Oligonucleotides having A-DNA form and B-DNA form conformational geometry
First Claim
1. An oligonucleotide comprising:
- at least one continuous sequence of nucleotides having A-form conformational geometry; and
at least one continuous sequence of nucleotides having B-form conformational geometry;
wherein at least one of said nucleotides having B-form conformational geometry is an arabinonucleotide.
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Accused Products
Abstract
Modified oligonucleotides containing both A-form conformation geometry and B-from conformation geometry nucleotides are disclosed. The B-form geometry allows the oligonucleotide to serve as substrates for RNase H when bound to a target nucleic acid strand. The A-form geometry imparts properties to the oligonucleotide that modulate binding affinity and nuclease resistance. By utilizing C2′ endo sugars or O4′ endo sugars, the B-form characteristics are imparted to a portion of the oligonucleotide. The A-form characteristics are imparted via use of either 2′-O-modified nucleotides that have 3′ endo geometries or use of end caps having particular nuclease stability or by use of both of these in conjunction with each other.
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Citations
38 Claims
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1. An oligonucleotide comprising:
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at least one continuous sequence of nucleotides having A-form conformational geometry; and
at least one continuous sequence of nucleotides having B-form conformational geometry;
wherein at least one of said nucleotides having B-form conformational geometry is an arabinonucleotide. - 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, 26, 27, 28, 29, 30, 31, 32, 33, 34, 35, 36, 37, 38)
wherein E is C1-C10 alkyl, N(Q1)(Q2) or N═
C(Q1)(Q2);
each Q1 and Q2 is, independently, H, C1-C10 alkyl, dialkylaminoalkyl, a nitrogen protecting group, a tethered or untethered conjugate group, a linker to a solid support, or Q1 and Q2, together, are joined in a nitrogen protecting group or a ring structure optionally containing at least one additional heteroatom selected from N and O;
R3 is OX, SX, or N(X)2;
each X is, independently, H, C1-C8 alkyl, C1-C8 haloalkyl, C(═
NH)N(H)Z, C(═
O)N(H)Z or OC(═
O)N(H)Z;
Z is H or C1-C8alkyl;
L1, L2 and L3 form a ring system having from about 4 to about 7 carbon atoms or having from about 3 to about 6 carbon atoms and 1 or 2 heteroatoms selected from oxygen, nitrogen and sulfur and wherein said ring system is aliphatic, unsaturated aliphatic, aromatic, or saturated or unsaturated heterocyclic;
Y is alkyl or haloalkyl having 1 to about 10 carbon atoms, alkenyl having 2 to about 10 carbon atoms, alkynyl having 2 to about 10 carbon atoms, aryl having 6 to about 14 carbon atoms, N(Q1)(Q2), O(Q1), halo, S(Q1), or CN;
each q1 is, independently, from 2 to 10;
each q2 is, independently, 0 or 1;
m is 0, 1 or 2;
p is from 0 to 10; and
q3 is from 1 to 10 with the proviso that when p is 0, q3 is greater than 1.
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8. The oligonucleotide of claim 7 wherein at least one nucleotide having A-form conformational geometry has the 2′
- -substituent group of formula I and p is 0.
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9. The oligonucleotide of claim 2 wherein at least one of said nucleotides having A-form conformational geometry is a 2′
- -F ribonucleotide, a 2′
-O—
(C1-C6 alkyl) ribonucleotide, or a 2′
-O—
(C1-C6 substituted alkyl) ribonucleotide wherein the substitution is C1-C6 ether, C1-C6 thioether, amino, amino(C1-C6 alkyl) or amino(C1-C6 alkyl)2.
- -F ribonucleotide, a 2′
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10. The oligonucleotide of claim 2 wherein at least one of said nucleotides having B-form conformational geometry is a 2′
- -CN arabinonucleotide, a 2′
-F arabinonucleotide, a 2′
-Cl arabinonucleotide, a 2′
-Br arabinonucleotide, a 2′
-N3 arabinonucleotide, a 2′
-OH arabinonucleotide, a 2′
-O—
CH3 arabinonucleotide or a 2′
-dehydro-2′
-CH3 arabinonucleotide.
