WO2021095875A1 - オリゴ核酸化合物の製造方法 - Google Patents

オリゴ核酸化合物の製造方法 Download PDF

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WO2021095875A1
WO2021095875A1 PCT/JP2020/042506 JP2020042506W WO2021095875A1 WO 2021095875 A1 WO2021095875 A1 WO 2021095875A1 JP 2020042506 W JP2020042506 W JP 2020042506W WO 2021095875 A1 WO2021095875 A1 WO 2021095875A1
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compound
general formula
alkyl
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French (fr)
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悠太朗 壽
光貴 加藤
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Nippon Shinyaku Co Ltd
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Nippon Shinyaku Co Ltd
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Priority to JP2021556192A priority Critical patent/JP7669280B2/ja
Priority to US17/776,356 priority patent/US12600744B2/en
Priority to CA3161589A priority patent/CA3161589A1/en
Priority to EP20888671.3A priority patent/EP4059943A4/en
Priority to CN202080092415.XA priority patent/CN114981280A/zh
Priority to KR1020227019574A priority patent/KR20220097986A/ko
Publication of WO2021095875A1 publication Critical patent/WO2021095875A1/ja
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Priority to JP2025067244A priority patent/JP2025111536A/ja
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    • C—CHEMISTRY; METALLURGY
    • C07—ORGANIC CHEMISTRY
    • C07H—SUGARS; DERIVATIVES THEREOF; NUCLEOSIDES; NUCLEOTIDES; NUCLEIC ACIDS
    • C07H21/00—Compounds containing two or more mononucleotide units having separate phosphate or polyphosphate groups linked by saccharide radicals of nucleoside groups, e.g. nucleic acids
    • C—CHEMISTRY; METALLURGY
    • C07—ORGANIC CHEMISTRY
    • C07H—SUGARS; DERIVATIVES THEREOF; NUCLEOSIDES; NUCLEOTIDES; NUCLEIC ACIDS
    • C07H21/00—Compounds containing two or more mononucleotide units having separate phosphate or polyphosphate groups linked by saccharide radicals of nucleoside groups, e.g. nucleic acids
    • C07H21/02—Compounds containing two or more mononucleotide units having separate phosphate or polyphosphate groups linked by saccharide radicals of nucleoside groups, e.g. nucleic acids with ribosyl as saccharide radical
    • C—CHEMISTRY; METALLURGY
    • C07—ORGANIC CHEMISTRY
    • C07H—SUGARS; DERIVATIVES THEREOF; NUCLEOSIDES; NUCLEOTIDES; NUCLEIC ACIDS
    • C07H1/00—Processes for the preparation of sugar derivatives
    • C—CHEMISTRY; METALLURGY
    • C07—ORGANIC CHEMISTRY
    • C07H—SUGARS; DERIVATIVES THEREOF; NUCLEOSIDES; NUCLEOTIDES; NUCLEIC ACIDS
    • C07H21/00—Compounds containing two or more mononucleotide units having separate phosphate or polyphosphate groups linked by saccharide radicals of nucleoside groups, e.g. nucleic acids
    • C07H21/04—Compounds containing two or more mononucleotide units having separate phosphate or polyphosphate groups linked by saccharide radicals of nucleoside groups, e.g. nucleic acids with deoxyribosyl as saccharide radical
    • Y—GENERAL TAGGING OF NEW TECHNOLOGICAL DEVELOPMENTS; GENERAL TAGGING OF CROSS-SECTIONAL TECHNOLOGIES SPANNING OVER SEVERAL SECTIONS OF THE IPC; TECHNICAL SUBJECTS COVERED BY FORMER USPC CROSS-REFERENCE ART COLLECTIONS [XRACs] AND DIGESTS
    • Y02—TECHNOLOGIES OR APPLICATIONS FOR MITIGATION OR ADAPTATION AGAINST CLIMATE CHANGE
    • Y02P—CLIMATE CHANGE MITIGATION TECHNOLOGIES IN THE PRODUCTION OR PROCESSING OF GOODS
    • Y02P20/00—Technologies relating to chemical industry
    • Y02P20/50—Improvements relating to the production of bulk chemicals
    • Y02P20/55—Design of synthesis routes, e.g. reducing the use of auxiliary or protecting groups

Definitions

  • the present invention relates to a method for producing a novel oligonucleic acid compound.
  • the solid-phase method is a heterogeneous reaction method in which a nucleic acid is extended while contacting a substrate supported on a solid-phase carrier with a solution containing a reaction reagent.
  • a so-called batch method is used in which a reaction vessel with a filter is used to carry out the reaction in the vessel (see, for example, Non-Patent Document 1 and Patent Document 1).
  • a pseudo flow synthesis method is also known, such as an automatic nucleic acid synthesizer (for example, DNA, RNA synthesizer), in which a solid-phase carrier is placed in a column and a solution containing a reaction reagent is allowed to flow through the column to cause a reaction.
  • the liquid phase method is a homogeneous reaction method in which nucleic acid is extended by reacting in a solution containing both a substrate and a reaction reagent.
  • a batch method in which the reaction is carried out in a container is used (see, for example, Patent Document 2 and Patent Document 3).
  • the chemical synthesis method of an oligonucleic acid compound removes the protecting group of an oxygen atom or an amino group on the nucleic acid compound.
  • the nucleic acid is extended by repeating the "deprotection” reaction and the “condensation” reaction of forming a bond between an oxygen atom or a nitrogen atom and a phosphorus atom that have been deprotected and can react.
  • controlling the reaction efficiency and reaction rate in the "condensation" reaction that forms a bond between a phosphorus atom and an oxygen atom or a nitrogen atom is very important in the production of oligonucleic acid compounds, and the conditions for this condensation reaction are It is an element that has a great impact on the production period of oligonucleic acid compounds.
  • the solid-phase method is a heterogeneous reaction between a solid-phase carrier and a solution, it is known that the reactivity of the condensation reaction decreases due to steric hindrance caused by the solid-phase carrier.
  • Polystyrene resin is generally used as the solid-phase carrier, but during the reaction, it swells due to the reaction solvent used, and its volume becomes larger than that in the dry state. The degree of swelling depends on the reaction solvent. Therefore, the reaction efficiency and reaction rate of the condensation reaction in the solid phase method depend on the reaction solvent used.
  • polystyrene resin does not have a large degree of swelling in a polar solvent such as acetonitrile, which is generally used for the synthesis of oligonucleic acid compounds, and the use of a polar solvent in the solid phase method means that the reaction efficiency of the condensation reaction is high. And it cannot be said that it is preferable from the aspect of improving the reaction speed.
  • a polar solvent such as acetonitrile
  • the liquid phase method is a homogeneous reaction method in which a reaction is carried out in a solution containing both a substrate and a reaction reagent. In order to remove the reaction solvent, column purification or the like is required. Similar to the liquid phase method, the synthetic method using a hydrophobic group-bonded nucleoside or a pseudo solid-phase protected nucleoside can react in a uniform system, so that the reaction efficiency is higher and the reaction rate is higher than that of the solid-phase method. fast.
  • non-protic solvents such as chloroform are used in the condensation reaction, as reported, for example, in the synthesis of morpholinon nucleic acids (see, eg, Patent Document 5). Since the condensation reaction in a non-protic solvent requires a very long time, it is preferable to use a non-protic solvent in the reaction of a homogeneous system from the viewpoint of improving the reaction efficiency and reaction rate of the condensation reaction. I can't say.
  • An object of the present invention is to provide a novel production method capable of shortening the production period of an oligonucleic acid compound.
  • the present inventors have found that the condensation reaction proceeds efficiently by forming a trivalent phosphorus bond in the condensation reaction of the oligonucleic acid compound, and completed the present invention.
  • substituted [3] represents; and A 2 is C 1-6 alkyl, mono- under basic conditions has been replaced with a leaving group capable of (C 1-6 alkyl) amino-C 1-6 alkyl, di (C 1-6 alkyl) amino-C 1-6 alkyl, tri (C 1-6 alkyl) ammonio-C 1-6 alkyl, under basic conditions Represents a removable group, aryl or heteroaryl. ) Represents a substituent represented by (hereinafter, referred to as "substituent [2]").
  • the compound [B] having a substituent containing a phosphorus atom represented by (hereinafter referred to as “substituent [1]”) is subjected to a condensation reaction, and the following general formula [C]:
  • W 0 and X are synonymous with the above;
  • A represents the residue obtained by removing one hydrogen atom of the hydroxyl group or the primary or secondary amino group of the compound from the compound [A];
  • B represents the substituent [1] from the compound [B].
  • a method for producing a compound represented by (hereinafter referred to as "Compound [C]”), which comprises a step of forming a trivalent phosphorus bond in the condensation reaction. Can be mentioned.
  • An oligonucleic acid compound is a compound having a structure in which two to a plurality of nucleoside units are linked via a phosphorus bond.
  • a condensation reaction many times to form a phosphorus bond between adjacent nucleoside units.
  • the present invention since phosphorus bonds can be efficiently formed, it can be expected that the production time of the oligonucleic acid compound will be shortened as a result.
  • FIG. 1 illustrates a schematic diagram of a reactor used for a continuous reaction.
  • F-1 to F-5 indicate solution vessels
  • P-1 to P-5 indicate pumps
  • R-1 to R-4 indicate flow reactors
  • S-1 to S-5 indicate supply streams. Show the road.
  • the present invention is a method for producing compound [C] by subjecting compound [A] having a hydroxyl group or a primary or secondary amino group and compound [B] having a substituent [1] to a condensation reaction. It is characterized in that a trivalent phosphorus bond is formed by the condensation reaction.
  • a compound containing one to a plurality of nucleoside units in its molecule can be mentioned.
  • a compound containing a nucleoside unit in the range of 1 to 50 is suitable, a compound containing a nucleoside unit in the range of 1 to 30 is preferable, and a compound containing a nucleoside unit in the range of 1 to 25 is preferable.
  • Compounds containing nucleoside units are more preferred.
  • nucleoside unit contained in the compound [A] examples include the following general formulas [4a] to [4d]: [During the ceremony, * Is (1) Bonding position of the phosphorus bond bonded to the oxygen atom at the 5'position of the adjacent nucleoside unit, (2) Bonding position with hydrogen atom or (3) The following general formula [6]: (During the ceremony, * Represents the binding position of compound [A] with a residue; G is (1) Silicon substituent, (2) Long-chain alkyl-carbonyl, (3) 1 to 5 benzoyls substituted with long-chain alkyloxy and / or long-chain alkenyloxy or (4) the following general formula [7]: (During the ceremony, * Represents the connection position with T; Z is (1) (Soluble polymer soluble in organic solvent) -Oxy, (2) (Soluble polymer soluble in organic solvent) -Amino, (3) Long-chain alkyloxy, (4) Solid-phase carrier or (5) The following general formulas [8A] to [
  • Substituents represented by (hereinafter, referred to as “substituent [9A]”, “substituent [9B]”, “substituent [9C]”, “substituent [9D]”, “substituent [9E]", respectively.
  • R 8d is the same or different, each of which is a hydrogen atom, a halogen, a long-chain alkyl optionally substituted with 1 to 13 halogens, or a long-chain alkyl optionally substituted with 1 to 13 halogens.
  • R 8e is (1) Long chain alkyl, Represents (2) long-chain alkyl-carbonyl or (3) benzoyl substituted with 1-5 long-chain alkyloxy and / or long-chain alkenyloxy; and R 8f (1) Long chain alkyl, Represents (2) long-chain alkyl-carbonyl or (3) long-chain alkenyl-carbonyl.
  • Substituents represented by hereinafter, "substituent [8A]”, “substituent [8B]”, “substituent [8C]”, “substituent [8D]", “substituent [8E]", respectively.
  • R 4b1 and R 4b2 represent the same or different hydrogen atoms or C 1-6 alkyl, respectively, or R 4b1 and R 4b2 combine with adjacent carbon atoms to form a carbonyl; and J Represents an oxygen atom or N-R 4b3 (R 4b3 stands for C 1-6 alkyl).
  • Examples thereof include nucleoside units represented by (hereinafter, referred to as “nucleoside unit [4a]”, “nucleoside unit [4b]”, “nucleoside unit [4c]”, and “nucleoside unit [4d]”, respectively).
  • Preferred embodiments of the nucleoside units [4a] to [4d] include, for example, the following general formulas [4a1] to [4d1]: [During the ceremony, d, BP , J, R 4a , R 4b1 and R 4b2 are synonymous with the above; * Is (1) Bonding position of the phosphorus bond bonded to the oxygen atom at the 5'position of the adjacent nucleoside unit, (2) Represents the bond position with a hydrogen atom or (3) the bond position with a substituent [6]; and ** are (1) Bonding position with a phosphorus bond bonded to an oxygen atom at the 3'position or a nitrogen atom at the 3'position of an adjacent nucleoside unit, It represents (2) a bond position with a hydrogen atom or (3) a bond position with a substituent [6].
  • nucleoside unit [4a1] examples of the nucleoside unit represented by (hereinafter, referred to as “nucleoside unit [4b1]”, “nucleoside unit [4c1]”, and “nucleoside unit [4d1]) can be mentioned.
  • nucleoside units in the compound [A] contains a plurality of nucleoside units in its molecule, it is preferable that adjacent nucleoside units in the compound are bonded to each other via a phosphorus bond.
  • the phosphorus bonds between the nucleoside units constituting the compound [A] are the same or different, for example, the following general formula [5]: [During the ceremony, X is synonymous with the above; One of * and ** represents the bond position of the nucleoside unit with the oxygen atom at the 3'position or the nitrogen atom of the 3'position, and the other represents the bond with the oxygen atom of the nucleoside unit at the 5'position different from the nucleoside unit.
  • W represents a lone pair of electrons, an oxygen atom or a sulfur atom.
  • Examples of the bond represented by (hereinafter referred to as “phosphorus bond [5]”) can be mentioned.
  • W preferably represents a lone pair of electrons or an oxygen atom, more preferably an oxygen atom.
  • (A-1) Compound [A] composed of one to a plurality of nucleoside units [4d]
  • nucleoside unit represented by the general formula [4d] *But, (1) the bond position with the phosphorus bond bonded to the oxygen atom at the 5'position of the adjacent nucleoside unit, or (2) the bond position with the hydrogen atom; **But, (1) The bond position with the phosphorus bond bonded to the nitrogen atom at the 3'position of the adjacent nucleoside unit, or (2) the bond position with the substituent [6].
  • examples of the phosphorus bond between each nucleoside unit constituting the compound [A] include the same or different phosphorus bond [5].
  • the phosphorus bond represented by the general formula [5] one of * and ** represents the bond position of the nucleoside unit with the nitrogen atom at the 3'position, and the other represents the bond position of the nucleoside unit different from the nucleoside unit. It represents the bond position with the oxygen atom at the 5'position.
  • the following general formula [A-1] [During the ceremony, BP , G, T, X and W are synonymous with the above; and n represents an integer from 1 to 50. ]
  • Examples of the compound represented by (hereinafter, referred to as “compound [A-1]”) can be mentioned.
  • An integer of 1 to 50 is suitable for n, an integer of 1 to 30 is preferable, and an integer of 1 to 25 is more preferable.
