A thiophenopyrimidine compound containing a thiophene methylene structure, a preparation method thereof and applications thereof

Abstract

The present invention relates to a thiophenopyrimidine compound containing a thiophene methylene structure, a preparation method and an application thereof. The compound has the structure shown in Formula I. The present invention also relates to a preparation method of a compound containing the structure of Formula I and a pharmaceutical composition. The present invention also provides an application of the above compound in the preparation of a drug for reducing uric acid. #imgabs0#

Description

Technical Field

[0001] The present invention belongs to the technical field of organic compound synthesis and pharmaceutical applications. Specifically, the present invention relates to a thiophenopyrimidine compound containing a thiophene methylene structure, a preparation method thereof, a pharmaceutical composition containing them, and their applications in medicine. Background Art

[0002] Gout is one of the most common chronic diseases in inflammatory arthritis, and hyperuricemia is the direct inducement for the formation of gout. The diagnostic criteria for hyperuricemia in the Chinese Gout Diagnosis and Treatment Guidelines are that the fasting blood uric acid on two non - same - day occasions is > 0.42 mmol / L (7.0 mg / dL), and the criteria for asymptomatic hyperuricemia in the American Gout Guidelines are that the blood uric acid is > 0.408 mmol / L (6.8 mg / dL). Under long - term hyperuricemia, sodium urate slowly deposits as needle - like crystals in soft tissues or joints. Urate crystals can activate the NLRP3 inflammasome in macrophages and monocytes, and then cause an inflammatory response, inducing gout. Gout treatment drugs are mainly divided into three categories according to their mechanisms of action: (1) Xanthine oxidase inhibitors that inhibit uric acid production, and the marketed drugs include allopurinol and febuxostat; (2) URAT1 inhibitors that promote uric acid excretion, and the marketed drugs include benzbromarone, lesinurad, and probenecid; (3) Uricase analogs that decompose uric acid, and the marketed drug includes polyethylene glycol recombinant uricase. Lesinurad is a new type of URAT1 inhibitor approved by the FDA in 2015, and its single - drug dose is 200 - 600 mg. Because of its severe renal toxicity, it is finally used in combination with a xanthine oxidase inhibitor at a dose of 200 mg to treat hyperuricemia with gout. When combined with allopurinol at 200 mg per day, only 55% of patients reach the treatment endpoint of < 0.36 mmol / L, which is extremely poor in efficacy compared with 43% of patients who reach the standard when using allopurinol alone. Therefore, structural modification of it is expected to obtain new anti - hyperuricemia and anti - gout drugs with better activity, higher safety, and independent intellectual property rights. Summary of the Invention

[0003] Aiming at the deficiencies of the prior art, the present invention provides a thiophenopyrimidine compound containing a thiophene methylene structure and a preparation method thereof. The present invention also provides the activity screening results and applications of the above - mentioned compound as a URAT1 inhibitor.

[0004] The technical solution of the present invention is as follows:

[0005] I. Thiophenopyrimidine Compounds Containing a Thiophene Methylene Structure

[0006] A thiophenopyrimidine compound containing a thiophene methylene structure or a pharmaceutically acceptable salt thereof, characterized in that it has a structure shown in the following general formula I:

[0007]

[0008]

[0009] Among them, Ar1 and Ar2 are thiophene or thiophene substituted by one or more R2; R is C 1-6 alkylene or C alkylene substituted by one or more R3 1-6 alkylene; the R2 is selected from H, C 1-6 alkyl, C 1-6 alkoxy, halogenated C 1-6 alkyl, halogenated C 1-6 alkoxy, C 3-6 cycloalkyl, halogen, -OH, -COOH, -SH or two R2 and the two carbon atoms to which they are attached together form an aromatic ring optionally substituted by R4 5-12 ; the R3 and R4 are selected from H, C 1-6 alkyl, C 1-6 alkoxy, halogenated C 1-6 alkyl, halogenated C 1-6 alkoxy, C 3-6 cycloalkyl, halogen, -OH, -COOH, -SH.

[0010] Preferably, R is C 1-3 alkylene or C alkylene substituted by one or more R3 1-3 alkylene; the R2 is selected from H, C 1-3 alkyl, C 1-3 alkoxy, halogenated C 1-3 alkyl, halogenated C 1-3 alkoxy, C 3-4 cycloalkyl, halogen, -OH, -COOH, -SH or two R2 and the two carbon atoms to which they are attached together form a benzene ring, thiophene, pyridine or furan optionally substituted by R4; the R3 and R4 are selected from H, C 1-3 alkyl, C 1-3 alkoxy, halogenated C1-3 alkyl, halogenated C 1-3 alkoxy, C 3-4 cycloalkyl, halogen, -OH, -COOH, -SH.

[0011] Furthermore, R is C 1-3 alkylene or C alkylene substituted by one or more R3 1-3 alkylene; the R2 is selected from H, C 1-3 alkyl, C 1-3 alkoxy, halogenated C 1-3 alkyl, halogen or two R2 and the two carbon atoms to which they are attached together form a benzene ring optionally substituted by R4; the R3 and R4 are selected from H, C 1-3Alkyl, C1-3 alkoxy, halo C1-3 alkyl, halo C 1-3 alkoxy, halogen.

