Search

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. [ABSTRACT FROM AUTHOR]

A strong immune response against transgenic cells is one important limitation for long-term expression after adenoviral gene transfer in mammals. Continuous pharmacological immunosuppression has been shown to ameliorate immune reactions and to prolong reporter gene expression. In this study, we explored the effect of short-term immunosuppression for long-term gene expression and its impact on antibody formation. Immunosuppression with FK 506 (1 mg/kg/day), cyclosporin A (20 mg/kg/day) and 15-deoxyspergualin (10 mg/kg/day) was performed in NMRI mice. Expression of the reporter gene human alpha-1-antitrypsin (hAAT) and antibody formation was monitored for 7 months. A 5-day course of 15-deoxyspergualin (15-DSG) markedly slowed the decline of reporter gene expression and a positive effect was still detectable 200 days after gene transfer. At the same time, antibody production was reduced by 50–60%. Continuous treatment with 15-DSG (10 mg/kg twice weekly) led to a further small increase of gene expression but reduced antibody formation by 80–90%. A short course of FK 506 and cyclosporin A (CsA), had conferred a negative effect on gene expression. Both groups showed an even faster reduction in gene expression compared with the control group. The results of this investigation suggest that 15-DSG could serve as an effective supplement for viral gene therapy protocols. [ABSTRACT FROM AUTHOR]