Preparation of 2'-deoxy-2',2'-difluorocytidine carbonate hydrazide and its use

Abstract

This invention relates to a series of derivatives of 2'-deoxy-2',2'-difluorocytidine (dFdC, gemcitabine) carbonate hydrazide esters, methods for preparing pharmaceutically acceptable salts thereof, and their applications in the therapeutic field. Furthermore, this invention provides a series of novel, easily prepared, and highly active antitumor 2'-deoxy-2',2'-difluorocytidine 5'-carbonic acid hydrazide ester derivatives (Formula I) and 2'-deoxy-2',2'-difluorocytidine 3'-carbonic acid hydrazide ester derivatives (Formula II). Their structures are primarily derived from the carbonate hydrazide ester at the 5'- or 3'-hydroxyl groups in the 2'-deoxy-2',2'-difluorocytidine structure. These compounds are easy to prepare, have good solubility, are stable in blood circulation, exhibit little or no activity in vitro, and release the original drug in vivo to exert therapeutic effects. They can be used as monotherapy, in combination therapy, or as antibody-drug conjugates for the treatment of cancer diseases.

Description

Technical Field

[0001] This invention relates to the pharmaceutical field. Specifically, this invention provides a series of novel preparations and applications of 2'-deoxy-2',2'-difluorocytidine carbonate hydrazides. Background Technology

[0002] 2'-Deoxy-2',2'-Difluorocytidine is a cytosine nucleoside derivative that is widely used in the treatment of various tumors.

[0003] However, severe drug resistance often occurs during the use of 2'-deoxy-2',2'-difluorocytidine. For example, the lack of nucleoside transporters (NTs) prevents the drug from entering tumor cells, the lack of deoxycytosine kinase (dCK) prevents the drug from forming the key dFdCMP, and the action of deoxycytidine deaminase (dCDA) leads to the deamination of the drug to form ineffective dFdU.

[0004] Therefore, there is an urgent need to develop a 2'-deoxy-2',2'-difluorocytidine derivative with high antitumor activity. Summary of the Invention

[0005] Based on the characteristic that hydrazine structures are easily oxidized and broken in tumor cells, this invention involves esterifying 2'-deoxy-2',2'-difluorocytidine at the 3' or 5'-OH group to form a series of 2'-deoxy-2',2'-difluorocytidine-3' or 5'-hydrazide ester derivatives. These compounds can circumvent deoxycytosine kinase resistance and escape the action of deoxycytidine deaminase, and instead undergo hydrazine oxidation within tumor cells to release active drug molecules and exert antitumor effects. Given the high antitumor activity and low toxicity of these derivatives, they can be used as single agents or in combination for tumor treatment.

[0006] In a first aspect of the invention, a 2'-deoxy-2',2'-difluorocytidine carbonate hydrazide derivative compound, its stereoisomers, prodrugs, or pharmaceutically acceptable salts thereof are provided, characterized in that the compound has a structure as shown in Formula I or Formula II:

[0007]

[0008] in,

[0009] R1 is selected from the following group: H, carboxyl, C1-C6 alkyl, C2-C6 alkenyl, C2-C6 alkynyl, C1-C6 haloalkyl, C1-C6 acyl, carboxylC1-C6 alkyl, C3-C8 saturated or partially unsaturated carbocyclic, 5-12 membered saturated or partially unsaturated heteroalkyl, 6-10 membered aryl, 5-12 membered heteroaryl. Wherein, R3 and R4 are selected from the following group: H, C1-C3 alkyl; n = 0, 1, 2, 3 or 4;

[0010] R2 is selected from the following group: H, C1-C6 acyl group;

[0011] In this group, R1, R2, R3, and R4 are each independently substituted by one or more substituents selected from the following group: halogen, cyano, hydroxyl, amino, carboxyl, C1-C6 alkyl, C2-C6 alkenyl, C2-C6 alkynyl, C1-C6 haloalkyl, C1-C6 alkoxy, C1-C6 haloalkoxy, C1-C6 acyl, carboxyl C1-C6 alkyl, C1-C6 alkylamine, C3-C8 saturated or partially unsaturated carbocyclic, 5-12 saturated or partially unsaturated heteroalkyl, 6-10 aryl, and 5-12 heteroaryl.

[0012] Furthermore, the stereoisomers include geometric isomers and optical isomers.

[0013] In some embodiments, R1 is selected from the group consisting of: H, C1-C6 alkyl, C3-C8 saturated or partially unsaturated carbocyclic groups, 5-12 membered saturated or partially unsaturated heterogroups, 6-10 membered aryl, and 5-12 membered heteroaryl.

[0014] The definitions of R3, R4, and n are as described above.

[0015] In some embodiments, R2 is selected from the group consisting of H, C1-C6 acyl groups.

[0016] In another preferred embodiment, R2 is H.

[0017] In some embodiments, R1 is selected from the group consisting of: H, phenyl,

[0018] In some implementations, R3 and R4 are H.

[0019] In some embodiments, the compounds are selected from the group consisting of:

[0020]

[0021]

[0022] In a second aspect of the invention, a method for preparing a compound as described in the first aspect of the invention is provided, characterized by comprising the steps of:

[0023] 1) Reaction of 2'-deoxy-2',2'-difluorocytidine with Boc-anhydride yields the corresponding 3',4-di-tert-butoxycarbonyl-2'-deoxy-2',2'-difluorocytidine;

[0024] 2) 3',4-di-tert-butoxycarbonyl-2'-deoxy-2',2'-difluorocytidine and N,N'-carbonyldiimidazole CDI were reacted, and then reacted with the corresponding hydrazine to obtain the corresponding 3',4-di-tert-butoxycarbonyl-2'-deoxy-2',2'-difluorocytidine-5-carbonate hydrazine derivatives.

[0025] 3) Reaction of 3',4-di-tert-butoxycarbonyl-2'-deoxy-2',2'-difluorocytidine-5'-carbonate hydrazide with trifluoroacetyl yields a 2'-deoxy-2',2'-difluorocytidine-5'-carbonate hydrazide derivative.

[0026] 4) Reaction of 2'-deoxy-2',2'-difluorocytidine with Boc-anhydride yields the corresponding 4-tert-butoxycarbonyl-2'-deoxy-2',2'-difluorocytidine;

[0027] 5) 4-tert-butoxycarbonyl-2'-deoxy-2',2'-difluorocytidine was reacted with TBDMS-Cl to give the corresponding product 5'-TBDMS-4-tert-butoxycarbonyl-2'-deoxy-2',2'-difluorocytidine;

[0028] 6) 5'-TBDMS-4-tert-butoxycarbonyl-2'-deoxy-2',2'-difluorocytidine reacts with CDI, and then reacts with the corresponding substituted hydrazine compounds to give the corresponding 5'-TBDMS-4-tert-butoxycarbonyl-2'-deoxy-2',2'-difluorocytidine-3'-carbonate hydrazine derivatives.

