Bioactive conjugates for oligonucleotide delivery
Provided herein are self-delivering oligonucleotides that are characterized by efficient RISC entry, minimum immune response and off-target effects, efficient cellular uptake without formulation, and efficient and specific tissue distribution.
1. A compound of formula (I):
wherein:
O is a double-stranded nucleic acid comprising a first oligonucleotide and a second oligonucleotide, wherein:
(1) the first oligonucleotide comprises at least 16 contiguous nucleotides, a 5′ end, a 3′ end and has complementarity to a target;
(2) the second oligonucleotide comprises at least 15 contiguous nucleotides, a 5′ end, a 3′ end, and has homology with a target; and
(3) a portion of the first oligonucleotide is complementary to a portion of the second oligonucleotide;
L is a divalent or trivalent linker;
X c is a hydrophobic moiety; and
Z c is selected from the group consisting of:
wherein X is O, S or BH 3 .
2. The compound of claim 1 , wherein L comprises an ethylene glycol chain, an alkyl chain, a peptide, RNA, DNA, a phosphodiester, a phosphorothioate, a phosphoramidate, an amide, a carbamate, or a combination thereof; and wherein L is attached to O via the second oligonucleotide.
3. The compound of claim 1 , wherein X c is selected from the group consisting of fatty acids, steroids, secosteroids, lipids, gangliosides and nucleoside analogs, and endocannabinoids.
4. The compound of claim 1 , wherein X c is a saturated or unsaturated moiety having fewer than three double bonds, or
wherein X c is a polyunsaturated moiety having three or more double bonds.
5. The compound of claim 1 , wherein:
(1) the first oligonucleotide comprises alternating 2′-methoxy-ribonucleotides and 2′-fluoro-ribonucleotides, wherein each nucleotide is a 2′-methoxy-ribonucleotide or a 2′-fluoro-ribonucleotide; and the nucleotides at positions 2 and 14 from the 5′ end of the first oligonucleotide are not 2′-methoxy-ribonucleotides;
(2) the second oligonucleotide comprises alternating 2′-methoxy-ribonucleotides and 2′-fluoro-ribonucleotides, wherein each nucleotide is a 2′-methoxy-ribonucleotide or a 2′-fluoro-ribonucleotide; and the nucleotides at positions 2 and 14 from the 5′ end of the second oligonucleotide are 2′-methoxy-ribonucleotides;
(3) the nucleotides of the first oligonucleotide are connected to adjacent nucleotides via phosphodiester or phosphorothioate linkages, wherein the nucleotides at positions 1-6 from the 3′ end, or positions 1-7 from the 3′ end are connected to adjacent nucleotides via phosphorothioate linkages; and
(4) the nucleotides of the second oligonucleotide are connected to adjacent nucleotides via phosphodiester or phosphorothioate linkages, wherein the nucleotides at positions 1 and 2 from the 3′ end are connected to adjacent nucleotides via phosphorothioate linkages.
6. The compound of claim 1 , wherein the first oligonucleotide comprises a moiety X at the 5′ end, wherein X is selected from the group consisting of:
7. The compound of claim 1 , wherein the first oligonucleotide has the structure of formula (Ia):
wherein:
X is selected from the group consisting of:
A, for each occurrence, independently is a 2′-methoxy-ribonucleotide;
B, for each occurrence, independently is a 2′-fluoro-ribonucleotide;
K, for each occurrence independently is a phosphodiester or phosphorothioate linker;
S is a phosphorothioate linker;
R is hydrogen, phosphate, vinylphosphonate, or a capping group;
j is 4, 5, 6 or 7;
r is 2 or 3; and
t is 0 or 1.
8. The compound of claim 1 , wherein the first oligonucleotide has the structure of formula (II a ):
wherein:
C-L is:
A, for each occurrence, independently is a 2′-methoxy-ribonucleotide;
B, for each occurrence, independently is a 2′-fluoro-ribonucleotide;
S is a phosphorothioate linker;
P is a phosphodiester linker;
R′ is hydrogen, phosphate, vinylphosphonate, or a capping group;
m′ is 0 or 1;
n′ is 4, 5 or 6;
q′ is 0 or 1;
r′ is 0 or 1; and
t′ is 0 or 1.
9. The compound of claim 1 , wherein the first oligonucleotide has the structure:
the second oligonucleotide has the structure:
and
the compound has the structure of formula (IIIa):
wherein each | represents a hydrogen bonding interaction wherein:
X is selected from the group consisting of:
C-L is:
A, for each occurrence, independently is a 2′-methoxy-ribonucleotide;
B, for each occurrence, independently is a 2′-fluoro-ribonucleotide;
S is a phosphorothioate linker;
P is a phosphodiester linker; R is hydrogen, phosphate, vinylphosphonate, or a capping group;
R′ is hydrogen, phosphate, vinylphosphonate, or a capping group.
