RNA preparations comprising purified modified RNA for reprogramming cells

摘要

The present invention provides compositions and methods for reprogramming somatic cells using purified RNA preparations comprising single-strand mRNA encoding an iPS cell induction factor. The purified RNA preparations are preferably substantially free of RNA contaminant molecules that: i) would activate an immune response in the somatic cells, ii) would decrease expression of the single-stranded mRNA in the somatic cells, and/or iii) active RNA sensors in the somatic cells. In certain embodiments, the purified RNA preparations are substantially free of partial mRNAs, double-stranded RNAs, un-capped RNA molecules, and/or single-stranded run-on mRNAs.

主权项

1. A method for generating a reprogrammed cell colony comprising reprogrammed cells that exhibit a second differentiated state, comprising:
introducing into human or mammalian cells that exhibit a first differentiated state a purified RNA preparation comprising in vitro-synthesized modified mRNA molecules that: (i) contain at least one pseudouridine that is not further modified and/or 1-methylpseudouridine in place of at least one unmodified uridine nucleoside, (ii) exhibit a 5′ cap, (iii) exhibit a 3′ poly(A) tail comprising 50-200 or greater than 150 nucleotides, and (iv) encode at least one reprogramming factor, wherein said purified RNA preparation is free of an amount of RNA contaminant molecules, including double-stranded RNA molecules, that would activate an immune response in said cells sufficient to prevent survival of said cells for at least 20 days in culture and the generation of a reprogrammed cell colony; and repeating said introducing on multiple days, under conditions such that at least one reprogrammed cell colony is generated that comprises reprogrammed cells that exhibit a second differentiated state.