Engineering Extracellular Vesicles as Delivery Systems in Therapeutic Applications

Abstract

Extracellular vesicles (EVs) are transport vesicles secreted by living cells and released into the extracellular environment. Recent studies have shown that EVs serve as "messengers" in intercellular and inter-organismal communication, in both normal and pathological processes. EVs, as natural nanocarriers, can deliver bioactivators in therapy with their endogenous transport properties. This review article describes the engineering EVs of sources, isolation method, cargo loading, boosting approach, and adjustable targeting of EVs. Furthermore, the review summarizes the recent progress made in EV-based delivery systems applications, including cancer, cardiovascular diseases, liver, kidney, nervous system diseases, and COVID-19 and emphasizes the obstacles and challenges of EV-based therapies and possible strategies.

Keywords: drug delivery; extracellular vesicles; nanocarrier; targeted therapy.

Figure 1

Overview of EV therapeutic application and potential sources. The analysis of EV inclusion tests based on human body fluid samples such as blood or urine helps in disease diagnosis. EV‐based therapies have been shown to be clinically effective in skeletal disorders, neurological disorders, cardiovascular disorders, and liver and kidney disorders. Nonhuman EVs based on bacteria, milk, and plants have been applied as drug carriers to achieve disease treatment.

Figure 2

EVs loading strategies and delivery mechanism. Engineered EVs can recognize target cells via biomarkers consisting of EV‐sorting domains fused with selective proteins and loading cargo by electroporation, transfection, incubation, and sonication. EVs are loadable with exogenous therapeutic cargos, including A) drugs, B) RNAs, C) proteins, and D) CRISPR/Cas9, similar to mainstream drug delivery systems like E) polymeric nanoparticles, F) polymeric micelles, and G) liposomes.

Figure 3

An overview of sources, isolation, boosting approach, and the application of EVs. A) EVs can be secreted by various cells with different biological functions. B) The methods of isolating EVs, including density‐gradient centrifugation, ultracentrifugation, ultrafiltration, size‐exclusion chromatography, precipitation, microfluidics, and immune‐affinity capture. C) The methods for boosting EV secretion, including physical signals, molecular interference, environmental factors and external inducers. D) The application of EVs as biomarkers or nanocarriers for disease diagnosis, prognosis, or treatment.

Figure 4

EVs‐mediated CRISPR/Cas9 system. The EVs can be engineered to target mutant genes or virus‐infected cells by fusing CD63 with GFP, while the loaded Cas9 proteins fused to the GFP nanobody. The delivery system can effectively inactivate target genes, inhibit tumor growth, and reduce virus replication rate.

Figure 5

The role of EVs as endogenous carriers in treating A) head and neck squamous cell carcinoma, B) nasopharyngeal carcinoma, and C) chronic myeloid leukemia.

Figure 6

The treatment of cardiovascular diseases. Vascular endothelial EVs increased the respiratory capacity of normoxic cardiomyocytes, and EVs rescued cardiomyocytes exposed to IRI, possibly by providing bioactive cargos to support multiple metabolic.

Figure 7

The role of EVs as endogenous carriers in neurodegenerative disease and preeclampsia. A) In Parkinson's disease, EVs are responsible for transporting the misfolded proteins to neighboring cells. In Alzheimer's disease, a fraction of β‐amyloid peptides were stored in the multivesicular bodies and released on binding to EVs. EV‐associated protein, Alix, was specifically enriched in amyloid plaques of patients with AD. B) Comparison between amniotic fluid EVs from healthy and preeclampsia pregnancies showed that the surface marker CD105 was especially upregulated, meaning that EVs from amniotic fluid of preeclamptic pregnancies might have the potential property of anti‐angiogenic.

Figure 8

The role of EVs in the mechanism of cytokine storm syndrome and COVID‐19. CSS is an EV‐involved profound inflammatory immune response in which the expression of IL‐2, IL‐6, IL‐7, TNF‐α, INF‐γ, MCP, MIP 10, and GSCF is upregulated. SARS‐CoV‐2 can induce the secretion of pro‐inflammatory cytokines to trigger alveolar edema, dyspnea, hypoxemia, and systemic inflammatory response syndrome. EVs targeting CD24 inhibit the NF‐κB pathway and restrain them from secreting proinflammatory cytokines, suggesting that CD24 is a potential immune target in COVID‐19.