Targeted Transduction of CD34 + Cells by Transdominant Negative Rev-Expressing Retrovirus Yields Partial Anti-HIV Protection of Progeny Macrophages

Abstract

Congenitally acquired HIV infection may be uniquely suited to treatment via genetic engineering of CD34⁺ hematopoietic stem/progenitor cells. However, current technologies yield only a small percentage of mature cells that carry the inserted genes, and expression is frequently suppressed. Since clinical trials employing these methodologies have been proposed for anti-HIV gene therapy of HIV-infected children, we wished to assess, by in vitro modeling, the expected limits of transduction efficiency, expression, and antiviral activity using currently available methods. We measured retrovirus-mediated transduction in cord blood progenitors and their in vitro-derived progeny macrophages by Mo-MuLV vectors expressing a transdominant negative Rev (Rev^TD). CFU-GM transduction efficiency ranged from 7 to 85%, with an average of 28%. Semiquantitative DNA PCR demonstrated ≤100 vector sequence copies per 1000 cells in monocyte/macrophage cultures, which were grown without selection to better model in vivo conditions. When challenged with the macrophage-tropic HIV-1_BaL isolate, cultured macrophages from mock-transduced CFU-GM colonies supported infection in eight of eight experimental cultures, control LXSN-transduced progenitors supported infection in six of eight cultures, while macrophages derived from Rev^TD-transduced CFU-GM colonies supported infection in four of eight cultures. Although these results support the ability of neo ^r retroviral vectors containing Rev^TD to inhibit HIV replication, they indicate that further optimization of transduction efficiency and sustained expression will be required for effective anti-HIV protection in vivo.

Overview summary

A major goal in gene therapy for AIDS is to modify hematopoietic stem/progenitor cells with anti-HIV constructs—to provide for antiviral protection in progeny macrophages and T lymphocytes. The transdominant negative Rev protein (Rev^TD) has exhibited potent anti-HIV activity in primary and transformed T lymphocytes. In this study, we demonstrated that transduction of primary CD34⁺ cells with a retroviral vector expressing Rev^TD confers anti-HIV activity to progeny macrophages. This study also indicates that further improvements in transduction efficiency and Rev^TD expression will be required for effective anti-HIV protection in vivo.