Method of synthesizing bilirubin

Abstract

The present invention relates to a method for synthesizing bilirubin, and more specifically, to a method for the first time chemically synthesizing bilirubin and pegylated bilirubin, which are useful as pharmaceuticals and the like, by comprising a step of coupling a compound represented by chemical formula 1 with a compound represented by chemical formula 2 to produce a compound represented by chemical formula 3.

Description

[Technical Field]

[0001] This invention relates to a novel method for synthesizing bilirubin. [Background technology]

[0002] Bilirubin is one of the components of bile and is mainly produced in the body from hemoglobin. Bilirubin is a yellow final metabolite formed from heme, and despite having many hydrophilic groups, it is extremely hydrophobic due to intramolecular hydrogen bonding.

[0003] Bilirubin was previously considered an unnecessary substance because high blood levels can cause jaundice. However, recent research has revealed that slightly elevated blood levels of bilirubin significantly reduce the risk of developing cardiovascular disease and cancer. Furthermore, animal studies have confirmed that bilirubin protects cells and tissues by performing functions such as removing various reactive oxygen species and regulating immune cells related to inflammation.

[0004] Despite its industrially useful properties, bilirubin has historically been obtained through animal extraction, and synthesis has never been successful. Extracting bilirubin from animals is difficult in large quantities and incurs high production costs. Furthermore, bilirubin extracted from animals is a mixture of three positional isomers, requiring additional separation and purification processes for use as a pharmaceutical. Therefore, there is an urgent need to develop a method for chemically producing bilirubin. [Overview of the project] [Problems that the invention aims to solve]

[0005] The present invention aims to provide a method for synthesizing bilirubin. [Means for solving the problem]

[0006] 1. A method for synthesizing bilirubin, comprising the step of coupling a compound represented by chemical formula 1 with a compound represented by chemical formula 2 to produce a compound represented by chemical formula 3. [ka] [ka] [ka] (In the above chemical formulas 1, 2, and 3, R1 and R2 are independently hydrogen, a C1-C12 alkyl group, a C6-C20 aryl group, a C2-C20 heteroaryl group, a C7-C20 arylalkyl group, or a C3-C20 heteroarylalkyl group; R3 is a vinyl group or an acetyl group; or an ethyl group substituted with a hydroxyl group, a carbamate, a selenide, or a sulfide; R4 is a hydrogen or nitrogen protecting group; and R5 is hydrogen, a tosyl group, or a mesyl group.)

[0007] 2. A method for synthesizing bilirubin, further comprising the step of reacting a compound represented by chemical formula 3 with polyethylene glycol (PEG) in item 1 above.

[0008] 3. A method for synthesizing bilirubin, wherein, in item 1 above, the compound represented by chemical formula 1 is reacted with polyethylene glycol (PEG), and then coupled with the compound represented by chemical formula 2.

[0009] 4. A method for synthesizing bilirubin, further comprising the step of dimerizing the compound represented by chemical formula 7 to produce the compound represented by chemical formula 1, as described in item 1 above. [ka] (In the formula, R1 is the same as R1 in chemical formula 1, and X is an arylalkyl ester group having 8 to 20 carbon atoms, -CH2OH, -COOH, a halogen atom, or hydrogen.)

[0010] 5. A method for synthesizing bilirubin, further comprising the step of oxidizing a compound represented by chemical formula 9 to produce a compound represented by chemical formula 2, as described in item 1 above. [ka] (In the formula, R4 is the same as R4 in chemical formula 2.)

[0011] 6. A method for synthesizing bilirubin, further comprising the step of reducing the acetyl group of the compound represented by chemical formula 10 to produce the compound represented by chemical formula 2, as described in item 1 above. [ka] (In the formula, R4 is the same as R4 in chemical formula 2.)

[0012] 7. A method for synthesizing bilirubin, further comprising the step of dehydrating the hydroxyl group of the compound represented by chemical formula 11 to produce the compound represented by chemical formula 2, as described in item 1 above. [ka] (In the formula, R4 is the same as R4 in chemical formula 2.)

[0013] 8. A method for synthesizing bilirubin, further comprising the step of producing a compound represented by chemical formula 2 by cyclization and halogen removal reactions of a compound represented by chemical formula 12 in item 1 above. [ka] (In the formula, R4 is the same as R4 in chemical formula 2.)

[0014] 9. A method for synthesizing bilirubin, further comprising the step of oxidizing and carbamate the compound represented by chemical formula 13 to produce the compound represented by chemical formula 2, as described in item 1 above. [ka] (In the formula, R is an alkyl group having 1 to 12 carbon atoms, and R4 is the same as R4 in chemical formula 2.)

[0015] 10. A method for synthesizing bilirubin, further comprising the step of cyclizing the compound represented by chemical formula 14 to produce the compound represented by chemical formula 2, as described in item 1 above. [ka] (In the formula, Y is selenide, and R4 is the same as R4 in chemical formula 2.)

[0016] 11. A method for synthesizing bilirubin, further comprising the step of oxidizing a compound represented by chemical formula 15 to produce a compound represented by chemical formula 2, as described in item 1 above. [ka] (In the formula, Z is a sulfide, and R4 and R5 are the same as R4 and R5 in chemical formula 2.)

[0017] 12. A method for synthesizing bilirubin, further comprising the step of producing bilirubin from a compound represented by chemical formula 3, as described in item 1 above.

[0018] 13. In item 1 above, the step is piperidine, N-methylpiperidine, N-ethylpiperidine, 2,6-dimethylpiperidine, 2,2,6,6-tetramethylpiperidine, 3-methylpiperidine, 3-ethylpiperidine, 1-methyl-4-(methylamino)piperidine, 4-aminopiperidine, pyrrolidine, 2-pyrrolidinecarboxamide, pyrrolidine-3-ol, piperazine, 2,6-dimethylpiperazine, 1-benzylpiperazine, 1-isopropylpiperazine, 2-ethylpiperazine, morpholine, 4-methylmorpholine, 2,6-di A method for synthesizing bilirubin, carried out in the presence of a base selected from the group consisting of methylmorpholine, ethylmorpholine, azepane, 2-methylazepane, 4-methylazepane, 2,2,7,7-tetramethylazepane, 1,2,2-trimethylazepane, 1,2-dimethylazepane, 2,7-dimethylazepane, methylazepane-4-carboxylate, azocanne, 2-methylazocanne, 1,2-dimethylazocanne, 1,2,2-trimethylazocanne, methylazocanne-2-carboxylate, 1-methylazocanne, and 2-(2-methylphenyl)azocanne.

[0019] 14. A method for synthesizing bilirubin, wherein the step in item 1 is carried out in the presence of a solvent selected from the group consisting of water, alcohols, ethers, ketones, aliphatic hydrocarbons, aromatic hydrocarbons, halogenated hydrocarbons, alkoxys, nitriles, and amides.

[0020] 15. A method for synthesizing bilirubin, wherein the step in item 1 is performed at a temperature of -20°C to 200°C.

[0021] 16. A method for synthesizing bilirubin, wherein the step in item 1 is carried out for 0.5 to 120 hours.

[0022] 17. A method for synthesizing bilirubin, wherein, in item 13 above, the base is added in amounts of 2 to 20 moles based on 1 mole of the compound represented by chemical formula 1. [Effects of the Invention]

[0023] The bilirubin synthesis method of the present invention can be carried out economically under mild conditions.

[0024] The bilirubin synthesis method of the present invention yields high yields and is suitable for mass production. [Brief explanation of the drawing]

[0025] [Figure 1] Figures 1-4 show the 2D NMR data of F-13a produced in Example 13. Figure 2 shows HSQC data, Figure 3 shows COSY data, and Figure 4 shows NOESY data. [Figure 2] Figures 1-4 show the 2D NMR data of F-13a produced in Example 13. Figure 2 shows HSQC data, Figure 3 shows COSY data, and Figure 4 shows NOESY data. [Figure 3] Figures 1-4 show the 2D NMR data of F-13a produced in Example 13. Figure 2 shows HSQC data, Figure 3 shows COSY data, and Figure 4 shows NOESY data. [Figure 4] Figures 1-4 show the 2D NMR data of F-13a produced in Example 13. Figure 2 shows HSQC data, Figure 3 shows COSY data, and Figure 4 shows NOESY data. [Modes for carrying out the invention]

[0026] This invention relates to a novel method for synthesizing bilirubin.

[0027] In this specification, the term "alkyl" refers to linear or branched, substituted or unsubstituted chain hydrocarbons. For example, methyl, ethyl, n-propyl, isopropyl 、n -butyl, sec-butyl, tert-butyl 、n -Pentyl, isopentyl, neopentyl, tert-pentyl 、n -Hexyl, Isohexyl In be.

[0028] The term "cycloalkyl" refers to monocyclic or bicyclic or multicyclic cyclic hydrocarbons, whether substituted or unsubstituted. Examples include cyclopropyl, cyclobutyl, cyclopentyl, cyclohexyl, cycloheptyl, and cyclooctyl.

[0029] The term "heterocycloalkyl" refers to a monocyclic or bicyclic or multicyclic, substituted or unsubstituted cyclic hydrocarbon containing one or more heteroatoms selected from B, N, O, S, P(=O), Si, and P. For example, these include tetrahydropyranyl group, azetidyl group, 1,4-dioxanyl group, piperazinyl group, piperidinyl group, pyrrolidinyl group, morpholinyl group, thiomorpholinyl group, dihydrofuranyl group, dihydroimidazolyl group, dihydroindolyl group, dihydroisoxazolyl group, dihydroisothiazolyl group, dihydrooxadiazolyl group, dihydroxazolyl group, dihydropyradinyl group, dihydropyrazolyl group, dihydropyridyl group, dihydropyrimidinyl group, dihydropyrrolyl group, dihydroquinolyl group, dihydrotetrazolyl group, dihydrothiadiazolyl group, dihydrothiazolyl group, dihydrothienyl group, dihydrotriazolyl group, dihydroazetidyl group, methylenedioxybenzoyl group, tetrahydrofuranyl group, or tetrahydrothienyl group.

[0030] The term "aryl" refers to monocyclic or bicyclic or multicyclic aromatic groups, whether substituted or unsubstituted. Examples include phenyl, biphenyl, terphenyl, naphthyl, binaphthyl, phenylnaphthyl, naphthylphenyl, phenylterphenyl, fluorenyl, phenylfluorenyl, diphenylfluorenyl, benzofluorenyl, dibenzofluorenyl, phenantrenyl, phenylphenantrenyl, anthracenyl, indenyl, triphenylenyl, pyrenyl, tetracerenyl, perilenyl, chrysenyl, naphthacenyl, fluoranthenyl, spirobifluorenyl, and azlenyl.

[0031] "Aryl" is, for example, phenyl, 1-naphthyl, 2-naphthyl, 1-anthryl, 2-anthryl, 9-anthryl, benzanthryl, 1-phenanthryl, 2-phenanthryl, 3-phenanthryl, 4-phenanthryl, 9-phenanthryl, naphthacenyl, pyrenyl, 1-crisenyl, 2-crisenyl, 3-crisenyl, 4-crisenyl, 5-crisenyl, 6-crisenyl, benzo[c]phenanthryl, benzo[g]crisenyl, 1-Triphenylenyl, 2-Triphenylenyl, 3-Triphenylenyl, 4-Triphenylenyl, 1-Fluorenyl, 2-Fluorenyl, 3-Fluorenyl, 4-Fluorenyl, 9-Fluorenyl, Benzofluorenyl, Dibenzofluorenyl, 2-Biphenylyl, 3-Biphenylyl, 4-Biphenylyl, o-Terphenyl, m-Terphenyl-4-yl, m-Terphenyl-3-yl, m-Terphenyl-2-yl, p-Terphenyl-4 -yl, p-terphenyl-3-yl, p-terphenyl-2-yl, m-quaterphenyl, 3-fluoranthenyl, 4-fluoranthenyl, 8-fluoranthenyl, 9-fluoranthenyl, benzofluoranthenyl, o-tolyl, m-tolyl, p-tolyl, 2,3-xylyl, 3,4-xylyl, 2,5-xylyl, mesityl, o-cumenyl, m-cumenyl, p-cumenyl, p-tert-butylphenyl, p-(2-phenylpropyl Examples include phenyl, 4'-methylbiphenylyl, 4''-tert-butyl-p-terphenyl-4-yl, 9,9-dimethyl-1-fluorenyl, 9,9-dimethyl-2-fluorenyl, 9,9-dimethyl-3-fluorenyl, 9,9-dimethyl-4-fluorenyl, 9,9-diphenyl-1-fluorenyl, 9,9-diphenyl-2-fluorenyl, 9,9-diphenyl-3-fluorenyl, and 9,9-diphenyl-4-fluorenyl.

[0032] The term "heteroaryl" refers to a monocyclic or bicyclic or multicyclic substituted or unsubstituted aromatic group containing one or more heteroatoms selected from B, N, O, S, P(=O), Si, and P. For example, these include benzothienyl, benzoxazolyl, benzofuranil, benzimidazolyl, benzthiazolyl, benzotriazolyl, cinnolinyl, furyl, imidazolyl, tetrazolyl, indazolyl, indolyl, isoxazolyl, isoquinolinyl, isothiazolyl, naphthilidinyl, oxadiazolyl, oxazolyl, isoxazolyl, flunyl, thiazolyl, isothiazolyl, thienopyridinyl, thienyl, thiadiazolyl, pyridinyl, pyridadinyl, pyrimidinyl, pyrazolyl, pyrrolyl, pyrido[2,3-d]pyrimidinyl, pyrrolo[2,3-b]pyrimidinyl, quinazolinyl, quinolinyl, thieno[2,3-c]pyrimidinyl, triazinyl, etc.

