Asymmetric auxiliary

Abstract

To provide a chiral reagent or salt thereof used for synthesizing a stereo-controlled phosphorous atom modified oligonucleotide derivative.SOLUTION: A chiral reagent has the following formula (I). In formula (I), G1 and G2 are H, a nitro group (-NO2), a halogen atom, a cyano group (-CN), a specific phenyl group or a group of (III), or G1 and G2 together form a ring.SELECTED DRAWING: None

Description

[Technical Field]

[0001] This invention relates to a chiral reagent used for synthesizing stereochemically controlled phosphorus atom-modified oligonucleotide derivatives. [Background technology]

[0002] Japanese Patent Publication No. 2005-89441 discloses a method for producing nucleotide derivatives called the oxazaphosphoridine method. However, the monomer isolation yield is low, and this method requires a special capping agent that is not commercially available. Furthermore, the resulting monomers are chemically unstable. Moreover, the isolation yield of oligonucleotide derivatives is not high. The low yield of oligonucleotide derivatives is thought to be due to the degradation reaction under the deprotection step.

[0003] International Publication No. 2010 / 064146 discloses a method for generating nucleotide derivatives. The method disclosed therein requires a special capping agent that is not commercially available. Furthermore, the isolation yield of oligonucleotide derivatives is not high. The low yield is thought to be due to the degradation reaction under the deprotection step. This tendency becomes more pronounced as the length of the oligonucleotide derivative increases.

[0004] International Publication No. 2012 / 039448 discloses asymmetric auxiliary groups used to generate stereocontrolled phosphorus atom-modified oligonucleotide derivatives. [Prior art documents] [Patent Documents]

[0005] [Patent Document 1] Japanese Patent Publication No. 2005-89441 [Patent Document 2] International Publication No. 2010 / 064146 Pamphlet [Patent Document 3] International Publication No. 2012 / 039448 Pamphlet [Overview of the project]

[0006] The first aspect of the present invention relates to a chiral reagent or a salt thereof. The chiral reagent has the following chemical formula (I).

[0007]

Chemical formula

[0008] In formula (I), G 1 and G 2 are independently a hydrogen atom, a nitro group (-NO2), a halogen atom, a cyano group (-CN), a group of formula (II), (III) or (V), or both G 1 and G 2 together form a group of formula (IV).

[0009]

Chemical formula

[0010] In formula (II), G 21 to G 23 are independently a hydrogen atom, a nitro group, a halogen atom, a cyano group or a C 1-3 alkyl group.

[0011]

Chemical formula

[0012] In formula (III), G 31 to G 33 are independently a C 1-4 alkyl group, a C 6-14 aryl group, a C 1-4 alkoxy group, a C 7-14 aralkyl group, a C 1-4 alkyl C 6-14 aryl group, a C 1-4 alkoxy C 6-14 aryl group, or a C 6-14 aryl C 1-4 alkyl group.

[0013] [ka]

[0014] In equation (IV), G 41 ~G 46 These are independently a hydrogen atom, a nitro group, a halogen atom, a cyano group, or C 1-3 It is an alkyl group.

[0015] [ka]

[0016] In equation (V), G 51 ~G 53 These are, independently, a hydrogen atom, a nitro group, a halogen atom, a cyano group, and C 1-3 Alkyl or C 1-3 It is an alkyloxy group.

[0017] G 3 and G 4 These are, independently, hydrogen atoms and C 1-3 Alkyl alkyl group, C 6-14 It is an aryl group, or G 3 and G 4 Together, they form a heteroatom-containing ring with 3 to 16 carbon atoms, along with the NH moiety in formula (I).

[0018] A preferred embodiment is one in which the chiral reagent has the following chemical formula (I').

[0019] [ka]

[0020] In equation (I'), G 1 and G 2 This is the same as above. That is, G 1 and G 2Independently, these are a hydrogen atom, a nitro group, a halogen atom, a cyano group, a group of formula (II) or (III), or G 1 and G 2 Together, they form the base of equation (IV).

[0021] A preferred embodiment is in which the chiral reagent has chemical formula (I') and G 1 and G 2 Each of these is a base of equation (II), and in this case, G 21 ~G 23 These are independently a hydrogen atom, a nitro group, a halogen atom, a cyano group, or C 1-3 It is an alkyl group.

[0022] A preferred embodiment is in which the chiral reagent has chemical formula (I') and G 1 and G 2 Each of these is a base of equation (II), and G 21 ~G 23 Each of these is a hydrogen atom.

[0023] A preferred embodiment is in which the chiral reagent has chemical formula (I') and G 1 G is a hydrogen atom, 2 is the basis of equation (II), and G 21 ~G 23 However, independently, hydrogen atoms, nitro groups, halogen atoms, cyano groups, or C 1-3 It is an alkyl group.

[0024] A preferred embodiment is in which the chiral reagent has chemical formula (I') and G 1 G is a hydrogen atom, 2 is the basis of equation (II), and G 21 and G 22 Each of them is a hydrogen atom, G 23 It is a nitro group.

[0025] A preferred embodiment is in which the chiral reagent has chemical formula (I') and G 1 G is a hydrogen atom, 2 is the basis of equation (III), and G 31 ~G 33is, independently, C 1-4 an alkyl group, C 6-14 an aryl group, C 7-14 an aralkyl group, C 1-4 an alkyl C 6-14 aryl group, C 1-4 an alkoxy C 6-14 aryl group, or C 6-14 an aryl C 1-4 alkyl group.

[0026] A preferred embodiment is that the chiral reagent has the chemical formula (I’), and G 1 is a hydrogen atom, G 2 is a group of formula (III), G 31 ~G 33 is, independently, C 1-4 an alkyl group, a C6 aryl group, C 7-10 an aralkyl group, C 1-4 an alkyl C6 aryl group, C 1-4 an alkoxy C6 aryl group, or a C6 aryl C 1-4 alkyl group.

[0027] A preferred embodiment is that the chiral reagent has the chemical formula (I’), and G 1 is a hydrogen atom, G 2 is a group of formula (III), G 31 ~G 33 is, independently, C 1-4 an alkyl group or a C6 aryl group. Examples of the C 1-4 alkyl group are a methyl group, an ethyl group, an n-propyl group, an isopropyl group, an n-butyl group and a tert-butyl group (tertiary butyl group).

[0028] A preferred embodiment is that the chiral reagent has the chemical formula (I’), and G 1 is a hydrogen atom, G 2 is a group of formula (III), G 31 ~G 33 are, independently, C 1-4 alkyl groups.

[0029] A preferred embodiment is that the chiral reagent has the chemical formula (I’), and G1 is a hydrogen atom, G 2 is a group of formula (III), G 31 and G 33 is a C6 aryl group, G 32 is C 1-4 alkyl group.

[0030] A preferred embodiment is that the chiral reagent has the chemical formula (I’), and G 1 and G 2 together form a group of formula (IV), and G 41 ~G 46 are each independently a hydrogen atom, a nitro group, a halogen atom, a cyano group or a C 1-4 alkyl group.

[0031] A preferred embodiment is that the chiral reagent has the chemical formula (I’), and G 1 and G 2 together form a group of formula (IV), wherein G 41 ~G 46 each is a hydrogen atom.

[0032] [[ID=4​​​​​​​​​​​​​​​​​​​​​​​​(S)-2-(methyldiphenylsilyl)-1-((S)-pyrrolidine-2-yl)ethanol(IIIa) (R)-2-(methyldiphenylsilyl)-1-((R)-1-pyrrolidine-2-yl)ethanol(III-b) (S)-2-(trimethylsilyl)-1-((S)-1-pyrrolidine-2-yl)ethanol(Va) (R)-2,2-diphenyl-1-((S)-pyrrolidine-2-yl)ethanol(VII-a) (S)-2,2-diphenyl-1-((R)-pyrrolidine-2-yl)ethanol(VII-b) (R)-2-(4-nitrophenyl)-1-((S)-pyrrolidine-2-yl)ethanol(IX-a) (S)-2-(4-nitrophenyl)-1-((R)-pyrrolidine-2-yl)ethanol(IX-b) (R)-(9H-fluorolene-9-yl)((S)-pyrrolidine-2-yl)methanol(XI-a) (S)-2-Tosyl-1-((S)-1-Tritylpyrrolidine-2-yl)ethanol(XIII-a) (R)-2-Tosyl-1-((R)-1-Tritylpyrrolidine-2-yl)ethanol(XIII-b) It is selected from one of the following.

[0034] A second aspect of the present invention relates to a nucleoside 3'-phosphoramidite derivative represented by formula (Va) or (Vb).

[0035] [ka]

[0036] In equations (Va) and (Vb), G 1 ~G 4 This is the same as above, G 5 B is a protecting group for the hydroxyl group, and Bs is a group selected from the group represented by the following formulas (VI) to (XI) or their derivatives.

[0037] [ka]

[0038] Examples of Bs include adenine, thymine, cytosine, guanine, uracil, 5-methylcytosine, or their derivatives.

[0039] R 2 These are hydrogen, -OH, -SH, and -NR d R d -N3, halogen, alkyl, alkenyl, alkynyl, alkyl-Y 1 -, Alkenil-Y 1 -, Alkinyl-Y 1 -, Aryl-Y 1 -, heteroaryl-Y 1 -, -OR b , or -SR b And in this case, R b This is the blocking section. Y 1 O, NR d It is S, or Se. R d These are independently hydrogen, alkyl, alkenyl, alkynyl, aryl, acyl, substituted silyl, carbamate, and -P(O)(R e )2, or -HP(O)(R e ) R e These are independently hydrogen, alkyl, aryl, alkenyl, alkynyl, and alkyl-Y 2 -, Alkenil-Y 2 -, Alkinyl-Y 2 -, Aryl-Y 2 -, or heteroaryl-Y 2 -, or, Na + Li + , or K + It is a positive ion. Y 2 O, NR d , or S.

[0040] R3 These are groups represented by -CH2-, -(CH2)2-, -CH2NH-, or -CH2N(CH3)-.

[0041] G 5 Examples include trityl, 4-monomethoxytrityl, 4,4'-dimethoxytrityl, 4,4',4”-trimethoxytrityl, 9-phenylxanthin-9-yl (Pixyl), and 9-(p-methoxyphenyl)xanthin-9-yl (MOX).

[0042] A preferred embodiment of the second aspect is that the nucleoside 3'-phosphoramidite derivative is represented by formula (Va') or (Vb').

[0043] [ka]

[0044] In equations (Va') and (Vb'), G 1 , G 2 , G 5 , Bs, R 2 , and R 3 This is the same as above.

[0045] A third aspect of the present invention relates to a method for synthesizing stereocontrolled phosphorus atom-modified oligonucleotide derivatives.

[0046] The first step involves reacting a molecule containing an achiral H-phosphonate moiety, a first activating reagent, and a chiral reagent or a salt thereof to form a monomer. The chiral reagent has chemical formula (I) or (I'), and its monomer can be represented by formula (formula)(Va), (Vb), (Va'), or (Vb'). The monomer reacts with a second activating reagent and a nucleoside to form a concentrated intermediate. The next step involves converting the concentrated intermediate into a nucleic acid containing a chiral X-phosphonate moiety.

[0047] Based on this method, it is possible to use stable commercially available materials as starting materials. Using achiral starting materials, it is possible to produce stereocontrolled phosphorus atom-modified oligonucleotide derivatives.

[0048] As shown in the examples, the method of the present invention does not cause degradation under the deprotection step. Furthermore, this method does not require special capping agents to produce phosphorus atom-modified oligonucleotide derivatives.

[0049] A fourth aspect of the present invention relates to a method for synthesizing stereocontrolled phosphorus atom-modified oligonucleotide derivatives using chiral monomers.

[0050] The first step involves reacting a nucleoside 3'-phosphoramidite derivative represented by (Va), (Vb), (Va'), or (Vb') with a second activating reagent and the nucleoside to form a concentrated intermediate. The second step involves converting the concentrated intermediate into a nucleic acid containing a chiral X-phosphonate moiety.

[0051] Reference All publications and patent applications disclosed herein are incorporated herein by reference in their entirety to the same extent that each individual publication or patent application is specifically and individually referred to. [Brief explanation of the drawing]

[0052] [Figure 1] This figure shows the UPLC profile when generating oligonucleotide derivatives using monomer 4b. [Figure 2] This figure shows the UPLC profile when generating oligonucleotide derivatives using 25 monomers. [Modes for carrying out the invention]

[0053] The term "nucleic acid" encompasses poly- or oligo-ribonucleotides (RNA) and poly- or oligo-deoxyribonucleotides (DNA); RNA or DNA derived from N-glycosides or C-glycosides of nucleic acid bases and / or modified nucleic acid bases; nucleic acids derived from sugars and / or modified sugars; and nucleic acids derived from phosphate bridges and / or modified phosphorus atom bridges. This term includes nucleic acids containing any combination of nucleic acid bases, modified nucleic acid bases, sugars, modified sugars, phosphate bridges, or modified phosphorus atom bridges. Examples include, but are not limited to, nucleic acids containing a ribose moiety, nucleic acids containing a deoxyribose moiety, nucleic acids containing both a ribose moiety and a deoxyribose moiety, and nucleic acids containing ribose and modified ribose moieties. The prefix "poly-" refers to nucleic acids containing approximately 1 to approximately 10,000 nucleotide monomer units, within which the prefix "oligo-" refers to nucleic acids containing approximately 1 to approximately 200 nucleotide monomer units.

[0054] The term "nucleic acid base" refers to the part of a nucleic acid involved in hydrogen bonding, which links a nucleic acid chain to another complementary chain in a sequence-specific manner. The most common naturally occurring nucleic acid bases are adenine (A), guanine (G), uracil (U), cytosine (C), 5-methylcytosine, and thymine (T).

[0055] The term "modified nucleic acid base" refers to a portion of a nucleic acid base that can be replaced. Modified nucleic acid bases mimic the spatial arrangement, electrical properties, or any other physicochemical properties of nucleic acid bases and retain the properties of hydrogen bonding, which links one nucleic acid chain to another in a sequence-specific manner. Modified nucleic acid bases can pair with any of the five naturally occurring bases (uracil, thymine, adenine, cytosine, or guanine) without substantially affecting melting behavior, recognition by intracellular enzymes, or the activity of the oligonucleotide double helix.

[0056] The term "nucleoside" refers to the portion of a nucleic acid base or modified nucleic acid base that is covalently bonded to a sugar or modified sugar.

[0057] The term "sugar" refers to ring-closed and / or ring-open monosaccharides. Examples of sugars, though not limited to, include ribose, deoxyribose, pentofuranose, pentopyranose, and hexopyranoses.

[0058] The term "modified sugar" refers to a portion of a sugar molecule that can be replaced by another sugar molecule. Modified sugars mimic the spatial arrangement, electrical properties, or other physicochemical properties of a sugar molecule.

[0059] The term "nucleotide" refers to the portion of a nucleic acid base or modified nucleic acid base that is covalently bonded to a sugar or modified sugar, and to a phosphate group or modified phosphorus atom.

[0060] The term "chiral reagent" refers to a compound that is chiral or enantiopurine and can be used for asymmetric induction in nucleic acid synthesis.

[0061] The term "chiral ligand" or "chiral auxiliary agent" refers to a part of a reaction that is chiral or enantiophorous and controls the stereochemical outcome of the reaction.

[0062] In condensation reactions, the term "activating reagent" refers to a reagent that activates less reactive sites, thereby increasing their susceptibility to attack by nucleophiles.

[0063] The term "blocking moiety" refers to a group that temporarily masks the reactivity of a functional group. The functional group can then be demasked by removing the blocking moiety.

