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CN-120005942-B - Cell vesicle co-delivery system, anti-tumor drug and application

CN120005942BCN 120005942 BCN120005942 BCN 120005942BCN-120005942-B

Abstract

The invention belongs to the field of tumor drug research, and particularly relates to a cell vesicle co-delivery system, an anti-tumor drug and application thereof. The cell vesicle co-delivery system is constructed according to the following steps that a tumor targeting protein or targeting peptide recombinant expression vector and a recombinant plasmid of the circRNA of a cell scorch related effector molecule are co-transfected into tool cells, and the cell vesicle co-delivery system is obtained by collecting the tool cells, wherein the tumor targeting protein or targeting peptide recombinant expression vector contains a DNA sequence for encoding MCP protein, and the recombinant plasmid of the circRNA of the cell scorch related effector molecule contains a DNA sequence for encoding MS2 protein. The cell vesicle co-delivery system provided by the invention can directly induce the death of tumor cells by delivering the cell death related effector molecule RNA through the cell vesicle, has high killing efficiency and small side effect, and can solve the problems of mass production and drug loading.

Inventors

  • XING YUQI
  • YANG GUODONG
  • ZHAO JING
  • ZHANG FEIYU
  • WEI MENGYING

Assignees

  • 中国人民解放军空军军医大学

Dates

Publication Date
20260505
Application Date
20241125

Claims (8)

  1. 1. A cell vesicle co-delivery system is characterized by being constructed according to the following steps of co-transfecting a recombinant plasmid of a tumor targeting protein or a recombinant expression vector of a targeting peptide and a circRNA of an effector molecule related to cell apoptosis into a tool cell, and collecting to obtain the cell vesicle co-delivery system; wherein the tumor targeting protein or the recombinant expression vector of the targeting peptide contains a DNA sequence encoding MCP protein and is used for expressing the MCP protein in tool cells; the recombinant plasmid of the circRNA of the effector molecule related to the apoptosis contains a DNA sequence encoding MS2 protein and is used for expressing MS2 protein in tool cells, wherein the MS2 protein is specifically combined with the MCP protein, so that the circRNA is loaded into the cell vesicle.
  2. 2. The cell-based vesicle co-delivery system of claim 1, wherein the recombinant expression vector for a tumor targeting protein or targeting peptide is obtained by ligating the sequence shown as SEQ ID No.3 to vector pcdna3.1.
  3. 3. The cell-based vesicle co-delivery system of claim 2, wherein the recombinant plasmid of the circRNA of the apoptosis-related effector molecule is obtained by ligating the sequence shown as SEQ ID No.6 to the vector pLC 5.
  4. 4. The cell type vesicle co-delivery system of claim 1, wherein the mass ratio of the recombinant expression vector of the tumor targeting protein or targeting peptide to the recombinant plasmid of the circRNA of the effector molecule related to cell apoptosis is 1:1-3.
  5. 5. The co-delivery system of cell vesicles according to claim 4 wherein the amount of transfection reagent is 2-4 times the total amount of recombinant plasmid of circRNA of effector molecules related to apoptosis of tumor targeting proteins or recombinant expression vectors of targeting peptides.
  6. 6. Use of the cell type vesicle co-delivery system according to any one of claims 1-5 for preparing an anti-tumor drug, wherein the cell type vesicle co-delivery system is used for preparing an anti-breast cancer drug.
  7. 7. An antitumor drug characterized in that it comprises the cell-based vesicle co-delivery system according to any one of claims 1 to 5 as the only active ingredient, and the antitumor drug is an antitumor drug against breast cancer.
  8. 8. The medicament of claim 7, wherein the medicament is formulated by the cell type vesicle co-delivery system with pharmaceutically acceptable excipients or carriers.