  • Equation [A-1-2] the following general formula [A-1-2]: Equation [A-1-2]: [During the ceremony, BP is a nucleobase that may be protected; Q 2 is a leaving group capable of either is H, or under acidic conditions; W represents a lone electron pair, an oxygen atom, or a sulfur atom, preferably a lone electron pair or an oxygen atom, and more preferably an oxygen atom; X, di (C 1-6 alkyl) amino; G is the following formula: [During the ceremony, * Represents the connection position with T. ] Selected from the group consisting of; T is a single bond; n is 1 to 25. ] Compounds include.
  • A-2) Compound [A] composed of one or more nucleoside units selected from the group consisting of nucleoside units [4a], nucleoside units [4b] and nucleoside units [4c]
  • nucleoside unit represented by the general formulas [4a], [4b] and [4c] *But, (1) the bond position with the phosphorus bond bonded to the oxygen atom at the 5'position of the adjacent nucleoside unit, or (2) the bond position with the substituent [6]; **But, It is (1) a bond position with a phosphorus bond bonded to an oxygen atom at the 3'position of an adjacent nucleoside unit, or (2) a bond position with a hydrogen atom.
  • the phosphorus bond [5] is suitable for the phosphorus bond between each nucleoside unit constituting the compound [A].
  • the phosphorus bond represented by the general formula [5] one of * and ** represents the oxygen atom at the 3'position of the nucleoside unit, and the other represents the oxygen atom at the 5'position of the nucleoside unit different from the nucleoside unit. Represents the bond position with an oxygen atom.
  • substituent [7] in the compound [A-1] and the compound [A-2] include the following substituents. [During the ceremony, * Represents the connection position with T. ]
  • a compound containing one to a plurality of nucleoside units in its molecule can be mentioned. More specifically, a compound containing a nucleoside unit in the range of 1 to 10 is suitable, a compound containing a nucleoside unit in the range of 1 to 7 is preferable, and a compound containing a nucleoside unit in the range of 1 to 5 is preferable. Compounds containing the nucleoside unit of are more preferred.
  • nucleoside unit contained in the compound [B] examples include the following general formulas [4e] to [4h]: [During the ceremony, d, BP , J, R 4a , R 4b1 and R 4b2 are synonymous with the above; *** is (1) Bonding position of the phosphorus bond bonded to the oxygen atom at the 5'position of the adjacent nucleoside unit, (2) Represents the bond position with the substituent [1] or (3) the bond position with a group that can be eliminated under acidic conditions; *** is (1) Bonding position with a phosphorus bond bonded to an oxygen atom at the 3'position or a nitrogen atom at the 3'position of an adjacent nucleoside unit, It represents (2) a bond position with a substituent [1] or (3) a bond position with a group that can be eliminated under acidic conditions.
  • nucleoside unit [4e] examples thereof include a nucleoside unit represented by (hereinafter, referred to as “nucleoside unit [4e]”, “nucleoside unit [4f]”, “nucleoside unit [4g]”, and “nucleoside unit [4h]", respectively).
  • Preferred embodiments of the nucleoside units [4e] to [4h] include, for example, the following general formulas [4e1] to [4h1]: [During the ceremony, d, BP , J, R 4a , R 4b1 and R 4b2 are synonymous with the above; *** is (1) Bonding position of the phosphorus bond bonded to the oxygen atom at the 5'position of the adjacent nucleoside unit, (2) Represents a bond position with a substituent [1] or (3) a bond position with a group that can be eliminated under acidic conditions; and *** (1) Bonding position with a phosphorus bond bonded to an oxygen atom at the 3'position or a nitrogen atom at the 3'position of an adjacent nucleoside unit, It represents (2) a bond position with a substituent [1] or (3) a bond position with a group that can be eliminated under acidic conditions.
  • nucleoside unit [4e1] examples of the nucleoside unit represented by (hereinafter, referred to as “nucleoside unit [4e1]”, “nucleoside unit [4f1]”, “nucleoside unit [4g1]”, and “nucleoside unit [4h1]) can be mentioned.
  • nucleoside unit [B] contains a plurality of nucleoside units in its molecule
  • adjacent nucleoside units in the compound are bonded to each other via a phosphorus bond.
  • examples of the phosphorus bond between each nucleoside unit constituting the compound [B] include the same or different phosphorus bond [5].
  • the phosphorus bond represented by the general formula [5] one of * and ** represents the oxygen atom at the 3'position of the nucleoside unit, and the other represents the oxygen atom at the 5'position of the nucleoside unit different from the nucleoside unit. Represents the bond position with an oxygen atom.
  • (B-1) Compound [B] composed of one to a plurality of nucleoside units [4h]
  • nucleoside unit represented by the general formula [4h] ***But, (1) the bond position with the phosphorus bond bonded to the oxygen atom at the 5'position of the adjacent nucleoside unit, or (2) the bond position with the group that can be eliminated under acidic conditions; ****But, (1) The bond position with the phosphorus bond bonded to the nitrogen atom at the 3'position of the adjacent nucleoside unit, or (2) the bond position with the substituent [1].
  • examples of the phosphorus bond between each nucleoside unit constituting the compound [B] include the same or different phosphorus bond [5].
  • the phosphorus bond represented by the general formula [5] one of * and ** represents the nitrogen atom at the 3'position of the nucleoside unit, and the other represents the 5'position of the nucleoside unit different from the nucleoside unit. Represents the bond position with an oxygen atom.
  • the following general formula [1A] [During the ceremony, D, W and X are synonymous with the above; and ** represent the binding position of compound [B] to a residue. ] It is appropriate to have a substituent containing a phosphorus atom represented by.
  • the following general formula [B-1] [During the ceremony, BP , D, X and W are synonymous with the above; p represents an integer from 1 to 10; and Q 1 represents a group that can be eliminated under acidic conditions. ] Examples of the compound represented by (hereinafter, referred to as “compound [B-1]”) can be mentioned.
  • An integer of 1 to 10 is suitable for p, an integer of 1 to 7 is preferable, and an integer of 1 to 5 is more preferable.
  • (B-2) Compound [B] composed of one to a plurality of nucleoside units selected from the group consisting of nucleoside units [4e], nucleoside units [4f] and nucleoside units [4 g].
  • nucleoside unit represented by the general formulas [4e], [4f] and [4g] ***But, (1) the bond position with the phosphorus bond bonded to the oxygen atom at the 3'position of the adjacent nucleoside unit, or (2) the bond position with the substituent [1]; ****But, It is (1) a bond position with a phosphorus bond bonded to an oxygen atom at the 5'position of an adjacent nucleoside unit, or (2) a bond position with a group that can be eliminated under acidic conditions.
  • compound [B] for example, a compound in which the oxygen atom at the 5'position of the nucleoside unit on the 5'-terminal side is replaced with a group that can be eliminated under acidic conditions can be mentioned. ..
  • examples of the phosphorus bond between each nucleoside unit constituting the compound [B] include the same or different phosphorus bond [5].
  • the phosphorus bond represented by the general formula [5] one of * and ** represents the oxygen atom at the 3'position of the nucleoside unit, and the other represents the oxygen atom at the 5'position of the nucleoside unit different from the nucleoside unit. Represents the bond position with an oxygen atom.
  • the following general formula [1B] [During the ceremony, D and X are synonymous with the above; and ** represent the binding position of compound [B] to a residue. ] It is appropriate to have a substituent containing a phosphorus atom represented by.
  • the following general formula [B-2] [During the ceremony, p, BP , D, Q 1 , R 4a , X and W are synonymous with the above. ] Examples of the compound represented by (hereinafter, referred to as “compound [B-2]”) can be mentioned.
  • compound [C] examples include compounds that can be produced by subjecting compound [A] and compound [B] to a condensation reaction.
  • (C-1) Compound [C] composed of one to a plurality of nucleoside units [4d] and one to a plurality of nucleoside units [4h].
  • the compound [C] for example, the following general formula [C-1]: [During the ceremony, n, p, BP , G, Q 1 , T, W and X are synonymous with the above. ]
  • Examples of the compound represented by (hereinafter, referred to as “compound [C-1]”) can be mentioned.
  • examples of the phosphorus bond newly formed in the method for producing the compound [C-1] by reacting the compound [A-1] with the compound [B-1] include a phosphorus bond [5]. Can be mentioned.
  • Examples of the phosphorus bond newly formed in the method for producing the compound [C-1-1] by reacting 1-1] include a trivalent phosphorus bond.
  • Compound [B-1-1] has a general formula: [In the formula, p, BP , Q 1 , W, and X are synonymous with the above. ] It is a compound represented by.
  • Compound [B-1-2] has a general formula: [In the formula, p, BP , Q 1 and X are synonymous with the above, W is a lone pair of electrons, and LG 1 is a leaving group such as halogen (chloro, bromo, iodo, especially chloro). is there. ] It is a compound represented by.
  • Compound [B-1-2] is a compound of compound [B-1], in which D is LG 1 and W is a lone electron pair.
  • C-2 One to a plurality of nucleoside units and nucleoside units [4e], nucleoside units [4f] and selected from the group consisting of nucleoside units [4a], nucleoside units [4b] and nucleoside units [4c].
  • Compound [C] composed of one to a plurality of nucleoside units selected from the group consisting of nucleoside units [4 g]
  • examples of the phosphorus bond newly formed in the method for producing the compound [C-2] by reacting the compound [A-2] with the compound [B-2] include the following general formula [ 5a]: [During the ceremony, X is synonymous with the above; and one of * and ** represents the bond position with the oxygen atom at the 3'position of the nucleoside unit, and the other represents the oxygen atom at the 5'position of the nucleoside unit different from the nucleoside unit. Represents the connection position of. ] A bond containing a phosphorus atom represented by (hereinafter, referred to as “phosphorus bond [5a]”) can be mentioned.
  • nucleobase examples include adenine, guanine, hypoxanthine, cytosine, thymine, uracil, and modified bases thereof.
  • modified bases include, for example, pseudouracil, 3-methyluracil, dihydrouracil, 5-alkylcytocin (eg, 5-methylcitosin), 5-alkyluracil (eg, 5-ethyluracil), 5-halouracil (5).
  • 6-azapyrimidine 6-alkylpyrimidine (6-methyluracil), 2-thiouracil, 4-thiouracil, 4-acetylcitosine, 5- (carboxyhydroxymethyl) uracil, 5'-carboxymethylaminomethyl 2-thiouracil, 5-carboxymethyl-aminomethyl uracil, 1-methyl-adenine, 1-methyl-hypoxanthine, 2,2-dimethyl guanine, 3-methylcytosine, 2-methyl-adenine, 2-methylguanine, N 6 - methyl Adenine, 7-methylguanine, 5-methoxyaminomethyl-2-thiouracil, 5-methylaminomethyluracil, 5-methylcarbonylmethyluracil, 5-methyloxyuracil, 5-methyl-2-thiouracil, 2-methylthio-N 6- Isopentenyladenin, uracil-5-oxyacetic acid, 2-thiocitosine, purine, 2,6-diamin
  • nucleobase related to BP may be protected.
  • optionally protected nucleobase includes both unprotected “nucleobase” and protected “nucleobase”, eg, amino groups and / or hydroxyl groups. Includes adenine, guanine, hypoxanthin, cytosine, thymine, uracil, etc., which are unprotected or protected.
  • the protecting group for the amino group is not particularly limited as long as it is used as a protecting group for nucleic acids.
  • examples thereof include phenoxyacetyl, 4-tert-butylphenoxyacetyl, 4-isopropylphenoxyacetyl, and (dimethylamino) methylene.
  • benzoyl, acetyl, phenylacetyl and 4-tert-butylphenoxyacetyl are preferable.
  • the protecting group for the hydroxyl group includes, for example, 2-cyanoethyl, 4-nitrophenethyl, phenylsulfonylethyl, methylsulfonylethyl, trimethylsilylethyl, and 1 to 5 electron-withdrawing groups at arbitrary substitutable positions.
  • As the protecting group for the hydroxyl group 2-cyanoethyl, 4-nitrophenethyl and 4- (tert-butylcarboxy) benzyl are preferable.
  • the protecting group for the 6-position hydroxyl group of guanine is preferably 2-cyanoethyl.
  • the protected nucleobases include, for example, those shown below. [In the formula, Pg represents a protecting group]. More specific protected nucleic acid base embodiments include benzoyl-protected adenine ( ABz ), amino groups benzoyl-protected cytosine ( CBz ), and hydroxyl groups protected by 2-cyanoethyl. And, but not limited to, guanine (GCE, Pac ) in which the amino group is protected with phenoxyacetyl.
  • the "long chain alkyl” refers, for example, to a linear or branched alkyl having 10 to 300 carbon atoms, preferably a linear or branched alkyl having 10 to 100 carbon atoms, and more. Preferred is a linear or branched chain alkyl having 10 to 30 carbon atoms.
  • the "long-chain alkyl” portion of the “long-chain alkyl-carbonyl” and “long-chain alkyloxy” can be the same as the "long-chain alkyl”.
  • “Long-chain alkenyl” For example, a linear or branched alkenyl having 10 to 300 carbon atoms, preferably a linear or branched alkenyl having 10 to 100 carbon atoms, more preferably.
  • the "long-chain alkenyl” portion of the “long-chain alkenyloxy” and the “long-chain alkenyl-carbonyl” can be the same as the "long-chain alkenyl".
  • Examples of the "halogen” include a fluorine atom, a chlorine atom, a bromine atom, and an iodine atom.
  • the "5- to 6-membered saturated cyclic amino” includes, for example, a 5- to 6-membered saturated cyclic amino having 1 or 2 N, which may have one O or S as a ring-constituting atom.
  • C 1-6 alkyl indicates a linear or branched alkyl having 1 to 6 carbon atoms, and specifically, for example, methyl, ethyl, n-propyl, isopropyl, n-butyl, isobutyl. , Se-butyl, tert-butyl, n-pentyl, n-hexyl.
  • C 1-6 alkoxy represents a linear or branched chain alkoxy having 1 to 6 carbon atoms, and specifically, for example, methoxy, ethoxy, n-propoxy, isopropoxy, n-butoxy, and the like. Examples thereof include isobutoxy, sec-butoxy, tert-butoxy, n-pentyloxy, and n-hexyloxy.
  • C 1-6 alkoxy moiety of the "C 1-6 alkoxy -C 1-6 alkyl” may include the same ones as the "C 1-6 alkoxy”.
  • di (C 1-6 alkyl) amino mono (amino -C 1-6 alkyl substituted with removable group under basic conditions) amino, di (removable under basic conditions groups
  • the "C 1-6 alkyl” portion of the alkyl) amino and the di (amino-C 1-6 alkyl) amino can be the same as the "C 1-6 alkyl" described above.
  • C 2-10 alkylene is a divalent product produced by removing one hydrogen atom bonded to a different constituent carbon atom from a linear or branched alkyl having 2 to 10 carbon atoms.
  • the group include an ethylene group, a propylene group, an isopropylene group, a butylene group, a pentylene group, and a hexylene group.
  • Such "alkylene” may be substituted with 1 to 12 halogens at any substitutable position. According to L 1 as “alkylene”, ethylene is particularly preferred.
  • C 6-10 arylene removes two hydrogen atoms bonded to two different ring-constituting carbon atoms from a monocyclic or polycyclic aromatic hydrocarbon having 6 to 10 carbon atoms. It is a divalent group produced by, and examples thereof include phenylene and naphthylene. Such “allylen” may be substituted with 1 to 6 halogens at any substitutable position. According to L 1 as “arylene”, phenylene are particularly preferred.
