Self-assembling protein nanoparticles for cytosolic delivery of nucleic acids and proteins

Abstract

Intracellular delivery of biomacromolecules is hampered by low efficiency and cytotoxicity. Here we report the development of elastin-based nanoparticles for therapeutic delivery (ENTER), a recombinant elastin-like polypeptide (ELP)-based delivery system for effective cytosolic delivery of biomacromolecules in vitro and in vivo. Through iterative design, we developed fourth-generation ELPs fused to cationic endosomal escape peptides (EEPs) that self-assemble into pH-responsive micellar nanoparticles and enable cytosolic entry of cargo following endocytic uptake. In silico screening of α-helical peptide libraries led to the discovery of an EEP (EEP13) with 48% improved protein delivery efficiency versus a benchmark peptide. Our lead ELP–EEP13 showed similar or superior performance compared to lipid-based transfection reagents in the delivery of mRNA-encoded, DNA-encoded and protein-form Cre recombinase and CRISPR gene editors as well as short interfering RNAs to multiple cell lines and primary cell types. Intranasal administration of ELP–EEP13 combined with Cre protein achieved efficient editing of lung epithelial cells in reporter mice.