An anti-tumor cinnamyl piperazine resveratrol conjugate or a pharmaceutically acceptable salt thereof and a preparation method and application thereof

Abstract

The present invention belongs to the technical field of compound synthesis and pharmaceutical application, and discloses an anti-tumor cinnamyl piperazine resveratrol conjugate or a pharmaceutically acceptable salt thereof, and a preparation method and application thereof. The present invention synthesizes cinnamyl piperazine resveratrol conjugate and a pharmaceutically acceptable salt thereof for the first time, and finds that the compound has good anti-tumor activity and can inhibit the proliferation and migration of various tumor cells. Therefore, the conjugate synthesized by the present invention has good application prospects in new anti-tumor drugs and is outstanding in innovation.

Description

Technical Field

[0001] The invention belongs to the technical field of compound synthesis and pharmaceutical application, in particular to an anti-tumor cinnamyl piperazine resveratrol conjugate or a pharmaceutically acceptable salt thereof, and a preparation method and application thereof. Background Art

[0002] Cancer seriously threatens human health and life span. Anti-tumor drugs have obvious toxic side effects and drug resistance. The research and development of new, highly effective and low-toxic anti-tumor drugs has always been a hot topic in the field of new drugs.

[0003] Cinnamylpiperazine-resveratrol conjugates refer to a new type of conjugates formed by coupling cinnamylpiperazine pharmacophores with resveratrol and its derivatives via a connecting bridge through a molecular hybridization strategy.

[0004] Resveratrol is a non-flavonoid natural polyphenol compound with pharmacological effects against colon cancer, cervical cancer, gastric cancer, acute T lymphocytic leukemia and liver cancer, and can significantly inhibit the proliferation of colorectal cancer stem cells in a dose-dependent manner. Pterostilbene is a methylated derivative of resveratrol, which has multiple biological activities such as antioxidant, anti-tumor, lipid-lowering and antibacterial, and has higher bioavailability and structural stability than resveratrol.

[0005] As an important raw material for drug intermediates, cinnamylpiperazine was mainly used in the early days to prepare analgesics and drugs for the treatment of cardiovascular and cerebrovascular diseases. For example, in 1955, Cinnarizine, launched by Janssen Pharmaceuticals of Belgium, is a typical vasodilator that has a certain therapeutic effect on diseases such as cerebral thrombosis, coronary artery sclerosis, cerebral hemorrhage, cerebral embolism, inner ear vertigo, and cerebral circulation disorders caused by hypertension. On this basis, the company launched Flunarizine Hydrochloride in 1982. Compared with Cinnarizine, the drug is safer and has better therapeutic effects on migraine and vertigo. In addition, Bucinazine Hydrochloride, as a moderate analgesic, can be used to treat trigeminal neuralgia, migraine, neuralgia, joint pain, postoperative pain, and cancer pain. 4-Thiourea-cinnamylpiperazine derivative LF3 (CAS 664969-54-4) is a Wnt signaling pathway inhibitor and a small molecule antagonist of the β-catenin / TCF4 interaction. It can competitively inhibit the binding of β-catenin to TCF4, inhibit the proliferation of colon cancer cells by inducing cell cycle arrest, and inhibit the self-renewal of tumor stem cells. It also has strong anti-tumor activity in colon cancer mouse xenograft tumor experiments and is a promising new target therapy for colon cancer. The hydrophobic tail of LF3, the cinnamylpiperazine fragment, is predicted to be inserted into the hydrophobic cleft arranged by the Cys466 and Pro463 residues of β-catenin to promote its inhibitory effect.

[0006] Breast cancer is the world's number one cancer, of which triple-negative breast cancer accounts for about 15%-20% of all breast cancers, with the characteristics of strong invasiveness, high recurrence rate and poor prognosis. Advanced triple-negative breast cancer is insensitive to targeted therapy and endocrine therapy, and lacks specific treatment methods. At present, chemotherapy is still the main treatment for triple-negative breast cancer in my country, but the efficacy of either single-drug or combined chemotherapy is poor, with a median survival of about 9-12 months and a 5-year survival rate of less than 30%. Therefore, the discovery of new specific therapeutic drugs for triple-negative breast cancer has important research value and market application potential. In addition, liver cancer ranks seventh in cancer incidence and third in cancer-related mortality, posing a major challenge to public health worldwide. Colon cancer is also one of the common tumors in my country, so the development of more efficient and less toxic new drugs for the treatment of liver cancer and colon cancer also has good market value.

[0007] In summary, with the goal of discovering new, highly effective and low-toxic anti-tumor drugs to make up for the shortcomings of existing technologies, the present invention selects the food-borne, low-toxic anti-tumor natural product resveratrol and the anti-tumor pharmacophore cinnamyl piperazine, adopts a molecular hybridization drug design strategy, and synthesizes a series of novel cinnamyl piperazine-resveratrol conjugates and their salts with anti-tumor activity through coupling with different connecting bridges, and evaluates their in vitro anti-tumor activity against a variety of tumor cells.

[0008] Through searching, the following public document related to the patent application of the present invention is found:

[0009] Li Rongxia, Synthesis and anti-tumor activity of cinnamyl piperazine conjugates, Master's thesis of Tianjin University of Science and Technology, 2023. This study is an earlier study of this research group. The compounds in this invention are new compounds discovered on the basis of this thesis after a lot of work and exploration. The structures are different from those of the compounds in the previous thesis. They are all new compounds with better anti-tumor activity, wider anti-tumor spectrum and further improved solubility. Summary of the invention

[0010] The purpose of the present invention is to overcome the deficiencies in the prior art and provide an anti-tumor cinnamyl piperazine resveratrol conjugate or a pharmaceutically acceptable salt thereof with a glycol chain as a connecting chain, and a preparation method and application thereof.

[0011] The technical solution adopted by the present invention to solve its technical problem is:

[0012] An anti-tumor cinnamyl piperazine resveratrol conjugate or a pharmaceutically acceptable salt thereof, the general structural formula of which is as follows:

[0013]

[0014] The linker is a straight or branched alkyl, ether or amine containing 1 to 6 atoms, R1 and R2 are the same or different, and R1 and R2 are hydrogen, hydroxyl, CH3O-, CH3 (CH2) n O-, CH3COO-, CH3 (CH2) m COO-, where n=1~3, m=1~2.

[0015] Furthermore, the anti-tumor cinnamyl piperazine resveratrol conjugate or a pharmaceutically acceptable salt thereof has a structural formula of one of the following:

[0016] .

[0017] The method for preparing the anti-tumor cinnamyl piperazine resveratrol conjugate or a pharmaceutically acceptable salt thereof as described above comprises the following steps:

[0018] First, 3, 5-dimethoxybenzaldehyde is subjected to a witting reaction to obtain compound 1; then 4-iodophenol is used as a raw material to connect acetyl chloride through an acylation reaction to obtain compound 2, and compound 1 is connected through a Heck reaction to obtain compound 3, which is hydrolyzed to obtain compound 4, and then connected with different connecting bridges through a Williams ether synthesis method to obtain compound 5a or compound 5b, and then connected with a nitrogen-containing compound such as N-tert-butyloxycarbonyl (Boc)-piperazine to obtain compound 6a or compound 6b, after removing the protecting group, an alkylation reaction is carried out with cinnamyl bromide to obtain compound 7 or compound 8, and hydrogen chloride gas is introduced into a methanol or acetone solution to obtain the corresponding hydrochloride salt 9 or 10. The reaction route is:

[0019]

[0020] ;

[0021] Alternatively, intermediate 4 is used as a raw material, connected with 2, 2'-dibromodiethyl ether to obtain compound 11, connected with N-Boc-piperazine to obtain compound 12, and finally after removing the tert-butyl ester, connected with cinnamyl bromide to obtain compound 13, and further introduced with acidic gas to obtain its hydrochloride 14. The reaction route is:

[0022] ;

[0023] Alternatively, triethylene glycol is activated to obtain compound 15, which is then subjected to nucleophilic substitution reaction with sodium iodide to obtain compound 16, and compound 4 is connected to obtain compound 17. Compound 17 is activated by hydroxyl to obtain compound 18 and compound 19, which are then connected to piperazine and cinnamyl to obtain compound 20, and acidic gas is further introduced to obtain its hydrochloride 21. The reaction route is:

[0024]

[0025] ;

[0026] Compound 7 was demethylated by BBr3 ether solution at low temperature to obtain resveratrol derivative 22;

[0027] .