[0064] The terms "boronating agent," "sulfur electrophile," and "selenium electrophile" refer to compounds useful in modification processes that are used to introduce BH3, S, and Se groups, respectively, for modification of phosphorus atoms.

[0065] The term "part" refers to a specific segment or functional group of a molecule. A chemical part is often a recognized chemical substance that is embedded in or attached to a molecule.

[0066] The term "solid-phase support" refers to a support that enables the mass production of nucleic acid synthesis and can be reused when needed. In this specification, this term refers to a polymer that is insoluble in the medium used in the reaction steps carried out to synthesize nucleic acids and is derivatized because it contains reactive groups.

[0067] The term "linking portion" refers to a portion optionally positioned between a terminal nucleoside and a solid support, or between a terminal nucleoside and another nucleoside, nucleotide, or nucleic acid.

[0068] In this specification, “treatment” or “to treat,” or “to alleviate” or “to induce remission” are used synonymously. These terms refer to an approach to obtain a beneficial or desirable outcome, including, but not limited to, therapeutic benefits and / or preventive benefits. Therapeutic benefits mean the eradication or remission of the underlying disease being treated. Therapeutic benefits are also achieved by the eradication or remission of one or more physiological signs associated with the underlying disease, such that improvement is observed in the patient, even though the patient may still be susceptible to the underlying disease. For preventive benefits, a composition may be administered to a patient at risk of developing a particular disease, or to a patient in whom one or more physiological signs of the disease have been reported, even if the disease has not been diagnosed.

[0069] In this specification, “therapeutic effect” includes therapeutic and / or preventive benefits as described above. Preventive effects include delaying or eliminating the onset of a disease or condition, delaying or eliminating the onset of signs of a disease or condition, slowing, stopping, or reversing the progression of a disease or condition, or any combination thereof.

[0070] An "alkyl" group refers to an aliphatic hydrocarbon group. The alkyl portion may be a saturated alkyl group (meaning it does not contain any unsaturated bonds, such as carbon-carbon double or carbon-carbon triple bonds) or an unsaturated alkyl group (meaning it contains at least one unsaturated bond). The alkyl portion may be saturated or unsaturated, branched or linear, or may contain a cyclic portion. The alkyl bond site is on a carbon atom that is not part of the ring.

[0071] The “alkyl” portion may have 1 to 10 carbon atoms (wherever it appears herein, the numerical range such as “1 to 10” refers to each integer within the given range; for example, “1 to 10 carbon atoms” means that the alkyl group may consist of 10 or fewer carbon atoms, such as 1 carbon atom, 2 carbon atoms, 3 carbon atoms, etc., but this definition also includes the term “alkyl” for which no numerical range is specified). Alkyl includes both branched alkyl groups and linear alkyl groups. Alkyl groups of compounds described herein may be designated “C1-C6 alkyl” or similar names. As just one example, “C1-C6 alkyl” indicates that the alkyl chain contains 1, 2, 3, 4, 5, or 6 carbon atoms; that is, the alkyl chain is selected from the group consisting of methyl, ethyl, propyl, isopropyl, n-butyl, iso-butyl, sec-butyl, and tert-butyl. Typical alkyl groups include, but are not limited to, methyl, ethyl, propyl, isopropyl, butyl, isobutyl, tert-butyl, pentyl, hexyl, allyl, cyclopropylmethyl, cyclobutylmethyl, cyclopentylmethyl, cyclohexylmethyl, and the like. In one embodiment, the alkyl group is a C1-C6 alkyl group.

[0072] C 1-3 An alkyl group refers to a linear or branched alkyl group having 1 to 3 carbon atoms. 1-3 Examples of alkyl groups are methyl, ethyl, propyl, and isopropyl. 1-4An alkyl group refers to a linear or branched alkyl group having 1 to 4 carbon atoms. 1-4 Examples of alkyl groups include methyl, ethyl, propyl, isopropyl, butyl, isobutyl, and tert-butyl.

[0073] In this specification, the term "aryl" refers to an aromatic ring in which each of the ring-forming atoms is a carbon atom. Aryl rings are formed by 5, 6, 7, 8, 9, or more than 9 carbon atoms. The aryl group can be substituted or unsubstituted. In one embodiment, the aryl is phenyl or naphthalenyl. Depending on the structure, the aryl group can be a monoradical or a diradical (i.e., an arylene group). In one embodiment, the aryl is C6-C 10 It is Ariel.

[0074] C 6-14 An aryl group refers to an aryl group that has 6 to 14 carbon atoms. 6-14 Examples of aryl groups include phenyl, biphenyl, naphthyl, anthrasyl, indanyl, phthalimidyl, naphthymidyl, phenanthridine, and tetrahydronaphthyl.

[0075] The term "aralkyl" refers to an alkyl group substituted with an aryl group. Suitable aralkyl groups include benzyl, picolyl, and similar groups, all of which may be substituted as desired.

[0076] The "acyl moiety" refers to an alkyl (C=O), aryl (C=O), or aralkyl (C=O) group. The acyl moiety may have an intervening moiety (Y) between the carbonyl group and the hydrocarbon group, which is oxy, amino, thio, or seleno. For example, an acyl group may be alkyl-Y-(C=O), aryl-Y-(C=O), or aralkyl-Y-(C=O).

[0077] An "alkenyl" group is a linear, branched, or cyclic hydrocarbon group containing at least one carbon-carbon double bond. Alkenyl groups can be substituted.

[0078] An "alkynyl" group is a straight-chain, branched-chain, and cyclic hydrocarbon group containing at least one carbon-carbon triple bond. The alkynyl group may be substituted.

[0079] An "alkoxy" group refers to an alkyl group bonded to oxygen, i.e., an (alkyl)-O- group, where alkyl is as defined herein. Examples include methoxy (-OCH3) or ethoxy (-OCH2CH3) groups.

[0080] An "alkenyloxy" group refers to an alkenyl group bonded to oxygen, i.e., an (alkenyl)-O- group, where alkenyl is as defined herein.

[0081] An "alkynyloxy" group refers to an alkynyl group bonded to oxygen, i.e., an (alkynyl)-O- group, where alkynyl is as defined herein.

[0082] An "aryloxy" group refers to an aryl group bonded to oxygen, i.e., an (aryl)-O- group, where aryl is as defined herein. An example is the phenoxy (-OC6H5) group.

[0083] The term "alkylseleno" refers to an alkyl group having a substituted seleno group bonded thereto, i.e., an (alkyl)-Se- group, where alkyl is as defined herein.

[0084] The term "alkenylseleno" refers to an alkenyl group having a substituted seleno group bonded thereto, i.e., an (alkenyl)-Se- group, where alkenyl is as defined herein.

[0085] The term "alkynylseleno" refers to an alkynyl group having a substituted seleno group bonded thereto, i.e., an (alkynyl)-Se- group, where alkenyl is as defined herein.

[0086] The term "alkylthio" refers to an alkyl group bonded to a bridging sulfur atom, i.e., an (alkyl)-S- group, where alkyl is defined herein. For example, alkylthios include methylthios and similar groups.

[0087] The term "alkenylthio" refers to an alkenyl group bonded to a bridging sulfur atom, i.e., an (alkenyl)-S- group, where alkenyl is defined herein.

[0088] The term "alkynylthio" refers to an alkynyl group bonded to a bridging sulfur atom, i.e., an (alkynyl)-S- group, where alkenyl is defined herein.

[0089] The term "alkylamino" refers to an amino group substituted with at least one alkyl group, i.e., -NH(alkyl) or -N(alkyl)2, where alkyl is defined herein.

[0090] The term "alkenylamino" refers to an amino group substituted with at least one alkenyl group, i.e., -NH(alkenyl) or -N(alkenyl)2, where alkenyl is defined herein.

[0091] The term "alkynylamino" refers to an amino group substituted with at least one alkynyl group, i.e., -NH(alkynyl) or -N(alkynyl)2, where alkynyl is defined herein.

[0092] The term "halogen" is intended to include fluorine, chlorine, bromine, and iodine.

[0093] A "fluorescent group" refers to a molecule that emits light of a different wavelength when excited by light of a selected wavelength. Fluorescent groups are not limited to, but include indole, fluorescein, tetramethylrhodamine, Texas Red, BODIPY, 5-[(2-aminoethyl)amino]naphthalene-1-sulfonic acid (EDANS), coumarin, and Lucifer Yellow.

[0094] The "ammonium ion" has the chemical formula NH4. + It is a positively charged polyatomic cation.

[0095] An "alkylammonium ion" is an ammonium ion in which at least one of its hydrogen atoms is replaced by an alkyl group, where alkyl is defined herein. Examples include the triethylammonium ion and the N,N-diisopropylethylammonium ion.

[0096] The "iminium ion" has a general structure R2C=NR2 + It has the following. The R group refers to an alkyl, alkenyl, alkynyl, or aryl group as defined herein. "Heteroaromatic iminium ion" refers to an iminium ion in which nitrogen and the R group to which it is bonded form an aromatic heterocycle. "Heterocyclic iminium ion" refers to an iminium ion in which nitrogen and the R group to which it is bonded form a heterocycle.

[0097] The term "amino" or "amine" refers to -N(R h ) refers to two radical groups, and in this case, each R h Unless otherwise specifically specified herein, these are independently hydrogen, alkyl, fluoroalkyl, carbocykryl, carbocykrylalkyl, aryl, aralkyl, heterocyclyl, heterocyclrylalkyl, heteroaryl, or heteroarylalkyl. -N(R h )Two units, in addition to hydrogen, have two R hIf they are present, they can combine with nitrogen atoms to form 4-membered, 5-membered, 6-membered, or 7-membered rings. For example, -N(R h )2 means including, but not limited to, 1-pyrrolidinyl and 4-morpholinyl. Any one or more of hydrogen, alkyl, fluoroalkyl, carbocykyl, carbocykylalkyl, aryl, aralkyl, heterocyclyl, heterocyclylalkyl, heteroaryl or heteroarylalkyl, optionally independently of alkyl, heteroalkyl, alkenyl, alkynyl, cycloalkyl, heterocycloalkyl, aryl, arylalkyl, heteroaryl, heteroarylalkyl, hydroxy, halo, cyano, trifluoromethyl, trifluoromethoxy, nitro, trimethylsilyl, -OR i , -SR i -OC(O)R i , -N(R i )2, -C(O)R i , -C(O)OR i ,-OC(O)N(R i )2, -C(O)N(R i )2, -N(R i )C(O)OR, -N(R i )C(O)R i , -N(R i )C(O)N(R i )2, N(R i )C(NR i )N(R i )2, -N(R i )S(O) t R i (wherein t is 1 or 2), -S(O), or -S(O) t N(R i )2 (where t is 1 or 2) is substituted with one or more substituents, in which case each R i These are independently hydrogen, alkyl, fluoroalkyl, carbocykyl, carbocykylalkyl, aryl, aralkyl, heterocyclyl, heterocyclylalkyl, heteroaryl, or heteroarylalkyl.

[0098] In this specification, "carbamate" refers to a molar group bonded to an amino group having the formula -C(O)OR, where R is alkyl, fluoroalkyl, carbocyclyl, carbocyclylalkyl, aryl, aralkyl, heterocyclyl, heterocyclylalkyl, heteroaryl, or heteroarylalkyl. Examples, but not limited to, include Boc(tert-butyl-OC(O)-), CBz(benzyl-OC(O)-), Teoc(Me3SiCH2CH2OC(O)-), alloc(allyl-OC(O)-), or Fmoc(9-fluorenylmethyl-OC(O)-).

[0099] In this specification, "substituted silyl" refers to the moiety having the formula R3Si-. Examples, but not limited to, include the TBDMS (tert-butyldimethylsilyl), TBDPS (tert-butyldiphenylsilyl), or TMS (trimethylsilyl) group.

[0100] The term "thiol" refers to the -SH group, which can be a substituted thiol group, namely -SR. J The base is included. In the above equation, R J Each of these is independently a substituted or unsubstituted alkyl, cycloalkyl, alkenyl, alkynyl, arylaralkyl, heterocyclyl, or heterocyclylalkyl group as defined herein.

[0101] A first aspect of the present invention relates to a chiral reagent or a salt thereof. The chiral reagent has the following chemical formula (I). The term "chiral reagent" refers to a chemical composition used to produce stereocontrolled phosphorus-modified nucleotide or oligonucleotide derivatives. The chiral reagent reacts with a nucleotide to form a chiral intermediate.

[0102] [ka]

[0103] In equation (I), G 1 and G 2is, independently, a hydrogen atom, a nitro group, a halogen atom, a cyano group (-CN), a group of formula (II), (III) or (V), or G 1 and G 2 both together form a group of formula (IV).

[0104] [Chemical formula]

[0105] In formula (II), G 21 ~G 23 are, independently, a hydrogen atom, a nitro group, a halogen atom, a cyano group or a C 1-3 alkyl group. A preferred example of G 21 ~G 23 is a hydrogen atom.

[0106] [Chemical formula]

[0107] In formula (III), G 31 ~G 33 are, independently, a C 1-4 alkyl group, a C 6-14 aryl group, a C 1-4 alkoxy group, a C 7-14 aralkyl group, a C 1-4 alkyl C 6-14 aryl group, a C 1-4 alkoxy C 6-14 aryl group, or a C 6-14 aryl C 1-4 alkyl group. Examples of C 1-4 alkyl C 6-14 aryl groups are the methylphenyl group and the ethylphenyl group. Examples of C 1-4 alkoxy C 6-14 aryl groups are the methoxyphenyl group and the ethoxyphenyl group. Examples of C 6-14 aryl C 1-4 alkyl groups are the benzyl group and the phenylethyl group. G 31 ~G 33Preferred examples are, independently, methyl and phenyl groups.

[0108] [ka]

[0109] In equation (IV), G 41 ~G 46 These are independently a hydrogen atom, a nitro group, a halogen atom, a cyano group, or C 1-3 It is an alkyl group. 41 ~G 46 A preferred example is the hydrogen atom.

[0110] [ka]

[0111] In equation (V), G 51 ~G 53 These are, independently, a hydrogen atom, a nitro group, a halogen atom, a cyano group, and C 1-3 Alkyl or C 1-3 It is an alkyloxy group.

[0112] G 3 and G 4 These are, independently, hydrogen atoms and C 1-3 Alkyl alkyl group, C 6-14 It is an aryl group, or G 3 and G 4 Both together form a heteroatom-containing ring with 3 to 16 carbon atoms. 3 and G 4 A preferred example is one in which, together with the NH moiety in formula (I), they form a heteroatom-containing ring having 3 to 16 carbon atoms.

[0113] A preferred embodiment is one in which the chiral reagent has the following chemical formula (I').

[0114] [ka]

[0115] In equation (I'), G 1 and G 2 This is the same as above, G 1 and G 2 Independently, these are a hydrogen atom, a nitro group, a halogen atom, a cyano group, a group of formula (II) or (III), or G 1 and G 2 Together, they form the basis of equation (IV).

[0116] A preferred embodiment is in which the chiral reagent has chemical formula (I') and G 1 and G 2 Each of these is a base of equation (II), and in this case, G 21 ~G 23 These are independently a hydrogen atom, a nitro group, a halogen atom, a cyano group, or C 1-3 It is an alkyl group.

[0117] A preferred embodiment is in which the chiral reagent has chemical formula (I') and G 1 and G 2 Each of these is a base of equation (II), and G 21 ~G 23 Each of these is a hydrogen atom.

[0118] A preferred embodiment is in which the chiral reagent has chemical formula (I') and G 1 G is a hydrogen atom, 2 is the basis of equation (II), and G 21 ~G 23 However, independently, hydrogen atoms, nitro groups, halogen atoms, cyano groups, or C 1-3 It is an alkyl group.

