Use of quantitative TaqMan real-time PCR to track the time-dependent distribution of gene transfer vectors in vivo

To assess the biodistribution and pharmacokinetics of gene transfer vectors , real-time PCR with fluorescent TaqMan chemistry was used to quantify tiss ue levels of adenovirus gene transfer vectors (Ad) following myocardial adm inistration. After optimizing the detection of the genome of Ad vectors exp ressing human vascular endothelial growth factor (Ad(GV)VEGF121.10) and Esc herichia coli cytosine deaminase (Ad(GV)CD.10), a comparison was made of in tramyocardial injection versus intracoronary delivery to the left ventricle of the pig. One hour post-intramyocardial administration, the left ventric ular Ad genome level was 6.2 copies per cellular genome, 26-fold higher tha n the level of 0.24 copies per cellular genome following intracoronary admi nistration. Relative to the vector levels after 1 h, the amount dropped 14- and 5.5-fold by 24 h following intramyocardial and intracoronary administr ation, respectively. Interestingly, the vector that escaped the left ventri cle after intracoronary or intramyocardial administration to pigs was found primarily within the lung, an observation in marked variance to the biodis tribution of Ad vector in rodents. In this regard, after intravenous inject ion 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 ex perimental animals for safety studies on administration of Ad to humans.