A class of compounds and their preparation methods and applications in treating thromboembolic diseases

Abstract

The present invention discloses a compound represented by any one of the general formulas (1) and (2), a method for preparing the compound, and its use in preparing a drug for inhibiting coagulation factor Xa, a drug having antithrombotic activity, or a drug for treating and / or preventing thromboembolic diseases. #imgabs0#

Description

Technical Field

[0001] The present invention relates to a class of compounds and a preparation method thereof and application thereof in preparing medicines for treating thromboembolic diseases, belonging to the fields of medicinal chemistry, pharmacology and preparations. Background Art

[0002] Thromboembolic disease is caused by blood clots that develop during blood flow. Thrombosis can lead to partial or complete interruption of blood flow, resulting in insufficient blood supply to the corresponding organs, functional imbalances, and ultimately thrombotic disease. Currently, the main anticoagulant drugs used clinically are coumarins and heparins. While they can achieve certain therapeutic effects, they can also cause adverse reactions such as bleeding, allergies, gastrointestinal discomfort, liver and kidney dysfunction, and respiratory distress. Coagulation factor Xa (FXa) is a serine protease that resides at the intersection of the intrinsic and extrinsic coagulation pathways, occupying a crucial position in the coagulation pathway. Factor Xa inhibitors not only block intrinsic coagulation but also inhibit extrinsic coagulation. Factor Xa simply promotes coagulation and does not have the side effects of thrombin inhibitors. Therefore, Factor Xa inhibitors have become a hot topic in anticoagulant drug research in recent years.

[0003] Siegesbeckia serrata, a member of the Asteraceae family, has the effects of dispelling rheumatism, strengthening tendons and bones, clearing away heat and detoxifying, and lowering blood pressure. Darutigenol, a compound extracted from the plant, has been shown in modern pharmacological experiments to possess anti-inflammatory, anti-allergic, antithrombotic, and antihistamine release activities. Diterpenes (primarily enantio-pimarane and enantio-kaurane diterpenes) underlie the anti-inflammatory and antithrombotic properties of Siegesbeckia serrata. Preliminary pharmacological experiments have shown that darutigenol significantly inhibits the maximum platelet aggregation and adhesion rates in rats, prolongs euglobulin lysis time, and bleeding and clotting times in mice. It also significantly reduces whole blood and plasma viscosity in rat models of blood stasis, improves abnormal hemorheology, and inhibits TXB2 synthesis in rat models of blood stasis, demonstrating significant antithrombotic, thrombolytic, anticoagulant, and hemorheological effects. Summary of the Invention

[0004] Purpose of the invention: The technical problem to be solved by the present invention is to provide a class of sirugradinol derivatives having antithrombotic activity, effectively inhibiting coagulation factor Xa, and being used for the treatment and prevention of thromboembolic diseases, as well as their preparation method and application.

[0005] Technical solution: To solve the above technical problems, the present invention provides a compound represented by any of the following general formulas:

[0006]

[0007] in, -OH, -Br or

[0008]

[0009] Wherein, the structural formula of the compound is any one of the following:

[0010]

[0011] The present invention also provides a method for preparing the compound, comprising the following steps:

[0012] (a) reacting dasyphylline with acetone under the catalysis of phosphomolybdic acid to obtain compound 1;

[0013] (b) oxidizing the acetone complex of siegetinol with pyridinium chlorochromate to obtain compound 2;

[0014] (c) Compound 2 reacts with ethyl formate, trifluoromethylethyl formate or pyridinium tribromide to introduce a carbonyl group at the α position. or -Br, to give compounds 3, 7, 10, and 11;

[0015] (d) Compound 3 reacts with hydroxylamine hydrochloride, hydrazine hydrate, and phenylhydrazine hydrochloride to obtain compounds 4, 5, and 6;

[0016] (e) Compound 7 reacts with hydroxylamine hydrochloride and hydrazine hydrate to obtain compounds 8 and 9;

[0017] (f) Compound 11 reacts with thiourea to obtain compound 12.

[0018] The present invention also provides the use of the compound in preparing a drug for inhibiting coagulation factor Xa.

[0019] The present invention also provides the use of the compound in preparing a medicine with antithrombotic activity.

[0020] The present invention also provides the use of the compound in preparing medicines for treating and / or preventing thromboembolic diseases.

[0021] Preferably, the structural formula of the compound is any one of the following:

[0022]

[0023] Preferably, the structural formula of the compound is any one of the following:

[0024]

[0025]

[0026] Beneficial effects: Compared with the prior art, the present invention has the following significant advantages: the present invention prepares a variety of new compounds with antithrombotic activity, which can effectively inhibit coagulation factor Xa and can be used for the treatment and prevention of thromboembolic diseases. BRIEF DESCRIPTION OF THE DRAWINGS

[0027] Figure 1 The docking diagrams of siuguilinol and coagulation factor Xa (FXa) under different modes;

[0028] Figure 2 Docking diagrams of compound 6 and coagulation factor Xa (FXa) under different modes;

[0029] Figure 3 Docking diagrams of compound 9 and coagulation factor Xa (FXa) under different modes;

[0030] Figure 4 Docking diagram of compound 12 and coagulation factor Xa (FXa) under different modes. DETAILED DESCRIPTION

[0031] The technical solution of the present invention will be further described below with reference to the accompanying drawings.