[0119] A preferred embodiment is in which the chiral reagent has chemical formula (I') and G 1 G is a hydrogen atom, 2 is the basis of equation (II), and G 21 and G 22 Each of them is a hydrogen atom, G 23 This is a nitro group (-NO2).

[0120] A preferred embodiment is in which the chiral reagent has chemical formula (I') and G 1 G is a hydrogen atom, 2 is the basis of equation (III), and G 31 ~G 33 However, independently, C 1-4 Alkyl alkyl group, C 6-14 Aryl group, C 7-14 Aralkyl group, C 1-4 Alkyl C 6-14 Aryl group, C 1-4 Alkoxy C 6-14 Aryl group, or C 6-14 Aryl C 1-4 It is an alkyl group.

[0121] A preferred embodiment is in which the chiral reagent has chemical formula (I') and G 1 G is a hydrogen atom, 2 is the basis of equation (III), and G 31 ~G 33 However, independently, C 1-4 Alkyl group, C6 aryl group, C 7-10 Aralkyl group, C 1-4 Alkyl C6 aryl group, C 1-4 Alkoxy C6 aryl group, or C6 aryl C 1-4 It is an alkyl group.

[0122] A preferred embodiment is in which the chiral reagent has chemical formula (I') and G 1 G is a hydrogen atom, 2 is the basis of equation (III), and G 31 ~G 33 However, independently, C 1-4 It is an alkyl group or a C6 aryl group (phenyl group). 1-4 Examples of alkyl groups include methyl, ethyl, n-propyl, isopropyl, n-butyl, and tert-butyl groups.

[0123] A preferred embodiment is in which the chiral reagent has chemical formula (I') and G 1 G is a hydrogen atom, 2 is the basis of equation (III), and G31 ~G 33 However, independently, C 1-4 It is an alkyl group.

[0124] A preferred embodiment is in which the chiral reagent has chemical formula (I') and G 1 G is a hydrogen atom, 2 is the basis of equation (III), and G 31 and G 33 However, it is a C6 aryl group (phenyl group), G 32 C 1-2 It is an alkyl group.

[0125] A preferred embodiment is in which the chiral reagent has chemical formula (I') and G 1 and G 2 However, together they form the basis of equation (IV), G 41 ~G 46 However, independently, hydrogen atoms, nitro groups, halogen atoms, cyano groups, or C 1-3 It is an alkyl group.

[0126] A preferred embodiment is in which the chiral reagent has chemical formula (I') and G 1 and G 2 However, together they form the basis of equation (IV), and in this case, G 41 ~G 46 Each of these is a hydrogen atom.

[0127] A preferred embodiment is in which the chiral reagent has chemical formula (I') and G 1 G is a hydrogen atom, 2 This is the basis of equation (V). Furthermore, G 51 ~G 53 Each of these is independently a hydrogen atom, a nitro group, a methyl group, or a methoxy group. A more preferred embodiment is G 1 G is a hydrogen atom, 2 This is the basis of equation (V), and in this case, G 51 and G 53 Each of them is a hydrogen atom, G 53 It is a 4-methyl group.

[0128] A preferred embodiment is one in which the chiral reagents are III-a, III-b, Va, VII-a, VII-b, IX-a, IX-b, XI-a, XIII-a and XIII-b: (S)-2-(methyldiphenylsilyl)-1-((S)-pyrrolidine-2-yl)ethanol(IIIa) (R)-2-(methyldiphenylsilyl)-1-((R)-1-pyrrolidine-2-yl)ethanol(III-b) (S)-2-(trimethylsilyl)-1-((S)-1-pyrrolidine-2-yl)ethanol(Va) (R)-2,2-diphenyl-1-((S)-pyrrolidine-2-yl)ethanol(VII-a) (S)-2,2-diphenyl-1-((R)-pyrrolidine-2-yl)ethanol(VII-b) (R)-2-(4-nitrophenyl)-1-((S)-pyrrolidine-2-yl)ethanol(IX-a) (S)-2-(4-nitrophenyl)-1-((R)-pyrrolidine-2-yl)ethanol(IX-b) (R)-(9H-fluorolene-9-yl)((S)-pyrrolidine-2-yl)methanol(XI-a) (S)-2-Tosyl-1-((S)-1-Tritylpyrrolidine-2-yl)ethanol(XIII-a) (R)-2-Tosyl-1-((R)-1-Tritylpyrrolidine-2-yl)ethanol(XIII-b) It is selected from one of the following.

[0129] Chiral reagents react with nucleic acids or modified nucleic acids that act as asymmetric auxiliary groups. Nucleoside 3'-phosphoramidite derivatives, which are intermediates for producing stereocontrolled phosphorus atom-modified oligonucleotide derivatives, are obtained by chiral reagents that react with nucleic acids or modified nucleic acids.

[0130] A second aspect of the present invention relates to a nucleoside 3'-phosphoramidite derivative represented by formula (Va) or (Vb). The compounds of formula (Va) and (Vb) are known as monomers used in the synthesis of ologonucleotide derivatives. These compounds are also known as oxazaphosphoridine monomers. The sugar moiety of the compound represented by formula (Vb) is known as BNA and LNA (R 3 (When it is a methylene group).

[0131] [ka]

[0132] In equations (Va) and (Vb), G 1 ~G 4 This is the same as above, G 5 B is a protecting group for the hydroxyl group, and Bs is a group selected from the groups represented by formulas (VI) to (XI) or their derivatives.

[0133] [ka]

[0134] Examples of Bs include adenine, thymine, cytosine, guanine, uracil, 5-methylcytosine, or their derivatives.

[0135] R 2 These are hydrogen, -OH, -SH, and -NR d R d -N3, halogen, alkyl, alkenyl, alkynyl, alkyl-Y 1 -, Alkenil-Y 1 -, Alkinyl-Y 1 -, Aryl-Y 1 -, heteroaryl-Y 1 -, -OR b , or -SR b And in this case, R b This is the blocking section. Y1 O, NR d It is S, or Se. R d These are independently hydrogen, alkyl, alkenyl, alkynyl, aryl, acyl, substituted silyl, carbamate, and -P(O)(R e )2, or -HP(O)(R e ) R e These are independently hydrogen, alkyl, aryl, alkenyl, alkynyl, and alkyl-Y 2 -, Alkenil-Y 2 -, Alkinyl-Y 2 -, Aryl-Y 2 -, or heteroaryl-Y 2 -, or, Na + Li + , or K + It is a positive ion. Y 2 O, NR d , or S. A preferred example of alkyl is C 1-10 A preferred example of an alkyl group is an alkenyl. 2-10 Alkenyls are a preferred example of alkinyls, and C 2-10 Alkinyl, and a preferred example of aryl is C 6-14 It is an aryl, and a preferred example of a heteroaryl is C 6-14 It is a heteroaryl compound.

[0136] R 3 These are groups represented by -CH2-, -(CH2)2-, -CH2NH-, or -CH2N(CH3)-.

[0137] G 5 Examples include trityl, 4-monomethoxytrityl, 4,4'-dimethoxytrityl, 4,4',4”-trimethoxytrityl, 9-phenylxanthin-9-yl (Pixyl), and 9-(p-methoxyphenyl)xanthin-9-yl (MOX).

[0138] Bs is adenine, thymine, cytosine, guanine, or a derivative thereof. Bs is a nucleic acid base or a modified nucleic acid base. Examples of derivatives are disclosed in Japanese Patent Application Publication No. 2005-89441 and are as follows:

[0139] [ka]

[0140] In the above equation, R 8 ~R 10 Each of them independently, C 1-10 Alkyl, C6-C 10 Ariel, C6-C 10 Aralkil, or C6-C 10 It is an aryloxyalkyl. 8 Preferred examples include methyl, isopropyl, phenyl, benzyl, and phenoxymethyl. 9 and R 10 A preferred example is C 1-4 It is an alkyl group.

[0141] A preferred embodiment of the second aspect is that the nucleoside 3'-phosphoramidite derivative is represented by formula (Va') or (Vb').

[0142] [ka]

[0143] In equations (Va') and (Vb'), G 1 , G 2 , G 5 , Bs, R 2 , and R 3 This is the same as above. Nucleoside 3'-phosphoramidite derivatives are chiral monomers used to produce stereocontrolled phosphorus-modified nucleotide and oligonucleotide derivatives.

[0144] Preferred examples of nucleoside 3'-phosphoramidite derivatives are represented by formulas 1a, 1b, 2a, 2b, 3a, 3b, 4a, 4b, 5a, 5b, 6a, 6b, 7a, 7b, 8a, 8b, 9a, 9b, 10a, 10b, 11a, 11b, 12a, 12b, 13a, 13b, 14a, 14b, 15a, 15b, 16a, 16b, 17a, 17b, 18a, 18b, 19a, 19b, 20a, 20b, 21a, 21b, 22a, 22b, 23a, 23b, or 24a. These formulas are described in the experimental section.

[0145] DMTr represents the 4,4'-dimethoxytrityl group, and TOM represents the triisopropylsiloxymethyl group.

[0146] An example of using a nucleoside 3'-phosphoramidite derivative is disclosed, for example, in Japanese Patent Application Publication No. 2005-89441. By repeating the condensation and deprotection steps, it is possible to lengthen the chain of the oligonucleotide derivative, as disclosed therein.

[0147] The formula for such an oligonucleotide derivative is shown in formula (X).

[0148] [ka]

[0149] In equation (X), X is a sulfide (=S), C 1-3 Alkyl, C 1-3 Alkoxy, C 1-3 Alkylthio, C6-C 10 Ariel, C6-C 10 Aralkil, or C6-C 10 This represents an aryloxyalkyl group. Preferably, X represents a sulfide (=S). "n" is an integer representing 1 to 150, 1 to 100, 1 to 50, or 1 to 30. "n" may preferably be 2 to 100, preferably 10 to 100, preferably 10 to 50, and more preferably 15 to 30.

[0150] A third aspect of the present invention relates to a method for synthesizing stereocontrolled phosphorus atom-modified oligonucleotide derivatives. The first step is to react a molecule containing an achiral H-phosphonate moiety, a first activating reagent, and a chiral reagent or a salt thereof to form a monomer. The chiral reagent has chemical formula (I) or (I'), and its monomer can be represented by formula (formula)(Va), (Vb), (Va'), or (Vb'). The monomer reacts with a second activating reagent and a nucleoside to form a concentrated intermediate. The next step is to convert the concentrated intermediate into a nucleic acid containing a chiral X-phosphonate moiety. This method is based on the disclosure in International Publication No. 2010 / 064146, where the basic steps are disclosed as pathways A and B. In this method, the chiral reagent of the present invention is used.

[0151] A first scheme for the synthesis of chiral oligonucleotides.

[0152] [ka]

[0153] Activation process The achiral H-phosphonate moiety is treated with a first activating reagent to form a first intermediate. In one embodiment, the first activating reagent is added to the reaction mixture during the condensation step. The use of the first activating reagent depends on the reaction conditions, such as the solvent used in the reaction. Examples of the first activating reagent include phosgene, trichloromethyl chloroformate, bis(trichloromethyl) carbonate (BTC), oxalyl chloride, Ph3PCl2, and (PhO)3PCl. 2、These are N,N'-bis(2-oxo-3-oxazolidinyl)phosphinate chloride (BopCl), 1,3-dimethyl-2-(3-nitro-1,2,4-triazole-1-yl)-2-pyrrolidine-1-yl-1,3,2-diazaphosphoridinium hexafluorophosphate (MNTP), or 3-nitro-1,2,4-triazole-1-yl-tris(pyrrolidine-1-yl)phosphonium hexafluorophosphate (PyNTP).

[0154] An example of the achiral H-phosphonate moiety is the compound shown in the scheme above. DBU represents 1,8-diazabicyclo[5.4.0]undeca-7-ene. + DBU may be, for example, an ammonium ion, an alkylammonium ion, a heteroaromatic iminium ion, or a heterocyclic iminium ion, all of which are primary, secondary, tertiary, or quaternary, or monovalent metal ions.

[0155] Reaction with chiral reagents Following the first activation step, the activated achiral H-phosphonate moiety reacts with a chiral reagent (represented by formula (I) or (I')) to form a chiral intermediate of formula (Va), (Vb), (Va'), or (Vb').

[0156] Stereospecific condensation process A chiral intermediate of formula Va((Vb), (Va'), or (Vb')) is treated with a second activating agent and a nucleoside to form a concentrated intermediate. The nucleoside may be coagulated. Examples of the second activating agent are 4,5-dicyanoimidazole (DCI), 4,5-dichloroimidazole, 1-phenylimidazolium triflate (PhIMT), benzimidazolium triflate (BIT), benztriazole, 3-nitro-1,2,4-triazole (NT), tetrazole, 5-ethylthiotetrazole (ETT), 5-benzylthiotetrazole (BTT), 5-(4-nitrophenyl)tetrazole, N-cyanomethylpyrrolidinium triflate (CMPT), N-cyanomethylpiperidinium triflate, and N-cyanomethyldimethylammonium triflate. The chiral intermediate of formula Va((Vb), (Va'), or (Vb')) can be isolated as a monomer. Typically, the chiral intermediate of Va((Vb), (Va'), or (Vb')) is not isolated but is reacted with a nucleoside or modified nucleoside in the same pot to provide a concentrated intermediate, the chiral phosfit compound. In other embodiments, when this method is carried out by solid-phase synthesis, the solid support containing the compound is filtered to separate by-products, impurities, and / or reagents.

[0157] Capping process If the final nucleic acid is larger than a dimer, the unreacted -OH portion can be capped with a blocking group, and the chiral auxiliaries in the compound can also be capped with a blocking group to form a capped concentrated intermediate. If the final nucleic acid is a dimer, the capping step is not necessary.

[0158] Modification process The compound is modified by reaction with an electrophile. The modification step can be carried out on a capped concentrated intermediate. In some embodiments of this method, the modification step is carried out using a sulfur electrophile, a selenium electrophile, or a boronating agent. Preferred examples of the modification step are oxidation and sulfidation steps.

[0159] In some embodiments of this method, the sulfur electrophile is a compound having one of the following formulas: S8 (Formula B), Z 1 -SSZ 2 , or Z 1 -SVZ 2 .

[0160] Z 1 and Z 2 These are independently alkyl, aminoalkyl, cycloalkyl, heterocyclic, cycloalkylalkyl, heterocycloalkyl, aryl, heteroaryl, alkyloxy, aryloxy, heteroaryloxy, acyl, amide, imide, or thiocarbonyl, or Z 1 and Z 2 These elements combine to form a 3- to 8-membered alicyclic or heterocyclic ring, which may be substituted or unsubstituted; V is SO2, O, or NR f And; R f These are hydrogen, alkyl, alkenyl, alkynyl, or aryl atoms.

[0161] In some embodiments of this method, the sulfur electrophile is a compound of the following formulas A, B, C, D, E, or F:

[0162] [ka]

[0163] In some embodiments of this method, the selenium electrophile is a compound having one of the following formulas: Se (formula G), Z 3 -Se-Se-Z 4 , or Z 3 -Se-VZ 4

[0164] Z 3 and Z 4These are independently alkyl, aminoalkyl, cycloalkyl, heterocyclic, cycloalkylalkyl, heterocycloalkyl, aryl, heteroaryl, alkyloxy, aryloxy, heteroaryloxy, acyl, amide, imide, or thiocarbonyl, or Z 3 and Z 4 These elements combine to form a 3- to 8-membered alicyclic or heterocyclic ring, which may be substituted or unsubstituted; V is SO2, S, O, or NR f And; R f These are hydrogen, alkyl, alkenyl, alkynyl, or aryl atoms.

[0165] In some embodiments of this method, the selenium electrophile is a compound of formula G, H, I, J, K, or L.

