PMID- 9048194
OWN - NLM
STAT- MEDLINE
DCOM- 19970821
LR  - 20071114
IS  - 1043-0342 (Print)
IS  - 1043-0342 (Linking)
VI  - 8
IP  - 3
DP  - 1997 Feb 10
TI  - Lack of site-specific integration of the recombinant adeno-associated virus 2 
      genomes in human cells.
PG  - 275-84
AB  - The adeno-associated virus 2 (AAV)-based vector system has been suggested for its 
      potential use in human gene therapy because the wild-type (wt) AAV genome appears 
      to integrate into the human chromosomal DNA in a site-specific manner. We 
      systematically investigated the integration patterns of the recombinant AAV 
      genomes lacking one or both the viral coding sequences. Four recombinant AAV 
      genomes were constructed containing the genes for resistance to tetracycline 
      (TcR) and the herpesvirus thymidine kinase (TK) promoter-driven gene for 
      resistance to neomycin (neoR; vTc.Neo), the genes for resistance to ampicillin 
      (ApR) and TK-neoR (vAp.Neo), the genes for AAV replication (rep) genes and 
      TK-neoR (vRep.Neo), and the AAV capsid (cap) genes and TK-neoR (vCap.Neo). The 
      integration pattern of each of the recombinant AAV genomes in individual clonal 
      isolates of the human nasopharyngeal carcinoma cell line (KB) analyzed on 
      Southern blots using a neo-specific DNA probe was distinctly different. In 
      addition, in none of the clones examined was the proviral genome covalently 
      linked to the previously described AAV right-junction (Rt.Jn.) human chromosomal 
      DNA fragment, the putative specific-site of integration for the wt AAV genome. 
      Furthermore, whereas a 276-bp DNA fragment could be readily amplified from each 
      of these clones, using a neo-specific primer-pair by polymerase chain reaction 
      (PCR), no amplified DNA product was obtained using the neo- and the Rt.Jn. 
      primer-pair under identical conditions. Fluorescence in situ hybridization (FISH) 
      analyses further revealed the lack of integration of the recombinant AAV into 
      human chromosome 19, even in the presence of a functional rep gene as determined 
      by rescue of the recombinant AAV genome in the presence of adenovirus. These data 
      suggest that the recombinant AAV genomes integrate at sites that are different 
      from that characterized for the wt AAV genome. These studies may have 
      implications in the development of the AAV-based vector system for its potential 
      use in human gene therapy.
FAU - Ponnazhagan, S
AU  - Ponnazhagan S
AD  - Department of Medicine, Indiana University School of Medicine, Indianapolis 
      46202-5120, USA.
FAU - Erikson, D
AU  - Erikson D
FAU - Kearns, W G
AU  - Kearns WG
FAU - Zhou, S Z
AU  - Zhou SZ
FAU - Nahreini, P
AU  - Nahreini P
FAU - Wang, X S
AU  - Wang XS
FAU - Srivastava, A
AU  - Srivastava A
LA  - eng
GR  - AI-26323/AI/NIAID NIH HHS/United States
GR  - HL-48342/HL/NHLBI NIH HHS/United States
GR  - HL-53586/HL/NHLBI NIH HHS/United States
GR  - etc.
PT  - Journal Article
PT  - Research Support, Non-U.S. Gov't
PT  - Research Support, U.S. Gov't, P.H.S.
PL  - United States
TA  - Hum Gene Ther
JT  - Human gene therapy
JID - 9008950
RN  - 0 (DNA Transposable Elements)
RN  - 0 (DNA, Recombinant)
SB  - IM
MH  - DNA Transposable Elements/*genetics
MH  - DNA, Recombinant/isolation & purification
MH  - Dependovirus/*genetics/isolation & purification
MH  - Genetic Vectors/genetics
MH  - Genome, Viral
MH  - Humans
MH  - Recombination, Genetic/*genetics
MH  - Tumor Cells, Cultured
MH  - *Virus Integration
EDAT- 1997/02/10 00:00
MHDA- 1997/02/10 00:01
CRDT- 1997/02/10 00:00
PHST- 1997/02/10 00:00 [pubmed]
PHST- 1997/02/10 00:01 [medline]
PHST- 1997/02/10 00:00 [entrez]
AID - 10.1089/hum.1997.8.3-275 [doi]
PST - ppublish
SO  - Hum Gene Ther. 1997 Feb 10;8(3):275-84. doi: 10.1089/hum.1997.8.3-275.