is provided, which includes forming an intramolecular disulfide cross-linkage from the compound represented by the following Chemical Formula 2.

[0086]

Chemical Formula

[0087]

Chem.

[0088]

Chem.

[0089]

Chem.

[0090] In Chemical Formulas 4, 3, 2 and 1’, R1, R2, R3, and R4 are the same as defined in Chemical Formula 1 described above, R5 is hydrogen, R is a protecting group, and n is 2 or 3.

[0091] As reaction reagents, the reagents described in the above method can be used in the embodiments of the present invention.

[0092] 〔Advantages of the Invention〕 The present invention provides a piperazinedione compound containing a -CH2-S-(S)n- bridge, and a pharmaceutically acceptable salt thereof. The compound of the present invention contains an intramolecular disulfide cross-linkage, which improves intracellular permeability, and is rapidly reduced in cells to a compound having two thiol groups, mimicking the function of 2-Cys-Prx. In particular, the compound of the present invention mimics the function of the PrxII isoform in arterial vascular cells to inhibit the migration and proliferation of PDGF-induced vascular smooth muscle cells and inhibit intimal hyperplasia, while promoting the migration and proliferation of VEGF-induced vascular endothelial cells to improve re-endothelialization. Therefore, the compound and the pharmaceutically acceptable salt thereof according to the present invention can be usefully used as a pharmaceutical composition for preventing or treating vascular diseases. In particular, the compound or the salt thereof of the present invention is useful for treating or preventing vascular diseases such as ischemic coronary artery disease, arteriosclerosis, vascular restenosis, and pulmonary hypertension.

[0093] 〔Brief Description of the Drawings〕 [Figure 1] Figures 1 to 3 show the results of evaluating the effect of the compound of the present invention on PDGF-induced tyrosine phosphorylation by immunoblot analysis in human aortic smooth muscle cells (HASMCs) and human pulmonary artery smooth muscle cells (PASMCs) in which PrxII has been removed using siRNA. Figure 1 shows the results of confirming the concentration-dependent effect of Compound 1 on the degree of Tyr857 phosphorylation of platelet-derived growth factor receptor-β (PDGFRβ) amplified by PrxII gene deficiency in HASMCs.

[0094] [Figure 2] This shows the results of evaluating the effects of Compounds 1, 2, 5, 6, 10, 15, 19, 20, 21, 22, and 23 on the degree of Tyr857 phosphorylation of platelet-derived growth factor receptor-β amplified by PrxII gene deficiency in HASMCs.

[0095] [Figure 3] This shows the results of evaluating the effect of Compound 8 on the degree of PDGF-induced intracellular tyrosine phosphorylation amplified by PrxII gene deficiency and the phosphorylation of signal transduction proteins such as platelet-derived growth factor receptor-β and downstream PLC-γ1 in PASMCs.

[0096] [Figure 4] Figures 4 to 6 show the results of evaluating the effect of the compound of the present invention on VEGF-induced tyrosine phosphorylation by immunoblot analysis in human aortic endothelial cells (HAECs) and human pulmonary artery endothelial cells (PAECs) in which PrxII has been removed using siRNA. Figure 4 shows the results of confirming the effect of Compound 1 on the degree of Tyr1175 phosphorylation of vascular endothelial growth factor receptor-2 (VEGFR2) amplified by PrxII gene deficiency in HAECs for each concentration.

[0097] [Figure 5] This shows the results of evaluating the effects of Compounds 1, 2, 5, 6, 10, 20, 21, and 23 on the degree of VEGF receptor phosphorylation amplified by PrxII gene deficiency in HAECs.

[0098] [Figure 6] Results of analyzing the degree of VEGF-induced intracellular tyrosine phosphorylation amplified by PrxII gene deficiency and the effect of Compound 8 on the phosphorylation of signal transduction proteins such as VEGF receptor-2 and downstream ERK2 in PAECs.

[0099] [Figure 7] Figures 7 to 9 show the results of preclinical efficacy evaluation using a pulmonary arterial hypertension rat model induced by SuGen and hypoxia. Figure 7 shows a significant decrease in right ventricular systolic pressure and inhibition of right ventricular hypertrophy compared to the vehicle control group.

[0100] [Figure 8] Results showing the reopening of occluded blood vessels in the vehicle control group.

[0101] [Figure 9] Results of immunofluorescence staining showing inhibition of pulmonary artery endothelial cell proliferation and smooth muscle cell hyperplasia.

[0102] [Mode for Carrying Out the Invention] Hereinafter, for the purpose of assisting in the understanding of the present invention, examples and the like will be given for detailed explanation. However, the examples according to the present invention can be deformed into various other forms, and it should not be construed that the scope of the present invention is limited to the following examples. The examples of the present invention are provided to more fully explain the present invention to those with average knowledge in the field to which the present invention belongs.

[0103] In the case of the amino acid used as a starting material in the production examples described later, the easily available L-form was used. However, it may also be synthesized using the D-form or racemic form.

[0104] Production Example 1: Synthesis of 1,6,8-trimethyl-2,3-dithia-6,8-diazabicyclo[3.2.2]nonane-7,9-dione (Compound 1)

[0105] [Chemical formula]

[0106] Step 1. Methyl N-(N-benzyloxycarbonyl)-O-t-butyl-L-seryl)-N-methyl-L-alanine (Intermediate 3) To a solution of Intermediate 2 (8.32 g, 54.2 mmol, 1.00 eq, HCl) in DMF (100 mL), DIPEA (35.0 g, 271 mmol, 47.2 mL, 5.00 eq) was added, followed by the addition of Intermediate 1 (16.0 g, 54.2 mmol, 1.00 eq) and HATU (30.9 g, 81.3 mmol, 1.50 eq). The reaction mixture was stirred at 25 °C for 12 h. Water (1000 mL) was added to the reaction mixture, and the mixture was extracted with ethyl acetate (100 mL × 2). The combined organic layers were washed with brine (200 mL), dried over Na2SO4, filtered, and concentrated under reduced pressure to obtain a residue. This residue was purified by chromatography on silica to give Intermediate 3 (20.0 g, 48.2 mmol, 88.9% yield) as a yellow oil. MS (ESI) m / z = 395.1 [M+1] + 。

[0107] Step 2. (3S,6S)-3-(t-Butoxymethyl)-1,6-dimethylpiperazine-2,5-dione (Intermediate 4) To a mixture of Intermediate 3 (35.0 g, 88.7 mmol, 1.00 eq) in MeOH (450 mL) was added Pd / C (3.00 g, 10.0% purity). The mixture was stirred at 50 °C for 12 h under H2 (50 Psi). After filtration, the filtrate was concentrated under reduced pressure to give Intermediate 4 (15.0 g, 65.7 mmol, 74.1% yield) as a pale yellow oil.

[0108] Step 3. (3S,6S)-3-(t-Butoxymethyl)-1,4,6-trimethylpiperazine-2,5-dione (Intermediate 5) Under nitrogen, NaH (3.99 g, 99.9 mmol, 60% purity, 1.20 eq) and MeI (186 g, 1.31 mmol, 81.4 mL, 15.7 eq) were added to a stirred solution of Intermediate 4 (19.0 g, 83.2 mmol, 1.00 eq) in DMF (500 mL) at 0 °C. The resulting solution was stirred for 12 hours under a nitrogen atmosphere. Water (2 L) was added to the reaction mixture, and the mixture was extracted with CH2Cl2 (300 mL × 3). The combined organic phases were washed with brine (500 mL × 2), dried over anhydrous Na2SO4, filtered, and concentrated in vacuo. The product was purified on silica to give Intermediate 5 (19.0 g, 78.4 mmol, 94.2% yield) as a pale yellow oil.

[0109] Step 4. (3S,6S)-3-(((tert-Butyldimethylsilyl)oxy)methyl)-1,4,6-trimethylpiperazine-2,5-dione (Intermediate 6) TFA (77.0 g, 675 mmol, 50.0 mL, 8.61 eq) was added to a mixture of Intermediate 5 (19.0 g, 78.4 mmol, 1.00 eq) in CH2Cl2 (50.0 mL). The mixture was stirred at 25 °C for 12 hours. The reaction mixture was concentrated under reduced pressure. The residue was dissolved in DMF (200 mL). TBSCl (14.6 g, 96.7 mmol, 11.9 mL, 1.20 eq) and imidazole (16.5 g, 242 mmol, 3.00 eq) were added to the mixture. The reaction mixture was stirred at 25 °C for 1 hour. Water (1000 mL) was added to the reaction mixture, and the mixture was extracted with CH2Cl2 (300 mL × 3). The combined organic phases were washed with brine (200 mL × 2), dried over anhydrous Na2SO4, filtered, and concentrated in vacuo. The residue was purified on silica gel to give Intermediate 6 (20.0 g, 66.6 mmol, 82.6% yield) as a pale yellow solid.

[0110] Step 5. 1,6,8-Trimethyl-2,3-dithia-6,8-diazabicyclo[3.2.2]nonane-7,9-dione (Compound 1) At room temperature, sulfur (853 mg, 26.63 mmol, 8 eq) was placed in anhydrous THF (53 mL). After slowly adding NaHMDS (10 mL, 1 M in THF, 10 mmol), the mixture was stirred for 10 minutes. Intermediate 6 (1 g, 3.33 mmol, 1 eq) dissolved in THF (12 mL) was added dropwise at room temperature for 10 minutes, and then NaHMDS (8.3 mL, 1 M in THF, 8.3 mmol) was gradually added dropwise. After stirring the reaction mixture for 75 minutes, ammonium chloride solution (200 mL) was added to the reaction mixture, and the mixture was extracted with CH2Cl2 (200 mL x 2). The combined organic phases were washed with brine (100 mL × 2), dried over anhydrous Na2SO4, filtered, and concentrated in vacuo. The residue was purified on silica gel to obtain the trisulfide cyclic intermediate 7, which was used in the next reaction.

[0111] Intermediate 7 (200 mg, 0.64 mmol) was dissolved in THF (10 mL) and MeOH (10 mL) at room temperature and then cooled to 0 °C. NaBH4 (0.12 g, 3.2 mmol, 5 eq) was gradually added dropwise at 0 °C, and then the temperature was raised to room temperature and stirred for about 30 minutes. The reaction mixture was concentrated under reduced pressure. Water (10 mL) and aqueous NaHCO3 solution (40 mL) were added to the reaction mixture and stirred at 0 °C. The mixture was washed with CH2Cl2 (100 mL × 3) to remove impurities. The aqueous layer was acidified with 1 N aqueous hydrochloric acid and then extracted with CH2Cl2 (200 mL × 3). The combined organic phases were washed with brine (100 mL × 2), dried over anhydrous Na2SO4, filtered, and concentrated in vacuo to 100 mL to obtain intermediate 8, which was used in the next reaction without purification immediately.

[0112] In another reaction vessel, 2,3-dichloro-5,6-dicyano-1,4-benzoquinone (DDQ, 120 mg, 0.8 eq) was dissolved in THF (10 mL) and CH₂Cl₂ (100 mL). The solution of Intermediate 8 obtained as described above was gradually added dropwise thereto, and then the reaction was carried out at room temperature for 30 minutes. An aqueous NaHCO₃ solution (200 mL) was added to the reaction mixture, and the mixture was extracted with CH₂Cl₂ (200 mL × 3). The combined organic phases were washed with brine (200 mL × 2), dried over anhydrous Na₂SO₄, filtered, and concentrated in vacuo. The residue was purified on silica gel to obtain 101 mg of Compound 1 as a pale yellow solid.

[0113] 1 H NMR (400 MHz, CDCl₃) δ 4.29 (s, 1H), 3.09 (s, 3H), 3.02 (s, 3H), 2.82 (dd, J = 12.8, 7.5 Hz, 2H), 1.86 (s, 3H). 13 C NMR (101 MHz, CDCl₃) δ 168.79 (s), 166.79 (s), 70.19 (s), 63.49 (s), 33.52 (s), 28.56 (s), 28.13 (s), 21.43 (s).。

[0114] Production Example 2: Synthesis of 6-benzyl-1,8-dimethyl-2,3-dithia-6,8-diazabicyclo[3.2.2]nonane-7,9-dione (Compound 2)

[0115]

Chemical Structure

[0116] Step 1. (3S,6S)-1-benzyl-6-(t-butoxymethyl)-3,4-dimethylpiperazine-2,5-dione (Intermediate 2) Using Intermediate 1 (the same substance as Intermediate 4 synthesized in Production Example 1), it was synthesized in the same manner as the production method of Intermediate 5 in Production Example 1. However, benzyl bromide was used instead of iodomethane.

[0117] 2.6-Benzyl-1,8-dimethyl-2,3-dithia-6,8-diazabicyclo[3.2.2]nonane-7,9-dione (Compound 2) It was produced by a synthetic method similar to Compound 1 in Production Example 1. That is, 1 g of Intermediate 3 was obtained using Intermediate 2 (1.9 g, 5.97 mmol), sulfur (1.53 g, 8 eq), and NaHMDS (29.8 mL, 1 M in THF, 29.8 mmol). Thereafter, Intermediate 4 was obtained using Intermediate 3 (0.8 g, 2.35 mmol) and NaBH4 (0.44 g, 5 eq), and it was used immediately in the next reaction without purification. Thereafter, 0.44 g of Compound 2 was obtained using Intermediate 4 and DDQ (0.426 g, 0.8 eq).

[0118] 1 H NMR (400 MHz, CDCl3) δ 7.37 - 7.29 (m, 9H), 7.26 (t, J = 7.8 Hz, 8H), 5.20 (d, J = 14.9 Hz, 1H), 4.75 (d, J = 14.7 Hz, 3H), 4.47 (d, J = 14.8 Hz, 3H), 4.30 (d, J = 4.8 Hz, 3H), 4.01 (d, J = 14.8 Hz, 1H), 3.88 (d, J = 14.3 Hz, 1H), 3.32 (dd, J = 14.4, 6.4 Hz, 1H), 3.09 (s, 8H), 2.98 (s, 2H), 2.59 (dt, J = 23.7, 9.9 Hz, 6H), 1.93 (s, 2H), 1.91 (s, 8H) 13 C NMR (101 MHz, CDCl3) δ 168.80 (s), 166.95 (s), 135.17 (s), 129.24 (s), 128.69 (s), 128.48 (s), 69.81 (s), 61.02 (s), 49.67 (s), 28.55 (d, J = 7.5 Hz), 21.61 (s).

