Nuclear cross-linked nano-drug based on dual-enhanced cascade reaction as well as preparation method and application of nuclear cross-linked nano-drug

Abstract

The invention discloses a core-crosslinked nano-drug based on a dual enhanced cascade reaction as well as a preparation method and application of the core-crosslinked nano-drug. The core-crosslinked nano-drug is prepared from polylysine-ferrocene, polyethylene glycol-poly (lysine-cyclodextrin)-polyglutamic acid, glucose oxidase and cis-platinum in a mass ratio of 1: 2: 1.2: 0.7. The core-crosslinked nano-drug based on the dual-enhanced cascade reaction can effectively avoid premature leakage of the drug and has good stability, and the preparation method is simple. Compared with the prior art, the nuclear cross-linked nano-drug based on the dual-enhanced cascade reaction combines chemotherapy, chemodynamic therapy and hunger therapy, and the anti-tumor treatment effect is greatly improved through a combined treatment system of a combined therapy of the chemotherapy, the chemodynamic therapy and the hunger therapy.

Description

Technical Field

[0001] The present invention belongs to the technical field of biomedicine, and in particular relates to a nuclear cross-linked nano drug based on a double enhanced cascade reaction, and a preparation method and application thereof. Background Art

[0002] Chemodynamic therapy can effectively convert endogenous hydrogen peroxide into highly toxic reactive oxygen species through the intracellular Fenton chemical reaction, ultimately causing cancer cell death. Given that the generation of reactive oxygen species is not affected by external excitation or local oxygen concentration, chemodynamic therapy is very suitable for inhibiting deep tumor growth, especially those hypoxic tumors that cannot be effectively treated by conventional photodynamic therapy. However, a significant obstacle to chemodynamic therapy is the lack of endogenous hydrogen peroxide in the tumor, which leads to inefficient Fenton reaction and affects the treatment effect of cancer. Therefore, other treatment methods can be integrated into traditional chemodynamic therapy strategies to solve the problem of insufficient endogenous hydrogen peroxide and enhance anti-cancer efficacy.

[0003] In addition, platinum drugs can significantly inhibit the growth and spread of cancer cells including liver cancer, ovarian cancer, lung cancer, bladder cancer, testicular cancer and colon cancer. Among them, cisplatin, as a platinum drug, can specifically promote the oxidation of nicotinamide adenine dinucleotide phosphate (NADPH) to NADP+ by activating nicotinamide adenine dinucleotide phosphate oxidase (NOXs), and then generate superoxide anions from one oxygen molecule. The generated superoxide anions are dismutated by superoxide dismutase (SOD), which can cause the level of hydrogen peroxide to rise. In addition, glucose oxidase has also been shown to consume glucose to cut off the energy supply of tumor cells for starvation therapy, and accelerate the generation of hydrogen peroxide through its catalytic activity. Therefore, platinum prodrug-mediated chemotherapy and glucose oxidase-mediated starvation therapy are expected to be combined with chemodynamic therapy to achieve the enhancement of Fenton-like reactions with self-supplied hydrogen peroxide, which is a promising tumor treatment strategy.

[0004] Literature search found that the research group of Lu Ligong of Zhuhai People's Hospital in my country disclosed a supramolecular spherical micelle constructed by cyclodextrin modified with photosensitizer dihydrochlorin e6 and ferrocene-modified polyethylene glycol in the paper (Self-Delivered Supramolecular Nanomedicine with Transformable Shape for Ferrocene-Amplified Photodynamic Therapy of Breast Cancer and Bone Metastases, Adv. Funct. Mater. 2021, 2104645), and studied its performance and application in tumor treatment. However, the ferrocene loaded in the above system can only use the limited endogenous hydrogen peroxide in tumor cells for Fenton reaction, which greatly reduces the efficacy of chemodynamic therapy and is difficult to transform and apply in clinical practice. Summary of the invention

[0005] In view of the deficiencies in the prior art, the present invention provides a core-crosslinked nanomedicine based on a double-enhanced cascade reaction and a preparation method and application thereof, so as to solve the current problems of instability of nanomedicines and low efficiency of chemodynamic therapy, as well as the lack of combined treatment methods for chemotherapy, chemodynamic therapy and starvation therapy.

