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Novel cell-permeable peptide capable of mediating drug delivery, and applications thereof

A technology of membrane-penetrating peptides and drugs, which is applied in the field of membrane-penetrating peptides and the application of the membrane-penetrating peptides in drug delivery and tumor treatment. Effect of small molecular weight and low immunogenicity

Inactive Publication Date: 2017-12-01
INST OF PROCESS ENG CHINESE ACAD OF SCI
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

However, there are common problems such as low import efficiency, difficulty in popularization, and easy to cause cell damage or even death.
All in all, there is still a lack of an efficient and feasible method for direct introduction of therapeutic drugs

Method used

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  • Novel cell-permeable peptide capable of mediating drug delivery, and applications thereof
  • Novel cell-permeable peptide capable of mediating drug delivery, and applications thereof
  • Novel cell-permeable peptide capable of mediating drug delivery, and applications thereof

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0019] Example 1: Synthesis of Penetrating Peptides

[0020] 1) Activation of the resin: Weigh 1000mg of Fmoc-Trp wang Resin or Fmoc-Pro wang Resin, add 10-15ml (immerse the entire resin) in DMF and soak for 30min to make it fully swell.

[0021] 2) Deprotection: soak the DMF of the resin to remove it by pressure filtration, add 10ml of DMF solution containing 20% ​​piperidine, react with nitrogen blowing and boiling for 15min, then remove it by pressure filtration, remove the FMOC group of the amino group, and wash the resin with 10ml of isopropanol Three times, and then wash three times with 10ml DMF, and then use the ninhydrin method to detect that the resin should turn black or purple.

[0022] 3) Condensation reaction: connect the next amino acid, weigh the amount of Fmoc-amino acid to be 1.4mmol / g resin, 910mg TBTU, add 10ml DMF and 0.45g HOBt and mix evenly, add 0.52ml DIEA to make the reaction solution, and blow it with nitrogen at room temperature Boiling reaction 2h...

Embodiment 2

[0027] Implementation Example 2: Cytotoxicity Experiment

[0028] 1) Take a 96-well plate and add 7×10 3 A cell culture medium, 37 ℃, 5% carbon dioxide incubator for 24 hours, so that the cells adhere to the wall.

[0029] 2) Prepare medium containing different concentrations of penetrating peptides, use medium without penetrating peptides as negative control wells (Control), and incubate at 37° C. with 5% carbon dioxide for 1-5 hours.

[0030] 3) Add 20 μl MTT to each well of the adherent cells, continue to incubate for 24 hours, discard the culture medium, add 150 μl DMSO (dimethyl sulfoxide) to each well, and shake for 10 minutes.

[0031] 4) Select a wavelength of 490nm or 570nm, and calculate the cell survival rate by the light absorption value on the microplate reader immunodetector. see results figure 1 .

[0032] The results showed that the penetrating peptide had no obvious cytotoxicity.

Embodiment 3

[0033] Implementation Example 3: Membrane-penetrating peptide and fluorescence detection experiment

[0034] 1) Add a medium containing a certain concentration of FITC fluorescently labeled penetrating peptide to the adherent cells, incubate for 30min-4h, then carefully suck off the medium, and wash the adherent cells three times with PBS.

[0035] 2) Add fixative, 10min-20min, fix the cells and wash the cells 2-3 times with PBS.

[0036] 3) Add DAPI staining solution to stain for 5min-10min, then wash with PBS 2-3 times.

[0037] 4) Observe with a fluorescence microscope under the excitation light of 488nm and 543nm respectively, the results are shown in figure 2 .

[0038] The results showed that the membrane-penetrating peptide successfully penetrated the membrane and delivered the fluorescent marker into the cell.

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Abstract

The invention discloses a novel cell-permeable peptide capable of efficiently mediate the delivery of drugs into cells, and further provides applications of the novel cell-permeable peptide in treatment of tumors.

Description

technical field [0001] The present invention relates to the field of biotechnology, more specifically, to a membrane-penetrating peptide and the application of the membrane-penetrating peptide in drug delivery and tumor treatment. Background technique [0002] Malignant tumors, commonly referred to as cancer, are abnormal growths of cells in the body that can invade adjacent parts of the body and spread to other organs, eventually leading to death. Tumor has become one of the most serious diseases threatening human life. In the current clinical treatment methods, chemotherapy, which uses chemical drugs to kill tumor cells and inhibit the proliferation of tumor cells, is the main means. However, many tumor therapeutic drugs cannot freely penetrate the cell membrane, so the delivery process of therapeutic drugs has become a key link in chemotherapy. [0003] At present, there are two main methods in the drug delivery process. One is the nano drug delivery system, including l...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): C07K7/06A61K47/42A61K49/00A61K31/337A61P35/00
CPCA61K31/337A61K47/42A61K49/0045C07K7/06
Inventor 徐霞韦宇平
Owner INST OF PROCESS ENG CHINESE ACAD OF SCI
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