A mitochondrion-targeted honokiol derivative, its preparation method and application

Abstract

The present invention belongs to the technical field of chemical medicine, and particularly relates to a mitochondrion-targeted honokiol derivative, a preparation method thereof and an application thereof. It is a compound represented by Formula I or Formula II, or a pharmaceutically acceptable salt of the compound represented by Formula I or Formula II, or a pharmaceutically acceptable ester of the compound represented by Formula I or Formula II, or a pharmaceutically acceptable solvate of the compound represented by Formula I or Formula II; #imgabs0# wherein, R1 is a straight-chain or branched-chain saturated alkyl group with 1 to 5 carbon atoms, R2 is a straight-chain or branched-chain saturated alkyl group with 1 to 5 carbon atoms, and n is an integer from 1 to 10. The mitochondrion-targeted honokiol derivative provided by the present invention not only has higher anti-tumor activity and cell selectivity, but also can improve the in vivo toxicity to the Caenorhabditis elegans model.

Description

Technical Field

[0001] The present invention belongs to the technical field of chemical medicine, and particularly relates to a mitochondrion-targeted honokiol derivative, a preparation method thereof, and an application thereof. Background Art

[0002] Disclosing the information of this background art section is only intended to increase the understanding of the overall background of the present invention, and is not necessarily regarded as an admission or an implication in any form that this information constitutes the prior art already known to those of ordinary skill in the art.

[0003] Chemotherapeutic drugs can inhibit the malignant proliferation of tumor cells, but are prone to cause toxic side effects and multidrug resistance. Mitochondria are the main organelles for energy supply and regulation of apoptosis in organisms. It has been found that there are abnormalities in the mitochondria of tumor cells. Designing drugs targeting mitochondria based on the differences between tumor cells and normal cells is one of the promising strategies to solve the problems of toxic side effects and drug resistance. The double-membrane structure and high negative charge of mitochondria make it difficult for therapeutic drugs to reach mitochondria. Lipophilic cations DLCs can utilize the abnormally increased mitochondrial membrane potential of tumor cells to achieve mitochondrial targeted aggregation. Conjugating natural products with anti-tumor activity to DLCs can improve the mitochondrial targeting effect of natural drugs, increase the bioavailability of natural drugs, and exert high anti-tumor activity.

[0004] Honokiol is a biphenyl diphenol compound extracted from the Chinese herbal medicine Magnolia officinalis. It can act on mitochondria and exert broad-spectrum anti-tumor activity by regulating mitochondrial energy metabolism and apoptosis. However, HN has no specific targeting to tumor cells. Structural modification is expected to obtain anti-tumor active derivatives that can enhance the efficacy and reduce toxicity compared with honokiol. Summary of the Invention

[0005] In order to solve the deficiencies of the prior art, the purpose of the present invention is to provide a mitochondrion-targeted honokiol derivative, a preparation method thereof, and an application thereof. Research shows that the mitochondrion-targeted honokiol derivative provided by the present invention not only has higher anti-tumor activity and cell selectivity, but also can improve the in vivo toxicity to the Caenorhabditis elegans model.

[0006] In order to achieve the above purpose, the technical solution of the present invention is as follows:

[0007] In the first aspect, a mitochondrion-targeted honokiol derivative is a compound represented by formula I or formula II, or a pharmaceutically acceptable salt of the compound represented by formula I or formula II, or a pharmaceutically acceptable ester of the compound represented by formula I or formula II, or a pharmaceutically acceptable solvate of the compound represented by formula I or formula II;

[0008]

[0009] Among them, R1 is a straight-chain or branched-chain saturated alkyl group with 1 to 5 carbon atoms, R2 is a straight-chain or branched-chain saturated alkyl group with 1 to 5 carbon atoms, and n is an integer from 1 to 10.

[0010] In the present invention, R1 and R2 can be the same or different.

[0011] In some embodiments of the present invention, R1 is a straight-chain saturated alkyl group with 1 to 5 carbon atoms, and R2 is a straight-chain saturated alkyl group with 1 to 5 carbon atoms. R1 is preferably methyl or propyl. R2 is preferably methyl or propyl.

[0012] In some embodiments of the present invention, n is an integer from 3 to 10. It is preferably 3, 6 or 10; particularly preferably 3.

[0013] In some embodiments of the present invention, it is selected from the following compounds:

[0014]

[0015] In the second aspect of the present invention, a method for preparing the mitochondrial-targeted honokiol derivative described in the first aspect includes the step of reacting honokiol as a starting material according to the following reaction route 1 or reaction route 2;

[0016] Reaction route 1:

[0017]

[0018] Reaction route 2:

[0019]

[0020] R1, R2, and n are as described above.

[0021] Specifically, the etherification reaction of the C-2-OH of honokiol with the iodoalkane R1-I obtains intermediate 2, and intermediate 2 reacts with the alkane brominated at both ends to carry out an etherification reaction to obtain intermediate 3, and then intermediate 3 reacts with F16 to obtain the compound of formula I.

[0022] More specifically, the conditions for the etherification reaction of honokiol with the iodoalkane R1-I are as follows: at room temperature, anhydrous potassium carbonate is added using DMF as a solvent. The molar ratio of honokiol to the iodoalkane R1-I is 1:1.5 to 2.5. The molar ratio of honokiol, the iodoalkane R1-I, and anhydrous potassium carbonate is 1:1.5 to 2.5:1.5 to 2.5. The conditional ratio of honokiol to DMF is 1:150 to 200, mol:mL.

[0023] More specifically, the conditions for the etherification reaction with the alkane brominated at both ends are as follows: at room temperature, anhydrous potassium carbonate is added using DMF as the solvent. The molar ratio of intermediate 2 to the alkane brominated at both ends is 1:1.5 - 2.5. The molar ratio of intermediate 2, the alkane brominated at both ends, and anhydrous potassium carbonate is 1:1.5 - 2.5:4.5 - 5.5. The conditional ratio of intermediate 2 to DMF is 1:150 - 200, mol:mL.

[0024] More specifically, the conditions for the etherification reaction with F16 are as follows: heating under reflux. The heating temperature is 80 - 90 °C. The solvent is one or more selected from N,N-dimethylformamide, n-butanol, and acetonitrile, preferably acetonitrile. The molar ratio of intermediate 3 to F16 is 1:2.0 - 3.0. The addition ratio of intermediate 3 to the solvent is 1:150 - 200, mol:mL.

[0025] Specifically, the C-4'-OH of honokiol undergoes an etherification reaction with the iodoalkane R2-I to obtain intermediate 2', and intermediate 2' undergoes an etherification reaction with the alkane brominated at both ends to obtain intermediate 3', and then intermediate 3' undergoes an etherification reaction with F16 to obtain the compound of formula II. The molar ratio of honokiol to the iodoalkane R2-I is 1:1.5 - 2.5. The molar ratio of honokiol, the iodoalkane R2-I, and anhydrous potassium carbonate is 1:1.5 - 2.5:1.5 - 2.5. The conditional ratio of honokiol to DMF is 1:150 - 200, mol:mL.

[0026] More specifically, the conditions for the etherification reaction of honokiol with the iodoalkane R2-I are as follows: at room temperature, anhydrous potassium carbonate is added using DMF as the solvent. The molar ratio of intermediate 2' to the alkane brominated at both ends is 1:1.5 - 2.5. The molar ratio of intermediate 2', the alkane brominated at both ends, and anhydrous potassium carbonate is 1:1.5 - 2.5:4.5 - 5.5. The conditional ratio of intermediate 2' to DMF is 1:150 - 200, mol:mL.

[0027] More specifically, the conditions for the etherification reaction with the alkane brominated at both ends are as follows: at room temperature, anhydrous potassium carbonate is added using DMF as the solvent. The molar ratio of intermediate '3 to F16 is 1:2.0 - 3.0. The addition ratio of intermediate 3' to the solvent is 1:150 - 200, mol:mL.

[0028] More specifically, the conditions for the etherification reaction with F16 are as follows: heating under reflux. The heating temperature is 80 - 90 °C. The solvent is one or more selected from N,N-dimethylformamide, n-butanol, and acetonitrile, preferably acetonitrile. The molar ratio of intermediate 3' to F16 is 1:2.0 - 3.0. The addition ratio of intermediate 3' to the solvent is 1:150 - 200, mol:mL.

