Virtually all cells in the organism secrete extracellular vesicles (EVs), a heterogeneous population of lipid bilayer membrane-enclosed vesicles that transport and deliver payloads of proteins and nucleic acids to recipient cells, thus playing central roles in cell-cell communications. Exosomes, nanosized EVs of endosomal origin, regulate many pathophysiological processes including immune responses and inflammation, tumour growth, and infection. Healthy subjects and patients with different diseases release exosomes with different RNA and protein contents into the circulation, which can be measured as biomarkers. The discovery of exosomes as natural carriers of functional small RNA and proteins has raised great interest in the drug delivery field, as it may be possible to harness these vesicles for therapeutic delivery of miRNA, siRNA, mRNA, lncRNA, peptides, and synthetic drugs. However, systemically delivered exosomes accumulate in liver, kidney, and spleen. Targeted exosomes can be obtained by displaying targeting molecules, such as peptides or antibody fragments recognizing target antigens, on the outer surface of exosomes. Display of glycosylphosphatidylinositol (GPI)-anchored nanobodies on EVs is a novel technique that enables EV display of a variety of proteins including antibodies, reporter proteins, and signaling molecules. However, naturally secreted exosomes show limited pharmaceutical acceptability. Engineered exosome mimetics that incorporate desirable components of natural exosomes into synthetic liposomes or nanoparticles, and are assembled using controllable procedures may be more acceptable pharmaceutically.
Bioengineered EVs for targeted drug delivery
Examples of EV targeting to specific recipient cells are shown. Targeting may be accomplished by engineering the parent cells using fusion constructs between a targeting ligand and an EV transmembrane protein, such as LAMP-2b or PDGFR TM domain. Targeting ligands may include naturally occurring molecules, such as EBV gp350 (which selectively binds to B cells via interaction with the B lineage marker CD21), RVG (which targets nACh receptor on neurons), and sialic acid residues (which mediate uptake by macrophages through binding to CD169). ICAM-1 contributes to B-cell binding through interaction with LFA-1. Examples of synthetic targeting peptides displayed on te EV surface include GE11 (which targets EGFR on tumour cells) and iRGD (which targets integrins and neuropilins). Anti-EGCF-nanobodies linked to GPI-anchors through GGGGS2 linkers have been used to target EGFR on tumour cells. VSV-G indiscriminately facilitates cell fusion. EVs may carry cargo small-molecule drugs and/or nucleic acids (Abbreviations: EBV, Epstein-Barr virus; EGFR, epidermal growth factor receptor; GPI, glycosylphosphatidylinositol; ICAM-1, intercellular adhesion molecule-1; iRGD, internalizing arginine-glycine-asparagine; LAMP-2, lysosome-associated membrane protein-2; LFA-1, lymphocyte function–associated antigen-1; nACh, nicotinic acetylcholine; PDGFR TM domain, platelet-derived growth factor receptor, transmembrane domain; RVG, rabies virus glycoprotein; VSV-G, vesicular stomatitis virus glycoprotein).