Vanadium carbide-coated doxorubicin NANO preparation and use thereof

Abstract

The present invention relates to the technical field of nanomedicine, in particular to a vanadium carbide-coated doxorubicin nano preparation and the use thereof. The vanadium carbide-coated doxorubicin nano preparation comprises a vanadium carbide nanosheet and doxorubicin, wherein the doxorubicin is wrapped in the vanadium carbide nanosheet. The preparation method is method I or method II. Method I comprises the following steps: mixing a V2C nanosheet, doxorubicin and a PBS buffer to dissolve same, and performing a vortex ultrasonic treatment to obtain the vanadium carbide-coated doxorubicin nano preparation. Method II comprises the following steps: dissolving poloxamer in a PBS buffer to obtain a poloxamer solution; mixing a V2C nanosheet, doxorubicin and the poloxamer solution to dissolve same, and performing a vortex ultrasonic treatment to obtain the vanadium carbide-coated doxorubicin nano preparation. By means of wrapping doxorubicin in the two-dimensional nano material vanadium carbide, the lethality of doxorubicin against tumor cells is improved, and the toxicity of doxorubicin to normal cells is reduced.

Description

A vanadium carbide-coated doxorubicin nanoformulation and its application Technical Field

[0001] The present invention relates to the technical field of nanomedicine, in particular to a vanadium carbide-coated doxorubicin nano preparation and application thereof. Background Art

[0002] Cancer is one of the leading causes of death worldwide, with breast cancer believed to account for 30% of all estimated new cancer cases. Studies have shown that epigenetic changes in several genes involved in tumor cell proliferation, invasion, metastasis, and apoptosis can lead to breast cancer, and these changes are not accompanied by alterations in the gene sequence. Therefore, they are considered reversible changes in gene expression. Unfortunately, traditional cancer treatments, such as chemotherapy, immunotherapy, and radiotherapy, are cell-selective, with limited efficacy and increased risks to normal and healthy tissues.

[0003] Doxorubicin (Dox) is an anthracycline antibiotic used in a variety of malignant tumors. As a first-line anticancer therapy, Dox has demonstrated clinical activity in a variety of cancer types, including breast, ovarian, lung, and liver cancers, as well as Hodgkin and non-Hodgkin lymphomas. However, long-term use of Dox, an anticancer drug, can cause side effects such as cardiotoxicity. Therefore, there is a need for a novel drug delivery system that increases tumor cell lethality while minimizing cytotoxic side effects in healthy cells.

[0004] Summary of the Invention

[0005] Based on the above content, the present invention provides a vanadium carbide-encapsulated nanoformulation of doxorubicin and its application. By encapsulating doxorubicin in a two-dimensional nanomaterial vanadium carbide, the lethality of doxorubicin to tumor cells (such as human breast cancer cells) is increased and its toxicity to normal cells is reduced.

[0006] To achieve the above object, the present invention provides the following solutions:

[0007] One of the technical solutions of the present invention is a vanadium carbide-encapsulated doxorubicin nanoformulation, comprising vanadium carbide nanosheets and doxorubicin; the doxorubicin is encapsulated in the vanadium carbide nanosheets; and the mass ratio of the doxorubicin to the vanadium carbide nanosheets is 2.5:1.

[0008] The second technical solution of the present invention is a method for preparing the above-mentioned vanadium carbide-coated doxorubicin nanoformulation, the preparation method being method one or method two;

[0009] The method 1 comprises the following steps:

[0010] The V2C nanosheets are mixed with doxorubicin and PBS buffer, dissolved, vortexed, and sonicated to obtain the vanadium carbide-coated doxorubicin nanoformulation;

[0011] The second method comprises the following steps:

[0012] Dissolve poloxamer in PBS buffer to obtain a poloxamer solution;

[0013] The V2C nanosheets are mixed and dissolved with doxorubicin and poloxamer solution, and then vortexed and ultrasonicated to obtain the vanadium carbide-coated doxorubicin nanoformulation.

[0014] The third technical solution of the present invention is the use of the above-mentioned vanadium carbide-coated doxorubicin nanoformulation in the preparation of anti-tumor drugs.

