KR101357234B1 - Novel Quinoxaline Derivates and Compositions Comprising the Same for Inducing Differentiation of Neural Precursor Cells or Stem Cells into Neural Cells - Google Patents

Abstract

본 발명에 따르면, 신규한 퀴녹살린 유도체 및 이를 포함하는 분화 유도용 조성물은 신경전구세포 또는 줄기세포를 신경세포로 분화시킬 수 있다. 또한, 본 발명에 의하여 분화 유도된 신경전구세포는 특정세포(예컨대, 도파민 신경세포) 또는 올리고덴드로사이트등으로 고효율로 분화할 수 있고, 차후 난치성 신경계 질환(예컨대, 파킨슨 병이나 척수손상)과 같은 난치성 신경계질환 적용할 수 있으며 신약계발에 있어서 기초적인 자료를 제공한다. The present invention relates to quinoxaline derivatives or salts thereof represented by the formula:

According to the present invention, the novel quinoxaline derivative and composition for inducing differentiation comprising the same can differentiate neural progenitor cells or stem cells into neurons. In addition, the neural progenitor cells induced by differentiation according to the present invention can be efficiently differentiated into specific cells (eg, dopamine neurons) or oligodendroseite, and the like. Refractory neurological diseases can be applied and provide basic data on drug development.

Description

Novel Quinoxaline Derivates and Compositions Comprising the Same for Inducing Differentiation of Neural Precursor Cells or Stem Cells into Neural Cells}

The present invention relates to a novel quinoxaline derivative and a composition for inducing differentiation of neural precursor cells or stem cells into neurons using the same.

Stroke, Alzheimer's disease, Parkinson's disease, demyelinating disease, spinal cord injury, and dementia are disorders of nerve function caused by nerve cell damage. Although a drug therapy or a surgical operation is generally used as a method of treating damaged nerve cells due to the diseases, such treatment is also pointed out as a problem due to damage to normal cells.

Recently, a cell therapeutic agent that supplies cells destroyed or damaged by diseases from the outside has been suggested to be effective. As a cell therapy agent, stem cells are in the spotlight.

Stem cells are generic to undifferentiated cells before they are differentiated into each cell constituting the tissue, and have the ability to differentiate into specific cells by specific differentiation stimulus (environment). Stem cells, unlike differentiated cells that cease cell division, are able to self-renewal by cell division, and differentiate into specific cells when differentiation stimulation is applied, and this differentiation is different in different environments or in different ways. It is characterized by the plasticity of differentiation, which can be differentiated into various cells by differentiation stimulation. Stem cells are embryonic stem cells, somatic cell nuclear transfer, adult stem cells, and recently reported induced pluripotent, depending on their cytological origin. stem cells (iPSCs). Adult stem cells are derived from each organ in the adult, such as bone marrow, brain, liver, pancreas, etc., while embryonic stem cells are derived from preimplantation fertilized eggs or developing fetal genital tissues. They are capable of differentiating into the cells that make up the human body, and are being spotlighted worldwide as a tool for treating intractable diseases and genetic diseases.

Unlike other tissues, cerebral nervous system tissue has almost no immune rejection reaction, so when transplanting stem cells, long-term survival of the transplanted cells can be expected, thus preventing various neurological diseases caused by nerve cell damage. Many studies have been made as a cell therapy for Korean. Thus, attempts are being made to apply stem cells to the treatment of diseases such as stroke, Alzheimer's disease, Parkinson's disease, demyelinating diseases, and spinal cord injury (Isacon O, Deacon T, Trends. Neurosci . , 10: 477-482 (1997). Studer et al ., Nat. Neurosci . , 1: 290-295 (1998). However, in order to increase the usefulness of stem cells as cell therapeutics, a technique for efficiently differentiating stem cells into specific cells is required. To this end, the present inventors synthesized a novel quinoxaline derivative compound, and identified the compounds of the present invention as a novel stem cell differentiation material.

Numerous papers and patent documents are referenced and cited throughout this specification. The disclosures of the cited papers and patent documents are incorporated herein by reference in their entirety to better understand the state of the art to which the present invention pertains and the content of the present invention.

The present inventors earnestly researched to develop a substance that induces differentiation of neural precursor cells or stem cells into neurons. As a result, the present inventors synthesized a substance having a molecular backbone similar to the quinoxaline derivative that we have already identified as a substance that induces differentiation into neurons, and found that these substances differentiate stem cells into specific cells. Furthermore, the present invention was completed by confirming that the efficiency was also quite high.

It is therefore an object of the present invention to provide novel quinoxaline derivatives or salts thereof.

Another object of the present invention is to provide a composition for inducing differentiation of neural precursor cells or stem cells into neurons.

Still another object of the present invention is to provide a method for differentiating neural progenitor cells or stem cells into neurons.

Still another object of the present invention is to provide a pharmaceutical composition for preventing or treating a neurological injury disease.

Other objects and advantages of the present invention will become more apparent from the following detailed description of the invention, claims and drawings.

According to one aspect of the present invention, the present invention provides a quinoxaline derivative represented by the following formula (1) or a salt thereof:

Formula 1

(A₁은 수소 또는 헤테로원자로서 산소, 황 또는 질소를 포함하는 C_4-5 헤테로사이클로알킬) 또는

(A₂는 수소 또는 C_1-6 알킬)이며; R₄는 인접한 고리탄소와 이중결합을 형성하는 경우는 산소이고, 단일결합을 형성하는 경우는 C_1-6 알콕시이고; n은 0-10의 정수이며;

는 단일결합 또는 이중결합을 나타내며; R₁이 비치환된 페닐기인 경우, R₂, R₃ 및 R₄는 각각

, H 및 산소가 아니다.
In Formula 1, R ₁ is C _6-10 aryl unsubstituted or substituted with C _1-6 alkoxy, C _1-6 alkyl or halo; R ₂ and R ₃ are each independently hydrogen,

(A ₁ is hydrogen or C _4-5 heterocycloalkyl comprising oxygen, sulfur or nitrogen as a heteroatom) or

(A ₂ is hydrogen or C _1-6 alkyl); R ₄ is oxygen when forming a double bond with an adjacent ring carbon, and C _1-6 alkoxy when forming a single bond; n is an integer from 0-10;

Represents a single bond or a double bond; When R ₁ is an unsubstituted phenyl group, R ₂ , R ₃ and R ₄ are each

, Not H and oxygen.

According to another aspect of the present invention, a composition for inducing differentiation of neural precursor cells or stem cells into neurons, comprising a quinoxaline derivative represented by Formula 1 or a salt thereof as an active ingredient. To provide.

The present inventors earnestly researched to develop a substance that induces differentiation of neural precursor cells or stem cells into neurons. As a result, the present inventors synthesized a substance having a molecular backbone similar to the quinoxaline derivative that we have already identified as a substance that induces differentiation into neurons, and found that these substances differentiate stem cells into specific cells. In addition, it was confirmed that the efficiency is also quite high.

The compound of the present invention represented by the formula (1) is chemically synthesized by mimicking the structure of the quinoxaline derivatives that have been identified by the present inventors to differentiate stem cells into specific cells.

As used herein, the term “halo” used to define a quinoxaline derivative of formula (1) or a salt thereof refers to a halogen group element, including, for example, fluoro, chloro, bromo and iodo.

The term “alkyl” refers to a straight or branched unsubstituted or substituted saturated hydrocarbon group, for example methyl, ethyl, propyl, isobutyl, pentyl, hexyl, heptyl, octyl, nonyl, decyl, undecyl, tridecyl , Pentadecyl and heptadecyl and the like. C _{1 -6} alkyl is meant an alkyl group having a carbon number of cases, the unit having an alkyl group having 1 to 6 carbon atoms, and, C _{1 -6} alkyl substituted with a substituent is not included. In the general formula _1, C 1 _-6 alkyl in R _1, R ₂ and R ₃ positions are preferably C _{1 -3} alkyl.

The term “heterocycloalkyl comprising oxygen, sulfur or nitrogen as a heteroatom” means a non-aromatic cyclic hydrocarbon group comprising carbon and hydrogen and at least one heteroatom (oxygen, sulfur or nitrogen). The heteroatom is preferably oxygen or sulfur, most preferably oxygen. The number of heteroatoms is preferably 1-4, more preferably 1-3, even more preferably 1-2, most preferably 1. C _{4 -5} heterocycloalkyl means a heterocycloalkyl carbon number is in the 4-5 to form a ring structure.

The term "alkoxy" means an -O group, and, for example, and the like ethoxy, methoxy, and it does not contain carbon atoms in the case where a C _{1 -6} alkoxy-substituted substituents.

The term “aryl” refers to a substituted or unsubstituted monocyclic or polycyclic carbon ring which is wholly or partially unsaturated. C _{6 -10} aryl group is not included in the carbon number of the substituent when the aryl group means a group having a carbon ring atom of a carbon number of 6 to 10, C _{6 -10} aryl-substituted. Preferably aryl is monoaryl or biaryl. It is preferable that monoaryl has 5-6 carbon atoms, and it is preferable that biaryl has 9-10 carbon atoms. Most preferably said aryl is substituted or unsubstituted phenyl. When monoaryl, such as phenyl, is substituted, substitutions may be made by various substituents at various positions, such as halo, hydroxy, nitro, cyano, C ₁ -C ₄ Substituted or unsubstituted straight or branched chain alkyl or C ₁ -C ₄ Lt; / RTI > may be substituted by straight chain or branched alkoxy.

According to a preferred embodiment of the present invention, R ₁ of Formula 1 is a C _{1 -3} alkoxy, or fluoro, and a more preferably methoxy or fluoro.

According to another preferred embodiment of the invention, A ₁ is hydrogen, tetrahydrofuran or tetrahydropyran, A ₂ is hydrogen or C _{1 -3} alkyl in the formula 1 R ₂ and R _3. More preferably A ₁ is hydrogen or tetrahydropyran and A ₂ is hydrogen or methyl.

According to another preferred embodiment of the invention, and it is oxygen when forming a ring carbon adjacent to R ₄ has the double bond of formula 1, in the case of forming a single bond is a C _{1 -3} alkoxy. More preferably, R ₄ is oxygen when forming a double bond with an adjacent ring carbon, and methoxy when forming a single bond.

