Metal coordination polycation gene vector and preparation method and application thereof

A polycation and gene carrier technology, applied in the field of metal-coordination polycation gene carrier and its preparation, can solve the problems of low transfection efficiency of stem cells, and achieve the effects of expanding applications, improving DNA binding ability, and improving endocytosis efficiency.

Active Publication Date: 2017-03-15
NANKAI UNIV
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

However, the transfection efficiency of these polycations, especially for stem cells, is very low

Method used

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  • Metal coordination polycation gene vector and preparation method and application thereof
  • Metal coordination polycation gene vector and preparation method and application thereof
  • Metal coordination polycation gene vector and preparation method and application thereof

Examples

Experimental program
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Effect test

Embodiment 1

[0025] A metal coordination polycation gene carrier, composed of a coordination metal, a coordination unit, a stimulus response unit and a polycation, the coordination metal is zinc (Zn), and the coordination unit is a lutidine amine group, The stimuli response unit is N,N'-bis(acryloyl)cystamine (CBA) containing disulfide bond monomer; the polycation is polyethyleneimine (PEI) with a weight average molecular weight of 1800, and the zinc coordination monomer modifies the polycation The Zn-PD structural formula obtained by PEI is as follows:

[0026]

[0027] In the structural formula: "M" represents the coordination metal Zn; "n" represents the number of metal ion ligands grafted on one polyethyleneimine (PEI) molecule, and n is 2 or 4.

[0028] A method for preparing the metal-coordinated polycation gene carrier Zn-PD, comprising the steps of:

[0029] 1) Preparation of stimuli-responsive ligand DDAC

[0030] Dissolve 16mmol of 1,4-p-dichlorobenzyl (Dx) in 20mL of dichlo...

Embodiment 2

[0035]A metal coordination polycation gene carrier, composed of a coordination metal, a coordination unit, a stimulus response unit and a polycation, the coordination metal is magnesium (Mg), and the coordination unit is a lutidine amine group, The stimuli response unit is bis(2-propylene) ethoxy disulfide (DSDA) containing disulfide bond monomer; the polycation is the first generation polyamide-amine (PAMAM), and the polycation PAMAM is obtained by modifying the polycation PAMAM with magnesium coordination monomer The structural formula of Mg-PAD is as follows:

[0036]

[0037] In the structural formula: "M" represents the coordination metal Mg; "n" represents the number of metal ion ligands grafted on one polyamide-amine (PAMAM) molecule, and n is 1 or 3.

[0038] A preparation method of the metal-coordinated polycation gene carrier Mg-PAD, comprising the steps of:

[0039] 1) Preparation of stimuli-responsive ligand DDAD

[0040] Dissolve 16mmol of 1,4-p-dichlorobenzy...

Embodiment 3

[0044] The application of the prepared metal-coordinating polycation gene carrier is used for non-viral gene carrier to deliver deoxyribonucleic acid (DNA) in cells. Let it stand for 10-30 minutes under high temperature to prepare a Zn-PD / DNA complex with a mass ratio of 5-60:1; the prepared complex is not only used to transfect conventional immortal cell lines, but also to transfect limited proliferation functional Cells - stem cells with low cytotoxicity, as follows:

[0045] Cell transfection steps of Zn-PD2 and Zn-PD4

[0046] 1) Preparation of Zn-PD / DNA complex:

[0047] The DNA used is the green fluorescent protein plasmid pCMV-GFP and the luciferase plasmid pCMV-GLuc. Dissolve Zn-PD2 or Zn-PD4 and DNA in acetate buffer at pH 5.2, mix equal volumes, and let stand at room temperature for 15 minutes to obtain a complex with a Zn-PD / DNA mass ratio of 5-20:1 ;

[0048] 2) In vitro cell transfection experiments:

[0049] The human embryonic kidney cell line 293T, rat bon...

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Abstract

The invention discloses a metal coordination polycation gene vector. The metal coordination polycation gene vector consists of a coordinated metal, a coordinated unit, a stimuli-responsive unit and polycations, and the structural general formula of the metal coordination polycation gene vector is as shown in the specification, wherein M represents the coordinated metal; a dimethyl pyridylamine component represents the coordinated unit; a disulfide bond component represents the stimuli-responsive unit; a ball represents the polycations; n represents the quantity of metal ion ligands grafted by one polycation molecule, n is 1 to 200; and the metal coordination polycation gene vector is used for non-virus gene vectors. The metal coordination polycation gene vector has the advantage that the metal ion ligands containing stimuli-responsive radicals are modified on the polycations, so that the DNA binding capacity of the polycations and the acting force with cytomembranes are greatly improved; after the compound enters cells, the metal ion ligands and the polycations are disconnected, DNAs are released and expressed so as to show extremely high transfection efficiency, so that the application of the polycations in the aspect of the non-virus gene vectors is greatly expanded due to metal coordination monomers in the invention.

Description

technical field [0001] The invention belongs to the field of gene therapy and new materials, in particular to a metal coordination polycation gene carrier and its preparation method and application. Background technique [0002] Gene therapy refers to the process of delivering functional nucleic acid molecules to specific tissue cells through certain means for disease treatment. It can be used for the treatment of many congenital genetic diseases and severe acquired diseases. However, the lack of high-efficiency gene carriers has become the most important obstacle restricting the development of gene therapy, especially the low transfection efficiency of some functional cells (such as stem cells), which seriously hinders the development of gene regulation. Stem cells are a kind of cell population with self-renewal ability and differentiation ability, which have attracted much attention in the fields of tissue engineering and cell therapy. Through gene regulation, stem cells ...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): C08G73/02C08G69/48C08G83/00C12N15/87
CPCC08G69/48C08G73/0213C08G83/004C12N15/87
Inventor 郭天瑛刘帅
Owner NANKAI UNIV
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