CN117164566B

CN117164566B - Oxopyridazine compound and preparation method and application thereof

Info

Publication number: CN117164566B
Application number: CN202311151425.8A
Authority: CN
Inventors: 黄龙; 付海霞
Original assignee: Chengdu Shibeikang Biological Medicine Technology Co ltd
Current assignee: Chengdu Shibeikang Biological Medicine Technology Co ltd
Priority date: 2023-09-07
Filing date: 2023-09-07
Publication date: 2025-06-06
Anticipated expiration: 2043-09-07
Also published as: CN117164566A

Abstract

The invention discloses a compound shown in a formula (I), a stereoisomer or a pharmaceutically acceptable salt thereof. The invention also provides application of the compound, stereoisomer or pharmaceutically acceptable salt thereof in preparing a medicament for treating and/or preventing diseases related to FXIa receptors, in particular application in preparing a medicament for treating and/or preventing cerebrovascular arterial diseases and/or peripheral arterial diseases, and has remarkable medicinal effect.

Description

Oxopyridazine compound and preparation method and application thereof

Technical Field

The invention relates to the field of pharmaceutical chemistry, in particular to an oxo-pyridine compound or salts and isomers thereof, a preparation method thereof and application thereof in preparing medicaments for treating and/or preventing diseases related to FXIa receptors, in particular to application in preparing medicaments for treating cerebrovascular arterial diseases and/or peripheral arterial diseases and the like.

Background

Thromboembolic diseases are diseases caused by abnormal blood clots formed in blood vessels of humans and animals during survival, and clinically may be manifested as myocardial infarction, stroke, deep vein thrombosis (deepveinthrombosis, DVT), pulmonary embolism, atrial fibrillation, cerebral infarction, etc., taking millions of people worldwide each year. Factor XI (FXI) is a plasma serine protease zymogen necessary for maintaining the endogenous pathway, and activated to form activated factor XIa (FXIa) plays a key role in the amplification of the coagulation cascade. Therefore, drugs against FXIa targets block endogenous pathways and inhibit amplification of the coagulation cascade, thus having antithrombotic effects.

The reported FXIa inhibitors mainly comprise monoclonal antibodies, antisense oligonucleotides, chemical small molecules, polypeptides or protein or polypeptide mimics and the like. Currently, milvexian, which was developed in combination with qigong, has completed a clinical phase II trial, which shows less risk of bleeding. Clinical phase I trials of the intravenous injection of the small molecule FXIa inhibitor BMS-962122 have been completed and development has been suspended. The small molecule oral FXIa inhibitor ONO-7684 developed by Japan Kochia company enters clinical phase I study. BAY-2433334 developed by Bayer has completed a clinical phase II trial and is currently the most promising small molecule FXIa inhibitor. The monoclonal antibody and the antisense oligonucleotide need to be injected and administrated, and have the defects of high price, slow effect, possibly difficult control and the like, and the chemical small molecules have the advantages of relatively good oral bioavailability, better patient compliance and the like.

Therefore, the research and development of new FXIa small molecule inhibitor drugs with safety, effectiveness, good specificity and strong activity can be used for overcoming the defect that the current clinical anticoagulation anti-thrombus drugs are easy to cause bleeding complications and meeting the clinical unmet demands.

Disclosure of Invention

The compound is a novel oxo-pyridine compound, and has good anticoagulation effect and in-vitro and in-vivo affinity to FXIa.

In one aspect, the present invention provides a compound of formula (I), a stereoisomer or a pharmaceutically acceptable salt thereof:

further, the pharmaceutically acceptable salt is a metal salt.

Further, the metal salt is selected from sodium salt, potassium salt, calcium salt, lithium salt, and magnesium salt.

In another aspect, the present invention provides a pharmaceutical composition of the above compound, a stereoisomer or a pharmaceutically acceptable salt thereof, which composition further comprises a pharmaceutically acceptable carrier and/or adjuvant.

In another aspect, the present invention provides a process for the preparation of the above compound, a stereoisomer or a pharmaceutically acceptable salt thereof, comprising the following route:

In another aspect, the invention also provides the use of any of the compounds, stereoisomers or pharmaceutically acceptable salts thereof, or a composition thereof, for the manufacture of a medicament for the treatment and/or prophylaxis of diseases associated with the FXIa receptor.

Further, the above-mentioned diseases related to FXIa receptor are selected from thrombosis or thromboembolic related disorders.

Further, the above-mentioned FXIa receptor-related diseases are selected from cerebrovascular arterial diseases and/or peripheral arterial diseases.

