US 8,013,136 B2 · Oligonucleotides comprising a non-phosphate backbone linkage

Abstract

One aspect of the present invention relates to a ribonucleoside substituted with a phosphonamidite group at the 3′-position. In certain embodiments, the phosphonamidite is an alkyl phosphonamidite. Another aspect of the present invention relates to a double-stranded oligonucleotide comprising at least one non-phosphate linkage. Representative non-phosphate linkages include phosphonate, hydroxylamine, hydroxylhydrazinyl, amide, and carbamate linkages. In certain embodiments, the non-phosphate linkage is a phosphonate linkage. In certain embodiments, a non-phosphate linkage occurs in only one strand. In certain embodiments, a non-phosphate linkage occurs in both strands. In certain embodiments, a ligand is bound to one of the oligonucleotide strands comprising the double-stranded oligonucleotide. In certain embodiments, a ligand is bound to both of the oligonucleotide strands comprising the double-stranded oligonucleotide. In certain embodiments, the oligonucleotide strands comprise at least one modified sugar moiety. Another aspect of the present invention relates to a single-stranded oligonucleotide comprising at least one non-phosphate linkage. Representative non-phosphate linkages include phosphonate, hydroxylamine, hydroxylhydrazinyl, amide, and carbamate linkages. In certain embodiments, the non-phosphate linkage is a phosphonate linkage. In certain embodiments, a ligand is bound to the oligonucleotide strand. In certain embodiments, the oligonucleotide comprises at least one modified sugar moiety.

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Claims35

Dependent claims

A double-stranded iRNA agent comprising a first strand and a second strand, wherein said first strand and said second strand are represented independently by formula II:

