Prodrug nano delivery system, drug containing prodrug nano delivery system and application thereof

Abstract

The present invention relates to the field of nanomedicine technology, and in particular to a prodrug nano delivery system, a drug containing the prodrug nano delivery system, and an application thereof. The preparation process of the prodrug nano delivery system is as follows: folic acid is mixed with triethylamine for reaction, and then the active ester of folic acid N-hydroxysuccinimide is obtained, and then the active ester is reacted with cysteamine to obtain a cysteamine-folate conjugate; a carboxylated ginsenoside substance is reacted with carboxymethyl chitosan to obtain a carboxymethyl chitosan-carboxylated ginsenoside graft; amino-terminated poly N-isopropylacrylamide and carboxymethyl chitosan-carboxylated ginsenoside graft are reacted as raw materials, and the obtained product solution is subjected to ultrasonic treatment and then a photosensitizer is added, and a nano-prodrug micelle is obtained and then mixed with a cysteamine-folate conjugate to obtain. In the prodrug nano delivery system, folic acid binds to the overexpressed folate receptor on cancer cells, and after loading the drug, the tumor is targeted for release, glutathione in liver cancer cells is consumed, and the effect of the combined treatment of drugs and photodynamic therapy is enhanced.

Description

Technical Field

[0001] The present invention belongs to the technical field of nano-drugs, and specifically relates to a prodrug nano-delivery system with a structure similar to a virus capsid particle, a drug containing the prodrug nano-delivery system, and an application thereof in the preparation of an anti-tumor drug. Background Art

[0002] Hepatocellular carcinoma is one of the most common primary tumors and has become the fourth leading cause of cancer death in the world. The 5-year overall survival rate of patients with hepatocellular carcinoma is only 10% to 18%. At present, the treatment of liver cancer mainly involves surgery, chemotherapy and radiotherapy, but liver cancer is usually discovered in the middle and late stages. Ordinary chemotherapy drugs have significant side effects and patients are in great pain. In order to reduce the side effects of chemotherapy drugs, more and more people are focusing their research on natural active drugs.

[0003] Ginsenoside CK is the main active substance in ginseng, which has excellent biological activities such as anti-aging, anti-inflammatory, antioxidant, liver protection, immunity enhancement and anti-tumor. When used in tumor treatment, patients have a high degree of acceptance. However, due to its poor water solubility, poor stability and low bioavailability, ginsenoside CK has greatly limited its application in the biomedical field.

[0004] Prodrugs are compounds that release active drugs in vivo after chemical structure modification. Compared with traditional drugs, prodrugs have the advantages of good solubility and high bioavailability. Prodrug design is an effective strategy to improve the stability and bioavailability of hydrophobic drugs and overcome the disadvantages of difficult delivery of hydrophobic drugs. Carboxymethyl chitosan is an amphiphilic derivative of chitosan with excellent biocompatibility and biodegradability. It is easy to process into nanoparticles, and its rich functional groups are easy to modify, making it commonly used in drug delivery systems, which is receiving more and more attention.

[0005] New indocyanine green (IR820) is a cyanine dye that is sensitive to near-infrared light and can absorb 808nm near-infrared light for photothermal conversion and laser-induced ROS singlet oxygen to kill tumor cells, but the new indocyanine green has a short half-life in the blood, rapid clearance, and non-specific biodistribution in the body. It is easy to cause systemic toxicity and low therapeutic efficiency during phototherapy. CN112641959A discloses a method for preparing a novel multifunctional nanoprobe, which uses gold nanoparticles to load gadolinium ions and new indocyanine green IR820 for photothermal and photodynamic synergistic therapy. However, compared with natural products and polymer compounds, gold nanoparticles are more expensive and easily oxidized, losing their characteristics, and nanoprobes do not add active or passive targeting functions, and their actual effects remain to be verified.

[0006] Folic acid (FA) can bind to overexpressed folate receptors on cancer cells, and targeted drug delivery through folate receptors becomes a biocompatible and targeted cancer treatment method. CN117122692A discloses a targeted nanocarrier modified with FA molecules, which promotes the enrichment of nanomaterials in tumor cells. Designing a nanocarrier with the function of targeting folate receptors and delivering targeted drugs to tumors after loading them can provide new ideas for efficient drug delivery. CN112451663A discloses a method for preparing a nanocomposite for fluorescent imaging navigation tumor resection and photothermal therapy, in which the tumor targeting protein TP is physically or chemically modified with a silicone layered structure loaded with near-infrared dye IR820 to form a nanocomposite. This patent takes into account the diagnosis and treatment of targeted tumor sites, but only relies on photothermal therapy and does not carry tumor treatment drugs.

[0007] In addition, in the process of killing cancer cells by indocyanine green laser-induced ROS, glutathione (GSH) maintains the dynamic balance between the generation and removal of intracellular ROS through its antioxidant function. Therefore, the GSH concentration in cancer cells can generally reach 10mM, which is at least 4 times or more than that of normal cells, and is more obvious in some drug-resistant cancer cells and cancer stem cells. CN112773895B discloses a nanoreactor composed of a metal organic framework, a photosensitizer, glucose oxidase and a polylactic acid-glycolic acid copolymer wrapped by a metal organic framework, which can catalyze oxygen production to achieve in situ oxygen supply to the tumor, remove glutathione and ATP, thereby inhibiting tolerance and enhancing the efficiency of photodynamic therapy. However, this effect must be based on the ability to target tumor cells, and the patent does not have relevant discussion.

[0008] Therefore, the present invention intends to develop a targeted drug delivery system that can overcome the above-mentioned defects of the prior art. Summary of the invention

[0009] In view of the shortcomings of the above-mentioned technologies, the present invention provides a prodrug nano drug delivery system with a virus-like capsid particle structure and its application. The prodrug nano micelle is a virus-like nucleocapsid structure. When the photosensitizer in the prodrug nano drug delivery system that reaches the lesion produces ROS to kill cancer cells when irradiated with near-infrared light, it can also perform photothermal conversion, causing the phase change of the thermosensitive agent, and the collapse of the polymer nucleocapsid to induce the release of the loaded chemotherapy drug ginsenoside, ensuring the release of the chemotherapy drug ginsenoside at the lesion site, and strengthening the combined medication effect of phototherapy and chemotherapy.