[0015] The present invention discloses the following technical effects:

[0016] (1) Vanadium carbide (V2C) has excellent biocompatibility and can be biodegraded by enzymes. The two-dimensional nanomaterial vanadium carbide nanosheets constructed by the present invention encapsulate the nanoformulation of doxorubicin, thereby improving the utilization rate of doxorubicin; vanadium carbide as a photothermal agent realizes photothermal conversion at a near-infrared wavelength of 808nm, and at the same time promotes the accumulation and release of doxorubicin in tumor cells through passive targeting; the present invention utilizes two-dimensional nanomaterial vanadium carbide nanosheets to encapsulate doxorubicin, thereby improving the killing rate of doxorubicin on tumor cells;

[0017] (2) Doxorubicin is a broad-spectrum anticancer drug. The present invention prepares the nanoformulation of doxorubicin coated with vanadium carbide into a solution (Example 2) or a gel formulation (Example 3). The solution formulation can enter all parts of the body and has a sustained-release characteristic; while the gel formulation can be passively fixed to the tumor site (avoiding the diffusion of the drug to other parts). The solution formulation and gel formulation of vanadium carbide coated doxorubicin nanoformulation constructed by the present invention can, on the one hand, avoid the premature release of the drug, and on the other hand, increase the drug loading of doxorubicin, improve the bioavailability of doxorubicin and avoid the release of doxorubicin in healthy parts, providing a basis and direction for the application combination of anti-tumor drugs and nano-biomedicine.

[0018] (3) The preparation method of the vanadium carbide-coated doxorubicin nanoformulation of the present invention has the characteristics of simple process and high yield. The solvent of the final product is water, which is non-toxic, environmentally friendly, and can achieve low-cost large-scale production;

[0019] (4) The vanadium carbide-coated doxorubicin nanoformulation of the present invention has the characteristics of integration, non-invasiveness, targeting, biocompatibility, and near-infrared response. The photothermal effect therapy strategy involved in the present invention can achieve passive targeting and release of drugs, overcoming the disadvantage of tumor drugs being highly toxic to normal cells, while having good biocompatibility, providing a new method and idea for designing tumor drug delivery carriers;

[0020] (5) The nanoformulation of doxorubicin coated with the two-dimensional nanomaterial vanadium carbide provided by the present invention is expected to be used as a drug delivery strategy in the treatment of diseases. BRIEF DESCRIPTION OF THE DRAWINGS

[0021] In order to more clearly illustrate the embodiments of the present invention or the technical solutions in the prior art, the following briefly introduces the drawings required for use in the embodiments. Obviously, the drawings described below are only some embodiments of the present invention. For ordinary technicians in this field, other drawings can be obtained based on these drawings without paying any creative work.

[0022] Figure 1 is a characterization diagram of V2C nanosheets, V2C-Dox, and V2C-Dox@Gel prepared in Examples 1-3; wherein, A is the particle size of V2C and V2C-Dox; B is the zeta potential of V2C and V2C-Dox; C is the transmission electron microscope image of V2C; D is the HRTEM image of V2C; E is the HRTEM image of V2C-Dox; F is the EDS elemental mapping image of V2C; G is the transparency of the gel and V2C-Dox@Gel at different temperatures; H is the LCST estimation of the hydrogel based on the transparency difference in the temperature range; I is the spectrum of the G' and G" moduli of the hydrogel; J is the viscosity change of the gel at different scanning frequencies; K is the viscosity change of the V2C-Dox@Gel at different scanning frequencies; L is the dynamic strain scan of the gel and V2C-Dox@Gel; M is the SEM image of the gel; and N is the SEM image of V2C-Dox@Gel.

[0023] Figure 2 is a safety assessment diagram of the V2C nanosheets prepared in Example 1 and the V2C-Dox prepared in Example 2; wherein A is the red blood cell hemolysis test; B is the MDA-MB-231 cell activity; C is the MCF-7 cell activity; D is the blood glucose test result; E is the blood lipid test result; F is the ion test result; G is the liver function test result; and H is the routine blood test result.

[0024] FIG3 is a graph showing spontaneous fluorescence generated by V2C-Dox prepared in Example 2 entering cells.

