Xanthine compounds and their uses

Abstract

The xanthine compounds and their uses have the structure shown in Formula Ia, where the definitions of each substituent are as described in the present specification and claims. The compounds of Formula Ia are inhibitors or agonists of TRPC4 and TRPC5 ion channels, and have potential application value in the prevention, delay, or treatment of diseases associated with abnormal TRPC4 / 5 function or expression. [Formula 1] TIFF2026507486000102.tif43170

Description

Detailed Description of the Invention

[0001] [Technical field] The present invention relates to the field of medicinal chemistry, specifically to xanthine compounds, their preparation methods and uses, which are inhibitors or agonists of TRPC4 and TRPC5 ion channels and have potential application value in the prevention, delay or treatment of diseases associated with abnormal TRPC4 / 5 function or expression.

[0002] [Background technology] Transient receptor potential (TRP) ion channels are a type of six-transmembrane channel protein widely distributed in the peripheral and central nervous systems. They are molecular sensory receptors that sense changes in the cellular environment, transmit signals, and maintain cellular homeostasis. Based on sequence similarity, mammalian TRP ion channels can be classified into six subfamilies: TRPA, TRPC, TRPM, TRPML, TRPP, and TRPV. Classical transient receptor potential (TRPC) channels are nonselective cation channels, transducing Ca. 2+ , Na + Mammalian TRPCs consist of six members: TRPC1, TRPC3, TRPC4, TRPC5, TRPC6, and TRPC7, where TRPC3, TRPC6, and TRPC7 share high amino acid sequence homology, and TRPC1, TRPC4, and TRPC5 share high sequence homology. TRPC family proteins exist as homozygous or heterozygous tetramers and function as ion channels, with TRPC5 forming heterozygous tetramers with TRPC1 and TRPC4.

[0003] TRPC4 / 5 are expressed in multiple brain regions, including the hippocampus, amygdala, and frontal cortex, and are involved in the development of neurological disorders such as anxiety, depression, epilepsy, and addiction. Knocking out the TRPC4 or TRPC5 gene or administering a TRPC4 / 5 inhibitor can alleviate symptoms of anxiety and depression in mice. The TRPC4 / 5 inhibitor BI-1358894, developed by Boehringer-Ingelheim Pharmaceuticals, is currently undergoing clinical trials as a treatment for psychiatric disorders such as depression, borderline personality disorder, and post-traumatic stress disorder.

[0004] Podocytes are specialized cells that form the glomerular filtration barrier. TRPC5 is also expressed in glomerular podocytes, and its dysfunction is associated with the development of chronic kidney disease. TRPC5 is involved in regulating angiotensin-stimulated cell migration and actin remodeling processes. In various animal models of kidney disease, TRPC5 gene knockout or TPRC5 inhibitor significantly reduced proteinuria levels, protected podocytes from damage, and inhibited the progression of kidney disease. The TRPC4 / 5 inhibitor GFB-887, developed by GoldfinchBoitechnology, is currently undergoing clinical trials for the treatment of kidney diseases such as diabetic nephropathy and focal segmental glomerulosclerosis.

[0005] TRPC4 / 5 are expressed in skin keratinocytes and play an important role in the process of regulating keratinocyte differentiation. Therefore, targeting and regulating TRPC4 / 5 may be a potential therapeutic strategy for treating the pathophysiological basis of keratinocyte dysfunction.

[0006] TRPC5 is expressed in the liver and portal vein bed, and endogenous phospholipids such as lysolecithin (LPC), which plays an important role in liver stasis syndrome, can significantly activate TRPC5. In a mouse model of bile acid-induced cholestatic liver disease, TRPC5 gene knockout alleviates liver injury and hepatic lipid metabolism abnormalities. This suggests that TRPC5 may also be a potential therapeutic target for liver disease.

[0007] TRPC4 / 5 is also expressed in peripheral sensory neurons and is involved in pain perception. Compared with wild-type animals, TRPC4- / - mice exhibit resistance to mustard oil-induced pain and an increased pain threshold. In various mouse models of neuropathic pain, inflammatory pain, and spontaneous pain, TRPC5 gene knockout or TRPC5 inhibitors can effectively alleviate mechanical allodynia, an effect essentially related to biological functions regulated by TRPC5.

[0008] TRPC4 / 5 is also involved in cardiovascular disease. Activation of the TRPC5 channel significantly promotes vascular smooth muscle cell proliferation and migration. Inhibition of TRPC5 significantly attenuates hypoxia-ischemia-induced cell apoptosis and oxidative stress in endothelial cells, reduces endothelium-dependent vasoconstriction, and inhibits atherosclerotic plaque formation. It also exerts anti-atherosclerotic, anti-cardiac hypertrophy, and anti-arrhythmic effects by reducing the expression of cardiac hypertrophy-related genes and maintaining the stability of cardiac electrical activity. Furthermore, TRPC5-mediated calcium ion influx is an important signaling mechanism released by various endothelium-derived contractile factors and contributes to the development of hypertension. TRPC5 promotes vascular smooth muscle cell proliferation and increases oxidative stress, thereby promoting the development of hypertension.

[0009] TRPC4 / 5 is involved in tumor angiogenesis, tumor resistance, and metastasis. TRPC5 is highly expressed in colorectal cancer and activates the Wnt5a / β-catenin pathway by promoting extracellular calcium influx, thereby reducing tumor differentiation and increasing tumor stem cells. TRPC5 expression levels are negatively correlated with the prognosis of colorectal cancer patients. TRPC5 can also promote tumor cell migration, invasion, and proliferation by reducing E-cadherin and increasing the expression of mesenchymal markers. Furthermore, calcium ion dysregulation caused by TRPC5 activation promotes anaerobe-inducible factor 1 (HIF-1)-mediated tumor angiogenesis, allowing cancer cells to escape from high-dose anticancer drug exposure sites and obtain additional nutrients that support tumor survival, thereby reducing the efficacy of chemotherapy. In breast cancer models, inhibition of TRPC5 can reduce chemotherapy resistance. However, activation of TRPC4 / 5 can also induce tumor cell death through calcium ion excess. For example, the natural product (-)-Englerin A is a potent TRPC4 / 5 agonist, which exhibits antiproliferative effects on various tumor cells, including those of kidney, lung, breast, and skin cancers.

[0010] TRPC4 / 5 are expressed in the small intestinal smooth muscle and myenteric plexus and are involved in the regulation of intestinal smooth muscle cell contraction, gastrointestinal motility, and enteric nerve signaling. Therefore, compounds that target the activity of TRPC4 / 5 channels have potential application in the treatment of intestinal diseases.

[0011] WO2014143799 discloses xanthine compounds as shown in the following general formula, in which the 8-position substituent R 2is defined as a C1-C6 alkyl group, a C1-C6 heteroalkyl group, a C1-C6 alkoxy group, a C3-C7 cycloalkoxy group, a C6-C10 aryl group, a C6-C10 aryloxy group, a C7-C16 arylalkoxy group, a heterocycloalkyl group, a heteroaryl group, a heteroaryloxy group, etc. Compound 260 in this application, the compound primarily studied, exhibits good TRPC5 inhibitory activity and also shows good efficacy in mouse models of depression and anxiety. In another comparative article (PLoS ONE. 2018, 13, e0191225.), compound 260 is also called HC-070, and this article details HC-070's inhibitory activity against TRPC4 / 5 channels and its anxiolytic and antidepressant effects in various animal models. However, HC-070 lacks selectivity for some other ion channels, and it can significantly inhibit the hERG potassium channel and TRPC3.

[0012] [ka]

[0013] WO2019011802 discloses compounds in which the 8-position of the xanthine is substituted with a "3-pyridyl group" or a "2-piperazinyl group" as shown below, which, compared to HC-070, have a reduced inhibitory effect on the hERG channel while maintaining high TRPC5 inhibitory activity.

[0014] [ka]

[0015] [Summary of the Invention] [Problem to be solved by the invention] The primary object of the present invention is to provide xanthine compounds useful as regulators of TRPC4 and TRPC5, their preparation methods and therapeutic uses.

[0016] [Means for solving the problem] In a first aspect, the present invention provides a fused pyrimidinedione compound represented by general formula (I), its stereoisomers and pharmaceutically acceptable salts thereof,

[0017] [ka]

[0018] wherein ring A is a saturated or partially unsaturated C3-C10 cyclic hydrocarbon group, including monocyclic, fused, spirocyclic, and bridged ring systems; X is N or CH; and R 1 is a C1-C6 alkyl group or a C1-C6 halogenated alkyl group, n is an integer of 1 to 4, and each R 2 are each independently H, deuterium, a halogen atom, a C1-C6 alkyl group, a C1-C6 halogenated alkyl group, a C1-C6 alkoxy group, a C1-C6 halogenated alkoxy group, or a cyano group; m is an integer of 1 to 6; and each R 3 are each independently H, deuterium, a C1-C6 alkyl group, a C1-C6 halogenated alkyl group, a C1-C6 alkoxy group, a C1-C6 halogenated alkoxy group, a C3-C8 cycloalkyl group, a C6-C10 aryl group, a 5- to 8-membered heteroaryl group, a 4- to 8-membered heterocyclic group, a halogen atom, a cyano group, a hydroxy group, -NR 4 R 5 or R 3 together with the carbon attached to ring A, form a 4-8 membered heterocyclic or C6-C10 aryl group, wherein the cycloalkyl group, aryl group, heteroaryl group, and heterocyclic group are unsubstituted or optionally substituted with 1 to 4 groups selected from the group consisting of a C1-C6 alkyl group, a C1-C6 alkoxy group, a halogen atom, and a cyano group; R 4 , R 5 are each independently H, a C1-C6 alkyl group, or a C3-C8 cycloalkyl group.

[0019] In a second aspect, the present invention further provides a xanthine-based compound represented by general formula (Ia), its stereoisomers and pharmaceutically acceptable salts thereof,

[0020] [ka]

[0021] wherein Ring A is a saturated or partially unsaturated C3-C10 cyclic hydrocarbon group, preferably including monocyclic, fused ring, spirocyclic and bridged ring systems; R 1 is a C1-C6 alkyl group or a C1-C6 halogenated alkyl group, n is an integer from 1 to 4, and each R 2 are each independently H, deuterium, halogen, a C1-C6 alkyl group, a C1-C6 halogenated alkyl group, a C1-C6 alkoxy group, a C1-C6 halogenated alkoxy group, or a cyano group; m is an integer from 1 to 6, and each R 3 are each independently H, deuterium, a C1-C6 alkyl group, a C1-C6 halogenated alkyl group, a C1-C6 alkoxy group, a C1-C6 halogenated alkoxy group, a C3-C8 cycloalkyl group, a C6-C10 aryl group, a 5- to 8-membered heteroaryl group, a 4- to 8-membered heterocyclic group, a halogen atom, a cyano group, a hydroxy group, -NR 4 R 5 or R 3 together with the carbon bonded to ring A, form a 4-8 membered heterocyclic or C6-C10 aryl group, and the cycloalkyl group, aryl group, heteroaryl group, or heterocyclic group is unsubstituted or optionally substituted with 1 to 4 groups selected from the group consisting of a C1-C6 alkyl group, a C1-C6 halogenated alkyl group, a C1-C6 alkoxy group, a C1-C6 halogenated alkoxy group, a halogen, and a cyano group; R 4 , R 5 are each independently H, a C1-C6 alkyl group, or a C3-C8 cycloalkyl group.

[0022] In another preferred example, ring A is a saturated or partially unsaturated C3-C8 monocyclic or bridged cyclic hydrocarbon group. In another preferred embodiment, the A ring is

[0023] [ka] is selected from the group consisting of:

[0024] In another preferred example, the A ring is a cyclohexyl group (

[0025] [ka] ) or a cyclohexenyl group (e.g.,

[0026] [ka] )

[0027] In another preferred embodiment, the compound has a structure as shown in general formula (Ib):

[0028] [ka]

[0029] where R 1 is a C1-C6 alkyl group, n is an integer from 1 to 4, and each R 2 are each independently H, CN, a C1-C6 halogenated alkyl group, or a halogen atom; Each R 3 are each independently H, deuterium, a C1-C6 alkyl group, a C1-C6 halogenated alkyl group, a C1-C6 alkoxy group, a C1-C6 halogenated alkoxy group, a C6-C10 aryl group, a C3-C8 cycloalkyl group, a 5- to 8-membered heteroaryl group, a 4- to 8-membered heterocyclic group, a halogen atom, a cyano group, a hydroxy group, -NR 4 R 5or two R 3 forms a 4- to 8-membered heterocyclic ring together with the carbon attached to ring A, R 4 , R 5 are each independently H or a C1-C6 alkyl group.

[0030] In another preferred embodiment, R 1 is a C1-C4 alkyl group. In another preferred example, R 1 is a methyl group, an ethyl group, or an isopropyl group. In another preferred example, n is 1, 2, or 3, and each R 2 are each independently H, deuterium, halogen, a C1-C4 alkyl group, a C1-C4 halogenated alkyl group, a C1-C4 alkoxy group, a C1-C4 halogenated alkoxy group, or a cyano group. In another preferred example, n is 1, 2, or 3, and each R 2 are each independently H, deuterium, F, Cl, Br, a methyl group, an ethyl group, or an isopropyl group. In another preferred example, n is 1 and R 2 is F, Cl, or Br located in the para position. In another preferred embodiment, n is 2 and each R 2 are independently F, Cl, or Br located in the para and meta positions In another preferred embodiment, R 2 is -F or -Cl.

[0031] In another preferred example, m is 1, 2, 3, or 4, and each R 3 are each independently H, deuterium, a C1-C4 alkyl group, a C1-C4 halogenated alkyl group, a C1-C4 alkoxy group, a C1-C4 halogenated alkoxy group, a C3-C6 cycloalkyl group, a phenyl group, a 5- to 6-membered heteroaryl group, a 5- to 6-membered heterocyclic group, F, Cl, Br, a cyano group, a hydroxy group, -NR 4 R 5 or R 3together with the carbon bonded to ring A, form a 5- to 6-membered heterocyclic or phenyl group, and the cycloalkyl group, phenyl group, heteroaryl group, and heterocyclic group are unsubstituted or optionally substituted with 1 to 4 groups selected from the group consisting of a C1-C6 alkyl group, a C1-C6 halogenated alkyl group, a C1-C6 alkoxy group, a C1-C6 halogenated alkoxy group, a halogen, and a cyano group.

[0032] In another preferred embodiment, each R 3 are each independently selected from H, a methyl group, an ethyl group, an n-propyl group, an isopropyl group, a trifluoromethyl group, a difluoromethyl group, a monofluoromethyl group, a phenyl group, a cyclobutyl group, a cyclopropyl group, a cyclopentyl group, a cyclohexyl group, a methoxy group, an ethoxy group, an n-propoxy group, an isopropoxy group, a hydroxy group, F, Cl, Br, a cyano group, an amino group, a methylamino group, a dimethylamino group, an oxetanyl group, a dioxolanyl group, a morpholinyl group, a piperazinyl group, a pyrrolidinyl group, and a piperidinyl group. In another preferred embodiment, each R 3 are each independently H, a C1-C6 alkyl group, a C1-C6 halogenated alkyl group, a C1-C6 alkoxy group, a C3-C8 cycloalkyl group, a phenyl group, a 4- to 8-membered heterocyclic group, a halogen atom, a cyano group, a hydroxy group, -NR 4 R 5 is selected from.

[0033] In another preferred embodiment, R 4 , R 5 are each independently H or a C1-C4 alkyl group. In another preferred embodiment, each R 3 are each independently selected from H, a C1-C3 halogenated alkyl group, or a halogen atom. 3 is -F.

[0034] In another preferred embodiment, R 3 together with the carbon attached to the A ring,

[0035] [ka] Form.

[0036] In another preferred example, the compound represented by formula (I), (Ia) or (Ib) is selected from example compounds E1 to E73-2. In a third aspect, the present invention provides a pharmaceutical composition comprising a therapeutically effective amount of at least one compound represented by general formula (I), (Ia), or (Ib), its stereoisomer, or a pharmaceutically acceptable salt thereof, and a pharmaceutically acceptable adjuvant, such as a pharmaceutically acceptable carrier, excipient, sustained-release agent, flavoring agent, or perfume.

[0037] In the pharmaceutical composition, the compound of the present invention, as an active ingredient, accounts for 0.1 to 99.9% by weight of the total weight of the pharmaceutical composition, with the remainder being pharmaceutically acceptable adjuvants. The preferred ratio of the compound of the present invention to the adjuvants is such that the compound of the present invention, as an active ingredient, accounts for 60% or more of the total weight, with the remainder accounting for 0 to 40% of the total weight, with the amount of the remainder being preferably 1 to 20%, and most preferably 1 to 10%.

[0038] The compounds or pharmaceutical compositions according to the present invention can be formulated into various dosage forms such as tablets, capsules, powders, syrups, solutions, suspensions, sprays, creams, ointments, gels, transdermal patches, etc., using conventional formulation techniques in the pharmaceutical formulation field, and can be present in a suitable solid or liquid carrier or diluent. The pharmaceutical compositions of the present invention can also be stored in a suitable sterile syringe or sterile injectable.

[0039] The compound or pharmaceutical composition according to the present invention can be administered to mammals, including humans and animals. The administration route includes oral administration, nasal inhalation, topical administration to the skin, intravenous injection, intramuscular injection, subcutaneous injection, etc. In another preferred embodiment, the preferred administration route of the compound or pharmaceutical composition according to the present invention is oral administration.

[0040] Solid dosage forms of the compounds or pharmaceutical compositions described in the present invention for oral administration include capsules, tablets, pills, powders, and granules. Solid carriers include starch, lactose, dicalcium phosphate, microcrystalline cellulose, sucrose, kaolin, etc., while liquid carriers include sterilized water, polyethylene glycol, nonionic surfactants, and edible oils (e.g., corn oil, peanut oil, and sesame oil), etc., depending on the properties of the appropriate active ingredient and the desired specific administration route. Adjuvants commonly used in the preparation of pharmaceutical compositions, such as flavors, dyes, preservatives, and antioxidants, such as vitamin E, vitamin C, BHT, and BHA, can also be advantageously included.

[0041] Liquid dosage forms for oral administration include pharmaceutically acceptable emulsions, solutions, suspensions, syrups, or tinctures. In addition to the active ingredient, liquid dosage forms can contain inert diluents conventionally used in the art, such as water or other solvents, and solubilizers and emulsifiers, such as ethanol, isopropanol, ethyl carbonate, ethyl acetate, propylene glycol, 1,3-butanediol, dimethylformamide, and oils, particularly cottonseed oil, peanut oil, corn germ oil, olive oil, castor oil, and sesame oil, or mixtures of these substances. In addition to these inert diluents, compositions can also contain auxiliary substances, such as wetting agents, emulsifying and suspending agents, sweeteners, flavoring agents, and fragrances.

[0042] In addition to the active ingredient, suspensions may contain suspending agents such as, for example, ethoxylated isostearyl alcohols, polyoxyethylene sorbitol and dehydrated sorbitan esters, microcrystalline cellulose, aluminum methoxide and agar-agar, or mixtures of these substances.

[0043] When used as a pharmaceutical preparation, the compounds of the present invention are preferably administered in unit doses, with each dose containing 0.01 mg to 200 mg of the active ingredient, preferably 0.5 mg to 50 mg, administered as a single dose or in batches. Regardless of the oral administration method, the optimal dose for an individual will vary depending on the specific treatment. Typically, a small dose is started and gradually increased until the optimal dose is found.

[0044] In a fourth aspect, the present invention provides a use of a compound represented by general formula (I), (Ia), or (Ib), a stereoisomer thereof, a pharmaceutically acceptable salt thereof, or a pharmaceutical composition thereof, wherein the compound is used as a modulator of TRPC4 and TRPC5 ion channels, and therefore can be used in the preparation of a medicament for preventing, delaying, or treating a disease associated with abnormalities in the function or expression of TRPC4 and / or TRPC5.

[0045] In the present invention, the regulators of the TRPC4 and TRPC5 ion channels include inhibitors and agonists. In a preferred embodiment, the modulators of TRPC4 and TRPC5 ion channels are TRPC4 inhibitors and TRPC5 inhibitors.

[0046] In the present invention, diseases associated with abnormalities in the function or expression of TRPC4 and / or TRPC5 include, but are not limited to, psychiatric disorders, neurodegenerative disorders, kidney diseases, pain, epilepsy, liver diseases, cardiovascular diseases, cancer, skin diseases, intestinal diseases, etc.

[0047] In a preferred embodiment, the disease associated with abnormalities in the function or expression of TRPC4 and / or TRPC5 is a psychiatric disorder, including depression-related disorders such as borderline personality disorder, melancholy, dysthymia, postpartum depression, and bipolar disorder, anxiety- and fear-related disorders such as post-traumatic stress disorder, panic disorder, agoraphobia, social phobia, generalized anxiety disorder, social anxiety disorder, and separation anxiety, and affective and psychotic disorders caused by various other causes such as schizophrenia, mania, obsessive-compulsive disorder, apathy, neurasthenia, and paranoia.

[0048] In another preferred embodiment, the psychiatric disorder is borderline personality disorder, depression, anxiety disorder, or post-traumatic stress disorder. The compounds described in the present invention or pharmaceutical compositions thereof can be used alone or in combination with other drugs with the same mechanism of action in the treatment of psychiatric disorders. Other drugs with the same mechanism of action include, but are not limited to, tricyclic antidepressants (TCAs), monoamine oxidase inhibitors (MAOIs), selective serotonin (5-HT) reuptake inhibitors (SSRIs), serotonin and noradrenaline reuptake inhibitors (SNRIs), noradrenaline-specific 5-HT antidepressants (NaSSAs), and 5-HT receptor antagonists and reuptake inhibitors (SARIs). SSRIs and SNRIs are preferred. Representative SSRIs include fluoxetine, paroxetine, citalopram, sertraline, fluvoxamine, and the like. Representative SNRIs include duloxetine, venlafaxine, milnacipran, and the like.