[0012] Preferably, the thiophenopyrimidine compounds containing a thiophenemethylene structure or pharmaceutically acceptable salts thereof are selected from the following formulas I-1 to I-5:

[0013]

[0014] Furthermore, the thiophenopyrimidine compounds containing a thiophenemethylene structure or pharmaceutically acceptable salts thereof are preferably selected from the following formulas I-1' to I-5':

[0015]

[0016] Preferably, R is selected from methylene, cyclopropyl-substituted methylene, cyclobutyl-substituted methylene, methylmethylene, dimethylmethylene; R2 is selected from H, methyl, ethyl, propyl, cyclopropyl, isopropyl, methoxy, ethoxy, trifluoromethyl, difluoromethyl, fluoromethyl, fluorine, chlorine, bromine; R4 is selected from H, methyl, ethyl, propyl, isopropyl, cyclopropyl, methoxy, ethoxy, trifluoromethyl, difluoromethyl, fluoromethyl, fluorine, chlorine, bromine.

[0017] Furthermore, the connection mode of Ar2 and methylene is thiophen-2-yl or thiophen-3-yl; the substituent R2 is one or more of methyl, ethyl, cyclopropyl or bromine.

[0018] As a preferred embodiment, the thiophenopyrimidine compounds containing a thiophenemethylene structure or pharmaceutically acceptable salts thereof are selected from the following compounds:

[0019] Structural formulas of Compounds 1-18 in Table 1

[0020]

[0021]

[0022] II. Preparation method of thiophenopyrimidine compounds containing a thiophenemethylene structure

[0023] The preparation method of the thiophenopyrimidine compounds containing a thiophenemethylene structure in the present invention includes the following steps:

[0024] (1) Using aminothiophenecarboxylate or substituted aminothiophenecarboxylate as raw materials, dichloromethane as a solvent, and triethylamine as an acid-binding agent, reacting with thiophosgene to generate isothiocyanate (M1);

[0025] (2) Using tetrahydrofuran as the solvent, M1 reacts with thiophene methylamine or substituted thiophene methylamine under the action of triethylamine to form a thiourea intermediate, and then cyclizes under the action of potassium hydroxide to form a thienopyrimidinone intermediate (M2);

[0026] (3) Using DMF as the solvent, M2 reacts with 2-bromo carboxylic acid under the action of potassium carbonate to form the target compound;

[0027]

[0028] Reaction conditions: (i) Thiophosgene, dichloromethane, triethylamine, 0 - 30 °C; (ii) a: Thiophene methylamine, tetrahydrofuran, triethylamine, room temperature; b: Methanol, potassium hydroxide, room temperature; (iii) N,N-Dimethylformamide, potassium carbonate, 2-bromo carboxylic acid, room temperature.

[0029] The room temperature mentioned in the present invention refers to 20 - 30 °C.

[0030] III. Application of Thienopyrimidine Compounds Containing Thiophene Methylene Structure

[0031] The present invention discloses the screening results of the inhibitory effect of thienopyrimidine compounds containing thiophene methylene structure on the uric acid transporter hURAT1 and their application as anti-hyperuricemic drugs. Activity screening experiments prove that the thienopyrimidine compounds containing thiophene methylene structure of the present invention all show a certain inhibitory effect on the uric acid transporter hURAT1.

[0032] Using Lesinurad as the positive control drug, the inhibitory effect of the compounds (numbered 1 - 12) synthesized in the present invention on the uric acid transporter hURAT1 was screened (single concentration experiment). The results showed that these 12 compounds all showed a certain inhibitory effect on the uric acid transporter hURAT1, and the inhibitory activities of 10 compounds were better than or equivalent to that of the positive control Lesinurad. The inhibitory effect IC 50 tests were carried out on compounds 3 and 4 with stronger single concentration inhibitory activities, and the inhibitory effect IC 50 of compounds 3 and 4 were measured to be 3.097 and 1.514 μmol / L respectively, and the activities were significantly better than that of the positive control Lesinurad (IC 50 = 7.3 μmol / L).

[0033] Therefore, the thienopyrimidine compounds containing thiophene methylene structure involved in the present invention have novel structures and strong inhibitory effects on the uric acid transporter hURAT1, and can be used as candidate compounds for the preparation of anti-hyperuricemic drugs and applied to the treatment of gout and hyperuricemia.

[0034] A uric acid-lowering pharmaceutical composition, comprising a thiophenopyrimidine compound containing a thiophene methylene structure of the present invention or a pharmaceutically acceptable salt thereof and one or more pharmaceutically acceptable carriers or excipients. Description of the Drawings

[0035] Figure 1 : Inhibitory effect of compounds 1-12 at 1 μmol / L on hURAT1

[0036] Figure 2 : Inhibitory effect of compound 3 on hURAT1

[0037] Figure 3 : Inhibitory effect of compound 4 on hURAT1 Detailed Description of the Invention

[0038] The following describes the embodiments of the present application through examples. Those skilled in the art should recognize that these specific examples only show the implementation technical solutions selected for the purpose of achieving the present application, and do not limit the technical solutions. According to the teachings of the present application, the improvement of the technical solutions of the present application in combination with the prior art is obvious and falls within the scope of protection of the present application.

