An ampk agonist and derivatives thereof, and methods of making and using the same
Patent Information
- Authority / Receiving Office
- CN · China
- Patent Type
- Patents(China)
- Current Assignee / Owner
- SHENYANG PHARMA UNIV
- Filing Date
- 2024-06-26
- Publication Date
- 2026-07-14
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Figure QLYQS_1 
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Abstract
Description
Technical Field
[0001] This invention relates to the field of pharmaceutical technology, particularly to the field of antitumor drug preparation technology, specifically to an AMPK agonist and its derivatives, their preparation methods, and uses. Background Technology
[0002] AMPK (AMP-activated protein kinase) is an enzyme that plays a crucial role in regulating cellular energy metabolism. It is activated when the ratio of AMP to ATP (adenosine triphosphate) in the cell increases, indicating a lower energy level. AMPK is present in all eukaryotes as a heterotrimeric αβγ complex. AMPK acts as a nutrient and energy sensor, maintaining cellular energy and metabolism, and playing a vital role in systemic energy homeostasis.
[0003] Metabolic alterations in cells are often hallmarks of chronic human diseases such as diabetes, Alzheimer's disease (AD), and cancer, and AMPK is closely related to cellular metabolism. Patients with metabolic disorders, such as insulin resistance, type 2 diabetes, and obesity, have a higher risk of developing cancer. Under energy stress, AMPK inhibits cell proliferation by activating catabolic processes and inhibiting anabolism. These observations suggest that AMPK is a tumor suppressor. However, AMPK mutations are rare in human cancers, suggesting that loss of AMPK function may be due to mutations in upstream kinases such as LKB1 or downstream targets such as TSC2.
[0004] Therefore, AMPK activation is a novel approach to cancer treatment. Activating AMPK through small molecules may be a feasible treatment method to restore energy and metabolic status and reverse disease phenotypes. Summary of the Invention
[0005] The purpose of this invention is to provide an AMPK agonist, its preparation method, and its uses. This AMPK agonist has excellent AMPK agonist activity and can be used as a novel antitumor drug for the prevention or treatment of tumors and their complications.
[0006] This invention provides an AMPK agonist or a pharmaceutically acceptable salt thereof, the general structural formula of which is shown in formula (I):
[0007]
[0008] in,
[0009] R1 is H, C1-C2 alkyl, C1-C2 alkoxy, halogen, cyano, hydroxy, or trifluoromethyl;
[0010] R2 is H, C1-C2 alkyl, C1-C2 alkoxy, halogen, cyano, hydroxy, amino, methyl, or trifluoromethyl; wherein, when X is C, R2 is H, C1-C2 alkyl, C1-C2 alkoxy, halogen, cyano, hydroxy, amino, methyl, or trifluoromethyl; when X is N, R2 is not present.
[0011] R3 is H, C1-C2 alkyl, C1-C2 alkoxy, halogen, cyano, hydroxy, or trifluoromethyl;
[0012] R4 is a carboxyl group, a carboxylic acid ester, an amide, a sulfonamide, or a methoxyoxime acid;
[0013] X is C or N;
[0014] Y is either C or N;
[0015] Z is OH or O;
[0016] It is a C3-C6 cycloalkyl or substituted or unsubstituted cycloheteroalkyl; wherein the heterocycle contains 1-3 N, O or S heteroatoms; the substitution is by 1, 2, 3 or 4 substituents selected from the following: C1-C2 alkoxy, halogen, cyano, hydroxy, amino, monofluoromethyl, difluoromethyl, trifluoromethyl, carboxyl or carboxymethyl ester.
[0017] Specifically, the AMPK agonists and their derivatives, or pharmaceutically acceptable salts thereof, described in this invention are specifically selected from the following compounds:
[0018]
[0019] The method for preparing the AMPK agonist provided by the present invention includes the following steps:
[0020] 1) React the compound of formula (II) with the compound of formula (III) to obtain the compound shown in formula (IV);
[0021]
[0022] 2) The compound of formula (V) is reacted with an acyl chloride and then cyclized, followed by reaction with pinacol diboronic acid ester to obtain the compound shown in formula (VI);
[0023]
[0024] 3) Couple the compound shown in formula (IV) with different substituted borate esters, i.e., formula (VI), to obtain the compound shown in general formula (VII);
[0025]
[0026] 4) Remove the protecting group of the compound shown in formula (VII) by SEM to obtain the compound shown in general formula (I);
[0027]
[0028] Further, the reaction process involved in step 1) is to dissolve the reactant shown in (II) in an organic solvent, then add a base and the compound shown in formula (III), react until the reaction is complete, then evaporate the solvent, extract, dry, concentrate the organic phase, and then purify it using a chromatography column.
