Claims for Patent: 9,370,582

Summary for Patent: 9,370,582

Title:	Carbohydrate conjugates as delivery agents for oligonucleotides
Abstract:	The present invention provides iRNA agents comprising at least one subunit of the formula (I): ##STR00001## wherein: A and B are each independently for each occurrence O, N(R.sup.N) or S; X and Y are each independently for each occurrence H, OH, a hydroxyl protecting group, a phosphate group, a phosphodiester group, an activated phosphate group, an activated phosphite group, a phosphoramidite, a solid support, --P(Z')(Z'')O-nucleoside, --P(Z')(Z'')O-oligonucleotide, a lipid, a PEG, a steroid, a lipophile, a polymer, --P(Z')(Z'')O-Linker-OP(Z''')(Z'''')O-oligonucleotide, a nucleotide, an oligonucleotide, --P(Z')(Z'')-formula (I), --P(Z')(Z'')- or -Linker-R; R is L.sup.G, -Linker-L.sup.G, or has the structure shown below: ##STR00002## L.sup.G is independently for each occurrence a carbohydrate, e.g., monosaccharide, disaccharide, trisaccharide, tetrasaccharide, oligosaccharide, polysaccharide; R.sup.N is independently for each occurrence H, methyl, ethyl, propyl, isopropyl, butyl, or benzyl; and Z', Z'', Z''' and Z'''' are each independently for each occurrence O or S.
Inventor(s):	Manoharan; Muthiah (Cambridge, MA), Rajeev; Kallanthottathil G. (Cambridge, MA), Nair; Jayaprakash (Cambridge, MA), Maier; Martin (Cambridge, MA)
Assignee:	ALNYLAM PHARMACEUTICALS, INC. (Cambridge, MA)
Application Number:	14/329,557
Patent Claims:	1. A modified PCSK9-targeting oligonucleotide comprising a structure shown below: ##STR00120## wherein L.sup.G is independently for each occurrence a ligand; and wherein the oligonucleotide targets the PCSK9 gene. 2. A modified PCSK9-targeting oligonucleotide comprising a structure shown in formula (II)-(V): ##STR00121## wherein: q.sup.2A, q.sup.2B, q.sup.3A, q.sup.3B, q.sup.4A, q.sup.4B, q.sup.5A, q.sup.5B and q.sup.5C for each represent independently occurrence 0-20 and wherein the repeating unit can be the same or different; P.sup.2A, P.sup.2B, P.sup.3A, P.sup.3B, P.sup.4A, P.sup.4B, P.sup.5A, P.sup.5B, P.sup.5C, T.sup.2A, T.sup.2B, T.sup.3A, T.sup.3B, T.sup.4A, T.sup.4B, T.sup.5A, T.sup.5B, and T.sup.5C are each independently for each occurrence absent, CO, NH, O, S, OC(O), NHC(O), CH.sub.2, CH.sub.2NH or CH.sub.2O; Q.sup.2A, Q.sup.2B, Q.sup.3A, Q.sup.3B, Q.sup.4A, Q.sup.4B, Q.sup.5A, Q.sup.5B, and Q.sup.5C are independently for each occurrence absent, alkylene, substituted alkylene wherein one or more methylenes can be interrupted or terminated by one or more of O, S, S(O), SO.sub.2, N(R.sup.N), C(R').dbd.C(R''), C.ident.C or C(O); R.sup.2A, R.sup.2B, R.sup.3A, R.sup.3B, R.sup.4A, R.sup.4B, R.sup.5A, R.sup.5B, and Q.sup.5C are each independently for each occurrence absent, NH, O, S, CH.sub.2, C(O)O, C(O)NH, NHCH(R.sup.a)C(O), --C(O)--CH(R.sup.a)--NH--, CO, CH.dbd.N--O, ##STR00122## or heterocyclyl; L.sup.2A, L.sup.2B, L.sup.3A, L.sup.3B, L.sup.4A, L.sup.4B, L.sup.5A, L.sup.5B and L.sup.5C are each independently for each occurrence a carbohydrate; R' and R'' are each independently H, C.sub.1-C.sub.6 alkyl, OH, SH or N(R.sup.N).sub.2; R.sup.N is independently for each occurrence methyl, ethyl, propyl, isopropyl, butyl or benzyl; and R.sup.a is H or amino acid side chain, wherein the oligonucleotide targets the PCSK9 gene. 3. The modified oligonucleotide of claim 1, wherein the modified oligonucleotide is double stranded. 4. The modified oligonucleotide of claim 3, wherein the structure is at the 3'-end of one of the strands. 5. The modified oligonucleotide of claim 4, wherein the structure is at the 3'-end of the sense strand. 6. The modified oligonucleotide of claim 1, wherein the structure is ##STR00123## 7. The modified oligonucleotide of claim 1, wherein the structure is ##STR00124## 8. The modified oligonucleotide of claim 1, wherein L.sup.G is carbohydrate. 9. The modified oligonucleotide of claim 8, wherein the carbohydrate is selected from the group consisting of monosaccharide, disaccharide, trisaccharide, tetrasaccharide and polysaccharide. 10. The modified oligonucleotide of claim 1, wherein L.sup.G is galactose or N-acetylgalactosamine. 11. The modified oligonucleotide of claim 1, wherein at least one of the nucleotide sugars of the modified oligonucleotide is modified at the 2' position to replace 2' hydroxyl. 12. The modified oligonucleotide of claim 11, wherein the 2' hydroxyl is replaced with a moiety selected from the group consisting of 2'-H, 2'-methoxyethyl, 2'-OCH.sub.3, 2'-O-allyl, 2'-C-allyl, and 2'-fluoro. 13. The modified oligonucleotide of claim 1, wherein the oligonucleotide comprises 2'-deoxy, 2'-methoxyethyl, and a phosphorothioate linkage. 14. The modified oligonucleotide of claim 1, wherein the oligonucleotide comprises 2'-deoxy, LNA, and a phosphorothioate linkage. 15. The modified oligonucleotide of claim 1, wherein the modified oligonucleotide is single stranded. 16. The modified oligonucleotide of claim 15, wherein the modified oligonucleotide is a hairpin or antisense. 17. A method of modulating the expression of a target PCSK9 gene in a cell, comprising providing to said cell the modified oligonucleotide of claim 1. 18. A pharmaceutical composition comprising the modified oligonucleotide of claim 1 alone or in combination with a pharmaceutically acceptable carrier or excipient. 19. A modified PCSK9-targeting oligonucleotide comprising a structure of ##STR00125## wherein the oligonucleotide targets the PCSK9 gene. 20. A modified PCSK9-targeting oligonucleotide comprising a structure of ##STR00126## wherein the oligonucleotide targets the PCSK9 gene.