[0039] Synthetic route of the compound:

[0040]

[0041] Reaction conditions: (i) Thionyl chloride, dichloromethane, triethylamine, 0-30 °C; (ii) a: Thiophene methylamine, tetrahydrofuran, triethylamine, room temperature; b: Methanol, potassium hydroxide, room temperature; (iii) N,N-Dimethylformamide, potassium carbonate, 2-bromo carboxylic acid, room temperature.

[0042] Example 1. Preparation of compound 1-M1

[0043]

[0044] Dissolve methyl 2-aminothiophene-3-carboxylate (2.36 g, 15 mmol) in 30 mL of dichloromethane in a 100 mL round-bottom flask, cool to 0 °C, and dropwise add thionyl chloride (2.07 g, 18 mmol). After stirring at this temperature for 10 min, dropwise add triethylamine (3.04 g, 30 mmol). After the addition is complete, gradually return to room temperature and monitor by TLC after 1 h. After the reaction is complete, pour the reaction system into 100 ml of water, extract three times with dichloromethane (60 mL), combine the organic phases, wash with saturated NaCl aqueous solution (100 ml), separate the layers, dry the organic phase with anhydrous sodium sulfate, and filter. The filtrate is concentrated under reduced pressure and purified by column chromatography (EA:PE = 1:20) to obtain intermediate compound 1-M1 (2.47 g) with a yield of 82.6%.

[0045] Example 2. Preparation of Compound 2-M1

[0046]

[0047] The operation was the same as that in Example 1, except that methyl 3-aminothiophene-2-carboxylate (2.36 g, 15 mmol) was reacted with thiophosgene (2.07 g, 18 mmol) to obtain intermediate compound 2-M1 (2.55 g) with a yield of 85.3%.

[0048] Example 3. Preparation of Compound 3-M1

[0049]

[0050] The operation was the same as that in Example 1, except that methyl 2-amino-4-methyl-3-carboxylate (2.57 g, 15 mmol) was reacted with thiophosgene (2.07 g, 18 mmol) to obtain intermediate compound 3-M1 (2.80 g) with a yield of 87.4%.

[0051] Example 4. Preparation of Compound 4-M1

[0052]

[0053] The operation was the same as that in Example 1, except that methyl 3-amino-4-methyl-2-carboxylate (2.57 g, 15 mmol) was reacted with thiophosgene (2.07 g, 18 mmol) to obtain intermediate compound 4-M1 (2.95 g) with a yield of 92.3%.

[0054] Example 5. Preparation of Compound 5-M1

[0055]

[0056] The operation was the same as that in Example 1, except that methyl 3-aminothiophene-4-carboxylate (2.36 g, 15 mmol) was reacted with thiophosgene (2.07 g, 18 mmol) to obtain intermediate compound 5-M1 (2.59 g) with a yield of 86.5%.

[0057] Example 6. Preparation of Compound 6-M1

[0058]

[0059] The operation was the same as that in Example 1, except that methyl 3-aminobenzo[b]thiophene-2-carboxylate (3.11 g, 15 mmol) was reacted with thiophosgene (2.07 g, 18 mmol) to obtain intermediate compound 6-M1 (3.13 g) with a yield of 83.7%.

[0060] Example 7. Preparation of Compound 1-M2

[0061]

[0062] Dissolve the intermediate compound 1-M1 (0.70 g, 3.5 mmol) in 100 ml round-bottom flask with tetrahydrofuran (30 ml). Sequentially add 2-thiophenecarboxamide (0.48 g, 4.2 mmol) and triethylamine (0.71 g, 7 mmol), and stir at room temperature. After 30 min, when the reaction is complete monitored by TLC, evaporate tetrahydrofuran. Add methanol (30 ml) to dissolve, add KOH (0.30 g, 5.3 mmol), and continue stirring at room temperature. After 30 min, the reaction is complete monitored by TLC. Pour the reaction system into 100 ml of water, extract three times with ethyl acetate (50 ml), combine the organic phases, wash the organic phase with saturated NaCl solution (50 ml), dry over anhydrous sodium sulfate and filter. Then recrystallize with ethyl acetate to obtain the intermediate compound 1-M2 (0.83 g), with a yield of 84.2%.

[0063] Example 8. Preparation of Compound 2-M2

[0064]

[0065] The operation is the same as in Example 7, except that the intermediate compound 2-M1 (0.70 g, 3.5 mmol) reacts with 2-thiophenecarboxamide (0.48 g, 4.2 mmol), and recrystallize with ethyl acetate to obtain the intermediate compound 2-M2 (0.81 g), with a yield of 82.6%.

