Five-membered heterocyclic indole derivatives

Abstract

This invention discloses a five-membered heterocyclic benzoindole derivative, the structural formula of which is shown in formula (I). Experiments have shown that the five-membered heterocyclic benzoindole derivative of this invention has an inhibitory effect on liver cancer cells.

Description

Technical Field

[0001] This invention belongs to the field of chemical pharmaceuticals, and in particular relates to five-membered heterocyclic indole derivatives and their application in the preparation of antitumor drugs. Background Technology

[0002] Cancer is a global health challenge.

[0003] Since the 20th century, cancer treatment has undergone three revolutions: the emergence of chemotherapy drugs, the development of targeted drugs, and the proposal of immunotherapy.

[0004] In the 1940s, the cytotoxic chemotherapy drugs nitrogen mustard and methotrexate proved effective in treating lymphoma and acute lymphoblastic leukemia, respectively, marking the beginning of chemotherapy for tumors. In the 1950s, cyclophosphamide, 5-fluorouracil, and other drugs were subsequently used to treat malignant tumors. In the 1970s and 80s, with the emergence of more chemotherapy drugs such as cisplatin and doxorubicin, the method of combining multiple anti-tumor drugs became increasingly mature. In the 1990s, natural drugs such as camptothecin and paclitaxel were extracted. Cytotoxic chemotherapy drugs have a rapid onset of action and are the mainstay of tumor treatment, but they lack selectivity and have significant toxic side effects on normal cells that divide and proliferate rapidly in the body. To overcome the toxicity caused by the selectivity of chemotherapy, the second revolution in tumor treatment was the search for highly selective targeted drugs with fewer toxic side effects.

[0005] Rituximab, the world's first molecularly targeted drug, opened the door to precision medicine for cancer. Epidermal growth factor receptor (EGFR) plays a crucial role in the spread and growth of non-small cell lung cancer, and imatinib mesylate, an EGFR tyrosine kinase inhibitor, is effective in treating chronic myeloid leukemia. The development and market launch of tyrosinib (TKI) drugs such as gefitinib and afatinib also demonstrate the rapid development of targeted therapies. However, as the clinical application of targeted drugs increases, their drawbacks are becoming increasingly apparent—slow onset of action and the emergence of drug resistance once the target mutation occurs. Developing dual-target or multi-target drugs is one strategy to address this problem.

[0006] The third revolution in anti-tumor drugs, and arguably the most promising turning point in current cancer treatment, is tumor immunotherapy. In 1958, immunologist Frank Macfrlane Burnet proposed the immune surveillance theory, suggesting that tumor cells appearing in the body under normal circumstances can be recognized and eliminated by the immune system. However, once immune escape occurs, tumor cells can survive at various stages of the anti-tumor immune response by employing different immunosuppressive strategies, preventing the body from killing them. Tumor immunotherapy works by activating the immune system to prevent tumor immune escape, thereby killing the tumor.

[0007] Despite significant advancements in cancer treatment methods over the past few decades, chemotherapy remains a primary treatment for cancer. Based on their mechanisms of action, commonly used chemotherapeutic drugs can be categorized as: antimetabolites, alkylating agents, mitotic spindle inhibitors, topoisomerase inhibitors, etc.

[0008] Heterocyclic compounds are an important source of pharmacologically active compounds, obtained through organic synthesis or isolated from natural products. Because heterocyclic compounds contain one or more heteroatoms such as nitrogen, oxygen, and sulfur in addition to at least one carbon atom in the ring, they are often used as hydrogen bond donors and acceptors. They can effectively bind to biological targets through intermolecular hydrogen bonds. Furthermore, they can modulate the lipid solubility of drug molecules or enhance their water solubility, thereby producing excellent therapeutic properties. Among various heterocyclic compounds, indole has attracted considerable attention due to its potential as a unique scaffold in drug discovery and development.

[0009] Indole is a planar bicyclic molecule. According to Huckel's rule (10 π electrons), it possesses aromatic properties. Due to the delocalization of π electrons, it readily undergoes electrophilic substitution reactions. Furthermore, because the N-H bond is weakly acidic, nucleophilic substitution reactions can also occur on the nitrogen atom under basic conditions. In addition, indole is widely found in some natural products (alkaloids, plant hormones, microbial hormones, etc.). For example, vincristine and vinblastine isolated from periwinkle can be used as microtubule inhibitors (Ki 0.178 and 0.085 mM, respectively) for the treatment of various cancers. Furthermore, a marine indole derivative, eudistomin K, has been reported to inhibit the P-388 tumor cell line (IC50). 50 =0.01ug / mL), it is considered a potential anticancer lead compound.

[0010] In the mid-to-late 19th century, with the advent of the Fischer indole synthesis method and various other indole synthesis methods, indole was regarded as the preferred skeleton for designing anticancer agents.

[0011] Currently, indole compounds have been used in various anti-tumor pathways, such as: acting on apoptosis (Mcl-1 inhibitors), intracellular signal transduction (Pim inhibitors), epigenetic modification (HDAC inhibitors and SIRT inhibitors), cell mitosis (tubulin inhibitors), etc.

[0012] Despite the promising anticancer properties of indole derivatives, the selectivity issue remains unresolved. For example, most HDAC inhibitors consist of an isohydroxamic acid group (ZBG), a cap group, and a linker (aliphatic or aromatic). The indole skeleton is commonly used as the cap group or aromatic spacer in HDAC inhibitor design. To obtain promising isomeric selective HDAC inhibitors and avoid adverse side effects such as fatigue, cardiotoxicity, and hematologic toxicity, effectively optimizing the indole skeleton, ZBG, and linker will be an important future research direction.

[0013] With advancements in drug design methods and screening technologies, more potent and selective indole derivatives can be obtained as anticancer drugs through further modification of natural products or synthetic lead compounds. Furthermore, for drug-resistant cancers, the combination of indole with other active moieties will be an important pathway for developing novel anticancer agents with higher efficacy. Summary of the Invention

[0014] The purpose of this invention is to overcome the shortcomings of the prior art and provide a five-membered heterocyclic indole derivative.

[0015] A second objective of this invention is to provide the application of five-membered heterocyclic indole derivatives in the preparation of antitumor drugs.

