Compositions and methods for inhibiting MAPT expression

Abstract

Provided herein are oligonucleotides that inhibit MAPT gene expression, including oligonucleotides conjugated to targeting ligands (e.g., lipid moieties). Also provided are compositions comprising the oligonucleotides and uses thereof, in particular to uses in treating diseases, disorders and / or conditions associated with the expression of the MAPT gene.

Description

[0001] CROSS-REFERENCE TO RELATED APPLICATIONS

[0002] This application claims priority under 35 U.S.C. §119(e) to U.S. Provisional Application No. 63 / 364,609, filed on May 12, 2022, which is incorporated herein by reference in its entirety. Technical Field

[0003] The present disclosure relates generally to biology and medicine, and more specifically to oligonucleotides and compositions comprising the oligonucleotides for inhibiting or reducing (i.e., regulating) microtubule-associated protein tau (MAPT) gene expression, and their use for treating diseases and disorders associated with MAPT gene expression. Background Art

[0004] Microtubules play several important roles in systemic cells. In the central nervous system (CNS), microtubules provide structural support and assist in transporting substances throughout the cell. Changes in microtubule quality, structure and pattern are known factors that lead to the development of many neurodegenerative diseases. Tau protein is an essential protein for the formation of microtubules, and its abnormal expression can lead to neurodegenerative diseases. Tau protein combines with tubulin to form microtubules. The alternative splicing of MAPT will produce different Tau proteins for microtubule assembly. Mutations (e.g., insertions and mismatches) that change Tau function and expression in MAPT are known causes of several diseases and disorders (e.g., Alzheimer's disease (AD), Parkinson's disease (PD) and tau protein disease) that affect CNS. Targeted MAPT gene expression is needed to prevent strategies for such diseases and disorders.

[0005] Mammal CNS is a complex system of tissue (comprising cell), fluid and chemical substance that synergistically interacts to realize multiple functions, and described function comprises motion, navigation, cognition, language, vision and emotion.Unfortunately, it is known that there are multiple CNS diseases and illness (for example, neurological disorder) and affect or destroy some or all of these functions.Usually, the treatment of CNS diseases and illness is limited to small molecule drugs, antibody and / or adaptive therapy or behavioral therapy.Continuous need develops the treatment of CNS diseases and illness relevant to improper MAPT gene expression. Summary of the invention

[0006] In order to meet this need, the present disclosure describes compositions and methods for treating diseases, disorders or illnesses related to MAPT gene expression. The present disclosure is at least partially based on double-stranded (ds) oligonucleotides, such as RNAi oligonucleotides, that effectively target and reduce the MAPT gene expression in CNS tissues and develop effective targets. Specifically, identify the target sequence in MAPT mRNA, and produce oligonucleotides that are bound to these target sequences and suppress MAPT mRNA expression. As demonstrated herein, oligonucleotides suppress the human and non-human primate (NHP) MAPT gene expression in CNS tissues. In addition, in CNS tissues related to AD or progressive supranuclear palsy (PSP), MAPT mRNA targeting oligonucleotides conjugated with N-acetylgalactosamine (GalNAc) and lipids are used, and MAPT mRNA expression is reduced. Not bound by theory, oligonucleotides described herein can be used to treat diseases, disorders or illnesses related to MAPT gene expression.

[0007] Therefore and in some aspects, the present disclosure provides an RNAi oligonucleotide for reducing MAPT gene expression, the oligonucleotide comprising a sense strand and an antisense strand, wherein the sense strand and the antisense strand form a duplex region, wherein the antisense strand comprises a complementary region to the MAPT mRNA target sequence of any one of SEQ ID NOs: 912-1295, and wherein the length of the complementary region is at least about 15 consecutive nucleotides.

[0008] In any of the foregoing or related aspects, the length of the sense strand is about 15 to about 50 nucleotides. In some aspects, the length of the sense strand is 18 to 36 nucleotides. In some aspects, the length of the antisense strand is about 15 to about 30 nucleotides. In some aspects, the length of the antisense strand is 22 nucleotides, wherein the antisense strand and the sense strand form a duplex region, the length of the duplex region is at least about 19 nucleotides, optionally at least 20 nucleotides in length. In some aspects, the length of the complementary region is at least about 19 consecutive nucleotides. In some aspects, the length of the complementary region is at least about 20 consecutive nucleotides.

[0009] In other aspects, the present disclosure provides a ds RNAi oligonucleotide for reducing MAPT gene expression, the oligonucleotide comprising:

[0010] (i) an antisense strand of about 19-30 nucleotides in length, wherein the antisense strand comprises a nucleotide sequence comprising a complementary region to a MAPT mRNA target sequence, wherein the complementary region is selected from the group consisting of SEQ ID NOs: 1296-1679, and

[0011] (ii) a sense strand of about 19-50 nucleotides in length, the sense strand comprising a complementary region to the antisense strand, wherein the antisense strand and the sense strand are separate strands forming an asymmetric duplex region having an overhang of 1-4 nucleotides at the 3' end of the antisense strand.

[0012] In some aspects, the 3' end of the sense strand comprises a stem loop as shown in S1-L-S2, wherein S1 is complementary to S2, and wherein L forms a loop of 3-5 nucleotides in length between S1 and S2. In some aspects, L is a tri-ring (triL) or a tetra-ring (tetraL). In some aspects, L is a tetraL. In some aspects, tetraL comprises a sequence 5'-GAAA-3'. In some aspects, the length of S1 and S2 is about 1 to about 10 nucleotides and has the same length. In some aspects, the length of S1 and S2 is 1 nucleotide, 2 nucleotides, 3 nucleotides, 4 nucleotides, 5 nucleotides, 6 nucleotides, 7 nucleotides, 8 nucleotides, 9 nucleotides or 10 nucleotides. In some aspects, the length of S1 and S2 is 6 nucleotides. In some aspects, the stem loop comprises a sequence 5'-GCAGCCGAAAGGCUGC-3' (SEQ ID NO: 1680).

[0013] In other aspects, the oligonucleotide comprises a blunt end. In some aspects, the blunt end comprises the 3' end of the sense strand. In some aspects, the sense strand is about 20-22 nucleotides. In some aspects, the sense strand is 20 nucleotides.

[0014] In any of the foregoing or related aspects, the antisense strand comprises a 3' overhang sequence of one or more nucleotides in length. In some aspects, the overhang comprises a purine nucleotide. In some aspects, the 3' overhang sequence is 2 nucleotides in length. In some aspects, the 3' overhang is selected from AA, GG, AG, and GA. In some aspects, the overhang is GG or AA. In some aspects, the overhang is GG.

[0015] In any one of the foregoing or related aspects, the oligonucleotide comprises at least one modified nucleotide. In some aspects, the modified nucleotide comprises a 2'-modification. In some aspects, the 2'-modification is selected from 2'-aminoethyl (EA), 2'-fluorine (2'-F), 2'-O-methyl (2'-OMe), 2'-O-methoxyethyl (2'-OME) and 2'-deoxy-2'-fluoro-β-d-arabinose nucleic acid (2'-FANA) modification. In some aspects, the modification is selected from 2'-F and 2'-OMe 2'-modification. In some aspects, about 18% to about 23% of the sense strand or 18%, 19%, 20%, 21%, 22% or 23% of the nucleotides comprise a 2'-F modification. In other aspects, about 38% to about 43% of the sense strand or 38%, 39%, 40%, 41%, 42% or 43% of the nucleotides comprise a 2'-F modification. In some aspects, about 25% to about 35% or 25%, 26%, 27%, 28%, 29%, 30%, 31%, 32%, 33%, 34% or 35% of the nucleotides of the antisense strand comprise a 2'-F modification. In some aspects, about 25% to about 35% or 25%, 26%, 27%, 28%, 29%, 30%, 31%, 32%, 33%, 34% or 35% of the nucleotides of the oligonucleotide comprise a 2'-F modification. In some aspects, about 35-45%, 35%, 36%, 37%, 38%, 39%, 40%, 41%, 42%, 43%, 44% or 45% of the nucleotides of the oligonucleotide comprise a 2'-F modification. In some aspects, the sense strand comprises 20 nucleotides from 5' to 3' at positions 1-20, wherein each of positions 3, 5, 8, 10, 12, 13, 15, and 17 comprises a 2'-F modification. In other aspects, the sense strand comprises 36 nucleotides from 5' to 3' at positions 1-36, wherein each of positions 3, 5, 8, 10, 12, 13, 15, and 17 comprises a 2'-F modification. In some aspects, the antisense strand comprises 22 nucleotides from 5' to 3' at positions 1-22, and wherein each of positions 2, 3, 4, 5, 7, 10, 14, 16, and 19 comprises a 2'-F modification. In some aspects, the remaining nucleotides comprise a 2'-OMe modification.

[0016] In any of the foregoing or related aspects, the oligonucleotide comprises at least one modified internucleotide bond. In some aspects, at least one modified internucleotide bond is a phosphorothioate bond. In some aspects, the antisense strand comprises a phosphorothioate bond at the following positions: (i) between positions 1 and 2 and between positions 2 and 3; or (ii) between positions 1 and 2, between positions 2 and 3, and between positions 3 and 4, wherein positions are numbered 1-4 from 5' to 3'. In some aspects, the antisense strand is 22 nucleotides in length, and wherein the antisense strand comprises a phosphorothioate bond between positions 20 and 21 and between positions 21 and 22, wherein positions are numbered 1-22 from 5' to 3'. In some aspects, the sense strand comprises a phosphorothioate bond between positions 1 and 2, wherein positions are numbered 1-2 from 5' to 3'. In some aspects, the sense strand is 20 nucleotides in length, and wherein the sense strand comprises phosphorothioate linkages between positions 1 and 2, between positions 18 and 19, and between positions 19 and 20, wherein positions are numbered 1-20 from 5' to 3'.

[0017] In any of the foregoing or related aspects, the 4'-carbon of the sugar of the 5'-nucleotide of the antisense strand comprises a phosphate analog. In some aspects, the phosphate analog is an oxymethylphosphonate, a vinylphosphonate, or a malonylphosphonate, optionally wherein the phosphate analog is a 4'-phosphate analog comprising a 4'-oxymethylphosphonate.

[0018] In any of the foregoing or related aspects, at least one nucleotide of the oligonucleotide is conjugated to one or more targeting ligands. In some aspects, each targeting ligand comprises a carbohydrate, an amino sugar, a lipid, cholesterol or a polypeptide. In some aspects, the stem loop comprises one or more targeting ligands conjugated to one or more nucleotides of the stem loop. In some aspects, one or more targeting ligands are conjugated to one or more nucleotides of the ring. In some aspects, the ring comprises 4 nucleotides numbered 1-4 from 5' to 3', wherein the nucleotides at positions 2, 3 and 4 each comprise one or more targeting ligands, wherein the targeting ligands are identical or different. In some aspects, each targeting ligand comprises a GalNAc moiety. In some aspects, the GalNac moiety is a monovalent GalNAc moiety, a divalent GalNAc moiety, a trivalent GalNAc moiety or a tetravalent GalNAc moiety. In some aspects, up to 4 nucleotides of the L of the stem loop are each conjugated to a monovalent GalNAc moiety.

[0019] In other aspects, one or more targeting ligands are lipid moieties. In some aspects, the lipid moiety is conjugated to the 5' terminal nucleotide of the sense strand. In some aspects, the lipid moiety is a hydrocarbon chain. In some aspects, the hydrocarbon chain is C8-C 30 In some aspects, the hydrocarbon chain is C 16 In some aspects, C 16The hydrocarbon chain is represented by:

[0020] In some aspects, the lipid moiety is conjugated to the 2' carbon of the ribose ring of the 5' terminal nucleotide.

[0021] In any of the foregoing or related aspects, the complementary region is fully complementary to the MAPT mRNA target sequence at nucleotide positions 2-8 of the antisense strand, wherein the nucleotide positions are numbered from 5' to 3'. In some aspects, the complementary region is fully complementary to the MAPT mRNA target sequence at nucleotide positions 2-11 of the antisense strand, wherein the nucleotide positions are numbered from 5' to 3'.

[0022] In any of the foregoing or related aspects, the sense strand comprises the nucleotide sequence of any one of SEQ ID NOs: 769-803 and 1681. In some aspects, the antisense strand comprises the nucleotide sequence of any one of SEQ ID NOs: 804-838.

[0023] In some aspects, the sense strand and the antisense strand comprise a nucleotide sequence selected from the group consisting of:

[0024] a) SEQ ID NO: 769 and 804, respectively;

[0025] b) SEQ ID NOs: 770 and 805, respectively;

[0026] c) SEQ ID NOs: 771 and 806, respectively;

[0027] d) SEQ ID NOs: 772 and 807, respectively;

[0028] e) SEQ ID NOs: 773 and 808, respectively;

[0029] f) SEQ ID NOs: 774 and 809, respectively;

[0030] g) SEQ ID NOs: 775 and 810, respectively;

[0031] h) SEQ ID NOs: 776 and 811, respectively;

[0032] i) SEQ ID NOs: 777 and 812, respectively;

[0033] j) SEQ ID NOs: 778 and 813, respectively;

[0034] k) SEQ ID NOs: 779 and 814, respectively;

[0035] 1) are SEQ ID NOs: 780 and 815, respectively;

[0036] m) are SEQ ID NOs: 781 and 816, respectively;

[0037] n) are SEQ ID NOs: 782 and 817, respectively;

[0038] o) are SEQ ID NOs: 783 and 818, respectively;

[0039] p) are SEQ ID NOs: 784 and 819, respectively;

[0040] q) are SEQ ID NOs: 785 and 820, respectively;

[0041] r) are SEQ ID NOs: 786 and 821, respectively;

[0042] s) are SEQ ID NOs: 787 and 822, respectively;

[0043] t) are SEQ ID NOs: 788 and 823, respectively;

[0044] u) are SEQ ID NOs: 789 and 824, respectively;

[0045] v) SEQ ID NOs: 790 and 825, respectively;

[0046] w) are SEQ ID NOs: 791 and 826, respectively;

[0047] x) are SEQ ID NOs: 792 and 827, respectively;

[0048] y) are SEQ ID NOs: 793 and 828, respectively;

[0049] z) are SEQ ID NOs: 794 and 829, respectively;

[0050] aa) are SEQ ID NOs: 795 and 830, respectively;

[0051] bb) are SEQ ID NOs: 796 and 831, respectively;

[0052] cc) are SEQ ID NOs: 797 and 832, respectively;

[0053] dd) are SEQ ID NOs: 798 and 833, respectively;

[0054] ee) are SEQ ID NOs: 799 and 834, respectively;

[0055] ff) are SEQ ID NOs: 800 and 835, respectively;

[0056] gg) are SEQ ID NOs: 801 and 836, respectively;

[0057] hh) are SEQ ID NOs: 802 and 837, respectively;

[0058] ii) SEQ ID NOs: 803 and 838, respectively; and

[0059] jj) are SEQ ID NOs: 1681 and 815, respectively.

[0060] In some aspects, the sense strand and the antisense strand comprise a nucleotide sequence selected from the group consisting of:

[0061] a) SEQ ID NO: 771 and 806, respectively;

[0062] b) SEQ ID NOs: 776 and 811, respectively;

[0063] c) SEQ ID NOs: 780 and 815, respectively;

[0064] d) SEQ ID NOs: 781 and 816, respectively;

[0065] e) SEQ ID NOs: 782 and 817, respectively;

[0066] f) SEQ ID NOs: 790 and 825, respectively;

[0067] g) SEQ ID NOs: 795 and 830, respectively;

[0068] h) SEQ ID NOs: 798 and 833, respectively;

[0069] i) SEQ ID NOs: 799 and 834, respectively;

[0070] j) SEQ ID NOs: 803 and 838, respectively;

[0071] k) are SEQ ID NOs: 1681 and 815, respectively.

[0072] In some aspects, the sense strand and the antisense strand comprise a nucleotide sequence selected from the group consisting of:

[0073] a) SEQ ID NO: 771 and 806, respectively;

[0074] b) SEQ ID NOs: 780 and 815, respectively;

[0075] c) SEQ ID NOs: 781 and 816, respectively;

[0076] d) SEQ ID NOs: 798 and 833, respectively;

[0077] e) SEQ ID NOs: 799 and 834, respectively;

[0078] f) SEQ ID NOs: 803 and 838, respectively; and

[0079] g) are SEQ ID NOs: 1681 and 815, respectively.

[0080] In some aspects, the sense strand comprises the nucleotide sequence set forth in SEQ ID NO: 771, and the antisense strand comprises the nucleotide sequence set forth in SEQ ID NO: 806. In some aspects, the sense strand comprises the nucleotide sequence set forth in SEQ ID NO: 780, and the antisense strand comprises the nucleotide sequence set forth in SEQ ID NO: 815. In some aspects, the sense strand comprises the nucleotide sequence set forth in SEQ ID NO: 781, and the antisense strand comprises the nucleotide sequence set forth in SEQ ID NO: 816. In some aspects, the sense strand comprises the nucleotide sequence set forth in SEQ ID NO: 798, and the antisense strand comprises the nucleotide sequence set forth in SEQ ID NO: 833. In some aspects, the sense strand comprises the nucleotide sequence set forth in SEQ ID NO: 799, and the antisense strand comprises the nucleotide sequence set forth in SEQ ID NO: 834. In some aspects, the sense strand comprises the nucleotide sequence set forth in SEQ ID NO: 803, and the antisense strand comprises the nucleotide sequence set forth in SEQ ID NO: 838. In some aspects, the sense strand comprises the nucleotide sequence set forth in SEQ ID NO:1681, and the antisense strand comprises the nucleotide sequence set forth in SEQ ID NO:815.

[0081] In any of the foregoing or related aspects, the antisense strand is 22 nucleotides in length. In some aspects, the antisense strand comprises a nucleotide sequence selected from SEQ ID NOs: 806, 815, 816, 833, 834, and 838. In some aspects, the sense strand is 36 nucleotides in length. In some aspects, the sense strand comprises a nucleotide sequence selected from SEQ ID NOs: 1130, 1095, 1096, 1119, 1120, and 1124. In some aspects, the sense strand comprises a nucleotide sequence selected from SEQ ID NOs: 771, 780, 781, 798, 799, and 803.

[0082] In any of the foregoing or related aspects, the sense strand comprises the nucleotide sequence of any one of SEQ ID NOs: 839-873 and 1682. In some aspects, the antisense strand comprises the nucleotide sequence of any one of SEQ ID NOs: 874-908.

[0083] In some aspects, the sense strand and the antisense strand comprise a nucleotide sequence selected from the group consisting of:

[0084] a) SEQ ID NO: 839 and 874, respectively;

[0085] b) SEQ ID NOs: 840 and 875, respectively;

[0086] c) SEQ ID NOs: 841 and 876, respectively;

[0087] d) SEQ ID NOs: 842 and 877, respectively;

[0088] e) SEQ ID NOs: 843 and 878, respectively;

[0089] f) SEQ ID NOs: 844 and 879, respectively;

[0090] g) SEQ ID NOs: 845 and 880, respectively;

[0091] h) SEQ ID NOs: 846 and 881, respectively;

[0092] i) SEQ ID NOs: 847 and 882, respectively;

[0093] j) SEQ ID NOs: 848 and 883, respectively;

[0094] k) SEQ ID NOs: 849 and 884, respectively;

[0095] 1) are SEQ ID NOs: 850 and 885, respectively;

[0096] m) are SEQ ID NOs: 851 and 886, respectively;

[0097] n) are SEQ ID NOs: 852 and 887, respectively;

[0098] o) are SEQ ID NOs: 853 and 888, respectively;

[0099] p) are SEQ ID NOs: 854 and 889, respectively;

[0100] q) are SEQ ID NOs: 855 and 890, respectively;

[0101] r) are SEQ ID NOs: 856 and 891, respectively;

[0102] s) are SEQ ID NOs: 857 and 892, respectively;

[0103] t) are SEQ ID NOs: 858 and 893, respectively;

[0104] u) are SEQ ID NOs: 859 and 894, respectively;

[0105] v) SEQ ID NOs: 860 and 895, respectively;

[0106] w) are SEQ ID NOs: 861 and 896, respectively;

[0107] x) are SEQ ID NOs: 862 and 897, respectively;

[0108] y) are SEQ ID NOs: 863 and 898, respectively;

[0109] z) are SEQ ID NOs: 864 and 899, respectively;

[0110] aa) are SEQ ID NOs: 865 and 900, respectively;

[0111] bb) are SEQ ID NOs: 866 and 901, respectively;

[0112] cc) are SEQ ID NOs: 867 and 902, respectively;

[0113] dd) are SEQ ID NOs: 868 and 903, respectively;

[0114] ee) are SEQ ID NOs: 869 and 904, respectively;

[0115] ff) are SEQ ID NOs: 870 and 905, respectively;

[0116] gg) are SEQ ID NOs: 871 and 906, respectively;

[0117] hh) are SEQ ID NOs: 872 and 907, respectively;

[0118] ii) SEQ ID NO: 873 and 908, respectively; and

[0119] jj) are SEQ ID NOs: 1682 and 885, respectively.

[0120] In other aspects, the sense strand and the antisense strand comprise a nucleotide sequence selected from the group consisting of:

[0121] a) SEQ ID NO: 860 and 895, respectively;

[0122] b) SEQ ID NO: 865 and 900, respectively;

[0123] c) SEQ ID NOs: 868 and 903, respectively;

[0124] d) SEQ ID NO: 869 and 904, respectively;

[0125] e) SEQ ID NOs: 873 and 908, respectively;

[0126] f) SEQ ID NOs: 841 and 876, respectively;

[0127] g) SEQ ID NOs: 846 and 881, respectively;

[0128] h) SEQ ID NOs: 850 and 885, respectively;

[0129] i) SEQ ID NOs: 851 and 886, respectively;

[0130] j) SEQ ID NOs: 852 and 887, respectively; and

[0131] k) are SEQ ID NOs: 1682 and 885, respectively.

[0132] In certain aspects, the sense strand and the antisense strand comprise a nucleotide sequence selected from the group consisting of:

[0133] a) SEQ ID NO: 841 and 876, respectively;

[0134] b) SEQ ID NOs: 850 and 885, respectively;

[0135] c) SEQ ID NOs: 851 and 886, respectively;

[0136] d) SEQ ID NO: 868 and 903, respectively;

[0137] e) SEQ ID NOs: 869 and 904, respectively;

[0138] f) SEQ ID NOs: 873 and 908, respectively; and

[0139] g) are SEQ ID NOs: 1682 and 885, respectively.

[0140] In some aspects, the sense strand comprises the nucleotide sequence set forth in SEQ ID NO: 841, and the antisense strand comprises the nucleotide sequence set forth in SEQ ID NO: 876. In some aspects, the sense strand comprises the nucleotide sequence set forth in SEQ ID NO: 850, and the antisense strand comprises the nucleotide sequence set forth in SEQ ID NO: 885. In some aspects, the sense strand comprises the nucleotide sequence set forth in SEQ ID NO: 851, and the antisense strand comprises the nucleotide sequence set forth in SEQ ID NO: 886. In some aspects, the sense strand comprises the nucleotide sequence set forth in SEQ ID NO: 868, and the antisense strand comprises the nucleotide sequence set forth in SEQ ID NO: 903. In some aspects, the sense strand comprises the nucleotide sequence set forth in SEQ ID NO: 869, and the antisense strand comprises the nucleotide sequence set forth in SEQ ID NO: 904. In some aspects, the sense strand comprises the nucleotide sequence set forth in SEQ ID NO: 873, and the antisense strand comprises the nucleotide sequence set forth in SEQ ID NO: 908. In some aspects, the sense strand comprises the nucleotide sequence set forth in SEQ ID NO:1682, and the antisense strand comprises the nucleotide sequence set forth in SEQ ID NO:885.

[0141] In some aspects, the sense strand comprises 5'-[mAs][mG][fA][mG][fU][mG][mU][fG][mG][fA][mA][fA][mA][fA][mA][mG][mA][mG][mC][mA][mG][mC][mG][ademA-GalNAc][ademA-GalNAc][ademAGalNAc][mG][mG][mC][mU][mG][mC]-3' (SEQ ID NO:841) and all modifications, and wherein the antisense strand comprises the sequence of 5'-[Me phosphonate-4O-mUs][fCs][fU][fU][fU][mU][fU][mU][fU][mU][mC][mC][fA][mC][fA][mC][mU][fC][mUs][mGs][mG]-3' (SEQ ID NO:876) and all modifications, wherein mC, mA, mG, mU = 2'-OMe ribonucleoside; fA, fC, fG, fU = 2'-F ribonucleoside; s = phosphorothioate, and wherein ademA-GalNAc =

[0142]

[0143] In some aspects, the sense strand comprises 5'-[mCs][mA][fG][mG][fU][mG][mG][fA][mA][fG][mU][fA][fA][mA][fA][mU][fC][mU][mG][mA][mG][mC][mA][mG][mC][mG][ademA-GalNAc][ademA-GalNAc][ademA-GalNAc][mG][mG][mC][mU][mG][mC]-3' (SEQ ID NO:850) and all modifications, and wherein the antisense strand comprises the sequence of 5'-[Me phosphonate-4O-mUs][fCs][fA][fG][fA][mU][fU][mU][mU][fA][mC][mU][mU][fC][mC][fA][mC][mC][fU][mGs][mGs][mG]-3' (SEQ ID NO:885) and all modifications, wherein mC, mA, mG, mU = 2'-OMe ribonucleoside; fA, fC, fG, fU = 2'-F ribonucleoside; s = phosphorothioate, and wherein ademA-GalNAc =

[0144]

[0145] In some aspects, the sense strand comprises 5'-[mAs][mG][fG][mU][fG][mG][mA][fA][mG][fU][mA][fA][fA][mA][fU][mC][fU][mG][mA][mA][mG][mC][mA][mG][mC][mC][mG][ademA-GalNAc][ademA-GalNAc][ademA-GalNAc][mG][mG][mC][mU][mG][mC]-3' (SEQ ID NO:851) and all modifications, and wherein the antisense strand comprises the sequence of 5'-[Me phosphonate-4O-mUs][fUs][fC][fA][fG][mA][fU][mU][mU][fU][mA][mC][mU][fU][mC][fC][mA][mC][fC][mUs][mGs][mG]-3' (SEQ ID NO:886) and all modifications, wherein mC, mA, mG, mU=2'-OMe ribonucleoside; fA, fC, fG, fU=2'-F ribonucleoside; s=phosphorothioate, and wherein ademA-GalNAc=

[0146]

[0147] In some aspects, the sense strand comprises 5'-[mAs][mG][fG][mA][fA][mA][mU][fA][mA][fA][mA][fA][fG][mA][fU][mU][fG][mA][mA][mA][mG][mC][mA][mG][mC][mG][ademA-GalNAc][ademA-GalNAc][ademA-GalNAc][mG][mG][mC][mU][mG][mC]-3' (SEQ ID NO:868) and all modifications, and wherein the antisense strand comprises the sequence of 5'-[Me phosphonate-4O-mUs][fUs][fU][fC][fA][mA][fU][mC][mU][fU][mU][mU][mU][fA][mU][fU][mU][mC][fC][mUs][mGs][mG]-3' (SEQ ID NO:903) and all modifications, wherein mC, mA, mG, mU = 2'-OMe ribonucleoside; fA, fC, fG, fU = 2'-F ribonucleoside; s = phosphorothioate, and wherein ademA-GalNAc =

[0148]

[0149] In some aspects, the sense strand comprises 5'-[mGs][mG][fA][mA][fA][mU][mA][fA][mA][fA][mA][fG][fA][mU][fU][mG][fA][mA][mA][mA][mG][mC][mA][mG][mC][mC][mG][ademA-GalNAc][ademA-GalNAc][ademA-GalNAc][mG][mG][mC][mU][mG][mC]-3' (SEQ ID NO:869) and all modifications, and wherein the antisense strand comprises the sequence of 5'-[Me phosphonate-4O-mUs][fUs][fU][fU][fC][mA][fA][mU][mC][fU][mU][mU][mU][fU][mA][fU][mU][mU][fC][mCs][mGs][mG]-3' (SEQ ID NO:904) and all modifications, wherein mC, mA, mG, mU = 2'-OMe ribonucleoside; fA, fC, fG, fU = 2'-F ribonucleoside; s = phosphorothioate, and wherein ademA-GalNAc =

[0150]

[0151] In some aspects, the sense strand comprises 5'-[mAs][mU][fA][mA][fA][mA][mA][fG][mA][fU][mU][fG][fA][mA][fA][mC][fC][mC][mA][mA][mG][mC][mC][mG][ademA-GalNAc][ademA-GalNAc][ademA-GalNAc][mG][mG][mC][mU][mG][mC]-3' (SEQ ID NO:873) and all modifications, and wherein the antisense strand comprises the sequence of 5'-[Me phosphonate-4O-mUs][fUs][fG][fG][fG][mU][fU][mU][mC][fA][mA][mU][mC][fU][mU][fU][mU][mU][fA][mUs][mGs][mG]-3' (SEQ ID NO:908) and all modifications, wherein mC, mA, mG, mU = 2'-OMe ribonucleoside; fA, fC, fG, fU = 2'-F ribonucleoside; s = phosphorothioate, and wherein ademA-GalNAc =

[0152]

[0153] In some aspects, the sense strand comprises 5'-[ademCs-C 16 ][mA][fG][mG][fU][mG][mG][fA][mA][fG][mU][fA][fA][mA][mU][fC][mUs][mGs][mA]-3' (SEQ ID NO: 1682) and all modifications, and wherein the antisense strand comprises the sequence of 5'-[Me phosphonate-4O-mUs][fCs][fA][fG][fA][mU][fU][mU][fA][mC][mU][mU][fC][mC][fA][mC][mC][fU][mGs][mGs][mG]-3' (SEQ ID NO: 885) and all modifications, wherein mC, mA, mG, mU = 2'-OMe ribonucleoside; fA, fC, fG, fU = 2'-F ribonucleoside; s = phosphorothioate, and [Cs-C][fA][fG][fA][mU][fU][mU][mU][fA][mC][mU][mU][fC][mC][fA][mC][mC][fU][mGs][mGs][mG]-3' 16 ] = conjugated to C 16 Cytosine in hydrocarbon chains:

[0154]

[0155] In some aspects, the present disclosure provides a pharmaceutical composition comprising an RNAi oligonucleotide described herein and a pharmaceutically acceptable carrier, delivery agent, or excipient.

[0156] In other aspects, the present disclosure provides a method for treating a subject having a disease, disorder or condition associated with MAPT gene expression, the method comprising administering to the subject a therapeutically effective amount of an RNAi oligonucleotide described herein or a pharmaceutical composition thereof, thereby treating the subject.

[0157] In other aspects, the present disclosure provides a method of delivering an oligonucleotide to a subject, the method comprising administering to the subject a pharmaceutical composition described herein.

[0158] In other aspects, the present disclosure provides a method for reducing MAPT gene expression in a cell, a cell population or a subject, the method comprising the steps of:

[0159] i. contacting the cell or the cell population with an RNAi oligonucleotide or a pharmaceutical composition described herein; or

[0160] ii. administering to the subject an RNAi oligonucleotide or pharmaceutical composition described herein.

[0161] In some aspects, reducing MAPT gene expression includes reducing the amount or level of MAPT mRNA, the amount or level of Tau protein or both. In some aspects, RNAi oligonucleotides described herein or pharmaceutical composition, experimenter suffers from the disease, disease or illness relevant to MAPT gene expression. In some aspects, the disease, disease or illness relevant to MAPT gene expression is AD, frontotemporal dementia (FTD), PSP, PD, Tau protein related diseases, primary age-related tauopathy, chronic traumatic encephalopathy, cortical basal ganglia degeneration, Lytico-bodig disease, ganglioglioma, meningioangiomatosis, postencephalitic parkinsonism or subacute sclerosing panencephalitis.

[0162] In any one of the aforementioned or related aspects, MAPT gene expression reduces in the tissue of one or more CNS regions, wherein the tissue is relevant to AD. In some aspects, the tissue relevant to AD is selected from: prefrontal cortex, motor cortex, temporal cortex, parietal cortex and hippocampus. In some aspects, MAPT gene expression reduces in the tissue of one or more CNS regions, wherein the tissue is relevant to PSP. In some aspects, the tissue relevant to PSPy is selected from: caudate nucleus, globus pallidus, thalamus, midbrain tegmentum, substantia nigra, pons, cerebellar white matter, cerebellar dentate nucleus, medulla, cervical spinal cord, thoracic spinal cord and lumbar spinal cord. In some aspects, MAPT gene expression is reduced in one or more CNS regions selected from the group consisting of: cervical spinal cord, thoracic spinal cord, lumbar spinal cord, frontal cortex, temporal cortex, cerebellum, midbrain, occipital cortex, parietal cortex, hippocampus, caudate nucleus, thalamus, brainstem, motor cortex, globus pallidus, midbrain tegmentum, substantia nigra, pons, cerebellar white matter, and cerebellar dentate nucleus.

[0163] In any of the foregoing or related aspects, the RNAi oligonucleotide or pharmaceutical composition is administered in combination with a second composition or therapeutic agent.

[0164] In other aspects, the present disclosure provides uses of the RNAi oligonucleotides or pharmaceutical compositions described herein for the manufacture of a medicament for treating a disease, disorder or condition associated with MAPT gene expression.

[0165] In other aspects, the present disclosure provides the RNAi oligonucleotides or pharmaceutical compositions described herein, which are used or suitable for treating a disease, disorder or condition associated with MAPT gene expression.

[0166] In some aspects, the present disclosure provides a kit comprising an RNAi oligonucleotide described herein, an optional pharmaceutically acceptable carrier, and a package insert comprising instructions for administration to a subject suffering from a disease, disorder, or condition associated with expression of a MAPT gene.

[0167] In any of the foregoing or related aspects, the disease, disorder or condition associated with MAPT gene expression is AD, FTD, PD, PSP, Tau protein-related disease, primary age-related tauopathy, chronic traumatic encephalopathy, cortical basal degeneration, Littiker-Bertiger disease, ganglioglioma, meningioangiomatosis, postencephalitis Parkinson's disease or subacute sclerosing panencephalitis. BRIEF DESCRIPTION OF THE DRAWINGS

[0168] Advantages, effects, features and objects other than those set forth above will become more apparent when considering the following detailed description. Such detailed description refers to the following drawings, wherein:

[0169] Figure 1A and Figure 1B Provided is a graph depicting the percentage (%) of human MAPT mRNA remaining in the liver of mice (hydrodynamic injection model) that exogenously express human MAPT after treatment with a MAPT oligonucleotide conjugated with GalNAc specific to human (Hs) MAPT or human and NHP (Hs-Mf; "double universal") MAPT. 3mg / kg of the designated GalNAc-conjugated MAPT targeting oligonucleotides prepared in PBS were subcutaneously administered to CD-1 mice. Four days after administration, DNA plasmids encoding human MAPT were hydrodynamically injected (HDI) into mice. The level of human MAPT mRNA was determined from the collected liver 18 hours later.

[0170] Figure 2A and Figure 2B Provides a description based on Figure 1A-1B Figure 2 is a graph of the inhibitory efficacy of selected GalNAc-conjugated MAPT targeting oligonucleotides and the dose response of GalNAc-conjugated MAPT targeting oligonucleotides specific for human (Hs), NHP (Mf) and murine (Mm) MAPT. Figure 1A-1B The percentage (%) of MAPT mRNA remaining in liver tissue was measured in CD-1 HDI mice as described in . The percentage (%) of mRNA remaining was determined in two groups after injection of 0.3 mg / kg, 1.0 mg / kg or 3.0 mg / kg of the specified GalNAc-conjugated MAPT targeting oligonucleotides, Figure 2A (Group I) and Figure 2B (Group II) Hs = construct is specific for human MAPT; Hs-Mf = construct is specific for human and monkey MAPT.

[0171] Figures 3A-3M Provided is a graph depicting the percentage (%) of human MAPT mRNA remaining in the CNS of NHPs after treatment with a GalNAc-conjugated MAPT targeting oligonucleotide. On day 0 and day 7 of the study, 50 mg of a specified GalNAc-conjugated MAPT targeting oligonucleotide prepared in artificial cerebrospinal fluid (aCSF) was injected into the cisterna magna (icm) to NHPs for administration. The percentage (%) of MAPT mRNA remaining in animals treated with aCSF was determined to determine the level of MAPT mRNA. The CNS tissues measured included the cervical spinal cord ( Figure 3A )、thoracic spinal cord( Figure 3B )、lumbar spinal cord( Figure 3C ), frontal cortex ( Figure 3D ), temporal cortex ( Figure 3E ), cerebellum( Figure 3F)、midbrain( Figure 3G ), occipital cortex( Figure 3H ), parietal cortex ( Fig. 3I ), hippocampus( Figure 3J ), caudate nucleus ( Figure 3K ),thalamus( Figure 3L ) and brainstem ( Figure 3M ).

[0172] Figure 4A-4B Lipid-conjugated RNAi oligonucleotides are provided ( Figure 4A ) and GalNAc-conjugated RNAi oligonucleotides ( Figure 4B ) schematic diagram.

[0173] Figure 5A-5B Provides a graph depicting the percentage (%) of NHP(Mf)MAPT mRNA remaining in CNS tissues associated with AD ( Figure 5A ) and the concentration of oligonucleotides ( Figure 5B ). Intrathecal administration of C-conjugated to NHP 16 MAPT-2357 with lipid or GalNAc, such as Figure 4A-4B Tissues were collected and analyzed 28 days after administration of the indicated oligonucleotides.

[0174] Figure 6A-6B Provided are graphs depicting the percentage (%) of NHP(Mf)MAPT mRNA remaining in CNS tissues associated with PSP ( Fig. 6A ) and the concentration of oligonucleotides ( Figure 6B ). Intrathecal administration of C-conjugated to NHP 16 MAPT-2357 with lipid or GalNAc, such as Figure 4A-4B Tissues were collected and analyzed 28 days after administration of the indicated oligonucleotides. DETAILED DESCRIPTION

[0175] According to some aspects, the disclosure provides oligonucleotides, such as RNAi oligonucleotides, that reduce the MAPT gene expression in CNS. In some embodiments, the oligonucleotides provided herein are designed to treat the disease relevant to the MAPT gene expression in CNS. In some aspects, the disclosure provides the method for treating the disease relevant to MAPT by reducing the MAPT gene expression in cells (e.g., cells of CNS).

[0176] Oligonucleotide inhibitors of MAPT gene expression

[0177] The present disclosure provides, inter alia, oligonucleotides (eg, RNAi oligonucleotides) that inhibit the expression of the MAPT gene. In some embodiments, the oligonucleotides that inhibit the expression of the MAPT gene target the MAPT mRNA.

[0178] MAPT target sequences

[0179] In some embodiments, the oligonucleotides herein (eg, RNAi oligonucleotides) target a target sequence comprising MAPT mRNA. In some embodiments, the oligonucleotides target a target sequence within the MAPT mRNA sequence.

[0180] In some embodiments, the oligonucleotide corresponds to a target sequence within the MAPT mRNA sequence. In some embodiments, the oligonucleotide or a portion, fragment or strand thereof (e.g., an antisense strand or guide strand of an RNAi oligonucleotide) is combined or bonded to a target sequence comprising MAPT mRNA, thereby inhibiting MAPT gene expression.

[0181] In some embodiments, the oligonucleotide targeting MAPT target sequence is used for the purpose of suppressing MAPT gene expression in vivo. In some embodiments, the amount or degree of the oligonucleotide targeting MAPT target sequence suppressing MAPT gene expression is associated with the effectiveness of the oligonucleotide. In some embodiments, the amount or degree of the oligonucleotide targeting MAPT target sequence suppressing MAPT gene expression is associated with the amount or degree of the therapeutic benefit in the subject or patient suffering from the disease, disorder or illness related to MAPT gene expression treated with the oligonucleotide.

[0182] In some embodiments, the sense strand of oligonucleotide comprises MAPT target sequence.In some embodiments, a part or region of the sense strand of oligonucleotide (e.g., RNAi oligonucleotide) comprises MAPT target sequence.In some embodiments, MAPT target sequence comprises the nucleotide sequence of any one of SEQ ID NO:912-1295 or consists of it. In some embodiments, the MAPT target sequence comprises or consists of the nucleotide sequence of any one of SEQ ID NOs: 1125, 1127, 1130, 1019, 1031, 1044, 1064, 1065, 1067, 1083, 915, 1095, 1096, 1102, 1110, 923, 925, 1025, 1039, 1049, 1061, 1070, 1072, 1075, 1081, 1108, 1111, 1114, 1119, 1120, 1121, 1122, 1123, 1124, and 924. In some embodiments, the MAPT target sequence comprises or consists of the nucleotide sequence of any one of SEQ ID NOs: 1061, 1108, 1119, 1120, 1124, 1130, 1065, 1095, 1096, and 1102. In some embodiments, the MAPT target sequence comprises or consists of the nucleotide sequence of any one of SEQ ID NOs: 1130, 1095, 1096, 1119, 1120, and 1124. In some embodiments, the MAPT target sequence comprises the nucleotide sequence set forth in SEQ ID NO: 1130. In some embodiments, the MAPT target sequence comprises the nucleotide sequence set forth in SEQ ID NO: 1095. In some embodiments, the MAPT target sequence comprises the nucleotide sequence set forth in SEQ ID NO: 1096. In some embodiments, the MAPT target sequence comprises the nucleotide sequence set forth in SEQ ID NO: 1119. In some embodiments, the MAPT target sequence comprises the nucleotide sequence set forth in SEQ ID NO: 1120. In some embodiments, the MAPT target sequence comprises the nucleotide sequence shown in SEQ ID NO:1124.