[0049] In another preferred embodiment, the disease associated with abnormalities in the function or expression of TRPC4 and / or TRPC5 is a neurodegenerative disease, including Alzheimer's disease (AD), Parkinson's disease, Huntington's disease, amyotrophic lateral sclerosis (ALS), memory impairment, amnesia, aphasia, chronic fatigue syndrome, Creutzfeldt-Jakob disease, dissociative amnesia, fuguetic amnesia, learning disorders, sleep disorders, and other brain disorders caused by trauma or aging.

[0050] In another preferred embodiment, the disease associated with abnormalities in the function or expression of TRPC4 and / or TRPC5 is a pain disorder. The pain disorder includes nociceptive pain, mechanical pain, inflammatory pain, cancer pain, and neuropathic pain (e.g., osteoarthritis pain, rheumatoid arthritis pain, postherpetic neuralgia, and pain caused by burns). The pain may be chronic or acute.

[0051] In another preferred embodiment, the disease associated with abnormalities in the function or expression of TRPC4 and / or TRPC5 is epilepsy. Epilepsy can be caused by excitotoxicity due to various causes, and excessive neuronal firing usually drives epileptic activity. Compounds that reduce the hyperexcitability of related neuronal populations have significant potential in reducing epileptic activity. Inhibiting TRPC5-mediated calcium ion influx can reduce hyperexcitability and, therefore, epileptic activity.

[0052] In another preferred embodiment, the disease associated with abnormalities in the function or expression of TRPC4 and / or TRPC5 is a kidney disease, including nephropathy caused by toxic substances, nephropathy caused by viral or bacterial infection, and polar and chronic kidney disorders caused by various causes such as hypertensive nephropathy, diabetic nephropathy, glomerulonephritis, lupus nephritis, IgA nephropathy, nephrotic syndrome, membranous nephropathy, and minimal change kidney disease, and further including hereditary kidney diseases such as polycystic kidney disease and focal segmental glomerulosclerosis.

[0053] In another preferred embodiment, the disease associated with abnormalities in the function or expression of TRPC4 and / or TRPC5 is a skin disease, which is associated with keratinocyte dysfunction, such as, but not limited to, psoriasis, ichthyosis, palmoplantar keratosis, Olmsted disease, toadflax, or climacteric keratosis.

[0054] In another preferred embodiment, the disease associated with abnormalities in the function or expression of TRPC4 and / or TRPC5 is a cardiovascular disease, including, but not limited to, hypertension, arteriosclerosis, coronary artery disease, angina pectoris, myocardial infarction, arrhythmia, stroke, and pulmonary hypertension.

[0055] In another preferred embodiment, the disease associated with abnormalities in the function or expression of TRPC4 and / or TRPC5 is a liver disease, including, but not limited to, cholestatic liver disease, alcoholic steatohepatitis, nonalcoholic steatohepatitis, viral hepatitis, autoimmune liver disease, chemical-induced liver injury, liver fibrosis, liver cirrhosis, and hepatocellular carcinoma.

[0056] In another preferred embodiment, the disease associated with abnormalities in the function or expression of TRPC4 and / or TRPC5 is cancer, including, but not limited to, renal cell carcinoma, breast cancer, colorectal cancer, etc.

[0057] In another preferred embodiment, the disease associated with abnormalities in the function or expression of TRPC4 and / or TRPC5 is an intestinal disease, including, but not limited to, ulcerative colitis, short bowel syndrome, irritable bowel syndrome, intestinal spasm, diarrhea, abdominal pain, etc.

[0058] Lysolecithin (LPC) can directly activate TRPC5, independent of other indirect activation mechanisms such as the GPCR-PLC pathway. A range of biological effects of LPC are related to TRPC5 activation. Therefore, TRPC5 inhibitors may be useful for treating diseases associated with dysregulated LPC levels, including, but not limited to, arteriosclerosis, cardiovascular disease, central and peripheral nervous system diseases, osteoarthritis, hepatitis, hepatocellular carcinoma, pneumonia, obesity, gastrointestinal diseases, bacterial infections, parasitic diseases, diabetes, tumors, and pain.

[0059] In a fifth aspect, the present invention provides a method for preparing the compound of the present invention, its stereoisomer, or its pharmaceutically acceptable salt, wherein the method is selected from one of the synthetic routes shown in Route A and Route B below.

[0060] When the compound of the present invention has the structure shown in general formula (II-a) to (II-d), it can be prepared according to the synthetic route shown in Route A, where halo is a halogen atom and R 3a is a C1-C6 alkyl group, THP is a tetrahydropyran protecting group, and the other substituents are the same as defined above;

[0061] [ka]

[0062] The reaction steps of Route A include: (1.1) Intermediate INT-A1 (synthesized with reference to WO2014143799) and a commercially available ketone raw material are subjected to lithium halogen exchange and nucleophilic addition to generate intermediate INT-A2. The reaction is carried out in an ultra-dry aprotic solvent, such as, but not limited to, dichloromethane (DCM) or tetrahydrofuran (THF). The preferred solvent is tetrahydrofuran. The reaction temperature is generally −40 to −100° C., preferably −78° C. (1.2) Intermediate INT-A2 is subjected to a fluorination reaction with diethylaminosulfur trifluoride (DAST), the reaction temperature is generally 0°C to room temperature, and intermediates INT-A3 and INT-A4 are produced. The reaction is carried out in an aprotic solvent such as DCM, the reaction temperature is generally 0°C to room temperature. (1.3) Intermediate INT-A2 and R 3a -halo and NaH in tetrahydrofuran, the reaction temperature is generally room temperature, to produce intermediate INT-A5, (1.4) The THP protecting group is removed from the intermediates INT-A2, INT-A3, INT-A4, and INT-A5 to produce compounds represented by general formulas (II-a), (II-b), (II-c), and (II-d). The deprotection reaction of the hydroxy group is usually carried out under the action of an acidic reagent, and the acidic reagent is trifluoroacetic acid, hydrogen chloride, or p-toluenesulfonic acid, but is not limited to these.

[0063] When the compound of the present invention has the structure shown in general formula (II-e), it can be prepared according to the following route B, wherein halo is a halogen atom, THP represents a tetrahydropyran protecting group, and the definitions of other substituents are the same as above,

[0064] [ka]

[0065] The reaction steps of Route B include: (2.1) Suzuki coupling reaction of intermediate INT-A1 with boron ester-based raw material (commercially available or synthesized according to literature methods) to generate intermediate INT-B1. The Suzuki coupling reaction is usually carried out under the influence of a palladium catalyst and a base. Examples of the palladium catalyst include, but are not limited to, Pd(PhP)Cl, Pd(dppf)Cl, and Pd(PhP). The base includes, but is not limited to, triethylamine, DIPEA, KCO, and CsCO. The reaction is carried out in a suitable organic solvent such as DMF or 1,4-dioxane, or a mixture of an organic solvent and water. The reaction temperature is generally between room temperature and 100°C. (2.2) The intermediate INT-B1 is subjected to a hydrogenation reduction reaction to produce an intermediate INT-B2, the reduction reaction being carried out under the catalysis of palladium / carbon, the reaction temperature is generally room temperature, and the reaction solvent includes protic solvents and aprotic solvents, such as, but not limited to, methanol, ethanol, ethyl acetate, THF, etc.

[0066] (2.3) The THP protecting group of intermediate INT-B2 is removed to produce a compound represented by general formula (II-e). The deprotection reaction of the hydroxy group is usually carried out under the action of an acidic reagent, such as, but not limited to, trifluoroacetic acid, hydrogen chloride, or p-toluenesulfonic acid.

[0067] Those skilled in the art can appropriately adjust the reaction conditions based on literature sources or the actual situations encountered during the synthesis process. [Effects of the invention] It should be understood that within the scope of the present invention, the above-mentioned technical features of the present invention and the technical features specifically described below (e.g., in the Examples) can be combined with each other to form new or preferred technical solutions. Each feature disclosed in the specification can be replaced by any alternative feature providing the same, equivalent, or similar purpose. Due to space limitations, they will not be repeated here. [Brief explanation of the drawings]

[0068] [Figure 1] The effect of compound E21 on TGF-β-induced hepatic stellate cell activation (α-SMA labeling) is shown. (A) Microscopic images. (B) Statistics of the ratio of α-SMA-positive cells. **, p<0.01, Mean±SEM. [Figure 2] The drug concentrations in various tissues of rats 2 hours after oral administration of 10 mg / kg of compound E68 are shown, Mean±SD, N=3. [Figure 3] In the marble-burying test, the difference in the number of buried marbles in mice before and after oral administration of Fluoxetine (10 mg / kg), HC-070 (1 mg / kg), and E68 (0.1, 0.3, 1 mg / kg) is shown. ns, p>0.05, *, p<0.05, **, p<0.01, Mean±SEM, N=12-13. [Figure 4] In the tail suspension test, the difference in immobility time in the tail suspension box of mice before and after oral administration of Fluoxetine (10 mg / kg), HC-070 (1 mg / kg), and E68 (0.1, 0.3, 1 mg / kg) is shown. ns, p>0.05, *, p<0.05, **, p<0.01, Mean±SEM, N=12-13. [Figure 5]In the marble-burying test, the difference in the number of buried marbles in mice before and after oral administration of HC-070 (0.1, 0.3 mg / kg) and E68 (0.03, 0.1 mg / kg) is shown. ns, p>0.05, **, p<0.01, Mean±SEM, N=12-13. [Figure 6] In the tail suspension test, the difference in immobility time in the tail suspension box between mice before and after oral administration of HC-070 (0.1, 0.3 mg / kg) and E68 (0.03, 0.1 mg / kg) is shown. ns, p>0.05, *, p<0.05, **, p<0.01, Mean±SEM, N=12-13. DETAILED DESCRIPTION OF THE INVENTION

[0069] After extensive and thorough research, the inventors of the present application have developed xanthine-based compounds substituted with a "cycloalkyl group" at the 8-position. Compared with HC-070, the compounds of the present invention have stronger inhibitory activity against TRPC4 and TRPC5, and weaker inhibition against hERG and TRPC3, thus exhibiting superior selectivity. In liver microsome stability tests, the compounds of the present invention also exhibit superior metabolic stability, thus exhibiting superior drug potential. Furthermore, some compounds of the present invention unexpectedly exhibit significant TRPC5 agonist activity, rather than inhibitory activity. Based on this, the present invention was completed.

[0070] term In the present invention, unless otherwise specified, the terms used have their ordinary meanings known to those skilled in the art.

[0071] In the present invention, the term "C1-C6" refers to having 1, 2, 3, 4, 5 or 6 carbon atoms, and the same applies hereinafter. "4 to 8-membered" refers to having 4, 5, 6, 7 or 8 ring atoms, and the same applies hereinafter.

[0072] As used herein, the term "alkyl group" refers to branched and straight chain hydrocarbon groups having a specific number of carbon atoms, and representative examples include, but are not limited to, methyl, ethyl, n-propyl, and isopropyl groups.

[0073] In the present invention, the term "alkoxy group" refers to an -O-alkyl group. For example, the term "C1-C6 alkoxy group" refers to a straight or branched alkoxy group having 1 to 6 carbon atoms, and representative examples include, but are not limited to, a methoxy group, an ethoxy group, an n-propoxy group, an isopropoxy group, and a butoxy group.

[0074] In the present invention, the terms "halogenated alkyl group" and "halogenated alkoxy group" refer to groups in which the hydrogen atoms in an "alkyl group" or "alkoxy group" having a specific number of carbon atoms are partially or completely substituted with "halogen atoms."

[0075] In the present invention, the "cyclic hydrocarbon group" refers to a non-aromatic cyclic aliphatic hydrocarbon group having a specific number of ring carbon atoms, and the "C3-C10 cyclic hydrocarbon group" refers to a cyclic aliphatic hydrocarbon group consisting of 3 to 10 ring carbon atoms. The "cyclic hydrocarbon group" described in the present invention includes not only monocyclic aliphatic hydrocarbon groups, but also fused rings, spiro rings, and bridged rings consisting of multiple cyclic aliphatic hydrocarbons. The "cyclic hydrocarbon group" described in the present invention includes not only aliphatic hydrocarbon groups whose carbon atoms are fully saturated (i.e., cycloalkyl), but also aliphatic hydrocarbon groups whose carbon atoms have unsaturated bonds (e.g., cycloalkenyl), but does not include aryl groups consisting of fully unsaturated carbons. Examples of the "cyclic hydrocarbon group" described in the present invention are:

[0076] [ka] Including, but not limited to:

[0077] For purposes of this invention, "aryl groups" are defined as monocyclic and bicyclic ring systems of a specified number of carbon atoms and conforming to the Huckel rules, including phenyl, naphthyl, and the like.

[0078] In the present invention, "heteroaryl group" is defined as a monocyclic or bicyclic ring system consisting of a specific number of ring-forming atoms and containing 1, 2, 3 or 4 heteroatoms (selected from N, O and S) at the same time according to the Huckel rules, and examples of "heteroaryl group" include, but are not limited to, pyridine, pyrrole, imidazole, thiophene, benzimidazole, benzothiophene, benzofuran, etc.

[0079] In the present invention, a "heterocyclic group" is defined as a saturated or partially unsaturated non-aromatic monocyclic or bicyclic ring system consisting of a specified number of ring-forming atoms and containing 1, 2, 3, or 4 heteroatoms (selected from N, O, and S). Examples of "heterocyclic groups" include, but are not limited to, oxetanyl, dioxolanyl, morpholinyl, piperazinyl, pyrrolidinyl, piperidinyl, decahydroquinolinyl, benzotetrahydrofuran, and the like.

[0080] In the present invention, the "halogen atom" includes fluorine, chlorine, bromine, and iodine. "Substituted" as used herein refers to being replaced by one or more groups (e.g., 2, 3, 4, or 5). If no specific atom is specified, it means that the number of substituents may be present on any atom that is not saturated. When multiple substituents are selected from the same series, they may be the same or different.

[0081] "Optionally" according to the present invention means that the defined group may or may not be selected from a set of possible groups. A "pharmaceutically acceptable salt" according to the present invention can be a salt formed between an anion and a positively charged group on a compound of formula (I), (Ia), or (Ib). Suitable anions include chloride, bromide, iodide, sulfate, nitrate, phosphate, citrate, methanesulfonate, trifluoroacetate, acetate, malate, toluenesulfonate, tartrate, fumarate, glutamate, glucuronate, lactate, glutarate, or maleate. Similarly, a salt can be formed between a cation and a negatively charged group on a compound of formula (I). Suitable cations include sodium, potassium, magnesium, calcium, and ammonium ions, such as tetramethylammonium.

[0082] In another preferred example, the term "pharmaceutically acceptable salt" refers to a salt of a compound of formula (I), (Ia) or (Ib) with hydrofluoric acid, hydrochloric acid, hydrobromic acid, phosphoric acid, acetic acid, oxalic acid, sulfuric acid, nitric acid, methanesulfonic acid, aminosulfonic acid, salicylic acid, trifluoromethanesulfonic acid, naphthalenesulfonic acid, maleic acid, citric acid, acetic acid, lactic acid, tartaric acid, succinic acid, oxalic acid, pyruvic acid, malic acid, glutamic acid, p-toluenesulfonic acid, naphthalenesulfonic acid, ethanesulfonic acid, naphthalenedisulfonic acid, malonic acid, fumaric acid, propionic acid, oxalic acid, trifluoromethanesulfonic acid, The term "salt system" refers to a salt system formed with an acid selected from the group consisting of sodium, potassium, calcium, aluminum or ammonium salts formed from compounds of formula (I), (Ia) or (Ib) and an inorganic base, such as acetic acid, stearic acid, pamoic acid, hydroxymaleic acid, phenylacetic acid, benzoic acid, glutamic acid, ascorbic acid, p-aminobenzenesulfonic acid, 2-acetoxybenzoic acid and isethionic acid, or from compounds of formula (I), (Ia) or (Ib) and an organic base, or from methylamine salts, ethylamine salts or ethanolamine salts formed from compounds of formula (I), (Ia) or (Ib) and an organic base.

[0083] The compounds of general formula (I), (Ia), or (Ib) of the present invention, or their pharmaceutically acceptable salts, can be obtained by distillation, crystallization, or recrystallization from water or an organic solvent, and the compounds may further contain the solvent molecules used. Furthermore, different crystallization conditions can result in different crystalline forms of the compounds. Therefore, the compounds of general formula (I), (Ia), or (Ib) or their pharmaceutically acceptable salts, including various chemical amounts of crystallization solvents and all crystalline forms, are all within the scope of the present invention.

[0084] In the present invention, the term "therapeutically effective amount" means that a subject treated with the dose achieves cure, improvement, effective prevention, or significant reduction in the incidence of lesions or side effects, etc., compared with a subject not treated with the dose, and further includes a dose effective for enhancing normal physiological functions.

[0085] In the present invention, the term "modulator" includes inhibitors and agonists. The term "inhibitor" may also be written as "antagonist" or "blocker," and has the same meaning in the present invention, and refers to a compound or mixture that can be used to reduce or inhibit biological activity. The term "agonist" may also be written as "activator," and has the same meaning in the present invention, and refers to a drug molecule or mixture that can be used to enhance biological activity.

[0086] TRPC family proteins exist not only in the form of homozygous tetramers but also often in the form of heterozygous tetramers. Take TRPC5 as an example, which forms not only TRPC5:C5 homozygous channels but also heterozygous channels such as TRPC4:C5, TRPC1:C5, and TRPC1:C4:C5. Therefore, the TRPC5 inhibitor or TRPC5 agonist described in the present invention refers not only to inhibitors or agonists of TRPC5 homozygous channels but also to inhibitors or agonists of TRPC5 heterozygous channels. Similarly, the same meaning applies to TRPC4.

[0087] Some compounds of the present invention represented by general formulas (I), (Ia), and (Ib) have chiral centers, potential chiral centers, or unsaturated bonds, and can form various stereoisomers, such as racemates, enantiomers, diastereomers, E / Z isomers, cis-trans isomers, and tautomers. Unless otherwise specified, a given chemical formula or name in this specification and the appended claims is intended to encompass all stereoisomers, as well as mixtures of individual isomers in various ratios, and pharmaceutically acceptable salts thereof. Those skilled in the art may separate compounds of the present invention containing asymmetric centers to obtain single isomers using separation methods commonly used in laboratories, but this does not detract from the novelty of the compounds of the present invention.

[0088] The replacement of hydrogen atoms with deuterium atoms to alter the physical and chemical properties of a compound is a structural modification method well known to those skilled in the art. Unless otherwise specified, the present invention is intended to encompass deuterated forms of the compounds represented by general formulas (I), (Ia), and (Ib).

[0089] The present invention will be further described below by way of examples. Note that these examples are intended only to illustrate the present invention and are not intended to limit the present invention in any way. All parameters and other descriptions in the examples are by weight unless otherwise specified. Unless otherwise specified, all packing materials used in column chromatography separations are silica gel. In the following examples, experimental methods for which specific conditions are not specified are generally carried out according to conventional conditions or manufacturer recommendations.

[0090] Unless otherwise defined, all technical and scientific terms used herein have the same meaning as those familiar to one of ordinary skill in the art. Furthermore, any methods and materials similar or equivalent to those described herein can be applied to the present invention.

[0091] Abbreviations: DCM: dichloromethane, DAST: diethylaminosulfur trifluoride; DMAP: 4-dimethylaminopyridine, DMF: N,N-dimethylformamide, DMSO: dimethyl sulfoxide, TFA: trifluoroacetic acid, THF: tetrahydrofuran.

[0092] 1. Preparation Examples Example E1: 5-(4-chlorobenzyl)-6-(cyclopent-1-en-1-yl)-3-(3-hydroxypropyl)-1-methyl-1,5-dihydro-2H-pyrrolo[3,2-d]pyrimidine-2,4(3H)-dione

[0093] [ka]

[0094] 6-Chlorouracil (10 g, 68.493 mmol) was dissolved in 40 mL of DMSO, and KCO (4.8 g, 34.783 mmol) and CHI (12.7 mL, 205.48 mmol) were added sequentially. The mixture was stirred at room temperature for 3 hours. When the reaction was complete as determined by TLC, 50 mL of H2O was added, the mixture was suction filtered, and the filter cake was washed with water to obtain 7.9 g of a white solid, intermediate INT 1-1.

[0095] Intermediate INT 1-1 (5 g, 31.25 mmol) was dissolved in 10 mL of DMF, and K2CO3 (8.65 g, 62.5 mmol) and 3-bromopropyl methyl ether (5.35 mL, 46.875 mmol) were added, followed by heating at 60 °C for 3 hours. When TLC showed the reaction was complete, the organic phase was separated with EA / saturated NaCl aqueous solution, dried over anhydrous sodium sulfate, and then separated on a flash silica gel chromatography column eluted with DCM / MeOH as the mobile phase. The desired product fraction was collected and evaporated to dryness to obtain 4 g of a gel-like substance, i.e., intermediate INT 1-2.

[0096] Intermediate INT 1-2 (1 g, 4.31 mmol) was dissolved in 3 mL of concentrated sulfuric acid and stirred at 0° C. for 5 minutes. 3 mL of concentrated nitric acid was slowly added dropwise and stirred at 0° C. for 1 hour. When TLC showed the reaction was complete, the mixture was separated with EA / ice water. The organic layer was washed four times with saturated brine and dried over anhydrous sodium sulfate. Then, the mixture was separated on a flash silica gel chromatography column using DCM / MeOH as the mobile phase. The desired product was collected and evaporated to dryness to obtain 800 mg of a gel-like substance, i.e., intermediate INT 1-3.

