Novel stable nucleoside phosphate and analogs thereof

Abstract

The present disclosure includes oligonucleotide and nucleic acid compositions having a stable phosphate moiety. Formula (Ia) and Formula (Ib) wherein: X is selected from the group consisting of a 3-to 5-membered cycloalkyl group,-CHCH-, a 3-to 5-membered heterocycle, and-CH3CH3-. # imgabs0 #

Description

[0001] CROSS-REFERENCE TO RELATED APPLICATIONS

[0002] This application claims priority to U.S. Provisional Application No. 63 / 376,579, filed on September 21, 2022, the entire contents of which are hereby incorporated by reference in their entirety. Technical Field

[0003] The present disclosure relates generally to the fields of chemistry, molecular biology, and genetic engineering, including oligonucleotides and nucleic acid moieties and molecules useful for stabilizing oligonucleotides, and the synthesis and use of oligonucleotide and nucleic acid moieties and molecules. Background Art

[0004] Effective gene silencing by RNAi in vivo requires recognition and binding of the 5'-phosphate of the siRNA guide strand by Argonaute proteins. It has been shown that the incorporation of 5'-E-vinyl-phosphonate 5'-(E)-VP increases the accumulation of siRNA, prolongs the duration of silencing, and protects siRNA from 5'-3'-exonucleases. ACS Chem.Biol.2013,8:71402-1406. Structural studies have shown that the 5' binding site in the MID domain of human Argonaute-2 (hAgo-2) is able to adjust key binding residues to compensate for changes introduced by modified nucleotides. Nucleic Acids Res.2017,45:63528-3536.

[0005] There remains a high unmet need for alternatives to 5'-(E)-VP. Summary of the invention

[0006] Therefore, there is an unmet need for alternatives to 5'-(E)-VP. The present invention meets this need by providing novel alternatives to 5'-(E)-VP.

[0007] In general aspects, the present disclosure relates to compounds represented by formula (Ia) or (Ib):

[0008]

[0009] Wherein: X is selected from 3- to 5-membered cycloalkyl, -CHCH-, 3- to 5-membered heterocyclic ring and -CHR 3 CHR 3 -; Y is selected from O or NR'; R' is a counterion, H or a protecting group; Z is selected from H, a counterion, an activating group and an oligonucleotide; A and the connecting dashed line are optional, and when present, A is selected from O, S and CR 4 R 4B is a nucleobase; each R is independently selected from an oligonucleotide, a counterion, H and a protecting group, such as a C1-C5 alkyl group and a POM or a C1-C5 alkyl group; each R 1 and R 2 is independently selected from H, F, OH and optionally substituted O-alkyl, or R 1 and R 2 To form an optionally substituted oxycyclobutene; each R 3 Independently selected from C1-C3 alkyl; each R 4 R is independently selected from H, F and C1-C5 alkyl; 5 and R 6 one of the groups is XP(O)(OR)2 and the other is H; provided that when A is O, then X is not -CHCH-, and when X is cyclopropyl, then at least one of A or Y is not O. In some embodiments, when A is present and X is selected from 3- to 5-membered cycloalkyl, -CHCH-, 3- to 5-membered heterocycle, and -CHR 3 CHR 3 -, then R 1 and R 2 Not each is H.

[0010] In some embodiments, the compound represented by the compound of formula (Ia) and (Ib) is represented by:

[0011]

[0012] In some embodiments, the compounds of Formula (Ia) and (Ib) are represented by:

[0013]

[0014] In some embodiments, the compound of formula (Ia) is represented by:

[0015] In some embodiments, X is cyclobutyl. In some embodiments, cyclobutyl is represented by:

[0016] Wherein the dotted line represents the point of connection with the adjacent atom. In some embodiments, X is -CHR 3 CHR 3 -, and R 3 In some embodiments, X is selected from the following:

[0017] Wherein the dotted line represents the point of connection with the adjacent atom. In some embodiments, X is a 3- to 5-membered heterocyclic ring selected from the following:

[0018]

[0019] Wherein the dotted lines represent the points of connection with adjacent atoms. 2 is an optionally substituted oxycyclobutene. In some embodiments, X is -CHCH-. In some embodiments, Y is O. In some embodiments, Z is an activating group. In some embodiments, the activating group is represented by:

[0020] Wherein the dotted line indicates the connection point with the adjacent atom. In some embodiments, Z is an oligonucleotide. In some embodiments, the oligonucleotide is the antisense strand of RNA, preferably the antisense strand of siRNA. In some embodiments, the 5' end of the oligonucleotide is connected to Y. In some embodiments, A is O. In some embodiments, A is CH2 or CHF. In some embodiments, X is -CHCH-. In some embodiments, B is uracil or thymine. In some embodiments, R 1 H and R 2 is OMe, OEt, MOE or F. In some embodiments, R is a protecting group, such as POM, Et, and Z is an activating group. In some embodiments, R is H and Z is an oligonucleotide. In other embodiments, R and Z are each an oligonucleotide, for example, the compounds of the present disclosure are not at the end of an oligonucleotide. In some embodiments, R 5 is XP(O)(OR)2. In other embodiments, R 6 It is XP(O)(OR)2.

[0021] In some embodiments, the compound is selected from:

[0022]

[0023] Or the compound of table, wherein the nucleobase is thymine or cytosine. In some embodiments, R 2 In some embodiments, R is Et and Z is an activating group. In some embodiments, R is H or a counterion and Z is an oligonucleotide.

[0024] Other aspects, features, and advantages of the invention will be apparent from the following disclosure, including the detailed description of the invention and preferred embodiments thereof, and the appended claims. DETAILED DESCRIPTION

[0025] Various publications, articles, and patents are cited or described in the background and throughout the specification; each of these references is incorporated herein by reference in its entirety. The discussion of documents, acts, materials, devices, articles, etc. included in this specification is intended to provide a context for the present invention. Such discussion is not an admission that any or all of these matters form part of the prior art with respect to any invention disclosed or claimed.

[0026] Unless otherwise defined, all technical and scientific terms used herein have the same meaning as commonly understood by one of ordinary skill in the art to which the invention belongs. Otherwise, certain terms used herein have the meanings described in this specification. All patents, published patent applications, and publications cited herein are incorporated herein by reference as if fully set forth herein.

[0027] It must be noted that as used herein and in the appended claims, the singular forms "a," "an," and "the" include plural referents unless the context clearly dictates otherwise.

[0028] Unless otherwise indicated, any numerical value, such as the % sequence identity or % sequence identity range described herein, is understood to be modified by the term "about" in all cases. Therefore, numerical values ​​generally include ± 10% of the value. For example, a dose of 10 mg includes 9 mg to 11 mg. Unless the context clearly indicates otherwise, as used herein, the numerical range used explicitly includes all possible subranges, all individual values ​​within the range, including integers within such ranges and fractions of these values.

[0029] As used herein, the connection term "and / or" between multiple listed elements is understood to include both single options and combined options. For example, where two elements are connected by "and / or", the first option refers to the application of the first element without the second element. The second option refers to the application of the second element without the first element. The third option refers to the application of the first element and the second element together. Any of these options is understood to fall within the meaning and therefore meets the requirements of the term "and / or" as used herein. The parallel applicability of more than one option is also understood to fall within the meaning and therefore meets the requirements of the term "and / or".

[0030] Unless otherwise indicated, the term "at least" preceding a series of elements should be understood to refer to each element in the series. Those skilled in the art will recognize, or will be able to ascertain using only routine experimentation, a variety of equivalents to the specific embodiments of the invention described herein. Such equivalents are intended to be encompassed by the present invention.

[0031] Throughout this specification and the claims that follow, unless the context requires otherwise, the word "comprise" and variations such as "include" and "contain" will be understood to imply the inclusion of the stated integers or steps or groups of integers or steps but not the exclusion of any other integers or steps or groups of integers or steps. When used herein, the term "comprise" may be replaced with the term "contains" or "includes", or sometimes when used herein, with the term "having".

[0032] When used herein, "consisting of excludes any element, step, or ingredient not specified in the claim elements. When used herein, "consisting essentially of does not exclude materials or steps that do not materially affect the basic and novel characteristics of the claim. Whenever used herein in the context of one aspect or embodiment of the present invention, any of the aforementioned terms "comprising," "containing," "including," and "having" may be replaced with the terms "consisting of" or "consisting essentially of" to alter the scope of the present disclosure.

[0033] As used herein, "nucleobases" may include unmodified, natural or modified nucleobases. "Unmodified" or "natural" nucleobases include the purine bases adenine (A) and guanine (G), and the pyrimidine bases thymine (T), cytosine (C) and uracil (U). "Modified nucleobases" include other synthetic and natural nucleobases, such as 5-methylcytosine (5-me-C), 5-hydroxymethylcytosine, xanthine, hypoxanthine, 2-aminoadenine, 6-methyl and other alkyl derivatives of adenine and guanine, 2-propyl and other alkyl derivatives of adenine and guanine, 2-thiouracil, 2-thiothymine and 2-thiocytosine, 5-halouracil and cytosine, 5-propynyl (-C≡C-CH3) uracil and other alkynyl derivatives of pyrimidine bases, 6-azouracil, Cytosine and thymine, 5-uracil (pseudouracil), 4-thiouracil, 8-halo, 8-amino, 8-thiol, 8-sulfanyl, 8-hydroxy and other 8-substituted adenines and guanines, 5-halo, especially 5-bromo, 5-trifluoromethyl and other 5-substituted uracils and cytosines, 7-methylguanine and 7-methyladenine, 2-F-adenine, 2-amino-adenine, 8-azaguanine and 8-azaadenine, 7-deazaguanine and 7-deazaadenine and 3-deazaguanine and 3-deazaadenine. Additional modified nucleobases include tricyclic pyrimidines such as phenoxazine cytidine (1H-pyrimido[5,4-b][1,4]benzoxazin-2(3H)-one), phenothiazine cytidine (1H-pyrimido[5,4-b][1,4]benzothiazin-2(3H)-one), G-tweezers such as substituted phenoxazine cytidines (e.g., 9-(2-aminoethoxy)-H-pyrimido[5,4-b][1,4]benzoxazin-2(3H)-one), carbazole cytidine (2H-pyrimido[4,5-b]indol-2-one), pyridoindole cytidine (H-pyrido[3,2,5]pyrrolo[2,3-d]pyrimidin-2-one). Modified nucleobases may also include those in which the purine or pyrimidine base is replaced with other heterocycles, for example, 7-deazaadenine, 7-deazaguanine, 2-aminopyridine, and 2-pyridone.

[0034] "Alkyl" refers to an optionally substituted saturated straight or branched hydrocarbon group. Examples include, but are not limited to, methyl, ethyl, propyl, isopropyl, butyl, isobutyl, tert-butyl, n-pentyl, and n-hexyl. "Cycloalkyl" refers to a cyclic alkyl group of 3 to 10 carbon atoms having a monocyclic or polycyclic ring, including, for example, adamantyl, cyclopropyl, cyclobutyl, cyclopentyl, cyclohexyl, cyclooctyl, etc. In some embodiments, the optionally substituted alkyl is substituted by one or more halogens (such as F).

[0035] As used herein, "heterocycle" refers to a saturated, unsaturated, or aromatic ring system of 3 to 18 atoms containing at least one N, O, S, or P. In some embodiments, the heterocycle contains one N, O, or S. In some embodiments, the heterocycle contains two N, O, or S.

[0036] As used herein, "protecting group" refers to a chemical modification of a functional group to obtain chemical selectivity in a subsequent chemical reaction. Examples of protecting groups include, for example, those disclosed in Greene Wuts, Protective Groups in Organic Synthesis.

[0037] The term "activating group" includes a part that increases the ability of the group to form a covalent bond with another molecule. The activating group includes a phosphite-triester, a phosphotriester, an H-phosphonate or preferably a phosphoramidite group on at least one oxygen atom of a sugar moiety. Preferably, the activating group is on the C-3' oxygen or C-5' oxygen of a nucleic acid monomer. Typically, the activating group is on the C-3' oxygen of a nucleic acid monomer for use in synthesizing probes in the 3' → 5' direction, and the oligonucleotide is attached to a support via the 3' end. The activating group is on the C-5' oxygen of a nucleic acid monomer for use in synthesizing probes in the 5' → 3' ("reverse") direction, and the oligonucleotide is attached to a support via the 5' end.

[0038] In order to help the reader of the application, the specification is divided into each paragraph or chapter, or relates to each embodiment of the application. These divisions should not be considered as the main content of a paragraph or chapter or embodiment being separated from the main content of another paragraph or chapter or embodiment. On the contrary, it will be appreciated by those skilled in the art that this specification has a wide range of applications, and encompasses all combinations of conceivable chapters, paragraphs and sentences. The discussion of any embodiment is intended to be exemplary only, and is not intended to indicate that the scope of the disclosure (including claims) is limited to these examples.

[0039] Compounds of the Disclosure

[0040] In general aspects, the present disclosure relates to compounds represented by formula (Ia) or (Ib):

[0041]

[0042] Wherein: X is selected from 3- to 5-membered cycloalkyl, -CHCH-, 3- to 5-membered heterocyclic ring and -CHR 3 CHR 3 -; Y is selected from O or NR'; R' is a counterion, H or a protecting group; Z is selected from H, a counterion, an activating group and an oligonucleotide; A and the connecting dashed line are optional, and when present, A is selected from O, S and CR 4 R 4B is a nucleobase; each R is independently selected from an oligonucleotide, a counterion, H and a protecting group, such as a C1-C5 alkyl group and a POM or a C1-C5 alkyl group; each R 1 and R 2 is independently selected from H, F, OH and optionally substituted O-alkyl, or R 1 and R 2 To form an optionally substituted oxycyclobutene; each R 3 Independently selected from C1-C3 alkyl; each R 4 independently selected from H, F, and C1-C5 alkyl; provided that when A is O, then X is not -CHCH-, and when X is cyclopropyl, then at least one of A or Y is not O. In some embodiments, when A is present and X is selected from 3- to 5-membered cycloalkyl, -CHCH-, 3- to 5-membered heterocycle, and -CHR 3 CHR 3 -, then R 1 and R 2 Not each is H.

[0043] In some embodiments, the compounds of Formula (Ia) and (Ib) are represented by:

[0044]

[0045] In some embodiments, the compounds of Formula (Ia) and (Ib) are represented by:

[0046]

[0047] In some embodiments, the compound of formula (Ia) is represented by:

[0048]

[0049] In some embodiments, X is cyclobutyl. In some embodiments, cyclobutyl is represented by:

[0050] Wherein the dotted line represents the point of connection with the adjacent atom. In some embodiments, X is -CHR 3 CHR 3 -, and R 3 In some embodiments, X is selected from the following:

[0051] Wherein the dotted line represents the point of connection with the adjacent atom. In some embodiments, X is a 3- to 5-membered heterocyclic ring having the following structure:

[0052] wherein the curve contains one or more heteroatoms (eg, N, O, or S). In some embodiments, X is a 3- to 5-membered heterocyclic ring selected from:

[0053]

[0054] wherein the dashed lines indicate points of attachment to adjacent atoms. In some embodiments, Y is O. In some embodiments, Y is O and X is not cyclopropyl.

[0055] In some embodiments, Z is an activating group. In some embodiments, Z is a phosphoramidite or a functionally similar moiety. In some embodiments, the activating group is represented by:

[0056] The dashed lines indicate the connection points with neighboring atoms.

[0057] In some embodiments, Z is an oligonucleotide. In some embodiments, the oligonucleotide is the antisense strand of RNA, preferably the antisense strand of siRNA. In some embodiments, the 5' end of the oligonucleotide is connected to Y.

[0058] In some embodiments, A is O. In some embodiments, A is O and X is not cyclopropyl. In some embodiments, A is CH2 or CHF.

[0059] In some embodiments, R 1 and R 2 independently selected from H, F, OH and optionally substituted O-alkyl, provided that when X is selected from 3- to 5-membered cycloalkyl, -CHCH-, 3- to 5-membered heterocyclic ring and -CHR 3 CHR 3 -, then R 1 and R 2 Not H or R 1 and R 2 To form an optionally substituted oxycyclobutene. In some embodiments, the substituted O-alkyl group includes -OMe, -OEt, -CH2CH2OCH3 (or MOE), -CF2CH2OCH3, -CH2CF2OCH3, -CH2CH2OCF3, -CF2CF2OCH3, -CH2CF2OCF3, -CF2CH2OCF3, -CF2CF2OCF3, -CHFCH2OCH3, -CHFCHFOCH3, -CHFCH2OCFH2, -CHFCH2OCHF2 and -CH2CHFOCH3, etc.

[0060] In some embodiments, B is uracil. In some embodiments, R 1 H and R 2In some embodiments, R is a protecting group, such as POM, C1-C5 alkyl (eg, Et), and Z is an activating group. In some embodiments, R is H and Z is an oligonucleotide.

[0061] In some embodiments, the compound is selected from:

[0062]

[0063]

[0064] Or a compound in the table, wherein the nucleobase is thymine or cytosine.

[0065] In some embodiments, R 2 is selected from H, F, OH and optionally substituted O-alkyl, such as OMe, OEt, MOE, etc. In some embodiments, R 2 In some embodiments, each R is independently selected from an oligonucleotide, a counterion, H, and a protecting group, such as POM or a C1-C5 alkyl group. In some embodiments, R is a protecting group, such as POM or a C1-C5 alkyl group (e.g., Et), and Z is an activating group, such as a phosphoramidite or a functionally similar part. In some embodiments, the activating group is represented by:

[0066] In some embodiments, R is H and Z is an oligonucleotide.

[0067] Example

[0068] The following examples are provided to illustrate but not to limit the invention of the present disclosure.One skilled in the art will recognize that the following procedures may be modified using methods known to those of ordinary skill in the art.