[0033] The term "arylalkyl" refers to an alkyl group in which at least one substituent is substituted with an aryl group, and "aryl" and "alkyl" are as described above. Examples include benzyl, phenylethyl, phenylpropyl, phenylbutyl, phenylhexyl, naphthylethyl, naphthylpropyl, naphthylbutyl, naphthylhexyl, anthracenylmethyl, anthracenylethyl, anthracenylpropyl, anthracenylbutyl, phenanthrylmethyl, phenanthrylethyl, phenanthrylpropyl, triphenylmethyl, triphenylethyl, triphenylpropyl, pyrenylmethyl, pyrenylethyl, pyrenylpropyl, phenylanthracenemethyl, phenylanthraceneethyl, phenylanthracenepropyl, perilenylmethyl, perilenylethyl, perilenylpropyl, crisenylmethyl, crisenylethyl, crisenylpropyl, fluorenylmethyl, fluorenylethyl, fluorenylpropyl, etc.

[0034] The term "heteroarylalkyl" refers to an alkyl group in which at least one substituent is replaced with a heteroaryl group, as described above. Examples include pyridinylmethyl, pyridinylethyl, pyridinylpropyl, pyridinylbutyl, pyrimidinylmethyl, pyrimidinylethyl, pyrimidinylpropyl, pyrazolylmethyl, pyrazolylethyl, pyrazolylmethyl, pyrazolylethyl, pyrazolylpropyl, quinolinylmethyl, quinolinylethyl, and quinolinylpropyl.

[0035] The term "substituted" means containing at least one substituted compound, such as one or more halogen atoms, nitro, hydroxy, cyano, amino, thiol, carboxyl, amide, nitrile, sulfide, disulfide, sulfenyl, formyl, formyloxy, formylamino, formylamino, aryl, or substituted aryls.

[0036] In the chemical formula of the present invention, if no substituent is listed at a site where a substituent is required, it means that a hydrogen substituent has been omitted.

[0037] The present invention relates to a method for synthesizing bilirubin, comprising the step of coupling a compound represented by chemical formula 1 with a compound represented by chemical formula 2 to produce a compound represented by chemical formula 3.

[0038] [ka]

[0039] [ka]

[0040] [ka]

[0041] In the above chemical formulas 1, 2, and 3, R1 and R2 are independently hydrogen, an alkyl group having 1 to 12 carbon atoms, an aryl group having 6 to 20 carbon atoms, a heteroaryl group having 2 to 20 carbon atoms, an arylalkyl group having 7 to 20 carbon atoms, or a heteroarylalkyl group having 3 to 20 carbon atoms.

[0042] The number of carbon atoms in R1 and R2 can be appropriately selected within a range that does not affect the coupling reaction between the compound represented by chemical formula 1 and the compound represented by chemical formula 2.

[0043] For example, R1 and R2 may independently be an alkyl group having 1 to 10 carbon atoms, an aryl group having 6 to 10 carbon atoms, a heteroaryl group having 2 to 10 carbon atoms, an arylalkyl group having 7 to 10 carbon atoms, or a heteroarylalkyl group having 3 to 10 carbon atoms.

[0044] Furthermore, R1 and R2 may independently be an alkyl group having 1 to 5 carbon atoms, an aryl group having 6 to 10 carbon atoms, a heteroaryl group having 4 to 10 carbon atoms, an arylalkyl group having 7 to 10 carbon atoms, or a heteroarylalkyl group having 5 to 10 carbon atoms.

[0045] R3 is a vinyl group or an acetyl group; or an ethyl group substituted with a hydroxyl group, carbamate, selenide, or sulfide.

[0046] Here, carbamate is a functional group having the structure shown in chemical formula 4 below.

[0047] [ka]

[0048] In chemical formula 4, R is an alkyl group having 1 to 12 carbon atoms or an alkyl group having 1 to 5 carbon atoms.

[0049] Selenides are functional groups having the structure of chemical formula 5 below, and sulfides are functional groups having the structure of chemical formula 6 below.

[0050] [ka]

[0051] [ka]

[0052] In chemical formulas 5 and 6, R X This may be hydrogen, or a substituted or unsubstituted linear or branched alkyl group, cycloalkyl group, heterocycloalkyl group, aryl group, heteroaryl group, arylalkyl group, or heteroarylalkyl group.

[0053] For example, R X These are alkyl groups having 1 to 12 carbon atoms, cycloalkyl groups having 5 to 20 carbon atoms, heterocycloalkyl groups having 2 to 20 carbon atoms, aryl groups having 5 to 20 carbon atoms, heteroaryl groups having 2 to 20 carbon atoms, arylalkyl groups having 6 to 20 carbon atoms, or heteroarylalkyl groups having 3 to 20 carbon atoms.

[0054] For example, R X This is a phenyl group or a p-tolyl group.

[0055] R3 may be an ethyl group substituted with a hydroxyl group. For example, it may be a functional group in which a hydroxyl group is substituted at the position of the first carbon atom of an ethyl group.

[0056] R3 may be a carbamate-substituted ethyl group. For example, it may be a functional group in which a carbamate is substituted at the position of the second carbon of an ethyl group.

[0057] R3 may be an ethyl group substituted with a selenide. For example, it may be a functional group in which a selenide is substituted at the position of the second carbon of an ethyl group.

[0058] R3 may be a sulfide-substituted ethyl group. For example, it may be a functional group in which a sulfide is substituted at the position of the second carbon of an ethyl group.

[0059] R4 is a hydrogen or nitrogen protecting group.

[0060] Here, the nitrogen protecting group is not limited to any particular substituent, as long as it serves to protect the nitrogen atom to which R4 is bonded. For example, -COOR x (R x The group may be selected from the group consisting of the following: ), tert-butyloxycarbonyl (Boc), trityl (-CPh3), tosyl group (SOOPhCH3), 9-fluorenylmethyloxycarbonyl (Fmoc), carboxybenzyl group (Cbz), p-methoxybenzylcarbonyl (Moz), acetyl (Ac), benzoyl (Bz), p-methoxybenzyl (PMB), 3,4-dimethoxybenzyl (DMPM), p-methoxyphenyl (PMP), 2-naphthylmethyl ether (Nap), and trichloroethyl chloroformate (Troc).

[0061] R5 is hydrogen, tosyl group ( [ka] , Ts or Tos) or mesyl group ( [ka] She is Ms.

[0062] After coupling the compound represented by chemical formula 1 with the compound represented by chemical formula 2, if the nitrogen protecting group R4 of chemical formula 2 remains, a further step may be required to remove this nitrogen protecting group.

[0063] The compound represented by chemical formula 1 and the compound represented by chemical formula 2 bond in a molar ratio of 1:2.

[0064] The compound represented by chemical formula 1 and the compound represented by chemical formula 2 can be added in molar ratios of 1:2-10, 1:2-5, 1:2-4, or 1:2-3 through the reaction.

[0065] Coupling reactions are carried out under solvent and base conditions.

[0066] The solvent is either an inorganic solvent or an organic solvent. Organic solvents include, for example, alcohols, ethers, ketones, aliphatic hydrocarbons, aromatic hydrocarbons, halogenated hydrocarbons, alkoxys, nitriles, or amides. Solvents belonging to these categories are shown, for example, in Table 1. An inorganic solvent is, for example, water.

[0067] [Table 1]

[0068] The base can be an organic or inorganic base. It is preferable to use a base that is stronger than the compound represented by chemical formula 2.

[0069] It is preferable to use an amine-based organic base. For example, it may be a chain-like amine-based organic base such as methylamine, ethylamine, dimethylamine, diethylamine, ethylmethylamine, propylamine, dipropylamine, methylpropylamine, ethylpropylamine, diisopropylamine, N-methylcyclohexylamine, or trimethylamine. Also, aziridine, azetidine, oxaziridine, azetidine, diazetidine, imidazolidine, pyrazolidine, oxazolidine, isoxazolidine, thiazolidine, isothiazolidine, piperidine, 2-methylpiperidine, 2-ethylpiperidine, 2,6-dimethylpiperidine, N-methylpiperazine, N-ethylpiperidine, 2,6-dimethylpiperidine, 2,2,6,6-tetramethylpiperidine, 3-methylpiperidine, 3-ethylpiperidine, 1-methyl-4-(methylamino) It may also be peridine, 4-aminopiperidine, pyrrolidine, 2-pyrrolidinecarboxamide, pyrrolidine-3-ol, piperazine, 2,6-dimethylpiperazine, 1-benzylpiperazine, 1-isopropylpiperazine, 2-ethylpiperazine, N-propylpiperazine, morpholine, thiomorpholine, 4-methylmorpholine, 2,6-dimethylmorpholine, ethylmorpholine, azepane, 2-methylazepane, 4-methylazepane, 2,2,7,7-tetramethylazepane, 1,2,2-trimethylazepane, 1,2-dimethylazepane, 2,7-dimethylazepane, azocan, 1,2-dimethylazocan, 1,2,2-trimethylazocan, methylazocan-2-carboxylate, 1-methylazocan, 2-(2-methylphenyl)azocan, or a cyclic amine organic base such as proline.

[0070] The organic base is preferably piperidine, pyrrolidine, morpholine, piperazine, azepane, azocane, N-methylpiperidine, N-ethylpiperidine, or proline.

[0071] The inorganic base may be, for example, LiOH, KOH, or NaOH.

[0072] The base is used in amounts of 2-20 moles, 2-15 moles, 2-10 moles, 4-20 moles, 4-15 moles, 4-10 moles, 5-20 moles, 5-15 moles, 5-10 moles, 6-20 moles, 6-15 moles, or 6-10 moles per mole of the compound represented by chemical formula 1.

[0073] The reaction temperature for the coupling reaction of the present invention is -20°C to 200°C. For example, 30°C to 180°C, 30°C to 150°C, 30°C to 120°C, 30°C to 100°C, 40°C to 150°C, 40°C to 140°C, 40°C to 120°C, 40°C to 100°C, 50°C to 150°C, 50°C to 120°C, or 50°C to 100°C. The optimal reaction temperature may vary depending on the solvent and base used.

[0074] The reaction time for the coupling reaction of this invention ranges from 10 minutes to 120 hours. For example, 1 hour to 72 hours, 1 hour to 48 hours, 1 hour to 24 hours, 3 hours to 72 hours, 3 hours to 48 hours, 3 hours to 24 hours, 6 hours to 72 hours, 6 hours to 48 hours, or 6 hours to 24 hours. The optimal reaction time may vary depending on the solvent and base used.

[0075] The bilirubin synthesis method of the present invention may further include the step of converting R1 and / or R2 of the compound represented by chemical formula 3 to hydrogen by a saponification reaction. For example, if R1 and R2 of the compound represented by chemical formula 3 are methyl groups, a base such as LiOH, KOH, or NaOH is added to the compound represented by chemical formula 3 to replace the methyl groups with hydrogen.

[0076] The solvent used in the saponification reaction is not particularly limited. The same solvent used in the coupling reaction can be used as the solvent for the saponification reaction. For example, methanol, ethanol, 2-propanol, tetrahydrofuran (THF), 2-methyltetrahydrofuran (ME-THF), dioxane, acetonitrile, N,N-dimethylformamide (DMF), t-butanol, dimethoxyethane (DME), dichloromethane (DCM), or isopropyl alcohol may be used.

[0077] Saponification reactions can be carried out under conditions known in this art. For example, they can be carried out at 10-150°C for 1-72 hours, or at 10-60°C for 1-48 hours.

[0078] The bilirubin synthesis method of the present invention may further include a pegylation step in which a compound represented by chemical formula 3 is reacted with polyethylene glycol (PEG).

[0079] The bilirubin synthesis method of the present invention may also include the step of reacting a compound represented by chemical formula 1 with polyethylene glycol (PEG) in a pegylation reaction, and then coupling the resulting product with a compound represented by chemical formula 2.

[0080] PEGylated bilirubin exhibits improved water solubility.

[0081] Polyethylene glycol is, for example, mPEG n This is -NH2 (methoxypolyethylene glycol-amine, n=5~60). n is the number of -CH2-CH2-O- repeating units of methoxypolyethylene glycol-amine, which may be 5~60, 10~50, 10~40, 20~40, 10~30, or 20~30.

[0082] Pegging includes both mono-PEGylation, where either O-R1 or O-R2 is pegged, and bi-PEGylation, where both are pegged.

[0083] In the PEGylation reaction, polyethylene glycol can be added in an appropriate amount, taking into account the number of moles of the compound represented by chemical formula 1 or chemical formula 3. For example, polyethylene glycol can be added in amounts of 0.1 to 10 moles, 0.1 to 8 moles, 0.1 to 5 moles, 0.3 to 8 moles, 0.3 to 5 moles, 0.3 to 4 moles, or 0.3 to 3 moles per mole of the compound represented by chemical formula 1 or chemical formula 3.