[0029] Furthermore, the reaction process involved in step 2) is as follows:
[0030] 2)-1: Dissolve the compound shown in formula (V) in an organic solvent, add alkali after dissolution, add acyl chloride under ice-water bath conditions, stop the reaction after the reaction is complete, dilute directly with water, extract, dry, filter, and concentrate the organic phase to obtain intermediate A;
[0031] 2)-2: Dissolve the obtained intermediate A in an organic solvent, slowly add NaH under ice-water bath conditions, react at room temperature until the reaction of the raw materials is completely quenched, dilute, extract, wash, dry, filter and concentrate to obtain intermediate B;
[0032] 2)-3: The obtained intermediate B was dissolved in an organic solvent, and pinacol diboronic acid ester and a base were added, followed by a catalyst. The reaction was carried out under nitrogen protection. After the reaction was completed, the reaction was stopped, and the solvent was evaporated to obtain a concentrated solution. The compound shown in formula (VI) was purified by silica gel column chromatography.
[0033] Furthermore, the reaction process shown in step 3) is as follows:
[0034] The compounds shown in formula (IV) and (VI) were dissolved in an organic solvent, and a catalyst, ligand, base, and water were added to a reaction flask. After the reaction mixture was fully reacted under nitrogen protection, it was diluted, extracted, washed, and purified by column chromatography to obtain the compound shown in formula (VII).
[0035] Further, the reaction process shown in step 4) is as follows: after the compound shown in formula (VII) is dissolved in an organic solvent, a THF solution of tetrabutylammonium fluoride is added dropwise, and the reaction is heated. During the reaction, a THF solution of tetrabutylammonium fluoride needs to be added once. After the reaction is complete, the compound shown in formula (I) is obtained.
[0036] Furthermore, the organic solvent is selected from tetrahydrofuran (THF), methyltetrahydrofuran (MeTHF), dioxane, and N,N-dimethylformamide (DMF).
[0037] Furthermore, the alkali is selected from potassium carbonate (K2CO3), cesium carbonate (Cs2CO3), sodium carbonate (Na2CO3), sodium acetate (CH3COONa), and triethylamine (TEA).
[0038] Furthermore, the catalyst is selected from Pd(dppf)Cl2 and PdCl2(PPh3)2.
[0039] Furthermore, the ligands are selected from S-Phos, X-Phos, and Xant-Phos.
[0040] Furthermore, in step 1), the reaction temperature is room temperature, the molar ratio of formula (II) to base is 1:(2-3), and the molar ratio of formula (II) to formula (III) is 1:(1.1-2).
[0041] Furthermore, in step 2):
[0042] 2)-1 The reaction temperature is room temperature, the molar ratio of formula (V) to base is 1:(1.2~2), and the molar ratio of formula (V) to acyl chloride is 1:(1.1~1.3);
[0043] 2)-2 The reaction temperature is room temperature, and the molar ratio of intermediate A to sodium hydride is 1:2;
[0044] 2) The reaction temperature of -3 is 100-110℃, the molar ratio of intermediate B to pinacol diboronic acid is 1:(1.2-2), and the molar ratio of intermediate B to base is 1:(1.3-3).
[0045] Furthermore, in step 3), the reaction temperature is 80-120℃, the molar ratio of formula (IV) to formula (V) is 1:(1.2-1.6), the molar ratio of ligand to catalyst is 1:(0.8-1.1), the molar ratio of formula (IV) to catalyst is 1:(0.03-0.1), the molar ratio of (IV) to base is 1:(3-5), and the amount of water used is 3%-10% of the volume of organic solvent.
[0046] Furthermore, in step 4), the reaction temperature is 60–90°C, and the molar ratio of formula (VII) to tetrabutylammonium fluoride is 1:(8–20).
[0047] This invention provides the use of the AMPK agonist and its derivatives or pharmaceutically acceptable salts thereof in the preparation of a medicament for treating tumors.
[0048] The present invention also provides a pharmaceutical composition comprising the AMPK agonist or a pharmaceutically acceptable salt thereof and a pharmaceutically acceptable carrier.
[0049] The present invention provides the use of the pharmaceutical composition in the preparation of a drug for treating tumors.
[0050] The beneficial effects of this invention are:
[0051] This invention provides an AMPK agonist and a method for preparing this derivative, which is simple and easy to operate. These compounds exhibit excellent AMPK agonist and antitumor activity, and have significant practical value and application prospects in the field of antitumor drug preparation. Detailed Implementation
[0052] The present invention will be described below through specific embodiments, but the present invention is not limited thereto.
[0053] Unless otherwise specified, the experimental methods described in the following examples are conventional methods; the reagents and biological materials described are commercially available unless otherwise specified.
