Antiviral and antibacterial n4-hydroxycytidine derivative, and use thereof and preparation method therefor

Abstract

Disclosed in the present invention are a compound derivative represented by formula I or an isotopically labeled compound thereof, or an optical isomer, geometric isomer, tautomer or mixture of isomer thereof, or a pharmaceutically acceptable salt thereof. On the basis of the application of a dual-protection strategy, diverse carrier fragments are introduced at the 5-hydroxyl group and hydroxylamine group of the N4-hydroxycytidine molecule to develop an N4-hydroxycytidine prodrug. Such a modification enhances the affinity for a pulmonary cell membrane (phospholipid membrane), rendering the physicochemical properties of the drug suitable for the physiological environment of lung tissue. Therefore, the compound of the present invention has a broad application prospect in the aspect of broad-spectrum anti-respiratory viruses.

Description

Antiviral antibacterial n4-hydroxycytidine derivatives, uses thereof and methods of preparation TECHNICAL FIELD

[0001] The present application belongs to the field of pharmacy, and particularly relates to a nucleoside N4-hydroxycytidine derivative or an isotopically labeled compound thereof, or an optical isomer, a geometric isomer, a tautomer or an isomer mixture thereof, or a pharmaceutically acceptable salt thereof, and a use and a preparation method thereof. BACKGROUND

[0002] Respiratory viral infections seriously threaten human health. Since the Spanish flu in 1918 caused about 1 billion people to be infected and nearly 40 million people to die, the outbreaks of H1N1, avian influenza, SARS-COV-1, MERs-COV, and the like have seriously threatened human health and caused huge social and economic losses. In particular, the outbreak of coronavirus (SARS-COV-2) in December 2019 has lasted for more than four years, and new variants of the SARS-COV-2 virus are still emerging. There is still a very realistic need to study broad-spectrum antiviral drugs.

[0003] N4-hydroxycytidine (NHC) is a broad-spectrum antiviral compound discovered in the mid-20th century. After entering the host cell, it is converted into a nucleotide analogue in the form of triphosphate, which acts as a substrate for triphosphate cytidine and triphosphate uridine in viral RNA, and is inserted as an error base during viral RNA replication, causing viral protein translation with the accumulation of mutations, and ultimately blocking viral replication, thereby exerting antiviral activity. However, N4-hydroxycytidine isobutyrate has a large polarity and poor membrane permeability. Therefore, in order to improve the balance of lipophilicity and hydrophilicity of the compound, the oral absorption and utilization degree are enhanced by increasing the liposolubility, and the isobutyric acid prodrug of NHC, Molnupiravir (EIDD-2801), was launched in 2019. Its chemical name is ((2S, 3R, 4S, 5S)-3, 4-dihydroxy-5-(4-(hydroxyamino)-2-oxopyrimidin-1 (2H)-yl)tetrahydrofuran-2-yl) methyl isobutyric acid. It has been proven to effectively inhibit MERS-CoV, SARS-CoV, and SARS-CoV-2 (including remdesivir-resistant strains) replication in human cell cultures and animal models, and has broad-spectrum antiviral activity against influenza viruses and various coronaviruses.

[0004] In previous studies (CN 114805458 B), the inventors found that lauroyl prodrugs (compound 19), myristoyl prodrugs (compound 20) and palmitoyl prodrugs (compound 21) greatly improve the biofilm permeability of EIDD-2801 by introducing long-chain alkyl groups on the hydroxylamine in the N4-hydroxycytidine isobutyric acid ester molecule of EIDD-2801. However, there are differences in cytotoxicity of different carbon chains, and the safety of carriers and prodrugs has not been elucidated. SUMMARY

[0005] Acute respiratory infections caused by coronaviruses and influenza viruses mainly occur in the respiratory tract, and oral administration has the disadvantages of slow onset, low bioavailability, and many adverse reactions, so there is an urgent need to develop new dosage forms to solve this problem. Inhalation administration is an effective way to improve the drug concentration in lung tissue, but lung tissue has a double blood circulation system, rich blood flow, fast blood flow, and good lung cell permeability, which makes it easy for drugs to diffuse to the whole body after inhalation administration, affecting the drug concentration in the target organ of respiratory viruses, and thus reducing the efficacy. Therefore, in order to achieve long-term residence of drugs in lung tissue, the present application aims to design N4-hydroxycytidine isobutyric acid ester as a prodrug, introduce a variety of carrier fragments on the hydroxylamine in the molecule, improve the affinity of lung cell membrane (phospholipid membrane), and make the physicochemical properties of the drug suitable for the requirements of the physiological environment of lung tissue. Therefore, the compounds of the present application have broad application prospects in the field of broad-spectrum antiviral. It is also unexpectedly found that the prodrug compounds provided by the present application have unexpected antibacterial activity, and the difference in antibacterial activity is related to the introduction of liposoluble carriers in the prodrug.

[0006] In addition to the influence of the carrier of the N4-hydroxycytidine prodrug on the lipid-water partition coefficient of the compound, thereby affecting the membrane permeability and bioavailability of the drug, the present application further found that different prodrug compounds have unexpected differences in antiviral activity, antibacterial activity and toxicity of the prodrug. Therefore, according to one aspect of the present application, one object of the present application is to provide a compound derivative represented by Formula I or an isotopically labeled compound thereof, or an optical isomer, geometric isomer, tautomer or isomer mixture thereof, or a pharmaceutically acceptable salt thereof:

[0007] wherein R1 is substituted or unsubstituted C2-C 10 alkyl, substituted or unsubstituted C 12 alkyl, substituted or unsubstituted C 14 alkyl, substituted or unsubstituted C 16 -C 27 alkyl, substituted or unsubstituted C2-C 27 alkyl, substituted or unsubstituted C2-C 27alkenyl, substituted or unsubstituted C3-C6cycloalkyl, wherein "substituted" means that said group contains 1 to 3 R a substituents selected from the group consisting of deuterium, hydroxy, amino, cyano, nitro, halogen, C1-C5alkoxy, C1-C5alkoxycarbonyl, C3-C6cycloalkyl, -NHCOOR a substituents selected from the group consisting of deuterium, hydroxy, amino, cyano, nitro, halogen, C1-C5alkoxy, C1-C5alkoxycarbonyl, C3-C6cycloalkyl, -NHCOOR a1 , C6-C 10 aryl, saturated or unsaturated three to six membered heterocyclyl containing 1 to 3 heteroatoms selected from N, O, S, three to six membered heteroaryl containing 1 to 3 heteroatoms selected from N, O, S, steroid structure unsubstituted or substituted by 1 to 2 R a2 substituents selected from the group consisting of deuterium, hydroxy, amino, cyano, nitro, halogen, C1-C5alkoxy, C1-C5alkoxycarbonyl, C3-C6cycloalkyl, -NHCOOR a1 substituents selected from the group consisting of deuterium, hydroxy, amino, cyano, nitro, halogen, C1-C5alkoxy, C1-C5alkoxycarbonyl, C3-C6cycloalkyl, -NHCOOR a2 selected from the group consisting of hydroxy, C2-C6ester group;

[0008] R2is substituted or unsubstituted C1-C6alkyl, wherein "substituted" means that said group contains 1 to 3 R b substituents selected from the group consisting of deuterium, hydroxy, amino, cyano, nitro, halogen, C1-C5alkoxy, C1-C5alkoxycarbonyl, C3-C6cycloalkyl, -NHCOOR b substituents selected from the group consisting of deuterium, hydroxy, amino, cyano, nitro, halogen, C1-C5alkoxy, C1-C5alkoxycarbonyl, C3-C6cycloalkyl, -NHCOOR

[0009] R1is preferably substituted or unsubstituted C4-C 10 alkyl, substituted or unsubstituted C 12 alkyl, substituted or unsubstituted C 14 alkyl, substituted or unsubstituted C 16 -C 20 alkyl, substituted or unsubstituted C3-C 17 alkenyl, substituted or unsubstituted C3-C 17 alkynyl, substituted or unsubstituted C5-C6cycloalkyl, wherein "substituted" means that said group contains 1 or 2 R a substituents selected from the group consisting of deuterium, hydroxy, amino, cyano, nitro, halogen, C1-C5alkoxy, C1-C5alkoxycarbonyl, C3-C6cycloalkyl, -NHCOOR a substituents selected from the group consisting of deuterium, hydroxy, amino, cyano, nitro, halogen, C1-C5alkoxy, C1-C5alkoxycarbonyl, C3-C6cycloalkyl, -NHCOOR a1 , C6-C 10 aryl, saturated or unsaturated three to six membered heterocyclyl containing 1 to 3 heteroatoms selected from N, O, S, three to six membered heteroaryl containing 1 to 3 heteroatoms selected from N, O, S, steroid structure unsubstituted or substituted by 1 to 2 R a2 substituents selected from the group consisting of deuterium, hydroxy, amino, cyano, nitro, halogen, C1-C5alkoxy, C1-C5alkoxycarbonyl, C3-C6cycloalkyl, -NHCOOR a1 substituents selected from the group consisting of deuterium, hydroxy, amino, cyano, nitro, halogen, C1-C5alkoxy, C1-C5alkoxycarbonyl, C3-C6cycloalkyl, -NHCOOR a2 selected from the group consisting of hydroxy, C2-C6ester group;

[0010] More preferably, R1 is a substituted or unsubstituted C7-C 10 Alkyl, substituted or unsubstituted C 12 Alkyl, substituted or unsubstituted C 14 Alkyl, substituted or unsubstituted C 16 -C 17 Alkyl, substituted or unsubstituted C8-C 17 Alkenyl, substituted or unsubstituted C8-C 17 Alkynyl, substituted or unsubstituted C3-C6 cycloalkyl, wherein the "substituted" means that the group contains 1 or 2 R a The substituent, R a Selected from deuterium, hydroxyl, amino, cyano, nitro, halogen, methoxy, ethoxy, propoxy, methoxycarbonyl, ethoxycarbonyl, propoxycarbonyl, cyclopentyl, cyclohexyl, -NHCOOR a1 , phenyl, naphthyl, a saturated or unsaturated five- to six-membered heterocyclic group containing 1 or 2 heteroatoms selected from N, O, and S, a five- to six-membered heteroaryl group containing 1 or 2 heteroatoms selected from N, O, and S, unsubstituted or replaced by 1 or 2 R a2 Substituted steroid structure, the R a1 is selected from methyl, ethyl, n-propyl, isopropyl, n-butyl, isobutyl, tert-butyl, said R a2 Selected from hydroxyl, CH3COO-, CH3CH2COO-, CH3CH2CH2COO-;

[0011] More preferably, the substituent R a wherein the residue is unsubstituted or is substituted with 1 or 2 R a2 The substituted steroid is selected from the following structures:

[0012] Preferably, R2 is a substituted or unsubstituted C1-C4 alkyl group, wherein the "substituted" means that the group contains 1 or 2 R b substituted, the R b is selected from deuterium, hydroxy, amino, cyano, nitro, halogen, C1-C3 alkoxy, C1-C3 alkyl substituted by halogen, and C3-C6 cycloalkyl;

[0013] More preferably, R2 is selected from methyl, ethyl, n-propyl, isopropyl, n-butyl, isobutyl, tert-butyl, n-pentyl, isopentyl, n-hexyl, cyclopropyl, cyclobutyl, cyclopentyl, cyclohexyl.

[0014] More preferably, R2 is isopropyl.

[0015] The compound of Formula I according to the present application or its isotopically labeled compound, or its optical isomer, geometric isomer, tautomer or mixture of isomers, or its pharmaceutically acceptable salt is selected from the following structures:

[0016] Another aspect of the present application provides a method for preparing the compound of Formula I according to the present application or its isotopically labeled compound, or its optical isomer, geometric isomer, tautomer or mixture of isomers, or its pharmaceutically acceptable salt, which can be carried out according to the contents disclosed in the prior art in combination with the conventional techniques in the art, as an example, the method shown in Reaction Scheme 1 or Reaction Scheme 2 can be used:

[0017] The method of Reaction Scheme 1 comprises the following steps:

[0018] i) protecting, substituting and deprotecting uridine to form compound 5;

[0019] ii) selectively esterifying compound 5 to form compound 6;

[0020] iii) hydroxylaminating compound 6 to form compound 9;

[0021] iv) oxyacylating the hydroxylamine group of compound 9 to form the compound of Formula I.

