3,8-disubstituted adenine derivative, preparation method therefor and use thereof

Abstract

The present invention belongs to the technical field of medicinal chemistry, and specifically relates to a 3,8-disubstituted adenine derivative, a preparation method therefor and the use thereof. The 3,8-disubstituted adenine derivative provided in the present invention has a good inhibitory effect on phosphodiesterase type 8, has a good liver microsomal stability and drug-like properties, can be used in the preparation of a drug for treating and / or preventing diseases associated with phosphodiesterase type 8, and has a good development potential, thereby increasing the available options of drugs for treating diseases associated with phosphodiesterase type 8.

Description

A 3,8-disubstituted adenine derivative, preparation method and application thereof Technical Field

[0001] The present invention belongs to the technical field of medicinal chemistry, and more specifically, relates to a 3,8-disubstituted adenine derivative and an application thereof. Background Art

[0002] Phosphodiesterases (PDEs) are the only superfamily of enzymes in the body responsible for the specific hydrolysis of cyclic adenosine monophosphate (cAMP) and cyclic guanosine monophosphate (cGMP). They catalyze the hydrolysis of 3′-phosphate bonds to generate 5′-AMP and 5′-GMP, respectively. cAMP and cGMP are important intracellular second messengers, regulating diverse signaling pathways, such as various intracellular physiological processes and neurobehavioral functions, through direct interactions with cAMP-dependent protein kinase A (PKA) and cGMP-dependent protein kinase G (PKG), respectively. To date, nearly twenty PDE inhibitors have been approved for marketing, such as the PDE5 inhibitor sildenafil and the PDE4 inhibitor apremilast. cAMP-specific PDE subtypes include PDE4, PDE7, and PDE8. PDE8 has the highest affinity for its cAMP substrate, with a Km value of 40-150 nM, over 40 times that of PDE4.

[0003] The biological functions of PDE8 are not yet fully understood, but based on its expression, it is speculated that PDE8 plays an important physiological role in various life processes, particularly in brain and thyroid function. Therefore, it may be a suitable therapeutic target for central nervous system diseases. Studies have shown that PDE8B knockout mice have enhanced spatial memory and motor coordination, and that PDE8 levels are overexpressed in the brains of aged rats. These findings suggest that inhibiting PDE8 may enhance cognitive function, prevent age-related motor coordination decline, and treat age-related diseases. Technical issues

[0004] The technical problems to be solved by the present invention are:

[0005] There is little research on PDE8 in the prior art, and no inhibitors with good effects have been found for clinical application. There are only a few studies on PDE8 inhibitors. For example, Patent No. WO2011058478A1 discloses "a substituted triazolopyrimidine inhibitor of PDE8 enzyme". This type of inhibitor has different degrees of inhibitory activity on PDE8 enzyme, but its clinical effect is still unknown, and it is impossible to explore the physiological effects of PDE8. Therefore, there is an urgent need to discover PDE8 inhibitors with strong activity, high selectivity and excellent properties to explore the physiological effects of PDE8. Technical Solutions

[0006] In order to solve the above technical problems, the present invention adopts the following technical solutions:

[0007] The present invention provides a compound represented by formula (I) or a pharmaceutically acceptable salt thereof,

[0008] in:

[0009] R1, R2 and R3 are each independently selected from hydrogen, halogen, substituted or unsubstituted C 1-6 One of the alkoxy groups;

[0010] R4 is selected from hydrogen, halogen or

[0011] R5 is hydrogen, substituted or unsubstituted C 1-6 alkyl.

[0012] Further, R1 and R3 are each independently selected from hydrogen or halogen; R2 is selected from halogen, substituted or unsubstituted C 1-6 One of the alkoxy groups.

[0013] Furthermore, the R1, R2 or R3 is C 1-6 Alkoxy, and C 1-6 At least one H on the alkoxy group is substituted by a halogen atom.

