Targeting DNA-LNPs to Endothelial Cells Improves Expression Magnitude, Duration, and Specificity

Area of Science:

• Biotechnology
• Nanomedicine
• Gene Therapy

Background:

• Unmodified DNA-lipid nanoparticles (DNA-LNPs) primarily accumulate in the liver after in vivo administration.
• Endothelial cells are crucial in various diseases, including pulmonary hypertension and stroke, making them a key therapeutic target.

Purpose of the Study:

• To develop extrahepatic DNA-LNP targeting strategies for endothelial cells.
• To investigate the organ- and cell-type specificity of targeted DNA-LNPs.

Main Methods:

• Conjugating DNA-LNPs with antibodies against PECAM-1 or VCAM-1 to target lung or brain/spleen endothelium, respectively.
• Comparing transgene expression levels and duration between targeted and untargeted LNPs.
• Evaluating the impact of antibody fragments (Fab) versus full-length antibodies on organ specificity.
• Utilizing single-cell expression analysis to elucidate the mechanism of improved targeting.

Main Results:

• Targeted DNA-LNPs demonstrated robust transgene expression in target organs (lungs, brain/spleen) with greater magnitude and duration than untargeted LNPs.
• PECAM-1 targeted LNPs showed increased organ specificity over two weeks, with declining off-target liver expression.
• Replacing full-length antibodies with Fab fragments further enhanced lung-to-liver expression ratio.
• Single-cell analysis revealed that long-lived endothelial cells in target organs dominated expression, while non-endothelial cells with shorter half-lives contributed to off-target liver expression.

Conclusions:

• Targeted DNA-LNPs achieve high levels of organ- and cell-type-specific transgene expression.
• This technology offers a promising therapeutic platform for numerous endothelial-centric diseases.
• Antibody-mediated targeting of endothelial cell surface proteins effectively redirects DNA-LNP delivery and expression.