[0029] In a third aspect, a pharmaceutical composition comprises an active ingredient and a pharmaceutically acceptable excipient, and the active ingredient is the mitochondrion-targeted honokiol derivative described in the first aspect.

[0030] The main purpose of administering the above pharmaceutical composition to a subject is to improve or treat symptoms of tumor-related diseases. In the pharmaceutical combination, the honokiol derivative, as the active ingredient, should be in an effective dose, and the "effective dose" means that it can effectively achieve the desired therapeutic or disease improvement effect in terms of dose and time period, and this dose can be routinely adjusted according to factors such as the subject and the administration method. Calculated based on the total mass of the pharmaceutical composition, the content of the derivative can be approximately 0.01 - 99%, 0.1 - 70%, 1 - 30%, 0.01 - 0.05%, 0.05 - 0.1%, 0.1 - 0.3%, 0.3 - 0.5%, 0.5 - 1%, 1 - 3%, 3 - 5%, 5 - 10%, 10 - 20%, 20 - 30%, 30 - 50%, 50 - 70%, or 70 - 99%. In a preferred embodiment, the subject is a mammal, including but not limited to rats, mice, rabbits, monkeys, dogs or humans, and most preferably humans.

[0031] Specifically, the pharmaceutically acceptable excipients include but are not limited to buffers, antioxidants, preservatives, bactericides, hydrophilic polymers, amino acids, monosaccharides, disaccharides and other carbohydrates, chelating agents, tonicity regulators, surfactants, salt-forming counterions, metal complexes and / or nonionic surfactants.

[0032] In a fourth aspect, there is provided the use of the mitochondrion-targeted honokiol derivative described in the first aspect or the pharmaceutical composition described in the third aspect in the preparation of a tumor-targeted therapeutic drug.

[0033] Specifically, the tumor-targeted therapeutic drug is for a subject in need of treatment to improve tumor-related symptoms. The administration methods include but are not limited to oral administration, nasal administration, inhalation administration, rectal administration, topical administration, injection administration or delivery to the lesion site by an interventional method.

[0034] Specifically, the tumor-targeted therapeutic drug is an anti-tumor drug, a model agent or a detection product.

[0035] The tumors described in the present invention include but are not limited to skin cancer, head and neck cancer, lung cancer, esophageal cancer, cervical cancer, uterine cancer, pancreatic cancer, breast cancer, kidney cancer, ureteral cancer, bladder cancer, pharyngeal squamous cell carcinoma, basal cell carcinoma or melanoma, tongue cancer, pharyngeal squamous cell carcinoma, malignant lymphoma, laryngeal squamous cell carcinoma, lung squamous cell carcinoma, small cell carcinoma, esophageal squamous cell carcinoma, cervical cancer, glioblastoma; preferably lung cancer.

[0036] Fifth aspect: An anti-tumor drug, which uses the mitochondrion-targeted honokiol derivative described in the first aspect as the active ingredient.

[0037] Specifically, the anti-tumor drug is used for treating lung cancer, and the dosage form is an oral preparation or an injection.

[0038] Compared with the prior art, the present invention has the following beneficial effects:

[0039] (1) High anti-tumor activity. The mitochondrion-targeted honokiol derivative provided by the present invention has strong in vitro anti-tumor activity, and its activity is significantly stronger than that of honokiol and F16; the cell selectivity is significantly improved. Some of the mitochondrion-targeted honokiol derivatives provided by the present invention show about 10-fold improved cell selectivity on lung cancer cells, and the differential effects on NCI-H446 cells and HBE cells have obvious time and concentration correlations.

[0040] (2) The mitochondrion-targeted honokiol derivative of the present invention can specifically target the mitochondria of tumor cells, disrupt the normal physiological functions of mitochondria such as free radical (such as reactive oxygen species ROS) overload and loss of membrane potential, resulting in abnormal metabolism of tumor cells.

[0041] (3) The mitochondrion-targeted honokiol derivative of the present invention can improve the in vivo toxicity to the Caenorhabditis elegans model. Description of the Drawings

[0042] The specification drawings forming a part of the present invention are used to provide a further understanding of the present invention. The schematic embodiments of the present invention and their descriptions are used to explain the present invention and do not constitute an improper limitation to the present invention.

[0043] Figure 1 It is the high-resolution mass spectrum of the derivative 2E-3-F16 of the present invention.

[0044] Figure 2 It is the nuclear magnetic resonance hydrogen spectrum of the derivative 2E-3-F16 of the present invention.

[0045] Figure 3 It is the nuclear magnetic resonance carbon spectrum of the derivative 2E-3-F16 of the present invention.

[0046] Figure 4 It is the cell growth curve data of the derivative 2E-3-F16 of the present invention on lung cancer cells NCI-H446 and human normal cells HBE;

[0047] Among them, Figure 4 (a) is 24 h; Figure 4 (b) is 48 h; Figure 4 (c) is 72 h.

[0048] Figure 5(a) is a graph of the treatment of NCI-H446 cells with 2E-3-F16 (0.25, 0.5, 1, 2 μM) for 24 h; (b) is a graph of the treatment of NCI-H446 cells for 24 h.

[0049] Figure 6 (a) is a flow cytometry detection chart of the mitochondrial membrane potential by 2E-3-F16 and HN; (b) is a quantitative analysis chart of the mitochondrial membrane potential by 2E-3-F16; (c) is a quantitative analysis chart of the mitochondrial membrane potential by HN.

[0050] Figure 7 This is the fluorescence image of the derivative 2E-3-F16 of the present invention in tumor cells.

[0051] Figure 8 (a) is a photographed image of the body length of the Caenorhabditis elegans model treated with 2E-3-F16 and HN (10, 20, 50, 100 μM) for 72 h; (b) is a data analysis chart of the body length of the Caenorhabditis elegans model by 2E-3-F16 and HN; (c) is a data analysis chart of the movement speed of the Caenorhabditis elegans model by 2E-3-F16 and HN. Detailed implementation manners

[0052] In order to enable those skilled in the art to more clearly understand the technical solution of the present invention, the technical solution of the present invention will be described in detail below in conjunction with specific embodiments.

[0053] Example 1 Preparation of compound 2B-3-F16

[0054] In this example, a preparation method of compound 2B-3-F16 is provided, and the synthetic route is as follows:

[0055]

[0056] Synthesis of 2B-3-F16: Under stirring, honokiol (2.12 g, 8 mmol) and anhydrous potassium carbonate (2.00 g, 16 mmol) were dissolved in 10 mL of DMF. 16 mmol of methyl iodide dissolved in 30 mL of DMF was slowly added dropwise to the reaction solution through a constant-temperature dropping funnel, and the reaction was stirred for 12 h in the dark. The reaction was monitored by TLC until completion. 700 mL of double-distilled water was added to the reaction solution, and the mixture was extracted with 150 mL of ethyl acetate three times. The organic phase was collected, back-extracted with saturated brine, dehydrated with anhydrous sodium sulfate, filtered under reduced pressure by vacuum, and concentrated by distillation to obtain a pale yellow viscous liquid. The liquid was purified by silica gel column chromatography (petroleum ether:ethyl acetate = 30:1) to obtain transparent viscous liquid compounds 2b and 2c with yields of 31.3% and 20.5% respectively. 2b (60.00 mg, 0.214 mmol) and anhydrous potassium carbonate (148.00 mg, 1.07 mmol) were dissolved in 3 mL of DMF solvent, and 0.428 mmol of 1,3-dibromopropane was slowly added. The reaction was stirred at room temperature for 12 h, and the reaction was monitored by TLC until completion. 60 mL of double-distilled water was added to the reaction solution, and the mixture was extracted with 20 mL of ethyl acetate three times. The organic phase was collected, back-extracted with saturated brine, dehydrated with anhydrous sodium sulfate, filtered by vacuum, and concentrated by distillation under reduced pressure to obtain a pale yellow viscous liquid. The liquid was purified by silica gel column chromatography (petroleum ether:ethyl acetate = 30:1) to obtain a transparent viscous compound 2b-3-Br with a yield of 82%. 2b-3-Br (78.10 mg, 0.195 mmol) and F16 (86.00 mg, 0.39 mmol) were dissolved in 5 mL of anhydrous acetonitrile and heated under reflux at 85 °C in an oil bath for 48 h. The reaction was monitored by TLC until completion, concentrated by distillation under reduced pressure, and the crude product was purified by silica gel column chromatography (dichloromethane:methanol = 30:1 - 10:1) to obtain a reddish-brown powdery compound with a yield of 36.5%. Spectral data:

