Ribose-modified capped analogs and uses thereof

Abstract

To provide a ribose-modified cap analog and a use thereof.SOLUTION: The present disclosure belongs to technical fields of chemical and biological engineering, the ribose-modified cap analog described herein has a structure of formula (I), and can improve stability of mRNA and / or translation efficiency of mRNA.SELECTED DRAWING: Figure 1

Description

[Technical Field]

[0001] The present invention is in the field of chemical and biotechnology and relates to ribose-modified capped analogs and their uses. [Background technology]

[0002] The chemical essence of the cap structure is a special structure located at the 5' end of mRNA, formed during the mRNA transcription process by modification: the m7GPPPN structure, also known as the methylguanosine cap. It is formed under the cooperative catalytic action of RNA triphosphatase, guanosyltransferase, mRNA (guanine-N7) methyltransferase, and mRNA (nucleoside-2') methyltransferase. Depending on the degree of methylation, three types of caps are formed: CAP0, CAP1, and CAP2, which are m7G5'ppp5'Np, m7G5'ppp5'NmpNp, and m7G5'ppp5'NmpNmpNp, respectively.

[0003] The cap structure is essential for mRNA translation initiation, providing a signal for ribosomes to recognize the mRNA and helping ribosomes bind to the mRNA so that translation starts at the AUG. At the same time, the cap structure can increase mRNA stability and protect it from 5'→3' exonuclease attack.

[0004] Simply put, the cap structure is like a steel helmet for mRNA, not only protecting it from toxins but also allowing chemical modifications to imprint the steel helmet to make it easier for other members to recognize. In addition to the native cap structure, cap analogs are often used to improve the stability of mRNA structures during in vitro transcription, and common ones include ARCA and Cap1 analogs.

[0005] Research has shown that the mRNA cap structure plays an important role in mRNA quality control and the body's innate immunity, so the development of novel capped analogs is of great importance for enhancing mRNA stability and improving mRNA translation efficiency. Summary of the Invention

[0006] In response to the shortcomings of existing technologies, the present invention provides ribose-modified capped analogs and uses thereof. The ribose-modified capped analogs described in the present application can enhance mRNA stability and / or mRNA translation efficiency.

[0007] To achieve this goal, the present invention adopts the following technical solutions.

[0008] In a first aspect, the present invention provides a ribose-modified capped analog having the structure of formula (I) or a stereoisomer, pharmaceutically acceptable salt, or solvate thereof.

[0009] [ka]

[0010] wherein n is selected from 1 or 0, and m is selected from 1 or 0; X1 is selected from -C(OH)H- or a single bond; X2 is -CH-, -O-, -CN, -C(O)-,

[0011] [ka] or a single bond; X3 is selected from -CH2, -CFH- or -O-; R1 is selected from -OH or -NHAc; R2 is selected from -CH3, -F, -H, or is absent; R3 is selected from -OR6, -NHC(O)R7, -N(R8), -CN, -F, -C(O)N(R9), substituted or unsubstituted C1-C3 alkyl, -H, or is absent; R4 is selected from -CH3, -F or -H; R5 is -OR 10 , -NHC(O)R 11 , -N(R 12 )2, -CF2H, -F, -C(O)N(R 13 ) 2, substituted or unsubstituted C1-C3 alkyl, or absent; R6, R8, R 10 , R 11 , R 12 and R 13 are each independently selected from unsubstituted C1-C3 alkyl or -H; R7 is -OR 14 , -NHC(O)R 15 , -F, or substituted or unsubstituted C1-C3 alkyl; R9 is selected from unsubstituted C2-C3 alkyl; R 14 and R 15 are each independently selected from unsubstituted C1-C3 alkyl.

[0012] Compared with the ribose-modified capped analogs of the prior art, the ribose-modified capped analogs of the present invention can significantly improve the in vitro transcription yield of mRNA, capping rate, mRNA translation efficiency, and mRNA protein expression level and duration in mice, while significantly reducing the decapping rate.

[0013] In a second aspect, the present invention provides the use of a ribose-modified capped analogue or a stereoisomer, pharmaceutically acceptable salt or solvate thereof described in the first aspect of the present invention in the preparation of an in vitro co-transcriptional capping reagent for mRNA.

[0014] In a third aspect, the present invention provides an RNA molecule comprising a ribose-modified capped analog described in one aspect of the present invention, or a stereoisomer, pharmaceutically acceptable salt, or solvate thereof, as a cap structure or cap structure fragment.

[0015] In a fourth aspect, the present invention provides a pharmaceutical composition comprising an RNA molecule according to the third aspect of the invention.

[0016] In a fifth aspect, the present invention provides a method for synthesizing an mRNA molecule for purposes other than disease diagnosis and treatment, comprising the step of co-incubating a ribose-modified capped analog described in the first aspect of the present invention, or a stereoisomer, pharmaceutically acceptable salt, or solvate thereof, with a polynucleotide template to perform templated transcription.

[0017] In a sixth aspect, the present invention provides a method for manufacturing a semiconductor device comprising: The present invention provides a capped mRNA transcription reaction system for purposes other than the diagnosis and treatment of disease, comprising: (1) a ribose-modified capped analog described in one aspect of the present invention, or a stereoisomer, pharmaceutically acceptable salt, or solvate thereof; and (2) a polynucleotide template, NTPs (nucleoside triphosphates), and an RNA polymerase.

[0018] In a seventh aspect, the present invention provides a kit comprising (1) a ribose-modified capped analog described in one aspect of the present invention, or a stereoisomer, pharmaceutically acceptable salt, or solvate thereof, and (2) a nucleoside triphosphate molecule and an RNA polymerase.

[0019] In an eighth aspect, the present invention provides a method for improving the intracellular stability of RNA, comprising incorporating into RNA a ribose-modified capped analog described in one aspect of the present invention, or a stereoisomer, pharmaceutically acceptable salt, or solvate thereof.

[0020] In a ninth aspect, the present invention provides a method for introducing RNA into a cell, comprising the step of contacting the cell with a ribose-modified capped analogue or a stereoisomer, pharmaceutically acceptable salt or solvate thereof described in the first aspect of the invention, or a pharmaceutical composition described in the fourth aspect of the invention.

[0021] In a tenth aspect, the present invention provides a method for inhibiting RNA translation in a cell, comprising contacting the cell with a ribose-modified capped analog or a stereoisomer, pharmaceutically acceptable salt or solvate thereof described in one aspect of the invention, or a pharmaceutical composition described in the fourth aspect of the invention.

[0022] In a tenth aspect, the present invention provides the use of a ribose-modified capped analogue or a stereoisomer, a pharmaceutically acceptable salt or solvate thereof according to the first aspect of the invention, or a pharmaceutical composition according to the fourth aspect of the invention, in the manufacture of a vaccine.

[0023] Compared with the prior art, the present invention has the following beneficial effects:

[0024] Compared with the ribose-modified capped analogs of the prior art, the ribose-modified capped analogs of the present invention can significantly improve the in vitro transcription yield of mRNA, capping rate, mRNA translation efficiency, and mRNA protein expression level and duration in mice, while significantly reducing the decapping rate. [Brief explanation of the drawings]

[0025] [Figure 1]1 is a graph showing the results of in vitro transcription yields of mRNA using YK-CAP-106, YK-CAP-107, YK-CAP-108, YK-CAP-109, YK-CAP-110, YK-CAP-111, YK-CAP-112, YK-CAP-113, YK-CAP-114, YK-CAP-115, YK-CAP-116, YK-CAP-117, YK-CAP-118, YK-CAP-119, YK-CAP-101, YK-CAP-102, YK-CAP-103, YK-CAP-104, YK-CAP-105, Compound 14, and Compound 5227 as capped analogs.

[0026] [Figure 2] 1 is a graph showing the results of mRNA transcript capping rates initiated with YK-CAP-106, YK-CAP-107, YK-CAP-108, YK-CAP-109, YK-CAP-110, YK-CAP-111, YK-CAP-112, YK-CAP-113, YK-CAP-114, YK-CAP-115, YK-CAP-116, YK-CAP-117, YK-CAP-118, YK-CAP-119, YK-CAP-101, YK-CAP-102, YK-CAP-103, YK-CAP-104, YK-CAP-105, Compound 14, and Compound 5227 as capped analogs.

[0027] [Figure 3] 1 is a graph showing the results of a relative fluorescence intensity test of capped mRNA using YK-CAP-106, YK-CAP-107, YK-CAP-108, YK-CAP-109, YK-CAP-110, YK-CAP-111, YK-CAP-112, YK-CAP-113, YK-CAP-114, YK-CAP-115, YK-CAP-116, YK-CAP-117, YK-CAP-118, YK-CAP-119, YK-CAP-101, YK-CAP-102, YK-CAP-103, YK-CAP-104, YK-CAP-105, 5227, CAP-2'O-ethyl, N-7113, Compound 14, HN3002, and m6A as capped analogs.

[0028] [Figure 4] 1 is a graph showing the results of decapping rates of DCP2 enzyme using YK-CAP-106, YK-CAP-107, YK-CAP-108, YK-CAP-109, YK-CAP-110, YK-CAP-111, YK-CAP-112, YK-CAP-113, YK-CAP-114, YK-CAP-115, YK-CAP-116, YK-CAP-117, YK-CAP-118, YK-CAP-119, YK-CAP-101, YK-CAP-102, YK-CAP-103, YK-CAP-104, YK-CAP-105, 5227, CAP-2′O-ethyl, N-7113, compound 14, HN3002, and m6A as capped analogs. DETAILED DESCRIPTION OF THE INVENTION

[0029] The technical solutions of the present invention are further described in the following specific embodiments, and those skilled in the art will understand that these embodiments are only to help understand the present invention and should not be considered as specifically limiting the present invention.

[0030] The present invention may be embodied in other specific forms without departing from the essential characteristics of the invention. It is to be understood that, unless inconsistent, any and all embodiments of the present invention may be combined with technical features of any one or more other embodiments to obtain additional embodiments. The present invention includes such additional embodiments obtained from such combinations.

[0031] Except in the examples, or where otherwise indicated, all numerical values ​​indicating quantitative properties, such as dosages, set forth in this disclosure are to be understood as being modified in all instances by the term "about." It is also to be understood that any numerical range set forth herein is intended to include all subranges within that range, as well as any combination of the individual endpoints of such ranges or subranges.

[0032] Unless otherwise defined, all terms (including technical and scientific terms) used in the present invention have the same meaning as commonly understood by a person skilled in the art to which the present invention belongs. Terms defined in commonly used dictionaries should be interpreted as having a meaning consistent with the meaning in the context of the relevant art, and should not be interpreted in an idealized or overly formal sense unless explicitly defined in the present invention.

[0033] As used herein, the term "C1-C3" refers to the group having any integer number of carbon atoms in the main chain ranging from 1 to 3, for example, 1, 2, or 3 carbon atoms. 6-15 The term "" refers to the number of carbon atoms in the group being any integer value between 6 and 15, e.g., 6, 7, 8, 9, 10, 11, 12, 13, 14, or 15 carbon atoms. Other carbon atom range limitations can be inferred from this, and all mean that the number of carbon atoms in the limited group can be any integer value within the limited range.

[0034] As used herein, the term "alkyl" refers to a saturated aliphatic hydrocarbon group having a straight or branched chain, non-limiting examples of which include methyl, ethyl, propyl, isopropyl, and the like.

[0035] As used herein, the term "salt" refers to the corresponding salt of a modified nucleoside compound (or nucleotide compound) of the invention that is convenient or desirable for preparation, purification, and / or handling, e.g., a pharmaceutically acceptable salt. Unless otherwise specified, a reference to a particular compound in the invention also includes its salt forms.

[0036] As used herein, "capped analog" refers to a special structure located at the 5' end of mature mRNA, i.e., the m7GPPPN structure, also known as the methylguanosine cap, formed by post-transcriptional modification in eukaryotes. Such structures may have the effect of preventing mRNA degradation at the 5' end, assisting RNA transcripts to pass through selective pores in the nuclear membrane into the cytoplasm, enhancing translation, and aiding in the completion of the overall shearing process.

[0037] As used herein, the terms "comprise," "contain," "include," and similar terms such as "comprise," mean that the elements appearing before the term include the elements listed after the term and their equivalents, and do not exclude elements not listed. As used herein, the terms "comprise" or "include" may refer to open, semi-closed, and closed systems. In other words, the terms also include "consisting essentially of" or "consisting of."

[0038] In the present invention, the term "pharmaceutically acceptable" refers to a compound or composition that is chemically and / or toxicologically compatible with other ingredients that make up the formulation and / or with humans or mammals in which it is used to prevent or treat a disease or disorder.

[0039] As used herein, the term "solvate" refers to a complex formed by combining a compound of formula (I) or a pharmaceutically acceptable salt thereof with a solvent (e.g., ethanol or water). Any solvate of a compound of formula (I) used to treat a disease or disorder may provide different properties (including pharmacokinetic properties), but upon absorption by a subject, the compound of formula (I) is obtained, and it should be understood that each use of a compound of formula (I) includes use of any solvate of a compound of formula (I).

[0040] It should further be understood that the compounds of formula (I) or pharmaceutically acceptable salts thereof may be isolated as solvates, and therefore, all such solvates are included within the scope of the present invention. For example, the compounds of formula (I) or pharmaceutically acceptable salts thereof may exist in unsolvated forms as well as solvated forms formed with pharmaceutically acceptable solvents (e.g., water, ethanol, etc.).

[0041] The present invention also includes salts of the compounds described herein, particularly pharmaceutically acceptable salts. Compounds of the present invention that have sufficiently acidic or sufficiently basic functional groups can react with many bases or acids to form salts. Alternatively, compounds that themselves carry a charge (e.g., compounds containing quaternary nitrogen) can form salts with a suitable counterion (e.g., a halogen ion such as bromide, chloride, or fluoride, especially bromide).

[0042] Pharmaceutically acceptable salts according to the present invention may be, for example, acid addition salts of the compounds of the present invention that are sufficiently basic and have a nitrogen atom in the chain or ring of the compound of the structure represented by formula (I), such as acid addition salts formed with inorganic acids such as hydrochloric acid, hydrobromic acid, hydroiodic acid, sulfuric acid, phosphoric acid or nitric acid, or with acids such as formic acid, acetic acid, acetoxyacetic acid, pyruvic acid, trifluoroacetic acid, propionic acid, butyric acid, hexanoic acid, heptanoic acid, undecylic acid, lauric acid, benzoic acid, salicylic acid, 2-(4-hydroxybenzoyl)benzoic acid, camphorsulfonic acid, cinnamic acid, cyclopentanepropionic acid, 3-hydroxy-2-naphthoic acid, nicotinic acid, pamoic acid, pectic acid, persulfate, 3-fluoroacetic acid, benzoic ... and acid addition salts formed with organic acids such as phenylpropionic acid, picric acid, pivalic acid, 2-hydroxyethanesulfonic acid, itaconic acid, sulfamic acid, trifluoromethanesulfonic acid, lauryl sulfuric acid, ethanesulfonic acid, benzenesulfonic acid, p-toluenesulfonic acid, methanesulfonic acid, 2-naphthalenesulfonic acid, naphthalenedisulfonic acid, camphorsulfonic acid, citric acid, tartaric acid, stearic acid, lactic acid, oxalic acid, malonic acid, succinic acid, malic acid, adipic acid, alginic acid, maleic acid, fumaric acid, D-gluconic acid, mandelic acid, ascorbic acid, glucoheptanoic acid, glycerophosphoric acid, aspartic acid, sulfosalicylic acid, and thiocyanic acid.

[0043] Furthermore, other suitable pharmaceutically acceptable salts of compounds of the invention that are sufficiently acidic are, for example, alkali metal salts such as sodium or potassium salts, alkaline earth metal salts such as calcium or magnesium salts, ammonium salts (e.g., salts formed with NH or aqueous ammonia), or salts formed with organic bases that provide a physiologically acceptable cation, such as, for example, salts formed with triethylamine, N-methylglucamine, dimethylglucamine, ethylglucamine, lysine, dicyclohexylamine, 1,6-diaminohexane, ethanolamine, glucosamine, sarcosine, serinol, trishydroxymethylaminomethane, 3-amino-1,2-propanediol, 1-amino-2,3,4-butanetriol, and the like. Additionally, basic nitrogen-containing groups can be quaternized with reagents such as lower alkyl halides, such as methyl, ethyl, propyl, and butyl chlorides, bromides, and iodides; dialkyl sulfates, such as dimethyl sulfate, diethyl sulfate, dibutyl sulfate, and dipentyl sulfate; long chain halides, such as decyl, lauryl, myristyl, stearyl, and the like chlorides, bromides, and iodides; aralkyl halides, such as benzyl bromide and phenethyl bromide.

[0044] Those skilled in the art will also recognize that acid addition salts of the compounds of formula (I) according to the present invention can be prepared by reacting said compounds with an appropriate inorganic or organic acid by any of the known methods. Alternatively, base addition salts of acidic compounds of the present disclosure are prepared by reacting them with an appropriate base by a variety of known methods.

[0045] The present invention includes all possible salts of the compounds of formula (I) according to the present invention, which may be a single salt or any mixture of said salts in any proportion.

[0046] Certain compounds of the present invention may exist in one or more stereoisomeric forms. Stereoisomers include geometric isomers, diastereomers, and enantiomers. Therefore, the compounds of Formula (I) described in the present invention also include racemic mixtures, single stereoisomers, and optically active mixtures. It should be understood by those skilled in the art that certain stereoisomers may have superior efficacy and / or fewer side effects than other stereoisomers. Single stereoisomers and optically active mixtures can be obtained by methods such as chiral synthesis, chiral catalysis, and chiral resolution. Racemates can be chirally separated by chromatographic or chemical separation. For example, chiral acid-based resolving agents, such as chiral tartaric acid or chiral malic acid, can be added to form salts with the compounds of the present invention, and the salts can be separated by utilizing the physical and chemical properties of the products, such as differences in solubility.

[0047] In the present invention, if there is a conflict between the name of a compound and its structural formula, the structural formula shall take precedence.

[0048] The present specification is to be construed in accordance with the laws and principles of chemical bonding. In some instances, hydrogen atoms may be removed to introduce substituents at certain positions.

[0049] It should be understood that the term "compounds of the invention" as used herein may include compounds of formula (I), solvates thereof, pharmaceutically acceptable salts thereof, stereoisomers thereof and mixtures thereof, depending on the context.

[0050] The present invention provides a ribose-modified capped analog having the structure of formula (I) or a stereoisomer, pharmaceutically acceptable salt, or solvate thereof.

[0051] [ka]

[0052] wherein n is selected from 1 or 0, and m is selected from 1 or 0; X1 is selected from -C(OH)H- or a single bond; X2 is -CH-, -O-, -CN, -C(O)-,

[0053] [ka] or a single bond; X3 is selected from -CH2, -CFH- or -O-; R1 is selected from -OH or -NHAc; R2 is selected from -CH3, -F, -H, or is absent; R3 is selected from -OR6, -NHC(O)R7, -N(R8), -CN, -F, -C(O)N(R9), substituted or unsubstituted C1-C3 alkyl, -H, or is absent; R4 is selected from -CH3, -F or -H; R5 is -OR 10 , -NHC(O)R 11 , -N(R 12 )2, -CF2H, -F, -C(O)N(R 13 ) 2, substituted or unsubstituted C1-C3 alkyl, or absent; R6, R8, R 10 , R 11 , R 12 and R 13 are each independently selected from unsubstituted C1-C3 alkyl or -H; R7 is -OR 14 , -NHC(O)R 15 , —F or substituted or unsubstituted C1-C3 alkyl; R9 is selected from unsubstituted C2-C3 alkyl; R 14 and R 15 are each independently selected from unsubstituted C1-C3 alkyl.

[0054] In some embodiments, X2 is -O-.

[0055] In some embodiments, X2 is -O- and R2 is absent.

[0056] In some embodiments, X2 is -O- and R3 is -H.

[0057] In some embodiments, X2 is -O-, R2 is absent, and R3 is -H.

[0058] In some embodiments, X2 is -CH-.

[0059] In some embodiments, X2 is -CH- and R2 is -H, -CH3, or -F.

[0060] In some embodiments, X2 is -CH- and R3 is -N(CH3)2, -C(O)CH3, -OCH3, -NHC(O)CH3, or -F.

[0061] In some embodiments, X2 is -CH-, R2 is -H, -CH3, or -F, and R3 is -N(CH3)2, -C(O)CH3, -OCH3, -NHC(O)CH3, or -F.

[0062] In some embodiments, X2 is -CN.

[0063] In some embodiments, X2 is -CN and R2 is absent.

[0064] In some embodiments, X2 is -CN, R2 is absent, and R3 is absent.

[0065] In some embodiments, X2 is

[0066] [ka] is.

[0067] In some embodiments, X2 is

[0068] [ka] and R2 does not exist.

[0069] In some embodiments, X2 is

[0070] [ka] and R3 does not exist.

[0071] In some embodiments, X2 is

[0072] [ka] where R2 is absent and R3 is absent.

[0073] In some embodiments, X2 is -C(O)-.

[0074] In some embodiments, X2 is -C(O)- and R2 is absent.

[0075] In some embodiments, X2 is -C(O)- and R3 is -N(CH2CH3)2 or -N(CH2CH2CH3)2.

[0076] In some embodiments, X2 is -C(O)-, R2 is absent, and R3 is -N(CH2CH3)2 or -N(CH2CH2CH3)2. In some embodiments, X3 is -O-.

[0077] In some embodiments, X3 is -O- and R7 is -CH3 or -OCH3.

[0078] In some embodiments, X3 is -CH2-.

[0079] In some embodiments, X3 is -CH2- and R7 is -OCH3, -NHC(O)CH3, or -F.

[0080] In some embodiments, X3 is -CFH-.

[0081] In some embodiments, X3 is -CFH- and R7 is -F.

[0082] In some embodiments, m is 1.

[0083] In some embodiments, m is 1 and R5 is -OCH3, -F or -CF2H, preferably, m is 1 and R5 is -OCH3.

[0084] In some embodiments, R6 is -H.

[0085] In some embodiments, the ribose-modified capped analog is as represented by the following formula: YK-CAP-101, YK-CAP-102, YK-CAP-103, YK-CAP-104, YK-CAP-105, YK-CAP-106, YK-CAP-107, YK-CAP-108, YK-CAP-109, YK-CAP-110, YK-CAP-111, YK-CAP-112, YK-CAP-113, YK-CAP-114, YK-CAP-115, YK-CAP-116, YK-CAP-117, YK-CAP-118, or YK-CAP-119.

[0086] [ka]

[0087] [ka]

[0088] [ka]

[0089] [ka]

[0090] [ka]

[0091] In a second aspect, the present invention provides the use of a ribose-modified capped analogue described in one aspect of the present invention, or a stereoisomer, pharmaceutically acceptable salt or solvate thereof, in the preparation of an in vitro co-transcriptional capping reagent for mRNA.

[0092] In a third aspect, the present invention provides an RNA molecule comprising, as a cap structure or cap structure fragment, a ribose-modified capped analog described in the first aspect of the present invention, or a stereoisomer, pharmaceutically acceptable salt, or solvate thereof.

[0093] In a fourth aspect, the present invention provides a pharmaceutical composition comprising an RNA molecule according to the third aspect of the invention.

[0094] In one embodiment, the pharmaceutical composition further comprises at least one RNA delivery agent.

[0095] The RNA delivery agent may be, for example, lipid nanoparticles (LNPs). Lipid nanoparticles are widely used for the delivery of small molecule drugs and nucleic acids, and mRNA coated with LNPs is protected from the effects of extracellular ribonucleases, facilitating intracellular delivery of the mRNA. For lipid nanoparticles, see the review article Chemistry of Lipid Nanoparticles for RNA Delivery. Acc Chem Res. 2022 Jan 4;55(1):2-12.

[0096] In one embodiment, the at least one RNA delivery agent comprises at least one cationic lipid.

[0097] The term cationic lipid used herein refers to the lipid that is positively charged at selected pH value.For example, refer to the cationic lipid disclosed in documents such as WO2023133946A1, CN115745820A, Chemistry of Lipid Nanoparticles for RNA Delivery.According to Chem Res.2022 Jan 4;55(1):2-12.

[0098] In one embodiment, the cationic lipid is selected from one or a combination of at least two of the following compounds:

[0099] (1) A compound represented by formula (II), or an -N-oxide, solvate, pharmaceutically acceptable salt, or stereoisomer thereof, wherein G1 is C 1~6 alkylene, and G2 is C 2~8 alkylene, and G3 is C 1~3 alkylene, and L1 is C 6~15 is a straight chain alkyl, and L2 is C 12~25 is a branched alkyl;

[0100] [ka]

[0101] (2) A compound represented by formula (III), or an -N-oxide, solvate, pharmaceutically acceptable salt, or stereoisomer thereof, wherein G1 is C 2~8 alkylene, and G2 is C 2~8 alkylene, L1 is -C(O)O- or -OC(O)-, L2 is -C(O)O- or -OC(O)-, and R1 is C 6~25 is a straight or branched chain alkyl, and R2 is C 6~25 is a straight or branched chain alkyl, G3 is HO(CH2)2- or HO(CH2)3-, G4 is HO(CH2)2- or HO(CH2)3-, and L is (CH2)2- or -(CH2)3- or -(CH2)4-;

[0102] [ka]

[0103] (3) A compound represented by formula (IV), or an -N-oxide, solvate, pharmaceutically acceptable salt, or stereoisomer thereof, wherein G1 is C 1~6 alkylene, and G2 is C 2~8 alkylene, and R1 is C 6~20 is a straight or branched chain alkyl, and R2 is C 12~25 is a branched alkyl, and G3 is HO(CH2)2N(CH3)(CH2)2-, HO(CH2)2N(CH2CH3)(CH2)2-, (HO(CH2)2)2N(CH2)2-, CHO(CH2)2N(CH3)(CH2)2-, (CH3)2N(CH2)3SC(O)O(CH2)2-, (CH3)2N(CH2)3SC(O)-, CH3NH(CH2)2N(CH3)(CH2)2- or CH3CH2NH(CH2)2-;

[0104] [ka]

[0105] (4) A compound represented by formula (V), or an -N-oxide, solvate, pharmaceutically acceptable salt, or stereoisomer thereof, wherein G1 is C 1~8 alkylene, and G2 is C 2~8 alkylene, and R1 is C 6~25 is a straight or branched chain alkyl, and R2 is C 12~25 is a straight or branched chain alkyl, G3 is HO(CH2)2N(R3)CH2CH(OH)CH2-, where R3 is -CH3 or -CH2CH3 or -CH2CH2OH;

[0106] [ka]

[0107] (5) A compound represented by formula (VI), or an -N-oxide, solvate, pharmaceutically acceptable salt, or stereoisomer thereof, wherein: 1 and G 2 are each independently unsubstituted C6 to C 10 alkylene, and G 3 is unsubstituted C1~C 12 alkylene, and R 1 and R 2 are C6 to C 24 Alkyl or C6-C 24 alkenyl, and R 3 is OR 5 , N, -C(=O)OR 4 , -OC(=O)R 4 or -NR 5 C(=O)R 4 and R 4 is C1~C 12 is hydrocarbyl, and R 5 is H or C1-C6 hydrocarbyl,

[0108] [ka]

[0109] (6) A compound represented by formula (VII), or an -N-oxide, solvate, pharmaceutically acceptable salt, or stereoisomer thereof, wherein R4 is -(CH2) n Q and -(CH2) n CHQR, where Q is -OR, -OH, or -O(CH2) n is selected from the group consisting of N(R), -OC(O)R, -CX, -CN, -N(R)C(O)R, -N(H)C(O)R, -N(R)S(O)R, -N(H)S(O)R, -N(R)C(O)N(R), -N(H)C(O)N(R), -N(H)C(O)N(H)(R), -N(R)C(S)N(R), -N(H)C(S)N(R), -N(H)C(S)N(H)(R), -N(R)S(O)R and heterocyclyl;

[0110] [ka]

[0111] (7) A compound represented by formula (VIII), or an -N-oxide, solvate, pharmaceutically acceptable salt or stereoisomer thereof.

[0112] [ka]

[0113] In one preferred embodiment, the cationic lipid is selected from one or a combination of at least two of YK-009, YK-401, YK-305, ALC0315, SM102, DLIN-MC3-DMA.

[0114] [ka]

[0115] In a more preferred embodiment, the cationic lipid is YK-009.

[0116] In one embodiment, the at least one RNA delivery agent further comprises at least one neutral lipid.

[0117] In the present invention, neutral lipids refer to auxiliary lipids that exist in uncharged or zwitterionic form at selected pH values, which may modulate the mobility of nanoparticles into lipid bilayer structures by promoting lipid phase transition, improving efficacy and potentially affecting target organ specificity.

[0118] In one embodiment, the neutral lipid comprises one or a combination of at least two of phosphatidylcholine, phosphatidylethanolamine, sphingomyelin, ceramide, sterol, and derivatives thereof.

