Targeted delivery carrier based on human papilloma virus-like particles and application of targeted delivery carrier in preparation of antitumor drugs

Abstract

The invention discloses a targeted delivery carrier based on human papilloma virus-like particles and application of the targeted delivery carrier in preparation of antitumor drugs, and belongs to the technical field of targeted delivery carriers. The targeted delivery carrier siRNA-pep-coated VLP-SP5-2 prepared by the invention simultaneously encapsulates two different types of antitumor drugs, namely Bcl-2siRNA and antitumor peptide D-pep, and has the advantages of high encapsulation efficiency, good targeting property, easiness in synthesis, strong stability, good biocompatibility and the like; compared with siRNA-pep, the total apoptosis rate of the cells is increased from 60% to 91%; compared with siRNA-pep (at) VLP, the anti-tumor inhibition rate is improved by about 40%. The targeted delivery carrier can target a non-small cell lung cancer cell line and has excellent tumor cell apoptosis promoting capability; the apoptosis of the non-small cell lung cancer cells is efficiently induced through the synergistic effect of the two drugs, targeted therapy is achieved, and the compound can be used for preparing anti-tumor drugs.

Description

Technical Field

[0001] The present invention belongs to the technical field of targeted delivery vectors, and specifically relates to a targeted delivery vector (siRNA_D-pep@VLP-SP5-2) for targeted delivery of siRNA and anti-tumor peptide D-LAK-120A (D-pep) based on human papillomavirus-like particles (HPVVLPs), and its application in the preparation of anti-tumor drugs. Background Art

[0002] Cancer has a high incidence and mortality rate. According to the World Health Organization, there will be nearly 20 million new cases of cancer and nearly 10 million deaths worldwide in 2022, posing a serious threat to life and health around the world. Therefore, it is a consensus around the world to find effective anti-tumor treatments. Cancer treatment usually involves the use of systemic administration methods, such as chemotherapy and some peptides with anti-tumor activity (such as D-LAK-120A, D-pep). However, chemotherapy drugs have no effect on many cancer patients or the therapeutic effect gradually decreases over time, partly because cancer cells develop resistance to apoptosis. Escape from apoptosis occurs through a variety of mechanisms, including non-homologous end-linked proteins that enhance DNA repair or overexpression of anti-apoptotic proteins, which can act alone or synergistically. A major anti-apoptotic factor is the B-cell lymphoma 2 (Bcl-2) protein family, which was discovered in non-Hodgkin's lymphoma nearly 30 years ago. Bcl-2 is located in the mitochondrial membrane and endoplasmic reticulum membrane, which can prevent the release of apoptosis-inducing factors and cytochrome c and inhibit caspase-mediated apoptosis. Due to the important role of Bcl-2 in the apoptosis pathway, several therapeutic strategies have been developed to inhibit or downregulate Bcl-2 protein. RNA interference (RNAi) is a new cancer treatment strategy that uses small interfering RNA (siRNA) to mediate gene silencing in cells to achieve the purpose of cancer treatment. siRNA used to silence Bcl-2 has successfully reduced the expression of Bcl-2 in a variety of tumors. However, siRNA itself has problems and disadvantages such as being easily degraded by nucleases, being easily cleared, having difficulty passing through the cell membrane, poor targeting, immunogenicity, and off-target effects.

[0003] Currently, one solution for efficient delivery of RNA to target cells is to use virus-like particles (VLPs) as targeted delivery vehicles. VLPs are engineered nanoparticles that mimic the properties of viruses and have strong tolerance to heat and proteases, but lack viral genomes and infectivity. VLPs are composed of multi-subunit structural proteins, have spontaneous self-assembly capabilities, and are easy to functionalize. They are ideal targeted delivery vehicles with almost all the advantages of real virus particles. Human papillomavirus-like particles (HPVVLPs) stand out among various VLPs due to their excellent expression levels and efficient self-assembly capabilities. HPV L1 (the monomer of HPV VLPs) self-assembles into HPV VLPs through disulfide bonds between 72 pentamers formed in the first stage. HPV VLPs have many advantages such as high stability, good biocompatibility, and easy functionalization, so they have extremely high practical application potential in the biomedical field.

