Ionizable lipids largely determine the biocompatibility of lipid nanoparticles (LNPs) and the efficacy for mRNA delivery. Rational design and combinatorial synthesis have led to the development of potent and biodegradable ionizable lipids, yet methodologies for the stepwise optimization of ionizable lipid structure are lacking. Here we show that iterative chemical derivatization and combinatorial chemistry, and in particular the amine–aldehyde–alkyne coupling reaction, can be leveraged to iteratively accelerate the structural optimization of propargylamine-based ionizable lipids (named A³-lipids) to improve their delivery activity and biodegradability. Through five cycles of such directed chemical evolution, we identified dozens of biodegradable and asymmetric A³-lipids with delivery activity comparable to or better than a benchmark ionizable lipid. We then derived structure−activity relationships for the headgroup, ester linkage and tail. Compared with standard ionizable lipids, the lead A³-lipid improved the hepatic delivery of an mRNA-based genome editor and the intramuscular delivery of an mRNA vaccine against SARS-CoV-2. Structural criteria for ionizable lipids discovered via directed chemical evolution may accelerate the development of LNPs for mRNA delivery.

可电离脂质在很大程度上决定了脂质纳米颗粒 （LNP） 的生物相容性和 mRNA 递送的功效。合理的设计和组合合成导致了有效和可生物降解的可电离脂质的开发，但缺乏逐步优化可电离脂质结构的方法。在这里，我们展示了迭代化学衍生化和组合化学，特别是胺-醛-炔烃偶联反应，可以迭代加速基于炔丙胺的可电离脂质（称为 A³-脂质）的结构优化，以提高其递送活性和生物降解性。通过这种定向化学进化的五个循环，我们鉴定了数十种可生物降解和不对称的 A³-脂质，其递送活性与基准可电离脂质相当或更好。然后，我们推导出头部基团、酯键和尾部的结构-活性关系。与标准可电离脂质相比，铅 A³-脂质改善了基于 mRNA 的基因组编辑器的肝脏递送和针对 SARS-CoV-2 的 mRNA 疫苗的肌肉内递送。通过定向化学释放发现的可电离脂质的结构标准可能会加速用于 mRNA 递送的 LNP 的开发。