[0069] the term

[0070] This article uses the following abbreviations:

[0071] m=2'-O-methyl modification

[0072] f=2'-fluorine modification

[0073] VP = Vinylphosphonate

[0074] (invabasic) = inverse abasic

[0075] ps = phosphorothioate

[0076] TEG-Chol = cholesterol-triethylene glycol

[0077] ecc); cc = carbocyclic ring; e refers to the geometric isomer of vinylphosphonate

[0078] VP(ecc)mU = carbocyclic mU e-vinylphosphonate

[0079] DMP(RS)=R,S-dimethylphosphonate

[0080] DMP(SR)=S,R-dimethylphosphonate

[0081] DMP(RR)=R,R-dimethylphosphonate

[0082] DMP(SS)=S,S-dimethylphosphonate

[0083] P(ccb) = cis-cyclobutylphosphonate

[0084] P(tcb) = trans-cyclobutylphosphonate

[0085] (UNAA) = should be (UNA-A)

[0086] (UNA-A) = Adenosine Unlocked Nucleic Acid

[0087] (J15AdaC) = should be J2-C15AdaC = 2'-O-[15-(adamantyl-1)pentadecyl]cytidine Embodiment 1: Synthesis of dimethyl phosphonate

[0088] Scheme 1. Synthesis of dimethyl phosphonate

[0089]

[0090] Scheme 2. Synthesis of cyclobutyl phosphonate

[0091]

[0092] Example 2: Synthesis of Phosphonates

[0093] Example 2-1

[0094]

[0095] Under an argon inert atmosphere, sec-butyl lithium (450 mL, 583.8 mmol, 3.0 equiv) was added dropwise to a solution of potassium tert-butoxide (65.5 g, 583.8 mmol, 3.0 equiv) in tert-butyl methyl ether (1300 mL), and stirred at -78°C for 2.5 hours. A solution of lithium bromide (101.5 g, 1167.5 mmol, 6.0 equiv) in tetrahydrofuran (1000 mL) was added dropwise to the resulting solution under stirring at -78°C. The resulting solution was stirred at -15°C for 30 minutes. The resulting solution was cooled to -78°C. A solution of copper bromide (I)-dimethyl sulfide (60 g, 291.9 mmol, 1.5 equiv) in 6-O-(triisopropylsilyl)-d-galactaldehyde cyclic carbonate (420 mL) was added to the resulting solution at -78°C. The resulting solution was stirred at -78 ° C for 1 hour. At -78 ° C, a solution of (3aR, 6aR) -2,2- dimethyl -3a, 6a- dihydro -4H- cyclopentadiene [d] [1,3] dioxol -4-one (30g, 194.6mmol, 1.0 equivalent) in tetrahydrofuran (300mL) was added to the resulting solution. The resulting solution was stirred at -30 ° C for 30 minutes. The reaction mixture was diluted with 2000mL tert-butyl methyl ether. The reaction mixture was quenched by adding 100mL acetic acid and methanol (v / v=1:1). The organic layer was washed with 3×3000mL mixed ammonium chloride and ammonia solution (PH=9). The organic phase was dried over anhydrous sodium sulfate, filtered and concentrated under reduced pressure. The crude product was applied to a silica gel column with petroleum ether / ethyl acetate (100:1-50:1). 35g (74% yield) of 21 was obtained as a white solid. MS m / z[M+NH4 + ] + (ESI):260.30. 1 HNMR(300MHz,DMSO-d6)δ4.60(d,J=5.4Hz,1H),4.11(d,J=5.4,1.1Hz,1H),3.49(dd,J=8.7,2.7Hz,1H),3.35-3.29 (m,1H),2.61(dd,J=17.7,8.9Hz,1H),2.49-2.42(m,1H),1.95-1.84(m,1H),1.33(s,3H),1.27(s,3H),1.06(s,9H).

[0096] At 0 ° C, borohydride (7.034 g, 185.94 mmol, 1.5 equivalents) was slowly added to a solution of 21 (20 g, 123.96 mmol, 1.0 equivalents) and cerium (III) chloride heptahydrate (46.172 g, 123.96 mmol, 1.0 equivalents) in methanol (180 mL) under an inert atmosphere of argon. The resulting solution was stirred for 4 hours at room temperature. The reaction mixture was quenched by adding water. The resulting solution was extracted with 300 mL of ethyl acetate. The organic layers were combined and washed with 300 mL of brine. The organic layer was dried over anhydrous sodium sulfate, filtered and concentrated under vacuum. The crude product was applied to a silica gel column with petroleum ether / ethyl acetate (50: 1-20: 1). 18 g (89% yield) of 21 was obtained as a light yellow liquid.

[0097] Benzoyl chloride (114.05 mL, 982.14 mmol, 1.1 equivalents) was added dropwise to a solution of pyrimidine-2,4 (1H, 3H)-dione (100 g, 892.85 mmol, 1.0 equivalents) in 840 mL of acetonitrile and 160 mL of pyridine at 0 ° C under an inert atmosphere of argon. The resulting solution was stirred at room temperature for 5 hours. The reaction mixture was quenched with 200 mL of 1N hydrochloric acid, diluted with 500 mL of dichloromethane, and washed with 3 × 200 mL of saturated sodium chloride aqueous solution. The organic layers were combined, dried over anhydrous sodium sulfate and concentrated under vacuum. The residue was applied to a silica gel column with petroleum ether / ethyl acetate (10: 1-1: 1). 60 g (31% yield) 19 was obtained as a white solid.

[0098] Under an argon inert atmosphere, benzyl chloromethyl ether (52g, 333.33mmol, 1.2 equivalents) was added to a solution of 19 (60g, 277.78mmol, 1.0 equivalents), 1,8-diazabicyclo [5.4.0] undec-7-ene (48mL) in acetonitrile (600mL). The resulting solution was stirred at room temperature for 2 hours. The reactant was then concentrated under vacuum. The residue was extracted with ethyl acetate (250mL) and washed with 1×200mL water, 1×200mL saturated sodium chloride aqueous solution. The organic layer was dried over anhydrous sodium sulfate and concentrated under vacuum. The residue was dissolved in 600mL sodium methoxide (2M sodium methoxide in methanol solution) and stirred at room temperature for 1 hour under an argon inert atmosphere. The reaction mixture was then evaporated to dryness. The residue was applied to a silica gel column with petroleum ether / ethyl acetate (2: 1-1: 3). 55g (85% yield) of 20 was obtained as a white solid. MS m / z[MH] - (ESI):231.00.

[0099] To a solution of 22 (28.5 g, 116.8 mmol, 1.0 equiv), 20 (40.6 g, 175.2 mmol, 1.5 equiv), triphenylphosphine (76.6 g, 292.0 mmol, 2.5 equiv) in tetrahydrofuran (1000 mL) was slowly added dropwise diethyl azodicarboxylate (50.85 g, 292.0 mmol, 2.5 equiv) at -78°C under an inert atmosphere of argon. The resulting solution was stirred at -78°C for 1 hour, then slowly warmed to room temperature and stirred for 16 hours. The resulting mixture was concentrated under vacuum. The residue was purified by flash preparative HPLC using the following conditions: column, C18 silica gel; mobile phase, water and acetonitrile (30% acetonitrile to 100% in 20 minutes), UV 254 nm. The fractions were diluted with dichloromethane, dried over anhydrous sodium sulfate and concentrated under reduced pressure. 36.5 g (68% yield) of 23 were obtained as a yellow oil. MS m / z [M+H] + (ESI):459.30. 1 H NMR (300MHz, chloroform-d) δ7.32(t,J=3.3Hz,5H),7.20(d,J=7.9Hz,1H),5.71(d,J =7.8Hz,1H),5.39-5.15(m,3H),5.03(dt,J=6.8,3.5Hz,1H),4.61(d,J=3.2 Hz,2H),4.52(t,J=6.6Hz,1H),3.62(dd,J=8.7,4.6Hz,1H),3.37-3.25(m,J =8.3,5.9Hz,1H),2.33-2.03(m,3H),1.52(s,3H),1.28(s,3H),1.17(s,9H).

[0100] To a solution of 23 (35 g, 76.4 mmol, 1.0 eq.) in ethyl acetate (350 mL) at room temperature under an inert atmosphere of hydrogen were added 10% Pd / C (w t / w t =10%, 3.5g), trifluoroacetic acid (35mL). The resulting solution was stirred at room temperature for 4 hours. The solution was filtered and concentrated under reduced pressure. The residue was dissolved in 350mL trifluoroacetic acid / water = 1:1 at room temperature. The resulting solution was stirred at 50°C under an inert atmosphere of argon for 2 hours. The solution was concentrated under reduced pressure. The residue was dissolved in a 1N methanol solution of ammonium hydroxide (200mL). The resulting mixture was concentrated in vacuo. The residue was purified by rapid preparative HPLC using the following conditions: column, C18 silica gel; mobile phase, water and acetonitrile (0% acetonitrile reached 30% in 15 minutes), UV 254nm. The fractions were concentrated under reduced pressure. 15.7g (85% yield) of -5 was obtained as a light yellow oil. MS m / z[M+Na]+ (ESI):265.00.

[0101] Under an inert atmosphere of argon, diphenyl carbonate (22.57 g, 105.37 mmol, 1.5 equiv) and sodium bicarbonate (590.38 mg, 7.02 mmol, 0.1 equiv) were added to a solution of 5 (17 g, 70.24 mmol, 1.0 equiv) in N, N-dimethylformamide (170 mL) in sequence. The reaction mixture was heated at 150 ° C for 3 hours until the solution turned dark red. The reaction mixture was cooled to room temperature and slowly poured into 3000 mL of ether under stirring. The precipitate was collected by filtration, redissolved in methanol, decolorized with activated carbon and filtered. The filtrate was concentrated under vacuum. 10.3 g (73% yield) of 6 was obtained as a yellow solid. MS m / z[MH] - (ESI):223.00.

[0102] To a solution of 6 (14.7 g, 65.62 mmol, 1.0 equiv) in methanol (150 mL) were added trimethyl borate (13.6 g, 131.25 mmol, 2.0 equiv), trimethoxymethane (6.96 g, 65.6 mmol, 1.0 equiv), sodium bicarbonate (1.1 g, 13.1 mmol, 0.2 equiv) in sequence. The reaction mixture was stirred at 150 ° C for 16 hours in a sealed tube. The reaction mixture was cooled to room temperature and concentrated under vacuum. The residue was purified by rapid preparative HPLC with the following conditions: column, C18 silica gel; mobile phase, water and acetonitrile (0% acetonitrile reached 40% in 20 minutes), UV 254nm. The fractions were concentrated under reduced pressure. 7.7 g (45% yield) of 7 was obtained as a light yellow oil. MS m / z [MH] - (ESI):255.00.

[0103] To a solution of 7 (5 g, 19.53 mmol, 1.0 equiv) in pyridine (50 mL) was added 4,4'-dimethoxytrityl chloride (6.94 g, 20.5 mmol, 1.05 equiv) at 0 ° C under an inert atmosphere of argon. The resulting solution was stirred at room temperature for 2 hours. The reaction mixture was quenched by 10 mL of methanol, diluted with 200 mL of dichloromethane, and washed with 2×150 mL of saturated aqueous sodium bicarbonate solution and 2×150 mL of saturated aqueous sodium chloride solution, respectively. The organic layer was dried over anhydrous sodium sulfate, filtered and concentrated under reduced pressure. The residue was purified by rapid preparative HPLC using the following conditions: column, C18 silica gel; mobile phase, water and acetonitrile (30% of acetonitrile reached 100% in 20 minutes), UV 254nm. The fractions were concentrated under reduced pressure. 8.4 g (77% yield) of 31 was obtained as a light yellow solid. MS m / z [MH] -(ESI):557.10. 1 H NMR(300MHz,DMSO-d6)δ11.13(s,1H),7.45-7.35(m,3H),7.35-7.18(m,7H), 6.93-6.84(m,4H),5.57(d,J=7.5Hz,1H),5.10-4.97(m,1H),4.37(d,J=7.3Hz ,1H),4.14-3.99(m,1H),3.90(t,1H),3.74(s,6H),2.89(t,J=7.9Hz,1H),3. 23-3.15(m,3H),1.96-1.69(m,2H),1.35-1.08(m,1H),0.83(t,J=7.3Hz,1H).

[0104] To a solution of 31 (8.4 g, 15.05 mmol, 1.0 equiv) in N,N-dimethylformamide (85 mL) was added imidazole (2.55 g, 37.59 mmol, 2.5 equiv) and tert-butyldimethylsilyl chloride (5.66 g, 37.59 mmol, 2.5 equiv) in sequence at room temperature under an inert atmosphere of argon. The resulting solution was stirred at room temperature for 2 hours. The reaction mixture was diluted with 500 mL of dichloromethane and washed with 2×200 mL of saturated aqueous sodium bicarbonate solution and 1×200 mL of saturated aqueous sodium chloride solution, respectively. The organic phase was dried over anhydrous sodium sulfate, filtered and concentrated under reduced pressure. The residue was purified by flash preparative HPLC using the following conditions: column, C18 silica gel; mobile phase, water and acetonitrile (30% of acetonitrile to 100% in 20 minutes), UV 254 nm. The fractions were concentrated under reduced pressure. 8.6 g (81% yield) of 32 were obtained as a light yellow oil. MS m / z [MH] - (ESI):671.30.

[0105] Under an inert atmosphere of argon, triethylsilane (5.2 g, 44.64 mmol, 3.0 equiv) was added to a solution of 32 (10.0 g, 14.88 mmol, 1.0 equiv) dissolved in 100 mL of 6% dichloroacetic acid in dichloromethane. The resulting solution was stirred at room temperature for 1 hour. The reaction mixture was diluted with 200 mL of dichloromethane and washed with 2×200 mL of saturated aqueous sodium bicarbonate solution, 1×100 mL of water and 1×200 mL of saturated aqueous sodium chloride solution, respectively. The organic phase was dried over anhydrous sodium sulfate, filtered and concentrated under reduced pressure. The residue was purified by rapid preparative HPLC using the following conditions: column, C18 silica gel; mobile phase, water and acetonitrile (30% of acetonitrile reached 100% in 20 minutes), UV 254 nm. The fractions were concentrated under reduced pressure. 4.2 g (76%) of 31 was obtained as a white solid. MS m / z [MH]- (ESI):369.15. 1 H NMR (300MHz, DMSO-d6) δ11.09(d,J=5.8Hz,1H),7.42(dd,J=7.6,5.8Hz,1H),5.57(dd,J =7.6,1.5Hz,1H),5.05(td,J=9.7,5.5Hz,1H),4.52(t,J=5.2Hz,1H),4.26(dd,J=6.7,5 .4Hz,1H),4.02(t,J=5.4Hz,1H),3.49-3.59(m,J=10.6,5.3Hz,1H),3.33-3.27(m,1H), 3.17(s,3H),1.90-2.03(m,J=7.5Hz,1H),1.86-1.56(m,2H),0.88(s,9H),0.06(d,6H).

[0106] At 0 ° C, Dess-Martin periodinane (4.0 g, 10.12 mmol, 1.5 equiv) was added to a solution of 9 (2.5 g, 6.74 mmol, 1.0 equiv) in 25 mL of dichloromethane under an inert atmosphere of argon. The resulting solution was stirred at room temperature for 1 hour. The reaction mixture was diluted with 300 mL of dichloromethane and washed with 1×200 mL of saturated aqueous sodium thiosulfate solution, 1×200 mL of saturated aqueous sodium bicarbonate solution, 2×100 mL of water and 1×200 mL of saturated aqueous sodium chloride solution, respectively. The organic phase was dried over anhydrous sodium sulfate, filtered and concentrated under reduced pressure. 2.1 g (84%) of 10 was obtained as a white solid. MS m / z [MH] - (ESI): 367.05. It was used in the next step without further purification.

[0107] To a solution of tetraethyl methylenebis(phosphonic acid) ester (2.13 g, 7.41 mmol, 1.3 equiv) in anhydrous tetrahydrofuran (20 mL) at 0°C was added potassium tert-butoxide (2.13 g, 5.70 mmol, 1.35 equiv). The reaction mixture was stirred at 0°C for 10 minutes and then warmed to room temperature for 30 minutes. A solution of 10 (2.10 g, 5.70 mmol, 1.0 equiv) in 20 mL of anhydrous tetrahydrofuran was then added dropwise at 0°C. The resulting solution was stirred at 0°C for 3 hours. The reaction mixture was diluted with 200 mL of dichloromethane and washed with 2×200 mL of saturated aqueous sodium bicarbonate. The organic phase was dried over anhydrous sodium sulfate, filtered and concentrated under reduced pressure. The residue was purified by flash preparative HPLC using the following conditions: column, C18 silica gel; mobile phase, water and acetonitrile (30% acetonitrile to 100% in 20 minutes), UV 254 nm. The fractions were diluted with an equal volume of dichloromethane. The organic layer was separated. The aqueous phase was extracted with 3×20 mL of dichloromethane. The organic phases were combined and dried over anhydrous sodium sulfate. The solid was filtered off. The filtrate was concentrated at 25° C. 2 g (68%) of 12 was obtained as a light yellow solid. MS m / z [M+H] + (ESI):503.20. 1 H NMR (300MHz, DMSO-d6) δ11.18(s,1H),7.45(d,J=7.6Hz,1H),6.65-6.48(m,J=21.9,17. 1,7.6Hz,1H),5.90-5.81(m,1H),5.61(d,J=7.5Hz,1H),5.06-4.95(m,1H),4.40(dd,J=9 .8,5.8Hz,1H),4.00-3.89(m,J=8.3,7.0,1.3Hz,4H),3.81(dd,J=5.9,2.6Hz,1H),3.21 (s,3H),2.75(s,1H),1.93-.73(m,2H),1.22(t,J=7.0Hz,6H),0.87(s,9H),0.03(s,6H). 31 PNMR(121MHz,DMSO-d6)δ17.62.

[0108] Under an argon inert atmosphere, 11 (2.00 g, 5.15 mmol, 1.0 equiv) was dissolved in 20 mL of formic acid and H2O (v / v=1:1). The mixture solution was stirred at room temperature for 1 hour. The resulting solution was concentrated under vacuum. The residue was purified by rapid preparative HPLC using the following conditions: column, C18 silica gel; mobile phase, water and acetonitrile (15% acetonitrile to 35% in 20 minutes), UV 254 nm. The fractions were concentrated under reduced pressure. 1.3 g (84%) of 12 was obtained as a white solid. MS m / z [M+H] + (ESI):389.25. 1 H NMR(300MHz,DMSO-d6)δ11.17(d,1H),7.48-7.38(m,1H),6.76-6.55(m,1H),5.87-5.72(m,1H),5.60(d,1H),5.02(td,1H),4.74( d,1H),4.17-4.07(m,1H),4.02-3.90(m,4H),3.88-3.83(m,1H),3.26(s,3H),2.75-2.58(m,1H),1.94-1.72(m,2H),1.23(td,6H). 31 P NMR (121MHz, DMSO-d6) δ18.08.

[0109] To a solution of 12 (1.7 g, 4.38 mmol, 1.0 equiv) in dichloromethane (170 mL) were added propylamino)phosphinooxy)propionitrile (1.72 g, 5.69 mmol, 1.3 equiv), 4,5-dicyanoimidazole (569 mg, 4.88 mmol, 1.1 equiv) in sequence. The resulting solution was stirred at room temperature for 40 minutes. The resulting solution was diluted with 500 mL of dichloromethane and washed with 1×300 mL of sodium bicarbonate and 1×300 mL of brine, respectively. The organic layer was dried over anhydrous sodium sulfate, filtered and concentrated under vacuum. The crude product was purified by flash preparative HPLC using the following conditions (IntelFlash-1): column, C18 silica gel; mobile phase, water and acetonitrile (30% acetonitrile to 100% in 10 minutes and maintained at 100% for 5 minutes); detector, UV 254 nm. The fractions were diluted with an equal volume of dichloromethane. The organic layer was separated. The aqueous phase was extracted with 3×50 mL of dichloromethane. The organic phases were combined and dried over anhydrous sodium sulfate. The solid was filtered off. The filtrate was concentrated at 25° C. to obtain the product, which was dried under reduced pressure at 25° C. for 4 hours. 1.85 g of 0 was obtained as a white solid. MS m / z [MH] - (ESI):587.10. 1H NMR(300MHz,DMSO-d6)δ11.14(s,1H),7.42(dd,J=7.6,1.1Hz,1H),6.67-6.49(m,1H ),5.95-5.69(m,1H),5.59(d,J=7.6Hz,1H),5.10-4.98(m,1H),4.53-4.29(m,1H),4. 02-3.83(m,5H),3.80-3.66(m,2H),3.63-3.47(m,2H),3.22(d,J=3.0Hz,3H),2.85( s,1H),2.79-2.71(m,2H),1.96-1.76(m,2H),1.24-1.17(m,6H),1.15-1.11(m,12H). 31 P NMR (121MHz, DMSO) δ147.53,146.78,17.69,17.50.