[0183] MAPT mRNA targeting sequences

[0184] In some embodiments, the oligonucleotide herein (e.g., RNAi oligonucleotide) has a complementary region of MAPT mRNA (e.g., within the target sequence of MAPT mRNA) for the purpose of targeting the mRNA in the cell and suppressing its expression. In some embodiments, the oligonucleotide comprises a MAPT mRNA target sequence (e.g., a ds oligonucleotide, such as the antisense strand or guide strand of an RNAi oligonucleotide) with a complementary region, which is bound or bonded to the MAPT target sequence by complementary (Watson-Crick (Watson-Crick)) base pairing. Targeting sequence or complementary region generally have a suitable length and base content, so that the oligonucleotide (or its chain) can be bound or bonded to the MAPT mRNA for the purpose of suppressing its expression. In some embodiments, the length of the targeting sequence or complementary region is at least about 12, at least about 13, at least about 14, at least about 15, at least about 16, at least about 17, at least about 18, at least about 19, at least about 20, at least about 21, at least about 22, at least about 23, at least about 24, at least about 25, at least about 26, at least about 27, at least about 28, at least about 29, or at least about 30 nucleotides. In some embodiments, the targeting sequence or complementary region is at least 12, at least 13, at least 14, at least 15, at least 16, at least 17, at least 18, at least 19, or at least 20 nucleotides. In some embodiments, the length of the targeting sequence or complementary region is about 12 to about 30 (e.g., 12 to 30, 12 to 22, 15 to 25, 17 to 21, 18 to 27, 19 to 27, or 15 to 30) nucleotides. In some embodiments, the length of the targeting sequence or complementary region is about 12, 13, 14, 15, 16, 17, 18, 19, 20, 21, 22, 23, 24, 25, 26, 27, 28, 29, or 30 nucleotides. In some embodiments, the length of the targeting sequence or complementary region is 18 nucleotides. In some embodiments, the length of the targeting sequence or complementary region is 19 nucleotides. In some embodiments, the length of the targeting sequence or complementary region is 20 nucleotides. In some embodiments, the length of the targeting sequence or complementary region is 21 nucleotides. In some embodiments, the length of the targeting sequence or complementary region is 22 nucleotides. In some embodiments, the length of the targeting sequence or complementary region is 23 nucleotides. In some embodiments, the length of the targeting sequence or complementary region is 24 nucleotides. In some embodiments, the oligonucleotide comprises a target sequence or complementary region that is complementary to the sequence of any one of SEQ ID NOs: 912-1295, and the length of the targeting sequence or complementary region is 18 nucleotides.In some embodiments, the oligonucleotide comprises a target sequence or complementary region complementary to a sequence of any one of SEQ ID NO:912-1295, and the length of the targeted sequence or complementary region is 19 nucleotides. In some embodiments, the oligonucleotide comprises a target sequence or complementary region complementary to a sequence of any one of SEQ ID NO:1-384, and the length of the targeted sequence or complementary region is 20 nucleotides. In some embodiments, the oligonucleotide comprises a target sequence or complementary region complementary to a sequence of any one of SEQ ID NO:1-384, and the length of the targeted sequence or complementary region is 21 nucleotides. In some embodiments, the oligonucleotide comprises a target sequence or complementary region complementary to a sequence of any one of SEQ ID NO:1-384, and the length of the targeted sequence or complementary region is 22 nucleotides. In some embodiments, the oligonucleotide comprises a target sequence or complementary region complementary to a sequence of any one of SEQ ID NO:1-384, and the length of the targeted sequence or complementary region is 23 nucleotides. In some embodiments, the oligonucleotide comprises a target sequence or a complementary region that is complementary to the sequence of any one of SEQ ID NOs: 1-384, and the targeting sequence or complementary region is 24 nucleotides in length.

[0185] In some embodiments, oligonucleotide comprises a targeting sequence or a complementary region (e.g., an antisense strand or a guide strand of a ds oligonucleotide) that is fully complementary to the MAPT mRNA target sequence. In some embodiments, the targeting sequence or the complementary region are partially complementary to the MAPT mRNA target sequence. In some embodiments, the oligonucleotide comprises a targeting sequence or a complementary region that is fully complementary to the sequence of any one of SEQ ID NO:912-1295. In some embodiments, the oligonucleotide comprises a targeting sequence or a complementary region that is partially complementary to the sequence of any one of SEQ ID NO:912-1295. In some embodiments, the oligonucleotide comprises a targeting sequence or a complementary region that is fully complementary to the sequence of any one of SEQ ID NO: 1125, 1127, 1130, 1019, 1031, 1044, 1064, 1065, 1067, 1083, 915, 1095, 1096, 1102, 1110, 923, 925, 1025, 1039, 1049, 1061, 1070, 1072, 1075, 1081, 1108, 1111, 1114, 1119, 1120, 1121, 1122, 1123, 1124, or 924. In some embodiments, the oligonucleotide comprises a targeting sequence or a complementary region that is complementary to a portion of the sequence of any one of SEQ ID NO: 1125, 1127, 1130, 1019, 1031, 1044, 1064, 1065, 1067, 1083, 915, 1095, 1096, 1102, 1110, 923, 925, 1025, 1039, 1049, 1061, 1070, 1072, 1075, 1081, 1108, 1111, 1114, 1119, 1120, 1121, 1122, 1123, 1124, or 924. In some embodiments, the oligonucleotide comprises a targeting sequence or a complementary region that is fully complementary to the sequence of any one of SEQ ID NOs: 1061, 1108, 1119, 1120, 1124, 1130, 1065, 1095, 1096, or 1102. In some embodiments, the oligonucleotide comprises a targeting sequence or a complementary region that is partially complementary to the sequence of any one of SEQ ID NOs: 1061, 1108, 1119, 1120, 1124, 1130, 1065, 1095, 1096, or 1102. In some embodiments, the oligonucleotide comprises a targeting sequence or a complementary region that is fully complementary to the sequence of any one of SEQ ID NOs: 1130, 1095, 1096, 1119, 1120, and 1124. In some embodiments, the oligonucleotide comprises a targeting sequence or a complementary region that is partially complementary to the sequence of any one of SEQ ID NOs: 1130, 1095, 1096, 1119, 1120, and 1124.In some embodiments, the oligonucleotide comprises a targeting sequence or a complementary region that is fully complementary to the sequence shown in SEQ ID NO: 1130. In some embodiments, the oligonucleotide comprises a targeting sequence or a complementary region that is fully complementary to the sequence shown in SEQ ID NO: 1095. In some embodiments, the oligonucleotide comprises a targeting sequence or a complementary region that is fully complementary to the sequence shown in SEQ ID NO: 1096. In some embodiments, the oligonucleotide comprises a targeting sequence or a complementary region that is fully complementary to the sequence shown in SEQ ID NO: 1119. In some embodiments, the oligonucleotide comprises a targeting sequence or a complementary region that is fully complementary to the sequence shown in SEQ ID NO: 1120. In some embodiments, the oligonucleotide comprises a targeting sequence or a complementary region that is fully complementary to the sequence shown in SEQ ID NO: 1124. In some embodiments, the oligonucleotide comprises a targeting sequence or a complementary region that is partially complementary to the sequence of any one of SEQ ID NO: 1130. In some embodiments, the oligonucleotide comprises a targeting sequence or a complementary region that is partially complementary to the sequence shown in SEQ ID NO: 1095. In some embodiments, the oligonucleotide comprises a targeting sequence or a complementary region that is partially complementary to the sequence shown in SEQ ID NO: 1096. In some embodiments, the oligonucleotide comprises a targeting sequence or a complementary region that is partially complementary to the sequence shown in SEQ ID NO: 1119. In some embodiments, the oligonucleotide comprises a targeting sequence or a complementary region that is partially complementary to the sequence shown in SEQ ID NO: 1120. In some embodiments, the oligonucleotide comprises a targeting sequence or a complementary region that is partially complementary to the sequence shown in SEQ ID NO: 1124.

[0186] In some embodiments, oligonucleotide comprises a targeting sequence or a complementary region complementary to a continuous nucleotide sequence comprising MAPT mRNA, wherein the length of the continuous nucleotide sequence is about 12 to about 30 nucleotides (e.g., a length of 12 to 30, 12 to 28, 12 to 26, 12 to 24, 12 to 20, 12 to 18, 12 to 16, 14 to 22, 16 to 20, 18 to 20 or 18 to 19 nucleotides). In some embodiments, oligonucleotide comprises a targeting sequence or a complementary region complementary to a continuous nucleotide sequence comprising MAPT mRNA, wherein the length of the continuous nucleotide sequence is 10, 11, 12, 13, 14, 15, 16, 17, 18, 19 or 20 nucleotides. In some embodiments, oligonucleotide comprises a targeting sequence or a complementary region complementary to a continuous nucleotide sequence comprising MAPT mRNA, wherein the length of the continuous nucleotide sequence is 19 nucleotides. In some embodiments, the oligonucleotide comprises a targeting sequence or a complementary region that is complementary to a contiguous nucleotide sequence comprising MAPT mRNA, wherein the contiguous nucleotide sequence is 20 nucleotides in length.

[0187] In some embodiments, the oligonucleotide comprises a targeting sequence or a complementary region that is complementary to a contiguous nucleotide sequence of any one of SEQ ID NOs: 912-1295, optionally wherein the contiguous nucleotide sequence is 19 nucleotides in length. In some embodiments, the oligonucleotide comprises a targeting sequence or a complementary region that is complementary to a contiguous nucleotide sequence of any one of SEQ ID NOs: 1125, 1127, 1130, 1019, 1031, 1044, 1064, 1065, 1067, 1083, 915, 1095, 1096, 1102, 1110, 923, 925, 1025, 1039, 1049, 1061, 1070, 1072, 1075, 1081, 1108, 1111, 1114, 1119, 1120, 1121, 1122, 1123, 1124, and 924, optionally wherein the contiguous nucleotide sequence is 19 nucleotides in length. In some embodiments, the oligonucleotide comprises a targeting sequence or a complementary region that is complementary to a contiguous nucleotide sequence of any one of SEQ ID NOs: 1061, 1108, 1119, 1120, 1124, 1130, 1065, 1095, 1096, and 1102, optionally wherein the contiguous nucleotide sequence is 19 nucleotides in length. In some embodiments, the oligonucleotide comprises a targeting sequence or a complementary region that is complementary to a contiguous nucleotide sequence of SEQ ID NOs: 1130, 1095, 1096, 1119, 1120, and 1124, optionally wherein the contiguous nucleotide sequence is 19 nucleotides in length.

[0188] In some embodiments, the targeting sequence or complementary region of the oligonucleotide is complementary to a continuous nucleotide sequence as shown in any one of SEQ ID NOs: 912-1295 and spans the entire length of the antisense strand. In some embodiments, the targeting sequence or complementary region of the oligonucleotide is complementary to a continuous nucleotide sequence as shown in any one of SEQ ID NOs: 912-1295 and spans a portion of the entire length of the antisense strand. In some embodiments, the oligonucleotide comprises a complementary region (e.g., completely) complementary to a continuous nucleotide stretch of nucleotides 1-20 spanning a sequence as shown in any one of SEQ ID NOs: 912-1295 (e.g., on the antisense strand of a ds oligonucleotide). In some embodiments, the targeting sequence or complementary region of the oligonucleotide is complementary to a continuous nucleotide sequence as shown in any one of SEQ ID NOs: 1-384 and spans the entire length of the antisense strand. In some embodiments, the complementary region of the oligonucleotide is complementary to a continuous nucleotide sequence as shown in any one of SEQ ID NOs: 1-384 and spans a portion of the entire length of the antisense strand. In some embodiments, the oligonucleotide comprises a complementary region (eg, on the antisense strand of the ds oligonucleotide) that is at least partially (eg, fully) complementary to a contiguous stretch of nucleotides spanning nucleotides 1-19 of a sequence as set forth in any one of SEQ ID NOs: 1-384.

[0189] In some embodiments, the oligonucleotide comprises a targeting sequence or complementary region with one or more base pairs (bp) mismatches with the corresponding MAPT mRNA target sequence. In some embodiments, the targeting sequence or complementary region can have up to about 1, up to about 2, up to about 3, up to about 4, up to about 5, etc. mismatches with the corresponding MAPT mRNA target sequence, provided that the targeting sequence or complementary region is combined or bonded to the ability of MAPT mRNA under appropriate hybridization conditions and / or the ability of the oligonucleotide to suppress MAPT gene expression is maintained. Or, in some embodiments, the targeting sequence or complementary region comprises no more than 1, no more than 2, no more than 3, no more than 4 or no more than 5 mismatches with the corresponding MAPT mRNA target sequence, provided that the targeting sequence or complementary region is combined or bonded to the ability of MAPT mRNA under appropriate hybridization conditions and / or the ability of the oligonucleotide to suppress MAPT gene expression is maintained. In some embodiments, the oligonucleotide comprises a targeting sequence or complementary region with 1 mismatches with the corresponding target sequence. In some embodiments, the oligonucleotide comprises a targeting sequence or complementary region with 2 mismatches with the corresponding target sequence. In some embodiments, the oligonucleotide comprises a targeting sequence or complementary region with 3 mismatches with the corresponding target sequence. In some embodiments, the oligonucleotide comprises a targeting sequence or complementary region with 4 mismatches with the corresponding target sequence. In some embodiments, the oligonucleotide comprises a targeting sequence or complementary region with 5 mismatches with the corresponding target sequence. In some embodiments, the oligonucleotide comprises a targeting sequence or complementary region with more than one mismatch (e.g., 2, 3, 4, 5 or more mismatches) with the corresponding target sequence, wherein at least 2 (e.g., all) mismatches are continuously positioned (e.g., 2, 3, 4, 5 or more mismatches in a row), or wherein mismatches are scattered throughout any position of the entire targeting sequence or complementary region. In some embodiments, an oligonucleotide comprises a targeting sequence or complementary region that has more than one mismatch (e.g., 2, 3, 4, 5 or more mismatches) with a corresponding target sequence, wherein at least 2 (e.g., all) of the mismatches are positioned consecutively (e.g., 2, 3, 4, 5 or more mismatches in a row), or wherein at least one or more non-mismatched base pairs are located between the mismatches, or a combination thereof.

[0190] In some embodiments, the oligonucleotide comprises a targeting sequence or a complementary region that is complementary to a contiguous nucleotide sequence of any one of SEQ ID NOs: 912-1295, wherein the targeting sequence or the complementary region may have at most about 1, at most about 2, at most about 3, at most about 4, at most about 5, etc. mismatches with the corresponding MAPT mRNA target sequence. In some embodiments, the oligonucleotide comprises a targeting sequence or a complementary region that is complementary to a contiguous nucleotide sequence of any one of SEQ ID NOs: 912-1295, wherein the targeting sequence or the complementary region may have no more than 1, no more than 2, no more than 3, no more than 4, or no more than 5 mismatches with the corresponding MAPT mRNA target sequence. In some embodiments, the oligonucleotide comprises a targeting sequence or complementary region that is complementary to a contiguous nucleotide sequence of any one of SEQ ID NOs: 1125, 1127, 1130, 1019, 1031, 1044, 1064, 1065, 1067, 1083, 915, 1095, 1096, 1102, 1110, 923, 925, 1025, 1039, 1049, 1061, 1070, 1072, 1075, 1081, 1108, 1111, 1114, 1119, 1120, 1121, 1122, 1123, 1124, and 924, wherein the targeting sequence or complementary region may have at most about 1, at most about 2, at most about 3, at most about 4, at most about 5, etc. mismatches with the corresponding MAPT mRNA target sequence. In some embodiments, the oligonucleotide comprises a targeting sequence or complementary region that is complementary to a contiguous nucleotide sequence of any one of SEQ ID NOs: 1125, 1127, 1130, 1019, 1031, 1044, 1064, 1065, 1067, 1083, 915, 1095, 1096, 1102, 1110, 923, 925, 1025, 1039, 1049, 1061, 1070, 1072, 1075, 1081, 1108, 1111, 1114, 1119, 1120, 1121, 1122, 1123, 1124, and 924, wherein the targeting sequence or complementary region may have no more than 1, no more than 2, no more than 3, no more than 4, or no more than 5 mismatches with the corresponding MAPT mRNA target sequence. In some embodiments, the oligonucleotide comprises a targeting sequence or a complementary region that is complementary to a contiguous nucleotide sequence of any one of SEQ ID NOs: 1061, 1108, 1119, 1120, 1124, 1130, 1065, 1095, 1096, and 1102, wherein the targeting sequence or the complementary region may have at most about 1, at most about 2, at most about 3, at most about 4, at most about 5, etc. mismatches with the corresponding MAPT mRNA target sequence.In some embodiments, the oligonucleotide comprises a targeting sequence or complementary region that is complementary to a contiguous nucleotide sequence of any one of SEQ ID NOs: 1061, 1108, 1119, 1120, 1124, 1130, 1065, 1095, 1096, and 1102, wherein the targeting sequence or complementary region may have no more than 1, no more than 2, no more than 3, no more than 4, or no more than 5 mismatches with the corresponding MAPT mRNA target sequence. In some embodiments, the oligonucleotide comprises a targeting sequence or complementary region that is complementary to a contiguous nucleotide sequence of any one of SEQ ID NOs: 1130, 1095, 1096, 1119, 1120, and 1124, wherein the targeting sequence or complementary region may have at most about 1, at most about 2, at most about 3, at most about 4, at most about 5, etc. mismatches with the corresponding MAPT mRNA target sequence. In some embodiments, the oligonucleotide comprises a targeting sequence or a complementary region that is complementary to a contiguous nucleotide sequence of any one of SEQ ID NOs: 1130, 1095, 1096, 1119, 1120, and 1124, wherein the targeting sequence or the complementary region may have no more than 1, no more than 2, no more than 3, no more than 4, or no more than 5 mismatches with the corresponding MAPT mRNA target sequence. In some embodiments, the oligonucleotide comprises a targeting sequence or a complementary region that is complementary to a contiguous nucleotide sequence of SEQ ID NO: 1095, wherein the targeting sequence or the complementary region may have at most about 1, at most about 2, at most about 3, at most about 4, at most about 5, etc. mismatches with the corresponding MAPT mRNA target sequence. In some embodiments, the oligonucleotide comprises a targeting sequence or a complementary region that is complementary to a contiguous nucleotide sequence of SEQ ID NO: 1095, wherein the targeting sequence or the complementary region may have no more than 1, no more than 2, no more than 3, no more than 4, or no more than 5 mismatches with the corresponding MAPT mRNA target sequence.

[0191] Types of Oligonucleotides

[0192] A variety of oligonucleotide types and / or structures can be used to target MAPT mRNA in the methods herein, including but not limited to RNAi oligonucleotides.Any oligonucleotide type described herein or elsewhere is contemplated as a framework to incorporate the MAPT mRNA targeting sequence herein for the purpose of inhibiting MAPT gene expression.

[0193] In some embodiments, the oligonucleotide herein suppresses MAPT gene expression by RNA interference (RNAi) pathway (e.g., RNAi oligonucleotide) in the upstream or downstream involved in Dicer. For example, RNAi oligonucleotides have been developed, wherein each chain has a size of about 19-25 nucleotides and at least one 3' overhang with about 1 to about 5 nucleotides (see, e.g., U.S. Patent No. 8,372,968). Longer oligonucleotides have also been developed, which are processed by Dicer to produce active RNAi products (see, e.g., U.S. Patent No. 8,883,996). Further work produces extended ds oligonucleotides, wherein at least one end of at least one chain extends beyond the duplex targeting region, including a structure in which one chain includes a thermodynamically stable tetraL structure (see, e.g., U.S. Patent No. 8,513,207 and 8,927,705, and International Patent Application Publication No. WO 2010 / 033225). Such structures can include single-stranded (ss) extensions (on one side or both sides of the molecule) and ds extensions.

[0194] In some embodiments, oligonucleotide participates in the RNAi pathway of the downstream of Dicer involved in (for example, Dicer cleavage).In some embodiments, oligonucleotide has overhang (for example, 1,2 or 3 nucleotides in length) at the 3' end of sense strand.In some embodiments, oligonucleotide comprises antisense strand and complementary sense strand of 21 nucleotides antisense to target mRNA (for example, MAPT mRNA), wherein two chains are bonded to form 19-bp duplex and overhang of 2 nucleotides at any one or two 3' ends.Also expected longer oligonucleotide design, including oligonucleotide with antisense strand of 23 nucleotides and sense strand of 21 nucleotides, wherein there is flat end on the right side of oligonucleotide (5' end of 3' end of sense strand / antisense strand) and there is 3' guide strand overhang of two nucleotides on the left side of oligonucleotide (5' end of sense strand / antisense strand).In such molecules, there is 21bp duplex region. See, e.g., U.S. Patent Nos. 9,012,138, 9,012,621, and 9,193,753.

[0195] In some embodiments, the oligonucleotide comprises a sense strand and an antisense strand both in the range of about 17 to about 36 (e.g., 17 to 26, 20 to 25, or 21-23) nucleotides in length. In some embodiments, the oligonucleotide comprises an antisense strand of 19-30 nucleotides in length and a sense strand of 19-50 nucleotides in length, wherein the antisense strand and the sense strand are independent strands forming an asymmetric duplex region, and the duplex region has an overhang of 1-4 nucleotides at the 3' end of the antisense strand. In some embodiments, the oligonucleotide comprises a sense strand and an antisense strand both in the range of about 19-22 nucleotides in length. In some embodiments, the sense strand and the antisense strand are equal in length. In some embodiments, the oligonucleotide comprises a sense strand and an antisense strand, so that a 3' overhang is present on either the sense strand or the antisense strand or on both the sense strand and the antisense strand. In some embodiments, for oligonucleotides having sense and antisense strands each in the range of about 21-23 nucleotides in length, the length of the 3' overhang on the sense strand, antisense strand, or both is 1 or 2 nucleotides. In some embodiments, the oligonucleotide has a guide strand of 22 nucleotides and a passenger strand of 20 nucleotides, with a blunt end on the right side of the molecule (3' end of the passenger strand / 5' end of the guide strand) and a 3' guide strand overhang of 2 nucleotides on the left side of the molecule (5' end of the passenger strand / 3' end of the guide strand). In such molecules, there is a 20 bp duplex region.

[0196] Other oligonucleotide designs for use in the compositions and methods herein include: 16-mer siRNA (see, e.g., NUCLEIC ACIDS IN CHEMISTRY AND BIOLOGY, Blackburn (ed.), Royal Society of Chemistry, 2006), shRNA (e.g., having a stem of 19 bp or less; see, e.g., Moore et al. (2010) METHODS MOL. BIOL. 629: 141-58), blunt siRNA (e.g., 19 bp in length; see, e.g., Kraynack and Baker (2006) RNA 12: 163-76), asymmetric siRNA (aiRNA; see, e.g., Sun et al. (2008) NAT. BIOTECHNOL. 26: 1379-82), asymmetric shorter duplex siRNA (see, e.g., Chang et al. (2009) MOL. THER. 17: 725-732), forked siRNA (see, e.g., Hohjoh (2004) FEBS LETT.557:193-98), single-stranded siRNA (Elsner (2012) NAT.BIOTECHNOL.30:1063), dumbbell-shaped circular siRNA (see, e.g., Abe et al. (2007) J.AM.CHEM.SOC.129:15108-09), and small internal segmented interfering RNA (siRNA; see, e.g., Bramsen et al. (2007) NUCLEIC ACIDS RES.35:5886-97). Other non-limiting examples of oligonucleotide structures that can be used to reduce or inhibit MAPT expression in some embodiments are microRNA (miRNA), short hairpin RNA (shRNA), and short siRNA (see, e.g., Hamilton et al. (2002) EMBO J.21:4671-79; see also U.S. Patent Application Publication No. 2009 / 0099115).

[0197] In addition, in some embodiments, the oligonucleotide used to reduce or inhibit the expression of MAPT gene herein is ss. Such structure may include but is not limited to ss RNAi molecules. Recent efforts have demonstrated the activity of ss RNAi molecules (see, for example, Matsui et al. (2016) Mol. Ther. 24: 946-955). However, in some embodiments, the oligonucleotide is an antisense oligonucleotide (ASO). ASO is an ss oligonucleotide with a core base sequence, when written or depicted in the 5' to 3' direction, the core base sequence comprises the reverse complement of the targeting segment of a specific nucleic acid and is appropriately modified (for example, as a gapmer) to induce the cleavage of its target RNA mediated by RNaseH in the cell or (for example, as a mixed polymer (mixer)) to inhibit the translation of target mRNA in the cell. ASO used herein can be modified in any suitable manner as known in the art, including, for example, shown in U.S. Patent No. 9,567,587 (including, for example, changes in the sugar moiety of the length, core base (pyrimidine, purine) and the heterocyclic moiety of the core base). In addition, ASOs have been used for decades to reduce the expression of specific target genes (see, e.g., Bennett et al. (2017) Annu. Rev. Pharmacol. 57:81-105).

[0198] In some embodiments, ASO shares complementary region with MAPT mRNA. In some embodiments, ASO targeting is identified as each region of the mankind MAPT of NM_001123066.3. In some embodiments, the length of ASO is about 15-50 nucleotides. In some embodiments, the length of ASO is about 15-25 nucleotides. In some embodiments, the length of ASO is 22 nucleotides. In some embodiments, ASO is complementary to any one of SEQ ID NO:912-1295. In some embodiments, the length of ASO is at least 15 continuous nucleotides. In some embodiments, the length of ASO is at least 19 continuous nucleotides. In some embodiments, the length of ASO is at least 20 continuous nucleotides. In some embodiments, ASO differs from target sequence by 1, 2 or 3 nucleotides.

[0199] Double-stranded RNAi oligonucleotides

[0200] In some aspects, the present disclosure provides ds RNAi oligonucleotides for targeting MAPT mRNA and suppressing MAPT gene expression (e.g., via RNAi pathway), the oligonucleotides comprising a sense strand (also referred to herein as a passenger strand) and an antisense strand (also referred to herein as a guide strand). In some embodiments, the sense strand and the antisense strand are independent strands and are not covalently linked. In some embodiments, the sense strand and the antisense strand are covalently linked. In some embodiments, the sense strand and the antisense strand form a duplex region, wherein the sense strand and the antisense strand or a part thereof are bound to each other in a complementary manner (e.g., by Watson-Crick base pairing).

[0201] In some embodiments, the sense strand has a first region (R1) and a second region (R2), wherein R2 comprises a first subregion (S1), L (e.g., tetraL or triL) and a second subregion (S2), wherein L is located between S1 and S2, and wherein S1 and S2 form a second duplex (D2). D2 can have various lengths. In some embodiments, the length of D2 is about 1 to about 6 bp. In some embodiments, the length of D2 is 2-6, 3-6, 4-6, 5-6, 1-5, 2-5, 3-5 or 4-5 bp. In some embodiments, the length of D2 is 1, 2, 3, 4, 5 or 6 bp. In some embodiments, the length of D2 is 6 bp.

[0202] In some embodiments, the R1 of the sense strand and the antisense strand form a first duplex (D1). In some embodiments, the length of D1 is at least about 15 (e.g., at least 15, at least 16, at least 17, at least 18, at least 19, at least 20, or at least 21) nucleotides. In some embodiments, the length of D1 is in the range of about 12 to about 30 nucleotides (e.g., 12 to 30, 12 to 27, 15 to 22, 18 to 22, 18 to 25, 18 to 27, 18 to 30, or 21 to 30 nucleotides in length). In some embodiments, the length of D1 is at least 12 nucleotides (e.g., at least 12, at least 15, at least 20, at least 25, or at least 30 nucleotides in length). In some embodiments, D1 is 12, 13, 14, 15, 16, 17, 18, 19, 20, 21, 22, 23, 24, 25, 26, 27, 28, 29 or 30 nucleotides in length. In some embodiments, D1 is 19 nucleotides in length. In some embodiments, D1 is 20 nucleotides in length. In some embodiments, D1 comprising a sense strand and an antisense strand does not span the entire length of the sense strand and / or antisense strand. In some embodiments, D1 comprising a sense strand and an antisense strand spans the entire length of either or both of the sense strand or antisense strand. In certain embodiments, D1 comprising a sense strand and an antisense strand spans the entire length of both the sense strand or the antisense strand.

[0203] In some embodiments, the sense strand described herein is 36 nucleotides in length and positions are numbered 1-36 from 5' to 3'. In some embodiments, the antisense strand described herein is 22 nucleotides in length and positions are numbered 1-22 from 5' to 3'. In some embodiments, the position numbering described herein follows this numbering format.

[0204] In some embodiments, the RNAi oligonucleotide comprises a sense strand having a sequence of any one of SEQ ID NOs: 1-384 and an antisense strand comprising a complementary sequence of any one of SEQ ID NOs: 385-768. In some embodiments, the RNAi oligonucleotide comprises a sense strand having a sequence of SEQ ID NOs: 912-1295 and an antisense strand comprising a complementary sequence of any one of SEQ ID NOs: 1296-1679.

[0205] In some embodiments, the RNAi oligonucleotide comprises a sense strand having a sequence of any one of SEQ ID NOs: 769-803 and an antisense strand comprising a complementary sequence of any one of SEQ ID NOs: 804-838. In some embodiments, the RNAi oligonucleotide comprises a sense strand having a sequence of any one of SEQ ID NOs: 769-803 and 1681 and an antisense strand comprising a complementary sequence of any one of SEQ ID NOs: 804-838.

[0206] In some embodiments, the RNAi oligonucleotide comprises a sense strand and an antisense strand, the sense strand and the antisense strand comprising a nucleotide sequence selected from the group consisting of:

[0207] a) SEQ ID NO: 769 and 804, respectively;

[0208] b) SEQ ID NOs: 770 and 805, respectively;

[0209] c) SEQ ID NOs: 771 and 806, respectively;

[0210] d) SEQ ID NOs: 772 and 807, respectively;

[0211] e) SEQ ID NOs: 773 and 808, respectively;

[0212] f) SEQ ID NOs: 774 and 809, respectively;

[0213] g) SEQ ID NOs: 775 and 810, respectively;

[0214] h) SEQ ID NOs: 776 and 811, respectively;

[0215] i) SEQ ID NOs: 777 and 812, respectively;

[0216] j) SEQ ID NOs: 778 and 813, respectively;

[0217] k) SEQ ID NOs: 779 and 814, respectively;

[0218] 1) are SEQ ID NOs: 780 and 815, respectively;

[0219] m) are SEQ ID NOs: 781 and 816, respectively;

[0220] n) are SEQ ID NOs: 782 and 817, respectively;

[0221] o) are SEQ ID NOs: 783 and 818, respectively;

[0222] p) are SEQ ID NOs: 784 and 819, respectively;

[0223] q) are SEQ ID NOs: 785 and 820, respectively;

[0224] r) are SEQ ID NOs: 786 and 821, respectively;

[0225] s) are SEQ ID NOs: 787 and 822, respectively;

[0226] t) are SEQ ID NOs: 788 and 823, respectively;

[0227] u) are SEQ ID NOs: 789 and 824, respectively;

[0228] v) SEQ ID NOs: 790 and 825, respectively;

[0229] w) are SEQ ID NOs: 791 and 826, respectively;

[0230] x) are SEQ ID NOs: 792 and 827, respectively;

[0231] y) are SEQ ID NOs: 793 and 828, respectively;

[0232] z) are SEQ ID NOs: 794 and 829, respectively;

[0233] aa) are SEQ ID NOs: 795 and 830, respectively;

[0234] bb) are SEQ ID NOs: 796 and 831, respectively;

[0235] cc) are SEQ ID NOs: 797 and 832, respectively;

[0236] dd) are SEQ ID NOs: 798 and 833, respectively;

[0237] ee) are SEQ ID NOs: 799 and 834, respectively;

[0238] ff) are SEQ ID NOs: 800 and 835, respectively;

[0239] gg) are SEQ ID NOs: 801 and 836, respectively;

[0240] hh) are SEQ ID NOs: 802 and 837, respectively;

[0241] ii) SEQ ID NOs: 803 and 838, respectively; and

[0242] jj) are SEQ ID NOs: 1681 and 815, respectively.

[0243] In some embodiments, the RNAi oligonucleotide comprises a sense strand and an antisense strand, the sense strand and the antisense strand comprising a nucleotide sequence selected from the group consisting of:

[0244] a) SEQ ID NO: 771 and 806, respectively;

[0245] b) SEQ ID NOs: 776 and 811, respectively;

[0246] c) SEQ ID NOs: 780 and 815, respectively;

[0247] d) SEQ ID NOs: 781 and 816, respectively;

[0248] e) SEQ ID NOs: 782 and 817, respectively;

[0249] f) SEQ ID NOs: 790 and 825, respectively;

[0250] g) SEQ ID NOs: 795 and 830, respectively;

[0251] h) SEQ ID NOs: 798 and 833, respectively;

[0252] i) SEQ ID NOs: 799 and 834, respectively;

[0253] j) SEQ ID NOs: 803 and 838, respectively; and

[0254] k) are SEQ ID NOs: 1681 and 815, respectively.

[0255] In some embodiments, the RNAi oligonucleotide comprises a sense strand and an antisense strand, the sense strand and the antisense strand comprising a nucleotide sequence selected from the group consisting of:

[0256] a) SEQ ID NO: 771 and 806, respectively;

[0257] b) SEQ ID NOs: 780 and 815, respectively;

[0258] c) SEQ ID NOs: 781 and 816, respectively;

[0259] d) SEQ ID NOs: 798 and 833, respectively;

[0260] e) SEQ ID NOs: 799 and 834, respectively;

[0261] f) SEQ ID NOs: 803 and 838, respectively; and

[0262] g) are SEQ ID NOs: 1681 and 815, respectively.

[0263] In some embodiments, the sense strand comprises the sequence of SEQ ID NO: 771, and the antisense strand comprises the sequence of SEQ ID NO: 806. In some embodiments, the sense strand comprises the sequence of SEQ ID NO: 780, and the antisense strand comprises the sequence of SEQ ID NO: 815. In some embodiments, the sense strand comprises the sequence of SEQ ID NO: 781, and the antisense strand comprises the sequence of SEQ ID NO: 816. In some embodiments, the sense strand comprises the sequence of SEQ ID NO: 798, and the antisense strand comprises the sequence of SEQ ID NO: 833. In some embodiments, the sense strand comprises the sequence of SEQ ID NO: 799, and the antisense strand comprises the sequence of SEQ ID NO: 834. In some embodiments, the sense strand comprises the sequence of SEQ ID NO: 803, and the antisense strand comprises the sequence of SEQ ID NO: 838. In some embodiments, the sense strand comprises the sequence of SEQ ID NO: 1681, and the antisense strand comprises the sequence of SEQ ID NO: 815.

[0264] It will be appreciated that in some embodiments, reference may be made to the sequences presented in the sequence listing when describing the structure of an oligonucleotide (e.g., an RNAi oligonucleotide) or other nucleic acid. In such embodiments, the actual oligonucleotide or other nucleic acid may have one or more alternative nucleotides (e.g., an RNA counterpart of a DNA nucleotide or a DNA counterpart of an RNA nucleotide) and / or one or more modified nucleotides and / or one or more modified internucleotide bonds and / or one or more other modifications when compared to a specified sequence while retaining a complementary property that is substantially identical or similar to the specified sequence.

[0265] In some embodiments, the RNAi oligonucleotide herein comprises a sense strand of 25 nucleotides and an antisense strand of 27 nucleotides, which, when acted upon by Dicer, produces an antisense strand that is incorporated into a mature RNA-induced silencing complex (RISC). In some embodiments, the sense strand of 25 nucleotides comprises a sequence selected from SEQ ID NO: 1-384. In some embodiments, the antisense strand of 27 nucleotides comprises a sequence selected from SEQ ID NO: 385-768. In some embodiments, the sense strand of the RNAi oligonucleotide is longer than 27 nucleotides (e.g., 28, 29, 30, 31, 32, 33, 34, 35, 36, 37, 38, 39, 40, 41, 42, 43, 44, 45, 46, 47, 48, 49, or 50 nucleotides). In some embodiments, the sense strand of the RNAi oligonucleotide is longer than 25 nucleotides (e.g., 26, 27, 28, 29, or 30 nucleotides). In some embodiments, the sense strand of the RNAi oligonucleotide comprises a nucleotide sequence selected from SEQ ID NOs: 912-1295, wherein the nucleotide sequence is longer than 27 nucleotides (e.g., 28, 29, 30, 31, 32, 33, 34, 35, 36, 37, 38, 39, 40, 41, 42, 43, 44, 45, 46, 47, 48, 49, or 50 nucleotides). In some embodiments, the sense strand of the RNAi oligonucleotide comprises a nucleotide sequence selected from SEQ ID NOs: 912-1295, wherein the nucleotide sequence is longer than 25 nucleotides (e.g., 26, 27, 28, 29, or 30 nucleotides).

[0266] In some embodiments, the RNAi oligonucleotide has a 5' end that is thermodynamically less stable when compared to the other 5' end. In some embodiments, an asymmetric RNAi oligonucleotide is provided, the oligonucleotide comprising a flat end at the 3' end of the sense strand and a 3' overhang at the 3' end of the antisense strand. In some embodiments, the 3' overhang on the antisense strand is about 1-8 nucleotides in length (e.g., 1, 2, 3, 4, 5, 6, 7, or 8 nucleotides in length). Typically, the RNAi oligonucleotide has an overhang of two nucleotides at the 3' end of the antisense (guide) strand; however, there may be other overhangs. In some embodiments, the overhang is a 3' overhang comprising a length of between about 1 to about 6 nucleotides, optionally 1 to 5, 1 to 4, 1 to 3, 1 to 2, 2 to 6, 2 to 5, 2 to 4, 2 to 3, 3 to 6, 3 to 5, 3 to 4, 4 to 6, 4 to 5, 5 to 6 nucleotides, or 1, 2, 3, 4, 5 or 6 nucleotides. However, in some embodiments, the overhang is a 5' overhang comprising a length of between about 1 to about 6 nucleotides, optionally 1 to 5, 1 to 4, 1 to 3, 1 to 2, 2 to 6, 2 to 5, 2 to 4, 2 to 3, 3 to 6, 3 to 5, 3 to 4, 4 to 6, 4 to 5, 5 to 6 nucleotides, or 1, 2, 3, 4, 5 or 6 nucleotides. In some embodiments, the RNAi oligonucleotide comprises a targeting sequence or complementary region that is complementary to a contiguous nucleotide sequence of any one of SEQ ID NOs: 912-1295, and a 5' overhang of between about 1 and about 6 nucleotides in length. In some embodiments, the RNAi oligonucleotide comprises a sense strand comprising a nucleotide sequence selected from SEQ ID NOs: 912-1295, wherein the RNAi oligonucleotide comprises a 5' overhang of between about 1 and about 6 nucleotides in length. In some embodiments, the RNAi oligonucleotide comprises an antisense strand comprising a nucleotide sequence selected from SEQ ID NOs: 1296-1679, wherein the RNAi oligonucleotide comprises a 5' overhang of between about 1 and about 6 nucleotides in length. In some embodiments, the RNAi oligonucleotide comprises a sense strand comprising a nucleotide sequence selected from SEQ ID NOs: 912-1295; and an antisense strand comprising a nucleotide sequence selected from SEQ ID NOs: 1296-1679, wherein the RNAi oligonucleotide comprises a 5' overhang between about 1 and about 6 nucleotides in length.

[0267] In some embodiments, the two terminal nucleotides on the 3' end of the antisense strand are modified. In some embodiments, the two terminal nucleotides on the 3' end of the antisense strand are complementary to target mRNA (e.g., MAPT mRNA). In some embodiments, the two terminal nucleotides on the 3' end of the antisense strand are not complementary to target mRNA. In some embodiments, the two terminal nucleotides on the 3' end of the antisense strand of the RNAi oligonucleotide herein are not paired. In some embodiments, the two terminal nucleotides on the 3' end of the antisense strand of the RNAi oligonucleotide herein include unpaired GG. In some embodiments, the two terminal nucleotides on the 3' end of the antisense strand of the RNAi oligonucleotide herein are not complementary to target mRNA. In some embodiments, the two terminal nucleotides on each 3' end of the RNAi oligonucleotide are GG. Typically, one or both of the two terminal GG nucleotides on each 3' end of the RNAi oligonucleotide are not complementary to target mRNA. In some embodiments, the two terminal nucleotides on each 3' end of the oligonucleotide are GG. In some embodiments, one or both of the two terminal GG nucleotides on each 3' end of the RNAi oligonucleotide are not complementary to the target mRNA. In some embodiments, the RNAi oligonucleotide comprises a targeting sequence or complementary region complementary to a continuous nucleotide sequence of any one of SEQ ID NO: 912-1295, wherein the two terminal nucleotides on the 3' end of the antisense strand of the oligonucleotide herein comprise unpaired GG. In some embodiments, the RNAi oligonucleotide comprises an antisense strand comprising a nucleotide sequence selected from SEQ ID NO: 1296-1679, wherein the two terminal nucleotides on the 3' end of the antisense strand of the RNAi oligonucleotide comprise unpaired GG. In some embodiments, the RNAi oligonucleotide comprises a sense strand comprising a nucleotide sequence selected from SEQ ID NO: 912-1295; and an antisense strand comprising a nucleotide sequence selected from SEQ ID NO: 1296-1679, wherein the two terminal nucleotides on the 3' end of the antisense strand of the RNAi oligonucleotide comprise unpaired GG.

[0268] In some embodiments, there are one or more (e.g., 1, 2, 3, 4, or 5) mismatches between the sense and antisense strands that make up the RNAi oligonucleotide. If there are more than one mismatch between the sense and antisense strands, they may be located consecutively (e.g., 2, 3, or more consecutively), or dispersed throughout the complementary region. In some embodiments, the 3' end of the sense strand contains one or more mismatches. In one embodiment, two mismatches are incorporated at the 3' end of the sense strand. In some embodiments, base mismatches or destabilization of a segment at the 3' end of the sense strand of the RNAi oligonucleotide improves or increases the efficacy of the oligonucleotide.