[0032] Example 1

[0033] Compound 1: Preparation of Siegesbeckia alcohol acetone complex

[0034] The structural formula of sigerinol acetone complex is as follows:

[0035]

[0036] Synthesis route:

[0037] Weigh siuguilinol (prepared in the laboratory, HPLC purity> 98%). 1 2 g (6.21 mmol) of the product (no obvious impurity peaks on the H NMR chromatogram) was placed in a 250 mL dry eggplant-shaped flask, 20 mL of dry acetone and 626 mg (0.34 mmol) of phosphomolybdic acid were added, and the reaction was stirred at room temperature for 4 h. The acetone was evaporated under reduced pressure, and the residue was dissolved in 30 mL of ethyl acetate. The product was washed three times with saturated NaHCO3 solution and water, each 30 mL each, and then dried over anhydrous Na2SO4. The Na2SO4 was removed by suction filtration, and the solvent was evaporated under reduced pressure to obtain 2.15 g of pure white product (95.6% yield).

[0038] The above pure white product was subjected to thin layer chromatography (TLC) and nuclear magnetic resonance detection, and the results were as follows: mp: 103.4-106.0°C. (c 0.1, MeOH). 1 H NMR (CDCl3, 400MHz): δΗ 5.06(1H,s,H-14),4.01(1H,t,J=6.9Hz,H-15),4.17(1H,dd,J=7.8,6.9Hz,H a -16),4.01(1H,t,J=7.8Hz,H b -16),3.26(1H,dd,J=11.6,4.2Hz,H-3),2.27(1H,m),2.02(1H,td,J=14.2,5.2Hz),1.77-1.50(9H,m),1.41(3H,s,Me-22) ,1.35(3H,s,Me-23),1.20-1.04(3H,m),1.00(3H,s,Me-18),0.91(3H,s,Me-17),0.82(3H,s,Me-19),0.77(3H,s,Me-20); 13 C NMR (CDCl3, 100 MHz): δ C 139.6(C-8),126.7(C-14),108.5(C-21),79.7(C-15),79.0(C-3),65.3(C-16),54.1,50.2,38.9,38.1,37.1,36 .0,35.8,32.0,28.5(C-18),27.5,26.3(C-22),25.2(C-23),22.4,22.2(C-17),18.2,15.7(C-19),14.9(C-20). ESI-MS m / z C 23 H 38 O3Na[M+Na] + :385.27. Compound 1 was identified as sigerinol acetonide.

[0039] Example 2

[0040] Compound 2: Preparation of ent-pimarane-8(14)-ene-3-one-15,16-propanone complex:

[0041] The structural formula of ent-pimarane-8(14)-ene-3-one-15,16-propanone complex is as follows:

[0042]

[0043] Synthesis route:

[0044] Weigh 2 g (5.56 mmol) of compound 1 (sichuanlin alcohol acetonide) prepared in Example 1 into a 250 mL dry eggplant-shaped flask. Add 30 mL of dichloromethane and stir to dissolve. Add 10 g of 60-100 mesh silica gel to the reaction flask, and then transfer the reaction apparatus to an ice-water bath. Dissolve 1.83 g (8.34 mmol) of pyridinium chlorochromate (PCC) in 15 mL of dichloromethane and slowly add it dropwise to the reaction solution. Stir at room temperature for 6 hours. The reaction solution is passed through a 200-300 mesh silica gel vacuum column, eluted with dichloromethane, and the eluents are combined and concentrated to obtain the crude product. The crude product is subjected to column chromatography, eluting with petroleum ether:ethyl acetate (10:1, v:v) to obtain 1.75 g of an oily substance, with a yield of 87.4%.

[0045] The above oil was subjected to TLC and NMR detection, and the results were as follows: mp: 94.9-95.1°C. (c0.1, MeOH). 1 H NMR (CDCl3, 400MHz):δ Η 5.14(1H,s,H-14),4.03(1H,t,J=6.9Hz,H-15),3.92(1H,dd,J=7.8,6.9Hz,H a -16),3.78(1H,t,J=7.8Hz,H b -16),2.66(1H,td,J=14.8,5.6),2.12-2.00(2H,m),1.78-1.74(2H,m),1.64-1.45(6H,m),1.42(3H,s,Me-22),1.36 (3H,s,Me-21),1.19-1.14(1H,m),1.09(3H,s,Me-18),0.98(3H,s,Me-17),1.06(3H,s,Me-19),0.93(3H,s,Me-20); 13 C NMR (Pyridine-d5, 100 MHz): δ C 216.7(C-3),138.7(C-8),127.6(C-14),108.6(C-21),80.0(C-15),65.4(C-16),55.3,49.4,47.8,38.1,37.6,3 6.0,35.8,34.8,31.8,26.2(C-22),25.8(C-18),25.2(C-23),23.2,22.5(C-17),22.3(C-19),18.4,14.9(C-20). ESI-MS m / z C 23 H 38 O3Na[M+Na] +:383.31. The compound 2 was determined to be ent-pimarane-8(14)-ene-3-one-15,16-propanone complex.

