RNA interference mediated inhibition of gene expression using chemically modified short interfering nucleic acid (siNA)
First Claim
1. A method of synthesizing a double stranded nucleic acid molecule consisting of a sense strand and an antisense strand that mediates cleavage of a target RNA encoded by a mammalian gene, wherein:
- (a) each strand of said double stranded nucleic acid molecule is 19 to 29 nucleotides in length;
(b) at least 19 nucleotides of the sense strand are complementary to the antisense strand (c) the antisense strand of said double stranded nucleic acid molecule has complementarity to said target RNA encoded by a mammalian gene;
(d) at least 35% of the internal nucleotides of each strand of said double stranded nucleic acid molecule are modified nucleosides having a sugar modification; and
(e) at least two of said sugar modifications are different from each other;
the method comprising (i) synthesizing two strands of nucleic acid molecules that have the structural features described in parts (a)-(e), and (ii) annealing the two complementary strands under conditions suitable to obtain a double-stranded nucleic acid molecule.
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Abstract
The present invention concerns methods and reagents useful in modulating gene expression in a variety of applications, including use in therapeutic, diagnostic, target validation, and genomic discovery applications. Specifically, the invention relates to synthetic chemically modified small nucleic acid molecules, such as short interfering nucleic acid (siNA), short interfering RNA (siRNA), double-stranded RNA (dsRNA), micro-RNA (miRNA), and short hairpin RNA (shRNA) molecules capable of mediating RNA interference (RNAi) against target nucleic acid sequences. The small nucleic acid molecules are useful in the treatment of any disease or condition that responds to modulation of gene expression or activity in a cell, tissue, or organism.
82 Citations
25 Claims
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1. A method of synthesizing a double stranded nucleic acid molecule consisting of a sense strand and an antisense strand that mediates cleavage of a target RNA encoded by a mammalian gene, wherein:
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(a) each strand of said double stranded nucleic acid molecule is 19 to 29 nucleotides in length;
(b) at least 19 nucleotides of the sense strand are complementary to the antisense strand (c) the antisense strand of said double stranded nucleic acid molecule has complementarity to said target RNA encoded by a mammalian gene;
(d) at least 35% of the internal nucleotides of each strand of said double stranded nucleic acid molecule are modified nucleosides having a sugar modification; and
(e) at least two of said sugar modifications are different from each other;
the method comprising (i) synthesizing two strands of nucleic acid molecules that have the structural features described in parts (a)-(e), and (ii) annealing the two complementary strands under conditions suitable to obtain a double-stranded nucleic acid molecule. - View Dependent Claims (2, 3, 4, 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, 15, 16, 17, 18, 19, 20, 21, 22, 23, 24, 25)
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Specification