wherein
X1 is H, —P(O)(OM)2, —P(O)(OM)-O—P(O)(OM)2, —P(O)(Oalkyl)2, —P(O)(Oalkyl)-O—P(O)(Oalkyl)2, or -A6-[A7-(A5)w]y;
M represents independently for each occurrence an alkali metal or a transition metal with an overall charge of +1;
R1 and R5 represent independently for each occurrence H, alkyl, or halogen;
R2 and R3 represent independently for each occurrence H, OH, F, —Oalkyl, —Oallyl, —O(C(R19)2)kOR19, —O(C(R19)2)kSR19, —O(C(R19)2)kN(R19)2, —O(C(R19)2)kC(O)N(R19)2, —N(R19)2, —S(C1-C6)alkyl, —O(C(R19)2)kO(C1-C6)alkyl, —O(C(R19)2)kS(C1-C6)alkyl, —O(C(R19)2)kO(C(R19)2)kN((C1-C6)alkyl)2, —O(C(R19)2)kON((C1-C6)alkyl)2, or —O-A6-[A7-(A5)w]y;
R4 represents independently for each occurrence H, OH, F, —Oalkyl, —Oallyl, —O(C(R19)2)kOR19, —O(C(R19)2)kSR19, —O(C(R19)2)kN(R19)2, —O(C(R19)2)kC(O)N(R19)2, —N(R19)2, —S(C1-C6)alkyl, —O(C(R19)2)kO(C1-C6)alkyl, —O(C(R19)2)kS(C1-C6)alkyl, —O(C(R19)2)kO(C(R19)2)kN((C1-C6)alkyl)2, or —O(C(R19)2)kON((C1-C6)alkyl)2;
R6, R7, and R9 represent independently for each occurrence H, alkyl, aryl, or aralkyl;
R8 represents independently for each occurrence alkyl, aryl, or aralkyl;
k represents independently for each occurrence 1, 2, 3, or 4;
n1 is 1, 2, or 3;
n2 is an integer in the range of about 15-28, inclusive;
w represents independently for each occurrence 1, 2, or 3 in accord with the rules of valence;
x represents independently for each occurrence 0, 1, 2, or 3;
y represents independently for each occurrence 1, 2, 3, 4, or 5 in accord with the rules of valence;
A1 represents independently for each occurrence:
A2 represents independently for each occurrence
A3 represents independently for each occurrence
A4 represents independently for each occurrence a bond, alkyl diradical, heteroalkyl diradical, alkenyl diradical, alkynyl diradical, alkylalkynyl diradical, aminoalkyl diradical, thioether, —C(O)—, —S(O)—, —S(O)2—, B1C(R)2B2, B1C(R)(B2)2, B1C(B2)3, B1N(R)(B2), B1N(B2)2, or has the formula:
B1 is a bond between A3 and A4;
B2 is a bond between A4 and A5;
R represents independently for each occurrence hydrogen or alkyl;
m represents independently for each occurrence 1, 2, 3, 4, 5, 6, 7, or 8;
m1 represents independently for each occurrence 0, 1, 2, 3, 4, 5, 6, 7, or 8;
Y represents independently for each occurrence an alkyl diradical, cycloalkyl diradical, heteroalkyl diradical, heterocycloalkyl diradical, alkenyl diradical, alkynyl diradical, aryl diradical, heteroaryl diradical, aralkyl diradical, heteroaralkyl diradical, —X2C(O)X2[C(R5)2]vX2—, —X2C(NR6)X2[C(R5)2]vX2—, —X2C(S)X2[C(R5)2]vX2—, —X2C(O)X2[C(R5)2]vX2C(O)X2—,
[C(R5)2]tN(R6)O[C(R5)2]t—, —[C(R5)2]tN(R6)N(R6)O[C(R5)2]t—, —[C(R5)2]tN(R7)C(O)[C(R5)2]t—, —[C(R5)2]tN(R)CO2[C(R5)2]t—, —[C(R5)2]tN(R7)C(S)[C(R5)2]t—, —[C(R5)2]tN(R7)C(S)O[C(R5)2]t—, —[C(R5)2]tOC(O)S[C(R5)2]t—, —[C(R5)2]tSN(R7)CO2[C(R5)2]t—, —[C(R5)2]t—OSi(R8)2—O[C(R5)2]t—, —[C(R5)2]t—OSO2N(R7)[C(R5)2]t—, —[C(R5)2]tN(R7)SO2N(R7)[C(R5)2]t—, —[C(R5)2]tSO2N(morpholino)-[C(R5)2]t—, —[C(R5)2]tSO2N(R7)[C(R5)2]t—, —[C(R1)2]tS[C(R5)2]t—, —[C(R5)2]t—OSO2[C(R5)2]t—, —[C(R5)2]tS[C(R5)2]y—O[C(R5)2]t—, —[C(R5)2]tO[C(R5)2]yO[C(R5)2]t—, —[C(R5)2]tO[C(R5)2]t—, —[C(R5)2]tN(R7)[C(R5)2]t—, —[C(R5)2]tC═NO[C(R5)2]t—, —[C(R5)2]tC(O)C(R5)═C(R5)[C(R5)2]t—, —[C(R5)2]tC(R5)═C(R5)[C(R5)2]t—, or —[C(R5)2]tX2C(O)X2[C(R5)2]t—;
X2 represents independently for each occurrence a bond, O, or N(R6);
Z1 represents independently for each occurrence O, S, or N(R8);