[0029] 7) 5'-TBDMS-4-tert-butoxycarbonyl-2'-deoxy-2',2'-difluorocytidine-3'-carbonate hydrazide was reacted with TBAF to give 4-tert-butoxycarbonyl-2'-deoxy-2',2'-difluorocytidine-3'-carbonate hydrazide;

[0030] 8) React 4-tert-butoxycarbonyl-2'-deoxy-2',2'-difluorocytidine-3'-carbonate hydrazide with trifluoroacetyl to obtain a 2'-deoxy-2',2'-difluorocytidine-3'-carbonate hydrazide derivative.

[0031] In some embodiments, the amount of Boc-anhydride used in step 1) is 2.1-2.4 molar equivalents relative to 2'-deoxy-2',2'-difluorocytidine.

[0032] In another preferred embodiment, the amount of CDI used in step 2) is 1-2 molar equivalents relative to 3',4-di-tert-butoxycarbonyl-2'-deoxy-2',2'-difluorocytidine.

[0033] In another preferred embodiment, the amount of CDI used in step 2) is 1.2-1.5 molar equivalents relative to 3',4-di-tert-butoxycarbonyl-2'-deoxy-2',2'-difluorocytidine.

[0034] In another preferred embodiment, the amount of the hydrazine compound used in step 2) is 1-2 molar equivalents relative to 3',4-di-tert-butoxycarbonyl-2'-deoxy-2',2'-difluorocytidine.

[0035] In another preferred embodiment, the amount of the hydrazine compound used in step 2) is 1.2-1.5 molar equivalents relative to 3',4-di-tert-butoxycarbonyl-2'-deoxy-2',2'-difluorocytidine.

[0036] In another preferred embodiment, the trifluoroacetic acid described in step 3) is a 1-30% DCM solution.

[0037] In another preferred embodiment, the trifluoroacetic acid described in step 3) is a 5-10% DCM solution.

[0038] In another preferred embodiment, the amount of Boc-anhydride used in step 4) is 1-1.1 molar equivalents relative to 2'-deoxy-2',2'-difluorocytidine.

[0039] In another preferred embodiment, the amount of TBDMS-Cl used in step 5) is 1-1.2 molar equivalents relative to 4-tert-butoxycarbonyl-2'-deoxy-2',2'-difluorocytidine.

[0040] In another preferred embodiment, the amount of TBAF used in step 6) is 1.2-1.5 molar equivalents relative to 5'-TBDMS-4-tert-butoxycarbonyl-2'-deoxy-2',2'-difluorocytidine.

[0041] In another preferred embodiment, the trifluoroacetic acid described in step 7) is a 1-30% DCM solution.

[0042] In another preferred embodiment, the trifluoroacetic acid described in step 7) is a 5-10% DCM solution.

[0043] In a third aspect of the invention, a pharmaceutical composition is provided, characterized in that the pharmaceutical composition comprises one or more of the following: a compound, a stereoisomer thereof, a prodrug, or a pharmaceutically acceptable salt thereof, or a mixture thereof, as described in the first aspect of the invention, and one or more pharmaceutically acceptable carriers, excipients, adjuvants, excipients, and / or diluents.

[0044] In a third aspect of the invention, there is provided the use of a compound, its stereoisomer, prodrug, or a pharmaceutically acceptable salt thereof as described in the first aspect of the invention, or a pharmaceutical composition as described in the third aspect of the invention, characterized in that it is used to treat tumor diseases.

[0045] It should be understood that, within the scope of this invention, the above-described technical features of this invention and the technical features specifically described below (such as in the embodiments) can be combined with each other to form new or preferred technical solutions. Due to space limitations, they will not be described in detail here. Detailed Implementation

[0046] Through extensive and in-depth research, the inventors unexpectedly discovered a series of phosphate ester amide derivatives of 2'-deoxy-2',2'-difluorocytidine. Bioactivity tests revealed that these derivatives exhibit high antitumor activity and low toxicity. Based on this, the present invention was completed.

[0047] definition

[0048] Unless otherwise defined, all technical and scientific terms used herein have the same meaning as commonly understood by one of ordinary skill in the art to which this invention pertains.

[0049] As used herein, the term "alkyl" includes straight-chain or branched alkyl groups. For example, C1-C6 alkyl groups refer to straight-chain or branched alkyl groups having 1-6 carbon atoms, such as methyl, ethyl, propyl, isopropyl, butyl, isobutyl, tert-butyl, etc.

[0050] As used herein, the term "alkenyl" includes straight-chain or branched alkenyl groups. For example, C2-C6 alkenyl refers to straight-chain or branched alkenyl groups having 2-6 carbon atoms, such as vinyl, allyl, 1-propenyl, isopropenyl, 1-butenyl, 2-butenyl, or similar groups.

[0051] As used herein, the term "alkynyl" includes straight-chain or branched alkynyl groups. For example, C2-C6 alkynyl refers to straight-chain or branched alkynyl groups having 2-6 carbon atoms, such as ethynyl, propynyl, butynyl, or similar groups.

[0052] As used herein, the term "cycloalkyl" refers to a cyclic saturated aliphatic hydrocarbon group having a specific number of carbon atoms. For example, C3-C 10 Alkenyl groups refer to cyclic saturated aliphatic hydrocarbon groups having 3-10 carbon atoms. They can be monocyclic, such as cyclopropyl, cyclobutyl, cyclopentyl, cyclohexyl, or similar groups. They can also be bicyclic, such as bridged or spirocyclic forms.

[0053] As used herein, the term "alkylamino" refers to an amino group substituted with an alkyl group. For example, "C1-C6 alkylamino" refers to an amino group substituted with a C1-C6 alkyl group, which can be monosubstituted or disubstituted; for example, methylamino, ethylamino, propylamino, isopropylamino, butylamino, isobutylamino, tert-butylamino, dimethylamino, diethylamino, dipropylamino, diisopropylamino, dibutylamino, diisobutylamino, ditert-butylamino, etc.

