Oxaliplatin multifunctional liposome as well as preparation method and application thereof

Abstract

The invention discloses a multifunctional oxaliplatin liposome as well as a preparation method and application thereof. The liposome is prepared from oxaliplatin, calcium peroxide nanoparticles, lecithin and ginsenoside Rg3. According to the oxaliplatin and calcium peroxide nano-particle co-loaded multifunctional liposome disclosed by the invention, the peroxide nano-particles improve a solid tumor hypoxia microenvironment and improve the anti-tumor curative effect of oxaliplatin; ginsenoside Rg3 replaces cholesterol and plays a targeting role, and meanwhile, ginsenoside Rg3 is used as a good chemotherapy adjuvant and is combined with a chemotherapy drug to improve an anti-tumor effect and reduce systemic toxicity; the IC50 value of the liposome to HCT116 cells is 5.37 + / -0.42 mu M, the tumor inhibition rate is 87.55 + / -9.35%, and the liposome has a good anti-colorectal cancer effect.

Description

Technical Field

[0001] The invention relates to a co-loaded multifunctional liposome, a preparation method and an application thereof, and in particular to a co-loaded multifunctional liposome of oxaliplatin and calcium peroxide nanoparticles, a preparation method and an application thereof. Background Art

[0003] Colorectal cancer (CRC) is a heterogeneous disease across the anatomical location of the tumor and is a multifactorial disease caused by lifestyle, genetic and environmental factors. Among them, genetic and environmental factors play an important role in the etiology of colorectal cancer. Generally, the subtypes of colorectal cancer are defined according to the anatomical location of the tumor in the three segments of the colorectum, and are divided into proximal colon cancer (cecum, ascending colon, hepatic flexure and transverse colon), distal colon cancer (splenic flexure, descending colon and sigmoid colon) and rectal cancer.

[0004] Platinum drugs are the main chemotherapy drugs for clinical anti-tumor treatment. Oxaliplatin (OXA) is a third-generation platinum anti-tumor drug developed by Debiopharm in Switzerland. It has broad-spectrum anti-tumor activity both in vivo and in vitro, but its therapeutic effect is inferior to that of cisplatin and carboplatin, and it has adverse reactions such as neurotoxicity and bone marrow suppression. In addition, oxaliplatin is a first-line chemotherapy drug for the treatment of colorectal cancer, which kills tumor cells by interfering with DNA replication and transcription, causing DNA damage. However, the clinical application of OXA is limited due to the unique microenvironment of solid tumors, such as hypoxia, acidosis, and high levels of glutathione. Recently, various metal peroxides, including calcium peroxide (CaO2), manganese dioxide (MnO2), magnesium peroxide (MgO2), and zinc peroxide (ZnO2), have been used to improve the hypoxic tumor microenvironment by reacting with H2O in tumor tissue to produce oxygen, thereby improving the chemotherapy effect. Among these metal peroxides, CaO2 has high biocompatibility and efficient oxygen production capacity, and is the most promising material for regulating tumor hypoxia.

[0005] However, since CaO2 is unstable in water and OXA is systemically toxic when administered directly, a drug carrier system needs to be designed for the delivery of CaO2 and OXA. Liposomes are a drug carrier with high biocompatibility, low toxicity and simple preparation. Cholesterol is the basic component of liposomes and has the ability to stabilize the phospholipid bilayer, but cholesterol may cause hyperlipidemia and allergic reactions. Therefore, it is of great significance to find structurally similar cholesterol substitutes to stabilize lipid membranes. Ginsenoside Rg3 is a steroid compound isolated from ginseng, which has a lipophilic steroid structure similar to cholesterol and can be used to replace cholesterol. In addition, Rg3 also has the functions of actively targeting glucose transporter 1 (Glut1) overexpressed on the surface of colorectal cancer cells and assisting chemotherapy, and can replace cholesterol as a membrane stabilizer to prepare multifunctional liposomes. Summary of the invention

[0006] The purpose of the present invention is to provide a multifunctional liposome co-loaded with oxaliplatin and calcium peroxide nanoparticles, a preparation method and an application thereof, which can target the glucose transporter 1 on the surface of HCT116 cells, increase the targeting and uptake of the liposome, and at the same time regulate the hypoxic microenvironment of the tumor, enhance the killing effect of chemotherapy drugs on colorectal cancer cells, have a good anti-colorectal cancer effect, and significantly reduce the tumor volume.