[0066] Example 9. Preparation of Compound 3-M2

[0067]

[0068] The operation is the same as in Example 7, except that the intermediate compound 3-M1 (0.75 g, 3.5 mmol) reacts with 2-thiophenecarboxamide (0.48 g, 4.2 mmol), and recrystallize with ethyl acetate to obtain the intermediate compound 3-M2 (0.88 g), with a yield of 85.6%.

[0069] Example 10. Preparation of Compound 4-M2

[0070]

[0071] The operation is the same as in Example 7, except that the intermediate compound 4-M1 (0.75 g, 3.5 mmol) reacts with 2-thiophenecarboxamide (0.48 g, 4.2 mmol), and recrystallize with ethyl acetate to obtain the intermediate compound 4-M2 (0.84 g), with a yield of 81.2%.

[0072] Example 11. Preparation of Compound 5-M2

[0073]

[0074] The operation was the same as that in Example 7, except that the intermediate compound 5-M1 (0.70 g, 3.5 mmol) was reacted with 2-thiophenemethylamine (0.48 g, 4.2 mmol), and recrystallized from ethyl acetate to obtain the intermediate compound 5-M2 (0.84 g) with a yield of 85.8%.

[0075] Example 12. Preparation of Compound 6-M2

[0076]

[0077] The operation was the same as that in Example 7, except that the intermediate compound 6-M1 (0.87 g, 3.5 mmol) was reacted with 2-thiophenemethylamine (0.48 g, 4.2 mmol), and recrystallized from ethyl acetate to obtain the intermediate compound 6-M2 (0.94 g) with a yield of 81.0%.

[0078] Example 13. Preparation of Compound 7-M2

[0079]

[0080] The operation was the same as that in Example 7, except that the intermediate compound 1-M1 (0.70 g, 3.5 mmol) was reacted with 2-thiophenemethylamine (0.48 g, 4.2 mmol), and recrystallized from ethyl acetate to obtain the intermediate compound 7-M2 (0.87 g) with a yield of 88.3%.

[0081] Example 14. Preparation of Compound 8-M2

[0082]

[0083] The operation was the same as that in Example 7, except that the intermediate compound 2-M1 (0.70 g, 3.5 mmol) was reacted with 2-thiophenemethylamine (0.48 g, 4.2 mmol), and recrystallized from ethyl acetate to obtain the intermediate compound 8-M2 (0.84 g) with a yield of 85.4%.

[0084] Example 15. Preparation of Compound 9-M2

[0085]

[0086] The operation was the same as that in Example 7, except that intermediate compound 3-M1 (0.75 g, 3.5 mmol) was reacted with 2-thiophene methylamine (0.48 g, 4.2 mmol), and recrystallized from ethyl acetate to obtain intermediate compound 9-M2 (0.89 g) with a yield of 86.4%.

[0087] Example 16. Preparation of Compound 10-M2

[0088]

[0089] The operation was the same as that in Example 7, except that intermediate compound 4-M1 (0.75 g, 3.5 mmol) was reacted with 2-thiophene methylamine (0.48 g, 4.2 mmol), and recrystallized from ethyl acetate to obtain intermediate compound 10-M2 (0.86 g) with a yield of 83.7%.

[0090] Example 17. Preparation of Compound 11-M2

[0091]

[0092] The operation was the same as that in Example 7, except that intermediate compound 5-M1 (0.70 g, 3.5 mmol) was reacted with 2-thiophene methylamine (0.48 g, 4.2 mmol), and recrystallized from ethyl acetate to obtain intermediate compound 11-M2 (0.78 g) with a yield of 79.6%.

[0093] Example 18. Preparation of Compound 12-M2

[0094]

[0095] The operation was the same as that in Example 7, except that intermediate compound 6-M1 (0.87 g, 3.5 mmol) was reacted with 2-thiophene methylamine (0.48 g, 4.2 mmol), and recrystallized from ethyl acetate to obtain intermediate compound 12-M2 (0.94 g) with a yield of 81.7%.

[0096] Example 19. Preparation of Compound 1

[0097]

[0098] Compound 1-M2 (0.14 g, 0.5 mmol) was dissolved in 10 ml of DMF in a 25 ml round-bottom flask, and ground K2CO3 (0.21 g, 1.5 mmol) was added, followed by stirring at room temperature. Bromoacetic acid (0.08 g, 0.55 mmol) was slowly added to the system, and TLC was used for monitoring. After the reaction was complete, the reaction system was poured into 50 ml of water, and the system was adjusted to weak acidity with dilute hydrochloric acid. The mixture was extracted three times with 50 ml of ethyl acetate. The organic phases were combined, washed with saturated NaCl solution (50 ml), dried over anhydrous sodium sulfate, and filtered. The filtrate was concentrated under reduced pressure and then purified by column chromatography (MeOH:DCM = 1:30) to obtain the product, which was recrystallized from ethyl acetate to obtain compound 1 (0.13 g) with a yield of 76.9%. The spectral data of compound 1 are as follows: 1 H NMR (400 MHz, DMSO-d6) δ 7.48 (dd, J = 9.1, 3.5 Hz, 1H), 7.37 (d, J = 5.8 Hz, 1H), 7.23 (d, J = 3.2 Hz, 1H), 7.00 (dd, J = 5.1, 3.5 Hz, 1H), 5.44 (s, 1H), 4.08 (s, 1H). 13 C NMR (100 MHz, DMSO-d6) δ 169.19, 162.66, 157.39, 156.96, 136.99, 128.47, 126.73, 126.60, 122.89, 121.76, 119.79, 42.28, 35.97.

