Derivatives of norzelaminaldehyde and their application in the preparation of anticancer drugs

Abstract

The present invention discloses norzelaminaldehyde derivatives and their application in the preparation of anticancer drugs, belonging to the field of pharmaceutical preparation. The norzelaminaldehyde derivatives are used to prepare drugs for non-small cell lung cancer, colon cancer, and cervical cancer, especially non-small cell lung cancer A549, colon cancer HCT116, and cervical cancer HeLa, and the drugs can be selected from at least one of tablets, capsules, granules, drop pills, suspensions, syrups, enteric preparations, emulsion suspensions, and injections. In the present invention, norzelaminaldehyde is a natural product with little toxicity and side effects, high bioavailability and clinical use value, and provides a candidate lead compound for the research and development of new inhibitors of anticancer drugs. At the same time, it has potential application value in the fields of targeted drugs, probe design, and fluorescent biomarkers for non-small cell lung cancer.

Description

Technical Field

[0001] The invention belongs to the field of medicine preparation, and particularly relates to norzelaminaldehyde derivatives and applications thereof in the preparation of anticancer drugs. Background Art

[0002] Malignant tumors are a type of disease that threatens human health and life. Currently, the treatment of malignant tumors often adopts a combination of surgery, radiotherapy, and chemotherapy. Chemotherapy is a systemic treatment that can eliminate cancer cells that have metastasized to distant locations, so it plays an important role in comprehensive treatment. However, clinically, chemotherapy drugs have problems with efficacy, side effects, and drug resistance of tumor cells. Therefore, it is still urgent to find highly effective and low-toxic anti-tumor drugs from different angles.

[0003] Natural products are an important source of drug lead compounds and functional organic molecules. Screening drug lead compounds from natural products has always been an effective way to develop new drugs. Natural products have always been an important source of drug research and development due to their wide range of biological activities and low toxic side effects. Therefore, the active ingredients in traditional Chinese medicine have become a current research hotspot. Among them, norzelaminaldehyde was isolated from the roots of Tripterygium wilfordii and has multiple biological activities, including anti-tumor, anti-inflammatory, and immunosuppressive effects. Among them, the antitumor activity of norzelaminaldehyde has been widely studied. Norzelaminaldehyde exhibits cytotoxicity against a variety of human cancer cell lines by inducing cell apoptosis. However, the antitumor efficacy (A549: 57.68±0.57μM, HCT116: 26.19±2.43μM, HeLa: 48.61±2.95μM) and physicochemical properties of norzelaminaldehyde need to be improved. By modifying C-2, C-3, C-4 and C-29 of demethylzelaminaldehyde and designing different substituents at different positions, the CCK8 method was used to measure the inhibitory effects of the compounds on three cancer cells, A549, HCT116 and HeLa, which can provide a good starting point for the development of new anti-tumor drugs. Summary of the invention

[0004] In view of this, the main purpose of the present invention is to provide a norzelaminaldehyde derivative.

[0005] Another object of the present invention is to provide the use of the above-mentioned nor-zearalenaldehyde derivatives in the preparation of anticancer drugs, and the nor-zearalenaldehyde derivatives have obvious anti-tumor cell activity.

[0006] The above-mentioned object of the present invention is achieved by the following technical solutions:

[0007] The present invention provides a norzelaminaldehyde derivative having a general structure as shown in Formula 1a or 1b:

[0008]

[0009] In formula 1a and 1b, R1, R2, R3, and R4 are respectively shown in the following table:

[0010]

[0011]

[0012] The present invention also provides a method for preparing the above-mentioned norzelaminaldehyde derivative, wherein the synthetic route is selected from any one of Formulas 2, 3 and 4:

[0013]

[0014]

[0015] The present invention also provides the use of the above-mentioned norzelaminaldehyde derivative or its pharmaceutically acceptable salt in the preparation of anticancer drugs.

[0016] Preferably, the cancer includes but is not limited to non-small cell lung cancer, colon cancer and cervical cancer.

[0017] The present invention provides an anti-cancer drug, which comprises a nor-zearalenaldehyde derivative or a pharmaceutically acceptable solvate thereof. The nor-zearalenaldehyde derivative has a structural formula as described above, has a significant inhibitory effect on cancer cells, and is a new type of anti-cancer drug.

[0018] Preferably, the weight percentage of the norzelaminaldehyde derivative or its pharmaceutically acceptable solvate in the drug is 5% to 100%.

[0019] Preferably, the medicine further comprises a pharmaceutically acceptable carrier or diluent.

[0020] "Pharmaceutically acceptable carrier" refers to inactive ingredients in drugs, including but not limited to calcium carbonate, calcium phosphate, various sugars such as lactose, mannitol, starch, cyclodextrin, magnesium stearate, cellulose, magnesium carbonate, acrylic acid polymers or methacrylic acid polymers, gelatin, water, polyethylene glycol, propylene glycol, ethylene glycol, castor oil or hydrogenated castor oil or polyethoxylated hydrogenated castor oil, sesame oil, corn oil, and peanut oil.

[0021] "Pharmaceutically acceptable diluents" include, but are not limited to, starch (such as corn starch, wheat starch, potato starch, etc.), lactose, dextrin, sucrose, pregelatinized starch, microcrystalline cellulose, inorganic salts (such as calcium hydrogen phosphate, calcium sulfate, residual calcium acid, etc.) and mannitol.

[0022] Preferably, the dosage form of the drug is selected from at least one of tablets, capsules, granules, pellets, suspensions, syrups, enteric-coated preparations, emulsion suspensions and injections.

[0023] The present invention proves through in vitro experiments that norzelaminaldehyde and its derivatives have strong activity against non-small cell lung cancer cell line A549, colon cancer HCT116, and cervical cancer HeLa, and can be used as a leading compound for treating cancer to prepare cancer drugs.

[0024] The present invention also provides the use of norzelaminaldehyde and its derivatives or pharmaceutically acceptable salts thereof in the preparation of A549, HCT116 and HeLa cell proliferation inhibitors.

[0025] Compared with the prior art, the present invention has the following beneficial effects:

[0026] (1) The present invention confirms through in vitro experiments that the nor-zearalenaldehyde derivatives have a significant inhibitory effect on the growth and cell activity of A549, HCT116, and HeLa cells, providing candidate lead compounds for the development of anticancer drugs.

[0027] (2) The norzelaminaldehyde derivatives of the present invention not only have the potential to be developed into a new generation of anticancer drugs, but also have potential application value in the fields of cancer targeted drugs, probe design and fluorescent biomarkers. BRIEF DESCRIPTION OF THE DRAWINGS

[0028] Figure 1 The inhibitory effect of compound 7 at different concentrations on the A549 cell cycle in Example 15 was detected.

[0029] Figure 2 The inhibitory effect of compound 7 at different concentrations on apoptosis of A549 cells in Example 16 was detected.

[0030] Figure 3 This is a graph showing the morphological changes of A549 cells after apoptosis at the same time under different concentrations of compound 7 in Example 17. DETAILED DESCRIPTION

[0031] In order to make the purpose, technical solution and advantages of the embodiment of the present invention clearer, the technical solution of the embodiment of the present invention is clearly and completely described below in conjunction with the accompanying drawings of the embodiment of the present invention. Obviously, the described embodiment is a part of the embodiment of the present invention, not all of the embodiments. Based on the described embodiment of the present invention, all other embodiments obtained by ordinary technicians in this field without creative work belong to the scope of protection of the present invention.

[0032] Unless otherwise specified in the following examples, all reagents and materials used were commercially available.

[0033] In the following examples, the norzelaminaldehyde derivatives have a general structure as shown in Formula 1a or 1b:

[0034]

[0035] In formula 1a and 1b, R1, R2, R3, and R4 are respectively shown in the following table:

[0036]

[0037]

[0038] There is no special requirement for the preparation method of norzelaminaldehyde derivatives in the following examples. The preparation method can be prepared by methods well known to those skilled in the art. The synthetic route is selected from Formula 2, 3 or 4. It is preferably prepared by the following steps:

[0039]

[0040]

[0041] Example 1

[0042] (1) Preparation of compound 1-3: Dissolve demethylzearalenaldehyde (45.2 mg, 0.1 mmol) in pyridine (1 mL), add propionic anhydride (76.6 μL, 0.6 mmol), butyric anhydride (97.4 μL, 0.6 mmol), valeric anhydride (122.9 μL, 0.6 mmol), and DMAP, respectively, and react at room temperature for 2–3 h.

