Cycloalkyne derivatives

Abstract

The present invention provides a novel strain-promoted azide alkynes cycloaddition reaction (strain-promoted azide alkynes cycloaddition reaction (SPAAC)), and a preparation method of the strain-promoted azide alkynes cycloaddition reaction (SPAAC), and the strain-promoted azide alkynes cycloaddition reaction (SPAAC) can be used for preparing the strain-promoted azide alkynes cycloaddition reaction. The present invention provides a novel cycloalkyne derivative having a spiro ring as a cycloalkyne derivative that can be used in SPAAC. Also provided is a method for producing a conjugate of a functional molecule using the cycloalkyne derivative.

Description

Technical Field

[0001] The present invention relates to cycloalkyne derivatives and their uses. Background Art

[0002] Click chemistry is a method of synthesizing new molecules by bonding relatively simple compounds with high reactivity and selectivity through carbon - heteroatom bond formation reactions (Non - Patent Document 1). The 1,3 - dipolar cycloaddition reaction of azides and alkynes (Azide alkyne cycloaddition: AAC) is a representative reaction of click chemistry and can construct stable 1,2,3 - triazole rings. Therefore, as a method for firmly connecting substrates to each other, it is used in the synthesis of pharmaceuticals and functional materials. In recent years, a strain - promoted azide - alkyne cycloaddition reaction (Strain - promoted azide alkyne cycloaddition (SPAAC)) that activates alkyne molecules without using a metal catalyst and reacts with azide compounds has been developed and has attracted attention (Patent Documents 1 - 3, Non - Patent Documents 2 - 11). For use in SPAAC, various cycloalkyne derivatives have been developed.

[0003] Prior Art Documents

[0004] Patent Documents

[0005] Patent Document 1: International Publication No. 2006 / 050262

[0006] Patent Document 2: Japanese Unexamined Patent Application Publication No. 2018 - 039773

[0007] Patent Document 3: Japanese Unexamined Patent Application Publication No. 2020 - 026476

[0008] Non - Patent Documents

[0009] Non - Patent Document 1: Kolb, H.C. et al., Angew. Chem. Int. Ed. 2001, 40, 2004.

[0010] Non - Patent Document 2: Dones, J.M., et al., J. Am. Chem. Soc. 2021, 143, 9489

[0011] Non - Patent Document 3: Danilkina, N.A. et al., J. Am. Chem. Soc. 2021, 143, 16519 Non - Patent Document 4: Ni, R. et al., Angew. Chem. Int. Ed. 2015, 54, 1190

[0012] Non-Patent Document 5: Kuzmin, A. et al., Bioconjugate Chem. 2010, 21, 2076

[0013] Non-Patent Document 6: Masuda, R. et al., Chem Asian J. 2020, 15, 742

[0014] Non-Patent Document 7: Debets, M. F. et al., Chem. Commun. 2010, 46, 97

[0015] Non-Patent Document 8: Jewett, J. C. et al., Chem. Soc. Rev. 2010, 39, 1272

[0016] Non-Patent Document 9: Agard, N. J. et al., J. Am. Chem. Soc. 2004, 126, 15046

[0017] Non-Patent Document 10: Kawasaki, Y. et al., Chemistry Letters, 2019, 48, 495

[0018] Non-Patent Document 11: Harris, T. et al., Mendeleev Commun. 2019, 29, 237.

[0019] Non-Patent Document 12: Teobald, B. J., Tetrahedron 2002, 58, 4133. Summary of the Invention

[0020] Technical Problem to be Solved by the Invention

[0021] The substrates of SPAAC are not only widely used as substrates for click chemistry, but also envisioned for the binding of carrier peptides and payloads in peptide-drug conjugates (PDCs). Therefore, new substrates for SPAAC are needed.

[0022] Technical Means for Solving the Problem

[0023] The inventors repeatedly conducted in-depth research and as a result, found a method for preparing a novel cycloalkyne compound having a spiro structure. In addition, it was found that the compound has the desired properties as a substrate for SPAAC, thus completing the present invention.

[0024] This specification includes the disclosure of the following inventions.

[0025] [1-1] A compound or a salt thereof, which is represented by formula (I),

[0026] Formula (I):

[0027] [Chemical Formula 1]

[0028]

[0029] In the formula, X 1 and X 2 are each independently an oxygen atom or -N(Y 1 ),

[0030] Q 1 is any one of the groups represented by [Chemical Formula 2],

[0031] [Chemical Formula 2]

[0032]

[0033] Q 2 is selected from C 2-4 alkylene; or

[0034] Q 1 is methylene, and Q 2 is any one of the groups represented by [Chemical Formula 3],

[0035] [Chemical Formula 3]

[0036]

[0037] Y 1 is selected from a hydrogen atom, a C 1-6 alkanesulfonyl optionally substituted with one or more halogen atoms, and a nitrobenzenesulfonyl,

[0038] R 1 is selected from a hydrogen atom, a C 1-6 alkyl, a (C 1-6 alkyl)carbonyl, a (C 1-6 alkoxy)carbonyl, a benzyloxycarbonyl, -CO-Q 3 -X 3 , -COO-Q 3 -X 3 , -CONR 2 -Q 3 -X 3 , -SO 2 -Q 3 -X 3 and -SO 2 NR 2 -Q 3 -X 3 ,

[0039] R 2 is a hydrogen atom or a C 1-6Alkyl

[0040] X 3 is - COR 5 、-OL 1 or - NR 3 R 4 ,

[0041] R 3 is a hydrogen atom or C 1-6 alkyl

[0042] R 4 is a hydrogen atom, (C 1-6 alkyl)carbonyl, (C 1-6 alkoxy)carbonyl, - COX 4 or benzyloxycarbonyl,

[0043] Q 3 is C 1-10 alkylene or -(CH 2 CH 2 O) n CH 2 CH 2 -

[0044] n is an integer from 1 to 10,

[0045] L 1 is a hydrogen atom or - COX 4 ,

[0046] R 5 is a hydroxyl group, X 4 or C 1-6 alkoxy,

[0047] X 4 is the group represented by [Chemical Formula 4],

[0048] [Chemical Formula 4]

[0049]

[0050] The triple bond of formula (I) is optionally protected by Co 2 (CO) 6 protection.

[0051] [1 - 2] The compound or its salt according to [1 - 1], wherein,

[0052] R 1 is selected from - CO - CH 2 - OH, benzyloxycarbonyl and tert - butoxycarbonyl.

[0053] [1 - 3] The compound or its salt according to [1 - 1] or [1 - 2], wherein,

[0054] Y 1 is C 1-6 an alkanesulfonyl group, a trifluoromethanesulfonyl group, or a 2-nitrobenzenesulfonyl group,

[0055] [1-4] The compound or its salt according to [1-3], wherein,

[0056] Y 1 is a methanesulfonyl group or a tert-butanesulfonyl group.

[0057] The compound or its salt according to any one of [1-1] to [1-4], wherein,

[0058] Q 1 is any one of the groups represented by [Chemical Formula 5].

[0059] [Chemical Formula 5]

[0060]

[0061] [1-6] The compound or its salt according to any one of [1-1] to [1-4], wherein,

[0062] Q 2 is any one of the groups represented by [Chemical Formula 6].

[0063] [Chemical Formula 6]

[0064]

[0065] [1-7] The compound or its salt according to any one of [1-1] to [1-4], wherein,

[0066] X 2 is an oxygen atom,

[0067] Q 1 is the group represented by [Chemical Formula 7],

[0068] [Chemical Formula 7]

[0069]

[0070] R 1 is benzyloxycarbonyl.

[0071] [1-8] The compound or its salt according to any one of [1-1] to [1-4], which is an 8- to 9-membered heterocycle, and Q 1 is the group represented by [Chemical Formula 8],

[0072] [Chemical Formula 8]

[0073] or

[0074] Q 2 is the group represented by [Chemical Formula 9],

[0075] [Chemical Formula 9]

[0076]

[0077] R 1 is benzyloxycarbonyl.

[0078] [1-9] The compound or a salt thereof according to [1-1] is selected from the compounds represented by [Chemical Formula 10].

[0079] [Chemical Formula 10]

[0080]

[0081] [1-10] A method for producing a conjugate of a functional molecule using the compound or a salt thereof according to any one of [1-1] to [1-9], comprising:

[0082] reacting a first functional molecule with a second functional molecule to obtain a conjugate in which the first functional molecule and the second functional molecule are linked by a linker represented by Formula IVa or Formula IVb,

[0083] The first functional molecule has a group represented by Formula (II),

[0084] Formula (II):

[0085] [Chemical Formula 11]

[0086]

[0087] wherein X 1 and X 2 are each independently an oxygen atom or -N(Y 1 ),

[0088] Q 1 is any one of the groups represented by [Chemical Formula 12],

[0089] [Chemical Formula 12]

[0090]

[0091] Q 2 is selected from C 2-4 alkylene; or

[0092] Q 1 is methylene, and Q 2 is any one of the groups represented by [Chemical Formula 13],

[0093] [Chemical Formula 13]

[0094]

[0095] Q 5 is a linker or a single bond bonded to the first functional molecule,

[0096] a second functional molecule having a group represented by formula (III),

[0097] Formula (III)

[0098] -Q 6 -N 3 (III)

[0099] Q 6 is a linker or a single bond bonded to the second functional molecule,

[0100] Formula IVa or Formula IVb:

[0101] [Chemical Formula 14]

[0102]

[0103] [1-11] The method according to [1-10], wherein,

[0104] the first functional molecule and the second functional molecule are each independently a pharmacologically active compound, a labeled compound, a peptide, a protein, a nucleic acid, a molecule used in a drug delivery system.

[0105] [1-12] The method according to [1-11], wherein,

[0106] either the first functional molecule or the second functional molecule is a pharmacologically active compound, a labeled compound, a peptide, a protein, a nucleic acid that has specific binding affinity for a target.

[0107] [1-13] A conjugate formed by connecting a first functional molecule and a second functional molecule through a linker,

[0108] the linker is represented by formula (IVa) or formula (IVb),

[0109] Formula (IVa) or formula (IVb):

[0110] [Chemical Formula 15]

[0111]

[0112] wherein, X 1 、X 2 、Q 1 、Q 2and Q 6 as defined in [1-10].

[0113] [1-14] The conjugate according to [1-13], wherein

[0114] the first functional molecule and the second functional molecule are each independently a pharmacologically active compound, a labeled compound, a peptide, a protein, a nucleic acid, a molecule used in a drug delivery system.

[0115] [1-15] The conjugate according to [1-14], wherein

[0116] either the first functional molecule or the second functional molecule is a pharmacologically active compound, a labeled compound, a peptide, a protein, a nucleic acid that has specific binding affinity for a target.

[0117] [1-16] A method for producing a compound represented by formula (I), comprising:

[0118] obtaining a corresponding ring-closed product by treating a compound represented by formula (V) in the presence of a Lewis acid,

[0119] Formula (I):

[0120] [Chemical formula 16]

[0121]

[0122] wherein X 1 , X 2 , Q 1 and Q 2 are as defined in [1-1], and the triple bond in formula (I) is optionally protected by Co 2 (CO) 6 ,

[0123] Formula (V):

[0124] [Chemical formula 17]

[0125]

[0126] wherein X 1 , X 2 , Q 1 and Q 2 are as defined in [1-1], and wherein, in the presence of a hydroxyl group and an amino group, they are protected by a protecting group, and L is a leaving group.

[0127] [1-17] The method according to [1-16], wherein

[0128] L is -OR 10 or a halogen atom, R10 is a hydrogen atom, (C 1-6 alkyl)carbonyl, acetyl, tris(C 1-6 alkyl)silyl, C 1-6 alkyl, benzenesulfonyl or C 1-6 alkanesulfonyl.

[0129] [2-1] A compound or its salt, which is represented by formula (I),

[0130] Formula (I):

[0131] [Chemical formula 18]

[0132]

[0133] In the formula, X 1 and X 2 are each independently an oxygen atom or -N(Y 1 ),

[0134] Q 1 is any one of the groups represented by [Chemical formula 19],

[0135] [Chemical formula 19]

[0136]

[0137] Q 2 is selected from C 2-4 alkylene; or

[0138] Q 1 is methylene, Q 2 is any one of the groups represented by [Chemical formula 20-1],

[0139] [Chemical formula 20-1]

[0140]

[0141] Y 1 is selected from a hydrogen atom, C 1-6 alkanesulfonyl and nitrobenzenesulfonyl optionally substituted with one or more halogen atoms,

[0142] R 1 is selected from a hydrogen atom, C 1-6 alkyl, (C 1-6 alkyl)carbonyl, (C 1-6 alkoxy)carbonyl, benzyloxycarbonyl, [(9H-fluoren-9-yl)methoxy]carbonyl, -CO-Q 3 -X 3 、-COO-Q 3 -X 3 、-CONR 2 -Q3 -X 3 、-SO 2 -Q 3 -X 3 and-SO 2 NR 2 -Q 3 -X 3 ,

[0143] R 2 A hydrogen atom or C 1-6 alkyl,

[0144] X 3 For-COR 5 , -OL 1 、-NR 3 R 4 or a group represented by [Chemical Formula 20-2],

[0145] [Chemical formula 20-2]

[0146]

[0147] R 3 A hydrogen atom or C 1-6 alkyl,

[0148] R 4 is a hydrogen atom, (C 1-6 alkyl)carbonyl, (C 1-6 Alkoxy)carbonyl, -COX 4 or benzyloxycarbonyl,

[0149] Q 3 C 1-10 Alkylene, Ring C 3-10 Alkylene, with ring C inserted 3-10 C of alkylene 2-10 Alkylene, -(ring C 3-10 Alkylene)-(C 1-10 Alkylene)-, -(C 1-10 Alkylene)-(ring C 3-10 alkylene)- or -(CH 2 CH 2 O) n CH 2 CH 2 -,

[0150] n is an integer from 1 to 10,

[0151] L 1 is a hydrogen atom, -COX 4 or (C 1-6 alkyl)diphenylsilyl,

[0152] R 5 is a hydroxyl group, X 4 or C 1-6 alkoxy group,

[0153] X 4 is the group shown in [Chemical Formula 21],

[0154] [Chemical Formula 21]

[0155]

[0156] The triple bond in formula (I) is optionally protected by Co 2 (CO) 6 protection.

[0157] [2-2] The compound or its salt according to [2-1], wherein,

[0158] Q 3 is ethylene or propylene.

[0159] [2-3] The compound or its salt according to [2-1] or [2-2], wherein,

[0160] R 1 is selected from -CO-CH 2 -OH, benzyloxycarbonyl, tert-butoxycarbonyl, [(9H-fluoren-9-yl)methoxy]carbonyl.

[0161] [2-4] The compound or its salt according to any one of [2-1] to [2-3], wherein,

[0162] Y 1 is C 1-6 alkylsulfonyl, trifluoromethanesulfonyl and 2-nitrobenzenesulfonyl,

[0163] [2-5] The compound or its salt according to [2-4], wherein,

[0164] Y 1 is mesyl or tert-butanesulfonyl.

[0165] [2-6] The compound or its salt according to any one of [2-1] to [2-5], wherein Q 1 is any one of the groups shown in [Chemical Formula 22].

[0166] [Chemical Formula 22]

[0167]

[0168] [2-7] The compound or its salt according to any one of [2-1] to [2-5], wherein Q 2 is any one of the groups shown in [Chemical Formula 23].

[0169] [Chemical formula 23]

[0170]

[0171] [2-8] The compound or its salt according to any one of [2-1] to [2-5], wherein Q 1 is a group represented by [Chemical formula 24],

[0172] [Chemical formula 24]

[0173]

[0174] R 1 is benzyloxycarbonyl.

[0175] [2-9] The compound or its salt according to any one of [2-1] to [2-5], wherein Q 1 is a group represented by [Chemical formula 25],

[0176] [Chemical formula 25]

[0177]

[0178] Q 2 is selected from C 2-3 alkylene or Q 2 is a group represented by [Chemical formula 26],

[0179] [Chemical formula 26]

[0180]

[0181] R 1 is benzyloxycarbonyl.

[0182] [2-10] The compound or its salt according to [2-1], which is selected from the compounds represented by [Chemical formula 27].

[0183] [Chemical formula 27]

[0184]

[0185] [2-11] A method for producing a conjugate of a functional molecule using the compound or its salt according to any one of [2-1] to [2-10], which comprises:

[0186] reacting a first functional molecule with a second functional molecule to obtain a conjugate in which the first functional molecule and the second functional molecule are linked by a linker represented by formula IVa or formula IVb,

[0187] The first functional molecule has a group represented by formula (II),

[0188] Formula (II):

[0189] [Chemical Formula 28]

[0190]

[0191] Wherein, X 1 and X 2 are each independently an oxygen atom or -N(Y 1 )-,

[0192] Q 1 is any one of the groups represented by [Chemical Formula 29],

[0193] [Chemical Formula 29]

[0194]

[0195] Q 2 is selected from C 2-4 alkylene; or

[0196] Q 1 is methylene, and Q 2 is any one of the groups represented by [Chemical Formula 30],

[0197] [Chemical Formula 30]

[0198]

[0199] Q 5 is a linker or a single bond bonded to the first functional molecule,

[0200] A second functional molecule having a group represented by formula (III),

[0201] Formula (III)

[0202] -Q 6 -N 3 (III)

[0203] Q 6 is a linker or a single bond bonded to the second functional molecule,

[0204] Formula IVa or Formula IVb:

[0205] [Chemical Formula 31]

[0206]

[0207] [2-12] According to the method described in [2-11], wherein,

[0208] The first functional molecule and the second functional molecule are each independently a pharmacologically active compound, a labeled compound, a peptide, a protein, a nucleic acid, or a molecule used in a drug delivery system.

[0209] [2-13] According to the method described in [2-12], wherein,

[0210] Either the first functional molecule or the second functional molecule is a pharmacologically active compound, a labeled compound, a peptide, a protein, a nucleic acid that has specific binding affinity for a target.

[0211] [2-14] A conjugate formed by connecting the first functional molecule and the second functional molecule through a linker,

[0212] The linker is represented by formula (IVa) or formula (IVb),

[0213] Formula (IVa) or formula (IVb):

[0214] [Chemical formula 32]

[0215]

[0216] In the formula, X 1 , X 2 , Q 1 , Q 2 and Q 6 are as defined in [2-11].

[0217] [2-15] According to the conjugate described in [2-14], wherein,

[0218] The first functional molecule and the second functional molecule are each independently a pharmacologically active compound, a labeled compound, a peptide, a protein, a nucleic acid, or a molecule used in a drug delivery system.

[0219] [2-16] According to the conjugate described in [2-15], wherein,

[0220] Either the first functional molecule or the second functional molecule is a pharmacologically active compound, a labeled compound, a peptide, a protein, a nucleic acid that has specific binding affinity for a target.