[0004] Due to the heterogeneity and complexity of cancer, precise targeting of tumors is very necessary. Targeted therapy can capture tumors faster and enrich them on the tumor surface faster, thereby significantly promoting drug enrichment at the tumor site; therefore, the concept of targeted delivery of nanomedicines is an emerging trend in the field of cancer treatment research. D-LAK-120A (D-pep), as a peptide with anti-tumor activity, has a wide range of cytotoxicity similar to other chemotherapy drugs, and can achieve targeted therapy by precisely targeting tumor cells; while siRNA itself has the disadvantages of being easily degraded, difficult to pass through the cell membrane, and poor targeting, and a reliable targeted delivery carrier is needed to achieve its therapeutic function on target cells. The present invention utilizes the large cavity (diameter of about 35nm) of HPV VLP to simultaneously encapsulate two different anti-tumor drugs, D-pep and Bcl-2siRNA, and modifies the targeting peptide SP5-2 on the outer surface of HPV VLP to improve its precise targeting to tumor cells, thereby achieving co-encapsulation and targeted delivery of the two drugs. Summary of the invention

[0005] The purpose of the present invention is to provide a targeted delivery vector (siRNA_D-pep@VLP-SP5-2) for the targeted delivery of Bcl-2 siRNA and anti-tumor peptide D-LAK-120A (D-pep) based on virus-like particles and its application in the preparation of anti-tumor drugs (siRNA_D-pep@VLP-SP5-2).

[0006] The targeted delivery vector (siRNA_D-pep@VLP-SP5-2) prepared by the present invention simultaneously encapsulates Bcl-2 siRNA and anti-tumor peptide D-pep, specifically, Bcl-2 siRNA and D-pep are mixed in a binding buffer (containing 40mM H2PO4) at a nitrogen-phosphorus ratio of 0.75 to 3:1. - / HPO42- , 100 mM NaCl and 0.1 mM dithiothreitol, pH = 7 aqueous solution) for 20 to 40 minutes to assemble the siRNA_D-pep complex; then, the HPVL1 pentamer (HPV L1-p) and the siRNA_D-pep complex were incubated in the assembly solution (containing 50 mM H2PO4) at a mass ratio of 0.3 to 0.8:1. - / HPO4 2- and 1.5MNaCl, pH=6.0 aqueous solution) to obtain siRNA_D-pep@VLP; then, siRNA_D-pep@VLP was mixed with targeting peptide SP5-2 at a molar ratio of 1:1-8, and then newly configured EDC and NHS solutions were added, with final concentrations of 3-5mM, and reacted at 3-5°C for 6-10 hours; finally, the obtained solution was added to a dialysis tube (100kDa), and dialyzed in PBS buffer with continuous stirring for 10-15 hours, repeated 3-5 times to remove free SP5-2 and other by-products, and a targeted delivery vector (siRNA_D-pep@VLP-SP5-2) solution was obtained in the dialysis tube, which can be used to target non-small cell lung cancer A549 cell line.

[0007] The targeted delivery vector (siRNA_D-pep@VLP-SP5-2) was co-incubated with the non-small cell lung cancer A549 cell line for 0.5 to 2 hours, and the green fluorescence intensity in the cells was analyzed by fluorescence microscopy to evaluate the ability of the targeted delivery vector (siRNA_D-pep@VLP-SP5-2) to target cells. The targeted delivery vector (siRNA_D-pep@VLP-SP5-2) prepared by the present invention can be used to promote apoptosis of the non-small cell lung cancer A549 cell line, and then used to prepare anti-tumor drugs.

[0008] The prepared targeted delivery carrier (siRNA_D-pep@VLP-SP5-2) based on human papillomavirus-like particles (HPV VLP) of the present invention simultaneously encapsulates two different types of anti-tumor drugs (Bcl-2siRNA and anti-tumor peptide D-pep), and the delivery carrier has the advantages of high encapsulation rate (71%), good targeting, easy synthesis, strong stability, good biocompatibility, etc. siRNA_D-pep@VLP-SP5-2 has a strong ability to promote apoptosis of A549 cells. The in vitro results show that the total apoptosis rate of siRNA_D-pep@VLP-SP5-2 is increased from 60% to 91% compared with siRNA_D-pep; the anti-tumor results in mice show that siRNA_D-pep@VLP-SP5-2 has a stronger tumor targeting ability and better therapeutic effect, and the anti-tumor inhibition rate is increased by about 40% relative to siRNA_D-pep@VLP. This targeted delivery vector can target non-small cell lung cancer cell lines, has excellent ability to promote tumor cell apoptosis, and enables HPV VLP to have the ability to target non-small cell lung cancer cell lines; through the synergistic effect of the two drugs, it can efficiently induce apoptosis of non-small cell lung cancer cells, thereby achieving targeted treatment. BRIEF DESCRIPTION OF THE DRAWINGS

