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Elastin-like polypeptide temperature-responsive nano-material, and preparation method and application thereof

A technology of elastin-like and nanomaterials, which is applied in the field of elastin-like polypeptide temperature-responsive nanomaterials and its preparation and application, and can solve problems such as easy disintegration of micelles, uncontrollable molecular weight, and complex polymer synthesis

Active Publication Date: 2017-11-21
中山华梓生物科技有限公司
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

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Problems solved by technology

However, these GNPs micelles are very unstable, especially in the complex environment in vivo, when the high-molecular interaction force is not enough to stabilize the high-density GNPs, the micelles are easily disintegrated.
At the same time, the synthesis of polymers is complicated, the cost is high, and the molecular weight is uncontrollable, which has great disadvantages in quality control.
Moreover, its biocompatibility and safety are also open to question

Method used

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  • Elastin-like polypeptide temperature-responsive nano-material, and preparation method and application thereof
  • Elastin-like polypeptide temperature-responsive nano-material, and preparation method and application thereof
  • Elastin-like polypeptide temperature-responsive nano-material, and preparation method and application thereof

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Experimental program
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Effect test

Embodiment 1

[0117] Embodiment 1, preparation of elastin-like polypeptide temperature-responsive nanomaterials (ELP-GNPs)

[0118] The elastin-like polypeptide temperature-responsive nanomaterials provided in this example are called ELP-GNPs, which are gold nanoparticles modified by elastin-like polypeptides; the name of the elastin-like polypeptide is ELP, and the amino acid sequence of ELP is as shown in the sequence listing As shown in 1, the repeat unit in the ELP is the 17th-21st amino acid of sequence 1, and the repeat unit in the ELP is repeated 60 times.

[0119] 1. Preparation of elastin-like polypeptide (ELP)

[0120] 1. Preparation of recombinant vector: use the method of PRe-RDL (McDaniel et al. Reconstruction Allows Rapid and Seamless Cloning of Oligomeric Genes, Biomacromolecules) was inserted into the DNA fragment shown in Sequence 2, and other sequences of the vector were kept unchanged to obtain a recombinant vector, which was named pET-24-ELP.

[0121] 2. Preparation of...

Embodiment 2

[0145] Example 2, ELP-GNPs have the effect of photothermal therapy to kill tumor cells

[0146] Human C8161 melanoma cells (C8161-GFP cells) transfected with green fluorescent protein (green fluorescent protein, GFP) were selected (Jolanta M Topczewska et al., Embryonic and tumorigenic pathways converge via Nodal signaling: role in melanoma aggressiveness, NATUREMEDICINE, VOLUME 12, NUMBER 8, AUGUST 2006) to test the effect of ELP-GNPs photothermal therapy on killing tumor cells by fluorescent staining and MTT method. The experiment was repeated three times, and the specific steps of each repeated experiment were as follows:

[0147] The revived C8161-GFP cells were cultured in culture medium 1 (medium 1 was obtained by adding FBS, penicillin and streptomycin to DMEM / F-12 medium, the mass percentage concentration of FBS was 10% FBS, and the concentration of penicillin was 200 U / mL , streptomycin concentration of 200 μg / mL medium), 5% CO 2 , and cultivated in a 37°C environme...

Embodiment 3

[0150] The cytotoxicity of embodiment 3, ELP-GNPs

[0151] Mouse 3T3 embryonic fibroblasts (3T3 cells) (Cell Bank of the Chinese Academy of Sciences, GNM 3) and human microvascular endothelial cells (HMEC-1 cells) (American Type Culture Collection, ATCC-CRL-3243) were selected, and the MTT assay was used. Test the toxicity of ELP-GNPs samples. The experiment was repeated three times, and the specific steps of each repeated experiment were as follows:

[0152] Inoculate 3T3 cell suspension in a 96-well plate (50 μL / well, 5000 cells / well, 3T3 cell suspension was obtained by resuspending 3T3 cells in medium 2, which was added FBS, penicillin and chain to DMEM medium The mass percentage concentration of FBS obtained by using 10% FBS, the concentration of penicillin is 100 U / mL, and the concentration of streptomycin is 100 μg / mL culture medium) and cultivated until adherent, and the medium is aspirated before the experiment. The ELP-GNPs of Example 1 were serially diluted with me...

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Abstract

The invention discloses an elastin-like polypeptide temperature-responsive nano-material, and a preparation method and application thereof. The nano-material disclosed by the invention is a material prepared by modifying a nanoparticle with an elastin-like polypeptide. The elastin-like polypeptide has responsiveness to a tumor microenvironment and is A1) or A2), wherein A1) refers to a protein with an amino acid sequence as shown in a sequence 1 in a sequence table, A2) refers to a protein which is obtained by inserting a cysteine residue or a polypeptide containing the cysteine residue into the amino acid sequence of a protein P and has same functions, and the protein P is prepared by subjecting a polypeptide as shown in site 17 to 21 in the sequence 1 to repetition 30 to 100 times. The nanoparticle has surface plasmon resonance effect and is a gold nanoparticle, silver nanoparticle, platinum nanoparticle or gold-silver nanoparticle. Experimental results show that the nano-material provided by the invention has a high photothermal conversion rate and good tumor inhibitory effect and can be used for treating tumors.

Description

technical field [0001] The invention relates to an elastin-like polypeptide temperature-responsive nano material in the field of biomedicine, a preparation method and application thereof. Background technique [0002] Photothermal therapy for tumors (photothermal therapy, PTT), as a method of using local hyperthermia to kill tumors, has been studied for a long time, and there are many kinds of nanomaterials that can be used for near-infrared laser hyperthermia. Including: organic compounds such as ICG, polymers such as polypyrrole, carbon nanomaterials (carbon nanotubes, C 60 ) and other conjugated nanosystems; inorganic materials such as gold nanomaterials (gold nanoshells, gold nanorods, gold nanostars), palladium nanoparticles, etc. These nanomaterials themselves have near-infrared absorption properties and can be directly used in photothermal therapy. However, they are either toxic or have complex and uncontrollable reaction processes, making them difficult to apply cl...

Claims

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Application Information

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IPC IPC(8): C07K14/78C07K1/113A61K41/00A61K47/62A61P35/00
CPCA61K41/0057C07K14/78
Inventor 高卫平孙蒙蒙
Owner 中山华梓生物科技有限公司
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