Cd73 inhibitors and their pharmaceutical applications

Abstract

Provided are CD73 (also known as ecto-5'-nucleotidase) inhibitor compounds, and compositions and uses thereof for treating and / or preventing CD73-associated diseases, disorders, and conditions, including cancer-associated and immune-associated disorders. CD73 inhibitor compounds include compounds having a structure according to Formula I': (I') and pharmaceutically acceptable esters or salts thereof.

Description

[0001] Cross-reference to related applications

[0002] This application claims the benefit of priority to Chinese Patent Application No. 202011346141.0, filed November 25, 2020, and U.S. Patent Application No. 17 / 133,348, filed December 23, 2020, which are incorporated herein by reference in their entirety. Technical Field

[0003] This invention relates to compounds and compositions that inhibit CD73 (extracellular 5'-nucleotidase), and their use in treating and / or preventing CD73-related or associated diseases, disorders and conditions (including cancer-related and immune-related disorders). Background Technology

[0004] Extracellular nucleotidases are a group of extracellular enzymes located on the cell surface. Members of the extracellular nucleotidase family include extracellular nucleotide pyrophosphatase / phosphodiesterase (E-NPP), extracellular nucleoside triphosphate diphosphate hydrolase (E-NTPDase), extracellular 5'-nucleotidase (e5NT, also known as CD73), and alkaline phosphatase (AP). These enzymes hydrolyze various extracellular nucleotides into nucleosides, including adenosine. Extracellular nucleotides are important signaling molecules that trigger cellular responses by acting on their respective receptors (e.g., adenosine activates the P1 receptor, and its nucleotides (ADP, ATP) activate the P2 receptor). Adenosine 5'-monophosphate (AMP) is the major substrate of CD73, which is hydrolyzed to adenosine. Adenosine is ubiquitous in the body and is a crucial regulator of purinergic cell signaling, essential for many physiological and pathophysiological processes.

[0005] Extensive data indicate that CD73 possesses enzymatic activity in cancer development and metastasis. CD73 is upregulated in many cancer cell types and tumors, suggesting that CD73 expression is associated with tumor angiogenesis, invasion, and metastasis. The hydrolytic cascade from extracellular ATP to adenosine is an important immunosuppressive regulatory pathway in the tumor microenvironment. CD73 overexpression impairs adaptive antitumor immune responses and promotes tumor growth and metastasis. Extracellular adenosine is also involved in the regulation of adaptive responses to hypoxia. It has been shown that reducing e5NT activity with monoclonal antibodies, siRNA, and small molecule inhibitors, including AMPCP (adenosine [(α,β)-methylene]diphosphate), can attenuate tumor growth and metastasis (see, for example, Zhou et al., Oncol. Rep. 17(2007):1341-1346; Stagg and Smyth, Oncogene, 29(2010):5346-5358). Tumor growth is also impaired in CD73-deficient mice, and these effects have been shown to be primarily attributed to reduced adenosine production in these mice. Therefore, the potential therapeutic applications of CD73 inhibitors for cancer have been actively explored (see, for example, M. al-Rashida et al., Eur. J. Med. Chem., 115(2016): 484-494 and the references cited therein).

[0006] Tumor cells partially overcome antitumor responses through immunosuppressive mechanisms. Several such immunomodulatory mechanisms exist. Among them, adenosine is a key factor produced by cancer cells and immune cells in the tumor microenvironment and used to suppress antitumor responses. Adenosine triphosphate (ATP) is catalyzed by two cell surface proteins, CD73 and CD39, to produce adenosine, and this process is enhanced under metabolic stress conditions, such as tumor hypoxia. Adenosine exerts its immunomodulatory function through four adenosine receptors (AR, referred to as A1, A2A, A2B, and A3, respectively) expressed on various immune cells. Overexpression of adenosine-producing enzymes (such as CD73 and AR) has been associated with tumor development in various cancer types. Since AR signaling increases tumor progression, regulation of this signaling represents a promising therapeutic approach for cancer (MHKazemi, et al., J. Cell. Physiol., 233(2018):2032-2057 and the references cited therein).

[0007] As mentioned above, extracellular nucleotidases are enzymes located on the cell surface that regulate purinergic (or pyrimidinergic) signaling pathways. There are four distinct families of extracellular nucleotidases: extracellular nucleotide triphosphate diphosphate hydrolase (CD39), extracellular nucleotide pyrophosphatase / phosphodiesterase, alkaline phosphatase, and extracellular 5'-nucleotidase (e5NT, also known as CD73). CD73 is a glycosylphosphatidylinositol-anchored zinc metallophosphatase. CD73 catalyzes the dephosphorylation of extracellular adenosine monophosphate (AMP) to generate adenosine. It collaborates with CD39 to form an extracellular enzyme cascade, thereby producing adenosine from ATP. The CD73-catalyzed conversion of AMP to adenosine is considered a major factor contributing to elevated extracellular adenosine levels in the tumor microenvironment (Stagg, J. et al., Proc. Natl. Acad. Sci. USA.:107(2010):1547-1552). CD73 expression is directly upregulated by hypoxia-inducible factor-1α, explaining the increased extracellular adenosine observed in hypoxic malignancies. CD73 is also expressed by regulatory T cells (Tregs) and promotes Treg-mediated immunosuppression (Stagg J, et al., Cancer Res. 71(2011):2892-2900). Furthermore, CD73 is induced by transforming growth factor-β (TGF-β), tumor necrosis factor-α (TNF-α), hepatocyte growth factor (HGF), interleukin-6 (IL-6), mitogen-activated protein kinase (MAPK), signal transduction and transcription activator 3 (STAT3), interleukin-2 (IL-2), retinoic acid, int / wingless (WNT), epithelial-mesenchymal transition, and p53 mutations. CD73 is overexpressed in various tumor types and promotes tumor cell invasion, metastasis, and adhesion. CD73 is also associated with immune tolerance and poor prognosis in cancer. Therefore, CD73 is a promising target for the development of anticancer drugs. In addition, CD73 inhibitors have potential in treating other diseases mediated by adenosine and its receptors (Y.-P. Gong, et al., Expert Opin. Ther. Pat., 28(2018):167-171).

[0008] The adenosine pathway is also considered a major immunosuppressive component in many human tumors (see Whiteside, TL, Expert Rev. Anticancer Ther., 17(2017):527-535 for a review). Adenosine and inosine are key immune checkpoints in cancer. The cooperation between the adenosine and PGE2 pathways in the tumor microenvironment helps suppress anti-tumor immune effector cells. Therefore, targeting the adenosine pathway with drug inhibitors or antibodies is a promising therapeutic strategy for cancer.

[0009] In preclinical in vivo studies, blocking the activity of extracellular nucleotidases or adenosine receptor signaling has successfully inhibited tumor growth and metastasis. Adenosine pathway blockade, alone or in combination with other immunotherapies (including gate inhibitors), is currently in initial Phase I clinical trials in patients with advanced malignancies.

[0010] Small molecule inhibitors of CD73 have been reported. For example, Adams et al. (International PCT Application Publication No. WO2017 / 098421) described substituted benzothiadiazine derivatives (CD73 inhibitors), pharmaceutical compositions thereof, and their use in the treatment of cancer, precancerous syndromes, and diseases associated with CD73 inhibition.

[0011] Debien et al. (International PCT Application Publication No. WO2017 / 120508; US Patent Application Publication No. US2017 / 0267710) disclosed compounds that regulate the conversion of AMP to adenosine by extracellular 5'-nucleotidase, compositions containing such compounds, methods for synthesizing such compounds, and uses of such compounds and compositions for the treatment and / or prevention of a variety of diseases mediated by extracellular 5'-nucleotidase.

[0012] Cacatian et al. (International PCT Application Publication No. WO2015 / 164573) described purine derivatives and pharmaceutical compositions thereof, which are inhibitors of CD73 and can be used to treat cancer.

[0013] Chen et al. (International PCT Application Publication No. WO 2018 / 049145) disclosed the preparation of nucleotides as inhibitors of extracellular nucleotidases and the use of these compounds in the treatment or prevention of cancer. Summary of the Invention

[0014] This invention relates to compounds and compositions comprising compounds that inhibit the activity of extracellular 5'-nucleotidases (also known as e5NT, CD73, NT5E, and 5NT). Inhibition of CD73 enzyme activity leads to inhibition or regulation of extracellular adenosine levels, thereby modulating the physiological environment of cells and tissues.

[0015] This invention also relates to the use of such compounds and compositions in the treatment and / or prevention of diseases, disorders, and conditions wholly or partially mediated by CD73. CD73 inhibitors have been used in the treatment of many diseases, including cancer, fibrosis, neurological and neurodegenerative diseases (e.g., depression and Parkinson's disease), ischemic cardiovascular diseases, immune-related diseases, and diseases with inflammatory components. In specific embodiments, the CD73 inhibitor compounds and compositions described herein can inhibit the immunosuppressive and / or anti-inflammatory activity of CD73, and can be used as a therapeutic or preventative therapy when such inhibition is required.

[0016] In the first general aspect, compounds of formula I' are provided, along with their pharmaceutically acceptable salts or esters:

[0017]

[0018] Where: W is oxygen; X' is -P(=O)(OR)-, where R' is hydrogen; Y is -PO3R'2, where R' is hydrogen; R 1 ' is a hydroxyl group (–OH); R 2 ' is chlorine (–Cl); and R 3 'and R 4 Together with the nitrogen atoms they are attached to, they form monocyclic, bicyclic, tricyclic, spirocyclic, or fused-ring systems, in which the ring system is substituted or unsubstituted.

[0019] In the second general aspect, compounds of formula I are provided, along with their pharmaceutically acceptable salts or esters:

[0020]

[0021] Where: R 1 and R 2 Independently selected from hydrogen, unsubstituted or substituted aryl, unsubstituted or substituted heteroaryl, unsubstituted or substituted 4-8 membered cyclic groups, and unsubstituted or substituted 4-8 membered heterocyclic groups; or, R 1 and R 2 Together with the carbon atoms they are attached to, they form 4-8 membered carbon rings or heterocycles, wherein the ring portion is a monocyclic ring, a ring fused with an aromatic ring, or a ring with a ketone functional group; m and n are independently selected from integers from 0 to 4, provided that the sum of m and n is equal to or greater than 2; when m>1, each R 3 Same or different, and when n>1, each R 4 Same or different; and each R 3 and each R 4 Each is independently selected from hydrogen, halogen, unsubstituted or substituted aryl, unsubstituted or substituted heteroaryl, and 4-8 membered carbon rings or heterocycles; or, when m is 2, 3, or 4, two adjacent Rs are selected from hydrogen, halogen, unsubstituted or substituted aryl, and heteroaryl, and each is selected from 4-8 membered carbon rings or heterocycles; or, when m is 2, 3, or 4, two adjacent Rs are selected from hydrogen, halogen, unsubstituted or substituted aryl, unsubstituted or substituted heteroaryl, and each is selected from 4-8 membered carbon rings or heterocycles; or, when m is 2, 3, or 4, two adjacent Rs are selected from hydrogen, halogen, unsubstituted or substituted aryl, and substituted heteroaryl, and each is selected from 4-8 membered carbon rings or heterocycles; or, when m is 2, 3, or 4, two adjacent Rs are selected from 3 Together with the carbon atoms they are attached to, they form unsubstituted or substituted aromatic rings, and R 4 Other R 3 (If present) independently selected from hydrogen or halogen; or when n is 2, 3, or 4, two adjacent R 4 Together with the carbon atoms they are attached to, they form unsubstituted or substituted aromatic rings, and R 3 Other R 4 (If present) Independently selected from hydrogen or halogen.

[0022] In another general aspect, compounds of formula II are provided, along with their pharmaceutically acceptable salts or esters:

[0023]

[0024] Where: p and q are independent integers from 0 to 3, provided that p and q are not both 0, and when p or q is 0, there is no carbon and no R group attached to it; r, s, and t are independent integers from 0 to 2; R 5 and R 6 Independently selected from H, C1 to C6 alkyl groups, substituted or unsubstituted C4 to C7 cycloalkyl groups, substituted or unsubstituted aryl groups, and unsubstituted or substituted arylalkyl groups; or, R 5 and R 6 Together with the carbon atoms they are attached to, they form unsubstituted or substituted aromatic rings; R 7 R 8 R 9 and R 10 Independently selected from hydrogen, C1 to C6 alkyl, substituted or unsubstituted C4 to C7 cycloalkyl, unsubstituted or substituted aryl, and unsubstituted or substituted arylalkyl; or R 8 and R 9 Together with the carbon atoms they are attached to, they form substituted or unsubstituted carbon rings, or unsubstituted or substituted aromatic rings; or R 7 It forms a carbonyl group together with the carbon it is attached to.

[0025] In another general aspect, compounds of formula III are provided, along with their pharmaceutically acceptable salts or esters:

[0026]

[0027] Where: R 5 Selected from hydrogen, C1 to C6 alkyl groups, unsubstituted or substituted C4 to C7 cycloalkyl groups, unsubstituted or substituted aryl groups, and unsubstituted or substituted arylalkyl groups; p and q are independently integers from 0 to 3, provided that p and q are not simultaneously 0, and when p or q is 0, there is no carbon and no R group attached thereto; and R 7 R 8 R 9 and R 10 Independently selected from hydrogen, C1 to C6 alkyl, substituted or unsubstituted C4 to C7 cycloalkyl, unsubstituted or substituted aryl, and unsubstituted or substituted arylalkyl; or, R 8 and R 9 Together with the carbon atoms they are attached to, they form substituted or unsubstituted carbon rings or unsubstituted or substituted aromatic rings; or R 7 It forms a carbonyl group together with the carbon it is attached to.

[0028] In another general aspect, compounds of formula IV are provided, along with their pharmaceutically acceptable salts or esters:

[0029]

[0030] Wherein: X is selected from hydrogen, halogen, amino, hydroxyl, and C1 to C6 alkyl groups; p and q are independently integers from 0 to 3, provided that p and q are not simultaneously 0, and when p or q is 0, there is no carbon and no R group attached thereto; r is an integer from 0 to 2; and R 7 R 8 R 9 and R 10 Independently selected from hydrogen, C1 to C6 alkyl, substituted or unsubstituted C4 to C7 cycloalkyl, unsubstituted or substituted aryl, and unsubstituted or substituted arylalkyl; or, R 8 and R 9 Together with the carbon atoms they are attached to, they form substituted or unsubstituted carbon rings or unsubstituted or substituted aromatic rings; or R 7 It forms a carbonyl group together with the carbon it is attached to.

[0031] In another general aspect, compounds of formula V are provided, along with their pharmaceutically acceptable salts or esters:

[0032]

[0033] Where: r and s are independently selected from integers from 0 to 2, provided that r and s are not both 0; p and q are independently selected from integers from 0 to 3, provided that p and q are not both 0, and when p or q is 0, there is no carbon and no R group attached to it; R 5 and R 6 Independently selected from hydrogen, C1 to C6 alkyl, substituted or unsubstituted C4 to C7 cycloalkyl, unsubstituted or substituted aryl, and unsubstituted or substituted arylalkyl; or, R 5 and R 6 Together with the carbon atoms they are attached to, they form unsubstituted or substituted aromatic rings; R 7 and R 10 Independently selected from H, C1 to C6 alkyl, substituted or unsubstituted C4 to C7 cycloalkyl, unsubstituted or substituted aryl, unsubstituted or substituted arylalkyl; or R 7 Together with the carbon atom to which it is attached, it forms a carbonyl group; and X is selected from hydrogen, halogen, amino, hydroxyl and C1 to C6 alkyl groups.

[0034] In another general aspect, compounds of formula VI are provided, along with their pharmaceutically acceptable salts or esters:

[0035]

[0036] Where r and s are independent integers from 0 to 2, provided that r and s are not both 0; R 5 and R 6 Independently selected from hydrogen, C1 to C6 alkyl, substituted or unsubstituted C4 to C7 cycloalkyl, unsubstituted or substituted aryl, and unsubstituted or substituted arylalkyl; or, R 5 and R 6 Together with the carbon atoms they are attached to, they form unsubstituted or substituted aromatic rings; R 11 and R 12 Independently selected from hydrogen, unsubstituted or substituted aryl, unsubstituted or substituted heteroaryl, and unsubstituted or substituted 4-8 membered carbon rings or heterocycles; or R 11 and R 12 Together with the carbon atoms they are attached to, they form unsubstituted or substituted 4-8 membered heterocycles.

[0037] In another general aspect, compounds of formula I' are provided, along with their pharmaceutically acceptable salts or esters:

[0038]

[0039] Wherein: W is oxygen, sulfur, nitrogen, or methylene; X' is a moiety selected from phosphoryl (-P(=O)(OR')-), sulfonyl (-S(=O)2-), and carbonyl (-C(=O)-), where R' is hydrogen, an esterifying group, or a protecting group; or X' and W together form –(CR 7 'R 8 ') n Where n is an integer from 0 to 3, and R 7 'and R 8 'Independently selected from hydrogen, halogen, hydroxyl group, and lower alkyl groups having 1 to 4 carbon atoms; Y is selected from phosphonate (-PO3R'2), sulfonate (-SO3R'), and carboxylate (-CO2R'), wherein R' is hydrogen, an esterifying group, or a protecting group; R 1 ' is hydroxyl and hydrogen; R 2 'is chlorine or hydrogen; and R 3 'and R 4 'Independently selected from hydrogen, alkyl, alkenyl and ynyl, wherein R 3 'and R 4 At least one of them has 1 to 30 carbon atoms, for example, but not limited to 1 to 10, 11 to 20, 11 to 30, or 21 to 30 carbon atoms, and wherein when W is O or S, the number of carbon atoms is not 1 to 10 (i.e., when W is O or S, R 3 'and R 4 At least one of them has 11 to 30 carbon atoms.

[0040] In one embodiment of formula I', R 3 'and R 4 'And the nitrogen atoms to which they are attached form heterocyclic systems, which are independently selected from monocyclic, bicyclic, tricyclic, spirocyclic, fused-ring, and bridged-ring systems.

[0041] In one embodiment of formula I', R 3 'and R 4 'Independently selected from hydrogen and cyclic systems, wherein the cyclic system is a carbocyclic system (aromatic or non-aromatic) or heterocyclic system containing monocyclic, bicyclic, tricyclic, spirocyclic, fused, or bridged rings, and wherein the cyclic system is substituted or unsubstituted, provided that R...' 3 'and R 4 'Not both of which are hydrogen, and the condition is that when W is O or S, the ring system is not a monocyclic ring.'

[0042] In another embodiment of formula I', R 3 'is hydrogen or a lower alkyl group (e.g., C10)' 1-6 ), and R 4 'for -C(=O)R 5 '、–C(=O)NHR 5 'Or -C(=O)OR 5 ', where R 5 'For C 1-30 Alkyl, C 2-30 alkenyl or C 2-30 Alkyne group, wherein when W is O or S, C 1-10 and C 2-10 The group is excluded (i.e., when W is O or S, R...). 5 'Is C 11-30 Alkyl, C 11-30 alkenyl or C 11-30 (Alkyne group).

[0043] In some embodiments of formula I', R 3 'is hydrogen or a lower alkyl group, and R 4 'for -C(=O)R 5 '、–C(=O)NHR 5 'Or -C(=O)OR 5 ', where R 5 'A ring system, wherein the ring system is a carbocyclic system (aromatic or non-aromatic) or heterocyclic system containing monocyclic, bicyclic, tricyclic, spirocyclic, fused, or bridged rings, and the carbocyclic or heterocyclic system is substituted or unsubstituted, provided that when W is O or S, R...' 5 The ring system is not a single-ring ring system.

[0044] In some embodiments of formula I', R3 'and R 4 It is not benzyl.

[0045] In some embodiments of Formula I', the bicyclic ring is not a biphenyl ring.

[0046] In some embodiments of formula I', R 3 'and R 4 It is not benzyl, and the bicyclic ring is not a biphenyl ring.

[0047] In one embodiment, compounds of formula II' and / or formula III', and their pharmaceutically acceptable salts or esters, are provided:

[0048]

[0049] Among them, W, X', Y, R 1 '、R 3 'and R 4 As specified above.

[0050] In another embodiment, compounds of formula IV' and their pharmaceutically acceptable salts or esters are provided:

[0051]

[0052] Among them, X', Y, R 1 '、R 2 '、R 3 'and R 4 As specified above.

[0053] In another embodiment, compounds of formula IV'a are provided, as well as pharmaceutically acceptable salts or esters thereof:

[0054]

[0055] Among them, X', Y, R 1 '、R 2 '、R 3 'and R 4 As specified above.

[0056] In another embodiment, the present invention provides compounds of formula IV'b, and pharmaceutically acceptable salts or esters thereof:

[0057]

[0058] Among them, X', Y, R 1 '、R 2 '、R 3 'and R 4 As specified above.

[0059] In another embodiment, the present invention provides compounds of formula IV'c, and pharmaceutically acceptable salts or esters thereof:

[0060]

[0061] Where m' is an integer from 0 to 3, X', Y, R 1 '、R 2 '、R 3 'and R 4 As specified above, R 7 'and R 8 'Independently selected from hydrogen, halogens, hydroxyl groups and lower alkyl groups having 1 to 4 carbon atoms.'

[0062] In yet another embodiment, compounds of formula V' and / or formula VI' are provided, as well as pharmaceutically acceptable salts or esters thereof:

[0063]

[0064] Among them, X', Y, R 1 '、R 3 'and R 4 As specified above.

[0065] In another embodiment, the present invention provides compounds of formula V'a and / or formula VI'a, and pharmaceutically acceptable salts or esters thereof:

[0066]

[0067] Where X', Y, R 1 '、R 2 '、R 3 'and R 4 As specified above.

[0068] In another embodiment, the present invention provides compounds of formula VII' and / or formula VIII', and their pharmaceutically acceptable salts or esters:

[0069]

[0070] Where R' is hydrogen, an esterifying group, or a protecting group; and R 1 '、R 2 '、R 3 'and R 4 As specified above.

[0071] In another embodiment, the present invention provides compounds of formula VII'a and / or formula VIII'a, and their pharmaceutically acceptable salts or esters:

[0072]

[0073] Where R' is hydrogen, an esterifying group, or a protecting group; and R 1 '、R 2 '、R 3 'and R 4 As specified above.

[0074] In yet another embodiment, the present invention provides compounds of formula IX', and pharmaceutically acceptable salts or esters thereof:

[0075]

[0076] Wherein, R' is hydrogen, an esterifying group, or a protecting group; and R 1 '、R 2 '、R 3 'and R 4 As specified above.

[0077] In yet another embodiment, the present invention provides compounds of formula IX'a, and pharmaceutically acceptable salts or esters thereof:

[0078]

[0079] Where: R' is hydrogen, an esterifying group, or a protecting group; and R 1 '、R 2 '、R 3 'and R 4 As specified above.

[0080] In embodiments of formula IX'a, R' is hydrogen, an esterifying group, or a protecting group. 1 ' is hydroxyl or hydrogen; R 2 'is hydrogen or chlorine; and R 3 'is hydrogen or a lower alkyl group, and R 4 'It is an alkyl, alkenyl, or alkynyl group having 1 to 30 carbon atoms; or R 3 'is hydrogen or a lower alkyl group, and R 4 'A substituent for a ring system containing monocyclic, bicyclic, tricyclic, or polycyclic rings, wherein the ring system is a fused ring system, a spirocyclic system, a bridged ring system, or a parallel ring system, and the ring system is a carbocyclic ring, an aliphatic ring, an aromatic ring, a heterocyclic ring, or a combination thereof; or R 3 'is hydrogen or a lower alkyl group, and R 4 'for -C(=O)R 5 '、-C(=O)NHR 5 'Or -C(=O)OR 5 ', where R 5 'It is an alkyl, alkenyl, or alkynyl group having 1 to 30 carbon atoms; or R 3'Hydrogen or lower alkyl groups, and R 4 'for -C(=O)R 5 'or -C(=O)NHR 5 ', where R 5 ' is a substituent for a carbocyclic or heterocyclic system, wherein the carbocyclic or heterocyclic system contains bicyclic, tricyclic, spirocyclic, fused, or bridged rings, wherein the carbocyclic system is aromatic or non-aromatic, and the heterocyclic system is substituted or unsubstituted, wherein the ring is a carbocyclic, aliphatic, aromatic, heterocyclic, or a combination thereof; or R 3 'is hydrogen or a lower alkyl group, and R 4 'is unsubstituted or substituted 1-adamantyl, α-naphthylmethyl or β-naphthylmethyl; or R 3 '、R 4 Together with the nitrogen atoms they are attached to, they form heterocyclic systems that are independently selected from monocyclic, bicyclic, tricyclic, spirocyclic, fused, or bridged rings.

[0081] In one embodiment of formula IX'a, R' is hydrogen, an esterifying group, or a protecting group; R 1 ' is hydroxyl or hydrogen; R 2 'is chlorine or hydrogen; R 3 'is hydrogen or a lower alkyl group; and R 4 ' is a group containing an adamantyl group. In some such embodiments, R 4 'It is a substituted or unsubstituted 1-adamantyl, or a substituted or unsubstituted 2-adamantyl.' In some such embodiments, R 4 'Refractory or unsubstituted 1-adamantylmethyl.' In some such embodiments, R 4 'It is substituted or unsubstituted 1-adamantylethyl, substituted or unsubstituted 1-adamantylpropyl, or substituted or unsubstituted 1-adamantylbutyl.'

[0082] In one embodiment of formula IX'a, R' is hydrogen, an esterifying group, or a protecting group. 1 ' is hydroxyl or hydrogen, R 2 'is chlorine or hydrogen; R 3 'is hydrogen or a lower alkyl group; and R 4 ' is a group containing a naphthyl moiety. In some such embodiments, R 4 'Substituted or unsubstituted α-naphthyl or substituted or unsubstituted β-naphthyl.' In some such embodiments, R 4 'Refers to substituted or unsubstituted α-naphthylmethyl, or substituted or unsubstituted β-naphthylmethyl.' In some such embodiments, R 4 'Selected from substituted or unsubstituted naphthylethyl, substituted or unsubstituted naphthylpropyl, and substituted or unsubstituted naphthylbutyl.'

[0083] In some embodiments of formula IX'a, the fused tricyclic structure is a substituted or unsubstituted carbazole moiety.

[0084] In one implementation, R 1 ' is a hydroxyl group (i.e., the carbohydrate moiety in the compound is a D-ribosyl moiety). In another embodiment, R 1 ' is hydrogen (i.e., the carbohydrate moiety in the compound is the 2-deoxy-D-ribosyl moiety).

[0085] In another embodiment, R 2 ' is hydrogen.' In yet another embodiment, R 2 ' is hydrogen, and R 1 ' is a hydroxyl group (i.e., the compound is an adenosine derivative). In another embodiment, R 2 'For hydrogen, R 1 ' is hydrogen (i.e., the compound is a deoxyadenosine derivative). In another embodiment, R 2 'For hydrogen, R 3 'and R 4 'None of them are hydrogen (i.e., the compound is an adenosine derivative or deoxyadenosine derivative with a substituent on the amino group of the adenine moiety). In another embodiment, R 2 ' is chlorine, and the compound is a 2-chloro-D-adenosine derivative or a 2-chloro-D-deoxyadenosine derivative.

[0086] In some implementations, R 3 'is hydrogen or a lower alkyl group (e.g., C10)' 1-6 ), and R 4 'A group consisting of an alkyl, alkenyl, or alkynyl group having 1 to 30 carbon atoms (i.e., C10, C20, C30, C40, C50, C60, C7 ...70, C70, 1-30 Alkyl, C 2-30 alkenyl or C 2-30 (alkynyl group), and when W is O or S, R 4 It has 11 to 30 carbon atoms. In some embodiments, R 3 'is hydrogen or a lower alkyl group, and R 4 ' is a group containing an adamantyl moiety. R 4 'It can be, for example, substituted or unsubstituted 1-adamantyl, substituted or unsubstituted 2-adamantyl, substituted or unsubstituted 1-adamantylmethyl, substituted or unsubstituted 1-adamantylethyl, substituted or unsubstituted 1-adamantylpropyl, or substituted or unsubstituted 1-adamantylbutyl. In some embodiments, R 3 'is hydrogen or a lower alkyl group, and R 4 ' is a group containing a naphthyl moiety. R 4'It can be, for example, substituted or unsubstituted α-naphthyl, substituted or unsubstituted β-naphthyl, substituted or unsubstituted α-naphthylmethyl, substituted or unsubstituted β-naphthylmethyl, substituted or unsubstituted naphthylethyl, substituted or unsubstituted naphthylpropyl, or substituted or unsubstituted naphthylbutyl.

[0087] In another embodiment, R 3 ' is hydrogen or a lower alkyl group, R 4 'A substituent containing a monocyclic, bicyclic, tricyclic, or polycyclic system, wherein the ring system is a fused ring, spirocyclic, bridged ring, or parallel ring system, and wherein the ring system is a carbocyclic, aliphatic, aromatic, heterocyclic, or a combination thereof.

[0088] In a further embodiment, R 3 ' is hydrogen or a lower alkyl group, R 4 'for -C(=O)R 5 '、-C(=O)NHR 5 'Or -C(=O)OR 5 ', where R 5 'It is an alkyl, alkenyl, or alkynyl group having 1 to 30 carbon atoms, wherein when W is O or S, C1 to C2 is present.' 10 Groups are excluded.

[0089] In some implementations, R 3 'is hydrogen or a lower alkyl group, and R 4 'For C(=O)R 5 '、-C(=O)NHR 5 'Or -C(=O)OR 5 ', where R 5 ' is a substituent containing a monocyclic, bicyclic, tricyclic, or polycyclic system, wherein the ring system is a fused ring, spirocyclic, bridged ring, or parallel ring system, and wherein the ring system is a carbocyclic, aliphatic, aromatic, heterocyclic, or combination thereof, wherein when W is O or S, R 5 It is not a single-ring system.

[0090] In one implementation, R 4 ' is a group containing an adamantyl group. In a further embodiment, R 4 'is a substituted or unsubstituted 1-adamantyl or 2-adamantyl group.' In yet another embodiment, R 4 'Refractory or unsubstituted 1-adamantylmethyl.' In some embodiments, R 4 'It is 1-adamantylethyl, 1-adamantylpropyl or 1-adamantylbutyl, wherein the adamantyl moiety may be substituted or unsubstituted.

[0091] In another implementation, R 4' is a group containing a naphthyl moiety. In a further embodiment, R 4 'Refers to substituted or unsubstituted α-naphthyl or β-naphthyl.' In other embodiments, R 4 'is α-naphthylmethyl or β-naphthylmethyl, with or without further substitution. In yet another embodiment, R 4 ' is selected from naphthylethyl, naphthylpropyl, and naphthylbutyl, wherein the naphthyl moiety may be unsubstituted or substituted.

[0092] In another embodiment, R 3 'and R 4 Together with the nitrogen to which they are attached, they form a tricyclic fused ring system, such as, but not limited to, substituted or unsubstituted carbazoyl moieties.

[0093] In another embodiment, R 3 'and R 4 Together with the nitrogen to which they are attached, they form heterocyclic systems independently selected from monocyclic, bicyclic, tricyclic, spirocyclic, fused, and bridged rings.

[0094] In some embodiments, the present invention provides the compounds listed in Table 1 and their pharmaceutically acceptable salts or esters.

[0095] Table 1. Structures of exemplary compounds

[0096]

[0097]

[0098]

[0099]

[0100]

[0101]

[0102]

[0103]

[0104]

[0105]

[0106] In some embodiments, the compounds in Table 1a and their pharmaceutically acceptable salts or esters are provided.

[0107] Table 1a. Structures of exemplary compounds

[0108]

[0109]

[0110]

[0111]

[0112]

[0113]

[0114]

[0115]

[0116]

[0117]

[0118] In some embodiments, the compounds in Table 1b and their pharmaceutically acceptable salts or esters are provided.

[0119] Table 1b: Structure of exemplary compounds

[0120]

[0121]

[0122]

[0123]

[0124]

[0125]

[0126]

[0127]

[0128]

[0129]

[0130] In some embodiments, the compounds in Table 1c and their pharmaceutically acceptable salts or esters are provided.

[0131] Table 1c: Structure of exemplary compounds

[0132]

[0133]

[0134]

[0135]

[0136]

[0137]

[0138]

[0139]

[0140]

[0141]

[0142]

[0143]

[0144]

[0145] In some embodiments, the compounds in Table 1d and their pharmaceutically acceptable salts or esters are provided.

[0146] Table 1d. Structures of exemplary compounds

[0147]

[0148]

[0149]

[0150]

[0151] In some embodiments, compounds are provided in which the C, H, O, and N atoms, as described herein, are each independently selected from atoms of natural abundance and isotopically enriched atoms. Examples of isotopically enriched atoms include, but are not limited to, for carbon, 12 C 13 C and 14 C; For hydrogen, 1 H, 2 H and 3 H; for oxygen, 16 O、 17 O and 18 O; for nitrogen, 14 N and 15 N.

[0152] In another general aspect, pharmaceutical compositions are also provided comprising: a compound as defined herein or a pharmaceutically acceptable salt or ester thereof, and a pharmaceutically acceptable carrier. In some embodiments, a pharmaceutical composition is provided comprising a compound of any one of formulas I to VI, I' to IX', IV'a, IV'b, IV'c, V'a, VI'a, VII'a, or VIII'a, or a pharmaceutically acceptable salt or ester thereof, and a pharmaceutically acceptable carrier. In some embodiments, a pharmaceutical composition is provided comprising a compound of any one of formulas I to VI, or a pharmaceutically acceptable salt or ester thereof, and a pharmaceutically acceptable carrier. In some embodiments, a pharmaceutical composition is provided comprising a compound of any one of formulas I' to IX', or a pharmaceutically acceptable salt or ester thereof, wherein, in the compound, when W is O or S, R... 3 'and R 4 One of them is not hydrogen or C1 to C. 10 The compounds are alkyl, alkenyl, or alkynyl. In some embodiments, pharmaceutical compositions are provided comprising compounds shown in Tables 1, 1a, 1b, 1c, and 1d, or pharmaceutically acceptable salts or esters thereof, and pharmaceutically acceptable carriers. In some embodiments, pharmaceutically acceptable carriers include creams, emulsions, gels, liposomes, or nanoparticles.

[0153] In some embodiments, the pharmaceutical composition is suitable for oral administration. In some such embodiments, the composition is in the form of hard-shell gelatin capsules, soft-shell gelatin capsules, flat capsules, pills, tablets, lozenges, powders, granules, pellets, pastures, or sugar-coated pills. In some embodiments, the composition is in the form of a solution, an aqueous liquid suspension, a non-aqueous liquid suspension, an oil-in-water liquid emulsion, an oil-in-water liquid emulsion, an elixir, or a syrup. In some embodiments, the composition has an enteric coating. In some embodiments, the composition is formulated for controlled release.

[0154] In some embodiments, the pharmaceutical composition is injectable.

[0155] In some embodiments, the pharmaceutically acceptable carrier further comprises at least one additional therapeutic agent, such as, but not limited to, a chemotherapeutic agent, an immunomodulatory and / or inflammatory modulator, an anti-hypercholesterol agent, an anti-infective agent, or an immune checkpoint inhibitor. In some embodiments, the at least one additional therapeutic agent is an immune checkpoint inhibitor. Non-limiting examples of immune checkpoint inhibitors include ipilimumab, nivolumab, and pembrolizumab.

[0156] In another general aspect, compounds, compositions, and methods for inhibiting CD73 activity in subjects of need are provided, including administering an effective amount of the compounds and / or pharmaceutical compositions described herein to the subject.

[0157] In specific embodiments, the compounds of this invention function to inhibit CD73 immunosuppressive activity and / or anti-inflammatory activity, and are used as therapeutic or prophylactic therapies when such inhibition is required. Unless otherwise stated, it should be understood in the description of the uses of the compounds of this invention that these compounds may be in the form of compositions (e.g., pharmaceutical compositions). As used herein, the terms “CD73 inhibitor,” “CD73 blocker,” “extracellular 5'-nucleotidase inhibitor of adenosine,” “NT5E inhibitor,” “5NT inhibitor,” and all other acceptable terms in the relevant fields are used interchangeably to refer to compounds that can directly or indirectly inhibit the CD73 receptor in in vitro tests, in vivo models, and / or other tests demonstrating CD73 inhibition and potential therapeutic or prophylactic efficacy. The term also refers to compounds that exhibit at least some therapeutic or prophylactic benefit in human subjects.

[0158] Although the compounds of the present invention are thought to act by inhibiting CD73, the implementation of the present invention does not require a precise understanding of the potential mechanism of action of the compounds. For example, the compounds may also act at least partially by modulating (e.g., inhibiting) other components of purinergic signaling pathways (e.g., CD39). The purinergic signaling system consists of transport proteins, enzymes, and (primarily) receptors for the synthesis, release, action, and extracellular inactivation of ATP and its extracellular breakdown product adenosine. Because inhibition of CD73 reduces adenosine production, CD73 inhibitors can be used to treat adenosine-mediated diseases or disorders, and they act on adenosine receptors (including A1, A2A, A2B, and A3).

[0159] In this invention, the described purinergic signaling process comprises the following components. The first component consists of purinergic receptors (P1, P2X, and P2Y), which are membrane receptors mediating various physiological functions as responses to ATP or adenosine release (e.g., relaxation of intestinal smooth muscle); generally, all cells possess the ability to release nucleotides into the extracellular environment by regulating exocytosis. The second component is a nucleoside transporter (NT), a membrane transporter that transports nucleoside substrates (e.g., adenosine) across the cell membrane; the extracellular concentration of adenosine can be regulated by NTs, possibly linking receptor signaling to transporter function in a feedback loop. As previously mentioned, the nucleotides released into the extracellular environment by extracellular nucleotidases (CD73 and CD39) also comprise additional components.

[0160] In some embodiments, the present invention provides a method for treating or preventing cancer in a subject (e.g., a human), comprising administering to the subject a therapeutically effective amount of at least one CD73 inhibitor compound or composition described herein. In some embodiments of such methods, at least one CD73 inhibitor compound or composition is administered to the subject in an amount that effectively reverses, slows, or prevents the progression of CD73-mediated immunosuppression. In some embodiments, CD73-mediated immunosuppression is mediated via antigen-presenting cells (APCs).

[0161] There are no particular limitations on the types of cancers or tumors that can be treated or prevented using the compounds and compositions described in this invention. Examples of cancers and tumors that can be treated or prevented using the compounds and compositions described in this invention include, but are not limited to: cancers of the prostate, colorectal, pancreas, cervix, stomach, endometrium, brain, liver, bladder, ovary, testis, head, neck, skin (including melanoma and basal carcinoma), mesothelioma, leukocytes (including lymphoma and leukemia), esophagus, breast, muscle, connective tissue, lung (including small cell lung cancer and non-small cell lung cancer), adrenal gland, thyroid, kidney, or bone, glioma, mesothelioma, renal cell carcinoma, gastric cancer, sarcoma, choriocarcinoma, basal cell carcinoma of the skin, and seminoma of the testis. In some embodiments of this invention, the cancer is melanoma, colon cancer, pancreatic cancer, breast cancer, prostate cancer, lung cancer, leukemia, brain tumor, lymphoma, sarcoma, ovarian cancer, or Kaposi's sarcoma.

[0162] In some embodiments, the present invention provides a method for treating subjects who have received bone marrow transplantation or peripheral blood stem cell transplantation, comprising administering a sufficient therapeutically effective amount of a CD73 inhibitor compound or composition to increase delayed-type hypersensitivity to tumor antigens, delay the time of recurrence of malignant tumors after transplantation, increase the recurrence-free survival rate after transplantation, and / or increase the long-term survival rate after transplantation.

[0163] In some embodiments, the present invention provides a method for treating or preventing an infectious condition (e.g., a viral infection) in a subject (e.g., a human), comprising administering to the subject a therapeutically effective amount of at least one CD73 inhibitor compound or composition of the present invention. In some embodiments, the infectious condition is a viral infection (e.g., a chronic viral infection), a bacterial infection, a fungal infection, or a parasitic infection. In some embodiments, the viral infection is human immunodeficiency virus or cytomegalovirus.

[0164] In other embodiments, the present invention provides methods for treating and / or preventing immune-related diseases, disorders and conditions, diseases with inflammatory components, and related disorders using at least one CD73 inhibitor compound or composition provided by the present invention.

[0165] Other diseases, disorders, and conditions that can be fully or partially treated or prevented by inhibiting CD73 activity are also candidate indications for the CD73 inhibitor compounds and compositions provided in this invention.

[0166] In some embodiments, the invention further provides the use of the CD73 inhibitor compounds and compositions described herein in combination with one or more additional agents. These additional agents may have some CD73-modulating activity and / or they may act through different mechanisms of action. In some embodiments, such agents comprise radiation (e.g., local or total radiotherapy) and / or other forms of treatment of a non-pharmacological nature. When using combination therapy, the CD73 inhibitor and an additional agent may be in the form of a single composition or multiple compositions, and the treatment may be administered simultaneously, sequentially, or through some other regimen. For example, in some embodiments, a treatment regimen is provided that follows a radiotherapy phase with a chemotherapy phase. Combination therapies may have additive or synergistic effects.

[0167] In some embodiments, the present invention provides the use of the CD73 inhibitor compounds or compositions described herein in combination with bone marrow transplantation, peripheral blood stem cell transplantation or other types of transplantation therapy.

[0168] In specific embodiments, this invention provides the use of a combination of the CD73 function inhibitor and an immune checkpoint inhibitor as described herein. It has been shown that blocking immune checkpoints (which lead to the amplification of antigen-specific T cell responses) is a promising approach in the treatment of human cancer. Non-limiting examples of immune checkpoints (ligands and receptors) include PD1 (programmed cell death protein 1), PDL1 (PD1 ligand), BTLA (B and T lymphocyte attenuator), CTLA4 (cytotoxic T lymphocyte-associated antigen 4), TIM3 (T cell membrane protein 3), LAG3 (lymphocyte activation gene 3), A2aR (adenosine A2a receptor A2aR), and cytotoxic inhibitory receptors; some of these immune checkpoints are selectively upregulated in various types of tumor cells and are candidates for blockade. Non-limiting examples of immune checkpoint inhibitors include ipilimumab, nivolumab, and pembrolizumab.

[0169] In other embodiments, the present invention provides a method of treating a subject's cancer, comprising administering to the subject a therapeutically effective amount of at least one CD73 inhibitor compound or a combination thereof and at least one chemotherapeutic agent, said chemotherapeutic agent including but not limited to: alkylating agents (e.g., aziridines, such as aziridine chlorate, cyclophosphamide, isofluramide, dichloromethyldiethylamine, melphalan, and uracil aziridine); aziridines, such as thiotepa; methanesulfonates, such as busulfan; nucleoside analogs, such as gemcitabine; nitrosoureas, such as... Carmustine, lomustine, and streptozotocin; topoisomerase 1 inhibitors, such as irinotecan; platinum complexes, such as cisplatin and carboplatin; bioreducing alkylating agents, such as mitomycin, procarbazine, dacarbazine, and hexamethylmelamine; DNA strand breaking agents (e.g., bleomycin); topoisomerase II inhibitors (e.g., acridine, daunorubicin, idarubicin, mitoxantrone, ruboxine, etoposide, and teniposide); DNA minor groove binding agents (e.g., Plicamydin); antimetabolites (e.g., folic acid antagonists). Antidotes, such as methotrexate and trimethoprim; pyrimidine antagonists, such as fluorouracil, fluorodeoxyuridine, CB3717, azacitidine, cytarabine, and fluorouridine; purine antagonists, such as mercaptopurine, 6-thioguanine, fludarabine, and pentostatin; asparaginases; and ribonucleotide reductase inhibitors, such as hydroxyurea; microtubule interactors (e.g., vincristine, estradiol, vinblastine, docetaxel, epormycin derivatives, and paclitaxel); hormones (e.g., estrogens; conjugated estrogens; ethinylestradiol); Diethylstilbestrol; chlormedroxyprogesterone; medroxyprogesterone; progestins, such as hydroxyprogesterone caproate, medroxyprogesterone acetate, and medroxyprogesterone acetate; and androgens, such as testosterone, testosterone propionate, flumethasone, and methyltestosterone; corticosteroids (e.g., prednisone; dexamethasone; methylprednisolone and prednisolone); luteinizing hormone-releasing hormone (LH-REM) or gonadotropin-releasing hormone (GnRH) antagonists (e.g., leuprorelin acetate and goserelin acetate); and anti-hormone antigens (e.g., tamoxifen, anti-androgens such as flutamide; and adrenaline-releasing agents such as mitotane and aminoglutethimide). This invention also provides the use of CD73 inhibitors in combination with other agents known in the art (e.g., arsenic trioxide) and other chemotherapeutic agents that may be developed in the future.