[0043] (2) The reaction mixture was filtered and the residue was washed with ethyl acetate (3×30 mL). Then, the ethyl acetate solution was washed with hydrochloric acid (3×30 mL), saturated sodium bicarbonate (3×30 mL) and saturated brine (3×30 mL), respectively. Subsequently, the organic layer was dried over anhydrous sodium sulfate, filtered, and the filtrate was evaporated under reduced pressure. Finally, it was purified by silica gel column chromatography to obtain pure target derivative 1-3 (1: 31.1 mg, yield 55.8%; 2: 44.2 mg, yield 72.6%; 3: 25.1 mg, yield 38.7%).

[0044] (3) Compound 1: 11H NMR (400 MHz, CDCl3) δ: 10.21 (s, 1H), 7.44 (s, 1H), 6.38 (s, 1H), 2.62–2.56 (m, 2H), 2.56–2.50 (m, 2H), 2.39–2.30 (m, 1H), 2.28–2.03 (m, 3H), 2.03–1.78 (m, 4H), 1.75–1.60 (m, 4H), 1.57 (s, 3H), 1.28 (s, 3H), 1.27–1.24 (m, 4H), 1.24–1.22 (m, 4H), 1.08 (s, 3H), 1.05 (s, 3H), 0.96–0.88 (m, 1H), 0.64 (s, 3H); 13 13C NMR (100 MHz, CDCl3) δ: 190.5, 184.7, 184.0, 177.0, 171.7, 171.4, 154.3, 146.6, 138.0, 132.9, 127.9, 124.0, 122.3, 45.7, 44.2, 40.9, 40.3, 39.4, 37.0, 36.2, 34.7, 33.6, 32.7, 31.6, 30.5, 30.6, 29.5, 29.4, 28.8, 27.6, 27.3, 20.7, 18.9, 9.1, 9.0. HRESIMS m / z: 593.3119 [M+H] + (calcd for C 35 H 44 O8, 593.3109).

[0045] (4) Compound 2: 1 1H NMR (400 MHz, CDCl3) δ: 10.21 (s, 1H), 7.44 (s, 1H), 6.38 (s, 1H), 2.62–2.56 (m, 2H), 2.56–2.50 (m, 2H), 2.36–2.32 (m, 1H), 2.28–2.05 (m, 2H), 2.03–1.78 (m, 4H), 1.78–1.70 (m, 5H), 1.67–1.59 (m, 3H), 1.57 (s, 3H), 1.51–1.43 (m, 2H), 1.28 (s, 3H), 1.23 (s, 3H), 1.08 (s, 3H), 1.06 (s, 3H), 1.03–1.00 (m, 4H), 0.96–0.88 (m, 1H), 0.64 (s, 3H); 13C NMR(100MHz, CDCl3)δ:190.4,184.7,184.2,176.9,170.9,170.5,154.3,146.7,138.0,133.0,127.8,124.0,122.3,45.7,44.2,40.9,4 0.3,39.4,37.0,36.2,34.7,33.6,32.7,31.6,30.6,30.5,29.8,29.5,28.8,27.6,27.3,20.7,18.9,18.4,18.2,13.73,13.70.HRESIMS m / z:621.3422[M+H] + (calcd for C 37 H 48 O8,621.3424).

[0046] (5) Compound 3: 1 H NMR(400MHz, CDCl3)δ:10.22(s,1H),7.33(s,1H),6.38(s,1H),2.56–2.54(m,2H) ,2.53–2.50(m,2H),2.42–2.35(m,1H),2.33–2.22(m,2H),2.06–1.95(m,4H),1.73 –1.69(m,5H),1.68–1.64(m,3H),1.59(s,3H),1.41(s,3H),1.36(s,3H),1.32–1. 31(m,4H),1.27(s,3H),1.24(s,3H),1.09(s,3H),0.96–0.94(m,4H),0.70(s,3H); 13 C NMR(100MHz, CDCl3)δ:190.4,184.7,183.3,176.8,171.1,170.3,154.2,146.7,138.0,133.0,127.9,124.4,122.3,45.7,44.3,40.9,40.3,3 9.4,37.0,36.3,35.1,34.5,33.88,33.86,33.6,32.0,31.6,30.5,30.2,29.8,29.4,26.8,26.7,22.8,22.2,20.7,18.9,14.2,13.8.HRESIMS m / z:649.3742[M+H] + (calcd for C 39 H 52 O8,649.3735).

[0047] Example 2

[0048] (1) Preparation of compound 4: Dissolve norzelaminaldehyde (49.0 mg, 0.1 mmol) in dichloromethane (1 mL), add isobutyryl chloride (25.0 μL, 0.2 mmol) and TEA (51.0 μL, 0.4 mmol), and react at room temperature for 3 h.

[0049] (2) The reaction mixture was filtered and the residue was washed with ethyl acetate (3×30 mL). Then, the ethyl acetate solution was washed with saturated sodium bicarbonate (3×30 mL) and saturated brine (3×30 mL), respectively. Subsequently, the organic layer was dried over anhydrous sodium sulfate, filtered, and the filtrate was evaporated under reduced pressure. Finally, it was purified by silica gel column chromatography to obtain the pure target derivative 4 (4: 47.5 mg, yield 75.0%).

[0050] (3) Compound 4: 1 H NMR(400MHz, CDCl3)δ:10.22(s,1H),7.45(s,1H),6.40(s,1H),2.85–2.79(m,1H), 2.79–2.73(m,1H),2.41–2.34(m,1H),2.30–2.09(m,2H),2.05–1.81(m,5H),1.77–1 .63(m,3H),1.60(s,3H),1.53–1.47(m,1H),1.33–1.31(m,4H),1.30–1.28(m,4H),1 .29(s,3H),1.28(s,3H),1.26–1.24(m,4H),1.13(s,3H),1.09(s,3H),0.69(s,3H); 13 CNMR(100MHz, CDCl3)δ:190.2,184.6,183.9,176.7,174.1,173.8,154.1,146.7,137.9,133.0,127.7,123.9,122.0,53.5,53.4,45.6,44 .1,40.8,40.2,39.3,36.9,36.2,34.1,33.8,32.6,31.6,30.4,29.7,29.5,29.4,28.7,20.6,18.85,18.83,18.77,18.66,18.65.HRESIMS m / z:621.3419[M+H] + (calcd for C 37 H 48 O8,621.3422).

[0051] Example 3

[0052] (1) Preparation of Compound 5: Demethylzeramaldehyde (50.0 mg, 0.1 mmol) was dissolved in dichloromethane (1 mL), and DMAP, 2-thiopheneyl chloride (37.5 μL, 0.2 mmol) and EDC·HCl (60.0 mg, 0.3 mmol) were added and reacted at room temperature for 7 h.

[0053] (2) The reaction mixture was filtered and the residue was washed with ethyl acetate (3×30 mL). Then, the ethyl acetate solution was washed with hydrochloric acid (3×30 mL), saturated sodium bicarbonate (3×30 mL) and saturated brine (3×30 mL), respectively. Subsequently, the organic layer was dried over anhydrous sodium sulfate, filtered, and the filtrate was evaporated under reduced pressure. Finally, it was purified by silica gel column chromatography to obtain the pure target derivative 5 (5: 27.0 mg, yield 35.6%).

[0054] (3) Compound 5: 1 H NMR (400 MHz, CDCl3) δ: 10.34 (s, 1H), 7.92–7.87 (m, 4H), 7.78–7.72 (m, 2H), 7.64 (dd, J = 0.8, 4.0 Hz, 1H), 7.63 (dd, J = 0.8, 4.0 Hz,1H),7.28(s,1H),7.12(d,J=3.6 Hz,1H),7.10(d,J=3.6 Hz,1H),6.46(s,1H),2.56–2.53(m,1H),2.42–2.28(m,2H),2.08–1.92(m,3H),1.91–1.71(m,6H),1.70(s,3 H),1.45(s,3H),1.39(s,1H),1.28(s,3H),1.19(s,3H),1.15–1.09(m,1H),1.01–0.98(m,1H),0.89(s,3H); 13 C NMR(100MHz, CDCl3)δ:190.0,184.5,176.3,172.9,159.2,158.9,154.3,14 6.5,137.5,135.9,135.6,135.5,134.9,134.6,134.3,133.3,128.6,128.2 ,128.0,124.1,122.1,45.6,44.3,41.9,41.0,39.5,36.9,36.2,34.6,33.5 ,31.9,31.8,31.6,30.5,30.1,29.8,29.7,29.4,28.7,20.6,19.2.HRESIMS m / z:729.2549[M+H] + (calcd for C 41 H44 O8S2,729.2550).

