Compound and application thereof

Abstract

The invention provides a compound with a structure as shown in a formula I, or a stereoisomer, a pharmaceutically acceptable salt, a solvate, a co-crystal or a deuterated substance of the compound. The compound has excellent enzyme inhibition activity, can be developed as a novel anti-CD73 drug, and has a wide application prospect.

Description

Technical Field

[0001] The present invention relates to the field of pharmaceutical technology, in particular to a compound and application thereof. Background Art

[0002] Cancer has become a major public health problem worldwide. According to statistics, in 2021, the number of deaths worldwide exceeded 10 million, and the number of deaths in China exceeded 3 million. At present, the number of tumor incidence and mortality in my country continues to rise, and it has become a veritable "cancer power". The medical expenses caused by malignant tumors exceed 220 billion each year, posing a serious threat to people's lives, health, and social property security.

[0003] Human CD73 is a multifunctional transmembrane glycoprotein composed of 523 amino acids encoded by the NT5E gene (located at 6q14-21), with a relative molecular mass of 70,000, which is anchored to the cell membrane through glycosylphosphatidylinositol (GPI). The CD73 protein consists of three domains. The N-terminal domain (27-317AA) contains two zinc ion binding sites and at least one N-glycosylation site located at aspartic acid 311; the C-terminal domain (337-549AA) contains a substrate binding site and is non-covalently bound to the plasma membrane through the GPI anchor; the N-terminal and C-terminal domains are connected by a short helix (318-336AA), which can control the movement of the enzyme. Therefore, CD73 has two conformations, "open" and "closed", and the catalytic reaction requires CD73 to convert from an open conformation to a closed conformation, the latter of which can expose the active binding site and allow substrate binding.

[0004] Studies have found that CD73 is widely expressed on the surface of various tissue cells in the human body, including lymphocytes, endothelial cells and epithelial cells, and controls a variety of physiological functions, such as epithelial ion exchange, fluid transport, platelet function, tissue hypoxia and vascular leakage. It mainly participates in the following physiological functions: (1) Affecting the salvage synthesis process of purine nucleotides. Adenosine produced by CD73 controls the generation of purine nucleotides through CD39 and CD73, thereby regulating the signal transduction of nucleotides. (2) As a signaling molecule and adhesion molecule, it regulates the interaction between cells and extracellular matrix components (such as laminin and fibronectin), mediating the invasion and metastasis characteristics of cancer. (3) Participating in T cell activation. Adenosine produced by CD73 hydrolysis can activate immune cells, affect the proliferation of immune cells, and regulate CD4+CD25+Treg cells to reduce the immune function of T cells.

[0005] In recent years, immunotherapy has become popular, including immune checkpoint inhibitors (PD-1 / PD-L1), tumor vaccines, cellular immune cell therapy, and nonspecific immunomodulators. As an emerging intervention, immunotherapy changes the patient's immune system to fight cancer by directly stimulating rejection-type processes or blocking inhibitory pathways. A large number of studies have confirmed that tumor cells can express CD73 and release adenosine, and its expression and activity are closely related to tumor invasion and metastasis. Moreover, in tumor cells, the extracellular adenosine produced by CD73 is sufficient to mediate immune escape and promote tumor growth and metastasis. In addition to the immune regulation of tumor cells on CD73, CD73 also affects multiple aspects of tumorigenesis, such as proliferation, adhesion, angiogenesis, and metastasis. It promotes the proliferation of tumor cells by regulating cell cycle, apoptosis, and signaling pathways such as EGFR, β-catenin / cyclin D1, VEGF, and AKT / ERK. Independent of its enzymatic function, CD73 can also promote cell-to-cell adhesion, migration, and cancer cell invasion.

[0006] Currently, there are no drugs targeting CD73 on the market, but the safety and effectiveness of drugs targeting this target have been preliminarily verified in clinical practice, which can change the patient's immune system to fight cancer by directly stimulating rejection-type processes or blocking inhibitory pathways. Therefore, the development of new, effective and safe CD73 inhibitors provides a new idea for anti-tumor therapy. Summary of the invention

[0007] In view of this, the technical problem to be solved by the present invention is to provide a compound and its application, wherein the compound has excellent CD73 inhibitory activity.

[0008] The present invention provides a compound having a structure as shown in Formula I, or a stereoisomer, a pharmaceutically acceptable salt, a solvate, a cocrystal or a deuterated compound thereof:

[0009] in," ” is a single bond or a double bond; R is selected from any of the following structures: or ; X1 and X2 are each independently N or C; X3 and X4 are each independently N or CH; X5 is selected from N or CH; R1, R2 are independently selected from hydrogen, C 1-6 Alkyl, C 1-6 The halogenated alkyl, C 1-6 alkyl, cycloalkyl, heterocycloalkyl, aryl, heteroaryl, C1-6 Alkyl aryl, C 1-6 Alkyl heteroaryl, C 1-6 Alkylcycloalkyl, C 1-6 Alkyl heterocycloalkyl, C 1-6 Acyl aryl, aryl and C 5-6 Cycloalkyl or C 5-6 wherein the above aryl, heteroaryl, heterocycloalkyl and heterocycloaryl may be substituted by one, two or three R 1b or R1 and R2 together with the nitrogen atom to which they are attached form a group optionally substituted by one, two or three R 1a Substituted heterocycloalkyl, heterocycloaryl, C 3-5 The heterocycloalkyl group is a 3- to 5-membered cycloalkyl group; R 1a , R 1b are independently selected from hydrogen, C 1-6 Alkyl, C 3-6 cycloalkyl or halogen; R3 is independently selected from halogen, thiol, C 3-6 The alkenyl group, C 2-6 Alkynylalkyl or C 2-6 wherein the alkyl group may be further substituted with a hydroxyl group; R4 is independently selected from hydrogen or C 1-6 The alkyl group of 1-6 The alkyl group is selected from methyl, ethyl, n-propyl, isopropyl, n-butyl, isobutyl or tert-butyl, preferably methyl; R 5a , R 5b , R 6a , R 6b are independently selected from hydrogen, halogen, hydroxyl, C 1-3 Alkyl, C 2-3 Alkenyl or C 2-3 The alkynyl group; R7, R8 are independently selected from hydrogen or -(CH2) m -O-(CH2) n R 11 ; m, n are independently selected from 0, 1 or 2; R 11 Independently selected from hydrogen, hydroxy, OCH3, heterocycloalkyl or heteroaryl including 1-4 heteroatoms; R9, R 10 are independently selected from hydrogen or C 1-10 alkyl.

[0010] Optionally, in the structure of formula I, the structural fragment Has any of the following structures: , .

[0011] Optionally, in the structure of Formula I, the structural fragment Has any of the following structures: , , .

[0012] Optionally, R3 is selected from halogen, thiol, or The halogen is F, Cl, Br or I, preferably Cl.

[0013] Optionally, R4 is hydrogen.

[0014] Optionally, the R 5a , R 5b , R 6a , R 6b are independently selected from hydrogen, hydroxy, halogen or C 2-3 The halogen is F, Cl, Br or I, preferably F.

[0015] More preferably, the R 5a , R 5b are independently selected from hydrogen, hydroxyl or F; said R 6a , R 6b are independently selected from hydroxy, ethynyl or propynyl.

[0016] Optionally, R7 and R8 are independently selected from hydrogen, , ,or .

[0017] Optionally, the R9, R 10 independently selected from hydrogen or C 1-6 Alkyl. 1-6 The alkyl group is selected from methyl, ethyl, n-propyl, isopropyl, n-butyl, isobutyl or tert-butyl, preferably methyl or ethyl.

[0018] When R is the following structure:

[0019] The compound has the structure shown in Formula I-a, or its stereoisomer, pharmaceutically acceptable salt, solvate, cocrystal or deuterated substance:

[0020] Formula Ⅰ-a In formula Ⅰ-a, the substituents X1, X2, X3, X4, X5, R1, R2, R3, R4, R 5a , R 5b , R 6a , R 6b , R7, R8, R9, R 10 The range is the same as that of formula Ⅰ.

[0021] Furthermore, the compound has any of the following structures: Formula I-1; Formula I-2; In Formula I-1 and Formula I-2, the substituents R1, R2, R3, R4, R 5a , R 5b , R 6a , R 6b , R7, R8, R9, R 10 , the range of X5 is the same as that of formula Ⅰ.

[0022] Optionally, the structural fragments in the structures of Formula I-1 and Formula I-2 are Has any of the following structures: , , .

[0023] Optionally, the R 5a , R 5b , R 6a , R 6b are independently selected from hydrogen, hydroxy, halogen or C 2-3 The alkynyl group.

[0024] Optionally, R7 and R8 are independently selected from hydrogen, , ,or .

[0025] Optionally, R4, R9, R 10 All are hydrogen.

[0026] Optionally, R3 is selected from halogen, thiol, or .

[0027] In some specific embodiments of the present invention, the compound has the following structure: Formula I-3; In formula Ⅰ-3, the ranges of substituents R1, R2, R3, R7, R8 and X5 are the same as those of formula Ⅰ.

[0028] Optional, structure fragment Has any of the following structures: , , .

[0029] Furthermore, the structural fragment Has any of the following structures: , .

[0030] Optionally, R3 is halogen, or .

[0031] Optionally, R7 and R8 are independently selected from , ,or .

[0032] Optionally, any one of R7 and R8 is selected from .

[0033] When R is the following structure:

[0034] The compound has the structure shown in Formula I-b, or its stereoisomer, pharmaceutically acceptable salt, solvate, cocrystal or deuterated form:

[0035] Formula I-b In formula I-b, the substituents X1, X2, X3, X4, X5, R1, R2, R3, R 5a , R 5b The range is the same as that of formula Ⅰ.

[0036] More preferably, in formula I-b, R1 and R2 are independently selected from hydrogen, cycloalkyl, C 1-6 Alkyl aryl, C 5-6 Cycloalkyl and aryl, aryl and C 5-6 The aryl group may be further substituted with halogen, C 1-3 Alkyl, C 3-5 is substituted by a cycloalkyl group.

[0037] R3 is independently selected from halogen; the halogen is F, Cl, Br or I, preferably Cl.

[0038] R 5a , R 5bIndependently selected from hydrogen, hydroxy or F, preferably hydroxy.

[0039] In formula Ⅰ-b, the structural fragment Has any of the following structures: , .

[0041] In formula Ⅰ-b, the structural fragment Has any of the following structures: , .

[0042] Optionally, the compound has any of the following structures: .

[0043] The present invention also provides a method for preparing the compound represented by the above general formula I, including but not limited to the following steps (the synthesis methods of some compounds are detailed in the Examples): Preparation method 1:

[0044] Preparation method 2:

[0045] Preparation method 3:

[0046] Preparation method 4:

[0047] Furthermore, the present invention also provides a method for preparing a target compound with a single chirality. As shown in the figure below, a pair of isomers II-M1 and II-M2 can be obtained by performing SFC separation on the product obtained by the sodium borohydride reduction reaction, and a subsequent reaction is performed to obtain a target compound with a determined chirality. Similarly, a single chiral isomer of compound IV or V can be successfully prepared according to a similar synthesis method.

[0048]

[0049] The present invention provides a pharmaceutical composition, comprising the above compound, and pharmaceutically acceptable adjuvants and / or carriers.

[0050] The above-mentioned pharmaceutical composition provided by the present invention can be used in combination with an anti-PD-1 antibody. Based on this, further, the above-mentioned pharmaceutical composition also includes an anti-PD-1 antibody.

[0051] The present invention provides the use of the above-mentioned compound or the above-mentioned pharmaceutical composition in the preparation of a drug for treating CD73-mediated diseases.

[0052] Optionally, the CD73-mediated disease is a tumor or an autoimmune disease.

[0053] The present invention also provides a method for treating a CD73-mediated disease, which comprises administering the above-mentioned compound of the present invention or the above-mentioned pharmaceutical composition, wherein the CD73-mediated disease is a tumor or an autoimmune disease. Furthermore, the method further comprises administering another drug, which is an anti-PD-1 antibody.

[0054] In the present invention, the following terms are used: Unless otherwise specified in the present invention, the terms of the present invention have the following meanings: "Halogen" in this article refers to F, Cl, Br, I, or isotopes thereof. "Halo" or "halogen substitution" refers to substitution by one or more selected from F, Cl, Br, I, or isotopes thereof, and the upper limit of the number of halogen substituents is equal to the sum of the number of hydrogens that can be replaced by the substituted group. Unless otherwise specified, the number of halogen substituents is any integer between 1 and the upper limit. When the number of halogen substituents is greater than 1, they can be substituted by the same or different halogens.

[0055] "Deuterated" or "deuterated substance" refers to the situation where hydrogen atoms on groups such as alkyl, cycloalkyl, alkylene, aryl, heteroaryl, thiol, heterocycloalkyl, alkenyl, alkynyl, etc. are replaced by at least one isotope deuterium, and the upper limit of the number of deuterations is equal to the sum of the number of hydrogen atoms that can be replaced. Unless otherwise specified, the number of deuterations is any integer between 1 and the upper limit, preferably 1-20 deuterium atoms are replaced, more preferably 1-10 deuterium atoms are replaced, more preferably 1-6 deuterium atoms are replaced, and further preferably 1-3 deuterium atoms are replaced.

[0056] "Alkyl" refers to a monovalent straight or branched chain saturated aliphatic hydrocarbon group, non-limiting examples of which include methyl, ethyl, n-propyl, isopropyl, n-butyl, sec-butyl, neobutyl, tert-butyl, n-pentyl, isopentyl, neopentyl, n-hexyl and various branched chain isomers thereof.

[0057] "Cycloalkyl" refers to a monovalent saturated, unsaturated, non-aromatic carbocyclic hydrocarbon group, which may be a monocyclic, bicyclic, spirocyclic, bridged, or cyclic group, and non-limiting examples include cyclopropyl, cyclobutyl, cyclopentyl, cyclohexyl, cycloheptyl, cyclopentenyl, etc. The cycloalkyl may be fused to a heteroaryl, heterocycloalkyl, or aryl ring, wherein the ring connected to the parent structure is a cycloalkyl ring. The cycloalkyl may be optionally further substituted by any substituent.

[0058] "Heterocycloalkyl" refers to a substituted or unsubstituted, saturated or unsaturated non-aromatic ring, which, when not specifically defined, contains 1 to 5 heteroatoms selected from N, O, P, Si or S, and further contains 1 to 4 or 1 to 3 heteroatoms selected from N, O, P, Si or S, and may be a monocyclic or polycyclic ring, and the polycyclic ring may be a bridged ring, a cyclic ring or a spirocyclic ring, and, when not specifically defined, is a 3- to 12-membered heterocyclic ring, more preferably a 4- to 12-membered heterocyclic ring, and more preferably a 4- to 10-membered heterocyclic ring. The N, S, and P selectively substituted in the heterocyclic ring are oxidized to various oxidation states. The heterocycloalkyl may be optionally further substituted by any substituent.

[0059] "Heterocycle" or "heterocyclic group" refers to a substituted or unsubstituted, saturated or unsaturated aromatic or non-aromatic ring, which, when not specifically defined, contains 1 to 5 heteroatoms selected from N, O or S, further contains 1 to 4 heteroatoms selected from N, O or S, and further contains 1 to 3 heteroatoms selected from N, O or S, including monocyclic heterocycles or polycyclic heterocycles, polycyclic heterocycles include bicyclic bridged heterocycles, bicyclic heterocycles and bicyclic spiro heterocycles, etc., and, when not specifically defined, is a 3- to 12-membered heterocycle, more preferably a 4- to 12-membered heterocycle, more preferably a 4- to 10-membered heterocycle, and further preferably a 4- to 7-membered heterocycle. Its definition includes heterocycloalkyl and heteroaryl. N and S in the heterocyclic ring can be oxidized to various oxidation states.

[0060] "Aryl" refers to a substituted or unsubstituted 5 to 15-membered aromatic carbon ring, including monocyclic aromatic groups and condensed aromatic groups. Preferably, the aromatic ring is 5 to 10 members, and more preferably, the aromatic ring is 5 to 8 members, and non-limiting examples thereof include phenyl, naphthyl, anthracenyl, and phenanthrenyl. The aryl ring may be fused to a heteroaryl, heterocycloalkyl, or cycloalkyl ring, wherein the ring connected to the parent structure is an aryl ring. The aryl may optionally be further substituted by any substituent.

[0061] "Heteroaryl" refers to a substituted or unsubstituted 5 to 15-membered aromatic ring, and contains 1 to 5 heteroatoms selected from N, O, P, Si or S and their heteroatoms and oxidized forms, preferably a 5 to 10-membered heteroaromatic ring, and further preferably 5 to 8 members. Non-limiting examples of heteroaryl include, but are not limited to, furanyl, oxazolyl, thienyl, N-alkylpyrrolyl, pyrazinyl, pyridazinyl, piperidinyl, morpholine, thiomorpholine, 1.3-dithiane, benzimidazole. The heteroaryl ring can be fused to an aryl, heterocycloalkyl or cycloalkyl ring, wherein the ring connected to the parent structure is a heteroaryl ring. The heteroaryl can be optionally further substituted by any substituent.

[0062] "Alkynyl" refers to a linear or branched monovalent unsaturated hydrocarbon group containing one or more carbon-carbon triple bonds. Unless otherwise specified, the alkynyl group contains 2-6 carbon atoms, preferably 2-4 carbon atoms. Non-limiting examples include ethynyl, propynyl, propargyl, etc.

[0063] "Alkenyl" refers to a linear or branched monovalent unsaturated hydrocarbon group containing one or more carbon-carbon double bonds. Unless otherwise specified, the alkenyl group contains 2-6 carbon atoms, preferably 2-4 carbon atoms. Non-limiting examples are ethenyl, propenyl, allyl, 2-butenyl, 1-butenyl, etc.

[0064] "Substitution" or "substituent" unless otherwise specified refers to any substitution at a position permitted by chemical theory, and the number of substituents complies with the chemical bond rules. Exemplary substituents include, but are not limited to, cyano, halogen, nitro, amino, hydroxyl, C 1-6 Alkyl, C 2-6 Alkenyl, C 2-6 Alkynyl, C 3-8 Heteroalkyl, C 5-12 Aryl, 5-12 membered heteroaryl, C 1-6 Alkoxy, C 5-12 Aryloxy, thiol, C 1-6 Alkylthio, C 1-6 Alkylthiocarbonyl, C 1-6 Alkylcarbamoyl, N-carbamoyl, silyl, sulfinyl, sulfonyl, sulfoxide, halogenated C 1-6 Alkyl, phosphonic acid, -CO2(C1-6 Alkyl), -OC(=O) (C 1-6 Alkyl), -OCO2(C 1-6 alkyl), -C(=O)NH2, C(=O)N(C 1-6 alkyl)2、OC(=O)NH(C 1-6 Alkyl), -NHC(=O)(C 1-6 Alkyl), -N(C 1-6 Alkyl)C(=O)(C 1-6 Alkyl), -NHCO2(C 1-6 alkyl), NHC(=O)N(C 1-6 Alkyl)2, -HC(=O)NH(C 1-6 Alkyl), -SO2N(C 1-6 Alkyl), - SO2NH(C 1-6 Alkyl), -SO2NH2, SO2C 1-6 Alkyl, etc.

[0065] "Optional" or "optionally" means that the event or circumstance described later may but need not occur, and the description includes occasions where the event or circumstance occurs or does not occur. For example, "alkyl optionally substituted with F" means that alkyl may but need not be substituted with F, and the description includes situations where alkyl is substituted with F and situations where alkyl is not substituted with F.

[0066] "Pharmaceutically acceptable salt" refers to a salt of the compound of the present invention which retains the biological effectiveness and properties of the free acid or free base, and the free acid is obtained by reacting with a non-toxic inorganic base or organic base, or the free base is obtained by reacting with a non-toxic inorganic acid or organic acid.

[0067] A "pharmaceutical composition" refers to a mixture of one or more compounds described herein, or stereoisomers, solvates, pharmaceutically acceptable salts or cocrystals thereof, with other ingredients, wherein the other ingredients include physiologically / pharmaceutically acceptable carriers and / or excipients.

[0068] "Carrier" refers to a system that does not cause significant irritation to the organism and does not eliminate the biological activity and properties of the administered compound, and can change the way the drug enters the human body and its distribution in the body, control the release rate of the drug and deliver the drug to the targeted organ. Non-limiting examples include microcapsules and microspheres, nanoparticles, liposomes, etc.

[0069] "Excipient" refers to: it is not a therapeutic agent itself, but is used as a diluent, adjuvant, binder and / or vehicle, and is used to add to a pharmaceutical composition to improve its handling or storage properties or to allow or facilitate the formation of a compound or pharmaceutical composition into a unit dosage form for administration. As known to those skilled in the art, pharmaceutical excipients can provide various functions and can be described as wetting agents, buffers, suspending agents, lubricants, emulsifiers, disintegrants, absorbents, preservatives, surfactants, colorants, flavoring agents and sweeteners.

[0070] "Stereoisomers" refer to isomers produced by different spatial arrangements of atoms in molecules, including cis-trans isomers, (dia)enantiomers and conformational isomers.

[0071] "Solvate" refers to a substance formed by a stoichiometric or non-stoichiometric amount of a solvent that is bound to the compound or salt of the present invention by non-covalent forces between molecules. When the solvent is water, it is a hydrate.

[0072] "Co-crystal" refers to a crystal formed by the active pharmaceutical ingredient (API) and the co-crystal former (CCF) under the action of hydrogen bonds or other non-covalent bonds, in which the pure state of API and CCF are solid at room temperature and there is a fixed stoichiometric ratio between the components. Co-crystal is a multi-component crystal, including binary eutectics formed between two neutral solids and multi-component eutectics formed between neutral solids and salts or solvates.

[0073] Compared with the prior art, the present invention provides a compound having a structure shown in Formula I, or a stereoisomer, a pharmaceutically acceptable salt, a solvate, a cocrystal or a deuterated product thereof. The compound has excellent CD73 enzyme inhibitory activity and tumor cell surface CD73 inhibitory activity, can be developed as a new anti-CD73 drug, and has broad application prospects. BRIEF DESCRIPTION OF THE DRAWINGS

[0074] Figure 1 :Effect of Test Example 4 on Tumor Volume in Lymphoma Mouse Tumor Model Figure 2 :Effect of Test Example 5 on Tumor Volume in Lymphoma Mouse Tumor Model DETAILED DESCRIPTION

[0075] In order to further illustrate the present invention, the compounds provided by the present invention, their preparation methods and applications are described in detail below in conjunction with examples.

[0076] If no specific conditions are specified in the examples, the experimental methods under conventional conditions are used. The examples are provided to better illustrate the content of the present invention, but it should not be understood that the content of the present invention is limited to the examples. Non-essential improvements and adjustments made by those skilled in the art to the implementation scheme according to the above-mentioned content of the invention still fall within the scope of protection of the present invention.

[0077] The full names corresponding to the abbreviations of the present invention are shown in Table 1.

[0078] Table 1 List of abbreviations abbreviation Full name abbreviation Full name MS Mass spectrometry NMR Nuclear Magnetic Resonance LCMS Liquid chromatography-mass spectrometry DMF N,N-Dimethylformamide DCM Dichloromethane DMSO Dimethyl sulfoxide <![CDATA[BF3.Et2O]]> Boron trifluoride etherate TESH Triethylsilane THF Tetrahydrofuran PPTS 4-Methylbenzenesulfonate pyridine DMP Dess-Martin Oxidant <![CDATA[CS2]]> Carbon disulfide <![CDATA[K2CO3]]> Potassium carbonate <![CDATA[NaHCO3]]> Sodium bicarbonate m-CPBA m-Chloroperbenzoic acid <![CDATA[NH2NH2.H2O]]> Hydrazine hydrate <![CDATA[Et3N]]> Triethylamine EtOH Ethanol NaH Sodium hydride <![CDATA[nBu3SnH]]> Tributyltin Hydrogen <![CDATA[CH3I]]> Methyl iodide AIBN Azobisisobutyronitrile KOH Potassium hydroxide TEA Triethylamine TFA Trifluoroacetic acid AcOH acetic acid LiHMDS Lithium Bis(trimethylsilyl)amide TBDPSCl Tert-Butyldiphenylsilyl chloride DMAP 4-Dimethylaminopyridine TxD p-Toluenesulfonic acid <![CDATA[Rh2(OAc)4]]> Rhodium Acetate Dimer Toluene Toluene MOMCl Chloromethyl methyl ether NaHMDS Sodium bis(trimethylsilyl)amide TBAI Tetrabutylammonium iodide <![CDATA[NaBH4]]> Sodium borohydride <![CDATA[CaCl2]]> Calcium chloride TMSBr Trimethylsilane <![CDATA[Cs2CO3]]> Cesium carbonate HCl hydrochloric acid <![CDATA[LiBH4]]> Lithium borohydride Imidazole Imidazole DIAD Diisopropyl azodicarboxylate <![CDATA[PPh3]]> Triphenylphosphine <![CDATA[KOsO4]]> Potassium Osate TBAF Tetrabutylammonium fluoride DIEA N,N-Diisopropylethylamine nBuLi n-Butyl lithium <![CDATA[CH3CN]]> Acetonitrile TBSCl tert-Butyldimethylsilyl chloride <![CDATA[Ac2O]]> Acetic anhydride

[0079] Example 1 (2-((2R,3S,4S,5R)-5-(6-chloro-4-(3,3-difluoroazetidin-1-yl)-1H-pyrazolo[3,4-d]pyrimidin-1-yl)-3,4-dihydroxytetrahydro-2H-pyran-2-yl)methoxy)-1-hydroxy-3-methoxypropane-2-yl)phosphonic acid

[0080] The preparation process is as follows:

[0081] Step 1: Synthesis of compound 1B (2R, 3S, 4R)-2-(acetoxymethyl)-3.4-dihydro-2H-pyran-3.4-diacetic acid diester 1A (20.0 g, 73.5 mmol) was dissolved in dichloromethane (200 mL), and triethylsilane (14.2 mL, 88.1 mmol) and boron trifluoride etherate (10.2 mL, 80.8 mmol) were added under ice bath, and the resulting solution was stirred at room temperature for 2 hours. Then 200 ml of saturated sodium bicarbonate aqueous solution was added to quench the reaction. The organic phases were combined and extracted twice with dichloromethane (100 mL), washed with saturated brine, dried over anhydrous sodium sulfate, filtered and concentrated under vacuum to obtain a light yellow oil 1B (15.7 g, yield: 100%). 1 H NMR (400 MHz, CDCl3) δ 5.93 (d, J = 10.4 Hz, 1H), 5.75 (d, J = 10.4Hz, 1H), 5.24 (d, J = 8.1 Hz, 1H), 4.25 – 4.12 (m, 4H), 3.77 – 3.65 (m, 1H), 2.08 (d, J = 8.6 Hz, 6H).

[0082] Step 2: Synthesis of Compound 1C Potassium carbonate (2.14 g, 15.5 mmol) was added to a solution of (2R, 3S)-3-acetoxy-3,6-dihydro-2H-pyran-2-yl) methyl acetate 1B (16.6 g, 77.5 mmol) in methanol (200 mL). The mixture was stirred at room temperature for 30 minutes. After the reaction was completed, the reaction solution was concentrated under vacuum and diluted with dichloromethane (150 mL). The solution was then filtered and the filter cake was washed with cold dichloromethane. The obtained filtrate was concentrated under reduced pressure, and the residue was purified by silica gel column chromatography (eluent: methanol / dichloromethane = 1 / 10, V / V) to obtain compound 1C (9.2 g, colorless oil, yield: 91.2%).

[0083] Step 3: Synthesis of Compound 1D Under an ice bath, m-CPBA (15.7 g, 77.8 mmol, 85%) was added in batches to a solution of (2R, 3S)-2-(hydroxymethyl)-3,6-dihydro-2H-pyran-3-ol 1C (9.2 g, 70.7 mmol) in dichloromethane (150 mL). The reaction solution was stirred at room temperature for 2 days. After the reaction was completed, the reaction solution was filtered and the filter cake was washed with cold dichloromethane. The filtrate was combined and concentrated under reduced pressure. The obtained oil was purified by silica gel column chromatography (eluent: methanol / dichloromethane=1 / 10, V / V) to obtain compound 1D (6.4 g, white solid, yield: 62.0%).

[0084] Step 4: Synthesis of Compound 1E 2,2-Dimethoxypropane (10.9 mL, 87.6 mmol) and pyridinium p-toluenesulfonate (11.0 g, 43.8 mmol) were added to (1S, 4R, 5R, 6R)-4-(hydroxymethyl)-3.7-dioxabicyclo[4.1.0]heptane-5-ol 1D (6.4 g, 43.8 mmol) in anhydrous tetrahydrofuran (100 mL). The reaction solution was stirred at room temperature for one day. After the reaction was completed, the reaction solution was diluted with ethyl acetate and washed with a saturated saline solution. The obtained organic phase was dried over anhydrous sodium sulfate, filtered, and concentrated. The obtained oil was purified by silica gel column chromatography (eluent: ethyl acetate / petroleum ether = 1 / 3, V / V) to obtain compound 1E (4.4 g, colorless oil, yield: 54.0%). 1 HNMR (400MHz, CDCl3) δ 4.07-3.93(m, 3H), 3.78(dd, J=10.3, 5.1 Hz, 1H), 3.65-3.45(m, 2H), 3.42-3.29(m, 2H), 1.49(s, 3H), 1.41(s,3H).

[0085] Step 5: Synthesis of Compound 1F Hydrazine hydrate (2.84 g, 70.9 mmol, 80% aqueous solution) was added to a solution of (1aS, 3aR, 7aR, 7bS)-6,6-dimethylhexahydrobenzo[2,3:4,5]pyrano[3,2-d][1,3]dioxin 1E (4.4 g, 23.6 mmol) in EtOH (50 mL). The reaction solution was stirred at 60 °C overnight. After the reaction was completed, the solution was concentrated under vacuum to obtain compound 1F (5.2 g, colorless oil). The product was used directly in the next step.

[0086] Step 6: Synthesis of Compound 1G At -60°C, slowly add a solution of (4aR,7R,8S,8aS)-7-hydrazino-2,2-dimethylhexahydropyrano[3,2-d][1,3]dioxin-8-ol 1F (5.2 g, 23.6 mmol) and DIPEA (7.8 mL, 47.3 mmol) in tetrahydrofuran (20 mL) to a solution of 2,4,6-trichloropyrimidine-5-carboxaldehyde (5 g, 23.65 mmol) in anhydrous tetrahydrofuran (50 mL). Slowly warm the reaction to room temperature and stir at room temperature for one hour. After the reaction is complete, the reaction solution is diluted with ethyl acetate and washed with a saturated saline solution. The organic phase is dried over anhydrous sodium sulfate, filtered, and concentrated under reduced pressure to obtain a crude product. The crude product was purified by silica gel column chromatography (eluent: ethyl acetate / petroleum ether = 1 / 5, V / V) to obtain compound 1G (4.7 g, colorless oil, yield: 52.6%). LC-MS m / z (ESI): 375 (M+H + ); 1 H NMR (400 MHz, CDCl3) δ 8.20 (s, 1H),5.05 (s, 1H), 4.43 (d, J = 9.0 Hz, 1H), 4.34 – 4.28 (m, 2H), 4.21 – 4.12 (m,1H), 3.96 – 3.79 (m, 3H), 2.82 (s, 1H), 1.52 (s, 3H), 1.42 (s, 3H).

[0087] Step 7: Synthesis of compound 1H (4aR, 7R, 8S, 8aS)-7-(4,6-dichloro-1H-pyrazolo[3,4-d]pyrimidin-1-yl)-2,2-dimethylhexahydropyrano[3,2-d][1,3]dioxin-8-ol 1G (3.0 g, 8.00 mmol) was dissolved in a mixed solution of ethyl acetate (30 mL) and ethanol (10 mL). 3,3-difluoroazetidine (0.82 g, 8.80 mmol) and triethylamine (3.3 mL, 24.0 mmol) were added in sequence under ice bath. The reaction solution was stirred at room temperature for 5 hours. After the reaction was completed, the mixture was diluted with ethyl acetate (200 mL) and washed with saturated brine. The organic phase was concentrated under reduced pressure to obtain a crude product, which was purified by silica gel column chromatography (eluent: ethyl acetate / petroleum ether = 1 / 3, V / V) to obtain compound 1H (3 g, yellow oil, yield 86.9%). LC / MS (ESI) m / z: 432.2 [M+H] + .