[0007] In order to achieve the above-mentioned object, the present invention provides a multifunctional liposome co-loaded with oxaliplatin and calcium peroxide nanoparticles. The liposome is composed of oxaliplatin, CaO2, lecithin, and ginsenoside Rg3, wherein the molar ratio of lecithin to ginsenoside Rg3 is 3-5:1; the molar ratio of lecithin to oxaliplatin is 15-20:1, the final concentration of CaO2 is 1-2 mg / mL, and the final concentration of OXA is 1-2 mg / mL.

[0008] The present invention provides a method for preparing multifunctional liposomes co-loaded with oxaliplatin and calcium peroxide nanoparticles, the method comprising the following steps:

[0009] Step 1, dissolving lecithin, ginsenoside Rg3 and CaO2 in a mixed solution of chloroform and anhydrous ethanol and performing rotary evaporation at 30-50° C. for 1-3 hours to form a transparent lipid film;

[0010] Step 2: Add oxaliplatin to the glucose aqueous solution, place on a shaker at 30-50° C. for hydration for 1-4 hours, then ultrasonicate at 100 W for 10-20 minutes, and squeeze the treated liposomes through a membrane with a pore size of 450 nm several times and through a membrane with a pore size of 220 nm several times;

[0011] The multifunctional liposomes co-loaded with oxaliplatin and calcium peroxide nanoparticles and their application of the present invention have the following advantages:

[0012] The multifunctional liposomes co-loaded with oxaliplatin and calcium peroxide nanoparticles provided by the present invention and the preparation method and application thereof combine chemotherapeutic drugs with peroxide nanoparticles, improve the hypoxic microenvironment of solid tumors, and improve the anti-tumor efficacy of oxaliplatin. The calcium peroxide nanoparticles provided by the present invention have a particle size of less than 25.0 nm and a PDI value of less than 0.2, indicating that the calcium peroxide nanoparticles provided by the present invention are uniformly distributed and do not aggregate.

[0013] The multifunctional liposomes co-loaded with oxaliplatin and calcium peroxide nanoparticles provided by the present invention and the preparation method and application thereof optimize the common liposomes, introduce ginsenoside Rg3 to replace cholesterol to reduce the liposome particle size, and improve the targeting of the liposomes. At the same time, ginsenoside Rg3 is a good chemotherapy adjuvant, which can increase the anti-tumor effect and reduce the systemic toxicity of chemotherapeutic drugs when used in combination with chemotherapeutic drugs. The particle sizes of common cholesterol liposomes (Lip-OXA / CaO2) and ginsenoside Rg3 liposomes (Rg3-Lip-OXA / CaO2) were compared, and the results showed that the particle size of Lip-OXA / CaO2 was 121.54±5.63nm, and the particle size of Rg3-Lip-OXA / CaO2 was 104.78±3.96nm. In addition, the particle sizes of Lip-OXA / CaO2 and Rg3-Lip-OXA / CaO2 were both less than 0.2, indicating that the particle size of the liposomes provided by the present invention was uniform and no coagulation occurred.

[0014] The multifunctional liposomes (Rg3-Lip-OXA / CaO2) co-loaded with oxaliplatin and calcium peroxide nanoparticles provided by the present invention are significantly different from free drugs (OXA), ordinary liposomes loaded with oxaliplatin (Lip-OXA) and ordinary liposomes (Lip-OXA / CaO2) co-loaded with oxaliplatin and calcium peroxide nanoparticles. The Rg3-Lip-OXA / CaO2 of the present invention has stronger cytotoxicity to HCT116 cells. Moreover, the Rg3-Lip-OXA / CaO2 of the present invention has the best inhibition rate on HCT116 cell tumors, the tumor volume is significantly reduced, and the tumor inhibition rate reaches more than 80%. In terms of toxicity, the weight of mice increased when using the Rg3-Lip-OXA / CaO2 of the present invention, and the coefficients of the main organs did not change significantly, indicating that the Rg3-Lip-OXA / CaO2 of the present invention has low toxicity.