[0006] The present invention is achieved through the following technical solutions:

[0007] A core-crosslinked nanomedicine based on a double-enhanced cascade reaction, wherein the core-crosslinked nanomedicine is prepared from polylysine-ferrocene, polyethylene glycol-poly(lysine-cyclodextrin)-polyglutamic acid, glucose oxidase and cisplatin in a mass ratio of 1:2:1.2:0.7; wherein the structural formula of the polyethylene glycol-poly(lysine-cyclodextrin)-polyglutamic acid is shown in the following formula III:

[0008]

[0009] In formula III: n is 20.

[0010] Preferably, the particle size of the core-crosslinked nanomedicine is less than 200 nm.

[0011] The method for preparing the core-crosslinked nanomedicine based on the double enhanced cascade reaction comprises the following steps:

[0012] Step 1) preparing poly-lysine-ferrocene;

[0013] Step 2) preparing polyethylene glycol-poly(lysine-cyclodextrin)-polyglutamic acid;

[0014] Step 3) polylysine-ferrocene, polyethylene glycol-poly(lysine-cyclodextrin)-polyglutamic acid, glucose oxidase and cisplatin are dissolved in N,N-dimethylformamide solvent in proportion, deionized water is added dropwise, stirred, and dialyzed to obtain the product.

[0015] Preferably, the process of preparing polylysine-ferrocene in step 1) is as shown in reaction formula II:

[0016]

[0017] The specific steps are as follows:

[0018] Step 1-1) 40.4 mg of polylysine, 405.4 mg of N-hydroxysuccinimide and 619.2 mg of 1-(3-dimethylaminopropyl)-3-ethylcarbodiimide hydrochloride were dissolved in 10 mL of anhydrous N,N-dimethylformamide and stirred for 4 h in a nitrogen atmosphere to obtain a reaction solution A;

[0019] Step 1-2) Add 107 mg of ferrocene carboxaldehyde to reaction solution A, and continue stirring the reaction for 24 hours to obtain reaction solution B;

[0020] Step 1-3) The reaction solution B was precipitated in 80 mL of anhydrous ether, and then centrifuged, repeated 3 times, and vacuum dried to obtain.

[0021] Preferably, the process of preparing polyethylene glycol-poly(lysine-cyclodextrin)-polyglutamic acid in step 2) is as shown in reaction formula IV:

[0022]

[0023] The specific steps are as follows:

[0024] Step 2-1) 200 mg of polyethylene glycol amino and 244.9 mg of ε-benzyloxycarbonyl-L-lysine anhydride were dissolved in 2 mL of anhydrous N,N-dimethylformamide, and stirred for reaction for 48 h in a nitrogen atmosphere to obtain a reaction solution a; 210.5 mg of γ-benzyl-glutamic acid anhydride was added to the reaction solution a, and the reaction was continued with stirring for 48 h to obtain a reaction solution b; the reaction solution b was precipitated in 16 mL of anhydrous ether, and then centrifuged, repeated 3 times, and vacuum dried to obtain polyethylene glycol-poly(ε-benzyloxycarbonyl-L-lysine)-poly(ε-benzyloxycarbonyl-L-glutamic acid);

[0025] Step 2-2) 100 mg of polyethylene glycol-poly(ε-benzyloxycarbonyl-L-lysine)-poly(ε-benzyloxycarbonyl-L-glutamic acid) was dissolved in 10 mL of a mixed solvent of glacial acetic acid / trifluoroacetic acid, and 1.1 mL of a mixed solution of hydrobromic acid / glacial acetic acid was added at 0° C. The reaction was continued for 1.5 h to obtain a reaction solution c; the reaction solution c was precipitated in 80 mL of anhydrous ether, and then centrifuged, repeated 3 times, and finally dried in a vacuum oven to obtain polyethylene glycol-polylysine-poly(ε-benzyloxycarbonyl-L-glutamic acid);