[0269] In some embodiments, the RNAi oligonucleotide comprises a sense strand and an antisense strand, the sense strand and the antisense strand comprising a nucleotide sequence selected from the group consisting of:

[0270] a) SEQ ID NO: 769 and 804, respectively;

[0271] b) SEQ ID NOs: 770 and 805, respectively;

[0272] c) SEQ ID NOs: 771 and 806, respectively;

[0273] d) SEQ ID NOs: 772 and 807, respectively;

[0274] e) SEQ ID NOs: 773 and 808, respectively;

[0275] f) SEQ ID NOs: 774 and 809, respectively;

[0276] g) SEQ ID NOs: 775 and 810, respectively;

[0277] h) SEQ ID NOs: 776 and 811, respectively;

[0278] i) SEQ ID NOs: 777 and 812, respectively;

[0279] j) SEQ ID NOs: 778 and 813, respectively;

[0280] k) SEQ ID NOs: 779 and 814, respectively;

[0281] 1) are SEQ ID NOs: 780 and 815, respectively;

[0282] m) are SEQ ID NOs: 781 and 816, respectively;

[0283] n) are SEQ ID NOs: 782 and 817, respectively;

[0284] o) are SEQ ID NOs: 783 and 818, respectively;

[0285] p) are SEQ ID NOs: 784 and 819, respectively;

[0286] q) are SEQ ID NOs: 785 and 820, respectively;

[0287] r) are SEQ ID NOs: 786 and 821, respectively;

[0288] s) are SEQ ID NOs: 787 and 822, respectively;

[0289] t) are SEQ ID NOs: 788 and 823, respectively;

[0290] u) are SEQ ID NOs: 789 and 824, respectively;

[0291] v) SEQ ID NOs: 790 and 825, respectively;

[0292] w) are SEQ ID NOs: 791 and 826, respectively;

[0293] x) are SEQ ID NOs: 792 and 827, respectively;

[0294] y) are SEQ ID NOs: 793 and 828, respectively;

[0295] z) are SEQ ID NOs: 794 and 829, respectively;

[0296] aa) are SEQ ID NOs: 795 and 830, respectively;

[0297] bb) are SEQ ID NOs: 796 and 831, respectively;

[0298] cc) are SEQ ID NOs: 797 and 832, respectively;

[0299] dd) are SEQ ID NOs: 798 and 833, respectively;

[0300] ee) are SEQ ID NOs: 799 and 834, respectively;

[0301] ff) are SEQ ID NOs: 800 and 835, respectively;

[0302] gg) are SEQ ID NOs: 801 and 836, respectively;

[0303] hh) are SEQ ID NOs: 802 and 837, respectively;

[0304] ii) SEQ ID NOs: 803 and 838, respectively; and

[0305] jj) are SEQ ID NOs: 1681 and 815, respectively, wherein there are one or more (e.g., 1, 2, 3, 4, or 5) mismatches between the sense strand and the antisense strand.

[0306] In some embodiments, the RNAi oligonucleotide comprises a sense strand and an antisense strand, the sense strand and the antisense strand comprising a nucleotide sequence selected from the group consisting of:

[0307] a) SEQ ID NO: 771 and 806, respectively;

[0308] b) SEQ ID NOs: 776 and 811, respectively;

[0309] c) SEQ ID NOs: 780 and 815, respectively;

[0310] d) SEQ ID NOs: 781 and 816, respectively;

[0311] e) SEQ ID NOs: 782 and 817, respectively;

[0312] f) SEQ ID NOs: 790 and 825, respectively;

[0313] g) SEQ ID NOs: 795 and 830, respectively;

[0314] h) SEQ ID NOs: 798 and 833, respectively;

[0315] i) SEQ ID NOs: 799 and 834, respectively;

[0316] j) SEQ ID NOs: 803 and 838, respectively; and

[0317] k) are SEQ ID NOs: 1681 and 815, respectively, wherein there are one or more (eg, 1, 2, 3, 4, or 5) mismatches between the sense strand and the antisense strand.

[0318] In some embodiments, the RNAi oligonucleotide comprises a sense strand and an antisense strand, the sense strand and the antisense strand comprising a sequence selected from the group consisting of:

[0319] a) SEQ ID NO: 771 and 806, respectively;

[0320] b) SEQ ID NOs: 780 and 815, respectively;

[0321] c) SEQ ID NOs: 781 and 816, respectively;

[0322] d) SEQ ID NOs: 798 and 833, respectively;

[0323] e) SEQ ID NOs: 799 and 834, respectively;

[0324] f) SEQ ID NOs: 803 and 838, respectively; and

[0325] g) are SEQ ID NOs: 1681 and 815, respectively, wherein there are one or more (eg, 1, 2, 3, 4, or 5) mismatches between the sense strand and the antisense strand.

[0326] Antisense strand

[0327] In some embodiments, the antisense strand of the oligonucleotides herein (e.g., RNAi oligonucleotides) is referred to as the "guide strand". The antisense strand engages with RISC and binds to an Argonaute protein, such as Ago2, or engages or binds to one or more similar factors, and directs the silencing of the target gene. In some embodiments, the sense strand complementary to the guide strand is referred to as the "passenger strand".

[0328] In some embodiments, the oligonucleotide comprises an antisense strand of at most about 50 nucleotides in length (e.g., at most 50, at most 40, at most 35, at most 30, at most 27, at most 25, at most 21, at most 19, at most 17, at most 15, or at most 12 nucleotides in length). In some embodiments, the oligonucleotide comprises an antisense strand of at least about 12 nucleotides in length (e.g., at least 12, at least 15, at least 19, at least 21, at least 22, at least 25, at least 27, at least 30, at least 35, or at least 38 nucleotides in length). In some embodiments, the oligonucleotide comprises an antisense strand ranging from about 12 to about 40 (e.g., 12 to 40, 12 to 36, 12 to 32, 12 to 28, 15 to 40, 15 to 36, 15 to 32, 15 to 30, 15 to 28, 17 to 22, 17 to 25, 19 to 27, 19 to 30, 20 to 40, 22 to 40, 25 to 40, or 32 to 40) nucleotides in length. In some embodiments, the oligonucleotide comprises an antisense strand ranging from about 15 to about 30 nucleotides in length. In some embodiments, the antisense strand of any of the oligonucleotides disclosed herein is 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, 39, or 40 nucleotides in length. In some embodiments, the oligonucleotide comprises an antisense strand of 22 nucleotides in length.

[0329] In some embodiments, the oligonucleotide for targeting MAPT comprises an antisense strand comprising or consisting of a sequence as shown in any one of SEQ ID NOs: 1296-1679. In some embodiments, the oligonucleotide comprises an antisense strand comprising at least about 12 (e.g., at least 12, at least 13, at least 14, at least 15, at least 16, at least 17, at least 18, at least 19, at least 20, at least 21, at least 22, or at least 23) consecutive nucleotide sequences as shown in any one of SEQ ID NOs: 1296-1679. In some embodiments, the oligonucleotide for targeting MAPT comprises an antisense strand comprising or consisting of a sequence as shown in any one of SEQ ID NOs: 385-768. In some embodiments, the oligonucleotide comprises an antisense strand comprising at least about 12 (e.g., at least 12, at least 13, at least 14, at least 15, at least 16, at least 17, at least 18, at least 19, at least 20, at least 21, at least 22, or at least 23) consecutive nucleotide sequences as shown in any one of SEQ ID NOs: 385-768. In some embodiments, the oligonucleotide for targeting MAPT comprises an antisense strand comprising or consisting of a sequence as shown in any one of SEQ ID NOs: 804-838. In some embodiments, the oligonucleotide comprises an antisense strand comprising at least about 12 (e.g., at least 12, at least 13, at least 14, at least 15, at least 16, at least 17, at least 18, at least 19, at least 20, at least 21, at least 22, or at least 23) consecutive nucleotide sequences as set forth in any one of SEQ ID NOs: 804-838. In some embodiments, the oligonucleotide for targeting MAPT comprises an antisense strand comprising or consisting of a sequence as shown in any one of SEQ ID NOs: 1509, 1511, 1514, 1403, 1415, 1428, 1448, 1449, 1451, 1467, 1299, 1479, 1480, 1486, 1494, 1307, 1309, 1409, 1423, 1433, 1445, 1454, 1456, 1459, 1465, 1492, 1495, 1498, 1503, 1504, 1505, 1506, 1507, 1508, and 1308.In some embodiments, the oligonucleotide comprises an antisense strand comprising at least about 12 (e.g., at least 12, at least 13, at least 14, at least 15, at least 16, at least 17, at least 18, at least 19, at least 20, at least 21, at least 22, or at least 23) sequences of SEQ ID The continuous nucleotide sequence shown in any one of NO:1509, 1511, 1514, 1403, 1415, 1428, 1448, 1449, 1451, 1467, 1299, 1479, 1480, 1486, 1494, 1307, 1309, 1409, 1423, 1433, 1445, 1454, 1456, 1459, 1465, 1492, 1495, 1498, 1503, 1504, 1505, 1506, 1507, 1508 and 1308. In some embodiments, the oligonucleotide for targeting MAPT comprises an antisense strand comprising or consisting of a sequence as shown in any one of SEQ ID NOs: 806, 811, 815, 816, 817, 825, 830, 833, 834, and 838. In some embodiments, the oligonucleotide comprises an antisense strand comprising at least about 12 (e.g., at least 12, at least 13, at least 14, at least 15, at least 16, at least 17, at least 18, at least 19, at least 20, at least 21, at least 22, or at least 23) consecutive nucleotide sequences as shown in any one of SEQ ID NOs: 806, 811, 815, 816, 817, 825, 830, 833, 834, and 838. In some embodiments, the oligonucleotide for targeting MAPT comprises an antisense strand comprising or consisting of a sequence as shown in any one of SEQ ID NOs: 806, 815, 816, 833, 834, and 838. In some embodiments, the oligonucleotide comprises an antisense strand comprising at least about 12 (e.g., at least 12, at least 13, at least 14, at least 15, at least 16, at least 17, at least 18, at least 19, at least 20, at least 21, at least 22, or at least 23) consecutive nucleotide sequences as shown in any one of SEQ ID NOs: 806, 815, 816, 833, 834, and 838.

[0330] Sense strand

[0331] In some embodiments, the oligonucleotides (e.g., RNAi oligonucleotides) for targeting MAPT mRNA include a sense strand comprising or consisting of a sequence as shown in any one of SEQ ID NOs: 912-1295. In some embodiments, the oligonucleotides have a sense strand comprising at least about 12 (e.g., at least 13, at least 14, at least 15, at least 16, at least 17, at least 18, at least 19, at least 20, at least 21, at least 22, or at least 23) consecutive nucleotide sequences as shown in any one of SEQ ID NOs: 912-1295. In some embodiments, the oligonucleotides include a sense strand sequence as shown in any one of SEQ ID NOs: 1-384. In some embodiments, the oligonucleotide has a sense strand comprising at least about 12 (e.g., at least 13, at least 14, at least 15, at least 16, at least 17, at least 18, at least 19, at least 20, at least 21, at least 22, or at least 23) consecutive nucleotide sequences as set forth in any one of SEQ ID NOs: 1-384. In some embodiments, the oligonucleotide comprises a sense strand sequence as set forth in any one of SEQ ID NOs: 769-803. In some embodiments, the oligonucleotide comprises a sense strand sequence as set forth in any one of SEQ ID NOs: 769-803 and 1681. In some embodiments, the oligonucleotide comprises a sense strand sequence as set forth in SEQ ID NOs: 1681. In some embodiments, the oligonucleotide has a sense strand comprising at least about 12 (e.g., at least 13, at least 14, at least 15, at least 16, at least 17, at least 18, at least 19, at least 20, at least 21, at least 22, or at least 23) consecutive nucleotide sequences as set forth in any one of SEQ ID NOs: 769-803. In some embodiments, the oligonucleotide comprises a sense strand sequence as set forth in any one of SEQ ID NOs: 1125, 1127, 1130, 1019, 1031, 1044, 1064, 1065, 1067, 1083, 915, 1095, 1096, 1102, 1110, 923, 925, 1025, 1039, 1049, 1061, 1070, 1072, 1075, 1081, 1108, 1111, 1114, 1119, 1120, 1121, 1122, 1123, 1124, and 924.In some embodiments, the oligonucleotide has a sense strand comprising at least about 12 (e.g., at least 13, at least 14, at least 15, at least 16, at least 17, at least 18, at least 19, at least 20, at least 21, at least 22, or at least 23) sequences of SEQ ID NO: 1125, 1127, 1130, 1019, 1031, 1044, 1064, 1065, 1067, 1083, 915, 1095, 1096, 1102, 1110, 923, 925, 1025, 1039, 1049, 1061, 1070, 1072, 1075, 1081, 1108, 1111, 1114, 1119, 1120, 1121, 1122, 1123, 1124, and 924. In some embodiments, the oligonucleotide comprises a sense strand sequence as set forth in any one of SEQ ID NOs: 771, 776, 780, 781, 782, 790, 795, 798, 799, and 803. In some embodiments, the oligonucleotide has a sense strand comprising at least about 12 (e.g., at least 13, at least 14, at least 15, at least 16, at least 17, at least 18, at least 19, at least 20, at least 21, at least 22, or at least 23) consecutive nucleotide sequences as set forth in any one of SEQ ID NOs: 771, 776, 780, 781, 782, 790, 795, 798, 799, and 803. In some embodiments, the oligonucleotide comprises a sense strand sequence as set forth in any one of SEQ ID NOs: 771, 780, 781, 798, 799, and 803. In some embodiments, the oligonucleotide has a sense strand comprising at least about 12 (e.g., at least 13, at least 14, at least 15, at least 16, at least 17, at least 18, at least 19, at least 20, at least 21, at least 22, or at least 23) consecutive nucleotide sequences as set forth in any one of SEQ ID NOs: 771, 780, 781, 798, 799, and 803.

[0332] In some embodiments, the oligonucleotide comprises a sense strand sequence as set forth in any one of SEQ ID NOs: 771, 776, 780, 781, 782, 790, 795, 798, 799, 803, and 1681. In some embodiments, the oligonucleotide has a sense strand comprising at least about 12 (e.g., at least 13, at least 14, at least 15, at least 16, at least 17, at least 18, at least 19, at least 20, at least 21, at least 22, or at least 23) contiguous nucleotide sequences as set forth in any one of SEQ ID NOs: 771, 776, 780, 781, 782, 790, 795, 798, 799, 803, and 1681. In some embodiments, the oligonucleotide comprises a sense strand sequence as set forth in any one of SEQ ID NOs: 771, 780, 781, 798, 799, 803, and 1681. In some embodiments, the oligonucleotide has a sense strand comprising at least about 12 (e.g., at least 13, at least 14, at least 15, at least 16, at least 17, at least 18, at least 19, at least 20, at least 21, at least 22, or at least 23) contiguous nucleotide sequences as set forth in any one of SEQ ID NOs: 771, 780, 781, 798, 799, 803, and 1681.

[0333] In some embodiments, the oligonucleotide comprises a sense strand that is at most about 50 nucleotides in length (e.g., at most 50, at most 40, at most 36, at most 30, at most 27, at most 25, at most 21, at most 19, at most 17, or at most 12 nucleotides in length). In some embodiments, the oligonucleotide can have a sense strand that is at least about 12 nucleotides in length (e.g., at least 12, at least 15, at least 19, at least 21, at least 25, at least 27, at least 30, at least 36, or at least 38 nucleotides in length). In some embodiments, the oligonucleotide can have a sense strand ranging from about 12 to about 50 (e.g., 12 to 50, 12 to 40, 12 to 36, 12 to 32, 12 to 28, 15 to 40, 15 to 36, 15 to 32, 15 to 28, 17 to 21, 17 to 25, 19 to 27, 19 to 30, 20 to 40, 22 to 40, 25 to 40, or 32 to 40) nucleotides in length. In some embodiments, the oligonucleotide comprises a sense strand of about 15 to about 50 nucleotides in length. In some embodiments, the oligonucleotide comprises a sense strand of 18 to 36 nucleotides in length. In some embodiments, the oligonucleotide can have a sense strand that is 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, 39, 40, 41, 42, 43, 44, 45, 46, 47, 48, 49, or 50 nucleotides in length. In some embodiments, the oligonucleotide comprises a sense strand that is 36 nucleotides in length.

[0334] In some embodiments, the oligonucleotide comprises a sense strand comprising a stem-loop structure at the 3' end of the sense strand. In some embodiments, the stem-loop is formed by intrachain base pairing. In some embodiments, the sense strand comprises a stem-loop structure at its 5' end. In some embodiments, the stem of the stem-loop comprises a duplex of 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, 12, 13 or 14 nucleotides in length. In some embodiments, the stem of the stem-loop comprises a duplex of 2 nucleotides in length. In some embodiments, the stem of the stem-loop comprises a duplex of 3 nucleotides in length. In some embodiments, the stem of the stem-loop comprises a duplex of 4 nucleotides in length. In some embodiments, the stem of the stem-loop comprises a duplex of 5 nucleotides in length. In some embodiments, the stem of the stem-loop comprises a duplex of 6 nucleotides in length. In some embodiments, the stem of the stem-loop comprises a duplex of 7 nucleotides in length. In some embodiments, the stem of the stem-loop comprises a duplex of 8 nucleotides in length. In some embodiments, the stem of the stem-loop comprises a duplex of 9 nucleotides in length. In some embodiments, the stem of the stem-loop comprises a duplex of 10 nucleotides in length. In some embodiments, the stem of the stem-loop comprises a duplex of 11 nucleotides in length. In some embodiments, the stem of the stem-loop comprises a duplex of 12 nucleotides in length. In some embodiments, the stem of the stem-loop comprises a duplex of 13 nucleotides in length. In some embodiments, the stem of the stem-loop comprises a duplex of 14 nucleotides in length.

[0335] In some embodiments, stem loop provides oligonucleotide protection for degradation (for example, enzymatic degradation), promotes or improves targeting and / or is delivered to target cell, tissue or organ (for example, liver or brain), or both.For example, in some embodiments, the ring of stem loop provides nucleotide comprising one or more modifications, and the one or more modifications promote, improve or increase targeting target mRNA (for example, MAPT mRNA), to the inhibition of target gene expression (for example, MAPT gene expression) and / or are delivered to target cell, tissue or organ (for example, CNS), or their combination.In some embodiments, stem loop itself or the modification to stem loop does not affect the intrinsic gene expression inhibition activity of oligonucleotide in general, but promotes, improves or increases stability (for example, provides protection for degradation) and / or oligonucleotide to target cell, tissue or organ (for example, CNS) delivery. In certain embodiments, the oligonucleotide comprises a sense strand comprising (e.g., at its 3' end) a stem loop as follows: S1-L-S2, wherein S1 is complementary to S2, and wherein the loop (L) forms an ss loop between S1 and S2 of up to about 10 nucleotides in length (e.g., 3, 4, 5, 6, 7, 8, 9, or 10 nucleotides in length). In some embodiments, L is 3 nucleotides in length. In some embodiments, L is 4 nucleotides in length. In some embodiments, L is 5 nucleotides in length. In some embodiments, L is 6 nucleotides in length. In some embodiments, L is 7 nucleotides in length. In some embodiments, L is 8 nucleotides in length. In some embodiments, L is 9 nucleotides in length. In some embodiments, L is 10 nucleotides in length.

[0336] In some embodiments, the oligonucleotide comprises a targeting sequence or a complementary region that is complementary to a contiguous nucleotide sequence of any one of SEQ ID NOs: 912-1295, and the oligonucleotide comprises a sense strand that comprises (e.g., at its 3' end) a stem-loop as follows: S1-L-S2, wherein S1 is complementary to S2, and wherein L forms an ss-loop of up to about 10 nucleotides in length (e.g., 3, 4, 5, 6, 7, 8, 9, or 10 nucleotides in length) between S1 and S2. In some embodiments, the oligonucleotide comprises a targeting sequence or a complementary region that is complementary to a contiguous nucleotide sequence of any one of SEQ ID NOs: 912-1295, and the oligonucleotide comprises a sense strand that comprises (e.g., at its 3' end) a stem-loop as follows: S1-L-S2, wherein S1 is complementary to S2, and wherein L forms an ss-loop of 4 nucleotides in length (i.e., tetraL) between S1 and S2.

[0337] In some embodiments, tetraL comprises the sequence 5'-GAAA-3'.In some embodiments, the stem-loop comprises the sequence 5'-GCAGCCGAAAGGCUGC-3' (SEQ ID NO: 1680).

[0338] In some embodiments, L of the stem-loop having the structure S1-L-S2 as described above is triL. In some embodiments, the oligonucleotide comprises a targeting sequence or a complementary region complementary to a contiguous nucleotide sequence of any one of SEQ ID NOs: 912-1295 and triL. In some embodiments, triL comprises ribonucleotides, deoxyribonucleotides, modified nucleotides, delivery ligands, and combinations thereof.

[0339] In some embodiments, L of the stem-loop having the structure S1-L-S2 as described above is tetraL as described in U.S. Patent No. 10,131,912 (e.g., within a nicked tetraL structure), which is incorporated herein by reference. In some embodiments, the oligonucleotide comprises a targeting sequence or complementary region complementary to a contiguous nucleotide sequence of any one of SEQ ID NOs: 912-1295 and tetraL. In some embodiments, tetraL comprises ribonucleotides, deoxyribonucleotides, modified nucleotides, delivery ligands, and combinations thereof.

[0340] Duplex length

[0341] In some embodiments, a duplex is formed between the sense strand and the antisense strand and is at least about 12 (e.g., at least 12, at least 13, at least 14, at least 15, at least 16, at least 17, at least 18, at least 19, at least 20, or at least 21) nucleotides in length. In some embodiments, the length of the duplex formed between the sense strand and the antisense strand is in the range of about 12 to about 30 nucleotides (e.g., 12 to 30, 12 to 27, 12 to 22, 15 to 25, 18 to 30, 18 to 22, 18 to 25, 18 to 27, 18 to 30, 19 to 30, or 21 to 30 nucleotides in length). In some embodiments, the length of the duplex formed between the sense strand and the antisense strand is 12, 13, 14, 15, 16, 17, 18, 19, 29, 21, 22, 23, 24, 25, 26, 27, 28, 29 or 30 nucleotides. In some embodiments, the length of the duplex formed between the sense strand and the antisense strand is 12 nucleotides. In some embodiments, the length of the duplex formed between the sense strand and the antisense strand is 13 nucleotides. In some embodiments, the length of the duplex formed between the sense strand and the antisense strand is 14 nucleotides. In some embodiments, the length of the duplex formed between the sense strand and the antisense strand is 15 nucleotides. In some embodiments, the length of the duplex formed between the sense strand and the antisense strand is 16 nucleotides. In some embodiments, the length of the duplex formed between the sense strand and the antisense strand is 17 nucleotides. In some embodiments, the length of the duplex formed between the sense strand and the antisense strand is 18 nucleotides. In some embodiments, the length of the duplex formed between the sense strand and the antisense strand is 19 nucleotides. In some embodiments, the length of the duplex formed between the sense strand and the antisense strand is 20 nucleotides. In some embodiments, the length of the duplex formed between the sense strand and the antisense strand is 21 nucleotides. In some embodiments, the length of the duplex formed between the sense strand and the antisense strand is 22 nucleotides. In some embodiments, the length of the duplex formed between the sense strand and the antisense strand is 23 nucleotides. In some embodiments, the length of the duplex formed between the sense strand and the antisense strand is 24 nucleotides. In some embodiments, the length of the duplex formed between the sense strand and the antisense strand is 25 nucleotides. In some embodiments, the length of the duplex formed between the sense strand and the antisense strand is 26 nucleotides. In some embodiments, the length of the duplex formed between the sense strand and the antisense strand is 27 nucleotides. In some embodiments, the length of the duplex formed between the sense strand and the antisense strand is 28 nucleotides. In some embodiments, the duplex formed between the sense strand and the antisense strand is 29 nucleotides in length.In some embodiments, the length of the duplex formed between the sense strand and the antisense strand is 30 nucleotides. In some embodiments, the duplex formed between the sense strand and the antisense strand does not span the entire length of the sense strand and / or the antisense strand. In some embodiments, the duplex between the sense strand and the antisense strand spans the entire length of either the sense strand or the antisense strand. In some embodiments, the duplex between the sense strand and the antisense strand spans the entire length of both the sense strand and the antisense strand.

[0342] In some embodiments, the duplex between the sense strand and the antisense strand spans the entire length of both the sense strand and the antisense strand. In some embodiments, the sense strand and the antisense strand of the oligonucleotide comprise a nucleotide sequence selected from the group consisting of:

[0343] a) SEQ ID NO: 769 and 804, respectively;

[0344] b) SEQ ID NOs: 770 and 805, respectively;

[0345] c) SEQ ID NOs: 771 and 806, respectively;

[0346] d) SEQ ID NOs: 772 and 807, respectively;

[0347] e) SEQ ID NOs: 773 and 808, respectively;

[0348] f) SEQ ID NOs: 774 and 809, respectively;

[0349] g) SEQ ID NOs: 775 and 810, respectively;

[0350] h) SEQ ID NOs: 776 and 811, respectively;

[0351] i) SEQ ID NOs: 777 and 812, respectively;

[0352] j) SEQ ID NOs: 778 and 813, respectively;

[0353] k) SEQ ID NOs: 779 and 814, respectively;

[0354] 1) are SEQ ID NOs: 780 and 815, respectively;

[0355] m) are SEQ ID NOs: 781 and 816, respectively;

[0356] n) are SEQ ID NOs: 782 and 817, respectively;

[0357] o) are SEQ ID NOs: 783 and 818, respectively;

[0358] p) are SEQ ID NOs: 784 and 819, respectively;

[0359] q) are SEQ ID NOs: 785 and 820, respectively;

[0360] r) are SEQ ID NOs: 786 and 821, respectively;

[0361] s) are SEQ ID NOs: 787 and 822, respectively;

[0362] t) are SEQ ID NOs: 788 and 823, respectively;

[0363] u) are SEQ ID NOs: 789 and 824, respectively;

[0364] v) SEQ ID NOs: 790 and 825, respectively;

[0365] w) are SEQ ID NOs: 791 and 826, respectively;

[0366] x) are SEQ ID NOs: 792 and 827, respectively;

[0367] y) are SEQ ID NOs: 793 and 828, respectively;

[0368] z) are SEQ ID NOs: 794 and 829, respectively;

[0369] aa) are SEQ ID NOs: 795 and 830, respectively;

[0370] bb) are SEQ ID NOs: 796 and 831, respectively;

[0371] cc) are SEQ ID NOs: 797 and 832, respectively;

[0372] dd) are SEQ ID NOs: 798 and 833, respectively;

[0373] ee) are SEQ ID NOs: 799 and 834, respectively;

[0374] ff) are SEQ ID NOs: 800 and 835, respectively;

[0375] gg) are SEQ ID NOs: 801 and 836, respectively;

[0376] hh) are SEQ ID NOs: 802 and 837, respectively;

[0377] ii) SEQ ID NOs: 803 and 838, respectively; and

[0378] jj) are SEQ ID NOs: 1681 and 815, respectively, wherein the length of the duplex formed between the sense strand and the antisense strand is in the range of about 12 to about 30 nucleotides (e.g., 12 to 30, 12 to 27, 12 to 22, 15 to 25, 18 to 30, 18 to 22, 18 to 25, 18 to 27, 18 to 30, 19 to 30, or 21 to 30 nucleotides in length).

[0379] In some embodiments, the duplex between the sense strand and the antisense strand spans the entire length of both the sense strand and the antisense strand. In some embodiments, the sense strand and the antisense strand of the oligonucleotide comprise a nucleotide sequence selected from the group consisting of:

[0380] a) SEQ ID NO: 771 and 806, respectively;

[0381] b) SEQ ID NOs: 776 and 811, respectively;

[0382] c) SEQ ID NOs: 780 and 815, respectively;

[0383] d) SEQ ID NOs: 781 and 816, respectively;

[0384] e) SEQ ID NOs: 782 and 817, respectively;

[0385] f) SEQ ID NOs: 790 and 825, respectively;

[0386] g) SEQ ID NOs: 795 and 830, respectively;

[0387] h) SEQ ID NOs: 798 and 833, respectively;

[0388] i) SEQ ID NOs: 799 and 834, respectively;

[0389] j) SEQ ID NOs: 803 and 838, respectively; and

[0390] k) are SEQ ID NOs: 1681 and 815, respectively, wherein the length of the duplex formed between the sense strand and the antisense strand is in the range of about 12 to about 30 nucleotides (e.g., 12 to 30, 12 to 27, 12 to 22, 15 to 25, 18 to 30, 18 to 22, 18 to 25, 18 to 27, 18 to 30, 19 to 30, or 21 to 30 nucleotides in length).

[0391] In some embodiments, the duplex between the sense strand and the antisense strand spans the entire length of both the sense strand and the antisense strand. In some embodiments, the sense strand and the antisense strand of the oligonucleotide comprise a nucleotide sequence selected from the group consisting of:

[0392] a) SEQ ID NO: 771 and 806, respectively;

[0393] b) SEQ ID NOs: 780 and 815, respectively;

[0394] c) SEQ ID NOs: 781 and 816, respectively;

[0395] d) SEQ ID NOs: 798 and 833, respectively;

[0396] e) SEQ ID NOs: 799 and 834, respectively;

[0397] f) SEQ ID NOs: 803 and 838, respectively; and

[0398] g) are SEQ ID NOs: 1681 and 815, respectively, wherein the length of the duplex formed between the sense strand and the antisense strand is in the range of about 12 to about 30 nucleotides (e.g., 12 to 30, 12 to 27, 12 to 22, 15 to 25, 18 to 30, 18 to 22, 18 to 25, 18 to 27, 18 to 30, 19 to 30, or 21 to 30 nucleotides in length).

[0399] Oligonucleotide ends

[0400] In some embodiments, the oligonucleotides herein (e.g., RNAi oligonucleotides) comprise a sense strand and an antisense strand, wherein the ends of any one or both strands comprise a flat end. In some embodiments, the oligonucleotide comprises a sense strand and an antisense strand, and the sense strand and the antisense strand are independent strands forming an asymmetric duplex region, and the duplex region has an overhang at the 3' end of the antisense strand. In some embodiments, the oligonucleotide comprises a sense strand and an antisense strand, wherein the ends of any one or both strands comprise an overhang containing one or more nucleotides. In some embodiments, the one or more nucleotides constituting the overhang are unpaired nucleotides. In some embodiments, the oligonucleotide comprises a sense strand and an antisense strand, wherein the 3' end of the sense strand and the 5' end of the antisense strand comprise a flat end. In some embodiments, the oligonucleotide comprises a sense strand and an antisense strand, wherein the 5' end of the sense strand and the 3' end of the antisense strand comprise a flat end.

[0401] In some embodiments, the oligonucleotide comprises a sense strand and an antisense strand, wherein the 3' end of either or both strands comprises a 3' overhang comprising one or more nucleotides. In some embodiments, the oligonucleotide comprises a sense strand and an antisense strand, wherein the sense strand comprises a 3' overhang comprising one or more nucleotides. In some embodiments, the oligonucleotide comprises a sense strand and an antisense strand, wherein the antisense strand comprises a 3' overhang comprising one or more nucleotides. In some embodiments, the oligonucleotide comprises a sense strand and an antisense strand, wherein both the sense strand and the antisense strand comprise a 3' overhang comprising one or more nucleotides.

[0402] In some embodiments, the 3' overhang is about 1 to about 20 nucleotides in length (e.g., about 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, 15, 16, 17, 18, 19, or 20 nucleotides in length). In some embodiments, the 3' overhang is 1 to 19, 1 to 18, 1 to 17, 1 to 16, 1 to 15, 1 to 14, 1 to 13, 1 to 12, 1 to 11, 1 to 10, 1 to 9, 1 to 8, 1 to 7, 1 to 6, 1 to 5, 1 to 4, 1 to 3, or 1 to 2 nucleotides in length. In some embodiments, the 3' overhang is 1 nucleotide in length. In some embodiments, the 3' overhang is 2 nucleotides in length. In some embodiments, the 3' overhang is 3 nucleotides in length. In some embodiments, the 3' overhang is 4 nucleotides in length. In some embodiments, the 3' overhang is 5 nucleotides in length. In some embodiments, the 3' overhang is 6 nucleotides in length. In some embodiments, the 3' overhang is 7 nucleotides in length. In some embodiments, the 3' overhang is 8 nucleotides in length. In some embodiments, the 3' overhang is 9 nucleotides in length. In some embodiments, the 3' overhang is 10 nucleotides in length. In some embodiments, the 3' overhang is 11 nucleotides in length. In some embodiments, the 3' overhang is 12 nucleotides in length. In some embodiments, the 3' overhang is 13 nucleotides in length. In some embodiments, the 3' overhang is 14 nucleotides in length. In some embodiments, the 3' overhang is 15 nucleotides in length. In some embodiments, the 3' overhang is 16 nucleotides in length. In some embodiments, the 3' overhang is 17 nucleotides in length. In some embodiments, the 3' overhang is 18 nucleotides in length. In some embodiments, the 3' overhang is 19 nucleotides in length. In some embodiments, the 3' overhang is 20 nucleotides in length.

[0403] In some embodiments, the oligonucleotide comprises a sense strand and an antisense strand, wherein the antisense strand comprises a 3' overhang, wherein the sense strand and the antisense strand of the oligonucleotide comprise a nucleotide sequence selected from the group consisting of:

[0404] a) SEQ ID NO: 769 and 804, respectively;

[0405] b) SEQ ID NOs: 770 and 805, respectively;

[0406] c) SEQ ID NOs: 771 and 806, respectively;

[0407] d) SEQ ID NOs: 772 and 807, respectively;

[0408] e) SEQ ID NOs: 773 and 808, respectively;

[0409] f) SEQ ID NOs: 774 and 809, respectively;

[0410] g) SEQ ID NOs: 775 and 810, respectively;

[0411] h) SEQ ID NOs: 776 and 811, respectively;

[0412] i) SEQ ID NOs: 777 and 812, respectively;

[0413] j) SEQ ID NOs: 778 and 813, respectively;

[0414] k) SEQ ID NOs: 779 and 814, respectively;

[0415] 1) are SEQ ID NOs: 780 and 815, respectively;

[0416] m) are SEQ ID NOs: 781 and 816, respectively;

[0417] n) are SEQ ID NOs: 782 and 817, respectively;

[0418] o) are SEQ ID NOs: 783 and 818, respectively;

[0419] p) are SEQ ID NOs: 784 and 819, respectively;

[0420] q) are SEQ ID NOs: 785 and 820, respectively;

[0421] r) are SEQ ID NOs: 786 and 821, respectively;

[0422] s) are SEQ ID NOs: 787 and 822, respectively;

[0423] t) are SEQ ID NOs: 788 and 823, respectively;

[0424] u) are SEQ ID NOs: 789 and 824, respectively;

[0425] v) SEQ ID NOs: 790 and 825, respectively;

[0426] w) are SEQ ID NOs: 791 and 826, respectively;

[0427] x) are SEQ ID NOs: 792 and 827, respectively;

[0428] y) are SEQ ID NOs: 793 and 828, respectively;

[0429] z) are SEQ ID NOs: 794 and 829, respectively;

[0430] aa) are SEQ ID NOs: 795 and 830, respectively;

[0431] bb) are SEQ ID NOs: 796 and 831, respectively;

[0432] cc) are SEQ ID NOs: 797 and 832, respectively;

[0433] dd) are SEQ ID NOs: 798 and 833, respectively;

[0434] ee) are SEQ ID NOs: 799 and 834, respectively;

[0435] ff) are SEQ ID NOs: 800 and 835, respectively;

[0436] gg) are SEQ ID NOs: 801 and 836, respectively;

[0437] hh) are SEQ ID NOs: 802 and 837, respectively;

[0438] ii) SEQ ID NOs: 803 and 838, respectively; and

[0439] jj) are SEQ ID NOs: 1681 and 815, respectively, wherein the antisense strand comprises a 3' overhang of about 1 to about 20 nucleotides in length (e.g., about 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, 15, 16, 17, 18, 19, or 20 nucleotides in length), optionally wherein the 3' overhang is 2 nucleotides in length.

[0440] In some embodiments, the oligonucleotide comprises a sense strand and an antisense strand, wherein the antisense strand comprises a 3' overhang, wherein the sense strand and the antisense strand of the oligonucleotide comprise a nucleotide sequence selected from the group consisting of:

[0441] a) SEQ ID NO: 771 and 806, respectively;

[0442] b) SEQ ID NOs: 776 and 811, respectively;

[0443] c) SEQ ID NOs: 780 and 815, respectively;

[0444] d) SEQ ID NOs: 781 and 816, respectively;

[0445] e) SEQ ID NOs: 782 and 817, respectively;

[0446] f) SEQ ID NOs: 790 and 825, respectively;

[0447] g) SEQ ID NOs: 795 and 830, respectively;

[0448] h) SEQ ID NOs: 798 and 833, respectively;

[0449] i) SEQ ID NOs: 799 and 834, respectively;

[0450] j) SEQ ID NOs: 803 and 838, respectively; and

[0451] k) are SEQ ID NOs: 1681 and 815, respectively, wherein the antisense strand comprises a 3' overhang of about 1 to about 20 nucleotides in length (e.g., about 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, 15, 16, 17, 18, 19, or 20 nucleotides in length), optionally wherein the 3' overhang is 2 nucleotides in length.

[0452] In some embodiments, the oligonucleotide comprises a sense strand and an antisense strand, wherein the antisense strand comprises a 3' overhang, wherein the sense strand and the antisense strand of the oligonucleotide comprise a nucleotide sequence selected from the group consisting of:

[0453] a) SEQ ID NO: 771 and 806, respectively;

[0454] b) SEQ ID NOs: 780 and 815, respectively;

[0455] c) SEQ ID NOs: 781 and 816, respectively;

[0456] d) SEQ ID NOs: 798 and 833, respectively;

[0457] e) SEQ ID NOs: 799 and 834, respectively;

[0458] f) SEQ ID NOs: 803 and 838, respectively; and

[0459] g) are SEQ ID NOs: 1681 and 815, respectively, wherein the antisense strand comprises a 3' overhang of about 1 to about 20 nucleotides in length (e.g., about 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, 15, 16, 17, 18, 19, or 20 nucleotides in length), optionally wherein the 3' overhang is 2 nucleotides in length.

[0460] In some embodiments, an oligonucleotide comprises a sense strand and an antisense strand, wherein the sense strand comprises a 5' overhang comprising one or more nucleotides.

[0461] In some embodiments, the oligonucleotide comprises a sense strand and an antisense strand, wherein the sense strand and the antisense strand of the oligonucleotide comprise a nucleotide sequence selected from the group consisting of:

[0462] a) SEQ ID NO: 769 and 804, respectively;

[0463] b) SEQ ID NOs: 770 and 805, respectively;

[0464] c) SEQ ID NOs: 771 and 806, respectively;

[0465] d) SEQ ID NOs: 772 and 807, respectively;

[0466] e) SEQ ID NOs: 773 and 808, respectively;

[0467] f) SEQ ID NOs: 774 and 809, respectively;

[0468] g) SEQ ID NOs: 775 and 810, respectively;

[0469] h) SEQ ID NOs: 776 and 811, respectively;

[0470] i) SEQ ID NOs: 777 and 812, respectively;

[0471] j) SEQ ID NOs: 778 and 813, respectively;

[0472] k) SEQ ID NOs: 779 and 814, respectively;

[0473] 1) are SEQ ID NOs: 780 and 815, respectively;

[0474] m) are SEQ ID NOs: 781 and 816, respectively;

[0475] n) are SEQ ID NOs: 782 and 817, respectively;

[0476] o) are SEQ ID NOs: 783 and 818, respectively;

[0477] p) are SEQ ID NOs: 784 and 819, respectively;

[0478] q) are SEQ ID NOs: 785 and 820, respectively;

[0479] r) are SEQ ID NOs: 786 and 821, respectively;

[0480] s) are SEQ ID NOs: 787 and 822, respectively;

[0481] t) are SEQ ID NOs: 788 and 823, respectively;

[0482] u) are SEQ ID NOs: 789 and 824, respectively;

[0483] v) SEQ ID NOs: 790 and 825, respectively;

[0484] w) are SEQ ID NOs: 791 and 826, respectively;

[0485] x) are SEQ ID NOs: 792 and 827, respectively;

[0486] y) are SEQ ID NOs: 793 and 828, respectively;

[0487] z) are SEQ ID NOs: 794 and 829, respectively;

[0488] aa) are SEQ ID NOs: 795 and 830, respectively;

[0489] bb) are SEQ ID NOs: 796 and 831, respectively;

[0490] cc) are SEQ ID NOs: 797 and 832, respectively;

[0491] dd) are SEQ ID NOs: 798 and 833, respectively;

[0492] ee) are SEQ ID NOs: 799 and 834, respectively;

[0493] ff) are SEQ ID NOs: 800 and 835, respectively;

[0494] gg) are SEQ ID NOs: 801 and 836, respectively;

[0495] hh) are SEQ ID NOs: 802 and 837, respectively;

[0496] ii) SEQ ID NOs: 803 and 838, respectively; and

[0497] jj) are SEQ ID NOs: 1681 and 815, respectively, wherein the antisense strand comprises a 3' overhang of about 1 to about 20 nucleotides in length (e.g., about 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, 15, 16, 17, 18, 19 or about 20 nucleotides in length), optionally wherein the 3' overhang is 2 nucleotides in length.