[0016] The technical solution of this invention is summarized as follows:

[0017] Five-membered heterocyclic benzoindole derivatives, the structural formula of which is shown in Formula I:

[0018]

[0019] in:

[0020] R1 is H or Br:

[0021] R2 is H,

[0022] The application of the above-mentioned five-membered heterocyclic indole derivatives in the preparation of antitumor drugs.

[0023] Advantages of this invention:

[0024] Experiments have shown that the five-membered heterocyclic indole derivative of this invention has an inhibitory effect on liver cancer cells. Detailed Implementation

[0025] The present invention will be further described below through specific embodiments.

[0026] The following abbreviations are used in this article: MeOH: methanol; Boc: tert-butyloxycarbonyl; THF: tetrahydrofuran; TEA: triethylamine; EA: ethyl acetate;

[0027] DMP: Dess-Martin reagent;

[0028] PE: Petroleum ether; Et2O: Diethyl ether; TFA: Trifluoroacetic acid; DMF: N,N-dimethylformamide; DCM: Dichloromethane;

[0029] HATU: 2-(7-azabenzotriazole)-N,N,N',N'-tetramethylurea hexafluorophosphate;

[0030] DIPEA: N,N-diisopropylethylamine;

[0031] Additionally, "L" represents naturally occurring amino acids. FBS: Fetal bovine serum; PBS solution: Phosphate buffer; PBST: Phosphate buffer plus Tween-20 (phosphate buffer at pH 7.4 containing 0.05% Tween-20);

[0032] ESMS: Electrospray ionization mass spectrometry; MS: Mass spectrometry; HPLC: High performance liquid chromatography.

[0033]

[0034] R1:H

[0035]

[0036] R1:Br

[0037]

[0038] Example 1

[0039] Preparation of 2-chloro-N-(4-morpholinylphenyl)acetamide (compound 1):

[0040]

[0041] 4-morpholinoaniline (3.00 g, 16.83 mmol) was added to a 100 mL dry round-bottom flask equipped with a stirrer. 40 mL of 1,4-dioxane was added as a solvent, and the mixture was stirred at room temperature. Triethylamine (TEA 0.18 g, 0.16 mmol) was added dropwise, followed by the slow addition of chloroacetyl chloride (1.92 g, 17.00 mmol) to induce a nucleophilic substitution reaction. The reaction was carried out at room temperature for 1 h. After the reaction was completed as monitored by TLC, 1,4-dioxane was removed by vacuum distillation. The reaction mixture was dissolved in dichloromethane, diluted with 100 mL of saturated sodium chloride solution, and extracted with dichloromethane (100 mL × 3). The organic phases were combined and dried over anhydrous sodium sulfate. Dichloromethane was removed by vacuum distillation. The crude product was purified by column chromatography (dichloromethane:methanol = 100:1 v / v) to give a brownish-yellow solid (compound 1) in approximately 75% yield. 1 H NMR (400MHz, DMSO-d6) δ10.09(s,1H),7.46(d,J=7.1Hz,2H),6.91(d,J=7.2Hz,2H),4.20(s,2H),3.73(s,4H),3.05(s,4H). 13 C NMR (100MHz, DMSO-d6) δ164.47,148.12,131.07,120.99,115.87,66.57,49.24,44.02.

[0042] Example 2

[0043] Preparation of compound (E)-2-(4-morpholinylphenyl)imino)thiazolidin-4-one (compound 2)

[0044]

[0045] Compound 1 (3.00 g, 11.80 mmol) and ammonium thiocyanate (1.80 g, 23.68 mmol) were added to a 100 mL dry round-bottom flask equipped with a stirrer. 40 mL of anhydrous ethanol (EtOH) was added as a solvent, and the mixture was heated to 80 °C and refluxed for 5 h. After the reaction was completed as monitored by TLC, the ethanol was removed by vacuum distillation. The crude product was then purified by column chromatography (dichloromethane:methanol = 100:1 v / v) to give a white solid (compound 2) in approximately 70% yield. 1H NMR(400MHz,DMSO-d6)δ11.23(d,J=170.6Hz,1H),7.55(d,J=8.9Hz,1H),6.99-6 .93(m,3H),3.98(s,1H),3.92(s,1H),3.75-3.72(m,4H),3.11-3.07(m,4H).13C NMR (100MHz, DMSO-d6) δ188.50,177.58,149.36,148.70,131.41,123.92,121.78,115.72,66.55,48.99,36.57.

[0046] Example 3

[0047] Preparation of compound (Z)-3-(4-fluorobenzyl)-2-(4-morpholinylphenyl)imino)thiazolidin-4-one (compound 3):

[0048]

[0049] Compound 2 (1.50 g, 5.42 mmol) and anhydrous potassium carbonate (0.75 g, 5.43 mmol) were added to a 100 mL dry round-bottom flask equipped with a stirrer. 40 mL of ultradry N,N-dimethylformamide (DMF) solvent was then added to the reaction mixture, and the mixture was stirred at room temperature for 30 min. 4-Fluorobenzyl bromide (1.23 g, 6.51 mmol) was added dropwise. After the addition was complete, the temperature was raised to 80 °C (or 75 °C or 85 °C), and the mixture was stirred for 12 h to allow a nucleophilic substitution reaction to occur. After the reaction was completed as monitored by TLC, the mixture was cooled to room temperature. The reaction mixture was diluted with 100 mL of saturated sodium chloride solution and extracted with ethyl acetate (100 mL × 3). The organic phases were combined and dried over anhydrous sodium sulfate. Ethyl acetate was removed by vacuum distillation. The crude product was purified by column chromatography (petroleum ether:ethyl acetate = 5:1 v / v) to give compound 3 as a yellow solid, with a yield of approximately 50%. 1H NMR (400MHz, DMSO-d6) δ7.43(dd,J=8.4,5.7Hz,2H),7.18(t,J=8.8Hz,2H),6.93(d,J=8.8Hz,2 H),6.83(d,J=8.8Hz,2H),4.89(s,2H),4.08(s,2H),3.75-3.71(m,4H),3.08-3.05(m,4H).13C NMR(100MHz,Chloroform-d)δ171.61,163.71,161.26,153.10,148.55,140.43,131 .94,131.19,131.11,121.91,116.52,115.40,115.19,66.95,49.70,45.53,32.70.