  • Examples of the "group that can be removed under acidic conditions” include trityl, monomethoxytrityl, tert-butyldimethylsilyl, and dimethoxytrityl.
  • Examples of the “group that can be removed under basic conditions” include trifluoroacetyl.
  • Examples of the “group that can be removed under neutral conditions” include a group that can be eliminated by the action of tetrabutylammonium fluoride or a hydrogen trifluoride / triethylamine salt.
  • 2 -Cyanoethoxymethoxy, 2-cyanoethoxy-2-ethoxy, tert-butyldimethylsilyl can be mentioned.
  • Examples of the "silicon substituent” include triphenylsilyl, diisopropylphenylsilyl, tert-butyldimethylsilyl, and tert-butyldiphenylsilyl.
  • Aryl can be, for example, phenyl.
  • Heteroaryl can include, for example, pyridyl, pyrimidyl, pyrariayl, pyrazinyl, thienyl, flanyl.
  • solid phase carrier generally, any substance that can be used for solid phase synthesis of nucleic acids, peptides, peptide nucleic acids, sugars and the like can be used without any particular problem. For example, polystyrene can be used.
  • Polyglass; CPG Oxalylated-Regular Glass (see, eg, Nucleic Acids Research, Vol. 19, 1527 (1991)), TentaGel Support-Aminopolystyrene Glycol Derivatized Support (eg, Tetrahedron Letters, Vol.) .34, 3373 (1993)), Polys-polystyrene / divinylbenzene copolymers, polystyrene-based resins, polyacrylamide-based resins.
  • soluble polymer soluble in an organic solvent include a styrene polymer having no crosslink and a polyethylene glycol derivative.
  • the "soluble polymer soluble in organic solvent” portion of "(soluble polymer soluble in organic solvent) -oxy" and “(soluble polymer soluble in organic solvent) -amino” is the above-mentioned "soluble polymer soluble in organic solvent”.
  • Examples of the “non-crosslinked styrene polymer” include derivatives (TentaGel series, ArgoGel series) of polystyrene not crosslinked with divinylbenzene and having a spacer such as polyethylene glycol.
  • Examples of the "polyethylene glycol derivative” include derivatives having a substituent on polyethylene glycol having a molecular weight of 100 to 40,000 (SUNBRIGHT (registered trademark) series).
  • the solvent that can be used in this production method is not particularly limited as long as it is a solvent usually used in the art, and the solvent may be used alone, or two or more solvents may be mixed and used. You may.
  • the solvent that can be used in this production method include aromatic solvents such as benzene, toluene, xylene, and mesitylene; ester solvents such as ethyl acetate and isopropyl acetate; hexane, pentane, heptane, octane, nonane, cyclohexane, and the like. Examples thereof include an aliphatic solvent and a halogen solvent. These solvents may be mixed and used.
  • halogen-based solvent examples include chloroform, dichloromethane, 1,1-dichloroethane, 1,2-dichloroethane, 1,1,2-trichloroethane, 1,2-dichloroethylene, or a mixed solvent thereof. Can be mentioned. Of these, chloroform, dichloromethane, 1,1-dichloroethane, and 1,2-dichloroethane are preferable. In this production method, a base may be used if necessary.
  • Examples of the "base” that can be used in this production method include diisopropylamine, N, N-diisopropylethylamine, triethylamine, N-ethylmorpholine, and 2,6-lutidine.
  • the amount of the base that can be used in this production method is, for example, appropriately in the range of 1 to 100 times the molar ratio with respect to 1 mol of the compound [A], and 1 to 10 times the amount.
  • the amount is preferably in the range of 1 to 5 times, and more preferably in the range of 1 to 5 times.
  • additives may be used if necessary.
  • the additive for example, LiBr, LiCl, LiI, and NaI are preferable.
  • the amount of the additive that can be used in this production method is, for example, appropriately in the range of 0.2 times to 6.0 times the molar ratio with respect to 1 mol of compound [A], and is 0.
  • the amount is preferably in the range of 4 times to 3.0 times, and more preferably in the range of 1.0 times to 2.5 times.
  • the reaction temperature is, for example, preferably in the range of ⁇ 78 ° C. to 130 ° C., preferably in the range of ⁇ 40 ° C. to 100 ° C., and more preferably in the range of 0 ° C. to 80 ° C.
  • the reaction time varies depending on the type of compound [A] used, the type of compound [B], the type of reaction solvent, the type of base, and the reaction temperature, but for example, the range of 1 minute to 300 minutes is appropriate. It is preferably in the range of 5 minutes to 120 minutes.
  • this production method can be applied by either a batch method or a flow method. Further, this production method can also be applied to a solid-phase method, a liquid-phase method, a liquid-phase method using a hydrophobic group-bonded nucleoside, a pseudo-solid-phase protected nucleoside, etc., which are known as methods for producing an oligonucleic acid compound.
  • compound [C] which is an oligonucleic acid compound
  • compound [C] can be produced by using the solid phase method, at the oxygen atom at the 3'position of the nucleoside unit on the 3'terminal side of compound [A] or at the 5'of compound [A].
  • the oxygen atom at the 5'position of the nucleoside unit on the terminal side, which is supported on a solid phase carrier, can be used.
  • the oligonucleic acid compound can be produced using the liquid phase method, at the oxygen atom at the 3'position of the nucleoside unit on the 3'end side of compound [A] or on the nucleoside unit on the 5'end side of compound [A].
  • those supported by a soluble polymer that dissolves in an organic solvent can be used.
  • an oligonucleic acid compound can be produced by using a liquid phase method using a hydrophobic group-bonded nucleoside, a pseudo-solid phase-protected nucleoside, or the like
  • the oxygen atom at the 3'position of the nucleoside unit on the 3'terminal side of compound [A] can be produced.
  • one having a hydrophobic group bonded or one supported on a pseudo solid phase can be used (for example,).
  • Compound [C-1] can be produced by subjecting compound [A-1] to a condensation reaction with compound [B-1].
  • the solvent that can be used in this production method is not particularly limited as long as it is a solvent usually used in the art, and the solvent may be used alone, or two or more solvents may be mixed and used. You may.
  • Examples of the solvent that can be used in this production method include aromatic solvents such as benzene, toluene, xylene, and mesityrene; ester solvents such as ethyl acetate and isopropyl acetate; hexane, pentane, heptane, octane, nonane, cyclohexane, and the like.
  • Alibo-based solvents such as acetonitrile and propionitrile, ethers such as THF, 1,4-dioxane and diethyl ether, amides such as dimethylformamide and dimethylacetamide, 1,3-dimethyl -2-Imidazolidinone and the like. These solvents may be mixed and used.
  • the halogen-based solvent that can be used in this production method include chloroform, dichloromethane, 1,1-dichloroethane, 1,2-dichloroethane, 1,1,2-trichloroethane, 1,2-dichloroethylene, or a mixed solvent thereof. Can be mentioned.
  • a base may be used if necessary.
  • the "base” that can be used in this production method include diisopropylamine, N, N-diisopropylethylamine, triethylamine, N-ethylmorpholine, and 2,6-lutidine.
  • the amount of the base that can be used in this production method is, for example, appropriately in the range of 1 to 100 times the molar ratio with respect to 1 mol of the compound [A], and 1 to 10 times the amount.
  • the amount is preferably in the range of 1 to 5 times, and more preferably in the range of 1 to 5 times.
  • additives may be used if necessary.
  • the "additive" that can be used in this production method for example, LiBr, LiCl, LiI, and NaI are preferable.
  • the amount of the additive that can be used in this production method is, for example, appropriately in the range of 0.2 times to 6.0 times the molar ratio with respect to 1 mol of compound [A], and is 0.
  • the amount is preferably in the range of 4 times to 3.0 times, and more preferably in the range of 1.0 times to 2.5 times.
  • the reaction temperature is, for example, preferably in the range of ⁇ 78 ° C.
  • reaction time varies depending on the type of compound [A] used, the type of compound [B], the type of reaction solvent, the type of base, and the reaction temperature, but for example, the range of 1 minute to 300 minutes is appropriate. It is preferably in the range of 5 minutes to 120 minutes.
  • compound [A-1] has a solid phase carrier in its molecule, that is, when G is a substituent [7] and Z is a solid phase carrier in compound [A-1], for example, ( 1) Fill a suitable column with compound [A-1] and elute the reaction solution containing compound [B-1], or (2) place compound [A-1] and compound [A-1] in a reaction vessel equipped with a filter. This condensation reaction can be carried out by shaking or stirring the reaction solution containing B-1].
  • G is substituted with (1) silicon substituent, (2) long chain alkyl-carbonyl, (3) 1-5 long chain alkyloxy and / or long chain alkenyloxy. If it is a benzoyl or (4) substituent [7] (except when Z is a solid phase carrier), for example, (1) compound [A-1] is placed in a suitable reaction vessel. [A-1] and compound [B-1] are stirred in the reaction solvent, or (2) the solution containing the compound [A-1] and the solution containing the compound [B-1] are independent of each other. This condensation reaction can be carried out by supplying the solution to the flow reactor or the reaction flow path via the supply flow path and mixing the solutions in the flow reactor or the like.
  • the "supply flow path” means a flow path for continuously supplying the solution
  • the “reaction flow path” means a flow path capable of reacting while flowing the solution.
  • the flow reactor means a reactor in which a solution is charged, a reaction is carried out, and a product is recovered at the same time, and the operation is continuous.
  • a pump for supplying a liquid usually used in this field specifically examples include a syringe pump, a plunger pump, a diaphragm pump, and a gear pump.
  • Examples of the distribution reactor include an in-line mixer such as a microreactor and a static mixer.
  • a multi-stage collision type micromixer As a means for guiding the solution containing the compound [A-1] and the solution containing the compound [B-1] from the supply flow path to the reaction flow path, for example, a multi-stage collision type micromixer can be mentioned.
  • the material of the supply channel and the reaction channel is selected from the group consisting of, for example, a fluororesin such as perfluoroalkoxy alkane (PFA), a vinyl chloride resin, a polyamide resin, and an aromatic polyetherketone resin.
  • PFA perfluoroalkoxy alkane
  • Examples include synthetic resin tubing or metal tubing selected from the group consisting of stainless steel, copper and its alloys and titanium and its alloys.
  • the inner diameters of the supply flow path and the reaction flow path may be appropriately selected from, for example, a size within the range of 0.1 mm to 1.0 mm, but for example, a size within the range of 0.2 mm to 1.0 mm. It is preferable to select from.
  • the process of forming the compound of Step 2) The compound of the general formula [B-1-2] can be used as a compound.
  • Step of forming the compound of, and step 3) A step of treating the compound of the general formula [C-1] with an oxidizing agent is included.
  • the general formula [C-1]: [In the formula, n, p, BP , Q 1 , G, T, W, and X are synonymous with the above.
  • Examples thereof include a method for preparing the compound of.
  • the process of forming the compound of Step 2') The compound of the general formula [A-1-3] can be used as the compound of the general formula [B-1-1]: [In the formula, p, BP , Q 1 , W, and X are synonymous with the above]
  • Step of forming the compound of, and step 3') A step of treating the compound of the general formula [C-1] with an oxidizing agent is included.
  • the general formula [C-1]: [In the formula, n, p, BP , Q 1 , G, T, W, and X are synonymous with the above.
  • Examples thereof include a method for preparing the compound of.
  • the amount of compound [P] is in the range of 0.6 times to 4.0 times the molar ratio of 1 mol of compound [A-1] or compound [B-1-1]. Therefore, it is preferable to use it in the range of 0.75 times to 1.5 times the amount.
  • a base may be used if necessary. Examples of the "base” that can be used in this step include N, N-diisopropylethylamine, triethylamine, 1,8-bis (dimethylamino) naphthalene, pyridine, 2,4,6-cholidine, N-ethylmorpholin, and diaza.
  • DBU Zabicycloundecene
  • DABCO 1,4-diazabicyclo [2.2.2] octane
  • N-methylimidazole preferably N, N-diisopropylethylamine, triethylamine, 1,8-bis ( Didimethylamino) naphthalene, pyridine, 2,4,6-colysine.
  • the amount of the base that can be used in this production method is, for example, appropriately in the range of 1 to 10 times the molar ratio with respect to 1 mol of compound [A] or compound [B], and 1 It is preferably in the range of double to five times the amount.
  • the solvent that can be used in this step is not particularly limited, but for example, dichloromethane, tetrahydrofuran, dimethyl sulfoxide, or a mixture thereof can be used, and dichloromethane, 10% tetrahydrofuran / dichloromethane, 10% dimethyl sulfoxide / dichloromethane can be used. Is preferred, and dichloromethane, 10% tetrahydrofuran / dichloromethane is even more preferred.
  • the reaction temperature in this step is not particularly limited, but is, for example, ⁇ 78 to 60 ° C.
  • the reaction time in this step is not particularly limited, but is, for example, 0.05 to 20 minutes, preferably 0.2 to 10 minutes.
  • the "base” that can be used in this step include N, N-diisopropylethylamine, triethylamine, 1,8-bis (dimethylamino) naphthalene, pyridine, 2,4,6-cholidine, N-ethylmorpholin, and diaza.
  • DBU Zabicycloundecene
  • DABCO 1,4-diazabicyclo [2.2.2] octane
  • N-methylimidazole preferably N, N-diisopropylethylamine, triethylamine, 1,8-bis ( Didimethylamino) naphthalene, pyridine, 2,4,6-colysine.
  • the amount of the base that can be used in this production method is, for example, within the range of 1 to 10 times the molar ratio with respect to 1 mol of the compound [A-1] or the compound [B-1-1]. Is appropriate, and is preferably in the range of 1 to 5 times the amount.
  • the solvent that can be used in this step is not particularly limited, but for example, dichloromethane, tetrahydrofuran2-methyl tetrahydrofuran, dimethyl sulfoxide, and a mixture thereof can be used, and dichloromethane, 10% tetrahydrofuran / dichloromethane, 10% can be used.
  • dichloromethane, 10% tetrahydrofuran / dichloromethane, 10% can be used.
  • 2-Methyl tetrahydrofuran / dichloromethane, 10% dimethyl sulfoxide / dichloromethane is preferable, and dichloromethane, 10% tetrahydrofuran / dichloromethane, 10% 2-methyl tetrahydrofuran / dichloromethane is more preferable.
  • the reaction temperature in this step is not particularly limited, but is, for example, ⁇ 78 to 60 ° C., preferably 0 to 60 ° C.
  • the reaction time of this step is not particularly limited, but is, for example, 0.5 to 60 minutes, preferably 2 to 30 minutes, and more preferably 1 to 30 minutes.
  • the amount of the oxidizing agent is in the range of 1 to 10 times the molar ratio with respect to 1 mol of the compound [C-1-1], and the amount is 1.5 to 3 times. Within the range is preferable.
  • the oxidizing agents are, for example, iodine, magnesium monoperoxyphthalate hexahydrate (MMPP), peracetic acid, metachloroperbenzoic acid (mCPBA), tert-butyl hydroperoxide (TBHP), N-methylmorpholine. Oxidizers, hydrogen peroxide and manganese dioxide can be used, with iodine and magnesium monoperoxyphthalate hexahydrate (MMPP) being preferred.