According to a preferred embodiment of the invention, the quinoxaline derivatives or salts thereof of the invention are represented by the group consisting of the following formulas (2) to (18):

(2)

(3)

Formula 4

Formula 5

6

Formula 7

8

Formula 9

10

Formula 11

Formula 12

Formula 13

Formula 14

Formula 15

Formula 16

Formula 17

18

The compounds of the present invention may have one or more chiral centers and / or geometric isomeric centers, so that the present invention includes all stereoisomers represented by Formula 1, ie, optical isomers, diastereomers and geometric isomers. do.

Quinoxaline derivatives of the general formula (1) of the present invention are excellent in the ability to differentiate neuronal progenitor cells or stem cells into neurons. Accordingly, the present invention provides a composition for inducing differentiation of neuroprogenitor cells or stem cells into neurons, comprising the quinoxaline derivative of Formula 1 or a salt thereof as an active ingredient.

As used herein, the term "neural cell" refers to neurons and / or glia of the central or peripheral nervous system, for example, astrocytes, oligodendrocytes and / or schwann. It means a cell (Schwann cell).

The term "neural precursor cell" is also a cell in the early stages of differentiation of neurons capable of differentiating into neurons, astrocytes and oligodendrosites. On the other hand, the neural stem cells have the ability to continue proliferation, that is, self-replicating ability, and multipotent cells that differentiate into three constituent cells such as neurons, asrocytes, and oligodendrocytes that constitute the central nervous system. It can be defined as having undifferentiated cells. Neural stem cells differentiate into neurons / neurons or glia cells through the steps of neural precursor cells or glia precursor cells that produce specific nervous system cells.

As used herein, the term “induction of differentiation of stem cells into neurons” refers to embryonic analogues formed not only when stem cells are fully differentiated into neurons but also at intermediate stages before stem cells are fully differentiated into neurons. It also includes the formation of an embryonic body. That is, the stem cell differentiation inducing composition of the present invention not only effectively achieves full differentiation of stem cells into specific cells, but also shows very great efficiency in forming embryonic-like structures in stem cells.

The stem cells to which the present invention is applied are not limited, and cells having the characteristics of stem cells, that is, undifferentiated, infinite proliferation, and ability to differentiate into specific cells, are cells to which the present invention can be applied. Stem cells to which the present invention is applied are broadly classified into two types: pluripotent stem cells and multipotent stem cells, including embryonic stem cells (ES) and embryonic germ cells (EG). Embryonic stem cells are derived from the internal cell mass (ICM) of the blastocyst, and embryonic germ cells are derived from primordial germ cells of the 5-10 week old gonadal ridge. Pluripotent stem cells, on the other hand, are found in embryonic, fetal and adult tissues, including adult stem cells. Pluripotent stem cells proliferate indefinitely in vitro and have the ability to differentiate into a variety of cells derived from all three embryonic layers (ectoderm, mesoderm and endoderm).

According to a preferred embodiment of the present invention, the stem cells include embryonic stem cells, adult stem cells, induced pluripotent stem cells, embryonic germ cells and embryonic tumor cells, preferably embryonic stem cells and induced pluripotent stem cells. The term 'induced pluripotent stem cells' is one of pluripotent stem cells artificially derived by inserting certain genes from non-pluripotent cells (eg, somatic cells). Induced pluripotent pluripotent cells are pluripotent stem cells in terms of stem cell gene and protein expression, chromosome methylation, doubling time, embryoid body formation, teratoma formation, viable chimeric formation, hybridization and differentiation For example, embryonic stem cells).

According to another aspect of the invention, the present invention provides a method for differentiating neural progenitor cells or stem cells into neurons, comprising culturing the composition with neural progenitor cells or stem cells.

The medium used in the culturing step is a universal medium used for culturing neural progenitor cells or stem cells in the prior art. For example, the medium may be Eagle's MEM [Eagle's minimum essensial medium, Eagle, H. Science 130: 142 (1959)], α-MEM [Stanner, CP et al., NAT . New Biol . 230: 52 (1971), Iscove's MEM [Iscove, N. et al., J. Exp . Med . 147: 923 (1978), 199 medium Morgan et al., Proc. Soc . Exp . BioMed . 73: 1 (1950), CMRL 1066, RPMI 1640 [Moore et al., J. Amer . Med . Assoc . 199: 519 (1967), F 12, Ham, Pro . Natl . Acad . Sci . USA 53: 288 (1965), F10 [Ham, RG Exp . Cell Res . 29: 515 (1963), DMEM (Dulbecco's modification of Eagle's medium, Dulbecco, R. et al., Virology 8: 396 (1959)), and mixtures of DMEM and F12 [Barnes, D. et al. , Anal . Biochem . 102: 225 (1980)], Way-mouth's MB752 / 1 [Waymouth, C. J. Natl . Cancer Inst . 22: 1003 (1959), McCoy's 5A [McCoy, TA, et al, Pro . soc . Exp . Bio . Med . 100: 115 (1959), MCDB's series [Ham, RG et al., In Vitro 14:11 (1978), and variations thereof. Detailed description of the medium can be found in R. Ian Freshney, Culture of Animal Cells , A Manual of Basic Technique, Alan R. Liss, Inc., New York, which is incorporated herein by reference. Sodium pyruvate, glutamine, etc. may be additionally added to the culture medium.

It is well known that stem cells can differentiate into different cell types. For example, the neural progenitor cells or stem cells may be differentiated into hematopoietic cells, nerve cells, beta cells, muscle cells, hepatocytes, chondrocytes, epithelial cells, urinary tract cells and similar cells under conditions for cell differentiation. Induced. Media conditions and methods for the differentiation of stem cells are described in Palacios, et al., PNAS . USA , 92: 7530-7537 (1995), Pedersen, J. Reprod . Fertil . Dev . 6; 543-552 (1994), and Bain et al., Dev . Biol , 168: 342-357 (1995).

The neural progenitor cells or stem cells are differentiated into neural cells by 'differentiation stimulation', which is commonly practiced in the art, for example, serum-free medium (Tropepe V et al., Direct neural fate specification). from embryonic stem cells: A primitive mammalian neural stem cell stage acquired through a default mechanism.Neuron. 30: 6578 (2001)), morphogens such as fibroblast growth factors (FGFs), Wnt and retinoic acid (RA). (Ying QL et al. Conversion of embryonic stem cells into neuroectodermal precursors in adherent monoculture. Nat Biotechnol. 21: 183186 (2003)), but is not limited thereto. Under the conditions for differentiation or cell differentiation, the neural progenitor cells or stem cells are differentiated into hematopoietic cells, neurons, beta cells, muscle cells, hepatocytes, chondrocytes, epithelial cells, urinary tract cells and similar cells. Induced. Media conditions and methods for the differentiation of stem cells are described in Palacios, et al., PNAS . USA , 92: 7530-7537 (1995), Pedersen, J. Reprod . Fertil . Dev . 6; 543-552 (1994), and Bain et al., Dev. Biol , 168: 342-357 (1995).

The quinoxaline derivatives or salts thereof of the invention may be used in their pharmaceutically effective amounts; And it can provide a pharmaceutical composition for preventing or treating neurological damage diseases, including a pharmaceutically acceptable salt.

As used herein, the term "nerve damage disease" refers to a disease in which the nerves involved in exercise and sensation are damaged and thus have abnormalities in behavioral functions including exercise and sensation, and such diseases include stroke, Alzheimer's disease, Parkinson's disease, spinal cord injury disease, dementia, demyelination disease, Huntington's disease and amyotrophic spinal sclerosis.

In addition, the term "treatment" as used herein refers to 1) the disease or disorder in which an animal, preferably a mammal, more preferably human, develops, although it has not yet been diagnosed as having a neurological disorder. Prophylaxis, 2) suppression of neurologically damaging diseases, i.e., inhibition of development, and 3) alleviation of neurologically damaging diseases. Specifically, when the pharmaceutical composition is administered in a therapeutically effective amount to a damaged part of a subject in need of treatment of a neuronal cell damage disease, nerve precursor cells or stem cells of the peripheral central nervous system or peripheral nervous system are neurons. Can be used to restore impaired nerve function and treat neurological disorders. Meanwhile, the administration subject may be a mammal including a human.

When the composition of the present invention is manufactured from a pharmaceutical composition, the pharmaceutical composition of the present invention includes a pharmaceutically acceptable carrier. Pharmaceutically acceptable carriers included in the pharmaceutical compositions of the present invention are those commonly used in the preparation, such as lactose, dextrose, sucrose, sorbitol, mannitol, starch, acacia rubber, calcium phosphate, alginate, gelatin, Calcium silicate, microcrystalline cellulose, polyvinylpyrrolidone, cellulose, water, syrup, methyl cellulose, methylhydroxybenzoate, propylhydroxybenzoate, talc, magnesium stearate and mineral oil, and the like It doesn't happen. The pharmaceutical composition of the present invention may further contain a lubricant, a wetting agent, a sweetening agent, a flavoring agent, an emulsifying agent, a suspending agent, a preservative, etc. in addition to the above components. Suitable pharmaceutically acceptable carriers and formulations are described in detail in Remington ' s Pharmaceutical Sciences (19th ed., 1995).

The pharmaceutical composition of the present invention may be administered orally or parenterally, and in the case of parenteral administration, it may be administered by intravenous injection, subcutaneous injection, intramuscular injection, intraperitoneal injection, transdermal administration, or the like.

The appropriate dosage of the pharmaceutical composition of the present invention may vary depending on factors such as the formulation method, administration method, age, body weight, sex, pathological condition, food, administration time, administration route, excretion rate, . The daily dose of the pharmaceutical composition of the present invention is, for example, 0.001-1000 mg / kg. However, the actual dosage of the active ingredient may be determined in consideration of several relevant factors such as the amount of neurons to be differentiated and multiplied, the route of administration, the patient's weight, age and gender, and therefore the dosage may be determined in any form. It does not limit the scope of the invention.

The pharmaceutical compositions of the present invention may be prepared in unit dose form by formulating with a pharmaceutically acceptable carrier and / or excipient according to methods which can be easily carried out by those skilled in the art. Or may be prepared by incorporation into a multi-dose container. The formulations may be in the form of solutions, suspensions, syrups or emulsions in oils or aqueous media, or in the form of excipients, powders, powders, granules, tablets or capsules, and may additionally contain dispersing or stabilizing agents.