[0022] In a specific embodiment, in step i), trimethylsilyl chloride is used as the protecting group, and after chlorination by phosphorus oxychloride in an ice bath, triazole substitution is carried out, and glacial acetic acid is used to deprotect the group, and the reaction temperature is 0-50°C.

[0023] In a specific embodiment, in step ii), selective esterification is carried out using acyl chloride compound R2(=O)Cl with N,N-dimethylacrylurea as the base, and the reaction temperature is 0-50°C.

[0024] In which the substituent R2in acyl chloride compound R2(=O)Cl is defined as the same as the definition in Formula I above.

[0025] In a specific embodiment, in step iii), hydroxylamination is carried out using hydroxylamine hydrochloride and a sodium bicarbonate aqueous solution, and the reaction temperature is 0-50°C.

[0026] In the specific embodiment, in step iv), EDCI is 1-ethyl-(3-dimethylaminopropyl) carbodiimide hydrochloride, DMAP is dimethylaminopyridine, DCM is chloromethane, EDCI is a condensing agent, and DCC dicyclohexylcarbodiimide, DIC diisopropylcarbodiimide, or HATU, HBTU can also be used instead; the reaction temperature is 0-50°C.

[0027] As shown in the above Reaction Scheme 2, the method comprises the following steps:

[0028] i) uridine is protected, substituted, and deprotected to generate compound 5;

[0029] ii) compound 5 is acetonide-protected and esterified to generate compound 7;

[0030] iii) compound 7 is hydroxylated to generate compound 8;

[0031] iv) compound 8 is deprotected to generate compound 9;

[0032] v) the hydroxylamine group of compound 9 is oxyacylated to generate the compound shown in formula I.

[0033] In the specific embodiment, in step i), trimethylchlorosilane is used as a protecting group, and after chlorination by phosphorus oxychloride in an ice bath, triazole substitution is performed, and glacial acetic acid is used to deprotect the group, and the reaction is performed at room temperature.

[0034] In the specific embodiment, in step ii), 2,2-dimethoxypropane is used for acetonide protection in the presence of sulfuric acid as a catalyst, and then esterification is performed by an acid anhydride compound (R2(C=O))2O, and the reaction is performed at room temperature.

[0035] The substituent R2 in the acid anhydride compound (R2(C=O))2O is defined in the same manner as in formula I above.

[0036] In the specific embodiment, in step iii), hydroxylamine hydrochloride and an aqueous solution of sodium bicarbonate are used for hydroxylamine, and the reaction is performed at room temperature.

[0037] In the specific embodiment, in step iv), sulfuric acid methanol aqueous solution is used for deprotection.

[0038] In the specific embodiment, in step v), the condensing agent EDCI is 1-ethyl-(3-dimethylaminopropyl) carbodiimide hydrochloride, DMAP is dimethylaminopyridine, DCM is chloromethane, and EDCI can also be replaced by DCC dicyclohexylcarbodiimide, DIC diisopropylcarbodiimide, or HATU, HBTU; the reaction temperature is 0-50°C.

[0039] In the reaction scheme, the definition of the substituent R1 in the carboxylic acid compound R1(C=O)OH is the same as that in the above Formula I.

[0040] For example, the carboxylic acid compound is selected from octanoic acid, DL-thioctic acid, decanoic acid, octadecenoic acid, linoleic acid, 4-methyloctanoic acid, undecanoic acid, undecenoic acid, palmitoleic acid, azelaic acid monomethyl ester, cyclopentane propionic acid, 6-phenylhexanoic acid, 10-undecynoic acid, 2-methylheptanoic acid, 4-ethyloctanoic acid, trans-4-butylcyclohexanecarboxylic acid, cyclohexylbutyric acid, BOC-11-aminoundecanoic acid prodrug, BOC-8-aminooctanoic acid, R-(+)-thioctic acid, chenodeoxycholic acid, ursodeoxycholic acid, lithocholic acid, deoxycholic acid, and the like.

[0041] wherein the chenodeoxycholic acid, ursodeoxycholic acid, lithocholic acid, deoxycholic acid, and the like are used to prepare a prodrug of N4-hydroxycytidine isobutyrate containing a steroid structure, such as Compound 0939, Compound 0941, Compound 0943, Compound 0945.

[0042] In the above preparation method, acetylation protection of the steroid 3-hydroxyl group can be involved, and the protection method is as follows in Reaction Scheme 3

[0043] wherein the starting material acid chenodeoxycholic acid, ursodeoxycholic acid, lithocholic acid, or deoxycholic acid is dissolved in a solvent, 4-dimethylaminopyridine is added, acetic anhydride is added with stirring, and stirring is performed at room temperature for 3 hours. The reaction solution is washed with 1 mol / L HCl (3 x 30 ml), saturated sodium bicarbonate solution, and saturated sodium chloride solution, respectively, the organic layer is concentrated, and the obtained residue is purified by column chromatography to obtain a white solid. The white solid is directly condensed with N4-hydroxycytidine isobutyrate, and column chromatography is performed to obtain the target product.

[0044] According to one aspect of the present application, one object of the present application is to provide a use of a compound represented by Formula I or an isotopically labeled compound thereof, or an optical isomer, a geometric isomer, a tautomer, or an isomer mixture thereof, or a pharmaceutically acceptable salt thereof, as a prodrug of N4-hydroxycytidine isobutyrate.

[0045] Another aspect of the present application is to provide a pharmaceutical composition comprising a therapeutically effective amount of the compound represented by Formula I or an isotopically labeled compound thereof, or an optical isomer, a geometric isomer, a tautomer, or an isomer mixture thereof, or a pharmaceutically acceptable salt thereof, and a pharmaceutically acceptable excipient.

[0046] Yet another aspect of the present application is to provide use of the compound of Formula I or an isotopically-labeled compound thereof, or an optical isomer, a geometric isomer, a tautomer, or a mixture of isomers thereof, or a pharmaceutically acceptable salt thereof, in the manufacture of a broad-spectrum antiviral or antibacterial medicament.

[0047] Preferably, the broad-spectrum antiviral or antibacterial medicament is for use in treating or preventing Gram-positive bacterial infection, including but not limited to Staphylococcus, Streptococcus, Pneumococcus, Bacillus anthracis, Corynebacterium diphtheriae, Clostridium tetani, etc.

[0048] Preferably, the broad-spectrum antiviral or antibacterial medicament is formulated into an inhalant, a patch, a transdermal, an oral immediate-release formulation, a sustained-release controlled-release formulation, a pill, a capsule, a granule, etc.

[0049] Preferably, the inhalant includes a dry powder inhalant, a solution inhalant, a mixed-rotation inhalant, etc.

[0050] Yet another aspect of the present application is to provide a method for treating or preventing a disease caused by viral or bacterial infection, the method comprising administering to a subject a therapeutically effective amount of the compound of Formula I or an isotopically-labeled compound thereof, or an optical isomer, a geometric isomer, a tautomer, or a mixture of isomers thereof, or a pharmaceutically acceptable salt thereof or the pharmaceutical composition according to the present application. BRIEF DESCRIPTION OF DRAWINGS

[0051] In order to more clearly illustrate the specific embodiments of the present application or the technical solutions in the prior art, the following will briefly introduce the drawings needed to be used in the specific embodiments or prior art description. Obviously, the drawings described below are some embodiments of the present application, and other drawings can also be obtained by those skilled in the art without creative labor on the basis of these drawings.

[0052] Figure 1 is the Beas-2B cell toxicity experiment result of N4-hydroxycytidine isobutyrate and its prodrugs. DETAILED DESCRIPTION

[0053] Hereinafter, the present application will be described in detail. Before proceeding with the description, it should be understood that the terms used in the specification and the appended claims should not be construed as being limited to the general and dictionary meanings and should be construed as having meanings and concepts corresponding to the technical aspects of the present application based on the principle that the inventor is allowed to define appropriate terms in order to best explain the application. Therefore, the description presented herein is merely a preferred example in all aspects and is not intended to limit the scope of the application, and it should be understood that other equivalents or modifications thereof can be obtained by those skilled in the art without departing from the spirit and scope of the present application.

[0054] As used herein, the terms "comprises," "comprising," "includes," "including," "has," "having," "contains," "containing," or any other similar term are intended to be open-ended. For example, a composition or article that comprises elements A, B, and C should be construed to mean that elements A, B, and C are present, but not excluding elements D and E. Further, unless otherwise specified, the use of "or" is to be interpreted as inclusive "or," meaning A or B or both. For example, the phrase "A or B" means "A or B or both." Also, unless otherwise specified, the use of "and" is to be interpreted as descriptive "and" rather than conjunctive "and." For example, the phrase "A and B" means "A and B, but not necessarily both." Moreover, the use of the term "including" as well as other forms such as "include," "includes," "comprise," "comprises," "comprising," "contain," "contains," "containing," "characterized by," "comprised of," and "comprises essentially of," are not to be construed as limiting. Rather, these terms are to be construed as meaning "comprising." Further, unless otherwise specified, the use of the term "about" is to be construed as meaning "approximately," "substantially," or "essentially."

[0055] All features or conditions described herein in terms of a numerical range or a percentage range are intended to be merely for convenience and brevity in providing some guidance as to the scope of the described features or conditions. Thus, descriptions in terms of a numerical range or a percentage range are to be construed as having specifically disclosed and encompassing all possible subranges and individual numerical values within the range, particularly integer values. For example, a range of "1 to 8" is to be construed as having specifically disclosed all subranges, particularly subranges defined by all integer values, such as 1 to 7, 2 to 8, 2 to 6, 3 to 6, 4 to 8, 3 to 8, and the like, and as having specifically disclosed individual values within the range, such as 1, 2, 3, 4, 5, 6, 7, 8, and the like. The foregoing interpretation is to apply to all descriptions of ranges, whether broad or narrow, throughout the specification, unless otherwise indicated.

[0056] If a numerical range or other numerical values or parameters are expressed herein as a range, a preferred range or a series of upper and lower limits, it is to be understood that every numerical value or parameter within the range or the series of upper and lower limits is specifically contemplated, even if not explicitly listed. Also, it is to be understood that the description herein of any range or numerical value is intended to serve as a parameter that can be combined with other parameters to define a new range or numerical value, even if not explicitly listed.

[0057] As used herein, numerical values are to be understood as having the precision of the number of significant figures in the numerical value. For example, the number 40.0 is to be understood as encompassing the range from 39.50 to 40.49.

[0058] In the event that Markush groups or optionals are used in describing the features or examples of the application herein, one of skill in the art will understand that subgroups of all the elements in the Markush group or any individual element in the Markush group or optionals list can also be used to describe the application. For example, if X is described as "selected from the group consisting of X1, X2, and X3," the proposition that X is X1and / or X2and / or X3has been fully described. Further, in the event that Markush groups or optionals are used in describing the features or examples of the application herein, one of skill in the art will understand that subgroups of all the elements in the Markush group or any combination of individual elements in the Markush group or optionals list can also be used to describe the application. Accordingly, for example, if X is described as "selected from the group consisting of X1, X2, and X3," and Y is described as "selected from the group consisting of Y1, Y2, and Y3," the proposition that X is X1or X2or X3and Y is Y1or Y2or Y3has been fully described.

[0059] Definitions

[0060] The compounds described herein can contain one or more asymmetric centers and thus can exist in various isomeric forms, e.g., enantiomeric and / or diastereomeric forms. For example, the compounds described herein can be in the form of an individual enantiomer, diastereomer, or geometric isomer, or can be in the form of a mixture of stereoisomers, including racemic mixtures and mixtures enriched in one or more stereoisomer. Isomers can be separated from mixtures by methods known to those skilled in the art, including chiral high pressure liquid chromatography (HPLC) and formation and crystallization of chiral salts; or, preferably, isomers can be prepared by asymmetric synthesis. See, e.g., Jacques et al., Enantiomers, Racemates and Resolutions (Wiley Interscience, New York, 1981); Wilen et al., Tetrahedron 33:2725 (1977); Eliel, Stereochemistry of Carbon Compounds (McGraw-Hill, NY, 1962); and Wilen, Tables of Resolving Agents and Optical Resolutions, p. 268 (E. L. Eliel, ed., University of Notre Dame Press, Notre Dame, IN 1972). The present disclosure additionally encompasses the compounds described herein as individual isomers substantially free of other isomers, or as mixtures of various isomers.

