CRISPR Cell Therapy for Immuno Oncology:
Editing in Immune Cells

CRISPR RNP gene knock-in techniques are currently being used to construct transgenic chimeric antigen receptor T (CAR-T) cells and T cell receptor T (TCR-T) cells for immunotherapy. Yet challenges remain to improve editing efficiency while reducing cytotoxicity.

Recently, significant efficiency improvements have been achieved using non-toxic GenExact single-stranded DNA (ssDNA) templates designed with a Cas9 Targeting Sequence (CTS) to bind the RNP for co-delivery via electroporation¹. This method achieved >90% knock-in efficiency with minimal cytotoxicity in multiple primary immune cell types.

NK cells are another attractive target for immune cell therapy. However, their highly sensitive and resistance to exogenous DNA limits the effectiveness of viral and plasmid-based approaches.

Non-viral CRISPR RNP systems are currently being optimized to engineer chimeric antigen receptor NK (CAR-NK) cells to improve cancer-killing activity and reduce tumor evasion^2-6. Non-viral approaches using Cas9 mRNA have also been applied to NK cell engineering to achieve >98% transfection efficiency with >90% cell viability⁷.

Antibody-producing B cells are another exciting target for immune cell therapy. While initial research focused on AAV-delivered antibody-expressing cassettes, B cells often produced insufficient antibody expression levels or duration⁸.

Therefore, CRISPR RNP-based workflows are currently being optimized for applications in B cell therapy^9,10. Specifically, targeted DNA insertion at the B cell receptor loci allows researchers to reprogram antibody specificity¹¹.