[0050] Example 4

[0051] Preparation of compound (2Z,5Z)-5-(1H-indol-3-yl)methylene)-3-(4-fluorobenzyl)-2-(4-morpholinylphenyl)imino)thiazolidin-4-one (I-1):

[0052]

[0053] Indole-3-carboxaldehyde (compound 4, 1.83 mmol) and 40 mL of anhydrous ethanol were added to a 100 mL dry round-bottom flask equipped with a stirrer. Then, a catalytic amount of piperidine (14.88 mg, 0.17 mmol) was added dropwise. The mixture was stirred at room temperature for 5 minutes, and then compound 3 (0.70 g, 1.82 mmol) was added to the reaction system. The mixture was stirred, and the temperature was raised to 80 °C and refluxed overnight to induce the Knoevenagel reaction. After cooling to room temperature, a large amount of yellow solid precipitated. The solid was filtered, washed twice with anhydrous ethanol, and dried to obtain yellow solid powder I-1, with a yield of approximately 60%. 1H NMR (400MHz, DMSO-d6) δ11.97(s,1H),8.04(s,1H),7.85(d,J=7.8Hz,1H),7.63(s,1H),7.51-7.4 6(m,3H),7.25-7.17(m,4H),6.96(s,4H),5.06(s,2H),3.76-3.73(m,4H),3.12-3.09(m,4H).13C NMR(100MHz,DMSO-d6)δ166.29,163.23,160.81,148.92,139.97,136.61,133.21,130.54,130.45,128.73,127.23 ,123.43,123.27,122.39,121.37,118.73,116.34,115.87,115.66,114.43,112.86,110.99,66.62,49.05,45.37.

[0054] Example 5

[0055] Preparation of tert-butyl carbamate (3-1): (3-(3-(Z)-(Z)-3-(4-fluorobenzyl)-2-(4-morpholinylphenyl)imino)-4-oxothiazolidinylbutan-5-ylidene)methyl)-1H-indole-1-propyl)carbamate (3-1)

[0056]

[0057] Intermediate I-1 (0.93 mmol) and sodium hydride (0.03 g, 1.25 mmol) were added to a 100 mL dry round-bottom flask equipped with a stirrer. 30 mL of ultra-dry DMF solvent was then added to the system, and the mixture was stirred at room temperature for 5 minutes. N-Boc-3-aminopropyl bromide (0.27 g, 1.13 mmol) was dissolved in 10 mL of ultra-dry DMF solvent and added dropwise to the reaction mixture. After the addition was complete, the temperature was raised to 80 °C, and the reaction was allowed to proceed for 4 h. The reaction was monitored by TLC. After the reaction was complete, the mixture was cooled to room temperature, diluted with 100 mL of saturated sodium chloride solution, and extracted with ethyl acetate (100 mL × 3). The organic phases were combined and dried over anhydrous sodium sulfate. Ethyl acetate was removed by vacuum distillation to obtain a yellow solid, 3-1.

[0058] Example 6

[0059] Preparation of (2Z,5Z)-5-(1-(3-aminopropyl)-1H-indole-3-methylene)-3-(4-fluorobenzyl)-2-(4-morpholinylphenyl)imino)thiazolidin-4-one (I-2)

[0060]

[0061] Add 3-1 to a 100 mL dry round-bottom flask equipped with a stir bar, add 30 mL of ultra-dry DCM (dichloromethane) solvent, stir, and then slowly add 15 mL of TFA (trifluoroacetic acid). Stir at room temperature for 2 hours, and monitor the reaction by TLC. After the reaction is complete, distill under reduced pressure to completely remove trifluoroacetic acid and dichloromethane. The crude product obtained is purified by column chromatography (dichloromethane:methanol = 20:1 v / v) to give yellow solid I-2, with a yield of approximately 75%. 1 H NMR (400MHz, DMSO-d6) δ8.01 (s, 1H), 7.89 (d, J = 7.9Hz, 1H), 7.83 (s, 2H), 7.73 (s,1H),7.64(d,J=8.2Hz,1H),7.47(d,J=2.8Hz,2H),7.31(t,J=7.4Hz,1H),7 .24-7.18(m,3H),6.96(d,J=9.9Hz,4H),5.06(s,2H),4.39(t,J=6.9Hz,2H),3 .76-3.74(m,4H),3.12-3.09(m,4H),2.75(t,J=7.5Hz,2H),2.05-2.00(m,2H). 13 C NMR(100MHz,DMSO-d6)δ166.33,163.24,160.82,148.72,148.65,139.93,136.33,133.20,131.06,130.56,130.48,127.87,1 23.68,122.64,122.29,121.80,116.36,115.89,115.67,114.76,111.29,110.42,66.63,49.03,45.47,43.82,36.94,28.32.

[0062] Example 7

[0063] Preparation of (3-(3-((Z)-(Z)-3-(4-fluorobenzyl)-2-(4-morpholinylphenyl)imino)-4-oxothiazolidin-5-imino)methyl)-1H-indole-1-propyl)amino)-4-methylthio-1-oxobutane-2-carbamate tert-butyl ester (4-1)

[0064]