[0046] Example 3

[0047] Compound 3: Preparation of enantio-1'-hydroxy-pimarane-2(1'),8(14)-diene-3-one-15,16-acetone complex

[0048] The structure of enantio-1'-hydroxy-pimarane-2(1'),8(14)-diene-3-one-15,16-propanone complex is as follows:

[0049]

[0050] Synthesis route:

[0051] 500 mg (1.39 mmol) of compound 2 (enantio-pimarane-8(14)-ene-3-one-15,16-acetonide) prepared in Example 2 and 600 mg (11.11 mmol) of sodium methoxide were weighed and placed in a 50 mL eggplant-shaped flask. 20 mL of dry tetrahydrofuran (THF) was added and stirred for 0.5 h. 0.3 g (4.16 mmol) of dry ethyl formate was slowly added dropwise and the reaction was allowed to proceed at room temperature overnight. The solvent in the reaction solution was evaporated under reduced pressure, and 30 mL of ethyl acetate was added to the residue to dissolve it. The mixture was washed with saturated brine and water three times, 30 mL each time, and then dried over anhydrous Na2SO4. Na2SO4 was removed by suction filtration, and the solvent was evaporated under reduced pressure to obtain a crude product. The product was chromatographed on a silica gel column and eluted with petroleum ether:ethyl acetate (10:1, V:V) to obtain 439 mg of pure product with a yield of 81.4%.

[0052] The above pure product was subjected to thin layer chromatography (TLC) and nuclear magnetic resonance detection, and the results were as follows: mp: 137.4-138.3°C. (c 0.1, MeOH). 1 H NMR (CDCl3, 400MHz): δ Η 14.91(1H,s,-OH),8.61(1H,s,=CH-OH),5.19(1H,s,H-15),3.99(1H,t,J=6.8Hz,H-15),3.91(1H,dd,J=7.7,6.8Hz,H a -16),4.01(1H,t,J=7.7Hz,H b-16),2.39-2.29(2H,m),2.09-2.01(2H,m),1.78-1.62(6H,m),1.42(3H,s,Me-22),1.35(3H,s,Me-23),1 .41-1.34(2H,overlap),1.21(3H,s,Me-18),1.12(3H,s,Me-19),0.95(3H,s,Me-17),0.76(3H,s,Me-20); 13 C NMR (CDCl3, 100 MHz): δ C 190.9(C-3),187.4(=CH-OH),138.3(C-8),126.8(C-14),108.5(C-21),105.7(C-2),80.5(C-15),65.4(C-16),51.5,47.6,4 0.3,38.2,37.1,36.0,35.9,31.4,29.0(C-18),26.2(C-22),25.1(C-23),24.0,22.4(C-19),21.5(C-17),18.2,13.7(C-20). ESI-MS m / z C 24 H 35 O4[MH] - :387.37. Compound 3 was identified as enantio-1'-hydroxy-pimarane-2(1'),8(14)-diene-3-one-15,16-propanone complex.

[0053] Example 4

[0054] Compound 4: Preparation of ent-pimarane-8(14)-ene-15,16-acetonide and [3,2,c]isoxazole

[0055] The structural formula of ent-pimarane-8(14)-ene-15,16-propanone ester and [3,2,c]isoxazole is as follows:

[0056]

[0057] Synthesis route:

[0058] 100 mg (0.26 mmol) of compound 3 (enantio-1'-hydroxy-pimarane-2(1'),8(14)-diene-3-one-15,16-acetonide) prepared in Example 3 and 56 mg (0.78 mmol) of hydroxylamine hydrochloride were weighed and placed in a 25 mL eggplant-shaped flask. 5 mL of pyridine was added and the mixture was refluxed for 4 h. The pyridine was evaporated under reduced pressure, and the residue was dissolved in 20 mL of ethyl acetate. The mixture was washed three times with saturated brine, each time with 30 mL of water, and then dried over anhydrous Na2SO4. The Na2SO4 was removed by suction filtration, and the solvent was evaporated under reduced pressure to obtain a crude product. 54 mg of the pure product was obtained by silica gel column chromatography, eluting with petroleum ether:ethyl acetate (10:1, v:v), with a yield of 54.1%.

[0059] The above pure product was subjected to thin layer chromatography (TLC) and nuclear magnetic resonance detection, and the results were as follows: mp: 129.9-131.1°C. (c 0.1, MeOH). 1 H NMR (CDCl3, 400MHz) showed characteristic signals: δ Η 8.07(1H,s,=CH-O),5.21(1H,s,H-15),3.99(1H,t,J=6.9Hz,H-15),3.91(1H,dd,J=7.7,6.9Hz,H a -16),4.01(1H,t,J=7.7Hz,H b -16),2.78(1H,d,J=15.0Hz),2.36-2.32(1H,m),2.14-2.04(2H,m),1.87(1H,t,J=6.4Hz),1.78-1.66(4H,m),1.57-1.44(2H,m),1.42( 3H,s,Me-22),1.39(3H,s,Me-18),1.35(3H,s,Me-23),1.31(3H,s,Me-19),1.25-1.20(1H,m),0.96(3H,s,Me-17),0.71(3H,s,Me-20); 13 C NMR (CDCl3, 100MHz):δ C167.8(C-3),153.2(=CH-O-),138.3(C-8),126.9(C-14),113.0(C-2),108.7(C-21),80.5(C-15),65.4(C-16),52.7,47.8 ,39.9,36.0(2×C),33.8,32.1(C-18),31.5,26.2(C-22),25.4(C-19),25.1(C-23),23.7,22.5(C-17),18.2,14.1(C-20). ESI-MS m / z C 24 H 36 NO3[M+H] + :386.42. Compound 4 was identified as ent-pimarane-8(14)-ene-15,16-propanone ester and [3,2,c]isoxazole.

