2-arylsulfanylmethyl-6-bromoindole compounds, synthesis and use thereof

Abstract

The present application provides a 2-arylthiomethyl-6-bromoindole compound based on the structure of arbidol, and synthesis and application thereof, and relates to the technical fields of chemistry and pharmacy. Experiments prove that the 2-arylthiomethyl-6-bromoindole compound has different antiviral activity from the hemagglutinin inhibitor arbidol, can be used as a protease inhibitor, and has inhibitory activity on 2019-nCoVM pro / 3CL pro protease, providing a new choice for anti-novel coronavirus.

Description

Technical Field

[0001] This invention relates to the fields of chemical and pharmaceutical technology, and in particular to a 2-arylthiomethyl-6-bromoindole compound and its synthesis and application. Background Technology

[0002] COVID-19, or pneumonia caused by the novel coronavirus, is caused by infection with the novel coronavirus (2019-nCoV). The novel coronavirus is a single-stranded positive-sense RNA virus belonging to the genus B of the subfamily Orthocoronavirinae in the family Coronaviridae of the order Nidovirales. It contains major structural genes such as ORF1a / b, N, and E. The novel coronavirus was discovered in December 2019 and caused the global COVID-19 pandemic. The novel coronavirus spreads rapidly, infects a wide range of people, and is difficult to control. Clinical manifestations include fever and fatigue; respiratory symptoms are mainly dry cough, and in severe cases, difficulty breathing may occur. A small number of patients become critically ill or even die.

[0003] The novel coronavirus (2019-nCoV) shares high homology with SARS-CoV. After infecting a host cell, with the help of the host cell, its genetic material RNA's ORF1a / b first translates and expresses two polyprotein precursors (pp1a and pp1ab). These polyprotein precursors undergo intramolecular cleavage under the action of the main protease (Mpro) and papain-like proteases, producing multiple non-structural proteins. These non-structural proteins participate in the production of viral subgenetic RNA and four structural proteins (envelope / E protein, membrane / M protein, spike / S protein, and nucleocapsid / N protein), thereby completing the proliferation and release of progeny viruses. Because the Mpro protease plays a crucial role in the viral life cycle and has no homologous protein in the human body, the Mpro main protease is an ideal target for antiviral drug development.

[0004] Arbidol is a broad-spectrum and safe antiviral drug. Experiments have shown that its efficacy and safety in treating COVID-19 are higher than those of antiviral drugs such as lopinavir. It can effectively improve the 2019-nCoV nucleic acid negative conversion rate, and early oral administration of arbidol in COVID-19 patients can alleviate symptoms, shorten the course of the disease, and reduce complications. However, arbidol is a hemagglutinin inhibitor, and it can only inhibit viral DNA and RNA synthesis by specifically inhibiting the contact, adhesion, and fusion of the viral lipid envelope with the host cell membrane, thus blocking the viral gene from entering the cell nucleus. It has virtually no inhibitory activity against the Mpro main protease.

[0005] Therefore, developing a new compound that inhibits the main Mpro protease is of great significance for enriching the drug pool against the novel coronavirus.

[0006] Summary of the Invention

[0007] One objective of this invention is to provide a 2-arylthiomethyl-6-bromoindole compound, and another objective is to provide its preparation method and application.

[0008] This invention is implemented as follows:

[0009] 2-Arylthiomethyl-6-bromoindole compounds or their pharmaceutically acceptable salts, having the following structures:

[0010]

[0011] The synthetic methods for the above-mentioned 2-arylthiomethyl-6-bromoindole compounds are as follows:

[0012] The synthetic route is as follows:

[0013]

[0014] Wherein, R1 is methylimidazole and R2 is n-propyl, benzyl, or thiophene; or R1 is methyl and R2 is n-hexyl or n-butyl.

[0015] Specifically, the preparation of the above-mentioned 2-arylthiomethyl-6-bromoindole compounds includes the following steps:

[0016] (1) Sodium hydroxide, HS-R1, and compound A were added sequentially to methanol and stirred to react. After the reaction was complete, the pH was adjusted to neutral with acetic acid, allowed to stand, filtered, the filter cake was dried, and the filtrate was extracted to obtain the crude product. The crude product was purified by column chromatography (dichloromethane:methanol = 120:1), and then recrystallized with EA to obtain pure compound B, which was a brownish-black solid. The molar ratio of sodium hydroxide, HS-R1, and compound A was 2.5:1.5:1.

