Bispecific VSV-G Engagers: A Platform for Cell-Type-Specific Delivery of Viral Vectors

VSV-G-pseudotyped lentiviral vectors (LVVs) enable highly efficient ex vivo gene delivery through the broadly expressed LDL receptor (LDLR) but lack the cell-type specificity required for in vivo applications. To provide cell-type specificity, we developed reversible engagers consisting of a cell-specific ligand or antibody fused to the ~40-amino-acid VSV-G-binding domains of LDLR. Following endocytosis, the engager dissociates, releasing unmodified VSV-G for efficient delivery of the vector cargo. An engager comprising the CEACAM8 N-terminal domain enables selective transduction of CEACAM6-positive epithelial cells. A trimeric CEACAM8-based engager matching the VSV-G homotrimeric spike blocked non-specific VSV infection through LDLR with an IC50 of 16.7 pM, while enabling effective VSV-mediated lysis of CEACAM6-positive tumor cells. An anti-CD3-based trimeric engager selectively transduced non-preactivated T cells in human PBMCs with high transduction efficiency. Engager-mediated in vivo CAR T cell generation resulted in sustained control of human OPM2 multiple myeloma tumors in NSG mice. This platform may further advance in vivo gene therapy, including in vivo CAR T-cell therapy and tumor-directed oncolytic virotherapy.