[0057] 2B-3-F16: reddish-brown solidity. 11H NMR (600 MHz, CDCl3) δ 11.77 (s, 1H, N-H), 7.82 (d, J = 6.1 Hz, 1H, H-2”), 7.72 - 7.68 (m, 2H, H-6”, H-10”), 7.65 (m, 2H, H-16”, H-5”), 7.50 (d, J = 8.1 Hz, 1H, H-3”), 7.28 (dd, J = 8.4, 2.3 Hz, 1H, H-6'), 7.25 (d, J = 2.3 Hz, 1H, H-2'), 7.11 (d, J = 7.4 Hz, 1H, H-6), 7.07 (d, J = 2.2 Hz, 1H, H-4), 7.06 - 6.99 (m, 2H, H-15”, H-14”), 6.96 (m, 2H, H-7”, H-8”), 6.91 (d, J = 8.4 Hz, 1H, H-3), 6.76 (d, J = 8.4 Hz, 1H, H-5'), 6.44 (d, J = 15.9 Hz, 1H, H-13”), 6.05 - 5.88 (m, 2H, H-8, H-8'), 5.14 - 4.98 (m, 4H, H-9, H-9'), 4.40 (t, J = 6.6 Hz, 2H, H-3”'), 3.84 (s, 3H, H-1””), 3.79 (t, J = 5.6 Hz, 2H, H-1”'), 3.42 (d, J = 6.8 Hz, 2H, H-7), 3.32 (d, J = 6.7 Hz, 2H, H-7'), 2.29 (m, 2H, H-2”'). 13 13C NMR (151 MHz, CDCl3) δ 156.71, 154.24, 153.22, 141.87, 137.70, 137.61, 137.47, 137.09, 133.60, 133.18, 131.10, 130.81, 130.74, 128.51, 128.42, 128.36, 125.15, 123.15, 121.68, 120.88, 120.18, 116.13, 115.89, 115.84, 114.11, 113.23, 110.37, 64.46, 57.04, 55.82, 53.52, 39.50, 34.64, 30.21. HR-ESI-MS m / z: 541.2845 (calcd for C 37 H 37 N2O2 + [M - Br] + , 541.2850).

[0058] Preparation of Compound 2B - 6 - F16 in Example 2

[0059] In this example, a preparation method of compound 2B-6-F16 is provided. The preparation method of this compound is the same as that of 2B-3-F16 in Example 1, with the difference lying in the selection of the bromide chain length. The specific synthetic route is as follows:

[0060]

[0061] Spectral data:

[0062] 2B-6-F16: reddish-brown solidity. 1 H NMR(400MHz,CDCl3)δ11.75(s,1H,N-H),7.95(d,J=6.1Hz,2H,H-2”,H-6”),7.82-7.72(s,1H,H-10”),7.75-7.67(m,2H,H-16”,H-5”),7.64-7.55(d,J=8.1Hz,1H,H-3”),7.37-7.28(m,2H,H-6',H-2'),7.15(m,H-15”,H-14”),7.10(d,J=2.3Hz,1H,H-8”),7.04(dd,J=8.3,2.3Hz,1H,H-7”),6.85(dd,J=12.7,8.2Hz,2H,H-3,H-5'),6.52(d,J=15.8Hz,1H,H-13”),5.96(dtt,J=16.7,10.0,6.6Hz,2H,H-8,H-8'),5.03(m,4H,H-9,H-9'),4.26(t,J=7.5Hz,2H,H-6”'),3.85(t,J=6.3Hz,2H,H-1”'),3.81(s,3H,H-1””),3.35(dd,J=14.8,6.7Hz,4H,H-7,H-7'),1.72(m,2H,H-5”'),1.63(m,2H,H-2”'),1.34(m,2H,H-3”'),1.26-1.23(m,2H,H-4”'). 1313C NMR (101 MHz, CDCl3) δ 156.37, 154.29, 141.57, 137.90, 137.64, 137.42, 137.16, 133.08, 132.55, 131.30, 131.04, 130.95, 130.57, 128.45, 128.10, 127.81, 125.11, 123.26, 121.86, 121.24, 120.38, 116.18, 115.69, 115.55, 114.10, 113.20, 112.93, 110.09, 68.26, 60.12, 55.72, 50.96, 39.55, 34.51, 31.15, 28.99, 25.75, 25.58. HR-ESI-MS m / z: 583.3318 (calcd for C 40 H 43 N2O2 + [M - Br] + , 583.3319).

[0063] Preparation of Compound 2B-10-F16 in Example 3

[0064] In this example, a preparation method of Compound 2B-10-F16 is provided. The preparation method of this compound is the same as that of 2B-3-F16 in Example 1, with the difference being the selection of the bromide chain length. The specific synthetic route is as follows:

[0065]

[0066] Spectral data:

[0067] 2B-10-F16: reddish-brown solidity. 11H NMR (600 MHz, CDCl3) δ 11.88 (s, 1H, N-H), 7.98 (d, J = 5.8 Hz, 2H, H-2”, H-6”), 7.77 (s, 1H, H-10”), 7.71 (m, 2H, H-16”, H-5”), 7.62 - 7.58 (d, J = 8.1 Hz, 1H, H-3”), 7.37 - 7.32 (m, 2H, H-6', H-2'), 7.21 - 7.18 (m, 2H, H-6, H-4), 7.16 (dd, J = 6.5, 2.8 Hz, 2H, H-15”, H-14”), 7.11 (d, J = 2.4 Hz, 1H, H-8”), 7.04 (dd, J = 8.3, 2.4 Hz, 1H, H-7”), 6.86 (dd, J = 8.3, 4.3 Hz, 2H, H-3, H-5'), 6.53 (d, J = 15.8 Hz, 1H, H-13”), 6.05 - 5.91 (m, 2H, H-8, H-8'), 5.11 - 4.98 (m, 4H, H-9, H-9'), 4.32 (t, J = 7.6 Hz, 2H, H-10”'), 3.88 (t, J = 6.5 Hz, 2H, H-1”'), 3.83 (s, 3H, H-1””), 3.39 (dd, J = 6.6, 1.5 Hz, 2H, H-7), 3.34 (d, J = 6.9 Hz, 2H, H-7'), 1.75 (m, 2H, H-9”'), 1.69 - 1.63 (m, 2H, H-2”'), 1.36 - 1.31 (m, 2H, H-3”'), 1.28 - 1.21 (m, 10H, H-4”', H-5”', H-6”', H-7”', H-8”). 13 13C NMR (151 MHz, CDCl3) δ 156.40, 154.54, 154.37, 141.63, 137.97, 137.73, 137.44, 137.22, 133.14, 132.36, 131.36, 131.01, 130.67, 128.41, 127.99, 127.83, 125.18, 123.24, 121.85, 121.30, 120.38, 116.18, 115.62, 115.46, 114.12, 113.27, 112.88, 110.07, 68.74, 60.43, 55.66, 50.94, 39.57, 34.49, 31.36, 29.83, 29.49, 29.40, 29.37, 29.12, 26.27, 26.22. HR-ESI-MS m / z: 639.3946 (calcd for C 44 H 51 N2O2 +[M-Br] + ,639.3945).

