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Adenovirus vector for idiopathy liver genetherapy and using method

A technology of adenovirus and recombinant adenovirus, applied in gene therapy, methods based on microorganisms, medical raw materials derived from viruses/phages, etc., can solve problems such as unclear mechanism and cell apoptosis, and achieve strong tumor killing specificity, highly effective effect

Active Publication Date: 2008-09-03
SHANGHAI SUNWAY BIOTECH
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

2. Inhibit tumors through a p53-independent mechanism, which may be related to the activation or inactivation of some genes in cells, thereby promoting cell apoptosis. The mechanism is still unclear (Ricardo SP et al., 1996)

Method used

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  • Adenovirus vector for idiopathy liver genetherapy and using method
  • Adenovirus vector for idiopathy liver genetherapy and using method
  • Adenovirus vector for idiopathy liver genetherapy and using method

Examples

Experimental program
Comparison scheme
Effect test

Embodiment I

[0095] The construction of embodiment 1 recombinant adenovirus AdHS4.AFP.E1a / TRAIL

[0096] The recombinant adenovirus AdHS4.AFP.E1a / TRAIL described below is a type 5 adenovirus vector with an alpha-fetoprotein promoter, and has an HS4 insulator gene upstream of the promoter, and an exogenous insertion gene downstream of the promoter Because the adenovirus early E1a gene related to tumor suppression and the tumor necrosis factor-related apoptosis-inducing ligand TRAIL gene related to apoptosis activation, and the two are connected through the internal ribosome entry site IRES, they share the same transcription unit, express separately.

[0097] Unless otherwise stated, the techniques used in the present invention are conventional techniques in the art, such as molecular cloning techniques, microbiology techniques, and cell biology techniques, and these techniques are fully explained in the literature. Such as Sambrook et al. "Molecular Cloning Laboratory Manual" Second Editio...

Embodiment II

[0103] Example II Study on the specific effect of recombinant adenovirus AdHS4.AFP.E1a / TRAIL in liver cancer cells

[0104] MTS method to detect the effect of recombinant adenovirus AdHS4.AFp.E1a / TRAIL on different tumor cells:

[0105] In order to detect the tumor-specific cytotoxicity of the recombinant adenovirus AdHS4.AFP.E1a / TRAIL, by using the MTS (Promega) kit to detect the cells of the virus in AFP negative / positive cells in live cells when the infection coefficient MOI is 100 (TCID50) Toxic specificity. HepG2, Hep3B, SMMC-7721, and BEL-7404 are liver cancer cell lines, L-02 is normal human liver cells, LS174T is human colon cancer cells, and Hek293 is human embryonic kidney cells. The above cells were inoculated in a 96-well plate to 104 cells per well. After 24 hours, they were infected with AdHS4.AFP.E1a / TRAIL, dl1520, or the culture medium was used as a control. MTS reagent was added at different time points and incubated at 37°C for one hour. After that, the 490...

Embodiment III

[0110] Example III Study on the antitumor effect of recombinant adenovirus AdHS4.AFP.E1a / TRAIL in nude mice bearing liver cancer xenografts

[0111] Anti-tumor effect of recombinant adenovirus AdHS4.AFP.E1a / TRAIL in nude mice bearing human hepatoma BEL-7404 tumors in vivo: 6-week-old BALB / C nude mice were inoculated with 1×107 BEL-7404 cell suspension 100μl. After tumor formation, when the transplanted tumor grew to 80-200mm3, they were randomly divided into groups, with 7-12 nude mice in each group, PBS, AdHS4.AFP.E1a / TRAIL (1.0×107 TCID50 / mm3), dl1520 (1.0×107 TCID50 / mm3), or DDP was directly injected into the transplanted tumor with an injection volume of 50 μl for 5 consecutive days and DDP for 7 consecutive days. The tumor volume was calculated by detecting the tumor diameter and measured twice a week on the 31st day. All tumor-bearing nude mice were sacrificed, and the tumor volume on the day of injection was set as 100%. Compared with the negative control group, the oth...

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Abstract

The invention of adenovirus carrier for primary liver cancer gene therapy and its use method provides a method based on oncolytic virus therapy and a composite for treating primary liver cancer. The invention constructs a gene recombinant adenovirus containing alpha-fetoprotein promotor gene and therapy gene. The research shows that the recombinant virus can specifically kill the primary liver cancer cell for expressing the alpha-fetoprotein.

Description

technical field [0001] The present invention relates to the field of gene therapy and virus therapy for tumors, and specifically relates to the technique of regulating adenoviral vectors to perform virus replication and exogenously inserted gene expression under specific conditions through tumor-specific promoters, that is, the use of alpha-fetoprotein promoters to regulate Adenoviral vector, which specifically carries out virus replication and foreign insertion gene expression in primary liver cancer cells expressing alpha-fetoprotein, so that in liver cancer cells expressing alpha-fetoprotein, it is specifically controlled by the alpha-fetoprotein promoter The technology of virus replication and exogenous insertion gene expression to kill liver cancer cells. Background technique [0002] Malignant tumors seriously threaten human health and are the second "killer" of humans after cardiovascular diseases. Traditional tumor treatment methods have considerable limitations. Th...

Claims

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Application Information

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IPC IPC(8): A61K48/00C12N15/861C12R1/92C12N7/01C12N15/11C12N15/12A61K35/76A61P35/00A61K35/761
Inventor 梁旻孟夏任晓维胡放
Owner SHANGHAI SUNWAY BIOTECH
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