Carrier for promoting Fenton reaction as well as preparation method and application thereof

Abstract

The invention provides a carrier for promoting Fenton reaction as well as a preparation method and application thereof. The carrier for promoting the Fenton reaction achieves the effects of improving gene delivery efficiency and cancer gene therapy by means of a plurality of efficacy experiments.

Description

Technical Field

[0001] The present invention relates to a carrier for promoting Fenton reaction and a preparation method and application thereof. Background Art

[0002] In recent years, gene therapy has been considered as a potential treatment for diseases. There are two ways of gene delivery: viral and non-viral. Viral vectors are used for gene delivery because of their high transfection efficiency, but their toxicity and immunogenicity limit their clinical application. Therefore, many non-viral forms of gene delivery have been studied to solve their limitations. However, most of these vectors face the problem of low transfection efficiency. Therefore, it is desirable to develop gene delivery vectors that can improve transfection efficiency.

[0003] In order to solve the above problems, technicians in this field are in urgent need of developing novel vectors that improve gene delivery efficiency and preparation methods and uses thereof to benefit the vast population with such needs. Summary of the invention

[0004] In view of this, an object of the present invention is to provide a carrier for promoting Fenton reaction, comprising a Prussian blue nanocube and a lipid component.

[0005] In one embodiment of the present invention, the lipid component encapsulates the Prussian blue nanocubes.

[0006] In one embodiment of the present invention, the lipid component comprises 1,2-dioleoyl-3-trimethylammonium-propane (DOTAP), methoxy-polyethylene-aldehyde (mPEG-CHO) and cholesterol.

[0007] In one embodiment of the present invention, the DOTAP is a cationic lipid component.

[0008] In one embodiment of the present invention, the molar ratio of the DOTAP, the mPEG-CHO and the cholesterol is 5:1:4.

[0009] In one embodiment of the present invention, the ratio of the mPEG-CHO to the carrier is 5 wt % to 40 wt %.

[0010] In one embodiment of the present invention, the ratio of the DOTAP to the carrier is 30 wt % to 50 wt %.

[0011] In one embodiment of the present invention, the ratio of the cholesterol to the carrier is 20 wt % to 35 wt %.

[0012] In one embodiment of the present invention, the ratio of the Prussian blue nanocubes to the carrier is 1 wt % to 10 wt %.

[0013] Another object of the present invention is to provide a method for preparing a carrier as described above, comprising the following steps: (a) dissolving polyvinylpyrrolidone (PVP) and potassium hexacyanoferrate (III) (K3[Fe(CN)6]) in deionized water to obtain a yellow solution; (b) adding hydrochloric acid to the yellow solution, stirring at room temperature, and then heating to obtain a blue solution; centrifuging the blue solution and washing with alcohol to collect the precipitate, then resuspending the precipitate in alcohol and drying to obtain the Prussian blue nanocube; and (d) dissolving the lipid component in alcohol, then adding the Prussian blue nanocube and evaporating the alcohol to obtain the carrier.

[0014] In one embodiment of the present invention, in step (d), after the alcohol is evaporated, a thin lipid film is formed, and then phosphate buffered saline (PBS) is added, and the thin lipid film is subjected to ultrasonic hydration.

[0015] Another object of the present invention is to provide a use of the aforementioned vector for improving gene delivery efficiency.

[0016] Another object of the present invention is to provide a use of the aforementioned vector for preparing a pharmaceutical composition for gene therapy of cancer.

[0017] In one embodiment of the present invention, the cancer is lung cancer.

[0018] Another object of the present invention is to provide a use of the aforementioned vector as a lung target vector.

[0019] In summary, the efficacy of the carrier for promoting Fenton reaction of the present invention is that the structure of cationic lipids is similar to that of natural lipids, and the positively charged polar head can bind to nucleic acids to stabilize nucleic acids and increase the ability of nucleic acids to be degraded. In addition, Prussian blue nanocubes are added to the lipid components to increase gene transfection and delivery efficiency by generating the ability of Fenton reaction, and have the potential to treat cancer through gene therapy.