Further, the above-mentioned cerebrovascular arterial diseases include, but are not limited to, transient Ischemic Attacks (TIAs), ischemic strokes or events resulting in thrombosis and/or thromboembolic origin of strokes or TIAs, and the above-mentioned peripheral arterial diseases include, but are not limited to, peripheral arterial occlusion, acute limb ischemia, amputation, reocclusion and restenosis after interventions such as angioplasty, stent implantation or surgery and bypass, and/or stent thrombosis.

Further, the ischemic stroke includes, but is not limited to, cardiac stroke, non-cardiac stroke, stroke due to aortic or arteriolar diseases, stroke due to adventitious causes, cryptogenic stroke, embolic stroke, or embolic stroke of adventitious origin.

Further, the cardiac stroke includes, but is not limited to, stroke due to atrial fibrillation, and the non-cardiac stroke includes, but is not limited to, interstitial stroke.

Compared with the prior art, the invention has the advantages that the FXIa inhibition effect is very good, in a FeCl2 induced rabbit carotid artery thrombosis model, the weight of the rabbit carotid artery thrombosis of the example compound 1 group is obviously reduced compared with that of the rabbit carotid artery thrombosis of the comparative example 1 group, and the invention has statistical significance and obvious curative effect.

Detailed Description

The present invention will be described in further detail with reference to the following examples and experimental examples, which are only for illustrating the technical scheme of the present invention, but not for limiting the present invention, and any equivalent substitution in the art according to the disclosure of the present invention shall fall within the scope of the present invention.

The structure of the compound was determined by nuclear magnetic resonance (¹ H NMR) or liquid chromatography-mass spectrometry (LC-MS).

The LC-MS is Agilent G6120B (matched with liquid Agilent 1260), the nuclear magnetic resonance (¹ HNMR) is Bruker AVANCE-400 or Bruker AVANCE-800, the nuclear magnetic resonance (¹ H NMR) displacement (delta) is given in parts per million (ppm), the internal standard is Tetramethylsilane (TMS), and the chemical displacement is given in units of 10 ^-6 (ppm).

The term "room temperature" in the present invention means that the temperature is between 10 and 30 ℃.

Example 1 preparation of (S) -4- (2- (4- (5-chloro-2- (1H-tetrazol-1-yl) phenyl) -3- (methoxy-d 3) -6-oxopyridazin-1 (6H) -yl) butyramide) -2-fluorobenzamide (Compound 1):

Step 1 preparation of intermediate b

6-Methoxypyridazin-3-ol (2 g,15.8 mmol) was taken, dissolved by adding 40ml DMF, cesium carbonate (10.3 g,31.6 mmol) was added, cooled to 0℃and deuterated iodomethane (3.5 g,24.1 mmol) was added dropwise over about 30 minutes, and the reaction was stirred at room temperature for 4 hours after the addition. EA, water, extraction, water washing, saturated brine washing, anhydrous sodium sulfate drying, filtration, evaporation of the solvent, column chromatography purification, 2g of intermediate b were obtained. The yield was 88.42% and the HPLC purity was 98.81%.

ESI-MS:m/z=144.1(M+H)⁺。

Step 2 preparation of intermediate c

Diisopropylamine (1.7 g,16.8 mmol) was dissolved in 20ml THF, cooled to below-60 ℃, 6.4ml of 2.5M n-butyllithium n-hexane solution was added dropwise, the addition was completed for about 1 hour, 5ml THF solution of compound b (2 g,14.0 mmol) was added dropwise over-60 ℃ for 15 minutes, the addition was completed for-60 ℃ for 2 hours, triisopropyl borate (2.9 g,15.4 mmol) was added dropwise over-30 minutes, the addition was completed, and the temperature was slowly raised to room temperature (20 ℃) for 30 minutes with stirring. A mixture of 3g of acetic acid and 15g of water was added dropwise to terminate the reaction. After the addition, stirring at room temperature for 30 minutes. The organic solvent was distilled off, a little water was added, stirred at room temperature for 15 minutes, filtered, and the cake was washed with water and dried under vacuum at 70℃to give 2.1g of solid. The yield was 80.22% and the HPLC purity was 98.12%.

ESI-MS:m/z=188.1(M+H)⁺。

Step 3 preparation of intermediate e

A mixed solution of compound d (2.59 g,10.0 mmol), pd (amphos) Cl2 (107.5 mg,0.15 mmol) in 25ml of t-amyl alcohol was taken, heated to 85℃and a reaction was carried out for 1 hour after the addition of a mixed solution of compound C (2.22 g,1.19 mmol), sodium carbonate (3.2 g,30.2 mmol) and 25ml of water for about 1 hour. Cooling to room temperature, adding EA/water, extracting, separating out water layer, washing organic layer with water and saturated salt water, drying with anhydrous sodium sulfate, filtering, and evaporating solvent. Purification by column chromatography gave 2.4g of intermediate e. Yield 74.59% and HPLC purity 98.63%.