[0166] [ka]

[0167] In some embodiments of this method, the boronating agent is borane-N,N-diisopropylethylamine (BH3DIPEA), borane-pyridine (BH3Py), borane-2-chloropyridine (BH3CPy), borane-aniline (BH3An), borane-tetrahydrofuran (tetrahydrofiirane) (BH3THF), or borane-dimethyl sulfide (BH3Me2S).

[0168] In some embodiments of this method, the modification step is an oxidation step. The oxidation step is disclosed, for example, in Japanese Patent Application Publication No. 2010-265304 and International Publication No. 2010 / 064146.

[0169] Chain extension cycle and deprotection process The capped concentrated intermediate is deblocked to remove the blocking group at the 5' end of the growing nucleic acid chain to obtain the compound. The compound is optionally subjected to another chain extension cycle to form a concentrated intermediate, a capped concentrated intermediate, a modified and capped concentrated intermediate, and a 5'-deprotected, modified and capped intermediate. After at least one round of chain extension cycles, the 5'-deprotected, modified and capped intermediate is further deblocked by removing chiral auxiliary ligands and other protecting groups, such as nucleic acid bases, modified nucleic acid bases, sugars, and modified sugar protecting groups, to obtain the nucleic acid. In other embodiments, the nucleoside containing the 5'-OH moiety is an intermediate from a previous chain extension cycle as described herein. In yet another embodiment, the nucleoside containing the 5'-OH moiety is an intermediate obtained from another known nucleic acid synthesis method. In embodiments using a solid support, the phosphorus-modified nucleic acid is then cleaved from the solid support. In certain embodiments, nucleic acids are left attached to a solid support for purification purposes and then cleaved from the solid support after purification.

[0170] Based on this method, it is possible to use stable, commercially available materials as starting materials. Using achiral starting materials, it is possible to generate stereocontrolled phosphorus atom-modified oligonucleotide derivatives.

[0171] As shown in the examples, the method of the present invention does not cause degradation under the deprotection step. Furthermore, this method does not require a special capping agent to produce phosphorus atom-modified oligonucleotide derivatives.

[0172] A fourth aspect of the present invention relates to a method for synthesizing stereocontrolled phosphorus atom-modified oligonucleotide derivatives using chiral monomers. The first step is to react a nucleoside 3'-phosphoramidite derivative represented by (Va), (Vb), (Va'), or (Vb') with a second activating reagent and the nucleoside to form a concentrated intermediate. The second step is to convert the concentrated intermediate into a nucleic acid containing a chiral X-phosphonate moiety.

[0173] The second scheme relates to the synthesis of chiral oligos using monomers of the formula Va((Vb), (Va'), or (Vb')). The second scheme is based on the method disclosed in Japanese Patent Application Publication No. 2005-89441.

[0174] [ka]

[0175] The detailed conditions of the above scheme are the same as those of the first scheme. The starting materials of formula Va(Vb), particularly formula Va'(or Vb'), are chemically stable. As shown in the examples, the method of the present invention does not cause decomposition under the deprotection step. Furthermore, the method does not require special capping agents to produce phosphorus atom-modified oligonucleotide derivatives.

[0176] The mechanism for removing the auxiliary agent is shown below:

[0177] [ka]

[0178] In the scheme above, Nu represents the nucleophile. The mechanism above is considered to be different from previous mechanisms for the removal of auxiliary agents. [Examples]

[0179] Abbreviation ac: acetyl bz: benzoyl CSO:(1S)-(+)-(10-camphorsulfonyl)oxaziridine DBU:1,8-Diazabicyclo[5.4.0]Undeca-7-En DCA: Dichloroacetic acid DCM: Dichloromethane, CH2Cl2 DMTr:4,4'-Dimethoxytrityl Tr: Trityl, Triphenylmethyl MeIm: N-methylimidazole NIS:N-iodosuccinimide pac: Phenoxyacetyl Ph: Phenyl PhIMT:N-phenylimidazolium triflate POS:3-phenyl-1,2,4-dithiazolin-5-one TBS: tert-butyldimethylsilyl TBDPS: tert-butyldiphenylsilyl TOM: Triisopropylsiloxymethyl TFA: Trifluoroacetic acid [Examples]

[0180] (S)-Tritylpyrrolidine-2-carbaldehyde(Ia).

[0181] [ka] Compound Ia was synthesized from L-proline according to the procedure described in the literature (Guga, P. Curr. Top. Med. Chem. 2007, 7, 695-713). [Examples]

[0182] (R)-1-Tritylpyrrolidine-2-carbaldehyde(Ib).

[0183] [ka] Compound Ib was synthesized from D-proline using the same method as compound Ia. [Examples]

[0184] (S)-2-(methyldiphenylsilyl)-1-((S)-1-tritylpyrrolidine-2-yl)ethanol(II-a).

[0185] [ka] A THF solution of methyldiphenylsilylmethylmagnesium chloride, prepared from chloromethyldiphenylmethylsilane (4.02 g, 16.3 mmol) and magnesium (402 mg, 16.3 mmol) in THF (14 mL), was added to a THF (30 mL) solution of Ia (2.79 g, 8.14 mmol) while cooling with ice. After stirring for 1.5 hours while cooling with ice, the mixture was warmed to room temperature and stirred for 30 minutes. A saturated NH4Cl aqueous solution (100 mL) was added to the reaction mixture at 0°C, and the mixture was extracted three times with diethyl ether (100 mL). The combined extract was dried over Na2SO4, filtered, and concentrated under reduced pressure. The residue was subjected to silica gel chromatography to obtain II-a as a colorless foam (3.91 g, 87%). 1 H NMR(300MHz,CDCl3)δ 7.48-7.08(25H,m),4.33-4.23(1H,m),3.16-2.89(3H,m),2.84(1H,brs),1.70-1.54(1H,m),1.35(1H,dd,J=14. 7,6.3Hz),1.10(1H,dd,J=14.7,8.1Hz),1.18-1.05(1H,m),1.04-0.90(1H,m),0.34(3H,s),-0.17--0.36(1H,m). [Examples]

[0186] (S)-2-(methyldiphenylsilyl)-1-((S)-pyrrolidine-2-yl)ethanol(III-a).

[0187] [ka]

[0188] II-a (3.91 g, 7.06 mmol) was dissolved in a solution of 3% DCA mixed in DCM (70 mL) and stirred at room temperature for 10 minutes. 1 M NaOH (200 mL) was added to the mixture, and extraction was performed three times with DCM (100 mL). The combined extract was dried over Na2SO4, filtered, and concentrated under reduced pressure. The residue was subjected to silica gel chromatography to obtain III-a as a pale yellow oily substance (1.99 g, 90%). 1 H NMR(300MHz,CDCl3)δ 7.57-7.52(5H,m),7.38-7.33(5H,m),3.77(1H,ddd,J=8.9,5.4,3.5Hz),3.01(1H,dt,J=7.4,3.6Hz),2.97-2.79( 2H,m),2.27(2H,brs),1.76-1.53(4H,m),1.38(1H,dd,J=15.0,9.0Hz),1.24(1H,dd,J=15.0,5.4Hz),0.65(3H,s); 13 C NMR(100.4MHz,CDCl3)δ 137.4,137.1,134.6,134.5,129.1,127.8,69.5,64.1,47.0,25.8,24.0,19.6,-3.4.MALDI TOF-MS m / z C 19 H 26 Calculated value for NOSi [M+H] + 312.18, measured value 312.06. [Examples]

[0189] (R)-2-(methyldiphenylsilyl)-1-((R)-1-tritylpyrrolidine-2-yl)ethanol(II-b).

[0190] [ka]

[0191] Compound II-b was obtained in the same manner as compound II-a, but by using Ib instead of Ia. 1H NMR(300MHz,CDCl3)δ 7.48-7.12(25H,m),4.33-4.24(1H,m),3.16-2.89(3H,m),2.86(1H,brs),1.69-1.52(1H,m),1.35(1H,dd,J=14. 4,6.0Hz),1.10(1H,dd,J=14.4,8.4Hz),1.18-1.05(1H,m),1.03-0.89(1H,m),0.33(3H,s),-0.19--0.39(1H,m); 13 C NMR(75.5MHz,CDCl3)δ MALDI TOF-MS m / z C 38 H 40 Calculated value for NOSi [M+H] + 554.29, actual measured value 554.09. [Examples]

[0192] (R)-2-(methyldiphenylsilyl)-1-((R)-1-pyrrolidine-2-yl)ethanol(III-b).

[0193] [ka]

[0194] Compound III-b was obtained by using II-b instead of II-a in the same manner as compound III-a. 1 H NMR(300MHz,CDCl3)δ 7.58-7.52(5H,m),7.38-7.33(5H,m),3.78(1H,ddd,J=9.0,5.1,3.6Hz),3.00(1H,dt,J=7.4,3.3Hz),2.97-2.78( 2H,m),2.19(2H,brs),1.76-1.53(4H,m),1.38(1H,dd,J=14.6,9.0Hz),1.24(1H,dd,J=14.6,5.1Hz),0.66(3H,s); 13C NMR(75.5MHz,CDCl3)δ 137.5,137.1,134.5,134.4,129.0,127.7,69.2,64.2,46.9,25.8,24.0,19.7,-3.4.MALDI TOF-MS m / z C 19 H 26 Calculated value for NOSi [M+H] + 312.18, measured value 312.09. [Examples]

[0195] (S)-2-(trimethylsilyl)-1-((S)-1-tritylpyrrolidine-2-yl)ethanol(IV-a).

[0196] [ka]

[0197] Compound IV-a was obtained in the same manner as compound II-a, by using chloromethyltrimethylsilane instead of chloromethyldiphenylmethylsilane. 1 H NMR(300MHz,CDCl3)δ 7.58-7.51(5H,m),7.31-7.14(10H,m),4.13(1H,dt,J=7.5,3.0Hz),3.39-3.31(1H,m),3.20-2.99(2H,m),2.84(1H,s) MALDI TOF-MS m / z C 28 H 36 Calculated value for NOSi [M+H] + 430.26, measured value 430.09. [Examples]

[0198] (S)-2-(trimethylsilyl)-1-((S)-1-pyrrolidine-2-yl)ethanol(Va).

[0199] [ka]

[0200] Compound Va was obtained in the same manner as compound III-a, but by using IV-a instead of II-a. 1 H NMR(300MHz,CDCl3)δ 3.76(1H,ddd,J=8.8,5.7,3.3Hz),3.08(1H,dt,J=7.8,3.3Hz),3.02-2.87(2H,m),2.48(2H,br s),1.81-1.58(4H,m),0.83(1H,dd,J=14.7,8.7Hz),0.68(1H,dd,J=14.7,6.0Hz),0.05(9H,s); 13 C NMR(75.5MHz,CDCl3)δ 69.6,64.3,46.9,25.8,23.9,22.0,-0.8.MALDI TOF-MS m / z C9H 22 Calculated value for NOSi [M+H] + 188.15, actual measured value 188.00. [Examples]

[0201] (R)-2,2-diphenyl-1-((S)-1-tritylpyrrolidine-2-yl)ethanol(VI-a).

[0202] [ka]

[0203] To a solution of diphenylmethane (6.7 mL, 40 mmol) in anhydrous THF (36 mL), n-BuLi (1.67 M solution in hexane, 24 mL, 40 mmol) was added dropwise at room temperature, and the mixture was stirred for 1 hour. To the mixture, Ia (3.41 g, 10 mmol), dried by repeated co-evaporation with toluene in anhydrous THF (40 mL), was slowly added at 0°C, and stirring was continued for 45 minutes. Next, saturated aqueous NH4Cl (100 mL) and Et2O (100 mL) were added, the organic layer was separated, and the aqueous layer was extracted with Et2O (2 × 100 mL). The organic layers were combined, dried over Na2SO4, filtered, and concentrated under reduced pressure. The residue was purified by chromatography on silica gel to obtain VI-a (1.41 g, 28%) as a white foam. 1 H NMR(300MHz,CDCl3)δ 7.45-7.01(23H,m),6.67-6.61(2H,m),4.80(1H,d,J=10.8Hz),3.63(1H,d,J=10.8Hz),3.36-3.27(1H,m), 3.23-3.09(1H,m),3.02-2.89(1H,m),2.66(1H,s),1.90-1.75(1H,m),1.32-1.04(2H,m),0--0.18(1H,m). [Examples]

[0204] (R)-2,2-diphenyl-1-((S)-pyrrolidine-2-yl)ethanol(VII-a).

[0205] [ka]

[0206] Compound VII-a was obtained in the same manner as compound III-a, but by using VI-a instead of II-a. 1H NMR(300MHz,CDCl3)δ 7.44-7.38(2H,m),7.33-7.14(8H,m),4.46(1H,dd,J=9.9,3.3Hz),3.91(1H,d,J= 9.9Hz),3.02-2.88(2H,m),2.81-2.69(1H,m),2.52(2H,brs),1.88-1.56(4H,m); 13 C NMR(75.5MHz,CDCl3)δ 142.3,142.0,128.6,128.5,128.4,128.2,126.5,126.4,73.5,60.1,55.8,46.6,25.8,23.4.MALDI TOF-MS m / z C 18 H 22 Calculated value for NO [M+H] + 268.17, measured value 268.06. [Examples]

[0207] (S)-2,2-diphenyl-1-((R)-1-tritylpyrrolidine-2-yl)ethanol(VI-b).

[0208] [ka]

[0209] Compound VI-b was obtained in the same manner as compound VI-a, but by using Ib instead of Ia. 1 H NMR(300MHz,CDCl3)δ 7.44-7.37(6H,m),7.30-7.01(17H,m),6.66-6.61(2H,m),4.80(1H,d,J=10.8Hz),3.63(1H,d,J=10.8Hz),3.36-3.28 (1H,m),3.22-3.09(1H,m),3.01-2.89(1H,m),2.66(1H,s),1.90-1.75(1H,m),1.29-1.04(2H,m),0.00--0.19(1H,m); 13C NMR(75.5MHz,CDCl3)δ MALDI TOF-MS m / z C 37 H 36 Calculated value for NO [M+H] + 510.28, measured value 510.11. [Examples]

[0210] (S)-2,2-diphenyl-1-((R)-pyrrolidine-2-yl)ethanol(VII-b).

[0211] [ka]

[0212] Compound VII-b was obtained in the same manner as compound VII-a, but by using VI-b instead of VI-a. 1 H NMR(300MHz,CDCl3)δ 7.45-7.14(10H,m),4.45(1H,dd,J=9.9,3.3Hz),3.91(1H,d,J=9.9Hz),3.00-2.89(2H,m),2.82-2.71(1H,m),2.40(2H,brs),1.87-1.55(4H,m); 13 C NMR(75.5MHz,CDCl3)δ 142.3,142.0,128.5,128.3,128.1,126.3,126.2,73.4,60.1,55.9,46.5,25.8,23.5.MALDI TOF-MS m / z C 18 H 22 Calculated value for NO [M+H] + 268.17, measured value 268.03. [Examples]

[0213] (R)-2-(4-nitrophenyl)-1-((S)-1-tritylpyrrolidine-2-yl)ethanol(VIII-a).

