Macrocyclic polyamine amphiphile compound based on green fluorescent protein chromophore bi and its preparation method and use

A green fluorescent protein and macrocyclic polyamine technology, which is applied in the field of macrocyclic polyamine amphiphiles and their preparation, can solve the problems of low transfection efficiency, single performance, and high toxicity

Active Publication Date: 2020-05-19
BEIJING NORMAL UNIVERSITY
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

However, there are still many problems in the clinical treatment of traditional non-viral delivery systems, such as compound accumulation in the body, endosome escape, single performance, high toxicity, and low transfection efficiency

Method used

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  • Macrocyclic polyamine amphiphile compound based on green fluorescent protein chromophore bi and its preparation method and use
  • Macrocyclic polyamine amphiphile compound based on green fluorescent protein chromophore bi and its preparation method and use
  • Macrocyclic polyamine amphiphile compound based on green fluorescent protein chromophore bi and its preparation method and use

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0046] A kind of macrocyclic polyamine amphiphile compound based on green fluorescent protein chromophore B1, the structural formula (I) of said compound is as follows:

[0047]

[0048] In formula (I), R is a hydrocarbyl structural unit.

[0049] Wherein, when R is a linear alkyl structural unit, and R is , the synthesized macrocyclic polyamine amphiphile based on the green fluorescent protein chromophore BI is denoted as BI-A; R is The synthesized macrocyclic polyamine amphiphile based on the green fluorescent protein chromophore BI is denoted as BI-B; R is The synthesized macrocyclic polyamine amphiphile based on the green fluorescent protein chromophore BI is denoted as BI-C; R is The synthesized macrocyclic polyamine amphiphile based on the green fluorescent protein chromophore BI is denoted as BI-D. Concrete synthetic route is as follows:

[0050]

[0051] In the formula, (i) bromoalkane RBr, potassium carbonate, acetone, reflux, 48 hours; (ii) 3,5-bis (azi...

Embodiment 2

[0070] Prepare solutions of different concentrations of compound BI-A~BI-D respectively, place different concentrations of BI-A~BI-D and pUC18 plasmid DNA (9μg / mL) in a 37°C water bath, incubate for 1h, and then carry out DNA agarose Gel retardation experiment The gel retardation results of different concentrations of compounds on pUC18 DNA were obtained.

[0071] figure 1 A~1H are the results of agarose gel retardation experiments on pUC18 DNA by compounds BI-A~BI-D and BI-A / DOPE~BI-D / DOPE (the ratio of compound to DOPE is 1:3) of the present invention, respectively; figure 1 The value marked above in A~1H is the test concentration (μM); figure 1 A~1H shows that compounds BI-A~BI-D and BI-A / DOPE~BI-D / DOPE (the ratio of compound to DOPE is 1︰3) can completely block the migration of DNA in agarose at lower concentrations .

[0072] It can be concluded from Example 2 that the amphiphilic compound based on the BI unit prepared in the present invention can effectively condense ...

Embodiment 3

[0074] Add ctDNA to the solution of cationic liposome BI-A / DOPE, BI-B / DOPE, BI-C / DOPE and BI-D / DOPE (the ratio of compound to DOPE is 1:3), test its fluorescence intensity and drawing, get figure 2 A~2D.

[0075] figure 2 A~2D are the fluorescence titration results of ctDNA to the complexes BI-A / DOPE, BI-B / DOPE, BI-C / DOPE and BI-D / DOPE (the ratio of compound to DOPE is 1︰3); among them, the X axis is the concentration of DNA, and the Y axis represents the fluorescence intensity. It can be concluded from Example 3 that the present invention artificially simulates the chromogenic process of green fluorescent protein.

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Abstract

The invention provides a macrocyclic polyamine amphipathic compound based on green fluorescent protein chromophore BI and a preparation method and application thereof. The compound provided by the invention is mainly prepared through carbonyl ammonia condensation, a 1,3 dipolar-cycloaddition reaction, an esterification reaction and a Click reaction. The compound provided by the invention has goodtwo-photon fluorescence properties, and has the advantages of being long in emission wavelength, high in 3D resolution ratio, low in self-illumination, and the like; chromophore mechanisms of green fluorescent proteins are artificially simulated; the compound provided by the invention is capable of forming a pH value response small molecule non-viral genetic vector together with dioleyl phosphatidylethanolamine (DOPE), then release of DNA (Deoxyribonucleic Acid) is promoted, and the compound has very high transfection efficiency and is capable of targeting cell nucleuses.

Description

technical field [0001] The invention relates to a non-viral gene carrier, in particular to a macrocyclic polyamine amphiphile compound based on the green fluorescent protein chromophore BI and its preparation method and application. Background technique [0002] Gene therapy is to repair or replace defective genes by introducing normal genes (eg, DNA, siRNA, mRNA and shRNA) into diseased cells. In recent decades, gene therapy, as a promising treatment method, has attracted widespread attention of scientific researchers. [0003] Vectors are divided into two categories: viral gene carriers and non-viral gene carriers. Viral gene vectors can transfect low-dose DNA very efficiently, but viral gene vectors also have certain risks of carcinogenicity and immunogenicity, limited vector capacity (usually 2-3kb) and high operational requirements. In order to solve these problems, researchers have developed a variety of non-viral gene vectors as virus substitutes. Non-viral gene ca...

Claims

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Application Information

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Patent Type & Authority Patents(China)
IPC IPC(8): C07D403/14C09K11/06C12N15/87
CPCC07D403/14C09K11/06C09K2211/1007C09K2211/1044C09K2211/1059C12N15/87
Inventor 卢忠林刘名轩马乐乐刘旭英
Owner BEIJING NORMAL UNIVERSITY
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