In vivo site-specific engineering to reprogram T cells
Menée à l'aide de lignées cellulaires, de modèles murins de leucémie lymphoblastique aiguë, de myélome multiple ou de sarcome et menée à l'aide de cellules immunitaires d'origine humaine, cette étude met en évidence l'intérêt d'une méthode utilisant un virus adéno-associé et un vecteur EDV (enveloppe utilisant des particules virales) pour délivrer respectivement des ribonucléoprotéines CRISPR-Cas9 et un transgène dans les lymphocytes T afin de produire in vivo des lymphocytes CAR-T
Engineered T cells, reprogrammed to express chimeric antigen receptors (CAR) or T cell receptors (TCR), have transformed cancer treatment and are being explored as therapeutics for autoimmune and infectious diseases. Enhancing T cell function through genome editing, either by disrupting endogenous genes or precisely inserting DNA payloads, has shown considerable promise1. However, the ex vivo manufacturing process is lengthy and costly, limiting accessibility of these therapies. In vivo generation of CAR T cells could overcome these barriers, but current methods rely either on transient expression with limited durability, or on random integration of DNA payloads that lack specificity. Here we demonstrate that stable and cell-specific transgene expression can be achieved through in vivo site-specific integration of large DNA payloads. We developed a two-vector system to deliver CRISPR–Cas9 ribonucleoproteins and a DNA donor template, using enveloped delivery vehicles and adeno-associated viruses, respectively. We optimized both vectors for T cell-specific delivery and gene-targeting efficiency. By integrating a CAR transgene into a T cell-specific locus, we generate therapeutic levels of CAR T cells in vivo in humanized mouse models of B cell aplasia, and haematological and solid malignancies. These findings offer a pathway to more efficient, precise and widely accessible T cell therapies.
Nature , article en libre accès, 2026