Towards developing HIV-2 lentivirus-based retroviral vectors for gene therapy: Dual gene expression in the context of HIV-2 LTR and Tat

Because of the distinct ability of retroviruses to integrate into the target cell genome and thus achieve long-term expression, retrovirus vectors hold great promise for stable gene transfer. Such vectors derived from human immunodeficiency retroviruses (HIVs) and other lentiviruses are envisioned to possess several advantages, especially for in vivo gene therapy of HIV infection and acquired immunodeficiency syndrome (AIDS) where targeting CD4+ T cells/macrophages and pluripotent non-dividing stem cells would be required. Among these is the ability of HIVs to transduce nondividing cells in contrast to the murine retroviruses which require target cell mitosis. The advantages of the lentivirus vectors will be further enhanced by the development of multigenic vectors carrying more than one gene in a dependent or independent transcriptional unit. Separate from the issue of transduction efficiency, information is needed about the impact of the configuration of the genes in a multigenic vector on their expression. Towards this end, we investigated the expression of genes specifically directed by the HIV-2 LTR and Tat in a prototypic minimal transfer vector. We found that the expression of a gene in a dual gene configuration depended upon its position in the transcriptional unit and that the insertion of an internal translational initiation mechanism improved the expression of the downstream gene. Apparently not sufficiently appreciated previously, these effects were promoter and cell-type dependent. Our data also suggest that the commonly used cellular or viral promoters may be orders of magnitude less effective than HIV-2 LTR in the presence of Tat, and thus may not be useful as internal promoters in the context of the HIV-2 LTR:Tat regulatory loop. (C) 1998 Wiley-Liss, Inc.dagger.