[0088] Step 8: Synthesis of Compound 1I (4aR, 7R, 8S, 8aS)-7-(6-chloro-4-(3,3-difluoroazetidin-1-yl)-1H-pyrazolo[3,4-d ]pyrimidin-1-yl)-2,2-dimethylhexahydropyrano[3,2-d ][1,3]dioxin-8-ol 1H (3 g, 6.95 mmol) was added to a mixed solution of acetic acid (10 mL) and water (10 mL). The reaction was stirred at 60°C for 1 h. After the reaction was completed, it was concentrated under reduced pressure, diluted with ethyl acetate (200 mL), and washed with saturated sodium bicarbonate (50 mL) and saturated sodium chloride in sequence. The organic phase was concentrated under reduced pressure to obtain a crude product, which was purified by silica gel column chromatography (eluent: methanol / dichloromethane = 1 / 10, V / V) to obtain the title compound 1I (2.6 g, white solid, yield: 95.5%). LC / MS (ESI) m / z: 392.2 [M+H] + .

[0089] Step 9: Synthesis of Compound 1J To a solution of (2R,3S,4S,5R)-5-(6-chloro-4-(3,3-difluoroazetidin-1-yl)-1H-pyrazolo[3,4-d]pyrimidin-1-yl)-2-(hydroxymethyl)tetrahydro-2H-pyran-3,4-diol 1I (2.6 g, 6.64 mmol) in acetone (30 ml) were added p-toluenesulfonic acid (3.42 g, 19.9 mmol) and 2,2-dimethoxypropane (4.15 g, 39.8 mmol). The reaction solution was stirred at room temperature overnight. After the reaction was completed, the reaction solution was diluted with ethyl acetate (200 mL) and washed twice with saturated sodium bicarbonate (100 mL) and once with saturated brine. The organic phase was dried over anhydrous sodium sulfate and concentrated under reduced pressure to obtain a crude product. The crude product was purified by silica gel column chromatography (eluent: ethyl acetate / petroleum ether = 1 / 3, V / V) to obtain the title compound 1J (1.9 g, yellow oil, yield: 66.3%). LC / MS (ESI) m / z: 432 [M+H] + .

[0090] Step 10: Synthesis of compound 1K To a solution of ((3aS, 4R, 7R, 7aS)-7-(6-chloro-4-(3,3-difluoroazetidin-1-yl)-1H-pyrazolo[3,4-d]pyrimidin-1-yl)-2,2-dimethyltetrahydro-4H-[1,3]dioxo[4,5-c]pyran-4-yl)methanol 1J (1.9 g, 4.40 mmol) in toluene (20 mL) were added 1-(diethoxyphosphoryl)-2-ethoxy-2-oxoethane-1-diazide (3.32 g, 13.2 mmol) and rhodium dimer acetate (0.10 g, 0.22 mmol) in sequence. The mixture was replaced with nitrogen three times and then stirred at 100°C for 3 h. After the reaction was completed, ethyl acetate (200 mL) was added to the reaction solution and washed once with saturated sodium bicarbonate and saturated brine. The organic phase was dried over anhydrous sodium sulfate and concentrated under reduced pressure to obtain a crude product. The crude product was purified by silica gel column chromatography (eluent: ethyl acetate / petroleum ether = 2 / 1, V / V) to obtain the title compound 1K (1.9 g, yellow oil, yield: 66.0%). LC / MS (ESI) m / z: 654 [M+H] + .

[0091] Step 11: Synthesis of Compound 1L Under N2 atmosphere, a solution of ethyl 2-((3aS, 4R, 7R, 7aS)-7-(6-chloro-4-(3,3-difluoroazetidin-1-yl)-1H-pyrazolo[3,4-d]pyrimidin-1-yl)-2,2-dimethyltetrahydro-4H-[1,3]dioxo[4,5-c]pyran-4-yl)methoxy)-2-(diethoxyphosphoryl)acetate 1K (1.4 g, 2.14 mmol) in THF (20 mL) was cooled to -15°C, and then NaHMDS (1.1 mL, 2.14 mmol, 2M in tetrahydrofuran solution) was slowly added dropwise at this temperature. The reaction solution was stirred at -15°C for 30 minutes. Then, TBAI (0.39 g, 1.07 mmol) was added to the reaction solution, and chloromethyl methyl ether (0.34 g, 4.28 mmol) was added dropwise. The reaction solution was stirred at -15 °C for 3 hours. After the reaction was completed, the reaction was quenched with saturated aqueous ammonium chloride solution and extracted three times with ethyl acetate (50 mL). The combined organic layer was washed once with saturated brine, dried over anhydrous sodium sulfate, filtered and concentrated under reduced pressure to obtain a crude product. The crude product was purified by silica gel column chromatography (eluent: ethyl acetate / petroleum ether = 2 / 1, V / V) to obtain the title compound 1L (0.8 g, yellow oil, yield: 53.5%). LC / MS (ESI) m / z: 698 [M+H] + .

[0092] Step 12: Synthesis of Compound 1M Under ice bath, calcium chloride (0.38 g, 3.44 mmol) and sodium borohydride (0.13 g, 34.4 mmol) were added to a solution of 1L (0.8 g, 1.15 mmol) of ethyl 2-((3aS, 4R, 7R, 7aS)-7-(6-chloro-4-(3,3-difluoroazetidin-1-yl)-1H-pyrazolo[3,4-d ]pyrimidin-1-yl)-2,2-dimethyltetrahydro-4H-[1,3]dioxo[4,5-c ]pyran-4-yl)methoxy)-2-(diethoxyphosphoryl)-3-methoxypropanoate in ethanol (8 mL) in sequence. The reaction solution was stirred at room temperature overnight. After the reaction was completed, the reaction was quenched with saturated aqueous ammonium chloride solution and extracted with ethyl acetate (100 mL) three times. The combined organic layer was washed once with saturated brine, dried over anhydrous sodium sulfate, filtered and concentrated under reduced pressure to obtain a crude product. The crude product was purified by silica gel column chromatography (eluent: methanol / dichloromethane = 1 / 10, V / V) to obtain the title compound 1M (0.5 g, yellow oil, yield: 66.5%). LC / MS (ESI) m / z: 656 [M+H] + .

[0093] Step 13: Synthesis of Compound 1 Under ice bath, triethylamine (0.30 mL, 2.13 mmol) and trimethylsilyl bromide (233 mg, 1.52 mmol) were added to a solution of diethyl (2-(((3aS, 4R, 7R, 7aS)-7-(6-chloro-4-(3,3-difluoroazetidin-1-yl)-1H-pyrazolo[3,4-d]pyrimidin-1-yl)-2,2-dimethyltetrahydro-4H-[1,3]dioxo[4,5-c ]pyran-4-yl)methoxy)-1-hydroxy-3-methoxypropane-2-yl)phosphonate 1M (100 mg, 0.152 mmol) in acetonitrile (5 mL) in sequence. The reaction mixture was stirred at room temperature overnight. After the phospholipids were completely hydrolyzed, the reaction mixture was concentrated under reduced pressure. Then trifluoroacetic acid / water (3 / 1, 2 mL) was added to the obtained crude product. The reaction solution was stirred at room temperature for one hour. After the reaction was completed, the reaction solution was concentrated under reduced pressure to obtain a residue, which was purified by reverse phase preparative chromatography (eluent: acetonitrile / 0.1% formic acid aqueous solution = 25% ~ 90%, V / V) to obtain compound 1 (53.3 mg, white solid, yield: 62.5%). LC / MS (ESI) m / z: 560.1 [M+H] + . 1 H NMR (400 MHz, DMSO- d 6) δ 8.23 ​​(d, J = 2.5 Hz, 1H), 5.17 – 4.75 (m, 6H), 4.38 – 4.32 (s, 1H), 4.14 – 3.72 (m, 11H), 3.37 (s, 6H). 31 P NMR (400 MHz, DMSO- d 6)δ 19.11 (s).

[0094] Example 2 2-(2R,3S,4S,5R)-5-(6-chloro-4-((S)-1-phenylethylamino)-1H-pyrazolo[3,4-d]pyrimidin-1-yl)-3,4-dihydroxytetrahydro-2H-pyran-2-yl)methoxy)-1-hydroxy-3-methoxypropane-2-yl)phosphonic acid

[0095] Compound 2 can be synthesized by referring to the synthesis steps of Example 1 using commercially available raw materials. LC / MS (ESI) [M+H] + m / z:588.2. 1 H NMR (400 MHz, CD3OD) δ 8.12 – 8.06 (s, 1H), 7.42 (d,J = 7.6 Hz, 2H),7.32 (t, J = 7.6 Hz, 2H), 7.25 – 7.20 (m, 1H), 5.56 – 5.48(m, 1H), 4.85 – 4.81(m, 1H), 4.45 – 4.35 (m, 1H), 4.25 – 4.00 (m, 5H), 4.00 – 3.86 (m, 3H), 3.86– 3.74 (m, 2H), 3.38 (s, 3H), 1.60 (d, J = 7.0 Hz, 3H).

[0096] Example 3 2-(2R,3S,4S,5R)-5-(6-chloro-4-(2,3-dihydro-1H-inden-2-yl)amino)-1H-pyrazolo[3,4-d]pyrimidin-1-yl)-3,4-dihydroxytetrahydro-2H-pyran-2-yl)methoxy)-1-hydroxy-3-methoxypropane-2-yl)phosphonic acid

[0097] Compound 3 can be synthesized by referring to the synthesis steps of Example 1 using commercially available raw materials. LC / MS (ESI) m / z: 600.2[M+H] + . 1 H NMR (400 MHz, CD3OD) δ 8.06 (s, 1H), 7.27 – 7.21 (m, 2H), 7.18 – 7.13 (m, 2H), 5.06 – 4.96 (m, 1H), 4.46 – 4.40 (m, 1H), 4.26 – 3.73 (m, 11H),3.46 – 3.39 (m, 2H), 3.37 (s, 3H), 2.99 (dd, J = 16.0, 5.8 Hz, 2H).

[0098] Example 4 2-((2R,3S,4S,5R)-5-(6-chloro-4-hexahydrocyclopentylpyrrol-2(1H)yl)-1H-pyrazolo[3,4-d]pyrimidin-1-yl)-3,4-dihydroxytetrahydro-2H-pyran-2-yl)methoxy)-1-hydroxy-3-methoxypropan-2-yl)phosphonic acid

[0099] Compound 4 can be synthesized by referring to the synthesis steps of Example 1 using commercially available raw materials. LC / MS (ESI) m / z: 578.2[M+H] + . 1 H NMR (400 MHz, CD3OD) δ 8.09 (s, 1H), 4.90 – 4.87 (m, 1H), 4.46 – 4.38 (m, 1H), 4.25 – 4.02 (m, 6H), 3.98 – 3.75 (m, 6H), 3.68 – 3.55 (m, 2H), 3.38 (s, 3H), 3.00 – 2.74 (m, 2H), 2.05 – 1.80 (m, 3H), 1.77 – 1.52 (m, 3H).

[0100] Example 5 (2-(2R,3S,4S,5R)-5-(6-chloro-4-((R)-1,2,3,4-tetrahydronaphthalen-1-amino)-1H-pyrazolo[3,4-d]pyrimidin-1-yl)-3,4-dihydroxytetrahydro-2H-pyran-2-yl)methoxy)-1-hydroxy-3-methoxypropane-2-yl)phosphonic acid

[0101] Compound 5 can be synthesized by referring to the synthesis steps of Example 1 using commercially available raw materials. LC / MS (ESI) m / z: 614.2[M+H]+. 1H NMR (400 MHz, DMSO- d 6) δ 9.06 (d, J = 8.4 Hz, 1H), 8.19 (d, J = 1.8 Hz,1H), 7.24 – 7.11 (m, 4H), 5.53 – 5.42 (m, 1H), 4.96 (s, 1H), 4.83 – 4.71 (m,1H), 4.28 – 4.20 (m, 1H), 3.95 – 3.61 (m, 10H), 3.28 (s, 3H), 2.90 – 2.73 (m, 2H), 2.09 – 1.78 (m, 4H).

[0102] Example 6 (2-(2R,3S,4S,5R)-5-(4-(3-azabicyclo[3.1.0]hexan-3-yl)-6-chloro-1H-pyrazolo[3,4-d]pyrimidin-1-yl)-3,4-dihydroxytetrahydro-2H-pyran-2-yl)methoxy)-1-hydroxy-3-methoxypropane-2-yl)phosphonic acid

[0103] Compound 6 can be synthesized by referring to the synthesis steps of Example 1 using commercially available raw materials. LC / MS (ESI) m / z: 550.2[M+H] + . 1 H NMR (400 MHz, DMSO- d 6) δ 8.19 (d, J = 3.0 Hz, 1H), 4.81 – 4.74 (m,1H), 4.27 – 4.21 (m, 1H), 4.01 – 3.85 (m, 7H), 3.83 – 3.72 (m, 2H), 3.72 –3.59 (m, 5H), 1.87 – 1.80 (m, 1H), 1.76 – 1.69 (m, 1H), 0.87 – 0.79 (m, 1H), 0.24 – 0.16 (q, J = 4.2 Hz, 1H).

[0104] Example 7 (2-(2R,3S,4S,5R)-5-(6-chloro-4-((S)-tetrahydrofuran-3-amino)-1H-pyrazolo[3,4-d]pyrimidin-1-yl)-3,4-dihydroxytetrahydro-2H-pyran-2-yl)methoxy)-1-hydroxy-3-methoxypropane-2-yl)phosphonic acid

[0105] Compound 7 can be synthesized by referring to the synthesis steps of Example 1 using commercially available raw materials. LC / MS (ESI) m / z: 554.1[M+H] + . 1 H NMR (400 MHz, CD3OD) δ 8.09 (s, 1H), 4.85 – 4.75 (m, 1H), 4.46 –4.40 (m, 1H), 4.24 – 3.71 (m, 15H), 3.38 (d, J = 0.9 Hz, 3H), 2.41 – 2.30 (m,1H), 2.06 – 1.96 (m, 1H).

[0106] Example 8 2-(2R,3S,4S,5R)-5-(6-chloro-4-cyclopropylmethylamino)-1H-pyrazolo[3,4-d]pyrimidin-1-yl)-3,4-dihydroxytetrahydro-2H-pyran-2-yl)methoxy)-1-hydroxy-3-methoxypropan-2-yl)phosphonic acid

[0107] Compound 8 can be synthesized by referring to the synthesis steps of Example 1 using commercially available raw materials. LC / MS (ESI) m / z: 538.2[M+H] + . 1 H NMR (400 MHz, DMSO- d 6) δ 8.84 (t, J = 5.0 Hz, 1H), 8.18 (d, J = 1.8 Hz,1H), 4.75 – 4.75 (m, 1H), 4.27 – 4.20 (m, 2H), 4.03 – 3.61 (m, 9H), 3.36 –3.30 (m, 2H), 3.28 (s, 3H), 1.14 – 1.03 (m, 1H), 0.54 – 0.46 (m, 2H), 0.32 –0.25 (m, 2H).

[0108] Example 9 2-(2R,3S,4S,5R)-5-(6-chloro-4-pyrazol-1-yl)-1H-pyrazolo[3,4-d]pyrimidin-1-yl)-3,4-dihydroxytetrahydro-2H-pyran-2-yl)methoxy)-1-hydroxy-3-methoxypropane-2-yl)phosphonic acid

[0109] Compound 9 can be synthesized by referring to the synthesis steps of Example 1 using commercially available raw materials. LC / MS (ESI) m / z: 535.3[M+H] + . 1 H NMR (400 MHz, DMSO- d 6) δ 8.80 (d, J = 2.8 Hz, 1H), 8.65 (d, J= 2.8 Hz,1H), 8.17 (s 1H), 6.80 – 6.76 (m, 1H), 4.99 – 4.95 (m, 1H), 4.33 – 4.31 (m,1H), 4.02 – 3.95 (m, 3H), 3.92 – 3.85 (m, 3H), 3.74 – 3.64 (m, 4H), 3.29 (s,3H).

[0110] Example 10 (2-(2R,3S,4S,5R)-5-(6-chloro-4-cyclopentylamino)-1H-pyrazolo[3,4-d]pyrimidin-1-yl)-3,4-dihydroxytetrahydro-2H-pyran-2-yl)methoxy)-1-hydroxy-3-methoxypropan-2-yl)phosphonic acid

[0111] Compound 10 can be synthesized by referring to the synthesis steps of Example 1 using commercially available raw materials. 1 H NMR (400 MHz, CD3OD)δ 8.08 (s, 1H), 4.95 – 4.85 (m, 1H), 4.59 – 4.50 (m, 1H), 4.45 – 4.39 (m,1H), 4.24 – 4.03 (m, 5H), 3.98 – 3.77 (m, 5H), 3.38 (s, 3H), 2.15 – 2.03 (m, 2H), 1.85 – 1.54 (m, 6H).

[0112] Embodiment 11 (2-(2R,3S,4S,5R)-5-(6-chloro-4-(2,3-dihydro-1H-inden-1-yl)amino)-1H-pyrazolo[3,4-d]pyrimidin-1-yl)-3,4-dihydroxytetrahydro-2H-pyran-2-yl)methoxy)-1-hydroxy-3-methoxypropane-2-yl)phosphonic acid

[0113] Compound 11 can be synthesized by referring to the synthesis steps of Example 1 using commercially available raw materials. 1 H NMR (400 MHz, CD3OD)δ 8.05 (s, 1H), 7.31 – 7.26 (m, 2H), 7.26-7.11 (m, 2H), 5.92 (t, J= 7.3 Hz,1H), 4.92-4.84 (m, 1H), 4.51-4.44 (m, 1H), 4.26 – 4.12 (m, 3H), 4.12 – 4.02(m, 2H), 3.99 – 3.86 (m, 3H), 3.80-3.71 (m, 2H), 3.38 (s, 3H), 3.15 - 3.09 (m, 1H), 2.94 - 2.80 (m, 1H), 2.70 - 2.58 (m, 1H), 2.09 - 1.95 (m, 1H).

[0114] Example 12 2-(2R,3S,4S,5R)-5-(6-chloro-4-(2,3-dihydro-1H-inden-4-ylamino)-1H-pyrazolo[3,4-d]pyrimidin-1-yl)-3,4-dihydroxytetrahydro-2H-pyran-2-yl)methoxy)-1-hydroxy-3-methoxypropane-2-yl)phosphonic acid

[0115] Compound 12 can be synthesized by referring to the synthesis steps of Example 1 using commercially available raw materials. 1 H NMR (400 MHz, CD3OD)δ 7.35-7.15 (m, 3H), 4.92-4.82 (m, 1H), 4.45-4.37 (m, 1H), 4.22 – 4.00 (m,5H), 3.98 – 3.74 (m, 5H), 3.36 (s, 3H), 3.00 (t, J = 7.2 Hz, 2H), 2.81 (t, J =7.2 Hz, 2H), 2.11-2.01 (m, 2H).

[0116] Example 13 (2-(2R,3S,4S,5R)-5-(6-chloro-4-((2,3-dihydro-1H-inden-5-yl)amino)-1H-pyrazolo[3,4-d]pyrimidin-1-yl)-3,4-dihydroxytetrahydro-2H-pyran-2-yl)methoxy)-1-hydroxy-3-methoxypropane-2-yl)phosphonic acid

[0117] Compound 13 can be synthesized by referring to the synthesis steps of Example 1 using commercially available raw materials. 1H NMR (400 MHz, CD3OD)δ 7.62 – 7.13 (m, 3H), 4.92-4.80 (m, 1H), 4.49-4.40 (m, 1H), 4.13 (m, 5H), 3.97 – 3.76 (m, 5H), 3.38 (s, 3H), 2.97-2.91 (m, 4H), 2.16-2.09 (m, 2H).

[0118] Embodiment 14 (2-(2R,3S,4S,5R)-5-(4-Benzamido-6-chloropyrazolo[3,4-d]pyrimidin-1-yl)-3,4-dihydroxytetrahydro-2H-pyran-2-yl)methoxy)-1-hydroxy-3-methoxypropan-2-yl)phosphonic acid

[0119] Compound 14 can be synthesized by referring to the synthesis steps of Example 1 using commercially available raw materials. 1 H NMR (400 MHz, CD3OD)δ 8.65 (s, 1H), 8.05 (d, J = 7.4 Hz, 2H), 7.67 (t, J = 7.4 Hz, 1H), 7.56 (t,J = 7.6 Hz, 2H), 5.05 (d, J = 4.5 Hz, 1H), 4.51 (dd, J = 9.4, 6.3 Hz, 1H), 4.30-4.01 (m, 5H), 4.03 – 3.88 (m, 3H), 3.88-3.75 (m, 2H), 3.39 (s, 3H).

[0120] Embodiment 15 (2-(2R,3S,4S,5R)-5-(6-chloro-4-(4-fluoro-2,3-dihydro-1H-inden-1-yl)amino)-1H-pyrazolo[3,4-d]pyrimidin-1-yl)-3,4-dihydroxytetrahydro-2H-pyran-2-yl)methoxy)-1-hydroxy-3-methoxypropane-2-yl)phosphonic acid

[0121] Compound 15 can be synthesized by referring to the synthesis steps of Example 1 using commercially available raw materials. 1 H NMR (400 MHz, DMSO- d6) δ 9.09 (d, J = 8.1 Hz, 1H), 8.16 (s, 1H), 7.26 (dd, J = 13.0, 7.7 Hz,1H), 7.11 (dd, J = 17.5, 8.2 Hz, 2H), 5.84 (q, J = 7.7 Hz, 1H), 4.78 (dd, J =11.3, 6.7 Hz, 1H), 4.25 (d, J = 7.5 Hz, 1H), 4.02 – 3.76 (m, 6H), 3.73 – 3.59(m, 4H), 3.28 (s, 3H), 3.14 – 3.03 (m, 1H), 2.93 (dd, J = 16.1, 8.0 Hz, 1H), 2.62 (dt, J = 8.2, 6.1 Hz, 1H), 2.02 (td, J = 16.2, 8.2 Hz, 1H).

[0122] Example 16 (2-(2R,3S,4S,5R)-5-(6-chloro-4-(4-fluoro-2,3-dihydro-1H-inden-1-yl)amino)-1H-pyrazolo[3,4-d]pyrimidin-1-yl)-3,4-dihydroxytetrahydro-2H-pyran-2-yl)methoxy)-1-hydroxy-3-methoxypropane-2-yl)phosphonic acid

[0123] Compound 16 can be synthesized by referring to the synthesis steps of Example 1 using commercially available raw materials. 1 H NMR (400 MHz, CD3OD)δ 8.04 (s, 1H), 7.24 – 7.08 (m, 4H), 5.62 (t, J = 5.8 Hz, 1H), 4.92 – 4.88(m, 1H), 4.50 – 4.40 (m, 1H), 4.27 – 3.73 (m, 10H), 3.37 (s, 3H), 2.96 – 2.75 (m, 2H), 2.17 – 1.84 (m, 4H).

[0124] Embodiment 17 2-((2R,3S,4S,5R)-5-(6-chloro-4-morpholinyl-1H-pyrazolo[3,4-d]pyrimidin-1-yl)-3,4-dihydroxytetrahydro-2H-pyran-2-yl)methoxy)-1-hydroxy-3-methoxypropan-2-yl)phosphonic acid

[0125] Compound 17 can be synthesized by referring to the synthesis steps of Example 1 using commercially available raw materials. 1 H NMR (400 MHz, CD3OD)δ 8.19 (d, J = 3.2 Hz, 1H), 4.92-4.80 (m, 1H), 4.46 – 4.39 (m, 1H), 4.22 – 4.15(m, 2H), 4.15 – 4.01 (m, 3H), 3.94 (dt, J = 24.3, 9.9 Hz, 7H), 3.85 – 3.77 (m,6H), 3.38 (d, J = 1.3 Hz, 3H).

[0126] Embodiment 18 2-(2R,3S,4S,5R)-5-(6-chloro-4-(5-fluoro-2,3-dihydro-1H-inden-1-yl)amino)-1H-pyrazolo[3,4-d]pyrimidin-1-yl)-3,4-dihydroxytetrahydro-2H-pyran-2-yl)methoxy)-1-hydroxy-3-methoxypropane-2-yl)phosphonic acid

[0127] Compound 18 can be synthesized by referring to the synthesis steps of Example 1 using commercially available raw materials. 1 H NMR (400 MHz, CD3OD)δ 8.05 (s, 1H), 7.32 – 7.27 (m, 1H), 7.00 (d, J = 8.0 Hz, 1H), 6.91 (m, 1H), 5.88 (t, J = 12 Hz, 1H), 4.92-4.80 (m, 1H), 4.44 (m, 1H), 4.24 – 4.06 (m,5H), 3.97 – 3.84 (m, 3H), 3.83 – 3.75 (m, 2H), 3.38 (s, 3H), 3.15 – 3.06 (m,1H), 2.94 (m, 1H), 2.72 – 2.58 (m, 1H), 2.07 (m, 1H).

[0128] Embodiment 19 2-(2R,3S,4S,5R)-5-(6-chloro-4-(4-methylpiperazin-1-yl)-1H-pyrazolo[3,4-d]pyrimidin-1-yl)-3,4-dihydroxytetrahydro-2H-pyran-2-yl)methoxy)-1-hydroxy-3-methoxypropan-2-yl)phosphonic acid

[0129] Compound 19 can be synthesized by referring to the synthesis steps of Example 1 using commercially available raw materials. 1 H NMR (400 MHz, CD3OD) δ 8.20 (s, 1H), 4.92 – 4.87 (m, 2H), 4.44 – 4.39 (m, 1H), 4.36 – 4.13 (m, 6H), 4.08 – 4.01 (m, 2H), 3.96 – 3.75 (m, 5H), 3.47 – 3.33 (m, 7H), 2.89 (s, 3H).

[0130] Embodiment 20 2-(2R,3S,4S,5R)-5-(6-chloro-4-(5-fluoro-2,3-dihydro-1H-inden-2-yl)amino)-1H-pyrazolo[3,4-d]pyrimidin-1-yl)-3,4-dihydroxytetrahydro-2H-pyran-2-yl)methoxy)-1-hydroxy-3-methoxypropane-2-yl)phosphonic acid

[0131] Compound 20 can be synthesized by referring to the synthesis steps of Example 1 using commercially available raw materials. 1 H NMR (400 MHz, CD3OD) δ8.05 (s, 1H), 7.25 – 7.19 (m, 1H), 6.99 (d, J = 8.7 Hz 1H), 6.89 (t, J = 8.9 Hz, 1H), 5.08 – 5.00 (m, 1H), 4.90-4.82 (m, 1H), 4.48-4.35 (m, 1H), 4.24 – 4.01 (m, 5H), 4.00 – 3.76 (m, 5H), 3.49 – 3.36 (m, 5H), 3.05-2.95 (m, 2H).

[0132] Embodiment 21 (1-(2H-tetrazolyl-5-yl)methoxy)-2-((2R,3S,4S,5R)-5-(6-chloro-4-(3,3-difluoroazetidin-1-yl)-1H-pyrazolo[3,4-d]pyrimidin-1-yl)-3,4-dihydroxytetrahydro-2H-pyran-2-yl)methoxy)-3-methoxypropane-2-yl)phosphonic acid

[0133]

[0134] Step 1: Synthesis of compound 21A Sodium hydroxide (24.4 mg, 0.61 mmol, 60% in mineral oil) was carefully added to a solution of diethyl (2-(((3aS,4R,7R,7aS)-7-(6-chloro-4-(3,3-difluoroazetidin-1-yl)-1H-pyrazolo[3,4-d]pyrimidin-1-yl)-2,2-dimethyltetrahydro-4H-[1,3]dioxo[4,5-c]pyran-4-yl)methoxy)-1-hydroxy-3-methoxypropan-2-yl)phosphonate 1M (200 mg, 0.305 mmol) and 5-(bromomethyl)-2-(2-(trimethylsilyl)ethoxy)methyl)-2H-tetrazole (179 mg, 0.610 mmol) in DMF (5 mL) under ice bath. The reaction solution was stirred at 0°C for 30 minutes and then quenched with saturated aqueous ammonium chloride solution. It was then extracted twice with ethyl acetate. The combined organic phase was washed once with saturated brine and dried over anhydrous sodium sulfate. The filtrate was filtered and concentrated under reduced pressure to obtain a crude product, which was purified by silica gel column chromatography (eluent: methanol / dichloromethane = 1 / 10, V / V) to obtain the title compound 21A (180 mg, 0.21 mmol, yield: 68.0%). LC / MS (ESI) m / z: 868 [M+H] + .

[0135] Step 2: Synthesis of compound 21 Triethylamine (0.22 mL, 1.61 mmol) and trimethylsilyl bromide (176 mg, 1.15 mmol) were added to a solution of diethyl (2-(((3aS,4R,7R,7aS)-7-(6-chloro-4-(3,3-difluoroazetidin-1-yl)-1H-pyrazolo[3,4-d]pyrimidin-1-yl)-2,2-dimethyltetrahydro-4H-[1,3]dioxo[4,5-c ]pyran-4-yl)methoxy)-1-methoxy-3-(2-(2-(2-(3-(trimethylsilyl)ethoxy)methyl)-2H-tetrazol-5-yl)methoxy)propan-2-yl)phosphonate 21A (130 mg, 0.15 mmol) in acetonitrile (5 mL) in an ice bath. The reaction mixture was stirred at room temperature overnight. After the phospholipids were completely hydrolyzed, the reaction solution was concentrated under reduced pressure. Then trifluoroacetic acid / water (3 / 1, 2 mL) was added to the obtained crude product. The reaction solution was stirred at room temperature for one hour. After the reaction was completed, the reaction solution was concentrated under reduced pressure to obtain a residue, which was purified by reverse phase preparative chromatography (eluent: acetonitrile / 0.1% formic acid aqueous solution = 25% ~ 90%, V / V) to obtain compound 21 (34.8 mg, yield: 36.2%). LC / MS (ESI) m / z: 642.2 [M+H] + . 1 H NMR (400 MHz, CD3OD) δ 7.98 –7.94 (s, 1H), 5.04 – 4.98 (m, 2H), 4.89 – 4.72(m 5H), 4.46 – 4.41 (m, 1H),4.29 – 3.70 (m, 10H), 3.36 (d, J = 4.0 Hz, 3H).

[0136] Embodiment 22 1-(2H-tetrazol-5-yl)methoxy)-2-((2R,3S,4S,5R)-5-(6-chloro-4-((S)-1-phenylethylamino)-1H-pyrazolo[3,4-d]pyrimidin-1-yl)-3,4-dihydroxytetrahydro-2H-pyran-2-yl)methoxy)-3-methoxypropane-2-yl)phosphonic acid

[0137] Compound 22 can be synthesized by referring to the synthesis steps of Example 21 using commercially available raw materials. LC / MS (ESI) m / z: 670.2[M+H] + . H NMR (400 MHz, CD3OD) δ 8.08 – 8.05(s, 1H), 7.41 (d, J= 7.4 Hz, 2H),7.32 (t, J = 7.6 Hz, 2H), 7.25 – 7.19 (m, 1H), 5.57 – 5.45 (m, 1H), 5.54 – 4.93(m, 2H), 4.42 – 4.36 (m, 1H), 4.21 – 4.73 (m, 11H), 3.41 – 3.32 (m, 3H), 1.61(d, J = 4.0 Hz, 3H).

[0138] Embodiment 23 1-(2H-tetrazolyl-5-yl)methoxy)-2-((2R,3S,4S,5R)-5-(6-chloro-4-hexahydrocyclopentylpyrrol-2(1H)yl)-1H-pyrazolo[3,4-d]pyrimidin-1-yl)-3,4-dihydroxytetrahydro-2H-pyran-2-yl)methoxy)-3-methoxypropane-2-yl)phosphonic acid

[0139] Compound 23 can be synthesized by referring to the synthesis steps of Example 21 using commercially available raw materials. LC / MS (ESI) m / z: 660.2[M+H] + . H NMR (400 MHz, CD3OD) δ 8.02 – 7.97 (s, 1H), 5.08 – 4.96 (m, 2H), 4.92–4.84 (m, 1H), 4.44 –4.38 (m, 1H), 4.29 – 3.73 (m, 12H), 3.64 – 3.53 (m, 2H),3.36 (d, J = 4.4 Hz, 3H), 3.03 – 2.88 (s, 1H), 2.87 – 2.76 (s, 1H), 2.04 – 1.54 (m, 6H).

