Method for preparing bicyclic phosphoramidites

Abstract

The present invention provides: a crystalline 2,4-crosslinked common intermediate useful for producing a plurality of ENA monomers; a method for stereoselectively producing said intermediate; and a method for efficiently producing individual ENA monomers using said intermediate.

Description

[Technical field]

[0001] The present invention relates to a novel crystalline common intermediate for producing multiple ENA monomers containing pyrimidine bases or purine bases such as A, G, T, and C, which are used as raw materials for oligonucleotides containing 2'-O,4'-C-ethylene-bridged nucleic acid (ENA), a method for stereoselectively synthesizing a β-adduct in glycosylation using the intermediate, a method for producing the production intermediate, and a method for producing an ENA monomer using the production intermediate. [Background technology]

[0002] ENA monomers are important compounds for the production of modified nucleic acid medicines and diagnostic agents.

[0003] The key steps in the production of ENAs are the 2,4-linking reaction and the glycosylation reaction to form the base skeleton.

[0004] Generally, in glycosylation reactions, when an acyloxy group is present at the 2-position, the neighboring group effect controls the stereochemistry and a β-nucleoside is obtained (see Patent Document 1). However, when no acyloxy group is present at the 2-position, such as in the case of 2,4-bridged or 2-deoxy nucleosides, it is known that an α-nucleoside is preferentially obtained (see Patent Document 2).

[0005] Therefore, when producing β-ENA, a method is used in which a base is introduced in a state in which an acyloxy group is provided at the 2-position before 2,4-ethyleneoxylation (see Patent Documents 3 and 4, and Non-Patent Documents 1 and 2).

[0006] However, the conventional method was not an industrially satisfactory production method because many intermediates, including the common intermediate before 2,4-bridge glycosylation (a compound having a hydroxyl group or an acetyloxy group at the 2-position and a protecting group -O-ethylene group at the 5-position), were oily compounds and required column purification, and the process from the common intermediate to each ENA monomer was long (see Non-Patent Document 2). [Prior art documents] [Patent documents]

[0007] [Patent Document 1] WO98 / 39352 [Patent Document 2] WO99 / 14226 [Patent Document 3] WO00 / 47599 [Patent Document 4] WO2013 / 191129 [Non-patent literature]

[0008] [Non-Patent Document 1] Bioorganic & Medicinal Chemistry Letters 12(2002) 73-76 [Non-Patent Document 2] Bioorganic & Medicinal Chemistry Letters 11(2003) 2211-2226 Summary of the Invention [Problem to be solved by the invention]

[0009] The object of the present invention is to provide a crystalline 2,4-bridged common intermediate useful for producing each ENA monomer, to provide a method for stereoselectively producing the intermediate, and to provide an efficient method for producing each ENA monomer using the intermediate. [Means for solving the problem]

[0010] As a result of intensive research into an efficient method for producing ENA monomers, the present inventors have found a novel crystalline 2,4-bridged common intermediate, a method for producing the common intermediate, and a method for producing ENA monomers using the common intermediate, and have completed the present invention. The common intermediate is crystalline and can be purified by crystallization, making it suitable for industrial production. In addition, the method for producing the common intermediate uses easily available and inexpensive raw materials and performs multiple steps without isolation, so that the common intermediate can be obtained in a few steps and in high yield. In addition, by using the common intermediate, a 2,4-bridged skeleton can be constructed before the glycosylation step, so that the steps after base introduction can be shortened and the yield can be improved, making it possible to efficiently produce multiple ENA monomers separately. Furthermore, in glycosylation using the intermediate, it has become possible to selectively produce the β-isomer by controlling the stereochemistry even without the neighboring group effect of the acyl group at the 2-position by substituting the hydroxyl group at the 1-position with an iodine atom or a bromine atom. In the amidite formation, the final step in the production of ENA monomer, the amount of amidite reagent required can be reduced by using a specific activator and desiccant, making it possible to efficiently produce ENA monomer.

[0011] That is, the present invention includes the following inventions. (1) General formula (I)

[0012] [ka]

[0013] [In the formula, Z 1 and Z 2 are the same or different and represent a protecting group for a hydroxyl group, R represents a hydrogen atom or an aliphatic acyl group, and n represents an integer of 0 to 4; (2) The compound according to (1), wherein R is a hydrogen atom or an acetyl group; (3) Z 1 and Z 2are the same or different and are an aliphatic acyl group, an aromatic acyl group, a methyl group substituted with 1 to 3 aryl groups, a methyl group substituted with 1 to 3 aryl groups in which the aryl group is substituted with a lower alkyl, a lower alkoxy, a halogen or a cyano group, or a silyl group; (4) Z 1 and Z 2 and are the same or different and are a benzyl group, a p-methoxybenzyl group, a t-butyldiphenylsilyl group, or a t-butyldimethylsilyl group; (5) Z 1 and Z 2 is a benzyl group; (6) The compound according to any one of (1) to (5), wherein n is 1; (7) A compound represented by formula (I');

[0014] [ka]

[0015] (8) A compound represented by formula (I'');

[0016] [ka]

[0017] (9a) General formula (III)

[0018] [ka]

[0019] [In the formula, Z 1 and Z 2are the same or different and represent a protecting group for a hydroxyl group, Y represents a methyl group substituted with 1 to 3 aryl groups, a methyl group substituted with 1 to 3 aryl groups in which the aryl ring is substituted with a lower alkyl, lower alkoxy, halogen or cyano group, a lower alkoxymethyl group, a tetrahydropyranyl group or a silyl group, and n represents an integer of 0 to 4. (i) General formula (XXIX)

[0020] [ka]

[0021] [wherein n is an integer of 0 to 4], (ii) the compound of general formula (XXX) obtained in step (i)

[0022] [ka]

[0023] [wherein Y and n are as defined above], (iii) the compound of general formula (XXXI) obtained in step (ii)

[0024] [ka]

[0025] [wherein Y and n are as defined above], (iv) The compound of general formula (XXXII) obtained in step (iii)

[0026] [ka]

[0027] [wherein Y and n are as defined above], (v) The compound of general formula (XXXIII) obtained in step (iv)

[0028] [ka]

[0029] [wherein Y and n are as defined above], and protecting a hydroxyl group of the compound represented by the formula: (10a)Z 1 and Z 2 and are the same or different and are an aliphatic acyl group, an aromatic acyl group, a methyl group substituted with one to three aryl groups, a methyl group substituted with one to three aryl groups in which the aryl group is substituted with a lower alkyl, a lower alkoxy, a halogen or a cyano group, or a silyl group. (11a)Z 1 and Z 2 and are the same or different and are a benzyl group, a p-methoxybenzyl group, a t-butyldiphenylsilyl group, or a t-butyldimethylsilyl group (9a). (12a)Z 1 and Z 2 is a benzyl group (9a). (13a) The method according to any one of (9a) to (12a), wherein Y is a t-butyldiphenylsilyl group, a t-butyldimethylsilyl group, a tetrahydropyran-2-yl group or a trityl group. (14a) The method according to any one of (9a) to (12a), wherein Y is a trityl group. (15a) The method according to any one of (9a) to (14a), wherein n is 1. (16a) The method according to any one of (9a) to (15a), characterized in that in each of steps (i) to (v), the organic layer of the solution containing the target compound after the reaction is washed with water, and the obtained organic layer is used as it is in each subsequent step. (9) General formula (II)

[0030] [ka]

[0031] [In the formula, Z 1 and Z 2 are the same or different and represent a protecting group for a hydroxyl group, and n represents an integer of 0 to 4. (i) General formula (III)

[0032] [ka]

[0033] [In the formula, Z 1 , Z 2 and n are as defined above, and Y is a methyl group substituted with one to three aryl groups, a methyl group substituted with one to three aryl groups in which the aryl ring is substituted with a lower alkyl, lower alkoxy, halogen or cyano group, a lower alkoxymethyl group, a tetrahydropyranyl group or a silyl group, in the presence of an acid catalyst in a lower alkyl alcohol solvent to deprotect Y; (ii) the compound of formula (IV) obtained in step (i)

[0034] [ka]

[0035] [In the formula, Z 1 , Z 2 and n are as defined above, and A is a lower alkyl group; (iii) the compound of general formula (V) obtained in step (ii)

[0036] [ka]

[0037] [In the formula, Z 1 , Z 2 wherein A and n are as defined above, A method comprising: (10) Z 1 and Z 2 are the same or different and are an aliphatic acyl group, an aromatic acyl group, a methyl group substituted with one to three aryl groups, a methyl group substituted with one to three aryl groups in which the aryl group is substituted with a lower alkyl, lower alkoxy, halogen or cyano group, or a silyl group; (11) Z 1 and Z 2 and are the same or different and are a benzyl group, a p-methoxybenzyl group, a t-butyldiphenylsilyl group, or a t-butyldimethylsilyl group; (12) Z 1 and Z 2 The method according to (9), wherein (13) The method according to any one of (9) to (12), wherein A is a methyl group, an ethyl group, or a propyl group; (14) The method according to any one of (9) to (12), wherein A is a methyl group; (15) The method according to any one of (9) to (14), wherein Y is a t-butyldiphenylsilyl group, a t-butyldimethylsilyl group, a tetrahydropyran-2-yl group, or a trityl group; (16) The method according to any one of (9) to (14), wherein Y is a trityl group; (17) The method according to any one of (9) to (16), wherein n is 1; (18) The method according to any one of (9) to (17), wherein the acid catalyst is sulfuric acid, p-toluenesulfonic acid or methanesulfonic acid; (19) The method according to any one of (9) to (18), wherein step (ii) is carried out using a trivalent phosphorus reagent and an azodicarboxylate; (20) The method according to (19), wherein the trivalent phosphorus reagent is triphenylphosphine or tri(n-butyl)phosphine; (21) The method according to (19) or (20), wherein the azodicarboxylate ester is diethyl azodicarboxylate, diisopropyl azodicarboxylate, or di-t-butyl azodicarboxylate; (22) The method according to any one of (9) to (21), wherein step (iii) is carried out using an acid; (23) The method according to (22), wherein the acid is hydrochloric acid, sulfuric acid, trifluoroacetic acid, methanesulfonic acid or p-toluenesulfonic acid; (9b)Z 1 and Z 2 is a benzyl group, A is a methyl group; Y is a trityl group; The method according to (9), wherein n is 1; (9c) The acid catalyst is sulfuric acid, p-toluenesulfonic acid or methanesulfonic acid; step (ii) is carried out using a trivalent phosphorus reagent and an azodicarboxylate; The process according to (9), wherein step (iii) is carried out using an acid; (9d) The acid catalyst is sulfuric acid, p-toluenesulfonic acid or methanesulfonic acid; step (ii) is carried out using a trivalent phosphorus reagent and an azodicarboxylate; The trivalent phosphorus reagent used in step (ii) is triphenylphosphine or tri(n-butyl)phosphine; The azodicarboxylate used in step (ii) is diethyl azodicarboxylate, diisopropyl azodicarboxylate or di-t-butyl azodicarboxylate; step (iii) is carried out with an acid; The process according to (9), wherein the acid used in step (iii) is hydrochloric acid, sulfuric acid, trifluoroacetic acid, methanesulfonic acid or p-toluenesulfonic acid; (9e)Z 1 and Z 2 is a benzyl group, A is a methyl group; Y is a trityl group; n is 1, the acid catalyst is sulfuric acid, p-toluenesulfonic acid or methanesulfonic acid; step (ii) is carried out using a trivalent phosphorus reagent and an azodicarboxylate; The trivalent phosphorus reagent used in step (ii) is triphenylphosphine or tri(n-butyl)phosphine; The azodicarboxylate used in step (ii) is diethyl azodicarboxylate, diisopropyl azodicarboxylate or di-t-butyl azodicarboxylate; step (iii) is carried out with an acid; The process according to (9), wherein the acid used in step (iii) is hydrochloric acid, sulfuric acid, trifluoroacetic acid, methanesulfonic acid or p-toluenesulfonic acid; (24) General formula (VI)

[0038] [ka]

[0039] [In the formula, R 1 represents a lower alkyl group or a hydrogen atom, R 2 represents a hydroxyl group, an amino group, or an amino group protected by an aliphatic acyl group or an aromatic acyl group; P 1 represents a trityl group optionally substituted by 1 to 3 lower alkoxy groups, and n represents an integer of 0 to 4; or a salt thereof, (i) General formula (II)

[0040] [ka]

[0041] [In the formula, Z 1 and Z 2 are the same or different and represent a protecting group for a hydroxyl group, and n represents an integer of 0 to 4.], with an activating agent in a solvent to convert the hydroxyl group at position 1 into a group that forms a leaving group; (ii) the compound of general formula (VII) obtained in step (i)

[0042] [ka]

[0043] [In the formula, Z 1 , Z 2 and n are as defined above; X 1 represents a group that forms a leaving group.

[0044] [ka]

[0045] [In the formula, R 1 and R 2 has the same meaning as defined above.] or a salt thereof is reacted in a solvent in the presence of a halogenating agent to obtain a compound represented by the general formula (IX):

[0046] [ka]

[0047] [In the formula, Z 1 , Z 2 , R 1 , R 2 and n are as defined above.] or a salt thereof, A method comprising: (25)Z 1 and Z 2 are the same or different and are an aliphatic acyl group, an aromatic acyl group, a methyl group substituted with one to three aryl groups, a methyl group substituted with one to three aryl groups in which the aryl ring is substituted with a lower alkyl, lower alkoxy, halogen or cyano group, or a silyl group; (26)Z 1 and Z 2and are the same or different and are a benzyl group, a p-methoxybenzyl group, a t-butyldiphenylsilyl group, or a t-butyldimethylsilyl group; (27)Z 1 and Z 2 The method according to (24), wherein (28)P 1 The method according to any one of (24) to (27), wherein (29)X 1 The method according to any one of (24) to (28), wherein is a halogen atom, an aliphatic acyloxy group, a halogen-substituted lower alkylimidooxy group, or a halogen-substituted lower alkylsulfonyloxy group; (30)X 1 The method according to any one of (24) to (28), wherein is an iodine atom, an acetoxy group, or a trichloroacetimidooxy group; (31) The method according to any one of (24) to (30), wherein n is 1; (32)R 1 The method according to any one of (24) to (31), wherein (33)R 2 The method according to any one of (24) to (32), wherein (34)R 1 is a methyl group, and R 2 The method according to any one of (24) to (31), wherein (35)R 1 is a methyl group, and R 2 The method according to any one of (24) to (31), wherein (36) (iii) The compound of general formula (IX) obtained in step (ii)

[0048] [ka]

[0049] [In the formula, Z 1 , Z 2, R 1 , R 2 and n are as defined above.] or a salt thereof is reacted with a deprotecting reagent for a hydroxyl group in a solvent to obtain Z 1 and Z 2 and deprotecting (iv) reacting the diol compound or a salt thereof obtained in the step (iii) with a protecting reagent for a primary hydroxyl group to obtain a compound represented by the general formula (VI)

[0050] [ka]

[0051] [In the formula, P 1 , R 1 , R 2 and n are as defined above; or a salt thereof; The method according to any one of (24) to (35), comprising: (37) The method according to any one of (24) to (36), wherein the activating agent is acetic anhydride, benzoic anhydride, trichloroacetonitrile, carbonyldiimidazole or diphenyl chlorophosphate; (38) The method according to any one of (24) to (37), wherein the halogenating agent is chlorotrimethylsilane, bromotrimethylsilane or iodotrimethylsilane; (24a)Z 1 and Z 2 is a benzyl group, P 1 is a 4,4'-dimethoxytrityl group, n is 1, R 1 is a methyl group, and R 2 is a benzoylamino group; (24b)Z 1 and Z 2 is a benzyl group, P 1 is a 4,4'-dimethoxytrityl group, X 1is an iodine atom, an acetoxy group, or a trichloroacetimidooxy group, n is 1, R 1 is a methyl group, and R 2 The method according to (24), wherein (24c) The activating agent is acetic anhydride, benzoic anhydride, trichloroacetonitrile, carbonyldiimidazole or diphenyl chlorophosphate; The method according to (24), wherein the halogenating agent is chlorotrimethylsilane, bromotrimethylsilane, or iodotrimethylsilane; (24d)Z 1 and Z 2 is a benzyl group, P 1 is a 4,4'-dimethoxytrityl group, X 1 is an iodine atom, an acetoxy group, or a trichloroacetimidooxy group, n is 1, R 1 is a methyl group, and R 2 is a benzoylamino group, the activating agent is acetic anhydride, benzoic anhydride, trichloroacetonitrile, carbonyldiimidazole or diphenyl chlorophosphate; The method according to (24), wherein the halogenating agent is chlorotrimethylsilane, bromotrimethylsilane, or iodotrimethylsilane; (39) General formula (X)

[0052] [ka]

[0053] [In the formula, R 3 represents an aliphatic acyl group or an aromatic acyl group, P 1 represents a trityl group optionally substituted by 1 to 3 lower alkoxy groups, and n represents an integer of 0 to 4; or a salt thereof, (i) General formula (II)

[0054] [ka]

[0055] [In the formula, Z 1 and Z 2 are the same or different and represent a protecting group for a hydroxyl group, and n has the same meaning as defined above, with an activating agent in a solvent to convert the hydroxyl group at 1-position into a group that forms a leaving group; (ii) the compound of formula (XI) obtained in step (i)

[0056] [ka]

[0057] [In the formula, Z 1 , Z 2 and n are as defined above; X 2 represents a group that forms a leaving group.

[0058] [ka]

[0059] [In the formula, R 3 or a salt thereof in a solvent in the presence of an acid reagent; (iii) Then, the compound represented by the general formula (XIII) is isomerized.

[0060] [ka]

[0061] [In the formula, Z 1 , Z 2 , R 3 and n are as defined above.] or a salt thereof, A method comprising: (39a) The method according to (39), wherein in step (iii), isomerization is carried out by heating; (40)Z 1 and Z 2 are the same or different and are an aliphatic acyl group, an aromatic acyl group, a methyl group substituted with one to three aryl groups, a methyl group substituted with one to three aryl groups in which the aryl ring is substituted with a lower alkyl, lower alkoxy, halogen or cyano group, or a silyl group; (41)Z 1 and Z 2 and are the same or different and are a benzyl group, a p-methoxybenzyl group, a t-butyldiphenylsilyl group, or a t-butyldimethylsilyl group; (42)Z 1 and Z 2 The method according to (39), wherein (43)P 1 The method according to any one of (39) to (42), wherein (44)X 2 The method according to any one of (39) to (43), wherein is a halogen atom, an aliphatic acyloxy group, a halogen-substituted lower alkylimidooxy group, or a halogen-substituted lower alkylsulfonyloxy group; (45)X 2 The method according to any one of (39) to (43), wherein (46) The method according to any one of (39) to (45), wherein n is 1; (47)R 3 The method according to any one of (39) to (46), wherein (48)R 3 The method according to any one of (39) to (46), wherein (49) (iv) The compound of general formula (XIII) obtained in step (iii)

[0062] [ka]

[0063] [In the formula, Z 1 , Z 2 , R 3 and n are as defined above.] or a salt thereof is reacted with a deprotecting reagent for a hydroxyl group in a solvent to obtain Z 1 and Z 2 and deprotecting (v) The diol compound or a salt thereof obtained in the step (iv) is reacted with a protecting reagent for a primary hydroxyl group to selectively protect the primary hydroxyl group, thereby obtaining a compound represented by the general formula (X):

[0064] [ka]

[0065] [In the formula, R 3 , n and P 1 has the same meaning as defined above] or a salt thereof; The method according to any one of (39) to (48), comprising: (50) The method according to any one of (39) to (49), wherein the activating agent is acetic anhydride, benzoic anhydride, trichloroacetonitrile, carbonyldiimidazole or diphenyl chlorophosphate; (51) The method according to any one of (39) to (50), wherein the acid reagent is trimethylsilyl trifluoromethanesulfonate and trifluoroacetic acid; (39b)Z 1 and Z 2 is a benzyl group, P 1 is a 4,4'-dimethoxytrityl group, n is 1, R 3 is a benzoyl group; (39c)Z 1 and Z 2 is a benzyl group, P 1 is a 4,4'-dimethoxytrityl group, X 2is an acetoxy group n is 1, R 3 is a benzoyl group; (39d) The activating agent is acetic anhydride, benzoic anhydride, trichloroacetonitrile, carbonyldiimidazole or diphenyl chlorophosphate; The method according to (39), wherein the acid reagent is trimethylsilyl trifluoromethanesulfonate and trifluoroacetic acid; (39e)Z 1 and Z 2 is a benzyl group, P 1 is a 4,4'-dimethoxytrityl group, X 2 is an acetoxy group n is 1, R 3 is a benzoyl group, the activating agent is acetic anhydride, benzoic anhydride, trichloroacetonitrile, carbonyldiimidazole or diphenyl chlorophosphate; The method according to (39), wherein the acid reagent is trimethylsilyl trifluoromethanesulfonate and trifluoroacetic acid; (52) General formula (XIV)

[0066] [ka]

[0067] [In the formula, Z 1 and Z 2 are the same or different and represent a protecting group for a hydroxyl group, and n represents an integer of 0 to 4, or a salt thereof; (53)Z 1 and Z 2 are the same or different and are an aliphatic acyl group, an aromatic acyl group, a methyl group substituted with 1 to 3 aryl groups, a methyl group substituted with 1 to 3 aryl groups in which the aryl ring is substituted with a lower alkyl, a lower alkoxy, a halogen or a cyano group, or a silyl group; (54)Z 1 and Z 2 and are the same or different and are a benzyl group, a p-methoxybenzyl group, a t-butyldiphenylsilyl group, or a t-butyldimethylsilyl group; (55)Z 1 and Z 2 is a benzyl group; (56) The compound or salt thereof according to any one of (52) to (55), wherein n is 1; (57) A compound represented by general formula (XIV') or a salt thereof;

[0068] [ka]

[0069] (58) General formula (X)

[0070] [ka]

[0071] [In the formula, P 1 represents a trityl group optionally substituted by one to three lower alkoxy groups; R 3 represents an aliphatic acyl group or an aromatic acyl group, and n represents an integer of 1 to 4, or a salt thereof, (i) General formula (XIV)

[0072] [ka]

[0073] [In the formula, Z 1 and Z 2 are the same or different and represent a hydroxyl-protecting group, and n represents an integer of 0 to 4; or a salt thereof, with an aminating agent to substitute the chlorine atom at position 6 of the purine ring with an amino group; (ii) The compound of formula (XV) obtained in step (i)

[0074] [ka]

[0075] [In the formula, Z 1 , Z 2 and n are as defined above.] or a salt thereof is reacted with a reducing agent in a solvent in the presence of a metal catalyst to substitute a hydrogen atom for the chlorine atom at the 2-position of the purine ring, thereby obtaining Z 1 and Z 2 By deprotecting the compound represented by the general formula (XVI)

[0076] [ka]

[0077] [wherein n is as defined above] or a salt thereof; A method comprising: (58b) The method according to (58), wherein the metal catalyst is a carbon-supported metal catalyst; (59)Z 1 and Z 2 are the same or different and are an aliphatic acyl group, an aromatic acyl group, a methyl group substituted with one to three aryl groups, a methyl group substituted with one to three aryl groups in which the aryl ring is substituted with a lower alkyl, lower alkoxy, halogen or cyano group, or a silyl group. method ; (60)Z 1 and Z 2 and are the same or different and are a benzyl group, a p-methoxybenzyl group, a t-butyldiphenylsilyl group, or a t-butyldimethylsilyl group; (61)Z 1 and Z 2 The method according to (58), wherein (62)P 1The method according to any one of (58) to (61), wherein (63) The method according to any one of (58) to (62), wherein n is 1; (64)R 3 The method according to any one of (58) to (63), wherein (65)R 3 The method according to any one of (58) to (63), wherein (66) (iii) The compound of general formula (XVI) obtained in step (ii)

[0078] [ka]

[0079] [wherein n is as defined above] or a salt thereof is reacted with a protecting reagent for a primary hydroxyl group to selectively protect the primary hydroxyl group; (iv) the compound of general formula (XVII) obtained in step (iii)

[0080] [ka]

[0081] [In the formula, P 1 and n are as defined above.] or a salt thereof is reacted with an acylating agent to obtain a compound represented by the general formula (X):

[0082] [ka]

[0083] [In the formula, P 1 , R 3 and n are as defined above; or a salt thereof; The method according to any one of (58) to (65), comprising: (67) The method according to any one of (58) to (66), wherein the aminating agent is ammonia, an aqueous ammonia solution, ammonium carbonate or ammonium acetate. (68) The method according to any one of (58) to (67), wherein the metal catalyst is palladium, palladium hydroxide or platinum; (69) The method according to any one of (58) to (68), wherein the reducing agent is hydrogen, formic acid or ammonium formate; (70) The method according to any one of (66) to (69), wherein the acylating agent is benzoyl chloride or benzoic anhydride; (58a) (iii-a) The compound of formula (XVI) obtained in step (ii) of (58)

[0084] [ka]

[0085] [wherein n is as defined above] or a salt thereof is reacted with a protecting reagent for a primary hydroxyl group to selectively protect the primary hydroxyl group; (iv-a) The compound of general formula (XVII) obtained in step (iii-a)

[0086] [ka]

[0087] [In the formula, P 1 and n are as defined above.] or a salt thereof is reacted with an acylating agent to obtain a compound represented by the general formula (X):

[0088] [ka]

[0089] [In the formula, P 1 , R 3 and n are as defined above. Z 1and Z 2 is a benzyl group, P 1 is a 4,4'-dimethoxytrityl group, n is 1, R 3 is a benzoyl group, the aminating agent is ammonia, an aqueous ammonia solution, ammonium carbonate or ammonium acetate; the metal catalyst is palladium, palladium hydroxide or platinum; the reducing agent is hydrogen, formic acid or ammonium formate; The method according to (58), wherein the acylating agent is benzoyl chloride or benzoic anhydride; (71) General formula (XVIII)

[0090] [ka]

[0091] [In the formula, P 1 represents a trityl group optionally substituted by one to three lower alkoxy groups; R 4 represents an aliphatic acyl group or an aromatic acyl group, and n represents an integer of 1 to 4; or a salt thereof, (i) General formula (XIV)

[0092] [ka]

[0093] [In the formula, Z 1 and Z 2 are the same or different and represent a protecting group for a hydroxyl group, and n represents an integer of 0 to 4.] or a salt thereof is reacted with benzyl alcohol which may be substituted with a lower alkyl, lower alkoxy, halogen or cyano group in a solvent in the presence of a base, to substitute the chlorine atom at the 6-position of the purine ring with a benzyloxy group which may be substituted with a lower alkyl, lower alkoxy, halogen or cyano group; (ii) the compound of formula (XIX) obtained in step (i)

[0094] [ka]

[0095] [In the formula, Z 1 , Z 2 and n are as defined above; R 5 represents a benzyl group which may be substituted with a lower alkyl, a lower alkoxy, a halogen or a cyano group.] or a salt thereof is cross-coupled with an amidating agent in a solvent in the presence of a palladium catalyst and a phosphine ligand to obtain a compound represented by the general formula (XX):

[0096] [ka]

[0097] [In the formula, Z 1 , Z 2 , R 4 , R 5 and n are as defined above; or a salt thereof; A method comprising: (72)Z 1 and Z 2 are the same or different and are an aliphatic acyl group, an aromatic acyl group, a methyl group substituted with one to three aryl groups, a methyl group substituted with one to three aryl groups in which the aryl ring is substituted with a lower alkyl, lower alkoxy, halogen or cyano group, or a silyl group; (73)Z 1 and Z 2 and are the same or different and are a benzyl group, a p-methoxybenzyl group, a t-butyldiphenylsilyl group, or a t-butyldimethylsilyl group; (74)Z 1 and Z 2 The method according to (71), wherein (75)P 1The method according to any one of (71) to (74), wherein (76) The method according to any one of (71) to (75), wherein n is 1; (77)R 4 The method according to any one of (71) to (76), wherein (78)R 5 The method according to any one of (71) to (77), wherein (79) (iii) The compound of formula (XX) obtained in step (ii)

[0098] [ka]

[0099] [In the formula, Z 1 , Z 2 , R 4 , R 5 and n are as defined above.] or a salt thereof is reacted with a deprotecting reagent for a hydroxyl group in a solvent to obtain Z 1 , Z 2 and R 5 Deprotecting the compound (iv) The compound of general formula (XXI) obtained in step (iii)

[0100] [ka]

[0101] [In the formula, R 4 and n are as defined above.] or a salt thereof is reacted with a protecting reagent for a primary hydroxyl group to selectively protect the primary hydroxyl group, thereby obtaining a compound represented by the general formula (XVIII):

[0102] [ka]

[0103] [In the formula, P 1 , R 4and n are as defined above; or a salt thereof; The method according to any one of (71) to (78), comprising: (80) The method according to any one of (71) to (79), wherein the base is sodium hydroxide, sodium carbonate, cesium carbonate, triethylamine, pyridine or 1,8-diazabicyclo[5.4.0]undec-7-ene; (81) The method according to any one of (71) to (80), wherein the palladium catalyst is tris(dibenzylideneacetone)(chloroform)dipalladium, palladium(II) acetate or tris(dibenzylideneacetone)dipalladium(0); (82) The method according to any one of (71) to (81), wherein the phosphine ligand is 4,5'-bis(diphenylphosphino)-9,9'-dimethylxanthene, 1,1'-bis(diphenylphosphino)ferrocene, 1,2-bis(diphenylphosphino)ethane or 2-dicyclohexylphosphino-2'-(N,N-dimethylamino)biphenyl; (83) The method according to any one of (71) to (82), wherein the amidating agent is acetylamide, benzoylamide or isobutyramide; (84) The method according to any one of (79) to (83), wherein the deprotection reagent for a hydroxyl group is a metal catalyst and a reducing agent; (84a) The method according to any one of (79) to (83), wherein the deprotection reagent for the hydroxyl group is a carbon-supported metal catalyst and hydrogen; (85) The method according to (84), wherein the metal catalyst is palladium, palladium hydroxide or platinum; (86) The method according to (84) or (85), wherein the reducing agent is hydrogen, formic acid or ammonium formate; (71a) (iii-a) The compound of formula (XX) obtained in step (ii) of (71)

[0104] [ka]

[0105] [In the formula, Z 1 , Z 2 , R 4 , R 5 and n are as defined above.] or a salt thereof is reacted with a deprotecting reagent for a hydroxyl group in a solvent to obtain Z 1 , Z 2 and R 5 Deprotecting the compound (iv-a) The compound of formula (XXI) obtained in step (iii-a)

[0106] [ka]

[0107] [In the formula, R 4 and n are as defined above.] or a salt thereof is reacted with a protecting reagent for a primary hydroxyl group to selectively protect the primary hydroxyl group, thereby obtaining a compound represented by the general formula (XVIII):

[0108] [ka]

[0109] [In the formula, P 1 , R 4 and n are as defined above; or a salt thereof, Z 1 and Z 2 is a benzyl group, P 1 is a 4,4'-dimethoxytrityl group, n is 1, R 4 is an isobutyryl group, the base is sodium hydroxide, sodium carbonate, cesium carbonate, triethylamine, pyridine or 1,8-diazabicyclo[5.4.0]undec-7-ene; the palladium catalyst is tris(dibenzylideneacetone)(chloroform)dipalladium, palladium(II) acetate or tris(dibenzylideneacetone)dipalladium(0); the phosphine ligand is 4,5'-bis(diphenylphosphino)-9,9'-dimethylxanthene, 1,1'-bis(diphenylphosphino)ferrocene, 1,2-bis(diphenylphosphino)ethane or 2-dicyclohexylphosphino-2'-(N,N-dimethylamino)biphenyl; the amidating agent is acetylamide, benzoylamide or isobutyramide; the deprotection reagent for the hydroxyl group is a carbon-supported metal catalyst and hydrogen; The method according to (71), wherein the metal catalyst is palladium, palladium hydroxide or platinum; (87) General formula (XXII)

[0110] [ka]

[0111] [In the formula, R 1 represents a lower alkyl group or a hydrogen atom, R 6 represents an aliphatic acyl group or an aromatic acyl group, P 1 represents a trityl group optionally substituted by 1 to 3 lower alkoxy groups, and n represents an integer of 0 to 4; or a salt thereof, (i) General formula (XXIII)

[0112] [ka]