[0061] When a range of values is listed, every value and sub-range within the range is intended to be included. For example, a range of "1 to 10" is intended to include any sub-range between (and including) the minimum and maximum values; e.g., 1-6.1, 2.3-8.1, 4.5- 7.5, etc. 1-6 " is intended to encompass C1, C2, C3, C4, C5, C6, C 1-6 , C 1-5 , C 1-4 , C 1-3 , C 1-2 , C 2-6 , C 2-5 , C 2-4 , C 2-3 , C 3-6 , C 3-5 , C 3- 4, C 4-6 , C 4-5 , and C 5-6 .

[0062] The term "alkyl" refers to a straight-chain or branched-chain saturated hydrocarbon group having from 1 to 10 carbon atoms ("C 1-10 alkyl"). In some embodiments, the alkyl group has from 1 to 9 carbon atoms ("C 1-9 alkyl"). In some embodiments, the alkyl group has from 1 to 8 carbon atoms ("C 1-8 alkyl"). In some embodiments, the alkyl group has from 1 to 7 carbon atoms ("C 1-7 alkyl"). In some embodiments, the alkyl group has from 1 to 6 carbon atoms ("C 1-6 alkyl"). In some embodiments, the alkyl group has from 1 to 5 carbon atoms ("C 1-5 alkyl"). In some embodiments, the alkyl group has from 1 to 4 carbon atoms ("C 1-4 alkyl"). In some embodiments, the alkyl group has from 1 to 3 carbon atoms ("C 1-3 alkyl"). In some embodiments, the alkyl group has from 1 to 2 carbon atoms ("C 1-2 alkyl"). In some embodiments, the alkyl group has 1 carbon atom ("C1alkyl"). In some embodiments, the alkyl group has from 2 to 6 carbon atoms ("C 2-6 alkyl"). C 1-6Examples of alkyl groups include methyl (Ci), ethyl (C2), propyl (C3) (e.g., n-propyl, isopropyl), butyl (C4) (e.g., n-butyl, t-butyl, sec-butyl, isobutyl), pentyl (C5) (e.g., n-pentyl, 3-pentyl, neopentyl, 3-methyl-2-butyl, t-pentyl), and hexyl (C6) (e.g., n-hexyl). Additional examples of alkyl groups include n-heptyl (C7), n-octyl (C8), and the like. Unless otherwise indicated, each instance of an alkyl group is independently unsubstituted (“unsubstituted alkyl”) or substituted (e.g., with one or more halogen, such as F) (“substituted alkyl”). In certain embodiments, an alkyl group is an unsubstituted C 1- 10 unsubstituted C 1-6 alkyl (e.g., unsubstituted C 1-10 alkyl (e.g., unsubstituted C 1-6 alkyl (e.g., unsubstituted C

[0063] “Alkenyl” refers to a straight or branched hydrocarbon group having from 2 to 20 carbon atoms, one or more carbon-carbon double bonds and no triple bonds (“C 2-20 alkenyl”). The carbon-carbon double bond(s) can be located at any position on the carbon chain that results in an alkene chemically, in some embodiments, an alkenyl group has 2 to 10 carbon atoms (“C 2-10 alkenyl”). In some embodiments, an alkenyl group has 2 to 9 carbon atoms (“C 2-9 alkenyl”). In some embodiments, an alkenyl group has 2 to 8 carbon atoms (“C 2-8 alkenyl”). In some embodiments, an alkenyl group has 2 to 7 carbon atoms (“C 2-7 alkenyl”). In some embodiments, an alkenyl group has 2 to 6 carbon atoms (“C 2-6 alkenyl”). In some embodiments, an alkenyl group has 2 to 5 carbon atoms (“C 2-5 alkenyl”). In some embodiments, an alkenyl group has 2 to 4 carbon atoms (“C 2-4 alkenyl”). In some embodiments, an alkenyl group has 2 to 3 carbon atoms (“C 2-3 alkenyl”). In some embodiments, an alkenyl group has 2 carbon atoms (“C2alkenyl”). The one or more carbon-carbon double bond(s) can be internal (e.g., in 2-butenyl) or terminal (e.g., in 1-butenyl). C 2-4 Examples of alkenyl groups include ethenyl (C2), 1-propenyl (C3), 2-propenyl (C3), 1-butenyl (C4), 2-butenyl (C4), butadienyl (C4), and the like. C 2-6 Examples of alkenyl groups include the foregoing C 2-4Alkenyl and pentenyl (C5), pentadienyl (C5), hexenyl (C6), and the like. Additional examples of alkenyl groups include heptenyl (C7), octenyl (C8), octatrienyl (C8), and the like. Unless otherwise specified, each instance of an alkenyl group is independently optionally substituted, i.e., unsubstituted (an “unsubstituted alkenyl”) or substituted (a “substituted alkenyl”) with one or more substituents. In certain embodiments, an alkenyl is an unsubstituted C 2-10 Alkenyl. In certain embodiments, an alkenyl is a substituted C 2-10 Alkenyl. In alkenyl groups, C=C double bonds for which stereochemistry is not specified (e.g., -CH=CHCH3or ) can be (E)- or (Z)-double bonds.

[0064] “Alkynyl” refers to a straight or branched hydrocarbon group (“C 2-20 alkynyl”) of from 2 to 20 carbon atoms having one or more carbon-carbon triple bonds and optionally one or more double bonds. The carbon-carbon triple bond(s) can be located at any position(s) of the carbon chain that the chemical structure allows, in some embodiments, an alkynyl group has 2 to 10 carbon atoms (“C 2-10 alkynyl”). In some embodiments, an alkynyl group has 2 to 9 carbon atoms (“C 2-9 alkynyl”). In some embodiments, an alkynyl group has 2 to 8 carbon atoms (“C 2-8 alkynyl”). In some embodiments, an alkynyl group has 2 to 7 carbon atoms (“C 2-7 alkynyl”). In some embodiments, an alkynyl group has 2 to 6 carbon atoms (“C 2-6 alkynyl”). In some embodiments, an alkynyl group has 2 to 5 carbon atoms (“C 2-5 alkynyl”). In some embodiments, an alkynyl group has 2 to 4 carbon atoms (“C 2-4 alkynyl”). In some embodiments, an alkynyl group has 2 to 3 carbon atoms (“C 2-3 alkynyl”). In some embodiments, an alkynyl group has 2 carbon atoms (“C2alkynyl”). The one or more carbon-carbon triple bonds can be internal (such as in 2-butynyl) or terminal (such as in 1-butynyl). C 2-4 Examples of alkynyl groups include, but are not limited to, ethynyl (C2), 1-propynyl (C3), 2-propynyl (C3), 1-butynyl (C4), 2-butynyl (C4), and the like. C 2-6 Examples of alkenyl groups include the above-described C 2-4alkynyl and pentynyl (C5), hexynyl (C6), and the like. Additional examples of alkynyl include heptynyl (C7), octynyl (C8), and the like. Unless otherwise indicated, each instance of an alkynyl group is independently optionally substituted, i.e., unsubstituted (“unsubstituted alkynyl”) or substituted (a “substituted alkynyl”) with one or more substituents. In certain embodiments, an alkynyl group is an unsubstituted C 2-10 alkynyl. In certain embodiments, an alkynyl group is a substituted C 2-10 alkynyl.

[0065] “Cycloalkyl” refers to a radical of a non-aromatic ring hydrocarbon having from 3 to 10 ring carbon atoms (“C 3-10 carbocyclyl”) and zero heteroatoms in the non-aromatic ring system. In some embodiments, a carbocyclyl group has 3 to 8 ring carbon atoms (“C 3-8 carbocyclyl”). In some embodiments, a carbocyclyl group has 3 to 7 ring carbon atoms (“C 3-7 carbocyclyl”). In some embodiments, a carbocyclyl group has 3 to 6 ring carbon atoms (“C 3-6 carbocyclyl”). In some embodiments, a carbocyclyl group has 5 to 10 ring carbon atoms (“C 5-10 carbocyclyl”). Exemplary C 3-6 carbocyclyl groups include, but are not limited to, cyclopropyl (C3), cyclopropenyl (C3), cyclobutyl (C4), cyclobutenyl (C4), cyclopentyl (C5), cyclopentenyl (C5), cyclohexyl (C6), cyclohexenyl (C6), cyclohexadienyl (C6), and the like. Exemplary C 3-8 carbocyclyl groups include, but are not limited to, the above-mentioned C 3-6 carbocyclyl groups as well as cycloheptyl (C7), cycloheptenyl (C7), cycloheptadienyl (C7), cycloheptatrienyl (C7), cyclooctyl (C8), cyclooctenyl (C8), bicyclo[2.2.1]heptyl (C7), bicyclo[2.2.2]octyl (C8), and the like. Exemplary C 3-10 carbocyclyl groups include, but are not limited to, the above-mentioned C 3- 8carbocyclyl groups as well as cyclononyl (C9), cyclononenyl (C9), cyclodecyl (C 10 ), cyclodecenyl (C 10 ), octahydro-lH-indenyl (C9), decahydronaphthyl (C 10 ), spiro[4.5]decyl (C 10) and the like. As shown in the foregoing examples, in certain embodiments, carbocyclyl is monocyclic ("monocyclic carbocyclyl") or contains fused, bridged, or spiro ring systems, such as bicyclic ring systems ("bicyclic carbocyclyl") and can be saturated or can be partially unsaturated. "Carbocyclyl" also includes ring systems in which the point of attachment of the carbocyclyl as defined above is to a ring carbon of one or more aryl or heteroaryl groups, and in this case the number of carbons refers to the number of carbons in the carbocyclyl ring system. Unless otherwise specified, each instance of a carbocyclyl group is independently optionally substituted, i.e., unsubstituted ("unsubstituted carbocyclyl") or substituted ( "substituted carbocyclyl") with one or more substituents. In certain embodiments, a carbocyclyl group is unsubstituted C 3-10 carbocyclyl. In certain embodiments, a carbocyclyl group is substituted C 3-10 carbocyclyl.

[0066] In some embodiments, "carbocyclyl" is a monocyclic saturated carbocyclyl group having 3 to 10 ring carbon atoms ("C 3-10 cycloalkyl"). In some embodiments, a cycloalkyl group has 3 to 8 ring carbon atoms ("C 3-8 cycloalkyl"). In some embodiments, a cycloalkyl group has 3 to 6 ring carbon atoms ("C 3-6 cycloalkyl"). In some embodiments, a cycloalkyl group has 5 to 6 ring carbon atoms ("C 5-6 cycloalkyl"). In some embodiments, a cycloalkyl group has 5 to 10 ring carbon atoms ("C 5-10 cycloalkyl").C 5-6 Examples of cycloalkyl groups include cyclopentyl (C5) and cyclohexyl (C6).C 3-6 Examples of cycloalkyl groups include the foregoing C 5-6 cycloalkyl groups as well as cyclopropyl (C3) and cyclobutyl (C4).C 3-8 Examples of cycloalkyl groups include the foregoing C 3-6 cycloalkyl groups as well as cycloheptyl (C7) and cyclooctyl (C8). Unless otherwise specified, each instance of a cycloalkyl group is independently unsubstituted ("unsubstituted cycloalkyl") or substituted ( "substituted cycloalkyl") with one or more substituents. In certain embodiments, a cycloalkyl group is unsubstituted C 3-10 cycloalkyl. In certain embodiments, a cycloalkyl group is substituted C 3-10 cycloalkyl.

