Phase-separating peptides for direct cytosolic delivery and redox-activated release of macromolecular therapeutics

Abstract Biomacromolecules are highly promising therapeutic modalities to treat various diseases. However, they suffer from poor cellular membrane permeability, limiting their access to intracellular targets. Strategies to overcome this challenge employ nanoscale carriers that often get trapped in endosome compartments. Here, we report on conjugated peptides forming pH- and redox-responsive coacervate microdroplets by liquid-liquid phase separation (LLPS) that readily cross the cell membrane. A wide range of macromolecules can be quickly recruited within the microdroplets, from small peptides to enzymes as large as 430 kDa to mRNAs. The therapeutic-loaded coacervates bypass endocytosis to directly enter in the cytosol, where they undergo glutathione-mediated release of payload whose bioactivity is retained in the cell, whereas mRNAs exhibit a high transfection efficiency. These peptide coacervates represent a promising platform for intracellular delivery of a large palette of macromolecular therapeutics that have potential in the treatment of various pathologies (e.g. cancers and metabolic diseases), or as carriers for mRNA-based vaccines.