[0009] Figure 1 : Fluorescence spectra of VLP (HPV VLP), FITC-SP5-2 and VLP-FITC-SP5-2; It can be seen that both FITC-SP5-2 and VLP-FITC-SP5-2 have FITC emission peaks at 510-520 nm, proving that FITC-SP5-2 has been successfully modified to the outer surface of VLP;

[0010] Figure 2 : Fluorescence microscopy images of the invasive ability of FITC-VLP and VLP-FITC-SP5-2 on A549 cells; Figure A is FITC-labeled VLP (FITC-VLP), and Figure B is VLP-FITC-SP5-2, and the invasion times are 0.5h, 1h and 2h, respectively. It can be seen that with the increase of invasion time, the invasive ability of FITC-VLP and VLP-FITC-SP5-2 on A549 cells is significantly improved; among them, VLP-FITC-SP5-2 shows a higher internalization rate, and compared with FITC-labeled VLP alone, VLP-FITC-SP5-2 enters A549 cells in a shorter time (1 hour).

[0011] Figure 3:The binding affinity diagram of siRNA and D-pep at different amino / phosphate ratios (D-pep:siRNA=0.75:1, 1.125:1, 1.5:1, 2.25:1, 3:1) was determined by agarose electrophoresis; it can be seen that when the different amino / phosphate ratios increased from 0.75:1 to 1.5:1, the corresponding siRNA band gradually shifted backward, indicating that siRNA and D-pep successfully assembled into a complex, and as the ratio increased, the amount of D-pep in the complex also increased; when the different amino / phosphate ratios increased from 1.5:1 to 3:1, the siRNA band did not change significantly, indicating that the amount of D-pep has reached saturation.

[0012] Figure 4 : Figure 4 A is the transmission electron microscopy photo and average particle size diagram of the siRNA_D-pep complex obtained with an amino group / phosphate group ratio of 1.5:1, showing that the average particle size is 12.6±3.3nm; Figure 4 B is the potential diagram of the siRNA_D-pep complex and siRNA_D-pep@VLP-SP5-2 obtained with an amino group / phosphate group ratio of 1.5:1, showing that the surface potential of siRNA_D-pep is -6.5 mV, and the surface potential of siRNA_D-pep@VLP-SP5-2 is 2.9 mV; Figure 4 C is the transmission electron microscopy photo and average particle size diagram of siRNA_D-pep@VLP-SP5-2 obtained with an amino group / phosphate group ratio of 1.5:1, showing that the average particle size of siRNA_D-pep@VLP-SP5-2 is 57.5±9nm;

[0013] Figure 5 : Figure 5 A is the fluorescence spectra of different concentrations of D-pep-FITC (3, 5, 7.5, 10, 15, 20 μg / mL) and siRNA_D-pep-FITC@VLP-SP5-2 (D-pep-FITC addition amount is 10.1 μL, D-pep-FITC addition concentration is 1 mg / mL) assembled by D-pep-FITC instead of D-pep, where the curves of D-pep-FITC concentration from small to large are numbered Ⅰ to VI (D-pep-FITC is an aqueous solution), and the volume is 1 mL. Figure 5 B is the linear relationship curve between the concentration (x) of D-pep-FITC and the fluorescence intensity (y) at 520 nm. The standard equation is y = 34.5x-21.68, R 2=0.99, and the actual concentration of D-pep-FITC in siRNA_D-pep-FITC@VLP-SP5-2 was calculated to be 7.4 μg / mL and the actual volume was 1 mL by the standard equation. The encapsulation efficiency (%) = actual concentration of D-pep-FITC × actual volume / added concentration of D-pep-FITC × added volume, and the calculated encapsulation efficiency was about 71%.

