A biomimetic cancer cell membrane engineered cisplatin nanomedicine and its preparation method and application
Through the double-encapsulation cancer cell membrane engineering technology, the problem of cisplatin drug inactivation and toxicity in the body is solved, tumor-targeted delivery and high-efficiency anti-cancer effects are achieved, making it suitable for clinical application.
Patent Information
- Application Number
- CN202411732514.6
- Authority / Receiving Office
- CN · China
- Patent Type
- Patents(China)
- Current Assignee / Owner
- Filing Date
- 2024-11-29
- Publication Date
- 2025-09-12
- Estimated Expiration
- 2044-11-29
Smart Images
Figure CN119679750B_ABST
Abstract
Description
Technical Field
[0001] The present invention belongs to the field of biomedicine, and in particular relates to a biomimetic cancer cell membrane engineered cisplatin nano anticancer drug and a preparation method and application thereof. Background Art
[0002] Cisplatin is a crucial first-line anticancer chemotherapy drug, playing a key role in over 70% of clinical cancer treatment regimens. However, the human body contains numerous macromolecular and micromolecular nucleophiles, such as DNA bases, protein residues, anions, and sulfur-containing biomolecules, which readily react with platinum ions. This is one of the main causes of cisplatin's systemic toxicity and inactivation, severely impacting its clinical efficacy.
[0003] In order to overcome the low efficacy and high toxicity caused by biodistribution, tumor-targeted delivery of cisplatin highlights its important value. Its research is currently focused on the design and preparation of cisplatin nanoformulations. Due to the water solubility of cisplatin drugs, it is not suitable for conventional physical encapsulation in nanocarriers. The common countermeasure is to bond cisplatin to a targeted polymer carrier through coordination chemistry and then form a nanodrug. However, the complex synthesis of polymer carriers limits its clinical application. In addition, the targeted delivery of cisplatin faces other challenges, including the immune system recognizing the drug carrier as a foreign body and clearing it prematurely, the heterogeneity and complexity of tumors leading to inefficient targeting or even off-target effects, especially the inactivation of cisplatin caused by nucleophilic substances such as proteins and sulfur-containing biological molecules mentioned above.
[0004] In order to reduce the non-specific binding of proteins, a protective hydrophilic corona composed of soluble polymer fragments (such as polyethylene glycol and polyanions) is usually designed on the surface of cisplatin nanoformulations to repel protein adsorption. However, this carrier molecule design cannot prevent the large number of small molecule nucleophiles present in the body from interacting with cisplatin. Taking cisplatin as an example, it is particularly easy to be rapidly neutralized by sulfur-containing compounds such as metallothionein and glutathione. Therefore, the development of cisplatin nanomedicines that can achieve tumor targeting and resist interference from nucleophiles in the body, although extremely challenging, is undoubtedly of great significance. In addition, the preparation process of such nanomedicines needs to be simplified to facilitate clinical application and transformation. Summary of the Invention
[0005] The present invention aims to provide a biomimetic cancer cell membrane-engineered cisplatin nanopharmaceutical, its preparation method, and its use in the preparation of anticancer drugs. This nanopharmaceutical, with its biomimetic cell structure, isolates cisplatin, ensuring that it circulates in the body without being inactivated by nucleophilic attack. It also targets homologous tumors, improving efficacy and reducing toxic side effects. This invention does not utilize any other artificial materials, resulting in excellent biosafety and high feasibility for clinical translation.
[0006] In order to solve the above problems, the present invention adopts the following technical solutions:
[0007] In the first aspect of the present invention, a bionic cancer cell membrane-engineered cisplatin nano-anticancer drug is provided. The nano-anticancer drug is formed by double encapsulation. The first encapsulation in the double encapsulation is to combine cisplatin with the first cancer cell membrane to form positively charged nanoparticles, and the second encapsulation in the double encapsulation is to compound the positively charged nanoparticles with the second cancer cell membrane.
[0008] According to the above scheme, the cisplatin is an anticancer drug approved by the State Food and Drug Administration.
[0009] According to the above scheme, in the first encapsulation, the mass ratio of the first cancer cell membrane to cisplatin is 1:1-1:4.
[0010] According to the above scheme, in the second encapsulation, the mass ratio of the positively charged nanoparticles to the second cancer cell membrane is 1:1-1:4.
