Emergence of Early Region 1-Containing Replication-Competent Adenovirus in Stocks of Replication-Defective Adenovirus Recombinants (ΔE1 + ΔE3) During Multiple Passages in 293 Cells

Abstract

Early region 1 (E1)-deleted human adenovirus (AV) recombinants have been shown to be powerful tools of gene transfer in vivo and in vitro and are considered for application in human gene therapy. We could detect increasing titers of E1-containing adenovirus in two independent E1+E3-deleted recombinant AV stocks during multiple passages in 293 cells, most likely due to a recombination event with the host cell genome. We show the deleterious effects of this E1-containing, mostly replication-competent AV subpopulation in vivo and compare different screening methods of AV stocks for its detection. These considerations are important for the safety of human gene therapy trials. The results presented in this paper show the emergence of a replication-competent AV subpopulation in different E1+E3-deleted recombinant AV stocks during multiple passages in 293 cells. This finding is most likely due to a recombination event between homologous sequences of the AV recombinants and 293 cells that include the E1 region. Different screening methods of recombinant AV stocks for the emergence of wild-type-like virus are compared, which are necessary for the safety in human gene therapy application.