[0214] [ka]

[0215] Compound VIII-a was obtained in the same manner as compound VI-a, by using "4-nitrobenzyl chloride" instead of "diphenylmethane". 1 H NMR(300MHz,CDCl3)δ 8.09-8.03(2H,m),7.49-7.43(6H,m),7.28-7.09(11H,m),4.23(1H,dd d,J=8.3,5.6,3.0Hz),3.43-3.33(1H,m),3.23-3.11(1H,m),3.07-2.9 6(1H,m),2.83(1H,brs),2.74(1H,dd,J=13.8,8.4Hz),2.49(1H,dd,J=13.8,5.1Hz),1.83-1.67(1H,m),1.41-1.17(2H,m),0.27-0.08(1H,m); 13 C NMR(75.5MHz,CDCl3)δ 147.3,146.3,144.3,129.8,129.6,127.5,126.3,123.4,77.9,74.8,63.5,53.2,39.5,25.0,24.9.MALDI TOF-MS m / z C 31 H 31 Calculated value for N2O3 [M+H] + 479.23, measured value 479.08. [Examples]

[0216] (R)-2-(4-nitrophenyl)-1-((S)-pyrrolidine-2-yl)ethanol(IX-a)

[0217] [ka]

[0218] Compound IX-a was obtained in the same manner as compound VII-a, but by using VIII-a instead of VI-a. 1 H NMR(300MHz,CDCl3)δ 8.15(2H,d,J=8.7Hz),7.42(2H,d,J=8.7Hz),3.86-3.79(1H,m),3.16-3.07(1H,m),2.99-2.68(6H,m),1.84-1.68(4H,m); 13 C NMR(75.5MHz,CDCl3)δ 147.4,146.2,129.9,123.2,72.4,62.0,46.6,40.4,25.7,24.4.MALDI TOF-MS m / z C 12 H 17 Calculated value for N2O3 [M+H] + 237.12, measured value 237.01. [Examples]

[0219] (S)-2-(4-nitrophenyl)-1-((R)-1-tritylpyrrolidine-2-yl)ethanol(VIII-b).

[0220] [ka]

[0221] Compound VIII-b was obtained in the same manner as compound VIII-a, but by using Ib instead of Ia. 1 H NMR(300MHz,CDCl3)δ 8.09-8.04(2H,m),7.49-7.43(6H,m),7.28-7.09(11H,m),4.22(1H,dd d,J=8.4,5.6,3.0Hz),3.43-3.33(1H,m),3.24-3.10(1H,m),3.08-2.9 4(1H,m),2.81(1H,brs),2.75(1H,dd,J=14.0,8.1Hz),2.49(1H,dd,J=14.0,5.1Hz),1.81-1.67(1H,m),1.40-1.16(2H,m),0.26-0.09(1H,m); 13C NMR(75.5MHz,CDCl3)δ 147.3,144.3,129.8,129.6,129.4,126.3,123.5,77.9,74.8,63.5,53.2,39.5,25.0,24.9.MALDI TOF-MS m / z C 31 H 31 Calculated value for N2O3 [M+H] + 479.23, measured value 479.08. [Examples]

[0222] (S)-2-(4-nitrophenyl)-1-((R)-pyrrolidine-2-yl)ethanol(IX-b).

[0223] [ka]

[0224] Compound IX-b was obtained in the same manner as compound IX-a, but by using VIII-b instead of VIII-a. 1 H NMR(300MHz,CDCl3)δ 8.19-8.13(2H,m),7.45-7.39(2H,m),3.83(1H,ddd,J=7.7,5.4,3.9Hz),3.14(1H,dt,J=7.7,3. 9Hz),3.01-2.87(2H,m),2.83(1H,d,J=3.3Hz),2.81(1H,s),2.62(2H,brs),1.79-1.72(4H,m); 13 C NMR(75.5MHz,CDCl3)δ 147.3,146.5,130.0,123.5,72.7,61.7,46.7,40.1,25.8,24.2.MALDI TOF-MS m / z C 12 H 17 Calculated value for N2O3 [M+H] + 237.12, measured value 237.02. [Examples]

[0225] (R)-(9H-fluoren-9-yl)((S)-1-tritylpyrrolidine-2-yl)methanol(Xa).

[0226] [ka]

[0227] Compound Xa was obtained in the same manner as compound VI-a, by using fluorene instead of diphenylmethane. 1 1H NMR (300MHz, CDCl3)δ 7.70(1H,d,J=7.5Hz),7.66(1H,d,J=7.8Hz),7.55(2H,d,J=7.5Hz),7.44-7. 09(18H,m),6.87-6.62(1H,m),4.55-4.48(1H,m),4.06(1H,d,J=7.5Hz),3.4 3-3.34(1H,m),3.18-3.06(1H,m),2.98-2.88(1H,m),2.85(1H,brs),1.42-1 .24(1H,m),1.18-1.04(1H,m),0.53-0.39(1H,m),-0.02--0.20(1H,m);MALDI TOF-MS m / z C 37 H 34 Calculated value for NO [M+H] + 508.26, measured value 508.12. [Examples]

[0228] (R)-(9H-fluorolene-9-yl)((S)-pyrrolidine-2-yl)methanol(XI-a).

[0229] [ka]

[0230] Compound XI-a was obtained in the same manner as compound III-a, but by using Xa instead of II-a. 1H NMR(300MHz,CDCl3)δ 7.76(2H,d,J=7.5Hz),7.68(2H,t,J=8.0Hz),7.43-7.35(2H,m),7.34-7.25(2H,m),4.28(1H,d,J=6.3Hz),4.03(1H,dd,J=6.5 MALDI TOF-MS m / z C 18 H 20 Calculated value for NO [M+H] + 266.15, measured value 266.04. [Examples]

[0231] (S)-2-Tosyl-1-((S)-1-Tritylpyrrolidine-2-yl)ethanol(XII-a).

[0232] [ka]

[0233] Compound XII-a was obtained in the same manner as compound II-a, by using chloromethyl p-tolylsulfone instead of chloromethyldiphenylmethylsilane. 1 H NMR(600MHz,CDCl3)δ 7.66(2H,d,J=8.4Hz),7.48-7.44(6H,m),7.35(2H,d,J=7.2Hz),7.21-7.13(9H,m),4.39-4.36(1H,m),3.33(1H,s),3.24-3.20(1H, m),3.19-3.10(2H,m),2.98-2.92(2H,m),2.49(3H,s),1.55-1.49(1H,m),1.33-1.26(1H,m),1.12-1.04(1H,m),0.22-0.14(1H,m); 13C NMR(150.9MHz, CDCl3)δ 144.6,144.5,136.3,129.9,129.5,128.1,127.5,126.2,78.0,69.1,63.9,60.2,52.6,25.5,24.7,21.7. [Examples]

[0234] (S)-2-Tosyl-1-((S)-1-Tritylpyrrolidine-2-yl)ethanol(XIII-a).

[0235] [ka]

[0236] Compound XIII-a was obtained by using XII-a instead of II-a in the same manner as compound III-a. 1 H NMR(600MHz,CDCl3)δ 7.82(2H,d,J=8.4Hz),7.37(2H,d,J=8.4Hz),4.01(1H,ddd,J=12.0,5.1,3.0Hz),3.32(1H,dd,J=14.4,3.0Hz),3.25(1H,dd,J=1 4.4,9.0Hz),3.16(1H,dt,J=7.8,5.1Hz),2.90-2.82(2H,m),2.46(3H,s),2.04(2H,brs),1.78-1.63(3H,m),1.62-1.55(1H,m); 13 C NMR(150.9MHz,CDCl3)δ 144.5,136.7,129.7,127.7,67.4,61.8,60.1,46.7,25.7,21.4.MALDI TOF-MS m / z C 13 H 20 Calculated value for NO3S [M+H] + 270.12, measured value 270.04. [Examples]

[0237] (R)-2-Tosyl-1-((R)-1-Tritylpyrrolidine-2-yl)ethanol(XII-b).

[0238] [ka]

[0239] Compound XII-b was obtained in the same manner as compound XII-a, but by using Ib instead of Ia. 1 H NMR(600MHz,CDCl3)δ 7.66(2H,d,J=8.4Hz),7.47-7.44(6H,m),7.35(2H,d,J=7.8Hz),7.21-7.13(9H,m),4.37(1H,dt,J=8.6,2.4Hz),3.33(1H,s),3.23-3.20 (1H,m),3.19-3.12(2H,m),2.98-2.92(2H,m),2.49(3H,s),1.56-1.49(1H,m),1.32-1.26(1H,m),1.11-1.03(1H,m),0.23-0.15(1H,m); 13 C NMR(150.9MHz, CDCl3)δ 144.6,144.5,136.3,129.9,129.6,128.1,127.6,126.2,78.0,69.1,63.9,60.2,52.6,25.5,24.7,21.7. [Examples]

[0240] (R)-2-Tosyl-1-((R)-1-Tritylpyrrolidine-2-yl)ethanol(XIII-b).

[0241] [ka]

[0242] Compound XIII-b was obtained by using XII-b instead of XII-a in the same manner as compound XIII-a. 1H NMR(600MHz,CDCl3)δ 7.82(2H,d,J=8.4Hz),7.37(2H,d,J=8.4Hz),4.01(1H,ddd,J=9.0,5.1,3.0Hz),3.32(1H,dd,J=14.4,3.0Hz),3.25(1H,dd,J=1 4.4,9.0Hz),3.17(1H,dt,J=7.2,5.1Hz),2.89-2.83(2H,m),2.46(3H,s),2.04(2H,brs),1.79-1.64(3H,m),1.62-1.55(1H,m); 13 C NMR(150.9MHz,CDCl3)δ 144.8,136.6,129.8,127.9,67.7,61.8,60.1,46.8,25.9,25.8,21.6.MALDI TOF-MS m / z C 13 H 20 Calculated value for NO3S [M+H] + 270.12, measured value 270.05. [Examples]

[0243] Oxazaphosphoridine monomer 3a.

[0244] [ka]

[0245] III-a (560 mg, 1.80 mmol) was dried by repeated co-evaporation with anhydrous toluene and dissolved in anhydrous diethyl ether (0.90 mL) under argon. N-methylmorpholine (400 mL, 3.60 mmol) was added to the solution, and the resulting solution was added dropwise to a solution of PCl3 (160 mL, 1.80 mmol) in anhydrous diethyl ether (0.90 mL) under argon at 0°C with stirring. Next, the mixture was warmed to room temperature and stirred for 30 minutes. The resulting N-methylmorpholine hydrochloride was removed by filtration under nitrogen, and the filtrate was concentrated to dryness under reduced pressure to obtain the crude 2-chloro-1,3,2-oxazaphosphoridine derivative. The crude product was dissolved in freshly distilled THF (3.6 mL) to prepare a 0.5 M solution, which was used to synthesize the nucleoside 3'-O-oxazaphosphoridine without further purification.

[0246] 5'-O-(DMTr)-2-N-(phenoxyacetyl)-6-O-(cyanoethyl)guanosine (636 mg, 0.84 mmol) was dried by repeated co-evaporation with anhydrous toluene and dissolved in freshly distilled THF (2.5 mL) under argon. Et3N (0.58 mL, 4.2 mmol) was added, and the mixture was cooled to -78°C. A 0.5 M solution of the corresponding crude 2-chloro-1,3,2-oxazaphosphoridine derivative in freshly distilled THF (3.6 mL, 1.80 mmol) was added dropwise by syringe, and the mixture was stirred at room temperature for 15 minutes. Next, saturated aqueous NaHCO3 (70 mL) and CHCl3 (70 mL) were added, the organic layer was separated, and washed with saturated aqueous NaHCO3 (2 × 70 mL). The combined aqueous layer was back-extracted with CHCl3 (70 mL). The organic layers were combined, dried over Na2SO4, filtered, and concentrated under reduced pressure. The residue was purified by chromatography using silica gel to obtain 3a (829 mg, 90%) as a white foam. 1H NMR(300MHz,CDCl3)δ 8.77(1H,brs),7.99(1H,s),7.54-6.98(24H,m),6.81-6.73(4H,m),6.35(1H,dd,J=8.0,6.3H z),4.89-4.73(4H,m),4.68(2H,brs),4.05-3.98(1H,m),3.75(6H,s),3.62-3.46(1H,m),3.4 1-3.20(3H,m),3.18-3.04(1H,m),3.08(2H,t,J=6.6Hz),2.58-2.36(2H,m),1.94-1.59(2H,m ),1.56(1H,dd,J=15.0,8.7Hz),1.43(1H,dd,J=15.0,5.7Hz),1.33-1.16(2H,m),0.62(3H,s); 31 P NMR(121.5MHz, CDCl3)δ 153.5(1P,s). [Examples]

[0247] Oxazaphosphoridine monomer 3b.

[0248] [ka]

[0249] Compound 3b was obtained in the same manner as compound 3a, but by using III-b instead of III-a. 1 H NMR(300MHz,CDCl3)δ 8.80(1H,brs),7.96(1H,s),7.54-6.96(24H,m),6.79-6.71(4H,m),6.19(1H,t,J= 6.6Hz),4.90-4.73(4H,m),4.66(2H,brs),4.16-4.08(1H,m),3.76(6H,s),3.60-3 .36(2H,m),3.29(1H,d,J=3.9Hz),3.27-3.12(2H,m),3.09(2H,t,J=6.6Hz),2.59- 2.46(1H,m),2.07-1.97(1H,m),1.94-1.41(5H,m),1.36-1.18(1H,m),0.65(3H,s); 31P NMR(121.5MHz, CDCl3)δ 157.1(1P,s). [Examples]

[0250] Oxazaphosphoridine monomer 1a.

[0251] [ka]

[0252] Compound 1a was obtained in the same manner as compound 3a, by using "5'-O-(DMTr)-6-N-(benzoyl)adenosine" instead of "5'-O-(DMTr)-2-N-(phenoxyacetyl)-6-O-(cyanoethyl)guanosine". 1 H NMR(600MHz,CDCl3)δ 8.71(1H,s),8.12(1H,s),8.04(2H,d,J=7.8Hz),7.62-7.15(23H,m),6.80-6.75(4H,m),6.37(1H,dd,J=7.8,6.0Hz),4.94-4.8 8(1H,m),4.80(1H,ddd,J=12.0,6.0,5.4Hz),4.07-4.04(1H,m),3.76(6H,s),3.58-3.49(1H,m),3.41-3.34(1H,m),3.33(1H,dd ,J=10.8,4.8Hz),3.25(1H,dd,J=10.8,4.8Hz),3.13-3.06(1H,m),2.66-2.58(1H,m),2.40-2.35(1H,m),1.91-1.84(1H,m),1. 73-1.66(1H,m),1.56(1H,dd,J=15.0,9.0Hz),1.44(1H,dd,J=15.0,5.4Hz),1.47-1.41(1H,m),1.30-1.23(1H,m),0.63(3H,s); 31 P NMR(243.0MHz, CDCl3)δ 151.8(1P,s). [Examples]

[0253] Oxazaphosphoridine monomer 1b.

[0254] [ka]

[0255] Compound 1b was obtained in the same manner as compound 1a, but by using III-b instead of III-a. 1 H NMR(300MHz,CDCl3)δ 9.06(1H,brs),8.76(1H,s),8.12(1H,s),8.07-7.99(2H,m),7.64-7.14(22H,m),6.83-6. 75(4H,m),6.25(1H,t,J=6.6Hz),4.86-4.75(2H,m),4.20-4.15(1H,m),3.77(6H,s),3.61- 3.38(2H,m),3.36(1H,dd,J=10.2,4.2Hz),3.27(1H,dd,J=10.2,4.2Hz),3.27-3.13(1H,m) ,2.71-2.59(1H,m),2.12-2.01(1H,m),1.94-1.42(5H,m),1.36-1.20(1H,m),0.67(3H,s); 31 P NMR(121.5MHz, CDCl3)δ 157.3(1P,s). [Examples]

[0256] Oxazaphosphoridine monomer 2a.