[0221] [2-17] A method for producing a compound represented by formula (I), which comprises:

[0222] By treating the compound represented by formula (V) in the presence of an acid to obtain the corresponding closed-loop body,

[0223] Formula (I):

[0224] [Chemical formula 33]

[0225]

[0226] In the formula, X 1 and X 2 , Q 1 and Q 2 are defined as in [2-1]. The triple bond of formula (I) is optionally protected by Co 2 (CO) 6 .

[0227] Formula (V):

[0228] [Chemical formula 34]

[0229]

[0230] In the formula, X 1 and X 2 , Q 1 and Q 2 are defined as in [2-1]. In the presence of hydroxyl and amino groups, they are protected by protecting groups, and L is a leaving group.

[0231] [2-18] According to the manufacturing method described in [2-17], wherein

[0232] the acid is a Lewis acid or a Bronsted acid.

[0233] [2-19] According to the manufacturing method described in [2-17], wherein

[0234] L is -OR 10 or a halogen atom, and R 10 is a hydrogen atom, (C 1-6 alkyl)carbonyl, acetyl, tris(C 1-6 alkyl)silyl, C 1-6 alkyl, benzenesulfonyl or C 1-6 alkanesulfonyl.

[0235] Advantages of the invention

[0236] The novel cycloalkyne derivative with a spiro ring of the present invention has high versatility as a cycloalkyne derivative applicable to SPAAC and is also useful for the conjugation of PDC. Description of the drawings

[0237] Figure 1 is the HPLC analysis result obtained by analyzing the synthesis of PN-S01-10 synthesized in [Example 13-1] under Analytical condition 1.

[0238] Figure 2 ​​are the HPLC analysis results obtained by analyzing the lyophilized product of PN-S01-10 obtained in [Example 13-4] under Analysis Condition 1.

[0239] Figure 3 are the molecular weight results obtained by analyzing the lyophilized product of PN-S01-10 obtained in [Example 13-4] under Analysis Condition 1.

[0240] Figure 4 represents the structural formula of a nucleic acid compound (synthesized according to the general synthesis method with reference to US20180010126. Hereinafter, it may be referred to as NS-S01-20). Detailed implementation mode

[0241] In one aspect of the present invention, a novel cycloalkyne derivative that can be used for SPAAC is provided.

[0242] In this specification, the "halogen atom" refers to a fluorine atom, a chlorine atom, a bromine atom, an iodine atom, etc. When a halogen atom is used as a substituent for an aryl group, a heteroaryl group, etc., for example, a fluorine atom, a chlorine atom, and a bromine atom can be mentioned. In this specification, when a halogen atom is used as a substituent for an alkyl group or a group containing an alkyl group as part of a group (alkoxy group, alkenyl group, alkylthio group, etc.), for example, a fluorine atom can be mentioned. Specific examples of the group having a halogen atom as a substituent include trifluoromethyl, pentafluoroethyl, trifluoromethoxy, pentafluoroethoxy, trifluoromethylthio, and pentafluoroethylthio, etc.

[0243] In this specification, "C 1-6 alkyl" refers to a monovalent group derived by removing any one hydrogen atom from a straight-chain or branched saturated aliphatic hydrocarbon having 1 to 6 carbon atoms. In specific examples, methyl, ethyl, n-propyl, isopropyl, n-butyl, isobutyl, sec-butyl, tert-butyl, 1-methylpropyl, n-pentyl, isopentyl, 2-methylbutyl, 1,1-dimethylpropyl, 1-ethylpropyl, hexyl, 4-methylpentyl, etc. can be mentioned.

[0244] In this specification, "C 1-6 alkoxy" refers to a C 1-6 alkyl - O - group, where C 1-6 alkyl is as defined above. Specific examples include methoxy, ethoxy, 1-propoxy, 2-propoxy, n-butoxy, isobutoxy, sec-butoxy, tert-butoxy, 1-pentyloxy, 1-hexyloxy, etc.

[0245] In this specification, "(C 1-6 alkyl)carbonyl" refers to a C 1-6 alkyl - C(O) - group, where C 1-6 ​​The alkyl group is as defined above. Specific examples include: methylcarbonyl, ethylcarbonyl, n-propylcarbonyl, isopropylcarbonyl, n-butylcarbonyl, isobutylcarbonyl, sec-butylcarbonyl, tert-butylcarbonyl, 1-methylpropylcarbonyl, n-pentylcarbonyl, isopentylcarbonyl, 2-methylbutylcarbonyl, 1,1-dimethylpropylcarbonyl, 1-ethylpropylcarbonyl, hexylcarbonyl, 4-methylpentylcarbonyl, 2-ethylbutylcarbonyl, and the like.

[0246] In this specification, “(C 1-6 alkyloxy)carbonyl” means a C 1-6 alkyloxy-C(O)- group, where C 1-6 alkyloxy is as defined above. Specific examples include: methoxycarbonyl, ethoxycarbonyl, 1-propoxycarbonyl, 2-propoxycarbonyl, n-butoxycarbonyl, isobutoxycarbonyl, sec-butoxycarbonyl, tert-butoxycarbonyl, 1-pentyloxycarbonyl, 1-hexyloxycarbonyl, and the like.

[0247] In this specification, “tri(C 1-6 alkyl)silyl” means a (C 1-6 alkyl) 3 Si- group, where C 1-6 alkyl is as defined above, and the three alkyl groups may be the same or different from each other. Specific examples include: trimethylsilyl, triethylsilyl, tripropylsilyl, tributylsilyl, tripentylsilyl, trihexylsilyl, and the like.

[0248] In this specification, “(C 1-6 alkyl)diphenylsilyl” means a (C 1-6 alkyl)(C 6 H 5 ) 2 Si- group, where C 1-6 alkyl is as defined above. Among specific examples, tert-butyldiphenylsilyl can be cited.

[0249] In this specification, “C 1-6 alkanesulfonyl” means a C 1-6 alkyl-SO 2 - group, where C 1-6 alkyl is as defined above. Among specific examples, methylsulfonyl, ethylsulfonyl, n-propylsulfonyl, etc. can be cited. For example, methylsulfonyl can be cited.

[0250] In this specification, “C 1-10 alkylene” means a group having a -(C n H 2n )- structure, where n is any integer from 1 to 10, and unless otherwise specified, it may be substituted or unsubstituted, and may be linear or branched. C 1-10 alkylene includes C1-6 Alkylene group. Specifically, methylene, ethylene, propylene, butylene, pentylene, hexylene, heptylene, octylene, nonylene, and decylene can be mentioned.

[0251] In addition, as the C in which a cyclic C 3-10 Alkylene group 2-10 Alkylene group, ethylene inserted with cyclohexylene, propylene inserted with cyclohexylene, butylene inserted with cyclohexylene, pentylene inserted with cyclohexylene, octylene inserted with cyclohexylene, nonylene inserted with cyclohexylene, decylene inserted with cyclohexylene, ethylene inserted with cycloheptylene, propylene inserted with cycloheptylene, butylene inserted with cycloheptylene, pentylene inserted with cycloheptylene, octylene inserted with cycloheptylene, nonylene inserted with cycloheptylene, decylene inserted with cycloheptylene, ethylene inserted with cyclooctylene, propylene inserted with cyclooctylene, butylene inserted with cyclooctylene, pentylene inserted with cyclooctylene, octylene inserted with cyclooctylene, nonylene inserted with cyclooctylene, decylene inserted with cyclooctylene can be mentioned.

[0252] In -(cyclic C 3-10 Alkylene group)-(C 1-10 Alkylene group)-, -(C 1-10 Alkylene group)- is connected to X 3 In -(C 1-10 Alkylene group)-(cyclic C 3-10 Alkylene group)-, -(cyclic C 3-10 Alkylene group)- is connected to X 3 As -(C 1-10 Alkylene group)-(cyclic C 3-10 Alkylene group)-, methylenecyclopropyl, methylenecyclobutyl, methylenecyclopentyl, methylenecyclohexyl, methylenecycloheptyl, methylenecyclooctyl, methylenecyclononyl, methylenecyclodecyl, ethylenecyclopropyl, ethylenecyclobutyl, ethylenecyclopentyl, ethylenecyclohexyl, ethylenecycloheptyl, ethylenecyclooctyl, ethylenecyclononyl, ethylenecyclodecyl, propylenecyclopropyl, propylenecyclobutyl, propylenecyclopentyl, propylenecyclohexyl, propylenecycloheptyl, propylenecyclooctyl, propylenecyclononyl, propylenecyclodecyl, butylenecyclopropyl, butylenecyclobutyl, butylenecyclopentyl, butylenecyclohexyl, butylenecycloheptyl, butylenecyclooctyl, butylenecyclononyl, butylenecyclodecyl, pentylenecyclopropyl, pentylenecyclobutyl, pentylenecyclopentyl, pentylenecyclohexyl, pentylenecycloheptyl, pentylenecyclooctyl, pentylenecyclononyl, pentylenecyclodecyl can be mentioned. For example, methylene(1,4-cyclohexylene)- can be mentioned. As -(cyclic C3-10 alkylene)-(C 1-10 alkylene)-, examples include cyclopropylmethylene, cyclobutylmethylene, cyclopentylmethylene, cyclohexylmethylene, cycloheptylmethylene, cyclooctylmethylene, cyclononylmethylene, cyclodecylmethylene, cyclopropylethylene, cyclobutylethylene, cyclopentylethylene, cyclohexylethylene, cycloheptylethylene, cyclooctylethylene, cyclononylethylene, cyclodecylethylene, cyclopropylpropyl, cyclobutylpropyl, cyclopentylpropyl, cyclohexylpropyl, cycloheptylpropyl, cyclooctylpropyl, cyclononylpropyl, cyclodecylpropyl, cyclopropylbutyl, cyclobutylbutyl, cyclopentylbutyl, cyclohexylbutyl, cycloheptylbutyl, cyclooctylbutyl, cyclononylbutyl, cyclodecylbutyl, cyclopropylpentyl, cyclobutylpentyl, cyclopentylpentyl, cyclohexylpentyl, cycloheptylpentyl, cyclooctylpentyl, cyclononylpentyl, cyclodecylpentyl.

[0253] In this specification, "C 2-4 alkylene" means a group having a -(C m H 2m )- structure, which can be linear or branched, for example linear. Here, m is any integer from 2 to 4. Specifically, examples include ethylene, propylene, and butylene.

[0254] In one aspect of the present invention, the compound of formula (I) is an 8- to 9-membered heterocyclic compound. In this specification, "8- to 9-membered heterocycle" means a heterocyclic group containing N or O as a heteroatom and having 8 to 9 ring-forming atoms. In this specification, the "8- to 9-membered heterocycle" may have one triple bond.

[0255] In this specification, the ● (black dot) in the chemical formula represents the bonding point.

[0256] In this specification, "optionally substituted" and "substituted" mean that, in the case where the number of substituents is not clearly described (for example: "one or more", "1 to 3", "1 or 2", "2", "1"), they are respectively "optionally substituted by one substituent" and "substituted by one substituent". For example, "B optionally substituted by A" and "B substituted by A" respectively mean "B optionally substituted by one A" and "B substituted by one A".

[0257] In this specification, a "functional molecule" refers to a molecule that, without particular limitation, confers a desired function through binding. Examples of the desired functions include, but are not limited to, pharmacological activity functions, labeling functions, purification functions, and delivery functions to target sites. The "functional molecules" in this specification include: pharmacologically active compounds, labeling compounds, peptides, proteins, nucleic acids, and molecules used in drug delivery systems. The molecules can be low-molecular-weight compounds, medium-molecular-weight compounds, or high-molecular-weight compounds.

[0258] In this specification, a "pharmacologically active compound" refers to a compound having pharmacological activity. A pharmacologically active compound is, for example, a low-molecular-weight compound having pharmacological activity.

[0259] In this specification, a "labeling compound" refers to a compound labeled with a pigment, a fluorescent substance, a tag, or a radioisotope. Among them, a compound labeled with a radioisotope refers to a compound such as a low-molecular-weight or medium-molecular-weight compound or an antibody labeled with a radioisotope. In addition, a "labeling compound" can be a pigment, a fluorescent substance, a tag, or a radioisotope itself. The radioisotope can form a coordination bond with a chelating agent such as DOTA.

[0260] In this specification, with respect to a "peptide", any peptide can be used as long as it exhibits a useful function in the body.

[0261] In this specification, with respect to a "protein", any protein can be used as long as it is a protein present in a living organism and exhibits a useful function in the body. For example, proteins having pharmacological effects, proteins having molecular recognition effects, etc. can be mentioned. Specifically, exportin / importin, fibronectin, avidin, antibodies, enzymes, etc. can be mentioned.

[0262] In this specification, with respect to a "nucleic acid", any nucleic acid can be used as long as it is a polymer of nucleotides. Specifically, DNA and RNA can be mentioned. In this specification, nucleic acids can be modified. For example, modifications such as 2'-methoxyethyl (MOE) modification, use of 2'-deoxynucleosides in RNA, internucleoside bond based on phosphorothioate, internucleoside bond based on phosphodiester, and conversion of cytosine to 5-methylcytosine can be mentioned.

[0263] In this specification, with respect to a "linker that binds to a functional molecule", a known linker can be used. For example, an amino acid linker (peptide linker), a fatty acid linker, a nucleic acid linker, a sugar linker, and other chemical linkers (alkyl linker, PEG linker, etc.) can be mentioned. In addition, for example, a complex of a chemical linker and a peptide linker, etc. can be used.

[0264] In this specification, a "drug delivery system" refers to a system that delivers an active ingredient into target cells. As examples, carriers such as water-soluble polymers, nanoparticles (nanospheres), liposomes, and micelles can be cited. The interior of a drug delivery system (DDS) molecule can further contain drugs such as low molecular weight compounds, proteins, peptides, nucleic acids, and vaccines.

[0265] In this specification, a "conjugate" refers to a complex of functional molecules containing two or more functional molecules. Here, the functional molecules contained in the "conjugate" can be the same functional molecules or a combination of different types of functional molecules.

[0266] In this specification, for the compound represented by formula (I), the R 1 can be a conventional protecting group such as an Fmoc group or a Boc group.

[0267] In this specification, for the compound represented by formula (I), the X 4 can be, for example, a leaving group such as p-nitrophenoxy.

[0268] In this specification, as salts of the compound represented by formula (I), acid addition salts or base addition salts can be cited. As acid addition salts, for example, hydrochloride, hydrobromide, hydroiodide, phosphate, phosphonate, sulfate, etc.; sulfonates such as methanesulfonate, ethanesulfonate, benzenesulfonate, and p-toluenesulfonate; carboxylates such as acetate, citrate, malate, tartrate, succinate, salicylate, maleate, fumarate, benzoate, malonate, glycolate, oxalate, glucuronate, adipate, glutarate, ketoglutarate, and hippurate, etc. As base addition salts, for example, they include: alkali metal salts such as sodium salt and potassium salt; alkaline earth metal salts such as magnesium salt and calcium salt; ammonium salts such as ammonium salt, alkylammonium salt, dialkylammonium salt, trialkylammonium salt, and tetraalkylammonium salt; amino acid salts such as lysine salt, arginine salt, glycine salt, valine salt, threonine salt, serine salt, proline salt, and alanine salt, etc. These salts are prepared by contacting the compound with an acid or a base that can be used in the manufacture of pharmaceuticals.

[0269] In this specification, the compound represented by formula (I) or its salt can be an anhydrate or can form a solvate such as a hydrate. The so-called "solvate" here refers to a solid in which compound molecules form a complex with solvent molecules. For example, when the solvent is water, it is called a hydrate. As solvates other than hydrates, solids containing alcohols (e.g., methanol, ethanol, n-propanol) or N,N-dimethylformamide, etc. can be cited.

[0270] In addition, the compounds represented by formula (I) and their salts may exist in several tautomeric forms, such as keto and enol forms, imine and enamine forms, and mixtures thereof. Tautomers exist as a mixture of tautomers in solution. In the solid form, usually one type of tautomer is dominant. Sometimes one tautomer is described, and the present invention includes all tautomers of the compounds of the present invention.

[0271] The present invention includes all stereoisomers of the compounds represented by formula (I) (e.g., enantiomers, diastereomers (including cis and trans geometric isomers)), racemates and other mixtures of said isomers. For example, the compounds of the present invention may have more than one asymmetric point, and the compounds of the present invention include racemic mixtures, diastereomeric mixtures and enantiomers of such compounds.

[0272] When the compound represented by formula (I) is obtained as a free form, it can be converted into the state of a salt, hydrate or solvate that the compound can form by a conventional method.

[0273] In addition, when the compound represented by formula (I) is obtained as a salt, hydrate or solvate of the compound, it can be converted into the free form of the compound by a conventional method.

[0274] The elements constituting the compounds represented by formula (I) may be any isotopes, and the present invention includes compounds of formula (I) containing isotopes. With respect to the isotopes of the compounds, they are isotopes obtained by substituting at least one atom with an atom having the same atomic number (number of protons) and a different mass number (sum of the number of protons and neutrons). Examples of isotopes included in the compounds of the present invention are hydrogen atoms, carbon atoms, nitrogen atoms, oxygen atoms, sulfur atoms, fluorine atoms, chlorine atoms, etc., respectively including: 2 H, 3 H, 13 C, 14 C, 15 N, 17 O, 18 O, 35 S, 18 F, 36 Cl, etc. In particular, radioactive isotopes such as 3 H and 14 C that decay by emitting radiation are useful in in vivo tissue distribution tests of drugs or compounds, etc. Stable isotopes do not decay, have an almost constant abundance, and have no radiation, so they can be used safely. The isotopes of the compounds of the present invention can be converted by a conventional method by replacing the reagents used in the synthesis with reagents containing the corresponding isotopes.

[0275] In this specification, when L in formula (V) is a halogen atom, examples of the halogen atom include a chlorine atom. Examples of R of the compound represented by formula (V) 10 include a hydrogen atom, a methyl group, an acetyl group, a tert-butyldimethylsilyl group, a trimethylsilyl group, a benzenesulfonyl group, a methanesulfonyl group, etc. In addition, when XH of the compound represented by formula (V) 1 is a hydroxyl group, the hydroxyl group can be protected by a silyl group such as TMS or TBS, etc.

[0276] In one aspect, the compound represented by formula (I) can be used to form a 1,2,3-triazole ring (Example 6) based on a 1,3-dipolar cycloaddition reaction with an azide. That is, the present invention provides a method for producing a conjugate of a functional molecule by forming a 1,2,3-triazole ring using the compound represented by formula (I).