[0020] The following will further illustrate the implementation mode of the present invention. The following examples are used to illustrate the present invention, not to limit the scope of the present invention. Anyone skilled in the art may make some changes and modifications without departing from the spirit and scope of the present invention. Therefore, the scope of protection of the present invention shall be determined according to the definition of the claims of this application. BRIEF DESCRIPTION OF THE DRAWINGS

[0021] Figure 1A and Figure 1B Characterization of the carrier (PB@LP) that promotes the Fenton reaction: Figure 1A )(a1) Prussian blue nanocube (PB); (a2) PB@lipidparticle (LP); (b1) iron oxide nanoparticles (IO) and (b2) lipid-modified iron oxide (IO@LP) transmission electron microscope (TEM) images; ( Figure 1B )Size distribution and zeta potential of PB, PB@LP, IO, IO@LP and 1,2-dioleoyl-3-trimethylammonium-propane (DOTAP). The DOTAP group contains DOTAP, mPEG-CHO and cholesterol as LP without PB or IO coating.

[0022] Figures 2A to 2D The PB@LP of the present invention generates reactive oxygen species (ROS) through the Fenton reaction. Figure 2A Schematic diagram of the ·OH catalytic process of PB in PB@LP; Figure 2B The UV-visible absorption spectra of PB@LP with different concentrations of PB after Fenton reaction of oxidized o-henylendiamine (ox OPD) are shown; Figure 2C UV-Vis absorption spectra and ( Figure 2D )image.

[0023] Figure 3AConfocal laser scanning microscope (CLSM) images showing green fluorescent protein (GFP) after different lipid-modified carriers were cultured with mouse skin melanoma B16F10 cells for 48 hours. (DAPI represents cell nuclei, GFP represents cells with green fluorescent protein. Scale bar: 100 μm; Merge represents merge; LNP represents a comparison group without PB or IO coating)

[0024] Figure 3B The transfection efficiency of the cationic lipid-only group, IO@LP, and PB@LP is shown.

[0025] Figure 4A and 4B After different lipid-modified carriers were loaded with programmed cell death 1 ligand 1 (PD-L1) siRNA and cultured with mouse skin melanoma B16F10 cells for 48 hours, the expression of PD-L1 protein on B16F10 cells was detected by (4A) conjugate microscopic image and (4B) flow cytometry. (DAPI represents the cell nucleus, PD-L1 represents the expression of programmed cell death 1 ligand 1 protein. Scale bar: 60μm; LNP represents the comparison group)

[0026] Figure 5 The accumulation of lipid-modified carriers PB@LP with different proportions of polyethylene glycol (PEG) in the lungs was detected using a non-invasive 3D in vivo molecular imaging (IVIS) system.

[0027] Figure 6 Fluorescence staining CLSM images showing the entry of the vector into the tumor after injection of PB@LP and IO@LP through the tail vein in an animal model of lung metastasis. (DAPI represents the cell nucleus; B16F10-GFP represents B16F10 cells with green fluorescent protein; DiI represents lipid carriers labeled with DiI dye; CD31 represents vascular endothelial cells. Scale bar: 20μm)

[0028] Figure 7 The figure shows that GFP mRNA was loaded onto different lipid-modified carriers and cultured with mouse skin melanoma B16F10 cells for 24 hours to observe the expression of green fluorescent protein (GFP). LNP represents the comparison group and Merge represents the merged group. DETAILED DESCRIPTION

[0029] The numerical values ​​used in this article are approximate values, and all experimental data are expressed in the range of ±20%, preferably in the range of ±10%, and most preferably in the range of ±5%.

[0030] Unless otherwise specified herein, the terms "a", "an", "the" and similar terms used in this specification (especially in the scope of the following patent application) should be understood to include both singular and plural forms.

[0031] According to the present invention, the term "Fenton reaction" is an inorganic chemical reaction in which hydrogen peroxide (H2O2) reacts with divalent iron ions Fe 2+ The mixed solution oxidizes many known organic compounds such as carboxylic acids, alcohols, and esters into inorganic forms. Cancer cells are killed by the production of reactive oxygen species (ROS) through the Fenton reaction.

[0032] According to the present invention, the chemical name of the term "Prussian blue" is ferric ferrocyanide; molecular formula: Fe7(CN) 18 14H2O is a dark blue pigment.

[0033] According to the present invention, the term "1,2-dioleoyl-3-trimethylammonium-propane (DOTAP)" is a cationic lipid.

[0034] According to the present invention, the molecular formula of the term "methoxy-polyethylene-aldehyde (mPEG-CHO)" is CH3O(CH2CH2O) n CH2CONHCH2CH2CHO needs to be stored at -20℃, in dry conditions, and protected by inert gas.

