Enhancing extracellular vesicle cargo loading and functional delivery by engineering protein-lipid interactions

Naturally generated lipid nanoparticles termed extracellular vesicles (EVs) hold significant promise as engineerable therapeutic delivery vehicles. However, active loading of protein cargo into EVs in a manner that is useful for delivery remains a challenge. Here, we demonstrate that by rationally designing proteins to traffic to the plasma membrane and associate with lipid rafts, we can enhance loading of protein cargo into EVs for a set of structurally diverse transmembrane and peripheral membrane proteins. We then demonstrate the capacity of select lipid tags to mediate increased EV loading and functional delivery of an engineered transcription factor to modulate gene expression in target cells. We envision that this technology could be leveraged to develop new EV-based therapeutics that deliver a wide array of macromolecular cargo. Understanding principles that govern protein association with extracellular vesicles should expand their potential as a therapeutic modality. Here, the authors show that by localizing proteins to the plasma membrane and lipid rafts, a variety of proteins can be preferentially loaded into extracellular vesicles.