[0014] Furthermore, the R5 is C 1-6 Alkyl, and C 1-6 At least one H on the alkyl group is replaced by a halogen atom, C 4-5 Heteroaryl, halogenated C 4-5 Heteroaryl, C6 aryl, halogenated C6 aryl, C 3-6 Cycloalkyl, halogenated C 3-6 Cycloalkyl or tetrahydropyran substitution.

[0015] Furthermore, the substituted or unsubstituted C 1-6 The alkoxy groups are each selected from one of -OCH3, -OCH2CH3, -OCH(CH3)2, -OCH2CH2CH3, -OCHF2, -OCH2CHF2, and -OCH2CH2CF3.

[0016] Furthermore, the substituted or unsubstituted C 1-6 Alkyl groups are selected from -CH3, -CH2CH3, -CH(CH3)2, -CH2CH2CH3, -CH2CH(CH3)2, -CH2CHF2, One of them.

[0017] Furthermore, the pharmaceutically acceptable salt is a salt obtained by reacting the compound of formula (I) with an acid, wherein the acid includes one or more of hydrochloric acid, hydrobromic acid, hydrofluoric acid, phosphoric acid, acetic acid, oxalic acid, sulfuric acid, methanesulfonic acid, salicylic acid, trifluoroacetic acid, trifluoromethanesulfonic acid, naphthalenesulfonic acid, maleic acid, fumaric acid, citric acid, tartaric acid, succinic acid, malic acid and glutamic acid.

[0018] Furthermore, the structure of the compound represented by formula (I) is shown as one of the following:

[0019] The present invention also provides a pharmaceutical composition, which comprises: the compound represented by formula (I) or a pharmaceutically acceptable salt thereof, and a pharmaceutically acceptable excipient.

[0020] In addition, the present invention also provides a method for preparing the compound represented by formula (I) or a pharmaceutically acceptable salt thereof. The synthetic route of the compound represented by formula (I) is as follows:

[0021] The specific steps include:

[0022] (1) When X in compound 1 is H:

[0023] (1) Compound 1 is dissolved in a solvent and reacted at 0-60°C for 12-18 hours under the action of liquid bromine. After the reaction is complete, compound 2 is obtained;

[0024] (2) dissolving the compound 2 obtained in step (1) in a solvent, reacting with a halogenated hydrocarbon at 0-60° C. for 8-16 hours in the presence of an alkaline reagent, and obtaining compound 3 after the reaction is complete;

[0025] (3) dissolving the compound 3 obtained in step (2) in a solvent and reacting at 100-150° C. for 20-36 hours under the action of thiourea to produce compound 4 after the reaction is complete;

[0026] (4) dissolving the compound 4 obtained in step (3) in a solvent, reacting with a halogenated hydrocarbon at 60-120° C. for 8-16 hours in the presence of an alkaline reagent, and obtaining the compound represented by formula (I) after the reaction is complete;

[0027] (2) When X in compound 1 is SH:

[0028] (1) Compound 1 is dissolved in a solvent and reacted with a halogenated hydrocarbon or a p-toluenesulfonate derivative at 80-150°C for 8-16 hours in the presence of an alkaline reagent. After the reaction is complete, compound 5 is obtained.

[0029] (2) Compound 5 is dissolved in a solvent and reacted with a halogenated hydrocarbon at 0-60° C. for 8-16 hours in the presence of an alkaline reagent. After the reaction is complete, the compound represented by formula (I) is obtained;

[0030] Wherein, the solvent involved in the above steps is one or more of water, N,N-dimethylformamide, N,N-dimethylacetamide, dimethyl sulfoxide, N-methylpyrrolidone, methanol, ethanol, isopropanol, acetonitrile, acetone, tetrahydrofuran, chloroform, ethyl acetate, dichloromethane, 1,2-dichloroethane, and 1,4-dioxane;

[0031] The alkaline reagent involved in the above steps is selected from one or more of diisopropylethylamine, triethylamine, 4-dimethylaminopyridine, piperidine, sodium bicarbonate, sodium carbonate, potassium carbonate, sodium hydroxide, potassium hydroxide, potassium tert-butoxide, sodium hydride, sodium methoxide, and sodium ethoxide.

