Extended dicer substrate agents and methods for the specific inhibition of gene expression

An isolated double stranded nucleic acid (dsNA) comprising a first oligonucleotide strand having a 5′ terminus and a 3′ terminus and a second oligonucleotide strand having a 5′ terminus and a 3′ terminus, wherein:

• said first strand is 31 nucleotide residues in length and said second strand is 31-35 nucleotide residues in length, wherein starting from the first nucleotide (position 1) at the 5′ terminus of the first strand, positions 1 to 23 of said first strand are ribonucleotides that base pair with ribonucleotides of said second strand to form a duplex;
• each of positions 24 to 31 of said first strand is a deoxyribonucleotide that base pairs with a deoxyribonucleotide of said second strand to form a duplex;
• the 3′ terminus of said first strand and the 5′ terminus of said second strand form a duplexed blunt end;
• said second strand is sufficiently complementary to a target RNA along at least 19 ribonucleotides of said second strand length to reduce target gene expression when said double stranded nucleic acid is introduced into a mammalian cell.

An isolated double stranded nucleic acid (dsNA) comprising a first oligonucleotide strand having a 5′ terminus and a 3′ terminus and a second oligonucleotide strand having a 5′ terminus and a 3′ terminus, wherein:

• said first strand is 33 nucleotide residues in length and said second strand is 33-37 nucleotide residues in length, wherein starting from the first nucleotide (position 1) at the 5′ terminus of the first strand, positions 1 to 23 of said first strand are ribonucleotides that base pair with ribonucleotides of said second strand to form a duplex;
• each of positions 24 to 33 of said first strand is a deoxyribonucleotide that base pairs with a deoxyribonucleotide of said second strand to form a duplex;
• the 3′ terminus of said first strand and the 5′ terminus of said second strand form a duplexed blunt end;
• said second strand is sufficiently complementary to a target RNA along at least 19 ribonucleotides of said second strand length to reduce target gene expression when said double stranded nucleic acid is introduced into a mammalian cell.

An isolated double stranded nucleic acid (dsNA) comprising a first oligonucleotide strand having a 5′ terminus and a 3′ terminus and a second oligonucleotide strand having a 5′ terminus and a 3′ terminus, wherein:

• said first strand is 35 nucleotide residues in length and said second strand is 35-39 nucleotide residues in length, wherein starting from the first nucleotide (position 1) at the 5′ terminus of the first strand, positions 1 to 23 of said first strand are ribonucleotides that base pair with ribonucleotides of said second strand to form a duplex;
• each of positions 24 to 35 of said first strand is a deoxyribonucleotide that base pairs with a deoxyribonucleotide of said second strand to form a duplex;
• the 3′ terminus of said first strand and the 5′ terminus of said second strand form a duplexed blunt end;
• said second strand is sufficiently complementary to a target RNA along at least 19 ribonucleotides of said second strand length to reduce target gene expression when said double stranded nucleic acid is introduced into a mammalian cell.

An isolated double stranded nucleic acid (dsNA) comprising a first oligonucleotide strand having a 5′ terminus and a 3′ terminus and a second oligonucleotide strand having a 5′ terminus and a 3′ terminus, wherein:

• said first strand is 37 nucleotide residues in length and said second strand is 37-41 nucleotide residues in length, wherein starting from the first nucleotide (position 1) at the 5′ terminus of the first strand, positions 1 to 23 of said first strand are ribonucleotides that base pair with ribonucleotides of said second strand to form a duplex;
• each of positions 24 to 37 of said first strand is a deoxyribonucleotide that base pairs with a deoxyribonucleotide of said second strand to form a duplex;
• the 3′ terminus of said first strand and the 5′ terminus of said second strand form a dublexed blunt end;
• said second strand is sufficiently complementary to a target RNA along at least 19 ribonucleotides of said second strand length to reduce target gene expression when said double stranded nucleic acid is introduced into a mammalian cell.

An isolated double stranded nucleic acid (dsNA) comprising a first oligonucleotide strand having a 5′ terminus and a 3′ terminus and a second oligonucleotide strand having a 5′ terminus and a 3′ terminus, wherein:

• said first strand is 31 to 49 nucleotide residues in length, wherein starting from the first nucleotide (position 1) at the 5′ terminus of the first strand, positions 1 to 23 of said first strand are ribonucleotides;
• said second strand is 31 to 53 nucleotide residues in length and comprises 23 consecutive ribonucleotides that base pair with the ribonucleotides of positions 1 to 23 of said first strand to form a duplex;
• the 5′ terminus of said first strand and the 3′ terminus of said second strand form a structure selected from the group consisting of a blunt end and a 1-4 nucleotide 3′ overhang,
• the 3′ terminus of said first strand and the 5′ terminus of said second strand form a blunt end;
• at least one of positions 24 to the 3′ terminal nucleotide residue of said first strand is a phosphorothioate-modified nucleotide (PS-NA) That base pairs with a deoxyribonucleotide of said second strand;
• and
• said second strand is sufficiently complementary to a target RNA along at least 19 ribonucleotides of said second strand length to reduce target gene expression when said double stranded nucleic acid is introduced into a mammalian cell.

An isolated double stranded nucleic acid (dsNA) comprising a first oligonucleotide strand having a 5′ terminus and a 3′ terminus and a second oligonucleotide strand having a 5′ terminus and a 3′ terminus, wherein:

• said first strand is 31 to 49 nucleotide residues in length, wherein starting from the first nucleotide (position 1) at the 5′ terminus of the first strand, positions 1 to 23 of said first strand are ribonucleotides;
• said second strand is 31 to 53 nucleotide residues in length and comprises 23 consecutive ribonucleotides that base pair with the ribonucleotides of positions 1 to 23 of said first strand to form a duplex;
• the 5′ terminus of said first strand and the 3′ terminus of said second strand form a structure selected from the group consisting of a blunt end and a 1-4 nucleotide 3′ overhang,
• the 3′ terminus of said first strand and the 5′ terminus of said second strand form a blunt end;
• at least one nucleotide of said second strand base pairs with a deoxyribonucleotide of positions 24 to the 3′ terminal nucleotide residue of said first strand and is a phosphorothioate-modified nucleotide (PS-NA);
• and
• said second strand is sufficiently complementary to a target RNA along at least 19 ribonucleotides of said second strand length to reduce target gene expression when said double stranded nucleic acid is introduced into a mammalian cell.

An isolated double stranded nucleic acid (dsNA) comprising a first oligonucleotide strand having a 5′ terminus and a 3′ terminus and a second oligonucleotide strand having a 5′ terminus and a 3′ terminus, wherein:

• said first strand is 31 to 49 nucleotide residues in length, wherein starting from the first nucleotide (position 1) at the 5′ terminus of the first strand, positions 1 to 23 of said first strand are ribonucleotides;
• said second strand is 31 to 53 nucleotide residues in length and comprises 23 consecutive ribonucleotides that base pair with the ribonucleotides of positions 1 to 23 of said first strand to form a duplex;
• the 5′ terminus of said first strand and the 3′ terminus of said second strand form a structure selected from the group consisting of a blunt end and a 1-4 nucleotide 3′ overhang,
• the 3′ terminus of said first strand and the 5′ terminus of said second strand form a blunt end;
• at least one of positions 24 to the 3′ terminal nucleotide residue of said first strand is a deoxyribonucleotide that base pairs with a phosphorothioate-modified nucleotide (PS-NA) of said second strand;
• and
• said second strand is sufficiently complementary to a target RNA along at least 19 ribonucleotides of said second strand length to reduce target gene expression when said double stranded nucleic acid is introduced into a mammalian cell.