Combinatorial Gene Therapy to Inhibit HIV: Improving Therapeutic Efficacy by Targeting Multiple Stages of the HIV-1 Replication Cycle Simultaneously

In these studies, we tested the effectiveness of combinatorial gene delivery to CCR5-expressing cell lines and primary cells to enhance resistance to HIV-1 infection. The transgenes used were chosen both to decrease membrane CCR5 and to inhibit HIV-1 replication, and were delivered using Tag-deleted SV40-derived vectors. rSV40s are very effective in transducing primary T cells and T cell lines. SV(RNAiR5), SV(RevM10.AU1) and SV(RNAiR5/RevM10.AU1) respectively deliver a small interfering RNA (siRNA) against CCR5, RevM10, to which a C-terminal AU1 epitope was added and which inhibits HIV-1 Rev, or both transgenes together. We used human cell lines (SupT1 and SupT1/CCR5) and primary human monocyte-derived macrophages (MDMs). We assessed RevM10 expression by flow cytometry (FACS). After transduction, we confirmed expression of each transgene by flow cytometry (FACS). Simultaneous delivery of both transgenes in a bifunctional vector protected both SupT1/CCR5 cells and MDMs from R5 tropic HIV-1Ba-L, better than either the monofunctional vectors, SV(RNAiR5) and SV(RevM10.AU1) individually. The bifunctional vector also protected from X4-tropic HIV-1, comparably to SV(RevM10.AU1). SV(RNAiR5) did not protect from X4-tropic HIV-1. Thus, combining these two transgenes in one vector protected from X4-tropic HIV-1 and provided enhanced protection for CCR5-bearing cells from R5-tropic strains of HIV-1. Combinatorial genetic therapy, by targeting of more then one HIV-1 function, may provide effective inhibition of HIV-1 replication.