[0032] The present invention also provides a use of a substance X in the preparation of a phosphodiesterase type 8 inhibitor, wherein the substance X is a compound represented by formula (I) or a pharmaceutically acceptable salt thereof, or the pharmaceutical composition.

[0033] The present invention also provides a use of a substance X in the preparation of a drug, wherein the substance X is a compound represented by formula (I) or a pharmaceutically acceptable salt thereof, or the pharmaceutical composition; the drug is a drug for treating and / or preventing diseases associated with phosphodiesterase type 8. Beneficial effects

[0034] Through the above technical solution, the present invention has the following beneficial effects:

[0035] The 3,8-disubstituted adenine derivatives provided by the present invention have a good inhibitory effect on phosphodiesterase type 8, high selectivity for other phosphodiesterase subtypes, and good liver microsomal stability and oral pharmacokinetic properties. They can be used in the preparation of drugs for treating and / or preventing diseases related to phosphodiesterase type 8, have good development potential, and increase the range of drugs available for treating diseases related to phosphodiesterase type 8. Best Mode for Carrying Out the Invention

[0036] The present invention will be further described below in conjunction with specific examples, but the examples do not limit the present invention in any form. Unless otherwise stated, the reagents, methods and equipment used in the present invention are conventional reagents, methods and equipment in the art. Unless otherwise stated, the reagents and materials used in the following examples are commercially available. Example

[0037] The preparation method of the 3,8-disubstituted adenine derivative (I) provided by the present invention is as follows:

[0038] The specific steps include:

[0039] (1) When X in compound 1 is H:

[0040] (1) Compound 1 is dissolved in N,N-dimethylformamide and reacted at 0-60°C for 12-18 hours under the action of liquid bromine. After the reaction is complete, compound 2 is obtained;

[0041] (2) Compound 2 obtained in step (1) is dissolved in N,N-dimethylacetamide, and reacted with a halogenated hydrocarbon at 0-60° C. for 8-16 hours under the action of diisopropylethylamine. After the reaction is complete, compound 3 is obtained;

[0042] (3) Compound 3 obtained in step (2) was dissolved in N,N-dimethylformamide and reacted at 120°C for 28 hours under the action of thiourea. After the reaction was complete, compound 4 was generated;

[0043] (4) dissolving the compound 4 obtained in step (3) in methanol and reacting with a halogenated hydrocarbon at 90° C. for 12 h in the presence of triethylamine. After the reaction is complete, the compound represented by formula (I) is obtained;

[0044] (2) When X in compound 1 is SH:

[0045] (1) Compound 1 is dissolved in water and reacted with a halogenated hydrocarbon or a p-toluenesulfonate derivative at 120°C for 12 h in the presence of 4-dimethylaminopyridine. Compound 5 is obtained after the reaction is complete.

[0046] (2) Compound 5 was dissolved in acetonitrile and reacted with a halogenated hydrocarbon at 30° C. for 12 h in the presence of piperidine. After the reaction was complete, the compound represented by formula (I) was obtained.

[0047] The synthesis process and effect verification of 3,8-disubstituted adenine derivatives are described below.

[0048] Modes for Carrying Out the Invention

[0049] Example 1: Synthesis of Compounds 1a-n

[0050] (1) Synthesis of intermediate 8-bromo-9H-purin-6-amine (M1)

[0051] Dissolve adenine (16 mmol) in water (200 mL) and slowly add liquid bromine (6 mL) dropwise. Allow to react at room temperature for 16 hours. Filter, wash with water to remove the remaining bromine, and dry to obtain a yellow solid. Yield: 28%. 1H NMR(400MHz, DMSO-d6)δ8.23(s,1H),8.06(s,2H).ESI:calculated for C5H4BrN5=211.96,213.96.Observed m / z[MH] - =212.00,214.00.

