A cispentacin immunoliposome, its preparation method and application

Abstract

This invention relates to an immunoliposome containing tebuconazole, its preparation method, and its application in anti-breast cancer treatment. The tebuconazole immunoliposome is composed of tebuconazole coated with phospholipids and cholesterol, and modified with trastuzumab. The preparation method of the tebuconazole immunoliposome of this invention is simple and exhibits excellent anti-tumor effects. The novel tebuconazole immunoliposome can overcome the drawback of tebuconazole's non-selectivity for tumor cells, thus enhancing its anti-tumor efficacy. Furthermore, the actively targeted immunoliposome with trastuzumab as a target exhibits significant targeting in vivo, and its anti-tumor effect is significantly different from that of non-targeted liposomes. This invention provides a new avenue for the anti-tumor application of tebuconazole and is of great significance for advancing the clinical application of tebuconazole.

Description

Technical Field

[0001] This invention relates to immune liposomes, specifically to immune liposomes containing tebufenozide, their preparation method, and their application in anticancer treatment, belonging to the field of biomedicine. Background Technology

[0002] In breast cancer patients, HER2 is highly expressed in approximately 30% of tumors, and in these patients, high HER2 expression is associated with decreased survival and shortened time to disease progression. High HER2 expression in human breast cancer cells increases their inherent metastatic potential; high HER2 expression or aberrant amplification makes breast cancer cells more aggressive, thus increasing the likelihood of brain metastasis. HER2-targeted therapy is promising because it targets a highly specific breast cancer growth receptor protein. Existing technologies have also made significant progress in molecularly targeted therapy by linking HER2 antibodies to drug molecules to form antibody-drug conjugates, improving the therapeutic index. For example, T-DM1 showed preliminary activity (ORR, 44%) in a phase 2 trial for treating advanced non-small cell lung cancer (NSCLC) with HER2 mutations (Tarantino P, Carmagnani Pestana R, Corti C, Modi S, Bardia A, Tolaney SM, Cortes J, Soria JC, Curigliano G. Antibody-drug conjugates: Smart chemotherapy delivery across tumor histologies. CA Cancer J Clin. 2022 Mar; 72(2):165-182).

[0003] Endiynes, naturally produced primarily by soil and marine microorganisms, are a class of small molecule compounds with potent antitumor activity containing conjugated alkyne-ene-yne ​​structures. Endiynes exhibit strong cytotoxicity against various cancer cell lines, including those resistant to clinically used chemotherapy. However, due to their non-selective cytotoxicity, enediynes themselves are unsuitable for chemotherapy. In fact, less than 1% of ADCs (anti-inflammatory drug complexes) reach human tumors; the remainder may cause unwanted toxicity.

[0004] Immunoliposomes have the potential to transfer large quantities of drug molecules to tumor cells, and drugs delivered via immunoliposomes exhibit antitumor activity similar to or higher than that of drugs alone. The ability of immunoliposomes to target tumor cells overcomes many limitations of traditional liposomes, providing a new strategy for tumor-targeted drug delivery. However, related studies have shown that the degree of improvement in drug efficacy delivered via immunoliposomes is unpredictable. For example, Michael Chen et al. developed a HER2-specific antibody to convert mPEG-coated liposomal doxorubicin (Lipo-Dox) into immunoliposomes. They compared the in vivo therapeutic effects between monoclonal antibody-modified and unmodified Lipo-Dox and showed that tumor suppression was not significantly improved in the anti-HER2-Lipo-Dox group (Chen M, Sheu MT, Cheng TL, Roffler SR, Lin SY, Chen YJ, Cheng YA, Cheng JJ, Chang HY, Wu TY, Kao AP, Ho YS, Chuang KH. A novel anti-tumor / anti-tumor-associated fibroblast / anti-mPEG tri-specific antibody to maximize the efficacy of mPEGylated nanomedicines against fibroblast-rich solidtumor. Biomother Sci. 2021 Dec 21;10(1):202-215).

[0005] Yanan Li et al. reported a HER2-modified thermosensitive liposome (immunoliposome) auxiliary complex, which obtained a novel biomass resonance structure carrier by reducing the gold nanoclusters on the surface (GTSL-CYC-HER2). The anti-tumor effect in orthotopic tumor-bearing mice was evaluated. The tumor volume growth inhibition rate of the GTSL-CYC-HER2@NIR group was increased by 0.5 times compared with the GTSL-CYC@NIR group (Li Y,Song W,Hu Y,Xia Y,Li Z,Lu Y,Shen Y. "Petal-like" size-tunable gold wrapped immunoliposome to enhance tumor deep penetration for multimodal guided two-step strategy. J Nanobiotechnology. 2021Sep27;19(1):293).