ESI-MS:m/z=322.1(M+H)⁺。

Step 4 preparation of intermediate f

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 20ml of isopropanol were taken and mixed, and reacted under reflux with heating for 16 hours. Cooled to room temperature, half of the solvent was distilled off, 30ml of water was added, stirred at room temperature for 15 minutes, filtered, and the cake was washed with water and dried under vacuum at 70 ℃ to give 1.61g of solid. The yield was 90.21% and the HPLC purity was 96.43%.

ESI-MS:m/z=308.1(M+H)⁺。

Step 5 preparation of intermediate h

A25 ml reaction flask was charged with compound f (803 mg,1.70 mmol), tetramethylguanidine (681 mg,5.91 mmol), isopropanol 6ml, acetone 1.5ml, and stirred at room temperature for 15 minutes, compound g (684 mg,1.9 mmol) was added, and the reaction was stirred at room temperature overnight. The next day, water is added to terminate the reaction, EA is added to extract, the water layer is separated, the organic layer is washed with saturated ammonium chloride, water, saturated saline water and anhydrous sodium sulfate in sequence, the mixture is dried, filtered, the solvent is evaporated, and the chromatographic column is used for separating and purifying, and the product is collected to obtain 832mg of pure product. The yield thereof was found to be 83.37%, the purity was 98.32%.

ESI-MS:m/z=587.2(M+H)⁺。

Step 6 preparation of intermediate j

A25 ml reaction flask was charged with compound h (587 mg,1 mmol), methanol 8ml, dissolved with stirring and cooled to 0 ℃. Lithium hydroxide monohydrate (84 mg,2 mmol) was weighed and dissolved in 4ml of water, and the mixture was added dropwise to a reaction flask and reacted at room temperature for 2 hours. Adding water to terminate the reaction, adjusting pH to weak acidity with 5% citric acid, adding EA to extract, separating water layer, washing organic layer with water, saturated NaCl, drying with anhydrous sodium sulfate, filtering, evaporating solvent, performing column chromatography, and collecting target product to obtain 432mg of intermediate j with yield 81.36% and purity of 98.30%.

ESI-MS:m/z=531.1(M+H)⁺。

Step 7 preparation of (S) -4- (2- (4- (5-chloro-2- (1H-tetrazol-1-yl) phenyl) -3- (methoxy-d 3) -6-oxopyridazin-1 (6H) -yl) butyramide) -2-fluorobenzamide (Compound 1)

50Ml reaction flask was charged with intermediate j (266 mg,0.50 mmol), ammonium chloride (71.6 mg,1.34 mmol), HBTU (0.31 g,0.8 mmol) and 30ml acetonitrile and cooled to 5-10 ℃. DIPEA (0.42 g,3.21 mmol) was added and reacted at 5-10℃for 1 hour and at room temperature for 30 minutes. Adding the reaction solution into cold water, extracting with ethyl acetate, 5% citric acid washing, saturated sodium bicarbonate washing, water washing, saturated saline water washing, anhydrous sodium sulfate drying, evaporating the solvent, and recrystallizing with acetone/water to obtain compound 1 with yield of 77.43% and purity of 98.39%.

ESI-MS:m/z=530.2(M+H)⁺。

¹HNMR(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).

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) butanamide) -2-fluorobenzamide

Synthesized according to the method described in patent CN108026072B, purity 98.5%.

ESI-MS:m/z=593.1(M+H)⁺。

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).

Test example 1 inhibition of coagulation factor FXIa

1. Test sample

Example compound 1 and comparative example 1.

2. Test procedure

1) The assay buffer (50 mM HEPES,5mM KCl,145mM NaCl,1mg/mlPEG8000, pH 7.4) was prepared and equilibrated to room temperature.

2) Preparing 10X compound working solution.

3) Preparing 0.8nM Human FXIa working solution (2X), and mixing.

4) Add 20. Mu.L of FXIa working fluid from step 3) to all experimental wells of 384 well plates (Coring, 3702), 200g, RT, centrifuge for 10s.

5) Add 4. Mu.L of the compound working fluid from step 2) to the corresponding experimental well in 384 well plates, 200g, RT, centrifuge for 10s, and then incubate the working plate at 25℃for 20min.

6) Preparing 750 mu M S-2366 working solution (2.5X), and uniformly mixing for later use.

7) Mu.L of the S-2366 working solution from step 6) was added to all experimental wells in 384 well plates, 200g, RT, centrifuged for 10S and the plates were incubated at 37℃for 45min.

8) After incubation was completed, absorbance at OD405nm was read using EnVision and data was collected.