[0065] Boc-L-methionine (0.20 mmol) and HATU (75.00 mg, 0.20 mmol) were added to a 50 mL dry round-bottom flask equipped with a stir bar. 10 mL of ultra-dry DMF solvent was added, and the mixture was stirred at room temperature for 5 minutes. DIPEA (N,N-diisopropylethylamine, 41.28 mg, 0.32 mmol) was then added dropwise, and the mixture was stirred at room temperature for 30 minutes. Intermediate I-2 (0.16 mmol) was dissolved in 5 mL of ultra-dry DMF solvent and added dropwise to the mixture. The mixture was stirred overnight at room temperature. The reaction was monitored by TLC. After the reaction was complete, the reaction mixture was diluted with 50 mL of saturated sodium chloride solution and extracted with ethyl acetate (50 mL × 3). The organic phases were combined and dried over anhydrous sodium sulfate. Ethyl acetate was removed by vacuum distillation. The crude product was purified by column chromatography (petroleum ether: ethyl acetate = 2:1 v / v) to give a yellow solid powder 4-1, with a yield of approximately 65%. 1 H NMR(400MHz,DMSO-d6)δ8.02(s,1H),7.96(s,1H),7.87(d,J=7.7Hz,1H),7.75(s,1 H),7.60(d,J=7.9Hz,1H),7.50-7.46(m,2H),7.29(d,J=6.7Hz,1H),7.21(d,J=9.5 Hz,3H),6.97(s,5H),5.07(s,2H),4.29(s,2H),3.99-3.93(m,1H),3.75(s,4H),3. 11(s,6H),2.44(d,J=7.1Hz,2H),2.02(s,3H),1.90-1.79(m,4H),1.36(s,9H).13C NMR(100MHz,DMSO-d6)δ172.33,166.32,163.22,160.80,155.94,148.71,148 .58,139.83,136.31,133.21,131.27,130.54,130.45,127.87,123.52,122.7 1,122.37,121.65,119.04,116.34,115.85,115.64,114.52,111.26,110.23,78.60,66.63,54.26,49.05,45.42,44.45,36.49,31.97,30.28,28.62,15.07.

[0066] Example 8

[0067] Preparation of (R)-2-amino-N-(3-(3-(Z)-3-(4-fluorobenzyl)-2-(4-morpholinylphenyl)imino)-4-oxothiazolidin-5-ylidene)methyl)-1H-indol-1-yl)propyl)-4-methylthiobutyramide (I-3)

[0068]

[0069] Intermediate 4-1 (50 mg) was added to a 50 mL dry round-bottom flask equipped with a stir bar, 10 mL of ultra-dry DCM solvent was added, and the mixture was stirred. Then, 5 mL of TFA was slowly added dropwise. The mixture was stirred at room temperature for 1 hour, and the reaction was monitored by TLC. After the reaction was completed, the mixture was distilled under reduced pressure to completely remove trifluoroacetic acid and dichloromethane, and the pure target compound I-3 was obtained as a yellow solid powder with a yield of about 75%. 1 HNMR(400MHz,DMSO-d6)δ8.63(t,J=5.3Hz,1H),8.26-8.23(m,2H),7.93(s,1H),7.79(d,J=7.9Hz,1H) ,7.65(s,1H),7.53(d,J=8.2Hz,1H),7.39(dd,J=8.4,5.6Hz,2H),7.21(t,J=7.5Hz,1H),7.16-7.08(m, 3H),6.94-6.86(m,4H),4.98(s,2H),4.26(t,J=6.9Hz,2H),3.78-3.73(m,1H),3.70-3.65(m,4H),3.09 (s,1H),3.04(s,5H),2.41-2.34(m,2H),1.93(s,3H),1.90(d,J=9.0Hz,2H),1.86(d,J=9.8Hz,2H).13C NMR(100MHz,DMSO-d6)δ168.52,166.32,163.23,160.81,148.63,148.42, 140.11,136.34,133.21,131.20,130.56,130.48,127.85,123.57,122.77 ,122.36,121.69,119.08,116.57,115.85,115.64,114.55,111.29,110.2 7,66.54,52.15,49.21,45.43,44.52,36.84,31.22,29.96,28.77,14.88.

[0070] Example 9

[0071] Preparation of (3-(3-((Z)-(Z)-3-(4-fluorobenzyl)-2-(4-morpholinylphenyl)imino)-4-oxothiazolidin-5-imino)methyl)-1H-indole-1-propyl)amino)-3-methyl-1-oxobutane-2-ylcarbamate (4-2)

[0072]

[0073] Boc-L-valine (0.20 mmol) and HATU (75.00 mg, 0.20 mmol) were added to a 50 mL dry round-bottom flask equipped with a stir bar. 10 mL of ultra-dry DMF solvent was added, and the mixture was stirred at room temperature for 5 minutes. DIPEA (41.28 mg, 0.32 mmol) was then added dropwise, and the mixture was stirred at room temperature for 30 minutes. Intermediate I-2 (0.16 mmol) was dissolved in 5 mL of ultra-dry DMF solvent and added dropwise to the mixture. The mixture was stirred overnight at room temperature. The reaction was monitored by TLC. After the reaction was complete, the reaction mixture was diluted with 50 mL of saturated sodium chloride solution and extracted with ethyl acetate (50 mL × 3). The organic phases were combined and dried over anhydrous sodium sulfate. Ethyl acetate was removed by vacuum distillation. The crude product was purified by column chromatography (petroleum ether: ethyl acetate = 2:1 v / v) to give a yellow solid powder 4-2, with a yield of approximately 65%. 1 H NMR (400MHz, DMSO-d6) δ8.01(s,1H),7.97(t,J=5.0Hz,1H),7.88(d,J=7.9Hz,1H),7.72(s,1H),7. 60(d,J=8.1Hz,1H),7.48(dd,J=8.6,5.6Hz,2H),7.30-7.18(m,4H),6.97(d,J=4.3Hz,4H),6.64(d ,J=8.6Hz,1H),5.07(s,2H),4.29(t,J=6.8Hz,2H),3.77-3.73(m,4H),3.73-3.68(m,1H),3.13-3. 10(m,4H),3.07(d,J=5.9Hz,2H),1.88(dt,J=16.1,8.2Hz,3H),1.36(s,9H),0.82(d,J=6.7Hz,6H). 13C NMR(100MHz,DMSO-d6)δ171.96,166.32,163.22,160.80,155.99,148.71,148.58 ,139.83,136.30,133.23,133.20,131.21,130.53,130.45,127.87,123.55,122.6 9,122.34,121.67,119.07,116.34,115.86,115.65,114.54,111.27,110.24,78.46,66.62,60.53,49.04,45.42,44.48,36.33,30.58,30.31,28.63,19.71,18.84.