[0035] According to the present invention, the pharmaceutical composition can be manufactured into a dosage form suitable for parenteral or oral administration using techniques well known to those skilled in the art, including, but not limited to: injections [e.g., sterile aqueous solutions or dispersions], sterile powders, tablets, troches, lozenges, pills, capsules, dispersible powders or granules, solutions, suspensions, emulsions, syrups, elixirs, slurries and the like.

[0036] The pharmaceutical composition according to the present invention can be administered via a parenteral route selected from the group consisting of: intraperitoneal injection, subcutaneous injection, intraepidermal injection, intradermal injection, intramuscular injection, intravenous injection, intralesional injection, sublingual administration and transdermal administration.

[0037] The pharmaceutical composition according to the present invention may include a pharmaceutically acceptable carrier widely used in drug manufacturing technology. For example, the pharmaceutically acceptable carrier may include one or more agents selected from the group consisting of: solvent, emulsifier, suspending agent, decomposer, binder, excipient, stabilizer, chelating agent, diluent, gelling agent, preservative, lubricant, absorption delaying agent, liposome and the like. The selection and amount of these agents fall within the professional qualities and routine technical scope of those skilled in the art.

[0038] According to the present invention, the pharmaceutically acceptable carrier comprises a solvent selected from the group consisting of: water, normal saline, phosphate buffered saline (PBS), sugar solution, aqueous solution containing alcohol, and combinations thereof.

[0039] The present invention is further illustrated by the following examples. These examples are provided for illustration only and are not intended to limit the scope of the present invention. The scope of the present invention is as shown in the claims.

[0040] Example 1. Preparation of Prussian blue nanocubes and iron oxide nanoparticles (IO)

[0041] 3g of polyvinylpyrrolidone (PVP) (a polymer compound generated by the polymerization of N-vinyl-2-pyrrolidone) (K30) and 226.7mg of potassium hexacyanoferrate (III) (K3[Fe(CN)6]) were dissolved in 40mL of deionized water and stirred under vigorous magnetic stirring to form a clear yellow solution. 35μL of concentrated hydrochloric acid (HCl, 12M) was added, and after stirring at room temperature for 30 minutes, the solution was placed in an oven at 85℃ for 20 hours to obtain a dark blue Prussian blue nanocube solution. The solution was centrifuged (12,000rpm, 10 minutes) and washed three times with 75% ethanol to collect the precipitate. The precipitate was resuspended in 3-5mL of 75% ethanol and dried at 60℃ overnight to obtain Prussian blue nanocube single crystals. The iron oxide in the comparison group was commercial iron oxide with a size of about 5 nanometers and surface modified with polyethylene.

[0042] Example 2. Preparation of carriers (lipid-modified Prussian blue nanocubes, PB@LP) and lipid-modified iron oxide (IO@LP) that promote Fenton reaction

[0043] To obtain PB@LP, lipid components (including 1,2-dioleoyl-3-trimethylammonium-propane (DOTAP), methoxy-polyethylene-aldehyde (mPEG-CHO) and cholesterol) (molar ratio of 5:1:4) were dissolved in ethanol, and 1-10% Prussian blue nanocubes or iron (IO) were added. The solution was allowed to evaporate the ethanol at room temperature to form a thin lipid film. Then, phosphate buffered saline (PBS) with pH 7.4 was added, and the lipid film was hydrated by ultrasound to obtain 5mM PB@LP. The prepared PB@LP was centrifuged at 12,000 rpm for 10 minutes to remove the carrier without Prussian blue nanocubes. In the present invention, the ratio of mPEG-CHO is greatly increased to 5wt% to 40wt%, which is different from the ratio of 1% to 2% in the prior art. In addition, the use of mPEG-CHO can effectively provide a tumor penetration effect, wherein the ratio of mPEG-CHO can also be adjusted from 5wt% to 40wt%, all of which have a corresponding gene delivery effect.

[0044] Table 1 shows the ratio of each component of PB@LP and IO@LP.