Description

Cell vesicle co-delivery system, anti-tumor drug and application Technical Field The invention belongs to the field of tumor drug research, and particularly relates to a cell vesicle co-delivery system, an anti-tumor drug and application thereof. Background Tumors are important diseases affecting human health and have become the second leading cause of death worldwide. Tumor treatment mainly comprises traditional operation treatment, radiation treatment, chemotherapy and novel treatment modes such as targeting treatment, immunotherapy and the like which are rapidly developed in recent years. Targeted drugs have been hot spots in the development of antitumor drugs, and can be broadly divided into two classes, monoclonal antibodies and small molecule compounds. In recent years, with the deep research, new targets are continuously emerging, and anti-tumor drug development is greatly advanced. Up to now, the united states food and drug administration has approved tumor targeted therapeutic drugs for more than 30 targets, many of which are approved for multiple indications, and the advent of these drugs has brought new promise to tumor patients. However, the problems of large therapeutic dose, high administration frequency and the like still exist, and inconvenience is brought to further transformation. Therefore, the strategy and application of targeting delivery effector molecule circRNA based on cell vesicles are of great significance for the development of tumor targeted therapy. Constructing a nano-delivery system is an effective solution to the problem of RNA drug delivery. Common nano-delivery systems include Lipid Nanoparticles (LNP), extracellular vesicles (extracellular vesicle, EV), and the like. EV is a membranous vesicle produced by cells and containing a lipid bilayer, including exosomes, microvesicles, apoptotic bodies, and the like. Because EV biocompatibility is good, immunogenicity is low, and the EV can cross the blood brain barrier, the EV is often used as a natural nano-carrier for drug delivery, but the EV is difficult to purify and low in yield, and limits practical application. The vesicle-like (VESICLE MIMETIC) technique is an alternative to solve this bottleneck problem. Vesicles-like means EV mimics obtained by artificial synthesis, physical extrusion of cells, etc., which are easier to produce while retaining similar biological properties to EV, and have been widely used for drug delivery. The circRNA has stability advantages in that it has no free end and is resistant to exonuclease degradation, and has a plasma half-life at least 2.5 times longer than homologous linear RNA. Therefore, the RNA drug structure is modified, and is optimized to be a circRNA configuration, so that the in vivo long circulation capacity can be improved. It is therefore urgent to begin with the discovery of an engineered cellular vesicle for targeted delivery of a circRNA drug that is effective in treating tumors with minimal side effects from the effector molecule, circRNA, in a cell death mode. However, the existing technology is more limited in exosome delivery strategies such as siRNA delivery and microRNA delivery, cell death is induced in an indirect mode, the killing speed is low, the exosome yield is low, and toxic and side effects caused by off-target problems are large. Disclosure of Invention Based on the technical problems, the invention provides a cell vesicle co-delivery system, which directly induces the death of tumor cells by delivering cell death related effector RNA through cell vesicles, has high killing efficiency and small side effect, and can solve the problems of mass production and drug loading. The specific technical scheme provided by the invention is as follows: the invention provides a cell vesicle co-delivery system, which is constructed according to the following steps: co-transfecting a recombinant expression vector of tumor targeting protein or targeting peptide and a recombinant plasmid of circRNA of effector molecules related to cell apoptosis to tool cells, and collecting to obtain a cell vesicle co-delivery system; the recombinant expression vector of the tumor targeting protein or the targeting peptide contains a DNA sequence for encoding MCP protein; the recombinant plasmid of the circRNA of the effector molecule related to the apoptosis contains a DNA sequence for encoding MS2 protein. As a preferred embodiment of the present invention, the recombinant expression vector of the tumor targeting protein or peptide is obtained by ligating the P1h3-IGSF8-MCP sequence shown in SEQ ID NO.3 to the vector PcDNA3.1. As a preferred embodiment of the invention, the recombinant plasmid of the circRNA of the effector molecule related to the apoptosis is obtained by ligating IRES-GSDMD-N-miR-145-MS2 sequence shown in SEQ ID NO.6 to vector pLC 5. As a preferred embodiment of the invention, the mass ratio of the recombinant expression vector of the tumor targeting protein or targeting