[0277] In one mode, the reaction can be carried out in a solvent. Examples of the solvent that can be used include dimethyl sulfoxide, acetonitrile, N,N-dimethylformamide, etc. Regarding the temperature when carrying out the reaction, for example, it is -78°C to 50°C, specifically 0°C to 25°C. In one mode, the compound represented by formula (I) can be used in an amount of, for example, 0.1 to 10 equivalents, specifically 0.5 to 1.5 equivalents, relative to the reactive azide group. The reaction time of the reaction can be appropriately set by those skilled in the art, for example, it is 10 minutes to 96 hours, more specifically 30 minutes to 2 hours.

[0278] In one aspect, for the first functional molecule having a group represented by formula (II), it can be used to form a conjugate based on a 1,3-dipolar cycloaddition reaction with a second functional molecule containing an azide group represented by formula (III), and the conjugate is formed by connecting functional molecules through a linker represented by formula (IVa) or (IVb) containing a 1,2,3-triazole ring. In one mode, the reaction can be carried out in a solvent. Examples of the solvent that can be used include dimethyl sulfoxide, N,N-dimethylformamide, N-methylpyrrolidone, acetonitrile, methanol, ethanol, tetrahydrofuran, dioxane, dichloromethane, a mixed solvent of the solvent and water, etc., for example, dimethyl sulfoxide or N,N-dimethylformamide. Regarding the temperature when carrying out the reaction, for example, it is 0°C to 100°C, specifically 0°C to 25°C. In one mode, the first functional molecule can be used in an amount of, for example, 0.5 to 1.5 equivalents, specifically 0.95 to 1.1 equivalents, relative to the reactive second functional molecule.

[0279] In one aspect, the compound represented by formula (I) can be obtained by subjecting the compound represented by formula (V) to ring closure through the Nicholas reaction (Non-Patent Document 12) and then treating it with ammonium cerium(IV) nitrate or tetrabutylammonium fluoride. That is, the present invention provides a method for producing the compound represented by formula (I), which includes: a step of subjecting the compound represented by formula (V) to ring closure through the Nicholas reaction and then treating it with ammonium cerium(IV) nitrate or tetrabutylammonium fluoride. In one mode, the Nicholas reaction can be carried out in a solvent. Examples of the solvent that can be used include dichloromethane, toluene, and tetrahydrofuran, and dichloromethane is, for example, used. In one mode, examples of the Lewis acid that can be used in the Nicholas reaction include boron trifluoride-diethyl ether complex, diethylaluminum chloride, ethylaluminum dichloride, trimethylsilyl trifluoromethanesulfonate, zinc chloride, and silver trifluoromethanesulfonate. Examples of the Bronsted acid that can be used in the Nicholas reaction include p-toluenesulfonic acid, trifluoroacetic acid, trifluoromethanesulfonic acid, and methanesulfonic acid. Regarding the temperature when carrying out the Nicholas reaction, for example, it is -78°C to 50°C, specifically 0°C to 25°C. The reaction time of the Nicholas reaction can be appropriately set by those skilled in the art. For example, it is 10 minutes to 24 hours, and more specifically 30 minutes to 2 hours.

[0280] In one aspect, regarding the asymmetric compound among the compounds represented by formula (I), a 1,2,3-triazole ring can be provided as a single positional isomer through a 1,3-dipolar cycloaddition reaction with a bulky azide compound (Example 7). In the conjugation of PDC, regarding the asymmetric compound among the compounds represented by formula (I), a 1,2,3-triazole ring can be provided as a single positional isomer through a 1,3-dipolar cycloaddition reaction with a second functional molecule having a group represented by formula (III), such as a peptide, a nucleic acid, etc. Therefore, the compounds of the present invention are characterized by easy purification during production.

[0281] In one aspect, regarding the symmetric compound among the compounds represented by formula (I), in the 1,3-dipolar cycloaddition reaction of an azide and an alkyne, a 1,2,3-triazole ring can be formed without generating multiple positional isomers. Therefore, the compounds of the present invention are characterized by easy purification during production.

[0282] In one aspect, the azide compound or the second functional molecule having a group represented by formula (III) is not particularly limited. A synthetic compound can be used, and a commercially available compound can be used as a reagent for the 1,3-dipolar cycloaddition reaction of an azide and an alkyne. In addition, a substance obtained by introducing an azide group into the functional molecule can be used.

[0283] Example

[0284] The following shows synthetic examples as examples, and the present invention will be described in more detail. However, the present invention is not limited to these examples.

[0285] In this specification, when compounds, reagents, etc. are represented by symbols, each represents a symbol based on the IUPAC-IUB Commission on Biochemical Nomenclature or a conventional symbol in the art.

[0286] It should be noted that the meanings of the abbreviated expressions in the synthetic examples are as follows.

[0287] tert: tertiary;

[0288] tBu: tert-butyl;

[0289] Ac: acetyl;

[0290] Boc: tert-butoxycarbonyl;

[0291] Fmoc: 9-fluorenylmethoxycarbonyl;

[0292] Cbz: benzyloxycarbonyl;

[0293] TMS: trimethylsilyl;

[0294] TBS: tert-butyldimethylsilyl;

[0295] TIPS: triisopropylsilyl;

[0296] tBuSO 2 : tert-butylsulfonyl;

[0297] nBuLi: n-butyllithium;

[0298] DMF: N,N-dimethylformamide;

[0299] DMSO: dimethyl sulfoxide;

[0300] NMP: N-methylpyrrolidone;

[0301] DIPEA: diisopropylethylamine;

[0302] EtOAc or AcOEt: ethyl acetate;

[0303] Na 2 SO 4 : sodium sulfate;

[0304] DCM: dichloromethane;

[0305] THF: Tetrahydrofuran;

[0306] TBAF: Tetrabutylammonium fluoride;

[0307] BF 3 ·OEt 2 : Boron trifluoride diethyl ether complex;

[0308] Et 2 AlCl: Diethylaluminum chloride;

[0309] EtAlCl 2 : Ethylaluminum dichloride;

[0310] Ms: Methanesulfonyl;

[0311] Ns: Nosyl (nitrobenzenesulfonyl);

[0312] mCPBA: Meta-chloroperoxybenzoic acid;

[0313] MeOH: Methanol;

[0314] CAN: Ammonium cerium(IV) nitrate;

[0315] KOH: Potassium hydroxide;

[0316] LHMDS: Lithium bis(trimethylsilyl)amide;

[0317] CO: Carbon monoxide;

[0318] H 2 O: Water;

[0319] TFA: Trifluoroacetic acid;

[0320] TsOH: p-Toluenesulfonic acid;

[0321] MsOH: Methanesulfonic acid;

[0322] Co 2 (CO) 8 : Dicobalt octacarbonyl;

[0323] HCl: Hydrochloric acid;

[0324] mL: Milliliter (unit);

[0325] M: Mole (unit: mol / L);

[0326] mM: Millimole (unit);

[0327] mm: Millimeter (unit);

[0328] nm: Nanometer (unit);

[0329] μm: Micrometer (unit);

[0330] angstrom (unit);

[0331] min: minute (unit);

[0332] MS: mass spectrometry;

[0333] mmol: millimole (unit);

[0334] g: gram (unit);

[0335] mg: milligram (unit);

[0336] NH silica gel: aminopropyl silica gel.

[0337] In the following synthesis examples, proton nuclear magnetic resonance ( 1 1H-NMR) was measured in deuterated chloroform or deuterated dimethyl sulfoxide solvent using JNM-ECP300 manufactured by JEOL Ltd., JNM-ECX300 manufactured by JEOL Ltd., or AscendTM500 manufactured by Bruker Corporation, and the chemical shift was expressed as the δ value (ppm) with tetramethylsilane as the internal standard (0.0 ppm).

[0338] In the description of the NMR spectrum, "s" means singlet, "d" means doublet, "t" means triplet, "q" means quartet, "dd" means two doublets, "dt" means double triplet, "m" means multiplet, "br" means broad, "J" means coupling constant, "Hz" means hertz, "CDCl 3 " means deuterated chloroform, "DMSO-d 6 " means deuterated dimethyl sulfoxide.

[0339] For high performance liquid chromatography / mass spectrometry, in the absence of special description, any one of ACQUITY UPLC H-Class / QDa manufactured by Waters Corporation, ACQUITY UPLC H-Class / SQD2 manufactured by Waters Corporation, or LC-20AD / Triple Tof5600 manufactured by Shimadzu Corporation was used for measurement.

[0340] In the description of high performance liquid chromatography / mass spectrometry, ESI+ means the positive ion mode of electrospray ionization method, (M+H) + means protonated ion. (M-MeOH+H) + means the ion based on protonation and methanol detachment. (M-C 4 H 8 SO 2 +H) +Refers to an ion based on protonation and detachment of tert-butylsulfonyl group. (M-C 5 H 8 O 2 +H) + Refers to an ion based on protonation and detachment of tert-butoxycarbonyl group. (M-C 4 H 8 +H) + Refers to an ion based on protonation and detachment of tert-butyl group. (M-H 2 O+H) + Refers to an ion based on protonation and detachment of water.

[0341] In the description of high performance liquid chromatography / mass spectrometry, ESI- refers to the negative ion mode of electrospray ionization method, and M-H refers to an ion based on proton detachment.

[0342] Analysis condition A

[0343] Column: Kinetex (registered trademark) 1.7μm EVO C18 2.6μm, 2.1×50mm,

[0344] Mobile phase A: 0.025% TFA in H 2 O

[0345] Mobile phase B: 0.025% TFA in CH 3 CN

[0346] Column temperature: 60°C

[0347] Gradient (% B): 5 - 95% over 2.10 minutes, then 95% from 2.10 minutes to 2.84 minutes, flow rate: 0.6 mL / min

[0348] Detection: UV 254nm.

[0349] [Example 1-1] Synthesis of tert-butyl 3,3-bis(hydroxymethyl)azetidine-1-carboxylate

[0350] [Chemical formula 35]

[0351]

[0352] Dissolve tert-butyl 3-formylazetidine-1-carboxylate (3.88 g, 20.95 mmol, CAS: 177947-96-5) in MeOH (105 mL), add H in which KOH (4.88 g, 87 mmol) is dissolved 2O (50 mL), 37% aqueous formaldehyde solution (17.2 mL, 230 mmol), after stirring at room temperature for 150 minutes, stirred at 40 °C for 24 hours. Methanol was distilled off from the reaction solution, saturated aqueous ammonium chloride solution was added, and extraction was carried out with ethyl acetate. The obtained organic layer was dried over sodium sulfate, filtered, and concentrated under reduced pressure. The obtained residue was purified by flash column chromatography using Isolera (Biotage; Sfaer HC Duo; mobile phase DCM, 3% - 15% MeOH gradient), and the fraction containing the target substance was concentrated under reduced pressure to obtain the title compound as a white powder (3.71 g, 17.08 mmol).

[0353] 1 H-NMR (500 MHz, CDCl 3 ) δ 3.89 (s, 4H), 3.69 (s, 4H), 1.44 (s, 9H).

[0354] [Example 1-2] Synthesis of tert-butyl 3,3-bis({N-[(tert-butoxy)carbonyl]methanesulfonamido}methyl)azetidine-1-carboxylate

[0355] [Chemical formula 36]

[0356]

[0357] tert-Butyl 3,3-bis(hydroxymethyl)azetidine-1-carboxylate (400 mg, 1.84 mmol) obtained in Example 1-1 was dissolved in DCM (9.2 mL), and N-(tert-butoxycarbonyl)methanesulfonamide (755 mg, 3.87 mmol, CAS: 147751-16-4), triphenylphosphine (1062 mg, 4.05 mmol), and diisopropyl azodicarboxylate (2.42 mL, 4.60 mmol, 1.9 M toluene solution, Tokyo Chemical Industry) were added under ice-cooling, and stirred overnight at room temperature. The reaction solution was purified by flash column chromatography using Isolera (Biotage; Sfaer HC Duo; mobile phase n-hexane, 0% - 100% ethyl acetate gradient), and the fraction containing the target substance was concentrated under reduced pressure to obtain the title compound as a colorless viscous substance (740 mg, 1.29 mmol).

[0358] Analysis condition A: retention time = 2.00 minutes; ESI-MS(+) observed value m / z = 572.4 (M + H) + Theoretical value m / z = 571.2.

[0359] [Example 1-3] Synthesis of benzyl 3,3-bis(methanesulfonamido)methyl)azetidine-1-carboxylate

[0360] [Chemical Formula 37]

[0361]

[0362] Dissolve tert-butyl 3,3-bis({N-[(tert-butoxy)carbonyl]methanesulfonamido}methyl)azetidine-1-carboxylate (740 mg, 1.29 mmol) obtained in Examples 1-2 in DCM (6.4 mL), add TFA (6.4 mL), and stir at room temperature for 2 hours. Add toluene to the reaction solution, concentrate, and dry under reduced pressure. Dissolve 400 mg of the 750 mg residue in 1,4-dioxane / water (3.08 mL / 0.34 mL), add sodium carbonate (218 mg, 2.06 mmol) and benzyl chloroformate (140 mg, 0.822 mmol) under ice-cooling, and stir at room temperature for 3 hours. Add saturated aqueous sodium bicarbonate solution to the reaction solution, and extract with DCM / MeOH. Wash the obtained organic layer with saturated brine, dry over sodium sulfate, filter, and concentrate under reduced pressure. Purify the obtained residue by flash column chromatography using Isolera (Biotage; Sfaer HC Duo; mobile phase: hexane, 0% - 100% ethyl acetate gradient, then mobile phase: DCM, 20% MeOH gradient), and concentrate the fraction containing the target substance under reduced pressure to obtain the title compound as a colorless viscous substance (346 mg, 0.700 mmol).

[0363] Analysis condition A: retention time = 1.33 minutes; ESI-MS(+) observed value m / z = 406.3 (M + H) + Theoretical value m / z = 405.1.

[0364] [Examples 1-4] Synthesis of benzyl 3-(methanesulfonamidomethyl)-{[N-(4-methoxybut-2-yn-1-yl)methanesulfonamido]methyl}azetidine-1-carboxylate

[0365] [Chemical Formula 38]

[0366]

[0367] Dissolve 3,3-bis(methanesulfonamidomethyl)azetidine-1-carboxylic acid benzyl ester (190 mg, 0.469 mmol) obtained in Examples 1-3 in a mixed solvent of THF (1.9 mL) and DMF (0.47 mL). Under ice cooling, add sodium hydride (18.7 mg, 0.469 mmol, 60%, dispersed in liquid paraffin), and stir as it is for 10 minutes. Add 1-bromo-4-methoxy-2-butyne (76 mg, 0.469 mmol, CAS: 693-26-5, synthesized by the method described in the literature Journal of Organic Chemistry 2005, 70, 4059-4063) to the reaction solution, and stir under ice cooling for 1 hour. Add water to the reaction solution, and extract with ethyl acetate. After washing the obtained organic layer with saturated brine, dry it over sodium sulfate, filter, and concentrate under reduced pressure. Purify the obtained residue by flash column chromatography using Isolera (Biotage; Sfaer HC Duo; mobile phase: n-hexane, 0% - 70% ethyl acetate gradient), and concentrate the fraction containing the target substance under reduced pressure to obtain the title compound as a colorless viscous substance (70 mg, 0.144 mmol).

[0368] Analysis condition A: retention time = 1.62 minutes; ESI-MS(+) observed value m / z = 488.3 (M+H) + Theoretical value m / z = 487.1.

[0369] [Example 1-5] 3-(methanesulfonamidomethyl)-3-[(N-{[4-(methoxymethyl)-1,2-dicobalt tricyclo[1.1.0.0 2,4 butane-3-yl]methyl}methanesulfonamide)methyl]azetidine-1-carboxylic acid benzyl ester; Synthesis of hexakis(methanidylidyneoxidanium)

[0370] [Chemical formula 39]

[0371]

[0372] Dissolve 3-(methanesulfonamidomethyl)-{[N-(4-methoxybut-2-yn-1-yl)methanesulfonamide]methyl}azetidine-1-carboxylic acid benzyl ester (66 mg, 0.135 mmol) obtained in Examples 1-4 in DCM (0.68 mL), and add Co 2 (CO) 8(51 mg, 0.149 mmol) was stirred at room temperature for 2 hours. The reaction solution was purified by flash column chromatography using Isolera (Biotage; Sfaer HC Duo; mobile phase: n-hexane, 0% - 70% ethyl acetate gradient). The fraction containing the target substance was concentrated under reduced pressure to obtain the title compound as a brown powder (86 mg, 0.111 mmol).

[0373] Analysis condition A: retention time = 2.10 minutes; ESI-MS(+) observed value m / z = 742.2 (M - MeOH + H) + Theoretical value m / z = 773.0.

[0374] [Example 1-6] 3’,7’-Dimethanesulfonyl-3’,7’-diazaspiro[azetidine-3,5’-tetracyclo[7.2.0.0 1,10 .0 9,11 undecane]-1-carboxylic acid benzyl ester; Synthesis of hexakis(methanidylidyneoxidanium)

[0375] [Chemical formula 40]

[0376]

[0377] 3-(Methanesulfonamidomethyl)-3-[(N-{[4-(methoxymethyl)-1,2-diazatricyclo[1.1.0.0 2,4 butan-3-yl]methyl}methanesulfonamide)methyl]azetidine-1-carboxylic acid benzyl ester; hexakis(methanidylidyneoxidanium) (76 mg, 0.098 mmol) obtained in Example 1-5 was dissolved in DCM (9.8 mL), boron trifluoride diethyl ether complex (0.037 mL, 0.295 mmol) was added, and the mixture was stirred at room temperature for 1 hour. A saturated aqueous sodium bicarbonate solution was added to the reaction solution, and the mixture was extracted with ethyl acetate. The obtained organic layer was washed with saturated brine, dried over sodium sulfate, filtered, and concentrated under reduced pressure. The obtained residue was purified by flash column chromatography using Isolera (Biotage; Sfaer HC Duo; mobile phase: n-hexane, 20% - 80% ethyl acetate gradient). The fraction containing the target substance was concentrated under reduced pressure to obtain the title compound as a brown powder (52 mg, 0.070 mmol).

[0378] Analysis condition A: retention time = 2.04 minutes; ESI-MS(+) observed value m / z = 742.2 (M + H) + Theoretical value m / z = 741.0.