[0067] “Heterocyclyl” refers to a group of 3- to 10-membered nonaromatic ring systems having ring carbon atoms and 1-4 ring heteroatoms, wherein each heteroatom is independently selected from nitrogen, oxygen, sulfur, boron, phosphorus, and silicon (“3-10 membered heterocyclyl”). In heterocyclyl groups containing one or more nitrogen atoms, the point of attachment can be a carbon atom or a nitrogen atom, as valence permits. Heterocyclyl groups can be monocyclic (“monocyclic heterocyclyl”) or fused, bridged, or spiro ring systems, e.g., bicyclic (“bicyclic heterocyclyl”), and can be saturated or can be partially unsaturated. Heterocyclyl bicyclic ring systems can contain one or more heteroatoms in one or both rings. “Heterocyclyl” also includes ring systems in which a heterocycle as defined above is fused with one or more carbocyclyl groups at a ring carbon of the carbocyclyl or heterocycle, or a heterocycle as defined above is fused with one or more aryl or heteroaryl groups at a ring heteroatom of the heterocycle, and in this case the number of ring members continues to refer to the number of ring members in the heterocycle. Unless otherwise specified, each instance of a heterocyclyl group is independently optionally substituted, i.e., unsubstituted (“unsubstituted heterocyclyl”) or substituted with one or more substituents (“substituted heterocyclyl”). In certain embodiments, a heterocyclyl group is an unsubstituted 3-10 membered heterocyclyl. In certain embodiments, a heterocyclyl group is a substituted 3-10 membered heterocyclyl.

[0068] In some embodiments, a heterocyclyl group is a 5-10 membered nonaromatic ring system having ring carbon atoms and 1-4 ring heteroatoms, wherein each heteroatom is independently selected from nitrogen, oxygen, sulfur, boron, phosphorus, and silicon (“5-10 membered heterocyclyl”). In some embodiments, a heterocyclyl group is a 5-8 membered nonaromatic ring system having ring carbon atoms and 1-4 ring heteroatoms, wherein each heteroatom is independently selected from nitrogen, oxygen, and sulfur (“5-8 membered heterocyclyl”). In some embodiments, a heterocyclyl group is a 5-6 membered nonaromatic ring system having ring carbon atoms and 1-4 ring heteroatoms, wherein each heteroatom is independently selected from nitrogen, oxygen, and sulfur (“5-6 membered heterocyclyl”). In some embodiments, a 5-6 membered heterocyclyl group has 1-3 ring heteroatoms selected from nitrogen, oxygen, and sulfur. In some embodiments, a 5-6 membered heterocyclyl group has 1-2 ring heteroatoms selected from nitrogen, oxygen, and sulfur. In some embodiments, a 5-6 membered heterocyclyl group has 1 ring heteroatom selected from nitrogen, oxygen, and sulfur.

[0069] “Aryl” refers to a group of a monocyclic or polycyclic (e.g., bicyclic or tricyclic) 4n+2 aromatic ring system (e.g., having 6, 10, or 14 π electrons shared in a cyclic array) having 6-14 ring carbon atoms and zero heteroatoms provided in the aromatic ring system (“C 6-14 In some embodiments, an aryl group has 6 ring carbon atoms (“C6 aryl”; e.g., phenyl). In some embodiments, an aryl group has 10 ring carbon atoms (“C 10In some embodiments, an aryl group has 14 ring carbon atoms ("C 14 "Aryl"; for example, anthracenyl). "Aryl" also includes ring systems in which an aryl ring as defined above is fused to one or more carbocyclyl or heterocyclyl groups, wherein the radical or point of attachment is on the aromatic ring, and in such cases, the number of carbon atoms continues to refer to the number of carbon atoms in the aromatic ring system. Unless otherwise specified, each instance of aryl is independently optionally substituted, i.e., unsubstituted (an "unsubstituted aryl") or substituted (a "substituted aryl") with one or more substituents. In certain embodiments, aryl is unsubstituted C 6-14 In certain embodiments, aryl is a substituted C 6-14 Aryl.

[0070] "Aralkyl" is a subset of alkyl and aryl and refers to an optionally substituted alkyl group substituted with an optionally substituted aryl group. In certain embodiments, the aralkyl group is an optionally substituted benzyl group. In certain embodiments, the aralkyl group is a benzyl group. In certain embodiments, the aralkyl group is an optionally substituted phenethyl group. In certain embodiments, the aralkyl group is a phenethyl group.

[0071] "Heteroaryl" refers to a group of a 5-10 membered monocyclic or bicyclic 4n+2 aromatic ring system (e.g., having 6 or 10 π electrons shared in a cyclic array) having ring carbon atoms and 1-4 ring heteroatoms provided in the aromatic ring system, wherein each heteroatom is independently selected from nitrogen, oxygen, and sulfur ("5-10 membered heteroaryl"). In heteroaryl groups containing one or more nitrogen atoms, the point of attachment can be a carbon atom or a nitrogen atom as valence permits. Heteroaryl bicyclic ring systems can contain one or more heteroatoms in one or both rings. "Heteroaryl" includes ring systems in which a heteroaryl ring as defined above is fused to one or more carbocyclyl or heterocyclyl groups, wherein the point of attachment is on the heteroaryl ring, and in such cases, the number of ring members continues to refer to the number of ring members in the heteroaryl ring system. "Heteroaryl" also includes ring systems in which a heteroaryl ring as defined above is fused to one or more aryl groups, wherein the point of attachment is on the aryl or heteroaryl ring, and in this case, the number of ring members refers to the number of ring members in the fused (aryl / heteroaryl) ring system. For bicyclic heteroaryl groups in which one ring contains no heteroatoms (e.g., indolyl, quinolyl, carbazolyl, etc.), the point of attachment can be on either ring, i.e., the ring with the heteroatom (e.g., 2-indolyl) or the ring without heteroatoms (e.g., 5-indolyl).

[0072] "Halogen" includes a halogen atom selected from F, Cl, Br and I.

[0073] The following examples are merely illustrative of the present application and do not in any way limit the scope of the application. Any modification obvious to those skilled in the art, without departing from the spirit and scope of the present application, shall fall within the scope of the present application. Unless otherwise specified, the reagents and instruments used in the following examples are commercially available products.

[0074] In addition, unless otherwise specified, the biochemical reagents used in this application are commercial products, such as those purchased from Anjieke Chemicals, Beijing Inokai Technology Co., Ltd. or Beijing Jin Ruimei Fragrance Technology Co., Ltd., with analytical grade, without further treatment for direct application.

[0075] Examples

[0076] Example 1: Preparation of octanoic acid prodrug (compound 0836)

[0077] First step: preparation of 1-((2R,3R,4S,5R)-3,4-dihydroxy-5-(hydroxymethyl)tetrahydrofuran-2-yl)-4-(1H-1,2,4-triazol-1-yl)pyrimidin-2(1H)-yl)(5)

[0078] After drying a 500 ml three-necked flask, uridine (30.00 g, 122.85 mmol), anhydrous acetonitrile (225 ml) and N-methylpyrrolidine (192.0 ml, 1.843 mol) were added, and TMSCl (78.0 ml, 614.25 mmol) was slowly added dropwise under nitrogen protection and 0°C ice bath, and stirred at room temperature for 1 h. Then, phosphorus oxychloride (22.5 ml, 245.7 mmol) was slowly added dropwise under 0°C ice bath, and stirred for 20 min under ice bath after the addition was completed. Triazole (84.60 g, 1.23 mol) was added to the reaction solution under 0°C ice bath, and stirred for 1 h under ice bath and 2 h at room temperature. After the reaction was completed, the reaction solution was poured into a prepared 600 ml saturated sodium chloride solution to quench the reaction, extracted with dichloromethane (200 ml) for three times, combined the organic phase, dried with anhydrous sodium sulfate, filtered, and concentrated to obtain an orange-red oil. Methyl acetate (v / v = 10:1) mixed solvent (120 mL) was added, stirred at room temperature overnight, filtered, and dried to obtain a light yellow solid 30.24 g with a yield of 83.4%.

[0079] Second step: preparation of methyl ((2R,3S,4R,5R)-3,4-dihydroxy-5-(2-oxo-4-(1H-1,2,4-triazol-1-yl)pyrimidin-1(2H)-yl)tetrahydrofuran-2-yl)isobutyrate (6)

[0080] To a solution of 1-((2R,3R,4S,5R)-3,4-dihydroxy-5-(hydroxymethyl)tetrahydrofuran- 2-yl)-4-(1H-1,2,4-triazol-1-yl)pyrimidin-2(1H)-yl)(5) (10 g, 33.87 mmol) in a mixture of 1,4-dioxane (40 ml) and N,N-dimethylformamide (20 ml), N,N- dimethylpropenylurea was added followed by slow dropwise addition of isobutyryl chloride (6.72 g, 67.74 mmol) at room temperature. The reaction was stirred for 8 h. The reaction mixture was extracted with ethyl acetate / water six times. The organic phase was concentrated to a syrupy solution and extracted with petroleum ether:methyl tert-butyl ether (20:1) three times, then with ethyl acetate: petroleum ether (1:10) to give an oily product 8.80 g in 71.31% yield.

[0081] Third step: Preparation of ((2R,3S,4R,5R)-3,4-dihydroxy-5-(4-(hydroxyamino)-2- oxopyrimidin-1(2H)-yl)tetrahydrofuran-2-yl)methyl isobutyrate (9)

[0082] To a solution of ((2R,3S,4R,5R)-3,4-dihydroxy-5-(2-oxo-4-(1H-1,2,4-triazol-1- yl)pyrimidin-1(2H)-yl)tetrahydrofuran-2-yl)methyl isobutyrate (6) (8.80 g, 24.10 mmol) in a 100 ml reaction flask, isopropanol was added and stirred to mix. Hydroxylamine hydrochloride (36.15 mmol) and sodium bicarbonate (36.15 mmol) were dissolved in 6 ml water to give a prepared hydroxylamine aqueous solution. The reaction was stirred for 3 h at room temperature. The isopropanol was removed by concentration under reduced pressure. The reaction mixture was extracted with ethyl acetate / water four times. The organic layer was concentrated to give an oily product 6.82 g in 85.89% yield.

[0083] Fourth step: Synthesis of octanoic acid prodrug (compound 0836)

[0084] To a solution of N4-hydroxycytidine isobutyrate (400 mg, 1.22 mmol) in a 50 ml reaction flask, dichloromethane (10 ml), octanoic acid (166 mg, 1.15 mmol) and condensing agent 1-(3-dimethylaminopropyl)-3-ethylcarbodiimide hydrochloride (256 mg, 1.34 mmol), 4-dimethylaminopyridine (122 mg, 0.49 mmol) were added. The reaction was stirred for 6 h at room temperature. The reaction mixture was extracted with dichloromethane / water. The organic layer was concentrated. The residue was purified by column chromatography (DCM:MeOH (v / v) = 80:1 to 50:1) to give a solid product in 30.2% yield.

[0085] 1 H NMR (600 MHz, chloroform-d) δ 8.82 (s, 1H), 7.09 (d, J = 8.3 Hz, 1H), 5.79 (d, J = 8.3 Hz, 1H), 5.76 (d, J = 4.0 Hz, 1H), 4.34 (d, J = 3.6 Hz, 2H), 4.28 (q, J = 3.9 Hz, 1H), 4.20 (t, J = 4.8 Hz, 1H), 4.17 (t, J = 5.2 Hz, 1H), 3.18 (s, 2H), 2.58 (p, J = 7.0 Hz, 1H), 2.48 (t, J = 7.5 Hz, 2H), 1.69 (p, J = 7.5 Hz, 2H), 1.37 - 1.25 (m, 8H), 1.18 (dd, J = 6.9, 1.2 Hz, 6H), 0.88 (t, J = 6.8 Hz, 3H). MS (ESI): m / z = 456.23 (M+H) + .

