The Overlooked Stereoisomers of the Ionizable Lipid ALC315

Lipid nanoparticles (LNPs) are a powerful delivery platform for nucleic acid therapeutics such as mRNA vaccines and gene therapies. Central to their success are ionizable lipids, which facilitate the cellular uptake and endosomal escape of nucleic acids. However, achieving a high delivery efficiency often comes with the drawback of increased cytotoxicity. Here, we report a chemical, biological, and toxicological investigation into the three stereoisomers of ALC315, a mixture of which constitutes one of the most successful marketed ionizable lipids for LNPs. We demonstrate that the individual stereoisomers of ALC315 can be accessed by either the asymmetric chemical synthesis or chromatography of an intermediate. An LNP formulation based on a single stereoisomer of ALC315 enhances mRNA transfer efficiency while reducing the associated cytotoxicity in human cell lines. Our results underscore the potential of stereochemically pure ionizable lipids as key components in the development of next-generation nucleic acid therapies, offering an enhanced delivery performance and better safety profiles.