[0119] In one embodiment, the neutral lipid is 1,2-dilinoleoyl-sn-glycero-3-phosphocholine (DLPC), 1,2-dimyristoyl-sn-glycero-3-phosphocholine (DMPC), 1,2-dioleoyl-sn-glycero-3-phosphocholine (DOPC), 1,2-dipalmitoyl-sn-glycero-3-phosphocholine (DPPC), 1,2-distaloyl-sn-glycero-3-phosphocholine (DSPC), 1,2-diundecanoyl-sn-glycero-3-phosphocholine (DUPC), 1-palmitoyl-2-oleoyl-sn-glycero-3-phosphocholine (POPC), 1,2-di-O-octadecenyl-sn-glycero-3-phosphocholine (18:0 Diether PC), 1-oleoyl-2-cholesterylhemisuccinoyl-sn-glycero-3-phosphocholine (OChemsPC), 1-hexadecyl-sn-glycero-3-phosphocholine (C16 Lyso PC), 1,2-dilinolenoyl-sn-glycero-3-phosphocholine, 1,2-diarachidonoyl-sn-glycero-3-phosphocholine, 1,2-didocosahexaenoyl-sn-glycero-3-phosphocholine, 1,2-dioleoyl-sn-glycero-3-phosphoethanolamine (DOPE), 1,2-diphytanoyl-sn-glycero-3-phosphoethanolamine (ME 16).0 PE), 1,2-distearoyl-sn-glycero-3-phosphoethanolamine, 1,2-dilinoleoyl-sn-glycero-3-phosphoethanolamine, 1,2-dilinolenoyl-sn-glycero-3-phosphoethanolamine, 1,2-diarachidonoyl-sn-glycero-3-phosphoethanolamine, 1,2-didocosahexaenoyl-sn-glycero-3-phosphoethanolamine, 1,2-dioleoyl-sn-glycero-3-phospho-rac-(1-glycerol) sodium salt (DOPG), dipalmitoylphosphatidylglycerol (DPPG), palmitoylphosphatidylethanolamine (POPE) The composition may comprise one or a combination of at least two of the following: distearoylphosphatidylethanolamine (DSPE), dipalmitoylphosphatidylethanolamine (DPPE), dimyristoylphosphatidylethanolamine (DMPE), 1-stearoyl-2-oleoyl-sn-glycero-3-phosphoethanolamine (SOPC), sphingomyelin, phosphatidylcholine, phosphatidylethanolamine, phosphatidylserine, phosphatidylinositol, phosphatidic acid, palmitoyloleoylphosphatidylcholine, lysophosphatidylcholine, and lysophosphatidylethanolamine (LPE).

[0120] In one preferred embodiment, the neutral lipid is DOPE and / or DSPC.

[0121] In one embodiment, the at least one RNA delivery agent further comprises a structured lipid.

[0122] In the present invention, a structured lipid refers to a lipid that enhances the stability of nanoparticles by filling the gaps between lipids.

[0123] In one embodiment, the structured lipid comprises one or a combination of at least two of cholesterol, a non-sterol, sitosterol, ergosterol, campesterol, stigmasterol, brassicasterol, tomatine, ursolic acid, α-tocopherol, or a corticosteroid.

[0124] In one preferred embodiment, the structured lipid is cholesterol.

[0125] In one embodiment, the at least one RNA delivery agent further comprises a polymer-conjugated lipid.

[0126] In the present invention, polymer-conjugated lipids are primarily polyethylene glycol (PEG)-modified lipids. Hydrophilic PEG stabilizes lipid nanoparticles (LNPs), regulates nanoparticle size by limiting lipid fusion, and extends the half-life of nanoparticles by reducing nonspecific interactions with macrophages.

[0127] In one embodiment, the polymer-conjugated lipid comprises one or a combination of at least two of distearoylphosphatidylethanolamine-polyethylene glycol 2000 (DSPE-PEG2000), dimyristoylglycerol-polyethylene glycol 2000 (DMG-PEG2000), and methoxypoly(ethylene glycol) ditetradecylacetamide (ALC-0159).

[0128] In one embodiment, the RNA delivery agent comprises a neutral lipid, a structured lipid, and a polymer-conjugated lipid, and the molar ratio of the cationic lipid, the neutral lipid, the structured lipid, and the polymer-conjugated lipid is (25-75):(5-25):(15-65):(0.5-10), for example, (35-49):(7.5-15):(35-55):(1-5).

[0129] In one embodiment, the pharmaceutical composition further comprises one or at least two cell-penetrating peptides.

[0130] The present invention provides a kit comprising: (1) a ribose-modified capped analog described in one aspect of the present invention, or a stereoisomer, pharmaceutically acceptable salt, or solvate thereof; and (2) a nucleotide triphosphate molecule and an RNA polymerase.

[0131] In one embodiment, the kit comprises an RNase inhibitor, inorganic pyrophosphatase, Mg 2+ , a crowding agent, or a buffer, or a combination of at least two of the above.

[0132] The present invention may be embodied in other specific forms without departing from the essential characteristics of the invention. It is to be understood that, unless inconsistent, any and all embodiments of the present invention may be combined with technical features of any one or more other embodiments to obtain additional embodiments. The present invention includes such additional embodiments obtained from such combinations.

[0133] Hereinafter, the embodiments of the present invention will be described in detail in combination with examples. Those skilled in the art will understand that the following examples are used only to illustrate the present invention and do not limit the scope of the present invention. If specific conditions are not described in the examples, they are carried out according to conventional conditions or conditions recommended by the manufacturer. If the manufacturers of the reagents or instruments used are not described, they are all commercially available conventional products.

[0134] The following abbreviations in Roman letters represent the following reagents: IBX: 2-iodoxybenzoic acid, BF3·Et2O: boron trifluoride diethyl ether, Allyltrimethylsilane: allyltrimethylsilane, TEA: triethylamine, Ac2O: acetic anhydride, HOAc: acetic acid, conc H2SO4: concentrated sulfuric acid, BSA: N,O-bis(trimethylsilyl)acetamide, TMSOTf: trimethylsilyl trifluoromethanesulfonate, Toluene: toluene, MeOH: methanol, Boc2O: di-tert-butyl dicarbonate, DIEA: N,N-diisopropylethylamine, DMAP: 4-dimethylaminopyridine, DMSO: dimethyl sulfoxide, HATU: O-(7-azabenzotriazol-1-yl)-N,N,N',N'-tetramethyluronium hexafluorophosphate, THF: tetrahydrofuran, TBSCl: tert-butyldimethylsilyl chloride, Imidazole: imidazole, DMF: N,N-dimethylformamide, TBAF: tetra-n-butylammonium fluoride, TBSOTf: tert-butyldimethylsilyl trifluoromethanesulfonate, NMO: N-methylmorpholine N-oxide, m-CPBA: metachloroperbenzoic acid, DIAD: diisopropyl azodicarboxylate, NCS: N-chlorosuccinimide, PO(MeO)3: trimethyl phosphate, PySSPy: 2,2'-dithiodipyridine, PPh3: triphenylphosphine, TEAP: triethylammonium phosphate, TEAB: triethylammonium bicarbonate, MTBE: methyl tert-butyl ether, DCM: dichloromethane, EA: ethyl acetate, DAST: diethylaminosulfur trifluoride, AcSH: thioacetic acid.

[0135] Example 1: 1. Synthesis of intermediate INT-I

[0136] [ka]

[0137] Step 1: Synthesis of INT-I-PM1 2-Amino-9H-purin-6-ol (50.0 g, 0.33 mol) was dissolved in N,N-dimethylacetamide (500 mL), acetic anhydride (100 mL, 1.06 mol) was added, and the mixture was heated to 160 °C and stirred until the solution became clear. After the reaction was complete, the mixture was stopped and allowed to cool to room temperature. A large amount of solid precipitated. The solid was filtered and washed with ethanol until it became white to give INT-I-PM1 (60.0 g, 0.31 mol, 94.1%). C7H7N5O2, MS (ES): m / z (M+H) + )194.1.

[0138] Step 2: Synthesis of INT-I INT-I-PM1 (60.0 g, 0.31 mol) was dissolved in pyridine (200 mL), N,N-diisopropylethylamine (120.2 g, 0.93 mol) was added, and the mixture was cooled to 0 °C. Diphenylcarbamic acid chloride (86.2 g, 0.37 mol) was dissolved in pyridine (100 mL) and slowly added dropwise to the reaction mixture in an ice bath. After the addition was complete, the ice bath was removed, and the mixture was allowed to warm to room temperature. The mixture was then stirred for 3 hours, and the reaction was complete, as monitored by LCMS to confirm that no raw materials remained. 100 mL of water was added to the reaction mixture, and the mixture was dried under reduced pressure. The residue was added to 800 mL of a mixed solvent of ethanol and water (1:1 volume ratio), heated to reflux for 2 hours, and then cooled to room temperature. A large amount of solid precipitated, which was filtered and washed with ethanol to give INT-I (59.7 g, 0.15 mol, 49.6%). 20 H 16 N6O3, MS(ES): m / z (M+H + )389.1.

[0139] 2. Synthesis of intermediate INT-II

[0140] [ka]

[0141] pA(2'-OMe)mpG·TEA (300.1 mg, 0.37 mmol), imidazole (347.2 mg, 5.10 mmol), dithiodipyridine (1123.6 mg, 5.10 mmol), and triethylamine (516.1 mg, 5.10 mmol) were dissolved in 2.0 mL of Superdry N,N-dimethylformamide. Triphenylphosphine (1337.7 mg, 5.10 mmol) was added under nitrogen gas protection, and the mixture was allowed to react at 25 °C for 4 hours. After completion of the reaction, the reaction solution was slowly added to a pre-cooled acetone solution containing sodium iodide (598.1 mg, 3.99 mmol) and allowed to crystallize at 25 °C for 30 minutes. After centrifugation, a white solid, INT-II (240.6 mg, 0.30 mmol, yield: 80.9%), was obtained. 24 H 30 N 12 O 13 P2, MS(ES): m / z(MH - ):755.2.

[0142] 3. Synthesis of YK-CAP-101

[0143] [ka]

[0144] Step 1: Synthesis of YK-CAP-101-PM1 2-Amino-6-chloropurine (6.29 g, 37.09 mmol), toluene (50 mL), and BSA (15.09 g, 74.19 mmol) were added to a 250 mL single-neck flask. The reaction mixture was heated to 80 °C and stirred until the reaction mixture became clear. The mixture was then allowed to cool to room temperature. α-D-Glucose pentaacetate (10.00 g, 25.62 mmol) was dissolved in 20 mL of toluene and added to the reaction mixture. The mixture was stirred at room temperature for 5 minutes. TMSOTf (8.24 g, 37.09 mmol) was added at room temperature, and the reaction mixture was heated to 110 °C and allowed to react at 110 °C for 3 hours. After the reaction was completed, the mixture was allowed to cool to room temperature, 100 mL of saturated sodium bicarbonate solution and 100 mL of ethyl acetate were added, stirred for 5 minutes, filtered through diatomaceous earth, separated, the aqueous phase was extracted with 100 mL of ethyl acetate, the ethyl acetate phases were combined, washed with 100 mL of saturated brine, dried over anhydrous sodium sulfate, filtered, and the filtrate was spin-dried under reduced pressure to obtain a viscous material, which was purified by silica gel column chromatography (0-100% ethyl acetate / n-hexane), and the product was collected and concentrated to obtain YK-CAP-101-PM1 (8.40 g, 16.80 mmol, 65.6%). 19 H 22 ClNO, MS (ES): m / z (M+H + )500.1.

[0145] Step 2: Synthesis of YK-CAP-101-PM2 YK-CAP-101-PM1 (8.40 g, 16.80 mmol) and aqueous sodium hydroxide (1 M, 40 mL, 40 mmol) were added to a 250 mL single-neck flask, and the reaction was heated to reflux with stirring for 3 hours. The reaction was cooled to room temperature and spun to dryness under reduced pressure. The residue was slurried in 100 mL (methanol:dichloromethane = 1:20) for 10 minutes and filtered to give 6.0 g of a brown solid, which was purified by high-pressure preparative liquid phase separation to give YK-CAP-101-PM2 (2.13 g, 6.80 mmol, 40.5%). 11 H 15 N5O6, MS(ES): m / z (M+H + )314.1. YK-CAP-101-PM2:1 HNMR(400MHz,D2O)δ8.71(s,1H),5.57(d,J=9.2Hz,1H),3.99(dd,J=9.2,9.2Hz,1H),3.86(dd,J=12.4,2.0Hz,1H),3.67-3.55(m,4H).

[0146] Step 3: Synthesis of YK-CAP-101-PM3 YK-CAP-101-PM2 (300 mg, 0.96 mmol) was dissolved in 3 mL of trimethyl phosphate. Under nitrogen gas protection, the mixture was cooled to 0°C, and phosphorus oxychloride (450 mg, 2.93 mmol) was slowly added dropwise. The mixture was stirred at 0°C for approximately 3 hours. After the reaction was complete, 5 mL of water was added, the mixture was cooled to room temperature, and the mixture was stirred for approximately 1.5 hours. After that, 10 mL of dichloromethane was added and the mixture was washed. The mixture was allowed to stand and separated. The upper aqueous phase was collected and concentrated under reduced pressure. The concentrated mixture was diluted to 180 mL with water and purified on a gel column (eluted with water:1.5 M TEAB = 10:1). The peak of the target product was collected, concentrated, lyophilized, and dried to obtain YK-CAP-101-PM3 (triethylamine salt, 340 mg, 0.69 mmol, 71.6%) as a white solid. 11 H 16 N5O9P, MS(ES): m / z(MH - )392.1.

[0147] Step 4: Synthesis of YK-CAP-101-PM4 YK-CAP-101-PM3 (340 mg, 0.69 mmol), imidazole (706 mg, 10.38 mmol), 2,2'-dithiodipyridine (2287 mg, 10.38 mmol), triethylamine (1050 mg, 10.38 mmol), and triphenylphosphine (2723 mg, 10.38 mmol) were dissolved in 4 mL of dry N,N-dimethylformamide. The mixture was stirred at room temperature for approximately 4 hours under nitrogen gas protection. After completion of the reaction, the reaction mixture was poured into a solution of sodium iodide (1225 mg, 8.17 mmol) in acetone (6 mL). The mixture was stirred at room temperature for 30 minutes and then centrifuged to obtain a crude precipitate. The crude precipitate was washed with acetone, separated, and the lower precipitate was collected and lyophilized to obtain a white solid, YK-CAP-101-PM4 (sodium salt, 309 mg, 0.66 mmol, 96.2%). 14 H 18 N7O8P, MS(ES): m / z(MH - )442.1.

[0148] Step 5: Synthesis of YK-CAP-101-PM5 YK-CAP-101-PM4 (309 mg, 0.66 mmol) and TEAP (458 mg, 2.30 mmol) were dissolved in dry N,N-dimethylformamide (5 mL), zinc chloride (215 mg, 1.58 mmol) was added, and the mixture was stirred at room temperature under nitrogen gas protection for approximately 21 hours. After the reaction was complete, MTBE (10 mL) was added to the reaction system, which was then washed with ultrasonic agitation. The mixture was then allowed to stand, the supernatant was poured off, and the mixture was then repeated once. The lower layer was collected and concentrated under reduced pressure. Water (80 mL) was added to dissolve the entire mixture, and the mixture was purified on a gel column (eluted with water and 1.5 M TEAB = 20:1). The peak of the target product was collected, concentrated, and lyophilized to yield YK-CAP-101-PM5 (triethylamine salt, 320 mg, 0.56 mmol, 84.4%) as a white solid. 11 H 17 N5O 12 P2, MS(ES): m / z(MH - )472.0.

[0149] Step 6: Synthesis of YK-CAP-101-PM6 YK-CAP-101-PM5 (320 mg, 0.56 mmol) and methyl iodide (960 mg, 6.76 mmol) were dissolved in dry N,N-dimethylformamide (4 mL) and stirred in an oil bath at 37 °C for approximately 23 hours. After the reaction was complete, water (5 mL) was added to the reaction mixture to dissolve it completely. The mixture was washed with EA (25 mL), separated, and the lower aqueous layer was collected and concentrated under reduced pressure. Water (50 mL) was added to dissolve it completely. The mixture was purified on a gel column (eluted with water and 1.5 M TEAB = 10:1). The target product peak was collected, concentrated, lyophilized, and further desalted by high-performance liquid chromatography (50 mM TEAB and methanol mobile phase system) to obtain YK-CAP-101-PM6 (triethylamine salt, 95 mg, 0.16 mmol, 28.8%) as a white solid. 12 H 19 N5O 12 P2, MS(ES): m / z(MH - )486.0.

[0150] Step 7: Synthesis of YK-CAP-101 YK-CAP-101-PM6 (95 mg, 0.16 mmol) and INT-II (219 mg, 0.27 mmol) were dissolved in dry dimethyl sulfoxide (1.2 mL), zinc chloride (518 mg, 3.80 mmol) was added, and the mixture was stirred in an oil bath at 37 °C for approximately 3 hours under nitrogen gas protection. After the reaction was complete, 0.25 M EDTA was added to dissolve the mixture completely. The pH was adjusted to 6-7 with 1.5 M TEAB, and the mixture was purified on a gel column (eluted with water and 1.5 M TEAB = 10:1). The peak of the target product was collected, concentrated, and lyophilized. Further purification by high-performance liquid chromatography yielded the final product YK-CAP-101 (20 mg, 16.30 μmol, 10.2%). 33 H 45 N 15 O 25 P4, MS(ES): m / z(MH - )1174.2. 1HNMR(400MHz,D2O)δ8.43(d,J=1.3Hz,1H),8.38(s,1H),8.06(d,J=6.0Hz,2H ),6.01(d,J=5.4Hz,1H),5.89(d,J=2.3Hz,1H),5.81(d,J=4.5Hz,1H),4.97- 4.86(m,3H),4.66(t,J=4.8Hz,1H),4.52-4.35(m,4H),4.32(d,J=4.5Hz,1H) ,4.21(s,3H),4.09(s,2H),3.97(s,3H),3.78(t,J=7.0Hz,1H),3.43(s,4H). 31 PNMR(D2O,162MHz)δ-0.92(s,1P),-11.11(d,J=19.3Hz,1P),-11.50(d,J=17.8Hz,1P),-22.23(t,J=17.8Hz,1P).

[0151] 4. Synthesis of YK-CAP-102

[0152] [ka]

[0153] Step 1: Synthesis of YK-CAP-102-PM1 Using α-D-glucosamine pentaacetate (10.0 g, 25.68 mmol) as a starting material, YK-CAP-102-PM1 (8.79 g, 17.62 mmol, 68.61%) was obtained using the same synthesis method as for YK-CAP-101-PM1. 19 H 23 ClNO, MS (ES): m / z (M+H + )499.1.

[0154] Step 2: Synthesis of YK-CAP-102-PM2 Using YK-CAP-102-PM1 (8.79 g, 17.62 mmol) as a raw material, YK-CAP-102-PM2 (2.37 g, 6.69 mmol, 37.96%) was obtained by the synthesis method of YK-CAP-101-PM2. 13 H 18N6O6, MS(ES): m / z (M+H + )355.2. YK-CAP-102-PM2: 1 HNMR(400MHz,DMSO-d6)δ8.41(s,1H),7.97(d,J=9.2Hz,1H),7.65(s,1H),6.76(s,2H),5.32(d,J=10. 2Hz,1H), 4.15(q,J=9.8Hz,1H),3.67(d,J=11.9Hz,1H),3.55-3.41(m,2H),3.25(s,2H),1.64(s,3H).

[0155] Step 3: Synthesis of YK-CAP-102-PM3 Using YK-CAP-102-PM2 (500 mg, 1.41 mmol) as a raw material, YK-CAP-102-PM3 (triethylamine salt, 702 mg, 1.31 mmol, 93.0%) was obtained via the synthetic route of YK-CAP-101-PM3. 13 H 19 N6O9P, MS(ES): m / z(MH - )433.1.

[0156] Step 4: Synthesis of YK-CAP-102-PM4 intermediate Using YK-CAP-102-PM3 (702 mg, 1.31 mmol) as a raw material, YK-CAP-102-PM4 (sodium salt, 521 mg, 1.03 mmol, 78.5%) was obtained via the synthetic route of YK-CAP-101-PM4. 16 H 21 N8O8P, MS(ES): m / z(MH - )483.1.

[0157] Step 5: Synthesis of YK-CAP-102-PM5 intermediate Using YK-CAP-102-PM4 (521 mg, 1.03 mmol) as a starting material, YK-CAP-102-PM5 (triethylamine salt, 338 mg, 0.55 mmol, 53.3%) was obtained via the synthetic route of YK-CAP-101-PM5. 13 H 20 NO 12 P2, MS(ES): m / z(MH- )513.0.

[0158] Step 6: Synthesis of YK-CAP-102-PM6 intermediate Using YK-CAP-102-PM5 (338 mg, 0.55 mmol) as a raw material, YK-CAP-102-PM6 (triethylamine salt, 158 mg, 0.25 mmol, 45.6%) was obtained via the synthetic route of YK-CAP-101-PM6. 14 H 22 NO 12 P2, MS(ES): m / z(MH - )527.1.

[0159] Step 7: Synthesis of YK-CAP-102 Using YK-CAP-102-PM6 (158 mg, 0.25 mmol) as a starting material, YK-CAP-102 (23.9 mg, 18.85 μmol, 7.5%) was obtained via the synthetic route of YK-CAP-101. 35 H 48 N 16 O 25 P4, MS(ES): m / z(MH - )1115.1. 1 HNMR(400MHz,D2O)δ8.42(d,J=1.3Hz,1H),8.38(s,1H),8.04(d,J=5.8Hz,2H), 6.00(d,J=5.4Hz,1H),5.88(d,J=2.3Hz,1H),5.81(d,J=4.4Hz,1H),4.95-4.83( m,3H),4.64(t,J=4.7Hz,1H),4.53-4.36(m,4H),4.30(d,J=4.4Hz,1H),4.20(s, 2H),4.06(s,2H),3.93(s,3H),3.75(t,J=7.0Hz,3H),3.62(s,3H),1.67(s,3H). 31 PNMR(D2O,162MHz)δ-0.92(s,1P),-11.21(d,J=19.2Hz,1P),-11.55(d,J=17.3Hz,1P),-23.43(t,J=17.6Hz,1P).

[0160] 5. Synthesis of YK-CAP-103

[0161] [ka]

[0162] Step 1: Synthesis of YK-CAP-103-PM1 (4R,5R)-5-((R)-1,2-dihydroxyethyl)-2,2-dimethyl-1,3-dioxolane-4-carbaldehyde (10.00 g, 52.58 mmol) and methyl 2-(triphenyl-5-phosphorimido)acetate (21.6 g, 64.60 mmol) were added to a single-neck flask containing acetonitrile (200 mL), heated to 90 °C, and stirred for 10 h. After completion of the reaction, the mixture was diluted with ethyl acetate (300 mL), washed with saturated brine, and dried over anhydrous sodium sulfate. The solvent was removed by vacuum decompression and spun down to give a yellow oily compound (26.7 g), which was used directly in the next step without further purification.

[0163] Step 2: Synthesis of YK-CAP-103-PM2 YK-CAP-103-PM1 (12.8 g, calculated as 25.21 mmol), tert-butyldiphenylchlorosilane (17.2 g, 62.42 mmol), and imidazole (5.31 g, 78.03 mmol) were sequentially added to a single-neck flask containing dichloromethane (200 mL) and stirred overnight at room temperature. After completion of the reaction, the mixture was diluted with dichloromethane (100 mL), washed with saturated brine (3 × 200 mL), dried over anhydrous sodium sulfate, filtered, and the filtrate was concentrated under reduced pressure. The residue was purified by silica gel column chromatography (0–17% ethyl acetate / n-hexane) to yield YK-CAP-103-PM2 (20.0 g, 41.27 mmol).

[0164] Step 3: Synthesis of YK-CAP-103-PM3 YK-CAP-103-PM2 (20.0 g, 41.27 mmol) was added to 150 mL of DCM in a single-neck flask and cooled to -78 °C. A toluene solution of diisobutylaluminum hydride (1.5 M, 63.2 mL, 94.8 mmol) was slowly added dropwise. After the addition was complete, the mixture was warmed to room temperature and stirred overnight. After the reaction was complete, 65 mL of methanol was slowly added in an ice bath, and a white fluffy solid formed. Sodium sulfate decahydrate was then added, and the mixture was stirred for 20 minutes. The solid was removed by suction filtration. The filtrate was spin-dried, and the residue was purified by silica gel column chromatography (0-30% ethyl acetate / n-hexane) to give YK-CAP-103-PM3 (15.00 g, 32.85 mmol, 79.6%).

[0165] Step 4: Synthesis of YK-CAP-103-PM4 YK-CAP-103-PM3 (15.00 g, 32.85 mmol), p-toluenesulfonyl chloride (7.5 g, 39.40 mmol), and triethylamine (5.0 g, 49.30 mmol) were sequentially added to a single-neck flask containing dichloromethane (60 mL) and stirred overnight at room temperature. After completion of the reaction, the mixture was diluted with dichloromethane (50 mL), washed with saturated brine (2 × 100 mL), dried over anhydrous sodium sulfate, filtered, and the filtrate was concentrated under reduced pressure. The residue was purified by silica gel column chromatography (0–20% ethyl acetate / n-hexane) to give YK-CAP-103-PM4 (17.00 g, 27.83 mmol, 84.7%).

[0166] Step 5: Synthesis of YK-CAP-103-PM5 YK-CAP-103-PM4 (17.00 g, 27.83 mmol) was dissolved in tetrahydrofuran (150 mL) in a single-neck flask and a 1 M solution of potassium tert-butoxide in tetrahydrofuran (61.0 mL, 61.0 mmol) was added dropwise at -40 °C. After the addition was complete, the reaction was stirred at room temperature for 3 h. The mixture was diluted with ethyl acetate (200 mL), washed with saturated brine (2 × 150 mL), dried over anhydrous sodium sulfate, filtered, and the filtrate was concentrated under reduced pressure. The residue was purified by silica gel column chromatography (0-11% ethyl acetate / n-hexane) to give YK-CAP-103-PM5 (3.29 g, 7.50 mmol, 27.0%).

[0167] Step 6: Synthesis of YK-CAP-103-PM6 YK-CAP-103-PM5 (3.29 g, 7.50 mmol) was dissolved in a mixture of tetrahydrofuran (25 mL) and water (5 mL). Potassium osmate(VI) dihydrate (140 mg, 0.38 mmol) and N-methylmorpholine N-oxide (1.05 g, 9.00 mmol) were added to the mixture, which was then heated to 40°C and stirred for 6 hours. After completion of the reaction, the mixture was diluted with ethyl acetate (100 mL), washed with saturated aqueous sodium sulfite (2 × 80 mL), dried over anhydrous sodium sulfate, filtered, and the solvent was removed under reduced pressure to give YK-CAP-103-PM6 (3.48 g, 7.36 mmol, 98.2%).

[0168] Step 7: Synthesis of YK-CAP-103-PM7 YK-CAP-103-PM6 (3.48 g, 7.36 mmol) was dissolved in a mixture of tetrahydrofuran (25 mL) and water (5 mL). Potassium periodate (2.54 g, 11.04 mmol) was then added to the mixture, which was then heated to 40°C and stirred for 6 hours. After completion of the reaction, the mixture was diluted with ethyl acetate (100 mL), washed with saturated aqueous sodium sulfite (2 x 50 mL), dried over anhydrous sodium sulfate, filtered, and concentrated under reduced pressure to give YK-CAP-103-PM7 (3.14 g, 7.13 mmol, 96.9%).

[0169] Step 8: Synthesis of YK-CAP-103-PM8 YK-CAP-103-PM7 (5.19 g, 11.78 mmol) was dissolved in a single-neck flask containing methanol (100 mL), and sodium borohydride (0.54 g, 13.18 mmol) was added in a batchwise manner under ice bath conditions. The mixture was then stirred at room temperature for 3 hours. After completion of the reaction, the mixture was diluted with water (150 mL) and extracted with ethyl acetate (2 × 150 mL). The combined organic phases were dried over anhydrous sodium sulfate, filtered, and concentrated under reduced pressure. The residue was purified by silica gel column chromatography (0-13% ethyl acetate / n-hexane) to give YK-CAP-103-PM8 (3.30 g, 7.46 mmol, 63.3%).

[0170] Step 9: Synthesis of YK-CAP-103-PM9 Using YK-CAP-103-PM8 (3.30 g, 7.46 mmol) as a raw material, YK-CAP-103-PM9 (3.42 g, 5.73 mmol, 76.8%) was obtained by the synthesis method of YK-CAP-103-PM4.