[0052] (2) Synthesis of intermediate 3-benzyl-8-bromo-3H-purin-6-amine (M2)

[0053] Intermediate M1 (10 mmol) was dissolved in N,N-dimethylformamide (15 mL), potassium carbonate (20 mmol) was added, and benzyl bromide (10 mmol) was added dropwise. The reaction was allowed to react at room temperature overnight. Water was added to the reaction system and extracted three times with ethyl acetate. The organic layer was collected, washed three times with saturated brine, and dried over anhydrous sodium sulfate. The solvent was removed by rotary evaporation under reduced pressure and purified by column chromatography to obtain a white solid. The yield was 33%. 1 H NMR (400MHz, DMSO-d6) δ8.54(s,1H),8.20(s,1H),8.08(s,1H),7.42–7.28(m,5H),5.47(s,2H).

[0054] (3) Synthesis of intermediate 6-amino-3-benzyl-3H-purine-8-thiol (M3)

[0055] Intermediate M2 (3 mmol) was dissolved in n-butanol (15 mL), and thiourea (24 mmol) was added. The mixture was refluxed for 24 hours. After the reaction was complete, the mixture was diluted with water, filtered, and the residue was washed with water. Drying afforded a white solid with a yield of 86%. 1 H NMR (400MHz, DMSO-d6) δ11.39(s,1H),8.51(s,1H),7.47–7.26(m,5H),5.42(s,2H).

[0056] (4) Synthesis of final products 1a-n

[0057] Intermediate M3 (0.3 mmol) and potassium hydroxide (0.45 mmol) were dissolved in a mixture of ethanol and water (2:1, 3 mL). Methyl iodide or an alkyl bromide or arylmethyl bromide (0.36 mmol) was added and allowed to react overnight at 80°C. After the reaction was complete, the mixture was diluted with water and extracted three times with ethyl acetate. The organic layers were combined, dried, and the solvent was evaporated by rotary evaporation. Purification by column chromatography yielded a white solid. The results are shown in Table 1.

[0058] Table 1

[0059] Example 2: Synthesis of Compound 2a-1

[0060] (1) Synthesis of intermediate 5-nitropyrimidine-4,6-diamine (M4)

[0061] Dissolve 4,6-dichloro-5-nitropyrimidine (26 mmol) in aqueous ammonia (15 mL) and react at 60°C for 5 hours. Filter, wash with water, and dry the residue to obtain a white solid with a yield of 95%. 1 H NMR (400MHz, DMSO-d6) δ8.49(s,2H),8.41(s,2H),7.88(s,1H).

[0062] (2) Synthesis of intermediate pyrimidine-4,5,6-triamine (M5)

[0063] Intermediate M4 (20 mmol) was dissolved in ethanol (400 mL), and Raney nickel (5 g) was added. 80% hydrazine hydrate solution (8 mL) was added dropwise. The mixture was reacted at room temperature for 40 hours. The mixture was filtered, the solvent was evaporated, and dried to obtain a white solid. The yield was 93%. 1 H NMR (400MHz, DMSO-d6) δ7.45(s,1H),5.55(s,4H),3.76(s,2H).

[0064] (3) Synthesis of intermediate 6-amino-9H-purine-8-thiol (M6)

[0065] Mix Intermediate M5 (10 mmol) with thiourea (35 mmol) and heat to 180°C for 1 hour. Add water while hot, crush the solid, and stir at room temperature overnight. Filter, wash with saturated brine, and dry the residue to obtain a yellow solid. Yield: 73%. 1 H NMR (400MHz, DMSO-d6) δ7.45(s,1H),5.55(s,4H),3.76(s,2H).

[0066] (4) Synthesis of intermediates M7a-e

[0067] Dissolve intermediate M6 (1 mmol) and potassium hydroxide (2 mmol) in a mixture of ethanol and water (2:1, 3 mL). Add a bromide or p-toluenesulfonate derivative (1.5 mmol) and allow to react overnight at room temperature. After the reaction is complete, remove the ethanol by rotary evaporation and adjust the pH to 5 with glacial acetic acid. Filter, wash with water, and dry the residue to obtain a yellow or white solid.

