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CN-116947936-B - Mitochondrion-targeted rotenone-dequalinium chloride-hyaluronic acid compound and preparation method and application thereof

CN116947936BCN 116947936 BCN116947936 BCN 116947936BCN-116947936-B

Abstract

The invention belongs to the technical field of pharmaceutical chemistry, and particularly relates to a mitochondrion-targeted rotenone-dequalinium chloride-hyaluronic acid compound and a preparation method and application thereof. The rotenone-dequalinium chloride-hyaluronic acid compound has the following structural formula, wherein n is 2. The invention is based on anticancer activity of rotenone, and adopts reduction reaction, esterification reaction and twice amide reaction to successfully connect rotenone, dequalinium chloride and hyaluronic acid through amide bond, then the rotenone, dequalinium chloride and hyaluronic acid are successfully self-assembled into spherical nano particles through ultrasound, thus constructing the rotenone-dequalinium chloride-hyaluronic acid nano system for tumor treatment, and after the nano particles are targeted and transported to tumor cells, the rotenone and dequalinium chloride can both act on mitochondria in a synergistic way to form a double-drug delivery system, so that a large amount of ROS can be generated, thereby promoting apoptosis of cells and effectively inhibiting growth and migration of tumors.

Inventors

  • HONG MIN
  • WANG JUAN
  • CHENG SHUANG

Assignees

  • 聊城大学

Dates

Publication Date
20260505
Application Date
20230725

Claims (10)

  1. 1. A mitochondrial-targeted roteol-dequalinium chloride-hyaluronic acid conjugate, characterized in that the roteol-dequalinium chloride-hyaluronic acid conjugate has the following structural formula: wherein n is 2; The rotenone and the dequalinium chloride are connected with the hyaluronic acid through an amide bond in the rotenone-dequalinium chloride-hyaluronic acid conjugate, and then are self-assembled into spherical nano particles through ultrasound.
  2. 2. A method of preparing a mitochondrial-targeted rotenone-dequetiamine-hyaluronic acid conjugate according to claim 1, comprising the steps of: S1, dissolving rotenone in a first solvent, adding sodium borohydride for room temperature reaction, and adding water in a stirring state after the reaction is finished to obtain a compound 1, wherein the structure of the compound 1 is shown as follows: ; S2, placing malonic acid in a container, adding 1, 4-dioxane under a protective gas atmosphere, dropwise adding thionyl chloride, performing a first oil bath reaction, removing a solvent after the reaction is finished to obtain residues, adding a compound 1 solution into the residues, dropwise adding triethylamine, performing a second oil bath reaction, removing the solvent after the reaction is finished, washing, and drying to obtain a compound 2, wherein the structure of the compound 2 is as follows: ; s3, placing the compound 2, 1- (3-dimethylaminopropyl) -3-ethylcarbodiimide and 1-hydroxybenzotriazole in a container, adding a second solvent under the protection of gas atmosphere, performing a first activation reaction, adding a dequalinium chloride solution after the reaction is finished, adjusting the pH of the system to 9, performing a third oil bath reaction, filtering, concentrating, recrystallizing and drying to obtain a compound 3, wherein the structure of the compound 3 is shown as follows: ; S4, placing hyaluronic acid and 1- (3-dimethylaminopropyl) -3-ethylcarbodiimide in a container, adding a third solvent under a protective gas atmosphere, performing a second activation reaction, adding N-hydroxysulfosuccinimide and a compound 3 solution after the reaction is finished, performing a fourth oil bath reaction, filtering, dialyzing, centrifuging and drying to obtain the roteol-dequalinium chloride-hyaluronic acid conjugate.
  3. 3. The method for preparing the mitochondrially targeted rotenone-dequalinium chloride-hyaluronic acid conjugate according to claim 2, wherein in S1, the mass volume ratio of rotenone, a first solvent, sodium borohydride and water is 270-330 mg:8-12 ml:120-140 mg:8-12 mL, the reaction time at room temperature is 3-5 h, and the first solvent is methanol.
  4. 4. The method for preparing the mitochondria-targeted rotenone-dequalinium chloride-hyaluronic acid conjugate according to claim 2, wherein in S2, malonic acid, 1, 4-dioxane, thionyl chloride, compound 1 and triethylamine are mixed in a mass-volume ratio of 40-60mg:10-12 mL:100-110 μl:230-240 mg:60-70 μl, and the compound 1 solution is obtained by mixing the compound and N, N-dimethylformamide in a mass-volume ratio of 237 mg:10 mL.
  5. 5. The method for preparing the mitochondrially targeted rotenone-dequalinium chloride-hyaluronic acid conjugate according to claim 2, wherein in S2, the first oil bath reaction is carried out at a temperature of 100-120 ℃ for a time of 12-15 h, and the second oil bath reaction is carried out at a temperature of 60-80 ℃ for a time of 12-15 h.
  6. 6. The method for preparing the mitochondrially targeted rotenone-dequalinium chloride-hyaluronic acid conjugate according to claim 2, wherein in S3, the compound 2, 1- (3-dimethylaminopropyl) -3-ethylcarbodiimide, 1-hydroxybenzotriazole, the second solvent and dequalinium chloride are mixed in a mass-volume ratio of 70-80 mg:120-140 mg:120-130 mg:25-30 mL:65-80 mg, dequalinium chloride solution is obtained by mixing dequalinium chloride and methanol in a mass-volume ratio of 76 mg:7 mL, and the second solvent is methanol.
  7. 7. The method for preparing the mitochondria-targeted rotenone-dequinuclidine chloride-hyaluronic acid conjugate according to claim 2, wherein in S3, the temperature of the first activation reaction is 0-4 ℃ and the time is 4-6 h, the pH of the system is adjusted to 9 by adopting N, N-diisopropylethylamine, the temperature of the third oil bath reaction is 37 ℃ and the time is 20-24 h.
  8. 8. The method for preparing the mitochondria-targeted rotenone-dequalinium chloride-hyaluronic acid conjugate according to claim 2, wherein in S4, hyaluronic acid, 1- (3-dimethylaminopropyl) -3-ethylcarbodiimide, a third solvent, N-hydroxysulfosuccinimide and compound 3 are mixed in a mass volume ratio of 220-250 mg:210-230mg:25-30 mL:30-35 mg:230-240 mg, the third solvent is N, N-dimethylformamide and water in a volume ratio of 2:3, and the compound 3 solution is obtained by mixing compound 3 and N, N-dimethylformamide in a mass volume ratio of 237 mg:15 mL.
  9. 9. The method for preparing the mitochondrially targeted rotenone-dequalinium chloride-hyaluronic acid conjugate according to claim 2, wherein in S4, the second activation reaction is performed at 50-80 ℃ for 1-2 hours, and the fourth oil bath reaction is performed at 50-80 ℃ for 10-15 h hours.
  10. 10. Use of a mitochondrial-targeted rotenol-dequalinium chloride-hyaluronic acid conjugate according to claim 1 for the preparation of a medicament for the treatment and/or prophylaxis of cancer, wherein the cancer is selected from lung cancer, cervical cancer or liver cancer.

