Engineered EVs designed to target diseases of the CNS.

Diseases of the central nervous system (CNS) are challenging to treat, mainly due to the blood-brain barrier (BBB), which restricts drugs in circulation from entering target regions in the brain. To address this issue extracellular vesicles (EVs) have gained increasing scientific interest as carriers able to cross the BBB with multiplex cargos. EVs are secreted by virtually every cell, and their escorted biomolecules are part of an intercellular information gateway between cells within the brain and with other organs. Scientists have undertaken efforts to safeguard the inherent features of EVs as therapeutic delivery vehicles, such as protecting and transferring functional cargo, as well as loading them with therapeutic small molecules, proteins, and oligonucleotides and targeting them to specific cell types for the treatment of CNS diseases. Here, we review current emerging approaches that engineer the EV surface and cargo to improve targeting and functional responses in the brain. We summarize existing applications of engineered EVs as a therapeutic delivery platform for brain diseases, some of which have been evaluated clinically.

Exogenous loading of extracellular vesicles, virus-like particles, and lentiviral vectors with supercharged proteins.

Cell membrane-based biovesicles (BVs) are important candidate drug delivery vehicles and comprise extracellular vesicles, virus-like particles, and lentiviral vectors. Here, we introduce a non-enzymatic assembly of purified BVs, supercharged proteins, and plasmid DNA called pDNA-scBVs. This multicomponent vehicle results from the interaction of negative sugar moieties on BVs and supercharged proteins that contain positively charged amino acids on their surface to enhance their affinity for pDNA. pDNA-scBVs were demonstrated to mediate floxed reporter activation in culture by delivering a Cre transgene. We introduced pDNA-scBVs containing both a CRE-encoding plasmid and a BV-packaged floxed reporter into the brains of Ai9 mice. Successful delivery of both payloads by pDNA-scBVs was confirmed with reporter signal in the striatal brain region. Overall, we developed a more efficient method to load isolated BVs with cargo that functionally modified recipient cells. Augmenting the natural properties of BVs opens avenues for adoptive extracellular interventions using therapeutic loaded cargo.