[0257] [ka]

[0258] Compound 2a was obtained in the same manner as compound 3a, by using "5'-O-(DMTr)-4-N-(isobutyryl)cytidine" instead of "5'-O-(DMTr)-2-N-(phenoxyacetyl)-6-O-(cyanoethyl)guanosine". 1H NMR(300MHz,CDCl3)δ 8.33(1H,brs),8.17(1H,d,J=7.5Hz),7.52-7.22(19H,m),7.07(1H,d,J=7.5Hz),6.8 8-6.81(4H,m),6.20(1H,t,J=6.2Hz),4.81-4.64(2H,m),3.93-3.87(1H,m),3.79(6H ,s),3.59-3.43(1H,m),3.39-3.29(3H,m),3.16-3.02(1H,m),2.69-2.52(2H,m),2.1 2-2.00(1H,m),1.91-1.50(3H,m),1.47-1.32(2H,m),1.27-1.16(7H,m),0.60(3H,s); 31 P NMR(121.5MHz, CDCl3)δ 154.8(1P,s). [Examples]

[0259] Oxazaphosphoridine monomer 2b.

[0260] [ka]

[0261] Compound 2b was obtained in the same manner as compound 2a, but by using III-b instead of III-a. 1 H NMR(300MHz,CDCl3)δ 8.33(1H,d,J=7.5Hz),8.23(1H,brs),7.57-7.22(19H,m),7.12(1H,d,J=7.5Hz),6.88-6 .81(4H,m),6.15(1H,dd,J=6.6,4.2Hz),4.82-4.63(2H,m),4.03-3.97(1H,m),3.80(6H,s ),3.55-3.26(4H,m),3.19-3.05(1H,m),2.59(1H,quintet,J=6.9Hz),2.39-2.27(1H,m), 2.21-2.10(1H,m),1.90-1.56(3H,m),1.50-1.32(2H,m),1.26-1.17(7H,m),0.66(3H,s); 31P NMR(121.5MHz, CDCl3)δ 157.2(1P,s). [Examples]

[0262] Oxazaphosphoridine monomer 4a.

[0263] [ka]

[0264] Compound 4a was obtained in the same manner as compound 3a, by using "5'-O-(DMTr)thymidine" instead of "5'-O-(DMTr)-2-N-(phenoxyacetyl)-6-O-(cyanoethyl)guanosine". 1 H NMR(300MHz,CDCl3)δ 7.58-7.23(21H,m),6.86-6.79(4H,m),6.35(1H,dd,J=8.1,5.7Hz),4.79-4.67(2H,m),3.83-3.7 8(1H,m),3.78(6H,s),3.59-3.43(1H,m),3.34(1H,dd,J=10.5,2.4Hz),3.35-3.24(1H,m),3.20( 1H,dd,J=10.5,2.4Hz),3.16-3.02(1H,m),2.36-2.26(1H,m),2.15-2.02(1H,m),1.92-1.77(1H, m),1.74-1.59(1H,m),1.52(1H,dd,J=14.7,9.0Hz),1.40(3H,s),1.45-1.15(3H,m),0.60(3H,s); 31 P NMR(121.5MHz, CDCl3)δ 153.7(1P,s). [Examples]

[0265] Oxazaphosphoridine monomer 4b.

[0266] [ka]

[0267] Compound 4b was obtained in the same manner as compound 4a, but by using III-b instead of III-a. 1 H NMR(300MHz,CDCl3)δ 8.46(1H,brs),7.59-7.20(20H,m),6.86-6.79(4H,m),6.26(1H,t,J=6.8Hz),4.78-4.6 5(2H,m),4.01-3.95(1H,m),3.78(6H,s),3.55-3.40(1H,m),3.42(1H,dd,J=10.5,2.7H z),3.40-3.28(1H,m),3.22(1H,dd,J=10.5,3.0Hz),3.19-3.06(1H,m),2.16-1.95(2H, m),1.90-1.54(3H,m),1.49-1.35(1H,m),1.43(3H,s),1.34-1.17(2H,m),0.67(3H,s); 31 P NMR(121.5MHz, CDCl3)δ 156.2(1P,s). [Examples]

[0268] Oxazaphosphoridine monomer 5a.

[0269] [ka]

[0270] Compound 5a was obtained in the same manner as compound 3a, by using "5'-O-(DMTr)-2'-O-methyl-6-N-(benzoyl)adenosine" instead of "5'-O-(DMTr)-2-N-(phenoxyacetyl)-6-O-(cyanoethyl)guanosine". 1H NMR(300MHz,CDCl3)δ 8.66(1H,s),8.13(1H,s),8.03(2H,d,J=7.2Hz),7.64-7.16(23H,m),6. 79(4H,d,J=8.7Hz),6.08(1H,d,J=6.3Hz),4.91-4.81(1H,m),4.77-4.6 9(1H,m),4.64-4.57(1H,m),4.15-4.10(1H,m),3.76(6H,s),3.60-3.23 (4H,m),3.35(3H,s),3.14-3.00(1H,m),1.90-1.19(6H,m),0.62(3H,s); 31 P NMR(121.5MHz, CDCl3)δ 155.8(1P,s). [Examples]

[0271] Oxazaphosphoridine monomer 5b.

[0272] [ka]

[0273] Compound 5b was obtained in the same manner as compound 5a, but by using III-b instead of III-a. 1 H NMR(300MHz,CDCl3)δ 9.12(1H,brs),8.73(1H,s),8.24(1H,s),8.07-8.01(2H,m),7.62-7.17(22H,m),6.83-6.77(4H,m),6.12(1H,d,J=4.8Hz),4.84-4.73(2H,m ),4.43(1H,t,J=4.8Hz),4.25-4.19(1H,m),3.77(6H,s),3.55-3.20(4 H,m),3.28(3H,s),3.16-3.03(1H,m),1.90-1.17(6H,m),0.65(3H,s); 31 P NMR(121.5MHz, CDCl3)δ 155.0(1P,s). [Examples]

[0274] Oxazaphosphoridine monomer 6a.

[0275] [ka]

[0276] Compound 6a was obtained in the same manner as compound 3a, by using "5'-O-(DMTr)-2'-O-methyl-4-N-(isobutyryl)cytidine" instead of "5'-O-(DMTr)-2-N-(phenoxyacetyl)-6-O-(cyanoethyl)guanosine". 1 H NMR(300MHz,CDCl3)δ 8.49(1H,d,J=7.2Hz),7.58-7.20(19H,m),6.96(1H,d,J=7.2Hz),6.90-6.82(4H,m),5.98(1H, s),4.84(1H,dd,J=13.1,7.5Hz),4.59(1H,dt,J=8.3,4.5Hz),4.19-4.13(1H,m),3.79(6H,s), 3.78-3.72(1H,m),3.63-3.40(3H,m),3.55(3H,s),3.36-3.24(1H,m),3.09-2.95(1H,m),2.59 (1H,septet,J=6.9Hz),1.85-1.53(5H,m),1.48-1.37(1H,m),1.24-1.17(6H,m),0.59(3H,s); 31 P NMR(121.5MHz, CDCl3)δ 155.2(1P,s). [Examples]

[0277] Oxazaphosphoridine monomer 6b.

[0278] [ka]

[0279] Compound 6b was obtained in the same manner as compound 6a, but by using III-b instead of III-a. 1H NMR(300MHz,CDCl3)δ 8.62(1H,d,J=7.5Hz),7.57-7.23(19H,m),7.02(1H,d,J=7.5Hz),6.89-6.81(4H,m), 5.92(1H,s),4.90(1H,dt,J=9.0,5.7Hz),4.61(1H,dt,J=8.7,4.8Hz),4.25-4.17(1H, m),3.81(6H,s),3.67(1H,d,J=4.5Hz),3.62-3.25(4H,m),3.38(3H,s),3.16-3.02(1 H,m),2.58(1H,septet,J=6.9Hz),1.87-1.40(6H,m),1.26-1.14(6H,m),0.64(3H,s); 31 P NMR(121.5MHz, CDCl3)δ 158.2(1P,s). [Examples]

[0280] Oxazaphosphoridine monomer 7a.

[0281] [ka]

[0282] Compound 7a was obtained in the same manner as compound 3a, by using "5'-O-(DMTr)-2'-O-methyl-2-N-(phenoxyacetyl)-6-O-(cyanoethyl)guanosine" instead of "5'-O-(DMTr)-2-N-(phenoxyacetyl)-6-O-(cyanoethyl)guanosine". 1H NMR(300MHz,CDCl3)δ 8.67(1H,brs),8.01(1H,s),7.56-7.16(24H,m),6.83-6.74(4H,m),6.08(1H,d,J=6.9Hz),4. 85-4.76(1H,m),4.84(2H,t,J=6.6Hz),4.65-4.56(1H,m),4.59(2H,brs),4.48(1H,dd,J=6.6 ,5.1Hz),4.09-4.05(1H,m),3.75(6H,s),3.60-3.42(2H,m),3.40-3.26(2H,m),3.35(3H,s), 3.18-3.05(1H,m),3.08(2H,t,J=6.6Hz),1.89-1.49(3H,m),1.48-1.16(3H,m),0.59(3H,s); 31 P NMR(121.5MHz, CDCl3)δ 156.9(1P,s). [Examples]

[0283] Oxazaphosphoridine monomer 7b.

[0284] [ka]

[0285] Compound 7b was obtained in the same manner as compound 7a, but by using III-b instead of III-a. 1 H NMR(300MHz,CDCl3)δ 8.74(1H,brs),8.09(1H,s),7.56-6.94(24H,m),6.84-6.71(4H,m),6.09(1H,d,J =4.8Hz),4.83-4.70(2H,m),4.83(2H,t,J=6.6Hz),4.63(2H,brs),4.35(1H,t,J=5 .0Hz),4.23-4.16(1H,m),3.75(6H,s),3.58-3.19(4H,m),3.32(3H,s),3.16-3.04 (1H,m),3.07(2H,t,J=6.6Hz),1.90-1.55(3H,m),1.48-1.15(3H,m),0.64(3H,s); 31P NMR(121.5MHz, CDCl3)δ 154.6(1P,s).

[0286] Oxazaphosphoridine monomer 8a.

[0287] [ka]

[0288] Compound 8a was obtained in the same manner as compound 3a, by using "5'-O-(DMTr)-2'-O-(methyl)uridine" instead of "5'-O-(DMTr)-2-N-(phenoxyacetyl)-6-O-(cyanoethyl)guanosine". 1 H NMR(300MHz,CDCl3)δ 7.91(1H,d,J=7.8Hz),7.58-7.20(19H,m),6.88-6.80(4H,m),5.96(1H,d,J=3. 3Hz),5.19(1H,d,J=7.8Hz),4.88-4.78(1H,m),4.66-4.57(1H,m),4.03-3.95(1 H,m),3.90-3.74(1H,m),3.78(6H,s),3.77-3.71(1H,m),3.58-3.29(2H,m),3.4 5(3H,s),3.13-2.82(2H,m),1.88-1.53(3H,m),1.49-1.16(3H,m),0.60(3H,s); 31 P NMR(121.5MHz, CDCl3)δ 155.3(1P,s). [Examples]

[0289] Oxazaphosphoridine monomer 8b.

[0290] [ka]

[0291] Compound 8b was obtained in the same manner as compound 8a, but by using III-b instead of III-a. 1H NMR(300MHz,CDCl3)δ 8.10(1H,d,J=8.4Hz),7.58-7.20(19H,m),6.87-6.79(4H,m),5.89(1H,d,J=1.5Hz) ,5.21(1H,d,J=8.4Hz),4.92-4.82(1H,m),4.73-4.63(1H,m),4.15-4.08(1H,m),3.8 9-3.73(1H,m),3.78(6H,s),3.66-3.62(1H,m),3.57-3.27(2H,m),3.30(3H,s),3.17 -2.82(2H,m),1.89-1.55(3H,m),1.55-1.40(1H,m),1.35-1.15(2H,m),0.66(3H,s); 31 P NMR(121.5MHz, CDCl3)δ 157.5(1P,s). [Examples]

[0292] Oxazaphosphoridine monomer 9a.

[0293] [ka]

[0294] Compound 9a was obtained in the same manner as compound 3a, by using "5'-O-(DMTr)-2'-deoxy-2'-fluoro-6-N-(benzoyl)adenosine" instead of "5'-O-(DMTr)-2-N-(phenoxyacetyl)-6-O-(cyanoethyl)guanosine". 1 H NMR(300MHz,CDCl3)δ 8.64(1H,s),8.14(1H,s),8.06-8.01(2H,m),7.63-7.07(23H,m),6.78-6.70(4H,m),6.12(1H,dd,J=18.0,2.4Hz),5.24-5.01(2H,m),4.9 4-4.84(1H,m),4.17-4.06(1H,m),3.73(6H,s),3.55-3.40(3H,m),3.30-3.22(1H,m),3.03-2.88(1H,m),1.92-1.19(6H,m),0.62(3H,s); 31P NMR(121.5MHz, CDCl3)δ 150.5(1P,d,J=7.7Hz). [Examples]

[0295] Oxazaphosphoridine monomer 9b.

[0296] [ka]

[0297] Compound 9b was obtained in the same manner as compound 9a, but by using III-b instead of III-a. 1 H NMR(300MHz,CDCl3)δ 9.07(1H,brs),8.80(1H,s),8.24(1H,s),8.08-8.01(2H,m),7.66-7.15(22H,m),6.81-6.75(4H,m),6.14(1H,dd,J= 18.0,1.8Hz),5.16-4.91(3H,m),4.28-4.21(1H,m),3.76(6H,s),3.57-3.11(5H,m),1.82-1.16(6H,m),0.65(3H,s); 31 P NMR(121.5MHz, CDCl3)δ 157.8(1P,d,J=5.6Hz). [Examples]

[0298] Oxazaphosphoridine monomer 10a.

[0299] [ka]

[0300] Compound 10a was obtained in the same manner as compound 3a, by using "5'-O-(DMTr)-2'-deoxy-2'-fluoro-4-N-(isobutyryl)cytidine" instead of "5'-O-(DMTr)-2-N-(phenoxyacetyl)-6-O-(cyanoethyl)guanosine". 1H NMR(300MHz,CDCl3)δ 8.66(1H,brs),8.41(1H,d,J=7.5Hz),7.55-7.20(19H,m),7.01(1H,d,J=7.5Hz),6.89-6.81(4H,m) ,6.06(1H,d,J=15.9Hz),4.85(1H,dd,J=51.4,3.9Hz),4.84(1H,dd,J=12.9,7.5Hz),4.77-4.59(1H, m),4.15-4.08(1H,m),3.79(6H,s),3.63-3.29(4H,m),3.10-2.96(1H,m),2.65(1H,septet,J=6.9H z),1.85-1.53(3H,m),1.48-1.17(3H,m),1.21(3H,d,J=4.8Hz),1.19(3H,d,J=4.8Hz),0.59(3H,s); 31 P NMR(121.5MHz, CDCl3)δ 155.5(1P,d,J=6.6Hz). [Examples]

[0301] [ka]

[0302] Oxazaphosphoridine monomer 10b. Compound 10b was obtained by using III-b instead of III-a in the same manner as compound 10a. 1 H NMR(300MHz,CDCl3)δ 8.53(1H,d,J=7.5Hz),7.57-7.23(20H,m),7.10(1H,d,J=7.5Hz),6.89-6.81(4H,m),6. 10(1H,d,J=15.9Hz),5.00-4.92(1H,m),4.84(1H,dd,J=51.5,3.3Hz),4.75-4.58(1H,m ),4.24(1H,d,J=9.3Hz),3.81(6H,s),3.65-3.39(3H,m),3.32-3.06(2H,m),2.59(1H,s eptet,J=6.9Hz),1.88-1.53(4H,m),1.49-1.34(2H,m),1.27-1.18(6H,m),0.65(3H,s);31 P NMR(121.5MHz, CDCl3)δ 159.0(1P,d,J=4.4). [Examples]

[0303] Oxazaphosphoridine monomer 11a.