[0119] Production Example 3: Synthesis of 1,8-dimethyl-6-(3,4,5-trimethoxybenzyl)-2,3-dithia-6,8-diazabicyclo[3.2.2]nonane-7,9-dione (Compound 3)

[0120] [Chemical formula]

[0121] Step 1. Methyl-(3,4,5-trimethoxybenzyl)-L-serine (Intermediate 1) After dissolving serine methyl ester (5 g, 32.1 mmol) in MeOH (50 mL), 3,4,5-trimethoxybenzaldehyde (3.25 g, 1 eq) was added and the mixture was cooled to 0 °C. Et3N (4.48 mL, 1 eq) was added and the mixture was stirred for 2 hours. Then, NaBH4 (2.41 g, 2 eq) was added dropwise at 0 °C and the mixture was stirred for 2 hours. After the reaction mixture was cooled to room temperature, the solvent was removed under reduced pressure and the residue was quenched with brine. The reaction mixture was extracted with CH2Cl2, and the organic layer was washed with brine and dried over anhydrous Na2SO4. After removing the solvent under reduced pressure, the obtained product was purified on silica to obtain Intermediate 1 (2.41 g).

[0122] Step 2. (3S,6S)-3-(Hydroxymethyl)-1,6-dimethyl-4-(3,4,5-trimethoxybenzyl)piperazine-2,5-dione (Intermediate 2) At room temperature, Intermediate 1 (2.41 g, 8.1 mmol) was dissolved in CH2Cl2 (150 mL). Then, NaHCO3 (1.0 g, 1.5 eq) dissolved in a small amount of water was added. Fmoc-N-Me-alanine chloride (2.77 g, 1 eq) dissolved in CH2Cl2 (40 mL) was added to the Intermediate 1 solution, and the mixture was stirred at room temperature for 3 hours. After adding water (100 mL) to the reaction vessel, the organic layer was washed with 1N HCl and brine, and then concentrated under reduced pressure. Ethanol (150 mL) and piperidine (0.8 mL, 8.1 mmol, 1 eq) were added to the obtained product, and the mixture was stirred for 4 hours while refluxing. After the reaction mixture was cooled to room temperature, the organic solvent was concentrated under reduced pressure, and the product was purified on silica to obtain Intermediate 2 (0.92 g), which was immediately used in the next reaction.

[0123] Step 3. (3S,6S)-3-(((tert-butyldimethylsilyl)oxy)methyl)-1,6-dimethyl-4-(3,4,5-trimethoxybenzyl)piperazine-2,5-dione (Intermediate 3) To a stirred solution of Intermediate 2 (0.91 g, 2.58 mmol) in anhydrous CH2Cl2 (20 mL) was added tert-butyldimethylsilyl chloride (TBSCl, 0.78 g, 2 eq) at 0 °C under a nitrogen atmosphere. Then, a solution of imidazole (0.35 g, 2 eq) in anhydrous CH2Cl2 (5 mL) was slowly added thereto. The mixture was stirred at 20 °C for 12 h and then quenched with water (20 mL). The organic layer was washed twice with water and brine (100 mL) and dried over anhydrous Na2SO4. The organic layer was concentrated to completely remove the solvent under reduced pressure, and the resulting substance was purified on silica to obtain Intermediate 3 (1.21 g).

[0124] Step 4. 1,8-Dimethyl-6-(3,4,5-trimethoxybenzyl)-2,3-dithia-6,8-diazabicyclo[3.2.2]nonane-7,9-dione (Compound 3) It was prepared by a synthetic method similar to that of Compound 1 in Production Example 1. That is, 0.53 g of Intermediate 4 was obtained using Intermediate 3 (1.2 g, 2.57 mmol), sulfur (0.66 g, 8 eq), and NaHMDS (14.1 mL, 1 M in THF, 14.1 mmol). After obtaining Intermediate 5 using Intermediate 4 (0.25 g, 0.58 mmol) and NaBH4 (0.1 g, 5 eq), it was used immediately in the next reaction without purification. Then, 0.125 g of Compound 3 was obtained using Intermediate 5 and DDQ (0.094 g, 0.42 mmol, 0.8 eq).

[0125] 11H NMR (400 MHz, CDCl3) δ 6.48 (s, 2H), 4.65 (d, J = 14.7 Hz, 1H), 4.43 (d, J = 14.7 Hz, 1H), 4.35 (dd, J = 5.7, 2.0 Hz, 1H), 3.84 (s, 6H), 3.83 (s, 3H), 3.10 (s, 3H), 2.57 (qd, J = 14.0, 3.9 Hz, 2H), 1.91 (s, 3H) 13 13C NMR (101 MHz, CDCl3) δ 168.85, 166.98, 153.82, 138.07, 130.98, 105.72, 69.85, 61.53, 61.05, 56.40, 50.22, 28.62, 28.39, 21.60。

[0126] Production Example 4: Synthesis of 6-(3,5-difluorobenzyl)-1,8-dimethyl-2,3-dithia-6,8-diazabicyclo[3.2.2]nonane-7,9-dione (Compound 4)

[0127]

Chemical Structure

[0128] Step 1. Methyl-(3,5-difluorobenzyl)-L-serine (Intermediate 1) It was produced by a synthetic method similar to Intermediate 1 of Production Example 3. However, 2,4-difluorobenzaldehyde was used instead of 3,4,5-trimethoxybenzaldehyde.

[0129] Step 2. (3S,6S)-1-(3,5-difluorobenzyl)-6-(hydroxymethyl)-3,4-dimethylpiperazine-2,5-dione (Intermediate 2) It was produced by a synthetic method similar to Intermediate 2 of Production Example 3. However, Fmoc-N-Me-alanine was used instead of Fmoc-N-Me-alanine chloride.

[0130] Step 3. (3S,6S)-3-(((tert-Butyldimethylsilyl)oxy)methyl)-4-(3,5-difluorobenzyl)-1,6-dimethylpiperazine-2,5-dione (Intermediate 3) It was produced by a synthetic method similar to Intermediate 3 of Production Example 3.

[0131] Step 4. 6-(3,5-Difluorobenzyl)-1,8-dimethyl-2,3-dithia-6,8-diazabicyclo[3.2.2]nonane-7,9-dione (Compound 4) It was produced by a synthetic method similar to Compound 1 of Production Example 1. That is, 0.26 g of Intermediate 4 was obtained using Intermediate 3 (0.86 g, 2.08 mmol), sulfur (0.54 g, 8 eq), and NaHMDS (11.5 mL, 1 M in THF, 11.5 mmol). After obtaining Intermediate 5 using Intermediate 4 (0.26 g, 0.69 mmol) and NaBH4 (0.13 g, 5 eq), it was used immediately in the next reaction without purification. 0.105 g of Compound 4 was obtained using the Intermediate 5 and DDQ (0.13 g, 0.8 eq).

[0132] 1 H NMR (400 MHz, CDCl3) δ 7.37 (dd, J = 15.0, 8.1 Hz, 1H), 6.86 (dt, J = 20.7, 9.2 Hz, 2H), 4.77 (d, J = 14.9 Hz, 1H), 4.44 (d, J = 14.9 Hz, 1H), 4.37 (d, J = 5.7 Hz, 1H), 3.08 (s, 3H), 2.75 (dd, J = 14.1, 5.8 Hz, 1H), 2.67 (d, J = 14.0 Hz, 1H), 1.88 (s, 3H) 1313C NMR (101 MHz, CDCl3) δ 168.62 (s), 167.03 (s), 164.21 (d, J = 12.3 Hz), 162.46 (d, J = 11.9 Hz), 161.71 (d, J = 12.3 Hz), 159.99 (d, J = 12.0 Hz), 132.44 (dd, J = 9.7, 5.3 Hz), 118.46 (d, J = 15.1 Hz), 112.38 (dd, J = 21.3, 3.7 Hz), 104.52 (s), 104.27 (s), 104.01 (s), 69.8, 61.6, 42.7 (d, J = 2.9 Hz), 28.8, 28.6, 21.5。

[0133] Production Example 5: Synthesis of 1,8-Dimethyl-6-(quinolin-2-ylmethyl)-2,3-dithia-6,8-diazabicyclo[3.2.2]nonane-7,9-dione (Compound 5)

[0134]

Chemical Structure

[0135] Step 1. (3S,6S)-3-(t-Butoxymethyl)-1,6-dimethyl-4-(quinolin-2-ylmethyl)piperazine-2,5-dione (Intermediate 2) Using Intermediate 1 (the same substance as Intermediate 4 synthesized in Production Example 1), it was synthesized in the same manner as the method for producing Intermediate 2 in Production Example 2. However, 2-quinolinemethyl chloride was used instead of iodomethane.

[0136] Step 2. 1,8-Dimethyl-6-(quinolin-2-ylmethyl)-2,3-dithia-6,8-diazabicyclo[3.2.2]nonane-7,9-dione (Compound 5) It was prepared by a synthetic method similar to that of Compound 1 in Production Example 1. That is, Intermediate 3 was obtained in a yield of 0.72 g (59%) using Intermediate 2 (1.15 g, 3.11 mmol), sulfur (0.79 g, 8 eq), and NaHMDS (15.6 mL, 1 M in THF, 15.6 mmol). After obtaining Intermediate 4 using Intermediate 3 (0.55 g, 1.44 mmol) and NaBH4 (0.26 g, 5 eq), it was used immediately in the next reaction without purification. Then, 0.25 g of Compound 5 was obtained using Intermediate 4 and DDQ (0.255 g, 0.8 eq).

[0137] 1 H NMR (400 MHz, CDCl3) δ 8.16 (d, J = 8.4 Hz, 1H), 8.02 (dd, J = 8.4, 0.7 Hz, 1H), 7.81 (dd, J = 8.1, 1.3 Hz, 1H), 7.72 (ddd, J = 8.4, 6.9, 1.4 Hz, 1H), 7.55 (ddd, J = 8.1, 6.9, 1.2 Hz, 1H), 7.45 (d, J = 8.4 Hz, 1H), 5.06 (d, J = 14.9 Hz, 1H), 4.71 (d, J = 14.9 H, 1H), 4.61 (dd, J = 6.1, 1.8 Hz, 1H), 3.11 (s, 3H), 2.81 (dd, J = 14.0, 1.8 Hz, 1H), 2.71 (dd, J = 14.0, 6.1 Hz, 1H), 1.91 (s, 3H) 13 C NMR (101 MHz, CDCl3) δ 168.9, 167.1, 155.8, 147.7, 137.6, 130.1, 129.3, 127.8, 127.7, 127.0, 120.7, 69.8, 62.0, 52.2, 28.8, 28.6, 21.6。

[0138] Production Example 6: Synthesis of 1,8-dimethyl-6-(pyridin-2-ylmethyl)-2,3-dithia-6,8-diazabicyclo[3.2.2]nonane-7,9-dione (Compound 6)

[0139] [Chemical formula]

[0140] Step 1. (3S,6S)-3-(t-Butoxymethyl)-1,6-dimethyl-4-(pyridin-2-ylmethyl)piperazine-2,5-dione (Intermediate 2) Using Intermediate 1 (the same substance as Intermediate 4 synthesized in Production Example 1), it was synthesized in the same manner as the method for producing Intermediate 2 in Production Example 2. However, 2-pyridinemethyl chloride was used instead of benzyl bromide.

[0141] Step 2. 1,8-Dimethyl-6-(pyridin-2-ylmethyl)-2,3-dithia-6,8-diazabicyclo[3.2.2]nonane-7,9-dione (Compound 6) It was produced by a synthetic method similar to that of Compound 1 in Production Example 1. That is, using Intermediate 2 (1.05 g, 3.29 mmol), sulfur (0.84 g, 8 eq), and NaHMDS (16.4 mL, 1 M in THF, 16.4 mmol), 0.68 g of Intermediate 3 was obtained. After obtaining Intermediate 4 using Intermediate 3 (0.58 g, 1.70 mmol) and NaBH4 (0.32 g, 5 eq), it was used immediately in the next reaction without purification. Then, using Intermediate 4 and DDQ (0.34 g, 0.8 eq), 0.23 g of Compound 6 was obtained.

[0142] 11H NMR (400 MHz, CDCl3) δ 8.52 (ddd, J = 4.9, 1.7, 0.9 Hz, 1H), 7.67 (td, J = 7.7, 1.8 Hz, 1H), 7.33 (d, J = 7.8 Hz, 1H), 7.22 (ddd, J = 7.5, 4.9, 1.1 Hz, 1H), 4.92 (d, J = 14.9 Hz, 1H), 4.56 (dd, J = 6.0, 1.9 Hz, 1H), 4.47 (d, J = 14.9 Hz, 1H), 3.09 (s, 3H), 2.81 (dd, J = 14.0, 1.9 Hz, 1H), 2.72 (dd, J = 14.0, 6.0 Hz, 1H), 1.87 (s, 3H) 13 13C NMR (101 MHz, CDCl3) δ 168.88, 166.95, 155.48, 149.68, 137.33, 123.24, 123.22, 69.81, 61.94, 51.56, 28.70, 28.56, 21.52。

[0143] Production Example 7: Synthesis of 11-Benzyltetrahydro-5H,7H-4,9a-(epiminomethano)pyrrolo[2,1-c][1,2,4]dithiazepine-5,10-dione (Compound 7)

[0144]

Chemical Structure

[0145] Step 1. Methyl N-((benzyloxycarbonyl)-O-(t-butyl)-L-seryl-L-proline (Intermediate 3) Prepared in the same manner as the synthesis of Intermediate 3 in Production Example 1. However, proline methyl ester was used instead of N-methylalanine methyl ester.

[0146] Step 2. (3S,8aR)-3-(t-butoxymethyl)hexahydropyrrolo[1,2-a]pyrazine-1,4-dione (Intermediate 4) To a mixture of Intermediate 3 (98.0 g, 1.00 eq) in MeOH (980 mL), Pd / C (9.80 g, 10% purity) was added, and the mixture was stirred at 70 °C for 12 h under H2 (50 psi). After filtration, the filtrate was concentrated. The residue was purified by column chromatography to obtain Intermediate 4 (35.0 g, 59.8% yield, 99.0% purity) as a white solid.

[0147] Step 3. (3S,8aR)-2-Benzyl-3-(t-butoxymethyl)hexahydropyrrolo[1,2-a]pyrazine-1,4-dione (Intermediate 5) It was synthesized in the same manner as the method for producing Intermediate 2 of Production Example 2.

