Compositions and Methods for Inhibiting MAPT Expression

Abstract

Oligonucleotides that inhibit the expression of the MAPT gene and contain oligonucleotides bound to a targeting ligand (e.g., a lipid moiety) are provided herein. Also provided are compositions containing the same and their use, particularly their use in the treatment of diseases, disorders, and / or conditions associated with the expression of the MAPT gene.

Description

Technical Field

[0001] Cross - reference to Related Applications This application claims priority under 35 U.S.C. § 119(e) to U.S. Provisional Application No. 63 / 364,609, filed May 12, 2022, the entire disclosure of which is incorporated herein by reference.

[0002] Technical Field The present disclosure generally relates to biology and medicine, and more particularly to oligonucleotides for inhibiting or reducing (i.e., modulating) the expression of the microtubule - associated protein tau (MAPT) gene and compositions containing the same, and their use for treating diseases and disorders associated with MAPT gene expression.

Background Art

[0003] Microtubules play several important roles within cells throughout the body. In the central nervous system (CNS), microtubules provide structural support and help transport substances throughout the cell. Changes in the amount, structure, and pattern of microtubules are known to contribute to the onset of many neurodegenerative diseases. Tau is an essential protein for microtubule formation, and its abnormal expression causes neurodegenerative diseases. Tau protein binds to tubulin to form microtubules. Alternative splicing of MAPT generates different tau proteins used for microtubule polymerization. Mutations (e.g., insertions and mismatches) in MAPT that alter the function and expression of tau are known causes of several diseases and disorders that affect the CNS (e.g., Alzheimer's disease (AD), Parkinson's disease (PD), and tauopathies). Strategies targeting MAPT gene expression are needed to prevent such diseases and disorders.

[0004] The mammalian central nervous system (CNS) is a complex system of tissues that includes cells, fluids, and chemicals, which interact simultaneously to enable a diverse range of functions, including movement, motor instruction, cognition, speech, vision, and emotion. Unfortunately, various diseases and disorders of the CNS (e.g., neuropathy) are known, which affect or disrupt some or all of these functions. Typically, the treatment of CNS diseases and disorders is limited to small molecule drugs, antibodies, and / or adaptive or behavioral therapies. There is a continuing need to develop therapies for CNS diseases and disorders associated with inappropriate MAPT gene expression.

Summary of the Invention

[0005] To address this need, the present disclosure describes compositions and methods of treating diseases, disorders, and / or conditions associated with MAPT gene expression. The present disclosure is based, at least in part, on the discovery and development of double-stranded (ds) oligonucleotides, such as RNAi oligonucleotides, that effectively target and reduce MAPT gene expression in CNS tissue. Specifically, target sequences within MAPT mRNA were identified, and oligonucleotides that bind to these target sequences and inhibit MAPT mRNA expression were generated. As demonstrated herein, the oligonucleotides inhibit MAPT gene expression in CNS tissue of humans and non-human primates (NHPs). Furthermore, MAPT mRNA expression was reduced in CNS tissue associated with Alzheimer's disease (AD) or progressive supranuclear palsy (PSP) by both GalNAc-conjugated and lipid-conjugated MAPT mRNA-targeting oligonucleotides. Without being bound by theory, the oligonucleotides described herein are useful for treating diseases, disorders, or conditions associated with MAPT gene expression.

[0006] Accordingly, and in some embodiments, the present disclosure provides an RNAi oligonucleotide for reducing the expression of the MAPT gene, the oligonucleotide comprising a sense strand and an antisense strand, wherein the sense strand and the antisense strand form a double-stranded region, and the antisense strand comprises a region complementary to any one of the MAPT mRNA target sequences of SEQ ID NOs: 912 to 1295, the complementary region being at least about 15 consecutive nucleotides in length.

[0007] In any of the foregoing embodiments or related embodiments, in some embodiments, the sense strand is about 15 to about 50 nucleotides in length, and in some embodiments, the sense strand is 18 to 36 nucleotides in length. In some embodiments, the antisense strand is about 15 to about 30 nucleotides in length. In some embodiments, the antisense strand is 22 nucleotides in length, and the antisense strand and the sense strand form a double-stranded region that is at least about 19 nucleotides in length, optionally at least 20 nucleotides in length. In some embodiments, the complementary region is at least about 19 consecutive nucleotides in length. In some embodiments, the complementary region is at least about 20 consecutive nucleotides in length.

[0008] In other embodiments, the present disclosure provides a dsRNAi oligonucleotide for reducing the expression of the MAPT gene, the oligonucleotide comprising (i) an antisense strand about 19 to 30 nucleotides in length, the antisense strand comprising a nucleotide sequence comprising a region complementary to a MAPT mRNA target sequence, the complementary region being selected from SEQ ID NOs: 1296 to 1679, the antisense strand, and (ii) a sense strand about 19 to 50 nucleotides in length comprising a region complementary to the region of the antisense strand, wherein the antisense strand and the sense strand are separate strands that form an asymmetric double-stranded region having an overhang of 1 to 4 nucleotides at the 3' end of the antisense strand.

[0009] In some embodiments, the 3' end of the sense strand includes a stem-loop defined as S1-L-S2, where S1 is complementary to S2 and L forms a loop between S1 and S2 that is 3 to 5 nucleotides in length. In some embodiments, L is a tri-loop (triL) or a tetra-loop (tetraL). In some embodiments, L is a tetraL. In some embodiments, the tetraL includes the sequence 5'-GAAA-3'. In some embodiments, S1 and S2 are nucleotides of about 1 to about 10 in length and have the same length. In some embodiments, S1 and S2 are 1 nucleotide, 2 nucleotides, 3 nucleotides, 4 nucleotides, 5 nucleotides, 6 nucleotides, 7 nucleotides, 8 nucleotides, 9 nucleotides, or 10 nucleotides in length. In some embodiments, S1 and S2 are 6 nucleotides in length. In some embodiments, the stem-loop includes the sequence 5'-GCAGCCGAAAGGCUGC-3' (SEQ ID NO: 1680).

[0010] In other embodiments, the oligonucleotide includes blunt ends. In some embodiments, the blunt ends include the 3' end of the sense strand. In some embodiments, the sense strand is about 20 to 22 nucleotides. In some embodiments, the sense strand is 20 nucleotides.

[0011] In any of the foregoing embodiments or related embodiments, the antisense strand includes a 3' overhang sequence of one or more nucleotides in length. In some embodiments, the overhang includes purine nucleotides. In some embodiments, the 3' overhang is 2 nucleotides in length. In some embodiments, the 3' overhang is selected from AA, GG, AG, and GA. In some embodiments, the overhang is GG or AA. In some embodiments, the overhang is GG.

[0012] In any 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 a modification selected from 2'-aminoethyl (EA), 2'-fluoro (2'-F), 2'-O-methyl (2'-OMe), 2'-O-methoxyethyl (2'-MOE), and 2'-deoxy-2'-fluoro-β-arabinonucleic acid (2'-FANA). In some aspects, the modification is a 2'-modification selected from 2'-F and 2'-OMe. In some aspects, about 18% to about 23%, or 18%, 19%, 20%, 21%, 22%, or 23% of the nucleotides of the sense strand comprise a 2'-F modification. In other aspects, about 38% to about 43%, or 38%, 39%, 40%, 41%, 42%, or 43% of the nucleotides of the sense 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 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% to about 45%, or 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 at positions 1 to 20 from 5' to 3', and each of positions 3, 5, 8, 10, 12, 13, 15, and 17 comprises a 2'-F modification. In some aspects, the sense strand comprises 36 nucleotides at positions 1 to 36 from 5' to 3', and 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 at positions 1 to 22 from 5' to 3', and 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.

[0013] In any of the foregoing or related embodiments, the oligonucleotide comprises at least one modified internucleotide linkage. In some embodiments, at least one modified internucleotide linkage is a phosphorothioate linkage. In some embodiments, the antisense strand comprises phosphorothioate linkages (i) between the 1st and 2nd positions, and between the 2nd and 3rd positions, or (ii) between the 1st and 2nd positions, between the 2nd and 3rd positions, and between the 3rd and 4th positions, where the positions are numbered 1 to 4 in the 5' to 3' direction. In some embodiments, the antisense strand is 22 nucleotides in length and comprises phosphorothioate linkages between the 20th and 21st positions, and between the 21st and 22nd positions, where the positions are numbered 1 to 22 in the 5' to 3' direction. In some embodiments, the sense strand comprises a phosphorothioate linkage between the 1st and 2nd positions, where the positions are numbered 1 to 2 in the 5' to 3' direction. In some embodiments, the sense strand is 20 nucleotides in length and comprises phosphorothioate linkages between the 1st and 2nd positions, between the 18th and 19th positions, and between the 19th and 20th positions, where the positions are numbered 1 to 20 in the 5' to 3' direction.

[0014] In any of the foregoing or related embodiments, the 4'-carbon of the sugar of the 5'-nucleotide of the antisense strand comprises a phosphate analog. In some embodiments, the phosphate analog is phosphonate oxymethyl, phosphonate vinyl, or phosphonate malonyl, and optionally, in that case, the phosphate analog is a 4'-phosphate analog comprising phosphonate 4'-oxymethyl.

[0015] In any of the foregoing or related aspects, at least one nucleotide of the oligonucleotide is bound 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 bound to one or more nucleotides of the stem-loop. In some aspects, one or more targeting ligands are bound to one or more nucleotides of the loop. In some aspects, the loop comprises four nucleotides numbered 1 to 4 from 5' to 3', wherein the nucleotides at positions 2, 3, and 4 each comprise one or more targeting ligands, and the targeting ligands are the same 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 four nucleotides of L of the stem-loop are each bound to a monovalent GalNac moiety.

[0016] In other aspects, one or more targeting ligands are lipid moieties. In some aspects, the lipid moiety is bound 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 a C8-C30 hydrocarbon chain. In some aspects, the hydrocarbon chain is a C16 hydrocarbon chain. In some aspects, the C16 hydrocarbon chain is represented by:

Chemical formula

[0017] In any of the foregoing embodiments or related embodiments, the complementary region is completely complementary to the MAPT mRNA target sequence at nucleotides 2 to 8 of the antisense strand, where the nucleotide positions are numbered 5' to 3'. In some embodiments, the complementary region is completely complementary to the MAPT mRNA target sequence at nucleotides 2 to 11 of the antisense strand, where the nucleotide positions are numbered 5' to 3'.

[0018] In any of the foregoing embodiments or related embodiments, the sense strand comprises any one of the nucleotide sequences of SEQ ID NOs: 769 to 803 and 1681. In some embodiments, the antisense strand comprises any one of the nucleotide sequences of SEQ ID NOs: 804 to 838.

[0019] In some embodiments, the sense strand and the antisense strand are: a) SEQ ID NO: 769 and SEQ ID NO: 804, respectively; b) SEQ ID NO: 770 and SEQ ID NO: 805, respectively; c) SEQ ID NO: 771 and SEQ ID NO: 806, respectively; d) SEQ ID NO: 772 and SEQ ID NO: 807, respectively; e) SEQ ID NO: 773 and SEQ ID NO: 808, respectively; f) SEQ ID NO: 774 and SEQ ID NO: 809, respectively; g) SEQ ID NO: 775 and SEQ ID NO: 810, respectively; h) SEQ ID NO: 776 and SEQ ID NO: 811, respectively; i) SEQ ID NO: 777 and SEQ ID NO: 812, respectively; j) SEQ ID NO: 778 and SEQ ID NO: 813, respectively; k) SEQ ID NO: 779 and SEQ ID NO: 814, respectively; l) SEQ ID NO: 780 and SEQ ID NO: 815, respectively; m) SEQ ID NO: 781 and SEQ ID NO: 816, respectively; n) SEQ ID NO: 782 and SEQ ID NO: 817, respectively; o) SEQ ID NO: 783 and SEQ ID NO: 818, respectively; p) SEQ ID NO: 784 and SEQ ID NO: 819, respectively; q) SEQ ID NO: 785 and SEQ ID NO: 820, respectively; r) SEQ ID NOs: 786 and 821, respectively; s) SEQ ID NOs: 787 and 822, respectively; t) SEQ ID NOs: 788 and 823, respectively; u) SEQ ID NOs: 789 and 824, respectively; v) SEQ ID NOs: 790 and 825, respectively; w) SEQ ID NOs: 791 and 826, respectively; x) SEQ ID NOs: 792 and 827, respectively; y) SEQ ID NOs: 793 and 828, respectively; z) SEQ ID NOs: 794 and 829, respectively; aa) SEQ ID NOs: 795 and 830, respectively; bb) SEQ ID NOs: 796 and 831, respectively; cc) SEQ ID NOs: 797 and 832, respectively; dd) SEQ ID NOs: 798 and 833, respectively; ee) SEQ ID NOs: 799 and 834, respectively; ff) SEQ ID NOs: 800 and 835, respectively; gg) SEQ ID NOs: 801 and 836, respectively; hh) SEQ ID NOs: 802 and 837, respectively ii) SEQ ID NOs: 803 and 838, respectively; and jj) a nucleotide sequence selected from the group consisting of SEQ ID NOs: 1681 and 815, respectively.

[0020] In some embodiments, the sense and antisense strands are: a) SEQ ID NOs: 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; k) It comprises a nucleotide sequence selected from the group consisting of SEQ ID NOs: 1681 and 815, respectively.

[0021] In some embodiments, the sense strand and the antisense strand are: a) SEQ ID NOs: 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) It comprises a nucleotide sequence selected from the group consisting of SEQ ID NOs: 1681 and 815, respectively.

[0022] In some embodiments, 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 embodiments, 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 embodiments, 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 embodiments, 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 embodiments, 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 embodiments, 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 embodiments, 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.

[0023] In any of the foregoing embodiments or related embodiments, the antisense strand is 22 nucleotides in length. In some embodiments, the antisense strand comprises a nucleotide sequence selected from SEQ ID NOs: 806, 815, 816, 833, 834, and 838. In some embodiments, the sense strand is 36 nucleotides in length. In some embodiments, the sense strand comprises a nucleotide sequence selected from SEQ ID NOs: 1130, 1095, 1096, 1119, 1120, and 1124. In some embodiments, the sense strand comprises a nucleotide sequence selected from SEQ ID NOs: 771, 780, 781, 798, 799, and 803.

[0024] In any of the foregoing embodiments or related embodiments, the sense strand comprises any one nucleotide sequence of SEQ ID NOs: 839-873 and 1682. In some embodiments, the antisense strand comprises any one nucleotide sequence of SEQ ID NOs: 874-908.

[0025] In some embodiments, the sense strand and the antisense strand are: a) SEQ ID NO: 839 and SEQ ID NO: 874 respectively; b) SEQ ID NO: 840 and SEQ ID NO: 875 respectively; c) SEQ ID NO: 841 and SEQ ID NO: 876 respectively; d) SEQ ID NO: 842 and SEQ ID NO: 877 respectively; e) SEQ ID NO: 843 and SEQ ID NO: 878 respectively; f) SEQ ID NO: 844 and SEQ ID NO: 879 respectively; g) SEQ ID NO: 845 and SEQ ID NO: 880 respectively; h) SEQ ID NO: 846 and SEQ ID NO: 881 respectively; i) SEQ ID NO: 847 and SEQ ID NO: 882 respectively; j) SEQ ID NO: 848 and SEQ ID NO: 883 respectively; k) SEQ ID NO: 849 and SEQ ID NO: 884 respectively; l) SEQ ID NO: 850 and SEQ ID NO: 885 respectively; m) SEQ ID NO: 851 and SEQ ID NO: 886 respectively; n) SEQ ID NO: 852 and SEQ ID NO: 887 respectively; o) SEQ ID NO: 853 and SEQ ID NO: 888 respectively; p) SEQ ID NO: 854 and SEQ ID NO: 889 respectively; q) SEQ ID NOs: 855 and 890, respectively; r) SEQ ID NOs: 856 and 891, respectively; s) SEQ ID NOs: 857 and 892, respectively; t) SEQ ID NOs: 858 and 893, respectively; u) SEQ ID NOs: 859 and 894, respectively; v) SEQ ID NOs: 860 and 895, respectively; w) SEQ ID NOs: 861 and 896, respectively; x) SEQ ID NOs: 862 and 897, respectively; y) SEQ ID NOs: 863 and 898, respectively; z) SEQ ID NOs: 864 and 899, respectively; aa) SEQ ID NOs: 865 and 900, respectively; bb) SEQ ID NOs: 866 and 901, respectively; cc) SEQ ID NOs: 867 and 902, respectively; dd) SEQ ID NOs: 868 and 903, respectively; ee) SEQ ID NOs: 869 and 904, respectively; ff) SEQ ID NOs: 870 and 905, respectively; gg) SEQ ID NOs: 871 and 906, respectively; hh) SEQ ID NOs: 872 and 907, respectively; ii) SEQ ID NOs: 873 and 908, respectively; and jj) a nucleotide sequence selected from the group consisting of SEQ ID NOs: 1682 and 885, respectively.

[0026] In other embodiments, the sense and antisense strands are: a) SEQ ID NOs: 860 and 895, respectively; b) SEQ ID NOs: 865 and 900, respectively; c) SEQ ID NOs: 868 and 903, respectively; d) SEQ ID NOs: 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) It comprises a nucleotide sequence selected from the group consisting of SEQ ID NOs: 1682 and 885, respectively.

[0027] In certain embodiments, the sense and antisense strands are: a) SEQ ID NOs: 841 and 876, respectively; b) SEQ ID NOs: 850 and 885, respectively; c) SEQ ID NOs: 851 and 886, respectively; d) SEQ ID NOs: 868 and 903, respectively; e) SEQ ID NOs: 869 and 904, respectively; f) SEQ ID NOs: 873 and 908, respectively; and g) It comprises a nucleotide sequence selected from the group consisting of SEQ ID NOs: 1682 and 885, respectively.

[0028] In some embodiments, 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 embodiments, 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 embodiments, 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 embodiments, 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 embodiments, 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 embodiments, 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 embodiments, 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.

[0029] In some embodiments, the sense strand comprises the sequence 5’-[mAs][mG][fA][mG][fU][mG][mU][fG][mG][fA][ mA][fA][fA][mA][fA][mA][fA][mA][mG][mA][mG][mC][mA][mG][mC][mC][mG][ademA-GalNAc][ademA-GalNAc][ademAGalNAc][mG][mG][mC][mU][mG][mC]-3’ (SEQ ID NO: 841) and all modifications, the antisense strand comprises the sequence 5’-[MePhosphonate-4O-mUs][fCs][fU][fU][fU][mU][fU][mU][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 =

Chemical formula

[0030] In some embodiments, the sense strand comprises the sequence 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, The antisense strand contains the sequence 5’-[MePhosphonate-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, where mC, mA, mG, mU = 2’-OMe ribonucleoside; fA, fC, fG, fU = 2’-F ribonucleoside; s = phosphorothioate, and where ademA-GalNAc = [Chemical formula] is as follows.

[0031] In some embodiments, the sense strand contains the sequence 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, The antisense strand contains the sequence 5’-[MePhosphonate-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, where mC, mA, mG, mU = 2’-OMe ribonucleoside; fA, fC, fG, fU = 2’-F ribonucleoside; s = phosphorothioate, and where ademA-GalNAc = [Chemical formula] is as follows.

[0032] In some embodiments, the sense strand comprises the sequence 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][mC][mG][ademA-GalNAc][ademA-GalNAc][ademA-GalNAc][mG][mG][mC][mU][mG][mC]-3’ (SEQ ID NO: 868) and all modifications, the antisense strand comprises the sequence 5’-[MePhosphonate-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, wherein ademA-GalNAc = [Chemical formula] is.

[0033] In some embodiments, the sense strand comprises the sequence 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, The antisense strand contains the sequence 5’-[MePhosphonate-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, where mC, mA, mG, mU = 2’-OMe ribonucleoside; fA, fC, fG, fU = 2’-F ribonucleoside; s = phosphorothioate, and where ademA-GalNAc =

Chemical formula

[0034] In some embodiments, the sense strand contains the sequence 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][ademA-GalNAc][ademA-GalNAc][ademA-GalNAc][mG][mG][mC][mU][mG][mC]-3’ (SEQ ID NO: 873) and all modifications, The antisense strand contains the sequence 5’-[MePhosphonate-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, where mC, mA, mG, mU = 2’-OMe ribonucleoside; fA, fC, fG, fU = 2’-F ribonucleoside; s = phosphorothioate, and where ademA-GalNAc =

Chemical formula

[0035] In some embodiments, the sense strand comprises the sequence 5’-[ademCs-C16][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, the antisense strand comprises the sequence 5’-[MePhosphonate-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 [ademCs-C16] = cytosine linked to a C16 hydrocarbon chain. [Chemical formula]

[0036] In some embodiments, the present disclosure provides a pharmaceutical composition comprising an RNAi oligonucleotide described herein and a pharmaceutically acceptable carrier, delivery agent, or excipient.

[0037] In other embodiments, the present disclosure provides a method of treating a subject having a disease, disorder, or condition associated with the expression of the MAPT gene, 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.

[0038] In a further embodiment, 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.

[0039] In yet a further embodiment, the present disclosure provides a method of reducing the expression of the MAPT gene in a cell, a population of cells, or a subject, the method comprising: i. contacting a cell or population of cells with an RNAi oligonucleotide or pharmaceutical composition described herein; or ii. administering an RNAi oligonucleotide or pharmaceutical composition described herein to a subject.

[0040] In some embodiments, reducing MAPT gene expression comprises reducing the amount or level of MAPT mRNA, the amount or level of tau protein, or both. In some embodiments, the RNAi oligonucleotide or pharmaceutical composition is as described herein and the subject has a disease, disorder, or condition associated with the expression of the MAPT gene. In some embodiments, the disease, disorder, or condition associated with MAPT gene expression is AD, frontotemporal dementia (FTD), PSP, PD, tau protein-related diseases, primary age-related tauopathy, chronic traumatic encephalopathy, corticobasal degeneration, corticobasal body disease, glioma, meningovascular angiomatosis, postencephalitic parkinsonism, or subacute sclerosing panencephalitis.

[0041] In any of the foregoing or related embodiments, the expression of the MAPT gene is reduced in the tissue of one or more regions of the CNS, wherein the tissue is associated with AD. In some embodiments, the tissue associated with AD is selected from the prefrontal cortex, motor cortex, temporal cortex, parietal cortex, and hippocampus. In some embodiments, the expression of the MAPT gene is reduced in the tissue of one or more regions of the CNS, wherein the tissue is associated with PSP. In some embodiments, the tissue associated with PSPy is selected from the caudate nucleus, putamen, thalamus, midbrain tegmentum, substantia nigra, pons, cerebellar white matter, dentate nucleus of the cerebellum, medulla oblongata, cervical spinal cord, thoracic spinal cord, and lumbar spinal cord. In some embodiments, the expression of the MAPT gene is reduced in one or more regions of the CNS selected from the cervical spinal cord, thoracic spinal cord, lumbar spinal cord, prefrontal cortex, temporal cortex, cerebellum, midbrain, occipital cortex, parietal cortex, hippocampus, caudate nucleus, thalamus, brainstem, motor cortex, putamen, midbrain tegmentum, substantia nigra, pons, cerebellar white matter, and dentate nucleus of the cerebellum.

[0042] In any of the foregoing or related embodiments, the RNAi oligonucleotide or pharmaceutical composition is administered in combination with a second composition or therapeutic agent.

[0043] In other aspects, the present disclosure provides the use of the RNAi oligonucleotides or pharmaceutical compositions described herein in the manufacture of a medicament for the treatment of a disease, disorder, or condition associated with the expression of the MAPT gene.

[0044] In a further aspect, the present disclosure provides the RNAi oligonucleotides or pharmaceutical compositions described herein for use in the treatment of a disease, disorder, or condition associated with the expression of the MAPT gene, or adaptable for use.

[0045] In some aspects, the present disclosure provides a kit comprising the RNAi oligonucleotides described herein, any pharmaceutically acceptable carrier, and a package insert containing instructions for administration to a subject having a disease, disorder, or condition associated with the expression of the MAPT gene.

[0046] In any of the foregoing aspects or related aspects, the disease, disorder, or condition associated with the expression of the MAPT gene is AD, FTD, PD, PSP, tau protein-related diseases, primary age-related tauopathy, chronic traumatic encephalopathy, corticobasal degeneration, corticobasal body disease, glioblastoma, meningovascular angiomatosis, postencephalitic parkinsonism, or subacute sclerosing panencephalitis.

[0047] Other advantages, effects, features, and objectives will become more readily apparent upon consideration of the following detailed description. Such detailed description refers to the following drawings (s). BRIEF DESCRIPTION OF THE DRAWINGS

[0048]

Figure 1A

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Figure 2A

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Figure 3A

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Figure 3C

Figure 3D

Figure 3E

Figure 3F

Figure 3G

Figure 3H

Figure 3I

Figure 3J

Figure 3K

Figure 3L

Figure 3M

Figure 4A

Figure 4B

Figure 5A

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Figure 6B

Mode for Carrying Out the Invention

[0049] According to some aspects, the present disclosure provides oligonucleotides such as RNAi oligonucleotides that reduce the expression of the MAPT gene in the CNS. In some embodiments, the oligonucleotides provided herein are designed to treat diseases associated with the expression of the MAPT gene in the CNS. In some respects, the present disclosure provides a method of treating MAPT-related diseases by reducing the expression of the MAPT gene in cells (e.g., cells of the CNS).

[0050] Oligonucleotide inhibitors of MAPT gene expression

[0051] The present disclosure provides, inter alia, oligonucleotides (e.g., RNAi oligonucleotides) that inhibit the expression of the MAPT gene. In some embodiments, the oligonucleotides that inhibit the expression of the MAPT gene target MAPT mRNA.

[0052] MAPT target sequence

[0053] In some embodiments, the oligonucleotides herein (e.g., RNAi oligonucleotides) target target sequences that make up MAPT mRNA. In some embodiments, the oligonucleotide targets a target sequence within the MAPT mRNA sequence.

[0054] 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., the antisense strand or guide strand of an RNAi oligonucleotide), binds or anneals to a target sequence that makes up MAPT mRNA, thereby inhibiting the expression of the MAPT gene.

[0055] In some embodiments, the oligonucleotide targets the MAPT target sequence for the purpose of inhibiting the expression of the MAPT gene in vivo. In some embodiments, the amount or degree of inhibition of MAPT gene expression by the oligonucleotide targeting the MAPT target sequence correlates with the potency of the oligonucleotide. In some embodiments, the amount or degree of inhibition of MAPT gene expression by the oligonucleotide targeting the MAPT target sequence correlates with the amount or degree of therapeutic effect in a subject or patient having a disease, disorder, or condition associated with MAPT gene expression treated with the oligonucleotide.

[0056] In some embodiments, the sense strand of the oligonucleotide comprises a MAPT target sequence. In some embodiments, a portion or region of the sense strand of an oligonucleotide (e.g., an RNAi oligonucleotide) comprises a MAPT target sequence. In some embodiments, the MAPT target sequence comprises, or consists of, any one nucleotide sequence of SEQ ID NOs: 912 to 1295. In some embodiments, the MAPT target sequence comprises, or consists of, any one nucleotide sequence 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, any one nucleotide sequence 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, any one nucleotide sequence of SEQ ID NOs: 1130, 1095, 1096, 1119, 1120, and 1124. In some embodiments, the MAPT target sequence comprises the nucleotide sequence represented by SEQ ID NO: 1130. In some embodiments, the MAPT target sequence comprises the nucleotide sequence represented by SEQ ID NO: 1095. In some embodiments, the MAPT target sequence comprises the nucleotide sequence represented by SEQ ID NO: 1096. In some embodiments, the MAPT target sequence comprises the nucleotide sequence represented by SEQ ID NO: 1119. In some embodiments, the MAPT target sequence comprises the nucleotide sequence represented by SEQ ID NO: 1120. In some embodiments, the MAPT target sequence comprises the nucleotide sequence represented by SEQ ID NO: 1124.

[0057] Sequence targeting MAPT mRNA

[0058] In some embodiments, the oligonucleotides herein (e.g., RNAi oligonucleotides) have regions complementary to MAPT mRNA (e.g., within the target sequence of MAPT mRNA) for the purpose of targeting mRNA in cells and inhibiting its expression. In some embodiments, the oligonucleotide has a MAPT mRNA target sequence (e.g., the antisense strand or guide strand of a ds oligonucleotide such as an RNAi oligonucleotide) having a complementary region that binds or anneals to the MAPT target sequence by complementary (Watson-Crick) base pairing. The complementary targeting sequence or region is generally of a length and base content suitable to enable binding or annealing of the oligonucleotide (or its strand) to MAPT mRNA for the purpose of inhibiting MAPT mRNA expression. In some embodiments, the complementary targeting sequence or 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 length. In some embodiments, the complementary targeting sequence or 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, at least 20 nucleotides in length. In some embodiments, the complementary targeting sequence or region is about 12 - 30 (e.g., 12 - 30, 12 - 22, 15 - 25, 17 - 21, 18 - 27, 19 - 27, or 15 - 30) nucleotides in length. In some embodiments, the complementary targeting sequence or 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 length. In some embodiments, the complementary targeting sequence or region is 18 nucleotides in length. In some embodiments, the complementary targeting sequence or region is 19 nucleotides in length.In some embodiments, the complementary targeting sequence or region is 20 nucleotides in length. In some embodiments, the complementary targeting sequence or region is 21 nucleotides in length. In some embodiments, the complementary targeting sequence or region is 22 nucleotides in length. In some embodiments, the complementary targeting sequence or region is 23 nucleotides in length. In some embodiments, the complementary targeting sequence or region is 24 nucleotides in length. In some embodiments, the oligonucleotide comprises a complementary targeting sequence or region that is complementary to any one of the sequences of SEQ ID NOs: 912 to 1295, and the complementary targeting sequence or region is 18 nucleotides in length. In some embodiments, the oligonucleotide comprises a complementary targeting sequence or region that is complementary to any one of the sequences of SEQ ID NOs: 912 to 1295, and the complementary targeting sequence or region is 19 nucleotides in length. In some embodiments, the oligonucleotide comprises a complementary targeting sequence or region that is complementary to any one of the sequences of SEQ ID NOs: 1 to 384, and the complementary targeting sequence or region is 20 nucleotides in length. In some embodiments, the oligonucleotide comprises a complementary targeting sequence or region that is complementary to any one of the sequences of SEQ ID NOs: 1 to 384, and the complementary targeting sequence or region is 21 nucleotides in length. In some embodiments, the oligonucleotide comprises a complementary targeting sequence or region that is complementary to any one of the sequences of SEQ ID NOs: 1 to 384, and the complementary targeting sequence or region is 22 nucleotides in length. In some embodiments, the oligonucleotide comprises a complementary targeting sequence or region that is complementary to any one of the sequences of SEQ ID NOs: 1 to 384, and the complementary targeting sequence or region is 23 nucleotides in length. In some embodiments, the oligonucleotide comprises a complementary targeting sequence or region that is complementary to any one of the sequences of SEQ ID NOs: 1 to 384, and the complementary targeting sequence or region is 24 nucleotides in length.