[0055] Example 4

[0056] (1) Preparation of compound 6-11: Dissolve norzelaminaldehyde (45.0 mg, 0.1 mmol) in acetone (1 mL), add potassium carbonate (68.0 mg, 0.5 mmol) and corresponding halogenated hydrocarbon or benzyl bromide (methyl iodide: 37.5 μL, 0.6 mmol; ethyl iodide: 23.0 μL, 0.6 mmol; propyl bromide: 43.1 μL, 0.6 mmol; benzyl bromide: 63.0 μL, 0.6 mmol; o-bromobenzyl bromide: 167.1 mg, 0.6 mmol; p-bromobenzyl bromide: 167.1 mg, 0.6 mmol), respectively. Heat at 65°C and reflux for 3-5 h.

[0057] (2) After the reaction was completed, the reaction mixture was filtered and the residue was washed with ethyl acetate (3×30 mL). Then, the ethyl acetate solution was washed with water (3×30 mL). Subsequently, the organic layer was dried over anhydrous sodium sulfate, filtered, and the filtrate was evaporated under reduced pressure. Finally, it was purified by silica gel column chromatography to obtain pure target derivatives 6-11 (6: 40.6 mg, yield 77.8%; 7: 33.2 mg, yield 59.5%; 8: 50.5 mg, yield 57.4%; 9: 60.9 mg, yield 78.0%; 10: 75.0 mg, yield 73.1%; 11: 63.1 mg, yield 54.2%).

[0058] (3) Compound 6: 1 H NMR(400 MHz, CDCl3)δ:10.42(s,1H),6.99(s,1H),6.32(s,1H),3.95(s,3H),3.84(s,3H),3.53(s,3H),2.45–2.39(m,1H),2.35–2.14(m,2H),2.04–1 .92(m,3H),1.77–1.65(m,6H),1.57(s,3H),1.32–1.29(m,1H),1.26–1 .23(m,4H),1.16(s,3H),1.09(s,3H),0.96–0.94(m,1H),0.58(s,3H); 1313C NMR (100 MHz, CDCl3) δ: 193.2, 184.7, 179.1, 175.6, 157.2, 153.5, 145.1, 134.3, 124.1, 123.0, 109.1, 62.8, 56.1, 51.7, 45.5, 44.4, 40.8, 40.6, 39.4, 37.0, 36.4, 34.9, 33.9, 32.9, 31.7, 31.0, 30.6, 29.8, 29.6, 28.9, 20.7, 18.3. HRESIMS m / z: 523.3355 [M+H] + (calcd for C 32 H 42 O6, 523.3054).

[0059] (4) Compound 7: 1 1H NMR (400 MHz, CDCl3) δ: 10.45 (s, 1H), 6.99 (s, 1H), 6.33 (s, 1H), 4.17 (qd,

[0060] J = 4.8, 2.0 Hz, 2H), 4.08 (qd, J = 4.8, 2.0 Hz, 2H), 4.04–3.94 (m, 2H), 2.49–2.43 (m, 1H), 2.29–2.20 (m, 2H), 2.01–1.83 (m, 4H), 1.79–1.66 (m, 5H), 1.59 (s, 3H), 1.52 (t, J = 8.0 Hz, 3H), 1.44–1.37 (m, 1H), 1.35 (t, J = 8.0 Hz, 3H), 1.33 (s, 3H), 1.28–1.26 (m, 1H), 1.21 (t, J = 4.8 Hz, 3H), 1.19 (s, 3H), 1.12 (s, 3H), 1.02–0.97 (m, 1H), 0.63 (s, 3H); 13 13C NMR (100 MHz, CDCl3) δ: 193.4, 184.8, 178.5, 175.4, 156.6, 153.1, 144.3, 135.6, 124.0, 122.7, 109.8, 70.9, 64.6, 60.3, 45.4, 44.3, 40.7, 40.4, 39.4, 36.8, 36.3, 34.8, 33.7, 32.9, 31.6, 30.8, 30.5, 29.7, 29.6, 28.8, 20.6, 18.4, 15.4, 14.7, 14.0. HRESIMS m / z: 565.3514 [M+H] + (calcd for C 35 H 48O6,565.3524).

[0061] (5) Compound 8: 1 H NMR (400MHz, CDCl3) δ: 10.45 (s, 1H), 6.97 (s, 1H), 6.32 (s, 1H), 4.03 (qd, J = 4.0, 8.0Hz, 2H), 3.96 (qd, J =4.0,8.0Hz,2H),3.87(qd,J=3.6,5.2Hz,2H),2.50–2.41(m,1H),2.31–2.17(m,2H),1.94–1.86(m,4H), 1.86–1.67(m,5H),1.63–1.59(m,3H),1.58(s,3H),1.44–1.38(m,1H),1.31(s,3H),1.27–1.25(m,2H), 1.18(s,3H),1.13–1.09(m,6H),1.08(s,1H),0.99(t,J=4.0Hz,4H),0.92(t,J=4.0Hz,4H),0.61(s,3H); 13 CNMR(100MHz, CDCl3)δ:193.4,184.7,177.7,175.4,156.8,153.1,144.3,133.62,124.0,122.6,109.7,70.46,70.45,66.1,45.4,44.3,40 .7,40.3,39.4,36.8,36.3,34.8,33.7,32.7,31.6,30.8,30.5,29.66,29.65,28.8,23.2,22.5,21.8,20.5,18.4,10.7,10.6,10.4.HRESIMS m / z:607.3992[M+H] + (calcd for C 38 H 54 O6,607.3993).

[0062] (6) Compound 9: 11H NMR (400 MHz, CDCl3) δ: 10.41 (s, 1H), 7.53–7.49 (m, 2H), 7.46–7.40 (m, 5H), 7.40–7.38 (m, 1H), 7.33–7.31 (m, 6H), 7.30–7.27 (m, 1H), 7.11 (s, 1H), 6.34 (s, 1H), 5.31–5.24 (m, 2H), 5.11–5.06 (m, 2H), 5.04 (d, J = 8.0 Hz, 1H), 4.97 (d, J = 8.0 Hz, 1H), 2.52–2.48 (m, 1H), 2.32–2.25 (m, 1H), 2.18–2.05 (m, 2H), 1.84–1.71 (m, 5H), 1.66–1.56 (m, 3H), 1.54 (s, 3H), 1.48–1.39 (m, 1H), 1.32 (s, 3H), 1.24 (s, 3H), 1.14 (s, 3H), 1.05–0.99 (m, 1H), 0.91–0.88 (m, 1H), 0.61 (s, 3H); 13 13C NMR (100 MHz, CDCl3) δ: 193.3, 184.7, 178.2, 175.7, 156.3, 153.4, 144.0, 136.9, 135.9, 135.7, 134.8, 128.91, 128.89, 128.83, 128.56, 128.54, 128.4, 128.23, 128.22, 128.09, 128.08, 128.06, 127.7, 127.6, 127.5, 127.0, 124.0, 123.1, 111.1, 71.2, 66.26, 66.25, 45.4, 44.3, 40.8, 40.5, 39.4, 36.8, 36.3, 34.7, 33.6, 32.9, 31.6, 30.8, 30.5, 29.8, 29.6, 28.8, 20.6, 18.5. HRESIMS m / z: 751.3992 [M + H] + (calcd for C 50 H 54 O6, 751.3993).

