A kind of oxopyridazine compound and its preparation method and use

Abstract

The present invention discloses a compound represented by formula (I), a stereoisomer thereof, or a pharmaceutically acceptable salt thereof. The present invention also provides the use of the compound, its stereoisomer or pharmaceutically acceptable salt in the preparation of a drug for treating and / or preventing diseases associated with FXIa receptors, especially in the preparation of a drug for treating and / or preventing cerebrovascular arterial disease and / or peripheral arterial disease, with significant efficacy.

Description

Technical Field

[0001] The present invention relates to the field of pharmaceutical chemistry, and specifically to oxopyridine compounds or their salts, isomers, and preparation methods thereof, as well as use of oxopyridine compounds in preparing drugs for treating and / or preventing diseases associated with FXIa receptors, especially use of oxopyridine compounds in preparing drugs for treating cerebrovascular arterial diseases and / or peripheral arterial diseases. Background Art

[0002] Thromboembolic disease is a disease caused by abnormal blood clots formed in blood vessels during the survival of humans and animals. Clinically, it may manifest as myocardial infarction, stroke, deep vein thrombosis (DVT), pulmonary embolism, atrial fibrillation and cerebral infarction, etc., claiming tens of millions of lives worldwide each year. Coagulation factor XI (FXI) is a plasma serine protease necessary for maintaining the intrinsic pathway. After activation, it generates activated coagulation factor XIa (FXIa), which plays a key role in the amplification of the coagulation cascade reaction. Therefore, drugs targeting FXIa can block the intrinsic pathway and inhibit the amplification of the coagulation cascade reaction, thereby having an anti-thrombotic effect.

[0003] The reported FXIa inhibitors mainly include monoclonal antibodies, antisense oligonucleotides, chemical small molecules, peptides or proteins and peptide mimetics. At present, milvexian, jointly developed by BMS and Johnson & Johnson, has completed the Phase II clinical trial, and the results show that it has a lower risk of bleeding. The Phase I clinical trial of BMS's intravenous small molecule FXIa inhibitor BMS-962122 has been completed and research and development has been suspended. ONO-7684, a small molecule oral FXIa inhibitor developed by Ono Co., Ltd. of Japan, has entered Phase I clinical research. BAY-2433334 developed by Bayer has completed Phase II clinical trials and has become the most promising small molecule FXIa inhibitor. Monoclonal antibodies and antisense oligonucleotides need to be injected, and have the disadvantages of being expensive, slow to take effect and possibly difficult to control. Chemical small molecules have the advantages of relatively good oral bioavailability and better patient compliance.

[0004] Therefore, the development of new FXIa small molecule inhibitors that are safe, effective, specific, and highly active may make up for the shortcomings of current clinical anticoagulant and antithrombotic drugs that are prone to bleeding complications and meet unmet clinical needs. Summary of the invention

[0005] The compound of the present invention is a novel oxopyridine compound, which exhibits good anticoagulant effect and in vitro and in vivo affinity to FXIa.

[0006] In one aspect, the present invention provides a compound represented by formula (I), a stereoisomer or a pharmaceutically acceptable salt thereof:

[0007]

[0008] Furthermore, the above-mentioned pharmaceutically acceptable salt is a metal salt.

[0009] Furthermore, the above-mentioned metal salt is selected from sodium salt, potassium salt, calcium salt, lithium salt and magnesium salt.

[0010] In another aspect, the present invention provides a pharmaceutical composition of the above-mentioned compound, its stereoisomer or pharmaceutically acceptable salt, wherein the composition further contains a pharmaceutically acceptable carrier and / or excipient.

[0011] On the other hand, the present invention provides a method for preparing the above-mentioned compound, its stereoisomer or pharmaceutically acceptable salt, comprising the following route:

[0012]

[0013] On the other hand, the present invention also provides the use of any one of the above compounds, stereoisomers or pharmaceutically acceptable salts thereof or a combination thereof in the preparation of a drug for treating and / or preventing diseases associated with FXIa receptors.

[0014] Furthermore, the above-mentioned disease associated with FXIa receptor is selected from thrombosis or thromboembolism-related diseases.