[0026] Step 2-3) 180 mg of cyclodextrin-carboxyl was dissolved in 10 mL of anhydrous N,N-dimethylformamide, and then 246 mg of N-hydroxysuccinimide and 408 mg of 1-(3-dimethylaminopropyl)-3-ethylcarbodiimide hydrochloride were added, and the mixture was stirred for 2 h in a nitrogen atmosphere to obtain a reaction solution d; then 57 mg of polyethylene glycol-polylysine-poly(ε-benzyloxycarbonyl-L-glutamic acid) was dissolved in 5 mL of anhydrous N,N-dimethylformamide and added dropwise to the reaction solution d, and the reaction was continued by stirring for 24 h to obtain a reaction solution e; the reaction solution e was precipitated in 80 mL of anhydrous ether, and then centrifuged, repeated 3 times, and vacuum dried to obtain polyethylene glycol-poly(lysine-cyclodextrin)-poly(ε-benzyloxycarbonyl-L-glutamic acid);

[0027] Step 2-4) 100 mg of polyethylene glycol-poly(lysine-cyclodextrin)-poly(ε-benzyloxycarbonyl-L-glutamic acid) was dissolved in 20 mL of 1 M NaOH solution and vigorously stirred at 100° C. under reflux overnight; the resulting reaction mixture was placed in a dialysis bag and dialyzed with deionized water for 48 h. After dialysis, the mixture was freeze-dried to obtain the product.

[0028] Preferably, in step 3), the concentration of the polylysine-ferrocene is 2.5 mg / mL; the concentration of the polyethylene glycol-poly(lysine-cyclodextrin)-polyglutamic acid is 5 mg / mL; the concentration of the glucose oxidase is 3.0 mg / mL; and the concentration of the cisplatin is 1.75 mg / mL.

[0029] Preferably, in step 3), the volume ratio of the N,N-dimethylformamide to the deionized water is 1:5.

[0030] Preferably, the dialysis conditions in step 3) are: using a dialysis bag with a molecular weight of 3500, 500 mL of deionized water × 1 time / 8 hours, and dialysis for 24 hours.

[0031] The use of the above-mentioned core-crosslinked nanomedicine based on the double-enhanced cascade reaction, or the core-crosslinked nanomedicine based on the double-enhanced cascade reaction prepared by the above-mentioned preparation method in the preparation of anti-tumor drugs.

[0032] An anti-tumor pharmaceutical composition comprises the above-mentioned core-crosslinked nanomedicine based on double-enhanced cascade reaction, or the core-crosslinked nanomedicine based on double-enhanced cascade reaction prepared by the above-mentioned preparation method, and pharmaceutically acceptable excipients.

[0033] The beneficial effects of the present invention are as follows:

[0034] (1) In the components of the core-crosslinked nanomedicine based on the double enhanced cascade reaction of the present invention, cisplatin is reduced and released in the high-concentration glutathione reducing environment in tumor cells to achieve chemotherapy, and at the same time, hydrogen peroxide is generated for subsequent chemodynamic therapy; glucose oxidase catalyzes glucose in liver cancer cells, blocks the energy supply in the cells, achieves starvation therapy, and at the same time, hydrogen peroxide is generated for subsequent chemodynamic therapy; ferrocene reacts with endogenous hydrogen peroxide in liver cancer cells and the generated hydrogen peroxide to produce Fenton reaction, kill tumor cells, and achieve chemodynamic therapy. The core-crosslinked nanomedicine is used in the integrated treatment system of chemotherapy-chemodynamic therapy-starvation therapy, which is simple in technical operation and only requires two intravenous injections to achieve effective treatment of tumors, and has important clinical application prospects.

[0035] (2) The core-crosslinked nanomedicine based on the double-enhanced cascade reaction of the present invention can effectively avoid premature leakage of drugs, has good stability, and the preparation method is simple and efficient, providing a good experimental platform for obtaining core-crosslinked nanomedicines for integrated chemotherapy-chemodynamic therapy-starvation therapy. Compared with the prior art, the core-crosslinked nanomedicine based on the double-enhanced cascade reaction of the present invention combines chemotherapy, chemodynamic therapy and starvation therapy, and greatly improves the effect of anti-tumor treatment through the treatment system of the three combined therapies. BRIEF DESCRIPTION OF THE DRAWINGS

[0036] Figure 1 is the NMR spectrum of polylysine-ferrocene in Example 1;