[0006] Xiaoyan Lu et al. reported that cetuximab was combined with liposomes loaded with afatinib to form immunoliposomes, and tumor growth experiments were conducted in tumor-bearing mice. The tumor volume growth inhibition rate of the LP-CTX group was 1-fold higher than that of the LP group (Lu X, Liu S, Han M, Yang X, Sun K, Wang H, Mu H, Du Y, Wang A, Ni L, Zhang C. Afatinib-loaded immunoliposomes functionalized with cetuximab: A novel strategy targeting the epidermal growth factor receptor for treatment of non-small-cell lung cancer. Int J Pharm. 2019 Apr 5; 560:126-135).

[0007] Tao Yang et al. investigated the antitumor effect of paclitaxel-loaded PEGylated immunoliposomes (PILs) in a breast cancer xenograft model. The tumor volume growth inhibition rate was 1-fold higher in the PIL group compared with that in the PL group (Yang T, Choi MK, Cui FD, Lee SJ, Chung SJ, Shim CK, Kim DD. Antitumor effect of paclitaxel-loaded PEGylated immunoliposomes against human breast cancer cells. Pharm Res. 2007 Dec; 24(12):2402-11).

[0008] Josimar O. Eloy et al. synthesized PTX / RAP co-loaded immunoliposomes modified with trastuzumab and evaluated their antitumor effects in a mouse xenograft model. The immunoliposome group showed a 2-3 fold increase in tumor volume growth inhibition compared to the liposome group (Eloy JO, Petrilli R, Chesca DL, Saggioro FP, Lee RJ, Marchetti JM. Anti-HER2 immunoliposomes for co-delivery of paclitaxel and rapamycin for breast cancer therapy. Eur J Pharm Biopharm. 2017 Jun; 115:159-167).

[0009] Huang Nian et al. disclosed a compound traditional Chinese medicine immunoliposome self-assembled from antibodies, lipid raw materials, and traditional Chinese medicine monomers bufotoxin and bee venom. The immunoliposome improved the efficiency by less than 1 times compared with the free drug group (A compound traditional Chinese medicine immunoliposome and its preparation method and application). Summary of the Invention

[0010] The present invention aims to provide a phenoxycin immunoliposome that specifically targets HER2, its preparation method, and its application in anticancer treatment. The phenoxycin immunoliposome has excellent antitumor effects and no obvious systemic toxic side effects.

[0011] To achieve the above objectives, the present invention provides the following solution:

[0012] A tiancimycin immunoliposome, wherein the immunoliposome is coated with tiancimycin and cholesterol and modified with trastuzumab.

[0013] Preferably, the phospholipid is a combination of natural phospholipids and polyethylene glycol-modified synthetic phospholipids.

[0014] Preferably, the natural phospholipid is soybean lecithin.

[0015] Preferably, the polyethylene glycol-modified synthetic phospholipids include DSPE-PEG2000 and DSPE-PEG2000-NHS.

[0016] Preferred ingredients include soybean lecithin, cholesterol, and DSPE-PEG. 2000 DSPE-PEG 2000 The molar ratio of NHS, trastuzumab, and teicoplanin was (56-58):(39-42):(5-10):1:(0.02-0.05):(0.004-0.006).

[0017] Preferably, the trastuzumab is a humanized monoclonal antibody.

[0018] Preferably, the trastuzumab is obtained by reacting with DSPE-PEG. 2000 -NHS reaction forms amide bonds, synthesizing DSPE-PEG. 2000 -trastuzumab then attaches to the surface of immune liposomes.

[0019] Preferably, the particle size of the immunoliposomes is 100-200 nm.

[0020] A method for preparing immune liposomes includes the following steps:

[0021] S1. Dissolve phospholipids, cholesterol and tebufenozide in anhydrous ethanol in parts by weight to form a lipid solution;

[0022] S2. Pour the lipid solution into the preheated PBS solution and vortex at 45-60℃ for 2-3 hours to form liposomes;

[0023] S3. Add trastuzumab, mix well, incubate, and then dialyze to obtain immunoliposomes.

[0024] Preferably, the dialysis time in step S3 is 24-48 hours.

[0025] Preferably, in step S3, the molecular weight cutoff of the dialysis bag is 300KD to remove trastuzumab that is not linked to the liposome.