[0498] In some embodiments, the oligonucleotide comprises a sense strand and an antisense strand, wherein the sense strand and the antisense strand of the oligonucleotide comprise a nucleotide sequence selected from the group consisting of:

[0499] a) SEQ ID NO: 771 and 806, respectively;

[0500] b) SEQ ID NOs: 776 and 811, respectively;

[0501] c) SEQ ID NOs: 780 and 815, respectively;

[0502] d) SEQ ID NOs: 781 and 816, respectively;

[0503] e) SEQ ID NOs: 782 and 817, respectively;

[0504] f) SEQ ID NOs: 790 and 825, respectively;

[0505] g) SEQ ID NOs: 795 and 830, respectively;

[0506] h) SEQ ID NOs: 798 and 833, respectively;

[0507] i) SEQ ID NOs: 799 and 834, respectively;

[0508] j) SEQ ID NOs: 803 and 838, respectively; and

[0509] k) are SEQ ID NOs: 1681 and 815, respectively, wherein the antisense strand comprises a 3' overhang of about 1 to about 20 nucleotides in length (e.g., about 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, 15, 16, 17, 18, 19 or about 20 nucleotides in length), optionally wherein the 3' overhang is 2 nucleotides in length.

[0510] In some embodiments, the oligonucleotide comprises a sense strand and an antisense strand, wherein the sense strand and the antisense strand of the oligonucleotide comprise a nucleotide sequence selected from the group consisting of:

[0511] a) SEQ ID NO: 771 and 806, respectively;

[0512] b) SEQ ID NOs: 780 and 815, respectively;

[0513] c) SEQ ID NOs: 781 and 816, respectively;

[0514] d) SEQ ID NOs: 798 and 833, respectively;

[0515] e) SEQ ID NOs: 799 and 834, respectively;

[0516] f) SEQ ID NOs: 803 and 838, respectively; and

[0517] g) are SEQ ID NOs: 1681 and 815, respectively, wherein the antisense strand comprises a 3' overhang of about 1 to about 20 nucleotides in length (e.g., about 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, 15, 16, 17, 18, 19 or about 20 nucleotides in length), optionally wherein the 3' overhang is 2 nucleotides in length.

[0518] In some embodiments, one or more (e.g., 2, 3, 4, 5, or more) nucleotides that make up the 3' end or 5' end of the sense strand and / or antisense strand are modified. For example, in some embodiments, one or two terminal nucleotides at the 3' end of the antisense strand are modified. In some embodiments, the last nucleotide at the 3' end of the antisense strand is modified, for example, comprising a 2' modification (e.g., 2'-OMe). In some embodiments, the last one or two terminal nucleotides at the 3' end of the antisense strand are complementary to the target. In some embodiments, the last one or two nucleotides at the 3' end of the antisense strand are not complementary to the target.

[0519] In some embodiments, the oligonucleotide comprises a sense strand and an antisense strand, wherein the 3' end of the sense strand comprises a stem loop and the 3' end of the antisense strand comprises a 3' overhang. In some embodiments, the oligonucleotide comprises a sense strand and an antisense strand forming a nicked tetraL structure, wherein the 3' end of the sense strand comprises a stem loop, wherein the loop is a tetraL, and wherein the 3' end of the antisense strand comprises a 3' overhang as described herein. In some embodiments, the 3' overhang is 2 nucleotides in length. In some embodiments, the 2 nucleotides constituting the 3' overhang both comprise a guanine (G) nucleobase. Typically, one or both of the nucleotides constituting the 3' overhang of the antisense strand are not complementary to the target mRNA.

[0520] Oligonucleotide modification

[0521] In some embodiments, oligonucleotides (e.g., RNAi oligonucleotides) comprise modifications. Oligonucleotides can be modified in various ways to improve or control specificity, stability, delivery, bioavailability, resistance to nuclease degradation, immunogenicity, base pairing properties, RNA distribution and cellular uptake, and other features relevant to therapeutic research uses.

[0522] In some embodiments, the modification is a modified sugar. In some embodiments, the modification is a 5' terminal phosphate group. In some embodiments, the modification is a modified internucleoside bond. In some embodiments, the modification is a modified base. In some embodiments, the modification is a reversible modification. In some embodiments, the oligonucleotide may comprise any one of the modifications described herein or any combination thereof. For example, in some embodiments, the oligonucleotide comprises at least one modified sugar, a 5' terminal phosphate group, at least one modified internucleoside bond, at least one modified base, and at least one reversible modification.

[0523] In some embodiments, the oligonucleotide comprises at least one modified sugar, a 5' terminal phosphate group, at least one modified internucleotide bond, and at least one modified base. In some embodiments, the sense strand and the antisense strand of the oligonucleotide comprise a nucleotide sequence selected from the group consisting of:

[0524] a) SEQ ID NO: 769 and 804, respectively;

[0525] b) SEQ ID NOs: 770 and 805, respectively;

[0526] c) SEQ ID NOs: 771 and 806, respectively;

[0527] d) SEQ ID NOs: 772 and 807, respectively;

[0528] e) SEQ ID NOs: 773 and 808, respectively;

[0529] f) SEQ ID NOs: 774 and 809, respectively;

[0530] g) SEQ ID NOs: 775 and 810, respectively;

[0531] h) SEQ ID NOs: 776 and 811, respectively;

[0532] i) SEQ ID NOs: 777 and 812, respectively;

[0533] j) SEQ ID NOs: 778 and 813, respectively;

[0534] k) SEQ ID NOs: 779 and 814, respectively;

[0535] 1) are SEQ ID NOs: 780 and 815, respectively;

[0536] m) are SEQ ID NOs: 781 and 816, respectively;

[0537] n) are SEQ ID NOs: 782 and 817, respectively;

[0538] o) are SEQ ID NOs: 783 and 818, respectively;

[0539] p) are SEQ ID NOs: 784 and 819, respectively;

[0540] q) are SEQ ID NOs: 785 and 820, respectively;

[0541] r) are SEQ ID NOs: 786 and 821, respectively;

[0542] s) are SEQ ID NOs: 787 and 822, respectively;

[0543] t) are SEQ ID NOs: 788 and 823, respectively;

[0544] u) are SEQ ID NOs: 789 and 824, respectively;

[0545] v) SEQ ID NOs: 790 and 825, respectively;

[0546] w) are SEQ ID NOs: 791 and 826, respectively;

[0547] x) are SEQ ID NOs: 792 and 827, respectively;

[0548] y) are SEQ ID NOs: 793 and 828, respectively;

[0549] z) are SEQ ID NOs: 794 and 829, respectively;

[0550] aa) are SEQ ID NOs: 795 and 830, respectively;

[0551] bb) are SEQ ID NOs: 796 and 831, respectively;

[0552] cc) are SEQ ID NOs: 797 and 832, respectively;

[0553] dd) are SEQ ID NOs: 798 and 833, respectively;

[0554] ee) are SEQ ID NOs: 799 and 834, respectively;

[0555] ff) are SEQ ID NOs: 800 and 835, respectively;

[0556] gg) are SEQ ID NOs: 801 and 836, respectively;

[0557] hh) are SEQ ID NOs: 802 and 837, respectively;

[0558] ii) SEQ ID NOs: 803 and 838, respectively; and

[0559] jj) are SEQ ID NOs: 1681 and 815, respectively, wherein the oligonucleotide comprises at least one modified sugar, a 5' terminal phosphate group, at least one modified internucleotide bond and at least one modified base.

[0560] In some embodiments, the sense strand and the antisense strand of the oligonucleotide comprise a nucleotide sequence selected from the group consisting of:

[0561] a) SEQ ID NO: 771 and 806, respectively;

[0562] b) SEQ ID NOs: 776 and 811, respectively;

[0563] c) SEQ ID NOs: 780 and 815, respectively;

[0564] d) SEQ ID NOs: 781 and 816, respectively;

[0565] e) SEQ ID NOs: 782 and 817, respectively;

[0566] f) SEQ ID NOs: 790 and 825, respectively;

[0567] g) SEQ ID NOs: 795 and 830, respectively;

[0568] h) SEQ ID NOs: 798 and 833, respectively;

[0569] i) SEQ ID NOs: 799 and 834, respectively;

[0570] j) SEQ ID NOs: 803 and 838, respectively; and

[0571] k) are SEQ ID NOs: 1681 and 815, respectively, wherein the oligonucleotide comprises at least one modified sugar, a 5' terminal phosphate group, at least one modified internucleotide linkage and at least one modified base.

[0572] In some embodiments, the sense strand and the antisense strand of the oligonucleotide comprise a nucleotide sequence selected from the group consisting of:

[0573] a) SEQ ID NO: 771 and 806, respectively;

[0574] b) SEQ ID NOs: 780 and 815, respectively;

[0575] c) SEQ ID NOs: 781 and 816, respectively;

[0576] d) SEQ ID NOs: 798 and 833, respectively;

[0577] e) SEQ ID NOs: 799 and 834, respectively;

[0578] f) SEQ ID NOs: 803 and 838, respectively; and

[0579] g) are SEQ ID NOs: 1681 and 815, respectively, wherein the oligonucleotide comprises at least one modified sugar, a 5' terminal phosphate group, at least one modified internucleotide linkage and at least one modified base.

[0580] The number of modifications on the oligonucleotide and the positions of those nucleotide modifications may affect the characteristics of the oligonucleotide. For example, the oligonucleotide can be delivered in vivo by being conjugated to a lipid nanoparticle (LNP) or a similar carrier or being included in a lipid nanoparticle (LNP) or a similar carrier. However, when the oligonucleotide is not protected by LNP or a similar carrier, at least some nucleotides may be advantageously modified. Therefore, in some embodiments, all or substantially all nucleotides of the oligonucleotide are modified. In some embodiments, more than half of the nucleotides are modified. In some embodiments, less than half of the nucleotides are modified. In some embodiments, the sugar moieties of all nucleotides constituting the oligonucleotide are modified at the 2' position. The modification may be reversible or irreversible. In some embodiments, the oligonucleotide as disclosed herein has the number and type of modified nucleotides sufficient to facilitate the desired characteristics (e.g., preventing enzymatic degradation, the ability and / or thermodynamic stability of the desired cells targeted after in vivo administration).

[0581] Sugar modification

[0582] In some embodiments, oligonucleotides include modified sugars. In some embodiments, modified sugars (also referred to herein as sugar analogs) include modified deoxyribose or ribose moieties, for example, wherein one or more modifications occur at 2', 3', 4' and / or 5' carbon positions of sugars. In some embodiments, modified sugars may also include non-natural alternative carbon structures, such as those present in the following structures: locked nucleic acids ("LNAs"; see, for example, Koshkin et al. (1998) TETRAHEDON 54:3607-30), non-locked nucleic acids ("UNA"; see, for example, Snead et al. (2013) MOL.THER-NUCL.ACIDS2:e103) and bridged nucleic acids ("BNAs"; see, for example, Imanishi and Obika (2002) CHEM COMMUN. (CAMB) 21:1653-59).

[0583] In some embodiments, the nucleotide modification in the sugar includes 2'-modification. In some embodiments, the 2'-modification can be 2'-O-propargyl, 2'-O-propylamine, 2'-amino, 2'-ethyl, 2'-F, EA, 2'-OMe, 2'-MOE, 2'-O-[2-(methylamino)-2-oxoethyl] (2'-O-NMA) or 2'-FANA. In some embodiments, the modification is 2'-F, 2'-OMe or 2'-MOE. In some embodiments, the modification in the sugar includes the modification of the sugar ring, which may include the modification of one or more carbons of the sugar ring. For example, the modification of the sugar of the nucleotide may include the 2'-oxygen of the sugar being connected to the 1'-carbon or 4'-carbon of the sugar, or the 2'-oxygen being connected to the 1'-carbon or 4'-carbon via an ethylene or methylene bridge. In some embodiments, the modified nucleotide has an acyclic sugar lacking a 2'-carbon to 3'-carbon bond. In some embodiments, the modified nucleotide has a thiol group, for example, at the 4'-position of the sugar.

[0584] In some embodiments, the oligonucleotides described herein comprise at least about 1 modified nucleotide (e.g., at least 1, at least 5, at least 10, at least 15, at least 20, at least 25, at least 30, at least 35, at least 40, at least 45, at least 50, at least 55, at least 60, or more). In some embodiments, the sense strand of the RNAi oligonucleotide comprises at least about 1 modified nucleotide (e.g., at least 1, at least 5, at least 10, at least 15, at least 20, at least 25, at least 30, at least 35, or more). In some embodiments, the antisense strand of the oligonucleotide comprises at least about 1 modified nucleotide (e.g., at least 1, at least 5, at least 10, at least 15, at least 20, or more).

[0585] In some embodiments, all nucleotides of the sense strand of an oligonucleotide are modified. In some embodiments, all nucleotides of the antisense strand of an oligonucleotide are modified. In some embodiments, all nucleotides of an oligonucleotide (i.e., both the sense strand and the antisense strand) are modified. In some embodiments, the modified nucleotides comprise a 2'-modification (e.g., 2'-F or 2'-OMe, 2'-MOE, and 2'-FANA). In some embodiments, the modified nucleotides comprise a 2'-modification (e.g., 2'-F or 2'-OMe).

[0586] In some embodiments, the present disclosure provides oligonucleotides with different modification patterns. In some embodiments, the modified oligonucleotides comprise a sense strand sequence having a modification pattern as listed in the Examples and Sequence Listing and an antisense strand having a modification pattern as listed in the Examples and Sequence Listing.

[0587] In some embodiments, the oligonucleotide comprises an antisense strand comprising 2'-F modified nucleotides. In some embodiments, the oligonucleotide comprises an antisense strand comprising 2'-F and 2'-OMe modified nucleotides. In some embodiments, the oligonucleotide comprises a sense strand comprising 2'-F modified nucleotides. In some embodiments, the oligonucleotide comprises a sense strand comprising 2'-F and 2'-OMe modified nucleotides.

[0588] In some embodiments, the oligonucleotide comprises a sense strand, wherein about 10-15%, 10%, 11%, 12%, 13%, 14%, or 15% of the nucleotides of the sense strand comprise a 2'-F modification. In some embodiments, the oligonucleotide comprises a sense strand, wherein about 18-23% (e.g., 18%, 19%, 20%, 21%, 22%, or 23%) of the nucleotides of the sense strand comprise a 2'-F modification. In some embodiments, the oligonucleotide comprises a sense strand, wherein about 38-43% (e.g., 38%, 39%, 40%, 41%, 42%, or 43%) of the nucleotides of the sense strand comprise a 2'-F modification. In some embodiments, about 11% of the nucleotides of the sense strand comprise a 2'-F modification. In some embodiments, about 22% of the nucleotides of the sense strand comprise a 2'-F modification. In some embodiments, about 40% of the nucleotides of the sense strand comprise a 2'-F modification. In some embodiments, the oligonucleotide comprises an antisense strand, wherein about 25% to about 35% (e.g., 25%, 26%, 27%, 28%, 29%, 30%, 31%, 32%, 33%, 34%, or 35%) of the nucleotides of the antisense strand comprise a 2'-F modification. In some embodiments, about 32% of the nucleotides of the antisense strand comprise a 2'-F modification. In some embodiments, the oligonucleotide has about 15% to about 25% (e.g., 15%, 16%, 17%, 18%, 19%, 20%, 21%, 22%, 23%, 24%, or 25%) of its nucleotides comprising a 2'-F modification. In some embodiments, the oligonucleotide has about 35-45% (e.g., 35%, 36%, 37%, 38%, 39%, 40%, 41%, 42%, 43%, 44%, or 45%) of its nucleotides comprising a 2'-F modification. In some embodiments, about 19% of the nucleotides in the oligonucleotide comprise a 2'-F modification. In some embodiments, about 29% of the nucleotides in the oligonucleotide comprise a 2'-F modification. In some embodiments, about 40% of the nucleotides in the oligonucleotide comprise a 2'-F modification.

[0589] In some embodiments, one or more of positions 8, 9, 10, or 11 of the 36-nucleotide sense strand is modified with a 2'-F group. In some embodiments, one or more of positions 8, 9, 10, or 11 of the sense strand comprising a stem-loop is modified with a 2'-F group. In some embodiments, the sugar moiety at each nucleotide at positions 1-7 and 12-20 of the 36-nucleotide sense strand is modified with 2'-OMe. In some embodiments, the sugar moiety at each nucleotide at positions 1-7 and 12-20 of the sense strand comprising a stem-loop is modified with 2'-OMe. In some embodiments, the sugar moiety at each nucleotide at positions 1-7 and 12-36 in the sense strand is modified with 2'-OMe.

[0590] In some embodiments, one or more of positions 3, 5, 8, 10, 12, 13, 15, and 17 of the sense strand are modified with 2'-F.

[0591] In some embodiments, the antisense strand has 3 nucleotides modified with 2'-F at the 2'-position of the sugar moiety. In some embodiments, the sugar moieties at positions 2, 5, and 14 of the antisense strand and optionally up to 3 nucleotides at positions 1, 3, 7, and 10 are modified with 2'-F. In some embodiments, the sugar moieties at positions 2, 5, and 14 of the antisense strand and optionally up to 3 nucleotides at positions 3, 4, 7, and 10 are modified with 2'-F. In other embodiments, the sugar moieties at each of positions 2, 5, and 14 of the antisense strand are modified with 2'-F. In other embodiments, the sugar moieties at each of positions 1, 2, 5, and 14 of the antisense strand are modified with 2'-F. In other embodiments, the sugar moieties at each of positions 2, 4, 5, and 14 of the antisense strand are modified with 2'-F. In other embodiments, the sugar moieties at each of positions 1, 2, 3, 5, 7, and 14 of the antisense strand are modified with 2'-F. In other embodiments, the sugar moieties at each of positions 1, 2, 3, 5, 7, and 14 of the antisense strand are modified with 2'-F. In other embodiments, the sugar moiety at each of positions 2, 3, 4, 5, 7, and 14 of the antisense strand is modified with 2'-F. In another embodiment, the sugar moiety at each of positions 1, 2, 3, 5, 10, and 14 of the antisense strand is modified with 2'-F. In other embodiments, the sugar moiety at each of positions 2, 3, 4, 5, 10, and 14 of the antisense strand is modified with 2'-F. In another embodiment, the sugar moiety at each of positions 2, 3, 4, 5, 7, 10, and 14 of the antisense strand is modified with 2'-F. In other embodiments, the sugar moiety at each of positions 2, 3, 4, 5, 7, 10, and 14 of the antisense strand that forms a duplex with the sense strand of 36 nucleotides is modified with 2'-F. In other embodiments, the sugar moiety at each of positions 2, 3, 4, 5, 7, 10, and 14 of the antisense strand that forms a duplex with the sense strand comprising a stem-loop is modified with 2'-F.

[0592] In some embodiments, an oligonucleotide comprises an antisense strand having a sugar moiety modified with 2'-F at positions 2 and 14. In some embodiments, an oligonucleotide comprises an antisense strand having a sugar moiety modified with 2'-F at positions 2, 5, and 14. In some embodiments, an oligonucleotide comprises an antisense strand having a sugar moiety modified with 2'-F at positions 1, 2, 5, and 14. In some embodiments, an oligonucleotide comprises an antisense strand having a sugar moiety modified with 2'-F at positions 2, 4, 5, and 14. In some embodiments, an oligonucleotide comprises an antisense strand having a sugar moiety modified with 2'-F at positions 1, 2, 3, 5, 7, and 14. In some embodiments, an oligonucleotide comprises an antisense strand having a sugar moiety modified with 2'-F at positions 2, 3, 4, 5, 7, and 14. In some embodiments, the oligonucleotide comprises an antisense strand having a sugar moiety modified with 2'-F at positions 1, 2, 3, 5, 10, and 14. In some embodiments, the oligonucleotide comprises an antisense strand having a sugar moiety modified with 2'-F at positions 2, 3, 4, 5, 10, and 14. In some embodiments, the oligonucleotide comprises an antisense strand having a sugar moiety modified with 2'-F at positions 2, 3, 4, 5, 10, and 14. In some embodiments, the oligonucleotide comprises a 36-nucleotide sense strand and an antisense strand, wherein the antisense strand comprises a sugar moiety modified with 2'-F at positions 2, 3, 4, 5, 7, 10, and 14. In some embodiments, the oligonucleotide comprises a sense strand and an antisense strand containing a stem-loop, wherein the antisense strand comprises a sugar moiety modified with 2'-F at positions 2, 3, 4, 5, 7, 10, and 14. In some embodiments, the oligonucleotide comprises an antisense strand having a sugar moiety modified with 2'-F at positions 2, 3, 4, 5, 7, 10, 14, 16, and 19.

[0593] In some embodiments, the oligonucleotide comprises an antisense strand having a sugar moiety with each nucleotide at positions 2, 5, and 14 of the antisense strand modified with a 2'-F modified sugar moiety and each remaining nucleotide of the antisense strand modified with a modification selected from the group consisting of: 2'-O-propargyl, 2'-O-propylamine, 2'-amino, 2'-ethyl, EA, 2'-OMe, 2'-MOE, 2'-O-NMA, and 2'-FANA.

[0594] In some embodiments, the oligonucleotide comprises an antisense strand having a sugar moiety with each nucleotide at positions 1, 2, 5, and 14 of the antisense strand modified with a 2'-F modified sugar moiety and each remaining nucleotide of the antisense strand modified with a modification selected from the group consisting of: 2'-O-propargyl, 2'-O-propylamine, 2'-amino, 2'-ethyl, EA, 2'-OMe, 2'-MOE, 2'-O-NMA, and 2'-FANA.

[0595] In some embodiments, the oligonucleotide comprises an antisense strand having a sugar moiety with each nucleotide at positions 2, 4, 5, and 14 of the antisense strand modified with a 2'-F modified sugar moiety and each remaining nucleotide of the antisense strand modified with a modification selected from the group consisting of: 2'-O-propargyl, 2'-O-propylamine, 2'-amino, 2'-ethyl, EA, 2'-OMe, 2'-MOE, 2'-O-NMA, and 2'-FANA.

[0596] In some embodiments, the oligonucleotide comprises an antisense strand having a sugar moiety with each nucleotide at positions 1, 2, 3, 5, 7, and 14 of the antisense strand modified with a 2'-F modified sugar moiety and each remaining nucleotide of the antisense strand modified with a modification selected from the group consisting of: 2'-O-propargyl, 2'-O-propylamine, 2'-amino, 2'-ethyl, EA, 2'-OMe, 2'-MOE, 2'-O-NMA, and 2'-FANA.

[0597] In some embodiments, the oligonucleotide comprises an antisense strand having a sugar moiety with each nucleotide at positions 2, 3, 4, 5, 7, and 14 of the antisense strand modified with a 2'-F modified sugar moiety and each remaining nucleotide of the antisense strand modified with a modification selected from the group consisting of: 2'-O-propargyl, 2'-O-propylamine, 2'-amino, 2'-ethyl, EA, 2'-OMe, 2'-MOE, 2'-O-NMA, and 2'-FANA.

[0598] In some embodiments, the oligonucleotide comprises an antisense strand having a sugar moiety with each nucleotide at positions 1, 2, 3, 5, 10, and 14 of the antisense strand modified with a 2'-F modified sugar moiety and each remaining nucleotide of the antisense strand modified with a modification selected from the group consisting of: 2'-O-propargyl, 2'-O-propylamine, 2'-amino, 2'-ethyl, EA, 2'-OMe, 2'-MOE, 2'-O-NMA, and 2'-FANA.

[0599] In some embodiments, the oligonucleotide comprises an antisense strand having a sugar moiety with each nucleotide at positions 2, 3, 4, 5, 10, and 14 of the antisense strand modified with a 2'-F modified sugar moiety and each remaining nucleotide of the antisense strand modified with a modification selected from the group consisting of: 2'-O-propargyl, 2'-O-propylamine, 2'-amino, 2'-ethyl, EA, 2'-OMe, 2'-MOE, 2'-O-NMA, and 2'-FANA.

[0600] In some embodiments, the oligonucleotide comprises an antisense strand having a sugar moiety with each nucleotide at positions 2, 3, 5, 7, 10, and 14 of the antisense strand modified with a 2'-F modified sugar moiety and each remaining nucleotide of the antisense strand modified with a modification selected from the group consisting of: 2'-O-propargyl, 2'-O-propylamine, 2'-amino, 2'-ethyl, EA, 2'-OMe, 2'-MOE, 2'-O-NMA, and 2'-FANA.

[0601] In some embodiments, the oligonucleotide comprises a sense strand and an antisense strand of 36 nucleotides, wherein the antisense strand comprises a sugar moiety modified with 2'-F at positions 2, 3, 4, 5, 7, 10, and 14 of the antisense strand and each of the remaining nucleotides of the antisense strand is modified with a sugar moiety modified with a modification selected from the group consisting of: 2'-O-propargyl, 2'-O-propylamine, 2'-amino, 2'-ethyl, EA, 2'-OMe, 2'-MOE, 2'-O-NMA, and 2'-FANA. In some embodiments, the oligonucleotide comprises a sense strand and an antisense strand comprising a stem-loop, wherein the antisense strand comprises a sugar moiety modified with 2'-F at positions 2, 3, 4, 5, 7, 10, and 14 of the antisense strand and each of the remaining nucleotides of the antisense strand is modified with a sugar moiety modified with a modification selected from the group consisting of: 2'-O-propargyl, 2'-O-propylamine, 2'-amino, 2'-ethyl, EA, 2'-OMe, 2'-MOE, 2'-O-NMA, and 2'-FANA.

[0602] In some embodiments, the oligonucleotide comprises an antisense strand having a sugar moiety with each nucleotide at positions 2, 3, 4, 5, 7, 10, 14, 16, and 19 of the antisense strand modified with a 2'-F modified sugar moiety and each remaining nucleotide of the antisense strand modified with a modification selected from the group consisting of: 2'-O-propargyl, 2'-O-propylamine, 2'-amino, 2'-ethyl, EA, 2'-OMe, 2'-MOE, 2'-O-NMA, and 2'-FANA.

[0603] In some embodiments, the oligonucleotide comprises an antisense strand having a sugar moiety modified with a 2'-F at position 1, position 2, position 3, position 4, position 5, position 6, position 7, position 8, position 9, position 10, position 11, position 12, position 13, position 14, position 15, position 16, position 17, position 18, position 19, position 20, position 21, or position 22.

[0604] In some embodiments, the oligonucleotide comprises an antisense strand having a sugar moiety modified with 2'-OMe at position 1, position 2, position 3, position 4, position 5, position 6, position 7, position 8, position 9, position 10, position 11, position 12, position 13, position 14, position 15, position 16, position 17, position 18, position 19, position 20, position 21, or position 22.

[0605] In some embodiments, the oligonucleotide comprises an antisense strand having a sugar moiety modified at position 1, position 2, position 3, position 4, position 5, position 6, position 7, position 8, position 9, position 10, position 11, position 12, position 13, position 14, position 15, position 16, position 17, position 18, position 19, position 20, position 21, or position 22 with a modification selected from the group consisting of: 2'-O-propargyl, 2'-O-propylamine, 2'-amino, 2'-ethyl, EA, 2'-OMe, 2'-MOE, 2'-O-NMA, and 2'-FANA.

[0606] In some embodiments, the oligonucleotide comprises a sense strand of 36 nucleotides having a sugar moiety modified with 2'-F at positions 8-11. In some embodiments, the oligonucleotide comprises a sense strand comprising a stem-loop and a sugar moiety modified with 2'-F at positions 8-11. In some embodiments, the oligonucleotide comprises a sense strand of 36 nucleotides having a sugar moiety modified with 2'-OMe at positions 1-7 and 12-17 or 12-20. In some embodiments, the oligonucleotide comprises a sense strand comprising a stem-loop and a sugar moiety modified with 2'-OMe at positions 1-7 and 12-17 or 12-20. In some embodiments, the oligonucleotide comprises a sense strand of 36 nucleotides having a sugar moiety modified with 2'-OMe at positions 1-7 and 12-17, 12-20, or 12-22. In some embodiments, the oligonucleotide comprises a sense strand comprising a stem-loop and a sugar moiety modified with 2'-OMe at positions 1-7 and 12-17, 12-20, or 12-22. In some embodiments, the oligonucleotide comprises a 36-nucleotide sense strand having a sugar moiety modified with a modification selected from the group consisting of 2'-O-propargyl, 2'-O-propylamine, 2'-amino, 2'-ethyl, EA, 2'-OMe, 2'-MOE, 2'-O-NMA, and 2'-FANA at each nucleotide at positions 1-7 and 12-17 or 12-20 of the sense strand. In some embodiments, the oligonucleotide comprises a sense strand comprising a stem-loop and having a sugar moiety modified with a modification selected from the group consisting of: 2'-O-propargyl, 2'-O-propylamine, 2'-amino, 2'-ethyl, EA, 2'-OMe, 2'-MOE, 2'-O-NMA, and 2'-FANA at each nucleotide at positions 1-7 and 12-17, 12-20, or 12-22 of the sense strand. In some embodiments, the oligonucleotide comprises a 36-nucleotide sense strand having a sugar moiety modified with a modification selected from the group consisting of: 2'-O-propargyl, 2'-O-propylamine, 2'-amino, 2'-ethyl, EA, 2'-OMe, 2'-MOE, 2'-O-NMA, and 2'-FANA at each nucleotide at positions 1-7 and 12-17, 12-20, or 12-22 of the sense strand. In some embodiments, the oligonucleotide comprises a sense strand comprising a stem-loop and a sugar moiety wherein each nucleotide at positions 1-7 and 12-17, 12-20, or 12-22 of the sense strand is modified with a modification selected from the group consisting of: 2'-O-propargyl, 2'-O-propylamine, 2'-amino, 2'-ethyl, EA, 2'-OMe, 2'-MOE, 2'-O-NMA, and 2'-FANA.

[0607] In some embodiments, the oligonucleotide comprises a sense strand having a sugar moiety modified with 2'-F at positions 3, 5, 8, 10, 12, 13, 15, and 17. In some embodiments, the oligonucleotide comprises a sense strand having a sugar moiety modified with 2'-OMe at positions 1, 2, 4, 6, 7, 9, 11, 14, 16, and 18-20. In some embodiments, the oligonucleotide comprises a sense strand having a sugar moiety modified with a modification selected from the group consisting of 2'-O-propargyl, 2'-O-propylamine, 2'-amino, 2'-ethyl, EA, 2'-OMe, 2'-MOE, 2'-O-NMA, and 2'-FANA at each nucleotide at positions 1, 2, 4, 6, 7, 9, 11, 14, 16, and 18-20 of the sense strand. In some embodiments, the oligonucleotide comprises a sense strand having a sugar moiety modified with a modification selected from the group consisting of: 2'-O-propargyl, 2'-O-propylamine, 2'-amino, 2'-ethyl, EA, 2'-OMe, 2'-MOE, 2'-O-NMA, and 2'-FANA at each nucleotide at positions 1-7 and 12-17, 12-20, or 12-22 of the sense strand.

[0608] In some embodiments, the oligonucleotide comprises a sense strand having a sugar moiety with a 2'-F modification at position 1, position 2, position 3, position 4, position 5, position 6, position 7, position 8, position 9, position 10, position 11, position 12, position 13, position 14, position 15, position 16, position 17, position 18, position 19, position 20, position 21, position 22, position 23, position 24, position 25, position 26, position 27, position 28, position 29, position 30, position 31, position 32, position 33, position 34, position 35, or position 36.

[0609] In some embodiments, the oligonucleotide comprises a sense strand having a sugar moiety modified with 2'-OMe at position 1, position 2, position 3, position 4, position 5, position 6, position 7, position 8, position 9, position 10, position 11, position 12, position 13, position 14, position 15, position 16, position 17, position 18, position 19, position 20, position 21, position 22, position 23, position 24, position 25, position 26, position 27, position 28, position 29, position 30, position 31, position 32, position 33, position 34, position 35, or position 36.

[0610] In some embodiments, the oligonucleotide comprises a sense strand having a sugar moiety modified at position 1, position 2, position 3, position 4, position 5, position 6, position 7, position 8, position 9, position 10, position 11, position 12, position 13, position 14, position 15, position 16, position 17, position 18, position 19, position 20, position 21, position 22, position 23, position 24, position 25, position 26, position 27, position 28, position 29, position 30, position 31, position 32, position 33, position 34, position 35, or position 36 with a modification selected from the group consisting of 2'-O-propargyl, 2'-O-propylamine, 2'-amino, 2'-ethyl, EA, 2'-OMe, 2'-MOE, 2'-O-NMA, and 2'-FANA.

[0611] In some embodiments, the oligonucleotide comprises an antisense strand having a sugar moiety with each nucleotide at positions 2, 3, 4, 5, 7, 10, and 14 of the antisense strand modified with a 2'-F modified sugar moiety and each remaining nucleotide of the antisense strand modified with a modification selected from the group consisting of: 2'-O-propargyl, 2'-O-propylamine, 2'-amino, 2'-ethyl, EA, 2'-OMe, 2'-MOE, 2'-O-NMA, and 2'-deoxy-2'-fluoro-β-d-arabinonucleotide ( and a sense strand of 36 nucleotides having a sugar moiety modified with 2'-F at each nucleotide at positions 8-11 of the sense strand and a sugar moiety modified with a modification selected from the group consisting of 2'-O-propargyl, 2'-O-propylamine, 2'-amino, 2'-ethyl, EA, 2'-OMe, 2'-MOE, 2'-O-NMA, and 2'-deoxy-2'-fluoro-β-d-arabino nucleic acid (2'-FANA). In some embodiments, the oligonucleotide comprises an antisense strand having a sugar moiety with each nucleotide at positions 2, 3, 4, 5, 7, 10, and 14 of the antisense strand modified with a 2'-F modified sugar moiety and each remaining nucleotide of the antisense strand modified with a modification selected from the group consisting of: 2'-O-propargyl, 2'-O-propylamine, 2'-amino, 2'-ethyl, EA, 2'-OMe, 2'-MOE, 2'-O-NMA, and 2'-FANA; and a sense strand comprising a stem-loop and a sugar moiety with each nucleotide at positions 8-11 of the sense strand modified with a 2'-F modified sugar moiety and each remaining nucleotide of the antisense strand modified with a modification selected from the group consisting of: 2'-O-propargyl, 2'-O-propylamine, 2'-amino, 2'-ethyl, 2'-EA, 2'-OMe, 2'-MOE, 2'-O-NMA, and 2'-FANA.

[0612] In some embodiments, the oligonucleotide comprises an antisense strand having a sugar moiety with each nucleotide at positions 2, 3, 4, 5, 7, 10, 14, 16, and 19 of the antisense strand modified with a 2'-F modified sugar moiety and each remaining nucleotide of the antisense strand modified with a modification selected from the group consisting of: 2'-O-propargyl, 2'-O-propylamine, 2'-amino, 2'-ethyl, EA, 2'-OMe, 2'-MOE, 2'-O-NMA, and 2'-O- '-FANA; and a sense strand having a sugar moiety with each nucleotide at positions 3, 5, 8, 10, 12, 13, 15 and 17 of the sense strand modified with a 2'-F modified sugar moiety and each remaining nucleotide of the antisense strand modified with a sugar moiety selected from the group consisting of: 2'-O-propargyl, 2'-O-propylamine, 2'-amino, 2'-ethyl, EA, 2'-OMe, 2'-MOE, 2'-O-NMA and 2'-FANA.

[0613] In some embodiments, the sense strand and the antisense strand of the oligonucleotide comprise a nucleotide sequence selected from the group consisting of:

[0614] a) SEQ ID NO: 769 and 804, respectively;

[0615] b) SEQ ID NOs: 770 and 805, respectively;

[0616] c) SEQ ID NOs: 771 and 806, respectively;

[0617] d) SEQ ID NOs: 772 and 807, respectively;

[0618] e) SEQ ID NOs: 773 and 808, respectively;

[0619] f) SEQ ID NOs: 774 and 809, respectively;

[0620] g) SEQ ID NOs: 775 and 810, respectively;

[0621] h) SEQ ID NOs: 776 and 811, respectively;

[0622] i) SEQ ID NOs: 777 and 812, respectively;

[0623] j) SEQ ID NOs: 778 and 813, respectively;

[0624] k) SEQ ID NOs: 779 and 814, respectively;

[0625] 1) are SEQ ID NOs: 780 and 815, respectively;

[0626] m) are SEQ ID NOs: 781 and 816, respectively;

[0627] n) are SEQ ID NOs: 782 and 817, respectively;

[0628] o) are SEQ ID NOs: 783 and 818, respectively;

[0629] p) are SEQ ID NOs: 784 and 819, respectively;

[0630] q) are SEQ ID NOs: 785 and 820, respectively;

[0631] r) are SEQ ID NOs: 786 and 821, respectively;

[0632] s) are SEQ ID NOs: 787 and 822, respectively;

[0633] t) are SEQ ID NOs: 788 and 823, respectively;

[0634] u) are SEQ ID NOs: 789 and 824, respectively;

[0635] v) SEQ ID NOs: 790 and 825, respectively;

[0636] w) are SEQ ID NOs: 791 and 826, respectively;

[0637] x) are SEQ ID NOs: 792 and 827, respectively;

[0638] y) are SEQ ID NOs: 793 and 828, respectively;

[0639] z) are SEQ ID NOs: 794 and 829, respectively;

[0640] aa) are SEQ ID NOs: 795 and 830, respectively;

[0641] bb) are SEQ ID NOs: 796 and 831, respectively;

[0642] cc) are SEQ ID NOs: 797 and 832, respectively;

[0643] dd) are SEQ ID NOs: 798 and 833, respectively;

[0644] ee) are SEQ ID NOs: 799 and 834, respectively;

[0645] ff) are SEQ ID NOs: 800 and 835, respectively;

[0646] gg) are SEQ ID NOs: 801 and 836, respectively;

[0647] hh) are SEQ ID NOs: 802 and 837, respectively;

[0648] ii) SEQ ID NOs: 803 and 838, respectively; and

[0649] jj) are SEQ ID NOs: 1681 and 815, respectively, wherein one or more of positions 3, 5, 8, 10, 12, 13, 15 or 17 of the sense strand are modified with a 2'-F group.

[0650] In some embodiments, the sense strand and the antisense strand of the oligonucleotide comprise a nucleotide sequence selected from the group consisting of:

[0651] a) SEQ ID NO: 771 and 806, respectively;

[0652] b) SEQ ID NOs: 776 and 811, respectively;

[0653] c) SEQ ID NOs: 780 and 815, respectively;

[0654] d) SEQ ID NOs: 781 and 816, respectively;

[0655] e) SEQ ID NOs: 782 and 817, respectively;

[0656] f) SEQ ID NOs: 790 and 825, respectively;

[0657] g) SEQ ID NOs: 795 and 830, respectively;

[0658] h) SEQ ID NOs: 798 and 833, respectively;

[0659] i) SEQ ID NOs: 799 and 834, respectively;

[0660] j) SEQ ID NOs: 803 and 838, respectively; and

[0661] k) are SEQ ID NOs: 1681 and 815, respectively, wherein one or more of positions 3, 5, 8, 10, 12, 13, 15 or 17 of the sense strand are modified with a 2'-F group.

[0662] In some embodiments, the sense strand and the antisense strand of the oligonucleotide comprise a nucleotide sequence selected from the group consisting of:

[0663] a) SEQ ID NO: 771 and 806, respectively;

[0664] b) SEQ ID NOs: 780 and 815, respectively;

[0665] c) SEQ ID NOs: 781 and 816, respectively;

[0666] d) SEQ ID NOs: 798 and 833, respectively;

[0667] e) SEQ ID NOs: 799 and 834, respectively;

[0668] f) SEQ ID NOs: 803 and 838, respectively; and

[0669] g) are SEQ ID NOs: 1681 and 815, respectively, wherein one or more of positions 3, 5, 8, 10, 12, 13, 15 or 17 of the sense strand are modified with a 2'-F group.

[0670] 5' terminal phosphate

[0671] In some embodiments, the oligonucleotide comprises a sense strand and an antisense strand, wherein the antisense strand comprises a 5' terminal phosphate. In some embodiments, the 5' terminal phosphate group of the oligonucleotide enhances the interaction with Ago2. However, the oligonucleotide comprising a 5'-phosphate group may be easily degraded via phosphatases or other enzymes, which may limit its in vivo bioavailability. In some embodiments, the oligonucleotide includes a 5'-phosphate analog that is resistant to this degradation. In some embodiments, the phosphate analog is an oxygen methyl phosphonate, a vinyl phosphonate or a malonyl phosphonate or a combination thereof. In certain embodiments, the 5' end of the oligonucleotide chain is attached to a chemical part that simulates the electrostatic and spatial properties of a natural 5'-phosphate group ("phosphate mimics").

[0672] In some embodiments, the oligonucleotide has a phosphate analog (referred to as a "4'-phosphate analog") at the 4'-carbon position of the sugar. See, for example, International Patent Application Publication No. WO 2018 / 045317. In some embodiments, the oligonucleotide comprises a 4'-phosphate analog at the 5' terminal nucleotide. In some embodiments, the phosphate analog is an oxymethylphosphonate, wherein the oxygen atom of the oxymethyl is bound to a sugar moiety (e.g., at its 4'-carbon) or an analog thereof. In other embodiments, the 4'-phosphate analog is a thiomethylphosphonate or an aminomethylphosphonate, wherein the sulfur atom of the thiomethyl or the nitrogen atom of the aminomethyl is bound to the 4'-carbon of a sugar moiety or its analog. In certain embodiments, the 4'-phosphate analog is an oxymethylphosphonate. In some embodiments, the oxymethylphosphonate is represented by the formula -O-CH2-PO(OH)2, -O-CH2-PO(OR)2, or -O-CH2-POOH(R), wherein R is independently selected from H, CH3, alkyl, CH2CH2CN, CH2OCOC(CH3)3, CHOCH2CH2Si(CH3)3, or a protecting group. In certain embodiments, the alkyl is CH2CH3. More typically, R is independently selected from H, CH3, or CH2CH3. In some embodiments, R is CH3. In some embodiments, the 4'-phosphate analog is 4'-oxymethylphosphonate. In some embodiments, the modified nucleotide having a 4'-phosphonate analog is uridine. In some embodiments, the modified nucleotide is 4'-O-monomethylphosphonate-2'-O-methyluridine.

