Abstract
Recombinant adenoviruses are presently the most efficient in vivo gene transfer system available. Targeting single organs or large tumors by adenoviral vectors requires an intravascular route of application. During the first pass of viral particles through the vascular bed of the target tissue, virus uptake is not quantitative and undefinite amounts of particles leak into circulation. To determine the amount of leaking particles and to calculate organ-specific uptake (in-/outflow ratio), it is necessary to titrate virus particles directly in blood. In preclinical and clinical trials titration is currently mostly done with blood plasma instead of full blood. However, this technique provides valid results only as long as there is no affinity between adenovirus particles and erythrocytes. In this study we demonstrate that Ad5 particles, as mostly employed for gene therapy, have a strong affinity to human erythrocytes. At 60 min after coincubation of human erythrocytes and Ad5 particles, more than 98% of the particles are attached to the surface of erythrocytes. Therefore, ignoring the amount of red cell bound particles by performing titration in plasma leads to severe miscalculation of organ-specific transfer rates or virus circulation half-life. The biological impact of an increased affinity between virus particles and erythrocytes will be discussed.
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Acknowledgements
This study was supported by the Bundesministerium fur Bildung und Forschung. We thank Kordelia Hummel and Jenny Schlesinger for their excellent technical assistance. We are grateful to the blood donors among our colleagues and Dr Schulze from the blood bank of the Helios-Klinik of the Charité in Berlin Buch for providing additional blood samples. We thank Dr Meißler for providing blood of domestic pigs. We are very grateful to Andrew Byrnes and Charles Coutelle for important suggestions.
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Cichon, G., Boeckh-Herwig, S., Kuemin, D. et al. Titer determination of Ad5 in blood: a cautionary note. Gene Ther 10, 1012–1017 (2003). https://doi.org/10.1038/sj.gt.3301961
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DOI: https://doi.org/10.1038/sj.gt.3301961
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