The features and advantages of the present invention are summarized as follows:

(Iii) The present invention provides a novel quinoxaline derivative and a composition for inducing differentiation from nerve precursor cells or stem cells to nerve cells comprising the same.

(Ii) Using the quinoxaline derivatives according to the present invention can differentiate neural progenitor cells or stem cells into neurons.

(Iii) In addition, the neural progenitor cells induced by differentiation according to the present invention can be efficiently differentiated into specific cells (eg, dopamine neurons) or oligodendroseite, and are subsequently refractory to neurological diseases (eg, Parkinson's disease or spinal cord injury). Refractory neurological diseases such as) can be applied and provide basic data on drug development.

1 is a result of observing the neural differentiation of mesenchymal stem cells using the compound of the present invention using the Nissle staining method. Each compound used 10 [mu] M and the control group used DMSO.
2 is a numerical representation of the degree of neuronal differentiation of the derivatives of FIG. 1. Compound 13 induced neuronal differentiation in mesenchymal stem cells at 10 μM and compound 15 also induced good neuronal differentiation.
Figure 3 is the result of confirming the expression level of neuronal differentiation markers of mesenchymal stem cells by the compound derivative of the present invention by western blotting.
Figure 4 shows the electrobiochemical results of neurons differentiated from mesenchymal stem cells.

Hereinafter, the present invention will be described in more detail with reference to Examples. It is to be understood by those skilled in the art that these embodiments are only for describing the present invention in more detail and that the scope of the present invention is not limited by these embodiments in accordance with the gist of the present invention .

Synthetic example

Synthetic example  1: Synthesis of Compound 2, Compound 3, Compound 4, and Compound 5

<1-1> 1- (3- Bromo profile )-4- Of methoxybenzene  synthesis

Scheme 1

In 15 ml of dichloromethane (DCM) (Junsei Chemical Co.), 1 g (6 mmol, 1 eq) (Aldrich) of 3- (4-methoxyphenyl) propan-1-ol was dissolved and nitrogen-substituted. Add 2 g (18 mmol, 3 eq) of CBr ₄ (Aldrich). The mixture is added 4.8 g (18 mmol, 3 eq) (Aldrich) PPh ₃ at 0 ° C. After stirring for 1 hour at room temperature, after confirming that all of the 3- (4-methoxyphenyl) propan-1-ol disappeared, NaHCO ₃ aqueous solution was added to extract the organic layer. The organic layer was dried over Na ₂ SO ₄ , concentrated and separated by column chromatography to obtain 460 mg (yield 35%) of the title compound in the form of a colorless oil: ¹ H NMR (CdCl _3, 300 MHz) δ 7.11 (d , J = 8.7 Hz, 2H), 6.80 (d, J = 8.7 Hz, 2H), 3.78 (s, 3H), 3.51 (t, J = 7.3 Hz, 2H), 2.72 (t, J = 7.4Hz, 2H ), 2.71-2.00 (m, 2H).

<1-2> methyl  5- (4- Methoxyphenyl )-2- Of oxopentanoate  synthesis

Scheme 2

Mg 47 mg (1.96 mmol, 1.2 eq) (Aldrich) and I ₂ 10 mg (Aldrich) were added to 10 ml of dried Tetrahydrofuran (THF) (Junsei Chemical Co.) and 1- (3-bromopropyl 450 mg (1.96 mmol, 1.2 eq) of 4-methoxybenzene was added thereto, followed by stirring at room temperature for 30 minutes to prepare a Grignard reagent. Meanwhile, 200 mg (1.63 mmol, 1 eq) (Aldrich) of dimethyl oxalate was dissolved in 5 ml of dried diethyl ether (Junsei Chemical Co.) in another flask, and then cooled to -78 ° C. After slowly adding the Grignard reagent to the flask, the temperature was slowly raised to room temperature. After stirring for 3 hours at 0 ° C., all of the starting materials disappeared, and 2 ml of 1 N HCl solution was added. Thereafter, the organic layer was extracted three times with ether. The organic layer was dried over Na ₂ SO ₄ and separated by column chromatography to give 210 mg (yield 45%) of the title compound in the form of a colorless oil: ¹ H NMR (CdCl _{3 ,} 300 MHz) δ 7.08 (d, J = 8.7 Hz, 2H), 6.82 (d, J = 8.7 Hz, 2H), 3.81 (s, 3H), 3.78 (s, 3H), 2.83 (t, J = 7.3 Hz, 2H), 2.60 (t, J = 7.3 Hz, 2H), 1.99-1.91 (m, 2H).

<1-3> containing isomers methyl  2- (3- (4- Methoxyphenyl ) Propyl) -3-oxo-3,4- Dihydroquinoxaline -6- Carboxylate  synthesis

Scheme 3

150 mg (0.89 mmol, 1 eq) of methyl 3,4-diaminobenzoate (Aldrich) and methyl 5- (4-methoxyphenyl) -2-oxopentanoate 210 in 7 ml of ethanol (Junsei Chemical Co.) Add mg (0.89 mmol, 1 eq), dissolve completely by heating, and stir at room temperature for 12 hours. The reaction mixture was filtered to give 250 mg (yield 80%, including isomer) of the target compound.

<1-4> 2- (3- (4- containing isomer Methoxyphenyl ) -Propyl) -3-oxo-3,4- Dihydroquinoxaline -6- Carboxylic acid  synthesis

Scheme 4

After completely dissolving 200 mg of methyl 2- (3- (4-methoxyphenyl) propyl) -3-oxo-3,4-dihydroquinoxalin-6-carboxylate (including isomers) in 5 ml of tetrahydrofuran, , LiOH.H ₂ O (190 mg, 8 eq) (aldrich), completely dissolved in 5 ml of water, was added. After 5 ml of methanol was dissolved in all, the mixture was stirred at 50 ° C. for 12 hours, and then all of the starting materials disappeared. After completion of the reaction, the reaction was concentrated to remove the solvent and diluted with water. After adjusting the pH to 3-4 with 1 N HCl solution, the organic layer was extracted three times with ethyl acetate. The organic layer was dried over Na ₂ SO ₄ to obtain 190 mg (including isomers, yield 99%) of the title compound as a white solid.

<1-5> 3- (3- (4- Methoxyphenyl ) Propyl) -2-oxo-N-(( Tetrahydro -2H-pyran-2-yl) Oxy ) -1,2- Dihydroquinoxaline -6- Carboxamide (Compound 2) and 2- (3- (4- Methoxyphenyl ) Propyl) -3-oxo-N-(( Tetrahydro -2H-pyran-2-yl) oxy) -3,4- Dihydroquinoxaline -6- Of carboxamide (compound 3)  synthesis

Scheme 5

200 mg (0.59 mmol, 1 eq) of 2- (3- (4-methoxyphenyl) propyl) -3-oxo-3,4-dihydroquinoxalin-6-carboxylic acid containing isomers was added to dimethylformamide. Dissolved in 2 ml of (Dimethylformamide, DMF) (Junsei Chemical Co.), 138 mg (1.18 mmol, 2 eq) (aldrich), 1-ethyl-3- (3- NH ₂ O-THP (Tetrahydropyran-2-yloxyamine) Dimethylaminopropyl) carbodiimide (EDCI) (aldrich) was dissolved by addition of 136 mg (0.71 mmol, 1.2 eq) and 80 mg (0.59 mmol, 1 eq) of hydroxybenzotriazole (HOBT) (aldrich). The mixture was then cooled in an ice bath, and 0.41 ml (2.36 mmol, 4 eq) of diisopropylethylamine (DIPEA) (aldrich) was added. The mixture was stirred at room temperature for 3 hours, diluted with ethyl acetate (Junsei Chemical Co.) and washed with aqueous NH ₄ Cl solution. The organic layer was then dried over Na ₂ SO ₄ and concentrated to separate by column chromatography. 3- (3- (4-methoxyphenyl) propyl) -2-oxo-N-((tetrahydro-2H-pyran-2-yl) oxy) -1,2-dihydroquinoxaline isomer from the organic layer -6-carboxamide (compound 2) and 2- (3- (4-methoxyphenyl) propyl) -3-oxo-N-((tetrahydro-2H-pyran-2-yl) oxy) -3, 4-dihydroquinoxaline-6-carboxamide (Compound 3) was isolated to give 96 mg (white solid, yield 38%) and 20 mg (white solid, yield 8%) of the target compound, respectively: 1) Compound 2: ¹ H NMR (MeOH-d 4 _, 300 MHz) δ 8.15 (d, J = 1.7 Hz, 1H), 7.86 (dd, J = 8.4, 1.7 Hz, 1H), 7.26 (d, J = 8.4 Hz, 1H ), 7.06 (d, J = 8.5 H, 2H), 5.08 (s, 1H), 4.17-4.10 (m, 1H), 3.70 (s, 3H), 3.65-3.61 (m, 1H), 2.84 (t, J = 7.5 Hz, 2H), 2.64 (t, J = 7.5 Hz, 2H), 2.09-1.99 (m, 2H) 1.93-1.61 (m, 6H). 2) Compound 3: ¹ H NMR (MeOH-d 4 _, 300 MHz) δ 7.83 (d, J = 8.3 Hz, 1H), 7.74 (d, J = 1.8 Hz, 1H), 7.66 (dd, J = 8.3, 1.8 Hz, 1H), 7.10 (d, J = 8.6 H, 2H), 6.79 (d, J = 8.6 H, 2H), 5.10 (s, 1H), 4.22-4.10 (m, 1H), 3.73 (s, 3H ), 3.67-3.64 (m, 1H), 2.92 (t, J = 7.5 Hz, 2H), 2.74 (t, J = 7.5 Hz, 2H), 2.13-2.08 (m, 2H) 1.92-1.64 (m, 6H ).