[0011] Cancer cell membranes were extracted from cancer cells according to the above protocol.
[0012] In a second aspect of the present invention, a method for preparing the above-mentioned biomimetic cancer cell membrane engineered cisplatin nano anticancer drug is provided, the method comprising:
[0013] Dispersing cancer cell membrane fragments and cisplatin in deionized water, respectively, to obtain cancer cell membrane dispersion and cisplatin solution;
[0014] The cancer cell membrane dispersion and the cisplatin solution are mixed, shaken, and then centrifuged to perform a first encapsulation to obtain positively charged nanoparticles;
[0015] The positively charged nanoparticles are mixed with the cancer cell membrane dispersion, and after ultrasonic dispersion, centrifugation is performed for a second encapsulation to obtain a biomimetic cancer cell membrane engineered cisplatin nano anticancer drug.
[0016] In the first encapsulation, the concentration of the cancer cell membrane dispersion is 0.5 mg / mL-1.5 mg / mL; the concentration of the cisplatin solution is 0.5 mg / mL-1.5 mg / mL.
[0017] In the third aspect of the present invention, there is provided a use of the above-mentioned biomimetic cancer cell membrane engineered cisplatin nanomedicine in the preparation of anticancer drugs.
[0018] Furthermore, the cancer treated by the anticancer drug includes at least one of breast cancer, colon cancer, and liver cancer.
[0019] The present invention has the following advantages and beneficial effects:
[0020] 1. The nanoparticle anticancer drug provided by this invention utilizes groups on the surface of cancer cell membranes to complex with cisplatin, forming a positively charged nanocore. This core then electrostatically interacts with the negatively charged cancer cell membrane to form a negatively charged nanoparticle. This double encapsulation effectively isolates the encapsulated cisplatin from nucleophiles in the body during the nanoparticle's circulation. Simultaneously, it achieves homologous targeting of the cancer cell membrane, significantly reducing the risk of drug inactivation and systemic toxicity, enabling precise cancer treatment.
[0021] 2. The nano anticancer drug provided by the present invention has simple ingredients, good biocompatibility, small toxic and side effects, obvious therapeutic effect, simple preparation process, low equipment requirements, and has good application prospects in the field of cancer treatment. BRIEF DESCRIPTION OF THE DRAWINGS
[0022] Figure 1 This is a scanning electron microscope image of the nano anticancer drug prepared in Example 1.
[0023] Figure 2 This is a diagram of the in vitro anticancer effect of the nano anticancer drug prepared in Example 1.
[0024] Figure 3 The efficacy of the nano-cisplatin anticancer drug prepared in Example 1 is tolerant to nucleophilic small molecules and proteins, where: A is the efficacy in the presence of glutathione; B is the efficacy in the presence of serum albumin (*P<0.05; **P<0.01; ***P<0.001).
[0025] Figure 4 This is the homologous tumor targeting effect of the nano anticancer drug prepared in Example 1 in tumor-bearing mice.
[0026] Figure 5 The nano anticancer drug prepared in Example 1 was used to treat tumor-bearing mice for 14 days and had an inhibitory effect on tumor growth (***P<0.001). DETAILED DESCRIPTION
[0027] The present invention will be described in detail below in conjunction with specific embodiments and examples, and the advantages and various effects of the present invention will be more clearly presented. It should be understood by those skilled in the art that these specific embodiments and examples are for illustrating the present invention, rather than for limiting the present invention.
[0028] Throughout this specification, unless otherwise specified, the terms used herein should be understood as having the same meaning as commonly used in the art. Therefore, unless otherwise defined, all technical and scientific terms used herein have the same meaning as commonly understood by one of ordinary skill in the art to which this invention belongs. In the event of any conflict, the present specification shall take precedence.
[0029] Unless otherwise specified, various raw materials, reagents, instruments and equipment used in the present invention can be purchased from the market or obtained through existing methods.
[0030] The present application will be described in detail below with reference to examples and experimental data.
[0031] Example 1: Biomimetic cancer cell membrane engineered cisplatin nanoparticle anticancer drug and its preparation method
[0032] The present invention provides a method for preparing a biomimetic cancer cell membrane-engineered cisplatin nano anticancer drug, which comprises the following steps:
[0033] (1) Cancer cell membranes extracted from mouse breast cancer cells were dispersed in deionized water to form a 1 mg / mL dispersion.