[0025] Figure 4 shows the inhibition of cell growth by V2C nanosheets prepared in Example 1 and V2C-Dox prepared in Example 2 under the combined application of near-infrared laser; wherein A is the cell viability of MDA-MB-231 cells within a specified time; B is the colony formation efficiency; and C is the cell apoptosis rate.

[0026] Figure 5 shows the inhibition of cell metastasis by V2C nanosheets prepared in Example 1 and V2C-Dox prepared in Example 2 under the combined application of near-infrared laser; A is the scratch image of MDA-MB-231 cells; B is the migration rate; C is the cell migration image; and D is the cell migration number image.

[0027] Figure 6 shows the tumor inhibition effect of V2C nanosheets, V2C-Dox, and V2C-Dox@Gel prepared in Examples 1-3 in animals under the combined application of near-infrared laser; A is a tumor growth size curve; B is a tumor size graph; C is a Ki67 staining graph; and D is a Ki67 statistical graph. DETAILED DESCRIPTION

[0028] Various exemplary embodiments of the present invention will now be described in detail. This detailed description should not be considered as limiting the present invention, but rather as a more detailed description of certain aspects, features, and embodiments of the present invention.

[0029] It should be understood that the terms described herein are intended only to describe particular embodiments and are not intended to limit the present invention. In addition, for numerical ranges herein, it should be understood that each intermediate value between the upper and lower limits of the range is also specifically disclosed. The intermediate value within any stated value or stated range, and each smaller range between any other stated value or intermediate value within the stated range, is also encompassed within the present invention. The upper and lower limits of these smaller ranges may be independently included or excluded within the scope.

[0030] Unless otherwise indicated, all technical and scientific terms used herein have the same meaning as commonly understood by those skilled in the art. Although only preferred methods and materials are described herein, any methods and materials similar or equivalent to those described herein may also be used in the practice or testing of the present invention. All documents mentioned in this specification are incorporated by reference to disclose and describe the methods and / or materials associated with the documents. In the event of any conflict with any incorporated document, the contents of this specification shall prevail.

[0031] It will be apparent to those skilled in the art that various modifications and variations may be made to the specific embodiments described herein without departing from the scope or spirit of the invention. Other embodiments will be apparent to those skilled in the art from the description of the invention. The description and examples are intended to be exemplary only.

[0032] The words “include,” “including,” “have,” “contain,” etc. used in this document are open-ended terms, meaning including but not limited to.

[0033] Unless otherwise specified, the "%" in the present invention refers to mass percentage.

[0034] The "room temperature" mentioned in the present invention, unless otherwise specified, refers to 20-30°C.

[0035] A first aspect of the present invention provides a vanadium carbide-encapsulated doxorubicin nanoformulation, comprising vanadium carbide nanosheets and doxorubicin; the doxorubicin is encapsulated in the vanadium carbide nanosheets; and the mass ratio of the doxorubicin to the vanadium carbide nanosheets is 2.5:1.

[0036] When the mass ratio of the doxorubicin to the vanadium carbide nanosheets decreases, the drug loading rate of the doxorubicin is low; when the mass ratio increases, the carrier decreases, which affects the photothermal effect and drug efficacy.

[0037] In the present invention, the dosage form of the vanadium carbide-coated doxorubicin nanoformulation includes solution or gel.

[0038] The second aspect of the present invention provides a method for preparing the above-mentioned vanadium carbide-coated doxorubicin nanoformulation, which is method one or method two;

[0039] The method 1 comprises the following steps:

[0040] The V2C nanosheets are mixed with doxorubicin and PBS buffer, dissolved, vortexed, and sonicated to obtain the vanadium carbide-coated doxorubicin nanoformulation;

[0041] The second method comprises the following steps:

[0042] Dissolve poloxamer in PBS buffer to obtain a poloxamer solution;

[0043] The V2C nanosheets are mixed and dissolved with doxorubicin and poloxamer solution, and then vortexed and ultrasonicated to obtain the vanadium carbide-coated doxorubicin nanoformulation.

