Injured Cardiac Tissue-Targeted Delivery of TGFβ1 siRNA by FAP Aptamer-Functionalized Extracellular Vesicles Promotes Cardiac Repair

Ji-Young Kang,1 Dasom Mun,1 Malgeum Park,1 Gyeongseo Yoo,2 Hyoeun Kim,3 Nuri Yun,4 Boyoung Joung1,2 1Division of Cardiology, Yonsei University College of Medicine, Seoul, 03722, Republic of Korea; 2Graduate School of Medical Science, Brain Korea 21 Project, Yonsei University College of Medicine, Seoul, 03722, Republic of Korea; 3Department of Biochemistry and Molecular Biology, Yonsei University College of Medicine, Seoul, 03722, Republic of Korea; 4GNTPharma Science and Technology Center for Health, Incheon, 21983, Republic of KoreaCorrespondence: Boyoung Joung, Division of Cardiology, Yonsei University College of Medicine, Seoul, 03722, Republic of Korea, Tel +82-2-2228-8447, Fax +82-2-2227-7732, Email [email protected] Nuri Yun, GNTPharma Science and Technology Center for Health, Incheon, 21983, Republic of Korea, Tel +82-31-8005-9910, Fax +82-31-8005-9917, Email [email protected]: Small‐interfering RNA (siRNA) therapy holds significant potential for treating cardiac injury; however, its clinical application is constrained by poor blood stability and insufficient cellular uptake. Extracellular vesicles (EVs) have emerged as an effective delivery system for siRNA in vivo; but their lack of specific cell or tissue‐targeting ability remains a major challenge. Thus, we aimed to develop an EV-based delivery system capable of targeted delivery of therapeutic siRNA to injured cardiac tissue for cardiac repair.Methods: To identify fibroblast activation protein (FAP) as a potential target for delivery to injured cardiac tissue, we analyzed cardiac tissues from patients with heart failure and angiotensin II (Ang II)-treated mice. Injured cardiac tissue-targeting EVs were developed by embedding a cholesterol-conjugated FAP aptamer, which specifically targets FAP, onto human serum-derived EVs (hEV).Results: Our findings revealed that FAP is upregulated after cardiac injury, highlighting its potential as a target for siRNA delivery to injured cardiac tissues. We successfully developed FAP aptamer-functionalized hEV (hEV@FAP) and confirmed their typical EV characteristics, including morphology, size distribution, zeta potential, and marker protein expression. In addition, hEV@FAP demonstrated high targeting selectivity to FAP-positive regions both in vitro and in vivo. To treat cardiac injury, hEV@FAP were loaded with TGFβ 1 siRNA (siTGFβ 1), identified as a molecular target for cardiac repair. In Ang II‐treated mice, intravenous administration of hEV@FAP-siTGFβ 1 effectively reduced Ang II‐induced TGFβ 1 expression in cardiac tissues, attributed to the protective and targeting capabilities of hEV@FAP. Consequently, hEV@FAP-siTGFβ 1 significantly improved cardiac function, reduced myocardial fibrosis, and decreased cardiomyocyte cross‐sectional area (P < 0.05) without inducing systemic toxicity.Conclusion: hEV@FAP represents a novel approach for targeted delivery of therapeutic siRNA to injured cardiac tissues, providing a promising nanomedicine for cardiac repair. Keywords: cardiac injury, fibroblast activation protein, extracellular vesicles, small-interfering RNA, transforming growth factor beta 1