A Cell Line for High-Yield Production of E1-Deleted Adenovirus Vectors without the Emergence of Replication-Competent Virus

Gao, Guangping; Engdahl, Ryan; Wilson, James M.

doi:10.1089/10430340050016283

Cited by 34 publications

(17 citation statements)

References 16 publications

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“…However, a disadvantage of the 293 cell line is that it allows the emergence of replicationcompetent adenovirus (RCA) as a result of homologous recombination between the host cell genome and the vector [21]. This has led to strategies to avoid RCA by creating rationally designed E1-complementing helper cell lines with minimal or no homologous sequences between the transfected E1 DNA and E1-deleted vector [22,23].…”

Section: Production and Purification Of Adenoviral Vectorsmentioning

confidence: 99%

Adenoviral vectors for gene therapy

Douglas

2007

Mol Biotechnol

148

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Vectors based on human adenovirus serotypes 2 (Ad2) and 5 (Ad5) of species C possess a number of features that have favored their widespread employment for gene delivery both in vitro and in vivo. However, the use of recombinant Ad2- and Ad5-based vectors for gene therapy also suffers from a number of disadvantages. These vectors possess the tropism of the parental viruses, which infect all cells that possess the appropriate surface receptors, precluding the targeting of specific cell types. Conversely, some cell types that represent important targets for gene transfer express only low levels of the cellular receptors, which lead to inefficient infection. Another major disadvantage of Ad2- and Ad5-based vectors in vivo is the elicitation of both an innate and an acquired immune response. Considerable attention has therefore been focused on strategies to overcome these limitations, thereby permitting the full potential of adenoviral vectors to be realized.

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Section: Production and Purification Of Adenoviral Vectorsmentioning

confidence: 99%

Adenoviral vectors for gene therapy

Douglas

2007

Mol Biotechnol

148

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“…Contamination with RCA is unacceptable for clinical applications and must be avoided during production steps. In order to minimize the RCA occurrence, E1 complementing cell lines of the next generation, such as PER.C6 human embryonic retinoblasts, 23 N52.E6 human amniocytes 24 or HeLa-derived GH329 cells, 25 were established. These cells contain a minimal E1 gene fragment with no sequence overlap with the rAd genome.…”

Section: Adenovirus and Adeno-associated Virus Packaging Systemsmentioning

confidence: 99%

Chromatographic purification of recombinant adenoviral and adeno-associated viral vectors: methods and implications

Burova

Ioffe

2005

Gene Ther

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“…8 GH354 cells (courtesy of Dr J. M. Wilson, University of Pennsylvania, Philadelphia) were grown as described. 19 …”

Section: Cell Culturesmentioning

confidence: 99%

Calreticulin Destabilizes Glucose Transporter-1 mRNA in Vascular Endothelial and Smooth Muscle Cells Under High-Glucose Conditions

Totary-Jain

Naveh-Many

Riahi

et al. 2005

Circulation Research

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Abstract-Substrate autoregulation of glucose transporter-1 (GLUT-1) mRNA and protein expression provides vascular endothelial and smooth muscle cells a sensitive mechanism to adapt their rate of glucose transport in response to changing glycemic conditions. Hyperglycemia-induced downregulation of glucose transport is particularly important in protecting these cells against an excessive influx of glucose and consequently increased intracellular protein glycation and generation of free radicals; both are detrimental in the development of vascular disease in diabetes. We aimed to investigate the molecular mechanism of high glucose-induced downregulation of GLUT-1 mRNA expression in primary bovine aortic vascular endothelial (VEC) and smooth muscle (VSMC) cell cultures. Using RNA mobility shift, UV cross-linking, and in vitro degradation assays, followed by mass-spectrometric analysis, we identified calreticulin as a specific destabilizing trans-acting factor that binds to a 10-nucleotide cis-acting element (CAE 2181(CAE -2190 ) in the 3Ј-untranslated region of GLUT-1 mRNA. Pure calreticulin accelerated the rate of GLUT-1 mRNA-probe degradation in vitro, whereas overexpression of calreticulin in vascular cells decreased significantly the total cell content of GLUT-1 mRNA and protein. The expression of calreticulin was augmented in vascular cells exposed to high glucose in comparison with low-glucose conditions. Similarly, increased expression of calreticulin was observed in aortae of diabetic Psammomys obesus in comparison with normoglycemic controls. These data suggest that CAE 2181-2190 -calreticulin complex, which is formed in VSMC and VEC exposed to hyperglycemic conditions, renders GLUT-1 mRNA susceptible to degradation. This interaction underlies the process of downregulation of glucose transport in vascular cells under high-glucose conditions. (Circ Res. 2005;97:1001-1008.)Key Words: calreticulin Ⅲ glucose transporter-1 Ⅲ hyperglycemia Ⅲ mRNA turnover Ⅲ vascular smooth muscle cells Ⅲ vascular endothelial cells H yperglycemia is a major risk factor in the development of cardiovascular complications associated with the metabolic syndrome and diabetes. 1,2 Chronic hyperglycemia alters the normal function of endothelial and smooth muscle cells in blood vessels, which undergo morphological and functional modifications, because of an excessive extra-and intracellular protein glycation and an uncontrolled production of free radicals. Collectively, these processes contribute to basement-membrane thickening, vascular occlusion, increased permeability, and initiation and progression of vascular disease and atherosclerosis. [3][4][5][6] The transport of glucose across the plasma membrane of cells is the rate-limiting step for subsequent glucose metabolism. A family of glucose transporters (GLUTs) mediates the entry of glucose into cells by facilitated diffusion. 7 Vascular endothelial (VEC) and vascular smooth muscle (VSMC) cells express predominantly the ubiquitous GLUT-1 and to a very small extent the insulin-sensi...

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A Cell Line for High-Yield Production of E1-Deleted Adenovirus Vectors without the Emergence of Replication-Competent Virus

Cited by 34 publications

References 16 publications

Adenoviral vectors for gene therapy

Adenoviral vectors for gene therapy

Chromatographic purification of recombinant adenoviral and adeno-associated viral vectors: methods and implications

Calreticulin Destabilizes Glucose Transporter-1 mRNA in Vascular Endothelial and Smooth Muscle Cells Under High-Glucose Conditions

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