- -CN arabinonucleotide, a 2′
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11. The oligonucleotide of claim 10 wherein at least one of said nucleotides having B-form conformational geometry is a 2′
- -F arabinonucleotide, a 2′
-OH arabinonucleotide or a 2′
-O—
CH3 arabinonucleotide.
- -F arabinonucleotide, a 2′
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12. The oligonucleotide of claim 11 wherein at least one of said nucleotides having B-form conformational geometry is a 2′
- -F arabinonucleotide or a 2′
-OH arabinonucleotide.
- -F arabinonucleotide or a 2′
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13. The oligonucleotide of claim 12 wherein at least one of said nucleotides having B-form conformational geometry is a 2′
- -F arabinonucleotide.
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14. The oligonucleotide of claim 2 wherein at least one of said nucleotides having B-form conformational geometry is a 2′
- -CN arabinonucleotide, a 2′
-F arabinonucleotide, a 2′
-Cl arabinonucleotide, a 2′
-Br arabinonucleotide, a 2′
-N3 arabinonucleotide, a 2′
-OH arabino-nucleotide, a 2′
-O—
CH3 arabinonucleotide or a 2′
-dehydro-2′
-CH3 arabinonucleotide; andat least one of said nucleotides having A-form conformational geometry is a 2′
-F ribonucleotide, a 2′
-O—
(C1-C6 alkyl) ribonucleotide, or a 2′
-O—
(C1-C6 substituted alkyl) ribonucleotide wherein the substitution is C1-C6 ether, C1-C6 thioether, amino, amino(C1-C6 alkyl) or amino(C1-C6 alkyl)2.
- -CN arabinonucleotide, a 2′
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15. The oligonucleotide of claim 14 wherein at least one of said nucleotides having B-form conformational geometry is a 2′
- -F arabinonucleotide or a 2′
-OH arabinonucleotide; andat least one of said nucleotides having A-form conformational geometry is a 2′
-O—
(C1-C6 substituted alkyl) ribonucleotide wherein the substitution is C1-C6 ether, C1-C6 thioether, amino, amino(C1-C6 alkyl) or amino(C1-C6 alkyl)2.
- -F arabinonucleotide or a 2′
-
16. The oligonucleotide of claim 15 wherein at least one of said nucleotides having A-form conformational geometry is a 2′
- -alkylamino substituted nucleotide.
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17. The oligonucleotide of claim 1 further comprising at least one nucleotide having A-form conformational geometry bound by an internucleotide linkage to a 5′
- position of at least one of said sequence of nucleotides having B-form conformational geometry.
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18. The oligonucleotide of claim 17 wherein said internucleotide linkage at the 5′
- end of said oligonucleotide is a 3′
-5′
phosphodiester, 2′
-5′
phosphodiester, phosphorothioate, Sp phosphorothioate, Rp phosphorothioate, phosphorodithioate, 3′
-deoxy-3′
-amino, phosphoroamidate, 3′
-methylenephosphonate, methylene(methylimino), dimethylhydrazino, amide 3, amide 4 or a boranophosphate.
- end of said oligonucleotide is a 3′
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19. The oligonucleotide of claim 18 wherein said internucleotide linkage is a 2′
- -5′
phosphodiester, 3′
-methylenephosphonate, Sp phosphorothioate or a methylene(methylimino) linkage.
- -5′
-
20. The oligonucleotide of claim 17 further comprising at least two nucleotides having A-form conformational geometry bound by an internucleotide linkage to a 5′
- position of at least one of said sequence of nucleotides having B-form conformational geometry.
