"Triplet" polycistronic vectors encoding Gata4, Mef2c, and Tbx5 enhances postinfarct ventricular functional improvement compared with singlet vectors

Megumi Mathison, Vivek P Singh, Robert P Gersch, Maricela O Ramirez, Austin Cooney, Stephen M Kaminsky, Maria J Chiuchiolo, Ahmed Nasser, Jianchang Yang, Ronald G Crystal, Todd K Rosengart
Journal of Thoracic and Cardiovascular Surgery 2014, 148 (4): 1656-1664.e2

OBJECTIVE: The in situ reprogramming of cardiac fibroblasts into induced cardiomyocytes by the administration of gene transfer vectors encoding Gata4 (G), Mef2c (M), and Tbx5 (T) has been shown to improve ventricular function in myocardial infarction models. The efficacy of this strategy could, however, be limited by the need for fibroblast targets to be infected 3 times--once by each of the 3 transgene vectors. We hypothesized that a polycistronic "triplet" vector encoding all 3 transgenes would enhance postinfarct ventricular function compared with use of "singlet" vectors.

METHODS: After validation of the polycistronic vector expression in vitro, adult male Fischer 344 rats (n=6) underwent coronary ligation with or without intramyocardial administration of an adenovirus encoding all 3 major vascular endothelial growth factor (VEGF) isoforms (AdVEGF-All6A positive), followed 3 weeks later by the administration to AdVEGF-All6A-positive treated rats of singlet lentivirus encoding G, M, or T (1×10(5) transducing units each) or the same total dose of a GMT "triplet" lentivirus vector.

RESULTS: Western blots demonstrated that triplet and singlet vectors yielded equivalent GMT transgene expression, and fluorescence activated cell sorting demonstrated that triplet vectors were nearly twice as potent as singlet vectors in generating induced cardiomyocytes from cardiac fibroblasts. Echocardiography demonstrated that GMT triplet vectors were more effective than the 3 combined singlet vectors in enhancing ventricular function from postinfarct baselines (triplet, 37%±10%; singlet, 13%±7%; negative control, 9%±5%; P<.05).

CONCLUSIONS: These data have confirmed that the in situ administration of G, M, and T induces postinfarct ventricular functional improvement and that GMT polycistronic vectors enhance the efficacy of this strategy.

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