  • the solvent that can be used in this step is not particularly limited, but may be appropriately selected depending on, for example, the oxidizing agent to be used.
  • the oxidizing agent for example, water, chloroethane, dichloromethane, chloroform, tetrahydrofuran, or a mixture thereof is used. be able to.
  • the solvent that can be used in this step when iodine is used as the oxidizing agent, for example, dichloromethane, chloroform, tetrahydrofuran, 0.2% water / tetrahydrofuran can be used, and when MMPP is used, for example, dichloromethane, chloroform, tetrahydrofuran, 0.2% water / tetrahydrofuran can be used.
  • water can be used, if peracetic acid is used, for example dichloromethane can be used, if mCPBA is used, for example dichloromethane can be used, if TBHP is used, for example, for example.
  • the reaction temperature in this step is not particularly limited, but is, for example, 0 to 25 ° C.
  • the reaction time of this step is not particularly limited, but is, for example, 1 to 60 minutes, preferably 5 to 30 minutes.
  • compound [A-1] has a solid phase carrier in its molecule, that is, when G is a substituent [7] and Z is a solid phase carrier in compound [A-1], for example, ( 1) Fill a suitable column with compound [A-1] and elute the reaction solution containing compound [B-1-1], or (2) place compound [A-1] in a reaction vessel with a filter.
  • the condensation reaction can be carried out by shaking or stirring the reaction solution containing the compound [B-1-1].
  • G is substituted with (1) silicon substituent, (2) long chain alkyl-carbonyl, (3) 1-5 long chain alkyloxy and / or long chain alkenyloxy. If it is a benzoyl or (4) substituent [7] (except when Z is a solid phase carrier), for example, (1) compound [A-1] is placed in a suitable reaction vessel. [A-1] and compound [B-1-1] are stirred in a reaction solvent, or (2) a solution containing compound [A-1] and a solution containing compound [B-1-1]. Can be independently supplied to the flow reactor or the reaction flow path via the supply flow path, and the solutions can be mixed in the flow reactor or the like to carry out the main condensation reaction.
  • the "supply flow path” means a flow path for continuously supplying the solution
  • the “reaction flow path” means a flow path capable of reacting while flowing the solution.
  • the flow reactor means a reactor in which a solution is charged, a reaction is carried out, and a product is recovered at the same time, and the operation is continuous.
  • a pump for supplying a liquid usually used in this field Specific examples thereof include a syringe pump, a plunger pump, a diaphragm pump, and a gear pump.
  • the distribution reactor include an in-line mixer such as a microreactor and a static mixer.
  • a multi-stage collision type micromixer As a means for guiding the solution containing the compound [A-1] and the solution containing the compound [B-1-1] from the supply flow path to the reaction flow path, for example, a multi-stage collision type micromixer can be mentioned. it can.
  • the material of the supply channel and the reaction channel is selected from the group consisting of, for example, a fluororesin such as perfluoroalkoxy alkane (PFA), a vinyl chloride resin, a polyamide resin, and an aromatic polyetherketone resin.
  • PFA perfluoroalkoxy alkane
  • vinyl chloride resin vinyl chloride resin
  • polyamide resin polyamide resin
  • aromatic polyetherketone resin examples include synthetic resin tubing or metal tubing selected from the group consisting of stainless steel, copper and its alloys and titanium and its alloys.
  • the inner diameters of the supply flow path and the reaction flow path may be appropriately selected from, for example, a size within the range of 0.1 mm to 1.0 mm, but for example, a size within the range of 0.2 mm to 1.0 mm. It is preferable to select from.
  • Compound [C-2] can be produced by subjecting compound [A-2] to a condensation reaction with compound [B-2].
  • a base may be used if necessary.
  • Examples of the "base” that can be used in this production method include diisopropylamine, N, N-diisopropylethylamine, triethylamine, N-ethylmorpholine, and 2,6-lutidine.
  • compound [A-2] has a solid phase carrier in its molecule, that is, when G is a substituent [7] and Z is a solid phase carrier in compound [A-2], for example, ( 1) Fill a suitable column with compound [A-1] and elute the reaction solution containing compound [B-2], or (2) place compound [A-2] and compound [A-2] in a reaction vessel with a filter.
  • the condensation reaction can be carried out by shaking or stirring the reaction solution containing B-2].
  • G is substituted with (1) silicon substituent, (2) long chain alkyl-carbonyl, (3) 1-5 long chain alkyloxy and / or long chain alkenyloxy. If it is a benzoyl or (4) substituent [7] (except when Z is a solid phase carrier), for example, (1) compound [A-2] is placed in a suitable reaction vessel. [A-2] and compound [B-2] are stirred in the reaction solvent, or (2) the solution containing the compound [A-2] and the solution containing the compound [B-2] are independent of each other. This condensation reaction can be carried out by supplying the solution to the flow reactor or the reaction flow path via the supply flow path and mixing the solutions in the flow reactor or the like.
  • the compound [C-2] is obtained by purifying the reaction mixture using a column, or (2) a precipitate obtained by adding an appropriate solvent to the reaction mixture is obtained.
  • Compound [C-2] can be obtained by sampling and washing with a suitable solvent.
  • Compound [C-2] is a compound in which each nucleoside unit constituting the compound is a nucleoside unit [4a] and a nucleoside unit [4e], but all or one of the nucleoside unit [4a] or the nucleoside unit [4e]. Even if the part is a compound in which the nucleoside unit [4b], the nucleoside unit [4c], the nucleoside unit [4f] or the nucleoside unit [4g] is replaced, it can be produced by the same method as described above.
  • compound [C] has a solid-phase carrier in its molecule
  • compound [C] is packed in an appropriate column, and compound [C] is washed with an appropriate solvent to remove unnecessary substances.
  • examples of such a compound include compounds in which G is a substituent [7] and Z is a solid phase carrier in the compound [C-1] and the compound [C-2].
  • G is a silicon substituent in compound [C-1] and compound [C-2]
  • the target compound is isolated by performing an operation such as column purification using an appropriate solvent. -Can be purified.
  • n, p, BP , G, Q 1 , T, W and X are synonymous with the above.
  • compound [C-1] when G is a substituent [7] and Z is a solid phase carrier, for example, (1) compound [C-1] is packed in an appropriate column and contains an acid.
  • the elimination reaction can be carried out by eluting the solution or (2) shaking or stirring the solution containing compound [C-1] and an acid in a reaction vessel equipped with a filter.
  • the solvent that can be used in this elimination reaction is not particularly limited as long as it is a solvent usually used in the art, and the solvent may be used alone, or two or more solvents may be mixed and used. You may.
  • Examples of the solvent that can be used in this production method include aromatic solvents such as benzene, toluene, xylene, and mesitylene; ester solvents such as ethyl acetate and isopropyl acetate; hexane, pentane, heptane, octane, nonane, cyclohexane, and the like. Examples thereof include an aliphatic solvent and a halogen solvent. These solvents may be mixed and used.
  • Examples of the "acid” that can be used in this desorption reaction include trifluoroacetic acid, cyanopyridine trifluoroacetic acid, triethylamine trifluoroacetic acid, cyanoacetic acid, trichloroacetic acid, phosphoric acid, methanesulfonic acid, and p-toluenesulfone. Acids and hydrochloric acids can be mentioned. When these acids are used, they may be used in combination with a base (for example, triethylamine) to adjust the acidity.
  • a base for example, triethylamine
  • the amount of the acid that can be used in this elimination reaction for example, the amount in the range of 1 to 500 times the molar ratio is appropriate with respect to 1 mol of the compound [C-1], and the amount is 2 times.
  • the amount is preferably in the range of about 200 times.
  • the acid that can be used in this elimination reaction is appropriately diluted with a suitable solvent to a concentration in the range of, for example, 5% to 80%, and is in the range of 5% to 50%. It is preferable to dilute to the concentration of.
  • the solvent for dissolving the acid that can be used in this desorption reaction is not particularly limited, and is, for example, chloroform, dichloromethane, 1,1-dichloroethane, 1,2-dichloroethane, 1,1,2-trichloroethane, 1 , 2-Dichloroethylene, 2,2,2-trifluoroethanol or a mixed solvent thereof.
  • a scavenger may be used if necessary. Examples of "scavengers” that can be used in this elimination reaction include ethanol, triisopropylsilane, 1-hydroxybenzotriazole, pyrrole, indole, 2,2,2-trifluoroethanol, methanol, anisole, mercaptoethanol, and thio.
  • anisole can be mentioned.
  • the amount of the scavenger that can be used in this elimination reaction for example, the amount in the range of 1 to 100 times the molar ratio is appropriate with respect to 1 mol of the compound [C-1], and the amount is 1 time. The amount is preferably in the range of about 50 times.
  • G is substituted with (1) silicon substituent, (2) long chain alkyl-carbonyl, (3) 1-5 long chain alkyloxy and / or long chain alkenyloxy. If it is a benzoyl or (4) substituent [7] (except when Z is a solid phase carrier), for example, (1) compound [C-1] is placed in a suitable reaction vessel. [C-1] and the acid are stirred in an appropriate reaction solvent, or (2) the solution containing the compound [C-1] and the solution containing the acid are independently distributed through the supply channel. This desorption reaction can be carried out by supplying the solution to the reactor or the reaction flow path and mixing the solutions in the flow reactor or the like.
  • the solvent that can be used in this elimination reaction is not particularly limited as long as it is a solvent usually used in the art, and the solvent may be used alone, or two or more solvents may be mixed and used. You may.
  • the solvent that can be used in this production method include aromatic solvents such as benzene, toluene, xylene, and mesitylene; ester solvents such as ethyl acetate and isopropyl acetate; hexane, pentane, heptane, octane, nonane, cyclohexane, and the like. Examples thereof include an aliphatic solvent and a halogen solvent. These solvents may be mixed and used.
  • Examples of the "acid” that can be used in this elimination reaction include the same as described above. When these acids are used, they may be used in combination with a base (for example, triethylamine) to adjust the acidity.
  • a base for example, triethylamine
  • the amount of the acid that can be used in this elimination reaction for example, the amount in the range of 1 to 500 times the molar ratio is appropriate with respect to 1 mol of the compound [C-1], and the amount is 2 times. The amount is preferably in the range of about 200 times.
  • the acid that can be used in this elimination reaction is appropriately diluted with a suitable solvent to a concentration in the range of, for example, 5% to 80%, and is in the range of 5% to 50%. It is preferable to dilute to the concentration of.
  • the solvent for dissolving the acid that can be used in this desorption reaction is not particularly limited, and is, for example, chloroform, dichloromethane, 1,1-dichloroethane, 1,2-dichloroethane, 1,1,2-trichloroethane, 1 , 2-Dichloroethylene, 2,2,2-trifluoroethanol or a mixed solvent thereof.
  • a scavenger may be used if necessary. Examples of the "scavenger" that can be used in this elimination reaction include the same as described above.
  • the amount of the scavenger that can be used in this elimination reaction for example, the amount in the range of 1 to 100 times the molar ratio is appropriate with respect to 1 mol of the compound [C-1], and the amount is 1 time. The amount is preferably in the range of about 50 times.
  • a pump for supplying a liquid usually used in this field specifically, for example, a syringe pump, a plunger pump, a diaphragm pump, etc.
  • Gear pumps can be mentioned.
  • the flow reactor that can be used in this elimination reaction include in-line mixers such as a microreactor and a static mixer.
  • a multi-stage collision type micromixer As a means for guiding from the supply flow path to the reaction flow path that can be used in this elimination reaction, for example, a multi-stage collision type micromixer can be mentioned.
  • the materials of the supply channel and the reaction channel that can be used in this desorption reaction include fluororesins such as perfluoroalkoxy alkane (PFA), vinyl chloride resins, polyamide resins, and aromatic polyetherketones.
  • PFA perfluoroalkoxy alkane
  • vinyl chloride resins vinyl chloride resins
  • polyamide resins polyamide resins
  • aromatic polyetherketones aromatic polyetherketones
  • synthetic resin tubes selected from the group consisting of resins, or metal tubes selected from the group consisting of stainless steel, copper and its alloys and titanium and its alloys.
  • the inner diameters of the supply flow path and the reaction flow path that can be used in this elimination reaction may be appropriately selected from, for example, a size within the range of 0.1 mm to 1.0 mm, and for example, 0.2 mm to 1 It is preferable to select from a size within the range of 0.0 mm.
  • the solvent that can be used in this continuous reaction is not particularly limited as long as it is a solvent usually used in the art, and the solvent may be used alone, or two or more solvents may be mixed and used. You may.
  • the solvent that can be used in this production method include aromatic solvents such as benzene, toluene, xylene, and mesitylene; ester solvents such as ethyl acetate and isopropyl acetate; hexane, pentane, heptane, octane, nonane, cyclohexane, and the like. Examples thereof include an aliphatic solvent and a halogen solvent. These solvents may be mixed and used.
  • Equation [A-1-1] [During the ceremony, BP is a nucleobase that may be protected; Q 1 represents a leaving group capable under acidic conditions; W is a lone pair of electrons, an oxygen atom or a sulfur atom; X is a di (C 1-6 alkyl) amino or general formulas [2-1] to [2-8]: [In the formula, * represents the connection position with P] It is selected from the substituents represented by, preferably di (C 1-6 alkyl) amino, more preferably dimethyl amino; G is the general formula [7]: (During the ceremony, * Represents the connection position with T; Z is a general formula [8A] to [8D], [8E], [8G], [8H], [8J], [8K], [8N]: (During the ceremony, * Represents the connection position with L; k represents an integer from 0 to 5; R 8a represents a hydrogen atom or C
  • R 8d is the same or different, each of which is a hydrogen atom, a halogen, a long-chain alkyl optionally substituted with 1 to 13 halogens, or a long-chain alkyl optionally substituted with 1 to 13 halogens.
  • R 8e is (1) Long chain alkyl, Represents (2) long-chain alkyl-carbonyl or (3) benzoyl substituted with 1-5 long-chain alkyloxy and / or long-chain alkenyloxy; and
  • R 8f (1) Long chain alkyl, Represents (2) long-chain alkyl-carbonyl or (3) long-chain alkenyl-carbonyl.
  • L is the general formula [10]: (During the ceremony, * Represents the connection position with Z; ** represents the position of bond with the oxygen atom; and L 1 represents the optionally substituted C 2-10 alkylene or the optionally substituted C 6-10 arylene. ) It is a substituent represented by. ) It is a substituent represented by; T is a single bond or the following general formula [11]: (During the ceremony, X and W are synonymous with the above; * Represents the connection position with O; ** represents the position of connection with G; and q represents an integer from 0 to 10. ) It is a substituent represented by n is 1 to 25.
  • Equation [A-1] [In the formula, BP , W, X, G, T, and n are synonymous with the above. ]
  • Equation [B-1] [In the formula, BP , Q 1 , W, X, G, and T are synonymous with the above; D is a halogen, p is an integer from 1 to 10]
  • Equation [C-1] [In the formula, n, p, BP , Q 1 , W, X, G, and T are synonymous with the above. ] Examples thereof include a method for preparing the compound of.
