A pterosin acid derivative, a preparation method and application thereof

Abstract

The present application relates to the technical field of medicine and chemical industry, in particular to a preparation method and application of a powder back fern acid derivative, wherein powder back fern acid is used as a substrate to react with piperazine under the action of a condensing agent to generate an intermediate product powder back fern piperazine amide, then the intermediate product powder back fern piperazine amide reacts with a cinnamic acid compound or an unsaturated carboxylic acid compound under the action of a condensing agent to generate a series of powder back fern acid derivatives, the powder back fern acid derivatives are tested for cytotoxicity on four common tumor cell lines, and the results show that most of the compounds exhibit obvious anti-proliferation activity on the four tumor cell lines, and the half-inhibition concentration (IC 50 ) of some of the compounds is less than 10 μM. The compound of the present application is simple and fast to synthesize, has high synthesis efficiency, is an effective way to synthesize new compounds by modifying the structure of powder back fern acid, and the powder back fern acid derivatives prepared by synthesis are expected to be applied in the field of anti-tumor drugs.

Description

TECHNICAL FIELD

[0001] The present application relates to the technical field of medicine and chemical industry, in particular to a pyrulopic acid derivative, a preparation method and application thereof. BACKGROUND

[0002] Natural products refer to biological secondary metabolites and endogenous physiological active compounds isolated from animals, plants, marine organisms and microorganisms. Natural products are an important source of drugs, which have a significant effect on anti-tumor and have advantages of high activity, low toxicity and good safety. So far, the structure and anticancer mechanism of many natural products have been elucidated. Due to the advantages of structural diversity, multi-target, high activity and low toxicity, natural products may be one of the best and most valuable resources for multi-target drugs.

[0003] Fern is a kind of natural product, there are about 12000 kinds of ferns in the world, mostly distributed in tropical and subtropical regions, about 2600 kinds in China, more than 300 kinds of which can be used as medicine, mainly distributed in southern and southwestern China. Silver powder back fern is a plant of the genus Cibotium in China. Pyrulopic acid is isolated from silver powder back fern and belongs to the semi-lanosta diterpenoid natural product. Although silver powder back fern has been used as Chinese herbal medicine in China for a long time, scientists also discovered and identified the structure of pyrulopic acid in the 1960s. Studies have shown that pyrulopic acid exhibits various biological activities such as antibacterial, antiparasitic, antiviral and antitumor activities, but the specific mechanism and pharmacological activity are not very clear.

[0004] Cinnamic acid, also known as beta-phenylpropenoic acid and 3-phenyl-2-propenoic acid, is an organic carboxylic acid compound isolated from cinnamon or storax, which is mainly used in the fields of essence and flavor, food additives, pharmaceutical industry, beauty, pesticides, organic synthesis and the like. Cinnamic acid is an effective inhibitor of human non-small cell lung cancer cell A549 and has great application value in anticancer. Piperazine, also known as hexahydropyrazine, is a six-membered ring diamine, which is an important pharmaceutical intermediate. Due to the presence of symmetrical secondary amine structure in the molecule, it is often used as a drug linker to participate in drug synthesis. The molecular structure has a certain rigidity, which can enhance the selectivity of drug and target binding to a certain extent. However, there is no research on the combination of pyrulopic acid and cinnamic acid and piperazine structure unit in the prior art and the anticancer activity.

[0005] In view of the above reasons, the present inventors have obtained the present application after a long period of research. SUMMARY

[0006] The purpose of the present application is to solve the problem of how to combine pyrulopic acid and cinnamic acid for the preparation of anticancer drugs. The present application discloses a pyrulopic acid derivative, a preparation method and application thereof.

[0007] In order to achieve the above object, the application discloses a powdery back fern acid derivative, and a structural general formula of the derivative is as follows:

[0008]

[0009] Wherein, R is cinnamyl, substituted cinnamyl or alkenyl.

[0010] The R is any one of cinnamyl, substituted cinnamyl, 1-methylvinyl or 1,3-pentadienyl. The derivative includes the following specific structures:

[0011]

[0012]

[0013] The application further discloses a preparation method of the powdery back fern acid derivative.

[0014] S1, the powdery back fern acid is dissolved in dichloromethane, then diisopropylethylamine, N,N,N',N'-tetramethylurea hexafluorophosphate and piperazine are sequentially added, the reaction is quenched with water after the reaction is completed, extraction, washing, drying of the organic phase, spin-drying, evaporation to dryness, then the obtained crude product is separated and purified by a silica gel column chromatography method to obtain an intermediate product powdery back fern piperazine amide;

[0015] S2, the powdery back fern piperazine amide obtained in step S1 and an organic carboxylic acid compound are dissolved in dichloromethane, then diisopropylethylamine, N,N,N',N'-tetramethylurea hexafluorophosphate and piperazine are sequentially added, stirring at room temperature, the reaction is quenched with water after the reaction is completed, extraction, washing, drying of the organic phase, spin-drying, evaporation to dryness, then the obtained crude product is separated and purified by a silica gel column chromatography method to obtain the target compound.

[0016] The organic carboxylic acid compound in step S2 is any one of cinnamic acid, substituted cinnamic acid, 2-methyl-1-propenoic acid and sorbic acid.

[0017] The application further discloses application of the powdery back fern acid derivative in preparation of an antitumor drug.

[0018] Compared with the prior art, the application has the beneficial effects that:

[0019] 1. The synthesis process is simple and convenient, and the synthesis efficiency is high, and the purity of the compound after separation is high;

[0020] 2、In the anti-tumor activity screening, most of the compounds show obvious anti-tumor activity, 16 of the 18 derivatives generally show strong inhibitory activity on four cell lines A549 (human non-small cell lung cancer cells), HepG2 (human liver cancer cells), MCF-7 (human breast cancer cells) and MDA-MB-231 (human triple negative breast cancer cells);

[0021] Among them, compounds 3b, 3e, 3f, 3g, 3h, 3j, 3k, 3m and 3n have better inhibitory activity on A549 cell lines, IC 50 values are less than 10 μM, compounds 3a, 3b, 3c, 3d, 3e, 3f, 3g, 3h, 3j, 3k, 3l, 3m and 3n have better inhibitory activity on HepG2 cell lines, IC 50 values are less than 10 μM;

[0022] Compounds 3a, 3b, 3c, 3e, 3f, 3g, 3h, 3j, 3k, 3l, 3m, 3n, 3o, 3p and 3r have better inhibitory activity on MCF-7 cell lines, IC 50 values are less than 10 μM;

[0023] Compounds 3a, 3b, 3c, 3d, 3e, 3f, 3g, 3h, 3j, 3k, 3l, 3m, 3n, 3o and 3r have better inhibitory activity on MDA-MB-231 cell lines, IC 50 values are less than 10 μM;

[0024] Among them, the IC 50 values of compound 3m on A549, HepG2, MCF-7 and MDA-MB-231 cell lines are 2.48±0.49 μM, 1.04±0.01 μM, 0.73±0.28 μM and 1.55±0.12 μM, respectively; among the 18 synthesized derivatives, compound 3m has the strongest cytotoxicity and is the best anti-tumor compound, which can be used as an anti-tumor lead compound for further research;

[0025] 3、The powder back fern acid derivative prepared by the application has potential application in the preparation of anti-tumor drugs. DETAILED DESCRIPTION

[0026] The above and other technical features and advantages of the present application will be more apparent from the following embodiments.