[0170] In some embodiments of methods relating to treating cancer, cancer survival rates observed when a therapeutically effective amount of a CD73 inhibitor is administered in combination with at least one chemotherapy agent are higher than those observed when either agent is administered alone. In other embodiments of methods relating to treating cancer, tumor size reduction or slowing of tumor growth observed when a therapeutically effective amount of a CD73 inhibitor is administered in combination with at least one chemotherapy agent are superior to those observed when either agent is administered alone.

[0171] In a further embodiment, the present invention provides a method for treating or preventing cancer in a subject, comprising administering to the subject a therapeutically effective amount of at least one CD73 inhibitor compound or composition, and at least one signal transduction inhibitor (STI). In a specific embodiment, the at least one STI is selected from the group consisting of bcr / abl kinase inhibitors, epidermal growth factor (EGF) receptor inhibitors, HER-2 / neu receptor inhibitors, and farnesyltransferase inhibitors (FTI).

[0172] In other embodiments, the present invention provides a method for enhancing the rejection of tumor cells in a subject, comprising the combined administration of a CD73 inhibitor compound or composition with at least one chemotherapeutic agent and / or radiotherapy, wherein the resulting rejection of tumor cells is superior to that obtained by administering a CD73 inhibitor, chemotherapeutic agent or radiotherapy alone.

[0173] In a further embodiment, the present invention provides a method for treating cancer in a subject, comprising administering to the subject a therapeutically effective amount of at least one CD73 inhibitor and at least one immunomodulatory agent other than a CD73 inhibitor. It should be understood that, as used herein, "CD73 inhibitor" refers to compounds provided by the present invention, such as compounds of any one of formulas I to VI, I' to IX', IV'a, IV'b, IV'c, V'a, VI'a, VII'a, or VIII'a, or compounds of any one of Tables 1, 1a, 1b, 1c, and 1d, or pharmaceutically acceptable salts or esters thereof, and pharmaceutical compositions thereof.

[0174] In some embodiments, the present invention provides a method for treating or preventing CD73-related diseases, dysregulations, or conditions in subjects in need, comprising administering to the subject a therapeutically effective amount of at least one CD73 inhibitor or a pharmaceutical composition thereof, thereby treating or preventing CD73-related diseases, dysregulations, or conditions in the subject. In some embodiments, the compound is administered in an amount that effectively reverses, slows, or stops CD73-mediated immunosuppression in the subject.

[0175] In some embodiments, CD73-related diseases, disorders, or conditions are cancers, such as, but not limited to, cancers of the prostate, colon, rectum, pancreas, cervix, stomach, endometrium, brain, liver, bladder, ovary, testis, head, neck, skin, mesothelial membrane, leukocytes, esophagus, breast, muscle, connective tissue, lung, adrenal gland, thyroid, kidney, or bone. In some embodiments, the cancer is glioblastoma, mesothelioma, renal cell carcinoma, gastric cancer, sarcoma, choriocarcinoma, basal cell carcinoma of the skin, or seminoma of the testis. In some embodiments, the cancer is melanoma, colon cancer, pancreatic cancer, breast cancer, prostate cancer, lung cancer, leukemia, brain tumor, lymphoma, ovarian cancer, or Kaposi's sarcoma.

[0176] In some implementations, the CD73-related diseases, disorders, or conditions are immune-related diseases, disorders, or conditions selected from rheumatoid arthritis, renal failure, lupus, asthma, psoriasis, colitis, pancreatitis, allergies, fibrosis, anemia, fibromyalgia, Alzheimer's disease, congestive heart failure, stroke, aortic stenosis, arteriosclerosis, osteoporosis, Parkinson's disease, infection, Crohn's disease, ulcerative colitis, allergic contact dermatitis, eczema, systemic sclerosis, and multiple sclerosis.

[0177] In some embodiments, the methods provided herein further include administering at least one additional therapeutic agent to the subject. This additional therapeutic agent may be administered simultaneously or sequentially with the compounds or compositions described herein. In some embodiments, the at least one additional therapeutic agent is a chemotherapeutic agent, an immunomodulatory and / or inflammatory modulator, an anti-hypercholesterol agent, or an anti-infective agent. In one embodiment, the at least one additional therapeutic agent is an immune checkpoint inhibitor, such as, but not limited to, ipilimumab, nivolumab, or pembrolizumab.

[0178] In some embodiments, the present invention provides a method for treating or preventing an infectious condition (e.g., viral infection) in a subject (e.g., a human), comprising administering to the subject a therapeutically effective amount of at least one CD73 inhibitor and a therapeutically effective amount of an anti-infective agent (e.g., one or more antimicrobial agents).

[0179] In another embodiment, treatment of the infectious disease is achieved through the combined administration of a vaccine and a therapeutically effective dose of the CD73 inhibitor provided by the present invention. In some embodiments, the vaccine is an antiviral vaccine, including, for example, an anti-HIV vaccine. In other embodiments, the vaccine is effective against tuberculosis or malaria. In other embodiments, the vaccine is a tumor vaccine (e.g., a vaccine effective against melanoma); the tumor vaccine may comprise genetically modified tumor cells or genetically modified cell lines, including genetically modified tumor cells or genetically modified cell lines that have been transfected to express granulocyte-macrophage stimulating factor (GM-CSF). In a specific embodiment, the vaccine comprises one or more immunogenic peptides and / or dendritic cells.

[0180] In some embodiments involving the treatment of infection by administering the CD73 inhibitor provided by the present invention and at least one other therapeutic agent, the symptoms of infection observed after administration of both the CD73 inhibitor and the other therapeutic agent are improved compared to the same symptoms of infection observed when either the CD73 inhibitor or the other therapeutic agent is administered alone. In some embodiments, the observed symptoms of infection may be a decrease in viral load, an increase in CD4+ T cell count, a reduction in opportunistic infections, an increase in survival time, eradication of chronic infections, or a combination thereof.

[0181] In some embodiments, the present invention provides a method of treating a subject's cancer, comprising administering to the subject an effective amount of the compound or composition described herein and an immune checkpoint inhibitor, thereby treating the subject's cancer. The compound or composition described herein and the immune checkpoint inhibitor may be administered in combination or sequentially. The compound or composition may be administered after or before the administration of the immune checkpoint inhibitor. In some embodiments, the compound or composition and / or the immune checkpoint inhibitor may be administered before, concurrently with, or after other anticancer treatments, such as, but not limited to, radiotherapy. In some embodiments, the immune checkpoint inhibitor is selected from ipilimumab, nivolumab, and pembrolizumab.

[0182] In another general aspect, the invention also provides kits comprising the compounds or compositions described herein. The kits may further comprise buffers or excipients, and / or instructions for use. In some embodiments, the kits also include at least one additional therapeutic agent, such as, but not limited to, chemotherapeutic agents, immunomodulators and / or inflammatory modulators, anti-hypercholesterol agents, anti-infective agents, or immune checkpoint inhibitors. Attached Figure Description

[0183] To better understand the present invention and more clearly demonstrate how to implement it, features of embodiments according to the present invention are now illustrated by way of example and with reference to the accompanying drawings, wherein:

[0184] Figure 1The CD73 inhibition rate curve for compound 9 (% inhibition vs. Log[Conc.] / nM);

[0185] Figure 2 The CD73 inhibition rate curve of compound 22;

[0186] Figure 3 The CD73 inhibition rate curve of compound d-1; and

[0187] Figure 4 The curve shows the CD73 inhibition rate of compound a. Detailed Implementation

[0188] The number of people diagnosed with and dying from cancer continues to rise. Traditional treatments, including chemotherapy and radiation therapy, are often unbearable for patients and become less effective as cancers (such as tumors) evolve to evade them. Recent experimental evidence suggests that CD73 inhibitors may represent an important new treatment approach for cancers such as breast cancer.

[0189] Promising data also suggest that CD73 function inhibitors possess anti-inflammatory and / or immunosuppressive activity against CD73, thus making them potential treatments for immunosuppressive diseases such as HIV and AIDS. CD73 inhibition may also be an important therapeutic strategy for subjects with neurological or neuropsychiatric disorders or dysregulations (e.g., depression).

[0190] This invention particularly provides small molecule compounds having CD73 inhibitory activity and compositions thereof, as well as methods for treating and preventing the diseases, disorders, and conditions described herein using the compounds and compositions. The compounds provided by this invention can be used as inhibitors of CD73 and therefore can be used to treat diseases, disorders, and conditions in which CD73 activity is present. Additionally, the compounds provided by this invention can be used as inhibitors of adenosine receptors (e.g., A2A receptors). Therefore, the compounds provided by this invention can be used to treat diseases, disorders, and conditions associated with the activity of one or more adenosine receptors.

[0191] In one embodiment, the present invention provides a method for treating a subject (e.g., a human) with cancer or a CD73-mediated disorder, comprising administering to the subject a therapeutically effective amount of a CD73 inhibitor provided by the present invention, such as a compound provided by the present invention or a pharmaceutically acceptable composition thereof.

[0192] It should be understood that the pharmaceutical composition comprises the compounds disclosed in this invention (or pharmaceutically acceptable salts or esters thereof) and pharmaceutically acceptable carriers, additives, or solvents. In some embodiments, the amount of the compound in the composition is such that it is effective as a CD73 inhibitor in biological samples (e.g., in vitro assays, in vivo models, etc.) or in subjects. In some embodiments, the composition is formulated for administration to a subject who requires such a composition. In some embodiments, the composition is an injectable formulation. In other embodiments, the composition is formulated for oral administration to a subject.

[0193] This invention also provides a method for treating subjects (e.g., humans) with cancer or adenosine receptor (e.g., A2AR)-mediated dysregulation, comprising administering to the subject a therapeutically effective amount of a CD73 inhibitor provided by this invention, such as a compound provided by this invention or a pharmaceutically acceptable composition thereof. In some embodiments, the amount of the compound in the composition is such that it is effective as an inhibitor of adenosine receptors (e.g., A2AR) in biological samples (e.g., in vitro assays, in vivo models, etc.) or in subjects. In some embodiments, the composition is formulated for administration to a subject who requires such a composition. In some embodiments, the composition is an injectable formulation. In other embodiments, the composition is formulated for oral administration to a subject. In some embodiments, the composition is in the form of hard-shell gelatin capsules, soft-shell gelatin capsules, capsules, pills, tablets, lozenges, powders, granules, pellets, flavored lozenges, or sugar-coated pills. In some embodiments, the composition is in the form of a solution, an aqueous liquid suspension, a non-aqueous liquid suspension, an oil-in-water liquid emulsion, an oil-in-water liquid emulsion, an elixir, or a syrup. In some embodiments, the composition is enteric-coated. In some embodiments, the composition is formulated for controlled release.

[0194] In another embodiment, the present invention provides a method for treating or preventing cancer in a subject, comprising administering to the subject a therapeutically effective amount of at least one CD73 inhibitor and at least one signal transduction inhibitor (STI). In one specific embodiment, the at least one STI is selected from the group consisting of bcr / abl kinase inhibitors, epidermal growth factor (EGF) receptor inhibitors, HER-2 / neu receptor inhibitors, and farnesyltransferase inhibitors (FTI). The present invention also provides a method for enhancing rejection of tumor cells in a subject, comprising co-administering a CD73 inhibitor with at least one chemotherapy agent and / or radiotherapy, wherein the resulting rejection of tumor cells is superior to that obtained by administering a CD73 inhibitor, chemotherapy agent, or radiotherapy alone. In a further embodiment, the present invention provides a method for treating cancer in a subject, comprising administering to the subject a therapeutically effective amount of at least one CD73 inhibitor and at least one immunomodulatory agent different from a CD73 inhibitor.

[0195] In other embodiments, the present invention provides a method for treating or preventing an infectious condition (e.g., viral infection) in a subject (e.g., a human), comprising administering to the subject a therapeutically effective amount of at least one CD73 inhibitor and a therapeutically effective amount of an anti-infective agent (e.g., one or more antimicrobial agents).

[0196] In another embodiment, treatment of the infectious disease is achieved through the combined administration of a vaccine and a therapeutically effective dose of the CD73 inhibitor provided herein. In some embodiments, the vaccine is an antiviral vaccine, including, for example, an anti-HIV vaccine. In other embodiments, the vaccine is effective against tuberculosis or malaria. In yet another embodiment, the vaccine is an oncology vaccine (e.g., a vaccine effective against melanoma); the oncology vaccine may comprise genetically modified tumor cells or genetically modified cell lines, including genetically modified tumor cells or genetically modified cell lines that have been transfected to express granulocyte-macrophage stimulating factor (GM-CSF). In a specific embodiment, the vaccine comprises one or more immunogenic peptides and / or dendritic cells.

[0197] In some embodiments involving the treatment of infection by administration of a CD73 inhibitor and at least one other therapeutic agent, the symptoms of infection observed after administration of both the CD73 inhibitor and the other therapeutic agent are improved compared to the same symptoms of infection observed when either the CD73 inhibitor or the other therapeutic agent is administered alone. In some embodiments, the observed symptoms of infection may be a decrease in viral load, an increase in CD4+ T cell count, a decrease in opportunistic infections, an increase in survival time, eradication of chronic infection, or a combination thereof.

[0198] definition

[0199] To provide a clear and consistent understanding of the terminology used in this specification, some definitions are provided below. Furthermore, unless otherwise specified, all technical and scientific terms used in this invention have the same meaning as commonly understood by one of ordinary skill in the art to which this invention pertains.

[0200] When used in conjunction with the term "comprising" in the claims and / or description, the word "a (a or an)" can mean "one / type," but it is also known to have the meanings of "one / type or more / types," "at least one / type," and "one / type or more than one / type." Similarly, the word "another / type" can mean at least a second / type or many / types.

[0201] As used in this specification and claims, the terms “comprising” (and any form of “comprising”, such as “comprise” and “comprises”), “having” (and any form of “having”, such as “have” and “has”), and “containing” (and any form of “containing”, such as “contain” and “contains”) are inclusive and open-ended and do not exclude additional unlisted elements or processing steps.

[0202] The term “about” is used to indicate that the value includes inherent error variations introduced by the instruments and methods used in determining the value.

[0203] The term "derivative" as used in this invention should be understood as another compound that is structurally similar to the compound but differs in some fine structures.

[0204] This specification involves many chemical terms and abbreviations used by those skilled in the art. However, for clarity and consistency, definitions of selected terms are provided.

[0205] As used in this invention, the term "alkyl" refers to a saturated hydrocarbon having 1 to 30 carbon atoms, including straight-chain, branched, and cyclic alkyl groups. Examples of alkyl groups include, but are not limited to, methyl, ethyl, propyl, butyl, pentyl, hexyl, heptyl, octyl, nonyl, decyl, isopropyl, tert-butyl, sec-butyl, isobutyl, cyclopropyl, cyclopentyl, cyclohexyl, cycloheptyl, cyclooctyl, etc. The term alkyl includes unsubstituted alkyl groups and substituted alkyl groups. The term "C1 to C3" refers to... nAlkyl (where n is an integer from 2 to 30) refers to an alkyl group having 1 to the indicated "n" carbon atoms. Alkyl residues can be substituted or unsubstituted. In some embodiments, for example, the alkyl group can be substituted with groups such as hydroxyl, amino, carboxyl, carboxylic acid ester, amide, carbamate, or aminoalkyl. In some specific embodiments, "alkyl" is modified by a series of carbon atom numbers, thus the size of the alkyl group is specifically defined. For example, C 11 -C 30 Alkyl means an alkyl group containing at least 11 carbon atoms and no more than 30 carbon atoms.

[0206] As used in this invention, the term "acyclic" refers to an organic moiety that does not have a ring system. The term "aliphatic group" includes organic moieties characterized by straight or branched chains, typically having 1 to 15 carbon atoms. Aliphatic groups include acyclic alkyl, alkenyl, and alkynyl groups.

[0207] As used in this invention, the term "alkenyl" refers to an unsaturated hydrocarbon having 2 to 30 carbon atoms, including straight-chain, branched, and cyclic non-aromatic alkenyl groups, and containing 1 to 6 carbon-carbon double bonds. Examples of alkenyl groups include, but are not limited to, vinyl, allyl, 1-propen-2-yl, 1-buten-3-yl, 1-buten-4-yl, 2-buten-4-yl, 1-penten-5-yl, 1,3-pentadien-5-yl, cyclopentenyl, cyclohexenyl, ethylcyclopentenyl, ethylcyclohexenyl, etc. The term alkenyl includes both unsubstituted and substituted alkenyl groups. The term "C2 to C3" refers to... n "Alkenyl" and "C" 2-n The term "alkenyl" (where n is an integer from 3 to 30) is used interchangeably and refers to an alkenyl group having 2 to the number of carbon atoms indicated by "n". In some specific embodiments, "alkenyl" is modified by a series of carbon atom numbers, thus the size of the alkenyl group is specifically defined. For example, C 11 -C 30 Alkenyl groups are defined as alkenyl groups containing at least 11 carbon atoms and no more than 30 carbon atoms.

[0208] As used in this invention, the term "alkynyl" refers to an unsaturated hydrocarbon having 2 to 30 carbon atoms, including straight-chain, branched, and cyclic non-aromatic alkynyl groups, and containing 1 to 6 carbon-carbon triple bonds. Examples of alkynyl groups include, but are not limited to, ethynyl, 1-propyn-3-yl, 1-butyn-4-yl, 2-butyn-4-yl, 1-pentyn-5-yl, 1,3-pentadiyn-5-yl, etc. The term alkynyl includes both unsubstituted and substituted alkynyl groups. The term "C2-C" is also used. n "Alkyne" and "C" 2-n The term "alkynyl" (where n is an integer from 3 to 30) is used interchangeably and refers to an alkynyl group having 2 to the number of carbon atoms indicated by "n". In some specific embodiments, "alkynyl" is modified by a series of carbon atom numbers, thus the size of the alkynyl group is specifically defined. For example, C 11-C 30 The alkynyl group refers to an alkynyl group containing at least 11 carbon atoms and no more than 30 carbon atoms.

[0209] Unless otherwise specified, the term “lower aliphatic,” “lower alkyl,” “lower alkenyl,” and “lower alkynyl” as used herein means that the moiety has at least one (or at least two for alkenyl and alkynyl) and equal to or less than six carbon atoms.

[0210] The terms "cycloalkyl," "alicyclic," "carbocyclic," and equivalent expressions refer to groups comprising saturated or partially unsaturated carbocyclic systems of monocyclic, spirocyclic (sharing a single atom), or fused (sharing at least one bond) carbon rings having 3 to 15 carbon atoms. Examples of cycloalkyl groups include, but are not limited to, cyclopropyl, cyclobutyl, cyclopentyl, cyclopenten-1-yl, cyclopenten-2-yl, cyclopenten-3-yl, cyclohexyl, cyclohexen-1-yl, cyclohexen-2-yl, cyclohexen-3-yl, cycloheptyl, bicyclo[4,3,0]nonyl, norbornyl, etc. The term cycloalkyl includes both unsubstituted and substituted cycloalkyl groups. The term "C3 to C4" refers to... n "Cycloalkyl" and "C" 3-n The term "cycloalkyl" (where n is an integer from 4 to 15) is used interchangeably and refers to a cycloalkyl group having 3 to the number of carbon atoms indicated by "n" in its ring structure. Unless otherwise specified, the term "lower cycloalkyl" as used in this invention refers to a group having at least 3 and equal to or less than 8 carbon atoms in its ring structure.

[0211] Cyclic alkyl residues may be saturated or contain one or more double bonds within the ring system. Specifically, they may be saturated or contain a single double bond within the ring system. In unsaturated cyclic alkyl residues, the double bond can be present in any suitable position. Monocyclic alkyl residues include, for example, cyclopropyl, cyclobutyl, cyclopentyl, cyclopentenyl, cyclohexyl, cyclohexenyl, cycloheptyl, cycloheptenyl, cyclooctyl, cyclononyl, cyclodecyl, cycloundecyl, cyclododecyl, or cyclotetradecyl, which may also be substituted with a C10. 1-4 Alkyl groups. Examples of substituted cycloalkyl residues are 4-methylcyclohexyl and 2,3-dimethylcyclopentyl. Examples of parent structures for bicyclic systems are norbornene, bicyclo[2.2.1]heptane, bicyclo[2.2.2]octane, and bicyclo[3.2.1]octane.

[0212] The term "heterocyclic alkyl" and its equivalents refer to a group containing a saturated or partially unsaturated carbocyclic ring in a monocyclic, spirocyclic (sharing a single atom), or fused-ring (sharing at least one bond) carbocyclic system, having 3 to 30 carbon atoms, including 1 to 6 heteroatoms (e.g., N, O, S, P) or heteroatom-containing groups (e.g., NH, NRx (Rx is alkyl, acyl, aryl, heteroaryl, or cycloalkyl), PO2, SO, SO2, etc.). Where possible, the heterocyclic alkyl group may be linked to a C atom or to a heteroatom (e.g., via a nitrogen atom). Examples of heterocyclic alkyl groups include, but are not limited to, pyrrolidinyl, tetrahydrofuranyl, tetrahydrodithienyl, tetrahydropyranyl, tetrahydrothiaranyl, piperidinyl, morpholinyl, thiomorpholinyl, thiaxyl, piperazine, aziridine, oxacyclobutane, thioheterobutane, thioheterobutane, high-piperidinyl, oxacyclopentane, thioheteropentane, oxazaphenyl, diazaphenyl, thiazolidinyl, 1,2,3,6-tetrahydropyridinyl, and 2-pyrrolidinyl. Phosphoryl, 3-pyrrololinyl, indololinyl, 2H-pyranyl, 4H-pyranyl, dioxanecycloyl, 1,3-dioxolanecycloyl, pyrazolinyl, dithiaalkyl, dithioheterocyclic pentylyl, dihydropyranyl, dihydrothiophenyl, dihydrofuranyl, pyrazolyl, imidazolinyl, imidazolinyl, 3-azabicyclo[3,1,0]hexyl, 3-azabicyclo[4,1,0]heptyl, 3H-indolyl, quinazinyl, and sugars, etc. The term heterocyclic alkyl includes unsubstituted and substituted heterocyclic alkyl groups. The term "C3-C" is used in conjunction with the preceding text. n Heterocyclic alkyl groups and C 3- "n-heterocyclic alkyl", where n is an integer from 4 to 30, and is used interchangeably to denote a heterocyclic alkyl having 3 to the number of atoms shown in the "n" in its ring structure, including at least one heterogroup or atom as defined above. Unless otherwise stated, "low heterocyclic alkyl" as used in this invention refers to having at least 3 and equal to or less than 8 carbon atoms in its ring structure.

[0213] The terms "aryl" and "aryl ring" refer to an aromatic group having "4n+2" (π) electrons and 6 to 14 ring atoms in a conjugated monocyclic or polycyclic system (fused or unfused), where n is an integer from 1 to 7. Polycyclic systems include at least one aromatic ring. Aryl groups can be directly linked or linked via C1-C6 alkyl groups (also called arylalkyl or aralkyl groups). Examples of aryl groups include, but are not limited to, phenyl, benzyl, phenethyl, 1-phenylethyl, tolyl, naphthyl, biphenyl, terphenyl, indene, benzocyclooctenyl, benzocycloheptenyl, azulel, acenaphthel, fluorenyl, phenanthrene, anthracene, etc. The term aryl includes both unsubstituted and substituted aryl groups. The term "C6-C..." n "Aryl" or "C" 6-nThe aryl group (where n is an integer from 6 to 30) is used interchangeably to denote an aryl group having 6 to the number of carbon atoms shown in the "n" in a ring structure, including at least one heterocyclic group or atom as defined above.

[0214] The terms "heteroaryl" and "heteroaryl ring" refer to an aromatic group having "4n+2" (π) electrons and 5 to 14 ring atoms in a conjugated monocyclic or polycyclic system (fused or unfused), where n is an integer from 1 to 7, and includes 1 to 6 heteroatoms (e.g., N, O, S) or groups including heteroatoms (e.g., NH, NRx (Rx is alkyl, acyl, aryl, heteroaryl, or cycloalkyl), SO, etc.). Polycyclic systems include at least one heteroaryl ring. Heteroaryl groups can be directly attached or linked via C1-C3 alkyl groups (also called heteroarylalkyl or heteroarylalkyl). Where possible, heteroaryl groups can be linked to carbon atoms or to heteroatoms (e.g., via nitrogen atoms). Examples of heteroaryl groups include, but are not limited to, pyridinyl, imidazolyl, pyrimidinyl, pyrazolyl, triazolyl, tetrazolyl, furanyl, thiophene; isoxazolyl, thiazolyl, oxazolyl, isothiazolyl, pyrrolidinyl, quinolinyl, isoquinolinyl, indoleyl, isoindoleyl, chromenyl, isocherenyl, benzimidazolyl, benzofuranyl, cenolinyl, indazoleyl, indazinyl, phthalazinyl, pyridazinyl, pyrazinyl, triazinyl, isoindoleyl Dolyl, pteridinyl, furanyl, benzofuranyl, benzothiophenyl, benzothiopheneyl, benzothiazolyl, benzoxazolinyl, quinolinyl, quinolinoneyl, isoquinolinoneyl, quinoxalinyl, naphthidyl, furan-pyridyl, carbazoleyl, phenanthridineyl, acridineyl, peryleneyl, phenanthrolinel, phenazinyl, phenothiazinyl, phenotoxazinyl, dibenzofuranyl, etc. The term heteroaryl includes both unsubstituted and substituted heteroaryl groups. The term "C5-C" is also used. n "Hybrid aryl" and "C" 5-n "Heteroaryl", where n is an integer from 6 to 29, used interchangeably to denote a heteroaryl having from 5 to the number of atoms shown in the "n" in a ring structure, including at least one heterocyclic group or atom as defined above.

[0215] The term "heterocyclic" or "heterocyclic" includes heterocyclic alkyl and heteroaryl groups. Examples of heterocycles include, but are not limited to, acridine, acridine, benzimidazolyl, benzofuranyl, benzothiofuranyl, benzothiophene, benzoxazolyl, benzothiazolyl, benzotriazolyl, benzotetrazole, benzoisoxazolyl, benzoisothiazolyl, benzimidazolinyl, carbazolyl 4αH-carbazolyl, carbazolyl, benzopyranyl (chromanyol), chromenyl, cinnamyl, decahydroquinolinyl, 2H,6H-1,5,2-dithiazinyl, dihydrofurano[2,3-b]tetrahydrofuran, furanyl, furazolyl, and imidazole. Alkyl, imidazolinyl, imidazolyl, 1H-indazole, indolenyl, dihydroindole, indolizinyl, indole, 3H-indole, isobenzofuranyl, isobenzopyranyl, isoindole, isoindolelinyl, isoindole, isoquinolinyl, isothiazolyl, isoxazolyl, methylenedioxyphenyl, morpholinyl, naphthidyl, octahydroisoquinolinyl, oxadiazolyl, 1,2,3-oxadiazolyl, 1,2,4-oxadiazolyl, 1,2,5-oxadiazolyl, 1,3,4- Oxadiazolyl, oxazolylalkyl, oxazolyl, oxazolylalkyl, pyrimidinyl, phenanthridine, phenanthrolinel, phenazinyl, phenothiazinyl, phenotoxazinyl, phthalazinyl, piperazinyl, piperidinyl, piperidinoneyl, 4-piperidinoneyl, piperinyl, pteridinyl, purine, pyranyl, pyrazinyl, pyrazolylalkyl, pyrazolinyl, pyrazolyl, pyridazinyl, pyridoxazole, pyridinium-imidazazole, pyridoxazole, pyridinyl, pyridyl, pyridyl, pyrimidinyl, pyrrolylalkyl, pyrrololinyl, 2H-pyrrolyl, pyrrolyl, quinazolinyl, quinolinyl 4H-quinazinyl, quinoxalinyl, quininecycloyl, tetrahydrofuranyl, tetrahydroisoquinolinyl, tetrahydroquinolinyl, tetrazolyl, 6H-1,2,5-thiadiazinyl, 1,2,3-thiadiazolyl, 1,2,4-thiadiazolyl, 1,2,5-thiadiazolyl, 1,3,4-thiadiazolyl, thianyl, thiazolyl, thienyl, thienothiazolyl, thienooxazolyl, thienoimidazolyl, thienyl, triazinyl, 1,2,3-triazolyl, 1,2,4-triazolyl, 1,2,5-triazolyl, 3,4-triazolyl, xanthonyl, etc. The term heterocyclic includes both unsubstituted and substituted heterocyclic groups.

[0216] As used in this invention, the term "amine" or "amino" refers to unsubstituted or substituted -NR. a R b The fragment, in which R a and R b Each is independently hydrogen, alkyl, aryl, or heterocyclic, or R a and R bTogether with the nitrogen atoms to which they are attached, they form heterocycles. The term amino refers to a compound or segment in which a nitrogen atom is covalently bonded to at least one carbon or heteroatom. Therefore, as used herein, the terms "alkylamino" and "dialkylamino" refer to amino groups having one and at least two C1-C6 alkyl groups attached thereto, respectively. The terms "arylamino" and "diarylamino" respectively include groups in which a nitrogen atom is bonded to at least one or two aryl groups. The terms "amide" or "aminocarbonyl" include compounds or segments containing a nitrogen atom structure bonded to a carbonyl or thiocarbonyl group. The term "acylamino" refers to an amino group directly bonded to an acyl group as defined herein.

[0217] The term "bicyclic" or "bicyclic" refers to a cyclic system with two rings sharing two carbon atoms, which can be located anywhere along either ring. The term typically refers to bicyclic hydrocarbon groups, bicyclic aromatic carbon ring structures, and saturated or partially unsaturated bicyclic carbon ring structures, wherein one or more ring carbon members have been substituted with heteroatoms such as O, S, or N atoms, allowing for structural stability. Bicyclic systems can be fused ring systems, such as bicyclic [4.4.0]decane or naphthalene, or bridged ring systems, such as bicyclic [2.2.2]octane.

[0218] The term "tricyclic" or "tricyclic" refers to a cyclic system with three rings sharing three carbon atoms, which are located anywhere along each ring. The term generally refers to tricyclic hydrocarbon groups, tricyclic aromatic carbon atom ring structures, and saturated or partially unsaturated tricyclic carbon atom ring structures, wherein one or more ring carbon atom members have been substituted with heteroatoms such as O, S, or N atoms, which are permissible for structural stability. Tricyclic systems can be fused rings, such as anthracene or tetradecahydroanthracene, or bridged rings, such as those in adamantane or tricyclic [3.3.1.1]decane.

[0219] The terms "multi-cycle," "multicyclic," "multi-cyclic," or "multi-cyclic" refer to a cyclic system with more than three rings sharing a common carbon atom, located at any position along any ring. This term typically refers to polycyclic hydrocarbon groups, polycyclic aromatic carbon ring structures, and saturated or partially unsaturated polycyclic carbon ring structures, wherein one or more carbon ring members have been replaced by heteroatoms, such as O, S, or N atoms, as permitted by structural stability.

[0220] The term "fused ring" or "fused polycyclic" refers to a polycyclic system containing a fused ring. Typically, a fused ring system comprises 2 or 3 rings, and / or up to 18 ring atoms. As mentioned above, cycloalkyl, aryl, and heterocyclic groups can form fused ring systems. Therefore, fused ring systems can be aromatic, partially aromatic, or non-aromatic and can contain heteroatoms. By this definition, spirocyclic systems are not fused polycyclic, but the fused polycyclic systems of the present invention can themselves have a spirocycle attached to a single ring atom of the system. Examples of fused ring systems include, but are not limited to, naphthyl (e.g., 2-naphthyl), indenyl, phenanthryl, anthraceneyl, pyrene, benzimidazole, benzothiazole, etc. The terms "fused ring" or "fused ring" are used interchangeably herein.

[0221] The term "spirocyclic" or "spirocyclic" refers to an organic compound exhibiting a twisted structure of two or more rings (cyclic systems), wherein two or three rings are linked together by a common atom. Spirocyclic compounds can be fully carbocyclic (all-carbon), such as spiro[5.5]undecane, or heterocyclic (having one or more non-carbon atoms), including but not limited to carbocyclic spirocyclic compounds, heterocyclic spirocyclic compounds, and polycyclic spirocyclic compounds.

[0222] The term "bridged ring" or "bridged" refers to a carbon ring or heterocyclic portion in which two or more atoms are shared in two or more ring structures, wherein the shared atoms are C, N, S, or other heteroatoms arranged in a chemically rationally substituted pattern. Alternatively, a "bridged" compound also refers to a carbon ring or heterocyclic structure in which an atom at any position on the main ring is bonded to a second atom on the main ring by a chemical bond or an atom other than a bond, and which does not actually constitute part of the main ring structure. The first and second atoms may be adjacent to each other or not adjacent in the main ring. The following are specific, non-limiting examples of bridged ring structures considered herein. Other carbon ring or heterocyclic bridged ring structures are also contemplated, including bridged rings in which the bridging atoms are C or other heteroatoms arranged in a chemically rationally substituted pattern, as known in the art.

[0223] The term "nitro" refers to -NO2; the terms "halogen" and "halogen" refer to bromine, chlorine, fluorine, or iodine substituents; the terms "thiol," "thio group," or "mercapto" refer to SH; and the term "hydroxyl group" or "hydroxyl group" refers to -OH. The term "alkathio" refers to an alkyl group having a mercapto group attached thereto. Suitable alkathio groups include groups having 1 to about 12 carbon atoms, preferably 1 to about 6 carbon atoms. As used in this invention, the term "alkylcarboxyl" refers to an alkyl group having a carboxyl group attached thereto.

[0224] As used in this invention, the term "alkoxy" or "lower alkoxy" refers to an alkyl group having an oxygen atom attached thereto. Representative alkoxy groups include those having 1 to 6 carbon atoms, such as methoxy, ethoxy, propoxy, tert-butoxy, etc. Examples of alkoxy groups include, but are not limited to, methoxy, ethoxy, isopropoxy, propoxy, butoxy, pentoxy, fluoromethoxy, difluoromethoxy, trifluoromethoxy, chloromethoxy, dichloromethoxy, trichloromethoxy, etc. The term "alkoxy" includes unsubstituted or substituted alkoxy groups, as well as perhaloalkoxy groups.

[0225] The terms "carbonyl" or "carboxyl" refer to compounds and segments containing a carbon atom bonded to an oxygen atom via a double bond. Examples of carbonyl-containing segments include aldehydes, ketones, carboxylic acids, amides, esters, and acid anhydrides.

[0226] The term "acyl" is derived from the carbon atom of the acyl group and the hydrogen atom (i.e., formyl group), an aliphatic group (C1-C1). 29 Alkyl, C1-C 29 Alkenyl, C1-C 29 A carbonyl group is attached to an alkynyl group, such as an acetyl group, a cycloalkyl group (C3-C8 cycloalkyl group), a heterocyclic group (C3-C8 heterocyclic alkyl group and C5-C6 heteroaryl group), and an aryl group (C6 aryl group, such as benzoyl group). The acyl group can be unsubstituted or substituted (e.g., salicylyl group).

[0227] It should be understood that the terms "substituted" or "substituted" include the implicit condition that such substitution, with changes in the valence of the substituent atom and the substituent, results in a stable compound (e.g., the compound cannot spontaneously undergo processes such as rearrangement, cyclization, or elimination). As used herein, the term "substituted" includes all permissible substituents in organic compounds. In a broad sense, permissible substituents include acyclic and cyclic, branched and unbranched, carbocyclic and heterocyclic, aromatic and non-aromatic substituents in organic compounds. Permissible substituents may be one or more. The term "substituted" refers to substituents such as acyl, amino (including simple amino, monoalkylamino and dialkylamino, monoaryl and diarylamino, and alkylarylamino), acylamino (including carbamoyl and urea), alkylcarbonyloxy, arylcarbonyloxy, alkoxycarbonyloxy, alkoxycarbonyl, carboxy, carboxylate, aminocarbonyl, and monoalkylaminocarbonyl. The substituents include alkyl and dialkylaminocarbonyl, cyano, azide, halogen, hydroxyl, nitro, trifluoromethyl, thio, alkylthio, arylthio, alkylthiocarbonyl, thiocarboxylic acid ester, lower alkyl, lower alkenyl, lower alkynyl, cycloalkyl, heterocycloalkyl, aryl, heteroaryl, lower alkoxy, aryloxy, aryloxycarbonyloxy, benzyloxy, benzyl, sulfinyl, alkylsulfinyl, sulfonyl, sulfate, sulfonate, sulfonamide, phosphate, phosphonate, phosphoryl, oxo, guanidine, imino, formyl, etc. If permitted, any of the above substituents may be further substituted, for example, by alkyl, aryl, or other groups.

[0228] The term "solvent" refers to the physical association of a compound with one or more solvent molecules (whether organic or inorganic). This physical association includes hydrogen bonding. In some cases, solvates can be separated, such as when one or more solvent molecules are incorporated into the crystal lattice. "Solvate" includes both a solution phase and a separable solvate. Exemplary solvates include, but are not limited to, hydrates, ethanolates, methanolates, semi-ethanolates, etc.

[0229] A “pharmaceutically acceptable salt” of a compound refers to a salt of a pharmaceutically acceptable compound. Ideally, a salt of a compound should retain or improve the bioavailability and properties of the parent compound as defined herein, or be beneficial to the inherent basicity, acidity, or charged functionality of the molecule, which would otherwise be biologically undesirable. Examples of pharmaceutically acceptable salts may be those mentioned by Berge et al. in “Pharmaceutical Salts”, J. Pharm. Sci. 66, 1-19 (1977). Non-limiting examples of such salts include, but are not limited to:

[0230] (1) An acid addition salt is formed on a basic or positively charged functional group by adding an inorganic acid or an organic acid. The inorganic acids that can be added include hydrochloric acid, hydrobromic acid, hydroiodic acid, sulfuric acid, aminosulfonic acid, nitric acid, phosphoric acid, carbonate forming agent, etc. Organic acids that can be added include acetic acid, propionic acid, lactic acid, oxalic acid, glycolic acid, neopentanoic acid, tert-butylacetic acid, β-hydroxybutyric acid, valeric acid, hexanoic acid, cyclopentanepropionic acid, pyruvic acid, malonic acid, succinic acid, malic acid, fumaric acid, tartaric acid, citric acid, benzoic acid, 3-(4-hydroxybenzoyl)benzoic acid, cinnamic acid, mandelic acid, methanesulfonic acid, ethanesulfonic acid, 1,2-ethanedisulfonic acid, 2-hydroxyethanesulfonic acid, cyclohexylaminosulfonic acid, benzenesulfonic acid, sulfanilic acid, 4-chlorobenzenesulfonic acid, 2-naphthalenesulfonic acid, 4-toluenesulfonic acid, camphorsulfonic acid, 3-phenylpropionic acid, laurylsulfonic acid, laurylsulfonic acid, oleic acid, palmitic acid, stearic acid, lauric acid, palmitic acid, and purine. (acids), pantothenic acid, lactobionic acid, alginic acid, galactoside, galacturonic acid, gluconic acid, glucoheponic acid, glutamic acid, naphthoic acid, hydroxynaphthoic acid, salicylic acid, ascorbic acid, stearic acid, mucoconic acid, etc.

[0231] (2) An acidic proton present in the parent compound is replaced by a metal ion or forms a base addition salt by coordination with an organic base, wherein the metal ion includes alkali metal ions (e.g., lithium, sodium, potassium), alkaline earth metal ions (magnesium, calcium, barium) or other metal ions such as aluminum, zinc, iron, etc.; the organic base is such as ammonia, ethylamine, diethylamine, ethylenediamine, N,N'-dibenzylethylenediamine, ethanolamine, diethanolamine, triethanolamine, aminobutanetriol, N-methylglucosamine, piperazine, chloroprocaine, procaine, choline, lysine, etc.

[0232] Pharmaceutically acceptable salts can be synthesized from parent compounds containing basic or acidic moieties using conventional chemical methods. Typically, such salts are prepared by reacting a compound in its free acid or basic form with an equistoichiometric amount of a suitable base or acid in water, an organic solvent, or a mixture of both. Salts can be prepared in situ during the final separation or purification of the compound, or independently by reacting a compound in its free acid or basic form with the desired corresponding base or acid, followed by separation of the resulting salt. The term "pharmaceutically acceptable salt" also includes zwitterionic compounds containing cationic groups covalently bonded to anionic groups; these are referred to as "internal salts." It should be understood that all acids, salts, bases, and other ionic and nonionic forms of the compounds of this invention are covered within the scope of this invention. For example, if the compound in this invention is an acid, the salt form of that compound is also covered within the scope of this invention. Similarly, if the compound in this invention is a salt, the acidic and / or basic forms of that compound are also covered within the scope of this invention.

[0233] The compounds provided by this invention may contain non-natural proportions of atomic isotopes at one or more atoms constituting such compounds. Non-natural proportions of isotopes can be defined as amounts ranging from those found in nature to those comprising 100% of the atoms in question. For example, the compounds may incorporate radioactive isotopes, such as tritium (…). 3 H), Iodine-125 125 I) or carbon-14 ( 14 C) or non-radioactive isotopes such as deuterium ( 2 H) or carbon-13 ( 13 C). Such isotopic variations can provide additional applications for those described elsewhere in this application. For example, isotopic variants of the compounds of the present invention can find additional uses, including but not limited to, as diagnostic and / or imaging agents, or as therapeutic agents for cytotoxicity / radiotoxicity. Additionally, isotopic variants can have altered pharmacokinetic and pharmacodynamic characteristics, which can contribute to enhanced safety, tolerability, or efficacy during treatment. All isotopic variants of the compounds provided in this invention, whether radioactive or not, are covered herein.

[0234] Isotope enrichment is an elemental form in which the relative abundance of one particular isotope is increased (i.e., enhanced) while other isotopes are reduced or depleted, by altering the relative abundance of isotopes of a given element. As used herein, a "compound or derivative enriched" refers to a compound in which one or more specific isotopic forms of one or more particular isotopes are increased, i.e., one or more specific isotopic elements are enriched (i.e., enhanced). Typically, in an isotope-enriched compound or derivative, a specific isotopic element at a specific position in the compound is increased. However, it should be understood that a compound may have two or more isotopic elements increased. Furthermore, an isotope-enriched compound may be a mixture of isotopic enrichments of more than one isotope, more than one element, or both. As used herein, an "isotope-enriched" compound or derivative has an isotopic form level higher than the natural abundance of that form. The level of isotopic enrichment will vary depending on the natural abundance of a particular isotopic form. In some embodiments, the level of isotopic enrichment of the compound or the level of isotopic enrichment of elements in the compound can be from about 2 to about 100 molar percentages (%), for example, about 2%, about 5%, about 17%, about 30%, about 51%, about 83%, about 90%, about 95%, about 96%, about 97%, about 98%, and greater than about 98%, about 99%, or 100%.

[0235] In this invention, the terms "naturally abundant element" or "naturally abundant atom" refer to the element or atom with the most abundant atomic mass in nature. For example, the naturally abundant hydrogen is... 1 H (protium), the nitrogen species with natural abundance is14 N, naturally abundant oxygen is 16 O, naturally abundant carbon is 12 C. "Non-isotopically enriched" compounds are compounds in which all atoms or elements are naturally abundant isotopes, meaning that all atoms or elements have the most abundant atomic mass in nature.

