Pyrimido pyrimidinedione compounds, and preparation method and application thereof

Abstract

The invention relates to the field of medicinal chemistry and pharmacotherapeutics, in particular to a pyrimidinopyrimidinedione compound with a structure shown in a general formula (I) or pharmaceutically acceptable salt or solvate or stereoisomer or prodrug molecule of the pyrimidinopyrimidinedione compound, the invention also relates to a preparation method of the compound and application of the compound or pharmaceutically acceptable salts, solvates, prodrug molecules and pharmaceutical compositions of the compound in preparation of drugs for treating neurological diseases and the like. # imgabs0 #

Description

Technical Field

[0001] The present invention relates to the fields of medicinal chemistry and pharmacotherapy, and in particular to a pyrimidopyrimidinedione compound, a preparation method thereof, and use thereof as a potassium channel agonist in the preparation of a medicament for treating epilepsy, developmental and epileptic encephalopathy, convulsions, neuropathic pain, anxiety, depression, bipolar disorder, autism, and neurodegenerative diseases. Background Art

[0002] The KCNQ potassium channel family is a family of voltage-gated potassium channels, encompassing five subtypes, KCNQ1, KCNQ5, and KCNQ2. They participate in the regulation of many important physiological functions and play a crucial role in maintaining cellular excitability and ion homeostasis. KCNQ2 and KCNQ3, which can form homo- or heterotetramers, are the primary molecular basis for the formation of neuronal M currents.

[0003]

[0004] Retigabine (RTG), the first KCNQ2 channel agonist approved by the US Food and Drug Administration (FDA) for the treatment of epilepsy in 2011, is primarily used as an adjunctive treatment for refractory partial-onset seizures in adults. It remains the only approved drug targeting KCNQ ion channels to date. Its mechanism of action is to activate KCNQ2 / 3 channels in the brain, thereby maintaining a sustained efflux of potassium ions, causing membrane hyperpolarization, which in turn reduces neuronal excitability and ultimately produces an anti-epileptic effect. This mechanism of action differs from that of previous anti-epileptic drugs. However, after its launch, retigabine was found to cause fundus abnormalities similar to those seen in retinal pigment dystrophy, which can lead to photoreceptor damage and vision loss. Consequently, the FDA imposed a black box warning on the drug in 2013, recommending its use only in patients who have failed or are intolerant to other anti-epileptic drugs. Subsequent long-term monitoring revealed that after four years of treatment, retinal pigment abnormalities were detectable in approximately one-third of patients, and some patients experienced recovery after discontinuation of the drug. In addition, some patients also observed symptoms such as macular abnormalities characterized by yolk-like lesions and bluish skin color (S. Clark et al., Ther. Adv. Drug Saf. 2015, 6 (1), 15-19). Although the drug has not been withdrawn from the market by the FDA, GlaxoSmithKline announced in June 2017 that it would permanently stop producing the drug due to limited applications and market shrinkage. Studies have shown that the pigment abnormalities caused by retigabine are due to its metabolism in the body to form dimeric pigments, which is closely related to the chemical structure of retigabine itself and has nothing to do with its target mechanism of action (C. Bock et al., Future Med. Chem. 2019, 11 (4), 337-355).

[0005] In addition to epilepsy, modulation of KCNQ2 channel function is a potential therapeutic strategy for many other diseases, such as developmental and epileptic encephalopathy, convulsions, neuropathic pain, anxiety, depression, bipolar disorder, autism, and neurodegenerative diseases, all of which are characterized by neuronal hyperexcitability (W. Dalby-Brown et al., Curr. Top. Med. Chem. 2006, 6(10), 999-1023).

[0006] In summary, the development of agonists that act on the KCNQ2 target but have a structure significantly different from retigabine will potentially provide safer and more effective innovative drugs for the clinical treatment of neurological diseases, which has important economic and social value. Summary of the Invention

[0007] One of the objects of the present invention is to provide a pyrimido[4,5-d]pyrimidine-2,4(1H,3H)-dione compound or a pharmaceutically acceptable salt or solvate thereof or a prodrug molecule thereof.

[0008] A second object of the present invention is to provide a method for preparing a pyrimido[4,5-d]pyrimidine-2,4(1H,3H)-dione compound or a pharmaceutically acceptable salt or solvate thereof or a prodrug molecule thereof.