[0026] Preferably, the encapsulated drug includes, but is not limited to, any one anthraquinone fused enediyne drug, including: tiancimycin and its derivatives (TNM B, TNM C), yangpumicin (YPM)A and its derivatives (YPM F, YPM G).

[0027] This invention also claims protection for the use of the aforementioned tiancimycin immunoliposomes in the preparation of antitumor drugs.

[0028] Preferably, the tumor is breast cancer, ovarian cancer, lung cancer, stomach cancer, prostate cancer, or bladder cancer.

[0029] The present invention will be further explained below:

[0030] In this invention, the tiancimycin immunoliposome is an active targeting liposome with trastuzumab as the target head, which has significant targeting in vivo. It can not only improve the disadvantage of tiancimycin's non-selectivity for tumor cells, but also enhance the in vivo anti-tumor effect of tiancimycin. The anti-tumor effect is significantly different from that of non-targeted agents.

[0031] According to the preparation method of the present invention, more than 80-90% of trastuzumab is successfully linked to immunoliposomes.

[0032] This invention relates to anti-HER2 immunoliposomes prepared by conjugating trastuzumab with PEG-stereostable liposomes, which have shown promising efficacy in treating HER2-overexpressing breast cancer. Specifically, PEGylation can increase the circulation time of the encapsulated drug in the blood, while the trastuzumab conjugate can increase the therapeutic index of the encapsulated drug by promoting selective delivery.

[0033] This invention also provides in vitro cytotoxicity data of tiancimycin immunoliposomes. Experiments show that tiancimycin immunoliposomes have good selectivity for tumor cells.

[0034] This invention also provides data from cellular uptake experiments of trastuzumab immunoliposomes, which show that conjugation with trastuzumab increases the internalization of immunoliposomes.

[0035] This invention also provides the anti-tumor effect of tiancimycin immunoliposomes on breast cancer in mice. Experiments show that tiancimycin immunoliposomes exhibit better anti-tumor effects than ordinary liposomes and can be used as an effective drug delivery strategy in the field of oncology medicine.

[0036] The beneficial effects of this invention are: the preparation method of tiancimycin immunoliposomes is simple and exhibits excellent anti-tumor effects. The novel tiancimycin immunoliposomes can overcome the drawback of tiancimycin's non-selectivity for tumor cells, thereby enhancing its anti-tumor efficacy. This invention provides a new avenue for the anti-tumor application of tiancimycin and is of great significance for promoting its clinical application. Attached Figure Description

[0037] Figure 1 The molecular weight of trastuzumab was detected by SDS-PAGE electrophoresis using the tiancimycin immunoliposome of the present invention.

[0038] Figure 2 This is a particle size distribution diagram of the Tiancimycin immunoliposomes of the present invention;

[0039] Figure 3 The in vitro cytotoxicity of free thymol TNM A, thymol liposome TNM A-Lipo, and thymol immunoliposome HER2-TNM A-ILs against SKBR3(A) and MCF-7(B);

[0040] Figure 4 This study investigated the cellular uptake of PI-Lipo and HER2-PI-ILs in KPL-4 cells at 2 and 4 hours.

[0041] Figure 5 The in vivo antitumor activity of free TNM A, TNM A-Lipo liposomes, and HER2-TNM A-ILs on BALB / c nude mice with subcutaneous KPL-4 cell tumors is shown in the following images: weight changes in nude mice after administration of TNM A, TNM A-Lipo, and HER2-TNM A-ILs (A); tumor volume changes in nude mice after administration of TNM A, TNM A-Lipo, and HER2-TNM A-ILs (B); tumor weight in nude mice after administration of TNM A, TNM A-Lipo, and HER2-TNM A-ILs (C); and tumor anatomy of nude mice obtained on day 10 after administration of TNM A, TNM A-Lipo, and HER2-TNM A-ILs (D). Detailed Implementation

[0042] The technical solution of the present invention will be further described in detail below through specific embodiments and in conjunction with the accompanying drawings. These descriptions are merely illustrative and are not intended to limit the scope of the invention. The protection scope of the present invention is not limited to the following embodiments. Unless otherwise specified, the raw materials and equipment used in this invention are commercially available or commonly used in the art. Unless otherwise specified, the methods in the following embodiments are conventional methods in the art.

