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Alpha1,3 galactosyltransferase-transfected material, its preparation method and application

A galactosyl and gene transfection technology, applied in the field of α1, can solve the problems of lack of targeting and inability to specifically act on tumor sites, and achieve good targeting effect

Inactive Publication Date: 2012-10-10
赵永祥 +1
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

[0005] The technical problem to be solved by the present invention is to provide a new vascular endothelial cell with good Targeted α1, 3GT gene transfection material and its preparation method and application

Method used

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  • Alpha1,3 galactosyltransferase-transfected material, its preparation method and application
  • Alpha1,3 galactosyltransferase-transfected material, its preparation method and application
  • Alpha1,3 galactosyltransferase-transfected material, its preparation method and application

Examples

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Effect test

Embodiment 1

[0055] Example 1: Preparation of α1,3GT gene transfection material with a particle size of 100nm, and characterizing the product, the specific preparation steps are as follows:

[0056] 1) Use chloroform to prepare POPC, DDAB, DSPE-PEG2000 and DSPE-PEG2000-maleimide to the corresponding concentrations of 20mg / mL, 5mg / mL, 20mg / mL and 10mg / mL respectively, and press 18.6μmol, 0.6μmol, Mix 0.6 μmol and 0.2 μmol in the Agilent chromatographic sample bottle; after fully mixing, pass nitrogen flow for 15 minutes to remove the organic solvent, and rotate the sample bottle continuously until a thin layer of phospholipids is formed; then place it in a rotary evaporator for vacuum evaporation for 2 hours .

[0057] 2) Take out the sample bottle, add 0.2mL TRIS-HCl buffer solution (50mM, pH7.0), rotate and oscillate slightly, place it in a water bath sonicator for 3 minutes, fully resuspend the phospholipid mixture, and avoid the generation of air bubbles as much as possible; Add enhanc...

Embodiment 2

[0062] Example 2: Preparation of α1,3GT gene transfection material with a particle size of 40nm, the specific preparation steps are as follows:

[0063] 1) Use chloroform to prepare POPC, DDAB, DSPE-PEG2000 and DSPE-PEG2000-maleimide to the corresponding concentrations of 20mg / mL, 5mg / mL, 20mg / mL and 10mg / mL respectively, and press 18.6μmol, 0.6μmol, Mix 0.6 μmol and 0.2 μmol in the Agilent chromatographic sample bottle; after fully mixing, pass nitrogen flow for 15 minutes to remove the organic solvent, and rotate the sample bottle continuously until a thin layer of phospholipids is formed; then place it in a rotary evaporator for vacuum evaporation for 2 hours .

[0064] 2) Take out the sample bottle, add 4mL TRIS-HCl buffer solution (50mM, pH7.0), rotate and oscillate slightly, place it in a water bath ultrasonicator for 3 minutes, fully resuspend the phospholipid mixture, and try to avoid the generation of air bubbles; Add dropwise enhanced green fluorescent protein (EGFP...

Embodiment 3

[0068] Example 3: preparation of α1,3GT / EGFP fusion gene transfection material, and used for the experimental study of intracellular expression. The specific preparation steps of the fusion gene transfection material are basically the same as those described in Example 1, except that after being treated with a water bath ultrasonic instrument in step 2, α1,3GT / EGFP fusion gene plasmid DNA (350 μg), 60% Ethanol (final concentration 35%, V / V), 200 mM CaCl2 solution (final concentration 4 mM), and then 5 cycles of repeated freezing and thawing. Steps 1, 3, 4 and 5 are all the same as in Example 1.

[0069] Figure 4 Shown is the expression of the prepared α1,3GT / EGFP fusion gene transfection material in the 293T cell line stably expressing Endoglin. The results in this figure show that the α1,3 galactosyltransferase gene transfection material of the present invention can specifically bind to Endoglin protein, and the gene plasmid loaded therein can be well expressed, and the ge...

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Abstract

The invention relates to an alpha1,3 galactosyltransferase-transfected material, comprising alpha1,3 galactosyltransferase galactosyltransferase gene plasmids, PEG-coupled nanoliposomes for loading the plasmids, and Endoglin antibody molecules coupled on the nanoliposomes through the PEG. The alpha1,3 galactosyltransferase-transfected material of the invention has good targeting to neovascular endothelial cells of tumor tissues, when the material specifically effects on the neovascular endothelial cells of tumor tissues, the loaded alpha1,3 galactosyltransferase gene can be expressed, and hyperacute rejection is caused in local tumor tissues, so that the purpose of treating tumors can be realized. The invention further relates to a preparation method and application of the alpha1,3 galactosyltransferase-transfected material.

Description

technical field [0001] The invention relates to the field of biomedical materials, in particular to an alpha 1,3 galactosyltransferase gene transfection material acting on neovascular endothelial cells of tumors, and a preparation method and application thereof. Background technique [0002] In my country, the incidence of tumor is high, and the number of cases is quite large. For a long time, people have been making unremitting efforts to conquer tumors, and have proposed various treatment methods, such as radiotherapy, chemotherapy, etc., but these treatment methods have more or less problems such as unsatisfactory curative effect, large side effects, and high treatment costs. Therefore, it is necessary to continue in-depth exploration of tumor treatment and develop more effective treatment methods. [0003] Hyperacute rejection (Hyperacute Rejection, HAR) is the activation of the complement system by the combination of pre-existing natural antibodies in the recipient (su...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): A61K48/00A61K47/34A61K47/42A61P35/00
Inventor 赵永祥卢小玲彭宜
Owner 赵永祥
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