[0015] Therefore, the Rg3-Lip-OXA / CaO2 of the present invention has a good tumor inhibition effect in vivo and has relatively low toxicity. The preparation provided by the present invention has important theoretical value and good application prospects. BRIEF DESCRIPTION OF THE DRAWINGS

[0016] Figure 1 Transmission electron microscope image of Example 1 of the present invention

[0017] Figure 2 The oxygen release curve of Example 1 in water of the present invention is

[0018] Figure 3 Transmission electron microscope image of liposomes of Example 2 of the present invention

[0019] Figure 4 The particle size and PDI value changes of Example 2 of the present invention in different media.

[0020] Figure 5 The leakage rates of drugs of the liposomes of Example 2 of the present invention and Comparative Examples 1-3 at different temperatures.

[0021] Figure 6 The release curves of the drugs in the liposomes of Example 2 of the present invention and Comparative Examples 1-3 in PBS are shown.

[0022] Figure 7 These are fluorescent photos of intracellular uptake after HCT116 cells were incubated with different liposomes for a certain period of time.

[0023] Figure 8 The cell survival rate curves of the liposomes of Example 2 of the present invention and Comparative Examples 1-3 are shown.

[0024] Fig. 9 The curves showing the changes in tumor volume and tumor inhibition rate of tumor-bearing nude mice after treatment with the liposomes of Example 2 of the present invention and Comparative Examples 1-3 are shown.

[0025] Fig.10 The figure shows the curve of body weight change of tumor-bearing nude mice after liposome treatment of Example 2 of the present invention and Comparative Examples 1-3. DETAILED DESCRIPTION

[0026] The technical solutions in the embodiments of the present invention are described clearly and completely below. Obviously, the described embodiments are only part of the embodiments of the present invention, not all of them. Based on the embodiments of the present invention, all other embodiments obtained by ordinary technicians in this field without creative work are within the scope of protection of the present invention.

[0027] Example 1

[0028] A method for preparing calcium peroxide nanoparticles, comprising:

[0029] 2 mL of 0.2 g / mL CaCl2 aqueous solution and 2 mL of 1 mol / L ammonia aqueous solution were added to 80 mL PEG200, stirred evenly, and then 2 mL of 30% hydrogen peroxide solution was slowly added to the mixed solution, and the mixture was stirred for 6 hours at room temperature. The pH value of the solution was adjusted to 11.5 with 1 mol / L NaOH solution, and the precipitate was collected by centrifugation at 9500 g for 10 minutes, and washed twice with 0.1 mol / L sodium hydroxide solution, water and anhydrous ethanol respectively to obtain white CaO2 nanoparticles, which were stored in anhydrous ethanol.

[0030] Example 2

[0031] A method for preparing multifunctional liposomes co-loaded with oxaliplatin and calcium peroxide nanoparticles, comprising:

[0032] 36 mg PC (egg yolk lecithin), 9 mg Rg3 (ginsenoside Rg3) and 1 mg CaO2 (calcium peroxide nanoparticles) were weighed and dissolved in a mixed solvent of 5 mL chloroform and 5 mL anhydrous ethanol.

[0033] The mixed solution was ultrasonically mixed for 5 to 10 minutes to fully dissolve PC and Rg3 in the organic solvent, and then the solvent was slowly vacuumed and evaporated at a speed of 100 to 120 rpm. The water bath temperature was 37°C, and a uniform transparent film was formed after 2 hours. Then, 1 mL of glucose solution containing oxaliplatin (the concentration of oxaliplatin was 1 mg / mL) was added and hydrated at 37°C for 2 hours at a speed of 100 rpm. Then, the probe was ultrasonicated for 3 seconds and cooled for 3 seconds in an ice water bath for a total of 10 minutes, and extruded three times through a polycarbonate 80 nm membrane through a liposome extruder to obtain Rg3-Lip-OXA / CaO2.

[0034] Comparative Example 1

[0035] A method for preparing a single oxaliplatin-loaded common liposome Lip-OXA, comprising:

[0036] Weigh 36 mg PC (egg yolk phosphatidylcholine) and 4.5 mg Chol (cholesterol) and dissolve them in a mixed solvent of 5 mL chloroform and 5 mL anhydrous ethanol.

