Toward Exosome-Based Therapeutics: Isolation, Heterogeneity, and Fit-for-Purpose Potency

Abstract

Exosomes are defined as submicron (30–150 nm), lipid bilayer-enclosed extracellular vesicles (EVs), specifically generated by the late endosomal compartment through fusion of multivesicular bodies with the plasma membrane. Produced by almost all cells, exosomes were originally considered to represent just a mechanism for jettisoning unwanted cellular moieties. Although this may be a major function in most cells, evolution has recruited the endosomal membrane-sorting pathway to duties beyond mere garbage disposal, one of the most notable examples being its cooption by retroviruses for the generation of Trojan virions. It is, therefore, tempting to speculate that certain cell types have evolved an exosome subclass active in intracellular communication. We term this EV subclass “signalosomes” and define them as exosomes that are produced by the “signaling” cells upon specific physiological or environmental cues and harbor cargo capable of modulating the programming of recipient cells. Our recent studies have established that signalosomes released by mesenchymal stem/stromal cells (MSCs) represent the main vector of MSC immunomodulation and therapeutic action in animal models of lung disease. The efficacy of MSC-exosome treatments in a number of preclinical models of cardiovascular and pulmonary disease supports the promise of application of exosome-based therapeutics across a wide range of pathologies within the near future. However, the full realization of exosome therapeutic potential has been hampered by the absence of standardization in EV isolation, and procedures for purification of signalosomes from the main exosome population. This is mainly due to immature methodologies for exosome isolation and characterization and our incomplete understanding of the specific characteristics and molecular composition of signalosomes. In addition, difficulties in defining metrics for potency of exosome preparations and the challenges of industrial scale-up and good manufacturing practice compliance have complicated smooth and timely transition to clinical development. In this manuscript, we focus on cell culture conditions, exosome harvesting, dosage, and exosome potency, providing some empirical guidance and perspectives on the challenges in bringing exosome-based therapies to clinic.

Proteomic and Post‐Translational Modification Profiling of Exosome‐Mimetic Nanovesicles Compared to Exosomes

“…In addition, the quantity and composition of exosomes can be influenced by variations in cell physiological behavior. Cell cultures need to be controlled during every passage to ensure that they do not alter the cargo of exosomes and therapeutic potency . The advantages of using cell‐derived M‐NVs allows for a larger scale production of vesicles from a continuously maintained cell system that is not limited by alterations in cell biology.…”

Engineering mesenchymal stem cells to improve their exosome efficacy and yield for cell‐free therapy

“…The culture condition alone may not be enough to address the additional limitations on the efficacy of MSC-exosomes though. Exosomes derived from different cell types may have preferential targeting towards certain cell types based on their membrane composition thus imparting a differential effect on body systems [37,41,45]. For example, oligodendroglia EVs are preferentially taken up by microglia [36].…”

Extracellular vesicles in cardiovascular homeostasis and disease

“…The cardioprotection conferred by these EVs is not observed when fibroblast EVs are injected into the same models, indicating the specificity of the effect. Stem cell-derived EVs have also shown remarkable cardioprotective and regenerative potential [34]. The reparative capacity of these EVs is likely conferred by efficient delivery of microRNAs that promote homeostatic gene expression and non-inflammatory phenotypes in diseased tissue [35].…”