[0097] Diethyl malonate (3.24 mL, 21.66 mmol) was dissolved in 10 mL of dry 1,4-dioxane, potassium t-butoxide (2.16 g, 19.494 mmol) was added, and the mixture was stirred at room temperature for 10 minutes. Intermediate INT 1-3 (3 g, 10.83 mmol) was added, and the mixture was stirred at room temperature for 1 hour. When TLC showed the reaction was complete, the mixture was acidified with 2 M HCl, and the organic phase was separated with EA / saturated NaCl solution. After drying over anhydrous sodium sulfate, the mixture was subjected to flash silica gel chromatography using DCM / MeOH as the mobile phase. The desired product fraction was collected and evaporated to dryness to obtain 3.2 g of a gel-like substance, i.e., intermediate INT 1-4.

[0098] Intermediate INT 1-4 (3 g, 7.463 mmol) was dissolved in 15 mL of acetic acid, heated to 60 °C, and zinc powder (7.3 g, 111.94 mmol) was added. The mixture was then reacted in a pressure tube at 120 °C for 12 hours. When TLC showed the reaction was complete, the mixture was evaporated to dryness under reduced pressure to obtain the crude product. The crude product was then separated by flash silica gel chromatography using MeOH / DCM as the mobile phase. The desired product fraction was collected and evaporated to dryness to obtain 1.8 g of a gel-like substance, i.e., intermediate INT 1-5.

[0099] Intermediate INT 1-5 (100 mg, 7.463 mmol) was dissolved in 3 mL of dry 1,4-dioxane, and phosphorus oxybromide (340 mg, 22.389 mmol) was added under nitrogen gas protection, followed by stirring at 100° C. for 1 hour. When TLC showed the reaction was complete, the organic phase was separated with EA / saturated NaHCO3 aqueous solution, dried over anhydrous sodium sulfate, and then separated on a flash silica gel chromatography column eluted with DCM / MeOH as the mobile phase. The desired product fraction was collected and evaporated to dryness to obtain 90 mg of a gel-like substance, i.e., intermediate INT 1-6.

[0100] Intermediate INT 1-6 (90 mg, 0.286 mmol) was dissolved in 5 mL of DMF, and KCO (1.73 g, 12.5 mmol) and 4-chlorobenzyl bromide (116 mg, 0.572 mmol) were added, followed by heating at 60° C. for 2 hours. When TLC showed the reaction was complete, the organic phase was separated with EA / saturated NaCl solution, dried over anhydrous sodium sulfate, and then separated on a flash silica gel chromatography column eluted with DCM / MeOH as the mobile phase. The desired product fraction was collected and evaporated to dryness to give 80 mg of a white solid, i.e., intermediate INT 1-7.

[0101] Intermediate INT 1-7 (40 mg, 0.091 mmol) was dissolved in 3 mL of dry DCM, and 182 μL of 1 M BBr solution was added at 0° C. and stirred at the same temperature for 1 hour. When TLC showed the reaction was complete, the crude product was evaporated to dryness under reduced pressure. The crude product was then separated by flash silica gel chromatography using MeOH / DCM as the mobile phase. The desired product fraction was collected and evaporated to dryness to obtain 30 mg of a white solid, i.e., intermediate INT 1-8.

[0102] INT 1-8 (25 mg, 0.049 mmol), 1-cyclopenteneboronic acid (13.7 mg, 0.122 mmol), Pd(dppf)Cl2 (5.38 mg, 0.007 mmol), Cs2CO3 (48 mg, 0.147 mmol), 10 mL of 1,4-dioxane, and 3 mL of HO were placed in a pressure tube and degassed with nitrogen gas for 3 minutes. The tube was sealed and heated at 100 °C for 2 hours. After cooling, the mixture was diluted with ethyl acetate, filtered to remove insoluble material, and the filtrate was evaporated to dryness under reduced pressure to obtain the crude product. The crude product was then separated by flash silica gel chromatography using MeOH / DCM as the mobile phase. The desired product fraction was collected and evaporated to dryness to obtain 25 mg of a gel-like substance. The crude product was purified by preparative liquid chromatography column eluting with acetonitrile / water system as the mobile phase, and the target product fraction was collected and evaporated to dryness to obtain 15 mg of a white solid, ie, Example E1. 1 H NMR(500MHz,Chloroform-d)δ 7.31-7.28(m,2H),6.96(d,J=8.5Hz,2H),5.98(s,1H),5.90-5.88(m,1H),5.74(s,2H),4.18(t,J=5Hz,2H),3. 52(s,3H),3.51-3.49(m,2H),2.76-2.64(m,2H),2.56-2.51(m,2H),1.99(p,J=7.5Hz,2H),1.93-1.86(m,2H). MS(ESI):m / z 414.1[M+H] + .

[0103] Example E2: 5-(4-chlorobenzyl)-6-cyclopentyl-3-(3-hydroxypropyl)-1-methyl-1,5-dihydro-2H-pyrrolo[3,2-d]pyrimidine-2,4(3H)-dione

[0104] [ka]

[0105] E1 (30 mg, 0.072 mmol) was dissolved in 3 ml of methanol, 10 mg of Pd / C was added, and the mixture was purged with hydrogen gas and reacted at room temperature for 2 hours. After suction filtration, the filtrate was concentrated to dryness and purified by preparative liquid chromatography using an acetonitrile / water system as the mobile phase. The target product was collected and evaporated to dryness to obtain 20 mg of a white solid, i.e., Example E2. 1 H NMR(500MHz,Chloroform-d)δ 7.30-7.28(m,2H),6.94(d,J=8.5Hz,2H),5.89(s,1H),5.66-5.64(m,2H),4.19-4.16(m,2H),3.58-3.53(m,2H),3. 52-3.49(m,3H),2.96(h,J=8.1Hz,1H),2.03-1.94(m,2H),1.94-1.87(m,2H),1.85-1.77(m,2H),1.72-1.51(m,4H). MS(ESI):m / z 416.2[M+H] + .

[0106] Example E3: 5-(4-chlorobenzyl)-6-(2-cyclopentylcyclopropyl)-3-(3-hydroxypropyl)-1-methyl-1,5-dihydro-2H-pyrrolo[3,2-d]pyrimidine-2,4(3H)-dione

[0107] [ka]

[0108] Bis(pinacolato)diboron (2.026 g, 7.976 mmol), Cu (36 mg, 0.571 mmol), and MeONa (62 mg, 1.142 mmol) were dissolved in EtOH sequentially. Under nitrogen gas protection, cyclopentylacetylene (500 mg, 5.319 mmol) was added and stirred at room temperature for 12 hours. When TLC showed the reaction was complete, the organic phase was separated with EA / saturated NH4Cl solution, dried over anhydrous sodium sulfate, and then separated on a flash silica gel chromatography column using PE / EA as the mobile phase. The desired product fraction was collected and evaporated to dryness to obtain 350 mg of a gel-like substance, i.e., intermediate INT 2-1.

[0109] KOH (1.5 g, 26.78 mmol) was dissolved in 3 mL of HO, 3 mL of ether was added, and methylnitrosourea (250 mg, 2.427 mmol) was added at 0 °C. After stirring for 5 minutes, the organic layer was transferred to an ether solution containing INT 2-1 (50 mg, 0.225 mmol) and Pd(OAc) (20 mg, 0.089 mmol). After stirring for 5 minutes at 0 °C, TLC showed the reaction was complete. Insoluble materials were removed by filtration, and the filtrate was evaporated to dryness under reduced pressure to obtain the crude product. The product was then separated using a flash silica gel chromatography column eluted with PE / EA as the mobile phase. The desired product fraction was collected and evaporated to dryness to obtain 30 mg of a gel-like substance, i.e., intermediate INT 2-2.

[0110] INT 1-8 (25 mg, 0.049 mmol), INT 2-2 (25 mg, 0.106 mmol), Pd(dppf)Cl2 (5.38 mg, 0.007 mmol), Cs2CO3 (48 mg, 0.147 mmol), 10 mL of 1,4-dioxane, and 3 mL of HO were placed in a pressure tube and degassed with nitrogen gas for 3 minutes. The tube was sealed and heated and stirred at 100 °C for 2 hours. After cooling, the mixture was diluted with ethyl acetate, filtered to remove insoluble material, and the filtrate was evaporated to dryness under reduced pressure to obtain the crude product. The crude product was then separated by flash silica gel chromatography using MeOH / DCM as the mobile phase. The desired product fraction was collected and evaporated to dryness to obtain 20 mg of a gel-like substance. The crude product was purified by preparative liquid chromatography column eluting with acetonitrile / water system as the mobile phase, and the target product fraction was collected and evaporated to dryness to obtain 15 mg of a white solid, Example E3. 1H NMR(500MHz,Chloroform-d)δ 7.31-7.29(m,2H),7.02(d,J=8.5Hz,2H),5.83-5.72(m,1H),5.71-5.52(m,2H), 4.21-4.15(m,2H),3.58-3.51(m,2H),3.49-3.43(m,3H),1.94-1.87(m,2H),1.78 -1.70(m,2H),1.68-1.61(m,2H),1.59-1.51(m,2H),1.49(dt,J=9.1,4.8Hz,1H) ,1.41(p,J=8.0Hz,1H),1.34-1.19(m,2H),1.15-1.00(m,1H),0.93-0.84(m,2H). MS(ESI):m / z 456.2[M+H] + .

[0111] Example E4: 7-(4-chlorobenzyl)-8-(2-cyclopentylcyclopropyl)-1-(3-hydroxypropyl)-3-methyl-3,7-dihydro-1H-purine-2,6-dione

[0112] [ka]

[0113] 8-Bromo-3-methyl-3,7-dihydro-purine-2,6-dione (10 g, 40.816 mmol) was dissolved in 30 ml of DMF, and KHCO (6.18 g, 61.8 mmol) and 4-chlorobenzyl bromide (8.37 g, 40.829 mmol) were added, followed by stirring at 60° C. for 2 hours. When TLC showed the reaction was complete, 100 ml of H2O was added, suction filtered, and the filter cake was dried to obtain 14.1 g of a white solid, intermediate INT 4-1.

[0114] INT 4-1 (14.1 g, 38.3 mmol) was dissolved in 30 ml of DMF, and KCO (10.6 g, 76.8 mmol) and 2-(3-bromopropoxy)tetrahydro-2H-pyran (7.8 ml, 45.978 mmol) were added, followed by stirring for 2 hours at 60° C. When TLC showed the reaction was complete, 100 ml of H2O was added, and the mixture was suction filtered and the filter cake was dried to obtain 20.3 g of a white solid, intermediate INT 4-2.

[0115] INT 4-2 (100 mg, 0.196 mmol), INT 2-2 (92.5 mg, 0.392 mmol), Pd(dppf)Cl2 (14.3 mg, 0.019 mmol), Cs2CO3 (191.1 mg, 0.588 mmol), 10 mL of 1,4-dioxane, and 3 mL of HO were placed in a pressure tube and degassed with nitrogen gas for 3 minutes. The tube was then sealed and heated and stirred at 100 °C for 2 hours. After cooling, the mixture was diluted with ethyl acetate, filtered to remove insoluble material, and the filtrate was evaporated to dryness under reduced pressure to obtain the crude product. The crude product was then separated using a flash silica gel chromatography column eluted with MeOH / DCM as the mobile phase. The desired product fraction was collected and evaporated to dryness to obtain 80 mg of a gel-like substance, i.e., intermediate INT 4-3.

[0116] INT 4-3 (80 mg, 0.148 mmol) was dissolved in 2 mL of DCM, and then 0.4 mL of TFA was added. The mixture was stirred at room temperature for 1 hour, and TLC showed the reaction was complete. The solvent was removed under reduced pressure, and the residue was purified by preparative liquid chromatography using an acetonitrile / water system as the mobile phase. A portion of the desired product was collected and evaporated to dryness to obtain 50 mg of a white solid, i.e., Example E4. 1H NMR (500 MHz, Chloroform-d) δ 7.35(d,J=8.5Hz,2H),7.19(d,J=8.5Hz,2H),5.61(d,J=4.5Hz,2H),4.19( t,J=5Hz,2H),3.55(s,3H),3.52(t,J=5.5Hz,2H),2.08-1.99(m,1H),1.93 -1.86(m,2H),1.81-1.72(m,1H),1.71-1.60(m,4H),1.58-1.51(m,2H),1. 50-1.39(m,2H),1.25-1.16(m,1H),1.03-0.98(m,1H),0.95-0.88(m,1H). MS(ESI): m / z 457.2[M+H] + .

[0117] Example E5: 5-(4-chlorobenzyl)-3-(3-hydroxypropyl)-1-methyl-6-(4-phenylcyclohexyl)-1,5-dihydro-2H-pyrrolo[3,2-d]pyrimidine-2,4(3H)-dione

[0118] [ka]

[0119] 4-Phenylcyclohexanone (870 mg, 5 mmol) was dissolved in ultra-dry DCM, NaCO3 was added, and the mixture was stirred for 10 minutes. TfO (1.55 g, 5.5 mmol) was added dropwise at 0 °C, and the mixture was stirred at room temperature for 3 hours. TLC showed the reaction was complete. 5 ml of MeOH was added, and the reaction mixture was evaporated to dryness under reduced pressure to obtain the crude product. The crude product was then separated by flash silica gel chromatography using PE / EA as the mobile phase. The desired product fraction was collected and evaporated to dryness to obtain 1 g of a colorless oily liquid, intermediate INT 5-1.

[0120] INT 5-1 (300 mg, 0.98 mmol), bis(pinacolato)diboron (498 mg, 1.96 mmol), Pd(dppf)Cl2 (71 mg, 0.097 mmol), KOAc (288 mg, 2.939 mmol), and 10 mL of 1,4-dioxane were placed in a pressure tube and degassed with nitrogen gas for 3 minutes. The tube was then sealed and heated and stirred at 80 °C for 2 hours. After cooling, the mixture was diluted with ethyl acetate, filtered to remove insoluble material, and the filtrate was evaporated to dryness under reduced pressure to obtain the crude product. The crude product was then separated using a flash silica gel chromatography column eluted with PE / EA as the mobile phase. The desired product fraction was collected and evaporated to dryness to obtain 260 mg of a gel-like substance, i.e., intermediate INT 5-2.

[0121] INT 1-8 (25 mg, 0.049 mmol), INT 5-2 (25 mg, 0.088 mmol), Pd(dppf)Cl2 (5.38 mg, 0.007 mmol), Cs2CO3 (48 mg, 0.147 mmol), 10 mL of 1,4-dioxane, and 3 mL of HO were placed in a pressure tube and degassed with nitrogen gas for 3 minutes. The tube was then sealed and heated and stirred at 100 °C for 2 hours. After cooling, the mixture was diluted with ethyl acetate, filtered to remove insoluble material, and the filtrate was evaporated to dryness under reduced pressure to obtain the crude product. The crude product was then separated using a flash silica gel chromatography column eluted with MeOH / DCM as the mobile phase. The desired product fraction was collected and evaporated to dryness to obtain 20 mg of a gel-like substance, i.e., intermediate INT 5-3.

[0122] INT 5-3 (20 mg, 0.04 mmol) was dissolved in 3 ml of methanol, 5 mg of Pd / C was added, and the mixture was reacted at room temperature for 2 hours after purging with hydrogen gas. After suction filtration, the filtrate was concentrated to dryness and purified by preparative liquid chromatography using an acetonitrile / water system as the mobile phase. The target product was collected and evaporated to dryness to obtain 15 mg of a white solid, i.e., Example E5. 1H NMR(500MHz,Chloroform-d)δ 7.38-7.34(m,1H),7.34-7.29(m,4H),7.27-7.20(m,2H),7.01-6.90( m,2H),5.93-5.89(m,1H),5.71-5.64(m,2H),4.23-4.13(m,2H),3.54- 3.51(m,2H),3.49(s,3H),2.94-2.86(m,1H),2.68-2.56(m,1H),2.14- 1.98(m,2H),1.96-1.81(m,4H),1.81-1.74(m,2H),1.70-1.62(m,2H). MS(ESI): m / z 506.2[M+H] + .

[0123] Example E6: 7-(4-chlorobenzyl)-1-(3-hydroxypropyl)-3-methyl-8-(1,2,3,6-tetrahydro-[1,1'-biphenyl]-4-yl)-3,7-dihydro-1H-purine-2,6-dione

[0124] [ka]

[0125] INT 4-2 (100 mg, 0.196 mmol), INT 5-2 (111.3 mg, 0.392 mmol), Pd(dppf)Cl2 (14.3 mg, 0.019 mmol), Cs2CO3 (191.1 mg, 0.588 mmol), 10 mL of 1,4-dioxane, and 3 mL of HO were placed in a pressure tube and degassed with nitrogen gas for 3 minutes. The tube was then sealed and heated and stirred at 100 °C for 2 hours. After cooling, the mixture was diluted with ethyl acetate, filtered to remove insoluble material, and the filtrate was evaporated to dryness under reduced pressure to obtain the crude product. The crude product was then separated using a flash silica gel chromatography column eluted with MeOH / DCM as the mobile phase. The desired product fraction was collected and evaporated to dryness to obtain 100 mg of a gel-like substance, intermediate INT 6-1.

[0126] INT 6-1 (100 mg, 0.17 mmol) was dissolved in 4 mL of DCM, and then 1 mL of TFA was added. The mixture was stirred at room temperature for 1 hour, and TLC showed the reaction was complete. The solvent was removed under reduced pressure, and the residue was purified by preparative liquid chromatography using an acetonitrile / water system as the mobile phase. The desired product was collected and evaporated to dryness to obtain 80 mg of a white solid, i.e., Example E6. 1 H NMR(500MHz,Chloroform-d)δ 7.40-7.32(m,3H),7.30-7.29(m,2H),7.27-7.25(m,2H),7.09(d,J=8.0Hz,2H),6.24-6.20(m,1H),5.62(s,2H),4.23-4.16(m,2H),3 .66-3.63(m,3H),3.56-3.52(m,2H),2.92-2.85(m,1H),2.66-2.46(m,3H),2.42-2.33(m,1H),2.19-2.08(m,1H),2.02-1.92(m,3H). MS(ESI):m / z 505.2[M+H] + .

[0127] Example E7: 7-(4-chlorobenzyl)-1-(3-hydroxypropyl)-8-(4-isopropoxycyclohexyl)-3-methyl-3,7-dihydro-1H-purine-2,6-dione

[0128] [ka]

[0129] 4-Isopropoxycyclohexanone (250 mg, 1.6 mmol) was dissolved in ultra-dry DCM, NaCO was added, and the mixture was stirred for 10 minutes. TfO (497 mg, 1.76 mmol) was added dropwise at 0 °C, and the mixture was stirred at room temperature for 3 hours. TLC showed the reaction was complete. 5 ml of MeOH was added, and the reaction mixture was evaporated to dryness under reduced pressure to obtain the crude product. The crude product was then separated by flash silica gel chromatography using PE / EA as the mobile phase. The desired product fraction was collected and evaporated to dryness to obtain 200 mg of a colorless oily liquid, intermediate INT 7-1.

[0130] INT 7-1 (190 mg, 0.66 mmol), bis(pinacolato)diboron (335 mg, 1.319 mmol), Pd(dppf)Cl2 (50 mg, 0.068 mmol), KOAc (241 mg, 1.975 mmol), and 10 mL of 1,4-dioxane were placed in a pressure tube and degassed with nitrogen gas for 3 minutes. The tube was then sealed and heated and stirred at 80 °C for 2 hours. After cooling, the mixture was diluted with ethyl acetate, filtered to remove insoluble material, and the filtrate was evaporated to dryness under reduced pressure to obtain the crude product. The crude product was then separated using a flash silica gel chromatography column eluted with PE / EA as the mobile phase. The desired product fraction was collected and evaporated to dryness to obtain 150 mg of a gel-like substance, intermediate INT 7-2.

[0131] INT 4-2 (100 mg, 0.196 mmol), INT 7-2 (104.3 mg, 0.392 mmol), Pd(dppf)Cl2 (14.3 mg, 0.019 mmol), Cs2CO3 (191.1 mg, 0.588 mmol), 10 mL of 1,4-dioxane, and 3 mL of HO were placed in a pressure tube and degassed with nitrogen gas for 3 minutes. The tube was then sealed and heated and stirred at 100 °C for 2 hours. After cooling, the mixture was diluted with ethyl acetate, filtered to remove insoluble material, and the filtrate was evaporated to dryness under reduced pressure to obtain the crude product. The crude product was then separated using a flash silica gel chromatography column eluted with MeOH / DCM as the mobile phase. The desired product fraction was collected and evaporated to dryness to obtain 120 mg of a gel-like substance, i.e., intermediate INT 7-3.

[0132] INT 7-3 (100 mg, 0.175 mmol) was dissolved in 5 ml of methanol, 20 mg of Pd / C was added, and the mixture was reacted at room temperature for 2 hours after purging with hydrogen gas. After suction filtration, the filtrate was concentrated to dryness to obtain 80 mg of a gel-like substance, i.e., intermediate INT 7-4.

[0133] INT 7-4 (80 mg, 0.14 mmol) was dissolved in 4 mL of DCM, and then 1 mL of TFA was added. The mixture was stirred at room temperature for 1 hour, and TLC showed the reaction was complete. The solvent was removed under reduced pressure, and the residue was purified by preparative liquid chromatography using an acetonitrile / water system as the mobile phase. The desired product was collected and evaporated to dryness to obtain 55 mg of a white solid, i.e., Example E7. 1 H NMR(500MHz,DMSO-d6)δ 7.42(d,J=8.5Hz,2H),7.21(d,J=8.5Hz,2H),5.60(s,2H),4.49-4.45(m,1H),3.95-3.85(m,2H),3.64-3.57(m,2H),3.50-3.43 (m,4H),2.97-2.80(m,1H),1.87-1.73(m,4H),1.72-1.63(m,2H),1.50-1.40(m,2H),1.35-1.28(m,2H),1.08(d,J=6.1Hz,6H). MS(ESI):m / z 489.2[M+H] + .