[0028] Use of the anti-tumor cinnamylpiperazine-resveratrol conjugate or a pharmaceutically acceptable salt thereof in the preparation of a drug for treating liver cancer.

[0029] Use of the anti-tumor cinnamyl piperazine-resveratrol conjugate or a pharmaceutically acceptable salt thereof in preparing a drug for treating colon cancer.

[0030] Use of the anti-tumor cinnamylpiperazine-resveratrol conjugate or a pharmaceutically acceptable salt thereof as described above in the preparation of drugs for treating breast cancer and triple-negative breast cancer.

[0031] Use of the anti-tumor cinnamylpiperazine-resveratrol conjugate or a pharmaceutically acceptable salt thereof in preparing a drug for treating tumors.

[0032] The advantages and positive effects achieved by the present invention are:

[0033] 1. The present invention synthesizes cinnamylpiperazine-resveratrol conjugates and pharmaceutically acceptable salts thereof for the first time, and finds that such compounds have good anti-tumor activity, can inhibit the proliferation of various tumor cells, can significantly inhibit the proliferation activity of liver cancer, colon cancer, breast cancer, and triple-negative breast cancer cells in vitro, and can inhibit the lateral and longitudinal migration ability of tumor cells. Therefore, the conjugates synthesized by the present invention have good application prospects in new anti-tumor drugs and are outstanding in innovation.

[0034] 2. The reaction steps of the method of the present invention are simple to operate, and the raw materials and reagents are cheap and easily available, which is suitable for large-scale production and development.

[0035] 3. Some compounds in the method of the present invention are prepared into hydrochloride salts, which have increased water solubility and improved lipid-water partition coefficient, and have good drug potential.

[0036] 4. The present invention discovers for the first time that cinnamyl piperazine-resveratrol conjugate and its hydrochloride can not only inhibit the proliferation of liver cancer, colon cancer, breast cancer and triple-negative breast cancer cells, but also inhibit the migration of liver cancer and triple-negative breast cancer cells. Currently, there is a lack of small molecule drugs for the treatment of triple-negative breast cancer in clinical applications. Therefore, the present invention has good market application prospects. BRIEF DESCRIPTION OF THE DRAWINGS

[0037] Figure 1 is the H NMR spectrum of compound 7 in deuterated chloroform;

[0038] Figure 2 is the carbon NMR spectrum of compound 7 in deuterated chloroform;

[0039] Figure 3 is the H NMR spectrum of compound 9 in deuterated methanol;

[0040] Figure 4 is the H NMR spectrum of compound 8 in deuterated chloroform;

[0041] Figure 5 is the carbon NMR spectrum of compound 8 in deuterated chloroform;

[0042] Figure 6is the H NMR spectrum of compound 13 in deuterated chloroform;

[0043] Figure 7 is the carbon NMR spectrum of compound 13 in deuterated chloroform;

[0044] Figure 8 is the H NMR spectrum of compound 20 in deuterated chloroform;

[0045] Fig. 9 is the carbon NMR spectrum of compound 20 in deuterated chloroform;

[0046] Fig.10 is the H NMR spectrum of compound 21 in deuterated methanol;

[0047] Fig.11 This is a graph showing the effect of compound 7 of the present invention on the lateral migration ability of HepG2 cells (scratch test);

[0048] Fig.12 This is a grayscale analysis diagram of the effect of compound 7 of the present invention on the lateral migration ability of HepG2 cells (scratch test); compared with the solvent blank group (DMSO group), ***P<0.001, **P<0.01;

[0049] Fig.13 This is a graph showing the effect of compound 22 of the present invention on the lateral migration ability of MDA-MB-231 cells (scratch test);

[0050] Fig.14 This is a grayscale analysis diagram of the effect of compound 22 of the present invention on the lateral migration ability of MDA-MB-231 cells (scratch test); compared with the solvent blank group (DMSO group), ***P<0.001;

[0051] Fig.15 This is a cell staining diagram showing the effect of compound 7 of the present invention on the longitudinal migration ability (transwell) of HepG2 cells;

[0052] Fig.16 This is a cell number analysis diagram of the effect of compound 7 of the present invention on the longitudinal migration ability (transwell) of HepG2 cells; compared with the solvent blank group (DMSO group), ***P<0.001;

[0053] Fig.17 This is a cell staining diagram showing the effect of compound 22 of the present invention on the longitudinal migration ability (transwell) of MDA-MB-231 cells;

[0054] Fig.18This is a cell number analysis chart of the effect of compound 22 of the present invention on the longitudinal migration ability (transwell) of MDA-MB-231 cells. Compared with the solvent blank group (DMSO group), **P<0.01; DETAILED DESCRIPTION

[0055] The present invention will be further described below in conjunction with the embodiments. The following embodiments are descriptive rather than restrictive, and the protection scope of the present invention cannot be limited by the following embodiments.

[0056] The various experimental operations involved in the specific embodiments are all routine techniques in the art. For parts not specially annotated in this document, ordinary technicians in the art can implement them by referring to various commonly used reference books, scientific and technological literature or related instructions, manuals, etc. before the filing date of this invention.

[0057] Unless otherwise specified, the reagents involved in the embodiments of the present invention are all commercially available products and can be purchased through commercial channels.

[0058] An anti-tumor cinnamyl piperazine resveratrol conjugate or a pharmaceutically acceptable salt thereof, the general structural formula of which is as follows:

[0059]

[0060] The linker is a straight or branched alkyl, ether or amine containing 1 to 6 atoms, R1 and R2 are the same or different, and R1 and R2 are hydrogen, hydroxyl, CH3O-, CH3 (CH2) n O-, CH3COO-, CH3 (CH2) m COO-, where n=1~3, m=1~2.

[0061] Preferably, the anti-tumor cinnamyl piperazine resveratrol conjugate or a pharmaceutically acceptable salt thereof has a structural formula of one of the following:

[0062] .