  • the continuous reaction includes, for example, in a solvent usually used in the art. Equation [A-1-1]: [During the ceremony, Q 1 is trityl, monomethoxytrityl, or dimethoxytrityl, and n, BP , W, X, G, and T are synonymous with the above. ] From the compound of In the presence of trifluoroacetic acid and 2,2,2-trifluoroethanol, optionally triisopropylsilane or ethanol, It can include removing Q1. This continuous reaction can be carried out in a distribution reactor.
  • a solution containing the compound of the general formula [A-1-1] and a solution containing an acid were supplied to the flow reactor to remove Q1 to obtain a compound of the formula [A-1], and then A solution containing the compound of the general formula [A-1] and a solution containing the compound of the general formula [B-1] are supplied to the next flow reactor, and the compound of the general formula [C-1] is supplied.
  • There is a method of preparing In some cases, a distribution reactor that supplies a solution containing a compound of formula [A-1] and a solution containing a scavenger, or a surplus compound of formula [B-1] and a compound of formula [C-1] are contained.
  • a flow reactor can be used that supplies the solution to be used and a solution containing at least one selected from the group consisting of morpholine, 1-methylpiperazine, and N-ethylmorpholine.
  • the compounds [C-2] is an unstable compound, the compound [C-2], the Q 1 that is substituted with an oxygen atom of a 5 'nucleoside distal its 5' before leaving First, the phosphorus atom on the phosphorus bond formed by the condensation reaction is oxidized from trivalent to pentavalent using an oxidizing agent, and the compound represented by the following general formula [D-2] (hereinafter, "compound"). It is preferable to convert to [D-2] ”).
  • n, p, BP , G, Q 1 , R 4a , T, W and X are synonymous with the above.
  • Step 1 In the production compound [C-2] of the compound [D-2] , when G is a substituent [7] and Z is a solid phase carrier, the oxidation reaction of the phosphorus atom is carried out by a method known per se. It can be carried out in accordance with (Curent Protocols in Nuclear Acid Chemistry).
  • Examples of the oxidizing agent that can be used in this step include a commercially available oxidizing solution for nucleic acid synthesis [oxidizing solution-2, 0.1 mol / L iodine / 78% tetrahydrofuran / 20% pyridine / 2% water, Fujifilm Wako Jun.
  • oxidizing agent examples include commercially available sulfide reagents for nucleic acid synthesis [3- ⁇ (N, N-dimethylaminomethylidene) amino ⁇ ) -3H-1,2,4-dithiazole-5. -Thion (DDTT), manufactured by Glen Research Co., Ltd .; 5-phenyl-3H-1,2,4-dithiazole-3-one for nucleic acid synthesis, manufactured by Fujifilm Wako Pure Chemical Industries, Ltd.].
  • DDTT sulfide reagents for nucleic acid synthesis
  • DDTT DDTT
  • 5-phenyl-3H-1,2,4-dithiazole-3-one for nucleic acid synthesis, manufactured by Fujifilm Wako Pure Chemical Industries, Ltd.
  • G is substituted with (1) silicon substituent, (2) long chain alkyl-carbonyl, (3) 1-5 long chain alkyloxy and / or long chain alkenyloxy.
  • substituent [7] except when Z is a solid phase carrier, the oxidation reaction of the phosphorus atom can be carried out according to a method known per se (excluding the case where Z is a solid phase carrier). See, for example, Nucleic Acids Research, Vol. 21, No. 5, 1231-1217 (1993)).
  • Examples of the oxidizing agent that can be used in this step include (+)-camforylsulfonyl oxaziridine (CSO) (+)-(8,8-dichlorocamphorylsulfonyl) -oxadylidine (DCSO) and methyl ethyl ketone peroxide. Oxides such as tert-butyl hydroperoxide (TBHP) can be mentioned.
  • Step 2 Production of compound [E-2] By reacting compound [D-2] with an acid, it is replaced with the oxygen atom at the 5'position of the nucleoside unit on the 5'terminal side of compound [D-2]. Q 1 can be detached.
  • Compounds from [D-2] produced by desorbing a to Q 1 in the molecule, the compound represented by the general formula [E-2] (hereinafter. Referred to as "Compound [E-2]" can do.
  • G is a substituent [7] and Z is a solid phase carrier in compound [D-2]
  • compound [D-2] is packed in an appropriate column to elute a solution containing an acid.
  • it can be carried out by shaking or stirring a solution containing compound [D-2] and an acid in a reaction vessel with a filter.
  • Elimination reaction for Q 1 in the molecule of the compound [D-2] can be carried out according to methods known per se (see, for example, Current Protocols in Nucleic Acid Chemistry.) .
  • Examples of the acid that can be used in this step include a commercially available deblocking solution for nucleic acid synthesis [for example, deblocking solution-1,3 w / v% trichloroacetic acid / dichloromethane solution (manufactured by Fujifilm Wako Pure Chemical Industries, Ltd.). Deblocking Mix 3% dichloroacetic acid / dichloromethane solution (manufactured by Glen Research Co., Ltd.)] can be mentioned.
  • deblocking solution-1,3 w / v% trichloroacetic acid / dichloromethane solution manufactured by Fujifilm Wako Pure Chemical Industries, Ltd.
  • Deblocking Mix 3% dichloroacetic acid / dichloromethane solution manufactured by Glen Research Co., Ltd.
  • G is substituted with (1) silicon substituent, (2) long chain alkyl-carbonyl, (3) 1-5 long chain alkyloxy and / or long chain alkenyloxy. If it is a benzoyl or (4) substituent [7] (except when Z is a solid phase carrier), for example, (1) compound [D-2] is placed in a suitable reaction vessel. [D-2] and the acid are stirred in an appropriate reaction solvent, or (2) the solution containing the compound [D-2] and the solution containing the acid are independently distributed through the supply flow path. It can be carried out by supplying the solution to the reactor or the reaction channel and mixing the solutions in the flow reactor or the like.
  • Elimination reaction for Q 1 in the molecule of the compound [D-2] can be carried out according to methods known per se (e.g., Nucleic Acids Research, Vol.21, No.5,1213-1217 (1993 ).).
  • Examples of the acid that can be used in this step include dichloroacetic acid and trichloroacetic acid.
  • the "compound, [C-1] the desorption process for Q 1 in the molecule of” the same as the “acid” described in desorption for Q 1 in the molecule of the "compound [C-2]
  • the compound [C-1] can be obtained by treating the compound [C-1] with the same "acid” as described in "Step 2 Production of the compound [E-2]” of "Method” or by treating with a solution obtained by diluting hydrochloric acid or acetic acid with an appropriate solvent.
  • a group that can be eliminated under acidic conditions which is substituted with a protective group for the amino group at the 3'position of the 3'-terminal nucleoside and a hydroxyl group at the 5'position of the 5'-terminal nucleoside of compound [D-2]. Can be removed.
  • the group that can be eliminated under acidic conditions which is substituted with the hydroxyl group at the 5'position of the nucleoside on the 5'terminal side of compound [D-2] after removing the protective group of the nucleic acid base portion.
  • a mixed solution of, for example, 20 mM triethylamine / acetate buffer and acetonitrile can be used as the elution solvent.
  • a mixed solution of 1 M saline solution and 10 mM sodium hydroxide aqueous solution or 0.3 M saline solution of 50 mM phosphate buffer is used. Can be used.
  • the compound [D-2] or the compound from which all the protective groups contained in the compound [E-2] have been removed can be obtained from the above reaction mixture by conventional separation and purification means, for example, extraction, concentration, neutralization, filtration, centrifugation, Recrystallization, C 8 to C 18 reverse phase column chromatography, C 8 to C 18 reverse phase cartridge column, cation exchange column chromatography, anion exchange column chromatography, gel filtration column chromatography, high speed liquid chromatography, It can be isolated and purified by using means such as dialysis and marginal filtration alone or in combination.
  • separation and purification means for example, extraction, concentration, neutralization, filtration, centrifugation, Recrystallization, C 8 to C 18 reverse phase column chromatography, C 8 to C 18 reverse phase cartridge column, cation exchange column chromatography, anion exchange column chromatography, gel filtration column chromatography, high speed liquid chromatography, It can be isolated and purified by using means such as dialysis and marginal filtration alone or in combination.
  • the “eluting solvent” examples include a single solvent of acetonitrile, methanol, ethanol, isopropyl alcohol or water, or a mixed solvent of any ratio of these solvents.
  • sodium phosphate, potassium phosphate, sodium chloride, potassium chloride, ammonium acetate, triethylammonium acetate, sodium acetate, potassium acetate, tris-hydrochloric acid or ethylenediamine tetraacetic acid are added as additives at a concentration of 1 mM to 2 M.
  • the pH of the solution can also be adjusted in the range of 1-9.
  • Step 1 Production of a compound represented by the above general formula [A-1a-Q1] (hereinafter, referred to as “compound [A-1a-Q1]”)
  • a compound represented by the above general formula [21] hereinafter, ""
  • a compound represented by the above general formula [20A] hereinafter referred to as “Compound [20A]”
  • a silicon substituent is attached to the hydroxyl group at the 5'end of the compound [21].
  • This silicon substituent introduction reaction can be carried out according to a method known per se.
  • Step 2 Production of Compound [A-1a]
  • Compound [A-1a] can be produced by treating compound [A-1a-Q1] with an acid.
  • the "acid” that can be used in this step include the same "acid” described in the above-mentioned “Method for desorbing Q 1 in the molecule of compound [C-1]".
  • the amount of the acid that can be used in this step for example, the amount in the range of 1 to 500 times the molar ratio is appropriate with respect to 1 mol of the compound [A-1a-Q1], and the amount is 2 times. The amount is preferably in the range of about 200 times.
  • the acid that can be used in this step may be diluted with an appropriate solvent and is not particularly limited.
  • chloroform for example, chloroform, dichloromethane, 1,1-dichloroethane, 1,2-dichloroethane, 1,1,2-trichloroethane, 1 , 2-Dichloroethylene, 2,2,2-trifluoroethanol or a mixed solvent thereof.
  • a scavenger may be used if necessary.
  • “Scavenger” which can be used in this step may include the same as the “scavenger", as described in the "compound desorption method for Q 1 in the molecule [C-1]".
  • the amount of the scavenger that can be used in this step for example, the amount in the range of 1 to 100 times the molar ratio is appropriate with respect to 1 mol of the compound [A-1a-Q1], and the amount is 1 time. The amount is preferably in the range of about 50 times.
  • the compound represented by the general formula [A-1b] (hereinafter, referred to as “compound [A-1b]”) has a G of (1) long-chain alkyl-carbonyl and (2) 1 to 5 G.
  • Compound [A-1] which is a benzoyl or (3) substituent [7] substituted with a long-chain alkyloxy and / or a long-chain alkenyloxy, and T is a single bond.
  • An example of a method for producing the compound [A-1b] will be described below.
  • Step 1 Production of a compound represented by the above general formula [A-1b-Q1] (hereinafter, referred to as "compound [A-1b-Q1]”)
  • the compound [21] is represented by the above general formula [20B].
  • the compound [A-1b-Q1] can be produced by condensing with a compound (hereinafter, referred to as "compound [20B]”).
  • the condensation reaction can be carried out according to a method known per se.
  • the compound [20B] in which Y is a hydroxyl group is used in this step, it can be carried out in the range of ⁇ 20 ° C. to 100 ° C. using a condensing agent in the presence or absence of a base.
  • the compound [20B] in which Y is halogen when used in this step, it can be carried out in the range of ⁇ 20 ° C. to 100 ° C. in the presence of a base.
  • the condensing agent that can be used in this step include 1,1'-oxalyldiimidazole, 1-ethyl-3- (3-dimethylaminopropyl) carbodiimide, dicyclohexylcarbodiimide, diethyl cyanophosphonate, and O- (benzotriazole).
  • bases that can be used in this step include organic bases of triethylamine, N, N-diisopropylethylamine, N, N-dimethylaniline, pyridine, and 1,8-diazabicyclo [5,4,0] -7-undecene.
  • the solvent that can be used in this step is not particularly limited, but is, for example, ethers such as THF, 1,4-dioxane and diethyl ether, amides such as dimethylformamide and dimethylacetamide, and nitriles such as acetonitrile and propionitrile.
  • Examples include hydrocarbons such as benzene and toluene, halogenated hydrocarbons such as chloroform and methylene chloride, and mixed solvents thereof.
  • an additive can be used if necessary.
  • Examples of the additive that can be used in this step include 4-dimethylaminopyridine, 1-hydroxybenzotriazole, and 1-hydroxy-7-azabenzotriazole.
  • the reaction time varies depending on the type of raw material used, the reaction temperature, etc., but is usually in the range of 10 minutes to 24 hours.
  • the appropriate amount of the compound [21] and the condensing agent to be used is, for example, in the range of 1 to 1.5 times the mole with respect to 1 mol of the compound [20B].
  • the amount of the base used is, for example, in the range of 1 equivalent to 10 equivalents, preferably in the range of 1 equivalent to 4 equivalents, relative to compound [20B].
  • Step 2 Production of Compound [A-1b]
  • Compound [A-1b] can be produced by treating compound [A-1b-Q1] with an acid.
  • Examples of the “acid” that can be used in this step include the same “acid” described in the above-mentioned “Method for desorbing Q 1 in the molecule of compound [C-1]".
  • the amount of the acid that can be used in this step for example, the amount in the range of 1 to 500 times the molar ratio is appropriate with respect to 1 mol of the compound [A-1b-Q1], and the amount is 2 times. The amount is preferably in the range of about 200 times.
  • the acid that can be used in this step may be diluted with an appropriate solvent and is not particularly limited.
  • chloroform for example, chloroform, dichloromethane, 1,1-dichloroethane, 1,2-dichloroethane, 1,1,2-trichloroethane, 1 , 2-Dichloroethylene, 2,2,2-trifluoroethanol or a mixed solvent thereof.
  • a scavenger may be used if necessary.
  • “Scavenger” which can be used in this step may include the same as the “scavenger", as described in the "compound desorption method for Q 1 in the molecule [C-1]".
  • the amount of the scavenger that can be used in this step for example, the amount in the range of 1 to 100 times the molar ratio is appropriate with respect to 1 mol of the compound [A-1b-Q1], and the amount is 1 time. The amount is preferably in the range of about 50 times.
  • the compound represented by the general formula [A-1c] (hereinafter, referred to as “compound [A-1c]”) has a G of (1) long-chain alkyl-carbonyl and (2) 1 to 5 G.
  • Compound [A-1] which is a benzoyl or (3) substituent [7] substituted with a long-chain alkyloxy and / or a long-chain alkenyloxy, wherein T is a substituent [11].
  • An example of a method for producing the compound [A-1c] will be described below.
  • Step 1 Production of a compound represented by the above general formula [23] (hereinafter, referred to as “Compound [23]”)
  • a compound represented by the above general formula [20C] (hereinafter, referred to as “Compound [20C]”).
  • the compound [23] can be produced by condensing the compound represented by the above general formula [22] (hereinafter, referred to as “compound [22]”).
  • compound [20C] is a carboxylic acid compound
  • a reactive derivative thereof can also be used in this step.
  • Examples of the reactive derivative of the compound [20C] include those usually used in an ester condensation formation reaction such as an acid halide (for example, acid chloride and acid bromide).