[0043] Example 1

[0044] Preparation of Tiancimycin Immunoliposomes

[0045] Experimental procedure: 5 mL of PBS solution was placed in a vial and preheated in a 55°C water bath. The solution was then magnetically stirred to create a vortex. Soy lecithin, cholesterol, and DSPE-PEG were weighed according to a molar ratio of 56:39:5:1:0.004. 2000 DSPE-PEG 2000 -NHS and tebufenozide were dissolved in 2 mL of anhydrous ethanol by sonication to form a lipid solution, which was then vortexed until homogeneous. The oil phase lipid solution was quickly injected into the aqueous phase PBS solution and vortexed at 55°C for 2 h to form liposomes. Trastuzumab solution was added to a dialysis bag (molecular weight cutoff of 100 KD) and placed in dialysis buffer (50 mM Na2HPO4, 50 mM NaH2PO4, 10 mM EDTA, 0.15 M NaCl, pH 7.4) at 4°C for 6 h, with the dialysis buffer changed every 2 h. The dialyzed trastuzumab was mixed with the above liposome solution and then DSPE-PEG was applied. 2000 The molar ratio of NHS to trastuzumab was 50:1. The mixture was incubated at room temperature with shaking for 3 hours. The trastuzumab that was not conjugated to the liposomes was removed by PBS dialysis (molecular weight cutoff of 300KD) to obtain tiancimycin immunoliposomes, which were named HER2-TNM A-ILs.

[0046] The preparation method of Tiancimycin liposomes includes the following steps:

[0047] S1. Take 5 mL of PBS solution in a vial, preheat it in a 55°C water bath, and stir the PBS solution magnetically to form a vortex. Weigh out soybean lecithin, cholesterol, DSPE-PEG2000, and tebufenozide, and dissolve them in 2 mL of anhydrous ethanol by sonication to form a lipid solution.

[0048] S2. Quickly pour the oil phase lipid solution into the aqueous phase PBS solution and vortex at 55°C for 2 hours to form TNM A-Lipo liposomes.

[0049] Example 2

[0050] Molecular weight determination of trastuzumab on tiancimycin liposomes

[0051] Experimental Procedure: Transfer the sample to a new EP tube and add 4×SDS loading buffer according to the specified ratio. Perform SDS-PAGE denaturing electrophoresis on a 15% separating gel. Load the same volume of sample into each lane and perform routine electrophoresis. The results are as follows: Figure 1 As shown.

[0052] The results showed that free trastuzumab exhibited two bands at 50 kDa and 25 kDa, representing the heavy chain CH1 and the light chain VH, respectively. No bands were observed on TNM A-Lipo, but similar bands at 50 kDa and 25 kDa appeared in the HER2-TNM A-ILs sample, indicating that trastuzumab had been successfully linked to DSPE-PEG via an amide bond. 2000 -NHS, and then spontaneously integrate into immune liposomes.

[0053] Example 3

[0054] Particle size distribution determination of tiancimycin liposomes

[0055] Experimental procedure: The dynamic dimensions of TNM A-Lipo and HER2-TNM A-ILs were determined using a Zetasizer Nano 90 (Malvern). The results are as follows: Figure 2 As shown.

[0056] The results showed that the average particle sizes of TNM A-Lipo and HER2-TNM A-ILs were 202.8 ± 1.5 nm and 182.8 ± 2.1 nm, respectively. Compared with TNM A-Lipo, the particle size of HER2-TNM A-ILs decreased by 20 nm after modification with trastuzumab, which may be due to the invagination of the phospholipid bilayer caused by antibody linkage.

[0057] Example 4

[0058] In vitro cytotoxicity of tiancimycin liposomes

[0059] Experimental Procedure: The in vitro cytotoxicity of free tiancimycin TNM A, tiancimycin liposome TNM A-Lipo, and tiancimycin immunoliposome HER2-TNM A-ILs on SKBR3 and MCF-7 cells was determined using the CCK-8 assay. MCF-7 cells (5 × 10⁻⁶) were... 3 Transfer SKBR3 cells (5 × 10⁶ cells / 100 μL) to a 96-well plate containing RPMI-1640 medium. 3Cells (100 μL / well) were transferred to 96-well plates containing DMEM medium. After overnight incubation, the cells were treated for 48 h in a 5% CO2 cell culture incubator with different concentrations of free tebufenozide TNM A, tebufenozide liposome TNM A-Lipo, and tebufenozide immunoliposome HER2-TNM A-ILs (0.0001, 0.01, 0.1, 0.5, 1, 5, 50, 100 nM). Then, 110 μL of medium containing 10 μL of CCK-8 was added to each well. The cells were then incubated at 37°C for 1 h, and the optical density (OD) was measured at 450 nm using a microplate reader. The average value of three replicates was taken for each group, and the cell inhibition rate was calculated. The results are shown below. Figure 3 As shown.