[0027] Example 1

[0028]

[0029] Preparation of (E)-5-([1R,4aS,6R,8aS]-6-hydroxy-5,5,8a-trimethyl-2- methylenedecalin-1-yl)-3-methyl-1-(piperazin-1-yl)pent-2-en-1-one (Compound 2):

[0030] To the powder of 320 mg of pinnatin (3) dissolved in 10 mL of dichloromethane, 195 mg of diisopropylethylamine, 432 mg of N,N,N',N'-tetramethyluronium hexafluorophosphate and 129 mg of piperazine were sequentially added, and stirred at room temperature. After the reaction was completed, the reaction was quenched with water, extracted with ethyl acetate, washed with saturated brine, dried over anhydrous sodium sulfate, and finally, the ethyl acetate was evaporated. The crude product obtained by evaporation was separated and purified by silica gel column chromatography to obtain the intermediate product of pinnatin piperazine amide (2).

[0031] 1 H NMR (400 MHz, CDC13) δ 5.72 (d, J = 12.6 Hz, 1H, 14-H), 4.87 (s, 1H, 17-H), 4.51 (s, 1H, 17-H), 4.01-3.51 (m, 6H, a-H), 3.25 (dd, J = 11.9, 4.2 Hz, 1H, 3-H), 3.21-3.14 (m, 2H, a-H), 2.42 (d, J = 17.0 Hz, 1H, 7-H), 2.27 (t, J = 15.3 Hz, 12-H), 2.02-1.92 (m, 2H, 7, 12-H), 1.90 (s, 3H, 16-CH3), 1.82-1.47 (m, 7H, 1-H, 2-CH2, 6-H, 9-H, 11-CH2), 1.40 (qd, J = 12.9, 4.3 Hz, 1H, 6-H), 1.16 (t, J = 13.2 Hz, 1H, 1-H), 1.10-1.03 (m, 1H, 5-H), 1.00 (s, 3H, 18-CH3), 0.77 (s, 3H, 19-CH3), 0.69 (s, 3H, 20-CH3).

[0032] Example 2

[0033]

[0034] Preparation of (E)-1-(4-cinnamoylpiperazin-1-yl)-5-((1R,4aS,6R,8aS)-6-hydroxy-5,5,8a- trimethyl-2-methylenedecalin-1-yl)-3-methylpent-2-en-1-one (Compound 3a):

[0035] To the intermediate product, powdered piperazineamide of pyknosorus acid (2) 19 mg was added 2 mL of dichloromethane to dissolve, then diisopropylethylamine 9.7 mg, N,N,N',N'-tetramethyluronium hexafluorophosphate 22.8 mg and cinnamic acid 8.9 mg were added in turn, stirred at room temperature, after the reaction was complete, the reaction was quenched with water, extracted with ethyl acetate, washed with saturated brine, dried over anhydrous sodium sulfate, and finally the ethyl acetate was rotary evaporated. After evaporation, the crude product was separated and purified by silica gel column chromatography to obtain the target compound 3a. Yield: 81%, white solid; melting point: 84-85°C; mass spectral data: C 33 H 46 N2O3[M+H] + Calcd: 519.3581, Found: 519.3555.

[0036] 1 H NMR (400 MHz, CDC13) δ 7.71 (d, J = 15.3 Hz, 1H, d-H), 7.55 - 7.50 (m, 2H, 2 x Ph-H), 7.39 - 7.35 (m, 3H, 3 x Ph-H), 6.86 (d, J = 15.4 Hz, 1H, c-H), 5.75 (s, 1H, 14-H), 4.87 (s, 1H, 17-H), 4.52 (s, 1H, 17-H), 3.74 - 3.58 (m, 8H, a-H), 3.25 (dd, J = 11.6, 4.4 Hz, 1H, 3-H), 2.41 (ddd, J = 12.7, 4.2, 2.4 Hz, 1H, 7-H), 2.30 - 2.24 (m, 1H, 12-H), 1.99 - 1.92 (m, 2H, 7-H, 12-H), 1.91 (s, 3H, 16-CH3), 1.82 - 1.53 (m, 7H, 1-H, 2-CH2, 6-H, 9-H, 11-CH2), 1.40 (qd, J = 13.0, 4.3 Hz, 1H, 6-H), 1.16 (td, J = 13.2, 3.9 Hz, 1H, 1-H), 1.07 (dd, J = 12.5, 2.7 Hz, 1H, 5-H), 0.99 (s, 3H, 18-CH3), 0.77 (s, 3H, 19-CH3), 0.69 (s, 3H, 20-CH3).

[0037] Example 3

[0038]

[0039] Preparation of (E)-5-((1R,4aS,6R,8aS)-6-hydroxy-5,5,8a-trimethyl-2- methylenedecahydro naphthalen-1-yl)-3-methyl-1-(4-((E)-3-(p-tolyl)acryloyl)piperazin-1- yl)pent-2-en-1-one (Compound 3b):

[0040] To the intermediate powder pellia piperazine amide (2) 19 mg was added 2 mL of dichloromethane to dissolve, then diisopropyl ethyl amine 9.7 mg, N,N,N',N'-tetramethyl urea hexafluorophosphate 22.8 mg and 4-methyl cinnamic acid 9.7 mg were added in turn, stirred at room temperature, after the reaction was completed, the reaction was quenched with water, extracted with ethyl acetate, washed with saturated brine, dried over anhydrous sodium sulfate, and finally the ethyl acetate was rotary evaporated. The crude product obtained after evaporation was separated and purified by silica gel column chromatography to obtain the target compound 3b. Yield: 78%, white solid; melting point: 186-187°C; mass spectral data: C 34 H 48 N2O3[M+H] + Calcd: 533.3738, Found: 533.3727.

[0041] 1 H NMR (400 MHz, CDC13) δ 7.68 (d, J = 15.3 Hz, 1H, d-H), 7.42 (d, J = 7.9 Hz, 2H, 2 x Ph-H), 7.18 (d, J = 7.8 Hz, 2H, 2 x Ph-H), 6.81 (d, J = 15.4 Hz, 1H, c-H), 5.75 (s, 1H, 14-H), 4.87 (s, 1H, 17-H), 4.53 (s, 1H, 17-H), 3.76 - 3.54 (m, 8H, a-H), 3.25 (dd, J = 11.7, 4.4 Hz, 1H, 3-H), 2.42 (ddd, J = 12.7, 4.3, 2.4 Hz, 1H, 7-H), 2.37 (s, 3H, Ph-CH3), 2.32 - 2.21 (m, 1H, 12-H), 2.01 - 1.91 (m, 2H, 7-H, 12-H), 1.91 (s, 3H, 16-CH3), 1.78 - 1.47 (m, 7H, 1-H, 2-CH2, 6-H, 9-H, 11-CH2), 1.40 (qd, J = 12.8, 4.3 Hz, 1H, 6-H), 1.17 (td, J = 13.2, 3.8 Hz, 1H, 1-H), 1.07 (dd, J = 12.5, 2.7 Hz, 1H, 5-H), 0.99 (s, 3H, 18-CH3), 0.77 (s, 3H, 19-CH3), 0.70 (s, 3H, 20-CH3).

[0042] Example 4

[0043]

[0044] Preparation of (E)-1-(4-((E)-3-(4-fluorophenyl)acryloyl)piperazin-1-yl)-5-((1R,4aS,6R,8aS)-6-hydroxy-5,5,8a-trimethyl-2-oxo- octahydrophenalen-1-yl)-3-methylpent-2-en-1-one (Compound 3c):

[0045] To the intermediate powder pellitorinamide (2) 19 mg was dissolved in 2 mL of dichloromethane, then diisopropylethylamine 9.7 mg, N,N,N',N'-tetramethyluronium hexafluorophosphate 22.8 mg and 4-fluorocinnamic acid 10.0 mg were added in turn, stirred at room temperature, after the reaction was completed, the reaction was quenched with water, extracted with ethyl acetate, washed with saturated brine, dried over anhydrous sodium sulfate, and finally the ethyl acetate was rotary evaporated. The crude product obtained after evaporation was separated and purified by silica gel column chromatography to obtain the target compound 3c. Yield: 78%, white solid, melting point: 61-62°C; mass spectral data: C 33 H 45 FN2O3[M+H] + , calculated value: 537.3487, measured value: 537.3463.

