Oligonucleotides

JP2024513131A5Pending Publication Date: 2025-04-16HUDSON INST OF MEDICAL RES
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Patent Information

Application Number
JP2023561877
Authority / Receiving Office
JP · JP
Patent Type
Applications
Current Assignee / Owner
Priority Date
2021-10-27
Filing Date
2022-04-08
Publication Date
2025-04-16

AI Technical Summary

Technical Problem

Current oligonucleotide therapeutics face challenges in effectively inhibiting or enhancing the activity of Toll-like receptors (TLR3, TLR7, TLR8, and TLR9) and cyclic GMP-AMP synthase (cGAS) due to a lack of understanding of immunosuppressive sequence determinants and the complex interactions with nucleic acid sensors, leading to potential immunostimulatory effects and off-target responses.

Method used

Designing and modifying oligonucleotides with specific structural motifs, such as 5'-[A/G]GU[A/C][U/C]C-3' and 5'-G[G/C]CCT[C/G]-3', to selectively inhibit or enhance the activity of TLR3, TLR7, TLR8, TLR9, and cGAS, while minimizing immunostimulatory effects.

Benefits of technology

The designed oligonucleotides effectively inhibit or enhance the targeted receptor activities, reducing off-target immune responses and improving the therapeutic efficacy of oligonucleotide-based treatments.

✦ Generated by Eureka AI based on patent content.

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Abstract

The present disclosure relates to methods of selecting, designing or modifying oligonucleotides such that the oligonucleotides inhibit cyclic GMP-AMP synthase (cGAS), Toll-like receptor 3 (TLR3), Toll-like receptor 7 (TLR7), Toll-like receptor 8 (TLR8) and / or Toll-like receptor 9 (TLR9) or enhance Toll-like receptor 8 (TLR8).Further, the present disclosure pertains to methods of selecting, designing or modifying oligonucleotides such that the oligonucleotides exhibit reduced cGAS inhibitory activity.
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Description

[Technical Field]

[0001] CROSS-REFERENCE TO RELATED APPLICATIONS This application claims priority from Australian Provisional Application No. 2021 / 901027, filed April 8, 2021, and No. 2021 / 903431, filed October 27, 2021, the entire contents of each of which are incorporated herein by reference in their entirety.

[0002] The present invention relates to methods for selecting, designing, or modifying oligonucleotides such that the oligonucleotides inhibit cyclic GMP-AMP synthase (cGAS), Toll-like receptor 3 (TLR3), Toll-like receptor 9 (TLR9), Toll-like receptor 8 (TLR8), and / or Toll-like receptor 7 (TLR7) or enhance TLR8. Additionally, the present invention pertains to methods for selecting, designing, or modifying oligonucleotides such that the oligonucleotides exhibit reduced cGAS inhibitory activity. [Background technology]

[0003] RNA-targeting therapeutics based on synthetic oligonucleotides have attracted considerable attention in recent years, receiving regulatory approval in the United States and the European Union (Yin and Rogge, 2019) and securing multibillion-dollar licensing agreements from major pharmaceutical companies (Byrne et al., 2020). To ensure their essential functions related to gene targeting activity, oligonucleotide-based therapeutics require both increased affinity for their targets and stabilization against nuclease activity through the incorporation of modified nucleotides (e.g., 2'-O-methyl [2'OMe], 2'-methoxyethyl [2'MOE], 2'-fluoro [2'F], or locked nucleic acid [LNA]) and modified internucleotide linkages (e.g., phosphorothioate [PS]).

[0004] The complex relationship between synthetic oligonucleotides and nucleic acid sensors in the innate immune system involved in early pathogen detection has been known for 20 years. For example, PS-modified unmethylated "CG" (CpG)-containing DNA oligonucleotides have the ability to activate the DNA sensor Toll-like receptor (TLR) 9 (Krieg et al., 1995; Hemmi et al., 2000) but can also block it in a sequence- and length-dependent manner (Krieg et al., 1998; Gursel et al., 2003; Barrat et al., 2005; Trieu et al., 2006). Similarly, 2'OMe-modified RNA blocks RNA sensing by TLR7 and TLR8 (Robbins et al., 2007; Sioud et al., 2007) and retinoic acid-inducible gene-I (RIG-I) (Devarkar et al., 2016). This knowledge has been important for the design of oligonucleotide therapeutics that can avoid the activation of innate immune sensors and otherwise lead to strong off-target pro-inflammatory immune responses in patients (Krieg et al., 1995; Judge et al., 2005; Judge et al., 2006). From this perspective, chemical modifications may have the dual benefit of increasing the targeting potency of oligonucleotides while decreasing their immunostimulatory effects.