[0059] In some embodiments, the oligonucleotide comprises a complementary targeting sequence or region (e.g., the antisense strand or guide strand of a ds oligonucleotide) that is completely complementary to the MAPT mRNA target sequence. In some embodiments, the complementary targeting sequence or region is partially complementary to the MAPT mRNA target sequence. In some embodiments, the oligonucleotide comprises a complementary targeting sequence or region that is completely complementary to any one of the sequences of SEQ ID NOs: 912 to 1295. In some embodiments, the oligonucleotide comprises a complementary targeting sequence or region that is partially complementary to any one of the sequences of SEQ ID NOs: 912 to 1295. In some embodiments, the oligonucleotide comprises a complementary targeting sequence or region that is completely complementary to any one of the sequences 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 comprises a complementary targeting sequence or region that is partially complementary to any one of the sequences 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 comprises a complementary targeting sequence or region that is completely complementary to any one of the sequences of SEQ ID NOs: 1061, 1108, 1119, 1120, 1124, 1130, 1065, 1095, 1096, or 1102.In some embodiments, the oligonucleotide comprises a complementary targeting sequence or region that is partially complementary to any one of the sequences of SEQ ID NO: 1061, 1108, 1119, 1120, 1124, 1130, 1065, 1095, 1096, or 1102. In some embodiments, the oligonucleotide comprises a complementary targeting sequence or region that is completely complementary to any one of the sequences of SEQ ID NO: 1130, 1095, 1096, 1119, 1120, and 1124. In some embodiments, the oligonucleotide comprises a complementary targeting sequence or region that is partially complementary to any one of the sequences of SEQ ID NO: 1130, 1095, 1096, 1119, 1120, and 1124. In some embodiments, the oligonucleotide comprises a complementary targeting sequence or region that is completely complementary to the sequence shown in SEQ ID NO: 1130. In some embodiments, the oligonucleotide comprises a complementary targeting sequence or region that is completely complementary to the sequence shown in SEQ ID NO: 1095. In some embodiments, the oligonucleotide comprises a complementary targeting sequence or region that is completely complementary to the sequence shown in SEQ ID NO: 1096. In some embodiments, the oligonucleotide comprises a complementary targeting sequence or region that is completely complementary to the sequence shown in SEQ ID NO: 1119. In some embodiments, the oligonucleotide comprises a complementary targeting sequence or region that is completely complementary to the sequence shown in SEQ ID NO: 1120. In some embodiments, the oligonucleotide comprises a complementary targeting sequence or region that is completely complementary to the sequence shown in SEQ ID NO: 1124. In some embodiments, the oligonucleotide comprises a complementary targeting sequence or region that is partially complementary to any one of the sequences of SEQ ID NO: 1130. In some embodiments, the oligonucleotide comprises a complementary targeting sequence or region that is partially complementary to the sequence of SEQ ID NO: 1095. In some embodiments, the oligonucleotide comprises a complementary targeting sequence or region that is partially complementary to the sequence of SEQ ID NO: 1096. In some embodiments, the oligonucleotide comprises a complementary targeting sequence or region that is partially complementary to the sequence of SEQ ID NO: 1119.In some embodiments, the oligonucleotide comprises a complementary targeting sequence or region that is partially complementary to the sequence of SEQ ID NO: 1120. In some embodiments, the oligonucleotide comprises a complementary targeting sequence or region that is partially complementary to the sequence of SEQ ID NO: 1124.

[0060] In some embodiments, the oligonucleotide comprises a complementary targeting sequence or region that is complementary to a contiguous sequence of nucleotides comprising MAPT mRNA, wherein the contiguous sequence of nucleotides is about 12 to about 30 nucleotides in length (e.g., 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, the oligonucleotide comprises a complementary targeting sequence or region that is complementary to a contiguous sequence of nucleotides comprising MAPT mRNA, wherein the contiguous sequence of nucleotides is 10, 11, 12, 13, 14, 15, 16, 17, 18, 19, or 20 nucleotides in length. In some embodiments, the oligonucleotide comprises a complementary targeting sequence or region that is complementary to a contiguous sequence of nucleotides comprising MAPT mRNA, wherein the contiguous sequence of nucleotides is 19 nucleotides in length. In some embodiments, the oligonucleotide comprises a complementary targeting sequence or region that is complementary to a contiguous sequence of nucleotides comprising MAPT mRNA, wherein the contiguous sequence of nucleotides is 20 nucleotides in length.

[0061] In some embodiments, the oligonucleotide comprises a complementary targeting sequence or region that is complementary to a contiguous sequence of any one of nucleotides of SEQ ID NOs: 912 to 1295, and optionally, the contiguous sequence of nucleotides is 19 nucleotides in length. In some embodiments, the oligonucleotide comprises a complementary targeting sequence or region that is complementary to a contiguous sequence of any one of nucleotides 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, and optionally the contiguous sequence of nucleotides is 19 nucleotides in length. In other embodiments, the oligonucleotide comprises a complementary targeting sequence or region that is complementary to a contiguous sequence of any one of nucleotides of SEQ ID NOs: 1061, 1108, 1119, 1120, 1124, 1130, 1065, 1095, 1096, or 1102, and optionally, the contiguous sequence of nucleotides is 19 nucleotides in length. In other embodiments, the oligonucleotide comprises a complementary targeting sequence or region that is complementary to a contiguous sequence of nucleotides of SEQ ID NOs: 1130, 1095, 1096, 1119, 1120, and 1124, and optionally, the contiguous sequence of nucleotides is 19 nucleotides in length.

[0062] In some embodiments, the targeted sequence or region of complementarity of the oligonucleotide is complementary to consecutive nucleotides of the sequence shown in any one of SEQ ID NOs: 912 to 1295 and spans the entire length of the antisense strand. In some embodiments, the targeted sequence or region of complementarity of the oligonucleotide is complementary to consecutive nucleotides of the sequence shown in any one of SEQ ID NOs: 912 to 1295 and spans a portion of the entire length of the antisense strand. In some embodiments, the oligonucleotide comprises (e.g., on the antisense strand of a ds oligonucleotide) a region of complementarity that is at least partially (e.g., fully) complementary to a consecutive stretch of nucleotides spanning nucleotides 1 to 20 of the sequence shown in any one of SEQ ID NOs: 912 to 1295. In some embodiments, the targeted sequence or region of complementarity of the oligonucleotide is complementary to consecutive nucleotides of the sequence shown in any one of SEQ ID NOs: 1 to 384 and spans the entire length of the antisense strand. In some embodiments, the complementary region of the oligonucleotide is complementary to consecutive nucleotides of the sequence shown in any one of SEQ ID NOs: 1 to 384 and spans a portion of the entire length of the antisense strand. In some embodiments, the oligonucleotide comprises (e.g., on the antisense strand of a ds oligonucleotide) a region of complementarity that is at least partially (e.g., fully) complementary to a consecutive stretch of nucleotides spanning nucleotides 1 to 19 of the sequence shown in any one of SEQ ID NOs: 1 to 384.

[0063] In some embodiments, the oligonucleotide comprises a complementary targeting sequence or region having one or more base pair (bp) mismatches with the corresponding MAPT mRNA target sequence. In some embodiments, the complementary targeting sequence or region may have mismatches with the corresponding MAPT mRNA target sequence of up to about 1, up to about 2, up to about 3, up to about 4, up to about 5, etc., provided that the ability of the complementary targeting sequence or region to bind or anneal to MAPT mRNA under appropriate hybridization conditions and / or the ability of the oligonucleotide to inhibit the expression of the MAPT gene is maintained. Alternatively, in some embodiments, the complementary targeting sequence or region comprises up to 1, up to 2, up to 3, up to 4, or up to 5 mismatches with the corresponding MAPT mRNA target sequence, provided that the ability of the complementary targeting sequence or region to bind or anneal to MAPT mRNA under appropriate hybridization conditions and / or the ability of the oligonucleotide to inhibit the expression of the MAPT gene is maintained. In some embodiments, the oligonucleotide comprises a complementary targeting sequence or region having one mismatch with the corresponding target sequence. In some embodiments, the oligonucleotide comprises a complementary targeting sequence or region having two mismatches with the corresponding target sequence. In some embodiments, the oligonucleotide comprises a complementary targeting sequence or region having three mismatches with the corresponding target sequence. In some embodiments, the oligonucleotide comprises a complementary targeting sequence or region having four mismatches with the corresponding target sequence. In some embodiments, the oligonucleotide comprises a complementary targeting sequence or region having five mismatches with the corresponding target sequence. In some embodiments, the oligonucleotide comprises a complementary targeting sequence or region having more than one mismatch (e.g., 2, 3, 4, 5 or more mismatches) with the corresponding target sequence, wherein at least two (e.g., all) of the mismatches are arranged consecutively (e.g., 2, 3, 4, 5 or more consecutive mismatches), or the mismatches are scattered at any position throughout the complementary targeting sequence or region.In some embodiments, the oligonucleotide comprises a complementary targeting sequence or region having more than one mismatch (e.g., 2, 3, 4, 5 or more mismatches) with the corresponding target sequence, wherein at least two (e.g., all) of the mismatches are arranged contiguously (e.g., 2, 3, 4, 5 or more contiguous mismatches), or at least one non-mismatched base pair is located between the mismatches, or a combination thereof.

[0064] In some embodiments, the oligonucleotide comprises a complementary targeting sequence or region that is complementary to a contiguous sequence of any one nucleotide of SEQ ID NOs: 912 to 1295, wherein the complementary targeting sequence or region may 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. In some embodiments, the oligonucleotide comprises a complementary targeting sequence or region that is complementary to a contiguous sequence of any one nucleotide of SEQ ID NOs: 912 to 1295, wherein the complementary targeting sequence or region may have 1 or less, 2 or less, 3 or less, about 4 or less, about 5 or less mismatches with the corresponding MAPT mRNA target sequence. In some embodiments, the oligonucleotide comprises a complementary targeting sequence or region that is complementary to a contiguous sequence of any one nucleotide 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 complementary targeting sequence or region may 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. In some embodiments, the oligonucleotide comprises a complementary targeting sequence or region that is complementary to a contiguous sequence of any one nucleotide 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 complementary targeting sequence or region may have 1 or less, 2 or less, 3 or less, 4 or less, 5 or less mismatches with the corresponding MAPT mRNA target sequence.In some embodiments, the oligonucleotide comprises a complementary targeting sequence or region that is complementary to a contiguous sequence of any one of the nucleotides of SEQ ID NO: 1061, 1108, 1119, 1120, 1124, 1130, 1065, 1095, 1096, or 1102, wherein the complementary targeting sequence or region may 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. In some embodiments, the oligonucleotide comprises a complementary targeting sequence or region that is complementary to a contiguous sequence of any one of the nucleotides of SEQ ID NO: 1061, 1108, 1119, 1120, 1124, 1130, 1065, 1095, 1096, or 1102, wherein the complementary targeting sequence or region may have 1 or fewer, 2 or fewer, 3 or fewer, 4 or fewer, 5 or fewer mismatches with the corresponding MAPT mRNA target sequence. In some embodiments, the oligonucleotide comprises a complementary targeting sequence or region that is complementary to a contiguous sequence of any one of the nucleotides of SEQ ID NO: 1130, 1095, 1096, 1119, 1120, and 1124, wherein the complementary targeting sequence or region may 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. In some embodiments, the oligonucleotide comprises a complementary targeting sequence or region that is complementary to a contiguous sequence of any one of the nucleotides of SEQ ID NO: 1130, 1095, 1096, 1119, 1120, and 1124, wherein the complementary targeting sequence or region may have 1 or fewer, 2 or fewer, 3 or fewer, 4 or fewer, 5 or fewer mismatches with the corresponding MAPT mRNA target sequence. In some embodiments, the oligonucleotide comprises a complementary targeting sequence or region that is complementary to a contiguous sequence of the nucleotides of SEQ ID NO: 1095, wherein the complementary targeting sequence or region may 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.In some embodiments, the oligonucleotide comprises a complementary targeting sequence or region that is complementary to a contiguous sequence of nucleotides of SEQ ID NO: 1095, wherein the complementary targeting sequence or region may have 1 or fewer, 2 or fewer, 3 or fewer, 4 or fewer, 5 or fewer mismatches with the corresponding MAPT mRNA target sequence.

[0065] Types of oligonucleotides

[0066] Various types and / or structures of oligonucleotides, including but not limited to RNAi oligonucleotides, are useful for targeting MAPT mRNA by the methods herein. Any of the types of oligonucleotides described herein or elsewhere are contemplated for use as a framework for incorporating MAPT mRNA targeting sequences herein for the purpose of inhibiting the expression of the MAPT gene.

[0067] In some embodiments, the oligonucleotides herein inhibit the expression of the MAPT gene (e.g., RNAi oligonucleotides) by participating in the RNA interference (RNAi) pathway either upstream or downstream of Dicer involvement. For example, RNAi oligonucleotides have been developed in which each strand has a size of about 19-25 nucleotides and has at least one 3' overhang of about 1-about 5 nucleotides (see, e.g., U.S. Patent No. 8,372,968). Longer oligonucleotides have also been developed that are processed by Dicer to yield active RNAi products (see, e.g., U.S. Patent No. 8,883,996). Further studies have produced extended ds oligonucleotides in which at least one end of at least one strand extends beyond the double-stranded targeting region, which includes structures in which one of the strands contains a thermodynamically stable tetra-L structure (see, e.g., U.S. Patent Nos. 8,513,207 and 8,927,705, and International Patent Application Publication No. WO2010 / 033225). Such structures may include ds extensions as well as single-stranded extensions (ss) (on one or both sides of the molecule).

[0068] In some embodiments, the oligonucleotide is involved in the RNAi pathway downstream of Dicer involvement (e.g., Dicer cleavage). In some embodiments, the oligonucleotide has an overhang (e.g., 1, 2, or 3 nucleotides in length) at the 3' end of the sense strand. In some embodiments, the oligonucleotide comprises a 21-nucleotide antisense strand that is antisense to a target mRNA (e.g., MAPT mRNA), and a complementary sense strand that anneals to both strands to form a 19-bp double strand and forms a 2-nucleotide overhang at the 3' end of either or both. Also contemplated is the design of longer oligonucleotides that include an antisense strand 23 nucleotides in length and a sense strand 21 nucleotides in length, with a blunt end on the right side of the oligonucleotide (3' end of the sense strand / 5' end of the antisense strand) and a 2-nucleotide 3' guide strand overhang on the left side of the oligonucleotide (5' end of the sense strand / 3' end of the antisense strand). Such molecules have a 21-bp double-stranded region. See, for example, U.S. Patent Nos. 9,012,138; 9,012,621, and 9,193,753.

[0069] In some embodiments, the oligonucleotide comprises a sense strand and an antisense strand, both having a length in the range of about 17 to about 36 (e.g., 17 - 26, 20 - 25, or 21 - 23) nucleotides. In some embodiments, the oligonucleotide comprises an antisense strand having a length of 19 - 30 nucleotides and a sense strand having a length of 19 - 50 nucleotides, wherein the antisense strand and the sense strand are separate strands that form an asymmetric double-stranded region having 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 having a length in the range of about 19 to about 22 nucleotides. In some embodiments, the sense strand and the antisense strand are of equal length. In some embodiments, since the oligonucleotide comprises a sense strand and an antisense strand, there is a 3' overhang on either the sense strand or the antisense strand, or both the sense strand and the antisense strand. In some embodiments, for an oligonucleotide having a sense strand and an antisense strand, both having a length in the range of about 21 - 23 nucleotides, the 3' overhang of the sense strand, the antisense strand, or both the sense strand and the antisense strand is 1 or 2 nucleotides in length. In some embodiments, the oligonucleotide has a 22-nucleotide guide strand and a 20-nucleotide passenger strand, with a blunt end (3' end of the passenger strand / 5' end of the guide strand) on the right side of the molecule and a 2-nucleotide 3' guide strand overhang (5' end of the passenger strand / 3' end of the guide strand) on the left side of the molecule. Such a molecule has a 20-bp double-stranded region.

[0070] Other oligonucleotide designs for use with the compositions and methods herein include 16-base siRNAs (see, e.g., “Nucleic Acids in Chemistry & Biology,”, Blackburn (ed.), Royal Society of Chemistry, 2006), shRNAs (e.g., those having a stem of 19 bp or shorter; see, e.g., Moore et al. (2010) Methods Mol. Biol. 629:141-58), blunt siRNAs (e.g., those of 19 bp length; see, e.g., Kraynack & Baker (2006) RNA, 12:163-76), asymmetric siRNAs (aiRNAs; see, e.g., Sun et al. (2008) Nat. Biotechnol. 26:1379-82), asymmetric short double-stranded siRNAs (e.g., Chang et al. (2009) Mol. Ther. 17:725-32), fork siRNAs (see, e.g., Hohjoh (2004) FEBS Lett. 557:193-98), single-stranded siRNAs (see, e.g., Elsner (2012), Nat. Biotechnol. 30:1063), dumbbell-shaped circular siRNAs (see, e.g., Abe et al. (2007) J. Am. Chem. Soc. 129:15108-09), and small internally segmented interfering RNAs (siRNAs; see, e.g., Bramsen et al. (2007), Nucleic Acids Res. 35:5886-97). Further non-limiting examples of oligonucleotide structures that may be used in some embodiments to reduce or inhibit the expression of MAPT are microRNAs (miRNAs), short hairpin RNAs (shRNAs), and short siRNAs (see, e.g., Hamilton et al. (2002) EMBO J. 21:4671-79; also see U.S. Patent Application Publication No. 2009 / 0099115).

[0071] However, in some embodiments, oligonucleotides for reducing or inhibiting the expression of the MAPT gene herein include, but are not limited to, such structures that are ss, such as ssRNAi molecules. The activity of ssRNAi molecules has been demonstrated by recent efforts (see, e.g., Matsui et al. (2016) Mol. Ther. 24:946-55). However, in some embodiments, the oligonucleotide is an antisense oligonucleotide (ASO). An ASO contains a complement of the targeted segment of a particular nucleic acid in the reverse direction when written or drawn in the 5' to 3' direction and is suitably modified to induce RNaseH-mediated cleavage of its target RNA in the cell or to inhibit translation of the target mRNA in the cell (e.g., as a mixmer or, e.g., as a gapmer) and is a ss oligonucleotide having a nucleobase sequence. ASOs for use herein may be modified in any suitable manner known in the art, including, for example, those shown in U.S. Patent No. 9,567,587 (including, e.g., variations in the length of the nucleobases (pyrimidines, purines), the sugar moiety, and the heterocyclic moiety of the nucleobases). Further, 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).

[0072] In some embodiments, the ASO shares a region complementary to the MAPT mRNA. In some embodiments, the ASO targets various regions of human MAPT identified as NM_001123066.3. In some embodiments, the ASO is about 15 to 50 nucleotides in length. In some embodiments, the ASO is 15 to 25 nucleotides in length. In some embodiments, the ASO is 22 nucleotides in length. In some embodiments, the ASO is complementary to any one of SEQ ID NOs: 912 to 1295. In some embodiments, the ASO is at least 15 consecutive nucleotides in length. In some embodiments, the ASO is at least 19 consecutive nucleotides in length. In some embodiments, the ASO is at least 20 consecutive nucleotides in length. In some embodiments, the ASO differs from the target sequence by only 1, 2, or 3 nucleotides.

[0073] Double-stranded RNAi oligonucleotide

[0074] In some aspects, the present disclosure provides a dsRNAi oligonucleotide for targeting MAPT mRNA and inhibiting the expression of the MAPT gene (e.g., via the RNAi pathway), which includes 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 separate 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 double-stranded region, wherein the sense strand and the antisense strand, or a portion thereof, bind to each other in a complementary manner (e.g., by Watson-Crick base pairing).

[0075] In some embodiments, the sense strand comprises a first region (R1) and a second region (R2), where R2 comprises a first sub-region (S1), L (e.g., tetra L or tri L), and a second sub-region (S2), and L is located between S1 and S2, and S1 and S2 form a second double strand (D2). D2 may have various lengths. In some embodiments, D2 is about 1 to about 6 bp in length. In other embodiments, D2 is 2 - 6, 3 - 6, 4 - 6, 5 - 6, 1 - 5, 2 - 5, 3 - 5, or 4 - 5 bp in length. In other embodiments, D2 is 1, 2, 3, 4, 5, or 6 bP in length. In some embodiments, D2 is 6 bp in length.

[0076] In some embodiments, R1 of the sense strand and the antisense strand form a first double strand (D1). In some embodiments, D1 is at least 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 length. In some embodiments, D1 is about 12 to about 30 nucleotides in length (e.g., 12 - 30, 12 - 27, 15 - 22, 18 - 22, 18 - 25, 18 - 27, 18 - 30, or 21 - 30 nucleotides in length). In some embodiments, D1 is at least 12 nucleotides in length (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 formed by the sense strand and the antisense strand does not cover the entire length of the sense strand and / or the antisense strand. In some embodiments, D1 formed by the sense strand and the antisense strand covers the entire length of either or both of the sense strand and the antisense strand. In some embodiments, D1 formed by the sense strand and the antisense strand covers the entire length of both the sense strand and the antisense strand.

[0077] In some embodiments, the sense strand described herein is 36 nucleotides in length and is numbered 1 to 36 from 5' to 3' in position. In some embodiments, the antisense strand described herein is 22 nucleotides in length and is numbered 1 to 22 from 5' to 3' in position. In some embodiments, the position numbers described herein follow this numbering format.

[0078] In some embodiments, the RNAi oligonucleotide comprises a sense strand having any one of the sequences of SEQ ID NOs: 1 to 384 and an antisense strand comprising any one of the complementary sequences of SEQ ID NOs: 385 to 768. In some embodiments, the RNAi oligonucleotide comprises a sense strand having any one of the sequences of SEQ ID NOs: 912 to 1295 and an antisense strand comprising any one of the complementary sequences of SEQ ID NOs: 1296 to 1679.

[0079] In some embodiments, the RNAi oligonucleotide comprises a sense strand having any one of the sequences of SEQ ID NOs: 769 to 803 and an antisense strand comprising any one of the complementary sequences of SEQ ID NOs: 804 to 838. In some embodiments, the RNAi oligonucleotide comprises a sense strand having any one of the sequences of SEQ ID NOs: 769 to 803 and 1681 and an antisense strand comprising any one of the complementary sequences of SEQ ID NOs: 804 to 838.

[0080] In some embodiments, the RNAi oligonucleotide is: a) SEQ ID NOs: 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; l) SEQ ID NOs: 780 and 815, respectively; m) SEQ ID NOs: 781 and 816, respectively; n) SEQ ID NOs: 782 and 817, respectively; o) SEQ ID NOs: 783 and 818, respectively; p) SEQ ID NOs: 784 and 819, respectively; q) SEQ ID NOs: 785 and 820, respectively; r) SEQ ID NOs: 786 and 821, respectively; s) SEQ ID NOs: 787 and 822, respectively; t) SEQ ID NOs: 788 and 823, respectively; u) SEQ ID NOs: 789 and 824, respectively; v) SEQ ID NOs: 790 and 825, respectively; w) SEQ ID NOs: 791 and 826, respectively; x) SEQ ID NOs: 792 and 827, respectively; y) SEQ ID NOs: 793 and 828, respectively; z) SEQ ID NOs: 794 and 829, respectively; aa) SEQ ID NOs: 795 and 830, respectively; bb) SEQ ID NOs: 796 and 831, respectively; cc) SEQ ID NOs: 797 and 832, respectively; dd) SEQ ID NOs: 798 and 833, respectively; ee) SEQ ID NOs: 799 and 834, respectively; ff) SEQ ID NOs: 800 and 835, respectively; gg) SEQ ID NOs: 801 and 836, respectively; hh) SEQ ID NOs: 802 and 837, respectively ii) SEQ ID NOs: 803 and 838, respectively; and jj) comprising a sense strand and an antisense strand comprising nucleotide sequences selected from SEQ ID NOs: 1681 and 815, respectively.

[0081] In some embodiments, the RNAi oligonucleotides are: a) SEQ ID NOs: 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) comprising a sense strand and an antisense strand comprising nucleotide sequences selected from SEQ ID NOs: 1681 and 815, respectively.

[0082] In some embodiments, the RNAi oligonucleotides are: a) SEQ ID NOs: 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) comprising a sense strand and an antisense strand comprising nucleotide sequences selected from SEQ ID NOs: 1681 and 815, respectively.

[0083] 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.

[0084] It should be understood that in some embodiments, the sequences presented in the sequence listing may be referred to when describing the structure of oligonucleotides (e.g., RNAi oligonucleotides) or other nucleic acids. In such embodiments, the actual oligonucleotide or other nucleic acid may have one or more alternative nucleotides (e.g., the RNA counterpart of a DNA nucleotide or the DNA counterpart of an RNA nucleotide) and / or one or more modified nucleotides and / or one or more modified internucleotide linkages and / or one or more other modifications compared to the specified sequence while retaining substantially the same or similar complementarity characteristics as the specified sequence.

[0085] In some embodiments, the RNAi oligonucleotides herein comprise a 25-nucleotide sense strand and a 27-nucleotide antisense strand that, when acted upon by the Dicer enzyme, result in an antisense strand that is incorporated into the mature RNA-induced silencing complex (RISC). In some embodiments, the 25-nucleotide sense strand comprises a sequence selected from SEQ ID NOs: 1-384. In some aspects, the 27-nucleotide antisense strand comprises a sequence selected from SEQ ID NOs: 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, and 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, and the nucleotide sequence is longer than 25 nucleotides (e.g., 26, 27, 28, 29, or 30 nucleotides).

[0086] In some embodiments, the RNAi oligonucleotide has one 5' end that is thermodynamically less stable compared to the other 5' end. In some embodiments, an asymmetric RNAi oligonucleotide is provided that includes a blunt 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 from about 1 to 8 nucleotides in length (e.g., 1, 2, 3, 4, 5, 6, 7, or 8 nucleotides in length). Typically, the RNAi oligonucleotide has a 2-nucleotide overhang at the 3' end of the antisense (guide) strand, although other overhangs are possible. In some embodiments, the overhang is a 3' overhang that includes from 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 that includes from 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 includes a complementary targeting sequence or region that is complementary to a contiguous sequence of any one of nucleotides of SEQ ID NOs: 912 - 1295 and a 5' overhang that includes nucleotides between about 1 and about 6 in length. In some embodiments, the RNAi oligonucleotide includes a sense strand that includes a nucleotide sequence selected from SEQ ID NOs: 912 - 1295, wherein the RNAi oligonucleotide includes a 5' overhang that includes nucleotides between about 1 and about 6 in length. In some embodiments, the RNAi oligonucleotide includes an antisense strand that includes a nucleotide sequence selected from SEQ ID NOs: 1296 - 1679, wherein the RNAi oligonucleotide includes a 5' overhang that includes nucleotides between about 1 and about 6 in length.In some embodiments, the RNAi oligonucleotide comprises a sense strand comprising a nucleotide sequence selected from SEQ ID NOs: 912 to 1295 and an antisense strand comprising a nucleotide sequence selected from SEQ ID NOs: 1296 to 1679, wherein the RNAi oligonucleotide comprises a 5' overhang comprising nucleotides between about 1 and about 6 in length.

[0087] In some embodiments, the two terminal nucleotides at the 3'-end of the antisense strand are modified. In some embodiments, the two terminal nucleotides at the 3'-end of the antisense strand are complementary to the target mRNA (e.g., the mRNA of MAPT). In some embodiments, the two terminal nucleotides at the 3'-end of the antisense strand are not complementary to the target mRNA. In some embodiments, the two terminal nucleotides at the 3'-end of the antisense strand of the RNAi nucleotides herein do not base pair. In some embodiments, the two terminal nucleotides at the 3'-end of the antisense strand of the RNAi nucleotides herein contain unpaired GG. In some embodiments, the two terminal nucleotides at the 3'-end of the antisense strand of the RNAi oligonucleotides herein are not complementary to the target mRNA. In some embodiments, the two terminal nucleotides at the 3'-end of each of the RNAi oligonucleotides are GG. Usually, one or both of the two terminal GG nucleotides at the 3'-end of each of the RNAi oligonucleotides are not complementary to the target mRNA. In some embodiments, the two terminal nucleotides at the 3'-end of each of the oligonucleotides are GG. In some embodiments, one or both of the two terminal GG nucleotides at the 3'-end of each of the RNAi oligonucleotides are not complementary to the target mRNA. In some embodiments, the RNAi oligonucleotide comprises a complementary target sequence or region that is complementary to a contiguous sequence of any one of nucleotides of SEQ ID NOs: 912 to 1295, wherein the two terminal nucleotides on the 3'-end of the antisense strand of the oligonucleotide herein contain unpaired GG. In some embodiments, the RNAi oligonucleotide comprises an antisense strand comprising a nucleotide sequence selected from SEQ ID NOs: 1296 to 1679, and the two terminal nucleotides on the 3'-end of the antisense strand of the RNAi oligonucleotide contain unpaired GG.In some embodiments, the RNAi oligonucleotide comprises a sense strand comprising a nucleotide sequence selected from SEQ ID NOs: 912 to 1295, and an antisense strand comprising a nucleotide sequence selected from SEQ ID NOs: 1296 to 1679, and the two terminal nucleotides on the 3' end of the antisense strand of the RNAi oligonucleotide comprise unpaired GG.