[0134] Example E8: 7-(4-chlorobenzyl)-1-(3-hydroxypropyl)-8-(4-isopropoxycyclohex-1-en-1-yl)-3-methyl-3,7-dihydro-1H-purine-2,6-dione

[0135] [ka]

[0136] INT 7-3 (40 mg, 0.07 mmol) was dissolved in 4 mL of DCM, and then 1 mL of TFA was added. The mixture was stirred at room temperature for 1 hour, and TLC showed the reaction was complete. The solvent was removed under reduced pressure, and the residue was purified by preparative liquid chromatography using an acetonitrile / water system as the mobile phase. A portion of the desired product was collected and evaporated to dryness to obtain 25 mg of a white solid, i.e., Example E8. 1H NMR(500MHz,Chloroform-d)δ 7.32(d,J=8.5Hz,2H),7.15(d,J=8.5Hz,2H),6.03-5.99(m,1H),5.58(s,2H),4.20(t,J=6.0Hz,2H),3.84-3.69(m,2H),3.62(s,3H), 3.56-3.50(m,2H),3.50-3.45(m,1H),2.62-2.37(m,3H),2.34-2.16(m,1H),2.01-1.88(m,3H),1.86-1.78(m,1H),1.23-1.18(m,6H). MS(ESI):m / z 487.2[M+H] + .

[0137] Example E9: 8-([1,1'-bis(cyclopropane)]-2-yl)-7-(4-chlorobenzyl)-1-(3-hydroxypropyl)-3-methyl-3,7-dihydro-1H-purine-2,6-dione

[0138] [ka]

[0139] KOH (1.5 g, 26.78 mmol) was dissolved in 3 mL of HO, 3 mL of ether was added, and methylnitrosourea (250 mg, 2.427 mmol) was added at 0 °C. After stirring for 5 min, the organic layer was transferred to an ether solution of (E)-2-cyclopropylvinylboronic acid pinacol ester (44 mg, 0.225 mmol) and Pd(OAc) (20 mg, 0.089 mmol). After stirring for 5 min at 0 °C, TLC showed the reaction was complete. Insoluble materials were removed by filtration, and the filtrate was evaporated to dryness under reduced pressure to obtain the crude product. The crude product was then separated by flash silica gel chromatography using PE / EA as the mobile phase. The desired product fraction was collected and evaporated to dryness to obtain 35 mg of a gel-like substance, intermediate INT 9-1.

[0140] INT 4-2 (100 mg, 0.196 mmol), INT 9-1 (81.5 mg, 0.392 mmol), Pd(dppf)Cl2 (14.3 mg, 0.019 mmol), Cs2CO3 (191.1 mg, 0.588 mmol), 10 mL of 1,4-dioxane, and 3 mL of HO were placed in a pressure tube and degassed with nitrogen gas for 3 minutes. The tube was then sealed and heated and stirred at 100 °C for 2 hours. After cooling, the mixture was diluted with ethyl acetate, filtered to remove insoluble material, and the filtrate was evaporated to dryness under reduced pressure to obtain the crude product. The crude product was then separated using a flash silica gel chromatography column eluted with MeOH / DCM as the mobile phase. The desired product fraction was collected and evaporated to dryness to obtain 100 mg of a gel-like substance, i.e., intermediate INT 9-2.

[0141] INT 9-2 (100 mg, 0.195 mmol) was dissolved in 4 mL of DCM, and then 1 mL of TFA was added. The mixture was stirred at room temperature for 1 hour, and TLC showed the reaction was complete. The solvent was removed under reduced pressure, and the residue was purified by preparative liquid chromatography using an acetonitrile / water system as the mobile phase. The desired product was collected and evaporated to dryness to obtain 70 mg of a white solid, i.e., Example E9. 1 H NMR(500MHz,Chloroform-d)δ 7.35(d,J=8.5Hz,2H),7.24(d,J=8.5Hz,2H),5.71-5.50(m,2H),4.24-4.13(m,2H) ),3.62-3.55(m,1H),3.54(s,3H),3.53-3.49(m,2H),1.93-1.86(m,2H),1.66(dt ,J=8.8,4.7Hz,1H),1.52-1.47(m,1H),1.37(dt,J=9.1,4.7Hz,1H),1.02-0.88(m ,1H),0.86-0.79(m,1H),0.57-0.47(m,1H),0.46-0.35(m,1H),0.22-0.06(m,2H). MS(ESI):m / z 429.2[M+H] + .

[0142] Example E10: 7-(4-chlorobenzyl)-8-(4,4-dimethylcyclohexyl)-1-(3-hydroxypropyl)-3-methyl-3,7-dihydro-1H-purine-2,6-dione

[0143] [ka]

[0144] INT 4-2 (100 mg, 0.196 mmol), 4,4-(dimethylcyclohexen-1-yl)boronic acid pinacol ester (92.5 mg, 0.392 mmol), Pd(dppf)Cl2 (14.3 mg, 0.019 mmol), Cs2CO3 (191.1 mg, 0.588 mmol), 10 mL of 1,4-dioxane, and 3 mL of HO were placed in a pressure tube and degassed with nitrogen gas for 3 minutes. The tube was then sealed and heated and stirred at 100 °C for 2 hours. After cooling, the mixture was diluted with ethyl acetate, filtered to remove insoluble material, and the filtrate was evaporated to dryness under reduced pressure to obtain the crude product. The crude product was then separated by flash silica gel chromatography using MeOH / DCM as the mobile phase. The desired product fraction was collected and evaporated to dryness to obtain 100 mg of a gel-like substance, intermediate INT 10-1.

[0145] INT 10-1 (100 mg, 0.185 mmol) was dissolved in 5 ml of methanol, 20 mg of Pd / C was added, and the mixture was reacted at room temperature for 2 hours after purging with hydrogen gas. After suction filtration, the filtrate was concentrated to dryness to obtain 80 mg of a gel-like substance, i.e., intermediate INT 10-2.

[0146] INT 7-4 (80 mg, 0.15 mmol) was dissolved in 4 mL of DCM, and then 1 mL of TFA was added. The mixture was stirred at room temperature for 1 hour, and TLC showed the reaction was complete. The solvent was removed under reduced pressure, and the residue was purified by preparative liquid chromatography using an acetonitrile / water system as the mobile phase. The desired product was collected and evaporated to dryness to obtain 60 mg of a white solid, i.e., Example E10. 1H NMR(500MHz,Chloroform-d)δ 7.34(d,J=8.5Hz,2H),7.12(d,J=8.5Hz,2H),5.54(s,2H),4.20(t,J=6.0Hz,2H),3.63(s,3H),3.58-3.51(m,2H),2.59 (tt,J=11.9,3.5Hz,1H),1.95-1.83(m,4H),1.51(td,J=7.5,3.6Hz,4H),1.30-1.19(m,2H),1.02(s,3H),0.97(s,3H). MS(ESI):m / z 459.2[M+H] + .

[0147] Example E11: 7-(4-chlorobenzyl)-8-(4,4-dimethylcyclohex-1-en-1-yl)-1-(3-hydroxypropyl)-3-methyl-3,7-dihydro-1H-purine-2,6-dione

[0148] [ka]

[0149] INT 7-3 (50 mg, 0.093 mmol) was dissolved in 4 mL of DCM, and then 1 mL of TFA was added. The mixture was stirred at room temperature for 1 hour, and TLC showed the reaction was complete. The solvent was removed under reduced pressure, and the residue was purified by preparative liquid chromatography using an acetonitrile / water system as the mobile phase. The desired product was collected and evaporated to dryness to obtain 25 mg of a white solid, i.e., Example E11. 1 H NMR(500MHz,Chloroform-d)δ 7.32(d,J=8.5Hz,2H),7.05(d,J=8.5Hz,2H),6.05-6.02(m,1H),5.59(s,2H),4.23-4.14(m,2H),3.64(s,3H),3.52(t ,J=5.5Hz,2H),2.42(dt,J=6.3,3.7Hz,2H),2.02-1.99(m,2H),1.93-1.86(m,2H),1.53(t,J=6.4Hz,2H),0.98(s,6H). MS(ESI):m / z 457.2[M+H] + .

[0150] Example E12: 7-(4-chlorobenzyl)-1-(3-hydroxypropyl)-3-methyl-8-(4-(trifluoromethyl)cyclohexyl)-3,7-dihydro-1H-purine-2,6-dione

[0151] [ka]

[0152] The synthesis method of Example E12 is the same as that of E10, except that 4-(trifluoromethyl)-1-cyclohexene-1-boronic acid pinacol ester is used as the starting material instead of 4,4-(dimethylcyclohexen-1-yl)boronic acid pinacol ester. 1 H NMR(500MHz,Chloroform-d)δ 7.34(d,J=8.5Hz,2H),7.08(d,J=8.5Hz,2H),5.56(s,2H),4.21(t,J=6.0Hz,2H),3.63(s,3H),3.56-3.5 2(m,2H),3.52-3.47(m,1H),3.07-3.02(m,1H),2.29-2.12(m,3H),1.95-1.87(m,4H),1.76-1.65(m,4H). MS(ESI):m / z 499.2[M+H] + .

[0153] Example E13: 7-(4-chlorobenzyl)-1-(3-hydroxypropyl)-3-methyl-8-(1,4-dioxacyclo[4.5]dec-8-yl)-3,7-dihydro-1H-purine-2,6-dione

[0154] [ka]

[0155] The synthesis method of Example E13 is the same as that of E10, except that 1,4-dioxa-spiro[4,5]dec-7-ene-8-boronic acid pinacol ester is used as the starting material instead of 4,4-(dimethylcyclohexen-1-yl)boronic acid pinacol ester. 1 H NMR(500MHz,Chloroform-d)δ 7.34(d,J=8.5Hz,2H),7.11(d,J=8.5Hz,2H),5.56(s,2H),4.20(t,J=6.0Hz,2H),4.00(d,J=1.7Hz,4H),3.61(s,3H),3.53(t,J=5.5Hz ,2H),2.72(tt,J=11.6,3.6Hz,1H),2.04(qd,J=13.4,3.6Hz,2H),1.94-1.85(m,4H),1.73-1.66(m,2H),1.58(td,J=13.4,4.1Hz,2H). MS(ESI):m / z 489.2[M+H] + .

[0156] Example E14: 8-([1,1'-bis(cyclopropane)]-2-yl)-7-(4-chlorobenzyl)-1-(3-hydroxypropyl)-3-isopropyl-3,7-dihydro-1H-purine-2,6-dione

[0157] [ka]

[0158] 8-Bromoxanthine (2 g, 8.658 mmol) was dissolved in 10 ml of DMF, and KHCO (1.298 g, 12.987 mmol) and 4-chlorobenzyl bromide (1.77 g, 8.658 mmol) were added, followed by stirring at 60° C. for 2 hours. When TLC showed the reaction was complete, 10 ml of H2O was added, and the mixture was suction filtered and the filter cake was dried to give 2.5 g of a white solid, intermediate INT 14-1.

[0159] INT 14-1 (1 g, 2.825 mmol) was dissolved in 10 ml of DMF, and KCO (428 mg, 3.1 mmol) and 2-iodopropane (480 mg, 2.825 mmol) were added, followed by stirring for 2 hours at 40° C. When the reaction was complete as indicated by TLC, 10 ml of H2O was added, and the mixture was suction filtered and the filter cake was dried to give 400 mg of a white solid, intermediate INT 14-2.

[0160] INT 14-2 (350 mg, 0.883 mmol) was dissolved in 5 mL of DMF, and K2CO3 (244 mg, 1.768 mmol) and 2-(3-bromopropoxy)tetrahydro-2H-pyran (236.5 mg, 1.06 mmol) were added. The mixture was stirred at 60 °C for 2 hours. TLC showed the reaction was complete. The reaction mixture was poured into saturated aqueous NaCl solution and extracted three times with ethyl acetate. The organic phases were combined and washed three times with saturated aqueous NaCl solution. After drying over anhydrous MgSO4, the mixture was separated on a flash silica gel chromatography column and eluted with MeOH / DCM as the mobile phase. The desired product fraction was collected to give 250 mg of a colorless oily liquid, i.e., intermediate INT 14-3.

[0161] INT 14-3 (100 mg, 0.185 mmol), INT 9-1 (81.5 mg, 0.392 mmol), Pd(dppf)Cl2 (14.3 mg, 0.019 mmol), Cs2CO3 (191.1 mg, 0.588 mmol), 10 mL of 1,4-dioxane, and 3 mL of HO were placed in a pressure tube and degassed with nitrogen gas for 3 minutes. The tube was then sealed and heated and stirred at 100 °C for 2 hours. After cooling, the mixture was diluted with ethyl acetate, filtered to remove insoluble material, and the filtrate was evaporated to dryness under reduced pressure to obtain the crude product. The crude product was then separated using a flash silica gel chromatography column eluted with MeOH / DCM as the mobile phase. The desired product fraction was collected and evaporated to dryness to obtain 60 mg of a gel-like substance, i.e., intermediate INT 14-4.

[0162] INT 14-4 (60 mg, 0.111 mmol) was dissolved in 4 mL of DCM, and then 1 mL of TFA was added. The mixture was stirred at room temperature for 1 hour, and TLC showed the reaction was complete. The solvent was removed under reduced pressure, and the residue was purified by preparative liquid chromatography using an acetonitrile / water system as the mobile phase. The desired product was collected and evaporated to dryness to obtain 30 mg of a white solid, i.e., Example E14. 1 H NMR(500MHz,Chloroform-d)δ 7.36(d,J=8.5Hz,2H),7.27(d,J=8.3Hz,2H),5.73-5.42(m,2H),5.20-5.10(m,1 H),4.18(t,J=5.9Hz,2H),3.70(t,J=7.2Hz,1H),3.57-3.48(m,2H),1.91-1.83(m ,2H),1.66(dt,J=8.8,4.7Hz,1H),1.55(d,J=6.9Hz,6H),1.52-1.47(m,1H),1.36 (dt,J=9.1,4.7Hz,1H),0.97-0.79(m,2H),0.57-0.38(m,2H),0.19-0.08(m,2H). MS(ESI):m / z 457.2[M+H] + .

[0163] Example E15: 7-(4-chlorobenzyl)-8-(2-cyclopentylcyclopropyl)-1-(3-hydroxypropyl)-3-isopropyl-3,7-dihydro-1H-purine-2,6-dione

[0164] [ka]

[0165] INT 14-2 (100 mg, 0.185 mmol), INT 9-1 (81.5 mg, 0.392 mmol), Pd(dppf)Cl2 (14.3 mg, 0.019 mmol), Cs2CO3 (191.1 mg, 0.588 mmol), 10 mL of 1,4-dioxane, and 3 mL of HO were placed in a pressure tube and degassed with nitrogen gas for 3 minutes. The tube was then sealed and heated and stirred at 100 °C for 2 hours. After cooling, the mixture was diluted with ethyl acetate, filtered to remove insoluble material, and the filtrate was evaporated to dryness under reduced pressure to obtain the crude product. The crude product was then separated using a flash silica gel chromatography column eluted with MeOH / DCM as the mobile phase. The desired product fraction was collected and evaporated to dryness to obtain 80 mg of a gel-like substance, i.e., intermediate INT 15-1.

[0166] INT 15-1 (80 mg, 0.141 mmol) was dissolved in 4 mL of DCM, and then 1 mL of TFA was added. The mixture was stirred at room temperature for 1 hour, and TLC showed the reaction was complete. The solvent was removed under reduced pressure, and the residue was purified by preparative liquid chromatography using an acetonitrile / water system as the mobile phase. The desired product was collected and evaporated to dryness to obtain 50 mg of a white solid, i.e., Example E15. 1 H NMR(500MHz,Chloroform-d)δ 7.35(d,J=8.5Hz,2H),7.23(d,J=8.3Hz,2H),5.20-5.11(m,1H),4.17(t,J=5.9Hz,2H),3.70(t,J=7.2Hz,1H),3.55-3.47(m,2H),1.93-1.86( m,2H),1.81-1.74(m,1H),1.71-1.61(m,5H),1.56(d,J=6.8,6H),1.49 -1.38(m,3H),1.36-1.28(m,2H),1.26-1.17(m,1H),1.03-0.96(m,1H). MS(ESI): m / z 485.2[M+H] + .

[0167] Example E16: 7-(4-chlorobenzyl)-8-cyclopentyl-1-(3-methoxypropyl)-3-methyl-3,7-dihydro-1H-purine-2,6-dione

[0168] [ka]

[0169] INT 1-6 (100 mg, 0.227 mmol), 1-cyclopentenylboronic acid pinacol ester (88.1 mg, 0.454 mmol), Pd(dppf)Cl2 (14.3 mg, 0.019 mmol), Cs2CO3 (191.1 mg, 0.588 mmol), 10 mL of 1,4-dioxane, and 3 mL of HO were placed in a pressure tube and degassed with nitrogen gas for 3 minutes. The tube was then sealed and heated and stirred at 100 °C for 2 hours. After cooling, the mixture was diluted with ethyl acetate, filtered to remove insoluble material, and the filtrate was evaporated to dryness under reduced pressure to obtain the crude product. The crude product was then separated using a flash silica gel chromatography column eluted with MeOH / DCM as the mobile phase. The desired product fraction was collected and evaporated to dryness to obtain 80 mg of a gel-like substance, i.e., intermediate INT 16-1.

[0170] INT 16-1 (80 mg, 0.187 mmol) was dissolved in 5 ml of methanol, 15 mg of Pd / C was added, and the mixture was reacted at room temperature for 2 hours after purging with hydrogen gas. After suction filtration, the filtrate was concentrated to dryness to obtain 80 mg of a gel-like substance, which was purified by preparative liquid chromatography using an acetonitrile / water system as the mobile phase. The target product was collected and evaporated to dryness to obtain 50 mg of a white solid, i.e., Example E16. 1 H NMR(500MHz,Chloroform-d)δ 7.28(d,J=9Hz,2H),6.95(d,J=8.3Hz,2H),5.84(s,1H),5.68(s,2H),4.09(t,J=7.65Hz,2H),3.49(s,3H),3.45 (t,J=6.5Hz,2H),3.32(s,3H),2.95(p,J=8.1Hz,1H),2.00-1.90(m,,4H),1.85-1.77(m,2H),1.69-1.59(m,4H). MS(ESI):m / z 431.2[M+H] + .

[0171] Example E17: 7-(4-chlorobenzyl)-8-(1-fluoro-3-phenylcyclobutyl)-1-(3-hydroxypropyl)-3-methyl-3,7-dihydro-1H-purine-2,6-dione

[0172] [ka]

[0173] INT 4-2 (250 mg, 0.49 mmol) was dissolved in 5 ml of ultra-dry tetrahydrofuran, and isopropylmagnesium chloride lithium chloride complex (490 μL, 0.735 mmol) was added dropwise at 0° C., followed by stirring for 10 minutes. 3-phenylcyclobutanone (130 μL, 0.98 mmol) was added, and the mixture was stirred at 0° C. for 2 hours. LC-MS showed the reaction was complete, and the solvent was removed under reduced pressure. The mixture was then separated by flash silica gel chromatography using MeOH / DCM as the mobile phase. The desired product fraction was collected and evaporated to dryness to obtain 100 mg of a gel-like substance, i.e., intermediate INT 17-1.

[0174] Intermediate INT 17-1 (100 mg, 0.173 mmol) was dissolved in 3 ml of ultra-dry DCM, and DAST (113 μL, 0.865 mmol) was added dropwise at 0° C. and stirred for 30 min at 0° C. LC-MS showed the reaction was complete, and the mixture was evaporated to dryness to give 80 mg of a gel-like material, intermediate INT 17-2.

[0175] INT 17-2 (80 mg, 0.138 mmol) was dissolved in 4 mL of DCM, and then 1 mL of TFA was added. The mixture was stirred at room temperature for 1 hour, and TLC showed the reaction was complete. The solvent was removed under reduced pressure, and the residue was purified by preparative liquid chromatography using an acetonitrile / water system as the mobile phase. The desired product was collected and evaporated to dryness to obtain 50 mg of a white solid, i.e., Example E17. 1H NMR(500MHz,Chloroform-d)δ 7.41-7.35(m,3H),7.34-7.30(m,4H),7.21(d,J=8.2Hz,2H),5.68(s,2H),4.23(t,J=6.0Hz,2H),3.68(s ,3H),3.60-3.52(m,2H),3.27(p,J=8.9Hz,1H),3.16-3.08(m,2H),2.79-2.66(m,2H),1.97-1.88(m,2H). MS(ESI):m / z 497.2[M+H] + .

[0176] Example E18: (E)-7-(4-chlorobenzyl)-1-(3-hydroxypropyl)-3-methyl-2,6-dioxo-2,3,6,7-tetrahydro-1H-purine-8-carbaldehyde-O-(t-butyl)oxime

[0177] [ka]

[0178] INT 4-2 (250 mg, 0.49 mmol) was dissolved in 5 ml of ultra-dry tetrahydrofuran, and 2.5 M n-butyllithium (235 μL, 0.588 mmol) was added dropwise at −78° C., followed by stirring for 30 minutes. Ultra-dry DMF (57 μL, 0.735 mmol) was added, and the mixture was stirred at −78° C. for 2 hours. LC-MS showed the reaction was complete, and the solvent was removed under reduced pressure. The mixture was then separated by flash silica gel chromatography using MeOH / DCM as the mobile phase. The desired product fraction was collected and evaporated to dryness to obtain 80 mg of a gel-like substance, i.e., intermediate INT 18-1.