[0060] Example 5

[0061] Compound 5: Preparation of ent-pimarane-8(14)-ene-15,16-acetonide and [3,2,c]pyrazole

[0062] The structural formula of ent-pimarane-8(14)-ene-15,16-propanone ester and [3,2,c]pyrazole is as follows:

[0063]

[0064] Synthesis route:

[0065] 100 mg (0.26 mmol) of compound 3 (enantio-1'-hydroxy-pimarane-2(1'),8(14)-diene-3-one-15,16-acetonide) prepared in Example 3 and 27 mg (0.52 mmol) of hydrazine hydrate were weighed and placed in a 25 mL eggplant-shaped flask. 5 mL of ethanol was added to dissolve the mixture. The mixture was refluxed for 3 h. The reaction solution was evaporated under reduced pressure, and the residue was dissolved in 20 mL of ethyl acetate. The mixture was washed three times with saturated brine, 30 mL each time, and then dried over anhydrous Na2SO4. The Na2SO4 was removed by suction filtration, and the solvent was evaporated under reduced pressure to obtain a crude product. 74 mg of the pure product was obtained by silica gel column chromatography, eluting with petroleum ether:ethyl acetate (10:1, v:v), with a yield of 74.1%.

[0066] The above pure product was subjected to thin layer chromatography (TLC) and nuclear magnetic resonance detection, and the results were as follows: mp: 144.9-146.5℃. (c 0.1, MeOH). 1 H NMR (CDCl3, 400MHz):δ Η7.30(1H,s,=CH-N),5.19(1H,s,H-14),4.01(1H,t,J=6.9Hz,H-15),3.91(1H,dd,J=7.7,6.9Hz,H a -16),4.01(1H,t,J=7.7Hz,H b -16),2.68(1H,d,J=15.0Hz),2.33(1H,brd,J=12.8Hz),2.22(1H,d,J=14.4Hz),2.08(1H,t,J=13.4Hz),1.88(1H,t,J=5.8Hz),1.80-1.66(4H,m ),1.59-1.44(2H,m),1.42(3H,s,Me-22),1.35(3H,s,Me-23),1.34(3H,s,Me-18),1.24(3H,s,Me-19),0.96(3H,s,Me-17),0.74(3H,s,Me-20); 13 C NMR (CDCl3, 100MHz):δ C 149.2(C-3),139.2(C-8),133.8(=CH-N),126.3(C-14),112.3(C-2),108.5(C-21),80.6(C-15),65.5(C-16),52.7,48.3,40 .3,36.2,36.0,34.3,33.5,31.8(C-18),31.7,26.3(C-22),25.2(C-23),24.7(C-19),23.7,22.5(C-17),18.2,14.3(C-20). ESI-MS m / z C 24 H 37 N2O2[M+H] + :385.34. Compound 5 was identified as ent-pimarane-8(14)-ene-15,16-propanone ester and [3,2,c]pyrazole.

[0067] Example 6

[0068] Compound 6: Preparation of ent-pimarane-8(14)-ene-15,16-propanone esters and [3,2,c][1'-phenylpyrazole]

[0069] The structural formula of ent-pimarane-8(14)-ene-15,16-propanone ester and [3,2,c][1'-phenylpyrazole] is as follows:

[0070]

[0071] 100 mg (0.26 mmol) of compound 3 (enantio-1'-hydroxy-pimarane-2(1'),8(14)-diene-3-one-15,16-acetonide) prepared in Example 3 and 55 mg (0.52 mmol) of phenylhydrazine hydrochloride were weighed and placed in a 25 mL eggplant-shaped flask. 5 mL of ethanol was added to dissolve the mixture. The mixture was refluxed for 3 h. The reaction solution was evaporated under reduced pressure, and the residue was dissolved in 20 mL of ethyl acetate. The mixture was washed three times with saturated brine, 20 mL each time, and then dried over anhydrous Na2SO4. The Na2SO4 was removed by suction filtration, and the solvent was evaporated under reduced pressure to obtain a crude product. 63 mg of the pure product was obtained by silica gel column chromatography, eluting with petroleum ether:ethyl acetate (15:1, v:v), with a yield of 52.7%.

[0072] The above pure product was subjected to thin layer chromatography (TLC) and nuclear magnetic resonance detection, and the results were as follows: mp: 174-176°C. (c 0.1, MeOH). 1 H NMR (CDCl3, 400MHz): δ Η 7.44-7.37(6H,m),5.19(1H,s,H-14),4.01(1H,t,J=6.9Hz,H-15),3.91(1H,dd,J=7.8,6.9Hz,H a -16),3.75(1H,t,J=7.8Hz,H b -16),2.72(1H,d,J=15.0Hz),2.32-2.27(2H,m),2.04(1H,t,J=13.4Hz),1.87(1H,t,J=5.8Hz),1.80-1.53(5H,m),1.46-1.39(1H,overlap), 1.42(3H,s,Me-22),1.35(3H,s,Me-21),1.28-1.18(1H,m),1.09(3H,s,Me-18),0.97(3H,s,Me-17),0.96(3H,s,Me-19),0.80(3H,s,Me-20); 13 C NMR (CDCl3, 100MHz):δ C146.0(C-3),138.9(C-8),137.9(=CH-N),126.2(C-14),114.1(C-2),108.6(C-21),80.7(C-15),65.5(C-16),53.9,48.3,39.9,36.1,36.0,35.0,31.6,29.9(C-18),26.2(C-22),25.1(C-23),23.7,22.7(C-19),22.5(C-17),18.1,14.4(C-20). Carbon signal on the benzene ring: δ C 142.4,129.1,129.1,128.9,128.4,128.4. ESI-MS m / z C 30 H 41 N2O2[M+H] + :461.33. Compound 6 was identified as ent-pimarane-8(14)-ene-15,16-propanone ester and [3,2,c][1'-phenylpyrazole].

