Abstract

Extracellular Vesicle-mediated RNA Delivery: From Mechanistic Insights Towards Therapeutic Applications

Pieter Vader, Research Fellow, University Medical Center Utrecht

Extracellular vesicles (EVs) form an endogenous system for information transfer between cells. Since the recent discovery that EVs are also capable of functionally transferring RNA molecules, they are increasingly being considered as therapeutic RNA delivery systems. Despite extensive research into the engineering of EVs for RNA delivery, our understanding of the pathways and mechanisms regulating EV-mediated RNA delivery and processing is limited. Moreover, little is known about how their intrinsic RNA delivery efficiency compares to current synthetic RNA delivery systems. Here, we developed a novel CRISPR/Cas9-based reporter system in which eGFP expression is activated upon functional delivery of targeting single guide RNAs (sgRNAs) that allows study of EV-mediated RNA transfer at single-cell resolution. We employed this system to compare the delivery efficiency of EVs to clinically approved state-of-the-art DLin-MC3-DMA lipid nanoparticles and several in vitro transfection reagents. We found that EVs delivered RNA several orders of magnitude more efficiently than these synthetic systems. In addition, we prepared EV-liposome hybrid nanoparticles and evaluated them as siRNA delivery systems in terms of cellular uptake, toxicity, and gene-silencing efficacy [4]. We show that hybrids combine benefits of both synthetic and biological drug delivery systems and might serve as future therapeutic carriers of siRNA. Our data underline the potential of EVs as RNA delivery vehicles and highlight the need to study the mechanisms by which EVs achieve their efficiency. This may in turn contribute to the development of more efficient EV-based RNA delivery systems and accelerate clinical adoption of therapeutic EVs.