[0010] Based on a general inventive concept, the present invention provides a prodrug nano drug delivery system with a virus-like capsid particle structure, which is prepared according to the following steps:

[0011] The folic acid and triethylamine are mixed for amidation reaction to obtain folic acid N-hydroxysuccinimide active ester; the folic acid N-hydroxysuccinimide active ester and cysteamine are used as raw materials for amidation reaction to obtain a cysteamine-folic acid conjugate;

[0012] Carboxylating the hydroxyl groups on the ginsenoside compounds to obtain carboxylated ginsenoside substances; further mixing the carboxylated ginsenoside substances with carboxymethyl chitosan for amidation reaction to obtain carboxymethyl chitosan-carboxylated ginsenoside grafts;

[0013] polymerizing N-isopropylacrylamide monomers to obtain amino-terminated poly-N-isopropylacrylamide;

[0014] Using the amino-terminated poly N-isopropylacrylamide and the carboxymethyl chitosan-carboxylated ginsenoside graft as raw materials, an amidation reaction is carried out to obtain a carboxymethyl chitosan-ginsenoside-poly N-isopropylacrylamide graft;

[0015] The solution of carboxymethyl chitosan-ginsenoside-poly (N-isopropylacrylamide) grafted product is ultrasonically treated to self-assemble into a capsid structure, and a photosensitizer is added to encapsulate the photosensitizer in the formed capsid structure to obtain nano-prodrug micelles;

[0016] The nano prodrug micelles are mixed with the cysteamine-folic acid conjugate for electrostatic adsorption to obtain a prodrug nano drug delivery system with a virus capsid particle structure.

[0017] As a preferred embodiment of the present invention, the preparation of the prodrug nano drug delivery system can be carried out according to the following steps:

[0018] Add folic acid to a mixed solution of DMSO and triethylamine in a volume ratio of 100:1, stir, mix with an activator, stir, and remove impurities to obtain folic acid N-hydroxysuccinimide active ester;

[0019] The solution of folic acid N-hydroxysuccinimide active ester is mixed with cysteamine, stirred, then washed with ether and dried to obtain a cysteamine-folic acid conjugate;

[0020] The ginsenoside compounds are dissolved in a mixed solvent of pyridine and dichloromethane, succinic anhydride is added, the mixture is stirred, rotary evaporated, pure water is added to the precipitate, filtered and vacuum dried to obtain white carboxylated ginsenoside powder;

[0021] The carboxylated ginsenoside powder is dissolved in methanol, mixed with an activator, and stirred to obtain a carboxylated ginsenoside active ester solution; then, O-carboxymethyl chitosan is added, the reaction is continued, and the dialyzed solution is dialyzed in excess methanol / pure water = volume ratio 1:1, the dialyzed solution is left overnight, and finally the dialyzate is dried to obtain an O-carboxymethyl chitosan-carboxylated ginsenoside graft;

[0022] The recrystallized N-isopropylacrylamide monomer is dissolved in a nitrogen atmosphere, mixed with an activator and a catalyst, stirred, dialyzed overnight, and dried to obtain amino-terminated poly-N-isopropylacrylamide;

[0023] Adding an activator to the aqueous solution of O-carboxymethyl chitosan-carboxylated ginsenoside, stirring, adding amino-terminated poly N-isopropylacrylamide, amidation reaction, dialyzing, and drying to obtain O-carboxymethyl chitosan-ginsenoside-poly N-isopropylacrylamide grafted product;

[0024] The solution of O-carboxymethyl chitosan-ginsenoside-poly (N-isopropylacrylamide) grafted product was mixed with IR820 dissolved in dimethyl sulfoxide, stirred, sonicated in an ice bath, dialyzed, and dried to obtain nano-prodrug micelles;

[0025] The aqueous solution of the nano-prodrug micelles is mixed with the cysteamine-folic acid conjugate, and reacted in the dark to obtain a folic acid-targeted prodrug nano-drug delivery system.

[0026] As a preferred embodiment of the present invention, the ginsenoside substance is any one of ginsenoside CK, ginsenoside Re, ginsenoside Rg3, ginsenoside Rg5, ginsenoside Rh2, ginsenoside Rh3, ginsenoside Rh1 or ginsenoside Rc.

[0027] As a preferred embodiment of the present invention, the molar ratio of the carboxymethyl chitosan to the carboxylated ginsenosides is 6-14:1.

[0028] More preferably, the molar ratio of the carboxymethyl chitosan to the carboxylated ginsenosides is 10:1.

[0029] As a preferred embodiment of the present invention, the mass ratio of the carboxymethyl chitosan-carboxylated ginsenoside graft to the amino-terminated poly (N-isopropylacrylamide) is 1-2.5:2.

[0030] More preferably, the mass ratio of the carboxymethyl chitosan-carboxylated ginsenoside graft to the amino-terminated poly (N-isopropylacrylamide) is 1:1.

[0031] As a preferred embodiment of the present invention, the photosensitizer is any one of new indocyanine green, pyropheophytin-a (Ppa) or mono-asparagine dihydrochlorin-e6 (NPe6). The photosensitizer generates cytotoxic reactive oxygen species under the irradiation of light of a specific wavelength. The photosensitizer can also be selected from other types that have a selective killing effect on tumor cells, and the present invention is not limited to this.

[0032] As a preferred embodiment of the present invention, the concentration of the solution of carboxymethyl chitosan-ginsenoside-poly N-isopropylacrylamide grafts is 1 mg / mL, the added concentration of the photosensitizer is 2 mg / mL, and the mixed volume ratio of the solution of carboxymethyl chitosan-ginsenoside-poly N-isopropylacrylamide grafts to the solution of the photosensitizer is 5:0.2-1.6.

[0033] More preferably, the mixing volume ratio of the solution of carboxymethyl chitosan-ginsenoside-poly (N-isopropylacrylamide) grafts to the solution of the photosensitizer is 5:0.8.

[0034] As a preferred embodiment of the present invention, the activator is a mixture of 1-(3-dimethylaminopropyl)-3-ethylcarbodiimide hydrochloride (EDC) and N-hydroxysuccinimide (NHS).

[0035] As a preferred embodiment of the present invention, the mass ratio of folic acid:EDC:NHS is 0.25g:0.1g:0.1g.

[0036] As a preferred embodiment of the present invention, the mass ratio of folic acid:cysteamine is 0.25g:0.1g.

[0037] As a preferred embodiment of the present invention, in the preparation process of amino-terminated poly (N-isopropylacrylamide), the activator used is 1-(3-dimethylaminopropyl)-3-ethylcarbodiimide hydrochloride, the catalyst is potassium persulfate, and the polymerization occurs during stirring at 60° C. and 600 r / min in a nitrogen atmosphere.

[0038] As a preferred embodiment of the present invention, the dialysis is performed using a dialysis bag with a MW of 12000-14000Da for 24-48 hours.

[0039] As a preferred embodiment of the present invention, the electrostatic adsorption is carried out under light-proof conditions at a stirring speed of 600 r / min for 3 to 6 hours; after the light-proof reaction is completed, the reaction solution is dialyzed for 24 hours using a dialysis membrane with a molecular weight of 8000 to 14000 Da.

[0040] As a preferred embodiment of the present invention, the average particle size of the prodrug nanomicelles is 239.3 nm.

[0041] The present invention also provides an application of the prodrug nano drug delivery system in the preparation of photodynamic therapy drugs.