<1-6> N- Hydroxy -2- (3- (4- Methoxyphenyl ) Propyl) -3-oxo-3,4- Dihydroquinoxaline  -6- Carboxamide  Synthesis (Compound 5)

Scheme 6

To 5 ml of methanol and 5 ml of dichloromethane, 2- (3- (4-methoxyphenyl) propyl) -3-oxo-N- (tetrahydro-2H-pyran-2-yloxy) -3,4-dihydro 96 mg (0.22 mmol, 1 eq) of quinoxaline-6-carboxamide was completely dissolved, and 0.32 ml (4.4 mmol, 20 eq) of trifluoroacetic acid (TFA) (aldrich) was added at room temperature. . The mixture was stirred at 35 ° C. for 10 hours and concentrated to give a solid compound which was washed with ethyl acetate to give 40 mg of the target compound (white solid, yield 51%): ¹ H NMR (DMSO-d 6 _, 300 MHz). ) δ 12.43 (brs, 1H), 11.25 (brs, 1H), 9.04 (s, 1H), 8.10 (s, 1H), 7.86 (d, J = 8.6 Hz, 1H), 7.27 (d, J = 8.6) Hz, 1H), 7.13 (d, J = 8.2 Hz, 2H), 6.81 (d, 8.2 Hz, 2H), 3.69 (s, 3H), 2.76 (t, J = 7.5 Hz, 2H), 2.61 (t, 7.5 Hz, 2H), 1.94-1.9 (m, 2H).

<1-7> N- Hydroxy -3- (3- (4- Methoxyphenyl ) Propyl) -2-oxo-1,2- Dihydroquinoxaline -6- Carboxamide  Synthesis (Compound 4)

Scheme 7

Compound 4 is synthesized in “Synthesis of N-hydroxy-2- (3- (4-methoxyphenyl) propyl) -3-oxo-3,4-dihydroquinoxalin-6-carboxamide (Compound 5)” Synthesized according to the described method: ¹ H NMR (MeOH-d 4 _, 300 MHz) δ 7.82 (d, J = 8.5 Hz, 1H), 7.69 (d, J = 1.7 Hz, 1H), 7.63 (dd, J = 8.5 , 1.7 Hz, 1H), 7.13 (d, J = 8.5 H, 2H), 6.78 (d, J = 8.5 H, 2H), 3.73 (s, 3H), 2.92 (t, J = 7.5 Hz, 2H), 2.79 (t, J = 7.5 Hz, 2H), 2.13-2.06 (m, 2H).

Synthetic example  2: Synthesis of Compound 6, Compound 7, Compound 8 and Compound 9

<2-1> methyl  2-oxo-4- Phenylbutanoate  synthesis

Scheme 8

To a mixture of 10 ml of dried tetrahydrofuran, 350 mg (1.2 eq) of Mg and 50 mg of I ₂ was added 2.13 ml (1.2 eq) of (2-bromoethyl) benzene (aldrich) for 30 minutes at room temperature. Stirring prepared the Grignard reagent. Meanwhile, 1.7 g (1 eq) of dimethyl oxalate was added to 10 ml of diethyl ether in another flask to dissolve, and the temperature was lowered to -78 ° C. After slowly adding the Grignard reagent to the flask, the temperature was slowly raised to room temperature. Thereafter, the mixture was stirred at 0 ° C. for 3 hours to confirm that all the starting materials disappeared, and 5 ml of 1 N HCl solution was added. The organic layer was extracted three times with ether. The organic layer was dried over Na ₂ SO ₄ , separated by column chromatography, and finally 760 mg (yield 30%) of the title compound was obtained as a colorless oil: ¹ H NMR (CdCl _{3 ,} 300 MHz) δ 7.32-7.18 (m, 5H), 3.85 (s, 3H), 3.19 (t, J = 7.4 Hz, 2H), 2.91 (t, J = 7.4 Hz, 2H).

<2-2> isomers included methyl  3-oxo-2- Penetil -3,4- Dihydroquinoxaline -6- Carboxylate  synthesis

Scheme 9

To 30 ml of ethanol, 657 mg (3.95 mmol, 1 eq) of methyl 3,4-diaminobenzoate and 760 mg (3.95 mmol, 1 eq) of methyl 2-oxo-4-phenylbutanoate were added and dissolved completely. After stirring at room temperature for 12 hours. The reaction was filtered to give 1100 mg (90% yield, including isomer) of the target compound.

<2-3> 3-oxo-2- with isomer Penetil -3,4- Dihydroquinoxaline -6- Carboxylic acid  synthesis

Scheme 10

After 5 ml of tetrahydrofuran, 175 mg (0.57 mmol, 1 eq) of methyl 3-oxo-2-phenethyl-3,4-dihydroquinoxaline-6-carboxylate (including isomers) was added thereto to completely dissolve. LiOH.H ₂ O (190 mg, 8 eq) was added completely dissolved in 5 ml of water. To the mixture was added 5 ml of methanol and stirred at 50 ° C. for 12 hours to confirm that all starting materials disappeared. After completion of the reaction, the reaction was concentrated to remove solvent and diluted with water. 1N HCl solution was added to the diluted solution to adjust the pH to 3-4, and the organic layer was extracted three times with ethyl acetate. The organic layer was dried over Na ₂ SO ₄ to give 160 mg (including isomers, yield 95%) of the title compound as a white solid.

<2-4> 3-oxo-2- Penetil -N- ( Tetrahydro -2H-pyran-2- Sake ) -3,4- Dihydroquinoxaline -6- Carboxamide (Compound 7) and 2-oxo-3- Penetil -N- ( Tetrahydro -2H-pyran-2- Sake ) -1,2- Dihydroquinoxaline -6- Of carboxamide (compound 6)  synthesis

Scheme 11

Dissolve 173 mg (0.59 mmol, 1 eq) of 3-oxo-2-phenethyl-3,4-dihydroquinoxaline-6-carboxylic acid with isomers in 2 ml of dimethylformamide and 138 mg NH ₂ OTHP (1.18 mmol, 2 eq), 136 mg (0.71 mmol, 1.2 eq) of EDCI and 80 mg (0.59 mmol, 1 eq) of HOBT were added and dissolved, and then transferred to an ice bath for cooling. 0.41 mL (2.36 mmol, 4 eq) of diisopropylethylamine was added to the cooled mixture, stirred at room temperature for 3 hours, and diluted with ethyl acetate. The diluted solution was washed with aqueous NH ₄ Cl solution, the organic layer was dried over Na ₂ SO ₄ , concentrated and separated by column chromatography. 3-oxo-2-phenethyl-N- (tetrahydro-2H-pyran-2-yloxy) -3,4-dihydroquinoxalin-6-carboxamide (Compound 7) and 2 isomers from the organic layer -Oxo-3-phenethyl-N- (tetrahydro-2H-pyran-2-yloxy) -1,2-dihydroquinoxaline-6-carboxamide (Compound 6) was isolated and 63 mg each (white Solid, yield 27%) and 20 mg (white solid, yield 8%) were obtained: 1) Compound 7: ¹ H NMR (MeOH-d 4 _, 300 MHz) δ 8.10 (d, J = 1.8 Hz, 1H) , 7.80 (dd, J = 8.5, 1.8 Hz, 1H), 7.24 (d, J = 8.5 Hz, 1H), 7.17-7.15 (m, 4H), 7.07-7.02 (m, 1H), 4.98 (s, 1H ), 4.09-4.01 (m, 1H), 3.57 -3.52 (m, 1H), 3.12-2.97 (m, 4H), 1.84-1.53 (m, 6H). 2) Compound 6: ¹ H NMR (MeOH-d 4 _, 300 MHz) δ7.74 (d, J = 8.4 Hz, 1H), 7.61 (d, J = 1.8 Hz, 1H), 7.56 (dd, J = 8.4, 1.8 Hz, 1H), 7.18-7.14 (m, 4H), 7.08-7.05 (m, 1H), 4.98 (s, 1H), 4.09-4.00 (m, 1H), 3.57-3.45 (m, 1H), 3.14- 2.97 (m, 4 H) 1.81-1.46 (m, 6 H).

<2-5> N- Hydroxy -3-oxo-2- Petetyl -3,4- Dihydroquinoxaline -6- Of carboxamides (compound 9)  synthesis

Scheme 12

63 g of 3-oxo-2-phenethyl-N- (tetrahydro-2H-pyran-2-yloxy) -3,4-dihydroquinoxaline-6-carboxamide in 4 ml of methanol and 4 ml of dichloromethane (0.16 mmol, 1 eq) was added to completely dissolve it, and then 0.24 mL (3.2 mmol, 20 eq) of tetrahydrofuran was added at room temperature. The mixture was stirred at 35 ° C. for 10 hours, concentrated to give a solid compound which was washed with ethyl acetate to give 25 mg (white solid, yield 51%) of the title compound: ¹ H NMR (DMSO-d6 _, 300 MHz) δ 12.42 (brs, 1H), 11.29 (brs, 1H), 9.05 (s, 1H), 8.12 (s, 1H), 7.87 (dd, J = 8.5, 1.7 Hz, 1H), 7.31 (s, 1H), 7.31 (s, 1H), 7.26-7.28 (m, 4H), 7.15-7.17 (m, 1H), 3.04-3.06 (m, 4H).

<2-6> N- Hydroxy -2-oxo-3- Penetil -1,2- Dihydroquinoxaline -6- Of carboxamide (compound 8)  synthesis

Scheme 13

Compound 8 was synthesized according to the method described in “Synthesis of N-hydroxy-3-oxo-2-pettetyl-3,4-dihydroquinoxalin-6-carboxamide (Compound 9)”: ¹ H NMR (MeOH-d 4 _, 300 MHz) δ 7.84 (d, J = 8.5 Hz, 1H), 7.71 (s, 1H), 7.64 (d, J = 8.5 Hz, 1H), 7.28-7.26 (m, 4H ), 7.19-7.16 (m, 1 H), 3.24-3.09 (m, 4 H).

Synthetic example  3: Synthesis of Compound 10, Compound 11, Compound 12, and Compound 13

<3-1> methyl  2-oxo-6- Of phenylhexanoate  synthesis

Scheme 14

To a mixture of 10 ml of dried tetrahydrofuran, 350 mg (1.2 eq) of Mg, and 50 mg of I _2, 1.27 ml (1.2 eq) of (4-bromobutyl) benzene was added, followed by stirring at room temperature for 30 minutes to give Greenide. Reagents were prepared. On the other hand, 10 ml of diethyl ether dried in 1.7 g (1 eq) of dimethyl oxalate was added to another flask and dissolved, and the mixture was cooled to -78 ° C. After slowly adding the Grignard reagent to the cooled mixture, the temperature was slowly raised to room temperature. After stirring for 3 hours at 0 ℃ to confirm that all the starting material disappeared, 5 ml of 1 N HCl solution was added. The organic layer was extracted three times with ether, and the organic layer was dried over Na ₂ SO ₄ and separated by column chromatography to obtain 1140 mg (yield 35%) of the target compound in the form of a colorless oil: ¹ H NMR (cdcl 3 _, 300 MHz). ) δ 7.30-7.15 (m, 5H), 3.85 (s, 3H), 2.89-2.84 (m, 2H), 2.66-2.61 (m, 2H), 1.69-1.6 (m, 4H).