[0017] (2) Compound B was dissolved in 1,4-dioxane, and 10% NaOH solution was added. Dimethyl sulfate was added dropwise at 25°C while stirring. After the reaction was complete, the solvent was evaporated, water was added, and the mixture was filtered. The filter cake was purified by column chromatography (developing solvent: dichloromethane:methanol = 80:1) to obtain pure compound C, which was a yellowish-white solid. The molar ratio of compound B, 10% NaOH, and dimethyl sulfate was 1:14:9.

[0018] (3) Compound C, NaOH, and water were dissolved in ethanol and refluxed at 80°C for 6 hours. After the reaction was completed, the mixture was cooled to room temperature, concentrated hydrochloric acid was added, and the solvent was evaporated to dryness to obtain the crude product. The crude product was purified by column chromatography (dichloromethane:methanol = 40:1) to obtain compound D, which was a white solid. The molar ratio of compound C, NaOH, water, and ethanol was 1:14:0.6:33.

[0019] (4) Under ice bath conditions, compound D was dissolved in dichloromethane and N,N-dimethylformamide (DMF), followed by the addition of 1-ethyl-3-(3-dimethylpropylamine)carbodiimide (EDCI) and 1-hydroxybenzotriazole (HoBt). After stirring until homogeneous, triethanolamine (TEA) and NH2-R2 were added. The reaction was carried out at room temperature for 8 hours. After the dichloromethane was evaporated, ice water was added, and the solid was allowed to precipitate. The solid was then filtered. The filter cake was purified by thin-layer chromatography (dichloromethane:methanol = 80:1) to obtain pure compound IV, which was a yellowish-white solid. Specifically, when R1 was methylimidazole and R2 was n-propyl, compound I was obtained; when R1 was methylimidazole and R2 was benzyl, compound II was obtained; when R1 was methylimidazole and R2 was thiophene, compound III was obtained; when R1 was methyl and R2 was n-hexyl, compound IV was obtained; and when R1 was methyl and R2 was n-butyl, compound V was obtained. The molar ratio of compounds D, EDCI, HoBt, TEA to NH2-R2 is 1:3.5:3.5:4:2.5.

[0020] This invention, based on the Arbidol structural core, provides a 2-arylthiomethyl-6-bromoindole compound that exhibits antiviral activity different from the hemagglutinin inhibitor Arbidol. Experiments demonstrate that the 2-arylthiomethyl-6-bromoindole compound of this invention can be used as a protease inhibitor against 2019-nCoV M. pro / 3CL pro The protease has inhibitory activity, providing a new option for combating the novel coronavirus.

[0021] The preparation method provided by this invention has a high yield, low cost, and is easy to operate. Detailed Implementation

[0022] The present invention will be further described below with reference to the embodiments. The embodiments described below are intended to illustrate the present invention and do not limit the scope of protection of the present invention in any way.

[0023] The processes and methods not described in detail in the following examples are conventional methods known in the art. All reagents used in the examples are analytically pure or chemically pure, and can be commercially available or prepared by methods known to those skilled in the art.

[0024] Example 1

[0025] Preparation of 1-methyl-N-n-propyl-2-(-methylimidazolium thiomethyl)-5-methoxy-6-bromoindole-3-carboxamide (I):

[0026]