[0037] The mixed solution was ultrasonically mixed for 5-10 minutes to fully dissolve PC and Chol in the organic solvent, and then the solvent was slowly vacuumed and rotary evaporated at a speed of 100-120 rpm, and the water bath temperature was 37°C. After 2 hours, a uniform and transparent film was formed. Then, 1 mL of glucose solution containing oxaliplatin (the concentration of oxaliplatin was 1 mg / mL) was added, and hydrated at 37°C for 2 hours at a speed of 100 rpm. Then, the probe was ultrasonicated for 3 seconds and cooled for 3 seconds in an ice water bath for a total of 10 minutes, and extruded three times through a polycarbonate 80 nm membrane through a liposome extruder to obtain Lip-OXA.

[0038] Comparative Example 2

[0039] A method for preparing single oxaliplatin-loaded multifunctional liposome Rg3-Lip-OXA, comprising:

[0040] Weigh 36 mg PC (egg yolk lecithin) and 9 mg Rg3 (ginsenoside Rg3) and dissolve them in a mixed solvent of 5 mL chloroform and 5 mL anhydrous ethanol.

[0041] The mixed solution was ultrasonically mixed for 5 to 10 minutes to fully dissolve PC and Rg3 in the organic solvent, and then the solvent was slowly vacuumed and rotary evaporated at a speed of 100 to 120 rpm. The water bath temperature was 37°C, and a uniform transparent film was formed after 2 hours. Then, 1 mL of glucose solution containing oxaliplatin (the concentration of oxaliplatin was 1 mg / mL) was added and hydrated at 37°C for 2 hours at a speed of 100 rpm. Then, the probe was ultrasonicated for 3 seconds and cooled for 3 seconds in an ice water bath for a total of 10 minutes, and extruded three times through a polycarbonate 80 nm membrane through a liposome extruder to obtain Rg3-Lip-OXA.

[0042] Comparative Example 3

[0043] A method for preparing common liposomes Lip-OXA / CaO2 co-loaded with oxaliplatin and calcium peroxide nanoparticles, comprising:

[0044] 36 mg PC (egg yolk lecithin), 4.5 mg Chol (cholesterol) and 1 mg CaO2 (calcium peroxide nanoparticles) were weighed and dissolved in a mixed solvent of 5 mL chloroform and 5 mL anhydrous ethanol.

[0045] The mixed solution was ultrasonically mixed for 5-10 minutes to fully dissolve PC and Chol in the organic solvent, and then the solvent was slowly vacuumed and rotary evaporated at a speed of 100-120 rpm, and the water bath temperature was 37°C. After 2 hours, a uniform and transparent film was formed. Then, 1 mL of glucose solution containing oxaliplatin (the concentration of oxaliplatin was 1 mg / mL) was added, and hydrated at 37°C for 2 hours at a speed of 100 rpm. Then, the probe was ultrasonicated for 3 seconds and cooled for 3 seconds in an ice water bath for a total of 10 minutes, and extruded three times through a polycarbonate 80 nm membrane through a liposome extruder to obtain Lip-OXA / CaO2.

[0046] The specific examples and comparative examples of the present invention are as follows:

[0047] 1. The settings of the embodiments and comparative examples are as follows:

[0048] Table 1 Abbreviations of Examples and Comparative Examples of the present invention

[0049]

[0050] 2. Morphological observation of calcium peroxide nanoparticles

[0051] The dried Example 1 was evenly dispersed in a methanol solution, and an appropriate amount of the sample was dropped onto a copper mesh. After drying, it was stained with phosphotungstic acid for half a minute, and the morphology of the calcium peroxide particles was observed using a transmission electron microscope. Figure 1 As shown, the calcium peroxide nanoparticles are uniformly spherical and evenly dispersed without aggregation.

[0052] 3. Determination of particle size and PDI of calcium peroxide nanoparticles

[0053] The diluted Example 1 was taken and the particle size and PDI value (Polydispersity Index) were measured using a Malvern particle size analyzer at 25° C. The results are shown in Table 2.

[0054] Table 2 Particle size and PDI of Example 1 of the present invention (n=3)

[0055]

[0056] The particle size of the calcium peroxide nanoparticles is 23.23±1.84 nm, which is less than 25 nm, and the PDI is 0.164±0.017, indicating that the particle size distribution of the calcium peroxide nanoparticles is narrow.