<3-2> methyl  2-oxo-3- (4- Phenylbutyl ) -1,2- Dihydroquinoxaline -6- Carboxylate  Synthesis (Compound 10)

Scheme 15

Into 30 ml of ethanol, 754 mg (4.54 mmol, 1 eq) of methyl 3,4-diaminobenzoate and 1000 mg (4.54 mmol, 1 eq) of methyl 2-oxo-6-phenylhexanoate were added and heated with a heat gun. After complete melting, the mixture was stirred at room temperature for 12 hours. The resulting white solid was filtered to give 760 mg (yield 50%, isomer separation) of the desired product: ¹ H NMR (dmso-d6, 300 MHz) δ 12.58 (s, 1H), 8.20 (d, J = 1.8 Hz, 1H), 8.01 (dd, J = 8.5, 1.8 Hz, 1H), 7.34 (d, J = 8.5 Hz, 1H), 7.29-7.13 (m, 5H), 3.87 (s, 3H), 2.83 (t, J = 7.3 Hz, 2H), 2.63 (t, J = 7.3 Hz, 2H), 1.76-1.64 (m, 4H).

<3-3> 2-oxo-3- (4- Phenylbutyl ) -1,2- Dihydroquinoxaline -6- Carboxylic acid  Synthesis (Compound 11)

Scheme 16

To 5 ml of tetrahydrofuran add 190 mg (0.57 mmol, 1 eq) of methyl 2-oxo-3- (4-phenylbutyl) -1,2-dihydroquinoxalin-6-carboxylate (including isomers) After dissolving, LiOH.H ₂ O (190 mg, 8 eq) was added completely dissolved in 5 ml of water. After adding 5 ml of methanol to the mixture to adjust the clear phase, the mixture was stirred at 50 ° C. for 12 hours to confirm that all the delamination materials disappeared. After the reaction was completed, the reaction was concentrated to remove the solvent and diluted with water. 1N HCl solution was added to the diluted solution to adjust the pH to 3-4, and the organic layer was extracted three times with ethyl acetate. The organic layer was dried over Na ₂ SO ₄ to give 150 mg (yield 80%) of the title compound as a white solid: ¹ H NMR (dmso-d 6, 300 MHz) δ 12.55 (s, 1H), 8.17 (d, J = 1.8 Hz, 1H), 7.98 (d, J = 8.5, 1.8 Hz, 1H), 7.25 (d, J = 8.5 Hz, 1H), 7.16-7.12 (m, 5H), 2.80 (t, J = 7.2 Hz, 2H), 2.61 (t, J = 7.2 Hz, 2H), 1.65-1.67 (m, 4H).

<3-4> 2-oxo-3- (4- Phenylbutyl ) -N- ( Tetrahydro -2H-pyran-2- Sake ) -1,2- Dihydroquinoxaline -6- Carboxamide  Synthesis (Compound 12)

Scheme 17

40 mg (0.12 mmol, 1 eq) of 2-oxo-3- (4-phenylbutyl) -1,2-dihydroquinoxaline-6-carboxylic acid was dissolved in 1 ml of dimethylformamide and 28 mg of NH ₂ OTHP. (0.24 mmol, 2 eq), 28 mg (0.14 mmol, 1.2 eq) of EDCI and 17 mg (0.12 mmol, 1 eq) of HOBT were added and dissolved. Thereafter, the mixture was transferred to an ice bath to lower the temperature, 0.085 mL (0.48 mmol, 4 eq) of diisopropylethylamine was added thereto, and the mixture was stirred at room temperature for 3 hours. The mixture was diluted with ethyl acetate and washed with aqueous NH ₄ Cl solution. The organic layer was then dried over Na ₂ SO ₄ and separated by column chromatography to give 30 mg (white solid, yield 60%) of the title compound: ¹ H NMR (MeOH-d 4 _, 300 MHz) δ 8.09 (d, J = 1.7 Hz, 1H), 7.80 (dd, J = 8.5, 1.7 Hz, 1H), 7.23 (d, J = 8.5 Hz, 1H), 7.15-7.02 (m, 5H), 4.98 (s, 1H), 4.07- 4.03 (m, 1 H), 3.55 -3.52 (m, 1 H), 2.82 (t J = 7.4 Hz, 2H), 2.57 (t J = 7.4 Hz, 2H), 1.83-1.49 (m, 10H).

<3-5> N- Hydroxy -2-oxo-3- (4- Phenylbutyl ) -1,2- Dihydroquinoxaline -6- Carboxamide  Synthesis (Compound 13)

Scheme 18

2-oxo-3- (4-phenylbutyl) -N- (tetrahydro-2H-pyran-2-yloxy) -1,2-dihydroquinoxaline-6-carboxin 4 ml of methanol and 4 ml of dichloromethane 30 mg (0.07 mmol, 1 eq) of radiamide was completely dissolved and 0.11 mL (1.42 mmol, 20 eq) of trifluoroacetic acid was added at room temperature. The mixture was stirred at 35 ° C. for 10 hours, concentrated to afford a solid compound which was washed with ethyl acetate to give 10 mg (white solid, yield 42%) of the title compound: ¹ H NMR (DMSO-d6 _, 300 MHz) δ 12.43 (brs, 1H), 11.28 (brs, 1H), 9.05 (s, 1H), 8.15 (d, J = 1.7 Hz, 1H), 7.87 (dd, J = 8.5, 1.7 Hz, 1H) , 7.30 (d, J = 8.5, Hz, 1H), 7.29-7.16 (m, 5H), 2.82 (t, J = 7.4 Hz, 2H), 2.64 (t, J = 7.4 Hz, 2H), 1.73-1.67 (m, 4 H).

Synthetic example  4: Synthesis of Compound 14 and Compound 15

<4-1> 3- (4- Fluorophenyl ) Propane-1-ol

Scheme 19

Lithium aluminum hydride (LAH) (aldrich) 720 mg (18 mmol, 3 eq) was added to 20 ml of dried tetrahydrofuran, transferred to an ice bath, and then 1 g (6) of 4-fluorocinnamic acid. mmol, 1 eq) (aldrich) was dissolved in 20 ml of dried tetrahydrofuran dropwise and refluxed for 4 hours. When all of the starting material had disappeared, 10 ml of methanol was added to terminate the reaction and concentrated. After neutralization with 1 N HCl, washing with ethyl acetate three times, concentration and column separation gave 470 mg (yield 50%) of the title compound: ¹ H NMR (CdCl _{3 ,} 300 MHz) δ 7.17-6.79 (m , 4H), 3.69 (t, J = 7.4 Hz, 2H), 2.70 (t, J = 7.4 Hz, 2H), 1.89-1.80 (m, 2H).

<4-2> 1- (3- Bromo profile )-4- Fluorobenzene  synthesis

Scheme 20

470 mg (3.05 mmol, 1 eq) of 3- (4-fluorophenyl) propan-1-ol was dissolved in 10 ml of dichloromethane, and nitrogen was substituted. Then, 3 g (9.15 mmol, 3 eq) of CBr ₄ was added thereto. . After placing the mixture on an ice bath, 2.4 g (9.15 mmol, 3 eq) of PPh ₃ was added thereto, and stirred at room temperature for 1 hour to confirm that all starting materials disappeared. Then, NaHCO ₃ solution was added to extract the organic layer. The organic layer was dried over Na ₂ SO ₄ , concentrated and separated by column chromatography to obtain 540 mg (yield 81%) of the target compound in the form of a colorless oil: ¹ H NMR (CdCl _{3 ,} 300 MHz) δ 7.17-7.00 (m, 4H), 3.41-3.37 (m, 2H), 2.78-2.75 (m, 2H), 2.75-2.20 (m, 2H).

<4-3> methyl  5- (4- Fluorophenyl )-2- Of oxopentanoate  synthesis

Scheme 21

540 mg (2.48 mmol, 1.2 eq) of 1- (3-bromopropyl) -4-fluorobenzene in a mixture of 5 ml dried tetrahydrofuran, 60 mg (2.48 mmol, 1.2 eq) and I ₂ 10 mg After the addition, the mixture was stirred at room temperature for 30 minutes to prepare a Grignard reagent. Meanwhile, 244 mg (2.06 mmol, 1 eq) of dimethyl oxalate was added to another 5 ml of dried diethyl ether, and the mixture was cooled to -78 ° C. After slowly adding the Grignard reagent to the cooled mixture, the temperature was slowly raised to room temperature. After stirring for 3 hours at 0 ℃ to confirm that all the starting material disappeared, 2 ml of 1 N HCl solution was added. The organic layer was extracted three times with ether, and the organic layer was dried over Na ₂ SO ₄ and separated by column chromatography to obtain 40 mg (yield 7%) of the target compound in the form of a colorless oil.

<4-4> methyl  3- (3- (4- Fluorophenyl ) Propyl) -2-oxo-1,2- Dihydroquinoxaline -6- Carboxylate  Synthesis (Compound 14)

Scheme 22

30 mg (0.18 mmol, 1 eq) of methyl 3,4-diaminobenzoate and 40 mg (0.18 mmol, 1 eq) of methyl 5- (4-fluorophenyl) -2-oxopentanate were added to 2 ml of ethanol. The mixture was heated with a heat gun and completely dissolved, followed by stirring at room temperature for 12 hours. Filtration of the reaction product white solid gave 7 mg (yield 12%) of the title compound: ¹ H NMR (MeOH-d 4 _, 300 MHz) δ 8.37 (d, J = 1.7 Hz, 1H), 8.07 (dd, J = 8.5, 1.7 Hz, 1H), 7.31 (d, J = 8.5 Hz, 1H), 7.24-7.20 (m, 2H), 6.98-6.92 (m, 2H), 3.93 (s, 3H), 2.89 (t, J = 7.5 Hz, 2H), 2.74 (t, J = 7.5 Hz, 2H), 2.17-2.05 (m, 2H).

