Project Summary
Intercellular communication based on the delivery of miRNAs from one cell to adjacent or remote cells is now
known to be a critical part of normal cellular/organismal function. Recent work has demonstrated that a new
class of extracellular nanoparticles, exomeres, may be more important for miRNA-mediated intercellular
communication than extracellular vesicles (EVs). Exomeres are small, non-membranous secreted particles
that are different from EVs. Compared to EVs, exomeres are smaller, have distinct protein and nucleic acid
content, and are not encapsulated by lipid membranes. Also, exomeres contain Argonaute-2 (Ago2), one of
the principal proteins of the RNA interference (RNAi) pathway. With respect to intercellular communication,
exomeres contain miRNAs, which are likely bound by Ago2, and the EGFR ligand, amphiregulin, which could
initiate receptor-mediated endocytosis. Thus, exomeres are a natural RNA-containing nanoparticle evolved to
have molecular and physical properties that facilitate efficient endocytosis and intracellular processing of the
miRNA cargo by recipient cells. In this work, we will begin to explore whether exomeres could be used for
delivery of exogenous miRNAs (or siRNAs) for therapeutic applications.
The mechanisms of exomere biogenesis, secretion, endocytosis, and intracellular trafficking remain poorly
understood. This is due, in part, to the challenges of isolating exomeres. Current exomere isolation approaches
have allowed for preliminary characterization of exomeres, but further study of exomeres, and their critical
roles in RNA-mediated intercellular communication, will require approaches for isolating exomeres that yield
high concentrations of reasonably homogeneous complexes. In this work, we will develop a novel, scalable
exomere isolation approach based on tangential flow filtration (TFF) that will enhance our ability to study
exomeres and the endocytosis and intracellular trafficking of exomeres by recipient cells. The overarching
goal of this work is to demonstrate the utility of TFF for exomere isolation and to demonstrate that exomeres
isolated by TFF are identical to those isolated by current approaches. Given that TFF is compatible with
possible industrial production methods for exomeres, our results will provide the foundation for future efforts to
use exomeres for RNA delivery in clinical applications.