[0110] Example 2-2

[0111]

[0112] To a solution of (3aR,5S,6R,6aR)-5-((R)-2,2-dimethyl-1,3-dioxolan-4-yl)-2,2-dimethyltetrahydrofuro[2,3-d][1,3]dioxol-6-ol (400 g, 1.54 mol, 1.0 equiv) in 4 L of tetrahydrofuran was added sodium hydride (55.2 g, 2.31 mol, 1.5 equiv) at 0°C under an inert atmosphere of nitrogen. The resulting solution was stirred at 0°C for 20 minutes. Then, 2-(bromomethyl)naphthalene (509.7 g, 2.31 mol, 1.5 equiv) was added dropwise at 0°C with stirring. The reaction mixture was stirred at room temperature for 3 hours. The reaction mixture was quenched with saturated aqueous ammonium chloride solution. The resulting mixture was diluted with 4 L of ethyl acetate and washed with 2×4 L of water and 2×4 L of saturated aqueous sodium chloride solution, respectively. The organic phase was dried over anhydrous sodium sulfate, filtered and concentrated under reduced pressure. The residue was applied to a silica gel column with ethyl acetate / petroleum ether (1:5-1:3). 490 g (80%) of 41 was obtained as a yellow oil. MS m / z [M+Na] + (ESI):423.08.

[0113] Under an inert atmosphere of nitrogen, 41 (490 g, 1.23 mol, 1.0 equiv) was dissolved in acetic acid (80% aqueous solution, 4900 mL) and stirred at room temperature overnight. The resulting mixture was concentrated under reduced pressure and diluted with 5000 mL of ethyl acetate. The resulting mixture was washed with 3×1500 mL of saturated sodium bicarbonate aqueous solution, respectively. The organic phase was dried over anhydrous sodium sulfate, filtered and concentrated under reduced pressure. The residue was applied to a silica gel column with ethyl acetate / petroleum ether (1:5-1:1). 390 g (85%) of 42 was obtained as a yellow oil. MS m / z[M+Na] + (ESI):383.21.

[0114] At 0 ° C, imidazole (146.8 g, 2.16 mol, 4.0 equivalents) and tert-butyldiphenylsilyl chloride (156 g, 0.55 mol, 1.0 equivalents) were added to a mixture solution of 42 (195 g, 0.54 mol, 1.0 equivalents) in 2000 mL of dichloromethane in an inert atmosphere of argon. The reaction mixture was warmed to room temperature and stirred at room temperature for 4 hours. The resulting solution was diluted with 5000 mL of dichloromethane and washed with 3×1500 mL of water and 3×1500 mL of saturated sodium chloride aqueous solution, respectively. The organic phase was dried over anhydrous sodium sulfate, filtered and concentrated under reduced pressure. The residue was applied to a silica gel column with ethyl acetate / petroleum ether (1:10-1:5). 190 g (60%) of 43 was obtained as a yellow oil. MS m / z[M+Na] + (ESI):621.25. 1 H NMR(300MHz,DMSO-d6)δ7.90-7.85(m,1H),7.79-7.71(m,3H),7.65-7.56(m,4H),7.50-7.4 5(m,2H),7.43-7.33(m,5H),7.29-7.22(m,2H),5.72(d,J=3.7Hz,1H),5.06(d,J=4.8Hz,1H) ,4.78-4.72(m,2H),4.57(d,J=11.9Hz,1H),4.19-4.15(m,1H),4.13-3.98(m,1H),3.87-3. 94(m,1H),3.83-3.77(m,1H),3.57(d,J=6.5Hz,1H),1.47(s,3H),1.30(s,3H),0.94(s,9H).

[0115] To a solution of oxalyl chloride (120.84 g, 0.953 mol, 1.5 eq) in dichloromethane (2000 mL) was added dimethyl sulfoxide (78.19 g, 0.95 mol, 3.0 eq) dropwise at -78°C with stirring. The resulting solution was stirred at -78°C for 30 minutes, and a solution of 43 (120 g, 158.86 mmol, 1.0 eq) in dichloromethane (500 mL) was added dropwise at -78°C with stirring. The resulting solution was allowed to react for another 1.5 hours at -78°C with stirring. Then, triethylamine (72.24 g, 714.75 mmol, 4.5 eq) was added dropwise at -78°C with stirring. The resulting solution was allowed to react for another 2 hours at -78°C with stirring. The resulting solution was diluted with 500 mL of dichloromethane. The resulting mixture was washed with 1×500 mL of saturated aqueous sodium bicarbonate solution and 1×500 mL of saturated aqueous sodium chloride solution, respectively. The mixture was dried over anhydrous sodium sulfate and concentrated under reduced pressure. The residue was applied to a silica gel column with ethyl acetate / petroleum ether (1:10-1:6). 80 g (85%) of 44 was obtained as a yellow oil. MS m / z [M+Na] + (ESI):619.57.

[0116] Under an inert atmosphere of nitrogen, sodium hydride (6.5 g, 0.269 mol, 2.0 equiv) was added to a solution of methyltriphenylphosphonium bromide (96.2 g, 0.269 mol, 2.0 equiv) in tetrahydrofuran (500 mL). The resulting solution was stirred at room temperature for 30 minutes. A solution of 44 (80 g, 134.7 mmol, 1.00 equiv) in tetrahydrofuran (500 mL) was added. The resulting solution was allowed to react for another 3 hours under stirring at room temperature. The reactants were then quenched by adding a saturated aqueous solution of ammonium chloride. The resulting solution was extracted with 4000 mL of ethyl acetate, and the organic layers were combined. The resulting mixture was washed with water and a saturated aqueous sodium chloride solution. The mixture was dried over anhydrous sodium sulfate and concentrated under reduced pressure. The residue was applied to a silica gel column with ethyl acetate / petroleum ether (1:20-1:5). 63 g (80%) of 45 was obtained as a yellow oil. MS m / z[M+Na] + (ESI):617.60.

[0117] At 0°C, borane-methyl sulfide complex (10M, 42.1mL, 2.0 equivalents) was added to a solution of 45 (125g, 210.4mmol, 1.0 equivalents) in tetrahydrofuran (1250mL). The resulting solution was stirred at room temperature for 2 hours. Sodium hydroxide (2N aqueous solution 630mL, 6.0 equivalents) was then added at room temperature, and hydrogen peroxide (30%, 143.1g, 6.0 equivalents) was added dropwise at room temperature with stirring. The resulting solution was allowed to react for another 2 hours at room temperature with stirring. The resulting solution was extracted with 2×2000mL of dichloromethane, and the organic layers were combined. The mixture was dried over anhydrous sodium sulfate and concentrated under reduced pressure. The residue was applied to a silica gel column with ethyl acetate / petroleum ether (1:20-1:5). 73g (56%) of 46 was obtained as a yellow oil. MS m / z[M+Na] + (ESI):635.62.

[0118] Triethylamine (391.5 g, 2.37 mol, 10.0 equiv) was added to a solution of triethylamine trihydrofluoride (381.6 g, 2.37 mol, 10.0 equiv) in 730 mL of anhydrous tetrahydrofuran at room temperature under an inert atmosphere of argon. The resulting solution was stirred at room temperature for 10 minutes. A solution of 46 (145 g, 236.54 mmol, 1.0 equiv) in 730 mL of anhydrous tetrahydrofuran was added to the resulting solution. The reaction mixture was stirred at room temperature for 12 hours, diluted with 3000 mL of dichloromethane, and washed with 2×1000 mL of saturated aqueous sodium bicarbonate solution and 2×1000 mL of saturated aqueous sodium chloride solution, respectively. The organic layer was dried over anhydrous sodium sulfate, filtered and concentrated under reduced pressure. The residue was purified by flash preparative HPLC using the following conditions: column, C18 silica gel; mobile phase, water (containing 0.1% FA) and acetonitrile (10% acetonitrile to 100% in 12 minutes), UV 254 nm. Fractions were diluted with dichloromethane, dried over anhydrous sodium sulfate and concentrated under reduced pressure. 70 g (80%) of 47 was obtained as a light yellow oil. MS m / z [M+Na] + (ESI): 397.15. 1H NMR (300 MHz, CHLOROFORM-d) δ 7.93-7.80 (m, 4H), 7.57-7.46 (m, 3H), 5.77 (d, J = 3.9 Hz, 1H), 5.00 (d, J = 11.4 Hz, 1H), 4.77-4.64 (m, 2H), 4.23 (m, 1H), 3.92-3.73 (m, 5H), 2.29 (s, 3H), 2.03-1.87 (m, 1H), 1.64 (s, 3H), 1.40 (s, 3H).

[0119] To a solution of 47 (70 g, 187.17 mmol, 1.0 equiv) in 700 mL of anhydrous pyridine was added 4-methylbenzenesulfonyl chloride (107.12 g, 561.51 mmol, 3.0 equiv) at 0°C under an inert atmosphere of nitrogen. The reaction mixture was then stirred at room temperature for 12 hours. The reactants were quenched by the addition of water and concentrated under reduced pressure. The reaction mixture was diluted with 3000 mL of dichloromethane and washed with 1×1000 mL of saturated sodium bicarbonate and 2×1000 mL of saturated sodium chloride, respectively. The organic phase was dried over anhydrous sodium sulfate. The solid was filtered off and concentrated under reduced pressure. The crude product was purified by rapid preparative HPLC using the following conditions (IntelFlash-1): column, C18 silica gel; mobile phase, water (containing 0.04% NH4HCO3) and acetonitrile (10% acetonitrile to 100% in 15 minutes and maintained at 100% for 5 minutes); detector, UV 254 nm. 90 g (70%) of 48 was obtained as a light yellow solid. [M+Na+] + (ESI):705.81

[0120] To a solution of diethyl ((phenylthio)methyl)phosphonate (22.62 g, 87.00 mmol, 2.00 equiv) in 150 mL of anhydrous tetrahydrofuran was added butyl lithium (2M, 43.50 mL 2.00 equiv) dropwise at -78°C with stirring under an inert atmosphere of nitrogen. The resulting solution was stirred at -30°C for 2 hours. A solution of 48 (30 g, 43.99 mmol, 1.00 equiv) in 150 mL of tetrahydrofuran was added dropwise at -78°C with stirring. The resulting solution was stirred at room temperature for 12 hours. The resulting solution was extracted with ethyl acetate and the organic layers were combined. The organic layers were washed with 1×500 mL of water and 2×500 mL of saturated sodium chloride, respectively. The mixture was dried over anhydrous sodium sulfate and concentrated under reduced pressure. The crude product was purified by flash preparative HPLC using the following conditions (IntelFlash-1): column, C18 silica gel; mobile phase, water (containing 0.04% NH4HCO3) and acetonitrile (30% acetonitrile to 100% in 10 minutes and maintained at 100% for 5 minutes); detector, UV 254nm. 12.5 g (50%) of 201 was obtained as a light yellow solid. [M+H+] + (ESI):599.66

[0121] To a solution of 201 (15 g, 25.08 mmol, 1.0 equiv) in 300 mL of toluene was added 2,2'-azobis(2-methylpropionitrile) (3.29 g, 20.07 mmol, 0.4 equiv) and tributyltinane (14.6 g, 50.16 mmol, 2.0 equiv) at room temperature under an inert atmosphere of nitrogen. The resulting solution was stirred at 110 °C for 1 hour. The resulting mixture was concentrated under reduced pressure. The crude product was purified by rapid preparative HPLC using the following conditions: column, C18 silica gel; mobile phase, water (containing 0.04% NH4HCO3) and acetonitrile (30% acetonitrile reached 100% in 10 minutes and maintained at 100% for 5 minutes); detector, UV 254 nm. 8.5 g (65%) of 91 was obtained as a light yellow solid. [M+H+] + (ESI):491.15.1H NMR (400MHz, DMSO-d6) δ7.96-7.82(m,4H),7.57-7.43(m,3H),5.80-5.71(m,1H),4.85-4.72(m,2H),4.64(d,J=12.2Hz,1H),4.02 -3.78(m,5H),3.51-3.44(m,1H),2.65-2.58(m,2H),2.21-1.85(m,4H),1.48(d,J=5.0Hz,3H),1.31(d,J=3.1Hz,3H),1.19(m,6H). 31P NMR (162MHz, DMSO) δ33.08, 30.36.

[0122] To a solution of 91 (17 g, 34.69 mmol, 1.0 equiv) in 170 mL of acetic acid was added acetoacetate (35.4 g, 346.94 mmol, 10.0 equiv) and sulfuric acid (679.9 mg, 6.94 mmol, 0.2 equiv) under an inert atmosphere of nitrogen. The resulting solution was stirred at room temperature for 2 hours. The reactants were then quenched by the addition of ice water. The resulting solution was extracted with ethyl acetate. The resulting mixture was washed with water and saturated aqueous sodium chloride solution. The mixture was dried over anhydrous sodium sulfate and concentrated under reduced pressure. The crude product was purified by rapid preparative HPLC using the following conditions: column, C18 silica gel; mobile phase, water and acetonitrile (20% acetonitrile reached 100% in 10 minutes and maintained at 100% for 5 minutes); detector, UV 254 nm. 14 g (75%) of 82 was obtained as a light yellow solid. MS m / z[M+H] + (ESI):535.72.

[0123] To a solution of uracil (5.87 g, 52.43 mmol, 2.0 equiv) in 140 mL of acetonitrile was added N,O-bis(trimethylsilyl)acetamide (23.95 g, 117.98 mmol, 4.5 equiv) at room temperature under an inert atmosphere of nitrogen. The resulting solution was stirred at 60°C for 0.5 h. 82 (14 g, 26.22 mmol, 1.00 equiv) was added thereto at 0°C, and then perchlorostannane (8.86 g, 34.08 mmol, 1.3 equiv) was added dropwise at 0°C with stirring. The resulting solution was allowed to react at 60°C for another hour with stirring. The reaction mixture was cooled to 0°C and quenched by adding 100 mL of saturated aqueous sodium bicarbonate solution. The resulting solution was extracted with 2×300 mL of ethyl acetate, and the organic layers were combined, dried over anhydrous sodium sulfate and concentrated under reduced pressure. The crude product was purified by flash preparative HPLC using the following conditions: column, C18 silica gel; mobile phase, water (containing 0.04% NH4HCO3) and acetonitrile (45% acetonitrile to 100% in 10 minutes and maintained at 100% for 5 minutes); detector, UV 254 nm. 9 g (60%) of 83 was obtained as a white solid. MS m / z [M+H] + (ESI):587.10.1H NMR (300MHz, DMSO-d6) δ11.43(s,1H),7.93(d,J=8.7Hz,3H),7.84(s,1H),7.68(d,J=13.4Hz, 1H),7.54(d,J=9.3Hz,2H),7.47(d,J=8.5Hz,1H),5.79(dd,J=12.3,3.8Hz,1H),5.68(d,J=8. 0Hz,1H),5.49-5.42(m,1H),4.68(d,J=2.7Hz,2H),4.11(t,J=6.3Hz,1H),3.98-3.90(m,5H), 2.78-2.56(m,2H),2.10(s,4H),2.04-1.98(m,1H),1.24(d,J=7.0Hz,2H),1.21-1.14(m,6H). 31P NMR (121MHz, DMSO) δ32.76, 30.94, 30.15, 30.14.

[0124] To a solution of 83 (9 g, 15.36 mmol, 1.0 eq.) in 90 mL of ethanol was added sodium methoxide (1.24 g, 23.04 mmol, 1.5 eq.) at 0°C under an inert atmosphere of nitrogen. The resulting solution was stirred at room temperature for 2 hours. The mixture was then adjusted to pH = 7 by Amberlite IR-120 (H+). The resulting mixture was filtered and the filter cake was washed with 3×50 mL of ethanol. The filtrate was concentrated under reduced pressure. The 9 g crude product was used in the next step without further purification. MS m / z [M+H] + (ESI):549.32.

[0125] At 0°C, sodium hydride (737.28 mg, 30.72 mmol, 2.0 equiv) was added to a solution of 84 (15.36 mmol, 1.0 equiv) in 70 mL of tetrahydrofuran under an inert atmosphere of nitrogen. The mixture was stirred at 0°C for 30 minutes. A solution of iodomethane (4.33 g, 30.72 mmol, 2.0 equiv) in 20 mL of tetrahydrofuran was added dropwise at 0°C. The resulting solution was stirred at room temperature for 2 hours. The reactants were quenched with 50 mL of water. The resulting solution was extracted with 200 mL of ethyl acetate, and the organic phases were combined. The organic phases were washed with water, then dried over anhydrous sodium sulfate and concentrated under reduced pressure. The crude product was purified by flash preparative HPLC using the following conditions (IntelFlash-1): column, C18 silica gel; mobile phase, water (containing 0.04% NH4HCO3) and acetonitrile (20% acetonitrile to 100% in 25 minutes and maintained at 100% for 5 minutes); detector, UV 254nm. 5 g (60% in two steps) of 416 were obtained as a white solid. MS m / z [M+H] + (ESI):559.41.

[0126] At room temperature, 2,3-dichloro-5,6-dicyano-1,4-benzoquinone (4.6 g, 20.06 mmol, 1.4 equiv) was added to a solution of 416 (6.0 g, 14.33 mmol, 1.0 equiv) in 72 mL of dichloromethane and 8 mL of water under an inert atmosphere of nitrogen. The resulting solution was stirred at room temperature for 1 hour. The reaction mixture was diluted with 40 mL of water and extracted with 200 mL of dichloromethane. The aqueous phase was concentrated under reduced pressure. The crude product was purified by rapid preparative HPLC using the following conditions: column, C18 silica gel; mobile phase, water (containing 0.04% NH4HCO3) and acetonitrile (5% acetonitrile reached 50% in 15 minutes and maintained at 100% for 5 minutes); detector, UV 254 nm. A mixture of 3 g (60%) was obtained as a white solid and separated by SFC. 1.5 g of 417 was obtained as a white solid. MSm / z[M+H] + (ESI):419.00.31 P NMR (162 MHz, DMSO) δ 30.27. 1 H NMR (400MHz, DMSO-d6) δ11.39(d,J=2.3Hz,1H),7.60(d,J=8.0Hz,1H),5.77-5.74(m,1H),5.66(dd,J=8.1,2.2Hz,1H),5.17(s,1H)4.07 -3.89(m,4H),3.89-3.79(m,2H),3.68(dd,J=7.4,5.1Hz,1H),3.34(s,3H),2.69-2.61(m,2H),2.15-1.97(m,4H),1.20(t,J=7.1Hz,6H).

[0127] 1.2 g of 417S (for 100) was obtained as a white solid. MS m / z [M+H] + (ESI):419.00. 31 P NMR (162 MHz, DMSO) δ 32.80 1 H NMR (400MHz, DMSO-d6) δ11.39(d,J=2.2Hz,1H),7.57(d,J=8.1Hz,1H),5.79-5.76(m,1H),5.67(dd,J=8.0,2.2Hz,1H),5.17(s,1H ),4.07-3.90(m,4H),3.90-3.77(m,3H),3.34(s,3H),2.63(q,J=8.8,7.4Hz,2H),2.26-2.02(m,4H),1.22(td,J=7.1,0.8Hz,6H).