[0054] Example 15: Synthesis of 6-chloro-5-[4-(2-hydroxypyrrolidone)-phenyl]-1H-benzo[d]imidazol-2-yl}oxy}-2-methylbenzoic acid (compound 1)
[0055]
[0056] Step 1) Synthesis of 4-(2-oxo-1-pyrrolidinyl)phenylboronic acid pinacol ester
[0057]
[0058] 5.01 g (29.07 mmol) of 4-bromoaniline was added to a 250 mL pear-shaped flask equipped with a magnetic stirrer, followed by 90 mL of THF for dissolution. After dissolution, 4.41 g (43.60 mmol) of triethylamine was added, and 4.92 g (34.88 mmol) of 4-chlorobutyryl chloride was slowly added dropwise under ice-water bath conditions. The reaction mixture was then stirred at 25 °C for 2 hours until the reactants were completely reacted. The mixture was diluted directly with 100 mL of water and extracted with ethyl acetate (2 × 50 mL). The organic phase was washed with saturated NaCl solution and dried with anhydrous sodium sulfate. After filtration, the organic phase was concentrated to obtain 7.20 g of N-(4-bromophenyl)-4-chlorobutyramide as a grayish-white solid, with a yield of 90.31%.
[0059] 7.20 g (26.03 mmol) of the obtained N-(4-bromophenyl)-4-chlorobutyramide was added to a 250 mL pear-shaped flask equipped with a magnetic stirrer, and 90 mL of THF was added to dissolve it. After dissolution, 1.25 g (52.07 mmol) of sodium hydride was slowly added under ice-water bath conditions, and then the mixture was stirred at 25 °C for 2 h until the reaction was complete. The reaction solution was quenched with 100 mL of saturated NH4Cl solution, diluted with 50 mL of water, and extracted with ethyl acetate (2 × 75 mL). The organic phase was washed with saturated NaCl solution, dried with anhydrous sodium sulfate, filtered, and concentrated to give 5.52 g of crude product 1-(4-bromophenyl)pyrrolidine-2-one, a grayish-white solid, with a yield of 89.02%.
[0060] 5.50 g (22.91 mmol) of the obtained 1-(4-bromophenyl)pyrrolidine-2-one was added to a 250 mL pear-shaped flask equipped with a magnetic stir bar, and 90 mL of THF was added to dissolve it. After dissolution, 8.73 g (34.36 mmol) of pinacol diboronate and 3.74 g (27.49 mmol) of cesium carbonate were added, followed by 0.80 g (1.14 mmol) of Pd(dppf)Cl2. Under nitrogen protection, the reaction was heated to 100 °C and stirred for 24 h. After the reaction was completed by TLC, the solvent was evaporated to obtain a concentrated solution. The solution was purified by silica gel column chromatography (20% ethyl acetate / petroleum ether) to obtain 4.23 g of compound 4-(2-oxo-1-pyrrolyl)phenylboronate pinacol ester, with a yield of 66.02%.
[0061] The substituted borate esters required in subsequent Examples 7, 8, 9, 10 and 11 were all prepared using different substituted starting materials via this route.
[0062] The structural verification data is as follows:
[0063] 1 H NMR (400MHz, DMSO-d6) δ7.69-7.62(m,4H),3.86-3.78(m,2H),2.51-2.44(m,2H),2.05(p,J=8.0Hz,2H),1.28(s,12H).
[0064] ESI-MS: (m / z) 288.0 [M+H] + .
[0065] Step 2) Synthesis of methyl 5-methyl-((5-bromo-6-chloro-1-((2-(trimethylsilyl)ethoxy)methyl)-1H-benzo[d]imidazol-2-yl)oxy)-2-benzoate
[0066]
[0067] 13.00 g (30.71 mmol) of 6-chloro-5-bromo-2-(methanesulfonyl)-1-((2-(trimethylsilyl)ethoxy)methyl)-1H-benzo[d]imidazole was added to a 500 mL round-bottom flask equipped with a magnetic stirrer and dissolved in 250 mL of DMF. After dissolution, 10.61 g (76.76 mmol) of K₂CO₃ and 7.01 g (46.06 mmol) of methyl 5-hydroxy-2-methylbenzoate were added. The reaction mixture was reacted at 25 °C for 24 hours until the reactants were completely reacted. The solvent was evaporated to dryness, diluted with 250 mL of water, and extracted with ethyl acetate (3 x 300 mL). The organic phase was washed with saturated NaCl solution and dried over anhydrous sodium sulfate. The concentrated organic phase was purified by column chromatography (25% ethyl acetate / petroleum ether) to give 11.40 g of a white solid, with a yield of 75.41%.
[0068] The structural verification data is as follows:
[0069] ESI-MS: (m / z) 511.0 [M+H] + .
[0070] Step 3) Synthesis of 5-{{6-chloro-5-[4-(2-hydroxypyrrolidone)-phenyl]-1H-benzo[d]imidazol-2-yl}oxy}-2-methylbenzoic acid
[0071]
[0072] 0.163 g (0.57 mmol) of the prepared 4-(2-oxo-1-pyrrolyl)phenylboronic acid pinacol ester and 0.20 g (0.38 mmol) of 5-methyl-((6-chloro-5-bromo-1-((2-(trimethylsilyl)ethoxy)methyl)-1H-benzo[d]imidazol-2-yl)oxy)-2-benzoate methyl ester were added to a 50 mL reaction flask. 25 mL of LDM was added to dissolve the ester, followed by the addition of Pd(dppf)Cl2 (0.013 g, 0.019 mmol), X-Phos 0.009 g (0.019 mmol), and 1 M K2CO3 solution (2.4 mL). The reaction mixture was heated at 120 °C for 4 h under nitrogen protection until the reactants reacted completely. The solvent was concentrated under reduced pressure, the concentrate was diluted with 60 mL of water, and then extracted with ethyl acetate (2 x 100 mL). The organic layer was washed with saturated NaCl solution and then concentrated. The concentrate was purified by column chromatography (10-30% ethyl acetate / petroleum ether) to give 0.11 g of a pale yellow solid.