[0005] Nevertheless, it has been clear for some time that select PS-modified oligonucleotides (ODNs) have broad immunosuppressive effects (Bayik et al., 2016). This is best exemplified by the "TTAGGG"-containing PS-ODN A151, which is involved in the inhibition of TLR9 (Gursel et al., 2003), TLR7 (Beignon et al., 2005), Absent In Melanoma 2 (AIM2) (Kaminski et al., 2013), and cyclic GMP-AMP synthase (cGAS) (Steinhagen et al., 2018). These effects are sequence-dependent, and some PS-DNA ODNs exhibit limited immunosuppressive activity against individual immune sensors (Barrat et al., 2005, Bayik et al., 2016). Similarly, 2'OMe oligonucleotides can exhibit sequence-dependent inhibitory effects on TLR7 / 8 sensing (Sarvestani et al., 2015). These observations suggest a complex picture of immunosuppression by chemically modified oligonucleotides, with sequences indicating their activity against nucleic acid sensors. To date, only a small number of ODNs have been studied across different receptors (Bayik et al., 2016; Steinhagen et al., 2018), so a detailed understanding of the immunosuppressive sequence determinants of ODNs is currently lacking. Furthermore, there is no understanding of the immunosuppressive effects of oligonucleotides that combine base and / or backbone modifications, as seen in most approved and in development oligonucleotide therapeutics. While potentially useful for the immunosuppressive effects of therapeutic oligonucleotides (McWhirter and Jefferies, 2020), it is becoming increasingly important to characterize the immunosuppressive effects of therapeutic oligonucleotides to help avoid increased susceptibility to infection in large patient populations beginning to receive ODN therapy (Byrne et al., 2020).

[0006] cGAS has recently emerged as an essential sensor of cytosolic DNA derived from pathogens and damaged endogenous nucleic acids (McWhirter and Jefferies, 2020). Upon activation by DNA, cGAS drives the formation of cyclic GMP-AMP (cGAMP), which binds to stimulator of interferon genes (STING) and promotes the transcriptional induction of IRF3-responsive genes, including CXCL10 (IP-10) and IFNB1. Because cGAS triggers harmful immune responses associated with a wide range of diseases, various approaches to therapeutically targeting cGAS are currently being investigated (An et al., 2018; Lama et al., 2019; Padilla-Salinas et al., 2020; Vincent et al., 2017; Zhao et al., 2020). To this end, the majority of drug design strategies have focused on developing small molecules that inhibit cGAS enzymatic activity (An et al., 2018; Lama et al., 2019; Padilla-Salinas et al., 2020; Vincent et al., 2017; Zhao et al., 2020; Dai et al., 2019; Hall et al., 2017; Wang et al., 2018), which tend to target cGAS systemically rather than in specific tissues. Similar to cGAS, TLR9 is a key factor in autoimmune diseases, and again, there is great interest in developing synthetic TLR9 antagonists to help regulate autoimmune inflammation.

[0007] Therefore, there is a need for alternative inhibitors of the immunostimulatory effects of cGAS and / or TLR9 activity.

[0008] Additionally, there is a need for new or improved inhibitors of Toll-like receptor 3 (TLR3), Toll-like receptor 9 (TLR9), Toll-like receptor 8 (TLR8), and / or Toll-like receptor 7 (TLR7) activity, or new or improved molecules that enhance TLR8 activity. Summary of the Invention

[0009] While designing and testing the oligonucleotides, the inventors observed structural features or motifs that aid in inhibiting cGAS, TLR3, TLR7, TLR8, and / or TLR9 activity. The inventors further observed structural features or motifs in these oligonucleotides that aid in maintaining cGAS activity. The inventors further observed structural features or motifs that aid in enhancing TLR8 activity.