[0128] At room temperature, 2-cyanoethyl N, N, N', N'-tetraisopropyl diaminophosphorate (1.41 g, 4.7 mmol, 1.3 equiv) and 4,5-dicyanoimidazole (467.3 mg, 3.9 mmol, 1.1 equiv) were added to a solution of 417 (1.5 g, 3.6 mmol, 1.0 equiv) in 15 mL of dichloromethane in sequence under an argon inert atmosphere. The reaction mixture was stirred at room temperature for 1 hour. The reaction solution was diluted with 100 mL of dichloromethane and washed with 1 × 20 mL of saturated sodium bicarbonate aqueous solution and 1 × 20 mL of saturated sodium chloride aqueous solution, respectively. The organic phase was dried over anhydrous sodium sulfate, filtered and concentrated until no residual solvent was left under reduced pressure. The residue was purified by flash preparative HPLC using the following conditions: column, C18 silica gel; mobile phase, water (containing 0.04% NH4HCO3) and acetonitrile (20% acetonitrile to 100% in 10 minutes, 100% acetonitrile for 5 minutes). The organic layer was separated. The aqueous phase was extracted with 3×50 mL of dichloromethane. The organic phases were combined and dried over anhydrous sodium sulfate. The solid was filtered out. The filtrate was concentrated at 25° C. to obtain the product, which was dried under reduced pressure at 25° C. for 6 hours. 1.38 g (64%) of 2-0 was obtained as a white solid. MS m / z [MH] - (ESI):617.15. 1 H NMR (300MHz, chloroform-d) δ9.24(s,1H),7.40-7.36(m,1H),5.82-5.72(m,2H),4.14-4.03(m,5H),4.01-3.81(m,3H), 3.76-3.56(m,3H),3.51(m,3H),2.80-2.59(m,4H),2.33-2.22(m,4H),1.33-1.28(m,6H),1.22-1.17(m,12H). 31 P NMR (121MHz, CDCl3) δ149.84, 149.66, 29.89, 29.55.

[0129]

[0130] At room temperature, 2-cyanoethyl N,N,N',N'-tetraisopropyl diaminophosphorate (1.1 g, 3.8 mmol, 1.3 equiv) and 4,5-dicyanoimidazole (376.4 mg, 3.2 mmol, 1.1 equiv) were added to a solution of 417 (1.2 g, 2.9 mmol, 1.0 equiv) in 20 mL of dichloromethane in sequence under an inert atmosphere of argon. The reaction mixture was stirred at room temperature for 1 hour. The reaction solution was diluted with 20 mL of dichloromethane and washed with saturated aqueous sodium bicarbonate solution and saturated aqueous sodium chloride solution, respectively. The organic phase was dried over anhydrous sodium sulfate, filtered and concentrated until no residual solvent was left under reduced pressure. The residue was purified by rapid preparative HPLC using the following conditions: column, C18 silica gel; mobile phase, water (containing 0.04% NH4HCO3) and acetonitrile (20% acetonitrile reached 100% in 10 minutes, 100% acetonitrile was maintained for 5 minutes). The organic phase layer was separated. The aqueous phase was extracted with 3×50 mL of dichloromethane. The organic phases were combined and dried over anhydrous sodium sulfate. The solid was filtered off. The filtrate was concentrated at 25° C. to obtain the product, which was dried under reduced pressure at 25° C. for 4 hours. 990.9 mg (70%) of 2-100 was obtained as a white solid. MS m / z [MH] - (ESI):617.25. 1 H NMR (300MHz, chloroform-d) δ9.28(s,1H),7.30-7.23(m,1H),5.85(dd,J=7.7,3.2Hz,1H),5.77(dd,J=8.1,1.2Hz,1H),4.19-4.05(m,5H),4.02 -3.55(m,6H),3.51(d,J=11.7Hz,3H),2.85-2.71(m,1H),2.69-2.58(m,3H),2.56-2.18(m,4H),1.37-1.27(m,6H),1.25-1.11(m,12H). 31 PNMR(121MHz,CDCl3)δ149.70,149.69,32.96,32.65.

[0131] Example 2-3

[0132]

[0133] At 0 ° C, sulfuric acid (39.19 g, 0.4 mol, 0.1 eq.) was slowly added to a solution of (3R, 4S, 5R)-5-(hydroxymethyl)tetrahydrofuran-2,3,4-triol (600 g, 3.997 mol, 1.0 eq.) in 6 L of methanol under an inert atmosphere of argon. The reaction mixture was allowed to warm to room temperature and stirred overnight. The mixture was adjusted to pH 7 with sodium bicarbonate, then filtered and concentrated under reduced pressure to give 21 (520 g, crude) as a yellow oil. MS m / z [M+H] + (ESI): 165.07. The crude product was used in the next step without further purification.

[0134] At 0 ° C, 1,3-dichloro-1,1,3,3-tetraisopropyldisiloxane (549.5 g, 1.742 mol, 1.1 equiv) was added to a solution of 21 (260 g, 1.584 mol, 1.0 equiv) in 2.6 L of pyridine under an inert atmosphere of argon. The mixture was stirred at room temperature for 1 hour. The resulting solution was extracted with 3×4 L of ethyl acetate, and the organic layers were combined. The organic layers were washed with 2×500 mL of water and 2×4 L of saturated sodium chloride, respectively. The mixture was dried over anhydrous sodium sulfate and concentrated under reduced pressure. The residue was dissolved in 100 mL of ethyl acetate and evaporated to dryness. The residue was applied to a silica gel column with ethyl acetate / petroleum ether (1:5-1:3). 195 g (33%) of 22 was obtained as a yellow oil. MS m / z[M+H] + (ESI):407.23.

[0135] Under an argon inert atmosphere, sodium hydride (23.1 g, 0.959 mol, 2.0 equiv) was added to 22 (195 g, 0.48 mol, 1.0 equiv) in a mixture solution of 2L 1,3-dimethyl-2-imidazolidinone and 1L iodomethane. The resulting solution was stirred at room temperature for 40 minutes. The resulting solution was diluted with dichloromethane and washed with water. The organic layer was dried over anhydrous sodium sulfate, filtered and concentrated under reduced pressure. The crude product was purified by column chromatography (silica gel, ethyl acetate / petroleum ether=1:5 to 1:1) to obtain 23 (390 g, crude product) as a yellow oil. MS m / z[M+H] + (ESI): 421.25. H NMR (400 MHz, CHLOROFORM-d) δ 4.77 (s, 1H), 4.50 (m, 1H), 4.01 (m, 2H), 3.94-3.84 (m, 1H), 3.61 (d, J = 4.3 Hz, 1H), 3.59 (s, 3H), 3.34 (s, 3H), 1.15-1.02 (m, 28H). The crude product was used in the next step without further purification.

[0136] At room temperature, triethylamine (3.7kg, 37.082mol, 20.0 equivalents) was added to a solution of triethylamine trifluoride (3kg, 18.541mol, 10.0 equivalents) in 5L of anhydrous tetrahydrofuran under an inert atmosphere of argon. The resulting solution was stirred at room temperature for 10 minutes. A solution of 23 (780g, 1.854mol, 1.0 equivalents) in 1L of anhydrous tetrahydrofuran was added to the resulting solution. The reaction mixture was stirred overnight at room temperature and diluted with 3L of dichloromethane, washed with 2×1L of saturated sodium bicarbonate aqueous solution and 2×1L of saturated sodium chloride aqueous solution, respectively. The organic layer was dried over anhydrous sodium sulfate, filtered and concentrated under reduced pressure. The crude product was purified by column chromatography (silica gel, ethyl acetate / petroleum ether=1:5 to 3:1) to obtain 24 (280g) as a yellow oil. MS m / z[M+H] + (ESI):179.11.

[0137] At 0 ° C, 4,4'-(chloro(phenyl)methylene)bis(methoxybenzene) (292.8 g, 0.865 mol, 1.1 equiv) was added to a solution of crude 24 (140 g, 0.786 mol, 1.0 equiv) in 1.4 L pyridine under an inert atmosphere of argon. The mixture solution was stirred at room temperature for 12 hours. The resulting solution was diluted with 3000 mL of dichloromethane and washed with 2×1000 mL of water. The organic phase was dried over anhydrous sodium sulfate, filtered and concentrated under reduced pressure. The residue was applied to a silica gel column with ethyl acetate / petroleum ether (1:50-1:1). 230 g (61%) of 25 was obtained as a yellow oil. MS m / z[M+H] + (ESI):481.23.

[0138] At room temperature, tert-butylchlorodiphenylsilane (289.4 g, 1.053 mol, 1.1 equiv) and imidazole (162.9 g, 2.393 mol, 2.5 equiv) were added to a solution of 25 (460 g, 0.957 mol, 1.00 equiv) in 4.6 L N,N-dimethylformamide under an inert atmosphere of nitrogen. The resulting solution was stirred at room temperature for 12 hours. The organic layer was washed with water and extracted with 3×4 L ethyl acetate. The organic layer was dried over anhydrous sodium sulfate and concentrated under reduced pressure. The residue was applied to a silica gel column with ethyl acetate / petroleum ether (1:20-1:5). 520 g (72%) of 26 was obtained as a yellow oil. MS m / z [M+H] + (ESI):719.35.

[0139] At room temperature, triethylsilane (500 mL) was added to a solution of 26 (520 g, 166.910 mmol, 1.0 equiv) in 5.2 L of dichloromethane. Trifluoroacetic acid (500 mL) was then added at 0 ° C, and the resulting solution was stirred at room temperature for 2 hours. The reactant was quenched by adding 5 L of saturated sodium bicarbonate and extracted with 3×2 L of dichloromethane. The organic layers were combined, dried over anhydrous sodium sulfate, filtered and concentrated under reduced pressure. The residue was applied to a silica gel column with ethyl acetate / petroleum ether (1: 10-5: 1). The fractions of the desired product were concentrated under reduced pressure. 260 g (82%) of 27 were obtained as a yellow oil. MS m / z[M+NH4] + (ESI):434.35. H NMR (400MHz, DMSO-d6) δ7.81-7.34(m,10H),4.79(d,J=1.5Hz,1H),4.65(s,1H),4.16(m,1H),3.91(m ,1H),3.44(dd,J=11.6,3.6Hz,1H),3.22(m,1H),3.19(s,3H),3.11(s,3H),3.03(m,1H),1.03(s,9H).

[0140] To a solution of oxalyl chloride (47.3 g, 372.6 mmol, 2.0 equiv) in 1.3 L of dichloromethane was added dimethyl sulfoxide (43.7 g, 558.9 mmol, 3.0 equiv) at -78°C under an inert atmosphere of nitrogen. The resulting solution was stirred at -78°C for 0.5 h, and then a solution of 27 (75 g, 186.3 mmol, 1.0 equiv) in 750 mL of dichloromethane was added dropwise at -78°C with stirring. The resulting solution was stirred at -78°C for 3 h. Triethylamine (84.8 g, 838.4 mmol, 4.5 equiv) was then added dropwise at -78°C with stirring over 0.5 h. The resulting solution was diluted with 2000 mL of dichloromethane. The resulting mixture was washed with 1×1 L of saturated aqueous sodium bicarbonate solution and 1×1 L of saturated aqueous sodium chloride solution, respectively. The organic phase was dried over anhydrous sodium sulfate, filtered and concentrated under reduced pressure. The above process was repeated twice to obtain 120 g (crude) 28 as a yellow oil. The crude product was used in the next step without further purification. MS m / z [M+H] + (ESI):415.21.

[0141] At -78 ° C, methylmagnesium bromide (103.549 g, 868.4 mmol, 3.0 equiv) was added to a solution of 28 (120 g, 289.5 mmol, 1.0 equiv) in 1.2 L of tetrahydrofuran under an inert atmosphere of nitrogen. The resulting solution was stirred at -78 ° C for 2 hours. The resulting solution was diluted with 3 L of dichloromethane and washed with 2×1000 mL of water. The organic phase was dried over anhydrous sodium sulfate and concentrated under reduced pressure. The residue was applied to a silica gel column with ethyl acetate / petroleum ether (1: 10-5: 1). The fractions of the desired product were concentrated under reduced pressure. 86 g (62%) of 3-1 were obtained as a yellow oil. MS m / z[M+H] + (ESI):431.25.

[0142] At -78 ° C, dimethyl sulfoxide (46.8 g, 599.12 mmol, 3.0 equivalents) was added to a solution of oxalyl chloride (50.7 g, 399.4 mmol, 2.0 equivalents) in 860 mL of dichloromethane under an inert atmosphere of nitrogen. The resulting solution was stirred at -78 ° C for 0.5 hours. And 3-1 (86 g, 199.7 mmol, 1.00 equivalents) was added. The resulting solution was stirred at -78 ° C for 3 hours. Then, at -78 ° C, triethylamine (90.9 g, 898.7 mmol, 4.5 equivalents) was added dropwise with stirring. The resulting solution was stirred at -78 ° C for 0.5 hours. The resulting solution was diluted with 1 L of dichloromethane and washed with 3×600 mL of water. The organic layer was dried over anhydrous sodium sulfate, filtered and concentrated under reduced pressure. The residue was applied to a silica gel column with ethyl acetate / petroleum ether (1:50-3:1). The desired product fractions were concentrated under reduced pressure. 72 g (80%) of 3-2 were obtained as a yellow oil. MS m / z [M+H] + (ESI):429.22. H NMR (300MHz, DMSO-d6) δ7.71-7.30 (m, 10H), 4.94 (d, J = 1.2Hz, 1H), 4.36 (m, 1H), 4. 27(d,J=6.8Hz,1H),3.12(s,3H),3.01(m,1H),1.99(d,J=4.9Hz,3H),1.01(s,9H).

[0143] Under an inert atmosphere of nitrogen, 3-2 (72g, 168.0mmol, 1.0 equivalent) was added to a solution of 720mL N,N-dimethylformamide dimethyl acetal. The resulting solution was stirred at 110°C for 17 hours, then diluted with 1L dichloromethane and washed with 2×600mL water. The resulting mixture was concentrated under reduced pressure. The residue was applied to a silica gel column with ethyl acetate / petroleum ether (1:50-5:1). The fractions of the desired product were concentrated under reduced pressure. 60g (70%) of 4-1 was obtained as a yellow oil. MS m / z[M+H] + (ESI):484.26.

[0144] At room temperature, diethyl phosphonate (17.1 g, 124.1 mmol, 2.0 equiv.), manganese triacetate dihydrate (49.9 g, 186.1 mmol, 3.0 equiv.) were added to a solution of 4-1 (30 g, 62.026 mmol, 1.0 equiv.) in 100 mL of acetic acid and 200 mL of 1.4-dioxane under an inert atmosphere of nitrogen. The resulting solution was stirred at 80° C. for 20 minutes. The resulting solution was extracted with 3×500 mL of ethyl acetate, and the organic layers were combined. The organic layers were washed with 2×200 mL of water and 2×200 mL of saturated sodium chloride, respectively. The mixture was dried over anhydrous sodium sulfate and concentrated under reduced pressure. The above procedure was repeated 2 times. The crude product was purified by flash preparative HPLC using the following conditions (IntelFlash-1): column, C18 silica gel; mobile phase, water and acetonitrile (30% acetonitrile to 100% in 10 minutes and maintained at 100% for 5 minutes); detector, UV 254 nm. 38 g (52%) of 4-52 was obtained as a yellow oil. MS m / z [M+H] + (ESI):565.25.

[0145] To a solution of 4-52 (38 g, 67.3 mmol, 1.0 equiv) in 380 mL of 1,1-dimethoxy-N,N-dimethylmethanamine. The resulting solution was stirred at 80 ° C for 12 hours. The resulting solution was diluted with 500 mL of dichloromethane and washed with 2×300 mL of water. The crude product was purified by rapid preparative HPLC using the following conditions (IntelFlash-1): column, C18 silica gel; mobile phase, water (containing 0.04% NH4HCO3) and acetonitrile (15% acetonitrile reached 100% in 25 minutes and maintained at 100% for 5 minutes); detector, UV 254nm. 27 g (62%) of 4-53 was obtained as a yellow solid. MS m / z[M+H] + (ESI):620.30.

[0146] To a solution of 4-53 (27 g, 43.56 mmol, 1.0 equiv) in 270 mL of ethanol was added hydroxylamine hydrochloride (9.1 g, 130.7 mmol, 3.0 equiv) and pyridine (34.5 g, 435.6 mmol, 10.0 equiv) at room temperature under an inert atmosphere of nitrogen. The final reaction mixture was stirred at 60 ° C for 1 hour. The resulting solution was diluted with 400 mL of dichloromethane and washed with 2×200 mL of water. The crude product was purified by fast preparative HPLC with the following conditions (IntelFlash-1): column, C18 silica gel; mobile phase, water and acetonitrile (30% acetonitrile reached 100% in 10 minutes and maintained at 100% for 5 minutes); detector, UV 254 nm. 16 g (59.2%) of 4-4 was obtained as a yellow oil. MS m / z [MH] - (ESI):588.25.

[0147] To a solution of 4-4 (16 g, 42.395 mmol, 1.0 equiv) in 160 mL of acetic acid and 80 mL of acetic anhydride was added sulfuric acid (133.1 mg, 1.357 mmol, 0.05 equiv) dropwise with stirring at 0°C. The resulting solution was stirred at room temperature for 1 hour, then diluted with 300 mL of dichloromethane and washed with 2×100 mL of water. The crude product was purified by flash preparative HPLC using the following conditions (IntelFlash-1): column, C18 silica gel; mobile phase, water (containing 0.04% NH4HCO3) and acetonitrile (15% acetonitrile to 100% in 25 minutes and maintained at 100% for 5 minutes); detector, UV 254 nm. 12 g (68%) of 4-24 was obtained as a yellow oil. MS m / z [M+H] + (ESI):618.24.

[0148] At room temperature, N,O-bis(trimethylsilyl)acetamide (2.305 g, 11.3 mmol, 3.5 equiv) was added to a solution of uracil (0.544 g, 4.86 mmol, 1.5 equiv) in 80 mL of acetonitrile under an inert atmosphere of nitrogen. The resulting solution was stirred at 60°C for 0.5 h. The reaction mixture was cooled to 0°C and 4-4 (2 g, 3.24 mmol, 1.0 equiv) was added. Then, trimethylsilyl trifluoromethanesulfonate (1.44 g, 6.48 mmol, 2.0 equiv) was added dropwise at 0°C with stirring. The resulting solution was reacted at 60°C with stirring for 1 h. The resulting solution was diluted with dichloromethane and washed with water. The above procedure was repeated 6 times. The crude product was purified by flash preparative HPLC using the following conditions (IntelFlash-1): column, C18 silica gel; mobile phase, water (containing 0.04% NH4HCO3) and acetonitrile (30% acetonitrile to 100% in 10 minutes and maintained at 100% for 5 minutes); detector, UV 254nm. 3.28 g (24%) of 4-5 was obtained as an off-white solid. MS m / z [M+H] + (ESI):670.25.

[0149] At room temperature, triethylamine (9.9 g, 97.9 mmol, 20.0 equiv) was added to a solution of triethylamine trihydrofluoride (7.9 g, 49.0 mmol, 10.0 equiv) in 20 mL of anhydrous tetrahydrofuran under an inert atmosphere of argon. The resulting solution was stirred at room temperature for 10 minutes. A solution of 4-5 (3.28 g, 4.9 mmol, 1.0 equiv) in 12 mL of anhydrous tetrahydrofuran was added to the resulting solution. The reaction mixture was stirred overnight at room temperature and diluted with 200 mL of dichloromethane, washed with 2×200 mL of saturated aqueous sodium bicarbonate solution and 2×200 mL of saturated aqueous sodium chloride solution, respectively. The organic phase was dried over anhydrous sodium sulfate, filtered and concentrated under reduced pressure. The crude product was purified by flash preparative HPLC using the following conditions (IntelFlash-1): column, C18 silica gel; mobile phase, water (containing 0.04% NH4HCO3) and acetonitrile (30% acetonitrile to 100% in 10 minutes and maintained at 100% for 5 minutes); detector, UV 254nm. 1.3 g (62%) of 4-6 was obtained as a yellow oil. MS m / z [M+H] +(ESI):432.15. HNMR (400MHz, DMSO-d6) δ11.50-11.29(m,1H),8.89(s,1H),7.76(d,J=8.1Hz,1H),6.45(d,J=5.0Hz,1H),6.05(d,J=6.0Hz, 1H), 5.66 (m, 1H), 5.52 (d, J = 6.1Hz, 1H), 4.68 (m, 1H), 4.16 (t, J = 4.8Hz, 1H), 4.07 (m, 3.7Hz, 4H), 3.36 (s, 3H), 1.24 (m, 6H).