<4-5> 3- (3- (4- Fluorophenyl ) Propyl) -2-oxo-1,2- Dihydroquinoxaline -6- Carboxylic acid  synthesis

Scheme 23

In 0.5 ml of tetrahydrofuran, 7 mg of methyl 3- (3- (4-fluorophenyl) propyl) -2-oxo-1,2-dihydroquinoxaline-6-carboxylate was completely dissolved, followed by LiOH.H. ₂ O (6 mg, 8 eq) was added completely in 0.5 ml of water. After adding 0.5 ml of methanol to adjust the clear phase (clear phase), the mixture was stirred for 12 hours at 50 ℃, it was confirmed that all the starting material disappeared. After completion of the reaction, the reaction was concentrated to remove solvent and diluted with water. After adjusting the pH to 3-4 with 1 N HCl solution, the organic layer was extracted three times with ethyl acetate. The organic layer was dried over Na ₂ SO ₄ to give 6 mg (yield 90%) of the title compound as a white solid: ¹ H NMR (MeOH-d 4 _, 300 MHz) δ 8.28 (d, J = 1.7 Hz, 1H), 8.00 (dd, J = 8.5, 1.7 Hz, 1H), 7.20 (d, J = 8.5 Hz, 1H), 7.14-7.10 (m, 2H), 6.88-6.82 (m, 2H), 2.78 (t, J = 7.5 Hz, 2H), 2.64 (t, J = 7.5 Hz, 2H), 2.03-1.98 (m, 2H).

<4-6> 3- (3- (4- Fluorophenyl ) Propyl) -2-oxo-N- ( Tetrahydro -2H-pyran-2- Sake ) -1,2- Dihydroquinoxaline -6- Carboxamide  synthesis

Scheme 24

6 mg (0.022 mmol, 1 eq) of 3- (3- (4-fluorophenyl) propyl) -2-oxo-1,2-dihydroquinoxaline-6-carboxylic acid was dissolved in 2 ml of dimethylformamide. Dissolved by adding 5 mg (0.043 mmol, 2 eq) of NH ₂ OTHP, 5 mg (0.026 mmol, 1.2 eq) of EDCI, and 3 mg (0.022 mmol, 1 eq) of HOBT. The mixture was transferred to an ice bath for cooling and 0.014 mL (0.086 mmol, 4 eq) of diisopropylethylamine was added. After stirring at room temperature for 3 hours, the mixture was diluted with ethyl acetate and washed with aqueous NH ₄ Cl solution. The organic layer was then dried over Na ₂ SO ₄ , concentrated and separated by column chromatography, and the desired compound was obtained 7 mg (yield 77%): ¹ H NMR (MeOH-d 4 _, 300 MHz) δ 8.19 (d, J = 1.8 Hz, 1H), 7.91 (dd, J = 8.6, 1.8 Hz, 1H), 7.31 (d, J = 8.6 Hz, 1H), 7.24-7.19 (m, 2H), 6.98-6.92 (m, 2H), 5.08 (s, 1H), 4.21-4.10 (m, 1H), 3.71-3.62 (m, 1H), 2.89 (t, J = 7.5 Hz, 2H), 2.73 (t, J = 7.5 Hz, 2H), 2.15 -2.05 (m, 2 H) 1.91-1.58 (m, 6 H).

<4-7> 3- (3- (4- Fluorophenyl ) Profile) -N- Hydroxy -2-oxo-1,2- Dihydroquinoxaline -6- Carboxamide  Synthesis (Compound 15)

Scheme 25

3- (3- (4-fluorophenyl) propyl) -2-oxo-N- (tetrahydro-2H-pyran-2-yloxy) -1,2-dihydro in 0.5 ml of methanol and 0.5 ml of dichloromethane After 7 mg (0.016 mmol, 1 eq) of quinoxaline-6-carboxamide was completely dissolved, 0.024 mL (0.33 mmol, 20 eq) of trifluoroacetic acid was added at room temperature. The mixture was stirred at 35 ° C. for 10 hours and the concentrated solid was washed with ethyl acetate to give 4 mg (white solid, yield 71%) of the title compound: ¹ H NMR (MeOH-d 4 _, 300 MHz) δ 8.18 (s, 1H), 7.91 (d, J = 8.5 Hz, 1H), 7.34 (d, J = 8.5 Hz, 1H), 7.26-7.21 (m, 2H), 7.00-6.94 (m, 2H), 2.92 (t, J = 7.4 Hz, 2H), 2.76 (t, J = 7.4 Hz, 2H), 2.15-2.10 (m, 2H).

Synthetic example  5: Synthesis of Compound 16

<5-1> 5- Phenylpentane Synthesis of -1-ol

Scheme 26

197 mg (0.0052 mol, 2 eq) of lithium ammonium hydride was added to 10 ml of dried tetrahydrofuran and transferred into an ice bath. Then, 500 mg (0.0026 mol, 1 eq) of methyl5-phenylpentanoate was dried. It was dissolved in 10 ml of furan and added dropwise. It was then refluxed for 4 hours. When all of the starting material had disappeared, 10 ml of methanol was added to terminate the reaction and concentrated. To the concentrated reaction was neutralized by addition of 1 N HCl, and then washed three times with ethyl acetate. The organic layer was then concentrated and separated by column chromatography to give 330 mg (yield 78%) of the title compound: ¹ H NMR (CdCl _{3 ,} 300 MHz) δ 7.30-7.25 (m, 2H), 7.19-7.14 (m, 3H ), 3.63 (t, J = 6.7 Hz, 2H), 2.62 (t, J = 7.5 Hz, 2H), 1.68-1.55 (m, 4H), 1.45-1.37 (m, 2H).

<5-2> (5- Bromopentyl Synthesis of Benzene

Scheme 27

330 mg (2 mmol, 1eq) of 5-phenylpentan-1-ol was dissolved in 10 ml of dichloromethane, and the resultant was nitrogen-substituted, and 2 g (6 mmol, 3 eq) of CBr ₄ was added thereto. After the mixture was transferred to an ice pear tm, PPh ₃ 1.6 g (2 mmol, 3 eq) was added, and stirred at room temperature for 1 hour to confirm that all SM disappeared. Thereafter, an aqueous NaHCO ₃ solution was added to extract the organic layer. The organic layer was dried over Na ₂ SO ₄ , concentrated and separated by column chromatography, and 450 mg (quantitative) of the title compound was obtained as a colorless oil: ¹ H NMR (CdCl _3, 300 MHz) δ 7.30-7.25 (m , 2H), 7.20-7.16 (m, 3H), 3.40 (t, J = 6.6 Hz, 2H), 2.62 (t, J = 7.5 Hz, 2H), 1.91-1.80 (m, 2H), 1.67-1.60 ( m, 2H), 1.52-1.45 (m, 2H).

<5-3> methyl  2-oxo-7- Of phenylheptanoate  synthesis

Scheme 28

To a mixture of 5 ml of dried tetrahydrofuran, 47 mg (1.98 mmol, 1.2 eq) and 5 mg of I ₂ was added 450 mg (1.98 mmol, 1.2 eq) of (5-bromopentyl) benzene, followed by room temperature. Stirring for 30 minutes gave Grignard reagent. Meanwhile, in another flask, 200 mg (1.65 mmol, 1eq) of dimethyl oxalate was added to 5 ml of dried diethyl ether, and the temperature was lowered to -78 ° C. After slowly adding the Grignard reagent to the cooled mixture, the temperature was slowly raised to room temperature. After stirring for 3 hours at 0 ℃ to confirm that all the starting material disappeared, 3 ml of 1 N HCl solution was added, and the organic layer was extracted three times with ether. The organic layer was dried over Na ₂ SO ₄ and separated by column chromatography to obtain 70 mg (yield 7%) of the title compound in the form of a colorless oil: ¹ H NMR (CdCl _{3 ,} 300 MHz) δ 7.30-7.24 (m , 2H), 7.20-7.14 (m, 3H), 3.85 (s, 3H), 2.83 (t, J = 7.3 Hz, 2H), 2.64-2.58 (m, 2H), 1.69-1.58 (m, 4H), 1.42-1.34 (m, 2 H).

<5-4> methyl  2-oxo-3- (5- Phenylpentyl ) -1,2- Dihydroquinoxaline -6- Carboxylate  synthesis

Scheme 29

33 mg (0.2 mmol, 1 eq) of methyl 3,4-diaminobenzoate and 70 mg (0.2 mmol, 1 eq) of methyl 2-oxo-7-phenylheptanoate were added to 3 ml of ethanol, and dissolved completely by heating. Stir at room temperature for 12 hours. The reaction mixture was filtered to give 20 mg (yield 28%) of the title compound: ¹ H NMR (MeOH-d 4 _, 300 MHz) δ 8.26 (d, J = 1.7 Hz, 1H), 7.98 (dd, J = 8.5, 1.7 Hz, 1H), 7.22 (d, J = 8.5 Hz, 1H), 7.13-6.97 (m, 5H), 3.84 (s, 3H), 2.77 (t, J = 7.7 Hz, 2H), 2.52 ( t, J = 7.7 Hz, 2H), 1.75-1.64 (m, 2H), 1.62-1.53 (m, 1.40-1.30 (m, 2H).

<5-5> 2-oxo-3- (5- Phenylpentyl ) -1,2- Dihydroquinoxaline -6- Carboxylic acid  synthesis

Scheme 30

20 ml of methyl 2-oxo-3- (5-phenylpentyl) -1,2-dihydroquinoxalin-6-carboxylate was completely dissolved in 1 ml of tetrahydrofuran, followed by LiOH.H ₂ O (18 mg, 8 eq) was added completely in 1 ml of water. Then 1 ml of methanol was added to adjust the clear phase. After stirring the mixture at 50 ° C. for 12 h, it was confirmed that all starting materials disappeared. After completion of the reaction, the reaction was concentrated to remove solvent and diluted with water. After adjusting the pH to 3-4 with 1 N HCl solution, the organic layer was extracted three times with ethyl acetate. The organic layer was dried over Na ₂ SO ₄ to give 13 mg (yield 80%) of the title compound as a white solid: ¹ H NMR (MeOH-d 4 _, 300 MHz) δ 8.30 (d, J = 1.7 Hz, 1H), 7.99 (dd, J = 8.5, 1.7 Hz, 1H), 7.21 (d, J = 8.5 Hz, 1H), 7.13-6.97 (m, 5H), 2.77 (t, J = 7.5 Hz, 2H), 2.52 (t, J = 7.5 Hz, 2H), 1.77-1.68 (m, 2H), 1.64-1.54 (m, 2H), 1.40-1.33 (m, 2H).