[0054] As used herein, the term "alkoxy" refers to a group having an alkyl-oxy group structure. For example, "C1-C6 alkoxy" refers to a straight-chain or branched alkoxy group having 1-6 carbon atoms, including methoxy, ethoxy, propoxy, isopropoxy, butoxy, isobutoxy, tert-butoxy, etc.

[0055] As used herein, the term "haloalkyl" refers to an alkyl group in which one or more hydrogen atoms are replaced by a halogen, wherein the definition of alkyl is as described above.

[0056] As used herein, the term "haloalkoxy" refers to an alkoxy group in which one or more hydrogen atoms are replaced by a halogen, wherein the definition of alkoxy is as described above.

[0057] As used herein, the term "heterocyclic group" or "heterocyclic alkyl group" refers to a saturated or partially saturated cyclic group having a specific number of ring atoms (e.g., 3-10 ring atoms), wherein 1-3 of these atoms are heteroatoms selected from N, S, and O. It can be monocyclic, bicyclic, or polycyclic, such as bridged or spirocyclic forms. Specific examples include oxobutyranyl, azabutyranyl, tetrahydro-2H-pyranyl, piperidinyl, tetrahydrofuranyl, morpholinyl, and pyrrolidinyl, etc.

[0058] As used in this article, the term "C6-C" 10 "Aryl" refers to an aryl group having 6-10 carbon atoms, such as phenyl or naphthyl groups.

[0059] As used herein, the term "5-12-membered heteroaryl" refers to a cyclic aromatic group having 5-12 atoms, of which 1-3 atoms are heteroatoms selected from the group consisting of N, S, and O. It can be monocyclic or fused-ring. Specific examples include pyridyl, pyridinyl, pyrimidinyl, pyrazinyl, triazinyl, pyrroleyl, pyrazolyl, imidazoleyl, (1,2,3)-triazolyl and (1,2,4)-triazolyl, tetrazolyl, furanyl, thiophenyl, isoxazolyl, thiazolyl, oxazolyl, etc.

[0060] Unless otherwise specified as "substituted or unsubstituted", the groups described in this invention may be substituted by substituents selected from the group consisting of: halogen, nitrile, nitro, hydroxyl, amino, C1-C6 alkyl-amino, C1-C6 alkyl, C2-C6 alkenyl, C2-C6 alkynyl, C1-C6 alkoxy, halo-C1-C6 alkyl, halo-C2-C6 alkenyl, halo-C2-C6 alkynyl, halo-C1-C6 alkoxy, allyl, benzyl, C6-C 12 Aryl, C1-C6 alkoxy-C1-C6 alkyl, C1-C6 alkoxy-carbonyl, phenoxycarbonyl, C2-C6 alkynyl-carbonyl, C2-C6 alkenyl-carbonyl, C3-C6 cycloalkyl-carbonyl, C1-C6 alkyl-sulfonyl, etc.

[0061] As used herein, "halogen" or "halogen atom" refers to F, Cl, Br, and I. More preferably, the halogen or halogen atom is selected from F, Cl, and Br. "Halogenated" means substituted by an atom selected from F, Cl, Br, and I.

[0062] Unless otherwise specified, the structural formulas described in this invention are intended to include all isomers (such as enantiomers, diastereomers, and geometric isomers (or conformational isomers)): for example, R and S configurations containing an asymmetric center, (Z) and (E) isomers with double bonds, etc. Therefore, any single stereochemical isomer of the compounds of this invention, or a mixture of its enantiomers, diastereomers, or geometric isomers (or conformational isomers), is within the scope of this invention.

[0063] As used herein, the term "tautomer" refers to structural isomers with different energies that can cross a low energy barrier and thus interconvert. For example, proton tautomers (i.e., proton shifts) include interconversion via proton migration, such as 1H-indazole and 2H-indazole. Valence tautomers include interconversion via some bonding electron recombination.

[0064] As used herein, the term "solvent complex" refers to a complex of the compound of the present invention coordinated with a solvent molecule in a specific ratio.

[0065] As used herein, the term "hydrate" refers to a complex formed by the coordination of the compound of the present invention with water.

[0066] The term "pharmaceutically acceptable salt" refers to a salt formed by the compounds of the present invention with an acid or base that is suitable for use as a medicine. Pharmaceutically acceptable salts include both inorganic and organic salts. A preferred class of salts are those formed by the compounds of the present invention with an acid. Acids suitable for forming salts include, but are not limited to: inorganic acids such as hydrochloric acid, hydrobromic acid, hydrofluoric acid, sulfuric acid, nitric acid, and phosphoric acid; organic acids such as formic acid, acetic acid, trifluoroacetic acid, propionic acid, oxalic acid, malonic acid, succinic acid, fumaric acid, maleic acid, lactic acid, malic acid, tartaric acid, citric acid, picric acid, benzoic acid, methanesulfonic acid, ethanesulfonic acid, p-toluenesulfonic acid, benzenesulfonic acid, and naphthalenesulfonic acid; and amino acids such as proline, phenylalanine, aspartic acid, and glutamic acid. Another preferred class of salts are salts formed by the compounds of the present invention with a base, such as alkali metal salts (e.g., sodium or potassium salts), alkaline earth metal salts (e.g., magnesium or calcium salts), ammonium salts (such as lower alkanol ammonium salts and other pharmaceutically acceptable amine salts), such as methylamine salts, ethylamine salts, propylamine salts, dimethylamine salts, trimethylamine salts, diethylamine salts, triethylamine salts, tert-butylamine salts, ethylenediamine salts, hydroxyethylamine salts, dihydroxyethylamine salts, trihydroxyethylamine salts, and amine salts formed from morpholine, piperazine, and lysine, respectively.

[0067] Active ingredients

[0068] In this invention, an active ingredient with high anti-tumor activity, namely a compound of formula (I) or formula (II), is provided that can be used to treat tumor diseases.

[0069] Experiments have shown that the active ingredients of this invention can effectively inhibit tumor growth, thereby treating tumor diseases.

[0070] It should be understood that the active ingredients of the present invention include compounds represented by formula (I) or formula (II), or pharmaceutically acceptable salts thereof, or prodrugs thereof. It should also be understood that the active ingredients of the present invention further include crystalline forms, amorphous compounds, and deuterated compounds of formula (I) or formula (II).

[0071] Pharmaceutical Compositions and Administration

[0072] Because the compounds of the present invention have excellent antitumor activity, the compounds of the present invention and their various crystal forms, pharmaceutically acceptable inorganic or organic salts, hydrates or solvates, and pharmaceutical compositions containing the compounds of the present invention as the main active ingredient can be used to treat tumor diseases.