Z2 represents independently for each occurrence alkyl, aryl, aralkyl, B(R9)3, —OM, —Oalkyl, —Oaryl, —Oaralkyl, —SM, —Salkyl, —Saryl, —Saralkyl, —[C(R5)2]mN(R6)2, —N(R10)R11, —N(R19)(C(R19)2)mN(R19)2, —N(R7)C(O)R8, H, —OC(O)R8, —CO2R8, F, Se, —SeR8, —(C(R19)2)mOR19, —(C(R19)2)mSR19, —N(R19)(C(R19)2)m—OR19, —N(R19)(C(R19)2)mSR19, —N(R19)(C(R19)2)mN(R19)C(O)alkyl, —(C(R19)2)mN(R19)C(O)alkyl, or -A8-[A9-(A5)w]y;
R10 and R11 are independently H, alkyl, or aryl;
or R10 and R11 taken together form a 3-, 4-, 5-, 6-, or 7-member ring;
R12 represents independently for each occurrence H, alkyl, or —NHCH2CH═CH2;
t represents independently for each occurrence 0, 1, 2, 3, or 4;
v represents independently for each occurrence 0, 1, 2, 3, 4, 5, 6, 7, or 8;
A5 represents independently for each occurrence aryl, aralkyl, or the radical of a steroid, bile acid, lipid, folic acid, pyridoxal, B12, riboflavin, biotin, polycyclic compound, crown ether, intercalator, cleaver molecule, protein-binding agent, carbohydrate, or an optionally substituted saturated 5-membered ring;
A6 represents independently for each occurrence a bond, alkyl diradical, heteroalkyl diradical, alkenyl diradical, aminoalkyl, —C(O)—, —S(O)—, —S(O)2—, or is represented by formula:
Z3 represents independently for each occurrence O or S;
Z4 represents independently for each occurrence —OM, —Oalkyl, —Oaryl, —Oaralkyl, —SM, —Salkyl, —Saryl, —Saralkyl, —N(R10)R11, —[C(R5)2]mN(R6)2, —N(R19)(C(R19)2)mN(R19)2, —(C(R19)2)mOR19, —(C(R19)2)mSR19, —N(R19)(C(R19)2)m—OR19, —N(R19)(C(R19)2)mSR19, —N(R19)(C(R19)2)mN(R19)C(O)alkyl, —(C(R19)2)mN(R19)C(O)alkyl, aryl, or alkyl;
R13 represent independently for each occurrence H, alkyl, cycloalkyl, heteroalkyl, aryl, aralkyl, acyl, silyl, or B3;
R14 represents independently for each occurrence alkyl, aryl, aralkyl, acyl, or silyl;
R15 represents independently for each occurrence hydrogen, alkyl, aryl, aralkyl, acyl, alkylsulfonyl, alkylsulfoxide, arylsulfonyl, arylsulfoxide, or silyl;
R16 represents independently for each occurrence cycloalkyl, heterocycloalkyl, aryl, or heteroaryl;
B3 is a bond between A6 and A7;
B4 is a bond between A6 and O;
n3 represents independently for each occurrence an integer in the range of 1-15, inclusive;
n4 represents independently for each occurrence 1, 2, 3, 4, or 5 in accord with the rules of valence;
A7 represents independently for each occurrence a bond, alkyl diradical, heteroalkyl diradical, —C(O)—, —S(O)—, —S(O)2—, B3C(R)2B5, B3C(R)(B5)2, B3C(B5)3, B3N(R)(B5), B3N(B5)2, or has the formula:
p represents independently for each occurrence 1, 2, 3, or 4;
B5 is a bond between A5 and A7;
A8 is a bond, alkyl diradical, heteroalkyl diradical, alkenyl diradical, aminoalkyl, or is represented by formula:
R17 represent independently for each occurrence H, alkyl, cycloalkyl, heteroalkyl, aryl, aralkyl, acyl, silyl, or B6;
R18 represents independently for each occurrence H, halogen, alkyl, alkoxyl, —N(R6)2, —CN, —[C(R5)2]vC(R5)═C(R5)2;
R19 represents independently for each occurrence H or alkyl;
B6 is a bond between A8 and A9;
B7 is a bond between A8 and P;
A9 is a bond, alkyl diradical, heteroalkyl diradical, —C(O)—, —S(O)—, —S(O)2—, B6C(R)2B8, B6C(R) (B8)2, B6C(B8)3, B6N(R)(B8), B6N(B8)2, or has the formula:
and
B8 is a bond between A5 and A9;
provided that at least one occurrence of Y is a P-alkylphosphonate.