[0057] 4. In vitro oxygen release of calcium peroxide nanoparticles

[0058] Take an appropriate amount of Example 1 and add it to PBS solutions of different pH values. Use a dissolved oxygen meter to measure the ability of Example 1 to release oxygen in water. The results are as follows: Figure 2 As shown in Figure 3, calcium peroxide nanoparticles release oxygen more quickly and completely in PBS with a pH of 5.5, indicating that the prepared calcium peroxide nanoparticles can stably and quickly release oxygen in an acidic environment.

[0059] 5. Determination of liposome encapsulation efficiency and drug loading

[0060] The liposomes of Example 2 and Comparative Examples 1-3 were demulsified, and the encapsulation efficiency (EE%) and drug loading (DL%) were determined by HPLC. The results are shown in Table 3.

[0061] Table 3 Encapsulation efficiency and drug loading of liposomes of Comparative Examples 1-3 and Example 2 of the present invention (n=3)

[0062]

[0063] In the co-loaded liposomes, the addition of calcium peroxide nanoparticles does not affect the encapsulation efficiency of oxaliplatin. Rg3 as a membrane material has a relatively high encapsulation efficiency and drug loading.

[0064] 6. Morphological observation of Rg3-Lip-OXA / CaO2 liposomes

[0065] The freshly prepared Example 2 was diluted 100 times, and an appropriate amount of the sample was dripped onto a copper mesh. After drying, it was stained with phosphotungstic acid for half a minute, and the morphology of the liposomes was observed using a transmission electron microscope. The results are shown in FIG. Figure 3 As shown, the liposomes are uniformly spherical, with a particle size of about 100-200 nm, and are evenly dispersed without aggregation.

[0066] 7. Determination of liposome particle size, PDI and Zeta potential

[0067] The diluted liposome solutions of Example 2 and Comparative Examples 1-3 were taken and the particle size, potential and PDI value (Polydispersity Index) were measured using a Malvern particle size analyzer at 25° C. The results are shown in Table 4.

[0068] Table 4 Particle size, PDI and Zeta potential of liposomes of Comparative Examples 1-3 and Example 2 of the present invention (n=3)

[0069]

[0070] As can be seen from Table 4, the particle size of the multifunctional liposomes with Rg3 added is smaller than that of ordinary liposomes. The values ​​of the dispersion coefficient PDI of each liposome are all less than 0.2, indicating that the liposomes are uniformly distributed. The zeta potential values ​​of the liposomes are all negative, indicating that the liposomes are relatively stable and do not aggregate.

[0071] 8. Stability of Rg3-Lip-OXA / CaO2 liposomes

[0072] The liposomes of Example 2 were placed in three release media to measure the changes in particle size and PDI value within 48 hours. The three media were PBS solution, serum, and complete cell culture medium (10% FBS + 90% culture medium), wherein PBS solution and serum were the most commonly used release media in in vitro release experiments, and the complete cell culture medium was used to simulate the growth environment of human cells. In addition, the liposomes of Example 2 and Comparative Examples 1-3 were placed at 4°C and room temperature, respectively, and the drug concentration encapsulated in the liposomes was measured at different time points (0h, 1h, 2h, 4h, 8h, 24h, 48h), recorded as Cn (n is a different time point), substituted into the formula to obtain the leakage percentage, and the time-leakage percentage curve was drawn. The calculation method of the leakage rate is: leakage rate = (1-Cn / C0) × 100%.

[0073] like Figure 4 As shown, A is the change in particle size of Rg3-Lip-OXA / CaO2 in Example 2 in different media, and B is the change in PDI value of Rg3-Lip-OXA / CaO2 in Example 2 in different media. It can be seen that the particle size and PDI value both increase slightly with time, but are still within the appropriate range, and the PDI value is still less than 0.25, indicating that the liposome has good dispersibility.

[0074] like Figure 5As shown, the leakage rates of OXA of the liposomes of Example 2 of the present invention and Comparative Examples 1-3 at different temperatures (A: 4°C, B: room temperature) show that the leakage rate is low at lower temperatures, indicating that the liposomes are more stable and have a lower leakage rate at 4°C.