Setting 5 concentrations, namely 200nM, 40nM, 8nM, 1.6nM and 0.32nM, and detecting IC ₅₀ values.

3. Data analysis

1)Z’factor=1-3*(SD_Max+SD_Min)/(Mean_Max-Mean_Min);

2)CV_Max＝(SD_Max/Mean_Max)*100%;

3)CVMin=(SD_Min/Mean_Min)*100%;

4)S/B=Singal/Background;

5) Blank control, 0.1% dmso, positive control, comparative example 1;

6) The calculation formula of IC ₅₀ is y=bottom+ (Top-Bottom)/(1+10 ((LogIC ₅₀ -X) HillSlope)).

X is the log value of the compound concentration, and Y is the Inhibition%.

4. Test results

The results of the test are shown in the following table, and the results show that the in vitro inhibition activity of the compound 1 of the present invention on FXIa is equivalent to that of the compound of comparative example 1 under the same molar concentration condition.

TABLE 1 determination of the Activity of inhibitors of the coagulation factor FXIa (IC 50)

Sample of	IC₅₀(nM)	Sample of	IC₅₀(nM)
				Compound 1	0.09	Comparative example 1	7.6

Test example 2 evaluation of in vivo efficacy of Rabbit arteriovenous shunt model

1. Test sample

Example compound 1 and comparative example 1.

2. Test method

Selecting New Zealand white rabbits, all male 30, 2.5-3.0kg. Divided into 3 groups, 10/group. The model group, the comparative example 1 group and the compound 1 group, respectively.

Comparative example 1 group and compound 1 group 6mg/kg of the compounds shown in comparative example 1 and compound 1, respectively, were administered via single injection into the femoral vein.

The test animals were anesthetized by intramuscular injection of xylazine (xylazine, 5 mg/kg) and ketamine (ketamine, 40 mg/kg), and the anesthetic effect was maintained by intravenous infusion of xylazine and ketamine (80 mg+800mg, 12ml formulated) through the right auricular vein (5 ml/h) of rabbits. One common carotid artery was exposed surgically and after 30min of intravenous administration, one patch was usedThe filter paper (10 mm. Times.10 mm) on the strip was wrapped around the carotid artery and the blood flow was not affected after wrapping, and the filter paper contained 100. Mu.l of a 13% strength FeCl ₂ aqueous solution. After 5min, the filter paper was removed and the vessel was rinsed 2 times with 0.9% sodium chloride injection. After 30min using filter paper, the injured carotid artery was excised, and the intravascular thrombus was removed and weighed.

3. Test results

As shown in table 2, in the FeCl 2-induced rabbit carotid thrombus model, the weight of rabbit carotid artery thrombus in the example compound 1 group was significantly reduced compared to that in the comparative example 1 group, and it was statistically significant.

Table 2 FeCl ₂ weight of arterial thrombus in rabbit carotid thrombosis model induced (unit: mg)

Animal numbering/group	Model group	Comparative example 1 group	Group 1 of compounds
				Mean	18.53	4.80⁺⁺⁺	2.34⁺⁺⁺Δ

⁺⁺⁺ P <0.001 compared to model group and DeltaP <0.01 compared to comparative example 1 group.

The above embodiment is only one of the preferred embodiments of the present invention, and should not be used to limit the scope of the present invention, but all the insubstantial modifications or color changes made in the main design concept and spirit of the present invention are still consistent with the present invention, and all the technical problems to be solved are included in the scope of the present invention.

Claims

1. A compound or pharmaceutically acceptable salt of formula (I):

。

2. The compound or pharmaceutically acceptable salt of claim 1, wherein the salt is a metal salt.

3. A compound or pharmaceutically acceptable salt according to claim 2, wherein the metal salt is selected from sodium, potassium, calcium, lithium or magnesium salts.

4. A pharmaceutical composition comprising a compound or pharmaceutically acceptable salt according to any one of claims 1 to 3, wherein the composition further comprises a pharmaceutically acceptable carrier and/or adjuvant.

5. A process for the preparation of a compound or pharmaceutically acceptable salt according to any one of claims 1 to 3, which comprises the following route:

。

6. Use of a compound or a pharmaceutically acceptable salt according to any one of claims 1 to 3 or a composition according to claim 4 for the manufacture of a medicament for the treatment and/or prophylaxis of diseases which are associated with the FXIa receptor.

7. The use according to claim 6, wherein the disease associated with FXIa receptor is selected from thrombosis or thromboembolic associated disorders.

8. The use according to claim 6, wherein the disease associated with FXIa receptor is selected from cerebrovascular arterial disease and/or peripheral arterial disease.

9. The use according to claim 6, wherein the disease associated with FXIa receptor is selected from transient ischemic attacks, ischemic strokes and/or peripheral arterial diseases.