[0063] The method for preparing the anti-tumor cinnamyl piperazine resveratrol conjugate or a pharmaceutically acceptable salt thereof as described above comprises the following steps:

[0064] First, 3, 5-dimethoxybenzaldehyde is subjected to a witting reaction to obtain compound 1; then 4-iodophenol is used as a raw material to connect acetyl chloride through an acylation reaction to obtain compound 2, and compound 1 is connected through a Heck reaction to obtain compound 3, which is hydrolyzed to obtain compound 4, and then connected with different connecting bridges through a Williams ether synthesis method to obtain compound 5a or compound 5b, and then connected with a nitrogen-containing compound such as N-tert-butyloxycarbonyl (Boc)-piperazine to obtain compound 6a or compound 6b, after removing the protecting group, alkylation reaction with cinnamyl bromide is carried out to obtain compound 7 or compound 8, and hydrogen chloride gas is introduced into a methanol or acetone solution to obtain the corresponding hydrochloride 9 or 10. The reaction route is as follows:

[0065] ;

[0066] Alternatively, intermediate 4 is used as a raw material, connected with 2, 2'-dibromodiethyl ether to obtain compound 11, connected with N-Boc-piperazine to obtain compound 12, and finally after removing the tert-butyl ester, connected with cinnamyl bromide to obtain compound 13, and further introduced with acidic gas to obtain its hydrochloride 14. The reaction route is:

[0067]

[0068] ;

[0069] Alternatively, triethylene glycol is activated to obtain compound 15, which is then subjected to nucleophilic substitution reaction with sodium iodide to obtain compound 16, and compound 4 is connected to obtain compound 17. Compound 17 is activated by hydroxyl to obtain compound 18 and compound 19, which are then connected to piperazine and cinnamyl to obtain compound 20, and acidic gas is further introduced to obtain its hydrochloride 21. The reaction route is:

[0070] ;

[0071] Compound 7 was demethylated by BBr3 ether solution at low temperature to obtain resveratrol derivative 22;

[0072] .

[0073] Use of the anti-tumor cinnamylpiperazine-resveratrol conjugate or a pharmaceutically acceptable salt thereof in the preparation of a drug for treating liver cancer.

[0074] Use of the anti-tumor cinnamyl piperazine-resveratrol conjugate or a pharmaceutically acceptable salt thereof in preparing a drug for treating colon cancer.

[0075] Use of the anti-tumor cinnamylpiperazine-resveratrol conjugate or a pharmaceutically acceptable salt thereof as described above in the preparation of drugs for treating breast cancer and triple-negative breast cancer.

[0076] Use of the anti-tumor cinnamylpiperazine-resveratrol conjugate or a pharmaceutically acceptable salt thereof in preparing a drug for treating tumors.

[0077] Specifically, the relevant preparation and detection are as follows:

[0078] The synthetic routes of compounds 1 to 22 are as follows:

[0079] Among them, the synthetic routes of compounds 7~10 are:

[0080]

[0081] Example 1 Synthesis of 1-cinnamyl-4-(2-(4-((E)-3,5-dimethoxyphenylvinylphenoxy)ethyl)piperazine (7) and its hydrochloride 9

[0082] As shown in the synthetic routes of the above compounds 7 to 10, the specific steps are as follows:

[0083] (1) Synthesis of 1,3-dimethoxy-5-vinylbenzene (1)

[0084] Under argon protection, methyl triphenylphosphonium bromide (10.93 g, 30.69 mmol) and sodium amide (1.596 g, 40.92 mmol) were first dissolved in 25 mL of anhydrous tetrahydrofuran (THF), stirred at room temperature for 10 h, and then 3,5-dimethoxybenzaldehyde (3.4 g, 20.46 mmol) was dissolved in 5 mL of THF and slowly added to the reaction solution at -10 °C. After 0.5 h, the mixture was heated to room temperature and stirred for 2-4 h. After the reaction was complete, the mixture was filtered, and the filtrate was concentrated under reduced pressure and separated and purified by silica gel column chromatography (PE: EA = 300:1-150:1, v / v, volume ratio) to obtain a colorless and transparent oily product 1 with a yield of 96.0%. 1 H-NMR (400 MHz, CDCl3) δ 6.62-6.69 (m, 1H), 6.58 (d, J = 2.4 Hz, 2H), 6.40 (t, J = 2.2 Hz, 1H), 5.74 (d, J = 17.6 Hz, 1H), 5.26 (d, J = 10.8 Hz, 1H), 3.81 (s, 6H).

[0085] (2) Synthesis of 4-iodophenylacetate (2)

[0086] At 0 °C, 4-iodophenol (2.2 g, 10.0 mmol) was dissolved in 10 mL of anhydrous dichloromethane (DCM), and acetyl chloride (0.85 mL, 12.0 mmol) and triethylamine (Et3N, 1.67 mL, 12.0 mmol) were added while stirring. After 0.5 h, the temperature was raised to room temperature and stirred for 2 h. After the reaction was complete, 10 mL of ice water was added to quench the reaction, and DCM was extracted (10 mL×3). The organic phases were combined, dried over anhydrous sodium sulfate, and purified on a silica gel column to obtain a white solid product 2 with a yield of 94.6%. 1 H-NMR (400 MHz, CDCl3)δ 7.67-7.70 (m, 2H), 6.85-6.88 (m, 2H), 2.29 (s, 3H).

[0087] (3) Synthesis of (E)-4-(3,5-dimethoxyphenylvinyl)phenylacetate (3)

[0088] Under argon protection, compound 2 (1.0 g, 3.82 mmol), compound 1 (752 mg, 4.58 mmol), tetrabutylammonium bromide (TBAB) (1.85 g, 5.73 mmol), palladium acetate (42.9 mg, 0.191 mmol), potassium acetate (562.3 mg, 5.73 mmol) were dissolved in 5 mL of anhydrous N,N-dimethylformamide (DMF) in sequence and heated at 80 °C for 3 h. After the reaction was completed, 20 mL of ice water was added to the system, and DCM was extracted (10 mL×3). The organic phases were combined, dried over anhydrous sodium sulfate, and the organic solvent was removed. The crude product was chromatographed on a silica gel column (PE: EA = 100:1 ~ 20:1, v / v, volume ratio) to obtain a white solid product 3 with a yield of 87.0%. 1 H-NMR (400 MHz, CDCl3) δ 7.51 (d, J = 8.4 Hz, 2H), 7.09 (d, J = 8.4 Hz, 2H), 7.02 (dd, J = 16.2 Hz, 33.4 Hz, 2H), 6.66 (d, J = 2.0 Hz, 2H), 6.40 (t,J = 2.2 Hz, 1H), 3.83 (s, 6H), 2.3 (s, 3H).

[0089] (4) Synthesis of (E)-4-(3,5-dimethoxyphenylvinyl)phenol (4)

[0090] Compound 3 (213.4 mg, 0.72 mmol) was dissolved in methanol (13 mL), and sodium methoxide (5 mol / L, 0.012 mL) was slowly added and stirred at room temperature for 2 h. After the reaction was completed, 10 mL of water was added, and DCM was extracted (10 mL×2). The organic phases were combined and dried over anhydrous sodium sulfate. The crude product was recrystallized from methanol (MeOH) to obtain a white solid product 4 with a yield of 90%. 1 H-NMR (400MHz, CDCl3) δ 7.40 (d, J = 8.8 Hz, 2H), 6.96 (dd, J = 16.0 Hz, 52.4 Hz, 2H), 6.83 (d, J = 8.8 Hz, 2H), 6.65 (d, J = 2.4 Hz, 2H), 6.38 (t, J = 2.2 Hz, 1H), 4.79 (s, 1H), 3.83 (s, 6H).