[0113] [In the formula, P 1 , R 1 and n are as defined above.] or a salt thereof is reacted with a hydroxyl-protecting reagent in a solvent to protect the hydroxyl group at the 3'-position; (ii) The compound of formula (XXIV) obtained in step (i)

[0114] [ka]

[0115] [In the formula, P 1 , R 1 and n are as defined above; Z 3 represents an aliphatic acyl group or an aromatic acyl group, or a salt thereof, with an activating agent in a solvent in the presence of a base and a catalyst; (iii) Then, by reacting with an aminating agent, a compound represented by the general formula (XXV)

[0116] [ka]

[0117] [In the formula, P 1 , R 1 , Z 3 and n are as defined above; or a salt thereof; A method comprising: (88)P 1 The method according to (87), wherein (89)Z 3 The method according to (87) or (88), wherein (90) The method according to any one of (87) to (89), wherein n is 1; (91)R 1 The method according to any one of (87) to (90), wherein (92)R 6 The method according to any one of (87) to (91), wherein (93)R 6 The method according to any one of (87) to (91), wherein (94) (iv) The compound of general formula (XXV) obtained in step (iii)

[0118] [ka]

[0119] [In the formula, P 1, R 1 , Z 3 and n are as defined above.] or a salt thereof is reacted with an acylating agent in a solvent; (v) The compound of general formula (XXVI) obtained in step (iv)

[0120] [ka]

[0121] [In the formula, P 1 , R 1 , R 6 , Z 3 and n are as defined above.] or a salt thereof is reacted with a deprotecting reagent for a hydroxyl group to obtain Z 3 By deprotecting only the compound represented by the general formula (XXII),

[0122] [ka]

[0123] [In the formula, P 1 , R 1 , R 6 and n are as defined above; or a salt thereof; The method according to any one of (87) to (93), comprising: (95) The method according to any one of (87) to (94), wherein the catalyst is N,N-dimethylaminopyridine or 1,8-diazabicyclo[5.4.0]undec-7-ene; (96) The method according to any one of (87) to (95), wherein the activating agent is p-toluenesulfonyl chloride or 2,4,6-triisopropylbenzenesulfonyl chloride; (97) The method according to any one of (87) to (96), wherein the aminating agent is ammonia, an aqueous ammonia solution, ammonium carbonate or ammonium acetate; (98) The method according to any one of (94) to (97), wherein the acylating agent is benzoyl chloride or benzoic anhydride; (87a)P 1 is a trityl group, Z 3 is an acetyl group, n is 1, R 1 is a methyl group or a hydrogen atom R 6 The method according to (87), wherein (87b) (iv-b) The compound of general formula (XXV) obtained in step (iii) of (87)

[0124] [ka]

[0125] [In the formula, P 1 , R 1 , Z 3 and n are as defined above.] or a salt thereof is reacted with an acylating agent in a solvent; (vb) the compound of general formula (XXVI) obtained in step (iv-b)

[0126] [ka]

[0127] [In the formula, P 1 , R 1 , R 6 , Z 3 and n are as defined above.] or a salt thereof is reacted with a deprotecting reagent for a hydroxyl group to obtain Z 3 By deprotecting only the compound represented by the general formula (XXII),

[0128] [ka]

[0129] [In the formula, P 1 , R 1 , R 6 and n are as defined above; or a salt thereof, the catalyst is N,N-dimethylaminopyridine or 1,8-diazabicyclo[5.4.0]undec-7-ene; the activating agent is p-toluenesulfonyl chloride or 2,4,6-triisopropylbenzenesulfonyl chloride; the aminating agent is ammonia, an aqueous ammonia solution, ammonium carbonate or ammonium acetate; The method according to (87), wherein the acylating agent is benzoyl chloride or benzoic anhydride; (87c) (iv-c) (87)

[0130] [ka]

[0131] [In the formula, P 1 , R 1 , Z 3 and n are as defined above.] or a salt thereof is reacted with an acylating agent in a solvent; (vc) the compound of general formula (XXVI) obtained in step (iv-c)

[0132] [ka]

[0133] [In the formula, P 1 , R 1 , R 6 , Z 3 and n are as defined above.] or a salt thereof is reacted with a deprotecting reagent for a hydroxyl group to obtain Z 3 By deprotecting only the compound represented by the general formula (XXII),

[0134] [ka]

[0135] [In the formula, P 1 , R1 , R 6 and n are as defined above; or a salt thereof, P 1 is a trityl group, Z 3 is an acetyl group, n is 1, R 1 is a methyl group or a hydrogen atom R 6 is a benzoyl group, the catalyst is N,N-dimethylaminopyridine or 1,8-diazabicyclo[5.4.0]undec-7-ene; the activating agent is p-toluenesulfonyl chloride or 2,4,6-triisopropylbenzenesulfonyl chloride; the aminating agent is ammonia, an aqueous ammonia solution, ammonium carbonate or ammonium acetate; The method according to (87), wherein the acylating agent is benzoyl chloride or benzoic anhydride; (99) General formula (XXVII)

[0136] [ka]

[0137] [In the formula, P 1 represents a trityl group which may be substituted by 1 to 3 lower alkoxy groups, B represents a 2-oxo-pyrimidin-1-yl group or a purin-9-yl group which may have one or more substituents selected from the following α group, and n represents an integer of 0 to 4.] or a salt thereof is reacted with an amidite reagent in a solvent in the presence of an activator and a desiccant, General formula (XXVIII)

[0138] [ka]

[0139] [In the formula, P 1and B and n are as defined above.] or a salt thereof; (α group) Hydroxyl group, A protected hydroxyl group, lower alkoxy group, Mercapto group, A protected mercapto group, lower alkylthio group, Amino group, A protected amino group, A lower alkylamino group, A lower alkyl group, and Halogen atoms; (100)P 1 The method according to (99), wherein (101) The method according to (99) or (100), wherein B is a 2-oxo-4-hydroxy-5-methylpyrimidin-1-yl group, a 2-oxo-4-amino-pyrimidin-1-yl group with a protected amino group, a 4-amino-5-methyl-2-oxo-pyrimidin-1-yl group with a protected amino group, a 6-aminopurin-9-yl group with a protected amino group, or a 2-amino-6-hydroxypurin-9-yl group with a protected amino group; (102) The method according to (99) or (100), wherein B is a 2-oxo-4-hydroxy-5-methylpyrimidin-1-yl group, a 2-oxo-4-benzoylamino-pyrimidin-1-yl group, a 4-benzoylamino-5-methyl-2-oxo-pyrimidin-1-yl group, a 6-benzoylaminopurin-9-yl group, or a 2-isobutyrylamino-6-hydroxypurin-9-yl group; (103) The method according to any one of (99) to (102), wherein n is 1; (104) The method according to any one of (99) to (103), wherein the activating agent is pyridine trifluoroacetate, N-methylimidazole trifluoroacetate, N-isopropylimidazole trifluoroacetate, 5-benzylthiotetrazole, 5-phenyltetrazole, 4,5-dicyanoimidazole, or 2,4,5-tetrabromoimidazole; (105) The method according to any one of (99) to (103), wherein the activating agent is 4,5-dicyanoimidazole; (106) The method according to any one of (99) to (105), wherein the amiditizing reagent is 2-cyanoethyl N,N,N',N'-tetraisopropyl phosphorodiamidite or 2-cyanoethyl diisopropyl chlorophosphoroamidite; (107) The method according to any one of (99) to (105), wherein the amiditizing reagent is 2-cyanoethyl N,N,N',N'-tetraisopropylphosphorodiamidite; (108) The method according to any one of (99) to (107), wherein the desiccant is molecular sieve 3A, molecular sieve 4A, or molecular sieve 5A; (99a)P 1 is a 4,4'-dimethoxytrityl group, B is a 2-oxo-4-hydroxy-5-methylpyrimidin-1-yl group, a 2-oxo-4-benzoylamino-pyrimidin-1-yl group, a 4-benzoylamino-5-methyl-2-oxo-pyrimidin-1-yl group, a 6-benzoylaminopurin-9-yl group, or a 2-isobutyrylamino-6-hydroxypurin-9-yl group, The method according to (99), wherein n is 1; (99b) The activating agent is pyridine trifluoroacetate, N-methylimidazole trifluoroacetate, N-isopropylimidazole trifluoroacetate, 5-benzylthiotetrazole, 5-phenyltetrazole, 4,5-dicyanoimidazole or 2,4,5-tetrabromoimidazole; the amiditizing reagent is 2-cyanoethyl N,N,N',N'-tetraisopropyl phosphorodiamidite or 2-cyanoethyl diisopropyl chlorophosphoroamidite; The method according to (99), wherein the desiccant is molecular sieve 3A, molecular sieve 4A, or molecular sieve 5A; (99c)P 1 is a 4,4'-dimethoxytrityl group, B is a 2-oxo-4-hydroxy-5-methylpyrimidin-1-yl group, a 2-oxo-4-benzoylamino-pyrimidin-1-yl group, a 4-benzoylamino-5-methyl-2-oxo-pyrimidin-1-yl group, a 6-benzoylaminopurin-9-yl group, or a 2-isobutyrylamino-6-hydroxypurin-9-yl group, n is 1, the activating agent is pyridine trifluoroacetate, N-methylimidazole trifluoroacetate, N-isopropylimidazole trifluoroacetate, 5-benzylthiotetrazole, 5-phenyltetrazole, 4,5-dicyanoimidazole or 2,4,5-tetrabromoimidazole; the amiditizing reagent is 2-cyanoethyl N,N,N',N'-tetraisopropyl phosphorodiamidite or 2-cyanoethyl diisopropyl chlorophosphoroamidite; The method according to (99), wherein the desiccant is molecular sieve 3A, molecular sieve 4A, or molecular sieve 5A; (109) A method for producing an oligonucleotide, comprising the steps of: (A) synthesizing an ENA monomer by the method according to any one of (99) to (108); and (B) A step of extending a nucleotide chain according to a desired sequence using the ENA monomer obtained in step (A), a phosphoramidite compound of another nucleic acid, and / or a phosphoramidite compound of a ligand. (110) The method according to (109), wherein the oligonucleotide has a sequence represented by any one of the formulas selected from the following DMD AO01 to DMD AO15. (DMD AO01) HO-C e2s -A m1s -G m1s -T e2s -T e2s -U m1s -G m1s -C e2s -C e2s -G m1s -C e2s -T e2s -G m1s-C e2s -C e2s -C e2s -A m1s -A m1s -CH2CH2OH (SEQ ID NO: 1) (DMD AO02) HO-T e2s -G m1s -T e2s -T e2s -C e2s -T e2s -G m1s -A m1s -C e2s -A m1s -A m1s -C e2s -A m1s -G m1s -T e2s -T e2s -T e2s -G m1s -CH2CH2OH (SEQ ID NO: 2) (DMD AO03) HO-C e2s -G m1s -C e2s -T e2s -G m1s -C m1s -C e2s -C e2s -A m1s -A m1s -T e2s -G m1s -C e2s -C e2s -A m1s -U m1s -C e2s -C e2s -CH2CH2OH (SEQ ID NO: 3) (DMD AO04) HO-C e2s -A m1s -T e2s -A m1s -A m1s -T e2s -G m1s -A m1s -A e2s -A m1s -A m1s -C e2s -G m1s -C m1s -C e2s -G m1s -C e2s -Ce2s -CH2CH2OH (array number 4) (DMD AO05) HO-T e2s -U m1s -C e2s -C m1s -C e2s -A m1s -A m1s -T e2s -U m1s -C m1s -T e2s -C e2s -A m1s -G m1s -G m1s -A e2s -A m1s -T e2s -CH2CH2OH (array number 5) (DMD AO06) HO-C e2s -C e2s -A m1s -U m1s -T e2s -U m1s -G m1s -T e2s -A m1s -U m1s -T e2s -T e2s -A m1s -G m1s -C e2s -A m1s -T e2s -G m1s -CH2CH2OH (array number 6) (DMD AO07) HO-G m1s -G m1s -C e2s -T e2s -G m1s -C m1s -T e2s -T e2s -U m1s -G m1s -C e2s -C m1s -C m1s -T e2s -C e2s -A m1s -G m1s -C e2s -CH2CH2OH (array number 7) (DMD AO08) HO-Gm1s -C e2s -T e2s -A m1s -G m1s -G m1s -T e2s -C e2s -A m1s -G m1s -G m1s -T e2s -G m1s -C m1s -T e2s -T e2s -U m1s -CH2CH2OH (SEQ ID NO: 8) (DMD AO09) HO-A m1s -C e2s -C e2s -G m1s -C m1s -C e2s -T e2s -U m1s -C m1s -C e2s -A m1s -C m1s -T e2s -C e2s -A m1s -G m1s -A e2s -G m1s -CH2CH2OH; (SEQ ID NO: 9) (DMD AO10) HO-G e2s -G e2s -C e2s -A e2s -T e2s -U m1s -U m1s -C m1s -U m1s -A m1s -G m1s -U m1s -U m1s -T e2s -G e2s -G e2s -A e2s -G e2s -CH2CH2OH (SEQ ID NO: 10) (DMD AO11) HO-G m1s -G m1s -C e2s -A m1s -T e2s -Te2s -U m1s -C e2s -T e2s -A m1s -G m1s -U m1s -T e2s -T e2s -G m1s -G m1s -A e2s -G m1s -CH2CH2OH (SEQ ID NO: 11) (DMD AO12) HO-A e2s -G m1s -T e2s -U m1s -T e2s -G m1s -G m1s -A e2s -G m1s -A m1s -T e2s -G m1s -G m1s -C e2s -A e2s -G m1s -T e2s -T e2s -CH2CH2OH (SEQ ID NO: 12) (DMD AO13) HO-C e2s -T e2s -C m1s -C e2s -T e2s -U m1s -C e2s -C e2s -A m1s -T e2s -G m1s -A m1s -C e2s -T e2s -C e2s -A m1s -A m1s -G m1s - CH2CH2OH (SEQ ID NO: 13) (DMD AO14) HO-C e2s -T e2s -G m1s -A m1s -A m1s -G m1s -G m1s -T e2s -Gm1s -T e2s -T e2s -C e2s -T e2s -T e2s -G m1s -T e2s -A m1s - C e2s - CH2CH2OH (SEQ ID NO: 14) (DMD AO15) HO-T e2s -T e2s -C m1s -C e2s -A m1s -G m1s -C e2s -C e2s -A m1s -T e2s -T e2s -G m1s -T e2s -G m1s -T e2s -T e2s -G m1s -A m1s - CH2CH2OH (SEQ ID NO: 15) [In the above formula, the left side represents the 5' end and the right side represents the 3' end, and A, G, C, U and T represent adenosine, guanosine, cytidine, uridine and thymidine, respectively, in which D-ribofuranose has been modified and the carbon atom at the 5' position is linked via a phosphorothioate bond to the structural unit shown on the left side. The e2s attached to each nucleotide or nucleoside represents that the D-ribofuranose is 2'-O,4'-C-ethylene bridged and the 3' position is -OP(=S)(-OH)-O- bonded to the 5' carbon atom of the adjacent nucleotide or nucleoside on the right side, the e2t represents that the D-ribofuranose is 2'-O,4'-C-ethylene bridged and the 3' position is -O- bonded to the hydrogen atom of the 3' end, the m1s represents that the D-ribofuranose is 2'-O-methylated and the 3' position is -OP(=S)(-OH)-O- bonded to the 5' carbon atom of the adjacent nucleotide or nucleoside on the right side, and the m1t represents that the D-ribofuranose is 2'-O-methylated and the 3' position is -O- bonded to the hydrogen atom of the 3' end. (111) The oligonucleotide has a sequence represented by any one of the formulas selected from GSD AO01 to GSD AO16 below, and the ligand is X 18 Or X 20 The method according to (109), characterized in that (GSD AO01) X 18 -A m1s -A e2s -U m1s -C m1s -C e2s -G m1s -A m1s -T e2s -G m1s -G m1s -C e2s -G m1s -A m1s -A e2s -G m1t -H (SEQ ID NO: 16) (GSD AO02) X 18 -A e2s -U m1s -C m1s -C e2s -G m1s -A m1s -T e2s -G m1s -G m1s -C e2s -G m1s -A m1s -A e2s -G m1s -C e2t -H (SEQ ID NO: 17) (GSD AO03) X 18 -U m1s -C m1s -C e2s -G m1s -A m1s -T e2s -G m1s -G m1s -C e2s -G m1s -A m1s -A e2s -G m1s -C e2s -U m1t -H (SEQ ID NO: 18) (GSD AO04) X 18 -A m1s-A m1s -A e2s -U m1s -C m1s -C e2s -G m1s -A m1s -T e2s -G m1s -G m1s -C e2s -G m1s -A m1s -A e2s -G m1t -H(SEQ ID NO: 19) (GSD AO05) X 18 -A m1s -A e2s -U m1s -C m1s -C e2s -G m1s -A m1s -T e2s -G m1s -G m1s -C e2s -G m1s -A m1s -A e2s -G m1s -C e2t -H(SEQ ID NO: 20) (GSD AO06) X 18 -A e2s -U m1s -C m1s -C e2s -G m1s -A m1s -T e2s -G m1s -G m1s -C e2s -G m1s -A m1s -A e2s -G m1s -C e2s -U m1t -H(SEQ ID NO: 21) (GSD AO07) X 18 -A m1s -A e2s -U m1s -C m1s -C e2s -G m1s -A e2s -U m1s -G m1s -G m1s -Ce2s -G m1s -A m1s -A e2s -G m1t -H (SEQ ID NO: 22) (GSD AO08) X 18 -A e2s -U m1s -C m1s -C e2s -G m1s -A e2s -U m1s -G m1s -G m1s -C e2s -G m1s -A m1s -A e2s -G m1s -C e2t -H (SEQ ID NO: 23) (GSD AO09) X 18 -U m1s -C m1s -C e2s -G m1s -A e2s -U m1s -G m1s -G m1s -C e2s -G m1s -A m1s -A e2s -G m1s -C e2s -U m1t -H (SEQ ID NO: 24) (GSD AO10) X 18 -A m1s -A m1s -A e2s -U m1s -C m1s -C e2s -G m1s -A e2s -U m1s -G m1s -G m1s -C e2s -G m1s -A m1s -A e2s -G m1t -H (SEQ ID NO: 25) (GSD AO11) X 18 -A m1s -A e2s -U m1s -Cm1s -C e2s -G m1s -A e2s -U m1s -G m1s -G m1s -C e2s -G m1s -A m1s -A e2s -G m1s -C e2t -H1 (SEQ ID NO: 26) (GSD AO12) X 18 -A e2s -U m1s -C m1s -C e2s -G m1s -A e2s -U m1s -G m1s -G m1s -C e2s -G m1s -A m1s -A e2s -G m1s -C e2s -U m1t -H (SEQ ID NO: 27) (GSD AO13) X 20 -A m1s -A e2s -U m1s -C m1s -C e2s -G m1s -A e2s -U m1s -G m1s -G m1s -C e2s -G m1s -A m1s -A e2s -G m1t -H (SEQ ID NO: 28) (GSD AO14) X 20 -A e2s -U m1s -C m1s -C e2s -G m1s -A e2s -U m1s -G m1s -G m1s -C e2s -G m1s -A m1s -A e2s -Gm1s -C e2t -H (SEQ ID NO: 29) (GSD AO15) X 20 -A m1s -A m1s -A e2s -U m1s -C m1s -C e2s -G m1s -A e2s -U m1s -G m1s -G m1s -C e2s -G m1s -A m1s -A e2s -G m1t -H (SEQ ID NO: 30) (GSD AO16) X 20 -A m1s -A e2s -U m1s -C m1s -C e2s -G m1s -A e2s -U m1s -G m1s -G m1s -C e2s -G m1s -A m1s -A e2s -G m1s -C e2t -H (SEQ ID NO: 31) [In the above formula, the left side represents the 5' end and the right side represents the 3' end, and A, G, C, U and T represent adenosine, guanosine, cytidine, uridine and thymidine, respectively, in which D-ribofuranose has been modified and the carbon atom at the 5' position is linked via a phosphorothioate bond to the structural unit shown on the left side. e2s attached to each nucleotide or nucleoside represents that D-ribofuranose is 2'-O,4'-C-ethylene bridged and the 3' position is -OP(=S)(-OH)-O- bonded to the 5' carbon atom of the adjacent nucleotide or nucleoside on the right side, e2t represents that D-ribofuranose is 2'-O,4'-C-ethylene bridged and the 3' position is -O- bonded to the hydrogen atom at the 3' end, m1s represents that D-ribofuranose is 2'-O-methylated and the 3' position is -OP(=S)(-OH)-O- bonded to the 5' carbon atom of the adjacent nucleotide or nucleoside on the right side, and m1t represents that D-ribofuranose is 2'-O-methylated and the 3' position is -O- bonded to the hydrogen atom at the 3' end. In the above formula, X 18 and X 20 represents a GalNAc unit represented by the following formula, in which the bond bonded to the phosphate group is bonded to the carbon atom at the 5' end of the oligonucleotide to form a phosphodiester bond.

[0140] [ka]

[0141] [ka] Effect of the Invention

[0142] The present invention makes it possible to efficiently produce multiple ENA monomers separately. DETAILED DESCRIPTION OF THE PREFERRED EMBODIMENTS

[0143] The present invention will now be described in further detail.

[0144] In the present invention, Z 1 and Z 2 The "hydroxyl group-protecting group" of the group 1 and the "protected hydroxyl group" of the group α refer to a protecting group that can be cleaved by a chemical method such as hydrogenolysis, hydrolysis, electrolysis, and photolysis, or a biological method such as hydrolysis in the human body. Examples of such protecting groups include formyl, acetyl, propionyl, butyryl, isobutyryl, pentanoyl, pivaloyl, valeryl, isovaleryl, octanoyl, nonanoyl, decanoyl, 3-methylnonanoyl, 8-methylnonanoyl, 3-ethyloctanoyl, 3,7-dimethyloctanoyl, undecanoyl, dodecanoyl, tridecanoyl, tetradecanoyl, pentadecanoyl, hexadecanoyl, 1-methylpentadecanoyl, 14-methylpentadecanoyl, 13,13-dimethyltetradecanoyl, heptadecanoyl, 15-methyloctanoyl, 16-methyloctanoyl, 17-methyloctanoyl, 18-methyloctanoyl, 21-methyloctanoyl, 22-methyloctanoyl, 23-methyloctanoyl, 24-methyloctanoyl, 25-methyloctanoyl, 26-methyloctanoyl, 27-methyloctanoyl, 28-methyloctanoyl, 29-methyloctanoyl, 30-methyloctanoyl, 31-methyloctanoyl, 32-methyloctanoyl, 33-methyloctanoyl, 34-methyloctanoyl, 35-methyloctanoyl, 36-methyloctanoyl, 37-methyloctanoyl, 38-methyloctanoyl, 39-methyloctanoyl, 40-methyloctanoyl, 41-methyloctanoyl, 42-methyloctanoyl, 43- "fatty acyl groups" such as alkylcarbonyl groups such as hexadecanoyl, octadecanoyl, 1-methylheptadecanoyl, nonadecanoyl, eicosanoyl, and henaicosanoyl, carboxylated alkylcarbonyl groups such as succinoyl, glutaroyl, and adipoyl, halogeno lower alkylcarbonyl groups such as chloroacetyl, dichloroacetyl, trichloroacetyl, and trifluoroacetyl, lower alkoxy lower alkylcarbonyl groups such as methoxyacetyl, and unsaturated alkylcarbonyl groups such as (E)-2-methyl-2-butenoyl; "aromatic acyl groups" such as arylcarbonyl groups such as benzoyl, α-naphthoyl, β-naphthoyl, halogenoarylcarbonyl groups such as 2-bromobenzoyl, 4-chlorobenzoyl, lower alkylated arylcarbonyl groups such as 2,4,6-trimethylbenzoyl, 4-toluoyl, lower alkoxylated arylcarbonyl groups such as 4-anisoyl, carboxylated arylcarbonyl groups such as 2-carboxybenzoyl, 3-carboxybenzoyl, 4-carboxybenzoyl, nitrated arylcarbonyl groups such as 4-nitrobenzoyl, 2-nitrobenzoyl; lower alkoxycarbonylated arylcarbonyl groups such as 2-(methoxycarbonyl)benzoyl, arylated arylcarbonyl groups such as 4-phenylbenzoyl; "Tetrahydropyranyl or tetrahydrothiopyranyl group" such as tetrahydropyran-2-yl, 3-bromotetrahydropyran-2-yl, 4-methoxytetrahydropyran-4-yl, tetrahydrothiopyran-2-yl, 4-methoxytetrahydrothiopyran-4-yl; "tetrahydrofuranyl or tetrahydrothiofuranyl group" such as tetrahydrofuran-2-yl, tetrahydrothiofuran-2-yl; "Silyl groups" such as tri-lower alkylsilyl groups, such as trimethylsilyl, triethylsilyl, isopropyldimethylsilyl, t-butyldimethylsilyl, methyldiisopropylsilyl, methyldi-t-butylsilyl, and triisopropylsilyl, and tri-lower alkylsilyl groups substituted by one or two aryl groups, such as diphenylmethylsilyl, diphenylbutylsilyl, diphenylisopropylsilyl, and phenyldiisopropylsilyl; "lower alkoxymethyl groups" such as methoxymethyl, 1,1-dimethyl-1-methoxymethyl, ethoxymethyl, propoxymethyl, isopropoxymethyl, butoxymethyl, and t-butoxymethyl; "lower alkoxylated lower alkoxymethyl groups" such as 2-methoxyethoxymethyl; "halogeno lower alkoxymethyl" such as 2,2,2-trichloroethoxymethyl, bis(2-chloroethoxy)methyl; "lower alkoxyethyl groups" such as 1-ethoxyethyl, 1-(isopropoxy)ethyl; "halogenated ethyl groups" such as 2,2,2-trichloroethyl; "Methyl groups substituted with one to three aryl groups" such as benzyl, α-naphthylmethyl, β-naphthylmethyl, diphenylmethyl, triphenylmethyl, α-naphthyldiphenylmethyl, and 9-anthrylmethyl; "Methyl groups substituted with one to three aryl groups, the aryl ring of which is substituted with a lower alkyl, lower alkoxy, halogen or cyano group", such as 4-methylbenzyl, 2,4,6-trimethylbenzyl, 3,4,5-trimethylbenzyl, 4-methoxybenzyl, 4-methoxyphenyldiphenylmethyl, 4,4'-dimethoxytriphenylmethyl, 2-nitrobenzyl, 4-nitrobenzyl, 4-chlorobenzyl, 4-bromobenzyl and 4-cyanobenzyl; "lower alkoxycarbonyl groups" such as methoxycarbonyl, ethoxycarbonyl, t-butoxycarbonyl, isobutoxycarbonyl; "Lower alkoxycarbonyl groups substituted with halogen or tri-lower alkylsilyl groups" such as 2,2,2-trichloroethoxycarbonyl and 2-trimethylsilylethoxycarbonyl; "Alkenyloxycarbonyl groups" such as vinyloxycarbonyl and aryloxycarbonyl; Examples of the aryloxycarbonyl group include 1 or 2 "aralkyloxycarbonyl groups in which the aryl ring may be substituted with a lower alkoxy or nitro group", such as benzyloxycarbonyl, 4-methoxybenzyloxycarbonyl, 3,4-dimethoxybenzyloxycarbonyl, 2-nitrobenzyloxycarbonyl and 4-nitrobenzyloxycarbonyl.

[0145] Z 1 , Z 2The "hydroxyl-protecting group" in the above is preferably an "aliphatic acyl group", an "aromatic acyl group", a "methyl group substituted with one to three aryl groups", a "methyl group substituted with one to three aryl groups in which the aryl ring is substituted with a lower alkyl, lower alkoxy, halogen or cyano group" or a "silyl group", more preferably an acetyl group, a benzoyl group, a benzyl group, a p-methoxybenzoyl group, a dimethoxytrityl group, a monomethoxytrityl group or a t-butyldiphenylsilyl group, and even more preferably a benzyl group.

[0146] The "protected hydroxyl group" of the α group is an "aliphatic acyl group" or an "aromatic acyl group", and more preferably a benzoyl group.

[0147] In the present invention, A, R 1 The "lower alkyl group" of the α group and the α group refers to a straight or branched alkyl group having 1 to 6 carbon atoms, and examples thereof include a methyl group, an ethyl group, an n-propyl group, an isopropyl group, an n-butyl group, an isobutyl group, an s-butyl group, a t-butyl group, an n-pentyl group, an isopentyl group, a 2-methylbutyl group, a neopentyl group, a 1-ethylpropyl group, an n-hexyl group, an isohexyl group, a 4-methylpentyl group, a 3-methylpentyl group, a 2-methylpentyl group, a 1-methylpentyl group, a 3,3-dimethylbutyl group, a 2,2-dimethylbutyl group, a 1,1-dimethylbutyl group, a 1,2-dimethylbutyl group, a 1,3-dimethylbutyl group, a 2,3-dimethylbutyl group, and a 2-ethylbutyl group.

[0148] The "lower alkyl group" of A is preferably a methyl group, an ethyl group or a propyl group, and more preferably a methyl group.

[0149] R 1 The "lower alkyl group" is preferably a methyl group.

[0150] In the present invention, the "lower alkoxy group" of the α group refers to a group in which the above-mentioned "lower alkyl group" is bonded to an oxygen atom, and examples thereof include methoxy, ethoxy, n-propoxy, isopropoxy, n-butoxy, isobutoxy, s-butoxy and t-butoxy, and preferably is a methoxy or ethoxy group.

[0151] In the present invention, examples of the protecting group for the "protected mercapto group" of the α group include, in addition to the above-mentioned protecting groups for the hydroxyl group, "groups that form disulfides" such as alkylthio groups, e.g., methylthio, ethylthio, and t-butylthio, and arylthio groups, e.g., benzylthio, and are preferably an "aliphatic acyl group" or an "aromatic acyl group", and more preferably a benzoyl group.

[0152] In the present invention, the "lower alkylthio group" of the α group refers to a group in which the above-mentioned "lower alkyl group" is bonded to a sulfur atom, and examples thereof include methylthio, ethylthio, propylthio, isopropylthio, butylthio, isobutylthio, s-butylthio, and t-butylthio, and is preferably a methylthio or ethylthio group.

[0153] In the present invention, examples of the protecting group for the "protected amino group" of the α group include formyl, acetyl, propionyl, butyryl, isobutyryl, pentanoyl, pivaloyl, valeryl, isovaleryl, octanoyl, nonanoyl, decanoyl, 3-methylnonanoyl, 8-methylnonanoyl, 3-ethyloctanoyl, 3,7-dimethyloctanoyl, undecanoyl, dodecanoyl, tridecanoyl, tetradecanoyl, pentadecanoyl, hexadecanoyl, 1-methylpentadecanoyl, 14-methylpentadecanoyl, 13,13-dimethyltetradecanoyl, heptadecanoyl, 1-methylpentadecanoyl, 14-methylpentadecanoyl, 13,13-dimethyltetradecanoyl, and 1-methylpentadecanoyl. "Aliphatic acyl groups" such as alkylcarbonyl groups such as canoyl, 15-methylhexadecanoyl, octadecanoyl, 1-methylheptadecanoyl, nonadecanoyl, eicosanoyl, and henaicosanoyl, carboxylated alkylcarbonyl groups such as succinoyl, glutaroyl, and adipoyl, halogeno lower alkylcarbonyl groups such as chloroacetyl, dichloroacetyl, trichloroacetyl, and trifluoroacetyl, lower alkoxy lower alkylcarbonyl groups such as methoxyacetyl, and unsaturated alkylcarbonyl groups such as (E)-2-methyl-2-butenoyl; "Aromatic acyl groups" such as arylcarbonyl groups such as benzoyl, α-naphthoyl, and β-naphthoyl, halogenoarylcarbonyl groups such as 2-bromobenzoyl and 4-chlorobenzoyl, lower alkylated arylcarbonyl groups such as 2,4,6-trimethylbenzoyl and 4-toluoyl, lower alkoxylated arylcarbonyl groups such as 4-anisoyl, carboxylated arylcarbonyl groups such as 2-carboxybenzoyl, 3-carboxybenzoyl, and 4-carboxybenzoyl, nitrated arylcarbonyl groups such as 4-nitrobenzoyl and 2-nitrobenzoyl; lower alkoxycarbonylated arylcarbonyl groups such as 2-(methoxycarbonyl)benzoyl, and arylated arylcarbonyl groups such as 4-phenylbenzoyl; "lower alkoxycarbonyl groups" such as methoxycarbonyl, ethoxycarbonyl, t-butoxycarbonyl, isobutoxycarbonyl; "Lower alkoxycarbonyl groups substituted with halogen or tri-lower alkylsilyl groups" such as 2,2,2-trichloroethoxycarbonyl and 2-trimethylsilylethoxycarbonyl; "Alkenyloxycarbonyl groups" such as vinyloxycarbonyl and aryloxycarbonyl; Examples of aryloxycarbonyl groups include aralkyloxycarbonyl groups in which the aryl ring is optionally substituted with one or two lower alkoxy or nitro groups, such as benzyloxycarbonyl, 4-methoxybenzyloxycarbonyl, 3,4-dimethoxybenzyloxycarbonyl, 2-nitrobenzyloxycarbonyl and 4-nitrobenzyloxycarbonyl, and are preferably an aliphatic acyl group or an aromatic acyl group, and more preferably a benzoyl group.