[0060] The results showed that SKBR3 is a human breast cancer cell line that highly expresses HER2, while MCF-7 is a human breast cancer cell line that lowly expresses HER2. HER2-TNM A-ILs exhibited significantly higher cytotoxicity in SKBR3 cells than TNM A-Lipo, indicating that conjugation of trastuzumab to the surface of TNM A-Lipo can enhance the targeting efficiency of TNM A in these cell lines. In SKBR3 cells, HER2-TNM A-ILs showed more pronounced cytotoxicity, with a half-maximal inhibitory concentration (IC50) of [missing value]. 50 The concentration was 0.24 nM, significantly lower than that of untargeted TNM A-Lipo and TNM A. In MCF-7 cells, HER2-TNM A-ILs exhibited low cytotoxicity, with a half-maximal inhibitory concentration (IC50) of 0.24 nM, far less than that of untargeted TNM A-Lipo and TNM A. 50 The concentration was 1.54 nM, which is smaller than that of untargeted TNM A-Lipo (1.89 nM) and TNM A (1.73 nM).

[0061] Example 5

[0062] Cellular uptake of Tiancimycin Liposomes

[0063] Experimental Procedure: Propidium iodide (PI) fluorescent dye was used instead of TNM A to prepare PI-Lipo and HER2-PI-ILs using the same method. KPL-4 cells were cultured at 2×10⁻⁶ cells / year. 5 Cells were seeded per well in 6-well plates and incubated for 24 hours. The cell culture medium was discarded, and the cells were gently washed three times with PBS buffer. Then, PI-Lipo and HER2-PI-ILs (200 ng / well) were added and incubated for 2 hours and 4 hours, respectively. The cells were washed three times with pre-cooled PBS buffer to remove any remaining drug solution. The cells were then fixed with 4% paraformaldehyde and stained with 1 μg / mL DAPI solution. Cell uptake of liposomes was observed using an inverted fluorescence microscope. The results are shown below. Figure 4 As shown.

[0064] The results showed that the efficacy of immunoliposomes depended on the internalization capacity of tumor cells. KPL-4 cells were incubated with PI-Lipo and HER2-PI-ILs at 200 ng / well. All samples were incubated for 2 hours and 4 hours. HER2-PI-ILs entered KPL-4 cells more readily than PI-Lipo, indicating that trastuzumab conjugation promoted immunoliposome internalization. Given the high expression of the HER2 receptor on the surface of KPL-4 cells, the addition of trastuzumab successfully enhanced the internalization of immunoliposomes by tumor cells.

[0065] Example 6

[0066] In vivo anti-breast cancer activity of tecipitin immunoliposomes

[0067] Experimental Procedure: Six-week-old BALB / c nude mice were acclimatized for 5 days. KPL-4 cells in the logarithmic growth phase were digested with trypsin and collected, then washed twice with PBS. KPL-4 cells (5 × 10⁶ cells in 50 μL PBS and 50 μL Matrigel) were subcutaneously injected into the right axilla of the BALB / c nude mice. 6 (1 cell) to establish a subcutaneous tumor-bearing model (O'HAGAN RC, HEYER J. Krasmouse models: modeling cancer harboring Kras mutations[J].Genes&Cancer, 2011, 2(3):335-343. Yan Kemin, Sun Jia, Wang Jiaojiao, Nie Yongzhan, Xiao Haijuan, Research progress on the application of PDX model in malignant tumors[J], Modern Oncology Medicine 2019, 27(09):1629-1634). The mental state, diet, defecation, weight and activity of mice were observed. The size of the tumor was measured on the 3rd day after the tumor formation following subcutaneous inoculation. The length (a) and width (b) of the tumor were measured daily with calipers. The tumor volume V was calculated by the following formula: V=1 / 2ab2 (the model was successfully established based on the physiological condition and tumor volume). When the tumor volume of each successfully established BALB / c nude mouse model reached about 100 mm. 3 At the time (evaluation process after modeling), they were randomly divided into 6 groups (n=5): untreated control group (0.9% saline), positive control group (trastuzumab (10mg / kg), TNM A (0.05mg / kg), TNM A-Lipo (0.02mg / kg), HER2-TNM A-ILs (0.02mg / kg) and HER2-TNM A-ILs (0.02mg / kg) + trastuzumab (10mg / kg) combined, administered once on day 0 for a total of 14 days.

