KR20090031333A - Beta-secretase inhibiting compounds - Google Patents

Abstract

A pharmaceutical composition including a compound for beta-secretase inhibition is provided to prevent and treat a neurodegenerative disease. A pharmaceutical composition comprises a compound represented by the formula 1. The compound realizes a function of beta-secretase inhibition. In the formula 1, n is 0~2 and A is represented by the formual 2. In the formula 2, W, X, Y and Z are independently carbon atom or one or two nitrogen atoms; R1 is -(CR6R7)m-R8, wherein m is 0~2, R6 is selected from the group consisting of hydrogen, alkyl and alkoxy, and R7 is selected from the group consisting of hydrogen, alkyl and - C(O)NHR6.

Description

Novel Beta-Secretase Inhibiting Compounds

The present invention relates to novel compounds that inhibit the activity of beta-secretase, pharmaceutically acceptable salts or isomers thereof, methods for their preparation, and pharmaceutical compositions containing them in pharmacologically effective amounts.

Alzheimer's disease (senile dementia) is a neurological disease that develops gradually with age, accounting for about 50-70% of all dementia patients. The main symptoms are memory loss and loss of cognitive function, and statistically, most of them develop before and after 65 years and die after about 9 years of progression. As society develops and ages, the number of patients suffering from Alzheimer's disease is increasing. It is estimated that there will be about 6 million patients in the US within the next decade, which will increase over time.

Korean society is also on the rise of the elderly, especially the social problems caused by senile dementia patients. However, to date, no therapeutic agent has been developed to treat the underlying cause of Alzheimer's disease, and acetylcholine esterase inhibitor is the only one that can be used as a general therapeutic agent. For example, Pfizer's Aricept ^™ , Norvatis' Exelon ^™ , and Janssen's Reminyl ^™ are commonly used as representative therapeutic agents.

These acetylcholine esterase inhibitors do not cure the underlying cause of Alzheimer's disease, and only provide temporary relief of symptoms in some patients (approximately 40-50%) and do not last long. Therefore, it is difficult to define it as a cure in a strict sense. In addition, the nature of the disease is required to take long-term, in the case of the drug, accompanied by various side effects, including liver toxicity has also been revealed as a problem.

Therefore, the development of a therapeutic agent that can prevent the progress of the disease is an urgent problem. As a way to find the cause of Alzheimer's disease, studies on patients with hereditary early on-set Alzheimer's disease have been underway for a long time, and as a result, mutations in several genes have been found to be the main cause. .

As a common physiological result of these genetic pathogens, it was found that the production of beta amyloid consisting of 42 amino acids (hereinafter sometimes abbreviated as 'Ab42') is increased, so that Ab42 production is a major cause of Alzheimer's disease. (causative agent). Therefore, when a method for reducing the production of Ab42 is developed, it is expected to be the most direct and effective therapeutic agent that can block the pathogenesis of Alzheimer's disease itself.

Beta amyloid proteins are produced by a series of cleavage by three secretases from a large molecular weight precursor called Amyloid Precursor Protein (APP), which is made in neurons. This process is carried out in a membrane carrier called Golgi (or Endosome) in the cytoplasm of neurons, and APP and its protease secretases are located and act on the membrane. The N-terminus of beta amyloid (Ab) corresponds to the 99th amino acid from the C-terminus of APP, and this region is abbreviated as beta-secretase (hereinafter, sometimes called 'BACE' (beta-site APP Cleaving Enzyme)). Beta amyloid (Ab) protein is cleaved by a gamma-secretase cleavage of the C-terminus of beta amyloid (Ab) located in the membrane, and then secreted out of neurons. The APP precursor may be cleaved at a completely different position via a separate pathway, with alpha-secretase cutting off the intermediate region of the Ab (between the 16th and 17th amino acids from the N-terminus). Molecular weight sAPPa is produced and secreted, and no further beta amyloid is produced.

The beta amyloid (Ab) protein of various lengths is produced by the BACE and gamma secretase, and 40 amino acids (Ab40) and 42 amino acids (Ab42) are predominant. Ab42, unlike Ab40, tends to aggregate more easily, and when accumulated in the brain of the patient, it promotes the formation of amyloid plaque, which causes neural cells in the surrounding area to be necrotic, which is Alzheimer's disease. It is estimated to be the main pathogenesis of the disease.

Ab40 and Ab42 are produced in a ratio of about 9: 1 under normal circumstances. However, the overall increase in the synthesis amount of the two types of amyloid protein or the selective increase of Ab42 by the aforementioned mutation of the Presenilin 1 & 2 gene further promotes the onset of Alzheimer's disease, and the symptoms are also known to be serious. have. Therefore, reducing the synthesis of Ab42 can be said to be the most important part of the development of Alzheimer's treatment, which requires the development of beta or gamma secretase inhibitors.

Many multinational pharmaceutical companies are investing heavily in R & D in this area, and beta or gamma secretase inhibitors that reduce the production of beta amyloid consisting of 42 amino acids, which are believed to be the primary cause of Alzheimer's disease. Development is the dominant. However, in view of the development of a therapeutic agent that can prevent the progression of the disease by inhibiting the production mechanism of beta amyloid itself in the world, there is no place to show the remarkable research results so far.

On the other hand, both beta and gamma secretase are known as aspartic protease and are known to be attached to the membrane. However, in the case of gamma secretase inhibitors, the gene has not yet been identified. In addition, the substrate is not limited to the APP precursor protein and is known to be involved in the cleavage of Notch protein, which is known to play an important role in cell-fate decision during differentiation. In particular, transgenic animals from which the gamma secretase gene itself has been removed have not only been shown to die in the fetus, but the prospects are unclear because of the recent clinical trial results of the gamma secretase inhibitors. Thus, much remains to be seen whether gamma secretase inhibitors can be developed as safe treatments for Alzheimer's disease.

On the other hand, in the case of BACE, several pharmaceutical companies have identified the gene in a variety of ways, and it was published in 1999 showing its activity as a beta secretase in vivo. In addition, the X-ray crystal structure has been found, and a high affinity peptide inhibitor for this is also well known. In particular, genetically deficient transgenic animals showed normal traits with no abnormalities, suggesting that BACE specific inhibitors could be developed as a safer and more effective treatment for dementia. Thus, compared to gamma secretase, BACE is a more promising target.

As discussed above, considering that existing commercially available drugs are only effective in alleviating symptoms and have no effect on the progress of the disease itself, the development of BACE specific inhibitors is a new concept for Alzheimer's disease. There is no doubt that this will be a breakthrough in drug development. In addition, BACE inhibitors have been studied relatively recently, and thus, a complete small molecule chemical compound has not been announced yet except for peptide variants.

Recently, as BACE inhibitors, Merck (WO 2006/078577, WO 2006/060109, Wo 2006/057983), Ellan (WO 2004/022523, WO 2005/095326), Schering-Plaugh (WO 2006/014762, Wo 2006 / 014944), BMS (WO 2005/182105, WO 2005/030758), Eli-Lilly (WO 2005/108358, WO 2006/034093), and others. These compounds were compounds showing small molecular weight and brain permeability. However, the compounds are cat. The limit was exposed due to selectivity to D and cytotoxicity. Recently, Merck compounds have been attracting attention, especially Merck's dicarbonyl compound introduced in the early Sunesis, followed by sulfone amide form to show excellent inhibitory effects and selectivity. Many companies are now announcing compounds in the form of Merck compounds.

However, these compounds, in particular sulfon amide compounds, have excellent BACE enzyme inhibitory effects and Cat. The developmental potential has been shown to be due to the excellent selectivity to D. However, the low oral potential and the brain permeability have difficulties in development, and there are few reports on compounds that have dropped Ab42 in actual animal models. Lilly showed a small molecular weight mimicking compound outside sulfon amide, showing its efficacy in SC in real animal models (reduction of Ab42), but with low oral potential and low brain permeability. It is the situation that the compound which shows the outstanding effect so far is not announced. Scheing-Plaugh, too, has announced a peptidic compound, but as these compounds are known as Pgp substrates, they do not show efficacy in the brain of real animal models.

In other words, peptide beta secretase inhibitors are limited to oral preparations due to their molecular specificity. In particular, sulfon amide-type compounds have low brain permeability, making it difficult to observe efficacy in animal models. .

Therefore, there is a need for development of a compound having an inhibitory effect of inhibiting excellent beta secretase out of the sulfon amide form.

Accordingly, an object of the present invention is to solve the problems of the prior art and the technical problem that has been requested from the past.

Specifically, an object of the present invention is to improve the brain permeability which is a technical problem of the conventional sulfone amide-based compounds, structurally out of the sulfon amide form, the inhibitory effect of beta secretase through the heterocyclic compound of 5-membered ring It is to provide a novel compound having a pharmaceutically acceptable salt or isomer thereof.

Another object of the present invention is to provide a method for preparing such novel compounds.

Still another object of the present invention is to provide a composition for inhibiting activity of beta-secretase, which comprises a pharmacologically effective amount of the compound as an active ingredient.

Another object of the present invention is to use such a novel compound as an active ingredient, to provide a use for the treatment or prevention of neurodegenerative diseases such as cognitive function improvement or Alzheimer's disease.

In order to achieve this object, the present invention provides a compound represented by the following formula (I), a pharmaceutically acceptable salt or isomer thereof.

(I)

(I)

Where

n is 0 to 2;

A is represented by the following formula (II),

(II)

(II)

Wherein W, X, Y and Z are each independently C or N atoms, one or two of which is N;

R ₁ is-(CR ₆ R ₇ ) _m -R ₈ , where m is 0 to 2, R ₆ is selected from the group consisting of hydrogen, alkyl and alkoxy, R ₇ is hydrogen, alkyl and -C (O NHR ₆ is selected from the group consisting of R ₈ is alkyl, alkoxy, arylalkoxy, cycloalkyl, heterocycle, heteroaryl, -C (O) R ₆ and -C (O) NR ₆ R ₇ Wherein R ₆ and R ₇ are as defined above;

R ₂ is selected from the group consisting of hydrogen, alkyl and-(CR ₆ R ₇ ) _m -R ₉ , wherein m is 0 to 2, R ₆ and R ₇ are as defined above and R ₉ is aryl ,

R ₃ is selected from the group consisting of hydrogen, alkyl, alkoxy and — (CR ₆ R ₇ ) _p —R ₁₀ , wherein p is 0 to 2, R ₆ and R ₇ are as defined above and R ₁₀ is cyclo Alkyl, aryl, heterocycle, and heteroaryl;

R ₄ is selected from the group consisting of hydrogen, alkyl, cycloalkyl and-(CH ₂ ) _p -A'-R ₁₁ , wherein A 'is aryl or heteroaryl, and R ₁₁ is hydrogen, halogen, hydroxy, alkyl , Alkoxy and —NR ₆ R ₇ wherein R ₆ and R ₇ are as defined above;

In the above, R ₆ and R ₇ are bonded to each other to form a cyclized structure or an acyclic ring.

The alkyl, alkoxy, aryl, cycloalkyl, heterocycle, and heteroaryl may have a substituted or unsubstituted structure, where the substituents are halogen, amino, alkylamino, dialkylamino, alkylacylamino, C _1- C ₄ It may be at least one selected from the group consisting of alkyl, hydroxy, C ₁ -C ₄ alkoxy, arylalkoxy, and oxo. In some cases, these substituents may also be substituted, in which case the substituents are as described above. In addition, the substituents may be bonded to each other to form a ring structure.

In addition, the heteroaryl and heterocycle are each independently a 4-8 membered ring containing 1 to 3 heteroatoms selected from the group consisting of O, N, and S, 0 to 2 double bonds, Preferably, it is a 5-6 membered ring containing 1 or 2 double bonds.

The compound of formula (I) according to the present invention has a structure different from that of beta secretase inhibitors, such as sulfonamide-based or peptide-based compounds, which are known in the art, and as shown in the following experimental examples, cognitive function is improved. Or an excellent inhibitory effect on human beta secretase related to the prevention and treatment of neurodegenerative diseases such as Alzheimer's disease.

In the definitions for the substituents of the compounds of formula (I) according to the invention, the term 'alkyl' means an aliphatic hydrocarbon group. The alkyl moiety can be a "saturated alkyl" group, meaning that it does not contain any alkenes or alkyne moieties. The alkyl moiety may be an "unsaturated alkyl" moiety, meaning that it contains at least one alkene or alkyne moiety. "Alkene" moiety means a group of at least two carbon atoms consisting of at least one carbon-carbon double bond, and an "alkyne" moiety means at least two carbon atoms having at least one carbon-carbon triple bond Means a group. The alkyl moiety, whether saturated or unsaturated, may be branched, straight chain or cyclic.

Alkyl groups may have from 1 to 20 carbon atoms. The alkyl group may preferably be C ₁ -C ₈ alkyl having 1 to 8 carbon atoms. For example, C ₁ -C ₄ alkyl is one to four carbon atoms in the alkyl chain, i.e., the alkyl chain is methyl, ethyl, propyl, iso-propyl, n-butyl, iso-butyl, sec-butyl and t It is selected from the group consisting of -butyl. In addition, when used alone or in combination with alkyloxy, respectively, it means a straight or branched chain hydrocarbon radical.

Typical alkyl groups include methyl, ethyl, propyl, isopropyl, butyl, isobutyl, t-butyl, pentyl, hexyl, ethenyl, propenyl, butenyl, cyclopropyl, cyclobutyl, cyclopentyl, cyclohexyl, etc. It is not limited only to these.

The term 'alkoxy' means oxo alkyl having 1 to 8 carbon atoms.

The term 'cycloalkyl' means an unsaturated aliphatic 3-10 membered ring including cyclohexyl.

The term 'aryl' refers to an aryl group having at least one ring having a shared pi electron field and comprising a carbocyclic aryl (eg phenyl) and a heterocyclic aryl group (eg pyridine) Means. The term includes monocyclic or fused ring polycyclic (ie rings that divide adjacent pairs of carbon atoms) groups. 4 to 10 membered, preferably 6 to 10 membered aromatic monocyclic or multicyclic ring group containing phenyl, naphthyl and the like.

The term 'heteroaryl' includes one to three heteroatoms selected from the group consisting of N, O and S, and benzo or C ₃ -C ₈ It means an aromatic 4-8 membered ring, preferably a 5-6 membered ring, which can be fused with cycloalkyl. Examples of monocyclic heteroaryl include thiazole, oxazole, thiophene, furan, pyrrole, imidazole, isoxazole, pyrazole, triazole, thiadiazole, tetrazole, oxadiazole, pyridine, pyridazine, Pyrimidine, pyrazine and similar groups, but is not limited to these. Examples of bicyclic heteroaryl include indole, indolin, benzothiophene, benzofuran, benzimidazole, benzoxazole, benzisoxazole, benzthiazole, benzthiadiazole, benztriazole, quinoline, isoquinoline, purine , Furypyridine and similar groups, but are not limited to these.

The term 'heterocycle' includes 1 to 2 heteroatoms selected from the group consisting of N, O and S, and is selected from benzo or C ₃ -C ₈ 3 to 10 membered ring, preferably 4 to 8 membered ring, more preferably 5 to 6 membered ring, which may be fused with cycloalkyl and are saturated or include 1 or 2 double bonds. Furan, thiophene, pyrrole, pyrroline, pyrrolidine, oxazole, thiazole, imidazole, imidazoline, imidazolidine, pyrazole, pyrazoline, pyrazolidine, isothiazole, triazole, thia Diazole, pyran, pyridine, piperidine, morpholine, thiomorpholine, pyridazine, pyrimidine, pyrazine, piperazine, triazine, hydrofuran and the like, but are not limited thereto.