[0010] Thus, in one aspect, the invention provides a method for selecting or designing an oligonucleotide that inhibits cGAS activity, comprising: i) 5'-[A / G]GU[A / C][U / C]C-3' (SEQ ID NO: 1), 5'-A[G / A][U / G]C[U / C]C-3' (SEQ ID NO: 2), 5'-A[G / A][U / G]C[U / C]C[U / C][C / A]U-3' (SEQ ID NO: 3), 5'-GGUAUA-3' (SEQ ID NO: 4), 5'-UGUUUC-3' (SEQ ID NO: 5), 5'-UGUGUC-3' (SEQ ID NO: 6), 5'-CGUUUC-3' (SEQ ID NO: 7), 5'-CGUGUC-3' (SEQ ID NO: 8), 5'-AUGGCCTTTCCGTGCCAAGG-3' (SEQ ID NO: 9), 5'-UCCGGCCTCGGAAGCUCUCU-3' (SEQ ID NO: 10), 5'-GCAUUCCGTGCGGAAGCCUU-3' (SEQ ID NO: 11), 5'-GGCCGAACTTTCCCGCCUUA-3' (SEQ ID NO: 12), 5'-GGUCUTGGCTTCGTGGAGCA-3' (SEQ ID NO: 13), 5'-GGAGCTTCGAGGCCCCAGGC-3' (SEQ ID NO: 14), 5'-GGUGGTCCACAACCCCUUUC-3' (SEQ ID NO: 15), 5'-CAUUAGGTGCAGAAAUCUUC-3' (SEQ ID NO: 16), 5'-UUCUGGGGACTTCCAGUUUA-3' (SEQ ID NO: 17), 5'-UGAUUCCAAAGCCAGGGUUA-3' (SEQ ID NO: 18), 5'-CUUUAGTCGTAGTTGCUUCC-3' (SEQ ID NO: 19), 5'-UUAAATAATCTAGTTUGAAG-3' (SEQ ID NO: 20), 5'-GUGUCCTTCATGCTTUGGAU-3' (SEQ ID NO: 21), 5'-AGAAAGAAGCAAAGAUUCAA-3' (SEQ ID NO: 22), 5'-AGAUUATCTTCTTTTAAUUU-3' (SEQ ID NO: 23), 5'-AAAAGATTATCTTCTUUUAA-3' (SEQ ID NO: 24), 5'-GAAAAGATTATCTTCUUUUA-3' (SEQ ID NO: 25), 5'-UGUGAAAAGATTATCUUCUU-3' (SEQ ID NO: 26), 5'-CUUGUGAAAAGATTAUCUUC-3' (SEQ ID NO: 27), 5'-GGUGGCCACAGGCAACGUCA-3' (SEQ ID NO: 28), 5'-CCAUGTCCCAGGCCTCCAGU-3' (SEQ ID NO: 29), 5'-GCAGUCTCCATGTCCCAGGC-3' (SEQ ID NO: 30), 5'-AGCAGTCTCCATGTCCCAGG-3' (SEQ ID NO: 31), 5'-AGUGGCACATACCACACCCU-3' (SEQ ID NO: 32), 5'-AUUUCCACATGCCCAGUGUU-3' (SEQ ID NO: 33), 5'-GGUCCCATCCCTTCTGCUGC-3' (SEQ ID NO: 34), 5'-UCUGGTCCCATCCCTUCUGC-3' (SEQ ID NO: 35), 5'-GGGUTCCTCCACACCCUUC-3' (SEQ ID NO: 36), 5'-GCAAGGCAGAGAAACUCCAG-3' (SEQ ID NO: 37), 5'-GAUGGTTCCAGTCCCUCUUC-3' (SEQ ID NO: 38), 5'-UGUUUCCCCGGAGAGCAAUG-3' (SEQ ID NO: 39), 5'-AGCCGAACAGAAGGAGCGUC-3' (SEQ ID NO: 40), 5'-GCGUAGTTTCTCTTCCUCCC-3' (SEQ ID NO: 41), 5'-GCGGUATCCATGTCCCAGGC-3' (SEQ ID NO: 42), 5'-GCGGUATACAGGTCCCAGGC-3' (SEQ ID NO: 43), 5'-GCUGUTTCCATGTCCCAGGC-3' (SEQ ID NO: 44), 5'-GCUGUGTCCATGTCCCAGGC-3' (SEQ ID NO: 45), 5'-GCCGUTTCCATGTCCCAGGC-3' (SEQ ID NO: 46), 5'-GCCGUGTCCATGTCCCAGGC-3' (SEQ ID NO: 47), 5'-GCGGUATCCATAGTCUCCAU-3' (SEQ ID NO: 48), 5'-GCGGUATCCATCAGAUAUCG-3' (SEQ ID NO: 49), 5'-CUUUAGTCGTAGTTGUCUCU-3' (SEQ ID NO: 50), 5'-UCCGGGTCGTAGTTGCUUCC-3' (SEQ ID NO: 51), 5'-UCCGGCCTCGGAGTCUCCAU-3' (SEQ ID NO: 52), 5'-UCCGGCCTCGGGAGAUCUCU-3' (SEQ ID NO: 53), 5'-GGUATCCCCCCCCCCCCCCC-3' (SEQ ID NO: 54), 5'-GGAUUAAAACAGATTAAUAC-3' (SEQ ID NO: 55), 5'-GGUAU-3' (SEQ ID NO: 56), 5'-GGUA-3' (SEQ ID NO: 57), 5'-GUAU-3' (SEQ ID NO: 58), 5'-GGU-3' (SEQ ID NO: 59), 5'-GUA-3' (SEQ ID NO: 60), 5'-TGTCTG-3' (SEQ ID NO: 61), 