Supramolecular nano-drug for activating Hippo pathway as well as preparation method and application of supramolecular nano-drug

A nano-drug and supramolecular technology, applied in the field of nano-biomedical materials, can solve the problems of weak intracellular retention ability and weak tumor penetration ability, achieve good biocompatibility, easy clinical transformation, and improve solubility

Active Publication Date: 2022-07-26
INST OF RADIATION MEDICINE CHINESE ACADEMY OF MEDICAL SCI
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

Studies have shown that spherical nanoparticles can diffuse and penetrate better in tumor sites, but their intracellular retention ability is relatively weak; while nanorods or nanofibers with high aspect ratios can obtain higher accumulation in cells, but their Its tumor penetration ability is relatively weak

Method used

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  • Supramolecular nano-drug for activating Hippo pathway as well as preparation method and application of supramolecular nano-drug
  • Supramolecular nano-drug for activating Hippo pathway as well as preparation method and application of supramolecular nano-drug
  • Supramolecular nano-drug for activating Hippo pathway as well as preparation method and application of supramolecular nano-drug

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0042] Preparation and characterization of supramolecular nanomedicines, including Fmoc-S(FA)FFYSV-SS-PEG (abbreviated as FASP), control material Fmoc-SFFYSV-SS-PEG without FA (abbreviated as FSSP), without histone deacetylation The control material Fmoc-S(FA)FFY-SS-PEG (abbreviated as FAYP) of the enzyme inhibitory peptide YSV, and the control material Fmoc-S(FA)FFYSV-CC-PEG (abbreviated as FAYP) without glutathione response FACP), taking FASP nanomedicine as an example, the specific preparation steps are as follows:

[0043] (1) Weigh 5 mg of pure polypeptide, dissolve it in 1 mL DMSO, and slowly add it dropwise to 9 mLddH in a ratio of 1:9. 2 In O / PBS, a drop of DMSO solvent containing peptides was added dropwise every 30 s, while stirring; after the dropwise addition, continued stirring at room temperature for 4 h to stabilize the micelles; the obtained product was placed in a dialysis bag (MWCO=1000 D) After dialysis for 48 h, the DMSO solvent was completely removed to o...

Embodiment 2

[0049] The growth inhibitory effects of the nano-drugs FASP, FSSP, FAYP, FACP and free drug FA prepared in Example 1 on tumor cells and normal cells in vitro were evaluated, and the specific implementation steps were as follows:

[0050] 1) Take MDA-MB-231, 4T1 and 3T3 cells in the logarithmic growth phase, and inoculate 6000 cells per well into a 96-well plate at 37°C, 5% CO 2 Cultured in a constant temperature incubator for 24 h;

[0051] 2) Dilute the nano-drugs FASP, FAYP, FACP, FSSP and free drug FA to a preset concentration with culture medium and incubate with the cells for 48 h;

[0052] 3) Add 10 μL of CCK-8 solution to each well under the dark condition, put it in the incubator for 2-4 hours, use a microplate reader to detect the absorbance at 450 nm, and calculate the different concentrations of each group. The survival rate of each cell after drug action;

[0053] 4) figure 2 It is shown that the nanomedicine FASP can significantly improve the growth inhibitory...

Embodiment 3

[0055] (1) Fluorescence quantitative PCR analysis was performed on the regulation of the key target genes of the Hippo pathway by the supramolecular nano-drugs and free drugs prepared in Example 1. The specific steps are as follows:

[0056]1) Take MDA-MB-231 cells in logarithmic growth phase and inoculate them in a 6 cm culture dish at a density of 100,000 per well, at 37°C, 5% CO 2 Cultured in a constant temperature incubator for 24 h;

[0057] 2) Discard the original medium and add fresh medium containing 50 μM of nano-drugs and free drugs, and incubate with cells for 48 h in the incubator;

[0058] 3) Wash the cells twice with PBS, and trypsinize to collect the cells;

[0059] 4) Wash once with PBS, centrifuge at 3000 rpm for 5 min, and discard the supernatant;

[0060] 5) Add 1 mL of 4°C pre-cooled Trizol reagent, and lyse at room temperature for 5 min;

[0061] 6) Add 200 μL of chloroform and let stand for 3 minutes;

[0062] 7) 13500 rpm, 15 min;

[0063] 8) Take t...

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Abstract

The invention discloses a supramolecular nano-drug for targeted activation of a Hippo pathway in cancer cells and application of the supramolecular nano-drug. The amphiphilic compound Fmoc-S (FA) FFYSV-SS-PEG (FASP) is obtained by connecting a hydrophobic oligopeptide fragment and a hydrophilic polyethylene glycol fragment, which contain double drug components, through a disulfide bond, and the amphiphilic compound Fmoc-S (FA) FFYSV-SS-PEG (FASP) is obtained. The compound can be self-assembled through intermolecular non-covalent interaction to form an inactive nano prodrug, the nano prodrug is subjected to morphological transformation after being taken by cancer cells, bioactive peptide nanofibers are formed in situ, and flufenamic acid (FA) is released, so that a disordered Hippo pathway is activated to be used for tumor chemoradiotherapy combined treatment and metastasis inhibition, and the compound has the characteristics that the synthesis is simple, the cost is low, and the compound is suitable for large-scale industrial production. The repeatability is high, the biocompatibility is good, and the clinical transformation potential is great. Meanwhile, compared with free FA, the supramolecular nano-drug obtained by the invention can be delivered through covalent linkage and released in situ in a responsive manner, so that the growth inhibition effect of FA on tumor cells is remarkably improved, the toxic and side effects of FA on normal cells are reduced, and the supramolecular nano-drug has good clinical transformation and application prospects.

Description

technical field [0001] The invention belongs to the field of nanometer biomedical materials, and relates to a preparation method of a supramolecular nanomedicine for targeting and activating Hippo pathway in cancer cells and its application in the combined treatment of tumor radiotherapy and chemotherapy. Background technique [0002] The Hippo pathway was first discovered in Drosophila and is highly conserved in mammals. It has been one of the research hotspots in the field of life medicine since its discovery. Studies have shown that the Hippo pathway plays a key role in a wide range of biological processes, including organ size control, cell proliferation, cancer development, and virus-induced diseases. The Hippo pathway is mainly composed of three parts, namely the upstream core kinase MST-LATS module, the YAP protein, a key factor in signal cascade transmission, and the downstream transcription factor family TEADs module. When the Hippo pathway is activated by upstream...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): A61K47/69A61K47/60A61K31/196A61K38/08A61K41/00A61P35/00A61P35/04
CPCA61K47/6907A61K47/60A61K41/0038A61K31/196A61K38/08A61P35/00A61P35/04Y02P20/55
Inventor 刘鉴峰任春华王志龙杨翠红王忠彦杨丽军肖萌张嘉敏
Owner INST OF RADIATION MEDICINE CHINESE ACADEMY OF MEDICAL SCI
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