Other terms may be interpreted as meanings commonly understood in the field to which the present invention belongs.

Compounds of formula (I) according to the invention are preferably

R_OneIs-(CR₆R₇)_m-R₈Where m is 0 to 2 and R₆Is hydrogen or C_One-C₈ Alkyl, R₇Silver hydrogen, C_One-C₈ Alkyl and -C (O) NHR₆Is selected from the group consisting of R₈Silver c_One-C₈ Alkyl, C_One-C₈ Alkoxy, arylalkoxy, cycloalkyl, heteroaryl, -C (O) R₆ And -C (O) NR₆R₇It is selected from the group consisting of;

R ₂ is selected from the group consisting of hydrogen, C ₁ -C ₈ alkyl and-(CR ₆ R ₇ ) _m -R ₉ , wherein m is 0 to 2 and R ₉ is phenyl;

R ₃ is selected from the group consisting of hydrogen, C ₁ -C ₈ alkyl, C ₁ -C ₈ alkoxy and — (CR ₆ R ₇ ) _p -R ₁₀ , wherein p is 0 to 2 and R ₁₀ is cycloalkyl , Aryl, heterocycle and heteroaryl;

R ₄ is selected from the group consisting of hydrogen, C ₁ -C ₆ alkyl, cycloalkyl of a 5 or 6 membered ring, and — (CH ₂ ) _p —A′-R ₁₁ , wherein A ′ is aryl or heteroaryl, R ₁₁ is hydrogen, halogen, hydroxy, C ₁ -C ₈ Alkyl is selected from the group consisting of C ₁ -C ₈ alkoxy and -NR ₆ R _7, R ₆ And R ₇ may be bonded to each other to effect cyclization.

A in Formula I is a 5-membered ring having the structure of Formula II and containing 1 or 2 N atoms. R ₁ and R ₂ in the substituent A may be bonded to the same ring element, may be bonded separately, or may be bonded to a ring element connected to the substituent.

A for each case may be any of the heterocycles of the formulas (i) to (v).

(i)

(ii)

(iii)

(iv)

Preferably, R ₁ is-(CR ₆ R ₇ ) _m -R ₈ , where m is 0 to 2, R ₆ is hydrogen or C ₁ -C ₈ alkyl, R ₇ is hydrogen, C ₁ -C ₈ Alkyl and -C (O) NHR ₆ , R ₈ is C ₁ -C ₈ alkyl, C ₁ -C ₈ alkoxy, arylalkoxy, cycloalkyl, heteroaryl, -C (O) R ₆ and -C (O) NR ₆ R ₇ .

R ₁ is more preferably-(CHR ₇ ) _m -R ₈ , where m is 0 or 1, R ₇ is hydrogen, C ₁ -C ₄ alkyl or -C (O) NHR ₆ , and R ₈ is C ₁ -C ₈ alkyl, arylalkoxy, cycloalkyl, -C (O) R ₆ and -C (O) NR ₆ R ₇ is selected from the group consisting of particularly preferred R ₁ is hydrogen, benzyl, dimethylamino-carbo Neyl, propyl-carbonyl, phenethyl, butyl, pentyl, ethylcarbamoyl, trifluorobenzyl, 4-trifluoromethylbenzyl, 2-trifluoromethylbenzyl phenoxyethyl, 2,4,5-trifluoro Lobenzyl, 4-fluorobenzyl, 3,4-difluorobenzyl, 2,4-difluorobenzyl, 3,5-difluorobenzyl 3,5-dimethoxybenzyl, 3-methoxybenzyl, 3 , 5-dibromophenyl, cyclohexylmethyl and 2-acetylaminophenyl.

R ₂ is preferably selected from the group consisting of hydrogen, C ₁ -C ₈ alkyl and benzyl, more preferably from the group consisting of hydrogen, methyl and benzyl.

R ₃ is preferably hydrogen, C ₁ -C ₈ Alkyl, cycloalkyl and aryl, wherein alkyl, cycloalkyl and aryl are each independently halogen, hydroxy, C ₁ -C ₄ Alkyl, C ₁ -C ₄ It may be a structure substituted or unsubstituted by one or more substituents selected from the group consisting of alkoxy and arylalkoxy. More preferred R ₃ is phenyl unsubstituted or substituted by one or more by a substituent selected from the group consisting of fluorine, chlorine and methyl, particularly preferably phenyl or 3,5-difluorobenzyl.

R ₄ is preferably selected from the group consisting of hydrogen, C ₁ -C ₆ alkyl, cycloalkyl, — (CH ₂ ) —A′-R ₁₁ , wherein A ′ is aryl or heteroaryl, and R ₁₁ is hydrogen , Halogen, hydroxy, alkyl, alkoxy and -NR ₆ R ₇ wherein R ₆ is selected from the group consisting of hydrogen, C ₁ -C ₃ alkyl and C ₁ -C ₃ alkoxy, R ₇ Is selected from the group consisting of hydrogen, C ₁ -C ₄ alkyl and —C (O) NHR ₆ . More preferred R ₄ is 3-dimethylaminobenzyl, 3-isopropylbenzyl, 3-trifluoromethoxybenzyl, 3-trifluoromethylbenzyl, 3-ethylbenzyl, 3-t-butylbenzyl, ethyl and cyclohexyl It is selected from the group consisting of.

The compounds according to the invention can also form pharmaceutically acceptable salts. The 'pharmaceutically acceptable salts' are salts with acids which form non-toxic acid addition salts containing pharmaceutically acceptable anions, such as sulfuric acid, hydrochloric acid, nitric acid, phosphoric acid, hydrobromic acid, hydroiodic acid, and the like. Organic carboxylic acids, such as tartaric acid, formic acid, citric acid, acetic acid, trichloroacetic acid, trifluoroacetic acid, gluconic acid, benzoic acid, lactic acid, fumaric acid, maleic acid, methanesulfonic acid, benzenesulfonic acid, p-toluenesulfonic acid or naphthalene Acid addition salts formed by sulfonic acids such as sulfonic acids and the like. These acid addition salts can be prepared by known techniques based on the structure of formula (I).

The compounds of the present invention or pharmaceutically acceptable salts thereof may also exist in the form of hydrates or solvates.

The 'isomer' refers to a compound of the present invention or a salt thereof having the same chemical formula or molecular formula but different optically or stericly. The compounds of formula (I) according to the invention may have an asymmetric carbon center and therefore exist as optical isomers (R or S isomers), racemates, diastereomeric mixtures and individual diastereomers, when having double bonds, Geometric isomers (trans, cis isomers) may also be present. All these isomers and mixtures thereof are also within the scope of the present invention.

Unless stated otherwise below, compounds of formula (I) include pharmaceutically acceptable salts and isomers thereof, all of which are to be construed as being within the scope of the present invention. For convenience of explanation, these are referred to herein simply as compounds of formula (I).

Representative compounds of formula I according to the present invention include the following compounds:

(3S) -1-Benzyl-N-[(2S, 3R) -4- (3-dimethylaminophenyl) methylamino-3-hydroxy-1-phenylbutan-2-yl] -5-oxopyrrolidine -3-carboxamide;

(3R) -1-benzyl-N-[(2S, 3R) -4- (3-dimethylaminophenyl) methylamino-3-hydroxy-1-phenylbutan-2-yl] -5-oxopyrrolidine -3-carboxamide;

(2S) -N-[(2S, 3R) -4- (3-dimethylaminophenyl) methylamino-3-hydroxy-1-phenylbutan-2-yl] -5-oxopyrrolidine-2-car Radiamide;

(2R) -N-[(2S, 3R) -4- (3-dimethylaminophenyl) methylamino-3-hydroxy-1-phenylbutan-2-yl] -5-oxopyrrolidine-2-car Radiamide;

2-((3S) -1-benzyl-5-oxopyrrolidin-3-yl) -N-[(2R, 3R) -4- (3-dimethylaminophenyl) methylamino-3-hydroxy-1- Phenylbutan-2-yl] acetamide;

2-((3R) -1-benzyl-5-oxopyrrolidin-3-yl) -N-[(2R, 3R) -4- (3-dimethylaminophenyl) methylamino-3-hydroxy-1- Phenylbutan-2-yl] acetamide;

(4S) -1-benzyl-N-[(2S, 3R) -4- (3-dimethylaminophenyl) methylamino-3-hydroxy-1-phenylbutan-2-yl] -2-oxoimidazoli Dine-4-carboxamide;

(4S) -N-[(2S, 3R) -4- (3-dimethylaminophenyl) methylamino-3-hydroxy-1-phenylbutan-2-yl] -2-oxoimidazolidine-4- Carboxamides;

(4S) -1- [2-[(2S, 3R) -4- (3-dimethylaminophenyl) methylamino-3-hydroxy-1-phenylbutan-2-yl] amino-2-oxoethyl]- N, N-dimethyl-2-oxoimidazolidine-4-carboxamide;

(4S) -1-butanoyl-N-[(2S, 3R) -4- (3-dimethylaminophenyl) methylamino-3-hydroxy-1-phenylbutan-2-yl] -2-oxoimida Zolidine-4-carboxamide;

(4S) -N- [4- (3-dimethylaminophenyl) methylamino-3-hydroxy-1-phenylbutan-2-yl] -2-oxo-1-phenethylimidazolidine-4-car Radiamide;

N-[()-4- (3-dimethylaminophenyl) methylamino-3-hydroxy-1-phenylbutan-2-yl] -2- (1-methyl-oxopyrrolidin-3-yl) acet amides;

1-butyl-N- [4- (3-dimethylaminophenyl) methylamino-3-hydroxy-1-phenylbutan-2-yl] -2-oxoimidazolidine-4-carboxamide;

(3S) -N-[(2S, 3R) -4- (3-dimethylaminophenyl) methylamino-3-hydroxy-1-phenylbutan-2-yl] -5-oxo-1-phenethylpyrrolidine -3-carboxamide;

(3R) -N-[(2S, 3R) -4- (3-dimethylaminophenyl) methylamino-3-hydroxy-1-phenylbutan-2-yl] -5-oxo-1-phenethylpyrrolidine -3-carboxamide;

1- (2-amino-2-oxoethyl) -N-[(2S, 3R) -4- (3-dimethylaminophenyl) methylamino-3-hydroxy-1-phenylbutan-2-yl] -5 Oxopyrrolidine-3-carboxamide;

(3S) -1-benzyl-N-{(2S, 3R) -3-hydroxy-1-phenyl-4-[(3-trifluoromethoxyphenyl) methylamino] butan-2-yl} -5- Oxopyrrolidine-3-carboxamide;

(4S) -N-[(2S, 3R) -4- (3-dimethylaminophenyl) methylamino-3-hydroxy-1-phenylbutan-2-yl] -N-ethyl-2-oxoimidazoli Dine-1,4-dicarboxamide;

(3S) -1-[(2S) -3- (4-chlorophenyl) -1-ethylamino-1-oxopropan-2-yl] -N-[(2S, 3R) -4- (3-dimethyl Aminophenyl) methylamino-3-hydroxy-1-phenylbutan-2-yl] -5-oxopyrrolidine-3-carboxamide;

(3R) -1-[(2S) -3- (4-chlorophenyl) -1-ethylamino-1-oxopropan-2-yl] -N-[(2S, 3R) -4- (3-dimethyl Aminophenyl) methylamino-3-hydroxy-1-phenylbutan-2-yl] -5-oxopyrrolidine-3-carboxamide;

(3S) -1-benzyl-N-{()-3-hydroxy-1-phenyl-4-[(3-trifluoromethoxyphenyl) methylamino] butan-2-yl} -2-oxoimida Zolidine-4-carboxamide;

(3S) -2,4-dibenzyl-N-[()-4- (3-dimethylaminophenyl) methylamino-3-hydroxy-1-phenylbutan-2-yl] -5-oxopyrrolidine -2-carboxamide;

(3R) -2,4-dibenzyl-N-[()-4- (3-dimethylaminophenyl) methylamino-3-hydroxy-1-phenylbutan-2-yl] -5-oxopyrrolidine -2-carboxamide;

(2S) -4-benzyl-N-[()-4- (3-dimethylaminophenyl) methylamino-3-hydroxy-1-phenylbutan-2-yl] -5-oxopyrrolidine-2- Carboxamides;

(4S) -1,4-dibenzyl-N-[()-4- (3-dimethylaminophenyl) methylamino-3-hydroxy-1-phenylbutan-2-yl] -2-oxoimidazoli Dine-4-carboxamide;

(4S) -N-[()-4- (3-dimethylaminophenyl) methylamino-3-hydroxy-1-phenylbutan-2-yl] -2-oxo-1-[(2,4,5 -Trifluorophenyl) methyl] imidazolidine-4-carboxamide;

(4S) -N-[()-4- (3-dimethylaminophenyl) methylamino-3-hydroxy-1-phenylbutan-2-yl] -2-oxo-1-[(4-trifluoro Methylphenyl) methyl] imidazolidine-4-carboxamide;

(4S) -N-[()-4- (3-dimethylaminophenyl) methylamino-3-hydroxy-1-phenylbutan-2-yl] -2-oxo-1-[(4-trifluorome Methoxyphenyl) methyl] imidazolidine-4-carboxamide;

(4S) -1-benzyl-N-[()-1- (3,5-difluorophenyl) -4- (3-dimethylaminophenyl) methylamino-3-hydroxybutan-2-yl]- 2-oxoimidazolidine-4-carboxamide;

(3S) -1-benzyl-N-[(2S, 3R) -1- (3,5-difluorophenyl) -4- (3-dimethylaminophenyl) methylamino-3-hydroxybutan-2- Il] -5-oxopyrrolidine-3-carboxamide;

(4S) -N-[(2S, 3R) -4- (3-dimethylaminophenyl) methylamino-3-hydroxy-1-phenylbutan-2-yl] -2-oxo-1- (2-phenoxy Cyethyl) imidazolidine-4-carboxamide;

(4R) -N-[(2S, 3R) -4- (3-dimethylaminophenyl) methylamino-3-hydroxy-1-phenylbutan-2-yl] -2-oxo-1- (2-phenoxy Cyethyl) imidazolidine-4-carboxamide;

(4S) -N-[(2S, 3R) -4- (3-dimethylaminophenyl) methylamino-3-hydroxy-1-phenylbutan-2-yl] -2-oxo-1-[(2- Trifluoromethylphenyl) methyl] imidazolidine-4-carboxamide;

(4S) -N-[(2S, 3R) -1- (3,5-difluorophenyl) -4- (3-dimethylaminophenyl) methylamino-3-hydroxybutan-2-yl] -2 -Oxo-1-[(2,4,5-trifluorophenyl) methyl] imidazolidine-4-carboxamide;

(4S) -1-benzyl-N-[(2S, 3R) -4- (cyclohexylamino) -3-hydroxy-1-phenylbutan-2-yl] -2-oxoimidazolidine-4- Carboxamides;

(4R) -1-benzyl-N-[(2S, 3R) -4- (3-dimethylaminophenyl) methylamino-3-hydroxy-1-phenylbutan-2-yl] -4-methyl-2- Oxoimidazolidine-4-carboxamide;

(4S) -1-benzyl-N-[(2S, 3R) -4- (3-dimethylaminophenyl) methylamino-3-hydroxy-1-phenylbutan-2-yl] -4-methyl-2- Oxoimidazolidine-4-carboxamide;

(4S) -N-[(2S, 3R) -4- (3-dimethylaminophenyl) methylamino-3-hydroxy-1-phenylbutan-2-yl] -1-[(4-fluorophenyl) Methyl] -2-oxoimidazolidine-4-carboxamide;