[0073] Example 7

[0074] Compound 7: Preparation of enantio-1'-trifluoromethyl-1'-hydroxy-pimarane-2(1'),8(14)-diene-3-one-15,16-acetone complex

[0075] The structural formula of enantio-1'-trifluoromethyl-1'-hydroxy-pimarane-2(1'),8(14)-diene-3-one-15,16-acetone complex is as follows:

[0076]

[0077] Synthesis route:

[0078] The dried ethyl formate in Example 3 was replaced with 4.16 mmol of ethyl trifluoromethyl formate, and the synthesis was carried out according to the method of Example 3 to obtain 513 mg of pure product with a yield of 80.9%.

[0079] The above pure product was subjected to thin layer chromatography (TLC) and nuclear magnetic resonance detection, and the results were as follows: mp: 105-108°C. (c 0.1, MeOH). 1 H NMR (CDCl3, 400MHz):δ Η 15.7(1H,s,-OH),5.21(1H,s,H-14),4.00(1H,t,J=6.9Hz,H-15),4.17(1H,dd,J=7.6,6.9Hz,H a -16),3.75(1H,t,J=7.6Hz,Hb -16),2.65(1H,d,J=13.6Hz),2.33(1H,brd,J=13.6Hz),2.09-2.01(2H,m),1.79-1.60(5H,m),1.50-1.38(2H,m,overlap),1.42( 3H,s,Me-22),1.35(3H,s,Me-23),1.22(1H,m),1.26(3H,s,Me-18),0.96(3H,s,Me-17),1.19(3H,s,Me-19),0.76(3H,s,Me-20); 13 C NMR (CDCl3, 100 MHz): δ C 196.0(C-3),179.2(=C-OH),117.5(-CF3),137.9(C-8),127.2(C-14),108.6(C-21),102.0(C-2),80.4(C-15),65.3(C-16),5 1.3,47.4,40.8,37.4,36.0,35.8,31.3,29.5(C-18),26.2(C-22),25.0(C-23),24.0,22.5(C-19),21.9(C-17),13.4(C-20). ESI-MS m / z C 25 H 35 F3O4[MH] - :455.31. Compound 7 was identified as enantio-1'-trifluoromethyl-1'-hydroxy-pimarane-2(1'),8(14)-diene-3-one-15,16-propanone complex.

[0080] Example 8

[0081] Compound 8: Preparation of ent-pimarane-8(14)-ene-15,16-propanone and 3,2,c-(3'-trifluoromethyl-1',2'isoxazol-3'-ol)

[0082] The structural formulas of ent-pimarane-8(14)-ene-15,16-propanone and 3,2,c-(3'-trifluoromethyl-1',2'isoxazol-3'-ol) are as follows:

[0083]

[0084] Synthesis route:

[0085] 50 mg (0.11 mmol) of compound 7 (enanti-1'-trifluoromethyl-1'-hydroxy-pimarane-2(1'),8(14)-diene-3-one-15,16-acetone complex) prepared in Example 7 was taken and the synthesis method of compound 4 in Example 4 was followed to obtain 32 mg of pure white product with a yield of 62.0%.

[0086] The above pure product was subjected to thin layer chromatography (TLC) and nuclear magnetic resonance detection, and the results were as follows: mp: 211-214°C. (c 0.1, MeOH). 1 H NMR (CDCl3, 400MHz) showed characteristic signals: δ Η 5.16(1H,s,H-14),4.02(1H,t,J=6.9Hz,H-15),3.92(1H,dd,J=7.6,6.9Hz,H a -16),3.77(1H,t,J=7.6Hz,H b -16),3.42(1H,dd,J=13.4,5.4Hz,H-2),2.34(1H,d,J=13.6Hz),2.06(1H,t,J =10.0Hz),1.97(1H,dd,J=13.2,5.4Hz),1.80-1.77(2H,m),1.72-1.50(4H,m), 1.42(3H,s,Me-22),1.36(3H,s,Me-23),1.34(3H,s,Me-18),1.34-1.23(2H,m ),1.18-1.16(1H,m,overlap),1.17(3H,s,Me-19),0.93(6H,s,Me-17,Me-20); 13 C NMR (CDCl3, 100 MHz): δ C 167.7(C-3),138.2(C-8),128.1(C-14),123.9(-CF3),108.8(C-21),102.6(OC-CF3),79.8(C-15),65.3(C-16),53.9,49.5,4 6.9,38.8,37.4,35.9,35.6,31.6,26.9(C-18),26.2(C-22),25.2(C-23),23.4(C-19),22.5(C-17),22.0,18.4,14.8(C-20). ESI-MS m / z C 25 H 35 F3NO4[MH] -:470.29. Compound 8 was identified as ent-pimarane-8(14)-ene-15,16-propanone and 1,2-difluoro[3,2,c](3'-trifluoromethyl-1',2'isoxazol-3'-ol).