[0154] In the present invention, the "lower alkylamino group" of the α group refers to a group in which one or two hydrogen atoms of an amino group are substituted by the above-mentioned "lower alkyl group", and examples thereof include methylamino, ethylamino, propylamino, isopropylamino, butylamino, isobutylamino, s-butylamino, t-butylamino, dimethylamino, diethylamino, dipropylamino, diisopropylamino, dibutylamino, diisobutylamino, di(s-butyl)amino, and di(t-butyl)amino, and are preferably methylamino, ethylamino, dimethylamino, diethylamino, or diisopropylamino.

[0155] In the present invention, X 1 , X 2 And the "halogen atom" of the α group can include, for example, a fluorine atom, a chlorine atom, a bromine atom or an iodine atom.

[0156] In the present invention, X 1 and X 2 Examples of the "group forming a leaving group" include a halogen atom, an aliphatic acyloxy group, a halogen-substituted lower alkylimidooxy group, and a halogen-substituted lower alkylsulfonyloxy group.

[0157] X 1 The "group forming a leaving group" is preferably an iodine atom, an acetoxy group, or a trichloroacetimidooxy group.

[0158] X 2 The "group forming a leaving group" is preferably an acetoxy group.

[0159] In the present invention, examples of the "methyl group substituted with 1 to 3 aryl groups" of Y include benzyl, α-naphthylmethyl, β-naphthylmethyl, diphenylmethyl, trityl, α-naphthyldiphenylmethyl, and 9-anthrylmethyl groups, and preferably a trityl group.

[0160] In the present invention, examples of "a methyl group substituted by 1 to 3 aryl groups in which the aryl ring is substituted by a lower alkyl, a lower alkoxy, a halogen or a cyano group" for Y include 4-methylbenzyl, 2,4,6-trimethylbenzyl, 3,4,5-trimethylbenzyl, 4-methoxybenzyl, 4-methoxyphenyldiphenylmethyl, 4,4'-dimethoxytriphenylmethyl (4,4'-dimethoxytrityl), 2-nitrobenzyl, 4-nitrobenzyl, 4-chlorobenzyl, 4-bromobenzyl and 4-cyanobenzyl groups.

[0161] In the present invention, the "lower alkoxymethyl group" of Y is a group in which a methyl group is bonded to the above-mentioned "lower alkoxy group". Examples include methoxymethyl, 1,1-dimethyl-1-methoxymethyl, ethoxymethyl, propoxymethyl, isopropoxymethyl, butoxymethyl, and t-butoxymethyl, and is preferably a methoxymethyl group.

[0162] In the present invention, examples of the "tetrahydropyranyl group" of Y include tetrahydropyran-2-yl, 3-bromotetrahydropyran-2-yl, or 4-methoxytetrahydropyran-4-yl groups, and preferably tetrahydropyran-2-yl groups.

[0163] In the present invention, examples of the "silyl group" of Y include tri-lower alkylsilyl groups such as trimethylsilyl, triethylsilyl, isopropyldimethylsilyl, t-butyldimethylsilyl, methyldiisopropylsilyl, methyldi-t-butylsilyl, and triisopropylsilyl, and tri-lower alkylsilyl groups substituted with one or two aryl groups such as diphenylmethylsilyl, diphenylbutylsilyl, diphenylisopropylsilyl, and phenyldiisopropylsilyl, and is preferably a t-butyldiphenylsilyl group or a t-butyldimethylsilyl group.

[0164] P 1 The "trityl group optionally substituted with 1 to 3 lower alkoxy groups" refers to a group in which 1 to 3 hydrogen atoms of the phenyl group in the trityl group are substituted with the above "lower alkoxy groups", and examples thereof include a trityl group, a monomethoxytrityl group, or a dimethoxytrityl group, and preferably a 4,4'-dimethoxytrityl group.

[0165] In the present invention, R, R 3 , R 4 , R 6 and Z 3Examples of the "aliphatic acyl group" include formyl, acetyl, propionyl, butyryl, isobutyryl, pentanoyl, pivaloyl, valeryl, isovaleryl, octanoyl, nonanoyl, decanoyl, 3-methylnonanoyl, 8-methylnonanoyl, 3-ethyloctanoyl, 3,7-dimethyloctanoyl, undecanoyl, dodecanoyl, tridecanoyl, tetradecanoyl, pentadecanoyl, hexadecanoyl, 1-methylpentadecanoyl, 14-methylpentadecanoyl, 13,13-dimethyltetradecanoyl, heptadecanoyl, 15 Examples of lower alkylcarbonyl groups include alkylcarbonyl groups such as 1-methylhexadecanoyl, octadecanoyl, 1-methylheptadecanoyl, nonadecanoyl, eicosanoyl and henaicosanoyl; carboxylated alkylcarbonyl groups such as succinoyl, glutaroyl and adipoyl; halogeno lower alkylcarbonyl groups such as chloroacetyl, dichloroacetyl, trichloroacetyl and trifluoroacetyl; lower alkoxy lower alkylcarbonyl groups such as methoxyacetyl; and unsaturated alkylcarbonyl groups such as (E)-2-methyl-2-butenoyl.

[0166] R, R 3 , R 6 and Z 3 The "aliphatic acyl group" is preferably an acetyl group.

[0167] R 4 The "aliphatic acyl group" is preferably an isobutyryl group.

[0168] In the present invention, R 3 , R 4 , R 6 and Z 3Examples of the "aromatic acyl group" include arylcarbonyl groups such as benzoyl, α-naphthoyl, and β-naphthoyl; halogenoarylcarbonyl groups such as 2-bromobenzoyl and 4-chlorobenzoyl; lower alkylated arylcarbonyl groups such as 2,4,6-trimethylbenzoyl and 4-toluoyl; lower alkoxylated arylcarbonyl groups such as 4-anisoyl; carboxylated arylcarbonyl groups such as 2-carboxybenzoyl, 3-carboxybenzoyl, and 4-carboxybenzoyl; nitrated arylcarbonyl groups such as 4-nitrobenzoyl and 2-nitrobenzoyl; lower alkoxycarbonylated arylcarbonyl groups such as 2-(methoxycarbonyl)benzoyl; and arylated arylcarbonyl groups such as 4-phenylbenzoyl.

[0169] R 3 , R 4 , R 6 and Z 3 The "aromatic acyl group" is preferably a benzoyl group.

[0170] In the present invention, R 2 The "amino group protected by an aliphatic acyl group or an aromatic acyl group" in the formula (1) refers to a group in which an amino group is substituted with the above-mentioned "aliphatic acyl group" or "aromatic acyl group". 2 Examples of the "amino group protected by an aliphatic acyl group" include formylamino, acetylamino, propionylamino, butyrylamino, isobutyrylamino, pentanoylamino, pivaloylamino, valerylamino, chloroacetylamino, dichloroacetylamino, trichloroacetylamino, trifluoroacetylamino, methoxyacetylamino, and (E)-2-methyl-2-butenoylamino groups. 2Examples of the "amino group protected by an aromatic acyl group" include benzoylamino, α-naphthoylamino, β-naphthoylamino, 2-bromobenzoylamino, 4-chlorobenzoylamino, 2,4,6-trimethylbenzoylamino, 4-toluoylamino, 4-anisoylamino, 2-carboxybenzoylamino, 3-carboxybenzoylamino, 4-carboxybenzoylamino, 4-nitrobenzoylamino, 2-nitrobenzoylamino, 2-(methoxycarbonyl)benzoylamino or 4-phenylbenzoylamino groups, and a benzoylamino group is preferred.

[0171] In the present invention, R 5 Examples of the "benzyl group optionally substituted with lower alkyl, lower alkoxy, halogen, or cyano group" include benzyl, 4-methylbenzyl, 2,4,6-trimethylbenzyl, 3,4,5-trimethylbenzyl, 4-methoxybenzyl, 4-chlorobenzyl, 4-bromobenzyl, and 4-cyanobenzyl, and a benzyl group is preferable.

[0172] In the above general formulae (XXVII) and (XXVIII), the "purin-9-yl group which may have one or more substituents selected from group α" of B is preferably 6-aminopurin-9-yl (i.e., adeninyl), 6-aminopurin-9-yl with a protected amino group, 2,6-diaminopurin-9-yl, 2-amino-6-chloropurin-9-yl, 2-amino-6-chloropurin-9-yl with a protected amino group, 2-amino-6-fluoropurin-9-yl, 2-amino-6-fluoropurin-9-yl with a protected amino group, 2-amino-6-bromopurin-9-yl, 2-amino-6-bromopurin-9-yl with a protected amino group, 2-amino-6-hydroxypurin-9-yl (i.e., guanyl), purin-9-yl) with a protected amino group, 2-amino-6-hydroxypurin-9-yl with a protected amino group, 2-amino-6-hydroxypurin-9-yl, 6-amino-2-methoxypurin-9-yl, 6-amino-2-chloropurin-9-yl, 6-amino-2-fluoropurin-9-yl, 2,6-dimethoxypurin-9-yl, 2,6-dichloropurin-9-yl or 6-mercaptopurin-9-yl group with a protected amino group, more preferably 6-aminopurin-9-yl group with a protected amino group or 2-amino-6-hydroxypurin-9-yl group with a protected amino group, and even more preferably 6-benzoylaminopurin-9-yl or 2-isobutyrylamino-6-hydroxypurin-9-yl.

[0173] In the above general formulas (XXVII) and (XXVIII), the "2-oxo-pyrimidin-1-yl group which may have one or more substituents selected from group α" of B is preferably 2-oxo-4-amino-pyrimidin-1-yl (i.e., cytosinyl), 2-oxo-4-amino-pyrimidin-1-yl with a protected amino group, 2-oxo-4-amino-5-fluoro-pyrimidin-1-yl, 2-oxo-4-amino-5-fluoro-pyrimidin-1-yl with a protected amino group, 4-amino-2-oxo-5-chloro-pyrimidin-1-yl, 2-oxo-4-methoxy-pyrimidin-1-yl, 2-oxo-4-mercapto-pyrimidin-1-yl, 2-oxo-4-hydroxy-pyrimidin-1-yl (i.e., uracinyl), 2-oxo-4-hydroxy-pyrimidin-1-yl, and 4-amino-5-methyl-2-oxo-pyrimidin-1-yl group with a protected amino group, more preferably 2-oxo-4-hydroxy-5-methylpyrimidin-1-yl group, 2-oxo-4-amino-pyrimidin-1-yl group with a protected amino group, or 4-amino-5-methyl-2-oxo-pyrimidin-1-yl group with a protected amino group, and even more preferably 2-oxo-4-hydroxy-5-methylpyrimidin-1-yl group, 2-oxo-4-benzoylamino-pyrimidin-1-yl group, or 4-benzoylamino-5-methyl-2-oxo-pyrimidin-1-yl group.

[0174] In the present invention, n is an integer of 0 to 4, preferably 0 or 1, and more preferably 1.

[0175] The term "salt thereof" refers to a salt of the compound of the present invention, since it can be made into a salt. Suitable examples of such salts include metal salts such as alkali metal salts, potassium salts, and lithium salts, alkaline earth metal salts, such as calcium salts and magnesium salts, aluminum salts, iron salts, zinc salts, copper salts, nickel salts, and cobalt salts; inorganic salts such as ammonium salts, t-octylamine salts, dibenzylamine salts, morpholine salts, glucosamine salts, phenylglycine alkyl ester salts, ethylenediamine salts, N-methylglucamine salts, guanidine salts, diethylamine salts, triethylamine salts, dicyclohexylamine salts, N,N'-dibenzylethylenediamine salts, chloroprocaine salts, procaine salts, diethanolamine salts, N-benzyl-phenyl Examples of the salt include amine salts, such as organic salts such as netylamine salts, piperazine salts, tetramethylammonium salts, and tris(hydroxymethyl)aminomethane salts; inorganic acid salts, such as hydrohalogen salts, such as hydrofluoride, hydrochloride, hydrobromide, and hydroiodide, nitrates, perchlorates, sulfates, and phosphates; organic acid salts, such as lower alkane sulfonates, such as methanesulfonates, trifluoromethanesulfonates, and ethanesulfonates, arylsulfonates, such as benzenesulfonates and p-toluenesulfonates, acetates, malates, fumarates, succinates, citrates, tartrates, oxalates, and maleates; and amino acid salts, such as glycine salts, lysine salts, arginine salts, ornithine salts, glutamate salts, and aspartate salts.

[0176] In the present invention, the carbonyl group of a thymine derivative or a guanine derivative can be in a "tautomer". A tautomer is one of two or more structural isomers that exist in equilibrium and can be easily converted from one isomer to another, existing as a mixture of tautomeric pairs in solution. Under conditions where tautomerization is possible, a chemical equilibrium of tautomers is reached, the exact ratio of which depends on several factors including temperature, solvent, and pH. The concept of tautomers that are interconvertible by tautomerization is called tautomerism.

[0177] Examples of amide-imidic acid tautomerism in thymine and guanine derivatives are shown below.

[0178] [ka]

[0179] In the present invention, the term "hydroxyl-protecting reagent" refers to a reagent used for introducing the above-mentioned "hydroxyl-protecting group" into the hydroxyl group of a nucleoside analogue or its intermediate. For example, the "hydroxyl-protecting group" may be (1) Benzyl group: benzyl chloride, benzyl bromide, benzyl iodide, benzyl trichloroacetimidate (2) In the case of a trityl group: trityl chloride, trityl trifluorosulfonate, (3) In the case of t-butyldimethylsilyl group: t-butyldimethylsilyl chloride, (4) In the case of acetyl group: acetic anhydride, acetyl chloride It is.

[0180] In the present invention, the "protecting reagent for a primary hydroxyl group" refers to a reagent used for introducing the above-mentioned "trityl group which may be substituted with 1 to 3 lower alkoxy groups" into the sugar hydroxyl group at the 5-position of a nucleoside analogue, and examples thereof include trityl chloride, trityl trifluorosulfonate, 4-monomethoxytrityl chloride, 4,4'-dimethoxytrityl chloride, and 4,4'-dimethoxytrityl trifluorosulfonate, with 4,4'-dimethoxytrityl chloride being preferred.

[0181] In the present invention, the term "hydroxyl group deprotection reagent" refers to a reaction auxiliary added to remove the above-mentioned "hydroxyl group protecting group". For example, the "hydroxyl group protecting group" is (1) In the case of benzyl group: metal catalysts such as palladium, palladium hydroxide, platinum, etc., and reducing agents such as hydrogen, formic acid, ammonium formate, etc., iron(III) chloride, boron trichloride, etc. (2) In the case of a trityl group: an acid catalyst such as sulfuric acid, p-toluenesulfonic acid, or methanesulfonic acid (3) In the case of an acetyl group: inorganic bases such as sodium hydroxide, potassium hydroxide, and sodium carbonate; and organic bases such as ammonia and alkylamines. In the present invention, the metal catalyst may be a metal catalyst supported on a carrier (preferably carbon) or not, and is preferably a metal catalyst supported on carbon (preferably palladium, palladium hydroxide, or platinum supported on carbon, more preferably palladium supported on carbon).

[0182] In the present invention, the term "acylating agent" refers to a reagent used for introducing the above-mentioned "amino group-protecting group" into the amino group of a nucleoside analogue, and examples thereof include: (1) In the case of acetyl group: acetic anhydride, acetyl chloride (2) In the case of benzoyl group: benzoyl chloride, benzoic anhydride, benzoic acid (3) In the case of isobutyryl group: isobutyl chloride isobutyric anhydride It is.

[0183] In the present invention, the "aminating agent" refers to a reagent used to replace the chlorine atom at the 6-position of a purine ring or the carbonyl group at the 4-position of a pyrimidine ring with an amino group, and is, for example, ammonia, an aqueous ammonia solution, or an ammonia salt such as ammonium carbonate or ammonium acetate, and is preferably an aqueous ammonia solution.

[0184] In the present invention, the "amidating agent" refers to a reagent used for converting the chlorine atom at the 6-position of a purine ring to an aliphatic amide or aromatic amide in a cross-coupling reaction, and examples of such an agent include acetylamide, benzoylamide, and isobutylamide, and preferably isobutylamide.

[0185] In the present invention, the "activating agent" in the glycosylation reaction is a reagent used to convert the hydroxyl group at position 1 of a sugar to the above-mentioned "group that forms a leaving group", and is, for example, acetic anhydride, benzoic anhydride, trichloroacetonitrile, carbonyldiimidazole, or diphenyl chlorophosphate, and is preferably acetic anhydride or trichloroacetonitrile.

[0186] In the present invention, the "halogenating agent" in the glycosylation reaction is a reagent used to halogenate the above-mentioned "group that forms a leaving group" in order to stereoselectively proceed with the glycosylation reaction, and examples of such a reagent include chlorotrimethylsilane (TMSCl), bromotrimethylsilane (TMSBr), and iodotrimethylsilane (TMSI), with TMSI being preferred.

[0187] In the present invention, the "activating agent" in the amination reaction is a reagent used for converting a hydroxyl group into a leaving group, and is, for example, p-toluenesulfonyl chloride or 2,4,6-triisopropylbenzenesulfonyl chloride, preferably 2,4,6-triisopropylbenzenesulfonyl chloride.

[0188] In the present invention, the "activator" in the amidite reaction is a reagent used to form an active intermediate of the amidite reagent, and examples thereof include 5-benzylthiotetrazole, 5-phenyltetrazole, dibromoimidazole, dicyanoimidazole, N-alkylimidazole trifluoroacetate, etc., and preferably 4,5-dicyanoimidazole. The amount of the activator used is 0.01 to 1.0 equivalent relative to the compound of general formula (XXVII), and preferably 0.1 to 0.5 equivalent.

[0189] In the present invention, the "amidite reagent" in the amiditization reaction is a reagent used to introduce a phosphorus atom-containing group useful for forming an internucleoside bond to the sugar hydroxyl group at the 3-position of a nucleoside analogue, and examples thereof include 2-cyanoethyl N,N,N',N'-tetraisopropyl phosphorodiamidite and 2-cyanoethyl diisopropyl chlorophosphoroamidite, and preferably 2-cyanoethyl N,N,N',N'-tetraisopropyl phosphorodiamidite. The amount of the amiditization reagent used is 1.0 to 1.5 equivalents, preferably 1.1 to 1.3 equivalents, relative to the compound of general formula (XXVII).

[0190] In the present invention, the "desiccant" in the amidite reaction is a reagent used to absorb moisture in the reaction solution, such as molecular sieve 3A, molecular sieve 4A, or molecular sieve 5A, and preferably molecular sieve 4A.

[0191] The compound (2A) of the present invention can be produced by Method A described below. (Method A)

[0192] [ka]

[0193] A method, A, Y, Z 1 and Z 2 has the same meaning as above. 1 , Z 2 represents a protecting group for a hydroxyl group that is stable under the deprotection conditions of Y.

[0194] Each step of Method A will be described in detail below. (A-1 process) This step is a step of producing compound (14A) by deprotecting the protecting groups at the 1,2-hydroxyl groups and Y of compound (13A), which can be produced by Method C or D described below, in an alcohol solvent in the presence of an acid catalyst. When n=0, this step can be omitted, and step A-2 can be carried out using compound (14A') produced by Method B described below.

[0195] The alcohol solvent to be used may be, for example, alcohols such as methanol, ethanol, and propanol, with methanol being preferred.

[0196] Examples of the acid catalyst to be used include sulfuric acid, p-toluenesulfonic acid, methanesulfonic acid, etc., with sulfuric acid being preferred.

[0197] The reaction temperature is usually from 0°C to 100°C, preferably from 40°C to 60°C.

[0198] The reaction time varies depending on the type and amount of the acid catalyst used, but is usually from 1 to 48 hours, preferably from 10 to 24 hours.

[0199] After the reaction is completed, the alcohol solution of the target compound (14A) of this reaction is, for example, neutralized with triethylamine, and the co-product methoxytriphenylmethane is crystallized by adding water, and the target compound is separated into an aqueous alcohol solution by filtering it off. The aqueous alcohol solution obtained by separating and washing with n-heptane is separated by adding water and toluene, and the obtained organic layer is concentrated to obtain a toluene solution, which can be used as it is in the next step.

[0200] The resulting compound can be further purified, if necessary, by a conventional method such as recrystallization or silica gel chromatography. (A-2 process) This step is a step of producing compound (15A) by carrying out a cyclization reaction of the diol moiety of compound (14A) produced in step A-1 using a trivalent phosphorus reagent and an azodicarboxylate in a solvent.

[0201] Examples of the solvent to be used include halogenated hydrocarbons such as methylene chloride, chloroform, and 1,2-dichloroethane, hydrocarbons such as benzene and toluene, and ethers such as tetrahydrofuran and dimethyl ether, with toluene being preferred.

[0202] The trivalent phosphorus reagent used may include triphenylphosphine, tri(n-butyl)phosphine, etc., with triphenylphosphine being preferred.

[0203] Examples of the azodicarboxylate that can be used include diethyl azodicarboxylate, diisopropyl azodicarboxylate, di-t-butyl azodicarboxylate, etc., with diisopropyl azodicarboxylate being preferred.

[0204] The reaction temperature is usually from 0°C to 50°C, preferably from 10 to 40°C.

[0205] The reaction time varies depending on the type and amount of the trivalent phosphorus reagent used, but is usually 1 to 24 hours, preferably 1 to 3 hours.

[0206] After the reaction is completed, the toluene solution of the target compound (15A) of this reaction can be filtered to remove the co-product phosphine oxide by, for example, adding magnesium chloride to the reaction solution and stirring to form a poorly soluble phosphine complex. In addition, the other co-product, hydrazine diester, can also be effectively removed by washing the toluene layer with methanol water. The toluene solution obtained by concentrating the resulting organic layer can be used as it is in the next step.

[0207] The resulting compound can be further purified, if necessary, by a conventional method such as recrystallization or silica gel chromatography. (A-3 process) This step is a step of producing compound (2A) by hydrolysis of the anomeric position of compound (15A) produced in step A-2 by the action of an acid in a solvent.

[0208] Examples of the solvent to be used include water-soluble solvents such as acetic acid, water, and alcohol; halogenated hydrocarbons such as methylene chloride, chloroform, and 1,2-dichloroethane; hydrocarbons such as benzene and toluene; and ethers such as tetrahydrofuran and dimethyl ether. Of these, acetic acid is preferred.

[0209] The acid to be used may include hydrochloric acid, sulfuric acid, trifluoroacetic acid, methanesulfonic acid, p-toluenesulfonic acid, etc., with hydrochloric acid being preferred.

[0210] The reaction temperature is usually from 0°C to 50°C, preferably from 20°C to 30°C.

[0211] The reaction time varies depending on the type and amount of the acid used, but is usually from 1 to 24 hours, preferably from 1 to 3 hours.

[0212] After completion of the reaction, the target compound (2A) of this reaction can be crystallized, for example, by adding water to the reaction mixture, cooling it, and stirring it.

[0213] The resulting compound can be further purified, if necessary, by a conventional method such as recrystallization or silica gel chromatography.

[0214] The above compound (14A) (when n=0) can be produced by the following Method B. (Method B)

[0215] [ka]

[0216] B method, A, Z 1 and Z 2 has the same meaning as above.

[0217] The process of Method B is explained in detail below. (B-1 process) This step is a step of producing compound (14A') using compound (1B) produced according to the production method of compound (7) in WO00 / 47599. Compound (14A') can be produced by deprotecting the protecting groups at the 1,2-position hydroxyl groups of compound (1B) according to step A-1 of Method A.

[0218] The above compound (13A) (when n=1) can be produced by the following Method C. (C method)

[0219] [ka]

[0220] In method C, X is a group that forms a leaving group together with the oxygen atom. 1 and Z 2 has the same meaning as above.

[0221] Examples of X include lower alkylsulfonyl groups such as methanesulfonyl and ethanesulfonyl, halogen-substituted lower alkylsulfonyl groups such as trifluoromethanesulfonyl, and arylsulfonyl groups such as p-toluenesulfonyl, and are preferably a methanesulfonyl group or a p-toluenesulfonyl group.

[0222] Each step of Method C will be explained in detail below. (C-1 process) This step is a step of producing compound (5) by reacting compound (4) in an acetone solvent in the presence of an acid catalyst.

[0223] Examples of the acid catalyst to be used include sulfuric acid, p-toluenesulfonic acid, methanesulfonic acid, etc., with sulfuric acid being preferred.

[0224] The reaction temperature varies depending on the acid catalyst used, but is usually from 0°C to 50°C, preferably from 30°C to 40°C.

[0225] The reaction time varies depending on the type and amount of the acid catalyst used, but is usually from 10 minutes to 24 hours, preferably from 10 hours to 20 hours.

[0226] After completion of the reaction, the target compound (5) of this reaction can be obtained, for example, by neutralizing the reaction mixture, concentrating the reaction mixture, and distilling off the solvent.

[0227] The resulting compound can be further purified, if necessary, by a conventional method such as recrystallization or silica gel chromatography. (C-2 process) This step is a step of producing compound (6) by reacting compound (5) produced in step C-1 with a leaving group introducing reagent in a solvent in the presence of a base.

[0228] Examples of the solvent to be used include amides such as dimethylacetamide and dimethylformamide, halogenated hydrocarbons such as methylene chloride, chloroform and 1,2-dichloroethane, hydrocarbons such as benzene and toluene, ethers such as tetrahydrofuran and dimethyl ether, and esters such as methyl acetate, ethyl acetate and propyl acetate, and preferably dimethylacetamide.

[0229] The base used may be, for example, triethylamine, pyridine, dimethylaminopyridine or 1-methylimidazole, with 1-methylimidazole being preferred.

[0230] The leaving group introducing reagent used includes, for example, p-toluenesulfonyl chloride, methanesulfonyl chloride, trifluoromethanesulfonic anhydride, etc., with p-toluenesulfonyl chloride being preferred.

[0231] The reaction temperature varies depending on the leaving group introducing reagent used, but is usually 0°C to 50°C, preferably 20°C to 30°C.

[0232] The reaction time varies depending on the type and amount of the leaving group introducing reagent used, but is usually from 10 minutes to 24 hours, preferably from 1 hour to 5 hours.

[0233] After completion of the reaction, the target compound (6) of this reaction can be obtained, for example, by adding water to the reaction mixture and crystallizing it.

[0234] The resulting compound can be further purified, if necessary, by a conventional method such as recrystallization or silica gel chromatography. (C-3 process) This step is a step of producing compound (7) by reacting compound (6) produced in step C-2 with a hydride reducing agent in a solvent.

[0235] Examples of the solvent to be used include halogenated hydrocarbons such as methylene chloride, chloroform, and 1,2-dichloroethane; hydrocarbons such as benzene and toluene; and ethers such as tetrahydrofuran and dimethyl ether. Among them, tetrahydrofuran is preferable.

[0236] Examples of the hydride reducing agent to be used include sodium bis(2-methoxyethoxy)aluminum hydride, lithium aluminum hydride, and diisobutylaluminum hydride, and preferably sodium bis(2-methoxyethoxy)aluminum hydride.

[0237] The reaction time varies depending on the type and amount of the hydride reducing agent used, but is usually from 10 minutes to 24 hours, preferably from 1 hour to 3 hours.

[0238] After completion of the reaction, the target compound (7) of this reaction can be obtained, for example, by adding acetone and an aqueous solution of sodium potassium L-tartrate to the reaction mixture, separating the organic layer containing the target compound, washing it with water, and then distilling off the solvent.

[0239] The resulting compound can be further purified, if necessary, by a conventional method such as recrystallization or silica gel chromatography. (C-4 process) This step is a step of producing compound (8) by selectively introducing a protecting group into the primary hydroxyl group of compound (7) produced in step C-3 in a solvent in the presence of a base.

[0240] Examples of the solvent to be used include amides such as dimethylacetamide and dimethylformamide, hydrocarbons such as benzene and toluene, halogenated hydrocarbons such as methylene chloride, chloroform and 1,2-dichloroethane, ethers such as tetrahydrofuran and dimethyl ether, and esters such as methyl acetate, ethyl acetate and propyl acetate, and toluene is preferred.

[0241] Examples of the base to be used include triethylamine, pyridine, dimethylaminopyridine, 1-methylimidazole, and 4-methylmorpholine, with 4-methylmorpholine being preferred.

[0242] The primary hydroxyl group protecting reagent used includes, for example, trityl chloride, 4,4'-dimethoxytrityl chloride, etc., with trityl chloride being preferred.

[0243] The reaction temperature varies depending on the protecting reagent used, but is usually 0°C to 50°C, preferably 20°C to 30°C.

[0244] The reaction time varies depending on the type and amount of the protecting reagent for the primary hydroxyl group, but is usually from 10 minutes to 24 hours, preferably from 1 hour to 5 hours.

[0245] After completion of the reaction, the target compound (8) of this reaction can be obtained, for example, by adding water to the reaction mixture, separating the organic layer containing the target compound, washing it with water, and then distilling off the solvent.

[0246] The resulting compound can be further purified, if necessary, by a conventional method such as recrystallization or silica gel chromatography. (C-5 process) This step is a step of producing compound (9) by oxidizing the secondary hydroxyl group at the 3-position of compound (8) produced in step C-4 in a solvent in the presence of a base, an oxidizing agent, an oxidation catalyst and a cocatalyst.

[0247] Examples of the solvent to be used include amides such as dimethylacetamide and dimethylformamide, hydrocarbons such as benzene and toluene, halogenated hydrocarbons such as methylene chloride, chloroform and 1,2-dichloroethane, ethers such as tetrahydrofuran and dimethyl ether, and esters such as methyl acetate, ethyl acetate and propyl acetate, and toluene is preferred.

[0248] Examples of the base to be used include inorganic bases such as sodium hydroxide, potassium hydroxide, sodium bicarbonate, and sodium carbonate, and organic bases such as triethylamine, pyridine, dimethylaminopyridine, 1-methylimidazole, and 4-methylmorpholine, with sodium bicarbonate being preferred.

[0249] Examples of the oxidation catalyst to be used include 2,2,6,6-tetramethylpiperidine 1-oxyl, 2-azaadamantane-N-oxyl, 9-azanoradamantane-N-oxyl, and the like, with 9-azanoradamantane-N-oxyl being preferred.

[0250] The cocatalyst to be used includes potassium bromide, tetrabutylammonium bromide, etc., and is preferably potassium bromide.

[0251] The oxidizing agent to be used includes sodium hypochlorite, iodobenzene diacetate, etc., with sodium hypochlorite being preferred.

[0252] The reaction temperature varies depending on the oxidation catalyst and cocatalyst used, but is usually from 0°C to 50°C, preferably from 0°C to 10°C.

[0253] The reaction time varies depending on the types and amounts of the oxidation catalyst and cocatalyst used, but is usually from 10 minutes to 24 hours, preferably from 1 hour to 3 hours.

[0254] After completion of the reaction, a toluene solution of the target compound (9) of this reaction can be obtained, for example, by allowing the reaction solution to stand and then removing the aqueous layer to separate the organic layer containing the target compound, followed by washing with water, and can be used as is in the next step.

[0255] The resulting compound can be further purified, if necessary, by a conventional method such as recrystallization or silica gel chromatography. (C-6 process) This step is a step of producing compound (10) by stereoselectively hydroxymethylating the 4-carbon of compound (9) produced in step C-5 in the presence of a base and an alkylating agent in a solvent, accompanied by epimerization.