[0091] (5) Synthesis of (E)-1-(4-(2-bromoethoxy)phenylvinyl)-3,5-dimethoxybenzene (5a)

[0092] Compound 4 (260.0 mg, 1.0 mmol), 1,2-dibromoethane (0.86 mL, 10.0 mmol), and potassium carbonate (552.8 mg, 4.0 mmol) were dissolved in 0.5 mL DMF and heated at 80 °C for 7 h. After the reaction was completed, 5 mL of water was added to quench the reaction, and the aqueous layer was extracted with ethyl acetate (EA) (10 mL × 3). The organic phases were combined, washed with saturated brine, dried over anhydrous Na2SO4, filtered, and the filtrate was concentrated under reduced pressure. The crude product was separated and purified by silica gel column chromatography (PE:EA = 30:1-10:1, v / v, volume ratio) to obtain a white solid product 5a with a yield of 79.3%. 1 H-NMR (400 MHz, CDCl3) δ 7.45 (d, J = 8.4Hz, 2H), 7.04 (d, J = 16.0 Hz, 1H), 6.90-7.06 (m, 3H), 6.65 (d, J = 2.0 Hz,2H), 6.38 (t, J = 2.2 Hz, 1H), 4.32 (t, J = 6.2 Hz, 2H), 3.83 (s, 6H), 3.65 (t, J = 6.2 Hz, 2H).

[0093] (6) Synthesis of tert-butyl (E)-4-(2-(4-(3,5-dimethoxyphenylvinyl)phenoxy)ethyl)piperazine-1-carboxylate (6a)

[0094] Compound 5a (100 mg, 0.275 mmol) was dissolved in 1 mL of acetonitrile, and 1-tert-butyloxycarbonylpiperazine (77.3 mg, 0.415 mmol), potassium iodide (22.9 mg, 0.138 mmol), and potassium carbonate (76.0 mg, 0.55 mmol) were added in sequence and refluxed at 80 °C for 6 h. After the reaction was completed, 5 mL of water was added, and the aqueous layer was extracted with EA (10 mL × 3). The organic phases were combined, washed with saturated brine, dried over anhydrous Na2SO4, filtered, and the filtrate was concentrated under reduced pressure. The crude product was chromatographed on a silica gel column (PE: EA = 10:1 to 1:1) to obtain a white solid product 6a with a yield of 98.7%. 1 H-NMR (400 MHz, CDCl3) δ 7.44 (d, J =8.8 Hz, 2H), 7.03 (d, J = 16.0 Hz, 1H), 6.66-6.92 (m, 3H), 6.65 (d, J = 2.0Hz, 2H), 6.38 (t, J = 2.0 Hz, 1H), 4.16 (s, 2H), 3.83 (s, 6H), 3.49 (s, 4H), 2.87 (s, 2H), 2.57 (s, 4H), 1.46 (s, 9H).

[0095] (7) Synthesis of 1-cinnamyl-4-(2-(4-((E)-3,5-dimethoxyphenylvinylphenoxy)ethyl)piperazine (7)

[0096] Compound 6a (99.6 mg, 0.213 mmol) was dissolved in 1 mL of anhydrous DCM under ice bath, and 0.2 mL of trifluoroacetic acid (TFA) solution was slowly added, and stirred at room temperature for 4 h. After the reaction was completed, the organic solvent and the remaining TFA were evaporated under reduced pressure. The obtained product was dissolved in 2 mL of acetonitrile, and the pH value of the reaction solution was adjusted to 8 with Et3N. Then, cinnamyl bromide (125.9 mg, 0.639 mmol) and anhydrous sodium carbonate (67.7 mg, 0.639 mmol) were added to the reaction solution, and stirred at room temperature overnight. After the reaction was completed, 5 mL of water was added, and the aqueous layer was extracted with EA (10 mL × 3). The organic phases were combined, washed with saturated brine, dried over anhydrous Na2SO4, filtered, and the filtrate was concentrated under reduced pressure. The crude product was chromatographed on a silica gel column (DCM: MeOH = 200:1~50:1, v / v, volume ratio) to obtain a white solid product 7 with a yield of 63.2%. 1H-NMR (400 MHz, CDCl3) δ 7.43 (d, J = 8.4 Hz, 2H), 7.38 (d, J =7.6 Hz, 2H), 7.31 (t, J = 7.4 Hz, 2H), 7.23 (t, J = 7.2 Hz, 1H), 7.03 (d, J =16.0 Hz, 1H), 6.88-6.92 (m, 3H), 6.65 (d, J = 2.4 Hz, 2H), 6.54 (d, J = 15.6Hz, 1H), 6.38 (t, J = 2.0 Hz, 1H), 6.26-6.33(m, 1H), 4.14 (t, J = 5.6 Hz,2H), 3.83 (s, 6H), 3.20 (d, J = 6.8 Hz, 2H), 2.86 (t, J = 5.8 Hz, 2H), 2.63-2.69 (m, 8H); 13 C-NMR (100 MHz, CDCl3) δ 161.1, 158.6, 139.8, 136.8, 133.8,130.2, 128.8, 128.7, 127.9, 127.8, 126.7, 126.5, 125.8, 114.9, 104.4, 99.7,66.0, 61.0, 57.2, 55.5, 53.4, 53.0; HRMS (ESI-TOF) m / z calcd. for C 31 H 37 N2O3 [M+H] + : 485.2804, found 485.2802. The H NMR spectrum of compound 7 is shown in Figure 1 , NMR carbon spectrum see Figure 2 .

[0097] (8) Synthesis of 1-cinnamyl-4-(2-(4-((E)-3,5-dimethoxyphenylvinylphenoxy)ethyl)piperazine hydrochloride (9)

[0098] Compound 7 (50 mg, 0.103 mmol) was dissolved in 5 mL of methanol, and a 2 M methanol solution of hydrogen chloride (250 μL, 0.5 mmol) was added. A white solid precipitated from the solution. The sealed reaction system was left at room temperature overnight and evaporated to dryness under reduced pressure to obtain a white solid product 9 with a yield of 100%. H NMR spectrum: Figure 3 .

[0099] Example 2 Synthesis of 1-cinnamyl-4-(4-((E)-3,5-dimethoxyphenylvinylphenoxy)butyl)piperazine (8) and its hydrochloride 10

[0100] As shown in the synthetic routes of the above compounds 7 to 10, the specific steps are as follows:

[0101] (1) Synthesis of (E)-1-(4-bromobutyloxy)phenylvinyl)-3,5-dimethoxybenzene (5b)

[0102] Using compound 4 (500 mg, 1.95 mmol) and 1,4-dibromobutane (2.3 mL, 19.5 mmol) as starting materials, the synthesis method was the same as compound 5a to obtain white solid product 5b with a yield of 85.2%. 1 H-NMR (400 MHz, CDCl3) δ7.44 (d, J = 8.8 Hz, 2H), 7.04 (d, J = 16.4 Hz, 1H), 6.87-6.92 (m, 3H), 6.65(d, J = 2.0 Hz, 2H), 6.38 (t, J = 2.2 Hz, 1H), 4.02 (t, J = 6.0 Hz, 2H), 3.83 (s, 6H), 3.50 (t, J = 6.6 Hz, 2H), 2.05-2.10 (m, 2H), 1.94-1.99 (m, 2H).

[0103] (2) Synthesis of tert-butyl (E)-4-(4-(4-(3,5-dimethoxyphenylvinyl)phenoxy)butyl)piperazine-1-carboxylate (6b)

[0104] Using compound 5b (130 mg, 0.332 mmol) and 1-tert-butyloxycarbonylpiperazine (92.8 mg, 0.50 mmol) as starting materials, the synthesis method was the same as compound 6a to obtain colorless transparent oily product 6b with a yield of 92.4%. 1H-NMR (400 MHz, CDCl3) δ 7.43 (d, J = 8.8 Hz, 2H), 7.03 (d, J = 16.0 Hz, 1H), 6.86-6.91 (m,3H), 6.64 (d, J = 2.0 Hz, 2H), 6.37 (t, J = 2.2 Hz, 1H), 3.99 (t, J = 6.2 Hz,2H), 3.82 (s, 6H), 3.43 (t, J = 4.8 Hz, 4H), 2.38-2.43 (m, 6H), 1.80-1.83 (m,2H), 1.64-1.71 (m, 2H), 1.46 (s, 9H).