55 more elements

A single-stranded iRNA agent represented by formula V:

wherein
X1 is H, —P(O)(OM)2, —P(O)(OM)-O—P(O)(OM)2, —P(O)(Oalkyl)2, —P(O)(Oalkyl)-O—P(O)(Oalkyl)2, or -A6-[A7-(A5)w]y;
M represents independently for each occurrence an alkali metal or a transition metal with an overall charge of +1;
R1 and R5 represent independently for each occurrence H, alkyl, or halogen;
R2 and R3 represent independently for each occurrence H, OH, F, —Oalkyl, —Oallyl, —O(C(R19)2)kOR19, —O(C(R19)2)kSR19, —O(C(R19)2)kN(R19)2, —O(C(R19)2)kC(O)N(R19)2, —N(R19)2, —S(C1-C6)alkyl, —O(C(R19)2)kO(C1-C6)alkyl, —O(C(R19)2)kS(C1-C6)alkyl, —O(C(R19)2)kO(C(R19)2)kN((C1-C6)alkyl)2, —O(C(R19)2)kON((C1-C6)alkyl)2, or —O-A6-[A7-(A5)w]y;
R4 represents independently for each occurrence H, OH, F, —Oalkyl, —Oallyl, —O(C(R19)2)kOR19, —O(C(R19)2)kSR19, —O(C(R19)2)kN(R19)2, —O(C(R19)2)kC(O)N(R19)2, —N(R19)2, —S(C1-C6)alkyl, —O(C(R19)2)kO(C1-C6)alkyl, —O(C(R19)2)kS(C1-C6)alkyl, —O(C(R19)2)kO(C(R19)2)kN((C1-C6)alkyl)2, or —O(C(R19)2)kON((C1-C6)alkyl)2;
R6, R7, and R9 represent independently for each occurrence H, alkyl, aryl, or aralkyl;
R8 represents independently for each occurrence alkyl, aryl, or aralkyl;
k represents independently for each occurrence 1, 2, 3, or 4;
n1 is 1, 2, or 3;
n2 is an integer in the range of about 15-28, inclusive;
w represents independently for each occurrence 1, 2, or 3 in accord with the rules of valence;
x represents independently for each occurrence 0, 1, 2, or 3;
y represents independently for each occurrence 1, 2, 3, 4, or 5 in accord with the rules of valence;
A1 represents independently for each occurrence:
A2 represents independently for each occurrence:
A3 represents independently for each occurrence
A4 represents independently for each occurrence a bond, alkyl diradical, heteroalkyl diradical, alkenyl diradical, alkynyl diradical, alkylalkynyl diradical, aminoalkyl diradical, thioether, —C(O)—, —S(O)—, —S(O)2—, B1C(R)2B2, B1C(R)(B2)2, B1C(B2)3, B1N(R)(B2), B1N(B2)2, or has the formula:
B1 is a bond between A3 and A4;
B2 is a bond between A4 and A5;
R represents independently for each occurrence hydrogen or alkyl;
m represents independently for each occurrence 1, 2, 3, 4, 5, 6, 7, or 8;
m1 represents independently for each occurrence 0, 1, 2, 3, 4, 5, 6, 7, or 8;
Y represents independently for each occurrence an alkyl diradical, cycloalkyl diradical, heteroalkyl diradical, heterocycloalkyl diradical, alkenyl diradical, alkynyl diradical, aryl diradical, heteroaryl diradical, aralkyl diradical, heteroaralkyl diradical, —X2C(O)X2[C(R5)2]vX2—, —X2C(NR6)X2[C(R5)2]vX2—, —X2C(S)X2[C(R5)2]vX2—, —X2C(O)X2-[C(R5)2]vX2C(O)X2—,
[C(R5)2]tN(R6)O[C(R5)]t—, —[C(R5)2]tN(R6)N(R6)O[C(R5)]t—, —[C(R5)2]tN(R7)C(O)[C(R5)2]t—, —[C(R5)2]tN(R)CO2[C(R)2]t—, —[C(R5)2]tN(R7)C(S)[C(R5)2]t—, —[C(R5)2]tN(R7)C(S)O[C(R5)2]t—, —[C(R5)2]t—OC(O)S[C(R5)2]t—, —[C(R5)2]tSN(R7)CO2[C(R5)2]t—, —[C(R5)2]tOSi(R8)2—O[C(R5)2]t—, —[C(R5)2]tOSO2N(R7)[C(R5)2]t—, —[C(R5)2]tN(R7)SO2N(R7)[C(R5)2]t—, —[C(R5)2]tSO2N(morpholino)-[C(R5)2]t—, —[C(R5)2]tSO2N(R7)[C(R5)2]t—, —[C(R1)2]tS[C(R5)2]t—, —[C(R5)2]t—OSO2[C(R5)2]t—, —[C(R5)2]tS[C(R5)2]y—O[C(R5)2]t—, —[C(R5)2]tO[C(R5)2]y—O[C(R5)2]t—, —[C(R5)2]tO[C(R5)2]t—, —[C(R5)2]tN(R7)[C(R5)2]t—, —[C(R5)2]tC═NO[C(R5)2]t—, —[C(R5)2]tC(O)C(R5)═C(R5)[C(R5)2]t—, —[C(R5)2]tC(R5)═C(R5)[C(R5)2]t—, or —[C(R5)2]tX2C(O)X2[C(R5)2]t—;
X2 represents independently for each occurrence a bond, O, or N(R6);
