TEM-1 targeted gene vaccine and construction and application thereof

A gene vaccine and targeting technology, applied in the field of targeting TEM-1 gene vaccine and its construction and application, can solve the problems of weak immunogenicity and achieve the effect of simple steps, small adverse reactions and wide range of effects

Inactive Publication Date: 2020-10-16
贵阳市第二人民医院
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

However, TEM-1 is a self-antigen, and the body forms immune tolerance due to its weak immunogenicity

Method used

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  • TEM-1 targeted gene vaccine and construction and application thereof
  • TEM-1 targeted gene vaccine and construction and application thereof
  • TEM-1 targeted gene vaccine and construction and application thereof

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0074] This embodiment provides a gene vaccine targeting TEM-1. The gene vaccine uses flagellin (SF) as an adjuvant and uses the rTEM-1 gene as an antigen to obtain the rTEM-1-SF fusion gene;

[0075] The rTEM-1 is obtained by screening the dominant epitope gene sequence rTEM-1 of T lymphocytes and B cells from the TEM-1 gene and then synthesizing the artificial rTEM-1 gene fragment by PCR;

[0076] The amino acid sequence of the rTEM-1 gene is as follows:

[0077] DLSCEDPCAQAPCEQQCEPGGPQGYSCHCRLGFRPAEDDPH RCVDTDECQIAGVCQQMCVNYVGGFECYCSEGHELEADGISCSP AGAMGAQASQDLRDELLDDGEEGEDEEEPWEDFDGTWTEEQGI LWLAPTHPPDFGLPYRPNFPQDGEPQRLHLEPTWPPPLSAPRGPY HSSVVSATRPMVISATRPTLPSAHKTSVISATRPPLSPVHPPAMAPA TPPAVFSEHQIPKIKANYPDLPFGHKPGITSATHPARSPPYQPPIIST NYPQVFPPHQAPMSPDTHTITYLPPVPPHLDPGDTTSKAHQHPLL PDAPGIRTQAPQLSVSALQPPLPTNSRSSVHETPVPAANQPPAFPS SPLPPQRPTNQTSSISPTHSYSRAPLVPREGVPSPKSVPQLPSVPST AAPTALAESGLAGQSQRDDRWLLVALLVPTCVFLVVLLALGIVY CTRCGSHAPNKRITDCYRWVTHAGNKSSTEPMPPRGSLTGVQTC RTSV

[0078] Th...

Embodiment 2

[0084] The construction method of the targeted TEM-1 gene vaccine comprises the steps of:

[0085] (1) Obtain rTEM-1-SF fusion gene

[0086] According to the full-length DNA sequence of TEM-1 registered in GenBank, the DNA Star bioinformatics software was used to analyze T lymphocyte and B cell epitopes, intercept the extracellular expression sequence and screen the dominant epitope gene sequence rTEM of T lymphocytes and B cells -1, synthesize artificial rTEM-1 gene fragments by PCR, and use flagellin protein as an adjuvant to obtain rTEM-1-SF fusion gene;

[0087] (2) enzyme-cut connection of vector and rTEM-1-SF fusion gene

[0088] Digest the rTEM-1-SF fusion gene and vector with NheI and XhoI, and then use T4 DNA ligase to connect the recovered rTEM-1-SF fusion gene fragment and vector digestion fragment to obtain the ligation product;

[0089] (3) Conversion of ligation products

[0090] Take the ligation product and Escherichia coli competent cells and add them to EP...

Embodiment 3

[0110] This example explores the application of a gene vaccine targeting TEM-1 in the prevention and treatment of breast cancer; the specific operations are as follows:

[0111] 1. Preparation of targeted TEM-1 gene vaccine

[0112] 1.1 Acquisition of rTEM-1-SF fusion gene

[0113] According to the full-length DNA sequence of TEM-1 registered in GenBank, the DNA Star bioinformatics software was used to analyze T lymphocyte and B cell epitopes, intercept the extracellular expression sequence and screen the dominant epitope gene sequence rTEM of T lymphocytes and B cells -1, synthesize artificial rTEM-1 gene fragments by PCR, and use flagellin protein as an adjuvant to obtain rTEM-1-SF fusion gene;

[0114] 1.2 Restriction ligation of vector and rTEM-1-SF fusion gene

[0115] The rTEM-1-SF fusion gene and pcDNA3.1 (Amp) vector were digested with NheI and XhoI, and then the recovered rTEM-1-SF fusion gene fragment and vector digestion fragment were ligated with T4 DNA ligase to...

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Abstract

The invention relates to the technical field of biology, in particular to a TEM-1 targeted gene vaccine and construction and application thereof. According to the gene vaccine, a flagellin (SF) is used as an adjuvant, rTEM-1 is used as an antigen, and a rTEM-1-SF fusion gene is obtained, and is constructed in eukaryotic expression plasmids pcDNA3.1. The rTEM-1/ flagellin fusion gene vaccine can restrain formation of tumor vessels of breast cancer tumor-bearing mice, the size of the tumor and transfer of the tumor, can restrain the transfer of breast cancer cells to lung tissue and liver tissue, and has the effects of preventing and treating the breast cancer.

Description

technical field [0001] The invention relates to the field of biotechnology, in particular to a gene vaccine targeting TEM-1 and its construction and application. Background technique [0002] The formation of new blood vessels inside the tumor is closely related to the occurrence, development, local invasion and metastasis of solid tumors. Tumor-associated vascular antigens are highly expressed in a variety of cancers, but are less expressed in normal tissues. Targeting tumor vascular endothelial cells has the following advantages: First, the drug in the blood is in direct contact with its target, avoiding crossing the endothelial cells. Drugs can easily reach the drug concentration at the effect site by coupling cytotoxic substances, pro-apoptotic substances, procoagulant substances, etc.; secondly, endothelial cells are highly genetically stable, and phenotypic mutations lead to drug resistance The possibility of this is greatly reduced; finally, each capillary can provid...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): A61K39/00A61K39/39A61P35/00A61P35/04C12N15/85C12N15/66C12N15/62
CPCA61K39/00A61K39/39A61P35/00A61P35/04C12N15/85C12N15/66C07K14/7056A61K2039/53A61K2039/812A61K2039/55516C12N2800/107C07K2319/40C07K2319/00
Inventor 袁军李伟张景李荣辉罗力卓召振
Owner 贵阳市第二人民医院
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