[0042] As a preferred embodiment of the present invention, the prodrug nano drug delivery system is used to prepare anti-tumor drugs.

[0043] As a preferred embodiment of the present invention, the prodrug nano drug delivery system is used to prepare anti-liver cancer drugs.

[0044] The present invention also provides a drug for treating tumors, the drug comprising the prodrug nano drug delivery system, and a pharmaceutically acceptable excipient or carrier. The selection of excipients and carriers is specifically selected according to the drug to be prepared. The drug is an external preparation, an oral preparation or an injection. Preferably, the external preparation is an external gel. The oral preparation is a granule, a tablet, an oral solution, etc. The injection is an intravenous injection containing the nano drug delivery system.

[0045] Compared with the prior art, the present invention has the following beneficial effects:

[0046] 1. The present invention provides a prodrug nano drug delivery system, which is a virus-like nucleocapsid structure. The "outer membrane" of the prodrug nano drug delivery system is a cysteamine-folic acid conjugate, the nano micelles self-assembled by carboxymethyl chitosan-poly N-isopropyl acrylamide polymer are the "capsid", and the chemotherapy drug ginsenosides and new indocyanine green are the "core". Folic acid can guide the prodrug nano drug delivery system to target tumor cells, and cysteamine forms a disulfide bond when acting with the "nucleocapsid", and the disulfide bond can respond to the cleavage of glutathione in tumor cells, thereby clearing glutathione in tumor cells. When irradiated with near-infrared light, the photosensitizer in the prodrug nano micelles that reach the lesion produces ROS to kill cancer cells, and at the same time, it can perform photothermal conversion, causing the phase change of the thermosensitive agent poly N-isopropyl acrylamide, and the polymer core-shell collapses to induce the release of the loaded chemotherapy drug, ensuring that the chemotherapy drug ginsenoside CK is released at the lesion site, achieving the synergistic effect of phototherapy and chemotherapy.

[0047] 2. The present invention uses ginsenoside compounds as the main chemotherapy drugs and photosensitizers such as indocyanine green as the main phototherapy drugs in a composite multifunctional nanosystem, utilizes the ultrasonic self-assembly characteristics of chitosan to form nanomicelles, and wraps them layer by layer with folic acid cysteamine conjugates for stabilization, thereby preparing a prodrug nano delivery system, which is a method for preparing nano prodrugs with simple process and high stability.

[0048] 3. The present invention provides an application of a prodrug nano drug delivery system, wherein the "outer membrane" of the prodrug nano drug delivery system is a folic acid cysteamine conjugate, and folic acid can guide the prodrug nano micelles to target tumor cells, increase the tumor cells' uptake of the coordination polymer, and thus further enhance the targeted delivery of the drug.

[0049] 4. In the prodrug nano-drug delivery system provided by the present invention, the disulfide bond formed by the cysteamine in the folic acid cysteamine conjugate when reacting with the "nucleocapsid" can be broken in response to glutathione in tumor cells, thereby consuming glutathione in tumor cells, and continuously consuming glutathione produced by the oxidative stress state caused by ROS, thereby overcoming cell tolerance, enhancing the killing efficiency of photodynamic therapy, and improving the therapeutic effect on tumors.

[0050] In order to more clearly illustrate the embodiments of the present invention or the technical solutions in the prior art, the present invention is further described in detail below with reference to the accompanying drawings and embodiments. BRIEF DESCRIPTION OF THE DRAWINGS

[0051] Figure 1 The Fourier transform infrared chromatograms of Cys and FA-Cys conjugate in Example 1 are shown.

[0052] Figure 2 This is the hydrogen nuclear magnetic resonance spectrum of FA-Cys in Example 1.

[0053] Figure 3 This is an electron microscopic morphology of the prodrug nano-delivery system with a virus-like capsid particle structure of Example 1.

[0054] Figure 4 This is the particle size distribution diagram of the prodrug nano-delivery system with a virus-like capsid particle structure in Example 1.

[0055] Figure 5 This is the Zeta potential distribution diagram of the prodrug nano-delivery system with a virus-like capsid particle structure in Example 1.

[0056] Figure 6 This is the singlet reactive oxygen species generation detection of the prodrug nano-delivery system with a virus-like capsid particle structure of Example 2.

[0057] Figure 7 This is the toxicity test of the virus-like capsid particle structure of Example 3 on liver cancer HepG-2 cells.

[0058] Figure 8 This is a laser confocal image of the prodrug nano-drug delivery system with a virus-like capsid particle structure in Example 4 after treating liver cancer HepG-2 cells.

[0059] Fig. 9 This is an image of HepG-2 cell apoptosis induced by the virus-like capsid particle prodrug structure nano-drug delivery system of Example 5. DETAILED DESCRIPTION

[0060] The present invention is further described below with the aid of non-limiting examples.

[0061] The present invention provides a prodrug nano drug delivery system with a virus-like capsid particle structure, which is prepared according to the following steps:

[0062] The folic acid and triethylamine are mixed for amidation reaction to obtain folic acid N-hydroxysuccinimide active ester; the folic acid N-hydroxysuccinimide active ester and cysteamine are used as raw materials for amidation reaction to obtain a cysteamine-folic acid conjugate;

[0063] Carboxylating the hydroxyl groups on the ginsenoside compounds to obtain carboxylated ginsenoside substances; further mixing the carboxylated ginsenoside substances with carboxymethyl chitosan for amidation reaction to obtain carboxymethyl chitosan-carboxylated ginsenoside grafts;

[0064] polymerizing N-isopropylacrylamide monomers to obtain amino-terminated poly-N-isopropylacrylamide;

[0065] Using the amino-terminated poly N-isopropylacrylamide and the carboxymethyl chitosan-carboxylated ginsenoside graft as raw materials, an amidation reaction is carried out to obtain a carboxymethyl chitosan-ginsenoside-poly N-isopropylacrylamide graft;

[0066] The solution of carboxymethyl chitosan-ginsenoside-poly (N-isopropylacrylamide) grafted product is ultrasonically treated to self-assemble into a capsid structure, and a photosensitizer is added to encapsulate the photosensitizer in the formed capsid structure to obtain nano-prodrug micelles;

[0067] The nano prodrug micelles are mixed with the cysteamine-folic acid conjugate for electrostatic adsorption to obtain a prodrug nano drug delivery system with a virus capsid particle structure.

[0068] The "outer membrane" of the prodrug nano drug delivery system with a virus-like capsid particle structure provided by the present invention is a cysteamine-folic acid conjugate, the nano micelles self-assembled by carboxymethyl chitosan-poly N-isopropylacrylamide polymer are the "capsid", and the chemotherapy drug ginsenosides and new indocyanine green are the "core". Folic acid can guide the prodrug nano drug delivery system to target tumor cells, and cysteamine forms a disulfide bond when acting with the "nucleocapsid", and the disulfide bond can respond to the cleavage of glutathione in tumor cells, thereby clearing glutathione in tumor cells. When irradiated with near-infrared light, when the photosensitizer in the prodrug nano micelles that reach the lesion produces ROS to kill cancer cells, it can also perform photothermal conversion, causing the phase change of the thermosensitive agent poly N-isopropylacrylamide, and the polymer core-shell collapses to induce the release of the loaded chemotherapy drug, ensuring that the chemotherapy drug ginsenoside CK is released at the lesion site, achieving the synergistic effect of phototherapy and chemotherapy.