[0044] In the present invention, in the method 1, the mass volume ratio of the V2C nanosheets to doxorubicin and PBS buffer is 1 mg:2.5 mg:2 mL.

[0045] In the present invention, in the second method, the concentration of poloxamer in the poloxamer solution is 200 mg / mL; the mass volume ratio of the V2C nanosheets to doxorubicin and the poloxamer solution is 1 mg:2.5 mg:2 mL. The poloxamer is poloxamer 407.

[0046] In the present invention, in the method 1 and the method 2, the vortexing time is 2 minutes; the ultrasound is specifically ice bath ultrasound for 30 minutes; and the pH value of the PBS buffer is 7.2.

[0047] In the present invention, in the method 1 and the method 2, the method for preparing the V2C nanosheets comprises the following steps:

[0048] V2AlC is added to an HF solution for etching, followed by centrifugation. The resulting precipitate is mixed and stirred with a TPAOH solution, and then centrifuged for purification to obtain the V2C nanosheets.

[0049] In the present invention, the HF solution is a concentrated HF solution; the etching process is specifically stirred for 72 hours; after the centrifugation, the step of washing the obtained precipitate until the pH value of the supernatant is not less than 5.0 is further included; the temperature of the mixing and stirring is room temperature and the time is 24 hours; after the centrifugal purification, the steps of degassing with argon and rinsing with water are also included.

[0050] The dosage form of the vanadium carbide-coated doxorubicin nanoformulation obtained by the above method 1 is a solution, and the dosage form of the vanadium carbide-coated doxorubicin nanoformulation obtained by the above method 2 is a gel.

[0051] The third aspect of the present invention provides the use of the above-mentioned vanadium carbide-coated doxorubicin nanoformulation in the preparation of anti-tumor drugs.

[0052] In the present invention, the anti-tumor drug includes but is not limited to chemotherapy drugs, targeted therapy drugs, endocrine therapy drugs, and anti-tumor angiogenesis drugs, etc. In a more preferred embodiment of the present invention, the anti-tumor drug is an anti-breast cancer drug.

[0053] An important feature of effective drug delivery is the ability to avoid premature release of therapeutic drugs. Vanadium carbide (V2C) nanoparticles are a good light absorber with excellent photothermal conversion capabilities. V2C also has good biocompatibility and can be biodegraded by enzymes. V2C also has a large drug loading capacity. Therefore, the V2C encapsulation of Dox in the present invention can regulate the diffusion of the active ingredient of the Dox drug. Under near-infrared light radiation, the vanadium carbide-encapsulated doxorubicin nanoformulation can convert light into heat energy, achieve passive targeting of the drug to tumors and promote drug release, while having good biocompatibility.

[0054] The vanadium carbide-coated nanoformulation of doxorubicin of the present invention can avoid the premature release of the drug and improve the bioavailability of doxorubicin. The vanadium carbide-coated nanoformulation of doxorubicin can be used as a photothermal agent to achieve photothermal conversion under the near-infrared wavelength of 808nm, while achieving the accumulation and release of doxorubicin in tumors (such as breast cancer tumors), and the gel dosage form can avoid the release of doxorubicin in healthy tissues. The present invention uses nude mice as an animal model for in vivo evaluation to systematically evaluate the safety of the vanadium carbide-coated nanoformulation of doxorubicin. The results show that the vanadium carbide-coated nanoformulation of doxorubicin does not produce toxic side effects on cells under normal conditions, and effectively inhibits tumor growth and metastasis under the near-infrared wavelength of 808nm. The preparation and production process of the vanadium carbide-coated nanoformulation of doxorubicin provided by the present invention are simple, have high yield, strong effect, good safety, are environmentally friendly, and can achieve low-cost large-scale production.

[0055] Unless otherwise specified, the materials involved in the embodiments of the present invention can be obtained through commercial channels.

[0056] The V2AlC powder, HF solution, TPAOH (tetrapropylammonium hydroxide), and poloxamer 407 used in the examples of the present invention were all purchased from Sinopharm Chemical Reagent Co., Ltd.; doxorubicin was purchased from Aladdin Biotechnology Co., Ltd.; MCF-10A (human normal mammary epithelial cells), MCF-7 (human breast cancer cells), and MDA-MB-231 (human breast cancer cells) cells were all purchased from the American Type Culture Collection Catalogue.