(4S) -1-[(3,4-difluorophenyl) methyl-N-[(2S, 3R) -4- (3-dimethylaminophenyl) methylamino-3-hydroxy-1-phenylbutane- 2-yl] -2-oxoimidazolidine-4-carboxamide;

(4S) -1-[(2,4-difluorophenyl) methyl-N-[(2S, 3R) -4- (3-dimethylaminophenyl) methylamino-3-hydroxy-1-phenylbutane- 2-yl] -2-oxoimidazolidine-4-carboxamide;

(4S) -1-benzyl-N-[(2S, 3R) -3-hydroxy-1-phenyl-4-[(3-trifluoromethylphenyl)] methylamino] butan-2-yl] -2- Oxoimidazolidine-4-carboxamide;

(4S) -1-benzyl-N-{(2S, 3R) -3-hydroxy-1-phenyl-4-[(3-propan-2-ylphenyl) methylamino] butan-2-yl} -2 Oxoimidazolidine-4-carboxamide;

(4S) -N-{(2S, 3R) -1- (3,5-difluorophenyl) -3-hydroxy-4-[(3-propan-2-ylphenyl) methylamino] butane-2 -Yl} -2-oxo-1-[(2,4,5-trifluorophenyl) methyl] imidazolidine-4-carboxamide;

(4S) -1- (3,5-dimethoxyphenyl) methyl-N-[(2S, 3R) -4- (3-dimethylaminophenyl) methylamino-3-hydroxy-1-phenylbutan-2- Il] -2-oxoimidazolidine-4-carboxamide;

(4S) -N-[(2S, 3R) -4- (3-ethylphenyl) methylamino-3-hydroxy-1-phenylbutan-2-yl] -2-oxo-1-[(2,4 , 5-trifluorophenyl) methyl] imidazolidine-4-carboxamide;

(4S) -N-[(2S, 3R) -4- (3-thibutylphenyl) methylamino-3-hydroxy-1-phenylbutan-2-yl] -2-oxo-1-[(2, 4,5-trifluorophenyl) methyl] imidazolidine-4-carboxamide;

(4S) -1-[(3,5-difluorophenyl) methyl-N-[(2S, 3R) -4- (3-dimethylaminophenyl) methylamino-3-hydroxy-1-phenylbutane- 2-yl] -2-oxoimidazolidine-4-carboxamide;

(4S) -N-[(2S, 3R) -4- (3-dimethylaminophenyl) methylamino-3-hydroxy-1-phenylbutan-2-yl] -1-[(3-methoxyphenyl) Methyl] -2-oxoimidazolidine-4-carboxamide;

(4S) -N-[(2S, 3R) -4- (3-dimethylaminophenyl) methylamino-3-hydroxy-1-phenylbutan-2-yl] -1-[(4-methoxyphenyl) Methyl] -2-oxoimidazolidine-4-carboxamide;

(4S) -N-[(2S, 3R) -4- (3-thibutylphenyl) methylamino-3-hydroxy-1-phenylbutan-2-yl] -1- (3,5-difluoro Phenyl) methyl-2-oxoimidazolidine-4-carboxamide;

(4S) -1- (3,5-difluorophenyl) methyl-N-{(2S, 3R) -3-hydroxy-1-phenyl-4-[(3-propan-2-ylphenyl) methyl Amino] butan-2-yl} -2-oxoimidazolidine-4-carboxamide;

(4S) -N-[(2S, 3R) -4- (3-thibutylphenyl) methylamino-1- (3,5-difluorophenyl) -3-hydroxybutan-2-yl] -1 -(3,5-difluorophenyl) methyl-2-oxoimidazolidine-4-carboxamide;

(4S) -N-((2S, 3R) -4-ethylamino-3-hydroxy-1-phenylbutan-2-yl) -2-oxo-1-[(2,4,5-trifluoro Phenyl) methyl] imidazolidine-4-carboxamide;

(4S) -1- (3,5-dibromo) phenyl-N-[(2S, 3R) -4- (3-dimethylaminophenyl) methylamino-3-hydroxy-1-phenylbutan-2- Il] -2-oxoimidazolidine-4-carboxamide;

(4S) -N-[(2S, 3R) -4- (3-dimethylaminophenyl) methylamino-3-hydroxy-1-phenylbutan-2-yl] -2-oxo-1-phenylimidazoli Dine-4-carboxamide;

(3S) -1-cyclohexylmethyl-N-[(2S, 3R) -4- (3-dimethylaminophenyl) methylamino-3-hydroxy-1-phenylbutan-2-yl] -5-oxopy Rollidine-3-carboxamide;

(3R) -1-cyclohexylmethyl-N-[(2S, 3R) -4- (3-dimethylaminophenyl) methylamino-3-hydroxy-1-phenylbutan-2-yl] -5-oxopy Rollidine-3-carboxamide;

(4S) -N-[(2S, 3R) -4- (3-thibutylphenyl) methylamino-3-hydroxy-1-phenylbutan-2-yl] -2-oxoimidazolidine-4- Carboxamides;

(4S) -N-[(2S, 3R) -4-[(3-dimethylaminophenyl) methylamino] -3-hydroxy-1-phenylbutan-2-yl] -5-oxo-1-pentylpy Rollidine-4-carboxamide;

(4R) -N-[(2S, 3R) -4-[(3-dimethylaminophenyl) methylamino] -3-hydroxy-1-phenylbutan-2-yl] -5-oxo-1-pentylpy Ralidin-4-carboxamide.

The present invention also relates to a method for preparing a compound of formula (I), and as shown in Scheme 1, the compound of formula (I) may be prepared by an amide coupling reaction of a compound of formula (2) with a compound of formula (3).

Scheme 1

In the above scheme, n, R ₁ , R ₂ , R ₃ , R ₄ , W, X, Y, and Z are as defined above. As a coupling agent used for an amidation reaction, a well-known coupling agent can be used, for example, dicyclohexyl carbodiimide (DCC), 1- (3-dimethylaminopropyl) -3-ethylcarbodiimide Carbodiimides such as (EDC) and 1,1'-dicarbonyldiimidazole (CDI) are mixed with 1-hydroxybenzotriazole (HOBT) or 1-hydroxy-7-azabenzotriazole (HOAT) Or bis- (2-oxo-3-oxazolidinyl) -phosphinic chloride (BOP-Cl), diphenylphosphoryl azide (DPPA), N- [dimethylamino-1H-1,2 , 3-triazole [4,5-b] pyridin-1-ylmethylene] -N-methylmethanealuminum (HATU) and the like, but are not limited thereto. In general, a solvent used in the amidation reaction may be DCM, DMF, DCE and the like, triethylamine, diisopropylethyl amine and the like can be used as a base for activation.

In the case of the compound of Formula 2, the compound of the simple lactam form is commercially available, and in the case of cyclic urea can be prepared in the same manner as in Scheme 2 below from a commercially available compound.

Scheme 2

Wherein R ₁ is as defined above and Z means cbz (benzyloxycarbonyl). The compound of Chemical Formula 4 is commercially available, and synthesizes an ester of Chemical Formula 5 using triphenyl phosphine (PPh ₃ ) and t-BuOH, and a compound substituted as in Chemical Formula 6 through alkylation or acylation reaction. After obtaining, the compound of Chemical Formula 2-1 may be prepared through deprotection reaction and hydrolysis under strong acid (HCl) conditions.

On the other hand, aryl cyclic substituted cyclic urea can be prepared by the method as shown in Scheme 3.

Scheme 3

Wherein R ₁ is as defined above. Protecting N from the serine with a trityl functional group to obtain a compound as shown in formula (8), a diamino compound as shown in formula (10) through an oxidation reaction and a reductive amination reaction, followed by cyclization reaction with triphosgene and desorption under strong acid The compound as shown in Chemical Formula 2-2 can be prepared through gelatinization.

Compounds of the lactam structure that are difficult to purchase commercially can be prepared by the same method as in Scheme 4 below.

Scheme 4

Wherein n = 0 or 1 and R ₁ Is as defined above. When the amine is added to the compound of Formula 12 to react, a cyclization reaction may occur at the same time to obtain a compound of Formula 13, and the hydrolysis method is the same as that of Compound 2-1.

In addition, a compound such as Chemical Formula 2-4 of Scheme 5 directly connected to N in the cyclic urea compound may be synthesized by a method similar to Scheme 5 below from a commercially available compound such as Chemical Formula 4.

Scheme 5

Wherein R represents an alkyl group, and Z represents cbz. First, a commercially available formula (4) may be obtained by amide linkage reaction to obtain a compound as shown in Formula 14, by introducing an acetyl group through an alkylation reaction, and then deprotecting the Z group under hydrogen conditions. You can get it. Hydrogen reaction can be carried out through hydrogen reaction using a Pd / C catalyst. In this case, methanol, ethanol, dioxane, etc. may be used as the solvent, and may be generally performed at 1 to 3 atmospheres.

 The compound of Formula 2-4 and the compound can be obtained by hydrolysis in the same manner as when obtaining a compound of the formula 2-1.

In the compound of Chemical Formula 3, when R ₃ is unsubstituted benzyl, it may be synthesized by the same method as in Chemical Scheme 6.

Scheme 6

Wherein R ₄ is as defined above and is commercially available in the case of erythro-N-BOC-L-phenylalanine epoxide, and the reaction is carried out by hydrogen reaction after introducing azide under saturated ammonium and alcohol solvent. The same amine compound can be obtained, and the compound of formula 3-1 can be prepared through a reducing amino reaction.

Reductive amination reactions are carried out using compounds containing carbonyl groups, for example ketones or aldehydes. Reducing agents that can be used include sodium borohydride, sodium cyanoborohydride, sodium triacetoxyborohydride and the like. In addition, an acid may be used as a catalyst to promote the reaction, and examples of the acid catalyst which may be used include inorganic acids such as hydrochloric acid, sulfuric acid, nitric acid and phosphoric acid; Organic carbon acids such as acetic acid, trifluoroacetic acid and the like; And amine salts such as ammonium chloride and the like, with hydrochloric acid or acetic acid being particularly preferred.

In addition, benzyl epoxide substituted with R ₃ may be synthesized through a general amino acid synthesis method as in Scheme 7.

Scheme 7

Wherein R ₃ is as defined above.

By using a chiral oxali as shown in the formula (20) to perform a stereoselective alkylation reaction, and through the hydrolysis can be prepared an amino acid derivative of the formula (22). Hydrolysis reaction to obtain a compound of formula 22 may be carried out using a base in a mixed solution of water and an organic solvent, wherein the organic solvent may be used THF, methanol, dioxane, etc., LiOH , KOH, NaOH and the like can be used. In addition, chloro iodo may be reacted with methane under a base to obtain a chlorocarbonyl compound represented by Chemical Formula 24, and an iodo hydrin compound represented by Chemical Formula 25 may be obtained through a reduction reaction. In the reduction reaction, NaBH ₄ , NaB (CN) H ₃ , and the like may be used. Further, an epoxide compound such as Chemical Formula 26 may be obtained through a cyclization reaction of the chlorohydrin compound under a base.

In the case of a compound of R ₄ , most commercially available compounds may be used, but for some compounds, bromide may be converted to an aldehyde in the same manner as in Scheme 8 below.

Scheme 8

Wherein R 'represents alkyl or aryl. The reaction may preferably be carried out in conventional solvents which do not adversely affect the reaction, particularly preferably the group consisting of DMF, dimethylacetamide, tetrahydrofuran, methylene chloride, dichloroethane, methanol, and water One or more solvents selected from may be used, but they are not limited thereto.

Separation of common mixtures can be carried out using column chromatography, and for final compounds can be separated by recrystallization or normal or reversed HPLC (Waters, Delta Pack, 300 × 50 mmI.D., C18 5 μm, 100 A). In the case of recrystallization or purification through HPLC, a compound can be obtained in the form of trifluoroacetic acid salt, and an ion exchange resin can be used in case of obtaining hydrochloride.

As mentioned above, after the reaction according to the method of the present invention is completed, the product can be separated and purified by conventional post-treatment methods such as chromatography, recrystallization and the like.

The present invention also relates to a pharmaceutical composition for inhibiting beta secretase, comprising a pharmaceutically effective amount of the compound of formula (I), a pharmaceutically acceptable salt or isomer thereof, and a pharmaceutically acceptable carrier as an active ingredient. In addition, the composition may additionally include a diluent or an excipient, as the case may be.

Since the compound of Formula I has an excellent inhibitory effect on beta secretase, it provides a composition of a beta secretase inhibitor comprising the same together with a pharmaceutically acceptable carrier. In particular, the composition according to the present invention exhibits an excellent effect on improving cognitive function or treating or preventing neurodegenerative diseases, and in particular, shows an excellent effect on preventing and treating Alzheimer's disease, but is not limited thereto.

The term 'pharmaceutical composition' means a mixture of a compound of the invention with other chemical components such as diluents or carriers. The pharmaceutical composition facilitates administration of the compound into the organism. There are a variety of techniques for administering compounds, including but not limited to oral, injection, aerosol, parenteral, and topical administration. Pharmaceutical compositions may also be obtained by reacting acid compounds such as hydrochloric acid, bromic acid, sulfuric acid, nitric acid, phosphoric acid, methanesulfonic acid, p-toluenesulfonic acid, salicylic acid and the like.

The term 'therapeutically effective amount' means that the amount of the compound administered relieves to some extent one or more symptoms of the disorder being treated. Thus, the pharmacologically effective amount may be selected from (1) the effect of reversing the rate of progression of the disease; (2) the effect of inhibiting or somewhat slowing further progression of the disease; And / or (3) an amount that has some effect of alleviating (preferably eliminating) one or more symptoms associated with the disease.

The term 'carrier' is defined as a compound that facilitates the addition of a compound into a cell or tissue. For example, dimethyl sulfoxide (DMSO) is a commonly used carrier that facilitates the incorporation of many organic compounds into organisms' cells or tissues.

The term 'diluent' is defined as a compound that not only stabilizes the biologically active form of the compound of interest, but also is diluted in water to dissolve the compound. Salts dissolved in buffer solutions are used as diluents in the art. A commonly used buffer solution is phosphate buffered saline, because it mimics the salt state of human solutions. Because buffer salts can control the pH of a solution at low concentrations, buffer diluents rarely modify the biological activity of a compound.

The term 'physiologically acceptable' is defined as a carrier or diluent that does not impair the biological activity and the properties of the compound.

The compounds used herein may be administered to human patients as such or as pharmaceutical compositions in combination with other active ingredients or with a suitable carrier or excipient, such as in a combination therapy. Techniques for formulation and administration of compounds in this application can be found in "Remington's Pharmaceutical Sciences," Mack Publishing Co., Easton, PA, 18th edition, 1990.

a) Route of administration

The compounds of the present invention can be administered by any route as desired. Injection, oral and nasal administration are preferred, but can also be administered through the skin, abdominal cavity, larynx and rectum.

b) Composition / Formulation

Pharmaceutical compositions of the invention can be prepared in a known manner, for example, by means of conventional mixing, dissolving, granulating, sugar-making, powdering, emulsifying, encapsulating, trapping and or lyophilizing processes. . Thus, pharmaceutical compositions for use according to the present invention comprise one or more pharmacologically acceptable compositions comprising excipients or auxiliaries which facilitate the treatment of the active compounds into formulations which can be used pharmaceutically. It may also be prepared by conventional methods using a carrier. Proper formulation is dependent upon the route of administration chosen. Any of the known techniques, carriers and excipients can be used suitably and as understood in the art, for example in Remingston's Pharmaceutical Sciences described above.

