Julian J. Albers, Tim Ammon, Dario Gosmann, Stefan Audem, Silvia Thoene, Christof Winter, Ramona Secci, Anja Wolf, Anja Stelzl, Katja Steiger, Jürgen Ruland, Florian Bassermann, Christian Kupatt, Martina Anton, Angela M. Krackhardt. (12-03-2019).

Department: 

Research Area C

Abstract: 

Adoptive transfer of T cell receptor (TCR) transgenic T cells holds great promise for treating various cancers. So far, mainly semi-randomly integrating vectors have been used to genetically modify T cells. These carry the risk of insertional mutagenesis and the sole addition of an exogenous TCR potentially results in the mispairing of TCR chains with endogenous ones. Existing approaches using non-viral vectors, such as transposons, already reduce the risk of insertional mutagenesis, but have not accomplished site-specific integration. Here, we used CRISPR-Cas9 ribonucleoproteins and Adeno-associated virus 6 for gene-targeting to deliver an engineered TCR gene specifically to the T cell receptor alpha constant (TRAC) locus, thus placing it under endogenous transcriptional control. Our data demonstrate that this approach replaces the endogenous TCR, functionally redirects the edited T cells’ specificity in vitro, and facilitates potent tumor rejection in an in vivo xenograft model.