[0063] (7) Compound 10: 11H NMR (400 MHz, CDCl3) δ: 10.44 (s, 1H), 7.68 (d, J = 8.0 Hz, 1H), 7.61 (d, J = 8.0 Hz, 1H), 7.53 (t, J = 4.0 Hz, 2H), 7.48 (d, J = 8.0 Hz, 1H), 7.38 (d, J = 8.0 Hz, 1H), 7.30–7.26 (m, 3H), 7.21 (t, J = 8.0 Hz, 1H), 7.18–7.11 (m, 3H), 7.09 (s, 1H), 5.32–5.30 (m, 2H), 5.24–5.22 (m, 2H), 5.12–5.02 (m, 2H), 2.53–2.46 (m, 1H), 2.34–2.18 (m, 3H), 1.94–1.80 (m, 3H), 1.77–1.60 (m, 5H), 1.54 (s, 3H), 1.49–1.43 (m, 1H), 1.33–1.31 (m, 4H), 1.26 (s, 3H), 1.13 (s, 3H), 1.05–0.99 (m, 1H), 0.57 (s, 3H); 13 13C NMR (100 MHz, CDCl3) δ: 192.9, 184.6, 178.1, 175.7, 155.8, 153.6, 143.9, 136.7, 135.2, 134.9, 134.7, 132.9, 132.8, 132.3, 130.3, 129.9, 129.81, 129.79, 129.5, 129.2, 127.9, 127.52, 127.47, 124.0, 123.9, 123.4, 122.7, 122.2, 110.9, 70.3, 66.0, 65.0, 45.4, 44.3, 40.8, 40.7, 39.4, 36.8, 36.3, 34.7, 33.6, 33.0, 31.9, 31.6, 30.8, 30.5, 29.7, 28.8, 20.6, 18.4. HRESIMS m / z: 985.3196 [M + H] + (calcd for C 50 H 51 Br3O6, 985.1309).

[0064] (8) Compound 11: 11H NMR (400 MHz, CDCl3) δ: 10.37 (s, 1H), 7.54–7.51 (m, 2H), 7.49–7.44 (m, 2H), 7.42–7.39 (m, 4H), 7.31–7.28 (m, 2H), 7.18–7.13 (m, 2H), 7.04 (s, 1H), 6.31 (s, 1H), 5.20–5.08 (m, 2H), 5.01–4.89 (m, 4H), 2.45–2.39 (m, 1H), 2.25–2.00 (m, 3H), 1.92–1.79 (m, 3H), 1.72–1.67 (m, 5H), 1.52 (s, 3H), 1.45–1.35 (m, 1H), 1.29–1.24 (m, 4H), 1.19 (s, 3H), 1.09 (s, 3H), 1.02–0.94 (m, 1H), 0.52 (s, 3H); 13 13C NMR (100 MHz, CDCl3) δ: 193.0, 184.6, 178.1, 175.5, 156.2, 153.5, 144.0, 139.9, 136.0, 134.9, 134.7, 132.04, 132.03, 131.71, 131.70, 131.55, 131.54, 130.35, 130.34, 129.94, 129.92, 129.38, 129.36, 124.1, 123.5, 122.7, 122.3, 121.5, 110.9, 70.6, 65.6, 64.7, 45.5, 44.3, 40.8, 40.6, 39.5, 36.8, 36.3, 34.8, 33.7, 32.9, 31.7, 30.8, 30.6, 29.8, 29.7, 28.8, 20.7, 18.5. HRESIMS m / z: 985.3215 [M+H] + (calcd for C 50 H 51 Br3O6, 985.1309).

[0065] Example 5

[0066] (1) Preparation of compounds 12, 13: 6, 7 (40.0 mg, 0.07 mmol) were dissolved in 1 M sodium hydroxide MeOH (5 mL) solution, and the mixture was heated to 90°C overnight. The reaction mixture was filtered and the residue was washed with ethyl acetate (3×30 mL). Then, the ethyl acetate solution was washed with 3 M hydrochloric acid (3×30 mL) and water (3×30 mL), respectively. Subsequently, the organic layer was dried over anhydrous sodium sulfate, filtered, and the filtrate was evaporated under reduced pressure. Finally, it was purified by silica gel column chromatography to obtain pure target derivatives 12, 13 (12: 39.7 mg, yield 51.1%; 13: 37.3 mg, yield 49.5%).

[0067] (2) Compound 12: 1 H NMR(400MHz, CDCl3)δ:10.41(s,1H),6.87(s,1H),6.39(s,1H),3.96(s,3H),3.86(s,3H),2.45–2.38(m,1H),2.30–2.13(m,3H),2.01–1.95(m,3H) ),1.93–1.67(m,5H),1.54(s,3H),1.43–1.32(m,1H),1.29(s,3H),1.18 (s,3H),1.11(s,3H),1.03–0.93(m,1H),0.91–0.86(m,1H),0.61(s,3H); 13 C NMR(100MHz, CDCl3)δ:192.8,185.1,182.3,176.4,157.2,153.7,144.8,134.0,124.0,122.3,108.8,62.7,56.1,4 5.6,44.3,40.9,40.2,39.4,36.8,36.4,34.6,33.7,32.4,31.6,30.8,30.4,29.7,29.6,28.7,20.5,18.9.HRESIMS m / z:509.2896[M+H] + (calcd for C 31 H 40 O6,509.2898).

[0068] (3) Compound 13: 1H NMR(400MHz, CDCl3)δ:10.40(s,1H),7.27(s,1H),6.42(s,1H),4.24–3.96(m,4 H),2.43–2.38(m,1H),2.28–2.10(m,3H),1.91–1.74(m,3H),1.73–1.58(m,5H), 1.52(s,3H),1.47–1.38(m,1H),1.34(t,J=8.0Hz,3H),1.28(s,3H),1.26(s,3H ),1.18(s,3H),1.10(s,3H),0.99–0.92(m,1H),0.90–0.85(m,1H),0.63(s,3H); 13 C NMR(100MHz, CDCl3)δ:193.0,185.0,182.3,176.1,156.8,156.7,153.4,134.2,124.0,122.1,109.5,64.6,62.4,45.5,4 4.3,40.8,40.1,39.5,36.7,36.3,34.7,33.6,32.4,31.6,30.8,30.4,29.7,29.6,28.7,20.5,18.9,15.4,14.7.HRESIMS m / z:537.3209[M+H] + (calcd for C 33 H 44 O6,537.3211).

[0069] Example 6

[0070] (1) Preparation of Compounds 14-17: Dissolve norzelaminaldehyde (50.0 mg, 0.1 mmol) in acetone (1 mL), add potassium carbonate (68.0 mg, 0.5 mmol) and the corresponding halogenated hydrocarbon or benzyl bromide (iodomethane: 37.5 μL, 0.6 mmol; 1,5-dibromopentane: 92.5 μL, 0.6 mmol; p-bromobenzyl bromide: 167.1 mg, 0.6 mmol; 4-methylbenzyl bromide: 131.1 μL, 0.6 mmol), and react at room temperature for 9-12 h.

[0071] (2) After the reaction was completed, the reaction mixture was filtered and the residue was washed with ethyl acetate (3×30 mL). Then, the ethyl acetate solution was washed with water (3×30 mL). Subsequently, the organic layer was dried over anhydrous sodium sulfate, filtered, and the filtrate was evaporated under reduced pressure. Finally, it was purified by silica gel column chromatography to obtain pure target derivatives 14-17 (14: 19.0 mg, yield 37.3%; 15: 72.4 mg, yield 49.7%; 16: 11.1 mg, yield 8.5%; 17: 39.5 mg, yield 59.6%).

[0072] (3) Compound 14: 1 H NMR(400MHz, CDCl3)δ:10.98(s,1H),7.10(s,1H),6.34(s,1H),3.98(s,3H),3.52(s,3H),2.44–2.37(m,1H),2.29–2.15(m,3H),2.01–1 .87(m,3H),1.75–1.64(m,5H),1.57(s,3H),1.36–1.34(m,2H),1.24(s,3H),1.16(s,3H),1.10(s,3H),1.02–0.95(m,1H),0.57(s,3H); 13 C NMR(100MHz, CDCl3)δ:200.1,186.0,179.0,173.7,152.7,152.4,149.7,125.4,123.2,117.0,112.1,56.3,51.7,4 5.2,44.3,40.7,40.6,39.4,36.5,36.4,34.9,33.9,32.9,31.7,31.6,31.0,30.6,29.8,28.7,20.6,18.3.HRESIMS m / z:509.2898[M+H] + (calcd for C 31 H 40 O6,509.2898).