[0009] A third object of the present invention is to provide a pyrimido[4,5-d]pyrimidine-2,4(1H,3H)-dione compound or a pharmaceutically acceptable salt thereof or a solvate thereof or a prodrug molecule thereof for use in preparing a novel KCNQ potassium channel agonist, particularly in preparing a pharmaceutical preparation for anti-epileptic, neuropathic pain relief, antianxiety, depression, bipolar disorder, autism and neurodegenerative diseases.

[0010] One of the objectives of the present invention is achieved through the following technical solutions:

[0011] In one aspect, the pyrimido[4,5-d]pyrimidine-2,4(1H,3H)-dione compound of the present invention, or a pharmaceutically acceptable salt thereof, or a solvate thereof, or a prodrug molecule thereof, has a structure represented by the general formula (I):

[0012]

[0013] Among them, R 1 、R 2 、R 3 Each independently selected from C1-C8 alkyl, R 4 and R 5 Each is independently selected from hydrogen, C1-C8 alkyl.

[0014] Some representative compounds of the pyrimido[4,5-d]pyrimidine-2,4(1H,3H)-dione compounds or pharmaceutically acceptable salts or solvates thereof or prodrug molecules of the present invention are as follows:

[0015]

[0016] According to the above description, only three specific compounds having the general formula (I) are cited to explain the above general formula in more detail, but they are not exhaustive of the compounds of the present invention.

[0017] The pharmaceutically acceptable salt of the compound of the present invention is a salt formed by the compound and an acid, wherein the acid is selected from maleic acid, succinic acid, citric acid, tartaric acid, fumaric acid, formic acid, acetic acid, propionic acid, malonic acid, oxalic acid, stearic acid, benzoic acid, phthalic acid, methanesulfonic acid, benzenesulfonic acid, toluenesulfonic acid, naphthalenesulfonic acid, 1,5-naphthalenedisulfonic acid, camphoric acid, camphorsulfonic acid, salicylic acid, acetylsalicylic acid, pamoic acid, aspartic acid, glutamic acid, lactic acid, gluconic acid, ascorbic acid, gallic acid, mandelic acid, malic acid, sorbic acid, trifluoroacetic acid, taurine, homotaurine, 2-hydroxyethanesulfonic acid, cinnamic acid, hydrochloric acid, hydrobromic acid, hydroiodic acid, sulfuric acid, nitric acid, phosphoric acid or perchloric acid.

[0018] The second object of the present invention is achieved through the following technical solutions:

[0019] A method for preparing a pyrimido[4,5-d]pyrimidine-2,4(1H,3H)-dione compound or a pharmaceutically acceptable salt or solvate thereof or a prodrug molecule thereof can be one of the following two preparation methods:

[0020] Method 1:

[0021] The method comprises two steps. The first step is: an acyl chloride A reacts with thiocyanide to in situ generate an acylthiocyanate B; an aminopyrimidinedione C is then added to the reaction system for condensation to obtain a thioamide D. The obtained thioamide D generally also contains a further ring-closed pyrimidopyrimidinedione E; the second step is: a mixture of the thioamide D and the pyrimidopyrimidinedione E obtained in the first step is subjected to a substitution reaction with a chloroamide F to obtain a compound represented by the general formula (I). In the second step, the thioamide D first undergoes a ring-closure reaction to generate the pyrimidopyrimidinedione E, which then undergoes a further substitution reaction.

[0022]

[0023] Method 2: Synthesis of compound salts,

[0024] The compound represented by general formula (I) of the present invention can be converted into its pharmaceutically acceptable salt by the following method: adding a solution of the corresponding acid to a solution of the above compound, and removing the solvent under reduced pressure after the salt is completely formed to obtain the corresponding salt of the compound of the present invention.

[0025] The third object of the present invention is achieved through the following technical solutions:

[0026] Disclosed is a use of a pyrimido[4,5-d]pyrimidine-2,4(1H,3H)-dione compound or a pharmaceutically acceptable salt or solvate thereof or a prodrug molecule thereof in preparing a novel KCNQ potassium channel agonist, particularly in preparing a medicament for treating neurological diseases, including epilepsy, developmental and epileptic encephalopathy, convulsions, neuropathic pain, anxiety, depression, bipolar disorder, autism, and neurodegenerative diseases.

[0027] Compared with the prior art, the present invention has the following beneficial effects:

[0028] The novel KCNQ potassium channel agonist of the pyrimido[4,5-d]pyrimidine-2,4(1H,3H)-dione compound described in the present invention is more stable than the existing drug retigabine and is found to have no deterioration or discoloration after being placed in air at room temperature for more than one year.