[0069] Example 1

[0070] The viral capsid-like prodrug nano-drug delivery system is prepared according to the following steps:

[0071] S1. Preparation of folic acid N-hydroxysuccinimide active ester:

[0072] A solution of DMSO and triethylamine in a volume ratio of 100:1 was stirred for 10 minutes. 0.25 g of folic acid was gradually added to the above mixture and stirred continuously overnight in the dark. The folic acid solution was mixed with 0.1 g of EDC (1-(3-dimethylaminopropyl)-3-ethylcarbodiimide hydrochloride) and 0.1 g of NHS (N-hydroxysuccinimide) and stirred for another 24 hours, and the by-products were removed by filtration. DMSO and triethylamine were evaporated in vacuo to obtain folic acid N-hydroxysuccinimide active ester, recorded as FA-NHS;

[0073] S2. Preparation of cysteamine-folic acid conjugate:

[0074] Vacuum-dried FA-NHS was dissolved in a mixture of DMSO and trimethylamine (2:1). Then, 0.1 g of cysteamine was added to the mixture and stirred overnight. The resulting yellow solid cysteamine-folate conjugate was filtered and then washed twice with ether to obtain cysteamine-folate conjugate, recorded as FA-Cys;

[0075] The reaction process is as follows:

[0076]

[0077] S3, dissolve 100 mg of ginsenoside CK in 20 mL of a 1:1 mixture of pyridine and dichloromethane, add 48 g of succinic anhydride, stir at 600 r / min, rotary evaporate, add 30 mL of pure water to the precipitate, filter and vacuum dry to obtain white CK-COOH powder;

[0078] S4. CK-COOH powder was dissolved in methanol again, mixed with 1-(3-dimethylaminopropyl)-3-ethylcarbodiimide hydrochloride and N-hydroxysuccinimide, and stirred for 1 hour to obtain a carboxylated ginsenoside CK active ester solution; then, O-carboxymethyl chitosan was added, the reaction was continued for 24 hours, and dialyzed in excess methanol / pure water = volume ratio of 1:1 for 48 hours. The MW of the dialysis membrane was 12000-14000Da. The dialyzed solution was placed in a -20°C refrigerator overnight, and finally the dialyzate was freeze-dried for 48 hours to obtain O-carboxymethyl chitosan-carboxylated ginsenoside CK (OCMC-CK) graft; the molar ratio of O-carboxymethyl chitosan: carboxylated ginsenoside CK active ester solution was 10:1.

[0079] S5, dissolving 5.794 g of recrystallized N-isopropylacrylamide monomer under a nitrogen atmosphere, mixing with 1.4 mg of 1-ethyl-3-(3-dimethylaminopropyl)carbodiimide hydrochloride and 7.4 mg of potassium persulfate dissolved in 10 mL of pure water, stirring at 600 r / min at 60° C. and a nitrogen atmosphere, dialyzing, placing in a refrigerator overnight, and freeze-drying for 48 h to obtain amino-terminated poly-N-isopropylacrylamide;

[0080] S6. The O-carboxymethyl chitosan-ginsenoside CK aqueous solution was mixed with 40 mg of 1-(3-dimethylaminopropyl)-3-ethylcarbodiimide hydrochloride and 24 mg of N-hydroxysuccinimide, and the mixture was stirred for 1 h. Then, amino-terminated poly (N-isopropylacrylamide) was added, and the reaction was continued for 24 h. The mixture was dialyzed, and the MW of the dialysis membrane was 12000-14000 Da. The mixture was freeze-dried for 48 h to obtain O-carboxymethyl chitosan-ginsenoside CK-poly (N-isopropylacrylamide) (PNIPAM-OCMC-CK) grafted product. The ratio of O-carboxymethyl chitosan-ginsenoside CK to amino-terminated poly (N-isopropylacrylamide) was 200 mg:200 mg.

[0081] The synthetic route of O-carboxymethyl chitosan-ginsenoside CK-poly N-isopropylacrylamide polymer carrier is as follows:

[0082]

[0083] S7. The PNIPAM-OCMC-CK solution and IR820 dissolved in dimethyl sulfoxide were stirred, ultrasonicated for 3 h in an ice bath at 120 W, dialyzed for 30 min, and the MW of the dialysis membrane was 12000-14000 Da. After freeze-drying, OCMC-CK-PNIPAM@IR820 nanoprodrug micelles were obtained; the concentration of the PNIPAM-OCMC-CK solution was 1 mg / mL, the concentration of the IR820 solution was 2 mg / mL, and the mixing ratio of the two solutions was 5 mL:0.8 mL.

[0084] S8. Mix the aqueous solution of OCMC-CK-PNIPAM@IR820 nanoprodrug micelles with cysteamine-folate conjugates, and react them in the dark for 6 hours at a stirring speed of 600 r / min. After the reaction is completed, the reaction solution is dialyzed for 24 hours using a dialysis membrane with a molecular weight of 8000-14000 Da to obtain a folic acid-targeted prodrug nano-delivery system FA-Cys / OCMC-CK-PNIPAM@IR820. The concentration of the aqueous solution of the nanoprodrug micelles is 2 mg / mL, the concentration of the cysteamine-folate conjugate solution is 0.5 mg / mL, and the mixing ratio of the two solutions is 5 mL:5 mL.

[0085] Example 2

[0086] The prodrug nano drug delivery system similar to the viral capsid particle is different from Example 1 only in the preparation process of the O-carboxymethyl chitosan-ginsenoside CK-poly N-isopropylacrylamide polymer, specifically:

[0087] S3, dissolve 100 mg of ginsenoside CK in 20 mL of a 1:1 mixture of pyridine and dichloromethane, add 48 g of succinic anhydride, stir at 600 r / min, rotary evaporate, add 30 mL of pure water to the precipitate, filter and vacuum dry to obtain white CK-COOH powder;

[0088] S4. CK-COOH powder was dissolved in methanol again, mixed with 1-(3-dimethylaminopropyl)-3-ethylcarbodiimide hydrochloride and N-hydroxysuccinimide, and stirred for 1 hour to obtain a carboxylated ginsenoside CK active ester solution; then, O-carboxymethyl chitosan was added, the reaction was continued for 24 hours, and dialyzed in excess methanol / pure water = volume ratio of 1:1 for 48 hours. The MW of the dialysis membrane was 12000-14000Da. The dialyzed solution was placed in a -20°C refrigerator overnight, and finally the dialyzate was freeze-dried for 48 hours to obtain O-carboxymethyl chitosan-carboxylated ginsenoside CK (OCMC-CK) graft; the molar ratio of O-carboxymethyl chitosan: carboxylated ginsenoside CK active ester solution was 6:1.