[0099] Example 20. Preparation of Compound 2

[0100]

[0101] The operation was the same as in Example 19, except that compound 2-M2 (0.14 g, 0.5 mmol) was reacted with bromoacetic acid (0.08 g, 0.55 mmol), and the product was recrystallized from ethyl acetate to obtain compound 2 (0.13 g) with a yield of 78.7%. The spectral data of compound 2 are as follows: 1 HNMR (400 MHz, DMSO-d6) δ 12.93 (s, 1H), 8.18 (d, J = 5.2 Hz, 1H), 7.47 (d, J = 5.1 Hz, 1H), 7.24 (d, J = 5.2 Hz, 1H), 7.23 (d, J = 3.1 Hz, 1H), 6.99 (dd, J = 5.0, 3.6 Hz, 1H), 5.46 (s, 2H), 4.10 (s, 2H). 1313C NMR (100 MHz, DMSO-d6) δ 169.21, 157.67, 156.88, 155.53, 136.94, 136.45, 128.51, 126.81, 126.64, 124.38, 118.45, 42.34, 34.62.

[0102] Example 21. Preparation of Compound 3

[0103]

[0104] The operation was the same as in Example 19, except that compound 3-M2 (0.15 g, 0.5 mmol) was reacted with bromoacetic acid (0.08 g, 0.55 mmol), and recrystallized from ethyl acetate to obtain compound 3 (0.15 g) with a yield of 86.2%. The spectral data of compound 3 are as follows: 1 1H NMR (400 MHz, DMSO-d6) δ 12.91 (s, 1H), 7.47 (dd, J = 5.1, 1.2 Hz, 1H), 7.22 (d, J = 3.4 Hz, 1H), 7.07 (d, J = 1.2 Hz, 1H), 7.00 (dd, J = 5.1, 3.5 Hz, 1H), 5.41 (s, 2H), 4.06 (s, 2H), 2.46 (d, J = 1.0 Hz, 3H). 13 13C NMR (100 MHz, DMSO-d6) δ 169.16, 163.11, 157.56, 156.64, 137.02, 133.61, 128.47, 126.74, 126.61, 118.23, 117.33, 42.11, 34.54, 16.09.

[0105] Example 22. Preparation of Compound 4

[0106]

[0107] The operation was the same as in Example 19, except that compound 4-M2 (0.15 g, 0.5 mmol) was reacted with bromoacetic acid (0.08 g, 0.55 mmol), and recrystallized from ethyl acetate to obtain compound 4 (0.15 g) with a yield of 84.6%. The spectral data of compound 4 are as follows: 1 1H NMR (400 MHz, DMSO-d6) δ 12.84 (s, 1H), 7.84 (d, J = 1.1 Hz, 1H), 7.48 (dd, J = 5.1, 1.2 Hz, 1H), 7.23 (dd, J = 3.5, 1.0 Hz, 1H), 7.00 (dd, J = 5.1, 3.5 Hz, 1H), 5.45 (s, 2H), 4.06 (s, 2H), 2.25 (d, J = 1.0 Hz, 3H).13 C NMR (100 MHz, DMSO-d6) δ 169.37, 157.46, 157.03, 154.42, 136.99, 132.92, 131.06, 128.50, 126.81, 126.64, 118.13, 42.28, 34.82, 12.08.

[0108] Example 23. Preparation of Compound 5

[0109]

[0110] The operation was the same as in Example 19, except that compound 5-M2 (0.14 g, 0.5 mmol) was reacted with bromoacetic acid (0.08 g, 0.55 mmol), and recrystallized from ethyl acetate to obtain compound 5 (0.15 g) with a yield of 88.7%. The spectral data of compound 5 are as follows: 1 1H NMR (400 MHz, DMSO-d6) δ 12.86 (s, 1H), 8.54 (d, J = 3.2 Hz, 1H), 7.60 (d, J = 3.2 Hz, 1H), 7.46 (dd, J = 5.1, 1.2 Hz, 1H), 7.20 (d, J = 3.3 Hz, 1H), 6.99 (dd, J = 5.1, 3.5 Hz, 1H), 5.38 (s, 2H), 4.04 (s, 2H). 13 C NMR (100 MHz, DMSO-d6) δ 169.34, 157.01, 153.80, 145.91, 137.88, 129.26, 128.04, 126.64, 126.53, 123.13, 116.50, 41.59, 34.50.

