A targeted delivery vector based on human papillomavirus-like particles and its application in the preparation of anti-tumor drugs

Abstract

A targeted delivery vector based on human papillomavirus-like particles and its application in the preparation of anti-tumor drugs belong to the field of targeted delivery vector technology. The targeted delivery vector siRNA_D-pep@VLP-SP5-2 prepared by the present invention simultaneously encapsulates two different types of anti-tumor drugs, namely Bcl-2siRNA and anti-tumor peptide D-pep, and has the advantages of high encapsulation rate, good targeting, easy synthesis, strong stability, and good biocompatibility; compared with siRNA_D-pep, the total cell apoptosis rate is increased from 60% to 91%; compared with siRNA_D-pep@VLP, the anti-tumor inhibition rate is increased by about 40%. This targeted delivery vector can target non-small cell lung cancer cell lines and has excellent ability to promote tumor cell apoptosis; through the synergistic effect of the two drugs, it efficiently induces apoptosis of non-small cell lung cancer cells, realizes targeted therapy, and can be used to prepare 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 the 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, nearly 20 million new cancer cases and 10 million cancer deaths will occur worldwide in 2022, posing a serious threat to global health and life. Therefore, the search for effective anti-tumor therapies is a global consensus. Cancer treatment typically involves systemic administration of drugs, such as chemotherapy and peptides with anti-tumor activity (such as D-LAK-120A and D-pep). However, many cancer patients are ineffective with chemotherapy or their efficacy decreases over time, in part because cancer cells develop resistance to apoptosis. Evasion of apoptosis occurs through multiple mechanisms, including enhanced DNA repair by non-homologous end-linking proteins or overexpression of anti-apoptotic proteins, which can act individually or synergistically. A major anti-apoptotic factor is the B-cell lymphoma 2 (Bcl-2) family of proteins, discovered nearly 30 years ago in non-Hodgkin's lymphoma. Bcl-2 is located in the mitochondrial and endoplasmic reticulum membranes and blocks the release of apoptosis-inducing factors and cytochrome c, inhibiting caspase-mediated apoptosis. Due to the crucial role of Bcl-2 in the apoptotic pathway, several therapeutic strategies have been developed to inhibit or downregulate Bcl-2 protein. RNA interference (RNAi), a novel cancer treatment strategy, utilizes small interfering RNA (siRNA) to mediate gene silencing in cells. siRNAs used to silence Bcl-2 have successfully reduced Bcl-2 expression in various tumors. However, siRNAs themselves have drawbacks such as susceptibility to nuclease degradation, easy clearance, difficulty crossing cell membranes, poor targeting, immunogenicity, and off-target effects.

[0003] Currently, one solution for efficiently delivering RNA to target cells is to use virus-like particles (VLPs) as targeted delivery vehicles. VLPs are engineered nanoparticles that mimic the characteristics of viruses and have strong resistance to heat and proteases, but lack viral genomes and infectivity. VLPs are composed of multi-subunit structural proteins, have the ability to spontaneously self-assemble, 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; however, siRNA itself has disadvantages such as easy degradation, difficulty in passing through the cell membrane, and poor targeting, requiring a reliable targeted delivery carrier to achieve its therapeutic function on target cells. The present invention utilizes the large cavity (diameter of approximately 35 nm) possessed by HPV VLP to simultaneously encapsulate two different anti-tumor drugs, D-pep and Bcl-2 siRNA, 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- , 100mM NaCl and 0.1mM dithiothreitol, pH = 7 aqueous solution) were co-incubated 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 mixed in the assembly solution (containing 50mM H2PO4) at a siRNA:HPV L1 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 in a molar ratio of 1:1 to 8, and then newly configured EDC and NHS solutions were added to a final concentration of 3 to 5 mM, and the mixture was reacted at 3 to 5°C for 6 to 10 hours; finally, the obtained solution was added to a dialysis tube (100 kDa) and dialyzed in PBS buffer with continuous stirring for 10 to 15 hours, and this was repeated 3 to 5 times to remove free SP5-2 and other byproducts, thereby obtaining a targeted delivery vector (siRNA_D-pep@VLP-SP5-2) solution in the dialysis tube, which can be used to target non-small cell lung cancer A549 cell lines.