[0068] The yield of 8-(propylthio)-9H-purin-6-amine (M7a) was 74%. 1 H NMR (400 MHz, DMSO-d6) δ 12.95 (s, 1H), 8.02 (s, 1H), 6.96 (s, 2H), 3.24 - 3.18 (m, 2H), 1.73 - 1.64 (m, 2H), 0.98 (t, J = 7.3 Hz, 3H).

[0069] The yield of 8-((2-(tetrahydro-2H-pyran-4-yl)ethyl)thio)-9H-purin-6-amine (M7b) was 54%. 1 H NMR (500 MHz, DMSO-d6) δ 12.98 (s, 1H), 8.02 (s, 1H), 7.00 (s, 2H), 3.82 (dd, J = 11.0, 3.6 Hz, 2H), 3.32 - 3.20 (m, 4H), 1.68–1.55 (m, 5H), 1.24–1.10 (m, 2H).

[0070] The yield of 8-(((4,4-difluorocyclohexyl)methyl)thio)-9H-purin-6-amine (M7c) was 55%. 1 H NMR (400 MHz, DMSO-d6) δ 12.96 (s, 1H), 8.05 (s, 1H), 6.98 (s, 2H), 3.25 (d, J = 6.7 Hz, 2H), 2.05 - 1.96 (m, 2H), 1.92 - 1.87 (m, 2H), 1.85 - 1.70 (m, 3H), 1.33 - 1.23 (m, 2H).

[0071] The yield of 8-(((tetrahydro-2H-pyran-4-yl)methyl)thio)-9H-purin-6-amine (M7d) was 49%. 1 H NMR (400 MHz, DMSO-d6) δ 12.88 (s, 1H), 8.04 (s, 1H), 6.94 (s, 2H), 3.84 (dd, J = 11.2, 3.0 Hz, 2H), 3.27 - 3.20 (m, 4H), 1.88 - 1.77 (m, 1H), 1.75 - 1.64 (m, 2H), 1.34 - 1.17 (m, 2H).

[0072] The yield of 8-((cyclohexylmethyl)thio)-9H-purin-6-amine (M7e) was 38%. 1H NMR (400MHz, DMSO-d6) δ12.82(s,1H),8.04(s,1H),6.96(s,2H),3.18(d,J=6.8Hz,2H),1.82( d,J=11.3Hz,2H),1.73-1.64(m,2H),1.62-1.48(m,2H),1.27-1.09(m,3H),1.07-0.92(m,2H).

[0073] (5) Synthesis of final product 2a-1

[0074] Intermediate M7a-e (0.5 mmol) was dissolved in N,N-dimethylformamide (1 mL), and cesium carbonate (1 mmol) was added, followed by a benzyl bromide derivative (0.5 mmol). The mixture was allowed to react overnight at room temperature. After completion, the mixture was diluted with water and extracted three times with ethyl acetate. The combined organic layers were washed three times with brine, dried, and the solvent was evaporated by rotary evaporation. The mixture was then purified by column chromatography to yield a white or pale yellow solid. The results are shown in Table 2.

[0075] Table 2

[0076] Example 3: Synthesis of Compounds 3a-f

[0077] The synthesis process of compounds 3a-f refers to the synthesis process of compounds 2a-1 in Example 2 above. The results are shown in Table 3.

[0078] Table 3

[0079] Example 4: Compound 3-(3,5-dichloro-4-(2,2-difluoroethoxy)benzyl)

[0080] Synthesis of 3H-purin-6-amine (4a)

[0081] (1) Synthesis of the intermediate N-(2,4-dimethoxybenzyl)-9H-purin-6-amine (M8)

[0082] Dissolve 6-chloropurine (20 mmol), 2,4-dimethoxybenzylamine (24 mmol), and N,N-diisopropylethylamine (40 mmol) in n-butanol (16 mL) and react at 100°C for 3 h. A white solid precipitates. Cool to room temperature, dilute with acetonitrile, filter, and wash to obtain a white solid. Yield: 96%. 1H NMR (400MHz, DMSO-d6) δ12.92(s,1H),8.12(d,J=19.2Hz,2H),7.76(s,1H),7.07( s,1H),6.56(s,1H),6.42(d,J=7.8Hz,1H),4.59(s,2H),3.82(s,3H),3.72(s,3H).