[0111] Example 24. Preparation of Compound 6

[0112]

[0113] The operation was the same as in Example 19, except that compound 6-M2 (0.17 g, 0.5 mmol) was reacted with bromoacetic acid (0.08 g, 0.55 mmol), and recrystallized from ethyl acetate to obtain compound 6 (0.17 g) with a yield of 85.3%. The spectral data of compound 6 are as follows: 1HNMR(400MHz, DMSO-d6) δ 12.97 (s, 1H), 8.18 (dd, J = 17.4, 7.9 Hz, 2H), 7.71–7.64 (m, 1H), 7.61 (t, J = 7.5 Hz, 1H), 7.50 (dd, J = 5.1, 1.1 Hz, 1H), 7.28 (d, J = 2.8 Hz, 1H), 7.02 (dd, J = 5.1, 3.5 Hz, 1H), 5.51 (s, 2H), 4.16 (s, 2H). 13 C NMR(100MHz, DMSO-d6) δ 169.56, 159.03, 157.38, 150.85, 140.71, 136.61, 133.34, 129.51, 128.77, 126.99, 126.69, 125.56, 123.98, 123.33, 117.91, 42.79, 35.06.

[0114] Example 25. Preparation of Compound 7

[0115]

[0116] The operation was the same as in Example 19, except that compound 7-M2 (0.14 g, 0.5 mmol) was reacted with bromoacetic acid (0.08 g, 0.55 mmol), and recrystallized from ethyl acetate to obtain compound 7 (0.15 g) with a yield of 89.3%. The spectral data of compound 7 are as follows: 1 HNMR(400MHz, DMSO-d6) δ 12.93 (s, 1H), 7.51 (d, J = 3.8 Hz, 1H), 7.48 (d, J = 5.7 Hz, 1H), 7.44 (s, 1H), 7.35 (d, J = 5.7 Hz, 1H), 7.13 (d, J = 4.5 Hz, 1H), 5.29 (s, 2H), 4.06 (s, 2H). 13 C NMR(100MHz, DMSO-d6) δ 169.18, 162.51, 157.20, 157.08, 135.75, 127.40, 126.69, 123.75, 122.89, 121.84, 119.96, 42.82, 34.56.

[0117] Example 26. Preparation of Compound 8

[0118]

[0119] The operation was the same as that in Example 19, except that compound 9-M2 (0.15 g, 0.5 mmol) was reacted with bromoacetic acid (0.08 g, 0.55 mmol), and recrystallized from ethyl acetate to obtain compound 8 (0.15 g) with a yield of 84.6%. The spectral data of compound 8 are as follows: 1 HNMR (400 MHz, DMSO-d6) δ 12.91 (s, 1H), 7.51 (dd, J = 4.8, 3.0 Hz, 1H), 7.44 (s, 1H), 7.13 (d, J = 4.9 Hz, 1H), 7.05 (s, 1H), 5.26 (s, 2H), 4.04 (s, 2H), 2.45 (s, 3H). 13 C NMR (100 MHz, DMSO-d6) δ 169.19, 163.10, 157.69, 156.98, 135.88, 133.66, 127.43, 126.64, 123.66, 118.27, 117.13, 42.57, 34.49, 16.09.

[0120] Example 27. Preparation of compound 9

[0121]

[0122] The operation was the same as that in Example 19, except that compound 11-M2 (0.14 g, 0.5 mmol) was reacted with bromoacetic acid (0.08 g, 0.55 mmol), and recrystallized from ethyl acetate to obtain compound 9 (0.15 g) with a yield of 90.3%. The spectral data of compound 9 are as follows: 1 HNMR (400 MHz, DMSO-d6) δ 12.84 (s, 1H), 8.50 (d, J = 3.2 Hz, 1H), 7.60 (d, J = 3.2 Hz, 1H), 7.50 (dd, J = 4.9, 3.0 Hz, 1H), 7.41 (d, J = 1.7 Hz, 1H), 7.11 (d, J = 4.9 Hz, 1H), 5.23 (s, 2H), 4.02 (s, 2H). 13 C NMR (100 MHz, DMSO-d6) δ 169.37, 157.14, 154.21, 146.03, 136.64, 129.04, 127.31, 126.60, 123.34, 123.30, 116.29, 42.09, 34.45.

[0123] Example 28. Preparation of compound 10

[0124]

[0125] The operation was the same as in Example 19, except that compound 8-M2 (0.14 g, 0.5 mmol) was reacted with bromoacetic acid (0.08 g, 0.55 mmol), and recrystallized from ethyl acetate to obtain compound 10 (0.14 g) with a yield of 82.5%. The spectral data of compound 10 are as follows: 1 H NMR (400 MHz, DMSO-d6) δ 12.90 (s, 1H), 8.19 (d, J = 5.2 Hz, 1H), 7.52 (dd, J = 5.0, 3.0 Hz, 1H), 7.45 (dd, J = 2.7, 1.0 Hz, 1H), 7.26 (d, J = 5.2 Hz, 1H), 7.13 (dd, J = 5.0, 1.2 Hz, 1H), 5.31 (s, 2H), 4.07 (s, 2H). 13 C NMR (100 MHz, DMSO-d6) δ 169.28, 158.00, 157.04, 155.53, 136.23, 135.81, 127.43, 126.71, 124.38, 123.80, 118.58, 42.83, 34.58.