[0084] The reagents used in the pegylation reaction include CDI (1,1-carbonyldiimidazole), CMPI (2-chloro-1-methylpyridinium iodide), BEP (2-bromo-1-ethyl-pyridinium tetrafluoroborate), EDCI (1-ethyl-3-(3-dimethylaminopropyl)carbodiimide), and HATU (1-[bis(dimethylamino)methylene]-1H-1,2,3-triazolo[4,5-b]pyridinium 3-oxidehexafluorophosphate). 3-oxide hexafluorophosphate (Hexafluorophosphate Azabenzotriazole Tetramethyl Uronium), DCC (N,N'-Dicyclohexylcarbodiimide), or HOBt (Hydroxybenzotriazole) may be used, but are not limited to these.

[0085] The reagent for the pegylation reaction can be added in amounts of 0.3 to 5 moles, 0.3 to 3 moles, 0.5 to 5 moles, 0.5 to 3 moles, 0.5 to 2.5 moles, or 0.5 to 2 moles per mole of the compound represented by chemical formula 1 or chemical formula 3, but is not limited to these amounts.

[0086] The solvent for the pegylation reaction is not particularly limited. The same solvent used for the coupling reaction can be used for the pegylation reaction. For example, DMSO (dimethyl sulfoxide), DMF (dimethylformamide), DMA (dimethylacetamide), or pyridine may be used.

[0087] The pegylation reaction can be carried out under a base. The base is selected from the range exemplified as bases in the coupling reaction described above, and is preferably DIPEA (N,N-diisopropylethylamine) or pyridine.

[0088] The pegylation reaction can be carried out at temperatures between 10°C and 100°C, for example, between 10°C and 80°C, 20°C and 60°C, 20°C and 50°C, or 20°C and 30°C.

[0089] The pegylation reaction can be carried out for 1 to 24 hours, 1 to 18 hours, or 1 to 12 hours, but is not limited to these durations.

[0090] In one embodiment, the pegylation reaction can be carried out at 20°C to 40°C for 0.5 to 24 hours by adding 0.3 to 5 moles of polyethylene glycol and 0.5 to 5 moles of a coupling reagent (such as CDI, EDCI, or CMPI) to 1 mole of a compound represented by chemical formula 1 or chemical formula 3.

[0091] The compounds represented by chemical formula 1 and chemical formula 2, which are reactants in the bilirubin synthesis method of the present invention, can be produced as follows.

[0092] The compound represented by chemical formula 1 can be produced by dimerizing the compound represented by chemical formula 7 and substituting X in the product with -C(=O)H.

[0093] [ka]

[0094] In chemical formula 7, R1 is the same as in chemical formula 1, and X is an arylalkyl ester group having 8 to 20 carbon atoms, -CH2OH, -COOH, a halogen atom, or hydrogen.

[0095] The arylalkyl ester group is R in chemical formula 8. Y The functional group is an arylalkyl group.

[0096] [ka]

[0097] The arylalkyl group of chemical formula 8 is the same as the arylalkyl group of chemical formula 1.

[0098] The number of carbon atoms in the arylalkyl ester group can be appropriately selected within a range that does not affect the dimerization reaction of the compound represented by chemical formula 7. For example, it may have 8 to 20 carbon atoms, 8 to 18 carbon atoms, 8 to 15 carbon atoms, or 8 to 12 carbon atoms.

[0099] Methods for substituting X in the product with an aldehyde group include, for example, the following (1) to (5).

[0100] (1) If X is an arylalkyl ester group, the arylalkyl ester group can be reduced to -COOH by a hydrogenation reaction, and after the -COOH is reduced or removed, -C(=O)H can be added to substitute X with an aldehyde group.

[0101] In one embodiment, the hydrogenation reaction can be carried out under a Pd / C catalyst. For example, the arylalkyl ester group is -C(=O)OBn (Bn=benzyl), and -C(=O)OBn is reduced to -COOH by a hydrogenation reaction under a Pd / C catalyst, and then -COOH is removed to replace the arylalkyl ester group with -C(=O)H.

[0102] (2) If X is -CH2OH, oxidize X by a known method and replace it with -C(=O)H.

[0103] (3) If X is -COOH, reduce or remove the -COOH group, then add -C(=O)H to replace X with an aldehyde group.

[0104] (4) If X is a halogen atom, the halogen atom is replaced with an aldehyde group by a carbonylation reaction. A known method can be used for the carbonylation reaction. For example, the carbonylation reaction can be carried out using carbon monoxide and palladium.

[0105] (5) If X is hydrogen, the hydrogen is replaced with an aldehyde group by an aldehyde addition reaction. Known methods can be used for the aldehyde addition reaction. For example, the aldehyde addition reaction can be carried out using BuLi and DMF.

[0106] The dimerization reaction of the compound represented by chemical formula 6 can be carried out, for example, under bromine (Br2) conditions. The solvent for the dimerization reaction is not particularly limited. For example, the solvents listed in Table 1, which exemplify solvents for coupling reactions, can be used as organic solvents.

[0107] The dimerization reaction can be carried out at temperatures between 10°C and 100°C, for example, between 10°C and 80°C, 20°C and 60°C, 20°C and 50°C, or 20°C and 30°C.

[0108] The dimerization reaction can be carried out for 1 to 24 hours, 1 to 18 hours, or 1 to 12 hours, but is not limited to these periods.

[0109] The compound represented by chemical formula 2 (R3 = acetyl) can be produced by oxidizing the compound represented by chemical formula 9.

[0110] [ka]

[0111] In the formula, R4 is the same as R4 in chemical formula 2.

[0112] The oxidation reaction of the compound represented by chemical formula 9 can be carried out under H2O2 and pyridine. The oxidation reaction of the compound represented by chemical formula 9 can be carried out within the range of solvent, temperature, time, etc., used in coupling reactions.

[0113] The compound represented by chemical formula 2 (R3 = ethyl group substituted with a hydroxyl group) can be produced by reducing the compound represented by chemical formula 10.

[0114] [ka]

[0115] In the formula, R4 is the same as R4 in chemical formula 2 above.

[0116] The reaction to reduce acetyl groups can be carried out by known methods, and the conditions can be within the range of solvent, temperature, and time used in coupling reactions. For example, acetyl groups can be reduced using MeOH as the solvent and NaBH4 and CeCl37H2O as the reagents for the reduction reaction, or acetyl groups can be reduced using THF as the solvent and DIBAL as the reagent for the reduction reaction.

[0117] The compound represented by chemical formula 2 (R3 = vinyl group) can be produced by dehydrating the hydroxyl group of the compound represented by chemical formula 11.

[0118] [ka]

[0119] In the formula, R4 is the same as R4 in chemical formula 2.

[0120] The dehydration reaction of hydroxyl groups can be carried out by known methods, and the conditions can be within the range of solvent, temperature, and time used in the aforementioned coupling reaction. For example, hydroxyl groups can be dehydrated using DCM (dichloromethane) as the solvent and POCl3 and TEA as the reaction reagents.

[0121] The compound represented by chemical formula 2 (R3 = vinyl group) can also be produced by cyclization and halogen removal reactions of the compound represented by chemical formula 12.

[0122] [ka]

[0123] In the formula, R4 is the same as R4 in chemical formula 2.

[0124] The cyclization reaction of the compound represented by chemical formula 12 can be carried out under CuCl and ACN (acetonitrile), and the halogen removal reaction can be carried out under DMF. The cyclization and halogen removal reactions of the compound represented by chemical formula 12 produce the compound represented by chemical formula 2 below.

[0125] The compound represented by chemical formula 2 (R3 = ethyl group substituted with a carbamate) can be produced by oxidizing and carbamate the compound represented by chemical formula 13.

[0126] [ka]

[0127] In the formula, R4 is the same as R4 in chemical formula 2, and R is the same as R in chemical formula 4.

[0128] The oxidation and carbamate of the compound represented by chemical formula 13 can be carried out by the following procedure.

[0129] [ka] The oxidation reaction of the compound represented by chemical formula 13 can be carried out, for example, under H2O2 and pyridine. Subsequently, carbamate can be carried out, for example, by treating with NH2NH2 followed by treatment with NaNO2 / HCl and an alcohol. Each reaction can be carried out within the range of solvent, temperature, time, etc., of the coupling reaction, but is not limited to these.

[0130] The compound represented by chemical formula 2 (R3 = ethyl group substituted with selenide) can be produced by cyclizing the compound represented by chemical formula 14.

[0131] [ka]

[0132] In the formula, Y is selenide, and R4 is the same as R4 in chemical formula 2.

[0133] The cyclization reaction of the compound represented by chemical formula 14 can be carried out under t-BuOK conditions and within the range of solvent, temperature, and time used in coupling reactions. Cyclization of the compound represented by chemical formula 14 yields the compound represented by chemical formula 2 below.

[0134] [ka]

[0135] The compound represented by chemical formula 2 (R3 = ethyl group substituted with sulfide) can be produced by oxidizing the compound represented by chemical formula 15.

[0136] [ka]

[0137] In the formula, Z is a sulfide, and R4 and R5 are the same as R4 and R5 in chemical formula 2.

[0138] The compound represented by chemical formula 2 (R3 = ethyl group substituted with sulfide) obtained by oxidizing the compound represented by chemical formula 15 is as follows:

[0139] [ka]

[0140] The oxidation reaction of the compound represented by chemical formula 15 can be carried out by treating it with TFA / H2O. The oxidation reaction of the compound represented by chemical formula 15 can be carried out within the range of solvent, temperature, time, etc., used in coupling reactions.

[0141] If R5 is a tosyl or mesyl group, the tosyl group can be removed before the coupling reaction with the compound represented by chemical formula 1 can be carried out. The tosyl or mesyl group can be removed by known methods, for example, by treating with NaBH4.

[0142] If R3 of the compound represented by chemical formula 3 of the present invention is an acetyl group; or an ethyl group substituted with a hydroxyl group, carbamate, selenide, or sulfide, then the step of converting these substituents to vinyl groups must be carried out further.

[0143] (1) When R3 is an acetyl group, the conversion to a vinyl group can be carried out by reduction and dehydration of the acetyl group. The reduction and dehydration of the acetyl group can be carried out by known methods.

[0144] (2) If R3 is an ethyl group substituted with a hydroxyl group, the conversion to a vinyl group can be carried out by a dehydration reaction of the hydroxyl group. The dehydration reaction of the hydroxyl group can be carried out by known methods.

[0145] (3) If R3 is a carbamate-substituted ethyl group, the conversion to a vinyl group can be carried out by a Hofmann removal reaction after the deprotection reaction. For example, the compound represented by chemical formula 3 (compound D-Gd) can be converted from a carbamate-substituted ethyl group to a vinyl group (compound F-13a) by removal of the protecting group and a Hofmann removal reaction under LiOH, as follows.

[0146] [ka]

[0147] (4) If R3 is an ethyl group substituted with selenide, the conversion to a vinyl group can be carried out by the oxidation reaction of selenide. For example, the compound of chemical formula 3 can be converted to a vinyl group by oxidation in the presence of HOAc and H2O2 as follows.

[0148] [ka]

[0149] (5) If R3 is an ethyl group substituted with a sulfide, the conversion to a vinyl group can be carried out by an oxidation reaction of the sulfide. For example, this can be carried out by an oxidation reaction of the compound of chemical formula 3 with mCPBA followed by a deprotection reaction with pyridine, as follows.

[0150] [ka]

[0151] The present invention will be described in more detail below with reference to examples.

[0152] <Examples> 1. Preparation of the compound represented by chemical formula 1 Compounds C, D, C1, and C2, corresponding to the compound represented by Chemical Formula 1 of the present invention, were prepared as follows (Examples 1-4).

[0153] Example 1: Preparation of Compound C (1-1) Preparation of compound A [ka] A mixture of compound SM1 (75.0 g, 238 mmol, 1.0 equivalent) and TBME (1125 mL) was mixed with a mixture of Br2 (53.2 g, 333 mmol, 1.4 equivalent) and TBME (375 mL) added dropwise under nitrogen conditions at 20°C. The mixture was stirred at 20°C for 1 hour, and complete reaction was confirmed by TLC. The solvent was removed under reduced pressure. Then, methanol (546 mL) was added to the mixture. The mixture was stirred at 50°C for 12 hours, and complete consumption of SM1 was confirmed by TLC. The mixture was cooled to 20°C and concentrated under reduced pressure. The mixture was polished with methanol (100 mL) at 20°C, and the filtered product was washed with methanol (50 mL x 2) to obtain compound A (59.0 g, 95.9 mmol, yield: 81%) as a gray solid.

[0154] 1 H NMR(400MHz,CDCl3)δ9.11(s,2H),7.41-7.25(m,10H),5.26(s,4H),3.97(s, 2H),3.58(s,6H),2.77(t,J=7.2Hz,4H),2.52(t,J=6.8Hz,4H),2.29(s,6H).