[0150] To a solution of 4-6 (1 g, 2.318 mmol, 1.0 equiv) in 10 mL of dichloromethane was added 2-cyanoethyl N,N,N',N'-tetraisopropyl diaminophosphorate (838.6 mg, 2.782 mmol, 1.2 equiv) and 4,5-dicyanoimidazole (273.8 mg, 2.318 mmol, 1.0 equiv) at room temperature. The resulting solution was stirred at room temperature for 1 hour. The reaction solution was diluted with 300 ml of dichloromethane and washed with 2×100 mL of saturated aqueous sodium bicarbonate solution and 1×100 mL of saturated aqueous sodium chloride solution, respectively. The organic phase was dried over anhydrous sodium sulfate, filtered and concentrated until no residual solvent was left under reduced pressure. The crude product was purified by flash preparative HPLC using the following conditions (IntelFlash-1): column, C18 silica gel; mobile phase, water (containing 0.04% NH4HCO3) and acetonitrile (30% acetonitrile to 100% in 15 minutes and maintained at 100% for 5 minutes); detector, UV 254nm. The fractions were diluted with an equal volume of dichloromethane. The organic phase layer was separated. The aqueous phase was extracted with 3×50 mL dichloromethane. The organic phases were combined and dried over anhydrous sodium sulfate. The solid was filtered off. The filtrate was concentrated at 25°C. 840.5 mg (57%) of 4-0 was obtained as a white solid. MS m / z [MH] -(ESI):630.20. H NMR (300MHz, DMSO-d6) δ11.43(s,1H),8.90(s,1H),7.65(m,1H),6.48(d,J=4.4Hz,1H),5.80-5.56(m,2H),4.95(m,1H),4.24(m,1H),4.16-3.95( m,4H),3.82-3.63(m,2H),3.62-3.40(m,2H),3.37(d,J=3.5Hz,3H),2.76 (m,2H),1.32-1.16(m,7H),1.09(t,J=6.8Hz,8H),0.96(d,J=6.8Hz,3H). P NMR (121MHz, DMSO) δ150.00,149.96,7.37,7.35.

[0151] Embodiment 2-4

[0152]

[0153] Under an argon inert atmosphere, calcium carbonate (125 g, 0.71 mol, 1.0 equiv) was slowly added to a solution of L-ascorbic acid (125 g, 0.71 mol, 1.0 equiv) in 1 L of water over 30 minutes. At 0 ° C, under stirring, hydrogen peroxide (250 mL, 30% aqueous solution) was added dropwise to the resulting solution within 1 hour. The reaction mixture was allowed to warm to room temperature and stirred overnight. Then, the reaction mixture was filtered and the filter cake was washed with 2 × 100 mL of water. The filtrate was treated with activated carbon (25 g) and then heated to 70 ° C. The hot suspension was filtered and the solid material was washed with 2 × 50 mL of water. The filtrates were combined and crystallized by adding 2 volume equivalents of methanol while stirring at 4 ° C for 16 hours. The solid material was filtered, washed with 2 × 50 mL of methanol and dried under high vacuum at 40 ° C. 88.1 g (75%) of 1 was obtained as a white solid. No MS signal. 1 H NMR (400MHz, D2O): δ3.98 (d, J = 2.2Hz, 1H), 3.88-3.72 (m, 1H), 3.62-3.49 (m, 2H).

[0154] At room temperature, anhydrous oxalic acid (28.8 g, 0.32 mol, 1.0 equiv) and p-toluenesulfonic acid monohydrate (1.0 g, 0.01 equiv) were added to a solution of 1 (100 g, 0.30 mol, 1.0 equiv) in 500 mL of anhydrous acetonitrile in an inert atmosphere of argon. The mixture was stirred at reflux for 3 hours. The hot mixture was cooled to room temperature and filtered. The filter cake was washed with 50 mL of acetonitrile, and the combined filtrate was evaporated under reduced pressure. The residue was dissolved in 100 mL of ethyl acetate and evaporated to dryness. 50.1 g (70%) of 2 was obtained as a white solid. MS m / z [M+H] + (ESI): 119. It was used in the next step without further purification.

[0155] Benzoyl chloride (72.0 mL, 0.62 mol, 1.1 equiv) was added dropwise to a mixture solution of 2 (66.5 g, 0.56 mol, 1.0 equiv) in 1200 mL of dichloromethane and 135 mL of anhydrous pyridine at 0 ° C. under an inert atmosphere of argon. The resulting solution was stirred at 0 ° C for 30 minutes. The reaction mixture was quenched with 1N HCl and washed with 3×200 mL of saturated aqueous sodium chloride solution. 2 volume equivalents of hexane were added to the organic layer within 1 hour and stirred at 0 ° C for 16 hours. The desired product was collected by filtration and dried under vacuum. 75.1 g (60%) of 3 was obtained as a white solid. MS m / z[M+H] + (ESI):223. 1 H NMR (400MHz, DMSO-d6): δ8.08-7.98(m,2H),7.78-7.67(m,1H),7.59-7.51(m,2H),6.12(d,J= 17.9Hz, 1H), 5.73 (d, J = 7.9Hz, 1H), 4.74-4.71 (m, 1H), 4.55-4.44 (m, 1H), 4.09-4.03 (m, 1H).

[0156] At 0°C, 4-dimethylaminopyridine (60 mg, 6.0 mmol, 0.01 equiv), imidazole (10.2 g, 150 mmol, 2.0 equiv) and tert-butyldiphenylsilyl chloride (21.4 g, 75.0 mmol, 1.05 equiv) were added to a solution of 3 (16.5 g, 74.3 mmol, 1.0 equiv) in 160 mL of dichloromethane in sequence under an inert atmosphere of argon. The reaction mixture was warmed to room temperature and stirred overnight. The resulting solution was evaporated under reduced pressure. The residue was dissolved in 300 mL of hexane and washed with 1×100 mL of 1N HCl, 3×100 mL of water and 3×100 mL of saturated aqueous sodium chloride solution, respectively. The organic phase was dried over anhydrous sodium sulfate, filtered and concentrated under reduced pressure. It was used in the next step without further purification. MS m / z[M+H] + (ESI):461.

[0157] At -78 ° C, under an argon inert atmosphere, diisobutylaluminum hydride (100 mL, 100 mmol, 1 M in toluene) was added dropwise to a solution of crude 4 (35.0 g, 76 mmol, 1.0 eq.) in 150 mL of 1,2-dimethoxyethane over 10 minutes. The mixture solution was stirred at -78 ° C for 30 minutes. The reaction was monitored by TLC and used in the next step without further purification.

[0158] A pre-prepared solution of acetic anhydride (35 mL, 367 mmol, 5.0 equiv) and 4-dimethylaminopyridine (14.0 g, 115 mmol, 1.5 equiv) in 40 mL of dichloromethane was added dropwise to the solution of 5 at -78°C under an inert atmosphere of argon. After stirring for 10 minutes, the reaction mixture was warmed to room temperature and stirred for 2 hours. The mixture solution was diluted with 200 mL of hexane and poured into 200 mL of cold 1N HCl aqueous solution. The organic layer was washed with 3×100 mL of water, 3×100 mL of saturated sodium bicarbonate aqueous solution and 3×100 mL of saturated sodium chloride aqueous solution, respectively. The organic layer was dried over anhydrous sodium sulfate, filtered and concentrated under reduced pressure. The crude product was purified by rapid preparative HPLC using the following conditions: column, C18 silica gel; mobile phase, water (containing 0.1% FA) and acetonitrile (20% acetonitrile to 100% in 10 minutes, 100% acetonitrile for 8 minutes), UV 254nm. The fractions were diluted with dichloromethane and dried over anhydrous sodium sulfate. The solid was filtered off and the filtrate was concentrated under reduced pressure. 21.1 g (56% over three steps) of 6 were obtained as a colorless oil. MS m / z [M+H] + (ESI):505. 1H NMR (300MHz, DMSO-d6): δ7.89-7.82(m,2H),7.71-7.62(m,5H),7.57-7.28(m,8H),6.44-6.09(m,1H),5. 48-5.25(m,1H),4.76-4.59(m,1H),4.19-3.83(m,2H),2.14(s,2H),1.86(s,1H),1.04(d,J=15.2Hz,9H).

[0159] At room temperature, under an inert atmosphere of argon, uracil (3.3 g, 29.7 mmol, 1.5 eq) and N,O-bis(trimethylsilyl)acetamide (8.1 g, 39.6 mmol, 2.0 eq) were added to a solution of 6 (10.0 g, 59.4 mmol, 1.0 eq) in 50 mL of anhydrous acetonitrile in sequence. The mixture was stirred at 60°C for 30 minutes. Trimethylsilyl trifluoromethanesulfonate (7.0 g, 31.7 mmol, 1.5 eq) was then added dropwise and stirred at 60°C for another 2 hours. The reaction mixture was cooled to room temperature, diluted with 200 mL of ethyl acetate and poured into 100 mL of cold saturated aqueous sodium bicarbonate solution. The organic layer was washed with 3×100 mL of water and 3×100 mL of saturated aqueous sodium chloride solution, respectively. The organic layer was dried over anhydrous sodium sulfate, filtered and concentrated under reduced pressure. The crude product was used directly in the next step without further purification. MS m / z[M+H] + (ESI):557.

[0160] To a solution of triethylamine trihydrofluoride (28.9 g, 17.9 mmol, 10.0 equiv) in 50 mL of anhydrous tetrahydrofuran was added triethylamine (36.4 g, 35.9 mmol, 20.0 equiv) at room temperature under an inert atmosphere of argon. The resulting solution was stirred at room temperature for 10 minutes. A solution of 7 (10.0 g, 53.9 mmol, 1.0 equiv) in 50 mL of anhydrous tetrahydrofuran was added to the resulting solution. The reaction mixture was stirred at room temperature overnight and diluted with 300 mL of dichloromethane, washed with 2×100 mL of saturated aqueous sodium bicarbonate solution and 2×100 mL of saturated aqueous sodium chloride solution, respectively. The organic layer was dried over anhydrous sodium sulfate, filtered and concentrated under reduced pressure. The residue was purified by rapid preparative HPLC using the following conditions: column, C18 silica gel; mobile phase, water (containing 0.1% FA) and acetonitrile (10% acetonitrile to 100% in 12 minutes), UV 254 nm. The fractions were diluted with dichloromethane, dried over anhydrous sodium sulfate and concentrated under reduced pressure. 5.0 g (80% over two steps) of 8 was obtained as a pale yellow solid. MS m / z [M+H] + (ESI):319. 1H NMR (400MHz, DMSO-d6): δ11.35(d,J=2.2Hz,1H),8.05-7.94(m,2H),7.76(d,J=8.2Hz,1H),7.73-7.65(m,1H),7.5 4-7.32(m,2H),5.92-5.75(m,2H),5.67-5.46(m,1H),5.26(d,J=1.8Hz,1H),4.36-4.25(m,1H),4.14-3.89(m,2H).

[0161] To a solution of 8 (10.0 g, 31.4 mmol, 1.0 equiv) in 80 mL of anhydrous dichloromethane was added 2,4,6-collidine (22.8 g, 188.1 mmol, 6.0 equiv) and 4,4'-dimethoxytrityl chloride (26.6 g, 78.7 mmol, 2.5 equiv) in sequence at room temperature under an inert atmosphere of argon. The resulting solution was stirred at room temperature for 24 h. The reaction mixture was quenched by 10 mL of methanol, diluted with 500 mL of dichloromethane, and washed with 2×200 mL of saturated aqueous sodium bicarbonate solution and 2×200 mL of saturated aqueous sodium chloride solution, respectively. The organic layer was dried over anhydrous sodium sulfate, filtered and concentrated under reduced pressure. The residue was dissolved in 400 mL of ammonia (7M NH3 in methanol) and stirred at room temperature under an inert atmosphere of argon for 24 h. The reaction mixture was concentrated under reduced pressure. The residue was purified by flash preparative HPLC using the following conditions: column, C18 silica gel; mobile phase, water (containing 0.04% NH4HCO3) and acetonitrile (15% acetonitrile to 100% in 10 minutes, 100% acetonitrile for 6 minutes), UV 254nm. Fractions were diluted with dichloromethane, dried over anhydrous sodium sulfate and concentrated under reduced pressure. 12.1 g (75%) of 81 was obtained as a light yellow oil. MS m / z [MH] - (ESI):515.

[0162] To a solution of 81 (10.0 g, 19.4 mmol, 1.0 equiv) in 100 mL of anhydrous N,N-dimethylformamide was added imidazole (3.9 g, 57.3 mmol, 3.0 equiv) and tert-butyldimethylsilyl chloride (8.8 g, 58.1 mmol, 3.0 equiv) in sequence at room temperature under an inert atmosphere of argon. The resulting solution was stirred at room temperature for 12 hours. The reaction mixture was diluted with 500 mL of dichloromethane and washed with 2×200 mL of saturated aqueous sodium bicarbonate solution and 1×200 mL of saturated aqueous sodium chloride solution, respectively. The organic phase was dried over anhydrous sodium sulfate, filtered and concentrated under reduced pressure. The residue was dissolved in 120 mL of dichloromethane, trifluoroacetic acid (21.7 g, 190.5 mmol, 10.0 equiv) and triethylsilane (11.1 g, 95.2 mmol, 5.0 equiv) were added sequentially at room temperature, and stirred at room temperature under an inert atmosphere of argon for 1 hour. The reaction mixture was diluted with 400 mL of dichloromethane and washed with 2×150 mL of saturated aqueous sodium bicarbonate solution and 1×150 mL of saturated aqueous sodium chloride solution, respectively. The organic phase was dried over anhydrous sodium sulfate, filtered and concentrated under reduced pressure. The residue was purified by rapid preparative HPLC using the following conditions: column, C18 silica gel; mobile phase, water (containing 0.1% FA) and acetonitrile (15% acetonitrile reached 100% in 10 minutes, 100% acetonitrile was maintained for 6 minutes), UV 254 nm. The fractions were diluted with dichloromethane, dried over anhydrous sodium sulfate and concentrated under reduced pressure. 4.7 g (75%) of 82 was obtained as a light yellow solid. MS m / z[MH] - (ESI):515. 1 H NMR (300MHz, DMSO-d6): δ11.29 (s, 1H), 7.62 (d, J = 8.2Hz, 1H), 5.58-5.55 (m, 2H ), 5.43 (d, J = 2.6Hz, 1H), 4.17-3.94 (m, 4H), 0.86 (s, 9H), 0.10 (d, J = 4.0Hz, 6H).

[0163] At room temperature, 4-methylbenzenesulfonic acid (diethoxyphosphoryl) methyl ester (7.7 g, 23.9 mmol, 2.0 equivalents) was added to a solution of 82 (4.0 g, 11.9 mmol, 1.0 equivalents) in 40 mL of anhydrous tetrahydrofuran under an inert atmosphere of argon. At 0 ° C, sodium hydride (1.9 g, 47.7 mmol, 4.0 equivalents) was added to the reaction mixture and stirred at the same temperature for 3 hours under an inert atmosphere of argon. The reactant was quenched with 6 mL of acetic acid (1 M), diluted with 150 mL of ethyl acetate, and washed with 1 × 50 mL of saturated sodium chloride aqueous solution, respectively. The organic phase was dried over anhydrous sodium sulfate, filtered and concentrated under reduced pressure. The residue was applied to a silica gel column with ethyl acetate / petroleum ether (1: 10-5: 1). The fraction of the desired product was concentrated under reduced pressure. 2.5 g (60%) of 53 was obtained as a light yellow oil. MS m / z[M+H] + (ESI):479. 1 H NMR (400MHz, DMSO-d6): δ11.32(s,1H),7.53(d,J=8.1Hz,1H),5.60(d,J=1.1Hz,1H),5.51(d,J=8.1H z,1H),4.35-4.28(m,2H),4.07-3.84(m,8H),1.24-1.20(m,6H),0.87(s,9H),0.13(d,J=2.5Hz,6H). 31 P NMR (162 MHz, DMSO): δ 20.74.

[0164] A mixture solution of 53 (3.2 g, 6.7 mmol, 1.0 equiv) in 32 mL of formic acid and H2O (v / v=1:1) was stirred at room temperature under an inert atmosphere of argon for 24 hours. The resulting solution was concentrated under vacuum. The crude product was diluted with 300 mL of ethyl acetate and washed with 2×50 mL of water and 1×50 mL of saturated aqueous sodium bicarbonate solution, respectively. The organic phase was dried over anhydrous sodium sulfate, filtered and concentrated under vacuum. The residue was purified by rapid preparative HPLC using the following conditions: column, C18 silica gel; mobile phase, water and acetonitrile (15% acetonitrile reached 35% in 20 minutes), UV 254 nm. The fraction was diluted with dichloromethane, dried over anhydrous sodium sulfate and concentrated under reduced pressure. 1.9 g (80%) of 14 was obtained as a light yellow oil. MS m / z[M+H] + (ESI):365. 1HNMR (300MHz, DMSO-d6): δ11.31(s,1H),7.55(d,J=8.1Hz,1H),5.94(d,J=4.1Hz,1H),5.68(d,J=1.4Hz,1H),5.5 3(d,J=8.1Hz,1H),4.28(d,J=10.2Hz,1H),4.17(s,1H),4.09-3.97(m,6H),3.95-3.84(m,2H),1.23-1.09(m,6H). 31 P NMR (121 MHz, DMSO): δ 20.84.

[0165] At room temperature, 2-cyanoethyl N, N, N', N'-tetraisopropyl diaminophosphorate (2.4 g, 7.8 mmol, 1.3 equiv) and 4,5-dicyanoimidazole (785 mg, 6.6 mmol, 1.1 equiv) were added to a solution of 14 (2.2 g, 6.0 mmol, 1.0 equiv) in 20 mL of dichloromethane in sequence under an argon inert atmosphere. The reaction mixture was stirred at room temperature for 1 hour. The reaction solution was diluted with 200 mL of dichloromethane and washed with 2 × 100 mL of saturated sodium bicarbonate aqueous solution and 1 × 100 mL of saturated sodium chloride aqueous solution, respectively. The organic phase was dried over anhydrous sodium sulfate, filtered and concentrated until no residual solvent was left under reduced pressure. The residue was purified by flash preparative HPLC using the following conditions: column, C18 silica gel; mobile phase, water (containing 0.04% NH4HCO3) and acetonitrile (20% acetonitrile to 100% in 10 minutes, 100% acetonitrile for 5 minutes). The fractions were diluted with 500 mL of dichloromethane. The organic phase was dried over anhydrous sodium sulfate, filtered and concentrated under reduced pressure. 2.2 g (70%) of 100 was obtained as a light yellow oil. MS m / z [M+H] + (ESI):565. 1 H NMR (300MHz, acetonitrile-d3): δ9.05(s,1H),7.57-7.54(m,1H),5.88-5.83(m,1H),5.57-5.47(m,1H),4.55-4.41(m,1H),4.39-4.29 (m,1H),4.23-4.00(m,6H),3.95-3.76(m,4H),3.73-3.57(m,2H),2.75-2.65(m,2H),1.31-1.27(m,6H),1.22-1.19(m,12H). 31 P NMR (121MHz, CD3CN): δ151.05, 150.14, 19.82, 19.71.