[0140] Embodiment 24 1-(2H-tetrazol-5-yl)methoxy)-2-((2R,3S,4S,5R)-5-(6-chloro-4-((2,3-dihydro-1H-inden-2-yl)amino)-1H-pyrazolo[3,4-d]pyrimidin-1-yl)-3,4-dihydroxytetrahydro-2H-pyran-2-yl)methoxy)-3-methoxypropane-2-yl)phosphonic acid

[0141] Compound 24 can be synthesized by referring to the synthesis steps of Example 21 using commercially available raw materials. LC / MS (ESI) m / z: 682.2[M+H] + . 1 H NMR (400 MHz, CD3OD) δ 8.01 – 7.99 (s, 1H), 7.27 – 7.21 (m, 2H), 7.18 – 7.21 (m, 2H), 5.05 – 4.96 (m, 3H), 4.90 – 4.82 (m, 1H), 4.44 – 4.39(m, 1H), 4.21 – 3.98 (m, 7H), 3.96 – 3.90 (m, 1H), 3.86 – 3.74 (m, 2H), 3.46 – 3.35 (m, 5H), 3.03 – 2.93 (m, 2H).

[0142] Embodiment 25 1-(2H-tetrazolyl-5-yl)methoxy)-2-((2R,3S,4S,5R)-5-(6-chloro-4-((R)-1,2,3,4-tetrahydronaphthalen-1-amino)-1H-pyrazolo[3,4-d]pyrimidin-1-yl)-3,4-dihydroxytetrahydro-2H-pyran-2-yl)methoxy)-3-methoxypropane-2-yl)phosphonic acid

[0143] Compound 25 can be synthesized by referring to the synthesis steps of Example 21 using commercially available raw materials. LC / MS (ESI) m / z: 696.3[M+H] + . 1 H NMR (400 MHz, CD3OD) δ 8.01-7.98 (s, 1H), 7.25 – 7.09 (m, 4H), 5.63– 5.58 (m, 1H), 5.03 – 4.98 (m, 2H), 4.91 – 4.85 (m, 1H), 4.46 – 4.40 (m,1H), 4.22 – 3.99 (m, 7H), 3.97 – 3.90 (m, 1H), 3.86 – 3.73 (m, 2H), 3.36 (d, J = 2.6 Hz, 3H), 2.95– 2.75 (m, 2H), 2.18 – 1.79 (m, 4H).

[0144] Embodiment 26 1-(2H-tetrazolyl-5-yl)methoxy)-2-((2R,3S,4S,5R)-5-(4-(3-azabicyclo[3.1.0]hexan-3-yl)-6-chloropyrazolo[3,4-d]pyrimidin-1-yl)-3,4-dihydroxytetrahydro-2H-pyran-2-yl)methoxy)-3-methoxypropane-2-yl)phosphonic acid

[0145] Compound 26 can be synthesized by referring to the synthesis steps of Example 21 using commercially available raw materials. LC / MS (ESI) m / z: 632.2[M+H] + . 1 H NMR (400 MHz, DMSO- d 6) δ 8.16 (d, J = 8.4 Hz, 1H), 5.02 – 4.92 (m,2H), 4.78 – 4.70 (m, 1H), 4.24 – 4.16 (m, 1H), 3.98 – 3.79 (m, 9H), 3.72 –3.57 (m, 5H), 3.27 (s, 3H), 1.89 – 1.80 (m, 1H), 1.79 – 1.69 (m, 1H), 0.88 –0.79 (m, 1H), 0.23 – 0.17 (m, 1H).

[0146] Embodiment 27 1-(2H-tetrazol-5-yl)methoxy)-2-((2R,3S,4S,5R)-5-(6-chloro-4-((S)-tetrahydrofuran-3-amino)-1H-pyrazolo[3,4-d]pyrimidin-1-yl)-3,4-dihydroxytetrahydro-2H-pyran-2-yl)methoxy)-3-methoxypropane-2-yl)phosphonic acid

[0147] Compound 27 can be synthesized by referring to the synthesis steps of Example 21 using commercially available raw materials. LC / MS (ESI) m / z: 636.2[M+H] + . 1 H NMR (400 MHz, CD3OD) δ 8.03 (d, J = 11.2 Hz, 1H), 5.02 – 4.97 (m, 2H), 4.81 – 4.75 (m, 1H), 4.44 – 4.40 (m, 1H), 4.24 – 3.98 (m, 10H), 3.94 – 3.74(m, 5H), 3.36 (d,J = 3.6 Hz, 3H), 2.41 – 2.30 (m, 1H), 2.08 – 1.98 (m, 1H).

[0148] Embodiment 28 1-(2H-tetrazol-5-yl)methoxy)-2-((2R,3S,4S,5R)-5-(6-chloro-4-cyclopropylmethylamino)-1H-pyrazolo[3,4-d]pyrimidin-1-yl)-3,4-dihydroxytetrahydro-2H-pyran-2-yl)methoxy)-3-methoxypropane-2-yl)phosphonic acid

[0149] Compound 28 can be synthesized by referring to the synthesis steps of Example 21 using commercially available raw materials. LC / MS (ESI) m / z: 620.3[M+H] + . 1 H NMR (400 MHz, CD3OD) δ 8.02 (d, J = 10.0 Hz, 1H), 5.06 – 4.98 (m, 2H), 4.92 – 4.78 (m, 1H), 4.45 – 4.38 (m, 1H), 4.24 – 3.93 (m, 8H), 3.89 – 3.72(m, 2H), 3.46 – 3.34 (m, 5H), 1.20 – 1.06 (m, 1H), 0.61 – 0.50 (m, 2H), 0.38 – 0.29 (m, 2H).

[0150] Embodiment 29 1-(2H-tetrazol-5-yl)methoxy)-2-((2R,3S,4S,5R)-5-(6-chloro-4-cyclopentylamino)-1H-pyrazolo[3,4-d]pyrimidin-1-yl)-3,4-dihydroxytetrahydro-2H-pyran-2-yl)methoxy)-3-methoxypropane-2-yl)phosphonic acid

[0151] Compound 29 can be synthesized by referring to the synthesis steps of Example 21 using commercially available raw materials. 1 H NMR (400 MHz, CD3OD) δ8.01 (s, 1H), 5.02 (s, 2H), 4.86 – 4.81 (m, 1H), 4.59 – 4.48 (m, 1H), 4.45 –4.38 (m, 1H), 4.23 – 4.12 (m, 2H), 4.10 – 4.00 (m, 5H), 3.97 – 3.89 (m, 1H), 3.87 – 3.73 (m, 2H), 3.36 (s, 3H), 2.17 – 2.03 (m, 2H), 1.86 – 1.54 (m, 6H).

[0152] Embodiment 30 1-(2H-tetrazol-5-yl)methoxy)-2-((2R,3S,4S,5R)-5-(6-chloro-4-((2,3-dihydro-1H-inden-1-yl)amino)-1H-pyrazolo[3,4-d]pyrimidin-1-yl)-3,4-dihydroxytetrahydro-2H-pyran-2-yl)methoxy)-3-methoxypropan-2-yl)phosphonic acid

[0153] Compound 30 can be synthesized by referring to the synthesis steps of Example 21 using commercially available raw materials. 1 H NMR (400 MHz, CD3OD)δ 8.00 (d, J = 11.0 Hz, 1H), 7.32 – 7.15 (m, 4H), 5.92 (t, J = 7.3 Hz, 1H), 5.00(d, J = 5.7 Hz, 2H), 4.92-4.82 (m, 1H), 4.43-4.30 (m, 1H), 4.22 – 4.00 (m, 7H), 3.94 – 3.74 (m, 3H), 3.36 (d, J = 2.8 Hz, 3H), 3.13 – 3.05 (m, 1H), 2.94-2.77(m, 1H), 2.68 – 2.58 (m, 1H), 2.08 – 1.95 (m, 1H).

[0154] Embodiment 31 1-(2H-tetrazol-5-yl)methoxy)-2-((2R,3S,4S,5R)-5-(4-benzamido-6-chloro-1H-pyrazolo[3,4-d]pyrimidin-1-yl)-3,4-dihydroxytetrahydro-2H-pyran-2-yl)methoxy)-3-methoxypropane-2-yl)phosphonic acid

[0155] Compound 31 can be synthesized by referring to the synthesis steps of Example 21 using commercially available raw materials. 1 H NMR (400 MHz, CD3OD)δ 8.65-8.57 (m, 1H), 8.05 (d, J = 7.4 Hz, 2H), 7.67 (t, J = 7.4 Hz, 1H), 7.56(t, J = 7.6 Hz, 2H), 5.10-4.95 (m 3H), 4.55-4.48 (m, 1H), 4.19-3.75 (m, 10H), 3.39-3.35 (m, 3H).

[0156] Embodiment 32 1-(2H-tetrazol-5-yl)methoxy)-2-((2R,3S,4S,5R)-5-(6-chloro-4-(2,3-dihydro-1H-inden-4-yl)amino)-1H-pyrazolo[3,4-d]pyrimidin-1-yl)-3,4-dihydroxytetrahydro-2H-pyran-2-yl)methoxy)-3-methoxypropane-2-yl)phosphonic acid

[0157] Compound 32 can be synthesized by referring to the synthesis steps of Example 21 using commercially available raw materials. 1 H NMR (400 MHz, CD3OD)δ 7.34 – 7.10 (m, 3H), 5.02 – 4.95 (m, 2H), 4.92 – 4.86 (m, 1H), 4.46 – 4.32(m, 1H), 4.18 – 3.75 (m, 10H), 3.35 (s, 3H), 3.00 (t, J = 7.4 Hz, 2H), 2.81 (t, J = 7.1 Hz, 2H), 2.13 – 1.99 (m, 2H).

[0158] Embodiment 33 1-(2H-tetrazol-5-yl)methoxy)-2-((2R,3S,4S,5R)-5-(6-chloro-4-((2,3-dihydro-1H-inden-5-yl)amino)-1H-pyrazolo[3,4-d]pyrimidin-1-yl)-3,4-dihydroxytetrahydro-2H-pyran-2-yl)methoxy)-3-methoxypropane-2-yl)phosphonic acid

[0159] Compound 33 can be synthesized by referring to the synthesis steps of Example 21 using commercially available raw materials. 1 H NMR (400 MHz, CD3OD)δ 7.71 – 7.12 (m, 3H), 5.00 (d, J = 5.8 Hz, 2H), 4.92-4.80 (m, 1H), 4.45-4.35(m, 1H), 4.20 – 3.76 (m, 10H), 3.36 (d, J = 3.1 Hz, 3H), 2.98-2.80 (m, 4H), 2.16-2.09 (m, 2H).

[0160] Embodiment 34 1-(2H-tetrazol-5-yl)methoxy)-2-((2R,3S,4S,5R)-5-(6-chloro-4-(4-fluoro-2,3-dihydro-1H-indene-1-amino)-1H-pyrazolo[3,4-d]pyrimidin-1-yl)-3,4-dihydroxytetrahydro-2H-pyran-2-yl)methoxy)-3-methoxypropane-2-yl)phosphonic acid

[0161] Compound 34 can be synthesized by referring to the synthesis steps of Example 21 using commercially available raw materials. 1 H NMR (400 MHz, DMSO)δ 9.09 (d, J = 8.1 Hz, 1H), 8.13 (m, 1H), 7.29-7.24 (m, 1H), 7.14-7.08 (m,2H), 5.84 (q, J = 7.7 Hz, 1H), 4.97 (s, 2H), 4.78-4.70 (m, 1H), 4.25-4.21 (m,1H), 4.02 – 3.76 (m, 6H), 3.73 – 3.59 (m, 4H), 3.28 (s, 3H), 3.14 – 3.03 (m,1H), 2.93-2.81 (m, 1H), 2.65-2.52 (m, 1H), 2.05-1.98 (m, 1H).

[0162] Embodiment 35 1-(2H-tetrazolyl-5-yl)methoxy)-2-((2R,3S,4S,5R)-5-(6-chloro-4-((S)-1,2,3,4-tetrahydronaphthalen-1-amino)-1H-pyrazolo[3,4-d]pyrimidin-1-yl)-3,4-dihydroxytetrahydro-2H-pyran-2-yl)methoxy)-3-methoxypropane-2-yl)phosphonic acid

[0163] Compound 35 can be synthesized by referring to the synthesis steps of Example 21 using commercially available raw materials. 1 H NMR (400 MHz, CD3OD)δ 7.99 (d, J = 12.0 Hz, 1H), 7.22 (d, J = 8 Hz, 1H), 7.17 – 7.10 (m, 3H), 5.62 (t, J = 12 Hz, 1H), 5.03 – 4.99 (m, 2H), 4.89 (s, 1H), 4.43 (m, 1H), 4.22 – 4.00 (m, 7H), 3.93 (m, 1H), 3.82 (m, 2H), 3.38 – 3.34 (m, 3H), 2.85 (m, 2H), 2.11 (m, 1H), 2.05 – 1.84 (m, 3H).

[0164] Embodiment 36 1-(2H-tetrazol-5-yl)methoxy)-2-((2R,3S,4S,5R)-5-(6-chloro-4-morpholinyl-1H-pyrazolo[3,4-d]pyrimidin-1-yl)-3,4-dihydroxytetrahydro-2H-pyran-2-yl)methoxy)-3-methoxypropane-2-yl)phosphonic acid

[0165] Compound 36 can be synthesized by referring to the synthesis steps of Example 21 using commercially available raw materials. 1 H NMR (400 MHz, CD3OD) δ 8.11 (d, J = 17.1 Hz, 1H), 5.07 – 4.97 (m, 2H), 4.92-4.80 (m, 1H), 4.45-4.35 (m, 1H), 4.28 – 4.20 (m, 1H), 4.18 – 3.89 (m, 12H), 3.83 (t, J = 7.1Hz, 5H), 3.37 (d, J = 4.2 Hz, 3H).

[0166] Embodiment 37 1-(2H-tetrazol-5-yl)methoxy)-2-((2R,3S,4S,5R)-5-(6-chloro-4-(5-fluoro-2,3-dihydro-1H-indene-1-amino)-1H-pyrazolo[3,4-d]pyrimidin-1-yl)-3,4-dihydroxytetrahydro-2H-pyran-2-yl)methoxy)-3-methoxypropan-2-yl)phosphonic acid

[0167] Compound 37 can be synthesized by referring to the synthesis steps of Example 21 using commercially available raw materials. 1 H NMR (400 MHz, CD3OD)δ 7.99 (d, J = 12 Hz, 1H), 7.31 (m, 1H), 7.00 (d, J = 12 Hz, 1H), 6.91 (t, J= 8 Hz, 1H), 5.87 (t, J = 8 Hz, 1H), 5.05 – 4.97 (m, 2H), 4.9 (m, 1H), 4.43 (m, 1H), 4.20 – 4.02 (m, 7H), 3.94 (m, 1H), 3.86 – 3.73 (m, 2H), 3.36 (d, J =8 Hz, 3H), 3.14 – 3.05 (m, 1H), 2.97 (m, 1H), 2.66 (m, 1H), 2.05 (m, 1H).

[0168] Embodiment 38 1-(2H-tetrazol-5-yl)methoxy)-2-((2R,3S,4S,5R)-5-(6-chloro-4-(5-fluoro-2,3-dihydro-1H-indene-2-amino)-1H-pyrazolo[3,4-d]pyrimidin-1-yl)-3,4-dihydroxytetrahydro-2H-pyran-2-yl)methoxy)-3-methoxypropane-2-yl)phosphonic acid

[0169] Compound 38 can be synthesized by referring to the synthesis steps of Example 21 using commercially available raw materials. 1 H NMR (400 MHz, CD3OD)δ 8.00 (d, J = 10.4 Hz, 1H), 7.26 – 7.19 (m, 1H), 6.98 (d, J = 8.4 Hz, 1H), 6.89 (t, J=8.8 Hz, 1H), 5.08 – 4.96 (m, 3H), 4.92-4.81 (m, 1H), 4.47 – 4.40 (m, 1H), 4.23 –3.99 (m, 7H), 3.93-.3.87 (m, 1H), 3.86 – 3.74 (m, 2H), 3.46 – 3.34 (m, 5H), 3.05 –2.90 (m, 2H).

[0170] Embodiment 39 (R)-2-(2R,3S,4S,5R)-5-(6-chloro-4-cyclopentylamino)-1H-pyrazolo[3,4-d]pyrimidin-1-yl)-3,4-dihydroxytetrahydro-2H-pyran-2-yl)methoxy)-1-hydroxy-3-methoxypropan-2-yl)phosphonic acid (39-1) (S)-2-(2R,3S,4S,5R)-5-(6-chloro-4-cyclopentylamino)-1H-pyrazolo[3,4-d]pyrimidin-1-yl)-3,4-dihydroxytetrahydro-2H-pyran-2-yl)methoxy)-1-hydroxy-3-methoxypropan-2-yl)phosphonic acid (39-2)

[0171] The reaction equation is as follows:

[0172] Step 1: Synthesis of compound 39A Cyclopentylamine (1.4 mL, 13.8 mmol) was slowly added dropwise to a solution of (4aR, 7R, 8S, 8aS)-7-(4,6-dichloro-1H-pyrazolo[3,4-d]pyrimidin-1-yl)-2,2-dimethylhexahydropyrano[3,2-d][1,3]dioxin-8-ol 1G (4.7 g, 12.5 mmol) and triethylamine (3.5 mL, 25.1 mmol) in ethanol (50 mL) under ice bath. Solution The reaction solution was slowly heated to room temperature and stirred at room temperature for one hour. After the reaction was completed, the reaction solution was concentrated, and the crude product was diluted with ethyl acetate and washed once with 10% citric acid, and then washed once with saturated sodium bicarbonate and saturated brine. The organic phase was dried over sodium sulfate, filtered, and concentrated under reduced pressure to give the title compound 39A (4.1 g, light yellow solid, yield: 77.2%). LC / MS (ESI) m / z: 424 (M+H) + ; 1 H NMR (400 MHz, DMSO- d 6) δ 8.64 (d, J= 7.0 Hz, 1H), 8.19 (s, 1H), 4.94 – 4.80 (m, 2H), 4.68 – 4.61 (m, 1H), 4.47 – 4.38 (m, 1H), 4.30 – 4.26 (m, 1H), 3.96 – 3.87 (m, 1H), 3.81 – 3.74 (m, 1H), 3.71 – 3.61 (m, 2H), 3.55 – 3.50(m, 1H), 2.05 – 2.00 (m, 1H), 1.78 – 1.52 (m, 7H), 1.50 (s, 3H), 1.28 (s,3H).

[0173] Step 2: Synthesis of compound 39B (4aR, 7R, 8S, 8aS)-7-(6-chloro-4-(cyclopentylamino)-1H-pyrazolo[3,4-d]pyrimidin-1-yl)-2,2-dimethylhexahydropyrano[3,2-d][1,3]dioxin-8-ol 39A (4.1 g, 9.67 mmol) was dissolved in acetic acid (10 mL) and water (10 mL). The reaction solution was stirred at 60°C for 1 hour. After the reaction was completed, the solvent was removed by concentration under reduced pressure, and then water (50 mL) was added to the residue, and the mixture was extracted with ethyl acetate (200 mL), and the organic phase was washed with saturated brine. The organic phase was dried over anhydrous sodium sulfate, filtered, and concentrated under reduced pressure. The crude product was purified by silica gel column chromatography (eluent: methanol / dichloromethane = 1 / 10, V / V) to obtain the title compound 39B (3.1 g, white solid, yield: 83.5%). LC-MS m / z(ESI): 384 (M+H + ).

[0174] Step 3: Synthesis of compound 39C (2R, 3S, 4S, 5R)-5-(6-chloro-4-(cyclopentylamino)-1H-pyrazolo[3,4-d]pyrimidin-1-yl)-2-(hydroxymethyl)tetrahydro-2H-pyran-3,4-diol 39B (2.9 g, 7.56 mmol) was dissolved in a dry tetrahydrofuran (50 mL) solution, and then TBSCl (1.71 g, 11.3 mmol) and imidazole (0.77 g, 11.3 mmol) were added under ice bath. The reaction solution was stirred at room temperature for 2 hours. After the reaction was completed, the mixture was diluted with ethyl acetate (100 mL) and washed with saturated brine. The organic phase was dried over anhydrous sodium sulfate, filtered, and concentrated under reduced pressure. The crude product was purified by silica gel column chromatography (eluent: methanol / dichloromethane = 1 / 20, V / V) to give the title compound 39C (3.5 g, light yellow oil, 93.0% yield). LC-MS m / z (ESI): 498 (M+H + ).

[0175] Step 4: Synthesis of compound 39D DIEA (4.6 mL, 28.1 mmol) and MOMCl (2.1 mL, 28.1 mmol) were added to a THF solution (50 mL) of (2R, 3S, 4S, 5R)-2-(((tert-butyldimethylsilyl)oxy)methyl)-5-(6-chloro-4-(cyclopentylamino)-1H-pyrazolo[3,4-d]pyrimidin-1-yl)tetrahydro-2H-pyran-3,4-diol 39C (3.50 g, 7.03 mmol). The reaction solution was stirred at 60°C overnight. After the reaction was completed, the reaction solution was diluted with ethyl acetate and washed with saturated brine. The organic phase was dried over anhydrous sodium sulfate, filtered, and concentrated under reduced pressure. The crude product was purified by silica gel column chromatography with an eluent (ethyl acetate / petroleum ether = 1 / 10, V / V) to give the title compound 39D (1.5 g, light yellow oil, yield 36.4%). LC-MS m / z (ESI):586 (M+H + ); 1 H NMR (400 MHz, DMSO- d 6) δ 8.60 (d, J= 6.8 Hz, 1H), 8.17 (s, 1H),4.92 – 4.85 (m, 1H), 4.72 (s, 2H), 4.56 – 4.50 (m, 2H), 4.47 – 4.29 (m, 2H),4.25 – 4.20 (m, 1H), 4.12 – 4.01 (m, 1H), 3.90 – 3.80 (m, 4H), 3.32 (s, 3H), 3.01 (s, 3H), 2.05 – 1.92 (m, 2H), 1.77 – 1.48 (m, 6H), 0.86 (s, 9H), 0.05(d, J = 6.3 Hz, 6H).

[0176] Step 5: Synthesis of compound 39E 1-(3R,4S,5S,6R)-6-(((tert-butyldimethylsilyloxy)methyl)-4,5-bis(methoxymethoxy)tetrahydro-2H-pyran-3-yl)-6-chloro-N-cyclopentyl-1H-pyrazolo[3,4-d]pyrimidin-4-amine 39D (1.5 g, 2.56 mmol) was added to a tetrahydrofuran solution of tetra-n-butylammonium fluoride (3.8 mL, 3.84 mmol, 1M). The reaction solution was stirred at room temperature for 1 hour. After the reaction was completed, the reaction solution was diluted with ethyl acetate (100 mL) and washed with saturated brine. The organic phase was dried over anhydrous sodium sulfate, filtered, and concentrated under reduced pressure. The crude product was purified by silica gel column chromatography (eluent: methanol / dichloromethane = 1 / 10, V / V) to give the title compound 39E (3.5 g, light yellow oil, yield: 93.0%). LC-MS m / z (ESI):472 (M+H + ).

[0177] Step 6: Synthesis of compound 39F Rhodium dimer acetate (53 mg, 0.12 mmol) was added to a toluene solution (15 mL) of (2R, 3S, 4S, 5R)-5-(6-chloro-4-(cyclopentylamino)-1H-pyrazolo[3,4-d]pyrimidin-1-yl)-3,4-bis(methoxymethoxy)tetrahydro-2H-pyran-2-yl)methanol 39E (1.1 g, 2.33 mmol) and 1-(diethoxyphosphoryl)-2-ethoxy-2-oxoethane-1-diazide (1.76 g, 6.99 mmol). The reaction solution was immediately replaced with nitrogen three times and then stirred at 100°C for 3 hours. After the reaction was completed, the mixture was diluted with ethyl acetate (100 mL) and washed once with saturated NaHCO3 and saturated brine. The organic phase was dried over anhydrous sodium sulfate, filtered, and concentrated under reduced pressure. The obtained crude product was purified by silica gel column chromatography (eluent: methanol / dichloromethane = 1 / 10, V / V) to give the title compound 39F (0.96 g, yellow oil, yield 59.3%). LC-MS m / z (ESI): 694 (M+H + ).

[0178] Step 7: Synthesis of compound 39G At -15 ° C, NaHMDS (3.5 mL, 3.5 mmol, 1 M in THF) was slowly added dropwise to a solution of ethyl 2-((2R, 3S, 4S, 5R)-5-(6-chloro-4-(cyclopentylamino)-1H-pyrazolo[3,4-d]pyrimidin-1-yl)-3,4-bis(methoxymethoxy)tetrahydro-2H-pyran-2-yl)methoxy)-2-(diethoxyphosphoryl)acetate 39F (600 mg, 0.86 mmol) in anhydrous tetrahydrofuran, stirred at this temperature for 15 minutes, and then TBAI (160 mg, 0.43 mmol) and MOMCl (0.4 mL, 5.19 mmol) were quickly added to the reaction solution. After the addition was complete, the reaction solution was stirred at this temperature for 1 hour. After the reaction was completed, the reaction solution was quenched with saturated ammonium chloride solution and extracted three times with ethyl acetate. The combined organic phase was washed with saturated brine, dried over anhydrous sodium sulfate, filtered, and concentrated under reduced pressure. The crude product was purified by silica gel column chromatography (eluent: methanol / dichloromethane = 1 / 10, V / V) to give the title compound 39G (480 mg, yellow oil, yield 71.0%). LC / MS (ESI) (m / z): 782 (M+H) + .

[0179] Step 8: Synthesis of compounds 39H-1 and 39H-2 Calcium chloride (0.81 g, 7.29 mmol) and sodium borohydride (0.25 g, 7.29 mmol) were added to an ethanol (25 mL) solution of ethyl 2-((2R, 3S, 4S, 5R)-5-(6-chloro-4-(cyclopentyl(methoxymethyl)amino)-1H-pyrazolo[3,4-d]pyrimidin-1-yl)-3,4-bis(methoxymethoxy)tetrahydro-2H-pyran-2-yl)methoxy)-2-(diethoxyphosphoryl)-3-methoxypropanoate 39G (1.9 g, 2.43 mmol) in an ice bath. The reaction mixture was stirred at room temperature overnight. After the reaction was completed, the reaction was quenched with saturated aqueous ammonium chloride solution and extracted with ethyl acetate (100 mL) three times. The combined organic layer was washed once with saturated brine, dried over anhydrous sodium sulfate, filtered and concentrated under reduced pressure, and purified by silica gel column chromatography (eluent: methanol / dichloromethane = 1 / 10, V / V) to obtain isomers (1.4 g). Then, chiral separation (column model: ChiralCel OX, 250×21.2mm ID, 5µm; mobile phase A: CO2, mobile phase B: MeOH (0.1%7mol / L NH3in MeOH); separation gradient: mobile phase B 0-42%; flow rate: 40mL / min) was performed to obtain the first point 39H-1 (0.9 g, yield: 50%) with an elution time of 2.5 minutes and the second point 39H-2 (0.4 g, yield: 22%) with an elution time of 3.79 minutes. LC / MS (ESI) (m / z): 782 (M+H) + .

[0180] Step 9: Synthesis of compound 39-1 Under ice bath, triethylamine (0.14 mL, 1.01 mmol) and trimethylsilyl bromide (0.09 mL, 0.68 mmol) were added to a solution of diethyl ((R)-2-((2R, 3S, 4S, 5R)-5-(6-chloro-4-(cyclopentyl(methoxymethyl)amino)-1H-pyrazolo[3,4-d]pyrimidin-1-yl)-3,4-bis(methoxymethoxy)tetrahydro-2H-pyran-2-yl)methoxy)-1-hydroxy-3-methoxypropane-2-yl)phosphonate 39H-1 (50 mg, 0.07 mmol) in acetonitrile (3 mL) in sequence. The reaction solution was stirred at room temperature overnight. After the phospholipids were completely hydrolyzed, the reaction solution was concentrated under reduced pressure. Then trifluoroacetic acid / water (3 / 1, 2 mL) was added to the obtained crude product. The reaction solution was stirred at room temperature for 3 hours. After the reaction was completed, the reaction solution was concentrated under reduced pressure to obtain a residue, which was purified by reverse phase preparative chromatography (eluent: acetonitrile / 0.1% trifluoroacetic acid aqueous solution = 25%-90%, V / V) to obtain the title compound 39-1 (7 mg, yield: 17.5%). LC / MS (ESI) (m / z): 552 (M+H) + ;1H NMR (400 MHz, CD3OD) δ 8.08 (s, 1H), 4.86 – 4.81 (m, 1H), 4.61 – 4.49 (m, 1H), 4.45 – 4.41 (m, 1H), 4.25 – 4.03 (m, 5H), 3.96 – 3.90 (m, 3H), 3.81 (d, J =10.6 Hz, 2H), 3.38 (s, 3H), 2.18 – 2.02 (m, 2H), 1.87 – 1.54 (m, 6H).

[0181] Compound 39-2. Compound 39-2 can be successfully prepared by using intermediate 39H-2 as a raw material and referring to the synthesis of compound 39-1. 1 H NMR (400 MHz, CD3OD) δ 8.07 (s, 1H), 4.85 – 4.83 (m, 1H), 4.59 – 4.49(m, 1H), 4.43 – 4.39 (m, 1H), 4.25 – 4.01 (m, 5H), 4.00 – 3.84 (m, 3H), 3.80(d, J = 11.0 Hz, 2H), 3.38 (s, 3H), 2.17 – 2.04 (m, 2H), 1.86 – 1.51 (m, 6H).

[0182] Embodiment 40 (R)-2-((2R,3S,4S,5R)-5-(6-chloro-4-piperazin-1-yl)-1H-pyrazolo[3,4-d]pyrimidin-1-yl)-3,4-dihydroxytetrahydro-2H-pyran-2-yl)methoxy)-1-hydroxy-3-methoxypropane-2-yl)phosphonic acid (40-1) (S)-2-((2R,3S,4S,5R)-5-(6-chloro-4-piperazin-1-yl)-1H-pyrazolo[3,4-d]pyrimidin-1-yl)-3,4-dihydroxytetrahydro-2H-pyran-2-yl)methoxy)-1-hydroxy-3-methoxypropan-2-yl)phosphonic acid (40-2)

[0183] Compounds 40-1 and 40-2 can be synthesized by referring to the synthesis steps of Example 39 using commercially available raw materials.

[0184] 40-1: 1 H NMR (400 MHz, D2O) δ 8.20 (s, 1H), 4.90-4.81 (m, 1H), 4.41(dd, J = 7.4, 3.2 Hz, 1H), 4.22 – 3.95 (m, 10H), 3.90 – 3.69 (m, 4H), 3.46 –3.35 (m, 4H), 3.31 (d, J = 3.3 Hz, 3H).

[0185] 40-2: 1 H NMR (400 MHz, D2O) δ 8.19 (s, 1H), 4.86 (m, 1H), 4.41 (dd, J =7.3, 3.3 Hz, 1H), 4.22 – 3.92 (m, 10H), 3.88 – 3.66 (m, 4H), 3.49 – 3.35 (m, 4H), 3.32 (s, 3H).

[0186] Embodiment 41 (R)-2-((2R,3S,4S,5R)-5-(4-amino-6-chloro-1H-pyrazolo[3,4-d]pyrimidin-1-yl)-3,4-dihydroxytetrahydro-2H-pyran-2-yl)methoxy)-1-hydroxy-3-methoxypropan-2-yl)phosphonic acid (41-1) (S)-2-((2R,3S,4S,5R)-5-(4-amino-6-chloro-1H-pyrazolo[3,4-d]pyrimidin-1-yl)-3,4-dihydroxytetrahydro-2H-pyran-2-yl)methoxy)-1-hydroxy-3-methoxypropan-2-yl)phosphonic acid (41-2)

[0187]

[0188] Step 1: Synthesis of Compound 41A In a 200 mL single-necked bottle, (4aR, 7R, 8S, 8aS)-7-(6-chloro-4-((2,3-dihydro-1H-inden-4-yl)amino)-1H-pyrazolo[3,4-d ]pyrimidin-1-yl)-2,2-dimethylhexahydropyrano[3,2-d ][1,3]dioxin-8-ol 1G (3 g, 8.00 mmol), triethylamine (3.3 mL, 24.0 mmol) and ethyl acetate (50 mL) were added, and then bis(2,4-dimethoxybenzyl)amine (2.79 g, 8.80 mmol) was added under ice bath. The reaction solution was stirred at room temperature for 1 hour. After the reaction was completed, the reaction solution was poured into water and extracted twice with ethyl acetate. The combined organic phase was washed with saturated brine, dried over anhydrous sodium sulfate, and concentrated under reduced pressure to obtain a crude product, which was purified by silica gel column chromatography (eluent: ethyl acetate / petroleum ether = 1 / 1, V / V) to obtain the title compound 41A (4.6 g, white solid, yield: 87.7%).