[0015] Furthermore, the above-mentioned disease associated with FXIa receptor is selected from cerebrovascular arterial disease and / or peripheral arterial disease.

[0016] Furthermore, the above-mentioned cerebrovascular arterial diseases include but are not limited to transient ischemic attack (TIA), ischemic stroke or events of thrombotic and / or thromboembolic origin leading to stroke or TIA; the above-mentioned peripheral arterial diseases include but are not limited to peripheral arterial occlusion, acute limb ischemia, amputation, reocclusion and restenosis after intervention (such as angioplasty, stent implantation or surgery and bypass), and / or stent thrombosis.

[0017] Furthermore, the above-mentioned ischemic stroke includes but is not limited to cardiogenic stroke, non-cardiogenic stroke, stroke caused by large artery or small artery disease, stroke caused by undetermined cause, cryptogenic stroke, embolic stroke or embolic stroke of undetermined source.

[0018] Furthermore, the aforementioned cardiogenic stroke includes but is not limited to stroke caused by atrial fibrillation; the aforementioned non-cardiogenic stroke includes but is not limited to lacunar stroke.

[0019] Beneficial effects: Compared with the prior art, the present invention has a good FXIa inhibitory effect, and in the FeCl2-induced rabbit carotid artery thrombosis model, the weight of the rabbit carotid artery thrombosis in the Example Compound 1 group is significantly reduced compared with the weight of the rabbit carotid artery thrombosis in the Comparative Example 1 group, which is statistically significant and has significant therapeutic effect. DETAILED DESCRIPTION

[0020] The present invention will be further described in detail below in conjunction with embodiments and experimental examples. The embodiments and experimental examples of the present invention are only used to illustrate the technical scheme of the present invention, and are not intended to limit the present invention. Any equivalent substitutions in the art made in accordance with the contents disclosed in the present invention shall fall within the protection scope of the present invention.

[0021] The structure of the compound is determined by NMR ( 1 H NMR) or liquid chromatography-mass spectrometry (LC-MS).

[0022] The liquid chromatography-mass spectrometer (LC-MS) was Agilent G6120B (used with Agilent 1260 liquid chromatography); the nuclear magnetic resonance instrument ( 1 HNMR) was Bruker AVANCE-400 or Bruker AVANCE-800, and nuclear magnetic resonance ( 1 H NMR) shifts (δ) are given in parts per million (ppm) with tetramethylsilane (TMS) as the internal standard. Chemical shifts are in 10 -6 (ppm) is given as the unit.

[0023] The term "room temperature" in the present invention means a temperature between 10 and 30°C.

[0024] Example 1: Preparation of (S)-4-(2-(4-(5-chloro-2-(1H-tetrazol-1-yl)phenyl)-3-(methoxy-d3)-6-oxopyridazine-1(6H)-yl)butyramido)-2-fluorobenzamide (Compound 1):

[0025]

[0026] Step 1: Preparation of intermediate b

[0027] Take 6-methoxypyridazine-3-ol (2g, 15.8mmol), add 40ml DMF to dissolve, add cesium carbonate (10.3g, 31.6mmol), cool to 0℃, add deuterated iodomethane (3.5g, 24.1mmol) dropwise, add for about 30 minutes, after addition, stir at room temperature for 4 hours. Add EA and water, extract, wash with water, wash with saturated salt water, dry with anhydrous sodium sulfate, filter, evaporate the solvent, purify by column chromatography, and obtain 2g of intermediate b. Yield: 88.42%, HPLC purity: 98.81%.

[0028] ESI-MS: m / z = 144.1 (M+H) + .

[0029] Step 2: Preparation of intermediate c

[0030] Take diisopropylamine (1.7g, 16.8mmol) and dissolve it in 20mlTHF, cool it to below -60℃, add 6.4ml, 2.5M n-butyl lithium n-hexane solution dropwise, add it in about 1 hour, stir and react at -60℃ for 15 minutes, add 5ml THF solution of compound b (2g, 14.0mmol) dropwise, take about 1 hour, add it, stir and react at -60℃ for 2 hours, add triisopropyl borate (2.9g, 15.4mmol) dropwise, take 30 minutes, add it, slowly warm up to room temperature (20℃), stir and react for 30 minutes. Add a mixture of 3g acetic acid and 15g water dropwise to terminate the reaction. After addition, stir at room temperature for 30 minutes. Distill off the organic solvent, add a little water, stir at room temperature for 15 minutes, filter, wash the filter cake with water, and vacuum dry at 70℃ to obtain 2.1g solid. Yield 80.22%, HPLC purity: 98.12%.