[0179] Intermediate INT 18-1 (80 mg, 0.174 mmol) and 0-butylhydroxylamine hydrochloride (65.6 mg, 0.522 mmol) were dissolved in 3 mL of DCM and stirred for 12 hours. When TLC showed the reaction was complete, the solvent was removed under reduced pressure. The mixture was then separated by flash silica gel chromatography using MeOH / DCM as the mobile phase. The desired product fraction was collected and evaporated to dryness to obtain 40 mg of a gel-like substance, i.e., intermediate INT 18-2.

[0180] NT 17-2 (40 mg, 0.075 mmol) was dissolved in 4 mL of DCM, followed by the addition of 1 mL of TFA and stirring at room temperature for 1 hour. TLC showed the reaction was complete. The solvent was removed under reduced pressure, and the residue was purified by preparative liquid chromatography using an acetonitrile / water system as the mobile phase. A portion of the desired product was collected and evaporated to dryness to obtain 30 mg of a white solid, i.e., Example E18. 1 H NMR(500MHz,Chloroform-d)δ 8.22(s,1H),7.38-7.34(m,2H),7.22(d,J=8.5Hz,2H),6.03(s,1H),4.28-4.20(m,2H) ),3.67(s,3H),3.61-3.54(m,2H),3.48-3.25(m,1H),2.07-1.83(m,2H),1.31(s,9H). MS(ESI):m / z 448.2[M+H] + .

[0181] Example E19: 7-(4-chlorobenzyl)-8-(1-fluoro-4-isopropoxycyclohexyl)-1-(3-hydroxypropyl)-3-methyl-3,7-dihydro-1H-purine-2,6-dione

[0182] [ka]

[0183] The synthesis method of Example E19 is similar to that of E17, except that 4-isopropoxycyclohexanone is used as the starting material instead of 3-phenylcyclobutanone. 1H NMR(500MHz,Chloroform-d)δ 7.34-7.31(m,2H),7.20-7.09(m,2H),5.76(s,2H),4.21-4.13(m,2H),3.80-3.64(m,2H),3.64-3.56(m ,3H),3.55-3.37(m,2H),2.50-2.28(m,2H),2.18-2.00(m,4H),1.97-1.79(m,4H),1.22-1.14(ms,6H). MS(ESI):m / z 507.2[M+H] + .

[0184] Example E20: 7-(4-chlorobenzyl)-1-(3-hydroxypropyl)-8-(1H-inden-2-yl)-3-methyl-3,7-dihydro-1H-purine-2,6-dione

[0185] [ka]

[0186] The synthesis method of Example E20 is the same as that of E6, except that 1H-indene-2-boronic acid pinacol ester is used as the starting material instead of 4,4,5,5-tetramethyl-2-(1,2,3,6-tetrahydro-[1,1′-biphenyl]-4-yl)-1,3,2-dioxaborolane. 1 H NMR(500MHz,Chloroform-d)δ 7.58-7.52(m,1H),7.47-7.42(m,1H),7.39-7.32(m,4H),7.25(s,1H),7.11(d,J=8.5Hz,2H),5.86 (s,2H),4.20(t,J=6.0Hz,2H),4.02(s,2H),3.70(s,3H),3.54(t,J=5.4Hz,2H),1.96-1.88(m,2H). MS(ESI):m / z 463.2[M+H] + .

[0187] Example 21: 7-(4-chlorobenzyl)-8-(1-fluoro-4-(trifluoromethyl)cyclohexyl)-1-(3-hydroxypropyl)-3-methyl-3,7-dihydro-1H-purine-2,6-dione

[0188] [ka]

[0189] INT 4-2 (250 mg, 0.49 mmol) was dissolved in 5 ml of ultra-dry tetrahydrofuran, and 2.5 M n-butyllithium (235 μL, 0.588 mmol) was added dropwise at −78° C., followed by stirring for 30 minutes. 4-(trifluoromethyl)cyclohexanone (122 mg, 0.735 mmol) was added, and the mixture was stirred at −78° C. for 2 hours. LC-MS showed the reaction was complete, and the solvent was removed under reduced pressure. The mixture was then separated by flash silica gel chromatography using MeOH / DCM as the mobile phase. The desired product fraction was collected and evaporated to dryness to obtain 120 mg of a gel-like substance, i.e., intermediate INT 21-1.

[0190] Intermediate INT 21-1 (120 mg, 0.201 mmol) was dissolved in 3 ml of ultra-dry DCM, and DAST (53 μL, 0.402 mmol) was added dropwise at 0° C. and stirred for 30 minutes at 0° C. LC-MS showed the reaction was complete, and the mixture was evaporated to dryness to give 80 mg of a gel-like material, intermediate INT 21-2.

[0191] INT 21-2 (80 mg, 0.133 mmol) was dissolved in 4 mL of DCM, and then 1 mL of TFA was added. The mixture was stirred at room temperature for 1 hour, and TLC showed the reaction was complete. The solvent was removed under reduced pressure, and the residue was purified by preparative liquid chromatography using an acetonitrile / water system as the mobile phase. The desired product was collected and evaporated to dryness to obtain 50 mg of a white solid, i.e., Example E21. 1H NMR(500MHz,Chloroform-d,3:4 isomer)δ 7.30 / 7.29(2×d,J=7Hz,2H),7.15 / 7.09(2×d,J=8.2Hz,2H),5.76 / 5.75(2×s,2H),4.22-4.16(m,2H),3.58 / 3.61(2×s,3H),3.56- 3.50(m,2H),3.42-3.24(m,1H),2.46-2.36(m,1H),2.31-2.17(m,1H),2.17-2.08(m,1H),2.07-1.85(m,7H),1.82-1.71(m,1H). MS(ESI):m / z 517.2[M+H] + .

[0192] Example E22: 7-(4-chlorobenzyl)-1-(3-hydroxypropyl)-8-(1-methoxy-4-(trifluoromethyl)cyclohexyl)-3-methyl-3,7-dihydro-1H-purine-2,6-dione

[0193] [ka]

[0194] Intermediate INT 21-1 (120 mg, 0.201 mmol) was dissolved in 5 ml of ultra-dry THF, and NaH (16.08 mg, 0.402 mmol) and CHCl (38 μL, 0.603 mmol) were added. The mixture was allowed to react at 35° C. for 12 hours. When TLC showed the reaction was complete, the solvent was removed under reduced pressure. The mixture was then separated using a flash silica gel chromatography column, eluting with MeOH / DCM as the mobile phase. The desired product fraction was collected and evaporated to dryness to obtain 80 mg of a gel-like substance, i.e., intermediate INT 22-1.

[0195] INT 22-1 (80 mg, 0.133 mmol) was dissolved in 4 mL of DCM, and then 1 mL of TFA was added. The mixture was stirred at room temperature for 1 hour, and TLC showed the reaction was complete. The solvent was removed under reduced pressure, and the residue was purified by preparative liquid chromatography using an acetonitrile / water system as the mobile phase. The desired product was collected and evaporated to dryness to obtain 50 mg of a white solid, i.e., Example E22. 1 H NMR(500MHz,Chloroform-d)δ 7.31(d,J=8.5Hz,2H),6.98(d,J=8.5 Hz,2H),5.87(s,2H),4.21-4.09(m,2H),3.63(s,3H),3.54-3.49(m,2H),3.32(t,J=7.1H z,1H),3.00(s,3H),2.23-2.19(m,1H),1.92-1.78(m,6H),1.67(td,J=12.4,3.6Hz,4H). MS(ESI):m / z 529.2[M+H] + .

[0196] Example E23: 7-(4-chlorobenzyl)-8-(1-hydroxy-4-(trifluoromethyl)cyclohexyl)-1-(3-hydroxypropyl)-3-methyl-3,7-dihydro-1H-purine-2,6-dione

[0197] [ka]

[0198] INT 21-1 (40 mg, 0.067 mmol) was dissolved in 4 mL of DCM, and then 1 mL of TFA was added. The mixture was stirred at room temperature for 1 hour, and TLC showed the reaction was complete. The solvent was removed under reduced pressure, and the residue was purified by preparative liquid chromatography using an acetonitrile / water system as the mobile phase. The desired product was collected and evaporated to dryness to obtain 20 mg of a white solid, i.e., Example E23. 1H NMR(500MHz,Chloroform-d)δ 7.29(d,J=8.5Hz,2H),7.01(d,J=8.5Hz,2H),5.93(s,2H),4.18(t,J=6.0Hz,2H),3.61(s,3H),3.51-3.46( m,2H),3.38(t,J=7.1Hz,1H),2.17-2.04(m,3H),1.98-1.92(m,2H),1.90-1.83(m,4H),1.81-1.69(m,2H). MS(ESI):m / z 515.2[M+H] + .

[0199] Example E24: 7-(4-chlorobenzyl)-8-(1-fluoro-4,4-dimethylcyclohexyl)-1-(3-hydroxypropyl)-3-methyl-3,7-dihydro-1H-purine-2,6-dione

[0200] [ka]

[0201] The synthesis method of Example E24 is the same as that of E21, except that 4,4-dimethylcyclohexanone is used as the starting material instead of 4-(trifluoromethyl)cyclohexanone. 1 H NMR(500MHz,Chloroform-d)δ 7.28(d,J=8.5Hz,2H),7.12(d,J=8.5Hz,2H),5.76(s,2H),4.19(t,J=6.0Hz,2H),3.61(s,3H),3.52(t,J=5.5Hz,2H),2.2 6(td,J=14.1,4.4Hz,2H),1.93-1.86(m,4H),1.64(dd,J=13.7,4.0Hz,2H),1.38(d,J=13.7Hz,2H),1.02(d,J=6.0Hz,6H). MS(ESI):m / z 477.2[M+H] + .

[0202] Example E25: 7-(4-chlorobenzyl)-8-(1-hydroxy-4,4-dimethylcyclohexyl)-1-(3-hydroxypropyl)-3-methyl-3,7-dihydro-1H-purine-2,6-dione

[0203] [ka]

[0204] INT 24-1 (40 mg, 0.072 mmol) was dissolved in 4 mL of DCM, and then 1 mL of TFA was added. The mixture was stirred at room temperature for 1 hour, and TLC showed the reaction was complete. The solvent was removed under reduced pressure, and the residue was purified by preparative liquid chromatography using an acetonitrile / water system as the mobile phase. A portion of the desired product was collected and evaporated to dryness to obtain 20 mg of a white solid, i.e., Example E25. 1 H NMR(500MHz,Chloroform-d)δ 7.30(d,J=8Hz,2H),7.04(d,J=8.5Hz,2H),5.92(s,2H),4.16(t,J=6.0Hz,2H),3.62(s,3H),3.54-3.47(m,2H),3.46-3.41(m,1H),2. 23(td,J=13.7,4.2Hz,2H),1.91-1.83(m,2H),1.71(d,J=14.2Hz,2H),1.57(td,J=13.5,3.9Hz,2H),1.41-1.29(m,2H),1.00(s,6H). MS(ESI):m / z 475.2[M+H] + .

[0205] Example E26: 7-(4-chlorobenzyl)-1-(3-hydroxypropyl)-8-(1-methoxy-4,4-dimethylcyclohexyl)-3-methyl-3,7-dihydro-1H-purine-2,6-dione

[0206] [ka]

[0207] The synthesis method of Example E26 is the same as that of E22, except that INT 24-1 is used as the starting material instead of INT 21-1. 1 H NMR(500MHz,Chloroform-d)δ 7.30(d,J=8.6Hz,2H),6.97(d,J=9Hz,2H,2H),5.88(s,2H),4.27-4.03(m,2H),3.65(s,3H),3.56-3.48(m,2H),2.97( s,3H),2.03-1.97(m,2H),1.94-1.85(m,4H),1.50(td,J=13.0,3.8Hz,2H),1.31-1.23(m,2H),0.97(d,J=3.7Hz,6H). MS(ESI):m / z 489.2[M+H] + .

[0208] Example E27: (E)-8-(1-(t-butoxyimino)ethyl)-7-(4-chlorobenzyl)-1-(3-hydroxypropyl)-3-methyl-3,7-dihydro-1H-purine-2,6-dione

[0209] [ka]

[0210] INT 4-2 (200 mg, 0.392 mmol) was dissolved in 5 mL of 1,4-dioxane, and under N2 protection, tributyl(1-ethoxyethylene)tin (267 μL, 0.784 mmol) and Pd(PPh3)4 (45 mg, 0.039 mmol) were added, followed by stirring at 110 °C for 12 hours. When TLC showed the reaction was complete, the solvent was removed under reduced pressure. The mixture was then separated by flash silica gel chromatography using MeOH / DCM as the mobile phase. The desired product fraction was collected and evaporated to dryness to obtain 180 mg of a gel-like substance, i.e., intermediate INT 27-1. INT 27-1 (180 mg, 0.358 mmol) was dissolved in 3 ml of methanol, and 1 ml of 1N HCl was added dropwise at 0° C. The mixture was stirred at room temperature for 12 hours. When TLC showed the reaction was complete, the solvent was removed under reduced pressure. The mixture was then separated by flash silica gel chromatography using MeOH / DCM as the mobile phase. The desired product fraction was collected and evaporated to dryness to obtain 140 mg of a gel-like substance, i.e., intermediate INT 27-2.

[0211] INT 27-2 (140 mg, 0.295 mmol) and 0-butylhydroxylamine hydrochloride (74.3 mg, 0.59 mmol) were dissolved in 3 mL of DCM and stirred for 12 hours. When TLC showed the reaction was complete, the solvent was removed under reduced pressure. The mixture was then separated by flash silica gel chromatography using MeOH / DCM as the mobile phase. The desired product fraction was collected and evaporated to dryness to obtain 90 mg of a gel-like substance, i.e., intermediate INT 18-2.

[0212] NT 17-2 (90 mg, 0.165 mmol) was dissolved in 4 mL of DCM, followed by the addition of 1 mL of TFA and stirring at room temperature for 1 hour. TLC showed the reaction was complete. The solvent was removed under reduced pressure, and the residue was purified by preparative liquid chromatography using an acetonitrile / water system as the mobile phase. A portion of the desired product was collected and evaporated to dryness to obtain 50 mg of a white solid, i.e., Example E27. 1 H NMR(500MHz,Chloroform-d)δ 7.28(d,J=6.5Hz,2H),7.10(d,J=8.5Hz,2H),6.00(s,2H),4.17(t,J=6.0Hz,2H), 3.64(s,3H),3.51(t,J=5.6Hz,2H),2.38(s,3H),1.91-1.83(m,2H),1.21(s,9H). MS(ESI):m / z 462.2[M+H] + .

[0213] Example E28: 7-(4-chlorobenzyl)-8-(2-fluoro-6,6-dimethylbicyclo[3.1.1]heptan-2-yl)-1-(3-hydroxypropyl)-3-methyl-3,7-dihydro-1H-purine-2,6-dione

[0214] [ka]

[0215] The synthesis method of Example E28 is the same as that of E21, except that (1R,5S)-6,6-dimethylbicyclo[3.1.1]heptan-2-one is used as the starting material instead of 4-(trifluoromethyl)cyclohexanone. 1 H NMR(500MHz,Chloroform-d)δ 7.33(d,J=8.5Hz,2H),7.07(d,J=8.5Hz,2H),5.69-5.50(m,2H),4.33-3.97(m,2H),3.63(s,3H),3.55-3. 49(m,2H),2.64-2.58(m,1H),2.41-2.29(m,2H),2.06-2.00(m,4H),1.92-1.84(m,3H),0.96-0.86(m,6H). MS(ESI):m / z 489.2[M+H] + .

[0216] Example E29: 7-(4-chlorobenzyl)-8-(2-fluoro-6,6-dimethylbicyclo[3.1.1]heptan-2-yl)-1-(3-hydroxypropyl)-3-methyl-3,7-dihydro-1H-purine-2,6-dione

[0217] [ka]

[0218] Intermediate INT 28-1 (80 mg, 0.14 mmol) was dissolved in 3 ml of ultra-dry DCM, and DAST (37 μL, 0.28 mmol) was added dropwise at 0° C. and stirred for 30 minutes at 0° C. LC-MS showed the reaction was complete, and the mixture was evaporated to dryness to give 20 mg of a gel-like substance, intermediate INT 29-1.

[0219] INT 29-1 (20 mg, 0.036 mmol) was dissolved in 4 mL of DCM, and then 1 mL of TFA was added. The mixture was stirred at room temperature for 1 hour, and TLC showed the reaction was complete. The solvent was removed under reduced pressure, and the residue was purified by preparative liquid chromatography using an acetonitrile / water system as the mobile phase. The desired product was collected and evaporated to dryness to obtain 10 mg of a white solid, i.e., Example E29. 1 H NMR(500MHz,Chloroform-d)δ 7.33(d,J=8.5Hz,2H),7.11-6.92(m,2H),6.18-6.05(m,1H),5.58(s,2H),4.21-4.15(m,2H),3.63(s,3H),3.55-3.44 m,2H),2.64-2.51(m,1H),2.44-2.33(m,1H),2.29-2.21(m,1H),2.07-2.22(m,1H),1.95-1.83(m,2H),1.81-1.77(m,2H),0.98-0.81(m,6H). MS(ESI):m / z 469.2[M+H] + .

[0220] Example E30: 7-(4-chlorobenzyl)-8-(4-ethyl-1-fluorocyclohexyl)-1-(3-hydroxypropyl)-3-methyl-3,7-dihydro-1H-purine-2,6-dione

[0221] [ka]

[0222] The synthesis method of Example E30 is similar to that of E21, except that 4-ethylcyclohexanone is used as the starting material instead of 4-(trifluoromethyl)cyclohexanone. 1H NMR(500MHz,Chloroform-d,3:1 isomer)δ 7.31(d,J=8.5Hz,2H),7.16 / 7.11(2×d,J=8.2Hz,2H),5.76 / 5.75(2×s,2H),4.21-4.15(m,2H),3.59 / 3.62(2×s,3H),3.52(t,J= 6.1Hz,2H),3.45-3.36(m,1H),2.15-1.99(m,4H),1.93-1.86(m,3H),1.78-1.72(m,2H),1.37-1.30(m,4H),0.96-0.89(m,3H). MS(ESI):m / z 477.2[M+H] + .

[0223] Example E31: 7-(4-chlorobenzyl)-8-(1-fluoro-4-isopropylcyclohexyl)-1-(3-hydroxypropyl)-3-methyl-3,7-dihydro-1H-purine-2,6-dione

[0224] [ka]

[0225] The synthesis method of Example E31 is the same as that of E21, except that 4-isopropylcyclohexanone is used as the starting material instead of 4-(trifluoromethyl)cyclohexanone. 1 H NMR(500MHz,Chloroform-d,3:1 isomer)δ 7.31(d,J=8.5Hz,2H),7.16 / 7.10(2×d,J=8.2Hz,2H),5.76 / 5.75(2×s,2H),4.21-4.13(m,2H),3.59 / 3.62(2×s,3H),3.55-3.46(m,2H),2.17-1.98(m,4H),1.94-1.84 (m,2H),1.75-1.67(m,2H),1.55-1.48(m,1H),1.47-1.39(m,1H),1.38-1.31(m,2H),0.92(2×d,J=7Hz,6H). MS(ESI): m / z 491.2[M+H] + .

[0226] Example E32: 7-(4-chlorobenzyl)-8-(1-fluoro-4-methoxycyclohexyl)-1-(3-hydroxypropyl)-3-methyl-3,7-dihydro-1H-purine-2,6-dione

[0227] [ka]

[0228] The synthesis method of Example E32 is similar to that of E21, except that 4-methoxycyclohexanone is used as the starting material instead of 4-(trifluoromethyl)cyclohexanone. 1 H NMR(500MHz,Chloroform-d)δ 7.31(d,J=8.5Hz,2H),7.14(d,J=8.4Hz,2H),5.76(s,2H),4.18(t,J=6Hz,2H),3.61(s,3H),3.56(s,1H),3 .54-3.49(m,2H),3.42-3.39(m,1H),3.38(s,3H),2.46-2.29(m,2H),2.08-1.90(m,4H),1.88-1.78(m,4H). MS(ESI):m / z 479.2[M+H] + .

[0229] Example E33: 7-(4-chlorobenzyl)-8-(1-hydroxy-4-methoxycyclohexyl)-1-(3-hydroxypropyl)-3-methyl-3,7-dihydro-1H-purine-2,6-dione

[0230] [ka]

[0231] The synthesis method of Example E33 is the same as that of E25, except that INT 32-1 is used as the starting material instead of INT 24-1. 1H NMR(500MHz,Chloroform-d)δ 7.31(d,J=8.5Hz,2H),7.05(d,J=8.5Hz,2H),5.93(s,2H),4.16(t,J=6.0Hz,2H),3.61(s,3H),3.53-3.47(m,2H),3.39(s,3H),3.32-3. 25(m,1H),2.14(td,J=13.5,3.9Hz,2H),2.06-2.03(m,1H),2.01-1.97(m,1H),1.97-1.90(m,2H),1.91-1.83(m,2H),1.70-1.64(m,2H). MS(ESI): m / z 477.2[M+H] + .

[0232] Example E34: 7-(4-chlorobenzyl)-8-(1-fluoro-4,4-dimethylcyclohex-2-en-1-yl)-1-(3-hydroxypropyl)-3-methyl-3,7-dihydro-1H-purine-2,6-dione

[0233] [ka]

[0234] The synthesis method of Example E34 is similar to that of E21, except that 4,4-dimethyl-2-cyclohexyl-1-one is used as the starting material instead of 4-(trifluoromethyl)cyclohexanone. 1 H NMR(500MHz,Chloroform-d)δ 7.33(d,J=8.5Hz,2H),7.06(d,J=8.5Hz,2H),6.00(d,J=3.4Hz,1H),5.64(d,J=3.2Hz2H),4.74-4.64(m,1H),4.20(t,J=6Hz,2H),3.63 (s,3H),3.57-3.51(m,2H),3.42(t,J=7.1Hz,1H),2.54-2.33(m,2H),2.07-2.04(m,2H),1.91-1.83(m,2H),1.03(s,3H),0.94(s,3H). MS(ESI):m / z 475.2[M+H] + .