[0379] [Examples 1-7] Synthesis of Benzyl 6,11-Dimesyl-2,6,11-triazaspiro[3.8]dodec-8-yne-2-carboxylate

[0380] [Chemical Formula 41]

[0381]

[0382] The 3’,7’-dimesyl-3’,7’-diazaspiro[azetidine-3,5’-tetracyclo[7.2.0.0 1,10 .0 9,11 undecane]-1-carboxylic acid benzyl ester obtained in Examples 1-6; hexakis(methanidylidyneoxidanium) (32 mg, 0.043 mmol) was suspended in diethyl ether (4.3 mL), and WAKOSIL (registered trademark) C-200 (1 g, CAS: 63231-67-4) and CAN (237 mg, 0.432 mmol) were added under ice cooling, followed by stirring at room temperature for 1 hour. The reaction solution was passed through NH silica gel, and the insoluble matter was filtered off. The target substance was eluted with ethyl acetate. The obtained organic layer was concentrated under reduced pressure. The obtained residue was purified by flash column chromatography using Isolera (Biotage; Sfaer HC Duo; mobile phase 100% ethyl acetate), and the fraction containing the target substance was concentrated under reduced pressure to obtain the title compound as a colorless viscous substance (10.1 mg, 0.022 mmol).

[0383] Analysis condition A: retention time = 1.53 minutes; ESI-MS(+) observed value m / z = 456.3 (M+H) + Theoretical value m / z = 455.1.

[0384] 1 H-NMR (500 MHz, CDCl 3 ) δ 7.38 - 7.29 (m, 5H), 5.10 (s, 2H), 4.03 (s, 4H), 3.98 (s, 4H), 3.63 (s, 4H), 2.89 (s, 6H).

[0385] [Example 2-1] Synthesis of Benzyl 3-Hydroxy-3-(3-methoxyprop-1-yn-1-yl)azetidine-1-carboxylate

[0386] [Chemical Formula 42]

[0387]

[0388] Dissolve methyl propargyl ether (2.05 g, 29.2 mmol, CAS: 627-41-8) in THF (122 mL). Add 2 M nBuLi (13.4 mL, 26.8 mmol, cyclohexane solution) at -78 °C and stir for 20 minutes. Then, warm to 0 °C and stir for 15 minutes. After cooling to -78 °C again, add a THF solution of benzyl 3-oxoazetidine-1-carboxylate (5.00 g, 24.36 mmol, CAS: 105258-93-3) and stir at -78 °C for 1 hour. Then, warm to 0 °C, stir for 30 minutes, stop the reaction with saturated brine, extract the aqueous layer with ethyl acetate, wash the organic layer with saturated brine, dry over sodium sulfate, filter, and concentrate under reduced pressure. Purify the resulting residue by flash column chromatography using Isolera (Biotage; Sfaer HCDuo; mobile phase: n-hexane, 10% - 70% ethyl acetate gradient) to obtain the title compound as a colorless viscous substance (5.30 g, 19.3 mmol).

[0389] Analysis condition A: retention time = 1.54 minutes; ESI-MS(+) observed m / z = 276.2 (M+H) + Theoretical m / z = 275.1.

[0390] [Example 2-2] Synthesis of benzyl 3-(3-hydroxypropoxy)-3-(3-methoxyprop-1-yn-1-yl)azetidine-1-carboxylate

[0391] [Chemical formula 43]

[0392]

[0393] Dissolve benzyl 3-hydroxy-3-(3-methoxyprop-1-yn-1-yl)azetidine-1-carboxylate (3.00 g, 10.9 mmol) obtained in Example 2-1 in THF (18 mL). Add sodium hydride (0.654 g, 16.4 mmol, 60%, dispersed in liquid paraffin) at 0 °C and stir for 20 minutes. Then, add a DMF solution (18 mL) of (3-bromopropoxy)(tert-butyl)dimethylsilane (4.14 g, 16.4 mmol, CAS: 89031-84-5) at 0 °C and stir overnight at room temperature. Then, stop the reaction with saturated aqueous ammonium chloride, extract the aqueous layer with ethyl acetate, wash the organic layer with saturated brine, dry over sodium sulfate, filter, and concentrate under reduced pressure. Dissolve the resulting residue in THF / water (9 / 1), then add TBAF·3H 2O (10.28 g, 32.7 mmol) was stirred overnight. The reaction solution was dissolved in ethyl acetate, the aqueous layer was extracted with ethyl acetate, the organic layer was washed with saturated brine, dried over sodium sulfate, filtered, and concentrated under reduced pressure. The obtained residue was purified by flash column chromatography using Isolera (Biotage; Sfaer HC Duo; mobile phase: dichloromethane, 1% - 40% methanol gradient) to obtain the title compound as a pale yellow viscous substance (3.00 g, 9.00 mmol).

[0394] Analysis condition A: retention time = 1.54 minutes; ESI-MS(+) observed m / z = 334.4 (M + H) + Theoretical m / z = 333.2

[0395] [Example 2 - 3] 3-(3-Hydroxypropoxy)-3-[4-(methoxymethyl)-1,2-dicobalt tricyclo[1.1.0.0 2,4 butan-3-yl]azetidine-1-carboxylic acid benzyl ester; Synthesis of hexakis(methanidylidyneoxidanium)

[0396] [Chemical formula 44]

[0397]

[0398] Dissolve 3-(3-Hydroxypropoxy)-3-(3-methoxyprop-1-yn-1-yl)azetidine-1-carboxylic acid benzyl ester (0.30 g, 0.90 mmol) obtained in Example 2 - 2 in dichloromethane (9 mL), add Co 2 (CO) 8 (0.40 g, 1.17 mmol) at room temperature and stir for 2 hours. After the reaction is completed, concentrate under reduced pressure. The obtained residue was purified by flash column chromatography using Isolera (Biotage; Sfaer HC Duo; mobile phase: n-hexane, 20% - 70% ethyl acetate gradient) to obtain the title compound as a brown viscous substance (0.49 g, 0.79 mmol).

[0399] Analysis condition A: retention time = 2.16 minutes; ESI-MS(+) observed m / z = 588.2 (M - MeOH + H) + Theoretical m / z = 619.0

[0400] [Example 2 - 4] 3’,7’-Dioxa-10’,11’-dicobalt spiro[azetidine-3,2’-tetracyclo[7.2.0.0 1 ,10 .09,11 Benzyl undecane-1-carboxylate; Synthesis of hexakis(methanidylidyneoxidanium)

[0401] [Chemical formula 45]

[0402]

[0403] Dissolve 3-(3-hydroxypropoxy)-3-[4-(methoxymethyl)-1,2-dicobalt tricyclo[1.1.0.0 2,4 butane-3-yl]azetidine-1-carboxylate benzyl ester; hexakis(methanidylidyneoxidanium) (0.46 g, 0.74 mmol) in dichloromethane (149 mL). Add boron trifluoride diethyl ether complex (0.14 mL, 1.11 mmol) at room temperature and stir for 30 minutes. Stop the reaction by adding saturated aqueous sodium bicarbonate solution to the reaction mixture. Extract the aqueous layer with dichloromethane, wash the organic layer with saturated brine, dry over sodium sulfate, filter, and concentrate under reduced pressure. Purify the resulting residue by flash column chromatography using Isolera (Biotage; SfaerHC Duo; mobile phase: n-hexane, 10% - 40% ethyl acetate gradient) to obtain the title compound as a brown viscous substance (0.174 g, 0.296 mmol).

[0404] Analysis condition A: Retention time = 2.25 minutes; ESI-MS(+) observed m / z = 588.2 (M + H) + Theoretical m / z = 587.0

[0405] [Example 2-5] Synthesis of benzyl 5,9-dioxa-2-azaspiro[3.8]dodec-11-yne-2-carboxylate

[0406] [Chemical formula 46]

[0407]

[0408] Dissolve 3’,7’-dioxa-10’,11’-dicobalt spiro[azetidine-3,2’-tetracyclo[7.2.0.0 1,10 .0 9,11 undecane-1-carboxylate benzyl ester; hexakis(methanidylidyneoxidanium) (0.027 g, 0.046 mmol) in a mixed solvent of (acetone / water = 15 / 1) (1.5 mL). Add TBAF·3H 2O (0.218 g, 0.690 mmol). After stirring for 2 hours at room temperature, water was added to the reaction solution. The aqueous layer was extracted with ethyl acetate. The obtained organic layer was washed with saturated brine, dried over sodium sulfate, filtered, and concentrated under reduced pressure. The obtained residue was purified by flash column chromatography using Isolera (Biotage; Sfaer HC Duo; mobile phase: n-hexane, 15% - 50% ethyl acetate gradient) to obtain the title compound as a colorless viscous substance (0.011 g, 0.036 mmol).

[0409] Analysis condition A: retention time = 1.75 minutes; ESI-MS(+) observed value m / z = 302.2 (M + H) + Theoretical value m / z = 301.1.

[0410] 1 H-NMR (500 MHz, CDCl 3 ) δ 7.40 - 7.28 (m, 5H), 5.10 (s, 2H), 4.15 (s, 2H), 4.15 - 4.06 (m, 4H), 3.88 - 3.80 (m, 4H), 1.76 - 1.68 (m, 2H).

[0411] [Example 3 - 1] Synthesis of Benzyl 3-(3 - methoxyprop - 1 - yn - 1 - yl)-3-[(2 - methylpropan - 2 - ylsulfinyl)amino]azetidine - 1 - carboxylate

[0412] [Chemical Formula 47]

[0413]

[0414] Dissolve methyl propargyl ether (0.59 g, 8.43 mmol, CAS: 627-41-8) in THF (65 mL), add 2 M nBuLi (3.9 mL, 7.78 mmol, cyclohexane solution) at -78 °C, warm to 0 °C and stir for 10 minutes. Add a THF solution of benzyl 3-[(2-methylpropan-2-sulfinyl)imino]azetidine-1-carboxylate (2.00 g, 6.49 mmol, CAS: 1638764-74-5, synthesized by the method described in US2020 / 0038378A1) to the reaction solution, stir at 0 °C for 20 minutes, and stop the reaction with saturated aqueous ammonium chloride. After extracting the aqueous layer with ethyl acetate, wash the obtained organic layer with saturated brine, dry over sodium sulfate, filter, and concentrate under reduced pressure. Purify the obtained residue by flash column chromatography using Isolera (Biotage; Sfaer HC Duo; mobile phase: n-hexane, 50% - 100% ethyl acetate gradient) to obtain the title compound as a yellow viscous substance (0.96 g, 2.54 mmol).

[0415] Analysis condition A: retention time = 1.72 minutes; ESI-MS(+) observed m / z = 379.3 (M + H) + Theoretical m / z = 378.2.

[0416] [Example 3-2] Synthesis of benzyl 3-({3-[(tert-butyldimethylsilyl)oxy]propyl}(2-methylpropan-2-sulfinyl)amino)-3-(3-methoxyprop-1-yn-1-yl)azetidine-1-carboxylate

[0417] [Chemical formula 48]

[0418]

[0419] Sodium hydride (42.3 mg, 60%, dispersed in liquid paraffin) was suspended in THF (2.6 mL) at 0 °C, and then 3-((tert-butyl(dimethyl)silyl)oxy)propyl 2-methylpropane-2-sulfinate (0.200 g, 0.528 mmol) obtained in Example 3-1 was added at 0 °C and stirred for 10 minutes. A DMF solution (2.6 mL) of (3-bromopropoxy)(tert-butyl)dimethylsilane (0.294 g, 1.16 mmol, CAS: 89031-84-5) was added to the reaction solution and stirred overnight at room temperature. After the reaction solution was cooled to 0 °C, the reaction was stopped with a saturated aqueous ammonium chloride solution. The aqueous layer was extracted with ethyl acetate, and the obtained organic layer was washed with saturated brine, dried over sodium sulfate, filtered, and concentrated under reduced pressure. The obtained residue was purified by flash column chromatography using Isolera (Biotage; Sfaer HC Duo; mobile phase: n-hexane, 20% - 100% ethyl acetate gradient) to obtain the title compound as a colorless viscous substance (0.080 g, 0.145 mmol).

[0420] Analysis condition A: retention time = 2.37 minutes; ESI-MS(+) observed m / z = 551.4 (M + H) + Theoretical m / z = 550.3.

[0421] [Example 3-3] Synthesis of Benzyl 3-(N-{3-[(tert-butyldimethylsilyl)oxy]propyl}-2-methylpropane-2-sulfonamido)-3-(3-methoxyprop-1-yn-1-yl)azetidine-1-carboxylate

[0422] [Chemical formula 49]

[0423]

[0424] 3-({3-[(tert-Butyldimethylsilyl)oxy]propyl}(2-methylpropan-2-sulfinyl)amino)-3-(3-methoxyprop-1-yn-1-yl)azetidine-1-carboxylic acid benzyl ester (0.080 g, 0.145 mmol) obtained in Example 3-2 was dissolved in dichloromethane (1.5 mL), and 30% aqueous mCPBA (54 mg, 0.218 mmol, CAS: 937-14-4) was added at room temperature and stirred for 2 hours. After the reaction solution was cooled to 0 °C, the reaction was stopped with saturated aqueous sodium bicarbonate, the aqueous layer was extracted with ethyl acetate, the obtained organic layer was washed with saturated brine, dried over sodium sulfate, filtered, and concentrated under reduced pressure. The obtained residue was purified by flash column chromatography using Isolera (Biotage; Sfaer HC Duo; mobile phase: hexane, 20% - 50% ethyl acetate gradient) to obtain the title compound as a colorless viscous substance (0.068 g, 0.12 mmol).

[0425] Analysis condition A: retention time = 2.45 minutes; ESI-MS(+) observed value m / z = 447.4 (M - C 4 H 8 SO 2 +H) + Theoretical value m / z = 566.3.

[0426] [Example 3-4] Synthesis of Benzyl 3-[N-(3-hydroxypropyl)-2-methylpropan-2-sulfonamide]-3-(3-methoxyprop-1-yn-1-yl)azetidine-1-carboxylate

[0427] [Chemical formula 50]

[0428]

[0429] Benzyl 3-(N-{3-[(tert-Butyldimethylsilyl)oxy]propyl}-2-methylpropan-2-sulfonamide)-3-(3-methoxyprop-1-yn-1-yl)azetidine-1-carboxylate (0.068 g, 0.12 mmol) obtained in Example 3-3 was dissolved in THF (1.5 mL), and 1 M TBAF (0.18 mL, 0.18 mmol, THF solution) was added at room temperature and stirred for 3 hours. The reaction solution was concentrated under reduced pressure, and the obtained residue was purified by flash column chromatography using Isolera (Biotage; Sfaer HC Duo; mobile phase: hexane, 30% - 100% ethyl acetate gradient) to obtain the title compound as a colorless viscous substance (0.053 g, 0.117 mmol).

[0430] Analysis condition A: retention time = 1.71 minutes; ESI-MS(+) observed value m / z = 333.3 (M - C 4 H 8 SO 2 +H) + Theoretical value m / z = 452.2.

[0431] [Example 3-5] 3'-(2-methylpropan-2-sulfonyl)-7'-oxa-3'-aza-10',11'-dicobalt spiro[azetidine-3,2'-tetracyclo[7.2.0.0 1,10 .0 9,11 undecane]-1-carboxylic acid benzyl ester; Synthesis of hexakis(methanidylidyneoxidanium)

[0432] [Chemical formula 51]

[0433]

[0434] Dissolve 3-[N-(3-hydroxypropyl)-2-methylpropan-2-sulfonamide]-3-(3-methoxyprop-1-yn-1-yl)azetidine-1-carboxylic acid benzyl ester (0.053 g, 0.117 mmol) obtained in Example 3-4 in dichloromethane (1.1 mL), and add Co 2 (CO) 8 (0.060 g, 0.176 mmol) at room temperature and stir for 3 hours. After the reaction is completed, concentrate under reduced pressure. Purify the obtained residue by flash column chromatography using Isolera (Biotage; Sfaer HC Duo; mobile phase: n-hexane, 30% - 80% ethyl acetate gradient) to obtain 3-[N-(3-hydroxypropyl)-2-methylpropan-2-sulfonamide]-3-[4-(methoxymethyl)-1,2-dicobalt tricyclo[1.1.0.0 2,4 butan-3-yl]azetidine-1-carboxylic acid benzyl ester; hexakis(methanidylidyneoxidanium) (0.082 g, 0.111 mmol). The obtained 3-[N-(3-hydroxypropyl)-2-methylpropan-2-sulfonamide]-3-[4-(methoxymethyl)-1,2-dicobalt tricyclo[1.1.0.0 2 ,4Benzyl [azetidin-1-yl]butane-3-carboxylate; hexakis(methanidylidyneoxidanium) (0.020 g, 0.027 mmol) was dissolved in dichloromethane (5 mL), boron trifluoride diethyl ether complex (0.005 mL, 0.041 mmol) was added at room temperature and stirred for 15 minutes. The reaction was stopped by adding saturated aqueous sodium bicarbonate solution, the aqueous layer was extracted with dichloromethane, the organic layer was washed with saturated brine, dried over sodium sulfate, filtered, and concentrated under reduced pressure. The resulting residue was purified by flash column chromatography using Isolera (Biotage; Sfaer HC Duo; mobile phase: n-hexane, 20% - 50% ethyl acetate gradient) to obtain the title compound as a brown viscous substance (0.013 g, 0.027 mmol).

[0435] Analysis condition A: retention time = 2.29 minutes; ESI-MS(+) observed m / z = 707.1 (M+H) + Theoretical m / z = 706.0

[0436] [Example 3-6] Synthesis of benzyl 5-(2-methylpropan-2-sulfonyl)-9-oxa-2,5-diazaspiro[3.8]dodec-11-yne-2-carboxylate

[0437] [Chemical formula 52]

[0438]

[0439] The 3'-(2-methylpropan-2-sulfonyl)-7'-oxa-3'-aza-10',11'-dico-baltaspiro[azetidine-3,2'-tetracyclo[7.2.0.0 1,10 .0 9,11 undecane]-1-carboxylate benzyl ester obtained in Example 3-5; hexakis(methanidylidyneoxidanium) (0.013 g, 0.018 mmol) was dissolved in a mixed solvent of (acetone / water = 15 / 1) (0.36 mL), and TBAF·3H 2 O (0.115 g, 0.365 mmol) was added at room temperature. After stirring at room temperature for 3 hours, water was added to the reaction solution, the aqueous layer was extracted with ethyl acetate, the resulting organic layer was washed with saturated brine, dried over sodium sulfate, filtered, and concentrated under reduced pressure. The resulting residue was purified by flash column chromatography using Isolera (Biotage; Sfaer HC Duo; mobile phase: n-hexane, 20% - 70% ethyl acetate gradient) to obtain the title compound as a white powder (0.005 g, 0.012 mmol).

[0440] Analysis of Condition A: Retention time = 1.84 minutes; Observed value of ESI-MS(+) m / z = 421.3 (M+H) + Theoretical value of m / z = 420.2.

[0441] 1 H-NMR(500MHz,CDCl 3 ) δ 7.39 - 7.30 (m, 5H), 5.11 (s, 2H), 4.52 - 4.46 (m, 2H), 4.27 - 4.19 (m, 4H), 3.90 - 3.84 (m, 2H), 3.53 - 3.45 (m, 2H), 2.11 - 2.03 (m, 2H), 1.41 (s, 9H).

