Efficient in vivo generation of CAR T cells using a retargeted fourth-generation lentiviral vector

Chimeric antigen receptor (CAR) T cell therapy has proved remarkably successful for the treatment of hematological malignancies. However, the bespoke manufacturing of autologous CAR T cells is complex and expensive. The development of methods for in vivo engineering of T cells will enable generation of CAR T cells directly within the patient, bypassing the need for ex vivo manufacturing and thereby enabling greater access for patients. Here, we describe development of an improved retargeted Nipah envelope system paired with a fourth-generation lentiviral vector capable of specifically targeting T cells with increased efficiency, which generates high levels of functional CAR T cells in vivo. The retargeted vectors exhibited greater specificity to T cells compared to the VSV-G pseudotyped vector. Vectors targeted to either CD3 or CD8 similarly generated high levels of CAR T cells, which rapidly eradicated B cells, suggesting that T cell receptor (TCR) engagement is not required for lentiviral vectors to efficiently transduce T cells in vivo. Furthermore, the fourth-generation lentiviral vector platform (referred to as the TetraVecta system) employs the TRiP system to prevent incorporation of CAR protein into the vector particles, minimizing the risk of inadvertent transduction of tumor cells.