[0086] Example 2: Preparation of DL-Thioctic Acid Prodrug (Compound 0838)

[0087] First Step: Preparation of 1-((2R,3R,4S,5R)-3,4-dihydroxy-5- (hydroxymethyl)tetrahydrofuran-2-yl)-4-(1H-1,2,4-triazol-1-yl)pyrimidin-2(1H)-yl) (5)

[0088] After drying a 500 ml three-necked flask, uridine (30.00 g, 122.85 mmol), anhydrous acetonitrile (225 ml) and N-methylpyrrolidine (192.0 ml, 1.843 mol) were added, and TMSCl (78.0 ml, 614.25 mmol) was slowly added dropwise under nitrogen protection and 0 °C ice bath, and stirred at room temperature for 1 h. Then, phosphorus oxychloride (22.5 ml, 245.7 mmol) was slowly added dropwise under 0 °C ice bath, and stirred for 20 min under ice bath after the addition was completed. Triazole (84.60 g, 1.23 mol) was added to the reaction solution under 0 °C ice bath, and stirred for 1 h under ice bath and 2 h at room temperature. After the reaction was completed, the reaction solution was poured into a prepared 600 ml saturated sodium chloride solution to quench the reaction, extracted with dichloromethane (200 ml) for three times, the organic phases were combined, dried over anhydrous sodium sulfate, filtered, and concentrated to obtain an orange-red oil. Methanol:acetic acid (v / v = 10:1) mixed solvent (120 mL) was added, stirred at room temperature overnight, filtered, and dried to obtain a light yellow solid 30.24 g with a yield of 83.4%.

[0089] Step 2: Preparation of ((3aR,4R,6R,6aR)-2,2-dimethyl-6-(2-oxo-4-(1H-1,2,4-triazol-1- yl)pyrimidin-1(2H)-yl)tetrahydrofuran[3,4-d][1,3]dioxol-4-yl)methyl isobutyrate (7)

[0090] The compound 1-((2R,3R,4S,5R)-3,4-dihydroxy-5-(hydroxymethyl)tetrahydrofuran-2-yl)-4-(1H- 1,2,4-triazol-1-yl)pyrimidin-2(1H)-yl) (5) (29.07 g, 98.46 mmol) obtained above was added to a round bottom flask, anhydrous acetonitrile 300 ml was added, 2,2-dimethoxypropane (24.21 ml, 196.41 mmol) was added with stirring, concentrated sulfuric acid (4.92 mmol) was added dropwise, and the reaction was allowed to proceed at room temperature for 6 h. The reaction solution was filtered under suction to obtain a light yellow solid. The light yellow solid was added to a reaction flask, anhydrous acetonitrile 350 ml was added, triethylamine (109.50 ml, 787.68 mmol), 4-dimethylaminopyridine (3.01 g, 24.61 mmol), and isobutyric anhydride (18.00 ml, 108.31 mmol) were added with stirring, and the reaction was allowed to proceed at room temperature for 1 h. Most of the solvent was removed by rotary evaporation, and the residue was dissolved in ethyl acetate and washed once with saturated aqueous sodium bicarbonate solution and saturated sodium chloride solution, respectively, and dried by rotary evaporation to obtain a light yellow solid 32.94, with a yield of 82.53%.

[0091] Step 3: Preparation of ((3aR,4R,6R,6aR)-6-(4-(hydroxyamino)-2-oxopyrimidin-1(2H)-yl)-2,2- dimethyltetrahydrofuran[3,4-d][1,3]dioxol-4-yl)methyl isobutyrate (8).

[0092] The compound ((3aR,4R,6R,6aR)-2,2-dimethyl-6-(2-oxo-4-(1H-1,2,4-triazol-1-yl)pyrimidin- 1(2H)-yl)tetrahydrofuran[3,4-d][1,3]dioxol-4-yl)methyl isobutyrate (7) (30 g, 74.00 mmol) obtained above was added to a 500 ml reaction flask, isopropanol 300 ml was added and stirred to mix, and a prepared hydroxylamine aqueous solution (hydroxylamine hydrochloride 111.00 mmol and sodium bicarbonate 111.00 mmol dissolved in 60 ml of water) was added, and the reaction was allowed to proceed at room temperature for 3 h. After most of the isopropanol was removed by concentration under reduced pressure, ice water 50 ml was added, and the mixture was slurried in an ice bath for 1 h, and filtered under suction to obtain a white solid 23.58 g, with a yield of 85.63%.

[0093] Step 4: Preparation of ((2R, 3S, 4R, 5R)-3, 4-dihydroxy-5-(4-(hydroxyamino)-2- oxopyrimidin-1 (2H)-yl)tetrahydrofuran-2-yl)methyl isobutyrate (9)

[0094] The compound ((3aR, 4R, 6R, 6aR)-6-(4-(hydroxyamino)-2-oxopyrimidin-1 (2H)-yl)- 2, 2-dimethyltetrahydrofuran[3, 4-d] [1, 3]dioxol-4-yl)methyl isobutyrate (8) (20 g, 54.15 mmol) obtained above was added to a 500 ml reaction flask, 100 ml of 25% methanol aqueous solution was added, and sulfuric acid (43.32 mmol) was slowly added dropwise with stirring. The reaction was stirred at 70°C for 1 h. After cooling to room temperature, sodium bicarbonate was added to adjust the pH to neutral, and suction filtration was performed. The filtrate was extracted with ethyl acetate 10 times (80 ml x 10), the solvent was rotary dried, and EA: DCM: MeOH = 15:5:2, 20 ml was used for slurry purification. A total of 12.52 g of the compound was obtained, and the yield was 70.2%.

[0095] Step 5: Synthesis of DL-lipoic acid prodrug (compound 0838)

[0096] N4-hydroxycytidine isobutyrate (400 mg, 1.22 mmol) was added to a 50 ml reaction flask, dichloromethane (10 ml) was added, lipoic acid (237 mg, 1.15 mmol) and condensing agent 1-(3-dimethylaminopropyl)-3-ethylcarbodiimide hydrochloride (256 mg, 1.34 mmol), 4-dimethylaminopyridine (122 mg, 0.49 mmol) were added, and stirring was performed at room temperature for 6 hours. Dichloromethane / water extraction was performed, the organic phase was concentrated, and the obtained residue was purified by column chromatography (DCM: MeOH (v / v) = 80:1 ~ 50:1) to obtain a solid product, and the yield was 48.9%.

[0097] 1 H NMR (400 MHz, Chloroform-d) δ 8.90 (s, 1H), 7.10 (d, J = 8.3 Hz, 1H), 5.81 - 5.73 (m, 2H), 4.35 (d, J = 3.5 Hz, 2H), 4.28 (q, J = 3.8 Hz, 1H), 4.18 (dq, J = 10.6, 5.2 Hz, 2H), 3.60 - 3.55 (m, 1H), 3.45 (s, 1H), 3.22 - 3.06 (m, 3H), 2.54 - 2.46 (m, 3H), 1.95 - 1.91 (m, 1H), 1.82 (s, 1H), 1.75 - 1.68 (m, 4H), 1.19 (d, J = 6.8 Hz, 6H). MS (ESI): m / z = 518.08 (M+H)+ .

[0098] Example 3: Preparation of decanoic acid prodrug (Compound 0839)

[0099] Compound 0839 was prepared in the same manner as Example 1 except that octanoic acid was replaced by decanoic acid. The solid product was obtained in 21.1% yield.

[0100] 1 H NMR (600 MHz, Chloroform-d) δ 8.88 (s, 1H), 7.09 (d, J = 8.3 Hz, 1H), 5.79 (dd, J = 8.3, 1.6 Hz, 1H), 5.76 (d, J = 4.1 Hz, 1H), 4.34 (d, J = 3.6 Hz, 2H), 4.27 (dd, J = 5.0, 3.4 Hz, 1H), 4.23 - 4.14 (m, 2H), 2.58 (hept, J = 7.0 Hz, 1H), 2.48 (t, J = 7.6 Hz, 2H), 1.69 (p, J = 7.5 Hz, 2H), 1.37 - 1.23 (m, 12H), 1.18 (dd, J = 6.9, 1.1 Hz, 6H), 0.88 (t, J = 7.0 Hz, 3H). MS (ESI): m / z = 484.26 (M+H) + .

[0101] Example 4: Preparation of octadecenoic acid prodrug (Compound 0844)

[0102] Compound 0844 was prepared in the same manner as Example 1 except that octanoic acid was replaced by octadecenoic acid. The solid product was obtained in 36.0% yield.

[0103] 1H NMR (600 MHz, Chloroform-d) δ 8.62 (s, 1H), 7.10 (d, J = 8.4 Hz, 1H), 5.79 (d, J = 8.3 Hz, 1H), 5.73 (d, J = 4.0 Hz, 1H), 5.36 - 5.33 (m, 2H), 4.34 (t, J = 3.2 Hz, 2H), 4.31 - 4.29 (m, 1H), 4.20 (dd, J = 5.5, 4.0 Hz, 1H), 4.17 (t, J = 5.1 Hz, 1H), 2.57 (p, J = 7.0 Hz, 1H), 2.47 (t, J = 7.6 Hz, 2H), 2.01 (dd, J = 7.2, 3.0 Hz, 4H), 1.70 (p, J = 7.5 Hz, 2H), 1.40 - 1.21 (m, 20H), 1.18 (dd, J = 6.9, 2.0 Hz, 6H), 0.88 (t, J = 7.0 Hz, 3H). MS (ESI): m / z = 594.37 (M+H) + .

[0104] Example 5: Preparation of linoleic acid prodrug (Compound 0845)

[0105] Compound 0845 was prepared in the same manner as Example 1 except that octanoic acid was replaced by linoleic acid. The final product was obtained as a solid with a yield of 37.2%.

[0106] 1 H NMR (600 MHz, Chloroform-d) δ 7.10 (d, J = 8.3 Hz, 1H), 5.79 (dd, J = 8.3, 2.0 Hz, 1H), 5.73 (d, J = 3.9 Hz, 1H), 5.39 - 5.30 (m, 4H), 4.34 (dd, J = 3.4, 1.8 Hz, 2H), 4.31 - 4.27 (m, 1H), 4.21 - 4.16 (m, 2H), 2.77 (t, J = 6.9 Hz, 2H), 2.57 (p, J = 7.0 Hz, 1H), 2.47 (t, J = 7.6 Hz, 2H), 2.07 - 2.02 (m, 4H), 1.72 - 1.67 (m, 2H), 1.37 - 1.29 (m, 14H), 1.18 (dd, J = 6.9, 1.7 Hz, 6H), 0.89 (t, J = 6.9 Hz, 3H). MS (ESI): m / z = 592.37 (M+H) + .

[0107] Example 6: Preparation of 4-methyloctanoic acid prodrug (Compound 0850)

[0108] Compound 0850 was prepared in the same manner as Example 1 except that octanoic acid was replaced by 4-methyloctanoic acid. The solid product was obtained in 44.5% yield.

[0109] 1 H NMR (400 MHz, DMSO-d6) δ 7.18 (d, J = 8.1 Hz, 1H), 5.76 - 5.71 (m, 2H), 5.47 (d, J = 5.5 Hz, 1H), 5.29 (d, J = 5.3 Hz, 1H), 4.23 - 4.15 (m, 2H), 4.06 - 4.02 (m, 1H), 3.98 - 3.92 (m, 2H), 2.62 - 2.56 (m, 1H), 2.44 (t, J = 7.5 Hz, 2H), 1.54 (dd, J = 10.4, 3.9 Hz, 2H), 1.25 (d, J = 5.2 Hz, 13H), 1.10 (d, J = 6.9 Hz, 5H), 0.86 (d, J = 6.2 Hz, 3H). MS (ESI): m / z = 498.27 (M+H)

[0110] MS (ESI): m / z = 470.25 (M+H) + .

[0111] Example 7: Preparation of undecanoic acid prodrug (Compound 0901)

[0112] Compound 0901 was prepared in the same manner as Example 1 except that octanoic acid was replaced by undecanoic acid. The solid product was obtained in 22.1% yield.

[0113] 1 H NMR (400 MHz, DMSO-d6) δ 7.18 (d, J = 8.1 Hz, 1H), 5.76 - 5.71 (m, 2H), 5.47 (d, J = 5.5 Hz, 1H), 5.29 (d, J = 5.3 Hz, 1H), 4.23 - 4.15 (m, 2H), 4.06 - 4.02 (m, 1H), 3.98 - 3.92 (m, 2H), 2.62 - 2.56 (m, 1H), 2.44 (t, J = 7.5 Hz, 2H), 1.54 (dd, J = 10.4, 3.9 Hz, 2H), 1.25 (d, J = 5.2 Hz, 13H), 1.10 (d, J = 6.9 Hz, 5H), 0.86 (d, J = 6.2 Hz, 3H). MS (ESI): m / z = 498.27 (M+H) + .

