Polypeptide drug for resisting Hepatitis B virus X protein

A hepatitis B virus and drug technology, applied in the direction of viral peptides, antiviral agents, viruses, etc., can solve the problems of short half-life of polypeptide fragments, influence on pharmacodynamics, and difficulty in finding polypeptide drugs, and achieve obvious pharmacodynamics. effect of action

Active Publication Date: 2012-09-19
TIANJIN TOPPHARM BIO SCI & TECH CO LTD
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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 for resisting Hepatitis B virus X protein
  • Polypeptide drug for resisting Hepatitis B virus X protein
  • Polypeptide drug for resisting Hepatitis B virus X protein

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0100] Synthetic polypeptide functional fragment Anti-HBxP3#:

[0101] The present invention synthesized the amino acid sequence Met-Glu-Pro-Gly-Ala-Gly-His-Leu-Asp-Gly-His-Arg-Ala-Gly-Ser-Pro (SEQ ID NO: 1) by artificial synthesis method The polypeptide (hereinafter referred to as Anti-HBxP3#). 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 one added to the amino acid sequence Met-Glu-Pro-Gly-Ala-Gly-His-Leu-Asp-Gly-His-Arg-Ala -The amino acid monomer of Gly-Ser-Pro is Pro at the C-terminus, then Ser, then Gly, until the last Glu and Met at the N-terminus, continuously adding, reacting, synthesizing, and fin...

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

[0191] 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, in particular to a polypeptide for resisting hepatitis B virus X protein and polynucleotide coding the polypeptide, and application of the polypeptide and the polynucleotide. Specifically, the invention relates to a polypeptide with functional activity of inhibiting hepatitis B virus X protein (HBX), and the polypeptide has activity of inhibiting HBX on a molecular level, a cell level and an animal level so as to inhibit hepatitis caused by infection of hepatitis B viruses, liver cirrhosis caused by repeated attacks of hepatitis, and liver cancer caused on the basis of liver cirrhosis. The polypeptide and a peptide stimulant of the polypeptide comprise functional fragments and functional variants of the polypeptide and the peptide stimulant, and genes for coding the polypeptide, the peptide stimulant or the functional fragments and the functional variants, and are widely used for preventing and treating liver diseases that attack after infection of Hepatitis B virus, including hepatitis, liver cirrhosis and liver cancer.

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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IPC IPC(8): C07K7/08C12N15/11C12N15/63C12N5/10C12N1/15C12N1/19C12N1/21A61K38/10A61K48/00A61P1/16A61P31/20
CPCC07K7/08C07K14/02A61K38/00C07K14/00A61P1/16A61P31/20A61P35/00C07K14/005C12N2730/10122
Inventor 张晓东叶丽虹
Owner TIANJIN TOPPHARM BIO SCI & TECH CO LTD
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