[0236] The terms “patient” and “subject” are used interchangeably in this document to refer to human or non-human animals (e.g., mammals).

[0237] When applied to, for example, a subject, cells, tissue, organ, or biological fluid, the terms "application," "applicationer," etc., refer to contacting, for example, a CD73 inhibitor, a pharmaceutical composition including the CD73 inhibitor, or a diagnostic agent with the subject, cells, tissue, organ, or biological fluid. In the case of cells, application includes contacting the reagent with the cells (e.g., in vitro or ex vivo) and contacting the reagent with a fluid, wherein the fluid contacts the cells.

[0238] The terms "treat," "treating," and "treatment" refer to actions (e.g., administration of CD73 inhibitors or pharmaceutical compositions containing them) initiated after a disease, disorder, or condition or its symptoms have been diagnosed or observed, in order to temporarily or permanently eliminate, reduce, suppress, alleviate, or improve at least one underlying cause of the disease, disorder, or condition affecting the subject, or symptoms associated with the disease, disorder, or condition affecting the subject. Therefore, treatment includes suppressing (e.g., preventing the development or further progression of a disease, disorder, or condition or its associated clinical symptoms) an active disease.

[0239] As used in this article, the term "need for treatment" refers to a judgment made by a physician or other caregiver that a subject needs or will benefit from treatment. This judgment is based on a variety of factors within the physician's or caregiver's professional field.

[0240] The terms “prevent,” “preventing,” and “prevention” generally refer to actions taken in a certain way (e.g., administering a CD73 inhibitor or a pharmaceutical composition containing it) in a subject who is susceptible to a particular disease, disorder, or condition, thereby temporarily or permanently preventing, suppressing, inhibiting, or reducing the subject’s risk of developing the disease, disorder, or condition (e.g., determined by the absence of clinical symptoms) or delaying its onset. In some cases, the term also refers to slowing the progression of a disease, disorder, or condition or inhibiting its development into a harmful or other undesirable state.

[0241] As used in this article, "need for prevention" refers to a judgment made by a physician or other caregiver that a subject needs or will benefit from preventive care. This judgment is based on a variety of factors within the physician's or caregiver's professional field.

[0242] The terms “therapeutic effective amount” and “effective amount” are used interchangeably herein and refer to an amount of a pharmaceutical agent, alone or as part of a pharmaceutical composition, administered to a subject, either as a single dose or as part of a series of doses, that produces any detectable positive effect on any symptom, aspect, or characteristic of a disease, disorder, or condition. Therapeutic effective amounts can be determined by measuring the associated physiological effects and can be adjusted according to the dosing regimen and diagnostic analysis of the subject’s condition. For example, measuring the serum level of a CD73 inhibitor (or, for example, its metabolites) at a specific time after administration can indicate whether a therapeutic effective amount has been used. In some embodiments, the terms “therapeutic effective amount” and “effective amount” refer to the amount or dose of a therapeutic agent, such as a compound, that provides the desired therapeutic, diagnostic, or prognostic effect in a subject after administration of a single or multiple doses. An effective amount can be readily determined by an attending physician or diagnostician using known techniques and by observing results obtained in similar circumstances. When determining the effective amount or dose of a compound to be administered, many factors are considered, including but not limited to: the size, age, and general health condition of the subject; the specific disease involved; the extent or severity of the disease or condition to be treated; the individual response of the subject; the specific compound administered; the administration method; the bioavailability characteristics of the administered formulation; the chosen dosing regimen; the use of concomitant medications; and other relevant considerations.

[0243] The term "substantially pure" is used herein to mean that a component constitutes more than about 50% of the total content of the composition, and typically more than about 60% of the total content. More typically, "substantially pure" means that the target component constitutes at least 75%, at least 85%, at least 90% or more of the total composition. In some cases, the component of interest will constitute more than about 90% or more of the total content of the composition.

[0244] As used herein, the terms “CD73-related disease, dysregulation, or condition” and “CD73-mediated disease, dysregulation, or condition” are used interchangeably to refer to any disease, dysregulation, or condition that may benefit from treatment with a CD73 inhibitor. Generally, CD73-related or mediated diseases, dysregulations, and conditions are those in which CD73 activity plays a biological, mechanistic, or pathological role. Such diseases, dysregulations, and conditions may also be associated with the activity of one or more adenosine receptors. Non-limiting examples of CD73-related diseases, dysregulations, and conditions include tumor-related dysregulations (cancer, tumors, etc.), immune-related dysregulations, inflammatory component conditions, microbiome-related conditions, CNS-related conditions and neurological conditions, and other diseases (e.g., but not limited to, cardiovascular diseases, gastrointestinal diseases, metabolic diseases, liver diseases, lung diseases, ophthalmic diseases, and kidney diseases).

[0245] For example, CD73 inhibitors can be used to prevent or treat proliferative disorders, cancer, or tumors; increase or enhance immune responses; improve immunization, including enhancing vaccine efficacy; and increase inflammation. The CD73 inhibitors disclosed in this invention can be used to treat immunodeficiency diseases associated with immunodeficiency, immunosuppressive drug therapy, acute and / or chronic infections, and aging. CD73 inhibitors can also be used to stimulate the immune system of patients with iatrogenic induced immunosuppression, including those who have undergone bone marrow transplantation, chemotherapy, or radiation therapy. In other embodiments, CD73 inhibitors can be used to treat or prevent any viral, bacterial, fungal, parasitic, or other infectious diseases, disorders, or conditions, including but not limited to HIV and AIDS.

[0246] In some implementations, CD73 inhibitors can be used to prevent or treat immune-related diseases, disorders, or symptoms selected from the group consisting of: rheumatoid arthritis, renal failure, lupus, asthma, psoriasis, colitis, pancreatitis, allergies, fibrosis, anemia, fibromyalgia, Alzheimer's disease, congestive heart failure, stroke, aortic stenosis, arteriosclerosis, osteoporosis, Parkinson's disease, infections, Crohn's disease, ulcerative colitis, allergic contact dermatitis, eczema, systemic sclerosis, and multiple sclerosis.

[0247] The pharmaceutical compositions provided herein can be formulated to be compatible with the intended method or route of administration; exemplary routes of administration are described herein. Furthermore, the pharmaceutical compositions can be used in combination with other therapeutically active agents or compounds described herein to treat or prevent the CD73-related diseases, disorders, and conditions discussed herein.

[0248] Pharmaceutical compositions containing an active ingredient (e.g., a CD73 inhibitor) can be in forms suitable for oral administration, such as tablets, capsules, troche, lozenges, aqueous or oily suspensions, dispersible powders or granules, emulsions, hard or soft capsules, or syrups, solutions, microbeads, or elixirs. Pharmaceutical compositions for oral administration can be prepared according to any method known in the art for manufacturing pharmaceutical compositions, and such compositions can contain one or more agents, such as sweeteners, flavoring agents, coloring agents, and preservatives, to provide a pharmaceutically acceptable formulation. Tablets, capsules, etc., typically contain the active ingredient mixed with a non-toxic, pharmaceutically acceptable carrier or excipient suitable for manufacturing tablets. These carriers or excipients can be, for example, diluents such as calcium carbonate, sodium carbonate, lactose, calcium phosphate, or sodium phosphate; granulating and disintegrants such as corn starch or alginate; binders such as starch, gelatin, or gum arabic; and lubricants such as magnesium stearate, stearic acid, or talc.

[0249] Tablets, capsules, etc., suitable for oral administration can be uncoated or coated using known techniques to delay disintegration and absorption in the gastrointestinal tract and thereby provide sustained action. For example, delay-release materials such as glyceryl monostearate or glyceryl distearate can be used. They can also be coated using techniques known in the art to form osmotic therapeutic tablets for controlled release. Other agents include biodegradable or biocompatible particles or polymers such as polyesters, polyamines, hydrogels, polyvinylpyrrolidone, polyanhydrides, polyglycolic acid, ethylene vinyl acetate, methylcellulose, carboxymethylcellulose, protamine sulfate, or lactide / glycolic acid copolymers, polylactide / glycolic acid copolymers, or ethylene vinyl acetate copolymers to control the delivery of the administered composition. For example, oral formulations can be embedded in microcapsules prepared using hydroxymethylcellulose via coagulation technology or via interfacial polymerization, or in gelatin microcapsules or poly(methyl methacrylate) microcapsules, or embedded in colloidal drug delivery systems. Colloidal dispersion systems include macromolecular complexes, nanocapsules, microspheres, microbeads, and lipid-based systems, including oil-in-water emulsions, micelles, mixed micelles, and liposomes. Methods for preparing the above formulations will be readily apparent to those skilled in the art.

[0250] Formulations intended for oral use may also be in the form of hard gelatin capsules, wherein the active ingredient is mixed with an inert solid diluent (e.g., calcium carbonate, calcium phosphate, kaolin, or microcrystalline cellulose); or in the form of soft gelatin capsules, wherein the active ingredient is mixed with water or an oily medium (e.g., peanut oil, liquid paraffin, or olive oil). Aqueous suspensions contain active material mixed with excipients suitable for their manufacture. Such excipients can be suspending agents, such as sodium carboxymethyl cellulose, methyl cellulose, hydroxypropyl methyl cellulose, sodium alginate, polyvinylpyrrolidone, tragacanth gum, and gum arabic; dispersing or wetting agents, such as naturally occurring phospholipids (e.g., lecithin), or condensation products of olefinic oxygen and fatty acids (e.g., polyoxyethylene stearate), or condensation products of ethylene oxide and long-chain fatty alcohols (e.g., heptadecaethyleneoxycetyl alcohol), or condensation products of ethylene oxide and esters derived from fatty acids and hexitols (e.g., polyoxyethylene sorbitan monooleate), or condensation products of ethylene oxide and esters derived from fatty acids and hexitol anhydrides (e.g., polyvinyl sorbitan monooleate). Aqueous suspensions may also contain one or more preservatives.

[0251] Oily suspensions can be formulated by suspending the active ingredient in vegetable oils (such as peanut oil, olive oil, sesame oil, or coconut oil) or mineral oils (such as liquid paraffin). Oily suspensions may contain thickeners such as beeswax, hard paraffin, or cetyl alcohol. Sweeteners, as mentioned above, and flavoring agents may be added to provide a palatable oral formulation.

[0252] Dispersible powders and granules suitable for preparing aqueous suspensions by adding water provide an active ingredient that can be mixed with a dispersant or wetting agent, a suspending agent, and one or more preservatives. Suitable dispersants or wetting agents and suspending agents are known in the art.

[0253] The pharmaceutical compositions of the present invention may also be in the form of an oil-in-water emulsion. The oily phase may be a vegetable oil (e.g., olive oil or peanut oil) or a mineral oil (e.g., liquid paraffin), or a mixture thereof. Suitable emulsifiers may be naturally occurring gums, such as gum arabic and gum tragali; naturally occurring phospholipids, such as soybean, lecithin, and esters or metaesters derived from fatty acids; hexitan anhydrides, such as sorbitan monooleate; and condensation products of metaesters and ethylene oxide, such as polyoxyethylene sorbitan monooleate.

[0254] Pharmaceutical compositions typically comprise a therapeutically effective amount of the CD84 inhibitor compound provided by this invention and one or more pharmaceutically and physiologically acceptable formulations. Suitable pharmaceutically or physiologically acceptable diluents, carriers, or excipients include, but are not limited to, antioxidants (e.g., ascorbic acid and sodium disulfide), preservatives (e.g., benzyl alcohol, methylparaben, ethylparaben, or n-propylparaben), emulsifiers, suspending agents, dispersants, solvents, fillers, fillers, detergents, buffers, solvents, diluents, and / or adjuvants. For example, suitable solvents may be physiological saline solutions or citrate-buffered saline solutions, possibly supplemented with other substances common in pharmaceutical compositions intended for parenteral administration. Neutral buffered saline solutions or saline solutions mixed with serum albumin are further exemplary solvents. Those skilled in the art will readily recognize the various buffers that may be used in the pharmaceutical compositions and dosage forms considered herein. Typical buffers include, but are not limited to, pharmaceutically acceptable weak acids, weak bases, or mixtures thereof. For example, the buffer component can be a water-soluble substance such as phosphoric acid, tartaric acid, lactic acid, succinic acid, citric acid, acetic acid, ascorbic acid, aspartic acid, glutamic acid, and their salts. Acceptable buffers include, for example, Tris buffer, N-(2-hydroxyethyl)piperazine-N'-(2-ethanesulfonic acid) (HEPES), 2-(N-morpholine)ethanesulfonic acid (MES), sodium 2-(N-morpholine)ethanesulfonate (MES), 3-(N-morpholine)propanesulfonic acid (MOPS), and Ntris[hydroxymethyl]methyl-3-aminopropanesulfonic acid (TAPS). After formulation, the pharmaceutical composition can be stored in sterile vials in the form of solution, suspension, gel, emulsion, solid, dehydrated, or lyophilized powder. This formulation can be stored in ready-to-use form, lyophilized form requiring reconstitution before use, liquid form requiring dilution before use, or other acceptable forms.

[0255] In some embodiments, the pharmaceutical composition is contained in a single-use container (e.g., a single-use vial, ampoule, syringe, or autoinjector), while in other embodiments, it is contained in a reusable container (e.g., a reusable vial).

[0256] The formulation may also include a carrier to protect the composition from rapid degradation or disappearance from the body, such as controlled-release formulations, including liposomes, hydrogels, and microencapsulated delivery systems. For example, delayed-release materials (e.g., glyceryl monostearate or glyceryl stearate alone) or in combination with waxes may be used. Any drug delivery device can be used to deliver CD73 inhibitors, including implants (e.g., implantable pumps) and catheter systems, slow-infusion pumps, and devices, all of which are well known to those skilled in the art.

[0257] Pharmaceutical compositions can also be in the form of sterile injectable aqueous or oily suspensions. These suspensions can be formulated using suitable dispersants or wetting agents and suspending agents mentioned herein, according to known techniques. Sterile injectable formulations can also be sterile injectable solutions or suspensions in non-toxic, parenteral-acceptable diluents or solvents, such as solutions in 1,3-butanediol. Acceptable diluents, solvents, and dispersion media that can be used include water, Ringer's solution, isotonic sodium chloride solution, Cremophor ELTM (BASF, Parsippany, NJ) or phosphate-buffered saline (PBS), ethanol, polyols (e.g., glycerol, propylene glycol, and liquid polyethylene glycol), and suitable mixtures thereof. Additionally, sterile fixative oils are commonly used as solvents or suspension media. For this purpose, any mild fixative oil can be used, including synthetic monoglycerides or diglycerides. Furthermore, fatty acids (such as oleic acid) can be used to prepare injectable formulations. Prolonged absorption of specific injectable formulations can be achieved by including agents that delay absorption (e.g., aluminum monostearate or gelatin).

[0258] The CD73 inhibitor compounds and compositions provided by this invention can be administered to subjects in any suitable manner known in the art. Suitable routes of administration include, but are not limited to: oral; parenteral, such as intramuscular, intravenous, subcutaneous (e.g., injection or implantation), intraperitoneal, intracisional, intraarticular, intracerebral (intraparenchymal and intraventricular); nasal; vaginal; sublingual; intraocular; rectal; local (e.g., transdermal); oral; and inhalation. Accumulated injection, typically administered subcutaneously or intramuscularly, can also be used to release the CD73 inhibitors disclosed herein over a defined time period. In some embodiments, the CD73 inhibitor compounds and compositions are administered orally to subjects in need.

[0259] The CD73 inhibitor compounds and compositions provided by this invention can be administered to subjects in amounts that depend on, for example, the administration target (e.g., desired resolution); the age, weight, sex, health, and physical condition of the subject administering the formulation; the route of administration; and the presence of disease, disorder, condition, or symptoms thereof. The dosing regimen also takes into account the presence, nature, and extent of any adverse effects associated with the administered agent. Effective doses and dosing regimens can be readily determined by, for example, safety and dose escalation studies, in vivo studies (e.g., animal models), and other methods known to those skilled in the art. Typically, dosing parameters determine a dose less than the amount that may have irreversible toxicity to the subject (maximum tolerated dose, MTD) and not less than the amount required to produce a measurable effect in the subject. These amounts are determined, for example, by pharmacokinetic and pharmacodynamic parameters related to ADME, taking into account the route of administration and other factors.

[0260] In some embodiments, the CD73 inhibitor may be administered once daily or multiple times daily (e.g., orally) at a dose level of 0.01 mg / kg to about 50 mg / kg, or about 1 mg / kg to about 25 mg / kg, based on the subject's body weight, to achieve the desired therapeutic effect. For oral administration, the composition may be provided in the form of tablets or capsules containing 1.0 mg to 1000 mg of the active ingredient, particularly containing 1 mg, 3 mg, 5 mg, 10 mg, 15 mg, 20 mg, 25 mg, 50 mg, 75 mg, 100 mg, 150 mg, 200 mg, 250 mg, 300 mg, 400 mg, 500 mg, 600 mg, 750 mg, 800 mg, 900 mg, or 1000 mg of the active ingredient.

[0261] In some implementations, the desired dose of the CD73 inhibitor is included in a “unit dosage form.” The phrase “unit dosage form” refers to a physically discontinuous unit, each unit containing a predetermined amount of a single CD73 inhibitor or in combination with one or more other agents, sufficient to produce the desired effect. It should be understood that the parameters of the unit dosage form will depend on the specific agent and the desired effect.

[0262] This invention also provides kits comprising CD73 inhibitor compounds or compositions. Kits are typically in the form of a physical structure containing various components and can be used, for example, to perform the methods provided herein. For example, a kit may include one or more CD73 inhibitors disclosed herein (e.g., provided in a sterile container), which may be in the form of a pharmaceutical composition suitable for administration to a subject. CD73 inhibitors may be provided in ready-to-use form (e.g., tablets or capsules) or in form requiring, for example, reconstitution or dilution before administration (e.g., powder). When the CD73 inhibitor is in a form requiring reconstitution or dilution by the user, the kit may also include diluents (e.g., sterile water), buffers, pharmaceutically acceptable excipients, etc., packaged together with or separately from the CD73 inhibitor. When using combination therapy, the kit may contain several therapeutic agents independently, or they may already be combined in the kit. Each component of the kit may be packaged in a separate container, and all the various containers may be in a single package. The kits of this invention may be designed to maintain the components contained therein under the conditions required (e.g., refrigeration or freezing).

[0263] The kit may also include a label or packaging insert containing identification information for its components and instructions for use (e.g., dosage parameters, clinical pharmacology of the active ingredient, including mechanism of action, pharmacokinetics and pharmacodynamics, adverse effects, contraindications, etc.). The label or insert may include manufacturer information, such as batch number and expiration date. The label or packaging insert may be integrated into the physical structure containing the components, contained independently within the physical structure, or attached to a component of the kit (e.g., ampoules, tubes, or vials).

[0264] Example

[0265] The present invention will be more readily understood by referring to the following embodiments, which are provided for illustrative purposes and should not be construed as limiting the scope of the invention in any way.

[0266] Unless otherwise defined or the context clearly requires, all technical and scientific terms used herein have the same meaning as commonly understood by one of ordinary skill in the art to which this invention pertains. It should be understood that any methods and materials similar to or equivalent to those described herein may be used in the practice or testing of this invention.

[0267] Compound Synthesis

[0268]

[0269] The amine compound RNH2 can be obtained from commercial sources or prepared using methods described in the literature.

[0270] Prepare triethylammonium bicarbonate buffer (TEAC). Prepare a 1M TEAC solution by slowly adding dry ice to a 1M triethylamine solution in water over several hours until the pH of the solution reaches approximately 7.4–7.6 (measured using a pH meter).

[0271] 1.0 mmol (1.0 eq.) of the 2-chloropurine nucleoside derivative Sx was dissolved in 10 mL of trimethyl phosphate. The solution was cooled in an ice bath. A solution of 4.0 eq. of bis(dichlorophosphine)methane in 5 mL of trimethyl phosphate was added to the cooled solution. The mixture was stirred at 0 °C for 2–4 h, and the reaction was monitored by thin-layer chromatography (TLC). The reaction was quenched dropwise with TEAC solution, and the pH of the reaction mixture was adjusted to 7–8. The mixture was extracted with dichloromethane (DCM), the aqueous phase was separated, and concentrated. The residue was purified by reversed-phase chromatography using a C18 column to give the product as a white solid.

[0272] Example 1: Synthesis of Compound 1

[0273] DIEA (diisopropylethylamine, 7.5 mmol, 969 mg, 1.5 eq.) was added dropwise to a solution of 2,6-dichloro-9-(2',3',5'-tri-O-acetyl-β-D-furanoribosyl)purine (5.0 mmol, 2236 mg, 1.0 eq.) and benzylamine (5.0 mmol, 536 mg, 1.0 eq.) in dioxane (25 mL). The reaction mixture was stirred overnight at room temperature. The solvent was evaporated under vacuum, and the residue was dissolved in DCM (100 mL) and washed with water (2 × 30 mL). The crude product was purified by column chromatography. The intermediate was dissolved in 50 mL of NH3 / CH3OH solution and stirred overnight at 35 °C. The solvent was evaporated under vacuum, and the crude product was purified by column chromatography to give 2-chloropurine nucleoside derivative S-1 (1818 mg).

[0274] S-1 (1.0 mmol, 392 mg, 1.0 eq.) was dissolved in trimethyl phosphate (10 mL) and then cooled in an ice bath. A solution of bis(dichlorophosphine)methane (4.0 eq.) in trimethyl phosphate (5 mL) was added to the cooled solution. The reaction mixture was then stirred at 0 °C for 2–4 h, and the reaction was monitored by TLC. The reaction was quenched with TEAC solution, and the pH of the reaction mixture was adjusted to 7–8. The mixture was extracted with DCM, the aqueous phase was separated, and concentrated. The residue was purified by reversed-phase column chromatography (C18 column) to give compound 1 (369 mg) as an off-white solid. 1 H NMR (500MHz, CD3OD-d4) δppm 2.46 (t, 2H), 4.23-4.74 (m, 7H), 6.01 (d, 1H), 7.19-7.38 (m, 5H), 8.59 (s, 1H); 13 C NMR(125MHz,CD3Cl-d3)δppm 40.12, 43.84, 63.92, 69.87, 74.64, 83.71, 88.50, 115.22, 126.92, 127.47, 128.13, 149.27, 154.08, 154.96, 160.55; 31 P NMR (200MHz, CD3Cl-d3) δppm 12.94, 18.11; m / z (ESI + )550.1.

[0275] Example 2: Synthesis of Compound 6

[0276] DIEA (7.5 mmol, 969 mg, 1.5 eq.) was added dropwise to a solution of 2,6-dichloro-9-(2',3',5'-tris-O-acetyl-β-D-furanoribosyl)purine (5.0 mmol, 2.2 g, 1.0 eq.) and 1-naphthylmethylamine (5.0 mmol, 786 mg, 1.0 eq.) in dioxane (25 mL). The reaction mixture was stirred overnight at room temperature. The solvent was evaporated under vacuum, and the residue was dissolved in DCM (100 mL) and washed with water (2 × 30 mL). The solvent was removed (by rotary evaporation), and the residue was purified by column chromatography. The intermediate was dissolved in 50 mL of NH3 / CH3OH solution and stirred overnight at 35 °C. The solvent was evaporated under vacuum, and the residue was purified by column chromatography to give S-6 (1.3 g).

[0277] S-6 (1.0 mmol, 442 mg, 1.0 eq.) was dissolved in trimethyl phosphate (10 mL) and then cooled in an ice bath. A solution of bis(dichlorophosphoryl)methane (4.0 eq.) in trimethyl phosphate (5 mL) was added to the cooled solution. The reaction mixture was then stirred at 0 °C for 2–4 h, and the reaction was monitored by TLC. The reaction was quenched with TEAC solution, and the pH of the reaction mixture was adjusted to 7–8. The mixture was extracted with DCM. The aqueous phase was separated and concentrated, and the residue was purified by reversed-phase column chromatography (C18 column) to give compound 6 (110 mg) as an off-white solid. 1 H NMR(500MHz,D2O)δppm0.88-0.91(m,3H),1.34-1.43(m,4H),1.67-1.68(m,2H),2.15-2.26(m,2H),4.16-4.22(m,2H) ),4.25-4.31(m,2H),4.38-4.41(m,1H),4.53-4.57(m,1H),4.74-4.76(m,H),6.13-1.15(m,1H),8.70-8.75(m,1H); 13 C NMR(125MHz,D2O)δppm 13.20,21.60,27.24,27.55,63.46,67.12,70.15,74.34,84.08,84.14,87.38,120.98,142.83,150.24,152.49,153.22; 31 PNMR(200MHz,D2O)δ16.15,18.97; m / z(ES - )571.8.

[0278] Example 3: Synthesis of Compound 7

[0279] DIEA (7.5 mmol, 969 mg, 1.5 eq.) was added dropwise to a solution of 2,6-dichloro-9-(2',3',5'-tri-O-acetyl-β-D-furanoribosyl)purine (5.0 mmol, 2.2 g, 1.0 eq.) and 2-naphthylmethylamine (5.0 mmol, 786 mg, 1.0 eq.) in dioxane (25 mL). The reaction was stirred overnight at room temperature. The solvent was evaporated under vacuum, and the residue was dissolved in DCM (100 mL) and washed with water (2 × 30 mL). The crude product was purified by column chromatography. The intermediate was dissolved in 50 mL of NH3 / CH3OH solution and stirred overnight at 35 °C. The solvent was evaporated under vacuum, and the residue was purified by column chromatography to give the 2-chloropurine nucleoside derivative S-7 (1.15 g).

[0280] S-7 (1.0 mmol, 442 mg, 1.0 eq.) was dissolved in trimethyl phosphate (10 mL) and then cooled in an ice bath. A solution of bis(dichlorophosphoryl)methane (4.0 eq.) in trimethyl phosphate (5 mL) was added to the cooled solution. The reaction mixture was then stirred at 0 °C for 2–4 h, and the reaction was monitored by TLC. The reaction was quenched with TEAC solution, and the pH of the reaction mixture was adjusted to 7–8. The mixture was extracted with DCM, the aqueous phase was separated, and concentrated. The residue was purified by reversed-phase column chromatography (C18 column) to give compound 7 (105 mg) as an off-white solid. 1 HNMR(500MHz,D2O)δppm2.11(t,J=19.7Hz,2H),4.10(s,2H),4.29(s,1H),4.42(s,1H),4.55(s,1H),4.88(s,2H), 5.79(s,1H),7.36-7.44(m,4H),7.71(d,J=7.9Hz,1H),7.76(d,J=7.2Hz,1H),7.87(d,J=7.8Hz,1H),8.13(s,1H); 13 C NMR(125MHz,D2O)δppm 27.57,42.23,63.39,70.05,74.16,83.59,86.92,117.78,122.74,125.48,125.88,126 .05,126.26,128.24,128.43,130.48,132.19,133.08,139.02,148.62,153.86,154.44. 31 P NMR(202MHz,D2O)δppm 15.17,19.58; m / z(ES -)598.2.

[0281] Example 4: Synthesis of Compound 8

[0282] 2-Chloropurinic nucleoside derivative S-8 (1.0 mmol, 415 mg, 1.0 eq.) was dissolved in trimethyl phosphate (10 mL), and the solution was then cooled in an ice bath. A solution of bis(dichlorophosphoryl)methane (4.0 eq.) in trimethyl phosphate (5 mL) was added to the cooled solution while maintaining an ice bath. The mixture was then stirred at 0 °C for 2–4 h, and the reaction was monitored by TLC. The reaction was quenched with TEAC solution, and the pH of the reaction mixture was adjusted to 7–8. The mixture was extracted with DCM, the aqueous phase was separated, and concentrated. The residue was purified by reversed-phase column chromatography (C18 column) to give the product (58 mg) as a white solid. 1 H NMR(500MHz,D2O)δppm 2.12(t,J=19.7Hz,2H),4.08(s,2H),4.27(s,1H),4.40(s,1H),4.53(s,1H),4.65( s,2H),5.75(s,1H),7.36(s,3H),7.65(s,2H),7.69(d,J=8.0Hz,2H),8.28(s,1H); 13 C NMR(125MHz,D2O)δppm 44.04,63.41,70.06,74.19,83.73,86.90,125.50,125.60,126.00,126 .34,127.38,128.15,132.13,132.71,135.18,139.21,153.96,154.73; 31 P NMR(202MHz,D2O)δppm 15.86,19.01; m / z(ES - 598.4.

[0283] Example 5: Synthesis of Compound 9

[0284] DIEA (12.5 mmol, 1.6 g, 2.5 eq.) was added dropwise to a solution of 2,6-dichloro-9-(2',3',5'-tri-O-acetyl-β-D-furanoside)purine (5.0 mmol, 2.2 g, 1.0 eq.) and memantine hydrochloride (5.0 mmol, 1.0 g, 1.0 eq.) in 25 mL of dioxane. The reaction mixture was stirred overnight at room temperature. The solvent was evaporated under vacuum, and the residue was dissolved in DCM (100 mL), washed with water (2 × 30 mL), and concentrated. The residue was purified by column chromatography to give an intermediate. This intermediate was dissolved in 50 mL of NH3 / CH3OH solution, and the mixture was stirred overnight at 35 °C. The solvent was evaporated under vacuum, and the residue was purified by column chromatography to give the 2-chloropurine nucleoside derivative S-9 (1.1 g).

[0285] S-9 (1.0 mmol, 463 mg, 1.0 eq.) was dissolved in trimethyl phosphate (10 mL), and the mixture was then cooled in an ice bath. A solution of bis(dichlorophosphoryl)methane (4.0 eq.) in trimethyl phosphate (5 mL) was added to the cooled mixture. The mixture was then stirred at 0 °C for 2–4 h, and the reaction was monitored by TLC. After the reaction was complete, it was quenched with TEAC solution. The pH of the mixture was adjusted to 7–8. The mixture was extracted with DCM, and the aqueous phase was separated. The aqueous phase was concentrated, and the residue was purified by reversed-phase column chromatography (C18 column) to give compound 9 (200 mg) as an off-white solid. 1 HNMR(500MHz,D2O)δppm 0.76(s,6H),0.98-1.12(m,2H),1.22(s,2H),1.31(d,J=11.3Hz,2H),1.70(dd,J=29.4,11.9Hz,4H),1.90(s,2H),2.11(t,J =19.9Hz,3H),4.07(s,2H),4.28(s,1H),4.44(dd,J=6.5,2.4Hz,1H),4.67-4.63(m,1H),5.91(d,J=5.7Hz,1H),8.33(s,1H); 13 C NMR(125MHz,D2O)δppm 25.96,26.95,29.97,32.08,39.23,42.32,50.32,55.08,63.73,70.04,74.33,83.78,87.45,116.10,138.34,148.62,153.57,154.16; 31 P NMR (200MHz, D2O) δppm 18.12; m / z (ES- )620.2.

[0286] Example 6: Synthesis of Compound 10

[0287] 2-Chloropurinic nucleoside derivative S-10 (1.0 mmol, 451 mg, 1.0 eq.) was dissolved in trimethyl phosphate (10 mL), and the solution was cooled in an ice bath. A solution of bis(dichlorophosphoryl)methane (4.0 eq.) in trimethyl phosphate (5 mL) was added to the cooled solution. The mixture was then stirred at 0 °C for 2–4 h, and the reaction was monitored by TLC. The reaction was quenched with TEAC solution, and the pH of the reaction mixture was adjusted to 7–8. The mixture was extracted with DCM, the aqueous phase was separated, and concentrated. The residue was purified by reversed-phase column chromatography (C18 column) to give compound 10 (60 mg) as an off-white solid. 1 HNMR(500MHz,D2O)δppm 2.27(t,J=19.4Hz,2H),4.17(s,2H),4.35(s,1H),4.46(s,1H),6.02(s,1H),7.27(s,4H),7.54(s,2H),7.97(s,2H),8.33(s,1H); 13 C NMR(125MHz,D2O)δppm16.73,25.47,26.48,27.50,57.39,63.69,70.09,74.31,83.84,8 7.70,113.16,120.00,122.84,124.66,126.42,138.03,143.08,148.86,152.70,154.12; 31 P NMR(200MHz,D2O)δppm 17.43,19.35-19.97; m / z(ES - 608.0.

[0288] Example 7: Synthesis of Compound 11

[0289] The 2-chloropurine nucleoside derivative S-11 (1.0 mmol, 564 mg, 1.0 eq.) was dissolved in trimethyl phosphate (10 mL). The solution was cooled in an ice bath, and a solution of bis(dichlorophosphoryl)methane (4.0 eq.) in trimethyl phosphate solution (5 mL) was added to the cooled solution. The mixture was then stirred at 0 °C for 2–4 h, and the reaction was monitored by TLC. The reaction was quenched with TEAC solution, and the pH of the reaction mixture was adjusted to 7–8. The mixture was extracted with DCM, the aqueous phase was separated and concentrated. The residue was purified by reversed-phase column chromatography (C18 column) to give compound 11 (100 mg) as an off-white solid. 1 H NMR(500MHz,D2O)δppm 0.77(d,J=6.5Hz,3H),1.22(s,14H),1.55(s,2H),1.92(s,4H),2.12(t,J=19.8Hz,2H),2.67(d,J=38.2Hz,4H), 4.03(s,2H),4.19(s,1H),4.40(s,1H),4.60(s,1H),5.20(d,J=5.6Hz,4H),5.88(d,J=4.4Hz,1H),8.51(s,1H); 13 CNMR(125MHz,D2O)δppm 13.90,22.48,24.71,25.52,27.10,29.26,29.64,31.41,36.86,37.49,63.89,70.44,7 4.37,84.15,86.83,119.95,127.82,129.70,149.34,152.45,152.86,164.88,174.66; 31 P NMR(200MHz,D2O)δppm 16.04,18.65; m / z(ES - )720.4.

[0290] Example 8: Synthesis of Compound 12

[0291] DIEA (7.5 mmol, 969 mg, 1.5 eq.) was added dropwise to a solution of 2,6-dichloro-9-(2',3',5'-tris-O-acetyl-β-D-furanoribosyl)purine (5.0 mmol, 2.23 g, 1.0 eq.) and 2-naphthylamine (5.0 mmol, 715 mg, 1.0 eq.) in 25 mL of dioxane. The reaction mixture was stirred overnight at room temperature. The solvent was evaporated under vacuum, and the residue was dissolved in DCM (100 mL) and washed with water (2 × 30 mL). The DCM solution was concentrated to dryness, and the residue was purified by column chromatography to give an intermediate. This intermediate was dissolved in 50 mL of NH3 / CH3OH solution, and the mixture was stirred overnight at 35 °C. The solvent was evaporated under vacuum, and the residue was purified by column chromatography to give compound S-12 (670 mg).

[0292] S-12 (1.0 mmol, 427 mg, 1.0 eq.) was dissolved in trimethyl phosphate (10 mL), and the solution was cooled in an ice bath. A solution of bis(dichlorophosphoryl)methane (4.0 eq.) in trimethyl phosphate (5 mL) was added to the cooled solution. The mixture was then stirred at 0 °C for 2–4 h, and the reaction was monitored by TLC. The reaction was quenched with TEAC solution, and the pH of the quenched reaction mixture was adjusted to 7–8. The mixture was extracted with DCM, the aqueous phase was separated, and concentrated. The residue was purified by reversed-phase column chromatography (C18 column) to give compound 12 (100 mg) as an off-white solid. 1 HNMR(D2O,500MHz)δppm 2.21-2.29(m,2H),4.20-4.22(m,2H),4.40-4.44(m,1H),4.56-4.60(m,1H),4.78-4.80(m,1H),6.09- 6.10(m,1H),7.58-7.64(m,3H),7.71-7.73(m,1H),7.97-7.99(m,1H)8.02-8.04(m,2H),8.55(s,1H); 13 C NMR(D2O,125MHz)δppm26.38,27.37,28.36,63.61,70.33,84.01,86.76,118. 06,121.57,122.25,125.52,126.23,126.41,139.82,149.74,153.39,153.47; 31 P NMR (D2O, 200MHz) δppm 16.14, 18.96; m / z (ES - 583.9.

[0293] Example 9: Synthesis of Compound 15

[0294] The 2-chloropurine nucleoside derivative S-15 (1.0 mmol, 513 mg, 1.0 eq.) was dissolved in trimethyl phosphate (10 mL). The mixture was cooled in an ice bath, and then bis(dichlorophosphoryl)methane (4.0 eq.) in a solution of trimethyl phosphate (5 mL) was added. The mixture was stirred at 0 °C for 2–4 h, and the reaction was monitored by TLC. The reaction was quenched with TEAC solution, and the pH of the reaction mixture was adjusted to 7–8. The mixture was extracted with DCM, the aqueous phase was separated, and concentrated. The residue was purified by reversed-phase column chromatography (C18 column) to give compound 15 (30 mg) as an off-white solid. 1 H NMR(500MHz,D2O)δppm 0.70(t,J=6.3Hz,3H),1.06(s,16H),1.27(s,2H),1.58(s,2H),2.10(t,J=19.7Hz,2H),4.12( d,J=32.1Hz,4H),4.27(s,1H),4.46(s,1H),4.66(s,1H),6.00(d,J=4.7Hz,1H),8.60(s,1H); 13 C NMR(125MHz,D2O)δppm 13.73,22.40,25.51,27.55,28.40,29.07,29.36,31.66,63.64,66.70 ,70.27,74.36,83.95,87.18,120.57,142.65,150.05,152.38,153.2; 31 PNMR(200MHz,D2O)δppm 15.64,18.89; m / z(ES - 670.1.

[0295] Example 10: Synthesis of Compound 16

[0296] The 2-chloropurine nucleoside derivative S-16 (1.0 mmol, 478 mg, 1.0 eq.) was dissolved in trimethyl phosphate (10 mL), and the mixture was cooled in an ice bath. Then, a solution of bis(dichlorophosphoryl)methane (4.0 eq.) in trimethyl phosphate (5 mL) was added. The mixture was stirred at 0 °C for 2–4 h, and the reaction was monitored by TLC. The reaction was quenched with TEAC solution, and the pH of the mixture was adjusted to 7–8. The mixture was extracted with DCM, the aqueous phase was separated, and concentrated. The residue was purified by reversed-phase column chromatography (C18 column) to give compound 16 (250 mg) as an off-white solid. 1H NMR(500MHz,D2O)δppm 1.64(s,11H),1.91(s,4H),2.16(t,J=19.8Hz,2H),2.29(s,2H),4.14(dddd,J=6.8,5.9,4 .1,1.6Hz,3H),4.30-4.37(m,1H),4.56-4.46(m,1H),6.11(d,J=5.1Hz,1H),8.72(s,1H); 13 C NMR(125MHz,D2O)δppm 16.75,27.23,28.38,33.41,36.07,42.03,51.44,63.47,70.17,74.37,84.19,87.49,115.91,121.90,143.36,149.28,153.15,173.66; 31 P NMR(200MHz,D2O)δppm 16.37,18.85; m / z(ES - )634.1.

[0297] Example 11: Synthesis of Compound 17

[0298] DIEA (12.5 mmol, 1.6 g, 2.5 eq.) was added dropwise to a solution of 2,6-dichloro-9-(2',3',5'-tri-O-acetyl-β-D-furanoribosyl)purine (5.0 mmol, 2.23 g, 1.0 eq.) and 2-adamantaneamine hydrochloride (5.0 mmol, 0.94 g, 1.0 eq.) in 25 mL of dioxane. The mixture was stirred overnight at room temperature. The solvent was evaporated under vacuum, and the residue was dissolved in DCM (100 mL) and washed with water (2 × 30 mL). The organic layer was concentrated, and the residue was purified by column chromatography to give an intermediate. The intermediate was dissolved in 50 mL of NH3 / CH3OH solution and stirred overnight at 35 °C. The solvent was evaporated, and the residue was then purified by column chromatography to give the 2-chloropurine nucleoside derivative S-17 (880 mg).

[0299] Compound S-17 (1.0 mmol, 435 mg, 1.0 eq.) was dissolved in trimethyl phosphate (10 mL), and the mixture was then cooled in an ice bath. A solution of bis(dichlorophosphoryl)methane (4.0 eq.) in trimethyl phosphate (5 mL) was added to the cooled mixture. The mixture was then stirred at 0 °C for 2–4 h, and the reaction was monitored by TLC. The reaction was quenched with TEAC solution, and the pH of the mixture was adjusted to 7–8. The mixture was extracted with DCM, the aqueous phase was separated, and concentrated. The residue was purified by reversed-phase column chromatography (C18 column) to give compound S-17 (30 mg) as an off-white solid. 1 H NMR(500MHz,D2O)δppm 1.60(d,J=12.7Hz,2H),1.71(s,2H),1.82(d,J=20.8Hz,7H),1.96(d,J=19.6Hz,4H),2.14(t,J=19.8Hz ,2H),4.10(s,2H),4.20(s,1H),4.31(s,1H),4.47(t,J=4.2Hz,1H),5.96(d,J=5.6Hz,1H),8.41(s,1H); 13 C NMR(125MHz,D2O)δppm 16.70,26.77,30.81,31.44,36.48,36.86,57.36,63.51,70.23,74.20,83.97,86.73,139.05,154.39; 31 P NMR(200MHz,D2O)δppm 15.85,19.04; m / z(ES - )592.0.

[0300] Example 12: Synthesis of Compound 18

[0301] The 2-chloropurine nucleoside derivative S-18 (1.0 mmol, 512 mg, 1.0 eq.) was dissolved in trimethyl phosphate (10 mL). The mixture was cooled in an ice bath, and then a solution of bis(dichlorophosphoryl)methane (4.0 eq.) in trimethyl phosphate solution (5 mL) was added. The mixture was then stirred at 0 °C for 2–4 h, and the reaction was monitored by thin-layer chromatography. The reaction was quenched with TEAC solution, and the pH of the mixture was adjusted to 7–8. The mixture was extracted with DCM, the aqueous phase was separated, and concentrated. The residue was purified by reversed-phase column chromatography (C18 column) to give compound 18 (70 mg) as an off-white solid.

[0302] 1H NMR(500MHz,CD3OD-d4)δppm 0.93(t,J=6.8Hz,3H),1.52(s,18H),1.60-1.74(m,2H),2.33(t,J=19.8Hz,2H),3.43(t,J=6.6Hz,2H) ,4.27(d,J=21.4Hz,3H),4.47-4.61(m,1H),4.71(t,J=5.3Hz,1H),6.13(d,J=5.4Hz,1H),8.76(s,1H); 13 C NMR(125MHz,CD3OD-d4)δppm 13.01,22.30,26.62,28.98,29.28,31.63,39.56,63.95,70.57,74.91,84.44,87.86,118.78,142.32,150.79,151.78,151.97,154.17; 31 P NMR (200MHz, CD3OD-d4) δppm16.03,20.25; m / z (ES - ):669.2.

[0303] Example 13: Synthesis of Compound 19

[0304] DIEA (7.5 mmol, 969 mg, 1.5 eq.) was added dropwise to a solution of 2,6-dichloro-9-(2',3',5'-tri-O-acetyl-β-D-furanoribosyl)purine (5.0 mmol, 2.23 g, 1.0 eq.) and di-n-dodecylamine (5.0 mmol, 1.8 g, 1.0 eq.) in 25 mL of dioxane. The reaction mixture was stirred overnight at room temperature. The solvent was evaporated under vacuum, and the residue was dissolved in DCM (100 mL) and washed with water (2 × 30 mL). The organic layer was concentrated to dryness, and the residue was purified by column chromatography to give an intermediate compound. This intermediate was dissolved in 50 mL of NH3 / CH3OH solution, and the mixture was stirred overnight at 35 °C. The solvent was evaporated under vacuum, and the residue was purified by column chromatography to give the 2-chloropurine nucleoside derivative S-19 (1.3 g).