[0155] (1-2) Preparation of compound B [ka] To a mixture of the previously produced Compound A (50.0 g, 81.3 mmol, 1.0 equivalent) and THF (650 mL), Pd / C (5.00 g, 10 mol%) was added under nitrogen conditions. The gas of the mixture was removed under vacuum conditions and filled with H2 several times. The mixture was stirred at 20 °C and H2 (15 psi) for 16 hours. A mixture of Na2CO3 (8.62 g, 81.3 mmol) and H2O (50 mL) was added to the said mixture and stirred for 0.5 hour. The mixture was filtered, and acetic acid (about 10 mL) was added to the filtrate to adjust the pH to 7. The precipitate was filtered and dried to obtain Compound B (34.0 g, 78.3 mmol, yield: 96%) as a peach-colored solid.

[0156] 1 1H NMR (400 MHz, DMSO-d6) δ 11.09 (s, 2H), 3.78 (s, 2H), 3.56 (s, 6H), 2.56 (t, J = 7.2 Hz, 4H), 2.16 - 2.10 (m, 10H).

[0157] (1 - 3) Production of Compound C

Chemical formula

[0158] 1 H NMR(400MHz,CDCl3)δ10.19-10.00(m,2H),9.47(s,2H),4.05(s,2H),3.71(s,6H),2.80(t,J=7.2Hz,4H),2.61-2.45(m,4H),2.29(s,6H).

[0159] Example 2: Preparation of Compound D [ka] Lithium hydroxide (LiOH·H2O) (2.75 g, 65.6 mmol, 6.6 equivalents) was added to a mixture of methanol (100 mL), water (100 mL), and compound C (4.00 g, 9.94 mmol, 1.0 equivalent) from Example 1. This mixture was stirred at 25°C for 16 hours and then diluted with water (100 mL). 1 M hydrochloric acid was added dropwise to this mixture to adjust the pH to 2-3. The precipitate was then filtered and dried to obtain compound D (3.49 g, 9.32 mmol, yield: 94%), corresponding to the compound represented by chemical formula 1 of the present invention, as a purple solid.

[0160] 1 H NMR(400MHz,CDCl3)δ12.03(brs,2H),11.51(s,2H),9.48(s,2H),3.91(s,2H),2.54(overlapped with DMSO-d6's signal,4H),2.18(s,6H),2.06(t,J=8.0Hz,4H).

[0161] C 19 H 22 N2OS m / z [M+H] + =375

[0162] Example 3: Preparation of Compound C1 [ka] To a mixture of compound D (100 mg, 0.26 mmol, 1.0 equivalent) from Example 2 with DMSO (3 mL), DIPEA (103.56 mg, 0.80 mmol, 3.0 equivalents), HOBt (108.28 mg, 0.80 mmol, 3.0 equivalents), and EDCI (153.61 mg, 0.80 mmol, 3.0 equivalents) were added and the mixture was stirred at 25°C for 30 minutes. 1-propanol (80.26 mg, 1.34 mmol, 5.0 equivalents) was added dropwise to the mixture and the mixture was stirred for 12 hours. HCl (140 mL) was added to the mixture, and it was washed with water (120 mL x 5). The mixed organic layer was washed with brine (120 mL), dried over anhydrous Na2SO4, filtered under reduced pressure, and concentrated. The obtained residue was polished with MeOH (1 mL) and then filtered under reduced pressure to obtain compound C1 (27 mg, 0.046 mmol, yield: 21%), which corresponds to the compound represented by chemical formula 1 of the present invention, as a pink solid.

[0163] 1 H NMR(400MHz,DMSO-d6)δ11.53(brs,2H),9.47(s,2H),3.91(t,J=6.8Hz,6H),2.54(overlapped with DMSO-d6's signal,4H),2.17(s,6H),2.06(t,J=8.0Hz,4H),1.58-1.49(m,4H),0.84(t,J=7.6Hz,3H).

[0164] Example 4: Preparation of Compound C2 [ka] To a mixture of compound D (160 mg, 0.42 mmol, 1.0 equivalent) from Example 2 with DMSO (5 mL), DIPEA (165.56 mg, 1.28 mmol, 3.0 equivalents), HOBt (173.24 mg, 1.28 mmol, 3.0 equivalents), and EDCI (254.78 mg, 1.28 mmol, 3.0 equivalents) were added and the mixture was stirred at 25°C for 30 minutes. Benzyl alcohol (231.07 mg, 2.14 mmol, 5.0 equivalents) was added dropwise to the mixture and the mixture was stirred for 12 hours. SiO (180 mL) was added to the mixture, and it was washed with water (130 mL x 5). The mixed organic layer was washed with brine (130 mL), dried over anhydrous Na2SO4, filtered under reduced pressure, and concentrated. The obtained residue was polished with MeOH / MTBE (v / v=1 / 1, 10 mL), filtered under reduced pressure, and compound C2 (110 mg, 0.198 mmol, yield: 47%), corresponding to the compound represented by chemical formula 1 of the present invention, was obtained as a brown solid.

[0165] 1 H NMR(400MHz,DMSO-d6)δ11.53(s,2H),9.45(s,2H),7.37-7.26(m,10H),5.03(s,4H),3.87(s,2H),2.53(overlap with DMSO-d6's signal,4H),2.17-2.14(m,10H).

[0166] 2. Preparation of the compound represented by chemical formula 2 Compounds corresponding to the compound represented by chemical formula 2 of the present invention were prepared as follows (Examples 5-12).

[0167] Example 5: Preparation of compound Ga (5-1) Preparation of compound Ga-2 [ka] To a mixture of compound Ga-1 (55.8 g, 326 mmol, 1.1 equivalents) prepared as described above and acetone (400 mL), K2CO3 (45.0 g, 326 mmol, 1.1 equivalents) and 1-bromobut-2-ene (40.0 g, 296 mmol, 1.0 equivalent) were added, and the mixture was stirred at 60°C for 30 hours under nitrogen conditions. The mixture was filtered under reduced pressure and concentrated. The residue was purified by silica gel chromatography to obtain compound Ga-2 (42.4 g, yield: 64%) as a yellow solid.

[0168] 1 H NMR(400MHz,CDCl3)δ7.75(d,J=8.4Hz,2H),7.30(d,J=8.0Hz,2H),5.59-5.51(m,1H),5 .36-5.28(m,1H),4.95-4.78(m,1H),3.62-3.48(m,2H),2.43(s,3H),1.60-1.53(m,3H).

[0169] (5-2) Preparation of compound Ga-3 [ka] To a mixture of compound Ga-2 (5.00 g, 22.2 mmol, 1.0 equivalent) prepared as described above and THF (50 mL), NaH (1.33 g, 33.3 mmol, 60% purity in mineral oil, 1.5 equivalents) was added at 0°C. A mixture of 2,2-dichloropropanoyl chloride (9.79 g, 60.7 mmol, 2.7 equivalents) and DCM (15 mL) was added to the mixture at 0°C and stirred at 15°C for 12 hours. Water (100 mL) was added to this mixture and extracted with DCM (200 mL x 2). The mixed organic layer was dried over anhydrous Na2SO4, filtered under reduced pressure, and concentrated. The residue was purified by silica gel chromatography to obtain compound Ga-3 (4.47 g, 4.6 mmol, yield: 21%) as a yellow solid.

[0170] 1H NMR(400MHz,CDCl3)δ7.80(d,J=8.4Hz,2H),7.24(d,J=8.0Hz,2H),5.90-5.81(m,1H) ,5.60-5.50(m,1H),4.99-4.86(m,2H),2.37(s,3H),2.13(s,3H),1.75-1.69(m,3H).

[0171] (5-3) Preparation of compound Ga-4 [ka] To a mixture of compound Ga-3 (9.29 g, 26.5 mmol, 1.0 equivalent) prepared as described above and ACN (40 mL), CuCl (1.05 g, 10.6 mmol, 0.4 equivalent) was added and stirred at 110 °C for 36 hours under nitrogen conditions. This mixture was concentrated under reduced pressure. The residue was purified by silica gel chromatography to obtain compound Ga-4 (8.50 g, 24.1 mmol, yield: 91%) as a yellow oil.

[0172] C 14 H 17 Cl2NO3S m / z [M+H] + =350.0

[0173] (5-4) Preparation of compound Ga-5 [ka] The compound Ga-4 (8.50 g, 24.3 mmol, 1.0 equivalent) prepared above was mixed with DMF (90 mL) and stirred at 130 °C for 12 hours under nitrogen conditions. SiO (250 mL) was added to the mixture and washed with water (50 mL x 4). The mixed organic layer was washed with brine (50 mL x 2), dried over anhydrous Na₂SO₄, and then filtered and concentrated under reduced pressure. The residue was purified by silica gel chromatography to obtain compound Ga-5 (4.70 g, 17.0 mmol, yield: 70%) as a yellow solid.

[0174] 11H NMR (400 MHz, CDCl3) δ 7.90 (d, J = 8.4 Hz, 2H), 7.26 (d, J = 8.0 Hz, 2H), 6.62 (dd, J = 17.6, 11.2 Hz, 1H), 5.51 (d, J = 18.0 Hz, 1H), 5.43 (d, J = 10.8 Hz, 1H), 4.41 (d, J = 1.2 Hz, 2H), 2.36 (s, 3H), 1.76 (s, 3H).

[0175] (5-5) Production of Compound Ga

Chem.

[0176] 1 1H NMR (400 MHz, CDCl3) δ 6.79 (brs, 1H), 6.73 (dd, J = 17.6, 11.2 Hz, 1H), 5.45 (d, J = 17.6 Hz, 1H), 5.37 (d, J = 10.8 Hz, 1H), 4.06 (s, 2H), 1.92 (s, 3H).

[0177] C7H9NO m / z [M+H] + = 124

[0178] Example 6: Preparation of compound Gb (6-1) Production of Compound Gb-1

Chem.

[0179] 1 H NMR (400 MHz, CDCl3) δ 8.74 (brs, 1H), 7.36 (s, 1H), 6.55 (s, 1H), 2.40 (s, 3H), 2.31 (s, 3H).

[0180] (6-2) Preparation of Compound Gb <0OO0911>

Chemical Structure

[0181] 1H NMR (400MHz, CDCl3) δ7.59(brs,1H),4.12(s,2H),2.45(s,3H),2.21(s,3H).

[0182] Example 7: Preparation of compound Gc [ka] To a mixture of compound Gb (23.2 mg, 0.17 mmol, 1.0 equivalent) prepared as described above and THF (1.6 mL), 1.0 M DIBAL in THF (0.36 mL, 0.36 mmol, 2.2 equivalents) was added dropwise at -10°C and the mixture was stirred for 1 hour. The reaction was terminated with aqueous NH4Cl solution (5.0 mL), extracted with DCM (15 mL x 5), dried over anhydrous Na2SO4, and then concentrated under reduced pressure to obtain compound Gc (6.9 mg, 0.049 mmol, yield: 29.4%) as a yellow oil.

[0183] C7H 11 NO2m / z [M+H] + =142

[0184] Example 8: Preparation of compound Gd (8-1) Preparation of compound Gd-1 [ka] A mixture of compound SM1 (5 g, 15.85 mmol, 1.0 equivalent) and acetic acid (120 mL) was stirred at 25°C for 30 minutes, then SO2Cl2 (1 M in DCM, 31.7 mmol, 31.7 mL, 2.0 equivalents) was added and the mixture was stirred at 25°C for 1 hour. Further addition of SO2Cl2 (1 M in DCM, 31.7 mmol, 31.7 mL, 2.0 equivalents) was made and the mixture was stirred at 25°C for 1 hour. After adding DCM (300 mL), the mixture was washed with water (400 mL x 8). The mixed organic layer was dried over anhydrous Na2SO4, filtered under reduced pressure, and concentrated to obtain compound Gd-1 (5.9 g, yield: >99%) as a brown oil.

[0185] 1H NMR(400MHz,CDCl3)δ9.56(s,1H),7.44-7.35(m,5H),5.34(s,2H),3.67(s,3H),3.06(t,J=8.1Hz,2H),2.55(t,J=8.0Hz,2H),2.30(s,3H).

[0186] (8-2) Preparation of compound Gd-2 [ka] To a mixture of compound Gd-1 (5.9 g, 17.08 mmol, 1.0 equivalent) prepared above with dioxane (250 mL), an aqueous solution of NaHCO3 (4.3 g, 51.24 mmol, 3.0 equivalents, 250 mL of water) was added, and the mixture was stirred at 25°C for 30 minutes. I2 (4.34 g, 17.08 mmol, 1.0 equivalent) and KI (7.1 g, 42.7 mmol, 2.5 equivalents) were added to the mixture, and the mixture was stirred at 60°C for 3 hours. After cooling to 25°C, an aqueous solution of NaHSO3 (100 mL) was added, and the mixture was extracted with DCM (250 mL). The mixed organic layer was washed with brine (200 mL x 2) and dried over anhydrous Na2SO4. The mixture was filtered and concentrated under reduced pressure to obtain compound Gd-2 (6.1 g, 14.2 mmol, yield: 83%) as a brown oil.

[0187] 1 H NMR(400MHz,CDCl3)δ9.35(s,1H),7.42-7.32(m,5H),5.31(s,2H),3.67(s,3H),2.70(t,J=7.5Hz,2H),2.56(t,J=7.4Hz,2H),2.32(s,3H).

