In-Cell Penetration Selection-Mass Spectrometry Produces Noncanonical Peptides for Antisense Delivery.
ACS Chem Biol
; 18(3): 615-628, 2023 03 17.
Article
em En
| MEDLINE
| ID: mdl-36857503
ABSTRACT
Peptide-mediated delivery of macromolecules in cells has significant potential therapeutic benefits, but no therapy employing cell-penetrating peptides (CPPs) has reached the market after 30 years of investigation due to challenges in the discovery of new, more efficient sequences. Here, we demonstrate a method for in-cell penetration selection-mass spectrometry (in-cell PS-MS) to discover peptides from a synthetic library capable of delivering macromolecule cargo to the cytosol. This method was inspired by recent in vivo selection approaches for cell-surface screening, with an added spatial dimension resulting from subcellular fractionation. A representative peptide discovered in the cytosolic extract, Cyto1a, is nearly 100-fold more active toward antisense phosphorodiamidate morpholino oligomer (PMO) delivery compared to a sequence identified from a whole cell extract, which includes endosomes. Cyto1a is composed of d-residues and two non-α-amino acids, is more stable than its all-l isoform, and is less toxic than known CPPs with comparable activity. Pulse-chase and microscopy experiments revealed that while the PMO-Cyto1a conjugate is likely taken up by endosomes, it can escape to localize to the nucleus without nonspecifically releasing other endosomal components. In-cell PS-MS introduces a means to empirically discover unnatural synthetic peptides for subcellular delivery of therapeutically relevant cargo.
Texto completo:
1
Coleções:
01-internacional
Base de dados:
MEDLINE
Assunto principal:
Peptídeos Penetradores de Células
Aspecto:
Implementation_research
Idioma:
En
Revista:
ACS Chem Biol
Ano de publicação:
2023
Tipo de documento:
Article
País de afiliação:
Estados Unidos