[0046] 1H NMR (400 MHz, CDC13) δ 7.67 (d, J = 15.4 Hz, 1H, d-H), 7.56 - 7.47 (m, 2H, 2 x Ph-H), 7.12 - 7.03 (m, 2H, 2 x Ph-H), 6.79 (d, J = 15.3 Hz, 1H, c-H), 5.76 (s, 1H, 14-H), 4.87 (s, 1H, 17-H), 4.52 (s, 1H, 17-H), 3.79 - 3.53 (m, 8H, a-H), 3.25 (dd, J = 11.7, 4.4 Hz, 1H, 3-H), 2.42 (ddd, J = 12.9, 4.3, 2.4 Hz, 1H, 7-H), 2.33 - 2.21 (m, 1H, 12-H), 1.99 - 1.92 (m, 2H, 7-H, 12-H), 1.91 (s, 3H, 16-CH3), 1.84 - 1.58 (m, 7H, 1-H, 2-CH2, 6-H, 9-H, 11-CH2), 1.40 (qd, J = 12.5, 2.7 Hz, 1H, 6-H), 1.17 (td, J = 13.1, 3.9 Hz, 1H, 1-H), 1.07 (dd, J = 12.5, 2.7 Hz, 1H, 5-H), 0.99 (s, 3H, 18-CH3), 0.77 (s, 3H, 19-CH3), 0.70 (s, 3H, 20-CH3).

[0047] Example 5

[0048]

[0049] Preparation of (E)-1-(4-((E)-3-(2-chlorophenyl)acryloyl)piperazin-1-yl)-5-((1R,4aS,6R,8aS)-6-hydroxy-5,5,8a-trimethyl-2-oxo-1,2,4,6,7,8,9,9a-octahydropicen-1-yl)-3-methylpent-2-en-1-one (Compound 3d):

[0050] To the intermediate powder pinnatinamide (2) 19 mg was added 2 mL of dichloromethane to dissolve, then diisopropylethylamine 9.7 mg, N,N,N',N'-tetramethyluronium hexafluorophosphate 22.8 mg and 2-chlorocinnamic acid 10.9 mg were added in turn, stirred at room temperature, after the reaction was completed, the reaction was quenched with water, extracted with ethyl acetate, washed with saturated brine, dried over anhydrous sodium sulfate, and finally the ethyl acetate was rotary evaporated. The crude product obtained after evaporation was separated and purified by silica gel column chromatography to obtain the target compound 3d. Yield: 83%, white solid; melting point: 155-157°C; mass spectral data: C 33 H 45 N2O3[M+H] +Calculated: 553.3191, Found: 553.3190. 1 H NMR (400 MHz, CDC13) δ 8.03 (d, J = 15.3 Hz, 1H, d-H), 7.62 - 7.56 (m, 1H, Ph-H), 7.43 - 7.38 (m, 1H, Ph-H), 7.32 - 7.25 (m, 2 x Ph-H), 6.84 (d, J = 15.5 Hz, 1H, c-H), 5.75 (s, 1H, 14-H), 4.87 (s, 1H, 17-H), 4.52 (s, 1H, 17-H), 3.80 - 3.50 (m, 8H, a-H), 3.24 (dd, J = 11.7, 4.4 Hz, 1H, 3-H), 2.41 (ddd, J = 12.7, 4.2, 2.3 Hz, 1H, 7-H), 2.32 - 2.21 (m, 1H, 12-H), 2.01 - 1.91 (m, 2H, 7-H, 12-H), 1.90 (s, 3H, 16-CH3), 1.81 - 1.55 (m, 7H 1-H, 2-CH2, 6-H, 9-H, 11-CH2), 1.39 (qd, J = 12.9, 4.3 Hz, 1H, 6-H), 1.16 (td, J = 13.2, 3.9 Hz, 1H, 1-H), 1.07 (dd, J = 12.5, 2.7 Hz, 1H, 5-H), 0.99 (s, 3H, 18-CH3), 0.77 (s, 3H, 19-CH3), 0.69 (s, 3H, 20-CH3).

[0051] Example 6

[0052]

[0053] Preparation of (E)-1-(4-((E)-3-(3-chlorophenyl)acryloyl)piperazin-1-yl)-5-((1R,4aS,6R,8aS)-6-hydroxy-5,5,8a-trimethyl-2-oxotetrahydroharman-1-yl)-3-methylpent-2- en-1-one (Compound 3e):

[0054] To the intermediate powder pinnatinamide (2) 19 mg was added 2 mL of dichloromethane to dissolve, then diisopropylethylamine 9.7 mg, N,N,N',N'-tetramethyluronium hexafluorophosphate 22.8 mg and 3-chlorocinnamic acid 10.9 mg were added in turn, stirred at room temperature, after the reaction was completed, the reaction was quenched with water, extracted with ethyl acetate, washed with saturated brine, dried over anhydrous sodium sulfate, and finally the ethyl acetate was rotary evaporated. The crude product obtained after evaporation was separated and purified by silica gel column chromatography to obtain the target compound 3e. Yield: 83%, white solid, melting point: 155-157°C; mass spectral data: C 33 H45 N2O3[M+H] + Calculated: 553.3191, Found: 553.3190.

[0055] 1 H NMR (400 MHz, CDC13) δ 7.63 (d, J = 15.4 Hz, 1H, d-H), 7.51 (s, 1H, Ph-H), 7.40 - 7.28 (m, 3H, 3 x Ph-H), 6.86 (d, J = 15.4 Hz, 1H, c-H), 5.75 (s, 1H, 14-H), 4.87 (d, J = 1.7 Hz, 1H, 17-H), 4.52 (s, 1H, 17-H), 3.78 - 3.50 (m, 8H, a-H), 3.24 (dd, J = 11.7, 4.4 Hz, 1H, 3-H), 2.41 (ddd, J = 12.8, 4.3, 2.4 Hz, 1H, 7-H), 2.33 - 2.19 (m, 1H, 12-H), 1.94 (m, 2H, 7-H, 12-H), 1.90 (s, 3H, 16-CH3), 1.83 - 1.57 (m, 7H, 1-H, 2-CH2, 6-H, 9-H, 11-CH2), 1.39 (qd, J = 12.9, 4.3 Hz, 1H, 6-H), 1.16 (td, J = 13.2, 3.9 Hz, 1H, 1-H), 1.07 (dd, J = 12.5, 2.8 Hz, 1H, 5-H), 0.99 (s, 3H, 18-CH3), 0.77 (s, 3H, 19-CH3), 0.69 (s, 3H, 20-CH3).