[0188] (8-3) Preparation of compound Gd-3 [ka] To a mixture of the compound Gd-2 (6.1 g, 14.28 mmol, 1.0 eq) prepared above in AcOH (142 mL) was added Zn (6.06 g, 92.69 mmol, 6.5 eq), and the mixture was stirred at 120 °C for 3 h. Zn was removed under reduced pressure. After adding DCM (300 mL) to the residue, it was washed several times with water (400 mL). The combined organic layers were dried over anhydrous Na2SO4, filtered, and concentrated under reduced pressure to obtain the compound Gd-3 (4.1 g, 13.60 mmol, yield: 95%) as a yellow oil.

[0189] 1 H NMR (400 MHz, CDCl3) δ 8.74 (s, 1H), 7.41 - 7.31 (m, 5H), 5.29 (s, 2H), 3.66 (s, 3H), 2.75 (t, J = 7.7 Hz, 2H), 2.54 (t, J = 7.7 Hz, 2H), 2.30 (s, 3H).

[0190] (8 - 4) Preparation of Compound Gd-4

Chemical formula

[0191] 1 H NMR (500 MHz, CDCl3) δ 8.94 (s, 1H), 6.76 (d, J = 3.0 Hz, 1H), 3.67 (s, 3H), 2.76 (t, J = 7.6 Hz, 2H), 2.55 (t, J = 7.6 Hz, 2H), 2.32 (s, 3H).

[0192] (8 - 5) Preparation of Compound Gd-5

Chemical formula

[0193] 1 H NMR(400MHz,CDCl3)δ9.52(s,1H),6.88(s,1H),3.62(s,3H),2.70(t,J=7.8Hz,2H),2.51(t,J=7.8Hz,2H),2.25(s,3H).

[0194] (8-6) Preparation of compound Gd-6 [ka] To a mixture of the compound Gd-5 (226 mg, 1.15 mmol, 1.0 equivalent) prepared above and methanol (5.5 mL), pyridine (0.24 mL, 2.89 mmol, 2.5 equivalents) and 30% H2O2 (0.65 mL, 5.79 mmol, 5.0 equivalents) were added, and the mixture was stirred at 60°C for 16 hours. The mixture was concentrated under reduced pressure and high temperature conditions, and the residue was purified by silica gel chromatography to obtain compound Gd-6 (122 mg, 0.66 mmol, yield: 57%).

[0195] 1 H NMR(400MHz,CDCl3)δ7.06(brs,1H),6.88(s,1H),3.8(d,J=1.3Hz,2H),3.67(s,3H),2.69(t,J=7.5Hz,2H),2.50(t,J=7.4Hz,2H),1.80(s,3H).

[0196] (8-7) Preparation of compound Gd-7 [ka] To a mixture of the compound Gd-6 (56.5 mg, 0.31 mmol, 1.0 equivalent) prepared as described above and methanol (3 mL), hydrazine monohydrate (0.38 mL, 6.78 mmol, 22.0 equivalents) was added and refluxed for 16 hours. The formation of compound Gd-7 was confirmed by LC-MS.

[0197] C8H 13 N3O2m / z [M+H] + =184

[0198] (8-8) Preparation of compound Gd [ka] The compound Gd-7 (58.6 mg, 0.32 mmol, 1.0 equivalent) prepared as described above was cooled to 0°C, and then 5N HCl aqueous solution (3.27 mmol, 10.0 equivalents) was added. 0.5M NaNO2 aqueous solution (0.36 mmol, 1.1 equivalents) was added dropwise to the mixture at 0°C and stirred for 20 minutes. After several extractions with diethyl ether, the mixed organic layer was dried over anhydrous Na2SO4 and concentrated under reduced pressure, leaving only 1 / 10 of the organic layer. Methanol (30 mL) was then added and refluxed. The mixture was concentrated under reduced pressure, and the residue was purified by silica gel chromatography to obtain compound Gd (5.9 mg, 0.029 mmol, yield: 9%), which corresponds to the compound represented by chemical formula 2 of the present invention.

[0199] 1 H NMR(500MHz,CDCl3)δ6.32(brs,1H),4.84(brs,1H),3.87(s,2H),3.66(s,3H),3.39-3.34(m,2H),2.60(t,J=6.7Hz,2H),1.81(s,3H).

[0200] Example 9: Preparation of compound Ge-7 (9-1) Preparation of compound Ge-2 [ka] To a mixture of compound 4-methylbenzenethiol (36.9 g, 297 mmol, 0.83 equivalents) in THF (300 mL) and water (150 mL), Ge-1 (20.1 g, 358 mmol, 1.0 equivalent) was added dropwise at 0°C, and the mixture was stirred at 25°C for 16 hours under nitrogen conditions. Aqueous solution of NaHCO3 (200 mL) was added to the mixture, and it was extracted with siRNA (200 mL x 2). The mixed organic layer was washed with brine (50 mL x 2), dried over anhydrous Na2SO4, filtered and concentrated under reduced pressure to obtain compound Ge-2 (59.4 g, 330 mmol, yield: 92%) as a yellow oil.

[0201] 1 H NMR(400MHz,CDCl3)δ9.74(s,1H),7.27(d,J=8.0Hz,2H),7.11(d,J=7.6Hz,2H),3.13(t,J=7.2Hz,2H),2.75-2.71(m,2H),2.32(s,3H).

[0202] (9-2) Preparation of compound Ge-3 [ka] A mixture of compound Ge-2 (58 g, 322 mmol, 1.0 equivalent) and DBU (4.90 g, 32.2 mmol, 0.1 equivalent) in THF (400 mL) was added at 0°C to a mixture of 1-nitroethane (24.0 g, 322 mmol, 1.0 equivalent) in THF (50 mL), and the mixture was stirred at 25°C for 16 hours. The mixture was diluted with water (200 mL) and extracted with siRNA (400 mL x 2). The mixed organic layer was washed with brine (200 mL), dried over anhydrous Na2SO4, and then filtered and concentrated under reduced pressure. The residue was purified by silica gel chromatography to obtain compound Ge-3 (51.7 g, 202 mmol, yield: 63%) as a yellow oil.

[0203] 1H NMR(400MHz,CDCl3)δ7.28(d,J=7.6Hz,2H),7.13(d,J=8.0Hz,2H),4.54-4.46(m,1H),4.15-4.12(m ,1H),3.15-3.08(m,1H),3.03-2.96(m,1H),2.33(s,3H),1.80-1.64(m,2H),1.53(t,J=8.0Hz,3H).

[0204] (9-3) Preparation of compound Ge-4 [ka] To a mixture of the compound Ge-3 (51.7 g, 202 mmol, 1.0 equivalent) prepared above and H2SO4 (199 mg, 2.02 mmol, 0.01 equivalent) in chloroform (500 mL), acetic anhydride (31.0 g, 304 mmol, 1.5 equivalents) was slowly added at 0°C, and the mixture was stirred at 25°C for 16 hours. After terminating the reaction with aqueous NaHCO3 solution (100 mL), the mixture was extracted with DCM (50 mL x 4). The mixed organic layer was washed with brine (50 mL x 2), dried over anhydrous Na2SO4, filtered and concentrated under reduced pressure to obtain compound Ge-4 (65.5 g, yield: >99%) as a brown oil.

[0205] 1 H NMR(400MHz,CDCl3)δ7.27(d,J=8.4Hz,2H),7.12(d,J=8.0Hz,2H),5.45-5.40(m,1H),4.75-4.66(m,1H) ,2.98-2.78(m,2H),2.33(s,3H),2.07(d,J=8.8Hz,3H),1.99-1.80(m,2H),1.49(dd,J=6.8,2.0Hz,3H).

[0206] (9-4) Preparation of compound Ge-5 [ka] To a mixture of the compound Ge-4 (3.61 g, 18.5 mmol, 1.0 equivalent) and DBU (5.63 g, 37.0 mmol, 2.0 equivalents) in ACN (50 mL), a mixture of TosMIC (5.00 g, 16.8 mmol, 0.9 equivalents) in ACN (10 mL) was added dropwise under nitrogen conditions at -40°C, and the mixture was stirred at 25°C for 16 hours. This mixture was diluted with water (100 mL) and then extracted with ELISA (100 mL x 2). The mixed organic layer was washed with brine (100 mL), dried over anhydrous Na2SO4, filtered and concentrated under reduced pressure to obtain the residue. The residue was purified by silica gel chromatography to obtain compound Ge-5 (3.47 g, 9.00 mmol, yield: 53%) as a red oil.

[0207] 1 H NMR(400MHz,CDCl3)δ8.99(s,1H),7.65(d,J=8.0Hz,2H),7.31(d,J=8.0Hz,2H),7.21(d,J=8.0Hz,2H), 7.14(d,J=8.0Hz,2H),6.69(d,J=2.0Hz,1H),2.87-2.81(m,4H),2.37(s,3H),2.35(s,3H),1.93(s,3H).

[0208] (9-5) Preparation of compound Ge-6 [ka] To a mixture of the compound Ge-5 (3 g, 7.78 mmol, 1.0 equivalent) prepared above in DCM (78 mL), PhMe3NBr3 (3.31 g, 8.56 mmol, 1.1 equivalent) was added at 0°C and the mixture was stirred for 1 hour. An aqueous solution of NaHSO3 (50 mL) was added to the mixture to terminate the reaction, and after extraction with DCM (80 mL), the organic layer was washed with water (50 mL). The mixed organic layers were dried over anhydrous Na2SO4, filtered and concentrated under reduced pressure to obtain compound Ge-6 (3.25 g, 7.00 mmol, yield: 90%).

[0209] 1H NMR(400MHz,CDCl3)δ8.92(brs,1H),7.64(d,J=8.3Hz,2H),7.29(d,J=8.1Hz,2H),7.22(d,J= 8.0Hz,2H),7.13(d,J=8.0Hz,2H),2.86-2.77(m,4H),2.37(s,3H),2.34(s,3H),1.85(s,3H).

[0210] (9-6) Preparation of compound Ge-7 [ka] To the compound Ge-6 (600 mg, 1.29 mmol, 1.0 equivalent) prepared as described above, TFA / H2O (v / v=5 / 1, 6.46 mL / 1.29 mL) was added and the mixture was stirred at 50°C for 4 hours. The reaction was terminated by neutralizing the mixture with an aqueous Na2CO3 solution. The organic layer was extracted by adding DCM (50 mL) and water (40 mL). The mixed organic layer was dried over anhydrous Na2SO4, filtered under reduced pressure, and concentrated. The residue was purified by silica gel chromatography to obtain compound Ge-7 (250 mg, 0.62 mmol, yield: 48%), which corresponds to the compound represented by chemical formula 2 of the present invention.

[0211] 1 H NMR(400MHz,CDCl3)δ7.54(d,J=6.5Hz,1H),7.20-7.17(m,4H),7.04(d,J=6.3Hz,2H),3.21-3.16(m,1H),3. 07-3.02(m,1H),2.99-2.94(m,1H),2.73-2.67(m,1H),2.37(s,3H),2.32(s,3H),2.24(s,3H),1.42(s,3H).

[0212] Example 10: Preparation of compound Ge [ka] To a mixture of the compound Ge-7 (47.2 mg, 0.12 mmol, 1.0 equivalent) prepared above with EtOH (5 mL), NaBH4 (5 mg, 0.13 mmol, 1.1 equivalent) was added and the mixture was stirred at 25°C for 30 minutes. An aqueous solution of NH4Cl (5 mL) was added to the mixture and extracted with DCM (20 mL). The mixed organic layer was dried over anhydrous Na2SO4, filtered under reduced pressure, and concentrated. The residue was purified by silica gel chromatography to obtain compound Ge (29 mg, 0.057 mmol, yield: 49%), which corresponds to the compound represented by chemical formula 2 of the present invention.

[0213] 1 H NMR(500MHz,CDCl3)δ7.19(d,J=8.1Hz,2H),7.05(d,J=8.0Hz,2H),3.78(s,2H),2.93(d,J=7.4Hz,2H),2.58(d,J=7.3Hz,2H),2.26(s,3H),1.69(s,3H).

[0214] Example 11: Preparation of compound Gf-7 (11-1) Preparation of compound Gf-1 [ka] A mixture of tert-butyl glycinate (1 g, 5.96 mmol, 1.0 equivalent) and 1 M aqueous solution of NaHCO3 (2.5 equivalents) was mixed with TsCl (1.25 g, 6.56 mmol, 1.1 equivalents) and stirred at 25°C for 24 hours. The reaction mixture was extracted with RINKAN (50 mL x 3). The mixed organic layer was washed with brine (50 mL), dried over anhydrous Na2SO4, and then filtered and concentrated under reduced pressure. The residue was purified by silica gel chromatography to obtain compound Gf-1 (1.45 g, 4.94 mmol, yield: 83%).

[0215] C 13 H 19 NO4S m / z [M+2H-C4H9] + =230

[0216] (11-2) Preparation of compound Gf-2 [ka] To a mixture of compound Gf-1 (1.3 g, 4.55 mmol, 1.0 equivalent) prepared above with THF (13 mL), 1.3 M LiHMDS in THF (4 mL, 5.23 mmol, 1.15 equivalents) was added dropwise at -78°C and the mixture was stirred for 1 hour. Propionic anhydride (0.75 mL, 5.92 mmol, 1.3 equivalents) was added at -78°C and the mixture was stirred for 1 hour, followed by stirring at 25°C for another 1 hour. The reaction was terminated with aqueous NH4Cl solution (30 mL) and extracted with DCM (40 mL). The mixed organic layer was dried over anhydrous Na2SO4, filtered under reduced pressure, and concentrated. The residue was purified by silica gel chromatography to obtain compound Gf-2 (1.4 g, 4.23 mmol, yield: 93%).