[0256] Examples of the solvent to be used include amides such as dimethylacetamide and dimethylformamide, hydrocarbons such as benzene and toluene, halogenated hydrocarbons such as methylene chloride, chloroform and 1,2-dichloroethane, ethers such as tetrahydrofuran and dimethyl ether, and esters such as methyl acetate, ethyl acetate and propyl acetate, and toluene is preferred.

[0257] Examples of the base to be used include inorganic bases such as sodium hydroxide, potassium hydroxide, sodium bicarbonate, and sodium carbonate, and organic bases such as triethylamine, pyridine, and 1,8-diazabicyclo[5.4.0]undec-7-ene, and preferably 1,8-diazabicyclo[5.4.0]undec-7-ene.

[0258] Examples of the alkylating agent that can be used include paraformaldehyde and an aqueous formaldehyde solution, with an aqueous formaldehyde solution being preferred.

[0259] The reaction temperature varies depending on the base used, but is usually from 0°C to 50°C, preferably from 20°C to 30°C.

[0260] The reaction time varies depending on the type and amount of base used, but is usually from 10 minutes to 24 hours, preferably from 1 hour to 3 hours.

[0261] After the reaction is completed, the toluene solution of the target compound (10) of this reaction can be obtained by, for example, leaving the reaction solution to stand, removing the aqueous layer, separating the organic layer containing the target compound, and washing with water, and can be used as it is in the next step. The cyclized product (11) partially produced in this step is converted to compound (12) in the next step.

[0262] The resulting compound can be further purified, if necessary, by a conventional method such as recrystallization or silica gel chromatography. (C-7 process) In this step, compound (10) produced in step C-6 is reacted with a hydride reducing agent in a solvent to produce compound (12) in which the hydroxyl group at the 3-position is sterically controlled.

[0263] Examples of the solvent to be used include halogenated hydrocarbons such as methylene chloride, chloroform, and 1,2-dichloroethane, hydrocarbons such as benzene and toluene, and ethers such as tetrahydrofuran and dimethyl ether, with toluene being preferred.

[0264] Examples of the hydride reducing agent to be used include sodium bis(2-methoxyethoxy)aluminum hydride, lithium aluminum hydride, diisobutylaluminum hydride, and sodium borohydride, and preferably sodium borohydride.

[0265] The reaction temperature varies depending on the hydride reducing agent used, but is usually from 0°C to 50°C, preferably from 20°C to 30°C.

[0266] The reaction time varies depending on the type and amount of the hydride reducing agent used, but is usually from 10 minutes to 24 hours, preferably from 1 hour to 3 hours.

[0267] After completion of the reaction, a toluene solution of the target compound (12) of this reaction can be obtained, for example, by leaving the reaction solution at rest and removing the aqueous layer to separate the organic layer containing the target compound, followed by washing with water, and can be used as it is in the next step.

[0268] The resulting compound can be further purified, if necessary, by a conventional method such as recrystallization or silica gel chromatography. (C-8 process) In this step, a hydroxyl-protecting reagent is reacted in a solvent in the presence of a base and a catalyst to protect the two hydroxyl groups of compound (12) produced in step C-7, thereby producing compound (13).

[0269] Examples of the solvent to be used include halogenated hydrocarbons such as methylene chloride, chloroform, and 1,2-dichloroethane, hydrocarbons such as benzene and toluene, and ethers such as tetrahydrofuran and dimethyl ether, with toluene being preferred.

[0270] Examples of the hydroxyl group-protecting reagent to be used include trityl chloride, t-butyldiphenylsilyl chloride, t-butyldimethylsilyl chloride, acetic anhydride, benzoyl chloride, and the like. Preferred is benzyl chloride or benzyl bromide, and more preferred is benzyl bromide.

[0271] The catalyst used may be, for example, tetrabutylammonium iodide, potassium iodide, sodium iodide, etc., with tetrabutylammonium iodide being preferred.

[0272] Examples of the base to be used include inorganic bases such as sodium hydroxide, potassium hydroxide, sodium bicarbonate, and sodium carbonate, and organic bases such as triethylamine, pyridine, and 1,8-diazabicyclo[5.4.0]undec-7-ene, and potassium hydroxide is preferred.

[0273] The reaction temperature varies depending on the hydroxyl-protecting reagent used, but is usually 20°C to 100°C, preferably 60°C to 80°C.

[0274] The reaction time varies depending on the type and amount of the hydroxyl-protecting reagent used, but is usually 1 to 48 hours, preferably 10 to 24 hours.

[0275] After completion of the reaction, a toluene solution of the target compound (13) of this reaction can be obtained, for example, by leaving the reaction solution to stand and then removing the aqueous layer to separate an organic layer containing the target compound, followed by washing with water. The obtained toluene layer can be subjected to solvent replacement with 1-propanol to cause crystallization.

[0276] The resulting compound can be further purified, if necessary, by a conventional method such as recrystallization or silica gel chromatography.

[0277] The above compound (13A) (when n=2 to 4) can be produced by Method D described below. (D method)

[0278] [ka]

[0279] In Method D, E represents ethylene, trimethylene, or tetramethylene, and Y and Z 1 and Z 2 has the same meaning as above.

[0280] Each step of Method D will be described in detail below. (D-1 process) This step is a step of producing compound (13A') by reacting compound (1D), produced according to the production method of compound (3b) in WO00 / 47599, with a hydroxyl-protecting reagent in a solvent in the presence of a base.

[0281] Examples of the solvent to be used include amides such as dimethylacetamide and dimethylformamide, hydrocarbons such as benzene and toluene, halogenated hydrocarbons such as methylene chloride, chloroform and 1,2-dichloroethane, ethers such as tetrahydrofuran and dimethyl ether, and esters such as methyl acetate, ethyl acetate and propyl acetate, and toluene is preferred.

[0282] Examples of the base to be used include triethylamine, pyridine, dimethylaminopyridine, 1-methylimidazole, and 4-methylmorpholine, with 4-methylmorpholine being preferred.

[0283] Examples of the hydroxyl-protecting reagent that can be used include trityl chloride, 4,4'-dimethoxytrityl chloride, etc., with trityl chloride being preferred.

[0284] The reaction temperature varies depending on the protecting reagent used, but is usually 0°C to 50°C, preferably 20°C to 30°C.

[0285] The reaction time varies depending on the type and amount of the protecting reagent used, but is usually from 10 minutes to 24 hours, preferably from 1 hour to 5 hours.

[0286] After completion of the reaction, the target compound (13A') of this reaction can be obtained, for example, by adding water to the reaction mixture, separating the organic layer containing the target compound, washing it with water, and then distilling off the solvent.

[0287] The resulting compound can be further purified, if necessary, by a conventional method such as recrystallization or silica gel chromatography.

[0288] Compound (5E) of the present invention can be produced by Method E described below. (Law E)

[0289] [ka]

[0290] E method, R 1 , R 2 , X 1 , Z 1 , Z 2 and P 1 has the same meaning as above.

[0291] The steps of Method E are explained in detail below. (E-1 process) This step is a step of producing compound (2E) by reacting compound (2A) with an activating agent in a solvent in the presence of a base.

[0292] Examples of the solvent to be used include halogenated hydrocarbons such as methylene chloride, chloroform, and 1,2-dichloroethane; hydrocarbons such as benzene and toluene; ethers such as tetrahydrofuran and dimethyl ether; and acetonitrile. Of these, toluene or acetonitrile is preferred.

[0293] Examples of the activating agent used include acetic anhydride, benzoic anhydride, trichloroacetonitrile, carbonyldiimidazole, and diphenyl chlorophosphate, and preferably acetic anhydride or trichloroacetonitrile.

[0294] The base to be used varies depending on the activating agent, but examples of the base include organic bases such as triethylamine, pyridine, and 1,8-diazabicyclo[5.4.0]undec-7-ene, and preferably pyridine and 1,8-diazabicyclo[5.4.0]undec-7-ene. After completion of the reaction, the target compound (2E) of this reaction can be used directly for glycosylation, or can be obtained by neutralizing the reaction solution, concentrating the reaction mixture, and distilling off the solvent.

[0295] The reaction temperature varies depending on the activating agent used, but is usually from 0°C to 50°C, preferably from 0°C to 30°C.

[0296] The reaction time varies depending on the type and amount of the activating agent used, but is usually from 10 minutes to 24 hours, preferably from 1 hour to 3 hours.

[0297] The resulting compound can be further purified, if necessary, by a conventional method such as recrystallization or silica gel chromatography. (E-2 process) This step is a step of producing compound (3E) by glycosylation of compound (2E) obtained in step E-1 with a pyrimidine base (such as thymine or cytosine protected by an acyl group) that has been silylated by a silylation reaction in a solvent in the presence of a halogenating agent.

[0298] Examples of solvents used in the silylation reaction and the glycosylation reaction include halogenated hydrocarbons such as methylene chloride, chloroform, and 1,2-dichloroethane, hydrocarbons such as benzene and toluene, ethers such as tetrahydrofuran and dimethyl ether, and acetonitrile, with acetonitrile being preferred.

[0299] Examples of the silylating agent to be used include N,O-bistrimethylsilylacetamide, hexamethyldisilazane, and the like, with N,O-bistrimethylsilylacetamide being preferred.

[0300] The temperature for the silylation reaction of the pyrimidine base varies depending on the silylating agent, but is usually 0°C to 50°C, preferably 20°C to 40°C.

[0301] The reaction time for the silylation reaction varies depending on the type and amount of the silylating agent used, but is usually from 10 minutes to 24 hours, preferably from 1 hour to 10 hours.

[0302] Examples of the halogenating agent used in the glycosylation reaction include chlorotrimethylsilane (TMSCl), bromotrimethylsilane (TMSBr), and iodotrimethylsilane (TMSI), with TMSI being preferred.

[0303] The reaction temperature for the glycosylation reaction varies depending on the structure of compound (2E) and the halogenating agent used, but is usually 0°C to 50°C, preferably 20°C to 40°C.

[0304] The reaction time for the glycosylation reaction varies depending on the type and amount of halogenating agent used, but is usually from 10 minutes to 24 hours, preferably from 10 hours to 20 hours.

[0305] After completion of the reaction, the target compound (3E) of this reaction can be obtained, for example, by neutralizing the reaction mixture, concentrating the reaction mixture, and distilling off the solvent.

[0306] The resulting compound can be further purified, if necessary, by a conventional method such as recrystallization or silica gel chromatography. (E-3 process) This step is a step in which compound (3E) is reacted with a deprotecting reagent for the hydroxyl group in a solvent to produce compound (4E) by deprotection reaction. (i)R 2 is a hydroxyl group, Z 1 and Z 2 When is a benzyl group, the deprotection reagent is a carbon supported metal catalyst and a reducing agent.

[0307] Examples of the solvent to be used include hydrocarbons such as benzene and toluene, ethers such as tetrahydrofuran and dimethyl ether, and alcohols such as methanol, ethanol and propanol, with methanol being preferred.

[0308] The metal catalyst to be used includes palladium, palladium hydroxide, platinum and the like, with palladium being preferred.

[0309] The reducing agent used may be hydrogen, formic acid, ammonium formate, etc., with hydrogen being preferred.

[0310] The reaction temperature varies depending on the metal catalyst, but is usually from 0°C to 70°C, preferably from 40°C to 60°C.

[0311] The reaction time varies depending on the type and amount of the metal catalyst and reducing agent used, but is usually from 10 minutes to 24 hours, preferably from 1 hour to 5 hours.

[0312] After completion of the reaction, the target compound (4E) of this reaction can be obtained, for example, by filtering the reaction mixture to remove the metal catalyst, concentrating the reaction mixture, and distilling off the solvent.

[0313] The resulting compound can be further purified, if necessary, by a conventional method such as recrystallization or silica gel chromatography. (ii)R 2 is an amino group protected with an acyl group, Z 1 and Z 2 When is a benzyl group, a deprotection reaction is carried out using a deprotection reagent.

[0314] The solvent to be used may include, for example, halogenated hydrocarbons such as methylene chloride, chloroform, and 1,2-dichloroethane, and hydrocarbons such as benzene and toluene, with methylene chloride being preferred.

[0315] The deprotecting reagent to be used includes iron(III) chloride, boron trichloride, etc., and preferably boron trichloride.

[0316] The reaction temperature varies depending on the deprotection reagent, but is usually -20°C to 30°C, preferably -20°C to 20°C.

[0317] The reaction time varies depending on the deprotection reagent, but is usually from 10 minutes to 10 hours, preferably from 1 hour to 5 hours.

[0318] After completion of the reaction, the target compound (4E) of this reaction can be obtained, for example, by neutralizing the reaction mixture, concentrating the reaction mixture, and distilling off the solvent.

[0319] The resulting compound can be further purified, if necessary, by a conventional method such as recrystallization or silica gel chromatography. (E-4 process) This step is a step of producing compound (5E) by reacting compound (4E) with a protecting reagent for the primary hydroxyl group in a solvent in the presence of a base.

[0320] Examples of the solvent to be used include halogenated hydrocarbons such as methylene chloride, chloroform, and 1,2-dichloroethane; hydrocarbons such as benzene and toluene; ethers such as tetrahydrofuran and dimethyl ether; esters such as ethyl acetate and propyl acetate; and acetonitrile. Among these, tetrahydrofuran or ethyl acetate is preferred.

[0321] Examples of the protecting reagent for the primary hydroxyl group that can be used include trityl chloride, 4-methoxytrityl chloride, 4,4'-dimethoxytrityl chloride, etc., with 4,4'-dimethoxytrityl chloride being preferred.

[0322] The base to be used includes aliphatic amines such as triethylamine and N-methylmorpholine, and aromatic amines such as imidazole and pyridine, and is preferably pyridine.

[0323] The reaction temperature varies depending on the protecting reagent, but is usually 0°C to 50°C, preferably 0°C to 20°C.

[0324] The reaction time varies depending on the protecting reagent, but is usually from 10 minutes to 10 hours, preferably from 1 hour to 5 hours.

[0325] After completion of the reaction, the target compound (5E) of this reaction can be obtained, for example, by neutralizing the reaction mixture, concentrating the reaction mixture, and distilling off the solvent.

[0326] The resulting compound can be further purified, if necessary, by a conventional method such as recrystallization or silica gel chromatography.

[0327] The compound (4F) of the present invention can be produced by Method F described below. (F method)

[0328] [ka]

[0329] F method, R 1 , R 6 , Z 3 and P 1 has the same meaning as above.

[0330] Each step of Method F will be described in detail below. (F-1 process) This step is a step of producing compound (1F) by reacting compound (5E') with a hydroxyl-protecting reagent in a solvent in the presence of a base and a catalyst.

[0331] Examples of the solvent to be used include halogenated hydrocarbons such as methylene chloride, chloroform, and 1,2-dichloroethane, hydrocarbons such as benzene and toluene, ethers such as tetrahydrofuran and dimethyl ether, and acetonitrile, with acetonitrile being preferred.

[0332] Examples of the hydroxyl-protecting reagent that can be used include acetic anhydride, acetyl chloride, benzoic anhydride, and benzoyl chloride, with acetic anhydride being preferred.

[0333] The base used is Hydroxyl group protecting reagent Although it varies depending on the type, examples of the base include organic bases such as triethylamine and pyridine, with triethylamine being preferred.

[0334] Examples of the catalyst used include N,N-dimethylaminopyridine, 1,8-diazabicyclo[5.4.0]undec-7-ene, etc., and N,N-dimethylaminopyridine is preferred. The reaction temperature varies depending on the base used, but is usually 0° C. to 50° C., and preferably 0° C. to 30° C.

[0335] The reaction time varies depending on the type and amount of the protecting reagent used, but is usually from 10 minutes to 24 hours, preferably from 1 hour to 3 hours.

[0336] After completion of the reaction, the target compound (1F) of this reaction can be obtained, for example, by neutralizing the reaction mixture, concentrating the reaction mixture, and distilling off the solvent.

[0337] The resulting compound can be further purified, if necessary, by a conventional method such as recrystallization or silica gel chromatography. (F-2 process) This step is a step of activating the hydroxyl group of compound (1F) using an activating agent in a solvent in the presence of a base and a catalyst, and then reacting the compound with an aminating agent to produce compound (2F).

[0338] Examples of the solvent to be used include halogenated hydrocarbons such as methylene chloride, chloroform, and 1,2-dichloroethane, hydrocarbons such as benzene and toluene, ethers such as tetrahydrofuran and dimethyl ether, and acetonitrile, with acetonitrile being preferred.

[0339] The base to be used varies depending on the activating agent, but examples thereof include organic bases such as triethylamine and pyridine, with triethylamine being preferred.

[0340] The catalyst used may include N,N-dimethylaminopyridine, 1,8-diazabicyclo[5.4.0]undec-7-ene, etc., and is preferably N,N-dimethylaminopyridine.

[0341] The activating agent used may be p-toluenesulfonyl chloride, 2,4,6-triisopropylbenzenesulfonyl chloride, or the like, with 2,4,6-triisopropylbenzenesulfonyl chloride being preferred.

[0342] The reaction temperature varies depending on the activating agent used, but is usually from 0°C to 50°C, preferably from 0°C to 30°C.

[0343] The reaction time varies depending on the type and amount of the activating agent used, but is usually from 10 minutes to 8 hours, preferably from 1 hour to 3 hours.

[0344] The aminating agent used may include ammonia, an aqueous ammonia solution, or an ammonia salt such as ammonium carbonate or ammonium acetate, and is preferably an aqueous ammonia solution.

[0345] The reaction temperature varies depending on the aminating agent used, but is usually from 0°C to 50°C, preferably from 0°C to 30°C.

[0346] The reaction time varies depending on the type and amount of the aminating agent used, but is usually from 10 minutes to 8 hours, preferably from 1 hour to 3 hours.

[0347] After completion of the reaction, the target compound (2F) of this reaction can be obtained, for example, by neutralizing the reaction mixture, concentrating the reaction mixture, and distilling off the solvent.

[0348] The resulting compound can be further purified, if necessary, by a conventional method such as recrystallization or silica gel chromatography. (F-3 process) This step is a step of producing compound (3F) by acylation of the amino group of compound (2F) with an acylating agent in a solvent in the presence of a base.

[0349] Examples of the solvent to be used include halogenated hydrocarbons such as methylene chloride, chloroform, and 1,2-dichloroethane, hydrocarbons such as benzene and toluene, ethers such as tetrahydrofuran and dimethyl ether, and acetonitrile, with acetonitrile being preferred.

[0350] Examples of the base to be used include inorganic bases such as sodium hydroxide, potassium hydroxide, sodium bicarbonate, and sodium carbonate, and organic bases such as triethylamine, pyridine, and 1,8-diazabicyclo[5.4.0]undec-7-ene, and preferably sodium hydroxide.

[0351] The acylating agent to be used includes benzoyl chloride, benzoic anhydride, etc., and is preferably benzoic anhydride.

[0352] The reaction temperature varies depending on the acylating agent used, but is usually from 0°C to 50°C, preferably from 10°C to 40°C.

[0353] The reaction time varies depending on the type and amount of the acylating agent used, but is usually from 1 to 48 hours, preferably from 2 to 20 hours.

[0354] After completion of the reaction, the target compound (3F) of this reaction can be obtained, for example, by neutralizing the reaction mixture, concentrating the reaction mixture, and distilling off the solvent.

[0355] The resulting compound can be further purified, if necessary, by a conventional method such as recrystallization or silica gel chromatography. (F-4 process) This step is a step of producing compound (4F) by selectively hydrolyzing the acyl group at position 3 of compound (3F) with a base in a solvent.

[0356] Examples of the solvent to be used include alcohols such as methanol, ethanol, and propanol, water, and acetonitrile, with water and acetonitrile being preferred.

[0357] Examples of the base to be used include inorganic bases such as sodium hydroxide, potassium hydroxide, sodium bicarbonate, and sodium carbonate, with sodium hydroxide being preferred.

[0358] The reaction temperature varies depending on the base used, but is usually from 0°C to 50°C, preferably from 0°C to 30°C.

[0359] The reaction time varies depending on the base used, but is usually from 10 minutes to 24 hours, preferably from 1 hour to 3 hours.

[0360] After completion of the reaction, the target compound (4F) of this reaction can be obtained, for example, by neutralizing the reaction mixture, concentrating the reaction mixture, and distilling off the solvent.

[0361] The resulting compound can be further purified, if necessary, by a conventional method such as recrystallization or silica gel chromatography.

[0362] Compound (3G) of the present invention can be produced by Method G described below. (G method)

[0363] [ka]

[0364] During G method, R 3 , X 2 , Z 1 , Z 2 and P 1 has the same meaning as above.

[0365] The steps of the G method are explained in detail below. (G-1 process) This step is a step of producing compound (1G) by reacting compound (2A) with an activating agent in a solvent in the presence of a base.

[0366] Examples of the solvent to be used include halogenated hydrocarbons such as methylene chloride, chloroform, and 1,2-dichloroethane, hydrocarbons such as benzene and toluene, ethers such as tetrahydrofuran and dimethyl ether, and acetonitrile, and preferably toluene and acetonitrile.

[0367] The reaction temperature varies depending on the activating agent used, but is usually from 0°C to 50°C, preferably from 0°C to 30°C.

[0368] Examples of the activating agent that can be used include acetic anhydride, benzoic anhydride, trichloroacetonitrile, carbonyldiimidazole, and diphenyl chlorophosphate, and preferably, acetic anhydride and benzoic anhydride.

[0369] The base to be used varies depending on the activating agent, but examples thereof include organic bases such as triethylamine, pyridine, N,N-dimethylaminopyridine, and 1,8-diazabicyclo[5.4.0]undec-7-ene, with N,N-dimethylaminopyridine being preferred.

[0370] The reaction temperature varies depending on the activating agent used, but is usually from 0°C to 50°C, preferably from 0°C to 30°C.

[0371] The reaction time varies depending on the type and amount of the activating agent used, but is usually from 10 minutes to 8 hours, preferably from 1 hour to 3 hours.

[0372] After completion of the reaction, the target compound (1G) of this reaction can be used, for example, directly for glycosylation or can be obtained by neutralizing the reaction solution, concentrating the reaction mixture, and distilling off the solvent.

[0373] The resulting compound can be further purified, if necessary, by a conventional method such as recrystallization or silica gel chromatography. (G-2 process) This step involves carrying out a glycosylation reaction of compound (1G) with a protected 6-aminopurin-9-yl group in a solvent in the presence of an acid reagent, followed by isomerization to produce compound (2G).

[0374] Examples of the solvent to be used include halogenated hydrocarbons such as methylene chloride, chloroform, and 1,2-dichloroethane, hydrocarbons such as benzene and toluene, ethers such as tetrahydrofuran, and acetonitrile, with acetonitrile being preferred.

[0375] Examples of the acid reagent to be used include dichlorodimethylsilane and trifluoromethanesulfonic acid, trimethylsilyl trifluoromethanesulfonate and trifluoromethanesulfonic acid, methanesulfonic acid, and trifluoroacetic acid, and preferably trimethylsilyl trifluoromethanesulfonate and trifluoroacetic acid.

[0376] The reaction temperature for glycosylation and isomerization varies depending on the structure of compound (1G) and the acid reagent used, but is usually 30°C to 70°C, preferably 40°C to 60°C.

[0377] The reaction time varies depending on the type and amount of Lewis acid reagent used, but is usually from 10 minutes to 24 hours, preferably from 1 hour to 5 hours.

[0378] After completion of the reaction, the target compound (2G) of this reaction can be obtained, for example, by neutralizing the reaction mixture, concentrating the reaction mixture, and distilling off the solvent.

[0379] The resulting compound can be further purified, if necessary, by a conventional method such as recrystallization or silica gel chromatography. (G-3 process) In this step, compound (2G) is reacted with a deprotection reagent in a solvent to give Z 1 and Z 2 This is a process for producing compound (3G) by deprotecting the compound (3G).

[0380] The solvent to be used may include, for example, halogenated hydrocarbons such as methylene chloride, chloroform, and 1,2-dichloroethane, and hydrocarbons such as benzene and toluene, with methylene chloride being preferred.

[0381] Z 1 and Z 2 When is a benzyl group, examples of the deprotecting reagent include iron(III) chloride, boron trichloride, etc., and preferably boron trichloride.

[0382] The reaction temperature varies depending on the deprotection reagent, but is usually -20°C to 30°C, preferably -20°C to 20°C.

[0383] The reaction time varies depending on the deprotection reagent, but is usually from 10 minutes to 10 hours, preferably from 1 hour to 5 hours.

[0384] After completion of the reaction, the target compound (3G) of this reaction can be obtained, for example, by neutralizing the reaction mixture, concentrating the reaction mixture, and distilling off the solvent.

[0385] The resulting compound can be further purified, if necessary, by a conventional method such as recrystallization or silica gel chromatography. (G-4 process) The target compound (4G) of this step can be prepared by protecting the primary hydroxyl group of compound (3G) according to Step E-4 of Method E.

[0386] Moreover, compound (5H) of the present invention can also be produced by Method H (glycosylation using dichloropurine) described below. (H method)

[0387] [ka]

[0388] During H method, R 3 , Z 1 , Z 2 and P 1 has the same meaning as above.

[0389] The steps of the H method are explained in detail below. (H-1 process) This step is a step of producing compound (1H) by glycosylation reaction of compound (1G) with dichloropurine that has been silylated by a silylation reaction in a solvent in the presence of a halogenating agent.

[0390] Examples of solvents used in the glycosylation reaction and the silylation reaction include halogenated hydrocarbons such as methylene chloride, chloroform, and 1,2-dichloroethane, hydrocarbons such as benzene and toluene, ethers such as tetrahydrofuran, and acetonitrile, with acetonitrile being preferred.

[0391] Examples of the silylating agent used in the silylation reaction include N,O-bistrimethylsilylacetamide, hexamethyldisilazane, and the like, with N,O-bistrimethylsilylacetamide being preferred.

[0392] The reaction temperature for the silylation reaction of dichloropurine varies depending on the silylating agent, but is usually from 0°C to 90°C, preferably from 50°C to 80°C.

[0393] The reaction time for the silylation reaction varies depending on the type and amount of the silylating agent used, but is usually from 10 minutes to 24 hours, preferably from 1 hour to 10 hours.

[0394] Examples of halogenating agents used in the glycosylation reaction include chlorotrimethylsilane (TMSCl), bromotrimethylsilane (TMSBr), and iodotrimethylsilane (TMSI), and TMSI is preferred. The reaction temperature for the glycosylation reaction varies depending on the structure of compound (2B) and the halogenating agent used, but is usually 0° C. to 90° C., and preferably 50° C. to 80° C.

[0395] The reaction time for the glycosylation reaction varies depending on the type and amount of halogenating agent used, but is usually from 10 minutes to 24 hours, preferably from 10 hours to 5 hours.

[0396] After completion of the reaction, the target compound (1H) of this reaction can be obtained, for example, by neutralizing the reaction mixture, concentrating the reaction mixture, and distilling off the solvent.

[0397] The resulting compound can be further purified, if necessary, by a conventional method such as recrystallization or silica gel chromatography. (H-2 process) This step is a step of producing compound (2H) by reacting compound (1H) with an aminating agent in a solvent.

[0398] Examples of the solvent to be used include halogenated hydrocarbons such as methylene chloride, chloroform, and 1,2-dichloroethane, hydrocarbons such as benzene and toluene, ethers such as tetrahydrofuran and dimethyl ether, and acetonitrile, and preferably tetrahydrofuran.

[0399] The aminating agent used may include ammonia, an aqueous ammonia solution, or an ammonia salt such as ammonium carbonate or ammonium acetate, and is preferably an aqueous ammonia solution.

[0400] The reaction temperature is usually from 0°C to 90°C, preferably from 30°C to 60°C.

[0401] The reaction time varies depending on the amount of the solvent and the aminating agent used, but is usually from 10 minutes to 24 hours, preferably from 1 hour to 10 hours.

[0402] After completion of the reaction, the target compound (2H) of this reaction can be obtained, for example, by neutralizing the reaction mixture, concentrating the reaction mixture, and distilling off the solvent.

[0403] The resulting compound can be further purified, if necessary, by a conventional method such as recrystallization or silica gel chromatography. (H-3 process) In this step, Z of compound (2H) is reacted in a solvent. 1 and Z 2 and hydrogenating the chlorine atom at the 2-position of the purine ring to produce compound (3H).

[0404] Examples of the solvent to be used include water, alcohols such as methanol, ethanol, 1-propanol, and 2-propanol, hydrocarbons such as benzene and toluene, ethers such as tetrahydrofuran and diethyl ether, and acetonitrile, and preferably ethanol.

[0405] Z 1 and Z 2 When is a benzyl group, Z is converted to 1 and Z 2 The deprotection reaction and the hydrogenation reaction at the 2-position of the purine ring can be carried out simultaneously.

[0406] The metal catalyst used may be palladium, palladium hydroxide, platinum (particularly, palladium supported on carbon, palladium hydroxide, or platinum), and is preferably palladium (particularly, palladium supported on carbon).

[0407] The reducing agent used may be hydrogen, formic acid, ammonium formate, etc., with hydrogen being preferred.

[0408] The reaction temperature varies depending on the metal catalyst, but is usually from 0°C to 70°C, preferably from 40°C to 60°C.

[0409] The reaction time varies depending on the type and amount of the metal catalyst and reducing agent used, but is usually from 10 minutes to 24 hours, preferably from 1 hour to 10 hours.

[0410] After completion of the reaction, the target compound (3H) of this reaction can be obtained, for example, by filtering the reaction mixture to remove the metal catalyst, concentrating the reaction mixture, and distilling off the solvent.

[0411] The resulting compound can be further purified, if necessary, by a conventional method such as recrystallization or silica gel chromatography. (H-4 process) In this step, the hydroxyl group of compound (3H) is protected with a hydroxyl-protecting reagent in a solvent, the amino group on the purine ring is acylated with an acylating agent, and the protecting group on the hydroxyl group is then removed with ammonia to produce compound (4H).

[0412] Examples of the solvent to be used include pyridine and acetonitrile, with pyridine being preferred.

[0413] Examples of the hydroxyl group-protecting reagent that can be used include chlorotrimethylsilane, trifluoromethanesulfonyltrimethylsilane, etc., with chlorotrimethylsilane being preferred.

[0414] The reaction temperature is usually from 0°C to 90°C, preferably from 0°C to 30°C.

[0415] The reaction time varies depending on the type and amount of the hydroxyl-protecting reagent used, but is usually from 10 minutes to 2 hours, preferably from 30 minutes to 1 hour.

[0416] The acylating agent to be used includes acetic anhydride, acetyl chloride, benzoic anhydride, benzoyl chloride, etc., and is preferably benzoyl chloride.

[0417] The temperature for the acylation reaction is usually 0°C to 90°C, preferably 0°C to 30°C.

[0418] The reaction time varies depending on the type and amount of the hydroxyl-protecting reagent used, but is usually 1 to 8 hours, preferably 1 to 3 hours.

[0419] The target compound (4H) of this reaction can be obtained, for example, by adding aqueous ammonia to the reaction mixture, concentrating the reaction mixture, and distilling off the solvent.

[0420] The resulting compound can be further purified, if necessary, by a conventional method such as recrystallization or silica gel chromatography. (H-5 process) The target compound (5H) of this step can be prepared by protecting the primary hydroxyl group of compound (4H) according to Step E-4 of Method E.

[0421] The compound (4I) of the present invention can be prepared by Method I described below. (I Method)

[0422] [ka]

[0423] In Law I, R 4 , R 5 , Z 1 , Z 2 and P 1 has the same meaning as above.

[0424] Each step of Method I will be described in detail below. (I-1 process) This step is a step of producing compound (1I) by subjecting compound (1H) to a substitution reaction with an optionally substituted benzyl alcohol in a solvent in the presence of a base.

[0425] Examples of the benzyl alcohol which may have a substituent include benzyl alcohol, 4-methylbenzyl alcohol, 2,4,6-trimethylbenzyl alcohol, 3,4,5-trimethylbenzyl alcohol, 4-methoxybenzyl alcohol, 4-chlorobenzyl alcohol, 4-bromobenzyl alcohol, and 4-cyanobenzyl alcohol, and benzyl alcohol is preferred.

[0426] Examples of the solvent to be used include halogenated hydrocarbons such as methylene chloride, chloroform, and 1,2-dichloroethane, hydrocarbons such as benzene and toluene, ethers such as tetrahydrofuran and dimethyl ether, and acetonitrile, and preferably tetrahydrofuran.