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

[0008] The human papillomavirus-like particle (HPV VLP)-based targeted delivery vehicle (siRNA_D-pep@VLP-SP5-2) prepared by the present invention simultaneously encapsulates two different types of anti-tumor drugs (Bcl-2 siRNA and anti-tumor peptide D-pep). This delivery vehicle has the advantages of a high encapsulation efficiency (71%), good targeting, easy synthesis, strong stability, and good biocompatibility. siRNA_D-pep@VLP-SP5-2 has a strong ability to promote apoptosis in A549 cells. In vitro results showed that the total apoptosis rate of siRNA_D-pep@VLP-SP5-2 increased from 60% to 91% compared with siRNA_D-pep. In vivo anti-tumor results in mice showed that siRNA_D-pep@VLP-SP5-2 has a stronger tumor targeting ability and better therapeutic effect, with an anti-tumor inhibition rate increased by approximately 40% compared with 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, 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 exhibit FITC emission peaks at 510-520 nm, proving that FITC-SP5-2 was successfully modified onto 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, with invasion times of 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 significantly increased; among them, VLP-FITC-SP5-2 showed a higher internalization rate, and compared with FITC-labeled VLP alone, VLP-FITC-SP5-2 entered A549 cells in a shorter time (1 hour).

[0011] Figure 3: The binding affinity diagram of siRNA and D-pep at different amino / phosphate group 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 group 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 the amount of D-pep in the complex also increased with the increase of the ratio; when the different amino / phosphate group ratios increased from 1.5:1 to 3:1, the siRNA band did not change significantly, indicating that the amount of D-pep had reached saturation.

[0012] Figure 4 : Figure 4 A is a transmission electron microscopy photograph and average particle size diagram of the siRNA_D-pep complex obtained with an amino group / phosphate group ratio of 1.5:1, showing an average particle size of 12.6 ± 3.3 nm; 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. It can be seen 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 a transmission electron microscopy photograph 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±9 nm;

[0013] Figure 5 : Figure 5 A is the fluorescence spectra of different D-pep-FITC concentrations (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 with D-pep-FITC instead of D-pep. The curves from small to large D-pep-FITC concentration 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 of D-pep-FITC (x) and the fluorescence intensity at 520 nm (y), the standard equation is y = 34.5x–21.68, R 2=0.99. Using this standard equation, the actual D-pep-FITC concentration in siRNA_D-pep-FITC@VLP-SP5-2 was calculated to be 7.4 μg / mL, with an actual volume of 1 mL. Encapsulation efficiency (%) = Actual D-pep-FITC concentration × Actual volume / Added D-pep-FITC concentration × Added volume, resulting in an encapsulation efficiency of approximately 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 Annexin V-FITC (AV) and propidium iodide (PI), respectively, and then the cell apoptosis rates in 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 functional modification of VLP 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 led to 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 the 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 Aiboshi 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 Beyotime 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] Unless otherwise specified, 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. The mixture was reacted at 4°C on a rocking table for 8 hours. The obtained VLP-FITC-SP5-2 was added to a dialysis tube (100 kDa) and dialyzed in PBS buffer with continuous stirring for 12 hours. This was repeated 3 times to remove free SP5-2 and other byproducts. 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 488 nm excitation light, while VLP-FITC-SP5-2 and FITC-SP5-2 both have emission peaks at 510-520 nm, 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 hours, and then washed three times with PBS solution. 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 showed that VLP-FITC-SP5-2 showed a higher internalization rate. Compared with FITC-labeled VLP alone, VLP-FITC-SP5-2 entered A549 cells in a shorter time (1 hour), and the cells had stronger fluorescence after 2 hours of co-incubation, proving that SP5-2 improved 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 linked together through an amide reaction mediated by NHS and EDC. In order to prove that the amide reaction can link 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- , 100mM NaCl, 0.1mM dithiothreitol, pH = 7) and incubated for 30 minutes to assemble the siRNA_D-pep complex; then the HPV L1 pentamer and siRNA_D-pep complex were added in the assembly solution (50mM H2PO4 - / HPO4 2-, 1.5M NaCl, pH = 6.0) to co-assemble 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 resulting solution was added to a dialysis tube (100kDa) and dialyzed in PBS buffer with continuous stirring for 12 hours. This was repeated three times to remove free SP5-2 and other byproducts, and the siRNA_D-pep@VLP-SP5-2 solution was obtained in the dialysis tube. The results are shown in FIG. 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 backward. When the N / P ratio continues to increase, the siRNA band position no longer shifts, 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 an N / P ratio of 1.5:1 was approximately 12.6±3.3 nm, and the surface charge was negative. The particle size of the assembled siRNA_D-pep@VLP-SP5-2 was approximately 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 in A549 cells:

