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Use of VEGF-C or VEGF-D gene or protein to prevent resitenosis

A VEGF-C, VEGF-D technology, applied in the field of cardiovascular medicine, can solve problems such as arterial intimal hyperplasia and deterioration

Inactive Publication Date: 2002-03-06
LUDWIG INST FOR CANCER RES +2
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

[0014] De Young and Dichek, Circulation Research, 82:306-313 (1998) pointed out that VEGF gene delivery is not currently aimed at the application in human coronary restenosis, and two independent studies have shown that VEGF delivery can actually make intraarterial Exacerbation of membrane hyperplasia

Method used

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  • Use of VEGF-C or VEGF-D gene or protein to prevent resitenosis
  • Use of VEGF-C or VEGF-D gene or protein to prevent resitenosis
  • Use of VEGF-C or VEGF-D gene or protein to prevent resitenosis

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0086] Application of adenovirus-mediated VEGF-C gene transfer to prevent restenosis

[0087] The following experiments were carried out in vivo in a rabbit model of restenosis to demonstrate the efficacy of adenovirus-mediated intravascular VEGF-C gene transfer in preventing restenosis after angioplasty.

[0088] AMaterials and methods

[0089] 1. Adenovirus Construction

[0090] An adenoviral plasmid comprising a cDNA encoding the entire human pre-VEGF-C open reading frame operably linked to the cytomegalovirus (CMV) promoter and human growth hormone polyadenylation signal sequence was constructed as follows. A DNA fragment containing the CMV promoter sequence was prepared by cutting the pcDNA3.1+ vector (Invitrogen) with SalI and filling in the 5' overhang with Klenow enzyme. The CMV promoter (5431-911 nucleotides) was digested and isolated from the vector with HindIII. With HindIII and XhoI, the full-length human VEGF-C cDNA (and less than 50 bases of noncoding sequence a...

Embodiment 2

[0104] Comparative examples confirm that the anti-restenotic effect of VEGF-C is stronger than that of VEGF

[0105] The experiments described below demonstrate the preventive effect of adenovirus-mediated intravascular VEGF and VEGF-C gene transfer on restenosis after angioplasty, and demonstrate that VEGF-C exhibits stronger therapeutic efficacy compared to VEGF.

[0106] AMaterials and methods

[0107] 1. Adenovirus Construction

[0108] Apply the same promoter as VEGF-C, and use the method similar to Example 1 to construct VEGF adenovirus (mouse VEGF-A 164 , SEQ IN NO: 18). Production of VEGF-A in 293T cells 164 Constructs, concentrated essentially as described in Example 1, were determined to be free of helper virus, lipopolysaccharide, and bacterial contaminants.

[0109] 2. Animal models

[0110] Sixty-three New Zealand white rabbits were divided into two groups, the first group was fed with 0.25% cholesterol diet for 2 weeks, and underwent aortic dissection before...

Embodiment 3

[0122] Expression of transfected VEGFs on blood vessel wall

[0123] Utilizing the aortic fragments of the same experimental animal as in Example 2, lacZ, VEGF-C and VEGF (mouse VEGF-A) in the aortic tissue after gene transfer were analyzed. 164 ) mRNA expression. Total RNA was extracted from transfected aortic fragments using Trizol reagent (Gibco-BRL), and 2 μg was used for cDNA synthesis. To distinguish endogenous genes from transduced genes, primers for lacZ, VEGF-C and VEGF were designed by selecting 5' primers from the CMV promoter and 3' primers from the coding region.

[0124] In the amplification of lacZ, the primers are: 5' primer 5'-TTGGAGGCCTAGGCTTTT GC-3' (SEQ ID NO: 5), 3' primer 5'-ATACTGTCGTCGTCCCCTCA-3' (SEQ ID NO: 6). The first PCR cycle was an initial incubation at 96°C for 4 minutes, followed by 3 minutes at 80°C, during which time DNA polymerase was added. This was followed by 30 cycles, each cycle consisting of 94°C for 45 seconds, 58°C for 45 seconds,...

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Abstract

The present invention provides materials and methods for preventing stenosis or restenosis of a blood vessel using Vascular Endothelial Growth Factor C (VEGF-C) and / or Vascular Endothelial Growth Factor D (VEGF-D) genes or proteins.

Description

field of invention [0001] The present invention provides materials and methods for preventing vascular stenosis and restenosis, and mainly relates to the field of cardiovascular medicine. Background of the invention [0002] Coronary artery disease remains a leading cause of morbidity and mortality worldwide, especially in the United States and Europe. For these diseases, percutaneous luminal coronary angioplasty (eg, balloon angioplasty with or without intracoronary stents) is currently a common and successful treatment for these diseases, accounting for Do it hundreds of times. However, restenosis occurs in one-third to one-half of these revascularizations, usually within 6 months after angioplasty. The estimated economic cost of restenosis is $2 billion per year in the United States alone. [Feldman, et al., Cardiovascular Research, 32: 194-207 (1996), which is hereby incorporated by reference], autopsy and atherectomy studies have identified intimal hyperplasia as the ...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): C12N15/09A61F2/82A61K31/711A61K38/19A61K38/22A61K48/00A61L31/16A61L33/00A61M25/00A61P7/02A61P41/00A61P43/00C07K14/475
CPCA61F2/82A61K38/1866A61K48/00A61P41/00A61P43/00A61P7/02A61P9/10
Inventor 塞波·拉赫图亚拉卡里·阿利塔洛米科·O·希尔图南马库·M·杰尔奇马克·G·阿彻
Owner LUDWIG INST FOR CANCER RES
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