Injectable preparations, for example, sterile injectable aqueous or oleaginous suspensions may be prepared using suitable dispersing, wetting, or suspending agents according to known techniques. Solvents that can be used for this are water, Ringer's solution and isotonic NaCl solution, and sterile fixed oils are also commonly used as solvents or suspending media. Any non-irritating fixed oil may be used for this purpose, including mono- and diglycerides, and fatty acids such as oleic acid may also be used in the preparation of injectables.

Solid dosage forms for oral administration include capsules, tablets, pills, powders, and granules. Particularly, capsules and tablets are useful. Tablets and pills are preferably prepared with enteric agents. Solid dosage forms can be prepared by mixing the active compounds of formula (I) according to the invention with one or more inert diluents such as sucrose, lactose, starch and the like and carriers such as lubricants such as magnesium stearate, disintegrants, binders and the like. .

c) Effective amount

Pharmaceutical compositions suitable for use in the present invention include compositions in which the active ingredients are contained in an amount effective to achieve their intended purpose. More specifically, a therapeutically effective amount means an amount of a compound effective to prolong the survival of the subject to be treated or to prevent, alleviate or alleviate the symptoms of a disease. Determination of a therapeutically effective amount is within the capabilities of those skilled in the art, in particular in terms of the detailed disclosure provided herein.

The total daily dose to be administered to the host in a single dose or in separate doses when administering a compound of the present invention for clinical purposes is preferably in the range of 10 to 100 mg / kg body weight, but specific dose levels for individual patients may be used. It may vary depending on the specific compound, the weight of the patient, sex, health condition, diet, the time of administration of the drug, the method of administration, the rate of excretion, the mixing of the drug and the severity of the disease.

The invention is explained in more detail by the following preparations and examples, although the scope of the invention is not limited in any way by them.

Description of the abbreviations and terms used in the reaction schemes, the following Preparation Examples and Examples in the name of the compound is shown in Table 1 below.

TABLE 1

The preparation of the reactants (intermediates) required for the synthesis of the example compounds according to the invention in the preparation examples below will be described in more detail.

Production Example  1: (S) -1-benzyl-2-oxo- Imidazolidine -4-carboxylic acid

Step A: (S) -2-oxo- Imidazolidine -1,5- Dicarboxylic acid  One- Benzyl esters  5-t- Butyl ester

Commercially available (S) -2-oxo-imidazolidine-1,5-dicarboxylic acid 1-benzyl ester (2.64 g, 10 mmol) was dissolved in THF (20 ml) and then PPh ₃ (2.62 g, 10 mmol) was added and t-BuOH (81 mg, 11 mmol) was added dropwise and stirred at room temperature for 3 hours. After completion of the reaction, 1N HCl was added dropwise, an organic layer was obtained with EtOAc, washed with water and brine, dried over MgSO _4, and concentrated under reduced pressure. The residue was purified by column chromatography (eluent: EtOAc: n-Hex = 1/4). Purification) yielded the title compound (2.94 g, 92%).

MS [M + 1] = 321 (M + 1)

Step B: (S) -3-benzyl-2-oxo- Imidazolidine -1,5- Dicarboxylic acid  One- Benzyl esters , 5-t- Butyl ester

(S) -2-oxo-imidazolidine-1,5-dicarboxylic acid 1-benzylester 5-t-butylester (2.24 g, 7 mmol) obtained in step A was dissolved in THF, followed by NaH (392). mg, 60% mineral oil, 8.4 mmol) was added dropwise at 0 ° C., and after 30 minutes benzyl chloride (970 mg, 7.7 mmol) was added dropwise. After the reaction solution was stirred at room temperature for 4 hours, saturated NH ₄ Cl aqueous solution was added and the organic material was extracted with EtOAc. The extracted organics were dried over MgSO ₄ and concentrated under reduced pressure. The residue was purified by column chromatography (eluent: EtOAc: n-Hex = 1/4) to give the title compound (2.43 g, 85%).

MS [M + 1] = 411 (M + 1)

Step C: (S) -1-benzyl-2-oxo- Imidazolidine --4-carboxylic acid

(S) -3-benzyl-2-oxo-imidazolidine-1,5-dicarboxylic acid 1-benzylester, 5-t-butylester (2.43 g, 5.93 mmol) obtained in step B was dissolved in 4N HCl. It was dissolved in an aqueous solution and stirred at room temperature for 3 hours. After the reaction was terminated, the mixture was neutralized with a saturated NaHCO ₃ aqueous solution, and then extracted with EtOAc, and crystallized to give the title compound (1.14 g, 87%).

H ¹ NMR (CDCl ₃ ) δ 7.37-7.27 (m, 5H), 4.43-4.30 (m, 2H), 4.23-4.21 (m, 1H), 3.62 (t, 1H, J = 8.0 Hz), 3.46-3.37 (m , 1H),

MS [M + 1] = 221 (M + 1)

Production Example  2 to 13

Preparation was carried out in the same manner as in Preparation Example 1 using a commercially available halide derivative and (S) -2-oxo-imidazolidine-1,5-dicarboxylic acid 1-benzyl ester, as shown in Table 2 below. The compounds of Examples 2 to 13 were synthesized.

(III)

(III)

TABLE 2

Production Example  14: (S) -1- Butyryl 2-oxo- Imidazolidine -4-carboxylic acid

Step A: (S) -3- Butyryl 2-oxo- Imidazolidine -1,5- Dicarboxylic acid  One- Benzyl esters  5-t- Butyl ester

(S) -2-oxo-imidazolidine-1,5-dicarboxylic acid 1-benzyl ester 5-t-butyl ester (620 mg, 2 mmol) obtained in step A of Preparation Example 1 was dissolved in DCM. , Et ₃ N (560 ul, 4 mmol) was added dropwise and propyl acyl chloride (297 mg, 2.8 mmol) was added thereto, followed by stirring at room temperature for 2 hours. After completion of the reaction, saturated aqueous NH ₄ Cl solution was added and the organic material was extracted with EtOAc. The extracted organic solution was dried over MgSO ₄ and concentrated under reduced pressure. The residue was purified by column chromatography (eluent: EtOAc: n-Hex = 1/5) to give the title compound (725 mg, 93%).

MS [M + 1] = 391 (M + 1)

Step B: (S) -1- Butyryl 2-oxo- Imidazolidine -4-carboxylic acid

Step of Preparation Example 1 using t-butyl (S) -3-butyryl-2-oxo-imidazolidine-1,5-dicarboxylic acid 1-benzylester 5-t-butylester obtained in step A By the same method as C, the title compound is obtained.

MS [M + 1] = 201 (M + 1)

Production Example  15: (S) -1- Ethylcarbamoyl 2-oxo- Imidazolidine -4-carboxylic acid

Preparation Example Using (S) -2-oxo-imidazolidine-1,5-dicarboxylic acid 1-benzyl ester 5-t-butylester and ethyl isocyanate obtained in Step A of Preparation Example 1 By the same method as 14, the title compound was obtained.

MS [M + 1] = 202 (M + 1)

Preparation Example 16; (S) -2-oxo-1- phenyl - imidazolidine- 4-carboxylic acid

Step A: (S) -3-oxo-2- ( Tritylamino ) -Propionic acid t- Butyl ester

Commercially available Tr-N-Ser-O (t-Bu) (2.02 g, 5 mmol) was dissolved in DCM and DMSO solution, Et ₃ N (2.8 ml, 20 mmol) and SO ₃ Py (1.59 g, 10 mmol) ) Was added and the reaction solution was stirred at room temperature for 2 hours. After the reaction was completed, DCM was distilled off under reduced pressure, saturated aqueous NH ₄ Cl solution was added, and the organic material was extracted with ethyl ester. The extracted organic solution was washed with water, dried over MgSO ₄ and concentrated under reduced pressure. The residue was purified by column chromatography (eluent: EtOAc: n-Hex = 1/2) to obtain the title compound (1.9 g, 95%). It was.

MS [M + 1] = 402 (M + 1)

Step B: (S) -3- Phenylamino -2- Tritylamino Propionic acid t Butyl ester

(S) -3-oxo-2- (tritylamino) -propionic acid t-butylester (1.5 g, 3.74 mmol) and aniline (465 mg, 5 mmol) obtained in step A were dissolved in dichloroethane, followed by NaBH ( OAc) ₃ (1.72 g, 7.48 mmol) was added and the reaction solution was stirred at room temperature for 4 hours. After the reaction was completed, the reaction solution was diluted with saturated NaHCO ₃ aqueous solution, and the organic material was extracted with DCM and EtOAc. The extracted organic solution was washed with brine, dried over MgSO ₄ , and concentrated under reduced pressure. The residue was purified by column chromatography (eluent: EtOAc: n-Hex = 1/2) to obtain the title compound (2.07 g, 87%). It was.

MS [M + 1] = 479 (M + 1)

Step C: (S) -2-oxo-1- Phenyl 3-tree Imidazolidine -4-carboxylic acid t- Butyl ester

(S) -3-phenylamino-2-tritylamino--propionic acid t-butylester (2.0 g, 4.18 mmol) obtained in step B was dissolved in DCM (10 ml) and phosgene (toluene solution, 2M, 4 ml) was added and stirred at room temperature for 4 hours. After completion of the reaction, the solvent was distilled off under reduced pressure and the residue was purified by column chromatography (eluent: EtOAc: n-Hex = 1/3) to give the title compound (1.09 g, 54%).

MS [M + 1] = 521 (M + 1)

Step D: (S) -2-oxo-1- phenyl - imidazolidine- 4-carboxylic acid

Obtaining the title compound in the same manner as in Step C of Preparation Example 1 using (S) -2-oxo-1-phenyl-3-tri-imidazolidine-4-carboxylic acid t-butylester obtained in step C. It was.

MS [M + 1] = 223 (M + 1)

Production Example  17: (S) -1- (3,5- Dibromophenyl ) -2-oxo- Imidazolidine -4-carboxylic acid

(S) -3-oxo-2- (tritylamino) -propionic acid t-butylester and 3,5-dibromo aniline obtained in step A of Preparation Example 16 were carried out in the same manner as in Preparation Example 16 This gave the title compound.

MS [M + 1] = 381 (M + 1)

Production Example  18: ((S) -4- Dimethyl carbamoyl 2-oxo- Imidazolidine -1-yl) acetic acid

Step A: (S) -5- Dimethyl carbamoyl 2-oxo- Imidazolidine -1-carboxylic acid Benzyl esters

A commercially available (S) -2-oxo-imidazolidine-1,5-dicarboxylic acid 1-benzyl ester (2. 64 g, 10 mmol) was dissolved in DHF (20 ml) and then EDC (2.33 g, 13 mmol) and HOBT (2.30 g, 15 mmol) were added and Et ₃ N (2.8 ml, 20 mmol) was added. After confirming that the pH of the solution was basic, dimethylamine hydrochloride (1.2 g, 15 mmol) was added, and then the pH was again measured to confirm the basicity of the solution, followed by stirring at room temperature for 12 hours. After completion of the reaction, saturated DMF was distilled off under reduced pressure, and the residue was diluted with EtOAc, washed with saturated aqueous NaHCO ₃ solution, 1N HCl, and water, and the filtrate was dried over MgSO ₄ , concentrated under reduced pressure, and the residue was purified by column chromatography. Purification with (eluent: EtOAc: n-Hex = 1/1) gave the title compound (2.50 g, 87%).

MS [M + 1] = 392 (M + 1)

Step B: (S) -3-t- Butoxycarbonylmethyl -5- Dimethyl carbamoyl 2-oxo- Imidazolidine -1-carboxylic acid benzyl ester

(S) -5-dimethylcarbamoyl-2-oxo-imidazolidine-1-carboxylic acid benzyl ester (2.0 g, 6.87 mmol) obtained in step A was dissolved in THF (20 ml) and the temperature of the reaction solution was 0. After lowering to < 0 > C, NaH was added and stirred for about 30 minutes. t-butylbromoacetate was dissolved in THF and added slowly, and the reaction solution was raised to room temperature and stirred for 4 hours. After completion of the reaction, saturated aqueous NH ₄ Cl solution was added, the organics were extracted with EtOAc, dried over MgSO ₄ and concentrated under reduced pressure. The residue was purified by column chromatography (eluent: EtOAc: n-Hex = 1/3). The title compound (2.50 g, 87%) was obtained.

MS [M + 1] = 406 (M + 1)

Step C: ((S) -4- Dimethyl carbamoyl 2-oxo- Imidazolidine -1-yl) acetic acid

In step C of Preparation Example 1 using (S) -3-t-butoxycarbonylmethyl-5-dimethylcarbamoyl-2-oxo-imidazolidine-1-carboxylic acid benzyl ester obtained in step B The same procedure was followed to yield the title compound.

MS [M + 1] = 216 (M + 1)

Production Example  19: 1-butyl-5-oxo- Pyrrolidine -3-carboxylic acid

Step A: 1-Butyl-5-oxo- Pyrrolidine -3-carboxylic acid Methyl ester

Dimethyl itaconate (1.58 g, 10 mmol) was dissolved in THF (30 ml), n-butyl amine (870 mg, 10 mmol) was added, and the reaction solution was stirred for 12 hours while heating to 50 ° C. After completion of the reaction, saturated aqueous NH ₄ Cl solution was added, the organics were extracted with EtOAc, dried over MgSO ₄ , concentrated under reduced pressure, and the residue was purified by column chromatography (eluent: EtOAc: n-Hex = 1/1). Purification gave the title compound (1.84 g, 80%).

MS [M + 1] = 246 (M + 1)

Step B: 1-Butyl-5-oxo- Pyrrolidine -3-carboxylic acid

Dissolve 1-butyl-5-oxo-pyrrolidine-3-carboxylic acid methyl ester (1 g, 4. 6 mmol) obtained in step A in MeOH (1 ml), add water (5 ml), and then add LiOH. (529 mg, 23 mmol) was added and the reaction solution was stirred at room temperature for 4 hours. After the reaction was completed, MeOH was removed by distillation under reduced pressure, the pH was adjusted to 2-3 with 1N HCl, and the organics were extracted with EtOAc to obtain the title compound (851 mg, 93%).

MS [M + 1] = 200 (M + 1)

Production Example  20 to 26

By performing the same method as Preparation Example 19 using a commercially available amine compound and dimethyl itaconate, the compounds of Preparation Examples 20 to 26 as shown in Table 3 were synthesized.

(IV)

(IV)

TABLE 3

Production Example  27: 1-benzyl-4- methyl 2-oxo- Imidazolidine -4-carboxylic acid

Step A: 3-benzyl-5- methyl 2-oxo- Imidazolidine -1,5- Dicarboxylic acid  One- Benzyl esters  5-t- Butyl ester

(S) -3-benzyl-2-oxo-imidazolidine-1,5-dicarboxylic acid 1-benzyl ester, 5-t-butyl ester (410 mg, 1 mmol) obtained in step B of Preparation Example 1 ) Was dissolved in THF (5 ml) and the reaction solution was cooled to -78 ° C. Then, LiHMDS (1N THF, 1.4 ml, 1.4 mmol) solution was added. After 10 minutes, methyl iodide (135 mg, 1 mmol) was added. Was added. The reaction solution was stirred at -78 ° C for 1 hour, and the reaction was terminated by addition of saturated aqueous NH ₄ Cl solution. The organics were extracted with EtOAc, dried over MgSO ₄ , concentrated under reduced pressure, and the residue was purified by column chromatography (eluent). : EtOAc: n-Hex = 1/2) to give the title compound (339 mg, 80%).

MS [M + 1] = 425 (M + 1)

Step B: 1-benzyl-4- methyl 2-oxo- Imidazolidine -4-carboxylic acid

Method as in Step C of Preparation Example 1 using 3-benzyl-5-methyl-2-oxo-imidazolidine-1,5-dicarboxylic acid 1-benzylester 5-t-butylester prepared in Step A To give the title compound.