[0105] (3) Synthesis of 1-cinnamyl-4-(4-((E)-3,5-dimethoxyphenylvinylphenoxy)butyl)piperazine (8)

[0106] Compound 6b (152.4 mg, 0.307 mmol) was synthesized in the same manner as compound 7 to obtain light yellow transparent oily product 8 with a yield of 54.7%. 1 H-NMR (400 MHz, CDCl3) δ 7.43 (d, J = 8.4 Hz, 2H), 7.38 (d, J= 7.6 Hz, 2H), 7.32 (t, J = 7.2 Hz, 2H), 7.24-7.28 (m, 1H), 7.03 (d, J = 16.4Hz, 1H), 6.85-6.92 (m, 3H), 6.65 (d, J = 2.0 Hz, 2H), 6.58 (d, J = 15.6 Hz, 2H), 6.38 (s, 1H), 6.23-6.31 (m, 1H), 4.00 (t, J = 5.4 Hz, 2H), 3.83 (s, 6H), 3.34 (d, J = 6.8 Hz, 2H), 2.73-2.91 (m, 10H), 1.84-1.86 (m, 4H); 13C-NMR (100MHz, CDCl3) δ 161.0, 158.6, 139.7, 136.1, 135.6, 130.0, 128.7, 128.1, 127.9,126.6, 126.6, 123.1, 114.7, 104.3, 99.7, 67.3, 60.2, 57.4, 55.4, 51.8, 51.0,26.9, 22.2; HRMS (ESI-TOF) m / z calcd. for C 33 H 41 N2O3 [M+H] + : 513.3112, found513.3107. The H NMR spectrum of compound 8 is shown in Figure 4 , NMR carbon spectrum see Figure 5 .

[0107] (4) Synthesis of 1-cinnamyl-4-(4-((E)-3,5-dimethoxyphenylvinylphenoxy)butyl)piperazine hydrochloride (10)

[0108] Compound 8 (50 mg) was dissolved in 5 mL of methanol, and a 2 M methanol solution of hydrogen chloride (250 μL, 0.5 mmol) was added. A light yellow solid precipitated from the solution. The sealed reaction system was left at room temperature overnight and evaporated to dryness under reduced pressure to obtain a light yellow solid product 10 with a yield of 100%. The H NMR spectrum was the same as that of compound 8, but it was insoluble in neutral solvents such as dichloromethane and soluble in methanol.

[0109] The synthetic routes of compounds 13 and 14 are as follows:

[0110]

[0111] Example 3 Synthesis of 1-cinnamoyl-4-(2-(4-((E)-3,5-dimethoxyphenylvinyl)phenoxy)ethoxy)ethyl)piperazine (13) and its hydrochloride 14

[0112] (1) Synthesis of (E)-1-(4-(2-(2-bromoethoxy)ethoxy)phenylvinyl)-3,5-dimethoxybenzene (11)

[0113] The synthesis method of compound 11 is the same as compound 5a, and the intermediate 4, namely compound 4, is reacted with 2, 2'-dibromodiethyl ether to obtain white solid compound 11 with a yield of 86.2%. 1H-NMR (400 MHz, CDCl3) δ 7.44 (d, J = 8.8 Hz, 2H), 7.04 (d, J = 16 Hz, 1H), 6.89-6.93 (m, 6H), 6.65 (d, J = 2.4 Hz, 2H), 6.38 (t, J = 3.0 Hz, 1H), 4.16 (m, 2H), 3.87-3.91 (m, 4H), 3.83 (s, 1H), 3.50 (t, J = 6.4 Hz, 1H).

[0114] (2) Synthesis of 1-cinnamoyl-4-(2-(4-((E)-3,5-dimethoxyphenylvinyl)phenoxy)ethoxy)ethyl)piperazine (13)

[0115] The synthesis method of compound 13 is the same as that of compound 7. Compound 11 is connected with N-tert-butyloxycarbonylpiperazine to obtain 12 with a yield of 78.9%. After removing the tert-butyl ester, it is alkylated with cinnamyl bromide to obtain compound 13 as a white solid with a yield of 74.1%. 1 H-NMR (400 MHz, CDCl3) δ 7.41 (d, J = 8.4 Hz, 2H), 7.36 (d, J = 7.6 Hz, 2H), 7.30 (t, J = 7.2 Hz, 2H), 7.23 (m, 1H), 7.02 (d, J = 16.0 Hz, 1H), 6.89(m, 3H), 6.64 (s, 2H), 6.52 (d, J = 15.6 Hz, 1H), 6.37 (s, 1H), 6.27 (m, 1H), 4.17 (m, 2H), 3.81 (s, 8H), 3.71 (m, 2H), 3.17 (m, 2H), 2.65 (m, 10H); 13 C-NMR (100 MHz, CDCl3) δ 161.0, 158.6, 139.7, 136.8, 133.5, 130.1, 128.7, 128.6,127.8, 127.6, 126.6, 126.4, 125.9, 114.8, 104.3, 99.6, 69.5, 69.0, 67.4,60.9, 57.7, 55.3, 53.4, 52.8. The H NMR spectrum of compound 13 is shown in Figure 6 , NMR carbon spectrum see Figure 7 .

[0116] (3) Synthesis of 1-cinnamoyl-4-(2-(4-((E)-3,5-dimethoxyphenylvinyl)phenoxy)ethoxy)ethyl)piperazine hydrochloride (14)

[0117] Compound 13 (50 mg) was dissolved in 5 mL methanol, and hydrogen chloride gas was slowly introduced. A white solid precipitated from the solution until the solid stopped increasing. The sealed reaction system was left at room temperature overnight and evaporated to dryness under reduced pressure to obtain a white solid product 13 with a yield of 100%. The H NMR spectrum was the same as that of compound 14.

[0118] Synthesis routes of compound 20 and compound 21:

[0119]

[0120] Example 4 Synthesis of 1-cinnamoyl-4-(2-(2-(2-(4-((E)-3,5-dimethoxyphenylvinyl)phenoxy)ethoxy)ethoxy)ethyl)piperazine (20) and its hydrochloride 21

[0121] (1) Synthesis of 2-(2-(2-hydroxyethoxy)ethoxy)ethyl 4-methylbenzenesulfonate (15)

[0122] At 0 °C, triethylene glycol (500 mg, 3.33 mmol), anhydrous DCM (6 mL), 4-toluenesulfonyl chloride (508 mg, 2.66 mmol) and Et3N (0.5 mL) were added to a 25 mL round-bottom flask in sequence. After 0.5 h, the mixture was warmed to room temperature and stirred for 3 h. After the reaction was completed, the crude product was separated by silica gel column chromatography (DCM: MeOH = 200:1 ~ 20:1, v / v, volume ratio) to obtain a yellow oily product 15 with a yield of 58.5%. 1 H-NMR (400 MHz, CDCl3) δ 7.73 (d, J = 8.4 Hz, 2H), 7.29 (d,J = 8.0 Hz, 2H), 4.10 (t, J = 4.6 Hz, 2H), 3.61-3.64 (m, 4H), 3.48-3.53 (m,6H), 2.87 (s, 1H), 2.38 (s, 3H).