[0171] Step 10: Synthesis of YK-CAP-103-PM10 2-Amino-6-chloroguanine (1.17 g, 6.90 mmol) and lithium hydride (55 mg, 6.88 mmol) were dissolved in 30 mL of DMSO and stirred at 90 °C for 1 hour. After that, a DMSO solution (15 mL) of YK-CAP-103-PM9 (3.42 g, 5.73 mmol) was added and stirred for 5 hours. After the reaction was complete, the mixture was diluted with water (100 mL) and extracted with dichloromethane (2 × 150 mL). The combined organic phases were dried over anhydrous sodium sulfate, filtered, and concentrated under reduced pressure. The residue was purified by silica gel column chromatography (0-25% ethyl acetate / dichloromethane) to give YK-CAP-103-PM10 (1.90 g, 3.20 mmol, 55.9%). 30 H 36 ClNOSi, MS (ES): m / z (M+H + )595.2.

[0172] Step 11: Synthesis of YK-CAP-103-PM11 YK-CAP-103-PM10 (1.90 g, 3.20 mmol) was dissolved in tetrahydrofuran (40 mL), and then 1 M HCl (80 mL) was added. The mixture was stirred at 90°C for 7 hours to react. After the reaction was completed, the solvent was removed under reduced pressure, and the residue was purified by high-performance preparative liquid chromatography to obtain the white solid compound YK-CAP-103-PM11 (587 mg, 1.97 mmol, 61.7%). 11 H 15 N5O5, MS(ES): m / z (M+H + )298.2. YK-CAP-103-PM11: 1 HNMR(400MHz,DMSO-d6)δ10.57(s,1H),7.68(s,1H),6.46(s,2H),5.00-4.53(m, 2H),4.17-4.09(m,1H),4.00-3.87(m,2H),3.77-3.61(m,3H),3.46-3.32(m,3H).

[0173] Step 12: Synthesis of YK-CAP-103-PM12 Using YK-CAP-103-PM11 (300 mg, 1.01 mmol) as a raw material, YK-CAP-103-PM12 (triethylamine salt, 350 mg, 0.73 mmol, 72.5%) was obtained via the synthetic route of YK-CAP-101-PM3. 11 H 16 N5O8P, MS(ES): m / z(MH - )376.1.

[0174] Step 13: Synthesis of YK-CAP-103-PM13 Using YK-CAP-103-PM12 (350 mg, 0.73 mmol) as a raw material, YK-CAP-103-PM13 (sodium salt, 320 mg, 0.71 mmol, 97.4%) was obtained via the synthetic route of YK-CAP-101-PM4. 14 H 18 N7O7P, MS(ES): m / z(MH - )426.1.

[0175] Step 14: Synthesis of YK-CAP-103-PM14 Using YK-CAP-103-PM13 (320 mg, 0.71 mmol) as a starting material, YK-CAP-103-PM14 (triethylamine salt, 250 mg, 0.45 mmol, 62.9%) was obtained via the synthetic route of YK-CAP-101-PM5. 11 H 17 N5O 11 P2, MS(ES): m / z(MH - )456.0.

[0176] Step 15: Synthesis of YK-CAP-103-PM15 Using YK-CAP-103-PM14 (250 mg, 0.45 mmol) as a starting material, YK-CAP-103-PM15 (triethylamine salt, 90 mg, 0.16 mmol, 34.9%) was obtained via the synthetic route of YK-CAP-101-PM6. 12 H 20 N5O 11 P2, MS(ES): m / z(MH - )470.1.

[0177] Step 16: Synthesis of YK-CAP-103 Using YK-CAP-103-PM15 (90 mg, 0.16 mmol) as a raw material, YK-CAP-103 (29 mg, 23.95 μmol, 15.0%) was obtained via the synthetic route of YK-CAP-101. 33 H 45 N 15 O 24 P4, MS(ES): m / z(MH - )1158.2. 1 HNMR(400MHz,D2O)δ8.41(d,J=1.2Hz,1H),8.33(d,J=2.3Hz,1H),8.02(d,J=4.8Hz, 2H),6.11(d,J=5.4Hz,1H),5.91(d,J=2.4Hz,1H),5.71(d,J=4.6Hz,1H),5.20-5.03 (m,2H),4.91-4.77(m,3H),4.63(t,J=4.7Hz,1H),4.53-4.35(m,4H),4.26(d,J=3.1 Hz,1H),4.10(s,3H),4.02(s,2H),3.92(s,3H),3.54(t,J=7.0Hz,1H),3.32(s,3H). 31 PNMR(D2O,162MHz)δ-0.91(s,1P),-11.22(d,J=19.4Hz,1P),-11.33(d,J=17.2Hz,1P),-24.43(t,J=17.7Hz,1P).

[0178] 6. Synthesis of YK-CAP-104

[0179] [ka]

[0180] Step 1: Synthesis of YK-CAP-104-PM1 1,2-O-Isopropylidene-α-D-ribofuranose (20.0 g, 0.11 mol) was dissolved in dichloromethane, and imidazole (11.6 g, 0.17 mol) and TBDPSCl (33.0 g, 0.12 mol) were added to the reaction mixture. The mixture was stirred at room temperature for 15 hours. The reaction solution was added to saturated sodium bicarbonate solution, extracted with dichloromethane, and the dichloromethane phase was washed once with saturated brine, dried over anhydrous sodium sulfate, filtered, and the filtrate was spin-dried under reduced pressure. The residue was purified by silica gel column chromatography (0-17% ethyl acetate / n-hexane) to give YK-CAP-104-PM1 (33.5 g, 78.16 mmol, 71.1%).

[0181] Step 2: Synthesis of YK-CAP-104-PM2 YK-CAP-104-PM1 (33.5 g, 78.16 mmol) was dissolved in acetonitrile, 2-iodoxybenzoic acid (28.5 g, 101.8 mmol) was added, and the mixture was heated to 90°C and stirred for 5 hours to react. The reaction solution was filtered, and the filtrate was spin-dried under reduced pressure to obtain YK-CAP-104-PM2 (32.7 g, 76.66 mmol, 98.1%).

[0182] Step 3: Synthesis of YK-CAP-104-PM3 (Bromomethyl)triphenylphosphonium bromide (60.3 g, 138.3 mmol) was dissolved in tetrahydrofuran (400 mL) and cooled to -78 °C. A 2.5 M solution of n-butyllithium in tetrahydrofuran (76 mL, 190 mmol) was slowly added dropwise. After the addition was complete, the reaction mixture was allowed to cool to 0 °C and stirred for 2 hours. The reaction mixture was again cooled to -78 °C, and a solution of YK-CAP-104-PM2 (48.0 g, 112.52 mmol) in tetrahydrofuran (100 mL) was slowly added dropwise. After the addition was complete, the reaction mixture was allowed to warm to room temperature and stirred overnight. The reaction mixture was quenched with saturated ammonium chloride solution (200 mL) and extracted with ethyl acetate (300 mL x 3). The combined organic phase was washed with saturated aqueous NaCl, dried over anhydrous sodium sulfate, filtered, and spin-dried. The residue was purified by silica gel column chromatography (0-20% ethyl acetate / n-hexane) to give YK-CAP-104-PM3 (39.0 g, 91.85 mmol, 81.6%).

[0183] Step 4: Synthesis of YK-CAP-104-PM4 YK-CAP-104-PM3 (34.0 g, 80.07 mmol) was dissolved in tetrahydrofuran (200 mL), tetra-n-butylammonium fluoride (52.0 g, 198.9 mmol) was added, and the mixture was stirred at room temperature for 1 hour. Saturated aqueous ammonium chloride was added to the reaction mixture, followed by extraction with ethyl acetate (200 mL x 3). The combined organic phase was washed with saturated aqueous NaCl, dried over anhydrous sodium sulfate, filtered, and spin-dried. The residue was purified by silica gel column chromatography (0-60% ethyl acetate / n-hexane) to yield YK-CAP-104-PM4 (13.5 g, 72.50 mmol, 90.5%).

[0184] Step 5: Synthesis of YK-CAP-104-PM5 YK-CAP-104-PM4 (13.5 g, 72.50 mmol) was dissolved in dichloromethane (150 mL), triethylamine (22.0 g, 217.4 mmol) was added, and the reaction mixture was cooled to 0 °C. Benzoyl chloride (11.2 g, 79.7 mmol) was slowly added dropwise, allowed to warm to room temperature, and then stirred for 1 hour. After completion of the reaction, the reaction mixture was quenched by adding saturated aqueous sodium bicarbonate (100 mL) and extracted with dichloromethane (100 mL x 3). The combined organic phases were washed with saturated aqueous NaCl, dried over anhydrous sodium sulfate, filtered, and spin-dried. The residue was purified by silica gel column chromatography (0-20% ethyl acetate / n-hexane) to give YK-CAP-104-PM5 (19.6 g, 67.51 mmol, 93.1%).

[0185] Step 6: Synthesis of YK-CAP-104-PM6 YK-CAP-104-PM5 (18.6 g, 64.07 mmol) was dissolved in a mixture of tetrahydrofuran (160 mL) and water (40 mL). N-methylmorpholine N-oxide (11.3 g, 96.5 mmol) and potassium osmate(VI) dihydrate (2.0 g, 6.4 mmol) were added sequentially and stirred at room temperature overnight. After completion of the reaction, the mixture was quenched with saturated aqueous sodium sulfite and extracted with ethyl acetate (200 mL x 3). The combined organic phases were washed with saturated aqueous sodium chloride, dried over anhydrous sodium sulfate, filtered, and spin-dried. The residue was purified by silica gel column chromatography (0-60% ethyl acetate / n-hexane) to give YK-CAP-104-PM6 (20.5 g, 63.21 mmol, 98.7%).

[0186] Step 7: Synthesis of YK-CAP-104-PM7 YK-CAP-104-PM6 (17.7 g, 54.57 mmol) was dissolved in acetonitrile (140 mL) and pyridine (140 mL). Imidazole (11.1 g, 163.8 mmol), triphenylphosphine (21.5 g, 81.9 mmol), and carbon tetrabromide (27.1 g, 81.9 mmol) were then weighed out. The reaction mixture was heated to 70 °C under a nitrogen atmosphere and stirred for 6 h. Complete reaction was monitored by thin-layer chromatography and spin-dried in vacuo. The residue was purified by silica gel column chromatography (0-50% ethyl acetate / n-hexane) to give YK-CAP-104-PM7 (13.9 g, 35.90 mmol, 65.8%).

[0187] Step 8: Synthesis of YK-CAP-104-PM8 YK-CAP-104-PM7 (13.4 g, 34.60 mmol) was dissolved in acetonitrile (120 mL), followed by the addition of potassium carbonate (14.4 g, 104.2 mmol) and dimethylamine hydrochloride (3.4 g, 41.1 mmol). The reaction mixture was heated to 70°C and reacted for 16 hours. Complete reaction was monitored by thin-layer chromatography. After cooling to room temperature, the organic phase was spin-dried under reduced pressure, and the residue was purified by silica gel column chromatography (0-50% ethyl acetate / n-hexane) to yield YK-CAP-104-PM8 (8.5 g, 24.18 mmol, 69.9%). 18 H 25 NO6, MS(ES): m / z (M+H + )352.1.

[0188] Step 9: Synthesis of YK-CAP-104-PM9 YK-CAP-104-PM8 (8.5 g, 24.18 mmol) was dissolved in dichloromethane (160 mL) and cooled to -40 °C. A solution of diethylaminosulfur trifluoride (4.7 g, 29.0 mmol) in dichloromethane (10 mL) was slowly added dropwise. After the addition was complete, the mixture was slowly warmed to 0 °C and stirred for 4 hours. The reaction was monitored for completeness by thin-layer chromatography. The reaction was quenched with saturated aqueous sodium bicarbonate (100 mL) and extracted with dichloromethane (100 mL x 3). The combined organic phases were washed with saturated aqueous NaCl, dried over anhydrous sodium sulfate, filtered, and spin-dried. The residue was purified by silica gel column chromatography (0-40% ethyl acetate / n-hexane) to give YK-CAP-104-PM9 (5.2 g, 14.71 mmol, 60.8%). 18 H 24 FNO5, MS(ES): m / z (M+H + )354.2.

[0189] Step 10: Synthesis of YK-CAP-104-PM10 YK-CAP-104-PM9 (4.7 g, 13.30 mmol) was dissolved in acetic acid (100 mL), acetic anhydride (16.4 g, 160.6 mmol) was added, and concentrated sulfuric acid (4.7 g, 26.8 mmol) was slowly added dropwise. After the addition was complete, the mixture was slowly heated to 40 °C and stirred for 16 h. The reaction was monitored for completeness by LCMS. The reaction was quenched with saturated aqueous sodium bicarbonate, the pH was adjusted to neutral, and the mixture was extracted with ethyl acetate (200 mL x 3). The combined organic phases were washed with saturated aqueous NaCl, dried over anhydrous sodium sulfate, filtered, and spin-dried. The residue was purified by silica gel column chromatography (0-40% ethyl acetate / n-hexane) to give YK-CAP-104-PM10 (2.5 g, 6.29 mmol, 47.3%). 19 H 24 FNO7, MS(ES): m / z (M+H + )398.1.

[0190] Step 11: Synthesis of YK-CAP-104-PM11 Intermediate INT-I (2.7 g, 6.9 mmol) was dissolved in 1,2-dichloroethane (50 mL), N,O-bis(trimethylsilyl)acetamide (2.8 g, 13.8 mmol) was added, and the reaction mixture was heated to 80 °C and stirred for 2 h. The mixture was then spin-dried under reduced pressure. The residue was dissolved in toluene (30 mL), and a solution of YK-CAP-104-PM10 (2.5 g, 6.29 mmol) in toluene (20 mL) was added. Trimethylsilyl trifluoromethanesulfonate (1.5 g, 6.7 mmol) was slowly added dropwise. The reaction mixture was heated to 70 °C and stirred for 2 h. The reaction was monitored for completeness by thin-layer chromatography. The reaction was quenched with saturated aqueous sodium bicarbonate and extracted with ethyl acetate (50 mL x 3). The combined organic phases were washed with saturated aqueous NaCl, dried over anhydrous sodium sulfate, filtered, and spun under reduced pressure. The residue was purified by silica gel column chromatography (0-70% ethyl acetate / dichloromethane) to give YK-CAP-104-PM11 (2.7 g, 3.72 mmol, 59.1%). 37 H 36 FN7O8, MS(ES): m / z (M+H + )726.2.

[0191] Step 12: Synthesis of YK-CAP-104-PM12 YK-CAP-104-PM11 (2.7 g, 3.72 mmol) was dissolved in a mixed solvent of NH3 / MeOH (20 mL) and water (4 mL), and the reaction mixture was heated to 50 °C and stirred for 10 hours. The reaction was monitored by LCMS for completeness, and the mixture was spin-dried. 2.5 g of the crude product was purified by high-performance liquid chromatography to obtain YK-CAP-104-PM12 (560 mg, 1.64 mmol, 44.0%). 13 H 19 FN6O4, MS(ES): m / z (M+H + )343.1. YK-CAP-104-PM12: 1HNMR(400MHz,DMSO-d6)δ9.98(s,1H),7.93(s,1H),6.52(s,2H),5.69(d,J=6.1Hz,1H),5.51(s,1H),4.97-4.8 5(m,1H),4.37(d,J=6.4Hz,1H),4.14(d,J=4.4Hz,1H),3.77(d,J=12.1Hz,1H),3.62(s,1H),2.59-2.33(m,8H).

[0192] Step 13: Synthesis of YK-CAP-104-PM13 Using YK-CAP-104-PM12 (560 mg, 1.64 mmol) as a raw material, YK-CAP-104-PM13 (triethylamine salt, 557 mg, 1.06 mmol, 64.9%) was obtained via the synthetic route of YK-CAP-101-PM3. 13 H 20 FN6O7P, MS(ES): m / z(MH - )421.1.

[0193] Step 14: Synthesis of YK-CAP-104-PM14 Using YK-CAP-104-PM13 (557 mg, 1.06 mmol) as a raw material, YK-CAP-104-PM14 (sodium salt, 414 mg, 0.84 mmol, 79.0%) was obtained via the synthetic route of YK-CAP-101-PM4. 16 H 23 FN8O6P, MS(ES): m / z(MH - )471.1.

[0194] Step 15: Synthesis of YK-CAP-104-PM15 Using YK-CAP-104-PM14 (414 mg, 0.84 mmol) as a starting material, YK-CAP-104-PM15 (triethylamine salt, 300 mg, 0.50 mmol, 59.2%) was obtained via the synthetic route of YK-CAP-101-PM5. 13 H 21 FN6O 10 P2, MS(ES): m / z(MH - )501.1.

[0195] Step 16: Synthesis of YK-CAP-104-PM16 Using YK-CAP-104-PM15 (300 mg, 0.50 mmol) as a starting material, YK-CAP-104-PM16 (triethylamine salt, 124 mg, 0.20 mmol, 40.2%) was obtained via the synthetic route of YK-CAP-101-PM6. 14 H 24 FN6O 10 P2, MS(ES): m / z(MH - )515.1.

[0196] Step 17: Synthesis of YK-CAP-104 Using YK-CAP-104-PM16 (124 mg, 0.20 mmol) as a starting material, YK-CAP-104 (ammonium salt, 21 mg, 16.72 μmol, 8.4%) was obtained via the synthetic route of YK-CAP-101. 35 H 49 FN 16 O 23 P4, MS(ES): m / z(MH - )1203.1. 1 HNMR(400MHz,D2O)δ8.52(d,J=1.2Hz,1H),8.44(d,J=2.2Hz,1H),7.86(d,J=4.4Hz, 2H),6.14(d,J=5.2Hz,1H),5.94(d,J=2.4Hz,1H),5.81(d,J=4.6Hz,1H),4.81-4.67 (m,3H),4.63(t,J=4.6Hz,1H),4.53-4.35(m,5H),4.23(d,J=3.1Hz,1H),4.19(s,3H) ),4.12(s,2H),3.78(s,3H),3.32(t,J=7.0Hz,1H),3.12(s,3H),2.55-2.35(m,8H). 31 PNMR(D2O,162MHz)δ-0.92(s,1P),-11.32(d,J=20.7Hz,1P),-12.33(d,J=17.5Hz,1P),-23.28(t,J=16.8Hz,1P).

[0197] 7. Synthesis of YK-CAP-105

[0198] [ka]

[0199] Step 1: Synthesis of YK-CAP-105-PM1 YK-CAP-104-PM4 (5.76 g, 30.93 mmol) was dissolved in acetic acid (8 mL). To the mixture was added acetic anhydride (31.6 g, 310 mmol) and sulfuric acid (500 μL), and the mixture was stirred at room temperature for 4 hours. The pH of the reaction mixture was adjusted to a weak acidic state with aqueous NaHCO3 solution, and the mixture was extracted twice with ethyl acetate. The organic phase was spin-dried, and the residue was purified by silica gel column chromatography (0-80% ethyl acetate / n-hexane) to yield YK-CAP-105-PM1 (5.9 g, 21.67 mmol, 70.1%).

[0200] Step 2: Synthesis of YK-CAP-105-PM2 Intermediate INT-I (9.27 g, 23.87 mmol) and N,O-bis(trimethylsilyl)acetamide (9.72 g, 47.8 mmol) were dissolved in 1,2-dichloroethane (60 mL) and stirred at 80 °C for 2 h. The solvent was then spin-dried. YK-CAP-105-PM1 (5.9 g, 21.67 mmol) was dissolved in toluene (80 mL) and added to the residue. Trimethylsilyl trifluoromethanesulfonate (5.31 g, 23.9 mmol) was added and the mixture was stirred at 70 °C for 2 h. Completion of the reaction was monitored by thin-layer chromatography. After filtration, the filtrate was spin-dried under reduced pressure. The residue was purified by silica gel column chromatography (0-80% ethyl acetate / n-hexane) to give YK-CAP-105-PM2 (10.00 g, 16.65 mmol, 76.8%).

[0201] Step 3: Synthesis of YK-CAP-105-PM3 YK-CAP-105-PM2 (8.00 g, 13.32 mmol) and [N,N'-(1,1,2,2-Tetramethylethylene)bis(3,5-di-tert-butylsalicylideneiminato)]cobalt(II) (403 mg, 0.67 mmol) were dissolved in 1,4-dioxane (30 mL), and the resulting mixture was added to the above solution. Benzenesulfonyl cyanide (72.30 g, 400 mmol) was added and stirred at room temperature for 5 minutes. Phenylsilane (1.73 g, 16 mmol) was dissolved in absolute ethanol (60 mL) and added to the above mixture, followed by stirring at room temperature for 3 hours. After the reaction was completed, the mixture was filtered, extracted twice with water and ethyl acetate, the organic phase was spin-dried, and the residue was purified by silica gel column chromatography (0-70% ethyl acetate / n-hexane) to obtain YK-CAP-105-PM3 (4.50 g, 7.17 mmol, 53.8%).

[0202] Step 4: Synthesis of YK-CAP-105-PM4 YK-CAP-105-PM3 (4.50 g, 7.17 mmol) was dissolved in 7 M ammonia-methanol (50 mL) and stirred at 50 °C for 4 hours. After the reaction was complete, the solvent was removed by spin-drying under reduced pressure, and the residue was purified by high-performance liquid chromatography to obtain YK-CAP-105-PM4 (800 mg, 2.61 mmol, 36.4%). 12 H 14 NO, MS (ES): m / z (M+H + )307.11. YK-CAP-105-PM4: 1 HNMR(400MHz,DMSO-d6)δ10.66(s,1H),7.97(s,1H),6.54(d,J=5.2Hz,3H),5.74(d,J=7.2Hz,1H) ,5.40(t,J=4.4Hz,1H),4.62-4.65(m,1H),4.31(t,J=3.2Hz,1H),3.59-3.69(m,2H),1.53(s,3H).

[0203] Step 5: Synthesis of YK-CAP-105-PM5 Using YK-CAP-105-PM4 (800 mg, 2.61 mmol) as a raw material, YK-CAP-105-PM5 (triethylamine salt, 994 mg, 2.04 mmol, 78.1%) was obtained via the synthetic route of YK-CAP-101-PM3. 12 H 15 N6O7P, MS(ES): m / z(MH - )385.1.

[0204] Step 6: Synthesis of YK-CAP-105-PM6 Using YK-CAP-105-PM5 (994 mg, 2.04 mmol) as a raw material, YK-CAP-105-PM6 (sodium salt, 661 mg, 1.44 mmol, 70.7%) was obtained via the synthetic route of YK-CAP-101-PM4. 15 H 17 N8O6P, MS(ES): m / z(MH - )435.1.

[0205] Step 7: Synthesis of YK-CAP-105-PM7 Using YK-CAP-105-PM6 (661 mg, 1.44 mmol) as a starting material, YK-CAP-105-PM7 (triethylamine salt, 612 mg, 1.08 mmol, 74.8%) was obtained via the synthetic route of YK-CAP-101-PM5. 12 H 16 NO 10 P2, MS(ES): m / z(MH - )465.0.

[0206] Step 8: Synthesis of YK-CAP-105-PM8 Using YK-CAP-105-PM7 (612 mg, 1.08 mmol) as a starting material, YK-CAP-105-PM8 (triethylamine salt, 325 mg, 0.56 mmol, 51.7%) was obtained via the synthetic route of YK-CAP-101-PM6. 13 H 19 NO 10 P2, MS(ES): m / z(MH - )479.1.

[0207] Step 9: Synthesis of YK-CAP-105 Using YK-CAP-105-PM8 (325 mg, 0.56 mmol) as a starting material, YK-CAP-105 (16 mg, 13.12 μmol, 2.3%) was obtained via the synthetic route of YK-CAP-101. 34 H 44 N 16 O 23 P4, MS(ES): m / z(MH - )1267.1. 1 HNMR(400MHz,D2O)δ8.49(d,J=1.4Hz,1H),8.44(d,J=2.5Hz,1H),7.88(d,J= 4.2Hz,2H),6.22(d,J=4.7Hz,1H),5.93(d,J=2.4Hz,1H),5.80(d,J=4.5Hz,1 H),4.81-4.66(m,3H),4.62-4.32(m,5H),4.22(d,J=3.1Hz,1H),4.18(s,2H) ,4.10(s,2H),3.77(s,3H),3.31(t,J=6.8Hz,1H),3.11(s,3H),1.53(s,3H). 31 PNMR(D2O,162MHz)δ-0.90(s,1P),-11.33(d,J=20.2Hz,1P),-12.12(d,J=17.1Hz,1P),-22.34(t,J=16.2Hz,1P).

[0208] 8. Synthesis of YK-CAP-106

[0209] [ka]

[0210] Step 1: Synthesis of YK-CAP-106-PM1 To YK-CAP-105-PM2 (23.00 g, 38.30 mmol), p-toluenesulfonyl azide (211.46 g, 1.07 mol) and cobalt catalyst ([N,N'-(1,1,2,2-tetramethylethylene)bis(3,5-di-tert-butylsalicylideneiminato)]cobalt(II)) (696 mg, 1.15 mmol) were added and stirred under nitrogen gas protection for 30 minutes. Phenylsilane (10.22 g, 45.96 mmol) dissolved in absolute ethanol (40 mL) was added dropwise at room temperature. The addition was completed within 30 minutes, and the reaction was allowed to proceed with stirring at room temperature for 2 hours. The reaction was monitored for completion by LC-MS, quenched, and extracted with EA and saturated aqueous sodium chloride solution. The reaction mixture was separated, the aqueous phase was back-extracted twice with EA, the organic phases were combined, the solvent was spun off, and the crude product was purified by flash column chromatography (EA / PE = 0-100%) to give YK-CAP-106-PM1 (4.90 g, 7.61 mmol, 19.9%). 30 H 29 NO, MS (ES): m / z (M+H + )644.3.

[0211] Step 2: Synthesis of YK-CAP-106-PM2 Triphenylphosphine (2.78 g, 10.59 mmol), water (4.9 mL), and tetrahydrofuran (49 mL) were added to YK-CAP-106-PM1 (4.90 g, 7.61 mmol), and the mixture was heated to 50°C and stirred for 8 hours. The reaction was monitored for completion by LC-MS, and the reaction was stopped. The solvent was removed by spinning off the reaction solution under reduced pressure. The crude product was purified by flash column chromatography (MeOH / DCM = 0-10%) to give YK-CAP-106-PM2 (4.22 g, 6.83 mmol, 89.8%). 30 H 31 N7O 8, MS(ES): m / z (M+H + )618.2.

[0212] Step 3: Synthesis of YK-CAP-106-PM3 Triethylamine (5.04 g, 49.84 mmol) and dichloromethane (82 mL) were added to YK-CAP-106-PM2 (4.10 g, 6.64 mmol), cooled to 0°C, and acetyl chloride (1.68 g, 21.36 mmol) was added dropwise. After the addition was complete, the mixture was warmed to room temperature and allowed to react for 24 hours. The reaction was monitored for completion by LC-MS, and the reaction was stopped. The reaction solution was poured into ice water, stirred, and separated. The aqueous phase was back-extracted twice with dichloromethane. The organic phases were combined, and the solvent was spun off under reduced pressure. The crude product was purified by flash column chromatography (MeOH / DCM = 0-10%) to give YK-CAP-106-PM3 (2.94 g, 4.46 mmol, 67.1%). 32 H 33 N7O9, MS(ES): m / z (M+H + )660.3.

[0213] Step 4: Synthesis of YK-CAP-106-PM4 YK-CAP-106-PM3 (2.94 g, 4.46 mmol) was added to 7 M ammonia-methanol solution (29 mL) and water (5.9 mL) and stirred at room temperature for 24 hours. Completion of the reaction was monitored by LC-MS, and the reaction was stopped. The solvent was spun off under reduced pressure. EA was added to the crude product, stirred, and filtered under suction to obtain a yellow solid, YK-CAP-106-PM4 (1.36 g, 4.02 mmol, 90.1%). 13 H 18 N6O5, MS(ES): m / z (M+H + )339.3. YK-CAP-106-PM4: 1 HNMR (400MHz, MeOD) δ7.99(s,1H),5.84(d,1H),4.77(s,1H),4.66(d,1H),3.93-3.82(m,2H),2.06(s,3H),1.63(s,3H).

[0214] Step 5: Synthesis of YK-CAP-106-PM5 Using YK-CAP-106-PM4 (500 mg, 1.48 mmol) as a starting material, YK-CAP-106-PM5 (triethylamine salt, 480 mg, 0.92 mmol, 62.4%) was obtained via the synthetic route of YK-CAP-101-PM3. 13 H 19 N6O8P, MS(ES): m / z(MH - )417.2.

[0215] Step 6: Synthesis of YK-CAP-106-PM6 Using YK-CAP-106-PM5 (480 mg, 0.92 mmol) as a raw material, YK-CAP-106-PM6 (sodium salt, 410 mg, 0.84 mmol, 90.9%) was obtained via the synthetic route of YK-CAP-101-PM4. 16 H 21 N8O7P, MS(ES): m / z(MH - )467.1.