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21. The oligonucleotide of claim 17 wherein at least two of said nucleotides having B-form conformational geometry are joined together by phosphate, phosphorothioate, phosphorodithioate or boranophosphate internucleotide linkages.
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22. The oligonucleotide of claim 17 wherein at least one of said nucleotides having
A-form conformational geometry is a 2′ - -fluoro nucleotide or a nucleotide having a 2′
-substituent having the formula I or II;
wherein E is C1-C10 alkyl, N(Q1)(Q2) or N═
C(Q1)(Q2);
each Q1 and Q2 is, independently, H, C1-C10 alkyl, dialkylaminoalkyl, a nitrogen protecting group, a tethered or untethered conjugate group, a linker to a solid support, or Q1 and Q2, together, are joined in a nitrogen protecting group or a ring structure optionally containing at least one additional heteroatom selected from N and O;
R3 is OX, SX, or N(X)2;
each X is, independently, H, C1-C8 alkyl, C1-C8 haloalkyl, C(═
NH)N(H)Z, C(═
O)N(H)Z or OC(═
O)N(H)Z;
Z is H or C1-C8 alkyl;
L1, L2 and L3 form a ring system having from about 4 to about 7 carbon atoms or having from about 3 to about 6 carbon atoms and 1 or 2 heteroatoms selected from oxygen, nitrogen and sulfur and wherein said ring system is aliphatic, unsaturated aliphatic, aromatic, or saturated or unsaturated heterocyclic;
Y is alkyl or haloalkyl having 1 to about 10 carbon atoms, alkenyl having 2 to about 10 carbon atoms, alkynyl having 2 to about 10 carbon atoms, aryl having 6 to about 14 carbon atoms, N(Q1)(Q2), O(Q1), halo, S(Q1), or CN;
each q1 is, independently, from 2 to 10;
each q2 is, independently, 0 or 1;
m is 0, 1 or 2;
p is from 0 to 10; and
q3 is from 1 to 10 with the proviso that when p is 0, q3 is greater than 1.
- -fluoro nucleotide or a nucleotide having a 2′
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23. The oligonucleotide of claim 22 wherein at least one nucleotide having A-form conformational geometry has the 2′
- -substituent group of formula I and p is 0.
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24. The oligonucleotide of claim 17 wherein at least one of said nucleotides having A-form conformational geometry is a 2′
- -F ribonucleotide, a 2′
-O—
(C1-C6 alkyl) ribonucleotide, or a 2′
-O—
(C1-C6 substituted alkyl) ribonucleotide wherein the substitution is C1-C6 ether, C1-C6 thioether, amino, amino(C1-C6 alkyl) or amino(C1-C6 alkyl)2.
- -F ribonucleotide, a 2′
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25. The oligonucleotide of claim 17 wherein at least one of said nucleotides having B-form conformational geometry is a 2′
- -CN arabinonucleotide, a 2′
-F arabinonucleotide, a 2′
-Cl arabinonucleotide, a 2′
-Br arabinonucleotide, a 2′
-N3 arabinonucleotide, a 2′
-OH arabinonucleotide, a 2′
-O—
CH3 arabinonucleotide or a 2′
-dehydro-2′
-CH3 arabinonucleotide.
- -CN arabinonucleotide, a 2′
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26. The oligonucleotide of claim 25 wherein at least one of said nucleotides having B-form conformational geometry is a 2′
- -F arabinonucleotide, a 2′
-OH arabinonucleotide or a 2′
-O—
CH3 arabinonucleotide.
- -F arabinonucleotide, a 2′
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27. The oligonucleotide of claim 26 wherein at least one of said nucleotides having B-form conformational geometry is a 2′
- -F arabinonucleotide or a 2′
-OH arabinonucleotide.
- -F arabinonucleotide or a 2′
-
28. The oligonucleotide of claim 27 wherein at least one of said nucleotides having B-form conformational geometry is a 2′
- -F arabinonucleotide.
