title: Use of quantitative TaqMan real-time PCR to track the time-dependent distribution of gene transfer vectors in vivo.
abstract: To assess the biodistribution and pharmacokinetics of gene transfer vectors, real-time PCR with fluorescent TaqMan chemistry was used to quantify tissue levels of adenovirus gene transfer vectors (Ad) following myocardial administration. After optimizing the detection of the genome of Ad vectors expressing human vascular endothelial growth factor (Ad(GV)VEGF121.10) and Escherichia coli cytosine deaminase (Ad(GV)CD.10), a comparison was made of intramyocardial injection versus intracoronary delivery to the left ventricle of the pig. One hour post-intramyocardial administration, the left ventricular Ad genome level was 6.2 copies per cellular genome, 26-fold higher than the level of 0.24 copies per cellular genome following intracoronary administration. Relative to the vector levels after 1 h, the amount dropped 14- and 5.5-fold by 24 h following intramyocardial and intracoronary administration, respectively. Interestingly, the vector that escaped the left ventricle after intracoronary or intramyocardial administration to pigs was found primarily within the lung, an observation in marked variance to the biodistribution of Ad vector in rodents. In this regard, after intravenous injection to the pig, 90% of the recovered vector was found in the lung, and even after intrahepatic portal vein injection, 55% of the recovered vector was in the lung. These data have important implications regarding the use of experimental animals for safety studies on administration of Ad to humans.
authors: Belfer Gene Therapy Core Facility, Weill Medical College of Cornell University, New York, New York, 10021, USA. geneticmedicine@mail.med.cornell.edu
authors: N R
authors: NR
authors: Hackett
authors: T
authors: T
authors: El Sawy
authors: L Y
authors: LY
authors: Lee
authors: I
authors: I
authors: Silva
authors: J
authors: J
authors: O'Leary
authors: T K
authors: TK
authors: Rosengart
authors: R G
authors: RG
authors: Crystal
mesh_terms: D000256:Adenoviridae / Q000235:genetics; D000818:Animals; D001483:Base Sequence; D043525:Cytosine Deaminase; D017931:DNA Primers; D016228:Endothelial Growth Factors / Q000235:genetics; D004926:Escherichia coli / Q000201:enzymology; D018014:Gene Transfer Techniques; D005822:Genetic Vectors; D006801:Humans; D008222:Lymphokines / Q000235:genetics; D008297:Male; D009700:Nucleoside Deaminases / Q000235:genetics; D016133:Polymerase Chain Reaction / Q000379:methods*; D013552:Swine; D042461:Vascular Endothelial Growth Factor A; D042442:Vascular Endothelial Growth Factors
chemical_list: D017931:DNA Primers; D016228:Endothelial Growth Factors; D008222:Lymphokines; C467484:VEGFA protein, human; D042461:Vascular Endothelial Growth Factor A; D042442:Vascular Endothelial Growth Factors; D009700:Nucleoside Deaminases; D043525:Cytosine Deaminase