[0304] [ka]

[0305] Compound 11a was obtained in the same manner as compound 3a, by using "5'-O-(DMTr)-2'-deoxy-2'-fluoro-2-N-(phenoxyacetyl)-6-O-(cyanoethyl)guanosine" instead of "5'-O-(DMTr)-2-N-(phenoxyacetyl)-6-O-(cyanoethyl)guanosine". 1 H NMR(300MHz,CDCl3)δ 8.74(1H,brs),8.03(1H,s),7.55-6.94(24H,m),6.80-6.69(4H,m),6.21(1H,dd,J =14.9,3.6Hz),5.34(1H,dt,J=52.3,3.6Hz),5.01-4.75(2H,m),4.84(1H,t,J=6.6 Hz),4.62(2H,brs),4.15-4.07(1H,m),3.73(6H,s),3.59-3.29(4H,m),3.15-3.00 (1H,m),3.07(2H,t,J=6.6Hz),1.90-1.49(3H,m),1.47-1.12(3H,m),0.58(3H,s); 31 P NMR(121.5MHz, CDCl3)δ 155.6(1P,d,J=10.9Hz). [Examples]

[0306] Oxazaphosphoridine monomer 11b.

[0307] [ka]

[0308] Compound 11b was obtained in the same manner as compound 11a, but by using III-b instead of III-a. 1 H NMR(300MHz,CDCl3)δ 8.81(1H,brs),8.06(1H,s),7.55-6.95(24H,m),6.77-6.69(4H,m),6.06 (1H,d,J=17.1Hz),5.24-5.08(1H,m),5.04-4.80(2H,m),4.87(1H,t,J=6. 6Hz),4.62(2H,brs),4.25-4.19(1H,m),3.73(6H,s),3.58-3.02(5H,m), 3.10(2H,t,J=6.6Hz),1.90-1.56(3H,m),1.50-1.15(3H,m),0.63(3H,s); 31 P NMR(121.5MHz, CDCl3)δ 158.0(1P,d,J=4.4Hz). [Examples]

[0309] Oxazaphosphoridine monomer 12a.

[0310] [ka]

[0311] Compound 12a was obtained in the same manner as compound 3a, by using "5'-O-(DMTr)-2'-deoxy-2'-fluorouridine" instead of "5'-O-(DMTr)-2-N-(phenoxyacetyl)-6-O-(cyanoethyl)guanosine". 1H NMR(300MHz,CDCl3)δ 7.85(1H,d,J=8.1Hz),7.58-7.20(19H,m),6.87-6.79(4H,m),5.98(1H,d,J=16.5Hz),5.23(1H,d,J=8.1Hz),4.86-4.61(3H,m),3.99(1H,d,J =6.9Hz),3.76(6H,d,J=3.0Hz),3.56-3.34(4H,m),3.10-2.96(1H,m),1.88-1.74(1H,m),1.72-1.52(2H,m),1.48-1.16(3H,m),0.61(3H,s); 31 P NMR(121.5MHz, CDCl3)δ 154.3(1P,d,J=8.9Hz). [Examples]

[0312] Oxazaphosphoridine monomer 12b.

[0313] [ka]

[0314] Compound 12b was obtained in the same manner as compound 12a, but by using III-b instead of III-a. 1 1H NMR (300MHz, CDCl3)δ 8.01(1H,d,J=8.4Hz),7.58-7.20(19H,m),6.87-6.79(4H,m),6.03(1H,d,J =16.2Hz),5.29(1H,d,J=8.4Hz),4.96(1H,dd,J=13.1,7.5Hz),4.80-4.54( 2H,m),4.15(1H,d,J=9.0Hz),3.78(6H,s),3.61-3.39(3H,m),3.37-3.25(1 H,m),3.23-3.09(1H,m),1.91-1.56(3H,m),1.51-1.13(3H,m),0.66(3H,s); 31 P NMR(121.5MHz, CDCl3)δ 158.9(1P,d,J=4.4Hz). [Examples]

[0315] Oxazaphosphoridine monomer 13a.

[0316] [ka]

[0317] Compound 13a was obtained in the same manner as compound 3a, by using "5'-O-(DMTr)-2'-O-TOM-6-N-(acetyl)adenosine" instead of "5'-O-(DMTr)-2-N-(phenoxyacetyl)-6-O-(cyanoethyl)guanosine". 1 H NMR(300MHz,CDCl3)δ 8.82(1H,brs),8.49(1H,s),8.10(1H,s),7.58-7.17(19H,m),6.83-6.73(4H,m) ,6.11(1H,d,J=6.6Hz),5.15(1H,dd,J=6.6,5.4Hz),4.98-4.77(4H,m),4.18-4. 11(1H,m),3.76(6H,s),3.59-3.25(4H,m),3.16-3.02(1H,m),2.62(3H,s),1.91 -1.53(3H,m),1.49-1.18(3H,m),0.96-0.80(3H,m),0.90(18H,s),0.62(3H,s); 31 P NMR(121.5MHz, CDCl3)δ 156.7(1P,s). [Examples]

[0318] Oxazaphosphoridine monomer 13b.

[0319] [ka]

[0320] Compound 13b was obtained by using III-b instead of III-a in the same manner as compound 13a. 1H NMR(300MHz,CDCl3)δ 8.56(1H,brs),8.55(1H,s),8.13(1H,s),7.57-7.17(19H,m),6.82-6.73(4H,m),6.16(1H,d,J =5.7Hz),5.06(1H,t,J=5.6Hz),4.93(1H,d,J=5.1Hz),4.83(1H,d,J=5.1Hz),4.81-4.69(2H,m ),4.27-4.19(1H,m),3.76(6H,s),3.55-3.40(2H,m),3.33-3.16(2H,m),3.12-2.97(1H,m),2. 63(3H,s),1.88-1.52(3H,m),1.45-1.16(3H,m),0.91-0.79(3H,m),0.86(18H,s),0.64(3H,s); 31 P NMR(121.5MHz, CDCl3)δ 154.8(1P,s). [Examples]

[0321] Oxazaphosphoridine monomer 14a.

[0322] [ka]

[0323] Compound 14a was obtained in the same manner as compound 3a, by using "5'-O-(DMTr)-2'-O-TOM-4-N-(acetyl)cytidine" instead of "5'-O-(DMTr)-2-N-(phenoxyacetyl)-6-O-(cyanoethyl)guanosine". 1H NMR(300MHz,CDCl3)δ 10.04(1H,brs),8.30(1H,d,J=7.5Hz),7.51-7.21(19H,m),6.99(1H,d,J=7.5Hz),6.89-6.81 (4H,m),6.12(1H,d,J=3.3Hz),5.07(1H,d,J=4.8Hz),5.05(1H,d,J=4.8Hz),4.84-4.75(1H,m ),4.62-4.52(1H,m),4.31-4.25(1H,m),4.08-4.01(1H,m),3.78(6H,d,J=3.0Hz),3.55-3.23 (4H,m),3.10-2.96(1H,m),2.24(3H,s),1.84-1.49(3H,m),1.46-0.96(24H,m),0.58(3H,s); 31 P NMR(121.5MHz, CDCl3)δ 156.5(1P,s). [Examples]

[0324] Oxazaphosphoridine monomer 14b.

[0325] [ka]

[0326] Compound 14b was obtained by using III-b instead of III-a in the same manner as compound 14a. 1H NMR(300MHz,CDCl3)δ 10.19(1H,brs),8.46(1H,d,J=7.5Hz),7.54-7.23(19H,m),7.01(1H,d,J=7.5Hz),6.88-6. 79(4H,m),6.19(1H,d,J=1.8Hz),5.11(1H,d,J=4.8Hz),5.07(1H,d,J=4.8Hz),4.81-4.71( 1H,m),4.60-4.51(1H,m),4.26-4.18(2H,m),3.79(6H,s),3.63-3.55(1H,m),3.48-3.28(2 H,m),3.21-2.94(2H,m),2.26(3H,s),1.81-1.49(3H,m),1.43-0.96(24H,m),0.62(3H,s); 31 P NMR(121.5MHz, CDCl3)δ 156.4(1P,s). [Examples]

[0327] Oxazaphosphoridine monomer 15a.

[0328] [ka]

[0329] Compound 15a was obtained in the same manner as compound 3a, by using "5'-O-(DMTr)-2'-O-TOM-2-N-(acetyl)guanosine" instead of "5'-O-(DMTr)-2-N-(phenoxyacetyl)-6-O-(cyanoethyl)guanosine". 1 H NMR(300MHz,CDCl3)δ 7.70(1H,s),7.63-7.13(21H,m),6.84-6.76(4H,m),5.77(1H,d,J=8.4Hz),5.41-5.33(1H,m),4.90(2H,s),4.78-4.68(2H,m),3. 86(1H,brs),3.75(3H,s),3.74(3H,s),3.56-3.41(2H,m),3.32-2.90(3H,m),1.92-1.10(9H,m),0.97-0.87(21H,m),0.52(3H,s); 31P NMR(121.5MHz, CDCl3)δ 158.1(1P,s). [Examples]

[0330] Oxazaphosphoridine monomer 15b.

[0331] [ka]

[0332] Compound 15b was obtained by using III-b instead of III-a in the same manner as compound 15a. 1 H NMR(300MHz,CDCl3)δ 7.77(1H,s),7.56-7.15(21H,m),6.82-6.75(4H,m),5.86(1H,d,J=7.5Hz),5.26 -5.17(1H,m),4.95(1H,d,J=5.4Hz),4.85(1H,d,J=5.4Hz),4.78-4.71(1H,m),4 .59-4.49(1H,m),4.10-4.05(1H,m),3.74(6H,s),3.52-3.37(2H,m),3.30-3.18 (1H,m),3.11-2.85(2H,m),1.85-1.15(9H,m),0.93-0.84(21H,m),0.62(3H,s); 31 P NMR(121.5MHz, CDCl3)δ 152.3(1P,s). [Examples]

[0333] Oxazaphosphoridine monomer 16a.

[0334] [ka]

[0335] Compound 16a was obtained in the same manner as compound 3a, by using "5'-O-(DMTr)-2'-O-TOM-uridine" instead of "5'-O-(DMTr)-2-N-(phenoxyacetyl)-6-O-(cyanoethyl)guanosine". 1 H NMR(300MHz,CDCl3)δ 7.76(1H,d,J=8.1Hz),7.55-7.18(20H,m),6.88-6.80(4H,m),6.11(1H,d,J=6.0Hz),5.32(1H,d,J=8.1Hz) ,4.99(1H,d,J=5.1Hz),4.93(1H,d,J=5.1Hz),4.84-4.75(1H,m),4.54-4.46(1H,m),4.38(1H,t,J=5.7Hz), 3.87-3.83(1H,m),3.78(3H,s),3.77(3H,s),3.56-3.42(1H,m),3.39-3.28(1H,m),3.36(1H,dd,J=11.0,2. 7Hz),3.25(1H,dd,J=11.0,2.7Hz),3.16-3.03(1H,m),1.88-1.12(6H,m),1.08-0.97(21H,m),0.59(3H,s); 31 P NMR(121.5MHz, CDCl3)δ 156.6(1P,s). [Examples]

[0336] Oxazaphosphoridine monomer 16b.

[0337] [ka]

[0338] Compound 16b was obtained by using III-b instead of III-a in the same manner as compound 16a. 1H NMR(600MHz,CDCl3)δ 7.87(1H,d,J=7.8Hz),7.52-7.48(4H,m),7.38-7.21(16H,m),6.83-6.7 9(4H,m),6.14(1H,d,J=4.8Hz),5.33(1H,d,J=7.8Hz),4.99(1H,d,J=5. 4Hz),4.89(1H,d,J=5.4Hz),4.67(1H,dd,J=13.8,7.2Hz),4.52(1H,dt,J=10.4,4.8Hz),4.31(1H,t,J=4.8Hz),4.06-4.03(1H,m),3.78(3H,s), 3.77(3H,s),3.47(1H,dd,J=10.4,2.4Hz),3.47-3.39(1H,m),3.22-3.1 7(2H,m),3.00(1H,ddd,J=19.5,10.4,4.8Hz),1.82-1.74(1H,m),1.68- 1.58(1H,m),1.56(1H,dd,J=14.4,8.4Hz),1.38(1H,dd,J=14.4,7.2Hz) ,1.31-1.25(1H,m),1.26-1.17(1H,m),1.08-0.98(21H,m),0.63(3H,s); 31 P NMR(243.0MHz, CDCl3)δ 154.3(1P,s). [Examples]

[0339] Oxazaphosphoridine monomer 17a.

[0340] [ka]

[0341] Compound 17a was obtained in the same manner as compound 3a, by using "5'-O-(DMTr)-2'-O,4'-C-methylene-6-N-(benzoyl)adenosine" instead of "5'-O-(DMTr)-2-N-(phenoxyacetyl)-6-O-(cyanoethyl)guanosine". 1H NMR(300MHz,CDCl3)δ 9.10(1H,brs),8.76(1H,s),8.32(1H,s),8.04(2H,d,J=7.2Hz),7.64-7.18(22H,m),6.84(4H,d,J=8.7Hz),6.10(1H,s),4.76(1H,d J=6.9Hz),4.58(1H,s),4.61-4.51(1H,m),3.91(1H,d,J=7.8Hz),3.77(1H,d,J=7.8Hz),3.75(6H,s), 3.50(1H,s),3.47-3.33(1H,m),3.31-3.19(1H,m),3.03-2.88(1H,m),1.84-1.09(6H,m),0.51(3H,s); 31 P NMR(121.5MHz, CDCl3)δ 152.9(1P,s). [Examples]

[0342] Oxazaphosphoridine monomer 17b.

[0343] [ka]

[0344] Compound 17b was obtained by using III-b instead of III-a in the same manner as compound 17a. 1 H NMR(300MHz,CDCl3)δ 8.81(1H,s),8.30(1H,s),8.07-8.00(2H,m),7.64-7.17(22H,m),6.86-6.79(4H,m),6.12(1H,s),4.81-4.72(1H,m),4.62(1H,d J=7.2Hz),4.57(1H,s),3.94(1H,d,J=7.8Hz),3.89(1H,d,J=7.8Hz),3.77(6H,s),3.48(2H,s),3.46- 3.32(1H,m),3.24-3.13(1H,m),3.10-2.97(1H,m),1.84-1.49(3H,m),1.42-1.09(3H,m),0.58(3H,s); 31 P NMR(121.5MHz, CDCl3)δ 157.3(1P,s). [Examples]

[0345] Oxazaphosphoridine monomer 18a.

[0346] [ka]

[0347] Compound 18a was obtained in the same manner as compound 3a, by using "5'-O-(DMTr)-2'-O,4'-C-methylene-4-N-(isobutyryl)-5-methylcytidine" instead of "5'-O-(DMTr)-2-N-(phenoxyacetyl)-6-O-(cyanoethyl)guanosine". 1 H NMR(300MHz,CDCl3)δ 7.88(1H,brs),7.58-7.18(20H,m),6.88-6.80(4H,m),5.65(1H,s),4.69-4.60(1H,m),4.52(1H, d,J=6.6Hz),4.49(1H,s),3.81-3.74(1H,m),3.75(3H,s),3.73(3H,s),3.64(1H,d,J=8.1Hz),3. 56(1H,d,J=11.1Hz),3.53(1H,d,J=8.1Hz),3.46(1H,d,J=11.1Hz),3.56-3.40(1H,m),3.32-3.2 0(1H,m),3.14-3.00(1H,m),1.85-1.12(6H,m),1.60(3H,s),1.19(6H,d,J=6.9Hz),0.55(3H,s); 31 P NMR(121.5MHz, CDCl3)δ 155.9(1P,s). [Examples]

[0348] Oxazaphosphoridine monomer 18b.