[0087] Example 9

[0088] Compound 9: Preparation of ent-pimarane-8(14)-ene-15,16-propanone ester conjugate [3,2,c](3'-trifluoromethyl-1',2'pyrazol-3'-ol)

[0089] The structural formula of ent-pimarane-8(14)-ene-15,16-propanone ester and [3,2,c](3'-trifluoromethyl-1',2'pyrazol-3'-ol) is as follows:

[0090]

[0091] 50 mg (0.11 mmol) of compound 7 (enanti-1'-trifluoromethyl-1'-hydroxy-pimarane-2(1'),8(14)-diene-3-one-15,16-acetone complex) prepared in Example 7 was taken and synthesized according to the method of compound 5 in Example 5 to obtain 28 mg of pure white product with a yield of 51.5%.

[0092] The above pure products were subjected to thin layer chromatography (TLC) and nuclear magnetic resonance detection, and the results are as follows: (c0.1, MeOH). 1 H NMR (CDCl3, 400MHz): δ Η 5.17(1H,s,H-14),4.03(1H,t,J=6.9Hz,H-15),3.92(1H,dd,J=7.6,6.9Hz,H a -16),3.78(1H,t,J=7.6Hz,H b -16),3.19(1H,dd,J=13.8,5.4Hz,H-2),2.34(1H,d,J=8.4Hz),2.09-2.03(1H,m),1 .97-1.92(1H,dd,J=12.8,5.6Hz),1.81-1.77(2H,m),1.72-1.60(3H,m),1.43-1.41 (1H,m,overlap),1.43(3H,s,Me-22),1.37(3H,s,Me-23),1.28(3H,s,Me-18),1.28 -1.26(1H,m,overlap),1.09(3H,s,Me-19),0.97(3H,s,Me-17),0.94(3H,s,Me-17); 13C NMR (CDCl3, 100MHz):δ C 163.6(C-3),138.4(C-8),127.9(C-14),125.3(-CF3),108.7(C-21),92.3(OC-CF3),79.8(C-15),65.3(C-16),54.0,49.7,44.7 ,38.9,37.7,35.9,35.8,35.2,31.7,27.2(C-18),26.3(C-22),25.2(C-23),23.7(C-19),22.5(C-17),22.4,18.4,14.9(C-20). ESI-MS m / z C 25 H 36 F3N2O3[MH] - :469.41. Compound 9 was identified as ent-pimarane-8(14)-ene-15,16-propanone conjugate and [3,2,c](3'-trifluoromethyl-1',2'pyrazol-3'-ol).

[0093] Example 10

[0094] Compound 10: Preparation of 2-hydroxypimarane-1,8(14)-diene-3-one-15,16-acetone complex

[0095] The structural formula of enantio-2-hydroxypimarane-1,8(14)-diene-3-one-15,16-propanone complex is as follows:

[0096]

[0097] Synthesis route:

[0098] 100 mg (0.28 mmol) of compound 2 (enantio-pimarane-8(14)-ene-3-one-15,16-acetone complex) prepared in Example 2 was taken, the dried ethyl formate in Example 3 was replaced with 500 mL of analytically pure ethyl formate, and the product was synthesized according to the method of Example 3 to obtain 78 mg of an oily compound with an enol structure having a double bond formed in the ring, with a yield of 75.1%.

[0099] The above oily compounds were subjected to thin layer chromatography (TLC) and nuclear magnetic resonance detection, and the results were as follows: (c 0.1, MeOH). 1 H NMR (Pyridine-d5, 400MHz):δ Η6.30(1H,s,H-1),5.20(1H,s,H-14),4.02(1H,t,J=6.7Hz,H-15),3.91(1H,dd,J=7.6,6.7Hz,H a -16),3.77(1H,t,J=7.6Hz,H b -16),2.35(1H,brd,J=14.0Hz),2.13(1H,td,J=14.0,4.8Hz),2.00(1H,t,J=7.4Hz),1.87-1.48(7H,m),1.23(3H ,s,Me-18),1.17(3H,s,Me-19),1.01(3H,s,Me-17),0.93(3H,s,Me-20),1.42(3H,s,Me-22),1.36(3H,s,Me-23); 13 C NMR (Pyridine-d5, 100 MHz): δ C 200.8(C-3),144.1(C-2),137.8(C-8),129.1(C-14),124.8(C-1),108.7(C-21),79.8(C-15),65.3(C-16),52.3,47.3,4 3.9,39.2,36.0,35.6,31.6,26.23(C-18),26.19(C-22),25.2(C-23),22.6(C-19),22.5(C-17),21.8,18.5,17.3(C-20). ESI-MS m / z C 23 H 33 O4[MH] - :373.43. Compound 10 was identified as 2-hydroxypimarane-1,8(14)-diene-3-one-15,16-propanone complex.

[0100] Example 11

[0101] Compound 11: Preparation of 2-bromo-enantio-pimarane-8(14)-ene-3-one-15,16-propanone complex

[0102] The structural formula of 2-bromo-enantio-pimarane-8(14)-ene-3-one-15,16-propanone complex is as follows:

[0103]

[0104] Synthesis route:

[0105] 1 g (2.78 mmol) of compound 2 (enantio-pimarane-8(14)-ene-3-one-15,16-acetonide) prepared in Example 2 was weighed and placed in a 100 mL dry eggplant-shaped flask. Dry tetrahydrofuran (THF) was added to dissolve the mixture. 1.12 g (3.33 mmol) of pyridinium tribromide was added under ice bath and stirred for 1 h. No starting material was detected by TLC. The reaction solution was evaporated under reduced pressure and the residue was dissolved in 30 mL of ethyl acetate. The mixture was washed with saturated NaHCO3 solution and water three times, 30 mL each time, and then dried over anhydrous Na2SO4. Na2SO4 was removed by suction filtration and the solvent was evaporated under reduced pressure to obtain a crude product. The crude product was chromatographed on a silica gel column using petroleum ether:ethyl acetate (5:1, v:v) as eluent to obtain 956 mg of the pure product with a yield of 78.5%.