[0037] Figure 2 is the NMR spectrum of polyethylene glycol-poly(ε-benzyloxycarbonyl-L-lysine)-poly(ε-benzyloxycarbonyl-L-glutamic acid) in Example 1;

[0038] Figure 3 is the NMR spectrum of polyethylene glycol-poly(lysine-cyclodextrin)-polyglutamic acid in Example 1;

[0039] Figure 4 Schematic diagram of the preparation of core-crosslinked nanomedicine based on double enhanced cascade reaction in Example 1;

[0040] Figure 5 The dynamic light scattering spectrum of the core-crosslinked nanomedicine based on the double-enhanced cascade reaction in Example 1;

[0041] Figure 6 The effects of each group of drugs in Example 2 at different concentrations on liver cancer cells;

[0042] Figure 7 The effect of each group of drugs in Example 3 on the tumor size of nude mice bearing liver cancer tumors over time;

[0043] Figure 8 The figure shows the effects of each group of drugs in Example 3 on the body weight of nude mice bearing liver cancer tumors over time. DETAILED DESCRIPTION

[0044] The present invention is further described in detail below with reference to the accompanying drawings and specific embodiments.

[0045] Unless otherwise specified, the technical means used in the following examples are all conventional means well known to those skilled in the art, and the experimental methods without specifying the specific conditions are all conventional methods in the art.

[0046] Unless otherwise specified, the materials and reagents used in the following examples can be obtained from commercial sources.

[0047] The experimental animals in the following examples are nude mice, which were provided by the Experimental Animal Center of Nantong University.

[0048] Example 1 Preparation of core-crosslinked nanomedicine (NP-CDDP / GOD) based on dual enhanced cascade reaction

[0049] 1. Preparation of polylysine-ferrocene (PPL-Fc)

[0050] The structural formula of PPL-Fc is shown in Formula I below:

[0051]

[0052] Its preparation process is shown in reaction formula II:

[0053]

[0054] The specific steps are as follows:

[0055] Take 40.4 mg of polylysine, 405.4 mg of N-hydroxysuccinimide and 619.2 mg of 1-(3-dimethylaminopropyl)-3-ethylcarbodiimide hydrochloride and dissolve them in 10 mL of anhydrous N,N-dimethylformamide (DMF), and stir them for 4 hours in a nitrogen atmosphere to obtain reaction solution A. Then, 107 mg of ferrocene carboxaldehyde is added to the reaction solution A, and the reaction is continued with stirring for 24 hours to obtain reaction solution B. After the reaction is completed, the reaction solution B is precipitated in 80 mL of anhydrous ether, centrifuged again, and repeated 3 times, and vacuum dried to obtain 79.4 mg of PPL-Fc (solid), with a yield of 79.5%.

[0056] The PPL-Fc prepared in this example is as follows Figure 1 As shown, the detailed peak positions are attributed to: 1 H-NMR (400MHz, DMSO-d6), δ (ppm) = 4.35-4.2 (m, 9H, Fc), 2.86-2.71 (m, 22H, COCHNH, CH 3(CH2)4CH2),1.77-1.45(m,168H,CH3(CH2)4,CN(CH2)4),1.42-1.19(m,3H,CH3CH2).

[0057] 2. Preparation of polyethylene glycol-poly(lysine-cyclodextrin)-polyglutamic acid (PEG-P(LL-CD)-PGlu) The structural formula of PEG-P(LL-CD)-PGlu is shown in Formula III below:

[0058]

[0059] Its preparation process is shown in reaction formula IV:

[0060]

[0061] In the above formulas I, II, III and IV, the value of n is 20.

[0062] The specific steps are as follows:

[0063] (1) Preparation of polyethylene glycol-poly(ε-benzyloxycarbonyl-L-lysine)-poly(ε-benzyloxycarbonyl-L-glutamic acid)

[0064] Take 200 mg of polyethylene glycol amino and 244.9 mg of ε-benzyloxycarbonyl-L-lysine NCA and dissolve them in 2 mL of anhydrous DMF, stir and react for 48 hours in a nitrogen atmosphere to obtain a reaction solution a. Then, add 210.5 mg of γ-benzyl-glutamic acid NCA to the reaction solution a, continue to stir and react for 48 hours, and obtain a reaction solution b. After the reaction is completed, the reaction solution b is precipitated in 16 mL of anhydrous ether, centrifuged again, repeated 3 times, and vacuum dried to obtain 350.4 mg of polyethylene glycol-poly(ε-benzyloxycarbonyl-L-lysine)-poly(ε-benzyloxycarbonyl-L-glutamic acid) (solid), with a yield of 75.4%.