[0073] The pharmaceutical compositions of the present invention comprise the compound of the present invention within a safe and effective range and a pharmaceutically acceptable excipient or carrier. "Safe and effective range" refers to an amount of the compound sufficient to significantly improve the condition without causing serious side effects. Typically, the pharmaceutical composition contains 1-2000 mg of the compound of the present invention per dose, more preferably, 10-200 mg of the compound of the present invention per dose. Preferably, "one dose" is one capsule or tablet.

[0074] "Pharmaceutically acceptable carriers" refer to one or more compatible solid or liquid fillers or gelling substances that are suitable for human use and must have sufficient purity and sufficiently low toxicity. "Compatibility" here means that the components in the composition can be mixed with and with the compounds of the present invention without significantly reducing the efficacy of the compounds. Examples of pharmaceutically acceptable carriers include cellulose and its derivatives (such as sodium carboxymethyl cellulose, sodium ethyl cellulose, cellulose acetate, etc.), gelatin, talc, solid lubricants (such as stearic acid, magnesium stearate), calcium sulfate, vegetable oils (such as soybean oil, sesame oil, peanut oil, olive oil, etc.), polyols (such as propylene glycol, glycerin, mannitol, sorbitol, etc.), emulsifiers (such as... Wetting agents (such as sodium dodecyl sulfate), colorants, flavoring agents, stabilizers, antioxidants, preservatives, pyrogen-free water, etc.

[0075] There are no particular limitations on the administration of the compounds or pharmaceutical compositions of the present invention. Representative administration methods include (but are not limited to): oral administration, parenteral administration (intravenous, intramuscular, or subcutaneous).

[0076] Solid dosage forms for oral administration include capsules, tablets, pills, powders, and granules. In these solid dosage forms, the active compound is mixed with at least one conventional inert excipient (or carrier), such as sodium citrate or dicalcium phosphate, or with the following components: (a) fillers or compatibilizers, such as starch, lactose, sucrose, glucose, mannitol, and silica; (b) binders, such as hydroxymethyl cellulose, alginate, gelatin, polyvinylpyrrolidone, sucrose, and gum arabic; (c) humectants, such as glycerin; (d) disintegrants, such as agar, calcium carbonate, potato starch or cassava starch, alginate, certain complex silicates, and sodium carbonate; (e) slowing agents, such as paraffin; (f) absorption accelerators, such as quaternary ammonium compounds; (g) wetting agents, such as cetyl alcohol and glyceryl monostearate; (h) adsorbents, such as kaolin; and (i) lubricants, such as talc, calcium stearate, magnesium stearate, solid polyethylene glycol, sodium dodecyl sulfate, or mixtures thereof. Buffers may also be included in the dosage forms of capsules, tablets, and pills.

[0077] Solid dosage forms such as tablets, sugar pills, capsules, pellets, and granules can be prepared using coatings and shells, such as casings and other materials known in the art. They may contain opacifying agents, and the release of the active compound or compound from such compositions can be delayed in a portion of the digestive tract. Examples of encapsulating components that can be used are polymeric substances and waxes. If necessary, the active compound may also be formed into microcapsules with one or more of the excipients described above.

[0078] Liquid dosage forms for oral administration include pharmaceutically acceptable emulsions, solutions, suspensions, syrups, or tinctures. In addition to the active compound, liquid dosage forms may contain inert diluents conventionally used in the art, such as water or other solvents, solubilizers and emulsifiers, e.g., ethanol, isopropanol, ethyl carbonate, ethyl acetate, propylene glycol, 1,3-butanediol, dimethylformamide, and oils, particularly cottonseed oil, peanut oil, corn germ oil, olive oil, castor oil, and sesame oil, or mixtures of these substances.

[0079] In addition to these inert diluents, the composition may also contain auxiliaries such as wetting agents, emulsifiers and suspending agents, sweeteners, flavoring agents and fragrances.

[0080] In addition to the active compound, the suspension may contain suspending agents such as ethoxylated isooctadecyl alcohol, polyoxyethylene sorbitol and dehydrated sorbitol esters, microcrystalline cellulose, aluminum methoxide and agar, or mixtures of these substances.

[0081] Compositions for parenteral injection may comprise physiologically acceptable sterile aqueous or anhydrous solutions, dispersions, suspensions, or emulsions, and sterile powders for reconstitution into sterile injectable solutions or dispersions. Suitable aqueous and non-aqueous carriers, diluents, solvents, or excipients include water, ethanol, polyols, and suitable mixtures thereof.

[0082] The compounds of this invention can be administered alone or in combination with other pharmaceutically acceptable therapeutic agents.

[0083] When administered in combination, the pharmaceutical composition further comprises one or more (two, three, four, or more) other pharmaceutically acceptable therapeutic agents. One or more (two, three, four, or more) of these other pharmaceutically acceptable therapeutic agents may be used simultaneously, separately, or sequentially with the compounds of the present invention to treat oncological diseases.

[0084] When using the pharmaceutical composition, a safe and effective amount of the compound of the present invention is applied to the mammal (such as a human) requiring treatment. The dosage administered is the pharmaceutically considered effective dose. For a person weighing 60 kg, the daily dose is typically 1–2000 mg, preferably 20–500 mg. Of course, the specific dosage should also take into account factors such as the route of administration and the patient's health condition, which are all within the scope of the skill of a skilled physician.

[0085] The main advantages of this invention include:

[0086] (1) All compounds of the present invention have good antitumor activity.

[0087] (2) The crystal form preparation method of the present invention is simple.

[0088] (3) The compounds of the present invention have good pharmacokinetic properties and good drug-likeness.

[0089] The present invention will be further illustrated below with reference to specific embodiments. It should be understood that these embodiments are for illustrative purposes only and are not intended to limit the scope of the invention. Experimental methods in the following embodiments, unless otherwise specified, are generally performed under conventional conditions or as recommended by the manufacturer. Unless otherwise stated, percentages and parts are weight percentages and parts by weight.

[0090] Unless otherwise defined, all technical and scientific terms used herein have the same meaning as are familiar to those skilled in the art. Furthermore, any methods and materials similar to or equivalent to those described herein may be applied to the methods of this invention. The preferred embodiments and materials described herein are for illustrative purposes only.

[0091] Unless otherwise specified, all experimental materials and reagents used in the following examples are available from commercially available sources.