[0148] Step 4. 11-Benzyltetrahydro-5H,7H-4,9a-(epiminomethano)pyrrolo[2,1-c][1,2,4]dithiazepine-5,10-dione (Compound 7) It was produced by a synthetic method similar to that of Compound 1 of Production Example 1. That is, using Intermediate 5 (1.5 g, 4.54 mmol), sulfur (1.16 g, 8 eq), and NaHMDS (22.7 mL, 1 M in THF, 22.7 mmol), 0.43 g of Intermediate 6 was obtained. After obtaining Intermediate 7 using Intermediate 6 (0.41 g, 1.16 mmol) and NaBH4 (0.22 g, 5 eq), it was used immediately in the next reaction without purification. Then, using Intermediate 7 and DDQ (0.24 g, 0.8 eq), 0.085 g of Compound 7 was obtained.

[0149] 11H NMR (400 MHz, CDCl3) δ 7.40 - 7.26 (m, 3H), 7.25 (d, J = 8.6 Hz, 2H), 4.64 (d, J = 14.8 Hz, 1H), 4.51 (d, J = 14.8 Hz, 1H), 4.23 (d, J = 6.0 Hz, 1H), 3.87 - 3.79 (m, 1H), 3.73 (dd, J = 19.0, 9.8 Hz, 1H), 2.69 (ddd, J = 13.8, 11.7, 8.0 Hz, 1H), 2.60 (dd, J = 14.1, 6.1 Hz, 1H), 2.52 (d, J = 14.1 Hz, 1H), 2.33 (dd, J = 14.0, 6.7 Hz, 1H), 2.17 (ddd, J = 19.9, 11.7, 4.1 Hz, 2H) 13 13C NMR (101 MHz, CDCl3) δ 167.98, 165.57, 135.31, 129.25, 128.71, 128.48, 72.95, 62.96, 49.29, 45.98, 35.16, 27.94, 21.19。

[0150] Production Example 8: Synthesis of 12 - benzyltetrahydro - 5H,10H - 4,10a - (epiminomethano)[1,4]oxazino[3,4 - c][1,2,4]dithiazepine - 5,11 - dione (Compound 8)

[0151]

Chemical Structure

[0152] Step 1. Methyl - 4 - (N - ((benzyloxy)carbonyl) - O - (t - butyl) - L - seryl)morpholine - 3 - carboxylate (Intermediate 3) It was produced in a similar manner to the synthesis of Intermediate 3 in Production Example 7. However, morpholine - 3 - carboxylic acid, methyl ester was used instead of proline methyl ester.

[0153] Step 2. (7S)-7-(t-Butoxymethyl)hexahydropyridazino[2,1-c][1.4]oxazine-6,9-dione (Intermediate 4) It was produced in a similar manner to the synthesis of Intermediate 4 in Production Example 7.

[0154] Step 3. (7S)-8-Benzyl-7-(t-butoxymethyl)hexahydropyridazino[2,1-c][1.4]oxazine-6,9-dione (Intermediate 5) It was produced in a similar manner to the synthesis of Intermediate 5 in Production Example 7. One of the isomers of the resulting compound (5.20 g, 22.4% yield, 99.3% purity) was obtained as a white solid, and the other isomer (5.10 g, 21.8% yield, 98.6% purity) was obtained as a yellow oil.

[0155] HPLC: 99.3% purity (Isomer 1) & 98.6% purity (Isomer 2) H NMR (Isomer 1): 400 MHz, CDCl3 δ 7.19 - 7.44 (m, 5 H), 5.33 (d, J = 15.2 Hz, 1 H), 4.50 (dd, J = 13.6, 2.0 Hz, 1 H), 4.25 - 4.37 (m, 2 H), 3.87 - 4.00 (m, 3 H), 3.71 - 3.84 (m, 2 H), 3.61 - 3.69 (m, 1 H), 3.47 (td, J = 11.6, 2.4 Hz, 1 H), 2.88 (td, J = 12.8, 3.6 Hz, 1 H), 1.17 ppm (s, 9 H). H NMR (Isomer 2): 400 MHz, CDCl3 δ 7.17 - 7.43 (m, 5 H), 5.21 (d, J = 15.2 Hz, 1 H), 4.48 (dd, J = 12.0, 4.2 Hz, 1 H), 4.20 - 4.32 (m, 2 H), 3.83 - 4.08 (m, 3 H), 3.71 (dd, J = 9.6, 1.6 Hz, 1 H), 3.60 (dd, J = 9.2, 2.4 Hz, 1 H), 3.31 - 3.55 (m, 2 H), 2.87 (td, J = 12.8, 4.0 Hz, 1 H), 1.11 ppm (s, 9 H).

[0156] Step 4. 12-Benzyltetrahydro-5H,10H-4,10a-(epiminomethano)[1,4]oxazino[3,4-c][1,2,4]dithiazepine-5,11-dione (Compound 8) It was produced by the same synthetic method as Compound 1 of Production Example 1. That is, Intermediate 6 was obtained in 1.24 g (58.3% yield) using Intermediate 5 (any isomer, 2.0 g, 5.77 mmol), sulfur (1.48 g, 8 eq), and NaHMDS (28.9 mL, 1 M in THF, 28.9 mmol). After obtaining Intermediate 7 using Intermediate 6 (1.2 g, 3.26 mmol) and NaBH4 (0.62 g, 5 eq), it was used immediately in the next reaction without purification. Then, Compound 7 was obtained in 0.45 g (41% yield) using Intermediate 7 and DDQ (0.67 g, 2.93 mmol, 0.8 eq).

[0157] 1 H NMR (400 MHz, CDCl3) δ 7.34 (t, J = 7.9 Hz, 3H), 7.28 - 7.21 (m, 2H), 4.71 (d, J = 14.8 Hz, 1H), 4.46 (d, J = 14.8 Hz, 1H), 4.26 (d, J = 3.5 Hz, 1H), 4.17 (d, J = 13.7 Hz, 1H), 4.13 - 4.05 (m, 2H), 3.96 (d, J = 13.5 Hz, 1H), 3.59 (td, J = 12.3, 2.5 Hz, 1H), 3.21 (td, J = 13.1, 4.0 Hz, 1H), 2.70 - 2.58 (m, 2H) 13 C NMR (101 MHz, CDCl3) δ 169.23, 165.90, 134.80, 129.33, 128.74, 128.74, 68.82, 65.81, 65.45, 60.51, 49.19, 38.20, 29.42 LCMS: RT = 0.342 min, m / z = 337.0 [M+H] + .

[0158] Production Example 9: Synthesis of 12-(pyridin-4-ylmethyl)tetrahydro-5H,10H-4,10a-(epiminomethano)[1,4]oxazino[3,4-c][1,2,4]dithiazepine-5,11-dione (Compound 9)

[0159] [Chemical formula]

[0160] Step 1. (7S)-7-(tert-butoxymethyl)-8-(pyridin-4-ylmethyl)hexahydropyrazino[2,1-c][1,4]oxazine-6,9-dione (Intermediate 2) Using Intermediate 1 (the same substance as Intermediate 4 synthesized in Production Example 8), it was synthesized in the same manner as the method for producing Intermediate 5 in Production Example 8. However, 4-pyridylmethyl chloride was used instead of benzyl bromide to obtain a brown oil.

[0161] Step 2. 12-(pyridin-4-ylmethyl)tetrahydro-5H,10H-4,10a-(epiminomethano)[1,4]oxazino[3,4-c][1,2,4]dithiazepine-5,11-dione (Compound 9) It was produced by a synthetic method similar to that of Compound 1 in Production Example 1. That is, using Intermediate 2 (0.2 g, 0.58 mmol), sulfur (0.14 g, 8 eq), and NaHMDS (1.14 mL, 2 M in THF, 5 eq), 0.68 g of a trisulfide intermediate was obtained as a yellow solid. After reducing it using the intermediate (0.68 g, 1.84 mmol) and NaBH4 (0.2 g, 2.9 eq), it was used immediately in the next reaction without purification. Then, using the reduced intermediate and iodine (0.47 g, 1.0 eq), 41 mg of the target compound was obtained as a white solid.

[0162] 11H NMR (400 MHz, CDCl3) δ 2.66 - 2.73 (m, 2 H) 3.17 (td, J = 13.20, 4.38 Hz, 1 H) 3.55 (td, J = 12.26, 3.00 Hz, 1 H) 3.89 (d, J = 13.63 Hz, 1 H) 4.00 - 4.13 (m, 3 H) 4.16 - 4.37 (m, 2 H) 4.75 (d, J = 15.63 Hz, 1 H) 7.11 (d, J = 5.75 Hz, 2 H) 8.43 - 8.68 (m, 2 H) LCMS: RT = 0.342 min, m / z = 337.9 [M+H] + 。

[0163] Production Example 10: Synthesis of 12 - ethyltetrahydro - 5H,10H - 4,10a-(epiminomethano)[1,4]oxazino[3,4 - c][1,2,4]dithiazepine - 5,11 - dione (Compound 10)

[0164]

Chemical Structure

[0165] Step 1. (7S)-7-(tert - butoxymethyl)-8 - ethylhexahydropyrazino[2,1 - c][1,4]oxazine - 6,9 - dione (Intermediate 2) Using Intermediate 1 (the same substance as Intermediate 4 synthesized in Production Example 8), it was synthesized in the same manner as the method for producing Intermediate 5 in Production Example 8. However, ethyl bromide was used instead of benzyl bromide.

[0166] Step 2. 12 - ethyltetrahydro - 5H,10H - 4,10a-(epiminomethano)[1,4]oxazino[3,4 - c][1,2,4]dithiazepine - 5,11 - dione (Compound 10) It was prepared by a synthetic method similar to that of Compound 1 in Production Example 1. That is, 0.35 g of a trisulfide intermediate was obtained using Intermediate 2 (0.57 g, 2.0 mmol), sulfur (0.51 g, 8 eq), and NaHMDS (5.0 mL, 2 M in THF, 5 eq). After reduction using the intermediate (0.35 g, 1.2 mmol) and NaBH4 (0.23 g, 5 eq), it was used immediately in the next reaction without purification. Thereafter, 130 mg of the target compound was obtained as a white solid using the reduced intermediate and DDQ (0.24 g, 0.9 eq).

[0167] 1 H NMR (400 MHz, DMSO- d6 ) δ 1.07 (t, 3H), 2.95 (m, 1H), 3.00 (t, 2H), 3.13 (m, 1H), 3.55 (m, 2H), 3.85 (d, 2H), 3.98 (m, 2H), 4.60 (dd, 1H)).

[0168] Production Example 11: Synthesis of 11-(pyridin-4-ylmethyl)tetrahydro-5H,7H-4,9a-(epiminomethano)pyrrolo[2,1-c][1,2,4]dithiazepine-5,10-dione (Compound 11)

[0169]

Chemical formula

[0170] Step 1. (7S)-7-(tert-butoxymethyl)-8-ethylhexahydropyridazino[2,1-c][1,4]oxazine-6,9-dione (Intermediate 2) Using Intermediate 1 (the same substance as Intermediate 4 synthesized in Production Example 7), it was synthesized in the same manner as the method for producing Intermediate 5 in Production Example 7. However, 4-pyridylmethyl chloride was used instead of benzyl bromide.

[0171] Step 2. 11-(pyridin-4-ylmethyl)tetrahydro-5H,7H-4,9a-(epiminomethano)pyrrolo[2,1-c][1,2,4]dithiazepine-5,10-dione (Compound 11) It was prepared by a synthetic method similar to that of Compound 1 in Production Example 1. That is, 0.6 g of a trisulfide intermediate was obtained as a yellow solid using Intermediate 2 (0.2 g, 0.60 mmol), sulfur (0.15 g, 8 eq), and NaHMDS (1.2 mL, 2 M in THF, 5 eq). After reduction using the intermediate (1.0 g, 2.83 mmol) and NaBH4 (0.37 g, 3.5 eq), it was used directly in the next reaction without purification. Thereafter, 40 mg of the target compound was obtained as a white solid using the reduced intermediate and iodine (0.72 g, 1.0 eq).

[0172] 1 H NMR (400 MHz, CDCl3) δ 2.09-2.28 (m, 2 H) 2.35 (ddd, J = 14.04, 6.60, 1.38 Hz, 1 H) 2.56-2.83 (m, 3 H) 3.66-3.98 (m, 2 H) 4.22 (dd, J = 5.44, 2.06 Hz, 1 H) 4.41 (d, J = 15.64 Hz, 1 H) 4.75 (d, J = 15.64 Hz, 1 H) 7.19 (d, J = 5.64 Hz, 2 H) 8.61 (br d, J = 5.40 Hz, 2 H) LCMS: RT=0.343 min, m / z = 321.9 [M+H] + 。

[0173] Production Example 12: Synthesis of 1,8-dimethyl-6-((6-methylpyridin-2-yl)methyl)-2,3-dithia-6,8-diazabicyclo[3.2.2]nonane-7,9-dione (Compound 12)

[0174]

Chemical formula

[0175] Step 1. (3S,6S)-3-(tert-butoxymethyl)-1,6-dimethyl-4-((6-methylpyridin-2-yl)methyl)piperazine-2,5-dione (Intermediate 2) Using Intermediate 1 (the same substance as Intermediate 4 synthesized in Production Example 1), it was synthesized in the same manner as the method for producing Intermediate 2 of Production Example 5. However, 6-methyl-2-pyridylmethyl chloride was used instead of quinoline-2-methyl chloride.

[0176] Step 2. 1,8-Dimethyl-6-((6-methylpyridin-2-yl)methyl)-2,3-dithia-6,8-diazabicyclo[3.2.2]nonane-7,9-dione (Compound 12) It was produced by a synthetic method similar to that of Compound 1 of Production Example 1. That is, using Intermediate 2 (1.0 g, 3 mmol), sulfur (0.77 g, 8 eq), and NaHMDS (7.5 mL, 2 M in THF, 5 eq), 0.47 g of the trisulfide intermediate was obtained as a yellow oil. After reducing using the intermediate (0.47 g, 1.32 mmol) and NaBH4 (0.15 g, 3.5 eq), it was used immediately in the next reaction without purification. Then, using the reduced intermediate and iodine (0.67 g, 2.0 eq), 140 mg of the target compound was obtained as a pale yellow solid.