[0073] (4) Compound 15: 11H NMR (400 MHz, CDCl3) δ: 10.97 (s, 1H), 7.08 (s, 1H), 6.33 (s, 1H), 4.10 (t, J = 6.5 Hz, 2H), 3.96–3.89 (m, 1H), 3.88–3.81 (m, 1H), 3.44 (t, J = 4.0 Hz, 2H), 3.37 (t, J = 4.0 Hz, 2H), 2.44–2.38 (m, 1H), 2.30–2.15 (m, 2H), 1.97–1.81 (m, 12H), 1.70–1.64 (m, 5H), 1.55 (s, 3H), 1.31–1.29 (m, 5H), 1.26 (s, 3H), 1.16 (s, 3H), 1.09 (s, 3H), 1.07–0.91 (m, 2H), 0.58 (s, 3H); 13 13C NMR (100 MHz, CDCl3) δ: 200.1, 186.0, 178.5, 173.5, 153.0, 151.9, 149.6, 125.4, 124.4, 123.1, 113.3, 69.1, 64.2, 45.1, 44.3, 40.7, 40.6, 39.4, 36.4, 34.9, 33.9, 33.6, 33.0, 32.4, 32.3, 31.7, 31.6, 30.9, 30.6, 30.2, 29.8, 29.4, 28.7, 28.3, 27.7, 24.8, 22.8, 20.6, 18.4. HRESIMS m / z: 777.2362 [M + H] + (calcd for C 39 H 54 Br2O6, 777.2360).

[0074] (5) Compound 16: 1 1H NMR (400 MHz, CDCl3) δ: 10.96 (s, 1H), 7.54–7.51 (m, 2H), 7.40–7.34 (m, 4H), 7.15–7.12 (m, 2H), 7.07 (s, 1H), 6.31 (s, 1H), 5.27–5.15 (m, 2H), 4.97–4.82 (m, 2H), 2.45–2.37 (m, 1H), 2.21–2.18 (m, 1H), 2.08–1.99 (m, 2H), 1.71–1.66 (m, 3H), 1.58–1.51 (m, 5H), 1.44 (s, 3H), 1.42–1.35 (m, 2H), 1.27 (s, 3H), 1.18 (s, 3H), 1.09 (s, 3H), 1.00–0.96 (m, 1H), 0.49 (s, 3H); 1313C NMR (100 MHz, CDCl3) δ: 200.0, 185.5, 178.1, 173.5, 153.3, 151.0, 149.2, 134.90, 134.87, 132.05, 132.04, 131.71, 131.69, 129.94, 129.92, 129.25, 129.24, 125.4, 123.8, 122.5, 122.1, 117.4, 115.1, 70.7, 65.6, 45.1, 44.3, 40.6, 40.5, 39.4, 36.30, 36.25, 34.8, 33.7, 32.9, 31.6, 30.8, 30.61, 30.60, 29.8, 28.7, 20.5, 18.4. HRESIMS m / z: 817.5851 [M+H]+ + (calcd for C 43 H 46 Br2O6, 817.1734).

[0075] (6) Compound 17: 1 1H NMR (400 MHz, CDCl3) δ: 10.97 (s, 1H), 7.37–7.32 (m, 2H), 7.25 (s, 1H), 7.21–7.11 (m, 4H), 7.09–7.02 (m, 2H), 6.30 (s, 1H), 5.30–5.15 (m, 2H), 4.94–4.87 (m, 2H), 2.47–2.42 (m, 1H), 2.33 (s, 3H), 2.22 (s, 3H), 2.09–1.98 (m, 3H), 1.88–1.81 (m, 3H), 1.60–1.48 (m, 5H), 1.42 (s, 3H), 1.36 (s, 3H), 1.33 (s, 1H), 1.17 (s, 3H), 1.08 (s, 3H), 1.07–0.89 (m, 2H), 0.50 (s, 3H); 13C NMR(100MHz, CDCl3)δ:200.0,186.0,178.3,173.7,153.3,151.3,149.5,138.4,138.0 ,132.8,132.7,129.6,129.3,128.3,127.7,127.2,126.9,125.3,124.4,123.6,123.4, 117.2,115.0,71.3,66.4,50.9,45.1,44.3,40.6,39.4,36.3,36.2,34.8,33.7,33.0,3 2.0,31.7,31.6,30.6,30.2,28.7,21.3,21.2,20.5,18.3.HRESIMSm / z:689.3837[M+H] + (calcd for C 45 H 52 O6,689.3837).

[0076] Example 7

[0077] (1) Preparation of Compound 18: Demethylzeramaldehyde (48.0 mg, 0.1 mmol) was dissolved in DCM (1 mL), EDC·HCl (34.0 mg, 0.2 mmol) and HOBt (42.0 mg, 0.2 mmol) were added, ethylamine (8.5 μL, 0.1 mmol) was added at 0°C, the mixture was reacted for 30 min, and then DIPEA (38.0 μL, 0.2 mmol) was added. The reaction was allowed to proceed overnight at room temperature.

[0078] (2) After the reaction was completed, the reaction mixture was filtered and the residue was washed with ethyl acetate (3×30 mL). Then, the ethyl acetate solution was washed with water (3×30 mL). Subsequently, the organic layer was dried over anhydrous sodium sulfate, filtered, and the filtrate was evaporated under reduced pressure. Finally, it was purified by silica gel column chromatography to obtain the pure target derivative 18 (18: 40.3 mg, yield 79.5%).

[0079] (3) Compound 18: 1H NMR(400MHz, CDCl3)δ:9.99(s,1H),7.43(s,1H),7.04(s,1H),6.28(s,1H),3.77–3.59(m,2H),2.81–2.69(m,1H),2.44–2.21(m,3H),2.16–1.95 (m,5H),1.88–1.75(m,3H),1.65(s,3H),1.58(s,3H),1.48–1.42(m,2H) ,1.37(s,3H),1.27(s,3H),1.21(s,3H),0.96–0.94(m,1H),0.09(s,3H); 13 C NMR(100MHz, CDCl3)δ:185.8,174.3,171.9,165.4,143.4,128.6,128.5,125.8,124.7,120.6,108.0,45.9,44.7,4 4.2,40.8,39.7,36.2,35.0,34.5,33.3,33.2,31.9,31.5,30.5,30.1,29.7,28.6,22.7,20.3,19.5,15.6.HRESIMS m / z:508.3060[M+H] + (calcd for C 31 H 41 NO5,508.3057).

[0080] Example 8

[0081] (1) Preparation of compound 19: 6 (52.0 mg, 0.1 mmol) was dissolved in 0.2 M THF (1 mL), and NaOtBu (24.0 mg, 0.2 mmol) was added and the mixture was allowed to stand at room temperature overnight.

[0082] (2) The reaction mixture was evaporated under reduced pressure to remove THF. The crude oil obtained was washed with ethyl acetate, then with water (3×30 mL) and brine (3×30 mL). Subsequently, the organic layer was dried over anhydrous sodium sulfate, filtered, and the filtrate was evaporated under reduced pressure. Finally, it was purified by silica gel column chromatography to obtain the pure target derivative 19 (19: 47.9 mg, yield 91.8%). 1H NMR(500MHz, CDCl3)δ:7.54(s,1H),6.91(s,1H),6.36(s,1H),4.30–4.14(m ,2H),3.97(s,3H),3.95(s,3H),3.54(s,3H),2.46–2.43(m,1H),2.34–2.16 (m,2H),2.11–1.77(m,5H),1.77–1.66(m,4H),1.58(s,3H),1.41–1.37(m,2 H),1.33(s,3H),1.18(s,3H),1.11(s,3H),1.02–0.96(m,1H),0.58(s,3H); 13 C NMR(125 MHz, CDCl3)δ:184.8,178.9,174.7,153.3,150.0,147.8,128.8,124.4,123.6,106.8,68.1,56.03,56.01,51.6,4 5.2,44.3,40.5,40.0,39.5,36.5,36.4,34.8,33.7,32.8,31.6,30.9,30.5,29.8,29.7,28.9,20.5,18.2.HRESIMS m / z:525.3492[M+H] + (calcd for C 32 H 44 O6,525.3211).

[0083] Example 9

[0084] (1) Preparation of compound 20: 7 (74.0 mg, 0.1 mmol) was dissolved in 0.2 M THF (1 mL), and NaOtBu (21.2 mg, 0.2 mmol) was added and the mixture was allowed to react at room temperature overnight.