[0216] Step 7: Synthesis of YK-CAP-106-PM7 Using YK-CAP-106-PM6 (410 mg, 0.84 mmol) as a starting material, YK-CAP-106-PM7 (triethylamine salt, 331 mg, 0.55 mmol, 65.7%) was obtained via the synthetic route of YK-CAP-101-PM5. 13 H 20 NO 11 P2, MS(ES): m / z(MH - )497.0.

[0217] Step 8: Synthesis of YK-CAP-106-PM8 Using YK-CAP-106-PM7 (331 mg, 0.55 mmol) as a starting material, YK-CAP-106-PM8 (triethylamine salt, 150 mg, 0.24 mmol, 44.5%) was obtained via the synthetic route of YK-CAP-101-PM6. 14 H 22 NO 11 P2, MS(ES): m / z(MH - )511.1.

[0218] Step 9: Synthesis of YK-CAP-106 Using YK-CAP-106-PM8 (150 mg, 0.24 mmol) as a starting material, YK-CAP-106 (32 mg, 25.56 μmol, 10.6%) was obtained via the synthetic route of YK-CAP-101. 35 H 48 N 16 O 24 P4, MS(ES): m / z(MH - )1199.1. 1 HNMR(400MHz,D2O)δ8.50(d,J=1.4Hz,1H),8.44(d,J=2.2Hz,1H),7.78(d,J=4.2H z,2H),6.24(d,J=4.6Hz,1H),5.94(d,J=2.3Hz,1H),5.81(d,J=4.4Hz,1H),4.86- 4.76(m,3H),4.61-4.55(m,4H),4.52(d,J=3.6Hz,2H),4.28(s,2H),4.20(s,1H), 3.93-3.82(m,5H),3.23(t,J=6.7Hz,1H),3.12(s,2H),2.03(s,3H),1.62(s,3H). 31 PNMR(D2O,162MHz)δ-0.92(s,1P),-11.33(d,J=20.1Hz,1P),-12.13(d,J=16.8Hz,1P),-22.11(t,J=16.4Hz,1P).

[0219] 9. Synthesis of YK-CAP-107

[0220] [ka]

[0221] Step 1: Synthesis of YK-CAP-107-PM1 A 1 M solution of borane in tetrahydrofuran (325 mL, 0.325 mol) was added to a three-neck flask, and a solution of YK-CAP-104-PM3 (60.00 g, 0.14 mol) in tetrahydrofuran (180 mL) was slowly added dropwise at 0 °C. After the addition was completed, the mixture was stirred at room temperature for 2 hours to allow the reaction to proceed. Then, THF / HO (1:1, 120 mL), 2 N NaOH (261 mL), and 30% hydrogen peroxide (271 mL) were slowly added dropwise in that order at 0 °C. After the dropwise addition was completed, the reaction was stirred overnight at room temperature. Completion of the reaction was monitored by TLC. The reaction was then quenched and extracted with water. The organic phase was washed successively with saturated aqueous sodium thiosulfate and saturated brine. The organic phase was dried over anhydrous sodium sulfate, filtered, and spun off to remove the solvent. The crude product was purified by flash column chromatography (PE:EA = 0-25%) to obtain the colorless oily compound YK-CAP-107-PM1 (48.80 g, 0.11 mol, 78.8%).

[0222] Step 2: Synthesis of YK-CAP-107-PM2 YK-CAP-107-PM1 (34.20 g, 77.27 mmol) was dissolved in acetonitrile, and 2-iodoxybenzoic acid (28.10 g, 100.5 mmol) was added to the reaction mixture. The mixture was heated to 90°C and stirred for 5 hours. After cooling to room temperature, the reaction solution was filtered, and the filtrate was spin-dried to obtain crude YK-CAP-107-PM2 (34.50 g) as a pale yellow oily liquid. No purification was required and the product was used directly in the next step.

[0223] Step 3: Synthesis of YK-CAP-107-PM3 YK-CAP-107-PM2 (32.90 g, calculated based on 74.67 mmol) was dissolved in tetrahydrofuran and cooled to 0 °C under nitrogen gas protection. A 1 M solution of methylmagnesium bromide in tetrahydrofuran (97.1 mL, 97.1 mmol) was slowly added dropwise. After the addition was complete, the mixture was stirred at room temperature for 3 h. After cooling the reaction solution to 0 °C, the reaction was quenched with saturated ammonium chloride, extracted with ethyl acetate, and the organic phase was spun dry. The crude product was purified by flash column chromatography (PE:EA = 0-41%) to give YK-CAP-107-PM3 (16.80 g, 36.79 mmol, 49.3%) as a pale yellow oil.

[0224] Step 4: Synthesis of YK-CAP-107-PM4 YK-CAP-107-PM3 (16.80 g, 36.79 mmol) was dissolved in THF and cooled to 0 °C under nitrogen gas protection. A THF solution of sodium tert-butoxide (11.20 g, 116.7 mmol) was slowly added to the reaction mixture. After stirring for 1.5 hours at room temperature, methyl iodide (27.60 g, 194.5 mmol) was slowly added dropwise to the reaction mixture. After the addition was complete, the reaction mixture was stirred for 3 hours. The reaction was monitored for completion by TLC and quenched with saturated ammonium chloride solution. The mixture was extracted with ethyl acetate, and the organic phase was washed 3-5 times with saturated brine, dried, and spin-dried. The crude product was purified by flash column chromatography (PE:EA = 0-12%) to obtain a pale yellow oily liquid YK-CAP-107-PM4 (13.16 g, 27.96 mmol, 76.0%).

[0225] Step 5: Synthesis of YK-CAP-107-PM5 YK-CAP-107-PM4 (13.16 g, 27.96 mmol) was dissolved in glacial acetic acid (130 mL), and acetic anhydride (17.13 g, 167.8 mmol) and concentrated sulfuric acid (0.52 mL) were sequentially added to the reaction system. Under the condition of room temperature, the mixture was stirred and reacted for 4 hours. The reaction system was diluted with ethyl acetate, washed once with water, and further washed three times with saturated sodium bicarbonate solution. The organic phase was dried and spin-dried to obtain a yellow oily liquid YK-CAP-107-PM5 (11.38 g), which was directly used in the next step.

[0226] Step 6: Synthesis of YK-CAP-107-PM6 2-Acetamido-9H-purin-6-yl diphenylcarbamate (6.30 g, 16.22 mmol) was dissolved in 1,2-dichloroethane (100 mL), N,O-bis(trimethylsilyl)acetamide (6.60 g, 32.4 mmol) was added, and the temperature was raised to 80 °C. After stirring and reacting for 1.5 hours, it was spin-dried to remove the solvent. Under the condition of room temperature, a toluene solution (100 mL) of YK-CAP-107-PM5 (calculated based on 6.00 g, 18.85 mmol) and TMSOTf (3.6 g, 16.2 mmol) were sequentially added. Then the temperature was raised to 70 °C, and the mixture was continuously stirred and reacted for 3.5 hours. The reaction system was diluted with ethyl acetate, washed once with saturated sodium bicarbonate solution, filtered to remove insoluble substances, the filtrate was separated, the organic phase was dried and spin-dried, and the crude product was purified by flash column chromatography (DCM:EA = 0~52%) to obtain a yellow solid YK-CAP-107-PM6 (3.20 g, 4.95 mmol, 26.3%). C 32 H 34 N6O9, MS(ES): m / z(M+H + ) 647.2. <s

[0227] Step 7: Synthesis of YK-CAP-107-PM7 YK-CAP-107-PM6 (3.20 g, 4.95 mmol) was dissolved in a 7 M ammonium chloride / methanol solution and water (5:1, 24 mL), heated to 50 °C, and stirred for 6 hours. The reaction solution was then spun dry, slurried twice with EA, and suction filtered to collect the cake, yielding a white solid, YK-CAP-107-PM7 (1.38 g, 4.24 mmol, 85.7%). 13 H 19 N5O5, MS(ES): m / z (M+H + )326.2. YK-CAP-107-PM7: 1 HNMR(400MHz,MeOD)δ8.07(s,1H),5.82(d,J=2.6Hz,1H),4.53(dd,J=6.1,2.7Hz,1H),4.36(ddd,J=8.3,3.9,2.6H z,1H),3.95(dd,J=12.1,2.4Hz,1H),3.78-3.67(m,2H),3.34(s,3H),2.11(q,J=7.4Hz,1H),1.23(d,J=6.1Hz,3H).

[0228] Step 8: Synthesis of YK-CAP-107-PM8 Using YK-CAP-107-PM7 (1.20 g, 3.69 mmol) as a starting material, YK-CAP-107-PM8 (triethylamine salt, 644 mg, 1.27 mmol, 34.4%) was obtained via the synthetic route of YK-CAP-101-PM3. 13 H 20 N5O8P, MS(ES): m / z(MH - )404.1.

[0229] Step 9: Synthesis of YK-CAP-107-PM9 Using YK-CAP-107-PM8 (644 mg, 1.27 mmol) as a starting material, YK-CAP-107-PM9 (sodium salt, 549 mg, 1.15 mmol, 90.6%) was obtained via the synthetic route of YK-CAP-101-PM4. 16 H 22 N7O7P, MS(ES): m / z(MH - )454.2.

[0230] Step 10: Synthesis of YK-CAP-107-PM10 Using YK-CAP-107-PM9 (549 mg, 1.15 mmol) as a starting material, YK-CAP-107-PM10 (triethylamine salt, 531 mg, 0.91 mmol, 78.7%) was obtained via the synthetic route of YK-CAP-101-PM5. 13 H 21 N5O 11 P2, MS(ES): m / z(MH - )484.0.

[0231] Step 11: Synthesis of YK-CAP-107-PM11 Using YK-CAP-107-PM10 (531 mg, 0.91 mmol) as a raw material, YK-CAP-107-PM11 (triethylamine salt, 163 mg, 0.27 mmol, 29.8%) was obtained via the synthesis route of YK-CAP-101-PM6. 14 H 23 N5O 11 P2, MS(ES): m / z(MH - )498.1.

[0232] Step 12: Synthesis of YK-CAP-107 Using YK-CAP-107-PM11 (163 mg, 0.27 mmol) as a starting material, YK-CAP-107 (35 mg, 28.25 μmol, 10.5%) was obtained via the synthetic route of YK-CAP-101. 35 H 49 N 15 O 24 P4, MS(ES): m / z(MH - )1186.1. 1HNMR(400MHz,D2O)δ8.43(d,J=1.2Hz,1H),8.34(d,J=2.1Hz,1H),7.73(d,J=4.1Hz,2H),6.21(d,J =4.5Hz,1H),5.74(d,J=2.3Hz,1H),5.71(d,J=4.4Hz,1H),4.76-4.61(m,3H),4.51-4.45(m,4H),4. 42(d,J=3.5Hz,2H),4.31(s,2H),4.10(s,2H),3.95-3.92(m,1H),3.90-3.84(m,2H),3.78-3.67(m, 2H),3.13(t,J=6.7Hz,1H),3.34(s,3H),3.10(s,2H),2.07(q,J=7.4Hz,1H),1.22(d,J=6.2Hz,3H). 31 PNMR(D2O,161MHz)δ-0.88(s,1P),-11.23(d,J=20.2Hz,1P),-12.22(d,J=16.6Hz,1P),-22.32(t,J=16.1Hz,1P).

[0233] 10.Synthesis of YK-CAP-108

[0234]

change

[0235] Synthesis of ステップ1:YK-CAP-108-PM1 YK-CAP-107-PM3 (17.5 g, 38.32 mmol) was dissolved in 300 mL of THF, and triphenylphosphine (12.1 g, 46.00 mmol) and phthalimide (6.76 g, 46.00 mmol) were added. Under nitrogen gas protection, the reaction system was cooled to 0 °C, and a THF solution (30 mL) of DEAD (9.3 g, 53.40 mmol) was added dropwise. After the addition was complete, the reaction was continued for 3 hours, and the reaction was monitored for completeness by TLC (DCM). After the reaction was complete, 30 mL of purified water was added dropwise to the reaction system at 0 °C to quench the reaction. After extraction with EA (200 mL × 3), the organic phase was washed successively with saturated brine (400 mL), dried over anhydrous sodium sulfate, and spin-dried. The residue was purified by normal-phase silica gel column chromatography (PE:EA = 0-40%) to give a yellow oil, YK-CAP-108-PM1 (18.8 g, 32.09 mmol, 83.8%).

[0236] Step 2: Synthesis of YK-CAP-108-PM2 YK-CAP-108-PM1 (18.8 g, 32.09 mmol) was dissolved in 200 mL of ethanol, and 85% hydrazine hydrate (37.8 g, 0.64 mol) was added. The reaction mixture was heated to 80 °C and reacted for 1 h. Complete reaction was monitored by TLC (PE / EA = 3:1). The reaction was stopped, cooled to room temperature, and 200 mL of purified water and EA (200 mL) were added. The mixture was stirred for 10 min and separated. The aqueous phase was extracted with EA (200 mL × 2). The organic phase was washed sequentially with saturated brine (300 mL), dried over anhydrous sodium sulfate, and spin-dried to give YK-CAP-108-PM2 (14.2 g, 31.16 mmol, 97.1%) as a pale yellow oil.

[0237] Step 3: Synthesis of YK-CAP-108-PM3 YK-CAP-108-PM2 (14.2 g, 31.16 mmol) was dissolved in 150 mL of DCM, and DIEA (12.1 g, 93.49 mmol) was added. Protected with nitrogen gas, the reaction mixture was cooled to 0 °C, and a DCM solution (20 mL) of acetyl chloride (2.9 g, 37.40 mmol) was added dropwise. After the addition was complete, the temperature was controlled at 0 °C and the reaction was allowed to proceed for 1 h. The reaction was monitored for completion by TLC (PE / EA = 3 / 1). The reaction was stopped, warmed to room temperature, and 200 mL of saturated sodium bicarbonate solution was added to quench the reaction, followed by separation. The aqueous phase was extracted with DCM (200 mL × 2), and the organic phase was washed successively with saturated brine (300 mL), dried over anhydrous sodium sulfate, and spin-dried. The residue was purified by normal-phase silica gel column chromatography (DCM:MeOH = 0-40%) to give a yellow oil, YK-CAP-108-PM3 (12.5 g, 25.12 mmol, 80.6%).

[0238] Step 4: Synthesis of YK-CAP-108-PM4 YK-CAP-108-PM3 (12.5 g, 25.12 mmol) was dissolved in 60 mL of THF. The reaction mixture was cooled to 0 °C under nitrogen gas protection, and 1 M TBAF (37.7 mL) was added dropwise. After the addition was complete, the mixture was warmed to room temperature and reacted for 2 h. The reaction was monitored for completeness by TLC (PE / EA = 3 / 1). The reaction was quenched, and the reaction solution was directly spun to dryness. The residue was purified by normal-phase silica gel column chromatography (DCM:MeOH = 0-40%) to give a yellow oil, YK-CAP-108-PM4 (5.8 g, 22.37 mmol, 89.0%).

[0239] Step 5: Synthesis of YK-CAP-108-PM5 YK-CAP-108-PM4 (4.5 g, 17.35 mmol) was dissolved in 15 mL of acetic acid, and acetic anhydride (35.4 g, 34.71 mmol) and p-toluenesulfonic acid (1.5 g, 8.67 mmol) were added. The reaction mixture was heated to 50 °C and monitored by LC-MS. After completion of the reaction, the mixture was cooled to room temperature, 100 mL of purified water was added, and the mixture was extracted with 100 mL of EA. The mixture was stirred for 10 min and separated. The aqueous phase was extracted with EA (100 mL x 2). The organic phase was washed sequentially with saturated brine (500 mL), dried over anhydrous sodium sulfate, and spin-dried. The residue was purified by normal-phase silica gel column chromatography (DCM:MeOH = 0-40%) to give YK-CAP-108-PM5 (5.5 g, 15.94 mmol, 91.8%) as a yellow oil.

[0240] Step 6: Synthesis of YK-CAP-108-PM6 Intermediate INT-I (6.8 g, 17.53 mmol) was dissolved in 1,2-dichloroethane (50 mL), N,O-bis(trimethylsilyl)acetamide (7.1 g, 35.06 mmol) was added, and the reaction mixture was heated to 80 °C and stirred for 2 h. The mixture was then spin-dried under reduced pressure. The residue was dissolved in toluene (30 mL), and a solution of YK-CAP-108-PM5 (5.5 g, 15.94 mmol) in toluene (20 mL) was added. Trimethylsilyl trifluoromethanesulfonate (3.9 g, 7.53 mmol) was slowly added dropwise. The reaction mixture was heated to 70 °C and stirred for 2 h. Completion of the reaction was monitored by thin-layer column chromatography. The reaction was quenched with saturated aqueous sodium bicarbonate and extracted with ethyl acetate (100 mL x 3). The combined organic phases were washed with saturated aqueous NaCl (300 mL), dried over anhydrous sodium sulfate, filtered, and spun under reduced pressure. The residue was purified by silica gel column chromatography (DCM:MeOH = 0-100%) to give YK-CAP-108-PM6 (3.8 g, 5.64 mmol, 35.4%). 33 H 35 N7O9, MS(ES): m / z (M+H + )674.3.

[0241] Step 7: Synthesis of YK-CAP-108-PM7 YK-CAP-108-PM6 (3.8 g, 5.64 mmol) was dissolved in a mixture of 4 M NH3 / MeOH (40 mL) and water (4 mL), stirred overnight at room temperature, and the reaction was monitored for completeness by LCMS. The mixture was then spun dry and slurried in EA (100 mL x 2) at room temperature to give an off-white solid, YK-CAP-108-PM7 (1.92 g, 5.45 mmol, 96.6%). 14 H 20 N6O5, MS(ES): m / z (M+H + )353.1. YK-CAP-108-PM7: 1 HNMR(400MHz,MeOD)δ8.02(s,1H),5.71(d,J=2.5Hz,1H),4.43(d,J=2.4Hz,1H),4.31-4.20 (m,1H),3.93-3.82(m,1H),3.78-3.67(m,2H),2.34-2.25(m,1H),2.06(s,3H),1.63(s,3H).

[0242] Step 8: Synthesis of YK-CAP-108-PM8 Using YK-CAP-108-PM7 (1.50 g, 4.26 mmol) as a starting material, YK-CAP-108-PM8 (triethylamine salt, 1.11 g, 2.08 mmol, 48.8%) was obtained via the synthetic route of YK-CAP-101-PM3. 14 H 21 N6O8P, MS(ES): m / z(MH - )431.1.

[0243] Step 9: Synthesis of YK-CAP-108-PM9 Using YK-CAP-108-PM8 (1.11 g, 2.08 mmol) as a starting material, YK-CAP-108-PM9 (sodium salt, 778 mg, 1.54 mmol, 74.2%) was obtained via the synthetic route of YK-CAP-101-PM4. 17 H 23 N8O7P, MS(ES): m / z(MH - )481.1.

[0244] Step 10: Synthesis of YK-CAP-108-PM10 Using YK-CAP-108-PM9 (778 mg, 1.54 mmol) as a starting material, YK-CAP-108-PM10 (triethylamine salt, 512 mg, 0.83 mmol, 54.2%) was obtained via the synthetic route of YK-CAP-101-PM5. 14 H 22 NO 11 P2, MS(ES): m / z(MH - )511.0.

[0245] Step 11: Synthesis of YK-CAP-108-PM11 Using YK-CAP-108-PM10 (512 mg, 0.83 mmol) as a starting material, YK-CAP-108-PM11 (triethylamine salt, 117 mg, 0.19 mmol, 22.5%) was obtained via the synthetic route of YK-CAP-101-PM6. 15 H 24 NO 11 P2, MS(ES): m / z(MH - )525.1.

[0246] Step 12: Synthesis of YK-CAP-108 Using YK-CAP-108-PM11 (117 mg, 0.19 mmol) as a starting material, YK-CAP-108 (18 mg, 14.22 μmol, 7.5%) was obtained via the synthetic route of YK-CAP-101. 36 H 50 N 16 O 24 P4, MS(ES): m / z(MH - )1213.1. 1HNMR(400MHz,D2O)δ8.43-8.32(m,1H),8.12(d,J=2.5Hz,1H),7.73-7.54(m,2H ),6.22(d,J=4.5Hz,1H),5.35(d,J=2.3Hz,1H),5.22-5.11(m,1H),4.72-4.51( m,3H),4.41-4.33(m,6H),4.21(s,2H),4.13(s,2H),3.90-3.84(m,3H),3.13-3 .11(m3H),3.10(s,2H),2.32-2.26(m,1H),2.17(s,3H),1.42(d,J=6.2Hz,3H). 31 PNMR(D2O,163MHz)δ-0.87(s,1P),-11.21(d,J=20.1Hz,1P),-11.62(d,J=16.6Hz,1P),-21.67(t,J=17.2Hz,1P).

[0247] 11. Synthesis of YK-CAP-109

[0248] [ka]

[0249] Step 1: Synthesis of YK-CAP-109-PM1 YK-CAP-107-PM3 (10.0 g, 21.90 mmol) was dissolved in dichloromethane, the reaction mixture was cooled to 0 °C, and DAST (7.1 g, 43.80 mmol) was slowly added dropwise. The reaction mixture was stirred at 0 °C for 4 hours. The reaction mixture was quenched by slowly adding saturated aqueous sodium bicarbonate solution, and DCM was added to separate the mixture. The organic phase was washed twice with saturated aqueous sodium bicarbonate solution and separated. The organic phase was washed two to three times with saturated brine, dried over anhydrous sodium sulfate, filtered, and spin-dried. The residue was purified by silica gel column chromatography (0-30% ethyl acetate / n-hexane) to give YK-CAP-109-PM1 (6.1 g, 13.30 mmol, 60.7%).

[0250] Step 2: Synthesis of YK-CAP-109-PM2 YK-CAP-109-PM1 (6.1 g, 13.30 mmol) was dissolved in acetic acid, and acetic anhydride (27.2 g, 266.0 mmol) was added. Concentrated sulfuric acid (280 μL) was then slowly added dropwise. After the addition was complete, the mixture was stirred at room temperature for 16 hours. 200 mL of water was added to the reaction solution, which was then extracted with ethyl acetate. The organic phase was washed three times with saturated aqueous sodium bicarbonate until the pH became basic. The organic phase was dried and spun to obtain the crude product YK-CAP-109-PM2 (5.80 g), which was a yellow oily liquid and was directly used in the next step.

[0251] Step 3: Synthesis of YK-CAP-109-PM3 Intermediate INT-I (6.2 g, 15.96 mmol) was dissolved in 1,2-dichloroethane, and N,O-bis(trimethylsilyl)acetamide (8.1 g, 39.9 mmol) was added. The mixture was heated to 80°C and stirred for 2 hours to react. The reaction solvent was then removed under reduced pressure, and the mixture was dissolved in toluene. At room temperature, a toluene solution of YK-CAP-109-PM2 (5.8 g, calculated based on 13.30 mmol) and TMSOTf (4.4 g, 19.9 mmol) were added in that order. The mixture was then heated to 70°C and stirred for another 2 hours to react. The reaction mixture was diluted with ethyl acetate, washed once with saturated sodium bicarbonate solution, filtered, the filtrate separated, the organic phase dried over anhydrous sodium sulfate, the solvent was removed under reduced pressure, and the residue was purified by silica gel column chromatography (0-30% ethyl acetate / DCM) to give YK-CAP-109-PM3 (3.1 g, 4.88 mmol). 31 H 31 FN6O8, MS(ES): m / z (M+H + )635.2.

[0252] Step 4: Synthesis of YK-CAP-109-PM4 YK-CAP-109-PM3 (3.1 g, 4.88 mmol) was dissolved in a 7 M ammonium chloride / methanol solution and water (5:1) and stirred at room temperature for 16 hours. The solvent was removed by spin-drying under reduced pressure, and the crude product was recrystallized from ethyl acetate to give YK-CAP-109-PM4 (1.1 g, 3.51 mmol, 71.9%). 12 H 16 FN5O4, MS(ES): m / z (M+H + )314.1. YK-CAP-109-PM4: 1 H NMR (400 MHz, DMSO-d 6 )δ11.45(s,1H),8.00(s,1H),δ6.40(s,2H),5.73-5.56(m,2H),4.83-4.81(m,1H),4.31-4.29(m,1H),4.17 -4.13(m,1H),4.60(s,1H),3.76-3.72(m,1H),3.63-3.50(m,1H),2.34-2.26(m,1H),1.42(d,J=6.2Hz,3H).

[0253] Step 5: Synthesis of YK-CAP-109-PM5 Phosphorus oxychloride (1.6 g, 10.5 mmol) was dissolved in 20 mL of trimethyl phosphate. Under nitrogen gas protection, the mixture was cooled to 0 °C, and YK-CAP-109-PM4 (1.1 g, 3.51 mmol) was slowly added to the reaction mixture. The mixture was stirred at 0 °C for approximately 4 hours. After the reaction was complete, 20 mL of ice water was added, washed twice with ethyl acetate, and the aqueous phase was adjusted to pH 3.5 with aqueous ammonia and placed in a refrigerator overnight. The next day, the pH was adjusted to 6.5, diluted to 400 mL, and loaded. The target product peak was purified on a gel column (eluted with water:1.5 M TEAB = 1:4), collected, concentrated, and lyophilized to obtain a white solid, YK-CAP-109-PM5 (triethylamine salt, 980 mg, 1.98 mmol, 56.5%). 12 H 17 FN5O7P, MS(ES): m / z(MH - )392.1.

[0254] Step 6: Synthesis of YK-CAP-109-PM6 Using YK-CAP-109-PM5 (980 mg, 1.98 mmol) as a raw material, YK-CAP-109-PM6 (sodium salt, 900 mg, 1.93 mmol, 97.6%) was obtained via the synthetic route of YK-CAP-101-PM4. 15 H 19 FN7O6P, MS(ES): m / z(MH - )442.1.

[0255] Step 7: Synthesis of YK-CAP-109-PM7 Using YK-CAP-109-PM6 (900 mg, 1.93 mmol) as a starting material, YK-CAP-109-PM7 (triethylamine salt, 850 mg, 1.48 mmol, 76.7%) was obtained via the synthetic route of YK-CAP-101-PM5. 12 H 18 FN5O 10 P2, MS(ES): m / z(MH - )472.2.

[0256] Step 8: Synthesis of YK-CAP-109-PM8 Using YK-CAP-109-PM7 (850 mg, 1.48 mmol) as a starting material, YK-CAP-109-PM8 (triethylamine salt, 450 mg, 0.76 mmol, 51.7%) was obtained via the synthetic route of YK-CAP-101-PM6. 13 H 20 FN5O 10 P2, MS(ES): m / z(MH - )486.1.

[0257] Step 9: Synthesis of YK-CAP-109 Using YK-CAP-109-PM8 (100 mg, 0.17 mmol) as a starting material, YK-CAP-109 (35 mg, 28.53 μmol, 16.8%) was obtained via the synthetic route of YK-CAP-101. 34 H 46 FN 15 O 23 P4, MS(ES): m / z(MH - )1174.2. 1 HNMR(400MHz,D2O)δ8.53-8.41(m,1H),8.14(d,J=2.6Hz,1H),7.63-7.55(m,2H) ,6.17(d,J=4.4Hz,1H),5.65(d,J=2.6Hz,1H),5.32-5.21(m,1H),4.62-4.42(m,3 H),4.36-4.33(m,4H),4.22-4.15(m,3H),4.11(s,2H),4.03(s,1H),3.91-3.82( m,3H),3.21-3.15(m,3H),3.11(s,2H),2.31-2.22(m,1H),1.52(d,J=6.1Hz,3H). 31 PNMR(D2O,163MHz)δ-0.89(s,1P),-11.21(d,J=20.8Hz,1P),-11.74(d,J=16.6Hz,1P),-22.12(t,J=18.0Hz,1P).

[0258] 12. Synthesis of YK-CAP-110

[0259] [ka]

[0260] Step 1: Synthesis of YK-CAP-110-PM1 YK-CAP-107-PM2 (14.4 g, 32.68 mmol) was dissolved in dichloromethane, the reaction mixture was cooled to 0 °C, and DAST (16.7 g, 103.5 mmol) was slowly added dropwise. The reaction mixture was stirred at 0 °C for 4 hours. Saturated aqueous sodium bicarbonate was slowly added to the reaction mixture to quench it. DCM was added and stirred, and the organic phase was washed twice with saturated aqueous sodium bicarbonate and separated. The organic phase was washed two to three times with saturated brine, dried over anhydrous sodium sulfate, filtered, and spin-dried. The residue was purified by silica gel column chromatography (0-30% ethyl acetate / n-hexane) to give YK-CAP-110-PM1 (10.6 g, 22.91 mmol, 70.1%).

[0261] Step 2: Synthesis of YK-CAP-110-PM2 YK-CAP-110-PM1 (5.6 g, 12.11 mmol) was dissolved in acetic acid, and acetic anhydride (24.7 g, 242.1 mmol) was added. Concentrated sulfuric acid (280 μL) was then slowly added dropwise. After the addition was complete, the mixture was stirred at room temperature for 16 hours. 200 mL of water was added to the reaction solution, and the mixture was extracted with ethyl acetate. The organic phase was washed three times with saturated aqueous sodium bicarbonate until the pH became basic. The organic phase was dried and spun to obtain the crude product YK-CAP-110-PM2 (6.0 g), which was a yellow oily liquid and used directly in the next step of the reaction.