  • Compound [22] can be produced according to a known method (see, for example, US Patent Application Publication No. 2014/0330006A1). Also, according to a known method (see, for example, International Publication No. 2014/077292A1), a benzoyl in which G is substituted with 1 to 5 long-chain alkyloxy and / or long-chain alkenyloxy. A compound [20C] can be produced.
  • Step 2 Production of compound represented by the above general formula [24] (hereinafter referred to as “compound [24]”)
  • Compound [24] is prepared by removing the trityl group in the molecule with an acid. Can be manufactured.
  • Step 3 Production of a compound represented by the above general formula [A-1c-Q1] (hereinafter, referred to as "compound [A-1c-Q1]”) Represented by the compound [24] and the above general formula [25].
  • Compound [A-1c-Q1] can be produced by condensing a compound (hereinafter referred to as “compound [25]”). The condensation reaction and deprotection reaction can be carried out according to a method known per se.
  • Step 4 Production of Compound [A-1c]
  • Compound [A-1c] can be produced by treating compound [A-1c-Q1] with an acid.
  • the "acid” that can be used in this step include the same "acid” described in the above-mentioned “Method for desorbing Q 1 in the molecule of compound [C-1]".
  • the amount of the acid that can be used in this step for example, the amount in the range of 1 to 500 times the molar ratio is appropriate with respect to 1 mol of the compound [A-1c-Q1], and the amount is 2 times. The amount is preferably in the range of about 200 times.
  • the acid that can be used in this step may be diluted with an appropriate solvent and is not particularly limited.
  • chloroform for example, chloroform, dichloromethane, 1,1-dichloroethane, 1,2-dichloroethane, 1,1,2-trichloroethane, 1 , 2-Dichloroethylene, 2,2,2-trifluoroethanol or a mixed solvent thereof.
  • a scavenger may be used if necessary.
  • “Scavenger” which can be used in this step may include the same as the “scavenger", as described in the "compound desorption method for Q 1 in the molecule [C-1]".
  • the amount of the scavenger that can be used in this step for example, the amount in the range of 1 to 100 times the molar ratio is appropriate with respect to 1 mol of the compound [A-1c-Q1], and the amount is 1 time. The amount is preferably in the range of about 50 times.
  • compound [A-1d] The compound represented by the general formula [A-1d] (hereinafter, referred to as “compound [A-1d]”) has a compound [A] in which G is a substituent [7] and T is a single bond. -1].
  • An example of a method for producing the compound [A-1d] will be described below.
  • Step 1 Production of a compound represented by the above general formula [A-1d-Q1] (hereinafter, referred to as "Compound [A-1d-Q1]”)
  • a compound represented by the above general formula [20D] (hereinafter, "" Compound [20D] ”) is referred to as a compound represented by the above general formula [21] (hereinafter, compound [21]).
  • the compound [A-1d-Q1] can be produced.
  • the condensation reaction can be carried out according to a method known per se.
  • compound [20D] is a carboxylic acid compound, a reactive derivative thereof can also be used in this step.
  • Examples of the reactive derivative of the compound [20D] include those usually used in an ester condensation formation reaction such as an acid halide (for example, acid chloride and acid bromide).
  • an acid halide for example, acid chloride and acid bromide
  • the reaction can be carried out in the range of ⁇ 20 ° C. to 100 ° C. using a condensing agent in the presence or absence of a base.
  • the condensing agent that can be used in this step include 1,1'-oxalyldiimidazole, 1-ethyl-3- (3-dimethylaminopropyl) carbodiimide, dicyclohexylcarbodiimide, diethyl cyanophosphonate, and O- (benzotriazole).
  • bases that can be used in this step include organic bases such as triethylamine, N, N-diisopropylethylamine, N, N-dimethylaniline, pyridine, and 1,8-diazabicyclo [5,4,0] -7-undecene.
  • the solvent that can be used in this step is not particularly limited, but is, for example, ethers such as THF, 1,4-dioxane and diethyl ether, amides such as dimethylformamide and dimethylacetamide, and nitriles such as acetonitrile and propionitrile.
  • additives can be used if necessary. Examples of the additive that can be used in this step include 4-dimethylaminopyridine, 1-hydroxybenzotriazole, and 1-hydroxy-7-azabenzotriazole.
  • the reaction time varies depending on the type of raw material used, the reaction temperature, etc., but is usually in the range of 10 minutes to 24 hours.
  • the appropriate amount of the compound [21] and the condensing agent to be used is, for example, in the range of 1-fold to 1.5-fold mol with respect to 1 mol of the compound [20D].
  • the amount of the base used is, for example, in the range of 1 equivalent to 10 equivalents, preferably in the range of 1 equivalent to 4 equivalents, relative to compound [20D].
  • Step 2 Production of compound [A-1d]
  • Compound [A-1d] can be produced by acid-treating compound [A-1d-Q1].
  • Examples of the "acid” that can be used in this step include the same “acid” described in the above-mentioned “Method for desorbing Q 1 in the molecule of compound [C-1]".
  • the amount of the acid that can be used in this step for example, the amount in the range of 1 to 500 times the molar ratio is appropriate with respect to 1 mol of the compound [A-1d-Q1], and the amount is 2 times.
  • the amount is preferably in the range of about 200 times.
  • the acid that can be used in this step may be diluted with an appropriate solvent and is not particularly limited.
  • scavenger which can be used in this step may include the same as the “scavenger”, as described in the "compound desorption method for Q 1 in the molecule [C-1]".
  • the amount of the scavenger that can be used in this step for example, the amount in the range of 1 to 100 times the molar ratio is appropriate with respect to 1 mol of the compound [A-1d-Q1], and the amount is 1 time.
  • the amount is preferably in the range of about 50 times.
  • compound [20D] can be produced according to the production method described below. [During the ceremony, L 1 and Z are defined as above; R represents C 1-6 alkyl. ]
  • Step 1 Production of a compound represented by the above general formula [28] (hereinafter, referred to as “Compound [28]”)
  • a compound represented by the above general formula [26] (hereinafter, referred to as “Compound [26]”).
  • Compound [27] can be produced by condensing compound [28] with a compound represented by the above general formula [27] (hereinafter, referred to as “compound [27]”).
  • the condensation reaction can be carried out according to a method known per se.
  • the same reagents as in the above-mentioned "Production of compound [A-1b-Q1]" can be used.
  • Step 2 Production of compound [20D]
  • Compound [20D] can be produced by ester-hydrolyzing compound [28].
  • the ester hydrolysis reaction can be carried out according to a method known per se.
  • the solvent that can be used in this step is not particularly limited, but is, for example, water, alcohols such as methanol and ethanol, ethers such as tetrahydrofuran, 1,4-dioxane and diethyl ether, and nitriles such as acetonitrile and propionitrile.
  • Hydrocarbons such as benzene and toluene, halogenated hydrocarbons such as chloroform and methylene chloride, or a mixed solvent thereof. This step is carried out in the range of 20 ° C.
  • reaction time varies depending on the type of raw material used, the reaction temperature, etc., but is usually in the range of 10 minutes to 24 hours.
  • compound [26] can be produced, for example, according to the methods described in (a) to (j) below.
  • A For example, by using a commercially available primary amine compound or by amifying a commercially available alkyl halide, Z is a substituent [8A] and R 8a.
  • Compound [26] can be prepared in which is a hydrogen atom and R 8b is a long-chain alkyl.
  • B For example, by alkylating a commercially available primary amine compound, Z is a substituent [8A] or a substituent [8B] and R 8a is a C 1-6 alkyl.
  • Compound [26] can be prepared in which R 8b is the same or different, respectively, and is a long-chain alkyl.
  • the alkylation reaction can be carried out according to a method known per se.
  • Z is a substituent according to a known method (see, for example, Cancer Res., 2008 Nov 1; 68 (21): 8843-8851, Chem. Sci., 2016, 7, 2308-2321).
  • Compound [26], which is [8C] can be produced.
  • D For example, a compound in which methyl phthalate is condensed with 1- (tert-butoxycarbonyl) piperazine, then the ester moiety is hydrolyzed with an alkali such as sodium hydroxide, and Z is a substituent [8A].
  • the compound [26] in which Z is a substituent [8D] can be produced by removing the tert-butoxycarbonyl group with an acid such as trifluoroacetic acid.
  • the condensation reaction, the hydrolysis reaction using an alkali, and the deprotection reaction of the tert-butoxycarbonyl group using an acid can be carried out according to a method known per se.
  • E For example, by alkylating one hydroxyl group of ethane-1,2-diol with an alkyl halide, Z is a substituent [8E] and R 8e is a long-chain alkyl group. Certain compounds [26] can be produced.
  • compound [26] in which one hydroxyl group of ethane-1,2-diol is long-chain alkyl-carbonylated so that Z is a substituent [8E] and R 8e is a long-chain alkyl-carbonyl. ] can be manufactured.
  • the compound used for long-chain alkyl-carbonylation for example, the corresponding carboxylic acid compound or a reactive derivative thereof can be used.
  • the reactive derivative include those usually used in an ester condensation formation reaction, such as an acid halide (for example, acid chloride and acid bromide).
  • Z is a substituent [8E] and R 8e is 1 to 5 long-chain alkyl.
  • Compound [26] which is a benzoyl group substituted with oxy and / or long chain alkenyloxy, can be prepared.
  • F For example, 2-amino-ethanol is used instead of ethane-1,2-diol, and Z is substituted according to the same method as that for producing the compound (18) in which Z is a substituent [8E].
  • the compound [26] which is a group [8F] can be produced.
  • (G) For example, 9-fluorenylmethyloxycarbonyl-phenylalanine is condensed with compound [26] in which Z is a substituent [8A], and then the 9-fluorenylmethyloxycarbonyl group is eliminated with piperidine. Thereby, the compound [26] in which Z is a substituent [8G] can be produced.
  • the condensation reaction and the deprotection reaction of the 9-fluorenylmethyloxycarbonyl group can be carried out according to a method known per se.
  • (H) For example, using 1- (tert-butoxycarbonyl) piperazine instead of ethane-1,2-diol, according to the same method as the method for producing the compound [26] in which Z is the substituent [8E].
  • Step 1 In the presence of a production base for the compound represented by the general formula [30] (hereinafter referred to as “compound [30]”), the compound represented by the general formula [29] (for example, International Publication No. 2). 91/09033A1) can be acetylated with acetic anhydride to produce compound [30].
  • the acetylation reaction can be carried out according to a method known per se.
  • Step 2 Production of the compound represented by the general formula [31] (hereinafter, referred to as “compound [31]”)
  • the compound [31] can be produced by acid-treating the compound [30]. Elimination reaction of Q 1 is can be performed according to methods known per se.
  • Step 3 The production compound [31] of the compound represented by the general formula [33] (hereinafter, referred to as “compound [33]”) is replaced with the compound represented by the general formula [32] (hereinafter, “Compound [32]”). ] ”),
  • the compound [33] can be produced.
  • the condensation reaction can be carried out according to a method known per se (see, for example, International Publication No. 91/09033A1).
  • Compound [32] can be produced, for example, according to a known method (see, for example, International Publication No. 91/09033A1).
  • Step 4 Production of Compound [21]
  • Compound [21] can be produced by selectively removing the acetyl group of compound [33] using, for example, an alkali metal alkoxide such as sodium methoxide. ..
  • the elimination reaction of acetyl can be carried out according to a method known per se (see, for example, Tetrahedron Letters, Vol. 50, 1751-1753 (2009)).
  • Step 1 Production of a compound represented by the above general formula [A-2a-Q1] (hereinafter referred to as “compound [A-2a-Q1]”)
  • a compound represented by the above general formula [34] (hereinafter referred to as “”
  • Compound [34] ” is referred to as compound [20A]
  • a silicon substituent is introduced into the hydroxyl group at the 3'position of the nucleoside unit on the 3'-terminal side of compound [34] to introduce compound [A-2a].
  • -Q1] can be manufactured.
  • This silicon substituent introduction reaction can be carried out according to a method known per se.
  • Step 2 Production of compound [A-2a]
  • Compound [A-2a] can be produced by acid-treating compound [A-2a-Q1].
  • “Acid” which can be used in this step may include the same as the “acid” described in the "elimination reaction for Q 1 in the molecule of the compound [D-2]".
  • the amount of the acid that can be used in this step for example, the amount in the range of 1 to 500 times the molar ratio is appropriate with respect to 1 mol of the compound [A-2a-Q1], and the amount is 2 times. The amount is preferably in the range of about 200 times.
  • the acid that can be used in this step may be diluted with an appropriate solvent and is not particularly limited.
  • chloroform for example, chloroform, dichloromethane, 1,1-dichloroethane, 1,2-dichloroethane, 1,1,2-trichloroethane, 1 , 2-Dichloroethylene, 2,2,2-trifluoroethanol or a mixed solvent thereof.
  • a scavenger may be used if necessary.
  • "Scavenger” which can be used in this step may include the same as the “scavenger", as described in the "elimination reaction for Q 1 in the molecule of the compound [D-2]".
  • the amount of the scavenger that can be used in this step for example, the amount in the range of 1 to 100 times the molar ratio is appropriate with respect to 1 mol of the compound [A-2a-Q1], and the amount is 1 time. The amount is preferably in the range of about 50 times.
  • the compound represented by the general formula [A-2b] (hereinafter, referred to as “compound [A-2b]”) has a G of (1) long-chain alkyl-carbonyl and (2) 1 to 5 G.
  • Step 1 Production of a compound represented by the above general formula [A-2b-Q1] (hereinafter referred to as “compound [A-2b-Q1]”) By condensing the compound [20B] with the compound [34].
  • Compound [A-2b-Q1] can be produced.
  • the condensation reaction can be carried out according to a method known per se.
  • the compound [20B] in which Y is a hydroxyl group is used in this step, it can be carried out in the range of ⁇ 20 ° C. to 100 ° C. using a condensing agent in the presence or absence of a base.
  • the compound [20B] in which Y is halogen is used in this step, it can be carried out in the range of ⁇ 20 ° C.
  • condensing agent examples include 1,1'-oxalyldiimidazole, 1-ethyl-3- (3-dimethylaminopropyl) carbodiimide, dicyclohexylcarbodiimide, diethyl cyanophosphonate, and O- (benzotriazole).
  • bases that can be used in this step include organic bases of triethylamine, N, N-diisopropylethylamine, N, N-dimethylaniline, pyridine, and 1,8-diazabicyclo [5,4,0] -7-undecene.
  • the solvent that can be used in this step is not particularly limited, but is, for example, ethers such as THF, 1,4-dioxane and diethyl ether, amides such as dimethylformamide and dimethylacetamide, and nitriles such as acetonitrile and propionitrile.
  • Examples include hydrocarbons such as benzene and toluene, halogenated hydrocarbons such as chloroform and methylene chloride, and mixed solvents thereof. Further, when the compound [20B] in which Y is a hydroxyl group is used in this step, an additive can be used if necessary. Examples of additives that can be used include 4-dimethylaminopyridine, 1-hydroxybenzotriazole, and 1-hydroxy-7-azabenzotriazole.
  • the reaction time varies depending on the type of raw material used, the reaction temperature, etc., but is usually in the range of 10 minutes to 24 hours.
  • the appropriate amount of the compound [20B] and the condensing agent to be used is, for example, in the range of 1-fold to 1.5-fold mol with respect to 1 mol of the compound [34].