[0014] Figure 6 : A bar graph showing the cell apoptosis rate after co-incubation with A549 cells for 12 hours and 24 hours; A549 cells were double stained with AnnexinV-FITC (AV) and propidium iodide (PI), respectively, and then the cell apoptosis rates of the experimental group and the groups using D-pep, VLP+D-pep, siRNA_D-pep, siRNA_D-pep@VLP, and siRNA_D-pep@VLP-SP5-2 were evaluated by flow cytometry analysis; Figure 6 A shows that after 12 hours of co-incubation, there was no significant difference in the apoptosis rate induced by D-pep and siRNA_D-pep. In contrast, siRNA_D-pep@VLP formed after VLP encapsulation of siRNA_D-pep was observed to significantly increase the apoptosis rate. After VLP was functionally modified with SP5-2 (i.e., siRNA_D-pep@VLP-SP5-2), the induced apoptosis rate was further increased. In addition, the combination of VLP and D-pep resulted in a higher apoptosis rate. Figure 6 B shows that the detection results after 24 hours of co-incubation are consistent with the results after 12 hours of co-incubation.

[0015] Figure 7 A is a photo of mouse tumor. Figure 7 B is the mouse tumor growth curve. Figure 7 C is a bar graph of mouse tumor weight. Figure 7 D is the weight change curve of mice. Figure 7 E is the in vivo imaging photo of siRNA_D-pep@VLP and siRNA_D-pep@VLP-SP5-2. DETAILED DESCRIPTION

[0016] The Bcl-2 siRNA used in the present invention was purchased from Shanghai Aibosi Biotechnology Co., Ltd. (purity ≥ 99%). Targeting peptides (SP5-2, FITC-SP5-2) and D-LAK-120A (D-pep) were purchased from Shanghai Chupeptide Biotechnology Co., Ltd. (purity ≥ 99.9%). Sodium dihydrogen phosphate (NaH2PO4), disodium hydrogen phosphate (Na2HPO4), and sodium chloride (NaCl) were purchased from Shanghai Aladdin Biochemical Technology Co., Ltd. Dithiothreitol was purchased from Beijing Solebold Technology Co., Ltd. The cell apoptosis kit was purchased from Biyuntian Biotechnology Co., Ltd. HPV L1-p protein was purified by Glutathione Sepharose affinity chromatography, and HPV L1-p was purified in assembly solution (50mM H2PO4 - / HPO4 2- , 1.5 M NaCl, pH = 6.0) to obtain HLP VLPs by self-assembly.

[0017] If not specifically mentioned, the solutions described in the present invention are all deionized water solutions.

[0018] Example 1

[0019] Preparation and verification of VLP-FITC-SP5-2:

[0020] 12.1 μL of FITC-SP5-2 solution (1 mg / mL) was added to 1 mL of HPV L1 solution (0.2 mg / mL), and after mixing, 68.2 μL of NHS and EDC solution (the initial concentration of the mother solution was 50 mM) were added respectively to make the final concentration 4 mM, and the mixture was reacted at 4°C on a flip shaker for 8 hours. The obtained VLP-FITC-SP5-2 was added to a dialysis tube (100 kDa), and the mixture was dialyzed in PBS buffer with continuous stirring for 12 hours. This was repeated 3 times to remove free SP5-2 and other byproducts, and VLP-FITC-SP5-2 was obtained in the dialysis tube. The results are shown in FIG. Figure 1 As shown, VLP alone has no emission peak under 488nm excitation light, while VLP-FITC-SP5-2 and FITC-SP5-2 both have emission peaks at 510-520nm, proving that SP5-2 has been successfully modified to the outer surface of VLP.

[0021] Example 2

[0022] Targeting verification of VLP-FITC-SP5-2:

[0023] The prepared concentration was 75 μg mL -1VLP-FITC-SP5-2 and FITC-labeled VLP (FITC-VLP) of the same concentration were co-incubated with A549 cells for 0.5 to 2 h, then washed three times with PBS solution, and the cellular internalization rate of VLP-FITC-SP5-2 was analyzed by fluorescence microscopy. Figure 2 As shown, Figure 2 A shows the verification of the invasive ability of FITC-VLP to A549 cell line. The results show that FITC-VLP has a certain invasive ability to A549 cell line; Figure 2 B shows the verification of the invasive ability of VLP-FITC-SP5-2 on the A549 cell line. The results show that VLP-FITC-SP5-2 shows a higher internalization rate. Compared with FITC-labeled VLP alone, VLP-FITC-SP5-2 enters A549 cells in a shorter time (1 hour), and the cells have stronger fluorescence after 2 hours of co-incubation, proving that SP5-2 improves the targeted invasion ability of VLP on the A549 cell line.