[0092] Preparation of intermediates 4-BocG and 3',4-dBocG

[0093]

[0094] 4-tert-butoxycarbonyl-2'-deoxy-2',2'-difluorocytidine (4-BocG) and 3',4-di(tert-butoxycarbonyl)-2'-deoxy-2',2'-difluorocytidine (3',4-dBocG) were prepared according to the literature (J.Org.Chem.1999,64(22),8319-8322): 2'-deoxy-2',2'-difluorocytidine (2.0 g, 7.6 mmol) was added to a reaction flask containing DMF (12 mL), and tert-butoxycarbonic anhydride (2.5 g, 11.4 mmol) was added with stirring. The mixture was heated to 50 °C and stirred for 24 hours. After cooling, the mixture was poured into water, the solid was filtered, and recrystallized from ethyl acetate to obtain a white solid product, 4-tert-butoxycarbonyl-2'-deoxy-2',2'-difluorocytidine (2.1 g, yield 77%).

[0095] The above-mentioned 4-tert-butoxycarbonyl-2'-deoxy-2',2'-difluorocytidine (2.5 g, 7.0 mmol), Na2CO3 (0.4 g, 35 mmol) in dioxane, water (5:1, 40 mL) and tert-butoxycarbonic anhydride (1.53 g, 7.0 mmol) were stirred at room temperature for 24 hours. The solvent was concentrated, and the mixture was recrystallized from petroleum ether / ethyl acetate to give the solid product 3',4-bis(tert-butoxycarbonyl)-2'-deoxy-2',2'-difluorocytidine (2.9 g, 92% yield).

[0096] Example 1

[0097] Synthesis of 2'-deoxy-2',2'-difluorocytidine-5'-carbonate hydrazide (1)

[0098]

[0099] 3',4-dBocG (3.0 g, 6.4 mmol), triethylamine (1.8 mL, 12.9 mmol), and CDI (2.1 g, 12.9 mmol) were added to a reaction flask containing DCM (30 mL). The mixture was stirred at room temperature for 4 hours. Then, hydrazine hydrate (0.5 g, 85%, 12.9 mmol) was added, and the mixture was stirred overnight at room temperature. The solvent was removed under reduced pressure, and the mixture was purified by silica gel column chromatography to give compound 3',4-bis(tert-butoxycarbonyl)-2'-deoxy-2',2'-difluorocytidine-5'-carbonate hydrazide (2.1 g, 65% yield).

[0100] Trifluoroacetic acid (4 mL) was added to a solution of 3',4-bis(tert-butoxycarbonyl)-2'-deoxy-2',2'-difluorocytidine-5'-carbonate hydrazide (2.1 g, 4.1 mmol) in DCM (16 mL). The mixture was stirred overnight at room temperature, the solvent was removed under reduced pressure, and the solution was purified by silica gel column chromatography to give a white solid compound 1 (1.0 g, yield 76%, HPLC: 96%). 1 H NMR(500MHz,DMSO-d6)δ8.38(s,1H),7.55-7.34(m,3H),6.46(s,1H),6.18(t,J=8.6Hz,1H), 5.84(d,J=7.9Hz,1H),4.36(d,J=12.4Hz,1H),4.27-4.05(m,4H),3.96(t,J=6.9Hz,1H); 13C NMR (126MHz, DMSO) δ166.13,158.34,155.12,141.55,123.24,95.62,77.97,70.13,69.95,62.57; LCMS: (M+H) + 322.01 (Calculated value: 321.09).

[0101] Example 2

[0102] Synthesis of 2'-deoxy-2',2'-difluorocytidine-5'-carbonate acyl(N'-hydroxyacetyl)hydrazine (2)

[0103] Compound 1 (200 mg, 0.3 mmol) and glycolic acid (30 mg, 0.3 mmol) were added to a reaction flask containing DCM (4 mL). Triethylamine (0.1 mL, 0.7 mmol), DCC (157 mg, 0.7 mmol), and HOBt (103 mg, 0.7 mmol) were added to the reaction flask. The reaction mixture was stirred overnight at room temperature. The solid was filtered off, the filtrate was washed with water, dried over anhydrous sodium sulfate, and purified by silica gel column chromatography to give a white solid compound 3',4-bis(tert-butoxycarbonyl)-2'-deoxy-2',2'-difluorocytidine-5'-carbonyl(N'-hydroxyacetyl)hydrazine (100 mg, 45% yield).

[0104] Trifluoroacetic acid (0.5 mL) was added to a solution of 3',4-bis(tert-butoxycarbonyl)-2'-deoxy-2',2'-difluorocytidine-5'-carbonyl(N'-hydroxyacetyl)hydrazine (85 mg, 0.14 mmol) in DCM (4 mL). The mixture was stirred overnight at room temperature, the solvent was removed under reduced pressure, and the solution was purified by silica gel column chromatography to give a white solid compound 2 (45 mg, 81% yield). 1 H NMR (600MHz, DMSO-d6) δ9.76(d,J=52.8Hz,2H),9.29(d,J=31.8Hz,2H),7.86(d,J=7.8Hz,1H),6.30-6. 10(m,2H),5.03-4.58(m,1H),4.54-4.40(m,1H),4.37-4.01(m,4H),3.94(s,1H),1.11(d,J=4.5Hz,1H); 13 C NMR (151MHz, DMSO) δ172.03,160.12,159.18,156.00,147.44,115.17,95.98,88.41,84.28,78.85,69.70,61.30; LCMS: (M+H) + 379.98 (Calculated value: 379.09).

[0105] Example 3

[0106] Synthesis of 2'-deoxy-2',2'-difluorocytidine-5'-carbonate acyl(N'-p-carboxyphenyl)hydrazine (3)

[0107] 3',4-dBocG (6.0 g, 12.9 mmol), triethylamine (3.6 mL, 25.9 mmol), and CDI (4.2 g, 25.9 mmol) were added to a reaction flask containing THF (45 mL). The mixture was stirred at room temperature for 4 hours. Then, 4-hydrazinobenzoic acid (1.96 g, 12.9 mmol) and DMF (15 mL) were added. The mixture was stirred overnight at room temperature. The THF was removed under reduced pressure, and the solid was poured into water. The solid was filtered off and dissolved in DCM. The solid was purified by silica gel column chromatography to give compound 3',4-bis(tert-butoxycarbonyl)-2'-deoxy-2',2'-difluorocytidine-5'-carbonylacyl(N'-p-carboxyphenyl)hydrazine (2.2 g, 27% yield).

