Superior In Vivo Efficacy for T Cell Engineering via Citronellol‐Tailored mRNA‐tLNPs

ABSTRACT Efficient in vivo generation of CAR T cells via mRNA‐LNP delivery necessitates high‐efficiency T cell engineering, yet current systems remain suboptimal. While lymphoid tropism is essential, it is demonstrated that high tropism alone is insufficient for effective in vivo engineering—some highly tropic LNPs frequently exhibit suboptimal T cell transfection. Crucially, intrinsic T cell transfection potency emerges as an orthogonal and equally critical determinant, consistent with the need for both T cell specificity and transfection efficiency. Through screening a tail‐varied ionizable lipid library, H 3 T 4 is identified, a citronellol‐tailed LNP with enhanced lymphoid tropism and superior intrinsic potency. The CD3‐targeted H 3 T 4 LNP (αH 3 T 4 ) achieves ~85% in vivo efficacy in T cell engineering, outperforming benchmark LNPs by ∼50% (40% vs. 85%). This proposes a new optimization strategy: effective engineering requires balancing potency and tropism, not maximizing tropism alone. In murine CD19 solid tumor models, αH 3 T 4 ‐generated CAR T cells achieved 83% tumor clearance, demonstrating potency‐driven efficacy. This work establishes intrinsic transfection potency as a primary tunable parameter for next‐generation CAR T platforms.