[0074] Example 10

[0075] Preparation of 2-amino-N-(3-(4-bromo-3-((Z)-((Z)-3-(4-fluorobenzyl)-2-(4-morpholinylphenyl)imino)-4-oxothiazolidin-5-ylidene)methyl)-1H-indol-1-yl)propyl)-3-methylbutyramide (I-4)

[0076]

[0077] Intermediate 4-2 (50 mg) was added to a 50 mL dry round-bottom flask equipped with a stir bar, 10 mL of ultra-dry DCM solvent was added, and the mixture was stirred. Then, 5 mL of TFA was slowly added dropwise. The mixture was stirred at room temperature for 1 hour, and the reaction was monitored by TLC. After the reaction was completed, the mixture was distilled under reduced pressure to completely remove trifluoroacetic acid and dichloromethane, and the pure target compound I-4 was obtained as a yellow solid powder with a yield of about 85%. 1 HNMR(400MHz,DMSO-d6)δ8.69(t,J=4.9Hz,1H),8.24-8.21(m,2H),8.01(s,1H),7.88( d,J=7.8Hz,1H),7.72(s,1H),7.62(d,J=8.1Hz,1H),7.48(dd,J=7.9,5.8Hz,2H),7.30 (t,J=7.5Hz,1H),7.24-7.17(m,3H),7.03-6.96(m,4H),5.07(s,2H),3.77(s,4H),3.5 8-3.53(m,1H),3.13(s,6H),2.08-2.02(m,1H),1.97-1.90(m,2H),0.93-0.89(m,6H). 13C NMR(100MHz,DMSO-d6)δ168.25,166.31,163.23,160.81,148.61,148.40,140 .12,136.33,133.20,131.17,130.56,130.47,127.84,123.59,122.75,122.36 ,121.70,119.09,118.24,116.59,115.86,115.65,115.31,114.56,111.32,110.28,66.53,58.06,49.23,45.43,44.55,36.68,30.09,30.01,18.75,18.41.

[0078] Example 11

[0079] (R)-N-(3-(3-(Z)-(Z)-3-(4-fluorobenzyl)-2-(4-morpholinylphenyl)imino)-4-oxothiazolidine-5-ylide)methyl)-1H-indole-1-propyl)-1λ 2 Preparation of pyrrolidine-2-carboxamide (I-5)

[0080]

[0081] L-proline (0.20 mmol) and HATU (75.00 mg, 0.20 mmol) were added to a 50 mL dry round-bottom flask equipped with a stir bar. 10 mL of ultra-dry DMF solvent was added, and the mixture was stirred at room temperature for 5 minutes. Then, DIPEA (41.28 mg, 0.32 mmol) was added dropwise, and the mixture was stirred at room temperature for 30 minutes. Intermediate I-2 (0.16 mmol) was dissolved in 5 mL of ultra-dry DMF solvent and added dropwise to the mixture. The mixture was stirred overnight at room temperature. The reaction was monitored by TLC. After the reaction was complete, the reaction mixture was diluted with 50 mL of saturated sodium chloride solution and extracted with ethyl acetate (50 mL × 3). The organic phases were combined and dried over anhydrous sodium sulfate. Ethyl acetate was removed by vacuum distillation. The crude product was purified by column chromatography (petroleum ether: ethyl acetate = 2:1 v / v) to give a yellow solid powder I-5 in approximately 65% ​​yield. 1H NMR(400MHz, DMSO-d6)δ9.60(s,1H),8.63(t,J=4.9Hz,1H),8.51(s,1H),8.02(s,1H),7.88(d,J=7.9 Hz,1H),7.73(s,1H),7.60(d,J=8.1Hz,1H),7.48(dd,J=8.1,5.8Hz,2H),7.30(t,J=7.5Hz,1H),7.23 -7.19(m,2H),6.98(q,J=8.9Hz,4H),5.07(s,2H),4.34(t,J=6.6Hz,2H),4.13(t,J=8.3Hz,1H),3.76 (s,4H),3.23-3.17(m,2H),3.12(s,6H),2.28-2.20(m,1H),1.97-1.89(m,2H),1.88-1.78(m,3H).13C NMR(100MHz,DMSO-d6)δ168.51,166.33,163.23,160.81,148.64,139.93,136.33,133.22,133.19,131.24,130.56,130.48,127.86,123.60,12 2.76,122.34,121.71,116.42,115.88,115.67,114.55,111.27,110.25 ,66.60,59.48,49.09,46.00,45.44,44.41,36.98,29.95,29.90,24.00.

[0082] Example 12

[0083] Preparation of (2Z,5Z)-5-(5-bromo-1H-indol-3-yl)methylene)-3-(4-fluorobenzyl)-2-(4-morpholinylphenyl)imino)thiazolidin-4-one (3-2)

[0084]

[0085] Add 1.83 mmol of 5-bromoindole-3-carboxaldehyde and 40 mL of anhydrous ethanol to a 100 mL dry round-bottom flask equipped with a stirrer, followed by the dropwise addition of a catalytic amount of piperidine (14.88 mg, 0.17 mmol). Stir at room temperature for 5 minutes, then add intermediate 3 (0.70 g, 1.82 mmol) to the reaction system, stir, and reflux at 80 °C overnight. After cooling to room temperature, a large amount of yellow solid precipitates. Filter, wash twice with anhydrous ethanol, and dry to obtain yellow solid powder 3-2, with a yield of approximately 60%. 1H NMR (400MHz, DMSO-d6) δ12.16(s,1H),8.06(s,1H),7.98(s,1H),7.61(s,1H),7.44(dd,J=8.3,4.5Hz,3H),7.33 (d,J=7.5Hz,1H),7.17(t,J=8.8Hz,2H),6.93(d,J=3.8Hz,4H),5.02(s,2H),3.72(s,4H),3.09-3.05(m,4H).13C NMR(100MHz,DMSO-d6)δ166.19,163.21,160.79,148.80,139.88,135.31,133.17,130.51,130.43,129.79,129.02 ,126.01,122.97,122.38,121.48,116.34,115.89,115.68,115.26,114.87,114.05,110.66,66.61,49.02,45.38.