[0068] Preparation of Compound 2C-3-F16 in Example 4

[0069] In this example, a preparation method of compound 2C-3-F16 is provided, and the specific synthetic route is as follows:

[0070]

[0071] Synthesis of 2C-3-F16: Dissolve 2c (60.00 mg, 0.214 mmol) and anhydrous potassium carbonate (148.00 mg, 1.07 mmol) in 3 mL of DMF solvent, and slowly add 0.428 mmol of 1,3-dibromopropane. The operation method is the same as that of 2b-3-Br to obtain a transparent viscous compound 2c-3-Br with a yield of 81.6%. Dissolve 2c-3-Br (70.02 mg, 0.175 mmol) and F16 (77.00 mg, 0.35 mmol) in 5 mL of anhydrous acetonitrile. The method is the same as that of 2B-3-F16. The crude product is purified by silica gel column chromatography (methylene chloride:methanol = 30:1 to 10:1) to obtain a reddish-brown powdery compound with a yield of 28.4%.

[0072] Spectral data:

[0073] 2C-3-F16: reddish-brown solidity. 11H NMR (600 MHz, CDCl3) δ 11.85 (s, 1H, N-H), 8.11 - 8.06 (m, 2H, H-2”, H-6”), 7.86 (s, 1H, H-10”), 7.82–7.65 (m, 2H, H-16”, H-5”), 7.60 (d, J = 6.1 Hz, 1H, H-3”), 7.30–7.27 (m, 2H, H-6', H-2'), 7.22 (m, 2H, H-6, H-4), 7.15 (d, J = 6.5 Hz, 2H, H-15”, H-14”), 7.09 - 7.06 (m, 2H, H-8”, H-7”), 6.87 (dd, J = 8.2, 1.7 Hz, 1H, H-3), 6.71 (d, J = 8.3 Hz, 1H, H-5'), 6.57 (d, J = 15.8 Hz, 1H, H-13”), 6.02–5.91 (m, 2H, H-8, H-8'), 5.10–5.00 (m, 4H, H-9, H-9'), 4.63–4.60 (m, 2H, H-3”'), 3.89 (d, J = 5.6 Hz, 2H, H-1”'), 3.74 (d, J = 1.7 Hz, 3H, H-1””), 3.34 (d, J = 6.2 Hz, 4H, H-7, H-7'), 2.36 (m, 2H, H-2”'). 13 13C NMR (151 MHz, CDCl3) δ 155.06, 142.06, 137.90, 137.41, 132.56, 131.66, 131.04, 128.81, 128.30, 127.74, 125.25, 123.36, 121.92, 121.31, 120.34, 116.20, 115.75, 115.69, 114.22, 113.36, 111.69, 111.03, 63.70, 57.34, 55.91, 53.52, 50.94, 39.52, 34.58, 30.78, 14.20. HR-ESI-MS m / z: 541.2848 (calcd for C 37 H 37 N2O2 + [M - Br] + , 541.2850).

[0074] Preparation of Compound 2C-6-F16 in Example 5

[0075] In this example, a preparation method of compound 2C-6-F16 is provided. The preparation method of this compound is the same as that of 2C-3-F16 in Example 4, and the difference lies in the selection of the bromide chain length. The specific synthetic route is as follows:

[0076]

[0077] Spectral data:

[0078] 2C-6-F16:reddish-brown solidity. 1 H NMR(600MHz,CDCl3)δ11.87(s,1H,N-H),8.02(d,J=6.0Hz,2H,H-2”,H-6”),7.78(s,1H,H-10”),7.72(d,J=7.6Hz,2H,H-16”,H-5”),7.61(d,J=7.5Hz,1H,H-3”),7.31(dd,J=8.3,2.3Hz,1H,H-6'),7.27(d,J=2.3Hz,1H,H-2'),7.24-7.20(m,2H,H-6,H-4),7.16(m,2H,H-15”,H-14”),7.08(d,J=7.9Hz,2H,H-8”,H-7”),6.87(d,J=8.0Hz,1H,H-3),6.81(d,J=8.4Hz,1H,H-5'),6.55(d,J=15.8Hz,1H,H-13”),5.97(dtt,J=17.1,10.4,6.7Hz,2H,H-8,H-8'),5.10-4.97(m,4H,H-9,H-9'),4.33(t,J=7.5Hz,2H,H-6”'),3.91(t,J=6.2Hz,2H,H-1”'),3.74(s,3H,H-1””),3.38(d,J=6.8Hz,2H,H-7),3.34(d,J=6.8Hz,2H,H-7'),1.80(m,2H,H-5”'),1.73(m,2H,H-2”'),1.48(m,2H,H-3”'),1.32(m,2H,H-4”'). 1313C NMR(151MHz,CDCl3)δ155.86,155.10,154.52,141.76,137.94,137.81,137.53,137.29,133.19,132.53,131.18,131.08,130.89,130.65,128.51,128.27,128.09,125.24,123.34,121.91,121.46,120.43,116.29,115.65,115.54,114.20,113.32,111.71,111.03,67.73,60.25,55.94,50.92,39.55,34.68,31.27,29.83,29.21,25.98,25.75.HR-ESI-MS m / z:583.3327(calcd for C 40 H 43 N2O2 + [M-Br] + ,583.3319).

[0079] Preparation of Compound 2C-10-F16 in Example 6

[0080] In this example, a preparation method of compound 2C-10-F16 is provided. The preparation method of this compound is the same as that of 2C-3-F16 in Example 4, and the difference lies in the selection of the bromide chain length. The specific synthetic route is as follows:

[0081]

[0082] Spectral data:

[0083] 2C-10-F16:reddish-brown solidity. 11H NMR (600 MHz, CDCl3) δ 11.90 (s, 1H, N-H), 7.99 (d, J = 6.0 Hz, 2H, H-2”, H-6”), 7.77 (s, 1H, H-10”), 7.73 - 7.67 (m, 2H, H-16”, H-5”), 7.59 (d, J = 4.8 Hz, 1H, H-3”), 7.32 (dd, J = 8.5, 2.3 Hz, 1H, H-6'), 7.28 (dd, J = 2.2 Hz, 1H, H-2'), 7.23 - 7.18 (m, 2H, H-6, H-4), 7.18 - 7.15 (m, 2H, H-15”, H-14”), 7.11 - 7.05 (m, 2H, H-8”, H-7”), 6.88 (d, J = 8.3 Hz, 1H, H-3), 6.84 (d, J = 8.4 Hz, 1H, H-5'), 6.53 (d, J = 15.8 Hz, 1H, H-13”), 6.05 - 5.92 (m, 2H, H-8, H-8'), 5.11 - 4.98 (m, 4H, H-9, H-9'), 4.31 (t, J = 7.6 Hz, 2H, H-10”'), 3.96 (t, J = 6.3 Hz, 2H, H-1”'), 3.76 (s, 3H, H-1””), 3.41 (d, J = 6.8 Hz, 2H, H-7), 3.35 (d, J = 6.7 Hz, 2H, H-7'), 1.77 (m, 4H, H-9”', H-2”'), 1.44 (m, 2H, H-3”'), 1.34 - 1.29 (m, 10H, H-4”', H-5”', H-6”', H-7”', H-8”'). 13 13C NMR (151 MHz, CDCl3) δ 156.03, 155.04, 154.37, 141.65, 137.95, 137.79, 137.34, 133.22, 132.42, 131.10, 131.06, 130.67, 130.62, 128.42, 128.35, 128.00, 125.20, 123.23, 121.84, 121.30, 120.37, 116.17, 115.64, 115.43, 114.12, 113.31, 111.54, 110.90, 68.10, 60.43, 55.88, 50.95, 39.55, 34.72, 31.38, 29.84, 29.54, 29.51, 29.43, 29.40, 29.14, 26.30, 26.27, 14.24. HR-ESI-MS m / z: 639.3946 (calcd for C 44 H 51 N2O2 + [M - Br]+ ,639.3945).