[0442] [Example 4-1] 3-(3-Methanesulfonamidopropoxy)-3-[4-(methoxymethyl)-1,2-dicobalt tricyclo[1.1.0.0 2,4 butan-3-yl]azetidine-1-carboxylic acid benzyl ester; Synthesis of hexakis(methanidylidyneoxidanium)

[0443] [Chemical Formula 53]

[0444]

[0445] Dissolve 3-(3-Hydroxypropoxy)-3-(3-methoxyprop-1-yn-1-yl)azetidine-1-carboxylic acid benzyl ester (0.12 g, 0.360 mmol) obtained in Example 2-2 in toluene (1.5 mL), add MsNH 2 (0.103 g, 1.08 mmol), cyanomethylenetributylphosphine (0.261 g, 1.08 mmol CAS: 157141-27-0), and stir overnight at room temperature. After the reaction is completed, concentrate under reduced pressure, and purify the resulting residue by flash column chromatography using Isolera (Biotage; Sfaer HC Duo; mobile phase: n-hexane, 40% - 80% ethyl acetate gradient) to obtain a red viscous intermediate. Dissolve the above intermediate in dichloromethane (1.8 mL), and add Co 2 (CO) 8(0.148 g, 0.433 mmol) and stirred for 30 minutes. After completion of the reaction, it was concentrated under reduced pressure. The resulting residue was purified by flash column chromatography using Isolera (Biotage; Sfaer HC Duo; mobile phase: n-hexane, 30% - 50% ethyl acetate gradient) to obtain the title compound as a brown viscous substance (0.196 g, 0.281 mmol).

[0446] Analysis condition A: retention time = 2.27 minutes; ESI-MS(+) observed m / z = 697.1 (M+H) + Theoretical m / z = 696.0.

[0447] [Example 4-2] 7'-Methanesulfonyl-3'-oxa-7'-aza-10',11'-dicobalt spiro[azetidine-3,2'-tetracyclo[7.2.0.0 1,10 .0 9,11 undecane]-1-carboxylic acid benzyl ester; Synthesis of hexakis(methanidylidyneoxidanium)

[0448] [Chemical formula 54]

[0449]

[0450] 3-(3-Methanesulfonamidopropoxy)-3-[4-(methoxymethyl)-1,2-dicobalt tricyclo[1.1.0.0 2,4 butan-3-yl]azetidine-1-carboxylic acid benzyl ester; hexakis(methanidylidyneoxidanium) (0.100 g, 0.144 mmol) obtained in Example 4-1 was dissolved in dichloromethane (57 mL), boron trifluoride diethyl ether complex (0.152 mL, 1.21 mmol) was added at 0 °C and stirred for 1 hour. The reaction was stopped by adding saturated aqueous sodium bicarbonate solution, the aqueous layer was extracted with dichloromethane, the organic layer was washed with saturated brine, dried over sodium sulfate, filtered, and concentrated under reduced pressure. The resulting residue was purified by flash column chromatography using Isolera (Biotage; Sfaer HC Duo; mobile phase: n-hexane, 20% - 60% ethyl acetate gradient) to obtain the title compound as a reddish-brown viscous substance (0.093 g, 0.140 mmol).

[0451] Analysis condition A: retention time = 2.19 minutes; ESI-MS(+) observed m / z = 665.1 (M+H) + Theoretical m / z = 664.0.

[0452] [Example 4-3] Synthesis of Benzyl 9-Methanesulfonyl-5-oxa-2,9-diazaspiro[3.8]dodec-11-yne-2-carboxylate

[0453] [Chemical Formula 55]

[0454]

[0455] Dissolve benzyl 7'-methanesulfonyl-3'-oxa-7'-aza-10',11'-dicozaspiro[azetidine-3,2'-tetracyclo[7.2.0.0 1,10 .0 9,11 undecane]-1-carboxylate obtained in Example 4-2 (0.200 g, 0.301 mmol) in diethyl ether (30 mL), add WAKOSIL (registered trademark) C-200 (5 g), and then add CAN (1.65 g, 3.01 mmol) at 0 °C. After stirring at 0 °C for 1 hour, the temperature is raised to room temperature and stirred for 1 hour. After the reaction is completed, the reaction solution is passed through NH silica gel, and the insoluble matter is filtered out, and then the filtrate is concentrated under reduced pressure. The obtained residue is purified by flash column chromatography using Isolera (Biotage; Sfaer HC Duo;; mobile phase: hexane, 30% - 80% ethyl acetate gradient, then mobile phase: dichloromethane, 1% - 20% methanol gradient) to obtain the title compound as a pale yellow solid (0.010 g, 0.036 mmol).

[0456] Analysis condition A: retention time = 1.71 minutes; ESI-MS(+) observed value m / z = 379.3 (M + H) + Theoretical value m / z = 378.1.

[0457] 1 1H-NMR (500 MHz, CDCl 3 ) δ 7.40 - 7.29 (m, 5H), 5.10 (s, 2H), 4.14 - 4.03 (m, 4H), 3.96 (s, 2H), 3.91 - 3.84 (m, 2H), 3.46 - 3.38 (m, 2H), 2.84 (s, 3H), 1.95 - 1.84 (m, 2H).

[0458] [Example 5-1] Synthesis of (2S)-6-(2-Bromo-2-methylpropanamide)-2-{[(tert-butoxy)carbonyl]amino}hexanoic Acid

[0459] [Chemical Formula 56]

[0460]

[0461] N-Boc-L-lysine (1.13 g, 4.57 mmol, CAS: 13734-28-6) was suspended in THF (11.6 mL), N-methyl-N-(trimethylsilyl)acetamide (1.53 mL, 9.57 mmol) was added, and the mixture was stirred at 50 °C for 1 h. The reaction solution was ice-cooled at 0 °C, DIPEA (0.89 mL, 5.22 mmol) and 2-bromo-2-methylpropanoyl bromide (1.00 g, 4.35 mmol, CAS: 20769-85-1) were added, and the mixture was stirred for 1 h. The reaction was stopped with 10% aqueous sodium carbonate solution, and the aqueous layer was washed twice with heptane. The aqueous layer was acidified by adding 1 M aqueous hydrochloric acid solution and extracted with ethyl acetate. The obtained organic layer was washed with saturated brine, dried over sodium sulfate, filtered, and concentrated under reduced pressure to obtain the title compound as a white powder (1.64 g, 4.15 mmol).

[0462] Analysis condition A: retention time = 1.62 min; ESI-MS(+) observed value m / z = 295.1 (M - C 5 H 8 SO 2 + H) + Theoretical value m / z = 394.1.

[0463] [Example 5-2] Synthesis of (2S)-6-(2-azido-2-methylpropanamido)-2-{[(tert-butoxy)carbonyl]amino}hexanoic acid

[0464] [Chemical formula 57]

[0465]

[0466] (2S)-6-(2-Bromo-2-methylpropanamido)-2-{[(tert-butoxy)carbonyl]amino}hexanoic acid (200 mg, 0.506 mmol) obtained in Example 5-1 was dissolved in DMSO (1.7 mL), sodium azide (66 mg, 1.01 mmol) was added, and the mixture was stirred at 50 °C for 3 h. Water was added to the reaction solution, and the mixture was extracted with ethyl acetate. The obtained organic layer was washed with 1 N aqueous hydrochloric acid solution and saturated brine, dried over sodium sulfate, filtered, and concentrated under reduced pressure to obtain the title compound as a colorless viscous substance (146 mg, 0.408 mmol).

[0467] 1 H-NMR (500 MHz, CDCl 3)δ 6.62 - 6.56 (m, 1H), 5.23 (d, J = 7.5 Hz, 1H), 4.30 - 4.22 (m, 1H), 3.32 - 3.17 (m, 2H), 1.95 - 1.85 (m, 1H), 1.77 - 1.69 (m, 1H), 1.58 - 1.55 (m, 1H), 1.53 (s, 6H), 1.45 (s, 9H), 1.44 - 1.32 (m, 3H)

[0468] [Example 5 - 3] Synthesis of (2S)-6-(2 - azido - 2 - methylpropanamide)-2 - {[(9H - fluoren - 9 - yl)methoxy]carbonyl}aminohexanoic acid

[0469] [Chemical Formula 58]

[0470]

[0471] (2S)-6-(2 - azido - 2 - methylpropanamide)-2 - {[(tert - butoxy)carbonyl]amino}hexanoic acid (146 mg, 0.408 mmol) obtained in Example 5 - 2 was cooled in ice, TFA (1.36 mL) was added, and the mixture was stirred as it was for 2 hours. Toluene was added to the reaction solution, concentrated, and dried under reduced pressure. The residue was dissolved in acetone / water (1.0 mL / 1.0 mL), sodium bicarbonate (274 mg, 3.26 mmol) and N - [(9H - fluoren - 9 - ylmethoxy)carbonyloxy]succinimide (151 mg, 0.448 mmol, CAS: 82911 - 69 - 1) were added, and the mixture was stirred overnight at room temperature. Ethyl acetate was added to the reaction solution for extraction. The obtained organic layer was washed with saturated brine, dried over sodium sulfate, filtered, and concentrated under reduced pressure. The obtained residue was purified by flash column chromatography using Isolera (Biotage; Sfaer HC Duo; mobile phase 100% ethyl acetate, then mobile phase DCM, 1% - 20% methanol gradient), and the fraction containing the target substance was concentrated under reduced pressure to obtain the title compound as a white powder (148 mg, 0.309 mmol).

[0472] Analysis condition A: Retention time = 1.68 minutes; ESI - MS(+) observed value m / z = 480.3 (M + H) + Theoretical value m / z = 479.2.

[0473] 1 H - NMR(500 MHz, CDCl 3)δ 7.77 - 7.73 (m, 2H), 7.62 - 7.52 (m, 2H), 7.42 - 7.37 (m, 2H), 7.32 - 7.26 (m, 2H), 6.68 - 6.52 (m, 1H), 5.73 - 5.62 (m, 1H), 4.52 - 4.15 (m, 4H), 3.32 - 3.17 (m, 2H), 1.98 - 1.87 (m, 1H), 1.83 - 1.72 (m, 1H), 1.58 - 1.47 (m, 7H), 1.46 - 1.28 (m, 3H).

[0474] [Example 6] Synthesis of (2S)-6-{1'-[(Benzyloxy)carbonyl]-5,9-dimesyl-4,5,6,8,9,10-hexahydro-1H-spiro[[1,2,3]triazolo[4,5-g][1,5]diazocine-7,3'-azetidine]-1-yl}-2-({[(9H-fluoren-9-yl)methoxy]carbonyl}amino)hexanoic acid

[0475] [Chemical Formula 59]

[0476]

[0477] Dissolve benzyl 6,11-dimesyl-2,6,11-triazaspiro[3.8]dodec-8-yne-2-carboxylate (13.5 mg, 0.030 mmol) obtained in Examples 1 - 7 in DMSO (0.59 mL), add (2S)-6-azido-2-({[(9H-fluoren-9-yl)methoxy]carbonyl}amino)hexanoic acid (12.9 mg, 0.033 mmol, CAS: 159610-89-6) and stir at room temperature for 30 hours. Add water to the reaction solution and extract with ethyl acetate. Wash the obtained organic layer with saturated brine, dry over sodium sulfate, filter, and concentrate under reduced pressure. Purify the obtained residue by flash column chromatography using Isolera (Biotage; Sfaer HC Duo; mobile phase 100% ethyl acetate, then mobile phase DCM, 0% - 30% methanol gradient), and concentrate the fraction containing the target substance under reduced pressure to obtain the title compound as a white powder (21 mg, 0.025 mmol).

[0478] Analysis condition A: retention time = 1.90 minutes; ESI-MS(+) observed m / z = 850.6 (M + H) + Theoretical m / z = 849.3.

[0479] 1 H-NMR(500MHz, DMSO-d 6)δ 12.83 - 12.34 (m, 1H), 7.91 - 7.87 (m, 2H), 7.73 - 7.67 (m, 2H), 7.44 - 7.39 (m, 2H), 7.38 - 7.28 (m, 7H), 5.01 (s, 2H), 4.63 - 4.53 (m, 4H), 4.36 - 4.19 (m, 5H), 3.79 - 3.58 (m, 7H), 3.26 - 3.24 (m, 2H), 3.18 (s, 3H), 3.04 (s, 3H), 1.82 - 1.66 (m, 3H), 1.66 - 1.56 (m, 1H), 1.35 - 1.21 (m, 2H).

[0480] The results show that benzyl 6,11 - dimethanesulfonyl - 2,6,11 - triazaspiro[3.8]dodec - 8 - yne - 2 - carboxylate, which is a compound of the present invention, undergoes a click reaction with an azide compound in the same manner as the alkynes used in known click chemistry.

[0481] [Example 7] Synthesis of (2S) - 6 - (2 - {1’ - [(benzyloxy)carbonyl] - 6,7,8,10 - tetrahydro - 1H - spiro[[1,5]dioxonino(dioxonino)[7,8 - d][1,2,3]triazol - 4,3’ - azetidine] - 1 - yl} - 2 - methylpropanamide) - 2 - ({[(9H - fluoren - 9 - yl)methoxy]carbonyl}amino)hexanoic acid

[0482] [Chemical Formula 60]

[0483]

[0484] Dissolve benzyl 5,9 - dioxo - 2 - azaspiro[3.8]dodec - 11 - yne - 2 - carboxylate (13.0 mg, 0.043 mmol) obtained in Examples 2 - 5 in DMSO (0.78 mL), add (2S) - 6 - (2 - azido - 2 - methylpropanamide) - 2 - ({[(9H - fluoren - 9 - yl)methoxy]carbonyl}amino)hexanoic acid (18.8 mg, 0.033 mmol) obtained in Example 5 - 3, and stir at room temperature for 18 hours. Add water to the reaction solution, and extract with ethyl acetate / Et 2 O (3 / 1). Wash the obtained organic layer with saturated brine, dry over sodium sulfate, filter, and concentrate under reduced pressure. Purify the obtained residue by flash column chromatography using Isolera (Biotage; Sfaer HC Duo; mobile phase DCM, 0% - 30% methanol gradient), and concentrate the fraction containing the target substance under reduced pressure to obtain the title compound as a white powder (23.2 mg, 0.030 mmol).

[0485] Analysis condition A: retention time = 2.04 minutes; ESI-MS(+) observed value m / z = 781.6 (M+H) + Theoretical value m / z = 780.4.

[0486] 1 H-NMR(500MHz, DMSO-d 6 ) δ 12.57 - 12.52 (m, 1H), 7.91 - 7.85 (m, 3H), 7.74 - 7.70 (m, 2H), 7.61 - 7.58 (m, 1H), 7.44 - 7.39 (m, 2H), 7.38 - 7.30 (m, 7H), 5.08 (s, 2H), 4.50 (s, 2H), 4.44 - 4.29 (m, 2H), 4.29 - 4.07 (m, 5H), 3.93 - 3.85 (m, 1H), 3.75 - 3.70 (m, 2H), 3.38 - 3.33 (m, 2H), 3.06 - 2.99 (m, 2H), 1.79 (s, 6H), 1.71 - 1.52 (m, 4H), 1.43 - 1.21 (m, 4H).

[0487] The present results show that, for benzyl 5,9-dioxa-2-azaspiro[3.8]dodec-11-yne-2-carboxylate, which is a compound of the present invention, it undergoes a click reaction with an azide compound in the same manner as the alkynes used in known click chemistry.

[0488] [Example 8-1] Synthesis of tert-butyl 3-hydroxy-3-(3-methoxyprop-1-yn-1-yl)azetidine-1-carboxylate

[0489] [Chemical formula 61]

[0490]

[0491] Dissolve methyl propargyl ether (4.26 g, 60.7 mmol, CAS: 627 - 41 - 8) in THF (156 mL). Add 2 M nBuLi (28.0 mL, 56.1 mmol, cyclohexane solution) at -78 °C and stir for 10 minutes. Then, warm to 0 °C and stir for 10 minutes. After cooling to -78 °C again, add a THF solution of tert-butyl 3-oxoazetidine-1-carboxylate (8.00 g, 46.7 mmol, CAS: 398489 - 26 - 4) and stir at -78 °C for 20 minutes. Then, warm to 0 °C and stir for 30 minutes. After that, stop the reaction with saturated ammonium chloride solution at -78 °C. Extract the aqueous layer with ethyl acetate, wash the organic layer with saturated brine, dry over sodium sulfate, filter, and concentrate under reduced pressure. Purify the obtained residue by flash column chromatography using Isolera (Biotage; Sfaer HC Duo; mobile phase: n-hexane, 20% - 50% ethyl acetate gradient) to obtain the title compound as a pale yellow viscous substance (11.28 g, 45.7 mmol).

[0492] Analysis condition A: retention time = 1.35 minutes; ESI-MS(+) observed m / z = 242.2 (M + H) + Theoretical m / z = 241.1.

[0493] 1 H-NMR(500MHz,CDCl 3 ) δ 4.21 - 4.17 (m, 2H), 4.15 (s, 2H), 4.05 - 4.01 (m, 2H), 3.39 (s, 3H), 2.77 (s, 1H), 1.44 (s, 9H).

[0494] [Example 8 - 2] Synthesis of tert-butyl 3-(3-hydroxypropoxy)-3-(3-methoxyprop-1-yn-1-yl)azetidine-1-carboxylate

[0495] [Chemical formula 62]

[0496]

[0497] Sodium hydride (2.56 g, 64.0 mmol, 60%, dispersed in liquid paraffin) was added to DMF (76 mL), and a solution of tert-butyl 3-hydroxy-3-(3-methoxyprop-1-yn-1-yl)azetidine-1-carboxylate (11.03 g, 45.7 mmol) obtained in Example 8-1 in THF (76 mL) was added at 0 °C and stirred for 20 minutes. Then, a solution of (3-bromopropoxy)(tert-butyl)dimethylsilane (17.37 g, 68.6 mmol, CAS: 89031-84-5) in DMF was added at 0 °C, and the mixture was stirred at room temperature for 6 hours. Then, the reaction was stopped with a saturated aqueous ammonium chloride solution, the aqueous layer was extracted with ethyl acetate, the organic layer was washed with saturated brine, dried over sodium sulfate, filtered, and concentrated under reduced pressure. The obtained residue was purified by flash column chromatography using Isolera (Biotage; Sfaer HC Duo; mobile phase: n-hexane, 0% - 30% ethyl acetate gradient) to obtain an intermediate as a colorless transparent viscous substance. The previously obtained intermediate was added to a solution of THF (147 mL), 1M TBAF (66.1 mL, 66.1 mmol) was added at 0 °C, and the mixture was stirred at room temperature for 3 hours. After completion of the reaction, the reaction was stopped with a saturated aqueous ammonium chloride solution, the aqueous layer was extracted with ethyl acetate, the organic layer was washed with saturated brine, dried over sodium sulfate, filtered, and concentrated under reduced pressure. The obtained residue was purified by flash column chromatography using Isolera (Biotage; Sfaer HC Duo; mobile phase: n-heptane, 0% - 80% ethyl acetate gradient) to obtain the title compound as a colorless transparent viscous substance (13.05 g, 43.6 mmol).

