Extended transgene expression from a nonintegrating adenoviral vector containing retroviral elements

Abstract

We studied the effects of specific retroviral elements in a first-generation serotype 5 adenoviral (Ad5) vector, AdLTR2 EF1α-hEPO. This vector contains 858 base pair (bp) [251-bp envelope sequence plus 607-bp long-terminal repeat (LTR)] from Moloney murine leukemia virus (MoMLV), upstream of the human elongation factor-1α (EF1α) promoter and human erythropoietin (hEPO) cDNA, with the LTR sequence downstream of the polyadenylation signal. We compared expression of AdLTR2EF1α-hEPO with corresponding expressions of two conventional Ad5 vectors, AdEF1α-hEPO and AdCMV-hEPO, in vivo in submandibular glands in rats. Both the conventional vectors yielded low serum hEPO levels by day 7, and little change in hematocrits. In contrast, after receiving AdLTR2EF1α-hEPO, the rats showed elevated hEPO levels and hematocrits for 1-3 months. In vitro studies showed that the integration efficiencies of all the vectors were similar (∼10-3). Approximately 0.1% of the vector genomes were present 1 year after delivery in the case of each of the three vectors, primarily as intact linear double-strand DNA. The unique results seen with AdLTR2EF1α-hEPO are partly because of LTR enhancer activity. However, other cis-acting activity, which is not immunomodulatory but nevertheless influences promoter methylation, appears to be involved. A vector such as AdLTR2 EF1α-hEPO may prove useful in clinical applications in which extended, but not "permanent," transgene expression is desirable.