[0235] Example E35: 7-(4-chlorobenzyl)-8-(1-hydroxy-4,4-dimethylcyclohex-2-en-1-yl)-1-(3-hydroxypropyl)-3-methyl-3,7-dihydro-1H-purine-2,6-dione

[0236] [ka]

[0237] The synthesis method of Example E35 is the same as that of E25, except that INT 34-1 is used as the starting material instead of INT 24-1. 1 H NMR (500 MHz, Chloroform-d) δ 7.32(d,J=8.5Hz,2H),7.06(d,J=8.5Hz,2H),6.01-5.98(m,1H),5.64(d,J= 3.2Hz,2H),5.42-5.30(m,1H),4.19(t,J=5.5Hz,2H),3.97-3.93(m,1H),3.6 3(s,3H),3.55-3.50(m,2H),3.42(t,J=7Hz,1H),2.38-2.43(m,2H),2.28-2 .20(m,1H),2.07-2.01(m,1H),1.91-1.83(m,2H),1.03(s,3H),0.94(s,3H). MS(ESI): m / z 473.2[M+H] + .

[0238] Example E36: ​​7-(4-chlorobenzyl)-8-(4,4-difluorocyclohexyl)-1-(3-hydroxypropyl)-3-methyl-3,7-dihydro-1H-purine-2,6-dione

[0239] [ka]

[0240] The synthesis method of Example E36 is the same as that of E10, except that 2-(4,4-difluorocyclohexen-1-en-1-yl)boronic acid pinacol ester is used as the starting material instead of 4,4-(dimethylcyclohexen-1-yl)boronic acid pinacol ester. 1 H NMR(500MHz,Chloroform-d)δ 7.35(d,J=8.5Hz,2H),7.10(d,J=8.5Hz,2H),5.58(s,2H),4.20(t,J=6.0Hz,2H),3.62(s,3H),3.57-3.51(m,2H), 3.49-3.45(m,1H),2.82-2.73(m,1H),2.32-2.20(m,2H),2.10-2.02(m,2H),1.93-1.86(m,2H),1.86-1.72(m,4H). MS(ESI):m / z 467.2[M+H] + .

[0241] Example E37: 7-(4-chlorobenzyl)-8-(4,4-difluorocyclohexen-1-en-1-yl)-1-(3-hydroxypropyl)-3-methyl-3,7-dihydro-1H-purine-2,6-dione

[0242] [ka]

[0243] The synthesis method of Example E37 is the same as that of E25, except that INT 36-1 is used as the starting material instead of INT 24-1. 1 H NMR(500MHz,Chloroform-d)δ 7.33(d,J=8.5Hz,2H),7.09(d,J=8.1Hz,2H),5.91(s,1H),5.59(s,2H),4.21(t,J=6.0Hz,2H),3.62(s ,3H),3.56-3.51(m,2H),3.43-3.34(m,1H),2.82-2.61(m,4H),2.27-2.15(m,2H),1.93-1.86(m,2H). MS(ESI):m / z 465.1[M+H] + .

[0244] Example E38: 7-(4-chlorobenzyl)-1-(3-hydroxypropyl)-3-methyl-8-(1,4,4-trifluorocyclohexyl)-3,7-dihydro-1H-purine-2,6-dione

[0245] [ka]

[0246] The synthesis method of Example E38 is similar to that of E21, except that 4,4-difluorocyclohexanone is used as the starting material instead of 4-(trifluoromethyl)cyclohexanone. 1 H NMR(500MHz,Chloroform-d)δ 7.33(d,J=8.5Hz,2H),7.14(d,J=8.2Hz,2H),5.78(s,2H),4.20(t,J=6.0Hz,2H),3.60( s, 3H), 3.54 (t, J=5.6Hz, 2H), 2.49-2.33 (m, 2H), 2.22-2.09 (m, 6H), 1.93-1.86 (m, 2H). MS(ESI):m / z 485.2[M+H] + .

[0247] Example E39: 7-(4-chlorobenzyl)-8-(1-fluoro-4-methylcyclohexyl)-1-(3-hydroxypropyl)-3-methyl-3,7-dihydro-1H-purine-2,6-dione

[0248] [ka]

[0249] The synthesis method of Example E39 is the same as that of E21, except that 4-methylcyclohexanone is used as the starting material instead of 4-(trifluoromethyl)cyclohexanone. 1H NMR(500MHz,Chloroform-d,8:1 isomer)δ 7.31(d,J=8.5Hz,2H),7.10 / 7.16(2×d,J=8.2Hz,2H),5.75 / 5.76(2×s,2H),4.26-4.08(m,2H),3.62 / 3.60(2×s,3H),3.51(t,J=5 .5Hz,2H),3.45-3.27(m,1H),2.41-2.24(m,2H),2.12-1.97(m,1H),1.93-1.75(m,6H),1.52-1.42(m,2H),1.02(d,J=6.7Hz,3H). MS(ESI):m / z 463.2[M+H] + .

[0250] Example E40: 4-(7-(4-chlorobenzyl)-1-(3-hydroxypropyl)-3-methyl-2,6-dioxo-2,3,6,7-tetrahydro-1H-purin-8-yl)-4-fluorocyclohexane-1-carbonitrile

[0251] [ka]

[0252] The synthesis method of Example E40 is similar to that of E21, except that 4-oxocyclohexanecarbonitrile is used as the starting material instead of 4-(trifluoromethyl)cyclohexanone. 1 H NMR(500MHz,Chloroform-d,7:10 isomer)δ 7.33 / 7.31(2×d,J=8.6Hz,2H),7.14 / 7.12(2×d,J=8.2Hz,2H),5.76(s,2H),4.19(t,J=6.0Hz,2H),3.58 / 3.62(2 ×s,3H),3.53(t,J=5.5Hz,2H),3.06(p,J=3.9Hz,1H),2.52-2.36(m,1H),2.18-2.04(m,5H),2.03-1.99(m,2H). 1.91-1.83(m,2H). MS(ESI):m / z 474.2[M+H] + .

[0253] Example E41: 7-(4-chlorobenzyl)-1-(3-hydroxypropyl)-3-methyl-8-(4-(trifluoromethyl)cyclohex-1-en-1-yl)-3,7-dihydro-1H-purine-2,6-dione

[0254] [ka]

[0255] The synthesis method of Example E41 is the same as that of E29, except that INT 21-1 is used as the starting material instead of INT 28-1. 1 H NMR(500MHz,Chloroform-d)δ 7.33(d,J=8.5Hz,2H),7.03(d,J=8.5Hz,2H),6.08-6.04(m,1H),5.65-5.47(m,2H),4.18(t,J=6.0Hz,2H),3.62(s,3H),3.52(t,J=5.5Hz, 2H),3.42-3.34(m,1H),2.67-2.59(m,1H),2.54-2.44(m,1H),2.44-2 .24(m,3H),2.20-2.12(m,1H),1.91-1.83(m,2H),1.75-1.70(m,1H). MS(ESI): m / z 497.15[M+H] + .

[0256] Example E42: 7-(4-chlorobenzyl)-1-(3-hydroxypropyl)-3-isopropyl-8-(1,4,4-trifluorocyclohexyl)-3,7-dihydro-1H-purine-2,6-dione

[0257] [ka]

[0258] The synthesis method of Example E42 is the same as that of E38, except that INT 14-3 is used as the starting material instead of INT 4-2. 1H NMR(500MHz,Chloroform-d)δ 7.33(d,J=8.5Hz,2H),7.17(d,J=8.2Hz,2H),5.77(s,2H),5.19(hept,J=6.8Hz,1H),4.17(t,J=6.0Hz,2H),3.57- 3.48(m,2H),3.45-3.36(m,1H),2.54-2.24(m,2H),2.21-2.03(m,6H),1.91-1.83(m,2H)),1.59(d,J=7.0Hz,6H). MS(ESI):m / z 513.2[M+H] + .

[0259] Example E43: 7-(4-chloro-3-fluorobenzyl)-1-(3-hydroxypropyl)-3-methyl-8-(1,4,4-trifluorocyclohexyl)-3,7-dihydro-1H-purine-2,6-dione

[0260] [ka]

[0261] The synthesis method of Example E43 is the same as that of E38, except that INT 43-3 is used as a raw material instead of INT 4-2, and the synthesis method of INT 43-3 is the same as that of INT 4-2, except that 4-chloro-3-fluorobenzyl bromide is used as a raw material instead of 4-chlorobenzyl bromide. 1 H NMR(500MHz,Chloroform-d)δ 7.41-7.34(m,2H),7.01-6.90(m,2H),5.76(s,2H),4.19(t,J=6.0Hz,2H),3.59(s,3H),3 .56-3.48(m,2H),3.28(s,1H),2.55-2.32(m,2H),2.25-2.08(m,6H),1.92-1.84(m,2H). MS(ESI):m / z 503.1[M+H] + .

[0262] Example E44: 7-(3,4-difluorophenyl)-1-(3-hydroxypropyl)-3-methyl-8-(1,4,4-trifluorocyclohexyl)-3,7-dihydro-1H-purine-2,6-dione

[0263] [ka]

[0264] The synthesis method of Example E44 is similar to that of E43, except that 3,4-difluorobenzyl bromide is used as the starting material instead of 4-chloro-3-fluorobenzyl bromide. 1 H NMR(500MHz,Chloroform-d)δ 7.16-7.09(m,1H),7.03(ddd,J=10.5,7.5,2.3Hz,1H),6.99-6.95(m,1H),5.72(s,2H),4.17(t,J=6.5Hz,2 H),3.57(s,3H),3.55-3.47(m,2H),3.28(s,1H),2.49-2.31(m,2H),2.23-2.05(m,6H),1.93-1.84(m,2H). MS(ESI):m / z 487.2[M+H] + .

[0265] Example E45: 7-(4-Fluorobenzyl)-1-(3-hydroxypropyl)-3-methyl-8-(1,4,4-trifluorocyclohexyl)-3,7-dihydro-1H-purine-2,6-dione

[0266] [ka]

[0267] The synthesis method of Example E45 is similar to that of E43, except that 4-fluorobenzyl bromide is used as the starting material instead of 4-chloro-3-fluorobenzyl bromide. 1H NMR(500MHz,Chloroform-d)δ 7.23-7.13(m,2H),7.04-6.97(m,2H),5.75(s,2H),4.17(t,J=6Hz,2H),3.57(s,3H),3. 53-3.46(m,2H),3.33(s,1H),2.47-2.27(m,2H),2.19-2.04(m,6H),1.92-1.83(m,2H). MS(ESI):m / z 469.2[M+H] + .

[0268] Example E46: 7-(4-Fluorobenzyl)-1-(3-hydroxypropyl)-3-isopropyl-8-(1,4,4-trifluorocyclohexyl)-3,7-dihydro-1H-purine-2,6-dione

[0269] [ka]

[0270] The synthesis of Example E46 is similar to that of E45, except that 2-iodopropane is used as the starting material instead of iodomethane. 1 H NMR(500MHz,Chloroform-d)δ 7.24-7.18(m,2H),7.06-6.98(m,2H),5.75(s,2H),5.16(h,J=6.9Hz,1H),4.16(t,J=6.0Hz,2H),3.54-3.47( m,2H),3.40(t,J=7.1Hz,1H),2.46-2.27(m,2H),2.20-2.06(m,6H),1.91-1.83(m,2H),1.56(d,J=7.0Hz,6H). MS(ESI):m / z 497.2[M+H] + .

[0271] Example E47: 7-(3,4-difluorophenyl)-1-(3-hydroxypropyl)-3-isopropyl-8-(1,4,4-trifluorocyclohexyl)-3,7-dihydro-1H-purine-2,6-dione

[0272] [ka]

[0273] The synthesis method of Example E47 is the same as that of E46, except that INT 47-2 is used as the starting material instead of INT 46-2. 1 H NMR(500MHz,Chloroform-d)δ 7.19-7.12(m,1H),7.07(ddd,J=10.6,7.5,2.2Hz,1H),7.04-6.99(m,1H),5.74(s,2H),5.19(hept,J=6.9Hz,1H),4.17(t,J= 6Hz,2H), 3.53(t,J=5.5Hz,2H),3.40(s,1H),2.50-2.31(m,2H),2.28-2.07(m,6H),1.94-1.84(m,2H),1.59(d,J=6.9Hz,6H). MS(ESI):m / z 515.2[M+H] + .

[0274] Example E48: 7-(4-chlorobenzyl)-8-(4-(dimethylamino)-1-fluorocyclohexyl)-1-(3-hydroxypropyl)-3-methyl-3,7-dihydro-1H-purine-2,6-dione

[0275] [ka]

[0276] The synthesis method of Example E48 is the same as that of E21, except that 4-dimethylaminocyclohexanone is used as the starting material instead of 4-(trifluoromethyl)cyclohexanone. 1 H NMR(500MHz,Chloroform-d)δ 7.28(d,J=8.5Hz,2H),7.09(d,J=8.4Hz,2H),5.74(s,2H),4.15(t,J=6.0Hz,2H),3.59(s,3H),3.5 2-3.46(m,2H),3.36(s,1H),2.48-2.32(m,2H),2.27(s,6H),2.22-2.16(m,1H),1.94-1.79(m,8H). MS(ESI):m / z 492.2[M+H] +.

[0277] Example E49: 7-(4-chlorobenzyl)-8-(4-(dimethylamino)cyclohex-1-en-1-yl)-1-(3-hydroxypropyl)-3-methyl-3,7-dihydro-1H-purine-2,6-dione

[0278] [ka]

[0279] The synthesis method of Example E49 is the same as that of E29, except that INT 48-1 is used as the starting material instead of INT 28-1. 1 H NMR(500MHz,Chloroform-d)δ 7.32(d,J=8.5Hz,2H),6.99(d,J=8.3Hz,2H),5.98-5.94(m,1H),5.66-5.46(m,2H),4.15(t,J=6.1Hz,2H),3.59(s,3H),3.50(t, J=5.5Hz,3H),3.44-3.33(m,1H),2.89-2.81(m,6H),2.73-2.63(m,1H),2.57-2.42(m,5H),2.35-2.27(m,1H),1.91-1.81(m,2H). MS(ESI):m / z 472.2[M+H] + .

[0280] Example E50: 7-(4-chlorobenzyl)-1-(3-hydroxypropyl)-3-methyl-8-(4-morpholinocyclohex-1-en-1-yl)-3,7-dihydro-1H-purine-2,6-dione

[0281] [ka]

[0282] The synthesis method of Example E50 is the same as that of Example E6, except that 4-(4-(4,4,5,5-tetramethyl-1,3,2-dioxaborolan-2-yl)-3-cyclohexene)morpholine is used as the starting material instead of 4,4,5,5-tetramethyl-2-(1,2,3,6-tetrahydro-[1,1'-biphenyl]-4-yl)-1,3,2-dioxaborolane. 1 H NMR(500MHz,Chloroform-d)δ 7.30(d,J=8.5Hz,2H),7.05(d,J=8.3Hz,2H),6.05-6.01(m,1H),5.61-5. 47(m,2H),4.20-4.12(m,2H),3.78-3.72(m,4H),3.60(s,3H),3.54-3.46 (m,2H),3.41(t,J=7.1Hz,1H),2.66-2.50(m,6H),2.45-2.30(m,2H),2.2 7-2.17(m,1H),2.14-2.07(m,1H),1.91-1.84(m,2H),1.57-1.53(m,1H). MS(ESI): m / z 514.2[M+H] + .

[0283] Example E51: 7-(4-chloro-3-fluorobenzyl)-3-ethyl-8-(1-fluoro-4-(trifluoromethyl)cyclohexyl)-1-(3-hydroxypropyl)-3,7-dihydro-1H-purine-2,6-dione

[0284] [ka]

[0285] The synthesis of Example E51 is similar to that of E43, except that 2-iodopropane is used as the starting material instead of iodoethane. 1H NMR(500MHz,Chloroform-d,5:11 isomer)δ 7.41-7.34(m,1H),7.03-6.90(m,2H),5.75 / 5.73(2×s,2H),4.28-4.07(m,4H),3.60-3.46(m,2H),2.4 9-2.39(m,1H),2.35-2.09(m,2H),2.07-1.82(m,7H),1.81-1.74(m,1H),1.37 / 1.36(2×t,J=7Hz,3H). MS(ESI):m / z 549.2[M+H] + .

[0286] Example E52: 7-(3,4-difluorophenyl)-8-(1-fluoro-4-(trifluoromethyl)cyclohexyl)-1-(3-hydroxypropyl)-3,7-dihydro-1H-purine-2,6-dione

[0287] [ka]

[0288] The synthesis method of Example E52 is the same as that of E21, except that INT 44-3 is used as the starting material instead of INT 4-2. 1 H NMR(500MHz,Chloroform-d,9:13 isomer)δ 7.16-7.08(m,1H),7.07-6.88(m,2H),5.70 / 5.72(2×s,2H),4.17 / 4.16(2×t,J=7.5Hz,2H),3.57(2×s,3H),3.54-3.47(m ,2H),3.35-3.26(m,1H),2.46-2.36(m,1H),2.32-2.19(m,1H),2.18-2.11(m,1H),2.05-1.82(m,7H),1.82-1.71(m,1H). MS(ESI):m / z 519.2[M+H] + .

[0289] Example E53: 7-(4-chlorobenzyl)-8-(1-fluoro-4-(trifluoromethyl)cyclohexyl)-1-(3-hydroxypropyl)-3-isopropyl-3,7-dihydro-1H-purine-2,6-dione

[0290] [ka]

[0291] The synthesis method of Example E53 is similar to that of E42, except that 4-(trifluoromethyl)cyclohexanone is used as the starting material instead of 4,4-difluorocyclohexanone. 1 H NMR(500MHz,Chloroform-d,8:1 isomer)δ 7.32(d,J=8.5Hz,2H),7.19 / 7.13(2×d,J=8.2Hz,2H),5.76(s,2H),5.25-5.12(m,1H),4.17 / 4.16(2×t,J=6Hz,2H),3.58-3.49(m,2H),3 .48-3.39(m,1H),2.32-2.11(m,4H),2.11-2.00(m,1H),2.00-1.92(m,2H),1.91-1.85(m,2H),1.84-1.72(m,2H),1.58(d,J=7.0Hz,6H). MS(ESI): m / z 545.2[M+H] + .

[0292] Example E54: 7-(4-chlorobenzyl)-8-(1-fluoro-4-phenylcyclohexyl)-1-(3-hydroxypropyl)-3-methyl-3,7-dihydro-1H-purine-2,6-dione

[0293] [ka]

[0294] The synthesis method of Example E54 is similar to that of E21, except that 4-phenylcyclohexanone is used as the starting material instead of 4-(trifluoromethyl)cyclohexanone. 1H NMR(500MHz,Chloroform-d,3:1 isomer)δ 7.38-7.30(m,4H),7.28-7.22(m,3H),7.19 / 7.09(2×d,J=8.2Hz,2H),5.79 / 5.77(2×s,2H),4.22-4.15(m,2H),3.61 / 3.65(2 ×s,3H),3.56-3.49(m,2H),2.83-2.64(m,1H),2.51-2.43(m,1H),2.35-2.24(m,1H),2.22-2.15(m,2H),1.99-1.86(m,6H). MS(ESI):m / z 525.2[M+H] + .

[0295] Example E55: 7-(4-chlorobenzyl)-8-(1-fluoro-4-(pyrrolidin-1-yl)cyclohexyl)-1-(3-hydroxypropyl)-3-methyl-3,7-dihydro-1H-purine-2,6-dione

[0296] [ka]

[0297] The synthesis method of Example E55 is similar to that of E21, except that 4-(1-pyrrolidinyl)cyclohexanone is used as the starting material instead of 4-(trifluoromethyl)cyclohexanone. 1 H NMR(500MHz,Chloroform-d)δ 7.31(d,J=8.5Hz,2H),7.08(d,J=8.2Hz,2H),5.73(s,2H),4.17(t,J=5.9Hz,2H),3.60(s,3H),3.51(t,J=5.5Hz,2H),3.09-2.99(m,1H) ,2.87-2.75(m,2H),2.64-2.51(m,2H),2.35-2.27(m,2H),2.21-2.11(m,4H),2.09-2.01(m,4H),1.99-1.92(m,2H),1.93-1.85(m,2H). MS(ESI): m / z 518.2[M+H] + .

[0298] Example E56: 7-(4-chloro-3-fluorobenzyl)-1-(3-hydroxypropyl)-3-isopropyl-8-(1,4,4-trifluorocyclohexyl)-3,7-dihydro-1H-purine-2,6-dione

[0299] [ka]

[0300] The synthesis method of Example E56 is the same as that of E47, except that INT 51-1 is used as the starting material instead of INT 44-1. 1 H NMR(500MHz,Chloroform-d)δ 7.39-7.35(m,1H),7.05-6.95(m,2H),5.75(s,2H),5.19(hept,J=7.0Hz,1H),4.17(t,J=6.0Hz,2H),3 .53(t,J=5.5Hz,2H),2.51-2.30(m,2H),2.24-2.11(m,6H),1.93-1.86(m,2H),1.59(d,J=6.9Hz,6H). MS(ESI):m / z 531.2[M+H] + .