[0073] Add 5 mL (2.08 mmol) of tetrabutylammonium fluoride (1.0 M THF solution) dropwise to a 20 mL THF solution containing 0.11 g (0.164 mmol) of the product from the previous step using a syringe to a 50 mL pear-shaped flask equipped with a magnetic stirrer. Place the reaction mixture in an oil bath and heat at 80 °C for 45 minutes. Then add 3 mL of tetrabutylammonium fluoride to the reaction mixture. Continue heating the reaction mixture for 4 hours until the reactants are completely reacted. After cooling, dilute the reaction mixture with 200 mL of ethyl acetate and saturated water containing KHSO4 (pH 3). Extract the aqueous phase with 100 mL of ethyl acetate. Wash the combined organic phases with water and saturated NaCl solution. Dry the organic phase with anhydrous sodium sulfate, filter, and concentrate the organic phase. Dissolve the concentrate in 50 mL of methanol. Add 10 mL of 2.5 N NaOH to the reaction mixture and heat at 45 °C for 2 hours until the reactants are completely reacted. After the reaction flask cooled, the reaction solution was diluted with 50 mL of water. The aqueous phase was washed with ethyl acetate (2 x 100 mL) and acidified to pH ~1 with 2N HCl. The aqueous phase was then extracted with ethyl acetate (2 x 100 mL), and the combined organic layers were washed with saturated NaCl solution. The organic layer was dried with anhydrous sodium sulfate, filtered, and the organic phase was concentrated. The product was eluted using a reversed-phase silica gel column with an eluent of (30% MeOH / H2O) to give 0.05 g of compound 1, with a yield of 61.20%.
[0074] The structural verification data is as follows:
[0075] 1 H NMR (400MHz, DMSO-d6) δ7.81(d,J=2.0Hz,1H),7.80(d,J=2.8Hz,1H),7.79(s,1H),7.61(s,1H),7.52(d,J=2.5Hz,1H),7.51(d,J=4 .2Hz,1H),7.50(s,1H),7.43(d,J=8.5Hz,1H),7.41(s,1H),3.96(t,J=7.0Hz,2H),2.62(s,3H),2.60(t,2H),2.17(p,J=7.4Hz,2H).
[0076] 13 C NMR(100MHz,DMSO-d6)δ174.38,169.01,158.41,151.50,139.13,136.09,135.65,133.08,133.0 2(2C),130.34(3C),124.56,123.28(2C),121.87(2C),119.32(3C),48.52,32.83,21.02,17.93.
[0077] HRMS (ESI) - )m / z calcd for C 25 H 20 ClN3O4[MH] - ,460.1070; found,460.1115.
[0078] Example 2 Synthesis of methyl 5-{{6-chloro-5-[4-(2-hydroxypyrrolidone)phenyl]-1H-benzo[d]imidazol-2-yl}oxy}-2-methylbenzoate (compound 2)
[0079]
[0080] The prepared 4-(2-oxo-1-pyrrolyl)phenylboronic acid pinacol ester (0.163 g, 0.57 mmol) and methyl 5-methyl-{{6-chloro-5-bromo-1-{[2-(trimethylsilyl)ethoxy]methyl}-1H-benzo[d]imidazol-2-yl}oxy}-2-benzoate (0.2 g, 0.38 mmol) were added to a 50 mL reaction flask, dissolved in 25 mL of LDM, and then Pd(dppf)Cl2 (0.013 g, 0.019 mmol), X-Phos (0.009 g, 0.019 mmol), and 1 M K2CO3 solution (2.4 mL) were added to the reaction flask. Under nitrogen protection, the reaction mixture was heated at 120 °C for 4 h until the reactants reacted completely. The solvent was concentrated under reduced pressure, the concentrate was diluted with 60 mL of water, and then extracted with ethyl acetate (2 x 100 mL). The organic layer was washed with saturated NaCl solution and then concentrated. The concentrate was purified by column chromatography (10-30% ethyl acetate / petroleum ether) to give 0.18 g of a pale yellow solid.
[0081] Subsequently, as in Example 1, tetrabutylammonium fluoride (1.0M THF solution) was added dropwise to the product solution from the previous step into a 50mL pear-shaped flask equipped with a magnetic stirrer using a syringe. The reaction mixture was placed in an oil bath and heated at 80°C for 45 minutes, after which tetrabutylammonium fluoride was added to the reaction mixture. The reaction mixture was then heated continuously for 4 hours until the reactants reacted completely. After cooling, the reaction mixture was diluted with ethyl acetate and KHSO4 (~pH3) saturated water, and the aqueous phase was extracted with ethyl acetate. Compound 2 was obtained.