  • the amount of the base used is, for example, in the range of 1 equivalent to 10 equivalents, preferably in the range of 1 equivalent to 4 equivalents, relative to compound [34].
  • Step 2 Production of compound [A-2b]
  • Compound [A-2b] can be produced by acid-treating compound [A-2b-Q1].
  • “Acid” which can be used in this step may include the same as the “acid” described in the "elimination reaction for Q 1 in the molecule of the compound [D-2]".
  • the amount of the acid that can be used in this step for example, the amount in the range of 1 to 500 times the molar ratio is appropriate with respect to 1 mol of the compound [A-2b-Q1], and the amount is 2 times. The amount is preferably in the range of about 200 times.
  • the acid that can be used in this step may be diluted with an appropriate solvent and is not particularly limited.
  • chloroform for example, chloroform, dichloromethane, 1,1-dichloroethane, 1,2-dichloroethane, 1,1,2-trichloroethane, 1 , 2-Dichloroethylene, 2,2,2-trifluoroethanol or a mixed solvent thereof.
  • a scavenger may be used if necessary.
  • "Scavenger” which can be used in this step may include the same as the “scavenger", as described in the "elimination reaction for Q 1 in the molecule of the compound [D-2]".
  • the amount of the scavenger that can be used in this step for example, the amount in the range of 1 to 100 times the molar ratio is appropriate with respect to 1 mol of the compound [A-2b-Q1], and the amount is 1 time. The amount is preferably in the range of about 50 times.
  • the compound represented by the general formula [A-2c] (hereinafter, referred to as “compound [A-2c]”) has a G of (1) long-chain alkyl-carbonyl and (2) 1 to 5 G.
  • Compound [A-2] which is a benzoyl or (3) substituent [7] substituted with a long-chain alkyloxy and / or a long-chain alkenyloxy, and T is a substituent [11].
  • Step 1 Production of the compound represented by the general formula [36] (hereinafter, referred to as “compound [36]”)
  • the compound [36] can be produced.
  • the condensation reaction and deprotection reaction can be carried out according to a method known per se.
  • Step 2 Production of a compound represented by the above general formula [A-2c-Q1] (hereinafter referred to as "compound [A-2c-Q1]”)
  • compound [A-2c-Q1] By allowing an oxidizing agent to act on the compound [36], the compound [ A-2c-Q1] can be produced.
  • the oxidation reaction can be carried out according to a method known per se.
  • the "oxidizing agent” include iodine and tert-butyl hydroperoxide.
  • the oxidizing agent that can be used in this step can be diluted with an appropriate solvent so as to have a concentration of 0.05 to 2M.
  • the solvent is not particularly limited, and examples thereof include pyridine, tetrahydrofuran, water, and a mixed solvent thereof.
  • iodine / water / pyridine-tetrahydrofuran, iodine / pyridine-acetic acid, or a peroxide agent can be used.
  • the reaction temperature is preferably 20 ° C to 50 ° C.
  • the reaction time varies depending on the type of oxidizing agent used and the reaction temperature, but is usually 1 to 30 minutes.
  • the amount of the oxidizing agent used is preferably 1 to 100 times the molar amount, more preferably 10 to 50 times the molar amount of the compound [36].
  • Step 3 Production of compound [A-2c]
  • Compound [A-2c] can be produced by acid-treating compound [A-2c-Q1].
  • “Acid” which can be used in this step may include the same as the “acid” described in the "elimination reaction for Q 1 in the molecule of the compound [D-2]".
  • the amount of the acid that can be used in this step for example, the amount in the range of 1 to 500 times the molar ratio is appropriate with respect to 1 mol of the compound [A-2c-Q1], and the amount is 2 times. The amount is preferably in the range of about 200 times.
  • the acid that can be used in this step may be diluted with an appropriate solvent and is not particularly limited.
  • scavenger which can be used in this step may include the same as the “scavenger", as described in the "elimination reaction for Q 1 in the molecule of the compound [D-2]".
  • the amount of the scavenger that can be used in this step for example, the amount in the range of 1 to 100 times the molar ratio is appropriate with respect to 1 mol of the compound [A-2c-Q1], and the amount is 1 time.
  • the amount is preferably in the range of about 50 times.
  • condensing agent in the presence or absence of a base.
  • condensing agent examples include 1,1'-oxalyldiimidazole, 1-ethyl-3- (3-dimethylaminopropyl) carbodiimide, dicyclohexylcarbodiimide, diethyl cyanophosphonate, and O- (benzotriazole).
  • bases that can be used in this step include organic bases of triethylamine, N, N-diisopropylethylamine, N, N-dimethylaniline, pyridine, and 1,8-diazabicyclo [5,4,0] -7-undecene.
  • the solvent that can be used in this step is not particularly limited, but is, for example, ethers such as THF, 1,4-dioxane and diethyl ether, amides such as dimethylformamide and dimethylacetamide, and nitriles such as acetonitrile and propionitrile.
  • additives can be used if necessary. Examples of the additive that can be used in this step include 4-dimethylaminopyridine, 1-hydroxybenzotriazole, and 1-hydroxy-7-azabenzotriazole.
  • the reaction time varies depending on the type of raw material used, the reaction temperature, etc., but is usually in the range of 10 minutes to 24 hours.
  • the appropriate amount of the compound [20D] and the condensing agent to be used is, for example, in the range of 1-fold to 1.5-fold mol with respect to 1 mol of the compound [34].
  • the amount of the base used is, for example, in the range of 1 equivalent to 10 equivalents, preferably in the range of 1 equivalent to 4 equivalents, relative to compound [34].
  • Step 2 The compound [A-2d] can be produced by treating the compound [A-2d-Q1] with an acid.
  • “Acid” which can be used in this step may include the same as the “acid” described in the "elimination reaction for Q 1 in the molecule of the compound [D-2]".
  • the amount of the acid that can be used in this step for example, the amount in the range of 1 to 500 times the molar ratio is appropriate with respect to 1 mol of the compound [A-2d-Q1], and the amount is 2 times. The amount is preferably in the range of about 200 times.
  • the acid that can be used in this step may be diluted with an appropriate solvent and is not particularly limited.
  • scavenger which can be used in this step may include the same as the “scavenger", as described in the "elimination reaction for Q 1 in the molecule of the compound [D-2]".
  • the amount of the scavenger that can be used in this step for example, the amount in the range of 1 to 100 times the molar ratio is appropriate with respect to 1 mol of the compound [A-2d-Q1], and the amount is 1 time.
  • the amount is preferably in the range of about 50 times.
  • Compound “A-2” is a compound in which each nucleoside unit constituting the compound is a nucleoside unit [4a], but all or part of the nucleoside unit [4a] is a nucleoside unit [4b] or a nucleoside unit [4c]. ] Can also be produced by using the same method as described above.
  • Step 1 Production of a compound represented by the above general formula [39] (hereinafter referred to as “Compound [39]”)
  • a compound represented by the above general formula [37] (hereinafter referred to as “Compound [37]”).
  • Compound [38] Can be produced by condensing compound [39] with a compound represented by the above general formula [38] (hereinafter, referred to as “compound [38]”).
  • the condensation reaction can be carried out according to a method known per se (see, for example, US Patent Application Publication No. 2014/0330006A1, International Publication No. 2012/0437330A1, International Publication No. 2013/082548A1).
  • Step 2 Production of Compound [B-1]
  • Compound [B-1] is produced by condensing a compound represented by the above general formula [40] (hereinafter referred to as “Compound [40]”) with Compound [39].
  • the condensation reaction is carried out by a method known per se (see, for example, US Patent Application Publication No. 2014/0330006A1, International Publication No. 2012/0437330A1, International Publication No. 2013/082548A1, International Publication No. 91/09033A1). It can be carried out according to the above.
  • the compound [40] can be produced by using the same method as the method for producing the compound [21].
  • Step 1 Production of the compound [37] produced by the compound represented by the general formula [42] (hereinafter, referred to as “compound [42]”)
  • the compound [37] represented by the general formula [41] (hereinafter, “Compound [41]”). ] ”)
  • the compound [42] can be produced.
  • the reaction is carried out by a method known per se (eg, Helvetica Chemica Acta, Vol. 70, 175-186 (1987), WO 2003/106468A1, Acta Nature, 6,116-118 (2014), Russian Journal of General. It can be carried out according to Chemistry, Vol. 67, No. 1, 62-64 (1997)).
  • Step 2 Production of compound [B-2]
  • compound [42] is allowed to act on a compound represented by the above general formula [43] (hereinafter, referred to as “compound [43]”) at the 3'end.
  • Compound “B-2] can be carried out by introducing a substituent containing a phosphorus atom into the hydroxyl group at the 3'position of the nucleoside unit on the side. If necessary, an activator can also be used in this step.
  • the solvent used in this step is not particularly limited, and examples thereof include acetonitrile and tetrahydrofuran.
  • the amount of the compound [42] to be used is appropriately 1 to 20 times the molar amount, preferably 1 to 10 times the molar amount with respect to the compound [43].
  • the "activator” include 1H-tetrazole, 5-ethylthiotetrazole, 4,5-dichloroimidazole, 4,5-dicyanoimidazole, benzotriazole triflate, imidazole triflate, pyridinium triflate, N, N-diisopropylethylamine. , 2,4,6-colysine / N-methylimidazole.
  • the amount of the "activator" to be used is appropriately 1 to 20 times the molar amount, preferably 1 to 10 times the molar amount with respect to the compound [43].
  • the appropriate reaction temperature is 0 ° C to 120 ° C.
  • the reaction time varies depending on the type of raw material used, the reaction temperature, etc., but is usually 30 minutes to 24 hours.
  • Compound [B-2] is a compound in which each nucleoside unit constituting the compound is a nucleoside unit [4e], but all or part of the nucleoside unit [4e] is a nucleoside unit [4f] or a nucleoside unit [4g]. ] Can also be produced according to the same method as described above.
  • the “conversion yield (%)” means the rate at which the raw material is converted to the target product, and the peak corresponding to the target product detected by " ⁇ high performance liquid chromatography (hereinafter referred to as” HPLC ”)". Area (%) ⁇ ⁇ ⁇ Peak area (%) corresponding to the raw material detected by HPLC + Peak area (%) ⁇ ⁇ 100 corresponding to the target product detected by HPLC.
  • HPLC conditions For the product, 0.5 mg was dissolved in acetonitrile or 20% water / acetonitrile, and HPLC analysis was performed under the following conditions.
  • Example 1 4- (Octadecylamino) -4-oxobutanoic acid [(2S, 6R) -6- (5-methyl-2,4-dioxo-3,4-dihydropyrimidine-1 (2H) -yl) morpholine-2-yl) ] Methyl (hereinafter referred to as "G1-suc-morT-OFF”)
  • Step 1 Production of 4- (octadecylamino) -4-oxobutanoic acid (hereinafter referred to as "G1-suc") Succinic anhydride (8) in a solution of octadecane-1-amine (21.94 g) in dichloromethane (500 mL). .96 g, 1.1 eq) and triethylamine (17 mL, 1.5 eq.) Were added and stirred at room temperature for 7 hours. The mixture was concentrated under reduced pressure, 150 mL of acetone was added to the residue, and the mixture was stirred for 16 hours. The precipitate was suction filtered, washed with acetone (400 mL), and dried under reduced pressure at 30 ° C. for 3 hours to obtain G1-suc (29.1 g, 96.6%) as a white powder.
  • G1-suc octadecylamino) -4-oxobutanoic acid
  • Step 2 4- (Octadecylamino) -4-oxobutanoic acid [(2S, 6R) -6- (5-methyl-2,4-dioxo-3,4-dihydropyrimidine-1 (2H) -yl) morpholine-2 -Il] Methyl (hereinafter referred to as "G1-suc-morT-OFF”) production G1-suc (14.4 g) in tetrahydrofuran (150 mL) solution with 1-ethyl-3- (3-dimethylaminopropyl) Carbodiimide hydrochloride (8.56 g, 1.2 eq.) was added and stirred at room temperature.
  • G1-suc-morT-OFF Methyl
  • Example 2 Succinic acid ⁇ [(2S, 6R) -6- (5-methyl-2,4-dioxo-3,4-dihydropyrimidine-1-yl) morpholin-2-yl] methyl ⁇ ⁇ 2-octadeca Noyloxy-1-[(octadecanoyloxymethyl) ethyl] ⁇ (hereinafter referred to as "G2-suc-morT-OFF”)
  • Step 1 4-(Production of ((1,3-bis (stearoyloxy) propan-2-yl) oxy) -4-oxobutanoic acid (hereinafter referred to as "G2-suc") 2-hydroxypropane-1,3- Dichloromethane (8 mL) was added to 1 g (1.60 mmol) of diyl disteaert, then 176 mg (1.76 mmol) of succinic anhydride and 293 mg (2.40 mmol) of 4- (N, N-dimethylamino) pyridine were added, and 16 at room temperature. Stirred for hours.
  • G2-suc 2-hydroxypropane-1,3- Dichloromethane
  • Step 2 Succinic acid ⁇ [(2S, 6R) -6- (5-methyl-2,4-dioxo-3,4-dihydropyrimidine-1-yl) -4-trityl-morpholine-2-yl] methyl ⁇ ⁇ 2-Octadecanoyloxy-1-[(octadecanoyloxymethyl) ethyl] ⁇ (hereinafter referred to as "G2-suc-morT-ON”) production G2-suc (900 mg, 1.24 mmol) and 1- Ethyl-3- (3-dimethylaminopropyl) carbodiimide hydrochloride 277 mg (1.45 mmol) was added with dichloromethane (5.2 mL), followed by morT-OH 500 mg (1.03 mmol), 4- (N, N-dimethylamino).
  • Step 3 Production of G2-suc-morT-OFF Dichloromethane (4.2 mL) was added to G2-suc-morT-ON, and the mixture was stirred at 0 ° C. Then, 127 ⁇ L (0.62 mmol) of triisopropylsilane and 64 ⁇ L (0.82 mmol) of trifluoroacetic acid were added at 0 ° C., and the mixture was stirred at room temperature for 1 hour. After completion of the reaction, a saturated aqueous sodium hydrogen carbonate solution was added to the reaction solution, the mixture was extracted with dichloromethane, dried over sodium sulfate, and the solvent was distilled off. The obtained residue was purified by silica gel chromatography to obtain G2-suc-morT-OFF (373 mg, 95%).
  • Step 1 [ ⁇ (2S, 6R) -6- (5-methyl-2,4-dioxo-3,4-dihydropyrimidine-1 (2H) -yl) -4-tritylmorpholine-2-yl ⁇ methyl] succinate Production of Acid 1,3-Bis (Ole Oil Oxy) Propane-2-yl (hereinafter referred to as "G3-suc-morT-ON”) Using 2-hydroxypropane-1,3-diyldiolate as a raw material, 4-((1,3-Bis (oleoyloxy) propan-2-yl) oxy) -4-oxobutanoic acid (hereinafter referred to as "G3-suc”) was prepared in the same manner as in Step 1 of Example 2. Manufactured. Then, G3-suc-morT-ON was produced in the same manner as in Step 2 of Example 2.
  • G3-suc-morT-ON was prepared in the same manner as in
  • Step 2 Manufacture of G3-suc-morT-OFF Example 2 Manufactured in the same manner as in Step 3.
