687. Development of a Safe and Effective Combinatorial Foamy Virus Vector for HIV Gene Therapy

The Berlin patient and successes in retroviral hematopoietic stem cell (HSC) gene therapy suggest that gene therapy may provide a functional cure for HIV. However, retroviral-HSC gene therapy has resulted in serious adverse side effects due to vector mediated genotoxicity, including leukemia. Foamy virus (FV) vectors have a promising integration profile and may be safer than retroviral vectors. FV vectors also can efficiently deliver anti-HIV transgenes that can reduce the titer of HIV-1 based lentiviral vectors. We report a novel combinatorial anti-HIV FV vector that uses a housekeeping elongation factor 1 alpha (EF1α) promoter but still potently blocks HIV infection. We first evaluated the relative potency of various previously described anti-HIV transgenes, including the C46 fusion inhibitor, the F12-Vif derivative Chim3, lens epithelium derived growth factor-integrase binding domain (LIBD) and TRIM5α-CyclophilinA fusion (TCypA) in a FV vector background. We found that C46 was the most potent anti-HIV transgene, followed by TCypA and LEDGF-IBD. Next, we hypothesized that using a less-genotoxic internal promoter to drive the transgenes would reduce vector-mediated genotoxicity. Therefore, a direct comparison was made between the efficacy of housekeeping gene promoters (EF1α and Ubiquitin C; UbC) and a highly genotoxic SFFV promoter in expressing the C46 transgene and subsequently blocking HIV replication. We observed that C46 EF1α had ~2 to 4 fold higher anti-HIV effect than C46 driven by either SFFV or UbC promoters, respectively. Based on these results, we designed a novel combinatorial FV vector expressing three anti-HIV transgenes: C46, TcypA and LIBD. The gene cassette was driven by an EF1α promoter and also has mCherry as a reporter gene (FV-E C46TLmC-W). This vector can be produced at high titer, 1.4 × 107 transducing units/ml which is critical for clinical translation. This novel combinatorial anti-HIV FV vector showed a higher potency in blocking HIV replication than C46 alone at a late time point, 21 days post HIV infection (Figure 1Figure 1). An in vitro competitive survival advantage assay indicated that cells transduced with our novel combinatorial anti-HIV FV vectors are highly resistant to HIV infection compared to cells transduced with FV expressing a control EGFP reporter gene alone (FV-EG-W) (Figure 2Figure 2). Further studies will be focused on the efficacy of our novel combinatorial anti-HIV FV vector in human CD34+ cells transplanted in a mouse xenotransplant in vivo model. Our goal is to develop a safe and potent combinatorial FV vector for clinical studies.View Large Image | Download PowerPoint SlideView Large Image | Download PowerPoint Slide