[0166] Embodiment 2-5

[0167]

[0168] At 0 ° C, BnBr (118.0 g, 690.1 mmol) was added to a solution of 1 (120.0 g, 461.5 mmol) and KOH (31.0 g, 553.6 mmol) in DMF (500.0 mL) within 30 minutes, and then stirred at room temperature overnight. LC-MS and TLC monitored that 1 was completely consumed. The reactant was dissolved with water, and the pH was adjusted to pH=9 by gradually adding solid NaHCO3, and then extracted by EA (700.0 mL×3). The organic layer was washed with water (1.0 L×2) and anhydrous NaCl, dried over anhydrous Na2SO4 and evaporated in vacuo to give the crude product 2 (150.0 g, crude product) as a white oil. ESI-MS: m / z 334.1.

[0169] The crude product 2 (150.0 g, crude) was dissolved in 70% AcOH in H2O (700.0 mL) at room temperature, and then stirred at room temperature overnight. TLC showed that 2 was completely consumed. The reactant was quenched with solid NaHCO3 aqueous solution and extracted with EA (700.0 mL×3). The organic layer was washed with water (1.0 L×2) and NaCl aqueous solution, dried over anhydrous Na2SO4 and evaporated in vacuo to give a crude product, which was purified by column chromatography (SiO2, PE / EA=2:1 to 0:1) to give 3 (110.0 g, 90.5% purity, 76.8% yield) as a light yellow solid. ESI-MS: m / z 333.1; 1 HNMR (400MHz, CDCl3): δ7.36 (m, 5H), 5.76 (d, J = 3.72Hz, 1H), 4.78 (d, J = 11.28Hz, 1H), 4.60 (t, J = 4.04Hz, 1H), 4.5 6(d,J=11.28Hz,1H),4.11(m,1H),4.00(m,1H),3.93(m,1H),3.67(m,2H),2.55(s,2H),1.59(s,3H),1.36(s,3H).

[0170] To a stirred solution of 3 (110.g, 350.0mmol) in DCM (1.2L) was added a solution of NaIO4 (153.0g, 710mmol) in H2O (600.0mL) at room temperature, which was then stirred at room temperature for 1.0 hour. TLC monitored the work-up. The reactants were dissolved in water and extracted with DCM (700.0mL×3). The organic layer was washed with water (1.0L×2) and aqueous NaCl solution, dried over anhydrous Na2SO4 and concentrated under reduced pressure to give 4 (92.0g, crude) as a light yellow oil. ESI-MS: m / z 262.1; 1 HNMR (400MHz, CDCl3): δ9.60(d,J=1.76Hz,1H),7.35(m,5H),5.81(d,J=3.44Hz,1H),4. 74(d,J=12.12Hz,1H),4.60(m,2H),4.48(dd,J=1.68Hz and 9.24Hz,1H),3.85(q,J=4.28Hz 1H),1.60(s,3H),1.37(s,3H).

[0171] At 0°C, a solution of MeLi in diethoxymethane (1.6M, 287.0 mL) was added dropwise to a solution of 4 (85.0 g, crude) in THF (300.0 mL) over 0.5 hours, and the reaction was then stirred at 0°C for 2.5 hours. TLC showed that 4 was completely consumed. The reactant was quenched by aqueous NH4Cl solution and extracted by EA (400.0 mL×3). The organic layer was washed with water (1.0 L×2) and aqueous NaCl solution, dried over anhydrous Na2SO4 and evaporated in vacuo to give 5 (86.6 g, crude) as a light yellow oil. ESI-MS: m / z 278.1; 1 HNMR (400MHz, CDCl3): δ7.36(m,5H),5.73(d,J=3.6Hz,1H),4.76(m,1H),4.58(m,2H),4.02( m,1H),3.91(m,1H),3.81(m,1H),1.59(s,3H),1.36(m,3H),1.25(dd,J=6.6Hz,19.2Hz,3H).

[0172] A mixture of 5 (86.6 g, crude) and 2-iodobenzoic acid (165.0 g, 589.3 mmol) in ACN (1.0 L) was stirred at 80 ° C for 2 hours, and then TLC showed that 5 was completely consumed. It was then filtered and the organic layer was concentrated to give a crude product, which was purified by column chromatography (SiO2, PE / EA=10:1 to 8:1) to give 6 (48.6 g, 90% purity, 56.5% yield) as a yellow solid. ESI-MS: m / z 276.1; 1 HNMR (400MHz, CDCl3) δ7.36 (m, 5H), 5.82 (d, J = 3.56Hz, 1H), 4.77 (d, J = 11.96Hz, 1H), 4. 60(m,2H),4.52(d,J=9.16Hz,1H),3.77(m,1H),2.19(s,3H),1.60(s,3H),1.37(s,3H).

[0173] At -70 ° C, n-BuLi (1.6M, 86.0mL) was added dropwise to a stirred solution of diethyl ethylphosphonate (20.7g, 124.7mmol) in anhydrous THF (600.0mL), and it was stirred at -70 ° C for 0.5 hours. At -70 ° C, a THF (300.0mL) solution of 6 (40.0g, 137.0mmol) was added within 20 minutes, and it was stirred at -70 ° C for 2 hours. LCMS shows that 6 is completely consumed. It is quenched by NH4Cl aqueous solution and extracted by EA (600mL×3). The organic layer is washed with water (600.0mL×2) and NaCl aqueous solution, dried and filtered by anhydrous Na2SO4, and then the organic layer is concentrated. The crude product is purified by column chromatography (SiO2, PE / EA=1:1) to obtain 7 (33.6g, 73.3mmol, 93% purity) as a yellow solid. ESI-MS: m / z 459.2[M+H] + ; 1 HNMR(400MHz, CDCl3)δ7.34(m,5H),5.77(m,1H),4.83-4.71(m,1H),4.64-4.5 2(m,2H),4.23-3.86(m,7H),1.75(m,1H),1.58(t,J=7.6Hz,3H),1.30(m,15H). 31 P-NMR (162MHz, CDCl3) δ = 33.74, 32.93, 32.19, 31.53.

[0174] In an ice bath, SOCl2 (86.5 g, 73.3 mmol) was added to a solution of 7 (33.6 g, 73.4 mmol) in pyridine (200.0 mL) within 30 minutes and stirred at room temperature for 30 minutes. LCMS showed that 7 was completely consumed. The mixture was quenched by NaHCO3 (200.0 mL) aqueous solution, extracted by EA (400.0 mL×2) and washed by NaCl aqueous solution. After drying and filtering over anhydrous Na2SO4, the organic layer was concentrated to give a crude product, which was purified by column chromatography (SiO2, PE / EA=1:1) to give 8 (25.0 g, 56.8 mmol) as a light yellow solid. ESI-MS: m / z 441.2[M+H] + ; 1 HNMR (400MHz, CDCl3) δ7.34(m,5H),5.78(t,J=3.32Hz,1H),5.43(m,1H),5.31(m,J=5.36Hz1H) ,4.75(m,1H),4.56(m,3H),4.07(m,4H),3.69(m,1H),2.81(m,1H),1.60(s,3H),1.30(m,12H); 31 P-NMR (162MHz, CDCl3) δ = 30.33, 29.97.

[0175] To a solution of 8 (25.5 g, 56.0 mmol) in CH3OH (200.0 mL) was added Pd / C (3.8 g, 15%) at room temperature, stirred under N2 atmosphere at room temperature for 30 min, and then filtered. Another portion of Pd / C (3.8 g, 15%) was added and stirred at 50 °C for 3 hours. LCMS showed that 8 was completely consumed. After filtration, the organic layer was concentrated to give a crude product, which was purified by column chromatography to give 9 (15.0 g, 34.0 mmol, 58.6% yield) as a light yellow oil. ESI-MS: m / z 443.5 [M+H] + ; 1 HNMR(400MHz, CDCl3)δ7.35(m,5H),5.69(m,1H),4.81-4.73(m,1H),4.60-4.49(m,2H),4 .14-4.01(m,5H),3.56-3.48(m,1H),2.47-2.10(m,2H),1.58(m,3H),1.37-1.23(m,15H). 31 P-NMR (162MHz, CDCl3) δ = 34.76, 34.17, 33.95, 33.55.

[0176] A solution of 9 (15.0 g, 34.0 mmol) in 60% AcOH / H2O (75.0 mL) was stirred at 100 ° C for 3 hours. TLC showed that 9 was completely consumed, and then it was concentrated in vacuo to obtain a crude intermediate. The crude intermediate and DMAP (828.0 mg, 6.8 mmol) were dissolved in anhydrous pyridine (60.0 mL), and then Ac2O (20.8 g, 203.9 mmol) was added at 0 ° C in 10 minutes. It was stirred at room temperature for 2 hours, and then LCMS monitored its completion. The mixture was quenched by NaHCO3 (100.0 mL) aqueous solution and extracted by EA (100.0 mL × 2). The organic layer was washed with a NaCl aqueous solution and dried by anhydrous Na2SO4. After filtration, the organic layer was concentrated to obtain a crude product, which was purified by column chromatography to obtain 10 (13.8 g, 28.4 mmol, 83.7% yield) as a yellow oil. ESI-MS: m / z 509.2[M+Na] + ; 1 HNMR (400MHz, CDCl3) δ7.36-7.23(m,5H),6.36-6.07(m,1H),5.36-5.04(m,1H),4.65-4.52(m,1H),4.49-4. 42(m,1H),4.34-4.84(m,6H),2.45-2.18(m,1H),2.13-2.04(m,6H),1.95-1.69(m,1H),1.35-0.95(m,12H). 31 PNMR (162MHz, CDCl3) δ = 34.43, 34.17, 34.15, 33.56, 33.44, 33.39, 32.61.

[0177] A solution of uracil (4.1 g, 36.3 mmol) and BSA (15.9 g, 77.7 mmol) in anhydrous ACN (80.0 mL) was stirred at 50 ° C until the mixture was clear. Then a solution of 10 (11.8 g, 24.3 mmol) in anhydrous ACN (40.0 mL) was added, and TMSOTf (5.5 g, 24.8 mmol) was added dropwise at room temperature within 30 minutes. The mixture was stirred at room temperature for 5 hours. LMCS showed that 10 was completely consumed. The mixture was quenched by NaHCO (100.0 mL) aqueous solution and extracted by EA (200.0 mL × 2). The organic layer was washed with a NaCl aqueous solution and dried by anhydrous Na2SO4. After filtration, the organic layer was concentrated to obtain a crude product, which was purified by column chromatography (SiO2, PE: EA = 1: 2) to obtain 11 (10.3 g, 19.1 mmol, 78.8% yield) as a yellow solid. ESI-MS: m / z 539.1[M+H] + ; 1 HNMR(400MHz, CDCl3)δ9.94-9.86(m,1H),7.37-7.27(m,6H),5.77-5.71(m,2H),5.45-5.29(m,1H),4.60-4.46(m,2H) ,4.32-4.26(m,1H),4.14-4.03(m,5H),2.38-2.34(m,1H),2.22-2.16(m,1H),2.11-2.08(m,3H),1.38-1.02(m,12H). 31 PNMR (162MHz, CDCl3) δ = 34.16, 33.56, 33.29, 33.03, 32.44.

[0178] A solution of 11 (10.3 g, 19.1 mmol) in CH3NH2 (100.0 mL, 30% in MeOH) was stirred at room temperature for 1 hour. LCMS showed that 11 was completely consumed. The mixture was then concentrated in vacuo to give 12 (11.0 g, crude). ESI-MS: m / z 497.2 [M+H] + ; 31 PNMR (162MHz, CDCl3) δ = 34.18, 33.17, 33.02, 33.03, 32.40.

[0179] To a stirred solution of 12 (9.1 g, crude) and DBU (8.4 g, 55.2 mmol) in anhydrous DMF (50.0 mL) was added benzyl chloromethyl ether (5.7 g, 36.4 mmol) at 0°C over 10 minutes. The mixture was then stirred overnight at room temperature. LCMS showed that 12 was completely consumed. The mixture was then quenched by aqueous NaHCO3 (50.0 mL) solution and extracted by EA (100.0 mL×2). The organic layer was washed with aqueous NaCl solution and dried over anhydrous Na2SO4. After filtration, the organic layer was concentrated to give a crude product, which was purified by column chromatography (SiO2, PE:EA=1:1) to give 13 (7.0 g, 11.4 mmol, 59.4% yield). ESI-MS: m / z 617.4 [M+H] + ; 1 H NMR (400MHz, CDCl3) δ9.94-9.86(m,1H),7.38-7.17(m,11H),5.83-5.74(m,2H),5.66-5.43(m,3H),4.78-4.55(m,4H),4.37 -4.29(m,1H),4.22-3.93(m,6H),3.10-2.99(m,1H),2.45-2.11(m,1H),2.04-1.93(m,3H),1.65(m,1H),1.34-1.03(m,12H). 31 PNMR (162MHz, CDCl3) δ = 34.15, 33.17, 33.03, 33.46.

[0180] A solution of 13 (7.0 g, 10.2 mmol), NaI (767.0 mg, 5.1 mmol) and Ag2O (3.6 g, 15.3 mmol) in MeI (30.0 mL) was stirred at 40°C for 1 hour, and LCMS showed that 13 was completely consumed. After filtration, the organic layer was concentrated to give a crude product, which was purified by column chromatography (SiO2, PE:EA=1:1) to give 14-P1 (3.9 g, 6.2 mmol) and 14-P2 (3.1 g, 4.9 mmol) as yellow solids. ESI-MS: m / z 631.3 [M+H] + ;14-P1: 31 PNMR (162MHz, CDCl3) δ = 33.03, 32.85; 14-P2: 31 PNMR (162MHz, CDCl3) δ = 34.27, 33.55.

[0181] A solution of 14-P1 (3.9 g, 6.2 mmol) in CF3COOH (20.0 mL) was stirred at 80 ° C for 1 hour, and LCMS showed that 14 was completely consumed. After concentration, the crude product was purified by column chromatography (SiO2, PE: EA = 0: 1) to give a mixture product (2.5 g, 93% purity, 96.2% yield). The mixture was separated by SFC to give isomer 1 (500.0 mg, 1.2 mmol, 20.0% yield) and isomer 2 (760 mg, 1.8 mmol, 30.4% yield) of 15 as a white solid. A solution of 14-P2 (3.1 g, 4.9 mmol) in CF3COOH (20.0 mL) was stirred at 80 ° C for 1 hour, and LCMS showed that 14-P2 was completely consumed. After concentration, the crude product was purified by column chromatography (SiO2, PE:EA = 0: 1) to give a mixture of products (1.6 g, 91% purity, 77.4% yield). The mixture was separated by SFC to give isomer 3 of 15 (130.0 mg, 0.3 mmol, 8.1% yield) and isomer 4 of 15 (800 mg, 1.9 mmol, 50.0% yield) as white solids.

[0182] Isomer 1 of 15: 1 HNMR(400MHz, CDCl3)δ8.98(s,1H),7.33(d,J=8.12Hz,1H),5.76(m,2H),4.24(t,J=7.9Hz,1H),4.11( m,5H),3.89(d,J=5.6Hz,1H),3.59(s,3H),2.38-2.27(m,1H),2.15-2.04(m,1H),1.35-1.18(m,12H). 31 PNMR (162MHz, CDCl3)δ=33.29; ESI-MS: m / z 421.2[M+H] +

[0183] Isomer 2 of 15: 1 HNMR (400MHz, CDCl3) δ8.69 (s, 1H), 7.40 (d, J = 8.1Hz, 1H), 5.72-5.69 (m, 2H), 4.04-3.97 (m, 5H), 3.83 (q,J=6.2Hz,1H),3.72(m,1H),3.52(s,3H),2.27-2.16(m,1H),2.03-1.88(m,1H),1.26-1.17(m,12H). 31 PNMR (162MHz, CDCl3)δ=33.56; ESI-MS: m / z 421.2[M+H] +

[0184] Isomer 3 of 15: 1 HNMR (400MHz, CDCl3) δ8.69 (s, 1H), 7.40 (d, J = 8.1Hz, 1H), 5.83-5.76 (m, 2H), 4.20-4.05 (m, 4H), 3.91-3.88 (m,1H),3.74-3.71(m,2H),3.61(s,3H),2.46-2.31(m,2H),1.35-1.32(t,J=7.04Hz,6H),1.19-1.12(m,6H). 31 PNMR (162MHz, CDCl3)δ=33.51; ESI-MS: m / z 421.2[M+H] +

[0185] Isomer 4 of 15: 1 HNMR (400MHz, CDCl3) δ8.89 (s, 1H), 7.33 (d, J = 8.04Hz, 1H), 5.82-5.70 (m, 2H), 4.11-4.02 (m, 4H), 3.88 ( m,1H),3.72(s,1H),3.63(t,J=6.2Hz,9.28Hz,1H),3.52(s,3H),2.38-2.23(m,2H),1.27-1.01(m,12H). 31 PNMR (162MHz, CDCl3)δ=34.18; ESI-MS: m / z 421.1[M+H] +

[0186] Synthesis of A

[0187] To a stirred solution of isomer 1 of 15 (500 mg, 1.2 mmol) and DCI (126.4 mg, 1.1 mmol) in anhydrous DCM (5.0 mL) was added CEP[N(iPr)2]2 (430.0 mg, 1.4 mmol) under N2 atmosphere. The mixture was stirred at 30°C for 1 hour. LCMS showed that isomer 1 of 15 was completely consumed. The solution was then diluted with DCM (10 mL) and washed with H2O (10 mL×3). After drying over anhydrous Na2SO4, the organic layer was concentrated under reduced pressure and the residue was purified by rapid preparative HPLC using the following conditions (IntelFlash-1): column, C18 silica gel; mobile phase, CH3CN / H2O (0.5% NH4HCO3) = 1 / 1, increasing to CH3CN / H2O (0.5% NH4HCO3) = 1 / 0 within 20 minutes, collecting the eluted product at CH3CN / H2O (0.5% NH4HCO3) = 6 / 1; detector, UV 254nm. A (500 mg, 0.8 mmol, 95% purity, 65% yield) was obtained as a white solid. ESI-MS: m / z 619.2 [MH] - ; 1 HNMR (400MHz, CDCl3) δ9.10 (s, 1H), 7.30 (m, J = 8.4Hz, 1H), 5.82 (t, J = 4.4Hz, 1H), 5.76 (d, J = 8.4Hz, 1H), 4.34 (m, 1H), 4.1 0(m,5H),3.84(m,5H),3.48(m,5H),3.50(m,3H),2.72-2.66(m,2H),2.20(m,1H),2.04-1.97(m,1H),1.34-1.17(m,24H); 31 P NMR (162MHz, CDCl3) δ = 149.59, 33.43, 34.03.