[0088] In some embodiments, there is one or more (e.g., 1, 2, 3, 4, 5) mismatches (plural possible) between the sense strand and the antisense strand that make up the RNAi oligonucleotide. When there are more than one mismatch between the sense strand and the antisense strand, they may be arranged continuously (e.g., 2, 3, or more in a row), or may be scattered 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, the base mismatch or destabilization of the segment at the 3' end of the sense strand of the RNAi oligonucleotide improves or enhances the efficacy of the oligonucleotide.

[0089] In some embodiments, the RNAi oligonucleotide is: a) SEQ ID NOs: 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; l) SEQ ID NOs: 780 and 815 respectively; m) SEQ ID NOs: 781 and 816, respectively; n) SEQ ID NOs: 782 and 817, respectively; o) SEQ ID NOs: 783 and 818, respectively; p) SEQ ID NOs: 784 and 819, respectively; q) SEQ ID NOs: 785 and 820, respectively; r) SEQ ID NOs: 786 and 821, respectively; s) SEQ ID NOs: 787 and 822, respectively; t) SEQ ID NOs: 788 and 823, respectively; u) SEQ ID NOs: 789 and 824, respectively; v) SEQ ID NOs: 790 and 825, respectively; w) SEQ ID NOs: 791 and 826, respectively; x) SEQ ID NOs: 792 and 827, respectively; y) SEQ ID NOs: 793 and 828, respectively; z) SEQ ID NOs: 794 and 829, respectively; aa) SEQ ID NOs: 795 and 830, respectively; bb) SEQ ID NOs: 796 and 831, respectively; cc) SEQ ID NOs: 797 and 832, respectively; dd) SEQ ID NOs: 798 and 833, respectively; ee) SEQ ID NOs: 799 and 834, respectively; ff) SEQ ID NOs: 800 and 835, respectively; gg) SEQ ID NOs: 801 and 836, respectively; hh) SEQ ID NOs: 802 and 837, respectively ii) SEQ ID NOs: 803 and 838, respectively; and jj) a sense strand and an antisense strand each comprising a nucleotide sequence selected from SEQ ID NOs: 1681 and 815, respectively, with one or more (e.g., 1, 2, 3, 4, 5) mismatches (s) between the sense strand and the antisense strand.

[0090] In some embodiments, the RNAi oligonucleotide is: a) SEQ ID NOs: 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) a sense strand and an antisense strand comprising nucleotide sequences selected from SEQ ID NOs: 1681 and 815, respectively, wherein there is one or more (e.g., 1, 2, 3, 4, 5) mismatches (plural possible) between the sense strand and the antisense strand.

[0091] In some embodiments, the RNAi oligonucleotide is: a) SEQ ID NOs: 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) a sense strand and an antisense strand comprising sequences selected from SEQ ID NOs: 1681 and 815, respectively, wherein there is one or more (e.g., 1, 2, 3, 4, 5) mismatches (plural possible) between the sense strand and the antisense strand.

[0092] Antisense strand

[0093] 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 with or binds to one or more similar factors to direct the silencing of the target gene. In some embodiments, the sense strand complementary to the guide strand is referred to as the "passenger strand".

[0094] In some embodiments, the oligonucleotide comprises an antisense strand that is up to about 50 nucleotides in length (e.g., up to 50, up to 40, up to 35, up to 30, up to 27, up to 25, up to 21, up to 19, up to 17, up to 15, or up to 12 nucleotides). In some embodiments, the oligonucleotide comprises an antisense strand that is at least 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 some embodiments, the oligonucleotide comprises an antisense strand in the range of about 12 to about 40 nucleotides in length (e.g., 12 - 40, 12 - 36, 12 - 32, 12 - 28, 15 - 40, 15 - 36, 15 - 32, 15 - 28, 17 - 22, 17 - 25, 19 - 27, 19 - 30, 20 - 40, 22 - 40, 25 - 40, or 32 - 40). In some embodiments, the oligonucleotide comprises an antisense of about 15 to about 30 nucleotides in length. In some embodiments, the antisense strand of any one 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 that is 22 nucleotides in length.

[0095] In some embodiments, the oligonucleotide for targeting MAPT comprises, or consists of, an antisense strand comprising the sequence shown in any one of SEQ ID NOs: 1296 to 1679. In some embodiments, the oligonucleotide comprises an antisense strand comprising at least 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 nucleotides of the sequence shown in any one of SEQ ID NOs: 1296 to 1679. In some embodiments, the oligonucleotide for targeting MAPT comprises, or consists of, an antisense strand comprising the sequence shown in any one of SEQ ID NOs: 385 to 768. In some embodiments, the oligonucleotide comprises an antisense strand comprising at least 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 nucleotides of the sequence shown in any one of SEQ ID NOs: 385 to 768. In some embodiments, the oligonucleotide for targeting MAPT comprises, or consists of, an antisense strand comprising the sequence shown in any one of SEQ ID NOs: 804 to 838. In some embodiments, the oligonucleotide comprises an antisense strand comprising at least 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 nucleotides of the sequence shown in any one of SEQ ID NOs: 804 to 838.In some embodiments, the oligonucleotide for targeting MAPT comprises, or consists of, an antisense strand comprising the sequence 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 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 nucleotides of the sequence 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 for targeting MAPT comprises, or consists of, an antisense strand comprising the sequence 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 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 nucleotides of the sequence 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, or consists of, an antisense strand comprising the sequence shown in any one of SEQ ID NOs: 806, 815, 816, 833, 834, and 838. In some embodiments, the oligonucleotides herein comprise an antisense strand comprising at least 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 nucleotides of the sequence shown in any one of SEQ ID NOs: 806, 815, 816, 833, 834, and 838.

[0096] Sense strand In some embodiments, the oligonucleotide (e.g., RNAi oligonucleotide) for targeting MAPT mRNA comprises, or consists of, a sense strand comprising a sequence as shown in any one of SEQ ID NOs: 912 to 1295. In some embodiments, the oligonucleotide has a sense strand comprising at least 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 nucleotides of the sequence shown in any one of SEQ ID NOs: 912 to 1295. In some embodiments, the oligonucleotide comprises a sense strand sequence shown in any one of SEQ ID NOs: 1 to 384. In some embodiments, the oligonucleotide has a sense strand comprising at least 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 nucleotides of the sequence shown in any one of SEQ ID NOs: 1 to 384. In some embodiments, the oligonucleotide comprises a sense strand sequence shown in any one of SEQ ID NOs: 769 to 803. In some embodiments, the oligonucleotide comprises a sense strand sequence shown in any one of SEQ ID NOs: 769 to 803 and 1681. In some embodiments, the oligonucleotide comprises a sense strand sequence shown in SEQ ID NO: 1681. In some embodiments, the oligonucleotide has a sense strand composed of at least 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 nucleotides of the sequence shown in any one of SEQ ID NOs: 769 to 803.In some embodiments, the oligonucleotide comprises a sense strand sequence shown 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 composed of at least 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 nucleotides of the sequence shown 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 comprises a sense strand sequence shown 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 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 nucleotides of the sequence shown 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 shown 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 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 nucleotides of the sequence shown in any one of SEQ ID NOs: 771, 780, 781, 798, 799, and 803.

[0097] In some embodiments, the oligonucleotide comprises the sense strand sequence shown 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 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 nucleotides of the sequence shown 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 the sense strand sequence shown 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 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 nucleotides of the sequence shown in any one of SEQ ID NOs: 771, 780, 781, 798, 799, 803, and 1681.

[0098] In some embodiments, the oligonucleotide comprises a sense strand of up to about 50 nucleotides in length (e.g., up to 50, up to 40, up to 36, up to 30, up to 27, up to 25, up to 21, up to 19, up to 17, or up to 12 nucleotides). In some embodiments, the oligonucleotide may have a sense 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 25, at least 27, at least 30, at least 36, or at least 38 nucleotides). In some embodiments, the oligonucleotide may have a sense strand in the range of about 12 to about 50 nucleotides in length (e.g., 12 - 50, 12 - 40, 12 - 36, 12 - 32, 12 - 28, 15 - 40, 15 - 36, 15 - 32, 15 - 28, 17 - 21, 17 - 25, 19 - 27, 19 - 30, 20 - 40, 22 - 40, 25 - 40, or 32 - 40). In some embodiments, the oligonucleotide comprises a sense strand of about 15 to about 30 nucleotides in length. In some embodiments, the oligonucleotide comprises a sense strand of 18 to 36 nucleotides in length. In some embodiments, the oligonucleotide may have a sense strand of 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 of 36 nucleotides in length.

[0099] In some embodiments, the oligonucleotide comprises a sense strand that includes a stem-loop structure at the 3' end of the sense strand. In some embodiments, the stem-loop is formed by intramolecular base pairing. In some embodiments, the sense strand includes a stem-loop structure at its 5' end. In some embodiments, the stem of the stem-loop comprises a double-stranded region 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 double-stranded region of 2 nucleotides in length. In some embodiments, the stem of the stem-loop comprises a double-stranded region of 3 nucleotides in length. In some embodiments, the stem of the stem-loop comprises a double-stranded region of 4 nucleotides in length. In some embodiments, the stem of the stem-loop comprises a double-stranded region of 5 nucleotides in length. In some embodiments, the stem of the stem-loop comprises a double-stranded region of 6 nucleotides in length. In some embodiments, the stem of the stem-loop comprises a double-stranded region of 7 nucleotides in length. In some embodiments, the stem of the stem-loop comprises a double-stranded region of 8 nucleotides in length. In some embodiments, the stem of the stem-loop comprises a double-stranded region of 9 nucleotides in length. In some embodiments, the stem of the stem-loop comprises a double-stranded region of 10 nucleotides in length. In some embodiments, the stem of the stem-loop comprises a double-stranded region of 11 nucleotides in length. In some embodiments, the stem of the stem-loop comprises a double-stranded region of 12 nucleotides in length. In some embodiments, the stem of the stem-loop comprises a double-stranded region of 13 nucleotides in length. In some embodiments, the stem of the stem-loop comprises a double-stranded region of 14 nucleotides in length.

[0100] In some embodiments, the stem-loop provides protection of the oligonucleotide against degradation (e.g., enzymatic degradation), and facilitates, or improves, targeting and / or delivery to a target cell, tissue, or organ (e.g., the liver or the brain), or both. For example, in some embodiments, the loop of the stem-loop contains nucleotides with one or more modifications that facilitate, improve, or enhance targeting to a target mRNA (e.g., MAPT mRNA), inhibition of target gene expression (e.g., MAPT gene expression), and / or delivery to a target cell, tissue, or organ (e.g., the CNS), or combinations thereof. In some embodiments, the stem-loop itself or a modification (s) to the stem-loop does not substantially affect the oligonucleotide's intrinsic gene expression inhibitory activity, but does facilitate, improve, or enhance stability (e.g., provide protection against degradation), and / or delivery of the oligonucleotide to a target cell, tissue, or organ (e.g., the CNS). In certain embodiments, the oligonucleotide comprises a sense strand that includes a stem-loop shown as S1-L-S2 (e.g., at its 3' end), where S1 is complementary to S2, and the loop (L) forms a 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, 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.

[0101] In some embodiments, the oligonucleotide comprises a complementary targeting sequence or region that is complementary to a contiguous sequence of any one of nucleotides of SEQ ID NOs: 912 to 1295, and the oligonucleotide comprises a sense strand that comprises a stem-loop shown as S1-L-S2 (e.g., at its 3' end), wherein S1 is complementary to S2, and L forms a ss loop of nucleotides up to about 10 in length (e.g., nucleotides 3, 4, 5, 6, 7, 8, 9, or 10 in length) between S1 and S2. In some embodiments, the oligonucleotide comprises a complementary targeting sequence or region that is complementary to a contiguous sequence of any one of nucleotides of SEQ ID NOs: 912 to 1295, and the oligonucleotide comprises a sense strand that comprises a stem-loop shown as S1-L-S2 (e.g., at its 3' end), wherein S1 is complementary to S2, and L forms a ss loop of 4 nucleotides in length between S1 and S2.

[0102] In some embodiments, tetra L comprises the sequence 5'-GAAA-3'. In some embodiments, the stem-loop comprises the sequence 5'-GCAGCCGAAAGGCUGC-3' (SEQ ID NO: 1680).

[0103] In some embodiments, L of the stem-loop having the structure S1-L-S2 described above is tri L. In some embodiments, the oligonucleotide comprises a complementary targeting sequence or region that is complementary to a contiguous sequence of any one of nucleotides of SEQ ID NOs: 912 to 1295 and tri L. In some embodiments, tri L comprises ribonucleotides, deoxyribonucleotides, modified nucleotides, delivery ligands, and combinations thereof.

[0104] In some embodiments, the L of the stem-loop having the structure S1-L-S2 as described above is a tetra L as described in U.S. Patent No. 10,131,912, which is incorporated herein by reference (e.g., within the tetra L structure with a nick). In some embodiments, the oligonucleotide comprises a complementary targeting sequence or region that is complementary to a contiguous sequence of any one of nucleotides 912 to 1295 of SEQ ID NO: and the tetra L. In some embodiments, the tetra L comprises ribonucleotides, deoxyribonucleotides, modified nucleotides, delivery ligands, and combinations thereof.

[0105] Double-stranded length

[0106] In some embodiments, the double strand is formed between a sense strand and an 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 double strand formed between the sense strand and the antisense strand ranges from about 12 to about 30 nucleotides in length (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 double strand 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 length. In some embodiments, the double strand formed between the sense strand and the antisense strand is 12 nucleotides in length. In some embodiments, the double strand formed between the sense strand and the antisense strand is 13 nucleotides in length. In some embodiments, the double strand formed between the sense strand and the antisense strand is 14 nucleotides in length. In some embodiments, the double strand formed between the sense strand and the antisense strand is 15 nucleotides in length. In some embodiments, the double strand formed between the sense strand and the antisense strand is 16 nucleotides in length. In some embodiments, the double strand formed between the sense strand and the antisense strand is 17 nucleotides in length. In some embodiments, the double strand formed between the sense strand and the antisense strand is 18 nucleotides in length. In some embodiments, the double strand formed between the sense strand and the antisense strand is 19 nucleotides in length. In some embodiments, the double strand formed between the sense strand and the antisense strand is 20 nucleotides in length. In some embodiments, the double strand formed between the sense strand and the antisense strand is 21 nucleotides in length. In some embodiments, the double strand formed between the sense strand and the antisense strand is 22 nucleotides in length.In some embodiments, the double-strand formed between the sense strand and the antisense strand is 23 nucleotides in length. In some embodiments, the double-strand formed between the sense strand and the antisense strand is 24 nucleotides in length. In some embodiments, the double-strand formed between the sense strand and the antisense strand is 25 nucleotides in length. In some embodiments, the double-strand formed between the sense strand and the antisense strand is 26 nucleotides in length. In some embodiments, the double-strand formed between the sense strand and the antisense strand is 27 nucleotides in length. In some embodiments, the double-strand formed between the sense strand and the antisense strand is 28 nucleotides in length. In some embodiments, the double-strand formed between the sense strand and the antisense strand is 29 nucleotides in length. In some embodiments, the double-strand formed between the sense strand and the antisense strand is 30 nucleotides in length. In some embodiments, the double-strand formed between the sense strand and the antisense strand does not cover the entire length of the sense strand and / or the antisense strand. In some embodiments, the double-strand between the sense strand and the antisense strand covers the entire length of either the sense strand or the antisense strand. In some embodiments, the double-strand between the sense strand and the antisense strand covers the entire length of both the sense strand and the antisense strand.

[0107] In some embodiments, the double-strand between the sense strand and the antisense strand covers 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 are: a) SEQ ID NO: 769 and 804, respectively; b) SEQ ID NO: 770 and 805, respectively; c) SEQ ID NO: 771 and 806, respectively; d) SEQ ID NO: 772 and 807, respectively; e) SEQ ID NO: 773 and 808, respectively; f) SEQ ID NO: 774 and 809, respectively; g) SEQ ID NO: 775 and 810, respectively; h) SEQ ID NO: 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; l) SEQ ID NOs: 780 and 815, respectively; m) SEQ ID NOs: 781 and 816, respectively; n) SEQ ID NOs: 782 and 817, respectively; o) SEQ ID NOs: 783 and 818, respectively; p) SEQ ID NOs: 784 and 819, respectively; q) SEQ ID NOs: 785 and 820, respectively; r) SEQ ID NOs: 786 and 821, respectively; s) SEQ ID NOs: 787 and 822, respectively; t) SEQ ID NOs: 788 and 823, respectively; u) SEQ ID NOs: 789 and 824, respectively; v) SEQ ID NOs: 790 and 825, respectively; w) SEQ ID NOs: 791 and 826, respectively; x) SEQ ID NOs: 792 and 827, respectively; y) SEQ ID NOs: 793 and 828, respectively; z) SEQ ID NOs: 794 and 829, respectively; aa) SEQ ID NOs: 795 and 830, respectively; bb) SEQ ID NOs: 796 and 831, respectively; cc) SEQ ID NOs: 797 and 832, respectively; dd) SEQ ID NOs: 798 and 833, respectively; ee) SEQ ID NOs: 799 and 834, respectively; ff) SEQ ID NOs: 800 and 835, respectively; gg) SEQ ID NOs: 801 and 836, respectively; hh) SEQ ID NOs: 802 and 837, respectively ii) SEQ ID NOs: 803 and 838, respectively; and jj) It contains nucleotide sequences selected from the group consisting of SEQ ID NO: 1681 and 815 respectively, and at this time, the double-stranded formed between the sense strand and the antisense strand is within the range of about 12 to about 30 nucleotides in length (for example, nucleotides of length 12 - 30, 12 - 27, 12 - 22, 15 - 25, 18 - 30, 18 - 22, 18 - 25, 18 - 27, 18 - 30, 19 - 30 or 21 - 30).

[0108] In some embodiments, the double-stranded between the sense strand and the antisense strand extends over 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 are: a) SEQ ID NO: 771 and 806 respectively; b) SEQ ID NO: 776 and 811 respectively; c) SEQ ID NO: 780 and 815 respectively; d) SEQ ID NO: 781 and 816 respectively; e) SEQ ID NO: 782 and 817 respectively; f) SEQ ID NO: 790 and 825 respectively; g) SEQ ID NO: 795 and 830 respectively; h) SEQ ID NO: 798 and 833 respectively; i) SEQ ID NO: 799 and 834 respectively; j) SEQ ID NO: 803 and 838 respectively; and k) It contains nucleotide sequences selected from the group consisting of SEQ ID NO: 1681 and 815 respectively, and at this time, the double-stranded formed between the sense strand and the antisense strand is within the range of about 12 to about 30 nucleotides in length (for example, nucleotides of length 12 - 30, 12 - 27, 12 - 22, 15 - 25, 18 - 30, 18 - 22, 18 - 25, 18 - 27, 18 - 30, 19 - 30 or 21 - 30).

[0109] In some embodiments, the double-stranded between the sense strand and the antisense strand extends over 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 are: 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) a nucleotide sequence selected from the group consisting of SEQ ID NOs: 1681 and 815, respectively, wherein the double-strand formed between the sense strand and the antisense strand is in the range of about 12 to about 30 nucleotides in length (e.g., 12 - 30, 12 - 27, 12 - 22, 15 - 25, 18 - 30, 18 - 22, 18 - 25, 18 - 27, 18 - 30, 19 - 30 or 21 - 30 nucleotides in length).

[0110] Ends of the oligonucleotide

[0111] In some embodiments, the oligonucleotides herein (e.g., RNAi oligonucleotides) comprise a sense strand and an antisense strand, and the ends of either or both strands comprise blunt ends. In some embodiments, the oligonucleotide is a separate strand that forms an asymmetric double-stranded region having an overhang at the 3' end of the antisense strand and comprises a sense strand and an antisense strand. In some embodiments, the oligonucleotide comprises a sense strand and an antisense strand, wherein the ends of either or both strands comprise an overhang comprising one or more nucleotides. In some embodiments, the one or more nucleotides comprising 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 blunt ends. 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 blunt ends.

[0112] 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.

[0113] In some embodiments, the 3' overhang is from about 1 to about 20 nucleotides in length (e.g., 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 from 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.

[0114] In some embodiments, the oligonucleotide comprises a sense strand and an antisense strand, wherein the antisense strand comprises a 3' overhang, and the sense and antisense strands of the oligonucleotide are: a) SEQ ID NO: 769 and 804, respectively; b) SEQ ID NO: 770 and 805, respectively; c) SEQ ID NO: 771 and 806, respectively; d) SEQ ID NO: 772 and 807, respectively; e) SEQ ID NO: 773 and 808, respectively; f) SEQ ID NO: 774 and 809, respectively; g) SEQ ID NO: 775 and 810, respectively; h) SEQ ID NO: 776 and 811, respectively; i) SEQ ID NO: 777 and 812, respectively; j) SEQ ID NO: 778 and 813, respectively; k) SEQ ID NO: 779 and 814, respectively; l) SEQ ID NO: 780 and 815, respectively; m) SEQ ID NO: 781 and 816, respectively; n) SEQ ID NO: 782 and 817, respectively; o) SEQ ID NO: 783 and 818, respectively; p) SEQ ID NO: 784 and 819, respectively; q) SEQ ID NO: 785 and 820, respectively; r) SEQ ID NO: 786 and 821, respectively; s) SEQ ID NO: 787 and 822, respectively; t) SEQ ID NO: 788 and 823, respectively; u) SEQ ID NO: 789 and 824, respectively; v) SEQ ID NO: 790 and 825, respectively; w) SEQ ID NO: 791 and 826, respectively; x) SEQ ID NO: 792 and 827, respectively; y) SEQ ID NO: 793 and 828, respectively; z) SEQ ID NO: 794 and 829, respectively; aa) SEQ ID NO: 795 and 830, respectively; bb) SEQ ID NO: 796 and 831, respectively; (cc) SEQ ID NOs: 797 and 832, respectively; (dd) SEQ ID NOs: 798 and 833, respectively; (ee) SEQ ID NOs: 799 and 834, respectively; (ff) SEQ ID NOs: 800 and 835, respectively; (gg) SEQ ID NOs: 801 and 836, respectively; (hh) SEQ ID NOs: 802 and 837, respectively (ii) SEQ ID NOs: 803 and 838, respectively; and (jj) a nucleotide sequence selected from the group consisting of SEQ ID NOs: 1681 and 815, respectively, wherein the antisense strand comprises a 3' overhang of about 1 to about 20 nucleotides (e.g., 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, 15, 16, 17, 18, 19, or 20 nucleotides in length), and optionally, the 3' overhang is 2 nucleotides in length.

[0115] In some embodiments, the oligonucleotide comprises a sense strand and an antisense strand, wherein the antisense strand comprises a 3' overhang, and the sense and antisense strands of the oligonucleotide are: (a) SEQ ID NOs: 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) comprising a nucleotide sequence selected from the group consisting of SEQ ID NO: 1681 and 815, wherein the antisense strand comprises a 3' overhang of about 1 to about 20 nucleotides (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), and optionally, the 3' overhang is 2 nucleotides in length.

[0116] In some embodiments, the oligonucleotide comprises a sense strand and an antisense strand, wherein the antisense strand comprises a 3' overhang, and the sense strand and the antisense strand of the oligonucleotide are: a) SEQ ID NO: 771 and 806, respectively; b) SEQ ID NO: 780 and 815, respectively; c) SEQ ID NO: 781 and 816, respectively; d) SEQ ID NO: 798 and 833, respectively; e) SEQ ID NO: 799 and 834, respectively; f) SEQ ID NO: 803 and 838, respectively; and k) comprising a nucleotide sequence selected from the group consisting of SEQ ID NO: 1681 and 815, wherein the antisense strand comprises a 3' overhang of about 1 to about 20 nucleotides (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), and optionally, the 3' overhang is 2 nucleotides in length.

[0117] In some embodiments, the oligonucleotide comprises a sense strand and an antisense strand, wherein the sense strand comprises a 5' overhang comprising one or more nucleotides.

[0118] In some embodiments, the oligonucleotide comprises a sense strand and an antisense strand, wherein the sense strand and the antisense strand of the oligonucleotide are: a) SEQ ID NO: 769 and 804, respectively; b) SEQ ID NO: 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; l) SEQ ID NOs: 780 and 815, respectively; m) SEQ ID NOs: 781 and 816, respectively; n) SEQ ID NOs: 782 and 817, respectively; o) SEQ ID NOs: 783 and 818, respectively; p) SEQ ID NOs: 784 and 819, respectively; q) SEQ ID NOs: 785 and 820, respectively; r) SEQ ID NOs: 786 and 821, respectively; s) SEQ ID NOs: 787 and 822, respectively; t) SEQ ID NOs: 788 and 823, respectively; u) SEQ ID NOs: 789 and 824, respectively; v) SEQ ID NOs: 790 and 825, respectively; w) SEQ ID NOs: 791 and 826, respectively; x) SEQ ID NOs: 792 and 827, respectively; y) SEQ ID NOs: 793 and 828, respectively; z) SEQ ID NOs: 794 and 829, respectively; aa) SEQ ID NOs: 795 and 830, respectively; bb) SEQ ID NOs: 796 and 831, respectively; cc) SEQ ID NOs: 797 and 832, respectively; dd) SEQ ID NOs: 798 and 833, respectively; ee) SEQ ID NOs: 799 and 834, respectively; ff) SEQ ID NOs: 800 and 835, respectively; gg) SEQ ID NOs: 801 and 836, respectively; hh) SEQ ID NO: 802 and 837, respectively ii) SEQ ID NO: 803 and 838, respectively; and k) comprising a nucleotide sequence selected from the group consisting of SEQ ID NO: 1681 and 815, respectively, wherein the antisense strand comprises a 3' overhang of about 1 to about 20 nucleotides (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), optionally, the 3' overhang is 2 nucleotides in length.

[0119] In some embodiments, the oligonucleotide comprises a sense strand and an antisense strand, wherein the sense strand and the antisense strand of the oligonucleotide are: a) SEQ ID NO: 771 and 806, respectively b) SEQ ID NO: 776 and 811, respectively c) SEQ ID NO: 780 and 815, respectively d) SEQ ID NO: 781 and 816, respectively e) SEQ ID NO: 782 and 817, respectively f) SEQ ID NO: 790 and 825, respectively g) SEQ ID NO: 795 and 830, respectively h) SEQ ID NO: 798 and 833, respectively i) SEQ ID NO: 799 and 834, respectively j) SEQ ID NO: 803 and 838, respectively; and k) comprising a nucleotide sequence selected from the group consisting of SEQ ID NO: 1681 and 815, respectively, the antisense strand comprises a 3' overhang of about 1 to about 20 nucleotides (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), optionally, the 3' overhang is 2 nucleotides in length.

[0120] In some embodiments, the oligonucleotide comprises a sense strand and an antisense strand, wherein the sense strand and the antisense strand of the oligonucleotide are: a) SEQ ID NOs: 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) a nucleotide sequence selected from the group consisting of SEQ ID NOs: 1681 and 815, respectively, wherein the antisense strand comprises a 3' overhang of about 1 to about 20 nucleotides (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), and optionally, the 3' overhang is 2 nucleotides in length.

[0121] In some embodiments, one or more (e.g., 2, 3, 4, 5 or more) nucleotides including the 3' or 5' end of the sense strand and / or the 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, e.g., including 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.

[0122] In some embodiments, the oligonucleotide comprises a sense strand and an antisense strand, the 3' end of the sense strand comprises a staple 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 that form a nicked tetra-L structure, wherein the 3' end of the sense strand comprises a stem-loop, the loop is a tetra-L, and the 3' end of the antisense strand comprises a 3' overhang as described herein. In some embodiments, the 3' overhang is two nucleotides in length. In some embodiments, both of the two nucleotides that make up the 3' overhang comprise a guanine (G) nucleobase. Typically, one or both of the nucleotides that make up the 3' overhang of the antisense strand are not complementary to the target mRNA.

[0123] Modification of the Oligonucleotide

[0124] In some embodiments, the oligonucleotide (RNAi oligonucleotide) comprises a modification. The oligonucleotide may 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 properties relevant to therapeutic or research applications.

[0125] 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 internucleotide linkage. 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 or any combination of the modifications described herein. For example, in some embodiments, the oligonucleotide comprises at least one modified sugar, a 5' terminal phosphate group, at least one modified internucleotide linkage, at least one modified base, and at least one reversible modification.

[0126] In some embodiments, 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. In some embodiments, the sense and antisense strands of the oligonucleotide are: a) SEQ ID NO: 769 and 804, respectively; b) SEQ ID NO: 770 and 805, respectively; c) SEQ ID NO: 771 and 806, respectively; d) SEQ ID NO: 772 and 807, respectively; e) SEQ ID NO: 773 and 808, respectively; f) SEQ ID NO: 774 and 809, respectively; g) SEQ ID NO: 775 and 810, respectively; h) SEQ ID NO: 776 and 811, respectively; i) SEQ ID NO: 777 and 812, respectively; j) SEQ ID NO: 778 and 813, respectively; k) SEQ ID NO: 779 and 814, respectively; l) SEQ ID NO: 780 and 815, respectively; m) SEQ ID NO: 781 and 816, respectively; n) SEQ ID NO: 782 and 817, respectively; o) SEQ ID NO: 783 and 818, respectively; p) SEQ ID NO: 784 and 819, respectively; q) SEQ ID NO: 785 and 820, respectively; r) SEQ ID NO: 786 and 821, respectively; s) SEQ ID NO: 787 and 822, respectively; t) SEQ ID NO: 788 and 823, respectively; u) SEQ ID NO: 789 and 824, respectively; v) SEQ ID NO: 790 and 825, respectively; w) SEQ ID NO: 791 and 826, respectively; x) SEQ ID NO: 792 and 827, respectively; y) SEQ ID NO: 793 and 828, respectively; z) SEQ ID NO: 794 and 829, respectively; aa) SEQ ID NO: 795 and 830, respectively; bb) SEQ ID NOs: 796 and 831, respectively; cc) SEQ ID NOs: 797 and 832, respectively; dd) SEQ ID NOs: 798 and 833, respectively; ee) SEQ ID NOs: 799 and 834, respectively; ff) SEQ ID NOs: 800 and 835, respectively; gg) SEQ ID NOs: 801 and 836, respectively; hh) SEQ ID NOs: 802 and 837, respectively ii) SEQ ID NOs: 803 and 838, respectively; and jj) a nucleotide sequence selected from the group consisting of 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.