[0083] (2) Synthesis of the intermediate 3-(3,5-dichloro-4-(2,2-difluoroethoxy)benzyl)-N-(2,4-dimethyl-benzyl)-3H-purin-6-amine (M9)

[0084] Intermediate M8 (7 mmol) and 5-bromomethyl-1,3-dichloro-2-(2,2-difluoroethoxy)benzene (7 mmol) were dissolved in N,N-dimethylformamide (40 mL) and reacted overnight at 110°C. The solvent was evaporated using an oil pump, and the mixture of dichloromethane and methanol was slurried, filtered, and washed to obtain a white solid. The yield was 74%. 1 H NMR (400 MHz, CDCl3) δ8.07 (s, 1H), 7.99 (s, 1H), 7.38 (s, 2H), 7.26-7.22 (m, 1H), 6.47 (d, J = 1.9 Hz, 1H), 6.41 (dd, J = 8.2, 1.9 Hz, 1H), 6.14 (tt, J = 55.2, 4.4 Hz, 1H), 5.45 (s, 2H), 4.80 (s, 2H), 4.21 (td, J = 12.9, 4.2 Hz, 2H), 3.85 (s, 3H), 3.79 (s, 3H). (5) Synthesis of the final product 3-(3,5-dichloro-4-(2,2-difluoroethoxy)benzyl)-3H-purin-6-amine (4a)

[0085] Intermediate M9 (5 mmol) was dissolved in a trifluoroacetic acid / dichloromethane mixture (1:3, 20 mL) and allowed to react overnight at room temperature. The solvent was evaporated, diluted with methanol, and the pH was adjusted to 8 with methanolic ammonia. The product was purified by column chromatography to obtain a white solid in a 97% yield. 1 H NMR (400MHz, MeOD) δ 8.56 (s, 1H), 8.08 (s, 1H), 7.53 (s, 2H), 6.19 (tt, J = 54.6, 4.2Hz, 1H), 5.54 (s, 2H), 4.23 (td, J = 13.6, 2.7Hz, 2H).

[0086] Example 5: Synthesis of compound 8-bromo-3-(3,5-dichloro-4-(2,2-difluoroethoxy)benzyl)-3H-purin-6-amine (4b)

[0087] Compound 4a (1 mmol) from Example 4 was dissolved in anhydrous N,N-dimethylformamide (2.5 mL). N-bromosuccinimide (1.2 mmol) was added and the mixture was allowed to react overnight at room temperature. The solvent was evaporated using an oil pump and the product was purified by column chromatography to obtain a white solid. The yield was 83%. 1 H NMR (500MHz, Acetone-d6) δ8.52(s,1H),7.69(s,2H),6.33(tt,J=54.7,3.8Hz,1H),5.57(s,2H),4.33(td,J=13.9,3.9Hz,2H). Industrial Applicability

[0088] 1. Biological activity assay

[0089] (1) Determination of the inhibitory activity of compounds on PDE8A enzyme

[0090] 3 H-cAMP was diluted to 20,000-30,000 cpm in assay buffer (20 mM Tris-HCl (pH 7.5), 10 mM MnCl2, and 1 mM DTT). The substrate, PDE8A protein, and test compound were incubated at room temperature for 15 minutes, and then the reaction was terminated by adding 0.2 M ZnSO4 and 0.2 M Ba(OH)2. The unreacted cAMP in the supernatant was measured using a PerkinElmer 2910 counter. 3 H-cAMP. Each test compound IC 50 The calculation used 8-10 different concentrations and repeated the test 3 times. The activity results are shown in Table 4.