[0027] At 25°C, 0.23 g (2.5 mol) of sodium hydroxide was first added to methanol, followed by 0.44 g (1.7 mol) of 2-mercapto-1-methylimidazole, and the mixture was stirred for 2 h. Finally, 1 g (1 mol) of A1 was added, and the mixture was stirred for 5 h. After the reaction was complete, the reaction solution was neutralized with acetic acid (pH = 6), precipitating a large amount of brownish-black solid. The solid was allowed to stand, filtered, the filter cake was dried, and the filtrate was extracted to obtain the crude product. The crude product was purified by column chromatography (dichloromethane:methanol = 120:1), and then recrystallized from EA to obtain compound B1, a brownish-black solid, with a yield of 80%. 1 HNMR(600MHz,DMSO)δ9.85(s,1H),7.72(s,1H),7.56(s,1H),7.26(s,1H),6.95(s,1H) ,5.75(s,3H),4.63(s,2H),4.20(q,J=7.1Hz,2H),3.51(s,3H),1.33(t,J=7.1Hz,3H); 13 C10 NMR (151MHz, DMSO) δ 164.03, 149.26, 143.80, 137.80, 131.34, 129.01, 126.01, 124.18, 114.27, 106.42, 106.18, 102.68, 59.12, 32.84, 29.63, 28.84, 14.29. Molecular formula C10 19 H 20 BrNO3S, molecular weight 421.0347.

[0028] 0.89 g (1 mol) of compound B1 was dissolved in 1,4-dioxane, and 0.9 g (10.65 mol) of 10% NaOH solution was added. Then, 0.7 mL (3.5 mol) of dimethyl sulfate was added dropwise at 25 °C, and the mixture was stirred for 5 h. After the reaction was complete, the solvent was evaporated to dryness, and the mixture was filtered with water (evolving solvent: dichloromethane:methanol = 80:1) to give pure compound C1 as a yellowish-white solid, yield: 75%. 1 H NMR (600MHz, CDCl3) δ7.68,7.48,7.09,6.91,4.73,4.38,4.37,4.36,4.35,3.96,3.95,3.53,3.31,1.45,1.44,1.42; 13C10 NMR (151MHz, CDCl3) δ 164.94, 151.92, 143.23, 139.59, 132.09, 129.86, 126.44, 123.17, 114.18, 108.30, 104.55, 103.69, 59.84, 56.53, 33.26, 29.84, 29.68, 14.48. Molecular formula C10 18 H 20 BrN3O3S, molecular weight 437.0409.

[0029] 0.77 g (1 mol) of product C1, 1 g (14 mol) of NaOH, and 0.6 mL of water were dissolved in ethanol and refluxed at 80 °C. The reaction was monitored by TLC for approximately 6 h. After the reaction was complete, the mixture was cooled to room temperature, concentrated hydrochloric acid was added, and the solvent was evaporated to dryness to obtain the crude product. The crude product was purified by column chromatography (dichloromethane:methanol = 40:1) to obtain product D1, which was a white solid. The yield was 70%.

[0030] 1 g (1 mol) of product D1 was dissolved in dichloromethane and DMF in an ice bath, followed by the addition of 1.4 g (3 mol) of EDCI and 1 g (3 mol) of HoBt. After stirring for 5 min, 1.3 mL (4 mol) of TEA and 0.5 mL (2.5 mol) of NH2-R2 were added. The reaction was carried out at room temperature for 8 h. After evaporating the dichloromethane, ice water was added, resulting in the precipitation of a solid. The solid was filtered. The filter cake was purified by thin-layer chromatography (dichloromethane:methanol = 80:1) to obtain pure compound I as a yellowish-white solid. Yield: 60%. 1 H NMR (600MHz, CDCl3) δ7.83 (s, 1H), 7.46 (s, 1H), 7.44 (s, 1H), 7.00 (d, J = 0.8Hz, 1H), 6.88 (s, 1H), 4.63 (s, 2H), 3.92(s,3H),3.68(s,3H),3.47(s,3H),3.45-3.42(m,2H),1.71(dq,J=14.6,7.3Hz,4H),1.03(t,J=7.4Hz,3H); 13 C10 NMR (151MHz, CDCl3) δ 164.18, 150.28, 138.44, 135.06, 131.53, 128.29, 124.90, 122.01, 112.90, 110.31, 107.39, 101.88, 55.76, 40.43, 32.28, 29.26, 27.62, 22.04, 10.74. Molecular formula C10 19 H 23 BrN4O2S, molecular weight 450.0725.