[0065] The polyethylene glycol-poly(ε-benzyloxycarbonyl-L-lysine)-poly(ε-benzyloxycarbonyl-L-glutamic acid) prepared in this example is as follows Figure 2 As shown, the detailed peak positions are attributed to: 1 H-NMR (400MHz, DMSO-d6), δ (ppm) = 8.12 (s, 40H, COCHNH), 7.4-7.11 (m, 200H, Ph), 5.11 -4.9(m,80H,COOCH2),4.33-4.13(m,42H,COCHNH,NHCH2),3.51(s,180H,CH3O(CH2CH2) 45 ),3.24(m,3H,CH3),2.94(m,40H,OOCNHCH2CH2),2.38(m,80H,NHCH2CH2,OOCCH2),2.06-1.75(m,40H,NHCH2CH2CH2),1.47-1.17(m,80H,CHCH2).

[0066] (2) Preparation of polyethylene glycol-polylysine-poly(ε-benzyloxycarbonyl-L-glutamic acid)

[0067] 100 mg of polyethylene glycol-poly(ε-benzyloxycarbonyl-L-lysine)-poly(ε-benzyloxycarbonyl-L-glutamic acid) was dissolved in 10 mL of a mixed solvent of glacial acetic acid / trifluoroacetic acid (volume ratio of 1:1), and 1.1 mL of a mixed solution of hydrobromic acid / glacial acetic acid (33 wt%) was added at 0°C, and the reaction was continued for 1.5 h to obtain a reaction solution c. After the reaction was completed, the reaction solution c was precipitated in 80 mL of anhydrous ether, centrifuged again, and repeated 3 times, and finally dried in a vacuum oven to obtain 66.8 mg of polyethylene glycol-polylysine-poly(ε-benzyloxycarbonyl-L-glutamic acid) (solid), with a yield of 86.8%.

[0068] (3) Preparation of polyethylene glycol-poly(lysine-cyclodextrin)-poly(ε-benzyloxycarbonyl-L-glutamic acid)

[0069] 180 mg of cyclodextrin-carboxyl was dissolved in 10 mL of anhydrous DMF, and then 246 mg of N-hydroxysuccinimide and 408 mg of 1-(3-dimethylaminopropyl)-3-ethylcarbodiimide hydrochloride were added, and stirred for 2 hours in a nitrogen atmosphere to obtain a reaction solution d. Then, 57 mg of polyethylene glycol-polylysine-poly(ε-benzyloxycarbonyl-L-glutamic acid) was dissolved in 5 mL of anhydrous DMF and added dropwise to the reaction solution d, and the reaction was continued by stirring for 24 hours to obtain a reaction solution e. After the reaction was completed, the reaction solution e was precipitated in 80 mL of anhydrous ether, centrifuged again, repeated 3 times, and vacuum dried to obtain 131 mg of polyethylene glycol-poly(lysine-cyclodextrin)-poly(ε-benzyloxycarbonyl-L-glutamic acid) (solid), with a yield of 83.2%.

[0070] (4) Preparation of PEG-P(LL-CD)-PGlu

[0071] 100 mg of polyethylene glycol-poly(lysine-cyclodextrin)-poly(ε-benzyloxycarbonyl-L-glutamic acid) was dissolved in 20 mL of 1M NaOH solution and vigorously stirred overnight under reflux at 100° C. The resulting reaction mixture was placed in a dialysis bag and dialyzed with deionized water for 48 h. After dialysis, it was lyophilized to obtain 95.9 mg of PEG-P(LL-CD)-PGlu with a yield of 89.4%.