[0030] siRNA_D-pep@VLP-SP5-2 was assembled from 3 μg of Bcl-2 siRNA and 4.1 μg of D-pep (nitrogen-to-phosphorus ratio of 1.5:1); 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 apoptosis by flow cytometry using an apoptosis kit. The results are shown in Figure 5. Figure 6 As shown, Figure 6A shows that after 12 hours of co-incubation with A549 cells, there was no significant difference in the cell 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 and exert its intended function. In contrast, siRNA_D-pep@VLP, formed after VLP encapsulation of siRNA_D-pep, was observed to significantly increase the cell apoptosis rate, from a total apoptosis rate of 60% to approximately 80%, demonstrating 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 cell apoptosis rate was further increased, from a total apoptosis rate of 80% to approximately 91%, demonstrating that modification of the targeting peptide is also essential. In summary, the total cell apoptosis rate of siRNA_D-pep@VLP-SP5-2 increased from 60% to 91% compared to siRNA_D-pep. Figure 6 Figure 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 the A549 cell line.

[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 When the tumor grows to 40-50 mm, 3 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. 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%, while 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 siRNA_D-pep@VLP-SP5-2 had the best therapeutic effect and increased the tumor inhibition rate by about 40% compared with siRNA_D-pep. 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 it would not accumulate at the tumor site, while siRNA_D-pep@VLP-SP5-2 completely accumulated at the tumor site (e.g. Figure 7 Figure E. Where I: PBS; II: siRNA_D-pep@VLP-SP5-2; III: siRNA_D-pep@VLP). This demonstrates that siRNA_D-pep@VLP-SP5-2 still has good targeting in mice.

[0034] The above examples demonstrate the successful construction of a co-targeted delivery vector for siRNA and D-pep, demonstrating excellent encapsulation efficiency. This targeted delivery vector is capable of targeting the A549 cell line, improving targeted delivery efficiency. It effectively promotes tumor cell apoptosis both in vitro and in vivo, exhibiting promising anti-tumor therapeutic effects, and therefore can be used in the preparation of 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 technical personnel in this field may make improvements or changes based on some of the above descriptions, but all such 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 binding buffer at a nitrogen-phosphorus ratio of 0.75-3:1 for 20-40 minutes to form a complex siRNA_D-pep; then HPV L1 was added at a mass ratio of 0.3-0.8:

1. The L1 pentamer and siRNA_D-pep complex were co-assembled in the assembly solution to obtain siRNA_D-pep@VLP. The siRNA_D-pep@VLP was then mixed with the targeting peptide SP5-2 at a molar ratio of 1:1-8, and then EDC and NHS solutions were added to a final concentration of 3-5 mM. The mixture was reacted at 3-5°C for 6-10 hours. The resulting solution was added to a 100 kDa dialysis tube and dialyzed against PBS buffer with continuous stirring for 10-15 hours. This was repeated 3-5 times to remove free SP5-2 and other byproducts. Finally, a targeted delivery carrier solution, designated as siRNA_D-pep@VLP-SP5-2 solution, was obtained in the dialysis tube. D-pep is the anti-tumor peptide D-LAK-120A.

2. The human papillomavirus-like particle-based targeted delivery vector according to claim 1, wherein: The binding buffer contains 40 mM H2PO4 - / HPO4 2- , 100 mM NaCl and 0.1 mM dithiothreitol, pH = 7 aqueous solution.

3. The 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 1.5 M NaCl, pH = 6.0 aqueous solution.

4. Use of a targeted delivery vector 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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