[0031] ESI-MS: m / z = 188.1 (M+H) + .

[0032] Step 3: Preparation of intermediate e

[0033] Compound d (2.59 g, 10.0 mmol) and Pd(amphos)Cl2 (107.5 mg, 0.15 mmol) were suspended in 25 ml of tert-amyl alcohol, heated to 85°C, and a mixed solution of compound c (2.22 g, 1.19 mmol), sodium carbonate (3.2 g, 30.2 mmol) and 25 ml of water was added dropwise for about 1 hour. After the addition was completed, the mixture was reacted at 85°C for 1 hour. The mixture was cooled to room temperature, extracted with EA / water, the aqueous layer was separated, and the organic layer was washed with water and saturated brine in turn, dried with anhydrous sodium sulfate, filtered, and the solvent was evaporated. Purification by column chromatography gave 2.4 g of intermediate e. The yield was 74.59%, and the HPLC purity was 98.63%.

[0034] ESI-MS: m / z = 322.1 (M+H) + .

[0035] Step 4: Preparation of intermediate f

[0036] Compound e (1.87 g, 5.80 mmol), anhydrous lithium chloride (1.3 g, 30.7 mmol), p-toluenesulfonic acid monohydrate (2.2 g, 11.6 mmol) and 20 ml of isopropanol were mixed and heated under reflux for 16 hours. After cooling to room temperature, half of the solvent was evaporated, 30 ml of water was added, stirred at room temperature for 15 minutes, filtered, the filter cake was washed with water, and vacuum dried at 70°C to obtain 1.61 g of solid. Yield: 90.21%, HPLC purity: 96.43%.

[0037] ESI-MS: m / z = 308.1 (M+H) + .

[0038] Step 5: Preparation of intermediate h

[0039] Add compound f (523 mg, 1.70 mmol), tetramethylguanidine (681 mg, 5.91 mmol), 6 ml of isopropanol, and 1.5 ml of acetone to a 25 ml reaction bottle, stir at room temperature for 15 minutes, add compound g (684 mg, 1.9 mmol), stir at room temperature overnight. The next day, add water to terminate the reaction, add EA to extract, separate the water layer, wash the organic layer with saturated ammonium chloride, water, and saturated salt water in turn, dry with anhydrous sodium sulfate, filter, evaporate the solvent, separate and purify with a chromatography column, collect the product, and obtain 832 mg of pure product. The yield is 83.37% and the purity is 98.32%.

[0040] ESI-MS: m / z = 587.2 (M+H) + .

[0041] Step 6: Preparation of intermediate j

[0042] Add compound h (587 mg, 1 mmol) and 8 ml of methanol to a 25 ml reaction bottle, stir to dissolve, and cool to 0°C. Weigh lithium hydroxide monohydrate (84 mg, 2 mmol) and dissolve in 4 ml of water, dropwise add to the reaction bottle, and react at room temperature for 2 h. Add water to terminate the reaction, adjust the pH to weak acidity with 5% citric acid, add EA to extract, separate the aqueous layer, wash the organic layer with water and saturated NaCl in turn, dry with anhydrous sodium sulfate, filter, evaporate the solvent, and perform column chromatography to collect the target product to obtain 432 mg of intermediate j, with a yield of 81.36% and a purity of 98.30%.

[0043] ESI-MS: m / z = 531.1 (M+H) + .

[0044] Step 7: Preparation of (S)-4-(2-(4-(5-chloro-2-(1H-tetrazol-1-yl)phenyl)-3-(methoxy-d3)-6-oxopyridazin-1(6H)-yl)butyramido)-2-fluorobenzamide (Compound 1)

[0045] Add intermediate j (266 mg, 0.50 mmol), ammonium chloride (71.6 mg, 1.34 mmol), HBTU (0.31 g, 0.8 mmol) and 30 ml acetonitrile to a 50 ml reaction bottle and cool to 5-10°C. Add DIPEA (0.42 g, 3.21 mmol) and react at 5-10°C for 1 hour and at room temperature for 30 minutes. Add the reaction solution into cold water, extract with ethyl acetate, wash with 5% citric acid, wash with saturated sodium bicarbonate, wash with water, wash with saturated brine, dry with anhydrous sodium sulfate, evaporate the solvent, and recrystallize with acetone / water to obtain compound 1 with a yield of 77.43% and a purity of 98.39%.