  • Step 2 4-oxo-4- (4-stearoylpiperazin-1-yl) butanoic acid ⁇ (2S, 6R) -6- (5-methyl-2,4-dioxo-3,4-dihydropyrimidine-1 (2H) ) -Il) -4-tritylmorpholine-2-yl ⁇ methyl (hereinafter referred to as "G4-suc-morT-ON”) production G4-suc (982 mg, 2.17 mmol) and 1-ethyl-3- () Tetrahydrofuran (10 mL) was added to 555 mg (2.90 mmol) of 3-dimethylaminopropyl) carbodiimide hydrochloride, and the mixture was stirred at 70 ° C.
  • Step 3 Manufacture of G4-suc-morT-OFF Manufactured in the same manner as in Step 3 of Example 2.
  • Example 5 4- (octadecylcarbamoyl) benzoic acid [(2S, 6R) -6- (5-methyl-2,4-dioxo-3,4-dihydropyrimidine-1 (2H) -yl) morpholin-2-yl) ] Methyl (hereinafter referred to as "G5-tpa-morT-OFF")
  • Step 1 4- (Octadecylcarbamoyl) benzoic acid [(2S, 6R) -6- (5-methyl-2,4-dioxo-3,4-dihydropyrimidine-1 (2H) -yl) -4-tritylmorpholine- 2-Il] Methyl (hereinafter referred to as "G5-tpa-morT-ON”) was produced using 4- (octadecylcarbamoyl) benzoic acid in the same manner as in Step 2 of Example 2.
  • Step 2 Manufacture of G5-tpa-morT-OFF Manufactured in the same manner as in Step 3 of Example 2.
  • Example 6 4- (4- (4- (octadecylcarbamoyl) benzoyl) piperazine-1-yl) -4-oxobutanoic acid ⁇ (2S, 6R) -6- (5-methyl-2,4-dioxo-3) , 4-Dihydropyrimidine-1 (2H) -yl) morpholine-2-yl ⁇ methyl (hereinafter referred to as "G6-suc-morT-OFF”)
  • Step 1 4- (4- (4- (Octadecylcarbamoyl) benzoyl) piperazine-1-yl) -4-oxobutanoic acid (hereinafter referred to as "G6-suc”) production 4- (octadecylcarbamoyl) instead of stearic acid ) Using benzoic acid, it was produced in the same manner as in Step 1 of Example 4.
  • Step 2 4- [4- ⁇ 4- (Okudadecylcarbamoyl) benzoyl ⁇ piperazine-1-yl] -4-oxobutanoic acid ⁇ (2S, 6R) -6- (5-methyl-2,4-dioxo-3, 4-Dihydropyrimidine-1 (2H) -yl) -4-tritylmorpholine-2-yl ⁇ methyl (hereinafter referred to as "G6-suc-morT-ON”) production The same method as in step 2 of Example 2. Manufactured in.
  • Step 3 Production of G6-suc-morT-OFF 4.6 mL of dichloromethane and 0.4 mL of 2,2,2-trifluoroethanol were added to 493 mg (0.47 mmol) of G6-suc-morT-ON, and the mixture was stirred at 0 ° C. Then, 145 ⁇ L (0.70 mmol) of triisopropylsilane and 53 ⁇ L (0.70 mmol) of trifluoroacetic acid were added at 0 ° C., and the mixture was stirred at room temperature for 1 hour.
  • Step 1 3,4,5-tris (octadecyloxy) benzoic acid ⁇ (2S, 6R) -6- (5-methyl-2,4-dioxo-3,4-dihydropyrimidine-1 (2H) -yl)- Production of 4-Trityl-Morpholine-2-yl ⁇ Methyl (hereinafter referred to as "G7-morT-ON") Using 3,4,5-trioctadecoxybenzoic acid, the same as in Step 2 of Example 2. Manufactured by the method.
  • Step 2 Manufacture of G7-morT-OFF Manufactured in the same manner as in Step 3 of Example 2.
  • Example 8 Succinic acid ⁇ (2S, 6R) -6- (5-methyl-2,4-dioxo-3,4-dihydropyrimidine-1 (2H) -yl) morpholine-2-yl ⁇ methyl (2-[ ⁇ 3,4,5-tris (octadecyloxy) benzoyloxy ⁇ oxy] ethyl) (hereinafter referred to as "G8-suc-morT-OFF”)
  • Step 1 Production of 2-Hydroxyethyl 3,4,5-trioctadecyloxybenzoate 1.5 g (1.60 mmol) of 3,4,5-trioctadecyloxybenzoate, 1-ethyl-3- (3-dimethylamino)
  • Step 2 Succinate ⁇ (2S, 6R) -6- (5-methyl-2,4-dioxo-3,4-dihydropyrimidine-1 (2H) -yl) -4-tritylmorpholine-2-yl ⁇ methyl ( 2- [ ⁇ 3,4,5-tris (octadecyloxy) benzoyloxy ⁇ oxy] ethyl) (hereinafter referred to as "G8-suc-morT-ON”) by the same method as in step 1 of Production Example 2.
  • G8-suc 4-Oxo-4- (2-[ ⁇ 3,4,5-tris (octadecyloxy) benzoyl ⁇ oxy] ethoxy) succinic acid (hereinafter referred to as "G8-suc"), and then Example 2.
  • G8-suc-morT-ON was obtained in the same manner as in Step 2.
  • Step 3 Manufacture of G8-suc-morT-OFF G8-suc-morT-OFF G8-suc-morT-OFF was obtained in the same manner as in Step 3 of Example 2.
  • Example 9 4- (dioctadecylamino) -4-oxobutanoic acid ⁇ (2S, 6R) -6- (5-methyl-2,4-dioxo-3,4-dihydropyrimidine-1 (2H) -yl) morpholine -2-Il ⁇ methyl (hereinafter referred to as "G9-suc-morT-OFF")
  • Step 1 4- (Dioctadecylamino) -4-oxobutanoic acid ⁇ (2S, 6R) -6- (5-methyl-2,4-dioxo-3,4-dihydropyrimidine-1 (2H) -yl) -4 Production of -tritylmorpholine-2-yl ⁇ methyl (hereinafter referred to as "G9-suc-morT-ON”) Using N-octadecyloctadecan-1-amine as a raw material, 4 in the same manner as in step 1
  • G9-suc -(Dioctadecylamino) -4-oxobutanoic acid
  • Step 2 Manufacture of G9-suc-morT-OFF Manufactured in the same manner as in Step 3 of Example 2.
  • Example 10 4-[ ⁇ 1- (octadecylamino) -1-oxo-3-phenylpropan-2-yl ⁇ amino] -4-oxobutanoic acid ⁇ (2S, 6R) -6- (5-methyl-2, 4-Dioxo-3,4-dihydropyrimidine-1 (2H) -yl) morpholine-2-yl ⁇ methyl (hereinafter referred to as "G10-suc-morT-OFF”)
  • Step 1 4-[ ⁇ 1- (octadecylamino) -1-oxo-3-phenylpropane-2-yl ⁇ amino] -4-oxobutanoic acid ⁇ (2S, 6R) -6- (5-methyl-2,4) -Dioxo-3,4-dihydropyrimidine-1 (2H) -yl) -4-tritylmorpholine-2-yl ⁇ methyl (hereinafter referred to as "G10-su
  • Step 2 Production of G10-suc-morT-OFF
  • the target product was obtained in the same manner as in Step 3 of Example 2.
  • Table 5 shows the chemical structural formulas of the compounds described in Examples 1 to 10 above.
  • Example 11 ((2S, 6R) -6- (4-benzamide-2-oxopyrimidine-1 (2H) -yl) -4-((dimethylamino) (((2S, 6R) -6- (5-) Methyl-2,4-dioxo-3,4-dihydropyrimidine-1 (2H) -yl) -4-tritylmorpholin-2-yl) methoxy) phosphoryl) morpholin-2-yl) methyl 4- (octadecylamino)- Production of 4-oxobutanoate (hereinafter referred to as G1-suc-PMO [CT] -ON (3)) In a 0.8 mL tube reactor after mixing 2 mL of liquid a shown below at a flow rate of 0.1 mL / min and 2 mL of liquid b shown below sent at a flow rate of 0.1 mL / min in a static mixer.
  • the reaction was carried out at room temperature for 4 minutes. Then, 2 mL of the following c solution fed at a flow rate of 0.1 mL / min was mixed, and the reaction was carried out at room temperature for 16.7 minutes in a 5 mL tube reactor. After that, the liquid d shown below was further mixed at a flow rate of 0.3 mL / min, and the reaction was carried out in a 10 mL tube reactor at room temperature for 16.7 minutes. The obtained solution was recovered with a 10% aqueous sodium thiosulfate solution, and then the organic layer was diluted 10-fold with acetonitrile and analyzed by HPLC (raw material Rt: 14.79 min, target product Rt: 17.13 min, conversion yield. 91.3%).
  • Solution a Dichloro (dimethylamino) phosphine 145 uL (1.26 mmol) was dissolved in 9 mL of dichloromethane.
  • Solution b 1-((2R, 6S) -6- (hydroxymethyl) -4-tritylmorpholine-2-yl) -5-methylpyrimidine-2,4 (1H, 3H) -dione (1) 609 mg (1) .26 mmol) and 549 ⁇ L (3.15 mmol) of N, N-diisopropylethylamine were dissolved in 9 mL of dichloromethane.
  • Solution c 4- (octadecylamino) -4-oxobutanoic acid [(2S, 6R) -6- (4-benzamide-2-oxopyrimidine-1 (2H) -yl) morpholine-2-yl] methyl (2) 430 mg (0.63 mmol) and 274 ⁇ L (1.58 mmol) of N, N-diisopropylethylamine were dissolved in 9 mL of dichloromethane. Liquid d: 651 mg (1.05 mmol) of Magnesium Monoperoxyphthalate Hexahydrate (MMPP) was dissolved in 42 mL of water.
  • MMPP Magnesium Monoperoxyphthalate Hexahydrate
  • Example 12 ((2S, 6R) -4-((dimethylamino) (((2S, 6R) -4-((dimethylamino)) (((2S, 6R) -6- (5-methyl-2,4) -Dioxo-3,4-dihydropyrimidine-1 (2H) -yl) -4-tritylmorpholine-2-yl) methoxy) phosphoryl) -6- (5-methyl-2,4-dioxo-3,4-dihydro Pyrimidine-1 (2H) -yl) morpholine-2-yl) methoxy) phosphoryl) -6- (5-methyl-2,4-dioxo-3,4-dihydropyrimidine-1 (2H) -yl) morpholin-2 -Il) Methyl 4- (octadecylamino) -4-oxobutanoate (hereinafter referred to as G1-suc-PMO [TT
  • Solution b 1-((2R, 6S) -6- (hydroxymethyl) -4-tritylmorpholine-2-yl) -5-methylpyrimidine-2,4 (1H, 3H) -dione (1) 169 mg (0) .35 mmol) and 137 ⁇ L (0.788 mmol) of N, N-diisopropylethylamine were dissolved in 2.5 mL of dichloromethane.
  • Solution c 162 mg (0.175 mmol) of G1-suc-PMO [TT] -OFF (4) and 76 ⁇ L (0.438 mmol) of N, N-diisopropylethylamine were dissolved in 2.5 mL of dichloromethane.
  • Example 13 ((2S, 6R) -6- (6-benzamide-9H-purine-9-yl) -4-((((2S, 6R) -6- (6-benzamide-9H-purine-9-)) Il) -4-tritylmorpholine-2-yl) methoxy) (dimethylamino) phosphoryl) morpholin-2-yl) methyl 4- (octadecylamino) -4-oxobutanoate (hereinafter, G1-suc-PMO [A] Bz- A Bz ] -ON (8)) After mixing 2 mL of liquid a shown below at a flow rate of 1 mL / min and 2 mL of liquid b shown below sent at a flow rate of 1 mL / min in a static mixer, the reaction was carried out in a 2 mL tube reactor at room temperature for 1 minute.
  • Solution b N-(9-((2R, 6S) -6- (hydroxymethyl) -4-tritylmorpholine-2-yl) -9H-purine-6-yl) benzamide (6) 55 mg (0.114 mmol) And 50 ⁇ L (0.290 mmol) of N, N-diisopropylethylamine was dissolved in 2.5 mL of dichloromethane.
  • Solution c 62 mg (0.0878 mmol) of G1-suc-morA-OFF (7) and 38 ⁇ L (0.220 mmol) of N, N-diisopropylethylamine were dissolved in 2.5 mL of dichloromethane.
  • Example 14 ((2S, 6R) -6- (6-benzamide-9H-purine-9-yl) -4-((dimethylamino) (((2S, 6R) -6- (5-methyl-2,,) 4-Dioxo-3,4-dihydropyrimidine-1 (2H) -yl) -4-tritylmorpholin-2-yl) methoxy) phosphoryl) morpholin-2-yl) methyl 4- (octadecylamino) -4-oxobuta Manufacture of Noate (hereinafter referred to as G1-suc-PMO [A Bz- T] -ON (9)) After mixing 2 mL of liquid a shown below at a flow rate of 1 mL / min and 2 mL of liquid b shown below sent at a flow rate of 1 mL / min in a static mixer, the reaction was carried out in a 2 mL tube reactor at room temperature for 1 minute.
  • Solution b 1-((2R, 6S) -6- (hydroxymethyl) -4-tritylmorpholine-2-yl) -5-methylpyrimidine-2,4 (1H, 3H) -dione (1) 110 mg (0) .216 mmol) and 73 ⁇ L (0.580 mmol) of N-ethylmorpholine were dissolved in 2.5 mL of dichloromethane.
  • Solution c 124 mg (0.176 mmol) of G1-suc-morA-OFF (7) and 76 ⁇ L (0.439 mmol) of N, N-diisopropylethylamine were dissolved in 2.5 mL of dichloromethane.
  • Example 15 ((2S, 6R) -4-((dimethylamino) (((2S, 6R) -6- (5-methyl-2,4-dioxo-3,4-dihydropyrimidine-1 (2H)-)- Il) -4-tritylmorpholine-2-yl) methoxy) phosphoryl) -6- (5-methyl-2,4-dioxo-3,4-dihydropyrimidine-1 (2H) -yl) morpholin-2-yl) Production of Methyl 4- (Octadecylamino) -4-oxobutanoate (hereinafter, G1-suc-PMO [TT] -ON (11)) 2 mL of liquid a shown below and 2 mL of liquid b shown below delivered at a flow rate of 0.5 mL / min were mixed in a static mixer and then brought to room temperature in a 2 mL tube reactor.
  • Solution b 1-((2R, 6S) -6- (hydroxymethyl) -4-tritylmorpholine-2-yl) -5-methylpyrimidine-2,4 (1H, 3H) -dione (1) 105 mg (0) .216 mmol) and 95 ⁇ L (0.549 mmol) of N, N-diisopropylethylamine were dissolved in 2.4 mL of dichloromethane.
  • Solution c 100 mg (0.166 mmol) of G1-suc-morT-OFF (10) and 72 ⁇ L (0.416 mmol) of N, N-diisopropylethylamine were dissolved in 2.4 mL of dichloromethane.

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WO2023127918A1 (ja) * 2021-12-27 2023-07-06 日本新薬株式会社 オリゴ核酸化合物の製造方法

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