[0127] In some embodiments, the sense and antisense strands of the oligonucleotide are: a) SEQ ID NOs: 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) a nucleotide sequence selected from the group consisting of 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.

[0128] In some embodiments, the sense and antisense strands of the oligonucleotide are: a) SEQ ID NOs: 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 h) a nucleotide sequence selected from the group consisting of 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.

[0129] The number of modifications on the oligonucleotide and the positions of those nucleotide modifications may affect the properties of the oligonucleotide. For example, the oligonucleotide may be delivered in vivo by conjugating it to be incorporated into lipid nanoparticles (LNPs) or similar carriers. However, if the oligonucleotide is not protected by an LNP or similar carrier, it may be advantageous for at least some of the nucleotides to be modified. Thus, in some embodiments, all or substantially all of the 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 moiety of all of the nucleotides that make up the oligonucleotide is modified at the 2' position. The modifications may be reversible or irreversible. In some embodiments, the oligonucleotides disclosed herein have a sufficient number and type of modified nucleotides to result in the desired properties (e.g., protection from enzymatic degradation, the ability to target desired cells after in vivo administration, and / or thermodynamic stability).

[0130] Sugar modification

[0131] In some embodiments, the oligonucleotide comprises at least one modified sugar. In some embodiments, the modified sugar (also referred to herein as a sugar analog) comprises a modified deoxyribose or ribose moiety, for example, in which one or more modifications occur at the 2′, 3′, 4′, and / or 5′ carbon of the sugar. In some embodiments, the modified sugar may also include non-natural alternative carbon structures such as those present in, for example, locked nucleic acids (“LNAs”; see, e.g., Koshkin et al. (1998), Tetrahedon, 54:3607-30), unlocked nucleic acids (“UNAs”; see, e.g., Snead et al. (2013), Mol. Ther-Nucl. Acids, 2:e103), and bridged nucleic acids (“BNAs”; see, e.g., Imanishi & Obika, (2002), Chem. Commun. (Camb), 21:1653-59).

[0132] In some embodiments, the nucleotide modification in the sugar comprises a 2′-modification. In some embodiments, the 2′-modification may be 2′-O-propynyl, 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 comprises a modification of the sugar ring, which may include a 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 attached to the 1′-carbon or 4′-carbon of the sugar, or the 2′-oxygen attached to the 1′-carbon or 4′-carbon via an ethylene bridge or a methylene bridge. In some embodiments, the modified nucleotide has an acyclic sugar lacking a bond between the 2′ and 3′ carbons. In some embodiments, the modified nucleotide has, for example, a thiol group at the 4′ position of the sugar.

[0133] The oligonucleotides described herein contain at least about 1 (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) modified nucleotides. In some embodiments, the sense strand of the RNAi oligonucleotide contains at least about 1 (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) modified nucleotides. In some embodiments, the antisense strand of the oligonucleotide contains at least about 1 (e.g., at least 1, at least 5, at least 10, at least 15, at least 20, or more) modified nucleotides.

[0134] In some embodiments, all nucleotides of the oligonucleotide are modified. In some embodiments, all nucleotides of the antisense strand of the oligonucleotide are modified. In some embodiments, all nucleotides of the oligonucleotide (i.e., both the sense strand and the antisense strand) are modified. In some embodiments, the modified nucleotides include 2'-modifications (e.g., 2'-F, 2'-OMe, 2'-MOE, and 2'-FANA). In some embodiments, the modified nucleotides include 2' modifications (e.g., 2'-F or 2'-OMe).

[0135] In some embodiments, the present disclosure provides oligonucleotides having different modification patterns. In some embodiments, the modified oligonucleotides include a sense strand sequence having the modification pattern shown in the Examples and Sequence Listing, and an antisense strand having the modification pattern shown in the Examples and Sequence Listing.

[0136] In some embodiments, the oligonucleotide comprises an antisense strand having nucleotides modified with 2'-F. In some embodiments, the oligonucleotide comprises an antisense strand comprising nucleotides modified with 2'-F and 2'-OMe. In some embodiments, the oligonucleotide comprises a sense strand having nucleotides modified with 2'-F. In some embodiments, the oligonucleotide comprises a sense strand comprising nucleotides modified with 2'-F and 2'-OMe.

[0137] In some embodiments, the oligonucleotide comprises a sense strand in which about 10-15%, 10%, 11%, 12%, 13%, 14%, or 15% of the nucleotides of the sense strand contain a 2'-F modification. In some embodiments, the oligonucleotide comprises a sense strand in which about 18-23% (e.g., 18%, 19%, 20%, 21%, 22%, or 23%) of the nucleotides of the sense strand contain a 2'-F modification. In some embodiments, the oligonucleotide comprises a sense strand in which about 38-43% (e.g., 38%, 39%, 40%, 41%, 42%, or 43%) of the nucleotides of the sense strand contain a 2'-F modification. In some embodiments, about 11% of the nucleotides of the sense strand contain a 2'-F modification. In some embodiments, about 22% of the nucleotides of the sense strand contain a 2'-F modification. In some embodiments, about 40% of the nucleotides of the sense strand contain a 2'-F modification. In some embodiments, the oligonucleotide comprises an antisense strand in which about 25%-about 35% (e.g., 25%, 26%, 27%, 28%, 29%, 30%, 31%, 32%, 33%, 34%, or 35%) of the nucleotides of the antisense strand contain a 2'-F modification. In some embodiments, about 32% of the nucleotides of the antisense strand contain a 2'-F modification. In some embodiments, the oligonucleotide has about 15%-about 25% (e.g., 15%, 16%, 17%, 18%, 19%, 20%, 21%, 22%, 23%, 24%, or 25%) of its nucleotides contain a 2'-F modification. In some embodiments, the oligonucleotide has 35%-45% (e.g., 35%, 36%, 37%, 38%, 39%, 40%, 41%, 42%, 43%, 44%, or 45%) of its nucleotides contain a 2'-F modification. In some embodiments, about 19% of the nucleotides in the oligonucleotide contain a 2'-F modification. In some embodiments, about 29% of the nucleotides in the oligonucleotide contain a 2'-F modification. In some embodiments, about 40% of the nucleotides in the oligonucleotide contain a 2'-F modification.

[0138] In some embodiments, one or more of the 8th, 9th, 10th, or 11th positions of the 36-nucleotide sense strand are modified with 2'-F. In some embodiments, one or more of the 8th, 9th, 10th, or 11th positions of the sense strand containing a stem-loop are modified with 2'-F. In some embodiments, the respective sugar moieties of the nucleotides at positions 1-7 and 12-20 of the 36-nucleotide sense strand are modified with 2'-OMe. In some embodiments, the respective sugar moieties of the nucleotides at positions 1-7 and 12-20 of the sense strand containing a stem-loop are modified with 2'-OMe. In some embodiments, the respective sugar moieties of the nucleotides at positions 1-7 and 12-36 of the sense strand are modified with 2'-OMe.

[0139] In some embodiments, one or more of the 3rd, 5th, 8th, 10th, 12th, 13th, 15th, and 17th positions of the sense strand are modified with 2'-F.

[0140] In some embodiments, the antisense strand has three nucleotides modified with 2'-F at the 2'-position of the sugar moiety. In some embodiments, the sugar moieties at the 2nd, 5th, and 14th positions of the antisense strand, and optionally up to three of the nucleotides at the 1st, 3rd, 7th, and 10th positions are modified with 2'-F. In some embodiments, up to three of the sugar moieties at the 2nd, 5th, and 14th positions of the antisense strand, and optionally the nucleotides at the 3rd, 4th, 7th, and 10th positions are modified with 2'-F. In other embodiments, the sugar moiety at each of the 2nd, 5th, and 14th positions of the antisense strand is modified with 2'-F. In other embodiments, the sugar moieties at each of the 1st, 2nd, 5th, and 14th positions of the antisense strand are modified with 2'-F. In other embodiments, the sugar moieties at each of the 2nd, 4th, 5th, and 14th positions of the antisense strand are modified with 2'-F. In still other embodiments, the sugar moieties at each of the 1st, 2nd, 3rd, 5th, 7th, and 14th positions of the antisense strand are modified with 2'-F. In other embodiments, the sugar moieties at each of the 2nd, 3rd, 4th, 5th, 7th, and 14th positions of the antisense strand are modified with 2'-F. In yet another embodiment, the sugar moieties at each of the 1st, 2nd, 3rd, 5th, 10th, and 14th positions of the antisense strand are modified with 2'-F. In other embodiments, the sugar moieties at each of the 2nd, 3rd, 4th, 5th, 10th, and 14th positions of the antisense strand are modified with 2'-F. In another embodiment, the sugar moieties at each of the 2nd, 3rd, 5th, 7th, 10th, and 14th positions of the antisense strand are modified with 2'-F. In other embodiments, the sugar moieties at each of the 2nd, 3rd, 4th, 5th, 7th, 10th, and 14th positions of the antisense strand that is double-stranded with a 36-nucleotide sense strand are modified with 2'-F. In other embodiments, the sugar moieties at each of the 2nd, 3rd, 4th, 5th, 7th, 10th, and 14th positions of the antisense strand that is double-stranded with a sense strand containing a stem-loop are modified with 2'-F.

[0141] In some embodiments, the oligonucleotide comprises an antisense strand having sugar moieties at positions 2 and 14 modified with 2'-F. In some embodiments, the oligonucleotide comprises an antisense strand having sugar moieties at positions 2, 5 and 14 modified with 2'-F. In some embodiments, the oligonucleotide comprises an antisense strand having sugar moieties at positions 1, 2, 5 and 14 modified with 2'-F. In some embodiments, the oligonucleotide comprises an antisense strand having sugar moieties at positions 2, 4, 5 and 14 modified with 2'-F. In some embodiments, the oligonucleotide comprises an antisense strand having sugar moieties at positions 1, 2, 3, 5, 7 and 14 modified with 2'-F. In some embodiments, the oligonucleotide comprises an antisense strand having sugar moieties at positions 2, 3, 4, 5, 7 and 14 modified with 2'-F. In some embodiments, the oligonucleotide comprises an antisense strand having sugar moieties at positions 1, 2, 3, 5, 10 and 14 modified with 2'-F. In some embodiments, the oligonucleotide comprises an antisense strand having sugar moieties at positions 2, 3, 4, 5, 10 and 14 modified with 2'-F. In some embodiments, the oligonucleotide comprises a 36-nucleotide sense strand and antisense strand, wherein the antisense strand comprises sugar moieties at positions 2, 3, 4, 5, 7, 10, and 14 modified with 2'-F. In some embodiments, the oligonucleotide comprises a sense strand and antisense strand comprising a stem-loop, wherein the antisense strand comprises sugar moieties at positions 2, 3, 4, 5, 7, 10, and 14 modified with 2'-F. In some embodiments, the oligonucleotide comprises an antisense strand having sugar moieties at positions 2, 3, 4, 5, 7, 10, 14, 16, and 19 modified with 2'-F.

[0142] In some embodiments, the oligonucleotide comprises an antisense strand having a sugar moiety of each nucleotide at positions 2, 5, and 14 of the antisense strand modified with 2'-F, and a sugar moiety of each of the remaining nucleotides of the antisense strand modified with a modification selected from the group consisting of 2'-O-propynyl, 2'-O-propylamine, 2'-amino, 2'-ethyl, EA, 2'-OMe, 2'-MOE, 2'-O-NMA, and 2'-FANA.

[0143] In some embodiments, the oligonucleotide comprises an antisense strand having a sugar moiety of each nucleotide at positions 1, 2, 5, and 14 of the antisense strand modified with 2'-F, and a sugar moiety of each of the remaining nucleotides of the antisense strand modified with a modification selected from the group consisting of 2'-O-propynyl, 2'-O-propylamine, 2'-amino, 2'-ethyl, EA, 2'-OMe, 2'-MOE, 2'-O-NMA, and 2'-FANA.

[0144] In some embodiments, the oligonucleotide comprises an antisense strand having a sugar moiety of each nucleotide at positions 2, 4, 5, and 14 of the antisense strand modified with 2'-F, and a sugar moiety of each of the remaining nucleotides of the antisense strand modified with a modification selected from the group consisting of 2'-O-propynyl, 2'-O-propylamine, 2'-amino, 2'-ethyl, EA, 2'-OMe, 2'-MOE, 2'-O-NMA, and 2'-FANA.

[0145] In some embodiments, the oligonucleotide comprises an antisense strand having a sugar moiety of each nucleotide at positions 1, 2, 3, 5, 7, and 14 of the antisense strand modified with 2'-F, and a sugar moiety of each of the remaining nucleotides of the antisense strand modified with a modification selected from the group consisting of 2'-O-propynyl, 2'-O-propylamine, 2'-amino, 2'-ethyl, EA, 2'-OMe, 2'-MOE, 2'-O-NMA, and 2'-FANA.

[0146] In some embodiments, the oligonucleotide comprises an antisense strand having the sugar moiety of each nucleotide at positions 2, 3, 4, 5, 7, and 14 of the antisense strand modified with 2'-F, and the sugar moiety of each of the remaining nucleotides of the antisense strand modified with a modification selected from the group consisting of 2'-O-propynyl, 2'-O-propylamine, 2'-amino, 2'-ethyl, EA, 2'-OMe, 2'-MOE, 2'-O-NMA, and 2'-FANA.

[0147] In some embodiments, the oligonucleotide comprises an antisense strand having the sugar moiety of each nucleotide at positions 1, 2, 3, 5, 10, and 14 of the antisense strand modified with 2'-F, and the sugar moiety of each of the remaining nucleotides of the antisense strand modified with a modification selected from the group consisting of 2'-O-propynyl, 2'-O-propylamine, 2'-amino, 2'-ethyl, EA, 2'-OMe, 2'-MOE, 2'-O-NMA, and 2'-FANA.

[0148] In some embodiments, the oligonucleotide comprises an antisense strand having the sugar moiety of each nucleotide at positions 2, 3, 4, 5, 10, and 14 of the antisense strand modified with 2'-F, and the sugar moiety of each of the remaining nucleotides of the antisense strand modified with a modification selected from the group consisting of 2'-O-propynyl, 2'-O-propylamine, 2'-amino, 2'-ethyl, EA, 2'-OMe, 2'-MOE, 2'-O-NMA, and 2'-FANA.

[0149] In some embodiments, the oligonucleotide comprises an antisense strand having the sugar moiety of each nucleotide at positions 2, 3, 5, 7, 10, and 14 of the antisense strand modified with 2'-F, and the sugar moiety of each of the remaining nucleotides of the antisense modified with a modification selected from the group consisting of 2'-O-propynyl, 2'-O-propylamine, 2'-amino, 2'-ethyl, EA, 2'-OMe, 2'-MOE, 2'-O-NMA, and 2'-FANA.

[0150] In some embodiments, the oligonucleotide comprises a sense strand and an antisense strand of 36 nucleotides, and the antisense strand comprises sugar moieties at positions 2, 3, 4, 5, 7, 10, and 14 of the antisense strand modified with 2'-F, and sugar moieties of each of the remaining nucleotides of the antisense strand modified with a modification selected from the group consisting of 2'-O-propynyl, 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 that includes a stem-loop, and the antisense strand comprises sugar moieties at positions 2, 3, 4, 5, 7, 10, and 14 of the antisense strand modified with 2'-F, and sugar moieties of each of the remaining nucleotides of the antisense modified with a modification selected from the group consisting of 2'-O-propynyl, 2'-O-propylamine, 2'-amino, 2'-ethyl, EA, 2'-OMe, 2'-MOE, 2'-O-NMA, and 2'-FANA.

[0151] In some embodiments, the oligonucleotide comprises an antisense strand having sugar moieties at positions 2, 3, 4, 5, 7, 10, 14, 16, and 19 of the antisense strand modified with 2'-F, and sugar moieties of each of the remaining nucleotides of the antisense strand modified with a modification selected from the group consisting of 2'-O-propynyl, 2'-O-propylamine, 2'-amino, 2'-ethyl, EA, 2'-OMe, 2'-MOE, 2'-O-NMA, and 2'-FANA.

[0152] In some embodiments, the oligonucleotide comprises an antisense strand having a sugar moiety at position 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, 15, 16, 17, 18, 19, 20, 21, or 22 modified with 2'-F.

[0153] In some embodiments, the oligonucleotide comprises an antisense strand having a sugar moiety at the 1st, 2nd, 3rd, 4th, 5th, 6th, 7th, 8th, 9th, 10th, 11th, 12th, 13th, 14th, 15th, 16th, 17th, 18th, 19th, 20th, 21st, or 22nd position modified with 2'-OMe.

[0154] In some embodiments, the oligonucleotide comprises an antisense strand having a sugar moiety at the 1st, 2nd, 3rd, 4th, 5th, 6th, 7th, 8th, 9th, 10th, 11th, 12th, 13th, 14th, 15th, 16th, 17th, 18th, 19th, 20th, 21st, or 22nd position modified with a modification selected from the group consisting of 2'-O-propynyl, 2'-O-propylamine, 2'-amino, 2'-ethyl, EA, 2'-OMe, 2'-MOE, 2'-O-NMA, and 2'-FANA.

[0155] In some embodiments, the oligonucleotide comprises a sense strand of 36 nucleotides having a sugar moiety at positions 8 to 11 modified with 2'-F. In some embodiments, the oligonucleotide comprises a sense strand comprising a stem-loop and a sugar moiety at positions 8 to 11 modified with 2'-F. In some embodiments, the oligonucleotide comprises a sense strand of 36 nucleotides having a sugar moiety at positions 1 to 7 and 12 to 17 or 12 to 20 modified with 2'-OMe. In some embodiments, the oligonucleotide comprises a sense strand comprising a stem-loop and a sugar moiety at positions 1 to 7 and 12 to 17 or 12 to 20 modified with 2'-OMe. In some embodiments, the oligonucleotide comprises a sense strand of 36 nucleotides having a sugar moiety at positions 1 to 7 and 12 to 17, 12 to 20 or 12 to 22 modified with 2'-OMe. In some embodiments, the oligonucleotide comprises a sense strand comprising a stem-loop and a sugar moiety at positions 1 to 7 and 12 to 17, 12 to 20 or 12 to 22 modified with 2'-OMe. In some embodiments, the oligonucleotide comprises a sense strand of 36 nucleotides having a respective sugar moiety of each nucleotide at positions 1 to 7 and 12 to 17 or 12 to 20 of the sense strand modified with a modification selected from the group consisting of 2'-O-propynyl, 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 stem-loop and comprises a sense strand having a respective sugar moiety of each nucleotide at positions 1 to 7 and 12 to 17 or 12 to 20 of the sense strand modified with a modification selected from the group consisting of 2'-O-propynyl, 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 36-nucleotide sense strand having the respective sugar moieties of the nucleotides at positions 1 to 7 and positions 12 to 17, 12 to 20, or 12 to 22 of the sense strand modified with a modification selected from the group consisting of 2'-O-propynyl, 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 a stem-loop, and the respective sugar moieties of the nucleotides at positions 1 to 7 and positions 12 to 17, 12 to 20, or 12 to 22 of the sense strand modified with a modification selected from the group consisting of 2'-O-propynyl, 2'-O-propylamine, 2'-amino, 2'-ethyl, EA, 2'-OMe, 2'-MOE, 2'-O-NMA, and 2'-FANA.

[0156] In some embodiments, the oligonucleotide comprises a sense strand having the sugar moieties at positions 2, 3, 5, 8, 10, 12, 13, 15, and 17 modified with 2'-F. In some embodiments, the oligonucleotide comprises a sense strand having the sugar moieties at positions 1, 2, 4, 6, 7, 9, 11, 14, 16, and 18 to 20 modified with 2'-OMe. In some embodiments, the oligonucleotide comprises a sense strand having the respective sugar moieties of the nucleotides at positions 1, 2, 4, 6, 7, 9, 11, 14, 16, and 18 to 20 of the sense strand modified with a modification selected from the group consisting of 2'-O-propynyl, 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 having the respective sugar moieties of the nucleotides at positions 1 to 7 and positions 12 to 17, 12 to 20, or 12 to 22 of the sense strand modified with a modification selected from the group consisting of 2'-O-propynyl, 2'-O-propylamine, 2'-amino, 2'-ethyl, EA, 2'-OMe, 2'-MOE, 2'-O-NMA, and 2'-FANA.

[0157] In some embodiments, the oligonucleotide comprises a sense strand having a sugar moiety at the 1st, 2nd, 3rd, 4th, 5th, 6th, 7th, 8th, 9th, 10th, 11th, 12th, 13th, 14th, 15th, 16th, 17th, 18th, 19th, 20th, 21st, 22nd, 23rd, 24th, 25th, 26th, 27th, 28th, 29th, 30th, 31st, 32nd, 33rd, 34th, 35th, or 36th position modified with 2'-F.

[0158] In some embodiments, the oligonucleotide comprises a sense strand having a sugar moiety at the 1st, 2nd, 3rd, 4th, 5th, 6th, 7th, 8th, 9th, 10th, 11th, 12th, 13th, 14th, 15th, 16th, 17th, 18th, 19th, 20th, 21st, 22nd, 23rd, 24th, 25th, 26th, 27th, 28th, 29th, 30th, 31st, 32nd, 33rd, 34th, 35th, or 36th position modified with 2'-OMe.

[0159] In some embodiments, the oligonucleotide comprises a sense strand having a sugar moiety at the 1st, 2nd, 3rd, 4th, 5th, 6th, 7th, 8th, 9th, 10th, 11th, 12th, 13th, 14th, 15th, 16th, 17th, 18th, 19th, 20th, 21st, 22nd, 23rd, 24th, 25th, 26th, 27th, 28th, 29th, 30th, 31st, 32nd, 33rd, 34th, 35th, or 36th position 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.

[0160] In some embodiments, the oligonucleotide comprises an antisense strand having the sugar moiety of each of the nucleotides at positions 2, 3, 4, 5, 7, 10, and 14 of the antisense strand modified with 2'-F and the sugar moiety of each of the remaining nucleotides of the antisense strand modified with a modification selected from the group consisting of 2'-O-propynyl, 2'-O-propylamine, 2'-amino, 2'-ethyl, EA, 2'-OMe, 2'-MOE, 2'-O-NMA, and 2'-deoxy-2'-fluoro-β-d-arabinonucleic acid (2'-FANA); and a sense strand having the sugar moiety of each of the nucleotides at positions 8-11 of the sense strand modified with 2'-F and the sugar moiety of each of the remaining nucleotides of the sense strand modified with a modification selected from the group consisting of 2'-O-propynyl, 2'-O-propylamine, 2'-amino, 2'-ethyl, EA, 2'-OMe, 2'-MOE, 2'-O-NMA, and 2'-FANA. In some embodiments, the oligonucleotide comprises an antisense strand having the sugar moiety of each of the nucleotides at positions 2, 3, 4, 5, 7, 10, and 14 of the antisense strand modified with 2'-F and the sugar moiety of each of the remaining nucleotides of the antisense strand modified with a modification selected from the group consisting of 2'-O-propynyl, 2'-O-propylamine, 2'-amino, 2'-ethyl, EA, 2'-OMe, 2'-MOE, 2'-O-NMA, and 2'-FANA; a stem-loop; and a sense strand having the sugar moiety of each of the nucleotides at positions 8-11 of the sense strand modified with 2'-F and the sugar moiety of each of the remaining nucleotides of the sense strand modified with a modification selected from the group consisting of 2'-O-propynyl, 2'-O-propylamine, 2'-amino, 2'-ethyl, EA, 2'-OMe, 2'-MOE, 2'-O-NMA, and 2'-FANA.

[0161] In some embodiments, the oligonucleotide comprises an antisense strand having the sugar moiety of each of the nucleotides at positions 2, 3, 4, 5, 7, 10, 14, 16, and 19 of the antisense strand modified with 2'-F, and the sugar moiety of each of the remaining nucleotides of the antisense strand modified with a modification selected from the group consisting of 2'-O-propynyl, 2'-O-propylamine, 2'-amino, 2'-ethyl, EA, 2'-OMe, 2'-MOE, 2'-O-NMA, and 2'-FANA; and a sense strand having the sugar moiety of each of the nucleotides at positions 3, 5, 8, 10, 12, 13, 15, and 17 of the sense strand modified with 2'-F, and the sugar moiety of each of the remaining nucleotides of the sense strand modified with a modification selected from the group consisting of 2'-O-propynyl, 2'-O-propylamine, 2'-amino, 2'-ethyl, EA, 2'-OMe, 2'-MOE, 2'-O-NMA, and 2'-FANA.

[0162] In some embodiments, the sense and antisense strands of the oligonucleotide are: a) SEQ ID NO: 769 and 804, respectively; b) SEQ ID NO: 770 and 805, respectively; c) SEQ ID NO: 771 and 806, respectively; d) SEQ ID NO: 772 and 807, respectively; e) SEQ ID NO: 773 and 808, respectively; f) SEQ ID NO: 774 and 809, respectively; g) SEQ ID NO: 775 and 810, respectively; h) SEQ ID NO: 776 and 811, respectively; i) SEQ ID NO: 777 and 812, respectively; j) SEQ ID NO: 778 and 813, respectively; k) SEQ ID NO: 779 and 814, respectively; l) SEQ ID NO: 780 and 815, respectively; m) SEQ ID NO: 781 and 816, respectively; n) SEQ ID NO: 782 and 817, respectively; o) SEQ ID NO: 783 and 818, respectively; p) SEQ ID NO: 784 and 819, respectively; q) SEQ ID NO: 785 and 820, respectively; r) SEQ ID NO: 786 and 821, respectively; s) SEQ ID NO: 787 and 822, respectively; t) SEQ ID NO: 788 and 823, respectively; u) SEQ ID NO: 789 and 824, respectively; v) SEQ ID NO: 790 and 825, respectively; w) SEQ ID NO: 791 and 826, respectively; x) SEQ ID NO: 792 and 827, respectively; y) SEQ ID NO: 793 and 828, respectively; z) SEQ ID NO: 794 and 829, respectively; aa) SEQ ID NO: 795 and 830, respectively; bb) SEQ ID NO: 796 and 831, respectively; cc) SEQ ID NO: 797 and 832, respectively; dd) SEQ ID NO: 798 and 833, respectively; ee) SEQ ID NO: 799 and 834, respectively; ff) SEQ ID NO: 800 and 835, respectively; gg) SEQ ID NO: 801 and 836, respectively; hh) SEQ ID NO: 802 and 837, respectively ii) SEQ ID NO: 803 and 838, respectively; and jj) a nucleotide sequence selected from the group consisting of SEQ ID NO: 1681 and 815, wherein one or more of the 3rd, 5th, 8th, 10th, 12th, 13th, 15th, or 17th positions of the sense strand are modified with a 2'-F group.

[0163] In some embodiments, the sense and antisense strands of the oligonucleotide are: a) SEQ ID NO: 771 and 806, respectively; b) SEQ ID NO: 776 and 811, respectively; c) SEQ ID NO: 780 and 815, respectively; d) SEQ ID NO: 781 and 816, respectively; e) SEQ ID NO: 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) a nucleotide sequence selected from the group consisting of 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 is modified with a 2'-F group.

[0164] In some embodiments, the sense and antisense strands of the oligonucleotide are: a) SEQ ID NOs: 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) a nucleotide sequence selected from the group consisting of 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 is modified with a 2'-F group.

[0165] 5'-terminal phosphate group

[0166] In some embodiments, the oligonucleotide comprises a sense strand and an antisense strand, and the antisense strand comprises a 5'-terminal phosphate group. In some embodiments, the 5'-terminal phosphate group of the oligonucleotide enhances the interaction with Ago2. However, oligonucleotides containing a 5'-terminal phosphate group may be susceptible to degradation by phosphatases or other enzymes, which may limit their bioavailability in vivo. In some embodiments, the oligonucleotide comprises an analog of the 5'-phosphate group that is resistant to such degradation. In some embodiments, the phosphate analog is phosphonate oxymethyl, vinyl phosphonate, or malonyl phosphonate, or a combination thereof. In certain embodiments, the 5'-end of the oligonucleotide strand is linked to a chemical moiety (a "phosphate mimic") that mimics the electrostatic and steric properties of the natural 5'-phosphate group.

[0167] In some embodiments, the oligonucleotide has a phosphate analog at the 4'-carbon of the sugar (referred to as a 4'-phosphate analog). See, for example, International Patent Application Publication No. 2018 / 045317. In some embodiments, the oligonucleotide comprises a 4'-phosphate analog at the 5'-terminal nucleotide. In some embodiments, the phosphate analog is phosphonate oxymethyl or an analog thereof in which the oxygen atom of the oxymethyl group is bonded to the sugar moiety (e.g., its 4'-carbon). In other embodiments, the 4'-phosphate analog is phosphonate thiomethyl or phosphonate aminomethyl or an analog thereof in which the sulfur atom of the thiomethyl group or the nitrogen atom of the aminomethyl group is bonded to the 4'-carbon of the sugar moiety. In certain embodiments, the 4'-phosphate analog is phosphonate oxymethyl. In some embodiments, phosphonate oxymethyl is represented by the formula -O-CH 2 -PO(OH) 2 , -O-CH 2 -PO(OR) 2 , or O-CH2-POOH(R), wherein R is independently H, CH 3 , an alkyl group, CH 2 CH 2 CN, CH2 OCOC(CH 3 ) 3 、CH 2 OCH 2 CH 2 Si(CH 3 ) 3 or selected from a protecting group. In certain embodiments, the alkyl group is CH 2 CH 3 . Further usually, R is independently H, CH 3 , or CH 2 CH 3 . In some embodiments, R is CH 3 . In some embodiments, the 4'-phosphate analog is phosphonic acid 4'-oxymethyl. In some embodiments, the modified nucleotide having a 4'-phosphonic acid analog is uridine. In some embodiments, the modified nucleotide is 4'-O-methylphosphonate-2'-O-methyluridine.