[0189] Step 2: Synthesis of compound 41B (4aR, 7R, 8S, 8aS)-7-(4-(bis(3,4-dimethoxybenzyl)amino)-6-chloro-1H-pyrazolo[3,4-d]pyrimidin-1-yl)-2,2-dimethylhexahydropyrano[3,2-d][1,3]dioxin-8-ol 41A (4.5 g, 6.86 mmol) was added to a mixed solution of acetic acid (15 mL) and water (15 mL). The reaction was stirred at 40°C for 1 h. After the reaction was completed, the mixture was concentrated under reduced pressure and purified by silica gel column chromatography (eluent: dichloromethane / methanol = 10 / 1, V / V) to obtain the title compound 41B (4.0 g, white solid, yield: 94.7%).

[0190] Step 3: Synthesis of compound 41C To a solution of (2R,3S,4S,5R)-5-(4-(bis(3,4-dimethoxybenzyl)amino)-6-chloro-1H-pyrazolo[3,4-d]pyrimidin-1-yl)-2-(hydroxymethyl)tetrahydro-2H-pyran-3,4-diol 41B (4 g, 6.49 mmol) in acetone (30 mL) were added p-toluenesulfonic acid (3.35 g, 19.5 mmol) and 2,2-dimethoxypropane (4.06 g, 39.0 mmol). The reaction mixture was stirred at room temperature overnight. After the reaction was completed, the reaction mixture was diluted with ethyl acetate (200 mL) and washed twice with saturated sodium bicarbonate (100 mL) and once with saturated brine. The organic phase was dried over anhydrous sodium sulfate and concentrated under reduced pressure to obtain a crude product. The crude product was purified by silica gel column chromatography (eluent: ethyl acetate / petroleum ether = 1 / 1, V / V) to obtain the title compound 41C (1.7 g, colorless oil, yield: 39.9%).

[0191] Step 4: Synthesis of compound 41D To a toluene (20 mL) solution of (3aS, 4R, 7R, 7aS)-7-(4-(bis(3,4-dimethoxybenzyl)amino)-6-chloro-1H-pyrazolo[3,4-d]pyrimidin-1-yl)-2,2-dimethyltetrahydro-4H-[1,3]dioxo[4,5-c ]pyran-4-yl)methanol 41C (1.5 g, 2.28 mmol) were added 1-(diethoxyphosphoryl)-2-ethoxy-2-oxoethane-1-diazide (1.72 g, 6.86 mmol) and rhodium dimer acetate (0.05 g, 0.114 mmol) in sequence. The mixture was replaced with nitrogen three times and then stirred at 100°C for 3 h. After the reaction was completed, ethyl acetate (200 mL) was added to the reaction solution and washed once with saturated sodium bicarbonate and saturated brine. The organic phase was dried over anhydrous sodium sulfate and concentrated under reduced pressure to obtain a crude product. The crude product was purified by silica gel column chromatography (eluent: ethyl acetate / petroleum ether = 4 / 1, V / V) to obtain the title compound 41D (1.2 g, yellow oil, yield: 59.8%). LC / MS (ESI) m / z: 878 [M+H] + .

[0192] Step 5: Synthesis of compound 41E Under N2 atmosphere, a solution of ethyl 2-((3aS,4R,7R,7aS)-7-(4-(bis(3,4-dimethoxybenzyl)amino)-6-chloro-1H-pyrazolo[3,4-d]pyrimidin-1-yl)-2,2-dimethyltetrahydro-4H-[1,3]dioxo[4,5-c]pyran-4-yl)methoxy)-2-(diethoxyphosphoryl)acetate 41D (1.2 g, 1.37 mmol) in THF (15 mL) was cooled to -20°C, and then NaHMDS (2.73 mL, 2.73 mmol, 1 M in tetrahydrofuran solution) was slowly added dropwise at this temperature. The reaction solution was stirred at -20°C for 30 min. Then TBAI (0.25 g, 0.683 mmol) was added to the reaction solution, and chloromethyl methyl ether (0.415 mL, 5.46 mmol) was added dropwise. The reaction solution was stirred at -20 °C for 1 hour. After the reaction was completed, the reaction was quenched with saturated aqueous ammonium chloride solution and extracted three times with ethyl acetate (20 mL). The combined organic layer was washed once with saturated brine, dried over anhydrous sodium sulfate, filtered and concentrated under reduced pressure to obtain a crude product. The crude product was purified by silica gel column chromatography (eluent: ethyl acetate / petroleum ether = 5 / 1, V / V) to obtain the title compound 41E (0.9 g, yellow oil, yield: 71.4%).

[0193] Step 6: Synthesis of Compound 41F Calcium chloride (324 mg, 2.93 mmol) and sodium borohydride (111 mg, 2.93 mmol) were added to a solution of ethyl 2-((3aS, 4R, 7R, 7aS)-7-(4-(bis(3,4-dimethoxybenzyl)amino)-6-chloro-1H-pyrazolo[3,4-d]pyrimidin-1-yl)-2,2-dimethyltetrahydro-4H-[1,3]dioxo[4,5-c]pyran-4-yl)methoxy)-2-(diethoxyphosphoryl)-3-methoxypropanoate 41E (900 mg, 0.976 mmol) in ethanol (10 mL) in an ice bath. The reaction mixture was stirred at room temperature overnight. After the reaction was completed, the reaction was quenched with saturated aqueous ammonium chloride and extracted with ethyl acetate (30 mL) three times. The combined organic layer was washed once with saturated brine, dried over anhydrous sodium sulfate, filtered and concentrated under reduced pressure to obtain a crude product. The crude product was purified by silica gel column chromatography (eluent: methanol / dichloromethane = 1 / 10, V / V) to obtain the title compound 41F (500 mg, yellow oil, yield: 58.2%).

[0194] Step 7: Synthesis of compounds 41-1 and 41-2 Triethylamine (0.3 mL, 2.39 mmol) and trimethylsilyl bromide (261 mg, 1.70 mmol) were added to a solution of diethyl (2-(((3aS, 4R, 7R, 7aS)-7-(4-(bis(3,4-dimethoxybenzyl)amino)-6-chloro-1H-pyrazolo[3,4-d ]pyrimidin-1-yl)-2,2-dimethyltetrahydro-4H-[1,3]dioxo[4,5-c ]pyran-4-yl)methoxy)-1-hydroxy-3-methoxypropane-2-yl)phosphonate 41F (150 mg, 0.170 mmol) in acetonitrile (5 mL) in an ice bath. The reaction mixture was stirred at room temperature overnight. After the phospholipids were completely hydrolyzed, the reaction mixture was concentrated under reduced pressure. Then trifluoroacetic acid / water (3 / 1, 2 mL) was added to the crude product. The reaction was stirred at room temperature for another hour. After the reaction was completed, the reaction solution was concentrated under reduced pressure to obtain a residue, which was purified by reverse phase preparative chromatography (eluent: acetonitrile / 0.1% formic acid aqueous solution = 25% ~ 90%, V / V) to obtain (S)-2-((2R, 3S, 4S, 5R)-5-(4-amino-6-chloro-1H-pyrazolo[3,4-d]pyrimidin-1-yl)-3,4-dihydroxytetrahydro-2H-pyran-2-yl)methoxy)-1-hydroxy-3-methoxypropane-2-yl)phosphonic acid (41-1, 9 mg, white solid, 10.9%) and (R)-2-((2R, 3S, 4S, 5R)-5-(4-amino-6-chloro-1H-pyrazolo[3,4-d]pyrimidin-1-yl)-3,4-dihydroxytetrahydro-2H-pyran-2-yl)methoxy) -1-hydroxy-3-methoxypropan-2-yl)phosphonic acid (41-2, 12 mg, white solid, yield: 14.6%).

[0195] 41-1: 1 H NMR (400 MHz, CD3OD) δ 8.06 (s, 1H), 4.92-4.80 (m, 1H), 4.45 (s, 1H), 4.25-4.19 (m, 2H), 4.18-4.02 (m, 3H), 3.92-4.80 (m, 3H), 3.84-3.75(m, 2H), 3.38(s, 3H).

[0196] 41-2: 1 H NMR (400 MHz, CD3OD) δ 8.06 (s, 1H), 4.92-4.82 (m, 1H), 4.43 (s, 1H), 4.25-3.75 (m, 10H), 3.38 (s, 3H).

[0197] Embodiment 42 (S)-1-(2H-tetrazol-5-yl)methoxy)-2-((2R,3S,4S,5R)-5-(6-chloro-4-cyclopentylamino)-1H-pyrazolo[3,4-d]pyrimidin-1-yl)-3,4-dihydroxytetrahydro-2H-pyran-2-yl)methoxy)-3-methoxypropane-2-yl)phosphonic acid

[0198]

[0199] Step 1: Synthesis of compound 42A Diethyl ((R)-2-((2R,3S,4S,5R)-5-(6-chloro-4-(cyclopentyl(methoxymethyl)amino)-1H-pyrazolo[3,4-d]pyrimidin-1-yl)-3,4-bis(methoxymethoxy)tetrahydro-2H-pyran-2-yl)methoxy)-1-hydroxy-3-methoxypropan-2-yl)phosphonate 39H-1 (0.85 g, 1.15 mmol) and 5-(bromomethyl)-2-(2-(trimethylsilyl)ethoxy)methyl)-2H-tetrazole (0.67 g, 2.30 mmol) were dissolved in dry DMF (8 mL). It was cooled to 0°C and then sodium hydroxide (140 mg, 3.45 mmol, 60% in mineral oil) was carefully added to the mixture. The reaction was stirred at 0°C for half an hour. After the reaction was completed, the reaction was quenched with a saturated aqueous ammonium chloride solution and extracted twice with ethyl acetate (50 mL). The combined organic phase was washed once with saturated brine, dried over anhydrous sodium sulfate, filtered, concentrated under reduced pressure, and purified by silica gel column chromatography (eluent: methanol / dichloromethane = 1 / 10, V / V) to obtain the title compound 42A (1.0 g, light yellow oil, yield: 91%). LC / MS (ESI) (m / z): 952 (M+H) + .

[0200] Step 2: Synthesis of compound 42 Triethylamine (2.19 mL, 15.8 mmol) and trimethylsilyl bromide (1.39 mL, 10.50 mmol) were added to a solution of diethyl ((S)-2-((2R, 3S, 4S, 5R)-5-(6-chloro-4-(cyclopentyl(methoxymethyl)amino)-1H-pyrazolo[3,4-d]pyrimidin-1-yl)-3,4-bis(methoxymethoxy)tetrahydro-2H-pyran-2-yl)methoxy)-1-methoxy-3-(2-(2-(2-(trimethylsilyl)ethoxy)methyl)-2H-tetrazol-5-yl)methoxy)propan-2-yl)phosphonate 42A (1.0 g, 1.05 mmol) in acetonitrile (3 mL) in an ice bath. The reaction mixture was stirred at room temperature overnight. After the phospholipids were completely hydrolyzed, the reaction mixture was concentrated under reduced pressure. Then trifluoroacetic acid / water (3 / 1, 2 mL) was added to the obtained crude product. The reaction solution was stirred at room temperature for 3 hours. After the reaction was completed, the reaction solution was concentrated under reduced pressure to obtain a residue, which was purified by reverse phase preparative chromatography (eluent: acetonitrile / 0.1% trifluoroacetic acid aqueous solution = 25%-90%, V / V) to obtain the title compound 42 (300 mg, yield: 45.1%). LC / MS (ESI) (m / z): 634.2 (M+H) + ; 1 H NMR (400 MHz, CD3OD) δ 8.01(s, 1H), 5.02 (s, 2H), 4.86 – 4.81 (m, 1H), 4.59 – 4.48 (m, 1H), 4.45 – 4.38(m, 1H), 4.23 – 4.12 (m, 2H), 4.10 – 4.00 (m, 5H), 3.97 – 3.89 (m, 1H), 3.87 – 3.73 (m, 2H), 3.36 (s, 3H), 2.17 – 2.03 (m, 2H), 1.86 – 1.54 (m, 6H).

[0201] Embodiment 43 (R)-1-(2H-tetrazolyl-5-yl)methoxy)-2-((2R,3S,4S,5R)-5-(6-chloro-4-cyclopentylamino)-1H-pyrazolo[3,4-d]pyrimidin-1-yl)-3,4-dihydroxytetrahydro-2H-pyran-2-yl)methoxy)-3-methoxypropane-2-yl)phosphonic acid

[0202] Compound 43 was synthesized from starting material 39H-2 according to the synthetic procedure of Example 42. LC-MS m / z (ESI): 634.2 (M+H + ). 1 H NMR (400 MHz, MeOD) δ 8.03 (s, 1H), 5.00 (s, 2H), 4.86 – 4.82 (m, 1H), 4.61 – 4.48 (m, 1H), 4.46 – 4.38 (m, 1H), 4.26 – 3.97 (m, 7H), 3.95 – 3.75(m, 3H), 3.37 (s, 3H), 2.17 – 2.03 (m, 2H), 1.86 – 1.53 (m, 6H).

[0203] Embodiment 44 (S)-1-(2H-tetrazol-5-yl)methoxy)-2-((2R,3S,4S,5R)-5-(6-chloro-4-(piperazin-1-yl)-1H-pyrazolo[3,4-d]pyrimidin-1-yl)-3,4-dihydroxytetrahydro-2H-pyran-2-yl)methoxy)-3-methoxypropane-2-yl)phosphonic acid (44-1) (R)-1-(2H-tetrazol-5-yl)methoxy)-2-((2R,3S,4S,5R)-5-(6-chloro-4-(piperazin-1-yl)-1H-pyrazolo[3,4-d]pyrimidin-1-yl)-3,4-dihydroxytetrahydro-2H-pyran-2-yl)methoxy)-3-methoxypropane-2-yl)phosphonic acid (44-2)

[0204] Compounds 44-1 and 44-2 were synthesized from starting materials 40-1 and 40-2 according to the synthesis procedure of Example 42.

[0205] 44-1: 1 H NMR (400 MHz, D2O) δ 8.17 (d, J = 45.2 Hz, 1H), 4.93 (s, 2H), 4.90-4.80 (m, 1H), 4.42 (dd, J = 7.3, 3.3 Hz, 1H), 4.22 – 3.89 (m, 12H), 3.85-3.78 (m, 1H), 3.73-3.70 (m, 1H), 3.46 – 3.36 (m, 4H), 3.29 (s, 3H).

[0206] 44-2: 1 H NMR (400 MHz, D2O) δ 8.30 – 8.00 (m, 1H), 5.00 – 4.83 (m, 2H), 4.78 – 4.74 (m, 1H), 4.40-3.35 (m, 1H), 4.24 – 4.12 (m, 4H), 4.11 – 3.77 (m,9H), 3.70-3.64 (m, 1H), 3.52-3.45 (m, 4H), 3.30 (s, 3H).

[0207] Embodiment 45 (S)-1-(2H-tetrazol-5-yl)methoxy)-2-((2R,3S,4S,5R)-5-(6-chloro-4-(4-methylpiperazin-1-yl)-1H-pyrazolo[3,4-d]pyrimidin-1-yl)-3,4-dihydroxytetrahydro-2H-pyran-2-yl)methoxy)-3-methoxypropane-2-yl)phosphonic acid (45-1) (R)-1-(2H-tetrazol-5-yl)methoxy)-2-((2R,3S,4S,5R)-5-(6-chloro-4-(4-methylpiperazin-1-yl)-1H-pyrazolo[3,4-d]pyrimidin-1-yl)-3,4-dihydroxytetrahydro-2H-pyran-2-yl)methoxy)-3-methoxypropane-2-yl)phosphonic acid (45-2)

[0208] Compounds 19-1 and 19-2 were synthesized by referring to step 9 in Example 39 with starting material 19; compounds 45-1 and 45-2 were synthesized by referring to the synthetic procedure of Example 42 with starting materials 19-1 and 19-2.

[0209] 45-1: 1 H NMR (400 MHz, D2O) δ 8.11 (s, 1H), 4.93 (s, 2H), 4.83 – 4.78 (m,3H), 4.42 (dd, J = 7.2, 2.9 Hz, 1H), 4.11 – 3.88 (m, 8H), 3.85 – 3.78 (m, 1H), 3.75 – 3.53 (m, 5H), 3.35 – 3.17 (m, 5H), 2.93 (s, 3H).

[0210] 45-2: 1 H NMR (400 MHz, D2O) δ 8.06 (s, 1H), 5.00 – 4.86 (m, 2H), 4.78 –4.73 (m, 3H), 4.36 (dd, J = 6.5, 3.0 Hz, 1H), 4.11 – 3.79 (m, 9H), 3.75 – 3.51(m, 5H), 3.35 – 3.17 (m, 5H), 2.93 (s, 3H).

[0211] Embodiment 46 (2-(2R,3S,4S,5R)-5-(2-chloro-4-(cyclopentylamino)pyrrolo[2,1-f][1,2,4]triazin-7-yl)-3,4-dihydroxytetrahydro-2H-pyran-2-yl)methoxy)-1-hydroxy-3-methoxypropan-2-yl)phosphonic acid

[0212]

[0213] Step 1: Synthesis of compound 46B (1aS, 3aR, 7aR, 7bS)-6,6-dimethylhexahydroxy[2', 3': 4,5]pyrano[3,2-d][1,3]dioxin 46A (5.0 g, 26.9 mmol) was dissolved in 4% potassium hydroxide aqueous solution (150 mL), and the reaction solution was stirred at 100°C for 1 hour. After the reaction was completed, the reaction solution was diluted with saturated saline solution, dichloromethane / isopropanol (5 / 1, V / V ) was extracted, and the organic phase was dried over anhydrous sodium sulfate, filtered, and concentrated under reduced pressure to give the title compound 46B (5 g, colorless oil, yield: 91.2%).

[0214] Step 2: Synthesis of compound 46C (4aR, 7R, 8S, 8aS)-2,2-dimethylhexahydropyrano[3,2-d][1,3]dioxin-7,8-diol 46B (5.0 g, 24.5 mmol) was dissolved in a mixed solvent of acetic acid (50 mL) and water (50 mL), and the reaction solution was stirred at 70°C for 1 hour. After the reaction was completed, the reaction solution was concentrated under reduced pressure to obtain the title compound 46C (4 g, colorless oil), which was used directly in the next step.

[0215] Step 3: Synthesis of compound 46D (2R, 3S, 4S, 5R)-2-(Hydroxymethyl)tetrahydro-2H-pyran-3,4,5-triol 46C (3.5 g, 21.3 mmol) was dissolved in THF (80 mL), triethylamine (8.90 mL, 64.0 mmol) and tert-butyldiphenylsilyl chloride (6.45 g, 23.5 mmol) were added, and the reaction solution was stirred at room temperature overnight. After the reaction was completed, the reaction solution was diluted with water, extracted with ethyl acetate, and the organic phase was dried over anhydrous sodium sulfate, filtered, concentrated under reduced pressure, and purified by silica gel column chromatography (eluent: methanol / dichloromethane = 1 / 10, V / V) to obtain the title compound 46D (8 g, colorless oil, yield: 93.3%).

[0216] Step 4: Synthesis of Compound 46E (2R, 3S, 4S, 5R)-2-((tert-butyldiphenylsilyloxy)methyl)tetrahydro-2H-pyran-3,4,5-triol 46D (8.0 g, 19.9 mmol) was dissolved in dichloromethane (100 mL), and dimethoxypropane (8.28 g, 79.5 mmol) and 4-methylbenzenesulfonate pyridinium (2.50 g, 9.94 mmol) were added, and the reaction solution was stirred at room temperature overnight. After the reaction was completed, the reaction solution was diluted with water, extracted with dichloromethane (200 mL), and the organic phase was dried over anhydrous sodium sulfate, filtered, concentrated under reduced pressure, and purified by silica gel column chromatography (eluent: ethyl acetate / petroleum ether = 1 / 3, V / V) to obtain the title compound 46E (7.5 g, colorless oil, yield: 85.3%).

[0217] Step 5: Synthesis of Compound 46F (3aR, 4R, 7R, 7aS)-4-(((tert-butyldiphenylsilyl)oxy)methyl)-2,2-dimethyltetrahydro-4H-[1,3]dioxo[4,5-c]pyran-7-ol 46E (3.5 g, 7.91 mmol) was dissolved in dichloromethane (100 mL), and Dess-Martin periodinane (10.1 g, 23.7 mmol) was added under ice bath conditions, and the reaction solution was stirred at room temperature for 2 hours. After the reaction was completed, the reaction solution was filtered to remove insoluble matter, and washed with dichloromethane, and the filtrate was washed with saturated sodium bicarbonate and saturated brine in sequence. The organic phase was dried over anhydrous sodium sulfate, filtered, concentrated under reduced pressure, and purified by silica gel column chromatography (eluent: ethyl acetate / petroleum ether = 1 / 3, V / V) to obtain the title compound 46F (3.4 g, colorless oil, yield: 97.6%).

[0218] Step 6: Synthesis of Compound 46G Tert-butyl (7-bromo-2-chloropyrrolo[2,1-f][1,2,4]triazin-4-yl)(cyclopentyl)carbamate 46F (8.02 g, 19.3 mmol) was dissolved in dry tetrahydrofuran (150 mL). After adding n-butyl lithium (12.1 mL, 19.3 mmol, 1.6 M) at -78°C, the mixture was stirred for half an hour at this temperature. Then, a tetrahydrofuran solution of (3aR, 4R, 7aR)-4-(((tert-butyldiphenylsilyl)oxy)methyl)-2,2-dimethyldihydro-4H-[1,3]dioxo[4,5-c ]pyran-7(6H)-one (3.4 g, 7.72 mmol) was slowly added dropwise to the mixture, and stirring was continued at this temperature for 2 hours. After the reaction was completed, the reaction solution was quenched with saturated ammonium chloride aqueous solution, extracted with ethyl acetate, and the organic phase was dried over anhydrous sodium sulfate, filtered, and concentrated under reduced pressure to obtain a crude product. The crude product was purified by silica gel column chromatography (eluent: ethyl acetate / petroleum ether = 1 / 10, V / V) to obtain the title compound 46G (4.8 g, yellow oil, yield: 80%).

[0219] Step 7: Synthesis of compound 46H Tert-butyl (7-((3aR, 4R, 7S, 7aR)-4-((((tert-butyldiphenylsilyl)oxy)methyl)-7-hydroxy-2,2-dimethyltetrahydro-4H-[1,3]dioxo[4,5-c]pyran-7-yl)-2-chloropyrrolo[2,1-f][1,2,4]triazin-4-yl)(cyclopentyl)carbamate 46G (6.5 g, 8.36 mmol) was dissolved in dry tetrahydrofuran (40 mL). Sodium hydride NaH (3.34 g, 83.6 mmol, 60% in mineral oil) was added under ice bath condition. After stirring at this temperature for half an hour, carbon disulfide (5.0 mL, 83.6 mmol) was added and stirring was continued for half an hour. Finally, iodomethane (6.79 mL, 83.6 mmol) and then continued to stir for half an hour. After the reaction was completed, the reaction solution was quenched with saturated ammonium chloride aqueous solution, extracted with ethyl acetate, and the organic phase was dried over anhydrous sodium sulfate, filtered, concentrated under reduced pressure, and purified by silica gel column chromatography (eluent: ethyl acetate / petroleum ether=1 / 10, V / V) to obtain the title compound 46H (5 g, yellow oil, yield: 68.9%).

[0220] Step 8: Synthesis of compound 46I Tert-butyl (7-((3aR, 4R, 7S, 7aR)-4-((((tert-butyldiphenylsilyl)oxy)methyl)-2,2-dimethyl-7-(((methylthio)carbonylthio)oxy)tetrahydro-4H-[1,3]dioxo[4,5-c]pyran-7-yl)-2-chloropyrrolo[2,1-f][1,2,4]triazin-4-yl)(cyclopentyl)carbamate 46H (5.0 g, 5.76 mmol), AIBN (0.09 g, 0.576 mmol) and tributyltin hydride (3.10 mL, 11.5 mmol) were dissolved in toluene (100 mL). The reaction mixture was heated at 140 °C. o C. and stirred for 1 hour. After the reaction was completed, the reaction solution was quenched with an icy 10% potassium fluoride aqueous solution, extracted with ethyl acetate, and the organic phase was dried over anhydrous sodium sulfate, filtered, concentrated under reduced pressure, and purified by silica gel column chromatography (eluent: ethyl acetate / petroleum ether = 1 / 10, V / V) to obtain the title compound 46I (4 g, yellow oil, yield: 91.2%).

[0221] Step 9: Synthesis of compound 46J Tert-butyl (7-((3aS, 4R, 7R, 7aS)-4-(((tert-butyldiphenylsilyl)oxy)methyl)-2,2-dimethyltetrahydro-4H-[1,3]dioxo[4,5-c]pyran-7-yl)-2-chloropyrrolo[2,1-f][1,2,4]triazin-4-yl)(cyclopentyl)carbamate 46I (4 g, 5.25 mmol) was dissolved in tetrahydrofuran (30 mL), TBAF (10.5 mL, 10.5 mmol, 1 M THF solution) was added, and the reaction solution was stirred at room temperature overnight. After the reaction was completed, the reaction solution was diluted with saturated aqueous ammonium chloride solution, extracted with ethyl acetate, and the organic phase was dried over anhydrous sodium sulfate, filtered, concentrated under reduced pressure, and purified by silica gel column chromatography (eluent: ethyl acetate / petroleum ether = 1 / 2, V / V) to obtain the title compound 46J (1 g, yellow solid, yield: 36.4%).

[0222] Step 10: Synthesis of compound 46K Tert-butyl (2-chloro-7-((3aS, 4R, 7R, 7aS)-4-(hydroxymethyl)-2,2-dimethyltetrahydro-4H-[1,3]dioxo[4,5-c]pyran-7-yl)pyrrolo[2,1-f][1,2,4]triazin-4-yl)(cyclopentyl)carbamate 46J (1 g, 1.91 mmol), ethyl 2-diazo-2-(diethoxyphosphoryl)acetate (1.92 g, 7.65 mmol) and rhodium dimer acetate (80 mg, 0.191 mmol) were dissolved in toluene (20 mL). The reaction mixture was replaced with nitrogen three times and stirred at 100°C for 3 hours. After the reaction was completed, the reaction solution was diluted with water, extracted with ethyl acetate, and the organic phase was dried over anhydrous sodium sulfate, filtered, and concentrated under reduced pressure to obtain a crude product. The crude product was purified by silica gel column chromatography (eluent: ethyl acetate / petroleum ether = 1 / 0, V / V) to give the title compound 46K (1.4 g, yellow oil, yield: 98.3%).

[0223] Step 11: Synthesis of Compound 46L Diethyl {1-[(3aS, 4R, 4R, 7aS)-7-[2-chloro-4-(cyclohexyl-cyclohexyl-benzobenzo-2-methylpropan-2-yl-2-methylpropyl-2-yl-2-methylpropyl-2-yloxy]carbonylamino]pyrrolo[2,1-f][1,2,4,4-triazin-7-yl]-2,2-dimethyl-4,6,7,7-tetrahydro-3-cyclocyclohexyl-4-cyclopentenylbenzobenzotriyl}phosphonate 46K (700 mg, 0.939 mmol) was dissolved in tetrahydrofuran (10 mL), sodium bis(trimethylsilyl)amide (1.85 mL, 1.88 mmol, 1 M tetrahydrofuran solution) was added at -20°C, and the mixture was stirred for half an hour at this temperature. Tetrabutylammonium iodide (173 mg, 0.470 mmol) and chloromethyl methyl ether (0.285 mL, 3.76 mmol) were added sequentially at this temperature and stirred for half an hour. After the reaction was completed, the reaction solution was quenched with saturated aqueous ammonium chloride solution, extracted with ethyl acetate, and the organic phase was dried over anhydrous sodium sulfate, filtered, and concentrated under reduced pressure to obtain a crude product. The crude product was purified by silica gel column chromatography (eluent: ethyl acetate / petroleum ether = 1 / 0, V / V) to obtain the title compound 46L (280 mg, yellow solid, yield: 37.8%).

[0224] Step 12: Synthesis of compound 46M Diethyl [4-({[3aS, 44R, 7aS, 7S, 7R)-7-[2-chloro-4-(cyclopentyl-cyclopentyl-benzodimethylmethyl-2-methylprop-2-yl)benzoate 46L (280 mg, 0.355 mmol) was dissolved in ethanol (5 mL), and calcium chloride (236 mg, 2.13 mmol) and sodium borohydride (80.5 mg, 2.13 mmol) were added under ice bath conditions. The reaction solution was stirred at room temperature overnight. After the reaction was completed, the reaction solution was diluted with water, extracted with ethyl acetate, and the organic phase was dried over anhydrous sodium sulfate, filtered, and concentrated under reduced pressure to obtain a crude product. The crude product was purified by silica gel column chromatography (eluent: methanol / dichloromethane = 1 / 10, V / V) to obtain the title compound 46M (250 mg, yellow solid, yield: 94.3%).

[0225] Step 13: Synthesis of Compound 46 Dissolve diethyl {1-[(3aS, 4R, 4R, 7aS, 7aS, 7R)-7-[2-chloro-4-(cyclohexylcyclohexylbenzothiazole)-2-methylmethyl-2-methylpropene-2-ylbenzothiazole benzoate 46M (80 mg, 0.107 mmol) in acetonitrile (5 mL), add triethylamine (0.22 mL, 1.61 mmol) and trimethylsilyl bromide (164 mg, 1.07 mmol) under ice bath conditions. The reaction solution was stirred at room temperature overnight. After the phospholipid hydrolysis was completed, the reaction solution was concentrated under reduced pressure, and then trifluoroacetic acid (1.5 mL) and water (0.5 mL) were added and stirred at room temperature for 1 hour. The reaction solution was concentrated under reduced pressure and purified by reverse phase preparative chromatography (eluent: acetonitrile / 0.1% trifluoroacetic acid aqueous solution = 25%-90%, V / V) to obtain the title compound 46 (12.9 mg, white solid, yield: 21.9%).

[0226] 1 H NMR (400 MHz, MeOD) δ 6.93 – 6.83 (m, 1H), 6.80 – 6.51 (m, 1H), 4.61 – 4.50 (m, 1H), 4.35 – 4.23 (m, 1H), 4.15 – 3.94 (m, 5H), 3.91 – 3.74(m, 5H), 3.67 – 3.60 (m, 1H), 3.45 – 3.36 (m, 3H), 2.14 – 2.02 (m, 2H), 1.85 – 1.75 (m, 2H), 1.71 – 1.58 (m, 4H).

[0227] Embodiment 47 1-(2H-tetrazol-5-yl)methoxy)-2-((2R,3S,4S,5R)-5-(2-chloro-4-cyclopentylamino)pyrrolo[2,1-f][1,2,4]triazin-7-yl)-3,4-dihydroxytetrahydro-2H-pyran-2-yl)methoxy)-3-methoxypropane-2-yl)phosphonic acid

[0228]

[0229] Compound 47 can be prepared by using compound 46M as a raw material and referring to the synthesis of compound 21. 1 H NMR (400MHz, CD3OD) δ 6.90 – 6.80 (m, 1H), 6.74 – 6.51 (m, 1H), 5.08 – 4.97 (m, 2H), 4.60 – 4.47 (m, 1H), 4.29 (s, 1H), 4.21 – 3.93 (m, 6H), 3.87 – 3.75 (m, 4H), 3.69 – 3.55 (m, 1H), 3.45 – 3.34 (m, 3H), 2.15 – 2.00 (m, 2H), 1.84 – 1.74(m, 2H), 1.70 – 1.56 (m, 4H).