[0085] (2) The reaction mixture was evaporated under reduced pressure to remove THF. The obtained crude oil was washed with ethyl acetate, and then with water (3×30 mL) and brine (3×30 mL). Subsequently, the organic layer was dried over anhydrous sodium sulfate, filtered, and the filtrate was evaporated under reduced pressure. Finally, it was purified by silica gel column chromatography to obtain the pure target derivative 20 (20: 35.8 mg, yield 48.2%). 1H NMR(500 MHz, CDCl3)δ:7.54(s,1H),6.89(s,1H),6.34(s,1H),5.30(s,2H),4.21–4.14(m,4H),4.02–3.91(m,2H),2.47–2.4 3(m,1H),2.30–2.17(m,2H),2.09–2.02(m,1H),1.99–1.79(m,3H),1.77–1.60(m,5H),1.56(s,3H),1.50(t,J=10.0 Hz,3H),1.47(t,J=10.0 Hz,3H),1.43–1.34(m,2H),1.32(s,3H),1.19(d,J=5.0 Hz,3H),1.17(s,3H),1.11(s,3H),1.01–0.96(m,1H),0.60(s,3H); 13 C NMR(125 MHz, CDCl3)δ:182.5,176.1,172.2,150.6,147.4,145.1,122.1,121.2,106.1,105.8,62.3,62.0,57.9,51.1,42.9,42.0,3 8.0,37.6,37.2,34.1,32.5,31.4,30.6,29.3,28.4,28.2,27.45,27.40,27.37,26.5,20.4,18.2,16.0,12.4,11.7.HRESIMS m / z:567.3589[M+H] + (calcd for C 35 H 50 O6,567.3680).

[0086] Example 10

[0087] (1) Preparation of compound 21: 9 (42.0 mg, 0.05 mmol) was dissolved in 0.2 M THF (1 mL), and NaOtBu (13.0 mg, 0.1 mmol) was added and the mixture was allowed to stand at room temperature overnight.

[0088] (2) The reaction mixture was evaporated under reduced pressure to remove THF. The obtained crude oil was washed with ethyl acetate, and then with water (3×30 mL) and brine (3×30 mL). Subsequently, the organic layer was dried over anhydrous sodium sulfate, filtered, and the filtrate was evaporated under reduced pressure. Finally, it was purified by silica gel column chromatography to obtain the pure target derivative 21 (21: 18.6 mg, yield 28.3%). 1H NMR(500 MHz, CDCl3)δ:7.66(s,1H),7.52–7.43(m,4H),7.40–7.35(m,4H),7.33–7.23(m,7H),6.92(s, 1H),6.31(s,1H),5.30–5.25(m,2H),5.25–5.19(m,4H),5.00–4.89(m,2H),2.47–2.44(m,1H) ,2.26–2.19(m,1H),2.09–1.99(m,2H),1.76–1.63(m,5H),1.59–1.49(m,3H),1.44(s,3H),1. 41–1.35(m,1H),1.28–1.25(m,4H),1.20(s,3H),1.08(s,3H),1.02–0.93(m,1H),0.55(s,3H); 13 C NMR(125 MHz, CDCl3)δ:183.6,177.0,173.6,151.7,148.9,146.6,135.8,135.6,134.8,127.60,127.5 9,127.48,127.47,127.01,127.00,126.85,126.84,126.83,126.37,126.36,126.35,126.22, 126.21,126.20,123.3,123.1,109.4,108.2,70.2,69.7,65.2,52.4,44.1,43.2,39.5,38.8, 38.4,35.4,35.3,33.7,32.4,31.8,30.6,29.8,29.5,28.72,28.66,27.8,19.4,17.3.HRESIMS m / z:753.4060[M+H] + (calcd for C 50 H 56 O6,753.4150).

[0089] Embodiment 11

[0090] (1) Preparation of compounds 22 and 23: 10 (40.0 mg, 0.04 mmol) and 11 (40.0 mg, 0.04 mmol) were dissolved in 0.2 M THF (1 mL), and NaOtBu (10.0 mg, 0.08 mmol) was added and the mixture was allowed to stand at room temperature overnight.

[0091] (2) The reaction mixture was evaporated under reduced pressure to remove THF. The crude oil obtained was washed with ethyl acetate, and then with water (3×30 mL) and brine (3×30 mL). Subsequently, the organic layer was dried over anhydrous sodium sulfate, filtered, and the filtrate was evaporated under reduced pressure. Finally, it was purified by silica gel column chromatography to obtain pure target derivatives 22, 23 (22: 19.2 mg, yield 23.3%; 23: 23.7 mg, yield 28.8%).

[0092] (3) Compound 22: 1 H NMR(500MHz, CDCl3)δ:7.70(s,1H),7.69–7.63(m,2H),7.61–7.56(m,2H),7.50–7.48(m,1H),7.35–7.31(m,3H) ),7.27–7.16(m,3H),7.11–7.05(m,1H),6.94(s,1H),6.32(s,1H),5.34–5.33(m,2H),5.31–5.28(m,2H),5.27 –5.25(m,2H),5.08–4.98(m,2H),2.47–2.45(m,1H),2.28–2.04(m,3H),1.79–1.63(m,5H),1.62–1.52(m,3H), 1.47(s,3H),1.43–1.40(m,1H),1.29–1.28(m,4H),1.23(s,3H),1.10(s,3H),1.03–0.94(m,1H),0.49(s,3H); 13 C NMR (125MHz, CDCl3) δ: 184.6, 177.9, 174.8, 151.9, 150.3, 147.4, 136.2, 135.6, 135.1, 132. 9,132.61,132.57,130.3,129.7,129.4,129.3,129.0,128.9,127.8,127.5,127.44,127.43, 124.3,123.9,122.4,122.1,110.2,109.4,70.27,70.23,65.9,53.4,45.2,44.3,40.5,39.9 ,39.4,36.5,36.3,34.7,33.4,32.9,31.6,30.7,30.5,29.8,29.7,28.8,20.4,17.9.HRESIMS m / z:987.3217[M+H] + (calcd for C 50 H 53 Br3O6,987.1465).

[0093] (4) Compound 23: 1 H NMR(500MHz, CDCl3)δ:7.63(s,1H),7.54–7.47(m,4H),7.44–7.39(m,2H),7.35–7.31(m,4H),7. 17–7.14(m,2H),6.92(s,1H),6.32(s,1H),5.22–5.18(m,2H),5.17–5.13(m,4H),4.89–4.84(m, 2H),2.46–2.40(m,1H),2.26–1.97(m,3H),1.75–1.65(m,5H),1.65–1.59(m,3H),1.48(s,3H),1 .43–1.37(m,1H),1.29–1.26(m,4H),1.19(s,3H),1.10(s,3H),1.02–0.93(m,1H),0.53(s,3H); 13 C NMR(125MHz, CDCl3)δ:184.5,177.9,174.5,152.6,150.1,147.4,135.8,135.5,134.8,131.77 ,131.76,131.67,131.65,131.62,131.61,129.80,129.79,128.99,128.98,128.94,128.93,12 4.4,124.2,122.1,122.0,121.8,110.1,109.3,70.4,69.9,65.4,53.5,45.2,44.2,40.5,39.9 ,39.4,36.32,36.29,34.7,33.5,32.8,31.6,30.8,30.5,29.7,29.7,28.8,20.4,18.4.HRESIMS m / z:987.3276[M+H] + (calcd for C 50 H 53 Br3O6,987.1465).

[0094] Example 12

[0095] (1) Preparation of compounds 24 and 25: 7 (44.0 mg, 0.08 mmol) was dissolved in anhydrous ethanol (1 mL), and benzohydrazide, valeryl hydrazide (11.0 mg, 0.08 mmol) and acetic acid (8.0 μL) were added respectively, and the mixture was heated under reflux at 60°C for 3 h.

[0096] (2) The reaction mixture was filtered and the residue was washed with ethyl acetate (3×30 mL). Then, the ethyl acetate solution was washed with water (3×30 mL). Subsequently, the organic layer was dried over anhydrous sodium sulfate, filtered, and the filtrate was evaporated under reduced pressure. Finally, it was purified by silica gel column chromatography to obtain pure target derivatives 24, 25 (24: 31.6 mg, yield 74.7%; 25: 33.4 mg, yield 64.7%).