MS [M + 1] = 235 (M + 1)

Production Example  28: 1,4- Dibenzyl 2-oxo- Imidazolidine -4-carboxylic acid

Using (S) -3-benzyl-2-oxo-imidazolidine-1,5-dicarboxylic acid 1-benzyl ester, 5-t-butyl ester and benzyl chloride obtained in step B of Preparation Example 1 The title compound was obtained in the same manner as in Preparation Example 27.

MS [M + 1] = 311 (M + 1)

Production Example  29: (1- methyl -5-oxo- Pyrrolidine -3-yl) -acetic acid

Step A: 3- Methanesulfonyl jade - Pentandioic  Acid Dimethyl Ester

The commercially available methyl 3-hydroxy-pentanedioic acid dimethyl ester (3.5 g, 20 mmol) was dissolved in DCM (50 ml), Et ₃ N (5.6 ml, 40 mmol) was added and the temperature of the reaction solution was increased. After lowering to 0 ° C., MsCl (2.75 g, 24 mmol) was added dropwise and stirred at room temperature for 2 hours. After the reaction was completed, NH ₄ Cl was added and EtOAc The organics were extracted, dried over MgSO ₄ , and concentrated under reduced pressure. The residue was purified by column chromatography (eluent: EtOAc: n-Hex = 1/5) to give the title compound (4.77 g, 94%).

MS [M + 1] = 241 (M + 1)

Step B: 3- Nitromethyl - Pentandioic  Acid Dimethyl Ester

3-Methanesulfonylocta-pentanedioic acid dimethyl ester (4.0 g, 15.7 mmol) obtained in step A was dissolved in DMF (950 ml), DMAP (9200 mg, 1.57 mmol) was added, followed by nitromethane (4.71 g). , 78.5 mmol) was added and stirred at 80 ° C. for 12 hours. After the reaction was complete, NH ₄ Cl was added and EtOAc The organics were extracted, dried over MgSO ₄ and concentrated under reduced pressure. The residue was recrystallized with EtOAc / n-Hex to give the title compound (2.78 g, 81%). MS [M + 1] = (M + 1)

Step C: 3- Aminomethyl - Pentandioic  Acid Dimethyl Ester

The 3-nitromethyl-pentanedioic acid dimethyl ester (2.19 g, 10 mmol) obtained in step B was dissolved in MeOH (20 ml), Pd / C (200 mg) was added, followed by reaction in a hydrogen reactor for 12 hours. It was. After the reaction was completed, the celite filter was used and then used in the next reaction without further purification.

MS [M + 1] = 176 (M)

Step D: (5-Oxo- Pyrrolidine -3-yl) -acetic acid Methyl ester

The title compound was obtained in the same manner as A of Preparation Example 19 using 3-aminomethyl-pentanedioic acid dimethylester obtained in step C.

MS [M + 1] = 144 (M + 1)

Step E: (1- methyl -5-oxo- Pyrrolidine -3 days) Methyl acetate

Using the (5-oxo-pyrrolidin-3-yl) -acetic acid methyl ester and methyl iodide obtained in step D in the same manner as in step B of Preparation Example 18, the title compound was obtained.

MS [M + 1] = 158 (M + 1)

Step F: (1- methyl -5-oxo- Pyrrolidine -3-yl) -acetic acid

(1-Methyl-5-oxo-pyrrolidin-3-yl) acetic acid methyl ester obtained in step E was carried out in the same manner as in Step C of Preparation Example 19 to obtain the title compound.

MS [M + 1] = 144 (M + 1)

Production Example  30: (1-benzyl-5-oxo- Pyrrolidine -3-yl) -acetic acid

Steps E and F of Preparation Example 22 were carried out using (5-oxo-pyrrolidin-3-yl) -acetic acid methylester and benzyl chloride obtained in Step D of Preparation Example 29 to obtain the title compound.

MS [M + 1] = 220 (M + 1)

Production Example  31: (S) -4-benzyl-5-oxo- Pyrrolidine 2-carboxylic acid

Step A: (S) -2-benzyl-4-t- Butoxycarbonylamino - Pentandioic  Acid Dimethyl Ester

The title compound was obtained in the same manner as in Step A of Preparation Example 27 using commercially available NBOC-ASP (OMe) -OMe and benzyl bromide.

MS [M + 1] = 366 (M + 1)

Step B: (S) -2-Amino-4-benzyl- Pentanedioic acid Methyl ester

(S) -2-benzyl-4-t-butoxycarbonylamino-pentanedioic acid dimethyl ester obtained in step A was dissolved in DCM, TFA was added and stirred at room temperature for 2 hours. After the reaction was completed, the solvent was removed by distillation under reduced pressure, diluted with DCM, and then removed under reduced pressure. The residue was taken up again in DCM and crystallized using diethylester to afford the title compound.

MS [M + 1] = 266 (M + 1)

Step C: (S) -4-benzyl-5-oxo- Pyrrolidine 2-carboxylic acid Methyl ester

Using the (S) -2-amino-4-benzyl-pentanedioic acid methyl ester obtained in step B in the same manner as in B of Preparation Example 19, the title compound was obtained.

MS [M + 1] = 235 (M + 1)

Step D: (S) -4-benzyl-5-oxo- Pyrrolidine 2-carboxylic acid

(S) -4-benzyl-5-oxo-pyrrolidine-2-carboxylic acid methyl ester obtained in step C was carried out in the same manner as C of Preparation Example 19 to obtain the title compound. MS [M + 1] = 221 (M + 1)

Production Example  32: (R) -2,4- Dibenzyl -5-oxo- Pyrrolidine 2-carboxylic acid

Dialkylation compound was obtained in the course of compound purification in step A of preparation example 31, which was carried out in the same manner as steps B, C, and D of preparation example 31 to obtain the title compound.

MS [M + 1] = 310 (M + 1)

Production Example  33: ((1S, 2R) -3-Amino-1-benzyl-2- Hydroxy -profile) Carbamic  Acid t- Butyl ester

Step A: ((1S, 2R) -3- A map -1-benzyl-2- Hydroxy -profile) Carbamic  Acid t- Butyl ester

(2S, 3S) -erythro-N-BOC-L-phenylalanine epoxide (10.4 g, 50 mmol) was dissolved in ethanol (150 ml) and saturated aqueous NH ₄ Cl (40 ml) solution was added. When the solution became homogeneous, NaN ₃ (9.83 g, 150 mmol) was added and stirred at 80 ° C. for 12 hours. After completion of the reaction, the organics were extracted with EtOAc, dried over MgSO ₄ and concentrated under reduced pressure. The residue was purified by column chromatography (eluent: EtOAc: n-Hex = 1/4) to give the title compound (13.2 g, 86). %) Was obtained.

MS [M + 1] = 207 (M + 1)

Step B: ((1S, 2R) -3-Amino-1-benzyl-2- Hydroxy -profile) Carbamic  Acid t- Butyl ester

In step A, ((1S, 2R) -3-azido-1-benzyl-2-hydroxy-propyl) carbamic acid t-butylester was carried out in the same manner as in Step C of Preparation Example 29 to give the title The compound was obtained.

MS [M + 1] = 181 (M + 1)

Production Example  34: ((1S, 2R) -3-Amino-1- (3,5- Difluorobenzyl )-2- Hydroxy -profile) Carbamic  Acid t- Butyl ester

Step A: (2S, 5R) -2- (3,5- Difluorobenzyl ) -5-isopropyl-3,6- Dimethoxy -2,5- Dihydropyrazine

(R) -2,5-dihydro-3,6-dimethoxy-2-isopropylpyrazine was dissolved in THF, n-BuLi was added at -78 ° C, followed by difluorobenzyl chloride. After the reaction was completed, NH ₄ Cl was added and the mixture was added with EtOAc. The organic material was extracted, dried over MgSO ₄ and concentrated under reduced pressure. The residue was purified by column chromatography (eluent: EtOAc: n-Hex = 1/1) to give the title compound (339 mg, 80%).

MS [M + 1] = 313 (M + 1)

Step B: (S) -2-amino-3- (3,5- Difluorophenyl Propionic acid Methyl ester

(2S, 5R) -2- (3,5-difluorobenzyl) -5-isopropyl-3,6-dimethoxy-2,5-dihydropyrazine obtained in step A was dissolved in acetonitrile and 2N After adding HCl, the mixture was stirred at room temperature for 4 hours. After the reaction was complete, NaHCO _{3 was} added and EtOAc The organic material was extracted, dried over MgSO ₄ and concentrated under reduced pressure to give the title compound (339 mg, 80%).

MS [M + 1] = 215 (M + 1)

Step C: (S) -2-t- Butoxycarbonylamino -3- (3,5- Difluorophenyl Propionic acid methyl  ester

(S) -2-amino-3- (3,5-difluorophenyl) propionic acid methyl ester obtained in step B was dissolved in water and NaOH was added followed by (BOC) ₂ C. After 12 h the reaction was terminated with 1N HCl and EtOAc The organic material was extracted, dried over MgSO ₄ , and concentrated under reduced pressure. The residue was purified by column chromatography (eluent: EtOAc: n-Hex = 1/8) to obtain the title compound (339 mg, 80%).

MS [M + 1] = 315 (M + 1)

Step D: [(3S) -1- (3,5- Difluorobenzyl ) -3- Chloro -2-oxo-propyl]- Carbamic acid  t- Butyl ester

Chloroiodo methane was dissolved in THF and LDA was added, followed by addition of (S) -2-t-butoxycarbonylamino-3- (3,5-difluorophenyl) propionic acid methyl ester obtained in step C. And stirred at room temperature for 12 hours. After completion of the reaction, NH ₄ Cl was added, the organics were extracted with EtOAc, dried over MgSO ₄ and concentrated under reduced pressure. The residue was purified by column chromatography (eluent: EtOAc: n-Hex = 1/5). To give the title compound (339 mg, 80%).

MS [M + 1] = 426 (M + 1)

Step E: [(1S, 2S) -1- (3,5- Difluoro  Benzyl) -2- Hydroxy -3- Chloro -profile] Carbamic  Acid t- Butyl ester

[(3S) -1- (3,5-Difluorobenzyl) -3-chloro-2-oxo-propyl] -carbamic acid t-butylester obtained in step D was dissolved in MeOH and NaBH ₄ was added thereto. Stir at room temperature for 12 hours. After the reaction was completed, water was added, 1N HCl was added, stirred for 30 minutes, an organic material was extracted with EtOAc, dried over MgSO ₄ , and concentrated under reduced pressure. The residue was purified by column chromatography (eluent: EtOAc: n-Hex = 1/3) gave the title compound (339 mg, 80%).

MS [M + 1] = 428 (M + 1)

Step F: [(S) -2- (3,5- Difluoro - Phenyl ) -1- (S)- Oxiranyl -ethyl]- Carbamic acid  t- Butyl ester

Dissolve [(1S, 2S) -1- (3,5-difluorobenzyl) -2-hydroxy-3-chloro-propyl] carbamic acid t-butylester obtained in step E in THF and add NaOH Stir at room temperature for 12 hours. After completion of the reaction, add 1N HCl and add EtOAc. The organic material was extracted, dried over MgSO ₄ and concentrated under reduced pressure. The residue was purified by column chromatography (eluent: EtOAc: n-Hex = 1/1) to give the title compound (339 mg, 80%).

MS [M + 1] = 300 (M + 1)

Step G: ((1S, 2R) -3-Amino-1- (3,5- Difluorobenzyl )-2- Hydroxy -profile) Carbamic  Acid t- Butyl ester

The title compound was obtained by the same method as A and B of Preparation Example 33 using (2R, 3S) -erythro-N-BOC-L- (3,5-difluoro) phenylalanine epoxide obtained in step F. Obtained.

MS [M + 1] = 217 (M + 1)

Production Example  35: 3-dimethylamino Benzaldehyde

The title compound was obtained in the same manner as Step A of Preparation Example 16 using 3-dimethylamino benz alcohol.

MS [M + 1] = 150 (M + 1)

Production Example  36: 3-t-butyl Benzaldehyde

3-t-butyl phenylbromide was dissolved in THF, n-BuLi was added, and then DMF was added and stirred at 0 ° C. for 3 hours. After the reaction was completed, NH ₄ Cl was added and the mixture was added with EtOAc. The organic material was extracted, dried over MgSO ₄ and concentrated under reduced pressure. The residue was purified by column chromatography (eluent: EtOAc: n-Hex = 1/3) to give the title compound (339 mg, 80%).

MS [M + 1] = 163 (M + 1)

Production Example  37: [(1S, 2R) -1-Benzyl-3- (3-dimethylamino- Benzylamino ) -2-hydroxy-propyl]- Carbamic  Acid t-butyl ester

((1S, 2R) -3-Amino-1-benzyl-2-hydroxy-propyl) carbamic acid t-butyl ester and 3-dimethylaminobenzaldehyde were carried out in the same manner as in Step B of Preparation Example 16 This gave the title compound.

MS [M + 1] = 414 (M + 1)

Production Example 38 to 50

Using the amine compound synthesized in Preparation Examples 33 and 34 and the aldehyde synthesized in Preparation Examples 35 and 36 or the commercially available aldehyde selectively, Table 4 represented by the following formula The compounds according to were obtained.

TABLE 4

Example  1: (2S) -N-[(2S, 3R) -4-[(3- Dimethylaminophenyl ) Methylamino ] -3-hydroxy-1- Phenylbutane -2- day] -5- Oxopyrrolidine -2- Carboxamide

Step A: (2R, 3S) -3-Amino-1- (3-dimethylamino- Benzylamino )-4- Phenyl Butan-2-ol

[(1S, 2R) -1-Benzyl-3- (3-dimethylamino-benzylamino) -2-hydroxy-propyl] -carbamic acid t-butyl ester obtained in Preparation Example 37 (414 mg, 1 mmol) ) Was dissolved in dichloromethane (5 ml) and TFA (2 ml) was added, followed by stirring at room temperature for 2 hours. At the end of the reaction, the solvent was removed under reduced pressure and dried to use for the next reaction without further purification.

Mass [M + 1] = 314 (M + H)

Step B: (2S) -N-[(2S, 3R) -4-[(3- Dimethylaminophenyl ) Methylamino ] -3-hydroxy-1- Phenylbutane -2- day] -5- Oxopyrrolidine -2- Carboxamide

Commercially available (S) -5-oxo-proline (129 mg, 1 mmol) was dissolved in DMF (5 ml) and Et ₃ N (280 uM, 2 mmol) was added. HOBT (229.5 mg, 1.5 mmol) and EDC (232 mg, 1.3 mmol) were added and stirred at room temperature for 30 minutes, followed by (2R, 3S) -3-amino-1- (3-dimethylamino -Benzylamino) -4-phenyl-butan-2-ol (313 mg, 1 mmol) was added. After completion of the reaction, the solvent was removed under reduced pressure and saturated NaHCO ₃ After diluting with aqueous solution, the organic layer was extracted with EtOAc. The extracted organic solution was washed with brine, dried over MgSO ₄ , distilled under reduced pressure, and the residue was purified by Prep-TLC (10% MeOH / 90% DCM) to obtain the title compound (305 mg, 72%).