[0045] Table 1

[0046]

[0047]

[0048] Figure 1A and Figure 1B Characterization of the carrier (PB@LP) that promotes the Fenton reaction: Figure 1A )(a1) Prussian blue nanocube (PB); (a2) PB@lipidparticle (LP); (b1) iron oxide nanoparticles (IO) and (b2) lipid-modified iron oxide (IO@LP) transmission electron microscope (TEM) images. Figure 1B )Size distribution and zeta potential of PB, PB@LP, IO, IO@LP and 1,2-dioleoyl-3-trimethylammonium-propane (DOTAP). The DOTAP group contains DOTAP, mPEG-CHO and cholesterol as LP without PB or IO coating.

[0049] To confirm whether the Prussian blue nanocubes and iron oxide nanoparticles coated with cationic lipids (including DOTAP, mPEG-CHO and cholesterol) were successfully synthesized, transmission electron microscopy (TEM) and dynamic light scattering (DLS) were used to detect the formation of particles. TEM imaging results showed that the Prussian blue nanocubes were cubic in shape, the Fe2O3 morphology was relatively rough, and the PB@LP and IO@LP morphologies were round, and their tetragonal structures were visible ( Figure 1A In addition, DLS showed that the cationic lipid nanoparticles were originally about 100 nanometers, and after being coated with Prussian blue nanocubes or Fe2O3, the size was about 500 nanometers. The surface charge of Prussian blue nanocubes and Fe2O3 was originally negative, but after being coated with cationic lipids, the surface charge became positive ( Figure 1B ), from the above results we can see that PB@LP and IO@LP are successfully synthesized.

[0050] Example 3. Evaluation of the ability of PB@LP to produce reactive oxygen species (ROS)

[0051] Figures 2A to 2DIt is shown that PB@LP generates reactive oxygen species (ROS) through the Fenton reaction. Figure 2A Schematic diagram of the OH catalysis process of PB in PB@LP. Figure 2B The UV-Vis absorption spectra of PB@LP with different concentrations of PB after Fenton reaction of oxidized o-phenylenediamine (ox OPD). The UV-Vis absorption spectra of oxidized o-phenylenediamine (ox OPD) after adding (1) PBS; (2) PBS+H2O2; (3) Fe2O3+H2O2; (4) PB+H2O2 and ( Figure 2D )image.

[0052] The PB@LP PB undergoes a Fenton reaction to catalyze hydrogen peroxide (H2O2) to produce hydroxyl radicals (-OH), which oxidize the colorless o-phenylendiamine (OPD) into a yellow dimer ( Figure 2A ), the amount of dimer generated was detected by ultraviolet-visible spectroscopy (UV-Vis) to evaluate the ability of PB@LP to produce reactive oxygen species (ROS). In this part of the experiment, different concentrations of PB@LP were added to 0.5mM OPD and 2.5mM H2O2, and after waiting for 20 minutes, the absorbance at 417nm was measured using UV-Vis. It can be seen that the higher the concentration of PB@LP, the higher the absorbance, indicating that PB@LP can produce ROS through the Fenton reaction ( Figure 2B ). In addition, to compare the ability of PB@LP to produce ROS through the Fenton reaction, iron oxide nanoparticles, which also contain iron, were used for comparison. The results showed that compared with iron oxide nanoparticles, PB@LP has a better ability to produce ROS ( Figure 2C , 2D ).

[0053] Example 4. Evaluation of the efficacy of PB@LP in gene delivery

[0054] To evaluate the effect of gene delivery, plasmid DNA (pDNA) carrying the green fluorescent protein (GFP) gene was loaded into PB@LP. Mouse skin melanoma B16F10 cells (BCRC60031) were cultured at 2.5×10 per well. 4The cells were seeded at a density of 100 cells / mL in a 24-well cell culture dish. After cell attachment, the cells were cultured with PB@LP loaded with pDNA-GFP at a concentration of 10 μg / mL in serum-free medium. After 6 hours, 10% fetal bovine serum (FBS) medium was added to each well to maintain cell growth for 48 hours. The expression of GFP after transfection was examined using a conjugate laser scanning microscope (CLSM).

[0055] Figure 3A The confocal laser scanning microscope (CLSM) images of green fluorescent protein (GFP) after culturing different lipid-modified carriers with mouse skin melanoma B16F10 cells for 48 hours are shown. (DAPI represents cell nuclei, GFP represents cells with green fluorescent protein. Scale bar: 100 μm; Merge represents merge; LNP represents a comparison group without PB or IO coating)

[0056] like Figure 3A and Figure 3B As shown, the transfection efficiency of the group with only cationic lipids was about 16%, IO@LP was about 28%, and PB@LP was about 41%. The results show that compared with IO@LP coated with the same iron element, the transfection efficiency of PB@LP, which can produce more ROS, is 13% higher (Figure 3).