[0086] Example 13

[0087] Preparation of tert-butyl carbamate (4-(4-bromo-3-(Z)-3-(4-fluorobenzyl)-2-(4-morpholinylphenyl)imino)-4-oxothiazolidin-5-ylidene)methyl)-1H-indol-1-yl)propyl)carbamate (4-3)

[0088]

[0089] Intermediate 3-2 (0.93 mmol) and sodium hydride (0.03 g, 1.25 mmol) were added to a 100 mL dry round-bottom flask equipped with a stirrer. 30 mL of ultra-dry DMF solvent was then added to the system, and the mixture was stirred at room temperature for 5 minutes. N-Boc-3-aminopropyl bromide (0.27 g, 1.13 mmol) was dissolved in 10 mL of ultra-dry DMF solvent and added dropwise to the reaction mixture. After the addition was complete, the temperature was raised to 80 °C, and the reaction was allowed to proceed for 4 h. The reaction was monitored by TLC. After the reaction was complete, the mixture was cooled to room temperature, diluted with 100 mL of saturated sodium chloride solution, and extracted with ethyl acetate (100 mL × 3). The organic phases were combined and dried over anhydrous sodium sulfate. Ethyl acetate was removed by vacuum distillation to obtain a yellow solid, 4-3.

[0090] Example 14

[0091] Preparation of (2Z, 5Z)-5-(1-(3-aminopropyl)-5-bromo-1H-indole-3-methylene)-3-(4-fluorobenzyl)-2-(4-morpholinylphenyl)imino)thiazolidin-4-one (5-1)

[0092]

[0093] Add 4-3 to a 100 mL dry round-bottom flask equipped with a stir bar, add 30 mL of ultra-dry DCM solvent, stir, and then slowly add 15 mL of TFA. Stir at room temperature for 2 hours, and monitor the reaction by TLC. After the reaction is complete, distill under reduced pressure to completely remove trifluoroacetic acid and dichloromethane. The crude product obtained is purified by column chromatography (dichloromethane:methanol = 20:1 v / v) to give a yellow solid 5-1, with a yield of approximately 75%. 1 H NMR (400MHz, DMSO-d6) δ8.17(s,1H),8.01(s,1H),7.82(s,2H),7.76(s,1H),7.63(d,J=8.7Hz,1H),7.49-7.42(m,3H),7.19(t,J=8.8Hz,2H),6 .96(d,J=10.1Hz,4H),5.06(s,2H),4.38(t,J=6.6Hz,2H),3.77-3.73( m,4H),3.13-3.09(m,4H),2.73(t,J=7.4Hz,2H),2.03-1.97(m,2H).13C NMR(100MHz,DMSO-d6)δ166.21,163.26,160.84,148.76,148.54,139.88,135.13,133.19,132.08,130.59,130.51,129.62,1 26.20,122.31,121.96,116.36,115.91,115.70,115.63,114.61,113.37,110.17,66.65,49.03,45.50,44.02,36.88,28.35.

[0094] Example 15

[0095] Preparation of (3-(4-bromo-3-(Z)-3-(4-fluorobenzyl)-2-(4-morpholinylphenyl)imino)-4-oxothiazolidine-5-ylidene)methyl)-1H-indol-1-yl)propyl)amino)-3-methyl-1-oxobutane-2-carbamate (6-1)

[0096]

[0097] Boc-L-valine (0.20 mmol) and HATU (75.00 mg, 0.20 mmol) were added to a 50 mL dry round-bottom flask equipped with a stir bar. 10 mL of ultra-dry DMF solvent was added, and the mixture was stirred at room temperature for 5 minutes. DIPEA (41.28 mg, 0.32 mmol) was then added dropwise, and the mixture was stirred at room temperature for 30 minutes. Intermediate 5-1 (0.16 mmol) was dissolved in 5 mL of ultra-dry DMF solvent and added dropwise to the mixture. The mixture was stirred overnight at room temperature. The reaction was monitored by TLC. After the reaction was complete, the reaction mixture was diluted with 50 mL of saturated sodium chloride solution and extracted with ethyl acetate (50 mL × 3). The organic phases were combined and dried over anhydrous sodium sulfate. Ethyl acetate was removed by vacuum distillation. The crude product was purified by column chromatography (petroleum ether: ethyl acetate = 2:1 v / v) to give a yellow solid powder 6-1 in approximately 65% ​​yield. 1 H NMR (400MHz, DMSO-d6) δ8.15(s,1H),8.01(s,1H),7.97(t,J=4.8Hz,1H),7.75(s,1H),7.59(d,J=8. 7Hz,1H),7.48(dd,J=8.3,5.7Hz,2H),7.40(d,J=8.6Hz,1H),7.20(t,J=8.8Hz,2H),6.97(d,J=4.4Hz ,4H),6.66(d,J=8.5Hz,1H),5.07(s,2H),4.29(s,2H),3.77-3.73(m,4H),3.71-3.67(m,1H),3.13-3 .09(m,4H),3.07-3.02(m,2H),1.86(dt,J=14.7,7.2Hz,3H),1.35(s,9H),0.81(d,J=6.7Hz,6H).13C NMR(100MHz,DMSO-d6)δ171.95,166.18,163.23,160.80,156.00,148.74,148 .42,139.76,135.09,133.20,132.21,130.54,130.45,129.58,126.06,122.3 5,121.83,116.33,115.87,115.66,115.46,114.45,113.33,109.97,78.44,66.63,60.55,49.04,45.45,44.64,36.26,30.58,30.27,28.63,19.71,18.84.

[0098] Example 16

[0099] Preparation of 2-amino-N-(3-(4-bromo-3-((Z)-((Z)-3-(4-fluorobenzyl)-2-(4-morpholinylphenyl)imino)-4-oxothiazolidine-5-ylidene)methyl)-1H-indol-1-yl)propyl)-3-methylbutyramide (I-6)

[0100]

[0101] Intermediate 6-1 (50 mg) was added to a 50 mL dry round-bottom flask equipped with a stir bar, 10 mL of ultra-dry DCM solvent was added, and the mixture was stirred. Then, 5 mL of TFA was slowly added dropwise, and the mixture was stirred at room temperature for 1 hour. The reaction was monitored by TLC. After the reaction was completed, the mixture was distilled under reduced pressure to completely remove trifluoroacetic acid and dichloromethane, and the pure target compound I-6 was obtained as a yellow solid powder with a yield of about 85%. 1 HNMR(400MHz,DMSO-d6)δ8.15(s,1H),8.00(s,2H),7.79(s,1H),7.59(d,J=8.7Hz,1H),7. 50-7.45(m,2H),7.40(d,J=8.6Hz,1H),7.20(t,J=8.6Hz,2H),6.97(q,J=8.8Hz,4H),5.07 (s,2H),4.31(t,J=5.9Hz,2H),3.75(s,4H),3.12(s,4H),3.07-3.02(m,2H),2.92(d,J=4. 8Hz,1H),2.13(s,1H),1.89-1.83(m,2H),0.84(d,J=6.6Hz,3H),0.77(d,J=6.6Hz,3H).13C NMR(100MHz,DMSO-d6)δ174.94,166.18,163.22,160.80,148.75,148.44,139.74,135.10,133.21,132.35,130.52,130.44,129.61,126.07,12 2.36,121.84,116.34,115.88,115.67,115.42,114.45,113.38,109.91 ,66.63,60.57,49.04,45.44,44.76,36.18,32.01,30.30,20.00,17.82.