[0427] Examples of the base to be used include inorganic bases such as sodium hydride and sodium carbonate, and organic bases such as triethylamine, pyridine and 1,8-diazabicyclo[5.4.0]undec-7-ene, and preferably sodium hydride.

[0428] The reaction temperature is usually from 0°C to 90°C, preferably from 0°C to 30°C.

[0429] The reaction time varies depending on the amount of the solvent and the base used, but is usually from 10 minutes to 24 hours, preferably from 1 hour to 10 hours.

[0430] After completion of the reaction, the target compound (1I) of this reaction can be obtained, for example, by neutralizing the reaction mixture, concentrating the reaction mixture, and distilling off the solvent.

[0431] The resulting compound can be further purified, if necessary, by a conventional method such as recrystallization or silica gel chromatography. (I-2 process) This step is a step of producing compound (2I) by a cross-coupling reaction of compound (1I) with an amidating agent in a solvent in the presence of a base, a palladium catalyst and a phosphine ligand.

[0432] Examples of the solvent to be used include halogenated hydrocarbons such as methylene chloride, chloroform, and 1,2-dichloroethane, hydrocarbons such as benzene and toluene, ethers such as tetrahydrofuran and dimethyl ether, and acetonitrile, and toluene is preferred.

[0433] Examples of the base to be used include inorganic bases such as sodium hydroxide, sodium carbonate, and cesium carbonate, and organic bases such as triethylamine, pyridine, and 1,8-diazabicyclo[5.4.0]undec-7-ene, with cesium carbonate being preferred.

[0434] The palladium catalyst used includes tris(dibenzylideneacetone)(chloroform)dipalladium, palladium(II) acetate, tris(dibenzylideneacetone)dipalladium(0) and the like, with tris(dibenzylideneacetone)(chloroform)dipalladium being preferred.

[0435] The phosphine ligand to be used includes 4,5'-bis(diphenylphosphino)-9,9'-dimethylxanthene, 1,1'-bis(diphenylphosphino)ferrocene, 1,2-bis(diphenylphosphino)ethane, 2-dicyclohexylphosphino-2'-(N,N-dimethylamino)biphenyl, and the like, and is preferably 4,5'-bis(diphenylphosphino)-9,9'-dimethylxanthene.

[0436] Examples of the amidating agent that can be used include acetylamide, benzoylamide, and isobutylamide, with isobutylamide being preferred.

[0437] The reaction temperature is usually from 20°C to 150°C, preferably from 90°C to 110°C.

[0438] The reaction time varies depending on the amount of the solvent and the amount of the palladium catalyst used, but is usually from 10 minutes to 24 hours, preferably from 5 hours to 15 hours.

[0439] After completion of the reaction, the target compound (2I) of this reaction can be obtained, for example, by neutralizing the reaction mixture, concentrating the reaction mixture, and distilling off the solvent.

[0440] The resulting compound can be further purified, if necessary, by a conventional method such as recrystallization or silica gel chromatography. (I-3 process) In this step, compound (2I) is reacted with a deprotection reagent for the hydroxyl group in a solvent to give Z 1 , Z 2 and R 5 This is a process for producing compound (3I) by deprotection of the above.

[0441] Z 1 and Z 2 When is a benzyl group, the deprotection reagent is a carbon supported metal catalyst and a reducing agent.

[0442] The metal catalyst to be used may be palladium, palladium hydroxide, platinum or the like, with palladium hydroxide being preferred.

[0443] The reducing agent used may be hydrogen, formic acid, ammonium formate, etc., with hydrogen being preferred.

[0444] The reaction temperature varies depending on the metal catalyst, but is usually from 0°C to 70°C, preferably from 40°C to 60°C.

[0445] The reaction time varies depending on the type and amount of the metal catalyst and reducing agent used, but is usually from 10 minutes to 24 hours, preferably from 1 hour to 10 hours.

[0446] After completion of the reaction, the target compound (3I) of this reaction can be obtained, for example, by filtering the reaction mixture to remove the metal catalyst, concentrating the reaction mixture, and distilling off the solvent.

[0447] The resulting compound can be further purified, if necessary, by a conventional method such as recrystallization or silica gel chromatography. (I-4 process) The target compound (4I) of this step can be prepared by protecting the primary hydroxyl group of compound (3I) according to Step E-4 of Method E.

[0448] Compound (2J) can be produced by Method J described below. (J method)

[0449] [ka]

[0450] J Law, P 1 and B are as defined above.

[0451] The steps of the J method are explained in detail below. (J-1 process) This step is a step in which compound (1J) is reacted with an amidite reagent in a solvent in the presence of a drying agent and an activating agent to produce compound (2J) by amidite reaction.

[0452] Examples of the solvent to be used include halogenated hydrocarbons such as methylene chloride, chloroform, and 1,2-dichloroethane; esters such as ethyl acetate, propyl acetate, and butyl acetate; hydrocarbons such as benzene and toluene; ethers such as tetrahydrofuran; and acetonitrile. Of these, methylene chloride is preferred.

[0453] The drying agent used may be molecular sieve 3A, molecular sieve 4A, or molecular sieve 5A, and preferably molecular sieve 4A.

[0454] Examples of the activating agent to be used include 5-benzylthiotetrazole, 5-phenyltetrazole, dibromoimidazole, dicyanoimidazole, and N-alkylimidazole trifluoroacetate, and preferably 4,5-dicyanoimidazole.

[0455] Examples of the amiditizing reagent used include 2-cyanoethyl N,N,N',N'-tetraisopropyl phosphorodiamidite and 2-cyanoethyl diisopropyl chlorophosphoroamidite, and preferably 2-cyanoethyl N,N,N',N'-tetraisopropyl phosphorodiamidite.

[0456] The reaction temperature varies depending on the amidite reagent used, but is usually 0°C to 50°C, preferably 0°C to 30°C.

[0457] The reaction time varies depending on the amiditizing reagent used, but is usually from 10 minutes to 24 hours, preferably from 1 hour to 16 hours.

[0458] After completion of the reaction, the target compound (2J) of this reaction can be obtained, for example, by neutralizing the reaction mixture, concentrating the reaction mixture, and distilling off the solvent.

[0459] The resulting compound can be further purified, if necessary, by a conventional method such as recrystallization or silica gel chromatography. In one embodiment, the present invention provides a method for producing an oligonucleotide having a desired sequence and structure by linking each ENA monomer produced by the above method, a phosphoramidite compound of a nucleoside corresponding to a commercially available nucleic acid or modified nucleic acid, and a phosphoramidite compound of a ligand unit by a phosphodiester bond or a phosphorothioate bond to extend the unit chain. Such an oligonucleotide can be produced by synthesizing using a commercially available synthesizer (e.g., Perkin Elmer's Model 392 by phosphoramidite method) according to the method described in the literature (Nucleic Acids Research, 12, 4539 (1984)). The phosphoramidite compound of each ENA nucleoside (i.e., 2'-O,4'-C-ethylene-bridged guanosine, 2'-O,4'-C-ethylene-bridged adenosine, 2'-O,4'-C-ethylene-bridged cytidine, 2'-O,4'-C-ethylene-bridged thymidine) used in this case can be a compound produced by the above method. Phosphoramidite compounds of natural nucleosides and 2'-O-methyl nucleosides (i.e., 2'-O-methylguanosine, 2'-O-methyladenosine, 2'-O-methylcytidine, 2'-O-methyluridine) can be prepared using commercially available reagents.

[0460] In the present invention, the nucleic acid base sequence can be described as adenine (A) or (a), guanine (G) or (g), cytosine (C) or (c), thymine (T) or (t), and uracil (U) or (u). 5-methylcytosine can be used instead of cytosine. Among the nucleic acid bases, uracil (U) or (u) and thymine (T) or (t) are interchangeable. Either uracil (U) or (u) or thymine (T) or (t) can be used to form a base pair with adenine (A) or (a) in the complementary strand.

[0461] After coupling of a phosphoramidite compound, an oligonucleotide having a phosphorothioate bond can be synthesized by reacting with a reagent such as sulfur, tetraethylthiuram disulfide (TETD, Applied Biosystems), Beaucage reagent (Glen Research), or xanthan hydride (Tetrahedron Letters, 32, 3005 (1991), J. Am. Chem. Soc. 112, 1253(1990), PCT / WO98 / 54198).

[0462] The controlled pore glass (CPG) used in the synthesizer can be a commercially available product with 2'-O-methyl nucleoside attached. 2'-O, 4'-C-methyleneguanosine, adenosine, 5-methylcytidine and thymidine can be attached to CPG according to the method described in WO99 / 14226, and 2'-O, 4'-C-ethyleneguanosine, adenosine, 5-methylcytidine and thymidine produced by the above method can be attached to CPG according to the literature (Oligonucleotide Synthesis, Edited by MJGait, Oxford University Press, 1984). By using a modified CPG (described in Example 12b of JP-A-7-87982), an oligonucleotide with a 2-hydroxyethyl phosphate group attached to the 3' end can be synthesized. In addition, 3'-amino-Modifier C3 CPG, 3'-amino-Modifier C7 CPG, Glyceryl CPG, (Glen Research), 3'-specer C3SynBase CPG 1000, and 3'-specer C9 SynBase CPG 1000 (linktechnologies) can be used to synthesize oligonucleotides with a hydroxyalkyl phosphate group or an aminoalkyl phosphate group attached to the 3' end.

[0463] The oligonucleotides produced according to the present invention may contain a ligand unit suitable for transporting nucleic acids to tissues. The ligand unit can be attached to the 5' end of an oligonucleotide by synthesizing an amidite compound in which the ligand portion contains a phosphate portion via a linker, in a manner similar to that of nucleotide elongation. For transport to the liver, an oligonucleotide in which GalNAc is attached to the phosphate portion via a linker is used, and the following X 18 Or X 20 The phosphoramidite compounds corresponding to GalNAc units, X 18 The phosphoramidite compound corresponding to the above is compound 39D of Reference Example 39 of WO2019 / 172286 ([(2R,3R,4R,5R,6R)-5-acetamido-6-[3-[[2-[[3-[[2-[3-[(2R,3R,4R,5R,6R)-3-acetamido-4,5-diacetoxy-6-[[bis(4-methoxyphenyl)-phenyl-methoxy]methyl]tetrahydropyran-2-yl]oxypropylamino]-2-oxo-ethyl]carbamoyloxy]-2-[2-cyanoethoxy-(diisopropylamino)phosphanyl]oxy-propoxy]carbonylamino]acetyl]amino]propoxy]-4-acetoxy-2-[[bis(4-methoxyphenyl)-phenyl-methoxy]methyl]tetrahydropyran-3-yl] acetate),

[0464] [ka]

[0465] X 20The corresponding phosphoramidite compound is compound 41D of Reference Example 41 of WO2019 / 172286 ([(2R,3R,4R,5R,6R)-5-acetamido-6-[3-[3-[[3-[[3-[3-[(2R,3R,4R,5R,6R)-3-acetamido-4,5-diacetoxy-6-[[bis(4-methoxyphenyl)-phenyl-methoxy]methyl]tetrahydropyran-2-yl]oxypropylamino]-3-oxo-propyl]carbamoyloxy]-2-[2-cyanoethoxy-(diisopropylamino)phosphanyl]oxy-propoxy]carbonylamino]propanoylamino]propoxy]-4-acetoxy-2-[[bis(4-methoxyphenyl)-phenyl-methoxy]methyl]tetrahydropyran-3-yl] acetate),

[0466] [ka]

[0467] can be used, respectively. One embodiment of the oligonucleotide produced by the method of the present invention is an oligonucleotide having the sequence and structure represented by the following formula, which is useful for treating Duchenne muscular dystrophy (see WO2004 / 048570).

[0468] (DMD AO01) HO-C e2s -A m1s -G m1s -T e2s -T e2s -U m1s -G m1s -C e2s -C e2s -G m1s -C e2s -T e2s -G m1s -C e2s -C e2s -C e2s -A m1s -A m1s -CH2CH2OH (SEQ ID NO: 1) (DMD AO02) HO-Te2s -G m1s -T e2s -T e2s -C e2s -T e2s -G m1s -A m1s -C e2s -A m1s -A m1s -C e2s -A m1s -G m1s -T e2s -T e2s -T e2s -G m1s -CH2CH2OH (array number 2) (DMD AO03) HO-C e2s -G m1s -C e2s -T e2s -G m1s -C m1s -C e2s -C e2s -A m1s -A m1s -T e2s -G m1s -C e2s -C e2s -A m1s -U m1s -C e2s -C e2s -CH2CH2OH (array number 3) (DMD AO04) HO-C e2s -A m1s -T e2s -A m1s -A m1s -T e2s -G m1s -A m1s -A e2s -A m1s -A m1s -C e2s -G m1s -C m1s -C e2s -G m1s -C e2s -C e2s -CH2CH2OH (array number 4) (DMD AO05) HO-T e2s -U m1s -C e2s -C m1s -Ce2s -A m1s -A m1s -T e2s -U m1s -C m1s -T e2s -C e2s -A m1s -G m1s -G m1s -A e2s -A m1s -T e2s -CH2CH2OH (SEQ ID NO: 5) (DMD AO06) HO-C e2s -C e2s -A m1s -U m1s -T e2s -U m1s -G m1s -T e2s -A m1s -U m1s -T e2s -T e2s -A m1s -G m1s -C e2s -A m1s -T e2s -G m1s -CH2CH2OH (SEQ ID NO: 6) (DMD AO07) HO-G m1s -G m1s -C e2s -T e2s -G m1s -C m1s -T e2s -T e2s -U m1s -G m1s -C e2s -C m1s -C m1s -T e2s -C e2s -A m1s -G m1s -C e2s -CH2CH2OH (SEQ ID NO: 7) (DMD AO08) HO-G m1s -C e2s -T e2s -A m1s -G m1s -G m1s -T e2s -C e2s -Am1s -G m1s -G m1s -T e2s -G m1s -C m1s -T e2s -T e2s -U m1s -CH2CH2OH (SEQ ID NO: 8) (DMD AO09) HO-A m1s -C e2s -C e2s -G m1s -C m1s -C e2s -T e2s -U m1s -C m1s -C e2s -A m1s -C m1s -T e2s -C e2s -A m1s -G m1s -A e2s -G m1s -CH2CH2OH; (SEQ ID NO: 9) (DMD AO10) HO-G e2s -G e2s -C e2s -A e2s -T e2s -U m1s -U m1s -C m1s -U m1s -A m1s -G m1s -U m1s -U m1s -T e2s -G e2s -G e2s -A e2s -G e2s -CH2CH2OH (SEQ ID NO: 10) (DMD AO11) HO-G m1s -G m1s -C e2s -A m1s -T e2s -T e2s -U m1s -C e2s -T e2s -A m1s -G m1s -U m1s -T e2s -Te2s -G m1s -G m1s -A e2s -G m1s -CH2CH2OH (SEQ ID NO: 11) (DMD AO12) HO-A e2s -G m1s -T e2s -U m1s -T e2s -G m1s -G m1s -A e2s -G m1s -A m1s -T e2s -G m1s -G m1s -C e2s -A e2s -G m1s -T e2s -T e2s -CH2CH2OH (SEQ ID NO: 12) (DMD AO13) HO-C e2s -T e2s -C m1s -C e2s -T e2s -U m1s -C e2s -C e2s -A m1s -T e2s -G m1s -A m1s -C e2s -T e2s -C e2s -A m1s -A m1s -G m1s - CH2CH2OH (SEQ ID NO: 13) (DMD AO14) HO-C e2s -T e2s -G m1s -A m1s -A m1s -G m1s -G m1s -T e2s -G m1s -T e2s -T e2s -C e2s -T e2s -T e2s -G m1s -T e2s -Am1s - C e2s - CH2CH2OH (SEQ ID NO: 14) (DMD AO15) HO-T e2s -T e2s -C m1s -C e2s -A m1s -G m1s -C e2s -C e2s -A m1s -T e2s -T e2s -G m1s -T e2s -G m1s -T e2s -T e2s -G m1s -A m1s - CH2CH2OH (SEQ ID NO: 15) [In the above formula, the left side represents the 5' end and the right side represents the 3' end, and A, G, C, U and T represent adenosine, guanosine, cytidine, uridine and thymidine, respectively, in which D-ribofuranose has been modified and the carbon atom at the 5' position is linked via a phosphorothioate bond to the structural unit shown on the left side. The e2s attached to each nucleotide or nucleoside represents that the D-ribofuranose is 2'-O,4'-C-ethylene bridged and the 3' position is -OP(=S)(-OH)-O- bonded to the 5' carbon atom of the adjacent nucleotide or nucleoside on the right side, the e2t represents that the D-ribofuranose is 2'-O,4'-C-ethylene bridged and the 3' position is -O- bonded to the hydrogen atom of the 3' end, the m1s represents that the D-ribofuranose is 2'-O-methylated and the 3' position is -OP(=S)(-OH)-O- bonded to the 5' carbon atom of the adjacent nucleotide or nucleoside on the right side, and the m1t represents that the D-ribofuranose is 2'-O-methylated and the 3' position is -O- bonded to the hydrogen atom of the 3' end. Furthermore, one embodiment of the oligonucleotide produced by the method of the present invention is an oligonucleotide having the sequence and structure represented by the following formula and useful for treating glycogen storage disease type 1a (see WO2019 / 172286). (GSD AO01) X 18 -Am1s -A e2s -U m1s -C m1s -C e2s -G m1s -A m1s -T e2s -G m1s -G m1s -C e2s -G m1s -A m1s -A e2s -G m1t -H (SEQ ID NO: 16) GSD AO02) X 18 -A e2s -U m1s -C m1s -C e2s -G m1s -A m1s -T e2s -G m1s -G m1s -C e2s -G m1s -A m1s -A e2s -G m1s -C e2t -H (SEQ ID NO: 17) (GSD AO03) X 18 -U m1s -C m1s -C e2s -G m1s -A m1s -T e2s -G m1s -G m1s -C e2s -G m1s -A m1s -A e2s -G m1s -C e2s -U m1t -H (SEQ ID NO: 18) (GSD AO04) X 18 -A m1s -A m1s -A e2s -U m1s -C m1s -C e2s -G m1s -A m1s -T e2s -G m1s -G m1s -C e2s -Gm1s -A m1s -A e2s -G m1t -H (SEQ ID NO: 19) (GSD AO05) X 18 -A m1s -A e2s -U m1s -C m1s -C e2s -G m1s -A m1s -T e2s -G m1s -G m1s -C e2s -G m1s -A m1s -A e2s -G m1s -C e2t -H (SEQ ID NO: 20) (GSD AO06) X 18 -A e2s -U m1s -C m1s -C e2s -G m1s -A m1s -T e2s -G m1s -G m1s -C e2s -G m1s -A m1s -A e2s -G m1s -C e2s -U m1t -H (SEQ ID NO: 21) (GSD AO07) X 18 -A m1s -A e2s -U m1s -C m1s -C e2s -G m1s -A e2s -U m1s -G m1s -G m1s -C e2s -G m1s -A m1s -A e2s -G m1t -H (SEQ ID NO: 22) (GSD AO08) X 18 -A e2s -U m1s -C m1s -Ce2s -G m1s -A e2s -U m1s -G m1s -G m1s -C e2s -G m1s -A m1s -A e2s -G m1s -C e2t -H (SEQ ID NO: 23) (GSD AO09) X 18 -U m1s -C m1s -C e2s -G m1s -A e2s -U m1s -G m1s -G m1s -C e2s -G m1s -A m1s -A e2s -G m1s -C e2s -U m1t -H (SEQ ID NO: 24) (GSD AO10) X 18 -A m1s -A m1s -A e2s -U m1s -C m1s -C e2s -G m1s -A e2s -U m1s -G m1s -G m1s -C e2s -G m1s -A m1s -A e2s -G m1t -H (SEQ ID NO: 25) (GSD AO11) X 18 -A m1s -A e2s -U m1s -C m1s -C e2s -G m1s -A e2s -U m1s -G m1s -G m1s -C e2s -G m1s -A m1s -A e2s -Gm1s -C e2t -H1 (SEQ ID NO: 26) (GSD AO12) X 18 -A e2s -U m1s -C m1s -C e2s -G m1s -A e2s -U m1s -G m1s -G m1s -C e2s -G m1s -A m1s -A e2s -G m1s -C e2s -U m1t -H (SEQ ID NO: 27) (GSD AO13) X 20 -A m1s -A e2s -U m1s -C m1s -C e2s -G m1s -A e2s -U m1s -G m1s -G m1s -C e2s -G m1s -A m1s -A e2s -G m1t -H (SEQ ID NO: 28) (GSD AO14) X 20 -A e2s -U m1s -C m1s -C e2s -G m1s -A e2s -U m1s -G m1s -G m1s -C e2s -G m1s -A m1s -A e2s -G m1s -C e2t -H (SEQ ID NO: 29) (GSD AO15) X 20 -A m1s -A m1s -A e2s -U m1s -C m1s -C e2s -Gm1s -A e2s -U m1s -G m1s -G m1s -C e2s -G m1s -A m1s -A e2s -G m1t -H (SEQ ID NO: 30) (GSD AO16) X 20 -A m1s -A e2s -U m1s -C m1s -C e2s -G m1s -A e2s -U m1s -G m1s -G m1s -C e2s -G m1s -A m1s -A e2s -G m1s -C e2t -H (SEQ ID NO: 31) [In the above formula, the left side represents the 5' end and the right side represents the 3' end, and A, G, C, U and T represent adenosine, guanosine, cytidine, uridine and thymidine, respectively, in which D-ribofuranose has been modified and the carbon atom at the 5' position is linked via a phosphorothioate bond to the structural unit shown on the left side. e2s attached to each nucleotide or nucleoside represents that D-ribofuranose is 2'-O,4'-C-ethylene bridged and the 3' position is -OP(=S)(-OH)-O- bonded to the 5' carbon atom of the adjacent nucleotide or nucleoside on the right side, e2t represents that D-ribofuranose is 2'-O,4'-C-ethylene bridged and the 3' position is -O- bonded to the hydrogen atom at the 3' end, m1s represents that D-ribofuranose is 2'-O-methylated and the 3' position is -OP(=S)(-OH)-O- bonded to the 5' carbon atom of the adjacent nucleotide or nucleoside on the right side, and m1t represents that D-ribofuranose is 2'-O-methylated and the 3' position is -O- bonded to the hydrogen atom at the 3' end. In the above formula, X 18 and X 20represents a GalNAc unit represented by the following formula, in which the bond bonded to the phosphate group is bonded to the carbon atom at the 5' end of the oligonucleotide to form a phosphodiester bond.

[0469] [ka]

[0470] [ka] EXAMPLES

[0471] The present invention will be described in more detail below with reference to examples, but the scope of the present invention is not limited to these examples. (Example 1) 2,6-anhydro-3-O-benzyl-4-[(benzyloxy)methyl]-5-deoxy-α-L-lyxo-hexofuranose Preparation of compound 2 (Example 1-1) Preparation of a dimethylacetamide solution of (3aR,3bR,6S,6aS,7aR)-6-hydroxy-2,2-dimethyltetrahydro-2H-furo[2',3':4,5]furo[2,3-d][1,3]dioxol-5(3bH)-one (Compound 5)

[0472] [ka]

[0473] To acetone (11.9 kg), commercially available (2R)-2-[(2S,3R,4S)-3,4-dihydroxy-5-oxo-tetrahydrofuran-2-yl]-2-hydroxy-acetaldehyde (compound 4) (1.00 kg, 5.678 mol) and concentrated sulfuric acid (114 g, 1.162 mol) were added in this order, and the mixture was stirred at 40 to 50 ° C for 17 hours. The reaction solution was cooled to 20 to 30 ° C, sodium hydrogen carbonate (0.19 kg, 2.271 mol) was added, and the mixture was stirred for 2 hours. The resulting white solid was removed by filtration, washed with acetone (2.37 kg), and the washing liquid was added to the filtrate. Dimethylacetamide (7.50 kg) and toluene (4.34 kg) were added to the solution thus obtained, and the mixture was concentrated under reduced pressure. The operation of adding toluene (4.34 kg) and concentrating under reduced pressure was repeated twice to obtain a dimethylacetamide solution of compound 5.

[0474] The structure was confirmed by NMR after partial silica gel column purification (hexane / ethyl acetate).

[0475] 1 H NMR (CDCl3): δ = 1.36 (3H, s), 1.53 (3H, s), 2.72 (1H, d, J = 9.5 Hz), 4.50 (1H, dd, J =4.5, 9.5 Hz),4.84 (2H, d, J = 2.5 Hz), 4.95 (1H, dd, J = 2.5,4.5 Hz), 5.99 (1H, d, J = 3.5 Hz). (Example 1-2) Preparation of (3aR,3bR,6S,6aR,7aR)-2,2-dimethyl-5-oxohexahydro-2H-furo[2',3':4,5]furo[2,3-d][1,3]dioxol-6-yl 4-methylbenzene-1-sulfonate (Compound 6)

[0476] [ka]

[0477] To the dimethylacetamide solution of compound 5 obtained in Example 1-1, 1-methylimidazole (0.56 kg, 6.814 mol) and p-toluenesulfonyl chloride (1.24 kg, 6.530 mol) were added in this order, and the mixture was stirred at 20 to 30°C for 3 hours. Water (5 kg) was added to the reaction solution, and the mixture was stirred for 3 hours, then cooled to 0 to 5°C and stirred for another 2 hours. The white solid generated in the reaction solution was filtered and washed with water / dimethylacetamide (1 / 1, 4 kg) and water (8 kg) in this order. The obtained solid was dried under reduced pressure (40°C) to obtain compound 6 (1.66 kg, yield 79.4%).

[0478] 1 H NMR (CDCl3): δ = 1.33 (3H, s), 1.49 (3H, s), 2.45 (3H, s), 4.78 (1H, d, J = 3.5 Hz), 4.83(1H, d, J = 3.0 Hz), 4.97 (1H, dd, J = 3.0, 4.5 Hz), 5.22 (1H, d, J = 4.5 Hz),5.97 (1H, d, J = 3.5 Hz), 7.37 (2H, d, J = 8.0 Hz), 7.89 (2H, m). (Example 1-3) Preparation of a toluene solution of 5-deoxy-1,2-O-(1-methylethylidene)-α-D-xylo-hexofuranose (Compound 7)

[0479] [ka]

[0480] To tetrahydrofuran (22.1 kg, 15.0 v / w) adjusted to 0 to 10 ° C, sodium bis(2-methoxyethoxy)aluminum hydride (4.53 kg, 15.69 mol) and compound 6 (1.66 kg, 4.48 mol) obtained in Example 1-2 were added in this order, and the mixture was stirred for 30 minutes, and then further stirred at 20 to 30 ° C for 1.5 hours. After the reaction, the reaction liquid was cooled to 0 to 10 ° C, acetone (1.05 kg, 0.8 v / w) and a 50% L-sodium potassium tartrate aqueous solution (17.0 kg, 8.0 v / w) were added dropwise, the reaction liquid was heated to 40 to 50 ° C, and the mixture was stirred for 14 hours, and then separated at 40 to 50 ° C. To the obtained organic layer, 20% saline (3.81 kg, 2.0 v / w) was added at 40 to 50 ° C, and a tetrahydrofuran layer was obtained by separation. The same separation operation was repeated twice, and toluene (7.20 kg, 5.0 v / w) was added to the obtained solution, which was then concentrated under reduced pressure until the volume reached 3.3 L. The same reduced pressure concentration operation was repeated three times, and toluene (20.0 kg, 13.6 w / w) was added to the obtained solution to obtain a toluene solution of compound 7.

[0481] The structure was confirmed by NMR after partial silica gel column purification (hexane / ethyl acetate).

[0482] 1 H NMR (CDCl3): δ = 1.32 (3H, s), 1.51 (3H, s), 1.94-2.06 (2H, m), 2.46 (1H, br), 3.26 (1H,br), 3.75 (1H, td, J = 3.5, 9.5 Hz), 3.87-3.91 (1H, m), 4.11 (1H, d, J = 7.5Hz), 4.26 (1H, td, J = 3.0, 6.0 Hz), 4.55 (1H, d, J = 4.0 Hz), 5.91 (1H, d, J =4.0 Hz).

[0483] (Example 1-4) Preparation of a toluene solution of 5-deoxy-1,2-O-(1-methylethylidene)-6-O-(triphenylmethyl)-α-D-xylo-hexofuranose (Compound 8)

[0484] [ka]

[0485] To the toluene solution of compound 7 obtained in Example 1-3, trityl chloride (0.87 kg, 3.14 mol) and 4-methylmorpholine (0.54 kg, 5.38 mol) were added in this order, and the mixture was stirred at 50 to 60°C for 3.5 hours. After the reaction, the reaction solution was cooled to 20 to 30°C, and 8% sodium bicarbonate water (3.42 kg, 2.0 v / w) was added and separated to obtain a toluene layer. 5% saline (10.26 kg, 6.0 v / w) was added to the obtained toluene layer and separated. The obtained toluene layer was subjected to the same separation operation three times to obtain a toluene solution of compound 8.

[0486] The structure was confirmed by NMR after partial silica gel column purification (hexane / ethyl acetate).

[0487] 1 H NMR (CDCl3): δ = 1.31 (3H, s), 1.48 (3H, s), 1.99-2.02 (2H, m), 2.77 (1H, d, J = 4.0 Hz),3.15-3.19 (1H, m), 3.40 (1H, ddd, J = 4.5, 5.0, 10.0 Hz), 4.08 (1H, t, J = 2.5Hz), 4.25 (1H, ddd, J = 2.5, 7.0, 7.5 Hz), 4.53 (1H, d, J = 3.5 Hz), 5.88 (1H,d, J = 3.5 Hz), 7.23-7.26 (3H, m), 7.29-7.32 (6H, m), 7.39-7.41 (6H, m).

[0488] Example 1-5a) 5-Deoxy-1,2-O-(1-methylethylidene)-6-O-(triphenylmethyl)-α-D-erythro- Hexofuranos-3-ulose Preparation of toluene solution of (Compound 9)

[0489] [ka]

[0490] To the toluene solution of compound 8 obtained in Example 1-4, 8% sodium bicarbonate water (5.13 kg, 3.0 v / w), potassium bromide (53.0 g, 0.45 mol) and 9-azanoradamantane-N-oxyl (6.2 g, 44.82 mmol) were added at room temperature, and the temperature of this liquid was adjusted to 0-10°C, after which sodium hypochlorite pentahydrate (2.21 kg, 13.45 mol) was added and stirred for 1.5 hours. After the reaction, the reaction liquid was separated to obtain a toluene layer. The obtained toluene layer was washed twice with 5% sodium sulfite aqueous solution (5.22 kg, 3.0 v / w) and 20% saline (3.80 kg, 2.0 v / w) in this order to obtain a toluene solution of compound 9.

[0491] The structure was confirmed by NMR after partial silica gel column purification (hexane / ethyl acetate).

[0492] 1 H NMR (CDCl3): δ = 1.36 (3H, s), 1.44 (3H, s), 1.96-2.01 (1H, m), 2.21-2.28 (1H, m), 3.06(1H, ddd, J = 3.0, 4.5, 10.0 Hz), 3.30 (1H, ddd, J = 4.0, 4.5, 9.0 Hz), 4.33(1H, dd, J = 1.0, 4.5 Hz), 4.49 (1H, dd, J = 3.5, 5.5 Hz), 5.60 (1H, d, J =4.5), 7.21-7.25 (3H, m), 7.29-7.32 (6H, m), 7.35-7.37 (6H, m). High performance liquid chromatography (HPLC) analysis conditions Column: Xbridge C18 3.5 μm, 4.6x150 mm, column temperature: 40℃ Mobile phase A: 10 mM ammonium acetate aqueous solution, Mobile phase B: acetonitrile Gradient conditions: B(%) 30% (0-3 min), 30→95% (3-20 min), 95%(20-23 min) Flow rate: 1.0 mL / min, detection wavelength: 210 nm Retention times for compound 9: 13.9 min and 17.5 min

[0493] (Example 1-5b) 5-Deoxy-1,2-O-(1-methylethylidene)-6-O-(triphenylmethyl)-α-D-erythro- Hexofuranos-3-ulose Preparation of toluene solution of (Compound 9) To the toluene solution (30 mL) of compound 8 obtained in Example 1-4, 8% sodium bicarbonate water (6.0 mL, 3.0 v / w), potassium bromide (60.0 mg, 0.54 mmol) and 9-azanoradamantane-N-oxyl (3.7 mg, 0.03 mmol) were added at room temperature, and the temperature of this liquid was adjusted to 0 to 10 ° C., and then sodium hypochlorite aqueous solution (6.48 g, 10.80 mmol) was added at 10 ° C. or less, and the mixture was stirred at 0 ° C. for 2 hours. After the reaction was completed, 15% sodium sulfite aqueous solution (12.0 mL, 6.0 v / w) was added and the mixture was stirred at room temperature for 1 hour. The reaction liquid was separated to obtain a toluene layer. The obtained toluene layer was washed with 20% saline (4.0 mL, 2.0 v / w) to obtain a toluene solution of compound 9. The retention time of the obtained compound in HPLC analysis was consistent with the retention time of the compound obtained in (Example 1-5a) in HPLC analysis.

