Optimized messenger RNA
The present invention is directed to a synthetic nucleic acid sequence which encodes a protein wherein at least one non-common codon or less-common codon is replaced by a common codon. The synthetic nucleic acid sequence can include a continuous stretch of at least 90 codons all of which are common codons.
1. A synthetic nucleic acid sequence which encodes a human Factor VIII or, wherein at least one non-common codon or less-common codon has been replaced by a common codon encoding the same amino acid residue as the non-common or less-common codon and wherein the synthetic nucleic acid has a continuous stretch of at least 150 codons all of which are common codons, and wherein by a common codon is meant the most common codon encoding each particular amino acid residue in highly expressed human genes as shown in FIGS. 14A and 14B .
2. The synthetic nucleic acid sequence of claim 1 where the factor VIII protein has one or more of the following characteristics:
a) the B domain is deleted (beta domain deleted (BDD) factor VIII);
b) it has a recognition site for an intracellular protease of the PACE/furin class; or
c) it is expressed in a non-transformed cell.
3. The synthetic nucleic acid sequence of claim 1 , wherein the number of non-common or less-common codons replaced or remaining is between one and 15.
4. The synthetic nucleic acid sequence of claim 1 , wherein all non-common and less-common codons are replaced with common codons.
5. A synthetic nucleic acid sequence which encodes a human Factor VIII, wherein at least one non-common codon or less-common codon has been replaced by a common codon encoding the same amino acid residue as the non-common or less-common codon and wherein the synthetic nucleic acid has a continuous stretch of common codons which comprise at least 60% of the codons of the synthetic nucleic acid sequence, and wherein by a common codon is meant the most common codon encoding each particular amino acid residue in highly expressed human genes as shown in FIGS. 14A and 14B .
6. The synthetic nucleic acid sequence of claim 5 where the factor VIII protein has one or more of the following characteristics:
a) the B domain is deleted (BDD factor VIII);
b) it has a recognition site for an intracellular protease of the PACE/furin class; or
c) it is expressed in a non-transformed cell.
7. The synthetic nucleic acid sequence of claim 5 , wherein the number of non-common or less-common codons replaced or remaining is between one and 15.
8. The synthetic nucleic acid sequence of claim 5 , wherein all non-common and less-common codons are replaced with common codons.
9. A synthetic nucleic acid sequence which encodes a human Factor VIII, wherein at least one non-common codon or less-common codon has been replaced by a common codon encoding the same amino acid residue as the non-common or less-common codon and wherein at least 98% or more of the codons in the sequence encoding the Factor VIII are common codons and the Factor VIII is at least 90 amino acid residues in length, and wherein by a common codon is meant the most common codon encoding each particular amino acid residue in highly expressed human genes as shown in FIGS. 14A and 14B .
10. The synthetic nucleic acid sequence of claim 9 where the factor VIII protein has one or more of the following characteristics:
a) the B domain is deleted (BDD factor VIII;
b) it has a recognition site for an intracellular protease of the PACE/furin class; and
c) it is expressed in a non-transformed cell.
11. The synthetic nucleic acid sequence of claim 9 , wherein the number of non-common or less-common codons replaced or remaining is between one and 15.
12. The synthetic nucleic acid sequence of claim 9 , wherein the number of non-common or less-common codons replaced or remaining, taken together, are equal or less than 2% of the codons in the synthetic nucleic acid sequence.
13. The synthetic nucleic acid sequence of claim 9 , wherein all non-common and less-common codons are replaced with common codons.
14. The synthetic nucleic acid sequence of claim 9 , wherein at least 99% of the codons in the synthetic nucleic acid sequence are common codons.
15. The synthetic nucleic acid sequence of claim 9 , wherein all of the codons are replaced with common codons.
16. A synthetic nucleic acid sequence which encodes human Factor IX, wherein at least one non-common codon or less-common codon has been replaced by a common codon encoding the same amino acid residue as the non-common or less-common codon and wherein the synthetic nucleic acid has a continuous stretch of at least 150 codons all of which are common codons, and wherein by a common codon is meant the most common codon encoding each particular amino acid residue in highly expressed human genes as shown in FIGS. 14A and 14B .
17. The synthetic nucleic acid sequence of claim 16 , wherein the Factor IX protein has one or more of the following characteristics:
a) it has a PACE/furin site at a pro-peptide mature protein junction; and
b) is expressed in a non-transformed cell.
18. The synthetic nucleic acid sequence of claim 16 , wherein the number of non-common or less-common codons replaced or remaining is between one and 15.
19. A synthetic nucleic acid sequence which encodes human Factor IX, wherein at least one non-common codon or less-common codon has been replaced by a common codon encoding the same amino acid residue as the non-common or less-common codon and wherein the synthetic nucleic acid has a continuous stretch of common codons which comprise at least 60% of the codons of the synthetic nucleic acid sequence, and wherein by a common codon is meant the most common codon encoding each particular amino acid residue in highly expressed human genes as shown in FIGS. 14A and 14B .
20. The synthetic nucleic acid sequence of claim 19 , wherein the number of non-common or less-common codons replaced or remaining is between one and 15.
21. The synthetic nucleic acid sequence of claim 19 , wherein the factor IX protein has one or more of the following characteristics:
a) it has a PACE/furin site at a pro-peptide mature protein junction; and
b) is expressed in a non-transformed cell.
22. A synthetic nucleic acid sequence which encodes human Factor IX, wherein at least one non-common codon or less-common codon has been replaced by a common codon encoding the same amino acid residue as the non-common or less-common codon and wherein at least 98% or more of the codons in the sequence encoding the Factor IX are common codons and the Factor IX is at least 90 amino acid residues in length, and wherein by a common codon is meant the most common codon encoding each particular amino acid residue in highly expressed human genes as shown in FIGS. 14A and 14B .
23. The synthetic nucleic acid sequence of claim 22 , wherein the factor IX protein has one or more of the following characteristics:
a) it has a PACE/furin site at a pro-peptide mature protein junction; and
b) is expressed in a non-transformed cell.
24. The synthetic nucleic acid sequence of claim 22 , wherein the number of non-common or less-common codons replaced or remaining is between one and 15.
25. The synthetic nucleic acid sequence of claim 22 , wherein the number of non-common or less-common codons replaced or remaining, taken together, are equal or less then 2% of the codons in the synthetic nucleic acid sequence.
26. The synthetic nucleic acid sequence of claim 22 , wherein all non-common and less-common codons are replaced with common codons.
27. The synthetic nucleic acid sequence of claim 22 , wherein at least 99% of the codons in the synthetic nucleic acid sequence are common codons.
28. The synthetic nucleic acid sequence of claim 22 , wherein all of the codons are replaced with common codons.
29. A vector comprising the synthetic nucleic acid sequence of claim 1 , 5 , 9 , 16 , 19 or 22 .
30. A cell comprising the nucleic acid sequence of claim 1 , 5 , 9 , 16 , 19 or 22 .