[0177] 1 H NMR (400 MHz, CDCl3) δ 7.56 (t, J = 7.60 Hz, 1 H), 7.10 (dd, J = 16.0, 7.60 Hz, 2 H), 4.90 (d, J = 14.8 Hz, 1 H), 4.58 (dd, J = 6.00, 1.60 Hz, 1 H), 4.43 (d, J = 14.8 Hz, 1 H), 3.10 (s, 3 H), 2.82 - 2.90 (m, 1 H), 2.68 - 2.77 (m, 1 H), 2.52 (s, 3 H), 1.89 ppm (s, 3 H). LCMS: RT = 0.376 min, m / z = 324.0 [M+H] + 。

[0178] Production Example 13: Synthesis of 1,8-dimethyl-6-((1-methyl-1H-pyrazol-4-yl)methyl)-2,3-dithia-6,8-diazabicyclo[3.2.2]nonane-7,9-dione (Compound 13)

[0179]

Chem.

[0180] Step 1. (3S,6S)-3-(tert-Butoxymethyl)-1,6-dimethyl-4-((1-methyl-1H-pyrazol-4-yl)methyl)piperazine-2,5-dione (Intermediate 2) Using Intermediate 1 (the same substance as Intermediate 4 synthesized in Production Example 1), it was synthesized in the same manner as the method for producing Intermediate 2 of Production Example 5. However, 1-methyl-4-pyrazolylmethyl chloride was used instead of quinoline-2-methyl chloride.

[0181] Step 2. 1,8-Dimethyl-6-((1-methyl-1H-pyrazol-4-yl)methyl)-2,3-dithia-6,8-diazabicyclo[3.2.2]nonane-7,9-dione (Compound 13) It was produced by a synthetic method similar to that of Compound 1 of Production Example 1. That is, using Intermediate 2 (1.0 g, 3.1 mmol), sulfur (0.80 g, 8 eq), and NaHMDS (7.75 mL, 2 M in THF, 5 eq), 0.5 g of a trisulfide intermediate was obtained as a yellow oil. After reducing using the intermediate (0.5 g, 1.45 mmol) and NaBH4 (0.16 g, 3 eq), it was used immediately in the next reaction without purification. Then, using the reduced intermediate and iodine (1.1 g, 3.0 eq), 201 mg of the target compound was obtained as a pale white solid.

[0182] 1 H NMR (400 MHz, CDCl3) δ 7.35 - 7.46 (m, 2 H), 4.54 - 4.66 (m, 1 H), 4.29 - 4.44 (m, 2 H), 3.90 (d, J = 1.60 Hz, 3 H), 3.10 (d, J = 1.60 Hz, 3 H), 2.63 - 2.76 (m, 2 H), 1.89 ppm (d, J = 1.60 Hz, 3 H). LCMS: RT = 0.409 min, m / z = 313.0 [M+H] + 。

[0183] Production Example 14: Synthesis of 1,8-dimethyl-6-(thiophen-2-ylmethyl)-2,3-dithia-6,8-diazabicyclo[3.2.2]nonane-7,9-dione (Compound 14)

[0184]

Chem.

[0185] Step 1. (3S,6S)-3-(tert-butoxymethyl)-1,6-dimethyl-4-(thiophen-2-ylmethyl)piperazine-2,5-dione (Intermediate 2) Using Intermediate 1 (the same substance as Intermediate 4 synthesized in Production Example 1), it was synthesized in the same manner as the method for producing Intermediate 2 in Production Example 5. However, 2-thiophenemethyl chloride was used instead of quinoline-2-methyl chloride.

[0186] Step 2. 1,8-dimethyl-6-(thiophen-2-ylmethyl)-2,3-dithia-6,8-diazabicyclo[3.2.2]nonane-7,9-dione (Compound 14) It was produced by a synthetic method similar to that of Compound 1 in Production Example 1. That is, using Intermediate 2 (1.0 g, 3.1 mmol), sulfur (0.79 g, 8 eq), and NaHMDS (7.7 mL, 2 M in THF, 5 eq), 0.5 g of the trisulfide intermediate was obtained as a yellow oil. After reducing using the intermediate (0.5 g, 1.45 mmol) and NaBH4 (0.16 g, 3 eq), it was used immediately in the next reaction without purification. Then, using the reduced intermediate and iodine (1.1 g, 3.0 eq), 201 mg of the target compound was obtained as a pale white solid.

[0187] 11H NMR (400 MHz, CDCl3) δ 7.29 (d, J = 5.20 Hz, 1H), 7.01 (d, J = 3.40 Hz, 1 H), 6.94 - 6.99 (m, 1 H), 4.79 - 4.89 (m, 1 H), 4.67 - 4.77 (m, 1 H), 4.41 (d, J = 6.00 Hz, 1 H), 3.09 (s, 3 H), 2.64 - 2.72 (m, 1 H), 2.52 - 2.60 (m, 1 H), 1.90 ppm (s, 3 H) LCMS: RT = 0.496 min, m / z = 315.1 [M+H] + 。

[0188] Production Example 15: Synthesis of 6-(benzo[d]thiazol-2-ylmethyl)-1,8-dimethyl-2,3-dithia-6,8-diazabicyclo[3.2.2]nonane-7,9-dione (Compound 15)

[0189]

Chemical Structure

[0190] Step 1. (3S,6S)-1-(benzo[d]thiazol-2-ylmethyl)-6-(tert-butoxymethyl)-3,4-dimethylpiperazine-2,5-dione (Intermediate 2) Using Intermediate 1 (the same substance as Intermediate 4 synthesized in Production Example 1), it was synthesized in the same manner as the method for producing Intermediate 2 in Production Example 5. However, 2-benzothiazolemethyl chloride was used instead of quinoline-2-methyl chloride.

[0191] Step 2. 6-(benzo[d]thiazol-2-ylmethyl)-1,8-dimethyl-2,3-dithia-6,8-diazabicyclo[3.2.2]nonane-7,9-dione (Compound 15) It was produced by a synthetic method similar to Compound 1 of Production Example 1. That is, 0.47 g of a trisulfide intermediate was obtained as a brown oil using Intermediate 2 (0.5 g, 1.33 mmol), sulfur (0.34 g, 8 eq), and NaHMDS (3.3 mL, 2 M in THF, 5 eq). After reduction using the intermediate (0.47 g, 1.18 mmol) and NaBH4 (0.13 g, 3 eq), it was used immediately in the next reaction without purification. Thereafter, 152 mg of the target compound was obtained as a yellow solid using the reduced intermediate and iodine (0.6 g, 2.0 eq).

[0192] 1 H NMR (400 MHz, CDCl3) δ 8.00 (d, J = 8.00 Hz, 1 H), 7.88 (d, J = 8.00 Hz, 1 H), 7.46 - 7.55 (m, 1 H), 7.38 - 7.46 (m, 1 H), 5.15 (d, J = 15.6 Hz, 1 H), 4.85 (d, J = 15.6 Hz, 1 H), 4.62 (dd, J = 5.60, 2.40 Hz, 1 H), 3.12 (s, 3 H), 2.71 - 2.87 (m, 2 H), 1.92 ppm (s, 3 H). LCMS: RT = 0.409 min, m / z = 313.0 [M+H] + 。

[0193] Production Example 16: Synthesis of 1,6 - dimethyl - 8 - (pyrimidin - 2 - ylmethyl) - 2,3 - dithia - 6,8 - diazabicyclo[3.2.2]nonane - 7,9 - dione (Compound 16)

[0194]

Chemical Structure

[0195] Step 1. Methyl O-(tert - butyl)-N-(2 - chloropropanoyl)-L - serinate (Intermediate 1) To a mixture of t-butoxycarbonylmethyl ester (39.5 g, 186 mmol, 1.00 eq, HCl) and TEA (37.8 g, 373 mmol, 51.9 mL, 2.00 eq) in DCM (400 mL) was added 2-chloropropanoyl chloride (26.1 g, 205 mmol). The reaction was stirred at 0 °C for 0.5 h and at 25 °C for 16 h. The reaction mixture was diluted with water (100 mL) and extracted with DCM (50 mL). The combined organic layers were washed with brine (100 mL), dried over Na2SO4, filtered, and concentrated under reduced pressure to give Intermediate 1 (48.6 g, 183 mmol, 97.9% yield) as a yellow liquid. It was used in the next step without further purification.

[0196] Step 2. 3-(tert-Butoxymethyl)-6-methyl-1-(pyrimidin-2-ylmethyl)piperazine-2,5-dione (Intermediate 2) To a mixture of Intermediate 1 (10.8 g, 40.6 mmol, 1.00 eq), K2CO3 (11.2 g, 81.3 mmol, 2.00 eq) and KI (3.37 g, 20.3 mmol, 0.50 eq) in DMF (120 mL) was added pyrimidin-2-ylmethanamine (4.44 g, 40.64 mmol, 1 eq), and the mixture was stirred for 12 h. The reaction solution was filtered and the filtrate was concentrated. The product was purified by prep-HPLC, dissolved in toluene, and stirred at 130 °C for 36 h with DMAP (433 mg, 0.2 eq). The reaction solution was filtered and the filtrate was concentrated. The product was purified by prep-HPLC to give Intermediate 2 (6.04 g, 17.7 mmol, 43.5% yield, 99.0% purity) as a yellow oil.

[0197] Step 3. 3-(tert-Butoxymethyl)-4,6-dimethyl-1-(pyrimidin-2-ylmethyl)piperazine-2,5-dione (Intermediate 3) Intermediate 2 (1.80 g, 5.88 mmol, 1.00 eq) was dissolved in DMF (18 mL). NaH (258 mg, 6.46 mmol, 60% purity, 1.10 eq) was added to the mixture. The mixture was stirred at 0 °C for 0.5 h. Subsequently, MeI (834 mg, 5.88 mmol, 365 μL, 1.00 eq) was added to the reaction mixture at 0 °C. The reaction was stirred at 25 °C for 2.5 h. MeOH (3 mL) was added to quench the reaction mixture, which was then concentrated under reduced pressure. The product was purified by prep-HPLC to obtain Intermediate 3 (1.6 g, 4.99 mmol, 85.0% yield) as a brown oil.

[0198] Step 4. 1,6-Dimethyl-8-(pyrimidin-2-ylmethyl)-2,3-dithia-6,8-diazabicyclo[3.2.2]nonane-7,9-dione (Compound 16) It was produced by a synthetic method similar to that of Compound 1 in Production Example 1. That is, 0.64 g of a trisulfide intermediate was obtained as a white solid using Intermediate 3 (1.2 g, 3.75 mmol), sulfur (0.85 g, 8 eq), and NaHMDS (14.2 mL, 2 M in THF, 5 eq). After reduction using the intermediate (0.25 g, 0.78 mmol) and NaBH4 (0.11 g, 4 eq), it was used immediately in the next reaction without purification. Thereafter, 110 mg of the target compound was obtained as a pale white solid using the reduced intermediate and iodine (0.28 g, 1.5 eq).

[0199] 1 H NMR (400 MHz, CDCl3) δ 8.70 (d, J = 4.88 Hz, 1 H) 7.20 (t, J = 4.88 Hz, 1 H) 5.65 (d, J = 17.26 Hz, 1 H) 4.38 - 4.48 (m, 2 H) 3.05 (s, 3 H) 2.81 - 2.94 (m, 2 H) 1.70 (s, 3 H) LCMS: RT = 0.384 min, m / z = 311.0 [M+H] + 。

[0200] Production Example 17: Synthesis of 1,6-Dimethyl-8-((1-methyl-1H-imidazol-4-yl)methyl)-2,3-dithia-6,8-diazabicyclo[3.2.2]nonane-7,9-dione (Compound 17)

[0201] [Chemical Formula]

[0202] Step 1. 3-(tert-Butoxymethyl)-6-methyl-1-(pyrimidin-2-ylmethyl)piperazine-2,5-dione (Intermediate 2) Synthesized in the same manner as the method for producing Intermediate 2 of Production Example 16. However, 1-methylimidazolemethanamine was used instead of pyrimidin-2-ylmethanamine.

[0203] Step 2. 3-(tert-Butoxymethyl)-4,6-dimethyl-1-((1-methyl-1H-imidazol-4-yl)methyl)piperazine-2,5-dione (Intermediate 3) Synthesized in the same manner as the method for producing Intermediate 3 of Production Example 16.

[0204] Step 3. 1,6-Dimethyl-8-((1-methyl-1H-imidazol-4-yl)methyl)-2,3-dithia-6,8-diazabicyclo[3.2.2]nonane-7,9-dione (Compound 17) Produced by a synthetic method similar to that of Compound 1 in Production Example 1. That is, using Intermediate 3 (0.5 g, 1.55 mmol), sulfur (0.4 g, 8 eq), and NaHMDS (5.9 mL, 2 M in THF, 5 eq), 0.36 g of the trisulfide intermediate was obtained as a yellow oil. After reduction using the intermediate (0.25 g, 0.73 mmol) and NaBH4 (0.13 g, 5 eq), it was used directly in the next reaction without purification. Then, using the reduced intermediate and iodine (0.18 g, 1 eq), 106 mg of the target compound was obtained as a pale white solid.

[0205] 11H NMR (400 MHz, CDCl3) δ 7.41 (s, 1 H) 6.96 (s, 1 H) 4.92 (d, J = 15.51 Hz, 1 H) 4.55 (d, J = 15.63 Hz, 1 H) 4.31 (t, J = 4.00 Hz, 1 H) 3.67 (s, 3 H) 3.01 (s, 3 H) 2.84 - 2.88 (m, 2 H) 2.04 (s, 3 H) LCMS: RT = 0.417 min, m / z = 313.0 [M+H] + 。

[0206] Production Example 18: Synthesis of Methyl 4 - ((1,6 - dimethyl - 7,9 - dioxo - 2,3 - dithia - 6,8 - diazabicyclo[3.2.2]nonan - 8 - yl)methyl)benzoate (Compound 18)

[0207]

Chemical Structure

[0208] Step 1. Methyl 4 - ((3 - (tert - butoxymethyl) - 6 - methyl - 2,5 - dioxopiperazin - 1 - yl)methyl)benzoate (Intermediate 2) It was synthesized in the same manner as the method for producing Intermediate 2 of Production Example 16. However, 4 - methoxyphenylmethanamine was used instead of pyrimidin - 2 - ylmethanamine.

[0209] Step 2. Methyl 4 - ((3 - (tert - butoxymethyl) - 4,6 - dimethyl - 2,5 - dioxopiperazin - 1 - yl)methyl)benzoate (Intermediate 3) It was synthesized in the same manner as the method for producing Intermediate 3 of Production Example 16.