[0072] The PEG-P(LL-CD)-PGlu prepared in this example is as follows Figure 3 As shown, the detailed peak positions are attributed to: 1 H-NMR (400MHz, DMSO-d6), δ (ppm) = 7.71 (s, 40H, COCHNH), 5.08 (m, 240H, CHCOCH), 4.88 (m ,200H,OHCH2),4.33(m,240H,OHCH),3.68(m,40H,COCH2NH),3.51(s,180H,CH3O(CH2CH2) 45 ),3.37(m,40H,CDCONHCH2),3.24(m,3H,CH3),3.12(m,40H,OOCNHCH2CH2),2.83(m,80H,HOOCCH2CH2,OOCCH2),1.24(m,120H,CHCH2CH2CH2).

[0073] 3. Preparation of NP-CDDP / GOD

[0074] like Figure 4As shown, 5 mg PPL-Fc, 10 mg NP-CDDP / GOD, 6 mg cisplatin and 3.5 mg glucose oxidase were dissolved in 2 mL DMF, stirred for 6 h, and then 10 mL deionized water was added dropwise thereto, and stirred for another 12 h. After the reaction, the solution was placed in a dialysis bag with a molecular weight of 3500 and dialyzed with 500 mL deionized water for 1 day, with the deionized water replaced every 8 h. After the dialysis, the solution was freeze-dried to obtain a core-crosslinked nanomedicine (NP-CDDP / GOD) based on a dual enhanced cascade reaction, with a yield of 82.3%.

[0075] The dynamic light scattering spectrum of NP-CDDP / GOD prepared in this example is as follows: Figure 5 As shown, the number average particle size is 175.4±1.3 nm and the PDI is 0.39±0.06.

[0076] Example 2 Effect of NP-CDDP / GOD on liver cancer cells

[0077] 1. Experimental steps

[0078] (1) Group experiment

[0079] The nanomedicine (NP-CDDP / GOD group) was prepared in the manner of Example 1, and the following nanomedicine was prepared as a control group:

[0080] NP / GOD group: the nanomedicine was prepared in the same manner as step 3 in Example 1, except that cisplatin was not added;

[0081] NP-CDDP group: the nanomedicine was prepared in the same manner as step 3 of Example 1, except that glucose oxidase was not added;

[0082] NP group: Nanomedicine was prepared according to step 3 of Example 1, except that cisplatin and glucose oxidase were not added.

[0083] (2) The NP-CDDP / GOD, NP / GOD, NP-CDDP, NP and drug cisplatin (CDDP group) prepared above were respectively prepared with cell culture medium to prepare cisplatin concentration (CDDP) of 0.25, 0.5, 1, 2, 4, 8 μg / mL or corresponding nanoparticle concentration (NP) of 2.3, 4.6, 9.2, 18.4, 36.8, 73.6 μg / mL, and then the cell culture medium of each group was cultured with HepG2 cells (liver cancer) for 48 h.

[0084] 2. Experimental results

[0085] The cell activity test was performed using the MTT method. Figure 6In all experimental groups, regardless of the drug concentration, NP-CDDP / GOD showed the strongest cytotoxicity.

[0086] The experimental results of this example show that NP-CDDP / GOD can show better anti-tumor effect on tumor cells through the combined treatment of chemotherapy, chemodynamic therapy and starvation therapy.

[0087] Example 3 Effect of NP-CDDP / GOD on HepG2 tumor growth

[0088] 1. Experimental steps

[0089] The mice inoculated with HepG2 tumor were divided into 6 groups and injected with the nano drug prepared in Example 2, as follows:

[0090] (1) PBS group: injection of normal saline (5 mg / kg) into the tail vein;

[0091] (2) CDDP group: tail vein injection of cisplatin (5 mg / kg);

[0092] (3) NP group: tail vein injection of nanodrug NP;

[0093] (4) NP-CDDP group: tail vein injection of nanodrug NP-CDDP;

[0094] (5) NP / GOD group: tail vein injection of nanodrug NP / GOD;

[0095] (6) NP-CDDP / GOD group: tail vein injection of nanodrug NP-CDDP / GOD;

[0096] The mice injected with nanomedicine in the above groups were injected with cisplatin at a concentration of 5 mg / kg. The mice in all experimental groups were injected with the drug once on day 0 and day 4. The mice were weighed and the tumor volume was measured every 2 days.