[0230] Embodiment 48 (2-(2R,3S,4S,5S,6S)-5-(6-chloro-4-cyclopentylamino)-1H-pyrazolo[3,4-d]pyrimidin-1-yl)-3,4-dihydroxy-6-methyltetrahydro-2H-pyran-2-yl)methoxy)-1-hydroxy-3-methoxypropane-2-yl)phosphonic acid

[0231]

[0232] Step 1: Synthesis of compound 48B (2S, 3R, 6S) -6-cyano-2-methyl-3,6-dihydro-2H-pyran-3-yl acetate 48A (12 g, 66.2 mmol) was dissolved in HCl / MeOH (40 mL, 2 M) solution. The reaction solution was stirred at 70°C for 4 hours. After the reaction was completed, the reaction solution was filtered, the filtrate was concentrated under reduced pressure, and the crude product was purified by silica gel column chromatography (eluent: ethyl acetate / petroleum ether = 1 / 3, V / V) to obtain the title compound (2S, 5R, 6S) -5-hydroxy-6-methyl-5,6-dihydropyran-2-carboxylic acid methyl ester 48B (6 g, yield: 52.6%).

[0233] Step 2: Synthesis of compound 48C (2S, 5R, 6S)-5-hydroxy-6-methyl-5,6-dihydropyran-2-carboxylic acid methyl ester 48B (6 g, 34.8 mmol) was dissolved in dry THF (60 mL). LiAlH4 (20.9 mL, 52.3 mmol, 2.5 M tetrahydrofuran solution) was slowly added dropwise under ice bath. The reaction solution was stirred at 0°C for 1 h. After the reaction was completed, sodium sulfate decahydrate was carefully added to quench the reaction. The solution was filtered, the filtrate was concentrated under reduced pressure, and purified by silica gel column chromatography (eluent: methanol / dichloromethane = 1 / 10, V / V) to obtain the title compound 6-hydroxymethyl-2-methyl-3,6-dihydropyran-3-ol 48C (3.8 g, yield: 75.6%).

[0234] Step 3: Synthesis of compound 48D ((2S, 3R, 6S)-6-(Hydroxymethyl)-2-methyl-3,6-dihydro-2H-pyran-3-ol 48C (3.8 g, 26.3 mmol) and imidazole (3.59 g, 52.7 mmol) were dissolved in dry tetrahydrofuran (40 mL), and TBDPSCl (7.97 g, 29.0 mmol) was added under ice bath. The reaction mixture was stirred at room temperature overnight. After the reaction was completed, water was added to quench the reaction, and the reaction solution was extracted with ethyl acetate and washed with saturated saline solution. The obtained organic phase was dried over anhydrous sodium sulfate, filtered, and concentrated under reduced pressure. The crude product was purified by silica gel column chromatography (eluent: ethyl acetate / petroleum ether = 1 / 3, V / V) to obtain the title compound 48D (7.5 g, yield: 74.4%).

[0235] Step 4: Synthesis of Compound 48E (2S,3R,6S)-6-(((tert-butyldiphenylsilyl)oxy)methyl)-2-methyl-3,6-dihydro-2H-pyran-3-ol 48D (7.5 g, 19.6 mmol), 6-chloro-N-cyclopentyl-1H-pyrazolo[3,4-d ]pyrimidin-4-amine (2.33 g, 9.80 mmol) and triphenylphosphine (7.71 g, 29.4 mmol) were added to dry tetrahydrofuran (100 mL). DIAD (5.95 g, 29.4 mmol) was slowly added dropwise under N2 in an ice bath. After the addition was complete, the reaction solution was stirred at 70°C for 12 h. After the reaction was completed, water was added to the reaction mixture, extracted with ethyl acetate (200 mL), and the obtained organic phase was dried over anhydrous sodium sulfate, filtered, concentrated under reduced pressure, and purified by silica gel column chromatography (eluent: ethyl acetate / petroleum ether = 1 / 3, V / V) to obtain the title compound 48E (6 g, 101%). LC / MS (ESI) m / z: 602 [M+H] + .

[0236] Step 5: Synthesis of Compound 48F 1-(2S,3S,6S)-6-((tert-butyldiphenylsilyloxy)methyl)-2-methyl-3,6-dihydro-2H-pyran-3-yl)-6-chloro-N-cyclopentyl-1H-pyrazolo[3,4-d]pyrimidin-4-amine 48E (6 g, 9.96 mmol) was added to a mixed solution of tetrahydrofuran (50 mL) and water (5 mL), followed by the addition of NMO (2.33 g, 19.9 mmol) and K2OsO4·2H2O (0.04 g, 0.10 mmol). The reaction mixture was stirred at room temperature for 48 hours. After the reaction was completed, Na2S2O3 solution was added to quench the reaction. Water was then added to the reaction mixture, and the mixture was extracted with ethyl acetate. The obtained organic phase was dried over anhydrous sodium sulfate, filtered, concentrated under reduced pressure, and purified by silica gel column chromatography (eluent: ethyl acetate / petroleum ether = 1 / 2, V / V) to obtain the title compound 48F (2R, 3S, 4S, 5S, 6S)-2-(tert-butyldiphenylsilyloxy)methyl)-5-(6-chloro-4-cyclopentylamino)-1H-pyrazolo[3,4-d]pyrimidin-1-yl)-6-methyltetrahydro-2H-pyran-3,4-diol (2.3 g, yield: 36.3%). LC / MS (ESI) m / z: 636 [M+H] + .

[0237] Step 6: Synthesis of Compound 48G (2R, 3S, 4S, 5S, 6S)-2-(tert-butyldiphenylsilyloxy)methyl)-5-(6-chloro-4-cyclopentylamino)-1H-pyrazolo[3,4-d]pyrimidin-1-yl)-6-methyltetrahydro-2H-pyran-3,4-diol 48F (2 g, 3.14 mmol) was dissolved in anhydrous dichloromethane (20 mL), and 2,2-dimethoxypropane (1.31 g, 12.6 mmol) and PPTS (0.79 g, 3.14 mmol) were added under ice bath. The reaction solution was stirred at room temperature for 24 hours. After the reaction was completed, it was extracted with dichloromethane (200 mL), and the obtained organic phase was dried over anhydrous sodium sulfate, filtered, and concentrated under reduced pressure. The crude product was purified by silica gel column chromatography (eluent: ethyl acetate / petroleum ether = 1 / 5, V / V) to give the title compound 1-((3aS, 4R, 6S, 7R, 7aS)-4-(((tert-butyldiphenylsilyl)oxy)methyl)-2,2,6-trimethyltetrahydro-4H-[1,3]dioxo[4,5-c]pyran-7-yl)-6-chloro-N-cyclopentyl-1H-pyrazolo[3,4-d ]pyrimidin-4-amine 48G (1 g, yield: 47.0%). LC / MS (ESI) m / z: 676[M+H] + .

[0238] Step 7: Synthesis of compound 48H 1-((3aS,4R,6S,7R,7aS)-4-(((tert-butyldiphenylsilyl)oxy)methyl)-2,2,6-trimethyltetrahydro-4H-[1,3]dioxo[4,5-c]pyran-7-yl)-6-chloro-N-cyclopentyl-1H-pyrazolo[3,4-d]pyrimidin-4-amine 48G (1.00 g, 1.48 mmol) was dissolved in a solution of TBAF in tetrahydrofuran (2.22 mL, 2.22 mmol, 1 M). The reaction mixture was stirred at 60°C for 2 h. After the reaction was completed, the reaction solution was concentrated under reduced pressure, and the crude product was purified by silica gel column chromatography (eluent: ethyl acetate / petroleum ether = 1 / 2, V / V) to obtain the title compound (3aS, 4R, 6S, 7R, 7aS)-7-(6-chloro-4-(cyclopentylamino)-1H-pyrazolo[3,4-d]pyrimidin-1-yl)-2,2,6-trimethyltetrahydro-4H-[1,3]dioxo[4,5-c]pyran-4-yl)methanol 48H (500 mg, yield: 77.2%). LC / MS (ESI) m / z: 438 [M+H] + .

[0239] Step 8: Synthesis of compound 48I Under N2 atmosphere, dimeric rhodium (II) acetate (25.2 mg, 0.06 mmol) and ethyl 2-diazo-2-(diethoxyphosphoryl)acetate (860 mg, 3.43 mmol) were added to a toluene solution (10 mL) of (3aS, 4R, 6S, 7R, 7aS)-7-(6-chloro-4-(cyclopentylamino)-1H-pyrazolo[3,4-d]pyrimidin-1-yl)-2,2,6-trimethyltetrahydro-4H-[1,3]dioxo[4,5-c ]pyran-4-yl)methanol 48H (500 mg, 1.14 mmol). After nitrogen replacement, the reaction mixture was stirred at 100°C for 3 hours. After the reaction was completed, the reaction solution was poured into water and extracted with ethyl acetate. The obtained organic phase was dried over anhydrous sodium sulfate, filtered and concentrated. The crude product was purified by silica gel column chromatography (eluent: ethyl acetate / petroleum ether = 3 / 1, V / V) to give the title compound ethyl 2-(3aS,4R,6S,7R,7aS)-7-(6-chloro-4-cyclopentylamino)-1H-pyrazolo[3,4-d]pyrimidin-1-yl)-2,2,6-trimethyltetrahydro-4H-[1,3]dioxolane[4,5-c]pyran-4-methoxy)-2-(diethoxyphospho)acetate 48I (470 mg, 0.71 mmol, yield: 62.4%). LC / MS (ESI) (m / z): 660 (M+H)+.

[0240] Step 9: Synthesis of compound 48J (Ethyl 2-(3aS,4R,6S,7R,7aS)-7-(6-chloro-4-cyclopentylamino)-1H-pyrazolo[3,4-d]pyrimidin-1-yl)-2,2,6-trimethyltetrahydro-4H-[1,3]dioxolane[4,5-c]pyran-4-methoxy)-2-(diethoxyphospho)acetate 48I (470 mg, 0.71 mmol) was dissolved in dry tetrahydrofuran (10 mL). Sodium bis(trimethylsilyl)amide (2.42 mL, 2.42 mmol, 1M tetrahydrofuran solution) was added dropwise under N2 atmosphere at -20°C, and the reaction solution was stirred at -20°C for 30 minutes. Tetrabutylammonium iodide (112 mg, 0.30 mmol) was added to the mixture, and then chloro(methoxy)methane (0.37 mL, 4.85 mmol) was added dropwise at -20°C. The reaction solution was stirred at -20°C for 1 hour. The reaction solution was quenched with saturated NH4Cl solution and extracted with ethyl acetate. The obtained organic phase was dried over anhydrous sodium sulfate, filtered, and concentrated. The crude product was purified by silica gel column chromatography (eluent: methanol / dichloromethane = 1 / 10, V / V) to give the title compound ethyl 2-(3aS,4R,6S,7R,7aS)-7-(6-chloro-4-cyclopentylmethoxymethylamino)-1H-pyrazolo[3,4-d]pyranimidazole-1-yl)-2,2,6-trimethyltetrahydro-4H-[1,3]dioxacyclo[4,5-c]pyran-4-yl)methoxy)-2-(diethoxyphospho)-3-methoxypropanoate 48J (280 mg, 0.37 mmol, yield: 52.6%). LC / MS (ESI) (m / z): 748 (M+H)+.

[0241] Step 10: Synthesis of compound 48K Ethyl 2-(3aS,4R,6S,7R,7aS)-7-(6-chloro-4-cyclopentylmethoxymethylamino)-1H-pyrazolo[3,4-d]pyranimidazole-1-yl)-2,2,6-trimethyltetrahydro-4H-[1,3]dioxacyclo[4,5-c]pyran-4-yl)methoxy)-2-(diethoxyphospho)-3-methoxypropanoate 48J (280 mg, 0.37 mmol) was dissolved in ethanol (10 mL), and calcium chloride (124 mg, 1.12 mmol) and sodium borohydride (42.5 mg, 1.12 mmol) were added at 0°C. The reaction solution was stirred at room temperature for 1 hour, quenched with saturated NH4Cl solution, and extracted with ethyl acetate. The obtained organic phase was dried over anhydrous sodium sulfate, filtered, and concentrated. The crude product was purified by silica gel column chromatography (eluent: methanol / dichloromethane = 1 / 10, V / V) to give the title compound diethyl ester 2-((3aS, 4R, 6S, 7R, 7aS)-7-(6-chloro-4-cyclopentylmethoxymethylamino)-1H-pyrazolo[3,4-d]pyrimidin-1-yl)-2,2,6-trimethyltetrahydro-4H-[1,3]dioxacyclo[4,5-c]pyran-4-yl)methoxy)-1-hydroxy-3-methoxypropane-2-yl)phosphonate 48K (240 mg, 0.34 mmol, yield: 90.8%). LC / MS (ESI) (m / z): 706 (M +H)+.

[0242] Step 11: Synthesis of compound 48 Diethyl 2-((3aS,4R,6S,7R,7aS)-7-(6-chloro-4-cyclopentylmethoxymethylamino)-1H-pyrazolo[3,4-d]pyrimidin-1-yl)-2,2,6-trimethyltetrahydro-4H-[1,3]dioxacyclo[4,5-c]pyran-4-yl)methoxy)-1-hydroxy-3-methoxypropane-2-yl)phosphonate 48K (240 mg, 0.34 mmol) was dissolved in acetonitrile, and trimethylsilyl bromide (0.15 mL, 1.13 mmol) and triethylamine (0.22 mL, 1.59 mmol) were added at 0°C. The reaction mixture was stirred at room temperature for 12 hours. After the phospholipid hydrolysis was completed, the reaction solution was concentrated under reduced pressure. Then water (0.5 mL) and trifluoroacetic acid (2 mL) were added to the mixture and stirring was continued for 1 hour. After the reaction was completed, the mixture was concentrated under reduced pressure. The crude product was purified by HPLC to obtain the title compound (2-((2R, 3 S, 4 S, 5 S, 6 S)-5-(6-chloro-4-(cyclopentylamino)-1H-pyrazolo[3,4-d]pyrimidin-1-yl)-3,4-dihydroxy-6-methyltetrahydro-2H-pyran-2-yl)methoxy)-1-hydroxy-3-methoxypropane-2-yl)phosphonic acid 48 (37 mg, 0.07 mmol, yield: 57.7%).

[0243] 1 H NMR (400 MHz, CD3OD) δ 8.10 (s, 1H), 4.62 – 4.52 (m, 2H), 4.43 –4.37 (m, 1H), 4.19 – 3.83 (m, 9H), 3.42 (d, J = 8.8 Hz, 3H), 2.17 – 2.06 (m,2H), 1.85 -1.75 (m, 2H), 1.74 – 1.57 (m, 4H), 0.85 (d, J = 5.1 Hz, 3H).

[0244] Embodiment 49 1-(2H-tetrazol-5-yl)methoxy)-2-((2R,3S,4S,5S,6S)-5-(6-chloro-4-cyclopentylamino)-1H-pyrazolo[3,4-d]pyrimidin-1-yl)-3,4-dihydroxy-6-methyltetrahydro-2H-pyran-2-yl)methoxy)-3-methoxypropan-2-yl)phosphonic acid

[0245]

[0246] Compound 49 can be prepared by using compound 48K as a raw material and referring to the synthesis of compound 21. 1 H NMR (400MHz, CD3OD) δ 8.03 (d, J = 21.5 Hz, 1H), 5.18 – 5.01 (m, 2H), 4.83 – 4.75 (m,1H), 4.54 – 4.49 (m, 2H), 4.38 – 4.26 (m, 1H), 4.19 – 4.01 (m, 4H), 4.01 –3.98 (m, 1H), 3.94 – 3.76 (m, 3H), 3.39 (d, J = 5.5 Hz, 3H), 2.15 – 2.00 (m,2H), 1.91 – 1.50 (m, 6H), 0.85 – 0.73 (m, 6H).

[0247] Embodiment 50 (1-(2R,3S,4S,5R)-5-(6-chloro-4-cyclopentylamino)-1H-pyrazolo[3,4-d]pyrimidin-1-yl)-3,4-dihydroxytetrahydro-2H-pyran-2-ylmethoxy)-2-hydroxyethyl)phosphonic acid

[0248]

[0249] Step 1: Synthesis of compound 50A 2-(((2R,3S,4S,5R)-5-(6-chloro-4-(cyclopentylamino)-1H-pyrazolo[3,4-d]pyrimidin-1-yl)-3,4-bis(methoxymethoxy)tetrahydro-2H-pyran-2-yl)methoxy]-2-(diethoxyphosphoryl)acetic acid ethyl ester 39F (250 mg, 0.36 mmol) was dissolved in ethanol (5 mL) and calcium chloride (120 mg, 1.08 mmol) and sodium borohydride (36.5 mg, 1.09 mmol) were added at 0°C, and the mixture was stirred at room temperature overnight. The reaction solution was quenched with saturated aqueous ammonium chloride solution and extracted three times with ethyl acetate. The combined organic layer was dried over anhydrous sodium sulfate, filtered, and concentrated to dryness under reduced pressure. The obtained residue was purified by silica gel column chromatography (eluent: methanol / dichloromethane = 1 / 10, V / V) to give the title compound 50A (150 mg, yellow oil, yield: 64%).

[0250] Step 2: Synthesis of compound 50 Diethyl (1-(((2R,3S,4S,5R)-5-(6-chloro-4-(cyclopentylamino)-1H-pyrazolo[3,4-d]pyrimidin-1-yl)-3,4-bis(methoxymethoxy)tetrahydro-2H-pyran-2-yl)methoxy-2-hydroxyethyl)phosphonate 50A (150 mg, 0.23 mmol) was dissolved in acetonitrile (5 mL) and triethylamine (0.5 mL, 3.45 mmol) was added, followed by trimethylsilyl bromide (0.3 mL, 2.30 mmol). The solution was stirred overnight and then concentrated under reduced pressure. The resulting residue was dissolved in a mixed solution of water and trifluoroacetic acid (1 / 1, 4 mL), and the reaction mixture was stirred at room temperature for 1 hour. The mixture was concentrated and purified by preparative high performance liquid chromatography (20-50% acetonitrile / water, 0.1% formic acid, gradient elution) to give the title compound 50 (36 mg, yield: 31%). 1 H NMR (400 MHz, MeOD) δ 8.07 (s, 1H), 4.85 – 4.81 (m, 1H), 4.60 – 4.47(m, 1H), 4.39 – 4.35 (m, 1H), 4.23 – 4.10 (m, 4H), 4.01 – 3.68 (m, 5H), 2.15– 2.06 (m, 2H), 1.87 – 1.52 (m, 6H).

[0251] Embodiment 51 1-((2R,3S,4R,5R)-5-(6-chloro-4-cyclopentylamino)-1H-pyrazolo[3,4-d]pyrimidin-1-yl)-3,4-dihydroxytetrahydro-2H-pyran-2-yl)methoxy)-2-hydroxyethyl)phosphonic acid

[0252]

[0253] Step 1: Synthesis of compound 51B Under ice bath conditions. Slowly add 48% hydrobromic acid acetic acid solution (70 mL) to a dichloromethane solution (500 mL) of (2R, 3R, 4S, 5R, 6S)-4,5,6-triacetoxy-2-(acetoxymethyl)tetrahydropyran-3-yl acetate 51A (50 g, 128 mmol), and stir the reaction solution at room temperature overnight. After the reaction is completed, the reaction solution is quenched with an ice-saturated sodium bicarbonate aqueous solution, extracted with dichloromethane, and the organic phase is dried over anhydrous sodium sulfate, filtered, and concentrated under reduced pressure to obtain a crude product. The crude product is slurried and filtered in ethanol to obtain the title compound 51B (38 g, white solid, yield: 72.1%).

[0254] Step 2: Synthesis of compound 51C (2R, 3R, 4S, 5R, 6S)-4,5-diacetoxy-2-(acetoxymethyl)-6-bromotetrahydropyran-3-yl acetate 51B (38 g, 92.4 mmol), azobisisobutyronitrile (0.30 g, 1.85 mmol) and tributyltin hydride (24.9 mL, 92.4 mmol) were dissolved in toluene (500 mL). The reaction solution was stirred at 65°C for 1 hour. After the reaction was completed, the reaction solution was quenched with a cold 10% potassium fluoride aqueous solution, extracted with ethyl acetate, and the organic phase was dried over anhydrous sodium sulfate, filtered, and concentrated under reduced pressure to obtain a crude product. The crude product was purified by silica gel column chromatography (eluent: ethyl acetate / petroleum ether = 1 / 1, V / V) to obtain the title compound 51C (28.5 g, white solid, yield: 92.8%).

[0255] Step 3: Synthesis of Compound 51D (2R, 3R, 4R, 5S)-4,5-diacetoxy-2-(acetoxymethyl)tetrahydropyran-3-yl acetate 51C (28.5 g, 85.8 mmol) and sodium methoxide (1.59 mL, 8.58 mmol, 5.4 M methanol solution) were dissolved in methanol (200 mL). The reaction solution was stirred at room temperature overnight. After the reaction was completed, the reaction solution was concentrated under reduced pressure to obtain a crude product, which was dissolved in N,N-dimethylformamide (200 mL), and dimethoxyphenylmethane (19.4 mL, 128 mmol) and 4-methylbenzenesulfonic acid (1.54 g, 25.7 mmol) were added. The reaction solution was stirred at 70°C for 1 hour. After the reaction was completed, the reaction solution was diluted with saturated brine, extracted with ethyl acetate, and the organic phase was dried over anhydrous sodium sulfate, filtered, and concentrated under reduced pressure to obtain a crude product. The crude product was purified by silica gel column chromatography (eluent: methanol / dichloromethane = 1 / 10, V / V) to give the title compound 51D (15 g, white solid, yield: 69.3%).

[0256] Step 4: Synthesis of Compound 51E (4aR, 7S, 8aS, 8R)-2-phenyl-4,4a,6,7,8,8a-hexahydropyran[3,2-d][1,3]dioxy-7,8-diol 51D (15 g, 59.5 mmol) and tert-butyldimethylsilyl chloride (13.4 g, 89.2 mmol) were dissolved in tetrahydrofuran (500 mL), and imidazole (6.07 g, 89.2 mmol) was added. The reaction solution was stirred at room temperature overnight. After the reaction was completed, the reaction solution was quenched with ice water, extracted with ethyl acetate, and the organic phase was dried over anhydrous sodium sulfate, filtered, and concentrated under reduced pressure to obtain a crude product. The crude product was purified by silica gel column chromatography (eluent: ethyl acetate / petroleum ether = 1 / 5, V / V) to obtain the title compound 51E (7.3 g, colorless oil, yield: 33.5%).

[0257] Step 5: Synthesis of Compound 51F (4aR, 7S, 8aR, 8R)-8-{[dimethyl(2-methylprop-2-yl)silanyl]oxy}-2-phenyl-4,4a,6,7,8,8a-hexahydropyrano[3,2d][1,3]dioxa-7-ol 51E (7.3 g, 19.9 mmol), 6-chloro-4-(cyclopentylamino)-1H-pyrazolo[3,4-d]pyrimidine (5.21 g, 21.9 mmol) and triphenylphosphine (15.7 g, 59.8 mmol) were dissolved in anhydrous tetrahydrofuran (200 mL). Diisopropyl azodicarboxylate (11.8 mL, 59.8 mmol) was added under ice bath conditions. The reaction solution was stirred at room temperature overnight. After the reaction was completed, it was concentrated under reduced pressure to obtain a crude product. The crude product was purified by silica gel column chromatography (eluent: ethyl acetate / petroleum ether = 1 / 1, V / V) to obtain the title compound 51F (11 g, colorless oil, yield: 94.2%).

[0258] Step 6: Synthesis of Compound 51G 6-Chloro-4-(cyclopentylamino)-1-[(4aR,8aR,8R)-8-{[dimethyl(2-methylprop-2-yl)silyl]oxy}-2-phenyl-4,4a,6,7,8,8a-hexahydropyrano[3,2-d][1,3]dioxa-7-yl]pyrazolo[3,4-d]pyrimidine 51F (4 g, 6.82 mmol) was dissolved in tetrahydrofuran (100 mL), and tetrabutylammonium fluoride (13.6 mL, 13.6 mmol, 1 M tetrahydrofuran solution) was added. The reaction solution was stirred at 60°C for 1 hour. After the reaction was completed, the reaction solution was diluted with saturated aqueous ammonium chloride solution, extracted with ethyl acetate, and the organic phase was dried over anhydrous sodium sulfate, filtered, and concentrated under reduced pressure to obtain a crude product. The crude product was purified by silica gel column chromatography (eluent: ethyl acetate / petroleum ether = 1 / 1, V / V) to obtain the title compound 51G (1.5 g, white solid, yield: 46.6%).

[0259] Step 7: Synthesis of Compound 51H (4aR, 8aS, 8R)-7-[6-chloro-4-(cyclopentylamino)pyrazolo[3,4-d]pyrimidin-1-yl]-2-phenyl-4,4a,6,7,8,8a-hexahydropyrano[3,2d][1,3]dioxa-8-ol 51G (1.5 g, 3.18 mmol) was dissolved in acetic acid / water (15 mL / 15 mL), and the reaction solution was stirred at 70°C for 1 hour. After the reaction was completed, the mixture was concentrated under reduced pressure to obtain a crude product. The crude product was purified by silica gel column chromatography (eluent: methanol / dichloromethane = 1 / 10, V / V) to obtain the title compound 51H (1 g, white solid, yield: 82.0%).

[0260] Step 8: Synthesis of Compound 51I (2R, 3S, 4R)-5-[6-chloro-4-(cyclopentylamino)pyrazolo[3,4-d]pyrimidin-1-yl]-2-(hydroxymethyl)tetrahydropyran-3,4-diol 51H (1.0 g, 2.61 mmol) and tert-butyldimethylsilyl chloride (0.59 g, 3.91 mmol) were dissolved in tetrahydrofuran (30 mL), and imidazole (0.35 g, 5.21 mmol) was added. The reaction solution was stirred at room temperature for 2 hours. After the reaction was completed, the reaction solution was diluted with water, extracted with ethyl acetate, and the organic phase was dried over anhydrous sodium sulfate, filtered, and concentrated under reduced pressure to obtain a crude product. The crude product was purified by silica gel column chromatography (eluent: ethyl acetate / petroleum ether = 1 / 1, V / V) to obtain the title compound 51I (1.15 g, white solid, yield: 88.6%).

[0261] Step 9: Synthesis of Compound 51J (2R, 3S, 4R)-5-[6-chloro-4-(cyclopentylamino)pyrazolo[3,4-d]pyrimidin-1-yl]-2-(3,3,4,4-tetramethyl-2-oxo-3-silapentyl-1-yl)tetrahydropyran-3,4-diol 51I (400 mg, 0.803 mmol) was dissolved in dichloromethane (10 mL), and triethylamine (1.00 mL, 7.23 mmol), acetic anhydride (491 mg, 4.82 mmol) and 4-dimethylaminopyridine (9.81 mg, 0.080 mmol) were added, and the reaction solution was stirred at room temperature for 1 hour. After the reaction was completed, the reaction solution was diluted with water, extracted with dichloromethane, and the organic phase was dried over anhydrous sodium sulfate, filtered, and concentrated under reduced pressure to obtain a crude product. The crude product was purified by silica gel column chromatography (eluent: ethyl acetate / petroleum ether = 1 / 1, V / V) to obtain the title compound 51J (460 mg, white solid, yield: 98.4%).

[0262] Step 10: Synthesis of compound 51K (2R, 3S, 4R)-4-acetoxy-5-[6-chloro-4-(cyclopentylamino)pyrazolo[3,4-d]pyrimidin-1-yl]-2-(3,3,4,4-tetramethyl-2-oxo-3-silanpent-1-yl)tetrahydropyran-3-yl acetate 51J (460 mg, 0.790 mmol) was dissolved in tetrahydrofuran (10 mL), and tetrabutylammonium fluoride (3.16 mL, 3.16 mmol, 1 M tetrahydrofuran solution) was added. The reaction solution was stirred under ice bath for 2 hours. After the reaction was completed, the reaction solution was diluted with saturated ammonium chloride aqueous solution, extracted with ethyl acetate, and the organic phase was dried over anhydrous sodium sulfate, filtered, and concentrated under reduced pressure to obtain a crude product. The crude product was purified by silica gel column chromatography (eluent: ethyl acetate / petroleum ether = 1 / 0, V / V) to give the title compound 51K (310 mg, white solid, yield: 82.1%).

[0263] Step 11: Synthesis of Compound 51L (2R, 3S, 4R)-4-acetoxy-5-[6-chloro-4-(cyclopentylamino)pyrazolo[3,4-d]pyrimidin-1-yl]-2-(hydroxymethyl)tetrahydropyran-3-yl acetate 51K (310 mg, 0.663 mmol), 1-(diethoxyphosphoryl)-2-ethoxy-2-oxoethane-1-diazo (665 mg, 2.65 mmol) and dimeric acetate (29.3 mg, 0.066 mmol) were dissolved in toluene (10 mL). After nitrogen substitution, the reaction solution was stirred at 100 ° C for 3 hours. After the reaction was completed, the reaction solution was diluted with water, extracted with ethyl acetate, and the organic phase was dried over anhydrous sodium sulfate, filtered, and concentrated under reduced pressure to obtain a crude product. The crude product was purified by silica gel column chromatography (eluent: ethyl acetate / petroleum ether = 1 / 0, V / V) to obtain the title compound 51L (190 mg, yellow oil, yield: 41.6%).

[0264] Step 12: Synthesis of Compound 51M {1-[(2R, 3S, 4R, 5R)-3,4-diacetoxy-5-[6-chloro-4-(cyclopentylamino)pyrazolo[3,4-d]pyrimidin-1-yl]-3,4,5,6-tetrahydro-2H-pyran-2-yl]-4-oxo-2,5-dioxahept-3-yl}phosphonic acid diethyl ester 51L (150 mg, 0.217 mmol) was dissolved in ethanol (5 mL), and calcium chloride (145 mg, 1.30 mmol) and sodium borohydride (44.1 mg, 1.30 mmol) were added under ice bath conditions. The reaction solution was stirred at room temperature overnight. After the reaction was completed, the reaction solution was diluted with water, extracted with ethyl acetate, and the organic phase was dried over anhydrous sodium sulfate, filtered, and concentrated under reduced pressure to obtain a crude product. The crude product was redissolved in methanol (5 mL), potassium carbonate (15.0 mg, 0.108 mmol) was added, and the mixture was stirred at room temperature for 2 hours. After the reaction was completed, the mixture was concentrated under reduced pressure to obtain a crude product, which was purified by silica gel column chromatography (eluent: methanol / dichloromethane = 1 / 10, V / V) to obtain the title compound 51M (40 mg, white solid, yield: 32.6%).

[0265] Step 13: Synthesis of compound 51 Diethyl [2-hydroxy-1-({[(2R,3S,4R,5R)-5-[6-chloro-4-(cyclopentylamino)pyrazolo[3,4-d]pyrimidin-1-yl]-3,4-dihydroxy-3,4,5,6-tetrahydro-2H-pyran-2-yl]methyl}oxy)ethyl]phosphonate 51M (40 mg, 0.071 mmol) was dissolved in acetonitrile (5 mL), and triethylamine (0.296 mL, 2.13 mmol) and trimethylsilyl bromide (217 mg, 1.42 mmol) were added under ice-cooling conditions. The reaction solution was stirred at room temperature overnight. After the reaction was completed, the reaction solution was concentrated under reduced pressure and purified by reverse phase preparation (eluent: acetonitrile / 0.1% trifluoroacetic acid aqueous solution = 25% to 90%, V / V) to obtain the title compound 51 (10.5 mg, yellow solid, yield: 29.2%).

[0266] 1 H NMR (400 MHz, CD3OD) δ 8.10 (s, 1H), 5.19 – 5.10 (m, 1H), 4.59 – 4.46 (m, 2H), 4.29 – 4.19 (m, 1H), 4.10 – 3.98 (m, 3H), 3.97 – 3.86 (m, 3H), 3.80 – 3.69 (m, 1H), 3.43 – 3.35 (m, 1H), 2.16 – 2.04 (m, 2H), 1.81 – 1.74 (m, 2H), 1.73 – 1.57 (m, 4H).

