Modified HIV-1 based lentiviral vectors have an effect on viral transduction efficiency and gene expression in vitro and in vivo

Gene transfer using lentiviral vectors has been recently shown to be enhanced with cis-acting elements in a cell-type-dependent manner in vivo. For this reason, the study reported here was designed to modify lentiviral vectors that express lacZ, human factor IX (FIX), or human alpha1-antitrypsin (AAT) to study the effect of different cis DNA elements on transduction efficiencies. We found that incorporation of the central polypurine tract sequence (cppt) increased transduction efficiency in vitro while increasing the transduction of non-cell-cycling hepatocytes in vivo. C57Bl/6 scid mice that were administered lentiviral vectors devoid of the cppt (2 X 10(8) transducing units (T.U.)/mouse) had 81% of their lacZ-transduced hepatocytes colabeled with the cell cycle marker 5'-bromo-2'-deoxyuridine (BrdU). In contrast, inclusion of the cppt reduced the colabeling in mouse hepatocytes by 50%. Further modifications in the lentiviral vectors were performed to enhance viral titer and gene expression. We found that the inclusion of a matrix attachment region (MAR) from immunoglobulin-kappa (Ig kappa) significantly increased the transduction efficiency, as measured by transgene protein expression and proviral DNA copy number, compared with vectors without Ig kappa MAR. In vitro studies using human hepatoma cells demonstrated a significant increase (two- to fourfold) in human AAT and human FIX production when the Ig kappa MAR was incorporated. In vivo transduction of partially hepatectomized C57Bl/6 mice given an optimized lentiviral vector containing the cppt and Ig kappa MAR (2 X 108 T.U./mouse) resulted in sustained therapeutic levels of serum FIX (similar to 65 ng/ml). Our study demonstrates the importance of cis-acting elements to enhancing the transduction ability of lentiviral vectors and the expression of vector transgenes.