[0494] (Example 1-6) 5-Deoxy-4-(hydroxymethyl)-1,2-O-(1-methylethylidene)-6-O-(triphenylmethyl)-β-L-threo- Hexofuranos-3-ulose Preparation of a mixed toluene solution of (Compound 10) and (3aS,3bR,7aR,8aR)-2,2-dimethyl-7a-[2-(triphenylmethoxy)ethyl]tetrahydro-2H,3bH,5H-[1,3]dioxolo[4,5]furo[3,2-d][1,3]dioxin-3b-ol (Compound 11)

[0495] [ka]

[0496] To the toluene solution of compound 9 obtained in Example 1-5, 37% formaldehyde aqueous solution (1.81 kg, 1.0 v / w) and 1,8-diazabicyclo[5.4.0]undec-7-ene (2.05 kg, 13.45 mol) were added, and the mixture was stirred at 20 to 30°C for 1.5 hours. After the reaction, the reaction solution was separated to obtain a toluene layer. To the obtained toluene layer, 20% citric acid aqueous solution (3.59 kg, 2.0 v / w) was added and separated to obtain a toluene layer. To the obtained toluene layer, 8% sodium bicarbonate water (1.71 kg, 1.0 v / w) was added and separated to obtain a toluene solution of compounds 10 and 11.

[0497] (Example 1-7) Preparation of a toluene solution of 5-deoxy-4-(hydroxymethyl)-1,2-O-(1-methylethylidene)-6-O-(triphenylmethyl)-β-L-lyxo-hexofuranose (Compound 12)

[0498] [ka]

[0499] To the toluene solution of compounds 10 and 11 obtained in Examples 1-6, water (4.98 kg, 3.0 v / w) and sodium borohydride (0.34 kg, 8.96 mol) were added at 20 to 30°C, and the mixture was stirred for 2 hours. After the reaction, the reaction solution was cooled to 0 to 10°C, and 20% citric acid aqueous solution (5.37 kg, 3.0 v / w) was added at the same temperature and separated to obtain a toluene layer. 8% sodium bicarbonate water (3.42 kg, 2.0 v / w) was added to the obtained toluene layer and separated to obtain a toluene solution of compound 12.

[0500] The structure was confirmed by NMR after partial silica gel column purification (hexane / ethyl acetate).

[0501] 1H NMR (CDCl3): δ = 1.33 (3H, s), 1.55 (3H, s), 1.99 (1H, ddd, J = 6.5, 8.0, 14.0 Hz), 2.20(1H, dt, J = 6.0, 14.5 Hz), 2.85 (1H, br), 3.20-3.25 (1H, m), 3.35-3.42 (2H,m), 3.51 (1H, d, J = 12.0 Hz), 4.21 (1H, d, J = 6.0 Hz), 4.63 (1H, dd, J = 4.0,6.0 Hz), 5.73 (1H, d, J = 4.0), 7.21-7.25 (3H, m), 7.27-7.30 (6H, m), 7.41-7.44(6H, m).

[0502] (Example 1-8) Preparation of 3-O-benzyl-4-[(benzyloxy)methyl]-5-deoxy-1,2-O-(1-methylethylidene)-6-O-(triphenylmethyl)-β-L-lyxo-hexofuranose (Compound 13)

[0503] [ka]

[0504] To the toluene solution obtained in Example 1-7, 48% potassium hydroxide aqueous solution (7.27 kg, 3.0 v / w), tetrabutylammonium iodide (0.20 kg, 0.54 mol) and benzyl bromide (1.53 kg, 8.96 mol) were added, and the mixture was stirred at 65 to 75 ° C for 23 hours. After the reaction, water (6.64 kg, 4.0 v / w) and N-acetyl-L-cysteine ​​(0.50 kg, 0.3 w / w) were added to the reaction solution at 45 to 55 ° C, and the mixture was stirred for 2 hours. After the reaction solution was cooled to 20 to 30 ° C, water (4.98 kg, 3.0 v / w) was added and the mixture was separated to obtain a toluene layer. To the obtained toluene layer, 8% sodium bicarbonate water (3.42 kg, 2.0 v / w) was added and the mixture was separated to obtain a toluene layer. The obtained toluene solution was added with 10% saline (3.55 kg, 2.0 v / w) and separated to obtain a toluene layer. The obtained toluene layer was concentrated to 3.3 L, 1-propanol (5.34 kg, 4.0 v / w) was added, insoluble matter was filtered, and the mixture was further concentrated under reduced pressure to 3.0 L. After adding 1-propanol (0.60 kg), the mixture was stirred at 45 to 55 ° C for 0.5 hours to confirm the precipitation of crystals. The mixture was then stirred at 20 to 30 ° C for 12.5 hours and at 0 to 10 ° C for 2 hours, and the precipitated crystals were collected by filtration. The obtained crystals were washed with 1-propanol (4.00 kg, 3.0 v / w) previously cooled to 0 ° C, and dried under reduced pressure (40 ° C) to obtain compound 13 (1.25 kg, 1.90 mol, yield 42.5%).

[0505] 1H NMR (CDCl3): δ = 1.29 (3H, s), 1.51 (3H, s), 1.90-1.96 (1H, m), 2.43-2.49 (1H, m),3.21-3.32 (3H, m), 3.47 (1H, d, J = 10.5 Hz), 4.10 (1H, d, J = 5.5 Hz), 4.30(1H, d, J = 12.0 Hz), 4.42 (1H, d, J = 11.5 Hz), 4.47 (1H, d, J = 12.0 Hz),4.57 (1H, dd, J = 5.5, 4.0 Hz), 4.69 (1H, d, J = 12.0 Hz), 5.69 (1H, d, J = 4.0Hz), 7.15-7.34 (19H, m), 7.39-7.41 (6H, m).

[0506] (Example 1-9) Methyl = 3-O- Benzyl Preparation of a toluene solution of 4-[(benzyloxy)methyl]-5-deoxy-α-L-lyxo-hexofuranoside (compound 14)

[0507] [ka]

[0508] Concentrated sulfuric acid (3.73 g, 0.0380 mmol) was added to a solution of compound 13 (250.0 g, 380.6 mmol) obtained in Example 1-8 in methanol (1250 mL), and the reaction solution was stirred at 40 to 50 ° C for 20 hours. The reaction solution was cooled to 20 to 30 ° C, methoxytriphenylmethane (25 mg) and triethylamine (13.56 g, 134.0 mmol) were added, and the mixture was stirred for 2 hours, and then water (250 mL) was added dropwise over 1.5 hours. After the reaction solution was stirred for 50 minutes, the resulting white solid was filtered off and washed with methanol / water (5 / 1, 500 mL). After adding n-heptane (1250 mL) to the obtained solution and separating the layers, n-heptane (500 mL) was added to the lower layer (aqueous layer) and separating the layers. To the lower layer, 20% saline (1250 mL) and toluene (1250 mL) were added and the mixture was separated to obtain a toluene layer and an aqueous layer. Toluene (1250 mL) was then added to the aqueous layer and the mixture was separated. The obtained toluene layers were combined and concentrated under reduced pressure to obtain a toluene solution of compound 14 (625 mL).

[0509] The structure was confirmed by NMR after partial silica gel column purification (hexane / ethyl acetate).

[0510] 1 H NMR (β-form, CDCl3): δ = 2.00-2.05 (1H, m), 2.10-2.16 (1H, m), 3.29 (3H, s),3.35 (1H, d, J = 9.0 Hz), 3.55 (1H, d, J = 9.0 Hz), 3.76-381 (1H, m), 3.85-3.90(1H, m), 4.08 (2H, dd, J = 5.0, 15.0 Hz), 4.54 (2H, s), 4.63 (2H, s), 4.87 (1H,s), 7.26-7.36 (10H, m).

[0511] (Example 1-10) Preparation of toluene solution of methyl 2,6-anhydro-3-O-benzyl-4-[(benzyloxy)methyl]-5-deoxy-α-L-lyxo-hexofuranoside (Compound 15)

[0512] [ka]

[0513] Triphenylphosphine (89.85g, 342.6mmol) was added to the toluene solution of compound 14 obtained in Example 1-9, and the mixture was cooled to 0-5°C. A solution of diisopropyl azodicarboxylate (69.27g, 342.6mmol) in toluene (180.3mL) was then added dropwise over 20 minutes, and the reaction solution was stirred at 20-30°C for 3 hours. Magnesium chloride (90.60g, 951.6mmol) was added to the reaction solution and stirred for 6 hours, then n-heptane (1500mL) was added and stirred for 23 hours, and the mixture was cooled to 0-5°C and stirred for 2 hours. The resulting white solid was filtered off and washed with toluene / n-heptane (1 / 2, 750mL), and the resulting solution was separated by adding methanol / water (3 / 2, 750mL). Methanol / water (3 / 2, 750 mL) was added to the obtained organic layer, and the layers were separated. The organic layer was then concentrated under reduced pressure to prepare a solution of compound 15 in toluene (625 mL).

[0514] The structure was confirmed by NMR after partial silica gel column purification (hexane / ethyl acetate).

[0515] 1 H NMR (β-form, CDCl3): δ = 1.51 (1H, dd, J = 3.5, 13.0 Hz), 2.19 (1H, ddd, J =7.5, 11.0, 13.5 Hz), 3.41 (3H, s), 3.53 (2H, dd, J = 10.5, 30.5 Hz), 3.86-3.95(3H, m), 4.08 (1H, d, J = 3.0 Hz), 4.53 (1H, d, J = 11.5 Hz), 4.61 (2H, dd, J =12.0, 27.5 Hz), 4.71 (1H, d, J = 6.5 Hz), 5.10 (1H, s), 7.26-7.35 (10H, m).

[0516] (Examples 1-11) Preparation of Compound 2

[0517] [ka]

[0518] Water (625 mL) was added to the toluene solution (625 mL) of compound 15 obtained in Example 1-10, and then the mixture was concentrated under reduced pressure to obtain a water mixture (625 mL) of compound 15. Acetic acid (750 mL) and concentrated hydrochloric acid (125 mL) were added to this mixture in this order, and the mixture was stirred at 20 to 30 ° C for 3 hours, and then compound 2 (25 mg) was added, cooled to 0 to 5 ° C, and stirred for 17 hours. Water (750 mL) was added to this mixture, and the mixture was stirred at 0 to 5 ° C for 17 hours, and the precipitated crystals were collected by filtration and washed with acetic acid / water (1 / 2, 500 mL) and water (500 mL) cooled to 0 to 5 ° C in advance in order to obtain a wet product of crude compound 2. The obtained crude compound 2 was dissolved in toluene (1250 mL) at 35 to 45 ° C, and the resulting aqueous layer was removed by liquid separation. The obtained organic layer was concentrated under reduced pressure to obtain a toluene solution of compound 2 (750 mL), and then n-heptane (375 mL) was added and cooled to 20-30 ° C. To this solution, seed crystals of compound 2 (25 mg) were added and stirred at 20-30 ° C for 2 hours, and then n-heptane (1125 mL) was added and stirred at 0-5 ° C for 19 hours. The precipitated crystals were collected by filtration, washed with toluene / n-heptane (1 / 2, 750 mL), and then washed with n-heptane (750 mL). The obtained crystals were dried under reduced pressure (40 ° C) to obtain compound 2 (76.84 g, 215.6 mmol, yield 56.6%). The seed crystals of compound 2 used in Examples 1-11 were obtained by the following method. Crude compound 2 was obtained as crystals by the same method as the first half of the steps in Examples 1-11. Using the obtained crude compound 2, compound 2 was obtained as crystals by the same method as the second half of the steps in Examples 1-11. Here, the previously obtained crude crystals of compound 2 were used as the seed crystals of compound 2. The crystals of compound 2 obtained by the above method were used as the seed crystals of compound 2 in Examples 1-11.

[0519] 1H NMR (CDCl3): δ = 1.30 (0.39H, d, J = 10.8 Hz), 1.45 (0.59H, dd, J = 10.6, 3.2 Hz),1.97-2.07 (0.39H, m), 2.13-2.22 (0.64H, m), 2.60 (0.34H, d, J = 1.2 Hz), 2.80(0.49H, brs), 3.22 (0.36H, d, J = 7.2 Hz), 3.45-3.59 (1.41H, m), 3.71 (0.37H,d, J = 7.2 Hz), 3.78 (0.37H, brs), 3.82 (0.37H, ddd, J = 9.2, 4.0, 1.6 Hz),3.85-3.95 (1.23H, m), 4.01-4.13 (1.30H, m), 4.16-4.27 (0.83H, m), 4.44-4.75(4.24H, m), 5.36 (0.07H, dd, J = 7.0, 1.6 Hz), 5.57 (0.57H, s), 7.24-7.39 (10H,m), 9.89 (0.36H, s).

[0520] (Examples 1-12) Purification of Compound 2 Crude compound 2 (50 g) obtained in Example 1-11 was dissolved in toluene (300 mL), n-heptane (300 mL) was added, and the mixture was stirred at 20 to 30°C for 3 hours, after which n-heptane (300 mL) was further added and stirred for 16 hours. The precipitated crystals were collected by filtration, washed with toluene / n-heptane (1 / 2, 250 mL), and then washed with n-heptane (250 mL). The obtained crystals were dried under reduced pressure (40°C) to obtain compound 2 (40.06 g, yield 80.1%).

[0521] (Example 2) Preparation of 1-(2,6-anhydro-4-{[bis(4-methoxyphenyl)(phenyl)methoxy]methyl}-3-O-{(2-cyanoethoxy)[di(propan-2-yl)amino]phosphanyl}-5-deoxy-α-L-lyxo-hexofuranosyl)-5-methylpyrimidine-2,4(1H,3H)-dione (Compound 1t) (Example 2-1) Preparation of a methanol solution of 1-{2,6-anhydro-3-O-benzyl-4-[(benzyloxy)methyl]-5-deoxy-α-L-lyxo-hexofuranosyl}-5-methylpyrimidine-2,4(1H,3H)-dione (Compound 3t)

[0522] [ka]

[0523] Thymine (8.85 g, 70.18 mmol) and N,O-bistrimethylsilylacetamide (28.6 g, 140.6 mmol) were added to acetonitrile (50 mL), and the mixture was stirred at 20 to 30 ° C for 1 hour to obtain solution A. In a separate reaction vessel, compound 2 (20.00 g, 56.12 mmol) obtained in Example 1 and trichloroacetonitrile (15.20 g, 105.27 mmol) were added to acetonitrile (100 mL), and then 1,8-diazabicyclo[5.4.0]undec-7-ene (1.07 g, 7.02 mmol) was added, and the mixture was stirred at 20 to 30 ° C for 1 hour to obtain solution B. Solution A was added to solution B, and then iodotrimethylsilane (17.55 g, 87.73 mmol) was added and the mixture was stirred at 20 to 30 ° C for 2 hours. A solution of sodium sulfite (17.68 g, 175.45 mmol) dissolved in 5% aqueous sodium hydrogen carbonate (160 mL) was added dropwise to the reaction solution, and then toluene (200 mL) and methanol (80 mL) were added, stirred at 35 to 45 ° C., and then separated. 40% aqueous methanol (200 mL) was added to the obtained organic layer, stirred at 35 to 45 ° C., and then separated. After another separation operation with 40% aqueous methanol (200 mL), the obtained organic layer was concentrated under reduced pressure to give a methanol solution (50 mL), and then methanol (200 mL) was added and concentrated under reduced pressure again to give a methanol solution (50 mL) of compound 3t.

[0524] The NMR spectrum of the obtained compound was consistent with the NMR spectrum of the compound described in Example 6 of WO00 / 47599.

[0525] (Example 2-2) Preparation of 1-[2,6-anhydro-5-deoxy-4-(hydroxymethyl)-α-L-lyxo-hexofuranosyl]-5-methylpyrimidine-2,4(1H,3H)-dione (Compound 16)

[0526] [ka]

[0527] Methanol (50mL) and palladium hydroxide (1.0g) were further added to the methanol solution (50mL) of compound 3t obtained in Example 2-1, and the mixture was stirred for 2 hours at 55-65°C under a hydrogen atmosphere, and the catalyst was filtered at the same temperature. The obtained filtrate was concentrated under reduced pressure to obtain a methanol solution (50mL), and then acetone (200mL) was added and the mixture was concentrated under reduced pressure to 50mL. Further, acetone (100mL) was added, and the mixture was cooled to 0-5°C and stirred for 2 hours. The resulting crystals were filtered and washed with acetone (50mL). The obtained crystals were dried under reduced pressure (40 °C) to obtain compound 16 (9.95 g, yield 62.4% (from compound 2)).

[0528] The NMR spectrum of the obtained compound was identical to that of the compound described in Example 7 of WO00 / 47599.

[0529] (Example 2-3) Preparation of 1-(2,6-anhydro-4-{[bis(4-methoxyphenyl)(phenyl)methoxy]methyl}-5-deoxy-α-L-lyxo-hexofuranosyl)-5-methylpyrimidine-2,4(1H,3H)-dione (Compound 17)

[0530] [ka]

[0531] Compound 16 (9.50 g, 33.42 mmol) obtained in Example 2-2 and pyridine (10.57 g, 133.68 mmol) were added to tetrahydrofuran (47.5 mL), followed by 4,4'-dimethoxytrityl chloride (13.59 g, 40.10 mmol), and the reaction solution was stirred at 20 to 30 ° C. for 2 hours. Methanol (1.62 mL, 40.10 mmol) was added to the reaction solution, and 15% sodium carbonate aqueous solution (28.5 mL), ethyl acetate (133 mL) and water (67 mL) were added in this order to separate and wash the organic layer obtained, which was then washed twice with 20% citric acid aqueous solution (47.5 mL), and then washed with 15% sodium carbonate aqueous solution (47.5 mL) and water (47.5 mL) in that order, and then concentrated under reduced pressure to 47.5 mL. Ethyl acetate (100 mL) was added to the concentrate obtained, and the mixture was concentrated under reduced pressure to 100 mL. The concentrate obtained was stirred at 55-65°C for 16 hours, further stirred at 20-30°C for 5 hours, n-heptane (50 mL) was added, and the mixture was further stirred for 16 hours, after which the precipitated solid was collected by filtration. The solid obtained was washed with ethyl acetate / n-heptane (2 / 1, 47.5 mL), and then dried under reduced pressure (40°C) to obtain compound 17 (17.35 g, 88.5%).

[0532] The NMR spectrum of the obtained compound was consistent with the NMR spectrum of the compound described in Example 8 of WO00 / 47599.

[0533] (Example 2-4) Preparation of Compound 1t

[0534] [ka]

[0535] 2-Cyanoethyl N,N,N',N'-tetraisopropylphosphorodiamidite (2.96g, 9.82mmol) and trifluoroacetic acid pyridine salt (1.74g, 9.01mmol) were added to a solution of compound 17 (5.00g, 8.17mmol) obtained in Example 2-3 in ethyl acetate (40mL), and the mixture was stirred at 20-30°C for 24 hours. 20% saline (17.5mL) was added to the obtained solution and the mixture was separated, and then 5% aqueous sodium hydrogen carbonate solution (12.5mL) and 20% aqueous sodium chloride (10mL) were added to the obtained organic layer and the mixture was separated. The obtained organic layer was cooled to 0-5°C, and 10% aqueous potassium dihydrogen phosphate solution (20mL) was added and the mixture was separated. The organic layer was separated by adding 5% aqueous sodium bicarbonate solution (12.5mL) and 20% saline (10mL), and the organic layer was separated by adding 20% ​​saline (17.5mL). The organic layer was concentrated under reduced pressure and dissolved in ethyl acetate (15mL). Ethyl acetate (25mL) and basic alumina (10.00g) were added to this solution and stirred at 20-30°C for 2 hours. Alumina was removed from the reaction mixture by filtration and washed with ethyl acetate (25 mL). The filtrate and the washings were combined and the resulting solution was concentrated under reduced pressure. The concentrate was dissolved in ethyl acetate (15mL). The ethyl acetate solution was added dropwise to a mixed solution of n-heptane (45mL) and diisopropyl ether (80mL) at 20-30°C over 35 minutes, and then n-heptane (90mL) was added and stirred at the same temperature for 30 minutes. The precipitated solid was collected by filtration, washed with n-heptane (25 mL), and dried under reduced pressure (40° C., 12 hours, then 50° C., 8 hours) to obtain the title compound 1t (5.18 g, 6.58 mmol, 80.6%).

[0536] The NMR spectrum of the obtained compound was consistent with the NMR spectrum of the compound described in Example 9 of WO00 / 47599.

[0537] (Example 3) Preparation of 1-(2,6-anhydro-4-{[bis(4-methoxyphenyl)(phenyl)methoxy]methyl}-3-O-{(2-cyanoethoxy)[di(propan-2-yl)amino]phosphanyl}-5-deoxy-α-L-lyxo-hexofuranosyl)-4-benzamido-5-methylpyrimidin-2(1H)-one (Compound 1c) (Example 3-1) Preparation of 1-{2,6-anhydro-3-O-benzyl-4-[(benzyloxy)methyl]-5-deoxy-α-L-lyxo-hexofuranosyl}-4-benzamido-5-methylpyrimidin-2(1H)-one (Compound 3c)

[0538] [ka]

[0539] Acetonitrile (25 mL) with N 4 -benzoyl-5-methylcytosine (8.04g, 35.08mmol), N,O-bistrimethylsilylacetamide (14.27g, 70.16mmol) were added, and the mixture was stirred at 20-30°C for 1 hour to obtain solution A. In a separate reaction vessel, compound 2 (10.00g, 28.06mmol) obtained in Example 1 and trichloroacetonitrile (6.08g, 42.09mmol) were added to acetonitrile (50mL), and then 1,8-diazabicyclo[5.4.0]undec-7-ene (0.41g, 2.81mmol) was added, and the mixture was stirred at 20-30°C for 1 hour to obtain solution B. After adding solution A to solution B, iodotrimethylsilane (7.02g, 35.08mmol) was added and the mixture was stirred at 5-15°C for 16 hours. A solution of sodium sulfite (8.84 g, 87.73 mmol) dissolved in 5% aqueous sodium bicarbonate (80 mL) was added dropwise to the reaction solution, and then toluene (100 mL) was added. After separation, a small amount of solid was removed by filtration. The obtained organic layer was washed twice with 50% aqueous methanol (50 mL). The obtained organic layer was concentrated under reduced pressure to obtain a methanol solution (50 mL), and the mixture was stirred at 20 to 30°C for 16 hours. The precipitated solid was filtered and washed with methanol (50 mL). The obtained solid was dried under reduced pressure (40°C) to obtain compound 3c (9.35 g, 58.7%).

[0540] The NMR spectrum of the obtained compound was consistent with that of the compound described in Example 19 of WO00 / 47599.

[0541] (Example 3-2) Preparation of 1-[2,6-anhydro-5-deoxy-4-(hydroxymethyl)-α-L-lyxo-hexofuranosyl]-4-benzamido-5-methylpyrimidin-2(1H)-one (Compound 18)

[0542] [ka]

[0543] Compound 3c (5.00 g, 8.81 mmol) obtained in Example 3-1 was added to methylene chloride (20 mL), and 1M-boron trichloride / methylene chloride solution (40.0 mL, 39.65 mmol) was added dropwise at -20 to -10 ° C., and the mixture was stirred for 16 hours. 50% Rochelle salt aqueous solution (20.0 mL) was added dropwise to the reaction solution, and ethyl acetate (20 mL) and water (20 mL) were added. After stirring at 20 to 30 ° C. for 16 hours, the precipitated solid was filtered and washed with water (20 mL) and ethyl acetate (20 mL). The obtained solid was dried under reduced pressure (40 ° C.) to obtain compound 18 (1.59 g, 46.6%).

[0544] The NMR spectrum of the obtained compound was consistent with that of the compound described in Example 11 of WO00 / 47599.

[0545] (Example 3-3) Preparation of 1-(2,6-anhydro-4-{[bis(4-methoxyphenyl)(phenyl)methoxy]methyl}-5-deoxy-α-L-lyxo-hexofuranosyl)-4-benzamido-5-methylpyrimidin-2(1H)-one (Compound 19)

[0546] [ka]

[0547] Compound 18 (1.00 g, 2.581 mmol) obtained in Example 3-2 was added to ethyl acetate (40 mL), concentrated under reduced pressure to 10 mL, pyridine (0.815 g, 10.324 mmol) was added, and 4,4'-dimethoxytrityl chloride (1.05 g, 3.098 mmol) was added, and the mixture was stirred at 20 to 30 ° C for 16 hours. Methanol (1.0 mL) was added to the reaction solution, followed by water (10 mL) and methanol (10 mL), and the mixture was stirred at 20 to 30 ° C for 16 hours. The precipitated solid was filtered and washed with water (5 mL) and ethyl acetate (5 mL). The obtained solid was dried under reduced pressure (40 ° C) to obtain hydrate crystals of compound 19 (1.53 g, 86.0%).

[0548] The NMR spectrum of the obtained compound was consistent with that of the compound described in Example 21 of WO00 / 47599.

[0549] (Example 3-4) Preparation of Compound 1c

[0550] [ka]

[0551] Dichloromethane (30 mL) was added to the hydrate of compound 19 obtained in Example 3-3 (3.00 g, 4.24 mmol), and the mixture was concentrated under reduced pressure to give a dichloromethane solution (9 mL). The procedure of adding dichloromethane (30 mL) to the obtained solution, concentrating under reduced pressure to give a dichloromethane solution (9 mL) was repeated twice. Dichloromethane (15 mL), N,N,N',N'-tetraisopropyl phosphorodiamidite (1.54 g, 5.11 mmol), and trifluoroacetic acid pyridine salt (0.902 g, 4.67 mmol) were added to the obtained solution, and the mixture was stirred at 20-30°C for 7 hours. The obtained solution was washed twice with 5% aqueous sodium hydrogen carbonate solution (10.5 mL), then washed twice with 5% saline (12 mL), and concentrated under reduced pressure to give a dichloromethane solution (9 mL). To the obtained solution, dichloromethane (15 mL) and basic alumina (6.00 g) were added, and the mixture was stirred at 20 to 30°C for 30 minutes. The alumina was removed by filtration, and the mixture was washed with dichloromethane (15 mL), and the obtained solution was concentrated under reduced pressure to obtain a dichloromethane solution (9 mL) of compound 1c. The obtained solution was added dropwise to a mixed solution of n-heptane (66 mL) and diisopropyl ether (15.6 mL) at 20 to 30°C over 35 minutes. After stirring at the same temperature for 2 hours, the precipitated solid was filtered and washed with n-heptane (15 mL). The obtained solid was dried under reduced pressure (40°C) to obtain compound 1c (3.33 g, 3.74 mmol, 88.3%).

[0552] The NMR spectrum of the obtained compound was consistent with the NMR spectrum of the compound described in Example 6 of WO00 / 47599.

[0553] (Example 4) Preparation of 9-(2,6-anhydro-4-{[bis(4-methoxyphenyl)(phenyl)methoxy]methyl}-3-O-{(2-cyanoethoxy)[di(propan-2-yl)amino]phosphanyl}-5-deoxy-α-L-lyxo-hexofuranosyl)-N-benzoyl-9H-purin-6-amine (Compound 1a) (Example 4-1-1) Preparation of 9-{2,6-anhydro-3-O-benzyl-4-[(benzyloxy)methyl]-5-deoxy-α-L-lyxo-hexofuranosyl}-N-benzoyl-9H-purin-6-amine (Compound 3a)

[0554] [ka]

[0555] Dichlorodimethylsilane (5.44g, 42.09mmol) was added to trifluoromethanesulfonic acid (7.44mL, 84.18mmol), and the mixture was stirred at 20-30°C for 0.5 hours. Then, acetonitrile (80mL) and N-benzoyladenine (10.06g, 42.09mmol) were added, and the mixture was stirred at 20-30°C for 3 hours to obtain solution A. In a separate reaction vessel, compound 2 (10.00g, 28.06mmol) obtained in Example 1 and trichloroacetonitrile (4.2mL, 42.09mmol) were added to acetonitrile (40mL), and 1,8-diazabicyclo[5.4.0]undec-7-ene (0.22g, 1.40mmol) was added, and the mixture was stirred at 20-30°C for 1 hour to obtain solution B. Solution B was added to solution A, and the mixture was stirred at 45-55°C for 2 hours. After adjusting the temperature to 0-10°C, toluene (100mL) and 20% potassium bicarbonate aqueous solution (70mL) were added to the reaction solution, and the solution was separated to obtain a toluene layer. A 20% citric acid aqueous solution (50mL) was added to the obtained toluene layer, and the solution was separated to obtain a toluene layer. The same separation operation was repeated twice, and water (30mL) and 20% potassium bicarbonate aqueous solution (25mL) were added to the obtained toluene layer, and the solution was separated to obtain a toluene layer. A 50% methanol aqueous solution (100mL) was added to the obtained toluene layer, and the solution was separated to obtain a toluene layer. The obtained toluene layer was concentrated under reduced pressure to obtain a toluene solution (80mL). Isobutyl alcohol (100mL) was added to the obtained solution, and the solution was concentrated under reduced pressure to obtain a toluene-isobutyl alcohol solution (100mL). Isobutyl alcohol (50mL) was added to the obtained solution, and the solution was concentrated under reduced pressure to obtain a toluene-isobutyl alcohol solution (50mL). Isobutyl alcohol (50 mL) was added to the obtained solution, and the mixture was stirred at 20 to 30° C. for 2 hours to confirm the precipitation of crystals. After adjusting the temperature to 0-10° C. over 0.5 hours and stirring for 1 hour, the precipitated crystals were filtered, washed with isobutyl alcohol (50 mL) pre-cooled to 0° C., and dried under reduced pressure (40° C.) to obtain compound 3a (7.82 g, 13.54 mmol, yield 48.3%).

[0556] The NMR spectrum of the obtained compound was consistent with the NMR spectrum of the compound described in Example 6 of WO00 / 47599.

[0557] (Example 4-1-2) Preparation of 1-O-acetyl-2,6-anhydro-3-O-benzyl-4-[(benzyloxy)methyl]-5-deoxy-α-L-glycero-hexofuranose (Compound 22)

[0558] [ka]

[0559] Toluene (1497 mL), compound 2 (149.73 g, 375.78 mmol) obtained in Example 1, acetic anhydride (64.33 g, 563.67 mmol), pyridine (49.85 g, 563.67 mmol), and 4-dimethylaminopyridine (2.57 g, 18.79 mmol) were added, and the mixture was stirred at 20 to 30 ° C for 2 hours. Water (749 mL) was added to the reaction solution, and the mixture was separated to obtain a toluene layer. A 20% aqueous citric acid solution (749 mL) was added to the obtained toluene layer, and the mixture was separated to obtain a toluene layer. The same separation operation was repeated again, and an 8% aqueous potassium hydrogen carbonate solution (449 mL) was added to the obtained toluene layer, and the mixture was separated to obtain a toluene layer. A 5% saline solution (449 mL) was added to the obtained toluene layer, and the mixture was separated to obtain a toluene layer. The obtained toluene layer was concentrated under reduced pressure to obtain a toluene solution (449 mL). Acetonitrile (1497 mL) was added to the obtained solution, and the mixture was concentrated under reduced pressure to obtain a toluene-acetonitrile solution (449 mL). Acetonitrile (1497 mL) was added to the obtained solution, and the mixture was concentrated under reduced pressure to obtain a toluene-acetonitrile solution (449 mL) of compound 22.