[0168] In some embodiments, the sense strand and the antisense strand of the oligonucleotide are: a) SEQ ID NO: 769 and 804, respectively; b) SEQ ID NO: 770 and 805, respectively; c) SEQ ID NO: 771 and 806, respectively; d) SEQ ID NO: 772 and 807, respectively; e) SEQ ID NO: 773 and 808, respectively; f) SEQ ID NO: 774 and 809, respectively; g) SEQ ID NO: 775 and 810, respectively; h) SEQ ID NO: 776 and 811, respectively; i) SEQ ID NO: 777 and 812, respectively; j) SEQ ID NO: 778 and 813, respectively; k) SEQ ID NO: 779 and 814, respectively; l) SEQ ID NO: 780 and 815, respectively; m) SEQ ID NO: 781 and 816, respectively; n) SEQ ID NO: 782 and 817, respectively; o) SEQ ID NO: 783 and 818, respectively; p) SEQ ID NOs: 784 and 819, respectively; q) SEQ ID NOs: 785 and 820, respectively; r) SEQ ID NOs: 786 and 821, respectively; s) SEQ ID NOs: 787 and 822, respectively; t) SEQ ID NOs: 788 and 823, respectively; u) SEQ ID NOs: 789 and 824, respectively; v) SEQ ID NOs: 790 and 825, respectively; w) SEQ ID NOs: 791 and 826, respectively; x) SEQ ID NOs: 792 and 827, respectively; y) SEQ ID NOs: 793 and 828, respectively; z) SEQ ID NOs: 794 and 829, respectively; aa) SEQ ID NOs: 795 and 830, respectively; bb) SEQ ID NOs: 796 and 831, respectively; cc) SEQ ID NOs: 797 and 832, respectively; dd) SEQ ID NOs: 798 and 833, respectively; ee) SEQ ID NOs: 799 and 834, respectively; ff) SEQ ID NOs: 800 and 835, respectively; gg) SEQ ID NOs: 801 and 836, respectively; hh) SEQ ID NOs: 802 and 837, respectively ii) SEQ ID NOs: 803 and 838, respectively; and jj) a nucleotide sequence selected from the group consisting of SEQ ID NOs: 1681 and 815, respectively, wherein the oligonucleotide comprises a 5' terminal phosphate group, optionally a 5' terminal phosphate analog.

[0169] In some embodiments, the sense and antisense strands of the oligonucleotide are: a) SEQ ID NOs: 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) comprising a nucleotide sequence selected from the group consisting of SEQ ID NOs: 1681 and 815, respectively, wherein the oligonucleotide comprises a 5'-terminal phosphate group, optionally a 5'-terminal phosphate analog.

[0170] In some embodiments, the sense and antisense strands of the oligonucleotide are: a) SEQ ID NOs: 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) comprising a nucleotide sequence selected from the group consisting of SEQ ID NOs: 1681 and 815, respectively, wherein the oligonucleotide comprises a 5'-terminal phosphate group, optionally a 5'-terminal phosphate analog.

[0171] In some embodiments, the oligonucleotide comprises an antisense strand comprising a 4'-phosphate analog at the 5'-terminal nucleotide, wherein the 5'-terminal nucleotide has the following structure:

Chemical formula

[0172] Modified nucleotide linkages

[0173] In some embodiments, the oligonucleotides herein (e.g., RNAi oligonucleotides) contain modified internucleotide linkages. In some embodiments, the modification or substitution of the phosphate results in an oligonucleotide containing at least one (e.g., at least 1, at least 2, at least 3, or at least 5) modified internucleotide linkages. In some embodiments, the oligonucleotide contains from about 1 to about 10 (e.g., 1 - 10, 2 - 8, 4 - 6, 3 - 10, 5 - 10, 1 - 5, 1 - 3, or 1 - 2) modified internucleotide linkages. In some embodiments, the oligonucleotide contains 1, 2, 3, 4, 5, 6, 7, 8, 9, or 10 modified internucleotide linkages.

[0174] The modified internucleotide linkage may be a phosphorodithioate linkage, a phosphorothioate linkage, a phosphotriester linkage, a thionoalkylphosphonate linkage, a thionoalkylphosphotriester linkage, a phosphoramidite linkage, a phosphonic acid linkage, and / or a boranophosphate linkage. In some embodiments, at least one modified internucleotide linkage of the oligonucleotide is a phosphorothioate linkage.

[0175] In some embodiments, the oligonucleotide has a phosphorothioate linkage 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 linkage 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 linkage between (i) positions 1 and 2 of the sense strand; and (ii) each of positions 1 and 2, 2 and 3, 3 and 4, 20 and 21, and 21 and 22 of the antisense strand.

[0176] In some embodiments, the sense and antisense strands of the oligonucleotide are: a) SEQ ID NO: 769 and 804, respectively; b) SEQ ID NO: 770 and 805, respectively; c) SEQ ID NO: 771 and 806, respectively; d) SEQ ID NO: 772 and 807, respectively; e) SEQ ID NO: 773 and 808, respectively; f) SEQ ID NO: 774 and 809, respectively; g) SEQ ID NO: 775 and 810, respectively; h) SEQ ID NO: 776 and 811, respectively; i) SEQ ID NO: 777 and 812, respectively; j) SEQ ID NO: 778 and 813, respectively; k) SEQ ID NO: 779 and 814, respectively; l) SEQ ID NO: 780 and 815, respectively; m) SEQ ID NO: 781 and 816, respectively; n) SEQ ID NO: 782 and 817, respectively; o) SEQ ID NO: 783 and 818, respectively; p) SEQ ID NO: 784 and 819, respectively; q) SEQ ID NO: 785 and 820, respectively; r) SEQ ID NO: 786 and 821, respectively; s) SEQ ID NO: 787 and 822, respectively; t) SEQ ID NO: 788 and 823, respectively; u) SEQ ID NO: 789 and 824, respectively; v) SEQ ID NO: 790 and 825, respectively; w) SEQ ID NO: 791 and 826, respectively; x) SEQ ID NO: 792 and 827, respectively; y) SEQ ID NO: 793 and 828, respectively; z) SEQ ID NO: 794 and 829, respectively; aa) SEQ ID NO: 795 and 830, respectively; bb) SEQ ID NO: 796 and 831, respectively; cc) SEQ ID NO: 797 and 832, respectively; dd) SEQ ID NO: 798 and 833, respectively; ee) nucleotide sequences of SEQ ID NO: 799 and 834, respectively; ff) nucleotide sequences of SEQ ID NO: 800 and 835, respectively; gg) nucleotide sequences of SEQ ID NO: 801 and 836, respectively; hh) nucleotide sequences of SEQ ID NO: 802 and 837, respectively ii) nucleotide sequences of SEQ ID NO: 803 and 838, respectively; and jj) comprising a nucleotide sequence selected from the group consisting of SEQ ID NO: 1681 and 815, respectively, wherein the oligonucleotide comprises modified internucleotide linkages.

[0177] In some embodiments, the sense and antisense strands of the oligonucleotide: a) SEQ ID NO: 771 and 806, respectively; b) SEQ ID NO: 776 and 811, respectively; c) SEQ ID NO: 780 and 815, respectively; d) SEQ ID NO: 781 and 816, respectively; e) SEQ ID NO: 782 and 817, respectively; f) SEQ ID NO: 790 and 825, respectively; g) SEQ ID NO: 795 and 830, respectively; h) SEQ ID NO: 798 and 833, respectively; i) SEQ ID NO: 799 and 834, respectively; j) SEQ ID NO: 803 and 838, respectively; and k) comprising a nucleotide sequence selected from the group consisting of SEQ ID NO: 1681 and 815, respectively, wherein the oligonucleotide comprises modified internucleotide linkages.

[0178] In some embodiments, the sense and antisense strands of the oligonucleotide are: a) SEQ ID NO: 771 and 806, respectively; b) SEQ ID NO: 780 and 815, respectively; c) SEQ ID NO: 781 and 816, respectively; d) SEQ ID NO: 798 and 833, respectively; e) SEQ ID NO: 799 and 834, respectively; f) SEQ ID NO: 803 and 838, respectively; and k) comprising a nucleotide sequence selected from the group consisting of SEQ ID NO: 1681 and 815, respectively, wherein the oligonucleotide comprises modified internucleotide linkages.

[0179] Modification of bases

[0180] In some embodiments, the oligonucleotides herein (e.g., RNAi oligonucleotides) have one or more modified nucleic acid bases. In some embodiments, the modified nucleic acid bases (also referred to herein as base analogs) are linked at the 1'-position of the nucleotide sugar moiety. In certain embodiments, the modified nucleic acid base is a nitrogen-containing base. In certain embodiments, the modified nucleic acid base does not contain a nitrogen atom. See, e.g., U.S. Patent Application Publication No. 2008 / 0274462. In some embodiments, the modified nucleotide comprises a universal base. In some embodiments, the modified nucleotide is base-free (abasic).

[0181] In some embodiments, the sense and antisense strands of the oligonucleotide are: a) SEQ ID NO: 769 and 804, respectively; b) SEQ ID NO: 770 and 805, respectively; c) SEQ ID NO: 771 and 806, respectively; d) SEQ ID NO: 772 and 807, respectively; e) SEQ ID NO: 773 and 808, respectively; f) SEQ ID NO: 774 and 809, respectively; g) SEQ ID NO: 775 and 810, respectively; h) SEQ ID NO: 776 and 811, respectively; i) SEQ ID NO: 777 and 812, respectively; j) SEQ ID NO: 778 and 813, respectively; k) SEQ ID NO: 779 and 814, respectively; l) SEQ ID NO: 780 and 815, respectively; m) SEQ ID NO: 781 and 816, respectively; n) SEQ ID NO: 782 and 817, respectively; o) SEQ ID NOs: 783 and 818, respectively; p) SEQ ID NOs: 784 and 819, respectively; q) SEQ ID NOs: 785 and 820, respectively; r) SEQ ID NOs: 786 and 821, respectively; s) SEQ ID NOs: 787 and 822, respectively; t) SEQ ID NOs: 788 and 823, respectively; u) SEQ ID NOs: 789 and 824, respectively; v) SEQ ID NOs: 790 and 825, respectively; w) SEQ ID NOs: 791 and 826, respectively; x) SEQ ID NOs: 792 and 827, respectively; y) SEQ ID NOs: 793 and 828, respectively; z) SEQ ID NOs: 794 and 829, respectively; aa) SEQ ID NOs: 795 and 830, respectively; bb) SEQ ID NOs: 796 and 831, respectively; cc) SEQ ID NOs: 797 and 832, respectively; dd) SEQ ID NOs: 798 and 833, respectively; ee) SEQ ID NOs: 799 and 834, respectively; ff) SEQ ID NOs: 800 and 835, respectively; gg) SEQ ID NOs: 801 and 836, respectively; hh) SEQ ID NOs: 802 and 837 ii) SEQ ID NOs: 803 and 838, respectively; and jj) a nucleotide sequence selected from the group consisting of SEQ ID NOs: 1681 and 815, respectively, wherein the oligonucleotide comprises one or more modified nucleobases.

[0182] In some embodiments, the sense and antisense strands of the oligonucleotide are: a) SEQ ID NOs: 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) comprising a nucleotide sequence selected from the group consisting of SEQ ID NOs: 1681 and 815, respectively, wherein the oligonucleotide comprises one or more modified nucleobases.

[0183] In some embodiments, the sense and antisense strands of the oligonucleotide are: a) SEQ ID NOs: 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; g) comprising a nucleotide sequence selected from the group consisting of SEQ ID NOs: 1681 and 815, respectively, wherein the oligonucleotide comprises one or more modified nucleobases.

[0184] In some embodiments, the universal base is a heterocyclic moiety located at the 1'-position of the nucleotide sugar moiety of a modified nucleotide, or an equivalent position of a nucleotide sugar moiety substitution, which can be paired with multiple types of bases without significantly altering the double-stranded structure when present in a double strand. In some embodiments, compared to a reference ss nucleic acid (e.g., an oligonucleotide) that is completely complementary to a target nucleic acid, an ss nucleic acid containing a universal base has a lower Tm when forming a double strand with a target nucleic acid having a Tm. m The ss nucleic acid containing a universal base forms a double strand with a target nucleic acid having a Tm that is lower than that of the double strand formed by a complementary nucleic acid. In some embodiments, compared to a reference ss nucleic acid in which a universal base has been replaced with a base to create one mismatch, the ss nucleic acid containing a universal base has a higher Tm when forming a double strand with a nucleic acid containing a mismatched base.m forms a double strand with a target nucleic acid having it.

[0185] Non-limiting examples of universal binding nucleotides include inosine, 1-β-D-ribofuranosyl-5-nitroindole, and / or 1-β-D-ribofuranosyl-3-nitropyrrole, but are not limited thereto (see, for example, 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 & Brown, (1994), Nucleic Acids Res. 22:4039-43).

[0186] Targeting ligand

[0187] In some embodiments, it is desirable to direct an oligonucleotide (e.g., an RNAi oligonucleotide) to one or more cells or one or more organs. Such strategies can help avoid unwanted effects in other organs or avoid more than necessary loss of the oligonucleotide to cells, tissues, or organs that do not benefit from the oligonucleotide. Thus, in some embodiments, the oligonucleotide is modified to promote targeting and / or delivery to a particular tissue, cell, or organ (e.g., to promote delivery of the oligonucleotide to the CNS). 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 (s). In some embodiments, the sense and antisense strands of the oligonucleotide are: a) SEQ ID NOs: 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; l) SEQ ID NOs: 780 and 815, respectively; m) SEQ ID NOs: 781 and 816, respectively; n) SEQ ID NOs: 782 and 817, respectively; o) SEQ ID NOs: 783 and 818, respectively; p) SEQ ID NOs: 784 and 819, respectively; q) SEQ ID NOs: 785 and 820, respectively; r) SEQ ID NOs: 786 and 821, respectively; s) SEQ ID NOs: 787 and 822, respectively; t) SEQ ID NOs: 788 and 823, respectively; u) SEQ ID NOs: 789 and 824, respectively; v) SEQ ID NOs: 790 and 825, respectively; w) SEQ ID NOs: 791 and 826, respectively; x) SEQ ID NOs: 792 and 827, respectively; y) SEQ ID NOs: 793 and 828, respectively; z) SEQ ID NOs: 794 and 829, respectively; aa) SEQ ID NOs: 795 and 830, respectively; bb) SEQ ID NOs: 796 and 831, respectively; cc) SEQ ID NOs: 797 and 832, respectively; dd) SEQ ID NOs: 798 and 833, respectively; ee) SEQ ID NOs: 799 and 834, respectively; ff) SEQ ID NOs: 800 and 835, respectively; gg) SEQ ID NOs: 801 and 836, respectively; hh) SEQ ID NOs: 802 and 837, respectively; and ii) It comprises a nucleotide sequence selected from the group consisting of SEQ ID NO: 803 and SEQ ID NO: 838, wherein the oligonucleotide comprises a targeting ligand attached to at least one nucleotide.

[0188] In some embodiments, the oligonucleotide comprises at least one nucleotide (e.g., 1, 2, 3, 4, 5, 6, or more nucleotides) attached to one or more targeting ligand(s). In some embodiments, the sense and antisense strands of the oligonucleotide are: a) SEQ ID NO: 771 and SEQ ID NO: 806, respectively; b) SEQ ID NO: 776 and SEQ ID NO: 811, respectively; c) SEQ ID NO: 780 and SEQ ID NO: 815, respectively; d) SEQ ID NO: 781 and SEQ ID NO: 816, respectively; e) SEQ ID NO: 782 and SEQ ID NO: 817, respectively; f) SEQ ID NO: 790 and SEQ ID NO: 825, respectively; g) SEQ ID NO: 795 and SEQ ID NO: 830, respectively; h) SEQ ID NO: 798 and SEQ ID NO: 833, respectively; i) SEQ ID NO: 799 and SEQ ID NO: 834, respectively; j) It comprises a nucleotide sequence selected from the group consisting of SEQ ID NO: 803 and SEQ ID NO: 838, wherein the oligonucleotide comprises a targeting ligand attached to at least one nucleotide.

[0189] In some embodiments, the oligonucleotide comprises at least one nucleotide (e.g., 1, 2, 3, 4, 5, 6, or more nucleotides) attached to one or more targeting ligand(s). In some embodiments, the sense and antisense strands of the oligonucleotide are: a) SEQ ID NO: 771 and SEQ ID NO: 806, respectively; b) SEQ ID NO: 780 and SEQ ID NO: 815, respectively; c) SEQ ID NO: 781 and SEQ ID NO: 816, respectively; d) SEQ ID NO: 798 and SEQ ID NO: 833, respectively; e) SEQ ID NO: 799 and SEQ ID NO: 834, respectively; and f) comprising a nucleotide sequence selected from the group consisting of SEQ ID NO: 803 and SEQ ID NO: 838, wherein the oligonucleotide comprises a targeting ligand attached to at least one nucleotide.

[0190] In some embodiments, the targeting ligand comprises a carbohydrate, an amino sugar, cholesterol, a peptide, a polypeptide, or a protein or a portion of a protein (e.g., an antibody or antibody fragment). In some embodiments, the targeting ligand is an aptamer. For example, the targeting ligand may be an RGD peptide used to target tumor vasculature or glioma cells, a CREKA peptide for targeting tumor vasculature or tumor stroma, transferrin, lactoferrin, or an aptamer for targeting the transferrin receptor expressed on the CNS vasculature, or an anti-EGFR antibody for 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.

[0191] In some embodiments, one or more (e.g., 1, 2, 3, 4, 5, or 6) nucleotides of the oligonucleotide are each conjugated to a separate targeting ligand. In some embodiments, 2 to 4 nucleotides of the oligonucleotide are each conjugated to a separate targeting ligand. In some embodiments, the targeting ligand is conjugated to 2 to 4 nucleotides at either end of either the sense or antisense strand (e.g., the targeting ligand is conjugated to an overhang or extension of 2 to 4 nucleotides at the 5' or 3' end of the sense or antisense strand), such that the targeting ligand resembles the bristles of a toothbrush and the oligonucleotide resembles a toothbrush. For example, the oligonucleotide may include a stem-loop at either the 5' or 3' end of the sense strand, and 1, 2, 3, or 4 nucleotides of the loop of the stem may be individually conjugated to the targeting ligand. In some embodiments, the oligonucleotide includes a stem-loop at the 3' end of the sense strand, wherein the loop of the stem-loop includes triL or tetraL, and the 3 or 4 nucleotides that make up triL or tetraL are each individually conjugated to the targeting ligand. In some embodiments, the oligonucleotide includes 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 includes 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.

[0192] GalNAc conjugation

[0193] GalNAc is a high-affinity ligand for the asialoglycoprotein receptor (ASGPR) that is predominantly expressed on the sinusoidal surface of hepatocytes and plays a major role in binding, internalizing, and subsequently excreting circulating glycoproteins (asialoglycoproteins) containing terminal galactose or GalNAc residues. A GalNAc moiety can be attached (either indirectly or directly) to the oligonucleotides herein and used to direct the oligonucleotides to ASGPR expressed on cells. In some embodiments, the oligonucleotide is attached to at least one GalNAc moiety, where the GalNAc moiety directs the oligonucleotide to ASGPR expressed on human liver cells (e.g., human hepatocytes). In some embodiments, the GalNAc moiety directs the oligonucleotide to the liver.

[0194] In some embodiments, the oligonucleotide is directly or indirectly attached to a monovalent GalNAc. In some embodiments, the oligonucleotide is directly or indirectly attached to more than one monovalent GalNAc (i.e., attached to 2, 3, or 4 monovalent GalNAc moieties, usually 3 or 4 monovalent GalNAc moieties). In some embodiments, the oligonucleotide is attached to one or more divalent, trivalent, or tetravalent GalNAc moieties. In some embodiments, the divalent, trivalent, or tetravalent GalNAc moiety is attached to the oligonucleotide via a branched linker. In some embodiments, a monovalent GalNAc moiety is attached to a first nucleotide and a divalent, trivalent, or tetravalent GalNAc moiety is attached to a second nucleotide via a branched linker.

[0195] In some embodiments, one or more (e.g., 1, 2, 3, 4, 5, or 6) nucleotides of the oligonucleotide are each attached to a GalNAc moiety. In some embodiments, 2 to 4 nucleotides of tetraL are each attached to a separate GalNAc. In some embodiments, 1 to 3 nucleotides of triL are each attached to a separate GalNAc. In some embodiments, the targeting ligand is attached to 2 to 4 nucleotides at either the 5' or 3' end of either the sense strand or the antisense strand (e.g., the ligand is attached to an overhang or extension of 2 to 4 nucleotides at the 5' or 3' end of the sense strand or the antisense strand), such that the GalNAc moieties resemble the bristles of a toothbrush and the oligonucleotides resemble a toothbrush. In some embodiments, the GalNAc moiety is attached to a nucleotide of the sense strand. For example, four GalNAc moieties can be attached to the nucleotides of tetraL of the sense strand, with each GalNAc moiety attached to one nucleotide.

[0196] In some embodiments, the oligonucleotide comprises tetraL, where 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 illustrated below (X = heteroatom):

Chemical formula

[0197] In some embodiments, tetraL comprises a monovalent GalNAc moiety attached to any one or more adenine nucleotides of the tetraloop via any linker described herein, as illustrated below (X = heteroatom):

Chemical formula

[0198] In some embodiments, the oligonucleotides herein (e.g., RNAi oligonucleotides) include monovalent GalNAc conjugated to a guanidine nucleotide, referred to as [ademG-GalNAc] or 2'-aminodiethoxymethanol-guanidine-GalNAc, as illustrated below:

Chemical formula

[0199] In some embodiments, the oligonucleotides herein include monovalent GalNAc conjugated to an adenine nucleotide, referred to as [ademA-GalNAc] or 2'-aminodiethoxymethanol-adenine-GalNAc, as illustrated below:

Chemical formula

[0200] An example of such a linkage is shown below for a loop containing the nucleotide sequence GAAA from 5' to 3' (L = linker, X = heteroatom). The stem attachment points are indicated. Such a loop may be present, for example, at positions 27 to 30 of any one of the sense strands listed in Table 4 or 5. In chemical formulas,

Chemical formula

Chemical formula

[0201] A targeting ligand can be linked to a nucleotide using a suitable method or chemical approach (e.g., click chemistry). In some embodiments, the targeting ligand is attached to the nucleotide using a click linker. In some embodiments, an acetal-based linker is used to attach the targeting ligand to any one of the nucleotides of the oligonucleotides described herein. Acetal-based linkers are disclosed, for example, in International Patent Application Publication No. WO2016 / 100401. In some embodiments, the linker is a labile linker. However, in other embodiments, the linker is stable. An example of a loop containing the nucleotides GAAA from 5' to 3' is shown below, in which the GalNAc moiety is attached to the 3rd or 4th nucleotide of the loop using an acetal linker. Such loops may be present, for example, at positions 27 to 30 of any one of the sense strands listed in Table 4 or 5. In chemical formula,

Chemical formula

Chemical formula

Chemical formula

[0202] As described above, a targeting ligand can be linked to a nucleotide using various suitable methods or chemical synthesis methods (e.g., click chemistry). In some embodiments, the targeting ligand is attached to the nucleotide using a click linker. In some embodiments, an acetal-based linker is used to attach the targeting ligand to any one of the nucleotides of the oligonucleotides described herein. Acetal-based linkers are disclosed, for example, in International Patent Application Publication No. WO2016 / 100401. In some embodiments, the linker is a labile linker. However, in other embodiments, the linker is stable.

[0203] In some embodiments, a double-stranded extension portion (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 oligonucleotides herein do not have GalNAc attached thereto.

[0204] In some embodiments, the sense and antisense strands of the oligonucleotide are: a) SEQ ID NO: 769 and 804, respectively; b) SEQ ID NO: 770 and 805, respectively; c) SEQ ID NO: 771 and 806, respectively; d) SEQ ID NO: 772 and 807, respectively; e) SEQ ID NO: 773 and 808, respectively; f) SEQ ID NO: 774 and 809, respectively; g) SEQ ID NO: 775 and 810, respectively; h) SEQ ID NO: 776 and 811, respectively; i) SEQ ID NO: 777 and 812, respectively; j) SEQ ID NO: 778 and 813, respectively; k) SEQ ID NO: 779 and 814, respectively; l) SEQ ID NO: 780 and 815, respectively; m) SEQ ID NO: 781 and 816, respectively; n) SEQ ID NO: 782 and 817, respectively; o) SEQ ID NO: 783 and 818, respectively; p) SEQ ID NO: 784 and 819, respectively; q) SEQ ID NO: 785 and 820, respectively; r) SEQ ID NO: 786 and 821, respectively; s) SEQ ID NO: 787 and 822, respectively; t) SEQ ID NO: 788 and 823, respectively; u) SEQ ID NO: 789 and 824, respectively; v) SEQ ID NO: 790 and 825, respectively; w) SEQ ID NO: 791 and 826, respectively; x) SEQ ID NOs: 792 and 827, respectively; y) SEQ ID NOs: 793 and 828, respectively; z) SEQ ID NOs: 794 and 829, respectively; aa) SEQ ID NOs: 795 and 830, respectively; bb) SEQ ID NOs: 796 and 831, respectively; cc) SEQ ID NOs: 797 and 832, respectively; dd) SEQ ID NOs: 798 and 833, respectively; ee) SEQ ID NOs: 799 and 834, respectively; ff) SEQ ID NOs: 800 and 835, respectively; gg) SEQ ID NOs: 801 and 836, respectively; hh) SEQ ID NOs: 802 and 837, respectively; and ii) a nucleotide sequence selected from the group consisting of SEQ ID NOs: 803 and 838, respectively, wherein the oligonucleotide comprises at least one GalNAc moiety attached to the nucleotide.

[0205] In some embodiments, the sense and antisense strands of the oligonucleotide are: a) SEQ ID NOs: 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) a nucleotide sequence selected from the group consisting of SEQ ID NOs: 803 and 838, respectively, wherein the oligonucleotide comprises at least one GalNAc moiety attached to the nucleotide.

[0206] In some embodiments, the sense and antisense strands of the oligonucleotide are: a) SEQ ID NOs: 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) a nucleotide sequence selected from the group consisting of SEQ ID NOs: 803 and 838, respectively, wherein the oligonucleotide comprises at least one GalNAc moiety attached to a nucleotide.

[0207] Lipid conjugation In some embodiments, one or more lipid moieties are attached to the 5'-terminal nucleotide of the sense strand. In some embodiments, one or more lipid moieties are attached to an adenine nucleotide. In some embodiments, one or more lipid moieties are attached to a guanine nucleotide. In some embodiments, one or more lipid moieties are attached to a cytosine nucleotide. In some embodiments, one or more lipid moieties are attached to a thymine nucleotide. In some embodiments, one or more lipid moieties are attached to a uracil nucleotide.

[0208] 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-chain. In some embodiments, the lipid moiety is C 8 ~C 30 hydrocarbon chain. In some embodiments, the lipid moiety is C 8 :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, C18 :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 a hydrocarbon chain. In some embodiments, C 16 the hydrocarbon chain is represented as follows:

Chemical Structure

[0209] In some embodiments, the sense strand is a nucleotide of length 20 - 22, and the lipid moiety is a hydrocarbon chain attached to the 5'-terminal nucleotide of the sense strand. In some embodiments, the sense strand is a nucleotide of length 20 - 22, and the hydrocarbon chain is attached to the 5'-terminal nucleotide of the sense strand. In some embodiments, the sense strand is a nucleotide of length 20 - 22, and C 14 - C 22 the hydrocarbon chain is attached to the 5'-terminal nucleotide of the sense strand. In some embodiments, the sense strand is a nucleotide of length 20 - 22, and C 16 the hydrocarbon chain is attached to the 5'-terminal nucleotide of the sense strand. In some embodiments, the sense strand is a nucleotide of length 20, and the lipid moiety is attached to the 5'-terminal nucleotide of the sense strand. In some embodiments, the sense strand is a nucleotide of length 20, and the lipid moiety is a hydrocarbon chain attached to the 5'-terminal nucleotide of the sense strand. In some embodiments, the sense strand is a nucleotide of length 20, and C 14 - C 22 the hydrocarbon chain is attached to the 5'-terminal nucleotide of the sense strand. In some embodiments, the sense strand is a nucleotide of length 20, and C 16 the hydrocarbon chain is attached to the 5'-terminal nucleotide of the sense strand.

[0210] In some embodiments, the oligonucleotide comprises: (i) a sense strand that is 20 to 22 nucleotides in length; (ii) an antisense strand that comprises a 3' overhang sequence that is 1 or more nucleotides in length; (iii) a blunt end that constitutes the 3' end of the sense strand; and (iv) a lipid moiety attached to the 5' terminal nucleotide of the sense strand. In some embodiments, the oligonucleotide comprises: (i) a sense strand that is 20 to 22 nucleotides in length; (ii) an antisense strand that comprises a 3' overhang sequence that is 1 or more nucleotides in length; (iii) a blunt end that constitutes the 3' end of the sense strand; and (iv) a hydrocarbon chain attached to the 5' terminal nucleotide of the sense strand. In some embodiments, the oligonucleotide comprises: (i) a sense strand that is 20 to 22 nucleotides in length; (ii) an antisense strand that comprises a 3' overhang sequence that is 1 or more nucleotides in length; (iii) a blunt end that constitutes the 3' end of the sense strand; and (iv) a C 14 ~C 22 hydrocarbon chain attached to the 5' terminal nucleotide of the sense strand. In some embodiments, the oligonucleotide comprises: (i) a sense strand that is 20 to 22 nucleotides in length; (ii) an antisense strand that comprises a 3' overhang sequence that is 1 or more nucleotides in length; (iii) a blunt end that constitutes the 3' end of the sense strand; and (iv) a C 16 hydrocarbon chain attached to the 5' terminal nucleotide of the sense strand.