[0123] (2) Synthesis of 2-(2-(2-iodoethoxy)ethoxy)ethane-1-ol (16)

[0124] Compound 15 (495.5 mg, 1.56 mmol) was dissolved in 6 mL of acetone, and sodium iodide (1.169 g, 7.8 mmol) was added to reflux at 55 °C for 3 h. After the reaction was completed, the post-treatment was the same as compound 15 to obtain a yellow oily product 16 with a yield of 78.7%. 1 H-NMR (400 MHz, CDCl3) δ 3.73-3.77 (m, 4H), 3.67-3.69 (m, 4H), 3.60-3.63 (m, 2H), 3.26 (t, J = 6.8 Hz, 2H), 2.43 (s, 1H).

[0125] (3) Synthesis of (E)-2-(2-(4-(3,5-dimethoxyphenylvinyl)phenoxy)ethoxy)ethoxy)ethoxy)ethane-1-ol (17)

[0126] Compound 4 (300.0 mg, 1.17 mmol), compound 16 (365.1 mg, 1.404 mmol), and anhydrous potassium carbonate (485.1 mg, 3.51 mmol) were dissolved in 1 mL DMF and heated at 110 °C for 2 h. After the reaction was completed, the post-treatment was the same as compound 5a to obtain a white solid product 17 with a yield of 100.0%. 1 H-NMR (400 MHz, CDCl3) δ 7.43(d, J = 8.4 Hz, 2H), 7.03 (d, J = 16.0 Hz, 1H), 6.90 (t, J = 8.4 Hz, 3H), 6.65 (t, J = 2.4 Hz, 2H), 6.38(t, J = 2.2 Hz, 1H), 4.15 (t, J = 4.8 Hz, 2H), 3.87 (t, J = 4.8 Hz, 2H), 3.82 (s, 6H), 3.69-3.75 (m, 6H), 3.62 (t, J = 4.4Hz, 2H), 2.43 (t, J = 2.8 Hz, 1H); 13 C-NMR (100 MHz, CDCl3) δ 161.1, 158.6,139.8, 130.3, 128.8, 127.9, 126.8, 115.0, 104.5, 99.8, 72.6, 71.0, 70.5,69.8, 67.5, 61.9, 55.5.

[0127] (4) Synthesis of ethyl (E)-2-(2-(4-(3,5-dimethoxyphenylvinyl)phenoxy)ethoxy)ethoxy)4-methylbenzenesulfonate (18)

[0128] Compound 17 (454.0 mg, 1.17 mmol), 4-toluenesulfonyl chloride (335.6 mg, 1.404 mmol) and Et3N (0.49 mL, 3.51 mmol) were used as starting materials. The synthesis method was the same as compound 15 to obtain a milky white oily product compound 18 with a yield of 95.9%. 1 H-NMR (400 MHz, CDCl3) δ 7.80 (d, J = 8.4 Hz, 2H), 7.43 (d, J =8.8 Hz, 2H), 7.32 (d, J = 8.0 Hz, 2H), 7.03 (m, d, J = 16.0 Hz, 1H), 6.88-6.92 (m, 3H), 6.65 (d, J = 2.4 Hz, 2H), 6.38 (t, J = 2.2 Hz, 1H), 4.12-4.17(m, 4H), 3.83-3.84 (m, 8H), 3.63-3,71 (m, 4H), 3.62 (t, J = 3.0 Hz, 2H), 2.43(s, 3H).

[0129] (5) Synthesis of (E)-1-(4-(2-(2-(2-iodoethoxy)ethoxy)ethoxy)phenylvinyl)-3,5-dimethoxybenzene (19)

[0130] Compound 18 (608.7 mg, 1.12 mmol) and sodium iodide (839.4 mg, 5.6 mmol) were used as starting materials. The synthesis method was the same as compound 16 to obtain a yellow oily product, compound 19, with a yield of 95.0%. 1H-NMR (400 MHz, CDCl3) δ 7.44 (d, J = 8.8 Hz, 2H), 7.04 (d, J = 16.0 Hz, 1H), 6.88-6.93 (m,3H), 6.65 (d, J = 2.4 Hz, 2H), 6.38 (t, J = 2.2 Hz, 1H), 4.16 (t, J = 4.8 Hz,2H), 3.88 (m, t, J = 4.8 Hz, 2H), 3.83 (s, 6H), 3.73-3.79 (m, 4H), 3.68-3.71(m, 2H), 3.27 ( t, J = 6.8 Hz, 2H).

[0131] (6) Synthesis of 1-cinnamyl-4-(2-(2-(4-((E)-3,5-dimethoxyphenylvinyl)phenoxy)ethoxy)ethoxy)ethoxy)ethyl)piperazine (20)

[0132] Compound 19 (200 mg, 0.4 mmol) was dissolved in 1 mL of acetonitrile, and compound N-Boc-piperazine (1.2 mmol) and anhydrous sodium carbonate (127.2 mg, 1.2 mmol) were added in sequence, and the mixture was heated to reflux at 80 °C for 3 h. After the reaction was completed, the post-treatment operation was the same as compound 6a, and the next step was the same as compound 7. The crude product was purified by silica gel column chromatography (DCM: MeOH = 100:1-30:1, v / v, volume ratio) to obtain yellow oily product compound 20 with a yield of 86.0%. 1H-NMR (400 MHz, CDCl3) δ 7.41(d, J = 8.8 Hz, 2H), 7.35 (d, J = 7.2 Hz, 2H), , 7.25-7.30(m, 2H), 7.21 (t, J= 7.2 Hz, 1H), 7.02 (d, J = 16.0 Hz, 1H), 6.86-6.90 (m, 3H), 6.64 (d, J = 2.0Hz, 2H), 6.50 (d, J = 15.6 Hz, 1H), 6.37 (s, 1H), 6.26-6.30 (m, 1H), 4.12 (t,J = 4.8 Hz, 2H), 3.84 (t, J = 4.8 Hz, 2H), 3.8 (s, 6H), 3.70-3.72 (m,2H),3.63 (t, J = 5.0 Hz, 4H), 3.14 (t, J = 6.8 Hz, 2H), 2.62 (t, J = 5.8 Hz, 2H), 2.57 (m, 8H). 13 C-NMR (100 MHz, CDCl3) δ 161.0, 158.6, 139.7, 136.9, 133.3,130.1, 128.7, 128.6, 127.8, 127.6, 126.7, 126.4, 114.9, 104.4, 99.7, 70.9,70.4, 69.7, 68.8, 67.5, 61.0, 57.8, 55.4, 53.5, 53.0; HRMS (ESI-TOF) m / zcalcd. for C 35 H 44 N2O5Na [M+Na] + : 595.3148, found 595.3142. H NMR spectrum: Figure 8 , NMR carbon spectrum see Fig. 9 .

[0133] (7) Synthesis of 1-cinnamyl-4-(2-(2-(4-((E)-3,5-dimethoxyphenylvinyl)phenoxy)ethoxy)ethoxy)ethoxy)ethyl)piperazine hydrochloride (21)

[0134] Take 20 mg of compound 20, dissolve it in methanol, add 2 M hydrogen chloride methanol solution (100 μL, 0.2 mmol), and a white solid gradually appears. Leave it at room temperature overnight, filter out the white precipitate, and obtain compound 21 with a yield of 100%. NMR is shown in Fig.10.