[0262] Step 3: Synthesis of YK-CAP-110-PM3 Intermediate INT-I (5.2 g, 13.4 mmol) was dissolved in 1,2-dichloroethane, and N,O-bis(trimethylsilyl)acetamide (7.4 g, 36.3 mmol) was added. The mixture was heated to 80°C and stirred for 2 hours. The reaction solvent was then removed under reduced pressure, and the mixture was dissolved in toluene again. At room temperature, a toluene solution of YK-CAP-110-PM2 (6.0 g, calculated based on 12.11 mmol) and TMSOTf (3.0 g, 13.4 mmol) were added in that order. The mixture was then heated to 70°C and stirred for another 2 hours. The reaction mixture was diluted with ethyl acetate, washed once with saturated sodium bicarbonate solution, filtered to remove insoluble materials, and the filtrate was separated. The organic phase was dried over anhydrous sodium sulfate, and the solvent was removed under reduced pressure. The residue was purified by silica gel column chromatography (0-30% ethyl acetate / DCM) to give YK-CAP-110-PM3 (3.5 g, 5.48 mmol). 30 H 28 F2N6O8, MS(ES): m / z (M+H + )639.1.

[0263] Step 4: Synthesis of YK-CAP-110-PM4 YK-CAP-110-PM3 (3.5 g, 5.48 mmol) was dissolved in a 4 M ammonia-methanol solution and water (5:1) and stirred at room temperature for 16 hours. The solvent was removed by spin-drying under reduced pressure, and the crude product was recrystallized from ethyl acetate to give YK-CAP-110-PM4 (1.3 g, 4.10 mmol, 74.7%). 11 H 13 F2N5O4, MS(ES): m / z (M+H + )318.1. YK-CAP-110-PM4: 1 H NMR (400 MHz, DMSO-d 6 )δ10.56(s,1H),8.00(s,1H),δ6.40(s,2H),5.71-5.56(m,2H),4.86-4.83(m,1H),4.36-4.32(m ,1H),4.19-4.15(m,1H),4.62(s,1H),3.75-3.73(m,1H),3.62-3.49(m,1H),3.27-3.22(m,1H).

[0264] Step 5: Synthesis of YK-CAP-110-PM5 Phosphorus oxychloride (1.9 g, 12.3 mmol) was dissolved in 20 mL of trimethyl phosphate. The mixture was cooled to 0 °C under nitrogen gas protection, and YK-CAP-110-PM4 (1.3 g, 4.10 mmol) was slowly added to the reaction mixture. The mixture was stirred at 0 °C for approximately 4 hours. After the reaction was complete, 20 mL of ice water was added, washed twice with ethyl acetate, and the aqueous phase was adjusted to pH 3.5 with aqueous ammonia and placed in a refrigerator overnight. The next day, the pH was adjusted to 6.5, diluted to 400 mL, and loaded. The product was purified on a gel column (eluted with water:1.5 M TEAB = 1:5). The peak of the target product was collected, concentrated, and lyophilized to obtain YK-CAP-110-PM5 (triethylamine salt, 1.1 g, 2.21 mmol, 53.8%) as a white solid. 11 H 14 F2N5O7P, MS(ES): m / z(MH - )396.1.

[0265] Step 6: Synthesis of YK-CAP-110-PM6 Using YK-CAP-110-PM5 (1.1 g, 2.21 mmol) as a raw material, YK-CAP-110-PM6 (sodium salt, 1.0 g, 2.13 mmol, 96.4%) was obtained via the synthetic route of YK-CAP-101-PM4. 14 H 16 F2N7O6P, MS(ES): m / z(MH - )446.1.

[0266] Step 7: Synthesis of YK-CAP-110-PM7 Using YK-CAP-110-PM6 (1.0 g, 2.13 mmol) as a starting material, YK-CAP-110-PM7 (triethylamine salt, 900 mg, 1.56 mmol, 73.1%) was obtained via the synthetic route of YK-CAP-101-PM5. 11 H 15 F2N5O 10 P2, MS(ES): m / z(MH - )476.1.

[0267] Step 8: Synthesis of YK-CAP-110-PM8 Using YK-CAP-110-PM7 (900 mg, 1.56 mmol) as a starting material, YK-CAP-110-PM8 (triethylamine salt, 500 mg, 0.84 mmol, 54.1%) was obtained via the synthetic route of YK-CAP-101-PM6. 12 H 17 F2N5O 10 P2, MS(ES): m / z(MH - )490.2.

[0268] Step 9: Synthesis of YK-CAP-110 Using YK-CAP-110-PM8 (100 mg, 0.17 mmol) as a starting material, YK-CAP-110 (25 mg, 22.03 μmol, 13.0%) was obtained via the synthetic route of YK-CAP-101. 33 H 43 F2N 15 O 23 P4, MS(ES): m / z(MH - )1178.1. 1HNMR(400MHz,D2O)δ8.43(d,J=1.5Hz,1H),8.36(d,J=2.6Hz,1H),8.12(d,J=4.6Hz,2H), 6.10(d,J=5.4Hz,1H),5.88(d,J=2.2Hz,1H),5.61(d,J=4.4Hz,1H),4.90-4.88(m,3H),4 .85-4.82(m,1H),4.62(t,J=4.7Hz,1H),4.51-4.38(m,4H),4.23(d,J=3.0Hz,1H),4.12( s,2H),4.04(s,2H),3.93(s,3H),3.64(t,J=7.0Hz,1H),3.41(s,3H),3.28-3.22(m,1H). 31 PNMR(D2O,162MHz)δ-0.93(s,1P),-11.17(d,J=19.2Hz,1P),-11.35(d,J=17.1Hz,1P),-24.38(t,J=17.6Hz,1P).

[0269] 13.Synthesis of YK-CAP-111

[0270] [ka]

[0271] Step 1: Synthesis of YK-CAP-111-PM1 YK-CAP-107-PM1 (35.0 g, 79.07 mmol) was dissolved in acetonitrile and water (1:1, 280 mL). Iodobenzene diacetate (53.5 g, 166.03 mmol), sodium bicarbonate (9.96 g, 118.56 mmol), and TEMPO (1.85 g, 11.85 mmol) were added sequentially in an ice bath and the mixture was allowed to react at room temperature for 2 hours. After completion of the reaction was monitored by TLC, the reaction was stopped and quenched by adding saturated aqueous sodium thiosulfite solution. The mixture was extracted with EA and separated. The organic phase was dried over anhydrous sodium sulfate, filtered, and the solvent of the organic phase was spun off to give YK-CAP-111-PM1 (68.0 g, crude brown oil) which was used directly in the next step without further purification.

[0272] Step 2: Synthesis of YK-CAP-111-PM2 YK-CAP-111-PM1 (34.0 g, calculated based on 39.54 mmol) was dissolved in acetonitrile and cooled to 0°C. DIEA (14.0 g, 108.33 mmol) and HATU (17.86 g, 46.97 mmol) were added. The mixture was stirred for 20 minutes, and then diethylamine (6.6 g, 90.24 mmol) was added. The mixture was warmed to room temperature and stirred for 4 hours. The reaction solution was concentrated under reduced pressure, and the residue was purified by silica gel column chromatography (0-20% ethyl acetate / n-hexane) to give YK-CAP-111-PM2 (17.5 g, 34.20 mmol, 86.5%).

[0273] Step 3: Synthesis of YK-CAP-111-PM3 YK-CAP-111-PM2 (17.5 g, 34.20 mmol) was dissolved in tetrahydrofuran, and TBAF (13.4 g, 51.25 mmol) was added. The mixture was stirred at room temperature for 2 hours. The reaction solution was spin-dried, and the residue was purified by silica gel column chromatography (0-90% ethyl acetate / n-hexane) to give YK-CAP-111-PM3 (8.9 g, 32.56 mmol, 95.2%).

[0274] Step 4: Synthesis of YK-CAP-111-PM4 YK-CAP-111-PM3 (4.0 g, 14.63 mmol) was dissolved in acetic acid, sulfuric acid (300 μL) was added, and the mixture was stirred at room temperature for 30 minutes. Acetic anhydride (30.0 g, 293.86 mmol) was then added. The mixture was stirred at room temperature for 18 hours. To the reaction solution, 200 mL of water was added, and the mixture was extracted with ethyl acetate. The organic phase was washed three times with saturated aqueous sodium bicarbonate until the pH became basic. The organic phase was dried and spun to obtain the crude product YK-CAP-111-PM4 (3.19 g), which was a yellow oily liquid and used directly in the next step of the reaction.

[0275] Step 5: Synthesis of YK-CAP-111-PM5 Intermediate INT-I (3.79 g, 9.76 mmol) was dissolved in 1,2-dichloroethane, N,O-bis(trimethylsilyl)acetamide (5.42 g, 26.64 mmol) was added, and the mixture was heated to 80°C and stirred for 2 hours to react. Next, the reaction solvent was removed under reduced pressure, and a toluene solution of YK-CAP-111-PM4 (3.19 g) and TMSOTf (2.96 g, 13.32 mmol) were added sequentially at room temperature, followed by heating to 70°C and stirring for another 2 hours to react. The reaction mixture was diluted with ethyl acetate, washed once with saturated sodium bicarbonate solution, filtered, the insoluble matter was removed, the filtrate was separated, the organic phase was dried over anhydrous sodium sulfate, the solvent was removed under reduced pressure, and the residue was purified by silica gel column chromatography (0–60% ethyl acetate / DCM) to give YK-CAP-111-PM5 (2.20 g, 3.20 mmol).

[0276] Step 6: Synthesis of YK-CAP-111-PM6 YK-CAP-111-PM5 (2.20 g, 3.20 mmol) was dissolved in a 4 M ammonia-methanol solution and water (5:1) and stirred at room temperature for 16 hours. The solvent was spun off under reduced pressure, and the crude product was recrystallized from ethyl acetate to give YK-CAP-111-PM6 (890 mg, 2.43 mmol, 75.9%). 15 H 22 N6O5, MS(ES): m / z (M+H + )367.1. YK-CAP-111-PM6: 1 H NMR (400 MHz, DMSO-d 6 )δ11.40(s,1H),8.31(s,1H),6.38(s,2H),6.16(d,J=4.3Hz,1H),5.38-5.41(m,1H),4.84-4.91(m,1H),4.28-4.31(m, 1H),3.98-4.02(m,1H),3.45-3.49(m,2H),3.33-3.21(q,J=4.1Hz,4H),2.65-2.62(m,1H),1.17-1.15(t,J=4.1Hz,6H).

[0277] Step 7: Synthesis of YK-CAP-111-PM7 Phosphorus oxychloride (1.3 g, 8.48 mmol) was dissolved in 15 mL of trimethyl phosphate. The mixture was cooled to 0 °C under nitrogen gas protection, and YK-CAP-111-PM6 (880 mg, 2.42 mmol) was slowly added to the reaction mixture. The mixture was stirred at 0 °C for 4 hours. After the reaction was complete, the mixture was added to 20 mL of ice water, washed twice with ethyl acetate, and the aqueous phase was adjusted to pH 3.5 with aqueous ammonia and left in the refrigerator overnight. The pH was then adjusted to 6.5, diluted to 400 mL, and loaded. The product was purified on a silica gel column (eluted with water:1.5 M TEAB = 5:1), and the peak of the target product was collected, concentrated, and lyophilized to obtain YK-CAP-111-PM7 (triethylamine salt, 900 mg, 1.64 mmol, 68.5%) as a white solid. 15 H 23 N6O8P, MS(ES): m / z(MH - )445.1.

[0278] Step 8: Synthesis of YK-CAP-111-PM8 Using YK-CAP-111-PM7 (900 mg, 1.64 mmol) as a starting material, YK-CAP-111-PM8 (sodium salt, 730 mg, 1.41 mmol, 85.7%) was obtained via the synthetic route of YK-CAP-101-PM4. 18 H 25 N8O7P, MS(ES): m / z(MH - )495.1.

[0279] Step 9: Synthesis of YK-CAP-111-PM9 Using YK-CAP-111-PM8 (720 mg, 1.39 mmol) as a raw material, YK-CAP-111-PM9 (triethylamine salt, 750 mg, 1.20 mmol, 86.1%) was obtained via the synthetic route of YK-CAP-101-PM5. 15 H 24 NO 11 P2, MS(ES): m / z(MH - )525.1.

[0280] Step 10: Synthesis of YK-CAP-111-PM10 YK-CAP-111-PM9 (400 mg, 0.64 mmol) was dissolved in HO (20 mL), and the pH was adjusted to 4.0 with glacial acetic acid. Dimethyl sulfate (800 μL, 8.45 mmol) was added within 30 min. The pH of the reaction mixture was maintained at 3.8–4.1 with 0.1 M NaOH aqueous solution and stirred at room temperature for 5 h. After completion of the reaction, the reaction solution was quenched twice with dichloromethane. The pH of the aqueous phase was adjusted to 6.5, the volume was diluted to 400 mL, and the mixture was purified on a gel column (eluted with water and 1 M TEAB = 3:2). The peak of the target product was collected, concentrated, and lyophilized to give YK-CAP-111-PM10 (triethylamine salt, 350 mg, 0.55 mmol, 85.6%) as a white solid. 16 H 26 NO 11 P2, MS(ES): m / z(MH - )539.1.

[0281] Step 11: Synthesis of YK-CAP-111 Using YK-CAP-111-PM10 (100 mg, 0.16 mmol) as a raw material, YK-CAP-111 (ammonium salt, 50 mg, 39.07 μmmol, 25.1%) was obtained via the synthetic route of YK-CAP-101. 37 H 52 N 16 O 24 P4, MS(ES): m / z(MH - )1127.2. 1HNMR(400MHz,D2O)δ8.43(d,J=1.3Hz,1H),8.35(s,1H),8.06(d,J=6.0Hz,2H),6.01(d,J=5.4Hz ,1H),5.86(d,J=2.3Hz,1H),5.82(d,J=4.5Hz,1H),4.97-4.84(m,3H),4.66(t,J=4.8Hz,1H),4. 51-4.35(m,4H),4.32(d,J=4.5Hz,1H),4.21(s,2H),4.09(s,2H),3.97(s,3H),3.72(t,J=7.0Hz ,1H),3.35-3.21(d,J=4.1Hz,4H),3.04(s,3H),1.94-1.87(m,1H),1.21-1.14(t,J=4.1Hz,6H). 31 PNMR(D2O,162MHz)δ-0.92(s,1P),-11.15(d,J=19.2Hz,1P),-11.62(d,J=17.1Hz,1P),-23.46(t,J=17.5Hz,1P).

[0282] 14. Synthesis of YK-CAP-112

[0283] [ka]

[0284] Step 1: Synthesis of YK-CAP-112-PM1 Using YK-CAP-111-PM1 (10.0 g, 21.90 mmol) and dipropylamine (5.5 g, 54.35 mmol) as raw materials, YK-CAP-112-PM1 (11.0 g, 20.38 mmol, 93.1%) was obtained via the synthetic route of YK-CAP-111-PM2.

[0285] Step 2: Synthesis of YK-CAP-112-PM2 Using YK-CAP-112-PM1 (11.0 g, 20.38 mmol) as a raw material, YK-CAP-112-PM2 (6.0 g, 19.91 mmol, 97.7%) was obtained via the synthetic route of YK-CAP-111-PM3.

[0286] Step 3: Synthesis of YK-CAP-112-PM3 Starting from YK-CAP-112-PM2 (6.0 g, 19.91 mmol), the crude product YK-CAP-112-PM3 (5.1 g) was obtained via the synthetic route of YK-CAP-111-PM4. This crude product was a yellow oily liquid and was used directly in the next step of the reaction.

[0287] Step 4: Synthesis of YK-CAP-112-PM4 Using YK-CAP-112-PM3 (5.1 g) as a raw material, YK-CAP-112-PM4 (3.9 g, 5.45 mmol) was obtained via the synthetic route of YK-CAP-111-PM5. 36 H 41 N7O9, MS(ES): m / z (M+H + )716.3.

[0288] Step 5: Synthesis of YK-CAP-112-PM5 Using YK-CAP-112-PM4 (3.9 g, 5.45 mmol) as a raw material, YK-CAP-112-PM5 (1.7 g, 4.31 mmol, 79.1%) was obtained via the synthetic route of YK-CAP-111-PM6. 17 H 26 N6O5, MS(ES): m / z (M+H + )395.2. YK-CAP-112-PM5: 1 H NMR (400 MHz, DMSO-d 6 )δ11.37(s,1H),8.41(s,1H),6.28(s,2H),6.19(d,J=4.1,1H),5.46(s,1H),5.01(s,1H),4.25-4.28(m,1H),3.99 -4.01(m,1H),3.41-3.43(m,2H),3.21-3.24(m,4H),2.56-2.58(m,1H),1.57-1.54(m,4H),0.87(t,J=1.8Hz,6H).

[0289] Step 6: Synthesis of YK-CAP-112-PM6 Using YK-CAP-112-PM5 (1.7 g, 4.31 mmol) as a raw material, YK-CAP-112-PM6 (triethylamine salt, 1.80 g, 3.13 mmol, 72.6%) was obtained via the synthetic route of YK-CAP-111-PM7. 17 H 27 N6O8P, MS(ES): m / z(MH - )473.1.

[0290] Step 7: Synthesis of YK-CAP-112-PM7 Using YK-CAP-112-PM6 (1.8 g, 3.13 mmol) as a raw material, YK-CAP-112-PM7 (sodium salt, 1.47 g, 2.69 mmol, 86.0%) was obtained via the synthetic route of YK-CAP-101-PM4. 20 H 29 N8O7P, MS(ES): m / z(MH - )523.1.

[0291] Step 8: Synthesis of YK-CAP-112-PM8 Using YK-CAP-112-PM7 (1.47 g, 2.69 mmol) as a starting material, YK-CAP-112-PM8 (triethylamine salt, 1.30 g, 1.98 mmol, 73.7%) was obtained via the synthetic route of YK-CAP-101-PM5. 17 H 28 NO 11 P2, MS(ES): m / z(MH - )553.1.

[0292] Step 9: Synthesis of YK-CAP-112-PM9 Using YK-CAP-112-PM8 (600 mg, 0.92 mmol) as a starting material, YK-CAP-112-PM9 (triethylamine salt, 200 mg, 0.30 mmol, 32.6%) was obtained via the synthetic route of YK-CAP-111-PM10. 18 H 30 NO 11 P2, MS(ES): m / z(MH - )567.1.

[0293] Step 10: Synthesis of YK-CAP-112 Using YK-CAP-112-PM9 (200 mg, 0.30 mmol) as a starting material, YK-CAP-112 (ammonium salt, 50 mg, 38.23 μmol, 12.8%) was obtained via the synthetic route of YK-CAP-101. 39 H 56 N 16 O 24 P4, MS(ES): m / z(MH - )1255.2. 1 HNMR(400MHz,D2O)δ8.43(d,J=1.3Hz,1H),8.34(s,1H),8.11(d,J=6.1Hz,2H),6.05(d,J=5.3Hz,1 H),5.87(d,J=2.3Hz,1H),5.81(d,J=4.4Hz,1H),4.91-4.82(m,2H),4.64(t,J=4.7Hz,1H),4.48-4. 37(m,5H),4.30(d,J=4.6Hz,1H),4.22(s,2H),4.11(s,2H),3.96(s,3H),3.72(t,J=7.0Hz,1H),3.3 3-3.26(t,J=4.1Hz,4H),3.11(s,3H),2.22-2.14(m,1H),1.58-1.54(m,4H),0.87(t,J=1.8Hz,6H). 31 PNMR(D2O,162MHz)δ-0.94(s,1P),-11.22(d,J=19.1Hz,1P),-11.52(d,J=17.2Hz,1P),-23.11(t,J=17.3Hz,1P).

[0294] 15. Synthesis of YK-CAP-113

[0295] [ka]

[0296] Step 1: Synthesis of YK-CAP-113-PM1 Methyl β-D-ribofuranoside (100.0 g, 0.61 mol) was dissolved in 1 L of anhydrous pyridine, and TIPDSCl (230.6 g, 0.73 mol) was added dropwise in an ice-water bath. After the addition was complete, the mixture was warmed to room temperature and stirred for 12 hours. After concentrating under reduced pressure to remove most of the solvent, the residue was purified by silica gel column chromatography (0-30% ethyl acetate / n-hexane) to give YK-CAP-113-PM1 (212.2 g, 0.52 mol, 85.7%).

[0297] Step 2: Synthesis of YK-CAP-113-PM2 YK-CAP-113-PM1 (212.2 g, 0.52 mol) was dissolved in 2 L of acetonitrile, and Dess-Martin reagent (485.1 g, 1.14 mol) was added. The mixture was heated to 40°C and stirred for 12 hours. After cooling to room temperature, the reaction solution was filtered, and the filtrate was spin-dried under reduced pressure to obtain YK-CAP-113-PM2 (208.8 g, 0.52 mol, 98.9%).

[0298] Step 3: Synthesis of YK-CAP-113-PM3 (Bromomethyl)triphenylphosphonium bromide (408.7 g, 1.14 mol) was dissolved in tetrahydrofuran (3000 mL) and cooled to -78 °C. A 2.5 M solution of n-butyllithium in tetrahydrofuran (456 mL, 1.14 mol) was slowly added dropwise. After the addition was complete, the reaction mixture was allowed to warm to 0 °C and stirred for 2 hours. The reaction mixture was again cooled to -78 °C, and a solution of YK-CAP-113-PM2 (208.8 g, 0.52 mol) in tetrahydrofuran (400 mL) was slowly added dropwise. After the addition was complete, the reaction mixture was allowed to warm to room temperature and stirred overnight. The reaction mixture was quenched with saturated ammonium chloride solution (2000 mL) and extracted with ethyl acetate (2000 mL x 3). The combined organic phase was washed with saturated aqueous NaCl, dried over anhydrous sodium sulfate, filtered, and spin-dried. The residue was purified by silica gel column chromatography (0-20% ethyl acetate / n-hexane) to obtain YK-CAP-113-PM3 (108.8 g, 0.27 mol, 52.4%).

[0299] Step 4: Synthesis of YK-CAP-113-PM4 1M 9-BBN tetrahydrofuran (540 mL, 0.54 mmol) was added to a three-neck flask and cooled to 0°C under nitrogen gas protection. A tetrahydrofuran solution of YK-CAP-113-PM3 (108.8 g, 0.27 mol) was slowly added dropwise to the reaction mixture. After the addition was complete, the mixture was stirred at room temperature for 2 hours. After the reaction mixture was cooled to 0°C again, water / tetrahydrofuran (1:1, 540 mL), 2N sodium hydroxide solution (540 mL), and 30% hydrogen peroxide (460 mL) were slowly added sequentially. After the addition was complete, the mixture was warmed to room temperature and stirred for an additional 3 hours. The reaction mixture was diluted with ethyl acetate, separated, and the aqueous phase was back-extracted once with ethyl acetate. The organic phases were combined, dried, filtered, and the filtrate was spun to dryness under reduced pressure. The residue was purified by silica gel column chromatography (0–17% ethyl acetate / n-hexane) to give YK-CAP-113-PM4 (98.0 g, 0.23 mol, 86.2%).

[0300] Step 5: Synthesis of YK-CAP-113-PM5 YK-CAP-113-PM4 (20.0 g, 47.54 mmol) and sodium hydride (1369 mg, 57.04 mmol) were dissolved in dry tetrahydrofuran (100 mL) and cooled to 0 °C. Methyl iodide (13.5 g, 95.11 mmol) was added dropwise under nitrogen gas protection and stirred for approximately 5 h. After completion of the reaction, the reaction was quenched by adding water (10 mL) and extracted with EA (200 mL x 3). The combined organic phase was washed with saturated aqueous NaCl, dried over anhydrous sodium sulfate, filtered, and spin-dried. The residue was purified by silica gel column chromatography (0-20% ethyl acetate / n-hexane) to give YK-CAP-113-PM5 (17.8 g, 40.95 mmol, 86.1%).

[0301] Step 6: Synthesis of YK-CAP-113-PM6 N6-Benzoyladenosine (39.2 g, 163.85 mmol) was dissolved in hexamethyldisilazane (500 mL), a catalytic amount of ammonium sulfate was added, and the mixture was heated to 130 °C under nitrogen protection and stirred for 12 hours. The solvent was then spin-dried. YK-CAP-113-PM5 (17.8 g, 40.95 mmol) was dissolved in 1,2-dichloroethane (300 mL) and added to the residue. Trimethylsilyl trifluoromethanesulfonate (10.1 g, 48.97 mmol) was added and the mixture was reacted at 80 °C for 5 hours. Completion of the reaction was monitored by thin-layer chromatography. After filtration, the filtrate was spin-dried under reduced pressure, and the residue was purified by silica gel column chromatography (0-80% ethyl acetate / n-hexane) to give YK-CAP-113-PM6 (14.5 g, 22.59 mmol, 55.2%). 31 H 47 N5O6Si2, MS(ES): m / z (M+H + )642.3.

[0302] Step 7: Synthesis of YK-CAP-113-PM7 YK-CAP-113-PM6 (14.5 g, 22.59 mmol) was dissolved in tetrahydrofuran (100 mL), tetra-n-butylammonium fluoride (23.6 g, 90.26 mmol) was added, and the mixture was stirred at room temperature for 1 hour. Saturated aqueous ammonium chloride was added to the reaction mixture, followed by extraction with ethyl acetate (200 mL x 3). The combined organic phase was washed with saturated aqueous NaCl, dried over anhydrous sodium sulfate, filtered, and spin-dried. The residue was purified by silica gel column chromatography (0-60% ethyl acetate / n-hexane) to yield YK-CAP-113-PM7 (7.5 g, 18.78 mmol, 83.1%). 19 H 21 N5O5, MS(ES): m / z (M+H + )400.2.

[0303] Step 8: Synthesis of YK-CAP-113-PM8 YK-CAP-113-PM7 (7.5 g, 18.78 mmol) was dissolved in pyridine (50 mL), and 4,4'-dimethoxytriphenylmethyl chloride (10.2 g, 30.10 mmol) was added at room temperature. The mixture was stirred at room temperature for 3 hours. Completion of the reaction was monitored by TLC, and the reaction solution was quenched by adding 10 mL of methanol. The mixture was stirred for 10 minutes and then rotary evaporated until the solvent ceased to evaporate, yielding the crude product. The crude product was purified by flash column chromatography to yield YK-CAP-113-PM8 (8.8 g, 12.54 mmol, 66.8%). 40 H 39 N5O7, MS(ES): m / z (M+H + )702.3.

[0304] Step 9: Synthesis of YK-CAP-113-PM9 YK-CAP-113-PM8 (8.8 g, 12.54 mmol) was dissolved in acetonitrile (100 mL), and N-methylimidazole (1.1 g, 13.40 mmol) and bis(diisopropylamino)(2-cyanoethoxy)phosphine (11.3 g, 37.49 mmol) were added sequentially. The mixture was stirred under nitrogen gas protection for 6 hours. Completion of the reaction was monitored by TLC. The reaction solution was diluted with ethyl acetate, and the organic phase was washed with saturated aqueous sodium bicarbonate and separated. The organic phase was washed once with water and separated. The combined aqueous phase was back-extracted once with ethyl acetate. The combined organic phase was dried over anhydrous sodium sulfate and filtered. The organic phase was rotary evaporated under reduced pressure until the solvent ceased to evaporate, yielding the crude product. The crude product was purified by flash column chromatography. After evaporating the solvent from the product components, the product was dissolved in ethyl acetate and added dropwise to ice-cold n-hexane and stirred for 10 minutes. Filtration gave YK-CAP-113-PM9 (8.1 g, 8.98 mmol, 71.6%). 49 H 56 N7O8P, MS(ES): m / z(MH - )900.4.

[0305] Step 10: Synthesis of YK-CAP-113-PM10 YK-CAP-113-PM9 (4.0 g, 4.43 mmol), guanosine-9-17N,N-(2-methyl-1-oxopropyl)-2',3'-diacetate (2.0 g, 4.57 mmol), and tetrazole (3.1 g, 44.25 mmol) were dissolved in acetonitrile (50 mL) and stirred at room temperature for 3 hours under nitrogen gas protection. A 0.1 M iodine solution (53 mL) was added to the reaction solution, followed by stirring for 1 hour. The reaction solution was diluted with 200 mL of brine and extracted with dichloromethane (200 mL x 3). The combined organic phases were dried over anhydrous sodium sulfate, filtered, and spin-dried. The organic phase was rotary evaporated under reduced pressure until the solvent ceased to evaporate, yielding YK-CAP-113-PM10 (5.2 g, 4.15 mmol, 93.5%). 61 H 64 N 11 O 17P, MS(ES): m / z(MH - )1252.4.