[0106] The above pure product was subjected to thin layer chromatography (TLC) and nuclear magnetic resonance detection, and the results were as follows: mp: 129-130°C. (c 0.1, MeOH). 1 H NMR (CDCl3, 400MHz):δ Η 5.69(1H,dd,J=13.8,5.2Hz,H-2),5.31(1H,s,H-14),4.19(1H,t,J=6.9Hz,H-15),4.09(1H,dd,J=7.7,6.9Hz,H a -16),3.94(1H,t,J=7.7Hz,H b -16),2.62(1H,dd,J=12.6,5.4Hz),2.27(1H,brd,J=14.2Hz),2.06-1.97(2H,m),1.90-1.86(1H,m),1.72(1H,J=7.6Hz),1.53(3H,s,Me -22),1.47(3H,s,Me-23),1.50-1.30(5H,m,overlap),1.21(3H,s,Me-18),1.15-1.08(1H,m),1.04(9H,overlap,Me-17,Me-19,Me-20); 13 C NMR (CDCl3, 100MHz):δ C206.0(C-3),138.2(C-8),128.5(C-14),108.7(C-21),80.3(C-15),65.6(C-16),55.5,54.6(C-2),50.5,49.3,49.1 ,41.1,36.2,35.6,31.9,26.6(C-22),26.4(C-18),25.5(C-23),23.0,22.9(C-19),22.0(C-17),18.5,14.8(C-20). ESI-MS m / z C 23 H 35 BrO3[MH] - :437.30. Compound 11 was identified as 2-bromo-en-pimarane-8(14)-ene-3-one-15,16-propanone complex.

[0107] Example 12

[0108] Compound 12: Preparation of ent-pimarane-8(14)-ene-15,16-propanone ester and [3,2,d] (2'-aminothiazole) The structural formula of ent-pimarane-8(14)-ene-15,16-propanone ester and [3,2,d] (2'-aminothiazole) is as follows:

[0109]

[0110] Synthesis route:

[0111] 50 mg (0.11 mmol) of compound 11 (2-bromo-enantio-pimarane-8(14)-ene-3-one-15,16-acetonide) prepared in Example 11 was weighed and placed in a 100 mL dry eggplant-shaped flask. 4 mL of ethanol and 12 mg (0.16 mmol) of thiourea were added and dissolved by stirring at room temperature. Heat and reflux for 3 h. The reaction solution was cooled and alkalized by adding saturated NaHCO3 solution. The reaction solution was concentrated under reduced pressure to remove ethanol from the reaction system. 20 mL of water was added to disperse the solution and the solution was extracted with ethyl acetate three times, each time with 20 mL. The ethyl acetate layers were combined and concentrated under reduced pressure to obtain a crude product. Purification by silica gel column chromatography with petroleum ether:ethyl acetate (5:1, v:v) as eluent gave 32 mg of the pure product with a yield of 67.4%.

[0112] The above pure products were subjected to thin layer chromatography (TLC) and nuclear magnetic resonance detection, and the results are as follows: (c0.1, MeOH). 1 H NMR (CDCl3, 400MHz) showed characteristic signals: δ Η5.18(1H,s,H-14),4.00(1H,t,J=6.7Hz,H-15),3.91(1H,dd,J=7.6,6.7Hz,H a -16),3.76(1H,t,J=7.6Hz,H b -16),2.58(1H,d,J=11.4Hz),2.36-2.30(2H,m),2.07(1H,t,J=11.4Hz),1.86(1H,m),1.76-1.60(4H,m),1.56-1.40(2H,m,overlap),1.42 (3H,s,Me-22),1.35(3H,s,Me-23),1.27(1H,m,overlap),1.22(3H,s,Me-18),1.13(3H,s,Me-19),0.95(3H,s,Me-17),0.84(3H,s,Me-20); 13 C NMR (CDCl3, 100MHz):δ C 164.6(-N=CS-),151.8(C-2),139.0(C-8),126.4(C-14),115.7(C-14),108.7(C-21),80.6(C-15),65.5(C-16),51.8,48.2,4 0.0,37.0,36.5,36.1,36.0,31.6,30.8(C-18),26.3(C-22),25.2(C-23),24.0,22.9(C-19),22.5(C-17),18.1,14.5(C-20). ESI-MS m / z C 24 H 36 N2O2S[M+H] + :417.32. Compound 12 was identified as ent-pimarane-8(14)-ene-15,16-propanone ester and [3,2,d] (2'-aminothiazole).

[0113] Example 13

[0114] Computer-aided drug design: computational and virtual screening based on FXa targets

[0115] (1) Principle: Factor Xa is the rate-limiting component of thrombin generation. Due to the amplification of biological signals during the blood coagulation cascade, it is estimated that one molecule of factor Xa inhibitor can inhibit the physiological effects of 138 thrombin molecules. Factor Xa inhibitors are more effective than thrombin inhibitors. Factor Xa simply promotes coagulation and does not have the side effects of thrombin inhibitors. Therefore, factor Xa inhibitors have become one of the hot topics in anticoagulant drug research in recent years.