[0046] ESI-MS: m / z = 530.2 (M+H) + .

[0047] 1 HNMR(400MHz,DMSO-d6)δ:10.79(s,1H),9.44(s,1H),7.89–7.81(m,2H),7.79(m,1H),7.74–7.60(m,2H),7.55(d ,J=11.3Hz,2H),7.43(dd,J=8.6,2.0Hz,1H),7.03(s,1H),5.52(m,1H),2.16–2.05(m,2H),0.78(t,J=7.2Hz,3H).

[0048] Comparative Example 1: Preparation of (S)-4-(2-(4-(5-chloro-2-(4-(trifluoromethyl)-1H-1,2,3-triazol-1-yl)phenyl)-5-methoxy-2-oxopyridin-1(2H)-yl)butyramido)-2-fluorobenzamide

[0049]

[0050] It was synthesized according to the method described in patent CN108026072B, with a purity of 98.5%.

[0051] ESI-MS: m / z = 593.1 (M+H) + .

[0052] 1H NMR(400MHz,DMSO-d6)δ:10.78(s,1H),9.14(s,1H),7.88–7.77(m,3H),7.72–7.61(m,2H),7.55(d,2H) ),7.37(dd,1H),7.13(s,1H),6.54(s,1H),5.52(dd,1H),3.25(s,3H),2.18–2.00(m,2H),0.78(t,3H).

[0053] Test Example 1: Inhibition of coagulation factor FXIa

[0054] 1. Test samples

[0055] Example compound 1 and comparative example 1.

[0056] 2. Experimental steps

[0057] 1) Prepare experimental buffer (50 mM HEPES, 5 mM KCl, 145 mM NaCl, 1 mg / ml PEG 8000, pH 7.4) and equilibrate to room temperature.

[0058] 2) Prepare 10X compound working solution.

[0059] 3) Prepare 0.8 nM Human FXIa working solution (2X), mix well and set aside.

[0060] 4) Add 20 μL of the FXIa working solution prepared in step 3) to all experimental wells of a 384-well plate (Coring, 3702), and centrifuge at 200 g, RT, for 10 s.

[0061] 5) Add 4 μL of the compound working solution in step 2) to the corresponding experimental wells in the 384-well plate, centrifuge at 200g, RT for 10 seconds, and then incubate the working plate at 25°C for 20 minutes.

[0062] 6) Prepare 750 μM S-2366 working solution (2.5X), mix well and set aside.

[0063] 7) Add 16 μL of the S-2366 working solution in step 6) to all experimental wells in the 384-well plate, centrifuge at 200 g, RT for 10 s, and then incubate the working plate at 37° C. for 45 min.

[0064] 8) After the incubation is completed, the absorbance value at OD405nm is read using EnVision and the data is collected.

[0065] Set 5 concentrations, namely: 200nM, 40nM, 8nM, 1.6nM, 0.32nM, and detect IC 50 value.

[0066] 3. Data Analysis

[0067] 1) Z'factor = 1-3*(SD Max +SD Min ) / (Mean Max -Mean Min );

[0068] 2)CV Max =(SD Max / Mean Max )*100%;

[0069] 3) CVMin = (SD Min / Mean Min )*100%;

[0070] 4) S / B = Singal / Background;

[0071] 5) Blank control: 0.1% DMSO; Positive control: Comparative Example 1;

[0072] 6) IC 50 Calculation formula: Y = Bottom + (Top-Bottom) / (1 + 10^((LogIC 50 -X)*HillSlope)).

[0073] X: log value of compound concentration; Y: Inhibition%.

[0074] 4. Test results

[0075] The test results are shown in the table below, which show that under the same molar concentration conditions, the in vitro inhibitory activity of compound 1 of the present invention on FXIa is equivalent to that of the compound of comparative example 1.