[0560] (Example 4-1-3a) Preparation of 9-{2,6-anhydro-3-O-benzyl-4-[(benzyloxy)methyl]-5-deoxy-α-L-lyxo-hexofuranosyl}-N-benzoyl-9H-purin-6-amine (Compound 3a)

[0561] [ka]

[0562] Acetonitrile (51 mL), N-benzoyladenine (5.09 g, 21.30 mmol), and trimethylsilyl trifluoromethanesulfonate (9.78 g, 44.01 mmol) were added, and the mixture was stirred at 20-30 °C for 40 minutes. A toluene-acetonitrile solution (11.65 g, 14.20 mmol) containing compound 22 obtained in Example 4-1-2 was added, and the mixture was stirred at 20-30 °C for 16 hours. Subsequently, trifluoroacetic acid (2.43 g, 21.30 mmol) was added, and the mixture was stirred at 20-30 °C for 1 hour, and then the mixture was heated to 45-55 °C and stirred for 3 hours. After adjusting the temperature to 20-30 °C, toluene (51 mL), 25% sodium hydroxide (10 mL), and 8% sodium bicarbonate aqueous solution (10 mL) were added to the reaction solution, and the mixture was stirred at 20-30 °C for 1 hour. Subsequently, a 10% aqueous trifluoroacetic acid solution (25 mL) was added, and the mixture was separated to obtain a toluene layer. The same separation operation was repeated three times to obtain a toluene layer, to which an 8% aqueous sodium bicarbonate solution (51 mL) was added, and the mixture was separated to obtain a toluene layer. The toluene layer obtained by repeating the same separation operation once was concentrated under reduced pressure to obtain a toluene solution (15 mL). Isobutyl alcohol (40 mL) was added to the obtained solution, and the solution was concentrated under reduced pressure to obtain a toluene-isobutyl alcohol solution (40 mL). Isobutyl alcohol (15 mL) was added to the obtained solution, and crystals of compound 3a (0.1 wt%) were inoculated, and the mixture was stirred at 35 to 45 ° C for 30 minutes to confirm the precipitation of crystals. The mixture was then concentrated under reduced pressure to give a toluene-isobutyl alcohol solution (40 mL), after which isobutyl alcohol (5 mL) was added, and the mixture was heated to 0-10°C over 2 hours and stirred for 17.5 hours. The precipitated crystals were filtered, washed with isobutyl alcohol (25 mL) pre-cooled to 0°C, and dried under reduced pressure (40°C) to obtain compound 3a (6.37 g, 11.03 mmol, 77.7% yield).

[0563] (Example 4-1-3b) Preparation of 9-{2,6-anhydro-3-O-benzyl-4-[(benzyloxy)methyl]-5-deoxy-α-L-lyxo-hexofuranosyl}-N-benzoyl-9H-purin-6-amine (Compound 3a) To acetonitrile (30 mL), N-benzoyladenine (3.02 g, 12.63 mmol) and trimethylsilyl trifluoromethanesulfonate (5.61 g, 25.25 mmol) were added, and the reaction mixture was stirred at 20-30 °C for 30 minutes. To the resulting reaction mixture, an acetonitrile solution (6 mL) containing compound 22 obtained in Example 4-1-2 and acetonitrile (3 mL) were added, and the reaction mixture was stirred at 20-30 °C for 2 hours. Subsequently, trifluoroacetic acid (1.44 g, 12.63 mmol) was added, and the reaction mixture was stirred at 35-45 °C for 6 hours. After the resulting reaction mixture was cooled to 0 °C, the reaction solution was adjusted to pH 6.5-7.5 using 25% sodium hydroxide (3 mL). Then, water (9 mL) was added to the reaction mixture at 20-30 °C, and crystals of compound 3a (0.1 wt%) were seeded. The reaction mixture was stirred at 20-30 °C for 19 hours, after which water (27 mL) was added and the mixture was stirred at 20-30 °C for 4 hours. The precipitated crystals were filtered, washed with 50% acetonitrile water (15 mL), and dried under reduced pressure (40 °C) to obtain compound 3a (3.59 g, 6.21 mmol, yield 73.8%).

[0564] The NMR spectrum of the obtained compound was consistent with the NMR spectrum of the compound described in Example 6 of WO00 / 47599.

[0565] (Example 4-2) Preparation of 9-[2,6-anhydro-5-deoxy-4-(hydroxymethyl)-α-L-lyxo-hexofuranosyl]-N-benzoyl-9H-purin-6-amine (Compound 20)

[0566] [ka]

[0567] Compound 3a (2.00 g, 3.46 mmol) obtained in Example 4-1-3 was added to 1M boron trichloride / methylene chloride solution (50.0 mL, 51.94 mmol) at -20 to -10°C, and the mixture was stirred for 0.5 hours. After confirming the completion of the reaction, the mixture was concentrated under reduced pressure to obtain a methylene chloride solution (20 mL).

[0568] A 50% aqueous potassium hydroxide solution (6.0 mL) was added dropwise to the obtained solution, n-heptane (6 mL) was added, and the mixture was stirred at 20 to 30°C for 15 minutes. The precipitated solid was filtered and washed with methylene chloride / n-heptane (5 / 1, 4 mL). Acetonitrile (30 mL) and water (4 mL) were added to the obtained solid. After stirring for 0.5 hours, the solid was filtered and washed with acetonitrile (4 mL). Methanol (30 mL) was added to the obtained solid, and the solid was filtered off and washed with methanol (10 mL) to obtain a methanol solution. The obtained solution was concentrated under reduced pressure to prepare a methanol solution (10 mL). Acetonitrile (20 mL) was added to the obtained solution, and the mixture was concentrated under reduced pressure to obtain an acetonitrile-methanol mixed solution (10 mL). The same vacuum concentration operation was repeated twice, and acetonitrile (10 mL) was added to the resulting acetonitrile-methanol mixed solution (10 mL), and the mixture was stirred for 16 hours. The solid was filtered and washed with acetonitrile (6 mL). The solid was dried under reduced pressure (40° C.) to obtain compound 20 (0.61 g, 44.3%).

[0569] The NMR spectrum of the obtained compound was consistent with that of the compound described in Example 11 of WO00 / 47599.

[0570] (Example 4-3) Preparation of 9-(2,6-anhydro-4-{[bis(4-methoxyphenyl)(phenyl)methoxy]methyl}-5-deoxy-α-L-lyxo-hexofuranosyl)-N-benzoyl-9H-purin-6-amine (Compound 21)

[0571] [ka]

[0572] Compound 20 (0.50 g, 1.26 mmol) obtained in Example 4-2 and 4,4'-dimethoxytrityl chloride (0.64 g, 1.89 mmol) were added to tetrahydrofuran (7.5 mL), pyridine (0.60 g, 7.55 mmol) was added, and the mixture was stirred at 50 ° C. for 2 hours. Toluene (5.0 mL) and water (2.5 mL) were added to the reaction solution, and the mixture was separated to obtain a toluene layer. The obtained toluene layer was separated three times with a 20% aqueous citric acid solution (2.5 mL), and a 20% aqueous potassium carbonate solution (1.0 mL) was added to separate the toluene layer. The obtained toluene layer was washed by adding water (2.5 mL) and then concentrated under reduced pressure to obtain a toluene solution (2.5 mL). Toluene (5.0 mL) was added to the obtained solution, and the mixture was concentrated under reduced pressure to obtain a toluene solution (2.5 mL). The same operation was repeated twice to obtain a toluene solution (2.5 mL). Toluene (2.5 mL) and water (50 μL) were added to the obtained solution, and the mixture was stirred at 20 to 30° C. for 3 hours. The precipitated solid was filtered and washed with toluene (2.5 mL). The obtained solid was dried under reduced pressure (40° C.) to obtain crude compound 21 (0.51 g, 58.0%).

[0573] Crude compound 21 (0.36 g, 0.51 mmol) and water (1.8 mL) were added to 2-propanol (1.8 mL) and stirred at 20-30°C for 0.5 hours. Water (1.8 mL) was further added and stirred for 15 minutes, after which the solid was filtered and washed with 2-propanol-water (1 / 3, 1.8 mL). The obtained solid was dried under reduced pressure (40°C) to obtain a hydrate of compound 21 (0.31 g, 86.1%).

[0574] The NMR spectrum of the obtained compound was consistent with that of the compound described in Example 13 of WO00 / 47599.

[0575] (Example 4-4) Preparation of Compound 1a

[0576] [ka]

[0577] Dichloromethane (1.5mL) was added to the hydrate of compound 21 obtained in Example 4-3 (150mg, 0.209mmol), and the mixture was concentrated under reduced pressure to dryness. Dichloromethane (1.5mL) was added to the obtained foam, and the mixture was concentrated under reduced pressure to dryness. Dichloromethane (1.2mL), N,N,N',N'-tetraisopropylphosphorodiamidite (77.9mg, 0.258mmol), and trifluoroacetic acid pyridine salt (44.4mg, 0.230mmol) were then added, and the mixture was stirred at 20-30°C for 4 hours. The obtained solution was washed twice with 5% aqueous sodium bicarbonate solution (0.525mL), then washed twice with 5% saline (0.600mL), and the mixture was concentrated under reduced pressure to dryness. Ethyl acetate (1.2mL) and neutral silica gel (300mg) were added to the obtained foam, and the mixture was stirred at 20-30°C for 1 hour. The silica gel was filtered off and washed with ethyl acetate (6 mL), and the resulting solution was concentrated under reduced pressure to dryness. The resulting foamy ethyl acetate (0.600 mL) solution was added dropwise to a mixed solution of n-heptane (1.35 mL) and diisopropyl ether (2.4 mL) at 20-30°C over 20 minutes. n-heptane (3.15 mL) was added and stirred at the same temperature for 1 hour, after which the precipitated solid was filtered and washed with n-heptane (6 mL). The resulting solid was dried under reduced pressure (40°C) to obtain compound 1a (132.5 mg, 0.147 mmol, 70.4%).

[0578] The NMR spectrum of the obtained compound was consistent with that of the compound described in Example 14 of WO00 / 47599.

[0579] Example 5: Preparation of Compound 20 (Example 5-1) Purification of Compound 22 The solution obtained in Example 4-1-2 was concentrated to dryness to obtain a crude mixture of compound 22 (6.57 g). The crude mixture was purified by silica gel column chromatography (hexane:ethyl acetate=94:6 to 50:50) to obtain the β-form of compound 22 (4.68 g). 1H NMR (β-form, CDCl3): δ = 1.43 (1H, dd, J = 3.5, 13.5Hz), 2.01 (3H, s), 2.17-2.24 (1H, m), 3.57 (2H, dd, J = 11.5, 22.0 Hz),3.94-3.97 (2H, m), 4.07 (1H, d, J = 3.0 Hz), 4.14 (1H, d, J = 3.0 Hz), 4.54(1H, d, J = 12.5 Hz), 4.61 (2H, dd, J = 12.5, 18.0 Hz), 4.74 (1H, d, J = 11.5Hz), 6.33 (1H, s), 7.26-7.37 (10H, m).

[0580] (Example 5-2) Preparation of 9-{2,6-anhydro-3-O-benzyl-4-[(benzyloxy)methyl]-5-deoxy-α-L-lyxo-hexofuranosyl}-2,6-dichloro-9H-purine (Compound 23)

[0581] [ka]

[0582] To a toluene solution (0.6 mL) of compound 22 (60.0 mg, 150.58 μmol) obtained in Example 5-1-2, N,O-bistrimethylsilylacetamide (40.5 μL, 165.64 μmol) and 2,6-dichloropurine (31.3 mg, 165.64 μmol) were added, and the mixture was stirred at 75° C. for 1 hour, after which iodotrimethylsilane (22.5 μL, 165.64 μmol) was added and stirred for 3 hours. After confirming the completion of the reaction, the mixture was cooled to room temperature, and ethyl acetate (18.0 mL) and 5% sodium bicarbonate water (3.0 mL) were added and separated. The obtained organic layer was concentrated to dryness, and the residue was purified by silica gel chromatography (hexane: ethyl acetate = 2: 1) and concentrated under reduced pressure to obtain compound 23 as a colorless oily compound (52.0 mg, 65.4%). 1H-NMR (CDCl3): δ = 8.79 (1H, s), 7.23-7.39 (10H, m),6.43 (1H, s), 4.57-4.65 (2H, m), 4.42-4.45 (1H, m), 4.15 (1H, d, J = 3.0 Hz),4.06-4.09 (2H, m), 3.71 (1H, d, J = 10.5 Hz), 3.56 (1H, d, J = 10.5 Hz),2.27-2.36 (2H, m), 1.41-1.44 (2H, m)

[0583] (Example 5-3) Preparation of 9-{2,6-anhydro-3-O-benzyl-4-[(benzyloxy)methyl]-5-deoxy-α-L-lyxo-hexofuranosyl}-2-chloro-9H-purin-6-amine (Compound 24)

[0584] [ka]

[0585] A 28% aqueous ammonia solution (0.5 mL) was added to a tetrahydrofuran solution (0.75 mL) of compound 23 (52.0 mg, 98.60 μmol) obtained in Example 5-2, and the mixture was stirred at 50° C. for 7 hours. After confirming the completion of the reaction, the mixture was cooled to room temperature, and ethyl acetate (2.0 mL) was added to separate the mixture. The obtained organic layer was washed with 20% saline (1.0 mL), and then concentrated to dryness. The residue was purified by silica gel chromatography (hexane:ethyl acetate=4:6) and concentrated under reduced pressure to obtain compound 24 as white crystals (44.7 mg, 89.2%). 1H-NMR (CDCl3): δ = 8.36 (1H, s), 7.27-7.38 (10H, m),6.39 (1H, s), 5.69 (2H, brs), 4.56-4.68 (2H, m), 4.44-4.48 (1H, m), 4.20 (1H,d, J = 2.5 Hz), 4.05-4.14 (2H, m), 3.70 (1H, d, J = 10.8 Hz), 3.57 (1H, d, J =10.8 Hz), 2.28-2.36 (1H, m), 1.53 (1H, m), 1.42-1.44 (1H, m), 1.25-1.27 (1H, m)

[0586] (Example 5-4) Preparation of 9-{2,6-anhydro-3-O-benzyl-4-[(benzyloxy)methyl]-5-deoxy-α-L-lyxo-hexofuranosyl}-9H-purin-6-amine (Compound 25)

[0587] [ka]

[0588] 20% palladium hydroxide carbon (20.0 mg) and 1N sodium hydroxide aqueous solution (103.2 μL, 103.16 μmol) were added to an ethanol solution (2.0 mL) of compound 24 (26.2 mg, 51.58 μmol) obtained in Example 5-3, and the mixture was stirred at 50° C. for 5 hours under a hydrogen pressure of 3.5 bar. After the reaction, the mixture was cooled to room temperature, and 1N hydrochloric acid aqueous solution (51.6 μL, 51.58 μmol) was added and filtered. Ethanol (10 mL) was added to the filtrate, and the mixture was concentrated to dryness to obtain compound 25 as white crystals (9.7 mg, 64.1%). 1H-NMR (CD3OD): δ = 8.72 (1H, s), 8.35 (1H, s), 6.48(1H, s), 4.37 (1H, d, J = 3.5 Hz), 4.32 (1H, d, J = 3.0 Hz), 4.01-4.13 (2H, m),3.78 (1H, d, J = 12.5 Hz), 3.72 (1H, d, J = 12.0 Hz), 2.17-2.24 (1H, m), 1.44(1H, dd, J = 13.0 Hz, 4.0 Hz)

[0589] (Example 5-5) Preparation of Compound 20

[0590] [ka]

[0591] Add chlorotrimethylsilane (5 equivalents) to the pyridine solution (30 v / w) of compound 25 obtained in Example 5-4, stir for 30 minutes, add benzoyl chloride (5 equivalents), and react at room temperature for 2 hours. After the reaction, cool to 0°C and add aqueous ammonia to make a 2M ammonia solution. After 30 minutes, concentrate under reduced pressure, add acetonitrile (30 v / w), concentrate to (3 v / w), and filter the precipitated crystals to obtain compound 20.

[0592] (Example 6) Preparation of 9-(2,6-anhydro-4-{[bis(4-methoxyphenyl)(phenyl)methoxy]methyl}-3-O-{(2-cyanoethoxy)[di(propan-2-yl)amino]phosphanyl}-5-deoxy-α-L-lyxo-hexofuranosyl)-2-(2-methylpropanamido)-1,9-dihydro-6H-purin-6-one (Compound 1g) (Example 6-1) Preparation of 9-{2,6-anhydro-3-O-benzyl-4-[(benzyloxy)methyl]-5-deoxy-α-L-lyxo-hexofuranosyl}-6-(benzyloxy)-2-chloro-9H-purine (Compound 26)

[0593] [ka]

[0594] Benzyl alcohol (4.63μL, 66.84μmol) and sodium hydride (2.5mg, 57.93μmol) were added to tetrahydrofuran (0.5mL) and stirred at 0℃ for 30 minutes, and then compound 23 (23.5mg, 44.56μmol) obtained in Example 5-2 was added and stirred for 3 hours. After confirming the completion of the reaction, acetic acid (3.6μL, 62.38μmol), 20% saline (0.5mL), and ethyl acetate (2.0mL) were added and separated. The obtained organic layer was concentrated to dryness, and the residue was purified by silica gel chromatography (hexane: ethyl acetate = 2: 1) and concentrated under reduced pressure to obtain compound 26 as a colorless oily compound (20.9mg, 78.3%). 1 H-NMR (CD3Cl): δ = 8.50 (1H, s), 7.55-7.60 (2H, m),7.24-7.40 (13H, m), 6.42 (1H, s), 5.68 (1H, d, J = 12.0 Hz), 5.63 (1H, d, J =11.5 Hz), 4.55-4.64 (3H, m), 4.43-4.44 (2H, m), 4.17 (1H, d, J = 2.5 Hz),4.06-4.08 (2H, m), 3.69 (1H, d, J = 10.0 Hz), 3.55 (1H, d, J = 10.5 Hz),2.26-2.32 (1H, m), 1.41-1.44 (1H, m)

[0595] (Example 6-2) Preparation of 9-{2,6-anhydro-3-O-benzyl-4-[(benzyloxy)methyl]-5-deoxy-α-L-lyxo-hexofuranosyl}-6-(benzyloxy)-N-(2-methylpropanoyl)-9H-purin-2-amine (Compound 27)

[0596] [ka]

[0597] Compound 26 (34.9 mg, 58.26 μmol) obtained in Example 6-1, isobutylamide (7.6 mg, 87.39 μmol), tris(dibenzylideneacetone)(chloroform)dipalladium (3.0 mg, 2.91 μmol), 4,5'-bis(diphenylphosphino)-9,9'-dimethylxanthene (3.4 mg, 5.83 μmol), and cesium carbonate (36.1 mg, 110.69 μmol) were added to a reaction vessel, and then toluene (0.7 mL) degassed under a nitrogen atmosphere was added. The reaction solution was heated to 110°C and stirred for 12 hours, after which tap water (1.0 mL) and ethyl acetate (4.0 mL) were added and the mixture was separated. The obtained organic layer was concentrated to dryness, and the residue was purified by silica gel chromatography (hexane:ethyl acetate=1:1) and concentrated under reduced pressure to obtain compound 27 as a colorless oily compound (18.1 mg, 47.8%). 1 H-NMR (CD3Cl): δ = 8.37 (1H, s), 7.83 (1H, brs),7.53-7.55 (2H, m), 7.24-7.39 (13H, m), 6.35 (1H, s), 5.67 (1H, d, J = 12.0 Hz),5.62 (1H, d, J = 12.0 Hz), 4.56-4.64 (3H, m), 4.43-4.45 (2H, m), 4.18 (1H, d, J= 2.5 Hz), 4.07-4.12 (2H, m), 3.70 (1H, d, J = 11.0 Hz), 3.56 (1H, d, J = 10.5Hz), 3.22 (1H, brs), 2.27-2.23 (1H, m), 1.43-1.46 (1H, m), 1.28 (6H, dd, J =6.8 Hz, 2.3 Hz)

[0598] (Example 6-3) Preparation of 9-[2,6-anhydro-5-deoxy-4-(hydroxymethyl)-α-L-lyxo-hexofuranosyl]-2-(2-methylpropanamido)-1,9-dihydro-6H-purin-6-one (Compound 28)

[0599] [ka]

[0600] 20% palladium hydroxide carbon (18.0 mg) was added to an ethanol solution (2.0 mL) of compound 27 (17.7 mg, 27.24 μmol) obtained in Example 6-2, and the mixture was stirred at 45° C. for 3 hours under a hydrogen pressure of 3.5 bar. After confirming the completion of the reaction, the mixture was cooled to room temperature and filtered. The filtrate was concentrated to dryness to obtain compound 28 as white crystals (11.7 mg, 101.0%). 1 H-NMR (DMSO-d6): δ = 12.10 (1H, s), 11.79 (1H, s), 8.31 (1H, s),6.21 (1H, s), 5.39 (1H, brs), 5.25 (1H, brs), 4.14 (1H, d, J = 3.5 Hz), 4.10(1H, d, J = 3.0 Hz), 3.88-3.90 (2H, m), 3.61 (1H, d, J = 12.5 Hz), 2.75-2.81(1H, m), 2.00-2.07 (1H, m), 1.12 (6H, d, J = 6.5 Hz)

[0601] (Example 6-4) Preparation of 9-(2,6-anhydro-4-{[bis(4-methoxyphenyl)(phenyl)methoxy]methyl}-5-deoxy-α-L-lyxo-hexofuranosyl)-2-(2-methylpropanamido)-1,9-dihydro-6H-purin-6-one (Compound 29)

[0602] [ka]

[0603] A solution (5 v / w) of compound 28 obtained in Example 6-3 in tetrahydrofuran is reacted with dimethoxytriphenylmethyl chloride (1.5 equivalents). Toluene (10 v / w) and water (5 v / w) are added to the reaction solution, which is then separated and concentrated. Compound 29 is obtained by purifying the mixture by silica gel column chromatography.

[0604] (Example 6-5) Preparation of Compound 1g

[0605] [ka]

[0606] Compound 29 obtained in Example 6-4 is added with dichloromethane (10 v / w), molecular sieve 4A (0.5 w / w), 2-cyanoethyl N,N,N',N'-tetraisopropyl phosphorodiamidite (1.5 equivalents), and 4,5-dicyanoimidazole (0.5 equivalents), and stirred at room temperature for 24 hours. Ethyl acetate (900 mL) and water (5 v / w) are added to the reaction solution, which is then separated and concentrated, and the mixture is purified by silica gel column chromatography to obtain compound 1g.

[0607] (Example 7) Preparation of Compound 17 (Glycosylation via Compound 22) (Example 7-1) Preparation of an acetonitrile solution of compound 22

[0608] [ka]

[0609] To a solution of compound 2 (3.00 g, 8.42 mmol) in toluene (30 mL), N,N-dimethylaminopyridine (0.05 g, 0.42 mmol), pyridine (1.02 mL, 12.63 mmol) and acetic anhydride (1.20 mL, 12.63 mmol) were added, and the mixture was stirred at 20 to 30 ° C for 1 hour. Then, water (15 mL) was added and the mixture was separated. The organic layer obtained was washed twice with 20% aqueous citric acid solution (15 mL), once with 8% aqueous sodium bicarbonate solution (9 mL) and once with water (9 mL). The organic layer obtained was concentrated under reduced pressure until it became 6 mL, and then solvent replacement by vacuum concentration with acetonitrile (30 mL) was performed twice to obtain an acetonitrile solution (6 mL) of compound 22.

[0610] ( Working Example 7-2) Preparation of 1-propanol solution of compound 3t

[0611] [ka]

[0612] Thymine (1.33 g, 10.52 mmol) and N,O-bistrimethylsilylacetamide (5.15 mL, 21.04 mmol) were added to acetonitrile (7.5 mL), and the solution was stirred at 20 to 30 ° C for 1 hour. The solution was added with an acetonitrile solution (6 mL) of the compound 22 obtained in Example 7-1, acetonitrile (9 mL) and iodotrimethylsilane (1.50 mL, 10.52 mmol) and stirred at 45 to 55 ° C for 4 hours. After adding an 8% aqueous sodium hydrogen carbonate solution (24 mL) and sodium sulfite (1.20 g), toluene (30 mL) was added, and the mixture was stirred at 20 to 30 ° C for 30 minutes, and then separated. The obtained organic layer was washed three times with 20% aqueous methanol (18 mL), and the obtained organic layer was concentrated under reduced pressure until it became 9 mL. To the resulting concentrated solution, 1-propanol (30 mL) was added, and the mixture was concentrated again under reduced pressure to obtain a 1-propanol solution (9 mL) of compound 3t.

[0613] The NMR spectrum of the obtained compound was consistent with the NMR spectrum of the compound described in Example 6 of WO00 / 47599.

[0614] (Example 7-3) Preparation of Compound 16

[0615] [ka]

[0616] To the 1-propanol solution (9 mL) of compound 3t, 1-propanol (15 mL), water (6 mL) and 5% palladium carbon (0.67 g) were further added, and the mixture was stirred for 2 hours at 55-65 ° C under a hydrogen atmosphere, after which the catalyst was filtered at the same temperature and washed with 80% 1-propanol water (6 mL). The obtained filtrate was concentrated under reduced pressure to obtain a 1-propanol solution (9 mL), and then acetonitrile (30 mL) was added and the mixture was concentrated under reduced pressure to obtain 9 mL of acetonitrile solution (A). Further acetonitrile (6 mL) was added and the mixture was stirred for 2 hours at 20-30 ° C. The resulting crystals were filtered and washed with acetonitrile (9 mL). The obtained solid was dried under reduced pressure (40 ° C) to obtain compound 16 (1.57 g, yield 65.6% (from compound 2)). In the above procedure, when the water content in the acetonitrile solution (A) exceeded 4%, the operation of adding acetonitrile and concentrating it to 9 mL was repeated, and the reproducibility of recovery by crystallization could be maintained by controlling the water content to 4% or less. The water content was confirmed by the Karl Fischer method.

[0617] The NMR spectrum of the obtained compound was identical to that of the compound described in Example 7 of WO00 / 47599.

[0618] (Example 7-4a) Preparation of Compound 17

[0619] [ka]

[0620] Pyridine (33.39 g) and 4,4'-dimethoxytrityl chloride (42.91 g) were added to a tetrahydrofuran solution (150 mL) of compound 16 (30 g), and the mixture was stirred at 20-30°C for 3 hours. Then, methanol (6 mL) was added and the mixture was stirred at 20-30°C for 30 minutes. A 15% aqueous sodium carbonate solution (150 mL), ethyl acetate (300 mL) and water (210 mL) were added to the obtained solution and the mixture was separated. The obtained organic layer was washed twice with a 20% aqueous citric acid solution (150 mL), once with a 5% aqueous sodium hydrogen carbonate solution (150 mL) and once with water (150 mL). The obtained organic layer was concentrated under reduced pressure until the volume was 150 mL, and then solvent replacement was performed twice with ethyl acetate (300 mL) by vacuum concentration, and ethyl acetate (90 mL) was added to obtain an ethyl acetate solution (240 mL) of compound 17. The obtained solution was added with n-heptane (90 mL) and heated to 50°C, then seed crystals (30 mg) were added and stirred at the same temperature for 2 hours. The obtained slurry was cooled to 20-30°C and stirred at the same temperature for 15 hours. The obtained slurry was added with n-heptane (120 mL) and stirred at the same temperature for another hour. The precipitated crystals were collected by filtration and washed with a 1:1 mixture of ethyl acetate and n-heptane (100 mL). The obtained crystals were dried under reduced pressure (50°C) to obtain compound 17 (60.7 g, yield 98%).

[0621] The NMR spectrum of the obtained compound was consistent with the NMR spectrum of the compound described in Example 8 of WO00 / 47599.

[0622] The seed crystals of compound 17 used were those precipitated by leaving an ethyl acetate solution of compound 17 obtained in Example 2-3 to stand. Example 8a: Preparation of Compound 1a

[0623] [ka]

[0624] The hydrate of compound 21 obtained in Example 4-3 (45.00 g, 64.31 mmol) was added to dichloromethane (675 mL), and the mixture was concentrated under reduced pressure to obtain a dichloromethane solution (225 mL). Dichloromethane (450 mL) was added to the obtained dichloromethane solution, and the mixture was concentrated under reduced pressure to obtain a dichloromethane solution (225 mL). The same operation was repeated, and dichloromethane (225 mL), molecular sieve 4A (22.50 g), dibutylhydroxytoluene (1.42 g, 6.44 mmol), 2-cyanoethyl N,N,N',N'-tetraisopropylphosphorodiamidite (21.32 g, 70.73 mmol), and 4,5-dicyanoimidazole (0.91 g, 7.71 mmol) were added to the obtained dichloromethane solution (225 mL), and the mixture was stirred at 20 to 30 °C for 16.5 hours. After confirming the completion of the reaction, ethyl acetate (900 mL) was added to the reaction solution, neutral silica gel (67.50 g) was added at -5 to 5 °C, and the mixture was stirred for 15 minutes. The neutral silica gel was removed by filtration, and the mixture was washed with dichloromethane-ethyl acetate (1 / 2, 225 mL) to obtain a dichloromethane-ethyl acetate solution. The obtained dichloromethane-ethyl acetate solution was concentrated under reduced pressure to obtain an ethyl acetate solution (225 mL). Methyl tert-butyl ether (135 mL) was added to the obtained solution to obtain solution A (360 mL). Methyl tert-butyl ether (315 mL) and n-heptane (1350 mL) were added to a separate reaction vessel, and solution A (360 mL) was added at -5 to 5 °C to confirm the precipitation of a solid. After washing with ethyl acetate (23 mL) and stirring for 0.5 hours, the precipitated solid was filtered and washed with methyl tert-butyl ether-n-heptane (1 / 3, 225 mL) and n-heptane (225 mL) precooled to 0° C. The solid was dried under reduced pressure (40° C.) to obtain compound 1a (48.69 g, 84.1%).

[0625] The NMR spectrum of the obtained compound was consistent with that of the compound described in Example 14 of WO00 / 47599.

[0626] Example 8b: Preparation of Compound 1a The hydrate (3.57 g) of compound 21 obtained in Example 4-3 was added to dichloromethane (45 mL) and concentrated under reduced pressure to obtain a dichloromethane solution (15 mL). Dichloromethane (15 mL), molecular sieves 4A (22.50 g), 2-cyanoethyl N,N,N',N'-tetraisopropyl phosphorodiamidite (1.42 g) and 4,5-dicyanoimidazole (100 mg) were added to the obtained dichloromethane solution, and the reaction mixture was stirred at 20 to 30 °C for 24 hours. After confirming the completion of the reaction, ethyl acetate (60 mL) was added to the reaction solution. The obtained solution was cooled to -5 to 5 °C, passed through a column filled with neutral silica gel (4.5 g), and washed with dichloromethane-ethyl acetate (1 / 2, 30 mL) to obtain a dichloromethane-ethyl acetate solution. The obtained dichloromethane-ethyl acetate solution was concentrated under reduced pressure to obtain an ethyl acetate solution (15 mL), and then toluene (60 mL) was added and concentrated again to obtain a toluene solution (15 mL). Methyl tert-butyl ether (9 mL) was added to the obtained solution to obtain solution A (24 mL). Methyl tert-butyl ether (6 mL) and n-heptane (90 mL) were added to another reaction vessel, and solution A (24 mL) was added dropwise at -5 to 5 °C to confirm the precipitation of a solid. The apparatus used for the dropwise addition was washed with toluene (1.5 mL), and the mixture was stirred for 2 hours, after which the precipitated solid was filtered and washed with n-heptane (24 mL). The obtained solid was dried under reduced pressure (40 °C) to obtain compound 1a (3.97 g, 86.5%).

[0627] The NMR spectrum of the obtained compound was consistent with that of the compound described in Example 14 of WO00 / 47599.