[0498] Analysis condition A: retention time = 1.51 minutes; ESI-MS(+) observed value m / z = 300.3 (M + H) + Theoretical value m / z = 299.4.

[0499] [Example 8-3] N-{3-[(1-{2-[(tert-Butyldiphenylsilyl)oxy]acetyl}-3-[4-(methoxymethyl)-1,2-dicobaltatricyclo[1.1.0.0 2,4 butan-3-yl]azetidin-3-yl)oxy]propyl}methanesulfonamide; Synthesis of hexakis(methanidylidyneoxidanium)

[0500] [Chemical formula 63]

[0501]

[0502] Dissolve tert-butyl 3-(3-hydroxypropoxy)-3-(3-methoxyprop-1-yn-1-yl)azetidine-1-carboxylate (3.00 g, 10.0 mmol) obtained in Example 8-2 in toluene (33.4 mL), add MsNH 2 (2.38 g, 25.1 mmol), cyanomethylenetributylphosphine (6.57 mL, 25.1 mmol, CAS: 157141-27-0), and stir at 80 °C for 2 hours. After completion of the reaction, concentrate under reduced pressure, and purify the obtained residue by flash column chromatography using Isolera (Biotage; Sfaer HC Duo; mobile phase: n-hexane, 30% - 100% ethyl acetate gradient) to obtain an intermediate as a red viscous substance. Dissolve the above-mentioned intermediate in dichloromethane (50 mL), add TFA (15.3 mL, 200 mmol), and stir at room temperature for 1.5 hours. After completion of the reaction, concentrate the reaction solution under reduced pressure and azeotrope with toluene. Dissolve the obtained residue in a mixed solvent of 1,4-dioxane (20 mL) / water (20 mL), add sodium carbonate (5.31 g, 50.1 mmol) at 0 °C, and then add a 1,4-dioxane solution of 2-[[(1,1-dimethylethyl)diphenylsilyl]oxy]acetyl chloride (4.66 g, 14.0 mmol, CAS: 93853-60-2) dropwise at 0 °C and stir at room temperature for 1 hour. After adding saturated brine to stop the reaction, extract the aqueous layer with ethyl acetate, wash the organic layer with saturated brine, dry over sodium sulfate, filter, and concentrate under reduced pressure. Purify the obtained residue by flash column chromatography using Isolera (Biotage; Sfaer HC Duo; mobile phase: n-heptane, 20% - 100% ethyl acetate gradient) to obtain an intermediate as a brown viscous substance. Dissolve the previously obtained intermediate in dichloromethane (100 mL), and then add Co 2 (CO) 8 (3.77 g, 11.02 mmol) at room temperature and stir for 2 hours. After completion of the reaction, concentrate under reduced pressure. Purify the obtained residue by flash column chromatography using Isolera (Biotage; Sfaer HC Duo; mobile phase: n-hexane, 20% - 50% ethyl acetate gradient) to obtain the title compound as a brown viscous substance (5.20 g, 6.06 mmol).

[0503] Analysis condition A: retention time = 2.36 minutes; ESI-MS(+) observed value m / z = 859.2 (M + H) + Theoretical value m / z = 858.1.

[0504] [Example 8-4] 2-[(tert-Butyldiphenylsilyl)oxy]-1-{7'-methylsulfonyl-3'-oxa-7'-aza-10',11'-dicobaltaspiro[azetidine-3,2'-tetracyclo[7.2.0.0 1,10 .0 9,11 undecane]-1-yl}ethan-1-one; Synthesis of hexakis(methanidylidyneoxidanium)

[0505] [Chemical Formula 64]

[0506]

[0507] Dissolve N-{3-[(1-{2-[(tert-Butyldiphenylsilyl)oxy]acetyl}-3-[4-(methoxymethyl)-1,2-dicobaltatricyclo[1.1.0.0 2,4 butan-3-yl)oxy]propyl}methanesulfonamide; hexakis(methanidylidyneoxidanium) (4.50 g, 5.24 mmol) obtained in Example 8-3 in dichloromethane (105 mL), add boron trifluoride diethyl ether complex (1.99 mL, 15.7 mmol) at 0 °C and stir for 3 hours. After adding saturated aqueous sodium bicarbonate solution to the reaction solution to stop the reaction, extract the aqueous layer with ethyl acetate, wash the organic layer with saturated brine, dry over sodium sulfate and filter, and concentrate under reduced pressure. Purify the obtained residue by flash column chromatography using Isolera (Biotage; Sfaer HC Duo; mobile phase n-heptane, 20% - 50% ethyl acetate gradient) to obtain the title compound as a reddish-brown viscous substance (2.88 g, 3.48 mmol).

[0508] Analysis Condition A: Retention time = 2.34 minutes; ESI-MS(+) observed m / z = 827.3 (M+H) + Theoretical value m / z = 826.1.

[0509] [Example 8-5] Synthesis of 2-hydroxy-1-{9-methylsulfonyl-5-oxa-2,9-diazaspiro[3.8]dodec-11-yn-2-yl}ethan-1-one

[0510] [Chemical Formula 65]

[0511]

[0512] 2-[(tert-Butyldiphenylsilyloxy)]-1-{7'-methanesulfonyl-3'-oxa-7'-aza-10',11'-dicobaltaspiro[azetidine-3,2'-tetracyclo[7.2.0.0 1,10 .0 9,11 undecane]-1-yl}ethan-1-one obtained in Example 8-4; hexakis(methanidylidyneoxidanium) (2.5 g, 3.0 mmol) was dissolved in THF (101 mL), and then CAN (9.95 g, 18.1 mmol) was added at 0 °C. After stirring at 0 °C for 3 hours, the reaction solution was added dropwise to a mixed solvent of ethyl acetate (50 mL) / saturated aqueous sodium hydrogen carbonate solution (50 mL) cooled to 0 °C to stop the reaction. The aqueous layer was extracted with ethyl acetate, the organic layer was washed with saturated brine, dried over sodium sulfate, filtered, and concentrated under reduced pressure. The obtained residue was purified by flash column chromatography using Isolera (Biotage; Sfaer HC Duo; mobile phase: n-hexane, 10% - 80% ethyl acetate gradient) to obtain a colorless solid compound. The obtained compound (1.0 g, 1.85 mmol) was dissolved in THF (18.5 mL), and then acetic acid (0.85 mL, 14.8 mmol) was added at 0 °C. Further, tetramethylammonium fluoride tetrahydrate (1.38 g, 14.8 mmol, CAS: 17787-40-5) was added to the reaction solution and stirred at room temperature for 18 hours. After concentrating the reaction solution under reduced pressure, the residue was washed with methanol, and the white solid was filtered and dried to obtain the title compound (391 mg, 1.29 mmol).

[0513] Analysis condition A: retention time = 1.25 minutes; ESI-MS(+) observed value m / z = 303.1 (M + H) + Theoretical value m / z = 302.4.

[0514] 1 H-NMR(500 MHz, CDCl 3 ) δ 4.25 - 4.10 (m, 4H), 4.05 - 3.95 (m, 4H), 3.94 - 3.88 (m, 2H), 3.46 - 3.40 (m, 2H), 3.02 - 2.96 (m, 1H), 2.86 (s, 3H), 1.96 - 1.88 (m, 2H).

[0515] [Example 8-6] Synthesis of 1-{[(2-{9-Methanesulfonyl-5-oxa-2,9-diazaspiro[3.8]dodec-11-yn-2-yl}-2-oxoethoxy)carbonyl]oxy}tetrahydropyrrole-2,5-dione

[0516] [Chemical Formula 66]

[0517]

[0518] Dissolve 2-hydroxy-1-{9-(methylsulfonyl)-5-oxa-2,9-diazaspiro[3.8]dodec-11-yn-2-yl}ethan-1-one (151 mg, 0.50 mmol) obtained in Example 8-5 in acetonitrile (10 mL). Add N,N-diisopropylethylamine (262 μL, 1.50 mmol) and bis(2,5-dioxopyrrolidin-1-yl) carbonate (384 mg, 1.50 mmol, CAS: 74124-79-1) at room temperature, and heat to 40 °C and stir for 1 hour. After concentrating the reaction solution under reduced pressure, dilute it with ethyl acetate and water and extract with ethyl acetate. Wash the organic layer with saturated brine, dry it over sodium sulfate, filter, and concentrate under reduced pressure. Purify the obtained residue by flash column chromatography using Isolera (Biotage; Sfaer HC Duo; mobile phase: hexane, 20% - 100% ethyl acetate gradient) to obtain the title compound as a white solid (130 mg, 0.29 mmol).

[0519] Analysis condition A: retention time = 1.86 minutes; ESI-MS(+) observed value m / z = 444.2 (M + H) + Theoretical value m / z = 443.4.

[0520] 1 1H-NMR (500 MHz, CDCl 3 ) δ 4.80 (s, 2H), 4.49 - 4.38 (m, 2H), 4.23 - 4.14 (m, 2H), 4.05 - 3.87 (m, 4H), 3.46 - 3.38 (m, 2H), 2.89 - 2.83 (m, 7H), 1.96 - 1.88 (m, 2H).

[0521] [Example 9-1] Synthesis of tert-butyl 3-{[(4-hydroxybut-2-yn-1-yl)oxy]methyl}-3-(hydroxymethyl)azetidine-1-carboxylate

[0522] [Chemical Formula 67]

[0523]

[0524] tert-Butyl 3,3-bis(hydroxymethyl)azetidine-1-carboxylate (8.11 g, 37.3 mmol) obtained by the method of Example 1-1 was dissolved in THF (93 mL) / DMF (93 mL) and cooled to 0 °C. Then, LHMDS (28.7 mL, 37.3 mmol) was added and stirred at 0 °C for 30 minutes. (1,1-Dimethylethyl)[(4-iodo-2-butyn-1-yl)oxy]dimethylsilane (11.8 g, 38.0 mmol, CAS: 219996-95-9) was added to the reaction solution, and the mixture was stirred at 0 °C for 3 hours. After adding a saturated aqueous ammonium chloride solution to stop the reaction, the aqueous layer was extracted with ethyl acetate. The organic layer was washed with saturated brine, dried over sodium sulfate, filtered, and concentrated under reduced pressure. The obtained residue was purified by flash column chromatography using Isolera (Biotage; Sfaer HCDuo; mobile phase: n-heptane, 0% - 50% ethyl acetate gradient) to obtain an intermediate as a brown viscous substance. The obtained intermediate was dissolved in THF (55.4 mL), and 1 M TBAF (54.8 mL, 54.8 mmol) was added at room temperature and stirred for 1 hour. After adding water to stop the reaction, the aqueous layer was extracted with dichloroethane. The organic layer was washed with saturated brine, dried over sodium sulfate, filtered, and concentrated under reduced pressure. The obtained residue was purified by flash column chromatography using Isolera (Biotage; Sfaer HC Duo; mobile phase: n-heptane, 80% - 100% ethyl acetate gradient, then mobile phase: dichloromethane, 15% methanol) to obtain the title light brown viscous substance (5.0 g, 17.5 mmol).

[0525] Analysis condition A: retention time = 1.24 minutes; ESI-MS(+) observed value m / z = 286.2 (M + H) + Theoretical value m / z = 285.2.

[0526] [Example 9-2] hexakis(methanidylidyneoxidanium); 3’,7’-dioxa-10’,11’-dicobalt spiro[azetidine-3,5’-tetracyclo[7.2.0.0 1,10 .0 9,11 undecane]-1-carboxylate tert-butyl ester synthesis

[0527] [Chemical formula 68]

[0528]

[0529] Dissolve tert-butyl 3-{[(4-hydroxybut-2-yn-1-yl)oxy]methyl}-3-(hydroxymethyl)azetidine-1-carboxylate (1.00 g, 3.50 mmol) obtained in Example 9-1 in dichloromethane (35 mL), and add Co 2 (CO) 8 (1.44 g, 4.21 mmol) at room temperature and stir for 1.5 hours. After completion of the reaction, concentrate under reduced pressure. Purify the obtained residue by flash column chromatography using Isolera (Biotage; Sfaer HC Duo; mobile phase: n-heptane, 50% - 80% ethyl acetate gradient) to obtain an intermediate as a brown viscous substance. Dissolve the obtained intermediate in dichloromethane (292 mL), add methanesulfonic acid (0.076 mg, 0.788 mmol, CAS: 75-75-2) at room temperature and stir for 3 hours. Stop the reaction by adding saturated aqueous sodium bicarbonate solution to the reaction mixture, extract the aqueous layer with dichloromethane, wash the organic layer with saturated brine, dry over sodium sulfate, filter, and concentrate under reduced pressure. Purify the obtained residue by flash column chromatography using Isolera (Biotage; Sfaer HC Duo; mobile phase: n-heptane, 10% - 30% ethyl acetate gradient) to obtain the title compound as a brown viscous substance (0.124 g, 0.224 mmol).

[0530] Analysis condition A: retention time = 2.08 minutes; ESI-MS(+) observed m / z = 498.0 (M - C 4 H 8 + H) + Theoretical m / z = 553.0.

[0531] 1 1H-NMR (500 MHz, CDCl 3 ) δ 5.03 (s, 4H), 3.99 (s, 4H), 3.64 (s, 4H), 1.43 (s, 9H).

[0532] [Example 9-3] Synthesis of tert-butyl 6,11-dioxa-2-azaspiro[3.8]dodec-8-yne-2-carboxylate

[0533] [Chemical formula 69]

[0534]

[0535] Dissolve hexakis(methanidylidyneoxidanium); 3’,7’-dioxa-10’,11’-dicobalt spiro[azetidine-3,5’-tetracyclo[7.2.0.01,10 .0 9,11 tert-Butyl undecane-1-carboxylate (0.124 g, 0.224 mmol) was dissolved in tetrahydrofuran (22 mL), and CAN (0.737 g, 1.35 mmol) was added thereto at 0 °C. After stirring at 0 °C for 1 hour, the reaction solution was added dropwise to a mixed solvent of ethyl acetate (200 mL) / saturated aqueous sodium hydrogen carbonate solution (100 mL) cooled to 0 °C to stop the reaction. The aqueous layer was extracted with ethyl acetate, the organic layer was washed with saturated brine, dried over sodium sulfate, filtered, and concentrated under reduced pressure. The obtained residue was purified by flash column chromatography using Isolera (Biotage; Sfaer HC Duo; mobile phase: n-heptane, 10% - 30% ethyl acetate gradient) to obtain the title compound as a colorless powder (0.026 g, 0.097 mmol).

[0536] Analysis condition A: retention time = 1.49 minutes; ESI-MS(+) observed value m / z = 212.1 (M - C 4 H 8 + H) + Theoretical value m / z = 267.2

[0537] 1 1H-NMR (500 MHz, CDCl 3 ) δ 4.16 (s, 4H), 3.99 (s, 4H), 3.59 (s, 4H), 1.43 (s, 9H).

[0538] [Example 9-4] Synthesis of 2-[(tert-butyldiphenylsilyl)oxy]-1-{6,11-dioxa-2-azaspiro[3.8]dodec-8-yn-2-yl}ethan-1-one

[0539] [Chemical formula 70]

[0540]

[0541] Dissolve tert-butyl 6,11-dioxo-2-azaspiro[3.8]dodec-8-yn-2-carboxylate (0.026 g, 0.097 mmol) obtained in Example 9-3 in dichloromethane (1.0 mL), and then add TFA (0.225 mL, 2.92 mmol) at room temperature. After stirring at room temperature for 1 hour, add toluene and concentrate under reduced pressure, and then perform azeotropy with toluene. Add 1,4-dioxane (1.0 mL) / water (1.0 mL) to the obtained residue, and add sodium carbonate (103 mg, 0.97 mmol) at 0 °C. Further, a solution of 2-[[(1,1-dimethylethyl)diphenylsilyl]oxy]acetyl chloride (0.080 mg, 0.243 mmol, CAS: 93853-60-2) prepared separately in 1,4-dioxane was added dropwise at 0 °C, and stirred at room temperature for 1 hour. After completion of the reaction, add a saturated aqueous sodium bicarbonate solution to stop the reaction. Extract the aqueous layer with ethyl acetate, wash the organic layer with saturated brine, dry over sodium sulfate, filter, and concentrate under reduced pressure. The obtained residue was purified by flash column chromatography using Isolera (Biotage; Sfaer HC Duo; mobile phase: n-heptane, 10% - 50% ethyl acetate gradient) to obtain the title compound as a colorless viscous substance (0.041 g, 0.088 mmol).

[0542] Analysis condition A: retention time = 2.04 minutes; ESI-MS(+) observed value m / z = 464.3 (M + H) + Theoretical value m / z = 463.6.

[0543] 1 H-NMR(500MHz,CDCl 3 )δ7.67 - 7.61(m,4H),7.48 - 7.37(m,6H),4.21 - 4.12(m,6H),4.02 - 3.90(m,6H),3.68(s,2H),1.07(s,9H).

[0544] [Example 9-5] Synthesis of 1-{6,11-dioxo-2-azaspiro[3.8]dodec-8-yn-2-yl}-2-hydroxyethan-1-one

[0545] [Chemical formula 71]

[0546]

[0547] Dissolve 2-[(tert-butyldiphenylsilyl)oxy]-1-{6,11-dioxa-2-azaspiro[3.8]dodec-8-yn-2-yl}ethan-1-one (0.041 g, 0.088 mmol) obtained in Example 9-4 in tetrahydrofuran (1.0 mL), and then add TBAF (0.132 mL, 0.13 mmol, 1 M THF solution) at room temperature. After stirring at room temperature for 1.5 hours, the reaction solution was concentrated under reduced pressure. The obtained residue was purified by flash column chromatography using Isolera (Biotage; Sfaer HC Duo; mobile phase: n-heptane, 50% - 100% ethyl acetate gradient, then mobile phase: ethyl acetate, 5% methanol gradient) to obtain the title compound as a colorless powder (0.018 g, 0.080 mmol).

[0548] Analysis condition A: retention time = 0.85 minutes; ESI-MS(+) observed value m / z = 226.2 (M + H) + Theoretical value m / z = 225.2.

