A double-amine ionizable lipid library enables highly efficient mRNA delivery to T cells at reduced lipid mass.

Abstract

mRNA-based nucleic acid therapeutics have gained significant attention and undergone extensive clinical investigation since the FDA's approval of Spikevax and Comirnaty for the treatment of SARS-CoV-2. Lipid nanoparticles (LNPs) are currently the most advanced delivery system for delivering mRNA in vivo. However, concerns persist regarding the tolerability of their repeated use due to lipid accumulation. Over time, lipids have been optimized through the introduction of enzymatically degradable ester linkers. Yet, this modification alone does not fully address lipid clearance from the body. To mitigate these concerns, we have designed a library of novel ionizable lipids that incorporates two nitrogen atoms within the lipid backbone. We hypothesize that the LNPs with these lipids can be potentially administered at lower dosages while maintaining the same payload delivery and translation efficiency. In this study, we identified five lead LNPs that exhibited remarkable transfection efficiency without cytotoxic effects in vitro. We further optimized the LNP formulations by incorporating various cholesterol analogs to fine-tune translation performance. Upon analyzing the endocytic and intracellular pathways of the lead LNPs, we found that multiple pathways were involved in delivering the payload to the cell. This set of novel lipids holds great promise for mRNA delivery and warrants further investigation.