[0306] Step 11: Synthesis of YK-CAP-113-PM11 YK-CAP-113-PM10 (5.2 g, 4.15 mmol) was dissolved in 80% aqueous acetic acid (20 mL) and stirred at room temperature for 2 hours under nitrogen gas protection. The reaction mixture was concentrated under reduced pressure to remove acetic acid, and the solvent was rotary evaporated until no more solvent was evaporated to obtain the crude product. The crude product was purified by flash column chromatography to obtain YK-CAP-113-PM11 (3.0 g, 3.15 mmol, 76.0%). 40 H 46 N 11 O 15 P, MS(ES): m / z(MH - )950.3.

[0307] Step 12: Synthesis of YK-CAP-113-PM12 YK-CAP-113-PM11 (3.0 g, 3.15 mmol) was dissolved in acetonitrile (30 mL), and N-methylimidazole (0.5 g, 6.09 mmol) and bis(diisopropylamino)(2-cyanoethoxy)phosphine (2.8 g, 9.29 mmol) were added sequentially. The reaction was stirred for 2 hours under nitrogen gas protection. A 0.1 M iodine solution (40 mL) was added to the reaction solution, followed by stirring for 1 hour. The reaction solution was diluted with 200 mL of brine and extracted with dichloromethane (200 mL x 3). The combined organic phases were dried over anhydrous sodium sulfate, filtered, and spin-dried. The organic phase was rotary evaporated under reduced pressure until the solvent ceased to evaporate, yielding YK-CAP-113-PM12 (1.8 g, 1.66 mmol, 52.6%). 43 H 50 N 12 O 18 P2, MS(ES): m / z(MH - )1083.3.

[0308] Step 13: Synthesis of YK-CAP-113-PM13 YK-CAP-113-PM12 (1.8 g, 1.66 mmol) was dissolved in a mixture of aqueous ammonia (10 mL) and methanol (5 mL). The mixture was heated to 50°C under nitrogen gas protection and stirred for 24 hours. The reaction mixture was concentrated under reduced pressure to remove the solvent, and rotary evaporated until the solvent stopped evaporating to obtain the crude product. Water (50 mL) was added to completely dissolve the product, and the product was purified on a gel column (eluted with water and 1.5 M TEAB = 10:1). The target product peak was collected, concentrated, lyophilized, and desalted by high-performance liquid chromatography (50 mM TEAB and methanol mobile phase system) to obtain a white solid, YK-CAP-113-PM13 (triethylamine salt, 660 mg, 0.80 mmol, 48.4%). 22 H 30 N 10 O 14 P2, MS(ES): m / z(MH - )719.1.

[0309] Step 14: Synthesis of YK-CAP-113 YK-CAP-113-PM11 (150 mg, 0.18 mmol) and Im-m7GDP (193 mg, 0.36 mmol) were used as raw materials to obtain YK-CAP-113 (ammonium salt, 40 mg, 33.04 μmol, 18.1%) by the synthesis method for YK-CAP-101. 33 H 45 N 15 O 24 P4, MS(ES): m / z(MH - )1158.2. 1HNMR(400MHz,D2O)δ8.44(d,J=1.3Hz,1H),8.33(s,1H),8.08(d,J=6.0Hz,2H),6.02(d ,J=5.4Hz,1H),5.90(d,J=2.3Hz,1H),5.82(d,J=4.4Hz,1H),4.95-4.82(m,3H),4.63( t,J=4.5Hz,1H),4.53-4.31(m,4H),4.26(d,J=4.5Hz,1H),4.21(s,3H),4.09(s,1H),3 .76(t,J=7.0Hz,1H),3.43(s,3H),3.35(s,3H),3.21-3.11(m,2H),2.42-2.35(m,1H). 31 PNMR(D2O,162MHz)δ-0.94(s,1P),-11.34(d,J=19.2Hz,1P),-11.82(d,J=17.3Hz,1P),-23.11(t,J=17.3Hz,1P).

[0310] 16. Synthesis of YK-CAP-114

[0311] [ka]

[0312] Step 1: Synthesis of YK-CAP-114-PM1 YK-CAP-113-PM4 (13.3 g, 31.61 mmol), phthalimide (5.6 g, 38.06 mmol), triphenylphosphine (16.6 g, 63.29 mmol), and DIAD (7.7 g, 38.08 mmol) were dissolved in dry tetrahydrofuran (150 mL), cooled to 0 °C, and stirred under nitrogen gas protection for 5 h. After completion of the reaction, the reaction system was quenched by adding water (10 mL) and extracted with EA (200 mL x 3). The combined organic phase was washed with saturated aqueous NaCl, dried over anhydrous sodium sulfate, filtered, and spin-dried. The residue was purified by silica gel column chromatography (0-20% ethyl acetate / n-hexane) to give YK-CAP-114-PM1 (15.9 g, 28.92 mmol, 91.5%). 27 H 43NO7Si2, MS(ES): m / z (M+H + )550.3.

[0313] Step 2: Synthesis of YK-CAP-114-PM2 YK-CAP-114-PM1 (15.9 g, 28.92 mmol) was dissolved in 300 mL of ethanol, and 50 mL of hydrazine hydrate was added. The temperature was raised to 80°C, and the reaction was allowed to proceed with stirring for 12 hours. After the reaction was completed, the reaction system was quenched by adding water (10 mL), extracted with EA (200 mL x 3), and the organic phases were combined. The organic phase was washed with saturated aqueous NaCl, dried over anhydrous sodium sulfate, filtered, and spin-dried. YK-CAP-114-PM2 (12.0 g, 28.59 mmol, 98.9%) was obtained. 19 H 41 NO5Si2, MS(ES): m / z (M+H + )420.3.

[0314] Step 3: Synthesis of YK-CAP-114-PM3 YK-CAP-114-PM2 (12.0 g, 28.59 mmol) and triethylamine (5.9 g, 58.31 mmol) were dissolved in dichloromethane (150 mL) and cooled to 0 °C. Acetyl chloride (3.3 g, 42.04 mmol) was slowly added dropwise. After the addition was complete, the reaction mixture was maintained at 0 °C and stirred for 3 hours. After the reaction was complete, the reaction mixture was quenched with water (20 mL) and extracted with DCM (200 mL x 3). The combined organic phases were washed with saturated aqueous NaCl, dried over anhydrous sodium sulfate, filtered, and spin-dried. The residue was purified by silica gel column chromatography (0-20% ethyl acetate / n-hexane) to give YK-CAP-114-PM3 (12.2 g, 26.42 mmol, 92.4%). 21 H 43 NO6Si2, MS(ES): m / z (M+H + )462.3.

[0315] Step 4: Synthesis of YK-CAP-114-PM4 Using YK-CAP-114-PM3 (12.2 g, 26.42 mmol) as a raw material, YK-CAP-114-PM4 (8.5 g, 12.71 mmol, 48.1%) was obtained using the synthesis method for YK-CAP-113-PM6. 32 H 48 N6O6Si2, MS(ES): m / z (M+H + )669.3.

[0316] Step 5: Synthesis of YK-CAP-114-PM5 Using YK-CAP-114-PM4 (8.5 g, 12.71 mmol) as a raw material, YK-CAP-114-PM5 (4.4 g, 10.32 mmol, 81.2%) was obtained using the synthesis method for YK-CAP-113-PM7. 20 H 22 N6O5, MS(ES): m / z (M+H + )427.2.

[0317] Step 6: Synthesis of YK-CAP-114-PM6 Using YK-CAP-114-PM5 (4.4 g, 10.32 mmol) as a raw material, YK-CAP-114-PM6 (5.5 g, 7.55 mmol, 73.1%) was obtained by the synthesis method of YK-CAP-113-PM8. 41 H 40 N6O7, MS(ES): m / z (M+H + )729.3.

[0318] Step 7: Synthesis of YK-CAP-114-PM7 Using YK-CAP-114-PM6 (5.5 g, 7.55 mmol) as a raw material, YK-CAP-114-PM7 (5.0 g, 5.38 mmol, 71.3%) was obtained using the synthesis method for YK-CAP-113-PM9. 50 H 57 N8O8P, MS(ES): m / z(MH - )927.4.

[0319] Step 8: Synthesis of YK-CAP-114-PM8 Using YK-CAP-114-PM7 (5.0 g, 5.38 mmol) as a raw material, YK-CAP-114-PM8 (5.7 g, 4.45 mmol, 82.7%) was obtained by the synthesis method of YK-CAP-113-PM10. 62 H 65 N 12 O 17 P, MS(ES): m / z(MH - )1279.4.

[0320] Step 9: Synthesis of YK-CAP-114-PM9 Using YK-CAP-114-PM8 (5.7 g, 4.45 mmol) as a starting material, YK-CAP-114-PM9 (3.5 g, 3.58 mmol, 80.4%) was obtained by the same synthesis method as for YK-CAP-113-PM11. 41 H 47 N 12 O 15 P, MS(ES): m / z(MH - )977.3.

[0321] Step 10: Synthesis of YK-CAP-114-PM10 Using YK-CAP-114-PM9 (3.5 g, 3.58 mmol) as a raw material, YK-CAP-114-PM10 (2.0 g, 1.80 mmol, 50.3%) was obtained by the synthesis method of YK-CAP-113-PM12. 44 H 51 N 13 O 18 P2, MS(ES): m / z(MH - )1110.3.

[0322] Step 11: Synthesis of YK-CAP-114-PM11 Using YK-CAP-114-PM10 (2.0 g, 1.80 mmol) as a raw material, YK-CAP-114-PM11 (triethylamine salt, 800 mg, 0.94 mmol, 52.4%) was obtained by the synthesis method for YK-CAP-113-PM13. 23 H 31 N 11 O 14 P2, MS(ES): m / z(MH - )746.2.

[0323] Step 12: Synthesis of YK-CAP-114 Using YK-CAP-114-PM11 (150 mg, 0.18 mmol) as a raw material, YK-CAP-114 (ammonium salt, 26 mg, 21.01 μmol, 11.9%) was obtained by the synthesis method of YK-CAP-113. 34 H 46 N 16 O 24 P4, MS(ES): m / z(MH - )1185.2. 1 HNMR(400MHz,D2O)δ8.42(d,J=1.4Hz,1H),8.32(s,1H),8.11(d,J=6.0Hz,2H),6.12(d ,J=5.4Hz,1H),5.92(d,J=2.3Hz,1H),5.84(d,J=4.3Hz,1H),4.92-4.86(m,3H),4.61( t,J=4.5Hz,1H),4.52-4.33(m,4H),4.21(d,J=4.5Hz,1H),4.16(s,3H),4.03(s,1H),3 .72(t,J=6.3Hz,1H),3.42(s,3H),3.32-3.23(m,2H),2.42-2.32(m,1H),1.88(s,3H). 31 PNMR(D2O,162MHz)δ-0.88(s,1P),-11.12(d,J=19.1Hz,1P),-12.21(d,J=17.2Hz,1P),-23.55(t,J=17.1Hz,1P).

[0324] 17. Synthesis of YK-CAP-115

[0325] [ka]

[0326] Step 1: Synthesis of YK-CAP-115-PM1 YK-CAP-113-PM4 (20.0 g, 47.54 mmol) was dissolved in dichloromethane (200 mL) and cooled to -40 °C. A solution of diethylaminosulfur trifluoride (9.2 g, 57.08 mmol) in dichloromethane (20 mL) was slowly added dropwise. After the addition was complete, the mixture was slowly warmed to 0 °C and stirred for 4 h. Completion was monitored by thin-layer chromatography. The reaction was quenched with saturated aqueous sodium bicarbonate (100 mL) and extracted with dichloromethane (200 mL x 3). The combined organic phases were washed with saturated aqueous NaCl, dried over anhydrous sodium sulfate, filtered, and spin-dried. The residue was purified by silica gel column chromatography (0-40% ethyl acetate / n-hexane) to give YK-CAP-115-PM1 (11.2 g, 26.50 mmol, 55.7%).

[0327] Step 2: Synthesis of YK-CAP-115-PM2 Using YK-CAP-115-PM1 (11.2 g, 26.50 mmol) as a raw material, YK-CAP-115-PM2 (8.6 g, 13.65 mmol, 51.5%) was obtained by the synthesis method of YK-CAP-113-PM6. 30 H 44 FN5O5Si2, MS(ES): m / z (M+H + )630.3.

[0328] Step 3: Synthesis of YK-CAP-115-PM3 Using YK-CAP-115-PM2 (8.6 g, 13.65 mmol) as a raw material, YK-CAP-115-PM3 (4.8 g, 12.39 mmol, 90.8%) was obtained using the synthesis method for YK-CAP-113-PM7. 18 H 18 FN5O4, MS(ES): m / z(MH - )388.1.

[0329] Step 4: Synthesis of YK-CAP-115-PM4 Using YK-CAP-115-PM3 (4.8 g, 12.39 mmol) as a raw material, YK-CAP-115-PM4 (5.6 g, 8.12 mmol, 65.5%) was obtained by the synthesis method of YK-CAP-113-PM8. 39 H 36 FN5O6, MS(ES): m / z(MH - )690.3.

[0330] Step 5: Synthesis of YK-CAP-115-PM5 Using YK-CAP-115-PM4 (5.6 g, 8.12 mmol) as a raw material, YK-CAP-115-PM5 (5.0 g, 5.62 mmol, 69.2%) was obtained using the synthesis method for YK-CAP-113-PM9. 48 H 53 FN7O7P, MS(ES): m / z(MH - )888.4.

[0331] Step 6: Synthesis of YK-CAP-115-PM6 Using YK-CAP-115-PM5 (5.0 g, 5.62 mmol) as a raw material, YK-CAP-115-PM6 (4.9 g, 3.94 mmol, 70.2%) was obtained by the synthesis method of YK-CAP-113-PM10. 60 H 61 FN 11 O 16 P, MS(ES): m / z(MH - )1240.4.

[0332] Step 7: Synthesis of YK-CAP-115-PM7 Using YK-CAP-115-PM6 (4.9 g, 3.94 mmol) as a raw material, YK-CAP-115-PM7 (2.3 g, 2.45 mmol, 62.0%) was obtained using the synthesis method for YK-CAP-113-PM11. 39 H 43 FN 11 O 14 P, MS(ES): m / z(MH - )938.3.

[0333] Step 8: Synthesis of YK-CAP-115-PM8 Using YK-CAP-115-PM7 (2.3 g, 2.45 mmol) as a raw material, YK-CAP-115-PM8 (2.0 g, 1.86 mmol, 76.2%) was obtained by the synthesis method of YK-CAP-113-PM12. 42 H 47 FN 12 O 17 P2, MS(ES): m / z(MH - )1071.3.

[0334] Step 9: Synthesis of YK-CAP-115-PM9 Using YK-CAP-115-PM8 (2.0 g, 1.86 mmol) as a raw material, YK-CAP-115-PM9 (triethylamine salt, 720 mg, 0.89 mmol, 47.7%) was obtained using the synthesis method for YK-CAP-113-PM13. 21 H 27 FN 10 O 13 P2, MS(ES): m / z(MH - )707.1.

[0335] Step 10: Synthesis of YK-CAP-115 Using YK-CAP-115-PM9 (150 mg, 0.19 mmol) as a raw material, YK-CAP-115 (ammonium salt, 31 mg, 25.86 μmol, 14.0%) was obtained by the synthesis method of YK-CAP-113. 32 H 42 FN 15 O 23 P4, MS(ES): m / z(MH - )1146.1. 1HNMR(400MHz,D2O)δ8.44(d,J=1.4Hz,1H),8.32(s,1H),8.12(d,J=6.0Hz,2H),6.1 2(d,J=5.2Hz,1H),5.95(d,J=2.2Hz,1H),5.82(d,J=4.2Hz,1H),4.95-4.82(m,3H) ,4.80-4.73(m,2H),4.53(t,J=4.5Hz,1H),4.41-4.34(m,4H),4.21(d,J=4.1Hz,1H ),4.13(s,3H),4.04(s,1H),3.61(t,J=6.3Hz,1H),3.42(s,3H),2.92-2.81(m,1H). 31 PNMR(D2O,162MHz)δ-0.86(s,1P),-11.62(d,J=19.1Hz,1P),-12.41(d,J=17.2Hz,1P),-22.42(t,J=17.0Hz,1P).

[0336] 18. Synthesis of YK-CAP-116

[0337] [ka]

[0338] Step 1: Synthesis of YK-CAP-116-PM1 YK-CAP-113-PM4 (20.0 g, 47.54 mmol) was dissolved in acetonitrile (200 mL), and 2-iodoxybenzoic acid (16.0 g, 57.14 mmol) was added. The mixture was heated to 90°C and stirred for 4 hours. After cooling to room temperature, the reaction solution was filtered, and the filtrate was spin-dried under reduced pressure to obtain YK-CAP-116-PM1 (18.4 g, 43.95 mmol, 92.4%).

[0339] Step 2: Synthesis of YK-CAP-116-PM2 YK-CAP-116-PM1 (18.4 g, 43.95 mmol) was dissolved in dichloromethane (200 mL). The reaction mixture was cooled to -40 °C and a solution of diethylaminosulfur trifluoride (21.3 g, 132.14 mmol) in dichloromethane (40 mL) was slowly added dropwise. After the addition was complete, the mixture was slowly warmed to 0 °C and stirred for 4 h. Completion was monitored by thin-layer chromatography. The reaction mixture was quenched with saturated aqueous sodium bicarbonate (200 mL) and extracted with dichloromethane (300 mL x 3). The combined organic phases were washed with saturated aqueous NaCl, dried over anhydrous sodium sulfate, filtered, and spin-dried. The residue was purified by silica gel column chromatography (0-40% ethyl acetate / n-hexane) to give YK-CAP-116-PM2 (12.8 g, 29.05 mmol, 66.1%).

[0340] Step 3: Synthesis of YK-CAP-116-PM3 Using YK-CAP-116-PM2 (12.8 g, 29.05 mmol) as a raw material, YK-CAP-116-PM3 (9.6 g, 14.82 mmol, 51.0%) was obtained using the synthesis method for YK-CAP-113-PM6. 30 H 43 F2N5O5Si2, MS(ES): m / z (M+H + )648.3.

[0341] Step 4: Synthesis of YK-CAP-116-PM4 Using YK-CAP-116-PM3 (9.6 g, 14.82 mmol) as a raw material, YK-CAP-116-PM4 (5.2 g, 12.83 mmol, 86.6%) was obtained by the synthesis method of YK-CAP-113-PM7. 18 H 17 F2N5O4, MS(ES): m / z (M+H + )406.1.

[0342] Step 5: Synthesis of YK-CAP-116-PM5 Using YK-CAP-116-PM4 (5.2 g, 12.83 mmol) as a raw material, YK-CAP-116-PM5 (6.4 g, 9.04 mmol, 70.5%) was obtained by the synthesis method of YK-CAP-113-PM8. 39 H 35 F2N5O6, MS(ES): m / z (M+H + )708.3.

[0343] Step 6: Synthesis of YK-CAP-116-PM6 Using YK-CAP-116-PM5 (6.4 g, 9.04 mmol) as a raw material, YK-CAP-116-PM6 (5.5 g, 6.06 mmol, 67.0%) was obtained using the synthesis method for YK-CAP-113-PM9. 48 H 52 F2N7O7P, MS(ES): m / z(MH - )906.4.

[0344] Step 7: Synthesis of YK-CAP-116-PM7 Using YK-CAP-116-PM6 (5.5 g, 6.06 mmol) as a raw material, YK-CAP-116-PM7 (5.3 g, 4.21 mmol, 69.4%) was obtained by the synthesis method of YK-CAP-113-PM10. 60 H 60 F2N 11 O 16 P, MS(ES): m / z(MH - )1258.4.

[0345] Step 8: Synthesis of YK-CAP-116-PM8 Using YK-CAP-116-PM7 (5.3 g, 4.21 mmol) as a raw material, YK-CAP-116-PM8 (2.8 g, 2.92 mmol, 69.5%) was obtained using the synthesis method for YK-CAP-113-PM11. 39 H 42 F2N 11 O 14 P, MS(ES): m / z(MH - )956.3.

[0346] Step 9: Synthesis of YK-CAP-116-PM9 Using YK-CAP-116-PM8 (2.8 g, 2.92 mmol) as a raw material, YK-CAP-116-PM9 (2.2 g, 2.02 mmol, 69.0%) was obtained by the synthesis method of YK-CAP-113-PM12. 42 H 46 F2N 12 O 17 P2, MS(ES): m / z(MH - )1089.3.

[0347] Step 10: Synthesis of YK-CAP-116-PM10 Using YK-CAP-116-PM9 (2.2 g, 2.02 mmol) as a raw material, YK-CAP-116-PM10 (triethylamine salt, 580 mg, 0.70 mmol, 34.7%) was obtained by the synthesis method of YK-CAP-113-PM13. 21 H 26 F2N 10 O 13 P2, MS(ES): m / z(MH - )725.1.

[0348] Step 11: Synthesis of YK-CAP-116 Using YK-CAP-116-PM10 (150 mg, 0.18 mmol) as a raw material, YK-CAP-116 (ammonium salt, 17 mg, 13.97 μmol, 7.7%) was obtained by the synthesis method of YK-CAP-113. 32 H 41 F2N 15 O 23 P4, MS(ES): m / z(MH - )1164.1. 1HNMR(400MHz,D2O)δ8.52(d,J=1.4Hz,1H),8.45(s,1H),8.25(d,J=6.0Hz,2H),6.2 9(d,J=5.2Hz,1H),6.11(d,J=2.1Hz,1H),5.89(d,J=4.2Hz,1H),4.91-4.81(m,3H) ,4.59(t,J=4.6Hz,1H),4.42-4.31(m,4H),4.25(d,J=4.1Hz,1H),4.11(s,2H),4.0 2(s,1H),3.86-3.73(m,2H),3.63(t,J=6.3Hz,1H),3.43(s,3H),2.41-2.27(m,1H). 31 PNMR(D2O,162MHz)δ-0.82(s,1P),-11.65(d,J=19.1Hz,1P),-12.27(d,J=17.8Hz,1P),-23.88(t,J=17.3Hz,1P).

[0349] 19. Synthesis of YK-CAP-117

[0350] [ka]

[0351] Step 1: Synthesis of YK-CAP-117-PM1 Using 1,2-O-(1-methylethylidene)-4-C-[(phenylmethoxy)methyl]-3-O-(phenylmethyl)-L-lyxofuranose (10.00 g, 24.97 mmol) as a starting material, YK-CAP-117-PM1 (8.62 g, 20.80 mmol, 83.3%) was obtained via the synthetic route of YK-CAP-107-PM4.

[0352] Step 2: Synthesis of YK-CAP-117-PM2 YK-CAP-117-PM1 (8.62 g, 20.80 mmol) was dissolved in acetic acid, acetic anhydride (21.23 g, 207.95 mmol) was added, and concentrated sulfuric acid (380 μL) was slowly added dropwise. After the addition was complete, the mixture was stirred at room temperature for 16 hours. 300 mL of water was added to the reaction solution, which was then extracted with ethyl acetate. The organic phase was washed three times with saturated aqueous sodium bicarbonate until the pH became basic. The organic phase was dried and spun dry. The residue was purified by silica gel column chromatography (0-40% ethyl acetate / n-hexane) to obtain a yellow oily liquid, YK-CAP-117-PM2 (8.13 g, 17.73 mmol, 85.3%).

[0353] Step 3: Synthesis of YK-CAP-117-PM3 Using YK-CAP-117-PM2 (8.13 g, 17.73 mmol) as a raw material, YK-CAP-117-PM3 (6.24 g, 7.93 mmol, 44.7%) was obtained via the synthetic route of YK-CAP-104-PM11. 43 H 42 N6O9, MS(ES): m / z (M+H + )787.3.

[0354] Step 4: Synthesis of YK-CAP-117-PM4 YK-CAP-117-PM3 (6.24 g, 7.93 mmol) was dissolved in dichloromethane (200 mL). The reaction mixture was cooled to -40°C under a nitrogen atmosphere and a 1M solution of boron trichloride (79.3 mL, 79.30 mmol) in dichloromethane was slowly added dropwise. After the addition was complete, the reaction mixture was slowly warmed to 0°C and stirred at that temperature for 3 hours. TLC indicated complete reaction, so the mixture was cooled again to -40°C, quenched with methanol, spun dry, and left at room temperature for 24 hours. A solid was added dropwise to DCM, which precipitated out. The brown crude product was filtered and purified by HPLC to give YK-CAP-117-PM4 (1.19 g, 3.64 mmol, 45.8%). 12 H 17 N6O5, MS(ES): m / z (M+H + )328.1. YK-CAP-117-PM4: 1 HNMR (400MHz, MeOD) δ8.15(s,1H),4.51(s,1H),4.41(s,1H),4.21-4.17(m,1H),3.81(s,2H),3.44-3.31(m,2H),2.97(s,3H).

[0355] Step 5: Synthesis of YK-CAP-117-PM5 Using YK-CAP-117-PM4 (1.19 g, 3.64 mmol) as a starting material, YK-CAP-117-PM5 (triethylamine salt, 1.23 g, 2.42 mmol, 66.5%) was obtained via the synthetic route of YK-CAP-101-PM3. 12 H 18 N5O9P, MS(ES): m / z(MH - )406.1.

[0356] Step 6: Synthesis of YK-CAP-117-PM6 Using YK-CAP-117-PM5 (1.23 g, 2.42 mmol) as a starting material, YK-CAP-117-PM6 (sodium salt, 993 mg, 2.07 mmol, 85.6%) was obtained via the synthetic route of YK-CAP-101-PM4. 15 H 20 N7O8P, MS(ES): m / z(MH - )456.1.

[0357] Step 7: Synthesis of YK-CAP-117-PM7 Using YK-CAP-117-PM6 (993 mg, 2.07 mmol) as a starting material, YK-CAP-117-PM7 (triethylamine salt, 794 mg, 1.35 mmol, 65.1%) was obtained via the synthetic route of YK-CAP-101-PM5. 12 H 19 N5O 12 P2, MS(ES): m / z(MH - )486.1.

[0358] Step 8: Synthesis of YK-CAP-117-PM8 Using YK-CAP-117-PM7 (794 mg, 1.35 mmol) as a raw material, YK-CAP-117-PM8 (triethylamine salt, 520 mg, 0.86 mmol, 64.0%) was obtained via the synthetic route of YK-CAP-101-PM6. 13 H 21 N5O 12 P2, MS(ES): m / z(MH - )500.1.

[0359] Step 9: Synthesis of YK-CAP-117 Using YK-CAP-117-PM8 (150 mg, 0.25 mmol) as a starting material, YK-CAP-117 (ammonium salt, 26 mg, 20.95 μmol, 8.4%) was obtained via the synthetic route of YK-CAP-101. 34 H 47 N 15 O 25 P4, MS(ES): m / z(MH - )1188.1. 1 HNMR(400MHz,D2O)δ8.44(d,J=1.5Hz,1H),8.26(d,J=2.5Hz,2H),8.15(d,J=4.5Hz, 1H),6.31(d,J=5.2Hz,1H),5.72(d,J=2.4Hz,1H),5.51(d,J=4.6Hz,1H),4.90-4.86 (m,3H),4.60(t,J=4.7Hz,1H),4.50-4.38(m,5H),4.22(d,J=2.9Hz,1H),4.13(s,2H) ),4.05(s,2H),3.92(s,3H),3.54(t,J=7.0Hz,2H),3.44-3.31(m,3H),2.97(s,3H). 31 PNMR(D2O,162MHz)δ-0.91(s,1P),-11.22(d,J=19.1Hz,1P),-11.32(d,J=16.4Hz,1P),-22.12(t,J=17.4Hz,1P).

[0360] 20. Synthesis of YK-CAP-118

[0361] [ka]

[0362] Step 1: Synthesis of YK-CAP-118-PM1 Using 1,2-O-(1-methylethylidene)-4-C-[(phenylmethoxy)methyl]-3-O-(phenylmethyl)-L-lyxofuranose (10.00 g, 24.97 mmol) as a starting material, YK-CAP-118-PM1 (9.11 g, 22.64 mmol, 90.6%) was obtained via the synthetic route of YK-CAP-109-PM1.

[0363] Step 2: Synthesis of YK-CAP-118-PM2 Using YK-CAP-118-PM1 (9.11 g, 22.64 mmol) as a raw material, YK-CAP-118-PM2 (7.18 g, 16.08 mmol, 71.0%) was obtained via the synthetic route of YK-CAP-117-PM2.

[0364] Step 3: Synthesis of YK-CAP-118-PM3 Using YK-CAP-118-PM2 (7.18 g, 16.08 mmol) as a raw material, YK-CAP-118-PM3 (6.64 g, 8.57 mmol, 53.3%) was obtained via the synthetic route of YK-CAP-104-PM11. 42 H 39 FN6O8, MS(ES): m / z (M+H + )775.3.