¹ H NMR (400 MHz, CDCl ₃ ) δ 7.26-7.14 (m, 6H), 6.72-6.64 (m, 3H), 4.23-4.19 (m, 1H), 3.96-3.92 (m, 1H), 3.87-3.76 ( m, 3H), 3.20-3.10 (m, 1H), 2.94 (s, 6H), 2.87-2.77 (m, 3H), 2.23-1.85 (m, 3H), 1.51-1.46 (m, 1H)

Mass [M + 1] = 425 (M + H)

Example  2: (2R) -N-[(2R, 3S) -4- (3- Dimethylaminophenyl ) Methylamino -3-hydroxy-1- Phenyl butane -2- day] -5- Oxopyrrolidine - Carboxamide

(R) -5-oxo-proline commercially available and (2R, 3S) -3-amino-1- (3-dimethylamino-benzylamino) -4-phenyl-butane obtained in step A of Example 1 Using the 2-ol, the same procedure as in Example 1 was carried out to obtain the title compound.

¹ H NMR (400 MHz, CDCl ₃ ) δ 7.26-7.14 (m, 6H), 6.72-6.64 (m, 3H), 4.23-4.19 (m, 1H), 3.96-3.92 (m, 1H), 3.87-3.76 ( m, 3H), 3.20-3.10 (m, 1H), 2.94 (s, 6H), 2.87-2.77 (m, 3H), 2.23-1.85 (m, 3H), 1.51-1.46 (m, 1H)

Mass [M + 1] = 425 (M + H)

Example  3: (4S) -1-benzyl-N- [1- (3,5- Difluorophenyl ) -4- (3- Dimethylaminophenyl ) Methylamino -3- Hydroxybutane -2- day] -2- Oxoimidazolidine -4- Carboxamide

[(1S, 2R) -1- (3,5-Diflo-benzyl) -3- (3-dimethylamino-benzylamino) -2-hydroxy-propyl] -carbamic acid t obtained in Preparation Example 50 The title compound was obtained in the same manner as in Example 1 using -butyl ester.

H ¹ NMR (CDCl ₃ ) δ 8.13 (brs, 1H), 7.28-7.14 (m, 6H), 6.79-6.72 (m, 4H), 6.67-6.65 (m, 1H), 6.58 (t, 1H, J = 8.0Hz ), 4.58 (d, 1H, J = 16 Hz), 4.27-3.90 (m, 3H), 3.77 (brs, 2H), 3.47-3.41 (m, 1H), 2.89 (s, 6H), 2.85-2.67 (m , 2H), 2.53 (brs, 1H), 2.30 (brs, 1H)

Mass [M + 1] = 516 (M + H)

Example 4 to 60

By using the acid synthesized in Preparation Examples 1 to 30 and the amine compound obtained in Preparation Examples 33 and 34 selectively, it was carried out in the same manner as in Example 3 to synthesize the following Example compounds.

(I)

(I)

TABLE 5

이고, (ii)는

이며, (iii)은

이고, (iv)는

이며; *는 이성질체의 형태를 의미한다) (In Table 5, (i) of Ar is

And (ii) is

(Iii) is

And (iv) is

Is; * Stands for isomeric form)

Experimental Example  1: recombinant beta Secretase 2  Enzyme Activity Measurement

Step A: Recombinant Beta Secretase 2  Preparation of Expression Vectors

CDNA (ATCC, Cat #: 6896840) based on the human BACE2 gene sequence (accession #: BC014453) known from Genebank, an existing public data base, was purchased. Cloning the portion corresponding to amino acid sequence 1-466 except the trnsmembrane domain and the cytoplasmic domain, ectodomain only, of the entire BACE gene, human immunoglobulin G ( The Fc partial nucleotide sequence corresponding to 230 amino acids (hIgG) (amino acid sequences 1-466) was attached. BACE2-Fc protein expression vector pCDNA3 BACE2 Fc was prepared by conjugating the BACE (ectodomain) -IgG Fc (hereinafter referred to as BACE-Fc) between BamHI and XhoI of a mammalian expression vector pCDNA3 (Invitrogen).

Step B: BACE2 - Fc  Fusion Protein Expression Mammalian Cell Line Preparation

A-MEM (a-minimum essential medium) (GIBCO) containing 10% fetal bovine serum (FBS) (GIBCO-BRL) containing Chinese hamster ovary (CHO) DHFR-cells (ATCC Accession # CRL9096) -BRL) culture was then transferred to a 100 mm culture plate, cells were grown to cover the bottom of the culture vessel, and then transformed into the BACE2-Fc protein expression vector pCDNA3 BACE2-Fc using Lipofectamine Plus (Life Technologies). Transformed cells were replaced every 4 days with 10% dialyzed fetal bovine serum (dFBS) (JRH) culture containing 1 mg / ml Geneticin (G418 sulfate) (GIBCO-BRL). Selective culture was carried out, and about 100 clones were isolated and cultured in 24-well culture plates. Of the clones, 20 clones with good cell growth rate were set to the same cell number (2 × 10 ⁵ cells / ml / 24-well) and incubated for another 3 days in a 24-well culture plate. The amount of BACE2-Fc protein secreted into the culture was quantified by ELISA method using goat anti-human IgG (Pierce), and the fastest growth rate and the amount of BACE2-Fc expression (1 liter culture reagent) Clone # 66, representing 3 mg).

Step C: BACE2 - Fc  Fusion Protein Production and Purification

The CHO DHFR- BACE2-Fc # 66 cell line was placed in a roller bottle containing 250 ml of a-MEM medium containing 10% dFBS at 2 X 10 ⁵ cells / ml, and at 40 rpm in a Roll-In cell incubator (Bellco). Incubated at 37 ° C. for 4 days while rotating at speed. Once the cells have grown to completely cover the culture vessel, wash once with 250 ml serum free medium (SFII; GIBCO-BRL) and then again with serum free containing 500 ml of insulin (0.5 ug / ml) (SIGMA) The medium was added and incubated for 3 days. After the culture was collected, 500 ml of serum-free medium was added again, followed by culturing for three days, and then all collected cell cultures were centrifuged at 7000 rpm for 20 minutes (Beckman, JA 10). rotor) and then only the washing liquid was separated. The washing solution was passed through a 0.45 um filter, passed through a Protein A sepharose chromatography column (Pharmacia) equilibrated with 20 mM sodium phosphata buffer (pH 7.0), and washed again with 20 mM sodium phosphata buffer (pH 7.0). All proteins that were not removed. 100 mM sodium acetate buffer (pH 3.5) was added to drop the adsorbed protein to obtain BACE2-Fc protein (MW 75 KDa.) With a purity of 95% or more.

Step D: Beta Using Fluorescent Label Specific Substrate Secretita 2  Activity test

In order to measure the enzyme activity of the beta secretase 2 and the inhibitory effect of the synthetic compounds, Fluorescence Resonance Energy Transfer (FRET) enzyme activity assay using the purified BACE2-Fc fusion protein and the fluorescence-labeled beta secretita 2 specific substrate was established. . This is briefly described as follows.

Peptide-specific substrates corresponding to the 10 amino acid region including the beta secretase ablation portion of the total amino acid sequence of Amyloid precursor protien (APP), which is known as the intracellular beta secretase 2 specific substrate, were identified as EDANS, which is a fluorophore. It was synthesized by attaching the quenching group DABCYL. When the fluorescent label substrate is reacted with BACE2-Fc, the quenching group is separated while the BACE2 action site is cleaved, and EDANS exhibits 510 nm wavelength fluorescence by 350 nm excitation light. The accuracy can be measured sensitively and simply.

Each synthetic compound was dissolved in DMSO at a concentration of 10 mM and stored at 20 ° C. In order to measure the activity, the 10 mM DMSO solution was first placed in the rightmost column of the 96 well plate, and then diluted twice with the same amount of DMSO to 9 steps. 10 ul of the diluted compound solution was added to a 96-well assay plate containing 600 uM fluorescent labeled BACE substrate dissolved in 15 ul of reaction buffer (50 mM sodiumacetate, pH 4.5, 0.05% CHAPS) and 10 ul of 50% DMSO. The addition resulted in a final DMSO concentration of 10% and an inhibitor treatment concentration ranging from 500 uM to 9 fold dilutions. After adding 65 ul of the purified BACE2-Fc fusion protein solution to a final concentration of 0.4 ug / ml, the reaction was performed at room temperature for 1 hour. The reaction product was measured using a fluorescent plate reader (SpectraMax Gemini XS, Molecular Device) as fluorescence at 350 nM excitation and 510 nm emission wavelength. By comparing this measurement with that in the control group without addition of the synthetic compound, the concentrations of the synthetic compound, namely IC ₅₀ and K i , which inhibited 50% of the beta secretase activity were determined.

Experimental Example  2: recombination Cascept Cindy  Enzyme Activity Measurement

In order to measure the enzyme activity of the casceptindi and the inhibitory effect of the synthetic compounds, Fluorescence Resonance Energy Transfer (FRET) enzyme assay using the casceptindi (Calbiochem, # 219401) and the fluorescently labeled casceptin specific substrate (Bachem # M-2455) Was established. This is briefly described as follows.

Known as a casceptin specific substrate, a peptide specific substrate corresponding to 10 amino acid sites is attached to Mca, a fluorophore, and Dnp, a quenching group. When the fluorescently labeled substrate reacts with CASE, the quenching group is cleaved as the CASE synthesizing site is cleaved and Mca emits 393 nm of fluorescence by excitation light of 328 nm. And can be measured easily.

Each synthetic compound was dissolved in DMSO at a concentration of 10 mM and stored at 20 ° C. In order to measure the activity, the 10 mM DMSO solution was first placed in the rightmost column of the 96 well plate, and then diluted twice with the same amount of DMSO to 9 steps. Final DMSO was added to a 96-well assay plate containing 7 uM fluorescently labeled Casceptin substrate dissolved in 50 ul of reaction buffer (50 mM sodiumacetate, pH 4.0) and 10 ul of 25% DMSO. The concentration was 5% and the inhibitor treatment concentration was from 2 u dilutions of 250 uM. 30 ul of the purified Casceptindi fusion protein solution was added to a final concentration of 0.75 ng / ml and the reaction was performed at 37 ° C. for 1 hour. The amount of reaction product was measured by using a fluorescent plate reader (SpectraMax Gemini XS, Molecular Device) with fluorescence at 328 nM excitation and 393 nm emission wavelength. By comparing this measurement with that in the control group without addition of the synthetic compound, the concentrations of the synthetic compound, i.e., IC ₅₀ and Ki, which inhibited 50% of the Casceptin activity was determined.

Experimental Example  3: intracellular SEAP ( Secreted alkaline phosphatase  Secreted alkali kinase) activity measurement

Step A: SEAP - APPsw - KK Permanent cell line expressing

Genes expressing SEAP and swedish mutant amyloid precursors (CRE-SEAP-APP695sweKK) under the control of cAMP response elements (CRE) were cloned into mammalian expression vector pcDNA3.1 (+) Neo (Invitrogen). Neuro-2a cells (ATCC Accession # CCL-131) were incubated with Dulbecco's minimum essential medium (DMEM) culture medium containing 10% FBS (GIBCO-BRL), then transferred to a 6-well culture plate and the cells were After overgrowth was transformed with the CRE-SEAP-APP695sweKK expression vector using Lipofectamine 2000 (Life Technologies). Each clone was isolated and incubated again on a 6-well culture plate. 100 clones of the selected clones with good cell growth rate were incubated for 3 days in a 24-well culture plate, then incubated for 6 hours in DMSO / 10uM Forskolin media, and then 50 μl of each well was removed and 50 ul Reaction with Attophos (Promega). Fluorescence at 450 nM excitation and 580 nm emission wavelength was measured with a fluorescent plate reader (SpectraMax Gemini XS, Molecular Device) at room temperature for 30 minutes. Among the selected clones, four clones with the highest SEAP activity in forskolin / DMSO with the highest SEAP activity were selected. Selected clones were incubated for 1 day with the same number of cells (2 X 10 ⁴ cells / 96-well), then treated with DMSO / Forskolin10uM to measure secreted SEAP activity after 6 hours to determine the best clone # 159. Screened.

Step B: SEAP  Activity Assay

N2A SEAP-APPsw-KK # 159 expressing the CRE-SEAP-APP695sweKK was placed in a 96-well culture vessel with 2 × 10 ⁴ cells / 80 ul of cells per well. Forskolin 10mM and DMSO were prepared and diluted 100-fold with culture medium, and 10 ul was added to 96-well each.

Each synthetic compound was dissolved in DMSO at a concentration of 10 mM and stored at 20 ° C. In order to measure the activity, the 10 mM DMSO solution was first placed in the rightmost column of the 96 well plate, and then diluted three times with the same amount of DMSO was performed sequentially to step 7. 10 ul of serially diluted compounds are diluted 10-fold with 90 ul of culture. The final DMSO concentration was 1.1% and the inhibitor treatment concentration was from 2 u dilutions of 100 uM. After all treatments were incubated for 5 hours in a 37 ℃ incubator in the presence of 6% CO ₂ . In order to measure the amount of SEAP secreted into the culture medium, the reaction was first performed at 65 ° C. for 30 minutes by heat to remove other alkaline phosphatase activity. 50 ul of the thermally inactivated culture and 50 ul of Attophos (Promega) were reacted at room temperature for 30 minutes, and fluorescence was measured at 450 nM excitation and 580 nm emission wavelength with a fluorescent plate reader (SpectraMax Gemini XS, Molecular Device). By comparing this measured value with the measured value in the control group to which the synthetic compound was not added, the concentration of the synthetic compound that inhibits 50% of the SEAP activity, namely IC ₅₀ , was determined.

Experimental Example  4: Genotyping In mice  Amyloid Beta Measurement in Neurons

Step A: Genetic Diversion  mouse( APP Of PS1dE9 Primary nerve cell culture

Experiments were performed with mice born from cross- genotyped (APP / PS1dE9) male and female mice. Take out the brains of rats 3-4 days old, remove the hippocampus tissue and the membranes at 4 ℃, and then chop them and treat them with DNA enzymes (Sigma, D5025) and protease (Sigma, P5147). Leave for 25 minutes. After separation into cells, use a culture medium (27.6 ml Neurobasal + 1.5 ml FBS + 600 ul B27 + 300 ul of 200 mM of L-Glutamine) in a 24-well plate covered with Poly-L-Lysine. ⁵ cells. After incubation for 7 days at 37 ℃ incubator.

Step B: Ab40  Activity measurement

Each synthetic compound was dissolved in DMSO at a concentration of 10 mM and stored at 20 ° C. To measure the activity, the 10 mM DMSO solution was first placed in the rightmost column of a 96 well plate, and then diluted three times with the same amount of DMSO up to six steps. The diluted compounds were diluted 250-fold with culture solution (neurobasal 29.1 ml + B27 600 ul + 200 mM L-Glutamine 300 ul + 100 mM L-Glutamate 7.5 ul) and 350 ul was added to the cultured primary neurons. The cells were incubated in a 37 ° C. incubator for hours. The expression level of beta amyloid peptide secreted into the culture solution was measured by sandwich ELISA method (Biosource, # KHB3482) using two antibodies specific for beta amyloid peptide.

Aqueous beta-amyloid or cell culture diluted to 50 ul of each concentration was added to the plate coated with the antibody while mixing with 50 ul of the identified antibody and reacted at room temperature for 3 hours (or at least 1 day at 4 ° C). After washing 5 times with 1-fold washing solution (Biosource, # KHB3482), 100 ul of HRP (horse radish peroxidase) -attached antibody diluted 100-fold with antibody dilution solution (Biosource, # KHB3482) was added, and the mixture was kept at room temperature for 30 minutes. Reacted. Then, the HRP antibody was removed, washed five times with a one-time wash solution, and then 100 ul of chromogen solution (Biosource, # KHB3482) was added and reacted at room temperature for 30 minutes. Then, 100 ul of a stop solution (Biosource, # KHB3482) was added and reacted at room temperature for 30 minutes, and the absorbance at 450 nm was measured using a microplate reader (SpectraMax 340, Molecular Device). This measurement was compared with that in the control treated with 0.4% DMSO without the addition of the synthetic compound to determine the concentration of the synthetic compound, i.e., IC ₅₀ , which inhibited 50% of intracellular beta secretase activity.