[0211] In some embodiments, the oligonucleotide comprises (i) a sense strand that is 20 nucleotides in length; (ii) an antisense strand that is 22 nucleotides in length and comprises a 3'-overhang sequence that is 2 nucleotides in length; (iii) a blunt end that constitutes the 3'-end of the sense strand; and (iv) a lipid moiety attached to the 5'-terminal nucleotide of the sense strand. In some embodiments, the oligonucleotide comprises (i) a sense strand that is 20 nucleotides in length; (ii) an antisense strand that is 22 nucleotides in length and comprises a 3'-overhang sequence that is 2 nucleotides in length; (iii) a blunt end that constitutes the 3'-end of the sense strand; and (iv) a hydrocarbon chain attached to the 5'-terminal nucleotide of the sense strand. In some embodiments, the oligonucleotide comprises (i) a sense strand that is 20 nucleotides in length; (ii) an antisense strand that is 22 nucleotides in length and comprises a 3'-overhang sequence that is 2 nucleotides in length; (iii) a blunt end that constitutes the 3'-end of the sense strand; and (iv) a C 14 ~C 22 hydrocarbon chain attached to the 5'-terminal nucleotide of the sense strand. In some embodiments, the oligonucleotide comprises (i) a sense strand that is 20 nucleotides in length; (ii) an antisense strand that is 22 nucleotides in length and comprises a 3'-overhang sequence that is 2 nucleotides in length; (iii) a blunt end that constitutes the 3'-end of the sense strand; and (iv) a lipid moiety attached to the 5'-terminal nucleotide of the sense strand.

[0212] In some embodiments, the oligonucleotide comprises (i) an antisense strand of 19 to 30 nucleotides comprising a region of complementarity to a MAPT mRNA target sequence selected from 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, 1121, 1122, 1123, and 1124; (ii) a sense strand of 19 to 25 nucleotides that forms a double-stranded region with the antisense strand; and (iii) a lipid moiety attached to the 5'-terminal nucleotide of the sense strand. In some embodiments, the oligonucleotide comprises (i) an antisense strand of 19 to 30 nucleotides comprising a region of complementarity to a MAPT mRNA target sequence selected from SEQ ID NOs: 1061, 1108, 1119, 1120, 1124, 1130, 1065, 1095, 1096, and 1102; (ii) a sense strand of 19 to 25 nucleotides that forms a double-stranded region with the antisense strand; and (iii) a lipid moiety attached to the 5'-terminal nucleotide of the sense strand. In some embodiments, the oligonucleotide comprises (i) an antisense strand of 19 to 30 nucleotides comprising a region of complementarity to a MAPT mRNA target sequence selected from SEQ ID NOs: 1130, 1095, 1096, 1119, 1120, and 1124; (ii) a sense strand of 19 to 25 nucleotides that forms a double-stranded region with the antisense strand; and (iii) a lipid moiety attached to the 5'-terminal nucleotide of the sense strand.

[0213] In some embodiments, the oligonucleotide comprises: (i) an antisense strand of 19 to 30 nucleotides comprising a region complementary to a MAPT mRNA target sequence selected from 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, and 1124; (ii) a sense strand of 19 to 25 nucleotides that forms a double-stranded region with the antisense strand; and (iii) a hydrocarbon chain attached to the 5'-terminal nucleotide of the sense strand. In some embodiments, the oligonucleotide comprises: (i) an antisense strand of 19 to 30 nucleotides comprising a region complementary to a MAPT mRNA target sequence selected from SEQ ID NOs: 1061, 1108, 1119, 1120, 1124, 1130, 1065, 1095, 1096, and 1102; (ii) a sense strand of 19 to 25 nucleotides that forms a double-stranded region with the antisense strand; and (iii) a hydrocarbon chain attached to the 5'-terminal nucleotide of the sense strand. In some embodiments, the oligonucleotide comprises: (i) an antisense strand of 19 to 30 nucleotides comprising a region complementary to a MAPT mRNA target sequence selected from SEQ ID NOs: 1130, 1095, 1096, 1119, 1120, and 1124; (ii) a sense strand of 19 to 25 nucleotides that forms a double-stranded region with the antisense strand; and (iii) a hydrocarbon chain attached to the 5'-terminal nucleotide of the sense strand.

[0214] In some embodiments, the oligonucleotide comprises (i) an antisense strand of 19 to 30 nucleotides comprising a region complementary to a MAPT mRNA target sequence selected from 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, and 1124; (ii) a sense strand of 19 to 25 nucleotides that forms a double-stranded region with the antisense strand; and (iii) a C 14 ~C 22 hydrocarbon chain attached to the 5'-terminal nucleotide of the sense strand. In some embodiments, the oligonucleotide comprises (i) an antisense strand of 19 to 30 nucleotides comprising a region complementary to a MAPT mRNA target sequence selected from SEQ ID NOs: 1061, 1108, 1119, 1120, 1124, 1130, 1065, 1095, 1096, and 1102; (ii) a sense strand of 19 to 25 nucleotides that forms a double-stranded region with the antisense strand; and (iii) a C 14 ~C 22 hydrocarbon chain attached to the 5'-terminal nucleotide of the sense strand. In some embodiments, the oligonucleotide comprises (i) an antisense strand of 19 to 30 nucleotides comprising a region complementary to a MAPT mRNA target sequence selected from SEQ ID NOs: 1130, 1095, 1096, 1119, 1120, and 1124; (ii) a sense strand of 19 to 25 nucleotides that forms a double-stranded region with the antisense strand; and (iii) a C 14 ~C 22 hydrocarbon chain attached to the 5'-terminal nucleotide of the sense strand.

[0215] In some embodiments, the oligonucleotide comprises: (i) an antisense strand of 19 to 30 nucleotides comprising a region of complementarity to a MAPT mRNA target sequence selected from 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, and 1124; (ii) a sense strand of 19 to 25 nucleotides that forms a double-stranded region with the antisense strand; and (iii) a C 16 hydrocarbon chain attached to the 5'-terminal nucleotide of the sense strand. In some embodiments, the oligonucleotide comprises: (i) an antisense strand of 19 to 30 nucleotides comprising a region of complementarity to a MAPT mRNA target sequence selected from SEQ ID NOs: 1061, 1108, 1119, 1120, 1124, 1130, 1065, 1095, 1096, and 1102; (ii) a sense strand of 19 to 25 nucleotides that forms a double-stranded region with the antisense strand; and (iii) a C 16 hydrocarbon chain attached to the 5'-terminal nucleotide of the sense strand. In some embodiments, the oligonucleotide comprises: (i) an antisense strand of 19 to 30 nucleotides comprising a region of complementarity to a MAPT mRNA target sequence selected from SEQ ID NOs: 1130, 1095, 1096, 1119, 1120, and 1124; (ii) a sense strand of 19 to 25 nucleotides that forms a double-stranded region with the antisense strand; and (iii) a C 16 hydrocarbon chain attached to the 5'-terminal nucleotide of the sense strand.

[0216] In some embodiments, the oligonucleotide comprises a sense strand comprising the nucleotide sequence of SEQ ID NO: 1681 and an antisense strand comprising the nucleotide sequence of SEQ ID NO: 815, wherein the sense strand comprises a lipid moiety attached to the 5'-terminal nucleotide of the sense strand. In some embodiments, the oligonucleotide comprises a sense strand comprising the nucleotide sequence of SEQ ID NO: 1681 and an antisense strand comprising the nucleotide sequence of SEQ ID NO: 815, wherein the sense strand comprises a hydrocarbon chain attached to the 5'-terminal nucleotide of the sense strand. In some embodiments, the oligonucleotide comprises a sense strand comprising the nucleotide sequence of SEQ ID NO: 1681 and an antisense strand comprising the nucleotide sequence of SEQ ID NO: 815, wherein the sense strand comprises a C 14 ~C 22 hydrocarbon chain attached to the 5'-terminal nucleotide of the sense strand. In some embodiments, the oligonucleotide comprises a sense strand comprising the nucleotide sequence of SEQ ID NO: 1681 and an antisense strand comprising the nucleotide sequence of SEQ ID NO: 815, wherein the sense strand comprises a C 16 hydrocarbon chain attached to the 5'-terminal nucleotide of the sense strand.

[0217] Exemplary RNAi oligonucleotides targeting MAPT

[0218] In some embodiments, the MAPT-targeting RNAi oligonucleotide for reducing the expression of the MAPT gene provided by the present disclosure includes a sense strand and an antisense strand, wherein all nucleotides constituting the sense strand and the antisense strand are modified, and the antisense strand includes a region complementary to any one of the MAPT mRNA target sequences of SEQ ID NOs: 912 to 1295, and the complementary region is a continuous nucleotide sequence of at least 15 in length. In some embodiments, the 5'-terminal nucleotide of the antisense strand includes 4'-O-methylphosphonate-2'-O-methyluridine [MePhosphonate-40-mU] as described herein. In some embodiments, the 5'-terminal nucleotide of the antisense strand includes a phosphorothioate bond. In some embodiments, the antisense strand and the sense strand include one or more 2'-F- and 2'-OMe modified nucleotides and at least one phosphorothioate bond. In some embodiments, the antisense strand includes four phosphorothioate bonds and the sense strand includes one phosphorothioate bond. In some embodiments, the antisense strand includes five phosphorothioate bonds and the sense strand includes one phosphorothioate bond.

[0219] In some embodiments, the oligonucleotide (e.g., RNAi oligonucleotide) includes a sense strand having any one of the sequences of SEQ ID NOs: 912 to 1295 and an antisense strand including a complementary sequence selected from SEQ ID NOs: 1296 to 1679.

[0220] In some embodiments, the oligonucleotide includes a sense strand having any one of the sequences of SEQ ID NOs: 1 to 384 and an antisense strand including a complementary sequence selected from SEQ ID NOs: 385 to 768.

[0221] In some embodiments, the oligonucleotide includes a sense strand having any one of the sequences of SEQ ID NOs: 769 to 803 and an antisense strand including a complementary sequence selected from SEQ ID NOs: 804 to 838.

[0222] In some embodiments, the oligonucleotide comprises a sense strand having any one of the sequences of SEQ ID NOs: 769 to 803 and 1681, and an antisense strand comprising a complementary sequence selected from SEQ ID NOs: 804 to 838.

[0223] In some embodiments, the oligonucleotide comprises a sense strand having any one of the sequences of SEQ ID NOs: 839 to 873, and an antisense strand comprising a complementary sequence selected from SEQ ID NOs: 874 to 908.

[0224] In some embodiments, the oligonucleotide comprises a sense strand having any one of the sequences of SEQ ID NOs: 839 to 873 and 1681, and an antisense strand comprising a complementary sequence selected from SEQ ID NOs: 874 to 908.

[0225] In some embodiments, the oligonucleotide (e.g., an RNAi oligonucleotide) for reducing the expression of the MAPT gene comprises 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; a phosphorothioate bond between positions 1 and 2; and a 36 - nucleotide sense strand; and 2'-F modified nucleotides at positions 2, 3, 4, 5, 7, 10, 14, 16, and 19; 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, 2 and 3, 20 and 21, and 21 and 22; a 5'-terminal nucleotide at position 1 containing a 4'-phosphate analog; and a 22 - nucleotide antisense strand, optionally wherein the 5'-terminal nucleotide comprises 4 - O - monomethylphosphonate - 2'-O - methyluridine [MePhosphonate - 4O - mU]; positions 1 - 20 of the antisense strand form a double - stranded region with positions 1 - 20 of the sense strand, positions 21 - 36 of the sense strand form a stem - loop, positions 27 - 30 form the loop of the stem - loop, optionally positions 27 - 30 contain tetraL, positions 21 and 22 of the antisense strand contain an overhang, and the sense and antisense strands are: a) SEQ ID NO: 769 and 804, respectively; b) SEQ ID NO: 770 and 805, respectively; c) SEQ ID NO: 771 and 806, respectively; d) SEQ ID NO: 772 and 807, respectively; e) SEQ ID NO: 773 and 808, respectively; f) SEQ ID NO: 774 and 809, respectively; g) SEQ ID NO: 775 and 810, respectively; h) SEQ ID NO: 776 and 811, respectively; i) SEQ ID NO: 777 and 812, respectively; j) SEQ ID NO: 778 and 813, respectively; k) SEQ ID NOs: 779 and 814, respectively; l) SEQ ID NOs: 780 and 815, respectively; m) SEQ ID NOs: 781 and 816, respectively; n) SEQ ID NOs: 782 and 817, respectively; o) SEQ ID NOs: 783 and 818, respectively; p) SEQ ID NOs: 784 and 819, respectively; q) SEQ ID NOs: 785 and 820, respectively; r) SEQ ID NOs: 786 and 821, respectively; s) SEQ ID NOs: 787 and 822, respectively; t) SEQ ID NOs: 788 and 823, respectively; u) SEQ ID NOs: 789 and 824, respectively; v) SEQ ID NOs: 790 and 825, respectively; w) SEQ ID NOs: 791 and 826, respectively; x) SEQ ID NOs: 792 and 827, respectively; y) SEQ ID NOs: 793 and 828, respectively; z) SEQ ID NOs: 794 and 829, respectively; aa) SEQ ID NOs: 795 and 830, respectively; bb) SEQ ID NOs: 796 and 831, respectively; cc) SEQ ID NOs: 797 and 832, respectively; dd) SEQ ID NOs: 798 and 833, respectively; ee) SEQ ID NOs: 799 and 834, respectively; ff) SEQ ID NOs: 800 and 835, respectively; gg) SEQ ID NOs: 801 and 836, respectively; hh) SEQ ID NOs: 802 and 837, respectively; and f) comprising a nucleotide sequence selected from the group consisting of SEQ ID NOs: 803 and 838, respectively.

[0226] In some embodiments, the oligonucleotide for reducing the expression of the MAPT gene (e.g., an RNAi oligonucleotide) comprises 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; a phosphorothioate bond between positions 1 and 2; a 36-nucleotide sense strand; and 2'-F modified nucleotides at positions 2, 3, 4, 5, 7, 10, 14, 16 and 19; 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, 2 and 3, 20 and 21, and 21 and 22; a 5'-terminal nucleotide at position 1 containing a 4'-phosphate analog; a 22-nucleotide antisense strand, optionally wherein the 5'-terminal nucleotide comprises 4-O-methylphosphonate-2'-O-methyluridine [MePhosphonate-4O-mU]; positions 1-20 of the antisense strand form a double-stranded region with positions 1-20 of the sense strand, positions 21-36 of the sense strand form a stem-loop, positions 27-30 form the loop of the stem-loop, optionally positions 27-30 contain a tetraloop, positions 21 and 22 of the antisense strand contain an overhang, and the sense and antisense strands are: a) SEQ ID NOs: 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) comprising nucleotide sequences selected from the group consisting of SEQ ID NOs: 803 and 838, respectively.

[0227] In some embodiments, the oligonucleotide (e.g., an RNAi oligonucleotide) for reducing the expression of the MAPT gene comprises 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-linked nucleotides at positions 28, 29, and 30; a phosphorothioate bond between positions 1 and 2; and a 36-nucleotide sense strand; and 2'-F modified nucleotides at positions 2, 3, 4, 5, 7, 10, 14, 16, and 19; 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, 2 and 3, 20 and 21, and 21 and 22; a 5'-terminal nucleotide at position 1 containing a 4'-phosphate analog; and a 22-nucleotide antisense strand, optionally wherein the 5'-terminal nucleotide contains 4-O-methylphosphonate-2'-O-methyluridine [MePhosphonate-4O-mU]; positions 1-20 of the antisense strand form a double-stranded region with positions 1-20 of the sense strand, positions 21-36 of the sense strand form a stem-loop, positions 27-30 form the loop of the stem-loop, optionally positions 27-30 contain a tetraloop, positions 21 and 22 of the antisense strand contain an overhang, and the sense and antisense strands are: a) SEQ ID NOs: 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; and f) nucleotide sequences selected from the group consisting of SEQ ID NOs: 803 and 838, respectively.

[0228] In some embodiments, the oligonucleotide (e.g., RNAi oligonucleotide) for reducing the expression of the MAPT gene comprises 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 C16 hydrocarbon chain attached to the nucleotide at position 1; phosphorothioate bonds between positions 1 and 2, between positions 18 and 19, and between positions 19 and 20; a 20-nucleotide sense strand; and 2'-F modified nucleotides at positions 2, 3, 4, 5, 7, 10, 14, 16 and 19; 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, between positions 2 and 3, between positions 20 and 21, and between positions 21 and 22; a 5'-terminal nucleotide at position 1 containing a 4'-phosphate analog, optionally wherein the 5'-terminal nucleotide contains 4-O-methylphosphonate-2'-O-methyluridine [MePhosphonate-4O-mU]; positions 1-20 of the antisense strand form a double-stranded region with positions 1-20 of the sense strand, positions 21 and 22 of the antisense strand contain overhangs, and the sense strand and the antisense strand each contain the nucleotide sequences of SEQ ID NOs: 1681 and 815, respectively.

[0229] In some embodiments, the oligonucleotide for reducing the expression of the MAPT gene comprises a sense strand comprising the nucleotide sequence set forth in SEQ ID NO: 771 and an antisense strand comprising the nucleotide sequence set forth in SEQ ID NO: 806. In some embodiments, the oligonucleotide for reducing the expression of the MAPT gene comprises a sense strand comprising the nucleotide sequence set forth in SEQ ID NO: 780 and an antisense strand comprising the nucleotide sequence set forth in SEQ ID NO: 815. In some embodiments, the oligonucleotide for reducing the expression of the MAPT gene comprises a sense strand comprising the nucleotide sequence set forth in SEQ ID NO: 781 and an antisense strand comprising the nucleotide sequence set forth in SEQ ID NO: 816. In some embodiments, the oligonucleotide for reducing the expression of the MAPT gene comprises a sense strand comprising the nucleotide sequence set forth in SEQ ID NO: 798 and an antisense strand comprising the nucleotide sequence set forth in SEQ ID NO: 833. In some embodiments, the oligonucleotide for reducing the expression of the MAPT gene comprises a sense strand comprising the nucleotide sequence set forth in SEQ ID NO: 799 and an antisense strand comprising the nucleotide sequence set forth in SEQ ID NO: 834. In some embodiments, the oligonucleotide for reducing the expression of the MAPT gene comprises a sense strand comprising the nucleotide sequence set forth in SEQ ID NO: 803 and an antisense strand comprising the nucleotide sequence set forth in SEQ ID NO: 838. In some embodiments, the oligonucleotide for reducing the expression of the MAPT gene comprises a sense strand comprising the nucleotide sequence set forth in SEQ ID NO: 1681 and an antisense strand comprising the nucleotide sequence set forth in SEQ ID NO: 815.

[0230] In some embodiments, an oligonucleotide (e.g., an RNAi oligonucleotide) for reducing the expression of the MAPT gene comprises: (i) a nucleotide sequence comprising an antisense strand that includes a region complementary to the MAPT mRNA target sequence, the complementary region being 19 to 30 nucleotides in length as set forth in SEQ ID NO: 1514; and (ii) a sense strand that is 19 to 50 nucleotides in length and includes a region complementary to the antisense strand, wherein the antisense strand and the sense strand are separate strands that form an asymmetric double-stranded region having an overhang of 1 to 4 nucleotides at the 3' end of the antisense strand.

[0231] In some embodiments, an oligonucleotide (e.g., an RNAi oligonucleotide) for reducing the expression of the MAPT gene comprises: (i) a nucleotide sequence comprising an antisense strand that includes a region complementary to the MAPT mRNA target sequence, the complementary region being 19 to 30 nucleotides in length as set forth in SEQ ID NO: 1479; and (ii) a sense strand that is 19 to 50 nucleotides in length and includes a region complementary to the antisense strand, wherein the antisense strand and the sense strand are separate strands that form an asymmetric double-stranded region having an overhang of 1 to 4 nucleotides at the 3' end of the antisense strand.

[0232] In some embodiments, an oligonucleotide for reducing the expression of the MAPT gene comprises: (i) a nucleotide sequence comprising an antisense strand that includes a region complementary to the MAPT mRNA target sequence, the complementary region being 19 to 30 nucleotides in length as set forth in SEQ ID NO: 1480; and (ii) a sense strand that is 19 to 50 nucleotides in length and includes a region complementary to the antisense strand, wherein the antisense strand and the sense strand are separate strands that form an asymmetric double-stranded region having an overhang of 1 to 4 nucleotides at the 3' end of the antisense strand.

[0233] In some embodiments, the MAPT-targeting RNAi oligonucleotide for reducing the expression of the MAPT gene comprises: (i) a nucleotide sequence in which the antisense strand contains a region complementary to the MAPT mRNA target sequence, the complementary region being shown in SEQ ID NO: 1503, and being an antisense strand 19 to 30 nucleotides in length; and (ii) a sense strand 19 to 50 nucleotides in length containing a region complementary to the antisense strand, the antisense strand and the sense strand being separate strands that form an asymmetric double-stranded region having an overhang of 1 to 4 nucleotides at the 3'-end of the antisense strand.

[0234] In some embodiments, the oligonucleotide (e.g., RNAi oligonucleotide) for reducing the expression of the MAPT gene comprises: (i) a nucleotide sequence in which the antisense strand contains a region complementary to the MAPT mRNA target sequence, the complementary region being shown in SEQ ID NO: 1504, and being an antisense strand 19 to 30 nucleotides in length; and (ii) a sense strand 19 to 50 nucleotides in length containing a region complementary to the antisense strand, the antisense strand and the sense strand being separate strands that form an asymmetric double-stranded region having an overhang of 1 to 4 nucleotides at the 3'-end of the antisense strand.

[0235] In some embodiments, the oligonucleotide (e.g., RNAi oligonucleotide) for reducing the expression of the MAPT gene comprises: (i) a nucleotide sequence in which the antisense strand contains a region complementary to the MAPT mRNA target sequence, the complementary region being shown in SEQ ID NO: 1508, and being an antisense strand 19 to 30 nucleotides in length; and (ii) a sense strand 19 to 50 nucleotides in length containing a region complementary to the antisense strand, the antisense strand and the sense strand being separate strands that form an asymmetric double-stranded region having an overhang of 1 to 4 nucleotides at the 3'-end of the antisense strand.

[0236] In some embodiments, an oligonucleotide (e.g., an RNAi oligonucleotide) for reducing the expression of the MAPT gene comprises: (i) a nucleotide sequence comprising an antisense strand that includes a region complementary to the MAPT mRNA target sequence, the complementary region being shown in SEQ ID NO: 1514, the antisense strand having a length of 19 to 30 nucleotides; and (ii) a sense strand having a length of 19 to 50 nucleotides that includes a region complementary to the antisense strand and a stem-loop at the 3' end, the stem-loop being defined as S1-L-S2, where S1 is complementary to S2, L forms a loop having a length of 3 to 5 nucleotides between S1 and S2, and the antisense strand and the sense strand are separate strands that form an asymmetric double-stranded region having an overhang of 1 to 4 nucleotides at the 3' end of the antisense strand.

[0237] In some embodiments, an oligonucleotide (e.g., an RNAi oligonucleotide) for reducing the expression of the MAPT gene comprises: (i) a nucleotide sequence comprising an antisense strand that includes a region complementary to the MAPT mRNA target sequence, the complementary region being shown in SEQ ID NO: 1479, the antisense strand having a length of 19 to 30 nucleotides; and (ii) a sense strand having a length of 19 to 50 nucleotides that includes a region complementary to the antisense strand and a stem-loop at the 3' end, the stem-loop being defined as S1-L-S2, where S1 is complementary to S2, L forms a loop having a length of 3 to 5 nucleotides between S1 and S2, and the antisense strand and the sense strand are separate strands that form an asymmetric double-stranded region having an overhang of 1 to 4 nucleotides at the 3' end of the antisense strand.

[0238] In some embodiments, an oligonucleotide (e.g., an RNAi oligonucleotide) for reducing the expression of the MAPT gene comprises: (i) a nucleotide sequence comprising an antisense strand that includes a region complementary to the MAPT mRNA target sequence, the complementary region being shown in SEQ ID NO: 1480, the antisense strand being 19 to 30 nucleotides in length; and (ii) a sense strand that is 19 to 50 nucleotides in length and includes a region complementary to the antisense strand and a 3'-terminal stem-loop, the stem-loop being defined as S1-L-S2, where S1 is complementary to S2, L forms a loop 3 to 5 nucleotides in length between S1 and S2, and the antisense strand and the sense strand are separate strands that form an asymmetric double-stranded region having a 1 to 4 nucleotide overhang at the 3'-terminus of the antisense strand.

[0239] In some embodiments, an oligonucleotide (e.g., an RNAi oligonucleotide) for reducing the expression of the MAPT gene comprises: (i) a nucleotide sequence comprising an antisense strand that includes a region complementary to the MAPT mRNA target sequence, the complementary region being shown in SEQ ID NO: 1503, the antisense strand being 19 to 30 nucleotides in length; and (ii) a sense strand that is 19 to 50 nucleotides in length and includes a region complementary to the antisense strand and a 3'-terminal stem-loop, the stem-loop being defined as S1-L-S2, where S1 is complementary to S2, L forms a loop 3 to 5 nucleotides in length between S1 and S2, and the antisense strand and the sense strand are separate strands that form an asymmetric double-stranded region having a 1 to 4 nucleotide overhang at the 3'-terminus of the antisense strand.

[0240] In some embodiments, an oligonucleotide (e.g., an RNAi oligonucleotide) for reducing the expression of the MAPT gene comprises: (i) a nucleotide sequence comprising an antisense strand that includes a region complementary to the MAPT mRNA target sequence, the complementary region being an antisense strand of 19 to 30 nucleotides shown in SEQ ID NO: 1504; and (ii) a sense strand of 19 to 50 nucleotides that includes a region complementary to the antisense strand and a stem-loop at the 3' end, the stem-loop being defined as S1-L-S2, where S1 is complementary to S2, L forms a loop of 3 to 5 nucleotides between S1 and S2, and the antisense strand and the sense strand are separate strands that form an asymmetric double-stranded region having an overhang of 1 to 4 nucleotides at the 3' end of the antisense strand.

[0241] In some embodiments, an oligonucleotide (e.g., an RNAi oligonucleotide) for reducing the expression of the MAPT gene comprises: (i) a nucleotide sequence comprising an antisense strand that includes a region complementary to the MAPT mRNA target sequence, the complementary region being an antisense strand of 19 to 30 nucleotides shown in SEQ ID NO: 1508; and (ii) a sense strand of 19 to 50 nucleotides that includes a region complementary to the antisense strand and a stem-loop at the 3' end, the stem-loop being defined as S1-L-S2, where S1 is complementary to S2, L forms a loop of 3 to 5 nucleotides between S1 and S2, and the antisense strand and the sense strand are separate strands that form an asymmetric double-stranded region having an overhang of 1 to 4 nucleotides at the 3' end of the antisense strand.

[0242] In some embodiments, an oligonucleotide (e.g., an RNAi oligonucleotide) for reducing the expression of the MAPT gene comprises: (i) a nucleotide sequence comprising an antisense strand that includes a region of complementarity to a MAPT mRNA target sequence, the region of complementarity being 19 to 30 nucleotides in length as set forth in SEQ ID NO: 1479; and (ii) a sense strand that is 19 to 25 nucleotides in length and includes a region of complementarity to the antisense strand, the oligonucleotide includes a blunt end that constitutes the 3' end of the sense strand, and the antisense and sense strands are separate strands that form an asymmetric double-stranded region having an overhang of 1 to 4 nucleotides at the 3' end of the antisense strand.

[0243] In some embodiments, an oligonucleotide (e.g., an RNAi oligonucleotide) for reducing the expression of the MAPT gene comprises: (i) a nucleotide sequence comprising an antisense strand that includes a region of complementarity to a MAPT mRNA target sequence, the region of complementarity being 19 to 30 nucleotides in length as set forth in SEQ ID NO: 1514; and (ii) a sense strand that is 19 to 50 nucleotides in length and includes a region of complementarity to the antisense strand, the region of complementarity to the antisense strand being as set forth in SEQ ID NO: 1130, and the antisense and sense strands are separate strands that form an asymmetric double-stranded region having an overhang of 1 to 4 nucleotides at the 3' end of the antisense strand.

[0244] In some embodiments, an oligonucleotide (e.g., an RNAi oligonucleotide) for reducing the expression of the MAPT gene comprises: (i) a nucleotide sequence comprising an antisense strand that includes a region of complementarity to a MAPT mRNA target sequence, the region of complementarity being 19 to 30 nucleotides in length as set forth in SEQ ID NO: 1479; and (ii) a sense strand that is 19 to 50 nucleotides in length and includes a region of complementarity to the antisense strand, the region of complementarity to the antisense strand being as set forth in SEQ ID NO: 1095, and the antisense and sense strands are separate strands that form an asymmetric double-stranded region having an overhang of 1 to 4 nucleotides at the 3' end of the antisense strand.

[0245] In some embodiments, the oligonucleotide (e.g., an RNAi oligonucleotide) for reducing the expression of the MAPT gene comprises: (i) a nucleotide sequence having an antisense strand that includes a region complementary to the MAPT mRNA target sequence, the complementary region being shown in SEQ ID NO: 1480, and the antisense strand having a length of 19 to 30 nucleotides; and (ii) a sense strand having a length of 19 to 50 nucleotides that includes a region complementary to the antisense strand, the complementary region to the antisense strand being shown in SEQ ID NO: 1096, and the antisense strand and the sense strand being separate strands that form an asymmetric double-stranded region having an overhang of 1 to 4 nucleotides at the 3'-end of the antisense strand.

[0246] In some embodiments, the oligonucleotide (e.g., an RNAi oligonucleotide) for reducing the expression of the MAPT gene comprises: (i) a nucleotide sequence having an antisense strand that includes a region complementary to the MAPT mRNA target sequence, the complementary region being shown in SEQ ID NO: 1503, and the antisense strand having a length of 19 to 30 nucleotides; and (ii) a sense strand having a length of 19 to 50 nucleotides that includes a region complementary to the antisense strand, the complementary region to the antisense strand being shown in SEQ ID NO: 1119, and the antisense strand and the sense strand being separate strands that form an asymmetric double-stranded region having an overhang of 1 to 4 nucleotides at the 3'-end of the antisense strand.

[0247] In some embodiments, an oligonucleotide (e.g., an RNAi oligonucleotide) for reducing the expression of the MAPT gene comprises: (i) an antisense strand comprising a nucleotide sequence having a region complementary to the MAPT mRNA target sequence, the complementary region being shown in SEQ ID NO: 1504, and the antisense strand having a length of 19 to 30 nucleotides; and (ii) a sense strand having a length of 19 to 50 nucleotides and having a region complementary to the antisense strand, the complementary region to the antisense strand being shown in SEQ ID NO: 1120, and the antisense strand and the sense strand being separate strands that form an asymmetric double-stranded region having an overhang of 1 to 4 nucleotides at the 3' end of the antisense strand.

