Formulation Strategies for Extracellular Vesicle Delivery

Stem cells therapies have been demonstrating their ability to elicit cell replacement in tissues where there has been a loss of function or damage. However, cell-secreted extracellular vesicles (EVs) have been shown to be responsible for much of these regenerative effects. In the scarred myocardium after an infarct, EVs have shown cardioprotective effects upon injection. However, administration via injection has severe limitations, since much of the administered dose is rapidly cleared from the site of action. Development of implantable EV formulations is then needed. EVs from foetal bovine serum and from mesenchymal stem cells culture media were isolated and characterised. A differential centrifugation workflow was assessed to identify a robust and reproducible isolation method. Both isolation and protein characterisation were found to be highly dependent on the starting material, indicating that a careful assessment is required for each sample. The effect of EVs on stressed cardiomyocytes in culture was assessed in vitro and found to be concentration dependent. FBS EVs were found to be relatively stable over the course of a month at the storage conditions of -80◦ C, 4◦ C and 20◦ C. The feasibility of formulating EVs into a scaffold by electrospinning was explored, initially using fast-dissolving polymers such as poly(vinyl pyrrolidone). Extensive optimisation of processing conditions was undertaken and no difference in pre and postprocessing EVs potency was noted in cell culture assays, confirming electrospinning to be suitable for EV formulation. Finally, development of sustained-release scaffolds for cardiac applications was investigated, based on poly(lactic-co-glycolic acid) and poly(dioxanone). A range of systems was prepared and morphology, flexibility, shrinkage behaviour and mechanical properties explored. However, scaffolds were found to have a burst release at 24 h with no more release detected over the 4 week period. Overall, the results obtained are encouraging and lay the formulation foundations for potential EV clinical applications