Peptide-encoding mRNA barcodes for the high-throughput in vivo screening of libraries of lipid nanoparticles for mRNA delivery

Developing safe and effective nanoparticles for the delivery of messenger RNA (mRNA) is slow and expensive, partly due to the lack of predictive power of in vitro screening methods and the low-throughput nature of in vivo screening. While DNA barcoding and batch analysis present methods for increasing in vivo screening throughput, they can also result in incomplete or misleading measures of efficacy. Here, we describe a high-throughput and accurate method for the screening of pooled nanoparticle formulations within the same animal. The method uses liquid chromatography with tandem mass spectrometry to detect peptide barcodes translated from mRNAs in nanoparticle-transfected cells. We show the method’s applicability by evaluating a library of over 400 nanoparticle formulations with 384 unique ionizable lipids using only nine mice to optimize the formulation of a biodegradable lipid nanoparticle for mRNA delivery to the liver. Barcoding lipid nanoparticles with peptide-encoding mRNAs may facilitate the rapid development of nanoparticles for mRNA delivery to specific cells and tissues.