[0305] S-19 (1.0 mmol, 637 mg, 1.0 eq.) was dissolved in trimethyl phosphate (10 mL). The mixture was cooled in an ice bath, and then a solution of bis(dichlorophosphoryl)methane (4.0 eq.) in trimethyl phosphate solution (5 mL) was added. The resulting mixture was then stirred at 0 °C for 2–4 h, and the reaction was monitored by TLC. The reaction was quenched with TEAC solution, and the pH of the reaction mixture was adjusted to 7–8. The mixture was extracted with DCM, the aqueous phase was separated, and concentrated. The residue was then subjected to reverse-phase chromatography. 18 Purification by C2 column chromatography yielded compound 19 (150 mg) as an off-white solid. 1 H NMR(500MHz,CD3OD-d4)δppm 0.93(t,J=6.6Hz,6H),1.32-1.46(m,36H),1.72(s,4H),2.36(t,J=20.0Hz,2H),3.70(s,2H) ,4.11-4.32(m,5H),4.47(s,1H),4.66(t,J=5.2Hz,1H),6.04(d,J=5.4Hz,1H),8.37(s,1H); 13 C NMR(125MHz,CD3OD-d4)δppm 13.08,22.34,26.44,29.08,29.34,31.68,64.24,70.51,74.51,83.84,87.46,118.27,137.84,151.64,153.46,154.25; 31 P NMR (200MHz, CD3OD-d4) δppm16.20,19.99; m / z (ES - ):794.6.

[0306] Example 14: Synthesis of Compound 20

[0307] DIEA (7.5 mmol, 969 mg, 1.5 eq.) was added dropwise to a solution of 2,6-dichloro-9-(2',3',5'-tri-O-acetyl-β-D-furanoribosyl)purine (5.0 mmol, 2.23 g, 1.0 eq.) and 2-aminoanthracene (5.0 mmol, 1.0 g, 1.0 eq.) in 25 mL of dioxane. The reaction mixture was stirred overnight at room temperature. The solvent was evaporated under vacuum, and the residue was dissolved in DCM (100 mL) and washed with water (2 × 30 mL). The organic layer was concentrated to dryness, and the residue was purified by column chromatography to give an intermediate compound. This intermediate was dissolved in 50 mL of NH3 / CH3OH solution and stirred overnight at 35 °C. The solvent was evaporated under vacuum, and the residue was then purified by column chromatography to give the 2-chloropurine nucleoside derivative S-20 (770 mg).

[0308] S-20 (1.0 mmol, 477 mg, 1.0 eq.) was dissolved in trimethyl phosphate (10 mL). The mixture was then cooled in an ice bath, followed by the addition of a solution of bis(dichlorophosphoryl)methane (4.0 eq.) in trimethyl phosphate (5 mL). The mixture was then stirred at 0 °C for 2–4 h, and the reaction was monitored by TLC. The reaction was quenched with TEAC solution, and the pH of the reaction mixture was adjusted to 7–8. The mixture was extracted with DCM, the aqueous phase was separated, and concentrated. The residue was purified by reversed-phase column chromatography (C18 column) to give compound 20 (80 mg) as an off-white solid. 1 H NMR(500MHz,D2O)δppm 2.25(t,J=19.4Hz,2H),4.28(d,J=58.1Hz,3H),4.46(s,2H),5.56(s,1H),7.19(s,3H),7.57(d,J=73.7Hz,4H),7.85(s,2H),8.21(s,1H); 31 P NMR(200MHz,D2O)δppm 18.42,19.15; m / z(ES - ):633.9.

[0309] Example 15: Synthesis of Compound 22

[0310] DIEA (7.5 mmol, 969 mg, 1.5 eq.) was added dropwise to a solution of 2,6-dichloro-9-(2',3',5'-tri-O-acetyl-β-D-furanoribosyl)purine (5.0 mmol, 2.23 g, 1.0 eq.) and 1-adamantaneamine (5.0 mmol, 756 mg, 1.0 eq.) in 25 mL of dioxane. The reaction mixture was stirred overnight at room temperature. The solvent was evaporated under vacuum, and the residue was dissolved in DCM (100 mL) and washed with water (2 × 30 mL). After removing the solvent, the residue was purified by column chromatography to give an intermediate compound. This intermediate was dissolved in 50 mL of NH3 / CH3OH solution, and the mixture was stirred overnight at 35 °C. The solvent was evaporated under vacuum, and the residue was then purified by column chromatography to give the 2-chloropurine nucleoside derivative S-22 (770 mg).

[0311] S-22 (1.0 mmol, 435 mg, 1.0 eq.) was dissolved in trimethyl phosphate (10 mL). The mixture was then cooled in an ice bath, followed by the addition of a solution of bis(dichlorophosphoryl)methane (4.0 eq.) in trimethyl phosphate (5 mL). The mixture was then stirred at 0 °C for 2–4 h, and the reaction was monitored by TLC. The reaction was quenched with TEAC solution, and the pH of the reaction mixture was adjusted to 7–8. The mixture was extracted with DCM, the aqueous phase was separated, and concentrated. The residue was purified by reversed-phase column chromatography (C18 column) to give compound 22 (100 mg) as an off-white solid. 1 H NMR(500MHz,D2O)δppm 1.64(s,6H),2.06(d,J=33.5Hz,9H),2.17(d,J=19.9Hz,2H),4.09(s,2H),4.30(d,J=0.9Hz,1 H),4.45(ddd,J=5.9,3.0,2.0Hz,1H),4.68-4.65(m,1H),5.94(d,J=5.2Hz,1H),8.38(s,1H); 13 C NMR (125MHz, D2O) δppm 26.22, 27.22, 29.28, 35.72, 40.80, 53.50, 63.57, 70.19, 74.20, 83.95, 86.81, 138.60, 148.74, 153.76, 154.43; 31 P NMR (200MHz, D2O) δppm 16.38, 18.81; m / z (ES-) 592.2.

[0312] Example 16: Synthesis of Compound 23

[0313] The 2-chloropurine nucleoside derivative S-23 (1.0 mmol, 481 mg, 1.0 eq.) was dissolved in trimethyl phosphate (10 mL), and the mixture was cooled in an ice bath. A solution of bis(dichlorophosphoryl)methane (4.0 eq.) in trimethyl phosphate (5 mL) was added to the cooled mixture. The mixture was then stirred at 0 °C for 2–4 h, and the reaction was monitored by TLC. The reaction was quenched with TEAC solution, and the pH of the reaction mixture was adjusted to 7–8. The mixture was extracted with DCM, the aqueous phase was separated, and concentrated. The residue was purified by reversed-phase column chromatography (C18 column) to give compound 23 (141 mg) as an off-white solid. 1 H NMR(D2O,500MHz)δppm 0.90-0.97(m,9H),1.18-1.20(m,1H),1.36-1.40(m,1H),1.75-1.80(m,1H),1.90-1.97(m,1H),2.00-2.30(m,2H),2.40-2.45(m,1 H),4.20-4.25(m,2H),4.40-4.44(m,1H),4.44-4.57(m,1H),4.72-4.73(m,1H),4.91-5.05(m,1H),6.43-6.45(m,1H),9.02(m,1H); 13 C NMR(D2O,125MHz)δ12.74,18.03,18.90,35.91,44.47,47.41,48.45,63.50,70.16,74 .34,83.22,84.06,84.13,87.37,120.84,142.72,150.23,152.41,153.24,153.45ppm; 31 P NMR (D2O, 200MHz) δ16.49, 18.79ppm; m / z (ES - 638.0.

[0314] Example 17: Synthesis of Compound 31

[0315] DIEA (7.5 mmol, 969 mg, 1.5 eq.) was added dropwise to a solution of 2,6-dichloro-9-(2',3',5'-tri-O-acetyl-β-D-furanoribosyl)purine (5.0 mmol, 2.23 g, 1.0 eq.) and 3-azaspirocyclo[4.5]decane (5.0 mmol, 696 mg, 1.0 eq.) in 25 mL of dioxane. The reaction mixture was stirred overnight at room temperature. The solvent was evaporated under vacuum, and the residue was dissolved in DCM (100 mL) and washed with water (2 × 30 mL). The organic layer was concentrated to dryness, and the residue was purified by column chromatography to give an intermediate compound. This intermediate was dissolved in 50 mL of NH3 / CH3OH solution and stirred overnight at 35 °C. The solvent was evaporated under vacuum, and the residue was then purified by column chromatography to obtain the corresponding 2-chloropurine nucleoside derivative S-31 (1.1 g).

[0316] S-31 (1.0 mmol, 423 mg, 1.0 eq.) was dissolved in trimethyl phosphate (10 mL), and the mixture was cooled in an ice bath. A solution of bis(dichlorophosphoryl)methane (4.0 eq.) in trimethyl phosphate (5 mL) was added to the cooled mixture. The mixture was then stirred at 0 °C for 2–4 h, and the reaction was monitored by TLC. The reaction was quenched with TEAC solution, and the pH of the mixture was adjusted to 7–8. The mixture was extracted with DCM, the aqueous phase was collected, and concentrated. The residue was purified by reversed-phase column chromatography (C18 column) to give compound 31 (140 mg) as an off-white solid. 1 H NMR(500MHz,D2O)δppm1.43(t,J=22.8Hz,10H),1.83(d,J=52.0Hz,2H),2.15(t,J=19.7Hz,2H),3.38(s,1H),3.58(s,1H),3.79(s,1H),4 0.1(s,1H),4.12(s,2H),4.32(s,1H),4.49(t,J=4.4Hz,1H),4.67-4.70(m,1H),5.97-5.98(d,J=5.4Hz,1H),8.36-8.37(d,J=6.8Hz,1H); 13 C NMR(125MHz,D2O)δppm 7.67,8.70,9.72,22.92,27.41,34.49,45.49,47.78,69.61,73.58,74.79,86.08,87.43,118.15,139.22,150.08,153.06,153.84; 31P NMR (200MHz, D2O) δppm15.71,19.11; m / z (ES - ):580.0.

[0317] Example 18: Synthesis of Compound 51

[0318] DIEA (7.5 mmol, 969 mg, 1.5 eq.) was added dropwise to a solution of 2,6-dichloro-9-(2',3',5'-tri-O-acetyl-β-D-furanoribosyl)purine (5.0 mmol, 2.23 g, 1.0 eq.) and nortropinol (5.0 mmol, 636 g, 1.0 eq.) in 25 mL of dioxane. The reaction mixture was stirred overnight at room temperature. The solvent was evaporated under vacuum, and the residue was dissolved in DCM (100 mL) and washed with water (2 × 30 mL). The residue was purified by column chromatography to give an intermediate compound. This intermediate was dissolved in 50 mL of NH3 / CH3OH solution, and the mixture was stirred overnight at 35 °C. After removing the solvent (under vacuum), the residue was purified by column chromatography to give the 2-chloropurine nucleoside derivative S-51 (1.2 g).

[0319] Compound S-51 (1.0 mmol, 411 mg, 1.0 eq.) was dissolved in trimethyl phosphate (10 mL). The mixture was cooled in an ice bath, and then a solution of bis(dichlorophosphoryl)methane (4.0 eq.) in trimethyl phosphate solution (5 mL) was added. The mixture was then stirred at 0 °C for 2–4 h, and the reaction was monitored by TLC. The reaction was quenched with TEAC solution, and the pH of the reaction mixture was adjusted to 7–8. The mixture was extracted with DCM, the aqueous phase was separated, and concentrated. The residue was purified by reversed-phase column chromatography (C18 column) to give compound S-51 (160 mg) as an off-white solid. 1 H NMR(500MHz,D2O)δppm 1.78-2.42(m,11H),4.06(s,1H),4.12(s,2H),4.28-4.37(m,1H),4.45-4.55(m,1H) ,4.82-4.88(m,1H),5.38(dd,J=3.4,2.4Hz,1H),5.99(d,J=5.6Hz,1H),8.40(s,1H); 13 C NMR(125MHz,D2O)δppm 26.05,27.10,27.68,37.08,38.20,53.79,54.40,64.38,70.20,74.09,83.99,86.71,117.78,138.54,151.12,154.22; 31P NMR(200MHz,D2O)δppm 16.90,18.63; m / z(ES - ):568.0.

[0320] Example 19: Synthesis of compound a-1

[0321]

[0322] In a 1000 mL round-bottom flask, add D-ribose (50 g, 333.05 mmol, 1 eq.), acetone (400 mL), 2,2-dimethoxypropane (100 mL), and HClO4 (25 g, 25.00 mL, 70% purity). Stir the mixture at room temperature for 2.5 hours, then add MeOH (30 mL) solution and stir overnight. After the reaction is complete, cool to -30 °C, then slowly add 30% Na2CO3 solution (75 mL) to keep the temperature below 10 °C. Remove acetone (400 mL) to obtain the crude product. Filter the precipitate and wash with ethyl acetate (50 mL). Concentrate the filtrate. Dilute the resulting liquid with ethyl acetate (300 mL). Wash the organic layer with brine (300 mL), then dry (Na2SO4), filter, and evaporate to dryness. The residue was purified by silica gel column chromatography (elution, PE / EA: 100:0 to 70:30) to give [(3aR,4R,6R,6aR)-4-methoxy-2,2-dimethyl-3a,4,6,6a-tetrahydrofurano[3,4-d][1,3]dioxono-6-yl]methanol (43 g, 63.22%). 1 H NMR(500MHz,CDCl3)δppm 1.29(s,3H),1.44(d,J=8.3Hz,3H),3.36-3.45(m,3H),3.52-3.72(m,2H),4.3 9(t,J=2.8Hz,1H),4.56(d,J=5.9Hz,1H),4.80(d,J=5.9Hz,1H),4.94(s,1H).

[0323] Pyridine (17.82 g, 225.25 mmol, 18.13 mL, 2.3 eq.) was added to a solution of [(3aR,4R,6R,6aR)-4-methoxy-2,2-dimethyl-3a,4,6,6a-tetrahydrofurano[3,4-d][1,3]dioxono-6-yl]methanol (20 g, 97.93 mmol, 1 eq.) in DCM (700 mL). The mixture was cooled to approximately -10 °C. Then, trifluoromethanesulfonic anhydride (58.03 g, 205.66 mmol, 34.60 mL, 2.1 eq.) was added dropwise to the mixture at approximately -10 °C. The mixture turned red and formed a large amount of solid, which was kept at -10 °C for approximately 4 hours. The organic layer was washed with water (400 mL), then with brine (300 mL), dried over Na₂SO₄, filtered, and evaporated to dryness. The residue was purified by silica gel column chromatography (elution, PE / EA: 100:0 to 85:15) to give a brown oily [(3aR,4R,6R,6aR)-4-methoxy-2,2-dimethyl-3a,4,6,6a-tetrahydrofurano[3,4-d][1,3]dioxono-6-yl]methyltrifluoromethanesulfonate (21.6 g, 65.59%).

[0324] Under N2 protection, a solution of 1-[ethoxy(methyl)phosphoryl]oxyethane (9.77 g, 64.23 mmol, 1.0 eq.) in THF (60 mL) was cooled to approximately -78 °C. Then, n-BuLi (2.5 M, 28.26 mL, 1.1 eq.) was added dropwise to the mixture at approximately -78 °C. The mixture was stirred at -78 °C for 25 minutes. At approximately -78 °C, a solution of [(3aR,4R,6R,6aR)-4-methoxy-2,2-dimethyl-3a,4,6,6a-tetrahydrofurano[3,4-d][1,3]dioxono-6-yl]methyltrifluoromethanesulfonate (21.6 g, 64.23 mmol, 1 eq.) in THF (20 mL) was added dropwise to the mixture. The mixture was stirred at -78°C for 1 hour, then quenched at -78°C with an aqueous solution of NH4Cl (60 mL). H2O (60 mL) and EtOAc (60 mL) were added to the mixture. The organic layer was separated, and the aqueous layer was extracted with EtOAc (60 mL). The organic layers were combined and concentrated, and the residue was purified by silica gel column chromatography, eluting with PE / EA from 100:0 to 0:100, to give (3aR,4R,6R,6aR)-6-(2-diethoxyphosphorylethyl)-4-methoxy-2,2-dimethyl-3a,4,6,6a-tetrahydrofurano[3,4-d][1,3]dioxamol (11 g, 50.62%) as a yellow oil. 1H NMR(500MHz, CDCl3)δppm:1.28-1.34(m,9H).1.47(s,3H),1.72-2.00(m,4H),3.35(t,J=1 0.7Hz,3H),4.01-4.19(m,5H),4.53(d,J=5.9Hz,1H),4.60(d,J=5.3Hz,1H),4.94(s,1H).

[0325] Under N2 protection, a solution of 2-[isopropoxy(methyl)phosphoryl]oxypropane (1.60 g, 8.87 mmol, 2 eq) in THF (6 mL) was cooled to approximately -78 °C. At -78 °C, n-BuLi (2.5 M, 4.43 mL, 2.5 eq.) was added dropwise to the mixture. The mixture was stirred at -78 °C for 1 hour. At approximately -78 °C, (3aR,4R,6R,6aR)-6-(2-diethoxyphosphorylethyl)-4-methoxy-2,2-dimethyl-3a,4,6,6a-tetrahydrofurano[3,4-d][1,3]dioxonol (1.5 g, 4.43 mmol, 1 equivalent) in THF (5 mL) was added to the mixture. The mixture was stirred at -78 °C for 10 minutes and then at room temperature for 2 hours. The mixture was quenched at room temperature by adding NH4Cl solution. The organic layer was separated, and the aqueous layer was extracted with EtOAc (50 mL). The organic layers were combined and concentrated to dryness, and the residue was purified by silica gel column chromatography, eluting with (EA / MeOH from 100:0 to 90:10) to give (3aR,6R,6aR)-6-[2-[diisopropoxyphosphorylmethyl(ethoxy)phosphoryl]ethyl]-4-methoxy-2,2-dimethyl-3a,4,6,6a-tetrahydrofurano[3,4-d][1,3]dioxonol (1 g, 47.74%). 1 H NMR (500MHz, CDCl3): δppm 1.25-1.40(m,21H),1.80-1.82(m,2H),1.94-2.05(m,2H),2.28-2.32(m,2H),3.27-3. 29(m,3H),4.05-4.09(m,3H),4.51-4.54(m,2H),4.68-4.70(m,2H),4.86-4.89(m,1H).

[0326] A solution of (2R,3S,4R,5R)-2-[2-[diisopropoxyphosphorylmethyl(ethoxy)phosphoryl]ethyl]-5-methoxy-tetrahydrofuran-3,4-diol (1.7 g, 3.93 mmol, 1 eq.) in 1,4-dioxane (7.6 mL) and aqueous sulfuric acid (1 M, 19 mL, 4.83 eq.) was heated under reflux for 2.5 h. After cooling the reaction to room temperature, the pH was adjusted to 7 with saturated NaHCO3 (38 mL), and the solution was concentrated to dryness. The residue was evaporated twice with dry THF to give a crude product, which did not require purification for the next step.

[0327] At 0 °C, acetic anhydride (5.0 mL) was added dropwise to a solution of (3R,4S,5R)-5-[2-[diisopropoxyphosphorylmethyl(ethoxy)phosphoryl]ethyl]tetrahydrofuran-2,3,4-triol in dry pyridine (25 mL). The reaction mixture was stirred at room temperature for 16 hours and then evaporated to dryness. The residue was adjusted to pH 2 with HCl solution (1 M, 15 mL). The organic layer was separated by treatment with DCM (50 mL), washed with brine (10 mL), dried, and concentrated. The residue was purified by silica gel column chromatography, eluting with (DCM / MeOH from 100:0 to 90:10) to give a buttery [(2R,3R,4R)-4,5-diacetoxy-2-[2-[diisopropoxyphosphorylmethyl(ethoxy)phosphoryl]ethyl]tetrahydrofuran-3-yl]acetate (1.5 g, 70.28%). 1 H NMR(500MHz,MeOD)δppm 1.34(dt,J=12.6,6.4,15H),1.93-2.17(m,13H),2.51-2.83(m,2H),4.13(dd,J=16.8,7.2,2H),4. 23(d,J=15.6,1H),4.66-4.80(m,2H),5.06-5.21(m,1H),5.30(t,J=15.2,1H),5.95-6.45(m,1H).

[0328] At -10°C, DBU (768.7 mg, 5.05 mmol, 572.93 μL, 1.1 eq.) and TMSOTf (2.25 g, 10.10 mmol, 1.83 mL, 2.2 eq.) were added to a solution of [(2R,3R,4R)-4,5-diacetoxy-2-[2-[diisopropoxyphosphorylmethyl(ethoxy)phosphoryl]ethyl]tetrahydrofuran-3-yl]acetate (2.5 g, 4.59 mmol, 1 eq.) and 2,6-dichloro-9H-purine (911.2 mg, 4.64 mmol, 1.05 eq.) in 25 mL of ACN. The mixture was stirred at 50°C for 1 hour. The reaction was monitored by TLC (EA / MeOH = 20:1) to indicate completion. The mixture was concentrated and purified by silica gel column chromatography, eluting with (EA / MeOH from 100:0 to 95:5) to give [(2R,3R,4R,5R)-4-acetoxy-5-(2,6-dichloropurin-9-yl)-2-[2-[diisopropoxyphosphorylmethyl(ethoxy)phosphoryl]ethyl]tetrahydrofuran-3-yl]acetate (1.5 g, 63.83%). 1 H NMR (500MHz, CDCl3): δppm 1.21-1.31(m,15H),1.65-2.06(m,10H),2.35-2.46(m,2H),4.07-4.14(m,2H) ,4.72-5.16(m,3H),5.28-5.77(m,1H),6.08-6.11(m,1H),8.40-9.20(m,1H).

[0329] DIPEA (143.94 mg, 1.11 mmol, 193.99 μL, 3 eq.) was added to a solution of [(2R,3R,4R,5R)-4-acetoxy-5-(2,6-dichloropurin-9-yl)-2-[2-[diisopropoxyphosphorylmethyl(ethoxy)phosphoryl]ethyl]tetrahydrofuran-3-yl] (250 mg, 371.24 μmol, 1 eq.) and benzylamine (47.74 mg, 445.49 μmol, 1.2 eq.) in 1,4-dioxane (2.5 mL). The mixture was stirred at 120 °C for 16 hours. The reaction was monitored by TLC (EtOAc:MeOH = 10:1) to indicate completion. The mixture was concentrated and purified by silica gel column chromatography, eluting with (EA / MeOH from 100:0 to 90:10) to give a yellow semi-solid [(2R,3R,4R,5R)-4-acetoxy-5-[6-(benzylamino)-2-chloro-purine-9-yl]-2-[2-[diisopropoxyphosphorylmethyl(ethoxy)phosphoryl]ethyl]tetrahydrofuran-3-yl]acetate (140 mg, 50.68%).

[0330] A solution of [(2R,3R,4R,5R)-4-acetoxy-5-[6-(benzylamino)-2-chloro-purine-9-yl]-2-[2-[diisopropoxyphosphorylmethyl(ethoxy)phosphoryl]ethyl]tetrahydrofuran-3-yl]acetate (140 mg, 188.14 μmol, 1 eq.) in NH3-MeOH (7 M, 1.5 mL, 57.56 eq.) was stirred for 2 hours at room temperature. The mixture was concentrated to remove the solvent, giving the crude product (2R,3R,4S,5R)-2-[6-(benzylamino)-2-chloro-purine-9-yl]-5-[2-[diisopropoxyphosphorylmethyl(ethoxy)phosphoryl]ethyl]tetrahydrofuran-3,4-diol, a yellow semi-solid. It was used directly in the next step.

[0331] Bromo(trimethyl)silane (432.05 mg, 2.82 mmol, 15 eq.) was added to a solution of (2R,3R,4S,5R)-2-[6-(benzylamino)-2-chloro-purin-9-yl]-5-[2-[diisopropoxyphosphorylmethyl(ethoxy)phosphoryl]ethyl]tetrahydrofuran-3,4-diol (124.18 mg, 188.14 μmol, 1 eq.) in DMF (4 mL). The mixture was stirred at 50 °C for 2 hours. The mixture was then quenched with TEAC solution until pH = 8 at 0 °C. The mixture was concentrated to dryness, H2O (5 mL) was added, and the mixture was purified by reversed-phase chromatography (C18 column) (H2O / ACN from 100:0 to 90:10) to give a-1 as a white solid (20 mg, 19.40%). 1 H NMR(500MHz,CD3OD)δppm:1.88-2.13(m,6H),4.04-4.07(m,1H),4.21-4.23(m,1H),4.61-4.63(m,1H),4 .77(s,2H),5.94(d,J=2.5Hz,1H),7.27-7.29(m,1H),7.33-7.36(m,2H),7.41-7.42(m,2H),8.30(s,1H); 13 C NMR (125MHz, CD3OD) δppm:27.85,28.11,28..89,31.15,31.75,32.07,32.66,45.09,74.36,75.62 ,86.30,86.43,69.41,119.67,128.33,128.84,129.56,139.98,141.01,151.25,155.72,156.40; 31 P NMR (203MHz, CD3OD) δppm: 14.99, 36.71; m / z (ESI+ ):548.0(M+H).

[0332] Example 20. Synthesis of compound a-9

[0333]

[0334] DIPEA (380.00 mg, 2.94 mmol, 512.12 μL, 3 eq.) was added to a solution of [(2R,3R,4R,5R)-4-acetoxy-5-(2,6-dichloropurin-9-yl)-2-[2-[diisopropoxyphosphorylmethyl(ethoxy)phosphoryl]ethyl]tetrahydrofuran-3-yl]acetate (660 mg, 980.08 μmol, 1 eq.) and [(3R,5S)-3,5-dimethyl-1-adamantyl]ammonium chloride (317.20 mg, 1.47 mmol, 1.5 eq.) in 1,4-dioxane (11 mL). The mixture was stirred at 130 °C for 16 hours. The reaction was monitored by TLC (EtOAc:MeOH = 10:1) to indicate completion. The mixture was concentrated and purified by silica gel column chromatography, eluting with (EA / MeOH from 100:0 to 92:8) to give a yellow semi-solid [(2R,3R,4R,5R)-4-acetoxy-5-[2-chloro-6-[[(3R,5S)-3,5-dimethyl-1-adamantyl]amino]purine-9-yl]-2-[2-[diisopropoxyphosphorylmethyl(ethoxy)phosphoryl]ethyl]tetrahydrofuran-3-yl]acetate (310 mg, 38.75%). 1 H NMR (500MHz, CD3OD): δppm: 0.90 (s, 6H), 1.13-1.50 (m, 20H), 1.79-2.26 (m, 14H), 2.64 (dd, J=31.9, 15.0Hz, 2H), 4.10 (qd, J=14.4 ,7.1Hz,2H),4.64-4.79(m,2H),5.50-5.61(m,1H),5.83-5.94(m,1H),6.09(d,J=4.9Hz,1H),6.99(s,1H),8.18(d,J=6.0Hz,1H).

[0335] [(2R,3R,4R,5R)-4-acetoxy-5-[2-chloro-6-[[(3R,5S)-3,5-dimethyl-1-adamantyl]amino]purine-9-yl]-2-[2-[diisopropoxyphosphorylmethyl(ethoxy)phosphoryl]ethyl]tetrahydrofuran-3-yl]acetate (310 mg, 379.78 μmol, 1 eq.) in NH3-MeOH (7 M, 6 mL, 110.59 eq.) was stirred for 3 hours at room temperature. The mixture was concentrated to remove the solvent. EtOA (35 mL) was added to the residue and the mixture was washed with brine. The organic phase was dried with Na₂SO₄ and concentrated to give the crude product (2R,3R,4S,5R)-2-[2-chloro-6-[[(3R,5S)-3,5-dimethyl-1-adamantyl]amino]purine-9-yl]-5-[2-[diisopropoxyphosphorylmethyl(ethoxy)phosphoryl]ethyl]tetrahydrofuran-3,4-diol (250 mg, 89.91%), as a yellow semi-solid. This solid was used directly in the next step.

[0336] Bromo(trimethyl)silane (993.18 mg, 6.49 mmol, 856.19 μL, 19 eq.) was added to (2R,3R,4S,5R)-2-[2-chloro-6-[[(3R,5S)-3,5-dimethyl-1-adamantyl]amino]purine-9-yl]-5-[2-[diisopropoxyphosphorylmethyl(ethoxy)phosphoryl]ethyl]tetrahydrofuran-3,4-diol (250 mg, 341 eq.). 44 μmol (1 eq.) was added to a solution of DMF (8 mL). The mixture was stirred at 50 °C for 2 hours. The mixture was then quenched with TEAC solution until pH = 8 at 0 °C. The mixture was concentrated to dryness, H2O (8 mL) was added, and the compound was purified by reversed-phase chromatography (C18 column) (H2O / ACN from 100:0 to 90:10) to give a-9 as a white solid (101 mg, 41.02%). 1 H NMR(500MHz,CD3OD)δppm:0.89(s,6H),1.13-1.23(m,2H),1.35(d,J=12.3H z,2H),1.45(d,J=12.0Hz,2H),1.81(d,J=11.8Hz,2H),1.85-2.01(m,5H),2 .03-2.15(m,5H),2.18(d,J=2.9Hz,1H),4.02(dd,J=12.2,5.3Hz,1H),4.17 (t,J=5.1Hz,1H),4.57(t,J=5.1Hz,1H),5.89(d,J=4.8Hz,1H),8.26(s,1H); 13C NMR (125MHz, CD3OD) δppm: 27.68, 27.94, 28.72, 30.75, 31.69, 33.44, 40.72, 43.75, 51. 75,55.91,74.37,75.65,86.15,86.29,89.47,119.70,140.55,150.78,154.97,155.88; 31 P NMR (203MHz, CD3OD) δppm: 14.98, 38.21; m / z (ESI + ):620.1(M+H).

[0337] Example 21. Synthesis of compound a-19

[0338]

[0339] DIPEA (143.94 mg, 1.11 mmol, 193.99 μL, 3 eq.) was added to a solution of [(2R,3R,4R,5R)-4-acetoxy-5-(2,6-dichloropurin-9-yl)-2-[2-[diisopropoxyphosphorylmethyl(ethoxy)phosphoryl]ethyl]tetrahydrofuran-3-yl]acetate (250 mg, 371.24 μmol, 1 eq.) and dodecane-1-amine (82.57 mg, 445.49 μmol, 1.2 eq.) in 1,4-dioxane (2.5 mL). The mixture was stirred at 120 °C for 16 hours. The reaction was monitored by TLC (EtOAc:MeOH = 10:1) to indicate completion. The mixture was concentrated and purified by silica gel column chromatography, eluting with (EA / MeOH from 100:0 to 90:10) to give a yellow semi-solid [(2R,3R,4R,5R)-4-acetoxy-5-[2-chloro-6-(dodecylamino)purine-9-yl]-2-[2-[diisopropoxyphosphorylmethyl(ethoxy)phosphoryl]ethyl]tetrahydrofuran-3-yl]acetate (150 mg, 49.14%).

[0340] A solution of [(2R,3R,4R,5R)-4-acetoxy-5-[2-chloro-6-(dodecylamino)purine-9-yl]-2-[2-[diisopropoxyphosphorylmethyl(ethoxy)phosphoryl]ethyl]tetrahydrofuran-3-yl]acetate (150 mg, 182.41 μmol, 1 eq.) in NH3-MeOH (7 M, 1.5 mL, 57.56 eq.) was stirred for 2 hours at room temperature. The mixture was concentrated to remove the solvent, giving the crude product (2R,3R,4S,5R)-2-[2-chloro-6-(dodecylamino)purine-9-yl]-5-[2-[diisopropoxyphosphorylmethyl(ethoxy)phosphoryl]ethyl]tetrahydrofuran-3,4-diol, a yellow semi-solid. This solid was used directly in the next step.

[0341] Bromo(trimethyl)silane (418.90 mg, 2.74 mmol, 15 eq.) was added to a solution of (2R,3R,4S,5R)-2-[2-chloro-6-(dodecylamino)purine-9-yl]-5-[2-[diisopropoxyphosphorylmethyl(ethoxy)phosphoryl]ethyl]tetrahydrofuran-3,4-diol (134.66 mg, 182.41 μmol, 1 eq.) in DMF (4 mL). The mixture was stirred at 50 °C for 2 h. The mixture was then quenched with TEAC solution until pH = 8 at 0 °C. The mixture was concentrated to dryness, H2O (5 mL) was added, and the compound was purified by reversed-phase chromatography (C18 column) (H2O / ACN from 100:0 to 70:30) to give a-19 as a white solid (20 mg, 17.51%). 1 H NMR(500MHz,CD3OD)δppm:0.86-0.089(m,3H),1.26-1.30(m,18H),1.64-1.67(m,2H),1.92-2.17(m,6H),1.81(d,J=11.8Hz,2H),1.85-2.01(m,5H) ,2.03-2.15(m,5H),3.51-3.54(m,2H),4.05-4.06(d,J=5.0,1H),4.19(t ,J=5.0Hz,1H),4.59(t,J=5.0Hz,1H),5.91(d,J=5.0Hz,1H),8.25(s,1H); 13C NMR (125MHz, CD3OD) δppm: 13.13, 22.33, 26.17, 26.45, 28.87, 28.97, 29.04, 29.26, 29. 32,31.64,40.20,72.95,74.25,84.86,87.88,118.09,139.24,149.41,154.44,155.17; 31 P NMR (203MHz, CD3OD) δppm: 15.01, 39.35; m / z (ESI + ):626.2(M+H).

[0342] Example 22. Synthesis of compound a-31

[0343]

[0344] DIPEA (1.01 g, 7.80 mmol, 1.36 mL, 3.0 eq.) was added to a solution of [(2R,3R,4R,5R)-4-acetoxy-5-(2,6-dichloropurin-9-yl)-2-[2-[diisopropoxyphosphorylmethyl(ethoxy)phosphoryl]ethyl]tetrahydrofuran-3-yl]acetate (1.5 g, 2.23 mmol, 1 eq.) and 2-azacyclo[4.5]decane (465.22 mg, 3.34 mmol, 1.5 eq.) in 1,4-dioxane (15 mL). The mixture was stirred at 130 °C for 16 hours. The reaction was monitored by TLC (EtOAc:MeOH = 20:1) to indicate completion. The mixture was concentrated and purified by silica gel column chromatography, eluting with (EA / MeOH from 100:0 to 95:5) to give [(2R,3R,4R,5R)-4-acetoxy-5-[6-(2-azacyclo[4.5]decaalkyl-2-yl)-2-chloro-purine-9-yl]-2-[2-[diisopropoxyphosphorylmethyl(ethoxy)phosphoryl]ethyl]tetrahydrofuran-3-yl]acetate (950 mg, 54.95%). 1H NMR (500MHz, CD3OD): δppm 1.26-1.36(m,17H),1.51-1.53(m,2H),1.83-2.17(m,12H),2.62-2.69(m,2H),3.39-3.53(m,1H),3.71-3.73(m,1H),3.93-3. 95(m,1H),4.13-4.27(m,4H),4.69-4.71(m,2H),5.58-5.59(m,1H),5.91-5.93(m,1H),6.10-6.12(m,1H),6.15-6.17(m,1H).

[0345] A solution of [(2R,3R,4R,5R)-4-acetoxy-5-[6-(2-azacyclo[4.5]decaalkyl-2-yl)-2-chloro-purine-9-yl]-2-[2-[diisopropoxyphosphorylmethyl(ethoxy)phosphoryl]ethyl]tetrahydrofuran-3-yl]acetate (950 mg, 1.22 mmol, 1 eq.) in NH3-MeOH (7 M, 10 mL, 57.19 eq.) was prepared. The mixture was stirred at room temperature for 16 hours. The mixture was then concentrated by adding EtOA (30 mL). The organic layer was washed with an aqueous NaCl solution, dried with Na2SO4, filtered, and evaporated to dryness to give (2R,3R,4S,5R)-2-[6-(2-azacyclo[4.5]decaalkyl-2-yl)-2-chloro-purine-9-yl]-5-[2-[diisopropoxyphosphorylmethyl(ethoxy)phosphoryl]ethyl]tetrahydrofuran-3,4-diol (650 mg, 76.73%). 1H NMR (500MHz, CD3OD): δppm 1.27-1.36(m,15H),1.52-1.54(m,10H),1.84-1.94(m,2H),2.05-2.18(m,4H),2.61-2.68(m,2H),3.52-3.54(m,1H),3.72 -3.74(m,1H),3.88-4.03(m,2H),4.12-4.14(m,3H),4.27-4.29(m,1H),4.62-4.73(m,3H),5.89(s,1H),8.14-8.15(m,1H).

[0346] At 0 °C, bromo(trimethyl)silane (331.79 mg, 2.17 mmol, 15 eq.) was added to a solution of (2R,3R,4S,5R)-2-[6-(2-azacyclo[4.5]decaalkyl-2-yl)-2-chloro-purin-9-yl]-5-[2-[diisopropoxyphosphorylmethyl(ethoxy)phosphoryl]ethyl]tetrahydrofuran-3,4-diol (100 mg, 144.48 μmol, 1 eq.) in DMF (4 mL). The mixture was stirred at 50 °C for 2 hours. The mixture was then quenched with TEAC solution until pH = 8 at 0 °C. The mixture was concentrated to dryness, H2O (10 mL) was added, and then purified by reversed-phase chromatography (C18 column) (H2O / ACN from 100:0 to 90:10) to give product a-31 (20 mg, 20.32%). 1H NMR (CD3OD, 500MHz): δppm 1.51-1.57(m,10H),1.83-2.10(m,8H),3.51-3.53(m,1H),3.71-3.73(m,1H),3.95-3.97(m,1H),4 .03-4.04(m,1H),4.16-4.19(m,2H),4.57-4.59(m,1H),5.90-5.91(d,1H),8.19-8.20(m,1H); 13C NMR(CD3OD,125MHz): δppm 23.02,25.87,26.18,26.40,27.18,29.21,29.80,30.13,30.73,34.89,40.37,4 2.67,72.94,74.05,84.70,87.93,118.71,138.65,150.96,153.21,153.83; 31P NMR (CD3OD, 203MHz): δ15.12, 38.73; m / z (ESI+): 580.1 (M+H).

[0347] Example 23. Synthesis of compound c-13

[0348]

[0349] 1 N H₂SO₄ (1 M, 25 mL, 3.38 eq) was added to a mixture of (3aR,4R,6R,6aR)-6-(2-diethoxyphosphorylethyl)-4-methoxy-2,2-dimethyl-3a,4,6,6a-tetrahydrofurano[3,4-d][1,3]dioxonol (2.50 g, 7.39 mmol, 1 eq.) in 1,4-dioxane (10 mL), and the mixture was heated under reflux for 2.5 h. The pH was adjusted to 7 with an aqueous solution of NaHCO₃ (50 mL), and the mixture was concentrated to dryness. Ac₂O (836.84 mg, 8.20 mmol, 4.2 mL) was added to the residue in pyridine (21 mL), and the mixture was stirred at room temperature for 16 h. The resulting solution was evaporated to dryness. The residue was purified by silica gel column chromatography, eluting with (DCM / MeOH from 100:0 to 95:05) to give [(2R,3R,4R)-4,5-diacetoxy-2-(2-diethoxyphosphorylethyl)tetrahydrofuran-3-yl]acetate (1.9 g, 62.67%). 1 H NMR (500MHz, CDCl3): δppm 1.36-1.54(m,6H), 1.81-2.30(m,13H), 1.94-2.05(m,2H), 4.13-4.31(m,5H), 4.90-5.23(m,2H), 6.16-6.41(m,1H).

[0350] At -10°C, DBU (775.39 mg, 5.09 mmol, 760.18 μL, 1.1 eq) and TMSOTf (2.26 g, 10.19 mmol, 1.84 mL, 2.2 eq) were added to a solution of [(2R,3R,4R)-4,5-diacetoxy-2-(2-diethoxyphosphorylethyl)tetrahydrofuran-3-yl]acetate (1.9 g, 4.63 mmol, 1 eq) in 219 mL of ACN. The mixture was stirred at 50°C for 1 hour. The mixture was concentrated and purified by silica gel column chromatography, eluting with (EA / MeOH from 100:0 to 95:5) to give [(2R,3R,4R,5R)-4-acetoxy-5-(2,6-dichloropurin-9-yl)-2-(2-diethoxyphosphorylethyl)tetrahydrofuran-3-yl]acetate (1.3 g, 52.06%). 1 H NMR (500MHz, CDCl3): δppm 1.24-1.34(m,6H),2.01-2.14(m,10H),4.09-4.12(m,5H),5.56-5.58(m,1H),5.88-5.90(m,1H),6.24-6.25(d,1H),8.68(s,1H).

[0351] DIPEA (934.61 mg, 7.23 mmol, 1.26 mL, 3 eq.) was added to a solution of [(2R,3R,4R,5R)-4-acetoxy-5-(2,6-dichloropurin-9-yl)-2-(2-diethoxyphosphorylethyl)tetrahydrofuran-3-yl]acetate (1.3 g, 2.41 mmol, 1 eq.) and [(3R,5S)-3,5-dimethyl-1-adamantyl]ammonium chloride (624.12 mg, 2.89 mmol, 1.2 eq.) in 1,4-dioxane (13 mL). The mixture was stirred at 130 °C for 16 hours. The reaction was monitored by TLC until completion. The mixture was concentrated and purified by silica gel column chromatography, eluting with (EA / MeOH from 100:0 to 90:10) to give [(2R,3R,4R,5R)-4-acetoxy-5-[2-chloro-6-[[(3R,5S)-3,5-dimethyl-1-adamantyl]amino]purine-9-yl]-2-(2-diisopropoxyphosphorylethyl)tetrahydrofuran-3-yl]acetate (700 mg, 42.57%). 1 H NMR (500MHz, CD3OD): δppm 0.84-0.91(m,6H),1.20-1.32(m,9H),1.36-1.52(m,4H),1.82-2.01(m,6H),2.07-2.14(m,10 H),4.08-4.11(m,5H),5.57-5.60(m,1H),5.87-5.89(m,1H),6.09-6.10(m,1H),8.16(m,1H).

[0352] NH3-MeOH (7M, 7 mL, 47.75 eq.) was added to a solution of [(2R,3R,4R,5R)-4-acetoxy-5-[2-chloro-6-[[(3R,5S)-3,5-dimethyl-1-adamantyl]amino]purine-9-yl]-2-(2-diisopropoxyphosphorylethyl)tetrahydrofuran-3-yl]acetate (700 mg, 1.03 mmol, 1 eq). The mixture was stirred at room temperature for 16 hours. The mixture was then concentrated by adding EtOA (30 mL). The organic layer was washed with an aqueous NaCl solution, dried with Na2SO4, filtered, and evaporated to give (2R,3R,4S,5R)-2-[2-chloro-6-[[(3R,5S)-3,5-dimethyl-1-adamantyl]amino]purine-9-yl]-5-(2-diisopropoxyphosphorylethyl)tetrahydrofuran-3,4-diol (400 mg, 65.18%). 1H NMR (500MHz, CD3OD): δppm 0.90-0.92(m,6H),1.20-1.26(m,2H),1.29-1.32(m,6H),1.36-1.39( m,2H),1.46-1.48(m,2H),1.82-1.84(m,2H),1.91-1.93(m,3H),1.96- 2.05(m,3H),2.10-2.12(m,2H),2.20-2.21(m,1H),4.03-4.11(m,5H), 4.27-4.29(m,1H),4.69-4.71(m,1H),5.86-5.87(m,1H),8.15(s,1H).

[0353] At 0 °C, bromo(trimethyl)silane (383.97 mg, 2.51 mmol, 331.01 μL, 15 eq) was added to a solution of (2R,3R,4S,5R)-2-[2-chloro-6-[[(3R,5S)-3,5-dimethyl-1-adamantyl]amino]purine-9-yl]-5-(2-diisopropoxyphosphorylethyl)tetrahydrofuran-3,4-diol (100 mg, 167.20 μmol, 1 eq) in 4 mL of DMF. The mixture was stirred at 50 °C for 2 hours. The mixture was then quenched with TEAC solution until pH = 8 at 0 °C. The mixture was concentrated, H2O (510 mL) was added, and the compound was then purified by reversed-phase chromatography (C18 column) (H2O / ACN from 100:0 to 70:30) to give c-13 (40 mg, 37.20%). 1 H NMR(CD3OD,500MHz): δppm 1.19-1.26(m,2H),1.36-1.38(m,2H),1.45-1.48(m,2H),1.63-1.74(m,2H),1.82-1.84(m,2H),1.90-1.95(m,2H),1.98- 2.10(m,4H),2.19-2.21(m,1H),4.02-4.03(m,1H),4.15-4.17(m,1H),4.55-4.56(m,1H),5.90-5.91(d,1H),8.22(s,1H). 13 C NMR(CD3OD,125MHz): δppm 25.41,26.49,29.21,30.75,31.70,33.45,40.71,43.76,51.76,55.91,74 .60,75.76,85.99,86.13,89.71,119.74,140.42,150.68,154.98,155.88; 31P NMR (CD3OD, 203MHz): δppm 24.12; m / z (ESI - ):540.2 (MH).