[0091] Table 4

[0092] (2) Selectivity test of compound 4b against other PDE subtypes

[0093] Compound 4b was used as an example to test its selectivity index for the PDEs family. The test method was the same as the above-mentioned determination of the PDE8A enzyme inhibitory activity. The test results are shown in Table 5. From the data in the table, it can be seen that compound 4b has good subselectivity for other PDEs.

[0094] Table 5

[0095] 2. Metabolic stability of compound 4b in rat liver microsomes

[0096] This study investigated the metabolic stability of compound 4b in liver microsomes by incubating compound 4b with liver microsomes in vitro and determining the percentage of the remaining compound using LC-MS / MS. The specific steps are as follows:

[0097] (1) Solution preparation

[0098] (1) Preparation of Tris / HCl (0.1 M, pH 7.4) buffer: Weigh 12.12 g of TRIS (tris-hydroxymethyl aminomethane) and dissolve it in 800 mL of water. Adjust the pH to 7.4 with HCl (2 M), and then dilute to 1000 mL with water.

[0099] (2) Preparation of MgCl2: Prepare MgCl2 (100 mM) solution using 0.1 M Tris / HCl buffer, aliquot and store at -20°C.

[0100] (3) Preparation of NADPH: Prepare NADPH (10 mM) solution using 0.1 M Tris / HCl buffer, aliquot and store at -20°C.

[0101] (4) Preparation of test compounds: Dissolve the compound in DMSO to obtain a 10 mM stock solution. Dilute to 1 mM with DMSO and then dilute with 0.1% BSA-water to obtain a 2 μM working solution. In the incubation system, the 2 μM working solution was diluted 20-fold to a final concentration of 0.1 μM. The DMSO concentration in the incubation system was ≤ 0.01%.

[0102] (5) Preparation of positive control compound: Dilute 2 mM VIVID stock solution 50-fold with 2 μM analyte working solution. The final concentration of VIVID in the incubation system is 2 μM.

[0103] (2) Experimental methods

[0104] Liver microsomes were incubated in 96-well plates in a 450 μL incubation volume. The medium consisted of 0.1 M Tris buffer (pH 7.4), containing liver microsomes at a final concentration of 0.33 mg / mL, 0.1 μM test drug, 5.0 mM MgCl2, 0.01% DMSO, 0.005% BSA, and 1.0 mM NADPH. Incubation was performed at 37°C. After a 10-minute preincubation, NADPH was added to initiate the reaction. The reaction was terminated by adding 50 μL of the incubation solution to the same volume of methanol at 0, 7, 17, 30, and 60 minutes. The results are shown in Table 6.

[0105] Table 6

[0106] 3. In vivo pharmacokinetic determination of compound 4b

[0107] The time course of the plasma concentration of compound 4b was observed after oral gavage and intravenous injection in rats, and the corresponding pharmacokinetic parameters and absolute bioavailability were estimated. The determination process and results were as follows:

[0108] (1) Measurement process

[0109] Six rats were divided into two groups and administered compound 4b via oral gavage and tail vein injection. Approximately 0.25 mL of blood was collected from the jugular vein 5, 15, 30, 1, 2, 4, 6, 8, and 24 hours after administration in the intravenous group and 5, 15, 30, 1, 2, 4, 6, 8, and 24 hours after administration in the oral gavage group. The concentration of LWWX22194 in rat plasma samples was determined by LC-MS / MS. PK parameters for each compound were statistically analyzed and calculated (WinNonlin V5.2, Pharsight) to demonstrate the pharmacokinetic properties of the compounds of this invention in rats.

[0110] (2) Measurement results

[0111] After a single intravenous injection of 2.5 mg / kg of compound 4b in rats, the main pharmacokinetic parameters of intravenous administration were: Cmax of 1847 ng / mL, Tmax of 0.083 h, T1 / 2 of 3.78 h, AUC0-T of 4900 hr*ng / mL, AUC0-∞ of 5028 hr*ng / mL, Vz of 2820 mL / kg, Cl of 512 mL / hr / kg, MRT0-t of 3.104 h, and MRT0-∞ of 3.53 h. The results are shown in Table 7 below.

