Method and medicament for inhibiting the expression of a given gene
View Patent ↗The invention relates to an isolated RNA that mediates RNA interference of an mRNA to which it corresponds and a method of mediating RNA interference of mRNA of a gene in a cell or organism using the isolated RNA.
1. A method for inhibiting expression of a target gene, comprising:
a) introducing into a mammalian cell an isolated double stranded RNA (dsRNA) comprising two complementary oligoribonucleotide strands, wherein a first strand of the dsRNA is complementary to an RNA transcript of at least part of the target gene and a second strand of the dsRNA is complementary to the first strand, wherein the dsRNA is 15 to 21 base pairs in length, and wherein dsRNA is introduced into the mammalian cell at a concentration that is lower by at least one order of magnitude than a concentration that would be required for the first strand of the dsRNA alone as single-stranded oligoribonucleotide to inhibit expression of the target gene; and
b) maintaining the cell produced in step a) with the concentration of dsRNA for a time sufficient to obtain degradation of an RNA transcript of the target gene, thereby inhibiting the expression of the target gene at the concentration of the dsRNA.
2. The method of claim 1 , wherein at least one of said strands comprises at least one chemically modified nucleotide.
3. The method of claim 2 , wherein said chemically modified nucleotide is a 2′-modified nucleotide.
4. The method of claim 2 , wherein said 2′-modified nucleotide is a 2′-methyl substituted nucleotide.
5. The method of claim 2 , wherein said 2′-modified nucleotide is a 2′-amino substituted nucleotide.
6. The method of claim 2 , wherein said chemically modified nucleotide is a locked nucleotide.
7. The method of claim 1 , wherein the dsRNA is enclosed by a micellar structure.
8. The method of claim 7 , wherein the micellar structure comprises a liposome.
9. The method of claim 1 , wherein the two complementary strands are fully complementary to each other.
10. The method of claim 1 , wherein said dsRNA specifically inhibits the expression of the target gene using dsRNA-mediated interference.
11. The method of claim 1 , wherein one of the two complementary oligoribonucleotide strands is a sense strand, and wherein the sense strand comprises a 2′-methyl substituted nucleotide.
12. The method of claim 1 , wherein one of the two complementary oligoribonucleotide strands is a sense strand, and wherein the sense strand comprises a plurality of 2′-methoxynucleotides.
13. The method of claim 1 , wherein the RNA transcript comprises a third strand, and wherein the dsRNA, when introduced into the presence of the third strand, reduces an amount of the third strand, the dsRNA thereby specifically inhibiting expression of the target gene.
14. The method of claim 1 , wherein introducing the dsRNA into the mammalian cell comprises introducing a pharmaceutical composition comprising the dsRNA into the mammalian cell.
15. A method for inhibiting expression of a target gene, comprising:
a) introducing into a mammalian cell a pharmaceutical composition comprising an isolated double stranded RNA (dsRNA) comprising two complementary oligoribonucleotide strands, wherein a first strand of the dsRNA is complementary to an RNA transcript of at least part of the target gene and a second strand of the dsRNA is complementary to the first strand, wherein the dsRNA is 15 to 21 base pairs in length, and wherein dsRNA is introduced into the mammalian cell at a concentration that is lower by at least one order of magnitude than a concentration that would be required for the first strand of the dsRNA alone as single-stranded oligoribonucleotide to inhibit expression of the target gene; and
b) maintaining the cell produced in step a) with the concentration of dsRNA for a time sufficient to obtain degradation of an RNA transcript of the target gene, thereby inhibiting the expression of the target gene at the concentration of the dsRNA.
16. The method of claim 15 , wherein the pharmaceutical composition introduced comprises a liposome enclosing the dsRNA.
17. The method of claim 2 , wherein the at least one chemically modified nucleotide is at the 3′ terminus, the 5′ terminus or combinations thereof, of at least one of the oligoribonucleotide strands.
18. The method of claim 17 , wherein the at least one chemically modified nucleotide is at the 3′ terminus.
19. The method of claim 17 or 18 , wherein the at least one chemically modified nucleotide is a 2′-methyl substituted nucleotide.
20. The method of claim 1 , wherein the mammalian cell is a primate cell.
21. The method of claim 15 , wherein the dsRNA comprises at least one chemically modified nucleotide at the 3′ terminus, the 5′ terminus or combinations thereof, of at least one of the oligoribonucleotide strands.
22. The method of claim 21 , wherein the at least one chemically modified nucleotide is at the 3′ terminus.
23. The method of claim 21 or 22 , wherein the at least one chemically modified nucleotide is a 2′-methyl substituted nucleotide.
24. The method of claim 15 , wherein the mammalian cell is a primate cell.