[0057] The pH value of the PBS buffer used in the embodiment of the present invention is 7.2.

[0058] Example 1 Preparation of two-dimensional V2C nanosheets

[0059] One gram of V2AlC powder was slowly added to 30 mL of a 50% concentrated HF solution at room temperature while stirring with a magnetic bar for 72 hours. After removing the Al layer from the V2AlC by HF etching, the suspension was centrifuged at 2292 × g for 10 minutes and washed three times with distilled water. The resulting precipitate was centrifuged at 500 × g for 10 minutes until the pH of the supernatant reached above 5.0, resulting in a large precipitate. This was then stirred and mixed with 40 mL of an 80% TPAOH aqueous solution at room temperature for 24 hours. Finally, the V2C was purified by centrifugation, degassed with argon (Ar), rinsed three times with distilled water to remove residual TPAOH, and then centrifuged at 1467 × g for 50 minutes. The colloidal supernatant was collected to obtain a V2C nanosheet dispersion.

[0060] Example 2 Preparation of V2C-encapsulated doxorubicin nanoformulation

[0061] The V2C nanosheet dispersion obtained in Example 1 was first centrifuged to obtain V2C nanosheets, which were then washed three times with PBS buffer and diluted with PBS buffer to a V2C nanosheet concentration of 480 μg / mL to obtain a V2C nanosheet dispersion. Dox was dissolved in PBS buffer to a concentration of 1.2 mg / mL to obtain a Dox solution. 2 mL of the Dox solution was added to 2 mL of the V2C nanosheet dispersion, vortexed for 2 minutes, and sonicated in an ice bath for 30 minutes to obtain a V2C-encapsulated doxorubicin nanoformulation (V2C-Dox) in the form of a solution, which was then stored in a 4°C refrigerator for later use.

[0062] Example 3 Preparation of V2C-encapsulated doxorubicin nanoformulation

[0063] Weigh 2 grams of poloxamer 407, add it to 10 mL of PBS buffer, place it in the refrigerator to swell for 12 hours, and obtain a 200 mg / mL poloxamer 407 solution (i.e., blank gel Gel). The V2C nanosheet dispersion prepared in Example 1 is first centrifuged, washed three times with the above-mentioned poloxamer 407 solution, and then diluted with poloxamer 407 solution to obtain a V2C nanosheet concentration of 480 μg / mL to obtain a V2C nanosheet dispersion. Dox is dissolved in poloxamer 407 solution to a Dox solution with a concentration of 1.2 mg / mL. 2 mL of Dox solution is added to 2 mL of V2C nanosheet dispersion, vortexed for 2 minutes, and ultrasonicated in an ice bath for 30 minutes to obtain a V2C-encapsulated doxorubicin nanoformulation (V2C-Dox@Gel) in the form of a gel, which is placed in a 4°C refrigerator for later use.

[0064] Effect Example 1

[0065] Morphological characterization of V2C nanosheets (abbreviated as V2C), V2C-Dox, and V2C-Dox@Gel prepared in Examples 1-3 The particle size distribution and zeta potential were measured using a Zetasizer Nano ZS90 system (Malvern Instruments, UK). The morphology of freeze-dried blank gel Gel and V2C-Dox@Gel was observed using a scanning electron microscope (SEM). Transmission electron microscopy (TEM) images were obtained using a FEI Tecnai G2 instrument. High-resolution TEM (HRTEM) images were obtained using a FEI Tecnai G2 F30 instrument. Corresponding elemental mapping imaging was performed on V2C. The rheological properties of the gel (V2C-Dox@Gel) were analyzed using a rheometer. Shear thinning tests were performed on the blank gel Gel and V2C-Dox@Gel at a shear rate of 0.1-100s-1 and a strain range of 0.01-100%. The storage modulus (G') and loss modulus (G) of the hydrogel were detected. The results are shown in Figure 1.