[0056] Example 7

[0057]

[0058] Preparation of (E)-l-(4-((E)-3-(4-chlorophenyl)acryloyl)piperazin-l-yl)-5-((lR,4aS,6R,8aS)-6-hydroxy-5,5,8a-trimethyl-2-oxo- octahydro naphthalen- 1 -yl)-3 -methylpent-2-en- 1 -one (Compound 3f):

[0059] To the intermediate powder pumiloside piperazine amide (2) 19 mg in 2 mL of dichloromethane, diisopropylethylamine 9.7 mg, N,N,N',N'-tetramethyluronium hexafluorophosphate 22.8 mg and 4-chlorocinnamic acid 10.9 mg were added successively, stirred at room temperature, after the reaction was completed, the reaction was quenched with water, extracted with ethyl acetate, washed with saturated brine, dried over anhydrous sodium sulfate, and finally the ethyl acetate was rotary evaporated. The crude product obtained after evaporation was separated and purified by silica gel column chromatography to obtain the target compound 3f. Yield: 82%, white solid, melting point: 185-186°C; mass spectral data: C 33 H 45 ClN2O3[M+H] + , calculated: 553.3191, found: 553.3186.

[0060] 1 H NMR (400 MHz, CDC13) δ 7.65 (d, J = 15.4 Hz, 1H, d-H), 7.50 - 7.30 (m, 4H, 4 x Ph-H), 6.84 (d, J = 15.1 Hz, 1H, c-H), 5.75 (s, 1H, 14-H), 4.87 (s, 1H, 17-H), 4.52 (s, 1H, 17-H), 3.80 - 3.47 (m, 8H, a-H), 3.25 (dd, J = 11.7, 4.4 Hz, 1H, 3-H), 2.41 (ddd, J = 12.7, 4.3, 2.4 Hz, 1H, 7-H), 2.31 - 2.22 (m, 1H, 12-H), 1.99 - 1.91 (m, 2H, 7-H, 12-H), 1.90 (s, 3H, 16-CH3), 1.82 - 1.47 (m, 7H, 1-H, 2-CH2, 6-H, 9-H, 11-CH2), 1.39 (qd, J = 12.9, 4.3 Hz, 1H, 6-H), 1.16 (td, J = 13.2, 3.8 Hz, 1H, 1-H), 1.07 (dd, J = 12.5, 2.7 Hz, 1H, 5-H), 0.99 (s, 3H, 18-CH3), 0.77 (s, 3H, 19-CH3), 0.69 (s, 3H, 20-CH3).

[0061] Example 8

[0062]

[0063] (E)-1-(4-((E)-3-(4-bromophenyl)acryloyl)piperazin-1-yl)-5-((1R,4aS,6R,8aS)-6-hydroxy-5,5,8a-trimethyl-2-oxo- 1,2,4,5,6,7,8,9,9a-hexahydronaphth-1-yl)-3-methylpent-2-en-1-one (Compound 3g)

[0064] To the intermediate product, powder of pellitorinamide (2) 19 mg was added 2 mL of dichloromethane to dissolve, then diisopropylethylamine 9.7 mg, N,N,N',N'-tetramethyluronium hexafluorophosphate 22.8 mg and 4-bromocinnamic acid 15.3 mg were added in turn, stirred at room temperature, after the reaction was completed, the reaction was quenched with water, extracted with ethyl acetate, washed with saturated brine, dried over anhydrous sodium sulfate, and finally the ethyl acetate was rotary evaporated. After evaporation, the crude product was separated and purified by silica gel column chromatography to obtain the target compound 3 g. Yield: 74%, white solid, melting point: 177-178°C; mass spectrometry data: C 33 H 45 BrN2O3[M+H] + Calculated: 597.2686, Found: 597.2690.

[0065] 1 H NMR (400 MHz, CDC13) δ 7.63 (d, J = 15.0 Hz, 1H, d-H), 7.53 - 7.36 (m, 4H, 4 x Ph-H), 6.85 (d, J = 15.3 Hz, 1H, c-H), 5.75 (s, 1H, 14-H), 4.87 (s, 1H, 17-H), 4.52 (s, 1H, 17-H), 3.69 (s, 8H, a-H), 3.25 (dd, J = 11.6, 4.2 Hz, 1H, 3-H), 2.42 (ddd, J = 12.8, 4.2, 2.3 Hz, 1H, 7-H), 2.32 - 2.20 (m, 1H, 12-H), 1.96 (dd, J = 14.4, 4.5 Hz, 2H, 7-H, 12-H), 1.91 (s, 3H, 16-CH3), 1.83 - 1.47 (m, 7H, 1-H, 2-CH2, 6-H, 9-H, 11-CH2), 1.40 (qd, J = 13.0, 4.2 Hz, 1H, 6-H), 1.17 (td, J = 13.2, 3.6 Hz, 1H, 1-H), 1.07 (dd, J = 12.5, 2.7 Hz, 1H, 5-H), 0.99 (s, 3H, 18-CH3), 0.77 (s, 3H, 19-CH3), 0.70 (s, 3H, 20-CH3).

[0066] Example 9

[0067]

[0068] (E)-1-(4-((E)-3-(3,4-difluorophenyl)acryloyl)piperazin-1-yl)-5-((1R,4aS,6R,8aS)-6-hydroxy-5,5,8a-trimethyl-2-oxo- 1,3,4,5,6,7,8,9,9a-hexahydrocyclohexa[c]benz[f]inden-1-yl)-3-methylpent-2- en-1-one (Compound 3h)

[0069] To the intermediate powder pellitorinamide (2) 19 mg in 2 mL of dichloromethane was added, followed by diisopropylethylamine 9.7 mg, N,N,N',N'-tetramethyluronium hexafluorophosphate 22.8 mg and 3,4-difluorocinnamic acid 11.0 mg successively, stirred at room temperature, quenched with water after the reaction was completed, extracted with ethyl acetate, washed with saturated brine, dried over anhydrous sodium sulfate, and finally the ethyl acetate was evaporated. The crude product obtained after evaporation was separated and purified by silica gel column chromatography to obtain the target compound 3h. Yield: 95%, white solid, melting point: 83-85°C; mass spectral data: C 33 H 44 F2N2O3[M+H] + Calcd: 555.3393, Found: 555.3372.

[0070] 1 H NMR (400 MHz, CDC13) δ 7.60 (d, J = 15.4 Hz, 1H, d-H), 7.34 (dd, J = 10.7, 7.6 Hz, 1H, Ph-H), 7.24 - 7.22 (m, 1H, Ph-H), 7.27 - 7.10 (m, 1H, Ph-H), 6.78 (d, J = 15.4 Hz, 1H, c-H), 5.75 (s, 1H, 14-H), 4.87 (s, 1H, 17-H), 4.52 (s, 1H, 17-H), 3.78 - 3.48 (m, 8H, a-H), 3.25 (dd, J = 11.7, 4.4 Hz, 1H, 3-H), 2.41 (ddd, J = 12.7, 4.3, 2.4 Hz, 1H, 7-H), 2.33 - 2.17 (m, 1H, 12-H), 1.99 - 1.92 (m, 2H, 7-H, 12-H), 1.91 (s, 3H, 16-CH3), 1.81 - 1.56 (m, 7H, 1-H, 2-CH2, 6-H, 9-H, 11-CH2), 1.39 (qd, J = 12.9, 4.3 Hz, 1H, 6-H), 1.16 (td, J = 13.2, 3.8 Hz, 1H, 1-H), 1.07 (dd, J = 12.5, 2.8 Hz, 1H, 5-H), 0.99 (s, 3H, 18-CH3), 0.77 (s, 3H, 19-CH3), 0.69 (s, 3H, 20-CH3).

