Exosomes as natural delivery carriers for programmable therapeutic nucleic acid nanoparticles (NANPs)

Abstract

With recent advances in nanotechnology and therapeutic nucleic acids (TNAs), various nucleic acid nanoparticles (NANPs) have demonstrated great promise in diagnostics and therapeutics. However, the full realization of NANPs' potential necessitates the development of a safe, efficient, biocompatible, stable, tissue-specific, and non-immunogenic delivery system. Exosomes, the smallest extracellular vesicles and an endogenous source of nanocarriers, offer these advantages while avoiding complications associated with manufactured agents. The lipid membranes of exosomes surround a hydrophilic core, allowing for the simultaneous incorporation of hydrophobic and hydrophilic drugs, nucleic acids, and proteins. Additional capabilities for post-isolation exosome surface modifications with imaging agents, targeting ligands, and covalent linkages also pave the way for their diverse biomedical applications. This review focuses on exosomes: their biogenesis, intracellular trafficking, transportation capacities, and applications with emphasis on the delivery of TNAs and programmable NANPs. We also highlight some of the current challenges and discuss opportunities related to the development of therapeutic exosome-based formulations and their clinical translation.

Keywords: Drug delivery; Exosomes; Extracellular vesicle; Immunorecognition; Nanotechnology; Nucleic acid nanoparticles (NANPs); Therapeutic Nucleic Acids (TNA).

Figure 1.

Proteins, lipids, and nucleic acids as representative biological components of exosome content.

Figure 2.

EV biogenesis and trafficking. (A) Exosomes are taken up by recipient cells via direct fusion of the exosomal membrane and the plasma membrane of the recipient cell, leading to direct release of their contents into the recipient cell’s cytoplasm. Alternatively, receptor-mediated endocytosis and micropinocytosis involve exosome uptake into endosomes, but the exosomal contents are released via back fusion of the exosomal membrane and the recipient cell’s endosomal membrane. (B) Biogenesis of microvesicles, exosomes, and apoptotic bodies. Microvesicles shed from the cell via budding of the plasma membrane. Exosome biogenesis begins with internalization of membrane proteins and lipid complexes via endocytosis and engulfment of cytosolic proteins and nucleic acids into the intraluminal vesicles (ILVs) via inward budding of the endosomal membrane. With endosome maturation, late endosomes enclose numerous ILVs to become MVBs. Some MVBs are degraded in the lysosome; exosome secretion occurs when MVB fuses with the plasma membrane. During apoptosis, cell disassembly generates apoptotic bodies, which are released via blebbing and protrusion.

Figure 3.

The endosomal sorting complex required for transport (ESCRT) pathway for cargo recognition and sorting. The ESCRT-0 complex recognizes and sequesters ubiquitylated cargo, whereas the ESCRT-I and -II complexes are responsible for membrane deformation which yields buds with ubiquitylated cargo. The ESCRT-III complex subsequently drives vesicle scission, resulting in ILV formation. Various ESCRT-accessory proteins participate in and assist with these processes.

Figure 4.

Schematic representations of some examples of exosome-mediated delivery of various cargos. (A) NGF mRNA delivered by exosomes decorated with RVG. (B) 3WJ RNA nanoparticles delivered by exosomes decorated with cholesterol and ligand. (C) RNA cube, RNA ring, fiber sense, and fiber antisense delivered by exosomes. (D) Dex delivered by exosomes decorated with Folic acid-PEG and cholesterol. (E) Biotinylated CpG DNA delivered by exosomes decorated with streptavidin-lactadherin fusion protein.

Figure 5.

Different methods for exosome isolation and drug encapsulation. (A) Commonly used methods for exosome isolation. (B) Commonly used methods for drug loading to exosomes.

Figure 6.

Schematic summary of the workflow of exosome-mediated delivery of different functionalized cargos. Exosomes are released by cells. After isolation and purification, exosome surfaces can be engineered for specific targeting, or modified with covalent linkages for various motifs. Therapeutic cargos are loaded into the exosome lumen or linked on the exosome surface. After uptake by the destination cells, the therapeutic motifs are able to elicit intended effects in target cells.