[0354] Example 24. Synthesis of compound d-1

[0355]

[0356] A mixture of indane-1-carboxylic acid (5 g, 30.83 mmol, 1 eq.) and H₂SO₄ (3.02 g, 30.83 mmol, 1.64 mL, 1 eq.) in MeOH (100 mL) was heated at 65 °C for 20 h. The reaction was indicated by TLC analysis. The resulting solvent was evaporated, and the residue was diluted with EtOAc (30 mL). The residue was then washed with brine, dried over Na₂SO₄, filtered, and evaporated to dryness to give the product methyl indane-1-carboxylate (4.7 g, 86.5%).

[0357] Under a nitrogen atmosphere at 0°C, K₂CO₃ (11.06 g, 80.02 mmol, 3 eq.) and HCHO (7.37 g, 80.02 mmol, 37% purity, 3 eq.) were added to methyl indane-1-carboxylate (4.7 g, 26.67 mmol, 1 eq.) in a solution of DMSO (47 mL). The mixture was stirred at room temperature for 19 hours, and the resulting solution was quenched with H₂O (50 mL). The mixture was extracted with EtOAc (50 mL). The aqueous phase was adjusted to pH 3 with 2.5 N hydrochloric acid and extracted with EtOAc (50 mL). The organic layer was washed with brine, dried over Na₂SO₄, filtered, and evaporated to dryness to give the product 1-(hydroxymethyl)indane-1-carboxylic acid (3.3 g, 64.4%).

[0358] EDCI (2.99 g, 15.61 mmol, 1.5 eq.), HOBT (2.11 g, 15.61 mmol, 1.5 eq.), and DIPEA (2.02 g, 15.61 mmol, 2.72 mL, 1.5 eq.) were added to a mixture of 1-(hydroxymethyl)indane-1-carboxylic acid (2 g, 10.41 mmol, 1 eq.) and (4-methoxyphenyl)methylamine (1.43 g, 10.41 mmol, 1 eq.) in DMF (20 mL). The mixture was stirred at room temperature for 18 hours. The reaction was indicated by TLC. The resulting solution was quenched with H2O (10 mL) and diluted with EtOAc (80 mL). The organic layer was washed with brine, dried over Na2SO4, filtered, and evaporated to dryness. The residue was purified by silica gel column chromatography (DCM / MeOH, from 100:0 to 95:5) to give 1-(hydroxymethyl)-N-[(4-methoxyphenyl)methyl]indane-1-carboxamide (2.4 g, 74.1%) in oil form.

[0359] Under a nitrogen atmosphere, PPh3 (2.78 g, 10.60 mmol, 1.5 eq.) and DEAD (1.85 g, 10.60 mmol, 1.67 mL) were sequentially added dropwise to a solution of 1-(hydroxymethyl)-N-[(4-methoxyphenyl)methyl]indane-1-carboxamide (2.2 g, 7.07 mmol, 1 eq.) in 55 mL of THF, while maintaining the temperature at 0 °C. The mixture was stirred at room temperature for 2 hours. TLC analysis showed... The reaction was complete. The resulting solution was quenched with 2 mL of H₂O. Then 20 mL of EtOAc was added. The organic layer was separated, washed with brine, dried over Na₂SO₄, filtered, and evaporated to dryness. The residue was purified by silica gel column chromatography (DCM / MeOH, from 100:0 to 98:2 to 98:3) to give 1.7 g (82.1%) of 1-[(4-methoxyphenyl)methyl]-spiro-[azacyclobutane-3,1'-indane]-2-one as a yellow solid.

[0360] At 0 °C, cerium ammonium nitrate (9.16 g, 16.70 mmol, 3.5 eq.) was added to a solution of 1-[(4-methoxyphenyl)methyl]-spiro-[azacyclobutane-3,1'-indane]-2-one (1.4 g, 4.77 mmol, 1 eq.) in acetonitrile (36 mL) and H₂O (4 mL). The mixture was stirred at room temperature for 16 hours. The reaction was indicated by TLC. The resulting solution was diluted with EtOAc (50 mL). The organic phase was separated, washed with water and brine, dried over Na₂SO₄, filtered, and evaporated to dryness. The residue was purified by silica gel column chromatography using PE / EA (eluting from 100:0 to 45:55) to give spirocyclic [azacyclobutane-3,1'-indane]-2-one (160 mg, 19.4%) as a yellow solid.

[0361] At 0 °C, LiAlH4 (39.44 mg, 1.04 mmol, 1 eq.) was added to a solution of spirocyclic [azacyclobutane-3,1'-indane]-2-one (180.00 mg, 1.04 mmol, 1 eq.) in THF (10 mL). The mixture was stirred overnight at 70 °C. The mixture was quenched with H2O (1 mL), followed by the addition of EtOAc (10 mL). The solid material was removed by filtration, and the organic phase was concentrated. The residue was purified by silica gel column chromatography (DCM / MeOH (100:0–80:20)) to give the product spirocyclic [azacyclobutane-3,1'-indane] (80 mg, 48.4%) as an oil.

[0362] Spiro[azacyclobutane-3,1'-indane] (80 mg, 502.43 μmol, 1 eq.) and DIPEA (162.33 mg, 1.26 mmol, 218.78 μL, 2.5 eq.) were added to a solution of [(2R,3R,4R,5R)-3,4-diacetoxy-5-(2,6-dichloropurin-9-yl)tetrahydrofuran-2-yl]methyl acetate (224.70 mg, 502.43 μmol, 1 eq.) in 1,4-dioxane (8 mL). The mixture was stirred overnight at 100 °C. TLC showed that the starting material was consumed and two new products were generated. The mixture was concentrated, and the residue was purified by silica gel column chromatography (elution, PE / EtOAc, from 100:0 to 60:40) to give [(2R,3R,4R,5R)-3,4-diacetoxy-5-(2-chloro-6-spirocyclic [azacyclobutane-3,1'-indane]-1-yl-purin-9-yl)tetrahydrofuran-2-yl]methyl acetate (100 mg, 34.9%).

[0363] NH3-MeOH (7M, 601.51 μL, 20 eq.) was added to a solution of [(2R,3R,4R,5R)-3,4-diacetoxy-5-(2-chloro-6-spiro[azacyclobutane-3,1'-indane]-1-yl-purine-9-yl)tetrahydrofuran-2-yl]methyl acetate (120 mg, 210.53 μmol, 1 eq.) in MeOH (2 mL). The mixture was stirred at room temperature for 4 hours. The solvent was removed by evaporation, and the residue was diluted with EtOAc (30 mL), washed with brine (30 mL), and dried over Na2SO4. The organic layer was concentrated to obtain the crude product (2R,3R,4S,5R)-2-(2-chloro-6-spirocyclic [azacyclobutane-3,1'-indane]-1-yl-purin-9-yl)-5-(hydroxymethyl)tetrahydrofuran-3,4-diol (90 mg, 202.76 μmol, 96.31% yield) as a pale yellow solid, which could be used without further purification.

[0364] 2,2-Dimethoxypropane (187.70 mg, 1.80 mmol, 10 eq.) and TsOH-H2O (35.69 mg, 180.23 μmol, 1.0 eq.) were added to a solution of (2R,3R,4S,5R)-2-(2-chloro-6-spirocyclic [azacyclobutane-3,1'-indane]-1-yl-purin-9-yl)-5-(hydroxymethyl)tetrahydrofuran-3,4-diol (80 mg, 180.23 μmol, 1 eq.) in acetone (20 mL). The mixture was stirred overnight at room temperature. The mixture was diluted with EtOAc (30 mL) and washed with an aqueous solution of NaHCO3 (40 mL) and brine (40 mL). The organic layer was concentrated, and the residue was purified by silica gel column chromatography (PE / EtOAc (100:0-45:55)) to give [(3aR,4R,6R,6aR)-4-(2-chloro-6-spirocyclic [azacyclobutane-3,1'-indane]-1-yl-purin-9-yl)-2,2-dimethyl-3a,4,6,6a-tetrahydrofurano[3,4-d][1,3]dioxono-6-yl]methanol (70 mg, 80.2%).

[0365] A solution of [(3aR,4R,6R,6aR)-4-(2-chloro-6-spiro[azacyclobutane-3,1'-indane]-1-yl-purin-9-yl)-2,2-dimethyl-3a,4,6,6a-tetrahydrofurano[3,4-d][1,3]dioxono-6-yl]methanol (70 mg, 144.64 μmol, 1 eq.) in PO(MeO)3 (4 mL) was cooled to approximately 0 °C, and then bis(dichlorophosphoryl)methane (72.26 mg, 289.29 μmol, 2.0 eq.) in PO(MeO)3 (1 mL) was added. The mixture was stirred at 0 °C for 6 hours. Water (3 mL) was added, and the mixture was further stirred overnight at room temperature. The reaction mixture was purified by reverse-phase C-18 silica gel column purification (0–30% ACN in water) to give compound d-1 (41 mg, yield 44.5%). 1 H NMR(500MHz,CD3OD)δppm 2.47(dt,J=31.3,12.1Hz,4H),2.96(t,J=7.1Hz,2H),4.24(s,1H),4.28(dd,J=11.2,6.5Hz,1H),4.34(d,J=3.6Hz,1H),4 .42(t,J=4.8Hz,1H),4.64(t,J=5.0Hz,1H),6.00(d,J=5.0Hz,1H),7.16-7.31(m,3H),7.48(d,J=7.3Hz,1H),8.38(s,1H); 13 C NMR (125MHz, CD3OD) δppm31.23,39.74,65.95,71.42,75.67,84.73,89.71,119 .06,123.26,125.49,128.29,128.80,141.30,144.69,147.40,151.90,155.69; 31 P NMR (203MHz, CD3OD) δppm 16.89,19.83; m / z (ESI + ):602.1(M+H).

[0366] Example 25. Synthesis of compound d-2

[0367]

[0368] Potassium tert-butoxide (1 M, 26.64 mL, 2.0 eq.) was added to a solution of 4-fluoroindene-1-one (2 g, 13.32 mmol, 1 eq.) cooled at 0 °C in EtOH (20 mL) and THF (20 mL), followed by the addition of TOSMIC (3.90 g, 19.98 mmol, 1.5 eq.) in EtOH / THF (1:1, 20 mL). The mixture was stirred overnight at room temperature, cooled to 0 °C, and then brine was added. The mixture was extracted with EtOAc, and the extract was concentrated. The residue was purified by silica gel column chromatography (elution: PE / EA = 100:0-83:17) to give the product 4-fluoroindene-1-carbonitrile (910 mg, 5.65 mmol, yield 42.39%).

[0369] 4-Fluorindene-1-carbamate (910 mg, 5.65 mmol, 1 eq.) was added to a solution of sodium hydroxide (677.52 mg, 16.94 mmol, 3.0 eq.) in water (15 mL). The mixture was stirred at 100 °C for 16 h. The mixture was cooled to room temperature, diluted with water (30 mL), and extracted with EtOAc (2 x 40 mL). The aqueous layer was adjusted to pH 2 and extracted with EtOAc (2 x 50 mL). The organic layers were combined and concentrated to dryness to give 4-fluoroindene-1-carboxylic acid (905 mg, 5.02 mmol, yield 88.96%) as a brown solid.

[0370] At room temperature, thionyl chloride (3.30 g, 27.75 mmol, 2.02 mL, 5 eq.) was added to 4-fluoroindane-1-carboxylic acid (1 g, 5.55 mmol, 1 eq.) in a solution of methanol (20 mL) and DMF (0.1 mL). After the addition was complete, the reaction mixture was stirred at room temperature for 3 hours, then concentrated, quenched with ice water (50 mL), and extracted with EtOAc (2 x 50 mL). The combined organic layers were washed with brine, dried over Na2SO4, and concentrated to dryness to give methyl 4-fluoroindane-1-carboxylic acid (1.02 g, 5.25 mmol, yield 94.63%).

[0371] Under a nitrogen atmosphere at 0 °C, K₂CO₃ (2.18 g, 15.76 mmol, 3 eq.) and HCHO (941.54 mg, 10.50 mmol, 38% purity, 2 eq.) were added to a mixture of methyl 4-fluoroindane-1-carboxylic acid (1.02 g, 5.25 mmol, 1 eq.) in DMSO (25 mL). The mixture was stirred at room temperature for 16 hours. The reaction was quenched with H₂O (75 mL). The mixture was extracted with ethyl acetate (50 mL). The pH of the aqueous solution was adjusted to 3 with 3N HCl. The mixture was extracted with ethyl acetate (3 × 80 mL). The combined organic layers were washed with brine, dried over Na₂SO₄, filtered, and evaporated to dryness to give 4-fluoro-1-(hydroxymethyl)indane-1-carboxylic acid (1.02 g, 4.85 mmol, yield 92.39%).

[0372] To a mixture of 4-fluoro-1-(hydroxymethyl)indane-1-carboxylic acid (1.02 g, 4.85 mmol, 1 eq.) and benzylamine (519.96 mg, 4.85 mmol, 1 eq.) in DMF (10 mL), EDCI (1.40 g, 7.28 mmol, 1.5 eq.), HOBT (2.29 g, 16.91 mmol, 1.5 eq.), and DIPEA (627.14 mg, 4.85 mmol, 845.20 μL, 1 eq.) were added. The mixture was stirred at room temperature for 18 hours, and the solvent was removed by evaporation. Water (50 mL) and EA (60 mL) were then added. The organic layer was washed with brine, dried over Na2SO4, filtered, and evaporated to dryness. The residue was purified by silica gel column chromatography (elution, PE / EA = 100:0 -50:50) to give N-benzyl-4-fluoro-1-(hydroxymethyl)indane-1-carboxamide (1.25 g, 4.18 mmol, yield 86.06%).

[0373] Under a nitrogen atmosphere at 0 °C, PPh3 (657.17 mg, 2.51 mmol, 1.5 eq.) was added to a mixture of N-benzyl-4-fluoro-1-(hydroxymethyl)indane-1-carboxamide (500 mg, 1.67 mmol, 1 eq.) and tetrahydrofuran (10 mL), followed by dropwise addition of DEAD (436.35 mg, 2.51 mmol, 394.53 μL, 1.5 eq.). The mixture was stirred at room temperature for 2 hours. The reaction was quenched with water (20 mL), followed by the addition of ethyl acetate (60 mL). The organic layer was washed with brine, dried over Na2SO4, filtered, and evaporated to dryness. The residue was purified by silica gel column chromatography (elution, PE / EA = 100:0 to 67:33) to give 1-benzyl-4'-fluorospirocyclic [azacyclobutane-3,1'-indane]-2-one (380 mg, 1.35 mmol, yield 80.87%).

[0374] At 0 °C, lithium aluminum hydride (153.78 mg, 4.05 mmol, 3 eq.) was added to a solution of aluminum trichloride (360.22 mg, 2.70 mmol, 2 eq.) cooled at 0 °C in tetrahydrofuran (4 mL). The mixture was stirred at 0 °C for 30 min, and then 1-benzyl-4'-fluoro-spirocyclic [azacyclobutane-3,1'-indanane]-2-one (380 mg, 1.35 mmol, 1 eq.) was added to tetrahydrofuran (3 mL). The mixture was stirred at room temperature for 16 h, diluted with EtOAc (20 mL), and then the pH was adjusted to 10 by slowly adding 15% sodium hydroxide aqueous solution at 0 °C. The organic layer was separated and dried over MgSO4. Insoluble matter was removed by filtration, and the filtrate was concentrated to dryness. The residue was purified by silica gel column chromatography (elution, PE / EA = 100:0 to 67:33) to give 1-benzyl-4'-fluoro-spirocyclic [azacyclobutane-3,1'-indane] (280 mg, 1.05 mmol, yield 77.54%).

[0375] Ammonium formate (99.07 mg, 1.57 mmol, 1.5 eq.) and Pd(OH)₂ (64.64 mg, 523.69 μmol, 0.5 eq.) were added to a methanol (15 mL) solution of 1-benzyl-4'-fluorospirocyclic [azacyclobutane-3,1'-indane] (280 mg, 1.05 mmol, 1 eq.). The mixture was stirred overnight at 60 °C under hydrogen atmosphere. The insoluble residue was filtered off and washed with methanol. The filtrate and washings were combined and concentrated to dryness to give the product 4'-fluorospirocyclic [azacyclobutane-3,1'-indane] (148 mg, 835.13 μmol, yield 79.74%).

[0376] Methyl acetate [(2R,3R,4R,5R)-3,4-diacetoxy-5-(2,6-dichloropurin-9-yl)tetrahydrofuran-2-yl]acetate (340 mg, 760.24 μmol, 1 eq.) in 1,4-dioxane (10 mL) was mixed with 4'-fluorospirocyclic [azacyclobutane-3,1'-indane] (148.20 mg, 836.27 μmol, 1.1 eq.) and DIPEA (442.14 mg, 3.42 mmol, 595.88 μL, 4.5 eq.). The mixture was stirred at 100 °C for 3 hours.

[0377] The mixture was concentrated, and the residue was purified by silica gel column chromatography (elution, PE / EA = 100:0-60:40) to give the product [(2R,3R,4R,5R)-3,4-diacetoxy-5-[2-chloro-6-(4'-fluorospirocyclic [azacyclobutane-3,1'-indane]-1-yl)purin-9-yl]tetrahydrofuran-2-yl]acetate (290 mg, 493.21 μmol, yield 64.88%).

[0378] Methyl acetate [(2R,3R,4R,5R)-3,4-diacetoxy-5-[2-chloro-6-(4'-fluorospirocyclo[azacyclobutane-3,1'-indane]-1-yl)purin-9-yl]tetrahydrofuran-2-yl]acetate (290 mg, 493.21 μmol, 1 eq.) was added to a mixture of methanol (5 mL) with NH3-MeOH (7 M, 1.76 mL, 25 eq.). The mixture was stirred at room temperature for 2 hours and concentrated to dryness to give (2R,3R,4S,5R)-2-[2-chloro-6-(4'-fluorospirocyclo[azacyclobutane-3,1'-indane]-1-yl)purin-9-yl]-5-(hydroxymethyl)tetrahydrofuran-3,4-diol (227 mg, 491.48 μmol, 99.65% yield). It can be used directly in the next step.

[0379] To a solution of (2R,3R,4S,5R)-2-[2-chloro-6-(4'-fluorospiro[azacyclobutane-3,1'-indane]-1-yl)purin-9-yl]-5-(hydroxymethyl)tetrahydrofuran-3,4-diol (227 mg, 491.48 μmol, 1 eq.) in acetone (10 mL), 2,2-dimethoxypropane (1.02 g, 9.83 mmol, 20 eq.) and TsOH-H2O (97.31 mg, 491.48 μmol, 1 eq.) were added. The mixture was stirred at room temperature for 1 hour. The solvent was removed by evaporation. The residue was diluted with EtOAc (50 mL), washed successively with aqueous NaHCO3 solution and brine, and then concentrated. The residue was purified by silica gel column chromatography (elution, PE / EA = 100:0-50:50) to give [(3aR,4R,6R,6aR)-4-[2-chloro-6-(4'-fluorospirocyclic [azacyclobutane-3,1'-indane]-1-yl)purin-9-yl]-2,2-dimethyl-3a,4,6,6a-tetrahydrofurano[3,4-d][1,3]dioxono-6-yl]methanol (170 mg, 338.69 μmol, yield 68.91%).

[0380] To a solution of [(3aR,4R,6R,6aR)-4-[2-chloro-6-(4'-fluorospiro[azacyclobutane-3,1'-indanedione]-1-yl)purin-9-yl]-2,2-dimethyl-3a,4,6,6a-tetrahydrofurano[3,4-d][1,3]dioxono-6-yl]methanol (170 mg, 338.69 μmol, 1 eq.) in PO(OEt)3 (2.5 mL) (74.65 mg, 298.84 μmol, 2.5 eq.) in PO(OEt)3 (2.5 mL), bis(dichlorophosphoryl)methane (74.65 mg, 298.84 μmol, 2.5 eq.) in PO(OEt)3 (2.5 mL) was added. The mixture was stirred at 0 °C for 4 hours, and then water (2 mL) was introduced into the reaction mixture. The mixture was stirred overnight at room temperature. The reaction mixture was purified by reversed-phase C-18 silica gel column chromatography (gradient elution, 0 to 25% ACN in water) to give compound d-2 (40.2 mg, yield 19.15%). 1 H NMR(500MHz,MeOD)δppm 2.52(dt,J=42.1,14.0Hz,4H),3.00(t,J=7.0Hz,2H),4.21–4.50(m,4H),4.65(s,1H),6.01(d, J=5.0Hz,1H),6.96(t,J=8.4Hz,1H),7.26–7.38(m,2H),8.40(s,1H).m / z(ESI+):620.3(M+H).

[0381] Example 26. Synthesis of compound d-3

[0382]

[0383] To a solution of 5-fluoroindanone (6.0 g, 39.96 mmol, 1 eq.) cooled at 0 °C in EtOH (20 mL) and THF (20 mL), t-BuOK (8.97 g, 79.92 mmol, 2.0 eq.) in THF (40 mL) was added, followed by TOSMIC (11.70 g, 59.94 mmol, 1.5 eq.) in a 1:1 EtOH and THF (50 mL) solution. The mixture was stirred overnight at room temperature, cooled to 0 °C, and then brine was added. The mixture was extracted with EtOAc (2 x 120 mL). The organic layers were combined, washed with brine, dried over Na₂SO₄, filtered, and concentrated. The residue was purified and separated by silica gel column chromatography (elution: PE / EA = 100:0 -80:20) to give 5-fluoroindene-1-carbamate (2.15 g, 13.34 mmol, 33.38% yield).

[0384] Sodium hydroxide (1.86 g, 46.53 mmol, 3 eq.) was added to a mixture of 5-fluoroindene-1-carbamate (2.5 g, 15.51 mmol, 1 eq.) and water (30 mL). The mixture was stirred at 100 °C for 16 h, then cooled to room temperature, followed by the addition of water (30 mL). The mixture was extracted with EtOAc (2 x 30 mL). The aqueous layer was adjusted to pH 2 and extracted with EtOAc (2 x 30 mL). The combined organic layers were washed with brine, dried over Na₂SO₄, filtered, and evaporated to dryness to give 5-fluoroindene-1-carboxylic acid (2.7 g, 14.99 mmol, 96.61% yield) as a brown solid.

[0385] One drop of DMF was added to a solution of 5-fluoroindane-1-carboxylic acid (2.7 g, 14.99 mmol, 1 eq.) cooled at 0 °C in methanol (40 mL), followed by the addition of SOCl2 (7.13 g, 59.94 mmol, 4.35 mL, 4 eq.). The reaction mixture was stirred at room temperature for 3 hours, and the solvent was removed by evaporation at a bath temperature of approximately 40 °C. Water (60 mL) and EtOAc (40 mL) were added to the residue and mixed thoroughly. The organic layer was separated, washed successively with NaHCO3 (a q.) and NaCl (a q.), dried (sodium sulfate), filtered, and evaporated to dryness to give methyl 5-fluoroindane-1-carboxylic acid (2.8 g, 14.42 mmol, 96.21% yield) as a brown oil.

[0386] To a solution of methyl 5-fluoroindane-1-carboxylate (2.8 g, 14.42 mmol, 1 eq.) in DMSO (30 mL), K₂CO₃ (5.98 g, 43.25 mmol, 3 eq.) and HCHO (3.98 g, 43.25 mmol, 37% purity, 3 eq.) were added. The mixture was stirred at room temperature for 16 hours. The reaction was quenched with H₂O (100 mL). The mixture was extracted with ethyl acetate (50 mL). The pH of the aqueous solution was adjusted to 3 with 3N HCl. The mixture was extracted with EtOAc (2 x 80 mL). The pH of the aqueous layer was adjusted to 3 with 6N HCl, followed by extraction with EtOAc (2 x 60 mL). The organic layers were combined, washed with brine, dried with Na2SO4, filtered, and evaporated to dryness to give 5-fluoro-1-(hydroxymethyl)indane-1-carboxylic acid (3.0 g, 14.27 mmol, 98.99% yield).

[0387] Acetophenone (1.53 g, 14.27 mmol, 1 eq.), DIPEA (3.32 g, 25.69 mmol, 4.47 mL, 1.8 eq.), HOBT (2.89 g, 21.41 mmol, 1.5 eq.), and EDCI (4.10 g, 21.41 mmol, 1.5 eq.) were added to a solution of 5-fluoro-1-(hydroxymethyl)indane-1-carboxylic acid (3.0 g, 14.27 mmol, 1 eq.) in DMF (40 mL). The mixture was stirred at room temperature for 5 hours. The reaction was quenched with water (20 mL), and then EtOAc (100 mL) was added. The organic layer was separated, washed with brine, and the solvent was removed by evaporation, followed by the addition of water (50 mL) and EA (60 mL). The organic layer was washed with brine, dried with Na2SO4, filtered, and evaporated to dryness. The residue was purified by silica gel column chromatography (elution, PE / EA = 100:0 to 50:50) to give N-benzyl-5-fluoro-1-(hydroxymethyl)indane-1-carboxamide (3.6 g, 12.03 mmol, 84.27% yield).

[0388] Under a nitrogen atmosphere at 0 °C, PPh3 (3.79 g, 14.43 mmol, 1.2 eq.) was added to a mixture of N-benzyl-5-fluoro-1-(hydroxymethyl)indane-1-carboxamide (3.6 g, 12.03 mmol, 1 eq.) and tetrahydrofuran (40 mL), followed by the dropwise addition of DEAD (2.51 g, 14.43 mmol, 2.27 mL, 1.2 eq.). The mixture was stirred at room temperature for 16 hours. The reaction was quenched with water (20 mL), followed by the addition of EtOAc (100 mL). The organic layer was separated, washed with brine, dried over Na2SO4, filtered, and evaporated to dryness. The residue was purified by silica gel column chromatography (elution, PE / EA = 100:0 to 70:30) to give 1-benzyl-5'-fluoro-spirocyclic [azacyclobutane-3,1'-indane]-2-one as a white solid (3.0 g, 10.66 mmol, 88.67% yield).

[0389] LiAlH4 (202.35 mg, 5.33 mmol, 3 eq.) was added to a solution of aluminum trichloride (473.97 mg, 3.55 mmol, 2 eq.) cooled at 0 °C in 15 mL of tetrahydrofuran. The mixture was stirred at 0 °C for 20 min, and then 1-benzyl-5'-fluoro-spirocyclic [azacyclobutane-3,1'-indane]-2-one (500 mg, 1.78 mmol, 1 eq.; dissolved in 5 mL of THF) was added. The mixture was stirred at room temperature for 5 h, diluted with THF (10 mL), and then slowly added at 0 °C in a 15% aqueous sodium hydroxide solution until the pH of the mixture was adjusted to 9. The organic layer was separated and dried with MgSO4. The insoluble matter was removed by filtration, and the filtrate was concentrated to dryness to give 1-benzyl-5'-fluoro-spirocyclic [azacyclobutane-3,1'-indane] (475 mg, 1.78 mmol, 99.97% yield).

[0390] To a solution of 1-benzyl-5'-fluorospirocyclic [azacyclobutane-3,1'-indane] (475 mg, 1.42 mmol, 1 eq.) in methanol (10 mL), ammonium formate (134.45 mg, 2.13 mmol, 1.5 eq.) and Pd(OH)₂ (50 mg, 20% supported on carbon, ca. 50% wetted with water) were added. The mixture was stirred overnight at 60 °C under hydrogen atmosphere. The insoluble matter was filtered off and washed with methanol. The filtrate and washings were combined and concentrated to dryness to give the product 5'-fluorospirocyclic [azacyclobutane-3,1'-indane] (250 mg, 99.25%), which could be used directly in the next step.

[0391] Methyl [(2R,3R,4R,5R)-3,4-diacetoxy-5-(2,6-dichloropurin-9-yl)tetrahydrofuran-2-yl]acetate (217.03 mg, 485.28 μmol, 1 eq.) and DIPEA (156.79 mg, 1.21 mmol, 211.31 μL, 2.5 eq.) were added to a solution of 5'-fluorospirocyclic [azacyclobutane-3,1'-indane] (86 mg, 97.056 μmol, 1 eq.) in 1,4-dioxane (10 mL). The mixture was stirred at 100 °C for 4 hours. The solvent was removed by evaporation. The residue was diluted with EtOAc (50 mL) and washed sequentially with water and brine. The organic layer was concentrated, and the residue was purified by silica gel column chromatography (elution, PE / EA = 100:0-50:50) to give methyl [(2R,3R,4R,5R)-3,4-diacetoxy-5-[2-chloro-6-(5'-fluorospirocyclic [azacyclobutane-3,1'-indane]-1-yl)purin-9-yl]tetrahydrofuran-2-yl]acetate as a white solid (190 mg, 323.14 μmol, 66.59% yield).

[0392] Add NH3-MeOH (7M, 1.38mL, 30eq.) to a mixture of [(2R,3R,4R,5R)-3,4-diacetoxy-5-[2-chloro-6-(5'-fluorospirocyclic [azacyclobutane-3,1'-indane]-1-yl)purin-9-yl]tetrahydrofuran-2-yl]acetate (190 mg, 323.14 μmol, 1 eq.) in methanol (2.00 mL). The mixture was stirred overnight at room temperature, the solvent was removed by evaporation, the residue was diluted with EtOAc (30 mL), washed with brine (30 mL), dried with Na2SO4, and concentrated to dryness to give (2R,3R,4S,5R)-2-[2-chloro-6-(5'-fluorospirocyclic [azacyclobutane-3,1'-indane]-1-yl)purin-9-yl]-5-(hydroxymethyl)tetrahydrofuran-3,4-diol (149 mg, 322.60 μmol, 99.83% yield) as a white solid.

[0393] To a solution of (2R,3R,4S,5R)-2-[2-chloro-6-(5'-fluorospirocyclo[azacyclobutane-3,1'-indane]-1-yl)purin-9-yl]-5-(hydroxymethyl)tetrahydrofuran-3,4-diol (149 mg, 322.60 μmol, 1 eq.) in acetone (10 mL), TsOH-H2O (61.36 mg, 322.60 μmol, 1 eq.) and 2,2-dimethoxypropane (671.96 mg, 6.45 mmol, 793.34 μL, 20 eq.) were added. The mixture was stirred at room temperature for 2 hours. The mixture was alkalized to pH 9 at 0 °C by slow addition of an aqueous solution of NaHCO3 (aq.). The solvent was removed by evaporation. The residue was extracted with EtOAc (2 x 30 mL). The organic layer was washed with brine and concentrated. The residue was purified by silica gel column chromatography (elution, PE / EA = 100:0-50:50) to give a white solid [(3aR,4R,6R,6aR)-4-[2-chloro-6-(5'-fluorospirocyclic [azacyclobutane-3,1'-indane]-1-yl)purin-9-yl]-2,2-dimethyl-3a,4,6,6a-tetrahydrofurano[3,4-d][1,3]dioxono-6-yl]methanol (138 mg, 274.94 μmol, 85.22% yield).

[0394] To a solution of [(3aR,4R,6R,6aR)-4-[2-chloro-6-(5'-fluorospirocyclo[azacyclobutane-3,1'-indanedione]-1-yl)purin-9-yl]-2,2-dimethyl-3a,4,6,6a-tetrahydrofurano[3,4-d][1,3]dioxono-6-yl]methanol (138 mg, 274.94 μmol, 1 eq.) in PO(OEt)3 (1.5 mL) ...2.5 eq.) (0 °C) (1.5 mL) (2.5 eq.) (0 °C) (1.5 mL) (1.5 mL) (2.5 eq.) (0 °C) (1.5 mL) (1.5 mL) (2.5 eq.) (0 °C) (1.5 mL) (1.5 mL) (2.5 mL) (1.5 mL) (0 °C) (1.5 mL) (2.5 mL) (1.5 mL) (1.5 mL) (2.5 mL) (1.5 mL)) (0 °C) (1.5 mL) (1.5 mL) (2.5 mL) (1.5 mL) (1.5 mL) (1.5 mL)) (0 °C) (1.5 mL) (1.5 mL) (1.5 mL) (1.5 mL) (1.5 mL) (1.5 mL) (1.5 mL) (1.5 mL) (1.5 mL)) (1.5 mL) (1.5 mL) (1.5 mL) (1.5 mL) (1.5 mL)) (1.5 mL) (1.5 mL) (1 The reaction mixture was purified by reversed-phase C-18 silica gel column chromatography (gradient elution, 0 to 30% ACN in water) to give compound d-3 (40 mg, 64.53 μmol, 23.55% yield). 1 H NMR(500MHz,MeOD)δppm 2.47–2.60(m,4H),3.00(t,J=6.8Hz,2H),4.25–4.43(m,4H),4.43–4.52(m,2H),4.68(m,3H),6 .04(d,J=4.8Hz,1H),7.00(d,J=9.0Hz,2H),7.53(s,1H),8.45(s,1H); m / z(ESI+):620.0(M+H).

[0395] Example 27. Synthesis of compound d-4

[0396]

[0397] To a solution of 6-fluoroindene-1-one (2 g, 13.32 mmol, 1 eq.) cooled at 0 °C in EtOH (9 mL) and THF (9 mL), t-BuOK (2.99 g, 26.64 mmol, 2.0 eq.) in THF (25 mL) was added, followed by a mixture of TOSMIC (3.90 g, 19.98 mmol, 1.5 eq.) in EtOH and THF (1:1, 25 mL). The mixture was stirred at room temperature for 16 hours, cooled to 0 °C, and then brine was added. The mixture was extracted with EtOAc and the extract was concentrated. The residue was purified by silica gel column chromatography (elution: PE / EA = 100:0 -85:15) to give 6-fluoroindene-1-carbonitrile (1.3 g, 60.6%).

[0398] Sodium hydroxide (1.94 g, 48.39 mmol, 3.0 eq.) was added to a mixture of 6-fluoroindene-1-carbamate (2.6 g, 16.13 mmol, 1 eq.) and water (30 mL). The mixture was stirred at 100 °C for 5 hours and then cooled to room temperature. The mixture was diluted with water (10 mL) and extracted with EtOAc (2 x 35 mL). The aqueous layer was adjusted to pH 2 and extracted with EtOAc (2 x 20 mL). The extracts were combined, washed with brine, dried over Na2SO4, and concentrated to give 6-fluoroindene-1-carboxylic acid (2.7 g, 92.9%) as a yellow solid.

[0399] One drop of DMF was added to a solution of 6-fluoroindane-1-carboxylic acid (2.8 g, 15.54 mmol, 1 eq.) cooled at 0 °C in methanol (40 mL), followed by the addition of SOCl2 (7.40 g, 62.16 mmol, 4.51 mL, 4.0 eq.). The reaction mixture was stirred overnight at room temperature. The solvent was removed by evaporation, and the residue was treated with water (50 mL) and EtOAc (100 mL). The organic layer was separated, washed first with an aqueous solution of NaHCO3 and then with brine, dried (sodium sulfate), and evaporated to give methyl 6-fluoroindane-1-carboxylic acid (3 g, 99.4%) as a brown oil.

[0400] To a solution of methyl 6-fluoroindane-1-carboxylic acid (3 g, 15.45 mmol, 1 eq.) in DMSO (30 mL), K₂CO₃ (7.05 g, 50.98 mmol, 3.3 eq.) and HCHO (4.27 g, 46.34 mmol, 37% purity, 3.0 eq.) were added. The mixture was stirred at room temperature for 16 hours. The reaction solution was quenched with H₂O (100 mL) and extracted with EtOAc (2 x 80 mL). The pH of the aqueous solution was adjusted to 3 with 6N HCl and extracted with EtOAc (2 x 60 mL). The organic layers were combined, washed with brine, dried over Na₂SO₄, filtered, and evaporated to give 6-fluoro-1-(hydroxymethyl)indane-1-carboxylic acid (2.37 g, 72.9% yield) as a pale yellow solid.

[0401] To a solution of 6-fluoro-1-(hydroxymethyl)indane-1-carboxylic acid (2.37 g, 11.27 mmol, 1 eq.) in DMF (27 mL), benzylamine (1.21 g, 11.27 mmol, 1.23 mL, 1.0 eq.), DIPEA (2.62 g, 20.29 mmol, 3.53 mL, 1.8 eq.), EDCI (3.24 g, 16.91 mmol, 1.5 eq.), and HOBT (2.29 g, 16.91 mmol, 1.5 eq.) were added. The mixture was stirred at room temperature for 5 hours. The mixture was then diluted with EtOAc (100 mL) and water (100 mL). The organic layer was separated, washed with brine (5 x 100 mL), dried with Na2SO4, and concentrated to give N-benzyl-6-fluoro-1-(hydroxymethyl)indane-1-carboxamide (3.2 g, 94.8%).

[0402] At 0 °C, PPh3 (1.05 g, 4.01 mmol, 1.2 eq.) was added to a solution of N-benzyl-6-fluoro-1-(hydroxymethyl)indane-1-carboxamide (1 g, 3.34 mmol, 1 eq.) and tetrahydrofuran (14 mL), followed by DEAD (698.14 mg, 4.01 mmol, 631.23 μL, 1.2 eq.). The mixture was stirred overnight at room temperature and diluted with EtOAc (40 mL). The organic layer was separated, washed with brine, and concentrated. The residue was purified by silica gel column chromatography (elution, PE / EA = 100:0 to 60:40) to give 1-benzyl-6'-fluoro-spirocyclic [azacyclobutane-3,1'-indane]-2-one (680 mg, 72.3%) as a yellow solid.

[0403] LiAlH4 (275.19 mg, 7.25 mmol, 3.0 eq.) was added to a solution of aluminum trichloride (644.60 mg, 4.83 mmol, 2.0 eq.) cooled at 0 °C in 15 mL of tetrahydrofuran. The mixture was stirred at 0 °C for 15 min, and then 1-benzyl-6'-fluoro-spirocyclic [azacyclobutane-3,1'-indane]-2-one (80 mg, 2.42 mmol, 1 eq.; in THF, 8 mL) was added. The mixture was stirred at room temperature overnight, the reaction was quenched with H2O (0.6 mL), and then 15% NaOH aqueous solution (2.5 mL) and EtOAc (20 mL) were added. The solids were removed by filtration, and the filtrate was concentrated to give 1-benzyl-6'-fluoro-spirocyclic [azacyclobutane-3,1'-indane] (600 mg, 92.8%).

[0404] Pd(OH)₂ (100 mg) and ammonium formate (212.29 mg, 3.37 mmol, 1.5 eq.) were added to a methanol (10 mL) solution of 1-benzyl-6'-fluorospirocyclic [azacyclobutane-3,1'-indane] (600 mg, 92.8%). The mixture was stirred at 60 °C for 5 hours under hydrogen atmosphere. The insoluble matter was filtered off and washed with methanol. The filtrate and washings were combined and concentrated to dryness to give 6'-fluorospirocyclic [azacyclobutane-3,1'-indane] (390 mg, 98.1%).

[0405] Methyl acetate [(2R,3R,4R,5R)-3,4-diacetoxy-5-(2,6-dichloropurin-9-yl)tetrahydrofuran-2-yl]acetate (500 mg, 1.12 mmol, 1 eq.) in 1,4-dioxane (20 mL) was mixed with 6'-fluorospirocyclic [azacyclobutane-3,1'-indane] (237.76 mg, 1.34 mmol, 1.2 eq.) and DIPEA (577.96 mg, 4.47 mmol, 778.93 μL, 4.0 eq.). The mixture was stirred at 100 °C for 3 hours. The mixture was concentrated, and the residue was purified by silica gel column chromatography (elution, PE / EtOAc = 100:0 to 60:40) to give methyl [(2R,3R,4R,5R)-3,4-diacetoxy-5-[2-chloro-6-(6'-fluorospirocyclic [azacyclobutane-3,1'-indane]-1-yl)purin-9-yl]tetrahydrofuran-2-yl]acetate (500 mg, 76.1%) as a white solid.

[0406] Add NH3-MeOH (7M, 3.64mL, 30eq.) to a methanol (2mL) solution of methyl [(2R,3R,4R,5R)-3,4-diacetoxy-5-[2-chloro-6-(6'-fluorospiro[azacyclobutane-3,1'-indane]-1-yl)purin-9-yl]tetrahydrofuran-2-yl]acetate (500mg, 850.36μmol, 1eq.). The mixture was stirred overnight at room temperature, the solvent was removed by evaporation, the residue was diluted with EtOAc, washed (first with water, then with brine), dried with Na2SO4, and concentrated to give (2R,3R,4S,5R)-2-[2-chloro-6-(6'-fluorospirocyclic [azacyclobutane-3,1'-indane]-1-yl)purin-9-yl]-5-(hydroxymethyl)tetrahydrofuran-3,4-diol (380 mg, 96.7%).

[0407] To a solution of (2R,3R,4S,5R)-2-[2-chloro-6-(6'-fluorospirocyclo[azacyclobutane-3,1'-indane]-1-yl)purin-9-yl]-5-(hydroxymethyl)tetrahydrofuran-3,4-diol (380 mg, 822.74 μmol, 1 eq.) in acetone (15 mL), p-TsOH (141.68 mg, 822.74 μmol, 1 eq.) and 2,2-dimethoxypropane (1.29 g, 12.34 mmol, 15 eq.) were added. The mixture was stirred at room temperature for 3 hours. The mixture was diluted with EtOAc (25 mL), washed with an aqueous solution of NaHCO3 (aq.), then washed with brine, and concentrated. The residue was purified by silica gel column chromatography (elution, PE / EtOAc = 100:0-60:40) to give a white solid [(3aR,4R,6R,6aR)-4-[2-chloro-6-(6'-fluorospirocyclic [azacyclobutane-3,1'-indane]-1-yl)purin-9-yl]-2,2-dimethyl-3a,4,6,6a-tetrahydrofurano[3,4-d][1,3]dioxono-6-yl]methanol (200 mg, 48.4%).

[0408] To a solution of [(3aR,4R,6R,6aR)-4-[2-chloro-6-(6'-fluorospiro[azacyclobutane-3,1'-indanedione]-1-yl)purin-9-yl]-2,2-dimethyl-3a,4,6,6a-tetrahydrofurano[3,4-d][1,3]dioxono-6-yl]methanol (200 mg, 398.46 μmol, 1 eq.) in PO(OEt)3 (2 mL) was added bis(dichlorophosphoryl)methane (248.82 mg, 996.14 μmol, 2.5 eq.) in PO(OEt)3 (2 mL). The mixture was stirred at 0 °C for 5 hours. Then, water (2 mL) was added to the mixture at 0 °C. The mixture was stirred overnight at 25 °C. The mixture was directly injected into a reversed-phase C-18 silica column for purification (gradient elution, 0 to 30% ACN in water) to give compound d-4 (75 mg, 30.3% yield). 1 H NMR(500MHz,CD3OD)δppm 2.41-2.57(m,4H),2.93(t,J=7.0Hz,2H),4.21-4.39(m,3H),4.42(t,J=4.7Hz,1H),4.64(t,J=4.9Hz,1H), 6.00(d,J=4.9Hz,1H),6.95(t,J=8.6Hz,1H),7.23(t,J=9.1Hz,2H),8.39(s,1H); m / z(ESI+):620.1(M+H).