[0071] Example 10

[0072]

[0073] Preparation of (E)-1-(4-((E)-3-(2,5-dichlorophenyl)acryloyl)piperazin-1-yl)-5-((1R,4aS,6R,8aS)-6-hydroxy-5,5,8a-trimethyl-2-oxo- octahydro naphthalen-1-yl)-3-methylpent-2-en-1-one (Compound 3i):

[0074] To the intermediate powder pellitorinamide (2) 19 mg was dissolved in 2 mL of dichloromethane, then diisopropylethylamine 9.7 mg, N,N,N',N'-tetramethyluronium hexafluorophosphate 22.8 mg and 2,5-dichlorocinnamic acid 12.9 mg were added in sequence, stirred at room temperature, after the reaction was completed, the reaction was quenched with water, extracted with ethyl acetate, washed with saturated brine, dried over anhydrous sodium sulfate, and finally the ethyl acetate was rotary evaporated. The crude product obtained after evaporation was separated and purified by silica gel column chromatography to obtain the target compound 3i. Yield: 82%, white solid, melting point: 160-162°C; Mass spectral data: C 33 H 44 Cl2N2O3[M+H] + Calcd: 587.2802, Found: 587.2802.

[0075] 1H NMR (400 MHz, CDC13) δ 7.95 (d, J = 15.4 Hz, 1H, d-H), 7.56 (d, J = 2.4 Hz, 1H, Ph-H), 7.35 (d, J = 8.5 Hz, 1H, Ph-H), 7.28 - 7.25 (m, 1H, Ph-H), 6.83 (d, J = 15.5 Hz, 1H, c-H), 5.76 (s, 1H, 14-H), 4.87 (d, J = 1.6 Hz, 1H, 17-H), 4.52 (s, 1H, 17-H), 3.66 (s, 8H, a-H), 3.25 (dd, J = 11.7, 4.4 Hz, 1H, 3-H), 2.41 (ddd, J = 12.7, 4.2, 2.4 Hz, 1H, 7-H), 2.33 - 2.22 (m, 1H, 12-H), 2.00 - 1.92 (m, 2H, 7-H, 12-H), 1.91 (s, 3H, 16-CH3), 1.83 - 1.43 (m, 7H, 1-H, 2-CH2, 6-H, 9-H, 11-CH2), 1.38 (qd, J = 12.5, 2.7 Hz, 1H, 6-H), 1.17 (td, J = 13.2, 4.1 Hz, 1H, 1-H), 1.07 (dd, J = 12.5, 2.7 Hz, 1H, 5-H), 0.99 (s, 3H, 18-CH3), 0.77 (s, 3H, 19-CH3), 0.69 (s, 3H, 20-CH3).

[0076] Example 11

[0077]

[0078] Preparation of (E)-l-(4-((E)-3-(3,4-dichlorophenyl)acryloyl)piperazin-l-yl)-5-((lR,4aS,6R,8aS)-6-hydroxy-5,5,8a-trimethyl-2-oxo- 1,2,4,5,6,7,8,9,9a,10-decahydrophenacene-l-yl)-3-methylpent-2-en-l-one (Compound 3j):

[0079] To the intermediate powder pellitorinamide (2) 19 mg was dissolved in 2 mL of dichloromethane, then diisopropylethylamine 9.7 mg, N,N,N',N'-tetramethyluronium hexafluorophosphate 22.8 mg and 3,4-dichlorocinnamic acid 12.9 mg were added in turn, stirred at room temperature, after the reaction was completed, the reaction was quenched with water, extracted with ethyl acetate, washed with saturated brine, dried over anhydrous sodium sulfate, and finally the ethyl acetate was rotary evaporated. The crude product obtained after evaporation was separated and purified by silica gel column chromatography to obtain the target compound 3j. Yield: 94%, white solid, melting point: 84-86°C; mass spectral data: C 33 H 44 Cl2N2O3[M+H]+ Calculated: 587.2808, Found: 587.2799.

[0080] 1 H NMR (400 MHz, CDC13) δ 7.59 (d, J = 15.3 Hz, 2H, 2 x Ph-H), 7.45 (d, J = 8.3 Hz, 1H, Ph-H), 7.34 (d, J = 8.2 Hz, 1H, d-H), 6.85 (d, J = 15.2 Hz, 1H, c-H), 5.75 (s, 1H, 14-H), 4.87 (s, 1H, 17-H), 4.52 (s, 1H, 17-H), 3.85 - 3.48 (m, 8H, a-H), 3.25 (dd, J = 11.7, 4.4 Hz, 1H, 3-H), 2.41 (ddd, J = 12.8, 4.3, 2.4 Hz, 1H, 7-H), 2.27 (m, J = 14.0, 9.8, 3.5 Hz, 1H, 12-H), 1.99 - 1.92 (m, 2H, 7-H, 12-H), 1.91 (s, 3H, 16-CH3), 1.82 - 1.55 (m, 7H, 1-H, 2-CH2, 6-H, 9-H, 11-CH2), 1.38 (qd, J = 12.4, 5.6 Hz, 1H, 6-H), 1.16 (td, J = 13.1, 3.8 Hz, 1H, 1-H), 1.07 (dd, J = 12.5, 2.7 Hz, 1H, 5-H), 0.99 (s, 3H, 18-CH3), 0.77 (s, 3H, 19-CH3), 0.69 (s, 3H, 20-CH3).

[0081] Example 12

[0082]

[0083] Preparation of (E)-5-((1R,4aS,6R,8aS)-6-hydroxy-5,5,8a-trimethyl-2- methylenedecahydronaphthalen-1-yl)-1-(4-((E)-3-(4-methoxyphenyl)propenoyl)piperazin-1-yl)-3-methylpent-2-en-1-one (Compound 3k):

[0084] To the intermediate powder pellitorinamide (2) 19 mg in 2 mL of dichloromethane, diisopropylethylamine 9.7 mg, N,N,N',N'-tetramethyluronium hexafluorophosphate 22.8 mg and 4-methoxycinnamic acid 10.7 mg were added successively, and stirred at room temperature. After the reaction was completed, the reaction was quenched with water, extracted with ethyl acetate, washed with saturated brine, dried over anhydrous sodium sulfate, and finally the ethyl acetate was evaporated. The crude product obtained after evaporation was separated and purified by silica gel column chromatography to obtain the target compound 3k. Yield: 88%, white solid, melting point: 170-172°C; mass spectrometry data: C 34 H 48 N2O4[M+H] + Calcd: 549.3687, Found: 549.3671.

[0085] 1 H NMR (400 MHz, CDC13) δ 7.67 (d, J = 15.2 Hz, 1H, d-H), 7.52 - 7.44 (m, 2H, 2 x Ph-H), 6.90 (d, J = 8.4 Hz, 2H, 2 x Ph-H), 6.73 (d, J = 15.2 Hz, c-H), 5.75 (s, 1H, 14-H), 4.87 (s, 1H, 17-H), 4.53 (s, 1H, 17-H), 3.83 (s, 3H, Ph-OCH3), 3.63 (d, J = 54.3 Hz, 8H, a-H), 3.25 (dd, J = 11.7, 4.3 Hz, 1H, 3-H), 2.41 (ddd, J = 12.7, 4.2, 2.4 Hz, 1H, 7-H), 2.30 - 2.23 (m, 1H, 12-H), 2.01 - 1.92 (m, 2H, 7-H, 12-H), 1.90 (s, 3H, 16-CH3), 1.84 - 1.50 (m, 7H, 1-H, 2-CH2, 6-H, 9-H, 11-CH2), 1.40 (qd, J = 12.9, 4.2 Hz, 1H, 6-H), 1.17 (td, J = 13.3, 3.9 Hz, 1H, 1-H), 1.07 (dd, J = 12.5, 2.7 Hz, 1H, 5-H), 0.99 (s, 3H, 18-CH3), 0.77 (s, 3H, 19-CH3), 0.70 (s, 3H, 20-CH3).

