Method and medicament for inhibiting the expression of a given gene
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 .- 3 . (canceled)
4 . A method for stabilizing a double stranded RNA (dsRNA) consisting of two separate oligoribonucleotide strands for use in reducing expression of a target gene in a cell comprising:
a. providing a first oligoribonucleotide strand, wherein said first oligoribonucleotide strand is between 15-25 nucleotides in length and comprises a nucleic acid sequence fully complementary to a portion of an mRNA transcript of said target gene,
b. providing a second oligoribonucleotide strand which comprises a nucleic acid sequence complementary to said first strand,
c. hybridizing said first strand to said second strand under conditions that allow the formation of a dsRNA, and
d. linking said first and second strands to reduce dissociation compared to non-linked RNA strands, wherein said linking is in addition to the cohesion caused by the nucleotide pairs in the double stranded region.
5 . The method of claim 4 , wherein said chemical linkage is formed by a covalent bond or hydrogen bound.
6 . The method of claim 4 , wherein at least one of said oligoribonucleotide strands comprises at least one chemically modified nucleotide.
7 . The method of claim 5 , wherein at least one of said oligoribonucleotide strands comprises at least one chemically modified nucleotide.
8 . The method of claim 6 , wherein said chemically modified nucleotide is a 2′-modified nucleotide.
9 . The dsRNA of claim 8 , wherein said 2′-modified nucleotide is a 2′-methyl substituted nucleotide.
10 . The dsRNA of claim 8 , wherein said 2′-modified nucleotide is a 2′-amino substituted nucleotide.
11 . The dsRNA of claim 6 , wherein said chemically modified nucleotide is a blocked nucleotide.
12 . The method of claim 4 , wherein said first oligoribonucleotide strand is between 15-21 nucleotides in length.
13 . The method of claim 4 , wherein said first oligoribonucleotide strand is 21 nucleotides long.
14 . The method of claim 4 , wherein said chemical linkage is formed by a linkage selected from the group consisting of poly(oxyphosphinicooxy-1,3-propandiol), polyethylene glycol, methylene blue, bis(2-chloroethyl)amine, and N-acetyl-N′-(p-glyoxyl-benzoyl)cystamine.
15 . The method of claim 4 , wherein said chemical linkage is a C18 linker.
16 . The method of claim 4 , wherein the target gene is a mammalian gene or a viral gene.
17 . The method of claim 4 , wherein the target gene is selected from the group consisting of an oncogene, a cytokine gene, an Id protein gene, a developmental gene, a PKR gene and a prion gene.
18 . The method of claim 4 , wherein said cell is a mammalian cell.