[0089] S5, dissolving 5.794 g of recrystallized N-isopropylacrylamide monomer under a nitrogen atmosphere, mixing with 1.4 mg of 1-ethyl-3-(3-dimethylaminopropyl)carbodiimide hydrochloride and 7.4 mg of potassium persulfate dissolved in 10 mL of pure water, stirring at 600 r / min at 60° C. and a nitrogen atmosphere, dialyzing, placing in a refrigerator overnight, and freeze-drying for 48 h to obtain amino-terminated poly-N-isopropylacrylamide;

[0090] S6. The O-carboxymethyl chitosan-ginsenoside CK aqueous solution was mixed with 40 mg of 1-(3-dimethylaminopropyl)-3-ethylcarbodiimide hydrochloride and 24 mg of N-hydroxysuccinimide, and the mixture was stirred for 1 h. Then, amino-terminated poly (N-isopropylacrylamide) was added, and the reaction was continued for 24 h. The mixture was dialyzed, and the MW of the dialysis membrane was 12000-14000 Da. The mixture was freeze-dried for 48 h to obtain O-carboxymethyl chitosan-ginsenoside CK-poly (N-isopropylacrylamide) (PNIPAM-OCMC-CK) grafted product. The ratio of O-carboxymethyl chitosan-ginsenoside CK to amino-terminated poly (N-isopropylacrylamide) was 200 mg:200 mg.

[0091] Example 3

[0092] The prodrug nano drug delivery system similar to the viral capsid particle is different from Example 1 only in the preparation process of the O-carboxymethyl chitosan-ginsenoside CK-poly N-isopropylacrylamide polymer, specifically:

[0093] S3, dissolve 100 mg of ginsenoside CK in 20 mL of a 1:1 mixture of pyridine and dichloromethane, add 48 g of succinic anhydride, stir at 600 r / min, rotary evaporate, add 30 mL of pure water to the precipitate, filter and vacuum dry to obtain white CK-COOH powder;

[0094] S4. CK-COOH powder was dissolved in methanol again, mixed with 1-(3-dimethylaminopropyl)-3-ethylcarbodiimide hydrochloride and N-hydroxysuccinimide, and stirred for 1 hour to obtain a carboxylated ginsenoside CK active ester solution; then, O-carboxymethyl chitosan was added, the reaction was continued for 24 hours, and dialyzed in excess methanol / pure water = volume ratio of 1:1 for 48 hours. The MW of the dialysis membrane was 12000-14000Da. The dialyzed solution was placed in a -20°C refrigerator overnight, and finally the dialyzate was freeze-dried for 48 hours to obtain O-carboxymethyl chitosan-carboxylated ginsenoside CK (OCMC-CK) graft; the molar ratio of O-carboxymethyl chitosan: carboxylated ginsenoside CK active ester solution was 14:1.

[0095] S5, dissolving 5.794 g of recrystallized N-isopropylacrylamide monomer under a nitrogen atmosphere, mixing with 1.4 mg of 1-ethyl-3-(3-dimethylaminopropyl)carbodiimide hydrochloride and 7.4 mg of potassium persulfate dissolved in 10 mL of pure water, stirring at 600 r / min at 60° C. and a nitrogen atmosphere, dialyzing, placing in a refrigerator overnight, and freeze-drying for 48 h to obtain amino-terminated poly-N-isopropylacrylamide;

[0096] S6. The O-carboxymethyl chitosan-ginsenoside CK aqueous solution was mixed with 40 mg of 1-(3-dimethylaminopropyl)-3-ethylcarbodiimide hydrochloride and 24 mg of N-hydroxysuccinimide, and the mixture was stirred for 1 h. Then, amino-terminated poly (N-isopropylacrylamide) was added, and the reaction was continued for 24 h. The mixture was dialyzed, and the MW of the dialysis membrane was 12000-14000 Da. The mixture was freeze-dried for 48 h to obtain O-carboxymethyl chitosan-ginsenoside CK-poly (N-isopropylacrylamide) (PNIPAM-OCMC-CK) grafted product. The ratio of O-carboxymethyl chitosan-ginsenoside CK to amino-terminated poly (N-isopropylacrylamide) was 200 mg:200 mg.

[0097] Example 4

[0098] The prodrug nano drug delivery system similar to the viral capsid particle is different from Example 1 only in the preparation process of the O-carboxymethyl chitosan-ginsenoside CK-poly N-isopropylacrylamide polymer, specifically:

[0099] S3, dissolve 100 mg of ginsenoside CK in 20 mL of a 1:1 mixture of pyridine and dichloromethane, add 48 g of succinic anhydride, stir at 600 r / min, rotary evaporate, add 30 mL of pure water to the precipitate, filter and vacuum dry to obtain white CK-COOH powder;

[0100] S4. CK-COOH powder was dissolved in methanol again, mixed with 1-(3-dimethylaminopropyl)-3-ethylcarbodiimide hydrochloride and N-hydroxysuccinimide, and stirred for 1 hour to obtain a carboxylated ginsenoside CK active ester solution; then, O-carboxymethyl chitosan was added, the reaction was continued for 24 hours, and dialyzed in excess methanol / pure water = volume ratio of 1:1 for 48 hours. The MW of the dialysis membrane was 12000-14000Da. The dialyzed solution was placed in a -20°C refrigerator overnight, and finally the dialyzate was freeze-dried for 48 hours to obtain O-carboxymethyl chitosan-carboxylated ginsenoside CK (OCMC-CK) graft; the molar ratio of O-carboxymethyl chitosan: carboxylated ginsenoside CK active ester solution was 12:1.

[0101] S5, dissolving 5.794 g of recrystallized N-isopropylacrylamide monomer under a nitrogen atmosphere, mixing with 1.4 mg of 1-ethyl-3-(3-dimethylaminopropyl)carbodiimide hydrochloride and 7.4 mg of potassium persulfate dissolved in 10 mL of pure water, stirring at 600 r / min at 60° C. and a nitrogen atmosphere, dialyzing, placing in a refrigerator overnight, and freeze-drying for 48 h to obtain amino-terminated poly-N-isopropylacrylamide;

[0102] S6. The O-carboxymethyl chitosan-ginsenoside CK aqueous solution was mixed with 40 mg of 1-(3-dimethylaminopropyl)-3-ethylcarbodiimide hydrochloride and 24 mg of N-hydroxysuccinimide, and the mixture was stirred for 1 h. Then, amino-terminated poly (N-isopropylacrylamide) was added, and the reaction was continued for 24 h. The mixture was dialyzed, and the MW of the dialysis membrane was 12000-14000 Da. The mixture was freeze-dried for 48 h to obtain O-carboxymethyl chitosan-ginsenoside CK-poly (N-isopropylacrylamide) (PNIPAM-OCMC-CK) grafted product. The ratio of O-carboxymethyl chitosan-ginsenoside CK to amino-terminated poly (N-isopropylacrylamide) was 100 mg:200 mg.