[0097] (3) Compound 24: 1 H NMR(400MHz, CDCl3)δ:8.84(s,1H),8.11–7.80(m,2H),7.64–7.20(m,3H),6.97(s,1H),6.07(s,1H) ,4.29–4.21(m,2H),4.21–4.13(m,2H),4.09–3.95(m,2H),2.47–2.42(m,1H),2.30–2.16(m,3H),2.1 2–1.90(m,3H),1.89–1.59(m,5H),1.58(s,3H),1.52(t,J=10.0Hz,3H),1.45–1.39(m,2H),1.28(s, 3H),1.27(s,3H),1.23(t,J=10.0Hz,3H),1.19(s,3H),1.10(s,3H),1.01–0.98(m,1H),0.60(s,3H); 13 C NMR (100MHz, CDCl3) δ: 185.8, 178.6, 174.1, 164.1, 156.5, 153.5, 146.7, 146. 2,133.7,131.6,128.55,128.54,127.65,127.64,125.05,125.04,123.1,109 .3,71.4,64.4,60.4,45.0,44.2,40.6,40.4,39.3,36.8,36.3,34.8,34.1,33 .0,31.6,30.8,30.4,29.7,29.6,28.5,20.4,18.3,15.5,14.7,14.1.HRESIMS m / z:683.4056[M+H] + (calcd for C 42 H 54 N2O6,683.4055).

[0098] (4) Compound 25: 1H NMR(400MHz, CDCl3)δ:8.20(s,1H),6.99(s,1H),6.27(s,1H),4.20–4.14(m,2H),4.13–4.03(m,2H),4.03–3.94(m, 2H),2.71–2.64(m,2H),2.47–2.42(m,1H),2.34–2.18(m,2H),2.10–2.02(m,1H),1.94–1.81(m,3H),1.78–1.64(m,5 H),1.58(s,3H),1.57–1.55(m,1H),1.52(t,J=5.0Hz,3H),1.44–1.39(m,2H),1.39–1.34(m,4H),1.33–1.29(m,3H), 1.27–1.25(m,2H),1.22–1.19(m,3H),1.18(s,3H),1.12(s,3H),1.02–0.95(m,1H),0.95–0.89(m,3H),0.62(s,3H); 13 C NMR (100MHz, CDCl3) δ: 185.6, 178.5, 175.2, 173.3, 156.4, 153.5, 145.6, 142.6, 127.9, 125.1, 123.0, 109.3, 70.2, 64.4, 60.3, 45.0, 44.2, 40. 6,40.3,39.2,37.1,36.4,34.8,33.9,32.9,32.3,31.5,30.7,30.5,29.7,29.6,28.7,26.5,22.5,20.6,18.4,15.5,14.7,14.0,13.9.HRESIMS m / z:663.4361[M+H] + (calcd for C 40 H 58 N2O6,663.4366).

[0099] Embodiment 13

[0100] (1) Preparation of compound 26: 7 (44.0 mg, 0.08 mmol) was dissolved in anhydrous ethanol (1 mL), and 4-hydrazinobenzoic acid (10.0 mg, 0.01 mmol) and acetic acid (8.0 μL) were added respectively, and the mixture was heated under reflux at 60° C. for 3 h.

[0101] (2) The reaction mixture was filtered and the residue was washed with ethyl acetate (3×30 mL). Then, the ethyl acetate solution was washed with water (3×30 mL). Subsequently, the organic layer was dried over anhydrous sodium sulfate, filtered, and the filtrate was evaporated under reduced pressure. Finally, it was purified by silica gel column chromatography to obtain the pure target derivative 26 (26: 26.7 mg, yield 49.0%). 1 H NMR (400MHz, CDCl3) δ: 8.36 (s, 1H), 7.94 (d, J = 10.0Hz, 2H), 7.03 (d, J = 10.0Hz, 2H), 6.99 (s,1H),6.28(s,1H),4.16–4.12(m,4H),4.05–3.94(m,2H),2.48–2.44(m,1H),2.33–2.1 8(m,3H),2.05–1.95(m,3H),1.76–1.65(m,5H),1.60(s,3H),1.56–1.52(m,3H),1.51–1. 49(m,1H),1.36–1.34(m,3H),1.32(s,3H),1.31–1.29(m,1H),1.27(s,3H),1.22–1.29(m 3H),1.12(s,3H),1.01–0.96(m,1H),0.63(s,3H); 13 C NMR(100MHz, CDCl3)δ:186.0,178.5,173.3,171.3,158.8,156.6,153.8,14 9.4,145.7,141.9,139.5,132.1,128.8,125.4,122.8,119.3,111.5,108.9, 70.6,64.4,60.4,45.0,44.2,40.7,40.3,39.2,37.2,36.3,34.8,33.9,32. 9,31.6,30.8,30.5,29.8,29.7,28.7,20.7,18.4,15.4,14.7,14.0.HRESIMS m / z:699.4626[M+H] + (calcd for C 42 H 54 N2O7,699.4004).

[0102] Embodiment 14

[0103] Inhibitory effects of norzelaminaldehyde and its derivatives on proliferation of A549, HCT116 and HeLa cells.

[0104] (1) Cell inoculation: Digest EGFR sensitive mutant A549, HCT116 and HeLa cells in the exponential growth phase. Prepare a suspension using DMEM / RPMI1640 medium containing 10% calf serum. Inoculate tumor cells at a seed cell count of 0.5×10 4 Each well of the plate was inoculated with 100 μL of the culture medium and cultured at 37°C in a CO2 incubator for 12 h.

[0105] (2) Dosing: The test compound was added with DMSO to prepare a stock solution of a certain concentration, and then prepared into 50.0 μM, 25.0 μM, 12.5.0 μM, 6.25 μM, and 3.125 μM solutions respectively using DMEM / RPMI1640 culture medium containing 10% calf serum. The solutions were inoculated in a 96-well cell culture plate, and three parallel wells were set for each concentration. The cells were cultured in a 37°C, 5% CO2 incubator for 24 h, and the cell morphology and growth changes were observed under an inverted microscope.

[0106] (3) Termination of culture: First, prepare a mixed solution containing 10% CCK-8, take 450 μL of CCK-8 and dilute it to 4500 μL with serum-free culture medium, then add 100 μL of CCK-8 and culture medium mixed solution to each well, culture in a 37°C, 5% CO2 incubator for 1.5 hours, place on a microplate reader to measure the absorbance A value at a wavelength of 450 nm, and calculate the inhibition rate according to the following formula. Inhibition rate (%) = (average absorbance A value of the control group - average absorbance A value of the drug-added group) / (average absorbance A value of the control group - average absorbance A value of the blank group) × 100%; the control group is the culture well with a concentration of 0 demethylzelaminaldehyde and its derivatives and DMSO added, and the blank group is the culture well with culture medium and CCK-8 solution but no cells. GraphPadPrism6 was used for data statistical analysis and IC was calculated. 50 .

[0107] The results of the CCK-8 experiment are shown in Table 1, indicating that norzelaminaldehyde and its derivatives have a significant inhibitory effect on the proliferation of A549, HCT116 and HeLa cells, among which it is shown that norzelaminaldehyde and its derivatives have the largest inhibition gradient on the non-small cell lung cancer cell line A549, and have a relatively strong inhibitory effect, which can be further used in the research and development of new anti-non-small cell lung cancer drugs targeting EGFR.

[0108] Table 1

[0109]

[0110]

[0111] Embodiment 15

[0112] Flow cytometry for cell cycle analysis

[0113] (1) Cell inoculation: A549 cells in the exponential growth phase were digested and made into a suspension using DMEM medium containing 10% calf serum. Tumor cells were seeded at a cell number of 10×10 4 Each well of the plate was inoculated with 2 mL of the culture medium and cultured at 37°C in a CO2 incubator for 12 h.

[0114] (2) Dosing: The test compound was added with DMSO to prepare a stock solution of a certain concentration, and then prepared into 5.0 μM, 15.0 μM, and 30.0 μM solutions using DMEM culture medium containing 10% calf serum. The solutions were inoculated into 6-well cell culture plates, and three parallel wells were set for each concentration. The cells were cultured in a 37°C, 5% CO2 incubator for 24 h, and the cell morphology and growth changes were observed under an inverted microscope.