[0210] Step 3. Methyl 4 - ((1,6 - dimethyl - 7,9 - dioxo - 2,3 - dithia - 6,8 - diazabicyclo[3.2.2]nonan - 8 - yl)methyl)benzoate (Compound 18) It was produced by a synthetic method similar to that of Compound 1 in Production Example 1. That is, 1.0 g of a trisulfide intermediate was obtained as a yellow oil using Intermediate 3 (1.57 g, 4.16 mmol), sulfur (1.1 g, 8 eq), and NaHMDS (15.7 mL, 1 M in THF, 5 eq). After reduction using the intermediate (0.13 g, 0.32 mmol) and NaBH4 (0.015 g, 1.2 eq), it was used immediately in the next reaction without purification. Thereafter, 113 mg of the target compound was obtained as a pale yellow solid using the reduced intermediate and iodine (0.08 mg, 1 eq).

[0211] 1 H NMR (400 MHz, CDCl3) δ 7.99 - 8.01 (m, 2 H) 7.37 (d, J = 8.25 Hz, 2 H) 5.44 (d, J = 16.26 Hz, 1 H) 4.43 (dd, J = 6.38, 0.88 Hz, 1 H) 4.29 - 4.33 (m, 2 H) 3.92 (s, 3 H) 3.07 (s, 3 H) 2.85 (dd, J = 14.88, 6.38 Hz, 1 H) 1.70 (s, 3 H) LCMS: RT = 0.503 min, m / z = 367.1 [M+H] + 。

[0212] Production Example 19: Synthesis of 1-benzyl-6,8-dimethyl-2,3-dithia-6,8-diazabicyclo[3.2.2]nonane-7,9-dione (Compound 19)

[0213]

Chemical formula

[0214] Step 1. 3-benzyl-6-(tert-butoxymethyl)-1,4-dimethylpiperazine-2,5-dione (Intermediate 2) To a solution of Compound 1 (15.0 g, 33.6 mmol, 1.00 eq) in DMF (100 mL) was added tBuOK (1 M, 134 mL, 4.00 eq) and MeI (29.2 g, 205 mmol, 12.8 mL, 6.12 eq). The mixture was stirred at 25 °C for 2 h. 300 mL of water was added at 0 °C to quench the reaction mixture, and the mixture was extracted with ethyl acetate (100 mL × 2). The combined organic layers were washed with aqueous NaCl (50.0 mL × 6), dried over Na2SO4, filtered, and concentrated under reduced pressure to obtain a residue. The residue was purified by reverse-phase HPLC (column: Kromasil Eternity XT 250×80 mm×10 μm; mobile phase: [water (NH4HCO3)-ACN]; B%: 29%-59%, 20 min) to obtain Intermediate 2 (3.20 g, 9.86 mmol, 14.7% yield, 98.1% purity) as a white solid.

[0215] Step 2. 1-Benzyl-6,8-dimethyl-2,3-dithia-6,8-diazabicyclo[3.2.2]nonane-7,9-dione (Compound 19) It was produced by the same synthetic method as Compound 1 in Production Example 1. That is, 0.5 g of a trisulfide intermediate was obtained as a yellow oil using Intermediate 2 (1.0 g, 3.14 mmol), sulfur (0.9 g, 8 eq), and NaHMDS (15.6 mL, 1 M in THF, 5 eq). After reduction using the intermediate (0.5 g, 1.47 mmol) and NaBH4 (0.3 g, 5 eq), it was used immediately in the next reaction without purification. Thereafter, 330 mg of the target compound was obtained as a white solid using the reduced intermediate and iodine (0.6 g, 1 eq).

[0216] 11H NMR (400 MHz, CDCl3) δ 2.85 (d, J = 5.60 Hz, 1 H) 2.88 - 2.96 (m, 1 H) 3.02 (s, 3 H) 3.10 (s, 3 H) 3.24 (d, J = 16.4 Hz, 1 H) 4.16 (d, J = 16.4 Hz, 1 H) 4.37 (dd, J = 5.6, 2.00 Hz, 1 H) 7.14 (d, J = 7.2 Hz, 2 H) 7.19 - 7.25 (m, 1 H) 7.27 - 7.32 (m, 2 H). LCMS: RT = 0.498 min, m / z = 309.0 [M+H] + 。

[0217] Production Example 20: Synthesis of 6,8 - diethyl - 1 - methyl - 2,3 - dithia - 6,8 - diazabicyclo[3.2.2]nonane - 7,9 - dione (Compound 20)

[0218]

Chemical Structure

[0219] Step 1. 3 - (tert - butoxymethyl) - 1,4 - diethyl - 6 - methylpiperazine - 2,5 - dione (Intermediate 2) It was produced in a similar manner to the synthesis of Intermediate 2 in Production Example 19 using Starting Material 1. However, iodoethane was used instead of iodomethane.

[0220] Step 2. 6,8 - diethyl - 1 - methyl - 2,3 - dithia - 6,8 - diazabicyclo[3.2.2]nonane - 7,9 - dione (Compound 20) It was produced by the same synthetic method as Compound 1 of Production Example 1. That is, 0.55 g of a trisulfide intermediate was obtained as a yellow oil using Intermediate 2 (0.65 g, 2.4 mmol), sulfur (0.8 g, 10 eq), and NaHMDS (13.2 mL, 1 M in THF, 5 eq). After reduction using the intermediate (0.55 g, 1.47 mmol) and NaBH4 (0.47 g, 6.6 eq), it was used immediately in the next reaction without purification. Thereafter, 216 mg of the target compound was obtained as a white solid using the reduced intermediate and iodine (0.9 g, 2 eq).

[0221] 1 H NMR (400 MHz, CDCl3) δ 4.40 (t, J = 3.80 Hz, 1 H), 3.68 - 3.83 (m, 2 H), 3.58 - 3.67 (m, 1 H), 3.20 (dd, J = 14.0, 7.20 Hz, 1 H), 2.80 (d, J = 3.60 Hz, 2 H), 1.80 (s, 3 H), 1.32 (t, J = 7.20 Hz, 3 H), 1.16 ppm (t, J = 7.20 Hz, 3 H). LCMS: RT = 0.451 min, m / z = 261.0 [M+H] + 。

[0222] Production Example 2 Synthesis of 12-benzyl-9-methylhexahydro-5H-4,10a-(epiminomethano)pyrazino[2,1-c][1,2,4]dithiazepine-5,11-dione (Compound 21)

[0223]

Chemical formula

[0224] Step 1. tert-Butyl 8-benzyl-7-(tert-butoxymethyl)-6,9-dioxooctahydro-2H-pyrazino[1,2-a]pyrazine-2-carboxylate (Intermediate 2) To a solution of compound 1 (20.0 g, 56.3 mmol, 1.00 eq) in THF (500 mL) was added NaH (6.75 g, 169 mmol, 60% purity, 3.00 eq) at 0 °C over 30 minutes. After the addition, BnBr (10.6 g, 61.9 mmol, 7.35 mL, 1.10 eq) was added dropwise at 0 °C and the mixture was stirred at 25 °C for 12 hours. 100 mL of H2O was added at 0 °C to quench the reaction mixture, and the mixture was extracted with ethyl acetate (100 mL × 2). The combined organic layers were washed with aqueous NaCl (100 mL × 1), dried over Na2SO4, filtered, and concentrated under reduced pressure to obtain a residue. The residue was purified by column chromatography (SiO2, petroleum ether:ethyl acetate = 2:1 to 1:3) to obtain intermediate 2 (15.0 g, 17.2 mmol, 76.5% purity) as a white solid.

[0225] Step 2. 8-Benzyl-7-(tert-butoxymethyl)-2-methylhexahydro-2H-pyrazino[1,2-a]pyrazine-6,9-dione (Intermediate 3) To a solution of intermediate 2 (10.0 g, 22.4 mmol, 1.00 eq) in EtOAc (70.0 mL) was added HCl / EtOAc (4 M, 30.0 mL, 5.35 eq), and the mixture was stirred at 25 °C for 2 hours. The residue obtained by concentrating the reaction mixture under reduced pressure was used in the next step without further purification (15 g). This compound was dissolved in MeOH (150 mL), then formaldehyde (17.6 g, 217 mmol, 16.2 mL, 37.0% purity, 5.00 eq) was added at 25 °C over 30 minutes, and then NaBH(OAc)3 (13.8 g, 65.1 mmol, 1.50 eq) was added. The resulting mixture was stirred at 25 °C for 1 hour. After concentrating the reaction mixture under reduced pressure to obtain a residue, it was purified by reverse-phase HPLC (column: Kromasil Eternity XT 250×80 mm×10 μm; mobile phase: [water (NH4HCO3)-ACN]; B%: 23%-53%, 20 minutes) to obtain intermediate 3 (5.50 g, 15.2 mmol, 34.9% yield, 100% purity) as a yellow oil. Two isomers (diastereomers) were formed. The two were separated respectively.

[0226] Isomer 1: 11H NMR (400 MHz, DMSO- d6 ) δ ppm 1.05 (s, 9 H) 1.76 - 1.90 (m, 1 H) 2.17 - 2.37 (m, 4 H) 2.65 - 2.80 (m, 2 H) 3.06 - 3.20 (m, 2 H) 3.58 (s, 2 H) 3.94 - 4.15 (m, 2 H) 4.24 - 4.39 (m, 2 H) 4.84 (d, J = 15.2 Hz, 1 H) 7.10 - 7.45 (m, 5 H). Isomer 2: 1 1H NMR (400 MHz, DMSO- d6 ) δ 0.99 - 1.10 (m, 9 H) 1.73 - 1.92 (m, 1 H) 2.04 - 2.33 (m, 4 H) 2.61 - 2.82 (m, 2 H) 3.09 (dd, J = 10.4, 2.69 Hz, 1 H) 3.58 (d, J = 2.00 Hz, 2 H) 3.88 - 4.03 (m, 1 H) 4.09 - 4.17 (m, 1 H) 4.18 - 4.39 (m, 2 H) 4.85 (d, J = 15.2 Hz, 1 H) 7.24 - 7.35 (m, 5 H) MS: m / z = 360.2。

[0227] Step 3. 12-Benzyl-9-methylhexahydro-5H-4,10a-(epiminomethano)pyrazino[2,1-c][1,2,4]dithiazepine-5,11-dione (Compound 21) Prepared by a synthetic method similar to that of Compound 1 in Production Example 1. That is, 0.48 g of a trisulfide intermediate was obtained as a yellow solid using Intermediate 3 (Isomer 1 or 2, 1.2 g, 3.3 mmol), sulfur (0.86 g, 8 eq), and NaHMDS (16.7 mL, 1 M in THF, 5 eq). After reduction using the intermediate (0.78 g, 1.47 mmol) and NaBH4 (0.14 g, 1.9 eq), it was used immediately in the next reaction without purification. Using the reduced intermediate and iodine (0.68 g, 1 eq), 101 mg of the target compound was obtained as a white solid.

[0228] 11H NMR (400 MHz, CDCl3) δ 2.14 (td, J = 12.0, 3.63 Hz, 1 H) 2.43 (s, 3 H) 2.56 - 2.80 (m, 3 H) 2.87 - 3.03 (m, 1 H) 3.16 - 3.37 (m, 2 H) 4.13 - 4.29 (m, 2 H) 4.44 (d, J = 14.8 Hz, 1 H) 4.82 (d, J = 14.8 Hz, 1 H) 7.26 - 7.31 (m, 3 H) 7.35 - 7.39 (m, 2 H). LCMS: RT = 0.520 min, m / z = 350.1 [M+H] + 。

[0229] Production Example 22: Synthesis of 12-Benzyl-9-methylhexahydro-5H-4,10a-(epiminomethano)pyrazino[2,1-c][1,2,4]dithiazepine-5,11-dione (Compound 21)

[0230]

Chemical Structure

[0231] Step 1. tert-Butyl 7-(tert-butoxymethyl)-6,9-dioxo-8-(pyridin-4-ylmethyl)octahydro-2H-pyrazino[1,2-a]pyrazine-2-carboxylate (Intermediate 2) It was produced in a similar manner to the synthesis of Intermediate 2 in Production Example 21. However, 4-pyridylmethyl bromide was used instead of benzyl bromide.

[0232] Step 2. 7-(tert-Butoxymethyl)-2-methyl-8-(pyridin-4-ylmethyl)hexahydro-2H-pyrazino[1,2-a]pyrazine-6,9-dione (Intermediate 3) It was produced in a similar manner to the synthesis of Intermediate 3 in Production Example 21. Two isomers (diastereomers) were formed. The two were separated respectively.

[0233] Isomer 1: 1 1H NMR (400 MHz, DMSO-d6 ) δ 1.05 (d, J = 2.80 Hz, 9 H) 1.75-1.98 (m, 2 H) 2.19-2.37 (m, 3 H) 2.59-2.90 (m, 2 H) 3.03-3.29 (m, 3 H) 3.44-3.66 (m, 2 H) 3.97-4.14 (m, 2 H) 4.17-4.59 (m, 2 H) 4.60-4.95 (m, 1 H) 7.28 (dd, J = 18.4, 5.76 Hz, 2 H) 8.51 (t, J = 6.00Hz, 2 H). Isomer 2: 1 H NMR (400 MHz, DMSO- d6 ) δ 1.04 (d, J = 2.80 Hz, 9 H) 1.75-2.00 (m, 2 H) 2.25 (d, J = 9.20 Hz, 3 H) 2.75 (br s, 2 H) 3.03-3.39 (m, 2 H) 3.47 (br dd, J = 9.20, 2.38 Hz, 1 H) 3.60 (d, J = 2.00 Hz, 1 H) 3.98-4.07 (m, 1 H) 4.09-4.17 (m, 1 H) 4.18-4.57 (m, 2 H) 4.59-4.92 (m, 1 H) 7.28 (dd, J = 18.0, 5.88 Hz, 2 H) 8.37-8.60 (m, 2 H).

[0234] Step 3. 12-Benzyl-9-methylhexahydro-5H-4,10a-(epiminomethano)pyrazino[2,1-c][1,2,4]dithiazepine-5,11-dione (Compound 22) It was prepared by a synthetic method similar to Compound 1 of Production Example 1. That is, 0.40 g of a trisulfide intermediate was obtained as a yellow solid using Intermediate 3 (isomer 1 or 2, 1.2 g, 3.3 mmol), sulfur (0.85 g, 8 eq), and NaHMDS (16.6 mL, 1 M in THF, 5 eq). After reduction using the intermediate (0.55 g, 1.44 mmol) and NaBH4 (0.18 g, 3.2 eq), it was used immediately in the next reaction without purification. Thereafter, 102 mg of the target compound was obtained as a yellow solid using the reduced intermediate and iodine (0.53 g, 1 eq).

