Method for reducing immunogenicity of RNA
The present invention relates to RNA therapy and, in particular, decreasing immunogenicity of RNA. Specifically, the present invention provides methods for decreasing immunogenicity of RNA, said methods comprising modifying the nucleotide sequence of the RNA by reducing the uridine (U) content, wherein said reduction of the U content comprises an elimination of U nucleosides from the nucleotide sequence of the RNA and/or a substitution of U nucleosides by nucleosides other than U in the nucleotide sequence of the RNA. Using RNA having decreased immunogenicity allows administration of RNA as a drug to a subject, e.g. in order to obtain expression of a pharmaceutically active peptide or protein, without eliciting an immune response which would interfere with therapeutic effectiveness of the RNA or induce adverse effects in the subject.
1. A modified RNA comprising an adenosine-rich first nucleotide sequence having an adenosine content of more than 25% and encoding at least one peptide or protein; wherein the peptide or protein encoded by the first nucleotide sequence is also encoded by a second nucleotide sequence of a second, naturally occurring RNA; and the first nucleotide sequence has a higher A content and a lower U content compared to the second nucleotide sequence; wherein the first nucleotide sequence comprises nucleotide residues selected from adenosine (A), guanosine (G), and cytidine (C) at positions corresponding to uracil (U) nucleotide residue positions in the second nucleotide sequence; thereby rendering the modified RNA at least 5% less immunogenic than the second RNA.
2. The modified RNA of claim 1 , wherein the peptide or protein encoded by the first and second nucleotide sequences is pharmaceutically active or antigenic.
3. The modified RNA of claim 1 , wherein the first nucleotide sequence comprises at least 10% fewer U nucleotide residues compared to the second nucleotide sequence.
4. The modified RNA of claim 1 , wherein the first nucleotide sequence comprises at least 20% fewer U nucleotide residues compared to the second nucleotide sequence.
5. The modified RNA of claim 1 , wherein the first nucleotide sequence comprises at least 30% fewer U nucleotide residues compared to the second nucleotide sequence.
6. The modified RNA of claim 1 , wherein the first nucleotide sequence comprises fewer U nucleotide residues compared to the second nucleotide sequence in 5′ untranslated regions thereof.
7. The modified RNA of claim 6 , wherein the first nucleotide sequence comprises fewer U nucleotide residues compared to the second nucleotide sequence in coding regions thereof.
8. The modified RNA of claim 7 , wherein the first nucleotide sequence comprises fewer U nucleotide residues compared to the second nucleotide sequence in 3′ untranslated regions thereof.
9. The modified RNA of claim 1 , wherein the first nucleotide sequence comprises fewer U nucleotide residues compared to the second nucleotide sequence in coding regions thereof.
10. The modified RNA of claim 9 , wherein the first nucleotide sequence comprises fewer U nucleotide residues compared to the second nucleotide sequence in 3′ untranslated regions thereof.
11. The modified RNA of claim 1 , wherein the first nucleotide sequence comprises fewer U nucleotide residues compared to the second nucleotide sequence in 3′ untranslated regions thereof.
12. The modified RNA of claim 6 , wherein the first nucleotide sequence comprises fewer U nucleotide residues compared to the second nucleotide sequence in 3′ untranslated regions thereof.
13. The modified RNA of claim 1 , wherein the first nucleotide sequence comprises A nucleotide residues at positions corresponding to U nucleotide residue positions in the second nucleotide sequence.
14. The modified RNA of claim 1 , wherein the first nucleotide sequence comprises codons having more A nucleotide residues and fewer U nucleotide residues compared to the second nucleotide sequence.
15. A pharmaceutical composition comprising the modified RNA of claim 1 and one or more pharmaceutically acceptable carriers, diluents and/or excipients.
16. The composition of claim 15 , wherein the modified RNA is complexed with liposomes.
17. A method of increasing protein production of a peptide or protein comprising transfecting cells with the modified RNA of claim 1 , wherein the cells transfected with the modified RNA express more of the peptide or protein compared to cells transfected with the second RNA that encodes the same peptide or protein.
18. The method of claim 17 , wherein the cells are transfected in vitro.
19. The method of claim 17 , wherein the cells are transfected by intravenously administering the RNA to a subject comprising the cells.
20. The method of claim 17 , wherein the modified RNA is complexed with liposomes.