Rapid receptor internalization potentiates CD7-targeted lipid nanoparticles for efficient mRNA delivery to T cells and in vivo CAR T-cell engineering

Abstract

Targeted lipid nanoparticles (tLNPs) enable efficient mRNA delivery to T cells, allowing for in situ generation of chimeric antigen receptor (CAR) T cells without ex vivo manipulation. This strategy has shown promising therapeutical efficacy in preclinical studies of cardiac fibrosis, cancer, and autoimmune diseases. While multiple T-cell surface receptors have been targeted across studies for tLNP-mediated in vivo CAR T-cell generation and exhibit diverse efficiencies, their comparative performance and the mechanisms underlying these differences remain unclear. Here, we systematically compared tLNPs with antibody-based moieties targeting T-cell receptors including CD2, CD4, CD5, CD7, CD8, or a CD4/8 dual-targeting combination under identical conditions, assessing their mRNA delivery efficiency in human T cells and PBMCs in vitro, and subsequently validating the best performer in vivo in humanized mice. Among all moieties tested, CD7-targeting tLNPs achieved the highest mRNA delivery to T cells and efficiently generated functional CAR T cells in vivo. Mechanistic analysis revealed that receptor internalization, rather than the receptor abundance, is the primary determinant of delivery efficiency, a property intrinsic to each receptor and largely independent of antibody clone. These findings provide a rational framework for selecting optimal targeting moiety to enable highly efficient in vivo CAR T-cell engineering. HighlightsO_LITargeting CD7 outperforms other receptors for tLNP-mRNA delivery to T cells C_LIO_LIReceptor abundance does not predict tLNP-mRNA delivery efficiency C_LIO_LIReceptor internalization kinetics governs tLNP-mRNA delivery efficiency C_LIO_LICD7-targeting LNP-mRNA enables efficient in vivo CAR T-cell engineering C_LI Graphical Abstract O_FIG O_LINKSMALLFIG WIDTH=200 HEIGHT=90 SRC="FIGDIR/small/701374v1_ufig1.gif" ALT="Figure 1"> View larger version (35K): org.highwire.dtl.DTLVardef@9c2c4eorg.highwire.dtl.DTLVardef@120ced7org.highwire.dtl.DTLVardef@eb9b5forg.highwire.dtl.DTLVardef@25b6f7_HPS_FORMAT_FIGEXP M_FIG C_FIG