[0115] (3) Collect cells: Wash the cells once with PBS, collect them by centrifugation at 1500 rpm for 5 min, and adjust the cell concentration to 1×10 6 / mL, take 1mL of single cell suspension. After centrifugation of the prepared single cell suspension, remove the supernatant, add 300μL of PBS, add the cells to 700μL of anhydrous pre-cooled ethanol, mix well, and fix for 24h.

[0116] (4) Staining: Before staining, wash away the fixative with PBS, add 100 μL of RNase A solution to the cell pellet, resuspend the cells, then add 400 μL of PI staining solution and mix well, and incubate at 4°C in the dark for 30 min.

[0117] (5) On-machine detection: record the red fluorescence at the excitation wavelength of 488 nm.

[0118] The flow cytometry results are as follows Figure 1 As shown, compared with the control group, compound 7 significantly increased the number of cells in the S group by 1.06 times (30.0 μM). At the same time, the proportion of cells in the G1 phase decreased from 63.26% (control group) to 65.07% (5.0 μM), 64.89% (15.0 μM), and 62.72% (30.0 μM), and the proportion of cells in the G2 phase decreased from 4.43% (control group) to 4.85% (5.0 μM), 3.48% (15.0 μM), and 2.98% (30.0 μM). The G1 and G2 phase rates showed a trend of change with the increase in the concentration of compound 7, but there was no significant difference. These results indicate that derivative 7 may have a weak effect on cell cycle arrest.

[0119] Example 16

[0120] Flow cytometry detection of cell apoptosis

[0121] (1) Cell inoculation: A549 cells in the exponential growth phase were digested and made into a suspension using DMEM medium containing 10% calf serum. Tumor cells were seeded at a cell number of 10×10 4 Each well of the plate was inoculated with 2 mL of the culture medium and cultured at 37°C in a CO2 incubator for 12 h.

[0122] (2) Dosing: The test compound was added with DMSO to prepare a stock solution of a certain concentration, and then prepared into 5.0 μM, 15.0 μM, and 30.0 μM solutions using DMEM culture medium containing 10% calf serum. The solutions were inoculated into 6-well cell culture plates, and three parallel wells were set for each concentration. The cells were cultured in a 37°C, 5% CO2 incubator for 48 h, and the cell morphology and growth changes were observed under an inverted microscope.

[0123] (3) Cell collection: Wash the cells once with PBS, collect them by centrifugation at 1500 rpm for 5 min, and adjust the cell concentration to 1×10 6 / mL, take 1mL of single cell suspension. After centrifugation of the prepared single cell suspension, remove the supernatant.

[0124] (4) Staining: Resuspend the cells with 100 μL of labeling solution, add 2.5 μL of Annexin V and 2.5 μL of PI, and incubate at room temperature for 20 min. Add 400 μL of labeling solution and resuspend the cells.

[0125] (5) On-machine detection: record the red fluorescence at the excitation wavelength of 488 nm and the green fluorescence at 560 nm.

[0126] The flow cytometry results are as follows Figure 2 As shown, compound 7 can significantly induce apoptosis of A549 cells in a dose-dependent manner. The apoptotic effect of compound 7 on the A549 cell line was observed by a control method. After treatment with 5.0, 15.0 and 30.0 μM, the early apoptosis rate of compound 7 gradually increased from 7.06% and 13.30% to 14.60%, respectively, while the control group was 6.13%, and the late apoptosis rate increased from 2.37% and 2.41% to 1.62%, respectively. The proportions of A549 apoptotic cells treated with compound 7 were 9.43% (5.0 μM), 15.71% (15.0 μM) and 16.22% (30.0 μM), respectively. Therefore, this suggests that derivative 7 induces early apoptosis and leads to a significant increase in apoptosis in a concentration-dependent manner.

[0127] Embodiment 17

[0128] The result diagram of the morphological changes of non-small cell lung cancer A549 cells after being treated with different concentrations of norzelaminaldehyde compound 7 for the same time, the specific steps are as follows:

[0129] (1) Cell inoculation: A549 cells in the exponential growth phase were digested and made into a suspension using DMEM medium containing 10% calf serum. Tumor cells were seeded at a cell number of 10×10 4 Each well of the plate was inoculated with 2 mL of the culture medium and cultured at 37°C in a CO2 incubator for 12 h.

[0130] (2) Dosing: The test compound was added with DMSO to prepare a stock solution of a certain concentration, and then prepared into 5.0 μM, 15.0 μM, and 30.0 μM solutions using DMEM medium containing 10% calf serum. The solutions were inoculated into 6-well cell culture plates, and three parallel wells were set for each concentration. The cells were cultured in a 37°C, 5% CO2 incubator for 48 h, and the cell morphology and growth changes were observed under an inverted microscope.

[0131] (3) Staining: Remove the culture medium, wash each well with 1 mL of PBS, add 250 μL of Hoechst33342 to each well, and incubate at room temperature for 5 min in a light-protected environment. Then observe the cells with a fluorescence microscope to detect the differences in the morphological characteristics of A549 cells.

[0132] The experimental results are as follows Figure 3 As shown in the figure, compound 7 has a significant and concentration-dependent effect on the cell morphology of human non-small cell lung cancer cell line A549. After 24 hours of treatment with 5.0 μM norzelaminaldehyde and its derivatives, the morphology of A549 cells changed and the cell density decreased; after 24 hours of treatment with 15.0 μM norzelaminaldehyde and its derivatives, A549 cells showed nuclear condensation and fragmentation, and the cell density decreased; as the drug concentration increased to 30.0 μM, adherent cells began to show apoptotic morphologies such as reduced volume, wrinkled and rounded, and shedding, and the cell density further decreased significantly.

[0133] In summary, the above embodiments disclose 26 nor-zearalenaldehyde derivatives, among which compound 7 has higher activity than nor-zearalenaldehyde at the cellular level. The synthesis conditions of nor-zearalenaldehyde derivatives are mild and they are insensitive to water, oxygen, etc. The reaction raw materials are easily available and inexpensive, the operation is simple, the products are easy to separate and purify, the yield is high, and they are easy to industrialize and produce.

[0134] The CCK8 method was used to determine the inhibitory effect of norzelaminaldehyde derivatives on tumor cell lines. The results showed that most compounds had a novel structure and significant anticancer activity against cancer cell lines such as human non-small cell lung cancer cell line A549, human colon cancer cell line HCT116 and human cervical cancer cell line HeLa (part of the anticancer IC 50 Values ​​are shown in Table 1).

[0135] Flow cytometry detection showed that compound 7 had a cell cycle arresting effect, causing cancer cells A549 to remain in the S phase, thereby achieving an anti-tumor effect; flow cytometry detection showed that compound 7 had the effect of inducing cancer cell apoptosis; through A549 cell apoptosis morphological analysis, compound 7 can block the cancer cell cycle and cause cancer cells to apoptosis through nuclear condensation and fragmentation, cell shrinkage and fragmentation into apoptotic bodies.

[0136] The above description of the embodiments is to facilitate the understanding and use of the present invention by those skilled in the art. It is obvious that those skilled in the art can easily make various modifications to these embodiments and apply the general principles described herein to other embodiments without creative work. Therefore, the present invention is not limited to the above embodiments. Improvements and modifications made by those skilled in the art based on the principles of the present invention without departing from the scope of the present invention should be within the protection scope of the present invention.

Claims

1. Use of norzelaminaldehyde derivatives or pharmaceutically acceptable salts thereof in the preparation of anticancer drugs; The cancers are non-small cell lung cancer and cervical cancer; The norzelaminaldehyde derivative has a general structure as shown in Formula 1a: In formula 1a, R1, R2, R3, and R4 are respectively shown in the following table:

2. The use according to claim 1, characterized in that: The weight percentage of the norzelaminaldehyde derivative in the anticancer drug is 5% to 100%.

3. The use according to claim 2, characterized in that: The anticancer drug also includes a pharmaceutically acceptable carrier or diluent, which is selected from at least one of tablets, capsules, granules, pellets, suspensions, syrups, enteric-coated preparations, emulsion suspensions and injections.

4. Use of a nor-zearalenaldehyde derivative or a pharmaceutically acceptable salt thereof in the preparation of an inhibitor of A549 or HeLa cell proliferation, wherein the nor-zearalenaldehyde derivative is as described in claim 1.

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