[0365] Step 4: Synthesis of YK-CAP-118-PM4 Using YK-CAP-118-PM3 (6.64 g, 8.57 mmol) as a starting material, YK-CAP-118-PM4 (2.31 g, 7.33 mmol, 85.5%) was obtained via the synthetic route of YK-CAP-117-PM4. 11 H 14 FN5O5, MS(ES): m / z (M+H + )316.1. YK-CAP-118-PM4: 1 HNMR (400MHz, MeOD) δ 8.12 (s, 1H), 4.51 (s, 1H), 4.40 (s, 1H), 4.27-4.22 (m, 1H), 3.88 (s, 2H), 3.72-3.62 (m, 2H).

[0366] Step 5: Synthesis of YK-CAP-118-PM5 Using YK-CAP-118-PM4 (1.30 g, 4.12 mmol) as a starting material, YK-CAP-118-PM5 (triethylamine salt, 1.15 g, 2.32 mmol, 56.2%) was obtained via the synthetic route of YK-CAP-101-PM3. 11 H 15 FN5O8P, MS(ES): m / z(MH - )394.1.

[0367] Step 6: Synthesis of YK-CAP-118-PM6 Using YK-CAP-118-PM5 (1.15 g, 2.32 mmol) as a starting material, YK-CAP-118-PM6 (sodium salt, 870 mg, 1.86 mmol, 80.4%) was obtained via the synthetic route of YK-CAP-101-PM4. 14 H 17 FN7O7P, MS(ES): m / z(MH - )444.1.

[0368] Step 7: Synthesis of YK-CAP-118-PM7 Using YK-CAP-118-PM6 (870 mg, 1.86 mmol) as a starting material, YK-CAP-118-PM7 (triethylamine salt, 664 mg, 1.15 mmol, 61.9%) was obtained via the synthetic route of YK-CAP-101-PM5. 11 H 16 FN5O 11 P2, MS(ES): m / z(MH - )474.1.

[0369] Step 8: Synthesis of YK-CAP-118-PM8 Using YK-CAP-118-PM7 (664 mg, 1.15 mmol) as a starting material, YK-CAP-118-PM8 (triethylamine salt, 419 mg, 0.71 mmol, 61.6%) was obtained via the synthetic route of YK-CAP-101-PM6. 12 H 18 FN5O 11 P2, MS(ES): m / z(MH - )488.1.

[0370] Step 9: Synthesis of YK-CAP-118 Using YK-CAP-118-PM8 (150 mg, 0.25 mmol) as a starting material, YK-CAP-118 (ammonium salt, 33 mg, 26.86 μmol, 10.7%) was obtained via the synthetic route of YK-CAP-101. 33 H 44 FN 15 O 24 P4, MS(ES): m / z(MH - )1176.1. 1 HNMR(400MHz,D2O)δ8.37(d,J=1.2Hz,1H),8.33-8.21(m,1H),8.14(d,J=4.5Hz, 2H),6.33(d,J=5.1Hz,1H),5.61(d,J=2.3Hz,1H),5.45(d,J=4.6Hz,1H),4.92-4 .83(m,3H),4.63-4.55(m,2H),4.50-4.38(m,4H),4.31(d,J=2.9Hz,1H),4.11(s ,2H),4.01(s,2H),3.90-3.76(m,3H),3.52(t,J=7.0Hz,2H),3.46-3.34(m,3H). 31 PNMR(D2O,162MHz)δ-0.90(s,1P),-11.23(d,J=19.1Hz,1P),-11.59(d,J=16.0Hz,1P),-22.33(t,J=17.2Hz,1P).

[0371] 21. Synthesis of YK-CAP-119

[0372] [ka]

[0373] Step 1: Synthesis of YK-CAP-119-PM1 1,2-O-(1-methylethylidene)-4-C-[(phenylmethoxy)methyl]-3-O-(phenylmethyl)-L-lyxofuranose (10.0 g, 24.97 mmol) was dissolved in 50 mL of acetonitrile, and 2-iodoxybenzoic acid (10.49 g, 37.46 mmol) was added. The mixture was heated to 70°C and stirred for 2 hours. Complete reaction was monitored by TLC. The reaction was stopped, and after the reaction solution cooled to room temperature, it was filtered through diatomaceous earth, the cake was rinsed with acetonitrile (50 mL), and the filtrate was spin-dried under reduced pressure and dried in vacuo to obtain a pale yellow liquid YK-CAP-119-PM1 (10.21 g), which was used directly in the next step (yield calculated as 100%).

[0374] Step 2: Synthesis of YK-CAP-119-PM2 Using YK-CAP-119-PM1 (10.21 g, calculated based on 24.97 mmol) as a raw material, YK-CAP-119-PM2 (8.16 g, 19.41 mmol, 77.7%) was obtained via the synthetic route of YK-CAP-110-PM1.

[0375] Step 3: Synthesis of YK-CAP-119-PM3 Using YK-CAP-119-PM2 (8.16 g, 19.41 mmol) as a raw material, YK-CAP-119-PM3 (6.55 g, 14.10 mmol, 72.7%) was obtained via the synthetic route of YK-CAP-117-PM2.

[0376] Step 4: Synthesis of YK-CAP-119-PM4 Using YK-CAP-119-PM3 (6.55 g, 14.10 mmol) as a starting material, YK-CAP-119-PM4 (5.77 g, 7.28 mmol, 51.6%) was obtained via the synthetic route of YK-CAP-104-PM11. 42 H 38 F2N6O8, MS(ES): m / z (M+H + )793.3.

[0377] Step 5: Synthesis of YK-CAP-119-PM5 Using YK-CAP-119-PM4 (5.77 g, 7.28 mmol) as a starting material, YK-CAP-119-PM5 (2.16 g, 6.48 mmol, 89.0%) was obtained via the synthetic route of YK-CAP-117-PM4. 11 H 13 F2N5O5, MS(ES): m / z (M+H + )334.1. YK-CAP-119-PM5: 1 HNMR (400MHz, MeOD) δ 8.24 (s, 1H), 4.77 (s, 1H), 4.56 (s, 1H), 4.51 (s, 1H), 4.27-4.22 (m, 1H), 3.72-3.62 (m, 2H).

[0378] Step 6: Synthesis of YK-CAP-119-PM6 Using YK-CAP-119-PM5 (1.50 g, 4.50 mmol) as a raw material, YK-CAP-119-PM6 (triethylamine salt, 1.29 g, 2.51 mmol, 55.7%) was obtained via the synthetic route of YK-CAP-101-PM3. 11 H 14 F2N5O8P, MS(ES): m / z(MH - )412.1.

[0379] Step 7: Synthesis of YK-CAP-119-PM7 Using YK-CAP-119-PM6 (1.29 g, 2.51 mmol) as a raw material, YK-CAP-119-PM7 (sodium salt, 991 mg, 2.04 mmol, 81.4%) was obtained via the synthetic route of YK-CAP-101-PM4. 14 H 16F2N7O7P, MS(ES): m / z(MH - )462.1.

[0380] Step 8: Synthesis of YK-CAP-119-PM8 Using YK-CAP-119-PM7 (991 mg, 2.04 mmol) as a starting material, YK-CAP-119-PM8 (triethylamine salt, 774 mg, 1.30 mmol, 63.8%) was obtained via the synthetic route of YK-CAP-101-PM5. 11 H 15 F2N5O 11 P2, MS(ES): m / z(MH - )492.0.

[0381] Step 9: Synthesis of YK-CAP-119-PM9 Using YK-CAP-119-PM8 (774 mg, 1.30 mmol) as a starting material, YK-CAP-119-PM9 (triethylamine salt, 533 mg, 0.88 mmol, 67.3%) was obtained via the synthetic route of YK-CAP-101-PM6. 12 H 17 F2N5O 11 P2, MS(ES): m / z(MH - )506.1.

[0382] Step 10: Synthesis of YK-CAP-119 Using YK-CAP-119-PM9 (150 mg, 0.25 mmol) as a starting material, YK-CAP-119 (ammonium salt, 32 mg, 25.67 μmol, 10.3%) was obtained via the synthetic route of YK-CAP-101. 33 H 43 F2N 15 O 24 P4, MS(ES): m / z(MH - )1194.1. 1HNMR(400MHz,D2O)δ8.35(d,J=1.2Hz,1H),8.30-8.19(m,1H),8.12(d,J=4.4Hz,2 H),6.25(d,J=5.2Hz,1H),5.65(d,J=2.2Hz,1H),5.27(d,J=4.5Hz,1H),4.93-4.8 8(m,3H),4.83-4.79(s,1H),4.66-4.57(m,2H),4.51-4.42(m,5H),4.22(d,J=2.9 Hz,1H),4.04(s,2H),3.88-3.75(m,3H),3.42(t,J=7.0Hz,1H),3.36-3.31(m,3H). 31 PNMR(D2O,162MHz)δ-0.93(s,1P),-12.13(d,J=19.1Hz,1P),-13.31(d,J=16.1Hz,1P),-24.57(t,J=17.4Hz,1P).

[0383] 22. Synthesis of compound 5227:

[0384] [ka]

[0385] Starting from 5227-S, 35 mg of compound 5227 was obtained by the synthesis method of YK-CAP-117.

[0386] 23.Synthesis of CAP-2'O-ethyl

[0387] [ka]

[0388] 47 mg of CAP-2'O-ethyl was obtained by the method of WO2023025073A1.

[0389] Example 2: In vitro transcription yield and capping rate of mRNA I. Structural Differences of Capped Analogues

[0390] [Table 1] JPEG0007805500000046.jpg176170JPEG0007805500000047.jpg186170JPEG00078055000 00048.jpg186170JPEG0007805500000049.jpg168170JPEG0007805500000050.jpg117170

[0391] From Table 1, it can be seen that the chemical structures of the compounds YK-CAP-101 to 119 of the present application are partially similar to and partially very different from the mRNA capped analogs disclosed in the prior art.

[0392] 1. The first sugar ring of compounds YK-CAP-101 and YK-CAP-102 of the present application is a six-membered ring, while N7113 is a five-membered ring, and the other structures are exactly the same.

[0393] 2. The group connecting the first sugar ring and guanine in the compound YK-CAP-103 of the present application is different from that in N7113, that is, there is one more methylene group at the C1 position, but the other structures are exactly the same.

[0394] 3. Compounds YK-CAP-104 to 106 of the present application differ from N7113 in that they have two substituents at C3 of the first sugar ring, i.e., the C3 substituents of N7113 are hydroxy and hydrogen, while the C3 substituents of YK-CAP-104 are dimethylaminomethyl and fluorine, respectively, the C3 substituents of YK-CAP-105 are cyano and methyl, respectively, and the C3 substituents of YK-CAP-106 are acetamido and methyl, respectively, and the other structures are exactly the same.

[0395] 4. The C3 substituent on the first sugar ring of compounds YK-CAP-107 to 112 of the present application is different from that of N7113 and HN3002. That is, the C3 substituents of N7113 and HN3002 are hydroxyl and methoxymethyl, while the C3 substituents of YK-CAP-107 to 112 are 1-methoxyethyl, 1-acetamidoethyl, 1-fluoroethyl, difluoromethyl, N,N-diacetamido, and N,N-di-n-propylamido, and the other structures are exactly the same.

[0396] 5. The C2 substituent on the second sugar ring of the compounds YK-CAP-113 to 116 of the present application is different from that of N7113 and CAP-2'O-ethyl, i.e., the C2 substituents of N7113 and CAP-2'O-ethyl are methoxy and ethoxy, while the C2 substituents of YK-CAP-113 to 116 are methoxymethyl, acetamidomethyl, 1-fluoromethyl, and difluoromethyl, respectively, and the other structures are exactly the same.

[0397] 6. The C4 substituent on the first sugar ring of compounds YK-CAP-117 to 119 of the present application is different from that of N7113 and 5227. That is, the C4 substituents of N7113 and 5227 are hydrogen and methoxy, while the C4 substituents of YK-CAP-117 to 119 are methoxymethyl, 1-fluoromethyl, and difluoromethyl, respectively. The other structures are exactly the same.

[0398] 7. The structures of compounds YK-CAP-101 to 119 of the present application are significantly different from those of compounds 14 and m6A. The first sugar ring of compound 14 is a locked nucleic acid sugar ring, i.e., there is a methylene bridge between 2'-O and C4', while the second base, adenine, of m6A is methylated.

[0399] II. Determination of in vitro transcription yield and capping rate of mRNA 1. Experimental Method

[0400] (1) Capping synthesis was carried out using capped analogues.

[0401] First, the plasmid was linearized using a plasmid linearizing enzyme, and then the linearized plasmid was purified.

[0402] (2) In vitro transcription and synthesis of mRNA YK-CAP-101 to 119, compound 5227, and CAP-2'O-ethyl, each synthesized in Example 1, were used as capped analogs, and the reaction system was as shown in Table 2 below.

[0403] [Table 2]

[0404] The experimental procedure first calculated the volume of materials required for the reaction system, then added the sample. DNase / RNase-free water was added to the reaction system, followed by 10x buffer, NTPs, and capped analogs. Mix thoroughly and gently centrifuge. Next, nuclease inhibitors, inorganic pyrophosphatase, T7 RNA polymerase, and linear DNA template were added. Mix thoroughly and gently centrifuge. The mixture was then incubated at 37°C for 2 hours. 1 U of DNase I was added, followed by a 30-minute incubation at 37°C. The mRNA precipitate was then washed with 75% ethanol, briefly evaporated, and redissolved in DNase / RNase-free water.

[0405] (3) The transcription products were purified, and the in vitro transcription yield of mRNA was recorded.

[0406] (4) The obtained mRNA and the probe were subjected to an annealing reaction.

[0407] Anneal in a PCR machine: 95°C for 5 minutes, 65°C for 2 minutes, 55°C for 2 minutes, 40°C for 2 minutes, 22°C for 2 minutes.

[0408] (5) Pretreatment of magnetic beads and binding of probes: 100 μL of magnetic beads were taken and placed on a magnetic stand for pretreatment. 120 μL of sample was added and incubated with the magnetic bead solution at room temperature for 30 minutes, and gently mixed during incubation.

[0409] (6) The mRNA was cleaved to obtain the 5' single-stranded sequence of the mRNA bound to the probe.

[0410] 20 μL of RNase H (5 U / μL) was added and the mixture was incubated at 37°C for 3 hours, with uniform mixing every 30 minutes. After incubation, the magnetic beads were washed and added to 100 μL of 75% methanol heated to 80°C. The mixture was heated to 80°C on a hot plate and held for 3 minutes. The supernatant was then absorbed and the mixture was placed on a magnetic stand. The supernatant was then dried at room temperature for 45 minutes in an evaporative centrifuge until the volume reached 10 μL. The sample was then resuspended in 50 μL of 100 μM EDTA / 1% MeOH and immediately subjected to LC-MS analysis to confirm the RNA capping status during the transcription reaction. Because there is a large difference in molecular weight between capped and uncapped bases, the molecular weight difference can be used to determine the capping rate of mRNA transcription initiated from different capped analogs.

[0411] 2. Experimental Results The measurement results of in vitro mRNA transcription yield and capping rate showed that the ribose-modified capping analogs of the present invention have significant differences in in vitro mRNA transcription yield and capping rate.Compared with the ribose-modified capped analogs of the prior art, the ribose-modified capped analogs of the present invention have significantly improved in vitro mRNA transcription yield and capping rate.

[0412] The specific in vitro transcription yields and capping rates of mRNA are shown in Table 3.

[0413] [Table 3]

[0414] 1) The ribose-modified capped analogs of the present application exhibit significant differences in the in vitro transcription yield and capping rate of mRNA, with the in vitro transcription yield and capping rate of YK-CAP-106 to 119 mRNA both being significantly higher than those of YK-CAP-101 to 105. YK-CAP-111 exhibits the highest transcription yield and capping rate, with the transcription yield being 5.3 times that of the lowest YK-CAP-101 and the capping rate being 3.2 times that of the lowest YK-CAP-102.

[0415] As can be seen from Table 3, all of the ribose-modified capped analogs of the present application were capable of transcribing mRNA. The differences in mRNA transcription activity between different ribose-modified capped analogs were highly significant. The in vitro transcription yields of mRNA for YK-CAP-106 to 119 were all very high, exceeding 150 μg. Among them, the yields for YK-CAP-107, YK-CAP-110, YK-CAP-111, YK-CAP-117, and YK-CAP-118 reached 170.7 μg, 172.1 μg, 173.5 μg, 153.2 μg, and 172.2 μg, respectively. The highest yield was YK-CAP-111, reaching 173.5 μg (Figure 1). The lowest in vitro mRNA transcription yield was YK-CAP-101, at only 32.5 μg, and the yields of YK-CAP-102 to 105 were also very low, at 44.1 μg, 88.0 μg, 73.2 μg, and 82.3 μg, respectively. The transcription yield of YK-CAP-111 was significantly improved, being 5.3-, 3.9-, 2.0-, 2.4-, and 2.1-fold that of YK-CAP-101 to 105, respectively.

[0416] The capping rates of YK-CAP-106 to 119 were all very high, exceeding 95%. Among them, the capping rates of YK-CAP-107, YK-CAP-110, YK-CAP-111, YK-CAP-117 and YK-CAP-118 were 97.3%, 97.4%, 98.1%, 95.1% and 97.2%, respectively. The lowest was YK-CAP-111, reaching 98.1%.

[0417] The capping rate of YK-CAP-102 was lowest at only 30.2%, while that of YK-CAP-101 and YK-CAP-103 to 105 was also very low at 42.8%, 71.1%, 77.2%, and 69.3%, respectively. The capping rate of YK-CAP-111 was significantly improved, being 2.3-fold, 3.2-fold, 1.4-fold, 1.3-fold, and 1.4-fold that of YK-CAP-101 to 105, respectively (Figure 2).

[0418] 2) Compared with the ribose-modified capped analogs of the prior art, the ribose-modified capped analogs of the present application significantly improved the in vitro transcription yield and capping rate of mRNA. For example, the transcription yield of YK-CAP-111 increased by 40.8% compared with compound 14, and the capping rate increased by 36.4% compared with compound 5227.

[0419] The mRNA in vitro transcription yield and capping rate of compound 14 were 123.2 μg and 78.3%, respectively, while the mRNA in vitro transcription yield and capping rate of YK-CAP-111 in this application were significantly improved by 40.8% and 25.3%, respectively, compared with compound 14.

[0420] The mRNA in vitro transcription yield and capping rate of 5227 were 134.2 μg and 71.9%, respectively, while the mRNA in vitro transcription yield and capping rate of YK-CAP-111 in this application were significantly improved by 29.3% and 36.4%, respectively, compared with 5227.

[0421] 3) Ribose-modified capped analogs with similar structures have large differences in in vitro mRNA transcription yields and capping rates, so it is not possible to predict the in vitro mRNA transcription yields and capping rates based on their structures.

[0422] The structures of the ribose-modified capped analogs YK-CAP-117–119 designed in this application are very similar, and this series of compounds is very similar to the structure of 5227, but there were significant differences in the in vitro transcription yield and capping rate of mRNA.

[0423] For example, the compounds YK-CAP-117-119 of the present application differ from 5227 only in the C4 substituent on the first sugar ring, i.e., the C4 substituent of 5227 is methoxy, while the C4 substituents of YK-CAP-117-119 are methoxymethyl, 1-fluoromethyl, and difluoromethyl, respectively, with the other structures being identical. However, the in vitro transcription yields of YK-CAP-117-119 mRNA were increased by 14.2%, 28.3%, and 14.9%, respectively, compared with 5227, and the capping rates were increased by 32.3%, 35.2%, and 33.9%, respectively, which were significantly improved.

[0424] This indicates that ribose-modified capped analogs with similar structures do not necessarily have similar mRNA transcription activity and capping rates, and that there is a very high possibility that there may be significant differences.

[0425] The in vitro mRNA transcription yield and capping rate of the ribose-modified capped analogs YK-CAP-106-119 of the present application were significantly increased compared to YK-CAP-101-105 of the present application or to compounds 14 and 5227 of the prior art. This demonstrates that the ribose modification of YK-CAP-106-119 has an excellent anti-reverse transcription effect in the in vitro transcription of mRNA, significantly increasing the cap structure and the binding ability of capping enzymes, thereby increasing the capping rate of the transcribed mRNA.

[0426] Example 3: Preparation and characterization of lipid nanoparticles 1. Experimental Method

[0427] Cationic lipid YK-009 (Youcare Pharmaceutical Group Drug Research Institute), DSPC (AVT, Shanghai), cholesterol (AVT, Shanghai), and DMG-PEG2000 were dissolved in ethanol at a molar ratio of 49:10:39.5:1.5, and mRNA was diluted to pH 4 with 50 mM citrate buffer. Using a microfluidic device, the ethanolic lipid solution was mixed with the aqueous Fluc mRNA solution prepared with various cap structures at a volume ratio of 1:3 at a flow rate of 10 mL / min to produce LNPs with a total lipid to mRNA weight ratio of approximately 15:1. The resulting liposomes were diluted 10 times with PBS and then ultrafiltered using a 300 kDa ultrafiltration tube to remove the ethanol. Next, it was dissolved in PBS to a predetermined volume, and finally, the lipid nanoparticles were filtered through a 0.2 μm sterile filter to obtain an LNP formulation of YK-009 / DSPC / cholesterol / DMG-PEG2000 (mol ratio is 49:10:39.5:1.5) encapsulating Fluc-mRNA.

[0428] Dynamic light scattering was used to measure particle size and polydispersity index (PDI) using a Malvern laser particle size analyzer. Ten microliters of liposome solution was diluted to 1 mL with RNase-free deionized water and added to the sample pool. Measurements were repeated three times for each sample. Measurement conditions were a scattering angle of 90° and 25°C. Lipid nanoparticle encapsulation rates were measured using the QuantitRibogreen RNA Quantitation Analysis Kit (ThermoFisherScientific, UK) according to the manufacturer's instructions.

[0429] 2. Experimental Results Characterization data for specific lipid nanoparticles are shown in Table 4.

[0430] [Table 4]

[0431] Table 4 shows that the Flu mRNA transcribed by the capped analogs YK-CAP-101-119 of the present application and the capped analogs 5227, CAP-2'O-ethyl, N-7113, compound 14, HN3002, and m6A disclosed in the prior art all produced good lipid nanoparticles. The particle size of all lipid nanoparticles was in the range of 66-88 nm, the PDI value was in the range of 0.023-0.078, and the encapsulation efficiency was over 90%.

[0432] Example 4: Translation efficiency of various capped luciferase mRNAs 1. Experimental Method

[0433] (1) HEK293T cells were cultured in DMEM medium containing 10% FBS and penicillin / streptomycin at 37°C and 5% CO2.

[0434] (2) The cells in the culture dish were digested and counted, seeded into a 96-well plate at 10,000 cells / well, and cultured overnight to allow the cells to adhere to the wall.

[0435] (3) When the cell density reached approximately 80%, transfection was performed by adding 0.5 μg of mRNA sample and Lipofectamine Messenger MAX Transfection Reagent (Invitrogen) to each well. The transfection step was performed according to the manufacturer's instructions.

[0436] (4) After transfection, the cells were incubated at 37°C, 5% CO for 24 hours. The growth medium was removed from the cells and the cells were rinsed with PBS. After centrifugation to remove the PBS, 50 μL of 1× lysis buffer was added, and the cells and all liquid were transferred to a microcentrifuge tube and centrifuged.

[0437] (5) Take 20 μL of sample and add 100 μL of Dual-LumiNova equilibrated to room temperature. TM II Firefly Luciferase Detection Reagent was added and mixed properly.

[0438] (6) The plate was incubated at room temperature (approximately 25°C) for 5 minutes to stabilize the luminescence signal. Chemiluminescence was detected using a multifunctional microplate reader equipped with chemiluminescence detection capabilities, and the data were recorded.

[0439] 2. Experimental Results The relative fluorescence readings of the capped mRNA are shown in Table 5, and the relative fluorescence intensity is directly proportional to the translation efficiency of the mRNA.

[0440] [Table 5] JPEG0007805500000055.jpg65170

[0441] 1) The ribose-modified capped analogs of the present application showed significant differences in mRNA translation efficiency, with the translation efficiency of YK-CAP-106 to 119 being significantly higher than that of YK-CAP-101 to 105. YK-CAP-111 had the highest translation efficiency, 9.6 times that of YK-CAP-101, which had the lowest translation efficiency.

[0442] Table 5 shows that there are significant differences in the relative fluorescence intensities (corresponding to mRNA translation efficiency) of the ribose-modified capped analogs of the present application. YK-CAP-106 to 119 all have higher relative fluorescence intensities (ranging from 1.4 to 2.2). Among them, the relative fluorescence intensities of YK-CAP-107, YK-CAP-110, YK-CAP-111, YK-CAP-117, and YK-CAP-118 are 1.62, 1.95, 2.12, 1.82, and 1.65, respectively. The highest relative fluorescence intensity is YK-CAP-111, reaching 2.12, followed by YK-CAP-110, reaching 1.95.

[0443] The lowest relative fluorescence intensity was YK-CAP-101, at only 0.22. The relative fluorescence intensities of YK-CAP-102 to 105 were also very low, at 0.34, 0.77, 0.81, and 0.72, respectively. The relative fluorescence intensities of YK-CAP-111 were 9.6-fold, 6.2-fold, 2.8-fold, 2.6-fold, and 2.9-fold those of YK-CAP-001 to 105, respectively, and the relative fluorescence intensities of YK-CAP-110 were 8.9-fold, 5.7-fold, 2.5-fold, 2.4-fold, and 2.7-fold those of YK-CAP-001 to 105, respectively (Figure 3).

[0444] 2) Compared with the ribose-modified capped analogs of the prior art, the ribose-modified capped analogs of the present application significantly improved the mRNA translation efficiency, for example, the translation efficiency of YK-CAP-111 was 5.6-fold higher than that of m6A.

[0445] The relative fluorescence intensities (corresponding to the mRNA translation efficiency) of N-7113, HN3002, compound 14, and m6A were 1.00, 1.12, 1.13, and 0.38, respectively. The relative fluorescence intensities of YK-CAP-111 of the present application were 2.1-fold, 1.9-fold, 1.9-fold, and 5.6-fold those of N-7113, HN3002, compound 14, and m6A, respectively. The relative fluorescence intensities of YK-CAP-110 of the present application were 1.9-fold, 1.7-fold, 1.7-fold, and 5.1-fold those of N-7113, HN3002, compound 14, and m6A, respectively.

[0446] 3) Ribose-modified capped analogs with similar structures have large differences in mRNA translation efficiency, so mRNA translation efficiency cannot be predicted based on structure.

[0447] The ribose-modified capped analogs YK-CAP-117-119 designed in this application are very similar in structure to 5227, and although this series of compounds is very similar in structure to 5227, there are significant differences in mRNA translation efficiency. For example, the C4 substituent on the first sugar ring of the YK-CAP-117-119 compounds in this application is different from that of 5227. That is, the C4 substituent of 5227 is methoxy, while the C4 substituents of YK-CAP-117-119 are methoxymethyl, 1-fluoromethyl, and difluoromethyl, respectively. The rest of the structures are identical. However, the translation efficiency of YK-CAP-117-119 mRNA was significantly improved, being 2.2-fold, 2.0-fold, and 1.8-fold that of 5227, respectively.

[0448] Similarly, the structures of the ribose-modified capped analogs YK-CAP-113-116 designed in this application are very similar, and these compounds are also very similar in structure to CAP-2'O-ethyl. However, the mRNA translation efficiency differs significantly. For example, the compounds YK-CAP-113-116 of this application differ from CAP-2'O-ethyl only in the C2 substituent of the second sugar ring. That is, the C2 substituent of CAP-2'O-ethyl is ethoxy, while the C2 substituents of YK-CAP-113-116 are methoxymethyl, acetamidomethyl, 1-fluoromethyl, and difluoromethyl, respectively, while the rest of the structures are identical. However, the mRNA translation efficiency of YK-CAP-113-116 was significantly improved, being 1.4-fold, 1.3-fold, 1.5-fold, and 1.2-fold higher than that of CAP-2'O-ethyl, respectively.

[0449] This indicates that the translation efficiencies of luciferase mRNAs from ribose-modified capped analogs with similar structures are not necessarily similar, and on the contrary, there is a very high possibility that there may be significant differences.

[0450] The translation efficiencies of various capped luciferase mRNAs showed that the ribose-modified capped analogs of the present application, including YK-CAP-106-119, exhibited significantly improved mRNA translation efficiency compared to structurally similar ribose-modified capped analogs (including YK-CAP-101-105 and 5227, CAP-2'O-ethyl, N-7113, and HN3002) or structurally distinct ribose-modified capped analogs (including compound 14 and m6A). This indicates that the modified ribose in YK-CAP-106-119 readily binds to the cap-binding protein (EIF4E), thereby improving the translation efficiency of target mRNA. Furthermore, the luciferase mRNA translation efficiencies of structurally similar ribose-modified capped analogs are not necessarily similar; in fact, significant differences may exist.

