Linked Life Data

19364285

Source:http://linkedlifedata.com/resource/pubmed/id/19364285

Statements in which the resource exists as a subject.
PredicateObject
rdf:type
lifeskim:mentions
pubmed:issue
8
pubmed:dateCreated
2009-8-11
pubmed:abstractText
Liver-based gene therapy approaches demonstrated that high-capacity adenoviral vectors (HC-AdVs) can persist life-long in mice and for 2 years or longer in rats, dogs, and nonhuman primates. However, the molecular status of episomal HC-AdV DNA molecules and the mechanism of vector genome maintenance have not been analyzed. HC-AdV lacks all viral coding sequences including early gene region 4 (E4), which prevents concatemerization in wild-type adenovirus. Therefore, we addressed whether concatemerization or circularization of HC-AdV DNA occurs in transduced cells. We employed pulsed-field gel electrophoresis and a sensitive concatemer/circle-specific polymerase chain reaction (PCR). To test for replication as a potential mechanism for maintenance, we developed a methylase/restriction endonuclease-based system using methylation-marked HC-AdV. We found that unlike DeltaE4 mutant virus, only monomers of HC-AdV genomes were observable in vitro. Using our methylase/restriction endonuclease-based system, no replication of HC-AdV was sensed in various cell lines. However, concatemer formation of HC-AdV could be induced after coinfection with an E4-deleted helper virus, indicating that linkage of genomes may be supported by replication. To examine HC-AdV DNA molecules in vivo, C57BL/6 mice were injected and vector DNA in liver was analyzed. In concordance with our in vitro results, exclusively linear monomers were detected. To sense the replication status of HC-AdV genomes, we established a sensitive real-time PCR. Our results indicated that the input transduced DNA genomes were the persistent molecules in murine liver. In summary, we demonstrated that HC-AdV genomes persist predominantly as replication-defective monomeric genomes.
pubmed:language
eng
pubmed:journal
pubmed:citationSubset
IM
pubmed:chemical
pubmed:status
MEDLINE
pubmed:month
Aug
pubmed:issn
1557-7422
pubmed:author
pubmed:issnType
Electronic
pubmed:volume
20
pubmed:owner
NLM
pubmed:authorsComplete
Y
pubmed:pagination
883-96
pubmed:meshHeading
pubmed:year
2009
pubmed:articleTitle
Persistence of high-capacity adenoviral vectors as replication-defective monomeric genomes in vitro and in murine liver.
pubmed:affiliation
Department of Virology, Max von Pettenkofer-Institute, Ludwig-Maximilian-University of Munich, Munich 80336, Germany.
pubmed:publicationType
Journal Article, Research Support, Non-U.S. Gov't