5'-GTCT-3' (SEQ ID NO: 62), 5'-TCTCCG-3' (SEQ ID NO: 63), 5'-CTCC-3' (SEQ ID NO: 64), 5'-[G / A][A / C]AG[G / C][T / C]T[C / A] (SEQ ID NO: 65), 5'-AAAGGTTA-3' (SEQ ID NO: 66), 5'-GAAGCTTC-3' (SEQ ID NO: 67), 5'-GCAGGCTC-3' (SEQ ID NO: 68), 5'-A[G / A]GGTT-3' (SEQ ID NO: 69), 5'-AGGGTT-3' (SEQ ID NO: 70), 5'-AAGGTT-3' (SEQ ID NO: 71), 5'-GGTT-3' (SEQ ID NO: 72), 5'-[A / G]GCT[T / C][T / C][G / C][T / A]-3' (SEQ ID NO: 73), 5'-AGCTTCCT-3' (SEQ ID NO: 74), 5'-AGCTTCGA-3' (SEQ ID NO: 75), 5'-GGCTTCGT-3' (SEQ ID NO: 76), 5'-TGCTTCCT-3' (SEQ ID NO: 77), 5'-AGCTCTCT-3' (SEQ ID NO: 78), 5'-G[G / C]TT-3' (SEQ ID NO: 79), 5'-GCTT-3' (SEQ ID NO: 80), 5'-CGGAGGTCTTGGCTTCGTGG-3' (SEQ ID NO: 81), 5'-AGGTCTTGGCTTCGTGGAGC-3' (SEQ ID NO: 82), 5'-GGGAAAAGGTTATGCAAGGTC-3' (SEQ ID NO: 83), 5'-CTGTGATCTTGACATGCTGC-3' (SEQ ID NO: 84), 5'-ACTGACTGTCTTGAGGGTTC-3' (SEQ ID NO: 85), 5'-GCGTGTCTGGAAGCTTCCTT-3' (SEQ ID NO: 86), 5'-GAGTCTCTGGAGCTTCCTCT-3' (SEQ ID NO: 87), 5'-AGTCGTAGTTGCTTCCTAAC-3' (SEQ ID NO: 88), 5'-GTCTTGGCTTCGTGGAGCAG-3' (SEQ ID NO: 89), 5'-TTGGCTCGGCTTGCCTACTT-3' (SEQ ID NO: 90), 5'-ACAGTGTTGAGATACTCGGG-3' (SEQ ID NO: 91), 5'-TCGCACTTCAGTCTGAGCAG-3' (SEQ ID NO: 92), 5'-GGTGTCCTTGCACGTGGCTT-3' (SEQ ID NO: 93), 5'-TTTGCACACTTCGTACCCAA-3' (SEQ ID NO: 94), 5'-GCTGACAAAGATTCACTGGT-3' (SEQ ID NO: 95), 5'-GCGGAGGTCTTGGCTTCGTG-3' (SEQ ID NO: 96), 5'-CCAAGATCAGCAGTCT-3' (SEQ ID NO: 97), 5'-CTTGAAGCATCGTATC-3' (SEQ ID NO: 98), 5'-GCACACTTCGTACCCA-3' (SEQ ID NO: 99), 5'-GATAGCACCTTCAGCA-3' (SEQ ID NO: 100), 5'-CGTATTATAGCCGATT-3' (SEQ ID NO: 101), 5'-GCAGGCTCAGTGATGT-3' (SEQ ID NO: 102), 5'-GAAAGGTTATGCAAGG-3' (SEQ ID NO: 103), 5'-ATGGCCTCCCATCTCC-3' (SEQ ID NO: 104), 5'-CGCTTTTCTGTCTGGT-3' (SEQ ID NO: 105), 5'-GTGTCTGGAAGCTTCC-3' (SEQ ID NO: 106), 5'-TGGCCTCCCATCTCCT-3' (SEQ ID NO: 107), 5'-ATCTGGCAGCCCATCA-3' (SEQ ID NO: 108), 5'-GAGGTCTTGGCTTCGT-3' (SEQ ID NO: 109), 5'-ACACTTCGTGGGGTCC-3' (SEQ ID NO: 110), 5'-TTCGTGGGGTCCTTTT-3' (SEQ ID NO: 111), 5'-CCACTTGGCAGACCAT-3' (SEQ ID NO: 112), 5'-CCATCCATGAGGTCCT-3' (SEQ ID NO: 113), 5'-TCCAACACTTCGTGGG-3' (SEQ ID NO: 114), 5'-TCTTCATCGGCCCTGC-3' (SEQ ID NO: 115), 5'-CCAGCAGGTCAGCAAA-3' (SEQ ID NO: 116), 5'-CGCTTTTCTCTCCGGT-3' (SEQ ID NO: 117), 5'-GGUAUAGGACTCCAGATGUUUCC-3' (SEQ ID NO: 118), and scanning the polynucleotide or its complement for a region having a variant of that motif or sequence having at least about 75% sequence identity thereto (where U can be T and / or T can be U); ii) generating one or more candidate oligonucleotides containing the motif; iii) testing the ability of one or more candidate oligonucleotides to inhibit cGAS activity; and and iv) selecting an oligonucleotide that inhibits cGAS activity.

