	US 12,357,675 B2
	Efficient protein expression in vivo using modified RNA (mod-RNA)
Kenneth R. Chien, Cambridge, MA (US); Leon M. Ptaszek, Newton, MA (US); Kathy Oi-Lan Lui, Boston, MA (US); Lior Zangi, Brookline, MA (US); Wataru Ebina, Boston, MA (US); and Derrick J. Rossi, Roslindale, MA (US)
Assigned to THE GENERAL HOSPITAL CORPORATION; and CHILDREN'S MEDICAL CENTER CORPORATION
Filed by THE GENERAL HOSPITAL CORPORATION, Boston, MA (US); and CHILDREN'S MEDICAL CENTER CORPORATION, Boston, MA (US)
Filed on Jul. 23, 2019, as Appl. No. 16/519,607.
Application 16/519,607 is a continuation of application No. 16/015,460, filed on Jun. 22, 2018, abandoned.
Application 16/015,460 is a continuation of application No. 14/009,351, granted, now 10,086,043, issued on Oct. 2, 2018, previously published as PCT/US2012/028802, filed on Mar. 12, 2012.
Claims priority of provisional application 61/471,584, filed on Apr. 4, 2011.
Claims priority of provisional application 61/471,166, filed on Apr. 3, 2011.
Prior Publication US 2020/0000881 A1, Jan. 2, 2020
This patent is subject to a terminal disclaimer.
Int. Cl. A61K 38/18 (2006.01); A01K 67/0276 (2024.01); A61K 31/4745 (2006.01); A61K 31/513 (2006.01); A61K 31/517 (2006.01); A61K 31/519 (2006.01); A61K 31/555 (2006.01); A61K 31/7068 (2006.01); A61K 31/7115 (2006.01); A61K 33/243 (2019.01); A61K 48/00 (2006.01); A61L 31/10 (2006.01); A61L 31/16 (2006.01); C12N 15/11 (2006.01)

CPC A61K 38/1866 (2013.01) [A01K 67/0276 (2013.01); A61K 31/4745 (2013.01); A61K 31/513 (2013.01); A61K 31/517 (2013.01); A61K 31/519 (2013.01); A61K 31/555 (2013.01); A61K 31/7068 (2013.01); A61K 31/7115 (2013.01); A61K 33/243 (2019.01); A61K 48/005 (2013.01); A61L 31/10 (2013.01); A61L 31/16 (2013.01); C12N 15/111 (2013.01); A01K 2207/05 (2013.01); A01K 2217/075 (2013.01); A01K 2227/105 (2013.01); A01K 2267/03 (2013.01); A01K 2267/0393 (2013.01); A61L 2300/258 (2013.01); C12N 2310/11 (2013.01); C12N 2310/14 (2013.01); C12N 2800/30 (2013.01)]

13 Claims

1. A method for increasing cardiovascular fate cells in cardiac tissue in a subject with myocardial infarction by expressing a VEGF-A protein in a cardiac tissue in vivo, the method comprising contacting the cardiac tissue in vivo by myocardial injection with a composition comprising a synthetic, modified RNA molecule encoding a VEGF-A¹⁶⁵polypeptide,

wherein the synthetic, modified RNA molecule comprises at least one or more nucleoside base modifications selected from the group consisting of: pseudouracil, 2 (thio)pseudouracil, 4 (thio)pseudouracil, 2,4-(dithio)psuedouracil, 5-(alkyl)pseudouracil, 5-(methyl)pseudouracil, 5-(alkyl)-2-(thio)pseudouracil, 5-(methyl)-2-(thio)pseudouracil, 5-(alkyl)-4 (thio)pseudouracil, 5-(methyl)-4 (thio)pseudouracil, 5-(alkyl)-2,4 (dithio)pseudouracil, 5-(methyl)-2,4 (dithio)pseudouracil, 1 substituted pseudouracil, 1 substituted 2(thio)-pseudouracil, 1 substituted 4 (thio)pseudouracil, 1 substituted 2,4-(dithio)pseudouracil, 1 (aminocarbonylethylenyl)-pseudouracil, 1 (aminocarbonylethylenyl)-2(thio)-pseudouracil, 1 (aminocarbonylethylenyl)-4 (thio)pseudouracil, 1 (aminocarbonylethylenyl)-2,4-(dithio)pseudouracil, 1 (aminoalkylaminocarbonylethylenyl)-pseudouracil, 1 (aminoalkylamino-carbonylethylenyl)-2(thio)-pseudouracil, 1 (aminoalkylaminocarbonylethylenyl)-4 (thio)pseudouracil, and 1 (aminoalkylaminocarbonylethylenyl)-2,4-(dithio)pseudouracil,

such that introducing said synthetic, modified RNA molecule to a cell in the cardiac tissue in vivo results in increasing cardiovascular fate cells and reduces fibrosis in the cardiac tissue, and also results in a reduced innate immune response relative to a cell in the cardiac tissue in vivo contacted with a synthetic RNA molecule encoding the polypeptide not comprising said modification, wherein the modified RNA molecule encoding a VEGF-A¹⁶⁵polypeptide promotes WT-1+ epicardial progenitor cells to differentiate into endothelial cells, vascular smooth muscle cells, or cardiomyocytes.