[0217] 1 H NMR (400MHz, CDCl3) δ7.88(d,J=8.3Hz,2H),7.33(d,J=8.0Hz,2H),4.50(s,2H),2.56(q,J=7.2Hz,2H),2.43(s,3H),1.44(s,9H),1.02(t,J=7.2Hz,3H).

[0218] (11-3) Preparation of compound Gf-3 [ka] To a mixture of compound Gf-2 (500 mg, 1.46 mmol, 1.0 equivalent) prepared as described above and DCM (5 mL), TFA (1.5 mL) was added at 0°C and the mixture was stirred at 25°C for 2 hours. The solid obtained by concentration under reduced pressure was filtered to obtain compound Gf-3 (384 mg, 1.34 mmol, yield: 92%).

[0219] 1H NMR(500MHz,DMSO-d6)δ13.18(brs,1H),7.88(d,J=8.1Hz,2H),7.43(d,J=8.0H z,2H),4.57(s,2H),2.47(q,J=7.2Hz,2H),2.40(s,3H),0.87(t,J=7.1Hz,3H).

[0220] (11-4) Preparation of compound Gf-4 [ka] Compound Gf-3 (100 mg, 0.35 mmol, 1.0 equivalent) was mixed with DCM (2 mL) and (COCl)2 (36 μL, 0.42 mmol, 1.2 equivalents) and DMF (2 drops) was added, and the mixture was stirred at 0°C for 1 hour under nitrogen conditions. Compound Gf-4 was obtained by concentrating under reduced pressure.

[0221] (To confirm the reactive Gf-4 by LC-MS, methanol was added to form the ester (C 13 H 17 I checked after switching to NO5S.

[0222] C 13 H 17 NO5S m / z [M+H] + =300

[0223] (11-5) Preparation of compound Gf-5 [ka] To a mixture of compound Gf-4 (1.7 mmol, 1.0 equivalent) and THF (10 mL), CuI (65 mg, 0.34 mmol, 0.2 equivalents) was added at -20°C. A 1.0 M vinylMgBr in THF solution (3.4 mmol, 3.5 mL, 2.0 equivalents) was added dropwise at -20°C, and the mixture was stirred at 25°C for 16 hours. The reaction was terminated with an aqueous NH4Cl solution (10 mL), and the mixture was extracted with ethyl acetate (30 mL). The mixed organic layer was washed with brine (20 mL), dried over anhydrous sodium 2SO4, and then filtered and concentrated under reduced pressure. The residue was purified by silica gel chromatography to obtain compound Gf-5 (40 mg, 0.56 mmol, 2-step yield: 38%).

[0224] 1 H NMR(500MHz,CDCl3)δ7.90(d,J=8.0Hz,2H),7.32(d,J=7.9Hz,2H),6.43(dd,J=17.0,10.9Hz,1H),5.5 4(d,J=17.1Hz,1H),5.46(d,J=10.8Hz,1H),4.75(s,2H),2.46(t,J=7.1Hz,2H),0.92(t,J=7.1Hz,3H).

[0225] (11-6) Preparation of compound Gf-6 [ka] To a mixture of compound Gf-5 (40 mg, 0.13 mmol, 1.0 equivalent) prepared as described above and pyridine (1.5 mL), benzeneselenol (71.5 μL, 0.65 mmol, 5.0 equivalents) was added and the mixture was stirred at 150°C in a microwave for 10 minutes. After removing the pyridine under reduced pressure and high temperature conditions, NH4Cl aqueous solution (20 mL) was added and the mixture was extracted with DCM (20 mL). The mixed organic layer was dried over anhydrous Na2SO4, filtered under reduced pressure, and concentrated to obtain compound Gf-6.

[0226] C 20 H 23 NO4SSe m / z [M+H] + =454

[0227] (11-7) Preparation of compound Gf-7 [ka] To a mixture of compound Gf-6 (30 mg, 0.066 mmol, 1.0 equivalent) and THF (1 mL), 1 M tBuOK in THF solution (0.13 mL, 0.13 mmol, 2.0 equivalents) was added dropwise at -10°C and the mixture was stirred for 30 minutes. The reaction mixture was diluted with water (15 mL) and extracted with DCM (20 mL). The mixed organic layer was washed with brine (15 mL x 2), dried over anhydrous Na2SO4, and then filtered and concentrated under reduced pressure. The residue was purified by silica gel chromatography to obtain compound Gf-7 (10 mg, 0.023 mmol, yield: 34%), which corresponds to the compound represented by chemical formula 2 of the present invention.

[0228] 1 H NMR(500MHz,CDCl3)δ7.94(d,J=10.4Hz,2H),7.48-7.46(m,2H),7.32(d,J=10.1Hz,2H),7.27-7.25( m,3H),4.27(d,J=2.1Hz,2H),2.98(t,J=9.3Hz,2H),2.74(t,J=9.2Hz,2H),2.42(s,3H),1.63(s,3H).

[0229] Example 12: Preparation of compound Gf [ka] To a mixture of compound Gf-7 (148.6 mg, 0.34 mmol, 1.0 equivalent) prepared as described above and THF (7 mL), 17.1 mL of 0.1 M SmI2in THF solution (5.0 equivalents) was added dropwise under nitrogen conditions at 0°C. The mixture was stirred at 25°C for 10 minutes, and the reaction was terminated with NaHCO3 aqueous solution (10 mL). The mixture was extracted with RINKAN (50 mL) and washed with water (30 mL x 2). The mixed organic layer was dried over anhydrous Na2SO4, filtered under reduced pressure, and concentrated. The residue was purified by silica gel chromatography to obtain compound Gf (62.3 mg, 0.22 mmol, yield: 65%), which corresponds to the compound represented by chemical formula 2 of the present invention.

[0230] 1 H NMR(400MHz,CDCl3)δ7.51-7.48(m,2H),7.28-7.26(m,2H),3.83(s,2H),3.00(t,J=7.4Hz,2H),2.75(t,J=7.5Hz,2H),1.75(s,3H).

[0231] 3. Preparation of the compound represented by chemical formula 3 The compound represented by chemical formula 3 was produced by coupling the compound represented by chemical formula 1 and the compound represented by chemical formula 2 of the present invention, as shown below (Examples 13-44).

[0232] 3.1. Production of compound F-13a by coupling of compound D and compound Ga Compound D from Example 2 and compound Ga from Example 5 were coupled to produce F-13a, which corresponds to the compound represented by chemical formula 3. [ka]

[0233] Example 13 A mixture of compound D (1.34 g, 3.57 mmol, 1.0 equivalent) and compound Ga (1.10 g, 8.93 mmol, 2.5 equivalents) in dioxane (15 mL) was mixed with piperidine (2.81 g, 28.3 mmol, 8.0 equivalents) and stirred at 100 °C for 12 hours. The mixture was concentrated under reduced pressure, and the residue was washed with CHCl3 (400 mL) and 0.1 M hydrochloric acid aqueous solution (80 mL x 2). The organic layer was separated, dried over anhydrous Na2SO4, filtered and concentrated under reduced pressure. The residue was polished with methanol (30 mL) / MTBE (30 mL) at 20 °C. Filtered under reduced pressure, compound F-13a (939 mg, 1.61 mmol, yield: 45%) was obtained as a red solid.

[0234] 1H NMR(400MHz,DMSO-d6)δ11.91(brs,2H),10.44(s,2H),10.03(s,2H),6.82(dd,J=17.6,12.0Hz,2H),6.09(s,2H),5.64(d, J=10.8Hz,2H),5.61(dd,J=8.8,1.6Hz,2H),3.98(s,2H),2.44-2.40(m,4H),2.00(s,6H),1.96-1.94(m,4H),1.92(s,6H).

[0235] 13 C NMR(400MHz,DMSO-d6)δ174.52,171.83,140.87,131.35,127.87,123.92, 123.68,122.57,122.54,120.02,99,66,34.82,24.01,19.77,9.98,9.63.

[0236] C 33 H 36 N4O6m / z [M+H] + =585

[0237] Example 14: Preparation of Compound F-13a A mixture of compound D (1.0 equivalent) and dioxane was mixed with azepane (8.0 equivalents) and compound Ga (2.5 equivalents), and the mixture was stirred at 100°C under nitrogen conditions for 16 hours. The reaction conversion rate was measured by liquid chromatography-mass spectrometry (LCMS). The standardized reaction conversion rate was calculated to be 12%.

[0238] Example 15: Preparation of Compound F-13a A mixture of compound D (1.0 equivalent) and THF was mixed with piperidine (8.0 equivalents) and compound Ga (2.5 equivalents), and stirred at 60°C under nitrogen for 16 hours. The mixture was concentrated under reduced pressure, and the residue was washed with 400 mL of CHCl3 and 0.1 M aqueous hydrochloric acid (80 mL x 2). The organic layer was separated, dried over anhydrous Na2SO4, filtered and concentrated under reduced pressure. The residue was polished with methanol (30 mL) / MTBE (30 mL) at 20°C. Filtered under reduced pressure, compound F-13a (yield: 31%) was obtained as a red solid.

[0239] Example 16: Preparation of Compound F-13a A mixture of compound D (1.0 equivalent) and methanol was mixed with piperidine (8.0 equivalents) and compound Ga (2.5 equivalents), and stirred at 100°C under nitrogen for 16 hours. The mixture was concentrated under reduced pressure, and the residue was washed with 400 mL of CHCl3 and 0.1 M aqueous hydrochloric acid (80 mL x 2). The organic layer was separated, dried over anhydrous Na2SO4, filtered and concentrated under reduced pressure. The residue was polished with methanol (30 mL) / MTBE (30 mL) at 20°C. Filtered under reduced pressure, compound F-13a (yield: 5%) was obtained as a red solid.

[0240] Example 17: Preparation of Compound F-13a A mixture of compound D (1.0 equivalent) and dioxane was mixed with piperidine (8.0 equivalents) and compound Ga (2.5 equivalents), and the mixture was stirred at 100°C under nitrogen conditions for 16 hours. After concentrating the mixture under reduced pressure, CHCl3 (400 mL) was added to the residue, and it was washed with 0.1 M hydrochloric acid aqueous solution (80 mL x 2). The organic layer was separated, dried over anhydrous Na2SO4, filtered and concentrated under reduced pressure. The residue was polished with methanol (30 mL) / MTBE (30 mL) at 20°C. Filtered under reduced pressure, compound F-13a (yield: 12%) was obtained as a red solid.

[0241] Example 18: Preparation of Compound F-13a A mixture of compound D (1.0 equivalent) and dioxane was mixed with piperazine (8.0 equivalents) and compound Ga (2.5 equivalents), and the mixture was stirred at 100°C under nitrogen conditions for 16 hours. After concentrating the mixture under reduced pressure, CHCl3 (400 mL) was added to the residue, and it was washed with 0.1 M hydrochloric acid aqueous solution (80 mL x 2). The organic layer was separated, dried over anhydrous Na2SO4, filtered and concentrated under reduced pressure. The residue was polished with methanol (30 mL) / MTBE (30 mL) at 20°C. Filtered under reduced pressure, compound F-13a (yield: 6%) was obtained as a red solid.

[0242] Example 19: Preparation of Compound F-13a A mixture of compound D (1.0 equivalent) and dioxane was mixed with azocane (8.0 equivalents) and compound Ga (2.5 equivalents), and the mixture was stirred under nitrogen conditions at 25°C for 16 hours. After concentrating the mixture under reduced pressure, CHCl3 (400 mL) was added to the residue, and it was washed with 0.1 M hydrochloric acid aqueous solution (80 mL x 2). The organic layer was separated, dried over anhydrous Na2SO4, filtered and concentrated under reduced pressure. The residue was polished with methanol (30 mL) / MTBE (30 mL) at 20°C. Filtered under reduced pressure, compound F-13a (939 mg, 1.61 mmol, yield: 22%) was obtained as a red solid.

[0243] Example 20: Preparation of compound F-13a A mixture of compound D (1.0 equivalent) and dioxane was mixed with azocane (8.0 equivalents) and compound Ga (2.5 equivalents), and stirred at 100°C under nitrogen for 16 hours. The mixture was concentrated under reduced pressure, and the residue was washed with 0.1 M hydrochloric acid aqueous solution (80 mL x 2) after adding CHCl3 (400 mL). The organic layer was separated, dried over anhydrous Na2SO4, filtered and concentrated under reduced pressure. The residue was polished with methanol (30 mL) / MTBE (30 mL) at 20°C. Filtered under reduced pressure, compound F-13a (yield: 20%) was obtained as a red solid.