[0248] In some embodiments, an oligonucleotide (e.g., an RNAi oligonucleotide) for reducing the expression of the MAPT gene comprises: (i) an antisense strand comprising a nucleotide sequence having a region complementary to the MAPT mRNA target sequence, the complementary region being shown in SEQ ID NO: 1508, and the antisense strand having a length of 19 to 30 nucleotides; and (ii) a sense strand having a length of 19 to 50 nucleotides and having a region complementary to the antisense strand, the complementary region to the antisense strand being shown in SEQ ID NO: 1124, and the antisense strand and the sense strand being separate strands that form an asymmetric double-stranded region having an overhang of 1 to 4 nucleotides at the 3' end of the antisense strand.

[0249] In some embodiments, an oligonucleotide (e.g., an RNAi oligonucleotide) for reducing the expression of the MAPT gene comprises: (i) a nucleotide sequence comprising an antisense strand that includes a region complementary to the MAPT mRNA target sequence, the complementary region being 19 to 30 nucleotides in length as shown in SEQ ID NO: 1514; and (ii) a sense strand that is 19 to 50 nucleotides in length and includes a region complementary to the antisense strand and a 3'-terminal stem-loop, the region complementary to the antisense strand being shown in SEQ ID NO: 1130, the stem-loop being defined as S1-L-S2, where S1 is complementary to S2 and L forms a loop 3 to 5 nucleotides in length between S1 and S2, and the antisense strand and the sense strand are separate strands that form an asymmetric double-stranded region having an overhang of 1 to 4 nucleotides at the 3'-end of the antisense strand.

[0250] In some embodiments, an oligonucleotide (e.g., an RNAi oligonucleotide) for reducing the expression of the MAPT gene comprises: (i) a nucleotide sequence comprising an antisense strand that includes a region complementary to the MAPT mRNA target sequence, the complementary region being 19 to 30 nucleotides in length as shown in SEQ ID NO: 1479; and (ii) a sense strand that is 19 to 50 nucleotides in length and includes a region complementary to the antisense strand and a 3'-terminal stem-loop, the region complementary to the antisense strand being shown in SEQ ID NO: 1095, the stem-loop being defined as S1-L-S2, where S1 is complementary to S2 and L forms a loop 3 to 5 nucleotides in length between S1 and S2, and the antisense strand and the sense strand are separate strands that form an asymmetric double-stranded region having an overhang of 1 to 4 nucleotides at the 3'-end of the antisense strand.

[0251] In some embodiments, an oligonucleotide (e.g., an RNAi oligonucleotide) for reducing the expression of the MAPT gene comprises: (i) a nucleotide sequence comprising an antisense strand that includes a region complementary to the MAPT mRNA target sequence, the complementary region being shown in SEQ ID NO: 1480, and the antisense strand having a length of 19 to 30 nucleotides; and (ii) a sense strand having a length of 19 to 50 nucleotides that includes a region complementary to the antisense strand and a stem-loop at the 3' end, the region complementary to the antisense strand being shown in SEQ ID NO: 1096, the stem-loop being defined as S1-L-S2, where S1 is complementary to S2, L forms a loop having a length of 3 to 5 nucleotides between S1 and S2, and the antisense strand and the sense strand are separate strands that form an asymmetric double-stranded region having an overhang of 1 to 4 nucleotides at the 3' end of the antisense strand.

[0252] In some embodiments, an oligonucleotide (e.g., an RNAi oligonucleotide) for reducing the expression of the MAPT gene comprises: (i) a nucleotide sequence comprising an antisense strand that includes a region complementary to the MAPT mRNA target sequence, the complementary region being shown in SEQ ID NO: 1503, and the antisense strand having a length of 19 to 30 nucleotides; and (ii) a sense strand having a length of 19 to 50 nucleotides that includes a region complementary to the antisense strand and a stem-loop at the 3' end, the region complementary to the antisense strand being shown in SEQ ID NO: 1119, the stem-loop being defined as S1-L-S2, where S1 is complementary to S2, L forms a loop having a length of 3 to 5 nucleotides between S1 and S2, and the antisense strand and the sense strand are separate strands that form an asymmetric double-stranded region having an overhang of 1 to 4 nucleotides at the 3' end of the antisense strand.

[0253] In some embodiments, an oligonucleotide (e.g., an RNAi oligonucleotide) for reducing the expression of the MAPT gene comprises: (i) a nucleotide sequence comprising an antisense strand that includes a region complementary to the MAPT mRNA target sequence, the complementary region being shown in SEQ ID NO: 1504, an antisense strand of 19 to 30 nucleotides in length; and (ii) a sense strand of 19 to 50 nucleotides in length that includes a region complementary to the antisense strand and a 3'-terminal stem-loop, the region complementary to the antisense strand being shown in SEQ ID NO: 1120, the stem-loop being defined as S1-L-S2, where S1 is complementary to S2, L forms a loop of 3 to 5 nucleotides between S1 and S2, and the antisense strand and the sense strand are separate strands that form an asymmetric double-stranded region having an overhang of 1 to 4 nucleotides at the 3'-end of the antisense strand.

[0254] In some embodiments, an oligonucleotide (e.g., an RNAi oligonucleotide) for reducing the expression of the MAPT gene comprises: (i) a nucleotide sequence comprising an antisense strand that includes a region complementary to the MAPT mRNA target sequence, the complementary region being shown in SEQ ID NO: 1508, an antisense strand of 19 to 30 nucleotides in length; and (ii) a sense strand of 19 to 50 nucleotides in length that includes a region complementary to the antisense strand and a 3'-terminal stem-loop, the region complementary to the antisense strand being shown in SEQ ID NO: 1124, the stem-loop being defined as S1-L-S2, where S1 is complementary to S2, L forms a loop of 3 to 5 nucleotides between S1 and S2, and the antisense strand and the sense strand are separate strands that form an asymmetric double-stranded region having an overhang of 1 to 4 nucleotides at the 3'-end of the antisense strand.

[0255] In some embodiments, an oligonucleotide (e.g., an RNAi oligonucleotide) for reducing the expression of the MAPT gene comprises: (i) a nucleotide sequence comprising an antisense strand that includes a region of complementarity to a MAPT mRNA target sequence, the region of complementarity being 19 to 30 nucleotides in length as set forth in SEQ ID NO: 1479; and (ii) a sense strand that is 19 to 25 nucleotides in length and includes a region of complementarity to the antisense strand, the region of complementarity to the antisense strand being as set forth in SEQ ID NO: 1095, the oligonucleotide includes a blunt end that constitutes the 3' end of the sense strand, and the antisense strand and the sense strand are separate strands that form an asymmetric double-stranded region having an overhang of 1 to 4 nucleotides at the 3' end of the antisense strand.

[0256] In some embodiments, an oligonucleotide (e.g., an RNAi oligonucleotide) for reducing the expression of the MAPT gene comprises a sense strand and an antisense strand according to the following: Antisense strand: 5’-[MePhosphonate-4O-mX]-S-fX-S-fX-fX-fX-mX-fX-mX-mX-fX-mX-mX-mX-fX-mX-fX-mX-mX-fX-mX-S-mX-S-mX-3’ hybridized to 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-GalNAc]-[ademX-GalNAc]-[ademX-GalNAc]-mX-mX-mX-mX-mX-mX-3’, wherein 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 conjugated to the nucleotide.

[0257] In some embodiments, oligonucleotides (e.g., RNAi oligonucleotides) for reducing the expression of the MAPT gene comprise a sense strand and an antisense strand according to the following: 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-S-mX-S-mX-3’ hybridized to 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-[ademX-GalNAc]-[ademX-GalNAc]-[ademX-GalNAc]-mX-mX-mX-mX-mG-mX-3’, wherein 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 conjugated to a nucleotide.

[0258] In some embodiments, oligonucleotides (e.g., RNAi oligonucleotides) for reducing the expression of the MAPT gene comprise a sense strand and an antisense strand according to the following: Antisense strand: 5’-[MePhosphonate-4O-mX]-S-fX-S-fX-fX-fX-mX-fX-mX-mX-fX-mX-mX-mX-fX-mX-fX-mX-mX-fX-mX-S-mX-S-mX-3’ hybridized to sense strand: 5’-[AdemX-L]-S-mX-fX-mX-fX-mX-mX-fX-mX-fX-mX-fX-fX-mX-fX-mX-fX-mX-S-mX-S-mX-3’, In the formula, 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 is a lipid moiety attached to the nucleotide.

[0259] In some embodiments, an oligonucleotide (e.g., an RNAi oligonucleotide) for reducing the expression of the MAPT gene comprises a sense strand and an antisense strand according to: Antisense strand: 5'-[MePhosphonate-4O-mX]-S-fX-S-fX-fX-fX-mX-fX-mX-mX-fX-mX-mX-mX-fX-mX-fX-mX-mX-fX-mX-S-mX-S-mX-3' and sense strand hybridized therewith: 5'-[AdemX-C16]-S-mX-fX-mX-fX-mX-mX-fX-mX-fX-mX-fX-fX-mX-fX-mX-fX-mX-S-mX-S-mX-3', In the formula, 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 is a C 16 hydrocarbon chain.

[0260] In some embodiments, the present disclosure provides an oligonucleotide (e.g., an RNAi oligonucleotide) for reducing the expression of the MAPT gene, wherein the oligonucleotide is: a) SEQ ID NOs: 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) SEQ ID NOs: 846 and 881, respectively; i) SEQ ID NOs: 847 and 882, respectively; j) SEQ ID NOs: 848 and 883, respectively; k) SEQ ID NOs: 849 and 884, respectively; l) SEQ ID NOs: 850 and 885, respectively; m) SEQ ID NOs: 851 and 886, respectively; n) SEQ ID NOs: 852 and 887, respectively; o) SEQ ID NOs: 853 and 888, respectively; p) SEQ ID NOs: 854 and 889, respectively; q) SEQ ID NOs: 855 and 890, respectively; r) SEQ ID NOs: 856 and 891, respectively; s) SEQ ID NOs: 857 and 892, respectively; t) SEQ ID NOs: 858 and 893, respectively; u) SEQ ID NOs: 859 and 894, respectively; v) SEQ ID NOs: 860 and 895, respectively; w) SEQ ID NOs: 861 and 896, respectively; x) SEQ ID NOs: 862 and 897, respectively; y) SEQ ID NOs: 863 and 898, respectively; z) SEQ ID NOs: 864 and 899, respectively; aa) SEQ ID NOs: 865 and 900, respectively; bb) SEQ ID NOs: 866 and 901, respectively; cc) SEQ ID NOs: 867 and 902, respectively; dd) SEQ ID NOs: 868 and 903, respectively; ee) SEQ ID NOs: 869 and 904, respectively; ff) SEQ ID NOs: 870 and 905, respectively; gg) SEQ ID NOs: 871 and 906, respectively; hh) SEQ ID NOs: 872 and 907, respectively; ii) SEQ ID NOs: 873 and 908, respectively; and jj) comprising a sense strand and an antisense strand comprising a nucleotide sequence selected from the group consisting of SEQ ID NOs: 1682 and 885, respectively.

[0261] In some embodiments, the oligonucleotides (e.g., RNAi oligonucleotides) for reducing the expression of the MAPT gene are: a) SEQ ID NO: 860 and 895, respectively; b) SEQ ID NO: 865 and 900, respectively; c) SEQ ID NO: 868 and 903, respectively; d) SEQ ID NO: 869 and 904, respectively; e) SEQ ID NO: 873 and 908, respectively; f) SEQ ID NO: 841 and 876, respectively; g) SEQ ID NO: 846 and 881, respectively; h) SEQ ID NO: 850 and 885, respectively; i) SEQ ID NO: 851 and 886, respectively; j) SEQ ID NO: 852 and 887, respectively; and k) comprising a sense strand and an antisense strand comprising nucleotide sequences selected from the group consisting of SEQ ID NO: 1682 and 885, respectively.

[0262] In some embodiments, the oligonucleotides (e.g., RNAi oligonucleotides) for reducing the expression of the MAPT gene are: a) SEQ ID NO: 841 and 876, respectively; b) SEQ ID NO: 850 and 885, respectively; c) SEQ ID NO: 851 and 886, respectively; d) SEQ ID NO: 868 and 903, respectively; e) SEQ ID NO: 869 and 904, respectively; f) SEQ ID NO: 873 and 908, respectively; and g) comprising a sense strand and an antisense strand comprising nucleotide sequences selected from the group consisting of SEQ ID NO: 1682 and 885, respectively.

[0263] In some embodiments, the oligonucleotide for reducing the expression of the MAPT gene comprises a sense strand comprising the nucleotide sequence shown in SEQ ID NO: 841 and an antisense strand comprising the nucleotide sequence shown in SEQ ID NO: 876. In some embodiments, the oligonucleotide for reducing the expression of the MAPT gene comprises a sense strand comprising the nucleotide sequence shown in SEQ ID NO: 850 and an antisense strand comprising the nucleotide sequence shown in SEQ ID NO: 885. In some embodiments, the oligonucleotide for reducing the expression of the MAPT gene comprises a sense strand comprising the nucleotide sequence shown in SEQ ID NO: 851 and an antisense strand comprising the nucleotide sequence shown in SEQ ID NO: 886. In some embodiments, the oligonucleotide for reducing the expression of the MAPT gene comprises a sense strand comprising the nucleotide sequence shown in SEQ ID NO: 868 and an antisense strand comprising the nucleotide sequence shown in SEQ ID NO: 903. In some embodiments, the oligonucleotide for reducing the expression of the MAPT gene comprises a sense strand comprising the nucleotide sequence shown in SEQ ID NO: 869 and an antisense strand comprising the nucleotide sequence shown in SEQ ID NO: 904. In some embodiments, the oligonucleotide for reducing the expression of the MAPT gene comprises a sense strand comprising the nucleotide sequence shown in SEQ ID NO: 873 and an antisense strand comprising the nucleotide sequence shown in SEQ ID NO: 908. In some embodiments, the oligonucleotide for reducing the expression of the MAPT gene comprises a sense strand comprising the nucleotide sequence shown in SEQ ID NO: 1682 and an antisense strand comprising the nucleotide sequence shown in SEQ ID NO: 885.

[0264] Formulation

[0265] Various formulations have been developed to facilitate the use of oligonucleotides. For example, oligonucleotides (e.g., RNAi oligonucleotides) can be delivered to a subject or cell environment using formulations that minimize degradation, promote delivery and / or uptake, or impart other beneficial properties to the oligonucleotides in the formulation. In some embodiments, provided herein are compositions comprising oligonucleotides that reduce the expression of the MAPT gene. Such compositions can be suitably formulated so that when administered to a subject either in the adjacent environment of the target cells or systemically, a sufficient portion of the oligonucleotide enters the cells to reduce the expression of the MAPT gene. Any of a variety of suitable oligonucleotide formulations can be used to deliver the oligonucleotides for reducing the expression of the MART gene disclosed herein. In some embodiments, the oligonucleotides are formulated in buffers such as phosphate buffered saline, liposomes, micellar structures, and capsids. In some embodiments, the oligonucleotides are formulated in a buffer such as phosphate buffered saline.

[0266] The use of formulations of oligonucleotides containing cationic lipids can promote the transfection of oligonucleotides into cells. For example, cationic lipids such as lipofectin, cationic glycerol derivatives, and polycationic molecules (e.g., polylysine) can be used. Suitable lipids include oligofectamine, lipofectamine (Life Technologies), NC388 (Ribozyme Pharmaceuticals, Inc., Boulder, Colo.), or FuGene6 (Roche), all of which can be used according to the manufacturer's instructions. In some embodiments, the oligonucleotides are not formulated with components that promote transfection into cells.

[0267] Accordingly, in some embodiments, the formulation comprises lipid nanoparticles. In some embodiments, the excipient comprises liposomes, lipids, complex lipids, microspheres, microparticles, nanospheres, or nanoparticles, or may be formulated in other forms for administration to the cells, tissues, organs, or body of a subject in need thereof (see, e.g., Remington: The Science and Practice of Pharmacy, 22nd edition, Pharmaceutical Press, 2013).

[0268] In some embodiments, the formulations herein comprise an excipient. In some embodiments, the excipient imparts improved stability, improved absorbability, improved solubility, and / or a therapeutic enhancing effect to the composition of the active ingredient. 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 buffer solution, petrolatum, dimethyl sulfoxide, or mineral oil). In some embodiments, the oligonucleotide is lyophilized to extend its shelf life and then made into a solution prior to use (e.g., administration to a subject). Thus, the excipient in a composition comprising any one of the oligonucleotides described herein may be a lyoprotectant (e.g., mannitol, lactose, polyethylene glycol, or polyvinylpyrrolidone), or a collapse temperature modifier (e.g., dextran, Ficoll™, or gelatin).

[0269] In some embodiments, the pharmaceutical composition is formulated to be compatible with its intended route of administration. Examples of routes of administration include parenteral administration (e.g., intravenous, intramuscular, intraperitoneal, intradermal, and subcutaneous), oral administration (e.g., inhalation), transdermal administration (e.g., topical), transmucosal administration, and rectal administration.

[0270] In some embodiments, the pharmaceutical composition is formulated for administration to the central nervous system. In some embodiments, the pharmaceutical composition is formulated for administration to 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 administration into the cerebral ventricle. In some embodiments, the pharmaceutical composition is formulated for the brainstem. In some embodiments, the pharmaceutical composition is formulated for intrasaccular magna administration.

[0271] Pharmaceutical compositions suitable for injection include sterile aqueous solutions (where water-soluble), dispersions, and sterile powders for the immediate preparation of sterile injectable solutions or dispersions. In the case of intravenous administration, suitable carriers include saline, bacteriostatic water, Cremophor EL™ (BASF), or phosphate buffered saline (PBS). The carrier can be a solvent or dispersion medium containing, for example, water, ethanol, polyols (such as glycerol, propylene glycol, liquid polyethylene glycol, etc.), and suitable mixtures thereof. In many cases, it will be preferable to include in the composition isotonic agents such as sugars, polyhydric alcohols like mannitol, sorbitol, and / or sodium chloride. Sterile injectable solutions can be prepared by incorporating the oligonucleotide in the required amount into a selected solvent, along with one or a combination of the ingredients enumerated above as required, and then filtering the solution to sterilize it.

[0272] In some embodiments, the proportion of the active ingredient(s) may be about 1% to about 80% by weight or volume of the total composition, but the composition may contain at least about 0.1% or more of a therapeutic agent (e.g., an RNAi oligonucleotide for reducing the expression of the MAPT gene). Factors such as solubility, bioavailability, biological half-life, route of administration, product shelf-life, as well as other pharmacological considerations are contemplated by those skilled in the art in the field of preparation of such pharmaceutical formulations, and thus, various dosages and therapeutic dosing regimens may be desirable.

[0273] Usage method

[0274] Reducing MAPT gene expression

[0275] In some embodiments, the present disclosure provides a method of contacting or delivering to a cell or population of cells an effective amount of any of the oligonucleotides (e.g., RNAi oligonucleotides) herein for reducing the expression of the MAPT gene. In some embodiments, a reduction in MAPT gene expression is determined by measuring a decrease in the amount or level of MAPT mRNA, tau protein, or tau activity in the cell. Methods include those described herein and known to those of skill in the art.

[0276] In some embodiments, the present disclosure provides a method of reducing the expression of the MAPT gene in the CNS. In some embodiments, the CNS includes the brain and spinal cord. In some embodiments, the expression of the MAPT gene is reduced in at least one region of the brain. In some embodiments, the regions of the brain are the cervical spinal cord, thoracic spinal cord, lumbar spinal cord, prefrontal cortex, temporal cortex, cerebellum, midbrain, occipital cortex, parietal cortex, hippocampus, caudate nucleus, thalamus, and brainstem. In some embodiments, the expression of the MAPT gene is reduced in at least one region of the spinal cord. In some embodiments, the regions of the spinal cord include the cervical spinal cord, thoracic spinal cord, and lumbar spinal cord. In some embodiments, the expression of the MAPT gene is reduced in at least one region of the brain and at least one region of the spinal cord. In some aspects, the expression of the MAPT gene is reduced in at least one of the cervical spinal cord, thoracic spinal cord, lumbar spinal cord, prefrontal cortex, temporal cortex, cerebellum, midbrain, occipital cortex, parietal cortex, hippocampus, caudate nucleus, thalamus, brainstem, motor cortex, globus pallidus, tegmentum, substantia nigra, pons, cerebellar white matter, and dentate nucleus of the cerebellum. In some embodiments, the expression of the MAPT gene is reduced in at least one of the lumbar spinal cord, thoracic spinal cord, and cervical spinal cord. In some embodiments, the expression of the MAPT gene is reduced in the brain and / or spinal cord tissue associated with Alzheimer's disease. In some embodiments, the tissues associated with AD include, but are not limited to, the prefrontal cortex, motor cortex, temporal cortex, parietal cortex, and hippocampus. In some embodiments, the expression of the MAPT gene is reduced in the brain and / or spinal cord tissue associated with progressive supranuclear palsy. In some embodiments, the tissues associated with AD include, but are not limited to, the caudate nucleus, globus pallidus, thalamus, tegmentum, substantia nigra, pons, cerebellar white matter, dentate nucleus of the cerebellum, medulla, cervical spinal cord, thoracic spinal cord, and lumbar spinal cord.

[0277] In some embodiments, the expression of the MAPT gene decreases about 1 week to about 12 weeks after administration of the oligonucleotides described herein. In some embodiments, the expression of the MAPT gene decreases 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, or 12 weeks after administration of the oligonucleotides described herein. In some embodiments, the expression of the MAPT gene decreases about 1 to about 4 months after administration of the oligonucleotides described herein. In some embodiments, the expression of the MAPT gene decreases about 1 to about 6 months after administration of the oligonucleotides described herein. In some embodiments, the expression of the MAPT gene decreases 1, 2, 3, or 4 months after administration of the oligonucleotides described herein. In some embodiments, the expression of the MAPT gene decreases 1, 2, 3, 4, 5, or 6 months after administration of the oligonucleotides described herein. In some embodiments, the expression of the MAPT gene decreases about 7 to about 91 days after administration of the oligonucleotides described herein. In some embodiments, the expression of the MAPT gene decreases 7, 14, 21, 28, 35, 42, 49, 56, 63, 70, 77, 84, or 91 days after administration of the oligonucleotides described herein.

[0278] In some embodiments, the expression of the MAPT gene is reduced in at least one region of the brain and / or at least one region of the spinal cord about 1 to about 12 weeks after administration of the oligonucleotides described herein. In some embodiments, the expression of the MAPT gene is reduced in at least one region of the brain and / or at least one region of the spinal cord 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, or 12 weeks after administration of the oligonucleotides described herein. In some embodiments, the expression of the MAPT gene is reduced in at least one region of the brain and / or at least one region of the spinal cord about 1 to about 4 months after administration of the oligonucleotides described herein. In some embodiments, the expression of the MAPT gene is reduced in at least one region of the brain and / or at least one region of the spinal cord about 1 to about 6 months after administration of the oligonucleotides described herein. In some embodiments, the expression of the MAPT gene is reduced in at least one region of the brain and / or at least one region of the spinal cord 1, 2, 3, or 4 months after administration of the oligonucleotides described herein. In some embodiments, the expression of the MAPT gene is reduced in at least one region of the brain and / or at least one region of the spinal cord 1, 2, 3, 4, 5, or 6 months after administration of the oligonucleotides described herein. In some embodiments, the expression of the MAPT gene is reduced in at least one region of the brain and / or at least one region of the spinal cord about 7 to about 91 days after administration of the oligonucleotides described herein. In some embodiments, the expression of the MAPT gene is reduced in at least one region of the brain and / or at least one region of the spinal cord 7, 14, 21, 28, 35, 42, 49, 56, 63, 70, 77, 84, or 91 days after administration of the oligonucleotides described herein.

[0279] The methods provided herein are useful in any suitable cell type. In some embodiments, the cells are any cells that express MAPT mRNA (e.g., oligodendrocytes). In some embodiments, the cells are primary cells obtained from a subject. In some embodiments, the primary cells have undergone a limited number of passages such that the cells maintain substantially their native phenotypic characteristics. In some embodiments, the cells to which the oligonucleotide is delivered are ex vivo or in vitro (i.e., cells in culture or capable of being delivered to an organism in which the cells are present).

[0280] In some embodiments, the oligonucleotides disclosed herein are delivered to a cell or cell population using nucleic acid delivery methods known in the art, including, but not limited to, injection of a solution or pharmaceutical composition containing the oligonucleotide, particle bombardment with particles coated with the oligonucleotide, exposure of a cell or cell population to a solution containing the oligonucleotide, or electroporation of the cell membrane in the presence of the oligonucleotide. Other methods known in the art for delivering oligonucleotides to cells, such as lipid-mediated carrier transport, chemical-mediated transport, cationic liposome transfection such as calcium phosphate, may also be used.

[0281] In some embodiments, the reduction in MAPT gene expression is determined by an assay or technique that evaluates one or more molecules, properties, or characteristics of a cell or cell population associated with MAPT gene expression, or by an assay or technique that evaluates a molecule that directly indicates MAPT gene expression in a cell or cell population (e.g., MAPT mRNA or tau protein). In some embodiments, the degree to which an oligonucleotide reduces MAPT gene expression is evaluated by comparing the MAPT gene expression in a cell or cell population contacted with the oligonucleotide to a control cell or cell population (e.g., a cell or cell population not contacted with the oligonucleotide or contacted with a control oligonucleotide). In some embodiments, since the control amount or level of MAPT gene expression in the control cell or cell population is predetermined, it is not necessary to measure the control amount or level in every case where the assay or technique is performed. The predetermined level or value can be in various forms. In some embodiments, the predetermined level or value can be a single cut-off value such as a median or an average value.

[0282] In some embodiments, reducing the expression of the MAPT gene is effected by contacting or delivering an oligonucleotide to a cell or cell population. In some embodiments, the reduction in the expression of the MAPT gene is relative to a control amount or level of expression of the MAPT gene in a cell or cell population that is not contacted with the oligonucleotide or is contacted with a control oligonucleotide. In some embodiments, the reduction in the expression of the MAPT gene is about 1% or less, about 5% or less, about 10% or less, about 15% or less, about 20% or less, about 25% or less, about 30% or less, about 35% or less, about 40% or less, about 45% or less, about 50% or less, about 55% or less, about 60% or less, about 70% or less, about 80% or less, or about 90% or less compared to a control amount or level of expression of the MAPT gene. In some embodiments, the control amount or level of expression of the MAPT gene is the amount or level of MAPT mRNA and / or tau protein in a cell or cell population that is not in contact with the oligonucleotides herein. In some embodiments, the effect of delivery of the oligonucleotide to a cell or cell population by the methods herein is evaluated after any finite period or time (e.g., minutes, hours, days, weeks, months). For example, in some embodiments, after contacting or delivering the oligonucleotide to a cell or cell population, 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 more have elapsed, the expression of the MAPT gene is measured in the cell or cell population. In some embodiments, the expression of the MAPT gene is measured 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.

[0283] In some embodiments, the oligonucleotide is delivered in the form of a transgene engineered to express the oligonucleotide or the strand(s) that make up the oligonucleotide (e.g., its sense and antisense strands) intracellularly. In some embodiments, the oligonucleotide is delivered using a transgene engineered to express any of the oligonucleotides disclosed herein. The transgene may be delivered using a viral vector (e.g., an adenovirus, retrovirus, vaccinia virus, poxvirus, adeno-associated virus, or herpes simplex virus), or a non-viral vector (e.g., a plasmid or synthetic mRNA). In some embodiments, the transgene can be directly injected into a subject.

[0284] Therapeutic methods

[0285] The present disclosure also provides oligonucleotides (e.g., RNAi oligonucleotides) for use in treating or being adapted for use in treating a subject (e.g., a human having a disease, disorder, or condition associated with MAPT gene expression) that would benefit from a decrease in MAPT gene expression. In some aspects, the present disclosure provides oligonucleotides for use in treating or being adapted for use in treating a subject having a disease, disorder, or condition associated with MAPT gene expression. The present disclosure also provides oligonucleotides for use in or being adapted for use in the manufacture of a drug or pharmaceutical composition for treating a disease, disorder, or condition associated with MAPT gene expression. In some embodiments, the oligonucleotide for use or being adapted for use targets MAPT mRNA and decreases the expression of the MAPT gene (e.g., via the RNAi pathway). In some embodiments, the oligonucleotide for use or being adapted for use targets MAPT mRNA and decreases the amount or level of MAPT mRNA, tau protein, and / or tau activity.

[0286] Furthermore, in some embodiments of the methods herein, a subject having or susceptible to a disease, disorder, or condition associated with MAPT gene expression is selected for treatment with the oligonucleotides (e.g., ds oligonucleotides) herein. In some embodiments, the method includes selecting an individual having a marker (e.g., a biomarker) of a disease, disorder, or condition associated with MAPT gene expression, such as, but not limited to, MAPT mRNA, tau protein, or a combination thereof, or an individual having a predisposition thereto. Similarly, as detailed below, some embodiments of the methods provided by the present disclosure measure or obtain a baseline value of a marker of MAPT gene expression (e.g., tau protein or tau activity), and then compare such obtained value to one or more other baseline values or values obtained after administering the oligonucleotide to the subject to evaluate the effectiveness of the treatment.

[0287] The present disclosure also provides methods of treating a subject having, suspected of having, or at risk of developing a disease, disorder, or condition associated with MAPT gene expression with the oligonucleotides provided herein. In some aspects, the present disclosure provides methods of treating, or attenuating the onset or progression of, a disease, disorder, or condition associated with MAPT gene expression using the oligonucleotides provided herein. In other aspects, the present disclosure provides methods of achieving one or more therapeutic effects in a subject having a disease, disorder, or condition associated with MAPT gene expression using the oligonucleotides provided herein. In some embodiments of the methods herein, the subject is treated by administering a therapeutically effective amount of any one or more of the oligonucleotides provided herein. In some embodiments, the treatment includes reducing the expression of the MAPT gene. In some embodiments, the individual is treated therapeutically. In some embodiments, the individual is treated prophylactically.

