A functionalized leucine zipper self-assembling hydrogel platform for site-specific extracellular vesicle delivery

Mesenchymal stem cell (MSC)-derived extracellular vesicles (EVs) exhibit significant therapeutic potential across various regenerative applications. When applied to targeted tissue repair such as bone regeneration, precise and site-specific delivery of EVs is essential to ensure optimal therapeutic efficacy. To address this, we evaluated a leucine zipper (LZ)-based self-assembling hydrogel incorporating function-specific motifs as a platform to support site-specific EV delivery in this study. Based on the integrin-mediated binding capability of EVs to extracellular matrix (ECM) components, we generated LZ chimeric proteins containing ECM-derived EV binding motifs and tested their efficacy to bind EVs. As a proof-of-concept approach, we tethered engineered osteoinductive MSC EVs (BMP2 EVs) to RGD-functionalized LZ hydrogels. The combination of these engineered EVs and hydrogels were characterized and evaluated in vitro and in vivo. In vitro results demonstrated the importance of tethering peptides for EV retention within hydrogels as well as the ability of the tethered BMP2 EVs to maintain their physicochemical characteristics and osteoinductive function. When applied in vivo in a rat calvarial defect model, the combination of BMP2 EVs and hydrogels significantly enhanced bone formation and bone quality. Overall, we present a versatile LZ-based self-assembling peptide platform with tunable properties for EV delivery and tissue regeneration.