[0066] In Figure 1, A is the particle size of V2C and V2C-Dox; B is the zeta potential of V2C and V2C-Dox; C is the transmission electron microscope image of V2C; D is the HRTEM image of V2C; E is the HRTEM image of V2C-Dox; F is the EDS elemental mapping image of V2C; G is the transparency of blank gel Gel and V2C-Dox@Gel at different temperatures; H is the LCST estimation of the hydrogel based on the transparency difference in the temperature range; I is the spectrum of the G' and G" moduli of the blank gel Gel; J is the viscosity change of the blank gel Gel at different scanning frequencies; K is the viscosity change of V2C-Dox@Gel at different scanning frequencies; L is the dynamic strain scan of the blank gel Gel and V2C-Dox@Gel; M is the SEM image of the blank gel Gel; N is the SEM image of V2C-Dox@Gel. As can be seen from Figure 1, the particle size of V2C and V2C-Dox was measured (A in Figure 1). and zeta potential (Figure 1B). Dynamic light scattering (DLS) experiments showed that the average fluid diameter of Dox-loaded V2C was 194.77±4.82nm, while the average fluid diameter of Dox-loaded V2C was 202.90±5.16nm. In addition, the zeta potential of V2C was measured to be -22.05±1.13mV, while the zeta potential of V2C-Dox was 4.47±0.33mV, indicating the successful loading of Dox. To further analyze the morphological characteristics and size of V2C, TEM was performed (Figure 1C); the sizes of the observed V2C and V2C-Dox nanosheets were consistent with the DL The results of S analysis were consistent. Subsequent HRTEM observations revealed that V2C and V2C-Dox possessed a layered crystalline structure with lattice fringes of 0.241 nm and 0.252 nm, respectively (Figure 1, DE). Energy dispersive spectroscopy (EDS) mapping revealed that the surface of the V2C nanosheets was covered with elements C and V (Figure 1, F). The temperature dependence of the hydrogel transparency was subsequently investigated, with the lower critical solution temperature (LCST) of the blank gel Gel and V2C-Dox@Gel being 22.5°C and 22.2°C, respectively (Figure 1, G—H). Frequency sweep tests revealed the frequency dependence of the G' and G" moduli of the blank gel Gel. Both G' and G" increased with increasing angular frequency, with G' exceeding G", indicating the formation of a stable solid hydrogel (Figure 1, I). The shear thinning properties of the prepared gels were tested using a static shear rate sweep, revealing that the viscosity of the blank gel Gel and V2C-Dox@Gel decreased within the shear rate range of 0.1–100 s⁻¹ (Figure 1, J—K). This indicates that both gels have excellent shear-thinning properties and are suitable for in situ tumor injection. Furthermore, within the strain range, the G' of the blank gel and V2C-Dox@Gel is greater than the G" (L in Figure 1), indicating that the gels are elastic and their structures are not damaged.After freeze-drying, blank gel Gel and V2C-Dox@Gel were subjected to SEM testing. The internal morphology of the gels was evaluated by SEM. Both freeze-dried gels showed a uniform and interconnected porous structure (MN in Figure 1). These results indicate that V2C-Dox and V2C-Dox@Gel were successfully prepared.

[0067] Effect Example 2

[0068] Safety evaluation of V2C prepared in Example 1 and V2C-Dox prepared in Example 2

[0069] After treating human red blood cells with different concentrations of V2C and V2C-Dox (equal to the concentration of V2C), the red blood cells did not hemolyze, indicating that V2C and V2C-Dox are non-toxic under conditions without laser irradiation (shown in A in Figure 2). In addition, the present invention treated breast cancer cells MDA-MB-231 and MCF-7 with 0.58 μg / mL Dox, 0.24 μg / mL V2C and V2C-Dox (equal amounts of Dox and V2C), and the cell viability was determined by CCK8 method after incubation for 12, 24, 48, 72, and 96 hours. The results are shown in BC in Figure 2 (Con in Figure 2 represents the blank control group). V2C and V2C-Dox did not affect the proliferation ability of breast cancer cells, indicating that V2C and V2C-Dox are safe and non-toxic under conditions without laser irradiation. In addition, the present invention further evaluated the safety of V2C and V2C-Dox on animals. After treating the animals with V2C and V2C-Dox, blood samples were collected and analyzed for glucose (Figure 2D), lipids (Figure 2E), ions (Figure 2F), liver function markers (Figure 2G), and blood cells (Figure 2H). Neither V2C nor V2C-Dox affected these parameters in the absence of laser irradiation, further demonstrating their safety and nontoxicity. In summary, under normal conditions, the V2C-Dox nanocarrier exhibits excellent safety and exhibits no toxic side effects on cells or animals.