[0135] Compound 7 was demethylated by BBr3 ether solution at low temperature to obtain compound 22.

[0136]

[0137] Example 5 Synthesis of 5-((E)-4-(2-(4-cinnamylpiperazin-1-yl)ethoxy)phenylvinyl)benzene-1,3-diol (22)

[0138] Compound 7 (48 mg, 0.1 mmol) was dissolved in ether and placed in a low-temperature bath at -20 o C, add 0.25 mmol of boron tribromide (BBr3) ether solution and react for 2 h to obtain compound 22 as a white solid with a yield of 46%. 1 H-NMR(400 MHz, CDCl3) δ 7.42 (d, J = 8.4 Hz, 2H), 7.36 (d, J = 7.6 Hz, 2H), 7.31(t, J = 7.4 Hz, 2H), 7.23 (t, J = 7.2 Hz, 1H), 7.03 (d, J = 16.0 Hz, 1H), 6.90 (m, 3H), 6.65 (d, J = 2.4 Hz, 2H), 6.54 (d, J = 16.0 Hz, 1H), 6.38 (t, J= 2.0 Hz, 1H), 6.30 (m, 1H), 4.14 (t, J = 5.6 Hz, 2H), 3.20 (d, J = 6.8 Hz,2H), 2.86 (t, J = 5.8 Hz, 2H), 2.65 (m, 8H).

[0139] Example 6 Evaluation of the in vitro inhibitory activity of cinnamylpiperazine-resveratrol conjugate on tumor cell proliferation

[0140] The in vitro cytotoxic activity evaluation method is as follows:

[0141] (1) HepG2, SW480, HCT116, MDA-MB-231, 4T1 and MCF-7 cells were cultured at 5 × 10 4 The cells were inoculated at a density of 100 μL / mL in a 96-well plate.

[0142] (2) Place the inoculated cells in a 37°C, 5% CO2 incubator for 24 hours, add different concentrations (100, 10, 1, 0.1, 0.01, 0 μM) of cinnamyl piperazine conjugate and positive control camptothecin (CPT), add 0.5 μL to each well, and set up three replicate wells for each drug concentration. After the addition of drugs, place the cells in an incubator and continue to culture for 48 hours.

[0143] (3) Add 20 μL of methyl tetrazolyl blue (MTT, 5 mg / mL) solution to all wells in a dark environment. Place the 96-well plate in an incubator. After 4 hours of exposure to MTT solution, remove the supernatant from the wells and add 100 μL of DMSO to each well.

[0144] (4) Incubate at 37°C for 10 min. Detect the OD value of each well using an ELISA reader at dual wavelengths of 492 nm and 630 nm, and calculate the cell survival rate:

[0145] Cell survival rate (%) = (OD of experimental group - OD of blank group) / (OD of control group - OD of blank group) × 100%

[0146] The in vitro antitumor activity results of cinnamyl piperazine-resveratrol conjugates are shown in Table 1. All synthesized conjugates have certain antitumor activity, especially good inhibitory activity against HepG2 liver cancer cells, among which compound 7 has the highest inhibitory activity against HepG2 (IC 50 =0.19 μM). In addition, compound 20 (MDA-MB-231: IC 50 =0.41 μM; 4T1:IC 50 =0.14 μM) and its hydrochloride 21 (MDA-MB-231: IC 50 =0.35 μM; 4T1:IC 50 =0.22 μM) significantly inhibited the proliferation activity of triple-negative breast cancer, even compared with the positive control camptothecin (MDA-MB-231: IC 50 =0.13 μM; 4T1:IC 50 =0.20 μM). Therefore, the cinnamyl piperazine fragment was coupled with the resveratrol derivative via a connecting bridge, and the resulting compound had significant antitumor activity.

[0147]

[0148] Note: a The experimental data are the mean values ​​of three independent parallel experiments; b CPT (Camptothecin) is the positive control drug; * is the hydrochloride;

[0149] The structural formula of the cinnamyl piperazine resveratrol conjugate in Table 1 is as follows:

[0150] ;

[0151] R1 in Table 1 is R1 in the above structural formula, R2 in Table 1 is R2 in the above structural formula, and the connecting bridge in Table 1 is the linker in the above structural formula.

[0152] Example 7 Effects of Compound 7 on the lateral migration ability of HepG2 cells and Compound 21 on MDA-MB-231 cells

[0153] Representative compounds 7 and 21 were selected to evaluate the effects on the migration ability of liver cancer HepG2 and triple-negative breast cancer MDA-MB-231 cells, respectively. The cell scratch assay was used to evaluate the effect of the compounds on the lateral migration ability of cancer cells, and the method was as follows:

[0154] (1) Place the scratch professional plug-in into the prepared 12-well plate in advance. Select HepG2 and MDA-MB-231 cells, digest them, count and dilute them, and use complete medium containing 10% FBS at 2×10 5 The cells were inoculated into the insert at a density of 100 μL / mL. 70 μL of culture medium was added to each well, and the 12-well plate was placed in a 37°C, 5% CO2 incubator for 24 h.

[0155] (2) After 24 h, observe the cell status under a microscope. When the cells grow to about 90%, use tweezers to gently pull out the plug, add phosphate buffered saline (1×PBS) to wash the cells three times, add 1 mL of IMDM culture medium containing 1% FBS, and continue to culture in a 37°C, 5% CO2 incubator for 4 h.

[0156] (3) Compound 7 was added to culture cells at concentrations of 1, 0.2, 0.04, 0.008, and 0 μM, and compound 21 was added to culture cells at concentrations of 0.4, 0.04, and 0 μM, respectively, with a dosing volume of 5 μL per well.

[0157] (4) After drug administration, the changes in the scratch width in each well of the 12-well plate were observed under a microscope at 0, 12, 24, and 48 h, and photos were taken at 0, 12, 24, and 48 h. The scratch width was recorded and the lateral migration ability of HepG2 and MDA-MB-231 cells on the culture plate was determined by the width changes.

[0158] like Fig.11The results of the photos taken at 0, 12, 24 and 48 h are shown in the figure. Compound 7 has a significant effect on the lateral migration of HepG2 cells at 24 and 48 h. The migration distance was further statistically analyzed. Fig.12 Compound 7 had a significant inhibitory effect on the lateral migration of HepG2 cells at 24 and 48 h. At 24 h, the migration distance of compound 7 at 1 μM concentration was 393.75±43.39 μm, at 0.2 μM concentration was 408.43±38.99 μm, and at 0.04 μM concentration was 428.00±78.27 μm, which was significantly different from the control DMSO group (517.67±37.60 μm) in the solvent (***P<0.001). At 48 h, the migration distance of compound 7 at 1 μM concentration was 445.13±30.87 μm, at 0.2 μM concentration was 677.57±50.72 μm, and at 0.04 μM concentration was 699.80±104.52 μm. μm, which was significantly different from the control DMSO group (835.83±363.92 μm) in the solvent (***P<0.001 and **P<0.01). This indicates that compound 7 has a good ability to inhibit the migration of HepG2 cells.

