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Recombinant adeno-associated virus vector for treating atherosclerosis, gene composition and application

A technology for atherosclerosis and viral vectors, applied in the direction of viruses/bacteriophages, the use of vectors to introduce foreign genetic material, applications, etc.

Active Publication Date: 2021-08-17
XIEHE HOSPITAL ATTACHED TO TONGJI MEDICAL COLLEGE HUAZHONG SCI & TECH UNIV
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

However, the application of this measure still cannot prevent the occurrence of severe cardiovascular diseases to a large extent, especially for patients with atherosclerosis in the middle and late stages, so it is imperative to find a new direction for the treatment of atherosclerosis

Method used

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  • Recombinant adeno-associated virus vector for treating atherosclerosis, gene composition and application
  • Recombinant adeno-associated virus vector for treating atherosclerosis, gene composition and application
  • Recombinant adeno-associated virus vector for treating atherosclerosis, gene composition and application

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0037] The preparation and packaging process of the adeno-associated virus vector of AAV8-TBG-LOX-1 in Example 1 are as follows:

[0038] (1) Obtaining the target gene: From the Gene database in NCBI, it was found that the LOX-1 gene is mainly expressed in human umbilical vein endothelial cells, and the cells were used to extract RNA (Trizol extraction), and after reverse transcription (reverse transcription kit) Use the synthesized primers with protective bases and restriction sites (see the end of the article for the design method of primers with restriction sites) for PCR (use high-fidelity enzymes for PCR to avoid mismatches). PCR conditions were adjusted and optimized as needed. Subsequently, the PCR product was subjected to agarose electrophoresis (1%), and the gel piece corresponding to the target gene fragment was excised, and the gel was recovered (using a gel recovery kit), and the DNA concentration was measured after the gel recovery. At this point, we have obtaine...

Embodiment 2

[0053]Embodiment two: the adeno-associated virus vector of the AAV8-TBG-LOX-1 that the virus titer prepared in embodiment one is 2×1013 vg (virus genomes) / ml is diluted to 2×1011 vg / ml with sterile PBS buffer solution Only / 100μl, take 10ul virus stock solution + 90ul sterile PBS, configure it in an ultra-clean bench, mix it with a pipette gun, and store it in an ice box at low temperature. The titer has an impact; then use an insulin needle to draw 100ul of the virus dilution and inject it into the mouse through the tail vein, waiting to observe the transfection effect. Under normal circumstances, the virus can be stored in a -80 refrigerator. After injection, the virus can be stably expressed in about 2 weeks after injection, and the duration can be stable for at least 2 months.

[0054] The expression of AAV8-TBG-LOX-1 adeno-associated virus vector in the liver, the transfection effect and the therapeutic effect on atherosclerosis will be studied in combination with differen...

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Abstract

The invention provides a recombinant adeno-associated virus vector for treating atherosclerosis, a gene composition and application of the gene composition. The recombinant adeno-associated virus vector is obtained by inserting a lectin-like oxidized low-density lipoprotein receptor 1 (LOX-1) into an adeno-associated virus vector, and the recombinant adeno-associated virus vector and a TBG promoter are combined together to form a gene composition for preparing a gene therapy injection for treating atherosclerosis. The sequence of the LOX-1 gene is shown as SEQ ID NO: 1 in a sequence table. According to the invention, the drug effect element can be efficiently introduced into the body through intravenous injection by utilizing the adeno-associated virus vector, the ectopic efficient expression of the therapeutic protein LOX-1 of the expression product of the drug effect element is realized, and the LOX-1 receptor which is not expressed in the liver is successfully expressed in liver cells through the TBG promoter; and overloaded OX-LDL in the circulation of a patient with atherosclerosis is phagocytized and cleared by means of the strong lipid metabolism capability and metabolic pathway of the liver, so that the progress of atherosclerosis is inhibited.

Description

technical field [0001] The invention relates to the field of biotechnology, in particular to a recombinant adeno-associated virus vector, gene composition and application for treating atherosclerosis, specifically an AAV virus vector system that mediates the specific ectopic expression of LOX-1 in the liver , the LOX-1 receptor that is not expressed in the liver is successfully expressed in the liver cells, and with the help of the liver's powerful lipid metabolism and metabolic pathways, the overloaded OX-LDL in the circulation of patients with atherosclerosis is swallowed and cleared, and then inhibited Progression of atherosclerosis. Background technique [0002] Atherosclerosis (atherosclerosis) is a systemic pathological change caused by abnormal deposition of cholesterol-rich lipoproteins on the vessel wall. Continued to rise by 21.1%. In more than 100 years of atherosclerosis research and practice, modern clinical practice has been able to use statin drugs that inhi...

Claims

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Application Information

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IPC IPC(8): C12N15/864C12N15/12A61K48/00A61K38/17A61P9/10
CPCC12N15/86C07K14/705A61K48/0008A61K48/0058A61K38/177A61P9/10C12N2750/14143C12N2800/107C12N2830/008
Inventor 王志文郭小朋魏宇淼曾状林孙谛曹炳鑫张青杜高辉陈娟陈磊任衍乔张卫华景熙瑞李嘉熙
Owner XIEHE HOSPITAL ATTACHED TO TONGJI MEDICAL COLLEGE HUAZHONG SCI & TECH UNIV
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