[0102] Example 17

[0103] Preparation of (3-(4-bromo-3-(Z)-3-(4-fluorobenzyl)-2-(4-morpholinylphenyl)imino)-4-oxothiazolidin-5-imino)methyl)-1H-indol-1-yl)propyl)amino)-4-methylthio-1-oxobutane-2-carbamate (6-2)

[0104]

[0105] Boc-L-methionine (0.20 mmol) and HATU (75.00 mg, 0.20 mmol) were added to a 50 mL dry round-bottom flask equipped with a stir bar. 10 mL of ultra-dry DMF solvent was added, and the mixture was stirred at room temperature for 5 minutes. DIPEA (41.28 mg, 0.32 mmol) was then added dropwise, and the mixture was stirred at room temperature for 30 minutes. Intermediate 5-1 (0.16 mmol) was dissolved in 5 mL of ultra-dry DMF solvent and added dropwise to the mixture. The mixture was stirred overnight at room temperature. The reaction was monitored by TLC. After the reaction was complete, the reaction mixture was diluted with 50 mL of saturated sodium chloride solution and extracted with ethyl acetate (50 mL × 3). The organic phases were combined and dried over anhydrous sodium sulfate. Ethyl acetate was removed by vacuum distillation. The crude product was purified by column chromatography (petroleum ether: ethyl acetate = 2:1 v / v) to give a yellow solid powder 6-2, with a yield of approximately 65%. 1H NMR(400MHz,DMSO-d6)δ8.15(s,1H),8.01(s,1H),7.94(s,1H),7.78(s,1H),7.59(d,J= 8.6Hz,1H),7.50-7.45(m,2H),7.39(d,J=8.5Hz,1H),7.20(t,J=8.8Hz,2H),6.97(s,5H) ,5.07(s,2H),4.29(s,2H),3.98-3.92(m,1H),3.75(s,4H),3.11(s,4H),3.08-3.01(m,2 H),2.47-2.39(m,2H),2.01(s,3H),1.83(dq,J=24.5,11.0,8.9Hz,4H),1.35(s,9H).13C NMR(100MHz,DMSO-d6)δ172.30,166.18,163.22,160.80,155.94,148.73,1 48.44,139.75,135.09,133.21,132.27,130.54,130.46,129.59,126.02,12 2.37,121.84,116.33,115.87,115.66,115.42,114.44,113.32,109.95,78.58,66.63,54.25,49.03,45.44,44.59,36.41,31.93,30.27,28.63,15.08.

[0106] Example 18

[0107] Preparation of (R)-2-amino-N-(3-(4-bromo-3-((Z)-((Z)-3-(4-fluorobenzyl)-2-(4-morpholinylphenyl)imino)-4-oxothiazolidin-5-ylidene)methyl)-1H-indol-1-yl)propyl)-4-methylthiobutyramide (I-7)

[0108]

[0109] Intermediate 6-2 (50 mg) was added to a 50 mL dry round-bottom flask equipped with a stir bar, 10 mL of ultra-dry DCM solvent was added, and the mixture was stirred. Then, 5 mL of TFA was slowly added dropwise. The mixture was stirred at room temperature for 1 hour, and the reaction was monitored by TLC. After the reaction was completed, the mixture was distilled under reduced pressure to completely remove trifluoroacetic acid and dichloromethane, and the pure target compound I-7 was obtained as a yellow solid powder with a yield of about 75%. 1HNMR(400MHz,DMSO-d6)δ8.15(s,1H),8.10(s,1H),8.01(s,1H),7.81(s,1H),7.59(d,J=8.7Hz,1H ),7.51-7.44(m,2H),7.40(d,J=8.7Hz,1H),7.20(t,J=8.6Hz,2H),7.01-6.94(m,4H),5.07(s,2H), 4.31(t,J=6.2Hz,2H),3.76(s,4H),3.26-3.22(m,1H),3.12(s,4H),3.07-3.01(m,2H),2.68(s,2H) ,2.51(s,2H),2.00(s,3H),1.91-1.85(m,2H),1.83-1.76(m,1H),1.60(dt,J=13.4,6.9Hz,1H).13C NMR(100MHz,DMSO-d6)δ174.91,166.19,163.21,160.80,148.74,148.46,139.74,135 .10,133.19,132.41,130.53,130.44,129.60,126.04,122.43,122.37,121.82,116.35 ,115.88,115.67,115.39,114.44,113.40,109.88,66.63,54.30,49.03,45.44,44.72,40.62,40.41,40.19,39.99,39.78,39.57,39.36,36.32,34.78,30.27,30.16,15.07.

[0110] Example 19

[0111] Preparation of tert-butyl((2R,3R)-1-(3-(4-bromo-3-(Z)-3-(4-fluorobenzyl)-2-((4-morpholinylphenyl)imino)-4-oxothiazolidinylbutyrate-5-ylidene)methyl)-1H-indol-1-yl)propyl)amino)-3-methyl-1-oxopentane-2-ylcarbamate (6-3)

[0112]