[0188] Synthesis of B

[0189] To a stirred solution of isomer 2 of 15 (760 mg, 1.8 mmol) and DCI (192.2 mg, 1.6 mmol) in anhydrous DCM (5.0 mL) was added CEP[N(iPr)2]2 (653.6 mg, 2.1 mmol) under N2 atmosphere. The mixture was stirred at 30°C for 1 hour. LCMS showed that isomer 2 of 15 was completely consumed. The solution was then diluted with DCM (10 mL) and washed with H2O (10 mL×3). After drying over anhydrous Na2SO4, the organic layer was concentrated under reduced pressure and the residue was purified by rapid preparative HPLC using the following conditions (IntelFlash-1): column, C18 silica gel; mobile phase, CH3CN / H2O (0.5% NH4HCO3) = 1 / 1, increasing to CH3CN / H2O (0.5% NH4HCO3) = 1 / 0 within 20 minutes, collecting the eluted product at CH3CN / H2O (0.5% NH4HCO3) = 6 / 1; detector, UV 254nm. B (500mg, 0.8mmol, 95% purity, 45% yield) was obtained as a white solid. ESI-MS: m / z 619.2 [MH] - ; 1 HNMR(400MHz, CDCl3)δ9.40(s,1H),7.45(m,1H),5.77(m,2H),4.27-3.60(m,11H),3. 49(d,J=14Hz,3H),2.66(m,2H),2.38-2.30(m,1H),2.01(m,1H),1.33-1.15(m,24H). 31 PNMR (162MHz, CDCl3) δ = 149.35, 32.76, 32.73.

[0190] Synthesis of C

[0191] To a stirred solution of isomer 3 of 15 (130 mg, 0.3 mmol) and DCI (32.9 mg, 0.3 mmol) in anhydrous DCM (2.0 mL) was added CEP[N(iPr)2]2 (111.8 mg, 0.4 mmol) under N2 atmosphere. The mixture was stirred at 30°C for 1 hour. LCMS showed that isomer 3 of 15 was completely consumed. The solution was then diluted with DCM (10 mL) and washed with H2O (10 mL×3). After drying over anhydrous Na2SO4, the organic layer was concentrated under reduced pressure and the residue was purified by rapid preparative HPLC using the following conditions (IntelFlash-1): column, C18 silica gel; mobile phase, CH3CN / H2O (0.5% NH4HCO3) = 1 / 1, increased to CH3CN / H2O (0.5% NH4HCO3) = 1 / 0 within 20 minutes, and the eluted product was collected at CH3CN / H2O (0.5% NH4HCO3) = 6 / 1; detector, UV 254nm. C (110 mg, 0.2 mmol, 95% purity, 57% yield) was obtained as a white solid. ESI-MS: m / z 619.2 [MH] - ; 1 HNMR (400MHz, CDCl3) δ8.50 (s, 1H), 7.31 (m, 1H), 5.84 (m, 1H), 5.75 (d, J = 8.4Hz ,1H),4.16-3.59(m,11H),2.70(m,2H),2.40-2.18(m,2H),1.41-1.12(m,24H). 31 PNMR (162MHz, CDCl3) δ = 150.21, 149.66, 33.61, 33.01.

[0192] Synthesis of D

[0193] To a stirred solution of isomer 4 of 15 (800 mg, 1.9 mmol) and DCI (202.3 mg, 1.7 mmol) in anhydrous DCM (5.5 mL) was added CEP[N(iPr)2]2 (688.0 mg, 2.3 mmol) under N2 atmosphere. The mixture was stirred at 30°C for 1 hour. LCMS showed that isomer 4 of 15 was completely consumed. The solution was then diluted with DCM (10 mL) and washed with H2O (10 mL×3). After drying over anhydrous Na2SO4, the organic layer was concentrated under reduced pressure and the residue was purified by rapid preparative HPLC using the following conditions (IntelFlash-1): column, C18 silica gel; mobile phase, CH3CN / H2O (0.5% NH4HCO3) = 1 / 1, increasing to CH3CN / H2O (0.5% NH4HCO3) = 1 / 0 within 20 minutes, collecting the eluted product at CH3CN / H2O (0.5% NH4HCO3) = 6 / 1; detector, UV 254nm. D (760mg, 1.23mmol, 95% purity, 64% yield) was obtained as a white solid. ESI-MS: m / z 619.2 [MH] - ; 1 HNMR (400MHz, CDCl3) δ9.40 (s, 1H), 7.45 (m, 1H), 6.03 (t, J = 4.8Hz, 1H), 5.70 (m, 1H), 4.22-4.06 (m, 5H), 3.94-3.61 (m, 6H), 3.48 (d, J = 10.8Hz, 3H), 2.64 (m, 2H), 2.49-2.29 (m, 2H), 1.39-1.29 (m, 6H), 1.24-1.15 (m, 12H), 1.12-1.04 (m, 6H). 31 P NMR (162MHz, CDCl3) δ = 150.3, 149.90, 34.52, 34.36.

[0194] Example 3: Exemplary Synthesis of Oxetane VP

[0195]

[0196]

[0197] Under N2 atmosphere, 1 (prepared according to: T. Jonckers et al. J. Med. Chem., 2016, 59, 5790-5798) (10.0 g, 25.6 mmol), imidazole (34.9 g, 512 mmol) and DMAP (0.94 g, 7.7 mmol) were dissolved in DMF (100 mL). TBSCl (30.9 g, 205 mmol) was added in batches at 0 ° C. After the addition was complete, the mixture was stirred at 80 ° C for 16 hours. Saturated NH4Cl aqueous solution was added to the reactant, and the aqueous layer was extracted with EtOAc (3×). The combined organic layers were washed with 5% LiCl aqueous solution (2×), dried over Na2SO4 and concentrated in vacuo to give 2 (21.1 g) as a crude product, which was used in the next step without further purification.

[0198]

[0199] 2 (15.8 g, 25.6 mmol) was dissolved in THF (51.7 mL) and distilled water (13.7 mL). The mixture was cooled to 0 ° C, and TFA (13.7 mL) was added dropwise over 15 minutes while the internal reaction temperature was kept below 2 ° C. The mixture was stirred at 0 ° C for 1 hour, then at -18 ° C for 14 hours. The reactant was warmed to 0 ° C and continued to stir for 4 hours, then stirred at 5 ° C for 4 hours. The reactant was alkalized to pH ~ 8 with NH4OH solution, diluted with water, and the aqueous layer was extracted with EtOAc (3 ×). The combined organic layers were dried over Na2SO4 and concentrated in vacuo. The crude product was purified by flash silica gel chromatography (DCM / MeOH, gradient from 100: 0 to 98: 2) to obtain 3 (7.6 g, 59%).

[0200]

[0201] Under N2 atmosphere, 3 (3.0 g, 6.0 mmol) was dissolved in DCM (100 mL). DMP (3.6 g, 8.4 mmol) was added at 0 ° C, and the mixture was stirred at 0 ° C for 1 hour, and then stirred for another 3 hours at room temperature. A mixture of 50 mL 30% Na2S2O3 aqueous solution and saturated NaHCO3 aqueous solution (1: 1) was added to the mixture, and the aqueous layer was extracted with EtOAc (3 ×). The combined organic layers were dried over Na2SO4 and concentrated in vacuo to give 4 (4.4 g) as a crude product, which was used in the next step without further purification.

[0202] Under N2 atmosphere, NaH (60% dispersion in mineral oil, 720mg, 18mmol) is suspended in THF (10mL), and the mixture is cooled to -78 ℃.Add 5 (9.5g, 14.5mmol) in THF (11mL) solution, and the mixture is stirred at -78 ℃ for 20 minutes.At -78 ℃, in the period of 30 minutes, drip 4 (3.0g, 6mmol) in THF (21mL) solution in this mixture, and continue to stir for 100 minutes at the same temperature.The mixture is warmed to 0 ℃ and stirred for 1 hour, then stirred for another 90 minutes at room temperature.The crude reaction mixture is poured into 100mL saturated NH4Cl aqueous solution, and the product is extracted into 100mL ethyl acetate (2×).The organic layer is washed with brine, through anhydrous Na2SO4 dry. The solution was then concentrated under reduced pressure and the residue was purified by flash silica gel chromatography (heptane:EtOAc, gradient from 100:0 to 0:100) to afford 6 (2.7 g, 56%) as a 90:10 mixture of E:Z isomers.

[0203]

[0204] 6 (2.7 g, 2.7 mmol) was dissolved in MeCN (26 mL) and distilled water (8.1 mL). CAN (7.5 g, 13.7 mmol) was added and the mixture was stirred at room temperature for 24 hours. The reactant was alkalized to pH 6-7 with NH4OH solution, poured into a saline solution (50 mL) and extracted with EtOAc (3×). The combined organic layers were dried over Na2SO4 and concentrated in vacuo. The crude product was purified by flash silica gel chromatography (DCM / MeOH, gradient from 100:0 to 90:10) to give 7 (823 mg, 52%).

[0205]

[0206] Under N2 atmosphere, 7 (952mg, 1.7mmol) and 4,5-dicyanoimidazole (DCI, 166mg, 1.4mmol) are dissolved in anhydrous DCM (9.2mL).Add 2-cyanoethyl-N, N, N', N'-tetraisopropyl diaminophosphoric acid ester (0.684mL, 2.2mmol), the mixture is stirred at room temperature for 16 hours.Add another portion of 2-cyanoethyl-N, N, N', N'-tetraisopropyl diaminophosphoric acid ester (0.158mL, 0.5mmol), and the mixture is stirred at room temperature for 3 hours.The organic layer is poured into water and washed with water (2×) and salt solution (1×).The organic layer is through Na2SO4Dry and concentrate in a vacuum. The crude product was purified by flash silica gel chromatography (heptane with 0.15% TEA / EtOAc with 0.15% TEA, gradient from 100:0 to 0:100) to afford 8 (532 mg, 41%). 1 HNMR(400MHz,DMSO-d6)δppm 1.12-1.16(m,18H),1.17-1.24(m,12H),2.52-2.78(m,H),2.79-2.89(m,2H),3.60-3.79(m,3H),3.79-3.91(m,1H),4.16-4.29(m,1H), 4.32-4.56(m,3H),5.56-5.67(m,5H),5.88-5.94(m,1H),6.05-6.21(m,1H),6.67-6.97(m,1H),7.57-7.68(m,1H),11.43-11.50(m,1H). 31 P NMR (162 MHz, DMSO-d6) δ ppm 16.4, 17.2, 148.7, 150.7 LCMS: Method A, Rt: 1.24, 775.5 [M+H] +

[0207]

[0208] At room temperature, chlorodimethylphenylsilane (183.90 g, 1077.25 mmol) and imidazole (173.35 g, 2546.24 mmol) and DCM (3000 mL) were added to a 5L 3-neck round-bottom flask. At 0°C, 9 (200 g, 979.32 mmol) was added dropwise to the above mixture over 2 hours. The resulting mixture was stirred overnight at room temperature. The reactants were quenched with water at room temperature. The resulting mixture was extracted with CH2Cl2 (2×2 L). The combined organic layers were washed with brine (1×2 L) and dried over anhydrous Na2SO4. After filtration, the filtrate was concentrated under reduced pressure. The residue was purified by flash silica gel chromatography eluting with PE / EA=10:1 to give 10 (260 g, 78%).

[0209]

[0210] At room temperature, LiBH4 (50.19 g, 2304.45 mmol) and THF (6500 mL) were added to a 10L 4-neck round-bottom flask. 10 (260 g, 768.15 mmol) was added dropwise to the above mixture over 1.5 hours at 0°C. The resulting mixture was stirred overnight at room temperature. The reactants were quenched at 0°C by adding saturated NH4Cl (aqueous solution) (3 L). The resulting mixture was extracted with CH2Cl2 (2×1 L). The combined organic layers were washed with brine (1×2 L) and dried over anhydrous Na2SO4. After filtration, the filtrate was concentrated under reduced pressure. The residue was purified by flash silica gel chromatography eluting with PE / EA (1:1) to give 11 (120 g, 61%).

[0211]

[0212] At room temperature, 11 (120 g, 471.69 mmol), DCM (6000 mL), pyridine (37.68 g, 476.41 mmol) and DMAP (5.76 g, 47.17 mmol) were added to a 10 L 4-neck round-bottom flask. Ac2O (48.64 g, 476.41 mmol) was added dropwise to the above mixture over 1 hour at 0°C. The resulting mixture was stirred at room temperature overnight. The reactants were quenched with NaHCO3 (aqueous solution) at room temperature. The resulting mixture was extracted with CH2Cl2 (2×2 L). The combined organic layers were washed with brine (1×2 L) and dried over anhydrous Na2SO4. After filtration, the filtrate was concentrated under reduced pressure. The residue was purified by flash silica gel chromatography eluting with PE / EA (1:1) to give 12 (64 g, 46%).

[0213]

[0214] At room temperature, uracil (70 g, 624.50 mmol), acetonitrile (210 mL) and pyridine (123.50 g, 1.56 mol) were added to a 1L 3-necked round-bottom flask. Benzyl chloroformate (234.38 g, 1.37 mol) was added dropwise to the above mixture at 0°C over 1.5 hours. The resulting mixture was stirred at room temperature overnight. The resulting mixture was extracted with EtOAc (2×500 mL). The combined organic layers were washed with brine (1×500 mL) and dried over anhydrous Na2SO4. After filtration, the filtrate was concentrated under reduced pressure.

[0215] The crude product was recrystallized from MeOH (200 mL) to afford 13 (80 g, 59%).

[0216]

[0217] At room temperature, 12 (64 g, 215.89 mmol), PPh3 (124.58 g, 474.97 mmol) and THF (1600 mL) were added to a 3L 3-neck round-bottom flask. 13 (93.35 g, 431.79 mmol) and DIAD (96.04 g, 474.97 mmol) were added dropwise to the above mixture over 2 hours at -10°C. The resulting mixture was stirred for another 3 hours at room temperature. The reactants were quenched by adding water / ice (1 L) at 0°C. The resulting mixture was extracted with EtOAc (2×1 L). The combined organic layers were washed with brine (1×1 L) and dried over anhydrous Na2SO4. After filtration, the filtrate was concentrated under reduced pressure. The residue was purified by flash silica gel chromatography eluting with PE / EA (1:1) to give 14 (80 g, 75%).

[0218]

[0219] To a 2L 3-necked round-bottom flask was added 14 (80 g, 161.74 mmol) and THF (800 mL) at room temperature. To the above mixture was added dropwise hydrazine hydrate (12.15 g, 242.61 mmol) at 0°C over 30 minutes. The resulting mixture was stirred at room temperature for another 3 hours. The resulting mixture was concentrated under reduced pressure. The residue was purified by flash silica gel chromatography eluting with PE / EA (1:1) to give 15 (55 g, 87%).

[0220]

[0221] At room temperature, 15 (55 g, 140.84 mmol) and THF (550 mL) were added to a 500 mL 3-neck round bottom flask. TBAF (36.82 g, 140.84 mmol) was added dropwise to the above mixture at 0°C over 0.5 h. The resulting mixture was stirred for another 3 h at room temperature. The resulting mixture was extracted with CH2Cl2 (2×1 L). The combined organic layers were washed with brine (1×1 L) and dried over anhydrous Na2SO4. After filtration, the filtrate was concentrated under reduced pressure. 16 (40 g, crude product) was obtained, which was used in the next step without further purification.

[0222] At room temperature, 16 (40 g, 156.09 mmol), imidazole (31.42 g, 468.27 mmol) and THF (400 mL) were added to a 1L 3-necked round-bottom flask. Tert-butyl (chloro) diphenylsilane (85.81 g, 312.18 mmol) was added dropwise to the above mixture at 0°C for 1 hour. The resulting mixture was stirred for another 3 hours at room temperature. The resulting mixture was extracted with EtOAc (2×400 mL). The combined organic layers were washed with brine (1×400 mL) and dried over anhydrous Na2SO4. After filtration, the filtrate was concentrated under reduced pressure. The residue was purified by flash silica gel chromatography eluting with PE / EA (1:1) to give 17 (52 g, 67%).

[0223]

[0224] At room temperature, 17 (52 g, 105.12 mmol) and methanol (500 mL) were added to a 1L 3-neck round-bottom flask. NaOH (2 M, 100 mL) was added dropwise to the above mixture over 1 hour at 0°C. The resulting mixture was stirred for another 3 hours at room temperature. The resulting mixture was extracted with EtOAc (2×200 mL). The combined organic layers were washed with brine (1×200 mL) and dried over anhydrous Na2SO4. After filtration, the filtrate was concentrated under reduced pressure. The residue was purified by grinding with MeOH (100 mL), and the solid was collected by filtration to give 18 (40.2 g, 84%).

[0225]

[0226] Under N2 atmosphere, 18 (2.4 g, 5 mmol) was dissolved in DCM (75 mL). DMP (3.0 g, 7 mmol) was added at 0 ° C, and the mixture was stirred for another 2 hours at room temperature. A mixture of 50 mL 30% Na2S2O3 aqueous solution and saturated NaHCO3 aqueous solution (1: 1) was added to the mixture, and the aqueous layer was extracted with EtOAc (3×). The combined organic layers were dried over Na2SO4 and concentrated in vacuo to give 19 as a crude product, which was used in the next step without further purification.

[0227] Under N2 atmosphere, NaH (60% dispersion in mineral oil, 440mg, 18mmol) is suspended in THF (10mL), and the mixture is cooled to -40 ℃. Tetraethyl methylene diphosphonate (3.0mL, 12mmol) is added, and the mixture is stirred for 30 minutes at -40 ℃. At -40 ℃, a solution of 19 (2.3g, 5mmol) in THF (25mL) is added dropwise to the mixture over a period of 10 minutes, and the mixture is stirred for 100 minutes while the mixture is warmed to -10 ℃. The crude reaction mixture is poured into water, and the product is extracted into ethyl acetate (3×). The organic layer is dried over anhydrous Na2SO4 and concentrated in vacuo. The crude product was purified by flash silica gel chromatography (heptane / EtOAc:EtOH (3:1), gradient from 100:0 to 0:100) to afford 20 as an inseparable mixture with tetraethyl methylenediphosphonate (2.1 g, 35%).

[0228]

[0229] Under N2 atmosphere, 20 (2.1 g, 1.8 mmol) was dissolved in THF (31 mL), TBAF (1 M THF solution, 2.3 mL, 2.3 mmol) was added at room temperature, and the mixture was stirred for 16 hours. The solvent was removed in vacuo, and the crude product was purified by flash silica gel chromatography (DCM / MeOH, gradient from 100:0 to 94:6) to obtain 21a and 21b (562 mg, 91%) as racemates. Purification was performed via preparative SFC (stationary phase: Chiralpak Diacel AD 20×250 mm, mobile phase: CO2, EtOH+0.4iPrNH2). 21a and 21b were each purified again via preparative SFC (stationary phase: Torus Diol 30×150 mm, mobile phase: CO2, MeOH+0.4iPrNH2) to obtain 21a (192 mg, 31%). 21b was further purified via preparative SFC (stationary phase: Chiralpak Diacel AD 20×250 mm, mobile phase: CO 2 , EtOH+0.4iPrNH 2 ) to give 21b (163 mg, 27%).