[0108] 3',4-bis(tert-butoxycarbonyl)-2'-deoxy-2',2'-difluorocytidine-5'-carbonyl(N'-p-carboxyphenyl)hydrazine (100 mg, 0.15 mmol) was dissolved in DCM (5 mL), and trifluoroacetic acid (0.5 mL) was added. The mixture was stirred overnight, the solvent was removed under reduced pressure, and the solid compound 3 (53 mg, 77% yield) was purified by silica gel column chromatography. 1 H NMR(500MHz,DMSO-d6)δ9.54(d,J=24.1Hz,2H),9.24-8.98(m,1H),8.48(s,1H),7.97-7.86(m,1H),7.81(d,J =8.5Hz,2H),6.76(dd,J=8.6,1.5Hz,2H),6.32-6.17(m,2H),4.55-4.38(m,3H),4.22(dt,J=32.9,9.0Hz,3H); 13 C NMR (126MHz, DMSO) δ167.75,160.68,159.46,156.74,153.43,143.82,131. 49,120.64,117.81,111.07,95.86,84.05,78.88,70.10,63.19; LCMS: (M+H) + :442.32 (Calculated value: 441.11).

[0109] Example 4

[0110] Synthesis of 2'-deoxy-2',2'-difluorocytidine-5'-carbonate acyl(N'-3-carboxy-6-pyridyl)hydrazine (4)

[0111] 2'-Deoxy-2',2'-difluorocytidine-5'-carbonyl(N'-3-carboxy-6-pyridyl)hydrazine was obtained by the method used to synthesize compound 4 (737 mg, yield 52%, HPLC: 96%); LCMS: (M+H) +:443.02 (Calculated value: 442.34).

[0112] Example 5

[0113] Synthesis of 2'-deoxy-2',2'-difluorocytidine-5'-carbonate acyl-N'-(aminoethylcarbamoyl-4-phenyl)hydrazine (5)

[0114] The synthesized 3',4-bis(tert-butoxycarbonyl)-2'-deoxy-2',2'-difluorocytidine-5'-carbonyl (N'-p-carboxyphenyl)hydrazine (2.06 g, 3.2 mmol) was dissolved in DCM (30 mL), and mono-Boc ethylenediamine (0.6 g, 3.8 mmol), triethylamine (0.9 mL, 6.4 mmol), DCC (1.3 g, 6.4 mmol), and HOBt (0) were added. 0.86 g (6.4 mmol), the reaction solution was stirred at room temperature for 15 hours, the solid was filtered off, the filtrate was washed three times with water, dried over anhydrous sodium sulfate, concentrated and purified by silica gel column chromatography to give a pale yellow solid compound 3',4-bis(tert-butoxycarbonyl)-2'-deoxy-2',2'-difluorocytidine-5'-carbonyl (N'-Boc-ethylenediaminecarbamoyl-4-phenyl)hydrazine (2.1 g, yield 84%, HPLC: 96%).

[0115] The above-mentioned 3',4-bis(tert-butoxycarbonyl)-2'-deoxy-2',2'-difluorocytidine-5'-carbonyl(N'-Boc-ethylenediaminecarbamoyl-4-phenyl)hydrazine (2.1 g, 2.7 mmol) was dissolved in DCM (15 mL), and trifluoroacetic acid (4 mL) was added. The mixture was stirred overnight at room temperature, the solvent was concentrated under reduced pressure, and the solution was purified by silica gel column chromatography to obtain 2'-deoxy-2',2'-difluorocytidine-5'-carbonyl-N'-(aminoethylaminecarbamoyl-4-phenyl)hydrazine (400 mg, yield 31%, HPLC: 98%). 1H NMR(500MHz,DMSO-d6)δ9.42(s,1H),8.35(t,J=5.6Hz,1H),8.25(s,1H),7.92(s,3H),7.71(d, J=8.3Hz,2H),7.55(d,J=7.1Hz,2H),7.45(s,1H),6.70(d,J=8.3Hz,2H),6.54(d,J=6.5Hz,1H), 6.23(t,J=8.6Hz,1H),5.85(d,J=7.5Hz,1H),4.44(d,J=12.4Hz,1H),4.31(dd,J=12.4,5.4Hz,1 H), 4.18 (t, J = 12.1Hz, 1H), 4.02 (t, J = 7.1Hz, 1H), 3.48 (t, J = 6.0Hz, 2H), 2.98 (t, J = 6.2Hz, 2H); 13 C NMR (126MHz, DMSO) δ167.25,166.05,156.86,155.05,152.37,141.65,128.69,124 .16,118.87,111.01,95.65,84.09,78.02,70.33,63.19,39.32,37.50; LCMS: (M+H) + :484.10 (Calculated value: 483.17).

[0116] Example 6

[0117] Synthesis of 2'-deoxy-2',2'-difluorocytidine-5'-carbonate acyl-N'-(4-aminobutyrcarbamoyl-4-phenyl)hydrazine (7)

[0118] Triethylamine (0.013 μL, 0.93 mmol), 1-N-Boc-butanediamine (105.64 mg, 0.56 mmol), dichloromethane (6 mL), 3',4-bis(tert-butoxycarbonyl)-2'-deoxy-2',2'-difluorocytidine-5'-carbonyl(N'-p-carboxyphenyl)hydrazine (0.3 g, 0.467 mmol, Example 3), DCC (192.9 mg, 0.93 mmol), and HOBT (143.2 mg, 0.93 mmol) were added to the reaction flask. The reaction mixture was stirred at room temperature for 12 h, the solvent was removed under reduced pressure, and the product was purified by silica gel column chromatography (DCM / MeOH = 10:1) to give the Boc-protected product (0.18 g, yield 48%).

[0119] The above Boc-protected product was dissolved in DCM (4 mL), TFA (1 mL) was added, the mixture was stirred at room temperature for 2 hours, the solvent was removed under reduced pressure, methyl tert-butyl ether was added, the precipitated solid was filtered, and dried to obtain product 7 (70 mg, yield 62%). 1 H NMR(500MHz,DMSO-d6)δ9.51(s,1H),9.33(d,J=5.5Hz,1H),8.12(d,J=5.7Hz,1H), 7.83-7.64(m,4H),7.60(d,J=8.1Hz,2H),6.60(d,J=8.3Hz,2H),6.17-5.99(m,2H), 4.35(d,J=12.5Hz,1H),4.26(dd,J=12.4,5.6Hz,2H),4.12(h,J=7.1Hz,2H),4.03(t ,J=6.6Hz,1H),3.16(q,J=6.0Hz,2H),2.73(q,J=6.3Hz,2H),1.47(d,J=5.4Hz,4H); 13 C NMR (126MHz, DMSO) δ166.53,161.05,159.29,156.82,152.06,143.76,128.95,124.87,118.08 ,115.73,111.06,95.88,84.28,78.87,70.13,63.14,39.14,38.73,26.77,25.04; LCMS: (M+H) + :512.17 (Calculated value: 511.20).

