Looking for breakthrough ideas for innovation challenges? Try Patsnap Eureka!

Recombination adenovirus for curing overexpression proto-oncogene neu/erbB2 malignancy

A technology of recombinant adenovirus and proto-oncogene, applied in the field of recombinant adenovirus construction co-expressing human monoclonal antibody and Mda-7

Inactive Publication Date: 2007-10-17
王尚武
View PDF0 Cites 2 Cited by
  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

However, although the humanized monoclonal antibody of the antigenic oncogene neu / erbB2 has been successfully used in the treatment of malignant breast cancer overexpressing the proto-oncogene neu / erbB2, clinical studies have shown that most metastatic breast cancers that are effectively treated with this antibody in the early stage Patients will become resistant to this antibody treatment within a year, and 15% of patients will still relapse

Method used

the structure of the environmentally friendly knitted fabric provided by the present invention; figure 2 Flow chart of the yarn wrapping machine for environmentally friendly knitted fabrics and storage devices; image 3 Is the parameter map of the yarn covering machine
View more

Image

Smart Image Click on the blue labels to locate them in the text.
Viewing Examples
Smart Image
  • Recombination adenovirus for curing overexpression proto-oncogene neu/erbB2 malignancy
  • Recombination adenovirus for curing overexpression proto-oncogene neu/erbB2 malignancy
  • Recombination adenovirus for curing overexpression proto-oncogene neu/erbB2 malignancy

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0015] The cloning process of the proto-oncogene neu / erbB2 humanized monoclonal antibody scFv (the same below) gene is as follows (including the connection between the light chain variable region and the heavy chain variable region of the human monoclonal antibody; cloning of the novel scFv gene expression plasmid ):

[0016] 1). The connection between the variable region of the light chain of the humanized monoclonal antibody and the variable region of the heavy chain:

[0017] a. Using the humanized monoclonal antibody cDNA as a template, carry out PCR reaction with artificially synthesized primers. First, amplify the light chain variable region of the antibody, add a Hind111 and Nhe1 endonuclease cut site to the 5'-N end of the light chain variable region, and splice the signal consisting of 20 amino acids of human interleukin-2 Peptide with a Kpn1 endonuclease cleavage site at its 3'-C terminus. This process is completed through the second PCR reaction, that is, the firs...

Embodiment 2

[0035] Example 2. Artificial synthesis and cloning of Mda-7 DNA fragment: According to the known gene sequence, the coding DNA fragment of Mda-7 was artificially synthesized according to conventional techniques, Smal1 was added at its 5'-end, and Smal1 was added at the 3'-not Xba1 restriction site was added at the end, and PCR amplification was carried out with the following primers.

[0036] Primer 1:

[0037] 5'-agcgggccctatgaattttcaacagaggctgcaaagcctgtgg-3'

[0038] Primer 2:

[0039] 5'-cgacagatctatcagagcttgtagaatttctgcatcc-3'

[0040] After tailing and adding adenine, the PCR amplification product was connected to the T-easy vector, transformed into bacteria, amplified, extracted, purified, and confirmed by DNA sequencing.

Embodiment 3

[0041] Example 3. Construction of an expression cassette composed of CMV promoter, scFv, internal ribosome binding site (IRES) and Mda-7. The Stratagene product used in this example is pShuttle-IRES, and the expression cassette was constructed using this vector as the backbone. The construction of this expression cassette was confirmed by multiple digestions with different endonucleases, ligation reactions, transformation of competent bacteria and sequencing. That is, the scFv fragment with the 5'-end as Nhe1 and the 3'-end as the Not1 endonuclease site is spliced ​​upstream of the IRES; the 5'-end as the Sma1 endonuclease site and the 3'-end as the Xba1 endonuclease site The Mda-7 gene fragment at the Dicer site is spliced ​​upstream of SV40 polyadenine and downstream of IRES; so far, the construction of the expression cassette has been completed.

the structure of the environmentally friendly knitted fabric provided by the present invention; figure 2 Flow chart of the yarn wrapping machine for environmentally friendly knitted fabrics and storage devices; image 3 Is the parameter map of the yarn covering machine
Login to View More

PUM

No PUM Login to View More

Abstract

The present invention relates to a method for constructing a recombinant adenovirus for expressing neu / erbB2 proto-oncogene human-source antibody variable region and Mda-7 / IL-24 expression box structure, and its uses. The expression box is characterized as 1) neu / erbB2 proto-oncogene human-source antibody variable region, 2) Mda-7 / IL-24 being 206 amino acid gene fragment, the 72 position amino acid converting from Ala to Val, 3) connecting neu / erbB2 proto-oncogene human-source antibody variable region and Mda-7 gene through inner ribosome binding site(IRES), 4) the adenovirus being duplication-defective serum A5 type adenovirus. The novel adenovirus expresses two kinds of anti-cancer gene, that is neu / erbB2 proto-oncogene human-source antibody variable region and Mda-7 gene, whose product has strong cell apotosis function and does no harm to normal cell so it has significance in gene therapy for malignant tumour of over expression for neu / erbB2 proto-oncogene.

Description

technical field [0001] The invention relates to the construction of a recombinant adenovirus co-expressing human monoclonal antibody and Mda-7 for treating tumors overexpressing proto-oncogene neu / erbB2, and its technical field is related to life science and biomedicine technology. Background technique [0002] ·Studies have shown that the proto-oncogene neu / erbB2 is an important marker gene of various malignant tumors, and its product is also called malignant protein. About 30% of breast cancer patients and some patients with other tumors, such as ovarian cancer, lung cancer and gastric cancer, The gene neu / erbB2 is highly expressed, which is one of the important reasons for the deterioration and distant metastasis of these tumors. Foreign countries have successfully developed a humanized monoclonal antibody against the proto-oncogene neu / erbB2. Approved by the US FDA for marketing and clinical application for many years, it has been shown that this therapeutic humanized m...

Claims

the structure of the environmentally friendly knitted fabric provided by the present invention; figure 2 Flow chart of the yarn wrapping machine for environmentally friendly knitted fabrics and storage devices; image 3 Is the parameter map of the yarn covering machine
Login to View More

Application Information

Patent Timeline
no application Login to View More
IPC IPC(8): C12N15/861C12N15/13C12N15/28C07K14/525C07K16/18A61K48/00A61P35/00
CPCC12N2799/04A61K35/13C12N15/62C07K2317/565C07K16/00C07K16/32C12N2840/203C12N2799/022A61K48/00C12N2830/36A61K2039/505C07K16/005C12N15/861C12N15/86C12N2710/10343A61P35/00
Inventor 王尚武
Owner 王尚武
Who we serve
  • R&D Engineer
  • R&D Manager
  • IP Professional
Why Patsnap Eureka
  • Industry Leading Data Capabilities
  • Powerful AI technology
  • Patent DNA Extraction
Social media
Patsnap Eureka Blog
Learn More
PatSnap group products

Browse by: Latest US Patents, China's latest patents, Technical Efficacy Thesaurus, Application Domain, Technology Topic, Popular Technical Reports.

© 2024 PatSnap. All rights reserved.Legal|Privacy policy|Modern Slavery Act Transparency Statement|Sitemap|About US| Contact US: help@patsnap.com