[0082] The structural verification data is as follows:
[0083] 1H NMR(400MHz,DMSO-d6)δ7.97–7.92(m,1H),7.89(dt,J=7.6,1.4Hz,1H),7.73 (d,J=2.3Hz,2H),7.71(d,J=2.4Hz,2H),7.64(t,J=7.9Hz,1H),7.56(s,1H), 7.44(d,J=2.1Hz,1H),7.42(d,J=1.6Hz,1H),7.35(s,1H),3.87(s,4H),3.84 (d,J=15.9Hz,2H),2.53(d,J=8.0Hz,2H),2.52(s,4H),2.09(q,J=7.4Hz,1H).
[0084] 13 C NMR(100MHz,DMSO-d6)δ167.39,167.26,152.00,151.83,137.83,137.40,134.07(2C),134.00(2C), 133.17,132.44(2C),131.39,131.35,129.59,125.31(2C),122.77(3C),53.07,30.93,21.36,19.58.
[0085] HRMS (ESI) - )m / z calcd for C 26 H 22 ClN3O4[MH] - ,474.1226;found,474.1147.
[0086] Example 35: Synthesis of 6-chloro-5-[4-(2-hydroxypyrrolidone)phenyl]-1H-benzo[d]imidazol-2-yl}oxy}-N-hydroxy-2-methylbenzamide
[0087]
[0088] Tetrahydropyranylhydroxylamine (1.5 eq) was added to a DMF solution of compound 1, followed by triethylamine (2.0 eq) and HATU (1.5 eq), and the mixture was stirred at room temperature for 14 hours. Extraction was performed with ethyl acetate (2 x 100 mL) and water, the organic layer was washed with saturated NaCl solution, dried over anhydrous sodium sulfate, and concentrated under reduced pressure to obtain several layers. The concentrate was dissolved in methanol and toluenesulfonic acid monohydrate (1.5 eq). The reaction was stirred at room temperature for 2 hours, followed by addition of saturated NaCl solution and extraction with ethyl acetate. The organic phase was dried over anhydrous sodium sulfate and concentrated. The concentrate was purified by column chromatography (0-10% MeOH-CH2Cl2) to obtain compound 3.
[0089] The structural verification data is as follows:
[0090] 1 H NMR(600MHz,DMSO-d6)δ11.00(s,1H),8.29(s,1H),8.00(s,1H),7.78(s, 1H),7.77(s,1H),7.59(s,1H),7.48(s,1H),7.47(s,1H),7.44(dd,J=8.2, 2.7Hz,1H),7.40(d,J=8.5Hz,1H),7.38(s,1H),7.36(d,J=2.5Hz,1H),3.9 3(t,J=6.9Hz,2H),2.58(d,J=7.9Hz,2H),2.41(s,3H),2.16–2.12(m,2H).
[0091] 13 C NMR (150MHz, DMSO-d6) δ174.38,165.52,162.78,158.27,151.31,139.13,136.19,135.62,133.44,133.1 1,132.24(2C),130.34(2C),124.58,121.94(2C),119.61(2C),119.32(2C),48.52,32.83,19.11,17.93.
[0092] HRMS (ESI) - )m / z calcd for C 25 H 21 ClN4O4[MH] - ,475.1179;found,475.1096.
[0093] Example 45: Synthesis of 6-chloro-5-[4-(2-hydroxypyrrolidone)phenyl]-1H-benzo[d]imidazol-2-yl}oxy}-2-methylbenzamide (compound 4)
[0094]
[0095] Using compound 1 (0.08 g, 0.173 mmol) as a starting material, a THF solution of NH3 was added to a reaction flask, and THF was added as a solvent. HATU (1.5 eq) and DIPEA (3 eq) were added to the reaction flask sequentially. After reacting at room temperature for 12 h, the starting material was completely reacted. The solvent was removed under vacuum, and the product was purified by column chromatography to obtain 0.1 g of compound 5-{{6-chloro-5-[4-(2-hydroxypyrrolidone)phenyl]-1H-benzo[d]imidazol-2-yl}oxy}-2-methylbenzamide (compound 4), with a yield of 58.67%.
[0096] The structural verification data is as follows:
[0097] 1 H NMR(600MHz,DMSO-d6)δ12.85(s,1H),8.01(s,1H),7.89(s,1H),7.78(dd,J=13 .9,8.1Hz,2H),7.59(d,J=36.9Hz,1H),7.52(s,1H),7.49(d,J=8.4Hz,1H),7.4 4(d,J=2.5Hz,1H),7.41(d,J=2.6Hz,1H),7.39(s,1H),7.37(d,J=9.9Hz,1H),3 .94(t,J=6.5Hz,2H),2.58(d,J=8.0Hz,2H),2.45(s,3H),2.14(t,J=7.8Hz,2H).
