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J Am Chem Soc DOI:10.1021/jacs.5b05694

Discovery and characterization of a peptide that enhances endosomal escape of delivered proteins in vitro and in vivo.

Publication TypeJournal Article
Year of Publication2015
AuthorsLi, M, Tao, Y, Shu, Y, LaRochelle, JR, Steinauer, A, Thompson, D, Schepartz, A, Chen, Z-Y, Liu, DR
JournalJ Am Chem Soc
Volume137
Issue44
Pages14084-93
Date Published2015 Nov 11
ISSN1520-5126
KeywordsAnimals, Antimicrobial Cationic Peptides, Cells, Cultured, Cytoplasm, Drug Delivery Systems, Ear, Inner, Endosomes, Green Fluorescent Proteins, Integrases, Mice
Abstract

The inefficient delivery of proteins into mammalian cells remains a major barrier to realizing the therapeutic potential of many proteins. We and others have previously shown that superpositively charged proteins are efficiently endocytosed and can bring associated proteins and nucleic acids into cells. The vast majority of cargo delivered in this manner, however, remains in endosomes and does not reach the cytosol. In this study we designed and implemented a screen to discover peptides that enhance the endosomal escape of proteins fused to superpositively charged GFP (+36 GFP). From a screen of peptides previously reported to disrupt microbial membranes without known mammalian cell toxicity, we discovered a 13-residue peptide, aurein 1.2, that substantially increases cytosolic protein delivery by up to ∼5-fold in a cytosolic fractionation assay in cultured cells. Four additional independent assays for nonendosomal protein delivery collectively suggest that aurein 1.2 enhances endosomal escape of associated endocytosed protein cargo. Structure-function studies clarified peptide sequence and protein conjugation requirements for endosomal escape activity. When applied to the in vivo delivery of +36 GFP-Cre recombinase fusions into the inner ear of live mice, fusion with aurein 1.2 dramatically increased nonendosomal Cre recombinase delivery potency, resulting in up to 100% recombined inner hair cells and 96% recombined outer hair cells, compared to 0-4% recombined hair cells from +36-GFP-Cre without aurein 1.2. Collectively, these findings describe a genetically encodable, endosome escape-enhancing peptide that can substantially increase the cytoplasmic delivery of cationic proteins in vitro and in vivo.

DOI10.1021/jacs.5b05694
Pubmed

http://www.ncbi.nlm.nih.gov/pubmed/26465072?dopt=Abstract

Alternate JournalJ. Am. Chem. Soc.
PubMed ID26465072
Grant ListR01 DC006908 / DC / NIDCD NIH HHS / United States
R01 GM095501 / GM / NIGMS NIH HHS / United States
/ / Howard Hughes Medical Institute / United States