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Recombinant chimeric eucaryotic expression vector Cb1-Grb7(SH2)-RING and use of anti-tumour gene drug

A technology of chimeric molecules and expression plasmids, which can be used in anti-tumor drugs, recombinant DNA technology, gene therapy, etc., and can solve the problem of not using recombinant Cbl ubiquitin ligase.

Inactive Publication Date: 2007-05-09
FOURTH MILITARY MEDICAL UNIVERSITY
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

However, there is no report on the application of recombinant Cbl ubiquitin ligase for gene therapy research.

Method used

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  • Recombinant chimeric eucaryotic expression vector Cb1-Grb7(SH2)-RING and use of anti-tumour gene drug
  • Recombinant chimeric eucaryotic expression vector Cb1-Grb7(SH2)-RING and use of anti-tumour gene drug
  • Recombinant chimeric eucaryotic expression vector Cb1-Grb7(SH2)-RING and use of anti-tumour gene drug

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Experimental program
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Effect test

Embodiment Construction

[0033] The following is the construction and identification of the recombinant chimeric Cbl ubiquitin ligase Cbl-Grb7(SH2)-RING eukaryotic expression plasmid (pcDNA3.1(+)-Cbl-Grb7(SH2)-RING) given by the inventor and its identification Inhibitory effect of plasmids on the growth of EGFR and HER2 positive tumor cells.

[0034] 1. Construction of recombinant chimeric Cbl ubiquitin ligase Cbl-Grb7(SH2)-RING eukaryotic expression plasmid

[0035] 1.1 Experimental materials

[0036] Both restriction enzymes and T4 ligase were purchased from Dalian Baosheng Biological Co., Ltd., and the eukaryotic cell expression plasmid pcDNA3.1(+) was purchased from Invitrogen Company. The plasmid map of pcDNA3.1(+) is shown in Figure 1. In the figure, PCMV: CMV promoter, base 232-819, T7: T7 promoter / primer site, base 863-882, polyclonal restriction site: base 895-1010, BGHpA: bovine Growth hormone polyadenylation sequence, 1028-1252 bases, f1 ori: f1 start site, 1296-1726 bases, SV40 ori: SV40...

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Abstract

This invention discloses a structure method for the Cb1-Grb7 (SH2)-RING eukaryote expression carrier of the recomposed inserting Cb1 ligase, wherein the composed expression plasmid pcDNA3.1 (+)-Cb1-Grb7 (SH2)-RING can be used as the gene medicament of the anti-EGFR or anti-HER2 masculine tumour. It is proved by the experiment that after the Cb1-Grb7 (SH2)-RING catching with the HER2 masculine clung carcinomaan cell A549 or the HER2 masculine galactophore carcinomaan cell SK-BR-3, it restrains from the cell proliferation coming from excessive activating with the EGFR or the HER2 by accelerating them to move down, so it has the anti-EGFR or anti-HER2 masculine tumour use.

Description

technical field [0001] The invention belongs to the field of tumor gene therapy, and relates to the construction of a recombinant chimeric Cbl ubiquitin ligase Cbl-Grb7(SH2)-RING eukaryotic expression vector and its use in gene medicine against EGFR-positive or HER2-positive tumors. Background technique [0002] Cbl (Casitas B-lineage lymphoma) is a kind of evolutionarily conserved and widely distributed cytoplasmic protein, which consists of 906 amino acids and contains multiple different structural domains, which can mediate the combination with different intracellular molecules, so In the process of cell activation, it can be used as an adapter molecule or a scaffold molecule to participate in signal transduction; at the same time, the RING domain in the molecule endows Cbl with the function of ubiquitin ligase, which promotes the ubiquitination and downregulation of its bound target molecules, thereby participating in the intracellular Mechanisms...

Claims

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Application Information

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Patent Type & Authority Patents(China)
IPC IPC(8): C12N15/79C12N15/66A61K48/00A61P35/00
Inventor 李霞张璟刘新平药立波
Owner FOURTH MILITARY MEDICAL UNIVERSITY