A simple strategy for retargeting lentiviral vectors to desired cell types via a disulfide-bond-forming protein-peptide pair

Despite recent improvements in the engineering of viral envelope proteins, it remains a significant challenge to create lentiviral vectors that allow targeted transduction to specific cell populations of interest. In this study, we developed a simple ‘plug and play’ strategy to retarget lentiviral vectors to any desired cell types through in vitro covalent modification of the virions with specific cell-targeting proteins (CTPs). This strategy exploits a disulfide bond-forming protein-peptide pair PDZ1 and its pentapeptide ligand (ThrGluPheCysAla, TEFCA). PDZ1 was incorporated into an engineered Sindbis virus envelope protein (Sind-PDZ1) and displayed on lentiviral particles while the TEFCA pentapeptide ligand was genetically linked to the CTP. Her2/neu-binding designed ankyrin repeat proteins (DARPin) were used as our model CTPs. DARPin-functionalized unconcentrated lentiviral vectors harboring Sind-PDZ1 envelope protein (Sind-PDZ1-pp) exhibited >800-fold higher infectious titer in HER2 + cells than the unfunctionalized virions (8.5 × 10 6 vs. <10 4 IU/mL). Moreover, by virtue of the covalent disulfide bond interaction between PDZ1 and TEFCA, the association of the CTP with the virions is nonreversible under non-reducing conditions (e.g. serum), making these functionalized virions potentially stable in an in vivo setting.