[0112] Table 7: Pharmacokinetic parameters of 4b in rats after intravenous injection (n=3)

[0113] After oral administration of 10 mg / kg 4b to rats, the main pharmacokinetic parameters of oral administration were: Cmax of 1763 ng / mL, Tmax of 5.33 h, T1 / 2 of 3.34 h, AUC0-t of 22932 hr*ng / mL, AUC0-∞ of 23354 hr*ng / mL, MRT0-t of 6.75 h, MRT0-∞ of 7.10 h, and oral bioavailability of 116.1%. The results are shown in Table 8 below.

[0114] Table 8: Pharmacokinetic parameters of 4b in rats after oral administration (n=3)

[0115] The contents of Tables 4 to 8 above indicate that the compounds provided herein exhibit excellent PDE8 inhibitory activity, PDE subtype selectivity, good liver microsomal stability, and favorable pharmacokinetic properties, among other drug-like properties, fully demonstrating that the 3,8-disubstituted adenine derivatives of the present invention possess excellent drug-like properties. Therefore, the 3,8-disubstituted adenine derivatives of the present invention have broad application prospects as phosphodiesterase type 8 (PDE8) inhibitors and can be used for further drug research and development.

[0116] The above embodiments are preferred implementation modes of the present invention, but the implementation modes of the present invention are not limited to the above embodiments. Any other changes, modifications, substitutions, combinations, and simplifications that do not deviate from the spirit and principles of the present invention should be considered as equivalent replacement methods and are included in the scope of protection of the present invention.

Claims

1. A compound represented by formula (I) or a pharmaceutically acceptable salt thereof, characterized in that: in: The R1, R2 or R3 is C 1-6 Alkoxy, H, halogen, and C 1-6 At least one H on the alkoxy group is replaced by a halogen atom; R4 is selected from R5 is C 1-6 Alkyl, and C 1-6 At least one H on the alkyl group is replaced by a halogen atom, C 4-5 Heteroaryl, halogenated C 4-5 Heteroaryl, halogenated C6 aryl, C 3-6 Cycloalkyl, halogenated C 3-6 Cycloalkyl or tetrahydropyran substitution.

2. The compound of formula (I) or a pharmaceutically acceptable salt thereof according to claim 1, characterized in that: The C 1-6 The alkoxy group is selected from one of -OCHF2, -OCH2CHF2, and -OCH2CH2CF3.

3. The compound of formula (I) or a pharmaceutically acceptable salt thereof according to claim 1, characterized in that: The C 1-6 Alkyl is selected from -CH2CHF2, One of them.

4. The compound of formula (I) or a pharmaceutically acceptable salt thereof according to claim 1, characterized in that: The pharmaceutically acceptable salt is a salt obtained by reacting a compound of formula (I) with an acid, wherein the acid includes one or more of hydrochloric acid, hydrobromic acid, hydrofluoric acid, phosphoric acid, acetic acid, oxalic acid, sulfuric acid, methanesulfonic acid, salicylic acid, trifluoroacetic acid, trifluoromethanesulfonic acid, naphthalenesulfonic acid, maleic acid, fumaric acid, citric acid, tartaric acid, succinic acid, malic acid and glutamic acid.

5. A pharmaceutical composition comprising: a compound represented by formula (I) according to any one of claims 1 to 4 or a pharmaceutically acceptable salt thereof, and a pharmaceutically acceptable excipient.

6. Use of a substance X in the preparation of a phosphodiesterase type 8 inhibitor, characterized in that: The substance X is a compound represented by formula (I) according to any one of claims 1 to 4 or a pharmaceutically acceptable salt thereof, or a pharmaceutical composition according to claim 5.

7. Use of a substance X in the preparation of a medicine, characterized in that: The substance X is a compound represented by formula (I) according to any one of claims 1 to 4 or a pharmaceutically acceptable salt thereof, or a pharmaceutical composition according to claim 5; the drug is a drug for treating and / or preventing diseases related to phosphodiesterase type 8.

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