<5-6> 2-oxo-3- (5- Phenylpentyl ) -N- ( Tetrahydro -2H-pyran-2- Sake ) -1,2- Dihydroquinoxaline -6- Carboxamide  synthesis

Scheme 31

13 mg (0.038 mmol, 1 eq) of 2-oxo-3- (5-phenylpentyl) -1,2-dihydroquinoxaline-6-carboxylic acid was dissolved in 2 ml of dimethylformamide and 9 mg of NH ₂ OTHP. (0.077 mmol, 2 eq), 8 mg (0.045 mmol, 1.2 eq) of EDCI and 5 mg (0.038 mmol, 1 eq) of HOBT were added and dissolved. It was then transferred to an ice bath and cooled, and 0.026 mL (0.152 mmol, 4 eq) of diisopropylethylamine was added. The mixture was stirred at room temperature for 3 hours, diluted with ethyl acetate and washed with aqueous NH ₄ Cl solution. The organic layer was then dried over Na ₂ SO ₄ and concentrated, which was then separated by column chromatography to give 10 mg (yield 62%) of the title compound: ¹ H NMR (MeOH-d 4 _, 300 MHz) δ 8.09 (d, J = 1.8 Hz, 1H), 7.80 (dd, J = 8.5, 1.8 Hz, 1H), 7.23 (d, J = 8.5 Hz, 1H), 7.13-6.99 (m, 5H), 4.98 (m, 1H), 4.12 -4.10 (m, 1H), 3.57-3.52 (m, 1H), 2.75 (t, J = 7.5 Hz, 2H), 2.52 (t, J = 7.5 Hz, 2H), 1.18-1.16 (m, 2H) 1.61 -1.13 (m, 10 H).

<5-7> N- Hydroxy -2-oxo-3- (5- Phenylpentyl ) -1,2- Dihydroquinoxaline -6- Carboxamide  Synthesis (Compound 16)

Scheme 32

To 1 ml of methanol and 1 ml of dichloromethane, 2-oxo-3- (5-phenylpentyl) -N- (tetrahydro-2H-pyran-2-yloxy) -1,2-dihydroquinoxaline-6-carr 10 mg (0.023 mmol, 1 eq) of radiamide was completely dissolved, and 0.034 mL (0.046 mmol, 20 eq) of tetrahydrofuran was added at room temperature. The mixture was stirred at 35 ° C. for 10 hours and the concentrated solid was washed with ethyl acetate to give 4 mg (white solid, yield 50%) of the title compound: ¹ H NMR (MeOH-d 4 _, 300 MHz) δ 8.15 (s, 1H), 7.87 (d, J = 8.5 Hz, 1H), 7.32 (d, J = 8.5 Hz, 1H), 7.23-7.09 (m, 5H), 2.87 (t, J = 7.4 Hz, 2H ), 2.62 (t, J = 7.4 Hz, 2H), 1.84-1.79 (m, 2H), 1.71-1.64 (m, 2H), 1.45-1.42 (m, 2H).

Synthetic example  6: Synthesis of Compound 17 and Compound 18

<6-1> methyl  One- methyl -2-oxo-3- (3- Phenylpropyl ) -1,2- Dihydroquinoxaline -6- Carboxylate  synthesis

Scheme 33

Methyl 2-oxo-3- (3-phenylpropyl) -1,2-dihydroquinoxaline-6-carboxylate (100 mg, 0.31 mmol) was dissolved in acetone (10 mL) (Junsei Chemical Co.) , K ₂ CO ₃ (52 mg, 0.372 mmol) (Junsei Chemical Co.) and MeI (excess, excess) (aldrich) were added, followed by reaction under reflux for 12 hours. After the reaction was completed, the organic layer was extracted with ethyl acetate / NaCl. The organic layer was dried over MgSO 4 and concentrated to methyl 1-methyl-2-oxo-3- (3-phenylpropyl) -1,2-dihydro by column chromatography (ethyl acetate: hexane = 1: 3). Quinoxaline-6-carboxylate (90 mg, 0.267 mmol, yield 86%) was obtained: ¹ H NMR (300 MHz, CDCl ₃ ): δ 8.51 (d, J = 2.1 Hz, 1H), 8.17 (dd) , J = 8.7Hz, 2.1Hz, 1H), 7.32 (d, J = 8.7Hz, 1H), 7.29 ~ 7.14 (m, 5H), 3.96 (s, 3H), 3.70 (s, 3H), 3.03 ~ 2.98 (m, 2H), 2.81-2.75 (m, 2H), 2.21-2.11 (m, 2H).

<6-2> 1- methyl -2-oxo-3- (3- Phenylpropyl ) -1,2- Dihydroquinoxaline -6- Carboxylic acid  synthesis

Scheme 34

Methyl 1-methyl-2-oxo-3- (3-phenylpropyl) -1,2-dihydroquinoxaline-6-carboxylate (390 mg, 1.16 mmol) was dissolved in tetrahydrofuran (10 mL) and methanol ( 10 mL), and a solution of LiOH (244 mg, 5.8 mmol) in water (10 mL) was added dropwise, followed by stirring at 50 ° C for 4 hours. After the reaction was completed, pH was adjusted to 2 by adding 2M HCl aqueous solution. Then, the organic layer was extracted with ethyl acetate, the organic layer was dried over MgSO 4, and concentrated to methyl 1-methyl-2-oxo-3- (3-phenylpropyl) -1,2-dihydroquinoxaline-6-carboxyl. Acid (150 mg, 0.46 mmol, yield 40%, white solid) was obtained: ¹ H NMR (300 MHz, CDCl ₃ ): δ 13.06 (br, OH), 8.21 (d, J = 1.7 Hz, 1H), 8.06 (dd, J = 8.8Hz, 1.7Hz, 1H), 7.59 (d, J = 8.8Hz, 1H), 7.28 ~ 7.12 (m, 5H), 3.60 (s, 3H), 2.84 ~ 2.79 (m, 2H ), 2.70-2.65 (m, 2H), 2.06-1.96 (m, 2H).

<6-3> 1- methyl -2-oxo-3- (3- Phenylpropyl ) -N- ( Tetrahydro -2H-pyran-2- Sake ) -1,2- Dihydroquinoxaline Synthesis of -6-carboxamide

Scheme 35

1-Methyl-2-oxo-3- (3-phenylpropyl) -1,2-dihydroquinoxaline-6-carboxylic acid (150 mg, 0.47 mmol) was dissolved in dichloromethane (1 mL), and then EDCI (109 mg, 0.57 mmol), 1-HOBT (64 mg, 0.47 mmol), NH ₂ OTHP (110 mg, 0.94 mmol) and diisopropylethylamine (243 mg, 1.88 mmol) were added at room temperature for 4 hours. Stirred. After the reaction was completed, the organic layer was extracted with ethyl acetate / NH ₄ Cl. The organic layer was dried over MgSO 4, concentrated and then purified by column chromatography (ethyl acetate: hexane = 4: 1) using 1-methyl-2-oxo-3- (3-phenylpropyl) -N- (tetrahydro). -2H-pyran-2-yloxy) -1,2-dihydroquinoxalin-6-carboxamide (100 mg, 0.237 mmol, yield 50%) was obtained: ¹ H NMR (300 MHz, DMSO-d6 ): δ 11.78 (br, NH), 8.20 (m, 1H), 7.97 ~ 7.94 (m, 1H), 7.61 ~ 7.58 (m, 1H), 7.29 ~ 7.13 (m, 5H), 5.01 (s, 1H) , 4.09 to 4.01 (m, 1H), 3.60 (s, 3H), 3.54 to 3.50 (m, 1H), 2.85 to 2.80 (m, 2H), 2.71 to 2.66 (m, 2H), 2.06 to 1.97 (m, 2H), 1.71-1.54 (m, 6H).

<6-4> N- Hydroxy1 - methyl -2-oxo-3- (3- Phenylpropyl ) -1,2- Dihydroquinoxaline -6- Carboxamide  Synthesis (Compound 17)

Scheme 36

1-methyl-2-oxo-3- (3-phenylpropyl) -N- (tetrahydro-2H-pyran-2-yloxy) -1,2-dihydroquinoxaline-6-carboxamide (100 mg , 0.237 mmol) was dissolved in 2 ml of methylene chloride (MC) (Junsei Chemical Co) and 2 ml of methanol, followed by addition of trifluoroacetic acid (540 mg, 4.74 mmol) for 12 hours at 50 ° C. . The reaction was concentrated and a solid compound was obtained under ether and hexane (Junsei Chemical Co), and then filtered to give N-hydroxy-1-methyl-2-oxo-3- (3-phenylpropyl) -1, 2-dihydroquinoxaline-6-carboxamide (22 mg) was obtained: ¹ H NMR (300 MHz, DMSO-d 6): δ 11.35 (br, OH), 9.08 (br, NH), 8.15 (m , 1H), 7.96 ~ 7.94 (m, 1H), 7.58 ~ 7.56 (m, 1H), 7.23 ~ 7.14 (m, 5H), 3.59 (s, 3H), 2.82 (s, 2H), 2.68 (s, 2H ), 2.01 (s, 2H).

<6-5> methyl  2- Chloro -3- (3- Phenylpropyl ) Quinoxaline -6- Carboxylate  synthesis

Scheme 37

Methyl-2-oxo-3- (3-phenylpropyl) -1,2-dihydroquinoxaline-6-carboxylate (400 mg, 1.24 mmol) was dissolved in POCl ₃ It was dissolved in (5 mL) (aldrich) and stirred under reflux for 3 hours. After the reaction was completed, the reaction solution was added to ice-water to which NaHCO ₃ (30 mL) was added to produce a solid material. The solid material was purified by column chromatography (ethylacetate: hexane = 1: 3) using methyl 2-chloro-3- (3-phenylpropyl) quinoxaline-6-carboxylate (340 mg, 0.99 mmol, Yield 80%) was obtained: ¹ H NMR (300 MHz, CDCl ₃ ): δ 8.76 (d, J = 1.7 Hz, 1H), 8.32 (dd, J = 8.7 Hz, 1.7 Hz, 1H), 8.01 (d , J = 8.7Hz, 1H), 7.32 ~ 7.19 (m, 5H), 4.01 (s, 3H), 3.21 ~ 2.16 (m, 2H), 2.84 ~ 2.79 (m, 2H), 2.30 ~ 2.22 (m, 2H ).

