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A lipoic acid-modified inherently disordered protein nanocarrier and its preparation method and application

A nano-carrier, lipoic acid technology, applied in the field of medicine, to achieve low cytotoxicity, good ability to co-carry genes and chemotherapy drugs, and enhance sensitivity

Active Publication Date: 2020-08-07
SHANGHAI WEI ER BIOPHARM TECH CO LTD +3
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

[0014] There is currently no lipoic acid-modified intrinsically disordered protein nanocarrier that can target breast cancer as a nanodelivery carrier

Method used

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  • A lipoic acid-modified inherently disordered protein nanocarrier and its preparation method and application
  • A lipoic acid-modified inherently disordered protein nanocarrier and its preparation method and application
  • A lipoic acid-modified inherently disordered protein nanocarrier and its preparation method and application

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0072] Example 1: Synthesis of 17 peptides modified by lipoic acid

[0073] Lipoic acid (LA) modified 17 peptide amino acid sequence: Lys Val Arg Val Arg Val Arg Val D ProThr Arg Val Arg Gln Arg Val Lys (Lys: Lysine, Val: Valine, D PRO: D-proline, Thr: threonine, Arg: arginine, Gln: glutamic acid) (SEQ ID NO: 1), LA-CL, from Shanghai Gil Biochemical Co., Ltd. using peptide solid-phase synthesis method It was synthesized and named as LA-CL, and the synthesized LA-CL was purified by preparative high-performance liquid chromatography, and its purity reached more than 95%. LA- is lipoic acid, CL: the sequence is KVRVRVRV D PTRVRERVK. Wherein K: lysine; V valine; R: arginine; D P: D-proline; T: threonine; E: glutamic acid, amino acids are connected by peptide bonds to form 17 peptides.

Embodiment 2

[0074] Example 2: Preparation of lipoic acid-modified polypeptide nanocarrier LA-CLss

[0075] Take 50 mg lipoic acid-modified polypeptide LA-CL and different amounts of cysteine ​​hydrochloride dissolved in 10 ml of methanol, stirring and reacting for 12 hours, the reaction temperature is 10-30°C. The amounts of cysteine ​​are: 2.5%, 5%, 10%, and 20%, respectively.

[0076] As shown in Table 1, LA-CLss with different molecular weights were prepared according to the ratio of LA-CL to cysteine. The resulting solution was dialyzed for 12 hours through a dialysis bag with a molecular weight cut-off of 1000, and the dialysate was distilled water, and the dialysate was changed every 4 hours. The obtained dialysis product was lyophilized and stored at -20°C, and the lyophilized carrier could be stored for a long time under reconstitution conditions. The synthesized carrier was subjected to H NMR spectroscopy 1 H-NMR (600M) detection, molecular weight through gel permeation chroma...

Embodiment 3

[0081] Example 3: Preparation of LA-CLss in pDNA nanomicelles

[0082] Dissolve the vector (LA-CLss) and the luciferase expression plasmid pEGFP (Shanghai Innovation Biotechnology Co., Ltd.) in water to prepare an aqueous solution, and the ratio of nitrogen to phosphorus (N / P) is 5, 10, 20, 40, 80, respectively. The nanocomposites were prepared and vortexed for 10 seconds, then left to stand for 30 minutes to obtain nanomicelles. The average particle size of nanomicelles is related to N / P. When N / P=40, the optimal particle size is obtained, and the particle size is between 100-300. See Figure 2 for details. The Zeta potential of nanomicelles increases with the increase of N / P, and is stable at 0-30mV when N / P is greater than 5. For details, see image 3 .

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Abstract

The invention relates to the technical field of medicines, in particular to a lipoic acid modified intrinsically disordered protein nano-carrier, a preparation method thereof and an application of the nano-carrier. The nano-carrier is crosslinked by disulfide bonds of lipoic acid, formed polypeptide polymer can be rapidly degraded in cells and cannot be accumulated in the cells, and amino acid forming a polypeptide carrier exists in vivo and has no toxic and side effects on the cells and a human body. CCK-8 cell proliferation assay indicates that the prepared nano-carrier has low cell toxicity and good gene and chemotherapeutic drug co-carrier capacity, can successfully deliver sensitivity of autophagic specificity enhanced cancer cells to chemotherapeutic drugs caused by siRNA suppressive chemotherapeutic drugs in breast cancer therapy, promotes breast cancer cell apoptosis and accordingly becomes a targeted, efficient and low-toxicity nano-scale delivery system in breast cancer therapy.

Description

technical field [0001] The invention relates to the technical field of medicine, in particular to a lipoic acid-modified inherently disordered protein nanocarrier and a preparation method and application thereof. Background technique [0002] Breast cancer is one of the malignant tumors that threaten women's health. At present, surgery is still the main treatment for breast cancer, combined with radiation therapy and chemotherapeutic drug therapy (chemotherapy for short). Chemotherapy plays a key role in the treatment of breast cancer. , but conventional chemotherapy and radiotherapy drugs are not selective and inevitably damage normal cells. Therefore, the development of targeting vectors for tumor cell therapy can greatly improve the effect on tumors. The introduction of nanotechnology not only realizes the targeted treatment of tumors, but also reduces the toxic and side effects of chemotherapy drugs. Therefore, it is of great significance to prepare a nano drug deliver...

Claims

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Application Information

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Patent Type & Authority Patents(China)
IPC IPC(8): C07K7/08C07K1/02A61K9/107A61K48/00A61K47/42A61K31/7088A61K31/337A61P35/00
CPCA61K9/1075A61K31/337A61K31/7088A61K47/42C07K7/08C07K19/00A61K2300/00
Inventor 管斐宫春爱高原武鑫
Owner SHANGHAI WEI ER BIOPHARM TECH CO LTD
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