[0011] In one embodiment, step i) comprises scanning the polynucleotide or its complement for a motif having the sequence 5'-[A / G]GU[A / C][U / C]C-3' (SEQ ID NO: 1), 5'-A[G / A][U / G]C[U / C]C-3' (SEQ ID NO: 2) or 5'-A[G / A][U / G]C[U / C]C[U / C][C / A]U-3' (SEQ ID NO: 3) (U can be T and / or T can be U). Preferably, the motif 5'-[A / G]GU[A / C][U / C]C-3' (SEQ ID NO: 1), 5'-A[G / A][U / G]C[U / C]C-3' (SEQ ID NO: 2) or 5'-A[G / A][U / G]C[U / C]C[U / C][C / A]U-3' (SEQ ID NO: 3) is at or towards the 5' end of the oligonucleotide.

[0012] In another embodiment, step i) comprises scanning the polynucleotide or its complement for a motif having the sequence 5'-GGUAUA-3' (SEQ ID NO: 4) or a variant having at least about 75% sequence identity thereto (U can be T and / or T can be U). Suitably, the 5'-GGUAUA-3' (SEQ ID NO: 4) motif is at or towards the 5' end of the oligonucleotide.

[0013] In still further embodiments, step i) comprises scanning the polynucleotide or its complement for a motif having the sequence 5'-GGUAU-3' (SEQ ID NO:56), 5'-GGUA-3' (SEQ ID NO:57), 5'-GUAU-3' (SEQ ID NO:58), 5'-GGU-3' (SEQ ID NO:59) or 5'-GUA-3' (SEQ ID NO:60), or a variant having at least about 75% sequence identity thereto (U can be T and / or T can be U). Suitably, the 5'-GGUAU-3' (SEQ ID NO:56), 5'-GGUA-3' (SEQ ID NO:57), 5'-GUAU-3' (SEQ ID NO:58), 5'-GGU-3' (SEQ ID NO:59) or 5'-GUA-3' (SEQ ID NO:60) motif is at or towards the 5' end of the oligonucleotide.