[0075] 9. In vitro release rate study

[0076] The liposomes of Example 2 and Comparative Examples 1-3 were placed in a dialysis bag for dialysis at a dialysis temperature of 37°C and a dialysis medium of PBS. Appropriate amounts were taken out at 0, 1, 2, 4, 8, 24, and 48 hours, and the OXA concentration was measured, recorded as Cn (n is a different time point). The release rate curve was drawn with time as the abscissa and the release rate as the ordinate. The release rate was calculated as follows: release rate = (1-Cn / C0) × 100%.

[0077] like Figure 6 As shown, the release curves of OXA in PBS from the liposomes of Example 2 of the present invention and Comparative Examples 1-3 show that all liposomes exhibit slow release behavior, among which the slow release behavior of Rg3-Lip-OXA / CaO2 is the most obvious.

[0078] 10. In vitro targeting studies

[0079] In order to observe the uptake of different liposome preparations by HCT116 cells, two liposome preparations loaded with the fluorescent dye rhodamine B (Rh B) were co-incubated with cells in an in vitro targeting experiment. Then, the cells were nuclear stained with Hoechst 33342 dye, and the changes in the uptake of liposomes by cells were observed under a fluorescence microscope. Hoechst 33342 showed blue fluorescence under a fluorescence microscope, and rhodamine B showed red fluorescence under a fluorescence microscope. Figure 7 The figure shows the cell fluorescence images of free Rh B, ordinary liposomes loaded with Rh B (Lip-Rh B) and multifunctional liposomes loaded with Rh B (Rg3-Lip-Rh B). It can be seen that the fluorescence intensity of free Rh B in cells is the weakest, while the fluorescence intensity of Lip-Rh B and Rg3-Lip-Rh B in cells is stronger than that of free Rh B. Among them, Rg3-Lip-Rh B has the strongest fluorescence intensity in cells, which is due to the targeting effect of Rg3 on glucose transporters on the surface of HCT116 cells. This result shows that the Rg3 targeting fragment can specifically recognize and bind to glucose transporters on the surface of HCT116 cells, enter cells through receptor-mediated endocytosis, and increase the uptake of liposomes.

[0080] 11. In vitro cytotoxicity

[0081] HCT116 cells in the logarithmic growth phase (cultured in McCoy's 5A medium containing 10% fetal bovine serum and 1% streptomycin and penicillin mixed solution) were placed in a 25 cm 2 After digestion, the cells were centrifuged at 1000 rpm for 5 min, and the culture medium was resuspended. 100 μL of the diluted cell suspension (2×10 3 10 μL of CCK-8 detection solution was added to each well, and then the cells were incubated for 1.5 to 2 hours and then detected at a wavelength of 450 nm using an ELISA reader. The relative cell survival rate was calculated, and the IC 50 .

[0082] like Figure 8 As shown, the cell survival rate curves of the liposomes of Example 2 of the present invention and Comparative Examples 1-3 show the in vitro cytotoxicity of different liposomes to HCT116, among which Rg3-Lip-OXA / CaO2 has the strongest cytotoxicity.

[0083] As shown in Table 5 below, the liposome IC of Example 2 of the present invention and Comparative Examples 1-3 are shown. 50 Among them, Rg3-Lip-OXA / CaO2 showed the strongest tumor cell killing ability and had the lowest IC 50 value.

[0084] Table 5 IC of Comparative Examples 1-3 and Example 2 of the present invention 50 value

[0085]

[0086] 12. In vivo anti-tumor effect and toxicity studies

[0087] Healthy female BALB / c nude mice weighing 18±2g were selected and raised in a clean environment at room temperature of 25°C with 12h day and night alternation. HCT116 cells were massively proliferated, and the cells were collected by centrifugation during the logarithmic growth phase. 0.1mL of cell suspension (5×106 cells / mouse) was subcutaneously injected into the flank of nude mice. After the tumor volume grew to more than 80mm3, the mice were intravenously injected with the drug through the tail vein, once every four days, for a total of 6 times. The physical signs of the mice were observed, and the weight and tumor volume of each group of mice were measured. After 30 days, the mice were killed, the tumor tissue was removed and weighed, and the tumor inhibition rate of each group was calculated. The dosage of Example 2 and Comparative Examples 1-3 was 5mg / kg OXA.

