Preparation of anti-cancer lipopeptide and application of anti-cancer lipopeptide in anti-tumor treatment

A lipopeptide and resin technology, which is used in the preparation of anticancer lipopeptides and the application field of antitumor therapy, can solve the problems of reducing hemolysis, low bioavailability, and reducing hydrophobic torque.

Pending Publication Date: 2022-02-08
BINZHOU MEDICAL COLLEGE
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

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Problems solved by technology

In addition, the increase of the hydrophobic amino acid of the polypeptide can improve the anticancer activity of the polypeptide. However, high hydrophobicity usually produces high hemolysis. By introducing hydrophilic residues on the hydrophobic side of the α-helical amphiphilic peptide, the hydrophobic moment is reduced, thereby reducing hemolytic
[0003] The shortcomings of peptide drugs such as short half-life, low bioavailability, poor stability, and easy hydrolysis by proteases limit their clinical application.

Method used

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  • Preparation of anti-cancer lipopeptide and application of anti-cancer lipopeptide in anti-tumor treatment
  • Preparation of anti-cancer lipopeptide and application of anti-cancer lipopeptide in anti-tumor treatment
  • Preparation of anti-cancer lipopeptide and application of anti-cancer lipopeptide in anti-tumor treatment

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0022] Embodiment 1: the preparation method of anticancer lipopeptide

[0023] The synthesis of the anticancer lipopeptide of the present invention is carried out based on the Fmoc-protected solid-phase polypeptide synthesis method. The sequence of the anticancer lipopeptide is C 8 h 15 O-Asp-Ser-Asp-Val-Trp-Trp-Gly-Gly-Arg-Arg-Leu-Leu-Arg-Arg-Leu-Arg-Arg-Leu, the specific synthesis steps are as follows:

[0024] (1) Fmoc-Asp(OtBu)-Ser(tBu)-Asp(OtBu)-Val-Trp(Boc)-Trp(Boc)-Gly-Gly-Arg(Pbf)-Arg(Pbf)-Leu-Leu- Synthesis of Arg(Pbf)-Arg(Pbf)-Leu-Arg(Pbf)-Arg(Pbf)-Leu-WangResin.

[0025] Weigh a certain amount of Fmoc-Leu-Wang Resin with a loading capacity of 0.535mmol / g and place it in a beaker, add anhydrous DMF to swell the resin for 30min. Take a small amount of the above resin for ninhydrin detection, if it is colorless, continue. According to the ratio of DMF: piperidine = 4:1, add piperidine solution, stir magnetically for 30 minutes to remove Fmoc group, perform suction...

Embodiment 2

[0028] Example 2: In vitro anticancer activity assay of anticancer lipopeptides

[0029] The in vitro anticancer activity of anticancer lipopeptides was tested by MTT assay. Select HepG2 cells in the logarithmic growth phase, inoculate the HepG2 cells into a 96-well plate, and then place the 96-well plate in 5% CO 2 , in an incubator at 37°C, and after 24 hours, when the cells covered the bottom of the well, the old medium was sucked out, the lipopeptide was prepared into different concentrations with DMEM medium, and 100 μL of drug-containing medium was added to each well. Reflecting situation 4 is true and reliable, and 5 parallel experiments are set for each concentration to ensure the reliability of the experiment. After culturing for 24 h, add 20 μL (5 mg·mL -1 ) The MTT solution was placed in a cell culture incubator for 4 hours. Discard the supernatant, then add 150 μL DMSO to each well to dissolve the blue-purple MTT crystals, wrap the 96-well plate with tin foil an...

Embodiment 3

[0030] Example 3: Determination of the secondary structure of anticancer lipopeptides

[0031] The secondary structure of lipopeptide in water and SDS solution was detected by circular dichroism spectrometer. The lipopeptide was prepared into an aqueous solution with a final concentration of 150 μM and a sodium dodecyl sulfate (SDS) solution (30 mM) with a final concentration of 150 μM, wherein the aqueous solution simulated a hydrophilic environment, and the SDS solution was used to simulate a hydrophobic environment such as a cell membrane. Quartz cuvette (optical path length is 0.5mm), test with circular dichroism spectrometer, wavelength range is set as 180-260nm, the measured data is converted into average residue ellipticity with the spectrum obtained by the following formula:

[0032] θ M = θ obs ×1000 / cln

[0033] where θ M is the average residue ellipticity; θ obs is the ellipticity obtained by buffer correction at a given wavelength; c represents the sample conc...

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Abstract

The invention relates to a preparation method of an anti-cancer lipopeptide C8H15O-Asp-Ser-Asp-Val-Trp-Trp-Gly-Gly-Arg-Arg-Leu-Leu-Arg-Arg-Leu-Arg-Arg-Leu, and an application of the anti-cancer lipopeptide C8H15O-Asp-Ser-Asp-Val-Trp-Trp-Gly-Arg-Arg-Leu in anti-tumor treatment, and belongs to the field of biological medicines. The anti-cancer lipopeptides are prepared based on key biological characteristics (positive charges, alpha-helical structure and amphipathy) of membrane lysis peptides. The invention provides a solid-phase synthesis method of the anticancer lipopeptide. The anti-cancer lipopeptide has the characteristics of small hydrophobic torque, low hemolytic activity and good serum stability, can effectively overcome the defect of high hemolytic toxicity of most anti-cancer peptides, and solves the problem of poor in-vivo stability of peptide drugs. An in-vitro anti-cancer experiment and an in-vivo anti-tumor experiment prove that the anti-cancer lipopeptide has a good anti-tumor effect and a good application prospect.

Description

technical field [0001] The invention belongs to the field of biomedicine, and specifically relates to the preparation of an artificially synthesized anticancer lipopeptide with excellent in vivo and in vitro anticancer activities and its application in antitumor therapy. Background technique [0002] Despite great advances in the treatment of cancer, it remains one of the deadliest diseases in humans. At present, chemotherapy and radiotherapy are mainly used to treat cancer, but the treatment is often accompanied by severe side effects and multidrug resistance. Anticancer peptides (ACPs) have the advantages of high selectivity, good biocompatibility, and high tumor permeability. Most ACPs are α-helical cationic amphiphilic peptides, whose activity is mainly determined by net charge, hydrophobicity, and helicity. Compared with normal cells, the surface of cancer cell membrane carries a large amount of anionic components such as phosphatidylserine and phosphatidylinositol, w...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): C07K7/08C07K1/08C07K1/06C07K1/04A61K38/10A61P35/00
CPCC07K7/08A61P35/00A61K38/00Y02P20/55
Inventor 闫苗苗魏光成霍业鸿
Owner BINZHOU MEDICAL COLLEGE
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