[0230]

[0231] Under N2 atmosphere, 7 (192mg, 0.55mmol) and 4,5-dicyanoimidazole (DCI, 56mg, 0.47mmol) are dissolved in anhydrous DCM (3mL).2-cyanoethyl-N, N, N', N'-tetraisopropyl diaminophosphoric acid ester (0.23mL, 0.72mmol) is added, and mixture is stirred at room temperature for 16 hours.Mixture is concentrated in a vacuum, and crude product is passed through flash silica gel chromatography (heptane and 0.15%TEA / EtOAc and 0.15%TEA, gradient from 100:0 to 0:100, then DCM / MeOH, gradient from 100:0 to 95:5) purifying, obtain 22a (205mg, 68%). 1 H NMR (400MHz, acetonitrile-d3) δppm 1.14–1.22(m,12H),1.23-1.30(m,6H),1.78–1.90(m,2H),2.50–2.65(m,2H),2.65–2.72(m,2H),3.56–3.78(m,4H),3.78–3. 94(m,2H),3.94–4.10(m,5H),5.49–5.57(m,1H),5.72–5.87(m,1H),6.59–6.80(m,1H),7.27–7.37(m,1H),8.79–9.00(m,1H). 31P NMR (162 MHz, acetonitrile-d3) δ ppm 16.9, 17.2, 146.8, 147.8. LCMS: Method A, Rt: 1.00, 545.2 [MH] - ,1.02,545.2[MH] -

[0232]

[0233] Under N2 atmosphere, 21b (163mg, 0.47mmol) and 4,5-dicyanoimidazole (DCI, 47mg, 0.40mmol) are dissolved in anhydrous DCM (2.6mL).2-cyanoethyl-N, N, N', N'-tetraisopropyl diaminophosphoric acid ester (0.19mL, 0.61mmol) is added, and the mixture is stirred at room temperature for 16 hours.Mixture is concentrated in a vacuum, and crude product is passed through flash silica gel chromatography (heptane and 0.15%TEA / EtOAc and 0.15%TEA, gradient from 100:0 to 0:100, then DCM / MeOH, gradient from 100:0 to 95:5) purifying, obtain 22a (196mg, 76%). 1 H NMR (400MHz, acetonitrile-d3) δppm 1.16-1.22(m,12H),1.23-1.30(m,6H),1.79-1.91(m,2H),2.48-2.62(m,2H),2.64-2.71(m,2H),3.56-3.76(m,4H),3.79-3. 94(m,2H),3.94-4.09(m,5H),5.50-5.58(m,1H),5.71-5.85(m,1H),6.58-6.79(m,1H),7.28-7.39(m,1H),8.81-9.03(m,1H). 31 P NMR (162 MHz, acetonitrile-d3) δ ppm 16.9, 17.2, 146.8, 147.8. LCMS: Method A, Rt: 0.96, 545.3 [MH] - ,0.99,545.3[MH] -

[0234]

[0235] At 10 ℃-25 ℃, Ac2O (3.1 mL, 32.8 mmol) was added to a solution of compound 23 (9.00 g, 32.8 mmol) in pyridine (90.0 mL). The mixture was stirred at 0 ℃ for 2 hours. The reaction mixture was concentrated under reduced pressure to remove the solvent. The residue was purified by flash silica gel chromatography (PE / EtOAc, gradient from 100:0 to 0:100) to give compound 13 (18.0 g, 86% yield).

[0236]

[0237] Imidazole (11.6 g, 171 mmol) was added to a solution of compound 24 (18.0 g, 56.9 mmol) in DCM (100 mL). Then 25.0 mL of a dichloromethane solution of TBSCl (20.9 mL, 170 mmol) was slowly added at 25 ° C. The mixture was stirred at 25 ° C for 12 hours. The reaction mixture was quenched by adding H2O 400 mL at 0 ° C, and then extracted with ethyl acetate 600 mL (200 mL × 3). The combined organic layer was washed with brine 200 mL, dried over Na2SO4, filtered and concentrated under reduced pressure. The crude product was purified by flash silica gel chromatography (PE / EtOAc, gradient from 100: 0 to 80: 20) to obtain compound 25 (21.0 g, 85% yield).

[0238]

[0239] K2CO3 (13.5 g, 97.5 mmol) was added to a solution of compound 25 (21.0 g, 48.7 mmol) in MeOH (150 mL). The mixture was stirred at 25 ° C for 0.5 hours. The reaction mixture was quenched by adding H2O 350 mL at 0 ° C, and then extracted with ethyl acetate 600 mL (200 mL×3). The combined organic layer was washed with brine 200 mL, dried over Na2SO4, filtered and concentrated under reduced pressure. The crude product was purified by flash silica gel chromatography (PE / EtOAc, gradient from 100:0 to 0:100) to give compound 25 (21.0 g, 85% yield). The crude product was further purified by reverse phase SFC (column: DAICEL CHIRALPAK AS (250 mm×50 mm, 10 μm); mobile phase: [Neu-ETOH]; B%: 20%-20%, min) to give 26 (14.5 g, 84%).

[0240]

[0241] Under N2 atmosphere, 26 (1.9 g, 5.0 mmol) was dissolved in DCM (50 mL). DMP (3.3 g, 7.8 mmol) was added, and the mixture was stirred at room temperature for 3 hours. A mixture of 50 mL 30% Na2S2O3 aqueous solution and saturated NaHCO3 aqueous solution (1: 1) was added to the mixture, and the aqueous layer was extracted with EtOAc (3×). The combined organic layers were dried over Na2SO4 and concentrated in vacuo to give 27 (2.5 g) as a crude product, which was used in the next step without further purification.

[0242] Under N2 atmosphere, NaH (60% dispersion in mineral oil, 588mg, 15mmol) is suspended in THF (17mL), and the mixture is cooled to -78 ℃.Add a solution of 5 (7.7g, 14.5mmol) in THF (9mL), and the mixture is stirred at -78 ℃ for 60 minutes.At -78 ℃, in the mixture, a solution of 27 (3.0g, 6mmol) in THF (17mL) is added dropwise over a period of 15 minutes, and stirring is continued at the same temperature for 1 hour.The mixture is warmed to 0 ℃ and stirred for 1 hour, and then stirred at room temperature for another 1 hour.The crude reaction mixture is poured into 50mL saturated NH4Cl aqueous solution, and the product is extracted into 50mL ethyl acetate (3×).The organic layer is washed with brine, over anhydrous Na2SO4 dry. The solution was then concentrated under reduced pressure and the residue was purified by flash silica gel chromatography (heptane:EtOAc, gradient from 100:0 to 0:100) to afford 28 (1.4 g, 40%).

[0243]

[0244] To a mixture of 28 (608 mg, 0.9 mmol) and distilled water (15 mL) was added formic acid (15 mL). The reactant was stirred at room temperature for 20 hours. The solvent was removed in vacuo, and the crude product was purified by flash silica gel chromatography (DCM / MeOH, gradient from 100:0 to 95:5) to give 29 (402 mg, 79%).

[0245]

[0246] Under N2 atmosphere, 7 (402mg, 0.7mmol) and 4,5-dicyanoimidazole (DCI, 70mg, 0.6mmol) are dissolved in anhydrous DCM (3.9mL).2-cyanoethyl-N, N, N', N'-tetraisopropyl diaminophosphoric acid ester (0.29mL, 0.9mmol) is added, and the mixture is stirred at room temperature for 16 hours.The organic layer is poured into water and washed with water (2×) and salt solution (1×).The organic layer is dried over Na2SO4 and concentrated in a vacuum.The crude product is purified by flash silica gel chromatography (heptane with 0.15%TEA / EtOAc and 0.15%TEA, gradient from 100:0 to 0:100), and another fraction (279mg, 36%, 70% purity) of 30 (163mg, 30%) and 30 is obtained. 1 H NMR (400 MHz, acetonitrile-d3) δ ppm 1.15-1.22 (m, 30H), 2.63-2.71 (m, 2H), 3.43-3.49 (m, 3H), 3.56-3.68 (m, 2H), 3.68-3.79 (m, 1H), 3.79-3.88 (m, 1H), 3.92-4.00 (m, 1H), 4.09-4.22 (m, 1H), 4.29-4.39 (m, 1H), 5.57-5.71 (m, 5H), 5.99-6.15 (m, 2H), 6.79-7.00 (m, 1H), 7.79-7.85 (m, 1H), 8.82-9.07 (m, 1H). 31 P NMR (162 MHz, acetonitrile-d3) δ ppm 15.7, 15.9, 149.9, 150.3. LCMS: Method A, Rt: 1.23, 779.4 [M+H] + ,1.24,779.4[M+H] + .

[0247] Example 4 - Example of Unlocking Vinyl Phosphonate Synthesis

[0248]

[0249] LCMS method :

[0250]

[0251] CPG (25 μmol, 30 mg) with the synthesized n-1 oligonucleotide with 5'-DMTr was placed in a 1 mL syringe equipped with a filter and a cap.

[0252] Step 1. Fill the syringe with 3% dichloroacetic acid in DCM and keep at ambient temperature for 20 minutes. Remove the solvent and wash the CPG 6 times with 0.6 mL acetonitrile, 3 times with anhydrous DCM, and then dry in vacuo for 30 minutes.

[0253] Step 2. Fill the syringe with 200 μL of 0.1M phosphonate amide solution in anhydrous acetonitrile and 200 μL of 0.25M 5-(ethylthio)-1H-tetrazole acetonitrile solution. Close the syringe and shake gently for 40 minutes at ambient temperature. Remove the solvent and repeat the second operation. Remove the solvent and wash CPG 6 times with 0.6mL acetonitrile. Add 0.05M iodine solution in pyridine / water 9:1 (v:v) and keep for 15 minutes, repeat twice. Wash CPG 6 times with 0.6mL acetonitrile, wash 3 times with anhydrous DCM, and then dry in vacuum for 30 minutes. Take an aliquot (2mg) of CPG, treat with AMA at 60°C for 10 minutes, and check the MW of the oligonucleotide obtained by LCMS. If the coupling is not completed, repeat step 2.

[0254] Step 3. At ambient temperature, the anhydrous CPG in the syringe was treated with 250 μL of 3.5% trimethylsilyl iodide in acetonitrile / pyridine 50:1 (v / v) for 20 minutes. This treatment was repeated 3 times. The solvent was removed, and the CPG was washed 6 times with 0.6 mL of acetonitrile, then treated 20 minutes with a solution of 4% mercaptoethanol in pyridine / triethylamine 1:1 (v:v) at ambient temperature. The solvent was removed, and the CPG was washed 6 times with 0.6 mL of acetonitrile, washed 3 times with anhydrous DCM, and then dried in a vacuum for 30 minutes. An aliquot (2 mg) of CPG was taken, treated with AMA at 60°C for 10 minutes, and the MW of the oligonucleotide obtained was checked by LCMS. If deprotection is not carried out to completion, step 3 is repeated.

[0255] Step 4. Treat the CPG in the syringe with 200 μL AMA for 3 hours and filter. Wash the solid support 5 times with 250 μL water. Combine the filtrate and washings, dilute to 10 mL, and separate the target compound by ion exchange chromatography. Column GESource 15Q 250×10 mm. Buffer A is 20 mM sodium phosphate in acetonitrile / water 9:1 (v:v), and buffer B is 1.8 M sodium borate in buffer A (pH 7.5). Use a gradient of 5% to 60% over 10 column volumes to elute the target compound at 25%-35% B. The obtained oligonucleotides are desalted and used for annealing.

[0256] Step 5. Annealing was achieved by mixing equimolar amounts of sense and antisense strands. The obtained siRNA was analyzed by LCMS under denaturing conditions at 80°C.

[0257] Examples of synthetic AS :

[0258]

[0259] Examples of Synthetic siRNA

[0260]

[0261] HTRAKD via stable phospho-reactivity in ARPE-19

[0262]

[0263]

[0264] The table above shows the percentage of HTRA remaining at 3 days in vitro for different concentrations of oligonucleotides with a recognized phosphate attached (n=3 samples), the oligonucleotides in 1 and 2 are different, and the oligonucleotides in the remaining samples are identical except for the recognized phosphate.

[0265] Novel stable phosphodiesterase KD at 4 weeks after intravitreal injection

[0266]

[0267] The table above shows the novel stable phospho-results KD at weeks 2 and 4 after intravitreal injection. The oligonucleotides in the samples were identical except for the recognized phospho-results.

[0268] Additional implementations include:

[0269] 1. A compound represented by formula (Ia) or (Ib):

[0270]

[0271] in:

[0272] X is selected from 3- to 5-membered cycloalkyl, -CHCH-, 3- to 5-membered heterocyclic ring and -CHR 3 CHR 3 -;

[0273] Y is selected from O or NR';

[0274] R' is a counter ion, H or a protecting group

[0275] Z is selected from H, a counterion, an activating group, and an oligonucleotide;

[0276] A is selected from O, S and CR 4 R 4 ;

[0277] B is a nucleobase;

[0278] Each R is independently selected from an oligonucleotide, a counterion, H, and a protecting group such as a C1-C5 alkyl group and POM or a C1-C5 alkyl group;

[0279] Each R 1 and R 2 are independently selected from H, F, OH and optionally substituted O-alkyl, provided that R 1 and R 2 Not each is H;

[0280] Each R 3 Independently selected from C1-C3 alkyl;

[0281] Each R 4 Independently selected from H, F and C1-C5 alkyl;

[0282] with the proviso that when A is O, then X is not -CHCH-, and when X is cyclopropyl, then at least one of A or Y is not O.

[0283] 2. The compound according to embodiment 1, wherein the compound of formula (Ia) is represented by:

[0284]

[0285] 3. The compound according to embodiment 1 or 2, wherein X is cyclobutyl.

[0286] 4. A compound according to embodiment 3, wherein the cyclobutyl group is represented by:

[0287] The dashed lines indicate the connection points with neighboring atoms.

[0288] 5. The compound according to embodiment 1 or 2, wherein X is -CHR 3 CHR 3 -, and R 3 For methylene.

[0289] 6. The compound according to embodiment 5, wherein X is selected from the following:

[0290] The dashed lines indicate the connection points with neighboring atoms.

[0291] 7. The compound according to embodiment 1 or 2, wherein X is a 3- to 5-membered heterocyclic ring selected from:

[0292] The dashed lines indicate the connection points with neighboring atoms.

[0293] 8. A compound according to any one of embodiments 1 to 7, wherein Y is O.

[0294] 9. A compound according to any one of embodiments 1 to 8, wherein Z is an activating group.

[0295] 10. The compound according to embodiment 9, wherein the activating group is represented by:

[0296] The dashed lines indicate the connection points with neighboring atoms.

[0297] 11. The compound according to any one of embodiments 1 to 8, wherein Z is an oligonucleotide.

[0298] 12. The compound according to embodiment 11, wherein the oligonucleotide is the antisense strand of RNA, preferably the antisense strand of siRNA.

[0299] 13. The compound according to embodiment 11 or 12, wherein the 5' end of the oligonucleotide is linked to Y.

[0300] 14. A compound according to any one of embodiments 1 to 13, wherein A is O.

[0301] 15. A compound according to any one of embodiments 1 to 13, wherein A is CH2 or CHF.

[0302] 16. The compound according to embodiment 15, wherein X is -CHCH-.

[0303] 17. The compound according to any one of embodiments 1 to 16, wherein B is uracil.

[0304] 18. A compound according to any one of embodiments 1 to 17, wherein R 1 H and R 2 is OMe, OEt, MOE or F.

[0305] 19. A compound according to any one of embodiments 1 to 18, wherein R is a protecting group, such as POM, Et, and Z is an activating group.

[0306] 20. The compound of any one of embodiments 1 to 18, wherein R is H and Z is an oligonucleotide.

[0307] As will be appreciated by those skilled in the art, for any and all purposes, specifically with respect to providing a written description, all ranges disclosed herein also encompass any and all possible subranges and combinations of their subranges. Any listed range can be easily understood to fully describe and allow the same range to be decomposed into at least equal halves, thirds, quarters, fifths, tenths, etc. As a non-limiting example, each range discussed herein can be easily decomposed into a lower third, a middle third, and an upper third, etc. It will also be appreciated by those skilled in the art that all language, such as "at most," "at least," "greater than," "less than," etc., includes the listed numbers and refers to a range that can be subsequently decomposed into subranges as described above. Finally, as will be appreciated by those skilled in the art, a range includes each individual member.

[0308] All publications, patent applications, issued patents, and other documents mentioned in this specification are incorporated herein by reference, as if each individual publication, patent application, issued patent, or other document was specifically and individually indicated to be incorporated by reference in its entirety. Definitions contained in text incorporated by reference are excluded to the extent that they conflict with definitions in this disclosure.

[0309] Other embodiments are set forth in the following claims.

Claims

1. A compound represented by formula (Ia) or (Ib): in: X is selected from 3- to 5-membered cycloalkyl, -CHCH-, 3- to 5-membered heterocyclic ring and -CHR 3 CHR 3 -; Y is selected from O or NR'; R' is a counterion, H or a protecting group; Z is selected from H, a counterion, an activating group, and an oligonucleotide; A and the connecting dashed line are optional, and when present, A is selected from O, S and CR 4 R 4 ; B is a nucleobase; Each R is independently selected from an oligonucleotide, a counterion, H, and a protecting group such as a C1-C5 alkyl group and POM or a C1-C5 alkyl group; Each R 1 and R 2 independently selected from H, F, OH and optionally substituted O-alkyl, provided that when X is selected from 3- to 5-membered cycloalkyl, -CHCH-, 3- to 5-membered heterocyclic ring and -CHR 3 CHR 3 -, then R 1 and R 2 Not H or R 1 and R 2 forming an optionally substituted oxycyclobutene; Each R 3 Independently selected from C1-C3 alkyl; Each R 4 Independently selected from H, F and C1-C5 alkyl; R 5 and R 6 One of them is XP(O)(OR)2 and the other is H; with the proviso that when A is O, then X is not -CHCH-, and when X is cyclopropyl, then at least one of A or Y is not O.

2. The compound according to claim 1, wherein the compound has formula (Ia) and is represented by:

3. The compound according to claim 1 or 2, wherein X is cyclobutyl.

4. The compound according to claim 3, wherein the cyclobutyl group is represented by: The dashed lines indicate the connection points with neighboring atoms.

5. The compound according to claim 1 or 2, wherein X is -CHR 3 CHR 3 -, and R 3 For methylene.

6. The compound according to claim 5, wherein X is selected from the following: The dashed lines indicate the connection points with neighboring atoms.

7. The compound according to claim 1 or 2, wherein X is a 3- to 5-membered heterocycle selected from the following: The dashed lines indicate the connection points with neighboring atoms.

8. The compound according to claim 1, wherein the compounds of formula (Ia) and (Ib) are represented by:

9. The compound according to claim 1 or 2, wherein R 2 is an optionally substituted oxycyclobutene.

10. The compound according to claim 9, wherein X is -CHCH-.

11. The compound according to any one of claims 1 to 10, wherein Y is O.

12. The compound according to any one of claims 1 to 11, wherein Z is an activating group.

13. The compound of claim 12, wherein the activating group is represented by: The dashed lines indicate the connection points with neighboring atoms.

14. The compound according to any one of claims 1 to 11, wherein Z is an oligonucleotide.

15. The compound according to claim 14, wherein the oligonucleotide is the antisense strand of RNA, preferably the antisense strand of siRNA.

16. The compound according to claim 14 or 15, wherein the 5' end of the oligonucleotide is linked to Y.

17. The compound according to any one of claims 1 to 16, wherein A is O.

18. A compound according to any one of claims 1 to 16, wherein A is CH2 or CHF.

19. The compound according to claim 18, wherein X is -CHCH-.

20. The compound according to any one of claims 1 to 19, wherein B is uracil or thymine.

21. A compound according to any one of claims 1 to 17, wherein R 1 H and R 2 is OMe, OEt, MOE or F.

22. A compound according to any one of claims 1 to 21, wherein R is a protecting group, such as POM, Et, and Z is an activating group.

23. A compound according to any one of claims 1 to 21, wherein R is H and Z is an oligonucleotide.

24. The compound according to claim 1, wherein the compound is selected from: Or a compound in the table, wherein the nucleobase is thymine or cytosine.

25. The compound according to claim 24, wherein R 2 It is -OMe.

26. The compound of claim 24 or 25, wherein R is Et and Z is an activating group.

27. The compound of claim 24 or 25, wherein R is H or a counterion and Z is an oligonucleotide.

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