[0409] Example 28. Synthesis of compound d-5

[0410]

[0411] 4 g (26.64 mmol, 1 eq.) of 7-fluoroindene-1-one cooled at 0 °C was added to a solution of EtOH (15 mL) and THF (15 mL), followed by t-BuOK (5.98 g, 53.28 mmol, 2.0 eq.) in THF (40 mL). Then, TOSMIC (7.80 g, 39.96 mmol, 1.5 eq.) in EtOH and THF (1:1, 50 mL). The mixture was stirred at room temperature for 16 h and cooled to 0 °C. After the addition of brine, the mixture was extracted with EtOAc. The organic layer was concentrated, and the residue was purified by silica gel column chromatography (elution: PE / EtOAc = 100:0 -85:15) to give a brown oily substance of 7-fluoroindene-1-carbonitrile (2.5 g, 58.2%).

[0412] Sodium hydroxide (1.86 g, 46.53 mmol, 3.0 eq.) was added to a mixture of 7-fluoroindene-1-carbamate (2.5 g, 15.51 mmol, 1 eq.) and water (25 mL). The mixture was stirred at 110 °C for 16 h and then cooled to room temperature. The mixture was diluted with water (10 mL) and extracted with EtOAc (2 x 35 mL). The pH of the aqueous layer was adjusted and extracted with EtOAc (2 x 20 mL). The extracts were combined, washed with brine, dried over Na2SO4, and concentrated to give 7-fluoroindene-1-carboxylic acid (2.76 g, 93.0%).

[0413] One drop of DMF was added to a solution of 6-fluoroindane-1-carboxylic acid (2.7 g, 14.99 mmol, 1 eq.) cooled at 0 °C in methanol (25 mL), followed by the addition of SOCl2 (7.13 g, 59.94 mmol, 4.35 mL, 4.0 eq.). The mixture was stirred at 25 °C for 4 hours. The solvent was removed by evaporation. The residue was diluted with water (50 mL) and extracted with EtOAc (100 mL). The organic layer was washed (first with an aqueous solution of NaHCO3, and subsequently with brine), dried (sodium sulfate), filtered, and evaporated to dryness to give methyl 7-fluoroindane-1-carboxylic acid (2.7 g, 92.8%) as a brown oil.

[0414] To a solution of methyl 7-fluoroindane-1-carboxylic acid (2.7 g, 13.90 mmol, 1 eq.) in DMSO (27 mL), K₂CO₃ (6.34 g, 45.88 mmol, 3.3 eq.) and HCHO (3.84 g, 41.71 mmol, 37% purity, 3.0 eq.) were added. The mixture was stirred overnight at room temperature. The reaction was quenched with H₂O (100 mL), and the mixture was extracted with EtOAc (2 x 80 mL). The pH of the aqueous solution was adjusted to 3 with 6N HCl, and the layer was extracted with EtOAc (2 x 60 mL). The organic layers were combined, washed with brine, dried over Na₂SO₄, filtered, and evaporated to give 7-fluoro-1-(hydroxymethyl)indane-1-carboxylic acid (2.2 g, 75.3%).

[0415] Aniline (1.12 g, 10.47 mmol, 1 eq.), DIPEA (2.43 g, 18.84 mmol, 3.28 mL, 1.8 eq.), HOBT (2.12 g, 15.70 mmol, 1.5 eq.), and EDCI (3.01 g, 15.70 mmol, 1.5 eq.) were added to a solution of 7-fluoro-1-(hydroxymethyl)indane-1-carboxylic acid (2.2 g, 10.47 mmol, 1 eq.) in DMF (25 mL). The mixture was stirred at room temperature for 16 hours. It was then diluted with EtOAc (100 mL) and water (100 mL). The organic layer was separated, washed with brine (5 x 100 mL), dried with Na2SO4, and concentrated to give N-benzyl-7-fluoro-1-(hydroxymethyl)indane-1-carboxamide (2.97 g, 94.8%) as a yellow solid.

[0416] At approximately 0 °C, PPh3 (1.14 g, 4.34 mmol, 1 eq.) and DEAD (756.32 mg, 4.34 mmol, 1.3 eq.) were added to a solution of N-benzyl-7-fluoro-1-(hydroxymethyl)indane-1-carboxamide (1 g, 3.34 mmol, 1 eq.) and tetrahydrofuran (15 mL). The mixture was stirred at room temperature for 3 hours and diluted with EtOAc (40 mL). The organic layer was washed with brine and concentrated. The residue was purified by silica gel column chromatography (elution, PE / EA = 100:0 to 60:40) to give 1-benzyl-7'-fluoro-spirocyclic [azacyclobutane-3,1'-indane]-2-one (880 mg, 93.6%) as a pale yellow solid.

[0417] LiAlH4 (364.22 mg, 9.60 mmol, 3.0 eq.) was added to a solution of aluminum trichloride (853.15 mg, 6.40 mmol, 2.0 eq.) cooled at 0 °C in 15 mL of tetrahydrofuran. The mixture was stirred at 0 °C for 15 min, and then 1-benzyl-7'-fluoro-spirocyclic [azacyclobutane-3,1'-indane]-2-one (900 mg, 3.20 mmol, 1 eq.) was added in 10 mL of THF. The mixture was stirred overnight at room temperature, the reaction was quenched with H2O (0.6 mL), and then 15% NaOH aqueous solution (3.5 mL) and EtOAc (20 mL) were added. The solids were removed by filtration, and the filtrate was concentrated to dryness to give 1-benzyl-7'-fluoro-spirocyclic [azacyclobutane-3,1'-indane] (730 mg, 85.4%).

[0418] To a methanol (15 mL) solution of 1-benzyl-7'-fluorospirocyclic [azacyclobutane-3,1'-indane] (730 mg, 2.73 mmol, 1 eq.), Pd(OH)₂ (130 mg, 2.73 mmol, 1 eq.) and ammonium formate (258.29 mg, 4.10 mmol, 1.5 eq.) were added. The mixture was stirred at 60 °C for 5 hours under hydrogen atmosphere. The insoluble matter was filtered off and washed with methanol. The filtrate and washings were combined and concentrated to dryness to give 7'-fluorospirocyclic [azacyclobutane-3,1'-indane] (480 mg, 99.2%).

[0419] Methyl [(2R,3R,4R,5R)-3,4-diacetoxy-5-(2,6-dichloropurin-9-yl)tetrahydrofuran-2-yl]acetate (500 mg, 1.12 mmol, 1 eq.) in 20 mL of 1,4-dioxane was mixed with 7'-fluorospirocyclic [azacyclobutane-3,1'-indane] (237.76 mg, 1.34 mmol, 1.2 eq.) and DIPEA (577.96 mg, 4.47 mmol, 778.93 μL, 4.0 eq.). The mixture was stirred at 100 °C for 3 hours. The mixture was concentrated, and the residue was purified by silica gel column chromatography (elution, PE / EtOAc = 100:0 to 60:40) to give methyl [(2R,3R,4R,5R)-3,4-diacetoxy-5-[2-chloro-6-(7'-fluorospirocyclic [azacyclobutane-3,1'-indane]-1-yl)purin-9-yl]tetrahydrofuran-2-yl]acetate (400 mg, 45.6%) as a white solid.

[0420] Methyl [(2R,3R,4R,5R)-3,4-diacetoxy-5-[2-chloro-6-(7'-fluorospiro[azacyclobutane-3,1'-indane]-1-yl)purin-9-yl]tetrahydrofuran-2-yl]acetate (300 mg, 510.22 μmol, 1 eq.) in methanol (2 mL) was added to NH3-MeOH (7 M, 2.19 mL, 30 eq.). The mixture was stirred overnight at room temperature. The solvent was removed by evaporation, the residue was diluted with EtOAc, washed successively with water and brine, dried with Na2SO4, and concentrated to dryness to give (2R,3R,4S,5R)-2-[2-chloro-6-(7'-fluorospirocyclic [azacyclobutane-3,1'-indane]-1-yl)purin-9-yl]-5-(hydroxymethyl)tetrahydrofuran-3,4-diol (200 mg, 84.9%).

[0421] To a solution of (2R,3R,4S,5R)-2-[2-chloro-6-(7'-fluorospirocyclo[azacyclobutane-3,1'-indane]-1-yl)purin-9-yl]-5-(hydroxymethyl)tetrahydrofuran-3,4-diol (200 mg, 433.02 μmol, 1 eq.) in acetone (10 mL), p-TsOH (74.57 mg, 433.02 μmol, 1 eq.) and 2,2-dimethoxypropane (676.47 mg, 6.50 mmol, 15 eq.) were added. The mixture was stirred at room temperature for 3 hours. The mixture was diluted with EtOAc (25 mL), washed successively with NaHCO3 (aq.) aqueous solution and brine, and then concentrated. The residue was purified by silica gel column chromatography (elution, PE / EtOAc = 100:0-60:40) to give a white solid [(3aR,4R,6R,6aR)-4-[2-chloro-6-(7'-fluorospirocyclic [azacyclobutane-3,1'-indane]-1-yl)purin-9-yl]-2,2-dimethyl-3a,4,6,6a-tetrahydrofurano[3,4-d][1,3]dioxono-6-yl]methanol (140 mg, 64.4%).

[0422] To a solution of [(3aR,4R,6R,6aR)-4-[2-chloro-6-(7'-fluorospiro[azacyclobutane-3,1'-indanedione]-1-yl)purin-9-yl]-2,2-dimethyl-3a,4,6,6a-tetrahydrofurano[3,4-d][1,3]dioxono-6-yl]methanol (140 mg, 278.92 μmol, 1 eq.) in PO(OEt)3 (1.5 mL) (1.5 mL) (1.5 mL) (1.5 mL) (1.5 mL) (1.5 mL) (1.5 mL) (1.5 mL) (1.5 mL) (1.5 mL) (1.5 mL) (2.5 eq.) (1.5 mL) (0 °C) (2.5 mL) (1.5 mL) (0 °C ...2.5 mL) (0 °C) (2.5 mL) (2.5 mL) (0 °C) (2.5 mL) (2.5 mL) (0 °C) (2.5 mL) (2.5 mL) (0 °C) (2.5 mL) (2.5 mL) (2.5 mL) (0 °C) (2.5 mL) (2.5 mL) (2.5 mL) (0 °C) (2.5 mL) (2.5 mL) (2.5 mL) (2.5 mL) (0 °C) (2.5 mL) (2.5 mL) ( The mixture was stirred overnight at 25°C and then directly injected into a reversed-phase C-18 silica column for purification (gradient elution, 0 to 25% ACN in water) to give compound d-5 (60 mg, 34.7% yield). 1 H NMR(500MHz,CD3OD)δppm 2.53(dt,J=41.8,14.0Hz,4H),3.02(t,J=7.2Hz,2H),4.22-4.38(m,3H),4.43(d,J=4.4Hz,2H),4.65(s,2H),4.76(s,1H),4.97(s,1H), 6.00(d,J=4.6Hz,1H),6.92(t,J=9.3Hz,1H),7.06(d,J=7.4Hz,1H),7.24(dd,J=12.9,7.7Hz,1H),8.39(s,1H); m / z(ESI+):620.1(M+H).

[0423] Example 29. Synthesis of compound d-6

[0424]

[0425] A mixture of tetrahydronaphthalene-1-carboxylic acid (0.5 g, 2.84 mmol, 1 eq.) and concentrated sulfuric acid (278.30 mg, 2.84 mmol, 151.25 μL, 1 eq.) in methanol (10 mL) was heated at 65 °C for 20 h. The reaction was monitored by TLC until complete. The resulting solution was evaporated. The residue was concentrated with EtOAc (30 mL), washed with brine, dried over Na2SO4, filtered, and evaporated to dryness to give methyl tetrahydronaphthalene-1-carboxylic acid (470 mg, 87.1%).

[0426] At -78 °C and under a nitrogen atmosphere, methyl tetrahydronaphthalene-1-carboxylate (470 mg, 2.47 mmol, 1 eq.) was added dropwise to a mixture of tetrahydrofuran (6 mL) with LDA (2 M, 1.48 mL, 1.2 eq.) and maintained at -78 °C, followed by the addition of bromoacetonitrile (592.69 mg, 4.94 mmol, 2 eq.) (at -78 °C). The mixture was stirred at room temperature for 4 hours. The completion of the reaction was indicated by TLC analysis. The resulting solution was quenched with 1 N HCl (3 mL) and diluted with EtOAc (30 mL). The organic layer was washed with brine, dried (Na2SO4), filtered, and evaporated to dryness. The residue was purified by silica gel column chromatography (elution, PE / EA = 100:0-70:30) to give methyl 1-(cyanomethyl)tetrahydronaphthalene-1-carboxylate (490 mg, 86.5%) as a yellow solid.

[0427] At 0 °C under a nitrogen atmosphere, cobalt dichloride (226.52 mg, 1.74 mmol, 2 eq.) and NaBH4 (330.00 mg, 8.72 mmol, 10 eq.) were added dropwise to a mixture of methyl 1-(cyanomethyl)tetrahydronaphthalene-1-carboxylic acid (200 mg, 872.32 μmol, 1 eq.) and ethanol (30 mL). The mixture was stirred overnight at room temperature. The reaction was indicated by TLC to indicate completion. The resulting solution was quenched with 1 N HCl (30 mL) and extracted with EtOAc (50 mL * 2). The organic layer was washed with brine, dried over Na2SO4, filtered, and evaporated to dryness. The residue was purified by silica gel column chromatography (elution: DCM / MeOH = 100:0-90:10) to give spirocyclic [pyrrolidine-4,1'-tetrahydronaphthalene]-2-one (50 mg, 28.5%) in solid form.

[0428] Lithium aluminum hydride (150.85 mg, 3.97 mmol, 2.0 eq.) was added to a mixture of spirocyclic [pyrrolidine-4,1'-tetrahydronaphthalene]-2-one (400 mg, 1.99 mmol, 1 eq.) and tetrahydrofuran (30 mL). The mixture was stirred overnight at 70 °C. TLC indicated the formation of a new product. The mixture was quenched with water (1 mL) and then ethyl acetate (20 mL) was added. The solids were removed by filtration, and the organic layer was concentrated to the residue. The residue was purified by silica gel column chromatography (elution, DCM / MeOH (100:0–80:20)) to give spirocyclic [pyrrolidine-3,1'-tetrahydronaphthalene] (230 mg, 61.8%) in solid form.

[0429] DIPEA (361.23 mg, 2.8 mmol, 486.83 μL, 2.5 eq.) was added to a mixture of [(2R,3R,4R,5R)-3,4-diacetoxy-5-(2,6-dichloropurin-9-yl)tetrahydrofuran-2-yl]acetate (500 mg, 1.12 mmol, 1 eq.) and spirocyclic [pyrrolidine-3,1'-tetrahydronaphthalene] (230.32 mg, 1.23 mmol, 1.1 eq.) in 1,4-dioxane (20 mL). The mixture was stirred overnight at 100 °C, diluted with ethyl acetate (40 mL), and washed successively with water and brine. The organic layer was concentrated, and the residue was purified by silica gel column chromatography (elution, PE / EtOAc = 100:0-60:40) to give methyl [(2R,3R,4R,5R)-3,4-diacetoxy-5-(2-chloro-6-spirocyclic [pyrrolidine-3,1'-tetrahydronaphthalene]-1-ylpurine-9-yl)tetrahydrofuran-2-yl]acetate (600 mg, 89.7%) as a yellow solid.

[0430] Methyl [(2R,3R,4R,5R)-3,4-diacetoxy-5-(2-chloro-6-spirocyclic [pyrrolidine-3,1'-tetrahydronaphthalene]-1-ylpurine-9-yl)tetrahydrofuran-2-yl]acetate (600 mg, 1.00 mmol, 1 eq.) in a methanol (8 mL) solution was added to NH3-MeOH (7 M, 4.30 mL, 30 eq.). The mixture was stirred overnight at room temperature. TLC showed the consumption of the starting material and the formation of a new product. The solvent was removed by evaporation; the residue was diluted with EtOAc (50 mL), washed successively with water (30 mL) and brine (30 mL), dried over Na2SO4, and concentrated to give (2R,3R,4S,5R)-2-(2-chloro-6-spirocyclic [pyrrolidine-3,1'-tetrahydronaphthalene]-1-ylpurine-9-yl)-5-(hydroxymethyl)tetrahydrofuran-3,4-diol (450 mg, 95.1%).

[0431] To a solution of (2R,3R,4S,5R)-2-(2-chloro-6-spirocyclic [pyrrolidine-3,1'-tetrahydronaphthalene]-1-ylpurine-9-yl)-5-(hydroxymethyl)tetrahydrofuran-3,4-diol (450 mg, 953.52 μmol, 1 eq.) in acetone (25 mL), 2,2-dimethoxypropane (1.49 g, 14.30 mmol, 15 eq.) and TsOH-H2O (188.80 mg, 953.52 μmol, 1 eq.) were added. The mixture was stirred at room temperature for 3 hours. The solvent was removed by evaporation; the residue was diluted with EtOAc (50 mL), washed with an aqueous solution of NaHCO3, and then washed with brine. The organic layer was concentrated, and the residue was purified by silica gel column chromatography (elution, PE / EtOAc = 100:0-50:50) to give a white solid [(3aR,4R,6R,6aR)-4-(2-chloro-6-spirocyclic[pyrrolidine-3,1'-tetrahydropyridine]-1-ylpurine-9-yl)-2,2-dimethyl-3a,4,6,6a-tetrahydrofurano[3,4-d][1,3]dioxono-6-yl]methanol (400 mg, yield 81.9%).

[0432] A solution of [(3aR,4R,6R,6aR)-4-(2-chloro-6-spirocyclic [pyrrolidine-3,1'-tetrahydropyridine]-1-ylpurine-9-yl)-2,2-dimethyl-3a,4,6,6a-tetrahydrofurano[3,4-d][1,3]dioxono-6-yl]methanol (400 mg, 781.25 μmol, 1 eq.) in PO(OEt)3 (6 mL) was cooled to ~0 °C, and then bis(dichlorophosphoryl)methane (390.29 mg, 1.56 mmol, 2.0 eq.) in PO(OEt)3 (3 mL) was added. The mixture was stirred at ~0 °C for 4 hours, then water (4 mL) was added, and the mixture was stirred at 40 °C for 1 hour, followed by stirring overnight at room temperature. The reaction mixture was purified by reversed-phase C-18 silica gel column chromatography (5–30% ACN in water) to give compound d-6 as a white solid (321 mg, 64.2%). 1 H NMR(500MHz,CD3OD)δppm 1.87(ddd,J=34.9,18.4,7.5Hz,4H),2.11(d,J=47.6Hz,1H),2.32-2.60(m,3H),2.82(d,J=5.5Hz,2H),3.86(dd,J=60.2,34 .8Hz,2H),4.11-4.46(m,6H),4.60(d,J=19.6Hz,1H),6.00(s,1H),7.03-7.19(m,3H),7.30(s,1H),8.37(d,J=25.3Hz,1H); 13C NMR(125MHz,CD3OD)δppm 21.34,26.47,27.52,28.57,31.19,35.75,39.69,41.72,44.72,46.89,48.04,62.29,63.22,65.91,7 1.32,75.76,84.65,89.81,118.81,127.37,130.35,138.74,140.07,141.21,152.10,154.03,155.58; 31 P NMR (203MHz, CD3OD) δppm 16.82, 19.71; m / z (ESI + ):630.18(M+H).

[0433] Example 30. Synthesis of compound d-7

[0434]

[0435] Under nitrogen protection and at -78°C, LiHMDS (1M, 33.05mL, 2.2eq.) was added dropwise to a mixture of indolin-2-one (2g, 15.02mmol, 1eq.) and tetrahydrofuran (30mL), followed by heating to -50°C for 30 minutes. The mixture was then cooled to -78°C, and a solution of 1,5-dibromopentane (3.45g, 15.02mmol, 2.05mL, 1eq.) in THF (15mL) was added. The mixture was stirred at room temperature for 3 hours, followed by reflux for 5 hours. The mixture was then stirred overnight at room temperature. The mixture was evaporated under reduced pressure, and the residue was partitioned between saturated ammonium chloride and ethyl acetate. The organic layer was concentrated, and the residue was purified by silica gel column chromatography (elution: PE / EtOAc = 100:0-60:40) to give a yellow solid spirocyclic [cyclohexane-1,3'-indoline]-2'-one (1.3 g, 43.0%).

[0436] Lithium aluminum hydride (188.56 mg, 4.97 mmol, 2.0 eq.) was added to a solution of spirocyclic [cyclohexane-1,3'-indoline]-2'-one (500 mg, 2.48 mmol, 1 eq.) in tetrahydrofuran (30 mL). The mixture was stirred overnight at 70 °C. The mixture was quenched with water (0.3 mL) and ethyl acetate (20 mL) was added. The solid was removed by filtration, the organic layer was concentrated, and the residue was purified by silica gel column chromatography (elution: PE / EtOAc = 100:0-80:20) to give spirocyclic [cyclohexane-1,3'-indoline] (401 mg, 86.2%) as a white solid.

[0437] Methyl [(2R,3R,4R,5R)-3,4-diacetoxy-5-(2,6-dichloropurin-9-yl)tetrahydrofuran-2-yl]acetate (450 mg, 1.01 mmol, 1 eq.) in 1,4-dioxane (25 mL) was mixed with spirocyclic [cyclohexane-1,3'-indoline] (207.29 mg, 1.11 mmol, 1.1 eq.) and DIPEA (325.10 mg, 2.52 mmol, 438.15 μL, 2.5 eq.). The mixture was stirred overnight at 100 °C. The mixture was concentrated, and the residue was purified by silica gel column chromatography (elution: PE / EtOAc = 100:0-60:40) to give methyl [(2R,3R,4R,5R)-3,4-diacetoxy-5-(2-chloro-6-spirocyclic[3a,7a-dihydro-2H-indol-3,1'-cyclohexane]-1-ylpurine-9-yl)tetrahydrofuran-2-yl]acetate (300 mg, yield 49.7%).

[0438] Methyl [(2R,3R,4R,5R)-3,4-diacetoxy-5-(2-chloro-6-spirocyclic [3a,7a-dihydro-2H-indol-3,1'-cyclohexane]-1-ylpurine-9-yl)tetrahydrofuran-2-yl]acetate (300 mg, 499.95 μmol, 1 eq.) was added to a methanol (5 mL) solution with MeOH-NH3 (7 M, 2.14 μL, 30 eq.). The mixture was stirred at room temperature for 5 hours. A large amount of solid formed. The solvent was removed by evaporation, and then ethyl acetate (100 mL) and water (70 mL) were added. After removing the aqueous layer, the formed solid was suspended in the organic layer. The product (2R,3R,4S,5R)-2-(2-chloro-6-spirocyclic[3a,7a-dihydro-2H-indole-3,1'-cyclohexane]-1-ylpurine-9-yl)-5-(hydroxymethyl)tetrahydrofuran-3,4-diol (236 mg, 99.6%) was obtained by concentrating the suspension layer.

[0439] Add 2,2-dimethoxypropane (1.04 g, 9.96 mmol, 20 eq.) and p-TsOH□H₂O (98.59 mg, 497.94 μmol, 1.0 eq.) to a solution of (2R,3R,4S,5R)-2-(2-chloro-6-spirocyclic [3a,7a-dihydro-2H-indol-3,1'-cyclohexane]-1-ylpurin-9-yl)-5-(hydroxymethyl)tetrahydrofuran-3,4-diol (236 mg, 497.94 μmol, 1 eq.) in acetone (20 mL). Stir the mixture overnight at room temperature. Remove the solvent by evaporation, and place the residue in ethyl acetate (50 mL). Wash the organic solution with an aqueous solution of NaHCO₃ and then with brine. The organic layer was concentrated, and the residue was purified by silica gel column chromatography (elution: PE / EtOAc = 100:0-50:50) to give [(3aR,4R,6R,6aR)-4-(2-chloro-6-spirocyclic[3a,7a-dihydro-2H-indol-3,1'-cyclohexane]-1-ylpurine-9-yl)-2,2-dimethyl-3a,4,6,6a-tetrahydrofurano[3,4-d][1,3]dioxono-6-yl]methanol (254 mg, yield 99.2%).

[0440] A solution of [(3aR,4R,6R,6aR)-4-(2-chloro-6-spirocyclo[3a,7a-dihydro-2H-indol-3,1'-cyclohexane]-1-ylpurine-9-yl)-2,2-dimethyl-3a,4,6,6a-tetrahydrofurano[3,4-d][1,3]dioxono-6-yl]methanol (298.82 mg, 583.64 μmol, 1 eq.) in PO(OEt)3 (4 mL) was cooled to ~0 °C, and then bis(dichlorophosphoryl)methane (291.57 mg, 1.17 mmol, 2.0 eq.) in PO(OEt)3 (3 mL) was added. The mixture was stirred at 0 °C for 5 hours. Then water (4 mL) was added to the reaction mixture. The mixture was stirred at 40 °C for 40 minutes, and then stirred overnight at room temperature. The reaction mixture was purified by reverse-phase C-18 silica gel column chromatography (0 to 25% ACN in water) to give compound d-7 (301 mg, 80.7%). 1H NMR(500MHz,CD3OD)δppm 1.31-1.81(m,10H),2.52(t,J=20.9Hz,2H),4.27(s,1H),4.30-4.42(m,2H),4.45(t,J=4.8Hz,1H),4.53-4.61(m ,2H),4.66(t,J=4.9Hz,1H),6.05(d,J=4.7Hz,1H),7.04(t,J=7.3Hz,1H),7.12-7.26(m,2H),8.41-8.49(m,2H); 13 CNMR(125MHz,CD3OD)δppm 24.12,26.53,27.51,38.39,45.84,62.14,66.16,71.48,75.72,84.56,89.72,11 8.98,120.30,123.56,128.51,140.84,142.88,143.18,152.54,153.35,154.56; 31 P NMR (203MHz, CD3OD) δppm 16.61, 19.95; m / z (ESI + ):630.27(M+H).

[0441] Example 31. Synthesis of compound d-8

[0442]

[0443] Methyl indane-1-carboxylate was prepared in the same manner as described in the d-1 section of compound.

[0444] Under nitrogen protection at -78°C, LDA (2M, 7.15mL, 1.2eq.) was added dropwise to a mixture of methyl indane-1-carboxylate (2.1g, 11.92mmol, 1eq.) and tetrahydrofuran (25mL). After 20 minutes, a solution of 2-bromoacetonitrile (2.86g, 23.84mmol, 2eq.) in THF (1mL) was added to the reaction mixture at -78°C. The resulting mixture was stirred at room temperature for 4 hours, quenched with 1N HCl (30mL), and then ethyl acetate (60mL) was added. The organic layer was washed with brine, dried over Na2SO4, filtered, and evaporated to dryness. The residue was purified by silica gel column chromatography (elution, PE / EtOAc = 100:0-70:30) to give methyl 1-(cyanomethyl)indane-1-carboxylate (2.4g, 93.5%) as a yellow oil.

[0445] Under nitrogen protection, cobalt dichloride (2.90 g, 22.30 mmol, 1 eq.) and sodium borohydride (4.22 g, 111.50 mmol, 10 eq.) were added dropwise to a mixture of 1-(cyanomethyl)indane-1-carboxylic acid methyl ester (2.4 g, 11.15 mmol, 1 eq.) and ethanol (25 mL) while maintaining the temperature at 0 °C. The reaction mixture was stirred overnight at room temperature, quenched with 1N HCl (150 mL), and extracted with EtOAc (2 x 200 mL). The organic layer was washed with brine, dried over Na2SO4, filtered, and evaporated to dryness. The residue was purified by silica gel column chromatography (elution, DCM / MeOH = 100:0-95:5) to give a yellow solid of spirocyclic [indane-1,3'-pyrrolidine]-2'-one (1.05 g, 50.3%).

[0446] Lithium aluminum hydride (405.37 mg, 10.68 mmol, 2.0 eq.) was added to a solution of spirocyclic [indane-1,3'-pyrrolidine]-2'-one (1 g, 5.34 mmol, 1 eq.) in tetrahydrofuran (40 mL). The mixture was stirred overnight at 70 °C, quenched with water (1 mL), and then added dropwise with 15% sodium hydroxide aqueous solution (1 mL), water (1 mL), and ethyl acetate (30 mL). The solid was removed by filtration, and the organic layer was concentrated to give spirocyclic [indane-1,3'-pyrrolidine] (800 mg, 86.5%) in solid form.

[0447] To a solution of [(2R,3R,4R,5R)-3,4-diacetoxy-5-(2,6-dichloropurin-9-yl)tetrahydrofuran-2-yl]methyl acetate (500 mg, 1.12 mmol, 1 eq.) and 1,4-dioxane (25 mL), spirocyclic [indane-1,3'-pyrrolidine] (232.44 mg, 1.34 mmol, 1.2 eq.) and DIPEA (361.23 mg, 2.8 mmol, 486.83 μL, 2.5 eq.) were added. The mixture was stirred at 100 °C for 2 hours and then concentrated to the residue. The residue was purified by silica gel column chromatography (elution, PE / EtOAc = 100:0-60:40) to give methyl [(2R,3R,4R,5R)-3,4-diacetoxy-5-(2-chloro-6-spirocyclic [1,3'-pyrrolidine]-1'-ylpurine-9-yl)tetrahydrofuran-2-yl]acetate (500 mg, 76.6%) as a white solid.

[0448] Methyl [(2R,3R,4R,5R)-3,4-diacetoxy-5-(2-chloro-6-spirocyclic [1,3'-pyrrolidine]-1'-ylpurine-9-yl)tetrahydrofuran-2-yl]acetate (500 mg, 856.14 μmol, 1 eq.) in methanol (6 mL) was added to MeOH-NH3 (7 M, 3.67 mL, 30 eq.). The mixture was stirred at room temperature for 4 hours. The solvent was removed by evaporation, and the residue was placed in ethyl acetate (100 mL), followed by the addition of water (70 mL). The organic layer was separated, washed with brine, dried with Na2SO4, and concentrated to obtain (2R,3R,4S,5R)-2-(2-chloro-6-spirocyclic [1,3'-pyrrolidine]-1'-ylpurine-9-yl)-5-(hydroxymethyl)tetrahydrofuran-3,4-diol (390 mg, 99.5%), a white solid.

[0449] Add 2,2-dimethoxypropane (1.33 g, 12.78 mmol, 1 eq.) and p-TsOH (146.66 mg, 851.70 μmol, 1.0 eq.) to a solution of (2R,3R,4S,5R)-2-(2-chloro-6-spirocyclic [1,3'-pyrrolidine]-1'-ylpurin-9-yl)-5-(hydroxymethyl)tetrahydrofuran-3,4-diol (390 mg, 851.70 μmol, 1 eq.) in acetone (20 mL). Stir the mixture overnight at room temperature. Remove the solvent by evaporation. Dilute the residue with EtOAc (50 mL), wash with an aqueous solution of NaHCO3 and then with brine. The organic layer was concentrated, and the residue was purified by silica gel column chromatography (elution, PE / EtOAc = 100:0-50:50) to give [(3aR,4R,6R,6aR)-4-(2-chloro-6-spirocyclic[1,3'-pyrrolidine]-1'-ylpurine-9-yl)-2,2-dimethyl-3a,4,6,6a-tetrahydrofurano[3,4-d][1,3]dioxono-6-yl]methanol (410 mg, 96.7%) as a white solid.

[0450] [(3aR,4R,6R,6aR)-4-(2-chloro-6-spirocyclic[1,3'-pyrrolidine]-1'-ylpurin-9-yl)-2,2-dimethyl-3a,4,6,6a-tetrahydrofurano[3,4-d][1,3]dioxono-6-yl]methanol (410 mg, 823.34 μmol, 1 eq.) cooled at 0 °C to a solution of PO(OEt)3 (4 mL) was mixed with bis(dichlorophosphoryl)methane (411.32 mg, 1.65 mmol, 2.0 eq) in PO(OEt)3 (4 mL). The mixture was stirred at 0 °C for 5 hours, and then water (6 mL) was added. The mixture was stirred at 40 °C for 1 hour, and then stirred overnight at room temperature. The reaction mixture was purified by reversed-phase C-18 silica gel column chromatography (gradient elution, 0 to 35% ACN in water) to give compound d-8 (301 mg, 58.4%). 1 H NMR (500MHz, CD3OD) δppm1.96-2.35(m,4H),2.44(dd,J=37.0,20.5Hz,2H),2.94(d,J=6.7Hz,2H),3.64-4.1 9(m,3H),4.21-4.45(m,5H),4.56(d,J=20.9Hz,1H),5.96(s,1H),7.12-7.29(m,4H),8.38(d,J=33.4Hz,1H); 13 C NMR(125MHz,CD3OD)δppm 26.44,27.50,28.55,31.11,37.22,38.81,39.17,53.92,56.03,59.67,60.67,65.91,71.32,75.73,84 .61,89.81,118.82,123.32,125.72,127.84,128.50,140.10,144.97,147.27,152.06,154.06,155.63; 31 P NMR (203MHz, CD3OD) δppm 16.96, 19.61; m / z (ESI + ):616.32(M+H).

[0451] Example 32. Synthesis of compound d-9

[0452]

[0453] A solution of 3-cyanoaniline-1-carboxylic acid tert-butyl ester (3.0 g, 16.46 mmol, 1 eq.) in THF (30 mL) was cooled to -78 °C, and then LiHMDS (1 M, 20.58 mL, 1.25 eq.) was added. The mixture was stirred for 20 min, and then a solution of 1-(bromomethyl)-2-iodobenzene (5.13 g, 17.29 mmol, 1.05 eq.) in THF (3 mL) was added. The mixture was stirred at -78 °C for 3 h, quenched with saturated ammonium chloride solution, and extracted with 50 mL of ethyl acetate. The organic layer was washed with brine and concentrated. The residue was purified by column chromatography (elution, PE / EA = 100:0-83:17) to give 3-cyano-3-(2-iodophenyl)azacyclobutane-1-carboxylic acid tert-butyl ester (6.44 g, 16.17 mmol, yield 98.22%) as a yellow oil.

[0454] 3-Cyano-3-(2-iodophenyl)azacyclobutane-1-carboxylic acid tert-butyl ester (6.44 g, 16.17 mmol, 1 eq.) was cooled to -78 °C in 60 mL of THF solution, and then n-BuLi (2.5 M, 12.94 mL, 2 eq.) was added dropwise at -78 °C. The mixture was stirred at -78 °C for 2 h. TLC showed that the starting material was completely consumed. The mixture was quenched with saturated ammonium chloride solution and extracted with ethyl acetate (75 mL * 2). The organic layer was washed with brine and concentrated. The residue was purified by column chromatography (elution, PE / EA = 100:0–83:17) to give 1'-oxospirocyclic [azacyclobutane-3,2'-indene]-1-carboxylic acid tert-butyl ester (3.4 g, 12.44 mmol, yield 76.92%) as a pale yellow solid.

[0455] 1'-oxospirocyclic [azacyclobutane-3,2'-indene]-1-carboxylic acid tert-butyl ester (1.0 g, 3.66 mmol, 1 eq.) was added to a solution of CH3OH / CH3COOH (5 mL + 10 mL) with Pd / C (100 mg, 10% purity). The mixture was stirred overnight at room temperature under hydrogen atmosphere. TLC showed that most of the starting material was consumed. The mixture was filtered and washed with MeOH. The filtrate was concentrated to remove most of the solvent. The mixture was then neutralized with saturated NaHCO3 and extracted with ethyl acetate (15 mL * 2). The organic layer was washed with brine and concentrated. The residue was purified by column chromatography (elution, PE / EA = 100:0–90:10) to give spirocyclic [azacyclobutane-3,2'-indene]-1-carboxylic acid tert-butyl ester (360 mg, 1.39 mmol, yield 37.94%) as a white solid.

[0456] Spirocyclic [azacyclobutane-3,2'-indene]-1-carboxylic acid tert-butyl ester (370 mg, 1.43 mmol, 1 eq.) was dissolved in HCl-EA (5 mL), and the mixture was stirred at room temperature for 3 hours. The mixture was concentrated and used in the next step.

[0457] Methyl acetate [(2R,3R,4R,5R)-3,4-diacetoxy-5-(2,6-dichloropurin-9-yl)tetrahydrofuran-2-yl] (523.74 mg, 1.17 mmol, 1 eq.) was added to a mixture of 1,4-dioxane (8 mL) and then [(2R,3R,4R,5R)-3,4-diacetoxy-5-(2,6-dichloropurin-9-yl)tetrahydrofuran-2-yl] was added. Methyl acetate (523.74 mg, 1.17 mmol, 1 eq.) and [(2R,3R,4R,5R)-3,4-diacetoxy-5-(2,6-dichloropurin-9-yl)tetrahydrofuran-2-yl]acetate (523.74 mg, 1.17 mmol, 1 eq.) and DIPEA (529.73 mg, 4.10 mmol, 713.92 μL, 3.5 eq.). The mixture was stirred overnight at 100 °C. TLC showed that the starting material was consumed. The solvent was removed by evaporation. The residue was placed in EtOAc (40 mL) and washed with water and then with brine. The organic phase was concentrated, and the residue was purified by silica gel column chromatography (elution, PE / EA = 100:0-65:35) to give methyl [(2R,3R,4R,5R)-3,4-diacetoxy-5-(2-chloro-6-spirocyclic [azacyclobutane-3,2'-indane]-1-yl-purin-9-yl)tetrahydrofuran-2-yl]acetate (620 mg, 1.09 mmol, yield 92.88%) as a yellow solid.

[0458] To a mixture of [(2R,3R,4R,5R)-3,4-diacetoxy-5-(2-chloro-6-spirocyclic [azacyclobutane-3,2'-indane]-1-ylpurin-9-yl)tetrahydrofuran-2-yl]methyl acetate (620 mg, 1.09 mmol, 1 eq.) in 6 mL of methanol, NH3-MeOH (7 M, 4.66 mL, 30 eq.) was added. The mixture was stirred at room temperature for 3 hours. LC-MS showed that the starting material disappeared. The solvent was removed by evaporation. The residue was placed in EtOAc (30 mL) and water (50 mL). The organic phases were combined, washed with brine, dried over Na2SO4, and concentrated to give (2R,3R,4S,5R)-2-(2-chloro-6-spirocyclic [azacyclobutane-3,2'-indane]-1-ylpurine-9-yl)-5-(hydroxymethyl)tetrahydrofuran-3,4-diol (459 mg, 1.03 mmol, yield 95.07%) as a white solid.

[0459] Add p-TsOH□H₂O (197.76 mg, 1.03 mmol, 1 eq.) and 2,2-dimethylpropane (2.15 g, 20.68 mmol, 20 eq.) to 30 mL of acetone solution of (2R,3R,4S,5R)-2-(2-chloro-6-spirocyclic [azacyclobutane-3,2'-indane]-1-ylpurin-9-yl)-5-(hydroxymethyl)tetrahydrofuran-3,4-diol (459 mg, 1.03 mmol, 1 eq.) and 2,2-dimethylpropane (2.15 g, 20.68 mmol, 20 eq.). Stir the mixture at room temperature for one week. Remove the solvent by evaporation. Place the residue in EtOAc (50 mL), wash with aqueous NaHCO₃ solution, and then wash with brine. The organic layer was concentrated, and the residue was purified by silica gel column chromatography (elution, PE / EA = 100:0–85:15) to give [(3aR,4R,6R,6aR)-4-(2-chloro-6-spirocyclic [azacyclobutane-3,2'-indane]-1-ylpurine-9-yl)-2,2-dimethyl-3a,4,6,6a-tetrahydrofurano[3,4-d][1,3]dioxono-6-yl]methanol (350 mg, 723.22 mmol, yield 69.94%) as a white solid.

[0460] A solution of [(3aR,4R,6R,6aR)-4-(2-chloro-6-spirocyclic [azacyclobutane-3,2'-indane]-1-ylpurin-9-yl)-2,2-dimethyl-3a,4,6,6a-tetrahydrofurano[3,4-d][1,3]dioxono-6-yl]methanol (348.55 mg, 720.22 μmol, 1 eq.) in PO(MeO)3 (4 mL) was cooled to 0 °C, and then bis(dichlorophosphoryl)methane (359.80 mg, 1.44 mmol, 2 eq.) in PO(MeO)3 (4 mL) was added. The mixture was stirred at 0 °C for 5 hours. After the starting material was consumed by LC-MS, 7 mL of water was added dropwise to the reaction system, and the mixture was stirred at 40 °C for 40 minutes, followed by stirring at room temperature overnight. The reaction mixture was purified by C-18 reversed-phase silica column chromatography (0–25% ACN in water) to give compound d-9 (260 mg, yield 58.63%). 1 H NMR(500MHz,MeOD)δppm 2.48(t,J=20.9Hz,2H),3.26(s,4H),4.24(s,2H),4.26-4.38(m,3H),4.42(t,J=4.7Hz,1H),4.55(s,1H), 4.63(t,J=4.9Hz,1H),6.00(d,J=4.8Hz,1H),7.15(dd,J=5.1,3.2Hz,2H),7.20-7.25(m,2H),8.39(s,1H); 31P NMR (203MHz, MeOD) δppm 16.85, 19.68; 13 C NMR(125MHz,MeOD)δppm 26.17,43.38,44.10,61.81,63.68,64.53,70.02,74.35,83.39,88.33,116.93,12 4.21,126.49,139.71,141.29,150.27,154.05,154.37; m / z(ESI+):602.31(M+H).

[0461] Example 33. Synthesis of compound d-10

[0462]

[0463] NBS (5.66 g, 31.78 mmol, 1.5 eq.) was added to a mixture of 4-fluoro-2-iodo-1-methylbenzene (5 g, 21.18 mmol, 1 eq.) and carbon tetrachloride (50 mL). The mixture was refluxed and stirred for 5 hours. The solvent was removed by evaporation, and the residue was purified by column chromatography (elution, PE / EA = 100:0–99:1) to give 1-(bromomethyl)-4-fluoro-2-iodobenzene (2.55 g, 8.10 mmol, yield 38.22%).

[0464] LiHMDS (1M, 9.19 mL, 1.25 eq.) was added dropwise to a solution of tert-butyl 3-cyanoazoalkyl-1-carboxylic acid (1.34 g, 7.35 mmol, 1 eq.) cooled at -78 °C in 20 mL of THF. The mixture was stirred at this temperature for 20 min, and then 1-(bromomethyl)-4-fluoro-2-iodobenzene (2.55 g, 8.09 mmol, 1.1 eq., dissolved in 5 mL of THF) was added. The mixture was stirred at -78 °C for 3 h. The reaction was quenched with a saturated ammonium chloride solution. The reaction mixture was extracted with ethyl acetate (2 x 25 mL). The organic phase was washed with brine and then concentrated. The residue was purified by silica gel column chromatography (elution, PE / EA = 100:0-75:25) to give tert-butyl 3-cyano-3-[(4-fluoro-2-iodophenyl)methyl]azacyclobutane-1-carboxylic acid (2.41 g, 5.79 mmol, yield 78.74%).

[0465] 3-Cyano-3-[(4-fluoro-2-iodophenyl)methyl]azacyclobutane-1-carboxylic acid tert-butyl ester (2.41 g, 5.79 mmol, 1 eq.) cooled at -78 °C was added dropwise to 25 mL of THF solution with n-BuLi (2.5 M, 4.63 mL, 2 eq.). The mixture was stirred at this temperature for 2 h. The reaction was quenched with saturated ammonium chloride solution, and the reaction mixture was extracted with ethyl acetate (2 x 100 mL). The organic phase was washed with brine and then concentrated. The residue was purified by silica gel column chromatography (elution, PE / EA = 100:0–70:30) to give 6'-fluoro-1'-oxospirocyclic [azacyclobutane-3,2'-indane]-1-carboxylic acid tert-butyl ester (1.21 g, 4.15 mmol, yield 71.74%).