[0097] 2. Experimental results

[0098] The experimental results are as follows Figure 7 , 8 As shown, for HepG2 tumors, the control groups (except the PBS group and the NP-CDDP / GOD group) had a certain inhibitory effect, but the inhibitory effect was significantly weaker than that of the NP-CDDP / GOD group. The NP-CDDP / GOD group showed a significant inhibitory effect on mouse tumors ( Figure 7 , P<0.01), and had almost no effect on the body weight of mice ( Figure 8 ).

[0099] The experimental results of this example show that the NP-CDDP / GOD of the present invention can be used to kill liver cancer cells (HepG2), and after two injections in the tail vein of mice, effective inhibition and treatment of HepG2 subcutaneous tumors can be achieved. Among the components of NP-CDDP / GOD, cisplatin-based chemotherapy, ferrocene-based chemodynamic therapy, glucose oxidase-based starvation therapy, and the combination of chemotherapy, chemodynamic therapy and starvation therapy have important clinical application prospects.

[0100] The embodiments described above are only some embodiments of the present invention, rather than all embodiments. The detailed description of the embodiments of the present invention is not intended to limit the scope of the invention claimed for protection, but merely represents selected embodiments of the present invention. The scope of protection of the present invention shall be subject to the scope required by the claims. Based on the embodiments of the present invention, all other embodiments obtained by ordinary technicians in this field without creative work shall fall within the scope of protection of the present invention.

Claims

1. A core-crosslinked nanomedicine based on a double-enhanced cascade reaction, characterized in that: The core-crosslinked nanomedicine is prepared from polylysine-ferrocene, polyethylene glycol-poly(lysine-cyclodextrin)-polyglutamic acid, glucose oxidase and cisplatin in a mass ratio of 1:2:1.2:0.7; wherein the structural formula of the polyethylene glycol-poly(lysine-cyclodextrin)-polyglutamic acid is shown in the following formula III: In formula III: n is 20.

2. A core-crosslinked nanomedicine based on a double enhanced cascade reaction according to claim 1, characterized in that: The particle size of the core-crosslinked nano drug is less than 200 nm.

3. A method for preparing a core-crosslinked nanomedicine based on a double enhanced cascade reaction according to claim 1 or 2, characterized in that: The following steps are involved: Step 1) preparing poly-lysine-ferrocene; Step 2) preparing polyethylene glycol-poly(lysine-cyclodextrin)-polyglutamic acid; Step 3) polylysine-ferrocene, polyethylene glycol-poly(lysine-cyclodextrin)-polyglutamic acid, glucose oxidase and cisplatin are dissolved in N,N-dimethylformamide solvent in proportion, deionized water is added dropwise, stirred, and dialyzed to obtain the product.

4. The method for preparing a core-crosslinked nanomedicine based on a double enhanced cascade reaction according to claim 3, characterized in that: The process of preparing polylysine-ferrocene in step 1) is shown in reaction formula II: The specific steps are as follows: Step 1-1) 40.4 mg of polylysine, 405.4 mg of N-hydroxysuccinimide and 619.2 mg of 1-(3-dimethylaminopropyl)-3-ethylcarbodiimide hydrochloride were dissolved in 10 mL of anhydrous N,N-dimethylformamide and stirred for 4 h in a nitrogen atmosphere to obtain a reaction solution A; Step 1-2) Add 107 mg of ferrocene carboxaldehyde to reaction solution A, and continue stirring the reaction for 24 hours to obtain reaction solution B; Step 1-3) The reaction solution B was precipitated in 80 mL of anhydrous ether, and then centrifuged, repeated 3 times, and vacuum dried to obtain.