[0114] Example 8: Preparation of undecenoic acid prodrug (Compound 0902)

[0115] Compound 0902 was prepared in the same manner as Example 1 except that octanoic acid was replaced by undecylenic acid. The solid product was obtained in 40.0% yield.

[0116] 1 H NMR (600 MHz, Chloroform-d) δ 8.84 (s, 1H), 7.09 (d, J = 8.3 Hz, 1H), 5.81 - 5.77 (m, 2H), 5.75 (d, J = 4.0 Hz, 1H), 4.99 (dt, J = 17.2, 1.9 Hz, 1H), 4.93 (dq, J = 10.2, 1.5 Hz, 1H), 4.34 (d, J = 3.7 Hz, 2H), 4.29 - 4.26 (m, 1H), 4.21 - 4.15 (m, 2H), 2.60 - 2.56 (m, 1H), 2.47 (t, J = 7.5 Hz, 2H), 2.04 - 2.02 (m, 2H), 1.68 (d, J = 7.6 Hz, 2H), 1.32 - 1.27 (m, 10H), 1.18 (dd, J = 7.1, 1.2 Hz, 6H). MS (ESI): m / z = 496.27 (M+H) + .

[0117] Example 9: Preparation of palmitoleic acid prodrug (Compound 0903)

[0118] Compound 0903 was prepared in the same manner as Example 1 except that octanoic acid was replaced by palmitoleic acid. The solid product was obtained in 40.7% yield.

[0119] 1 H NMR (600 MHz, Chloroform-d) δ 8.74 (s, 1H), 7.10 (d, J = 8.3 Hz, 1H), 5.79 (dd, J = 8.3, 1.7 Hz, 1H), 5.74 (d, J = 4.0 Hz, 1H), 5.34 (q, J = 5.5 Hz, 2H), 4.34 (d, J = 3.5 Hz, 2H), 4.31 - 4.25 (m, 1H), 4.23 - 4.13 (m, 2H), 2.58 (dq, J = 14.0, 7.0 Hz, 1H), 2.47 (t, J = 7.6 Hz, 2H), 2.04 - 1.98 (m, 4H), 1.69 (p, J = 7.5 Hz, 2H), 1.39 - 1.24 (m, 18H), 1.18 (dd, J = 7.0, 1.5 Hz, 6H), 0.88 (t, J = 6.9 Hz, 3H). MS (ESI): m / z = 566.35 (M+H) + .

[0120] Example 10: Preparation of azelaic acid monomethyl ester prodrug (Compound 0904)

[0121] Compound 0904 was prepared in the same manner as Example 1 except that caprylic acid was replaced by azelaic acid monomethyl ester. The solid product was obtained in 23.1% yield.

[0122] 1 H NMR (400 MHz, DMSO-d6) δ 10.88 (s, 1H), 7.11 (d, J = 8.2 Hz, 1H), 5.70 - 5.65 (m, 2H), 5.40 (d, J = 5.6 Hz, 1H), 5.22 (d, J = 5.3 Hz, 1H), 4.17 - 4.09 (m, 2H), 3.97 (t, J = 5.2 Hz, 1H), 3.91 - 3.85 (m, 2H), 3.34 (s, 2H), 2.52 (td, J = 6.9, 1.5 Hz, 1H), 2.39 (t, J = 7.8 Hz, 2H), 1.69 - 1.63 (m, 3H), 1.56 - 1.36 (m, 8H), 1.03 (d, J = 7.0 Hz, 6H). MS (ESI): m / z = 454.22 (M+H) + .

[0123] Example 11: Preparation of cyclopentane propanoic acid prodrug (Compound 0905)

[0124] Compound 0905 was prepared in the same manner as Example 1 except that caprylic acid was replaced by cyclopentane propanoic acid. The solid product was obtained in 27.6% yield.

[0125] 1 H NMR (400 MHz, DMSO-d6) δ 10.88 (s, 1H), 7.11 (d, J = 8.2 Hz, 1H), 5.70 - 5.65 (m, 2H), 5.40 (d, J = 5.6 Hz, 1H), 5.22 (d, J = 5.3 Hz, 1H), 4.17 - 4.09 (m, 2H), 3.97 (t, J = 5.2 Hz, 1H), 3.91 - 3.85 (m, 2H), 3.34 (s, 2H), 2.52 (td, J = 6.9, 1.5 Hz, 1H), 2.39 (t, J = 7.8 Hz, 2H), 1.69 - 1.63 (m, 3H), 1.56 - 1.36 (m, 8H), 1.03 (d, J = 7.0 Hz, 6H). MS (ESI): m / z = 454.22 (M+H) + .

[0126] Example 12: Preparation of 6-phenylhexanoic acid prodrug (Compound 0906)

[0127] Compound 0906 was prepared in the manner of Example 1 except that octanoic acid was replaced with 6-phenylhexanoic acid. The final product was obtained as a solid with a yield of 45.8%.

[0128] 1 H NMR (400 MHz, DMSO-d6) δ 7.29 - 7.15 (m, 6H), 5.77 - 5.67 (m, 2H), 5.47 (d, J = 5.5 Hz, 1H), 5.29 (d, J = 5.3 Hz, 1H), 4.27 - 4.13 (m, 2H), 4.05 (q, J = 5.3 Hz, 1H), 4.01 - 3.90 (m, 2H), 3.39 (s, 19H), 2.58 (q, J = 7.3, 6.9 Hz, 3H), 2.45 (t, J = 7.5 Hz, 2H), 1.59 (q, J = 7.6 Hz, 4H), 1.31 (td, J = 7.2, 3.9 Hz, 2H), 1.10 (dd, J = 7.0, 3.9 Hz, 6H). MS (ESI): m / z = 504.25 (M+H) + .

[0129] Example 13: Preparation of 10-undecynoic acid prodrug (Compound 0907)

[0130] Compound 0907 was prepared in the manner of Example 1 except that octanoic acid was replaced with 10-undecynoic acid. The final product was obtained as a solid with a yield of 16.0%.

[0131] 1 H NMR (600 MHz, Chloroform-d) δ 8.77 (s, 1H), 7.09 (d, J = 8.3 Hz, 1H), 5.79 (dd, J = 8.3, 2.0 Hz, 1H), 5.74 (d, J = 3.9 Hz, 1H), 4.35 - 4.34 (m, 2H), 4.29 - 4.27 (m, 1H), 4.22 - 4.16 (m, 2H), 2.58 (dt, J = 14.0, 7.0 Hz, 1H), 2.48 (t, J = 7.6 Hz, 2H), 2.19 - 2.17 (m, 2H), 1.94 (t, J = 2.6 Hz, 1H), 1.69 (d, J = 7.5 Hz, 2H), 1.54 - 1.50 (m, 2H), 1.40 - 1.31 (m, 8H), 1.18 (dd, J = 7.1, 1.4 Hz, 6H). MS (ESI): m / z = 494.25 (M+H) + .

[0132] Example 14: Preparation of 2-methylheptanoic acid prodrug (Compound 0908)

[0133] Compound 0908 was prepared in the same manner as Example 1 except that octanoic acid was replaced by 2-methylheptanoic acid. The solid product was obtained in 33.2% yield.

[0134] 1H NMR (600 MHz, Chloroform-d) δ 8.49 (s, 1H), 7.08 (dd, J = 8.2, 2.2 Hz, 1H), 5.82 (d, J = 8.2 Hz, 1H), 5.75 (dd, J = 7.8, 4.0 Hz, 1H), 4.34 (d, J = 3.6 Hz, 2H), 4.28 (q, J = 3.8 Hz, 1H), 4.23 - 4.16 (m, 2H), 2.59 (ddt, J = 24.1, 14.0, 7.0 Hz, 2H), 1.48 (ddt, J = 13.2, 10.0, 6.4 Hz, 1H), 1.33 - 1.26 (m, 6H), 1.23 (d, J = 6.9 Hz, 3H), 1.18 (dd, J = 7.0, 1.4 Hz, 6H), 0.88 (td, J = 7.0, 2.2 Hz, 3H). MS (ESI): m / z = 456.23 (M+H) + .

[0135] Example 15: Preparation of 4-ethyloctanoic acid prodrug (Compound 0909)

[0136] Compound 0909 was prepared in the same manner as Example 1 except that octanoic acid was replaced by 4-ethyloctanoic acid. The solid product was obtained in 36.0% yield.

[0137] 1 1H NMR (600 MHz, Chloroform-d) δ 8.69 (s, 1H), 7.10 (d, J = 8.2 Hz, 1H), 5.79 (d, J = 8.3 Hz, 1H), 5.73 (d, J = 3.9 Hz, 1H), 4.35 (dd, J = 3.5, 2.0 Hz, 2H), 4.30 - 4.28 (m, 1H), 4.21 - 4.16 (m, 2H), 2.60 - 2.56 (m, 1H), 2.46 (dd, J = 9.0, 6.9 Hz, 2H), 1.68 - 1.64 (m, 2H), 1.32 - 1.25 (m, 9H), 1.18 (dd, J = 6.9, 1.6 Hz, 6H), 0.91 - 0.86 (m, 6H). MS (ESI): m / z = 484.28 (M+H) + .

[0138] Example 16: Preparation of trans-4-butylcyclohexanecarboxylic acid prodrug (Compound 0910)

[0139] Compound 0910 was prepared in the manner of Example 1 except that octanoic acid was replaced by trans-4-butylcyclohexanecarboxylic acid. The final product was obtained as a solid in 30.6% yield.

[0140] 1 H NMR (600 MHz, Chloroform-d) δ 8.45 (s, 1H), 7.08 (d, J = 8.2 Hz, 1H), 5.81 (d, J = 8.3 Hz, 1H), 5.75 (d, J = 4.1 Hz, 1H), 4.34 (d, J = 3.6 Hz, 2H), 4.28 (q, J = 3.8 Hz, 1H), 4.19 (dt, J = 16.1, 5.3 Hz, 2H), 2.61 - 2.56 (m, 1H), 2.01 - 1.97 (m, 2H), 1.85 (dd, J = 13.5, 3.3 Hz, 2H), 1.52 (dddd, J = 17.2, 13.1, 8.6, 4.1 Hz, 2H), 1.30 - 1.20 (m, 9H), 1.18 (dd, J = 6.9, 1.6 Hz, 6H), 0.90 - 0.87 (m, 3H). MS (ESI): m / z = 496.28 (M+H) + .

[0141] Example 17: Preparation of cyclohexylbutyric acid prodrug (Compound 0912)

[0142] Compound 0912 was prepared in the manner of Example 1 except that octanoic acid was replaced by trans-cyclohexylbutyric acid. The final product was obtained as a solid in 17.8% yield.

[0143] 1 H NMR (600 MHz, Chloroform-d) δ 8.90 (s, 1H), 7.09 (d, J = 8.3 Hz, 1H), 5.78 (dd, J = 12.2, 6.1 Hz, 2H), 4.34 (d, J = 3.5 Hz, 2H), 4.26 (t, J = 4.1 Hz, 1H), 4.18 (dt, J = 25.4, 5.2 Hz, 2H), 2.58 (p, J = 7.0 Hz, 1H), 2.45 (t, J = 7.6 Hz, 2H), 1.73 - 1.67 (m, 7H), 1.28 - 1.21 (m, 6H), 1.18 (d, J = 7.1 Hz, 6H), 0.88 (dt, J = 10.0, 5.3 Hz, 2H). MS (ESI): m / z = 482.28 (M+H) + .

[0144] Example 18: Preparation of BOC-11-aminoundecanoic acid prodrug (Compound 0923)

[0145] Compound 0923 was prepared in the manner of Example 1 except that octanoic acid was replaced by BOC-11-aminoundecanoic acid. The solid product was obtained in the end with a yield of 12.6%.