[0076] Table 1 Activity determination of coagulation factor FXIa inhibitor (IC50)

[0077] sample <![CDATA[IC 50 (nM)]]> sample <![CDATA[IC 50 (nM)]]> Compound 1 0.09 Comparative Example 1 7.6

[0078] Experimental Example 2: In vivo efficacy evaluation in rabbit arteriovenous shunt model

[0079] 1. Test samples

[0080] Example compound 1 and comparative example 1.

[0081] 2. Test methods

[0082] Thirty all-male New Zealand white rabbits, weighing 2.5-3.0 kg, were selected and divided into three groups, 10 rabbits per group, namely, a model group, a comparative example group 1, and a compound group 1.

[0083] The comparative example 1 group and the compound 1 group were given a single injection of 6 mg / kg of the compound represented by the comparative example 1 and the compound 1 respectively via the femoral vein.

[0084] The experimental animals were anesthetized by intramuscular injection of xylazine (5 mg / kg) and ketamine (40 mg / kg), and the anesthesia was maintained by intravenous drip of xylazine and ketamine (80 mg + 800 mg, prepared in 12 ml) through the right ear vein (5 ml / h). The common carotid artery on one side was exposed surgically, and after intravenous injection for 30 minutes, a tablet was used. The filter paper (10 mm × 10 mm) on the strip was wrapped around the carotid artery without affecting blood flow. The filter paper contained 100 μl of 13% FeCl 2 After 5 minutes, the filter paper was removed and the blood vessels were rinsed twice with 0.9% sodium chloride injection. After using the filter paper for 30 minutes, the injured carotid artery was removed, and the intravascular thrombus was removed and weighed.

[0085] 3. Test results

[0086] As shown in Table 2, in the FeCl2-induced rabbit carotid artery thrombosis model, the weight of the carotid artery thrombosis in the rabbits in the Example Compound 1 group was significantly reduced compared with the weight of the carotid artery thrombosis in the rabbits in the Comparative Example 1 group, which was statistically significant.

[0087] Table 2 FeCl 2 Weight of arterial thrombus in the induced rabbit carotid artery thrombosis model (unit: mg)

[0088] Animal No. / Group Model Group Comparative Example 1 Compound 1 Mean 18.53 <![CDATA[4.80 +++ ]]> <![CDATA[2.34 +++ D]]>

[0089] Compared with the model group: +++ P<0.001: compared with control group 1: ΔP<0.01.

[0090] The above embodiment is only one of the preferred implementation modes of the present invention and should not be used to limit the protection scope of the present invention. Any changes or modifications that are made to the main design concept and spirit of the present invention and have no substantive significance, and the technical problems they solve are still consistent with the present invention, should be included in the protection scope of the present invention.

Claims

1. A compound or a pharmaceutically acceptable salt thereof as represented by formula (I): 。 2. The compound or pharmaceutically acceptable salt according to claim 1, It is characterized in that The salt is a metal salt.

3. The compound or pharmaceutically acceptable salt according to claim 2, It is characterized in that The metal salt is selected from sodium salt, potassium salt, calcium salt, lithium salt or magnesium salt.

4. A pharmaceutical composition comprising the compound or pharmaceutically acceptable salt according to any one of claims 1 to 3, It is characterized in that The composition further contains a pharmaceutically acceptable carrier and / or excipient.

5. A method for preparing the compound or pharmaceutically acceptable salt according to any one of claims 1 to 3, It is characterized in that The method includes the following route: 。 6. Use of the compound or pharmaceutically acceptable salt according to any one of claims 1 to 3 or the composition according to claim 4 in the preparation of a medicament for treating and / or preventing diseases associated with FXIa receptor.

7. The use according to claim 6, It is characterized in that The disease associated with FXIa receptor is selected from thrombosis or thromboembolism related disorders.

8. The use according to claim 6, It is characterized in that The disease associated with FXIa receptor is selected from cerebrovascular arterial disease and / or peripheral arterial disease.

9. The use according to claim 6, It is characterized in that The disease associated with FXIa receptor is selected from transient ischemic attack, ischemic stroke and / or peripheral arterial disease.

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