5. The method for preparing a core-crosslinked nanomedicine based on a double enhanced cascade reaction according to claim 3, characterized in that: The process of preparing polyethylene glycol-poly(lysine-cyclodextrin)-polyglutamic acid in step 2) is shown in reaction formula IV: The specific steps are as follows: Step 2-1) 200 mg of polyethylene glycol amino and 244.9 mg of ε-benzyloxycarbonyl-L-lysine anhydride were dissolved in 2 mL of anhydrous N,N-dimethylformamide, and stirred for reaction for 48 h in a nitrogen atmosphere to obtain a reaction solution a; 210.5 mg of γ-benzyl-glutamic acid anhydride was added to the reaction solution a, and the reaction was continued with stirring for 48 h to obtain a reaction solution b; the reaction solution b was precipitated in 16 mL of anhydrous ether, and then centrifuged, repeated 3 times, and vacuum dried to obtain polyethylene glycol-poly(ε-benzyloxycarbonyl-L-lysine)-poly(ε-benzyloxycarbonyl-L-glutamic acid); Step 2-2) 100 mg of polyethylene glycol-poly(ε-benzyloxycarbonyl-L-lysine)-poly(ε-benzyloxycarbonyl-L-glutamic acid) was dissolved in 10 mL of a mixed solvent of glacial acetic acid / trifluoroacetic acid, and 1.1 mL of a mixed solution of hydrobromic acid / glacial acetic acid was added at 0° C. The reaction was continued for 1.5 h to obtain a reaction solution c; the reaction solution c was precipitated in 80 mL of anhydrous ether, and then centrifuged, repeated 3 times, and finally dried in a vacuum oven to obtain polyethylene glycol-polylysine-poly(ε-benzyloxycarbonyl-L-glutamic acid); Step 2-3) 180 mg of cyclodextrin-carboxyl was dissolved in 10 mL of anhydrous N,N-dimethylformamide, and then 246 mg of N-hydroxysuccinimide and 408 mg of 1-(3-dimethylaminopropyl)-3-ethylcarbodiimide hydrochloride were added, and stirred for 2 h in a nitrogen atmosphere to obtain a reaction solution d; then 57 mg of polyethylene glycol-polylysine-poly(ε-benzyloxycarbonyl-L-glutamic acid) was dissolved in 5 mL of anhydrous N,N- Dimethylformamide is added dropwise to the reaction solution d, and the reaction is continued with stirring for 24 hours to obtain a reaction solution e; the reaction solution e is precipitated in 80 mL of anhydrous ether, centrifuged again, repeated 3 times, and vacuum dried to obtain polyethylene glycol-poly(lysine-cyclodextrin)-poly(ε-benzyloxycarbonyl-L-glutamic acid); step 2-4) 100 mg of polyethylene glycol-poly(lysine-cyclodextrin)-poly(ε-benzyloxycarbonyl-L-glutamic acid) is dissolved in 20 mL of 1M NaOH solution, and vigorously stirred at reflux at 100° C. overnight; the resulting reaction mixture is placed in a dialysis bag, dialyzed with deionized water for 48 hours, and freeze-dried after dialysis to obtain.

6. The method for preparing a core-crosslinked nanomedicine based on a double enhanced cascade reaction according to claim 3, characterized in that: Step 3) The concentration of the polylysine-ferrocene is 2.5 mg / mL; the concentration of the polyethylene glycol-poly(lysine-cyclodextrin)-polyglutamic acid is 5 mg / mL; the concentration of the glucose oxidase is 3.0 mg / mL; and the concentration of the cisplatin is 1.75 mg / mL.

7. The method for preparing a core-crosslinked nanomedicine based on a double enhanced cascade reaction according to claim 3, characterized in that: Step 3) The volume ratio of the N,N-dimethylformamide to the deionized water is 1:

5.

8. The method for preparing a core-crosslinked nanomedicine based on a double enhanced cascade reaction according to claim 3, characterized in that: Step 3) The dialysis conditions are: using a dialysis bag with a molecular weight of 3500, 500 mL of deionized water × 1 time / 8 hours, and dialysis for 24 hours.

9. Use of the core-crosslinked nanomedicine based on a double enhanced cascade reaction as described in claim 1 or 2, or the core-crosslinked nanomedicine based on a double enhanced cascade reaction prepared by the preparation method according to any one of claims 3 to 8 in the preparation of anti-tumor drugs.

10. An anti-tumor pharmaceutical composition, characterized in that: It comprises the core-crosslinked nanomedicine based on the double-enhanced cascade reaction as described in claim 1 or 2, or the core-crosslinked nanomedicine based on the double-enhanced cascade reaction prepared by the preparation method according to any one of claims 3 to 8, and pharmaceutically acceptable excipients.

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