[0084] Preparation of Compound 2D-3-F16 in Example 7

[0085] In this example, a preparation method of compound 2D-3-F16 is provided, and the specific synthetic route is as follows:

[0086]

[0087] Synthesis of 2D-3-F16: Under stirring, honokiol (2.12 g, 8 mmol) and anhydrous potassium carbonate (2.00 g, 16 mmol) were dissolved in 10 mL of DMF. 16 mmol of iodopropane dissolved in 30 mL of DMF was slowly added dropwise through a constant-temperature dropping funnel, and the reaction was stirred for 12 h in the dark. The reaction was monitored by TLC until completion. 700 mL of double-distilled water was added to the reaction solution, and the mixture was extracted with 150 mL × 3 of ethyl acetate. The organic phase was collected, back-extracted with saturated brine, dehydrated with anhydrous sodium sulfate in the organic phase, filtered by vacuum suction, and concentrated by distillation to obtain a pale yellow viscous liquid. It was purified by silica gel column chromatography (petroleum ether:ethyl acetate = 40:1) to obtain compounds 2d and 2e with yields of 20.7% and 15.7% respectively. 2d (60.00 mg, 0.195 mmol) and anhydrous potassium carbonate (135.00 mg, 0.975 mmol) were dissolved in 3 mL of DMF solvent, and 0.585 mmol of 1,3-dibromopropane was slowly added in the same method as 2b-3-Br to obtain a transparent viscous compound 2d-3-Br with a yield of 81.9%. 2d-6-Br (52.00 mg, 0.11 mmol) and F16 (60.00 mg, 0.27 mmol) were dissolved in 5 mL of anhydrous acetonitrile in the same method as 2B-3-F16. The crude product was purified by silica gel column chromatography (methylene chloride:methanol = 30:1 - 10:1) to obtain a reddish-brown powdery compound with a yield of 26.6%.

[0088] Spectral data:

[0089] 2D-3-F16:reddish-brown solidity. 11H NMR (600 MHz, CDCl3) δ 11.79 (s, 1H, N-H), 7.79 (d, J = 2.8 Hz, 1H, H-2”), 7.70 (d, J = 6.4 Hz, 2H, H-6”, H-10”), 7.69–7.63 (m, 2H, H-16”, H-5”), 7.51 (d, J = 8.0 Hz, 1H, H-3”), 7.27 (d, J = 2.3 Hz, 2H, H-6', H-2'), 7.13–7.09 (m, 1H, H-6), 7.08 (d, J = 2.3 Hz, 1H, H-4), 7.05 (d, J = 7.7 Hz, 1H, H-15”), 7.02 (dd, J = 8.4, 2.5 Hz, 1H, H-14”), 6.95 (d, J = 6.3 Hz, 2H, H-7”, H-8”), 6.90 (d, J = 8.1 Hz, 1H, H-3), 6.77 (d, J = 8.4 Hz, 1H, H-5'), 6.45 (d, J = 15.9 Hz, 1H, H-13”), 6.06–5.88 (m, 2H, H-8, H-8'), 5.13–5.02 (m, 4H, H-9, H-9'), 4.42 (t, J = 6.5 Hz, 2H, H-3”'), 3.95 (t, J = 6.4 Hz, 2H, H-1”'), 3.79 (t, J = 5.5 Hz, 2H, H-1””), 3.43 (d, J = 6.8 Hz, 2H, H-7), 3.33 (d, J = 6.7 Hz, 2H, H-7'), 2.31 (m, 2H, H-2”'), 1.81 (m, 2H, H-2””), 1.03 (t, J = 7.4 Hz, 3H, H-3””). 13 13C NMR (151 MHz, CDCl3) δ 156.23, 154.21, 153.20, 141.93, 137.74, 137.61, 137.44, 137.21, 133.62, 133.13, 131.10, 130.82, 130.59, 128.53, 128.43, 128.37, 125.15, 123.16, 121.66, 120.82, 120.12, 116.09, 115.87, 115.76, 114.07, 113.26, 111.18, 69.96, 64.38, 56.99, 53.52, 50.85, 39.51, 34.85, 30.18, 22.88, 10.84. HR-ESI-MS m / z: 569.3163 (calcd for C 39 H 41 N2O2 + [M - Br] + , 569.3163).

[0090] Preparation of Compound 2D-6-F16 in Example 8

[0091] In this example, a preparation method of compound 2D-6-F16 is provided. The preparation method of this compound is the same as that of 2D-3-F16 in Example 7, and the difference lies in the selection of the bromide chain length. The specific synthetic route is as follows:

[0092]

[0093] Spectral data:

[0094] 2D-6-F16:reddish-brown solidity. 1 H NMR(400MHz,CDCl3)δ11.83(s,1H,N-H),7.96(d,J=6.5Hz,2H,H-2”,H-6”),7.75(s,1H,H-10”),7.69(d,2H,H-16”,H-5”),7.61-7.57(d,J=8.0Hz,1H,H-3”),7.31(d,J=2.3Hz,1H,H-6'),7.30(d,J=2.3Hz,1H,H-2'),7.18-7.13(m,4H,H-6,H-4,H-15”,H-14”),7.10(d,J=2.3Hz,1H,H-8”),7.04(dd,J=8.3,2.3Hz,1H,H-7”),6.84(dd,J=8.3,3.0Hz,2H,H-3,H-5'),6.52(d,J=15.9Hz,1H,H-13”),5.97(dddt,J=16.8,12.0,10.0,6.7Hz,2H,H-8,H-8'),5.12-4.94(m,4H,H-9,H-9'),4.28(t,J=7.4Hz,2H,,H-6”'),3.92(t,J=6.4Hz,2H,H-1”'),3.86(t,J=6.2Hz,2H,H-1””),3.39(d,J=6.7Hz,2H,H-7),3.34(d,J=6.7Hz,2H,H-7'),1.78(m,6H,H-5”',H-2”',H-2””),1.61(m,2H,H-3”'),1.37(m,2H,H-4”'),11.03(t,J=7.4Hz,3H,H-3””). 1313C NMR (151 MHz, CDCl3) δ 155.97, 154.48, 154.39, 141.71, 137.92, 137.80, 137.55, 137.37, 133.08, 132.65, 131.29, 131.07, 130.86, 130.81, 128.40, 128.15, 128.06, 125.21, 123.31, 121.89, 121.32, 120.35, 116.16, 115.66, 115.42, 114.16, 113.32, 113.15, 111.08, 69.90, 68.39, 60.17, 53.52, 50.92, 39.57, 34.75, 31.11, 29.83, 29.01, 25.75, 25.58, 22.93, 14.33, 10.88. HR-ESI-MS m / z: 611.3624 (calcd for C 42 H 47 N2O2 + [M - Br] + , 611.3632).

[0095] Preparation of Compound 2D - 10 - F16 in Example 9

[0096] In this example, a preparation method of compound 2D - 10 - F16 is provided. The preparation method of this compound is the same as that of 2D - 3 - F16 in Example 7, with the difference being the choice of bromide chain length. The specific synthetic route is as follows:

[0097]

[0098] Spectral data:

[0099] 2D - 10 - F16: reddish - brown solidity. 11H NMR (600 MHz, CDCl3) δ 11.89 (s, 1H, N-H), 8.00 (d, J = 6.4 Hz, 2H, H-2”, H-6”), 7.75 (s, 1H, H-10”), 7.73 - 7.67 (m, 2H, H-16”, H-5”), 7.62 - 7.57 (d, J = 8.0 Hz, 1H, H-3”), 7.35 (d, J = 2.3 Hz, 1H, H-6'), 7.33 - 7.29 (d, J = 2.3 Hz, 1H, H-2'), 7.19 (d, J = 6.3 Hz, 2H, H-6, H-4), 7.18 - 7.15 (m, 2H, H-15”, H-14”), 7.11 (d, J = 2.3 Hz, 1H, H-8”), 7.03 (dd, J = 8.3, 2.4 Hz, 1H, H-7”), 6.84 (m, 2H, H-3, H-5'), 6.54 (d, J = 15.8 Hz, 1H, H-13”), 6.04 - 5.91 (m, 2H, H-8, H-8'), 5.10 - 4.97 (m, 4H, H-9, H-9'), 4.33 (t, J = 7.6 Hz, 2H, H-10”'), 3.96 - 3.92 (m, 2H, H-1”'), 3.88 (t, J = 6.5 Hz, 2H, H-1””), 3.40 (d, J = 6.7 Hz, 2H, H-7), 3.34 (d, J = 6.7 Hz, 2H, H-7'), 1.83 - 1.79 (m, 2H, H-9”'), 1.76 (m, 2H, H-2”'), 1.67 (t, J = 7.3 Hz, 2H, H-2””), 1.35 (m, 2H, H-3”'), 1.26 - 1.22 (m, 10H, H-4”', H-5”', H-6”', H-7”', H-8”'), 1.04 (t, J = 7.3, 1.1 Hz, 3H, H-3””). 1313C NMR (151 MHz, CDCl3) δ 155.88, 154.52, 154.36, 141.64, 137.97, 137.74, 137.36, 133.08, 132.33, 131.27, 130.98, 130.74, 130.70, 128.31, 128.03, 127.91, 125.14, 123.22, 121.83, 121.27, 120.34, 116.14, 115.56, 115.34, 114.08, 113.28, 112.84, 110.88, 69.71, 68.70, 60.43, 53.55, 50.88, 39.56, 34.70, 31.34, 29.81, 29.47, 29.39, 29.37, 29.35, 29.09, 26.24, 26.19, 22.89, 10.87. HR-ESI-MS m / z: 667.4259 (calcd for C 46 H 55 N2O2 + [M - Br] + , 667.4258).