Experimental Example  5: intracellular beta Secretase  Activity measurement

Inhibition of intracellular beta secretase activity of synthetic compounds can be measured using a cell line making beta amyloid peptides from amyloid precursors (APP).

Step A: Establish Permanent Cell Line Producing Amyloid Precursor

Mutant amyloid precursor gene (APP751NFEV) under the control of TRE (Tet-response element) was cloned into PBI-L vector (CloneTech), a mammalian expression vector expressing the luciferase gene under the control of TRE (Tet-response element). Neuro-2a cells (ATCC Accession # CCL-131) were incubated with Dulbecco's minimum essential medium (DMEM) culture medium containing 10% FBS (GIBCO-BRL), then transferred to a 6-well culture plate and the cells were Overgrowth was then transformed with the pBI-L APP751 NFEV expression vector using Lipofectamine 2000 (Life Technologies). Each clone was isolated and incubated again on a 6-well culture plate. One hundred clones with good cell growth rate among the selected clones were incubated for one day in a 96-well culture plate, exchanged with a culture solution containing 1 ug / ml Doxicycline, and then incubated for 24 hours. After treatment with 50 μl of Bright-Glo Luciferase Reagent (Promega), the solution was left at room temperature for 15 minutes, and then the luminescence of each well was measured using a luminometer (Luminometer, Victor). Four clones with the highest luciferase expression among the selected clones were incubated for 1 day in a 24-well culture plate at the same cell count (3 X 10 ⁵ cells / ml / 24-well), then 1 ug / After exchange with OPTI-MEM (GIBCO-BRL) culture containing ml Doxicycline and incubated for 24 hours, the amount of beta amyloid peptide secreted into the culture was quantified by ELISA method using an antibody selective to beta amyloid peptide. As a result, clone # 79 was selected that exhibited the fastest growth rate and the highest amount of beta amyloid peptide expression.

Step B: Aqueous Beta-amyloid Precursor ( sAPP ) ELISA  Method

Neuro-2a APP751 NFEV # 79 cells, a cell line expressing the mutant amyloid precursors, were placed in a 24-well incubator at 3 × 10 ⁵ cells per well in a 37 ° C. incubator for 24 hours in the presence of 6% CO ₂ . Incubated for Once the cells were grown to completely cover the culture vessel, exchange them with Opti-MEM (GIBCO-BRL) containing 300 ul of 1 ug / ml Doxicycline and a beta secretase inhibitor (Example compound) diluted at each concentration. Incubate again for 24 hours. Aqueous beta-amyloid precursor expression amount secreted into the culture was measured according to the manufacturer's experimental plan using the Human sAPP assay kit (IBL), briefly described as follows.

Aqueous beta-amyloid precursors or cell cultures diluted to 100 ul of each concentration were added to the plate coated with the antibody and reacted at room temperature for 4 hours (or at least 1 day at 4 ° C). After washing 7 times with washing solution (PBS with 0.05% Tween20), 100 ul of HRP (horse radish peroxidase) antibody diluted 30-fold with antibody dilution solution (1% BSA, PBS with 0.05% Tween20) was added. After the addition, the mixture was reacted at room temperature for 30 minutes. After washing 9 times with the washing solution and 100 ul of TMB solution was added and reacted for 30 minutes at room temperature. After 100 ul of 1N sulfuric acid (H ₂ SO ₄ ) solution was added and reacted at room temperature for 30 minutes, the absorbance at 450 nm was measured using a microplate reader (SpectraMax 340, Molecular Device). This measurement was compared with the measurement in the control treated with only 1% DMSO without addition of the synthetic compound to determine the concentration of the synthetic compound, i.e., IC ₅₀ , which inhibits 50% of intracellular beta secretase activity.

Experimental Example  6: beta amyloid protein assay

Beta amyloid protein (beta-amyloid) was quantitated by enzyme-linked immune antibody assay (ELISA) using two antibodies (Human beta amyloid 1-40 colorimetric immunoassay kit, Biosource, California, USA). The two antibodies used in the enzyme-linked immune antibody assay consist of an antibody that specifically recognizes the N-terminus of the beta amyloid protein truncated by beta secretase and an antibody that binds to the C-terminus. After reacting the two antibodies and the beta amyloid protein at room temperature for three hours (or at least one day at 4 ° C.), they are washed four times with a washing solution and then subjected to a horseradish peroixdase (HRP) attached anti-rabbit IgG's peroxidase antibody. The reaction was carried out for 30 minutes. After washing four times with the wash solution, tetramethylbenzidine, a substrate of HRP, was added and reacted at room temperature for 30 minutes, and then absorbance at 450 nm was measured using a microplate reader (SpectraMax 340, Molecular Device). The degree of reduction of beta amyloid protein was determined by comparing this measurement with that in the control.

The secreted amyloid precursor protein beta (sAPPbeta) protein is also carried out in the same manner as the enzyme-linked immuno antibody assay used to quantify beta amyloid protein.

Experimental Example  7: animal test ( in vivo assay )

In order to confirm that the activity of beta secretase was inhibited, the degree of inhibition of production of beta amyloid protein, a product of beta secretase, was tested in animals. The animals used in the test were transgenic mice (Jankosky JL et al) containing a Swedish mutant (chimeric mouse / human amyloid precursor protein 695swe) of the beta amyloid protein precursor and a mutant gene of presenilin 1 (presenilin 1-dE9). , Biomolecular engineering, 17 (6), 157-165, 2001). Beta-secretase inhibitors are administered at concentrations that are expected to reduce amyloid protein but not toxicity through intraperitoneal or subcutaneous administration. Animals were anesthetized at a defined time after drug administration and blood and brain tissue separated. Blood was collected in a heparin-coated tube via cardiac blood collection, centrifuged at 13,000 rpm for 10 minutes (Eppendorf), and then only plasma was separated and stored at 80 ° C until analyzed with the extracted brain tissue (cerebral cortex and hippocampus). . For analysis, the plasma was diluted five-fold and the inhibitory effect of amyloid protein in plasma was confirmed by the enzyme-linked immune antibody assay mentioned above. After the brain tissue was extracted, the brain tissue was pulverized in a 4 times volume of PBS, and finally reacted with 8.2 M guanidine / 82 mM Tris HCl (pH 8.0) at 5M Guanidine at room temperature for 4 hours. Beta amyloid protein was extracted and analyzed by diluting the beta amyloid protein 1: 500 using BSAT-DPBS (Dulbecco's phosphate buffered saline with 5% BSA and 0.03% Tween-20).

Experimental Example  8: Dosing and formulation

The drug was dissolved in 10% HPCD (hydroxypropyl-beta-cyclodextrin) and then generally administered at a concentration of 15 mg / kg to 100 mg / kg once or three to five times daily.

The compounds of the present invention have a range of Ki from about 0.001 uM to 500 uM, preferably from about 0.001 uM to 1 uM, more preferably from about 0.001 uM to 0.1 uM. Ki values of representative example compounds are shown in Table 6 below.

TABLE 6

In addition, the IC ₅₀ of SEAP Cell activity is in the range of about 0.1 uM to 100 uM, preferably 0.1 uM to 10 uM, and more preferably 0.1 uM to 1.0 uM. IC ₅₀ values related to SEAP Cell activity of representative example compounds are shown in Table 7 below.

TABLE 7

In general, the example compounds are Cat. Selectivity to D is shown, typically Example 12 is about 10 times, Example 27 is about 15.2 times and Example 29 is about 29.5 times.

In addition, these compounds also have excellent cell activity against primary neuronal cells, and in Example 13, the IC ₅₀ value was about 0.050 uM.

Some compounds have excellent brain permeability, and Example 3 effectively reduces Ab40 in brain and plasma in double mutant transgenic mice, and effectively decreases sAPPb in brain.

As described above, the compounds of formula (I) according to the present invention exert an excellent inhibitory effect on human beta cecritas. Therefore, the compound according to the present invention can be used as a medicament for improving cognitive function or preventing and treating neurodegenerative diseases such as Alzheimer's disease.

Those skilled in the art to which the present invention pertains will be able to perform various applications and modifications within the scope of the present invention based on the above contents.

Claims (17)

A compound represented by formula (I), a pharmaceutically acceptable salt or isomer thereof:

(I) Where n is 0 to 2; A is represented by the following formula (II),

(II) Wherein W, X, Y and Z are each independently C or N atoms, one or two of which is N; R ₁ is-(CR ₆ R ₇ ) _m -R ₈ , where m is 0 to 2, R ₆ is selected from the group consisting of hydrogen, alkyl and alkoxy, R ₇ is hydrogen, alkyl and -C (O NHR ₆ is selected from the group consisting of R ₈ is alkyl, alkoxy, arylalkoxy, cycloalkyl, heterocycle, heteroaryl, -C (O) R ₆ and -C (O) NR ₆ R ₇ Wherein R ₆ and R ₇ are as defined above; R ₂ is selected from the group consisting of hydrogen, alkyl and-(CR ₆ R ₇ ) _m -R ₉ , wherein m is 0 to 2, R ₆ and R ₇ are as defined above and R ₉ is aryl , R ₃ is selected from the group consisting of hydrogen, alkyl, alkoxy and — (CR ₆ R ₇ ) _p —R ₁₀ , wherein p is 0 to 2, R ₆ and R ₇ are as defined above and R ₁₀ is cyclo Alkyl, aryl, heterocycle, and heteroaryl; R ₄ is selected from the group consisting of hydrogen, alkyl, cycloalkyl and-(CH ₂ ) _p -A'-R ₁₁ , wherein A 'is aryl or heteroaryl, and R ₁₁ is hydrogen, halogen, hydroxy, alkyl , Alkoxy and —NR ₆ R ₇ wherein R ₆ and R ₇ are as defined above; Wherein R ₆ and R ₇ are bonded to each other to form a cyclized structure or an acyclic ring; The alkyl, alkoxy, aryl, cycloalkyl, heterocycle, and heteroaryl may have a substituted or unsubstituted structure, where the substituents are halogen, amino, alkylamino, dialkylamino, alkylacylamino, C _1- C ₄ At least one selected from the group consisting of alkyl, hydroxy, C ₁ -C ₄ alkoxy, arylalkoxy and oxo; The heteroaryl and heterocycle are each independently a 4-8 membered ring containing 1 to 3 heteroatoms selected from the group consisting of O, N and S, and 0 to 2 double bonds. The method of claim 1, R_OneIs-(CR₆R₇)_m-R₈Where m is 0 to 2 and R₆Is hydrogen or C_One-C₈ Alkyl, R₇Silver hydrogen, C_One-C₈ Alkyl and -C (O) NHR₆Is selected from the group consisting of R₈Silver c_One-C₈ Alkyl, C_One-C₈ Alkoxy, arylalkoxy, cycloalkyl, heteroaryl, -C (O) R₆ And -C (O) NR₆R₇It is selected from the group consisting of; R ₂ is selected from the group consisting of hydrogen, C ₁ -C ₈ alkyl and-(CR ₆ R ₇ ) _m -R ₉ , wherein m is 0 to 2 and R ₉ is phenyl; R ₃ is selected from the group consisting of hydrogen, C ₁ -C ₈ alkyl, C ₁ -C ₈ alkoxy and — (CR ₆ R ₇ ) _p -R ₁₀ , wherein p is 0 to 2 and R ₁₀ is cycloalkyl , Aryl, heterocycle and heteroaryl; R ₄ is selected from the group consisting of hydrogen, C ₁ -C ₆ alkyl, cycloalkyl of a 5 or 6 membered ring, and — (CH ₂ ) _p —A′-R ₁₁ , wherein A ′ is aryl or heteroaryl, R ₁₁ is hydrogen, halogen, hydroxy, C ₁ -C ₈ Alkyl is selected from the group consisting of C ₁ -C ₈ alkoxy and -NR ₆ R _7, R ₆ And R ₇ is a structure linked to and cyclized to each other, wherein the compound represented by the formula (I), a pharmaceutically acceptable salt or isomer thereof. A compound represented by the formula (I), a pharmaceutically acceptable salt or isomer thereof, according to claim 1, wherein A is any one of the heterocycles of the formulas (i) to (iv): (i)

(ii)

(iii)

(iv)