[0029] The pyrimido[4,5-d]pyrimidine-2,4(1H,3H)-dione compounds of the present invention have KCNQ2 potassium channel agonist activity similar to or superior to that of retigabine. DETAILED DESCRIPTION

[0030] The present invention will be further described in detail below with reference to specific examples, but the present invention is not limited to these examples.

[0031] 1. Preparation Examples of Compounds

[0032] Preparation Example 1: Synthesis of 7-n-butyl-1,3-dimethyl-5-(2-oxo-2-aminoethylthio)pyrimido[4,5-d]pyrimidine-2,4(1H,3H)-dione

[0033]

[0034] To a mixture of KSCN (9.7 g, 100 mmol) and acetone (500 mL) was added n-valeryl chloride (12 mL, 100 mmol) dropwise with stirring at room temperature. Stirring was continued for 1 hour after the addition was complete. A solution of the commercial reagent 6-amino-1,3-dimethyluracil (15.5 g, 100 mmol) in DMF (150 mL) was added to the reaction mixture, and stirring was continued at room temperature for 12 hours. The mixture was filtered through celite, and the filtrate was concentrated under reduced pressure to remove the acetone. Purified water (450 mL) was added to the resulting concentrate. Solids gradually precipitated with stirring at room temperature. After stirring for 30 minutes, the mixture was filtered to yield 4.5 g of a yellow powder, a mixture of 6-amino-1,3-dimethyl-5-(n-valerylaminothioformyl)uracil and 7-n-butyl-5-mercapto-1,3-dimethylpyrimido[4,5-d]pyrimidine-2,4(1H,3H)-dione.

[0035] To a mixture of 0.45 g of the above mixture, anhydrous KCO (0.25 g, 1.80 mmol), and DMF (2.5 mL) was added a solution of the commercial reagent 2-chloroacetamide (0.21 g, 2.2 mmol) in DMF (2.5 mL) dropwise with stirring at room temperature. Stirring was continued for 17 hours after the addition was complete. Dilute with water (15 mL), filter, and recrystallize the filter cake from methanol to obtain 0.26 g of 7-n-butyl-1,3-dimethyl-5-(2-oxo-2-aminoethylthio)pyrimido[4,5-d]pyrimidine-2,4(1H,3H)-dione as an off-white solid.

[0036] 1 H NMR(300MHz, CDCl3)δ6.53(br,1H),5.29(br,1H),3.89(s,2H),3.67(s,3H),3.4 6(s,3H),2.92(t,J=7.5Hz,2H),1.82(m,2H),1.40(m,2H),0.96(t,J=7.2Hz,3H).

[0037] HRMS(ESI):m / z 338.1261[M+H] + (calcd for C 14 H 20 N5O3S + Preparation Example 2: Synthesis of 7-isopropyl-1,3-dimethyl-5-(2-oxo-2-diethylaminoethylthio)pyrimido[4,5-d]pyrimidine-2,4(1H,3H)-dione

[0038]

[0039] To a mixture of KSCN (1.94 g, 20 mmol) and acetone (160 mL) was added isobutyryl chloride (2.07 mL, 20 mmol) dropwise with stirring at room temperature. Stirring was continued for 1 hour after the addition was complete. A solution of the commercial reagent 6-amino-1,3-dimethyluracil (3.1 g, 20 mmol) in DMF (40 mL) was added to the reaction mixture, and stirring was continued at room temperature for 16 hours. The mixture was filtered through celite, and the filtrate was concentrated under reduced pressure to remove the acetone. Purified water (150 mL) was added to the resulting concentrate. Solids gradually precipitated with stirring at room temperature. After stirring for 30 minutes, the mixture was filtered to yield 2.03 g of a yellow powder, a mixture of 6-amino-1,3-dimethyl-5-(isobutyrylaminothioformyl)uracil and 7-isopropyl-5-mercapto-1,3-dimethylpyrimido[4,5-d]pyrimidine-2,4(1H,3H)-dione.

[0040] To a mixture of 0.42 g of the above mixture, anhydrous K₂CO₃ (0.25 g, 1.8 mmol), and DMF (3 mL) was added dropwise a solution of the commercial reagent N,N-diethylchloroacetamide (0.27 g, 1.8 mmol) in DMF (2 mL) with stirring at room temperature. Stirring was continued for 20 hours after the addition was complete. The mixture was diluted with water (15 mL) and extracted with ethyl acetate (25 mL x 3). The combined organic phases were dried over anhydrous Na₂SO₄, filtered, and the filtrate concentrated to yield a pale yellow oil. Column chromatography (dichloromethane / methanol = 20:1 v / v) and recrystallization from methanol afforded 0.39 g of 7-isopropyl-1,3-dimethyl-5-(2-oxo-2-diethylaminoethylthio)pyrimido[4,5-d]pyrimidine-2,4(1H,3H)-dione as a white solid.

