The evolving paradigm of extracellular vesicles in intercellular signaling and delivery of therapeutic RNAs

Extracellular vesicles (EVs) are approximately 30- to 150-nM particles that are generated by cells and are thought to mediate the transfer of bioactive cargo.¹ Substantial evidence suggests that an under-appreciated role of EVs is the mediation of intercellular signaling via microRNAs (miRNAs).^1,2 However, the concept that miRNAs packaged into EVs mediate intercellular signaling has recently been called into question by Albanese and Chen et al.,³ who report that miRNAs do not seem to be packaged appreciably into EVs and, furthermore, when they are, they have little to no effect on recipient cells. These observations call into question myriad publications over the last decade that suggested that EV-packaged miRNAs alter target cell function and play a role in disease development and even, for example, in reprogramming of cells toward an oncogenic phenotype.^2,4

Interestingly, the findings from Albanese and Chen et al. are supported to a large extent by observations from others who actively try to engineer EVs to deliver therapeutic cargo: not only miRNAs, but also other small non-coding RNAs,⁵ gene-encoding mRNAs,⁶ zinc finger nucleases,^7,8 and even entire CRISPR/Cas systems.^9,10 Indeed, a common theme from these studies is that native EVs are not necessarily efficient in cargo delivery. Instead, significant engineering of the EV-producing cells or alteration of EVs post-production seems to be required for the functional packaging of therapeutically relevant RNA and other cargos and their release into recipient cells.6, 7, 8 These manipulations may include fusion of RNA-binding domains to transmembrane proteins (e.g., CD63) and introduction of elements to promote endosomal escape.^2,3

Collectively, the need for extensive manipulation of EV-producing cells and the observations by Albanese and Chen et al. suggest that native, unmanipulated EVs may not be the universally effective intercellular communicators that so many previously published studies have suggested them to be. Perhaps EVs represent agents mostly involved in expunging cell byproducts and/or in a metabolic recycling pathway rather than an intercellular signaling pathway after all?⁷ Although these demonstrated EV roles of “taking out the trash” and “putting food on the table” are often disregarded as unimportant characteristics of EV biology in the literature, they are vital functions in the life of the cell. However, meagre cell fusion and cargo delivery abilities also do not necessarily preclude other roles in signaling. EVs may indeed serve as a scaffold for “kiss-and-run” interactions, in which the coordination of targeting and signaling ligands achieves more potent transduction than soluble ligands alone.⁷

If past is prologue, one thing is clear: our understanding of the inner workings of the cell and the role and function of EVs within it is merely a model that is constantly challenged and evolving. No doubt, future studies will provide greater insights, and invariably EVs may prove to be multifactorial elements involved in various aspects of cellular biology that can be taken advantage of for therapeutic gain. They are already emerging as the next-generation delivery agent that offer unique properties not imbued in lipid nanoparticles or viral vectors, ultimately resulting in a significant expansion of the gene therapy delivery tool kit.