[0126] Example 29. Preparation of compound 11

[0127]

[0128] The operation was the same as in Example 19, except that compound 10-M2 (0.15 g, 0.5 mmol) was reacted with bromoacetic acid (0.08 g, 0.55 mmol), and recrystallized from ethyl acetate to obtain compound 11 (0.16 g) with a yield of 92.2%. The spectral data of compound 11 are as follows: 1 H NMR (400 MHz, DMSO-d6) δ 12.83 (s, 1H), 7.83 (d, J = 1.1 Hz, 1H), 7.52 (dd, J = 5.2, 3.1 Hz, 1H), 7.44 (dd, J = 3.1, 1.3 Hz, 1H), 7.12 (dd, J = 5.2, 1.4 Hz, 1H), 5.29 (s, 2H), 4.04 (s, 2H), 2.26 (d, J = 0.8 Hz, 3H). 13 C NMR (100 MHz, DMSO-d6) δ 169.42, 157.77, 157.18, 154.40, 135.83, 132.88, 130.82, 127.43, 126.69, 123.81, 118.25, 42.74, 34.78, 12.08.

[0129] Example 30. Preparation of compound 12

[0130]

[0131] The operation was the same as that in Example 19, except that compound 12-M2 (0.17 g, 0.5 mmol) was reacted with bromoacetic acid (0.08 g, 0.55 mmol), and recrystallized from ethyl acetate to obtain compound 12 (0.17 g) with a yield of 89.7%. Spectral data of compound 12: 1 H NMR (400 MHz, DMSO-d6) δ 8.21 (d, J = 7.6 Hz, 1H), 8.15 (d, J = 8.1 Hz, 1H), 7.71–7.64 (m, 1H), 7.64–7.58 (m, 1H), 7.54 (dd, J = 4.9, 3.0 Hz, 1H), 7.50 (dd, J = 2.7, 1.1 Hz, 1H), 7.17 (dd, J = 5.0, 1.3 Hz, 1H), 5.35 (s, 2H), 4.14 (s, 2H). 13 C NMR (100 MHz, DMSO-d6) δ 169.60, 159.33, 157.54, 150.83, 140.68, 135.51, 133.42, 129.42, 127.49, 126.79, 125.50, 124.03, 123.93, 123.30, 118.03, 43.26, 35.01.

[0132] Example 31. Screening test for the inhibitory effect of compounds on uric acid transporter hURAT1

[0133] Test materials and methods:

[0134] (1) Cell line: Human uric acid transporter (hURAT1) highly expressed cell line (HEK293-URAT1).

[0135] (2) Experimental conditions:

[0136]

[0137] (3) Preparation of test drugs:

[0138] 1) Preparation of 20 mmol / L stock solution

[0139] Each compound was prepared into a 20 mmol / L stock solution using DMSO as the solvent.

[0140] 2) Preparation of working solution

[0141] ① Single concentration: (1 μmol / L Using DMSO as the solvent, the stock solutions of No. 1-12 were diluted to a 200-fold working solution of the administration concentration, i.e., 200 μmol / L, and the stock solution of Lesinurad (Les, positive control) was diluted to 200 μmol / L.

[0142] ②IC50: Using DMSO as the solvent, dilute the stock solutions of No. 3 and No. 4 by 200 times to prepare working solutions with concentrations of 6000, 2000, 600, 200, 60, and 20 μmol / L.

[0143] ③Using HBSS (without Cl-) buffer solution as the solvent, dilute the 200-fold working solutions in steps ① and ② by 100 times to obtain 2-fold working solutions with various dosing concentrations.

[0144] ④Prepare a 2-fold working solution of the radiolabeled substrate C-Uric acid using HBSS (without Cl-) buffer solution, and then mix it with the 2-fold working solutions in step ③ in equal volumes to obtain the dosing working solutions.

[0145] ④Prepare the radiolabeled substrate 14 C-Uric acid 2-fold working solution, and then mix it with the 2-fold working solutions in step ③ in equal volumes to obtain the dosing working solutions.

[0146] (4) Administration method

[0147] After the cell line (HEK293-URAT1) expressing hURAT1 is resuscitated and passaged, select the well-grown adherent cells, digest them with trypsin into single-cell suspensions, adjust the cell density to about 1.5×105 cells / mL with the medium, inoculate 1 mL / well into a 24-well cell culture plate, and culture in an incubator at 37°C, 5% CO2, and saturated air humidity for 2 - 3 days.

[0148] After the cells cover each well, remove the culture medium, wash twice with pre-warmed HBSS at 37°C, add 1 mL of HBSS buffer at 37°C to each well and incubate for 10 min. Then replace the HBSS with 500 μL of the dosing working solution containing the radiolabeled probe substrate; after 2 min, terminate the reaction with cold HBSS buffer, add 300 μL of 0.1 mol / L NaOH to each well to lyse the cells; take the cell lysate into a scintillation vial, add 1.5 mL of scintillation fluid, and measure the radioactivity intensity in the sample using a Tri-Carb 2910TR liquid scintillation counter. Set up 2 replicates (n = 2) for each compound, positive control, and blank control.