[0098] 13 C NMR(150MHz,DMSO-d6)δ170.43,162.79,151.30,139.16,138.54,132.93,132.27(2C),130.34(2C),124. 58,121.65(2C),120.08,119.38(2C),119.31(2C),118.52,112.96,111.62,48.52,31.24,19.48,18.41.
[0099] HRMS (ESI) - )m / z calcd for C 25 H 21 ClN4O3[MH] -,459.1229;found,459.1157.
[0100] Example 55 Synthesis of 6-chloro-5-[4-(2-hydroxypyrrolidone)phenyl]-1H-benzo[d]imidazol-2-yl}oxy}-N,2-dimethylbenzamide (compound 5)
[0101]
[0102] 0.08 g of 5-{{6-chloro-5-[4-(2-hydroxypyrrolidone)phenyl]-1H-benzo[d]imidazol-2-yl}oxy}-N,2-dimethylbenzamide (compound 5) was prepared by the same method as that used to prepare compound 4, with a yield of 68.74%.
[0103] The structural verification data is as follows:
[0104] 1 H NMR (400MHz, DMSO-d6) δ12.67(s,1H),8.30(q,J=4.6Hz,1H),7.77(d,J=8.4Hz,2H),7.59(s,1H),7.50–7.45(m,2H),7.41(d,J =2.5Hz,1H),7.39(s,3H),3.93(t,J=7.0Hz,2H),2.81–2.78(m,3H),2.58(d,J=8.0Hz,2H),2.40(s,3H),2.15(q,J=7.4Hz,2H).
[0105] 13 C NMR(100MHz,DMSO-d6)δ174.41,168.91,158.42,151.35,139.17,138.68,135.56,132.24(2C),130.34( 2C),124.62,121.65(2C),119.36(2C),119.33(3C),118.56,112.92,48.52,32.82,26.42,19.31,17.93.
[0106] HRMS (ESI) - )m / z calcd for C 26 H 23 ClN4O3[MH] - ,473.1386; found,473.1299.
[0107] Example 65: Synthesis of 6-chloro-5-[4-(2-hydroxypyrrolidone)phenyl]-1H-benzo[d]imidazol-2-yl}oxy}-N-cyclopropyl-2-methylbenzamide (compound 6)
[0108]
[0109] Compound 5-{{6-chloro-5-[4-(2-hydroxypyrrolidone)phenyl]-1H-benzo[d]imidazol-2-yl}oxy}-N-cyclopropyl-2-methylbenzamide (compound 6) was prepared by the same method as compound 4, with a yield of 61.25%.
[0110] The structural verification data is as follows:
[0111] 1 H NMR (400MHz, DMSO-d6) δ12.60(s,1H),8.37(d,J=4.3Hz,1H),7.78(d,J=26.1Hz,1H),7.71(d,J=9. 0Hz,2H),7.56(d,J=5.8Hz,1H),7.47(d,J=14.5Hz,1H),7.43–7.41(m,1H),7.35(d,J=2.9Hz,1H),7 .30(d,J=3.1Hz,1H),7.29(d,J=2.9Hz,1H),3.89(dd,J=7.2,4.0Hz,2H),2.85–2.80(m,1H),2.53(d ,J=8.1Hz,2H),2.33(d,J=2.7Hz,3H),2.12–2.07(m,2H),0.94–0.81(m,2H),0.67(d,J=7.4Hz,2H).
[0112] 13 C NMR(100MHz,DMSO-d6)δ174.40,169.58,151.29,139.18,138.66,135.55,133.19,132.91,132.14(2C),130.34 (2C),124.62,121.71,119.41,119.32,118.55,112.89,111.57,48.52,32.83,29.50,19.17,17.93,6.08(2C).
[0113] HRMS (ESI) - )m / z calcd for C 28 H 25 ClN4O3[MH] -,499.1542; found,499.1458.
[0114] Example 75 - Synthesis of 6-chloro-5-[3-methyl-4-(2-oxopyrrolidone-1-yl)phenyl]-1H-benzo[d]imidazol-2-yl}oxy}-2-methylbenzoic acid (compound 7)
[0115]
[0116] Using the method in step 1 of Example 1, 3-methyl-4-(2-oxo-1-pyrrolidinyl)phenylboronic acid pinacol ester was prepared from 2-methyl-4-bromoaniline as the starting material.
[0117] Compound 5-{{6-chloro-5-[3-methyl-4-(2-oxopyrrolidone-1-yl)phenyl]-1H-benzo[d]imidazol-2-yl}oxy}-2-methylbenzoic acid (compound 7) was prepared by the same method as compound 1, with a yield of 50.08%.