Z1 represents independently for each occurrence O, S, or N(R8);
Z2 represents independently for each occurrence alkyl, aryl, aralkyl, B(R9)3, —OM, —Oalkyl, —Oaryl, —Oaralkyl, —SM, —Salkyl, —Saryl, —Saralkyl, —[C(R5)2]mN(R6)2, —N(R10)R11, —N(R19)(C(R19)2)mN(R19)2, —N(R7)C(O)R8, H, —OC(O)R8, —CO2R8, F, Se, —SeR8, —(C(R19)2)m—OR19, —(C(R19)2)mSR19, —N(R19)(C(R19)2)mOR19, —N(R19)(C(R19)2)mSR19, —N(R19)(C(R19)2)mN(R19)C(O)alkyl, —(C(R19)2)mN(R19)C(O)alkyl, or -A8-[A9-(A5)w]y;
R10 and R11 are independently H, alkyl, or aryl;
or R10 and R11 taken together form a 3-, 4-, 5-, 6-, or 7-member ring;
R12 represents independently for each occurrence H, alkyl, or —NHCH2CH═CH2;
t represents independently for each occurrence 0, 1, 2, 3, or 4;
v represents independently for each occurrence 0, 1, 2, 3, 4, 5, 6, 7, or 8;
A5 represents independently for each occurrence aryl, aralkyl, or the radical of a steroid, bile acid, lipid, folic acid, pyridoxal, B12, riboflavin, biotin, polycyclic compound, crown ether, intercalator, cleaver molecule, protein-binding agent, carbohydrate, or an optionally substituted saturated 5-membered ring;
A6 represents independently for each occurrence a bond, alkyl diradical, heteroalkyl diradical, alkenyl diradical, aminoalkyl, —C(O)—, —S(O)—, —S(O)2—, or is represented by formula:
Z3 represents independently for each occurrence O or S;
Z4 represents independently for each occurrence —OM, —Oalkyl, —Oaryl, —Oaralkyl, —SM, —Salkyl, —Saryl, —-Saralkyl, —N(R10)R11, —[C(R5)2]mN(R6)2, —N(R19)(C(R19)2)mN(R19)2, —(C(R19)2)mOR19, —(C(R19)2)mSR19, —N(R19)(C(R19)2)m—OR19, —N(R19)(C(R19)2)mSR19, —N(R19)(C(R19)2)mN(R19)C(O)alkyl, —(C(R19)2)mN(R19)C(O)alkyl, aryl, or alkyl;
R13 represent independently for each occurrence H, alkyl, cycloalkyl, heteroalkyl, aryl, aralkyl, acyl, silyl, or B3;
R14 represents independently for each occurrence alkyl, aryl, aralkyl, acyl, or silyl;
R15 represents independently for each occurrence hydrogen, alkyl, aryl, aralkyl, acyl, alkylsulfonyl, alkylsulfoxide, arylsulfonyl, arylsulfoxide, or silyl;
R16 represents independently for each occurrence cycloalkyl, heterocycloalkyl, aryl, or heteroaryl;
B3 is a bond between A6 and A7;
B4 is a bond between A6 and O;
n3 represents independently for each occurrence an integer in the range of 1-15, inclusive;
n4 represents independently for each occurrence 1, 2, 3, 4, or 5 in accord with the rules of valence;
A7 represents independently for each occurrence a bond, alkyl diradical, heteroalkyl diradical, —C(O)—, —S(O)—, —S(O)2—, B3C(R)2B5, B3C(R)(B5)2, B3C(B5)3, B3N(R)(B5), B3N(B5)2, or has the formula:
p represents independently for each occurrence 1, 2, 3, or 4;
B5 is a bond between A5 and A7;
A8 is a bond, alkyl diradical, heteroalkyl diradical, alkenyl diradical, aminoalkyl, or is represented by formula:
R17 represent independently for each occurrence H, alkyl, cycloalkyl, heteroalkyl, aryl, aralkyl, acyl, silyl, or B6;
R18 represents independently for each occurrence H, halogen, alkyl, alkoxyl, —N(R6)2, —CN, —[C(R5)2]vC(R5)═C(R5)2;
R19 represents independently for each occurrence H or alkyl;
B6 is a bond between A8 and A9;
B7 is a bond between A8 and P;
A9 is a bond, alkyl diradical, heteroalkyl diradical, —C(O)—, —S(O)—, —S(O)2—, B6C(R)2B8, B6C(R)(B8)2, B6C(B8)3, B6N(R)(B8), B6N(B8)2, or has the formula:
and
B8 is a bond between A5 and A9;
provided that at least one occurrence of Y is a P-alkylphosphonate.