[0466] At 0 °C, sodium borohydride (392.82 mg, 10.38 mmol, 2.5 eq.) was added fractionally to a solution of 6'-fluoro-1'-oxospirocyclic [azacyclobutane-3,2'-indane]-1-carboxylic acid tert-butyl ester (1.21 g, 4.15 mmol, 1 eq.) in methanol (5 mL). The mixture was stirred at this temperature for 2 hours. The solvent was removed by evaporation, and the residue was extracted with ethyl acetate (2 x 100 mL), washed with brine (100 mL), and the organic layer was concentrated to dryness to give 6'-fluoro-1'-hydroxyspirocyclic [azacyclobutane-3,2'-indane]-1-carboxylic acid tert-butyl ester (1.21 g, 4.13 mmol, yield 99.31%) as a white solid.

[0467] At -40 °C, boron trifluoride diethyl ether (4 mL) and triethylsilane (1.21 g, 10.40 mmol, 1 eq.) were added to a mixture of 6'-fluoro-1'-hydroxyspirocyclic [azacyclobutane-3,2'-indane]-1-carboxylic acid tert-butyl ester (305 mg, 1.04 mmol, 1 eq.) in DCM (10 mL). The mixture was stirred at 40 °C for 16 h. The reaction was then quenched with water (20 mL) and ammonium chloride solution (20 mL). The pH of the mixture was adjusted to 10 with 15% NaOH. The mixture was extracted with DCM (60 mL * 2). The combined organic layers were washed with brine (100 mL), dried with Na2SO4, filtered, and evaporated to give 5'-fluorospirocyclic [azacyclobutane-3,2'-indane] (100 mg, 394.99 μmol, yield 37.99%, purity 70%).

[0468] Methyl [(2R,3R,4R,5R)-3,4-diacetoxy-5-(2,6-dichloropurin-9-yl)tetrahydrofuran-2-yl]acetate (252 mg, 1 eq.) was added to 5 mL of dioxane solution along with 5'-fluorospirocyclic [azacyclobutane-3,2'-indane] (99.86 mg, 563.47 μmol, 1 eq.) and DIPEA (182.06 mg, 1.41 mmol, 245.36 μL, 2.5 eq.). The mixture was stirred at 100 °C for 3 hours. The concentrated mixture and the residue were purified by silica gel column chromatography (elution, PE / EA = 100:0-58:42) to give methyl [(2R,3R,4R,5R)-3,4-diacetoxy-5-[2-chloro-6-(5'-fluorospirocyclic [azacyclobutane-3,2'-indane]-1-yl]purin-9-yl]tetrahydrofuran-2-yl]acetate (200 mg, 136.06 μmol, yield 24.15%, purity 40%).

[0469] Methyl [(2R,3R,4R,5R)-3,4-diacetoxy-5-[2-chloro-6-(5'-fluorospirocyclo[azacyclobutane-3,2'-indane]-1-yl]purin-9-yl]tetrahydrofuran-2-yl]acetate (200 mg, 136.06 μmol, 1 eq.) in 5 mL of methanol was added to NH3-MeOH (7 M, 485.92 μL, 25 eq.). The mixture was incubated at room temperature. Stir for 2 hours. The residue was purified by silica gel column chromatography (elution, DCM / MeOH = 100:0-93:7) to give (2R,3R,4R,4S,4S,5R)-2-[2-chloro-6-(5'-fluorospirocyclic [azacyclobutane-3,2'-indane]-1-ylpurine-9-yl]-5-(hydroxymethyl)tetrahydrofuran-3,4-diol (60 mg, 129.91 μmol, yield 95.48%).

[0470] To a solution of (2R,3R,4R,4S,4S,5R)-2-[2-chloro-6-(5'-fluorospirocyclic [azacyclobutane-3,2'-indane]-1-ylpurin-9-yl]-5-(hydroxymethyl)tetrahydrofuran-3,4-diol (60 mg, 129.91 μmol, 1 eq.) in 10 mL of acetone, p-TsOH (22.37 mg, 129.91 μmol, 1 eq.) and 2,2-dimethylpropane (270.59 mg, 2.60, 2.60 mmol, 20 eq.) were added. The mixture was stirred at room temperature for 1 hour. The precipitate was removed by evaporation. Solvent. The residue was diluted with 50 mL of EA, washed successively with NaHCO3 aqueous solution and brine, and then concentrated. The residue was purified by silica gel column chromatography (elution, PE / EA = 100:0 to 55:45) to give [(3aR,4R,6R,6aR)-4-[2-chloro-6]-(5'-fluorospirocyclic [azacyclobutane-3,2'-indane]-1-ylpurine-9-yl]-2,2-dimethyl-3a,4,6,6a-tetrahydrofurano[3,4-d][1,3]dioxono-6-yl]methanol (60 mg, 119.54 μmol, yield 92.02%).

[0471] A solution of [(3aR,4R,6R,6aR)-4-[2-chloro-6]-(5'-fluorospirocyclic [azacyclobutane-3,2'-indane]-1-ylpurine-9-yl]-2,2-dimethyl-3a,4,6,6a-tetrahydrofurano[3,4-d][1,3]dioxono-6-yl]methanol (60 mg, 119.54 μmol, 1 eq.) cooled at 0 °C in PO(OEt)3 (2 mL) Bis(dichlorophosphoryl)methane (74.65 mg, 298.84 μmol, 2.5 eq) was added to PO(OEt)3 (2 mL). The mixture was stirred at 0 °C for 4 hours, and then water (3 mL) was added. The mixture was stirred at room temperature overnight. The reaction mixture was purified by reverse-phase C-18 silica gel column chromatography (gradient elution, 0 to 35% ACN in water) to give compound d-10 (7.1 mg, yield 9.58%). 1 H NMR(500MHz,MeOD)δppm 2.47(t,J=20.8Hz,2H),3.25(d,J=19.8Hz,4H),4.19–4.37(m,5H),4.42(d,J=8.2Hz,1H),4.49(d,J=58.8Hz,2H),4.64(s,1H) ,6.00(d,J=4.8Hz,1H),6.88(t,J=8.7Hz,1H),6.97(d,J=8.2Hz,1H),7.14–7.27(m,1H),8.40(s,1H); m / z(ESI+):620.3(M+H).

[0472] Example 34. Synthesis of compound d-11

[0473]

[0474] NBS (5.66 g, 31.78 mmol, 1.5 eq.) and benzoyl peroxide (256.57 mg, 1.06 mmol, 0.05 eq.) were added to a mixture of 1-fluoro-3-iodo-2-toluene (5.0 g, 21.18 mmol, 1 eq.) and carbon tetrachloride (80 mL). The mixture was refluxed and stirred for 5 hours. The solvent was removed by evaporation, and the residue was extracted with dichloromethane (50 mL x 2). The organic layer was washed with brine, dried over Na2SO4, filtered, and then concentrated. The residue was purified by silica gel column chromatography (elution, PE / EA = 100:0–99:1) to give the product 2-(bromomethyl)-1-fluoro-3-iodobenzene (1.97 g, 6.26 mmol, yield 29.53%).

[0475] LiHMDS (1M, 7.56 mL, 1.25 eq.) was added dropwise to a solution of tert-butyl 3-cyanoaniline-1-carboxylic acid (1.1 g, 6.05 mmol, 1 eq.) cooled at -78 °C in 10 mL of tetrahydrofuran. The mixture was stirred at -78 °C for 20 min, and then a solution of 2-(bromomethyl)-1-fluoro-3-iodobenzene (2.0 g, 6.35 mmol, 1.05 eq.) in 10 mL of THF was added. The mixture was stirred at -78 °C for 3 h, and the reaction was quenched with saturated ammonium chloride. Extraction of the mixture was completed using ethyl acetate (50 mL * 2). The combined organic layers were washed with brine, dried over Na₂SO₄, filtered, and concentrated. The residue was purified by silica gel column chromatography (elution, PE / EA = 100:0-80:20) to give tert-butyl 3-cyano-3-(2-fluoro-6-iodophenyl)azacyclobutane-1-carboxylic acid (1.92 g, 4.61 mmol, yield 76.27%) as a yellow oil.

[0476] To a solution of 3-cyano-3-(2-fluoro-6-iodophenyl)azacyclobutane-1-carboxylic acid tert-butyl ester (1.92 g, 4.61 mmol, 1 eq.) cooled at -78 °C, a mixture of n-BuLi (2.5 M, 3.69 mL, 2 eq.) was added dropwise to 30 mL of THF solution and stirred at -78 °C for 3 h. The reaction was quenched with saturated ammonium chloride solution, and the reaction mixture was extracted with ethyl acetate (2 x 30 mL). The organic phase was washed with brine, dried over Na₂SO₄, filtered, and concentrated. The residue was dissolved in a 1:1 mixture of THF and water (20 mL) and stirred at room temperature for 4 days, followed by extraction of the mixture with ethyl acetate (2 x 30 mL). The combined organic phases were washed with brine, dried over Na₂SO₄, filtered, and concentrated. The residue was purified by silica gel column chromatography (elution, PE / EA = 100:0-80:20) to give 4'-fluoro-1'-oxo-1',3'-dihydrospirocyclic [azacyclobutane--3,2'-indene]-1-carboxylic acid tert-butyl ester (430 mg, 1.48 mmol, yield 32%).

[0477] At 0 °C, 4'-fluoro-1'-oxy-1',3'-dihydrospirocyclic [azacyclobutane-3,2'-indene]-1-carboxylic acid tert-butyl ester (430 mg, 1.48 mmol, 1 eq.) was reacted with sodium borohydride (167.52 mg, 4.43 mmol, 3.0 eq.) in a methanol (10 mL) solution. The mixture was stirred at this temperature for 2 h. The solvent was removed by evaporation, and the residue was extracted with ethyl acetate (30 mL * 2). The combined organic layers were washed with brine, dried over Na₂SO₄, filtered, and concentrated to give 4'-fluoro-1'-hydroxy-1',3'-dihydrospirocyclic [azacyclobutane-3,2'-indene]-1-carboxylic acid tert-butyl ester (410 mg, 1.40 mmol, yield 94.69%) as a white solid.

[0478] A mixture of 4'-fluoro-1'-hydroxy-1',3'-dihydrospirocyclic [azacyclobutane-3,2'-indene]-1-carboxylic acid tert-butyl ester (410 mg, 1.40 mmol, 1 eq.) in acetic acid (10 mL) was added with boron trifluoride diethyl ether (2 mL) and triethylsilane (1.30 g, 11.18 mmol, 1.79 mL, 8 eq.). The mixture was stirred at 60 °C for 16 h, then cooled to 0 °C, and the pH of the mixture was adjusted to 9 by slow addition of 15% sodium hydroxide solution. The mixture was then extracted with ethyl acetate (30 mL x 3). The organic layers were combined, washed successively with NH4Cl (aq.) and brine, dried over Na2SO4, filtered, and concentrated to dryness to give 4'-fluoro-1',3'-dihydrospirocyclic [azacyclobutane-3,2'-indene] (245 mg, 1.38 mmol, yield 98.91%).

[0479] To a mixture of 4'-fluoro-1',3'-dihydrospirocyclic [azacyclobutane-3,2'-indene] (72 mg, 406.28 μmol, 1.2 eq.) in 1,4-dioxane (10 mL), methyl [(2R,3R,4R,5R)-3,4-diacetoxy-5-(2,6-dichloropurin-9-yl)tetrahydrofuran-2-yl]acetate (151.42 mg, 338.57 μmol, 1 eq.) and DIPEA (109.39 mg, 846.42 μmol, 147.43 μL, 2.5 eq.) was added. The mixture was stirred overnight at 100 °C. The solvent was removed by evaporation. The residue was diluted in EtOAc (50 mL), washed with brine, dried over Na2SO4, filtered, and concentrated. The residue was purified by silica gel column chromatography (elution, PE / EA = 100:0-50:50) to give (2R,3R,4R,5R)-2-(acetoxymethyl)-5-(2-chloro-6-(4'-fluoro-1',3'-dihydrospirocyclic [azacyclobutane-3,2'-indene]-1-yl)-9H-purin-9-yl)tetrahydrofuran-3,4-diacetate (100 mg, 170.07 μmol, yield 50.23%) as a white solid.

[0480] To a mixture of (2R,3R,4R,5R)-2-(acetoxymethyl)-5-(2-chloro-6-(4'-fluoro-1',3'-dihydrospirocyclic [azacyclobutane-3,2'-indene]-1-yl)-9H-purin-9-yl)tetrahydrofuran-3,4-diacetate (100 mg, 170.17 μmol, 1 eq.) in methanol (2 mL), NH3-MeOH (7 M, 728.88 μL, 30 eq.) was added. The mixture was stirred at room temperature for 2 hours. The solvent was removed by evaporation, and the residue was extracted with ethyl acetate (2 x 20 mL). The organic phases were combined, washed with brine, dried over Na2SO4, and concentrated to dryness to give (2R,3R,4S,5R)-2-(2-chloro-6-(4'-fluoro-1',3'-dihydrospirocyclic [azacyclobutane-3,2'-indene]-1-yl)-9H-purin-9-yl)-5-(hydroxymethyl)tetrahydrofuran-3,4-diol (78 mg, 168.88 μmol, yield 99.30%) as a white solid.

[0481] Add p-TsOH□H₂O (32.12 mg, 168.88 μmol, 1 eq.) and 2,2-dimethoxypropane (351.76 mg, 3.38 mmol, 20 eq.) to 10 mL of acetone solution of (2R,3R,4S,5R)-2-(2-chloro-6-(4'-fluoro-1',3'-dihydrospirocyclic[azacyclobutane-3,2'-indene]-1-yl)-9H-purine-9-yl)-5-(hydroxymethyl)tetrahydrofuran-3,4-diol (78 mg, 168.88 μmol, 1 eq.) and 2,2-dimethoxypropane (351.76 mg, 3.38 mmol, 20 eq.). Stir the mixture overnight at room temperature. Adjust the pH of the mixture to 9 by slowly adding NaHCO₃ at 0 °C. Remove the solvent by evaporation, and extract the residue with ethyl acetate (2 x 30 mL). Combine the organic layers, wash with brine, dry with Na₂SO₄, and concentrate. The residue was purified by silica gel column chromatography (elution, PE / EA = 100:0-50:50) to give a white solid product ((3aR,4R,6R,6aR)-6-(2-chloro-6-(4'-fluoro-1',3'-dihydrospirocyclic[azacyclobutane-3,2'-indene]-1-yl)-9H-purin-9-yl)-2,2-dimethyltetrahydrofurano[3,4-d][1,3]dioxono-4-yl)methanol (57 mg, 113.56 μmol, yield 67.24%).

[0482] To a solution of ((3aR,4R,6R,6aR)-6-(2-chloro-6-(4'-fluoro-1',3'-dihydrospirocyclic[azacyclobutane-3,2'-indene]-1-yl)-9H-purin-9-yl)-2,2-dimethyltetrahydrofurano[3,4-d][1,3]dioxono-4-yl)methanol (57 mg, 113.56 μmol, 1 eq.) cooled at 0 °C in PO(MeO)3 (1 mL), bis(dichlorophosphoryl)methane (70.91 mg, 283.90 μmol, 2.5 eq.) in PO(MeO)3 (1 mL) was added. The mixture was stirred at 0 °C for 4 hours. Then, 1.5 mL of water was slowly added at 0 °C. The mixture was stirred at 40 °C for 40 minutes, followed by stirring at 25 °C overnight. The reaction mixture was purified by C-18 reversed-phase silica column chromatography (0–25% ACN in water) to give compound d-11 (10 mg, yield 14.25%). 1HNMR(500MHz,MeOD)δppm 2.53(t,J=20.6Hz,2H),3.35(s,4H),4.25–4.41(m,5H),4.45(s,1H),4.60(s,2H),4.67(s,1H),6.04(d,J=4.0Hz ,1H),6.91(t,J=8.5Hz,1H),7.09(d,J=7.1Hz,1H),7.22(d,J=5.4Hz,1H),8.47(s,1H); m / z(ESI+):620.1(M+H).

[0483] Example 35. Synthesis of compound d-12

[0484]

[0485] A mixture of tetrahydronaphthalene-1-carboxylic acid (2 g, 11.35 mmol, 1 eq.) and H₂SO₄ (1.11 g, 11.35 mmol, 605.00 μL, 1 eq.) in methanol (20 mL) was heated at 65 °C for 20 hours. The resulting solution was concentrated. The residue was dissolved in ethyl acetate (50 mL), and the solution was washed with brine, dried over Na₂SO₄, filtered, and concentrated to dryness to give methyl tetrahydronaphthalene-1-carboxylic acid (2.1 g, 97.2%).

[0486] Under nitrogen protection, potassium carbonate (5.03 g, 36.43 mmol, 3.3 eq.) and HCHO (3.05 g, 33.12 mmol, 37% purity, 3 eq.) were added to a mixture of tetrahydronaphthalene-1-carboxylic acid methyl ester (2.1 g, 11.04 mmol, 1 eq.) and DMSO (25 mL). The mixture was stirred at room temperature for 19 h. TLC showed that the starting material was consumed and the reaction was quenched with water (75 mL). The mixture was extracted with ethyl acetate (50 mL). After adjusting the pH of the aqueous layer to 3 with 3N hydrochloric acid, ethyl acetate (80 mL * 3) was added for extraction. The organic layers were combined, washed with brine, dried over Na2SO4, filtered, and evaporated to dryness to give 1-(hydroxymethyl)tetrahydronaphthalene-1-carboxylic acid (1.5 g, 65.9%).

[0487] To a mixture of 1-(hydroxymethyl)tetrahydronaphthalene-1-carboxylic acid (1.5 g, 7.27 mmol, 1 eq.) and benzylamine (779.34 mg, 1 eq.), in DMF (15 mL), EDCI (2.09 g, 10.91 mmol, 1.5 eq.), HOBT (1.47 g, 10.91 mmol, 1.5 eq.), and DIPEA (1.41 g, 10.91 mmol, 1.90 mL, 1.5 eq.) were added. The mixture was stirred at room temperature for 18 hours. The solvent was removed by evaporation, and then 50 mL of water was added to react the mixture, followed by ethyl acetate (60 mL). The organic layer was washed with brine, dried over Na2SO4, filtered, and evaporated to dryness. The residue was purified by silica gel column chromatography (elution, PE / EtOAc = 100:0-70:30) to give N-benzyl-1-(hydroxymethyl)tetrahydronaphthalene-1-carboxamide (1.3 g, yield 60.5%) in oil form.

[0488] Triphenylphosphine (1.73 g, 6.60 mmol, 1 eq.) was added to a mixture of N-benzyl-1-(hydroxymethyl)tetrahydronaphthalene-1-carboxamide (1.3 g, 4.40 mmol, 1 eq.) and tetrahydrofuran (20 mL) under nitrogen protection at 0 °C, followed by the addition of DEAD (1.15 g, 6.60 mmol, 1.04 mL, 1.5 eq.). The mixture was stirred at room temperature for 2 hours. The reaction was confirmed by TLC to be complete. The resulting solution was quenched with water (20 mL) and then ethyl acetate (60 mL) was added. The organic layer was separated, washed with (saline) Na₂SO₄, dried, filtered, and evaporated to dryness. The residue was purified by silica gel column chromatography (elution, PE / EtOAc = 100:0-67:33) to give the product 1-benzylspirocyclic [azacyclobutane-3,1'-tetrahydronaphthalene]-2-one (1 g, yield 81.9%).

[0489] A solution of aluminum trichloride (961.50 mg, 7.21 mmol, 2.0 eq.) in tetrahydrofuran (15 mL) was cooled to 0 °C, and then lithium aluminum hydride (410.48 mg, 10.82 mmol, 3.0 eq.) was added. The mixture was stirred at 0 °C for 30 min, followed by the addition of 1-benzylspirocyclic [azacyclobutane-3,1'-tetrahydronaphthalene]-2-one (1 g, 3.61 mmol, 1 eq.) in 4 mL of THF. The mixture was stirred overnight at room temperature. The reaction was quenched with water (1 mL), followed by the addition of 15% aqueous sodium hydroxide solution (4 mL) and ethyl acetate (20 mL). The solid was removed by filtration, and the organic layer was concentrated to give 1-benzylspirocyclic [azacyclobutane-3,1'-tetrahydronaphthalene] (800 mg, yield 84.2%).

[0490] Ammonium formate (287.32 mg, 4.56 mmol, 1.5 eq.) and palladium hydroxide (100 mg) were added to a solution of 1-benzylspirocyclic [azacyclobutane-3,1'-tetrahydronaphthalene] (800 mg, 3.04 mmol, 1 eq.) in methanol (30 mL). The mixture was stirred overnight at 60 °C under hydrogen atmosphere. The mixture was filtered, and the residue was washed with methanol. The filtrate and washings were combined and concentrated to give spirocyclic [azacyclobutane-3,1'-tetrahydronaphthalene] (500 mg, 95.0%).

[0491] Methyl acetate [(2R,3R,4R,5R)-3,4-diacetoxy-5-(2,6-dichloropurin-9-yl)tetrahydrofuran-2-yl]acetate (500 mg, 1.12 mmol, 1 eq.) in 1,4-dioxane (20 mL) was mixed with spirocyclic [azacyclobutane-3,1'-tetrahydronaphthalene] (251.81 mg, 1.45 mmol, 1.3 eq.) and DIPEA (577.96 mg, 4.47 mmol, 778.93 μL, 4 eq.). The mixture was stirred at 100 °C for 3 hours. The mixture was concentrated, and the residue was purified by silica gel column chromatography (elution, PE / EtOAc = 100:0-50:50) to give methyl [(2R,3R,4R,5R)-3,4-diacetoxy-5-(2-chloro-6-spirocyclic [azacyclobutane-3,1'-tetrahydronaphthalene]-1-ylpurine-9-yl)tetrahydrofuran-2-yl]acetate (500 mg, yield 76.5%) as a white solid.

[0492] Methyl [(2R,3R,4R,5R)-3,4-diacetoxy-5-(2-chloro-6-spirocyclic [azacyclobutane-3,1'-tetrahydronaphthalene]-1-ylpurine-9-yl)tetrahydrofuran-2-yl]acetate (500 mg, 856.14 μmol, 1 eq.) was added to a methanol (4 mL) solution of MeOH-NH3 (7 M, 3.67 mL, 30 eq.). The mixture was stirred at room temperature for 3 hours. The solvent was removed by evaporation, the residue was diluted with ethyl acetate (100 mL), the organic layer was washed with water (70 mL), then washed with brine (60 mL), dried over Na2SO4, and concentrated to give (2R,3R,4S,5R)-2-(2-chloro-6-spirocyclic [azacyclobutane-3,1'-tetrahydronaphthalene]-1-ylpurine-9-yl)-5-(hydroxymethyl)tetrahydrofuran-3,4-diol (390 mg, 851.99.4%).

[0493] To a solution of (2R,3R,4S,5R)-2-(2-chloro-6-spirocyclic [azacyclobutane-3,1'-tetrahydronaphthalene]-1-ylpurine-9-yl)-5-(hydroxymethyl)tetrahydrofuran-3,4-diol (391 mg, 853.88 μmol, 1 eq.) in acetone (30 mL), 2,2-dimethoxypropane (1.78 g, 17.08 mmol, 20 eq.) and p-TsOH (147.04 mg, 853.88 μmol, 1 eq.) were added. The mixture was stirred at room temperature for 3 hours. The solvent was removed by evaporation. The residue was diluted with EtOAc (50 mL), washed with an aqueous solution of NaHCO3, and then with brine. The organic layer was concentrated, and the residue was purified by silica gel column chromatography (elution, PE / EtOAc = 100:0-50:50) to give [(3aR,4R,6R,6aR)-4-(2-chloro-6-spirocyclic [azacyclobutane-3,1'-tetrahydronaphthyl]-1-ylpurine-9-yl)-2,2-dimethyl-3a,4,6,6a-tetrahydrofurano[3,4-d][1,3]dioxono-6-yl]methanol (350 mg, yield 82.3%) as a white solid.

[0494] [(3aR,4R,6R,6aR)-4-(2-chloro-6-spirocyclic [azacyclobutane-3,1'-tetrahydronaphthalene]-1-ylpurine-9-yl)-2,2-dimethyl-3a,4,6,6a-tetrahydrofurano[3,4-d][1,3]dioxono-6-yl]methanol (350 mg, 702.85 μmol, 1 eq.) in PO(OEt)3 (4 mL) was cooled to 0 °C, and then bis(dichlorophosphoryl)methane (351.12 mg, 1.41 mmol, 2.0 eq.) in PO(OEt)3 (3 mL) was added. The mixture was stirred at 0 °C for 5 hours, and then water (5 mL) was added at 0 °C. The mixture was stirred at 40 °C for 30 minutes, and then stirred overnight at room temperature. The reaction mixture was purified by reversed-phase C-18 silica gel column chromatography (gradient elution, 0 to 25% ACN in water) to give compound d-12 (253 mg, yield 57.4%). 1HNMR (500MHz, CD3OD) δppm 2.04(s,2H),2.38(s,2H),2.67(t,J=20.9Hz,2H),3.02(t,J=6.1Hz,2H),4.4 5(s,1H),4.47-4.52(m,1H),4.55(dd,J=10.4,7.0Hz,1H),4.62(t,J=4.8Hz, 1H),4.84(t,J=5.0Hz,1H),6.21(d,J=4.8Hz,1H),7.28(d,J=7.5Hz,1H),7.3 4(t,J=7.4Hz,1H),7.42(t,J=7.5Hz,1H),7.81(d,J=7.9Hz,1H),8.58(s,1H); 13 C NMR(125MHz,CD3OD)δppm 21.29,26.46,27.51,28.56,30.92,37.23,40.00,65.91,71.33,75.71,84.63,89.84 ,118.67,127.05,127.89,130.14,138.07,140.81,141.23,151.77,155.59,155.76; 31 P NMR (203MHz, CD3OD) δppm16.86,19.80; m / z (ESI + ):616.3(M+H).

[0495] Example 36. Synthesis of compound d-13

[0496]

[0497] At -78 °C, LiHMDS (1 M, 33.05 mL, 2.2 eq.) was added dropwise to a mixture of indolin-2-one (2 g, 15.02 mmol, 1.0 eq.) in THF (30 mL). The reaction temperature was raised to -50 °C and maintained at -50 °C for 30 min. The mixture was then cooled again to -78 °C, followed by the addition of 1,4-dibromobutane (3.24 g, 15.02 mmol, 1 eq.) in 15 mL of THF. The mixture was stirred at room temperature for 2 h, then stirred under reflux for 2 h, and finally stirred at room temperature for 16 h. The mixture was concentrated under reduced pressure, and the residue was partitioned between ethyl acetate and saturated ammonium chloride. The concentrated organic layer was purified by silica gel column chromatography (elution, PE / EtOAc = 100:0-60:40) to obtain a yellow solid spirocyclic [cyclopentane-1,3'-indoline]-2'-one (1.29 g, 45.87%).

[0498] LiAlH4 (522.92 mg, 13.78 mmol, 2.0 eq.) was added to a solution of spirocyclic [cyclopentane-1,3'-indoline]-2'-one (1.29 g, 6.89 mmol, 1 eq.) in THF (30 mL). The mixture was stirred overnight at 70 °C. The mixture was quenched with water (1 mL) and EtOAc (20 mL) was added. The solid was removed by filtration and the organic layer was concentrated. The residue was purified by silica gel column chromatography (elution, PE / EtOAc = 100:0-80:20) to give a white solid of spirocyclic [cyclopentane-1,3'-indoline] (1.02 g, 85.45%).

[0499] To a solution of methyl [(2R,3R,4R,5R)-3,4-diacetoxy-5-(2,6-dichloropurin-9-yl)tetrahydrofuran-2-yl]acetate (1.2 g, 2.68 mmol, 1 eq.) in 1,4-dioxane (25 mL), spirocyclic [cyclopentane-1,3'-indoline] (511 mg, 2.95 mmol, 1.10 eq.) and DIPEA (866.94 mg, 6.71 mmol, 1.17 mL, 2.5 eq.) were added. The mixture was stirred overnight at 100 °C. The mixture was concentrated and purified by silica gel column chromatography (elution, PE / EtOAc = 100:0-60:40) to give methyl [(2R,3R,4R,5R)-3,4-diacetoxy-5-(2-chloro-6-spiro[cyclopentan-1,3'-indoline]-1'-ylpurine-9-yl)tetrahydrofuran-2-yl]acetate (1.49 g, yield 95.08%).

[0500] Ammonia-methanol (7M, 5.81 mL, 25 eq.) was added to a solution of [(2R,3R,4R,5R)-3,4-diacetoxy-5-(2-chloro-6-spiro[1,3'-indoline]-1'-ylpurine-9-yl)tetrahydrofuran-2-yl]acetate (950 mg, 1.63 mmol, 1 eq.) in methanol (15 mL). The mixture was stirred at room temperature for 4 hours. The solid was obtained by filtration, and the filter cake was washed with methanol (20 mL) to give the product (2R,3R,4S,5R)-2-(2-chloro-6-spirocyclo[1,3'-indoline]-1'-ylpurine-9-yl)-5-(hydroxymethyl)tetrahydrofuran-3,4-diol (580 mg, 77.87% yield).

[0501] Add 2,2-dimethoxypropane (1.98 g, 19.00 mmol, 15 eq.) and p-toluenesulfonic acid (218.11 mg, 1.27 mmol, 1 eq.) to a mixture of (2R,3R,4S,5R)-2-(2-chloro-6-spirocyclopentan-1,3'-indole]-1'-ylpurine-9-yl)-5-(hydroxymethyl)tetrahydrofuran-3,4-diol (580 mg, 1.27 mmol, 1 eq.) in acetone (20 mL). Stir the mixture at room temperature for 2 hours. Evaporate the solvent, dilute the residue with EtOAc (50 mL), wash with an aqueous sodium bicarbonate solution, and then wash with brine. The organic layer was concentrated, and the residue was purified by silica gel column chromatography (elution, PE / EtOAc = 100:0-50:50) to give [(3aR,4R,6R,6aR)-4-(2-chloro-6-spirocyclopentan-1,3'-indoline]-1'-ylpurine-9-yl)-2,2-dimethyl-3a,4,6,6a-tetrahydrofurano[3,4-d][1,3]dioxono-6-yl]methanol (540 mg, yield 85.61%).

[0502] A solution of [(3aR,4R,4R,6R,6aR)-4-(2-chloro-6-spirocyclopentan-1,3'-indoline]-1'-ylpurine-9-yl)-2,2-dimethyl-3a,4,4,6,6a-tetrahydrofurano[3,4-d][1,3]dioxono-6-yl]methanol (540 mg, 1.08 mmol, 1 eq.) in trimethyl phosphate (3 mL) was cooled to 0 °C, followed by the addition of a solution of bis(dichlorophosphoryl)methane (812.60 mg, 3.25 mmol, 3.0 eq.) in trimethyl phosphate (3 mL). The mixture was stirred at 0 °C for 5 h. Water (4 mL) was then added to the reaction mixture. The mixture was stirred overnight at room temperature. The reaction mixture was purified by C-18 reversed-phase silica gel (0 to 25% ACN in water) to give compound d-13 (149.9 mg, yield 21.99%). 1 H NMR(500MHz,MeOD)δppm 1.90(dd,J=17.7,12.4Hz,8H),2.52(t,J=20.9Hz,2H),4.28(s,1H),4.36(dtd,J=15.0,11.2,4.7Hz,2H),4.46(t,J=4.8Hz, 1H),4.55(s,2H),4.66(t,J=4.9Hz,1H),6.06(d,J=4.7Hz,1H),7.07(t,J=7.4Hz,1H),7.18-7.27(m,2H),8.40-8.53(m,2H); 31P NMR (203MHz, MeOD) δppm16.81,19.86; 13 C NMR(125MHz,MeOD)δppm 24.48,26.15,27.21,40.47,51.52,64.71,64.80,70.10,74.33,83.26,88.31,117.29,119.01,12 1.96,123.89,126.97,139.49,140.10,142.36,151.13,152.00,153.21; m / z(ESI+):616.3(M+H).

[0503] Example 37. Synthesis of compound d-14

[0504]

[0505] A solution of 4-fluoroindolin-2-one (1 g, 6.62 mmol, 1 eq.) in tetrahydrofuran (20 mL) was cooled to -78 °C, and then LiHMDS (1 M, 14.56 mL, 2.2 eq.) was added. The mixture was stirred at -50 °C for 30 min, and then cooled again to -78 °C. At -78 °C, a solution of 1,4-dibromobutane (1.43 g, 6.62 mmol, 1 eq. dissolved in 10 mL of tetrahydrofuran) was added to the mixture. The mixture was stirred at room temperature for 2 h, then at 70 °C for 3 h, and then stirred overnight at room temperature. After completion, the reaction was quenched with an aqueous solution of ammonium chloride (50 mL) and diluted with ethyl acetate (60 mL). The organic layer was separated, washed with brine, dried with Na2SO4, filtered, and purified by silica gel column chromatography (elution, PE / EA = 100:0-55:45) to give 4'-fluorospirocyclic [cyclopentane-1,3'-indoline]-2'-one (750 mg, 55.23%) as a yellow solid.

[0506] Lithium aluminum hydride (369.83 mg, 9.75 mmol, 4.0 eq.) was added to a solution of 4'-fluorospirocyclic [cyclopentane-1,3'-indoline]-2'-one (500 mg, 2.44 mmol, 1 eq.) in tetrahydrofuran (15 mL). The mixture was stirred at 70 °C for 2 hours. The reaction was quenched by adding water (0.3 mL) and 15% sodium hydroxide aqueous solution (0.6 mL), followed by the addition of ethyl acetate (30 mL). The solid was removed by filtration, and the organic layer was concentrated to give 4'-fluorospirocyclic [cyclopentane-1,3'-indoline] (440 mg, yield 94.4%).

[0507] DIPEA (693.56 mg, 5.37 mmol, 934.71 μL, 3.0 eq.) and 4'-fluorospirocyclic [cyclopentane-1,3'-indoline] (410.52 mg, 2.15 mmol, 1.2 eq.) were added to a solution of [(2R,3R,4R,5R)-3,4-diacetoxy-5-(2,6-dichloropurin-9-yl)tetrahydrofuran-2-yl]acetate (800 mg, 1.79 mmol, 1 eq.) in 1,4-dioxane (20 mL). The mixture was stirred overnight at 100 °C and then concentrated. The residue was purified by silica gel column chromatography (elution, PE / EA = 100:0-60:40) to give methyl [(2R,3R,4R,5R)-3,4-diacetoxy-5-[2-chloro-6-(4'-fluorospiro[cyclopentan-1,3'-indoline]-1'-yl)purin-9-yl]tetrahydrofuran-2-yl]acetate (0.9 g, yield 83.58%) as a pale yellow solid.

[0508] Methyl [(2R,3R,4R,5R)-3,4-diacetoxy-5-[2-chloro-6-(4'-fluorospiro[cyclopentan-1,3'-indoline]-1'-yl)purin-9-yl]tetrahydrofuran-2-yl]acetate (900 mg, 1.49 mmol, 1 eq.) in a methanol (6 mL) solution was added to ammonia-methanol (7 M, 6.41 mL, 30 eq.). The mixture was stirred overnight at room temperature. The solid obtained by filtration was (2R,3R,4S,5R)-2-[2-chloro-6-(4'-fluorospiro[cyclopentan-1,3'-indoline]-1'-yl)purin-9-yl]-5-(hydroxymethyl)tetrahydrofuran-3,4-diol (700 mg, 98.4%) as a white solid.

[0509] To a solution of (2R,3R,4S,5R)-2-[2-chloro-6-(4'-fluorospiro[cyclopentan-1,3'-indoline]-1'-yl)purin-9-yl]-5-(hydroxymethyl)tetrahydrofuran-3,4-diol (700 mg, 1.47 mmol, 1 eq.) in acetone (20 mL), 2,2-dimethoxypropane (2.30 g, 22.06 mmol, 2.71 mL, 15 eq.) and p-toluenesulfonic acid (253.29 mg, 1.47 mmol, 1 eq.) were added. The mixture was stirred at room temperature for 3 hours. The mixture was diluted with EtOAc (50 mL), and the organic layer was washed with an aqueous sodium bicarbonate solution, followed by washing with brine, and then concentrated. The residue was purified by silica gel column chromatography (elution, PE / EtOAc = 100:0-50:50) to give [(3aR,4R,6R,6aR)-4-[2-chloro-6-(4'-fluorospiro[cyclopentan-1,3'-indoline]-1'-yl)purin-9-yl]-2,2-dimethyl-3a,4,6,6a-tetrahydrofurano[3,4-d][1,3]dioxono-6-yl]methanol (600 mg, 79.06%).

[0510] A solution of [(3aR,4R,6R,6aR)-4-[2-chloro-6-(4'-fluorospiro[cyclopentan-1,3'-indoline]-1'-yl)purin-9-yl]-2,2-dimethyl-3a,4,6,6a-tetrahydrofurano[3,4-d][1,3]dioxono-6-yl]methanol (300 mg, 581.44 μmol, 1 eq.) in triethyl phosphate (3 mL) was cooled to 0 °C, and then bis(dichlorophosphoryl)methane (290.47 mg, 1.16 mmol, 2.0 eq.) in 2 mL of trimethyl phosphate was added. The mixture was stirred at room temperature for 4 hours, and then water (3 mL) was added. The mixture was stirred at room temperature overnight. The reaction mixture was purified by C-18 reversed-phase silica gel (0 to 30% ACN in water) to give compound d-14 (195 mg, 52.9% yield). 1 H NMR(500MHz,CD3OD)δppm 1.84(d,J=20.7Hz,6H),2.08(d,J=31.3Hz,2H),2.49(t,J=20.3Hz,2H),4.30(s,3H),4.46(d,J=10.2Hz,3H),4.6 5(s,1H),6.04(d,J=4.6Hz,1H),6.73(t,J=9.0Hz,1H),7.15(d,J=5.9Hz,1H),8.19(d,J=8.1Hz,1H),8.46(s,1H); 13C NMR(125MHz,CD3OD)δppm 25.99,27.50,40.04,51.81,65.57,67.04,71.73,75.77,88.96,112.08,1 14.79,120.06,130.02,140.94,145.47,152.11,152.26,153.25,154.30; 31 P NMR (203MHz, DMSO-d6) δppm 14.72, 18.24; m / z (ESI + ):634.2(M+H).

[0511] Example 38. Synthesis of compound d-15

[0512]

[0513] At -78°C, LiHMDS (1M, 7.28mL, 2.2eq.) was added dropwise to a mixture of 5-fluoroindolin-2-one (500.00mg, 3.31mmol, 1eq.) and THF (30mL). The mixture was then heated to -50°C and held at -50°C for 30 minutes. It was then cooled again to -78°C. 1,4-Dibromobutane (714.29mg, 3.31mmol, 1eq.) in THF (15mL) was then added. The mixture was first stirred at room temperature for 1 hour, then stirred under reflux for 3 hours, and finally stirred at room temperature for 16 hours. The mixture was concentrated under reduced pressure, and the residue was partitioned between ethyl acetate and saturated ammonium chloride. The organic layer was concentrated, and the residue was purified by silica gel column chromatography (elution, PE / EtOAc = 100:0-60:40) to give 5'-fluorospirocyclic [cyclopentane-1,3'-indoline]-2'-one (350 mg, yield 51.55%).

[0514] LiAlH4 (161.80 mg, 4.26 mmol, 2.5 eq.) was added to a solution of 5'-fluorospirocyclic [cyclopentane-1,3'-indoline]-2'-one (350.00 mg, 1.71 mmol, 1 eq.) in THF (30 mL). The mixture was stirred at 70 °C for 2 h. The reaction was quenched at 0 °C with water (1 mL) and 15% sodium hydroxide aqueous solution (1 mL), followed by the addition of ethyl acetate (40 mL). The solid was removed by filtration and the organic layer was concentrated. The residue was purified by silica gel column chromatography (elution, PE / EtOAc = 100:0-78:22) to give 5'-fluorospirocyclic [cyclopentane-1,3'-indoline] (210 mg, yield 64.39%) as a white solid.

[0515] Add 5'-fluorospirocyclopentane-1,3'-indoline](210.26 mg, 1.10 mmol, 1.1 eq.) and DIPEA(322.94 mg, 2.50 mmol, 435.22 μL, 2.5 eq.) to a solution of [(2R,3R,4R,5R)-3,4-diacetoxy-5-(2,6-dichloropurin-9-yl)tetrahydrofuran-2-yl]acetate (447 mg, 999.49 μmol, 1 eq.) in 1,4-dioxane (25 mL) to a solution of methyl [(2R,3R,4R,5R)-3,4-diacetoxy-5-(2,6-dichloropurin-9-yl)tetrahydrofuran-2-yl]acetate (210.26 mg, 1.10 mmol, 1.1 eq.) and DIPEA (322.94 mg, 2.50 mmol, 435.22 μL, 2.5 eq.) in 1,4-dioxane (25 mL). Stir the mixture at 100 °C for 4 hours. The mixture was concentrated and purified by silica gel column chromatography (elution, PE / EtOAc = 100:0-58:42) to give methyl [(2R,3R,4R,5R)-3,4-diacetoxy-5-[2-chloro-6-(5'-fluorospirocyclopentan-1,3'-indoline]-1'-yl]purin-9-yl]tetrahydrofuran-2-yl]acetate (490 mg, 813.94 μmol, yield 81.44%).

[0516] Ammonia-methanol (7M, 2.91 mL, 25 eq.) was added to a solution of methyl [(2R,3R,4R,5R)-3,4-diacetoxy-5-[2-chloro-6-(5'-fluorospiro[cyclopentan-1,3'-indoline]-1'-yl)purin-9-yl]tetrahydrofuran-2-yl]acetate (490.00 mg, 813.94 μmol, 1 eq.) in methanol (15 mL). The mixture was stirred at room temperature for 4 hours. The solid was obtained by filtration, and the filter cake was washed with methanol (10 mL) to give the product (2R,3R,4S,5R)-2-[2-chloro-6-(5'-fluorospiro[1,3'-indole]-1'-yl)purin-9-yl]-5-(hydroxymethyl)tetrahydrofuran-3,4-diol (360 mg, 756.46 μmol, yield 92.94%).

[0517] Add 2,2-dimethoxypropane (1.18 g, 11.35 mmol, 15 eq.) and p-toluenesulfonic acid (130.26 mg, 756.46 μmol, 1 eq.) to a mixture of (2R,3R,4S,5R)-2-[2-chloro-6-(5'-fluorospirocyclopentan-1,3'-indole]-1'-yl)purine-9-yl]-5-(hydroxymethyl)tetrahydrofuran-3,4-diol (360 mg, 756.46 μmol, 1 eq.) in acetone (25 mL). Stir the mixture at room temperature for 2 hours. Remove the solvent by evaporation, dilute the residue with EA (50 mL), wash with an aqueous sodium bicarbonate solution, and then wash with brine. The organic layer was concentrated and purified by silica gel column chromatography (elution, PE / EtOAc = 100:0-50:50) to give [(3aR,4R,6R,6aR)-4-[2-chloro-6-(5'-fluorospirocyclopentan-1,3'-indoline]-1'-yl)purin-9-yl]-2,2-dimethyl-3a,4,6,6a-tetrahydrofurano[3,4-d][1,3]dioxono-6-yl]methanol (310 mg, 600.82 μmol, yield 79.42%).

[0518] A solution of [(3aR,4R,6R,6aR)-4-[2-chloro-6-(5'-fluorospirocyclopentan-1,3'-indole]-1'-yl)purine-9-yl]-2,2-dimethyl-3a,4,6,6a-tetrahydrofurano[3,4-d][1,3]dioxono-6-yl]methanol (310 mg, 600.82 μmol, 1 eq.) in trimethyl phosphate (3 mL) was cooled to 0 °C, and then a solution of bis(dichlorophosphoryl)methane (450.23 mg, 1.80 mmol, 3 eq.) in trimethyl phosphate (3 mL) was added to the mixture. The mixture was stirred at 0 °C for 5 h. Then water (4 mL) was added to the reaction mixture. The mixture was stirred overnight at room temperature. The reaction mixture was purified by C-18 reversed-phase silica gel (0 to 25% ACN in water) to give compound d-15 (129.9 mg, yield 33.99%, purity 99.65%). 1 HNMR(500MHz,MeOD)δppm 1.90(d,J=25.6Hz,8H),2.52(t,J=20.9Hz,2H),4.28(s,1H),4.34(dd,J=22.5,15.9Hz,2H),4.46(d,J=4.6Hz,1H),4.60( s,2H),4.66(s,1H),6.06(d,J=4.8Hz,1H),6.95(d,J=2.5Hz,1H),7.01–7.08(m,1H),8.43(s,1H),8.49(d,J=4.1Hz,1H);13 C NMR(126MHz,MeOD)δppm 24.40,40.31,51.52,69.49,70.68,73.69,74.89,87.63,88.95,117.58,118.92,138.71,140.45,142.59,150.89,151.99,153.16; 31 P NMR (203MHz, MeOD) δppm 16.76, 20.03; m / z (ESI + ):634.2(M+H).

