Cell membrane-based biovesicles (BVs) are important candidate drug delivery vehicles and comprise extracellular vesicles, virus-like particles, and lentiviral vectors. Here, Harvard Medical School researchers 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. The researchers 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, they have 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.
Two component delivery with supercharged lentivius vectors (scLVVs) to single cell demonstrated through bioluminescent reporter activity in vitro and in vivo
a FLEx reports on complex payload delivery with pDNA-scLVVs. Illustration describing that a plasmid (pDNA) encoding for CRE is associated with our supercharging procedure to a lentiviral vector (LVV) encoding for a FLEx reporter. b pDNA-scLVV delivers both Cre and FLEx transgenes to recipient cells. c FLEx activity post-pDNA-LVVs uptake by recipient cells confirmed with qPCR. d Delivery of multi-component payload to the brain with pDNA-scLVVs. e Activation of pDNA-scLVV delivered FLEx reporter in the brain. f FLEx reporter activity at pDNA-scLVV injection site. g FLEx reporter activity outside pDNA-scLVV injection site. h Mouse Ai9 reporter to demonstrate local delivery of pDNA-scLVV transgenes in striatum. i tdTomato expression levels as quantitative measure of Ai9 reporter activity with pDNA-scLVVs in the brain. j Floxing events to detect Ai9 reporter mice post-pDNA-scLVVs mediated delivery of Cre.