[0235] 1 H NMR (400 MHz, CDCl3) δ 2.16 (br d, J = 3.60 Hz, 1 H) 2.43 (s, 3 H) 2.62 (d, J = 13.6 Hz, 1 H) 2.69 - 2.86 (m, 2 H) 2.89 - 3.04 (m, 1 H) 3.16 - 3.35 (m, 2 H) 4.14 - 4.38 (m, 3 H) 4.92 (d, J = 15.6 Hz, 1 H) 7.19 (d, J = 5.60 Hz, 2 H) 8.61 (d, J = 5.60 Hz, 2 H). LCMS: RT = 0.531 min, m / z = 351.3 [M+H] + 。

[0236] Production Example 23: Synthesis of 12-benzyl-9-methylhexahydro-5H-4,10a-(epiminomethano)pyrazino[2,1-c][1,2,4]dithiazepine-5,11-dione (Compound 23)

[0237]

Chemical formula

[0238] Step 1. tert-Butyl 7-(tert-butoxymethyl)-8-methyl-6,9-dioxooctahydro-2H-pyrazino[1,2-a]pyrazine-2-carboxylate (Intermediate 2) It was produced in a similar manner to the synthesis of Intermediate 2 in Production Example 21. However, methyl iodide was used instead of benzyl bromide.

[0239] Step 2. 7-(tert-Butoxymethyl)-2,8-dimethylhexahydro-2H-pyrazino[1,2-a]pyrazine-6,9-dione (Intermediate 3) It was produced in a similar manner to the synthesis of Intermediate 3 in Production Example 21.

[0240] Step 3. 12-Benzyl-9-methylhexahydro-5H-4,10a-(epiminomethano)pyrazino[2,1-c][1,2,4]dithiazepine-5,11-dione (Compound 23) It was produced by a synthetic method similar to that of Compound 1 in Production Example 1. That is, 2.1 g of a trisulfide intermediate was obtained as a white solid using Intermediate 3 (4.0 g, 14.1 mmol), sulfur (3.6 g, 8 eq), and NaHMDS (70.6 mL, 1 M in THF, 5 eq). After reduction using the intermediate (0.50 g, 1.44 mmol) and NaBH4 (0.26 g, 5 eq), it was used immediately in the next reaction without purification. Then, 125 mg of the target compound was obtained as a pale yellow solid using the reduced intermediate and DDQ (0.33 g, 0.9 eq).

[0241] 1 H NMR (400 MHz, CDCl3) δ 4.24 (dd, J = 4.9, 3.1 Hz, 1H), 4.19 (dd, J = 13.4, 2.0 Hz, 1H), 3.30 - 3.22 (m, 2H), 3.03 (s, 3H), 2.97 - 2.87 (m, 3H), 2.53 (d, J = 13.6 Hz, 1H), 2.40 (s, 3H), 2.10 (td, J = 12.2, 3.6 Hz, 1H). 13 C NMR (126 MHz, CDCl3) δ 169.49, 166.15, 68.29, 62.86, 59.28, 52.72, 46.14, 40.01, 33.07, 29.37. MS: m / z = 274.0715 [M+H]+ .

[0242] Production Example 24: Synthesis of 6-Benzyl-1,4,8-trimethyl-2,3-dithia-6,8-diazabicyclo[3.2.2]nonane-7,9-dione (Compound 24)

[0243] [Chemical Formula]

[0244] Step 1. (6S)-3-((R)-1-(tert-Butoxy)ethyl)-1,6-dimethylpiperazine-2,5-dione (Intermediate 1) Prepared in a similar manner to the synthesis of Intermediate 4 in Production Example 7.

[0245] Step 2. (3S)-1-Benzyl-6-((S)-1-(tert-butoxy)ethyl)-3,4-dimethylpiperazine-2,5-dione (Intermediate 2) Prepared by a method similar to the synthesis of Intermediate 2 in Production Example 2.

[0246] Step 3. 6-Benzyl-1,4,8-trimethyl-2,3-dithia-6,8-diazabicyclo[3.2.2]nonane-7,9-dione (Compound 24) Prepared by a synthetic method similar to that of Compound 1 in Production Example 1. That is, using Intermediate 2 (0.9 g, 2.71 mmol), sulfur (0.7 g, 8 eq), and NaHMDS (10.9 mL, 1 M in THF, 5 eq), a trisulfide intermediate was obtained as a yellow solid. After reduction using the intermediate (0.49 g, 1.38 mmol) and NaBH4 (0.16 g, 3 eq), it was used directly in the next reaction without purification. Subsequently, the reduced intermediate and iodine (0.7 g, 2 eq) were used to obtain the target compound. Two isomers (diastereomers) were formed, with isomer 1 obtained as a yellow solid (157 mg) and isomer 2 obtained as a white solid (13 mg).

[0247] Isomer 1: 11H NMR (400 MHz, CDCl3) δ δ 7.22 - 7.33 (m, 3 H), 7.15 - 7.22 (m, 2 H), 4.41 - 4.68 (m, 2 H), 3.88 (s, 1 H), 3.02 (s, 3 H), 2.67 (q, J = 7.20 Hz, 1 H), 1.83 (s, 3 H), 1.11 (d, J = 7.20 Hz, 3 H) Isomer 2: 1 1H NMR (400 MHz, CDCl3) δ 7.28 - 7.38 (m, 3 H), 7.10 - 7.18 (m, 2 H), 5.45 (d, J = 15.2 Hz, 1 H), 4.19 (d, J = 6.00 Hz, 1 H), 3.84 (d, J = 15.2 Hz, 1 H), 3.08 (s, 3 H), 2.96 - 3.04 (m, 1 H), 1.91 (s, 3 H), 1.57 - 1.60 (m, 3 H) LCMS (Isomer 1): RT = 1.925 min, m / z = 323 [M+H] + LCMS (Isomer 2): RT = 1.894 min, m / z = 323 [M+H] + 。

[0248] Production Example 25: Synthesis of 6-benzyl-1,4,8-trimethyl-2,3-dithia-6,8-diazabicyclo[3.2.2]nonane-7,9-dione (Compound 25)

[0249]

Chemical Structure

[0250] Step 1. (6S)-3-((S)-1-(tert-butoxy)ethyl)-1,6-dimethyl-4-(pyridin-2-ylmethyl)piperazine-2,5-dione (Intermediate 2) It was produced in a similar manner to the synthesis of Intermediate 2 in Production Example 2. However, 2-pyridylmethyl bromide was used instead of benzyl bromide.

[0251] 2.6-Benzyl-1,4,8-trimethyl-2,3-dithia-6,8-diazabicyclo[3.2.2]nonane-7,9-dione (Compound 25) It was produced by a synthetic method similar to that of Compound 1 in Production Example 1. That is, 1.67 g of a trisulfide intermediate was obtained as a yellow oil using Intermediate 2 (3.1 g, 9.3 mmol), sulfur (2.4 g, 8 eq), and NaHMDS (37 mL, 1 M in THF, 5 eq). After reduction using the intermediate (1.67 g, 4.7 mmol) and NaBH4 (0.53 g, 3 eq), it was used immediately in the next reaction without purification. Thereafter, 373 mg of the target compound was obtained as a white solid using the reduced intermediate and iodine (1 g, 1 eq).

[0252] 1 H NMR (400 MHz, CDCl3) δ 8.52 (d, J = 4.60 Hz, 1 H) 7.62 (td, J = 7.60, 1.60 Hz, 1 H) 7.12 - 7.23 (m, 2 H) 5.44 (d, J = 15.6 Hz, 1 H) 4.49 (d, J = 15.6 Hz, 1 H) 3.95 - 4.09 (m, 3 H) 2.96 (s, 3 H) 1.68 (d, J = 7.20 Hz, 3 H) 1.33 (d, J = 6.00 Hz, 3 H) 1.21 (s, 9 H) LCMS: RT = 0.684 min, m / z = 323.9 [M+H] + 。

[0253] Experimental Example 1: Materials Rabbit polyclonal antibodies specific for peroxiredoxin type 2 (PrxII or Prdx2) were obtained from AbFrontier (Seoul, Korea). The sequences of the human PrxII-specific siRNAs used in the present invention are as follows.

[0254] 5'-CGCUUGUCUGAGGAUUACGUU-3' (Prx II-1) 5'-AGGAAUAUUUCUCCAAACAUU-3' (Prx II-2). The phospho-tyrosine antibody (4G10) and PDGF-BB were obtained from Upstate. PrxII, Prx-SO 2 / 3 and the phospho-PDGFR-β (pY857) rabbit polyclonal antibody were prepared according to the description in the previous literature (M.H. Choi et al., Nature 2005 May 19;435(7040):347-53). The phospho-VEGFR2 (pY1175), VEGFR2, phospho-PLCγ1 (pY783), PLCγ1, pTpY-Erk, and Erk2 antibodies were obtained from Cell Signaling Technology (CST), the α-tubulin antibody was obtained from Sigma, and VEGF-A was obtained from R&D.

[0255] Experimental Example 2: Cell Culture Human Aortic Endothelial Cells (HAEC), Human Pulmonary Artery Endothelial Cells (PAEC), Human Aortic Smooth Muscle Cells (HASMC) and Human Pulmonary Artery Smooth Muscle Cells (PASMC) were obtained from Clonetics-Bio Whittaker (Belgium). These were seeded on 0.1% gelatin-coated culture plates. Then, endothelial basal medium (Endothelial Basal Medium; EBM TM -2) containing 10% fetal bovine serum (FBS) and full media supplement (Clonetics-Bio Whittaker Cat.no.cc-4176 for HAECs; Clonetics-Bio Whittaker Cat.no.cc-4149 for HASMCs) and smooth muscle cell basal medium (Smooth Muscle Cell Basal Medium; SmBM TM ) were used for culturing respectively. Cell culture was carried out in a humidified incubator at 37°C with 5% carbon dioxide. In the present invention, cells at passages 5 to 7 were used.

[0256] Experimental Example 3: Analysis of Peroxidase Activity Analysis of the peroxidase activity of the compound according to the present invention to reduce hydrogen peroxide (H2O2) was carried out as follows by a known method. The measurement of peroxiredoxin activity was performed in a reaction solution (volume 200 μL) containing 50 mM HEPES buffer (pH 7.0), 1 mM EDTA, 250 μM NADPH, 3 μM yeast thioredoxin (Trx), 1.5 μM yeast Trx reductase (TR), 50 μM test compound, and 1.2 mM hydrogen peroxide (H2O2).

[0257] The enzyme reaction was started by adding H2O2, and the decrease in absorbance was measured at 340 nm using an Agilent UV8453 spectrophotometer (Hewlett-Packard, USA) at 30 °C for 12 minutes. The initial reaction rate was expressed as the amount of NADPH oxidized per minute by calculating the slope of the linear part of the curve.

[0258] Experimental Example 4: Measurement of Intracellular Hydrogen Peroxide Removal Ability The ability of the compound of the present invention to remove intracellular hydrogen peroxide was measured as follows. NIH-3T3 cells (ATCC) were seeded in 35 mm culture dishes at a concentration of 2 × 10 5 cells / well. After culturing for 24 hours, they were starved for 6 hours in phenol red-free basal DMEM containing 0.5% FBS. Starved NIH 3T3 cells were pretreated with the test compound for 30 minutes according to the concentrations (0, 12.5, 25, 50, 100 μM) diluted in phenol red-free basal DMEM, and then treated with 20 mU of glucose oxidase (Gox) for 30 minutes. CM-H2DCF-DA, a reactive oxygen species-specific fluorescent substance, was prepared in a 1 mM DMSO stock, diluted 500-fold in phenol red-free basal DMEM, and treated with the cells pretreated with the test compound for 5 minutes. Then, the residual fluorescent diluent was quickly washed with the basal medium. Images were taken using a fluorescence microscope with an Ex497 nm / Em518 nm setting. Three images at 10× magnification were taken for each concentration sample, and the fluorescence values of more than 30 individual cells per captured image were measured and used as representative values.

[0259] Experimental Example 5: Analysis of Cytotoxicity To analyze cytotoxicity, human aortic endothelial cells (HAEC) and human aortic smooth muscle cells (HASMC) were seeded into 96-well culture plates at a concentration of 4,000 cells / well with a final volume of 100 μL. After culturing the cells for 24 hours to stabilize them, they were further cultured in serum-free medium for 18 hours. The test substances were serially diluted in EBM-2 culture medium and added to the cells at 100 μl / well, followed by culturing for 24 hours. The number of living cells was measured using a WST-1 cell viability assay kit (Roche Diagnostics, USA), and the cell count was expressed as the value obtained by subtracting the turbidity measured at 600 nm from the absorbance measured at 450 nm. The data used the average value of the measured values in three wells.

[0260] Experimental Example 6: Analysis of Immunoblot The cells were quickly washed with ice-cold phosphate buffer saline (PBS) and lysed in an extraction buffer containing 20 mM Hepes (pH 7.0), 1% Triton X-100, 150 mM NaCl, 10% glycerol, 1 mM EDTA, 2 mM EGTA, 1 mM DTT, 5 mM Na3VO4, 5 mM NaF, 1 mM AEBSF, aprotinin (5 μg / ml), and leupeptin (5 μg / ml). After centrifugation at 12,000×g, the purified cell extracts were used for immunoblot analysis. When necessary, the nitrocellulose membrane used in the primary blot was reacted by shaking in a 67 mM Tris (pH 6.7), 2% SDS, 100 mM 2-mercaptoethanol solution at 60 °C for 30 minutes to remove the antibody, and then washed three times with a Tris-buffered saline (TBS) solution containing 1% Triton X-100. Subsequently, additional immunoblots were performed using other desired antibodies.

[0261] Experimental Example 7: Measurement of Redox Potential As shown in Table 1 below, 10 kinds of Redox buffer solutions (0.5 mL) were prepared on the day of the experiment by mixing a 50 mM solution of reduced dithiothreitol (DTT) and its oxidized DTT (trans-1,2-dithiane-4,5-diol) at a certain ratio. A 10 mM test substance was diluted 20-fold with each buffer solution. After reacting the mixture of the reaction solution and the test substance in a constant temperature water bath at 30 °C for 3 hours, it was analyzed by HPLC. The peak areas (%) of the specific retention times of the oxidized test substance and the reduced test substance on HPLC were calculated. After completing the graph with the Nernst equation value (mV) on the X-axis and the ratio (%) of the reduced test substance on the y-axis, the redox potential value corresponding to a ratio of 50 (%) of the reduced test substance was calculated using the graph in the linear region, and this was taken as the median redox potential, which is the representative value of the redox potential of the test substance.