[0519] Example 39. Synthesis of compound d-16

[0520]

[0521] Under a nitrogen atmosphere, at -78 °C, LiHMDS (1 M, 7.28 mL, 2.2 eq.) was added dropwise to a mixture of 6-fluoroindolin-2-one (500 mg, 3.31 mmol, 1 eq.) and tetrahydrofuran (5 mL). The mixture was then heated to -50 °C and maintained at that temperature for 30 minutes. The mixture was cooled again to -78 °C, and then a solution of 1,4-dibromobutane (714.30 mg, 3.31 mmol, 1.0 eq.) in THF (5 mL) was added. The mixture was first stirred at room temperature for 1 hour, and then stirred under reflux for 2 hours. The mixture was quenched with saturated ammonium chloride solution and extracted with ethyl acetate (30 mL * 2). The organic layer was washed with brine and concentrated. The residue was purified by silica gel column chromatography (elution, PE / EtOAc = 100:0-70:30) to give the product 6'-fluorospirocyclic [cyclopentane-1,3'-indole]-2'-one (282 mg, 1.37 mmol, yield 41.54%).

[0522] A solution of AlCl3 (493.79 mg, 3.70 mmol, 2 eq.) in 10 mL of THF was cooled to 0 °C, and then LiAlH4 (210.80 mg, 5.55 mmol, 3 eq.) was added. The mixture was stirred at 0 °C for 30 minutes. Then, a solution of 6'-fluorospirocyclic [cyclopentane-1,3'-indole]-2'-one (380 mg, 1.85 mmol, 1.0 eq.) in 4 mL of THF was added. The mixture was stirred at room temperature overnight. The mixture was diluted with 10 mL of THF, and the reaction was quenched at 0 °C by slowly adding 15% NaOH (aq.) until the pH reached 9. The organic layer was dried over magnesium sulfate. The solids were removed by filtration. The filtrate was concentrated, and the residue was purified by silica gel column chromatography (elution, PE / EA = 100:0-90:10) to give the product 6'-fluorospirocyclic [cyclopentane-1,3'-indoline] (250 mg, 1.31 mmol, yield 70.60%).

[0523] Methyl [(2R,3R,4R,5R)-3,4-diacetoxy-5-(2,6-dichloropurin-9-yl)tetrahydrofuran-2-yl]acetate (487.19 mg, 1.09 mmol, 1 eq.) and DIPEA (351.97 mg, 2.72 mmol, 474.36 μL, 2.5 eq.) were added to a mixture of 6'-fluorospirocyclic [cyclopentane-1,3'-indoline] (250 mg, 1.31 mmol, 1.2 eq.) and 1,4-dioxane (15 mL). The mixture was stirred overnight at 100 °C. TLC showed approximately 30% of the starting material remaining and the formation of a new product. The mixture was stirred at 120 °C for 3 hours and then cooled to room temperature. The solvent was removed by evaporation. The residue was diluted with ethyl acetate (40 mL), washed first with water and then with brine. The organic layer was concentrated, and the residue was purified by silica gel column chromatography (elution, PE / EA = 100:0-60:40) to give (2R,3R,4R,5R)-2-(acetoxymethyl)-5-(2-chloro-6-(6'-fluorospiro[cyclopentane-1,3'-indole]-1'-yl)-9H-purin-9-yl)tetrahydrofuran-3,4-diacetate (520 mg, 863.77 μmol, yield 79.29%) as a light yellow solid.

[0524] Ammonia-methanol (7M, 3.70 mL, 30 eq.) was added to a mixture of (2R,3R,4R,5R)-2-(acetoxymethyl)-5-(2-chloro-6-(6'-fluorospiro[cyclopentan-1,3'-indole]-1'-yl)-9H-purin-9-yl)tetrahydrofuran-3,4-diacetate (520 mg, 863.77 μmol, 1 eq.) in methanol (5 mL). The mixture was stirred at room temperature for 4 hours. LC-MS showed that the intermediate was not consumed, and the mixture was then stirred overnight at room temperature. At this point, analysis (LC-MS) indicated that the reaction was complete. The solvent was removed by evaporation, and the residue was used directly in the next step without further purification.

[0525] To a solution of (2R,3R,4S,5R)-2-(2-chloro-6-(6'-fluorospiro[cyclopentan-1,3'-indole]-1'-yl)-9H-purin-9-yl)-5-(hydroxymethyl)tetrahydrofuran-3,4-diol (410 mg, 861.53 μmol, 1 eq.) in acetone (20 mL), p-toluenesulfonic acid monohydrate (165.41 mg, 861.53 μmol, 1 eq.) and 2,2-dimethoxypropane (1.79 g, 17.23 mmol, 2.12 mL, 20 eq.) were added. The mixture was stirred at room temperature for 2 hours. LC-MS showed SM consumption. The pH of the mixture was adjusted to 9 at 0 °C by slow addition of 15% aqueous sodium hydroxide solution. The solvent was removed by evaporation, and the residue was extracted with EtOAc (2 x 30 mL). The organic layer was washed with brine and concentrated. The residue was purified by silica gel column chromatography (elution, PE / EA = 100:0-50:50) to give [(3aR,4R,6R,6aR)-6-(2-chloro-6-(6'-fluorospirocyclopentane-1,3'-indole]-1'-yl)purin-9-yl]-2,2-dimethyl-3a,4,6,6a-tetrahydrofurano[3,4-d][1,3]dioxono-6-yl)methanol (410 mg, 794.63 μmol, yield 92.24%) as a white solid.

[0526] A solution of [(3aR,4R,6R,6aR)-6-(2-chloro-6-(6'-fluorospirocyclopentan-1,3'-indole]-1'-yl)purin-9-yl]-2,2-dimethyl-3a,4,6,6a-tetrahydrofurano[3,4-d][1,3]dioxono-6-yl)methanol (380 mg, 736.49 μmol, 1 eq.) in triethyl phosphate (3 mL) was cooled to 0 °C, and then a solution of bis(dichlorophosphoryl)methane (367.93 mg, 1.47 mmol, 2.0 eq.) in 3 mL of trimethyl phosphate was added. The mixture was stirred at 0 °C for 4 hours. LC-MS showed that only a small amount of the starting material remained. Water (3 mL) was slowly added dropwise to the mixture at 0 °C. The mixture was first stirred at 40 °C for 40 minutes, and then stirred at room temperature overnight. LC-MS showed that the intermediate was consumed and the product was detected. The reaction mixture was purified by C-18 reversed-phase silica gel (0 to 30% ACN in water) to give compound d-16 (255 mg, yield 54.24%, with a purity of 99.29%). 1 H NMR(500MHz,MeOD)δppm 1.91(s,8H),2.57(s,2H),4.31(s,1H),4.34–4.46(m,2H),4.49(s,1H),4.57(s,2H),4.69(s,1H),6.08 (d,J=4.6Hz,1H),6.77(dd,J=11.7,5.2Hz,1H),7.20(dd,J=8.0,5.7Hz,1H),8.20(s,1H),8.46(s,1H); 31 P NMR(203MHz,MeOD)δppm16.85,20.04; 13 C NMR(126MHz,MeOD)δppm 24.31,25.03,26.09,27.15,40.61,50.88,64.73,65.35,70.07,74.34,83.17,88.33,104.93,10 9.85,118.96,122.60,135.54,139.88,143.43,150.89,152.06,153.00,161.09,162.99; m / z(ESI + ):634.3(M+H).

[0527] Example 40. Synthesis of compound d-17

[0528]

[0529] A solution of 7-fluoroindolin-2-one (600 mg, 3.97 mmol, 1 eq.) in tetrahydrofuran (25 mL) was cooled to approximately -78 °C, and then LiHMDS (1 M, 8.73 mL, 2.2 eq.) was added. The mixture was stirred at approximately -50 °C for 30 minutes, and then cooled to -78 °C. At -78 °C, a solution of 1,4-dibromobutane (857.16 mg, 3.97 mmol, 1 eq.) in THF (5 mL) was added to the cooled mixture. The mixture was stirred at room temperature for 2 hours, then at 70 °C for 3 hours, and finally stirred overnight at room temperature. The reaction was quenched by adding an aqueous solution of ammonium chloride (30 mL), and the mixture was washed with ethyl acetate (60 mL) to separate the layers. After concentration of the organic layer, the product was purified and separated by silica gel column chromatography (petroleum ether: ethyl acetate = 100:0-75:25) to obtain 7'-fluorospirocyclic [cyclopentane-1,3'-indoline]-2'-one (590 mg, yield 72.42%).

[0530] Lithium aluminum hydride (436.40 mg, 11.50 mmol, 4.0 eq.) was added to a solution of 7'-fluorospirocyclic [cyclopentane-1,3'-indoline]-2'-one (590 mg, 2.87 mmol, 1 eq.) in tetrahydrofuran (20 mL). The mixture was stirred at 70 °C for 2 hours. The mixture was quenched by adding water (0.3 mL) and a 15% sodium hydroxide aqueous solution (0.6 mL). Ethyl acetate (40 mL) was added to the mixture. The solid was removed by filtration, the organic layer was concentrated, and the residue was purified by silica gel column chromatography (elution, PE / EtOAc = 100:0-75:25) to give 7'-fluorospirocyclic [cyclopentane-1,3'-indoline] (300 mg, 54.57%) as a white solid.

[0531] Methyl [(2R,3R,4R,5R)-3,4-diacetoxy-5-(2,6-dichloropurin-9-yl)tetrahydrofuran-2-yl]acetate (350 mg, 782.60 μmol, 1 eq.) was added to a solution of 7'-fluorospirocyclic [cyclopentane-1,3'-indoline] (149.67 mg, 782.60 μmol, 1 eq.) and DIPEA (202.29 mg, 1.57 mmol, 272.62 μL, 2.0 eq.) in 20 mL of NMP. The mixture was stirred overnight at 140 °C. The mixture was diluted with ethyl acetate (30 mL), washed with water, and then washed with brine (30 mL x 3). The organic layer was concentrated, and the residue was purified by silica gel column chromatography (elution, PE / EtOAc = 100:0-60:40) to give methyl [(2R,3R,4R,5R)-3,4-diacetoxy-5-[2-chloro-6-(7'-fluorospirocyclic [cyclopentan-1,3'-indoline]-1'-yl)purin-9-yl]tetrahydrofuran-2-yl]acetate (100 mg, 21.23%) as a brown solid.

[0532] Methyl [(2R,3R,4R,5R)-3,4-diacetoxy-5-[2-chloro-6-(7'-fluorospiro[cyclopentan-1,3'-indoline]-1'-yl)purin-9-yl]tetrahydrofuran-2-yl]acetate (350 mg, 581.39 μmol, 1 eq.) was added to a methanol (3 mL) solution containing ammonia-methanol (7 M, 2.49 mL, 30 eq.). The mixture was stirred overnight at room temperature and then concentrated. The residue was diluted with ethyl acetate (30 mL), washed successively with water and brine, dried with Na2SO4, and concentrated to give (2R,3R,4S,5R)-2-[2-chloro-6-(7'-fluorospirocyclo[cyclopentan-1,3'-indoline]-1'-yl)purin-9-yl]-5-(hydroxymethyl)tetrahydrofuran-3,4-diol (250 mg, 90.36%) as a brown solid.

[0533] To a solution of (2R,3R,4S,5R)-2-[2-chloro-6-(7'-fluorospiro[cyclopentan-1,3'-indoline]-1'-yl)purin-9-yl]-5-(hydroxymethyl)tetrahydrofuran-3,4-diol (270 mg, 567.35 μmol, 1 eq.) in acetone (10 mL), 2,2-dimethoxypropane (886.31 mg, 8.51 mmol, 1.05 mL, 15 eq.) and p-toluenesulfonic acid (97.70 mg, 567.35 μmol, 1 eq.) were added. The mixture was stirred at room temperature for 3 hours. The mixture was diluted with ethyl acetate (50 mL), washed first with an aqueous solution of NaHCO3, and then with brine. The organic layer was concentrated, and the residue was purified by silica gel column chromatography (elution, PE / EtOAc = 100:0-50:50) to give [(3aR,4R,6R,6aR)-4-[2-chloro-6-(7'-fluorospirocyclopentan-1,3'-indoline]-1'-yl)purin-9-yl]-2,2-dimethyl-3a,4,6,6a-tetrahydrofurano[3,4-d][1,3]dioxono-6-yl]methanol (290 mg, yield 99.07%).

[0534] A solution of [(3aR,4R,6R,6aR)-4-[2-chloro-6-(7'-fluorospiro[cyclopentan-1,3'-indoline]-1'-yl)purin-9-yl]-2,2-dimethyl-3a,4,6,6a-tetrahydrofurano[3,4-d][1,3]dioxono-6-yl]methanol (290 mg, 562.06 μmol, 1 eq.) in triethyl phosphate (3 mL) was cooled to 0 °C, and then bis(dichlorophosphoryl)methane (280.79 mg, 1.12 mmol, 2.0 eq.) in 2 mL of trimethyl phosphate was added. The mixture was stirred at room temperature for 4 hours, and then water (3 mL) was added to the mixture. The mixture was stirred at room temperature overnight. The reaction mixture was purified by reversed-phase C-18 silica gel column chromatography (gradient elution, 0 to 30% ACN in water) to give compound d-17 (198 mg, yield 55.7%). 1 H NMR (500MHz, CD3OD) δppm1.81(dd,J=36.0,29.1Hz,8H),2.51(t,J=20.9Hz,2H),4.27(s,1H),4.32(d,J=6.6Hz,1H),4.37(s,1H),4.45(d,J=9. 7Hz,3H),4.68(t,J=5.0Hz,1H),6.07(d,J=4.8Hz,1H),7.01(t,J=9.2Hz,1H),7.09(d,J=6.9Hz,1H),7.15(dd,J=7.8,4.2Hz,1H),8.51(s,1H);13 C NMR(125MHz,CD3OD)δppm 25.94,27.47,39.77,55.20,65.92,68.14,71.50,75.63,84.72,89.77,116.28,1 18.88,121.40,127.55,130.21,141.98,146.35,152.28,153.71,154.31,154.75; 31 P NMR (203MHz, CD3OD) δppm 16.73, 20.18; m / z (ESI + ):634.1(M+H).

[0535] Example 41. Synthesis of compound d-18

[0536]

[0537] Under nitrogen protection and at -78°C, LiHMDS (1M, 21.98mL, 2.2eq.) was added dropwise to a mixture of indolin-2-one (1.33g, 9.99mmol, 1.0eq.) in THF (20mL). The temperature was raised to -50°C and maintained at this temperature for 30 minutes. The mixture was cooled to -78°C, and then 1-bromo-2-(2-bromoethoxy)ethane (2.32g, 9.99mmol, 1eq.) in THF (15mL) was added. The mixture was stirred at room temperature for 2 hours, then under reflux for 2 hours, and finally stirred at room temperature for 16 hours. The mixture was concentrated under reduced pressure, and the residue was partitioned between saturated ammonium chloride and ethyl acetate. The organic layer was concentrated, and the residue was separated by silica gel column chromatography (elution: PE / EtOAc = 100:0-60:40) to give a yellow solid spirocyclic [indoline-3,4'-tetrahydropyran]-2-one (420 mg, yield 20.69%).

[0538] LiAlH4 (153.12 mg, 4.03 mmol, 2.0 eq.) was added to a THF (15 mL) solution of spirocyclic [indoline-3,4'-tetrahydropyran]-2-one (410 mg, 2.02 mmol, 1 eq.). The mixture was stirred overnight at 70 °C. The mixture was quenched with water (0.5 mL) and EtOAc (20 mL) was added. The solid was removed by filtration and the organic layer was concentrated. The residue was purified by silica gel column chromatography (elution: PE / EtOAc = 100:0-75:25) to give spirocyclic [indoline-3,4'-tetrahydropyran] (310 mg, yield 81.20%) as a pink solid.

[0539] To a solution of [(2R,3R,4R,5R)-3,4-diacetoxy-5-(2,-dichloropurin-9-yl)tetrahydrofuran-2-yl]acetate (666 mg, 1.49 mmol, 1 eq.) in 1,4-dioxane (15 mL), spirocyclic [indoline-3,4'-tetrahydropyran] (310.01 mg, 1.1 eq.) and DIPEA (481.15 mg, 3.72 mmol, 2.5 eq.) were added. The mixture was stirred overnight at 100 °C and then concentrated. The residue was purified by silica gel column chromatography (elution: PE / EtOAc = 100:0-10:90) to give methyl [(2R,3R,4R,5R)-3,4-diacetoxy-5-(2-chloro-6-spirocyclic [indoline-3,4'-tetrahydropyran]-1-ylpurine-9-yl)tetrahydrofuran-2-yl]acetate (550 mg, yield 61.55%).

[0540] Ammonia-methanol (7M, 3.27 mL, 25 eq.) was added to a solution of [(2R,3R,4R,5R)-3,4-diacetoxy-5-(2-chloro-6-spiro[cyclopentan-1,3'-indoline]-1'-ylpurine-9-yl)tetrahydrofuran-2-yl]acetate (550 mg, 916.64 μmol, 1 eq.) in methanol (5 mL). The mixture was stirred at room temperature for 5 hours. The solid was obtained by filtration and washed with methanol (15 mL) to give (2R,3R,4S,5R)-2-(2-chloro-6-spiro[indoline-3,4'-tetrahydropyran]-1-yl-purine-9-yl)-5-(hydroxymethyl)tetrahydrofuran-3,4-diol (310 mg, yield 71.36%).

[0541] Add 2,2-dimethoxypropane (1.02 g, 9.81 mmol, 15 eq.) and p-toluenesulfonic acid (112.64 mg, 654.13 μmol, 1 eq.) to a mixture of (2R,3R,3R,4S,4S,5R)-2-(2-chloro-6-spirocyclic [indoline-3,4'-tetrahydropyran]-1-ylpurine-9-yl)-5-(hydroxymethyl)tetrahydrofuran-3,4-diol (310 mg, 654.13 μmol, 1 eq.) in acetone (10 mL). Stir the mixture at room temperature for 2 hours. Evaporate the solvent, dilute the residue with EtOAc (50 mL), wash with saturated sodium bicarbonate solution, and then wash with brine. The organic layer was concentrated and purified by passing it through a silica gel column (elution: PE / EtOAc = 100:0-5:95) to obtain [(3aR,4R,6R,6aR)-4-(2-chloro-6-spirocyclic [indoline-3,4'-tetrahydropyran]-1-ylpurine-9-yl)-2,2-dimethyl-3a,4,6,6a-tetrahydrofurano[3,4-d][1,3]dioxono-6-yl]methanol (252 mg, yield 74.95%).

[0542] A solution of [(3aR,4R,4R,6R,6aR)-4-(2-chloro-6-spirocyclic [indoline-3,4'-tetrahydropyran]-1-ylpurin-9-yl)-2,2-dimethyl-3a,4,4,6,6a-tetrahydrofurano[3,4-d][1,3]dioxono-6-yl]methanol (252 mg, 490.30 μmol, 1 eq.) in trimethyl phosphate (3 mL) was cooled to approximately 0 °C, and then a solution of bis(dichlorophosphoryl)methane (306.17 mg, 1.23 mmol, 2.5 eq.) in trimethyl phosphate (3 mL) was added. The mixture was stirred at room temperature for 5 hours. Water (4 mL) was then added to the reaction mixture, and the mixture was stirred overnight at room temperature. The residual reaction mixture was purified by reverse-phase C-18 silica gel column chromatography (0 to 25% ACN in water) to give compound d-18 (39.0 mg, yield 12.43%). 1HNMR (500MHz, MeOD) δppm 1.61(d,J=13.5Hz,2H),2.04(t,J=13.1Hz,2H),2.53(t,J=21.0Hz,2H),3.73 (dd,J=16.3,8.0Hz,2H),3.98(d,J=10.5Hz,2H),4.29(s,1H),4.31-4.43(m,2 H),4.46(t,J=4.6Hz,1H),4.66-4.75(m,3H),6.06(d,J=4.6Hz,1H),7.10(t,J =7.4Hz,1H),7.24(t,J=7.7Hz,1H),7.29(d,J=7.4Hz,1H),8.45-8.54(m,2H); 31 P NMR (203MHz, MeOD) δppm 16.77, 20.00; 13 C NMR(125MHz,MeOD)δppm 26.14,37.03,42.16,60.27,64.65,70.13,74.32,83.28,88.33,117.70,118.98,122. 34,123.83,127.62,139.74,142.05,151.14,152.05,153.17; m / z(ESI+):632.3(M+H).

[0543] Example 42. Synthesis of compound d-19

[0544]

[0545] A solution of 3-cyanoaniline-1-carboxylic acid tert-butyl ester (3.0 g, 16.46 mmol, 1 eq.) in THF (30 mL) was cooled to -78 °C, and then LiHMDS (1 M, 20.58 mL, 1.25 eq.) was added dropwise at -78 °C. The mixture was stirred at -78 °C for 20 min, and then a solution of 1-(bromomethyl)-2-iodobenzene (5.13 g, 17.29 mmol, 1.05 eq.) in THF (3 mL) was added. The mixture was stirred at -78 °C for 3 h, and then quenched with saturated ammonium chloride. The mixture was extracted with ethyl acetate (50 mL * 2). The organic layer was washed with brine and concentrated. The residue was purified by column chromatography (elution, PE / EA = 100:0-83:17) to give tert-butyl 3-cyano-3-[(2-iodophenyl)methyl]azacyclobutane-1-carboxylic acid (6.44 g, 16.17 mmol, yield 98.22%) as a yellow oil.

[0546] 3-Cyano-3-[(2-iodophenyl)methyl]azacyclobutane-1-carboxylic acid tert-butyl ester (6.44 g, 16.17 mmol, 1 eq.) was cooled to -78 °C in 60 mL of THF solution, and then n-BuLi (2.5 M, 12.94 mL, 2 eq.) was added dropwise at -78 °C. The mixture was stirred at -78 °C for 2 h, and the reaction was quenched with saturated ammonium chloride. The mixture was extracted with ethyl acetate (75 mL * 2). The organic layer was washed with brine and concentrated. The residue was purified by column chromatography (elution, PE / EA = 100:0-83:17) to give 1'-oxospirocyclic [azacyclobutane-3,2'-indene]-1-carboxylic acid tert-butyl ester (3.4 g, 12.44 mmol, yield 76.92%) as a pale yellow solid.

[0547] 1'-oxospirocyclic [azacyclobutane-3,2'-indene]-1-carboxylic acid tert-butyl ester (350 mg, 1.28 mmol, 1 eq.) was dissolved in HCl-EA (4 mL). The mixture was stirred at room temperature for 3 hours, concentrated, and used directly in the next step.

[0548] Methyl [(2R,3R,4R,5R)-3,4-diacetoxy-5-(2,6-dichloropurin-9-yl)tetrahydrofuran-2-yl]acetate (408.31 mg, 912.99 μmol, 1 eq.) was added to 20 mL of a mixture of 1,4-dioxanes, along with spirocyclic [azacyclobutane-1-onthium-3,2'-indene]-1'-one chloride (268 mg, 1.28 mmol, 1.4 eq.) and DIPEA (412.98 mg, 3.20 mmol, 556.58 μL, 3.5 eq.). The mixture was stirred at 100 °C for 4 hours. The solvent was removed by evaporation, and the residue was diluted with ethyl acetate (40 mL). The residue was then washed with water and brine, and the organic phase was concentrated. The residue was purified by column chromatography (elution, PE / EA = 100:0-50:50) to give methyl [(2R,3R,4R,5R)-3,4-diacetoxy-5-[2-chloro-6-(1'-oxospirocyclic [azacyclobutane-3,2'-indene]-1-yl)purin-9-yl)tetrahydrofuran-2-yl]acetate (440 mg, 753.46 μmol, yield 82.53%) as a white solid.

[0549] To a mixture of methyl [(2R,3R,4R,5R)-3,4-diacetoxy-5-[2-chloro-6-(1'-oxospirocyclic [azacyclobutane-3,2'-indene]-1-yl)purin-9-yl)tetrahydrofuran-2-yl]acetate (337 mg, 577.08 μmol, 1 eq.) in 5.00 mL of methanol, NH3-MeOH (7 M, 2.47 mL, 30 eq.) was added. The mixture was stirred at room temperature for 3 hours to give a crude product, which was extracted with ethyl acetate (50 mL x 2) and washed with 50 mL of water. The organic layer was washed with brine, dried with sodium sulfate, and concentrated to give 1-[2-chloro-9-[(2R,3R,4S,5R)-3,4-dihydroxy-5-(hydroxymethyl)tetrahydrofuran-2-yl]purin-6-yl]spirocyclic [azacyclobutane-3,2'-indene]-1'-one (264 mg, 576.59 μmol, yield 99.91%).

[0550] At 0 °C, 2,2-dimethylpropane (1.36 g, 13.10 mmol, 1.61 mL, 20 eq.) and p-TsOH□H₂O (125.80 mg, 655.21 μmol, 1 eq.) were added to a solution of 1-[2-chloro-9-[(2R,3R,4S,5R)-3,4-dihydroxy-5-(hydroxymethyl)tetrahydrofuran-2-yl]purin-6-yl]spirocyclic[azacyclobutane-3,2'-indene]-1'-one (300 mg, 655.21 μmol, 1 eq.) in 30 mL of acetone. The mixture was stirred at room temperature for 3 hours, followed by removal of the solvent. The residue was diluted with EA (50 mL), then washed first with an aqueous solution of sodium bicarbonate (2 x 50 mL), followed by washing with 50 mL. The organic layer was concentrated. The residue was purified by column chromatography (elution, PE / EA = 100:0-60:40) to give 1-[9-[(3aR,4R,6R,6aR)-6-(hydroxymethyl)-2,2-dimethyl-3a,4,6,6a-tetrahydrofurano[3,4-d][1,3]dioxono-4-yl]-2-chloro-purin-6-yl]-spirocyclic[azacyclobutane-3,2'-indene]-1'-one (270 mg, yield 82.76%) as a white solid.

[0551] A solution of 1-[9-[(3aR,4R,6R,6aR)-6-(hydroxymethyl)-2,2-dimethyl-3a,4,6,6a-tetrahydrofurano[3,4-d][1,3]dioxono-4-yl]-2-chloro-purine-6-yl]-spirocyclic [azacyclobutane-3,2'-indene]-1'-one (270 mg, 542.25 μmol, 1 eq.) in PO(MeO)3 (4 mL) was cooled to 0 °C, and then bis(dichlorophosphoryl)methane (270.89 mg, 1.08 mmol, 2 eq.) in PO(MeO)3 (4 mL) was added. The mixture was stirred at 0 °C for 5 hours. LC-MS analysis showed that the reaction did not proceed in large quantities. Add bis(dichlorophosphoryl)methane (135.4 mg, 0.54 mmol, 1 eq.) to the mixture in PO(MeO)3 (2 mL). Stir the mixture overnight at room temperature. LC-MS analysis showed that approximately 50% of the feed remained. The mixture was used directly for the next step.

[0552] Water (7 mL) was added to the above mixture at 0 °C. The mixture was stirred overnight at room temperature. The mixture was purified by preparative HPLC. ¹H NMR analysis showed impurities in the crude product (containing PO(OMe)₃). The crude product was purified by C-18 reversed-phase silica column chromatography (0–25% ACN in water) to give compound d-19 (60.6 mg, 98.11 μmol, yield 22.38%, purity 99.71%). 1 H NMR(500MHz,MeOD)δppm 2.52(t,J=20.9Hz,2H),3.68(s,2H),4.25-4.43(m,4H),4.47(t,J=4.7Hz,2H),4.74(d,J=68.8Hz,3H),6.04(d,J=4 .7Hz,1H),7.49(t,J=7.5Hz,1H),7.61(d,J=7.7Hz,1H),7.74(t,J=7.5Hz,1H),7.80(d,J=7.6Hz,1H),8.44(s,1H); 31 P NMR (203MHz, MeOD) δppm 16.84, 19.73; 13 C NMR(125MHz,MeOD)δppm15.90,26.14,38.96,46.25,56.11,58.97,60.87,64.58,70.07,74.31,83.31,88.34,117 .89,123.72,126.42,127.67,134.91,135.65,140.10,150.63,152.95,154.15,206.67; m / z(ESI+):616.3(M+H).

[0553] Example 43. Synthesis of compound d-20

[0554]

[0555] Under a nitrogen atmosphere and at -78°C, LiHMDS (1M, in THF, 16.52 mL, 2.2 eq.) was added dropwise to a mixture of indoline-2-one (1 g, 7.51 mmol, 1 eq.) in 8 mL of THF. The reaction temperature was raised to -50°C and maintained at this temperature for 30 min. The mixture was then cooled again to -78°C, followed by the dropwise addition of 1,3-dibromopropane (1.52 g, 7.51 mmol, 1 eq.) in 8 mL of THF. The mixture was stirred at room temperature for 1 h, and then stirred under reflux for 3 h. The mixture was concentrated under reduced pressure, and the residue was partitioned between ethyl acetate and saturated ammonium chloride. The organic layer was concentrated, and the residue was purified by silica gel column chromatography (elution, PE / EA = 100:0–85:15) to give spirocyclic [cyclobutane-1,3'-indoline]-2'-one (345 mg, 1.99 mmol, yield 26.52%).

[0556] LiAlH4 (151.18 mg, 3.98 mmol, 2 eq.) was added to a solution of spirocyclic [cyclobutane-1,3'-indoline]-2'-one (345 mg, 1.99 mmol, 1 eq.) in 20 mL of THF. The mixture was stirred at 70 °C for 4 h. The reaction was quenched at 0 °C with 15% NaOH (aq., 5 mL), followed by the addition of EtOAc (20 mL). The organic layer was dried over magnesium sulfate. After removing the solid by filtration, the filtrate was concentrated to give spirocyclic [cyclobutane-1,3'-indoline] (317 mg, 1.99 mmol, yield 99.95%).

[0557] To a solution of [(2R,3R,4R,5R)3,4-diacetoxy-5-(2,6-dichloropurin-9-yl)tetrahydrofuran-2-yl]acetate (593.58 mg, 1.33 mmol, 1 eq.) in 1,4-dioxane (30 mL), spirocyclic [cyclobutane-1,3'-indoline] (317 mg, 1.99 mmol, 1.5 eq.) and DIPEA (428.83 mg, 3.32 mmol, 577.94 μL, 2.5 eq.) were added. The mixture was stirred at 100 °C for 5 hours. The solvent was removed by evaporation. The residue was diluted with ethyl acetate (50 mL), washed with water, and then washed with brine. The organic phase was concentrated, and the residue was purified by column chromatography (elution, PE / EA = 100:0-65:35) to give methyl [(2R,3R,4R,5R)-3,4-diacetoxy-5-[2-chloro-6-spirocyclic [cyclobutane-1,3'-indoline]-1'-yl-purine-9-yl)tetrahydrofuran-2-yl]acetate (586 mg, 1.03 mmol, yield 77.46%) as a pale yellow solid.

[0558] Methyl [(2R,3R,4R,5R)-3,4-diacetoxy-5-[2-chloro-6-spirocyclic [cyclobutane-1,3'-indoline]-1'-yl-purine-9-yl)tetrahydrofuran-2-yl]acetate (586 mg, 1.03 mmol, 1 eq.) in methanol (5 mL) was added to NH3-MeOH (7 M, 4.41 mL, 30 eq.). The mixture was stirred at room temperature for 3 hours. LC-MS analysis showed that the starting material was consumed, the molecular ion of the product was detected, and a certain amount of intermediate was present. NH3-MeOH (2 mL) was added to the mixture, and the mixture was stirred overnight at room temperature. The solvent was removed by evaporation, and the residue was diluted with ethyl acetate (2 × 30 mL) and water (30 mL). The organic layer was washed with brine, dried with Na2SO4, and concentrated to give (2R,3R,4S,5R)-2-(2-chloro-6-(spirocyclic [cyclobutane-1,3'-indoline]-1'-yl-purine-9-yl)-5-(hydroxymethyl)tetrahydrofuran-3,4-diol (450 mg, 1.01 mmol, yield 98.61%) as a yellow solid.

[0559] At 0 °C, 2,2-dimethylpropane (2.11 g, 20.28 mmol, 2.49 mL, 20 eq.) and p-TsOH□H2O (193.92 mg, 1.01 mmol, 1 eq.) were added to a solution of (2R,3R,4S,5R)-2-(2-chloro-6-(spirocyclobutane-1,3'-indole]-1'-yl)-9H-purine-9-yl)-5-(hydroxymethyl)tetrahydrofuran-3,4-diol (450 mg, 1.01 mmol, 1 eq.) in 30 mL of acetone. The mixture was stirred at room temperature for 3 hours. The solvent was removed by evaporation, the residue was diluted with EA (50 mL), washed first with NaHCO3 (aq, 2 x 50 mL), then with brine (50 mL), and concentrated. The residue was purified by column chromatography (elution, PE / EA = 100:0-60:40) to give a pale yellow solid of [(3aR,4R,6R,6aR)-4-(2-chloro-6-spirocyclic [cyclobutane-1,3'-indoline]-1'-yl)-purine-9-yl)-2,2-dimethyl-3a,4,6,6a-tetrahydrofurano[3,4-d][1,3]dioxono-6-yl)methanol (370 mg, 764.55 μmol, yield 75.42%).

[0560] A solution of [(3aR,4R,6R,6aR)-4-(2-chloro-6-spirocyclo[cyclobutane-1,3'-indoline]-1'-yl)-purine-9-yl)-2,2-dimethyl-3a,4,6,6a-tetrahydrofurano[3,4-d][1,3]dioxono-6-yl)methanol (370 mg, 764.55 μmol, 1 eq.) in trimethyl phosphate (4 mL) was cooled to 0 °C, and then bis(dichlorophosphoryl)methane (381.95 mg, 1.53 mmol, 2 eq.) in trimethyl phosphate (4 mL) was added. The mixture was stirred at 0 °C for 5 h. LC-MS showed that approximately 50% of the starting material remained. Bis(dichlorophosphoryl)methane (191 mg, 0.765 mmol, 1 eq.) in trimethyl phosphate (2 mL) was added at 0 °C, and the reaction was continued by stirring the mixture at room temperature for 2 hours. At this point, LC-MS showed that the starting material had disappeared. 7 mL of water was added dropwise to the mixture at 0 °C, and the mixture was stirred at 40 °C for 40 minutes, then stirred overnight at room temperature. The reaction mixture was purified by C-18 reversed-phase silica gel (0 to 25% ACN in water) to give compound d-20 (106 mg, 176.12 μmol, yield 23.04%, purity 98.33%). 1H NMR(500MHz,MeOD)δppm 2.10-2.22(m,2H),2.38(dd,J=15.8,9.9Hz,2H),2.48(d,J=8.5Hz,2H),2.56(t,J= 20.9Hz,2H),4.32(s,1H),4.34-4.39(m,1H),4.40-4.46(m,1H),4.49(t,J=4.8Hz,1 H),4.70(t,J=4.8Hz,1H),4.84(s,2H),6.08(d,J=4.6Hz,1H),7.14(t,J=7.4Hz,1H ),7.23(t,J=7.8Hz,1H),7.54(d,J=7.4Hz,1H),8.46(d,J=8.2Hz,1H),8.48(s,1H); 31 P NMR (203MHz, MeOD) δppm 16.80, 19.93; 13 C NMR(125MHz,MeOD)δ15.43,26.15,35.69,46.27,64.66,70.07,74.33,83.21,88.34,117.15,118.93 ,122.13,124.01,127.29,139.54,139.80,141.79,150.91,151.93,153.18; m / z(ESI+):602.2(M+H).

[0561] Example 44. Synthesis of compound d-21

[0562]

[0563] A solution of tert-butyl 4-cyanopiperidine-1-carboxylate (2.0 g, 9.51 mmol, 1 eq.) in THF (30 mL) was cooled to -78 °C, followed by dropwise addition of LiHMDS (1 M, 11.89 mL, 1.25 eq.) at -78 °C with stirring for 20 min. Then, a solution of 1-(bromomethyl)-2-iodobenzene (3.11 g, 10.46 mmol, 1.1 eq.) in THF (5 mL) was added dropwise to the mixture. The mixture was stirred at -78 °C for 3 h. The reaction was quenched with saturated ammonium chloride and extracted with ethyl acetate (50 mL * 2). The organic layer was washed with brine and concentrated. The residue was purified by silica gel column chromatography (elution, PE / EA = 100:0–83:17) to give tert-butyl 4-cyano-4-[(2-iodophenyl)methyl]piperidine-1-carboxylate (3.7 g, yield 91.25%).

[0564] A solution of 4-cyano-4-[(2-iodophenyl)methyl]piperidine-1-carboxylic acid tert-butyl ester (3.7 g, 8.68 mmol, 1 eq.) in THF (30 mL) was cooled to approximately -78 °C, followed by dropwise addition of n-butyllithium (2.5 M, 6.94 mL, 2 eq.) at approximately -78 °C. The mixture was stirred at this temperature for 2 hours, and the reaction mixture was quenched with saturated ammonium chloride. The mixture was extracted with EA (80 mL x 2). The organic layer was washed with brine and concentrated. The residue was purified by silica gel column chromatography (elution, PE / EA petroleum ether: ethyl acetate = 100:0-75:25) to give 1-oxopyrrole[indene-2,4'-piperidine]-1'-carboxylic acid tert-butyl ester (1.55 g, 5.14 mmol, yield 59.25%).

[0565] At 0 °C, sodium borohydride (486.40 mg, 12.86 mmol, 2.5 eq.) was added in portions to a solution of 1-oxopyrrole[indene-2,4'-piperidine]-1'-carboxylic acid tert-butyl ester (1.55 g, 5.14 mmol, 1 eq.) in methanol (20 mL). The mixture was stirred at this temperature for 2 hours. The solvent was removed by evaporation, and the residue was extracted with ethyl acetate (100 mL * 2). The organic layer was washed with brine (100 mL) and concentrated to give 1-hydroxyspirocyclic[indene-2,4'-piperidine]-1'-carboxylic acid tert-butyl ester (1.54 g, 5.08 mmol, yield 98.69%).

[0566] Pd / C (200 mg, 1.65 mmol) was added to a solution of 1-hydroxyspirocyclic [indene-2,4'-piperidine]-1'-carboxylic acid tert-butyl ester (950 mg, 3.13 mmol, 1 eq.) in CH3OH / CH3COOH = 1:4 (40 mL). The mixture was stirred overnight at room temperature under a hydrogen atmosphere. Insoluble matter was removed by filtration and washed with methanol. The filtrate and washings were combined, concentrated by evaporation, neutralized with saturated sodium bicarbonate solution, and extracted with DCM (80 mL * 2). The organic layer was washed with brine and concentrated. The residue was purified by silica gel column chromatography (elution, PE / EA = 100:0–90:10) to give spirocyclic [indene-2,4'-piperidine]-1'-carboxylic acid tert-butyl ester (190 mg, yield 21.11%).

[0567] Spirocyclic [indene-2,4'-piperidine]-1'-carboxylic acid tert-butyl ester (330 mg, 1.15 mmol, 1 eq.) was dissolved in dioxane hydrochloride (5 mL). After stirring at room temperature for 3 h, the mixture was concentrated, and the residue was used directly for the next step.

[0568] To a solution of methyl [(2R,3R,4R,5R)-3,4-diacetoxy-5-(2,6-dichloropurin-9-yl)tetrahydrofuran-2-yl]acetate (466.8 mg, 1.04 mmol, 1 eq.) in 1,4-dioxane (10 mL), spirocyclic [indene-2,4'-piperidine-1-onium] chloride (257 mg, 1.15 mmol, 1.10 eq.) and DIPEA (472.14 mg, 3.65 mmol, 636.30 μL, 3.5 eq.) were added. The mixture was stirred at 100 °C for 4 hours and concentrated by evaporation. The residue was purified by silica gel column chr...

Claims

1. A compound of formula IV or a pharmaceutically acceptable salt thereof: (IV) in, X is selected from halogens; p and q are independently selected from integers from 0 to 3, provided that p and q are not both 0 at the same time; r is 1; and R 7 R 8 R 9 and R 10 It is independently selected from hydrogen.

2. A compound of formula V or a pharmaceutically acceptable salt thereof: （V） in, r and s are independently selected from integers from 0 to 2, provided that r and s are not both 0 at the same time; p and q are independently selected from integers from 0 to 3, provided that p and q are not both 0 at the same time; R 5 and R 6 Independently selected from hydrogen; R 7 and R 10 Independently selected from H; and X is selected from halogens.

3. Selected from the following compounds: ， Or its pharmaceutically acceptable salt.

4. A pharmaceutical composition comprising the compound of any one of claims 1 to 3 or a pharmaceutically acceptable salt thereof, and a pharmaceutically acceptable carrier.

5. The pharmaceutical composition according to claim 4, wherein, Pharmaceutically acceptable carriers include emulsions, gels, liposomes, or nanoparticles.

6. The pharmaceutical composition according to claim 4, wherein, Pharmaceutically acceptable carriers include creams.

7. The pharmaceutical composition according to claim 4 or 5, wherein, The composition is suitable for oral administration.

8. The pharmaceutical composition according to claim 4 or 5, wherein, The composition is in the form of hard-shell gelatin capsules, soft-shell gelatin capsules, pills, tablets, lozenges, powders, or granules.

9. The pharmaceutical composition according to claim 4 or 5, wherein, The composition is in the form of a capsule, pellet, flavoring tablet, or sugar-coated pill.

10. The pharmaceutical composition according to claim 4 or 5, wherein, The composition is in the form of a solution, an aqueous liquid suspension, a non-aqueous liquid suspension, an oil-in-water liquid emulsion, an oil-in-water liquid emulsion, an elixir, or a syrup.

11. The pharmaceutical composition according to claim 4 or 5, wherein, The composition has an enteric coating.

12. The pharmaceutical composition according to claim 4 or 5, wherein, The composition is formulated for controlled release.

13. The pharmaceutical composition according to claim 4 or 5, wherein, The composition is injectable.

14. The pharmaceutical composition according to claim 4 or 5 further comprises at least one additional therapeutic agent.

15. The pharmaceutical composition according to claim 14, wherein, The at least one additional therapeutic agent is a chemotherapeutic agent, an immunomodulatory and / or inflammatory modulator, an anti-hypercholesterol agent, or an anti-infective agent.

16. The pharmaceutical composition according to claim 14, wherein, The aforementioned at least one additional therapeutic agent is an immune checkpoint inhibitor.

17. A kit comprising a compound according to any one of claims 1 to 3 or a pharmaceutically acceptable salt thereof, and at least one additional therapeutic agent.

18. The kit according to claim 17, wherein, The at least one additional therapeutic agent is a chemotherapeutic agent, an immunomodulatory and / or inflammatory modulator, an anti-hypercholesterol agent, or an anti-infective agent.

19. The kit according to claim 17, wherein, The at least one additional therapeutic agent is an immune checkpoint inhibitor.

20. The kit according to any one of claims 17 to 19, further comprising an excipient and / or instructions for use thereof.

21. The kit according to any one of claims 17 to 19, further comprising a buffer and / or instructions for use thereof.

22. The kit according to claim 19, wherein, The immune checkpoint inhibitors are selected from a group consisting of ipilimumab, nivolumab, and pembrolizumab.

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