PROJECT SUMMARY The proposed research project addresses the need for a bioengineering approach to a smart and targeted exosome-based therapeutic delivery platform that can promote tissue repair after myocardial infarction (MI). Accumulating evidence in both preclinical and clinical settings has consistently indicated the exosomes secreted from adult stem/stromal cells orchestrate the principal modes of action of cell therapy to repair the injured heart muscle following MI. Exosome-based therapeutics represent a paradigm shift from conventional cell-based approaches to cardiovascular therapy. However, current exosome-based therapeutics suffer from several problems including lack of targetability, complicated fabrication process to load additional therapeutic cargos, and expense. In addition, technologies that enable controlled delivery of exosome-based products have yet to be explored. To overcome these challenges, we propose to develop a smart cardiotherapeutic nanocapsule (CardioXo) for the targeted and controlled delivery of both mesenchymal stem cell-derived exosomes (MSC-Exo) and therapeutic proteins to promote heart repair post-MI. Our central hypotheses are that (1) the CardioXo functionalized with both exosomes and cardioprotective proteins (e.g., prokineticin 2, agrin, insulin-like growth factor 1, myeloid-derived growth factor) can be engineered via an electrostatic layer-by-layer assembly process combined with a novel enzyme-mediated radical polymerization (EMRP) technology; and (2) the CardioXo can enable the delivery of combination therapeutics in response to the upregulation of matrix metalloproteinases (MMPs), a hallmark of cardiac ischemic injury, and trigger the reparative signaling pathways mediated by therapeutic proteins to repair the ischemic heart. The innovation of the proposed CardioXo strategy includes (1) targeting to the injured cardiomyocytes; (2) ability to deliver combination therapeutics in an on- demand fashion by leveraging cardiac pathology in the ischemic heart; (3) dual-regenerative mechanism for targeted heart repair that combines the exosome-mediated paracrine mechanism similar to stem cell therapy with the cardioprotective protein-mediated signaling pathways. We will fabricate and optimize the CardioXo loaded with various cardioprotective proteins and assess the in vitro bioactivity of CardioXo. The safety, functional benefits, and the potential modes of action of CardioXo therapy will be investigated in rats with ischemic heart injury. Successful execution of the proposed research project will establish a mild and facile approach to exosome functionalization and form the foundation of an innovative and off-the-shelf exosome- based therapy based on smart and targeted nanomedicine for the repair and regeneration of the post-MI heart. Although this particular grant application targets the heart, the CardioXo strategy represents a platform technology that can be generalized to the repair and regeneration of mul...