[0451] Example 5. Decap enzyme stability test 1. Experimental Method

[0452] Thirty pmol of purified RNA was extracted by polyacrylamide gel electrophoresis (PAGE) and reacted with 50 μM mRNA decapping enzyme (New England Biolabs) in 1x MDE buffer at 37°C for 45 minutes. The reaction products were subjected to PAGE electrophoresis and stained with SYBR Green II (Lonza). The gel patterns were then visualized using a Typhoon FLA7000 (GE Healthcare) instrument. The ratio of the electrophoretic band intensities of capped and uncapped RNA was calculated using ImageQuant (GE Healthcare) software, and the decapping rate of the decapping enzyme was calculated.

[0453] 2. Experimental Results

[0454] [Table 6]

[0455] 1) The ribose-modified capped analogs of this application had significant differences in decap rates, with the decap rates of YK-CAP-106 to 119 being significantly lower than those of YK-CAP-101 to 105. YK-CAP-117 had the lowest decap rate, a 36.3% decrease from the highest rate of YK-CAP-104.

[0456] The data in Table 6 show that the ribose-modified capped analogs YK-CAP-101 to 119 of the present application have very different decap rates. YK-CAP-106 to 119 all have low decap rates, while YK-CAP-107, YK-CAP-110, YK-CAP-111, YK-CAP-117, and YK-CAP-118 have decap rates of 11.5%, 10.2%, 11.5%, 9.8%, and 11.9%, respectively. The lowest decap rate was YK-CAP-117, at only 9.8%, followed by YK-CAP-110 at 10.2%.

[0457] The decap rate for YK-CAP-104 was the highest, reaching 46.1%, followed by YK-CAP-101, YK-CAP-102, YK-CAP-103, and YK-CAP-105, which were also higher at 32.4%, 41.2%, 36.5%, and 33.9%. The decap rate for YK-CAP-117 was 22.6%, 31.4%, 26.7%, 36.3%, and 24.1% lower than that for YK-CAP-101-105, respectively. The decap rate for YK-CAP-110 was 22.2%, 31.0%, 26.3%, 35.9%, and 23.7% lower than that for YK-CAP-101-105, respectively (Figure 4).

[0458] 2) The ribose-modified capped analogs of the present application exhibited significantly reduced decap rates compared to prior art ribose-modified capped analogs with similar or very different structures, e.g., the decap rate of YK-CAP-117 was 34.0% lower than that of N-7113.

[0459] The decap rates of N-7113, compound 14, HN3002, and m6A were 43.8%, 26.8%, 23.3%, and 33.8%, respectively. The decap rates of YK-CAP-117 of the present application were reduced by 34.0%, 17.0%, 13.5%, and 24.0% compared to N-7113, compound 14, HN3002, and m6A, respectively. The decap rates of YK-CAP-110 were reduced by 33.6%, 16.6%, 13.1%, and 23.6% compared to N-7113, compound 14, HN3002, and m6A, respectively.

[0460] 3) Ribose-modified capped analogs with similar structures have large differences in decapping rates, making it impossible to predict the decapping rate of mRNA based on its structure.

[0461] The structures of the ribose-modified capped analogs YK-CAP-117-119 designed by the present application are very similar, and the structures of this series of compounds are also very similar to that of 5227, but the mRNA decapping rates were significantly different. For example, the compounds YK-CAP-117-119 of the present application differ from 5227 only in the C4 substituent on the first sugar ring. That is, the C4 substituent of 5227 is methoxy, while the C4 substituents of YK-CAP-117-119 are methoxymethyl, 1-fluoromethyl, and difluoromethyl, respectively, while the other structures are exactly the same. However, the mRNA decapping rates of YK-CAP-117-119 were significantly reduced, being 13.4%, 11.3%, and 11.8% lower than those of 5227, respectively.

[0462] Similarly, the structures of the ribose-modified capped analogs YK-CAP-113-116 designed by the present application are very similar, and the structures of this series of compounds are also very similar to that of CAP-2'O-ethyl. However, the mRNA decapping rates are significantly different. For example, the compounds YK-CAP-113-116 of the present application differ from CAP-2'O-ethyl only in the C2 substituent on the second sugar ring. That is, the C2 substituent of CAP-2'O-ethyl is ethoxy, while the C2 substituents of YK-CAP-113-116 are methoxymethyl, acetamidomethyl, 1-fluoromethyl, and difluoromethyl, respectively. The rest of the structures are identical. However, the mRNA decapping rates of YK-CAP-113-116 were significantly reduced compared to CAP-2'O-ethyl, with capping rates reduced by 13.0%, 14.1%, 13.0%, and 14.0%, respectively.

[0463] This demonstrates that structurally similar ribose-modified capped analogs do not necessarily have similar decap rates, but rather are very likely to have large differences.

[0464] The decap rates of the DCP2 enzyme were significantly reduced for the ribose-modified capped analogs of the present application, including YK-CAP-106-119, compared with structurally similar ribose-modified capped analogs (including YK-CAP-101-105 of the present application and 5227, CAP-2'O-ethyl, N-7413, and HN3002 of the prior art) or structurally significantly different ribose-modified capped analogs (including compound 14 and m6A). Furthermore, the decap rates of structurally similar ribose-modified capped analogs are not necessarily similar; conversely, there may be significant differences.

[0465] Example 6: Animal Experiments 1. Experimental Method

[0466] An LNP formulation containing 5 μg of Fluc-mRNA transcribed from the capped analog was intramuscularly injected into female BALB / C mice aged 4-6 weeks and weighing 17-19 g. At specific time points after administration (6 h, 12 h, 24 h, 48 h, 96 h, and 168 h), the mice were intraperitoneally injected with a fluorescent imaging substrate. After allowing the mice to move freely for 5 minutes, the total radiation intensity (corresponding to the amount of protein expressed) of the protein expressed by the mRNA delivered by the LNPs in the mice was detected using an IVIS Spectrum small animal in vivo imager.

[0467] 2. Experimental Results The detection results are shown in Table 7. In the in vivo imaging experiment of mice, the ratio of the total radiation intensity of each group of mice to the m6A group is shown in Table 7 (where the total radiation intensity is the listed value × 10 8 p / s).

[0468] [Table 7]

[0469] [Table 8]

[0470] 1) The ribose-modified capped analogs of this application showed significant differences in the total radiation intensity and duration of protein expression in mice, with YK-CAP-106-119 being significantly higher than YK-CAP-101-105. YK-CAP-110 had the highest total radiation intensity, 11.0-fold and 12.1-fold higher than the lowest YK-CAP-101 at 12 hours and 48 hours, respectively.

[0471] The data in Table 7 show that the total radiation intensity of the proteins expressed in vivo in mice for the mRNAs of different ribose-modified cap analogs of this application varies greatly. The total radiation intensity of YK-CAP-106-119 in vivo imaging in mice was all very high, with YK-CAP-110 having the highest radiation intensity at 10.55×10 at 12 h. 8 p / s, and still 2.30×10 8 p / s, followed by YK-CAP-111, with 9.34 × 10 8 p / s, reaching 1.95 × 10 8 p / s was reached.

[0472] YK-CAP-101 had the lowest total radiation intensity, 0.96 × 10 8 p / s, and only 0.19 × 10 8 p / s. The total radiation intensity of YK-CAP-110 was 11.0 times that of YK-CAP-101 at 12 hours and 12.1 times that at 48 hours. The total radiation intensity of YK-CAP-111 was 9.7 times that of YK-CAP-101 at 12 hours and 10.3 times that at 48 hours.

[0473] 2) The present ribose-modified capped analogs significantly improved the total radiation intensity and duration of protein expression in mice compared with similar or very different structurally related ribose-modified capped analogs of the prior art. For example, the total radiation intensity of YK-CAP-110 was 5.0-fold higher than that of m6A at 12 hours and 4.4-fold higher at 48 hours.

[0474] The total radiation intensity for N-7113 and m6A in 12 hours was 4.33 × 10 8 p / s and 2.12×10 8 p / s, and 0.49 × 10 8 p / s and 0.52×10 8 It was p / s.

[0475] The total radiation intensity of YK-CAP-110 of the present application was 2.4 and 5.0 times that of N-7113 and m6A, respectively, at 12 h, and 4.7 and 4.4 times that of N-7113 and m6A, respectively, at 48 h.

[0476] The total radiation intensity of YK-CAP-111 of the present application was 2.2 and 4.4 times that of N-7113 and m6A, respectively, at 12 h, and 4.0 and 3.8 times that at 48 h.

[0477] 3) Ribose-modified capped analogs with similar structures have significantly different total radiation intensities and durations of protein expression in the mouse body from mRNA, making it impossible to predict the total radiation intensities and durations of protein expression in the mouse body from mRNA based on their structures.

[0478] The structures of the ribose-modified capped analogs YK-CAP-117-119 designed by the present application are very similar, and the structures of these compounds are also very similar to that of 5227. However, there were significant differences in the total radiation intensity of the proteins expressed in vivo from the mRNA of mice. For example, the C4 substituent on the first sugar ring of the present application's compounds YK-CAP-117-119 differs from that of 5227. That is, the C4 substituent of 5227 is methoxy, while the C4 substituents of YK-CAP-117-119 are methoxymethyl, 1-fluoromethyl, and difluoromethyl, respectively. The other structures are identical. However, the total radiation intensity of the proteins expressed in vivo from the mRNA of YK-CAP-117-119 was significantly improved at 6 h, being 3.8-fold, 3.0-fold, and 3.1-fold that of 5227, respectively.

[0479] Similarly, the structures of the ribose-modified capped analogs YK-CAP-113-116 designed by the present application are very similar, and these compounds are also very similar in structure to CAP-2'O-ethyl. However, the total radiation intensities of the proteins expressed in vivo in mice from mRNAs were significantly different. For example, the compounds YK-CAP-113-116 of the present application differ from CAP-2'O-ethyl only in the C2 substituent on the second sugar ring. That is, the C2 substituent of CAP-2'O-ethyl is ethoxy, while the C2 substituents of YK-CAP-113-116 are methoxymethyl, acetamidomethyl, 1-fluoromethyl, and difluoromethyl, respectively. The rest of the structures are identical. However, the total radiation intensities of the proteins expressed in vivo in mice from YK-CAP-114 and YK-CAP-116 mRNAs were significantly improved at 6 h, being 1.6- and 1.7-fold higher than those of CAP-2'O-ethyl, respectively.

[0480] This indicates that the total radiation intensity and duration of protein expressed in mice from Fluc mRNA produced with a ribose-modified capped analogue, which has a similar structure, are not necessarily similar, and in fact, there is a very high possibility that there are significant differences.

[0481] Animal experiments have shown that the ribose-modified capped analogs of the present application, including YK-CAP-106-119, significantly improve the total radiation intensity and duration of protein expression in mice compared to structurally similar ribose-modified capped analogs (including YK-CAP-101-105 of the present application, and 5227, N-7413 and CAP-2'O-ethyl of the prior art) or structurally different ribose-modified capped analogs (m6A).

[0482] In vivo experiments further demonstrated that mRNA transcribed by the YK-CAP-106-119 gene of the present application can be effectively delivered to the body by the LNP delivery vector and expressed efficiently and continuously.Furthermore, the total radiation intensity and duration of protein expression in mice of Fluc mRNA produced with structurally similar ribose-modified capped analogs were not necessarily similar; in fact, there was a very high possibility of significant differences.

[0483] In summary, the ribose-modified capped analogs YK-CAP-106-119 of the present application significantly improved the in vitro mRNA transcription yield, capping rate, mRNA translation efficiency, stability of decapping enzymes, and the amount and duration of protein expression in animals compared to prior art ribose-modified capped analogs (including 5227, CAP-2'O-ethyl, N-7113, compound 14, HN3002, and m6A). This indicates that the YK-CAP-106-119 cap structure provided by the present invention can significantly improve the resistance of modified ribose structures to decapping enzymes and the binding affinity for capping enzymes, providing a novel and efficient ribose-modified cap structure for in vitro mRNA transcription.

[0484] 1. The chemical structures of the compounds of the present application are very similar, and some of these compounds have similar structures and some are significantly different structures compared to mRNA capped analogs disclosed in the prior art.

[0485] 1) The first sugar ring of compounds YK-CAP-101 and YK-CAP-102 of the present application is a six-membered ring, while compound N7113 is a five-membered sugar ring, and the other structures are exactly the same.

[0486] 2) The group connecting the first sugar ring and guanine in the compound YK-CAP-103 of the present application was different from that in N7113, that is, there was one more methylene at the C1 position, but the other structures were exactly the same.

[0487] 3) Compounds YK-CAP-104 to 106 of the present application have two substituents at C3 of the first sugar ring, which differ from N7113. That is, the C3 substituents of N7113 are hydroxy and hydrogen, while the C3 substituents of YK-CAP-104 are dimethylaminomethyl and fluorine, respectively, the C3 substituents of YK-CAP-105 are cyano and methyl, respectively, and the C3 substituents of YK-CAP-106 are acetamido and methyl, respectively, and the other structures are exactly the same.

[0488] 4) The C3 substituents of the first sugar ring of compounds YK-CAP-107 to 112 of the present application are different from those of N7113 and HN3002. That is, the C3 substituents of N7113 and HN3002 are hydroxy and methoxymethyl, while the C3 substituents of YK-CAP-107 to 112 are 1-methoxyethyl, 1-acetamidoethyl, 1-fluoroethyl, difluoromethyl, N,N-diacetamido, and N,N-di-n-propylamido, respectively. The other structures are exactly the same.

[0489] 5) The C2 substituents on the second sugar ring of the compounds YK-CAP-113 to 116 of the present application are different from those of N7113 and CAP-2'O-ethyl. That is, the C2 substituents of N7113 and CAP-2'O-ethyl are methoxy and ethoxy, while the C2 substituents of YK-CAP-113 to 116 are methoxymethyl, acetamidomethyl, 1-fluoromethyl, and difluoromethyl, respectively, and the other structures are exactly the same.

[0490] 6) The C4 substituent on the first sugar ring of compounds YK-CAP-117 to 119 of the present application is different from that of N7113 and 5227. That is, the C4 substituents of N7113 and 5227 are hydrogen and methoxy, while the C4 substituents of YK-CAP-117 to 119 are methoxymethyl, 1-fluoromethyl, and difluoromethyl, respectively, and the other structures are exactly the same.

[0491] 7) The structures of compounds YK-CAP-101 to 119 of the present application are significantly different from those of compounds 14 and m6A. The first sugar ring of compound 14 is a locked nucleic acid sugar ring, i.e., there is a methylene bridge between 2'-O and C4', while the second base, adenine, of m6A is methylated.

[0492] 2. The ribose-modified capped analogs of the present application showed significant differences in the in vitro transcription yield and capping rate of mRNA. Compared with the ribose-modified capped analogs of the prior art, the ribose-modified capped analogs of the present application showed significantly improved in vitro transcription yield and capping rate of mRNA.

[0493] 1) The ribose-modified capped analogs of this application showed significant differences in the in vitro transcription yield and capping rate of mRNA, with the in vitro transcription yield and capping rate of YK-CAP-106-119 mRNA both being higher than those of YK-CAP-101-105. The transcription yield and capping rate of YK-CAP-111 were the highest, with a transcription yield 5.3 times that of the lowest YK-CAP-101 and a capping rate 3.2 times that of the lowest YK-CAP-102.

[0494] 2) The ribose-modified capped analogs of the present application have significantly improved in vitro mRNA transcription yield and capping rate compared to prior art ribose-modified capped analogs. For example, the transcription yield of YK-CAP-111 is 40.8% higher than that of compound 14, and the capping rate is 36.4% higher than that of 5227.

[0495] 3) Ribose-modified capped analogs with similar structures showed significant differences in in vitro mRNA transcription yield and capping rate. For example, the in vitro transcription yield of YK-CAP-117~119 mRNA was improved by 14.2%, 28.3%, and 14.9%, respectively, compared to 5227, and the capping rate was improved by 32.3%, 35.2%, and 33.9%, respectively.

[0496] 3. The ribose-modified capped analogs of the present application showed significant differences in mRNA translation efficiency. Compared with the ribose-modified capped analogs of the prior art, the ribose-modified capped analogs of the present application showed significantly improved mRNA translation efficiency.

[0497] 1) The ribose-modified capped analogs of the present application showed significant differences in mRNA translation efficiency, with the translation efficiency of YK-CAP-106 to 119 being significantly higher than that of YK-CAP-101 to 105. YK-CAP-111 had the highest translation efficiency, 9.6 times that of YK-CAP-101, the lowest.

[0498] 2) The ribose-modified capped analogs of the present application significantly improved mRNA translation efficiency compared with ribose-modified capped analogs of similar or very different structures in the prior art. For example, the translation efficiency of YK-CAP-111 mRNA was 5.6-fold higher than that of m6A.

[0499] 3) The mRNA translation efficiencies of structurally similar ribose-modified capped analogs differed significantly. For example, the translation efficiencies of YK-CAP-117~119 mRNA were 2.2-, 2.0-, and 1.8-fold higher than those of 5227, respectively.

[0500] 4. The ribose-modified capped analogs of the present application exhibited significantly different decap rates. Compared with ribose-modified capped analogs of similar or very different structures in the prior art, the ribose-modified capped analogs of the present application exhibited significantly reduced decap rates.

[0501] 1) The ribose-modified capped analogs of the present application had significant differences in decap rates, with the decap rates of YK-CAP-106 to 119 being significantly lower than those of YK-CAP-101 to 105. YK-CAP-117 had the lowest decap rate, 36.3% lower than that of YK-CAP-104, which had the highest decap rate.

[0502] 2) The ribose-modified capped analogs of the present application all exhibited significantly reduced decap rates compared to ribose-modified capped analogs of similar or very different structures in the prior art, for example, the decap rate of YK-CAP-117 was 34.0% lower than that of N-7113.

[0503] 3) Ribose-modified capped analogs with similar structures showed significant differences in decapping rates. For example, the decapping rates of YK-CAP-113~116 mRNA were reduced by 13.0%, 14.1%, 13.0%, and 11.0%, respectively, compared to CAP-2'O-ethyl.

[0504] 5. The ribose-modified capped analogs of the present application showed significant differences in the total radiation intensity (corresponding to the protein expression level) and duration of protein expression in mice from mRNA. Compared with ribose-modified capped analogs of the prior art with similar or very different structures, the ribose-modified capped analogs of the present application showed significantly improved protein expression levels and duration in mice from mRNA.

[0505] 1) The ribose-modified capped analogs of the present application showed significant differences in the total radiation intensity and duration of protein expressed in the mouse mRNA, with YK-CAP-106-119 being significantly higher than YK-CAP-101-105. YK-CAP-110 had the highest total radiation intensity, 11.0-fold and 12.1-fold higher than the lowest YK-CAP-101 at 12 hours and 48 hours, respectively.

[0506] 2) The present ribose-modified capped analogs significantly improved the total radiation intensity and duration of protein expression in mice compared with similar or very different structurally related ribose-modified capped analogs in the prior art. For example, the total radiation intensity of YK-CAP-110 was 5.0-fold higher than that of m6A at 12 hours and 4.4-fold higher at 48 hours.

[0507] 3) Ribose-modified capped analogs with similar structures showed significant differences in the total radiation intensity and duration of protein expression in mice. For example, the total radiation intensity of the protein expressed in mice from YK-CAP-117~119 mRNA was 3.8-fold, 3.0-fold, and 3.1-fold higher than that of 5227 at 6 hours.

[0508] The applicant declares that the present invention has described the ribose-modified caprylic acid analogues of the present invention and their uses through the above examples, but the present invention is not limited to the above examples, that is, the present invention does not mean that the present invention must be implemented by relying on the above examples. Those skilled in the art should understand that any improvements to the present invention, equivalent substitution of raw materials and addition of auxiliary ingredients to the products of the present invention, and selection of specific methods are all within the protection scope and disclosure of the present invention.

Claims

1. 1. A ribose-modified capped analog or a pharmaceutically acceptable salt thereof, characterized in that the ribose-modified capped analog has a structure of the following formula: YK-CAP-106, YK-CAP-107, YK-CAP-108, YK-CAP-109, YK-CAP-110, YK-CAP-111, YK-CAP-112, YK-CAP-113, YK-CAP-114, YK-CAP-115, YK-CAP-116, YK-CAP-117, YK-CAP-118, or YK-CAP-119. 【Chemistry 1】 【Chemistry 2】 【Transformation 3】 【Chemistry 4】

2. An RNA molecule, characterized in that it comprises the ribose-modified capped analog of claim 1 as a capped structure.

3. A pharmaceutical composition comprising the RNA molecule of claim 2.

4. The pharmaceutical composition of claim 3, further comprising at least one RNA delivery agent.

5. The pharmaceutical composition of claim 4, wherein the at least one RNA delivery agent comprises at least one cationic lipid.

6. The cationic lipid is selected from one or more of the following compounds: (1) A compound represented by formula (II) or a pharmaceutically acceptable salt thereof, wherein G １ is C １～６ alkylene, and G ２ is C ２～８ alkylene, and G ３ is C １～３ alkylene, and L １ is C ６～１５ is a linear alkyl; L ２ is C １２～２５ is a branched alkyl 【Transformation 5】 (2) A compound represented by formula (III) or a pharmaceutically acceptable salt thereof, wherein: １ is C ２～８ alkylene, and G ２ is C ２～８ alkylene, and L １ is —C(O)O— or —OC(O)—, and L ２ is —C(O)O— or —OC(O)—, and R １ is C ６～２５ is a straight or branched chain alkyl; R ２ is C ６～２５ is a straight or branched chain alkyl; G ３ is HO(CH ２ ) ２ - or HO(CH ２ ) ３ - and G ４ is HO (CH ２ ) ２ - or HO(CH ２ ) ３ - and L is -(CH ２ ) ２ - or - (CH ２ ) ３ - or - (CH ２ ) ４ -is, 【Transformation 6】 (3) A compound represented by formula (IV) or a pharmaceutically acceptable salt thereof, wherein G １ is C １～６ alkylene, and G ２ is C ２～８ alkylene, and R １ is C ６～２０ is a straight or branched chain alkyl; R ２ is C １２～２５ is a branched alkyl; G ３ is HO (CH ２ ) ２ N (CH ３ ) (CH ２ ) ２ -, HO(CH ２ ) ２ N (CH ２ CH ３ ) (CH ２ ) ２ -, (HO(CH ２ ) ２ ) ２ N (CH ２ ) ２ -, CH ３ O (CH ２ ) ２ N (CH ３ ) (CH ２ ) ２ -, (CH ３ ) ２ N (CH ２ ) ３ SC(O)O(CH ２ ) ２ -, (CH ３ ) ２ N (CH ２ ) ３ SC(O)-, CH ３ NH (CH ２ ) ２ N (CH ３ ) (CH ２ ) ２ - or CH ３ CH ２ NH (CH ２ ) ２ -is, 【Transformation 7】 (4) A compound represented by formula (V) or a pharmaceutically acceptable salt thereof, wherein G １ is C １～８ alkylene, and G ２ is C ２～８ alkylene, and R １ is C ６～２５ is a straight or branched chain alkyl; R ２ is C １２～２５ is a straight or branched chain alkyl; G ３ is HO (CH ２ ) ２ N (R ３ ) CH ２ CH(OH)CH ２ -, where R ３ Ha-CH ３ , -CH ２ CH ３ or -CH ２ CH ２ OH, 【Transformation 8】 (5) A compound represented by formula (VI) or a pharmaceutically acceptable salt thereof, wherein G １ and G ２ are each independently an unsubstituted C ６ ~C １０ alkylene, and G ３ is unsubstituted C １ ~C １２ alkylene, and R １ and R ２ are each independently C ６ ~C ２４ Alkyl or C ６ ~C ２４ alkenyl, and R ３ Ha-OR ５ , -C(=O)OR ４ , -OC(=O)R ４ or -NR ５ C(=O)R ４ and R ４ is C １ ~C １２ is hydrocarbyl, and R ５ is H or C １ ~C ６ is a hydrocarbyl, 【Chemistry 9】 (6) A compound represented by formula (VII) or a pharmaceutically acceptable salt thereof, wherein R ４ Ha-(CH ２ ) ｎ Q, where Q is —OH or —CN, and n is 1, 2, or 3; 【Chemistry 10】 (7) A compound represented by formula (VIII) or a pharmaceutically acceptable salt thereof: 【Chemistry 11】 The pharmaceutical composition according to claim 5.

7. 7. The pharmaceutical composition of claim 6, wherein the cationic lipid is selected from one or a combination of at least two of YK-009, YK-401, YK-305, ALC-0315, SM-102, or DLIN-MC3-DMA. 【Chemistry 12】

8. 8. The pharmaceutical composition according to claim 7, wherein the cationic lipid is YK-009.

9. The pharmaceutical composition of claim 5 , wherein the at least one RNA delivery agent further comprises at least one neutral lipid.

10. 10. The pharmaceutical composition according to claim 9, wherein the neutral lipid comprises one or a combination of at least two of phosphatidylcholine, phosphatidylethanolamine, sphingomyelin, ceramide, or sterol.

11. The neutral lipid is 1,2-dilinoleoyl-sn-glycero-3-phosphocholine, 1,2-dimyristoyl-sn-glycero-phosphocholine, 1,2-dioleoyl-sn-glycero-3-phosphocholine, 1,2-dipalmitoyl-sn-glycero-3-phosphocholine, 1,2-distaloyl-sn-glycero-3-phosphocholine, 1,2-diundecanoyl-sn-glycero-phosphocholine, 1-palmitoyl-2-oleoyl-sn-glycero-3-phosphocholine, 1,2-di-O-octadecenyl-sn-glycero-3-phosphocholine, 1-oleoyl-2-cholesterylhemisuccinoyl-sn-glycero-3-phosphocholine, 1-hexadecyl-sn-glycero-3-phosphocholine, 1,2-dilinolenoyl- sn-glycero-3-phosphocholine, 1,2-diarachidonoyl-sn-glycero-3-phosphocholine, 1,2-didocosahexaenoyl-sn-glycero-3-phosphocholine, 1,2-dioleoyl-sn-glycero-3-phosphoethanolamine, 1,2-diphytanoyl-sn-glycero-3-phosphoethanolamine, 1,2-distearoyl-sn-glycero-3-phosphoethanolamine, 1,2-dilinoleoyl-sn-glycero-3-phosphoethanolamine, 1,2-dilinolenoyl-sn-glycero-3-phosphoethanolamine, 1,2-diarachidonoyl-sn-glycero-3-phosphoethanolamine, 1,2-didocosahexaenoyl-sn-glycero-3-phosphoethanolamine, 1,The pharmaceutical composition according to claim 10, characterized in that the phosphatidylcholine is selected from one or a combination of at least two of 2-dioleoyl-sn-glycero-3-phospho-rac-(1-glycerol) sodium salt, dipalmitoylphosphatidylglycerol, palmitoyloleylphosphatidylethanolamine, distearoylphosphatidylethanolamine, dipalmitoylphosphatidylethanolamine, dimyristoylphosphatidylethanolamine, 1-stearoyl-2-oleoyl-sn-glycero-3-phosphoethanolamine, 1-stearoyl-2-oleoyl-phosphatidylcholine, sphingomyelin, phosphatidylcholine, phosphatidylethanolamine, phosphatidylserine, phosphatidylinositol, phosphatidic acid, palmitoyloleoylphosphatidylcholine, lysophosphatidylcholine, and lysophosphatidylethanolamine.

12. 12. The pharmaceutical composition according to claim 11, wherein the neutral lipid is DOPE and / or DSPC.

13. 10. The pharmaceutical composition of claim 9, wherein the at least one RNA delivery agent further comprises a structured lipid.

14. 14. The pharmaceutical composition according to claim 13, characterized in that the structured lipids are selected from one or a combination of at least two of cholesterol, non-sterols, sitosterol, ergosterol, campesterol, stigmasterol, brassicasterol, tomatine, ursolic acid, α-tocopherol, or corticosteroids.

15. 15. The pharmaceutical composition according to claim 14, characterized in that the structured lipid is cholesterol.

16. The pharmaceutical composition of claim 4, wherein the at least one RNA delivery agent further comprises a polymer-conjugated lipid.

17. 17. The pharmaceutical composition according to claim 16, wherein the polymer-conjugated lipid is selected from one or a combination of at least two of distearoylphosphatidylethanolamine-polyethylene glycol 2000, dimyristoylglycero-3-methoxypolyethylene glycol-2000, and methoxypolyethylene glycol ditetradecylacetamide.

18. The pharmaceutical composition according to any one of claims 3 to 17, characterized in that the pharmaceutical composition further comprises one or at least two cell-penetrating peptides.

19. (1) The ribose-modified capped analog of claim 1 or a pharmaceutically acceptable salt thereof; (2) a nucleoside triphosphate molecule and an RNA polymerase; A kit comprising:

20. The kit comprises an RNase inhibitor, inorganic pyrophosphatase, Mg ２＋ 20. The kit of claim 19, further comprising one or a combination of at least two of the following: a soluble soluble protein, a crowding agent, or a buffer.

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