55 more elements

Dependencies are inferred from the claim text.

Description64,288 words

Summary2,098 words

#Oligonucleotide compounds have important therapeutic applications in medicine. Oligonucleotides can be used to silence genes that are responsible for a particular disease. Gene-silencing prevents formation of a protein by inhibiting translation. Importantly, gene-silencing agents are a promising alternative to traditional small, organic compounds that inhibit the function of the protein linked to the disease. siRNA, antisense RNA, and micro-RNA are oligonucleotides that prevent the formation of proteins by gene-silencing.

#siRNA

#RNA interference (RNAi) is an evolutionarily conserved gene silencing mechanism, originally discovered in studies of the nematode Caenorhabditis elegans (Lee et al, Cell 75:843 (1993); Reinhart et al., Nature 403:901 (2000)). It is triggered by introducing dsRNA into cells expressing the appropriate molecular machinery, which then degrades the corresponding endogenous mRNA. The mechanism involves conversion of dsRNA into short RNAs that direct ribonucleases to homologous mRNA targets (summarized, Ruvkun, Science 2294:797 (2001)). This process is related to normal defense against viruses and the mobilization of transposons.

Brief description of the drawings13 figures

#FIG. 1 depicts various oligonucleotides that are conjugated to a ligand. Note that NA is an oligonucleotide (or a nucleic acid) comprising of either RNA or DNA or chimeric RNA-DNA, DNA-RNA, RNA-DNA-RNA or DNA-RNA-DNA. In certain instances, at least one among R1, R2 and R3 is aromatic or substituted aromatic, when R1 is aromatic or substituted aromatic, R2 is either H or any organic substituent, and R3 is either H or any organic substituent.

#FIG. 2 depicts various oligonucleotides that are conjugated to a ligand. Note: In certain instances, at least one among R1, R2 and R3 is aromatic or substituted aromatic. For rows A-E: NA=DNA or RNA. For row A: racemic and R and S isomers. For rows B and C: racemic and all four stereo isomers (RR, RS, SR and SS). For rows D and E: R═H or OH.