[0070] Effect Example 3

[0071] The V2C-Dox prepared in Example 2 can be taken up by cells and enter the cells

[0072] Breast cancer MDA-MB-231 cells were seeded in a six-well plate. When the cell density reached 70%, V2C-Dox was added. After incubation for 24 hours, autofluorescence was observed under an inverted fluorescence microscope, as shown in Figure 3 (Con in Figure 3 represents the blank control group). The results showed that V2C-Dox-treated MDA-MB-231 cells expressed fluorescence, indicating that V2C-Dox can be taken up into the cells and further exert its effect.

[0073] Effect Example 4

[0074] Under PTT treatment, V2C-Dox prepared in Example 2 inhibited tumor growth and metastasis

[0075] The CCK8 assay was used to analyze the effect of V2C-Dox on cell proliferation under PTT treatment: breast cancer cells MDA-MB-231 and MCF-7 were seeded in 96-well plates and treated with Dox (0.58 μg / mL), V2C (0.24 μg / mL), and V2C-Dox (0.58 μg / mL) and laser irradiation. After treatment for different time periods (24 h, 48 h, and 72 h), 10 μL of CCK8 reagent was added and incubated for one hour. The absorbance of each well at 450 nm was measured and the inhibition rate was calculated. A colony formation assay was also used to analyze the effect of V2C-Dox on cell proliferation under PTT treatment: MDA-MB-231 cells were seeded in 6-well plates and treated with Dox (0.58 μg / mL), V2C (0.24 μg / mL), and V2C-Dox (0.58 μg / mL) and laser irradiation. After two weeks of incubation, images were taken and the colony formation rate was calculated. In addition, the present invention uses the AnnexinV-PI double staining method to detect the effect of V2C-Dox on cell apoptosis under PTT treatment: Dox (0.58μg / mL), V2C (0.24μg / mL), and V2C-Dox (0.58μg / mL) were treated with breast cancer cells for 24 hours respectively, and the cells were collected and stained with AnnexinV and PI at room temperature in the dark, and the apoptosis rate was analyzed by flow cytometry. The results are shown in Figure 4. Under the action of PTT, V2C-Dox significantly inhibited tumor growth (A and B in Figure 4) and induced its apoptosis (C in Figure 4). In Figure 4, Con represents the blank control group, V2C+L represents the laser irradiation group after adding V2C, and V2C-Dox+L represents the laser irradiation group after adding V2C-Dox.

[0076] To evaluate the effect of V2C-Dox on tumor cell migration under PTT treatment, the present invention conducted a wound healing experiment: MDA-MB-231 cells grown to 80-90% were wounded and treated with Dox (0.58 μg / mL), V2C (0.24 μg / mL), and V2C-Dox (0.58 μg / mL), followed by laser irradiation. Cell migration was observed 24 hours later (Figure 5, AB). In addition, a migration experiment was conducted to further evaluate the effect of V2C-Dox on cell migration under PTT treatment: MDA-MB-231 cells in the logarithmic growth phase were seeded in Transwell chambers and treated with Dox (0.58 μg / mL), V2C (0.24 μg / mL), and V2C-Dox (0.58 μg / mL), followed by laser irradiation. Migration rates were calculated 12 hours later (Figure 5, CD). As shown in Figure 5, V2C-Dox significantly inhibited tumor metastasis under PTT. In Figure 5 , Con represents the control group, V2C+L represents the group irradiated with laser after the addition of V2C, and V2C-Dox+L represents the group irradiated with laser after the addition of V2C-Dox.

[0077] The above results show that V2C-Dox+Laser (i.e., V2C-Dox+L) has a more obvious tumor cell killing effect than Dox alone when it has good compatibility, and does not produce cytotoxic effects on other normal cells and their organs.