[0628] Example 9: Preparation of Compound 1t

[0629] [ka]

[0630] Compound 17 (5.00 g, 8.52 mmol) obtained in Example 2-3 was added to ethyl acetate (40 mL), molecular sieves 4A (2.5 g), 2-cyanoethyl N,N,N',N'-tetraisopropyl phosphorodiamidite (2.83 g, 9.38 mmol), and 4,5-dicyanoimidazole (0.20 g, 1.70 mmol) were added, and the mixture was stirred at 20 to 30 °C for 24 hours. After confirming the completion of the reaction, neutral silica gel (10.00 g) was added to the reaction solution, and the mixture was stirred for 30 minutes. The neutral silica gel was removed by filtration, and the mixture was washed with ethyl acetate (100 mL) to obtain an ethyl acetate solution. The obtained ethyl acetate solution was concentrated under reduced pressure to obtain an ethyl acetate solution (35 mL). The obtained ethyl acetate solution was added with tert-butyl ether (100 mL), and the mixture was concentrated under reduced pressure to obtain a methyl tert-butyl ether solution (35 mL). The obtained methyl tert-butyl ether solution was added with seed crystals of compound 1t (2.5 mg), stirred for 2 hours, added with n-heptane (100 mL), cooled to 0-5°C, and stirred for another 2 hours. The precipitated solid was filtered and washed with n-heptane (100 mL) pre-cooled to 0°C. The obtained solid was dried under reduced pressure (40 °C) to obtain compound 1t (6.12 g, 91.3%).

[0631] The NMR spectrum of the obtained compound was consistent with the NMR spectrum of the compound described in Example 9 of WO00 / 47599.

[0632] The seed crystals of Compound 1t were obtained by precipitating Compound 1t obtained in Example 2-4 by leaving a mixed solution of ethyl acetate, diisopropyl ether, and n-heptane to stand. Example 10: Preparation of Compound 1c (Example 10-1) Preparation of 1-(3-O-acetyl-2,6-anhydro-4-{[bis(4-methoxyphenyl)(phenyl)methoxy]methyl}-5-deoxy-α-L-lyxo-hexofuranosyl)-5-methylpyrimidine-2,4(1H,3H)-dione (Compound 30)

[0633] [ka]

[0634] Compound 17 (80.00 g, 136.37 mmol) obtained in Example 2-3, N,N-dimethylaminopyridine (1.67 g, 13.64 mmol) and triethylamine (27.60 g, 272.74 mmol) were added to ethyl acetate (320 mL), cooled to 0-5°C, and acetic anhydride (16.71 g, 163.64 mmol) was added at the same temperature. After stirring at 0-5°C for 1 hour, water (160 mL) was added and the organic layer was separated and washed with 10% citric acid aqueous solution (160 mL), 5% sodium bicarbonate aqueous solution (160 mL) and 10% saline (160 mL). The organic layer was concentrated under reduced pressure to 240 mL, and then solvent replacement by vacuum concentration with acetonitrile (400 mL) was performed twice to obtain an acetonitrile solution of compound 30 (240 mL).

[0635] (Example 10-2) Preparation of 1-(3-O-acetyl-2,6-anhydro-4-{[bis(4-methoxyphenyl)(phenyl)methoxy]methyl}-5-deoxy-α-L-lyxo-hexofuranosyl)-4-amino-5-methylpyrimidin-2(1H)-one (Compound 31)

[0636] [ka]

[0637] Acetonitrile (160 mL), N,N-dimethylaminopyridine (3.33 g, 27.27 mmol) and triethylamine (55.20 g, 545.51 mmol) were added to the solution of compound 30 obtained in Example 10-1, and the mixture was cooled to 5-15°C, and then 2,4,6-triisopropylbenzenesulfonyl chloride (47.50 g, 156.83 mmol) was added at the same temperature. After stirring at 10°C for 2 hours, 25% aqueous ammonia (100 kg, 156.83 mmol) was added and stirred at 20°C for 2 hours. The resulting solution was concentrated under reduced pressure to about 320 mL, and then ethyl acetate (400 mL) and water (120 mL) were added and separated. The resulting organic layer was washed twice with 10% saline (160 mL), and then acetonitrile (400 mL) was added and the mixture was concentrated under reduced pressure to about 320 mL. Acetonitrile (400 mL) was added to the obtained concentrated solution and concentrated under reduced pressure to obtain an acetonitrile solution (B) (240 mL) of compound 31. In the above procedure, when the water content in the acetonitrile solution (B) exceeded 0.4%, the procedure of adding acetonitrile to the acetonitrile solution (B) and concentrating and dehydrating it was repeated until the water content in the acetonitrile solution (B) became 0.4% or less, whereby the next reaction could be rapidly carried out. The water content was confirmed by the Karl Fischer method.

[0638] (Example 10-3a) Preparation of Compound 19

[0639] [ka]

[0640] To the acetonitrile solution of compound 31 obtained in Example 10-2, THF (80 mL) and benzoic anhydride (67.87 g, 300.01 mmol) were added, and the mixture was stirred at 40° C. for 5 hours, cooled to 25° C., and further stirred for 15 hours. The obtained slurry was dissolved in water (80 mL), 25% aqueous sodium hydroxide solution (176 g) and THF (80 mL) at 5-15 ° C over 1 hour or more. THF (160 mL) was added to the resulting mixture, and the mixture was stirred at 20 ° C for 2 hours, and the pH of the solution was adjusted to 6.8 using acetic acid. The hydrate crystals (80 mg) of compound 19 obtained in Example 3-3 were added as seed crystals to the resulting solution, and the mixture was stirred for 10 hours, and water (400 mL) was added dropwise at 15-25 ° C over 2 hours or more. The resulting slurry was stirred at 20 ° C for 1 hour, and the precipitated crystals were filtered and washed with 40% THF water (400 mL). The resulting crystals were dried under reduced pressure at 50 ° C to obtain compound 19 (78.35 g, 83.3% G / G (from compound 17)).

[0641] The NMR spectrum of the obtained compound was consistent with that of the compound described in Example 21 of WO00 / 47599.

[0642] (Example 10-3b) Preparation of Compound 19 THF (80 mL) and benzoic anhydride (23.14 g, 102.3 mmol) were added to the acetonitrile solution (90 mL) of compound 31 obtained in Example 10-2, and the mixture was stirred at 40°C for 5 hours, then cooled to 25°C and stirred for another 20 hours. THF (30 mL) and potassium acetate (23.09 g, 235.3 mmol) were added to the obtained slurry, and the mixture was stirred for 1 hour, and then 8% saline (135 mL) was added and the mixture was separated. THF (15 mL), water (15 mL) and 25% aqueous sodium hydroxide solution (33 g) were added to the obtained organic layer, and the mixture was cooled to 0-10°C, and then 25% aqueous sodium hydroxide solution (15 g) and water (6 mL) were added, and the mixture was stirred at 0-10°C for 4 hours. After confirming the completion of the reaction, the pH of the obtained solution was adjusted to 6.5 using acetic acid. The obtained solution was added with salt (6.0 g) and THF (60 mL) and separated, and the obtained organic layer was added with THF (30 mL), water (45 mL), and the hydrate crystals (30 mg) of compound 19 obtained in Example 3-3 as seed crystals, and after stirring for 16 hours, water (180 mL) was added dropwise at 20-30 ° C for more than 2 hours. The obtained slurry was stirred at 20-30 ° C for 3 hours, and the precipitated crystals were filtered and washed with 40% THF water (150 mL). The obtained crystals were dried under reduced pressure at 50 ° C to obtain compound 19 (28.48 g, 80.7% G / G (from compound 17)). In the above procedure, by using potassium acetate, the amount of acetic acid used for adjusting the pH during crystallization could be controlled to a smaller amount than in the procedure described in Example 10-3a, thereby shortening the time required for crystallization.

[0643] The NMR spectrum of the obtained compound was consistent with that of the compound described in Example 21 of WO00 / 47599.

[0644] Example 11: Preparation of Compound 1c

[0645] [ka]

[0646] Compound 19 (20.00 g, 29.00 mmol) obtained in Example 10-3 was added to dichloromethane (400 mL), and the mixture was concentrated under reduced pressure to obtain a dichloromethane solution (200 mL). Dichloromethane (200 mL) was added to the obtained dichloromethane solution, and the mixture was concentrated under reduced pressure to obtain a dichloromethane solution (200 mL). The same operation was repeated, and molecular sieves 4A (10.00 g), dibutylhydroxytoluene (0.64 g, 2.90 mmol), 2-cyanoethyl N,N,N',N'-tetraisopropylphosphorodiamidite (9.61 g, 31.89 mmol), and 4,5-dicyanoimidazole (1.03 g, 8.70 mmol) were added to the obtained dichloromethane solution (200 mL), and the mixture was stirred at 20 to 30 °C for 4 hours. After confirming the completion of the reaction, ethyl acetate (100 mL) was added to the reaction solution, neutral silica gel (40.00 g) was added at -5 to 5 °C, and the mixture was stirred for 30 minutes. The neutral silica gel was removed by filtration, and the mixture was washed with dichloromethane-ethyl acetate (2 / 1, 100 mL) to obtain a dichloromethane-ethyl acetate solution. The obtained dichloromethane-ethyl acetate solution was concentrated under reduced pressure to obtain a dichloromethane-ethyl acetate solution (100 mL). Ethyl acetate (100 mL) was added to the obtained dichloromethane-ethyl acetate solution, and the mixture was concentrated under reduced pressure to obtain an ethyl acetate solution (100 mL). The same operation was performed again, and methyl tert-butyl ether (100 mL) was added to the obtained solution, and the mixture was concentrated under reduced pressure to obtain an ethyl acetate-methyl tert-butyl ether solution (100 mL). Methyl tert-butyl ether (100 mL) was added to the obtained ethyl acetate-methyl tert-butyl ether solution, and the mixture was concentrated under reduced pressure to obtain a methyl tert-butyl ether solution (100 mL). The same operation was repeated, and n-heptane (200 mL) was added to the obtained solution and stirred for 1.5 hours. After further addition of n-heptane (100 mL) and stirring for 1 hour, n-heptane (200 mL) was added again and stirred for 1.5 hours to confirm the precipitation of a solid. The precipitated solid was filtered and washed with n-heptane (200 mL) pre-cooled to 0 ° C. The obtained solid was dried under reduced pressure (40 ° C) to obtain compound 1c (22.40 g, 86.8%).

[0647] The NMR spectrum of the obtained compound was consistent with the NMR spectrum of the compound described in Example 6 of WO00 / 47599.

[0648] Example 12: Stereoselective glycosylation of compound 22

[0649] [ka]

[0650] 1,2-dichloroethane (0.3 mL) was added to compound 22 (30 mg, 0.075 mmol) and bistrimethylsilylthymine (40.8 mg, 0.151 mmol) obtained in Example 5-1, and the mixture was stirred at room temperature. Then, 2.0 equivalents (0.151 mmol) of chlorotrimethylsilane (TMSCl), bromotrimethylsilane (TMSBr), iodotrimethylsilane (TMSI), and trimethylsilyl trifluoromethanesulfonate (TMSOTf) were added as activators relative to the raw materials, and the mixture was stirred at the temperature shown in the table. The reaction solution was sampled, and the raw materials, α-adducts, and β-adducts were analyzed by HPLC. The results are shown in Table 1.

[0651] [Table 1]

[0652] As a result of the investigation, it was found that the α-adduct was stereoselectively obtained under the conditions using TMSOTf, while the β-adduct was stereoselectively obtained under the conditions using TMSBr and TMSI. In particular, the reaction proceeded smoothly even at room temperature when TMSI was used.

[0653] (Example 13) Screening of activators for the amidite formation process

[0654] [ka]

[0655] Compound 21 hydrate (100 mg, 0.14 mmol) obtained in Example 4-3, 2-cyanoethyl N,N,N',N'-tetraisopropyl phosphorodiamidite (51.7 mg, 0.17 mmol), and activators 5-benzylthiotetrazole, 5-phenyltetrazole, 4,5-dicyanoimidazole, and 2,4,5-tetrabromoimidazole were added to dichloromethane (1 mL) in an amount of 1.15 equivalents (0.16 mmol) relative to the raw materials, and the mixture was stirred at 20 to 30 ° C. The results of the reaction completion confirmed by HPLC are shown in Table 2.

[0656] [Table 2]

[0657] As a result of the investigation, good results were obtained using 4,5-dicyanoimidazole and 2,4,5-tribromoimidazole.

[0658] (Example 14) Verification of molecular sieve effect

[0659] [ka]

[0660] Compound 17 obtained in Example 2-3, dibutylhydroxytoluene (0.10 equivalents), 2-cyanoethyl N,N,N',N'-tetraisopropylphosphorodiamidite (1.10 equivalents), and 4,5-dicyanoimidazole (1.0 equivalents) were added to dichloromethane (10 v / w), and stirred at 20 to 30 ° C. in the presence or absence of molecular sieve 4A (0.5 wt %). The progress of the reaction was confirmed by HPLC and the results are shown in Table 3.

[0661] [Table 3]

[0662] By using molecular sieves, compound 17 was converted to compound 1t without any residue.

[0663] Example 15: Synthesis of oligonucleotides Oligonucleotides having the desired sequence and structure can be synthesized by the following method.

[0664] The synthesis is carried out using an automatic nucleic acid synthesizer ("ABI 394 DNA / RNA Synthesizer" manufactured by Applied Biosystems) according to the phosphoramidite method (Nucleic Acids Research, 12, 4539 (1984)). The reagents used were Activator Solution-3 (0.25 mol / L 5-benzylthio-1H-tetrazole in acetonitrile, Wako Pure Chemical Industries, product No. 013-20011), CAP A for AKTA (1-methylimidazole in acetonitrile, Sigma-Aldrich, product No. L040050), Cap B1 for AKTA (acetic anhydride in acetonitrile, Sigma-Aldrich, product No. L050050), Cap B2 for AKTA (pyridine in acetonitrile, Sigma-Aldrich, product No. L050150), or DCA Deblock (dichloroacetic acid in toluene, Sigma-Aldrich, product No. L023050). As a thiolation reagent for forming phosphorothioate bonds, phenylacetyl disulfide (Carbosynth, product No. FP07495) was dissolved in a 1:1 (v / v) solution of acetonitrile (dehydrated, Kanto Chemical, product No. 01837-05) and pyridine (dehydrated, Kanto Chemical, product No. 11339-05) to a concentration of 0.2 M. As an amidite reagent, 2'-O-Me nucleoside phosphoramidites (adenosine product No. ANP-5751, cytidine product No. ANP-5752, guanosine product No. ANP-5753, uridine product No. ANP-5754) manufactured by ChemGenes were used.ENA monomer 9-(2,6-anhydro-4-{[bis(4-methoxyphenyl)(phenyl)methoxy]methyl}-3-O-{(2-cyanoethoxy)[di(propan-2-yl)amino]phosphanyl}-5-deoxy-α-L-lyxo-hexofuranosyl)-N-benzoyl-9H-purin-6-amine (compound 1a), 1-(2,6-anhydro-4-{[bis(4-methoxyphenyl)(phenyl)methoxy]methyl}-3-O-{(2-cyanoethoxy)[di(propan-2-yl)amino]phosphanyl}-5 -deoxy-α-L-lyxo-hexofuranosyl)-4-benzamido-5-methylpyrimidin-2(1H)-one (compound 1c), and 1-(2,6-anhydro-4-{[bis(4-methoxyphenyl)(phenyl)methoxy]methyl}-3-O-{(2-cyanoethoxy)[di(propan-2-yl)amino]phosphanyl}-5-deoxy-α-L-lyxo-hexofuranosyl)-5-methylpyrimidin-2,4(1H,3H)-dione (compound 1t) are the compounds synthesized in the above examples. Glen Unysupport 0.1 μmol (GlenResearch) is used as a solid phase support to synthesize oligonucleotides having the desired sequence. The program used was for 0.2 μmol scale attached to the automatic nucleic acid synthesizer, with the proviso that the time required for condensation of the amidite was 600 seconds, and the time required for thiolation was 150 seconds. An oligonucleotide having a ligand unit at the 5' end can be synthesized by similarly reacting a phosphoramidite of the ligand unit following synthesis of a nucleotide chain according to the method described in WO2019 / 172286. 18 and X 20 The corresponding phosphoramidite compounds used are compound 39D of Reference Example 39 and compound 41D of Reference Example 41 of WO2019 / 172286, respectively.

[0665] The protected oligonucleotide analogue having the target sequence is treated with 300 μL of concentrated aqueous ammonia to cleave the oligomer from the support, and the protecting groups on the phosphorus atom (cyanoethyl group) and the nucleic acid bases (nucleobases) are removed. The oligomer is purified using a Clarity QSP (Phenomenex) according to the attached protocol. [Industrial Applicability]

[0666] According to the present invention, a crystalline 2,4-bridged common intermediate useful for producing various ENA monomers and a stereoselective method for producing the intermediate have been provided, which makes it possible to efficiently produce various ENA monomers.

Claims

1. General formula (I) 【Chemistry 1】 [In the formula, Z 1 and Z 2 are the same or different and represent a protecting group for a hydroxyl group, R represents a hydrogen atom or an aliphatic acyl group, and n represents an integer of 1 to 4.

2. A compound represented by formula (I'): 【Chemistry 2】

3. A compound represented by formula (I''): 【Chemistry 3】

4. General formula (II) 【Chemistry 4】 [In the formula, Z 1 and Z 2 are the same or different and represent a protecting group for a hydroxyl group, and n represents an integer of 0 to 4. (i) General formula (III) 【Chemistry 5】 [In the formula, Z 1 , Z 2 and n are as defined above, and Y is a methyl group substituted with 1 to 3 aryl groups, a methyl group substituted with 1 to 3 aryl groups in which the aryl ring is substituted with a lower alkyl, lower alkoxy, halogen or cyano group, a lower alkoxymethyl group, a tetrahydropyranyl group or a silyl group, in the presence of an acid catalyst in a lower alkyl alcohol solvent to deprotect Y; (ii) The compound of general formula (IV) obtained in step (i) 【Chemistry 6】 [In the formula, Z 1 , Z 2 and n are as defined above, and A is a lower alkyl group; (iii) The compound represented by formula (V) obtained in step (ii) 【Chemistry 7】 [In the formula, Z 1 , Z 2 wherein A and n are as defined above, A method comprising:

5. Z 1 and Z 2 is a benzyl group, A is a methyl group; Y is a trityl group; n is 1, the acid catalyst is sulfuric acid, p-toluenesulfonic acid or methanesulfonic acid; Step (ii) is carried out using a phosphorus reagent and an azodicarboxylate, the phosphorus reagent being triphenylphosphine or tri(n-butyl)phosphine, and the azodicarboxylate being diethyl azodicarboxylate, diisopropyl azodicarboxylate or di-t-butyl azodicarboxylate; and The process according to claim 4, wherein step (iii) is carried out using an acid, the acid being hydrochloric acid, sulfuric acid, trifluoroacetic acid, methanesulfonic acid or p-toluenesulfonic acid.

6. General formula (VI) 【Chemistry 8】 [In the formula, R 1 represents a lower alkyl group or a hydrogen atom, R 2 represents a hydroxyl group, an amino group, or an amino group protected by an aliphatic acyl group or an aromatic acyl group; P 1 represents a trityl group which may be substituted by 1 to 3 lower alkoxy groups, and n represents an integer of 0 to 4, or a salt thereof, comprising the steps of: (i) General formula (II) 【Chemistry 9】 [In the formula, Z 1 and Z 2 are the same or different and represent a protecting group for a hydroxyl group, and n represents an integer of 0 to 4.], with an activating agent in a solvent to convert the hydroxyl group at position 1 into a group that forms a leaving group; (ii) The compound represented by formula (VII) obtained in step (i) 【Chemistry 10】 [In the formula, Z 1 , Z 2 and n are as defined above; X 1 represents a group that forms a leaving group. 【Chemistry 11】 [In the formula, R 1 and R 2 has the same meaning as defined above.] or a salt thereof is reacted in a solvent in the presence of a halogenating agent to obtain a compound represented by the general formula (IX): 【Chemistry 12】 [In the formula, Z 1 , Z 2 , R 1 , R 2 and n are as defined above, (iii) The compound of formula (IX) obtained in step (ii) 【Chemistry 13】 [In the formula, Z 1 , Z 2 , R 1 , R 2 and n are as defined above.] or a salt thereof is reacted with a deprotecting reagent for a hydroxyl group in a solvent to obtain Z 1 and Z 2 and deprotecting (iv) reacting the diol compound or a salt thereof obtained in the step (iii) with a protecting reagent for a primary hydroxyl group to obtain a compound represented by the general formula (VI): 【Chemistry 14】 [In the formula, P 1 , R 1 , R 2 and n are as defined above, A method comprising:

7. Z 1 and Z 2 is a benzyl group, P 1 is a 4,4'-dimethoxytrityl group, X 1 is an iodine atom, an acetoxy group, or a trichloroacetimidooxy group, n is 1, R 1 is a methyl group, R 2 is a hydroxyl group or a benzoylamino group, the activating agent is acetic anhydride, benzoic anhydride, trichloroacetonitrile, carbonyldiimidazole or diphenyl chlorophosphate; and 7. The method of claim 6, wherein the halogenating agent is chlorotrimethylsilane, bromotrimethylsilane or iodotrimethylsilane.

8. General formula (X) 【Chemistry 15】 [In the formula, R 3 represents an aliphatic acyl group or an aromatic acyl group; P 1 represents a trityl group which may be substituted by 1 to 3 lower alkoxy groups, and n represents an integer of 0 to 4, or a salt thereof, comprising the steps of: (i) General formula (II) 【Chemistry 16】 [In the formula, Z 1 and Z 2 and n is the same or different and represents a protecting group for a hydroxyl group, and is as defined above, with an activating agent in a solvent to convert the hydroxyl group at position 1 into a group that forms a leaving group; (ii) The compound represented by formula (XI) obtained in step (i) 【Chemistry 17】 [In the formula, Z 1 , Z 2 and n are as defined above; X 2 represents a group that forms a leaving group. 【Chemistry 18】 [In the formula, R 3 or a salt thereof in a solvent in the presence of an acid reagent; (iii) Then, the compound is isomerized to obtain a compound of the general formula (XIII): 【Chemistry 19】 [In the formula, Z 1 , Z 2 , R 3 and n are as defined above, (iv) The compound represented by the general formula (XIII) obtained in step (iii) 【Chemistry 20】 [In the formula, Z 1 , Z 2 , R 3 and n are as defined above.] or a salt thereof is reacted with a deprotecting reagent for a hydroxyl group in a solvent to obtain Z 1 and Z 2 and deprotecting (v) reacting the diol compound or a salt thereof obtained in the step (iv) with a protecting reagent for a primary hydroxyl group to selectively protect the primary hydroxyl group, thereby obtaining a compound represented by the general formula (X): 【Chemistry 21】 [In the formula, R 3 , n and P 1 has the same meaning as defined above, or a salt thereof; A method comprising:

9. Z 1 and Z 2 is a benzyl group, P 1 is a 4,4'-dimethoxytrityl group, X 2 is an acetoxy group, n is 1, R 3 is a benzoyl group, the activating agent is acetic anhydride, benzoic anhydride, trichloroacetonitrile, carbonyldiimidazole or diphenyl chlorophosphate; and 9. The method of claim 8, wherein the acid reagent is trimethylsilyl trifluoromethanesulfonate and trifluoroacetic acid.

10. General formula (XIV) 【Chemical 22】 [In the formula, Z 1 and Z 2 are the same or different and represent a protecting group for a hydroxyl group, and n represents an integer of 0 to 4; or a salt thereof, (i) General formula (II) 【Chemistry 23】 [wherein Z 1 and Z 2 are the same or different and each represents a protecting group for a hydroxyl group, and n is as defined above] is reacted with an activating agent in a solvent to convert the hydroxyl group at position 1 into a group that forms a leaving group; (ii) The compound represented by formula (XI) obtained in step (i) 【Chemistry 24】 [wherein Z 1 , Z 2 and n are as defined above, and X 2 is a group forming a leaving group.] is reacted with silylated dichloropurine in a solvent in the presence of a halogenating agent to obtain a compound represented by the general formula (XIV): 【Chemistry 25】 [wherein Z 1 , Z 2 and n are as defined above] or a salt thereof; A method comprising:

11. The method according to claim 10, wherein the compound represented by general formula (XIV) is represented by formula (XIV'). 【Chemistry 26】

12. General formula (XVIII) 【Chemical 27】 [In the formula, P 1 represents a trityl group optionally substituted by 1 to 3 lower alkoxy groups; R 4 represents an aliphatic acyl group or an aromatic acyl group, and n represents an integer of 0 to 4, or a salt thereof, comprising the steps of: (0) A compound represented by the general formula (XIV) according to the method of claim 10 【Chemistry 28】 [wherein Z 1 and Z 2 are the same or different and each represents a protecting group for a hydroxyl group, and n represents an integer of 0 to 4] or a salt thereof, (i) The compound represented by formula (XIV) prepared in step (0) 【Chemical 29】 [wherein Z 1 , Z 2 and n are as defined above] or a salt thereof is reacted with benzyl alcohol which may be substituted with a lower alkyl, lower alkoxy, halogen or cyano group in a solvent in the presence of a base to substitute the chlorine atom at the 6-position of the purine ring with a benzyloxy group which may be substituted with a lower alkyl, lower alkoxy, halogen or cyano group; (ii) The compound represented by formula (XIX) obtained in step (i) 【Chemistry 30】 [In the formula, Z 1 , Z 2 and n are as defined above; R 5 represents a benzyl group which may be substituted with a lower alkyl, a lower alkoxy, a halogen or a cyano group, or a salt thereof, is cross-coupled with an amidating agent in a solvent in the presence of a palladium catalyst and a phosphine ligand to obtain a compound represented by the general formula (XX): 【Chemistry 31】 [In the formula, Z 1 , Z 2 , R 4 , R 5 and n are as defined above, (iii) The compound of formula (XX) obtained in step (ii) 【Chemistry 32】 [In the formula, Z 1 , Z 2 , R 4 , R 5 and n are as defined above.] or a salt thereof is reacted with a deprotecting reagent for a hydroxyl group in a solvent to obtain Z 1 , Z 2 and R 5 and deprotecting (iv) The compound of general formula (XXI) obtained in step (iii) 【Chemical Formula 33】 [In the formula, R 4 and n are as defined above.] or a salt thereof is reacted with a protecting reagent for a primary hydroxyl group to selectively protect the primary hydroxyl group, thereby obtaining a compound represented by the general formula (XVIII): 【Chemical 34】 [In the formula, P 1 , R 4 and n are as defined above, A method comprising:

13. General formula (XXII) 【Chemistry 35】 [In the formula, R 1 represents a lower alkyl group or a hydrogen atom, R 6 represents an aliphatic acyl group or an aromatic acyl group; P 1 represents a trityl group which may be substituted by 1 to 3 lower alkoxy groups, and n represents an integer of 0 to 4, or a salt thereof, comprising the steps of: （０） The method according to claim 6, 【Chemical 36】 [wherein R 1 represents a lower alkyl group or a hydrogen atom, R 2 represents a hydroxyl group, P 1 represents a trityl group which may be substituted with 1 to 3 lower alkoxy groups, and n represents an integer of 0 to 4] or a salt thereof; (i) A compound represented by general formula (XXIII), which is a tautomer of the compound represented by general formula (VI) or a salt thereof produced in step (0), 【Chemical 37】 [In the formula, P 1 , R 1 and n are as defined above, or a salt thereof, with a hydroxyl-protecting reagent in a solvent to protect the hydroxyl group at the 3'-position; (ii) The compound represented by general formula (XXIV) obtained in step (i) 【Chemical Formula 38】 [In the formula, P 1 , R 1 and n are as defined above; Z 3 represents an aliphatic acyl group or an aromatic acyl group, or a salt thereof, with an activating agent in a solvent in the presence of a base and a catalyst; (iii) Then, by reacting with an aminating agent, a compound represented by the general formula (XXV) 【Chemical Formula 39】 [In the formula, P 1 , R 1 , Z 3 and n are as defined above, (iv) The compound of general formula (XXV) obtained in step (iii) 【Chemistry 40】 [In the formula, P 1 , R 1 , Z 3 and n are as defined above, or a salt thereof, with an acylating agent in a solvent; (v) The compound represented by general formula (XXVI) obtained in step (iv) 【Chemistry 41】 [In the formula, P 1 , R 1 , R 6 , Z 3 and n are as defined above.] or a salt thereof is reacted with a deprotecting reagent for a hydroxyl group to deprotect only Z 3 , thereby obtaining a compound represented by the general formula (XXII): 【Chemistry 42】 [wherein P 1 , R 1 , R 6 and n are as defined above] or a salt thereof. A method comprising:

14. P 1 is a trityl group, Z 3 is an acetyl group, n is 1, R 1 is a methyl group or a hydrogen atom, R 6 is a benzoyl group, and The method of claim 13, wherein the activating agent is p-toluenesulfonyl chloride or 2,4,6-triisopropylbenzenesulfonyl chloride.

15. By the method according to claim 6, a compound represented by the general formula (XXVII) 【Chemistry 43】 [In the formula, P 1 represents a trityl group which may be substituted by 1 to 3 lower alkoxy groups, and B represents a 2-oxo-4-hydroxy-5-methylpyrimidin-1-yl group, a 2-oxo-4-amino-pyrimidin-1-yl group in which the amino group is protected, or a 4-amino-5-methyl-2-oxo-pyrimidin-1-yl group in which the amino group is protected, with the proviso that the protecting group for the amino group of B is an aliphatic acyl group or an aromatic acyl group, and n represents an integer of 0 to 4.] or a salt thereof, and reacting the compound or a salt thereof with an amidite reagent in a solvent in the presence of an activator and a desiccant, General formula (XXVIII) 【Chemistry 44】 [In the formula, P 1 , B and n are as defined above. or a salt thereof.

16. P 1 is a 4,4'-dimethoxytrityl group, B is a 2-oxo-4-hydroxy-5-methylpyrimidin-1-yl group, a 2-oxo-4-benzoylamino-pyrimidin-1-yl group, a 4-benzoylamino-5-methyl-2-oxo-pyrimidin-1-yl group, n is 1, the activator is 4,5-dicyanoimidazole; The amiditizing reagent is 2-cyanoethyl N,N,N',N'-tetraisopropyl phosphorodiamidite, and 16. The method of claim 15, wherein the desiccant is molecular sieve 3A, molecular sieve 4A, or molecular sieve 5A.

17. By the method according to claim 8, a compound represented by the general formula (XXVII) 【Chemistry 45】 [In the formula, P 1 represents a trityl group which may be substituted by 1 to 3 lower alkoxy groups, B represents a 6-aminopurin-9-yl group in which the amino group is protected, with the proviso that the protecting group for the amino group of B represents an aliphatic acyl group or an aromatic acyl group, and n represents an integer of 0 to 4, or a salt thereof, and reacting the compound or the salt thereof with an amidite reagent in a solvent in the presence of an activator and a desiccant, General formula (XXVIII) 【Chemistry 46】 [In the formula, P 1 , B and n are as defined above. or a salt thereof.

18. P 1 is a 4,4'-dimethoxytrityl group, B is a 6-benzoylaminopurin-9-yl group; n is 1, the activator is 4,5-dicyanoimidazole; The amiditizing reagent is 2-cyanoethyl N,N,N',N'-tetraisopropyl phosphorodiamidite, and 18. The method of claim 17, wherein the desiccant is molecular sieve 3A, molecular sieve 4A, or molecular sieve 5A.

19. By the method according to claim 12, a compound represented by the general formula (XXVII) 【Chemistry 47】 [In the formula, P 1 represents a trityl group which may be substituted by 1 to 3 lower alkoxy groups, B represents a 2-amino-6-hydroxypurin-9-yl group in which the amino group is protected, with the proviso that the protecting group for the amino group of B represents an aliphatic acyl group or an aromatic acyl group, and n represents an integer of 0 to 4, or a salt thereof, and reacting the compound or the salt thereof with an amidite reagent in a solvent in the presence of an activator and a desiccant, General formula (XXVIII) 【Chemistry 48】 [In the formula, P 1 , B and n are as defined above. or a salt thereof.

20. P 1 is a 4,4'-dimethoxytrityl group, B is a 2-isobutyrylamino-6-hydroxypurin-9-yl group; n is 1, the activator is 4,5-dicyanoimidazole; The amiditizing reagent is 2-cyanoethyl N,N,N',N'-tetraisopropyl phosphorodiamidite, and 20. The method of claim 19, wherein the desiccant is molecular sieve 3A, molecular sieve 4A, or molecular sieve 5A.

21. A method for producing an oligonucleotide, comprising the steps of: (A) synthesizing an ENA (2'-O,4'-C-ethylene-bridged nucleic acid) monomer represented by general formula (XXVIII) by the method according to any one of claims 15 to 20; and (B) A step of extending a nucleotide chain according to a desired sequence using the ENA monomer obtained in step (A), a phosphoramidite compound of another nucleic acid, and / or a phosphoramidite compound of a ligand.

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