[0262]

Table 1

[0263] In Experimental Examples 1 to 7, other specific methods and conditions were the same as those disclosed in International Publication WO2013-077709 or International Publication WO2018-008984. All the disclosure matters of International Publication WO2013-077709 and International Publication WO2018-008984 are incorporated herein by reference.

[0264] Experimental Example 8: Test for regulating growth factor-induced proliferation of vascular smooth muscle cells and vascular endothelial cells The ability of the test compound to regulate the proliferation of vascular smooth muscle cells and vascular endothelial cells was measured.

[0265] HASMC and PASMC were transfected with PrxII - specific siRNA using the RNAi MAX transfection reagent according to the manufacturer's protocol. After 24 hours, the transfected cells (3,000 cells / well) were seeded into 96 - well culture plates. Twelve hours after seeding, the cells were further serum - starved for 18 hours in SmBM basal medium containing 0.5% FBS. The test compounds were serially diluted in the same basal medium containing 0.5% FBS and added to the cells (100 μL / well) and cultured for 2 hours. After treatment, the cells were placed in fresh SmBM medium containing 0.5% FBS and PDGF - BB (25 ng / mL) for 24 hours for growth factor stimulation and further cultured.

[0266] HAEC and PAEC were transfected with PrxII - specific siRNA using the RNAi MAX transfection reagent according to the manufacturer's protocol. After 24 hours, the transfected cells (3,000 cells / well) were seeded into 96 - well culture plates. Twelve hours after seeding, the cells were further serum - starved for 18 hours in EBM basal medium containing 0.5% FBS. The test compounds were serially diluted in the same basal medium containing 0.5% FBS and added to the cells (100 μL / well) and cultured for 2 hours. After treatment, the cells were placed in fresh EBM medium containing 0.5% FBS and VEGF - A (25 ng / mL) for 24 hours for growth factor stimulation and further cultured.

[0267] The degree of cell proliferation was measured using the Cell Titer - GLO kit (Roche Diagnostics, USA). The data were presented as the percentage of the luminescence intensity averaged from 3 replicate wells and the percentage of the untreated control group.

[0268] Experimental Example 9: Efficacy Evaluation Test in a Pre - clinical Rat Model of Pulmonary Arterial Hypertension This rat study protocol was approved by the Institutional Animal Care and Use Committee of Ewha Womans University, Korea, and adhered to the ARRIVE guidelines. Six-week-old rats were acclimated in the laboratory for 1 week. Subsequently, rats were subcutaneously injected with SuGen5416 (Sigma-Aldrich) at a dose of 20 mg / kg. The administered subjects were maintained in a normobaric hypoxia (10% O2) chamber (Chamber A, Biosphenix) for 3 weeks. Rats were transferred to a normoxic condition and further orally administered with control group vehicle or test compound for 5 weeks (oral, once a day, 0.1 mg per kg body weight).

[0269] Right ventricular systolic pressure (RVSP) was measured by echocardiography using a catheter-tip micromanometer (AD Instruments, UK) attached to a PowerLab 2 / 26 system. Rats were anesthetized with 2% isoflurane inhalation. The catheter was inserted into the right jugular vein, and pressure was measured for 10 s in a stable pattern. After echocardiography, rats were perfused with saline and the heart was carefully excised. The excised heart was sectioned into the right ventricle (RV), the remaining part of the septum (S; septum), and the left ventricle (LV). The weight of each heart section was measured using a microbalance. The RV / (LV+S) value was calculated and presented as the right ventricular hypertrophy index (Fulton's index).

[0270] Experimental Example 10: Histological Analysis Rats were anesthetized by inhalation of 2% isoflurane and fixed by perfusion through the heart with an aqueous heparin solution containing 37% formaldehyde. Subsequently, the left lung lobe was incised and fixed in 10% NBF for 3 days. The fixed tissues were embedded in paraffin and sectioned using a rotary microtome (Leica HistoCore MULTICUT). Two consecutive tissue sections (4 μm thick) were stained with hematoxylin and eosin (HE) on glass slides. Approximately 100 pulmonary arteries (20 - 100 μm in diameter) per tissue section were selected and measured for analysis. The lumen, internal elastic lamina, and external elastic lamina regions were quantified using NIH ImageJ v1.62. The intimal area and medial area were calculated as the values obtained by subtracting the area of the internal elastic lamina from the lumen area and the area of the external elastic lamina from the area of the internal elastic lamina, respectively. The calculated values were averaged and used for calculating the thickness of the pulmonary artery vessels.

[0271] Experimental Example 11: Immunofluorescence Staining Paraffin sections were blocked with 5% normal donkey serum (Vector Laboratories) in PBST (PBS solution containing 0.3% Triton X - 100) at room temperature for 1 hour. Subsequently, the lung tissue sections were maintained with antigens against Alexa Fluor 568 - conjugated smooth muscle α - actin (SMA; diluted 1:300) and Alexa Fluor 488 - conjugated von willebrand factor (vWF; diluted 1:200) at 4°C for 12 hours. The nuclei were labeled with DAPI. Fluorescent images were recorded at a screen magnification of 60× for more than 10 pulmonary artery vessels per tissue section in an arbitrary region using an LSM880 confocal microscope equipped with argon and helium - neon lasers.

[0272] Result 1: Peroxidase Activity against Hydrogen Peroxide - Reducing Ability The hydrogen peroxide - reducing peroxidase activities of the novel compounds of the present invention were measured respectively in reaction solutions containing thioredoxin (Trx) / thioredoxin reductase (TR). The results are summarized in the following table.

[0273]

Table 2

[0274] A5:

[0275]

Chem.

[0276] As shown in Table 2, the compounds used in the test showed almost excellent hydrogen peroxide-reducing peroxidase activity in the presence of the Trx / TR redox system. This indicates that the novel compounds have peroxidase activity that specifically mimics peroxiredoxin.

[0277] In particular, compared with the epidithiodioxopiperazine derivative compound (A5), the preferred compounds of the present invention showed similar or better peroxidase activity.

[0278] Result 2: Measurement of the ability to remove intracellular hydrogen peroxide The ability of the novel compounds of the present invention to remove intracellular hydrogen peroxide was measured using a fluorescence probe for live cell imaging (CM-H2DCF-DA). The removal ability according to the concentration of each compound was calculated as the percentage compared to the untreated group and graphed, and the concentration (EC 50 ) of the compound having 50% intracellular hydrogen peroxide removal ability is shown in the following table.

[0279]

Table 3

[0280] As shown in Table 3, the test compounds showed excellent reactive oxygen species removal ability compared to the conventional epidithiodioxopiperazine derivative compound (A5).

[0281] Result 3: Measurement of the redox potential The redox potential value of the novel compound was analyzed based on the potential values embodied in a Redox buffer solution in which reduced DTT and oxidized DTT were mixed at various ratios. The results are shown in the following table.

[0282]

Table 4

[0283] As shown in Table 4, the compounds of the present invention generally showed very low redox potential values. When compared with the potential values of Trx or GSH, it was confirmed that the novel compounds are peroxiredoxin mimetic compounds capable of a redox reaction specialized for the Trx / TR redox system in cells.

[0284] Result 4: Cytotoxicity test Cytotoxicity tests were performed on vascular endothelial cells, vascular smooth muscle cells, and hepatocytes with respect to the novel compounds. After treating each compound at various dilution concentrations, the concentration of the compound corresponding to 50% cell viability (CC 50 ) was measured. The results are shown in the following table.

[0285]

Table 5

[0286] As a result of the measurement, except for Compound 6, all CC 50 showed very low cytotoxicity with all being 100 μM or more. In particular, in the case of some compounds, there was no toxicity in the tested concentration range (10 μM to 1 mM). This represents very excellent safety compared to the conventional episulfide dioxopiperazine derivative compound (A5).

[0287] Result 5: Analysis of PDGF-induced signal transduction in smooth muscle cells In human aortic smooth muscle cells (HASMCs) and human pulmonary artery smooth muscle cells (PASMCs) in which PrxII was knocked out by injecting the PrxII siRNA, it was evaluated whether a test compound could replace the cellular function of PrxII. That is, the efficacy of the novel compound was verified in the signal transduction pathway induced by platelet-derived growth factor (PDGF) that regulates the growth and migration of aortic and pulmonary artery vascular smooth muscle cells. Activation of PDGF-induced signal transduction was analyzed by an immunoblot method using an antibody that specifically binds to phosphorylation at the 857th residue, which is the major signal transduction tyrosine residue of the PDGF receptor-β (PDGFRβ). The results are shown in FIGS. 1 to 3.

[0288] Figure 1 shows the results of confirming the effect of Compound 1 on the degree of Tyr857 phosphorylation of PDGFRβ amplified by PrxII gene deficiency in aortic smooth muscle cells (HASMCs) for each concentration. It shows that phosphorylation significantly decreases from a concentration of 2.5 nM of Compound 1.

[0289] Figure 2 shows the results of evaluating the effects of Compounds 1, 2, 5, 6, 10, 15, 19, 20, 21, 22, and 23 on the degree of Tyr857 phosphorylation of PDGFRβ amplified by PrxII gene deficiency in aortic smooth muscle cells (HASMCs). As a result of the evaluation, all of the compounds showed the ability to decrease and regulate, replacing PrxII.

[0290] Figure 3 shows the results of evaluating the effect of Compound 8 on the degree of total tyrosine phosphorylation and the phosphorylation of a signal transduction protein (e.g., PLC-γ1) downstream of PDGFRβ induced by PDGF stimulation in pulmonary artery smooth muscle cells (PASMCs). As a result of the evaluation, Compound 8 showed a significant decreasing ability.

[0291] As a result of the evaluation, the novel compound showed a result that the PDGF signal regulation ability was improved about 10-fold compared with the conventional episulfide dioxopiperazine derivative compound. It also showed that the abnormal PDGF signal transduction induced by PrxII deficiency was restored to a normal level.

[0292] Result 6: Analysis of VEGF-induced Signaling in Vascular Endothelial Cells In human aortic endothelial cells (HAECs) and human pulmonary artery endothelial cells (PAECs) in which PrxII was knocked out by injecting PrxII siRNA, it was evaluated whether a novel compound could replace the cellular function of PrxII. That is, the efficacy of the novel compound was verified in the signaling pathway induced by vascular endothelial growth factor (VEGF), which regulates the growth and migration of vascular endothelial cells. Activation of the VEGF-induced signaling pathway was analyzed by an immunoblotting method using an antibody that specifically binds to phosphorylation of Tyr1175, the major signaling tyrosine residue of VEGF receptor 2 (VEGFR2). The results are shown in Figures 4 to 6.

[0293] Figure 4 shows the results of confirming the effect of Compound 1 on the degree of Tyr1175 phosphorylation of VEGF receptor-2 amplified by PrxII gene deletion in aortic endothelial cells (HAECs) for each concentration. As a result of the confirmation, Compound 1 was shown to decrease to the normal signal from a concentration of 2.5 nM.

[0294] Figure 5 shows the results of analyzing the effects of Compounds 1, 2, 5, 6, 10, 20, 21, and 23 on the degree of VEGF receptor-2 Tyr phosphorylation amplified by PrxII gene deletion in human aortic endothelial cells (HAECs). As shown, all of these compounds strongly inhibited VEGFR2 phosphorylation in HAECs with PrxII gene deletion.

[0295] Figure 6 shows the results of analyzing the effect of Compound 8 on the degree of total tyrosine phosphorylation and phosphorylation of signaling proteins (e.g., ERK2) downstream of VEGFR2 induced by VEGF stimulation in pulmonary artery endothelial cells (PAECs). As shown, Compound 8 significantly inhibited the phosphorylation induced by VEGF.

[0296] As a result of the evaluation, the ability of the novel compound according to the present invention to regulate VEGF signaling showed a result of being about 10 times improved compared to the conventional episulfidodioxopiperazine derivative compound (A5), indicating that it restores the inactive VEGF signaling induced by PrxII deficiency to a normal level.

[0297] Result 7: Analysis of the ability of the novel compound to inhibit PDGF-induced cell proliferation in smooth muscle cells In human aortic smooth muscle cells (HASMC) and human pulmonary artery smooth muscle cells (PASMC) lacking the PrxII gene, the inhibitory ability of a novel compound that suppresses the proliferation of vascular smooth muscle cells induced by PDGF was analyzed. The results are shown in Table 6 below. In Table 6, the degree of proliferation of the compound-treated group compared to the degree of PDGF-induced proliferation in the group without treatment with the novel compound is shown as a percentage. The concentration corresponding to 50% growth inhibition is IC 50 as described in.

[0298] [Table 6]

[0299] As shown in the above table, the compound of the present invention suppresses the proliferation of HASMC and PASMC in a concentration-dependent manner and exhibits growth inhibitory activity from the lowest concentration of 2.5 nM.

[0300] Result 8: Analysis of the ability of the novel compound to promote VEGF-induced cell proliferation activity in endothelial cells In human aortic endothelial cells (HAEC) and human pulmonary artery endothelial cells (PAEC) lacking the PrxII gene, the ability of a novel compound that promotes the proliferation of vascular endothelial cells induced by VEGF was evaluated. The results are shown in Table 7 below. In Table 7, the degree of proliferation of the compound-treated group compared to the degree of VEGF-induced proliferation in the group without treatment with the novel compound is shown as a percentage. The concentration corresponding to 50% growth recovery is EC 50 as described in.

[0301] [Table 7]

[0302] As shown in the above table, the compounds of the present invention increased the aortic and pulmonary artery endothelial cells in a concentration-dependent manner. In particular, the compounds of the present invention showed an increasing trend in growth from the lowest concentration of 2.5 nM.

[0303] Result 9: Evaluation test of preclinical efficacy using a pulmonary hypertension model The therapeutic effect of the novel compound on pulmonary hypertension was evaluated in a preclinical SuHx rat model. To establish the PAH model, rats were administered Sugen (VEGFR2 inhibitor) and placed in a normal atmospheric pressure hypoxic state (10% O2) for 3 weeks. The rats were transferred to a normoxic state, and the control group vehicle or compound 1 or 8 (P.O., once a day, 0.1 mg / kg) was orally administered for 5 weeks. Then, on the day after the end of the administration, the right ventricular systolic pressure (RVSP) and the right ventricular hypertrophy index (Fulton's index) were measured. The results are shown in FIGS. 7 to 9.