[0267] Embodiment 52 (2-(2R,3S,4R,5R)-5-(6-chloro-4-cyclopentylamino)-1H-pyrazolo[3,4-d]pyrimidin-1-yl)-3,4-dihydroxytetrahydro-2H-pyran-2-yl)methoxy)-1-hydroxy-3-methoxypropan-2-yl)phosphonic acid

[0268]

[0269] Step 1: Synthesis of compound 52A (2R, 3S, 4R)-5-[6-chloro-4-(cyclopentylamino)pyrazolo[3,4-d]pyrimidin-1-yl]-2-(3,3,4,4-tetramethyl-2-oxo-3-silanpent-1-yl)tetrahydropyran-3,4-diol 51I (1.5 g, 3.01 mmol) was dissolved in 1,4-dioxane (30 mL), and N,N-diisopropylethylamine (19.9 mL, 120 mmol) and chloromethyl methyl ether (4.58 mL, 60.2 mmol) were added. The reaction solution was stirred at 100°C overnight. After the reaction was completed, the reaction solution was quenched with saturated ammonium chloride aqueous solution, extracted with ethyl acetate, and the organic phase was dried over anhydrous sodium sulfate, filtered, and concentrated under reduced pressure to obtain a crude product. The crude product was purified by silica gel column chromatography (eluent: ethyl acetate / petroleum ether = 1 / 4, V / V) to give the title compound 52A (900 mg, yellow solid, yield: 47.7%).

[0270] Step 2: Synthesis of compound 52B 1-[(4R, 5S, 6R)-4,5-bis[(methoxymethyl)oxy]-6-(3,3,4,4-tetramethyl-2-oxa-3-silanpentyl-1-yl)-3,4,5,6-tetrahydro-2H-pyran-3-yl]-6-chloro-4-(cyclopentylamino)pyrazolo[3,4-d]pyrimidine 52A (900 mg, 1.43 mmol) was dissolved in tetrahydrofuran (10 mL), and tetrabutylammonium fluoride (2.86 mL, 2.86 mmol, 1M tetrahydrofuran solution) was added. The reaction solution was stirred at 60°C for 1 hour. After the reaction was completed, the reaction solution was diluted with saturated aqueous ammonium chloride solution, extracted with ethyl acetate, and the organic phase was dried over anhydrous sodium sulfate, filtered, and concentrated under reduced pressure to obtain a crude product. The crude product was purified by silica gel column chromatography (eluent: methanol / dichloromethane = 1 / 10, V / V) to give the title compound 52B (600 mg, yellow solid, yield: 81.4%).

[0271] Step 3: Synthesis of compound 52C [(2R, 3S, 4R)-5-[6-chloro-4-(cyclopentylamino)pyrazolo[3,4-d]pyrimidin-1-yl]-3,4-bis[(methoxymethyl)oxy]-3,4,5,6-tetrahydro-2H-pyran-2-yl]methanol (450 mg, 0.872 mmol), 1-(diethoxyphosphoryl)-2-ethoxy-2-oxoethane-1-diazo 52B (876 mg, 3.49 mmol) and dimeric acetate (38.6 mg, 0.087 mmol) were dissolved in toluene (10 mL), the reaction solution was replaced with nitrogen, and stirred at 100°C for 3 hours. After the reaction was completed, the reaction solution was diluted with water, extracted with ethyl acetate, and the organic phase was dried over anhydrous sodium sulfate, filtered, and concentrated under reduced pressure to obtain a crude product. The crude product was purified by silica gel column chromatography (eluent: ethyl acetate / petroleum ether = 1 / 0, V / V) to give the title compound 52C (550 mg, yellow solid, yield: 85.4%).

[0272] Step 4: Synthesis of Compound 52D Diethyl {1-[(2R,3S,4R,5R)-5-{6-chloro-4-[cyclopentyl(methoxymethyl)amino]pyrazolo[3,4-d]pyrimidin-1-yl}-3,4-bis[(methoxymethyl)oxy]-3,4,5,6-tetrahydro-2H-pyran-2-yl]-4-oxo-2,5-dioxahept-3-yl}phosphonate 52C (350 mg, 0.474 mmol) was dissolved in tetrahydrofuran (10 mL), sodium bis(trimethylsilyl)amide (1.90 mL, 1.90 mmol, 1M tetrahydrofuran solution) was added at -20°C, and the mixture was stirred at this temperature for half an hour, and tetrabutylammonium iodide (87.6 mg, 0.237 mmol) and chloromethyl methyl ether (0.11 mL, 1.42 mmol) were added in sequence. Continue stirring at this temperature for 1 hour. After the reaction is completed, the reaction solution is quenched with saturated aqueous ammonium chloride solution, extracted with ethyl acetate, and the organic phase is washed with saturated brine, dried over anhydrous sodium sulfate, filtered, and concentrated under reduced pressure to obtain a crude product. The crude product is purified by silica gel column chromatography (eluent: ethyl acetate / petroleum ether = 1 / 0, V / V) to obtain the title compound 52D (80 mg, yellow solid, yield: 21.6%).

[0273] Step 5: Synthesis of Compound 52E Diethyl [4-({[(2R, 3S, 4R, 5R)-5-{6-chloro-4-[cyclopentyl(methoxymethyl)amino]pyrazolo[3,4-d]pyrimidin-1-yl}-3,4-bis[(methoxymethyl)oxy]-3,4,5,6-tetrahydro-2H-pyran-2-yl]methyl}oxy)-5-oxo-2,6-dioxaoctane-4-yl]phosphonate 52D (80 mg, 0.102 mmol) was dissolved in ethanol (5 mL), and calcium chloride (34.0 mg, 0.307 mmol) and sodium borohydride (0.010 mL, 0.307 mmol) were added under ice bath, and the reaction solution was stirred at room temperature overnight. After the reaction was completed, the reaction solution was diluted with water, extracted with ethyl acetate, and the organic phase was washed once with saturated brine, dried over anhydrous sodium sulfate, filtered, and concentrated under reduced pressure to obtain a crude product. The crude product was purified by silica gel column chromatography (eluent: methanol / dichloromethane = 1 / 10, V / V) to obtain the title compound 52E (70 mg, yellow oil, yield: 92.5%).

[0274] Step 6: Synthesis of compound 52 Diethyl {1-[(2R,3S,4R,5R)-5-{6-chloro-4-[cyclopentyl(methoxymethyl)amino]pyrazolo[3,4-d]pyrimidin-1-yl}-3,4-bis[(methoxymethyl)oxy]-3,4,5,6-tetrahydro-2H-pyran-2-yl]-3-(hydroxymethyl)-2,5-dioxahexan-3-yl}phosphonate 52E (70 mg, 0.095 mmol) was dissolved in acetonitrile (5 mL). Triethylamine (0.40 mL, 2.85 mmol) and trimethylsilyl bromide (291 mg, 1.90 mmol) were added under ice-cooling conditions, and the reaction solution was stirred at room temperature overnight. After the reaction was completed, the reaction solution was concentrated under reduced pressure, redissolved in trifluoroacetic acid (1 mL), and stirred at room temperature for 1 hour. The reaction solution was concentrated under reduced pressure and purified by reverse preparation (eluent: acetonitrile / 0.1% trifluoroacetic acid aqueous solution = 25% ~ 90%, V / V) to obtain the title compound 52 (14.1 mg, white solid, yield: 27.0%).

[0275] 1H NMR (400 MHz, CD3OD) δ 8.09 (s, 1H), 5.11 (s, 1H), 4.61 – 4.45 (m,1H), 4.35 – 4.22 (m, 1H), 4.16 – 3.75 (m, 9H), 3.52 – 3.44 (m, 1H), 3.43 – 3.35 (m, 3H), 2.14 – 2.01 (m, 2H), 1.86 – 1.76 (m, 2H), 1.73 – 1.55 (m, 4H).

[0276] Examples 53-54 (1-(2R,3S,5S)-5-(2-chloro-4-cyclopentylamino)pyrrolo[2,1-f][1,2,4]triazin-7-yl)-3-hydroxytetrahydro-2H-pyran-2-ylmethoxy)-2-hydroxyethyl)phosphonic acid (Example 53) (1-((2R,3S,5R)-5-(2-chloro-4-(cyclopentylamino)pyrrolo[2,1-f][1,2,4]triazin-7-yl)-3-hydroxytetrahydro-2H-pyran-2-ylmethoxy)-2-hydroxyethyl)phosphonic acid (Example 54)

[0277]

[0278] Step 1: Synthesis of compound 53B Dess-Martin reagent (8.08 g, 19.0 mmol) was added in batches to a solution of 2-phenylhexahydropyran [3, 2-d] [1, 3] dioxin-7-ol 53A (3 g, 12.7 mmol) in anhydrous dichloromethane (30 mL). The reaction mixture was stirred at room temperature for 2 hours. After the reaction was completed, the reaction solution was poured into a saturated aqueous sodium thiosulfate solution to quench it, and then extracted twice with ethyl acetate. The combined organic phase was washed with saturated brine, dried over anhydrous sodium sulfate, filtered, and concentrated under reduced pressure to obtain a crude product. The crude product was purified by silica gel column chromatography (eluent: ethyl acetate / petroleum ether = 1 / 5, V / V) to obtain the title compound 53B (2.2 g, 9.40 mmol, yield: 74.0%).

[0279] Step 2: Synthesis of compound 53C In a dry ice acetone bath, n-butyl lithium (28.1 mL, 44.9 mmol, 1.6 M in n-hexane) was slowly added dropwise to a solution of tert-butyl (7-bromo-2-chloropyrrolo[2,1-f ][1,2,4]triazin-4-yl)(cyclopentyl)carbamate (9.76 g, 23.5 mmol) in tetrahydrofuran (150 mL). The mixture was stirred at this temperature for 15 minutes, and then a solution of 4aR,8aS-2-phenyltetrahydropyrano[3,2-d][1,3]dioxin-7(6H)one 53B (2.2 g, 9.40 mmol) in tetrahydrofuran was added dropwise to the above solution. The reaction solution was stirred at -78°C for 2 hours, and then slowly heated to room temperature. After the reaction was completed, saturated NH4Cl solution was carefully added to quench the reaction, and then the reaction solution was extracted with ethyl acetate. The obtained organic phase was washed with saturated brine, dried over anhydrous sodium sulfate, filtered and concentrated. The crude product was purified by silica gel column chromatography (eluent: ethyl acetate / petroleum ether = 1 / 10, V / V) to obtain the title compound 53C (3 g, 5.25 mmol, yield: 55.9%).

[0280] Step 3: Synthesis of compound 53D Sodium hydroxide (100 mg, 10.5 mmol, 60% in mineral oil) was added to a solution of tert-butyl 2-chloro-7-hydroxy-2-phenylhexahydropyrano[3,2-d][1,3]dioxin-7-yl)(cyclopentyl)carbamate 53C (2.4 g, 4.20 mmol) in tetrahydrofuran (20 mL) at -30°C. After the mixture was stirred at -30°C for 20 minutes, carbon disulfide (0.80 g, 10.5 mmol) was added to the mixture and stirring was continued at -30°C for 20 minutes. Then, iodomethane (1.49 g, 10.5 mmol) was added to the mixture and stirred for 1 hour. The reaction solution was quenched with saturated NH4Cl solution and extracted with ethyl acetate. The obtained organic phase was washed with saturated brine, dried over anhydrous sodium sulfate, filtered and concentrated. The crude product was purified by silica gel column chromatography (eluent: ethyl acetate / petroleum ether = 1 / 2, V / V) to obtain the title compound 53D (890 mg, 1.35 mmol, yield: 32.03%).

[0281] Step 4: Synthesis of Compound 53E Dissolve tert-butyl 2-chloro-7-(methylthio)thiocarbonyloxy)-2-phenylhexahydropyrano[3,2-d][1,3]dioxin-7-yl)pyrrolo[2,1-F][1,2,4]triazine-4-yl)(cyclopentyl)carbamate 53D (890 mg, 1.35 mmol) in toluene (10 mL), add tributyltin hydride (1.09 mL, 4.04 mmol) and azobisisobutyronitrile (110 mg, 0.67 mmol). The reaction solution was reacted at 100°C for 3 hours. After the reaction was completed, the reaction solution was poured into a saturated potassium fluoride aqueous solution and extracted with ethyl acetate. The organic phase was washed once with saturated brine, dried over anhydrous sodium sulfate, filtered, and concentrated. The crude product was purified by silica gel column chromatography (eluent: ethyl acetate / petroleum ether = 1 / 3, V / V) to obtain the title compound 53E (530 mg, 0.96 mmol, yield: 70.9%).

[0282] Step 5: Synthesis of Compound 53F Tert-butyl 2-chloro-7-(4aR,8aS)-2-phenylhexahydropyrano[3,2-d][1,3]dioxin-7-yl)pyrrolo[2,1-f][1,2,4]triazepine-4-yl(cyclopentyl)carbamate 53E (530 mg, 0.955 mmol) was dissolved in a mixed solvent of trifluoroacetic acid (2 mL) and water (2 mL). The reaction solution was reacted at 80°C for 1 hour. After the reaction was completed, the reaction solution was concentrated under reduced pressure, and the crude product was purified by silica gel column chromatography (eluent: methanol / dichloromethane = 1 / 10, V / V) to obtain the title compound 53F (270 mg, 0.74 mmol, yield: 77.1%).

[0283] Step 6: Synthesis of Compound 53G Under nitrogen atmosphere, dimeric rhodium (II) acetate (81.3 mg, 0.37 mmol) and ethyl 2-diazo-2-(diethoxyphosphoryl)acetate (239 mg, 0.96 mmol) were added to a toluene solution (10 mL) of (2R, 3S)-5-(2-chloro-4-cyclopentylamino)pyrrolo[2,1-f][1,2,4]triazine-7-yl)-2-hydroxymethyltetrahydro-2H-pyran-3-ol 53F (270 mg, 0.74 mmol). The reaction mixture was stirred at 100° C. for 3 hours. After the reaction was completed, it was concentrated under reduced pressure. The crude product was purified by silica gel column chromatography (eluent: ethyl acetate / petroleum ether = 1 / 1, V / V) to obtain the title compound 53G (260 mg, 0.44 mmol, yield: 60.0%).

[0284] Step 7: Synthesis of Compound 53H Ethyl 2-(2R,3S)-5-(2-chloro-4-cyclopentylamino)pyrrolo[2,1-f][1,2,4]triazin-7-yl)-3-hydroxytetrahydro-2H-pyran-2-yl)methoxy)-2-(diethoxyphospho)acetate 53G (260 mg, 0.44 mmol) was dissolved in ethanol (10 mL), and calcium chloride (147 mg, 1.32 mmol) and sodium borohydride (44.8 mg, 1.32 mmol) were added at 0°C. The reaction mixture was stirred at room temperature for 1 hour. Saturated NH4Cl solution was added to the reaction solution, and the mixture was extracted with ethyl acetate. The organic phase was washed with saturated brine, dried over anhydrous sodium sulfate, filtered, and concentrated. The crude product was purified by silica gel column chromatography (eluent: methanol / dichloromethane = 1 / 10, V / V) to give the title compound 53H (180 mg, 0.33 mmol, yield: 74.6%).

[0285] Step 8: Synthesis of compounds 53 and 54 Diethyl (1-((2R,3S)-5-(2-chloro-4-(cyclopentylamino)pyrrolo[2,1-f][1,2,4]triazine-7-yl)-3-hydroxytetrahydro-2H-pyran-2-ylmethoxy)-2-hydroxyethyl)phosphonate 53H (80 mg, 0.15 mmol) was dissolved in acetonitrile, trimethylsilyl bromide (224 mg, 1.46 mmol) and triethylamine (0.29 mL, 2.05 mmol) were added at 0°C, and stirred at room temperature for 12 hours. After the reaction was completed, the reaction solution was concentrated under reduced pressure. The crude product was purified by reverse preparation (eluent: acetonitrile / 0.1% trifluoroacetic acid aqueous solution = 25% to 90%, V / V) to give the title compounds 53 (2.3 mg, 0.005 mmol, yield: 3.20%) and 54: (14.8 mg, 0.03 mmol, yield: 20.6%).

[0286] Compound 53: 1 H NMR (400 MHz, CD3OD) δ 6.86 (s, 1H), 6.75 (s, 1H), 4.59 – 4.47 (m, 1H), 4.20 – 4.08 (m, 1H), 3.94 – 3.61 (m, 8H), 3.51 – 3.37 (m, 1H), 2.66 – 2.54 (m, 1H), 2.11 – 1.97 (m, 2H), 1.97 – 1.86 (m, 1H), 1.84 – 1.53 (m, 6H).

[0287] Compound 54: 1H NMR (400 MHz, CD3OD) δ 6.88 (s, 1H), 6.69 – 6.43 (m, 1H), 4.58 – 4.48 (m, 1H), 4.25 – 4.12 (m, 2H), 4.02 – 3.45 (m, 7H), 3.40 – 3.33(m, 1H), 2.42 – 2.33 (m, 1H), 2.15 – 1.99 (m, 2H), 1.86 – 1.74 (m, 3H), 1.70 – 1.52 (m, 4H).

[0288] Embodiment 55 2-(2H-tetrazol-5-yl)methoxy)-1-((2R,3S,4S,5R)-5-(6-chloro-4-cyclopentylamino)-1H-pyrazolo[3,4-d]pyrimidin-1-yl)-3,4-dihydroxytetrahydro-2H-pyran-2-yl)methoxy)ethyl)phosphonic acid

[0289] Compound 55 can be prepared by using compound 50A as the starting material and referring to the synthesis of Example 21. 1 H NMR (400MHz, CD3OD) δ 8.07 – 8.01 (m, 1H), 5.05 – 4.97 (m, 2H), 4.95 – 4.83 (m, 1H), 4.58 – 4.48 (m, 1H), 4.41 – 4.33 (m, 1H), 4.16 – 3.82 (m, 9H), 2.15 – 2.02 (m, 2H), 1.86 – 1.53 (m, 6H).

[0290] Embodiment 56 1-(2H-tetrazol-5-yl)methoxy)-2-((2R,3S,4S,5R)-5-(4-amino-6-chloro-1H-pyrazolo[3,4-d]pyrimidin-1-yl)-3,4-dihydroxytetrahydro-2H-pyran-2-yl)methoxy)-3-methoxypropane-2-yl)phosphonic acid

[0291] Compound 56 can be prepared by using compound 41F as a raw material and referring to the synthesis of Example 21. 1 H NMR (400MHz, CD3OD) δ 8.01 (d, J= 10.2 Hz, 1H), 5.05 – 4.98 (m, 2H), 4.45 – 4.39 (m,1H), 4.23 – 3.98 (m, 8H), 3.94 – 3.75 (m, 3H), 3.37 (d, J = 4.0 Hz, 3H).

[0292] Embodiment 57 (2-(2R,3S,4S,5R)-5-(6-cyclopropylethynyl)-4-(2,3-dihydro-1H-inden-2-yl)amino)-1H-pyrazolo[3,4-d]pyrimidin-1-yl)-3,4-dihydroxytetrahydro-2H-pyran-2-yl)methoxy)-1-hydroxy-3-methoxypropane-2-yl)phosphonic acid

[0293]

[0294] Step 1: Synthesis of compound 57B In a 100 mL single-necked bottle, diethyl (2-((3aS,4R,7R,7aS)-7-(6-chloro-4-((2,3-dihydro-1H-inden-2-yl)(methoxymethyl)amino)-1H-pyrazolo[3,4-d]pyrimidin-1-yl)-2,2-dimethyltetrahydro-4H-[1,3]dioxo[4,5-c]pyran-4-yl)methoxy)-1-hydroxy-3-methoxypropan-2-yl)phosphonate 57A (350 mg, 0.473 mmol), cyclopropylacetylene (156 mg, 2.36 mmol), Pd(PPh3)2Cl2 (33.2 mg, 0.047 mmol) and cuprous iodide (18.0 mg, 0.095 mmol) were added to DMF (6 mL) and triethylamine (1 mL). mL) solution. After nitrogen replacement of the reaction liquid three times, the reaction was continued at 100°C for 15 hours. After the reaction was completed, the reaction liquid was diluted with water and extracted twice with ethyl acetate. The combined organic phase was washed once with saturated brine, dried over anhydrous sodium sulfate, filtered, concentrated under reduced pressure, and purified by silica gel column chromatography (eluent: methanol / dichloromethane = 1 / 10, V / V) to obtain the title compound 57B (350 mg, brown oil, yield: 96.15%).

[0295] Step 2: Synthesis of compound 57 Under ice bath, triethylamine (0.5 mL, 3.90 mmol) and trimethylsilyl bromide (398 mg, 2.60 mmol) were added successively to a solution of diethyl (2-(((3aS,4R,7R,7aS)-7-(6-(cyclopropylethynyl)-4-((2,3-dihydro-1H-inden-2-yl)(methoxymethyl)amino)-1H-pyrazolo[3,4-d]pyrimidin-1-yl)-2,2-dimethyltetrahydro-4H-[1,3]dioxo[4,5-c]pyran-4-yl)methoxy)-1-methoxy-3-(2-(2-(2-(3-methylsilyl)ethoxy)methyl)-2H-tetrazolyl-5-methoxy)propan-2-yl)phosphonate 57B (200 mg, 0.260 mmol) in acetonitrile (5% acetonitrile). The reaction solution was stirred at room temperature overnight. After the phospholipids were completely hydrolyzed, the reaction solution was concentrated under reduced pressure. Then trifluoroacetic acid / water (3 / 1, 2 mL) was added to the obtained crude product. The reaction solution was continued to be stirred at room temperature for 1 hour. After the reaction was completed, the reaction solution was concentrated under reduced pressure to obtain a residue, which was purified by reverse phase preparative chromatography (eluent: acetonitrile / 0.1% trifluoroacetic acid aqueous solution = 25%-90%, V / V) to obtain the title compound 57 (15.2 mg, yield: 9.29%). 1 H NMR (400 MHz, CD3OD) δ8.54 – 8.08 (m, 1H), 7.33 – 7.11 (m, 4H), 5.04 – 4.97 (m, 1H), 4.96 – 4.88(m, 1H), 4.49 – 4.34 (m, 1H), 4.25 – 3.76 (m, 10H), 3.45 (dd, J = 16.0, 7.1Hz, 2H), 3.38 (s, 3H), 3.16 – 2.95 (m, 2H), 1.72 – 1.48 (m, 1H), 1.13 – 0.86(m, 4H).

[0296] Embodiment 58 1-(2H-tetrazol-5-yl)methoxy)-2-((2R,3S,4S,5R)-5-(6-cyclopropylethynyl)-4-((2,3-dihydro-1H-inden-2-yl)amino)-1H-pyrazolo[3,4-d]pyrimidin-1-yl)-3,4-dihydroxytetrahydro-2H-pyran-2-yl)methoxy)-3-methoxypropane-2-yl)phosphonic acid

[0297] Compound 58 can be prepared by using compound 57B as the starting material and referring to the synthesis of Example 21. 1H NMR (400MHz, CD3OD) δ 8.46 – 8.03 (m, 1H), 7.36 – 7.07 (m, 4H), 5.07 – 4.93 (m, 3H), 4.92 – 4.89 (m, 1H), 4.47 – 4.36 (m, 1H), 4.21 – 3.73 (m, 10H), 3.50 – 3.40 (m, 2H), 3.39 – 3.34 (m, 3H), 3.17 – 2.97 (m, 2H), 1.68 – 1.51 (m, 1H), 1.07 – 0.87 (m, 4H).

[0298] Embodiment 59 (2-(2R,3S,4S,5R)-5-(4-((2,3-dihydro-1H-inden-2-yl)amino)-6-propyl-1-propyl-1-yl-1H-pyrazolo[3,4-d]pyrimidin-1-yl)-3,4-dihydroxytetrahydro-2H-pyran-2-yl)methoxy)-1-hydroxy-3-methoxypropane-2-yl)phosphonic acid

[0299] Compound 59 can be prepared by referring to the synthesis of Example 57. 1 H NMR (400 MHz, CD3OD) δ 8.50-8.15 (m, 1H), 7.29 – 7.23 (m, 2H), 7.21 – 7.14 (m, 2H), 5.07 – 5.00 (m, 1H), 4.96-4.88 (m, 1H), 4.47-4.37 (m, 1H), 4.25 – 4.03 (m, 5H), 4.00 –3.75 (m,5H), 3.52-3.38 (m, 2H), 3.38 (s, 3H), 3.10-2.96 (m, 2H), 2.14 (s, 3H).

[0300] Embodiment 60 1-(2H-tetrazol-5-yl)methoxy)-2-((2R,3S,4S,5R)-5-(4-((2,3-dihydro-1H-inden-2-yl)amino)-6-(propyl-1-ynyl)-1H-pyrazolo[3,4-d]pyrimidin-1-yl)-3,4-dihydroxytetrahydro-2H-pyran-2-yl)methoxy)-3-methoxypropane-2-yl)phosphonic acid

[0301] Compound 60 can be prepared by referring to the synthesis of Example 58. 1 H NMR (400 MHz, CD3OD) δ 8.10(d, J = 7.2 Hz, 1H), 7.26 (s, 2H), 7.18 (d, J = 3.0 Hz, 2H), 5.10-4.95 (m, 3H), 4.90-4.81 (m, 1H), 4.42 (m, 1H), 4.21 – 3.99 (m, 7H), 3.95 – 3.65 (m, 3H), 3.48 – 3.39 (m, 2H), 3.36 (d, J = 2.4 Hz, 3H), 3.10-3.01 (m, 2H), 2.14 (s, 3H).

[0302] Embodiment 61 2-(2R,3S,4S,5R)-5-(6-chloro-4-((2,3-dihydro-1H-inden-2-yl)amino)-1H-pyrazolo[3,4-d]pyrimidin-1-yl)-3,4-dihydroxytetrahydro-2H-pyran-2-yl)methoxy)-1,3-dimethoxypropane-2-yl)phosphonic acid

[0303]

[0304] Step 1: Synthesis of compound 61A 2-((3aS,4R,7R,7aS)-7-(6-chloro-4-((2,3-dihydro-1H-inden-2-yl)(methoxymethyl)amino)-1H-pyrazolo[3,4-d]pyrimidin-1-yl)-2,2-dimethyltetrahydro-4H-[1,3]dioxacyclo[4,5-c]pyran-4-yl)methoxy)-1-hydroxy-3-methoxypropan-2-yl)phosphonate 57A (200 mg, 0.264 mmol) and iodomethane (0.2 mL, 2.70 mmol) were dissolved in anhydrous N,N-dimethylformamide (5 mL). Sodium hydride (19.4 mg, 0.810 mmol, 60% mineral oil) was then added under ice bath and the resulting solution was stirred at room temperature for 1 hour. Then 5 mL of saturated aqueous ammonium chloride solution was added to quench the reaction. The organic phases were combined and extracted twice with ethyl acetate (10 mL), washed with saturated brine, dried over anhydrous sodium sulfate, filtered and concentrated under vacuum. The resulting oil was purified by silica gel column chromatography (eluent: methanol / dichloromethane = 1 / 20, V / V) to give the title compound 61A (160 mg, 0.212 mmol, yellow oil, yield: 78.5%).

[0305] Step 2: Synthesis of compound 61 2-((3aS,4R,7R,7aS)-7-(6-chloro-4-((2,3-dihydro-1H-inden-2-yl)(methoxymethyl)amino)-1H-pyrazolo[3,4-d]pyrimidin-1-yl)-2,2-dimethyltetrahydro-4H-[1,3]dioxa[4,5-c]pyran-4-yl)methoxy)-1,3-dimethoxypropan-2-yl)phosphonate 61A (160 mg, 0.212 mmol) was dissolved in anhydrous acetonitrile (5 mL), and then trimethylsilyl bromide (324 mg, 2.12 mmol) and triethylamine (0.4 mL, 2.97 mmol) were added, and the resulting solution was stirred at room temperature overnight. After the phospholipid hydrolysis was completed, the reaction solution was concentrated under vacuum. Then trifluoroacetic acid (1.5 mL) and water (0.5 mL) were added to the mixture. The resulting solution was stirred at room temperature for 1 h. After the reaction was completed, the reaction solution was concentrated under vacuum. The resulting residue was purified by reverse preparation (eluent: acetonitrile / 0.1% trifluoroacetic acid aqueous solution = 25% to 90%, V / V) to obtain the title compound 61 (60 mg, yield: 46.1%). 1 H NMR (400 MHz, CD3OD) δ 8.05 (s, 1H), 7.27 – 7.21 (m, 2H), 7.16 (dd, J = 5.4, 3.3 Hz, 2H), 5.12-4.95 (m, 1H), 4.90-4.81 (m, 1H), 4.51-4.43 (m, 1H), 4.21 – 4.13 (m, 2H), 4.10 – 3.99 (m, 3H), 3.98 – 3.94 (m, 1H), 3.77 (d, J = 10.1 Hz, 4H), 3.45 – 3.35 (m, 8H), 2.99 (dd, J = 15.8, 5.8 Hz, 2H).

[0306] Embodiment 62 2-(2R,3S,4S,5R)-5-(6-chloro-4-((5-cyclopropyl-2,3-dihydro-1H-inden-2-yl)amino)-1H-pyrazolo[3,4-d]pyrimidin-1-yl)-3,4-dihydroxytetrahydro-2H-pyran-2-yl)methoxy)-1-hydroxy-3-methoxypropane-2-yl)phosphonic acid

[0307] Compound 62 can be synthesized by referring to the synthesis steps of Example 1 using commercially available raw materials. 1H NMR (400 MHz, CD3OD)δ 8.05 (s, 1H), 7.10 (d, J = 7.7 Hz, 1H), 6.95 (s, 1H), 6.90 (d, J = 7.8 Hz,1H), 5.04 – 4.96 (m, 1H), 4.92-4.81 (m, 1H), 4.42 (m, 1H), 4.17 – 3.78 (m,10H), 3.37 (m, 5H), 2.97-2.88 (m, 2H), 1.91-1.84 (m, 1H), 0.95 – 0.90 (m,2H), 0.66-0.61 (m, 2H).

[0308] Embodiment 63 (2-(2R,3S,4S,5R)-5-(6-chloro-4-cyclopentylamino)-1H-pyrazolo[3,4-d]pyrimidin-1-yl)-3-ethynyl-3,4-dihydroxytetrahydro-2H-pyran-2-yl)methoxy)-1-hydroxy-3-methoxypropane-2-yl)phosphonic acid

[0309]

[0310] Step 1: Synthesis of compound 63B At -78 ° C, n-butyl lithium (56.2 mL, 140 mmol, 2.5 M in n-hexane) was slowly added dropwise to a solution of trimethylethynylsilane (16.3 g, 166 mmol) in tetrahydrofuran (400 mL). After stirring at this temperature for 0.5 hours, a solution of (1S, 4R, 6S)-4-(((tert-butyldimethylsilyl)oxy)methyl)-3,7-dioxabicyclo[4.1.0]heptane-5-one 63A (33 g, 128 mmol) in tetrahydrofuran was then added dropwise to the reaction solution. The reaction solution was slowly warmed to room temperature and continued to stir at room temperature for 2 hours. An aqueous ammonium chloride solution was slowly added dropwise to the reaction solution and extracted twice with ethyl acetate. The combined organic phase was washed once with saturated sodium chloride, dried over anhydrous sodium sulfate, filtered, and the filtrate was concentrated. The residue was purified by silica gel column chromatography (eluent: PE / EA = 1 / 0 to 3 / 1) to give the title compound 63B (15 g, 42.1 mmol, yield 32.9%) as a colorless oil. 1 H NMR (400 MHz, CDCl3) δ 4.30 (s, 1H), 4.05 (dd, J = 13.4, 3.7 Hz, 1H),3.88 (d, J= 6.1 Hz, 1H), 3.86 (d, J = 9.4 Hz, 1H), 3.73 (dd, J = 9.2, 5.3 Hz, 1H),3.49 (d, J = 4.3 Hz, 1H), 3.44 (dd, J = 9.4, 5.3 Hz, 1H), 3.41 (t, J = 4.0 Hz, 1H),0.86 – 0.83 (m, 9H), 0.15 – 0.11 (m, 9H), 0.04 (d, J = 11.7 Hz, 6H).