<6-6> 2- Methoxy -3- (3- Phenylpropyl ) Quinoxaline -6- Carboxylic acid  synthesis

Scheme 38

Methyl 2-chloro-3- (3-phenylpropyl) quinoxaline-6-carboxylate (330 mg, 0.97 mmol) was dissolved in tetrahydrofuran (10 mL) and methanol (10 mL), followed by water (10 mL). ) Was added dropwise a solution of LiOH (204 mg, 4.85 mmol) and stirred at 50 ° C. for 4 hours.

After the reaction was completed, pH was adjusted by adding 2 M HCl aqueous solution. The organic layer was extracted from the reaction with ethyl acetate. The organic layer was dried over MgSO 4 and concentrated to afford 2-methoxy-3- (3-phenylpropyl) quinoxalin-6-carboxylic acid (300 mg, 0.92 mmol, 94% yield, white solid): ¹ H NMR (300 MHz, CDCl ₃ ): δ 13.19 (br, OH), 8.42 (d, J = 1.8 Hz, 1H), 8.12 (dd, J = 8.6 Hz, 1.8 Hz, 1H), 7.83 (d, J = 8.6Hz, 1H), 7.29 ~ 7.12 (m, 5H), 4.04 (s, 3H), 2.93 ~ 2.88 (m, 2H), 2.72 ~ 2.67 (m, 2H), 2.12 ~ 2.02 (m, 2H) .

<6-7> 2- Methoxy -3- (3- Phenylpropyl ) -N- ( Tetrahydro -2H-pyran-2- Sake ) Quinoxaline -6- Carboxamide  synthesis

Scheme 39

2-methoxy-3- (3-phenylpropyl) quinoxaline-6-carboxylic acid (300 mg, 0.92 mmol) was dissolved in dimethylformamide (1 mL), then EDCI (212 mg, 1.1 mmol), 1 -HOBT (125 mg, 0.92 mmol), NH ₂ OTHP (216 mg, 1.84 mmol) and diisopropylethylamine (476 mg, 3.68 mmol) were added and reacted by stirring at room temperature for 4 hours. After the reaction was completed, the organic layer was extracted from the reaction with ethyl acetate / NH ₄ Cl. The organic layer was dried over MgSO 4, concentrated, and then 2-methoxy-3- (3-phenylpropyl) -N- (tetrahydro-2H-) using column chromatography (ethyl acetate: hexane = 1: 1). Pyran-2-yloxy) quinoxaline-6-carboxamide (385 mg, 0.9 mmol, yield 98%) was obtained: ¹ H NMR (300 MHz, DMSO-d 6): δ 11.86 (br, NH), 8.36 (m, 1H), 8.03-8.00 (m, 1H), 7.86-7.83 (m, 1H), 7.29-7.13 (m, 5H), 5.03 (s, 1H), 4.04 (s, 3H + 1H), 3.54-3.51 (m, 1H), 2.94-2.89 (m, 2H), 2.72-2.67 (m, 2H), 2.14-2.04 (m, 2H), 1.72-1.54 (m, 6H).

<6-8> N- Hydro -2- Methoxy -3- (3- Phenylpropyl ) Quinoxaline -6- Carboxamide  Synthesis (Compound 18)

Scheme 40

2-Methoxy-3- (3-phenylpropyl) -N- (tetrahydro-2H-pyran-2-yloxy) quinoxaline-6-carboxamide (100 mg, 0.235 mmol) was added methylene chloride (2 mL). ) And methanol (2 mL), trifluoroacetic acid (536 mg, 4.7 mmol) was added thereto, and reacted at 50 ° C. for 12 hours. The reaction was concentrated, to give a solid compound under ether and hexane, and then filtered to give N-hydroxy-2-methoxy-3- (3-phenylpropyl) quinoxaline-6-carboxamide (25 mg ) ¹ H NMR (300 MHz, DMSO-d 6): δ 11.42 (br, OH), 9.14 (br, NH), 8.31 (d, J = 1.4 Hz, 1H), 8.00 (dd, J = 8.6Hz, 1.4Hz, 1H), 7.82 (d, J = 8.6Hz, 1H), 7.29 ~ 7.13 (m, 5H), 4.03 (s, 3H), 2.93 ~ 2.88 (m, 2H), 2.72 ~ 2.67 ( m, 2H), 2.12-2.02 (m, 2H).

Experimental Example

Experimental Example  1: isolated from mouse femur Mesenchymal  Induction of differentiation into neurons

In vitro experiments were performed to confirm the differentiation capacity of stem cells into neurons by the compound according to the present invention. First, bone marrow cells were isolated from femurs of Fisher mice (Central experimental animals) of 8 weeks old, and then cultured at 37 ° C. in DMEM medium. The cultured cells were placed in a T75 culture vessel (Nunc) at a concentration of 1 x 10 ⁵ cells and cultured under conditions of changing medium once every 3 days at 37 ° C for 10 days. Then, the cells were removed, placed in a culture vessel containing the same culture medium at a concentration of 1 x 10 ⁵ cells, and further cultured for 10 days under conditions of changing the medium once every 3 days at 37 ° C. This process was repeated five times to obtain mesenchymal stem cells. The obtained mesenchymal stem cells were placed in a culture dish having a diameter of 100 mm containing DMEM medium at a concentration of 1 × 10 ⁵ cells, and 10 μM of the compound of the above synthesis example was incubated for 2 days, and then the differentiation form of neurons was obtained. Observation was performed by Nissle staining (FIG. 1), and the differentiated cells were quantified (FIG. 2). Briefly, the method of dyeing dyes is as follows: i) Treated differentiated cells with 4% paraformaldehyde for 20 minutes at room temperature to fix the cells, and ii) wash them three times with PBS (phosphate buffer salin) solution. After treatment with nistle solution (0.5% cresyl solution) and reacted at room temperature for 5 minutes to remove the remaining solution, i) wash with PBS solution three times and observe the cells differentiated by optical microscope. At this time, cells treated with DMSO were used as a control.

As a result of observing the differentiation form of neurons by Nissle staining method, the ratio of differentiated cells to total stem cells in the mesenchymal stem cells treated with Compound 13, Compound 17, Compound 18 or Compound 15 was 66.6%, 5%, 10% and 33.1% (Figure 1). In particular, the best neuronal differentiation was observed in mesenchymal stem cells treated with Compound 13, and Compound 15 also showed excellent differentiation ability.

Experimental Example 2 Eruption induced In mesenchymal stem cells Neuronal Differentiation Markers  Observe protein changes

For the compound of Experimental Example 1, expression of neuronal differentiation marker, neuron-specific enolase (NSE) and beta III tubulin, was observed at the protein level through western blotting.

First, cells were treated with the compound at a concentration of 10 μM, and then whole cell proteins were extracted. Proteins were denatured by applying heat in a solution containing sodium dodesyl sulfate (SDS), and the proteins were electrically listed by size on SDS-PAGE. The proteins listed by size were transferred to a nylon filter (Bio-red) and then treated with neuronal differentiation marker antibodies, anti-beta III tubulin and anti-NSE (abcam), and reacted at 4 ° C. for 24 hours. At this time, a secondary antibody (Santacruz) having a fluorescence generating enzyme that recognizes a neuronal differentiation marker antibody was attached. The expression of neuronal differentiation marker protein was confirmed using a fluorescence measurement device (image analyzer 3000, Fuji), and the results are shown in FIG. 3. As shown in FIG. 3, the expression of neuronal differentiation markers NSE and beta III tubulin was shown, indicating that the compounds of the present invention can induce neuronal differentiation. That is, beta III tubulin and NSE, which are neuronal differentiation markers, were overexpressed in mesenchymal stem cells treated with Compound 13, Compound 17, Compound 18, or Compound 15, and thus, only cells treated with the compound of the present invention were derived from bone marrow-derived mesenchymal stem. It was confirmed that the cells differentiated into neurons, and in particular, compound 13 was found to have a very good neuronal differentiation ability.

Experimental Example  3: electro biochemical experiment

To conduct electrochemical experiments on whole cells, cells were cultured in special culture solutions (NaCl (150 mM), KCl (5 mM), MgCl ₂ ). (1 mM), CaCl ₂ (2.2 mM), Hepes (10 mM), pH 7.3) and glucose to adjust osmotic pressure 310 mOsM. The solution in the pipette for the electrical measurement was aspartic acid potassium salt (120 mM), MgCl ₂ (5 mM), ethylene glycol tetraacetic acid (EGTA, 0.5 mM), Hepes (10 mM), ATP (2 mM), GTP (0.3 mM). Whole cell electrical analysis was performed with the instrument of Axopatch 200B (Molecular Devices Corportion) (FIG. 4). As a result, mesenchymal stem cells treated with Compound 13 showed excellent biochemical properties.

While the present invention has been particularly shown and described with reference to exemplary embodiments thereof, it is to be understood that the same is by way of illustration and example only and is not to be construed as limiting the scope of the present invention. Accordingly, the actual scope of the present invention will be defined by the appended claims and their equivalents.

Claims (9)

delete delete delete delete A quinoxaline derivative represented by Formula 13, Formula 15, Formula 16, or Formula 18, or a salt thereof:
Formula 13

Formula 15

Formula 16

Formula 18

The compound according to claim 5, wherein the quinoxaline derivative or salt thereof is selected from the group consisting of the following Chemical Formulas 13, 15 and 18:
Formula 13

Formula 15

Formula 18

A composition for inducing differentiation of neuroprogenitor cells or stem cells into neurons, comprising the quinoxaline derivative of claim 5 or 6 or a salt thereof as an active ingredient.
8. The composition of claim 7, wherein the stem cells are embryonic stem cells, embryonic germ cells, embryonic tumor cells, adult stem cells and induced pluripotent stem cells.
8. The method of differentiating neural progenitor cells or stem cells into neurons, comprising culturing the composition of claim 7 together with neural progenitor cells or stem cells.

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