A Novel Non-Viral Platform Combining A LEG Nuclease And mbDNA™ For High-Efficiency T-Cell Engineering

Non-viral gene editing could help make engineered T-cell therapies safer and more accessible, but efficient delivery, precise integration, and cell recovery remain persistent challenges. A platform combining a compact LEG nuclease with a cell-free mbDNA template achieved at least 80% TRAC knockout using either mRNA or ribonucleoprotein complexes. LNP delivery produced greater than 95% knockout with minimal toxicity, while mbDNA generated twice as many anti-NYESO-1 TCR T cells as the plasmid-based approach and supported the integration of larger DNA sequences. Although donor DNA caused an initial decline in viability and reduced expansion capacity, edited cells recovered to greater than 95% viability by harvest.

Explore how nuclease format, delivery method, and HDR template design can shape the efficiency of non-viral T-cell engineering.