[0159] In addition, if Fig.13 As shown, compound 21 had a significant effect on the lateral migration of MDA-MB-231 cells at 24 and 48 h. Further statistical analysis of the migration distance was performed, and the results were shown in Fig.14 Compound 21 had a significant inhibitory effect on the lateral migration of MDA-MB-231 cells at 24 and 48 h. At 24 h, the migration distance of compound 21 at 0.04 μM concentration was 128.50±2.12 μm, and the migration distance at 0.4 μM concentration was 91.50±6.36 μm, which was significantly different from the control DMSO group in the solvent (197.50±3.54 μm) (***P<0.001); at 48 h, the migration distance of compound 21 at 0.04 μM concentration was 269.00±4.24 μm, and the migration distance at 0.4 μM concentration was 186.00±21.21 μm, which was significantly different from the control DMSO group in the solvent (478.00±8.48 μm) (***P<0.001). This shows that compound 21 has a good ability to inhibit the migration of triple-negative breast cancer cells.

[0160] Example 8 Effects of Compound 7 on the Longitudinal Migration of HepG2 Cells and Compound 21 on the Longitudinal Migration of MDA-MB-231 Cells

[0161] The effects of compound 7 on the longitudinal migration of HepG2 cells and compound 21 on MDA-MB-231 cells were evaluated using the Transwell assay as follows:

[0162] (1) The Transwell chamber was irradiated for 30 min in a clean bench. 600 μL of culture medium containing 20% ​​serum was added to the lower chamber of the chamber. Cells in the logarithmic growth phase were taken and serum-free cell solution was used at a concentration of 1×10 5 The density of 100 μL / mL was evenly spread in the upper chamber of the chamber, with 200 μL in each upper chamber.

[0163] (2) After the cells adhered to the wall, 1 μL of different concentrations of compound 7 (0.04, 0.2, 1 μM) and compound 22 (0.04, 0.4 μM) were added.

[0164] (3) After 24 h, discard the culture medium in the upper and lower chambers and wash the inside and outside of the chambers with phosphate buffered saline (1× PBS) for a total of three times.

[0165] (4) Fix with 4% paraformaldehyde at room temperature for 30 min, discard the 4% paraformaldehyde, and wash twice with 1× PBS. Add 200 μL of 0.1% crystal violet solution to the upper chamber and 600 μL of 0.1% crystal violet solution to the lower chamber and stain for 10 min.

[0166] (5) After staining, wash the chamber with PBS, dry it, and then take a photo.

[0167] like Fig.15 As shown in the figure, at the tested concentration, compound 7 could significantly inhibit the longitudinal migration of HepG2 cells. The number of migrating cells was further statistically analyzed (e.g. Fig.16 As shown in the figure, the number of migrated cells of compound 7 at concentrations of 1 μM, 0.2 μM and 0.04 μM were 28, 49 and 281, respectively, which were significantly different from those of the solvent control DMSO group (541) (***P<0.001), indicating that compound 7 could significantly inhibit the longitudinal migration of HepG2 cells.

[0168] like Fig.17 As shown, compound 21 can significantly inhibit the longitudinal migration of triple-negative breast cancer cells. The number of migrating cells was further statistically analyzed (e.g. Fig.18 As shown in the figure, the number of migrating cells of compound 21 at 0.04 μM and 0.4 μM concentrations were 276 and 80, respectively. The number of migrating cells at 0.4 μM concentration was significantly different from that of the solvent control DMSO group (576) (**P<0.01), indicating that compound 21 can significantly inhibit the longitudinal migration of MDA-MB-231 cells.

[0169] In summary, the present invention synthesized a series of cinnamyl piperazine resveratrol derivatives for the first time. The synthesized compounds have a significant inhibitory effect on the proliferation of various tumor cells, among which compound 7 and compound 21 showed good inhibitory activity on liver cancer cells and triple-negative breast cancer cells, respectively. In particular, compound 21 has an activity comparable to that of the clinical drug camptothecin on triple-negative breast cancer. In addition, representative compounds 7 and 21 were selected to affect the migration ability of tumor cells, and the effect was obvious at the tested concentration. The present invention discovered for the first time that cinnamyl piperazine resveratrol conjugates have good activity in inhibiting the proliferation of various cancer cells, and the synthesized hydrochloride has good solubility and good drug potential. Therefore, this type of compound has good application prospects in anti-tumor.

[0170] Although the embodiments of the present invention are disclosed for illustrative purposes, those skilled in the art will appreciate that various substitutions, changes and modifications are possible without departing from the spirit and scope of the present invention and the appended claims. Therefore, the scope of the present invention is not limited to the contents disclosed in the embodiments.

Claims

1. An anti-tumor cinnamylpiperazine-resveratrol conjugate or a pharmaceutically acceptable salt thereof, characterized in that: Its general structure is as follows: ； Wherein, the linker is a straight or branched alkyl or ether containing 1 to 6 atoms, R1 and R2 are the same or different, and R1 and R2 are hydrogen, hydroxyl, CH3O-, CH3 (CH2) n O-, where n=1~3.

2. The anti-tumor cinnamyl piperazine-resveratrol conjugate or a pharmaceutically acceptable salt thereof according to claim 1, characterized in that: The structural formula of the anti-tumor cinnamyl piperazine resveratrol conjugate or a pharmaceutically acceptable salt thereof is one of the following:

3. The method for preparing the anti-tumor cinnamylpiperazine-resveratrol conjugate or a pharmaceutically acceptable salt thereof according to claim 1 or 2, characterized in that: The steps include: First, 3, 5-dimethoxybenzaldehyde is subjected to a witting reaction to obtain compound 1; then 4-iodophenol is used as a raw material to connect acetyl chloride through an acylation reaction to obtain compound 2, compound 1 is connected through a Heck reaction to obtain compound 3, and compound 4 is hydrolyzed, and then different connecting bridges are connected through a Williams ether synthesis method to obtain compound 5a or compound 5b, and then a nitrogen-containing compound is connected to obtain compound 6a or compound 6b, after removing the protecting group, an alkylation reaction is carried out with cinnamyl bromide to obtain compound 7 or compound 8, and hydrogen chloride gas is introduced into a methanol or acetone solution to obtain the corresponding hydrochloride 9 or 10. The reaction route is as follows: ; Alternatively, intermediate 4 is used as a raw material, connected with 2, 2'-dibromodiethyl ether to obtain compound 11, connected with N-Boc-piperazine to obtain compound 12, and finally after removing tert-butyl ester, connected with cinnamyl bromide to obtain compound 13, and further introduced with acidic gas to obtain its salt compound 14. The reaction route is: ; Alternatively, triethylene glycol is activated to obtain compound 15, which is then subjected to nucleophilic substitution reaction with sodium iodide to obtain compound 16, which is then connected to compound 4 to obtain compound 17, and compound 17 is activated by hydroxyl to obtain compounds 18 and 19, which are then connected to piperazine and cinnamyl to obtain compound 20, and further acidic gas is introduced to obtain its hydrochloride 21. The reaction route is: ; Compound 7 was demethylated by BBr3 ether solution at low temperature to obtain resveratrol derivative 22; .

4. Use of the anti-tumor cinnamylpiperazine-resveratrol conjugate or a pharmaceutically acceptable salt thereof as claimed in claim 1 or 2 in the preparation of a drug for treating liver cancer.

5. Use of the anti-tumor cinnamylpiperazine-resveratrol conjugate or a pharmaceutically acceptable salt thereof as claimed in claim 1 or 2 in the preparation of a drug for treating colon cancer.

6. Use of the anti-tumor cinnamylpiperazine-resveratrol conjugate or a pharmaceutically acceptable salt thereof as claimed in claim 1 or 2 in the preparation of a drug for treating breast cancer.

7. Use of the anti-tumor cinnamylpiperazine-resveratrol conjugate or a pharmaceutically acceptable salt thereof as claimed in claim 1 or 2 in the preparation of a drug for treating triple-negative breast cancer.

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