High-titer human packaging cells producing γ-retroviral vectors pseudotyped with engineered baboon endogenous...

Lentiviral, α-retroviral, and γ-retroviral vector particles pseudotyped with envelope (Env) proteins of baboon endogenous retrovirus (BaEV) were shown to mediate highly efficient gene transfer into a range of primary human hematopoietic cell types in gene therapeutic strategies. However, the fusogenicity of utilized Env proteins mediated syncytia formation, hindering the establishment of stable viral packaging cell lines (VPCLs). Consequently, we used another BaEV Env variant, termed BaEV-TR-previously reported not to induce cell-to-cell fusion-and assessed its utility in stable producer cell line development. The absence of syncytia formation was reconfirmed in human 293T and HT1080 cell lines. Using HT1080 host cells, the stable VPCL HT/BaEV-TR was established. Conducting a Western blot analysis, BaEV-TR protein incorporation into vector particles was readily detected. Murine leukemia virus (MLV) pseudotype vectors efficiently transduced a variety of human target cell lines, reaching high vector titers of more than 3.0 × 10⁶ transducing units per mL (TU/mL) in 293T target cells. In summary, this proof-of-concept study underlines the utility of the Env variant BaEV-TR for the establishment of stable pseudotype VPCLs and the utilization of HT1080 cells as an alternative to frequently used 293T host cells.