[0086] Example 13

[0087]

[0088] Preparation of (E)-5-((1R,4aS,6R,8aS)-6-hydroxy-5,5,8a-trimethyl-2- methylenedecahydro naphthalen-1-yl)-3-methyl-1-(4-((E)-3-(4- (methylthio)phenyl)acryloyl)piperazin-1-yl)pent-2-en-1-one (Compound 3l):

[0089] To the intermediate powder pellitorinamide (2) 19 mg was added 2 mL of dichloromethane to dissolve, then diisopropylethylamine 9.7 mg, N,N,N',N'-tetramethyluronium hexafluorophosphate 22.8 mg and 4-methylthiocinnamic acid 11.6 mg were added in turn, stirred at room temperature, after the reaction was completed, the reaction was quenched with water, extracted with ethyl acetate, washed with saturated brine, dried over anhydrous sodium sulfate, and finally the ethyl acetate was rotary evaporated. The crude product obtained after evaporation was separated and purified by silica gel column chromatography to obtain the target compound 3l. Yield: 92%, white solid, melting point: 195-196°C; mass spectral data: C 34 H 48 N2O3S[M+H] + Calcd: 565.3458, Found: 565.3450.

[0090] 1 H NMR (400 MHz, CDC13) δ 7.66 (d, J = 15.3 Hz, 1H, d-H), 7.44 (d, J = 8.1 Hz, 2H, 2 x Ph-H), 7.22 (d, J = 8.2 Hz, 2H, 2 x Ph-H), 6.81 (d, J = 15.3 Hz, 1H, c-H), 5.75 (s, 1H, 14-H), 4.87 (s, 1H, 17-H), 4.52 (s, 1H, 17-H), 3.63 (m, 8H, a-H), 3.25 (dd, J = 11.6, 4.4 Hz, 1H, 3-H), 2.50 (s, 3H, Ph-SCH3), 2.42 (ddd, J = 12.7, 4.2, 2.4 Hz, 1H, 7-H), 2.33 - 2.21 (m, 1H, 12-H), 2.01 - 1.92 (m, 2H, 7-H, 12-H), 1.91 (s, 3H, 16-CH3), 1.84 - 1.55 (m, 7H, 1-H, 2-CH2, 6-H, 9-H, 11-CH2), 1.40 (qd, J = 12.8, 4.3 Hz, 2H, 6-H), 1.16 (td, J = 13.1, 3.8 Hz, 1H, 1-H), 1.07 (dd, J = 12.5, 2.7 Hz, 1H, 5-H), 0.99 (s, 3H, 20-CH3), 0.77 (s, 3H, 19-CH3), 0.69 (s, 3H, 20-CH3).

[0091] Example 14

[0092]

[0093] (E)-5-((1R,4aS,6R,8aS)-6-hydroxy-5,5,8a-trimethyl-2- methylenedecalin-1-yl)-3-methyl-1-(4-((E)-3-(3,4,5-trimethoxyphenyl)allyl) piperazin-1-yl)pent-2-en-1-one (Compound 3m) was prepared as follows:

[0094] To the intermediate powder pellitorinamide (2) 19 mg was dissolved in 2 mL of dichloromethane, followed by the addition of diisopropylethylamine 9.7 mg, N,N,N',N'-tetramethyluronium hexafluorophosphate 22.8 mg and 3,4,5-trimethoxycinnamic acid 14.2 mg, stirred at room temperature, after the reaction was completed, quenched with water, extracted with ethyl acetate, washed with saturated brine, dried over anhydrous sodium sulfate, and finally the ethyl acetate was rotary evaporated. The crude product obtained after evaporation was separated and purified by silica gel column chromatography to obtain the target compound 3m. Yield: 79%, white solid, melting point: 81-83 °C; Mass spectral data: C 36 H 52 N2O6[M+H] + Calcd: 609.3898, Found: 609.3893.

[0095] 1H NMR (400 MHz, CDC13) δ 7.58 (d, J = 15.3 Hz, 1H, d-H), 6.76 (d, J = 16.3 Hz, 3H, c-H, 2 x Ph-H), 5.74 (s, 1H, 14-H), 4.85 (s, 1H, 17-H), 4.50 (s, 1H, 17-H), 3.88 (s, 6H, 2 x Ph-OCH3), 3.85 (s, 3H, Ph-OCH3), 3.68 (m, 8H, a-H), 3.23 (dd, J = 11.7, 4.4 Hz, 1H, 3-H), 2.39 (ddd, J = 12.8, 4.3, 2.4 Hz, 1H, 7-H), 2.24 (m, J = 14.0, 9.6, 3.7 Hz, 1H, 12-H), 1.97 - 1.88 (m, 2H, 7-H, 12-H), 1.87 (s, 3H, 16-CH3), 1.81 - 1.48 (m, 7H, 1-H, 2-CH2, 6-H, 9-H, 11-CH2), 1.38 (qd, J = 12.4, 5.6 Hz, 1H, 6-H), 1.14 (td, J = 13.1, 3.9 Hz, 1H, 1-H), 1.05 (dd, J = 12.4, 2.7 Hz, 1H, -H), 0.97 (s, 3H, 18-CH3), 0.75 (s, 3H, 19-CH3), 0.67 (s, 3H, 20-CH3).

[0096] Example 15

[0097]

[0098] Preparation of (E)-5-((1R,4aS,6R,8aS)-6-hydroxy-5,5,8a-trimethyl-2- methylenedecahydro naphthalen-1-yl)-3-methyl-1-(4-((E)-3-(4- (trifluoromethyl)phenyl)prop-2-enoyl)piperazin-1-yl)pent-2-en-1-one (Compound 3n):

[0099] To the intermediate powder pinnatin piperazine amide (2) 19 mg was added 2 mL of dichloromethane to dissolve, then diisopropylethylamine 9.7 mg, N,N,N',N'-tetramethyluronium hexafluorophosphate 22.8 mg and 4-(trifluoromethyl)cinnamic acid 13.0 mg were added in turn, stirred at room temperature, after the reaction was completed, the reaction was quenched with water, extracted with ethyl acetate, washed with saturated brine, dried over anhydrous sodium sulfate, and finally the ethyl acetate was rotary evaporated. The crude product obtained after evaporation was separated and purified by silica gel column chromatography to obtain the target compound 3n. Yield: 85%, white solid, melting point: 139-140°C; mass spectral data: C 34 H 45 F3N2O3[M+H]+ Calculated: 587.3455, Found: 587.3442.

[0100] 1 H NMR (400 MHz, CDC13) δ 7.71 (d, J = 15.4 Hz, 1H, d-H), 7.66 - 7.60 (m, 4H, 2 x Ph-H), 6.93 (d, J = 15.4 Hz, 1H), 5.76 (s, 1H, c-H, 14-H), 4.88 (s, 1H, 17-H), 4.53 (s, 1H, 17-H), 3.80 - 3.55 (m, 8H, a-H), 3.25 (dd, J = 11.7, 4.4 Hz, 1H, 3-H), 2.42 (ddd, J = 12.8, 4.3, 2.4 Hz, 1H, 7-H), 2.32 - 2.22 (m, 1H, 12-H), 2.00 - 1.93 (m, 2H, 7-H, 12-H), 1.92 (s, 3H, 16-CH3), 1.82 - 1.48 (m, 7H, 1-H, 2-CH2, 6-H, 9-H, 11-CH2), 1.40 (qd, J = 13.0, 4.3 Hz, 1H, 6-H), 1.17 (td, J = 13.2, 3.8 Hz, 1, 1-H), 1.08 (dd, J = 12.5, 2.8 Hz, 1H, 5-H), 1.00 (s, 3H, 18-CH3), 0.78 (s, 3H, 19-CH3), 0.70 (s, 3H, 20-CH3).