[0673] In some embodiments, the sense strand and the antisense strand of the oligonucleotide comprise a nucleotide sequence selected from the group consisting of:

[0674] a) SEQ ID NO: 769 and 804, respectively;

[0675] b) SEQ ID NOs: 770 and 805, respectively;

[0676] c) SEQ ID NOs: 771 and 806, respectively;

[0677] d) SEQ ID NOs: 772 and 807, respectively;

[0678] e) SEQ ID NOs: 773 and 808, respectively;

[0679] f) SEQ ID NOs: 774 and 809, respectively;

[0680] g) SEQ ID NOs: 775 and 810, respectively;

[0681] h) SEQ ID NOs: 776 and 811, respectively;

[0682] i) SEQ ID NOs: 777 and 812, respectively;

[0683] j) SEQ ID NOs: 778 and 813, respectively;

[0684] k) SEQ ID NOs: 779 and 814, respectively;

[0685] 1) are SEQ ID NOs: 780 and 815, respectively;

[0686] m) are SEQ ID NOs: 781 and 816, respectively;

[0687] n) are SEQ ID NOs: 782 and 817, respectively;

[0688] o) are SEQ ID NOs: 783 and 818, respectively;

[0689] p) are SEQ ID NOs: 784 and 819, respectively;

[0690] q) are SEQ ID NOs: 785 and 820, respectively;

[0691] r) are SEQ ID NOs: 786 and 821, respectively;

[0692] s) are SEQ ID NOs: 787 and 822, respectively;

[0693] t) are SEQ ID NOs: 788 and 823, respectively;

[0694] u) are SEQ ID NOs: 789 and 824, respectively;

[0695] v) SEQ ID NOs: 790 and 825, respectively;

[0696] w) are SEQ ID NOs: 791 and 826, respectively;

[0697] x) are SEQ ID NOs: 792 and 827, respectively;

[0698] y) are SEQ ID NOs: 793 and 828, respectively;

[0699] z) are SEQ ID NOs: 794 and 829, respectively;

[0700] aa) are SEQ ID NOs: 795 and 830, respectively;

[0701] bb) are SEQ ID NOs: 796 and 831, respectively;

[0702] cc) are SEQ ID NOs: 797 and 832, respectively;

[0703] dd) are SEQ ID NOs: 798 and 833, respectively;

[0704] ee) are SEQ ID NOs: 799 and 834, respectively;

[0705] ff) are SEQ ID NOs: 800 and 835, respectively;

[0706] gg) are SEQ ID NOs: 801 and 836, respectively;

[0707] hh) are SEQ ID NOs: 802 and 837, respectively;

[0708] ii) SEQ ID NOs: 803 and 838, respectively; and

[0709] jj) are SEQ ID NOs: 1681 and 815, respectively, wherein the oligonucleotide comprises a 5' terminal phosphate, optionally a 5' terminal phosphate analog.

[0710] In some embodiments, the sense strand and the antisense strand of the oligonucleotide comprise a nucleotide sequence selected from the group consisting of:

[0711] a) SEQ ID NO: 771 and 806, respectively;

[0712] b) SEQ ID NOs: 776 and 811, respectively;

[0713] c) SEQ ID NOs: 780 and 815, respectively;

[0714] d) SEQ ID NOs: 781 and 816, respectively;

[0715] e) SEQ ID NOs: 782 and 817, respectively;

[0716] f) SEQ ID NOs: 790 and 825, respectively;

[0717] g) SEQ ID NOs: 795 and 830, respectively;

[0718] h) SEQ ID NOs: 798 and 833, respectively;

[0719] i) SEQ ID NOs: 799 and 834, respectively;

[0720] j) SEQ ID NOs: 803 and 838, respectively; and

[0721] k) are SEQ ID NO: 1681 and 815, respectively, wherein the oligonucleotide comprises a 5' terminal phosphate, optionally a 5' terminal phosphate analog.

[0722] In some embodiments, the sense strand and the antisense strand of the oligonucleotide comprise a nucleotide sequence selected from the group consisting of:

[0723] a) SEQ ID NO: 771 and 806, respectively;

[0724] b) SEQ ID NOs: 780 and 815, respectively;

[0725] c) SEQ ID NOs: 781 and 816, respectively;

[0726] d) SEQ ID NOs: 798 and 833, respectively;

[0727] e) SEQ ID NOs: 799 and 834, respectively;

[0728] f) SEQ ID NOs: 803 and 838, respectively; and

[0729] g) are SEQ ID NO: 1681 and 815, respectively, wherein the oligonucleotide comprises a 5' terminal phosphate, optionally a 5' terminal phosphate analog.

[0730] In some embodiments, the oligonucleotide comprises an antisense strand comprising a 4'-phosphate analog at the 5' terminal nucleotide, wherein the 5' terminal nucleotide comprises the following structure:

[0731]

[0732] 4'-O-monomethylphosphonate-2'-O-methyluridine phosphorothioate [Mephosphonate-4O-mUs].

[0733] Modified internucleotide bonds

[0734] In some embodiments, the oligonucleotides herein (e.g., RNAi oligonucleotides) comprise modified internucleotide bonds. In some embodiments, phosphate modification or substitution produces an oligonucleotide comprising at least about 1 (e.g., at least 1, at least 2, at least 3, or at least 5) modified internucleotide bonds. In some embodiments, the oligonucleotide comprises about 1 to about 10 (e.g., 1 to 10, 2 to 8, 4 to 6, 3 to 10, 5 to 10, 1 to 5, 1 to 3, or 1 to 2) modified internucleotide bonds. In some embodiments, the oligonucleotide comprises 1, 2, 3, 4, 5, 6, 7, 8, 9, or 10 modified internucleotide bonds.

[0735] The modified internucleotide bond can be a phosphorodithioate bond, a phosphorothioate bond, a phosphotriester bond, a thiocarbonylalkylphosphonate bond, a thiocarbonylalkylphosphotriester bond, a phosphoramidite bond, a phosphonate bond, or a boranophosphate bond. In some embodiments, at least one modified internucleotide bond of an oligonucleotide is a phosphorothioate bond.

[0736] In some embodiments, the oligonucleotide has a phosphorothioate bond between one or more of positions 1 and 2 of the sense strand, positions 1 and 2 of the antisense strand, positions 2 and 3 of the antisense strand, positions 3 and 4 of the antisense strand, positions 20 and 21 of the antisense strand, and positions 21 and 22 of the antisense strand. In some embodiments, the oligonucleotides described herein have a phosphorothioate bond between each of positions 1 and 2 of the sense strand, positions 1 and 2 of the antisense strand, positions 2 and 3 of the antisense strand, positions 20 and 21 of the antisense strand, and positions 21 and 22 of the antisense strand. In some embodiments, the oligonucleotides described herein have a phosphorothioate bond between each of: (i) positions 1 and 2 of the sense strand; and (ii) positions 1 and 2, positions 2 and 3, positions 3 and 4, positions 20 and 21, and positions 21 and 22 of the antisense strand.

[0737] In some embodiments, the sense strand and the antisense strand of the oligonucleotide comprise a nucleotide sequence selected from the group consisting of:

[0738] a) SEQ ID NO: 769 and 804, respectively;

[0739] b) SEQ ID NOs: 770 and 805, respectively;

[0740] c) SEQ ID NOs: 771 and 806, respectively;

[0741] d) SEQ ID NOs: 772 and 807, respectively;

[0742] e) SEQ ID NOs: 773 and 808, respectively;

[0743] f) SEQ ID NOs: 774 and 809, respectively;

[0744] g) SEQ ID NOs: 775 and 810, respectively;

[0745] h) SEQ ID NOs: 776 and 811, respectively;

[0746] i) SEQ ID NOs: 777 and 812, respectively;

[0747] j) SEQ ID NOs: 778 and 813, respectively;

[0748] k) SEQ ID NOs: 779 and 814, respectively;

[0749] 1) are SEQ ID NOs: 780 and 815, respectively;

[0750] m) are SEQ ID NOs: 781 and 816, respectively;

[0751] n) are SEQ ID NOs: 782 and 817, respectively;

[0752] o) are SEQ ID NOs: 783 and 818, respectively;

[0753] p) are SEQ ID NOs: 784 and 819, respectively;

[0754] q) are SEQ ID NOs: 785 and 820, respectively;

[0755] r) are SEQ ID NOs: 786 and 821, respectively;

[0756] s) are SEQ ID NOs: 787 and 822, respectively;

[0757] t) are SEQ ID NOs: 788 and 823, respectively;

[0758] u) are SEQ ID NOs: 789 and 824, respectively;

[0759] v) SEQ ID NOs: 790 and 825, respectively;

[0760] w) are SEQ ID NOs: 791 and 826, respectively;

[0761] x) are SEQ ID NOs: 792 and 827, respectively;

[0762] y) are SEQ ID NOs: 793 and 828, respectively;

[0763] z) are SEQ ID NOs: 794 and 829, respectively;

[0764] aa) are SEQ ID NOs: 795 and 830, respectively;

[0765] bb) are SEQ ID NOs: 796 and 831, respectively;

[0766] cc) are SEQ ID NOs: 797 and 832, respectively;

[0767] dd) are SEQ ID NOs: 798 and 833, respectively;

[0768] ee) are SEQ ID NOs: 799 and 834, respectively;

[0769] ff) are SEQ ID NOs: 800 and 835, respectively;

[0770] gg) are SEQ ID NOs: 801 and 836, respectively;

[0771] hh) are SEQ ID NOs: 802 and 837, respectively;

[0772] ii) SEQ ID NOs: 803 and 838, respectively; and

[0773] jj) are SEQ ID NOs: 1681 and 815, respectively, wherein the oligonucleotide comprises a modified internucleotide bond.

[0774] In some embodiments, the sense strand and the antisense strand of the oligonucleotide comprise a nucleotide sequence selected from the group consisting of:

[0775] a) SEQ ID NO: 771 and 806, respectively;

[0776] b) SEQ ID NOs: 776 and 811, respectively;

[0777] c) SEQ ID NOs: 780 and 815, respectively;

[0778] d) SEQ ID NOs: 781 and 816, respectively;

[0779] e) SEQ ID NOs: 782 and 817, respectively;

[0780] f) SEQ ID NOs: 790 and 825, respectively;

[0781] g) SEQ ID NOs: 795 and 830, respectively;

[0782] h) SEQ ID NOs: 798 and 833, respectively;

[0783] i) SEQ ID NOs: 799 and 834, respectively;

[0784] j) SEQ ID NOs: 803 and 838, respectively; and

[0785] k) are SEQ ID NOs: 1681 and 815, respectively, wherein the oligonucleotide comprises a modified internucleotide bond.

[0786] In some embodiments, the sense strand and the antisense strand of the oligonucleotide comprise a nucleotide sequence selected from the group consisting of:

[0787] a) SEQ ID NO: 771 and 806, respectively;

[0788] b) SEQ ID NOs: 780 and 815, respectively;

[0789] c) SEQ ID NOs: 781 and 816, respectively;

[0790] d) SEQ ID NOs: 798 and 833, respectively;

[0791] e) SEQ ID NOs: 799 and 834, respectively;

[0792] f) SEQ ID NOs: 803 and 838, respectively; and

[0793] g) are SEQ ID NOs: 1681 and 815, respectively, wherein the oligonucleotide comprises a modified internucleotide bond.

[0794] Base modification

[0795] In some embodiments, the oligonucleotides herein (e.g., RNAi oligonucleotides) have one or more modified nucleobases. In some embodiments, the modified nucleobases (also referred to herein as base analogs) are connected at the 1' position of the nucleotide sugar moiety. In certain embodiments, the modified nucleobases are nitrogenous bases. In certain embodiments, the modified nucleobases do not contain nitrogen atoms. See, e.g., U.S. Patent Application Publication No. 2008 / 0274462. In some embodiments, the modified nucleotides include universal bases. In some embodiments, the modified nucleotides do not contain nucleobases (abasic).

[0796] In some embodiments, the sense strand and the antisense strand of the oligonucleotide comprise a nucleotide sequence selected from the group consisting of:

[0797] a) SEQ ID NO: 769 and 804, respectively;

[0798] b) SEQ ID NOs: 770 and 805, respectively;

[0799] c) SEQ ID NOs: 771 and 806, respectively;

[0800] d) SEQ ID NOs: 772 and 807, respectively;

[0801] e) SEQ ID NOs: 773 and 808, respectively;

[0802] f) SEQ ID NOs: 774 and 809, respectively;

[0803] g) SEQ ID NOs: 775 and 810, respectively;

[0804] h) SEQ ID NOs: 776 and 811, respectively;

[0805] i) SEQ ID NOs: 777 and 812, respectively;

[0806] j) SEQ ID NOs: 778 and 813, respectively;

[0807] k) SEQ ID NOs: 779 and 814, respectively;

[0808] 1) are SEQ ID NOs: 780 and 815, respectively;

[0809] m) are SEQ ID NOs: 781 and 816, respectively;

[0810] n) are SEQ ID NOs: 782 and 817, respectively;

[0811] o) are SEQ ID NOs: 783 and 818, respectively;

[0812] p) are SEQ ID NOs: 784 and 819, respectively;

[0813] q) are SEQ ID NOs: 785 and 820, respectively;

[0814] r) are SEQ ID NOs: 786 and 821, respectively;

[0815] s) are SEQ ID NOs: 787 and 822, respectively;

[0816] t) are SEQ ID NOs: 788 and 823, respectively;

[0817] u) are SEQ ID NOs: 789 and 824, respectively;

[0818] v) SEQ ID NOs: 790 and 825, respectively;

[0819] w) are SEQ ID NOs: 791 and 826, respectively;

[0820] x) are SEQ ID NOs: 792 and 827, respectively;

[0821] y) are SEQ ID NOs: 793 and 828, respectively;

[0822] z) are SEQ ID NOs: 794 and 829, respectively;

[0823] aa) are SEQ ID NOs: 795 and 830, respectively;

[0824] bb) are SEQ ID NOs: 796 and 831, respectively;

[0825] cc) are SEQ ID NOs: 797 and 832, respectively;

[0826] dd) are SEQ ID NOs: 798 and 833, respectively;

[0827] ee) are SEQ ID NOs: 799 and 834, respectively;

[0828] ff) are SEQ ID NOs: 800 and 835, respectively;

[0829] gg) are SEQ ID NOs: 801 and 836, respectively;

[0830] hh) are SEQ ID NOs: 802 and 837, respectively;

[0831] ii) SEQ ID NOs: 803 and 838, respectively; and

[0832] jj) are SEQ ID NOs: 1681 and 815, respectively, wherein the oligonucleotide comprises one or more modified nucleobases.

[0833] In some embodiments, the sense strand and the antisense strand of the oligonucleotide comprise a nucleotide sequence selected from the group consisting of:

[0834] a) SEQ ID NO: 771 and 806, respectively;

[0835] b) SEQ ID NOs: 776 and 811, respectively;

[0836] c) SEQ ID NOs: 780 and 815, respectively;

[0837] d) SEQ ID NOs: 781 and 816, respectively;

[0838] e) SEQ ID NOs: 782 and 817, respectively;

[0839] f) SEQ ID NOs: 790 and 825, respectively;

[0840] g) SEQ ID NOs: 795 and 830, respectively;

[0841] h) SEQ ID NOs: 798 and 833, respectively;

[0842] i) SEQ ID NOs: 799 and 834, respectively;

[0843] j) SEQ ID NOs: 803 and 838, respectively; and

[0844] k) are SEQ ID NOs: 1681 and 815, respectively, wherein the oligonucleotide comprises one or more modified nucleobases.

[0845] In some embodiments, the sense strand and the antisense strand of the oligonucleotide comprise a nucleotide sequence selected from the group consisting of:

[0846] a) SEQ ID NO: 771 and 806, respectively;

[0847] b) SEQ ID NOs: 780 and 815, respectively;

[0848] c) SEQ ID NOs: 781 and 816, respectively;

[0849] d) SEQ ID NOs: 798 and 833, respectively;

[0850] e) SEQ ID NOs: 799 and 834, respectively;

[0851] f) SEQ ID NOs: 803 and 838, respectively;

[0852] g) are SEQ ID NOs: 1681 and 815, respectively, wherein the oligonucleotide comprises one or more modified nucleobases.

[0853] In some embodiments, the universal base is a heterocyclic moiety located at the 1' position of the nucleotide sugar moiety in the modified nucleotide or at an equivalent position in a nucleotide sugar moiety substitution, which, when present in a duplex, can be positioned relative to more than one type of base without substantially altering the structure of the duplex. In some embodiments, the ss nucleic acid containing the universal base forms a duplex with the target nucleic acid, compared to a reference ss nucleic acid (e.g., an oligonucleotide) that is fully complementary to the target nucleic acid, wherein the T of the duplex m In some embodiments, when compared to a reference ss nucleic acid in which the universal base has been replaced by a base to produce a single mismatch, the ss nucleic acid containing the universal base forms a duplex with the target nucleic acid, the duplex having a T of m Higher than duplexes formed with nucleic acids containing mismatched bases.

[0854] Non-limiting examples of universal binding nucleotides include, but are not limited to, inosine, 1-β-D-ribofuranosyl-5-nitroindole and / or 1-β-D-ribofuranosyl-3-nitropyrrole (see U.S. Patent Application Publication No. 2007 / 0254362; Van Aerschot et al. (1995) NUCLEIC ACIDS RES. 23:4363-4370; Loakes et al. (1995) NUCLEIC ACIDS RES. 23:2361-66; and Loakes and Brown (1994) NUCLEIC ACIDS RES. 22:4039-43).

[0855] Targeting ligands

[0856] In some embodiments, it is desirable to target an oligonucleotide (e.g., an RNAi oligonucleotide) to one or more cells or one or more organs. This strategy can help avoid undesirable effects in other organs, or avoid excessive loss of oligonucleotides to cells, tissues, or organs that will not benefit from oligonucleotides. Therefore, in some embodiments, oligonucleotides are modified to promote targeting and / or delivery to specific tissues, cells, or organs (e.g., to promote delivery of oligonucleotides to CNS). In some embodiments, an oligonucleotide comprises at least one nucleotide (e.g., 1, 2, 3, 4, 5, 6, or more nucleotides) conjugated to one or more targeting ligands. In some embodiments, the sense strand and antisense strand of an oligonucleotide comprise a nucleotide sequence selected from the group consisting of:

[0857] a) SEQ ID NO: 769 and 804, respectively;

[0858] b) SEQ ID NOs: 770 and 805, respectively;

[0859] c) SEQ ID NOs: 771 and 806, respectively;

[0860] d) SEQ ID NOs: 772 and 807, respectively;

[0861] e) SEQ ID NOs: 773 and 808, respectively;

[0862] f) SEQ ID NOs: 774 and 809, respectively;

[0863] g) SEQ ID NOs: 775 and 810, respectively;

[0864] h) SEQ ID NOs: 776 and 811, respectively;

[0865] i) SEQ ID NOs: 777 and 812, respectively;

[0866] j) SEQ ID NOs: 778 and 813, respectively;

[0867] k) SEQ ID NOs: 779 and 814, respectively;

[0868] 1) are SEQ ID NOs: 780 and 815, respectively;

[0869] m) are SEQ ID NOs: 781 and 816, respectively;

[0870] n) are SEQ ID NOs: 782 and 817, respectively;

[0871] o) are SEQ ID NOs: 783 and 818, respectively;

[0872] p) are SEQ ID NOs: 784 and 819, respectively;

[0873] q) are SEQ ID NOs: 785 and 820, respectively;

[0874] r) are SEQ ID NOs: 786 and 821, respectively;

[0875] s) are SEQ ID NOs: 787 and 822, respectively;

[0876] t) are SEQ ID NOs: 788 and 823, respectively;

[0877] u) are SEQ ID NOs: 789 and 824, respectively;

[0878] v) SEQ ID NOs: 790 and 825, respectively;

[0879] w) are SEQ ID NOs: 791 and 826, respectively;

[0880] x) are SEQ ID NOs: 792 and 827, respectively;

[0881] y) are SEQ ID NOs: 793 and 828, respectively;

[0882] z) are SEQ ID NOs: 794 and 829, respectively;

[0883] aa) are SEQ ID NOs: 795 and 830, respectively;

[0884] bb) are SEQ ID NOs: 796 and 831, respectively;

[0885] cc) are SEQ ID NOs: 797 and 832, respectively;

[0886] dd) are SEQ ID NOs: 798 and 833, respectively;

[0887] ee) are SEQ ID NOs: 799 and 834, respectively;

[0888] ff) are SEQ ID NOs: 800 and 835, respectively;

[0889] gg) are SEQ ID NOs: 801 and 836, respectively;

[0890] hh) are SEQ ID NOs: 802 and 837, respectively; and

[0891] ii) are SEQ ID NOs: 803 and 838, respectively, wherein the oligonucleotide comprises a targeting ligand conjugated to at least one nucleotide.

[0892] In some embodiments, the oligonucleotide comprises at least one nucleotide (e.g., 1, 2, 3, 4, 5, 6 or more nucleotides) conjugated to one or more targeting ligands. In some embodiments, the sense strand and the antisense strand of the oligonucleotide comprise a nucleotide sequence selected from the group consisting of:

[0893] a) SEQ ID NO: 771 and 806, respectively;

[0894] b) SEQ ID NOs: 776 and 811, respectively;

[0895] c) SEQ ID NOs: 780 and 815, respectively;

[0896] d) SEQ ID NOs: 781 and 816, respectively;

[0897] e) SEQ ID NOs: 782 and 817, respectively;

[0898] f) SEQ ID NOs: 790 and 825, respectively;

[0899] g) SEQ ID NOs: 795 and 830, respectively;

[0900] h) SEQ ID NOs: 798 and 833, respectively;

[0901] i) SEQ ID NOs: 799 and 834, respectively; and

[0902] j) are SEQ ID NOs: 803 and 838, respectively, wherein the oligonucleotide comprises a targeting ligand conjugated to at least one nucleotide.

[0903] In some embodiments, the oligonucleotide comprises at least one nucleotide (e.g., 1, 2, 3, 4, 5, 6 or more nucleotides) conjugated to one or more targeting ligands. In some embodiments, the sense strand and the antisense strand of the oligonucleotide comprise a nucleotide sequence selected from the group consisting of:

[0904] a) SEQ ID NO: 771 and 806, respectively;

[0905] b) SEQ ID NOs: 780 and 815, respectively;

[0906] c) SEQ ID NOs: 781 and 816, respectively;

[0907] d) SEQ ID NOs: 798 and 833, respectively;

[0908] e) SEQ ID NOs: 799 and 834, respectively; and

[0909] f) are SEQ ID NOs: 803 and 838, respectively, wherein the oligonucleotide comprises a targeting ligand conjugated to at least one nucleotide.

[0910] In some embodiments, the targeting ligand comprises a carbohydrate, an amino sugar, cholesterol, a peptide, a polypeptide, or a portion of a protein or a protein (e.g., an antibody or antibody fragment). In some embodiments, the targeting ligand is an aptamer. For example, the targeting ligand can be an RGD peptide for targeting tumor vasculature or glioma cells, a CREKA peptide for targeting tumor vasculature or stoma, transferrin, lactoferrin, or an aptamer targeting transferrin receptor expressed on CNS vasculature, or an anti-EGFR antibody targeting EGFR on glioma cells. In certain embodiments, the targeting ligand is one or more GalNAc moieties. In some embodiments, the targeting ligand is one or more lipid moieties.

[0911] In some embodiments, one or more (e.g., 1, 2, 3, 4, 5 or 6) nucleotides of an oligonucleotide are each conjugated to an independent targeting ligand. In some embodiments, 2 to 4 nucleotides of an oligonucleotide are each conjugated to an independent targeting ligand. In some embodiments, a targeting ligand is conjugated to 2 to 4 nucleotides at either end of a sense strand or an antisense strand (e.g., a targeting ligand is conjugated to 2 to 4 nucleotides at the 5' or 3' end of a sense strand or an antisense strand, overhanging or extending), so that the targeting ligand is similar to the bristles of a toothbrush and the oligonucleotide is similar to a toothbrush. For example, an oligonucleotide can include a stem loop at either end of the 5' or 3' end of the sense strand, and 1, 2, 3 or 4 nucleotides of the stem loop can be individually conjugated to a targeting ligand. In some embodiments, an oligonucleotide includes a stem loop at the 3' end of the sense strand, wherein the ring of the stem loop includes triL or tetraL, and wherein 3 or 4 nucleotides constituting triL or tetraL are individually conjugated to a targeting ligand, respectively. In some embodiments, the oligonucleotide comprises a blunt end at the 3' end of the oligonucleotide and one or more targeting ligands conjugated to at least one nucleotide. In some embodiments, the oligonucleotide comprises a blunt end at the 3' end of the oligonucleotide and one or more targeting ligands conjugated to the 5' terminal nucleotide of the sense strand.

[0912] GalNAc conjugation

[0913] GalNAc is a high affinity ligand of the asialoglycoprotein receptor (ASGPR), which is expressed primarily on the sinusoidal surface of hepatocytes and plays a major role in binding, internalization, and subsequent clearance of circulating glycoproteins (asialoglycoproteins) containing terminal galactose or GalNAc residues. The conjugation (indirect or direct) of the GalNAc moiety to the oligonucleotides herein can be used to target it to the ASGPR expressed on the cell. In some embodiments, the oligonucleotide is conjugated to at least one or more GalNAc moieties, wherein the GalNAc moieties target the oligonucleotide to the ASGPR expressed on human liver cells (e.g., human hepatocytes). In some embodiments, the GalNAc moieties target the oligonucleotide to the liver.

[0914] In some embodiments, the oligonucleotide is directly or indirectly conjugated to a monovalent GalNAc. In some embodiments, the oligonucleotide is directly or indirectly conjugated to more than one monovalent GalNAc (i.e., conjugated to 2, 3 or 4 monovalent GalNAc moieties, and generally conjugated to 3 or 4 monovalent GalNAc moieties). In some embodiments, the oligonucleotide is conjugated to one or more divalent GalNAc, trivalent GalNAc or tetravalent GalNAc moieties. In some embodiments, the divalent, trivalent or tetravalent GalNAc moieties are conjugated to the oligonucleotide via a branched joint. In some embodiments, the monovalent GalNAc moiety is conjugated to the first nucleotide, and the divalent, trivalent or tetravalent GalNAc moiety is conjugated to the second nucleotide via a branched joint.

[0915] In some embodiments, one or more (e.g., 1, 2, 3, 4, 5, or 6) nucleotides of an oligonucleotide are each conjugated to a GalNAc moiety. In some embodiments, 2 to 4 nucleotides of a tetraL are each conjugated to an independent GalNAc. In some embodiments, 1 to 3 nucleotides of a triL are each conjugated to an independent GalNAc. In some embodiments, a targeting ligand is conjugated to 2 to 4 nucleotides at either end of a sense strand or an antisense strand (e.g., a ligand is conjugated to 2 to 4 nucleotides overhanging or extending at the 5' or 3' end of a sense strand or an antisense strand), such that the GalNAc moiety is similar to the bristles of a toothbrush and the oligonucleotide is similar to a toothbrush. In some embodiments, the GalNAc moiety is conjugated to the nucleotides of the sense strand. For example, 4 GalNAc moieties can be conjugated to the nucleotides in the tetraL of the sense strand, wherein each GalNAc moiety is conjugated to 1 nucleotide.

[0916] In some embodiments, the oligonucleotide comprises tetraL, wherein tetraL is any combination of adenine (A) and guanine (G) nucleotides. In some embodiments, tetraL comprises a monovalent GalNAc moiety attached to any one or more guanine (G) nucleotides of the tetraloop via any linker described herein, as depicted below (X = heteroatom):

[0917]

[0918] In some embodiments, tetraL has a monovalent GalNAc attached to any one or more adenine nucleotides of the tetraloop via any linker described herein, as depicted below (X = heteroatom):

[0919]

[0920] In some embodiments, an oligonucleotide herein (e.g., an RNAi oligonucleotide) comprises a monovalent GalNAc attached to a guanine nucleotide, referred to as [ademG-GalNAc] or 2'-aminodiethoxymethanol-guanine-GalNAc, as depicted below:

[0921]

[0922] In some embodiments, the oligonucleotides herein comprise a monovalent GalNAc attached to an adenine nucleotide, referred to as [ademA-GalNAc] or 2'-aminodiethoxymethanol-adenine-GalNAc, as depicted below:

[0923]

[0924] An example of such a conjugation is shown below for a loop from 5' to 3' containing the nucleotide sequence GAAA (L = linker, X = heteroatom), wherein the stem attachment point is shown. Such a loop can be present, for example, at positions 27-30 of the sense strand of any of the sense strands listed in Tables 4 and 5. In the chemical formula, Used to describe the point of attachment to an oligonucleotide chain:

[0925]

[0926] The targeting ligand can be connected to the nucleotide using appropriate methods or chemistry (e.g., click chemistry). In some embodiments, the targeting ligand is conjugated to the nucleotide using a click linker. In some embodiments, an acetal-based linker is used to conjugate the targeting ligand to the nucleotide of any one of the oligonucleotides described herein. Acetal-based linkers are disclosed in, for example, International Patent Application Publication No. WO 2016 / 100401. In some embodiments, the linker is an unstable linker. However, in other embodiments, the linker is stable. The following shows an example of a loop comprising the nucleotide GAAA from 5' to 3', in which the GalNAc moiety is attached to 3 or 4 nucleotides of the loop using an acetal linker. Such a loop may be present at positions 27-30 of any one of the sense strands listed in, for example, Tables 4 and 5. In the chemical formula, is the point of attachment to the oligonucleotide chain:

[0927]

[0928]

[0929] As mentioned, the targeting ligand can be connected to the nucleotide using various appropriate methods or chemical synthesis techniques (e.g., click chemistry). In some embodiments, the targeting ligand is conjugated to the nucleotide using a click linker. In some embodiments, an acetal-based linker is used to conjugate the targeting ligand to the nucleotide of any one of the oligonucleotides described herein. Acetal-based linkers are disclosed in, for example, International Patent Application Publication No. WO 2016 / 100401. In some embodiments, the linker is an unstable linker. However, in other embodiments, the linker is a stable linker.

[0930] In some embodiments, a duplex extension (e.g., up to 3, 4, 5, or 6 bp in length) is provided between the targeting ligand (e.g., a GalNAc moiety) and the RNAi oligonucleotide. In some embodiments, the oligonucleotide herein does not have GalNAc conjugated thereto.

[0931] In some embodiments, the sense strand and the antisense strand of the oligonucleotide comprise a nucleotide sequence selected from the group consisting of:

[0932] a) SEQ ID NO: 769 and 804, respectively;

[0933] b) SEQ ID NOs: 770 and 805, respectively;

[0934] c) SEQ ID NOs: 771 and 806, respectively;

[0935] d) SEQ ID NOs: 772 and 807, respectively;

[0936] e) SEQ ID NOs: 773 and 808, respectively;

[0937] f) SEQ ID NOs: 774 and 809, respectively;

[0938] g) SEQ ID NOs: 775 and 810, respectively;

[0939] h) SEQ ID NOs: 776 and 811, respectively;

[0940] i) SEQ ID NOs: 777 and 812, respectively;

[0941] j) SEQ ID NOs: 778 and 813, respectively;

[0942] k) SEQ ID NOs: 779 and 814, respectively;

[0943] 1) are SEQ ID NOs: 780 and 815, respectively;

[0944] m) are SEQ ID NOs: 781 and 816, respectively;

[0945] n) are SEQ ID NOs: 782 and 817, respectively;

[0946] o) are SEQ ID NOs: 783 and 818, respectively;

[0947] p) are SEQ ID NOs: 784 and 819, respectively;

[0948] q) are SEQ ID NOs: 785 and 820, respectively;

[0949] r) are SEQ ID NOs: 786 and 821, respectively;

[0950] s) are SEQ ID NOs: 787 and 822, respectively;

[0951] t) are SEQ ID NOs: 788 and 823, respectively;

[0952] u) are SEQ ID NOs: 789 and 824, respectively;

[0953] v) SEQ ID NOs: 790 and 825, respectively;

[0954] w) are SEQ ID NOs: 791 and 826, respectively;

[0955] x) are SEQ ID NOs: 792 and 827, respectively;

[0956] y) are SEQ ID NOs: 793 and 828, respectively;

[0957] z) are SEQ ID NOs: 794 and 829, respectively;

[0958] aa) are SEQ ID NOs: 795 and 830, respectively;

[0959] bb) are SEQ ID NOs: 796 and 831, respectively;

[0960] cc) are SEQ ID NOs: 797 and 832, respectively;

[0961] dd) are SEQ ID NOs: 798 and 833, respectively;

[0962] ee) are SEQ ID NOs: 799 and 834, respectively;

[0963] ff) are SEQ ID NOs: 800 and 835, respectively;

[0964] gg) are SEQ ID NOs: 801 and 836, respectively;

[0965] hh) are SEQ ID NOs: 802 and 837, respectively; and

[0966] ii) are SEQ ID NOs: 803 and 838, respectively, wherein the oligonucleotide comprises at least one GalNAc moiety conjugated to the nucleotide.

[0967] In some embodiments, the sense strand and the antisense strand of the oligonucleotide comprise a nucleotide sequence selected from the group consisting of:

[0968] a) SEQ ID NO: 771 and 806, respectively;

[0969] b) SEQ ID NOs: 776 and 811, respectively;

[0970] c) SEQ ID NOs: 780 and 815, respectively;

[0971] d) SEQ ID NOs: 781 and 816, respectively;

[0972] e) SEQ ID NOs: 782 and 817, respectively;

[0973] f) SEQ ID NOs: 790 and 825, respectively;

[0974] g) SEQ ID NOs: 795 and 830, respectively;

[0975] h) SEQ ID NOs: 798 and 833, respectively;

[0976] i) SEQ ID NOs: 799 and 834, respectively; and

[0977] j) are SEQ ID NOs: 803 and 838, respectively, wherein the oligonucleotide comprises at least one GalNAc moiety conjugated to the nucleotide.

[0978] In some embodiments, the sense strand and the antisense strand of the oligonucleotide comprise a nucleotide sequence selected from the group consisting of:

[0979] a) SEQ ID NO: 771 and 806, respectively;

[0980] b) SEQ ID NOs: 780 and 815, respectively;

[0981] c) SEQ ID NOs: 781 and 816, respectively;

[0982] d) SEQ ID NOs: 798 and 833, respectively;

[0983] e) SEQ ID NOs: 799 and 834, respectively; and

[0984] f) are SEQ ID NOs: 803 and 838, respectively, wherein the oligonucleotide comprises at least one GalNAc moiety conjugated to the nucleotide.

[0985] Lipid conjugation

[0986] In some embodiments, one or more lipid moieties are conjugated to the 5' terminal nucleotide of the sense strand. In some embodiments, one or more lipid moieties are conjugated to an adenine nucleotide. In some embodiments, one or more lipid moieties are conjugated to a guanine nucleotide. In some embodiments, one or more lipid moieties are conjugated to a cytosine nucleotide. In some embodiments, one or more lipid moieties are conjugated to a thymine nucleotide. In some embodiments, one or more lipid moieties are conjugated to a uracil nucleotide.

[0987] In some embodiments, the lipid moiety is a hydrocarbon chain. In some embodiments, the hydrocarbon chain is saturated. In some embodiments, the hydrocarbon chain is unsaturated. In some embodiments, the hydrocarbon chain is branched. In some embodiments, the hydrocarbon chain is straight. In some embodiments, the lipid moiety is C8-C 30 In some embodiments, the lipid moiety is C8:0, C 10 :0, C 11 :0, C 12 :0, C 14 :0, C 16 :0, C 17 :0, C 18 :0, C 18 :1, C 18 :2, C 22 :5, C 22 :0, C 24 :0, C 26 :0, C 22 :6, C 24 :1. Diacyl C 16 :0 or diacyl C 18 : 1. In some embodiments, the lipid moiety is C 16 In some embodiments, C 16 The hydrocarbon chain is represented by:

[0988]

[0989] In some embodiments, the sense strand is 20-22 nucleotides in length and the lipid moiety is a hydrocarbon chain conjugated to the 5' terminal nucleotide of the sense strand. In some embodiments, the sense strand is 20-22 nucleotides in length and the hydrocarbon chain is conjugated to the 5' terminal nucleotide of the sense strand. 14 -C 22 The hydrocarbon chain is conjugated to the 5' terminal nucleotide of the sense strand. In some embodiments, the sense strand is 20-22 nucleotides in length and C 16 The hydrocarbon chain is conjugated to the 5' terminal nucleotide of the sense strand. In some embodiments, the sense strand is 20 nucleotides in length and the lipid moiety is conjugated to the 5' terminal nucleotide of the sense strand. In some embodiments, the sense strand is 20 nucleotides in length and the lipid moiety is a hydrocarbon chain conjugated to the 5' terminal nucleotide of the sense strand. In some embodiments, the sense strand is 20 nucleotides in length and the lipid moiety is a hydrocarbon chain conjugated to the 5' terminal nucleotide of the sense strand. 14 -C 22 The hydrocarbon chain is conjugated to the 5' terminal nucleotide of the sense strand. In some embodiments, the sense strand is 20 nucleotides in length and C 16 The hydrocarbon chain is conjugated to the 5' terminal nucleotide of the sense strand.

[0990] In some embodiments, the oligonucleotide comprises (i) a sense strand of 20-22 nucleotides in length; (ii) an antisense strand comprising a 3' overhang sequence of one or more nucleotides in length; (iii) a blunt end comprising the 3' end of the sense strand; and (iv) a lipid portion conjugated to the 5' terminal nucleotide of the sense strand. In some embodiments, the oligonucleotide comprises (i) a sense strand of 20-22 nucleotides in length; (ii) an antisense strand comprising a 3' overhang sequence of one or more nucleotides in length; (iii) a blunt end comprising the 3' end of the sense strand; and (iv) a hydrocarbon chain conjugated to the 5' terminal nucleotide of the sense strand. In some embodiments, the oligonucleotide comprises (i) a sense strand of 20-22 nucleotides in length; (ii) an antisense strand comprising a 3' overhang sequence of one or more nucleotides in length; (iii) a blunt end comprising the 3' end of the sense strand; and (iv) a C chain conjugated to the 5' terminal nucleotide of the sense strand. 14 -C 22 In some embodiments, the oligonucleotide comprises (i) a sense strand of 20-22 nucleotides in length; (ii) an antisense strand comprising a 3' overhang sequence of one or more nucleotides in length; (iii) a blunt end comprising the 3' end of the sense strand; and (iv) a C nucleotide conjugated to the 5' terminal nucleotide of the sense strand. 16 Hydrocarbon chain.

[0991] In some embodiments, the oligonucleotide comprises (i) a sense strand of 20 nucleotides in length; (ii) an antisense strand of 22 nucleotides in length and comprising a 3' overhang sequence of two nucleotides in length; (iii) a blunt end comprising the 3' end of the sense strand; and (iv) a lipid portion conjugated to the 5' terminal nucleotide of the sense strand. In some embodiments, the oligonucleotide comprises (i) a sense strand of 20 nucleotides in length; (ii) an antisense strand of 22 nucleotides in length and comprising a 3' overhang sequence of two nucleotides in length; (iii) a blunt end comprising the 3' end of the sense strand; and (iv) a hydrocarbon chain conjugated to the 5' terminal nucleotide of the sense strand. In some embodiments, the oligonucleotide comprises (i) a sense strand of 20 nucleotides in length; (ii) an antisense strand of 22 nucleotides in length and comprising a 3' overhang sequence of two nucleotides in length; (iii) a blunt end comprising the 3' end of the sense strand; and (iv) a C chain conjugated to the 5' terminal nucleotide of the sense strand. 14 -C 22 In some embodiments, the oligonucleotide comprises (i) a sense strand of 20 nucleotides in length; (ii) an antisense strand of 22 nucleotides in length and comprising a 3' overhang sequence of two nucleotides in length; (iii) a blunt end comprising the 3' end of the sense strand; and (iv) a C nucleotide conjugated to the 5' terminal nucleotide of the sense strand. 16 Hydrocarbon chain.

[0992] In some embodiments, the oligonucleotide comprises (i) an antisense strand of 19-30 nucleotides, the antisense strand comprising a complementary region to a MAPT mRNA target sequence selected from the group consisting of SEQ ID NO: 1125, 1127, 1130, 1019, 1031, 1044, 1064, 1065, 1067, 1083, 915, 1095, 1096, 1102, 1110, 923, 925, 1025, 1039, 1049, 1061, 1070, 1072, 1075, 1081, 1108, 1111, 1114, 1119, 1120, 1121, 1122, 1123 and 1124; (ii) a sense strand of 19-25 nucleotides that forms a duplex region with the antisense strand; and (iii) a lipid moiety conjugated to the 5' terminal nucleotide of the sense strand. In some embodiments, the oligonucleotide comprises (i) an antisense strand of 19-30 nucleotides, the antisense strand comprising a complementary region to a MAPT mRNA target sequence selected from SEQ ID NO: 1061, 1108, 1119, 1120, 1124, 1130, 1065, 1095, 1096 and 1102; (ii) a sense strand of 19-25 nucleotides, the sense strand forming a duplex region with the antisense strand; and (iii) a lipid portion conjugated to the 5' terminal nucleotide of the sense strand. In some embodiments, the oligonucleotide comprises (i) an antisense strand of 19-30 nucleotides, the antisense strand comprising a complementary region to a MAPT mRNA target sequence selected from SEQ ID NOs: 1130, 1095, 1096, 1119, 1120, and 1124; (ii) a sense strand of 19-25 nucleotides, the sense strand forming a duplex region with the antisense strand; and (iii) a lipid portion conjugated to the 5' terminal nucleotide of the sense strand.

