Fusion protein for treating breast cancer and application thereof

A fusion protein, breast cancer technology, applied in the fusion protein and its application field, can solve the problems that have not been completely solved, and achieve the effect of breaking through the gene inhibition effect, inhibiting growth and metastasis

Inactive Publication Date: 2009-11-25
SUN YAT SEN UNIV +2
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

However, many problems restricting siRNA t

Method used

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  • Fusion protein for treating breast cancer and application thereof
  • Fusion protein for treating breast cancer and application thereof
  • Fusion protein for treating breast cancer and application thereof

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0053] Example 1: Preparation of fusion protein of Her2 single-chain fragment antibody and protamine polypeptide

[0054] After finding the full-length sequence of the Her2 single-chain fragment antibody from the gene library, use primer software to design relevant primers.

[0055] Designed Her2-ScFv primers:

[0056] Forward primer: NcoI is an endonuclease

[0057] 5'-GGGCCATGGCCCAGGTGCAGCTGTTGCAGTCTGGGGCAGAG-3'

[0058] Anti-primer: NotI is an endonuclease

[0059] 5'-TTGCGGCCGCTCCGGAATTCACCTAGGACGGTCAGCTTGGTCCC-3'

[0060] Designed protidine peptide primers:

[0061] Positive primer: NotI is an endonuclease

[0062] 5'-CCGGAGCGGCCGCAATGGCCAGGTACAGATGCTG-3'

[0063] Anti-primer: PpuM I as endonuclease, stop codon and 6 histidines

[0064] 5'-GCCGGGTCCCAGGAAAGGATCAGATCTGCATTAATGGTGGTGGTGATGATGAGATCTGTGTCTTTCTACATCTCGGTCTG-3'

[0065] Clone relevant target fragments by PCR, Her2-ScFv PCR reaction system 1:50ul system:

[0066] wxya 2 O 37ul

[0067] dNTP 5ul

[00...

Embodiment 2

[0123] Example 2: Inhibitory effect of F5-P fusion protein targeted delivery of PLK1-siRNA therapy on orthotopic tumor growth of breast cancer

[0124] The specific steps for cell expansion, culture and inoculation of nude mice to make breast cancer tumor orthotopic and metastasis models are as follows: (BT 474-EGFP cells used in the metastasis model were transfected with EGFP expressing green fluorescence in our laboratory and screened by flow cytometry cell line)

[0125] 1 Human breast cancer cell BT 474 was cultured in DMEM / F12 medium containing 10% calf serum in an incubator with 5% CO2 at 37°C and saturated humidity, and the cells grew in a monolayer adherent manner;

[0126] 2 When the cells cover 95% of the bottom of the bottle, digest with 0.25% trypsin for 1 min. Under an inverted microscope, the cell space increases and the cytoplasm shrinks, and the digestion stops. Suck up and discard the trypsin solution;

[0127] 3. Add a small amount of serum culture medium an...

Embodiment 3

[0136] Example 3: The effect of F5-P fusion protein targeted delivery of PLK1-siRNA therapy on the expression, proliferation and apoptosis of tumor target genes in breast cancer

[0137] F5-P fusion protein targeted delivery of PLK1-siRNA to treat breast cancer tumor in situ and metastasis models, the specific steps are as follows:

[0138] 1. Treat tumor-bearing mice from the second day after injecting breast cancer cells;

[0139] 2 Randomly grouping the breast cancer model mice, with 8 nude mice in each group;

[0140] Breast cancer orthotopic model mice group 4: F5-P fusion protein plus PLK1-siRNA group

[0141] Single-use F5-P fusion protein group

[0142] Single use PLK1-siRNA group

[0143] F5-P fusion protein plus NC-siRNA group

[0144] Four groups of breast cancer metastasis model mice: F5-P fusion protein plus PLK1-siRNA group

[0145] Single-use F5-P fusion protein group

[01...

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Abstract

The present invention discloses a fusion protein for treating a breast cancer, wherein the fusion protein is Her2 single-chain fragment antibody-polypeptide, the Her2 single-chain fragment antibody is capable of targeting a Her2 receptor locating on a surface of a breast cancer cell, and the polypeptide is capable of carrying siRNA. The polypeptide includes a protamine polypeptide. The siRNA includes PLK1-siRNA, which is capable of restraining an expression of a knubble malignant phenotype gene. The fusion protein for treating the breast cancer is capable of carrying the PLK1-siRNA, and targeting and importing a Her2 positive breast cancer cell specifically, so as to bring the medicament into the cell. The treatment of the breast cancer is capable of generating a strong effect for restraining the breast cancer growth and transfer, may break a gene suppression effect of the existing antisense oligonucleotide and ribozyme, and becames a novel anti-Her2 gene medicament for treating the breast cancer.

Description

technical field [0001] The invention relates to a fusion protein and its application, in particular to a fusion protein for treating breast cancer and its application. Background technique [0002] Breast cancer is one of the most common malignant tumors in women, and its incidence rate has leapt to the first among female malignant tumors, and it is the "first killer" of women. Although many patients had their primary tumor resected and comprehensive treatment was performed after surgery, many patients eventually failed the treatment due to metastasis. Different breast cancers may have different genotypes, and these genotypes often show large differences in response to treatment. [0003] Her2 is one of the typical malignant phenotype genes of breast cancer, and its overexpression in breast cancer is positively correlated with the proliferation, metastasis and chemotherapy resistance of cancer cells. About 20-30% of breast cancer patients' cancer tissue cells express this ...

Claims

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Application Information

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IPC IPC(8): C07K19/00A61K48/00A61K47/42A61P35/00A61P15/14
Inventor 宋尔卫姚燕丹王均孙天盟张佩琢
Owner SUN YAT SEN UNIV
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