[0024] VLP-FITC-SP5-2 is a product of VLP and FITC-SP5-2 connected together through an amide reaction mediated by NHS and EDC. In order to prove that the amide reaction can connect SP5-2 to VLP, FITC-SP5-2 was used instead of SP5-2 for verification.

[0025] Example 3

[0026] Preparation and purification of siRNA_D-pep@VLP-SP5-2:

[0027] Bcl-2 siRNA and D-LAK-120A were mixed in a binding buffer (40 mM H2PO4) at a nitrogen-phosphorus ratio of 0.75 to 3:1. - / HPO4 2- , 100 mM NaCl, 0.1 mM dithiothreitol, pH = 7) and incubated for 30 minutes to assemble the complex siRNA_D-pep; then the HPV L1 pentamer and siRNA_D-pep complex with a mass ratio of siRNA: HPV L1 of 0.5:1 were added in the assembly solution (50 mM H2PO4 - / HPO4 2-, 1.5MNaCl, pH=6.0) to obtain siRNA_D-pep@VLP; then siRNA_D-pep@VLP was mixed with targeting peptide SP5-2 at a molar ratio of 1:4, and then newly prepared EDC and NHS solutions were added, with the final concentration of each solution being 4mM, and reacted at 4°C for 8 hours. The obtained solution was added to a dialysis tube (100kDa) and dialyzed in PBS buffer with continuous stirring for 12 hours, repeated 3 times to remove free SP5-2 and other byproducts, and siRNA_D-pep@VLP-SP5-2 solution was obtained in the dialysis tube. The results are shown in Figure 3 As shown in the figure, when the N / P ratio increases from 0.75:1 to 1.5:1, the siRNA band gradually shifts backwards, and when the N / P ratio continues to increase, the siRNA band position no longer continues to shift, and the siRNA band brightness also gradually decreases. This proves that the N / P ratio of 1.5:1 is the optimal ratio. Figure 4 The particle size of siRNA_D-pep with N / P ratio of 1.5:1 was about 12.6±3.3 nm, and the surface charge was negative. The particle size of assembled siRNA_D-pep@VLP-SP5-2 was about 57.5±9 nm, and the surface charge was positive. Figure 5 The encapsulation efficiency calculated based on D-pep was approximately 71%.

[0028] Example 4

[0029] siRNA_D-pep@VLP-SP5-2 induces apoptosis of A549 cells:

[0030] 3 μg of Bcl-2 siRNA and 4.1 μg of D-pep (nitrogen-phosphorus ratio of 1.5:1) were assembled to obtain siRNA_D-pep@VLP-SP5-2; and siRNA_D-pep@VLP, siRNA_D-pep, D-pep, and D-pep+VLP assembled at the same concentration were incubated with A549 cells at 37°C for 12 to 24 hours, and the cells were collected and analyzed for cell apoptosis by flow cytometry using an apoptosis kit. The results are shown in Figure 6 As shown, Figure 6A shows that after 12 hours of co-incubation with A549 cells, there is no significant difference in the apoptosis rate induced by D-pep and siRNA_D-pep. This may be because only D-pep exerts a biological effect, while siRNA without a carrier cannot enter the cell to exert its intended function. In contrast, siRNA_D-pep@VLP formed after VLP encapsulation of siRNA_D-pep was observed to significantly increase the apoptosis rate, from 60% to about 80%, proving the important role of VLP in encapsulating and delivering siRNA_D-pep. After functional modification of VLP with SP5-2 (i.e., siRNA_D-pep@VLP-SP5-2), it was found that the induced apoptosis rate was further increased, from 80% to about 91%, proving that modification of the targeting peptide is also essential. In summary, the total apoptosis rate of siRNA_D-pep@VLP-SP5-2 cells increased from 60% to 91% compared with siRNA_D-pep. Figure 6 B shows that the detection results after 24 hours of co-incubation are consistent with those after 12 hours of co-incubation, which proves that siRNA_D-pep@VLP-SP5-2 can effectively induce apoptosis in A549 cell lines.