[0993] In some embodiments, the oligonucleotide comprises (i) an antisense strand of 19-30 nucleotides, the antisense strand comprising a complementary region to a MAPT mRNA target sequence selected from the group consisting of SEQ ID NO: 1125, 1127, 1130, 1019, 1031, 1044, 1064, 1065, 1067, 1083, 915, 1095, 1096, 1102, 1110, 923, 925, 1025, 1039, 1049, 1061, 1070, 1072, 1075, 1081, 1108, 1111, 1114, 1119, 1120, 1121, 1122, 1123 and 1124; (ii) a sense strand of 19-25 nucleotides that forms a duplex region with the antisense strand; and (iii) a hydrocarbon chain conjugated to the 5' terminal nucleotide of the sense strand. In some embodiments, the oligonucleotide comprises (i) an antisense strand of 19-30 nucleotides, the antisense strand comprising a complementary region to a MAPT mRNA target sequence selected from SEQ ID NO: 1061, 1108, 1119, 1120, 1124, 1130, 1065, 1095, 1096 and 1102; (ii) a sense strand of 19-25 nucleotides, the sense strand forming a duplex region with the antisense strand; and (iii) a hydrocarbon chain conjugated to the 5' terminal nucleotide of the sense strand. In some embodiments, the oligonucleotide comprises (i) an antisense strand of 19-30 nucleotides, the antisense strand comprising a complementary region to a MAPT mRNA target sequence selected from SEQ ID NOs: 1130, 1095, 1096, 1119, 1120, and 1124; (ii) a sense strand of 19-25 nucleotides, the sense strand forming a duplex region with the antisense strand; and (iii) a hydrocarbon chain conjugated to the 5' terminal nucleotide of the sense strand.

[0994] In some embodiments, the oligonucleotide comprises (i) an antisense strand of 19-30 nucleotides, the antisense strand comprising a complementary region to a MAPT mRNA target sequence selected from the group consisting of SEQ ID NO: 1125, 1127, 1130, 1019, 1031, 1044, 1064, 1065, 1067, 1083, 915, 1095, 1096, 1102, 1110, 923, 925, 1025, 1039, 1049, 1061, 1070, 1072, 1075, 1081, 1108, 1111, 1114, 1119, 1120, 1121, 1122, 1123 and 1124; (ii) a sense strand of 19-25 nucleotides that forms a duplex region with the antisense strand; and (iii) a C conjugated to the 5' terminal nucleotide of the sense strand. 14 -C 22In some embodiments, the oligonucleotide comprises (i) an antisense strand of 19-30 nucleotides, the antisense strand comprising a complementary region to a MAPT mRNA target sequence selected from SEQ ID NO: 1061, 1108, 1119, 1120, 1124, 1130, 1065, 1095, 1096, and 1102; (ii) a sense strand of 19-25 nucleotides, the sense strand forming a duplex region with the antisense strand; and (iii) a C nucleotide conjugated to the 5' terminal nucleotide of the sense strand. 14 -C 22 In some embodiments, the oligonucleotide comprises (i) an antisense strand of 19-30 nucleotides, the antisense strand comprising a complementary region to a MAPT mRNA target sequence selected from SEQ ID NOs: 1130, 1095, 1096, 1119, 1120, and 1124; (ii) a sense strand of 19-25 nucleotides, the sense strand forming a duplex region with the antisense strand; and (iii) a C nucleotide conjugated to the 5' terminal nucleotide of the sense strand. 14 -C 22 Hydrocarbon chain.

[0995] In some embodiments, the oligonucleotide comprises (i) an antisense strand of 19-30 nucleotides, the antisense strand comprising a complementary region to a MAPT mRNA target sequence selected from the group consisting of SEQ ID NO: 1125, 1127, 1130, 1019, 1031, 1044, 1064, 1065, 1067, 1083, 915, 1095, 1096, 1102, 1110, 923, 925, 1025, 1039, 1049, 1061, 1070, 1072, 1075, 1081, 1108, 1111, 1114, 1119, 1120, 1121, 1122, 1123 and 1124; (ii) a sense strand of 19-25 nucleotides that forms a duplex region with the antisense strand; and (iii) a C conjugated to the 5' terminal nucleotide of the sense strand. 16 In some embodiments, the oligonucleotide comprises (i) an antisense strand of 19-30 nucleotides, the antisense strand comprising a complementary region to a MAPT mRNA target sequence selected from SEQ ID NO: 1061, 1108, 1119, 1120, 1124, 1130, 1065, 1095, 1096, and 1102; (ii) a sense strand of 19-25 nucleotides, the sense strand forming a duplex region with the antisense strand; and (iii) a C nucleotide conjugated to the 5' terminal nucleotide of the sense strand. 16In some embodiments, the oligonucleotide comprises (i) an antisense strand of 19-30 nucleotides, the antisense strand comprising a complementary region to a MAPT mRNA target sequence selected from SEQ ID NOs: 1130, 1095, 1096, 1119, 1120, and 1124; (ii) a sense strand of 19-25 nucleotides, the sense strand forming a duplex region with the antisense strand; and (iii) a C nucleotide conjugated to the 5' terminal nucleotide of the sense strand. 16 Hydrocarbon chain.

[0996] In some embodiments, the oligonucleotide comprises a sense strand comprising a nucleotide sequence of SEQ ID NO: 1681 and an antisense strand comprising a nucleotide sequence of SEQ ID NO: 815, wherein the sense strand comprises a lipid moiety conjugated to the 5' terminal nucleotide of the sense strand. In some embodiments, the oligonucleotide comprises a sense strand comprising a nucleotide sequence of SEQ ID NO: 1681 and an antisense strand comprising a nucleotide sequence of SEQ ID NO: 815, wherein the sense strand comprises a hydrocarbon chain conjugated to the 5' terminal nucleotide of the sense strand. In some embodiments, the oligonucleotide comprises a sense strand comprising a nucleotide sequence of SEQ ID NO: 1681 and an antisense strand comprising a nucleotide sequence of SEQ ID NO: 815, wherein the sense strand comprises a C chain conjugated to the 5' terminal nucleotide of the sense strand. 14 -C 22 In some embodiments, the oligonucleotide comprises a sense strand comprising a nucleotide sequence of SEQ ID NO: 1681 and an antisense strand comprising a nucleotide sequence of SEQ ID NO: 815, wherein the sense strand comprises a C nucleotide conjugated to the 5' terminal nucleotide of the sense strand. 16 Hydrocarbon chain.

[0997] Exemplary MAPT-targeting RNAi oligonucleotides

[0998] In some embodiments, the MAPT targeting RNAi oligonucleotide for reducing MAPT gene expression provided by the present disclosure comprises a sense strand and an antisense strand, wherein all nucleotides constituting the sense strand and the antisense strand are modified, wherein the antisense strand comprises the complementary region of the MAPT mRNA target sequence of any one of SEQ ID NO:912-1295, and wherein the length of the complementary region is at least 15 continuous nucleotides. In some embodiments, the 5' terminal nucleotide of the antisense strand comprises 4'-O-monomethylphosphonate-2'-O-methyluridine [Me phosphonate-4O-mU], as described herein. In some embodiments, the 5' terminal nucleotide of the antisense strand comprises a thiophosphate bond. In some embodiments, the antisense strand and the sense strand comprise nucleotides and at least one thiophosphate bond modified by one or more 2'-F and 2'-OMe. In some embodiments, the antisense strand comprises 4 thiophosphate bonds and the sense strand comprises 1 thiophosphate bond. In some embodiments, the antisense strand comprises 5 phosphorothioate linkages and the sense strand comprises 1 phosphorothioate linkage.

[0999] In some embodiments, an oligonucleotide (eg, an RNAi oligonucleotide) comprises a sense strand having a sequence of any one of SEQ ID NOs: 912-1295 and an antisense strand comprising a complementary sequence selected from SEQ ID NOs: 1296-1679.

[1000] In some embodiments, the oligonucleotide comprises a sense strand having a sequence of any one of SEQ ID NOs: 1-384 and an antisense strand comprising a complementary sequence selected from SEQ ID NOs: 385-768.

[1001] In some embodiments, the oligonucleotide comprises a sense strand having a sequence of any one of SEQ ID NOs: 769-803 and an antisense strand comprising a complementary sequence selected from SEQ ID NOs: 804-838.

[1002] In some embodiments, the oligonucleotide comprises a sense strand having a sequence of any one of SEQ ID NOs: 769-803 and 1681 and an antisense strand comprising a complementary sequence selected from SEQ ID NOs: 804-838.

[1003] In some embodiments, the oligonucleotide comprises a sense strand having a sequence of any one of SEQ ID NOs: 839-873 and an antisense strand comprising a complementary sequence selected from SEQ ID NOs: 874-908.

[1004] In some embodiments, the oligonucleotide comprises a sense strand having a sequence of any one of SEQ ID NOs: 839-873 and 1681 and an antisense strand comprising a complementary sequence selected from SEQ ID NOs: 874-908.

[1005] In some embodiments, the oligonucleotides (eg, RNAi oligonucleotides) used to reduce MAPT gene expression comprise:

[1006] a sense strand of 36 nucleotides, the sense strand comprising 2'-F modified nucleotides at positions 3, 5, 8, 10, 12, 13, 15, and 17; 2'-OMe modified nucleotides at positions 1, 2, 4, 6, 7, 9, 11, 14, 16, 18-27, and 31-36; GalNAc conjugated nucleotides at positions 28, 29, and 30; and a phosphorothioate bond between positions 1 and 2; and

[1007] The invention relates to an antisense strand of 22 nucleotides, the antisense strand comprising nucleotides modified with 2'-F at positions 2, 3, 4, 5, 7, 10, 14, 16 and 19; nucleotides modified with 2'-OMe at positions 1, 6, 8, 9, 11, 12, 13, 15, 17, 18 and 20-22; phosphorothioate bonds between positions 1 and 2, positions 2 and 3, positions 20 and 21, and positions 21 and 22; and a 5' terminal nucleotide at position 1 comprising a 4'-phosphate analog, optionally wherein the 5' terminal nucleotide comprises a 4'-phosphate analog, wherein the 5' terminal nucleotide comprises 4'-O-monomethylphosphonate-2'-O-methyluridine [Mephosphonate-4O-mU]; wherein positions 1-20 of the antisense strand form a duplex region with positions 1-20 of the sense strand, wherein positions 21-36 of the sense strand form a stem-loop, wherein positions 27-30 form a loop of the stem-loop, optionally wherein positions 27-30 comprise a tetraloop, wherein positions 21 and 22 of the antisense strand comprise an overhang, and wherein the sense strand and the antisense strand comprise a nucleotide sequence selected from the group consisting of:

[1008] a) SEQ ID NO: 769 and 804, respectively;

[1009] b) SEQ ID NOs: 770 and 805, respectively;

[1010] c) SEQ ID NOs: 771 and 806, respectively;

[1011] d) SEQ ID NOs: 772 and 807, respectively;

[1012] e) SEQ ID NOs: 773 and 808, respectively;

[1013] f) SEQ ID NOs: 774 and 809, respectively;

[1014] g) SEQ ID NOs: 775 and 810, respectively;

[1015] h) SEQ ID NOs: 776 and 811, respectively;

[1016] i) SEQ ID NOs: 777 and 812, respectively;

[1017] j) SEQ ID NOs: 778 and 813, respectively;

[1018] k) SEQ ID NOs: 779 and 814, respectively;

[1019] 1) are SEQ ID NOs: 780 and 815, respectively;

[1020] m) are SEQ ID NOs: 781 and 816, respectively;

[1021] n) are SEQ ID NOs: 782 and 817, respectively;

[1022] o) are SEQ ID NOs: 783 and 818, respectively;

[1023] p) are SEQ ID NOs: 784 and 819, respectively;

[1024] q) are SEQ ID NOs: 785 and 820, respectively;

[1025] r) are SEQ ID NOs: 786 and 821, respectively;

[1026] s) are SEQ ID NOs: 787 and 822, respectively;

[1027] t) are SEQ ID NOs: 788 and 823, respectively;

[1028] u) are SEQ ID NOs: 789 and 824, respectively;

[1029] v) SEQ ID NOs: 790 and 825, respectively;

[1030] w) are SEQ ID NOs: 791 and 826, respectively;

[1031] x) are SEQ ID NOs: 792 and 827, respectively;

[1032] y) are SEQ ID NOs: 793 and 828, respectively;

[1033] z) are SEQ ID NOs: 794 and 829, respectively;

[1034] aa) are SEQ ID NOs: 795 and 830, respectively;

[1035] bb) are SEQ ID NOs: 796 and 831, respectively;

[1036] cc) are SEQ ID NOs: 797 and 832, respectively;

[1037] dd) are SEQ ID NOs: 798 and 833, respectively;

[1038] ee) are SEQ ID NOs: 799 and 834, respectively;

[1039] ff) are SEQ ID NOs: 800 and 835, respectively;

[1040] gg) are SEQ ID NOs: 801 and 836, respectively;

[1041] hh) are SEQ ID NOs: 802 and 837, respectively; and

[1042] ii) are SEQ ID NOs: 803 and 838, respectively.

[1043] In some embodiments, the oligonucleotides (eg, RNAi oligonucleotides) used to reduce MAPT gene expression comprise:

[1044] a sense strand of 36 nucleotides, the sense strand comprising 2'-F modified nucleotides at positions 3, 5, 8, 10, 12, 13, 15, and 17; 2'-OMe modified nucleotides at positions 1, 2, 4, 6, 7, 9, 11, 14, 16, 18-27, and 31-36; GalNAc conjugated nucleotides at positions 28, 29, and 30; and a phosphorothioate bond between positions 1 and 2; and

[1045] The invention relates to an antisense strand of 22 nucleotides, the antisense strand comprising nucleotides modified with 2'-F at positions 2, 3, 4, 5, 7, 10, 14, 16 and 19; nucleotides modified with 2'-OMe at positions 1, 6, 8, 9, 11, 12, 13, 15, 17, 18 and 20-22; phosphorothioate bonds between positions 1 and 2, positions 2 and 3, positions 20 and 21, and positions 21 and 22; and a 5' terminal nucleotide at position 1 comprising a 4'-phosphate analog, optionally wherein the 5' terminal nucleotide comprises a 4'-phosphate analog, wherein the 5' terminal nucleotide comprises 4'-O-monomethylphosphonate-2'-O-methyluridine [Mephosphonate-4O-mU]; wherein positions 1-20 of the antisense strand form a duplex region with positions 1-20 of the sense strand, wherein positions 21-36 of the sense strand form a stem-loop, wherein positions 27-30 form a loop of the stem-loop, optionally wherein positions 27-30 comprise a tetraloop, wherein positions 21 and 22 of the antisense strand comprise an overhang, and wherein the sense strand and the antisense strand comprise a nucleotide sequence selected from the group consisting of:

[1046] a) SEQ ID NO: 771 and 806, respectively;

[1047] b) SEQ ID NOs: 776 and 811, respectively;

[1048] c) SEQ ID NOs: 780 and 815, respectively;

[1049] d) SEQ ID NOs: 781 and 816, respectively;

[1050] e) SEQ ID NOs: 782 and 817, respectively;

[1051] f) SEQ ID NOs: 790 and 825, respectively;

[1052] g) SEQ ID NOs: 795 and 830, respectively;

[1053] h) SEQ ID NOs: 798 and 833, respectively;

[1054] i) SEQ ID NOs: 799 and 834, respectively; and

[1055] j) are SEQ ID NOs: 803 and 838, respectively.

[1056] In some embodiments, the oligonucleotides (eg, RNAi oligonucleotides) used to reduce MAPT gene expression comprise:

[1057] a sense strand of 36 nucleotides, the sense strand comprising 2'-F modified nucleotides at positions 3, 5, 8, 10, 12, 13, 15, and 17; 2'-OMe modified nucleotides at positions 1, 2, 4, 6, 7, 9, 11, 14, 16, 18-27, and 31-36; GalNAc conjugated nucleotides at positions 28, 29, and 30; and a phosphorothioate bond between positions 1 and 2; and

[1058] The invention relates to an antisense strand of 22 nucleotides, the antisense strand comprising nucleotides modified with 2'-F at positions 2, 3, 4, 5, 7, 10, 14, 16 and 19; nucleotides modified with 2'-OMe at positions 1, 6, 8, 9, 11, 12, 13, 15, 17, 18 and 20-22; phosphorothioate bonds between positions 1 and 2, positions 2 and 3, positions 20 and 21, and positions 21 and 22; and a 5' terminal nucleotide at position 1 comprising a 4'-phosphate analog, optionally wherein the 5' terminal nucleotide comprises a 4'-phosphate analog, wherein the 5' terminal nucleotide comprises 4'-O-monomethylphosphonate-2'-O-methyluridine [Mephosphonate-4O-mU]; wherein positions 1-20 of the antisense strand form a duplex region with positions 1-20 of the sense strand, wherein positions 21-36 of the sense strand form a stem-loop, wherein positions 27-30 form a loop of the stem-loop, optionally wherein positions 27-30 comprise a tetraloop, wherein positions 21 and 22 of the antisense strand comprise an overhang, and wherein the sense strand and the antisense strand comprise a nucleotide sequence selected from the group consisting of:

[1059] a) SEQ ID NO: 771 and 806, respectively;

[1060] b) SEQ ID NOs: 780 and 815, respectively;

[1061] c) SEQ ID NOs: 781 and 816, respectively;

[1062] d) SEQ ID NOs: 798 and 833, respectively;

[1063] e) SEQ ID NOs: 799 and 834, respectively; and

[1064] f) are SEQ ID NOs: 803 and 838, respectively.

[1065] In some embodiments, the oligonucleotides (eg, RNAi oligonucleotides) used to reduce MAPT gene expression comprise:

[1066] A 20-nucleotide sense strand comprising 2'-F modified nucleotides at positions 3, 5, 8, 10, 12, 13, 15, and 17; 2'-OMe modified nucleotides at positions 2, 4, 6, 7, 9, 11, 14, 16, and 18-20; a C conjugated to the nucleotide at position 1 16 a hydrocarbon chain; and a phosphorothioate bond between positions 1 and 2, between positions 18 and 19, and between positions 19 and 20; and

[1067] 2'-OMe modified nucleotides at positions 1, 6, 8, 9, 11, 12, 13, 15, 17, 18 and 20-22; phosphorothioate bonds between positions 1 and 2, positions 2 and 3, positions 20 and 21, and positions 21 and 22; and a 5' terminal nucleotide at position 1 comprising a 4'-phosphate analog, optionally wherein the 5' terminal nucleotide comprises 4'-O-monomethylphosphonate-2'-O-methyluridine [Mephosphonate-4O-mU]; wherein positions 1-20 of the antisense strand form a duplex region with positions 1-20 of the sense strand, wherein positions 21 and 22 of the antisense strand comprise overhangs, and wherein the sense strand and the antisense strand comprise the nucleotide sequences of SEQ ID NOs: 1681 and 815, respectively.

[1068] In some embodiments, the oligonucleotides for reducing the expression of the MAPT gene comprise a sense strand containing a nucleotide sequence as shown in SEQ ID NO: 771 and an antisense strand containing a nucleotide sequence as shown in SEQ ID NO: 806. In some embodiments, the oligonucleotides for reducing the expression of the MAPT gene comprise a sense strand containing a nucleotide sequence as shown in SEQ ID NO: 780 and an antisense strand containing a nucleotide sequence as shown in SEQ ID NO: 815. In some embodiments, the oligonucleotides for reducing the expression of the MAPT gene comprise a sense strand containing a nucleotide sequence as shown in SEQ ID NO: 781 and an antisense strand containing a nucleotide sequence as shown in SEQ ID NO: 816. In some embodiments, the oligonucleotides for reducing the expression of the MAPT gene comprise a sense strand containing a nucleotide sequence as shown in SEQ ID NO: 798 and an antisense strand containing a nucleotide sequence as shown in SEQ ID NO: 833. In some embodiments, the oligonucleotides for reducing the expression of the MAPT gene comprise a sense strand containing a nucleotide sequence as shown in SEQ ID NO: 799 and an antisense strand containing a nucleotide sequence as shown in SEQ ID NO: 834. In some embodiments, the oligonucleotides for reducing the expression of the MAPT gene comprise a sense strand containing a nucleotide sequence as shown in SEQ ID NO: 803 and an antisense strand containing a nucleotide sequence as shown in SEQ ID NO: 838. In some embodiments, the oligonucleotides for reducing the expression of the MAPT gene comprise a sense strand containing a nucleotide sequence as shown in SEQ ID NO: 1681 and an antisense strand containing a nucleotide sequence as shown in SEQ ID NO: 815.

[1069] In some embodiments, the oligonucleotides (e.g., RNAi oligonucleotides) used to reduce MAPT gene expression comprise (i) an antisense strand of 19-30 nucleotides in length, wherein the antisense strand comprises a nucleotide sequence comprising a complementary region to a MAPT mRNA target sequence, wherein the complementary region is as shown in SEQ ID NO: 1514; and (ii) a sense strand of 19-50 nucleotides in length, wherein the sense strand comprises a complementary region to the antisense strand, wherein the antisense strand and the sense strand are independent strands forming an asymmetric duplex region having an overhang of 1-4 nucleotides at the 3' end of the antisense strand.

[1070] In some embodiments, the oligonucleotides (e.g., RNAi oligonucleotides) for reducing MAPT gene expression comprise (i) an antisense strand of 19-30 nucleotides in length, wherein the antisense strand comprises a nucleotide sequence comprising a complementary region to a MAPT mRNA target sequence, wherein the complementary region is as shown in SEQ ID NO: 1479; and (ii) a sense strand of 19-50 nucleotides in length, wherein the sense strand comprises a complementary region to the antisense strand, wherein the antisense strand and the sense strand are independent strands forming an asymmetric duplex region having an overhang of 1-4 nucleotides at the 3' end of the antisense strand.

[1071] In some embodiments, the oligonucleotides for reducing MAPT gene expression comprise (i) an antisense strand of 19-30 nucleotides in length, wherein the antisense strand comprises a nucleotide sequence comprising a complementary region to a MAPT mRNA target sequence, wherein the complementary region is as shown in SEQ ID NO: 1480; and (ii) a sense strand of 19-50 nucleotides in length, wherein the sense strand comprises a complementary region to the antisense strand, wherein the antisense strand and the sense strand are independent strands forming an asymmetric duplex region having an overhang of 1-4 nucleotides at the 3' end of the antisense strand.

[1072] In some embodiments, the MAPT targeting RNAi oligonucleotide for reducing MAPT gene expression comprises (i) an antisense strand of 19-30 nucleotides in length, wherein the antisense strand comprises a nucleotide sequence containing a complementary region to a MAPT mRNA target sequence, wherein the complementary region is as shown in SEQ ID NO: 1503; and (ii) a sense strand of 19-50 nucleotides in length, wherein the sense strand comprises a complementary region to the antisense strand, wherein the antisense strand and the sense strand are independent strands forming an asymmetric duplex region having an overhang of 1-4 nucleotides at the 3' end of the antisense strand.

[1073] In some embodiments, the oligonucleotides (e.g., RNAi oligonucleotides) used to reduce MAPT gene expression comprise (i) an antisense strand of 19-30 nucleotides in length, wherein the antisense strand comprises a nucleotide sequence comprising a complementary region to a MAPT mRNA target sequence, wherein the complementary region is as shown in SEQ ID NO: 1504; and (ii) a sense strand of 19-50 nucleotides in length, wherein the sense strand comprises a complementary region to the antisense strand, wherein the antisense strand and the sense strand are independent strands forming an asymmetric duplex region having an overhang of 1-4 nucleotides at the 3' end of the antisense strand.

[1074] In some embodiments, the oligonucleotides (e.g., RNAi oligonucleotides) used to reduce MAPT gene expression comprise (i) an antisense strand of 19-30 nucleotides in length, wherein the antisense strand comprises a nucleotide sequence comprising a complementary region to a MAPT mRNA target sequence, wherein the complementary region is as shown in SEQ ID NO: 1508; and (ii) a sense strand of 19-50 nucleotides in length, wherein the sense strand comprises a complementary region to the antisense strand, wherein the antisense strand and the sense strand are independent strands forming an asymmetric duplex region having an overhang of 1-4 nucleotides at the 3' end of the antisense strand.

[1075] In some embodiments, the oligonucleotides (e.g., RNAi oligonucleotides) for reducing MAPT gene expression comprise (i) an antisense strand of 19-30 nucleotides in length, wherein the antisense strand comprises a nucleotide sequence comprising a complementary region to a MAPT mRNA target sequence, wherein the complementary region is as shown in SEQ ID NO: 1514; and (ii) a sense strand of 19-50 nucleotides in length, wherein the sense strand comprises the complementary region to the antisense strand and a stem loop at the 3' end, wherein the stem loop is as shown in S1-L-S2, wherein S1 is complementary to S2 and wherein L forms a loop of 3 to 5 nucleotides in length between S1 and S2, wherein the antisense strand and the sense strand are independent strands forming an asymmetric duplex region having an overhang of 1-4 nucleotides at the 3' end of the antisense strand.

[1076] In some embodiments, the oligonucleotides (e.g., RNAi oligonucleotides) for reducing MAPT gene expression comprise (i) an antisense strand of 19-30 nucleotides in length, wherein the antisense strand comprises a nucleotide sequence comprising a complementary region to a MAPT mRNA target sequence, wherein the complementary region is as shown in SEQ ID NO: 1479; and (ii) a sense strand of 19-50 nucleotides in length, wherein the sense strand comprises the complementary region to the antisense strand and a stem loop at the 3' end, wherein the stem loop is as shown in S1-L-S2, wherein S1 is complementary to S2 and wherein L forms a loop of 3 to 5 nucleotides in length between S1 and S2, wherein the antisense strand and the sense strand are independent strands forming an asymmetric duplex region having an overhang of 1-4 nucleotides at the 3' end of the antisense strand.

[1077] In some embodiments, the oligonucleotides (e.g., RNAi oligonucleotides) for reducing MAPT gene expression comprise (i) an antisense strand of 19-30 nucleotides in length, wherein the antisense strand comprises a nucleotide sequence comprising a complementary region to a MAPT mRNA target sequence, wherein the complementary region is as shown in SEQ ID NO: 1480; and (ii) a sense strand of 19-50 nucleotides in length, wherein the sense strand comprises the complementary region to the antisense strand and a stem loop at the 3' end, wherein the stem loop is as shown in S1-L-S2, wherein S1 is complementary to S2 and wherein L forms a loop of 3 to 5 nucleotides in length between S1 and S2, wherein the antisense strand and the sense strand are independent strands forming an asymmetric duplex region having an overhang of 1-4 nucleotides at the 3' end of the antisense strand.

[1078] In some embodiments, the oligonucleotides (e.g., RNAi oligonucleotides) for reducing MAPT gene expression comprise (i) an antisense strand of 19-30 nucleotides in length, wherein the antisense strand comprises a nucleotide sequence comprising a complementary region to a MAPT mRNA target sequence, wherein the complementary region is as shown in SEQ ID NO: 1503; and (ii) a sense strand of 19-50 nucleotides in length, wherein the sense strand comprises the complementary region to the antisense strand and a stem loop at the 3' end, wherein the stem loop is as shown in S1-L-S2, wherein S1 is complementary to S2 and wherein L forms a loop of 3 to 5 nucleotides in length between S1 and S2, wherein the antisense strand and the sense strand are independent strands forming an asymmetric duplex region having an overhang of 1-4 nucleotides at the 3' end of the antisense strand.

[1079] In some embodiments, the oligonucleotides (e.g., RNAi oligonucleotides) for reducing MAPT gene expression comprise (i) an antisense strand of 19-30 nucleotides in length, wherein the antisense strand comprises a nucleotide sequence comprising a complementary region to a MAPT mRNA target sequence, wherein the complementary region is as shown in SEQ ID NO: 1504; and (ii) a sense strand of 19-50 nucleotides in length, wherein the sense strand comprises the complementary region to the antisense strand and a stem loop at the 3' end, wherein the stem loop is as shown in S1-L-S2, wherein S1 is complementary to S2 and wherein L forms a loop of 3 to 5 nucleotides in length between S1 and S2, wherein the antisense strand and the sense strand are independent strands forming an asymmetric duplex region having an overhang of 1-4 nucleotides at the 3' end of the antisense strand.

[1080] In some embodiments, the oligonucleotides (e.g., RNAi oligonucleotides) for reducing MAPT gene expression comprise (i) an antisense strand of 19-30 nucleotides in length, wherein the antisense strand comprises a nucleotide sequence comprising a complementary region to a MAPT mRNA target sequence, wherein the complementary region is as shown in SEQ ID NO: 1508; and (ii) a sense strand of 19-50 nucleotides in length, wherein the sense strand comprises the complementary region to the antisense strand and a stem loop at the 3' end, wherein the stem loop is as shown in S1-L-S2, wherein S1 is complementary to S2 and wherein L forms a loop of 3 to 5 nucleotides in length between S1 and S2, wherein the antisense strand and the sense strand are independent strands forming an asymmetric duplex region having an overhang of 1-4 nucleotides at the 3' end of the antisense strand.

[1081] In some embodiments, the oligonucleotides (e.g., RNAi oligonucleotides) for reducing MAPT gene expression comprise (i) an antisense strand of 19-30 nucleotides in length, wherein the antisense strand comprises a nucleotide sequence comprising a complementary region to a MAPT mRNA target sequence, wherein the complementary region is as shown in SEQ ID NO: 1479; and (ii) a sense strand of 19-25 nucleotides in length, wherein the sense strand comprises a complementary region to the antisense strand, wherein the oligonucleotide comprises a blunt end comprising the 3' end of the sense strand, wherein the antisense strand and the sense strand are independent strands forming an asymmetric duplex region having an overhang of 1-4 nucleotides at the 3' end of the antisense strand.

[1082] In some embodiments, the oligonucleotides (e.g., RNAi oligonucleotides) for reducing MAPT gene expression comprise (i) an antisense strand of 19-30 nucleotides in length, wherein the antisense strand comprises a nucleotide sequence comprising a complementary region to a MAPT mRNA target sequence, wherein the complementary region is as shown in SEQ ID NO: 1514; and (ii) a sense strand of 19-50 nucleotides in length, wherein the sense strand comprises a complementary region to the antisense strand, wherein the complementary region to the antisense strand is as shown in SEQ ID NO: 1130, wherein the antisense strand and the sense strand are independent strands forming an asymmetric duplex region having an overhang of 1-4 nucleotides at the 3' end of the antisense strand.

[1083] In some embodiments, the oligonucleotides (e.g., RNAi oligonucleotides) for reducing MAPT gene expression comprise (i) an antisense strand of 19-30 nucleotides in length, wherein the antisense strand comprises a nucleotide sequence comprising a complementary region to a MAPT mRNA target sequence, wherein the complementary region is as shown in SEQ ID NO: 1479; and (ii) a sense strand of 19-50 nucleotides in length, wherein the sense strand comprises a complementary region to the antisense strand, wherein the complementary region to the antisense strand is as shown in SEQ ID NO: 1095, wherein the antisense strand and the sense strand are independent strands forming an asymmetric duplex region having an overhang of 1-4 nucleotides at the 3' end of the antisense strand.

[1084] In some embodiments, the oligonucleotides (e.g., RNAi oligonucleotides) for reducing MAPT gene expression comprise (i) an antisense strand of 19-30 nucleotides in length, wherein the antisense strand comprises a nucleotide sequence comprising a complementary region to a MAPT mRNA target sequence, wherein the complementary region is as shown in SEQ ID NO: 1480; and (ii) a sense strand of 19-50 nucleotides in length, wherein the sense strand comprises a complementary region to the antisense strand, wherein the complementary region to the antisense strand is as shown in SEQ ID NO: 1096, wherein the antisense strand and the sense strand are independent strands forming an asymmetric duplex region having an overhang of 1-4 nucleotides at the 3' end of the antisense strand.

[1085] In some embodiments, the oligonucleotides (e.g., RNAi oligonucleotides) for reducing MAPT gene expression comprise (i) an antisense strand of 19-30 nucleotides in length, wherein the antisense strand comprises a nucleotide sequence comprising a complementary region to a MAPT mRNA target sequence, wherein the complementary region is as shown in SEQ ID NO: 1503; and (ii) a sense strand of 19-50 nucleotides in length, wherein the sense strand comprises a complementary region to the antisense strand, wherein the complementary region to the antisense strand is as shown in SEQ ID NO: 1119, wherein the antisense strand and the sense strand are independent strands forming an asymmetric duplex region having an overhang of 1-4 nucleotides at the 3' end of the antisense strand.

[1086] In some embodiments, the oligonucleotides (e.g., RNAi oligonucleotides) for reducing MAPT gene expression comprise (i) an antisense strand of 19-30 nucleotides in length, wherein the antisense strand comprises a nucleotide sequence comprising a complementary region to a MAPT mRNA target sequence, wherein the complementary region is as shown in SEQ ID NO: 1504; and (ii) a sense strand of 19-50 nucleotides in length, wherein the sense strand comprises a complementary region to the antisense strand, wherein the complementary region to the antisense strand is as shown in SEQ ID NO: 1120, wherein the antisense strand and the sense strand are independent strands forming an asymmetric duplex region having an overhang of 1-4 nucleotides at the 3' end of the antisense strand.

[1087] In some embodiments, the oligonucleotides (e.g., RNAi oligonucleotides) for reducing MAPT gene expression comprise (i) an antisense strand of 19-30 nucleotides in length, wherein the antisense strand comprises a nucleotide sequence comprising a complementary region to a MAPT mRNA target sequence, wherein the complementary region is as shown in SEQ ID NO: 1508; and (ii) a sense strand of 19-50 nucleotides in length, wherein the sense strand comprises a complementary region to the antisense strand, wherein the complementary region to the antisense strand is as shown in SEQ ID NO: 1124, wherein the antisense strand and the sense strand are independent strands forming an asymmetric duplex region having an overhang of 1-4 nucleotides at the 3' end of the antisense strand.

[1088] In some embodiments, the oligonucleotides (e.g., RNAi oligonucleotides) for reducing MAPT gene expression comprise (i) an antisense strand of 19-30 nucleotides in length, wherein the antisense strand comprises a nucleotide sequence comprising a complementary region to a MAPT mRNA target sequence, wherein the complementary region is as shown in SEQ ID NO: 1514; and (ii) a sense strand of 19-50 nucleotides in length, wherein the sense strand comprises the complementary region to the antisense strand and a stem loop at the 3' end, wherein the complementary region to the antisense strand is as shown in SEQ ID NO: 1130, wherein the stem loop is as shown in S1-L-S2, wherein S1 is complementary to S2 and wherein L forms a loop of 3 to 5 nucleotides in length between S1 and S2, wherein the antisense strand and the sense strand are independent strands forming an asymmetric duplex region having an overhang of 1-4 nucleotides at the 3' end of the antisense strand.

[1089] In some embodiments, the oligonucleotides (e.g., RNAi oligonucleotides) for reducing MAPT gene expression comprise (i) an antisense strand of 19-30 nucleotides in length, wherein the antisense strand comprises a nucleotide sequence comprising a complementary region to a MAPT mRNA target sequence, wherein the complementary region is as shown in SEQ ID NO: 1479; and (ii) a sense strand of 19-50 nucleotides in length, wherein the sense strand comprises the complementary region to the antisense strand and a stem loop at the 3' end, wherein the complementary region to the antisense strand is as shown in SEQ ID NO: 1095, wherein the stem loop is as shown in S1-L-S2, wherein S1 is complementary to S2 and wherein L forms a loop of 3 to 5 nucleotides in length between S1 and S2, wherein the antisense strand and the sense strand are independent strands forming an asymmetric duplex region having an overhang of 1-4 nucleotides at the 3' end of the antisense strand.

[1090] In some embodiments, the oligonucleotides (e.g., RNAi oligonucleotides) for reducing MAPT gene expression comprise (i) an antisense strand of 19-30 nucleotides in length, wherein the antisense strand comprises a nucleotide sequence comprising a complementary region to a MAPT mRNA target sequence, wherein the complementary region is as shown in SEQ ID NO: 1480; and (ii) a sense strand of 19-50 nucleotides in length, wherein the sense strand comprises the complementary region to the antisense strand and a stem loop at the 3' end, wherein the complementary region to the antisense strand is as shown in SEQ ID NO: 1096, wherein the stem loop is as shown in S1-L-S2, wherein S1 is complementary to S2 and wherein L forms a loop of 3 to 5 nucleotides in length between S1 and S2, wherein the antisense strand and the sense strand are independent strands forming an asymmetric duplex region having an overhang of 1-4 nucleotides at the 3' end of the antisense strand.

[1091] In some embodiments, the oligonucleotides (e.g., RNAi oligonucleotides) for reducing MAPT gene expression comprise (i) an antisense strand of 19-30 nucleotides in length, wherein the antisense strand comprises a nucleotide sequence comprising a complementary region to a MAPT mRNA target sequence, wherein the complementary region is as shown in SEQ ID NO: 1503; and (ii) a sense strand of 19-50 nucleotides in length, wherein the sense strand comprises the complementary region to the antisense strand and a stem loop at the 3' end, wherein the complementary region to the antisense strand is as shown in SEQ ID NO: 1119, wherein the stem loop is as shown in S1-L-S2, wherein S1 is complementary to S2 and wherein L forms a loop of 3 to 5 nucleotides in length between S1 and S2, wherein the antisense strand and the sense strand are independent strands forming an asymmetric duplex region having an overhang of 1-4 nucleotides at the 3' end of the antisense strand.

[1092] In some embodiments, the oligonucleotides (e.g., RNAi oligonucleotides) for reducing MAPT gene expression comprise (i) an antisense strand of 19-30 nucleotides in length, wherein the antisense strand comprises a nucleotide sequence comprising a complementary region to a MAPT mRNA target sequence, wherein the complementary region is as shown in SEQ ID NO: 1504; and (ii) a sense strand of 19-50 nucleotides in length, wherein the sense strand comprises the complementary region to the antisense strand and a stem loop at the 3' end, wherein the complementary region to the antisense strand is as shown in SEQ ID NO: 1120, wherein the stem loop is as shown in S1-L-S2, wherein S1 is complementary to S2 and wherein L forms a loop of 3 to 5 nucleotides in length between S1 and S2, wherein the antisense strand and the sense strand are independent strands forming an asymmetric duplex region having an overhang of 1-4 nucleotides at the 3' end of the antisense strand.

[1093] In some embodiments, the oligonucleotides (e.g., RNAi oligonucleotides) for reducing MAPT gene expression comprise (i) an antisense strand of 19-30 nucleotides in length, wherein the antisense strand comprises a nucleotide sequence comprising a complementary region to a MAPT mRNA target sequence, wherein the complementary region is as shown in SEQ ID NO: 1508; and (ii) a sense strand of 19-50 nucleotides in length, wherein the sense strand comprises the complementary region to the antisense strand and a stem loop at the 3' end, wherein the complementary region to the antisense strand is as shown in SEQ ID NO: 1124, wherein the stem loop is as shown in S1-L-S2, wherein S1 is complementary to S2 and wherein L forms a loop of 3 to 5 nucleotides in length between S1 and S2, wherein the antisense strand and the sense strand are independent strands forming an asymmetric duplex region having an overhang of 1-4 nucleotides at the 3' end of the antisense strand.

[1094] In some embodiments, oligonucleotides for reducing MAPT gene expression (e.g., RNAi oligonucleotides) comprise (i) an antisense strand of 19-30 nucleotides in length, wherein the antisense strand comprises a nucleotide sequence comprising a complementary region to a MAPT mRNA target sequence, wherein the complementary region is as shown in SEQ ID NO: 1479; and (ii) a sense strand of 19-25 nucleotides in length, wherein the sense strand comprises a complementary region to the antisense strand, wherein the complementary region to the antisense strand is as shown in SEQ ID NO: 1095, wherein the oligonucleotide comprises a blunt end comprising the 3' end of the sense strand, wherein the antisense strand and the sense strand are independent strands forming an asymmetric duplex region having an overhang of 1-4 nucleotides at the 3' end of the antisense strand.

[1095] In some embodiments, an oligonucleotide for reducing MAPT gene expression (eg, an RNAi oligonucleotide) comprises a sense strand and an antisense strand according to:

[1096] Sense strand: 5'-mX-S-mX-fX-mX-fX-mX-mX-fX-mX-fX-mX-fX-fX-mX-fX-mX-fX-mX-mX-mX-mX-mX-mX-mX-mX-mX-mX-[ademX-Gal NAc]-[ademX-GalNAc]-[ademX-GalNAc]-mX-mX-mX-mX-mX-mX-3', hybridized to:

[1097] Antisense strand: 5'-[Mephosphonate-4O-mX]-S-fX-S-fX-fX-fX-mX-fX-mX-fX-mX-mX-fX-mX-fX-mX-fX-mX-fX-mX-fX-mX-mX-S-mX-S-mX-3', where mX=2'-OMe modified nucleotide, fX=2'-F modified nucleotide, -S-=phosphorothioate bond, -=phosphodiester bond, [Mephosphonate-4O-mX]=4'-O-monomethylphosphonate-2'-O-methyl modified nucleotide, and ademX-GalNAc=GalNAc attached to the nucleotide.