[0113] Boc-L-isoleucine (0.20 mmol) and HATU (75.00 mg, 0.20 mmol) were added to a 50 mL dry round-bottom flask equipped with a stir bar. 10 mL of ultra-dry DMF solvent was added, and the mixture was stirred at room temperature for 5 minutes. DIPEA (41.28 mg, 0.32 mmol) was then added dropwise, and the mixture was stirred at room temperature for 30 minutes. Intermediate 5-1 (0.16 mmol) was dissolved in 5 mL of ultra-dry DMF solvent and added dropwise to the mixture. The mixture was stirred overnight at room temperature. The reaction was monitored by TLC. After the reaction was complete, the reaction mixture was diluted with 50 mL of saturated sodium chloride solution and extracted with ethyl acetate (50 mL × 3). The organic phases were combined and dried over anhydrous sodium sulfate. Ethyl acetate was removed by vacuum distillation. The crude product was purified by column chromatography (petroleum ether: ethyl acetate = 2:1 v / v) to give a yellow solid powder 6-3, with a yield of approximately 60%. 1 H NMR (400MHz, DMSO-d6) δ8.15(s,1H),8.01(s,1H),7.97(s,1H),7.76(s,1H),7.58(d,J=8.7Hz,1H),7. 50-7.46(m,2H),7.39(d,J=8.4Hz,1H),7.20(t,J=8.7Hz,2H),6.97(d,J=3.5Hz,4H),6.70(d,J=8.4Hz, 1H),5.07(s,2H),4.29(s,2H),3.75(s,4H),3.71(s,1H),3.11(s,4H),3.05(s,2H),1.86(dt,J=17.7, 9.3Hz,2H),1.69-1.62(m,1H),1.35(s,9H),1.24(s,1H),1.12-1.04(m,1H),0.79(t,J=7.9Hz,6H).13C NMR (100MHz, DMSO-d6) δ172.02,166.18,163.23,160.80,155.94,148.74,148. 42,139.75,135.08,133.19,132.18,130.53,130.45,129.57,126.07,122.35,1 21.84,116.33,115.87,115.66,115.45,114.45,113.31,109.97,78.45,66.62,59.41,49.03,45.44,44.62,36.59,36.25,30.25,28.62,25.01,15.92,11.42.

[0114] Example 20

[0115] Preparation of 2-amino-N-(3-(4-bromo-3-((Z)-((Z)-3-(4-fluorobenzyl)-2-(4-morpholinylphenyl)imino)-4-oxothiazolidin-5-ylidene)methyl)-1H-indole-1-propyl)-3-methylpentanamide (I-8)

[0116]

[0117] Intermediate 6-3 (50 mg) was added to a 50 mL dry round-bottom flask equipped with a stir bar, 10 mL of ultra-dry DCM solvent was added, and the mixture was stirred. Then, 5 mL of TFA was slowly added dropwise, and the mixture was stirred at room temperature for 1 hour. The reaction was monitored by TLC. After the reaction was completed, the mixture was distilled under reduced pressure to completely remove trifluoroacetic acid and dichloromethane, and the pure target compound I-8 was obtained as a yellow solid powder with a yield of about 80%. 1 HNMR(400MHz, DMSO-d6)δ8.16(s,1H),8.01(s,1H),7.95(t,J=5.1Hz,1H),7.78(s,1H),7.58(d,J=8.7Hz,1H),7.48 (dd,J=7.9,5.8Hz,2H),7.40(d,J=8.7Hz,1H),7.20(t,J=8.7Hz,2H),6.97(d,J=5.5Hz,4H),5.07(s,2H),4.30(t,J =6.7Hz,2H),3.77-3.73(m,4H),3.13-3.10(m,4H),3.04(q,J=6.0Hz,2H),2.94(d,J=5.5Hz,1H),1.98(s,2H),1.89 -1.83(m,2H),1.59-1.53(m,1H),1.39(dd,J=12.2,4.4Hz,1H),1.03(dd,J=13.6,7.9Hz,1H),0.82-0.78(m,6H).13C NMR(100MHz,DMSO-d6)δ168.17,166.18,163.24,160.82,148.71,148.45,139 .83,135.12,133.19,132.11,130.58,130.50,129.58,126.11,122.44,122.3 2,121.94,116.36,115.88,115.67,115.53,114.50,113.31,110.04,66.62,57.12,55.39,49.05,45.46,44.70,36.69,36.43,30.02,24.63,15.03,11.56.

[0118] Example 21

[0119] Cell counting method for detecting HepG2 (commercial) liver cancer cells. Drug treatment IC 50

[0120] Taking a 24-well plate as an example, each well is filled with 2×10 4 HepG2 cells were incubated in Dulbecco's Modified Eagle medium. Three replicates were set up for each drug concentration, following the example of a blank control group (DMSO). The following day, drugs were added according to a concentration gradient. After 72 hours of treatment, the DMEM medium (commercial) in the 24-well plate was aspirated. Adhering cells were rinsed with 200 μL of PBS solution (pH 7.4), and the PBS solution was discarded. 200 μL of trypsin was added to each well. The plate was capped and shaken a few times to ensure the trypsin fully contacted the cells. The trypsin was then aspirated from each well. The plate was incubated for 2 minutes. 200 μL of DMEM medium containing 10% FBS was added to each well. The cells were then pipetted with the medium to completely detach the cells, yielding a HepG2 cell suspension.

[0121] Prepare several sterile 1.5 mL centrifuge tubes, labeling the tube caps with the drug name, drug concentration, and replicate well number. Pipette 10 μL of HepG2 cell suspension into each tube and add 10 μL of 0.08% trypan blue solution. Mix thoroughly by pipetting and use a hemocytometer to count viable cells. Compare the data from the drug-treated group with the blank control DMSO group to obtain the cell proliferation inhibition rate and drug IC50. 50 The values ​​were analyzed and compared based on the data from the three replicates. The results are shown in the table below:

[0122] Compound serial number <![CDATA[IC 50 (μM)]]> Ⅰ-1 19.78μM Ⅰ-2 5.136μM Ⅰ-3 18.76μM Ⅰ-4 1.37μM Ⅰ-5 2.46μM Ⅰ-6 3.57μM Ⅰ-7 >100μM Ⅰ-8 >100μM

[0123] Formulations, such as tablets, capsules, or injections, made from any one of the compounds I-1 to I-8 of the present invention with pharmaceutically acceptable excipients using conventional techniques, may also be used in the preparation of antitumor drugs.

Claims

1. Five-membered heterocyclic benzoindole derivatives, characterized by: The structural formula is shown in equation (I): in: R1 is H or Br; R2 is H, 2. The use of the five-membered heterocyclic indole derivative of claim 1 in the preparation of anti-liver cancer drugs.

Images