[0311] Step 2: Synthesis of compound 63C Hydrazine hydrate (164 mg, 2.8 mmol, 85%) was added to an ethanol solution of (1S, 4R, 5R, 6S)-4-(((tert-butyldimethylsilyl)oxy)methyl)-5-((trimethylsilyl)ethynyl)-3,7-dioxabicyclo[4.1.0]heptane-5-ol 63B (1.0 g, 2.8 mmol), and the reaction solution was stirred at room temperature under nitrogen protection at 60°C overnight. After the reaction was completed, the crude product was concentrated under reduced pressure and used directly in the next step.

[0312] Step 3: Synthesis of compound 63D At -78 ° C, a tetrahydrofuran solution of 2,4,6-trichloropyrimidine-5-carboxaldehyde (0.54 g, 2.6 mmol) and DIEA (0.8 mL, 5.1 mmol) was stirred for 5 minutes, and then the tetrahydrofuran solution of the crude product in the previous step was added dropwise to the above solution. The resulting solution was slowly warmed to room temperature and continued to stir at room temperature for 1 hour. The reaction solution was concentrated under reduced pressure, diluted with ethyl acetate, and washed once with saturated sodium chloride water. The organic phase was dried over sodium sulfate, filtered, and concentrated under reduced pressure. The crude product was purified by column chromatography (eluent: PE / EA = 1 / 0 to 10 / 1) to obtain the title compound 63D (320 mg, yellow solid, 0.59 mmol). LC / MS (ESI) m / z: 545 [M+H] + .

[0313] Step 4: Synthesis of Compound 63E Cyclopentylamine (40.6 mg, 0.48 mmol) was added dropwise to a solution of (2R, 3S, 4S, 5R)-2-(((tert-butyldimethylsilyl)oxy)methyl)-5-(4,6-dichloro-1H-pyrazolo[3,4-d]pyrimidin-1-yl)-3-(trimethylsilyl)ethynyl)tetrahydro-2H-pyran-3,4-diol 63D (200 mg, 0.366 mmol) and triethylamine (74.0 mg, 0.732 mmol) in ethyl acetate (10 mL) under ice bath. The reaction solution was slowly warmed to room temperature and stirred at room temperature for one hour. After the reaction was completed, ethyl acetate was added to dilute, and the mixture was washed once with 10% citric acid, and then washed once with saturated sodium bicarbonate and saturated brine. The organic phase was dried over anhydrous sodium sulfate, filtered, and concentrated under reduced pressure. The crude product was purified by silica gel column chromatography (eluent: petroleum ether / ethyl acetate = 4 / 1) to obtain the title compound 63E (200 mg, 0.34 mmol, light yellow solid, yield: 91.8%). LC / MS (ESI) m / z: 594 [M+H] + .

[0314] Step 5: Synthesis of Compound 63F At zero degrees, sodium hydroxide (674 mg, 16.8 mmol) was slowly added to a solution of (2R, 3S, 4S, 5R)-2-(((tert-butyldimethylsilyl)oxy)methyl)-5-(6-chloro-4-(cyclopentylamino)-1H-pyrazolo[3,4-d ]pyrimidin-1-yl)-3-(trimethylsilyl)ethynyl)tetrahydro-2H-pyran-3,4-diol 63E (2.0 g, 3.37 mmol) in anhydrous tetrahydrofuran (50 mL). The resulting mixture was stirred at zero degrees for 0.5 hours, and then methoxymethyl chloride (1.35 g, 16.8 mmol) was added dropwise. The reaction solution was allowed to react for 1 hour. After the reaction was completed, the reaction solution was poured into ice water and extracted twice with ethyl acetate. The combined organic phase was washed once with saturated sodium chloride, dried over anhydrous sodium sulfate, filtered, and concentrated under reduced pressure to obtain a crude product. The crude product was purified by (eluent: petroleum ether / ethyl acetate = 3 / 1) to give the title compound 63F (1.2 g, yellow oil, 1.65 mmol). LC / MS (ESI) m / z: 726 [M+H] + .

[0315] Step 6: Synthesis of Compound 63G 1-(3R, 4S, 5S, 6R)-6-(((tert-butyldimethylsilyl)oxymethyl)-4,5-bis(methoxymethoxy)-5-(trimethylsilyl)ethynyl)tetrahydro-2H-pyran-3-yl)-6-chloro-N-cyclopentyl-N-(methoxymethyl)-1H-pyrazolo[3,4-d]pyrimidin-4-amine 63F (1.2 g, 1.65 mmol) was dissolved in a tetrahydrofuran solution of TBAF (2.5 mL, 2.5 mmol). The reaction solution was stirred at room temperature for 2 h. After the reaction was completed, the reaction solution was concentrated under reduced pressure, and the crude product was purified by silica gel column chromatography (eluent: ethyl acetate / petroleum ether = 1 / 2, V / V) to obtain the title compound 63G (650 mg, yield: 72.9%) as a yellow oil. LC / MS (ESI) m / z: 540 [M+H] + .

[0316] Step 7: Synthesis of compound 63H To a toluene (15 mL) solution of (2R, 3S, 4S, 5R) -5-(6-chloro-4-(cyclopentyl(methoxymethyl)amino) -1H-pyrazolo[3,4-d ]pyrimidin-1-yl) -3-ethynyl-3,4-bis(methoxymethoxy)tetrahydro-2H-pyran-2-yl) methanol 63G (650 mg, 1.2 mmol) were added 1-(diethoxyphosphoryl) -2-ethoxy-2-oxoethane-1-diazide (903 mg, 3.6 mmol) and rhodium dimer acetate (53.2 mg, 0.12 mmol) in sequence. The mixture was replaced with nitrogen three times and then stirred at 100°C for 3h. After the reaction was completed, ethyl acetate (20 mL) was added to the reaction solution and washed with saturated sodium bicarbonate and saturated brine in sequence. The organic phase was dried over anhydrous sodium sulfate and concentrated under reduced pressure to obtain a crude product. The crude product was purified by silica gel column chromatography (eluent: ethyl acetate / petroleum ether = 4 / 1, V / V) to obtain the title compound 63H (480 mg, 0.63 mmol, yellow oil, yield: 52.3%). LC / MS (ESI) m / z: 762 [M+H] + .

[0317] Step 8: Synthesis of compound 63I Sodium bis(trimethylsilyl)amide (3.5 mL, 3.5 mmol, 1M THF solution) was slowly added dropwise to a solution of ethyl 2-((2R, 3S, 4S, 5R)-5-(6-chloro-4-(cyclopentyl(methoxymethyl)amino)-1H-pyrazolo[3,4-d]pyrimidin-1-yl)-3-ethynyl-3,4-bis(methoxymethoxy)tetrahydro-2H-pyran-2-yl)methoxy)-2-(diethoxyphosphoryl)acetate 63H (480 mg, 0.63 mmol) in anhydrous tetrahydrofuran (10 mL) at -30°C, and stirred at the same temperature for 30 minutes, followed by the addition of tetrabutylammonium iodide (116 mg, 0.32 mmol) and methoxymethyl chloride (225 mg, 3.1 mmol). The resulting reaction solution was stirred at the same temperature for 1 hour. After the reaction was completed, the reaction solution was quenched with saturated ammonium chloride solution and extracted three times with ethyl acetate. The combined organic phase was washed with saturated brine, dried over anhydrous sodium sulfate, filtered, and concentrated under reduced pressure. The crude product was purified by silica gel column chromatography (eluent: methanol / dichloromethane = 1 / 10, V / V) to give the title compound 63I (280 mg, yellow oil, yield: 55.2%). LC / MS (ESI) (m / z): 806 (M+H) + .

[0318] Step 9: Synthesis of compound 63J Under ice bath, calcium chloride (118 mg, 1.08 mmol) and sodium borohydride (40.8 mg, 1.08 mmol) were added to ethyl 2-((2R, 3S, 4S, 5R)-5-(6-chloro-4-(cyclopentyl(methoxymethyl)amino)-1H-pyrazolo[3,4-d]pyrimidin-1-yl)-3-ethynyl-3,4-bis(methoxymethoxy)tetrahydro-2H-pyran-2-yl)methoxy)-2-(diethoxyphosphoryl)-3-methoxypropanoate 63I (250 mg, 0.36 mmol) dissolved in ethanol (5 mL). The reaction solution was stirred at room temperature for 2 hours. The reaction solution was diluted with ethyl acetate, washed once with saturated sodium chloride, dried over anhydrous sodium sulfate, filtered, concentrated under reduced pressure, and purified by silica gel column chromatography (eluent: methanol / dichloromethane = 1 / 10, V / V) to obtain the title compound 63J (100 mg, colorless oil, yield: 36.4%). LC / MS (ESI) m / z: 764 [M+H] + .

[0319] Step 10: Synthesis of compound 63 Under ice bath, triethylamine (286 mg, 2.28 mmol) and trimethylsilyl bromide (360 mg, 2.4 mmol) were added to a solution of diethyl (2-((2R, 3 S, 4 S, 5 R)-5-(6-chloro-4-(cyclopentyl(methoxymethyl)amino)-1H-pyrazolo[3,4-d ]pyrimidin-1-yl)-3-ethynyl-3,4-bis(methoxymethoxy)tetrahydro-2H-pyran-2-yl)methoxy)-1-hydroxy-3-methoxypropane-2-yl)phosphonate 63J (90 mg, 0.12 mmol) in acetonitrile (6 mL) in sequence. The reaction solution was stirred at room temperature overnight. After the phospholipids were completely hydrolyzed, the reaction solution was concentrated under reduced pressure. Then trifluoroacetic acid (2 mL) and water (0.5 mL) were added to the obtained crude product. The reaction solution was stirred at room temperature for another hour. After the reaction was completed, the reaction solution was concentrated under reduced pressure to obtain a residue, which was purified by reverse phase preparative chromatography (eluent: acetonitrile / 0.1% formic acid aqueous solution = 25% ~ 90%, V / V) to obtain the title compound 63 (15 mg, white solid, yield: 22.1%). 1 H NMR (400 MHz, CD3OD) δ 8.10 (s, 1H), 5.05-4.98 (m, 1H), 4.97 – 4.89 (m, 1H), 4.74 – 4.65 (m, 1H), 4.61 – 4.40 (m, 3H), 4.37-4.25 (m, 1H), 4.09-3.93 (m, 3H), 3.91-3.81(m, 3H), 3.41 – 3.34 (m, 3H), 2.93-2.90 (d, J = 3.7 Hz, 1H), 2.16 – 2.04 (m,2H), 1.85-1.75 (m, 2H), 1.73 – 1.54 (m, 4H).

[0320] Embodiment 64 1-(2H-tetrazol-5-yl)methoxy)-2-((2R,3S,4S,5R)-5-(6-chloro-4-cyclopentylamino)-1H-pyrazolo[3,4-d]pyrimidin-1-yl)-3-ethynyl-3,4-dihydroxytetrahydro-2H-pyran-2-yl)methoxy)-3-methoxypropane-2-yl)phosphonic acid

[0321] Compound 64 can be prepared by referring to the synthesis of Example 21 using starting material 63J. 1H NMR (400 MHz, CD3OD)δ 8.08 (s, 1H), 5.06-4.97 (m, 2H), 4.97 – 4.89 (m, 1H), 4.84 – 4.82 (m, 1H), 4.66 – 4.36 (m, 4H), 4.33-4.22 (m, 1H), 4.14-4.00 (m, 3H), 3.91-3.78 (m, 3H),3.41 – 3.34 (m, 3H), 2.90-2.86 (m, 1H), 2.16 – 2.04 (m, 2H), 1.80 (s, 2H),1.73 – 1.54 (m, 4H).

[0322] Embodiment 65 1-(2H-tetrazol-5-yl)methoxy)-2-((2R,3S,4S,5R)-5-(6-chloro-4-(5-cyclopropyl-2,3-dihydro-1H-indene-2-amino)-1H-pyrazolo[3,4-d]pyrimidin-1-yl)-3,4-dihydroxytetrahydro-2H-pyran-2-yl)methoxy)-3-methoxypropane-2-yl)phosphonic acid

[0323] Compound 65 can be prepared by referring to the synthesis of Example 21 using commercially available raw materials. 1 H NMR (400 MHz, CD3OD)δ 8.02 – 7.97 (m, 1H), 7.10 (d, J = 7.6 Hz, 1H), 6.95 (s, 1H), 6.90 (d, J = 7.6Hz, 1H), 5.04 – 4.94 (m, 3H), 4.94 – 4.81 (m, 1H), 4.44 – 4.38 (m, 1H), 4.21 – 3.99 (m, 7H), 3.97 – 3.88 (m, 1H), 3.85 – 3.71 (m, 2H), 3.41 – 3.33 (m,5H), 2.99 – 2.85 (m, 2H), 1.91 – 1.82 (m, 1H), 0.95 – 0.87 (m, 2H), 0.66 –0.59 (m, 2H).

[0324] Embodiment 66 1-(2H-tetrazol-5-yl)methoxy)-2-((2R,3S,4S,5R)-5-(6-chloro-4-cyclopentylamino)-1H-pyrazolo[3,4-d]pyrimidin-1-yl)-3,4-dihydroxytetrahydro-2H-pyran-2-yl)methoxy)-3-hydroxypropan-2-yl)phosphonic acid

[0325]

[0326] Step 1: Synthesis of compound 66B Ethyl 2-(((3aS,4R,7R,7aS)-7-(6-chloro-4-(cyclopentylamino)-1H-pyrazolo[3,4-d]pyrimidin-1-yl)-2,2-dimethyltetrahydro-4H-[1,3]dioxo[4,5-c]pyran-4-yl)methoxy)-2-(diethoxyphosphoryl)acetate 66A (2.0 g, 3.10 mmol) was dissolved in tetrahydrofuran (25 mL) and cooled to -15°C. Sodium bis(trimethylsilyl)amide (9.30 mL, 9.30 mmol, 1 M in tetrahydrofuran) was added under nitrogen and the mixture was stirred at -15°C for 30 min. Tetrabutylammonium iodide (0.57 g, 1.55 mmol) was added to the above mixture, and then 2-(trimethylsilyl)ethoxymethyl chloride (2.58 g, 15.48 mmol) was added dropwise at -15°C and stirred at -15°C for 1 hour. The reaction was quenched with saturated aqueous ammonium chloride and extracted three times with ethyl acetate. The combined organic layer was washed with brine, dried over anhydrous sodium sulfate, filtered and concentrated to dryness under reduced pressure. The residue was purified by silica gel column chromatography (eluent: ethyl acetate / petroleum ether = 1 / 1, V / V) to obtain the title compound 66B (1 g, yield: 35.6%) as a colorless oil.

[0327] Step 2: Synthesis of compound 66C Ethyl 2-(((3aS,4R,7R,7aS)-7-(6-chloro-4-(cyclopentyl((2-(trimethylsilyl)ethoxy)methyl)amino)-1H-pyrazolo[3,4-d]pyrimidin-1-yl)-2,2-dimethyltetrahydro-4H-[1,3]dioxo[4,5-c]pyran-4-yl)methoxy)-2-(diethoxyphosphoryl)-3-(2-(trimethylsilyl)ethoxy)propanoate 66B (1.0 g, 1.10 mmol) was dissolved in ethanol (10 mL), calcium chloride (0.37 g, 3.31 mmol) and sodium borohydride (0.13 g, 3.3 mmol) were added at 0 °C and stirred at room temperature for 2 h. The reaction mixture was quenched with saturated aqueous ammonium chloride solution and extracted three times with ethyl acetate. The combined organic layer was dried over anhydrous sodium sulfate, filtered, and concentrated to dryness under reduced pressure. The residue was purified by silica gel column chromatography (eluent: methanol / dichloromethane = 1 / 10, V / V) to give the title compound 66C (800 mg, yield: 84%) as a colorless oil. LC / MS (ESI) (m / z): 864 (M+H + ).

[0328] Step 3: Synthesis of compound 66D Diethyl (2-(((3aS,4R,7R,7aS)-7-(6-chloro-4-(cyclopentyl((2-(trimethylsilyl)ethoxy)methyl)amino)-1H-pyrazolo[3,4-d]pyrimidin-1-yl)-2,2-dimethyltetrahydro-4H-[1,3]dioxo[4,5-c]pyran-4-yl)methoxy)-1-hydroxy-3-(2-(trimethylsilyl)propan-2-yl)phosphonate 66C (400 mg, 0.46 mmol) and 5-(bromomethyl)-2-((2-(trimethylsilyl)ethoxy)methyl)-2H-tetrazole (271 mg, 0.93 mmol) were dissolved in N,N-dimethylformamide (5 mL) solution and cooled to 0°C. Sodium hydride (56 mg, 1.39 mmol, The solution was stirred at 0°C for 30 minutes and then quenched with saturated aqueous ammonium chloride solution (50 mL). The mixture was extracted twice with ethyl acetate (50 mL), and the combined organic layer was washed with brine and dried over sodium sulfate. The solution was filtered and concentrated, and the residue was purified by silica gel column chromatography (eluent: methanol / dichloromethane = 1 / 10, V / V) to give the title compound 66D (460 mg, yield: 92.3%) as a colorless oil. LC-MS m / e (ESI): 1076 (M+H + ).

[0329] Step 4: Synthesis of compound 66 Diethyl (2-(((3aS,4R,7R,7aS)-7-(6-chloro-4-(cyclopentyl((2-(trimethylsilyl)ethoxy)methyl)amino)-1H-pyrazolo[3,4-d]pyrimidin-1-yl)-2,2-dimethyltetrahydro-4H-[1,3]dioxo[4,5-c]pyran-4-yl)methoxy)-1-(2-(trimethylsilyl)ethoxy)-3-(2-(2-(trimethylsilyl)ethoxy)methyl)-2H-tetrazol-5-yl)methyl)propan-2-yl)phosphonate 66D (460 mg, 0.43 mmol) was dissolved in acetonitrile (5 mL) and triethylamine (0.9 mL, 6.41 mmol) was added, followed by trimethylsilane bromide (0.6 mL, 4.27 mmol). The solution was stirred overnight and then concentrated under reduced pressure. The resulting residue was dissolved in water / trifluoroacetic acid (2 / 3, 5 mL), and the reaction mixture was stirred at room temperature overnight. The mixture was concentrated and purified by reverse phase preparative chromatography (eluent: acetonitrile / 0.1% formic acid aqueous solution = 20% ~ 70%, V / V) to obtain the title product 66 (80 mg, yield: 30.2%). 1 H NMR (400 MHz, CD3OD) δ 8.13 – 7.95 (m, 1H), 5.08 – 4.97 (m, 2H), 4.84 – 4.79 (m, 1H), 4.62 – 4.48(m, 1H), 4.44 – 3.74 (m, 11H), 2.23 – 1.97 (m, 2H), 1.90 – 1.50 (m, 6H).

[0330] Embodiment 67 2-((2R,3S,4S,5R)-5-(6-chloro-4-((2,3-dihydro-1H-inden-2-yl)amino)-1H-pyrazolo[3,4-d]pyrimidin-1-yl)-3,4-dihydroxytetrahydro-2H-pyran-2-yl)methoxy)-1-hydroxy-3-methoxypropan-2-yl)phosphonic acid methyl ester

[0331]

[0332] Step 1: Synthesis of compound 67B [3aS, 4R, 7aS, 7R)-7-[6-chloro-4-(2,3-dihydro-1H-inden-2-ylamino)pyrazolo[3,4-d]pyrimidin-1-yl]-2,2-dimethyl-4,6,7,7a-tetrahydro-3aH-[1,3]dioxo[4,5-c]pyran-4-yl]methanol 67A (1 g, 2.12 mmol) was dissolved in toluene (15 mL), followed by the addition of methyl 2-diazo-2-(dimethoxyphosphoryl)acetate (1.33 g, 6.36 mmol) and rhodium dimer acetate (0.05 g, 0.106 mmol), and the mixture was replaced with nitrogen three times. The resulting solution was stirred at 100°C for 3 hours. The reaction solution was diluted with ethyl acetate and washed with a saturated saline solution. The obtained organic phase was dried over anhydrous sodium sulfate, filtered, and concentrated. The obtained oil was purified by silica gel column chromatography (eluent: ethyl acetate / petroleum ether = 1 / 1, V / V) to obtain the title compound 67B (900 mg, 1.380 mmol, yellow oil, yield: 65.1%). LC / MS (ESI) m / z: 652 [M+H] + .

[0333] Step 2: Synthesis of Compound 67C Methyl 2-(3aS,4R,7R,7aS)-7-(6-chloro-4-((2,3-dihydro-1H-inden-2-yl)amino)-1H-pyrazolo[3,4-d]pyrimidin-1-yl)-2,2-dimethyltetrahydro-4H-[1,3]dioxacyclo[4,5-c]pyran-4-yl)methoxy)-2-(dimethoxyphospho)acetate 67B (900 mg, 1.38 mmol) was dissolved in anhydrous tetrahydrofuran (15 mL) solution, and then sodium bis(trimethylsilyl)amide (6.90 mL, 6.90 mmol) was added at -20°C, and the resulting solution was stirred at -20°C for 0.5 hours. Tetrabutylammonium iodide (255 mg, 0.690 mmol) and chloromethyl methyl ether (1.05 mL, 13.8 mmol) were then added to the mixture and the resulting solution was stirred at -20 °C for 1 h. 20 ml of saturated aqueous ammonium chloride solution was then added to quench the reaction. The reaction solution was diluted with ethyl acetate and washed with saturated aqueous salt solution. The obtained organic phase was dried over anhydrous sodium sulfate, filtered, and concentrated. The obtained oil was purified by silica gel column chromatography (eluent: ethyl acetate / petroleum ether = 1 / 1, V / V) to give the title compound 67C (600 mg, 0.811 mmol, yellow oil, yield: 58.7%). LC / MS(ESI) m / z: 740 [M+H] + .

[0334] Step 3: Synthesis of compound 67D Methyl 2-((3aS,4R,7R,7aS)-7-(6-chloro-4-((2,3-dihydro-1H-inden-2-yl)(methoxymethyl)amino)-1H-pyrazolo[3,4-d]pyrimidin-1-yl)-2,2-dimethyltetrahydro-4H-[1,3]dioxacyclo[4,5-c]pyran-4-yl)methoxy)-2-(dimethoxyphospho)-3-methoxypropanoate 67C (600 mg, 0.811 mmol) was dissolved in anhydrous ethanol (10 mL), and then sodium borohydride (92.0 mg, 2.43 mmol) and calcium chloride (270 mg, 2.43 mmol) were added under ice bath, and the resulting solution was stirred at room temperature for 1 hour. Then, 10 mL of saturated aqueous ammonium chloride solution was added to quench the reaction. Extracted twice with ethyl acetate, the combined organic phase was washed with saturated brine, dried over anhydrous sodium sulfate, filtered and concentrated under vacuum, and the obtained oil was purified by silica gel column chromatography (eluent: methanol / dichloromethane = 1 / 10, V / V) to obtain the title compound 67D (480 mg, 0.674 mmol, colorless oil, yield: 83.1%). LC / MS (ESI) m / z: 712 [M+H] + .

[0335] Step 4: Synthesis of compound 67E 2-((3aS,4R,7R,7aS)-7-(6-chloro-4-((2,3-dihydro-1H-inden-2-yl)(methoxymethyl)amino)-1H-pyrazolo[3,4-d]pyrimidin-1-yl)-2,2-dimethyltetrahydro-4H-[1,3]dioxacyclo[4,5-c]pyran-4-yl)methoxy)-1-hydroxy-3-methoxypropan-2-yl)phosphonate 67D (140 mg, 0.197 mmol) was dissolved in acetone (10 mL) and sodium iodide (59.1 mg, 0.394 mmol) was added. The resulting solution was stirred at 60 °C for 12 hours and then concentrated under reduced pressure. The residue was dissolved in water, acidified to pH about 1, and extracted twice with ethyl acetate. The combined organic layers were dried, filtered, and concentrated to dryness under reduced pressure to give the crude title compound 67E (130 mg).

[0336] Step 5: Synthesis of compound 67 Methyl hydrogen (2-((3aS,4R,7R,7aS)-7-(6-chloro-4-((2,3-dihydro-1H-inden-2-yl)(methoxymethyl)amino)-1H-pyrazolo[3,4-d]pyrimidin-1-yl)-2,2-dimethyltetrahydro-4H-[1,3]dioxacyclo[4,5-c]pyran-4-yl)methoxy)-1-hydroxy-3-methoxypropan-2-yl)phosphonate 67E (130 mg) was dissolved in trifluoroacetic acid (1.5 mL) and water (0.5 mL). The resulting solution was stirred at room temperature for 1 h. After completion of the reaction, the reaction solution was concentrated under vacuum. The obtained residue was purified by reverse preparation (eluent: acetonitrile / 0.1% trifluoroacetic acid aqueous solution = 25% to 90%, V / V) to obtain the title compound 67 (62 mg, 0.101 mmol, white solid, yield: 51.36%). 1 H NMR (400 MHz, CD3OD) δ8.04 (s, 1H), 7.27 – 7.13 (m, 4H), 5.05 – 4.97 (m, 1H), 4.92-4.81 (m, 1H), 4.48-4.41 (m, 1H), 4.25 – 3.81 (m, 8H), 3.80 -3.65 (m, 5H), 3.45 – 3.36 (m, 5H), 3.05-2.95 (m, 2H).

[0337] Embodiment 68 1-(2H-tetrazol-5-yl)methoxy)-2-((2R,3S,4S,5R)-5-(6-chloro-4-((2,3-dihydro-1H-inden-2-yl)amino)-1H-pyrazolo[3,4-d]pyrimidin-1-yl)-3,4-dihydroxytetrahydro-2H-pyran-2-yl)methoxy)-3-methoxypropan-2-yl)phosphonic acid methyl ester

[0338] Compound 68 can be prepared by using 67D as the starting material and referring to the synthesis of Example 21. 1 H NMR (400 MHz, CD3OD) δ 7.99 (d, J = 8.5 Hz, 1H), 7.27 – 7.13 (m, 4H), 4.99 (s, 3H), 4.42 (s, 1H), 4.22 – 4.10 (m, 2H), 4.08 – 3.85 (m, 7H), 3.81 – 3.71 (m, 5H), 3.45 – 3.34 (m, 5H), 3.02-2.97 (m, 2H).

[0339] Embodiment 69 2-(2R,3S,4S,5R)-5-(6-chloro-4-((2,3-dihydro-1H-inden-2-yl)amino)-1H-pyrazolo[3,4-b]pyridin-1-yl)-3,4-dihydroxytetrahydro-2H-pyran-2-yl)methoxy)-1-hydroxy-3-methoxypropane-2-yl)phosphonic acid

[0340] Compound 69 can be synthesized by referring to the synthesis steps of Example 1 using commercially available raw materials. 1 H NMR (400 MHz, CD3OD) δ8.11 (d, J = 2.7 Hz, 1H), 7.32 – 7.21 (m, 2H), 7.16 (dd, J = 5.4, 3.3 Hz, 2H), 6.35 (s, 1H), 4.94 – 4.91 (m, 1H), 4.54 (s, 1H), 4.46 – 4.38 (m, 1H), 4.26 – 4.01 (m, 5H), 4.00 – 3.77 (m, 5H), 3.44 (dd, J = 16.1, 7.1 Hz, 2H), 3.37 (s, 3H), 3.03 (dd, J = 16.0, 4.7 Hz, 2H).

[0341] Embodiment 70 1-(2H-tetrazol-5-yl)methoxy)-2-((2R,3S,4S,5R)-5-(6-chloro-4-((2,3-dihydro-1H-inden-2-yl)amino)-1H-pyrazolo[3,4-b]pyridin-1-yl)-3,4-dihydroxytetrahydro-2H-pyran-2-yl)methoxy)-3-methoxypropane-2-yl)phosphonic acid

[0342] Compound 70 can be synthesized by referring to the synthesis steps of Example 21 using commercially available raw materials. 1H NMR (400 MHz, CD3OD)δ 8.08 – 8.03 (m, 1H), 7.27 – 7.21 (m, 2H), 7.18 – 7.13 (m, 2H), 6.34 – 6.31(m, 1H), 5.04 – 4.89 (m, 3H), 4.57 – 4.47 (m, 1H), 4.44 – 4.37 (m, 1H), 4.24 – 4.00 (m, 7H), 3.95 – 3.87 (m, 1H), 3.85 – 3.73 (m, 2H), 3.50 – 3.39 (m, 2H), 3.36 (d, J = 3.3 Hz, 3H), 3.06 – 2.97 (m, 2H).

[0343] Embodiment 71 ((2R,3S,4S,5R)-5-(6-chloro-4-cyclopentylamino)-1H-pyrazolo[3,4-b]pyridin-1-yl)-3,4-dihydroxytetrahydro-2H-pyran-2-yl)methoxy(hydroxy)phospho)methyl)phosphonic acid

[0344]

[0345] Step 1: Synthesis of compound 71 N,N-Diisopropylethylamine (67.1 mg, 0.52 mmol) was added to a suspension of methylenephosphonium dichloride (354 mg, 1.42 mmol) in tetrahydrofuran (5 mL) at 0°C. To the resulting mixture was added a solution of (3aS,4R,7R,7aS)-7-(6-chloro-4-(cyclopentylamino)-1H-pyrazolo[3,4-b]pyridin-1-yl)-2,2-dimethyltetrahydro-4H-[1,3]dioxo[4,5-c]pyran-4-yl)methanol 71A (200 mg, 0.473 mmol) in tetrahydrofuran (2 mL) dropwise over 2 min. After the addition was complete, the resulting mixture was stirred at 0°C for an additional 15 min before the solution was transferred via cannula to a pre-cooled (0°C) flask containing 2 M hydrochloric acid (2.5 mL). The reaction mixture was heated to 30°C and stirred at 30°C for 16 hours. After completion, it was purified by reverse phase preparative chromatography (eluent: acetonitrile / 0.1% formic acid aqueous solution = 0 ~ 70%, V / V) to obtain the title compound 71 (140 mg, yield: 54.7%) as a white solid. 1H NMR (400 MHz, CD3OD) δ 8.13 (s,1H), 6.24 (s, 1H), 4.95 – 4.85 (m, 1H), 4.43 – 4.35 (m, 1H), 4.34 – 4.24 (m,2H), 4.20 – 4.09 (m, 3H), 4.06 – 3.90 (m, 2H), 2.51 (t, J = 21.0 Hz, 2H), 2.15–2.03 (m, 2H), 1.85–1.60 (m, 6H).

[0346] Embodiment 72 (2R,3S,4S,5R)-5-(6-chloro-4-((S)-1-(2-fluorophenyl)ethyl)amino)-1H-pyrazolo[3,4-b]pyridin-1-yl)-3,4-dihydroxytetrahydro-2H-pyran-2-yl)methoxy(hydroxy)phospho)methyl)phosphonic acid

[0347]

[0348] Step 1: Synthesis of compound 72B 1-(((3aS,4R,7R,7aS)-4-(((tert-butyldiphenylsilyl)oxy)methyl)-2,2-dimethyltetrahydro-4H-[1,3]dioxacyclo[4,5-c]pyran-7-yl)-4,6-dichloro-1H-pyrazolo[3,4-b]pyridine 72A (500 mg, 0.816 mmol) was dissolved in dimethyl sulfoxide (5 mL) solution and triethylamine (0.34 mL, 2.45 mmol) and (S)-1-(2-fluorophenyl)ethan-1-amine (170 mg, 1.22 mmol) were added. The mixture was stirred at 80 °C overnight. After the reaction was completed, the mixture was diluted with ethyl acetate (50 mL) and washed twice with brine (30 mL). The organic phase was concentrated under reduced pressure and the residue was purified by silica gel column chromatography (eluent: ethyl acetate / petroleum ether = 1 / 3, V / V) to afford the title compound 72B (370 mg, yield: 63.4%) as a white solid.

[0349] Step 2: Synthesis of compound 72C 1-((3aS,4R,7R,7aS)-4-(((tert-butyldiphenylsilyl)oxy)methyl)-2,2-dimethyltetrahydro-4H-[1,3]dioxacyclo[4,5-c]pyran-7-yl)-6-chloro-N-((S)-1-(2-fluorophenyl)ethyl)-1H-pyrazolo[3,4-b]pyridin-4-amine 72B (370 mg, 0.517 mmol) was dissolved in tetrahydrofuran (5 mL) and tetrabutylammonium fluoride solution (0.78 mL, 0.78 mmol, 1 M in tetrahydrofuran) was added. The reaction was stirred at room temperature overnight. Upon completion, it was diluted with ethyl acetate (50 mL) and washed twice with water (30 mL), dried over anhydrous sodium sulfate, filtered and concentrated. The residue was purified by silica gel column chromatography (eluent: ethyl acetate / petroleum ether = 1 / 1, V / V) to give the title compound 72C (150 mg, yield: 60.8%) as a white solid.