[1098] In some embodiments, an oligonucleotide for reducing MAPT gene expression (eg, an RNAi oligonucleotide) comprises a sense strand and an antisense strand according to:

[1099] Sense strand: 5'-mX-S-mX-mX-mX-mX-mX-mX-fX-fX-fX-fX-mX-mX-mX-mX-mX-mX-mX-mX-mX-mX-mX-mX-mX -mX-mX-mX-[ademX-GalNAc]-[ademX-GalNAc]-[ademX-GalNAc]-mX-mX-mX-mX-mG-mX-3', hybridized to:

[1100] Antisense strand: 5'-[Mephosphonate-4O-mX]-S-fX-S-fX-S-fX-fX-mX-fX-mX-mX-fX-mX-mX-mX-fX-mX-mX-mX-mX-mX-mX-mX-S-mX-S-mX-3', where mX=2'-OMe modified nucleotide, fX=2'-F modified nucleotide, -S-=phosphorothioate bond, -=phosphodiester bond, [Mephosphonate-4O-mX]=4'-O-monomethylphosphonate-2'-O-methyl modified nucleotide, and ademX-GalNAc=GalNAc attached to the nucleotide.

[1101] In some embodiments, an oligonucleotide for reducing MAPT gene expression (eg, an RNAi oligonucleotide) comprises a sense strand and an antisense strand according to:

[1102] Sense strand: 5'-[AdemX-L]-S-mX-fX-mX-fX-mX-fX-mX-fX-mX-fX-fX-mX-fX-fX-mX-fX-mX-S-mX-S-mX-3', hybridizes to:

[1103] Antisense strand: 5'-[Mephosphonate-4O-mX]-S-fX-S-fX-fX-fX-mX-fX-mX-fX-mX-fX-mX-mX-fX-mX-fX-mX-fX-mX-fX-mX-mX-S-mX-S-mX-3', where mX=2'-OMe modified nucleotide, fX=2'-F modified nucleotide, -S-=phosphorothioate bond, -=phosphodiester bond, [Mephosphonate-4O-mX]=4'-O-monomethylphosphonate-2'-O-methyl modified nucleotide, and ademX-L=lipid moiety attached to the nucleotide.

[1104] In some embodiments, an oligonucleotide for reducing MAPT gene expression (eg, an RNAi oligonucleotide) comprises a sense strand and an antisense strand according to:

[1105] Sense strand: 5'-[AdemX-C16]-S-mX-fX-mX-fX-mX-fX-mX-fX-fX-mX-fX-fX-mX-fX-mX-fX-mX-S-mX-S-mX-3', hybridizes to:

[1106] Antisense strand: 5'-[Mephosphonate-4O-mX]-S-fX-S-fX-fX-fX-mX-fX-mX-fX-mX-fX-mX-mX-fX-mX-fX-mX-fX-mX-fX-mX-S-mX-S-mX-3', where mX = 2'-OMe modified nucleotide, fX = 2'-F modified nucleotide, -S- = phosphorothioate bond, - = phosphodiester bond, [Mephosphonate-4O-mX] = 4'-O-monomethylphosphonate-2'-O-methyl modified nucleotide, and ademX-C16 = C16 attached to the nucleotide. 16 Hydrocarbon chain.

[1107] In some embodiments, the present disclosure provides an oligonucleotide (e.g., an RNAi oligonucleotide) for reducing MAPT gene expression, wherein the oligonucleotide comprises a sense strand and an antisense strand, and the sense strand and the antisense strand comprise a nucleotide sequence selected from the group consisting of:

[1108] a) SEQ ID NO: 839 and 874, respectively;

[1109] b) SEQ ID NOs: 840 and 875, respectively;

[1110] c) SEQ ID NOs: 841 and 876, respectively;

[1111] d) SEQ ID NOs: 842 and 877, respectively;

[1112] e) SEQ ID NOs: 843 and 878, respectively;

[1113] f) SEQ ID NOs: 844 and 879, respectively;

[1114] g) SEQ ID NOs: 845 and 880, respectively;

[1115] h) SEQ ID NOs: 846 and 881, respectively;

[1116] i) SEQ ID NOs: 847 and 882, respectively;

[1117] j) SEQ ID NOs: 848 and 883, respectively;

[1118] k) SEQ ID NOs: 849 and 884, respectively;

[1119] 1) are SEQ ID NOs: 850 and 885, respectively;

[1120] m) are SEQ ID NOs: 851 and 886, respectively;

[1121] n) are SEQ ID NOs: 852 and 887, respectively;

[1122] o) are SEQ ID NOs: 853 and 888, respectively;

[1123] p) are SEQ ID NOs: 854 and 889, respectively;

[1124] q) are SEQ ID NOs: 855 and 890, respectively;

[1125] r) are SEQ ID NOs: 856 and 891, respectively;

[1126] s) are SEQ ID NOs: 857 and 892, respectively;

[1127] t) are SEQ ID NOs: 858 and 893, respectively;

[1128] u) are SEQ ID NOs: 859 and 894, respectively;

[1129] v) SEQ ID NOs: 860 and 895, respectively;

[1130] w) are SEQ ID NOs: 861 and 896, respectively;

[1131] x) are SEQ ID NOs: 862 and 897, respectively;

[1132] y) are SEQ ID NOs: 863 and 898, respectively;

[1133] z) are SEQ ID NOs: 864 and 899, respectively;

[1134] aa) are SEQ ID NOs: 865 and 900, respectively;

[1135] bb) are SEQ ID NOs: 866 and 901, respectively;

[1136] cc) are SEQ ID NOs: 867 and 902, respectively;

[1137] dd) are SEQ ID NOs: 868 and 903, respectively;

[1138] ee) are SEQ ID NOs: 869 and 904, respectively;

[1139] ff) are SEQ ID NOs: 870 and 905, respectively;

[1140] gg) are SEQ ID NOs: 871 and 906, respectively;

[1141] hh) are SEQ ID NOs: 872 and 907, respectively;

[1142] ii) SEQ ID NO: 873 and 908, respectively; and

[1143] jj) are SEQ ID NOs: 1682 and 885, respectively.

[1144] In some embodiments, the oligonucleotide for reducing MAPT gene expression (eg, RNAi oligonucleotide) comprises a sense strand and an antisense strand, wherein the sense strand and the antisense strand comprise a nucleotide sequence selected from the group consisting of:

[1145] a) SEQ ID NO: 860 and 895, respectively;

[1146] b) SEQ ID NO: 865 and 900, respectively;

[1147] c) SEQ ID NOs: 868 and 903, respectively;

[1148] d) SEQ ID NO: 869 and 904, respectively;

[1149] e) SEQ ID NOs: 873 and 908, respectively;

[1150] f) SEQ ID NOs: 841 and 876, respectively;

[1151] g) SEQ ID NOs: 846 and 881, respectively;

[1152] h) SEQ ID NOs: 850 and 885, respectively;

[1153] i) SEQ ID NOs: 851 and 886, respectively;

[1154] j) SEQ ID NOs: 852 and 887, respectively; and

[1155] k) are SEQ ID NOs: 1682 and 885, respectively.

[1156] In some embodiments, the oligonucleotide for reducing MAPT gene expression (eg, RNAi oligonucleotide) comprises a sense strand and an antisense strand, wherein the sense strand and the antisense strand comprise a nucleotide sequence selected from the group consisting of:

[1157] a) SEQ ID NO: 841 and 876, respectively;

[1158] b) SEQ ID NOs: 850 and 885, respectively;

[1159] c) SEQ ID NOs: 851 and 886, respectively;

[1160] d) SEQ ID NO: 868 and 903, respectively;

[1161] e) SEQ ID NOs: 869 and 904, respectively;

[1162] f) SEQ ID NOs: 873 and 908, respectively; and

[1163] g) are SEQ ID NOs: 1682 and 885, respectively.

[1164] In some embodiments, the oligonucleotides for reducing the expression of the MAPT gene comprise a sense strand containing a nucleotide sequence as shown in SEQ ID NO: 841 and an antisense strand containing a nucleotide sequence as shown in SEQ ID NO: 876. In some embodiments, the oligonucleotides for reducing the expression of the MAPT gene comprise a sense strand containing a nucleotide sequence as shown in SEQ ID NO: 850 and an antisense strand containing a nucleotide sequence as shown in SEQ ID NO: 885. In some embodiments, the oligonucleotides for reducing the expression of the MAPT gene comprise a sense strand containing a nucleotide sequence as shown in SEQ ID NO: 851 and an antisense strand containing a nucleotide sequence as shown in SEQ ID NO: 886. In some embodiments, the oligonucleotides for reducing the expression of the MAPT gene comprise a sense strand containing a nucleotide sequence as shown in SEQ ID NO: 868 and an antisense strand containing a nucleotide sequence as shown in SEQ ID NO: 903. In some embodiments, the oligonucleotides for reducing the expression of the MAPT gene comprise a sense strand containing a nucleotide sequence as shown in SEQ ID NO: 869 and an antisense strand containing a nucleotide sequence as shown in SEQ ID NO: 904. In some embodiments, the oligonucleotides for reducing the expression of the MAPT gene comprise a sense strand containing a nucleotide sequence as shown in SEQ ID NO: 873 and an antisense strand containing a nucleotide sequence as shown in SEQ ID NO: 908. In some embodiments, the oligonucleotides for reducing the expression of the MAPT gene comprise a sense strand containing a nucleotide sequence as shown in SEQ ID NO: 1682 and an antisense strand containing a nucleotide sequence as shown in SEQ ID NO: 885.

[1165] preparation

[1166] A variety of preparations have been developed to promote the use of oligonucleotides. For example, oligonucleotides (e.g., RNAi oligonucleotides) can be delivered to a subject or a cell environment using a preparation that minimizes degradation, promotes delivery and / or uptake, or provides another beneficial property for the oligonucleotides in the preparation. In some embodiments, compositions comprising oligonucleotides that reduce MAPT gene expression are provided herein. Such compositions can be suitably prepared so that when administered to a subject in the direct environment of a target cell or systemically, enough oligonucleotides of a portion enter cells to reduce MAPT gene expression. Suitable oligonucleotide preparations of any kind can be used to deliver oligonucleotides for reducing MAPT gene expression as disclosed herein. In some embodiments, oligonucleotides are prepared in buffer solutions, such as phosphate buffered saline solutions, liposomes, micellar structures, and capsids. In some embodiments, oligonucleotides are prepared in buffer solutions, such as phosphate buffered saline solutions.

[1167] The preparation of oligonucleotide and cationic lipid can be used to promote oligonucleotide transfection into cell.For example, cationic lipid, such as lipofectin, cationic glycerol derivative and polycationic molecule (for example, polylysine) can be used.Suitable lipid includes oligofectamine (Oligofectamine), lipofectamine (Lipofectamine) (Life Technologies), NC388 (Ribozyme Pharmaceuticals, Inc., Boulder, Colo.) or FuGene 6 (Roche), all of which can be used according to the manufacturer's instructions.In some embodiments, oligonucleotide is not prepared together with the component that promotes transfection into cell.

[1168] Thus, in some embodiments, the formulation comprises lipid nanoparticles. In some embodiments, the excipient comprises a liposome, a lipid, a lipid complex, a microsphere, a microparticle, a nanosphere or a nanoparticle, or can be otherwise formulated for use in administering to a cell, a tissue, an organ or a body of a subject in need thereof (see, e.g., Remington: THE SCIENCE AND PRACTICE OF PHARMACY, 22nd ed., Pharmaceutical Press, 2013).

[1169] In some embodiments, the formulations herein include excipients. In some embodiments, the excipients impart improved stability, improved absorption, improved solubility, and / or therapeutic enhancement of the active ingredient to the composition. In some embodiments, the excipient is a buffer (e.g., sodium citrate, sodium phosphate, tris base, or sodium hydroxide) or a vehicle (e.g., a buffered solution, paraffin, dimethyl sulfoxide, or mineral oil). In some embodiments, the oligonucleotides are lyophilized to extend their shelf life and then made into a solution before use (e.g., administration to a subject). Therefore, the excipient in the composition comprising any of the oligonucleotides described herein can be a lyoprotectant (e.g., mannitol, lactose, polyethylene glycol, or polyvinyl pyrrolidone) or a disintegration temperature regulator (e.g., dextran, Ficoll TM or gelatin).

[1170] In some embodiments, the pharmaceutical composition is formulated to be compatible with its intended route of administration. Examples of routes of administration include parenteral (e.g., intravenous, intramuscular, intraperitoneal, intradermal, subcutaneous), oral (e.g., inhalation), transdermal (e.g., topical), transmucosal, and rectal administration.

[1171] In some embodiments, the pharmaceutical composition is formulated for administration into the central nervous system. In some embodiments, the pharmaceutical composition is formulated for administration into the cerebrospinal fluid. In some embodiments, the pharmaceutical composition is formulated for administration to the spinal cord. In some embodiments, the pharmaceutical composition is formulated for intrathecal administration. In some embodiments, the pharmaceutical composition is formulated for administration to the brain. In some embodiments, the pharmaceutical composition is formulated for intraventricular administration. In some embodiments, the pharmaceutical composition is formulated for use in the brainstem. In some embodiments, the pharmaceutical composition is formulated for administration within the cisterna magna.

[1172] Pharmaceutical compositions suitable for injectable use include sterile aqueous solutions (where water soluble) or dispersions and sterile powders for the extemporaneous preparation of sterile injectable solutions or dispersions. For intravenous administration, suitable carriers include physiological saline, bacteriostatic water, Cremophor EL TM (BASF) or phosphate buffered saline (PBS). The carrier can be a solvent or dispersion medium containing, for example, water, ethanol, polyols (e.g., glycerol, propylene glycol, liquid polyethylene glycol, etc.) and suitable mixtures thereof. In many cases, it is preferred that isotonic agents, for example, sugars, polyols (such as mannitol, sorbitol) and / or sodium chloride are included in the composition. Sterile injectable solutions can be prepared by incorporating the desired amount of the oligonucleotide into a selected solvent together with one or a combination of ingredients listed above, followed by filtration sterilization as necessary.

[1173] In some embodiments, the composition can contain at least about 0.1% of the therapeutic agent (e.g., RNAi oligonucleotides for reducing MAPT gene expression) or more, but the percentage of active ingredient can be between about 1% to about 80% of the total composition weight or volume or more. Factors such as solubility, bioavailability, biological half-life, route of administration, product shelf life, and other pharmacological considerations will be expected by the technicians preparing such pharmaceutical preparations, and therefore may require multiple dosages and treatment regimens.

[1174] How to use

[1175] Reduce MAPT gene expression

[1176] In some embodiments, the disclosure provides for making cell or cell colony contact or to cell or cell colony delivery effective amount of oligonucleotide herein (for example, RNAi oligonucleotide) any one to reduce the method for MAPT gene expression.In some embodiments, by measuring the amount of MAPT mRNA, Tau protein or Tau activity in the cell or the reduction of level determines the reduction of MAPT gene expression.Described method includes those methods described herein and known to those of ordinary skill in the art.

[1177] In some embodiments, the disclosure provides a method for reducing the MAPT gene expression in the CNS. In some embodiments, the CNS includes brain and spinal cord. In some embodiments, the MAPT gene expression is reduced in at least one brain region. In some embodiments, brain regions include cervical spinal cord, thoracic spinal cord, lumbar spinal cord, frontal cortex, temporal cortex, cerebellum, midbrain, occipital cortex, parietal cortex, hippocampus, caudate nucleus, thalamus and brainstem. In some embodiments, the MAPT gene expression is reduced in at least one spinal cord region. In some embodiments, spinal cord regions include cervical spinal cord, thoracic spinal cord and lumbar spinal cord. In some embodiments, the MAPT gene expression is reduced in at least one brain region and at least one spinal cord region. In some embodiments, MAPT gene expression is reduced in at least one of the following: cervical spinal cord, thoracic spinal cord, lumbar spinal cord, frontal cortex, temporal cortex, cerebellum, midbrain, occipital cortex, parietal cortex, hippocampus, caudate nucleus, thalamus, brainstem, motor cortex, globus pallidus, midbrain tegmentum, substantia nigra, pons, cerebellar white matter and cerebellar dentate nucleus. In some embodiments, MAPT gene expression is reduced in at least one of lumbar spinal cord, thoracic spinal cord and cervical spinal cord. In some embodiments, MAPT gene expression is reduced in brain and / or spinal cord tissues associated with Alzheimer's disease. In some embodiments, tissues associated with AD include but are not limited to prefrontal cortex, motor cortex, temporal cortex, parietal cortex and hippocampus. In some embodiments, MAPT gene expression is reduced in brain and / or spinal cord tissues associated with progressive supranuclear palsy. In some embodiments, tissues associated with AD include, but are not limited to, caudate nucleus, globus pallidus, thalamus, midbrain tegmentum, substantia nigra, pons, cerebellar white matter, cerebellar dentate nucleus, medulla, cervical spinal cord, thoracic spinal cord, and lumbar spinal cord.

[1178] In some embodiments, after using oligonucleotides described herein, MAPT gene expression reduction continues from about 1 week to about 12 weeks. In some embodiments, after using oligonucleotides described herein, MAPT gene expression reduction continues 1,2,3,4,5,6,7,8,9,10,11 or 12 weeks. In some embodiments, after using oligonucleotides described herein, MAPT expression reduction continues from about 1 to about 4 months. In some embodiments, after using oligonucleotides described herein, MAPT expression reduction continues from about 1 to about 6 months. In some embodiments, after using oligonucleotides described herein, MAPT gene expression reduction continues 1,2,3 or 4 months. In some embodiments, after using oligonucleotides described herein, MAPT gene expression reduction continues 1,2,3,4,5 or 6 months. In some embodiments, after using oligonucleotides described herein, MAPT gene expression reduction continues from about 7 to about 91 days. In some embodiments, the reduction in MAPT gene expression persists for 7, 14, 21, 28, 35, 42, 49, 56, 63, 70, 77, 84, or 91 days after administration of an oligonucleotide described herein.

[1179] In some embodiments, after administering the oligonucleotides described herein, MAPT gene expression is reduced in at least one brain region and / or at least one spinal cord region for about 1 to about 12 weeks. In some embodiments, after administering the oligonucleotides described herein, MAPT gene expression is reduced in at least one brain region and / or at least one spinal cord region for 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 11 or 12 weeks. In some embodiments, after administering the oligonucleotides described herein, MAPT gene expression is reduced in at least one brain region and / or at least one spinal cord region for about 1 to about 4 months. In some embodiments, after administering the oligonucleotides described herein, MAPT gene expression is reduced in at least one brain region and / or at least one spinal cord region for about 1 to about 6 months. In some embodiments, after administering the oligonucleotides described herein, MAPT gene expression is reduced in at least one brain region and / or at least one spinal cord region for 1, 2, 3 or 4 months. In some embodiments, after administering the oligonucleotides described herein, MAPT gene expression is reduced in at least one brain region and / or at least one spinal cord region for 1, 2, 3, 4, 5 or 6 months. In some embodiments, after using the oligonucleotides described herein, MAPT gene expression is reduced in at least one brain region and / or at least one spinal cord region for about 7 to about 91 days. In some embodiments, after using the oligonucleotides described herein, MAPT gene expression is reduced in at least one brain region and / or at least one spinal cord region for 7, 14, 21, 28, 35, 42, 49, 56, 63, 70, 77, 84 or 91 days.

[1180] The method provided herein can be used for any appropriate cell type. In some embodiments, the cell is any cell (e.g., oligodendritic cell) expressing MAPT mRNA. In some embodiments, the cell is a primary cell obtained from a subject. In some embodiments, the primary cell has been passed down a limited number of times so that the cell generally maintains natural phenotypic characteristics. In some embodiments, the cell delivered by the oligonucleotide is ex vivo or in vitro (i.e., it can be delivered to the cell in culture or the organism in which the cell is located).

[1181] In some embodiments, the oligonucleotides disclosed herein are delivered to cells or cell colonies using nucleic acid delivery methods known in the art, including but not limited to injection of a solution or pharmaceutical composition containing the oligonucleotide, bombardment with particles covered with the oligonucleotide, exposing the cell or cell colony to a solution containing the oligonucleotide, or electroporating the cell membrane in the presence of the oligonucleotide. Other methods known in the art for delivering oligonucleotides to cells can be used, such as lipid-mediated vector transport, chemical-mediated transport, and cationic liposome transfection, such as calcium phosphate, etc.

[1182] In some embodiments, by evaluating the mensuration or technology of one or more molecules, characteristic or features of the cell or cell colony relevant to MAPT gene expression, or by evaluating the mensuration or technology of the molecule (for example, MAPT mRNA or Tau protein) of the MAPT gene expression in the direct indication cell or cell colony, determine the reduction of MAPT gene expression.In some embodiments, by comparing the MAPT gene expression in the cell or cell colony contacted with oligonucleotide with control cells or cell colony (for example, the cell or cell colony that does not contact oligonucleotide or contact control oligonucleotide) to evaluate the degree that oligonucleotide reduces MAPT gene expression.In some embodiments, the control amount or the level of MAPT gene expression in control cells or cell colony are predetermined, so that the control amount or the level do not need to be measured in each instance of measuring or technology.Predetermined level or value can adopt various forms.In some embodiments, predetermined level or value can be a single cutoff value, such as median or mean value.

[1183] In some embodiments, oligonucleotide is contacted with cell or cell colony or oligonucleotide is delivered to cell or cell colony to promote MAPT gene expression to reduce.In some embodiments, the reduction of MAPT gene expression is relative to the control amount or level of MAPT gene expression in cell or cell colony that is not contacted with oligonucleotide or contact control oligonucleotide.In some embodiments, relative to the control amount or level of MAPT gene expression, the reduction of MAPT gene expression is about 1% or lower, about 5% or lower, about 10% or lower, about 15% or lower, about 20% or lower, about 25% or lower, about 30% or lower, about 35% or lower, about 40% or lower, about 45% or lower, about 50% or lower, about 55% or lower, about 60% or lower, about 70% or lower, about 80% or lower or about 90% or lower.In some embodiments, the control amount or level of MAPT gene expression is the amount or level of MAPT mRNA and / or Tau protein in cell or cell colony that has not yet contacted the oligonucleotide herein. In some embodiments, the effect of oligonucleotide delivery to cell or cell colony is assessed according to the method herein after any limited time period or time amount (for example, minute, hour, day, week, month).For example, in some embodiments, after making oligonucleotide contact with cell or cell colony or oligonucleotide delivery to cell or cell colony at least about 4 hours, about 8 hours, about 12 hours, about 18 hours, about 24 hours; Or at least about 1 day, about 2 days, about 3 days, about 4 days, about 5 days, about 6 days, about 7 days, about 8 days, about 9 days, about 10 days, about 11 days, about 12 days, about 13 days, about 14 days, about 21 days, about 28 days, about 35 days, about 42 days, about 49 days, about 56 days, about 63 days, about 70 days, about 77 days, about 84 days or longer time, in cell or cell colony, determine MAPT gene expression. In some embodiments, MAPT gene expression is determined in a cell or cell population at least about 1 month, about 2 months, about 3 months, about 4 months, about 5 months, about 6 months, or more after contacting or delivering the oligonucleotide to the cell or cell population.

[1184] In some embodiments, the oligonucleotide is delivered in the form of a transgene that is engineered to express the oligonucleotide or a strand comprising the oligonucleotide (e.g., its sense strand and antisense strand) in a cell. In some embodiments, the oligonucleotide is delivered using a transgene that is engineered to express any of the oligonucleotides disclosed herein. The transgene can be delivered using a viral vector (e.g., adenovirus, retrovirus, vaccinia virus, poxvirus, adeno-associated virus, or herpes simplex virus) or a non-viral vector (e.g., plasmid or synthetic mRNA). In some embodiments, the transgene can be injected directly into the subject.

[1185] Treatment

[1186] The disclosure also provides oligonucleotides (e.g., RNAi oligonucleotides) for or applicable to treating a subject (e.g., a person suffering from a disease, disorder, or illness associated with MAPT gene expression) that will benefit from reducing MAPT gene expression. In some aspects, the disclosure provides oligonucleotides for or applicable to treating a subject suffering from a disease, disorder, or illness associated with MAPT gene expression. The disclosure also provides oligonucleotides for or applicable to manufacturing a medicament or pharmaceutical composition for treating a disease, disorder, or illness associated with MAPT gene expression. In some embodiments, the oligonucleotide targeting MAPT mRNA is used or is suitable for use and reduces MAPT gene expression (e.g., via RNAi pathway). In some embodiments, the oligonucleotide targeting MAPT mRNA is used or is suitable for use and reduces the amount or level of MAPT mRNA, Tau protein, and / or Tau activity.

[1187] In addition, in some embodiments of the method herein, select to suffer from the disease, disease or illness related to MAPT gene expression or be susceptible to the subject of the disease, disease or illness to treat with the oligonucleotides (e.g., ds oligonucleotides) herein. In some embodiments, the method includes selecting a marker (e.g., biomarker) for a disease, disease or illness related to MAPT gene expression or being susceptible to the subject of the disease, disease or illness, the marker such as but not limited to MAPT mRNA, Tau protein or a combination thereof. Similarly and as described in detail below, some embodiments of the method provided by the present disclosure include steps such as the following: measure or obtain the baseline value of the marker (e.g., Tau protein or Tau activity) of MAPT gene expression, then compare the value obtained with one or more other baseline values ​​or the value obtained after applying oligonucleotides to the subject to assess the effectiveness of treatment.

[1188] The disclosure also provides methods for treating a subject suffering from, suspected of suffering from, or at risk of suffering from a disease, disorder, or illness related to MAPT gene expression with the oligonucleotides provided herein. In some aspects, the disclosure provides methods for treating or alleviating the onset or progression of a disease, disorder, or illness related to MAPT gene expression using the oligonucleotides provided herein. In other aspects, the disclosure provides methods for achieving one or more therapeutic benefits in a subject suffering from a disease, disorder, or illness related to MAPT gene expression using the oligonucleotides provided herein. In some embodiments of the methods herein, a subject is treated by administering any one or more oligonucleotides provided herein of a therapeutically effective amount. In some embodiments, treatment includes reducing MAPT gene expression. In some embodiments, a subject is treated in a therapeutic manner. In some embodiments, a subject is treated in a preventive manner.

[1189] In some embodiments of the methods herein, oligonucleotides (e.g., RNAi oligonucleotides) or pharmaceutical compositions comprising the oligonucleotides are administered to subjects suffering from diseases, disorders or illnesses related to MAPT gene expression, so that the MAPT gene expression in the subject is reduced, thereby treating the subject. In some embodiments, the amount or level of MAPTmRNA in the subject is reduced. In some embodiments, the amount or level of Tau protein in the subject is reduced.

[1190] In some embodiments of the methods herein, an oligonucleotide or a pharmaceutical composition comprising the oligonucleotide is administered to a subject suffering from a disease, disorder or condition associated with MAPT gene expression such that MAPT gene expression in the subject is reduced by at least about 30%, about 35%, about 40%, about 45%, about 50%, about 55%, about 60%, about 65%, about 70%, about 75%, about 80%, about 85%, about 90%, about 95%, about 99% or greater than 99% when compared to MAPT gene expression before administration of the oligonucleotide or pharmaceutical composition. In some embodiments of the methods herein, an oligonucleotide or a pharmaceutical composition comprising the oligonucleotide is administered to a subject suffering from a disease, disorder or condition associated with MAPT gene expression such that MAPT gene expression in the subject is reduced by at least about 30%, about 35%, about 40%, about 45%, about 50%, about 55%, about 60%, about 65%, about 70%, about 75%, about 80%, about 85%, about 90%, about 95%, about 99%, or greater than 99% for about 1 week to about 12 weeks, about 1 month to about 6 months, or about 7 days to about 91 days, compared to the MAPT gene expression before administration of the oligonucleotide or pharmaceutical composition. In some embodiments, MAPT gene expression in a subject is reduced by at least about 30%, about 35%, about 40%, about 45%, about 50%, about 55%, about 60%, about 65%, about 70%, about 75%, about 80%, about 85%, about 90%, about 95%, about 99%, or greater than 99% when compared to MAPT gene expression in a subject (e.g., a reference or control subject) that did not receive the oligonucleotide or pharmaceutical composition or received a control oligonucleotide, pharmaceutical composition, or treatment. In some embodiments, MAPT gene expression in a subject is reduced by at least about 30%, about 35%, about 40%, about 45%, about 50%, about 55%, about 60%, about 65%, about 70%, about 75%, about 80%, about 85%, about 90%, about 95%, about 99%, or greater than 99% for about 1 week to about 12 weeks, about 1 month to about 6 months, or about 7 days to about 91 days when compared to MAPT gene expression in a subject (e.g., a reference or control subject) that did not receive the oligonucleotide or pharmaceutical composition or received a control oligonucleotide, pharmaceutical composition, or treatment.

[1191] In some embodiments of the methods herein, an oligonucleotide or pharmaceutical composition is administered to a subject suffering from a disease, disorder or condition associated with MAPT gene expression such that the amount or level of MAPT mRNA in the subject is reduced by at least about 30%, about 35%, about 40%, about 45%, about 50%, about 55%, about 60%, about 65%, about 70%, about 75%, about 80%, about 85%, about 90%, about 95%, about 99%, or greater than 99% when compared to the amount or level of MAPT mRNA before administration of the oligonucleotide or pharmaceutical composition. In some embodiments of the methods, an oligonucleotide or pharmaceutical composition is administered to a subject having a disease, disorder, or condition associated with MAPT gene expression such that the amount or level of MAPT mRNA in the subject is reduced by at least about 30%, about 35%, about 40%, about 45%, about 50%, about 55%, about 60%, about 65%, about 70%, about 75%, about 80%, about 85%, about 90%, about 95%, about 99%, or greater than 99% for about 1 week to about 12 weeks, about 1 month to about 6 months, or about 7 days to about 91 days, compared to the amount or level of MAPT mRNA before administration of the oligonucleotide or pharmaceutical composition. In some embodiments, the amount or level of MAPT mRNA in a subject is reduced by at least about 30%, about 35%, about 40%, about 45%, about 50%, about 55%, about 60%, about 65%, about 70%, about 75%, about 80%, about 85%, about 90%, about 95%, about 99%, or greater than 99% when compared to the amount or level of MAPT mRNA in a subject (e.g., a reference or control subject) that did not receive the oligonucleotide or pharmaceutical composition or received a control oligonucleotide, pharmaceutical composition, or treatment. In some embodiments, the amount or level of MAPT mRNA in a subject is reduced by at least about 30%, about 35%, about 40%, about 45%, about 50%, about 55%, about 60%, about 65%, about 70%, about 75%, about 80%, about 85%, about 90%, about 95%, about 99%, or greater than 99% for about 1 week to about 12 weeks, about 1 month to about 6 months, or about 7 days to about 91 days when compared to the amount or level of MAPT mRNA in a subject that did not receive the oligonucleotide or pharmaceutical composition or received a control oligonucleotide, pharmaceutical composition, or treatment (e.g., a reference or control subject).

[1192] In some embodiments of the methods, an oligonucleotide or pharmaceutical composition is administered to a subject having a disease, disorder or condition associated with MAPT gene expression such that the amount or level of Tau protein in the subject is reduced by at least about 30%, about 35%, about 40%, about 45%, about 50%, about 55%, about 60%, about 65%, about 70%, about 75%, about 80%, about 85%, about 90%, about 95%, about 99%, or greater than 99% when compared to the amount or level of Tau protein before administration of the oligonucleotide or pharmaceutical composition. In some embodiments of the methods herein, an oligonucleotide herein or a pharmaceutical composition comprising the oligonucleotide is administered to a subject having a disease, disorder, or condition associated with MAPT gene expression such that the amount or level of Tau protein in the subject is reduced by at least about 30%, about 35%, about 40%, about 45%, about 50%, about 55%, about 60%, about 65%, about 70%, about 75%, about 80%, about 85%, about 90%, about 95%, about 99%, or greater than 99% for about 1 week to about 12 weeks, about 1 month to about 6 months, or about 7 days to about 91 days, compared to the amount or level of Tau protein before administration of the oligonucleotide or pharmaceutical composition. In some embodiments, the amount or level of Tau protein in a subject is reduced by at least about 30%, about 35%, about 40%, about 45%, about 50%, about 55%, about 60%, about 65%, about 70%, about 75%, about 80%, about 85%, about 90%, about 95%, about 99%, or greater than 99% when compared to the amount or level of Tau protein in a subject not receiving the oligonucleotide or pharmaceutical composition or receiving a control oligonucleotide, pharmaceutical composition, or treatment (e.g., a reference or control subject). In some embodiments, the amount or level of Tau protein in a subject is reduced by at least about 30%, about 35%, about 40%, about 45%, about 50%, about 55%, about 60%, about 65%, about 70%, about 75%, about 80%, about 85%, about 90%, about 95%, about 99%, or greater than 99% for about 1 week to about 12 weeks, about 1 month to about 6 months, or about 7 days to about 91 days when compared to the amount or level of Tau protein in a subject not receiving the oligonucleotide or pharmaceutical composition or receiving a control oligonucleotide, pharmaceutical composition, or treatment (e.g., a reference or control subject).

[1193] In some embodiments of the methods, an oligonucleotide or pharmaceutical composition is administered to a subject having a disease, disorder, or condition associated with MAPT gene expression such that the amount or level of Tau activity in the subject is reduced by at least about 30%, about 35%, about 40%, about 45%, about 50%, about 55%, about 60%, about 65%, about 70%, about 75%, about 80%, about 85%, about 90%, about 95%, about 99%, or greater than 99% when compared to the amount or level of Tau activity before administration of the oligonucleotide or pharmaceutical composition. In some embodiments, an oligonucleotide or pharmaceutical composition is administered to a subject having a disease, disorder, or condition associated with MAPT gene expression such that the amount or level of Tau activity in the subject is reduced by at least about 30%, about 35%, about 40%, about 45%, about 50%, about 55%, about 60%, about 65%, about 70%, about 75%, about 80%, about 85%, about 90%, about 95%, about 99%, or greater than 99% for about 1 week to about 12 weeks, about 1 month to about 6 months, or about 7 days to about 91 days, compared to the amount or level of Tau activity before administration of the oligonucleotide or pharmaceutical composition. In some embodiments, the amount or level of Tau activity in a subject is reduced by at least about 30%, about 35%, about 40%, about 45%, about 50%, about 55%, about 60%, about 65%, about 70%, about 75%, about 80%, about 85%, about 90%, about 95%, about 99%, or greater than 99% when compared to the amount or level of Tau activity in a subject not receiving the oligonucleotide or pharmaceutical composition or receiving a control oligonucleotide, pharmaceutical composition, or treatment (e.g., a reference or control subject). In some embodiments, the amount or level of Tau activity in a subject is reduced by at least about 30%, about 35%, about 40%, about 45%, about 50%, about 55%, about 60%, about 65%, about 70%, about 75%, about 80%, about 85%, about 90%, about 95%, about 99%, or greater than 99% for about 1 week to about 12 weeks, about 1 month to about 6 months, or about 7 days to about 91 days when compared to the amount or level of Tau activity in a subject not receiving the oligonucleotide or pharmaceutical composition or receiving a control oligonucleotide, pharmaceutical composition, or treatment (e.g., a reference or control subject).

[1194] Suitable methods for determining the amount or level of MAPT gene expression, MAPT mRNA, Tau protein, and / or Tau activity in a sample or from a subject are known in the art. In addition, the embodiments described herein illustrate exemplary methods for determining MAPT gene expression.

[1195] In some embodiments, the expression of the MAPT gene, the amount or level of MAPT mRNA, the amount or level of Tau protein, the amount or level of Tau activity, or any combination thereof in cells (e.g., oligodendrocytes), cell colonies or a group of cells (e.g., organoids), organs (e.g., frontal cortex), blood or a portion thereof (e.g., plasma), tissues (e.g., brain tissue), samples (e.g., brain biopsy samples), or any other biological material obtained or separated from a subject is reduced. In some embodiments, the expression of the MAPT gene, the amount or level of MAPT mRNA, the amount or level of Tau protein, the amount or level of Tau activity, or any combination thereof, in more than one type of cells (e.g., oligodendrocytes and one or more other types of cells), more than one group of cells, more than one organ (e.g., brain and one or more other organs), more than one blood portion (e.g., plasma and one or more other blood portions), more than one type of tissue (e.g., brain tissue and one or more other types of tissues), more than one type of sample (e.g., brain biopsy samples and one or more other types of biopsy samples) obtained or separated from a subject is reduced. In some embodiments, the MAPT gene expression in one or more of cervical spinal cord, thoracic spinal cord, lumbar spinal cord, frontal cortex, temporal cortex, cerebellum, midbrain, occipital cortex, parietal cortex, hippocampus, caudate nucleus, thalamus, brain stem, motor cortex, globus pallidus, midbrain tegmentum, substantia nigra, pons, cerebellar white matter and cerebellar dentate nucleus, the amount or level of MAPT mRNA, the amount or level of Tau protein, the amount or level of Tau activity or any combination thereof is reduced. In some embodiments, the MAPT gene expression in the brain and / or spinal cord tissue related to AD, the amount or level of MAPT mRNA, the amount or level of Tau protein, the amount or level of Tau activity or any combination thereof is reduced. In some embodiments, the tissue related to AD includes but is not limited to prefrontal cortex, motor cortex, temporal cortex, parietal cortex and hippocampus. In some embodiments, the MAPT gene expression in brain and / or spinal cord tissues associated with PSP, the amount or level of MAPT mRNA, the amount or level of Tau protein, the amount or level of Tau activity, or any combination thereof is reduced. In some embodiments, the tissues associated with AD include but are not limited to the caudate nucleus, globus pallidus, thalamus, midbrain tegmentum, substantia nigra, pons, cerebellar white matter, cerebellar dentate nucleus, medulla, cervical spinal cord, thoracic spinal cord, and lumbar spinal cord.

[1196] The example of the disease, illness or disease related to MAPT gene expression includes but is not limited to AD, FTD, PD, PSP and Tau protein related diseases (for example, primary age-related tau disease, chronic traumatic encephalopathy, cortical basal ganglia degeneration, Litti-Bertiger disease, ganglioglioma, meningioangiomatosis, postencephalitis Parkinsonism and subacute sclerosing panencephalitis), it has the abnormal MAPT gene expression that causes the morbidity of these diseases. In MAPT, it is known that there are more than 50 kinds of missense, silence and intron mutations (Ghetti et al. (2015) NEUROPATHOL.APPL.NEUROBIOL.41:24-46) that cause these diseases.

[1197] Due to its high specificity, the oligonucleotide herein (e.g., RNAi oligonucleotide) specifically targets the mRNA of the target gene of cells, tissues or organs (e.g., brain). In preventing diseases, the target gene can be a gene required for initiating or maintaining the disease or identified as being associated with a higher risk of infectious diseases. In treating diseases, oligonucleotides can be contacted with cells, tissues or organs (e.g., brain) that exhibit or are responsible for mediating the disease. For example, all or part of substantially the same oligonucleotides of the wild type (i.e., native) or mutant genes related to the disorder or illness associated with MAPT gene expression can be contacted with a cell or tissue type (such as oligodendritic cells or other brain cells) of interest or introduced therein.

[1198] In some embodiments, the target gene can be a target gene from any mammal, such as a human.Any gene can be silenced according to the methods described herein.

[1199] The methods described herein generally involve administering to a subject a therapeutically effective amount of an oligonucleotide (e.g., an RNAi oligonucleotide), i.e., an amount capable of producing a desired therapeutic outcome. A therapeutically acceptable amount can be an amount that can be used to treat a disease or condition in a therapeutic manner. The appropriate dosage for any one subject will depend on certain factors, including the subject's size, body surface area, age, specific composition to be administered, active ingredients in the composition, time and route of administration, general health, and other drugs administered simultaneously.

[1200] In some embodiments, any of the compositions herein is administered to a subject enterally (e.g., orally, by a gastric feeding tube, by a duodenal feeding tube, via gastrostomy, or rectally), parenterally (e.g., subcutaneous injection, intravenous injection or infusion, intraarterial injection or infusion, intraosseous infusion, intramuscular injection, intracerebral injection, intraventricular injection, or intrathecal), topically (e.g., epidermal, inhalation, via eye drops, or transmucosal), or by direct injection into a target organ (e.g., the brain of a subject). Typically, the oligonucleotide is administered intravenously or subcutaneously. In some embodiments, the oligonucleotide is administered to the cerebrospinal fluid. In some embodiments, the oligonucleotide described herein is administered intrathecally. In some embodiments, the oligonucleotide is administered intraventricularly. In some embodiments, the oligonucleotide is administered by cisternal injection.

[1201] As a non-limiting example, the oligonucleotides will be administered typically quarterly (once every three months), bimonthly (once every two months), monthly, or weekly. For example, the oligonucleotides may be administered weekly or at intervals of two or three weeks. Alternatively, the oligonucleotides may be administered daily. In some embodiments, one or more loading doses of the oligonucleotides are administered to the subject, followed by one or more maintenance doses of the oligonucleotides.

[1202] In some embodiments, the subject to be treated is a human or NHP or other mammalian subject.Other exemplary subjects include domestic animals such as dogs and cats; livestock such as horses, cattle, pigs, sheep, goats and chickens; and animals such as mice, rats, guinea pigs and hamsters.

[1203] Reagent test kit

[1204] In some embodiments, the present disclosure provides a kind of kit comprising oligonucleotides (for example, RNAi oligonucleotides) and instructions for use herein.In some embodiments, the kit comprises oligonucleotides and a package insert containing instructions for use of the kit and / or any of its components.In some embodiments, the kit comprises oligonucleotides, one or more controls and various buffers, reagents, enzymes and other standard components well known in the art in a suitable container.In some embodiments, the container includes at least one vial, well, test tube, flask, bottle, syringe or other container tool, in which the oligonucleotide is placed and in some cases is appropriately divided.In some embodiments providing additional components, the kit contains additional containers in which such components are placed.The kit can also include a tool for accommodating oligonucleotides and any other reagents in a confined space for commercial sale.Such containers can include injection-molded or blow-molded plastic containers, in which required vials are retained.Containers and / or kits can include labels with instructions for use and / or warnings.