[0146] 1 H NMR (400 MHz, DMSO-d6) δ 10.93 (s, 1H), 6.76 (s, 1H), 5.75 (d, J = 3.1 Hz, 1H), 5.47 (d, J = 5.4 Hz, 1H), 5.29 (d, J = 5.2 Hz, 1H), 4.26 - 4.13 (m, 3H), 4.03 (t, J = 5.1 Hz, 1H), 3.98 - 3.91 (m, 2H), 2.88 (d, J = 6.5 Hz, 2H), 2.60 - 2.56 (m, 1H), 2.44 (t, J = 7.5 Hz, 2H), 1.54 (s, 2H), 1.36 (s, 11H), 1.23 (s, 12H), 1.10 (d, J = 7.0 Hz, 6H). MS (ESI): m / z = 613.36 (M+H) + .

[0147] Example 19: Preparation of BOC-8-aminooctanoic acid prodrug (Compound 0924)

[0148] Compound 0923 was prepared in the manner of Example 1 except that octanoic acid was replaced by BOC-8-aminooctanoic acid. The solid product was obtained in the end with a yield of 15.4%.

[0149] 1 H NMR (400 MHz, DMSO-d6) δ 10.93 (s, 1H), 7.18 (d, J = 8.2 Hz, 1H), 6.78 (s, 1H), 5.75 (d, J = 4.2 Hz, 1H), 5.47 (d, J = 5.5 Hz, 1H), 5.29 (d, J = 5.4 Hz, 1H), 4.26 - 4.14 (m, 3H), 4.04 (q, J = 5.5 Hz, 1H), 3.98 - 3.91 (m, 2H), 2.89 (d, J = 6.5 Hz, 2H), 2.61 - 2.56 (m, 1H), 2.44 (t, J = 7.5 Hz, 2H), 1.55 - 1.51 (m, 2H), 1.37 (s, 11H), 1.29 - 1.24 (m, 6H), 1.10 (d, J = 6.9 Hz, 6H). MS (ESI): m / z = 571.31 (M+H) + .

[0150] Example 20: Preparation of R-(+)-lipoic acid prodrug (compound 0926)

[0151] Compound 0926 was prepared in the same manner as Example 1 except that octanoic acid was replaced by R-(+)-lipoic acid. The final product was obtained as a solid with a yield of 28.9%.

[0152] 1 H NMR (400 MHz, DMSO-d6) δ 10.94 (s, 1H), 7.19 (d, J = 8.2 Hz, 1H), 5.75 - 5.73 (m, 1H), 5.47 (d, J = 5.6 Hz, 1H), 5.29 (d, J = 5.3 Hz, 1H), 4.19 (dtt, J = 20.8, 8.8, 3.5 Hz, 3H), 4.04 (q, J = 5.3 Hz, 1H), 3.98 - 3.91 (m, 2H), 3.24 - 3.10 (m, 3H), 2.46 (q, J = 7.1 Hz, 3H), 1.88 (dd, J = 12.9, 6.6 Hz, 1H), 1.71 - 1.66 (m, 1H), 1.62 - 1.54 (m, 4H), 1.43 - 1.38 (m, 2H), 1.10 (d, J = 7.0 Hz, 6H). MS (ESI): m / z = 518.20 (M+H) + .

[0153] Example 21: Preparation of chenodeoxycholic acid prodrug (compound 0939)

[0154] 1. Dissolve chenodeoxycholic acid in dichloromethane, add 4-dimethylaminopyridine, add acetic anhydride under stirring, stir at room temperature for 3 hours. Wash the reaction solution with 1 mol / L HC1 (3 x 30 ml), saturated sodium bicarbonate solution, saturated sodium chloride solution respectively, concentrate the organic layer, purify the obtained residue by column chromatography (PE:EA (v / v) = 5:1) to obtain a white solid.

[0155] 2. Dissolve N4-hydroxycytidine isobutyric acid ester in dichloromethane, add the above white solid (hydroxyl-protected acid) and condensing agent 1-(3-dimethylaminopropyl)-3-ethylcarbodiimide hydrochloride, 4-dimethylaminopyridine, stir at room temperature for 6 hours, extract with dichloromethane / water, spin dry, and purify by column chromatography (DCM:MeOH (v / v) = 80:1, 50:1) to obtain a white solid product with a yield of 18.9%.

[0156] 1H NMR (400 MHz, Chloroform-d) δ 7.11 (d, J = 8.3 Hz, 1H), 5.80 (dd, J = 8.3, 2.0 Hz, 1H), 5.74 (d, J = 3.6 Hz, 1H), 4.58 (tt, J = 11.3, 4.5 Hz, 1H), 4.35 (d, J = 3.5 Hz, 2H), 4.28 (dt, J = 5.1, 3.5 Hz, 1H), 4.21 - 4.13 (m, 2H), 3.86 (q, J = 3.0 Hz, 1H), 2.58 (dt, J = 14.0, 7.0 Hz, 1H), 2.51 (dt, J = 9.8, 4.9 Hz, 1H), 2.41 (dd, J = 9.4, 6.4 Hz, 1H), 2.30 (d, J = 12.4 Hz, 1H), 2.01 (s, 3H), 1.88 - 1.43 (m, 20H), 1.25 - 1.13 (m, 11H), 0.96 (d, J = 6.3 Hz, 3H), 0.92 (s, 3H), 0.67 (s, 3H). MS (ESI): m / z = 746.48 (M+H) + .。

[0157] Example 22: Preparation of Ursodeoxycholic acid prodrug (Compound 0941)

[0158] Compound 0941 was prepared in the same manner as Example 21 except that chenodeoxycholic acid was replaced by ursodeoxycholic acid. The final product was obtained as a solid with a yield of 11.3%.

[0159] 1 H NMR (400 MHz, Chloroform-d) δ 7.11 (d, J = 8.3 Hz, 1H), 5.80 (dd, J = 8.3, 2.0 Hz, 1H), 5.74 (d, J = 3.6 Hz, 1H), 4.58 (tt, J = 11.3, 4.5 Hz, 1H), 4.35 (d, J = 3.5 Hz, 2H), 4.28 (dt, J = 5.1, 3.5 Hz, 1H), 4.21 - 4.13 (m, 2H), 3.86 (q, J = 3.0 Hz, 1H), 2.58 (dt, J = 14.0, 7.0 Hz, 1H), 2.51 (dt, J = 9.8, 4.9 Hz, 1H), 2.41 (dd, J = 9.4, 6.4 Hz, 1H), 2.30 (d, J = 12.4 Hz, 1H), 2.01 (s, 3H), 1.88 - 1.43 (m, 20H), 1.25 - 1.13 (m, 11H), 0.96 (d, J = 6.3 Hz, 3H), 0.92 (s, 3H), 0.67 (s, 3H). MS (ESI): m / z = 746.48 (M+H) + .

[0160] Example 23: Lithocholic acid prodrug (Compound 0943)

[0161] Compound 0941 was prepared in the same manner as Example 21 except that chenodeoxycholic acid was replaced by lithocholic acid. The solid product was obtained in 12.2% yield.

[0162] 1 H NMR (400 MHz, Chloroform-d) δ 7.10 (d, J = 8.1 Hz, 1H), 5.79 (dd, J = 8.3, 2.0 Hz, 1H), 5.75 (d, J = 3.7 Hz, 1H), 4.72 (dt, J = 11.3, 6.3 Hz, 1H), 4.35 (d, J = 3.4 Hz, 2H), 4.27 (dd, J = 5.0, 3.3 Hz, 1H), 4.17 (dq, J = 10.5, 5.4 Hz, 3H), 2.62 - 2.50 (m, 2H), 2.39 (td, J = 9.7, 4.8 Hz, 1H), 2.03 (s, 3H), 1.88 - 1.38 (m, 21H), 1.19 (d, J = 6.9 Hz, 6H), 1.10 - 1.03 (m, 5H), 0.95 - 0.92 (m, 6H), 0.65 (s, 3H). MS (ESI): m / z = 730.43 (M+H) + .

[0163] Example 24: Preparation of deoxycholic acid prodrug (Compound 0945)

[0164] Compound 0945 was prepared in the same manner as Example 21 except that chenodeoxycholic acid was replaced by deoxycholic acid. The solid product was obtained in 13.9% yield.

[0165] 1 H NMR (400 MHz, Chloroform-d) δ 7.20 (d, J = 8.2 Hz, 1H), 5.79 (dd, J = 8.3, 1.7 Hz, 1H), 5.71 (d, J = 1.7 Hz, 1H), 4.73 (dq, J = 11.0, 5.5, 4.7 Hz, 1H), 4.43 - 4.36 (m, 2H), 4.25 (dt, J = 6.6, 3.2 Hz, 1H), 4.18 (d, J = 3.8 Hz, 2H), 4.00 (d, J = 3.1 Hz, 1H), 2.62 - 2.56 (m, 1H), 2.51 - 2.31 (m, 3H), 2.01 (s, 3H), 1.85 - 1.47 (m, 19H), 1.20 (dd, J = 7.0, 1.7 Hz, 6H), 1.17 - 1.03 (m, 5H), 1.01 (d, J = 6.0 Hz, 3H), 0.93 (s, 3H), 0.69 (s, 3H). MS (ESI): m / z = 746.48 (M+H) + .

[0166] Test Example 1: Anti-VSV virus cell activity screening

[0167] Test principle: African green monkey kidney cells (Vero) were used as host cells to measure the green fluorescence signal of recombinant vesicular stomatitis virus inserted with green fluorescent protein (GFP), so as to measure the inhibition efficiency of the sample.

[0168] Test materials and methods:

[0169] Vesicular stomatitis virus (VSV) was donated by Professor Zhang Weina's laboratory of Military Medical Research Institute.

[0170] Cell culture: African green monkey kidney cells (Vero) were used as host cells and cultured in MEM medium (CM50011, MACGENE) containing 10% fetal bovine serum (HQ30071-T500, Hongquan Biological)

[0171] Virus strain: Vesicular stomatitis virus (containing green fluorescent protein reporter gene), cultured and passaged in African green monkey kidney cells (Vero), -80℃.

[0172] Sample treatment: The sample was configured into a mother liquor with DMSO, and then diluted with cell culture solution.

[0173] Positive control drug: Molnupiravir

[0174] Test method: Vero cells were inoculated in a 96-well plate (2×104 cells / well) and incubated at 37℃, 5% CO2 for 12h. After the cells adhered, the old culture solution was replaced with the culture solution containing the drug, and three replicate wells were set, namely the solvent control group, the virus control group, and the drug administration group. MDCK cells were infected with vesicular stomatitis virus with a virus titer of 50 PFU in the virus control group and the drug administration group, and incubated at 37℃, 5% CO2. After 24h, the GFP signal was detected at an excitation wavelength of 488nm and an emission wavelength of 535nm using a fluorescence microplate reader. The inhibition rate calculation formula is: inhibition rate = 1-[(drug group fluorescence intensity-background fluorescence intensity) / (DMSO group fluorescence intensity-background fluorescence intensity)].

[0175] The anti-VSV virus test results are shown in Table 1 below.

[0176] Test Example 2: MDCK cell toxicity test

[0177] Test method: MDCK cells (purchased from ATCC company) were cultured in high-glucose DMEM medium (CM10017, MACGENE) containing 10% fetal bovine serum (10091148, Gibco), and when the cell density was about 90%, they were plated in a 96-well plate, 2×104 Cells were seeded in 96-well plates at a density of 5000 cells / well and incubated at 37°C in 5% CO2 for 12 h. After the cells adhered, the old medium was discarded, and 200 μl of the corresponding concentration of the compound was added to each well, with three replicates. A solvent control group was also set up. After incubation at 37°C for 72 h, the supernatant was discarded, and 100 μl of freshly prepared MTS (G3580, Promega) detection solution was added to each well, with a background control group. The 96-well plate was incubated in the incubator for 1 h, and the OD value at 490 nm was measured by the enzyme label instrument (Thermo Scientific). The value measured by the enzyme label instrument was automatically subtracted from the background value. Cell viability (%) = OD value of the drug test group / OD value of the solvent control group x 100%.

[0178] The effect of the drug on cell viability is shown in Table 2 below.