10. The compound of claim 9 , wherein:
X is
L is
X c is docosahexaenoic acid (DHA);
Z c is Z c1 ;
R is hydrogen or phosphate; and
R′ is hydrogen or phosphate.
11. The compound of claim 1 , wherein the first oligonucleotide has structure:
the second oligonucleotide has the structure:
and
the compound has the structure of compound (IIIb):
wherein each | represents a hydrogen bonding interaction wherein:
C-L is:
X is selected from the group consisting of:
A, for each occurrence, independently is a 2′-methoxy-ribonucleotide;
B, for each occurrence, independently is a 2′-fluoro-ribonucleotide;
S is a phosphorothioate linker;
P is a phosphodiester linker;
R is hydrogen, phosphate, vinylphosphonate, or a capping group; and
R′ is hydrogen, phosphate, vinylphosphonate, or a capping group.
12. The compound of claim 11 , wherein:
X is
L is
X c is DHA;
Z c is Z c1 ;
R is hydrogen or phosphate; and
R′ is hydrogen or phosphate.
13. The compound of claim 1 , wherein when X c is DHA, Z c is not Z c1 .
14. The compound of claim 1 , wherein when Z c is Z c1 , X c is not DHA.
15. A compound of formula (I):
wherein:
O is a double-stranded nucleic acid comprising a first oligonucleotide and a second oligonucleotide, wherein:
(1) the first oligonucleotide comprises at least 16 contiguous nucleotides, a 5′ end, a 3′ end and has complementarity to a target;
(2) the second oligonucleotide comprises at least 15 contiguous nucleotides, a 5′ end, a 3′ end, and has homology with a target; and
(3) a portion of the first oligonucleotide is complementary to a portion of the second oligonucleotide;
wherein the first oligonucleotide has structure:
the second oligonucleotide has the structure:
and
the compound has the structure of compound (IIIb):
wherein each | represents a hydrogen bonding interaction wherein:
C-L is:
X c is DHA;
Z c is Z c1 ;
A, for each occurrence, independently is a 2′-methoxy-ribonucleotide;
B, for each occurrence, independently is a 2′-fluoro-ribonucleotide;
S is a phosphorothioate linker;
P is a phosphodiester linker;
X is
L is
R is hydrogen or phosphate; and
R′ is hydrogen or phosphate.
16. A compound of formula (I):
wherein:
O is a double-stranded nucleic acid comprising a first oligonucleotide and a second oligonucleotide, wherein:
(1) the first oligonucleotide comprises at least 16 contiguous nucleotides, a 5′ end, a 3′ end and has complementarity to a target;
(2) the second oligonucleotide comprises at least 15 contiguous nucleotides, a 5′ end, a 3′ end, and has homology with a target; and
(3) a portion of the first oligonucleotide is complementary to a portion of the second oligonucleotide;
wherein the first oligonucleotide has structure:
the second oligonucleotide has the structure:
and
the compound has the structure of compound (IIIb):
wherein each | represents a hydrogen bonding interaction wherein:
C-L is:
X c is eicosapentanoic acid (EPA);
Z c a phosphodiester or phosphodiester derivative, or is absent;
A, for each occurrence, independently is a 2′-methoxy-ribonucleotide;
B, for each occurrence, independently is a 2′-fluoro-ribonucleotide;
S is a phosphorothioate linker;
P is a phosphodiester linker;
X is
L is
R is hydrogen or phosphate; and
R′ is hydrogen or phosphate.
17. A compound of formula (I):
wherein:
O is a double-stranded nucleic acid comprising a first oligonucleotide and a second oligonucleotide, wherein:
(1) the first oligonucleotide comprises at least 16 contiguous nucleotides, a 5′ end, a 3′ end and has complementarity to a target;
(2) the second oligonucleotide comprises at least 15 contiguous nucleotides, a 5′ end, a 3′ end, and has homology with a target; and
(3) a portion of the first oligonucleotide is complementary to a portion of the second oligonucleotide;
wherein the first oligonucleotide has structure:
the second oligonucleotide has the structure:
and
the compound has the structure of compound (IIIb):
wherein each | represents a hydrogen bonding interaction wherein:
C-L is:
X c is Docosanoic Acid (DCA);
Z c a phosphodiester or phosphodiester derivative, or is absent;
A, for each occurrence, independently is a 2′-methoxy-ribonucleotide;
B, for each occurrence, independently is a 2′-fluoro-ribonucleotide;
S is a phosphorothioate linker;
P is a phosphodiester linker;
X is
L is
R is hydrogen or phosphate; and
R′ is hydrogen or phosphate.
18. The compound of claim 1 , wherein X c is selected from the group consisting of DHA, EPA, DCA, and calciferol.