Method of minimizing off-target effects of siRNA molecules
The invention relates to a method of minimizing off-target effects of siRNA, comprising preparing double-stranded RNA (dsRNA) having a sense strand that is homologous to a sequence of a target gene and an anti-sense strand that is complementary to said sense strand, and having at least one pyrimidine replaced by a 5′-methyl-pyrimidine, and contacting said dsRNA with a cell capable of expressing said target gene.
1 . A method of minimizing off-target effects of siRNA, comprising preparing double-stranded RNA (dsRNA) having a sense strand that is homologous to a sequence of a target gene and an anti-sense strand that is complementary to said sense strand, and having at least one pyrimidine replaced by a 5′-methyl-pyrimidine, and contacting said dsRNA with a cell capable of expressing said target gene.
2 . The method of claim 1 , wherein the 5′-methyl-pyrimidine is ribothymidine.
3 . The method of claim 2 , wherein at least three of the uridines of the siRNA sequence are replaced by ribothymidines.
4 . The method of claim 2 , wherein at least three of the uridines of the sense strand of the siRNA sequence are replaced by ribothymidines.
5 . The method of claim 2 , wherein at least three of the uridines of the antisense strand of the siRNA sequence are replaced by ribothymidines.
6 . The method of claim 2 , wherein all of the uridines in the siRNA sequence are replaced by ribothymidines.
7 . The method of claim 2 , wherein the siRNA molecule has a 3′ overhang.
8 . The method of claim 2 , wherein the siRNA molecule is blunt ended.
9 . A method of increasing stability of siRNA, comprising preparing double-stranded RNA (dsRNA) having a sense strand that is homologous to a sequence of a target gene and an anti-sense strand that is complementary to said sense strand, and having at least one pyrimidine replaced by a 5′-methyl-pyrimidine, and contacting said dsRNA with a biological sample.
10 . The method of claim 9 , wherein the biological sample is blood serum or plasma.
11 . The method of claim 9 , wherein at least three of the uridines of the sense strand of the siRNA sequence are replaced by ribothymidines.
12 . The method of claim 9 , wherein at least three of the uridines of the antisense strand of the siRNA sequence are replaced by ribothymidines.
13 . The method of claim 9 , wherein at least three of the uridines in the siRNA sequence are replaced by ribothymidines.