[0120] Example 7

[0121] Synthesis of 2'-deoxy-2',2'-difluorocytidine-5'-carbonate acyl-N'-(hydrazinoyl-4-phenyl)hydrazine (8)

[0122] The synthesized 3',4-bis(tert-butoxycarbonyl)-2'-deoxy-2',2'-difluorocytidine-5'-carbonyl (N'-p-carboxyphenyl)hydrazine (2.4 g, 3.7 mmol) was dissolved in DCM (30 mL), and mono-Boc-hydrazine (0.59 g, 4.5 mmol), triethylamine (1.0 mL, 7.4 mmol), DCC (1.5 g, 7.4 mmol), and HOBt (1.0 g, 7.4 mmol) were added. The reaction mixture was stirred overnight at room temperature. The solid was filtered off, and the filtrate was washed three times with water, dried over anhydrous sodium sulfate, concentrated, and purified by silica gel column chromatography to give the pale yellow solid compound 3',4-bis(tert-butoxycarbonyl)-2'-deoxy-2',2'-difluorocytidine-5'-carbonyl (N'-Boc-hydrazylformyl-4-phenyl)hydrazine (2.4 g, yield 87%, HPLC: 98%).

[0123] Trifluoroacetic acid (5 mL) was added to a solution containing 20 mL of DCM containing 3',4-bis(tert-butoxycarbonyl)-2'-deoxy-2',2'-difluorocytidine-5'-carbonyl(N'-Boc-hydrazylformyl-4-phenyl)hydrazine (0.4 g, 3.1 mmol). The mixture was stirred at room temperature for 12 hours, the solvent was removed under reduced pressure, and the solution was purified by silica gel column chromatography to obtain 2'-deoxy-2',2'-difluorocytidine-5'-carbonyl(N'-hydrazylformyl-4-phenyl)hydrazine (800 mg, yield 56%, HPLC 98%). 1 H NMR (500MHz, DMSO-d6) δ9.41(d,J=23.3Hz,2H),8.18(s,1H),7.66(d,J=8.3Hz,2H),7.57-7.40(m,3H),6.67(d,J=8.3Hz ,2H),6.49(d,J=6.5Hz,1H),6.23(s,1H),5.84(d,J=7.4Hz,1H),4.49-4.26(m,4H),4.17(t,J=14.8Hz,1H),4.02(s,1H); 13 C NMR (126MHz, DMSO) δ166.62,166.13,156.87,155.10,152.05,141.57,128. 74,123.54,123.21,111.12,95.62,77.96,70.31,70.13,63.14; LCMS: (M+H) + :456.39 (Calculated value: 455.14).

[0124] Example 8

[0125] Synthesis of 2'-deoxy-2',2'-difluorocytidine-3'-carbonate hydrazide (11)

[0126] TBDMS-Cl (2.27 g, 15.1 mmol) was added to a reaction flask containing intermediate 4-BocG (5 g, 13.7 mmol), imidazole (1.87 g, 27.5 mmol), and DMF (40 mL). The mixture was stirred overnight at room temperature, poured into water, extracted twice with methyl tert-butyl ether, dried over anhydrous sodium sulfate, and purified by silica gel column chromatography to obtain 4-Boc-5-TBDMS-gemcitabine (4.8 g, 74% yield).

[0127] Acetonitrile (5 mL), triethylamine (0.26 mL, 1.86 mmol), 4-Boc-5-TBDMS-gemcitabine (0.3 g, 0.62 mmol), and di(N-hydroxysuccinimide) carbonate (0.24 g, 0.93 mmol) were added to a reaction flask. The mixture was stirred overnight at room temperature. After removing the solvent under reduced pressure, the mixture was extracted with ethyl acetate, washed with saturated brine, and dissolved in DCM (4 mL) to remove the solvent. Hydrazine (35 μL, 0.7 mmol) was added, and the mixture was stirred for 10 min at room temperature. After removing the solvent under reduced pressure, the mixture was purified by silica gel column chromatography to obtain 4-Boc-5-TBDMS-gemcitabine-3-carbonate hydrazide (0.1 g, 32% yield).

[0128] The above compound 4-Boc-5-TBDMS-gemcitabine-3-carbonate hydrazide (0.1 g, 0.18 mmol) was dissolved in methanol (2 mL), concentrated hydrochloric acid (0.1 mL) was added, and the mixture was stirred at room temperature for 2 h. The solvent was removed under reduced pressure, and the crude product was purified by silica gel column chromatography to obtain product 11 (0.04 g, yield 65%). 1 H NMR (500MHz, DMSO-d6) δ11.03(s,1H),10.31(s,1H),9.07(d,J=13.2Hz,1H),8.15(d,J=7.7Hz,1H),7.56-7.26(m,1H),6.34(d,J=7 .8Hz,1H),6.22(q,J=9.5,8.7Hz,1H),5.38-5.32(m,1H),4.33-4.17(m,2H),3.77(td,J=12.6,3.0Hz,2H),3.65(d,J=13.0Hz,1H); 13 C NMR (126MHz, DMSO) δ160.06,154.42,147.25,144.46,121.75,102.87,95.51,79.74,71.22,59.37; LCMS: (M+H) + 322.21 (Calculated value: 321.09).

[0129] Test example: Inhibition of tumor cell growth activity

[0130] Human esophageal cancer cells OE33 (human breast adenocarcinoma cells SK-BR-3, or human gastric cancer cells NCI-N87) were cultured in RPMI1640 (Cellmax) containing 10% fetal bovine serum (Cellmax). Tumor cells in the exponential growth phase were diluted with culture medium to 1×10⁵ cells / mL, and 100 μL was added to each well of a 96-well cell culture plate. The plates were then incubated overnight at 37°C with 5% CO₂. The next day, the compound was diluted with culture medium to 10000 nM, 2000 nM, 400 nM, 80 nM, 16 nM, 3.2 nM, 0.64 nM, and 0.13 nM. 2 μL of the diluted compound was added to each well of a 96-well cell culture plate, with three replicates for each concentration. 2 μL of the diluted compound was added to each well of the negative control and blank control group (no compound added). After sample addition, the plate was returned to an incubator at 37°C and 5% CO2 for 72 hours of incubation. After incubation, the cell culture plate was removed, and the culture medium was aspirated from the plate using a pipette. 100 μL of medium containing 10% CCK-8 was added to each well, and the plate was incubated at 37°C for 3 hours. After incubation, the plate was removed, protected from light, and placed in a microplate. The absorbance was measured using 630 nm as the reference wavelength and 450 nm as the measurement wavelength. Based on the absorbance values, the IC50 value was calculated using four-parameter regression in GraphPad (Table 2).