[0349] [ka]

[0350] Compound 18b was obtained by using III-b instead of III-a in the same manner as compound 18a. 1 H NMR(300MHz,CDCl3)δ 7.86(1H,brs),7.56-7.19(20H,m),6.88-6.79(4H,m),5.69(1H,s),4.86-4.76(1H,m),4.46(1H,s ),4.45(1H,d,J=7.5Hz),3.80-3.75(1H,m),3.79(6H,s),3.74(1H,d,J=8.1Hz),3.69(1H,d,J=8.1 Hz),3.51(1H,d,J=11.1Hz),3.44-3.30(1H,m),3.39(1H,d,J=11.1Hz),3.29-3.17(1H,m),3.11-2 .97(1H,m),1.86-1.52(3H,m),1.64(3H,s),1.45-1.10(3H,m),1.21(6H,d,J=6.6Hz),0.62(3H,s); 31 P NMR(121.5MHz, CDCl3)δ 158.2(1P,s). [Examples]

[0351] Oxazaphosphoridine monomer 19a.

[0352] [ka]

[0353] Compound 19a was obtained in the same manner as compound 3a, by using "5'-O-(DMTr)-2'-O,4'-C-methylene-2-N-(phenoxyacetyl)-6-O-(cyanoethyl)guanosine" instead of "5'-O-(DMTr)-2-N-(phenoxyacetyl)-6-O-(cyanoethyl)guanosine". 1H NMR(300MHz,CDCl3)δ 8.71(1H,brs),8.16(1H,s),7.50-7.17(21H,m),7.09-7.01(3H,m),6.86-6.79(4H,m),6.03(1H, s),4.84(2H,t,J=6.6Hz),4.72(2H,s),4.68(1H,d,J=7.2Hz),4.55-4.46(1H,m),4.50(1H,s),3.9 0(1H,d,J=7.8Hz),3.77(1H,d,J=7.8Hz),3.75(6H,s),3.51(1H,d,J=10.8Hz),3.47(1H,d,J=10.8 Hz),3.45-3.21(2H,m),3.08(2H,t,J=6.6Hz),3.03-2.89(1H,m),1.80-1.08(6H,m),0.47(3H,s); 31 P NMR(121.5MHz, CDCl3)δ 153.2(1P,s). [Examples]

[0354] Oxazaphosphoridine monomer 19b.

[0355] [ka]

[0356] Compound 19b was obtained in the same manner as compound 19a, but by using III-b instead of III-a. 1H NMR(300MHz,CDCl3)δ 8.86(1H,brs),8.13(1H,s),7.55-7.17(21H,m),7.08-6.98(3H,m),6.95-6.78(4H,m),6.01(1H,s ),4.86(2H,t,J=6.6Hz),4.82-4.73(1H,m),4.70(2H,s),4.64(1H,d,J=7.5Hz),4.49(1H,s),3.94( 1H,d,J=7.8Hz),3.89(1H,d,J=7.8Hz),3.77(6H,s),3.46(2H,s),3.45-3.30(1H,m),3.24-3.12(1 H,m),3.09(2H,t,J=6.6Hz),3.09-2.96(1H,m),1.81-1.50(3H,m),1.41-1.06(3H,m),0.58(3H,s); 31 P NMR(121.5MHz, CDCl3)δ 157.4(1P,s). [Examples]

[0357] Oxazaphosphoridine monomer 20a.

[0358] [ka]

[0359] Compound 20a was obtained in the same manner as compound 3a, by using "5'-O-(DMTr)-2'-O,4'-C-methylene-5-methyluridine" instead of "5'-O-(DMTr)-2-N-(phenoxyacetyl)-6-O-(cyanoethyl)guanosine". 1H NMR(300MHz,CDCl3)δ 7.71(1H,d,J=0.9Hz),7.50-7.17(20H,m),6.87-6.80(4H,m),5.61(1H,s),4.69-4.60(1H,m), 4.55(1H,d,J=6.9Hz),4.41(1H,s),3.74(3H,s),3.73(3H,s),3.64(1H,d,J=7.8Hz),3.55(1H,d ,J=7.8Hz),3.53(1H,d,J=10.8Hz),3.46(1H,d,J=10.8Hz),3.56-3.42(1H,m),3.35-3.24(1H,m ),3.13-3.00(1H,m),1.85-1.45(3H,m),1.55(3H,d,J=0.9Hz),1.41-1.12(3H,m),0.56(3H,s); 31 P NMR(121.5MHz, CDCl3)δ 155.1(1P,s). [Examples]

[0360] Oxazaphosphoridine monomer 20b.

[0361] [ka]

[0362] Compound 20b was obtained by using III-b instead of III-a in the same manner as compound 20a. 1 H NMR(300MHz,CDCl3)δ 7.69(1H,s),7.56-7.19(20H,m),6.88-6.79(4H,m),5.66(1H,s),4.87-4.77(1H,m),4. 47(1H,d,J=7.8Hz),4.40(1H,s),3.78(6H,s),3.74(1H,d,J=7.8Hz),3.68(1H,d,J=7.8 Hz),3.50(1H,d,J=10.8Hz),3.46-3.32(1H,m),3.39(1H,d,J=10.8Hz),3.30-3.19(1H, m),3.12-2.98(1H,m),1.85-1.56(3H,m),1.59(3H,s),1.46-1.12(3H,m),0.63(3H,s);31 P NMR(121.5MHz, CDCl3)δ 158.1(1P,s). [Examples]

[0363] Oxazaphosphoridine monomer 21a.

[0364] [ka]

[0365] Compound 21a was obtained in the same manner as compound 3a, by using "5'-O-(DMTr)-2'-O-methoxyethyl-5-methyluridine" instead of "5'-O-(DMTr)-2-N-(phenoxyacetyl)-6-O-(cyanoethyl)guanosine". 1 H NMR(300MHz,CDCl3)δ 7.62-7.18(21H,m),6.84(4H,d,J=8.7Hz),6.07(1H,d,J=5.7Hz),4.86-4.76(1H,m),4.63-4.54(1H,m),4.20(1H,t,J=5.4Hz),3.95-3.89 (1H,m),3.78(6H,s),3.78-3.71(2H,m),3.60-3.48(2H,m),3.44-3.02(5H,m),3.31(3H,s),1.88-1.15(6H,m),1.35(3H,s),0.58(3H,s); 31 P NMR(121.5MHz, CDCl3)δ 156.3(1P,s). [Examples]

[0366] Oxazaphosphoridine monomer 21b.

[0367] [ka]

[0368] Compound 21b was obtained in the same manner as compound 21a, but by using III-b instead of III-a. 1 H NMR(300MHz,CDCl3)δ 7.71(1H,d,J=1.2Hz),7.55-7.22(20H,m),6.86-6.78(4H,m),5.99(1H,d,J=3.9Hz),4.7 8-4.62(2H,m),4.13-4.08(1H,m),4.07-4.02(1H,m),3.77(6H,s),3.77-3.70(1H,m),3.6 5-3.56(1H,m),3.52-3.36(4H,m),3.33-3.14(2H,m),3.29(3H,s),3.08-2.94(1H,m),1. 86-1.72(1H,m),1.71-1.55(2H,m),1.30(3H,d,J=1.2Hz),1.47-1.16(3H,m)0.64(3H,s); 31 P NMR(121.5MHz, CDCl3)δ 155.6(1P,s). [Examples]

[0369] Oxazaphosphoridine monomer 22a.

[0370] [ka]

[0371] Compound 22a was obtained in the same manner as compound 4a, but by using VII-a instead of III-a. 1H NMR(300MHz,CDCl3)δ 7.57(1H,d,J=0.9Hz),7.37-6.94(20H,m),6.87-6.78(4H,m),6.48(1H,dd,J=8.6,5.7Hz),5.42(1H,d d,J=11.0,5.1Hz),4.81-4.71(1H,m),4.02(1H,d,J=11.0Hz),3.83(1H,d,J=2.1Hz),3.79(6H,s),3.6 1-3.41(2H,m),3.24-3.09(1H,m),3.16(1H,dd,J=10.8,2.4Hz),3.02(1H,dd,J=10.8,2.4Hz),2.54-2 .44(1H,m),2.34-2.22(1H,m),1.94-1.79(1H,m),1.74-1.56(1H,m),1.38(3H,s),1.38-1.28(2H,m); 31 P NMR(121.5MHz, CDCl3)δ 160.9(1P,s). [Examples]

[0372] [ka]

[0373] Oxazaphosphoridine monomer 22b. Compound 22b was obtained in the same manner as compound 22a, but by using VII-b instead of VII-a. 1 H NMR(300MHz,CDCl3)δ 7.57(1H,d,J=1.5Hz),7.43-7.11(20H,m),6.85-6.78(4H,m),6.48(1H,dd,J=7.5,5. 7Hz),5.58(1H,dd,J=11.4,5.1Hz),4.82-4.73(1H,m),4.17-4.02(2H,m),3.78(6H,s ),3.56-3.40(3H,m),3.32(1H,dd,J=10.7,2.4Hz),3.22-3.07(1H,m),2.26-2.04(2H ,m),1.95-1.81(1H,m),1.74-1.56(1H,m),1.40(3H,d,J=1.5Hz),1.44-1.34(2H,m); 31P NMR(121.5MHz, CDCl3)δ 162.2(1P,s). [Examples]

[0374] Oxazaphosphoridine monomer 23a.

[0375] [ka]

[0376] Compound 23a was obtained in the same manner as compound 4a, but by using IX-a instead of III-a. 1 H NMR(300MHz,CDCl3)δ 9.22(1H,brs),8.05-7.99(2H,m),7.52(1H,d,J=1.2Hz),7.41-7.19(11H,m),6.87-6. 79(4H,m),6.37(1H,dd,J=8.4,5.7Hz),4.88-4.75(2H,m),3.86-3.80(1H,m),3.79(6H, s),3.64-3.49(2H,m),3.27-3.12(3H,m),2.97(2H,d,J=6.6Hz),2.51-2.41(1H,m),2. 33-2.20(1H,m),2.03-1.75(2H,m),1.72-1.59(1H,m),1.46-1.36(1H,m),1.40(3H,s); 31 P NMR(121.5MHz, CDCl3)δ 157.5(1P,s). [Examples]

[0377] Oxazaphosphoridine monomer 23b.

[0378] [ka]

[0379] Compound 23b was obtained in the same manner as compound 23a, but by using IX-b instead of IX-a. 1H NMR(300MHz,CDCl3)δ 8.67(1H,brs),8.18-8.11(2H,m),7.57(1H,d,J=1.2Hz),7.47-7.22(11H,m),6.86-6.79(4H,m), 6.29(1H,t,J=6.6Hz),4.87(1H,dt,J=7.5,5.7Hz),4.80-4.72(1H,m),4.11-4.05(1H,m),3.79(6 H,s),3.67-3.47(2H,m),3.43(1H,dd,J=10.8,2.7Hz),3.27(1H,dd,J=10.8,2.4Hz),3.25-3.13( 1H,m),3.07-2.99(2H,m),2.19-2.12(2H,m),2.03-1.62(3H,m),1.46-1.30(1H,m),1.41(3H,s); 31 P NMR(121.5MHz, CDCl3)δ 158.1(1P,s). [Examples]

[0380] Oxazaphosphoridine monomer 24a.

[0381] [ka]

[0382] Compound 24a was obtained in the same manner as compound 4a, but by using XIII-a instead of III-a. 1H NMR(600MHz,CDCl3)δ 7.76(2H,d,J=9.0Hz),7.62(1H,d,J=1.2Hz),7.40(2H,d,J=7.2Hz),7.32-7.23(10H,m),6.85(4H,d,J=8.4Hz),6.41 (1H,dd,J=8.4,5.4Hz),4.94(1H,dd,J=12.3,5.4Hz),4.84-4.79(1H,m),4.03-4.01(1H,m),3.79(6H,s),3.59-3.53( 1H,m),3.52-3.44(2H,m),3.41(1H,dd,J=14.7,7.2Hz),3.37-3.30(2H,m),3.13(1H,ddd,J=19.3,10.3,4.1Hz),2.50 -2.44(1H,m),2.39(3H,s),2.35-2.29(1H,m),1.91-1.72(2H,m),1.64-1.59(1H,m),1.40(3H,s),1.12-1.05(1H,m); 31 P NMR(243.0MHz, CDCl3)δ 154.2(1P,s).

[0383] General procedure for the synthesis of chiral oligonucleotides: Automated solid-phase synthesis of chiral oligonucleotides was performed according to the cycle shown in Table 1. After synthesis, the resins were treated with 1 mL of 25% NH3 aqueous solution at 55°C for 12 hours. The mixture was allowed to cool to room temperature, and the resins were removed by membrane filtration. The filtrate was concentrated to dryness under reduced pressure. The residue was dissolved in 3 mL of H2O and analyzed by RP-UPLC-MS at a rate of 0.3 mL / min in 0.1 M triethylammonium acetate buffer (pH 7.0) at 50°C using a linear gradient of acetonitrile (0-50% / 30 min).

[0384] [Table 1] Comparative Example 1

[0385] [ka]

[0386] Oligonucleotides were generated using compound 25, which represents a conventional monomer. Figure 2 shows a table of products obtained in Comparative Example 1.

[0387] analysis The monomers in the examples were chemically stable. The monomer isolation yield exceeded 80%, which was higher than that of conventional methods.

[0388] The inventors synthesized oligonucleotide derivatives using the chiral reagents of the above examples based on the second general procedure and the monomers of the above examples based on the first general procedure. As shown in Figure 2, conventional monomers result in incomplete deprotection products, by-products, and defective sequences. On the other hand, the method of the present invention, as shown in Figure 1, results in defective sequences but produces almost no incomplete deprotection products or by-products. It is clear that the method of the present invention can reduce incomplete deprotection products and by-products. Because the present invention can reduce undesirable products, it was easy to isolate the target oligonucleotide derivative.

Claims

1. A combination comprising a chiral reagent or a salt thereof and an activating reagent, The chiral reagent has the following chemical formula (III-a): A combination of items. 【Chemistry 1】

2. A combination comprising a chiral reagent or a salt thereof and an activating reagent, The chiral reagent has the following chemical formula (III-b): A combination of items. 【Chemistry 2】

3. A combination comprising a chiral reagent or a salt thereof and an activating reagent, The chiral reagent has the following chemical formula (V-a): A combination of items. 【Transformation 3】

4. A combination comprising a chiral reagent or a salt thereof and an activating reagent, The chiral reagent has the following chemical formula (VII-a): A combination of items. 【Chemistry 4】

5. A combination comprising a chiral reagent or a salt thereof and an activating reagent, The chiral reagent has the following chemical formula (VII-b): A combination of items. 【Transformation 5】

6. A combination comprising a chiral reagent or a salt thereof and an activating reagent, The chiral reagent has the following chemical formula (IX-a): A combination of items. 【Transformation 6】

7. A combination comprising a chiral reagent or a salt thereof and an activating reagent, The chiral reagent has the following chemical formula (IX-b): A combination of items. 【Transformation 7】

8. A combination comprising a chiral reagent or a salt thereof and an activating reagent, The chiral reagent has the following chemical formula (XI-a): A combination of items. 【Transformation 8】

9. A combination comprising a chiral reagent or a salt thereof and an activating reagent, The chiral reagent has the following chemical formula (XIII-a): A combination of items. 【Chemistry 9】

10. A combination comprising a chiral reagent or a salt thereof and an activating reagent, The chiral reagent has the following chemical formula (XIII-b): A combination of items. 【Chemistry 10】

Images