[0549] 1 H-NMR(500MHz,CDCl 3 )δ4.17(s,4H),4.06 - 3.96(m,4H),3.97(s,2H),3.82 - 3.61(m,4H),3.06(s,1H).

[0550] [Example 10-1] Synthesis of tert-butyl 4,4-bis(hydroxymethyl)piperidine-1-carboxylate

[0551] [Chemical formula 72]

[0552]

[0553] Dissolve tert-butyl 4-formylpiperidine-1-carboxylate (10.0 g, 46.9 mmol, CAS: 137076-22-3) in MeOH (150 mL), and add H in which KOH (10.92 g, 195 mmol) is dissolved 2O (50 mL), 37% aqueous formaldehyde solution (38.4 mL, 516 mmol), after stirring at room temperature for 150 minutes, stirred at 40 °C for 5 hours. Methanol was removed by distillation under reduced pressure from the reaction solution, saturated aqueous ammonium chloride solution was added, and extraction was carried out with ethyl acetate. The obtained organic layer was dried over sodium sulfate, filtered, and concentrated under reduced pressure. The obtained residue was purified by flash column chromatography using Isolera (Biotage; Sfaer HC Duo; mobile phase DCM, 3% - 15% MeOH gradient), and the fraction containing the target substance was concentrated under reduced pressure to obtain the title compound as a colorless powder (10.0 g, 40.8 mmol).

[0554] 1 H-NMR(500MHz,CDCl 3 )δ3.67(s,4H),3.44 - 3.36(m,4H),2.39(s,2H),1.49 - 1.44(m,13H).

[0555] [Example 10 - 2] Synthesis of (9H - Fluoren - 9 - yl)methyl 4,4 - bis(hydroxymethyl)piperidine - 1 - carboxylate

[0556] [Chemical formula 73]

[0557]

[0558] tert - Butyl 4,4 - bis(hydroxymethyl)piperidine - 1 - carboxylate (1.00 g, 4.08 mmol) obtained in Example 10 - 1 was dissolved in 1,4 - dioxane (10.2 mL), hydrogen chloride 1,4 - dioxane solution (4 M, 10.2 mL, 40.8 mmol) was added, and after stirring at room temperature for 5 hours, concentrated under reduced pressure. The obtained residue and DIPE A (1.05 g, 8.15 mmol) were dissolved in dichloromethane (20 mL), Fmoc - OSu (1.25 g, 3.87 mmol) was added at room temperature, and stirred as it was for 2 hours. Dichloromethane was added to the reaction solution for extraction, and the obtained organic layer was concentrated under reduced pressure. The obtained residue was purified by flash column chromatography using Isolera (Biotage; Sfaer HC Duo; mobile phase DCM, 0% - 15% MeOH gradient), and the fraction containing the target substance was concentrated under reduced pressure to obtain the title compound as a colorless viscous substance (1.45 g, 3.95 mmol).

[0559] Analysis condition A: retention time = 1.56 minutes; ESI - MS(+) observed value m / z = 368.2 (M + H) + Theoretical value m / z = 367.2.

[0560] [Example 10-3] hexakis(methanidylidyneoxidanium); 3’,7’-dioxa-10’,11’-dicobalt spiro[piperidine-4,5’-tetracyclo[7.2.0.0 1,10 .0 9,11 undecane]-1-carboxylic acid (9H-fluoren-9-yl) methyl ester synthesis

[0561] [Chemical formula 74]

[0562]

[0563] For a solution of 4,4-bis(hydroxymethyl)piperidine-1-carboxylic acid (9H-fluoren-9-yl) methyl ester (250 mg, 0.68 mmol) obtained in Example 10-2 and methanesulfonic acid (0.131 g, 1.36 mmol) dissolved in dichloromethane (54.4 mL), a dichloromethane solution (13.6 mL) of hexakis(methanidylidyneoxidanium); [4-(hydroxymethyl)-1,2-dicobalt tricyclo[1.1.0.0 2,4 butane-3-yl]methanol (253 mg, 0.68 mmol, CAS: 55975-76-3, synthesized by the method described in Organic Letters (2015), 17(16), 4086-4089) was added dropwise at room temperature over 90 minutes. In addition, after stirring at room temperature for 30 minutes, 10% aqueous sodium bicarbonate solution was added to stop the reaction, and extraction was carried out with dichloromethane. The obtained organic layer was concentrated under reduced pressure. The obtained residue was purified by flash column chromatography using Isolera (Biotage; Sfaer HCDuo; mobile phase: n-heptane, 10% - 30% ethyl acetate gradient), and the fraction containing the target substance was concentrated under reduced pressure to obtain the title compound as a brown powder (280 mg, 0.398 mmol).

[0564] Analysis condition A: retention time = 2.22 minutes; ESI-MS(+) observed value m / z = 704.2 (M+H) + Theoretical value m / z = 703.0

[0565] [Example 10-4] 8,13-dioxa-3-azaspiro[5.8]tetradec-10-yne-3-carboxylic acid (9H-fluoren-9-yl) methyl ester synthesis

[0566] [Chemical formula 75]

[0567]

[0568] The methyl (9H-fluoren-9-yl) ester of 3’,7’-dioxo-10’,11’-dicobaltspiro[piperidine-4,5’-tetracyclo[7.2.0.0 1,10 .0 9,11 undecane]-1-carboxylic acid obtained in Example 10-3 (0.100 g, 0.142 mmol) was dissolved in tetrahydrofuran (14.2 mL), and then CAN (0.234 g, 0.426 mmol) was added at 0 °C. After stirring at 0 °C for 2 hours, the reaction was stopped by adding diethyl ether and 10% aqueous sodium hydrogen carbonate solution. The aqueous layer was extracted with diethyl ether, the organic layer was washed with saturated brine, dried over sodium sulfate and filtered, and concentrated under reduced pressure. The resulting residue was purified by flash column chromatography using Isolera (Biotage; Sfaer HC Duo; mobile phase: n-heptane, 10% - 40% ethyl acetate gradient) to obtain the title compound as a colorless viscous substance (0.025 g, 0.060 mmol).

[0569] Analysis condition A: retention time = 1.87 minutes; ESI-MS(+) observed m / z = 418.3 (M+H) + Theoretical m / z = 417.2.

[0570] 1 H-NMR(500 MHz, CDCl 3 ) δ 7.76 (d, J = 7.5 Hz, 2H), 7.57 (d, J = 7.5 Hz, 2H), 7.43 - 7.36 (m, 2H), 7.34 - 7.28 (m, 2H), 4.42 (d, J = 6.9 Hz, 2H), 4.24 (t, J = 6.9 Hz, 1H), 4.20 - 4.05 (m, 4H), 3.91 - 3.71 (m, 2H), 3.67 - 3.54 (m, 2H), 3.51 - 3.37 (m, 4H), 1.57 - 1.39 (m, 2H), 1.36 - 1.16 (m, 2H).

[0571] [Example 11-1] Synthesis of tert-butyl 4-{[(tert-butyldimethylsilyl)oxy]methyl}-4-(hydroxymethyl)piperidine-1-carboxylate

[0572] [Chemical formula 76]

[0573]

[0574] Dissolve tert-butyl 4,4-bis(hydroxymethyl)piperidine-1-carboxylate (1.00 g, 4.08 mmol) obtained in Example 10-1 in tetrahydrofuran (40.8 mL). Under ice cooling, add sodium hydride (163 mg, 4.08 mmol, 60%, dispersed in liquid paraffin) at 0 °C, warm up to room temperature and stir for 30 minutes. Add tert-butyldimethylchlorosilane (614 mg, 4.08 mmol, CAS: 18162-48-6) to the reaction solution and stir overnight at room temperature. Under ice cooling, add saturated ammonium chloride aqueous solution to the reaction solution, extract with ethyl acetate, wash the obtained organic layer with saturated brine, dry over sodium sulfate, filter, and concentrate under reduced pressure. Purify the obtained residue by flash column chromatography using Isolera (Biotage; Sfaer HCDuo; mobile phase: n-heptane, 10% - 50% ethyl acetate gradient), and concentrate the fraction containing the target substance under reduced pressure to obtain the title compound as a colorless powder (1.45 g, 4.03 mmol).

[0575] Analysis condition A: retention time = 2.00 minutes; ESI-MS(+) observed value m / z = 360.3 (M + H) + Theoretical value m / z = 359.3

[0576] [Example 11-2] Synthesis of tert-butyl 4-[({4-[(tert-butyldimethylsilyl)oxy]but-2-yn-1-yl}oxy)methyl]-4-{[(tert-butyldimethylsilyl)oxy]methyl}piperidine-1-carboxylate

[0577] [Chemical formula 77]

[0578]

[0579] Dissolve tert-butyl 4-{[(tert-butyldimethylsilyl)oxy]methyl}-4-(hydroxymethyl)piperidine-1-carboxylate (0.50 g, 1.39 mmol), obtained in Example 11-1, in a mixed solvent of tetrahydrofuran (11.1 mL) and DMF (2.78 mL). Under ice cooling, add sodium hydride (55.6 mg, 1.39 mmol, 60%, dispersed in liquid paraffin). After warming to room temperature, stir for 10 minutes. Add [(4-bromobut-2-yn-1-yl)oxy](tert-butyl)dimethylsilane (732 mg, 2.78 mmol, CAS: 110796-98-0) to the reaction solution and stir at room temperature for 7 hours. Add saturated aqueous ammonium chloride solution to the reaction solution, extract with ethyl acetate, wash the obtained organic layer with saturated brine, dry over sodium sulfate, filter, and concentrate under reduced pressure. Purify the obtained residue by flash column chromatography using Isolera (Biotage; Sfaer HC Duo; mobile phase: n-heptane, 10% - 40% ethyl acetate gradient), and concentrate the fraction containing the target substance under reduced pressure to obtain the title compound as a yellow oil (0.192 g, 0.354 mmol).

[0580] Analysis condition A: retention time = 2.71 minutes; ESI-MS(+) observed value m / z = 542.4 (M + H) + Theoretical value m / z = 541.4.

[0581] [Example 11-3] Synthesis of tert-butyl 4-{[(4-hydroxybut-2-yn-1-yl)oxy]methyl}-4-(hydroxymethyl)piperidine-1-carboxylate

[0582] [Chemical formula 78]

[0583]

[0584] Dissolve tert-butyl 4-[({4-[(tert-butyldimethylsilyl)oxy]but-2-yn-1-yl}oxy)methyl]-4-{[(tert-butyldimethylsilyl)oxy]methyl}piperidine-1-carboxylate (0.195 g, 0.360 mmol), obtained in Example 11-2, in tetrahydrofuran (3.6 mL). Add 1 M TBAF (0.792 mL, 0.792 mmol, THF solution) at room temperature. After stirring at room temperature for 2 hours, concentrate the reaction solution under reduced pressure. Purify the obtained residue by flash column chromatography using Isolera (Biotage; Sfaer HC Duo; mobile phase: n-heptane, 10% - 80% ethyl acetate gradient) to obtain the title compound as a pale yellow oil (0.102 g, 0.325 mmol).

[0585] Analysis condition A: retention time = 1.40 minutes; ESI-MS(+) observed value m / z = 314.3 (M+H) + Theoretical value m / z = 313.2

[0586] [Example 11-4] hexakis(methanidylidyneoxidanium); 4-(hydroxymethyl)-4-({[4-(hydroxymethyl)-1,2-dicobalt tricyclo[1.1.0.0 2,4 butane-3-yl]methoxy}methyl)piperidine-1-carboxylic acid tert-butyl ester synthesis

[0587] [Chemical formula 79]

[0588]

[0589] Dissolve 4-{[(4-hydroxybut-2-yn-1-yl)oxy]methyl}-4-(hydroxymethyl)piperidine-1-carboxylic acid tert-butyl ester (0.102 g, 0.325 mmol) obtained in Example 11-3 in dichloromethane (3.3 mL), and add Co 2 (CO) 8 (0.134 g, 0.391 mmol) at room temperature and stir for 2 hours. After the reaction is completed, concentrate under reduced pressure. Purify the obtained residue by flash column chromatography using Isolera (Biotage; Sfaer HC Duo; mobile phase: n-heptane, 10% - 80% ethyl acetate gradient) to obtain the title compound as a pale brown oily substance (0.170 g, 0.284 mmol).

[0590] Analysis condition A: retention time = 1.96 minutes; ESI-MS(+) observed value m / z = 582.1 (M-H 2 O+H) + Theoretical value m / z = 599.0

[0591] [Example 11-5] hexakis(methanidylidyneoxidanium); 3’,7’-dioxa-10’,11’-dicobalt spiro[piperidine-4,5’-tetracyclo[7.2.0.0 1,10 .0 9,11 undecane]-1-carboxylic acid tert-butyl ester synthesis

[0592] [Chemical formula 80]

[0593]

[0594] Hexakis(methanidylidyneoxidanium) obtained in Example 11-4; 4-(hydroxymethyl)-4-({[4-(hydroxymethyl)-1,2-dicobalt tricyclo[1.1.0.0 2,4 butane-3-yl]methoxy}methyl)piperidine-1-carboxylic acid tert-butyl ester (0.138 g, 0.230 mmol) was dissolved in dichloromethane (7.7 mL), and methanesulfonic acid (0.020 g, 0.207 mmol) was added at room temperature and stirred for 3 hours. Triethylamine (0.117 g, 1.15 mmol, CAS: 121-44-8) and di-tert-butyl dicarbonate (0.101 g, 0.461 mmol, CAS: 24424-99-5) were added to the reaction solution, and the mixture was stirred at room temperature for 20 minutes. The reaction solution was purified by flash column chromatography using Isolera (Biotage; Sfaer HC Duo; mobile phase: n-heptane, 5% - 80% ethyl acetate gradient) to obtain the title compound as a brown powder (0.055 g, 0.095 mmol).

[0595] Analysis condition A: retention time = 2.19 minutes; ESI-MS(+) observed m / z = 526.0 (M - C 4 H 8 +H) + Theoretical m / z = 581.0.

[0596] [Example 11-6] Synthesis of tert-butyl 8,13-dioxa-3-azaspiro[5.8]tetradec-10-yne-3-carboxylate

[0597] [Chemical formula 81]

[0598]

[0599] Hexakis(methanidylidyneoxidanium) obtained in Example 11-5; 3’,7’-dioxa-10’,11’-dicobalt spiro[piperidine-4,5’-tetracyclo[7.2.0.0 1,10 .0 9,11tert-Butyl [[undecane]]-1-carboxylate (0.060 g, 0.103 mmol) was dissolved in tetrahydrofuran (10.0 mL), and then CAN (0.340 g, 0.619 mmol) was added at 0 °C. After stirring at 0 °C for 1 hour, the reaction solution was added dropwise to a mixed solvent of ethyl acetate / 10% aqueous sodium bicarbonate solution cooled to 0 °C to stop the reaction. The aqueous layer was extracted with ethyl acetate, the organic layer was washed with saturated brine, dried over sodium sulfate, filtered, and concentrated under reduced pressure. The obtained residue was purified by flash column chromatography using Isolera (Biotage; Sfaer HC Duo; mobile phase: n-hexane, 10% - 40% ethyl acetate gradient) to obtain the title compound as a white powder (0.017 g, 0.057 mmol).

[0600] Analysis condition A: retention time = 1.69 minutes; ESI-MS(+) observed value m / z = 240.1 (M - C 4 H 8 +H) + Theoretical value m / z = 295.2.

[0601] 1 1H-NMR (500 MHz, CDCl 3 ) δ 4.26 - 4.03 (m, 4H), 3.80 (s, 2H), 3.60 (s, 2H), 3.39 (s, 4H), 1.58 - 1.12 (m, 13H).

[0602] [Example 11 - 7] Synthesis of 8,13 - dioxo - 3 - azaspiro[5.8]tetradec - 10 - yne hydrochloride

[0603] [Chemical formula 82]

[0604]

[0605] tert-Butyl 8,13 - dioxo - 3 - azaspiro[5.8]tetradec - 10 - yne - 3 - carboxylate (0.012 g, 0.041 mmol) obtained in Example 11 - 6 was dissolved in 1,4 - dioxane (0.41 mL), and then hydrogen chloride 1,4 - dioxane solution (4 M, 50.8 μL, 0.203 mmol) was added at room temperature. After stirring at room temperature for 1 hour, hydrogen chloride 1,4 - dioxane solution (4 M, 85 μL, 0.340 mmol) was added. After further stirring at room temperature for 1 hour, hydrogen chloride 1,4 - dioxane solution (4 M, 270 μL, 1.080 mmol) was added. After stirring at room temperature for 1 hour, it was concentrated and dried under reduced pressure to obtain the title compound as a white powder.

[0606] 1H-NMR (500 MHz, DMSO-d 6 ) δ 8.56 - 8.36 (m, 2H), 4.30 - 4.03 (m, 4H), 3.75 - 3.50 (m, 4H), 3.07 - 2.97 (m, 4H), 1.71 - 1.18 (m, 4H).

[0607] [Example 12-1] Synthesis of Methyl 3-(3-Hydroxypropoxy)-3-(3-methoxyprop-1-yn-1-yl)azetidine-1-carboxylate (9H-Fluoren-9-yl)

[0608] [Chemical Formula 83]

[0609]

[0610] A solution of tert-butyl 3-hydroxy-3-(3-methoxyprop-1-yn-1-yl)azetidine-1-carboxylate (1.20 g, 4.97 mmol) obtained in Example 8-1 in THF (8.3 mL) was added to DMF (8.3 mL). Sodium hydride (0.298 g, 7.46 mmol, 60%, dispersed in liquid paraffin) was added at 0 °C and stirred for 20 minutes. Then, a DMF solution of (3-bromopropoxy)(tert-butyl)dimethylsilane (1.89 g, 7.46 mmol, CAS: 89031-84-5) was added at 0 °C, and the mixture was stirred at room temperature for 14 hours. After adding an aqueous solution of isopropyl acetate / water (100 mL / 100 mL) to stop the reaction, the aqueous layer was extracted with isopropyl acetate. The organic layer was washed with saturated brine, dried over sodium sulfate, filtered, and concentrated under reduced pressure to obtain a colorless liquid intermediate. The obtained intermediate (1.90 g, 4.59 mmol) was dissolved in 11.5 mL of dichloromethane. After cooling to 0 °C, TFA (11.5 mL, 150 mmol) was added and stirred at room temperature for 2 hours. The reaction solution was concentrated under reduced pressure. Potassium bicarbonate (1.43 g, 14.3 mmol), water (10.6 mL), and 1,4-dioxane (10.6 mL) were added to the obtained residue and stirred. Then, a solution of (N-9-fluorenylmethoxycarbonyloxy)succinimide (1.45 g, 4.29 mmol) in 1,4-dioxane (10.6 mL) was added dropwise at 0 °C. After the addition was completed, the temperature was raised to room temperature and stirred overnight. Isopropyl acetate (100 mL) and water (100 mL) were added to the reaction solution. The aqueous layer was extracted with isopropyl acetate. The organic layer was washed with saturated brine, dried over sodium sulfate, filtered, and concentrated under reduced pressure. The obtained residue was purified by flash column chromatography using Isolera (Biotage; Sfaer HCDuo; mobile phase: n-heptane, 20% - 65% ethyl acetate gradient) to obtain the title compound as a brown viscous substance (264 mg, 0.63 mmol).