[0288] In some embodiments of the methods herein, an oligonucleotide (e.g., an RNAi oligonucleotide), or a pharmaceutical composition comprising an oligonucleotide, is administered to a subject having a disease, disorder or condition associated with the expression of the MAPT gene such that the expression of the MAPT gene is decreased in the subject, thereby treating the subject. In some embodiments, the amount or level of MAPT mRNA is decreased in the subject. In some embodiments, the amount or level of tau protein is decreased in the subject.

[0289] In some embodiments of the methods herein, an oligonucleotide or a pharmaceutical composition comprising an oligonucleotide is administered to a subject having a disease, disorder, or condition associated with MAPT gene expression such that the expression of the MAPT gene is reduced in the subject 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 more than 99% compared to the MAPT gene expression prior to administration of the oligonucleotide or pharmaceutical composition. In some embodiments of the methods herein, an oligonucleotide or a pharmaceutical composition comprising an oligonucleotide is administered to a subject having a disease, disorder, or condition associated with MAPT gene expression for about 1 week to about 12 weeks, about 1 month to about 6 months, or about 7 days to about 91 days such that the MAPT gene expression is reduced in the subject 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 more than 99% compared to the MAPT gene expression prior to administration of the oligonucleotide or pharmaceutical composition. In some embodiments, the expression of the MAPT gene is reduced in the subject 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 more than 99% compared to the expression of the MAPT gene in a subject not administered the oligonucleotide or pharmaceutical composition or receiving a control oligonucleotide, pharmaceutical composition, or treatment (e.g., a reference subject or control subject). In some embodiments, the expression of the MAPT gene is reduced in the subject 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 more 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 in a subject not administered the oligonucleotide or pharmaceutical composition or receiving a control oligonucleotide, pharmaceutical composition, or treatment (e.g., a reference subject or control subject).

[0290] In some embodiments of the methods herein, the amount or level of MAPT mRNA in a subject is 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 more than 99% lower than the amount or level of MAPT mRNA before administration of the oligonucleotide or pharmaceutical composition, and the oligonucleotide or pharmaceutical composition is administered to a subject having a disease, disorder, or condition associated with MAPT gene expression. In some embodiments of the methods, the amount or level of MAPT mRNA in a subject is 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 more than 99% lower than the amount or level of MAPT mRNA before administration of the oligonucleotide or pharmaceutical composition for about 1 week to about 12 weeks, about 1 month to about 6 months, or about 7 days to about 91 days, and the oligonucleotide or pharmaceutical composition is administered to a subject having a disease, disorder, or condition associated with MAPT gene expression. In some embodiments, the amount or level of MAPT mRNA in a subject is 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 more than 99% lower than the amount or level of MAPT mRNA in a subject not administered the oligonucleotide or pharmaceutical composition or a subject receiving a control oligonucleotide, pharmaceutical composition, or treatment (reference or control subject). In some embodiments, the amount or level of MAPT mRNA in a subject is 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 more than 99% lower than the amount or level of MAPT mRNA in a subject not administered the oligonucleotide or pharmaceutical composition or a subject receiving a control oligonucleotide, pharmaceutical composition, or treatment (reference subject or control subject) for about 1 week to about 12 weeks, about 1 month to about 6 months, or about 7 days to about 91 days.

[0291] In some embodiments of the methods, the amount or level of tau protein in a subject is 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 more than 99% lower than the amount or level of tau protein before administration of the oligonucleotide or pharmaceutical composition to the subject. The oligonucleotide or pharmaceutical composition is administered to a subject having a disease, disorder, or condition associated with MAPT gene expression. In some embodiments of the methods herein, the amount or level of tau protein in the subject is 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 more than 99% lower than the amount or level of tau protein before administration of the oligonucleotide or pharmaceutical composition, for about 1 week to about 12 weeks, about 1 month to about 6 months, or about 7 days to about 91 days in the subject. The oligonucleotide or pharmaceutical composition comprising the oligonucleotide herein is administered to a subject having a disease, disorder, or condition associated with MAPT gene expression. In some embodiments, the amount or level of tau protein in the subject is 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 more than 99% lower than the amount or level of tau protein in a subject not administered the oligonucleotide or pharmaceutical composition or a subject receiving a control oligonucleotide, pharmaceutical composition, or treatment (reference subject or control subject). In some embodiments, the amount or level of tau protein in the subject is 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 more than 99% lower than the amount or level of tau protein in a subject not administered the oligonucleotide or pharmaceutical composition or a subject receiving a control oligonucleotide, pharmaceutical composition, or treatment (e.g., reference subject or control subject), for about 1 week to about 12 weeks, about 1 month to about 6 months, or about 7 days to about 91 days in the subject.

[0292] In some embodiments of the method, the amount or level of tau activity in a subject is 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 more than 99% lower than the amount or level of tau activity before administration of the oligonucleotide or pharmaceutical composition. The oligonucleotide or pharmaceutical composition is administered to a subject having a disease, disorder, or condition associated with MAPT gene expression. In some embodiments, the amount or level of tau activity in a subject is 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 more than 99% lower than the amount or level of tau activity before administration of the oligonucleotide or pharmaceutical composition for about 1 week to about 12 weeks, about 1 month to about 6 months, or about 7 days to about 91 days. The oligonucleotide or pharmaceutical composition is administered to a subject having a disease, disorder, or condition associated with MAPT gene expression. In some embodiments, the amount or level of tau activity in a subject is 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 more than 99% lower than the amount or level of tau activity in a control subject (e.g., a reference subject or a control subject) that has not received the oligonucleotide or pharmaceutical composition or has received a control oligonucleotide, pharmaceutical composition, or treatment. In some embodiments, the amount or level of tau activity in a subject is 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 more than 99% lower than the amount or level of tau activity in a control subject (a reference subject or a control subject) that has not received the oligonucleotide or pharmaceutical composition or has received a control oligonucleotide, pharmaceutical composition, or treatment for about 1 week to about 12 weeks, about 1 month to about 6 months, or about 7 days to about 91 days.

[0293] Suitable methods for determining the expression of the MAPT gene, the amount or level of MAPT mRNA, the amount or level of tau protein, and / or the amount or level of tau activity in or from a subject sample are known in the art. Further, the examples provided herein illustrate exemplary methods for determining the expression of the MAPT gene.

[0294] In some embodiments, MAPT gene expression, 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 decreased in a cell (e.g., an oligodendrocyte), a cell population or group (e.g., an organoid), an organ (e.g., the prefrontal cortex), blood or a fraction thereof (e.g., plasma), a tissue (e.g., brain tissue), a sample (e.g., a brain biopsy sample), or other biological material obtained or isolated from a subject. In some embodiments, MAPT gene expression, 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 decreased in multiple cell types (e.g., oligodendrocytes and one or more other types of cells), multiple cell populations, multiple organs (e.g., the brain and one or more other organs), multiple fractions of blood (e.g., plasma and one or more other blood fractions), multiple types of tissue (e.g., brain tissue and one or more other types of tissue), multiple types of samples (e.g., a brain biopsy sample and one or more other types of biopsy samples obtained or isolated from a subject). In some embodiments, MAPT gene expression, 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 decreased in one or more of the cervical spinal cord, thoracic spinal cord, lumbar spinal cord, prefrontal 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 dentate nucleus of the cerebellum. In some embodiments, MAPT gene expression, 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 decreased in tissues of the brain and / or spinal cord associated with AD. In some embodiments, tissues associated with AD include, but are not limited to, the prefrontal cortex, motor cortex, temporal cortex, parietal cortex, and hippocampus. 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, is decreased in tissues of the brain and / or spinal cord associated with PSP.In some embodiments, tissues associated with AD include, but are not limited to, the caudate nucleus, globus pallidus, thalamus, midbrain tegmentum, substantia nigra, pons, cerebellar white matter, dentate nucleus of the cerebellum, medulla, cervical spinal cord, thoracic spinal cord, and lumbar spinal cord.

[0295] Examples of diseases, disorders, or conditions associated with MAPT gene expression include, but are not limited to, AD, FTD, PD, PSP, and tau protein-related diseases (e.g., primary age-related tauopathy, chronic traumatic encephalopathy, corticobasal degeneration, corticobasal body disease, glioma, meningovascular angiomatosis, postencephalitic parkinsonism, and subacute sclerosing panencephalitis), which have abnormal MAPT gene expression leading to the pathologies of these diseases. More than 50 missense, silencing, and intron mutations leading to these diseases are known in MAPT (Ghetti et al. (2015), Neuropathol. Appl. Neurobiol. 41:24-46).

[0296] Due to their high specificity, the oligonucleotides (e.g., RNAi oligonucleotides) herein specifically target the mRNA of target genes in cells, tissues, and / or organs (e.g., the liver). In disease prevention, the target gene may be one required for disease onset or maintenance or identified as being associated with a high risk of developing the disease. In disease treatment, the oligonucleotides can be contacted with cells, tissues, and / or organs (e.g., the brain) presenting or involved in mediating the disease. For example, oligonucleotides substantially identical to all or part of the wild-type (i.e., native) or mutant gene associated with a disorder or condition related to MAPT gene expression may be contacted with or introduced into target cells or tissue types such as oligodendrocytes or other brain cells.

[0297] In some embodiments, the target gene may be a target gene from any mammal, such as a human. Any gene may be silenced according to the methods described herein.

[0298] The methods described herein typically involve administering to a subject a therapeutically effective amount of an oligonucleotide (e.g., an RNAi oligonucleotide), i.e., an amount of the oligonucleotide capable of producing a desired therapeutic result. A therapeutically acceptable amount may be an amount capable of treating a disease or disorder. The appropriate dosage for any given subject will depend on a variety of factors including the size of the subject, body surface area, age, the particular composition being administered, the active ingredient(s) in the composition, the time and route of administration, general health, and other drugs being administered concurrently.

[0299] In some embodiments, a subject is administered a composition of the present disclosure to a target organ (e.g., the brain of the subject) either enterally (e.g., orally, by a gastric feeding tube, by a duodenal feeding tube, via a gastrostomy, or rectally), parenterally (e.g., by subcutaneous injection, intravenous injection or infusion, intra-arterial injection or infusion, intramedullary infusion, intramuscular injection, intracerebral injection, intraventricular injection, or intrathecal injection), topically (e.g., transdermally, by inhalation, via eye drops, or via mucosa), or by direct injection into the target organ (e.g., the brain of the subject). Typically, the oligonucleotide is administered intravenously or subcutaneously. In some embodiments, the oligonucleotide is administered into the cerebrospinal fluid. In some embodiments, the oligonucleotides described herein are administered intrathecally. In some embodiments, the oligonucleotide is administered intraventricularly. In some embodiments, the oligonucleotide is administered by intracapsular magna injection.

[0300] As a non-limiting set of examples, the oligonucleotide is typically administered quarterly (once every 3 months), bi-monthly (once every 2 months), monthly, or weekly. For example, the oligonucleotide may be administered weekly, or at intervals of 2 or 3 weeks. Alternatively, the oligonucleotide may be administered daily. In some embodiments, a subject is administered one or more maintenance doses of the oligonucleotide following one or more loading doses of the oligonucleotide.

[0301] In some embodiments, the subject to be treated is a human or NHP or other mammalian subject. Other exemplary subjects include companion animals such as dogs and cats, livestock such as horses, cows, pigs, sheep, goats, and chickens, and animals such as mice, rats, guinea pigs, and hamsters.

[0302] Kit

[0303] In some embodiments, the present disclosure provides a kit comprising the oligonucleotides herein (e.g., RNAi oligonucleotides) and instructions for use. In some embodiments, the kit comprises an oligonucleotide and a package insert containing instructions for use of the kit and / or any of its components. In some embodiments, the kit comprises an oligonucleotide, one or more controls, various buffers well known in the art, reagents, enzymes, and other standard components, within a suitable container. In some embodiments, the container comprises at least one vial, well, test tube, flask, bottle, syringe, or other container means into which the oligonucleotide is placed and optionally dispensed appropriately. In other embodiments where additional components are provided, the kit contains an additional container into which this component is placed. The kit can also include means for hermetically containing the oligonucleotide and other reagents for commercial sale. Such containers may include injection-molded or blow-molded plastic containers that hold the desired vials. The container and / or the kit can include a label with instructions and / or warnings.

[0304] 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 the progression of a disease, disorder, or condition associated with the expression of the MAPT gene in a subject in need thereof.

[0305] 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 the expression of the MAPT gene in at least one region of the brain and / or at least one region of the spinal cord in a subject in need thereof.

[0306] Definitions

[0307] As used herein, "about" or "approximately" as applied to one or more values of interest refers to a value similar to the reference value being described. In certain embodiments, "about" means within 25%, 20%, 19%, 18%, 17%, 16%, 15%, 14%, 13%, 12%, 11%, 10%, 9%, 8%, 7%, 6%, 5%, 4%, 3%, 2%, 1% in either direction (greater than or less than) of the stated reference value, unless otherwise stated or otherwise apparent from the content (except where such numbers can exceed 100% of the possible values).

[0308] As used herein, "administer", "administering", "administration", etc. refer to providing a substance (e.g., an oligonucleotide) to a subject by a pharmacologically useful (e.g., to treat the condition of the subject) method.

[0309] As used herein, "asialoglycoprotein receptor" or "ASGPR" refers to a dimeric C-type lectin formed by a 48 kDa major subunit (ASGPR-1) and a 40 kDa non-major subunit (ASGPR-2). ASGPR is mainly expressed on the sinusoidal surface of hepatocytes and plays a major role in the binding, internalization, and subsequent excretion of circulating glycoproteins (asialoglycoproteins) containing terminal galactose or GalNAc residues.

[0310] As used herein, "attenuate", "attenuating", "attenuation" and the like refer to weakening or effectively stopping. As a non-limiting example, one or more of the treatments herein may attenuate or effectively stop the onset or progression of a disease associated with the expression of the MAPT gene in a subject (e.g., a tau-related disease). This attenuation may be exemplified, for example, by a decrease in one or more aspects of a disease associated with MAPT gene expression (e.g., a tau-related disease) (e.g., symptoms, tissue characteristics, and cellular activity, inflammatory activity, or immune activity, etc.), the absence of a detectable progression (worsening) of one or more aspects of the disease, or the non-detection of a detectable aspect of the disease in a subject where it would otherwise be expected to be detected.

[0311] As used herein, "complementarity" refers to the structural relationship between two nucleotides that allows (e.g., on two opposing nucleic acids or on opposing regions of a single nucleic acid strand) the two nucleotides to form base pairs with each other. For example, a purine nucleotide of one nucleic acid that is complementary to a pyrimidine nucleotide of an opposing nucleic acid may 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 for the formation of a stable double strand. In some embodiments, two nucleic acids may form complementary regions having regions of multiple nucleotides that are complementary to each other as described herein.

[0312] As used herein, "deoxyribonucleotide" refers to a nucleotide that has a hydrogen in place of a hydroxyl at the 2'-position of its pentose sugar as compared to a ribonucleotide. A modified deoxyribonucleotide is a deoxyribonucleotide having one or more modifications or substitutions at atoms other than the 2'-position, including modifications or substitutions of the sugar, phosphate group, or base.

[0313] As used herein, "double-stranded oligonucleotide" or "ds oligonucleotide" refers to an oligonucleotide that is substantially in a double-stranded form. In some embodiments, the complementary base pairing in the double-stranded region(s) of a ds oligonucleotide is formed between the antiparallel sequences of nucleotides of covalently separated nucleic acid strands. In some embodiments, the complementary base pairing in the double-stranded region(s) of a ds oligonucleotide is formed between the antiparallel sequences of nucleotides of covalently linked nucleic acid strands. In some embodiments, the complementary base pairing in the double-stranded region(s) of a ds oligonucleotide is formed from a single nucleic acid strand that is folded (e.g., via a hairpin) to provide the complementary antiparallel sequences of nucleotides that base pair together. In some embodiments, a ds oligonucleotide comprises two covalently separated nucleic acid strands that are fully double-stranded with each other. However, in some embodiments, a ds oligonucleotide comprises two covalently separated nucleic acid strands that are partially double-stranded (e.g., having overhangs at one or both ends). In some embodiments, a ds oligonucleotide has one or more mismatches, which may include internal or terminal mismatches, since it comprises antiparallel sequences of nucleotides that are partially complementary.

[0314] As used herein, with respect to a nucleic acid (e.g., an oligonucleotide), "double-stranded" refers to a structure formed through complementary base pairing of two antiparallel sequences of nucleotides.

[0315] As used herein, "excipient" refers to a non-therapeutic agent that may be included in a composition, for example, to provide or contribute to a desired concentration or stabilizing effect.

[0316] As used herein, "labile linker" refers to a linker that can be cleaved (e.g., by an acidic pH). "Fairly stable linker" refers to a linker that cannot be cleaved.

[0317] As used herein, "loop" refers to the unpaired region of a nucleic acid (e.g., oligonucleotide) sandwiched between two anti-parallel regions of nucleic acids that are sufficiently complementary to each other, such that under appropriate hybridization conditions (e.g., in a phosphate buffer solution, intracellularly), the two anti-parallel regions sandwiching the unpaired region hybridize to form a double strand (referred to as a "stem").

[0318] As used herein, "modified internucleotide linkage" refers to an internucleotide linkage having one or more chemical modifications compared to a reference internucleotide linkage that includes a phosphodiester linkage. In some embodiments, the modified nucleotide is a non-naturally occurring linkage. Typically, a modified internucleotide linkage imparts one or more desirable properties to a nucleic acid in which the modified internucleotide linkage is present. For example, the modified nucleotide may improve thermal stability, resistance to degradation, nuclease resistance, solubility, bioavailability, biological activity, reduction of immunogenicity, etc.

[0319] As used herein, "modified nucleotide" refers to a nucleotide having one or more chemical modifications compared to the corresponding reference nucleotide selected from adenosine ribonucleotide, guanosine ribonucleotide, cytosine ribonucleotide, uracil ribonucleotide, adenosine deoxyribonucleotide, guanosine deoxyribonucleotide, cytosine deoxyribonucleotide, and thymidine deoxyribonucleotide. In some embodiments, the modified nucleotide is a non-naturally occurring nucleotide. In some embodiments, the modified nucleotide has one or more chemical modifications to its sugar, nucleobase, and / or phosphate group. In some embodiments, the modified nucleotide has one or more chemical moieties attached to the corresponding reference nucleotide. Typically, a modified nucleotide imparts one or more desirable properties to a nucleic acid in which the modified nucleotide is present. For example, the modified nucleotide may improve thermal stability, resistance to degradation, nuclease resistance, solubility, bioavailability, biological activity, reduction of immunogenicity, etc.

[0320] As used herein, the term "tetraloop structure with a nick" refers to the structure of an RNAi oligonucleotide characterized in that the sense (passenger) strand and the antisense (guide) strand are separate, the sense strand has a region complementary to the antisense strand, and at least one of the strands, generally the sense strand, has a tetraL configured to stabilize adjacent stem regions formed within at least one strand.

[0321] As used herein, the term "oligonucleotide" refers to a short nucleic acid (e.g., less than about 100 nucleotides in length). The oligonucleotide may be ss or ds. The oligonucleotide may or may not have a double-stranded region. By way of a non-limiting series of examples, the oligonucleotide may be, but is not limited to, small interfering RNA (siRNA), microRNA (miRNA), short hairpin RNA (shRNA), dicer substrate interfering RNA (dsiRNA), antisense oligonucleotide (ASO), short siRNA, or ss siRNA. In some embodiments, the ds oligonucleotide is an RNAi oligonucleotide.

[0322] As used herein, the term "overhang" refers to the terminal base pair non-forming nucleotide(s) resulting from one strand or region extending beyond the end of the complementary strand with which it forms a duplex. In some embodiments, the overhang comprises one or more non-pairing nucleotides extending from the duplex region at the 5' or 3' end of the ds oligonucleotide. In certain embodiments, the overhang is a 3' or 5' overhang on the antisense or sense strand of the ds oligonucleotide.

[0323] As used herein, "phosphate analog" refers to a chemical moiety that mimics the electrostatic and / or steric properties of a phosphate group. In some embodiments, the phosphate analog is positioned at the 5'-terminal nucleotide of the oligonucleotide instead of the 5'-phosphate, which is often susceptible to enzymatic removal. In some embodiments, the 5'-phosphate analog contains a phosphatase-resistant linkage. Examples of phosphate analogs include, but are not limited to, 5'-phosphonates such as 5'-methylenephosphonate (5'-MP) and 5'-(E)-vinylphosphonate (5'-VP). In some embodiments, the oligonucleotide has a phosphate analog (referred to as a "4'-phosphate analog") at the 4'-carbon position of the sugar of the 5'-terminal nucleotide. An example of a 4'-phosphate analog is an oxymethylphosphonate or an analog thereof in which the oxygen atom of the oxymethyl group is bonded to the sugar moiety (e.g., its 4'-carbon). See, for example, U.S. Provisional Patent Application Nos. 62 / 383,207 (filed Sep. 2, 2016) and 62 / 393,401 (filed Sep. 12, 2016). Other modifications to the 5'-terminus of oligonucleotides have been developed (see, for example, International Patent Application No. WO2011 / 133871; U.S. Patent No. 8,927,513; and Prakash et al. (2015), Nucleic Acids Res. 43:2993-3011).

[0324] As used herein, "MAPT" refers to microtubule-associated protein tau. MAPT transcripts undergo several types of alternative splicing to generate various mRNA species and tau proteins. There are six known tau isoforms generated by splicing of MAPT mRNA. Expression of the MAPT gene is mainly seen in the axons of neurons in the CNS. Tau protein interacts with tubulin to generate microtubules that are involved in several cellular processes. The MAPT mRNA encoding wild-type human tau protein is shown in SEQ ID NO: 909. The MAPT mRNA encoding mouse tau protein is shown in SEQ ID NO: 910. The MAPT mRNA encoding monkey tau protein is shown in SEQ ID NO: 911. However, one of ordinary skill in the art will understand that additional examples of MAPT mRNA sequences are readily available using public databases such as GenBank and UniProt, for example.

[0325] As used herein, "decreased expression" of a gene (e.g., MAPT) refers to a decrease in the amount or level of the RNA transcript (e.g., MAPT mRNA) or protein encoded by that gene, and / or a decrease in the amount or level of the activity of that gene in a cell, cell population, sample, or subject, as compared to an appropriate control (e.g., a reference cell, cell population, sample, or subject). For example, the act of contacting a cell with an oligonucleotide herein (e.g., an oligonucleotide comprising an antisense strand having a nucleotide sequence complementary to the nucleotide sequence that makes up MAPT mRNA) may result in a decrease in the amount or level of MAPT mRNA, tau protein, and / or tau activity (e.g., via inactivation and / or degradation of MAPT mRNA by the RNAi pathway) as compared to a cell not treated with the ds oligonucleotide. Similarly, and as used herein, "decrease in expression" refers to an act that results in a decrease in the expression of a gene (e.g., MAPT).

[0326] As used herein, "decreased MAPT gene expression" refers to a decrease in the amount or level of MAPT mRNA, tau protein, and / or tau activity in a cell, cell population, sample, or subject as compared to an appropriate reference (e.g., a reference cell, cell population, sample, or subject).

[0327] As used herein, "complementary region" refers to the nucleotide sequence of a nucleic acid (e.g., a ds oligonucleotide) that is sufficiently complementary to an antiparallel sequence of nucleotides to permit hybridization between two sequences of nucleotides under appropriate hybridization conditions (e.g., in a phosphate buffer, intracellularly). In some embodiments, the oligonucleotides herein include a targeting sequence having a region complementary to an mRNA target sequence.

[0328] As used herein, "ribonucleotide" refers to a nucleotide having ribose as its pentose sugar and containing a hydroxyl group at the 2'-position. A modified ribonucleotide is a ribonucleotide having one or more modifications or substitutions of atoms other than the 2'-position, including modifications or substitutions in the ribose, phosphate group, or base, or including its own modification or substitution.

[0329] As used herein, "RNAi oligonucleotide" refers to either (a) a ds oligonucleotide having a sense strand (passenger) and an antisense strand (guide), wherein the antisense strand or a portion of the antisense strand is used by the argonaute 2 (Ago2) endonuclease to cleave a target mRNA (e.g., MAPT mRNA), or (b) an ss oligonucleotide having a single-stranded antisense strand, wherein the antisense strand (or a portion of the antisense strand) is used by the Ago2 endonuclease to cleave a target mRNA (e.g., MAPT mRNA).

[0330] As used herein, "strand" refers to a single continuous sequence of nucleotides linked together via internucleotide linkages (e.g., phosphodiester linkages, phosphorothioate linkages). In some embodiments, a strand has two free ends (e.g., a 5' end and a 3' end).

[0331] As used herein, "subject" means a...

Claims

1. An RNAi oligonucleotide comprising an antisense strand and a sense strand forming a double-stranded region, The aforementioned sense chain is 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-GalN Ac][ademA-GalN The sequence is Ac][ademA-GalNAc][mG][mG][mC][mU][mG][mC]-3' (sequence number 850), The antisense strand is the sequence 5'-[MePhosphonate-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), mA is 2'-OMe adenosine, mC is 2'-OMe cytosine, mG is 2'-OMe guanosine, mU is 2'-OMe uridine, fA is 2'-F adenosine, fC is 2'-F cytosine, fG is 2'-F guanosine, fU is 2'-F-uridine, mCs are 2'-OMe cytosine having a 3'-phosphorothioate bond. mGs is 2'-OMe guanosine having a 3'-phosphorothioate bond, mUs is a 2'-OMe uridine having a 3'-phosphorothioate bond, fCs are 2'-F-cytosine having a 3'-phosphorothioate bond. ademA-GalNAc is 2'-aminodiethoxymethanol-adenine-GalNAc, MePhosphonate-4O-mUs has the following structure: 【Chemistry 1】 This is an RNAi oligonucleotide.

2. A pharmaceutical composition comprising an RNAi oligonucleotide containing an antisense strand and a sense strand forming a double-stranded region, The aforementioned sense chain is 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-GalN Ac][ademA-GalN The sequence is Ac][ademA-GalNAc][mG][mG][mC][mU][mG][mC]-3' (sequence number 850), The antisense strand is the sequence 5'-[MePhosphonate-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), mA is 2'-OMe adenosine, mC is 2'-OMe cytosine, mG is 2'-OMe guanosine, mU is 2'-OMe uridine, fA is 2'-F adenosine, fC is 2'-F cytosine, fG is 2'-F guanosine, fU is 2'-F-uridine, mCs are 2'-OMe cytosine having a 3'-phosphorothioate bond. mGs is 2'-OMe guanosine having a 3'-phosphorothioate bond, mUs is a 2'-OMe uridine having a 3'-phosphorothioate bond, fCs are 2'-F-cytosine having a 3'-phosphorothioate bond. ademA-GalNAc is 2'-aminodiethoxymethanol-adenine-GalNAc, MePhosphonate-4O-mUs has the following structure: 【Chemistry 2】 A pharmaceutical composition.

3. The pharmaceutical composition according to claim 2, further comprising a carrier suitable for intravenous administration.

4. The pharmaceutical composition according to claim 3, wherein the carrier comprises water.

5. The pharmaceutical composition according to claim 3, wherein the carrier comprises phosphate-buffered saline.

6. The pharmaceutical composition according to claim 2, further comprising a carrier suitable for intrathecal administration.

7. The pharmaceutical composition according to claim 6, wherein the carrier contains water.

8. The pharmaceutical composition according to claim 6, wherein the carrier comprises phosphate-buffered saline.

9. A pharmaceutical composition according to any one of claims 2 to 8 for reducing the expression of the microtubule-associated protein tau (MAPT) gene in cells, cell populations, or subjects.

10. The pharmaceutical composition according to claim 9, wherein reducing the expression of the MAPT gene includes reducing the amount or level of MAPT mRNA, the amount or level of tau protein, or both.

11. The pharmaceutical composition according to claim 9, wherein the expression of the MAPT gene is reduced in tissue in one or more regions of the central nervous system (CNS), and the tissue is associated with Alzheimer's disease (AD).

12. The pharmaceutical composition according to claim 11, wherein the tissue associated with AD is selected from the prefrontal cortex, motor cortex, temporal cortex, parietal cortex, and hippocampus.

13. The pharmaceutical composition according to claim 9, wherein the expression of the MAPT gene is reduced in tissue in one or more regions of the central nervous system (CNS), and the tissue is associated with progressive supranuclear palsy (PSP).

14. The pharmaceutical composition according to claim 13, wherein the tissue associated with PSP is selected from 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.

15. The pharmaceutical composition according to claim 9, wherein the expression of the MAPT gene is reduced in one or more regions of the central nervous system (CNS) selected from the 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.

16. The pharmaceutical composition according to claim 9, wherein the subject has a disease, disorder, or condition related to the expression of the MAPT gene.

17. The pharmaceutical composition according to claim 16, wherein the disease, disorder, or condition related to MAPT activity is selected from the group consisting of Alzheimer's disease (AD), frontotemporal dementia, progressive supranuclear palsy (PSP), tau protein-related disease, primary age-related tauopathy, chronic traumatic encephalopathy, corticobasal degeneration, Ritico Bodig disease, ganglioglioma, meningeal hemangioma, Parkinson's disease (PD), post-encephalitis parkinsonism, and subacute sclerosing panencephalitis.

18. The pharmaceutical composition according to claim 9, wherein the RNAi oligonucleotide or the pharmaceutical composition is administered in combination with a second composition or therapeutic agent.

19. A method for reducing the expression of the MAPT gene in cells or a population of cells in vitro, comprising contacting the cells or the population of cells with the RNAi oligonucleotide described in claim 1 or the pharmaceutical composition described in claim 2.

20. The method according to claim 19, wherein reducing MAPT activity includes reducing the amount or level of MAPT mRNA, the amount or level of tau protein, or both.

21. A kit comprising an RNAi oligonucleotide according to claim 1 or a pharmaceutically acceptable salt thereof, an optional pharmaceutically acceptable carrier, and a package insert including instructions for use in administering to an individual having a disease, disorder, or condition related to microtubule-associated protein tau (MAPT) activity.

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