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.
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