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Polypeptide drug against hepatitis B virus X protein (HBx)

A hepatitis B virus and drug technology, which is applied in the direction of viral peptides, antiviral agents, and antitumor drugs, can solve the problems of short half-life of polypeptide fragments, affecting pharmacodynamic effects, and difficulty in discovering polypeptide drugs, and achieve obvious drug discovery. The effect of dynamism

Active Publication Date: 2012-09-19
TIANJIN TOPTECH BIO SCI & TECH
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

However, the discovery of peptide drugs is difficult because most of the peptide fragments have a short half-life in the body, which directly affects the pharmacodynamic effect.

Method used

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  • Polypeptide drug against hepatitis B virus X protein (HBx)
  • Polypeptide drug against hepatitis B virus X protein (HBx)
  • Polypeptide drug against hepatitis B virus X protein (HBx)

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0100] Synthetic polypeptide functional fragment anti-HBxP1#:

[0101] The present invention has synthesized the polypeptide (hereinafter referred to as anti-HBxP1) whose amino acid sequence is Gly-Ser-Ala-Val-Met-Phe-Ser-Ser-Lys-Glu-Arg-Gly (SEQ ID NO: 1) by means of artificial synthesis. #). The preparation of the polypeptide adopts a solid-phase synthesis method, such as using an AAPPTECAPex396 polypeptide synthesis instrument (purchased from Hong Kong Universal Analytical and Testing Instrument Co., Ltd.), in a closed explosion-proof glass reactor to make amino acids according to the sequence shown in SEQ ID NO: 1, From C-terminus-carboxyl-terminus to N-terminus-amino-terminus, this refers to the first amino acid to be added to the amino acid sequence Gly-Ser-Ala-Val-Met-Phe-Ser-Ser-Lys-Glu-Arg-Gly The monomer is C-terminal Gly, then Arg, then Glu, until the last Ser and N-terminal Gly, which are continuously added, reacted, synthesized, and operated to finally obtain the...

Embodiment 2

[0109] Two methods are used to detect the anti-HBx activity of the polypeptide in Example 1 in vitro: the first method is to use molecular cloning technology to clone the cDNA expressing the polypeptide in Example 1 into the eukaryotic expression vector pcDNA3.1 On (+), through gene transfection, the purpose of expressing the studied polypeptide in liver cancer cells is realized, and then the effect of the researched polypeptide on inhibiting HBx is observed; the second method is to use artificially synthesized polypeptides and directly add them to cultured liver cancer cells In the culture medium, observe the effect of polypeptide on inhibiting HBx.

[0110] There are two kinds of liver cancer cells used in the experiment: one is liver cancer HepG2-X cells constitutively expressing HBx (hepatoma HepG2 cells stably transfected with HBx); the other is liver cancer HepG2.2.15 cells constitutively expressing the whole gene of hepatitis B virus (hepatoma HepG2 cells stably transfe...

Embodiment 3

[0190] HepG2-X cells or HepG2.2.15 cells in the logarithmic growth phase were digested with trypsin to make a cell suspension, the number of cells was calculated, and the cells were diluted to 1×10 with sterile normal saline. 7 cells / ml and stored in ice water. Twelve 4- to 6-week-old female BALB / C nude mice were then randomly divided into 2 groups: ① Control group, subcutaneously inject 0.2ml of the above-mentioned diluted cells in the armpit of the right forelimb of each mouse, and only inject 0.5 ml sterilized distilled water (without polypeptide drugs); ② experimental group (administration dose is 10 mg / kg body weight). Subcutaneously inject 0.2ml of the above-mentioned diluted cells into the armpit of the right forelimb of each mouse, and after 7 days of injection, the tumor volume (V=L×W 2 ×0.5) up to 100mm 3 , and then subcutaneously inject the above polypeptide drugs (dissolve the freeze-dried polypeptide drugs with 0.5ml sterilized distilled water), inject once ever...

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Abstract

The invention relates to the field of polypeptide drugs, and in particular relates to polypeptide against hepatitis B virus X protein (HBx), polynucleotide coding the polypeptide and application of the polypeptide and the polynucleotide. In particular, the invention relates to polypeptide with the functional activity of inhibiting HBx. The polypeptide has the activity of inhibiting HBx at molecular level, cellular level and animal level; therefore, the polypeptide can inhibit hepatitis caused by HBV infection, liver cirrhosis caused by recurrent attacks of hepatitis and liver cancers occurring on the basis of liver cirrhosis. The polypeptide and peptide mimics thereof comprise own functional fragments and functional varieties and genes coding the polypeptide and the peptide mimics or the functional fragments and functional varieties of the polypeptide and the peptide mimics and can be widely used for preventing and treating liver diseases caused by HB infection, including hepatitis, liver cirrhosis and liver cancers.

Description

technical field [0001] The invention relates to the field of polypeptide medicine, in particular to a polypeptide against hepatitis B virus X protein, a polynucleotide encoding the polypeptide, and their application. Background technique [0002] Liver cancer is one of the malignant tumors that lead to the death of patients. Its malignancy is high. In my country, the death rate of liver cancer is second only to gastric cancer, ranking second in the death rate of malignant tumors in my country. According to statistics, there are 300,000 new cases of liver cancer each year in my country, and 110,000 deaths from liver cancer each year. Hepatitis B virus (HBV) infection can lead to hepatitis, liver cirrhosis, and primary liver cancer. In my country, more than 80% of liver cancer patients are HCC after HBV infection, which can be called post-HBV HCC. [0003] HBV is a DNA virus with a length of about 3.2Kb, and its open reading frame expresses hepatitis B virus surface antigen (...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): C07K7/08C12N15/11C12N15/63C12N5/10C12N1/15C12N1/19C12N1/21A61K38/10A61K48/00A61P1/16A61P31/20
CPCC07K14/02C07K7/08C07K14/00A61K38/00A61P1/16A61P31/20A61P35/00A61K39/292C12N15/79
Inventor 张晓东叶丽虹
Owner TIANJIN TOPTECH BIO SCI & TECH
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