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

Yue Sun,Farid Ghadessy,Zhi Wei Lim,Sze Yi Lau,Shi Chieh Chang,Anthony Partridge,Ali Miserez

doi:10.1038/s41557-021-00854-4

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 often employ nanoscale carriers that can get trapped in endosomal compartments. Here we report conjugated peptides that form pH- and redox-responsive coacervate microdroplets by liquid-liquid phase separation that readily cross the cell membrane. A wide range of macromolecules can be quickly recruited within the microdroplets, including small peptides, enzymes as large as 430 kDa and messenger RNAs (mRNAs). The therapeutic-loaded coacervates bypass classical endocytic pathways to enter the cytosol, where they undergo glutathione-mediated release of payload, the bioactivity of which is retained in the cell, while mRNAs exhibit a high transfection efficiency. These peptide coacervates represent a promising platform for the intracellular delivery of a large palette of macromolecular therapeutics that have potential for treating various pathologies (for example, cancers and metabolic diseases) or as carriers for mRNA-based vaccines.

Highlights

Biomacromolecules are highly promising therapeutic modalities to treat various diseases
We have found that micrometre-sized peptide coacervates obtained by liquid–liquid phase separation (LLPS), within which both proteins[22] and low-molecular-weight compounds[23] can be recruited, are capable of crossing the cell membrane through an endocytosis-independent pathway[23], potentially opening new avenues for intracellular delivery of therapeutics
We hypothesized that peptide coacervates could be used for the intracellular delivery of a broad palette of macromolecular therapeutics featuring a wide range of molecular weights and isoelectric points (IEPs)

Summary

Introduction

Biomacromolecules are highly promising therapeutic modalities to treat various diseases. The therapeutic-loaded coacervates bypass classical endocytic pathways to enter the cytosol, where they undergo glutathione-mediated release of payload, the bioactivity of which is retained in the cell, while mRNAs exhibit a high transfection efficiency These peptide coacervates represent a promising platform for the intracellular delivery of a large palette of macromolecular therapeutics that have potential for treating various pathologies (for example, cancers and metabolic diseases) or as carriers for mRNA-based vaccines. Biomacromolecules, including peptides[1], proteins[2,3] and RNAs4–6, are promising therapeutic modalities for the treatment of various diseases owing to key advantages such as high potency, specificity and safety[7] Their therapeutic potential has not yet been fully realized due to their poor cell membrane permeability and/or endosomal entrapment that limits their intracellular exposure[8]. Therapeutic release these robust conjugated peptide coacervates can be used as general intracellular delivery vehicles for a broad range of macromolecular therapeutics

Methods

Results

Conclusion