#FIG. 3 depicts various NA building blocks with a serinol linker (see row A in FIG. 2) having aralkyl ligands linked through alkyl and PEG tethers. Each ligand shown is either racemic or optically enriched or pure R or S isomer.

Detailed description61,570 words

#One aspect of the present invention relates to a ribonucleoside substituted with a phosphonamidite group at the 3′-position. These compounds can be used to prepare oligonucleotides used in gene therapy. Oligonucleosides prepared from 3′-phosphonamidite substituted nucleosides have phosphonate linkages which are less prone to degradation in vivo. In certain instances, the phosphonamidite is an alkyl phosphonamidite. In addition, the 2′-position of the ribonucleoside can be protected with a protecting group that can be easily removed under mild conditions. One example of a protecting group that can be removed under mild conditions is a silyl protecting group. In a preferred embodiment, the protecting group is tert-butyldimethylsilyl.

#Another aspect of the present invention relates to a double-stranded oligonucleotide comprising at least one non-phosphate linkage. The non-phosphate moiety renders the oligonucleotide less prone to degradation in vivo. A large number of non-phosphate functional groups are known in the art and are amenable to the present invention. The non-phosphate linkage can be a functional group that contains a phosphorous atom, or a functional group that does not contain a phosphorous atom. Representative non-phosphate linkages amenable to the present invention are phosphonate, hydroxylamine, hydroxylhydrazinyl, amide, and carbamate linkages. In certain embodiments, the non-phosphate linkage is a phosphonate linkage. The non-phosphate linkage can occur in only one strand or in both strands. In certain instances, there are about 1-5 non-phosphate linkages per double-stranded oligonucleotide. In certain instances, there are about 5-10 non-phosphate linkages per double-stranded oligonucleotide. In certain instances, there are about 10-20 non-phosphate linkages per double-stranded oligonucleotide. In certain instances, there are about 1-2 non-phosphate linkages per strand in the double-stranded oligonucleotide. In certain instances, there are about 3-5 non-phosphate linkages per strand in the double-stranded oligonucleotide. In certain instances, there are about 5-10 non-phosphate linkages per strand in the double-stranded oligonucleotide. In certain instances, there are about 10-15 non-phosphate linkages per strand in the double-stranded oligonucleotide. A non-phosphate linkage can be located near the terminus of the oligonucleotide strand or in the interior of the oligonucleotide strand. In certain instances, a non-phosphate linkage is located between the first and second nucleoside at the 3′-terminus of the oligonucleotide strand. In certain instances, a non-phosphate linkage is located between the first and second nucleoside at the 5′-terminus of the oligonucleotide strand. In certain instances, a non-phosphate linkage is located between the first and second nucleoside at the 3′-terminus of the oligonucleotide strand, and a non-phosphate linkage is located between the first and second nucleoside at the 5′-terminus of the oligonucleotide strand. In certain instances, there are two adjacent non-phosphate linkages.

#Additional examples of non-phosphate linkages include, for example, aminoalkylphosphotriesters, methyl and other alkyl phosphonates including 3′-alkylene phosphonates, 5′-alkylene phosphonates and chiral phosphonates, phosphinates, phosphoramidates including 3′-amino phosphoramidate and aminoalkylphosphoramidates, thionophosphoramidates, thionoalkylphosphonates, thionoalkylphosphotriesters, selenophosphates and boranophosphates having normal 3′-5′ linkages, 2′-5′ linked analogs of these, and those having inverted polarity wherein one or more internucleotide linkages is a 3′ to 3′, 5′ to 5′, or 2′ to 2′ linkage. In certain instances, the oligonucleotides have inverted polarity comprising a single 3′ to 3′ linkage at the 3′-most internucleotide linkage i.e. a single inverted nucleoside residue which may be abasic (the nucleobase is missing or has a hydroxyl group in place thereof). Various salts, mixed salts and free acid forms are also included.

Oligonucleotides comprising a non-phosphate backbone linkage

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Oligonucleotides comprising a non-phosphate backbone linkage

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Abstract

Drawings

Drawings

Claims35

Description64,288 words

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Description64,288 words