[0118] The structural verification data is as follows:
[0119] 1 H NMR (400MHz, DMSO-d6) δ12.68(s,1H),7.78(d,J=2.7Hz,1H),7.56(s,1H),7.51(dd,J=8.3,2.8Hz,1H),7.41(d,J=8.5Hz,1H ),7.33(s,2H),7.27(s,2H),3.72(t,J=6.9Hz,2H),2.55(s,3H),2.44(t,J=8.0Hz,2H),2.20(s,3H),2.14(q,J=7.5Hz,2H).
[0120] 13 C NMR(100MHz,DMSO-d6)δ173.92,168.26,151.51,139.18,137.65,136.82,135.42,133.40(2C),133.10,132. 34(2C),132.11,128.35(2C),126.67(2C),124.55,124.27,122.24(2C),50.52,31.24,21.23,19.18,18.20.
[0121] HRMS (ESI) - )m / z calcd for C 26 H 23 ClN4O3[MH] -,473.1386; found,473.1299.
[0122] Example 85: Synthesis of 6-chloro-5-[3-fluoro-4-(2-oxopyrrolidone-1-yl)phenyl]-1H-benzo[d]imidazol-2-yl}oxy}-2-methylbenzoic acid (compound 8)
[0123]
[0124] Using the method in step 1 of Example 1, 3-fluoro-4-(2-oxo-1-pyrrolidinyl)phenylboronic acid pinacol ester was prepared from 2-fluoro-4-bromoaniline as the starting material.
[0125] Compound 5-{{6-chloro-5-[3-fluoro-4-(2-oxopyrrolidone-1-yl)phenyl]-1H-benzo[d]imidazol-2-yl}oxy}-2-methylbenzoic acid (compound 8) was prepared by the method of preparing compound 1, with a yield of 61.69%.
[0126] The structural verification data is as follows:
[0127] 1 H NMR (400MHz, DMSO-d6) δ12.68(s,1H),7.78(d,J=2.7Hz,1H),7.56(s,1H),7.51(dd,J=8.3,2.8Hz,1H),7.41(d,J=8.5Hz,1H ),7.33(s,2H),7.27(s,2H),3.72(t,J=6.9Hz,2H),2.55(s,3H),2.44(t,J=8.0Hz,2H),2.20(s,3H),2.14(q,J=7.5Hz,2H).
[0128] 13 C NMR(100MHz,DMSO-d6)δ174.30,168.74,158.55,157.55,151.48,140.25,136.58,133.25(2C),131.99,131.77, 129.14,127.61,126.49,126.46,125.98,124.37,123.89,122.12,118.17,117.96,49.85,30.98,21.07,19.07.
[0129] HRMS (ESI) - )m / z calcd for C 26 H 22 ClN3O4[MH] -,474.1226;found,474.1145.
[0130] Example 95 - Synthesis of 6-chloro-5-[6-(2-oxopyrrolidone-1-yl)pyridin-3-yl]-1H-benzo[d]imidazol-2-yl}oxy}-2-methylbenzoic acid (compound 9)
[0131]
[0132] 2-(2-oxo-1-pyrrolidinyl)-5-bromopyridine was prepared from 2-amino-5-bromopyridine using the method in step 1 of Example 1, and then borate ester was prepared by the same method.
[0133] Compound 5-{{6-chloro-5-[6-(2-oxopyrrolidone-1-yl)pyridin-3-yl]-1H-benzo[d]imidazol-2-yl}oxy}-2-methylbenzoic acid (compound 9) was prepared by the same method as compound 1, with a yield of 58.74%.
[0134] The structural verification data is as follows:
[0135] 1 H NMR (400MHz, DMSO-d6) δ12.79(s,1H),8.47(d,J=2.5Hz,1H),8.41(d,J=8.6Hz,1H),7.94(dd,J=8.7,2.5Hz,1H),7.84(d,J=2.7Hz,1H),7.62(s,1H) ,7.55(dd,J=8.3,2.7Hz,1H),7.46(d,J=3.4Hz,1H),7.44(s,1H),4.09(t ,J=7.1Hz,2H),2.65(t,J=8.0Hz,2H),2.59(s,3H),2.12(p,J=7.7Hz,2H).
[0136] 13 C NMR(100MHz,DMSO-d6)δ175.18,168.29,151.48,151.00,139.35(2C),136.83,133.39(2C),132.20, 131.39,129.87,129.13,124.85,124.23(2C),122.23(2C),113.16(2C),47.45,33.52,21.07,17.68.
[0137] HRMS (ESI) - )m / z calcd for C 24 H 19ClN4O4[MH] - ,461.1022;found,461.0932.
[0138] Example 105: Synthesis of 6-chloro-5-[3-chloro-4-(2-oxopyrrolidone-1-yl)phenyl]-1H-benzo[d]imidazol-2-yl}oxy}-2-methylbenzoic acid (compound 10)
[0139]
[0140] Using the method in step 1 of Example 1, 3-chloro-4-(2-oxo-1-pyrrolidinyl)phenylboronic acid pinacol ester was prepared from 4-bromo-2-chloroaniline as the starting material.
