Editing T cell specificity towards leukemia by zinc finger nucleases and lentiviral gene transfer

Abstract

The transfer of high-avidity T-cell receptor (TCR) genes isolated from rare tumor-specific lymphocytes into polyclonal T cells is an attractive cancer immunotherapy strategy. However, TCR gene transfer results in competition for surface expression and inappropriate pairing between the exogenous and endogenous TCR chains, resulting in suboptimal activity and potentially harmful unpredicted specificities. We designed zinc-finger nucleases (ZFNs) promoting the disruption of endogenous TCR β and α chain genes. ZFN-treated lymphocytes lacked CD3/TCR surface expression and expanded with IL-7 and IL-15. Upon lentiviral transfer of a TCR for the WT1 tumor antigen, these TCR-edited cells expressed the new TCR at high levels, were easily expanded to near-purity, and proved superior in specific antigen recognition to matched TCR-transferred cells. In contrast to TCR-transferred cells, TCR edited lymphocytes did not mediate off-target reactivity while maintaining anti-tumor activity in vivo, thus demonstrating that complete editing of T-cell specificity generate tumor-specific lymphocytes with improved biosafety profile.

Is autoimmunity the Achilles' heel of cancer immunotherapy?

“…Genome editing to create CAR T cells or TCR transgenic T cells devoid of the endogenous TCR in order to prevent GVHD and enhance the function of the transgenic TCR has been reported 64,100,101 .…”

A Cut above the Rest: Targeted Genome Editing Technologies in Human Pluripotent Stem Cells

“…Other than elucidating gene function and modeling human diseases, genome editing technologies can facilitate a variety of novel studies, such as improving directed differentiation of hiPSCs by generating lineage-specific reporter lines, engineering dendritic cell-directed cancer vaccines and T cell immunotherapies (58,59), and generating human cell lines with enhanced production of biomolecules for biotechnology and pharmaceutical industries.…”

Emerging applications of gene edited T cells for the treatment of leukemia

“…The ability to use banked CAR T-cells, generated from non-HLA matched donors in an 'off-the-shelf' manner has been an attractive prospect that has driven attempts to overcome HLA-barriers, both in terms of allo-reactivity from infused cells and host-mediated rejection [6][7][8]. T-cells encode highly specific heterodimeric αβ T-cell receptors, and these are the key mediators of major histocompatibility complex (MHC) recognition leading to GVHD.…”

“…T-cells encode highly specific heterodimeric αβ T-cell receptors, and these are the key mediators of major histocompatibility complex (MHC) recognition leading to GVHD. Strategies to disrupt T-cell receptor (TCR) expression have used a variety of reagents including RNA interference (RNAi) [9] and targeted gene disruption using directed DNA nucleases such as Zinc Finger Nucleases [6,7], Meganucleases [10], MegaTALs [11,12], and Transcription activator like effector nucleases (TALENs) [8,13,14]. The latter operate at the genomic level, directing engineered endonucleases to highly specific loci to create double stranded DNA cleavage.…”