[1205] In some embodiments, the kit comprises an oligonucleotide and a pharmaceutically acceptable carrier or a pharmaceutical composition comprising the oligonucleotide, and instructions for treating or delaying progression of a disease, disorder, or condition associated with MAPT gene expression in a subject in need thereof.

[1206] In some embodiments, the kit comprises an oligonucleotide and a pharmaceutically acceptable carrier or a pharmaceutical composition comprising the oligonucleotide, and instructions for administering the oligonucleotide or the pharmaceutical composition to the cerebrospinal fluid to reduce MAPT gene expression in at least one brain region and / or at least one spinal cord region of a subject in need thereof.

[1207] definition

[1208] As used herein, "approximately" or "about" when applied to one or more values ​​of interest refers to a value similar to the reference value recited. In certain embodiments, unless otherwise specified or otherwise apparent from the context (unless such a numerical value will exceed 100% of a possible value), "about" refers to a range of values ​​within 25%, 20%, 19%, 18%, 17%, 16%, 15%, 14%, 13%, 12%, 11%, 10%, 9%, 8%, 7%, 6%, 5%, 4%, 3%, 2%, 1% or less of the reference value recited in either direction (greater than or less than).

[1209] As used herein, "administer," "administering," "administration," and the like refer to providing a substance (eg, an oligonucleotide) to a subject in a manner that is pharmacologically useful (eg, to treat a condition in the subject).

[1210] As used herein, "asialoglycoprotein receptor" or "ASGPR" refers to a bisected C-type lectin formed by a major 48 kDa subunit (ASGPR-1) and a minor 40 kDa subunit (ASGPR-2). ASGPR is primarily expressed on the sinusoidal surface of hepatocytes and plays a major role in the binding, internalization, and subsequent clearance of circulating glycoproteins containing terminal galactose or GalNAc residues (asialoglycoproteins).

[1211] As used herein, "attenuate", "attenuating", "attenuation", etc. refer to reduction or effective prevention. As a non-limiting example, one or more treatments herein can reduce or effectively prevent the onset or progression of a disease (e.g., Tau-related disease) associated with MAPT gene expression in a subject. Such attenuation can be exemplified by the following: for example, one or more aspects (e.g., symptoms, tissue characteristics, and cells, inflammatory or immune responses, etc.) of a disease (e.g., Tau-related disease) associated with MAPT gene expression are reduced, the progression (deterioration) of one or more aspects of the disease is not detected, or aspects of the disease that may otherwise be expected are not detected in the subject.

[1212] As used herein, "complementarity" refers to a structural relationship between two nucleotides (e.g., on two relative nucleic acids or on relative regions of single-stranded nucleic acids), which allows two nucleotides to form base pairs with each other. For example, the purine nucleotides of a nucleic acid complementary to the pyrimidine nucleotides of the relative nucleic acid can base pair together by forming hydrogen bonds with each other. In some embodiments, complementary nucleotides can base pair in a Watson-Crick manner or in any other manner that allows the formation of a stable duplex. In some embodiments, two nucleic acids can have a region of multiple nucleotides, and the regions are complementary to each other to form a complementary region, as described herein.

[1213] As used herein, "deoxyribonucleotide" refers to a nucleotide having a hydrogen replacing the hydroxyl group at the 2' position of its pentose when compared to a ribonucleotide. A modified deoxyribonucleotide is a deoxyribonucleotide having one or more modifications or substitutions of atoms other than the 2' position, including modifications or substitutions within or within the sugar, phosphate group, or base.

[1214] As used herein, "double-stranded oligonucleotide" or "ds oligonucleotide" refers to an oligonucleotide that is generally in the form of a duplex. In some embodiments, complementary base pairing of the duplex region of a ds oligonucleotide is formed between the antiparallel sequences of the nucleotides of the covalently separated nucleic acid chains. In some embodiments, complementary base pairing of the duplex region of a ds oligonucleotide is formed between the antiparallel sequences of the nucleotides of the covalently linked nucleic acid chains. In some embodiments, complementary base pairing of the duplex region of a ds oligonucleotide is formed by a single-stranded nucleic acid that is folded (e.g., via a hairpin) to provide complementary antiparallel sequences of nucleotides that are base-paired together. In some embodiments, a ds oligonucleotide comprises two covalently separated nucleic acid chains that are completely duplexed with each other. However, in some embodiments, a ds oligonucleotide comprises two covalently separated nucleic acid chains that are partially duplexed (e.g., having an overhang at one or both ends). In some embodiments, a ds oligonucleotide comprises an antiparallel sequence of partially complementary nucleotides, and therefore may have one or more mismatches, which may include internal mismatches or terminal mismatches.

[1215] As used herein, "duplex" with respect to nucleic acids (eg, oligonucleotides) refers to the structure formed by complementary base pairing of two antiparallel sequences of nucleotides.

[1216] As used herein, "excipient" refers to a non-therapeutic agent that can be included in the composition, for example, to provide or aid in desired consistency or stabilization.

[1217] As used herein, a "labile linker" refers to a linker that can be cleaved (eg, by acidic pH). A "fairly stable linker" refers to a linker that is not cleavable.

[1218] As used herein, "loop" refers to an unpaired region of a nucleic acid (e.g., an oligonucleotide) that is flanked by two antiparallel regions of the nucleic acid that are sufficiently complementary to each other so that under appropriate hybridization conditions (e.g., in phosphate buffer, in cells), the two antiparallel regions flanking the unpaired region hybridize to form a duplex (referred to as the "stem").

[1219] As used herein, "modified internucleotide bonds" refers to internucleotide bonds having one or more chemical modifications when compared to a reference internucleotide bond comprising a phosphodiester bond. In some embodiments, the modified nucleotide is a non-naturally occurring bond. Typically, the modified internucleotide bond confers one or more desired properties to the nucleic acid in which the modified internucleotide bond is present. For example, the modified nucleotide can improve thermal stability, resistance to degradation, nuclease resistance, solubility, bioavailability, biological activity, reduced immunogenicity, etc.

[1220] As used herein, "modified nucleotides" refer to nucleotides having one or more chemical modifications compared to the corresponding reference nucleotides selected from the group consisting of adenine ribonucleotides, guanine ribonucleotides, cytosine ribonucleotides, uracil ribonucleotides, adenine deoxyribonucleotides, guanine deoxyribonucleotides, cytosine deoxyribonucleotides, and thymidine deoxyribonucleotides. In some embodiments, modified nucleotides are non-naturally occurring nucleotides. In some embodiments, modified nucleotides have one or more chemical modifications in their sugar, nucleobase, and / or phosphate groups. In some embodiments, modified nucleotides have one or more chemical moieties conjugated to the corresponding reference nucleotides. Typically, modified nucleotides confer one or more desired properties to the nucleic acid in which the modified nucleotides are present. For example, modified nucleotides can improve thermal stability, degradation resistance, nuclease resistance, solubility, bioavailability, biological activity, reduced immunogenicity, etc.

[1221] As used herein, "nicked tetraloop structure" refers ...

Claims

1. An RNAi oligonucleotide for reducing MAPT gene expression, said oligonucleotide comprising a sense strand and an antisense strand, wherein said sense strand and said antisense strand form a duplex region, wherein said antisense strand comprises a complementary region to a MAPT mRNA target sequence of any one of SEQ ID NO:912-1295, and wherein said complementary region has a length of at least 15 continuous nucleotides.

2. The RNAi oligonucleotide of claim 1, wherein the sense strand is 15 to 50 nucleotides in length.

3. The RNAi oligonucleotide of claim 1 or 2, wherein the sense strand is 18 to 36 nucleotides in length.

4. The RNAi oligonucleotide of any one of claims 1 to 3, wherein the antisense strand is 15 to 30 nucleotides in length.

5. The RNAi oligonucleotide of any one of claims 1 to 4, wherein the antisense strand is 22 nucleotides in length, and wherein the antisense strand and the sense strand form a duplex region having a length of at least 19 nucleotides, optionally at least 20 nucleotides in length.

6. The RNAi oligonucleotide of any one of claims 1 to 5, wherein the length of the complementary region is at least 19 consecutive nucleotides.

7. The RNAi oligonucleotide of any one of claims 1 to 6, wherein the length of the complementary region is at least 20 consecutive nucleotides.

8. A double-stranded RNAi oligonucleotide for reducing MAPT gene expression, the oligonucleotide comprising: (i) an antisense strand of 19-30 nucleotides in length, wherein the antisense strand comprises a nucleotide sequence comprising a complementary region to a MAPT mRNA target sequence, wherein the complementary region is selected from SEQ ID NOs: 1296-1679, and (ii) a sense strand of 19-50 nucleotides in length, the sense strand comprising a complementary region to the antisense strand, wherein the antisense strand and the sense strand are independent strands forming an asymmetric duplex region having an overhang of 1-4 nucleotides at the 3' end of the antisense strand.

9. The RNAi oligonucleotide of any one of claims 1-8, wherein the 3' end of the sense strand comprises a stem loop as shown in S1-L-S2, wherein S1 is complementary to S2, and wherein L forms a loop of 3-5 nucleotides in length between S1 and S2.

10. The RNAi oligonucleotide of claim 9, wherein L is a tricyclic or tetracyclic ring.

11. The RNAi oligonucleotide of claim 10, wherein L is a tetracyclic ring.

12. The RNAi oligonucleotide of claim 11, wherein the tetraloop comprises the sequence 5'-GAAA-3'.

13. The RNAi oligonucleotide of any one of claims 9 to 12, wherein the S1 and S2 are 1 to 10 nucleotides in length and have the same length.

14. The RNAi oligonucleotide of claim 13, wherein the length of S1 and S2 is 1 nucleotide, 2 nucleotides, 3 nucleotides, 4 nucleotides, 5 nucleotides, 6 nucleotides, 7 nucleotides, 8 nucleotides, 9 nucleotides or 10 nucleotides.

15. The RNAi oligonucleotide of claim 14, wherein S1 and S2 are 6 nucleotides in length.

16. The RNAi oligonucleotide of any one of claims 9-15, wherein the stem-loop comprises the sequence 5'-GCAGCCGAAAGGCUGC-3' (SEQ ID NO: 1680).

17. The RNAi oligonucleotide of any one of claims 1-8, wherein the oligonucleotide comprises a blunt end.

18. The RNAi oligonucleotide of claim 17, wherein the blunt end comprises the 3' end of the sense strand.

19. The RNAi oligonucleotide of claim 17 or 18, wherein the sense strand is 20-22 nucleotides.

20. The RNAi oligonucleotide of claim 19, wherein the sense strand is 20 nucleotides.

21. The RNAi oligonucleotide of any one of claims 1 to 20, wherein the antisense strand comprises a 3' overhang sequence of one or more nucleotides in length.

22. The RNAi oligonucleotide of claim 21, wherein the overhang comprises a purine nucleotide.

23. The RNAi oligonucleotide of claim 21 or 22, wherein the 3' overhang sequence is 2 nucleotides in length.

24. The RNAi oligonucleotide of claim 23, wherein the 3' overhang is selected from the group consisting of AA, GG, AG and GA.

25. The RNAi oligonucleotide of claim 24, wherein the 3' overhang is GG or AA.

26. The RNAi oligonucleotide of claim 25, wherein the 3' overhang is GG.

27. The RNAi oligonucleotide of any one of the preceding claims, wherein the oligonucleotide comprises at least one modified nucleotide.

28. The RNAi oligonucleotide of claim 27, wherein the modified nucleotide comprises a 2'-modification.

29. The RNAi oligonucleotide of claim 28, wherein the 2'-modification is a modification selected from 2'-aminoethyl, 2'-fluoro, 2'-O-methyl, 2'-O-methoxyethyl and 2'-deoxy-2'-fluoro-β-d-arabino nucleic acid.

30. The RNAi oligonucleotide of any one of claims 28-29, wherein the modification is a 2'-modification selected from 2'-fluoro and 2'-O-methyl.

31. The RNAi oligonucleotide of any one of claims 29-30, wherein about 18-23%, 18%, 19%, 20%, 21%, 22% or 23% of the nucleotides of the sense strand comprise a 2'-fluoro modification.

32. The RNAi oligonucleotide of any one of claims 29-30, wherein about 38-43%, 38%, 39%, 40%, 41%, 42% or 43% of the nucleotides of the sense strand comprise a 2'-fluoro modification.

33. The RNAi oligonucleotide of any one of claims 29-32, wherein about 25-35%, 25%, 26%, 27%, 28%, 29%, 30%, 31%, 32%, 33%, 34% or 35% of the nucleotides of the antisense strand comprise a 2'-fluoro modification.

34. The RNAi oligonucleotide of any one of claims 29-33, wherein about 25-35%, 25%, 26%, 27%, 28%, 29%, 30%, 31%, 32%, 33%, 34% or 35% of the nucleotides of the oligonucleotide comprise a 2'-fluoro modification.

35. The RNAi oligonucleotide of any one of claims 29-33, wherein about 35-45%, 35%, 36%, 37%, 38%, 39%, 40%, 41%, 42%, 43%, 44% or 45% of the nucleotides of the oligonucleotide comprise a 2'-fluorine modification.

36. The RNAi oligonucleotide of any one of claims 29-35, wherein the sense strand comprises 36 nucleotides from 5' to 3' at positions 1-36, wherein each of positions 3, 5, 8, 10, 12, 13, 15 and 17 comprises a 2'-fluoro modification.

37. The RNAi oligonucleotide of any one of claims 29-35, wherein the sense strand comprises 20 nucleotides from 5' to 3' at positions 1-20, wherein each of positions 3, 5, 8, 10, 12, 13, 15 and 17 comprises a 2'-fluoro modification.

38. The RNAi oligonucleotide of any one of claims 29-37, wherein the antisense strand comprises 22 nucleotides from 5' to 3' at positions 1-22, and wherein each of positions 2, 3, 4, 5, 7, 10, 14, 16 and 19 comprises a 2'-fluoro modification.

39. The RNAi oligonucleotide of any one of claims 31-38, wherein the remaining nucleotides comprise a 2'-O-methyl modification.

40. The RNAi oligonucleotide of any one of the preceding claims, wherein the oligonucleotide comprises at least one modified internucleotide bond.

41. The RNAi oligonucleotide of claim 40, wherein the at least one modified internucleotide bond is a phosphorothioate bond.

42. The RNAi oligonucleotide of claim 41, wherein the antisense strand comprises phosphorothioate bonds at: (i) between positions 1 and 2 and between positions 2 and 3; or (ii) between positions 1 and 2, between positions 2 and 3, and between positions 3 and 4, wherein positions are numbered 1-4 from 5' to 3'.

43. The RNAi oligonucleotide of claim 41 or 42, wherein the antisense strand is 22 nucleotides in length, and wherein the antisense strand comprises phosphorothioate bonds between positions 20 and 21 and between positions 21 and 22, and wherein positions are numbered 1-22 from 5' to 3'.

44. The RNAi oligonucleotide of any one of claims 41-43, wherein the sense strand comprises a phosphorothioate bond between positions 1 and 2, and wherein positions are numbered 1-2 from 5' to 3'.

45. The RNAi oligonucleotide of any one of claims 41-43, wherein the sense strand is 20 nucleotides in length, wherein the sense strand comprises phosphorothioate bonds between positions 1 and 2, between positions 18 and 19, and between positions 19 and 20, and wherein positions are numbered 1-20 from 5' to 3'.

46. ​​The RNAi oligonucleotide of any of the preceding claims, wherein the 4'-carbon of the sugar of the 5'-nucleotide of the antisense strand comprises a phosphate analog.

47. The RNAi oligonucleotide of claim 46, wherein the phosphate analog is oxymethylphosphonate, vinylphosphonate or malonylphosphonate, optionally wherein the phosphate analog is a 4'-phosphate analog comprising 4'-oxymethylphosphonate.

48. The RNAi oligonucleotide of any of the preceding claims, wherein at least one nucleotide of the oligonucleotide is conjugated to one or more targeting ligands.

49. The RNAi oligonucleotide of claim 48, wherein each targeting ligand comprises a carbohydrate, an amino sugar, a lipid, cholesterol, or a polypeptide.

50. The RNAi oligonucleotide of any one of claims 9-16 and 21-49, wherein the stem-loop comprises one or more targeting ligands conjugated to one or more nucleotides of the stem-loop.

51. The RNAi oligonucleotide of claim 50, wherein the one or more targeting ligands are conjugated to one or more nucleotides of the loop.

52. The RNAi oligonucleotide of claim 51, wherein the loop comprises 4 nucleotides numbered 1-4 from 5' to 3', wherein the nucleotides at positions 2, 3 and 4 each comprise one or more targeting ligands, and wherein the targeting ligands are the same or different.

53. The RNAi oligonucleotide of claim 52, wherein each targeting ligand comprises an N-acetylgalactosamine (GalNAc) moiety.

54. The RNAi oligonucleotide of claim 53, wherein the GalNac moiety is a monovalent GalNAc moiety, a divalent GalNAc moiety, a trivalent GalNAc moiety, or a tetravalent GalNAc moiety.

55. The RNAi oligonucleotide of any one of claims 9-16 and 21-54, wherein up to 4 nucleotides of L of the stem-loop are each conjugated to a monovalent GalNAc moiety.

56. The RNAi oligonucleotide of any one of claims 17-48, wherein the one or more targeting ligands is a lipid moiety.

57. The RNAi oligonucleotide of claim 56, wherein the lipid moiety is conjugated to the 5' terminal nucleotide of the sense strand.

58. The RNAi oligonucleotide of claim 56 or 57, wherein the lipid moiety is a hydrocarbon chain.

59. The RNAi oligonucleotide of claim 58, wherein the hydrocarbon chain is C8-C 30 Hydrocarbon chain.

60. The RNAi oligonucleotide of claim 58 or 59, wherein the hydrocarbon chain is C 16 Hydrocarbon chain.

61. The RNAi oligonucleotide of claim 60, wherein the C 16 The hydrocarbon chain is represented by:

62. The RNAi oligonucleotide of any one of claims 56-61, wherein the lipid moiety is conjugated to the 2' carbon of the ribose ring of the 5' terminal nucleotide.

63. The RNAi oligonucleotide of any one of claims 1-62, wherein the complementary region is fully complementary to the MAPT mRNA target sequence at nucleotide positions 2-8 of the antisense strand, and wherein nucleotide positions are numbered from 5' to 3'.

64. The RNAi oligonucleotide of any one of claims 1-62, wherein the complementary region is fully complementary to the MAPT mRNA target sequence at nucleotide positions 2-11 of the antisense strand, and wherein nucleotide positions are numbered from 5' to 3'.

65. The RNAi oligonucleotide of any one of claims 1 to 64, wherein the sense strand comprises the nucleotide sequence of any one of SEQ ID NOs: 769-803 and 1681.

66. The RNAi oligonucleotide of any one of claims 1 to 65, wherein the antisense strand comprises the nucleotide sequence of any one of SEQ ID NOs: 804-838.

67. The RNAi oligonucleotide of any one of claims 1 to 66, wherein the sense strand and the antisense strand comprise a nucleotide sequence selected from the group consisting of: a) SEQ ID NO: 769 and 804, respectively; b) SEQ ID NOs: 770 and 805, respectively; c) SEQ ID NOs: 771 and 806, respectively; d) SEQ ID NOs: 772 and 807, respectively; e) SEQ ID NOs: 773 and 808, respectively; f) SEQ ID NOs: 774 and 809, respectively; g) SEQ ID NOs: 775 and 810, respectively; h) SEQ ID NOs: 776 and 811, respectively; i) SEQ ID NOs: 777 and 812, respectively; j) SEQ ID NOs: 778 and 813, respectively; k) SEQ ID NOs: 779 and 814, respectively; 1) are SEQ ID NOs: 780 and 815, respectively; m) are SEQ ID NOs: 781 and 816, respectively; n) are SEQ ID NOs: 782 and 817, respectively; o) are SEQ ID NOs: 783 and 818, respectively; p) are SEQ ID NOs: 784 and 819, respectively; q) are SEQ ID NOs: 785 and 820, respectively; r) are SEQ ID NOs: 786 and 821, respectively; s) are SEQ ID NOs: 787 and 822, respectively; t) are SEQ ID NOs: 788 and 823, respectively; u) are SEQ ID NOs: 789 and 824, respectively; v) SEQ ID NOs: 790 and 825, respectively; w) are SEQ ID NOs: 791 and 826, respectively; x) are SEQ ID NOs: 792 and 827, respectively; y) are SEQ ID NOs: 793 and 828, respectively; z) are SEQ ID NOs: 794 and 829, respectively; aa) are SEQ ID NOs: 795 and 830, respectively; bb) are SEQ ID NOs: 796 and 831, respectively; cc) are SEQ ID NOs: 797 and 832, respectively; dd) are SEQ ID NOs: 798 and 833, respectively; ee) are SEQ ID NOs: 799 and 834, respectively; ff) are SEQ ID NOs: 800 and 835, respectively; gg) are SEQ ID NOs: 801 and 836, respectively; hh) are SEQ ID NOs: 802 and 837, respectively; ii) are SEQ ID NOs: 803 and 838, respectively; and jj) are SEQ ID NOs: 1681 and 815, respectively.

68. The RNAi oligonucleotide of any one of claims 1 to 66, wherein the sense strand and the antisense strand comprise a nucleotide sequence selected from the group consisting of: a) SEQ ID NO: 771 and 806, respectively; b) SEQ ID NOs: 776 and 811, respectively; c) SEQ ID NOs: 780 and 815, respectively; d) SEQ ID NOs: 781 and 816, respectively; e) SEQ ID NOs: 782 and 817, respectively; f) SEQ ID NOs: 790 and 825, respectively; g) SEQ ID NOs: 795 and 830, respectively; h) SEQ ID NOs: 798 and 833, respectively; i) SEQ ID NOs: 799 and 834, respectively; j) SEQ ID NOs: 803 and 838, respectively; and k) are SEQ ID NOs: 1681 and 815, respectively.

69. The RNAi oligonucleotide of any one of claims 1 to 66, wherein the sense strand and the antisense strand comprise a nucleotide sequence selected from the group consisting of: a) SEQ ID NO: 771 and 806, respectively; b) SEQ ID NOs: 780 and 815, respectively; c) SEQ ID NOs: 781 and 816, respectively; d) SEQ ID NOs: 798 and 833, respectively; e) SEQ ID NOs: 799 and 834, respectively; f) SEQ ID NOs: 803 and 838, respectively; and g) are SEQ ID NOs: 1681 and 815, respectively.

70. The RNAi oligonucleotide of any one of claims 1 to 66, wherein the sense strand comprises the nucleotide sequence shown in SEQ ID NO: 771, and wherein the antisense strand comprises the nucleotide sequence shown in SEQ ID NO:

806.

71. The RNAi oligonucleotide of any one of claims 1 to 66, wherein the sense strand comprises the nucleotide sequence shown in SEQ ID NO: 780, and wherein the antisense strand comprises the nucleotide sequence shown in SEQ ID NO:

815.

72. The RNAi oligonucleotide of any one of claims 1 to 66, wherein the sense strand comprises the nucleotide sequence shown in SEQ ID NO: 781, and wherein the antisense strand comprises the nucleotide sequence shown in SEQ ID NO:

816.

73. The RNAi oligonucleotide of any one of claims 1 to 66, wherein the sense strand comprises the nucleotide sequence shown in SEQ ID NO: 798, and wherein the antisense strand comprises the nucleotide sequence shown in SEQ ID NO:

833.

74. The RNAi oligonucleotide of any one of claims 1 to 66, wherein the sense strand comprises the nucleotide sequence shown in SEQ ID NO: 799, and wherein the antisense strand comprises the nucleotide sequence shown in SEQ ID NO:

834.

75. The RNAi oligonucleotide of any one of claims 1 to 66, wherein the sense strand comprises the nucleotide sequence shown in SEQ ID NO: 803, and wherein the antisense strand comprises the nucleotide sequence shown in SEQ ID NO:

838.

76. The RNAi oligonucleotide of any one of claims 1 to 66, wherein the sense strand comprises the nucleotide sequence shown in SEQ ID NO: 1681, and wherein the antisense strand comprises the nucleotide sequence shown in SEQ ID NO:

815.

77. The RNAi oligonucleotide of any one of claims 1-64, wherein the antisense strand is 22 nucleotides in length.

78. The RNAi oligonucleotide of claim 77, wherein the antisense strand comprises a nucleotide sequence selected from the group consisting of SEQ ID NOs: 806, 815, 816, 833, 834, and 838.

79. The RNAi oligonucleotide of any one of claims 1-64 and 77-78, wherein the sense strand comprises a nucleotide sequence selected from the group consisting of SEQ ID NOs: 1130, 1095, 1096, 1119, 1120 and 1124.

80. The RNAi oligonucleotide of any one of claims 1-64 and 77-79, wherein the sense strand is 36 nucleotides in length.

81. The RNAi oligonucleotide of claim 80, wherein the sense strand comprises a nucleotide sequence selected from the group consisting of SEQ ID NOs: 771, 780, 781, 798, 799 and 803.

82. The RNAi oligonucleotide of any one of claims 1-64, wherein the sense strand comprises the nucleotide sequence of any one of SEQ ID NOs: 839-873 and 1682.

83. The RNAi oligonucleotide of any one of claims 1 to 64 and 82, wherein the antisense strand comprises the nucleotide sequence of any one of SEQ ID NOs: 874-908.

84. The RNAi oligonucleotide of any one of claims 1-64, wherein the sense strand and the antisense strand comprise a nucleotide sequence selected from the group consisting of: a) SEQ ID NO: 839 and 874, respectively; b) SEQ ID NOs: 840 and 875, respectively; c) SEQ ID NOs: 841 and 876, respectively; d) SEQ ID NOs: 842 and 877, respectively; e) SEQ ID NOs: 843 and 878, respectively; f) SEQ ID NOs: 844 and 879, respectively; g) SEQ ID NOs: 845 and 880, respectively; h) are SEQ ID NOs: 846 and 881, respectively; i) are SEQ ID NOs: 847 and 882, respectively; j) are SEQ ID NOs: 848 and 883, respectively; k) are SEQ ID NOs: 849 and 884, respectively; 1) are SEQ ID NOs: 850 and 885, respectively; m) are SEQ ID NOs: 851 and 886, respectively; n) are SEQ ID NOs: 852 and 887, respectively; o) are SEQ ID NOs: 853 and 888, respectively; p) are SEQ ID NOs: 854 and 889, respectively; q) are SEQ ID NOs: 855 and 890, respectively; r) are SEQ ID NOs: 856 and 891, respectively; s) are SEQ ID NOs: 857 and 892, respectively; t) are SEQ ID NOs: 858 and 893, respectively; u) are SEQ ID NOs: 859 and 894, respectively; v) SEQ ID NOs: 860 and 895, respectively; w) are SEQ ID NOs: 861 and 896, respectively; x) are SEQ ID NOs: 862 and 897, respectively; y) are SEQ ID NOs: 863 and 898, respectively; z) are SEQ ID NOs: 864 and 899, respectively; aa) are SEQ ID NOs: 865 and 900, respectively; bb) are SEQ ID NOs: 866 and 901, respectively; cc) are SEQ ID NOs: 867 and 902, respectively; dd) are SEQ ID NOs: 868 and 903, respectively; ee) are SEQ ID NOs: 869 and 904, respectively; ff) are SEQ ID NOs: 870 and 905, respectively; gg) are SEQ ID NOs: 871 and 906, respectively; hh) are SEQ ID NOs: 872 and 907, respectively; ii) SEQ ID NO: 873 and 908, respectively; and jj) are SEQ ID NOs: 1682 and 885, respectively.

85. The RNAi oligonucleotide of any one of claims 1-64, wherein the sense strand and the antisense strand comprise a nucleotide sequence selected from the group consisting of: a) SEQ ID NO: 860 and 895, respectively; b) SEQ ID NO: 865 and 900, respectively; c) SEQ ID NOs: 868 and 903, respectively; d) SEQ ID NO: 869 and 904, respectively; e) SEQ ID NOs: 873 and 908, respectively; f) SEQ ID NOs: 841 and 876, respectively; g) SEQ ID NOs: 846 and 881, respectively; h) SEQ ID NOs: 850 and 885, respectively; i) SEQ ID NOs: 851 and 886, respectively; j) SEQ ID NOs: 852 and 887, respectively; and k) are SEQ ID NOs: 1682 and 885, respectively.

86. The RNAi oligonucleotide of any one of claims 1-64, wherein the sense strand and the antisense strand comprise a nucleotide sequence selected from the group consisting of: a) SEQ ID NO: 841 and 876, respectively; b) SEQ ID NOs: 850 and 885, respectively; c) SEQ ID NOs: 851 and 886, respectively; d) SEQ ID NO: 868 and 903, respectively; e) SEQ ID NOs: 869 and 904, respectively; f) SEQ ID NOs: 873 and 908, respectively; and g) are SEQ ID NOs: 1682 and 885, respectively.

87. The RNAi oligonucleotide of any one of claims 1 to 64, wherein the sense strand comprises the nucleotide sequence shown in SEQ ID NO:841, and wherein the antisense strand comprises the nucleotide sequence shown in SEQ ID NO:

876.

88. The RNAi oligonucleotide of any one of claims 1 to 64, wherein the sense strand comprises the nucleotide sequence shown in SEQ ID NO:850, and wherein the antisense strand comprises the nucleotide sequence shown in SEQ ID NO:

885.

89. The RNAi oligonucleotide of any one of claims 1 to 64, wherein the sense strand comprises the nucleotide sequence shown in SEQ ID NO: 851, and wherein the antisense strand comprises the nucleotide sequence shown in SEQ ID NO:

886.

90. The RNAi oligonucleotide of any one of claims 1 to 64, wherein the sense strand comprises the nucleotide sequence shown in SEQ ID NO: 868, and wherein the antisense strand comprises the nucleotide sequence shown in SEQ ID NO:

903.

91. The RNAi oligonucleotide of any one of claims 1 to 64, wherein the sense strand comprises the nucleotide sequence shown in SEQ ID NO: 869, and wherein the antisense strand comprises the nucleotide sequence shown in SEQ ID NO:

904.

92. The RNAi oligonucleotide of any one of claims 1 to 64, wherein the sense strand comprises the nucleotide sequence shown in SEQ ID NO: 873, and wherein the antisense strand comprises the nucleotide sequence shown in SEQ ID NO:

908.

93. The RNAi oligonucleotide of any one of claims 1 to 64, wherein the sense strand comprises the nucleotide sequence shown in SEQ ID NO: 1682, and wherein the antisense strand comprises the nucleotide sequence shown in SEQ ID NO:

885.

94. The RNAi oligonucleotide of claim 1, wherein the sense strand comprises 5'-[mAs][mG][fA][mG][fU][mG][mU][fG][mG][fA][mA][fA][mA][fA][mA][fA][mA][mG][mA][mG][mC][mA][mG][mC][mG][adem A-GalNAc][ademA-GalNAc][ademA-GalNAc][mG][mG][mC][mU][mG][mC]-3' (SEQ ID NO:841) and all modifications, wherein the antisense strand comprises the sequence of 5'-[Me phosphonate-4O-mUs][fCs][fU][fU][fU][mU][fU][mU][mU][fU][mC][mC][fA][mC][fA][mC][mU][fC][mUs][mGs][mG]-3' (SEQ ID NO:876) and all modifications, wherein mC, mA, mG and mU = 2'-OMe ribonucleoside; fA, fC, fG and fU = 2'-F ribonucleoside; s = phosphorothioate, and wherein ademA-GalNAc = 95. The RNAi oligonucleotide of claim 1, wherein the sense strand comprises 5'-[mCs][mA][fG][mG][fU][mG][mG][fA][mA][fG][mU][fA][fA][mA][fA][mU][fC][mU][mG][mA][mG][mC][mA][mG][mC][mC][mG][ademA-GalNAc][ademA-GalNAc][ademA-GalNAc][mG][mG][mC][mU][mG][mC]-3' (SEQ ID NO:850) and all modifications, wherein the antisense strand comprises the sequence of 5'-[Me phosphonate-4O-mUs][fCs][fA][fG][fA][mU][fU][mU][mU][fA][mC][mU][mU][fC][mC][fA][mC][mC][fU][mGs][mGs][mG]-3' (SEQ ID NO:885) and all modifications, wherein mC, mA, mG and mU = 2'-OMe ribonucleoside; fA, fC, fG and fU = 2'-F ribonucleoside; s = phosphorothioate, and wherein ademA-GalNAc = 96. The RNAi oligonucleotide of claim 1, wherein the sense strand comprises 5'-[mAs][mG][fG][mU][fG][mG][mA][fA][mG][fU][mA][fA][fA][mA][fU][mC][fU][mG][mA][mA][mG][mC][mA][mG][mC][mC][mG][adem A-GalNAc][ademA-GalNAc][ademA-GalNAc][mG][mG][mC][mU][mG][mC]-3' (SEQ ID NO:851) and all modifications, wherein the antisense strand comprises the sequence of 5'-[Me phosphonate-4O-mUs][fUs][fC][fA][fG][mA][fU][mU][mU][fU][mA][mC][mU][fU][mC][fC][mA][mC][fC][mUs][mGs][mG]-3' (SEQ ID NO:886) and all modifications, wherein mC, mA, mG and mU = 2'-OMe ribonucleoside; fA, fC, fG and fU = 2'-F ribonucleoside; s = phosphorothioate, and wherein ademA-GalNAc = 97. The RNAi oligonucleotide of claim 1, wherein the sense strand comprises 5'-[mAs][mG][fG][mA][fA][mA][mU][fA][mA][fA][fA][fG][mA][fU][mU][fG][mA][mA][mA][mG][mC][mA][mG][mC][mC][mG][adem A-GalNAc][ademA-GalNAc][ademA-GalNAc][mG][mG][mC][mU][mG][mC]-3' (SEQ ID NO:868) and all modifications, wherein the antisense strand comprises the sequence of 5'-[Me phosphonate-4O-mUs][fUs][fU][fC][fA][mA][fU][mC][mU][fU][mU][mU][mU][fA][mU][fU][mU][mC][fC][mUs][mGs][mG]-3' (SEQ ID NO:903) and all modifications, wherein mC, mA, mG and mU = 2'-OMe ribonucleoside; fA, fC, fG and fU = 2'-F ribonucleoside; s = phosphorothioate, and wherein ademA-GalNAc = 98. The RNAi oligonucleotide of claim 1, wherein the sense strand comprises 5'-[mGs][mG][fA][mA][fA][mU][mA][fA][mA][fA][mA][fG][fA][mU][fU][mG][fA][mA][mA][mA][mG][mC][mA][mG][mC][mC][mG][adem A-GalNAc][ademA-GalNAc][ademA-GalNAc][mG][mG][mC][mU][mG][mC]-3' (SEQ ID NO:869) and all modifications, wherein the antisense strand comprises the sequence of 5'-[Me phosphonate-4O-mUs][fUs][fU][fU][fC][mA][fA][mU][mC][fU][mU][mU][mU][fU][mA][fU][mU][mU][fC][mCs][mGs][mG]-3' (SEQ ID NO:904) and all modifications, wherein mC, mA, mG and mU = 2'-OMe ribonucleoside; fA, fC, fG and fU = 2'-F ribonucleoside; s = phosphorothioate, and wherein ademA-GalNAc = 99. The RNAi oligonucleotide of claim 1, wherein the sense strand comprises 5'-[mAs][mU][fA][mA][fA][mA][mA][fG][mA][fU][mU][fG][fA][mA][fA][mC][fC][mC][mA][mA][mG][mC][mA][mG][mC][mC][mG][adem A-GalNAc][ademA-GalNAc][ademA-GalNAc][mG][mG][mC][mU][mG][mC]-3' (SEQ ID NO:873) and all modifications, wherein the antisense strand comprises the sequence of 5'-[Me phosphonate-4O-mUs][fUs][fG][fG][fG][mU][fU][mU][mC][fA][mA][mU][mC][fU][mU][fU][mU][mU][fA][mUs][mGs][mG]-3' (SEQ ID NO:908) and all modifications, wherein mC, mA, mG and mU = 2'-OMe ribonucleoside; fA, fC, fG and fU = 2'-F ribonucleoside; s = phosphorothioate, and wherein ademA-GalNAc = 100. The RNAi oligonucleotide of claim 1, wherein the sense strand comprises 5'-[ademCs-C 16 ][mA][fG][mG][fU][mG][mG][fA][mA][fG][mU][fA][fA][mA][fA][mU][fC][mUs][mGs][mA]-3' (SEQ ID NO: 1682) and all modifications, wherein the antisense strand comprises 5'-[Me phosphonate-4O-mUs][fCs][fA][fG][fA][mU][fU][mU][mU][fA][mC][mU][mU][fC][mC][fA][mC][mC][fU][mGs][mG]-3' (SEQ ID NO: 1684). NO:885) and all modifications, wherein mC, mA, mG, and mU = 2'-OMe ribonucleosides; fA, fC, fG, and fU = 2'-F ribonucleosides; s = phosphorothioate, and [ademCs-C16] = conjugated to C 16 Hydrocarbon chain of cytosine.

101. A pharmaceutical composition comprising the RNAi oligonucleotide of any one of claims 1 to 100 and a pharmaceutically acceptable carrier, delivery agent or excipient.

102. A method for treating a subject suffering from a disease, disorder or condition associated with MAPT gene expression, the method comprising administering to the subject a therapeutically effective amount of the RNAi oligonucleotide of any one of claims 1 to 100 or the pharmaceutical composition of claim 101, thereby treating the subject.

103. A method of delivering an oligonucleotide to a subject, the method comprising administering to the subject the pharmaceutical composition of claim 101.

104. A method for reducing MAPT gene expression in a cell, a cell population or a subject, the method comprising the steps of: i. contacting the cell or the cell population with the RNAi oligonucleotide of any one of claims 1 to 100 or the pharmaceutical composition of claim 101; or ii. administering the RNAi oligonucleotide of any one of claims 1 to 100 or the pharmaceutical composition of claim 101 to the subject.

105. The method of claim 104, wherein reducing MAPT gene expression comprises reducing the amount or level of MAPT mRNA, the amount or level of Tau protein, or both.

106. The method of claim 104 or 105, wherein the subject suffers from a disease, disorder or condition associated with MAPT gene expression.

107. methods as claimed in claim 102 or 106, wherein the disease, disorder or illness associated with MAPT gene expression is Alzheimer's disease (AD), frontotemporal dementia, progressive supranuclear palsy (PSP), Tau protein related diseases, primary age-related tau disease, chronic traumatic encephalopathy, cortical basal ganglia degeneration, Litiko-Bertiger disease, ganglioglioma, meningioangiomatosis, Parkinson's disease (PD), postencephalitic parkinsonism or subacute sclerosing panencephalitis.

108. The method of any one of claims 102-107, wherein MAPT is reduced in tissue of one or more regions of the central nervous system (CNS), wherein the tissue is associated with AD.

109. The method of claim 108, wherein the AD-associated tissue is selected from the group consisting of prefrontal cortex, motor cortex, temporal cortex, parietal cortex, and hippocampus.

110. The method of any one of claims 102-107, wherein MAPT is reduced in tissue of one or more regions of the central nervous system (CNS), wherein the tissue is associated with PSP.

111. The method of claim 110, wherein the tissue associated with PSP is selected from the group consisting of the caudate nucleus, globus pallidus, thalamus, midbrain tegmentum, substantia nigra, pons, cerebellar white matter, cerebellar dentate nucleus, medulla, cervical spinal cord, thoracic spinal cord, and lumbar spinal cord.

112. methods as described in any one in claims 102-107, wherein MAPT reduces in one or more regions of CNS, and described one or more regions are selected from cervical spinal cord, thoracic spinal cord, lumbar spinal cord, frontal cortex, temporal cortex, cerebellum, midbrain, occipital cortex, parietal cortex, hippocampus, caudate nucleus, thalamus, brainstem, motor cortex, globus pallidus, midbrain tegmentum, substantia nigra, pons, cerebellar white matter and cerebellar dentate nucleus.

113. The method of any one of claims 102-112, wherein the RNAi oligonucleotide or the pharmaceutical composition is administered in combination with a second composition or therapeutic agent.

114. Use of the RNAi oligonucleotide of any one of claims 1 to 100 or the pharmaceutical composition of claim 101 for the manufacture of a medicament for treating a disease, disorder or condition associated with MAPT gene expression.

115. The RNAi oligonucleotide of any one of claims 1 to 100 or the pharmaceutical composition of claim 101, which is used or suitable for treating a disease, disorder or condition associated with MAPT gene expression.

116. A kit comprising the RNAi oligonucleotide of any one of claims 1 to 100, an optional pharmaceutically acceptable carrier, and a package insert comprising instructions for administration to a subject suffering from a disease, disorder or condition associated with expression of the MAPT gene.

117. The use of claim 114, the RNAi oligonucleotide or pharmaceutical composition for use or suitable for use as claimed in claim 115, or the kit as claimed in claim 116, wherein the disease, disorder or condition associated with MAPT gene expression is Alzheimer's disease (AD), frontotemporal dementia, progressive supranuclear palsy (PSP), Tau protein-related disease, primary age-related tauopathy, chronic traumatic encephalopathy, cortical basal degeneration, Litiko-Bertiger disease, ganglioglioma, meningioangiomatosis, Parkinson's disease (PD), postencephalitic Parkinsonism or subacute sclerosing panencephalitis.

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