[0101] Example 16

[0102]

[0103] Preparation of (E)-5-((1R,4aS,6R,8aS)-6-hydroxy-5,5,8a-trimethyl-2- methylenedecahydro naphthalen-1-yl)-3-methyl-1-(4-((E)-3-(4-nitrophenyl) acryloyl)piperazin-1-yl)pent-2-en-1-one (Compound 3o):

[0104] To the intermediate powder pellia acid piperazine amide (2) 19 mg was added 2 mL of dichloromethane to dissolve, then diisopropyl ethyl amine 9.7 mg, N,N,N',N'-tetramethyl urea hexafluorophosphate 22.8 mg and 4-nitro acid 11.6 mg were added in turn, stirred at room temperature, after the reaction was completed, the reaction was quenched with water, extracted with ethyl acetate, washed with saturated brine, dried over anhydrous sodium sulfate, and finally the ethyl acetate was rotary evaporated. After evaporation, the crude product was separated and purified by silica gel column chromatography to obtain the target compound 3o. Yield: 79%, white solid, melting point: 108-110°C; Mass spectral data: C 33 H45 N3O5[M+H] + Calculated: 564.3432, Found: 564.3427.

[0105] 1 H NMR (400 MHz, CDC13) δ 8.24 (d, J = 8.6 Hz, 2H, 2 x Ph-H), 7.77 - 7.64 (m, 3H, d-H, 2 x Ph-H), 7.00 (d, J = 15.3 Hz, 1H, c-H), 5.76 (s, 1H, 14-H), 4.87 (s, 1H, 17-H), 4.52 (s, 1H, 17-H), 3.80 - 3.53 (m, 8H, a-H), 3.25 (dd, J = 11.7, 4.4 Hz, 1H, 3-H), 2.42 (ddd, J = 12.7, 4.3, 2.4 Hz, 1H, 7-H), 2.33 - 2.22 (m, 1H, 12-H), 2.01 - 1.92 (m, 2H, 7-H, 12-H), 1.91 (s, 3H, 16-CH3), 1.82 - 1.54 (m, 7H, 1-H, 2-CH2, 6-H, 9-H, 11-CH2), 1.39 (qd, J = 12.5, 2.7 Hz, 1H, 6-H), 1.16 (td, J = 13.2, 3.8 Hz, 1H, 1-H), 1.07 (dd, J = 12.5, 2.7 Hz, 1H, 5-H), 0.99 (s, 3H, 18-CH3), 0.77 (s, 3H, 19-CH3), 0.69 (s, 3H, 20-CH3).

[0106] Example 17

[0107]

[0108] Preparation of (E)-5-((lR,4aS,6R,8aS)-6-hydroxy-5,5,8a-trimethyl-2- methylenedecalin-l-yl)-3-methyl-l-(4-((E)-2-methyl-3-phenylacryloyl)piperazin-l- yl)pent-2-en-l-one (Compound 3p):

[0109] To the intermediate powder pumiloside piperazine amide (2) 19 mg was added 2 mL of dichloromethane to dissolve, then diisopropylethylamine 9.7 mg, N,N,N',N'-tetramethyluronium hexafluorophosphate 22.8 mg and (E)-2-methyl-3-phenylacrylic acid 9.7 mg were added in turn, stirred at room temperature, after the reaction was completed, the reaction was quenched with water, extracted with ethyl acetate, washed with saturated brine, dried over anhydrous sodium sulfate, and finally the ethyl acetate was rotary evaporated. After evaporation, the crude product was separated and purified by silica gel column chromatography to obtain the target compound 3p. Yield: 88%, white solid, melting point: 191-192°C; mass spectral data: C 34 H 48 N2O3[M+H] + , calculated value: 533.3738, measured value: 533.3718.

[0110] 1 H NMR (400 MHz, CDC13) δ 7.44 - 7.29 (m, 5H, 5 x Ph-H), 6.55 (s, 1H, d-H), 5.74 (s, 1H, 14-H), 4.86 (s, 1H, 17-H), 4.52 (s, 1H, 17-H), 3.73 - 3.48 (m, 8H, a-H), 3.23 (dd, J = 11.7, 4.4 Hz, 1H, 3-H), 2.41 (ddd, J = 12.8, 4.3, 2.4 Hz, 1H, 7-H), 2.30 - 2.22 (m, 1H, 12-H), 2.11 (d, J = 1.5 Hz, 3H, e-CH3), 1.95 (m, J = 13.7, 5.8 Hz, 2H, 7-H, 12-H), 1.90 (s, J = 1.2 Hz, 3H, 16-CH3), 1.82 - 1.54 (m, 7H, 1-H, 2-CH2, 6-H, 9-H, 11-CH2), 1.39 (qd, J = 12.9, 4.2 Hz, 1H, 6-H), 1.15 (td, J = 13.2, 3.9 Hz, 1H, 1-H), 1.06 (dd, J = 12.5, 2.7 Hz, 1H, 5-H), 0.98 (s, 3H, 18-CH3), 0.77 (s, 3H, 19-CH3), 0.69 (s, 3H, 20-CH3).

[0111] Example 18

[0112]

[0113] Preparation of (E)-5-((1R,4aS,6R,8aS)-6-hydroxy-5,5,8a-trimethyl-2- methylenedecalin-1-yl)-1-(4-isobutenoylpiperazin-1-yl)-3-methylpent-2-en-1-one (Compound 3q):

[0114] To the intermediate powder pumiloside piperazine amide (2) 19 mg was added 2 mL of dichloromethane to dissolve, then diisopropylethylamine 9.7 mg, N,N,N',N'-tetramethyluronium hexafluorophosphate 22.8 mg and isobutyryl acid 5.2 mg were added in turn, stirred at room temperature, after the reaction was completed, the reaction was quenched with water, extracted with ethyl acetate, washed with saturated brine, dried over anhydrous sodium sulfate, and finally the ethyl acetate was rotary evaporated. After evaporation, the crude product was separated and purified by silica gel column chromatography to obtain the target compound 3q. Yield: 97%, white solid, melting point: 184-186°C; mass spectral data: C 28 H 44 N2O3[M+H] + Calcd: 457.3425, Found: 457.3415.

[0115] 1 H NMR (400 MHz, CDC13) δ 5.73 (s, 1H, d-H, 14-H), 5.23 (s, 1H, d-H), 5.05 (s, 1H, d-H), 4.86 (s, 1H, 17-H), 4.51 (s, 1H, 17-H), 3.57 (s, 8H, a-H), 3.23 (dd, J = 11.7, 4.4 Hz, 1H, 3-H), 2.40 (ddd, J = 12.7, 4.3, 2.4 Hz, 1H, 7-H), 2.30 - 2.16 (m, 1H, 12-H), 1.95 (s, 3H, e-CH3), 1.95 - 1.89 (m, 2H, 7-H, 12-H), 1.88 (s, 3H, 16-CH3), 1.82 - 1.45 (m, 7H, 1-H, 2-CH2, 6-H, 9-H, 11-CH2), 1.38 (qd, J = 12.9, 4.3 Hz, 1H, 6-H), 1.15 (td, J = 13.2, 3.9 Hz, 1H, 1-H), 1.06 (dd, J = 12.5, 2.7 Hz, 1H, 5-H), 0.98 (s, 3H, 18-CH3), 0.76 (s, 3H, 19-CH3), 0.68 (s, 3H, 20-CH3).