[0244] 3.2. Production of compound F-13a by coupling compound D and compound Gb Compound D from Example 2 and compound Gb from Example 6 were coupled to produce D-Gb, which corresponds to the compound represented by chemical formula 3, and from there compound F-13a was produced. [ka]

[0245] Example 21: Preparation of compound D-Gb A mixture of compound D (7.4 mg, 0.02 mmol, 1.0 equivalent) and compound Gb (11.0 mg, 0.08 mmol, 2.0 equivalents) in dioxane (1 mL) was mixed with azepane (24 mg, 0.24 mmol, 6.0 equivalents) and stirred at 60°C for 12 hours. After concentration under reduced pressure, CHCl3 (50 mL) was added to the residue and washed with 0.2 N HCl aqueous solution (50 mL x 2). The organic layer was separated, dried over anhydrous Na2SO4, filtered and concentrated under reduced pressure. The residue was purified by silica gel chromatography to obtain compound D-Gb (16 mg, 0.043 mmol, yield: 22%) as a yellow solid.

[0246] C 33 H 36 N4O8m / z [M+H] + =618

[0247] 3.3. Production of compound F-13a by coupling compound D and compound Gc Compound D from Example 2 and compound Gc from Example 7 were coupled to produce D-Gc, which corresponds to the compound represented by chemical formula 3, and from there compound F-13a was produced. [ka]

[0248] Example 22: Preparation of compound D-Gc A mixture of compound D (3.66 mg, 0.0097 mmol, 1.0 equivalent) and dioxane (0.5 mL) was mixed with azepane (0.066 mL, 0.058 mmol, 6.0 equivalents) and compound Gc (6.9 mg, 0.048 mmol, 5.0 equivalents), and the mixture was stirred at 45°C under nitrogen conditions for 18 hours. The formation of compound D-Gc was confirmed by LC-MS.

[0249] C 33 H 40 N4O8m / z [M+H] + =621

[0250] 3.4. Production of compound F-13a by coupling compound D and compound Gd Compound D from Example 2 and compound Gd from Example 8 were coupled to produce D-Gd, which corresponds to the compound represented by chemical formula 3, and from there compound F-13a was produced. [ka]

[0251] Example 23: Preparation of compound D-Gd A mixture of compound D (37.0 mg, 0.099 mmol, 1.0 equivalent) and compound Gd (49.0 mg, 0.247 mmol, 2.5 equivalents) in dioxane (3 mL) was mixed with azepane (59.4 mg, 0.593 mmol, 6.0 equivalents) and stirred at 60°C for 12 hours. After concentration under reduced pressure, CHCl3 (50 mL) was added to the residue and washed with 0.2 N HCl aqueous solution (50 mL x 2). The organic layer was separated, dried over anhydrous Na2SO4, filtered and concentrated under reduced pressure. The residue was purified by silica gel chromatography to obtain compound D-Gd as a red solid.

[0252] C 37 H 46 N6O 10 m / z [M+H] + =736

[0253] Example 24: Preparation of compound D-Gda from compound D-Gd [ka] The purified compound D-Gd (6.6 mg, 0.022 mmol, 1.0 equivalent) was dissolved in MeOH (3 mL), and then 1 M LiOH aqueous solution (0.132 ml, 6.0 equivalents) was added. The mixture was stirred at 60°C for 4 hours, then diluted with CHCl3 (50 mL), and washed with 0.2 N HCl aqueous solution (50 mL x 2). The organic layer was separated, dried over anhydrous Na2SO4, filtered under reduced pressure, and concentrated to obtain the orange compound D-Gda.

[0254] C 33 H 42 N6O6m / z [M+H] + =620

[0255] 3.5. Production of compound F-13a by coupling compound D and compound Ge Compound D from Example 2 and compound Ge from Example 10 were coupled to produce D-Ge, which corresponds to the compound represented by chemical formula 3, and from there compound F-13a was produced. [ka]

[0256] Example 25: Preparation of compound D-Ge To a mixture of compound D (36.0 mg, 0.096 mmol, 1.0 equivalent) and dioxane (5 mL), azepane (65.6 μL, 0.58 mmol, 6.0 equivalents) was added and the mixture was stirred at 25°C for 10 minutes. Compound Ge (23.8 mg, 0.096 mmol, 1.0 equivalent) was added to the mixture and the mixture was stirred at 80°C for 12 hours. After concentration under reduced pressure, CHCl3 (40 mL) was added to the residue and washed with 0.2 M hydrochloric acid aqueous solution (20 mL x 2). The organic layer was separated, dried over anhydrous Na2SO4, filtered and concentrated under reduced pressure. The residue was polished with DCM (5 mL) / Hexanes (20 mL) and filtered under reduced pressure to obtain compound D-Ge (30 mg, 0.036 mmol, yield: 37%) as a brown solid.

[0257] C 47 H 52 N4O6S2m / z [M+H] + =834

[0258] Example 26: Preparation of compound F-13a from compound D-Ge A mixture of compound D-Ge (26 mg, 0.031 mmol, 1.0 equivalent) and dioxane (8 mL) was cooled to 0°C, then m-CPBA (6.9 mg, 0.04 mmol, 1.3 equivalents) was added, and the mixture was stirred at 0°C for 1 hour. NaHSO3 aqueous solution (10 mL) was added to the mixture, and it was extracted with CHCl3 (20 mL x 2). The mixed organic layer was dried over anhydrous Na2SO4, filtered, and concentrated under reduced pressure. The residue was dissolved in DMF (8 mL), pyridine (3 mL) was added, and the mixture was refluxed for 2 hours. The formation of compound F-13a was confirmed by LC-MS.

[0259] C 33 H 36 N4O6m / z [M+H] + =585

[0260] 3.6. Preparation of compound F-13a by coupling compound D and compound Gf Compound D from Example 2 and compound Gf from Example 12 were coupled to produce D-Gf, which corresponds to the compound represented by chemical formula 3, and from there compound F-13a was produced. [ka]

[0261] Example 27: Preparation of compound D-Gf To a mixture of compound D (37.3 mg, 0.099 mmol, 1.0 equivalent) and dioxane (10 mL), azepane (68.1 μL, 0.58 mmol, 6.0 equivalents) was added and the mixture was stirred at 25°C for 10 minutes. Compound Gf (55.9 mg, 0.20 mmol, 2.0 equivalents) was added to the mixture and the mixture was stirred at 80°C for 14 hours. After concentration under reduced pressure, CHCl3 (60 mL) was added to the residue and washed with 0.2 M hydrochloric acid aqueous solution (40 mL x 2). The organic layer was separated, dried over anhydrous Na2SO4, filtered and concentrated under reduced pressure. The residue was polished with RINKAN (5 mL) / Hexanes (50 mL) and filtered under reduced pressure to obtain compound D-Gf (20 mg, 0.022 mmol, yield: 22%) as a red solid.

[0262] C 45 H 48 N4O6Se2m / z [M+H] + =900

[0263] Example 28: Preparation of compound F-13 from compound D-Gf To a mixture of D-Gf (15 mg, 0.017 mmol, 1.0 eq) in THF (2 mL), HOAc (3.0 mg, 0.050 mmol, 3.0 eq) and H2O2 (7.7 mg, 0.068 mmol, 4.0 eq) were added. After that, the mixture was stirred at 25 °C for 2 hours. It was confirmed by LCMS that compound F-13a was formed.

[0264] C 33 H 36 N4O6 m / z [M+H] + =585

[0265] 3.7. Production of Compound F-13a by Coupling of Compound C and Compound Ga Compound C of Example 1 and Compound Ga of Example 5 were coupled to produce Compound C-Ga corresponding to the compound represented by Chemical Formula 3, and then Compound F-13a was produced.

Chemical Formula

[0268] C 35 H 40 N4O6m / z [M+H] + =613

[0269] Example 30: Preparation of compound C-Ga A mixture of compound C (1.0 equivalent) and dioxane was mixed with pyrrolidine (8.0 equivalents) and compound Ga (2.5 equivalents), and the mixture was stirred at 20°C under nitrogen conditions for 16 hours. The reaction conversion rate was measured by liquid chromatography-mass spectrometry (LCMS). The standardized reaction conversion rate was calculated to be 10%.

[0270] Example 31: Preparation of compound C-Ga A mixture of compound C (1.0 equivalent) and dioxane was mixed with pyrrolidine (8.0 equivalents) and compound Ga (2.5 equivalents), and the mixture was stirred at 100°C under nitrogen conditions for 1 hour. The reaction conversion rate was measured by liquid chromatography-mass spectrometry (LCMS). The standardized reaction conversion rate was calculated to be 23%.

[0271] Example 32: Preparation of compound C-Ga A mixture of compound C (1.0 equivalent) and dioxane was mixed with piperazine (8.0 equivalents) and compound Ga (2.5 equivalents), and the mixture was stirred at 20°C under nitrogen conditions for 16 hours. The reaction conversion rate was measured by liquid chromatography-mass spectrometry (LCMS). The standardized reaction conversion rate was calculated to be 14%.

[0272] Example 33: Preparation of compound C-Ga A mixture of compound C (1.0 equivalent) and dioxane was mixed with piperazine (8.0 equivalents) and compound Ga (2.5 equivalents), and the mixture was stirred at 100°C under nitrogen conditions for 3 hours. The reaction conversion rate was measured by liquid chromatography-mass spectrometry (LCMS). The standardized reaction conversion rate was calculated to be 31%.

[0273] Example 34: Preparation of compound C-Ga A mixture of compound C (1.0 equivalent) and dioxane was mixed with morpholine (8.0 equivalents) and compound Ga (2.5 equivalents), and the mixture was stirred at 25°C under nitrogen conditions for 16 hours. The reaction conversion rate was measured by liquid chromatography-mass spectrometry (LCMS). The standardized reaction conversion rate was calculated to be 4%.

[0274] Example 35: Preparation of compound C-Ga A mixture of compound C (1.0 equivalent) and dioxane was mixed with morpholine (8.0 equivalents) and compound Ga (2.5 equivalents), and the mixture was stirred at 100°C under nitrogen conditions for 16 hours. The reaction conversion rate was measured by liquid chromatography-mass spectrometry (LCMS). The standardized reaction conversion rate was calculated to be 18%.

[0275] Example 36: Preparation of compound C-Ga A mixture of compound C (1.0 equivalent) and dioxane was mixed with piperidine (8.0 equivalents) and compound Ga (2.5 equivalents), and the mixture was stirred at 20°C under nitrogen conditions for 16 hours. The reaction conversion rate was measured by liquid chromatography-mass spectrometry (LCMS). The standardized reaction conversion rate was calculated to be 27%.

[0276] Example 37: Preparation of compound C-Ga A mixture of compound C (1.0 equivalent) and dioxane was mixed with azepane (8.0 equivalents) and compound Ga (2.5 equivalents), and the mixture was stirred at 100°C under nitrogen conditions for 16 hours. The reaction conversion rate was measured by liquid chromatography-mass spectrometry (LCMS). The standardized reaction conversion rate was calculated to be 21%.

[0277] Example 38: Preparation of compound C-Ga A mixture of compound C (1.0 equivalent) and dioxane was mixed with azocane (8.0 equivalents) and compound Ga (2.5 equivalents), and the mixture was stirred at 100°C under nitrogen conditions for 16 hours. The reaction conversion rate was measured by liquid chromatography-mass spectrometry (LCMS). The standardized reaction conversion rate was calculated to be 22%.

[0278] Example 39: Preparation of compound C-Ga A mixture of compound C (1.0 equivalent) and dioxane was mixed with azepane (8.0 equivalents) and compound Ga (2.5 equivalents), and the mixture was stirred at 100°C under nitrogen conditions for 16 hours. The reaction conversion rate was measured by liquid chromatography-mass spectrometry (LCMS). The standardized reaction conversion rate was calculated to be 13%.

[0279] Example 40: Preparation of compound F-13a from compound C-Ga To a mixture of compound C-Ga (100 mg, 0.16 mmol, 1.0 equivalent) in methanol (2.5 mL) and water (0.5 mL), LiOH·H2O (41.09 mg, 0.98 mmol, 6.0 equivalents) was added and the mixture was stirred at 60°C for 2 hours. A 1 M aqueous HCl solution was added dropwise to the mixture to adjust the pH to 2-3. The resulting solid was filtered under reduced pressure to obtain a brown solid. This solid was polished with MeOH (5 mL) to obtain compound F-13a (67 mg, 0.11 mmol, yield: 70%) as a red solid.

[0280] 1 H NMR(400MHz,DMSO-d6)δ10.58(brs,2H),6.79(dd,J=17.2,11.6Hz,2H),6.05(s,2H),5.63-5.58(m,4H),3.93(s,2H),2.50(m,overlap with DMSO-d6's signal,4H),2.05(m,4H),1.98(s,6H),1.90(s,4H).

[0281] 3.8. Production of compound F-13a by coupling of compound C1 and compound Ga Compound C1 from Example 3 and compound Ga from Example 5 were coupled to produce compound C1-Ga, which corresponds to the compound represented by chemical formula 3, and from there compound F-13a was produced. [Chemical]

[0282] Example 41: Preparation of compound C1-Ga To a mixture of Compound C1 (100 mg, 0.21 mmol, 1.0 equivalent) in dioxane (1.5 mL), piperidine (0.18 g, 2.1 mmol, 10.0 equivalents) and Compound Ga (78.5 mg, 0.64 mmol, 3.0 equivalents) were added, and the mixture was stirred at 101 °C for 16 hours under nitrogen conditions. CHCl3 (40 mL) was added to the mixture, and it was washed with 0.1 M aqueous HCl solution (25 mL × 2). The combined organic layers were washed with brine (25 mL), dried over anhydrous Na2SO4, filtered under reduced pressure, and concentrated. The residue was purified by silica gel chromatography to obtain Compound C1-Ga (32.3 mg, 0.048 mmol, yield: 23%) as a red solid.