[0179] Table 1: Anti-VSV efficacy test results of the example compounds

[0180] Table 2. MDCK cell toxicity test results of the example compounds

[0181] Test Example 3: Anti-Staphylococcus aureus experiment

[0182] Reagents:

[0183] MHB medium Beijing Aoboxing Biotechnology Co., Ltd. Batch number: 20200806

[0184] Brain heart infusion (BHI) broth Beijing Aoboxing Biotechnology Co., Ltd. Batch number: 20210813

[0185] Potassium penicillin for injection Sichuan Jishan Family Pharmaceutical Co., Ltd. Batch number: 20190905

[0186] The control compound ZXC-0323 is the lauroyl prodrug (compound 19) in CN114805458B.

[0187] Strains:

[0188] Gram-positive bacteria: Staphylococcus aureus ATCC 29213

[0189] Gram-negative bacteria: Escherichia coli ATCC 25922

[0190] Experimental method:

[0191] The minimum inhibitory concentration (MIC) of the compound was determined by microdilution method (CLSI standard, 2017). A single colony of bacteria was picked and cultured in BHI broth at 37°C until the bacteria reached the logarithmic growth phase. The turbidity of the bacteria was adjusted to 0.5 McFarland turbidity, and the bacteria were diluted 100 times (about 1.0 x 106 CFUs / mL) with MHB medium for standby. The drug to be tested was diluted with MHB medium, 100 μL of which was added to a 96-well U-shaped plate, followed by the addition of 100 μL of the diluted bacteria to be tested to each well. After incubation at 37°C for 18 h, the MIC results were read, and the minimum drug concentration that could inhibit bacterial growth with the naked eye was taken as the MIC value of the drug. The test results are shown in Table 3 below.

[0192] Table 3 Test results of antibacterial activity

[0193] Test Example 4 Beas-2B cell toxicity experiment

[0194] A cell: digest the cells, remove the culture medium, wash with 3 ml of normal saline, and then wash the culture medium. Add 2 ml of trypsin and incubate at 37°C for 3 min. Then add 2 ml of culture medium to neutralize the cells. Use a 1 ml pipette to blow off the adherent cells and transfer them to a 15 ml centrifuge tube. Centrifuge (800 r, 3 min) and discard the supernatant. Add 2 ml of culture medium and mix to count: prepare 1.5 ml EP tubes, add 10 μl of trypan blue and cell suspension to each tube, mix by blowing, and then take 10 μl and add it to the counting plate. Assuming that 8*106 cell active cells are needed per well of a 96-well plate, and 4 plates are needed, 8 concentrations, and 6 rows per plate, the cell suspension needed is 4*8*6*(8*106) / the number of active cells after counting. Each well of the 96-well plate requires 100 μl, 4 plates, 6 rows, and 8 concentrations, so the total liquid needed is 4*6*8*100. Add the cell suspension and culture medium in proportion, mix, and then use a pipette to add it to the 96-well plate.

[0195] Drug addition: weigh 5 mg of each drug, and use a molar calculator to calculate the required volume of DMSO (mass = concentration * volume * molecular weight). The concentration is 100 mM (the required volume is too small, so it is converted to 10 mM). The 8 concentrations are -- NT, 0 μM, 5 μM, 10 μM, 20 μM, 40 μM, 80 μM, and 160 μM. Add 500 μl of culture medium to an EP tube, prepare drugs of different concentrations by adding them to 500 μl of culture medium, label them, and then turn off the wind. Remove the culture medium from the 96-well plate, and then add the prepared culture medium of different concentrations to each well, 100 μl per well, and incubate in a 37°C incubator for 24-48 h.

[0196] Detection: Then draw out 96-well plate culture medium, add 100 μl of the mixed solution of the newly prepared culture medium and MTS (G3580, Promega) at a ratio of 5:1 as detection solution to each well, set up a background control group at the right upper corner of each plate, and place the 96-well plate in a 37℃ incubator for 1-4h of incubation. Measure the OD value at 490nm by using an enzyme marker (Thermo Scientific), and the value measured by the enzyme marker is the value after the background value is automatically subtracted. Cell viability (%) = (OD value of the drug experimental group / OD value of the solvent control group) x 100%. The results are shown in the following Figure 1.

[0197] The above merely provides a specific implementation of the present application, but the protection scope of the present application is not limited thereto, any person skilled in the art can easily think of changes or replacements within the technical range disclosed by the present application, which shall be covered within the protection scope of the present application. Therefore, the protection scope of the present application shall be subject to the protection scope of the claims.

Claims

1. A compound derivative represented by formula I or an isotope-labeled compound thereof, or an optical isomer, geometric isomer, tautomer or isomer mixture thereof, or a pharmaceutically acceptable salt thereof: in, R1 is substituted or unsubstituted C2-C 10 Alkyl, substituted or unsubstituted C 12 Alkyl, substituted or unsubstituted C 14 Alkyl, substituted or unsubstituted C 16 -C 27 Alkyl, substituted or unsubstituted C2-C 27 Alkenyl, substituted or unsubstituted C2-C 27 Alkynyl, substituted or unsubstituted C3-C6 cycloalkyl, wherein the "substituted" means that the group contains 1 to 3 R a The substituent, R a Selected from deuterium, hydroxyl, amino, cyano, nitro, halogen, C1-C5 alkoxy, C1-C5 alkoxycarbonyl, C3-C6 cycloalkyl, -NHCOOR a1 、C6-C 10 Aryl, saturated or unsaturated three to six-membered heterocyclic group containing 1 to 3 heteroatoms selected from N, O, S, three to six-membered heteroaryl containing 1 to 3 heteroatoms selected from N, O, S, unsubstituted or with 1 to 2 R a2 Substituted steroid structure, the R a1 is selected from C1-C6 alkyl, said R a2 Selected from hydroxyl group, C2-C6 ester group; R2 is a substituted or unsubstituted C1-C6 alkyl group, wherein the "substituted" means that the group contains 1 to 3 R b substituted, the R b Selected from deuterium, hydroxy, amino, cyano, nitro, halogen, C1-C5 alkoxy, C1-C5 alkyl substituted by halogen, and C3-C6 cycloalkyl.

2. The compound derivative represented by formula I or its isotope-labeled compound according to claim 1, or its optical isomers, geometric isomers, tautomers or isomer mixtures, or its pharmaceutically acceptable salts, characterized in that: R1 is substituted or unsubstituted C4-C 10 Alkyl, substituted or unsubstituted C 12 Alkyl, substituted or unsubstituted C 14 Alkyl, substituted or unsubstituted C 16 -C 20 Alkyl, substituted or unsubstituted C3-C 17 Alkenyl, substituted or unsubstituted C3-C 17 Alkynyl, substituted or unsubstituted C5-C6 cycloalkyl, wherein the "substituted" means that the group contains 1 or 2 R a The substituent, R a Selected from deuterium, hydroxyl, amino, cyano, nitro, halogen, C1-C3 alkoxy, C1-C3 alkoxycarbonyl, C5-C6 cycloalkyl, -NHCOOR a1 、C6-C 10 Aryl, saturated or unsaturated five- to six-membered heterocyclic group containing 1 or 2 heteroatoms selected from N, O, and S, five- to six-membered heteroaryl group containing 1 or 2 heteroatoms selected from N, O, and S, unsubstituted or with 1 to 2 R a2 Substituted steroid structure, the R a1 is selected from C1-C4 alkyl, said R a2 Selected from hydroxyl group, C2-C6 ester group.

3. The compound derivative represented by formula I or its isotope-labeled compound according to claim 1, or its optical isomers, geometric isomers, tautomers or isomer mixtures, or its pharmaceutically acceptable salts, characterized in that: More preferably, R1 is a substituted or unsubstituted C7-C 10 Alkyl, substituted or unsubstituted C 12 Alkyl, substituted or unsubstituted C 14 Alkyl, substituted or unsubstituted C 16 -C 17 Alkyl, substituted or unsubstituted C8-C 17 Alkenyl, substituted or unsubstituted C8-C 17 Alkynyl, substituted or unsubstituted C3-C6 cycloalkyl, wherein the "substituted" means that the group contains 1 or 2 R a The substituent, R a Selected from deuterium, hydroxyl, amino, cyano, nitro, halogen, methoxy, ethoxy, propoxy, methoxycarbonyl, ethoxycarbonyl, propoxycarbonyl, cyclopentyl, cyclohexyl, -NHCOOR a1 , phenyl, naphthyl, a saturated or unsaturated five- to six-membered heterocyclic group containing 1 or 2 heteroatoms selected from N, O, and S, a five- to six-membered heteroaryl group containing 1 or 2 heteroatoms selected from N, O, and S, unsubstituted or replaced by 1 or 2 R a2 Substituted steroid structure, the R a1 is selected from methyl, ethyl, n-propyl, isopropyl, n-butyl, isobutyl, tert-butyl, said R a2 Selected from hydroxyl, CH3COO-, CH3CH2COO-, CH3CH2CH2COO-; More preferably, the substituent R a wherein the residue is unsubstituted or is substituted with 1 or 2 R a2 The substituted steroid is selected from the following structures:

4. The compound derivative represented by formula I or its isotope-labeled compound according to claim 1, or its optical isomers, geometric isomers, tautomers or isomer mixtures, or its pharmaceutically acceptable salts, characterized in that: R2 is a substituted or unsubstituted C1-C4 alkyl group, wherein the "substituted" means that the group contains 1 or 2 R b substituted, the R b Selected from deuterium, hydroxy, amino, cyano, nitro, halogen, C1-C3 alkoxy, C1-C3 alkyl substituted by halogen, and C3-C6 cycloalkyl.

5. The compound derivative represented by formula I or its isotope-labeled compound according to claim 1, or its optical isomers, geometric isomers, tautomers or isomer mixtures, or its pharmaceutically acceptable salts, characterized in that: More preferably, R2 is selected from methyl, ethyl, n-propyl, isopropyl, n-butyl, isobutyl, tert-butyl, n-pentyl, isopentyl, n-hexyl, cyclopropyl, cyclobutyl, cyclopentyl, cyclohexyl; More preferably, R2 is isopropyl.

6. The compound derivative represented by formula I according to claim 1 or its isotope-labeled compound, or its optical isomers, geometric isomers, tautomers or isomer mixtures, or its pharmaceutically acceptable salts are selected from the following structures:

7. Use of a compound derivative represented by formula I according to any one of claims 1 to 6, or an isotope-labeled compound thereof, or an optical isomer, geometric isomer, tautomer or isomer mixture thereof, or a pharmaceutically acceptable salt thereof as a prodrug of N4-hydroxycytidine.

8. A pharmaceutical composition comprising a therapeutically effective amount of a compound derivative represented by formula I according to any one of claims 1 to 6 or an isotope-labeled compound thereof, or an optical isomer, geometric isomer, tautomer or isomer mixture thereof, or a pharmaceutically acceptable salt thereof, and a pharmaceutically acceptable excipient.

9. Use of a compound derivative represented by formula I according to any one of claims 1 to 6, or an isotope-labeled compound thereof, or an optical isomer, geometric isomer, tautomer or isomer mixture thereof, or a pharmaceutically acceptable salt thereof in the preparation of a broad-spectrum antiviral or antibacterial drug; Preferably, the broad-spectrum antiviral or antibacterial drug is used to treat or prevent Gram-positive bacterial infection; Preferably, the Gram-positive bacteria include but are not limited to Staphylococcus, Streptococcus, Pneumococcus, Bacillus anthracis, Corynebacterium diphtheriae, and Corynebacterium tetani; Preferably, the broad-spectrum antiviral or antibacterial drug is prepared into an inhalant, a patch, a transdermal agent, an oral immediate-release preparation, a sustained-release or controlled-release preparation, a pill, a capsule, or a granule; Preferably, the inhalant includes a dry powder inhaler, a solution inhaler, or a vortex inhaler.

10. A method for treating or preventing a disease caused by a viral or bacterial infection, the method comprising administering to a subject a therapeutically effective amount of a compound derivative represented by formula I according to any one of claims 1 to 6, or an isotope-labeled compound thereof, or an optical isomer, geometric isomer, tautomer or isomer mixture thereof, or a pharmaceutically acceptable salt thereof, or a pharmaceutical composition according to claim 8.

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