[0116] FXa consists of two chains, a heavy chain (B chain, 241 amino acid residues) and a light chain (A chain, 139 amino acid residues). FXa is mainly divided into four subunits: S1, S2, S3, and S4. S1 is the specific binding pocket, and S4 is the aromatic pocket of FXa. S1 and S4 are the main sites for binding to inhibitors.

[0117] (2) Process: The Glide docking module in Maestro.10.6 software was used for simulation calculations, with G-score as the docking result evaluation score, and the docking was performed using the default standard method of Glide. With the original ligand as the center, the generated length was The workspace ligand was set to XP (Extra precision). The docking results were evaluated using the Docking score function in Glide.

[0118] (3) The results are as follows Figure 1-4 As shown: 15-OH of sirucinol acts as a hydrogen bond donor to form a hydrogen bond with the oxygen atom of the Glu 97 side chain, and 16-OH of the ligand acts as a hydrogen bond donor to form a hydrogen bond interaction with the oxygen atom on the Lys96 side chain ( Figure 1 The benzene ring of compound 6 forms Pi-Pi interactions with Trp215, Tyr 99, and Phe174, and the thiazole ring forms Pi-Pi interactions with Tyr 99 ( Figure 2 ). The 5-OH group of the isoxazole of compound 9 and the oxygen atom on the side chain of Glu 97 form a hydrogen bond interaction ( Figure 3 The amino group on the thiophene ring of compound 12 acts as a hydrogen bond donor and forms a hydrogen bond with the oxygen atom on the side chain of Thr 98. The imine on the thiophene ring is ionized to form a positive ion conjugated structure, forming an electrostatic interaction with Trp 215, Phe 174, and Tyr99 ( Figure 4 ).

[0119] (4) The docking result scores are shown in Table 1:

[0120] Table 1 Docking result scoring table

[0121]

[0122] Example 14

[0123] Pharmacodynamics test: In vitro inhibition test of compound on coagulation factor Xa

[0124] The Docking score considers hydrogen bonding, electrostatic interactions, van der Waals forces, and hydrophilic / hydrophobic interactions between the ligand and the receptor protein, and is a comprehensive evaluation of these factors. Therefore, the Docking score is valuable for screening compounds that effectively bind to the target. Based on previous pharmacological activity studies and the results of this Schrodinger Glide docking, we selected derivatives of siuguirenol and some derivatives of kiwanoalanol for in vitro inhibition of coagulation factor Xa, hoping to identify lead compounds with antithrombotic activity.

[0125] 1. Test instruments and reagents

[0126] (1) Instruments: FORMA 700 ultra-low temperature refrigerator, Thermo Fisher Scientific; YC-300L medicine storage cabinet, Zhongke Meiling Cryogenic Technology Co., Ltd.; Direct-Q ultrapure water analyzer with pump, Millopore; M200 full-wavelength microplate reader, Tecan.

[0127] (2) Reagents: Human FXa, Kordia (purchased by Jinan Xinlianxin Biotechnology Co., Ltd.); S-2765 chromogenic substrate, Boatman Biotech (purchased by Jinan Xinlianxin Biotechnology Co., Ltd.); bovine serum albumin (BSA), China National Pharmaceutical Group Co., Ltd.; DMSO, tris(hydroxymethyl)aminomethane hydrochloride (Tris), Sigma.

[0128] 2. Test methods

[0129] (1) Prepare BSA Buffer: Take 0.005 mol (0.605 g) of Tris, 0.01 mol (0.58 g) of NaCl, and 0.10 g of BSA, add 100 mL of triple-distilled water, and shake well to obtain a final concentration of 0.05 M Tris-0.1 M NaCl-0.1% BSA buffer (pH = 7.4). Set aside.

[0130] (2) Preparation of compound solution: Take 10 mmol of compound, add 100 μL of DMSO, shake to dissolve and mix, and obtain 10 5 mol / L stock solution of the compound. Dilute quantitatively to the specified concentration using the doubling dilution method.

[0131] (3) Determination of FXa inhibitory activity: In a 96-well plate, 10 μL of the prepared solution of different concentrations (10 nM, 100 nM, 1000 nM, 10000 nM, 100000 nM) was added to each well (10 μL of 0.1% DMSO was added to the blank group) and 25 μL of 0.003 IU / mL human FXa was mixed with 40 μL of BSA buffer (pH 7.4). Incubate at 37°C for 15 min. Then, 40 μL of the chromogenic substrate S-2765 was added, shaken for 10 s, and incubated at room temperature for 1 h. The absorbance of each well was measured at 405 nm using a microplate reader.

[0132] (4) Data statistics: FXa inhibitory activity = 1-[(OD / min)sample / (OD / min)control]. IC 50 The values ​​were calculated and analyzed using SPSS 19.0 software.

[0133] 3. The test results are shown in Table 2:

[0134] Table 2 Inhibitory effects of test compounds on FXa

[0135]

[0136]

[0137]

[0138] The test results show that the present invention evaluated the in vitro FXa inhibitory activity of a series of dauriculinol derivatives (compounds 1-12), and the results showed that compound 9 and compound 12 had significant inhibitory activity against FXa.

Claims

1. A compound, characterized in that The structural formula of the compound is any one of the following: 。 2. Use of the compound according to claim 1 in the preparation of a medicament for inhibiting coagulation factor Xa.

3. Use of the compound according to claim 1 in the preparation of a drug having antithrombotic activity.

4. Use of the compound according to claim 1 in the preparation of a medicament for treating and / or preventing thromboembolic diseases.

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