[0034] (2) Dissolve cisplatin in deionized water to form a 1 mg / mL solution.
[0035] (3) The cisplatin solution and the cancer cell membrane dispersion were mixed at a mass ratio of 1:2 between the cancer cell membrane and cisplatin, shaken, and centrifuged to obtain positively charged nanoparticles.
[0036] (4) The positively charged nanoparticles are dispersed in deionized water to form a 1 mg / mL dispersion, and the dispersion is mixed with a cancer cell membrane dispersion at a mass ratio of 1:2, ultrasonicated, and centrifuged to obtain a biomimetic cancer cell membrane engineered cisplatin nano anticancer drug.
[0037] Figure 1 The scanning electron microscope image of the nano anticancer drug prepared in this example is shown in Figure 2. The prepared nano anticancer drug dispersion was added dropwise to a silicon wafer and observed after natural drying. Figure 1 As shown, the nanoparticles are regular spherical in shape and have a size of about 100 nanometers.
[0038] Example 2: In vitro anticancer experiment
[0039] Test method: Mouse breast cancer cells were seeded in a 96-well plate at a density of 6000 cells / well and cultured with 100 μL of culture medium for 24 hours. Then, 100 μL of a gradient dispersion of nanomedicine prepared in culture medium was added to each well. All cells were cultured at 37°C for 48 hours. Subsequently, 20 μL of 5 mg / m MTT (MTT dissolved in PBS buffer) was added to each well. After 4 hours of co-culture, the culture medium was aspirated and 150 μL of dimethyl sulfoxide was added. The absorbance at 570 nm in each well was measured using a microplate reader, and the cell survival rate was calculated to obtain the toxicity of the nano anticancer drug to mouse breast cancer cells.
[0040] The results of the in vitro anticancer effects of nano anticancer drugs are as follows Figure 2 As shown, the survival rate of mouse breast cancer cells gradually decreased with the increase of nanomedicine concentration, showing a good anti-cancer effect.
[0041] Example 3: Detection of the effect of nano-anticancer drugs in isolating cisplatin from binding to nucleophilic molecules and proteins in the body
[0042] Mouse breast cancer cells were seeded in a 96-well plate at a density of 6000 cells / well and cultured with 100 μL of culture medium for 24 hours. Cisplatin and the prepared nanomedicine were mixed with glutathione and serum protein respectively and placed for 6 hours. Then, they were dispersed in the culture medium to prepare a concentration gradient dispersion and added to each well. All cells were cultured at 37°C for 48 hours. Subsequently, 20 μL of 5 mg / mL MTT (MTT dissolved in PBS buffer) was added to each well. After co-culture for 4 hours, the culture medium was aspirated and 150 μL of dimethyl sulfoxide was added. The absorbance value at 570 nm in each well was measured with an enzyme-linked microplate reader, and the cell survival rate was calculated to obtain the effect of the nano anticancer drug in isolating cisplatin from the binding of nucleophilic molecules and proteins in the body.
[0043] The effect of testing the nano anticancer drug on isolating cisplatin from binding to nucleophilic molecules and proteins (glutathione or serum albumin) in the body is shown in the figure. Figure 3 As shown, at the same concentration of nucleophilic substances, the ability of nanomedicine to kill cancer cells is significantly greater than that of the small molecule drug cisplatin, reflecting its excellent ability to tolerate nucleophilic molecules and proteins.
[0044] Example 4: In vivo anticancer experiment
[0045] Mouse breast cancer cells (4T1) and mouse colon cancer cells (CT26) were inoculated subcutaneously on the right back of 6-week-old female BALB / c mice (5×10 5 cells). When the tumor grows to 150 mm 3 The cy5-labeled nano anticancer drug (10 mg / kg) was intravenously injected into mice bearing 4T1 and CT26 tumors. The mice were then monitored in real time using an in vivo imaging system at time points of 0, 0.5, 1, 2, 4, 6, 8, 10, 12, 24, 36, and 48 hours to detect the fluorescence intensity at the tumor site of the mice.
[0046] The fluorescence intensity of mouse tumors is as follows Figure 4 As shown, after 48 hours, fluorescence was significantly enriched in the 4T1 tumor site. Throughout the observation period, the fluorescence intensity in the 4T1 tumor was significantly higher than that in the CT26 tumor, and the trend became increasingly pronounced. The prepared nanoparticle anticancer drug exhibited significant tumor targeting and homologous tumor selectivity.