[0100] Preparation of Compound 2E-3-F16 in Example 10

[0101] In this example, a preparation method of compound 2E-3-F16 is provided, and the specific synthetic route is as follows:

[0102]

[0103] Synthesis of 2E-3-F16: Dissolve 2e (68.26 mg, 0.222 mmol) and anhydrous potassium carbonate (153.00 mg, 1.11 mmol) in 3 mL of DMF solvent, and slowly add 0.443 mmol of 1,3-dibromopropane. The method is the same as that of 2b-3-Br to obtain a transparent viscous compound 2e-3-Br with a yield of 84.3%. Dissolve 2e-3-Br (82.13 mg, 0.191 mmol) and F16 (105.00 mg, 0.479 mmol) in 5 mL of anhydrous acetonitrile. The method is the same as that of 2B-3-F16, and the crude product is purified by silica gel column chromatography (methylene chloride:methanol = 30:1 - 10:1) to obtain a reddish-brown powdery compound with a yield of 25.4%.

[0104] Spectral data:

[0105] 2E-3-F16: reddish-brown solidity. 11H NMR (600 MHz, CDCl3) δ 11.81 (s, 1H, N-H), 8.08 (d, J = 6.1 Hz, 2H, H-2”, H-6”), 7.90 (s, 1H, H-10”), 7.78 (d, J = 16.1 Hz, 1H, H-16”), 7.73 - 7.69 (m, 1H, H-5”), 7.64 - 7.59 (d, J = 8.0 Hz, 1H, H-3”), 7.36 (d, J = 2.2 Hz, 1H, H-6'), 7.29 (dd, J = 8.4, 2.3 Hz, 1H, H-2'), 7.26 - 7.22 (m, 2H, H-6, H-4), 7.16 - 7.13 (m, 2H, H-15”, H-14”), 7.12 - 7.02 (m, 2H, H-8”, H-7”), 6.86 (d, J = 8.4 Hz, 1H, H-3), 6.71 (d, J = 8.4 Hz, 1H, H-5'), 6.57 (d, J = 15.8 Hz, 1H, H-13”), 5.97 (tdt, J = 16.7, 10.1, 6.5 Hz, 2H, H-8, H-8'), 5.09 - 5.00 (m, 4H, H-9, H-9'), 4.60 (t, J = 6.9 Hz, 2H, H-3”'), 3.91 - 3.83 (m, 4H, H-1”', H-1””), 3.33 (d, J = 6.6 Hz, 4H, H-7, H-7'), 2.35 (d, J = 9.2 Hz, 2H, H-2”'), 1.70 (t, J = 7.0 Hz, 2H, H-2””), 0.94 (t, J = 7.4 Hz, 3H, H-3””). 13 13C NMR (151 MHz, CDCl3) δ 154.85, 154.65, 154.55, 141.96, 137.95, 137.75, 137.39, 133.42, 132.41, 131.83, 130.96, 130.39, 128.68, 128.20, 127.43, 125.24, 123.31, 121.88, 121.46, 120.43, 116.31, 115.74, 115.63, 114.25, 113.29, 112.91, 110.96, 70.35, 63.75, 57.30, 39.55, 34.58, 30.82, 22.79, 10.91. HR-ESI-MS m / z: 569.3163 (calcd for C 44 H 51 N2O2 + [M - Br] + , 569.3163).

[0106] Preparation of Compound 2E-6-F16 in Example 11

[0107] In this example, a preparation method of compound 2E-6-F16 is provided. The preparation method of this compound is the same as that of 2E-3-F16 in Example 10, and the difference lies in the selection of the bromide chain length. The specific synthetic route is as follows:

[0108]

[0109] Spectral data:

[0110] 2E-6-F16: reddish-brown solidity. 1 H NMR(600MHz,CDCl3)δ11.94(s,1H,N-H),7.99(d,J=6.2Hz,2H,H-2”,H-6”),7.77(s,1H,H-10”),7.72-7.68(m,2H,H-16”,H-5”),7.63-7.58(d,J=8.0Hz,1H,H-3”),7.36(d,J=2.4Hz,1H,H-6'),7.32(dd,J=8.4,2.3Hz,1H,H-2'),7.20-7.14(m,4H,H-6,H-4,H-15”,H-14”),7.10(d,J=2.3Hz,1H,H-8”),7.05(dd,J=8.3,2.3Hz,1H,H-7”),6.86(d,J=8.3Hz,1H,H-3),6.80(d,J=8.5Hz,1H,H-5'),6.53(d,J=15.9Hz,1H,H-13”),5.97(dddt,J=16.8,13.2,10.0,6.6Hz,2H,H-8,H-8'),5.10-4.98(m,4H,H-9,H-9'),4.33(t,J=7.4Hz,2H,H-6”'),3.94-3.89(m,2H,H-1”'),3.86(t,J=6.4Hz,2H,H-1””),3.38(d,J=6.7Hz,2H,H-7),3.34(d,J=6.7Hz,2H,H-7'),1.79(m,2H,H-5”'),1.71(m,4H,H-2””,H-2”'),1.51-1.39(m,2H,H-3”'),1.32(m,2H,H-4”'),0.95(t,J=7.4Hz,3H,H-3””). 1313C NMR(151MHz,CDCl3)δ155.69,154.53,154.41,141.66,137.99,137.81,137.48,137.27,133.31,132.34,131.38,130.95,130.62,128.42,128.01,127.87,125.19,123.27,121.86,121.33,120.39,116.20,115.61,115.55,114.15,113.32,112.81,110.87,70.27,67.64,60.22,39.57,34.71,31.28,29.83,29.21,25.97,25.74,22.78,10.94.HR-ESI-MS m / z:611.3633(calcd for C 42 H 47 N2O2 + [M-Br] + ,611.3632).

[0111] Preparation of Compound 2E-10-F16 in Example 12

[0112] In this example, a preparation method of compound 2E-10-F16 is provided. The preparation method of this compound is the same as that of 2E-3-F16 in Example 10, and the difference lies in the selection of the bromide chain length. The specific synthetic route is as follows:

[0113]

[0114] Spectral data:

[0115] 2E-10-F16:reddish-brown solidity. 11H NMR (600 MHz, CDCl3) δ 11.88 (s, 1H, N-H), 7.98 (d, J = 6.3 Hz, 2H, H-2”, H-6”), 7.81 - 7.79 (s, 1H, H-10”), 7.75 - 7.69 (m, 2H, H-16”, H-5”), 7.61 - 7.59 (d, J = 8.0 Hz, 1H, H-3”), 7.37 (d, J = 2.3 Hz, 1H, H-6'), 7.33 (dd, J = 8.4, 2.3 Hz, 1H, H-2'), 7.21 (d, J = 6.3 Hz, 2H, H-6, H-4), 7.16 (dt, J = 5.7, 2.1 Hz, 2H, H-15”, H-14”), 7.11 (d, J = 2.3 Hz, 1H, H-8”), 7.05 (dd, J = 8.3, 2.3 Hz, 1H, H-7”), 6.88 - 6.82 (m, 2H, H-3, H-5'), 6.53 (d, J = 15.8 Hz, 1H, H-13”), 6.05 - 5.91 (m, 2H, H-8, H-8'), 5.09 - 5.00 (m, 4H, H-9, H-9'), 4.30 (t, J = 7.6 Hz, 2H, H-10”'), 3.96 (t, J = 6.4 Hz, 2H, H-1”'), 3.87 (t, J = 6.4 Hz, 2H, H-1””), 3.41 - 3.39 (d, J = 6.7 Hz, 2H, H-7), 3.35 (d, J = 6.7 Hz, 2H, H-7'), 1.78 - 1.70 (m, 6H, H-9”', H-2”', H-2””), 1.50 - 1.41 (m, 2H, H-3”'), 1.33 - 1.25 (m, 10H, H-4”', H-5”', H-6”', H-7”', H-8”'), 0.96 (t, J = 7.4 Hz, 3H, H-3””). 1313C NMR(151 MHz, CDCl3) δ 155.87, 154.54, 154.34, 141.61, 137.99, 137.70, 137.40, 137.33, 133.14, 132.28, 131.28, 130.99, 130.75, 130.72, 128.33, 127.99, 127.91, 125.17, 123.22, 121.83, 121.33, 120.41, 116.22, 115.57, 115.41, 114.13, 113.23, 112.78, 110.83, 70.26, 68.10, 60.39, 50.90, 39.57, 34.73, 31.36, 29.52, 29.43, 29.14, 26.27, 22.77, 10.93. HR-ESI-MS m / z: 667.4253 (calcd for C 46 H 55 N2O2 + [M - Br] + , 667.4258).

[0116] Example 13: Evaluation of the Antitumor Activity of the Derivative

[0117] To verify the antitumor activity of the honokiol mitochondrial targeting derivative, the MTT colorimetric method was used to determine its growth inhibitory effects on human non - small cell lung cancer cell line A549, human normal liver cell line HL - 7702, human small cell lung cancer cell line NCI - H446, human bronchial epithelial cell line HBE, and human liver cancer cell line HepG - 2. The F16 and honokiol (HN) raw material drug groups were set up for control experiments.

[0118] Cell culture: NCI - H446, HBE, and HL - 7702 cells were cultured in 1640 medium with 10% fetal bovine serum (FBS), A549 cells were cultured in F12K with 10% FBS, and HepG - 2 cells were cultured in DMEM with 10% FBS. The culture conditions were in an incubator at 37°C and 5% CO2.

[0119] MTT colorimetric method: The tumor cells and normal cells were seeded at a density of 5×10 4 ~6×10 4Inoculate at a density of cells / mL into a 96-well plate, add 100 μL to each well, incubate overnight in an incubator with 5% CO2 at 37 °C until the cells adhere and grow to cover the bottom of the well. Dilute the drug dissolved in dimethyl sulfoxide with 2% medium to the target concentration, add 10 μL to each well of the 96-well plate, gently shake evenly to mix the drug with the medium, set up a blank control and 3 replicate well controls for each concentration. After incubating in the incubator for 48 h, add 10 μL of 0.5 mg / mL MTT to each well, continue to incubate for 4 h, and then aspirate all the medium in the well plate. Add 100 μL of dimethyl sulfoxide to each well to dissolve the water-insoluble purple crystalline formazan, and detect it at a wavelength of 570 nm with an enzyme-linked immunosorbent assay reader. The experiment is carried out three times at different culture times, and the average value and SD value are obtained. The anti-tumor activity of the honokiol mitochondrial-targeted compound is evaluated by the half-lethal dose IC 50 as follows, and the specific data are shown in Table 1.

[0120] Table 1 Evaluation of the anti-tumor activity of mitochondrial-targeted derivatives

[0121]

[0122] Note: IC 50 refers to the drug concentration corresponding to when the growth inhibition rate of the drug on the cells reaches 50% after the drug acts on the cells. All experimental results are repeated more than three times, and these values are expressed as the mean ± standard deviation.

[0123] The results are shown in Table 1. The derivatives show different cell activities: compared with the parent compound honokiol, the cytotoxicity of all derivatives has increased; especially, the cell selectivity of 2E-3-F16 is better than that of other derivatives and the raw materials HN and F16. Calculated by the selectivity index SI (IC 50,HBE / IC 50,NCI-H446 ), 2E-3-F16 shows approximately 10-fold higher tumor cell selectivity (IC 50,NCI-H446 = 2.87 μM, IC 50,HBE = 23.09 μM), while there is no significant difference between the raw materials HN and F16 in the two cell lines.

[0124] Meanwhile, in this example, the growth curves of NCI-H446 lung cancer cells and HBE normal human cells were also measured using 2E-3-F16 as an example. The results are shown in Figure 4 as follows, according to Figure 4It can be seen that 2E-3-F16 inhibits the cell viability of NCI-H446 cells in a time- and concentration-dependent manner, showing different degrees of inhibition. At a low concentration (1 μM), it shows different inhibitions between NCI-H446 and the normal cell line HBE, has a stronger killing effect and cell selectivity on the NCI-H446 cell line, and this difference becomes more and more obvious with the increase of time and concentration, showing significance.

[0125] This example tested the effect of the derivative on reactive oxygen species (ROS).

[0126] Experimental method: After the cells were incubated with the drug, the fluorescence probe DCFH-DA was used to detect the level of reactive oxygen species (ROS). The specific operation was to add 500 μL of serum-free cell culture medium containing 10 μM DCFH-DA to each sample, and culture it in a 37 °C cell incubator for 30 min. After the probe loading was completed, the culture medium was aspirated and the cells were washed twice with PBS. The ROS positive control agent Rosup was added to the positive control wells 4 hours before loading the probe, 1.5 mL per well, with a concentration of 50 mg / mL, and no addition was required for other groups. The cells were digested, centrifuged and collected, resuspended in PBS, and detected by flow cytometry (FL1 channel).

[0127] The results are shown in Figure 5 According to Figure 5 It can be seen that 2E-3-F16 at a low concentration (0.25 μM) can significantly stimulate the increase of ROS in NCI-H446 lung cancer cells, and with the increase of the drug concentration, the expression of ROS also shows a concentration-dependent increase; while there is no change trend in the expression of ROS in the cells treated with 2 μM HN, and it is significantly weaker than the positive control group.

[0128] This example also tested the effect of the derivative on mitochondrial membrane potential.

[0129] Experimental method: After the cells were incubated with the drug, the fluorescence probe JC-1 was used to detect the change of mitochondrial membrane potential. The specific operation was to add 1.5 mL of cell culture medium containing 5 μg / mL JC-1 to each sample, incubate it in a 37 °C cell incubator for 30 minutes, wash the cells three times with PBS, digest, centrifuge and collect the stained cells, resuspend them in PBS, mix well and place them on ice, protected from light, and detect by flow cytometry (FL2, FL3 channels).

[0130] Among them, Figure 6 shows the disruption of mitochondrial membrane potential of NCI-H446 cells by 2E-3-F16. 2E-3-F16 at a low concentration (0.25 μM) can cause a significant decrease in MMP, and with the increase of the drug concentration, MMP shows a concentration-dependent decrease to a relatively low level; compared with 2E-3-F16, the cells treated with HN at the same concentration (0.25 μM) did not cause a significant decrease in MMP.

[0131] Example 14: Targeted Mitochondrial Localization of Derivatives

[0132] To verify the mitochondrial targeting of the derivatives, the cellular localization of the compounds was performed using a laser confocal microscope.

[0133] Experimental procedure: After the administration of the NCI-H446 cells, 1 mL of Mito-tracker red CMXRos (200 nM) was added for staining, and incubated in the dark for 30 min. After the staining was completed, the staining solution was aspirated, and the cells were washed with PBS for 5 min × 3 times. Then, 1 mL of PBS was added, and the cells were photographed using a laser confocal microscope at 63× Oil. The mitochondrial dye excited red fluorescence and 2E-3-F16 excited green fluorescence, and the color overlap was observed.

[0134] The results are shown in Figure 7 . The green fluorescence excited by 2E-3-F16, which can enter tumor cells, significantly overlapped with the red fluorescence of Mito-Tracker Red CMXRos, indicating that 2E-3-F16 can not only enter tumor cells but also target and accumulate in the mitochondria of tumor cells.