[0057] Example 5. Evaluation of the efficacy of PB@LP in inhibiting the expression of programmed cell death 1 ligand 1 (PD-L1) protein in cancer cells

[0058] If PD-L1 siRNA effectively enters cells and exerts its effect, the PD-L1 protein on the surface of cancer cells will decrease. In order to evaluate the effect of siRNA delivery, PD-L1 siRNA was first loaded into PB@LP, then added to the cell culture medium and cultured with B16F10 cells for 48 hours, and finally immunohistochemical staining and flow cytometry were used to detect the expression of PD-L1.

[0059] Figure 4A and 4BAfter different lipid-modified carriers were loaded with programmed cell death 1 ligand 1 (PD-L1) siRNA and cultured with B16F10 cells for 48 hours, the expression of PD-L1 protein on B16F10 cells was detected by (4A) conjugate microscopy and (4B) flow cytometry. (DAPI represents the cell nucleus, PD-L1 represents the expression of programmed cell death 1 ligand 1 protein. Scale bar: 60μm; LNP represents the comparison group)

[0060] like Figure 4A and 4B As shown in the figure, the lipid carrier loaded with PD-L1 siRNA showed lower PD-L1 expression than the control group. According to the results of flow cytometry, compared with the control group, the PD-L1 expression of the LNP group decreased by nearly 40%, while the PD-L1 expression of the IO@LP group decreased by 64.2%, but the PB@LP group showed almost no decrease ( Figure 4B ). This shows that the lipid carrier PB@LP loaded with PD-L1 siRNA can effectively inhibit the expression of PD-L1 protein on cancer cells, and may be able to increase the activity of T cells in immunotherapy in the future.

[0061] Example 6. Evaluation of the efficacy of PB@LP in gene therapy of in vivo organ cancer

[0062] In order to evaluate whether the PB@LP lipid carrier can successfully stay in the lungs for the subsequent treatment of lung metastasis cancer, lipid carriers stained with DiI (a cyan dye used in cell biology) were injected into the tail vein of an animal model of lung metastasis cancer to observe whether the lipid carriers can enter the lungs. On day 0, 5×10 B16F10 melanoma cells carrying GFP were inoculated into the lungs. 6 The concentration of cells / mL was injected into C57BL / 6 mice through the tail vein. On the 13th day, 100μL of 5mM lipid carrier was injected, and the mice were sacrificed by perfusion 24 hours later. The heart, liver, spleen, lung, kidney and other major organs of the mice were removed and non-invasive 3D in vivo molecular imaging (IVIS) was performed.

[0063] Figure 5 The results show that the accumulation of lipid-modified vectors PB@LP with different proportions of polyethylene glycol (PEG) (i.e. mPEG-CHO) in the lungs was detected using a non-invasive 3D in vivo molecular imaging (IVIS) system. The results showed that the transfection rate of lipid-modified vectors PB@LP increased by 50% to 80%, and as the concentration of PEG increased, the accumulation in the lungs also increased ( Figure 5), but no significant accumulation was observed in cancers of other organs.

[0064] Example 7. Evaluation of the efficacy of PB@LP in in vivo cancer gene therapy

[0065] In order to evaluate whether PB@LP lipid carriers can be used for subsequent in vivo cancer gene therapy, DiI-stained PB@LP and IO@LP were injected into the tail vein of an animal model of lung metastasis cancer to observe whether PB@LP and IO@LP could enter the tumor. On day 0, GFP-carrying B16F10 melanoma cells were inoculated at 1×10 6 The concentration of 100 μL of 5 mM lipid carrier was injected into C57BL / 6 mice through the tail vein. On the 13th day, 100 μL of 5 mM lipid carrier was injected, and the mice were sacrificed by perfusion 24 hours later. The lungs of the mice were frozen and sectioned, and immunohistochemical staining was performed to observe the accumulation of PB@LP and IO@LP in the lungs using confocal microscopy.