[0078] Effect Example 5

[0079] Under PTT treatment, V2C, V2C-Dox, and V2C-Dox@Gel prepared in Examples 1-3 inhibited the growth of breast cancer implants in animals

[0080] Each mouse was injected with 10 7 MDA-MB-231 cells were injected intratumorally with Dox, V2C, V2C-Dox, and V2C-Dox@Gel after tumor formation, and then laser irradiated. The tumor size was monitored every other day and a tumor growth curve was drawn. Fifteen days later, the mice were sacrificed, and the tumor sections were sliced ​​and immunohistochemistry was performed to observe the expression of the proliferation indicator Ki67. The results are shown in Figure 6. V2C-Dox and V2C-Dox@Gel significantly inhibited the growth of mouse breast cancer tumors under the action of PTT (AB in Figure 6) and inhibited the expression of Ki67 in the tumor (CD in Figure 6). In the figure, Con represents the control group, V2C+Laser represents the laser irradiation group after adding V2C, V2C-Dox+Laser represents the laser irradiation group after adding V2C-Dox, and V2C-Dox@Gel+Laser represents the laser irradiation group after adding V2C-Dox.

[0081] The embodiments described above are merely descriptions of preferred embodiments of the present invention and are not intended to limit the scope of the present invention. Without departing from the spirit of the present invention, various modifications and improvements made to the technical solutions of the present invention by persons skilled in the art shall fall within the scope of protection defined by the claims of the present invention.

Claims

1. A vanadium carbide-coated doxorubicin nanoformulation, characterized in that: The invention comprises vanadium carbide nanosheets and doxorubicin; the doxorubicin is wrapped in the vanadium carbide nanosheets; and the mass ratio of the doxorubicin to the vanadium carbide nanosheets is 2.5:

1.

2. The vanadium carbide-coated doxorubicin nanoformulation according to claim 1, characterized in that: The dosage form of the vanadium carbide-coated doxorubicin nanoformulation is solution or gel.

3. A method for preparing the vanadium carbide-coated doxorubicin nanoformulation according to claim 1 or 2, characterized in that: The preparation method is method 1 or method 2; The method 1 comprises the following steps: The V2C nanosheets are mixed and dissolved with doxorubicin and PBS buffer, and then vortexed and sonicated to obtain the vanadium carbide-encapsulated doxorubicin nanoformulation; The second method comprises the following steps: Dissolving poloxamer in PBS buffer to obtain a poloxamer solution; The V2C nanosheets are mixed and dissolved with doxorubicin and poloxamer solution, and then vortexed and ultrasonicated to obtain the vanadium carbide-encapsulated doxorubicin nanoformulation.

4. The method for preparing the vanadium carbide-coated doxorubicin nanoformulation according to claim 3, characterized in that: In the method 1, the mass volume ratio of the V2C nanosheets to doxorubicin and PBS buffer is 1 mg:2.5 mg:2 mL.

5. The method for preparing the vanadium carbide-coated doxorubicin nanoformulation according to claim 3, characterized in that: In the second method, the concentration of poloxamer in the poloxamer solution is 200 mg / mL; the mass volume ratio of the V2C nanosheets to doxorubicin and the poloxamer solution is 1 mg:2.5 mg:2 mL.

6. The method for preparing the vanadium carbide-coated doxorubicin nanoformulation according to claim 3, characterized in that: In the method 1 and the method 2, the vortexing time is 2 minutes; and the ultrasound is specifically ice bath ultrasound for 30 minutes.

7. The method for preparing the vanadium carbide-coated doxorubicin nanoformulation according to claim 3, characterized in that: In the method 1 and the method 2, the method for preparing the V2C nanosheets comprises the following steps: V2AlC is added to an HF solution for etching, followed by centrifugation, and the resulting precipitate is mixed and stirred with a TPAOH solution, and centrifuged for purification to obtain the V2C nanosheet.

8. Use of the vanadium carbide-coated doxorubicin nanoformulation as claimed in claim 1 or 2 in the preparation of anti-tumor drugs.

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