[0135] Example 15: Toxicity Detection of Derivatives

[0136] An in vivo animal model of Caenorhabditis elegans was used to detect the movement speed and body length of the nematodes after 72 h of drug treatment, and to evaluate the toxicity of the derivatives before and after modification.

[0137] General culture and synchronization of Caenorhabditis elegans: Wild-type Caenorhabditis elegans N2 was purchased from the Caenorhabditis Genetics Center (CGC) and cultured at 20 °C on NGM (Nematode growth media) coated with Escherichia coli OP50. OP50, a uracil-leaky mutant strain of Escherichia coli, was used as the main food source for culturing Caenorhabditis elegans and was purchased from CGC.

[0138] After the nematodes grew to the adult stage and began to lay eggs and a small number of larvae hatched, they were washed from the surface of the NGM medium into a 1 mL centrifuge tube with M9 buffer. After the adults settled naturally, the supernatant larvae were aspirated for subculture. Alkaline lysis buffer was added to the adults in the precipitate, and vortexed for 5 - 7 min until the adults were completely lysed into fragments. Then, centrifuged at 5000 rpm for 1 min, and the supernatant was discarded. The eggs were washed twice with M9 buffer, each time at 5000 rpm for 1 min. Subsequently, the eggs were placed at 20 °C for incubation for 24 - 48 h.

[0139] Measurement of movement speed and body length: Synchronized L1-stage larvae were inoculated on NGM medium with or without drugs and cultured at 20 °C for 72 h. Nematodes were collected with M9 buffer, and residual OP50 was washed away. Then they were inoculated on the surface of NGM without OP50 and left standing at 20 °C for 30 min. A 1-min movement video of nematodes was taken using a continuous zoom stereo microscope (SZ650, Chongqing Opto-Electronic Instrument Co., Ltd.). After the video recording was completed, nematodes were collected with M9, dropped onto a glass slide, paralyzed with 20 mM sodium azide, covered with a coverslip, and photographed under a continuous zoom stereo microscope. There were at least 50 nematodes in each group. The movement speed was measured using the wrMTrck plugin of ImageJ software, the body length of nematodes was measured using ImageView software, and statistical graphs were made using GraphPad Prism 9.0 software. Experimental data were expressed as mean ± standard deviation (Mean ± SD), and P < 0.05 was considered statistically significant.

[0140] The results are shown in Figure 8 , compared with the blank group, 20, 50, and 100 μM of HN significantly inhibited the growth of nematodes ( # P < 0.05). However, 2E-3-F16 only showed a weak inhibitory effect on nematode growth when the concentration reached 100 μM, and compared with the HN group, 50 and 100 μM of 2E-3-F16 showed less toxicity (*P < 0.05). Compared with the blank group, 100 μM of HN significantly reduced the movement speed of nematodes. Compared with the HN group, 20, 50, and 100 μM of 2E-3-F16 significantly improved the movement speed of nematodes. The modified derivative significantly improved the growth inhibition and movement function of HN on nematodes. The nematodes showed a better health status and movement vitality, indicating that the in vivo toxicity of the modified derivative was improved.

[0141] The above are only the preferred embodiments of the present invention and are not used to limit the present invention. For those skilled in the art, the present invention can have various changes and modifications. Any modification, equivalent replacement, improvement, etc. made within the spirit and principle of the present invention shall be included within the protection scope of the present invention.

Claims

1. A mitochondrial-targeted honokiol derivative, characterized in that it is A compound represented by Formula I or Formula II, or a pharmaceutically acceptable salt of the compound represented by Formula I or Formula II; wherein, R1 is a C1-C5 straight-chain or branched-chain saturated alkyl group, R2 is a C1-C5 straight-chain or branched-chain saturated alkyl group, and n is an integer from 1 to 10.

2. The mitochondrial-targeted honokiol derivative according to claim 1, wherein R1 is a C1-C5 straight-chain saturated alkyl group, and R2 is a C1-C5 straight-chain saturated alkyl group.

3. The mitochondrial-targeted honokiol derivative according to claim 2, characterized in that, R1 is methyl or propyl.

4. The mitochondrial-targeted honokiol derivative according to claim 2, wherein, R2 is methyl or propyl.

5. The mitochondrial-targeted honokiol derivative according to claim 1, characterized in that, n is an integer from 3 to 10.

6. The mitochondrial-targeted honokiol derivative according to claim 5, characterized in that, n is 3, 6 or 10.

7. The mitochondrial-targeted honokiol derivative according to claim 6, characterized in that, n is 3.

8. The mitochondrial-targeted honokiol derivative according to claim 1, wherein selected from the following compounds:

9. A method for preparing the mitochondrion-targeted honokiol derivative according to claim 1, characterized in that, including the step of reacting magnolol as a starting material according to the following Reaction Route 1 or Reaction Route 2; Reaction Route 1: Reaction Route 2: wherein, R1, R2, and n are as described in Claim 1.

10. The preparation method of the mitochondrion-targeted honokiol derivative according to claim 9, characterized in that, Performing an etherification reaction between the C-2-OH of magnolol and an iodoalkane R1-I to obtain Intermediate 2, performing an etherification reaction between Intermediate 2 and an alkane brominated at both ends to obtain Intermediate 3, and then performing an etherification reaction between Intermediate 3 and F16 to obtain a compound of Formula I; or, performing an etherification reaction between the C-4'-OH of magnolol and an iodoalkane R2-I to obtain Intermediate 2', performing an etherification reaction between Intermediate 2' and an alkane brominated at both ends to obtain Intermediate 3', and then performing an etherification reaction between Intermediate 3' and F16 to obtain a compound of Formula II.

11. A pharmaceutical composition comprising an active ingredient and a pharmaceutically acceptable excipient, characterized in that, The active ingredient is a mitochondrion-targeted magnolol derivative as described in any one of Claims 1 to 8.

12. The pharmaceutical composition according to claim 11, wherein The pharmaceutically acceptable excipients are buffers, antioxidants, preservatives, bactericides, hydrophilic polymers, amino acids, monosaccharides, disaccharides and other carbohydrates, chelating agents, tonicity regulators, surfactants, salt-forming counterions and / or metal complexes.

13. The pharmaceutical composition according to claim 12, characterized in that, The surfactant is a non-ionic surfactant.

14. Use of a mitochondrion-targeted magnolol derivative as described in any one of Claims 1 to 8 or a pharmaceutical composition as described in any one of Claims 11-13 in the preparation of a tumor-targeted therapeutic drug.

15. The application according to claim 14, characterized in that, The tumor-targeted therapeutic drug is used for a subject in need of treatment to improve tumor-related symptoms.

16. The application according to claim 14, characterized in that, The administration mode of the tumor-targeted therapeutic drug is oral administration, nasal administration, inhalation administration, rectal administration, topical administration, injection administration or delivery to the lesion site by an interventional method.

17. The application according to claim 14, characterized in that, The tumor-targeted therapeutic drug is an anti-tumor drug, a model agent or a detection product.

18. The application according to claim 14, characterized in that, The tumors are skin cancer, head and neck cancer, lung cancer, esophageal cancer, cervical cancer, uterine cancer, pancreatic cancer, breast cancer, kidney cancer, ureteral cancer, bladder cancer, basal cell carcinoma or melanoma, malignant lymphoma, small cell carcinoma or glioma.

19. The application according to claim 18, characterized in that, The head and neck cancer is pharyngeal squamous cell carcinoma, tongue cancer or laryngeal squamous cell carcinoma.

20. The application according to claim 18, characterized in that, The lung cancer is squamous cell carcinoma of the lung.

21. The application according to claim 18, characterized in that, The esophageal cancer is esophageal squamous cell carcinoma.

22. The application according to claim 18, wherein, The tumor is lung cancer.

23. An anti-tumor drug, characterized in that, Using a mitochondrion-targeted magnolol derivative as described in any one of Claims 1 to 8 as the active ingredient.

24. The drug according to claim 23, characterized in that, The anti-tumor drug is used for the treatment of lung cancer, and the dosage form is an oral preparation or an injection.

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