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Target type polypeptide nanometer gene carrier compound

A technology of gene carriers and complexes, applied in the field of gene carriers, can solve the problems of non-targeting of carriers, non-specificity of arginine receptors, restriction of targeted transfection, etc., and achieve easy operation, simple process, and improved target effect on ability

Active Publication Date: 2017-06-27
THE NAT CENT FOR NANOSCI & TECH NCNST OF CHINA
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

However, the arginine receptor has no specificity due to the non-targeting of the published vector, which limits its targeted transfection to specific cells and in vivo transfection

Method used

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  • Target type polypeptide nanometer gene carrier compound
  • Target type polypeptide nanometer gene carrier compound
  • Target type polypeptide nanometer gene carrier compound

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0038] Example 1 Preparation of amphiphilic conjugates

[0039] Using dichlorotrityl chloride resin (purchased from Jill Biochemical (Shanghai) Co., Ltd.), and Fmoc-protected L-amino acids (Fmoc-Arg(Pbf)-OH and Fmoc-Gly-OH, purchased from Jill Biochemical (Shanghai) ) Co., Ltd.) as raw materials, under the conditions of the presence of various coupling and cleavage reagents, the polymer of polyarginine is prepared on the resin by solid phase synthesis. After a simple cleavage reaction, the peptide is separated from the resin, and the pure white peptide powder is obtained after subsequent precipitation, washing, and drying.

[0040]The specific experimental process is as follows: First, take 1.01 g of dichlorotrityl chloride resin (purchased from Jill Biochemical (Shanghai) Co., Ltd.) to the peptide synthesis device (purchased from Sigma-Aldrich Company), and add dry N, N -Dimethylformamide (purchased from Sigma-Aldrich Company) soaked the resin for half an hour to make it ful...

Embodiment 2

[0041] Example 2 Preparation of PNP / siRNA / antibody complexes

[0042] The method described in this example is also applicable to the preparation of PNP / DNA / antibody complexes. During preparation, the siRNA in this example can be replaced with the corresponding DNA.

[0043] PNP / siRNA / antibody complexes were prepared according to the following steps:

[0044] (1) Firstly, the amphiphilic conjugate prepared above (molecular weight: 2423) was prepared into a 10 mg / mL stock solution of dimethyl sulfoxide (purchased from Sinopharm Chemical Reagent Beijing Co., Ltd.). Dilute 5 μL of the stock solution in 100 μL of DEPC water, and sonicate for 5 minutes to form uniform and stable polypeptide nanoparticles PNP (ie, the nanogene carrier described above).

[0045] (2) Dissolve negative control siRNA (synthesized by Shanghai Gemma Co., Ltd.) in DEPC water, shake and mix well, and prepare 0.13mg / mL working solution. Immediately add 50 μL siRNA solution to 50 μL polypeptide nanoparticle ...

Embodiment 3

[0049] Embodiment 3 TEM observation morphology

[0050] Take 10 μL of the PNP / siRNA / antibody complex solution prepared in Example 2 for TEM testing. Carefully drop the compound onto a clean copper grid (purchased from Beijing Zhongjing Science and Technology Co., Ltd.), suck up the sample after a while, dry it at room temperature, and stain with uranyl acetate (purchased from Beijing Zhongjing Science and Technology Co., Ltd.) After drying for at least 1 hour at room temperature for 8 minutes, observe with an electron microscope (Hitachi, Japan, model HT7700).

[0051] See the experimental results figure 2 . Depend on figure 2 It can be seen that the PNP / DNA / antibody complex forms spherical nanoscale particles with uniform particle size and good dispersion.

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Abstract

The invention relates to the field of gene carriers, and particularly discloses a target type polypeptide nanometer gene carrier compound. The target type polypeptide nanometer gene carrier compound is prepared from a nano gene carrier with positive charges, a gene medicine and a tumor targeted antibody, wherein the nano gene carrier is adsorbed with the gene medicine and the tumor targeted antibody; and the nano gene carrier is prepared by coupling polypeptides with hydrophobic functional molecules. According to the target type polypeptide nanometer gene carrier compound, the nano gene carrier is formed by self-assembling and carries small interfering nucleic acid; the surface of the nano gene carrier is adsorbed with the tumor tissue targeted antibody; the nano gene carrier with the rich positive charges is used for adsorbing the targeted antibody with negative charges, so that the nano gene carrier with the positive charges can be cleared away in vivo circulation, and the targeting capacity can be also improved. The method is simple in process, mild in reaction conditions and easy to operate. Moreover, no covalent bonds are generated in the self-assembling process, so that no reverse reaction is caused, and highly ordered nano structures can be formed; and the target type polypeptide nanometer gene carrier compound is particularly suitable for constructing a biological medical material.

Description

technical field [0001] The invention relates to the field of gene carriers, in particular to a targeted polypeptide nanometer gene carrier complex. Background technique [0002] Gene therapy is the delivery of genes (mainly DNA or siRNA) into specific cells, thereby promoting or inhibiting the expression of target proteins to achieve the purpose of treating human diseases, because DNA or siRNA is easily degraded by nucleases, and is often negatively charged. It is difficult for it to pass through the negatively charged cell membrane, so it is necessary to use a gene carrier to protect and deliver the nucleic acid into the cell. An ideal gene carrier should ensure high transfection efficiency while having good biocompatibility and low cytotoxicity. In addition, transfection to specific cell types is also one of the characteristics that gene carriers need to possess. Therefore, the development of gene carriers with high efficiency, low toxicity and targeting function is an im...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): A61K48/00A61K47/42A61K47/28A61K31/7088A61P35/00
CPCA61K31/7088A61K47/28A61K47/42
Inventor 赵颖聂广军郎佳妍季天骄丁艳萍
Owner THE NAT CENT FOR NANOSCI & TECH NCNST OF CHINA
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