[0068] On the last day of the experiment, all BALB / c nude mice were euthanized, solid tumors were dissected, weighed, and photographed for recording. The heart, liver, spleen, lungs, and kidneys were isolated, and blood was collected. After sampling, the gross condition of the organs was observed, such as surface color, hardness, elasticity, surface changes, and presence of hemorrhage. The tissues were then fixed in 10% neutral formaldehyde to prepare paraffin sections for histological examination (HE staining, Masson staining). The remaining tissues were stored at -80°C for later use. Blood was also used for inflammatory analysis. The results are as follows: Figure 5 As shown.

[0069] Results showed that the therapeutic effect of HER2-TNM A-ILs was investigated using a KPL-4 human breast cancer subcutaneous xenograft model via intravenous administration. We administered TNM A-Lipo (0.02 mg / kg) and HER2-TNM A-ILs (0.02 mg / kg) to 5 female BALB / c nude mice carrying tumors, with trastuzumab (10 mg / kg) and free TNM A (0.05 mg / kg) serving as controls. Single-dose treatment with HER2-TNM A-ILs and HER2-TNM A-ILs plus trastuzumab significantly inhibited tumor growth in nude mice. Figure 5 No significant weight loss was observed in any of the treatment groups, indicating that these treatment regimens were well tolerated. Figure 5 A). Conversely, single doses of free TNM A, TNM A-Lipo, and trastuzumab showed only slight inhibitory effects on KPL-4 tumors. However, two lower doses of HER2-TNM A-ILs or HER2-TNM A-ILs + trastuzumab significantly inhibited tumor growth in treated nude mice without weight loss during treatment. Figure 5 B). The HER2-TNM A-ILs group had the smallest tumor weight, confirming the optimal antitumor effect of HER2-TNM A-ILs, and the effect of HER2-TNM A-ILs was far greater than the sum of TNM A-Lipo (0.02 mg / kg) and trastuzumab (10 mg / kg). Figure 5 C). Importantly, TNM A-Lipo and HER2-TNM A-ILs exhibited stronger antitumor activity than trastuzumab, which showed only weak tumor-suppressive activity upon single-dose administration. TNM A-Lipo also showed only antitumor activity comparable to high-dose free TNM A. Figure 5 B and Figure 5D). Therefore, in this invention, trastuzumab and tiancimycin liposomes exhibit a significant synergistic effect.

[0070] The above description is merely a preferred embodiment of the present invention. The scope of protection of the present invention is not limited to the above embodiments. All technical solutions falling within the scope of the present invention's concept are within the scope of protection of the present invention. It should be noted that for those skilled in the art, any improvements and modifications made without departing from the principles of the present invention should also be considered within the scope of protection of the present invention.

Claims

1. A liposome immunovaccine of caelumycin, characterized in that, The immunoliposome is coated with phospholipid and cholesterol, and is modified with trastuzumab; the phospholipid is a combination of natural phospholipid and polyethylene glycol synthetic phospholipid; The natural phospholipid is soy lecithin, and the pegylated synthetic phospholipid comprises DSPE-PEG 2000 and DSPE-PEG 2000 -NHS; The trastuzumab is reacted with DSPE-PEG2000-NHS to form an amide bond, and DSPE-PEG2000-trastuzumab is synthesized to be connected to the surface of the immunoliposome; The soy lecithin, cholesterol, DSPE-PEG 2000 , DSPE-PEG 2000 The molar ratio of the soy lecithin, cholesterol, DSPE-PEG -NHS, trastuzumab, and calicheamicin is (56-58):(39-42):(5-10):1:(0.02-0.05):(0.004-0.006).

2. The natamycin immunoliposome according to claim 1, characterized in that, The particle size of the liposome is 100-200 nm.

3. A method of preparing the natamycin immunoliposome according to claim 1 or 2, characterized in that, The method comprises the following steps: S1, according to weight parts, dissolving phospholipid, cholesterol and calicheamicin in anhydrous ethanol to form a lipid solution; S2, pouring the lipid solution into a preheated PBS solution, stirring at 45-60 ℃ for 2-3 h to form a liposome; S3, adding trastuzumab, mixing, incubating and then dialyzing to obtain an immunoliposome.

4. The production method according to claim 3, characterized by, The dialysis time in step S3 is 24-48 h.

5. The use of calicheamicin immunoliposome according to claim 1 or 2 in the preparation of an anti-breast cancer drug.

Images