[0304] FIG. 7 shows that compounds 1 and 8 significantly reduce the right ventricular systolic pressure and right ventricular hypertrophy compared to the vehicle control group.

[0305] FIG. 8 shows that compound 1 of the present invention dilates the severely constricted pulmonary artery vessels in the vehicle control group. According to the data, normal blood flow is induced in the vascular lumen by administering compound 1, and the right ventricular systolic pressure and right ventricular hypertrophy are reduced.

[0306] FIG. 9 shows the immunostaining images of the endothelial layer and the intimal SMC layer of the pulmonary artery vessels. Specifically, the lung tissue sections were immunostained with an endothelial cell-specific vWF antibody and a smooth muscle cell-specific SMA antibody. The results show that endothelial damage and medial thickness due to SMC hyperplasia occur in the vehicle control group. However, when compound 1 is administered, the endothelial layer (vWF-labeled endothelial cells (EC)) recovers and the medial thickness (SMA-labeled smooth muscle cells (SMC)) decreases.

Brief Description of the Drawings

[0307]

Figure 1

Figure 2

Figure 3

Figure 4

Figure 5

Figure 6

Figure 7

Figure 8

Figure 9

Claims

1. A compound represented by the following Chemical Formula 1 or Chemical Formula 2, or a pharmaceutically acceptable salt thereof. 【Chemical 1】 [[Chemical 2]] In Chemical Formula 1 and Chemical Formula 2, n is an integer from 1 to 3, R １ and R ２ are, independently of each other, C １－３ alkyl, C １－３ alkoxy-C １－３ alkyl, -(CH ２ ) １－３ -C(R')(R")OH, -(CH ２ ) １－３ -N(R')(R"), -(CH ２ ) ０－３ -alkenyl, -(CH ２ ) ０－３ -alkynyl, -(CH ２ ) ０－３ -C(R')(R")CO ２ H, -(CH ２ ) ０－５ -heterocycloalkyl, -(CH ２ ) ０－５ -cycloalkyl, -(CH ２ ) ０－５ -aryl, or -(CH ２ ) ０－５ -heteroaryl, where the alkyl, heterocycloalkyl, cycloalkyl, aryl and heteroaryl are unsubstituted or substituted with one or more substituents selected from the group consisting of C １－３ alkyl, -CF ３ , C １－３ alkoxy, -OCF ３ , halogen, CN, amino, -N(R')(R"), -OH, -COOH, -COO-C １－３ alkyl, and =O, where R' and R" are, independently of each other, hydrogen or C １－３ alkyl, R ３ is C １－３ alkyl, -(CH ２ ) ０－３ -aryl, or -(CH ２ ) ０－３ -heteroaryl, where the aryl or heteroaryl is unsubstituted or is substituted with one or more substituents selected from the group consisting of C １－３ alkyl, -CF ３ , C １－３ alkoxy, -OCF ３ , halogen, -CN, amino, -OH, and -COOH; or, R ２ and R ３ are linked to each other to form any one of the following structures fused with the piperazinedione present in Chemical Formula 1: [Chemical Formula 3] Here, X is S, SO ２ , CH ２ , O or NR ６ , where R ６ is hydrogen or C １－３ alkyl, R ４ is hydrogen or C １－３ alkyl, and R ５ is hydrogen, C １－３ alkyl, -(CH ２ ) １－２ -aryl, or -(CH ２ ) １－２ -heteroaryl.

2. n is an integer from 1 to 3, R １ and R ２ are, independently of one another, C １－３ alkyl, -(CH ２ ) １－２ -heterocycloalkyl, -(CH ２ ) １－２ -aryl, or -(CH ２ ) １－２ -heteroaryl, where the alkyl, heterocycloalkyl, aryl, and heteroaryl are unsubstituted or substituted with one or more substituents selected from the group consisting of C １－３ alkyl, -CF ３ , C １－３ alkoxy, CN, halogen, -OH, -COOH, and -COO-C １－３ alkyl, R ３ is C １－３ alkyl, -CH ２ -aryl, or -CH ２ -heteroaryl, where aryl or heteroaryl is unsubstituted or substituted with one or more substituents selected from the group consisting of methyl, methoxy, halogen, -CN, amino, -OH, and -COOH; or R ２ and R ３ are each linked to each other to form any one of the following structures fused with the piperazinedione present in Chemical Formula 1: 【Chemical 4】 Here, X is S, SO ２ , CH ２ , O or NR ６ , and R ６ is hydrogen or C １－３ alkyl, R ４ is hydrogen or C １－３ alkyl, and R ５ is hydrogen, C １－３ alkyl, or -(CH ２ ) １－２ -aryl, The compound according to Claim 1, or a pharmaceutically acceptable salt thereof.

3. n is 1, R １ and R ２ are, independently of each other, C １－３ alkyl, -CH ２ -piperidyl, -CH ２ -morpholinyl, -CH ２ -piperazinyl, -CH ２ -phenyl, -CH ２ -naphthyl, -CH ２ -pyridyl, -CH ２ -quinolinyl, -CH ２ -pyrazolyl, -CH ２ -thiophen-2-yl, -CH ２ -benzo[d]thiazol-2-yl, -CH ２ -pyrimidyl, -CH ２ -1H-imidazol-4-yl, -CH ２ -1H-imidazol-2-yl, -CH ２ -thiazol-4-yl, -CH ２ -thiazol-5-yl, -CH ２ -isoxazolyl, -CH ２ -indol-2-yl, -CH ２ -indol-3-yl, -CH ２ -benzimidazol-5-yl, -CH ２ -quinolin-4-yl, -CH ２ -quinazolin-2-yl, or -CH ２ -quinazolin-4-yl, wherein said piperidyl, morpholinyl, piperazinyl, phenyl, naphthyl, pyridyl, quinolinyl, pyrazolyl, thiophene, benzo[d]thiazole, pyrimidyl, imidazole, thiazole, isoxazolyl, indole, benzimidazole, quinoline, and quinazoline are unsubstituted or are substituted with one or more substituents selected from the group consisting of C １－３ alkyl, -CF ３ 、C １－３ alkoxy, CN, halogen, and -COO-C １－３ alkyl, R ３ is C １－３ alkyl or -CH ２ -aryl, or R ２ and R ３ are each linked to each other to form any one of the following structures fused with the piperazinedione present in Chemical Formula 1: 【Chemical Formula 5】 Here, X is O or NR ６ wherein, R ６ is methyl, R ４ is hydrogen, and R ５ is hydrogen, The compound according to Claim 2, or a pharmaceutically acceptable salt thereof.

4. The compound is 1,6,8 - trimethyl - 2,3 - dithia - 6,8 - diazabicyclo[3.2.2]nonane - 7,9 - dione (Compound 1), 6 - benzyl - 1,8 - dimethyl - 2,3 - dithia - 6,8 - diazabicyclo[3.2.2]nonane - 7,9 - dione (Compound 2), 1,8 - dimethyl - 6 - (3,4,5 - trimethoxybenzyl) - 2,3 - dithia - 6,8 - diazabicyclo[3.2.2]nonane - 7,9 - dione (Compound 3), 6 - (3,5 - difluorobenzyl) - 1,8 - dimethyl - 2,3 - dithia - 6,8 - diazabicyclo[3.2.2]nonane - 7,9 - dione (Compound 4), 1,8 - dimethyl - 6 - (quinolin - 2 - ylmethyl) - 2,3 - dithia - 6,8 - diazabicyclo[3.2.2]nonane - 7,9 - dione (Compound 5), 1,8 - dimethyl - 6 - (pyridin - 2 - ylmethyl) - 2,3 - dithia - 6,8 - diazabicyclo[3.2.2]nonane - 7,9 - dione (Compound 6), 11 - benzyltetrahydro - 5H,7H - 4,9a - (epiminomethano)pyrrolo[2,1 - c][1,2,4]dithiazepin - 5,10 - dione (Compound 7), 12 - benzyltetrahydro - 5H,10H - 4,10a - (epiminomethano)[1,4]oxazino[3,4 - c][1,2,4]dithiazepin - 5,11 - dione (Compound 8), 12 - (pyridin - 4 - ylmethyl)tetrahydro - 5H,10H - 4,10a - (epiminomethano)[1,4]oxazino[3,4 - c][1,2,4]dithiazepin - 5,11 - dione (Compound 9), 12 - ethyltetrahydro - 5H,10H - 4,10a - (epiminomethano)[1,4]oxazino[3,4 - c][1,2,4]dithiazepin - 5,11 - dione (Compound 10), 11 - (pyridin - 4 - ylmethyl)tetrahydro - 5H,7H - 4,9a - (epiminomethano)pyrrolo[2,1 - c][1,2,4]dithiazepin - 5,10 - dione (Compound 11), 1,8-Dimethyl-6-((6-methylpyridin-2-yl)methyl)-2,3-dithia-6,8-diazabicyclo[3.2.2]nonane-7,9-dione (Compound 12), 1,8-Dimethyl-6-((1-methyl-1H-pyrazol-4-yl)methyl)-2,3-dithia-6,8-diazabicyclo[3.2.2]nonane-7,9-dione (Compound 13), 1,8-Dimethyl-6-(thiophen-2-ylmethyl)-2,3-dithia-6,8-diazabicyclo[3.2.2]nonane-7,9-dione (Compound 14), 6-(Benzo[d]thiazol-2-ylmethyl)-1,8-dimethyl-2,3-dithia-6,8-diazabicyclo[3.2.2]nonane-7,9-dione (Compound 15), 1,6-Dimethyl-8-(pyrimidin-2-ylmethyl)-2,3-dithia-6,8-diazabicyclo[3.2.2]nonane-7,9-dione (Compound 16), 1,6-Dimethyl-8-((1-methyl-1H-imidazol-4-yl)methyl)-2,3-dithia-6,8-diazabicyclo[3.2.2]nonane-7,9-dione (Compound 17), Methyl 4-((1,6-dimethyl-7,9-dioxo-2,3-dithia-6,8-diazabicyclo[3.2.2]nonan-8-yl)methyl)benzoate (Compound 18), 1-Benzyl-6,8-dimethyl-2,3-dithia-6,8-diazabicyclo[3.2.2]nonane-7,9-dione (Compound 19), 6,8-Diethyl-1-methyl-2,3-dithia-6,8-diazabicyclo[3.2.2]nonane-7,9-dione (Compound 20), 12-Benzyl-9-methylhexahydro-5H-4,10a-(epiminomethano)pyrazino[2,1-c][1,2,4]dithiazepine-5,11-dione (Compound 21), 12-Benzyl-9-methylhexahydro-5H-4,10a-(epiminomethano)pyrazino[2,1-c][1,2,4]dithiazepine-5,11-dione (Compound 22), 12-Benzyl-9-methylhexahydro-5H-4,10a-(epiminomethano)pyrazino[2,1-c][1,2,4]dithiazepine-5,11-dione (Compound 23), 6-Benzyl-1,4,8-trimethyl-2,3-dithia-6,8-diazabicyclo[3.2.2]nonane-7,9-dione (Compound 24), or 6-Benzyl-1,4,8-trimethyl-2,3-dithia-6,8-diazabicyclo[3.2.2]nonane-7,9-dione (Compound 25), The compound according to claim 1 or a pharmaceutically acceptable salt thereof.

5. A method for treating or preventing a vascular disease, comprising administering a therapeutically effective amount of the compound according to any one of claims 1 to 4 or a pharmaceutically acceptable salt thereof to an individual in need of treatment or prevention of a vascular disease or suspected of having a vascular disease.

6. The method according to claim 5, wherein the vascular disease is selected from the group consisting of hypertension, ischemic coronary artery disease, cerebral artery occlusion, arteriosclerosis, peripheral artery occlusion disease, thromboembolism, diabetic foot lesion, venous ulcer, deep vein thrombosis, vasoconstriction, arteritis, and vascular restenosis.

7. The method according to claim 6, wherein the vascular disease is ischemic coronary artery disease, arteriosclerosis, vascular restenosis, or pulmonary hypertension.

8. Use of the compound according to any one of claims 1 to 4 or a pharmaceutically acceptable salt thereof, for the manufacture of a medicament for the treatment or prevention of a vascular disease.

9. The use according to claim 8, wherein the vascular disease is selected from the group consisting of hypertension, ischemic coronary artery disease, cerebral artery occlusion, arteriosclerosis, peripheral artery occlusion disease, thromboembolism, diabetic foot lesion, venous ulcer, deep vein thrombosis, vasoconstriction, arteritis, and vascular restenosis.

10. The use according to claim 9, wherein the vascular disease is ischemic coronary artery disease, arteriosclerosis, vascular restenosis, or pulmonary hypertension.

11. A 6-(1-hydroxyalkyl)piperazine-2,5-dione derivative represented by the following chemical formula 4 is reacted with sulfur (S ８ ), LiHMDS (lithium bis(trimethylsilyl)amide) or NaHMDS (sodium bis(trimethylsilyl)amide), and a method for producing a compound represented by the following chemical formula 3, characterized by the above reaction. 【Chemical Formula 6】 【Chemical Formula 7】 In the above chemical formula 4, R １ , R ２ , R ３ and R ４ are the same as those in chemical formula 1 described in claim 1, R ５ is hydrogen, and R is a protecting group, In the above chemical formula 3, R １ , R ２ , R ３ and R ４ are the same as the chemical formula 1 according to claim 1, R ５ is hydrogen, and n is 2 or 3.

12. (S1) Reacting a compound represented by the following chemical formula 4 with (a) sulfur (S ８ ) and (b) LiHMDS (lithium bis(trimethylsilyl)amide) or NaHMDS (sodium bis(trimethylsilyl)amide) to produce a compound represented by the following chemical formula 3; (S2) reducing the compound of the following Chemical Formula 3 to produce a compound represented by the following Chemical Formula 2; and (S3) forming an intramolecular disulfide crosslink bond from the compound represented by the following Chemical Formula 2, A method for producing a compound represented by the following Chemical Formula 1'. 【Chemical 8】 【Chemical Formula 9】 【Chemical Formula 10】 【Chemical 11】 In the above chemical formulas 4, 3, 2, and 1', R １ , R ２ , R ３ and R ４ are the same as the chemical formula 1 described in claim 1, R ５ is hydrogen, R is a protecting group, and n is 2 or 3.

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