[0103] Example 5

[0104] The prodrug nano drug delivery system similar to the viral capsid particle is different from Example 1 only in the preparation process of the O-carboxymethyl chitosan-ginsenoside CK-poly N-isopropylacrylamide polymer, specifically:

[0105] S3, dissolve 100 mg of ginsenoside CK in 20 mL of a 1:1 mixture of pyridine and dichloromethane, add 48 g of succinic anhydride, stir at 600 r / min, rotary evaporate, add 30 mL of pure water to the precipitate, filter and vacuum dry to obtain white CK-COOH powder;

[0106] S4. CK-COOH powder was dissolved in methanol again, mixed with 1-(3-dimethylaminopropyl)-3-ethylcarbodiimide hydrochloride and N-hydroxysuccinimide, and stirred for 1 hour to obtain a carboxylated ginsenoside CK active ester solution; then, O-carboxymethyl chitosan was added, the reaction was continued for 24 hours, and dialyzed in excess methanol / pure water = volume ratio of 1:1 for 48 hours. The MW of the dialysis membrane was 12000-14000Da. The dialyzed solution was placed in a -20°C refrigerator overnight, and finally the dialyzate was freeze-dried for 48 hours to obtain O-carboxymethyl chitosan-carboxylated ginsenoside CK (OCMC-CK) graft; the molar ratio of O-carboxymethyl chitosan: carboxylated ginsenoside CK active ester solution was 8:1.

[0107] S5, dissolving 5.794 g of recrystallized N-isopropylacrylamide monomer under a nitrogen atmosphere, mixing with 1.4 mg of 1-ethyl-3-(3-dimethylaminopropyl)carbodiimide hydrochloride and 7.4 mg of potassium persulfate dissolved in 10 mL of pure water, stirring at 600 r / min at 60° C. and a nitrogen atmosphere, dialyzing, placing in a refrigerator overnight, and freeze-drying for 48 h to obtain amino-terminated poly-N-isopropylacrylamide;

[0108] S6. The O-carboxymethyl chitosan-ginsenoside CK aqueous solution was mixed with 40 mg of 1-(3-dimethylaminopropyl)-3-ethylcarbodiimide hydrochloride and 24 mg of N-hydroxysuccinimide, and the mixture was stirred for 1 h. Then, amino-terminated poly (N-isopropylacrylamide) was added, and the reaction was continued for 24 h. The mixture was dialyzed, and the MW of the dialysis membrane was 12000-14000 Da. The mixture was freeze-dried for 48 h to obtain O-carboxymethyl chitosan-ginsenoside CK-poly (N-isopropylacrylamide) (PNIPAM-OCMC-CK) grafted product. The ratio of O-carboxymethyl chitosan-ginsenoside CK to amino-terminated poly (N-isopropylacrylamide) was 250 mg:200 mg.

[0109] Example 6

[0110] The prodrug nano drug delivery system similar to the viral capsid particle is different from that of Example 1 only in step S7, which is specifically:

[0111] S7. The PNIPAM-OCMC-CK solution and IR820 dissolved in dimethyl sulfoxide were stirred, ultrasonicated for 6 hours in an ice bath at 120W, dialyzed for 30 minutes, and the MW of the dialysis membrane was 12000-14000Da. After freeze-drying, OCMC-CK-PNIPAM@IR820 nanoprodrug micelles were obtained; the concentration of the PNIPAM-OCMC-CK solution was 1 mg / mL, the concentration of the IR820 solution was 2 mg / mL, and the mixing ratio of the two solutions was 5 mL:0.2 mL.

[0112] Example 7

[0113] The prodrug nano drug delivery system similar to the viral capsid particle is different from that of Example 1 only in step S7, which is specifically:

[0114] S7. The PNIPAM-OCMC-CK solution and IR820 dissolved in dimethyl sulfoxide were stirred, ultrasonicated for 3 h in an ice bath at 120 W, dialyzed for 30 min, and the MW of the dialysis membrane was 12000-14000 Da. After freeze-drying, OCMC-CK-PNIPAM@IR820 nanoprodrug micelles were obtained; the concentration of the PNIPAM-OCMC-CK solution was 1 mg / mL, the concentration of the IR820 solution was 2 mg / mL, and the mixing ratio of the two solutions was 5 mL:1.6 mL.

[0115] Example 8

[0116] The prodrug nano drug delivery system similar to the viral capsid particle is different from that of Example 1 only in step S7, which is specifically:

[0117] S7. The PNIPAM-OCMC-CK solution and IR820 dissolved in dimethyl sulfoxide were stirred, ultrasonicated for 3 h in an ice bath at 120 W, dialyzed for 30 min, and the MW of the dialysis membrane was 12000-14000 Da. After freeze-drying, OCMC-CK-PNIPAM@IR820 nanoprodrug micelles were obtained; the concentration of the PNIPAM-OCMC-CK solution was 1 mg / mL, the concentration of the IR820 solution was 2 mg / mL, and the mixing ratio of the two solutions was 5 mL:0.8 mL.

[0118] Example 9

[0119] The viral capsid-like prodrug nano-drug delivery system is different from Example 1 only in that the photosensitizer is pyropheophytin-a (Ppa).

[0120] Example 10

[0121] The viral capsid-like prodrug nano-delivery system is different from Example 1 only in that the photosensitizer is mono-asparagine chlorin-e6 (NPe6).

[0122] Since Examples 1 to 10 all prepared prodrug nano-delivery systems with the expected effects of the present invention, and the performance and effects of the prodrug nano-delivery systems prepared in each example are basically the same, the present invention hereinafter only takes the prodrug nano-delivery system prepared in Example 1 as an example to illustrate its performance and anti-tumor effect.

[0123] The FA-Cys synthesized in Example 1 was characterized by Fourier transform infrared absorption spectrometry. Figure 1 As shown in FA, 1335cm -1 The band at 1607 cm corresponds to the asymmetric stretching vibration of -NH2. -1 and 1696cm -1 The position corresponds to the C=O stretching in carboxylic acid. As shown in FA-Cys, the -1 The bands between are due to the stretching vibration bands of FA and Cys and the NH stretching vibration. Therefore, these results can indicate the successful grafting of FA and Cys.

[0124] The FA-Cys synthesized in Example 1 was characterized by hydrogen nuclear magnetic resonance spectroscopy. Figure 2 There are NH chemical shifts at 8.01 ppm and SH chemical shifts at 2.1 ppm in FA-Cys, indicating the successful grafting of FA onto Cys.