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29. The oligonucleotide of claim 17 wherein at least one of said nucleotides having B-form conformational geometry is a 2′
- -CN arabinonucleotide, a 2′
-F arabinonucleotide, a 2′
-Cl arabinonucleotide, a 2′
-Br arabinonucleotide, a 2′
-N3 arabinonucleotide, a 2′
-OH arabinonucleotide, a 2′
-O—
CH3 arabinonucleotide or a 2′
-dehydro-2′
-CH3 arabinonucleotide; andat least one of said nucleotides having A-form conformational geometry is a 2′
-F ribonucleotide, a 2′
-O—
(C1-C6 alkyl) ribonucleotide, or a 2′
-O—
(C1-C6 substituted alkyl) ribonucleotide wherein the substitution is C1-C6 ether, C1-C6 thioether, amino, amino(C1-C6 alkyl) or amino(C1-C6 alkyl)2.
- -CN arabinonucleotide, a 2′
-
30. The oligonucleotide of claim 29 wherein at least one of said nucleotides having B-form conformational geometry is a 2′
- -F arabinonucleotide or a 2′
-OH arabinonucleotide; andat least one of said nucleotides having A-form conformational geometry is a 2′
-O—
(C1-C6 substituted alkyl) ribonucleotide wherein the substitution is C1-C6 ether, C1-C6 thioether, amino, amino(C1-C6 alkyl) or amino(C1-C6 alkyl)2.
- -F arabinonucleotide or a 2′
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31. The oligonucleotide of claim 30 wherein at least one of said nucleotides having A-form conformational geometry is a 2′
- -alkylamino substituted nucleotide.
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32. The oligonucleotide of claim 1 further comprising at least one nucleotide having B-form conformational geometry bound by an internucleotide linkage to a 3′
- or a 5′
position of at least one of said sequence of nucleotides having A-form conformational geometry.
- or a 5′
-
33. The oligonucleotide of claim 32 wherein said nucleotide having B-form conformational geometry is bound to said oligonucleotide by a 3′
- -5′
phosphodiester, 2′
-5′
phosphodiester, phosphorothioate, Sp phosphorothioate, Rp phosphorothioate, phosphorodithioate, 3′
-deoxy-3′
-amino phosphoroamidate, 3′
-methylenephosphonate, methylene(methylimino), dimethylhydrazino, amide 3, amide 4 or a boranophosphate internucleotide linkage.
- -5′
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34. The oligonucleotide of claim 33 wherein said internucleotide linkage is a 2′
- -5′
phosphodiester, 3′
-methylenephosphonate, Sp phosphorothioate or a methylene(methylimino).
- -5′
-
35. The oligonucleotide of claim 1 wherein at least one nucleotide of said sequence of nucleotides has a C3′
- endo type pucker; and
at least one of said nucleotides has a C2′
endo type pucker or an O4′
endo type pucker, provided that said nucleotides are not 2′
-deoxy-erythro-pentofuranosyl nucleotides.
- endo type pucker; and
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36. The oligonucleotide of claim 35 wherein at least one of said nucleotides having said C3′
- endo type pucker is bound to a 3′
position to at least one of said nucleotides having said C2′
endo type pucker or O4′
endo type pucker.
- endo type pucker is bound to a 3′
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37. The oligonucleotide of claim 35 wherein at least one of said nucleotides having said C3′
- endo type pucker is bound to a 5′
position to at least one of said nucleotides having said C2′
endo type pucker or O4′
endo type pucker.
- endo type pucker is bound to a 5′
-
38. The oligonucleotide of claim 35 wherein at least one of said nucleotides having said C3′
- endo type pucker is bound to a 3′
position to at least one of said nucleotides having said C2′
endo type pucker or O4′
endo type pucker; andat least one of said nucleotides having said C3′
endo type pucker is bound to a 5′
position to at least one of said nucleotides having said C2′
endo type pucker or O4′
endo type pucker.
- endo type pucker is bound to a 3′
Specification