[0283] 1 H NMR (400 MHz, DMSO-d6) δ 10.53 (brs, 2H), 10.01 (brs, 2H), 6.81 (dd, J = 17.6, 11.6 Hz, 2H), 6.06 (s, 2H), 5.65 - 5.60 (m, 4H), 3.96 (s, 2H), 3.76 (t, J = 6.8 Hz, 4H), 2.50 - 2.43 (m, overlap with DMSO-d6’s signal, 4H), 1.97 (s, 6H), 1.92 (s, 6H), 1.82 (t, J = 8.0 Hz, 4H), 1.42 (q, J = 7.2 Hz, 4H), 0.75 (t, J = 7.2 Hz, 6H).

[0284] C 39 H 48 N4O6 m / z [M + H] + = 669.5

[0285] Example 42: Preparation of compound F-13a from compound C1-Ga A mixture of compound C1-Ga (60 mg, 0.089 mmol, 1.0 equivalent) and methanol (1 mL) was mixed with a mixture of LiOH·H2O (22.14 mg, 0.53 mmol, 6.0 equivalents) and water (0.5 mL), and the mixture was stirred at 60°C for 2 hours under nitrogen conditions. After adding CHCl3 (25 mL) to the mixture, a 0.1 M aqueous HCl solution (15 mL) was added to adjust the pH to acid. The mixed organic layer was dried over anhydrous Na2SO4, filtered under reduced pressure, and concentrated. DCM / Hexane was added to the residue, and the precipitated solid was filtered under reduced pressure to obtain compound F-13a (10.4 mg, 0.025 mmol, yield: 20%) as an orange solid.

[0286] C 33 H 36 N4O6m / z [M+H] + =585

[0287] 3.9. Production of compound F-13a by coupling of compound C2 and compound Ga Compound C2 from Example 4 and compound Ga from Example 5 were coupled to produce compound C2-Ga, which corresponds to the compound represented by chemical formula 3, and from there compound F-13a was produced. [ka]

[0288] Example 43: Preparation of compound C2-Ga To a mixture of compound C2 (100 mg, 0.17 mmol, 1.0 equivalent) and dioxane (2 mL), piperidine (0.14 g, 1.7 mmol, 10.0 equivalents) and compound Ga (59.1 mg, 0.48 mmol, 3.0 equivalents) were added and the mixture was stirred at 101°C for 12 hours under nitrogen conditions. CHCl3 (40 mL) was added to the mixture and washed with 0.1 M aqueous HCl (30 mL x 2). The mixed organic layer was washed with brine (40 mL), dried over anhydrous Na2SO4, filtered under reduced pressure, and concentrated. Ether / Hexanes was added to the residue, and the precipitated dark orange solid was filtered to obtain compound C2-Ga (65 mg, 0.085 mmol, yield: 50%).

[0289] 1 H NMR(400MHz,DMSO-d6)δ10.52(s,2H),10.03(s,2H),7.30-7.19(m,10H),6.75(dd,J=17.6,1 1.6Hz,2H),6.06(s,2H),5.65-5.57(m,4H),4.91(s,4H),3.97(s,2H),2.50-2.45(m,overlap with DMSO-d6's signal,4H),2.02-1.97(m,4H),1.95(s,6H),1.91(s,6H).

[0290] C 47 H 48 N4O6m / z [M+H] + =765.6

[0291] Example 44: Preparation of compound F-13a from compound C2-Ga A mixture of compound C2-Ga (30 mg, 0.039 mmol, 1.0 equivalent) and THF (1 mL) was mixed with Pd / C (2.5 mg, 10 mol%) under nitrogen conditions. The mixture was degassed under vacuum conditions and then filled several times with H2. The mixture was stirred for 4 hours at 25°C under H2 conditions. Compound F-13a was confirmed to have been formed by LC-MS.

[0292] C 33 H 36 N4O6m / z [M+H] + =585

[0293] 4. Pegylation of compounds represented by chemical formula 3 Compound FP-13a was produced by pegyrating F-13a, which corresponds to the compound represented by chemical formula 3 (Examples 45-50).

[0294] Example 45: Pegylation of compound F-13a [ka] Compound F-13a (90 mg, 0.15 mmol, 1.0 equivalent) was dissolved in DMSO (5 mL) and stirred at 25°C for 15 minutes. A mixture of CDI (37.44 mg, 0.23 mmol, 1.5 equivalents) in DMSO (2 mL) was added dropwise to the mixture and stirred at 25°C for 2 hours. mPEG was added to this mixture. 36 A mixture of -NH2 (99.56 mg, 0.061 mmol, 0.4 equivalents) and DMSO (1 mL) was added and stirred at 25°C for 4 hours. Na2CO3 aqueous solution (100 mL) was added to the mixture and extracted with CHCl3 (50 mL x 2). The separated organic layer was dried over anhydrous Na2SO4, filtered, and concentrated under reduced pressure. The residue was polished with MTBE (50 mL), and the resulting solid was dissolved in CHCl3 (50 mL) and concentrated under reduced pressure. H2O (10 mL) was added to the resulting oily residue, freeze-dried, and then purified with Sephadex LH-20 to obtain compound FP-13a (35 mg, 0.016 mmol, yield: 26%) as a red solid.

[0295] C 106 H 183 N5O 41 m / z [M] + =2182

[0296] Example 46: Pegging of F-13a A DMA mixture of compound F-13a (1.0 equivalent) was stirred at 25°C for 15 minutes. A DMA mixture of CDI (1.2 equivalents) was added dropwise to this mixture. After stirring at 25°C for 2 hours, mPEG was used. 36 A DMA mixture containing -NH2 (0.4 equivalents) was added and the mixture was stirred at 25°C for 16 hours. The reaction conversion rate was measured by liquid chromatography-mass spectrometry (LCMS). The standardized reaction conversion rate was calculated to be 34%.

[0297] Example 47: Pegging of F-13a A DMSO mixture of compound F-13a (1.0 equivalent) was stirred at 25°C for 15 minutes. A DMSO mixture of CMPI (1.2 equivalents) was added dropwise to this mixture. After stirring at 25°C for 1 hour, mPEG was added. 36A DMSO mixture containing -NH2 (0.4 equivalents) was added and stirred at 0°C for 4 hours. The reaction conversion rate was measured by liquid chromatography-mass spectrometry (LCMS). The standardized reaction conversion rate was calculated to be 26%.

[0298] Example 48: Pegging of F-13a A DMF mixture of compound F-13a (1.0 equivalent) was stirred at 25°C for 15 minutes. A DMF mixture of EDCI (1.1 equivalents), pentafluorophenol (1.1 equivalents), and DIPEA (1.1 equivalents) was added dropwise to this mixture. After stirring at 25°C for 2 hours, mPEG was used. 36 A DMF mixture containing -NH2 (0.4 equivalents) was added and the mixture was stirred at 25°C for 3 hours. The reaction conversion rate was measured by liquid chromatography-mass spectrometry (LCMS). The standardized reaction conversion rate was 10%.

[0299] Example 49: Pegging of F-13a A DCM / DMA mixture of compound F-13a (1.0 equivalent) was stirred at 25°C for 15 minutes. A DCM / DMA mixture of DCC (1.0 equivalent) and HOAt (0.2 equivalents) was added dropwise to this mixture. After stirring at 25°C for 2 hours, mPEG was performed. 36 A DCM / DMA mixture of -NH2 (0.4 equivalents) was added and the mixture was stirred at 25°C for 16 hours. The reaction conversion rate was measured by liquid chromatography-mass spectrometry (LCMS). The standardized reaction conversion rate was calculated to be 15%.

[0300] Example 50: Pegging of F-13a A DMA mixture of compound F-13a (1.0 equivalent) was stirred at 25°C for 15 minutes. A DMA mixture of DCC (1.5 equivalents) was added dropwise to this mixture. After stirring at 25°C for 2 hours, mPEG was used. 36 A DMA mixture containing -NH2 (0.4 equivalents) was added and the mixture was stirred at 25°C for 16 hours. The reaction conversion rate was measured by liquid chromatography-mass spectrometry (LCMS). The standardized reaction conversion rate was calculated to be 18%.

Claims

1. A method for synthesizing bilirubin, comprising the step of coupling a compound represented by the following chemical formula 1 and a compound represented by the following chemical formula 2 using an ether solvent and a cyclic amine organic base to produce a compound represented by chemical formula 3: 【Chemistry 1】 【Chemistry 2】 【Transformation 3】 In the above chemical formulas 1, 2, and 3, R １ and R ２ R is independently of hydrogen, a C1-C12 alkyl group, a C6-C20 aryl group, a C2-C20 heteroaryl group, a C7-C20 arylalkyl group, or a C3-C20 heteroarylalkyl group. ３ R is a vinyl group or an acetyl group; or an ethyl group substituted with a hydroxyl group, carbamate, selenide or sulfide. ４ is a hydrogen or nitrogen protecting group, R ５ R is hydrogen, a tosyl group, or a mesyl group, however, ５ If the group is a tosyl or mesyl group, it is assumed that a removal reaction to remove the tosyl or mesyl group will be performed before the coupling reaction.

2. A method for synthesizing PEGylated bilirubin, comprising the steps of synthesizing the compound represented by chemical formula 3 by the synthesis method described in claim 1, and reacting the synthesized compound represented by chemical formula 3 with polyethylene glycol (PEG).

3. The method for synthesizing PEGylated bilirubin according to claim 2, wherein the polyethylene glycol (PEG) is aminated polyethylene glycol (PEG).

4. A method for synthesizing bilirubin according to claim 1, further comprising the step of dimerizing a compound represented by the following chemical formula 7 to produce a compound represented by the chemical formula 1: 【Chemistry 4】 In the formula, R １ is the same as R １ in Chemical Formula 1, and X is an arylalkyl ester group having 8 to 20 carbon atoms, -CH ２ OH, -COOH, a halogen atom or hydrogen.

5. A method for synthesizing bilirubin according to claim 1, further comprising the step of oxidizing a compound represented by the following chemical formula 9 to produce a compound represented by the chemical formula 2: 【Transformation 5】 In the formula, R ４ R is the R of chemical formula 2 mentioned above. ４ It is the same as this.

6. A method for synthesizing bilirubin according to claim 1, further comprising the step of oxidizing and carbamate a compound represented by the following chemical formula 13 to produce a compound represented by the chemical formula 2: 【Transformation 6】 In the formula, R is an alkyl group having 1 to 12 carbon atoms, ４ R is the R of chemical formula 2 mentioned above. ４ It is the same as this.

7. A method for synthesizing bilirubin according to claim 1, further comprising the step of cyclizing a compound represented by the following chemical formula 14 to produce a compound represented by the chemical formula 2: 【Transformation 7】 In the formula, Y is selenide, and R ４ R is the R of chemical formula 2 mentioned above. ４ It is the same as this.

8. A method for synthesizing bilirubin according to claim 1, further comprising the step of oxidizing a compound represented by the following chemical formula 15 to produce a compound represented by the chemical formula 2: 【Transformation 8】 In the formula, Z is a sulfide, and R ４ and R ５ This is R of the chemical formula 2. ４ and R ５ It is the same as this.

9. The aforementioned cyclic amine organic bases are piperidine, N-methylpiperidine, N-ethylpiperidine, 2,6-dimethylpiperidine, 2,2,6,6-tetramethylpiperidine, 3-methylpiperidine, 3-ethylpiperidine, 1-methyl-4-(methylamino)piperidine, 4-aminopiperidine, pyrrolidine, 2-pyrrolidinecarboxamide, pyrrolidine-3-ol, piperazine, 2,6-dimethylpiperazine, 1-benzylpiperazine, 1-isopropylpiperazine, 2-ethylpiperazine, morpholine, 4-methylmorpholine, 2,6-dimethyl A method for synthesizing bilirubin according to claim 1, comprising selecting from the group consisting of rumorpholine, ethylmorpholine, azepane, 2-methylazepane, 4-methylazepane, 2,2,7,7-tetramethylazepane, 1,2,2-trimethylazepane, 1,2-dimethylazepane, 2,7-dimethylazepane, methylazepane-4-carboxylate, azocan, 2-methylazocan, 1,2-dimethylazocan, 1,2,2-trimethylazocan, methylazocan-2-carboxylate, 1-methylazocan, and 2-(2-methylphenyl)azocan.

10. The method for synthesizing bilirubin according to claim 1, wherein the ether solvent is selected from the group consisting of diethyl ether, tetrahydrofuran (THF), 2-methyltetrahydrofuran, and dioxane.

11. The method for synthesizing bilirubin according to claim 1, wherein the coupling is performed at a temperature of -20°C to 200°C.

12. The method for synthesizing bilirubin according to claim 1, wherein the coupling is carried out for 0.5 to 120 hours.

13. The method for synthesizing bilirubin according to claim 1, wherein the base is added in amounts of 2 to 20 moles based on 1 mole of the compound represented by chemical formula 1.

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