[0141] Compound 5-{{6-chloro-5-[3-chloro-4-(2-oxopyrrolidone-1-yl)phenyl]-1H-benzo[d]imidazol-2-yl}oxy}-2-methylbenzoic acid (compound 10) was prepared by the same method as compound 1, with a yield of 65.61%.
[0142] The structural verification data is as follows:
[0143] 1 H NMR(400MHz,DMSO-d6)δ7.73(s,1H),7.64(s,1H),7.61(s,1H),7.51(s,2H),7.44(s,1H),7.41(s ,1H),7.37(s,1H),3.79(t,J=6.9Hz,2H),2.55–2.54(m,3H),2.49(s,2H),2.21(t,J=7.4Hz,2H).
[0144] 13 C NMR(100MHz,DMSO-d6)δ174.38,168.78,158.38,151.51,139.13,136.27,135.65,133.72,133.11 ,133.09,130.34(3C),124.58,123.54(2C),121.96(2C),119.31(3C),48.52,32.82,21.02,17.93.
[0145] HRMS (ESI) - )m / z calcd for C 25 H 19 Cl2N3O4[MH] - ,494.0680; found,494.0608.
[0146] Example 115 Synthesis of 6-chloro-5-[4-(2-oxopiridine-1-yl)phenyl]-1H-benzo[d]imidazol-2-yl}oxy}-2-methylbenzoic acid (compound 11)
[0147]
[0148] Using p-bromoaniline as the starting material and 5-chlorobutyryl chloride as the starting reaction, 4-(2-oxo-1-piperidinyl)phenylboronic acid pinacol ester was prepared by the method in step 1 of Example 1.
[0149] The compound 5-{{6-chloro-5-[4-(2-oxopiridine-1-yl)phenyl]-1H-benzo[d]imidazol-2-yl}oxy}-2-methylbenzoic acid (11) was prepared by the same method as that used to prepare compound A1, with a yield of 67.11%.
[0150] 1 H NMR (400MHz, DMSO-d6) δ12.74(s,1H),7.83(d,J=2.7Hz,1H),7.59(s,1H),7.55(dd,J=8.3,2.6Hz,1H),7.49(s, 1H),7.47(s,2H),7.40(s,2H),7.38(s,1H),3.70(t,J=5.6Hz,2H),2.59(s,3H),2.46(t,2H),1.95–1.87(m,4H).
[0151] 13 C NMR(100MHz,DMSO-d6)δ169.42,168.27,162.80,158.36,151.51,143.15,137.79,136.83,133.40(2C),133. 09,132.14,130.41(2C),126.19(2C),124.56,124.27(2C),122.24(2C),51.25,33.10,23.50,21.36,21.07.
[0152] HRMS (ESI) - )m / z calcd for C 26 H 22 ClN3O4[MH] - ,474.1226;found,474.1333.
[0153] Activity Assay Example 1: AMPK (α1 / β1 / γ1) Kinase Agonistancy Test
[0154] 1) Dilute the compound to 100 times the final concentration using 100% dimethyl sulfoxide. Transfer 40 μL of the compound dilution to a 384-well echo plate.
[0155] 2) Transfer 200 nL per well from the 384-well echo plate to the 384-well analysis plate via echo.
[0156] 3) Except for the control wells without enzyme, add 10 μC of kinase solution to each well.
[0157] 4) Add substrate and ATP to 1x kinase base buffer to prepare 2x substrate solution.
[0158] 5) Add 10 μL of 2x substrate solution to each well to start the reaction.
[0159] 6) RT incubation reaction.
[0160] 7) Prepare a detection solution with a final concentration of 2 times in antibody dilution buffer.
[0161] 8) Add 20 μL of detection solution to each well, stop the reaction, and incubate at room temperature for 60 minutes.
[0162] 9) Copy the Lance signal ratio from the Envision program (665nm / 615nm).
[0163] 10) Convert the ratio value to a percentage activation value.
[0164] a. Activation percentage = (sample Lance signal ratio - max) / (max - min) * 100.
[0165] b. “min” indicates the proportion controlled without enzymes, and “max” indicates the proportion controlled by DMSO.
[0166] 11) The data is represented in MS Excel, and the curves are fitted using the XLFit Excel add-in version 5.4.0.8. 50 The calculation formula is: Y = Bottom + (Top - Bottom) / (1 + (IC) 50 / X)^HillSlope)
[0167]
[0168]
[0169] The results of kinase assays show that the compounds of this invention have very good AMPK agonist activity, reaching nanomolar levels, and have great development potential.
Claims
1. A compound or a pharmaceutically acceptable salt thereof, characterized in that, Selected from the following compounds: 。 2. The use of the compound of claim 1 or a pharmaceutically acceptable salt thereof in the preparation of an AMPK agonist.
3. A pharmaceutical composition comprising the compound of claim 1 or a pharmaceutically acceptable salt thereof and a pharmaceutically acceptable carrier.
4. The use of the pharmaceutical composition of claim 3 in the preparation of AMPK agonists.