[0031] Example 2

[0032] Preparation of 1-methyl-N-benzylamino-2-(-methylimidazolium thiomethyl)-5-methoxy-6-bromoindole-3-carboxamide (II)

[0033]

[0034] The procedure was the same as in Example 1, and the reaction was carried out for 8 hours to obtain pure compound II, with a yield of 55%. 1 H NMR (600MHz, CDCl3) δ7.88(s,1H),7.45(s,1H),7.29(s,1H),7.29(s,1H),7.28(s,3H),7.28(s,1H),7.21(dd,J=5.5,2.2Hz,1H),6 .90(s,1H),6.85(s,1H),4.47(s,2H),3.83(s,3H),3.78(dd,J=12.4,6.8Hz,3H),3.63(s,3H),3.43(s,3H),3.03(t,J=6.9Hz,3H); 13 C10 NMR (151MHz, CDCl3) δ 164.06, 150.25, 138.46, 138.25, 135.16, 131.46, 128.05, 127.60, 126.95, 126.18, 124.97, 121.82, 112.86, 109.96, 107.33, 101.81, 52.40, 42.56, 32.20, 29.35, 27.38. Molecular formula C10 23 H 23 BrN4O2S, molecular weight 498.0725.

[0035] Example 3

[0036] Preparation of 1-methyl-N-2-thiophenemethylamino-2-(-methylimidazolium thiomethyl)-5-methoxy-6-bromoindole-3-carboxamide (III)

[0037]

[0038] The procedure was the same as in Example 1, and the reaction was carried out for 8 hours to obtain pure compound III, with a yield of 68%. 1H NMR (600MHz, CDCl3) δ8.76(s,1H),7.47(s,1H),7.46(s,1H),7.22(d,J=5.1Hz,1H),7.08(d,J=3.3Hz,1H),6.97-6 .96(m,1H),6.82(s,1H),6.79(s,1H),4.86(d,J=5.4Hz,3H),4.64(s,2H),3.90(s,4H),3.68(s,4H),3.44(s,3H); 13 C NMR (151MHz, CDCl3) δ 163.93, 150.24, 141.17, 138.42, 135.26, 131.40, 128.02, 125.80, 124.93, 124.75, 123.78, 121.83, 112.83, 109.74, 107.30, 101.75, 55.67, 37.33, 32.23, 29.36, 27.26. Molecular formula C21H21BrN4O2S2, molecular weight 504.0289.

[0039] Example 4

[0040] Preparation of 1-methyl-N-hexyl-2-(-methylthiomethyl)-5-methoxy-6-bromoindole-3-carboxamide (IV)

[0041]

[0042] The procedure was the same as in Example 1, and the reaction was carried out for 8 hours to obtain pure compound IV, with a yield of 70%. 1 H NMR (600MHz, CDCl3) δ7.49(s,1H),7.36(s,1H),6.32(s,1H),4.14(s,1H),3.92(s,3H),3.70(s,2H),3.49(dd,J=12. 8,6.9Hz,3H),2.14(s,2H),1.67-1.63(m,3H),1.33(d,J=3.5Hz,2H),1.24(d,J=14.2Hz,3H),0.90(t,J=7.0Hz,4H); 13 C10 NMR (151MHz, CDCl3) δ 164.45, 150.31, 137.24, 131.23, 124.15, 113.11, 109.12, 106.99, 101.13 52.40, 38.62, 30.56, 28.93, 28.82, 26.44, 25.89, 21.63, 14.14, 13.02. Molecular formula C10 19 H 27BrN2O2S, molecular weight 426.0977.

[0043] Example 5

[0044] Preparation of 1-methyl-N-n-butyl-2-(-methylthiomethyl)-5-methoxy-6-bromoindole-3-carboxamide (V)

[0045]

[0046] The procedure was the same as in Example 4, and the reaction was carried out for 8 hours to obtain pure compound V, with a yield of 50%. 1 H NMR (600MHz, CDCl3) δ7.48(s,1H),7.36(s,1H),6.33(s,1H),4.13(s,2H),3.92(s,4H),3.69(s,3H),3.4 9(d,J=6.0Hz,2H),2.14(s,3H),1.66-1.62(m,3H),1.47(dd,J=15.0,7.5Hz,3H),0.98(t,J=7.4Hz,4H); 13 C10 NMR (151MHz, CDCl3) δ 164.45, 150.27, 137.17, 131.19, 124.15, 113.08, 109.10, 106.94, 101.09, 52.41, 38.29, 30.89, 28.92, 26.45, 19.33, 14.14, 12.80. Molecular formula C10 17 H 23 BrN2O2S, molecular weight 398.0664.