[0014] In a related aspect, the invention provides a method for increasing the cGAS inhibitory activity of an oligonucleotide by modifying the oligonucleotide, wherein the modified oligonucleotide comprises: 5'-[A / G]GU[A / C][U / C]C-3' (SEQ ID NO: 1), 5'-A[G / A][U / G]C[U / C]C-3' (SEQ ID NO: 2), 5'-A[G / A][U / G]C[U / C]C[U / C][C / A]U-3' (SEQ ID NO: 3), 5'-GGUAUA-3' (SEQ ID NO: 4), 5'-UGUUUC-3' (SEQ ID NO: 5), 5'-UGUGUC-3' (SEQ ID NO: 6), 5'-CGUUUC-3' (SEQ ID NO: 7), 5'-CGUGUC-3' (SEQ ID NO: 8), 5'-AUGGCCTTTCCGTGCCAAGG-3' (SEQ ID NO: 9), 5'-UCCGGCCTCGGAAGCUCUCU-3' (SEQ ID NO: 10), 5'-GCAUUCCGTGCGGAAGCCUU-3' (SEQ ID NO: 11), 5'-GGCCGAACTTTCCCGCCUUA-3' (SEQ ID NO: 12), 5'-GGUCUTGGCTTCGTGGAGCA-3' (SEQ ID NO: 13), 5'-GGAGCTTCGAGGCCCCAGGC-3' (SEQ ID NO: 14), 5'-GGUGGTCCACAACCCCUUUC-3' (SEQ ID NO: 15), 5'-CAUUAGGTGCAGAAAUCUUC-3' (SEQ ID NO: 16), 5'-UUCUGGGGACTTCCAGUUUA-3' (SEQ ID NO: 17), 5'-UGAUUCCAAAGCCAGGGUUA-3' (SEQ ID NO: 18), 5'-CUUUAGTCGTAGTTGCUUCC-3' (SEQ ID NO: 19), 5'-UUAAATAATCTAGTTUGAAG-3' (SEQ ID NO: 20), 5'-GUGUCCTTCATGCTTUGGAU-3' (SEQ ID NO: 21), 5'-AGAAAGAAGCAAAGAUUCAA-3' (SEQ ID NO: 22), 5'-AGAUUATCTTCTTTTAAUUU-3' (SEQ ID NO: 23), 5'-AAAAGATTATCTTCTUUUAA-3' (SEQ ID NO: 24), 5'-GAAAAGATTATCTTCUUUUA-3' (SEQ ID NO: 25), 5'-UGUGAAAAGATTATCUUCUU-3' (SEQ ID NO: 26), 5'-CUUGUGAAAAGATTAUCUUC-3' (SEQ ID NO: 27), 5'-GGUGGCCACAGGCAACGUCA-3' (SEQ ID NO: 28), 5'-CCAUGTCCCAGGCCTCCAGU-3' (SEQ ID NO: 29), 5'-GCAGUCTCCATGTCCCAGGC-3' (SEQ ID NO: 30), 5'-AGCAGTCTCCATGTCCCAGG-3' (SEQ ID NO: 31), 5'-AGUGGCACATACCACACCCU-3' (SEQ ID NO: 32), 5'-AUUUCCACATGCCCAGUGUU-3' (SEQ ID NO: 33), 5'-GGUCCCATCCCTTCTGCUGC-3' (SEQ ID NO: 34), 5'-UCUGGTCCCATCCCTUCUGC-3' (SEQ ID NO: 35), 5'-GGGUTCCTCCACACCCUUC-3' (SEQ ID NO: 36), 5'-GCAAGGCAGAGAAACUCCAG-3' (SEQ ID NO: 37), 