Gene Therapy with miR-199a-3p for Myocardial Infarction

One of the striking differences between the adult and the embryonic heart in mammals is the incapacity of cardiomyocytes to proliferate as a response to damage. It has been previously shown that microRNA hsa-miR-199a-3p promotes cell cycle re-entry of rodent adult cardiomyocytes, ex vivo, and favors almost complete recovery of cardiac functional parameters when injected in infarcted mouse hearts by stimulating cardiac regeneration.1 The aim of this study was to test the therapeutic action of miR-199a in a pre-clinical, large animal model of myocardial infarction (MI). The precursor DNA encoding for hsa-mir-199a was encapsidated in serotype 6 adeno-associated viral vectors (AAV6) for cardiac delivery, in vivo. MI was induced in pigs by occluding the left anterior descending artery (LAD) immediately below the 1st diagonal branch for 90 minutes and a total of 2x1013 AAV6-hsa-mir-199a-3p (n=10) or 2x1013empty-AAV6 (containing and empty polylinker; n=9) was injected intramyocardially during reperfusion in 20 different sites along the infarct border zone. Non-infarcted pigs (n=6) were used as normal controls. Pigs underwent cardiac magnetic resonance imaging (MRI) without and with gadolinium-delayed contrast enhancement at 2, 7, 28 and 56 days post-MI. The infarct area was not significantly different between the two infarcted groups at 2 days post-MI. However, at 28 days, the scar involved a percentage of the total left ventricle myocardium of 22.95±2.5% in the empty AAV6 group, and only 11.49±1.28% in the AAV6-hsa-mir-199a-3p group. Consistently, ejection fraction was 65.3±1.97% (n.s. vs normal control) in the AAV6-hsa-mir-199a-3p group, markedly preserved compared to 53.7±3.04% in the empty AAV6 group (P<0.05). Finally, a 12-segment analysis of left ventricular circumferential and radial shortening, as measured by tagging-MRI, was performed to generate a 12-point curve. The area under the curve (in arbitrary units) for Ecc (circumferential shortening) was -259.3±19.8 in the AAV6-hsa-mir-199a-3p group (n.s. vs normal control AUC=-285±6.13) and -177.6±26.2 in the empty-AAV6 group (P<0.05 vs normal control, and treated group). The AUC for ERR (Radial shortening) was 338.15±67.7 in the AAV6-hsa-mir-199a-3p group, compared to 213.4±35.18 (P<0.05) in the empty AAV6 group. In both infarcted groups, the AUC of ERR was lower compared to control animals (P<0.05 vs 540±24.6). Histological revealed that AAV6-has-miR-199a-3p treatment does not increase cardiomyocyte hypertrophy at day 56 after infarction. BrdU (Bromodeoxyuridine) and phospho histone 3 quantification assays did not show any difference between two groups. The expression analyses of genes involved in homeostatic response to heart failure revealed ANP (atrial natriuretic peptide) downregulation and MYH6/MYH7 (Myosin heavy chain6/7) overespression, which may undirectly indicate improved myocardial contractility. All the above mentioned parameters indicate extensive recovery of regional contractility in hearts receiving miR-199a-3p. These are the first data to show, in a clinically relevant animal model, the efficacy of a new therapeutic strategy for acute MI based on cardiac delivery of a small regulatory RNA.