[0350] Step 3: Synthesis of compound 72 N,N-Diisopropylethylamine (42.3 mg, 0.33 mmol) was added to a suspension of methylenephosphonium dichloride (204 mg, 0.82 mmol) in tetrahydrofuran (1 mL) at 0° C. To the resulting mixture was added dropwise a solution of ((3aS,4R,7R,7aS)-7-(6-chloro-4-(((S)-1-(2-fluorophenyl)ethyl)amino)-1H-pyrazolo[3,4-b]pyridin-1-yl)-2,2-dimethyltetrahydro-4H-[1,3]dioxo[4,5-c]pyran-4-yl)methanol 72C (130 mg, 0.273 mmol) in tetrahydrofuran (2 mL) over 2 minutes. After the addition was complete, the resulting mixture was stirred at 0°C for another 15 minutes, and then the solution was transferred to a precooled (0°C) flask containing 0.2M hydrochloric acid (2 mL) via cannula. The reaction mixture was warmed to 30°C and stirred at 30°C for 16 hours. After completion, it was purified by reverse phase preparative chromatography (eluent: acetonitrile / 0.1% formic acid in water = 0 ~ 70%, V / V) to obtain the title compound 72 (63 mg, yield: 38.9%) as a white solid. 1H NMR (400 MHz, CD3OD) δ 8.18(s, 1H), 7.42 – 7.33 (m, 1H), 7.31 – 7.24 (m, 1H), 7.15 – 7.07 (m, 2H), 6.00(s, 1H), 5.12 – 5.01 (m, 1H), 4.95 – 4.85 (m, 1H), 4.43 – 4.23 (m, 3H), 4.18 – 4.05 (m, 3H), 3.98 – 3.88 (m, 1H), 2.73 – 2.72 (m, 1H), 2.51 (t, J = 21.0 Hz,2H), 1.64 (d, J = 6.8 Hz, 3H).

[0351] Embodiment 73 (2-(2R,3S,4S,5R)-5-(6-chloro-4-cyclopentylamino)-1H-pyrazolo[3,4-b]pyridin-1-yl)-3,4-dihydroxytetrahydro-2H-pyran-2-yl)methoxy)-1-hydroxy-3-methoxypropan-2-yl)phosphonic acid

[0352] Compound 73 can be synthesized by referring to the synthesis steps of Example 1 using commercially available raw materials. 1 H NMR (400 MHz, DMSO- d 6) δ 8.19 (s, 1H), 7.57 (d, J = 6.5 Hz, 1H), 6.18 (s, 1H), 4.83 – 4.77 (m,1H), 4.25 (dd, J = 7.1, 2.9 Hz, 1H), 4.02-3.93 (m, 3H), 3.91-3.85 (m, 2H), 3.81-3.74 (m, 2H), 3.72-3.62 (m, 4H), 3.28 (s, 3H), 2.03-1.95 (m, 2H), 1.74-1.65 (m, 2H), 1.64-1.53 ​​(m, 4H).

[0353] Embodiment 74 1-(2H-tetrazol-5-yl)methoxy)-2-((2R,3S,4S,5R)-5-(6-chloro-4-cyclopentylamino)-1H-pyrazolo[3,4-b]pyridin-1-yl)-3,4-dihydroxytetrahydro-2H-pyran-2-yl)methoxy)-3-methoxypropane-2-yl)phosphonic acid

[0354] Compound 74 can be synthesized by referring to the synthesis steps of Example 21 using commercially available raw materials. 1 H NMR (400 MHz, DMSO- d 6) δ 8.17 (d, J = 5.5 Hz, 1H), 7.56 (d, J = 6.6 Hz, 1H), 6.17 (s, 1H), 4.99-4.95 (m, 2H), 4.80 – 4.73 (m, 1H), 4.24 – 4.18 (m, 1H), 4.00-3.94 (m, 2H), 3.93-3.78 (m, 5H), 3.75-3.61 (m, 4H), 3.27 (s, 3H), 2.05 – 1.94 (m, 2H), 1.75-1.66 (m, 2H), 1.64-1.54 (m, 4H).

[0355] Embodiment 75 1-(2H-tetrazol-5-yl)methoxy)-2-((2R,3S,4S,5R)-5-(6-chloro-4-cyclopentylamino)-1H-pyrazolo[3,4-b]pyridin-1-yl)-3,4-dihydroxytetrahydro-2H-pyran-2-yl)methoxy)-3-hydroxypropan-2-yl)phosphonic acid

[0356] Compound 75 can be synthesized by referring to the synthesis steps of Example 66 using commercially available raw materials. 1 H NMR (400 MHz, DMSO- d 6) δ 8.21 – 8.15 (m, 1H), 7.57 (d, J = 6.7 Hz, 1H), 6.18 (s, 1H), 4.99 (s,2H), 4.81 – 4.73 (m, 1H), 4.21 – 4.18 (m, 1H), 4.01 – 3.95 (m, 3H), 3.92 –3.80 (m, 6H), 3.79 – 3.63 (m, 3H), 2.06 – 1.95 (m, 2H), 1.76 – 1.51 (m, 6H).

[0357] Embodiment 76 2-(2R,3S,4S,5R)-5-(6-chloro-4-((S)-1-(2-fluorophenyl)ethyl)amino)-1H-pyrazolo[3,4-b]pyridin-1-yl)-3,4-dihydroxytetrahydro-2H-pyran-2-yl)methoxy)-1-hydroxy-3-methoxypropane-2-yl)phosphonic acid

[0358] Compound 76 can be synthesized by referring to the synthesis steps of Example 1 using commercially available raw materials. 1 H NMR (400 MHz, CD3OD)δ 8.18 (s, 1H), 7.36 (t, J = 7.6 Hz, 1H), 7.32 – 7.25 (m, 1H), 7.17 – 7.08 (m,2H), 6.01 (s, 1H), 5.11 -5.00 (m, 1H), 4.93 – 4.85 (m, 1H), 4.42 – 4.35 (m,1H), 4.24 – 3.90 (m, 8H), 3.87 – 3.75 (m, 2H), 3.39 (d, J = 2.6 Hz, 3H), 1.64(d, J = 6.8 Hz, 3H).

[0359] Embodiment 77 1-(2H-tetrazol-5-yl)methoxy)-2-((2R,3S,4S,5R)-5-(6-chloro-4-((S)-1-(2-fluorophenyl)ethyl)amino)-1H-pyrazolo[3,4-b]pyridin-1-yl)-3,4-dihydroxytetrahydro-2H-pyran-2-yl)methoxy)-3-methoxypropane-2-yl)phosphonic acid

[0360] Compound 77 can be synthesized by referring to the synthesis steps of Example 21 using commercially available raw materials. 1 H NMR (400 MHz, CD3OD)δ 8.12 (d, J= 8.7 Hz, 1H), 7.40 -7.23 (m, 6.6 Hz, 2H), 7.16 – 7.07 (m, 2H), 6.00 (s, 1H), 5.11 – 4.93 (m, 4H), 4.44 -4.34 (m, 1H), 4.23 – 3.90 (m, 8H), 3.88 – 3.72 (m, 2H), 3.41 – 3.20 (m, 6H), 1.63 (d, J = 6.8 Hz, 3H).

[0361] Embodiment 78 (2-(((2R,3S,4S,5R)-5-(6-chloro-4-(cyclopentylamino)-1H-pyrazolo[3,4-d]pyrimidin-1-yl)-3,4-dihydroxytetrahydro-2H-pyran-2-yl)methoxy)-1,3-dihydroxypropane-2-yl)phosphonic acid

[0362]

[0363] Diethyl (2-(((3aS,4R,7R,7aS)-7-(6-chloro-4-(cyclopentyl((2-(trimethylsilyl)ethoxy)methyl)amino)-1H-pyrazolo[3,4-d]pyrimidin-1-yl)-2,2-dimethyltetrahydro-4H-[1,3]dioxo[4,5-c]pyran-4-yl)methoxy)-1-hydroxy-3-(2-(trimethylsilyl)propan-2-yl)phosphonate (200 mg, 0.23 mm ol) was dissolved in acetonitrile (5 mL) and triethylamine (0.5 mL, 3.47 mmol) was added, followed by trimethylsilyl bromide (0.3 mL, 2.31 mmol). After the reaction solution was stirred overnight, the phospholipid hydrolysis was completed and concentrated under reduced pressure. The resulting residue was dissolved in water / trifluoroacetic acid (1 / 2, 3 mL), and the reaction mixture was stirred at room temperature for 6 hours. The mixture was concentrated and purified by reverse phase preparative chromatography (eluent: acetonitrile / 0.1% formic acid aqueous solution = 20% to 70%, V / V) to obtain compound 78 (46 mg, yield: 37.0%). 1 H NMR (400 MHz, CD3OD) δ 8.07 (s, 1H), 4.84 – 4.80 (m, 1H), 4.60 – 4.48 (m, 1H), 4.45 – 4.37 (m, 1H), 4.26 – 3.88 (m, 10H), 2.17 – 2.02 (m, 2H),1.86 – 1.53 (m, 6H).

[0364] Embodiment 79 (2-(((2R,3S,4S,5R)-5-(6-chloro-4-(cyclopentylamino)-1H-pyrazolo[3,4-d]pyrimidin-1-yl)-3,4-dihydroxytetrahydro-2H-pyran-2-yl)methoxy)-1,3-dihydroxypropane-2-yl)phosphonic acid

[0365]

[0366] Step 1: Synthesis of compound 79A Ethyl 2-((3aR,4R,6R,6aS)-6-(6-chloro-4-((2,3-dihydro-1H-inden-2-yl)amino)-1H-pyrazolo[3,4-b]pyridin-1-yl)-2,2-dimethyltetrahydro-4H-cyclopentyl[d][1,3]dioxolanyl)-2-(diethoxyphospho)acetate 66A (1 g, 1.55 mmol) was dissolved in anhydrous tetrahydrofuran (10 mL) solution, and then sodium bis(trimethylsilyl)amide (7.74 mL, 7.74 mmol, 1 M tetrahydrofuran solution) was added at -20°C, and the resulting solution was stirred at -20°C for 0.5 hours. Tetrabutylammonium iodide (285 mg, 0.77 mmol) and chloromethyl methyl ether (1.25 g, 15.5 mmol) were then added to the mixture and the resulting solution was stirred at -20 °C for 1 h. 20 ml of saturated aqueous ammonium chloride solution was then added to quench the reaction. The reaction solution was extracted twice with ethyl acetate, and the combined organic phase was washed with saturated sodium chloride. The organic phase was then dried over anhydrous sodium sulfate, filtered, and concentrated. The resulting oil was purified by silica gel column chromatography (eluent: ethyl acetate / petroleum ether = 4 / 5, V / V) to give the title compound 79A (440 mg, yield: 38.7%).

[0367] Step 2: Synthesis of compound 79B Ethyl 2-((3aS,4R,7R,7aS)-7-(6-chloro-4-cyclopentylmethoxymethylamino)-1H-pyrazolo[3,4-d]pyrimidin-1-yl)-2,2-dimethyltetrahydro-4H-[1,3]dioxacyclo[4,5-c]pyran-4-yl)methoxy)-2-(diethoxyphospho)-3-methoxypropanoate 79A (440 mg, 0.60 mmol) was dissolved in anhydrous ethanol (10 mL). Sodium borohydride (68 mg, 1.8 mmol) and calcium chloride (200 mg, 1.8 mmol) were then added under ice bath, and the resulting solution was stirred at room temperature for 1 hour. Then, 10 mL of saturated aqueous ammonium chloride solution was added to quench the reaction. The combined organic phase was extracted twice with ethyl acetate, washed with saturated brine, dried over anhydrous sodium sulfate, filtered and concentrated under vacuum. The resulting oil was purified by silica gel column chromatography (eluent: methanol / dichloromethane = 1 / 10, V / V) to give the title compound 79B (330 mg, yield: 79.6%).

[0368] Step 3: Synthesis of compound 79C Diethyl 2-(((3aS,4R,7R,7aS)-7-(6-chloro-4-(cyclopentylmethoxymethyl)amino)-1H-pyrazolo[3,4-d]pyrimidin-1-yl)-2,2-dimethyltetrahydro-4H-[1,3]dioxacyclo[4,5-c]pyran-4-yl)methoxy)-1-hydroxy-3-methoxypropan-2-yl)phosphonate 79B (330 mg, 0.48 mmol) and iodomethane (102 mg, 0.71 mmol) were dissolved in anhydrous N,N-dimethylformamide (5 mL). Sodium hydride (28.6 mg, 0.71 mmol, 60% in mineral oil) was then added under ice-cooling and the resulting solution was stirred at room temperature for 1 hour. The reaction was then quenched by the addition of 5 mL of saturated aqueous ammonium chloride. The mixture was extracted twice with ethyl acetate and the combined organic phases were washed with saturated brine, dried over anhydrous sodium sulfate, filtered and concentrated in vacuo to give the crude title compound 79C (300 mg).

[0369] Step 4: Synthesis of compound 79 Diethyl (2-(((3aS, 4R, 7R, 7aS)-7-(6-chloro-4-(cyclopentyl(methoxymethyl)amino)-1H-pyrazolo[3,4-d]pyrimidin-1-yl)-2,2-dimethyltetrahydro-4H-[1,3]dioxo[4,5-c]pyran-4-yl)methoxy)-1,3-dimethoxypropan-2-yl)phosphonate 79C (300 mg, 0.42 mmol) was dissolved in anhydrous acetonitrile (5 mL), and then trimethylsilyl bromide (650 mg, 4.25 mmol) and triethylamine (0.83 mL, 5.95 mmol) were added, and the resulting solution was stirred at room temperature overnight. After the phospholipid hydrolysis was completed, the reaction solution was concentrated under vacuum, and then trifluoroacetic acid (1.5 mL) and water (0.5 mL) were added to the residue, and the resulting solution was stirred at room temperature for 1 h. The reaction solution was concentrated under reduced pressure, and the resulting residue was purified by reverse phase preparation (eluent: acetonitrile / 0.1% trifluoroacetic acid aqueous solution = 25% to 90%, V / V) to obtain the title compound 79 (57 mg, yield: 23.7%). 1 H NMR (400 MHz, CD3OD) δ 8.07 (s, 1H), 4.89 – 4.75 (m,1H), 4.59 – 4.49 (m, 1H), 4.47 – 4.44 (m, 1H), 4.20 – 4.14 (m, 2H), 4.08 –3.96 (m, 4H), 3.83 – 3.77 (m, 4H), 3.40 – 3.31 (m, 6H), 2.14 – 2.04 (m, 2H), 1.85 – 1.78 (m, 2H), 1.77 – 1.57 (m, 1H).

[0370] Biological Assessment

[0371] Test Example 1: In vitro inhibitory activity test of the compounds of the present invention on CD73 enzyme

[0372] 1. Experimental materials are shown in Table 2: Table 2 name supplier Part Number CD73 BPS 71184 AB680 MCE HY-125286 AMP-Glo ​​Kit Promege V5012 Tris-HCl, pH 7.5 Invitrogen 15567027 <![CDATA[MgCl2]]> Sigma 63020 DTT Sigma D0632 BSA Sigma D0632 Tween-20 Sigma P2287 DMSO Sigma 474382 Echo Qualified 384-Well Polypropylene Microplate 2.0,Clear,Flat Bottom LABCYTE 001-14555 White 384-shallow well Microplate PerkinElmer 6008289 .

[0373] 2. Experimental instruments are shown in Table 3: Table 3 Chinese name English name model factory SN Automatic micropipette Precision PRC384U BioTek 205532 Ultrasonic Nanoliter Liquid Handling System Echo 650 Liquid Handler Echo 650 Labcyte E6XX-21146 Centrifuge Centrifuge Avanti J-15R Beckman Coulter JBR20A055 ELISA reader Envision 2104 Multilabel Reader PerkinElmer 1041604 .

[0374] 3. Experimental Methods The experiment used the AMP-Glo ​​Assay kit according to the instructions. Prepare the reaction buffer containing 25mMTrispH7.5, 5mM MgCl2, 2mM DTT, 0.01% BSA and 0.005% Tween-20. Dissolve the test compound in DMSO to prepare a 10 mM stock solution, and dilute it with DMSO to 100 times the final concentration of the reaction in a 384-well Echo plate according to the starting concentration of the experiment. Transfer 50 nL of the compound dilution in the 84-well Echo plate to the 384-well reaction plate using an ultrasonic nanoliter liquid handling system, and set up a DMSO control (positive control) and a control without protein (negative control). Add 2.5 µL of reaction buffer containing CD73 protein (final concentration 0.025 nM) to each reaction well, centrifuge for 1 minute, and incubate the mixture at 37 ºC for 60 minutes. Then, add 2.5 µL of 5'-AMP substrate solution (final concentration 5µM, provided by the AMP-Glo ​​kit) prepared in reaction buffer to each reaction well, centrifuge for 1 minute, and react at 37ºC for 45 minutes. At the same time, prepare a 5'-AMP standard curve (20 µM~0.02 µM), add 5 µL of AMP-Glo ​​reagent 1 containing AB680 (final concentration 400nM) to each reaction well and standard curve well, centrifuge for 1 minute, and incubate at room temperature for 1 hour. Add 10 µL of AMP-Glo ​​reagent 2 to each reaction well and standard curve well to terminate the reaction, centrifuge for 1 minute, and let stand at room temperature for 60 minutes. Read the luminescence values ​​of all samples on the Envision2104 multifunctional reader in a 384-well plate, copy the luminescence value RLU, convert the luminescence reading value to the amount of AMP through the standard curve formula, and convert the amount of AMP to the inhibition percentage through the formula, inhibition rate (%) = (maximum value-sample value) / (maximum value-minimum value)×100%, where the maximum value refers to the reading value of the positive control and the minimum value refers to the reading value of the negative control. Use the formula Y=Bottom + (Top-Bottom) / (1+(IC50 / X)^HillSlope) to fit the curve and obtain IC 50 Value, Y is the inhibition rate, and X is the concentration of the compound.

[0375] 4. The experimental results are shown in Table 4: Table 4 Compound <![CDATA[hCD73 IC 50 (nM)]]> Compound <![CDATA[hCD73 IC 50 (nM)]]> 1 C 41-1 C 2 C 41-2 D 3 A 42 A 4 B 43 B 5 B 44-1 A 6 B 44-2 C 7 C 45-1 A 8 B 45-2 D 9 C 46 C 10 B 47 A 11 B 48 D 12 D 49 B 13 D 50 C 14 D 51 C 15 B 52 D 16 C 55 A 17 C 56 A 18 B 57 A 19 D 58 A 20 A 59 A 21 A 60 A 22 A 61 A 23 A 62 A 24 A 63 C 25 A 64 A 26 A 65 A 27 A 66 A 28 A 67 D 29 A 68 C 30 A 69 A 31 B 70 A 32 A 71 D 33 B 72 D 34 A 73 B 35 A 74 A 36 A 75 A 37 A 76 C 38 A 77 A 39-1 A 78 B 39-2 B 79 A 40-1 C AB680 A 40-2 C

[0376] A: IC50 ≤ 0.1 nM; B:0.1 nM <IC50 ≤ 1 nM C: 1 nM <IC50 ≤ 10 nM D: 10 nM <IC50 ≤ 100 nM The above results show that the compounds of the embodiments of the present invention have strong inhibitory activity against CD73 enzyme.

[0377] Test Example 2: Test of the inhibitory activity of the compounds of the present invention on CD73 on the surface of tumor cells A375 1. Experimental materials are shown in Table 5: Table 5 name factory Batch number / article number A375 Nanjing Kebai DMEM medium CORNING 34021010 FBS gibco 2217479CP Dual Antibody gibco 2321147 PBS Solarbio 20220104 Pancreatin gibco 2310191 Malachite Green Phosphate Detection Kit R&D Systems P318632 HEPES Absin 411A002 NaCl Chongqing Chuandong Chemical Industry Group 20210301 KCl Chengdu Kelong Chemicals Co., Ltd. 2022042501 CaCl2 Chongqing Chuandong Chemical Industry Group 20160501 NaHCO3 Chengdu Kelong Chemicals Co., Ltd. 2022011601 Glucose Absin J02D21 96-well plate YEASEN 84015ES51 .

[0378] 2. Experimental instruments are shown in Table 6 Table 6 name factory Multifunctional ELISA reader Molecular Devices Centrifuge cence Xiangyi Electronic balance SARTORIUS .

[0379] 3. Experimental methods Compound IC50 determination When human melanoma A375 cells were cultured to an appropriate density using complete medium (DMEM medium containing 10% FBS), the cell suspension was plated on a 96-well plate, 100 uL per well, and placed in an incubator at 37°C and 5% CO2 for overnight culture. The supernatant was removed, and the plate was washed once with Buffer (containing HEPES, NaCl, KCl, CaCl2, NaHCO3, Glucose), and then 90 uL of compound solutions of different concentrations prepared with Buffer were added. The blank control was Buffer, and 500 uM AMP solution was added to the reaction wells, 10 uL per well, and incubated in a 37°C incubator for 4 hours. Malachite Green Reagent A solution was added to each well and incubated at room temperature for 10 minutes; then 10 μL of Malachite Green Reagent B was added, and the plate was incubated at room temperature for 60 minutes, and the absorbance value was measured with an enzyme marker.

[0380] The inhibition rate was calculated using the absorbance value, and the concentration and inhibition rate were fitted with a nonlinear regression curve using Graphpad Prism software to obtain the IC 50 value.

[0381] The experimental results are shown in Table 7. Table 7 Compound <![CDATA[A375 IC 50 (nM)]]> Compound <![CDATA[A375 IC 50 (nM)]]> AB680 B 42 A 3 C 47 C 4 C 49 C 5 C 55 B 8 B 57 B 10 B 58 C 11 C 60 B 21 B 61 C 22 C 62 B 23 B 64 B 24 B 66 B 25 B 69 B 27 B 70 A 28 B 74 A 29 A 75 B 30 B 77 C 38 B 78 C 39-1 B 79 B A: IC 50 ≤ 0.1 nM; B:0.1 nM <IC 50 ≤ 1 nM C: 1 nM <IC 50 ≤ 10 nM The IC values ​​of the compounds of the present invention for inhibiting CD73 on the surface of A375 cells were measured by the above test method. 50 The value is less than 10 nM, indicating that the compound of the present invention has a strong inhibitory activity on CD73 on the surface of A375 cells.

[0382] Test Example 3: Test of the inhibitory activity of the compounds of the present invention on CD73 on the surface of tumor cells H1568

[0383] 1. Experimental materials are shown in Table 8: Table 8 name factory Batch number / article number H1568 Wuhan Punosai 1640 medium Yisheng Biology 41402ES76 FBS gibco 2217479CP Dual Antibody gibco 2321147 PBS Solarbio 20220104 Pancreatin gibco 2310191 Malachite Green Phosphate Detection Kit R&D Systems P318632 HEPES Absin 411A002 NaCl Chongqing Chuandong Chemical Industry Group 20210301 KCl Chengdu Kelong Chemicals Co., Ltd. 2022042501 CaCl2 Chongqing Chuandong Chemical Industry Group 20160501 NaHCO3 Chengdu Kelong Chemicals Co., Ltd. 2022011601 Glucose Absin J02D21 96-well plate YEASEN 84015ES51

[0384] 2. Experimental instruments are shown in Table 9 Table 9 name factory Multifunctional ELISA reader Molecular Devices Centrifuge cence Xiangyi Electronic balance SARTORIUS .

[0385] 3. Experimental methods Compound IC50 determination When human non-small cell lung cancer cells H1568 were cultured to an appropriate density using complete medium (1640 medium containing 10% FBS), the cell suspension was plated on a 96-well plate, 100 uL per well, and placed in an incubator at 37°C and 5% CO2 for overnight culture. The supernatant was removed, and washed once with Buffer (containing HEPES, NaCl, KCl, CaCl2, NaHCO3, Glucose), and then 90 uL of compound solutions of different concentrations prepared with Buffer were added. The blank control was Buffer, and 500 uM AMP solution was added to the reaction wells, 10 uL per well, and incubated in a 37°C incubator for 4 hours. Malachite Green Reagent A solution was added to each well and incubated at room temperature for 10 minutes; then 10 μL of Malachite Green Reagent B was added, and incubated at room temperature for 60 minutes, and the absorbance value was measured with an enzyme marker.

[0386] The inhibition rate was calculated using the absorbance value, and the concentration and inhibition rate were fitted with a nonlinear regression curve using Graphpad Prism software to obtain the IC 50 value.

[0387] 4. The experimental results are shown in Table 10: Table 10 Compound <![CDATA[H1568 IC 50 (nM)]]> 38 B 42 B 66 A A: IC 50 ≤ 0.1 nM; B:0.1 nM <IC 50 ≤ 1 nM C: 1 nM <IC50 ≤ 10 nM The IC values ​​of the compounds of the present invention for inhibiting CD73 on the surface of A375 cells were measured by the above test method. 50 The value is less than 10 nM, indicating that the compound of the present invention has a strong inhibitory activity on CD73 on the surface of A375 cells.

[0388] Test Example 4: Antitumor Activity Test of the Compounds of the Invention Female C57 mice (5-6 weeks old) were housed in a specific pyrogen-free room at 20-25°C with 12h:12h light-dark lighting, and they had free access to water and were fed with autoclaved food.

[0389] 100 μL (5×105) of E.G7-OVA cell suspension was injected subcutaneously into the axilla of the forelimb of mice, and the animals were divided into a control group and a treatment group (5 mice / group) by random block method according to the average weight of the mice. The control group was orally administered with solvent twice a day, and the treatment group was orally administered with Example 66 or CB708 twice a day for 15 days. The tumor volume of mice in each group was measured twice a week.

[0390] Table 11 Effect on tumor volume

[0391] Note: CB708 was prepared according to the synthesis method of compound 49 in Journal of Medicinal Chemistry 2023, 66, 1, 345-370.

[0392] The experimental results show that the compounds of the present invention have strong tumor inhibition activity on lymphoma mice and are superior to the clinical drug CB-708 (such as Figure 1 as shown).

[0393] Test Example 5: Antitumor Activity Test of the Compounds of the Invention Female C57 mice (6-7 weeks old) were housed in a specific pyrogen-free room at 20-25°C with 12h:12h light-dark lighting, and they had free access to water and were fed with autoclaved food.

[0394] 100 μL (5×105) E.G7-OVA cell suspension was injected subcutaneously into the axilla of the forelimb of mice, and the animals were divided into a control group and a treatment group (5 mice / group) by random block method according to the average weight of the mice. The control group was orally administered with solvent twice a day, and the treatment group was orally administered with Example 42 or ORIC533 twice a day for 17 days. The tumor volume of mice in each group was measured twice a week.

[0395] Table 12 Effect on tumor volume

[0396] Note: ORIC533 was prepared by referring to the synthesis method of Example 25 in CN114929721A.

[0397] The experimental results show that the compounds of the present invention have strong anti-tumor activity against lymphoma mice and are superior to the clinical drug ORIC533 (such as Figure 2 as shown).

[0398] The above embodiments are only used to help understand the method and core idea of ​​the present invention. It should be noted that, for those skilled in the art, several improvements and modifications can be made to the present invention without departing from the principles of the present invention, and these improvements and modifications also fall within the scope of protection of the claims of the present invention.

Claims

1. A compound having the structure shown in Formula I, or a stereoisomer, pharmaceutically acceptable salt, solvate, cocrystal or deuterated form thereof: ， in, " ” is a single bond or a double bond; R is selected from any of the following structures: or , X1 and X2 are each independently N or C; X3 and X4 are each independently N or CH; X5 is selected from N or CH; R1, R2 are independently selected from hydrogen, C 1-6 Alkyl, C 1-6 The halogenated alkyl, C 1-6 alkyl, cycloalkyl, heterocycloalkyl, aryl, heteroaryl, C 1-6 Alkyl aryl, C 1-6 Alkyl heteroaryl, C 1-6 Alkylcycloalkyl, C 1-6 Alkyl heterocycloalkyl, C 1-6 Acyl aryl, aryl and C 5-6 Cycloalkyl or C 5-6 wherein the above aryl, heteroaryl, heterocycloalkyl and heterocycloaryl may be substituted by one, two or three R 1b or R1 and R2 together with the nitrogen atom to which they are attached form a group optionally substituted by one, two or three R 1a Substituted heterocycloalkyl, heterocycloaryl, C 3-5 The heterocycloalkyl group is a 3- to 5-membered cycloalkyl group; R 1a , R 1b are independently selected from hydrogen, C 1-6 Alkyl, C 3-6 cycloalkyl or halogen; R3 is independently selected from halogen, thiol, C 3-6 The alkenyl group, C 2-6 Alkynylalkyl or C 2-6 wherein the alkyl group may be further substituted with a hydroxyl group; R4 is independently selected from hydrogen or C 1-6 The alkyl group; R 5a , R 5b , R 6a , R 6b are independently selected from hydrogen, halogen, hydroxyl, C 1-3 Alkyl, C 2-3 Alkenyl or C 2-3 The alkynyl group; R7, R8 are independently selected from hydrogen or -(CH2) m -O-(CH2) n R 11 ; m, n are independently selected from 0, 1 or 2; R 11 Independently selected from hydrogen, hydroxy, OCH3, heterocycloalkyl or heteroaryl including 1-4 heteroatoms; R9, R 10 are independently selected from hydrogen or C 1-10 alkyl.

2. The compound according to claim 1, characterized in that In the structure of Formula I, the structural fragment Has any of the following structures: 、 。 3. The compound according to claim 1, characterized in that In the structure of Formula I, the structural fragment Has any of the following structures: 、 、 。 4. The compound according to claim 1, characterized in that The R3 is selected from halogen, thiol, or .

5. The compound according to claim 1, characterized in that The R4 is hydrogen.

6. The compound according to claim 1, characterized in that The R 5a , R 5b , R 6a , R 6b are independently selected from hydrogen, hydroxy, halogen or C 2-3 The alkynyl group.

7. The compound according to claim 1, characterized in that The R7 and R8 are independently selected from hydrogen, , ,or .

8. The compound according to claim 1, characterized in that The R is .

9. The compound according to claim 1 or 8, characterized in that R9, R 10 independently selected from hydrogen or C 1-6 alkyl.

10. The compound according to claim 1, characterized in that Has any of the following structures: Formula I-1; Formula I-2; R1, R2, R3, R4, R 5a , R 5b , R 6a , R 6b , R7, R8, R9, R 10 , X5 is defined as in claim 1.

11. The compound according to claim 10, characterized in that Structure fragment Has any of the following structures: 、 、 。 12. The compound according to claim 10, characterized in that The R 5a , R 5b , R 6a , R 6b are independently selected from hydrogen, hydroxy, halogen or C 2-3 The alkynyl group.

13. The compound according to claim 10, characterized in that The R7 and R8 are independently selected from hydrogen, , ,or .

14. The compound according to claim 10, characterized in that R4, R9, R 10 For hydrogen.

15. The compound according to claim 10, characterized in that The R3 is selected from halogen, thiol, or .

16. The compound according to claim 1, characterized in that Has the following structure: Formula I-3; R1, R2, R3, R7, R8 and X5 are defined as in claim 1.

17. The compound according to claim 16, characterized in that Structure fragment Has any of the following structures: 、 、 。 18. The compound according to claim 16 or 17, characterized in that Structure fragment Has any of the following structures: 、 。 19. The compound according to claim 16, characterized in that R3 is halogen, or .

20. The compound according to claim 16, characterized in that The R7 and R8 are independently selected from , ,or .

21. The compound according to claim 16 or 20, characterized in that Any one of R7 and R8 is selected from .

22. The compound according to claim 1, characterized in that Has any of the following structures:

23. A pharmaceutical composition comprising the compound according to any one of claims 1 to 22, and a pharmaceutically acceptable excipient and / or carrier.

24. The pharmaceutical composition according to claim 23, characterized in that Also included are anti-PD-1 antibodies.

25. Use of the compound according to any one of claims 1 to 22, or the pharmaceutical composition according to any one of claims 23 to 24, in the preparation of a medicament for treating a CD73-mediated disease.

26. The use according to claim 25, characterized in that The CD73-mediated disease is a tumor or an autoimmune disease.

Images