[0301] Example E57: 7-(4-chloro-3-fluorobenzyl)-3-ethyl-1-(3-hydroxypropyl)-8-(4-(trifluoromethyl)cyclohex-1-en-1-yl)-3,7-dihydro-1H-purine-2,6-dione

[0302] [ka]

[0303] The synthesis method of Example E57 is the same as that of E29, except that INT 51-4 is used as the starting material instead of INT 28-1. 1H NMR(500MHz,Chloroform-d)δ 7.42-7.37(m,1H),6.88(ddd,J=12.3,8.9,2.1Hz,2H),6.05(s,1H),5.63-5.49(m,2H),4.28-4.09(m,4H),3.52(t ,J=5.5Hz,2H),2.72-2.63(m,1H),2.55-2.26(m,4H),2.23-2.13(m,1H),1.96-1.83(m,3H),1.39(t,J=7.0Hz,3H). MS(ESI):m / z 529.2[M+H] + .

[0304] Example E58: 7-(3,4-difluorophenyl)-1-(3-hydroxypropyl)-3-methyl-8-(4-(trifluoromethyl)cyclohex-1-en-1-yl)-3,7-dihydro-1H-purine-2,6-dione

[0305] [ka]

[0306] The synthesis method of Example E58 is the same as that of E29, except that INT 52-1 is used as the starting material instead of INT 28-1. 1 H NMR(500MHz,Chloroform-d)δ 7.21-7.12(m,1H),6.92(ddd,J=10.3,7.4,2.3Hz,1H),6.88-6.83(m,1H),6.10-6.04(m,1H),5.64-5.49(m,2H),4.27-4.12(m,2H), 3.62(s,3H),3.57-3.47(m,2H),3.38(t,J=7.0Hz,1H),2.69-2.61(m,1H),2.56-2.27(m,4H),2.23-2.13(m,1H),1.94-1.86(m,3H). MS(ESI):m / z 499.2[M+H] + .

[0307] Example E59: 8-(1-fluoro-4-(trifluoromethyl)cyclohexyl)-7-(4-fluorobenzyl)-1-(3-hydroxypropyl)-3-methyl-3,7-dihydro-1H-purine-2,6-dione

[0308] [ka]

[0309] The synthesis method of Example E59 is the same as that of E45, except that 4-(trifluoromethyl)cyclohexanone is used as the starting material instead of 4,4-difluorocyclohexanone. 1 H NMR(500MHz,Chloroform-d,3:1 isomer)δ 7.23 / 7.16(2×m,2H),7.06-7.00(m,2H),5.77 / 5.75(2×s,2H),4.23-4.15(m,2H),3.58 / 3.61(2×s,3H),3.56-3.49(m,2 H),3.39-3.30(m,1H),2.48-2.35(m,1H),2.31-2.09(m,3H),2.08-2.01(m,1H),2.00-1.85(m,5H),1.83-1.74(m,1H). MS(ESI):m / z 501.2[M+H] + .

[0310] Example E60: 1-(3-hydroxypropyl)-3-methyl-8-(1,4,4-trifluorocyclohexyl)-7-(4-(trifluoromethyl)benzyl)-3,7-dihydro-1H-purine-2,6-dione

[0311] [ka]

[0312] 8-Bromo-3-methyl-3,7-dihydro-purine-2,6-dione (2 g, 8.13 mmol) was dissolved in 10 mL of DMF, and KHCO (1.298 g, 12.987 mmol) and 4-(trifluoromethyl)benzyl bromide (1.8 g, 7.531 mmol) were added, followed by stirring at 60 °C for 2 hours. When TLC showed the reaction was complete, 10 mL of H2O was added, and the mixture was suction filtered and dried to obtain 2.5 g of a white solid, intermediate INT 60-1. INT 60-1 (350 mg, 0.87 mmol) was dissolved in 5 mL of DMF, and KCO (244 mg, 1.768 mmol) and 2-(3-bromopropoxy)tetrahydro-2H-pyran (236.5 mg, 1.06 mmol) were added, followed by stirring at 60 °C for 2 hours. TLC showed the reaction was complete. The reaction mixture was poured into saturated aqueous NaCl solution, extracted three times with ethyl acetate, and the combined organic phases were washed three times with saturated aqueous NaCl solution. After drying over anhydrous MgSO4, the mixture was separated on a flash silica gel chromatography column, eluted with MeOH / DCM as the mobile phase, and the desired product was collected to obtain 300 mg of a colorless oily liquid, i.e., intermediate INT 60-2.

[0313] The subsequent synthesis of Example E60 is similar to that of E38, except that INT 60-2 is used as the starting material instead of 4-2. 1 H NMR(500MHz,Chloroform-d)δ 7.59(d,J=8.1Hz,2H),7.29-7.24(m,2H),5.84(s,2H),4.15(t,J=6.0Hz,2H),3.58(s,3H) ,3.53-3.48(m,2H),3.26(s,1H),2.51-2.31(m,2H),2.22-2.06(m,6H),1.91-1.83(m,2H). MS(ESI):m / z 519.2[M+H] + .

[0314] Example E61: 7-(3,5-difluorobenzyl)-1-(3-hydroxypropyl)-3-methyl-8-(1,4,4-trifluorocyclohexyl)-3,7-dihydro-1H-purine-2,6-dione

[0315] [ka]

[0316] The synthesis method of Example E61 is the same as that of E60, except that 3,5-difluorobenzyl bromide is used as the starting material instead of 4-(trifluoromethyl)benzyl bromide. 1 H NMR(500MHz,Chloroform-d)δ 6.74(tt,J=2.3,8.7Hz,1H),6.70-6.64(m,2H),5.75(s,2H),4.16(t,J=6.1Hz,2H),3.58(s,3H),3 .54-3.48(m,2H),3.21(t,J=7.0Hz,1H),2.51-2.32(m,2H),2.21-2.06(m,6H),1.93-1.83(m,2H). MS(ESI):m / z 487.2[M+H] + .

[0317] Example E62: 7-benzyl-1-(3-hydroxypropyl)-3-methyl-8-(1,4,4-trifluorocyclohexyl)-3,7-dihydro-1H-purine-2,6-dione

[0318] [ka]

[0319] The synthesis method of Example E62 is similar to that of E60, except that benzyl bromide is used as the starting material instead of 4-(trifluoromethyl)benzyl bromide. 1 H NMR(500MHz,Chloroform-d)δ 7.36-7.28(m,3H),7.18-7.11(m,2H),5.80(s,2H),4.17(t,J=6.0Hz,2H),3.57(s ,3H),3.54-3.48(m,2H),2.43-2.26(m,2H),2.17-2.03(m,6H),1.91-1.84(m,2H). MS(ESI):m / z 451.2[M+H] + .

[0320] Example E63: 7-benzyl-8-(1-fluoro-4-(trifluoromethyl)cyclohexyl)-1-(3-hydroxypropyl)-3-methyl-3,7-dihydro-1H-purine-2,6-dione

[0321] [ka]

[0322] The synthesis method of Example E63 is similar to that of E62, except that 4-(trifluoromethyl)cyclohexanone is used as the starting material instead of 4,4-difluorocyclohexanone. 1 H NMR(500MHz,Chloroform-d)δ 7.36-7.28(m,3H),7.13-7.06(m,2H),5.79(s,2H),4.15(t,J=6.0Hz,2H),3.60(s,3H),3.52-3.45(m,2 H),3.33(t,J=7.1Hz,1H),2.45-2.34(m,2H),2.29-2.18(m,1H),1.99-1.91(m,4H),1.89-1.82(m,4H). MS(ESI):m / z 483.2[M+H] + .

[0323] Example E64: 7-(2,4-Difluorobenzyl)-1-(3-hydroxypropyl)-3-methyl-8-(1,4,4-trifluorocyclohexyl)-3,7-dihydro-1H-purine-2,6-dione

[0324] [ka]

[0325] The synthesis method of Example E64 is the same as that of E60, except that 2,4-difluorobenzyl bromide is used as the starting material instead of 4-(trifluoromethyl)benzyl bromide. 1H NMR(500MHz,Chloroform-d)δ 6.89-6.84(m,1H),6.83-6.73(m,2H),5.82(s,2H),4.16(t,J=6.0Hz,2H),3.58(s ,3H),3.53-3.47(m,2H),2.47-2.30(m,2H),2.19-2.04(m,6H),1.91-1.83(m,2H). MS(ESI):m / z 487.2[M+H] + .

[0326] Example E65: 7-(2,4-difluorobenzyl)-8-(1-fluoro-4-(trifluoromethyl)cyclohexyl)-1-(3-hydroxypropyl)-3-methyl-3,7-dihydro-1H-purine-2,6-dione

[0327] [ka]

[0328] The synthesis method of Example E65 is the same as that of E64, except that 4-(trifluoromethyl)cyclohexanone is used as the starting material instead of 4,4-difluorocyclohexanone. 1 H NMR(500MHz,Chloroform-d)δ 6.89-6.83(m,1H),6.82-6.75(m,1H),6.72-6.64(m,1H),5.79(s,2H),4.14(t,J=6.0Hz,2H ),3.60(s,3H),3.52-3.44(m,2H),2.45-2.33(m,2H),2.31-2.21(m,1H),2.02-1.82(m,8H). MS(ESI):m / z 519.2[M+H] + .

[0329] Example E66: 7-(2-chloro-4-fluorobenzyl)-1-(3-hydroxypropyl)-3-methyl-8-(1,4,4-trifluorocyclohexyl)-3,7-dihydro-1H-purine-2,6-dione

[0330] [ka]

[0331] The synthesis method of Example E66 is the same as that of E60, except that 2-chloro-4-fluorobenzyl bromide is used as the starting material instead of 4-(trifluoromethyl)benzyl bromide. 1 H NMR(500MHz,Chloroform-d)δ 7.19(dd,J=2.6,8.2Hz,1H),6.92-6.85(m,1H),6.47-6.42(m,1H),5.82(s,2H),4.13(t,J=6.0H z,2H),3.60(s,3H),3.52-3.45(m,2H),2.50-2.31(m,2H),2.21-2.05(m,6H),1.89-1.82(m,2H). MS(ESI):m / z 503.1[M+H] + .

[0332] Example E67: 7-(2-chloro-4-fluorobenzyl)-8-(1-fluoro-4-(trifluoromethyl)cyclohexyl)-1-(3-hydroxypropyl)-3-methyl-3,7-dihydro-1H-purine-2,6-dione

[0333] [ka]

[0334] The synthesis method of Example E67 is the same as that of E66, except that 4-(trifluoromethyl)cyclohexanone is used as the starting material instead of 4,4-difluorocyclohexanone. 1 H NMR(500MHz,Chloroform-d)δ 7.19(dd,J=2.6,8.2Hz,1H),6.92-6.84(m,1H),6.47-6.42(m,1H),5.80(s,2H),4.12(t,J=6.0Hz,2H),3.62(s,3 H),3.52-3.44(m,2H),3.17(s,1H),2.48-2.37(m,2H),2.30-2.19(m,1H),2.01-1.88(m,6H),1.87-1.82(m,2H). MS(ESI):m / z 535.1[M+H]+ .

[0335] Example E68: 7-(3-chloro-4-fluorobenzyl)-1-(3-hydroxypropyl)-3-methyl-8-(1,4,4-trifluorocyclohexyl)-3,7-dihydro-1H-purine-2,6-dione

[0336] [ka]

[0337] The synthesis method of Example E68 is the same as that of E60, except that 3-chloro-4-fluorobenzyl bromide is used as the starting material instead of 4-(trifluoromethyl)benzyl bromide. 1 H NMR(500MHz,Chloroform-d)δ 7.29-7.26(m,1H),7.10(d,J=6.9Hz,2H),5.72(s,2H),4.17(t,J=6.0Hz,2H),3.57( s, 3H), 3.54-3.47 (m, 2H), 2.49-2.32 (m, 2H), 2.24-2.07 (m, 6H), 1.92-1.84 (m, 2H). MS(ESI):m / z 503.1[M+H] + .

[0338] Example E69: 7-(3-chloro-4-fluorobenzyl)-8-(1-fluoro-4-(trifluoromethyl)cyclohexyl)-1-(3-hydroxypropyl)-3-methyl-3,7-dihydro-1H-purine-2,6-dione

[0339] [ka]

[0340] The synthesis method of Example E69 is the same as that of E68, except that 4-(trifluoromethyl)cyclohexanone is used as the starting material instead of 4,4-difluorocyclohexanone. 1H NMR(500MHz,Chloroform-d)δ 7.25-7.21(m,1H),7.13-7.04(m,2H),5.69(s,2H),4.16(t,J=6.0Hz,2H),3.59(s,3H),3.53-3.47(m,2 H),3.25(t,J=7.0Hz,1H),2.48-2.35(m,2H),2.32-2.21(m,1H),2.11-1.96(m,4H),1.94-1.83(m,4H). MS(ESI):m / z 535.1[M+H] + .

[0341] Example E70: 7-(4-chloro-2-fluorobenzyl)-1-(3-hydroxypropyl)-3-methyl-8-(1,4,4-trifluorocyclohexyl)-3,7-dihydro-1H-purine-2,6-dione

[0342] [ka]

[0343] The synthesis method of Example E70 is the same as that of E60, except that 4-chloro-2-fluorobenzyl bromide is used as the starting material instead of 4-(trifluoromethyl)benzyl bromide. 1 H NMR(500MHz,Chloroform-d)δ 7.17-7.12(m,1H),7.08-7.03(m,1H),6.71-6.65(m,1H),5.82(s,2H),4.14(t,J=6.0Hz,2H ),3.58(s,3H),3.53-3.47(m,2H),2.48-2.31(m,2H),2.19-2.05(m,6H),1.90-1.82(m,2H). MS(ESI):m / z 503.1[M+H] + .

[0344] Example E71: 7-(4-chloro-2-fluorobenzyl)-8-(1-fluoro-4-(trifluoromethyl)cyclohexyl)-1-(3-hydroxypropyl)-3-methyl-3,7-dihydro-1H-purine-2,6-dione

[0345] [ka]

[0346] The synthesis method of Example E71 is the same as that of E70, except that 4-(trifluoromethyl)cyclohexanone is used as the starting material instead of 4,4-difluorocyclohexanone. 1 H NMR(500MHz,Chloroform-d)δ 7.13(dd,J=2.1,9.8Hz,1H),7.08-7.03(m,1H),6.71-6.66(m,1H),5.82(s,2H),4.15(t,J=6.0Hz,2H),3.58(s,3H) ,3.51-3.46(m,2H),2.26-2.10(m,4H),2.09-2.01(m,1H),1.97-1.91(m,2H),1.90-1.83(m,2H),1.79-1.67(m,2H). MS(ESI):m / z 535.1[M+H] + .

[0347] Example E72: 4-((1-(3-hydroxypropyl)-3-methyl-2,6-dioxo-8-(1,4,4-trifluorocyclohexyl)-1,2,3,6-tetrahydro-7H-purin-7-yl)methyl)benzonitrile

[0348] [ka]

[0349] The synthesis method of Example E72 is the same as that of E60, except that 4-cyanobenzyl bromide is used as the starting material instead of 4-(trifluoromethyl)benzyl bromide. 1H NMR(500MHz,Chloroform-d)δ 7.63(d,J=8.2Hz,2H),7.25(d,J=8.7Hz,2H),5.83(s,2H),4.15(t,J=6.1Hz,2H),3.58(s,3H) ),3.52-3.44(m,2H),3.17(s,1H),2.50-2.32(m,2H),2.20-2.03(m,6H),1.91-1.81(m,2H). MS(ESI):m / z 476.2[M+H] + .

[0350] Example E73: 4-((8-(1-fluoro-4-(trifluoromethyl)cyclohexyl)-1-(3-hydroxypropyl)-3-methyl-2,6-dioxo-1,2,3,6-tetrahydro-7H-purin-7-yl)methyl)benzonitrile

[0351] [ka]

[0352] The synthesis method of Example E73 is similar to that of E72, except that 4-(trifluoromethyl)cyclohexanone is used as the starting material instead of 4,4-difluorocyclohexanone. 1 H NMR(500MHz,Chloroform-d)δ 7.62(d,J=8.0Hz,2H),7.22(d,J=8.0Hz,2H),5.81(s,2H),4.13(t,J=6.0Hz,2H),3.60(s,3H) ),3.52-3.47(m,2H),3.19(s,1H),2.47-2.36(m,2H),2.33-2.21(m,1H),2.04-1.80(m,8H). MS(ESI):m / z 508.2[M+H] + .

[0353] 2. Pharmacological Examples 1. Evaluation of ion channel activity 1.1.TRPC3 / 4 / 5 inhibitory activity test HEK-293 cells expressing hTRPC3, hTRPC4, or hTRPC5 were plated at 2 × 10 per well in polylysine (PDL)-coated black-bottom 96-well plates. 4 After inoculation with cells and culturing for 8 hours, the original medium was discarded, and immediately after that, 60 μL of Fluo-4 / AM dye was added at a final concentration of 4 μM and incubated at 37°C for 60 minutes. After that, the cells were washed five times with calcium flux assay buffer, and the cell plates were transferred to a FLIPR tube preheated to 30°C. (登録商標)TETRA The fluorescent signal was continuously recorded at a sampling rate of 1 s in the range of 515–535 nm with excitation at 488 nm (Molecular Devices, Sunnyvale, CA, USA). After 60 s of recording, the solvent control, test compound, and positive inhibitor HC608 (compound C31 in WO2014143799, final concentration 100 nM) were added, and the signal was continuously collected for 300 s. Subsequently, the agonist Englerin A (EA) (MedChemExpress, Shanghai, China, final concentration 0.3 nM) was added, and the fluorescent signal was continuously collected for 600 s. The fluorescent signal in the trace plot was expressed as F / F, where F is the fluorescent signal at various time points, and F is the basal fluorescent signal, i.e., the average value of the fluorescent signal at the first 10 time points. In the dose-effect plot, F / F0 = 1 was used as the baseline, and the area under the curve of the change in fluorescence intensity after the addition of EA was calculated. The area under the curve was then combined with the logarithm of the compound concentration, and the IC was calculated using the log[Inhibitor] vs. response-variable slope. 50 Calculate the value.

[0354] 1.2.TRPC5 agonist activity test HEK-293 cells expressing hTRPC5 were plated at 2 x 10 cells per well in a PDL-coated black-bottom 96-well plate. 4After inoculation with cells and culturing for 8 hours, the original medium was discarded, and immediately after that, 60 μL of Fluo-4 / AM dye was added at a final concentration of 4 μM and incubated at 37°C for 60 minutes. After that, the cells were washed five times with calcium flux assay buffer, and the cell plates were transferred to a FLIPR tube preheated to 30°C. (登録商標)TETRA The fluorescent signal was continuously recorded at a sampling rate of 1 s in the range of 515–535 nm with excitation at 488 nm (Molecular Devices, Sunnyvale, CA, USA). After 360 s of recording, the solvent control and test compound were added, and the fluorescent signal was continuously collected for 600 s. The fluorescent signal in the trace plot was expressed as F / F, where F is the fluorescent signal at various time points, and F is the basal fluorescent signal, i.e., the average value of the fluorescent signal at the first 10 time points. In the dose-effect plot, F / F = 1 was used as the baseline, and the area under the curve of the change in fluorescence intensity after the addition of the test compound was calculated. The logarithmic value of the area under the curve and the compound concentration were combined, and the EC was calculated using the logarithmic slope of the agonist vs. response variable. 50 Calculate the value.

[0355] 1.3. hERG channel blocking effect test (1)Cell preparation CHO cells expressing hERG protein were cultured at 175 cm 2 When the cell density reached 60-80%, the culture medium was removed, the cells were washed once with 7 mL of PBS, and then 3 mL of Detachin was added for digestion. After digestion was complete, the cells were added to 7 mL of culture medium for neutralization, followed by centrifugation. The supernatant was removed by aspiration, and 5 mL of culture medium was added to resuspend the cells. The cells were then allowed to grow until the cell density reached 2-5 × 10. 6 / mL.

[0356] (2) Solution preparation

[0357] [Table 1]

[0358] (3) Electrophysiological recording process The Qpatch instrument automatically performed single-cell high-impedance sealing and whole-cell patterning. After acquiring the whole-cell recording mode, the cells were clamped at -80 mV and subjected to a 5-second +20 mV depolarizing stimulus. A 50-ms pre-voltage of -50 mV was applied, followed by a 5-second repolarization to -50 mV and then a 2-minute return to -80 mV. This voltage stimulus was applied every 15 seconds, and after 2 minutes of recording, extracellular solution was added and recording continued for 2 minutes. The dosing process then began, with compound concentrations being administered for 2 minutes, starting with the lowest test concentration. After all concentrations had been administered, 10 μM of the positive control compound Cisapride was administered. At least three cells were tested for each concentration (n ≥ 3).

[0359] (4) Preparation of compounds The compound stock solution was diluted with extracellular solution, and 5 μL of 20 mM compound stock solution was added to 2495 μL of extracellular solution, followed by a 500-fold dilution to 40 μM. The compound was then serially diluted 3-fold in 0.2% DMSO-containing extracellular solution to the final test concentration. The highest test concentration of the compound was 40 μM, followed by six successive concentrations of 40, 13.33, 4.44, 1.48, 0.49, and 0.16 μM, respectively. The highest test concentration of the positive compound, Cisapride, was 3 μM, followed by six successive concentrations of 3, 1, 0.333, 0.111, 0.037, and 0.012 μM, respectively. The DMSO content in the final test concentrations did not exceed 0.2%, as this concentration of DMSO does not affect the hERG potassium channel.

[0360] (5) Data analysis: The experimental data are analyzed by XLFit software. 1.4. Liver microsome stability test First, a 0.1 M Tris buffer solution (pH 7.4) was prepared, followed by the preparation of a 100 mM MgCl2 solution and a 10 mM NADPH solution. The test compound was first prepared as a stock solution in DMSO, which was then diluted with water and 0.1% BSA to the working concentration. For measurement, liver microsomes in TRIS buffer (final concentration: 0.33 mg / mL microsomal protein), MgCl2 solution (final concentration: 5 mM), test compound solution (final concentration: 1 μM), and NADPH solution (final concentration: 1 mM) were incubated at 37°C, and the reaction was stopped by adding methanol at 0, 7, 17, 30, and 60 minutes, respectively. The residual concentration of the test compound was measured by LC / MS / MS.