[0088] like Fig. 9As shown, the change curve of tumor volume and tumor inhibition rate of tumor-bearing nude mice after liposome treatment of Example 2 of the present invention and Comparative Examples 1-3, wherein the model group (Control) was injected with normal saline. It can be seen that the tumor volume of the Rg3-Lip-OXA / CaO2 group was significantly reduced, and the tumor inhibition rate was the highest.

[0089] like Fig.10 As shown, it is the weight change curve of tumor-bearing nude mice after liposome treatment of Example 2 of the present invention and Comparative Examples 1-3. It can be seen that the weight of mice in each group did not change much, but the weight of mice in the Rg3-Lip-OXA / CaO2 group of the present invention increased.

Claims

1. An oxaliplatin multifunctional liposome, characterized in that: The liposome is made of oxaliplatin, calcium peroxide nanoparticles, lecithin and ginsenoside Rg3.

2. The oxaliplatin multifunctional liposome according to claim 1, wherein the particle size is 104.78±3.96 nm, the PDI value is 0.152±0.011, the Zeta potential is -29.67±0.59, and the encapsulation efficiency of oxaliplatin is 43.96±3.18%.

3. A method for preparing oxaliplatin multifunctional liposomes, characterized in that: Lecithin and ginsenoside Rg3 are dissolved in a mixed solvent of chloroform and anhydrous ethanol and calcium peroxide nanoparticles are added, and rotary evaporation is performed on a rotary evaporator at 30-50°C for 1-3 hours to form a transparent lipid film; then an aqueous solution of oxaliplatin in glucose is added to make the final concentration of OXA 1-2 mg / mL, and the mixture is placed on a shaker at 30-50°C for hydration for 1-4 hours, and then ultrasonically treated at 100W for 10-20 minutes. The treated liposomes are extruded through a membrane with a pore size of 450nm several times and through a membrane with a pore size of 220nm several times to obtain.

4. The method for preparing an oxaliplatin multifunctional liposome according to claim 3, wherein the method for preparing calcium peroxide nanoparticles comprises the following steps: adding 2 mL of a 0.1 g / mL CaCl2 aqueous solution to 80 mL of PEG 200 After stirring evenly, 1 mL of 1 mol / L ammonia solution was added, and then 1 mL of 30% hydrogen peroxide solution was slowly added to the mixture at a rate of 3 drops per minute under rapid stirring. The mixture was stirred at room temperature for 6 h. Subsequently, 1 mol / L NaOH solution was added under the action of ultrasound to adjust the pH value of the solution to 11.

5. The precipitate was collected by centrifugation at 9500 g for 5 minutes, and washed twice with 0.1 mol / L sodium hydroxide solution and ethanol, respectively, to obtain white calcium peroxide nanoparticles.

5. The method for preparing an oxaliplatin multifunctional liposome according to claim 3, wherein the molar ratio of lecithin to ginsenoside Rg3 is 3-5:1; the molar ratio of lecithin to oxaliplatin is 15-20:1, and the final concentration of CaO2 is 1-2 mg / mL.

6. An application of oxaliplatin multifunctional liposomes, characterized in that: The peroxide nanoparticles in the liposome can improve the hypoxic microenvironment of solid tumors and enhance the anti-tumor efficacy of oxaliplatin; the ginsenoside Rg3 in the liposome replaces cholesterol and reduces the particle size of the liposome; the ginsenoside Rg3 in the liposome acts as a targeting fragment to specifically recognize and bind to the glucose transporter on the surface of HCT116 cells, enters the cell through receptor-mediated endocytosis, and increases the uptake of the liposome; the ginsenoside Rg3 in the liposome acts as a good chemotherapy adjuvant and is used in combination with chemotherapy drugs to increase the anti-tumor effect and reduce systemic toxicity.

7. Use of oxaliplatin multifunctional liposomes in colorectal cancer, characterized in that: The liposome is used as a drug for the preparation of colorectal cancer treatment, and can enhance the anti-tumor effect of chemotherapy drugs and reduce the systemic toxicity of chemotherapy drugs. The liposome has an IC 50 The value was 5.37±0.42μM, and the tumor inhibition rate was 87.55±9.35%.

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