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Life sciences · Journal article
Angewandte Chemie International Edition · October 27, 2026
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ABSTRACT Efficient, biocompatible, and flexible application of CRISPR‐Cas9 remains a major challenge in gene therapy. This study addresses a novel formulation method, focusing on organic solvent‐free lipid nanoparticles for the efficient delivery of CRISPR‐Cas9 components. Therefore, novel stealth lipids comprising the lipid anchor cholesterol hemisuccinate conjugated to poly(2‐ethyl‐2‐oxazoline) (PEtOx) were synthesized and evaluated as a promising in vitro alternative to commercially available poly(ethylene glycol) (PEG)‐based lipids. Our study identified PEtOx‐lipids as markedly more biocompatible compared to PEG‐based analogues (12‐fold reduction in cytotoxicity; 99% reduced hemolytic effect) with high efficiency for mRNA gene delivery. We demonstrate successful CRISPR‐Cas9 activity ex vivo and efficient uptake of PEtOx nanoparticles by macrophages in vivo (66.5% pos. F4/80 + macrophages). Our data demonstrate that PEtOx‐based stealth lipids are a promising alternative to PEG. The resulting LNPs, optimized for fully aqueous manufacturing, address key limitations of current LNP systems, including immunogenicity and the reliance on ethanol‐based production processes.