US 11,739,300 B2
RNA preparations comprising purified modified RNA for reprogramming cells
Katalin Kariko, Rydal, PA (US); Drew Weissman, Wynnewood, PA (US); Gary Dahl, Madison, WI (US); Anthony Person, Madison, WI (US); Judith Meis, Fitchburg, WI (US); and Jerome Jendrisak, Madison, WI (US)
Filed by The Trustees of the University of Pennsylvania, Philadelphia, PA (US)
Filed on Mar. 23, 2020, as Appl. No. 16/827,098.
Application 16/827,098 is a continuation of application No. 15/994,093, filed on May 31, 2018, granted, now 11,028,370.
Application 15/994,093 is a continuation of application No. 15/160,062, filed on May 20, 2016, granted, now 10,006,007, issued on Jun. 26, 2018.
Application 15/160,062 is a continuation of application No. 14/801,075, filed on Jul. 16, 2015, granted, now 9,371,511, issued on Jun. 21, 2016.
Application 14/801,075 is a continuation of application No. 14/644,680, filed on Mar. 11, 2015, granted, now 9,163,213, issued on Oct. 20, 2015.
Application 14/644,680 is a continuation of application No. 12/962,468, filed on Dec. 7, 2010, granted, now 9,012,219, issued on Apr. 21, 2015.
Claims priority of provisional application 61/267,312, filed on Dec. 7, 2009.
Prior Publication US 2021/0024895 A1, Jan. 28, 2021
This patent is subject to a terminal disclaimer.
Int. Cl. C12N 5/074 (2010.01); C12N 15/87 (2006.01); C12N 15/85 (2006.01); A61K 38/17 (2006.01); A61K 38/18 (2006.01); A61K 38/44 (2006.01); A61K 38/46 (2006.01); A61K 38/50 (2006.01); A61K 48/00 (2006.01); C07K 14/47 (2006.01); C07K 14/505 (2006.01); C12N 9/02 (2006.01); C12N 9/22 (2006.01); C12N 9/78 (2006.01); C12N 15/117 (2010.01)
CPC C12N 5/0696 (2013.01) [A61K 38/1709 (2013.01); A61K 38/1816 (2013.01); A61K 38/44 (2013.01); A61K 38/465 (2013.01); A61K 38/50 (2013.01); A61K 48/005 (2013.01); A61K 48/0075 (2013.01); C07K 14/47 (2013.01); C07K 14/4712 (2013.01); C07K 14/505 (2013.01); C12N 9/0075 (2013.01); C12N 9/22 (2013.01); C12N 9/78 (2013.01); C12N 15/117 (2013.01); C12N 15/85 (2013.01); C12N 15/87 (2013.01); C12N 2310/17 (2013.01); C12N 2310/335 (2013.01); C12N 2320/30 (2013.01); C12N 2501/602 (2013.01); C12N 2501/603 (2013.01); C12N 2501/604 (2013.01); C12N 2501/605 (2013.01); C12N 2501/606 (2013.01); C12N 2501/608 (2013.01); C12N 2506/02 (2013.01); C12Y 114/13039 (2013.01); C12Y 301/04012 (2013.01); C12Y 305/04004 (2013.01)] 14 Claims
 
1. A method comprising:
administering a single dose of a purified RNA preparation to mammalian cells or a mammalian subject without triggering a detectable innate immune response,
wherein said purified RNA preparation was made by a process comprising purifying a preparation of in vitro-synthesized RNA molecules obtained by a process comprising in vitro transcription (IVT),
wherein said in vitro-synthesized RNA molecules encode at least one recombinant protein and comprise a modified nucleoside selected from ψ, m1ψ, m5U, mo5U, and s2U in place of at least a portion of uridine nucleosides in said in vitro-synthesized RNA molecules,
wherein said purifying uses a purification process that removes RNA contaminant molecules comprising double-stranded RNA (dsRNA) molecules that are toxic to mammalian cells by inducing an innate immune response,
wherein said purified RNA preparation is free of said RNA contaminant molecules such that less than 0.01% of the total RNA in said purified RNA preparation consists of said RNA contaminant molecules based on dsRNA dot blotting assays that use a dsRNA-specific monoclonal antibody (mAb) selected from J2 mAb and K1 mAb to quantify the amount of said RNA contaminant molecules in said purified RNA preparation that is spotted on a membrane.