Microenvironment double-response chitosan gene carrier as well as preparation method and application thereof

A dual-response, gene carrier technology, applied in the direction of using the carrier to introduce foreign genetic material, recombinant DNA technology, etc., to achieve the effect of solving the contradiction between gene compression and release, low cytotoxicity, and high gene transfection efficiency

Active Publication Date: 2016-08-31
DALIAN NATIONALITIES UNIVERSITY
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

According to the research results of the current literature, there has not been any report to crosslink chitosan with acid and reduction sensitive chemical bonds, synthesize acid and reducing environment responsive macromolecule crosslinked chitosan, and then conduct polyethylenimine (PEI) and polyethene Ethylene glycol (PEG) grafting to construct a chitosan-based non-viral gene carrier with a "proton sponge effect" and long-term cycle stability, reducing environment and acidic environment dual response controlled release genes

Method used

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  • Microenvironment double-response chitosan gene carrier as well as preparation method and application thereof
  • Microenvironment double-response chitosan gene carrier as well as preparation method and application thereof
  • Microenvironment double-response chitosan gene carrier as well as preparation method and application thereof

Examples

Experimental program
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Effect test

Embodiment 1

[0040] Low-molecular-weight chitosan (CSO) 0.8554 g was dissolved in 50 mL of NaAc / HAc (pH=4.5) buffer solution, and ultrasonicated for 30 min to dissolve it. Dissolve 0.1093 g of sodium periodate in 50 mL of NaAc / HAc (pH=4.5) buffer solution, and sonicate for 30 min. The above two solutions were placed in an ice bath, and degassed with high-purity nitrogen for 30 minutes. The two solutions were mixed, stirred and reacted at 0-4°C for 26 hours, and 10 mL of ethylene glycol was added to stop the reaction. The reaction solution was transferred to a dialysis bag (MWCO=3000Da), dialyzed in NaAc / HAc buffer solution for 24 hours, then dialyzed in deionized water for 48 hours, and freeze-dried to obtain aldehyde-based chitosan. Weigh 0.16g of aldehyde-based chitosan, 0.10g of 3,3'-dithiobis(hydrazine propionate), three drops of glacial acetic acid, 30mL of absolute ethanol, under nitrogen protection, stir and reflux at 68°C for 42h, vacuum rotary evaporation The solvent was removed,...

Embodiment 2

[0043] Add the prepared SRCS into [BMIM]Cl ionic liquid, 80 °C oil bath N 2 Fully stir and dissolve under air protection, add CDI in an equimolar amount to the chitosan glucosamine unit, N 2 Stir and react at 80°C for 2 h under air protection, then add 0.3 mol of mPEG-NH with chitosan glucosamine unit 2 , N 2 Under air protection, react at 80°C for 2 hours, and finally add 0.45mol PEI of chitosan glucosamine unit, N 2 Stir the reaction at 80°C for 2h under gas protection. Add an appropriate amount of deionized water to the reaction solution, transfer the reaction solution mixture to a dialysis bag (MWCO=8000-14000Da), dialyze in deionized water for 3 days, and freeze-dry to obtain SRCS-g-PEI-g-PEG polymer.

Embodiment 3

[0045] Add the prepared SRCS into DMF, 40°C N 2 Stir fully under air protection to disperse evenly, add CDI in an equimolar amount to the chitosan glucosamine unit, N 2 Stir and react at 40°C for 12 hours under air protection, then add 0.3mol chitosan-glucosamine unit mPEG-NH 2 , N 2 Under air protection, react at 40°C for 12h, and finally add 0.45mol PEI of chitosan glucosamine unit, N 2 The reaction was stirred at 40°C for 12h under the protection of air. Add an appropriate amount of deionized water to the reaction solution, transfer the reaction solution mixture to a dialysis bag (MWCO=8000-14000Da), dialyze in deionized water for 3 days, and freeze-dry to obtain SRCS-g-PEI-g-PEG polymer.

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Abstract

The invention provides a microenvironment double-response chitosan gene carrier as well as a preparation method and application thereof. The method is started from oligosaccharides of chitosan, and synthesizes macromolecular chitosan (SRCS) containing an acylhydrazone bond and a disulfide bond. The method adopts a covalent grafting method to carry out polyethyleneimine (PEI) grafting and polyethylene glycol (PEG) grafting on the SRCS, thus obtaining oxidation reduction microenvironment and acid microenvironment double-response SRCS-g-PEI-g-PEG polymer. The SRCS-g-PEI-g-PEG polymer prepared by the invention has performance of microenvironment double-response gene release control. The invention also provides the application of the SRCS-g-PEI-g-PEG polymer obtained by the method as a gene carrier in gene transfection; the SRCS-g-PEI-g-PEG polymer has good biocompatibility and high gene transfection efficiency, thus being a good carrier for gene transfection and having a wide application prospect.

Description

technical field [0001] The invention relates to a chitosan gene carrier with dual response to microenvironment, in particular to a polymer of acylhydrazone bond and disulfide bond cross-linked chitosan grafted with polyethyleneimine and polyethylene glycol and its The preparation method and application belong to the preparation method and technology of a novel high-efficiency non-viral gene carrier in the field of gene therapy. Background technique [0002] Gene therapy introduces foreign genes into the nucleus of target cells to repair defective genes that cause diseases or inhibit harmful genes that cause diseases, so as to restore the body to normal functions and achieve the purpose of treating diseases [1. Chinese invention patent, publication number CN 102140171A , the name of the invention is: Glutathione-modified chitosan copolymer used as a non-viral gene carrier material and its preparation and application]. Gene therapy technology provides new treatment methods fo...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): C08G81/00C08B37/08C12N15/85
CPCC08B37/003C08G81/00C12N15/85
Inventor 张树彪陈会英崔韶晖海华
Owner DALIAN NATIONALITIES UNIVERSITY
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