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Genetic vector system of nanoparticle with multiple oxidation-reduction stimulus response as well as preparation method and application of genetic vector system

A gene carrier and system technology, applied in the field of nanoparticle gene carrier, can solve the problems of low transfection efficiency, compressed DNA, high surface positive charge, etc.

Active Publication Date: 2015-04-22
SICHUAN UNIV
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

[0004] Polyethyleneimine (PEI) with a molecular weight of 25kDa is widely used as a gold standard polymer carrier in non-viral carriers, but the inherent cytotoxicity and high positive surface charge of PEI limit its application in vivo
OEI with a molecular weight of 800Da has almost no cytotoxicity, but it cannot compress DNA very well, and the transfection efficiency is low

Method used

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  • Genetic vector system of nanoparticle with multiple oxidation-reduction stimulus response as well as preparation method and application of genetic vector system
  • Genetic vector system of nanoparticle with multiple oxidation-reduction stimulus response as well as preparation method and application of genetic vector system
  • Genetic vector system of nanoparticle with multiple oxidation-reduction stimulus response as well as preparation method and application of genetic vector system

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0061] Example 1: Preparation of gene carrier (OEI-SS / DNA / HA-SS-COOH) with double redox stimulus responsiveness

[0062] Dissolve the plasmid DNA in sterile HBG buffer (20 mmol of 4-hydroxyethylpiperazineethanesulfonic acid, 5% glucose) to prepare a DNA solution with a concentration of 0.1 mg / mL; Polyethylenimine (OEI-SS) was dissolved in HBG buffer to prepare OEI-SS solution with a concentration of 0.1-10 mg / mL; the masking system containing disulfide bonds (HA-SS-COOH) was dissolved in HBG HA-SS-COOH solution with a concentration of 0.01-1 mg / mL in the buffer solution.

[0063] The above OEI-SS solution and the plasmid DNA solution were mixed, and the mixed solution was incubated at room temperature for 20 minutes to obtain the OEI-SS / DNA binary complex. Then add HA-SS-COOH solution, and incubate the resulting mixed solution at room temperature for 20 minutes to obtain the gene carrier OEI-SS / DNA / HA- SS-COOH ternary complex.

Embodiment 2

[0064] Example 2: Gene carrier with dual redox stimulus responsiveness (OEI-SeSe x / DNA / HA-SeSe-COOH) preparation

[0065] Dissolve the plasmid DNA in sterile HBG buffer (20 mmol of 4-hydroxyethylpiperazineethanesulfonic acid, 5% glucose) to prepare a DNA solution with a concentration of 0.1 mg / mL; Polyethyleneimine (OEI-SeSe x ) was dissolved in HBG buffer to prepare OEI-SeSe at a concentration of 0.1-10 mg / mL x Solution: Dissolve the shielding system (HA-SeSe-COOH) containing diselenium bonds in HBG buffer to prepare a HA-SeSe-COOH solution with a concentration of 0.01-1 mg / mL.

[0066] The above OEI-SeSe x The solution solution and the plasmid DNA solution were mixed, and the mixed solution was incubated at room temperature for 20 minutes to obtain OEI-SeSe x / DNA binary complex. After adding HA-SeSe-COOH solution, the resulting mixed solution was incubated at room temperature for 20 minutes, and the gene carrier OEI-SeSe with multiple redox stimulus responsiveness c...

Embodiment 3

[0067] Example 3: Gene carrier with dual redox stimulus responsiveness (OEI-SeSe x / DNA / HA-SS-COOH) preparation

[0068] Dissolve the plasmid DNA in sterile HBG buffer (20 mmol of 4-hydroxyethylpiperazineethanesulfonic acid, 5% glucose) to prepare a DNA solution with a concentration of 0.1 mg / mL; Polyethyleneimine (OEI-SeSe x ) was dissolved in HBG buffer to prepare OEI-SeSe at a concentration of 0.1-10 mg / mL x Solution: Dissolve the shielding system (HA-SS-COOH) containing disulfide bonds in HBG buffer to prepare a HA-SS-COOH solution with a concentration of 0.01-1 mg / mL.

[0069] The above OEI-SeSe x The solution solution and the plasmid DNA solution were mixed, and the mixed solution was incubated at room temperature for 20 minutes to obtain OEI-SeSe x / DNA binary complex. After adding HA-SS-COOH solution, the resulting mixed solution was incubated at room temperature for 20 minutes, and the gene carrier OEI-SeSe with multiple redox stimulus responsiveness was obtain...

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Abstract

The invention discloses a genetic vector system of a nanoparticle with multiple oxidation-reduction stimulus response as well as a preparation method and application of the genetic vector system. The genetic vector system is composed of a shielding system, a cation material and foreign plasmid DNA (deoxyribonucleic acid); and the shielding system and the cation material both have oxidation-reduction stimulus response, so that a ternary complex nanoparticle with multiple oxidation-reduction stimulus response is formed. The genetic vector of the ternary complex nanoparticle is low in cytotoxicity, and capable of well compressing and compounding the plasmid DNA under physiological conditions, successfully transferring a supported genetic substance into a cell and entering the cell; and the reduction environment in the cell can be more rapidly broken to release the supported gene, so that the expression of gene substances is realized, the transfection process is finished, and the gene transfection efficiency can be remarkably increased.

Description

technical field [0001] The invention belongs to the field of biomedical materials, and relates to a nanoparticle gene carrier with multiple redox stimulation response characteristics. Background technique [0002] Viruses are currently the most effective gene carriers in the treatment of genetic diseases and tumors, but their non-negligible safety hazards limit the clinical application of virus vectors. Most viruses are nanoscale particles with a core-shell structure consisting of a core with a compressed genome and a protein shell. The virus exists stably outside the cell, and it can recognize the signal provided by the cell, and carry out step-by-step molecular transformation according to changes in the body or the microenvironment of the cell, thereby initiating the disassembly response, which provides a good design for the design of non-viral gene vectors enlightenment. People have also designed nucleocapsid "artificial viruses" that mimic virus disassembly responses, ...

Claims

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Application Information

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Patent Type & Authority Patents(China)
IPC IPC(8): C12N15/85A61K48/00A61P35/00A61P11/06A61P9/00
Inventor 顾忠伟聂宇何一燕程刚谢丽
Owner SICHUAN UNIV
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