Description

Mitochondrion-targeted rotenone-dequalinium chloride-hyaluronic acid compound and preparation method and application thereof Technical Field The invention belongs to the technical field of pharmaceutical chemistry, and particularly relates to a mitochondrion-targeted rotenone-dequalinium chloride-hyaluronic acid compound and a preparation method and application thereof. Background Since 1932, 47 rotenone compounds have been extracted and isolated from plants such as rotenone. The compound has insecticidal activity because it can inhibit reduced Nicotinamide Adenine Dinucleotide (NADH) and block mitochondrial respiration. Research shows that rotenone can reduce environmental pollution, stimulate plant growth and delay pest's drug resistance effectively. However, rotenone is easy to decompose and lose efficacy under the conditions of sunlight, oxygen, high temperature and the like, so that the application of the rotenone is greatly limited. To reduce the limitations of rotenone, it may be structurally modified or chemically modified to construct a targeted prodrug. The design of the rotenone derivatives with high efficiency and low toxicity and the improvement of the bioactivity thereof are the problems to be solved urgently at present. Rotenone and its derivatives have been widely studied and used in practical applications as natural botanical insecticides and acaricides, but their studies on anticancer are very few, and there are studies showing that it can induce apoptosis by activating Reactive Oxygen Species (ROS), JNK/p38 and mitogen-activated protein kinase in human breast cancer MCF-7 cells. However, low doses of rotenone (< 10 nM) lead to oxidative damage and death of dopaminergic neurons, whereas long-term intake leads to parkinson's disease. In addition, there are limitations such as poor stability, multiple organ toxicity, and the like. Generally speaking, rotenone and its derivatives are still in an early stage in the application of targeted anticancer, and certain structural transformation and modification are necessary for better development of their application in anticancer field. In particular, anticancer studies of conjugates of targeting molecules with rotenone have not been reported. Therefore, the design and synthesis of the rotenone targeting molecule as the inhibitor of the mitochondrial respiratory chain have great application potential in the anticancer field. Disclosure of Invention In order to solve the problems, the invention provides a rotenone-dequalinium chloride-hyaluronic acid compound targeting mitochondria, a preparation method and application thereof, and a tumor targeting anticancer prodrug system based on anticancer activity of rotenone and combining a cancer cell targeting function of hyaluronic acid and a mitochondria targeting and cancer cell apoptosis promoting function of dequalinium chloride. Firstly starting from raw material rotenone, adopting reduction reaction, esterification reaction and two times of amide reaction to successfully connect rotenol, dequalinium chloride and hyaluronic acid through an amide bond, and then successfully self-assembling into spherical nano particles through ultrasound. A nanosystem (roteol-dequetiamide-hyaluronic acid) for tumor treatment was constructed. After the nano particles of the rotenone and the dequalinium chloride and the hyaluronic acid are targeted and transported to tumor cells, the rotenone and the dequalinium chloride can act on mitochondria in a synergistic way to form a double-drug delivery system, so that a large amount of ROS are generated, the apoptosis of the cells is promoted, and the growth and migration of the tumor are effectively inhibited. After that, by constructing a 4T1 tumor-bearing model mouse, the in vivo stability of the medicine is proved to be obviously superior to that of raw materials and other intermediate products, and the medicine has no obvious side effect on the mouse. The invention solves the technical problems through the following technical proposal. A first object of the present invention is to provide a mitochondrial-targeted roteol-dequalinium chloride-hyaluronic acid compound having the following structural formula: Wherein n is 2. A second object of the present invention is to provide a method for preparing the above-mentioned mitochondrial-targeted rotenone-dequetiazine-hyaluronic acid compound, comprising the steps of: S1, dissolving rotenone in a first solvent, adding sodium borohydride to react at room temperature, and adding water in a stirring state after the reaction is finished to obtain a compound 1; S2, placing malonic acid in a container, adding 1, 4-dioxane under a protective gas atmosphere, dropwise adding thionyl chloride, performing a first oil bath reaction, removing a solvent after the reaction is finished to obtain residues, adding a compound 1 solution into the residues, dropwise adding triethylamine, performing a second oil bath reaction, removing the solvent aft