Orthogonal analysis reveals inconsistencies in cargo loading of extracellular vesicles.

Autor: Lowe NM; Department of Biomedical Engineering University of California Davis California USA., Mizenko RR; Department of Biomedical Engineering University of California Davis California USA., Nguyen BB; Department of Biomedical Engineering University of California Davis California USA., Chiu KL; Department of Biomedical Engineering University of California Davis California USA., Arun V; Department of Biomedical Engineering University of California Davis California USA., Panitch A; Department of Biomedical Engineering University of California Davis California USA.; Wallace H. Coulter Department of Biomedical Engineering Georgia Institute of Technology and Emory University Atlanta Georgia USA., Carney RP; Department of Biomedical Engineering University of California Davis California USA.
Jazyk: angličtina
Zdroj: Journal of extracellular biology [J Extracell Biol] 2024 Aug 23; Vol. 3 (8), pp. e70003. Date of Electronic Publication: 2024 Aug 23 (Print Publication: 2024).
DOI: 10.1002/jex2.70003
Abstrakt: Since extracellular vesicles (EVs) have emerged as a promising drug delivery system, diverse methods have been used to load them with active pharmaceutical ingredients (API) in preclinical and clinical studies. However, there is yet to be an engineered EV formulation approved for human use, a barrier driven in part by the intrinsic heterogeneity of EVs. API loading is rarely assessed in the context of single vesicle measurements of physicochemical properties but is likely administered in a heterogeneous fashion to the detriment of a consistent product. Here, we applied a suite of single-particle resolution methods to determine the loading of rhodamine 6G (R6G) surrogate cargo mimicking hydrophilic small molecule drugs across four common API loading methods: sonication, electroporation, freeze-thaw cycling and passive incubation. Loading efficiencies and alterations in the physical properties of EVs were assessed, as well as co-localization with common EV-associated tetraspanins (i.e., CD63, CD81 and CD9) for insight into EV subpopulations. Sonication had the highest loading efficiency, yet significantly decreased particle yield, while electroporation led to the greatest number of loaded API particles, albeit at a lower efficiency. Moreover, results were often inconsistent between repeated runs within a given method, demonstrating the difficulty in developing a rigorous loading method that consistently loaded EVs across their heterogeneous subpopulations. This work highlights the significance of how chosen quantification metrics can impact apparent conclusions and the importance of single-particle characterization of EV loading.
Competing Interests: The authors declare no conflicts of interest.
(© 2024 The Author(s). Journal of Extracellular Biology published by Wiley Periodicals LLC on behalf of International Society for Extracellular Vesicles.)
Databáze: MEDLINE
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