[0041] 1 H NMR(300MHz, CDCl3)δ4.13(s,2H),3.66(s,3H),3.49-3.38(m,4H),

[0042] 3.44(s,3H),3.09(m,1H),1.31(d,J=6.9Hz,6H),1.27(m,3H),1.40(t,

[0043] J = 6.6 Hz, 3H).

[0044] HRMS(ESI):m / z 380.1757[M+H] + (calcd for C 17 H 26 N5O3S +:380.1751) Preparation Example 3: Synthesis of 3,7-di-n-butyl-1-methyl-5-(2-oxo-2-dimethylaminoethylthio)pyrimido[4,5-d]pyrimidine-2,4(1H,3H)-dione

[0045]

[0046] To a mixture of KSCN (0.65 g, 6.6 mmol) and acetone (55 mL) was added n-valeryl chloride (0.80 mL, 6.6 mmol) dropwise with stirring at room temperature. Stirring was continued for 1 hour after the addition was complete. A solution of 6-amino-3-n-butyl-1-methyluracil (1.30 g, 6.6 mmol, prepared according to Journal of Medicinal Chemistry 2008, 51, 2267-2278) in DMF (15 mL) was added to the reaction mixture and stirring was continued at room temperature for 12 hours. The mixture was filtered through celite, and the filtrate was concentrated under reduced pressure to remove the acetone. Pure water (45 mL) was added to the obtained concentrated solution, and solids gradually precipitated under stirring at room temperature. After stirring for 30 minutes, the mixture was filtered to obtain 0.45 g of a yellow powder, which was a mixture of 6-amino-3-n-butyl-1-methyl-5-(n-pentanoylaminothioformyl)uracil and 3,7-di-n-butyl-5-mercapto-1-methylpyrimido[4,5-d]pyrimidine-2,4(1H,3H)-dione.

[0047] To a mixture of 0.34 g of the above mixture, anhydrous KCO (0.17 g, 1.2 mmol), and DMF (3 mL) was added a solution of the commercial reagent N,N-dimethylchloroacetamide (0.15 g, 1.2 mmol) in DMF (2 mL) with stirring at room temperature. Stirring was continued for 20 hours after the addition was complete. Water (25 mL) was added to dilute the mixture, washing out a light yellow powdery solid. Filtering and recrystallization from methanol afforded 0.23 g of 3,7-di-n-butyl-1-methyl-5-(2-oxo-2-dimethylaminoethylthio)pyrimido[4,5-d]pyrimidine-2,4(1H,3H)-dione as a white solid.

[0048] 1 H NMR (300MHz, CDCl3) δ4.09 (s, 2H), 4.02 (t, J = 7.2Hz, 2H), 3.63 (s,

[0049] 3H),3.19(s,3H),3.00(s,3H),2.84(t,J=7.2Hz,2H),1.78(m,2H),

[0050] 1.63(m,2H),1.39(brs,4H),0.96-0.95(m,6H).

[0051] HRMS(ESI):m / z 408.2076[M+H] + (calcd for C 19 H 30 N5O3S + :408.2064). 2. Target Test Experimental Example: FLIPR Thallium Ion Flux Assay of the Compounds of the Present Invention on KCNQ2 Potassium Channels Expressed in Chinese Hamster Ovary Cells (CHO)

[0052] 1. Thallium flux assay to test the effects of compounds

[0053] The CHO-KCNQ2 stable cell line was routinely cultured in DMEM / F12 medium containing 10% FBS and 500 μg / mL G418. The day before the experiment, CHO-KCNQ2 cells were seeded at 12,000 cells per well in a black bottom transparent 384-well cell culture plate (Nunc, USA) and incubated overnight at 37°C with 5% CO2. For the determination of thallium ion flux, the medium in the cell plate was replaced with 20 uL / well thallium dye solution dissolved in HBSS buffer (Hank's balanced salt solution, Gibco, 14175, supplemented with 20 mM HEPES, pH 7.4) and the cell plate was incubated in the dark at room temperature for 1 hour according to the manufacturer's instructions. The compounds to be tested were prepared in a dose-dependent manner (8 doses, with the highest dose starting at 30 μM and diluted 3 times), transferred to the cell plate, and incubated with the cells for 20 minutes. The real-time fluorescence detection and analysis system (FLIPR Tetra , Molecular Devices), record the basal fluorescence signal for 10 seconds (F0), then transfer the KCNQ2 channel stimulation buffer containing 2.5mM Tl2SO4, 2.5mM K2SO4 to the cell plate, and record the KCNQ2 fluorescence signal for 120 seconds (F0). For data analysis, the relative fluorescence signal F / F0 is output from each well. The activity (%) of the test compound is defined as follows: 100*[(F / F0 test compound-F / F0 control) / F / F0 control]. If the activity of the test compound is greater than 3 times the standard deviation (SD) of the control activity, the compound is designated as a KCNQ2 channel agonist. If a dose-dependent increase in activity is shown, the half-effective activation concentration EC is obtained. 50 .