[0031] Example 5

[0032] Evaluation of the anti-tumor effect of siRNA_D-pep@VLP-SP5-2 in vivo:

[0033] Blab / c nude mice were injected subcutaneously with 1×10 7 A549 cells, when the tumor grows to 40-50 mm 3 At the time of the experiment, the mice were divided into 4 groups, each with 4 mice, namely control group, siRNA_D-pep group, siRNA_D-pep@VLP group and siRNA_D-pep@VLP-SP5-2 group, and PBS, siRNA_D-pep, siRNA_D-pep@VLP and siRNA_D-pep@VLP-SP5-2 were injected intravenously, respectively. The dosage of siRNA was 1.5 mg / kg per mouse, and that of D-pep was 2 mg / kg per mouse. The injection volume of PBS, siRNA_D-pep, siRNA_D-pep@VLP and siRNA_D-pep@VLP-SP5-2 was 100 μL per mouse. The tumor size and mouse weight were recorded every two days. Figure 7 A~ Figure 7D, the tumor inhibition rate of the siRNA_D-pep group was 38.5%, and the tumor inhibition rates of the siRNA_D-pep@VLP group and the siRNA_D-pep@VLP-SP5-2 group were 49.25% and 78.2%, respectively, indicating that the therapeutic effect of siRNA_D-pep@VLP-SP5-2 was the best, and the tumor inhibition rate was increased by about 40% compared with siRNA_D-pep. In addition, the in vivo imaging of mice using Cy5-labeled siRNA_D-pep@VLP and siRNA_D-pep@VLP-SP5-2 showed that siRNA_D-pep@VLP could also enter the tumor, but some of them would not accumulate at the tumor location, while siRNA_D-pep@VLP-SP5-2 completely accumulated at the tumor location (such as Figure 7 E. Among them, I: PBS; II: siRNA_D-pep@VLP-SP5-2; III: siRNA_D-pep@VLP). This proves that siRNA_D-pep@VLP-SP5-2 still has good targeting in mice.

[0034] The above examples illustrate that we have successfully constructed a co-targeted delivery vector of siRNA and D-pep, which has a good encapsulation rate. The targeted delivery vector has the ability to target the A549 cell line, and the targeted delivery efficiency is improved. It can effectively promote tumor cell apoptosis both in vitro and in vivo, and has a good anti-tumor therapeutic effect, so it can be used to prepare anti-tumor drugs.

[0035] It should also be noted that the specific embodiments of the present invention are only used for illustrative purposes and do not limit the scope of protection of the present invention in any way. Relevant technicians in this field can make improvements or changes based on some of the above descriptions, but all these improvements and changes should fall within the scope of protection of the claims of the present invention.

Claims

1. A targeted delivery vector based on human papillomavirus-like particles, characterized in that: Bcl-2 siRNA and D-pep were co-incubated in a binding buffer at a nitrogen-phosphorus ratio of 0.75 to 3:1 for 20 to 40 minutes to assemble the complex siRNA_D-pep; then, HPV L1 pentamer and siRNA_D-pep complex were co-assembled in the assembly solution at a siRNA:HPV L1 mass ratio of 0.3 to 0.8:1 to obtain siRNA_D-pep@VLP; then, siRNA_D-pep@VLP was mixed with targeting peptide SP5-2 at a molar ratio of 1:1 to 8, and then EDC and NHS solutions were added, both with a final concentration of 3 to 5 mM, and reacted at 3 to 5°C for 6 to 10 hours; finally, the obtained solution was added to a 100 kDa dialysis tube, and the solution was continuously stirred and dialyzed in a PBS buffer for 10 to 15 hours, and the process was repeated 3 to 5 times to remove free SP5-2 and other byproducts, and finally a targeted delivery carrier solution was obtained in the dialysis tube, which was recorded as siRNA_D-pep@VLP-SP5-2 solution.

2. A targeted delivery vector based on human papillomavirus-like particles according to claim 1, characterized in that: The binding buffer contained 40 mM H2PO4 - / HPO4 2- , 100 mM NaCl and 0.1 mM dithiothreitol, pH=7 aqueous solution.

3. A targeted delivery vector based on human papillomavirus-like particles according to claim 1, characterized in that: The assembly solution contains 50mM H2PO4 - / HPO4 2- and an aqueous solution of 1.5 M NaCl, pH = 6.

0.

4. Use of a targeted delivery carrier based on human papillomavirus-like particles according to any one of claims 1 to 3 in the preparation of anti-tumor drugs.

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