[0125] The microscopic morphology of the prodrug nano-delivery system with a virus-like capsid structure synthesized in Example 1 was characterized by transmission electron microscopy. Figure 3 As shown in the figure, the micelles in water medium are nearly spherical in shape, with smooth surface, uniform size and particle size of about 200 nm.

[0126] The near-infrared fluorescence imaging distribution results of the prodrug nano-delivery system with a virus-like capsid structure synthesized in Example 1 are shown in the attached figure. Figure 4 The Zeta potential results are shown in the attached Figure 5As shown. The average particle size of the prepared FA-Cys / OCMC-CK-PNIPAM@IR820 micelles is 239.3±3.4nm, with uniform distribution and good dispersion; the Zeta potential of FA-Cys / OCMC-CK-PNIPAM@IR820 micelles is -17.2±0.31mv. This shows that the self-assembled nanomicelles have good stability. And compared with the OCMC-CK-PNIPAM@IR820 micelles, the potential is reduced, which is caused by the positive charge carried by FA-Cys, indicating that FA-Cys / OCMC-CK-PNIPAM@IR820 is successfully synthesized.

[0127] Example 2

[0128] This example shows the generation of singlet reactive oxygen species by the prodrug nano-delivery system FA-Cys / OCMC-CK-PNIPAM@IR820 with a virus-like capsid structure prepared in Example 1 under near-infrared light irradiation: 2 mL of the prodrug nano-delivery system aqueous solution (50 μg / mL) was mixed with 2 mL of 1,3-diphenylisobenzofuran (DPBF) DMF solution (0.1 mM) in the dark. After thorough mixing, the mixed solution was continuously irradiated with a near-infrared laser for 5 minutes, and the UV-visible absorption spectrum of the solution was measured every 1 minute to measure the changes in the UV-visible absorption spectrum of the solution.

[0129] The singlet reactive oxygen species generation of the FA-Cys / OCMC-CK-PNIPAM@IR820 nano drug delivery system synthesized in Example 1 is shown in the attached figure. Figure 6 As shown in Figure 2, under the irradiation of near-infrared light, the UV-visible absorption peak of DPBF has no obvious change in the absence of FA-Cys adsorption, indicating that the adsorption of FA-Cys does not affect the generation of the prodrug nano-delivery system. 1 O2 capacity, the drug delivery system still has good photothermal response ability.

[0130] Example 3

[0131] This example is a toxicity test of the FA-Cys / OCMC-CK-PNIPAM@IR820 nano drug delivery system prepared in Example 1 on liver cancer cells HepG-2, which is divided into a chemotherapy group alone and a photodynamic therapy and chemotherapy combined treatment group.

[0132] The chemotherapy group alone was: according to the density of 1.0×10 5HepG-2 cells were seeded in a 96-well plate / well and cultured at 37°C and 5% carbon dioxide for 24 hours. After the culture was completed, the culture medium in the well was aspirated, and 100 μL of ginsenoside CK and OCMC-CK@IR820 nanomicelles were added and cultured for 24 hours. After the culture was completed, the sample solution in the well was aspirated, MTT solution (10 μL) was added to each well, and then 90 μL of culture medium was added and cultured for 4 hours. After the culture was completed, the MTT solution in the well was aspirated, and 110 μL of Formazan dissolving solution was added to dissolve the crystals. The absorbance of each sample was recorded at a wavelength of 490 nm using a BIO-DL-K3 ​​Plus microplate reader. The cytotoxicity calculation formula is as follows:

[0133]

[0134] Among them, A experimental group represents the absorbance of the experimental group (including cells, MTT, Formazan solution and sample solution), A blank group represents the absorbance of the blank group (including MEM culture medium, MTT solution and Formazan solution), and A control group represents the absorbance of the control group (including cells, MTT solution and Formazan solution).

[0135] Combined treatment group: After adding 100 μL of IR820 and OCMC-CK@IR820 nanomicelles of different concentrations into the culture medium, the cells were first illuminated with 808 nm, 1 W / cm 2 The cells were irradiated with near-infrared laser for 3 min and then cultured for 24 h. The rest of the steps were the same as those in the chemotherapy alone group.

[0136] Example 1 The toxicity of the synthesized nano drug delivery system to HepG2 cells is shown in the attached figure. Figure 7 As shown. As the concentration of the encapsulated ginsenoside CK increased from 3.125μg / mL to 50.000μg / mL, the inhibitory effect of FA-Cys / PNIPAM-OCMC-CK@IR820 on liver cancer cells HepG2 continued to increase. When irradiated with near-infrared rays, the inhibitory effect of FA-Cys / PNIPAM-OCMC-CK@IR820 on liver cancer cells was enhanced, reaching 38.28% at 12.5mg / mL and 7.31% at 25mg / mL. This shows that the prodrug nanodelivery system prepared by the invention has the effect of enhancing the combined treatment effect of photodynamic therapy and ginsenoside CK chemotherapy.

[0137] Example 4

[0138] This example shows the uptake of the FA-Cys / PNIPAM-OCMC-CK@IR820 prodrug nano-drug delivery system prepared in Example 1 by liver cancer cells. The FA-Cys / PNIPAM-OCMC-CK@IR820 prodrug nano-micelles were co-incubated with HepG-2 cells, and the in vitro cell uptake of human liver cancer cells HepG-2 was detected using an inverted confocal laser scanning microscope. Fluorescence imaging was performed as shown in FIG. Figure 8 shown.

[0139] Depend on Figure 8 It can be seen that after FA-Cys / PNIPAM-OCMC-CK@IR820 prodrug nanomicelles acted on HepG-2 cells, the cells in the merged fluorescence imaging showed blue and red fluorescence at the same time, and as the incubation time increased, the red fluorescence intensity in the cell nucleus increased. In addition, the brightness of FA-Cys / PNIPAM-OCMC-CK@IR820 prodrug nanomicelles was enhanced compared with PNIPAM-OCMC-CK@IR820. This shows that FA-Cys / PNIPAM-OCMC-CK@IR820 prodrug nanomicelles with folic acid-cysteamine conjugates are more easily taken up by cells than PNIPAM-OCMC-CK@IR820, indicating that FA-grafted prodrug nanomicelles have a stronger ability to target liver cancer cells and are more conducive to the uptake of IR820 by cells.

[0140] Example 5

[0141] This example shows the cell apoptosis of HepG-2 cells after the prodrug nano drug delivery system prepared in Example 1 was used to treat the cells.

[0142] HepG-2 cells were cultured at 1×10 5 Cells were inoculated per well in a 6-well plate. After culturing for 24 h, 1 mL of ginsenoside CK of different concentrations and micelle solution of the corresponding concentration were added. After treatment for 24 h, the cells were digested with 0.25% EDTA trypsin, and the cells were collected by centrifugation at 1000 rpm for 4 min. The cells were washed three times with pre-cooled PBS, and then 100 μL 1×Binding Buffer was added to resuspend the cells. Finally, 5 μL Annexin V-FITC and 5 μL PI were added to each tube, and 400 μL 1×Binding Buffer was added to each tube after incubation in the dark for 15 min, and the cell apoptosis rate was detected by C6 Plus flow cytometer.