[0054] 2. Experimental results:

[0055] EC 50It represents the half-maximal activation concentration of the compound on the fluorescence signal in the FLIPR experiment.

[0056] Table 1. Activity results of compounds on KCNQ2 potassium channels

[0057]

[0058] Results and Discussion: From the above functional analysis experimental results of KCNQ2 potassium channels, it can be seen that the affinity of the compounds disclosed in the present invention for KCNQ2 channels is similar to or better than that of retigabine.

Claims

1. A pyrimidopyrimidinedione compound having a structure represented by general formula (I) or a pharmaceutically acceptable salt or solvate thereof or a prodrug molecule thereof, in, R 1 、R 2 、R 3 Each independently selected from C1-C8 alkyl, R 4 and R 5 Each is independently selected from hydrogen, C1-C8 alkyl.

2. The compound according to claim 1, or a pharmaceutically acceptable salt thereof, or a solvate thereof, or a prodrug molecule thereof, characterized in that: The compound is:

3. The compound according to any one of claims 1 and 2, or a pharmaceutically acceptable salt thereof, or a solvate thereof, or a prodrug molecule thereof, characterized in that: The pharmaceutically acceptable salt of the compound is a salt formed by the compound and an acid, wherein the acid is selected from maleic acid, succinic acid, citric acid, tartaric acid, fumaric acid, formic acid, acetic acid, propionic acid, malonic acid, oxalic acid, stearic acid, benzoic acid, phthalic acid, methanesulfonic acid, benzenesulfonic acid, toluenesulfonic acid, naphthalenesulfonic acid, 1,5-naphthalenedisulfonic acid, camphoric acid, camphorsulfonic acid, salicylic acid, acetylsalicylic acid, pamoic acid, aspartic acid, glutamic acid, lactic acid, gluconic acid, ascorbic acid, gallic acid, mandelic acid, malic acid, sorbic acid, trifluoroacetic acid, taurine, homotaurine, 2-hydroxyethanesulfonic acid, cinnamic acid, hydrochloric acid, hydrobromic acid, hydroiodic acid, sulfuric acid, nitric acid, phosphoric acid or perchloric acid.

4. A method for preparing the compound according to any one of claims 1 and 2, characterized in that: The method comprises two steps. The first step is: an acyl chloride A reacts with thiocyanide to generate an acylthiocyanate B in situ; an aminopyrimidinedione C is then added to the reaction system for condensation to obtain a thioamide D. The obtained thioamide D generally contains a further ring-closed pyrimidopyrimidinedione E; the second step is: a mixture of the thioamide D and the pyrimidopyrimidinedione E obtained in the first step is subjected to a substitution reaction with a chloroamide F to obtain a compound represented by the general formula (I); R 1 、R 2 、R 3 、R 4 and R 5 The limitation is the same as that described in any one of claims 1-2.

5. A method for preparing a pharmaceutically acceptable salt of the compound according to claim 3, characterized in that: The compound represented by general formula (I) of the present invention can be converted into its pharmaceutically acceptable salt by the following method: adding a solution of the corresponding acid to a solution of the above compound, and removing the solvent under reduced pressure after the salt is completely formed to obtain the corresponding salt of the compound of the present invention.

6. Use of the compound according to any one of claims 1 to 3, or a pharmaceutically acceptable salt thereof, or a solvate thereof, or a prodrug molecule thereof in the preparation of a KCNQ potassium channel agonist.

7. Use of the compound according to any one of claims 1 to 3, or a pharmaceutically acceptable salt thereof, or a solvate thereof, or a prodrug molecule thereof, in the preparation of a medicament for treating a neurological disease.

8. The use according to claim 7, characterized in that Such neurological disorders include epilepsy, developmental and epileptic encephalopathy, convulsions, neuropathic pain, anxiety, depression, bipolar disorder, autism, and neurodegenerative diseases.

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