[0047] Mouse breast cancer cells (4T1) were inoculated subcutaneously on the right back of 6-week-old female BALB / c mice (5×10 5 cells). When the tumor grows to 65 mm 3 At 10 pm, the nano anticancer drug (10 mg / kg) was intravenously injected into mice bearing 4T1 tumors, and the size of the tumors in the mice was recorded on time.
[0048] The effect of nano anticancer drugs on inhibiting tumor growth after 14 days of treatment in tumor-bearing mice is shown in the figure below. Figure 5 As shown, the nano anticancer drug significantly inhibited the growth of tumors.
[0049] The above embodiments are preferred implementations of the present invention, but the implementations of the present invention are not limited to the above embodiments. Any other changes, modifications, substitutions, combinations, and simplifications that do not deviate from the spirit and principles of the present invention should be considered equivalent replacement methods and are included in the scope of protection of the present invention.
[0050] Finally, it should be noted that the terms "comprises," "comprising," or any other variations thereof are intended to cover non-exclusive inclusion, such that a process, method, article, or apparatus that includes a list of elements includes not only those elements, but also other elements not expressly listed, or elements inherent to such process, method, article, or apparatus.
[0051] Although preferred embodiments of the present invention have been described, those skilled in the art may make additional changes and modifications to these embodiments once they are aware of the basic inventive concepts. Therefore, the appended claims are intended to be interpreted as including the preferred embodiments and all changes and modifications that fall within the scope of the invention. Obviously, those skilled in the art may make various changes and modifications to the present invention without departing from the spirit and scope of the invention. Thus, the present invention is intended to include such changes and modifications as fall within the scope of the claims and their equivalents.
Claims
1. A biomimetic cancer cell membrane engineered cisplatin nanomedicine, characterized in that: The nanomedicine is formed by double encapsulation, wherein the first encapsulation in the double encapsulation is to combine cisplatin with a first cancer cell membrane to form positively charged nanoparticles, and the second encapsulation in the double encapsulation is to combine the positively charged nanoparticles with a second cancer cell membrane; The preparation method of the nano drug comprises: Dispersing cancer cell membrane fragments and cisplatin in deionized water, respectively, to obtain cancer cell membrane dispersion and cisplatin solution; The cancer cell membrane dispersion and the cisplatin solution are mixed, shaken, and then centrifuged to perform a first encapsulation to obtain positively charged nanoparticles; wherein the mass ratio of the cancer cell membrane to the cisplatin is 1:2; The positively charged nanoparticles are mixed with the cancer cell membrane dispersion, ultrasonically dispersed and then centrifuged for a second encapsulation to obtain a biomimetic cancer cell membrane-engineered cisplatin nanomedicine, wherein the mass ratio of the positively charged nanoparticles to the cancer cell membrane is 1:2, and the cancer cells are breast cancer.
2. A method for preparing the biomimetic cancer cell membrane engineered cisplatin nanomedicine according to claim 1, characterized in that: The method comprises: Dispersing cancer cell membrane fragments and cisplatin in deionized water, respectively, to obtain cancer cell membrane dispersion and cisplatin solution; The cancer cell membrane dispersion and the cisplatin solution are mixed, shaken, and then centrifuged for a first encapsulation to obtain positively charged nanoparticles, wherein the mass ratio of the cancer cell membrane to the cisplatin is 1:2; The positively charged nanoparticles are mixed with the cancer cell membrane dispersion, ultrasonically dispersed and then centrifuged for a second encapsulation to obtain a biomimetic cancer cell membrane-engineered cisplatin nanomedicine, wherein the mass ratio of the positively charged nanoparticles to the cancer cell membrane is 1:2, and the cancer cells are breast cancer.
3. The preparation method according to claim 2, characterized in that In the first encapsulation, the concentration of the cancer cell membrane dispersion is 0.5 mg / mL-1.5 mg / mL; the concentration of the cisplatin solution is 0.5 mg / mL-1.5 mg / mL.
4. The preparation method according to claim 2, characterized in that The cancer cell membrane fragments are extracted from cancer cells.
5. Use of the bionic cancer cell membrane engineered cisplatin nanomedicine according to claim 1 in the preparation of an anticancer drug, wherein the cancer treated by the anticancer drug is breast cancer.