[0047] Example 6

[0048] The 2-arylthiomethyl-6-bromoindole-3-carboxamide compounds obtained in Examples 1-5 of this invention are effective against 2019-nCoV M. pro / 3C pro Inhibitory activity of proteases.

[0049] Sample: 2-arylthiomethyl-6-bromoindole-3-carboxamide compounds obtained in Examples 1-5 of this invention.

[0050] Reagent: Novel Coronavirus M pro / 3CL pro Inhibitor screening kit (2019-nCoVM) pro / 3C proThe Inhibitor Screening Kit includes Assay Buffer 125 mL, Substrate 1 mL, Ebselen (10 mM) (100 μL), and 2019-nCoV M. pro / 3C pro Store (500 μL) at -20°C. Substrate should be stored away from light. BeyoGold TM All black 96-well cell culture plates (FCP966) were provided by Beyotime Biotechnology. DMSO (Tianjin Kemeio Chemical Reagent Co., Ltd.)

[0051] Experimental procedure:

[0052] (1) Sample preparation

[0053] Take an appropriate amount of the samples obtained in Examples 1-5 and Arbidol standards to be tested, and prepare a solution with a concentration of 2 μmol / L using DMSO; prepare a solution of Ebselen, the positive control inhibitor provided in the kit, with a concentration of 2 μmol / L using DMSO.

[0054] (2) Experimental Operation

[0055] Novel coronavirus (2019-nCoV) M pro / 3CL pro The inhibitor screening kit uses fluorescence resonance energy transfer (FRET) to detect the inhibitory activity of the test samples. First, 92 μL of Assay Buffer and 1 μL of 2019-nCoV M are pipetted according to the sample quantity. pro / 3CL pro Prepare an appropriate amount of AssayReagent. Set up each group using a 96-well blackboard: blank control (93 μL Assay Buffer + 5 μL DMSO), 100% enzyme activity control (93 μL Assay Reagent + 5 μL LDMSO), positive inhibitor control (93 μL Assay Reagent + 5 μL LEbselen solution), and sample (93 μL Assay Reagent + 5 μL sample to be tested). Then, quickly add 2 μL of Substrate to each well and mix well. For more reliable results, each sample should be tested in at least two replicates. After incubation at 37°C in the dark for 5 minutes, the signal will stabilize. Perform fluorescence measurement using a multifunctional enzyme label. The excitation wavelength is 360 nm, and the emission wavelength is 528 nm.

[0056] (3) Calculation

[0057] a. Calculate the average fluorescence value for each sample well and blank control well, and record it as RFU blank control, RFU 100% enzyme activity control, RFU positive control, and RFU sample. (RFU, Relative Fluorescence Unit).

[0058] b. Calculate the inhibition percentage for each sample.

[0059] The calculation formula is as follows:

[0060] Inhibition rate (%) = (RFU 100% enzyme activity control - RFU sample) / (RFU 100% enzyme activity control - RFU blank control) × 100%, and the results are shown in Table 1 below.

[0061] Table 1

[0062]

[0063] The above results indicate that compounds I, II, III, IV, V, and VI of this invention are effective against 2019-nCoV M. pro / 3CL pro The protease has inhibitory activity, while Arbidol is effective against 2019-nCoV M. pro / 3CL pro The protease has virtually no inhibitory activity.

Claims

1. A bromoindole compound or a pharmaceutically acceptable salt thereof, having the following structure: ###0001### 、 、 、 、 。 2. The synthesis method of the bromoindole compound of claim 1, characterized in that, the synthesis route is: ###0002### wherein, R1 is methyl imidazole, R2 is n-propyl, benzyl or thienylmethyl; or R1 is methyl, R2 is n-hexyl or n-butyl. ； ​ 3. Use of the bromoindole compound or pharmaceutically acceptable salt thereof according to claim 1 for the manufacture of an anti-coronavirus drug, characterized in that, for use in the preparation of a 2019-nCoV M pro / 3CL pro protease potential inhibitor drug.

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