[0611] Analysis condition A: retention time = 1.85 minutes; ESI-MS(+) observed value m / z = 422.4 (M + H) + Theoretical value m / z = 421.4.

[0612] [Example 12-2] 3-(3-Hydroxypropoxy)-3-[4-(methoxymethyl)-1,2-dicobalt tricyclo[1.1.0.0 2 ,4 butan-3-yl]azetidine-1-carboxylic acid (9H-fluoren-9-yl)methyl ester; synthesis of hexakis(methanidylidyneoxidanium)

[0613] [Chemical Formula 84]

[0614]

[0615] Dissolve methyl (9H-fluoren-9-yl) 3-(3-hydroxypropoxy)-3-(3-methoxyprop-1-yn-1-yl)azetidine-1-carboxylate (803 mg, 1.91 mmol) obtained in Example 12-1 in dichloromethane (9.5 mL), and add Co 2 (CO) 8 (782 mg, 2.29 mmol) at room temperature and stir for 0.5 hour. After completion of the reaction, concentrate under reduced pressure. Purify the resulting residue by flash column chromatography using Isolera (Biotage; Sfaer HC Duo; mobile phase: dichloromethane, 1% - 15% methanol gradient) to obtain an intermediate as a brown viscous substance (1.25 g, 1.77 mmol).

[0616] Analysis condition A: retention time = 2.29 minutes; ESI-MS(+) observed m / z = 676.2 (M-MeOH+H) + Theoretical m / z = 707.4

[0617] [Example 12-3] Synthesis of methyl 3',7'-dioxo-10',11'-dicobaltaspiro[azetidine-3,2'-tetracyclo[7.2.0.0 1 ,10 .0 9.11 undecane]-1-carboxylate; hexakis(methanidylidyneoxidanium)

[0618] [Chemical Formula 85]

[0619]

[0620] Dissolve methyl 3-(3-hydroxypropoxy)-3-[4-(methoxymethyl)-1,2-dicobaltatricyclo[1.1.0.0 2,4(3-Butanyl)azetidine-1-carboxylic acid (9H-fluoren-9-yl)methyl ester; hexakis(methanidylidyneoxidanium) (1.0 g, 1.41 mmol) was dissolved in dichloromethane (283 mL), boron trifluoride diethyl ether complex (0.21 mL, 1.70 mmol) was added at 0 °C and stirred for 2.5 hours. The reaction was stopped by adding saturated aqueous sodium bicarbonate solution, the aqueous layer was extracted with dichloromethane, the organic layer was washed with saturated brine, dried over sodium sulfate and filtered, and concentrated under reduced pressure. The obtained residue was purified by flash column chromatography using Isolera (Biotage; Sfaer HCDuo; mobile phase: n-heptane, 10% - 60% ethyl acetate gradient) to obtain the title compound as a brown viscous substance (123 mg, 0.181 mmol).

[0621] Analysis condition A: retention time = 2.38 minutes; ESI-MS(+) observed m / z = 676.2 (M+H) + Theoretical m / z = 675.4.

[0622] [Example 12-4] Synthesis of (9H-Fluoren-9-yl)methyl 5,9-dioxo-2-azaspiro[3.8]dodec-11-yn-2-carboxylate

[0623] [Chemical formula 86]

[0624]

[0625] To the 3',7'-dioxo-10',11'-dicobaltspiro[azetidine-3,2'-tetracyclo[7.2.0.0 1,10 .0 9.11Methyl (9H-fluoren-9-yl) undecane-1-carboxylate; Diisopropyl ether (4.4 mL) and WAKOSIL (registered trademark)-c200 (FUJIFILM Wako Pure Chemical Corporation; 2.5 g, CAS: 63231-67-4) were added to hexakis(methanidylidyneoxidanium) (150 mg, 0.22 mmol), and then CAN (1.22 g, 2.22 mmol) was added at 0 °C. The temperature was then raised to room temperature and the mixture was stirred for 3 hours. After completion of the reaction, the reaction mixture was passed through WAKOSIL (registered trademark)-c200 (FUJIFILM Wako Pure Chemical Corporation), and the insoluble matter was filtered off. The filtrate was then concentrated under reduced pressure. The resulting residue was purified by flash column chromatography using Isolera (Biotage; Sfaer HC Duo;; mobile phase: hexane, 50% - 80% ethyl acetate gradient, then mobile phase: dichloromethane, 1% - 20% methanol gradient) to obtain the title compound as a pale yellow powder (0.050 g, 0.128 mmol).

[0626] Analysis condition A: retention time = 2.03 minutes; ESI-MS(+) observed value m / z = 390.3 (M+H) + Theoretical value m / z = 389.4

[0627] The following shows synthesis examples of peptide-nucleic acid conjugates as examples, and the present invention will be described in more detail, but the present invention is not limited to these examples.

[0628] It should be noted that the peptides, nucleic acids, and click reagents for conjugation are the following compounds.

[0629] · Peptide compound (CAS: 2699074-52-5. Refer to WO2021167107. Hereinafter, sometimes referred to as peptide F).

[0630] [Chemical formula 87]

[0631]

[0632] · Nucleic acid compound (synthesized by referring to US20180010126 and according to a conventional synthesis method. Hereinafter, sometimes referred to as NS-S01-20. The structural formula of the nucleic acid compound is as Figure 4 shown)

[0633] Regarding 1-{[(2-{9-Methylsulfonyl-5-oxa-2,9-diazaspiro[3.8]dodec-11-yn-2-yl}-2-oxoethoxy)carbonyl]oxy}pyrrolidine-2,5-dione obtained in Example 8-6, it is a click reagent compound. (Hereinafter, sometimes referred to as click reagent X).

[0634] [Chemical Formula 88]

[0635]

[0636] The compound obtained by introducing click reagent X into NS-S01-20 is as follows. (Hereinafter, it may be referred to as the click reagent X-introduced product).

[0637] [Chemical Formula 89]

[0638]

[0639] The compound obtained by conjugating the click reagent X-introduced product with peptide F is as follows. (Hereinafter, it may be referred to as PN-S01-10).

[0640] [Chemical Formula 90]

[0641]

[0642] Analysis condition 1 (LC-MS)

[0643] Column: XBridge Premier Oligonucleotide BEH C18 column (2.5 μm), 2.1 mm × 150 mm (Waters)

[0644] Mobile phase A: Aqueous solution containing 400 mM 1,1,1,3,3,3-hexafluoro-2-propanol and 15 mM triethylamine Mobile phase B: Methanol solution containing 400 mM 1,1,1,3,3,3-hexafluoro-2-propanol and 15 mM triethylamine Flow rate: 0.2 mL / min

[0645] Column temperature: 60 °C

[0646] Detection wavelength: 260 nm

[0647] Gradient condition: 10% B (0 min) → 85% B (22.5 min) → 100% B (23 min) → 100% B (25 min) → 10% B (26 min) → 10% B (30 min)

[0648] Purification condition 1

[0649] Column: Triart Prep C18-S column (10 μm), 10 mm × 250 mm (YMC)

[0650] Mobile phase A: 100 mM aqueous solution of triethylamine acetate

[0651] Mobile phase B: Acetonitrile

[0652] Flow rate: 6.0 mL / min

[0653] Column temperature: Room temperature

[0654] Detection wavelength: 260 nm

[0655] Gradient condition: 20% B (0 min) → 40% B (80 min)

[0656] Desalting condition 1

[0657] Column: Triart Prep C18-S column (10 μm), 10 mm × 250 mm (YMC Co., Ltd.)

[0658] Mobile phase A: Water for injection

[0659] Mobile phase B: Acetonitrile

[0660] Flow rate: 6.0 mL / min

[0661] Column temperature: Room temperature

[0662] Detection wavelength: 260 nm

[0663] Gradient condition: 0% B (6.5 min)

[0664] Elution condition 1

[0665] Column: Triart Prep C18-S column (10 μm), 10 mm × 250 mm (YMC Co., Ltd.)

[0666] Mobile phase A: Water for injection

[0667] Mobile phase B: Acetonitrile

[0668] Flow rate: 6.0 mL / min

[0669] Column temperature: Room temperature

[0670] Detection wavelength: 260 nm

[0671] Gradient condition: 0% B (0 min) → 45% B (30 min)

[0672] [Example 13-1] Synthesis of PN-S01-10

[0673] 1) Synthesis of click reagent X-introducing body

[0674] NS-S01-20 (29.4 mg, 4.0 μmol) in a glass vial was dissolved in 150 mM phosphate buffer (560 μL, pH 7.2) containing 68 μL of DMSO to prepare an NS-S01-20 solution. In another glass vial, click reagent X (5.3 mg, 12 μmol) was dissolved in DMSO (690 μL) to prepare a click reagent X solution. 345 μL (1.5 eq) of the click reagent X solution was added to all of the NS-S01-20 solution, and the mixture was stirred at room temperature for 2 hours. Stirring was stopped when the peak of NS-S01-20 was confirmed to be 3% or less under analytical condition 1.

[0675] Retention time (analytical condition 1): NS-S01-20: 9.2 minutes; click reagent X: 9.5 minutes

[0676] 2) Synthesis of PN-S01-10

[0677] Peptide F (13.8 mg, 6.0 μmol, 1.5 eq.) in a glass vial was dissolved in DMSO (1425 μL) to prepare a peptide F solution. The peptide F solution (1450 μL, 1.5 eq.) was added to the solution containing the click reagent X-introducing body, and the mixture was stirred at room temperature for 2 hours. Stirring was stopped when the peak of the click reagent X-introducing body was confirmed to be 3% or less under analytical condition 1. A crude PN-S01-10 solution with a purity of 64% was obtained.

[0678] Retention time (analytical condition 1): click reagent X-introducing body: 9.5 minutes; PN-S01-10: 10.8 minutes

[0679] [Example 13-2] Purification of PN-S01-10

[0680] The solution obtained by adding mobile phase A (118 mL) of purification condition 1 to the crude PN-S01-10 solution and diluting it was used as the injection solution, and separation and purification were carried out under purification condition 1. The components with a separation retention time of 76 minutes to 85 minutes were separated, and all of the separated solutions were combined. The separated solution was analyzed under analytical condition 1 to obtain a purified PN-S01-10 solution with a purity of 90.7% (recovery rate 100%).

[0681] [Example 13-3] Desalting / Elution of PN-S01-10

[0682] The solution obtained by adding the mobile phase A (100 mL) of desalting condition 1 to the PN-S01-10 purification solution and diluting it was used as the injection solution, and desalting was carried out under desalting condition 1. After injection into the column, two column volumes of water for injection were passed through to confirm that the conductivity was 50 μS / cm or less. Then, the PN-S01-10 retained in the column was eluted under elution condition 1. The components with a retention time of 10 minutes to 13 minutes were separated, and all the separated solutions were combined. The separated solution was analyzed under analysis condition 1 to obtain a desalted solution of PN-S01-10 with a purity of 92.2% (recovery rate 100%).

[0683] [Example 13-4] Freeze-drying of PN-S01-10

[0684] Water for injection (42 mL) was added to the desalted solution of PN-S01-10 (42 mL) and diluted. After freezing the diluted solution, drying was carried out using a freeze dryer for 3 days. After drying, white solid PN-S01-10 (16.1 mg, purity 90.4%, yield 40.4%) was obtained.

Claims

1. A compound or a salt thereof, which is represented by formula (I), Formula (I): [Chemical formula 1] wherein, X 1 and X 2 are each independently an oxygen atom or -N(Y 1 ), Q 1 is any one of the groups represented by [Chemical Formula 2]. [Chemical formula 2] Q 2 selected from C 2-4 an alkylene group; or Q 1 is methylene, Q 2 is any one of the groups represented by [Chemical Formula 3-1], [Chemical formula 3-1] Y 1 selected from a hydrogen atom, a C 1-6 alkanesulfonyl group and a nitrobenzenesulfonyl group which are optionally substituted with one or more halogen atoms, R 1 selected from a hydrogen atom, C 1-6 alkyl, (C 1-6 alkyl)carbonyl, (C 1-6 alkoxy)carbonyl, benzyloxycarbonyl, [(9H-fluoren-9-yl)methoxy]carbonyl, -CO-Q 3 -X 3 、-COO-Q 3 -X 3 、-CONR 2 -Q 3 -X 3 、-SO 2 -Q 3 -X 3 and -SO 2 NR 2 -Q 3 -X 3 , R 2 is a hydrogen atom or a C 1-6 alkyl group X 3 is -COR 5 、-OL 1 、-NR 3 R 4 or the group shown in [Chemical Formula 3-2], [Chemical formula 3-2] R 3 is a hydrogen atom or a C 1-6 alkyl group R 4 is a hydrogen atom, (C 1-6 alkyl)carbonyl, (C 1-6 alkoxy)carbonyl, -COX 4 or benzyloxycarbonyl, Q 3 is C 1-10 alkylene, cyclo C 3-10 alkylene, inserted with cyclo C 3-10 alkylene of C 2-10 alkylene, -(cyclo C 3-10 alkylene)-(C 1-10 alkylene)-, -(C 1-10 alkylene)-(cyclo C 3-10 alkylene)- or -(CH 2 CH 2 O) n CH 2 CH 2 -, n is an integer from 1 to 10, L 1 is a hydrogen atom, -COX 4 or (C 1-6 alkyl)diphenylsilyl, R 5 is a hydroxyl group, X 4 or C 1-6 alkoxy group X 4 is the group shown in [Chemical Formula 4], [Chemical formula 4] The triple bond of formula (I) is optionally protected by Co 2 (CO) 6 groups.

2. The compound or a salt thereof according to claim 1, wherein, Q 3 is ethylene or propylene.

3. The compound or a salt thereof according to claim 1 or 2, wherein, R 1 Selected from -CO-CH 2 -OH, benzyloxycarbonyl, tert-butoxycarbonyl, [(9H-fluoren-9-yl)methoxy]carbonyl.

4. The compound or a salt thereof according to any one of claims 1 to 3, wherein, Y 1 is C 1-6 alkylsulfonyl, trifluoromethanesulfonyl and 2-nitrobenzenesulfonyl.

5. The compound or a salt thereof according to claim 4, wherein, Y 1 is mesyl or tert-butanesulfonyl.

6. The compound or a salt thereof according to any one of claims 1 to 5, wherein, Q 1 is any one of the groups represented by [Chemical Formula 5] [Chemical formula 5] 7. The compound or a salt thereof according to any one of claims 1 to 5, wherein, Q 2 is any one of the groups represented by [Chemical Formula 6], [Chemical formula 6] 8. The compound or a salt thereof according to any one of claims 1 to 5, wherein, Q 1 is the group represented by [Chemical Formula 7], [Chemical formula 7] R 1 is benzyloxycarbonyl.

9. The compound or a salt thereof according to any one of claims 1 to 5, wherein, Q 1 is the group represented by [Chemical Formula 8], [Chemical formula 8] Q 2 Selected from C 2-3 an alkylene group or Q 2 is the group shown in [Chemical Formula 9], [Chemical formula 9] R 1 is benzyloxycarbonyl.

10. The compound or a salt thereof according to claim 1, which is selected from the compounds represented by [Chemical formula 10], [Chemical formula 10] 11. A method for producing a conjugate of a functional molecule using the compound or a salt thereof according to any one of claims 1 to 10, which comprises: reacting a first functional molecule with a second functional molecule to obtain a conjugate in which the first functional molecule and the second functional molecule are linked by a linker represented by formula IVa or formula IVb, The first functional molecule has a group represented by formula (II), formula (II): [Chemical formula 11] wherein, X 1 and X 2 are each independently an oxygen atom or -N(Y 1 ), Q 1 is any one of the groups represented by [Chemical Formula 12], [Chemical formula 12] Q 2 selected from C 2-4 an alkylene; or Q 1 is methylene, Q 2 is any one of the groups represented by [Chemical Formula 13], [Chemical formula 13] Q 5 is a linker or single bond bonded to the first functional molecule, and a second functional molecule having a group represented by the formula (III), formula (III): -Q 6 -N 3 (III) Q 6 is a linker or single bond bonded to the second functional molecule, of formula IVa or formula IVb: [Chemical formula 14] 12. The method according to claim 11, wherein, The first functional molecule and the second functional molecule are each independently a pharmacologically active compound, a labeled compound, a peptide, a protein, a nucleic acid, a molecule used in a drug delivery system.

13. The method according to claim 12, wherein, Either the first functional molecule or the second functional molecule is a pharmacologically active compound, a labeled compound, a peptide, a protein, a nucleic acid that has specific binding affinity for a target.

14. A conjugate formed by linking a first functional molecule and a second functional molecule through a linker, The linker is represented by formula (IVa) or formula (IVb), Formula (IVa) or formula (IVb): [Chemical formula 15] wherein X 1 , X 2 , Q 1 , Q 2 and Q 6 are as defined in claim 11.

15. The conjugate according to claim 14, wherein, The first functional molecule and the second functional molecule are each independently a pharmacologically active compound, a labeled compound, a peptide, a protein, a nucleic acid, a molecule used in a drug delivery system.

16. The conjugate according to claim 15, wherein, Either the first functional molecule or the second functional molecule is a pharmacologically active compound, a labeled compound, a peptide, a protein, a nucleic acid that has specific binding affinity for a target.

17. A method for producing a compound represented by formula (I), which comprises: obtaining a corresponding ring-closed product by treating a compound represented by formula (V) in the presence of an acid, Formula (I): [Chemical formula 16] wherein X 1 and X 2 , Q 1 and Q 2 are as defined in claim 1, and the triple bond of formula (I) is optionally protected by Co 2 (CO) 6 ​ Formula (V): [Chemical formula 17] wherein X 1 , X 2 , Q 1 and Q 2 are as defined in claim 1 wherein, In the presence of hydroxyl and amino groups, they are protected by protecting groups, and L is a leaving group.

18. The manufacturing method according to claim 17, wherein, the acid is a Lewis acid or a Bronsted acid.

19. The manufacturing method according to claim 17, wherein, L is -OR 10 or a halogen atom, R 10 is a hydrogen atom, (C 1-6 alkyl)carbonyl, acetyl, tris(C 1-6 alkyl)silyl, C 1-6 alkyl, benzenesulfonyl or C 1-6 alkanesulfonyl.

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