[0116] Example 19

[0117]

[0118] Preparation of (2E,4E)-1-(4-((E)-5-((1R,4aS,6R,8aS)-6-hydroxy-5,5,8a- trimethyl-2-methylenodecyl-1-yl)-3-methylpent-2-enoyl)piperazin-1-yl)hexa-2,4- dien-1-one (Compound 3r):

[0119] To the intermediate powder pellitorinamide (2) 19 mg was added 2 mL of dichloromethane to dissolve, then diisopropylethylamine 9.7 mg, N,N,N',N'-tetramethyluronium hexafluorophosphate 22.8 mg and sorbic acid 6.1 mg were added in turn, stirred at room temperature, after the reaction was completed, the reaction was quenched with water, extracted with ethyl acetate, washed with saturated brine, dried over anhydrous sodium sulfate, and finally the ethyl acetate was rotary evaporated. After evaporation, the crude product was separated and purified by silica gel column chromatography to obtain the target compound 3r. Yield: 93%, white solid, melting point: 100-102°C; mass spectral data: C 30 H 46 N2O3[M+H] + Calcd: 483.3581, Found: 483.3557.

[0120] 1 H NMR (400 MHz, CDC13) δ 7.31 - 7.24 (m, 1H, d-H), 6.24 - 6.06 (m, 3H, c-H, e-H, f-H), 5.73 (s, 1H, 14-H), 4.86 (s, 1H, 17-H), 4.51 (s, 1H, 17-H), 3.74 - 3.45 (m, 8H, a-H), 3.24 (dd, J = 11.7, 4.4 Hz, 1H, 3-H), 2.40 (ddd, J = 12.7, 4.3, 2.4 Hz, 1H, 7-H), 2.25 (m, J = 14.1, 10.0, 3.6 Hz, 1H, 12-H), 1.99 - 1.90 (m, 2H, 7-H, 12-H), 1.88 (s, 3H, 16-CH3), 1.84 (d, J = 6.2 Hz, 3H, g-CH3), 1.80 - 1.53 (m, 7H, 1-H, 2-CH2, 6-H, 9-H, 11-CH2), 1.38 (qd, J = 12.9, 4.3 Hz, 1H, 6-H), 1.15 (td, J = 13.1, 3.9 Hz, 1H, 1-H), 1.06 (dd, J = 12.5, 2.7 Hz, 1H, 5-H), 0.99 (s, 3H, 18-CH3), 0.76 (s, 3H, 19-CH3), 0.68 (s, 3H, 20-CH3).

[0121] The 18 synthesized derivatives 3a-3r were subjected to anti-tumor activity studies.

[0122] Anti-tumor activity experiment steps:

[0123] (1) Collect cells in logarithmic growth phase, adjust the concentration of cell suspension, add 100 μL per well, and plate to adjust the density of the cells to be tested to 3000 cells per well (the edge wells are filled with sterile PBS);

[0124] (2) Incubate at 37℃ under 5% CO2 for 24 hours until the cells are completely adherent, then add different concentration gradients of drugs, generally 5 gradients, 100 μL per well, and set 3 replicate wells;

[0125] (3) Incubate at 37℃ under 5% CO2 for 72 hours, and observe under an inverted microscope;

[0126] (4) Add 10 μL of MTT solution (5 mg / mL, i.e. 0.5% MTT) per well, and incubate for 2 hours. After two hours, discard the medium in each well, and add 150 μL of DMSO. Use an enzyme-linked immunoassay instrument to measure the absorbance value of each well at 490 nm;

[0127] (5) Set zero adjustment wells (culture medium, MTT, dimethyl sulfoxide) and control wells (cells, same concentration of drug dissolution medium, culture solution, MTT, dimethyl sulfoxide) at the same time.

[0128] The IC50 of each compound on human non-small cell lung cancer cells A549, human hepatoma cells HepG2, human breast cancer cells MCF-7, and human triple-negative breast cancer cells MDA-MB-231 50 As shown in Table 1.

[0129] Table 1 Anti-tumor activity results of compounds (3a-3r)

[0130]

[0131] The present application takes pellucin as a substrate, first pellucin reacts with piperazine under the action of a condensing agent to generate an intermediate product pellucin piperazine amide, then the intermediate product pellucin piperazine amide reacts with a cinnamic acid compound or an unsaturated carboxylic acid compound under the action of a condensing agent to generate a series of 18 pellucin derivatives. The structures of these compounds are derived through physical and chemical properties and various spectral methods. Finally, using cisplatin as a positive control, the cytotoxicity of the compounds on A549, HepG2, MCF-7, and MDA-MB-231 cell lines is determined by the MTT method.

[0132] In the anti-tumor activity screening, most of the compounds showed obvious anti-tumor activity, 16 of the 18 derivatives generally showed strong inhibitory activity on the four cell lines A549, HepG2, MCF-7 and MDA-MB-231. Among them, compounds 3b, 3e, 3f, 3g, 3h, 3j, 3k, 3m and 3n showed better inhibitory activity on A549 cell line, IC 50 values were all less than 10 μM; compounds 3a, 3b, 3c, 3d, 3e, 3f, 3g, 3h, 3j, 3k, 3l, 3m and 3n showed better inhibitory activity on HepG2 cell line, IC 50 values were all less than 10 μM; compounds 3a, 3b, 3c, 3e, 3f, 3g, 3h, 3j, 3k, 3l, 3m, 3n, 3o, 3p and 3r showed better inhibitory activity on MCF-7 cell line, IC 50 values were all less than 10 μM; compounds 3a, 3b, 3c, 3d, 3e, 3f, 3g, 3h, 3j, 3k, 3l, 3m, 3n, 3o and 3r showed better inhibitory activity on MDA-MB-231 cell line, IC 50 values were all less than 10 μM. Among them, compound 3m showed IC 50 values of 2.48±0.49 μM, 1.04±0.01 μM, 0.73±0.28 μM and 1.55±0.12 μM, respectively. Among the 18 synthesized derivatives, compound 3m had the strongest cytotoxicity and was the best anti-tumor compound, which can be used as an anti-tumor lead compound for further research.

[0133] The above description is merely preferred embodiments of the present application, only to illustrate but not limit the present application. Those skilled in the art understand that many changes, modifications, and even equivalents can be made to the present application within the spirit and scope defined by the claims of the present application.

Claims

1. A pinnatin acid derivative, characterized by, The derivative includes the following specific structure:

2. A process for the preparation of a pinnatin derivative according to claim 1, characterized in that, The method comprises the following steps: S1, the powder of the powder back fern acid is dissolved in dichloromethane, then diisopropyl ethylamine, N, N, N', N'-tetramethyl urea hexafluorophosphate and piperazine are added in sequence, the reaction is quenched with water after the reaction is completed, extraction, washing, drying of the organic phase, spin-drying, evaporation to dryness, then the obtained crude product is separated and purified by silica gel column chromatography to obtain the intermediate product of powder back fern piperazine amide; S2, the powder back fern piperazine amide obtained in step S1 and the organic carboxylic acid compound are dissolved in dichloromethane, then diisopropyl ethylamine, N, N, N', N'-tetramethyl urea hexafluorophosphate and piperazine are added in sequence, stirring at room temperature, the reaction is quenched with water after the reaction is completed, extraction, washing, drying of the organic phase, spin-drying, evaporation to dryness, then the obtained crude product is separated and purified by silica gel column chromatography to obtain the target compound.

3. The method for preparing a fern acid derivative as described in claim 2, characterized in that, The organic carboxylic acid compound in the step S2 is any one of cinnamic acid, substituted cinnamic acid, 2-methyl-1-propenoic acid and sorbic acid.

4. Use of the powder back fern acid derivative of claim 1 in the preparation of an antitumor drug.

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