[0143] like Fig. 9As shown, micelles of different concentrations can induce apoptosis of HepG-2 cells, and the apoptosis rate is significantly positively correlated with the treatment concentration. Under the treatment of FA-Cys / PNIPAM-OCMC-CK@IR820 at a concentration of 50 μg / mL, the percentage of apoptotic cells (the sum of early apoptotic cells and late apoptotic cells) was 59.78±2.12%.

[0144] The above results are similar to those of cytotoxicity experiments, which show that FA-Cys / PNIPAM-OCMC-CK@IR820 can effectively inhibit the growth of liver cancer cells and has great potential for the treatment of liver cancer.

[0145] The present invention utilizes the characteristics that folic acid (FA) can bind to the overexpressed folic acid receptor on cancer cells, PNIPAM-OCMC-CK has thermosensitivity, and phase change occurs at 38°C, which is conducive to the release of active controlled release prodrug micelles. The amino groups of cross-linked carboxymethyl chitosan and the carboxyl groups of carboxylated ginsenoside CK and the carboxyl groups of carboxymethyl chitosan and the amino groups of terminal amino groups of poly N-isopropyl acrylamide form amide bond-connected polymers, which are self-assembled into spherical micelles to encapsulate IR820 through ultrasound. The FA-Cys conjugate is adsorbed on the surface of the PNIPAM-OCMC-CK@IR820 prodrug nano-micelles under electrostatic action and cross-linked to form disulfide bonds. Through folic acid receptor targeted drug delivery, when the drug-loaded nanoparticles reach the tumor site, the disulfide bonds consume the glutathione in the liver cancer cells and break; when the photosensitizer IR820 is irradiated with near-infrared light, the photothermal conversion causes the PNIPAM phase change to actively release ginsenoside CK, and the release of the controlled release prodrug micelles is precisely treated. The present invention enhances the effect of combined chemotherapy and photodynamic therapy, and provides a certain reference for improving the bioavailability of ginsenoside CK and promoting photodynamic therapy.

[0146] It should be noted that the prodrug nano drug delivery system provided by the present invention can not only load ginsenoside CK, but also other anti-tumor drugs containing hydroxyl groups can be loaded after carboxylation, and better anti-tumor effects can be obtained according to the concept of the present invention. Exemplary, the anti-tumor drugs containing hydroxyl groups that can be selected are ginsenoside Re, ginsenoside Rg3, ginsenoside Rg5, ginsenoside Rh2, ginsenoside Rh3, ginsenoside Rh1 or ginsenoside Rc. The type of anti-tumor that the final prodrug nano drug delivery system prepared according to the scheme of the present invention can be different depending on the type of hydroxyl-containing drug loaded.

[0147] Obviously, it is only a preferred embodiment of the present invention and does not limit the present invention in any way. Any simple modification, change and equivalent change made to the above embodiment according to the technical essence of the invention still falls within the protection scope of the technical solution of the present invention.

Claims

1. A prodrug nano drug delivery system, characterized in that: It is prepared according to the following steps: The folic acid and triethylamine are mixed for amidation reaction to obtain folic acid N-hydroxysuccinimide active ester; the folic acid N-hydroxysuccinimide active ester and cysteamine are used as raw materials for amidation reaction to obtain a cysteamine-folic acid conjugate; The carboxylated ginsenoside drug is mixed with carboxymethyl chitosan for amidation reaction to obtain a carboxymethyl chitosan-carboxylated ginsenoside graft; the ginsenoside drug is any one of ginsenoside CK, ginsenoside Re, ginsenoside Rg3, ginsenoside Rg5, ginsenoside Rh2, ginsenoside Rh3, ginsenoside Rh1 or ginsenoside Rc; polymerizing N-isopropylacrylamide monomers to obtain amino-terminated poly-N-isopropylacrylamide; Using the amino-terminated poly N-isopropylacrylamide and the carboxymethyl chitosan-carboxylated ginsenoside graft as raw materials, an amidation reaction is carried out to obtain a carboxymethyl chitosan-ginsenoside-poly N-isopropylacrylamide graft; The amino group of carboxymethyl chitosan forms an amide bond with the carboxyl group of carboxylated ginsenoside drugs, and the carboxyl group of carboxymethyl chitosan forms an amide bond with the amino group of the amino-terminated poly N-isopropyl acrylamide, thereby obtaining the carboxymethyl chitosan-ginsenoside-poly N-isopropyl acrylamide graft; The solution of carboxymethyl chitosan-ginsenoside-poly (N-isopropylacrylamide) grafted product is ultrasonically treated to self-assemble into a capsid structure, and a photosensitizer is added to encapsulate the photosensitizer in the formed capsid structure to obtain nano-prodrug micelles; The nano prodrug micelles are mixed with the cysteamine-folic acid conjugate for electrostatic adsorption to obtain a prodrug nano drug delivery system with a virus capsid particle structure.

2. The prodrug nano drug delivery system according to claim 1, characterized in that: The molar ratio of the carboxymethyl chitosan to the carboxylated ginsenoside drugs is 6-14:

1.

3. The prodrug nano drug delivery system according to claim 2, characterized in that: The mass ratio of the carboxymethyl chitosan-carboxylated ginsenoside grafted product to the amino-terminated poly N-isopropylacrylamide is 1-2.5:

2.

4. The prodrug nano drug delivery system according to claim 3, characterized in that: The photosensitizer is any one of new indocyanine green, pyropheophytin-a or mono-asparagine chlorin-e6.

5. The prodrug nano drug delivery system according to claim 4, characterized in that: The concentration of the carboxymethyl chitosan-ginsenoside-poly N-isopropylacrylamide graft solution is 1 mg / mL, the concentration of the photosensitizer added is 2 mg / mL, and the mixed volume ratio of the carboxymethyl chitosan-ginsenoside-poly N-isopropylacrylamide graft solution to the photosensitizer solution is 5:0.2-1.6; The mixing mass ratio of the nano prodrug micelle to the cysteamine-folic acid conjugate is 2:0.

5.

6. Use of the prodrug nano drug delivery system according to any one of claims 1 to 5 in the preparation of photodynamic therapy drugs.

7. The use according to claim 6, characterized in that: The prodrug nano drug delivery system is used for preparing anti-tumor drugs.

8. The use according to claim 7, characterized in that: The prodrug nano drug delivery system is used for preparing anti-liver cancer drugs.

9. An anti-tumor drug, characterized in that: The invention comprises the prodrug nano drug delivery system according to any one of claims 1 to 5, and pharmaceutically acceptable excipients.

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