[0066] Figure 6 Fluorescence staining CLSM images showing the entry of the vector into the tumor after injection of PB@LP and IO@LP through the tail vein in an animal model of lung metastasis. (DAPI represents the cell nucleus; B16F10-GFP represents B16F10 cells with green fluorescent protein; DiI represents lipid carriers labeled with DiI dye; CD31 represents vascular endothelial cells. Scale bar: 20μm)

[0067] like Figure 6 As shown, there is accumulation of DiI-labeled PB@LP and IO@LP in the tumor area with green fluorescence, indicating that PB@LP can enter the tumor through the blood circulation and is conducive to subsequent gene delivery therapy in the future ( Figure 6 ).

[0068] Example 8. Application of PB@LP to other gene delivery systems

[0069] In order to test whether lipid carriers can be applied to other gene delivery systems, this example uses GFP mRNA to be loaded onto lipid carriers and cultured with B16F10 cells for 24 hours to express green fluorescent protein.

[0070] Figure 7 The results show that GFP mRNA was loaded onto different lipid-modified carriers and cultured with B16F10 cells for 24 hours to observe the expression of green fluorescent protein (GFP). LNP represents the comparison group and Merge represents the merger. The results show that lipid carriers can effectively deliver mRNA into cells and express their corresponding proteins. Therefore, PB@LP can effectively deliver mRNA ( Figure 7 ).

[0071] In summary, the carrier of the present invention that promotes the Fenton reaction does have the following effects: the structure of the cationic lipid is similar to that of natural lipid, and the positively charged polar head can bind to nucleic acid to stabilize the nucleic acid and increase the ability of nucleic acid to be degraded. In addition, Prussian blue nanocubes are added to the lipid component to increase the gene transfection and delivery efficiency by generating the Fenton reaction, and have the potential to treat cancer through gene therapy.

[0072] The above description is for illustrative purposes only and is not intended to be limiting. Any equivalent modifications or changes made thereto without departing from the spirit and scope of the present invention shall be included in the claims of the present invention.

[0073]

Explanation of symbols

[0074] none.

Claims

1. A carrier for promoting Fenton reaction, characterized in that: The invention comprises a Prussian blue nanocube and a lipid component.

2. The carrier according to claim 1, characterized in that The lipid component encapsulates the Prussian blue nanocubes.

3. The carrier according to claim 1, characterized in that The lipid component comprises 1,2-dioleoyl-3-trimethylammonium-propane (DOTAP), methoxy-polyethylene-aldehyde (mPEG-CHO) and cholesterol.

4. The carrier according to claim 3, characterized in that The DOTAP is a cationic lipid component.

5. The carrier according to claim 4, characterized in that The molar ratio of the DOTAP, the mPEG-CHO and the cholesterol is 5:1:

4.

6. The carrier according to claim 4, characterized in that The ratio of mPEG-CHO to the carrier is 5wt% to 40wt%.

7. The carrier according to claim 4, characterized in that The proportion of DOTAP in the carrier is 30 wt% to 50 wt%.

8. The carrier according to claim 4, characterized in that The proportion of cholesterol in the carrier is 20 wt% to 35 wt%.

9. The carrier according to claim 1, characterized in that The ratio of the Prussian blue nanocubes to the carrier is 1 wt% to 10 wt%.

10. A method for preparing the carrier according to any one of claims 1 to 9, characterized in that The following steps are involved: (a) polyvinylpyrrolidone (PVP) and potassium hexacyanoferrate (III) (K3[Fe(CN)6]) are dissolved in deionized water to obtain a yellow solution; (b) adding hydrochloric acid to the yellow solution, followed by stirring at room temperature, and then heating to obtain a blue solution; (c) centrifuging the blue solution and washing it with alcohol to collect a precipitate, and then resuspending the precipitate in alcohol and drying it to obtain the Prussian blue nanocubes; and (d) dissolving the lipid component in alcohol, then adding the Prussian blue nanocubes and evaporating the alcohol to obtain the carrier.

11. The method according to claim 10, characterized in that In step (d), after the alcohol is evaporated, a thin lipid film is formed, and then phosphate buffered saline (PBS) is added, and the thin lipid film is subjected to ultrasonic hydration treatment.

12. Use of the vector according to any one of claims 1 to 9 for improving gene delivery efficiency.

13. Use of the vector according to any one of claims 1 to 9 for preparing a pharmaceutical composition for gene therapy of cancer.

14. The use according to claim 13, characterized in that Wherein the cancer is lung cancer.

15. Use of the vector of any one of claims 1 to 9 as a lung-targeting vector.

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