The compound of claim 1, wherein R ₁ is — (CR ₆ R ₇ ) _m -R ₈ , wherein m is 0 to 2, R ₆ is hydrogen or C ₁ -C ₈ alkyl, R ₇ is hydrogen, C ₁ -C ₈ Alkyl and —C (O) NHR ₆ , wherein R ₈ is C ₁ -C ₈ alkyl, C ₁ -C ₈ alkoxy, arylalkoxy, cycloalkyl, heteroaryl, -C (O) R ₆ And -C (O) NR ₆ R ₇ , a pharmaceutically acceptable salt or isomer thereof. The compound of claim 1, wherein R ₁ is — (CHR ₇ ) _m -R ₈ , wherein m is 0 or 1, R ₇ is hydrogen, C ₁ -C ₄ alkyl or —C (O) NHR ₆ , and R ₈ is C ₁ -C ₈ alkyl, arylalkoxy, cycloalkyl, -C (O) R ₆ and -C (O) NR ₆ R ₇ A compound, characterized in that it is pharmaceutically acceptable Salts or isomers of. The compound of claim 5, wherein R ₁ is hydrogen, benzyl, dimethylamino-carbonyl, propyl-carbonyl, phenethyl, butyl, pentyl, ethylcarbamoyl, trifluorobenzyl, 4-trifluoromethylbenzyl, 2- Trifluoromethylbenzyl phenoxyethyl, 2,4,5-trifluorobenzyl, 4-fluorobenzyl, 3,4-difluorobenzyl, 2,4-difluorobenzyl, 3,5-difluoro Compound, characterized in that selected from the group consisting of lobenzyl 3,5-dimethoxybenzyl, 3-methoxybenzyl, 3,5-dibromophenyl, cyclohexylmethyl and 2-acetylaminophenyl, pharmaceutically acceptable Its salts or isomers. The compound, pharmaceutically acceptable salt or isomer thereof of claim 1, wherein R ₂ is selected from the group consisting of hydrogen, C ₁ -C ₈ alkyl and benzyl. 8. A compound, pharmaceutically acceptable salt or isomer thereof according to claim 7, wherein R ₂ is selected from the group consisting of hydrogen, methyl and benzyl. The compound of claim 1, wherein R ₃ is hydrogen, C ₁ -C ₈ Alkyl, cycloalkyl, and aryl, wherein alkyl, cycloalkyl, and allyl are each independently halogen, hydroxy, C ₁ -C ₄ Alkyl, C ₁ -C ₄ A compound represented by the following formula (I), a pharmaceutically acceptable salt or isomer thereof, characterized in that the structure is unsubstituted or substituted by one or more substituents selected from the group consisting of alkoxy and arylalkoxy. The compound of formula (I) according to claim 9, wherein R ₃ is phenyl unsubstituted or substituted with one or more substituents selected from the group consisting of fluorine, chlorine and methyl. Its salts or isomers. 11. A compound represented by the formula (I), a pharmaceutically acceptable salt or isomer thereof, according to claim 10, wherein R ₃ is phenyl or 3,5-difluoro phenyl. The compound of claim 1, wherein R ₄ is selected from the group consisting of hydrogen, C ₁ -C ₆ alkyl, cycloalkyl and — (CH ₂ ) —A′-R ₁₁ , wherein A ′ is aryl or heteroaryl, R ₁₁ is selected from the group consisting of hydrogen, halogen, hydroxy, alkyl, alkoxy and -NR ₆ R ₇ , wherein R ₆ is selected from the group consisting of hydrogen, C ₁ -C ₃ alkyl and C ₁ -C ₃ alkoxy , R ₇ is selected from the group consisting of hydrogen, C ₁ -C ₄ alkyl and -C (O) NHR ₆ , a compound represented by the formula (I), a pharmaceutically acceptable salt or isomer thereof. The compound of claim 12, wherein R ₄ is 3-dimethylaminobenzyl, 3-isopropylbenzyl, 3-trifluoromethoxybenzyl, 3-trifluoromethylbenzyl, 3-ethylbenzyl, 3-t-butylbenzyl, ethyl And cyclohexyl, a compound represented by the formula (I), a pharmaceutically acceptable salt or isomer thereof. The compound of formula I, a pharmaceutically acceptable salt or isomer thereof, according to claim 1, wherein the compound of formula I is any one selected from the following compounds: (3S) -1-Benzyl-N-[(2S, 3R) -4- (3-dimethylaminophenyl) methylamino-3-hydroxy-1-phenylbutan-2-yl] -5-oxopyrrolidine -3-carboxamide; (3R) -1-benzyl-N-[(2S, 3R) -4- (3-dimethylaminophenyl) methylamino-3-hydroxy-1-phenylbutan-2-yl] -5-oxopyrrolidine -3-carboxamide; (2S) -N-[(2S, 3R) -4- (3-dimethylaminophenyl) methylamino-3-hydroxy-1-phenylbutan-2-yl] -5-oxopyrrolidine-2-car Radiamide; (2R) -N-[(2S, 3R) -4- (3-dimethylaminophenyl) methylamino-3-hydroxy-1-phenylbutan-2-yl] -5-oxopyrrolidine-2-car Radiamide; 2-((3S) -1-benzyl-5-oxopyrrolidin-3-yl) -N-[(2R, 3R) -4- (3-dimethylaminophenyl) methylamino-3-hydroxy-1- Phenylbutan-2-yl] acetamide; 2-((3R) -1-benzyl-5-oxopyrrolidin-3-yl) -N-[(2R, 3R) -4- (3-dimethylaminophenyl) methylamino-3-hydroxy-1- Phenylbutan-2-yl] acetamide; (4S) -1-benzyl-N-[(2S, 3R) -4- (3-dimethylaminophenyl) methylamino-3-hydroxy-1-phenylbutan-2-yl] -2-oxoimidazoli Dine-4-carboxamide; (4S) -N-[(2S, 3R) -4- (3-dimethylaminophenyl) methylamino-3-hydroxy-1-phenylbutan-2-yl] -2-oxoimidazolidine-4- Carboxamides; (4S) -1- [2-[(2S, 3R) -4- (3-dimethylaminophenyl) methylamino-3-hydroxy-1-phenylbutan-2-yl] amino-2-oxoethyl]- N, N-dimethyl-2-oxoimidazolidine-4-carboxamide; (4S) -1-butanoyl-N-[(2S, 3R) -4- (3-dimethylaminophenyl) methylamino-3-hydroxy-1-phenylbutan-2-yl] -2-oxoimida Zolidine-4-carboxamide; (4S) -N- [4- (3-dimethylaminophenyl) methylamino-3-hydroxy-1-phenylbutan-2-yl] -2-oxo-1-phenethylimidazolidine-4-car Radiamide; N-[(2S, 3R) -4- (3-dimethylaminophenyl) methylamino-3-hydroxy-1-phenylbutan-2-yl] -2- (1-methyl-oxopyrrolidine-3- I) acetamide; 1-butyl-N- [4- (3-dimethylaminophenyl) methylamino-3-hydroxy-1-phenylbutan-2-yl] -2-oxoimidazolidine-4-carboxamide; (3S) -N-[(2S, 3R) -4- (3-dimethylaminophenyl) methylamino-3-hydroxy-1-phenylbutan-2-yl] -5-oxo-1-phenethylpyrrolidine -3-carboxamide; (3R) -N-[(2S, 3R) -4- (3-dimethylaminophenyl) methylamino-3-hydroxy-1-phenylbutan-2-yl] -5-oxo-1-phenethylpyrrolidine -3-carboxamide; 1- (2-amino-2-oxoethyl) -N-[(2S, 3R) -4- (3-dimethylaminophenyl) methylamino-3-hydroxy-1-phenylbutan-2-yl] -5 Oxopyrrolidine-3-carboxamide; (3S) -1-benzyl-N-{(2S, 3R) -3-hydroxy-1-phenyl-4-[(3-trifluoromethoxyphenyl) methylamino] butan-2-yl} -5- Oxopyrrolidine-3-carboxamide; (4S) -N-[(2S, 3R) -4- (3-dimethylaminophenyl) methylamino-3-hydroxy-1-phenylbutan-2-yl] -N-ethyl-2-oxoimidazoli Dine-1,4-dicarboxamide; (3S) -1-[(2S) -3- (4-chlorophenyl) -1-ethylamino-1-oxopropan-2-yl] -N-[(2S, 3R) -4- (3-dimethyl Aminophenyl) methylamino-3-hydroxy-1-phenylbutan-2-yl] -5-oxopyrrolidine-3-carboxamide; (3R) -1-[(2S) -3- (4-chlorophenyl) -1-ethylamino-1-oxopropan-2-yl] -N-[(2S, 3R) -4- (3-dimethyl Aminophenyl) methylamino-3-hydroxy-1-phenylbutan-2-yl] -5-oxopyrrolidine-3-carboxamide; (3S) -1-benzyl-N-{()-3-hydroxy-1-phenyl-4-[(3-trifluoromethoxyphenyl) methylamino] butan-2-yl} -2-oxoimida Zolidine-4-carboxamide; (3S) -2,4-dibenzyl-N-[()-4- (3-dimethylaminophenyl) methylamino-3-hydroxy-1-phenylbutan-2-yl] -5-oxopyrrolidine -2-carboxamide; (3R) -2,4-dibenzyl-N-[()-4- (3-dimethylaminophenyl) methylamino-3-hydroxy-1-phenylbutan-2-yl] -5-oxopyrrolidine -2-carboxamide; (2S) -4-benzyl-N-[()-4- (3-dimethylaminophenyl) methylamino-3-hydroxy-1-phenylbutan-2-yl] -5-oxopyrrolidine-2- Carboxamides; (4S) -1,4-dibenzyl-N-[()-4- (3-dimethylaminophenyl) methylamino-3-hydroxy-1-phenylbutan-2-yl] -2-oxoimidazoli Dine-4-carboxamide; (4S) -N-[()-4- (3-dimethylaminophenyl) methylamino-3-hydroxy-1-phenylbutan-2-yl] -2-oxo-1-[(2,4,5 -Trifluorophenyl) methyl] imidazolidine-4-carboxamide; (4S) -N-[()-4- (3-dimethylaminophenyl) methylamino-3-hydroxy-1-phenylbutan-2-yl] -2-oxo-1-[(4-trifluoro Methylphenyl) methyl] imidazolidine-4-carboxamide; (4S) -N-[()-4- (3-dimethylaminophenyl) methylamino-3-hydroxy-1-phenylbutan-2-yl] -2-oxo-1-[(4-trifluorome Methoxyphenyl) methyl] imidazolidine-4-carboxamide; (4S) -1-benzyl-N-[()-1- (3,5-difluorophenyl) -4- (3-dimethylaminophenyl) methylamino-3-hydroxybutan-2-yl]- 2-oxoimidazolidine-4-carboxamide; (3S) -1-benzyl-N-[(2S, 3R) -1- (3,5-difluorophenyl) -4- (3-dimethylaminophenyl) methylamino-3-hydroxybutane-2 -Yl] -5-oxopyrrolidine-3-carboxamide; (4S) -N-[(2S, 3R) -4- (3-dimethylaminophenyl) methylamino-3-hydroxy-1-phenylbutan-2-yl] -2-oxo-1- (2-phenoxy Cyethyl) imidazolidine-4-carboxamide; (4R) -N-[(2S, 3R) -4- (3-dimethylaminophenyl) methylamino-3-hydroxy-1-phenylbutan-2-yl] -2-oxo-1- (2-phenoxy Cyethyl) imidazolidine-4-carboxamide; (4S) -N-[(2S, 3R) -4- (3-dimethylaminophenyl) methylamino-3-hydroxy-1-phenylbutan-2-yl] -2-oxo-1-[(2- Trifluoromethylphenyl) methyl] imidazolidine-4-carboxamide; (4S) -N-[(2S, 3R) -1- (3,5-difluorophenyl) -4- (3-dimethylaminophenyl) methylamino-3-hydroxybutan-2-yl] -2 -Oxo-1-[(2,4,5-trifluorophenyl) methyl] imidazolidine-4-carboxamide; (4S) -1-benzyl-N-[(2S, 3R) -4- (cyclohexylamino) -3-hydroxy-1-phenylbutan-2-yl] -2-oxoimidazolidine-4- Carboxamides; (4R) -1-benzyl-N-[(2S, 3R) -4- (3-dimethylaminophenyl) methylamino-3-hydroxy-1-phenylbutan-2-yl] -4-methyl-2- Oxoimidazolidine-4-carboxamide; (4S) -1-benzyl-N-[(2S, 3R) -4- (3-dimethylaminophenyl) methylamino-3-hydroxy-1-phenylbutan-2-yl] -4-methyl-2- Oxoimidazolidine-4-carboxamide; (4S) -N-[(2S, 3R) -4- (3-dimethylaminophenyl) methylamino-3-hydroxy-1-phenylbutan-2-yl] -1-[(4-fluorophenyl) Methyl] -2-oxoimidazolidine-4-carboxamide; (4S) -1-[(3,4-difluorophenyl) methyl-N-[(2S, 3R) -4- (3-dimethylaminophenyl) methylamino-3-hydroxy-1-phenylbutane- 2-yl] -2-oxoimidazolidine-4-carboxamide; (4S) -1-[(2,4-difluorophenyl) methyl-N-[(2S, 3R) -4- (3-dimethylaminophenyl) methylamino-3-hydroxy-1-phenylbutane- 2-yl] -2-oxoimidazolidine-4-carboxamide; (4S) -1-benzyl-N-[(2S, 3R) -3-hydroxy-1-phenyl-4-[(3-trifluoromethylphenyl)] methylamino] butan-2-yl] -2- Oxoimidazolidine-4-carboxamide; (4S) -1-benzyl-N-{(2S, 3R) -3-hydroxy-1-phenyl-4-[(3-propan-2-ylphenyl) methylamino] butan-2-yl} -2 Oxoimidazolidine-4-carboxamide; (4S) -N-{(2S, 3R) -1- (3,5-difluorophenyl) -3-hydroxy-4-[(3-propan-2-ylphenyl) methylamino] butane-2 -Yl} -2-oxo-1-[(2,4,5-trifluorophenyl) methyl] imidazolidine-4-carboxamide; (4S) -1- (3,5-dimethoxyphenyl) methyl-N-[(2S, 3R) -4- (3-dimethylaminophenyl) methylamino-3-hydroxy-1-phenylbutan-2- Il] -2-oxoimidazolidine-4-carboxamide; (4S) -N-[(2S, 3R) -4- (3-ethylphenyl) methylamino-3-hydroxy-1-phenylbutan-2-yl] -2-oxo-1-[(2,4 , 5-trifluorophenyl) methyl] imidazolidine-4-carboxamide; (4S) -N-[(2S, 3R) -4- (3-thibutylphenyl) methylamino-3-hydroxy-1-phenylbutan-2-yl] -2-oxo-1-[(2, 4,5-trifluorophenyl) methyl] imidazolidine-4-carboxamide; (4S) -1-[(3,5-difluorophenyl) methyl-N-[(2S, 3R) -4- (3-dimethylaminophenyl) methylamino-3-hydroxy-1-phenylbutane- 2-yl] -2-oxoimidazolidine-4-carboxamide; (4S) -N-[(2S, 3R) -4- (3-dimethylaminophenyl) methylamino-3-hydroxy-1-phenylbutan-2-yl] -1-[(3-methoxyphenyl) Methyl] -2-oxoimidazolidine-4-carboxamide; (4S) -N-[(2S, 3R) -4- (3-dimethylaminophenyl) methylamino-3-hydroxy-1-phenylbutan-2-yl] -1-[(4-methoxyphenyl) Methyl] -2-oxoimidazolidine-4-carboxamide; (4S) -N-[(2S, 3R) -4- (3-thibutylphenyl) methylamino-3-hydroxy-1-phenylbutan-2-yl] -1- (3,5-difluoro Phenyl) methyl-2-oxoimidazolidine-4-carboxamide; (4S) -1- (3,5-difluorophenyl) methyl-N-{(2S, 3R) -3-hydroxy-1-phenyl-4-[(3-propan-2-ylphenyl) methyl Amino] butan-2-yl} -2-oxoimidazolidine-4-carboxamide; (4S) -N-[(2S, 3R) -4- (3-thibutylphenyl) methylamino-1- (3,5-difluorophenyl) -3-hydroxybutan-2-yl] -1 -(3,5-difluorophenyl) methyl-2-oxoimidazolidine-4-carboxamide; (4S) -N-((2S, 3R) -4-ethylamino-3-hydroxy-1-phenylbutan-2-yl) -2-oxo-1-[(2,4,5-trifluoro Phenyl) methyl] imidazolidine-4-carboxamide; (4S) -1- (3,5-dibromo) phenyl-N-[(2S, 3R) -4- (3-dimethylaminophenyl) methylamino-3-hydroxy-1-phenylbutan-2- Il] -2-oxoimidazolidine-4-carboxamide; (4S) -N-[(2S, 3R) -4- (3-dimethylaminophenyl) methylamino-3-hydroxy-1-phenylbutan-2-yl] -2-oxo-1-phenylimidazoli Dine-4-carboxamide; (3S) -1-cyclohexylmethyl-N-[(2S, 3R) -4- (3-dimethylaminophenyl) methylamino-3-hydroxy-1-phenylbutan-2-yl] -5-oxopy Rollidine-3-carboxamide; (3R) -1-cyclohexylmethyl-N-[(2S, 3R) -4- (3-dimethylaminophenyl) methylamino-3-hydroxy-1-phenylbutan-2-yl] -5-oxopy Rollidine-3-carboxamide; (4S) -N-[(2S, 3R) -4- (3-thibutylphenyl) methylamino-3-hydroxy-1-phenylbutan-2-yl] -2-oxoimidazolidine-4- Carboxamides; (4S) -N-[(2S, 3R) -4-[(3-dimethylaminophenyl) methylamino] -3-hydroxy-1-phenylbutan-2-yl] -5-oxo-1-pentylpy Rollidine-4-carboxamide; (4R) -N-[(2S, 3R) -4-[(3-dimethylaminophenyl) methylamino] -3-hydroxy-1-phenylbutan-2-yl] -5-oxo-1-pentylpy Ralidin-4-carboxamide. A pharmaceutical composition for inhibiting beta secretase, comprising as an active ingredient a compound of formula (I) according to claim 1, a pharmaceutically acceptable salt or isomer thereof, and a pharmaceutically acceptable carrier. The pharmaceutical composition for inhibiting beta secretase according to claim 15, wherein the pharmaceutical composition is used to improve cognitive function or to treat or prevent neurodegenerative diseases. 17. The pharmaceutical composition for inhibiting beta secretase according to claim 16, wherein the composition is used to treat Alzheimer's disease.