[0149] (5) Data processing

[0150] Define the transport value Uc of the dosing group (control) cells containing only the radiolabeled substrate as 100%, and calculate the percentage (In) of the transport value U of each dosing group after adding the test compound to Uc based on this standard, that is, the transport ratio. Use this transport ratio to characterize the inhibitory effect of the compound on the transporter (the lower the transport ratio In, the stronger the activity). The formula is as follows:

[0151]

[0152] Each group was set with 2 replicate wells (n = 2), and Mean±standard error (SD) was calculated using Excel statistical formulas, and IC was calculated using GraphPad Prism software 50 .

[0153] Test results:

[0154] (1) Single-concentration activity inhibition

[0155] As Figure 1 shown, under the condition of a single concentration of 1 μmol / L, compounds 1-12 all had different inhibitory effects on hURAT1, and the activity of hURAT1 could be inhibited to 82.30%, 81.15%, 61.97%, 58.89%, 76.13%, 78.90%, 66.77%, 67.63%, 79.93%, 65.81%, 72.15%, 87.47% respectively. Compared with the positive control drug Lesinurad (80.03%) at the same concentration (1 μmol / L), the inhibitory activities of 10 of these compounds were better than or equivalent to Lesinurad

[0156] (2) IC 50 test results

[0157] Compounds 3 and 4 showed strong inhibitory activities on hURAT1, and inhibitory effect IC of these two compounds was 50 tested. As Figure 2 and Figure 3 shown, the inhibitory effect IC of compounds 3 and 4 on hURAT1 50 was 3.097 and 1.514 μmol / L respectively, and the activity was significantly better than that of the positive control Lesinurad (IC 50 = 7.3 μmol / L).

[0158] Conclusion: The single-concentration activity inhibition test showed that compounds 1-12 all had different inhibitory effects on hURAT1, and the inhibitory activities of 10 of these compounds were better than or equivalent to Lesinurad; compounds 3 and 4 had strong inhibitory activities, and the inhibitory effect IC of compounds 3 and 4 on hURAT1 50 was 3.097 and 1.514 μmol / L respectively, and the activity was significantly better than that of the positive control Lesinurad (IC 50 = 7.3 μmol / L), and they can be used as candidate drugs for the treatment of gout and hyperuricemia

Claims

1. A thiophenopyrimidine compound containing a thiophene methylene structure or a pharmaceutically acceptable salt thereof, characterized in that, It has the structure shown by the following general formula I: Among them, Ar1 is thiophene or thiophene substituted by one or more R2; Ar2 is thiophene; the R2 is selected from H, C 1-6 alkyl or a benzene ring formed by two R2 together with the two carbon atoms to which they are attached; R is selected from methylene, isopropyl, cyclopropyl, cyclobutyl.

2. The thiophenopyrimidine compound containing a thiophene methylene structure or a pharmaceutically acceptable salt thereof according to claim 1, characterized in that, Selected from the following formulas I-1 to I-5:

3. The thiophenopyrimidine compound containing a thiophene methylene structure or a pharmaceutically acceptable salt thereof according to claim 1, characterized in that, Selected from the following formulas I-1' to I-5':

4. The thiophenopyrimidine compound containing a thiophene methylene structure or a pharmaceutically acceptable salt thereof according to claim 1, characterized in that, The connection mode of Ar2 and methylene is thiophen-2-yl or thiophen-3-yl.

5. The thiophenopyrimidine compound containing a thiophene methylene structure or a pharmaceutically acceptable salt thereof according to claim 1, characterized in that, Selected from the following compounds:

6. The preparation method of the thiophenopyrimidine compound containing a thiophene methylene structure as described in claim 1, characterized in that, Comprising the following steps: (1) Using aminothiophenecarboxylate or substituted aminothiophenecarboxylate as raw materials, dichloromethane as solvent, and triethylamine as acid-binding agent, reacting with thiophosgene to generate isothiocyanate (M1); (2) Using tetrahydrofuran as solvent, M1 reacts with thiophene methylamine under the action of triethylamine to generate a thiourea intermediate, and then cyclizes under the action of potassium hydroxide to generate a thieno[2,3-d]pyrimidin-4(3H)-one intermediate (M2); (3) Using DMF as solvent, M2 reacts with 2-bromocarboxylic acid under the action of potassium carbonate to generate the target compound; 7. Use of the thieno[2,3-d]pyrimidine compound containing a thiophenemethylene structure according to claim 5 or a pharmaceutically acceptable salt thereof in the preparation of a drug for reducing uric acid.

8. A drug composition for reducing uric acid, comprising the thieno[2,3-d]pyrimidine compound containing a thiophenemethylene structure according to claim 7 or a pharmaceutically acceptable salt thereof and one or more pharmaceutically acceptable carriers or excipients.

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