[0361] Half-life calculation: T1 / 2=0.693 / k e , Intrinsic clearance calculation:

[0362]

number

[0363] In vivo clearance calculation:

[0364]

number

[0365] Calculation of hepatic clearance:

[0366]

number

[0367] Calculating metabolic utilization:

[0368]

number

[0369] k e : The slope of the regression line (absolute value of slope) when plotted on a semi-logarithmic scale. The semi-logarithm of the substrate residual rate is taken and plotted against the reaction time. P: Microsomal protein concentration (mg / mL), Houston: Houston factor (45 mg microsomal protein / g liver), LW: Liver weight (g), HBF: Hepatic blood flow (mL / min), fu: Unbound ratio (usually fu=1).

[0370] 1.5. Test Results (1) As shown in Table 1, the compounds of the present invention have moderate to strong inhibitory effects on the TRPC5 channel, and the activity of some compounds exceeds that of the reference compound HC-070. Therefore, the compounds of the present invention have potential application value in the treatment of diseases associated with excessive activation of the TRPC5 channel or overexpression of the TRPC5 protein.

[0371] [Table 2]

[0372] (2) As shown in Table 2, some compounds of the present invention have moderate agonistic activity against the TRPC5 channel. Some compounds with agonistic activity have very similar structures to other compounds with inhibitory activity; for example, compound Example 14 is different from compounds Example 15 and Example 9, and compounds Example 7 and Example 17 are different from compounds Example 8. Such subtle structural differences unexpectedly result in significant changes in biological activity. Therefore, the compounds of the present invention can be used to inhibit the TRPC5 channel and treat diseases associated with TRPC5 protein deficiency, or to activate the TRPC5 channel, causing calcium ion overload and inducing cell death, thereby treating certain types of tumors.

[0373] [Table 3]

[0374] (3) As shown in Table 3, the compounds of the present invention also have good inhibitory effects on TRPC4, which is consistent with the biological properties of the highly homologous TRPC4 and TRPC5 proteins. Therefore, the compounds of the present invention have potential application value in the treatment of diseases associated with excessive activation of the TRPC4 channel or overexpression of the TRPC4 protein.

[0375] On the other hand, compared with HC-070, the compounds of the present invention have lower inhibitory activity against the TRPC3 channel, higher relative selectivity against the TRPC5 channel, and lower off-target risk.

[0376] On the other hand, compared to HC-070, the compounds of the present invention have lower inhibitory activity against the hERG channel and higher relative selectivity against the TRPC5 channel. In the field of medicinal chemistry, hERG channel blockade is considered an indicator of cardiotoxicity risk and should be avoided as much as possible.Compared with existing publicly known technologies, the distinctive structural feature of the compounds of the present invention is that the 8-position of the xanthine is substituted with a "cycloalkyl group" such as an aryl group, an aryloxy group, an alkyl group, an alkoxy group, or a cycloalkoxy group.Unexpectedly, this structural difference significantly weakens the inhibitory effect of the compounds on the hERG channel, suggesting that the compounds of the present invention have a lower cardiotoxicity risk.

[0377] [Table 4]

[0378] 4) As shown in Table 4, the reference compound HC-070 exhibits moderate metabolic stability in human liver microsomes, but is rapidly metabolized in mouse liver microsomes and has a shorter half-life. Compared with HC-070, the compounds of the present invention have a longer half-life and higher metabolic availability, which indicates better metabolic properties and superior drug-like properties.

[0379] [Table 5]

[0380] 2. Evaluation of the inhibitory effect of compounds on hepatic stellate cell activation 2.1. Experimental Method Primary cultured mouse hepatic stellate cells (HSCs) were plated in a 96-well black-bottom plate for 24 hours, pre-incubated with 1 μM Compound E21 or 0.1% DMSO (solvent control) for 1 hour, then added TGF-β (final concentration: 10 ng / mL) and cultured for 24 hours in a CO2 incubator. The medium was then aspirated, washed three times with PBS at 37°C, fixed with 4% paraformaldehyde for 30 minutes, permeabilized with 0.15% Triton X-100 for 10 minutes, blocked with 5% BSA for 1 hour at room temperature, added anti-α-SMA (1:300) in 5% BSA, and incubated overnight at 4°C. The next day, Alexa-488 anti-rabbit fluorescent secondary antibody (1:1000) in 5% BSA was added and incubated in the dark for 1 hour at room temperature. After incubation with the secondary antibody, the cells were washed three times with PBS, Hoechst solution was added, and the cells were incubated at room temperature in the dark for 20 minutes. After incubation, the cells were washed three times with PBS and fluorescent images were taken. A FITC filter was used to capture green fluorescence, and a DAPI filter was used to capture blue fluorescence. The total number of cells per field was calculated using ImageJ software, and the number of α-SMA-positive cells was manually counted. Data were processed and analyzed using GraphPad (Version 6.01) software.

[0381] 2.2.Experimental Results As can be seen from Figure 1, under TGF-β stimulation, the proportion of α-SMA-positive hepatic stellate cells increased from 0.3±0.04% to 11.76±1.47% (P<0.01), indicating an increased degree of hepatic stellate cell activation. The TRPC4 / 5 inhibitor E21 significantly reduced the proportion of α-SMA-positive cells to 5.20±0.74% (P<0.01), significantly inhibiting the activation of hepatic stellate cells, suggesting that the compounds of the present invention are effective in treating liver diseases.

[0382] 3. Preliminary pharmacokinetic evaluation 3.1.Experimental Method Male ICR mice (Nanjing Anuokang Biotech Co., Ltd.) aged 6 to 8 weeks were housed in an SPF-grade animal care facility with a 1:1 light / dark ratio, room temperature controlled at 23±2°C, and humidity controlled at 55%. Mice were allowed free access to food and water. Test compounds were prepared in a 2:2:96 (v / v / v) DMSO / Tween 80 / 0.5% (m / v) CMC-Na aqueous solution at a concentration of 1 mg / mL. The test compounds were then administered orally to the mice at a dose of 10 mL / kg (3 mice per compound). Approximately 50 μL of blood was collected from the orbital venous plexus of the mice 1 and 4 hours after administration and placed in a centrifuge tube containing EDTA-K2 anticoagulant. The blood samples are centrifuged at 3000 rpm for 15 minutes, and the supernatant is collected to obtain plasma samples, which are then measured for drug concentrations in the plasma using liquid chromatography-tandem mass spectrometry (LC-MS / MS).

[0383] 3.2. Test Results As shown in Table 5, the compounds of the present invention had higher plasma drug concentrations at 1 hour and 4 hours after oral administration to mice compared with HC-070, preliminarily suggesting that the compounds of the present invention have superior oral absorption properties, plasma exposure, and pharmacokinetic properties.

[0384] [Table 6]

[0385] 4. Pharmacokinetics and tissue distribution evaluation in rats 4.1. Drug Preparation Intravenous administration (IV): Test drug was dissolved in 5:5:90 (v / v / v) DMSO / Solutol / physiological saline at a concentration of 0.2 mg / mL and the dose was 5 mL / kg. Oral administration (PO): Test drug was dissolved in 2:2:96 (v / v / v) DMSO / Tween 80 / 0.5% CMC-Na at a concentration of 1 mg / mL and the dose was 10 mL / kg.

[0386] 4.2.Experimental Method Male SD rats weighing 180-280 g were fasted for 12 hours before the experiment, allowed free access to water, and fed 4 hours after administration. In the IV group, rats were administered 1 mg / kg via the tail vein. Blood samples were taken at 0.083, 0.25, 0.5, 1.0, 2.0, 4.0, 6.0, 8.0, 12, and 24 hours after administration. In the PO group, rats were administered 10 mg / kg via oral gavage. Blood samples were taken at 0.25, 0.5, 1.0, 2.0, 4.0, 6.0, 8.0, 12, and 24 hours after administration. At the above time points, 0.2 mL of blood was collected from the jugular vein and placed in an anticoagulant tube containing EDTA-K2. After collection, the whole blood was temporarily stored in an ice-water bath and centrifuged at 11,000 rpm for 5 minutes within 30 minutes to separate the plasma, which was then frozen in a refrigerator at -70°C for testing. For the tissue distribution group, rats were given 10 mg / kg of benzodiazepine orally by gavage. Two hours after administration, the rats were anesthetized and sacrificed by exsanguination via the abdominal aorta. They were then immediately dissected to collect the brain, heart, liver, spleen, lungs, kidneys, colon, and muscle. A portion of the whole blood was collected and placed in an ice-water bath. The whole blood was centrifuged at 11,000 rpm for 5 minutes within 30 minutes to separate the plasma (approximately 200 μL). After collection, the tissues and plasma were stored at or below -60°C for testing. After accurately weighing each tissue, homogenize it using acetonitrile-water (1:1, v / v) as the homogenizing agent (tissue:homogenizing agent = 1:4 (g:mL)). After homogenization, sonicate for 10 minutes. After centrifugation, collect the supernatant and detect. LC-MS / MS method is used to quantitatively detect the drug concentration in the sample. WinNonlin software is used to measure the relevant pharmacokinetic parameter T. max , C max , AUC 0-∞ , t 1 / 2 Calculate the following:

[0387] 4.3. Test Results As shown in Table 6, after oral administration of compound E68 to rats at 10 mg / kg, the maximum plasma drug concentration was 1600 ng / mL, the half-life was 4.88 hours, and the bioavailability was 62.2%, indicating good oral pharmacokinetic properties. Figure 2 shows the tissue distribution of compound E68 2 hours after oral administration of 10 mg / kg to rats, and it can be seen that E68 showed higher concentrations in each tissue, which were significantly higher than the plasma concentration. Therefore, when the compounds of the present invention are used to treat diseases such as psychiatric disorders, neurodegenerative disorders, liver diseases, and kidney diseases, they can be effectively distributed to target tissues.

[0388] [Table 7]

[0389] 5. Efficacy tests of anti-anxiety and anti-depressant drugs 5.1. Drug Preparation The test drugs were dissolved in a 2:2:96 (v / v / v) mixture of DMSO, Tween 80, and distilled water and then administered orally to the mice at a dose of 10 mL / kg. Dosage: positive control Fluoxetine, 10 mg / kg; HC-070, 0.1, 0.3, or 1 mg / kg; compound E68, 0.03, 0.1, 0.3, or 1 mg / kg. The blank control group received only the above solvent at a volume of 10 mL / kg.

[0390] 5.2.Experimental Method (1) Marble burial test The mouse marble-burying test is a behavioral assay used to screen for anxiolytic drugs. It utilizes the natural digging behavior of mice in their environment, such as burrows or escape tunnels, and is sensitive to subtle behavioral changes induced by genetic manipulation, disease, or drug treatment. The test has been shown to effectively detect anxiolytic effects in drug intervention and transgenic mouse models, and to accurately and sensitively measure repetitive and compulsive behaviors in rodents.

[0391] A 33cm x 24cm x 28cm cardboard box was covered with 5cm of sawdust, and 20 black marbles (14mm in diameter) were neatly arranged on the floor in a 4x5 pattern (4cm apart). A mouse was placed facing the corner of the cardboard box for 30 minutes, after which it was carefully removed and the number of marbles buried was counted. When more than two-thirds of the marbles were buried, the marbles were considered buried. First, a blank experiment was conducted. Before the experiment, the mouse was brought into the experimental environment and allowed to acclimate for one hour. After the experiment, the mouse was returned to its cage and the number of marbles buried was calculated. The next day, the mouse was brought into the experimental environment and allowed to acclimate for one hour before the experiment. The marble-burying experiment was then conducted one hour after administration. After the experiment, the mice were carefully returned to their cages to prevent them from moving about and affecting the results of the experiment. After counting, the black marbles were removed, sprayed with alcohol, wiped, and the bedding was turned over to remove the odor of the previous mouse and prevent it from affecting the next mouse experiment. The marbles were then neatly arranged for the next mouse experiment.

[0392] (2) Tail suspension test The mouse tail suspension test (TST) is a widely used experimental method for evaluating the efficacy of antidepressants and other psychotropic drugs. The principle of the test is to suspend a mouse by its tail and observe its behavior. The mouse initially attempts to escape, but when it realizes it cannot escape, it eventually stops struggling and becomes immobile, suggesting despair. This immobility behavior is used to measure depression-like states, and the severity of the condition is evaluated by recording the duration of immobility. Antidepressants and stimulants can significantly shorten this duration of immobility, which is why the TST is used for preliminary screening of these drugs. This test is highly sensitive to most antidepressants, and its efficacy is significantly correlated with clinical response, making it a standard initial test for antidepressant drugs.

[0393] Immediately after the marble-burying test, the same mouse was subjected to the tail suspension test. A tail suspension test box was constructed using a 55 cm high, 15 cm long, and 11.5 cm wide black frosted acrylic plate. The top and three surrounding sides of the box were covered with frosted blackboards, and one side was used for video recording. The mouse's tail was wrapped 2 cm from the tip with yellow tape and fixed upside down to the center of the upper acrylic plate. The tip of the tail was approximately 3 cm away from the acrylic plate, and the body was suspended in mid-air without touching the surrounding walls. The test period for each mouse lasted 6 minutes, and the time spent immobile within that time was recorded. After the experiment, the mouse was returned to its cage, and the inside of the box and the surface of the table were wiped with alcohol to remove the odor of the previous mouse and prevent it from affecting the next mouse test. The criteria for immobility in the tail suspension test were that the mouse stopped struggling, there was no bending or twisting of the body, the hind limbs were immobile, and the forelimbs and head were slightly moving.

[0394] 5.3.Experimental Results As shown in Figures 3 and 4, in the marble-burying test, the number of buried marbles in the blank control group did not change significantly before and after administration, while the number of buried marbles in the mice was significantly reduced after administration of Fluoxetine (10 mg / kg), HC-070 (1 mg / kg), and E68 (0.1-1 mg / kg), indicating that the drugs have a significant anxiolytic effect. In the tail suspension test, the immobility time of the blank control group did not change significantly before and after administration, while the immobility time of the mice was significantly reduced after administration of compounds E68, HC-070, or Fluoxetine, indicating that the drugs have a significant antidepressant effect.

[0395] In another experiment conducted after reducing the dosage (Figures 5 and 6), when mice were orally administered 0.03 mg / kg of compound E68, the number of buried marbles was significantly reduced, and the effect was even more pronounced when 0.1 mg / kg of E68 was orally administered, and was superior to the same dose of HC-070. Furthermore, when 0.03 mg / kg of compound E68 was orally administered, the immobility time of mice in the tail suspension box was also significantly reduced, and the effect was equivalent to that of 0.1 mg / kg of HC-070. It can be seen that compound E68 of the present invention has stronger anxiolytic and antidepressant effects than HC-070.

[0396] All documents mentioned in this application are incorporated by reference in this application as if each document were incorporated by reference individually. Furthermore, after reading the above teachings of the present invention, those skilled in the art will be able to make various changes or modifications to the present invention, and these equivalents will also fall within the scope defined by the appended claims of this application.

Claims

1. Compounds represented by general formula (I-a), their stereoisomers, and pharmaceutically acceptable salts thereof, 【Chemistry 1】 Here, R １ is a C1-C6 alkyl group or a C1-C6 halogenated alkyl group. n is an integer from 1 to 4, and each R ２ These are, independently, H, deuterium, halogen, C1-C6 alkyl group, C1-C6 halogenated alkyl group, C1-C6 alkoxy group, C1-C6 halogenated alkoxy group, and cyano group. Ring A is a saturated or partially unsaturated C3-C10 cyclic hydrocarbon group. m is an integer from 1 to 6, and each R ３ is, independently of one another, H, deuterium, a C1-C6 alkyl group, a C1-C6 halogenated alkyl group, a C1-C6 alkoxy group, a C1-C6 halogenated alkoxy group, a C3-C8 cycloalkyl group, a C6-C10 aryl group, a 5- to 8-membered heteroaryl group, a 4- to 8-membered heterocyclic group, a halogen, a cyano group, a hydroxy group, -NR ４ R ５ is selected from the group consisting of, or R ３ is combined with the carbon to which it is attached to the A ring to form a 4- to 8-membered heterocyclic or C6-C10 aryl group, and the cycloalkyl group, aryl group, heteroaryl group, heterocyclic group are unsubstituted or are optionally substituted by 1 to 4 groups selected from the group consisting of C<​​​​​​​ R ４ , R ５ The compound is characterized in that each of these elements is independently H, a C1-C6 alkyl group, and a C3-C8 cycloalkyl group.

2. Ring A is characterized by being a saturated or partially unsaturated C3-C8 monocyclic or bridging cyclic hydrocarbon group. The compound according to claim 1.

3. R １ It is characterized by being a C1-C4 alkyl group. The compound according to claim 1.

4. n is 1, 2, or 3, and each R ２ Each of these is independently characterized by being H, deuterium, halogen, C1-C4 alkyl group, C1-C4 halogenated alkyl group, C1-C4 alkoxy group, C1-C4 halogenated alkoxy group, and cyano group. The compound according to claim 1.

5. m is 1, 2, 3 or 4, and each R ３ These are, independently, H, deuterium, C1-C4 alkyl group, C1-C4 halogenated alkyl group, C1-C4 alkoxy group, C1-C4 halogenated alkoxy group, C3-C6 cycloalkyl group, phenyl group, 5-6 membered heteroaryl group, 5-6 membered heterocyclic group, F, Cl, Br, cyano group, hydroxyl group, -NR ４ R ５ Selected from or R ３ The carbon bonded to the A ring forms a 5-6 membered heterocyclic or phenyl group, and the cycloalkyl group, phenyl group, heteroaryl group, and heterocyclic group are either unsubstituted or optionally substituted with 1 to 4 groups selected from the group consisting of C1-C6 alkyl groups, C1-C6 halogenated alkyl groups, C1-C6 alkoxy groups, C1-C6 halogenated alkoxy groups, halogens, and cyano groups. The compound according to claim 1.

6. It has a structure as shown in general formula (I-b), 【Chemistry 2】 Here, R １ It is a C1-C6 alkyl group, n is an integer from 1 to 4, and each R ２ Each of these is independently H, a halogen atom, a C1-C6 alkyl halide, or CN. Each R ３ These are, independently, H, deuterium, C1-C6 alkyl group, C1-C6 halogenated alkyl group, C1-C6 alkoxy group, C1-C6 halogenated alkoxy group, C6-C10 aryl group, C3-C8 cycloalkyl group, 5-8 membered heteroaryl group, 4-8 membered heterocyclic group, halogen atom, cyano group, hydroxyl group, -NR ４ R ５ Either selected from or two R ３ It, together with the carbon bonded to the A ring, forms a 4- to 8-membered heterocyclic structure. R ４ , R ５ Each of these is independently characterized as H and C1-C6 alkyl groups. The compound according to claim 1.

7. The aforementioned compound is characterized by being selected from the following: The compound according to claim 1. 【Transformation 3】 【change】

8. A pharmaceutical composition, The pharmaceutical composition comprising the compound described in claim 1, its stereoisomers and pharmaceutically acceptable salts thereof, and a pharmaceutically acceptable carrier.

9. Use of the compound according to claim 1, The use is characterized by being used for the preparation of inhibitors or agonists of TRPC4 and TRPC5 channels, or for the preparation of drugs for preventing, delaying, or treating diseases associated with abnormal function or expression of TRPC4 and / or TRPC5.

10. The aforementioned disease is characterized by being a mental illness, neurodegenerative disease, kidney disease, pain, epilepsy, liver disease, cardiovascular disease, cancer, skin disease, or intestinal disease. Use as described in claim 9.

11. The mental disorder is borderline personality disorder, depression, dysthymia, postpartum depression, bipolar disorder, post-traumatic stress disorder, panic disorder, agoraphobia, social phobia, generalized anxiety disorder, social anxiety disorder, separation anxiety, schizophrenia, mania, obsessive-compulsive disorder, apathy, neurasthenia, or delusional disorder. The aforementioned neurodegenerative diseases include Alzheimer's disease, Parkinson's disease, Huntington's disease, amyotrophic lateral sclerosis, memory impairment, amnesia, aphasia, chronic fatigue syndrome, Creutzfeldt-Jakob disease, dissociative amnesia, fugue amnesia, learning disabilities, sleep disorders, or other brain disorders caused by trauma or aging. Kidney diseases include kidney damage caused by toxic substances, kidney damage caused by viral or bacterial infections, hypertensive nephropathy, diabetic nephropathy, glomerulonephritis, lupus nephritis, IgA nephropathy, nephrotic syndrome, membranous nephropathy, minimal change kidney, polycystic kidney disease, or focal segmental glomerulosclerosis, etc. The aforementioned pain is nociceptive pain, mechanical pain, inflammatory pain, cancer pain, or neuropathic pain. The aforementioned liver diseases include cholestatic liver disease, alcoholic steatohepatitis, non-alcoholic steatohepatitis, viral hepatitis, autoimmune liver disease, chemical liver injury, hepatic fibrosis, cirrhosis, or hepatocellular carcinoma. The aforementioned cardiovascular diseases include hypertension, arteriosclerosis, coronary artery disease, angina pectoris, myocardial infarction, arrhythmia, stroke, or pulmonary hypertension. The aforementioned cancers are renal cell carcinoma, breast cancer, or colorectal cancer. The aforementioned skin diseases are psoriasis, ichthyosis, palmoplantar keratosis, Olmsted's disease, ranula, or menopausal keratosis. The aforementioned intestinal disease is characterized by being ulcerative colitis, short bowel syndrome, irritable bowel syndrome, intestinal spasms, diarrhea, or abdominal pain. The use described in claim 10.

12. The disorder is characterized in that it is borderline personality disorder, depression, anxiety disorder, or post-traumatic stress disorder. Use as described in claim 9.

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