Abstract 471: Development of a Cardiomyocyte Targeting Delivery System Utilizing Cardiosphere-Derived Cell Exosomes

Abstract

Research has shown that treating the heart with stem cell populations, including cardiosphere-derived cells (CDCs), post myocardial infarction stimulates regeneration, angiogenesis, and functional improvement. While this treatment has shown promise in early stage clinical trials, there remains a gap in the ability to efficiently deliver tissue-specific agents directly to the heart while avoiding nonspecific delivery to other organs. Our aim was to develop an efficient delivery system that can target cardiomyocytes and transport drugs, siRNA, and/or miRNA to aid in the treatment of cardiovascular diseases. Methods: Lamp-2b was cloned with cDNA from C2C12 cells and ligated downstream of the CMV promoter in a lentiviral transfer plasmid. Primers designed to encode a cardiomyocyte targeting peptide were used to introduce the targeting ligand between XhoI and BspEI at the N terminus of Lamp-2b forming the final vector (LV-CMP). We subsequently generated CMP lentivirus by triple transfection of LV-CMP with helper plasmids pLP/VSVG and psPAX2 into 293T cells. CDCs were transduced at 1 MOI in the presence of polybrene. Exosomes were isolated from infected and non-infected CDCs and labelled with a nucleic acid dye. Cultured mouse cardiomyocytes were incubated with 1 ug of fluorescent labeled CDC-exosomes or CMP CDC-exosomes for 24 hours then imaged. Results: To achieve exosomal cardiomyocyte targeting, we engineered CDCs to express Lamp-2b, an exosomal membrane protein, fused to a cardiomyocyte specific peptide. Primary cardiomyocytes showed a 3.8-fold increase in uptake of targeted CMP CDC-exosomes in vitro when compared with their non-engineered CDC control, establishing the efficacy of our system and laying the groundwork for further experiments to assess the efficiency of cardiac localization and cardiomyocyte uptake in vivo. Conclusions: Cardiomyocytes show enhanced in vitro uptake of CDC-derived exosomes engineered to express a cardiomyocyte targeting peptide fused with Lamp-2b. Improving exosomal uptake by cardiomyocytes following myocardial infarction would allow for improved delivery of beneficial drug and gene therapies and may result in an increase in viable myocardium and improved functional capacity of the heart.