Abstract
Retroviral vectors have shown their curative potential in clinical trials correcting monogenetic disorders. However, therapeutic benefits were compromised due to vector-induced dysregulation of cellular genes and leukemia development in a subset of patients. Bromodomain and extraterminal domain (BET) proteins act as cellular cofactors that tether the murine leukemia virus (MLV) pre-integration complex to host chromatin via interaction with the MLV integrase (IN) and thereby define the typical gammaretroviral integration distribution. We engineered next-generation BET-independent (Bin) MLV vectors to retarget their integration to regions where they are less likely to dysregulate nearby genes. We mutated MLV IN to uncouple BET protein interaction and fused it with chromatin-binding peptides. The addition of the CBX1 chromodomain to MLV INW390A efficiently targeted integration away from gene regulatory elements. The retargeted vector produced at high titers and efficiently transduced CD34+ hematopoietic stem cells, while fewer colonies were detected in a serial colony-forming assay, a surrogate test for genotoxicity. Our findings underscore the potential of the engineered vectors to reduce the risk of insertional mutagenesis without compromising transduction efficiency. Ultimately, combined with other safety features in vector design, next-generation BinMLV vectors can improve the safety of gammaretroviral vectors for gene therapy.