[0131] Table 2. IC50s (nM) values ​​of some compounds inhibiting tumor cell growth

[0132]

[0133]

[0134] All documents mentioned in this invention are incorporated herein by reference as if each document were individually incorporated by reference. Furthermore, it should be understood that after reading the foregoing teachings of this invention, those skilled in the art can make various alterations or modifications to this invention, and these equivalent forms also fall within the scope defined by the appended claims.

Claims

1. A 2'-deoxy-2',2'-difluorocytidine carbonate hydrazide derivative compound or a pharmaceutically acceptable salt thereof, characterized in that, The compound has a structure as shown in Formula I or Formula II: (AND) (II) in, In equation (I), R1 is selected from the following group: Where R3 and R4 are H; n = 0, 1, 2, 3 or 4; in equation (II), R1 is H; R2 is H.

2. The compound of claim 1 or a pharmaceutically acceptable salt thereof, characterized in that, Compound I is selected from , or ; Compounds of Formula II are selected from .

3. The method for preparing the compound according to claim 1, characterized in that, Including the following steps: Formula I is prepared as follows: 1) 2'-deoxy-2',2'-difluorocytidine is reacted with Boc-anhydride to give the corresponding 3',4-di-tert-butoxycarbonyl-2'-deoxy-2',2'-difluorocytidine; 2). 3',4-Di-tert-butoxycarbonyl-2'-deoxy-2',2'-difluorocytidine and N,N'-carbonyldiimidazole CDI were reacted, and then reacted with the corresponding hydrazine to obtain the corresponding 3',4-di-tert-butoxycarbonyl-2'-deoxy-2',2'-difluorocytidine-5'-carbonate hydrazine derivatives. 3). Reaction of 3',4-di-tert-butoxycarbonyl-2'-deoxy-2',2'-difluorocytidine-5'-carbonate hydrazide with trifluoroacetic acid yields a 2'-deoxy-2',2'-difluorocytidine-5'-carbonate hydrazide derivative. Formula II is prepared as follows: 4). 2'-Deoxy-2',2'-Difluorocytidine is reacted with Boc-anhydride to give the corresponding 4-tert-butoxycarbonyl-2'-deoxy-2',2'-difluorocytidine; 5). 4-tert-butoxycarbonyl-2'-deoxy-2',2'-difluorocytidine was reacted with TBDMS-Cl to give the corresponding product 5'-TBDMS-4-tert-butoxycarbonyl-2'-deoxy-2',2'-difluorocytidine; 6) 5'-TBDMS-4-tert-butoxycarbonyl-2'-deoxy-2',2'-difluorocytidine reacts with CDI, and then reacts with the corresponding substituted hydrazine compounds to give the corresponding 5'-TBDMS-4-tert-butoxycarbonyl-2'-deoxy-2',2'-difluorocytidine-3'-carbonate hydrazine derivatives. 7). 5'-TBDMS-4-tert-butoxycarbonyl-2'-deoxy-2',2'-difluorocytidine-3'-carbonate hydrazide was reacted with TBAF to give 4-tert-butoxycarbonyl-2'-deoxy-2',2'-difluorocytidine-3'-carbonate hydrazide; 8) Reaction of 4-tert-butoxycarbonyl-2'-deoxy-2',2'-difluorocytidine-3'-carbonate hydrazide with trifluoroacetic acid yields a 2'-deoxy-2',2'-difluorocytidine-3'-carbonate hydrazide derivative.

4. The method as described in claim 3, characterized in that, The amount of Boc-anhydride used in step 1) is 2.1-2.4 molar equivalents relative to 2'-deoxy-2',2'-difluorocytidine.

5. The method as described in claim 3, characterized in that, The amount of CDI used in step 2) is 1-2 molar equivalents relative to 3',4-di-tert-butoxycarbonyl-2'-deoxy-2',2'-difluorocytidine.

6. The method as described in claim 3, characterized in that, The amount of CDI used in step 2) is 1.2-1.5 molar equivalents relative to 3',4-di-tert-butoxycarbonyl-2'-deoxy-2',2'-difluorocytidine.

7. The method as described in claim 3, characterized in that, The amount of hydrazine used in step 2) is 1-2 molar equivalents relative to 3',4-di-tert-butoxycarbonyl-2'-deoxy-2',2'-difluorocytidine.

8. The method as described in claim 3, characterized in that, The amount of hydrazine used in step 2) is 1.2-1.5 molar equivalents relative to 3',4-di-tert-butoxycarbonyl-2'-deoxy-2',2'-difluorocytidine.

9. The method as described in claim 3, characterized in that, The trifluoroacetic acid mentioned in step 3) is a 1-30% DCM solution.

10. The method as described in claim 3, characterized in that, The trifluoroacetic acid mentioned in step 3) is a 5-10% DCM solution.

11. The method as described in claim 3, characterized in that, The amount of Boc-anhydride used in step 4) is 1-1.1 molar equivalents relative to 2'-deoxy-2',2'-difluorocytidine.

12. The method as described in claim 3, characterized in that, The amount of TBDMS-Cl used in step 5) is 1-1.2 molar equivalents relative to 4-tert-butoxycarbonyl-2'-deoxy-2',2'-difluorocytidine.

13. The method as described in claim 3, characterized in that, The trifluoroacetic acid mentioned in step 8) is a 1-30% DCM solution.

14. The method as described in claim 3, characterized in that, The trifluoroacetic acid mentioned in step 8) is a 5-10% DCM solution.

15. A pharmaceutical composition, characterized in that, The pharmaceutical composition comprises one or more of the compound of claim 1 or a pharmaceutically acceptable salt thereof, or a mixture thereof, and one or more pharmaceutically acceptable excipients.

16. The use of the compound of claim 1 or a pharmaceutically acceptable salt thereof, or the pharmaceutical composition of claim 15, characterized in that, Used to prepare drugs for the treatment of esophageal cancer, breast cancer, or stomach cancer.

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