5'-GAUGGTTCCAGTCCCUCUUC-3' (SEQ ID NO: 38), 5'-UGUUUCCCCGGAGAGCAAUG-3' (SEQ ID NO: 39), 5'-AGCCGAACAGAAGGAGCGUC-3' (SEQ ID NO: 40), 5'-GCGUAGTTTCTCTTCCUCCC-3' (SEQ ID NO: 41), 5'-GCGGUATCCATGTCCCAGGC-3' (SEQ ID NO: 42), 5'-GCGGUATACAGGTCCCAGGC-3' (SEQ ID NO: 43), 5'-GCUGUTTCCATGTCCCAGGC-3' (SEQ ID NO: 44), 5'-GCUGUGTCCATGTCCCAGGC-3' (SEQ ID NO: 45), 5'-GCCGUTTCCATGTCCCAGGC-3' (SEQ ID NO: 46), 5'-GCCGUGTCCATGTCCCAGGC-3' (SEQ ID NO: 47), 5'-GCGGUATCCATAGTCUCCAU-3' (SEQ ID NO: 48), 5'-GCGGUATCCATCAGAUAUCG-3' (SEQ ID NO: 49), 5'-CUUUAGTCGTAGTTGUCUCU-3' (SEQ ID NO: 50), 5'-UCCGGGTCGTAGTTGCUUCC-3' (SEQ ID NO: 51), 5'-UCCGGCCTCGGAGTCUCCAU-3' (SEQ ID NO: 52), 5'-UCCGGCCTCGGGAGAUCUCU-3' (SEQ ID NO: 53), 5'-GGUATCCCCCCCCCCCCCCC-3' (SEQ ID NO: 54), 5'-GGAUUAAAACAGATTAAUAC-3' (SEQ ID NO: 55), 5'-GGUAU-3' (SEQ ID NO: 56), 5'-GGUA-3' (SEQ ID NO: 57), 5'-GUAU-3' (SEQ ID NO: 58), 5'-GGU-3' (SEQ ID NO: 59), 5'-GUA-3' (SEQ ID NO: 60), 5'-TGTCTG-3' (SEQ ID NO: 61), 5'-GTCT-3' (SEQ ID NO: 62), 5'-TCTCCG-3' (SEQ ID NO: 63), 5'-CTCC-3' (SEQ ID NO: 64), 5'-[G / A][A / C]AG[G / C][T / C]T[C / A] (SEQ ID NO: 65), 5'-AAAGGTTA-3' (SEQ ID NO: 66), 5'-GAAGCTTC-3' (SEQ ID NO: 67), 5'-GCAGGCTC-3' (SEQ ID NO: 68), 5'-A[G / A]GGTT-3' (SEQ ID NO: 69), 5'-AGGGTT-3' (SEQ ID NO: 70), 5'-AAGGTT-3' (SEQ ID NO: 71), 5'-GGTT-3' (SEQ ID NO: 72), 5'-[A / G]GCT[T / C][T / C][G / C][T / A]-3' (SEQ ID NO: 73), 5'-AGCTTCCT-3' (SEQ ID NO: 74), 5'-AGCTTCGA-3' (SEQ ID NO: 75), 5'-GGCTTCGT-3' (SEQ ID NO: 76), 5'-TGCTTCCT-3' (SEQ ID NO: 77), 5'-AGCTCTCT-3' (SEQ ID NO: 78), 5'-G[G / C]TT-3' (SEQ ID NO: 79), 5'-GCTT-3' (SEQ ID NO: 80), 5'-CGGAGGTCTTGGCTTCGTGG-3' (SEQ ID NO: 81), 5'-AGGTCTTGGCTTCGTGGAGC-3' (SEQ ID NO: 82), 5'-GGGAAAAGGTTATGCAAGGTC-3' (SEQ ID NO: 83), 5'-CTGTGATCTTGACATGCTGC-3' (SEQ ID NO: 84), 5'-ACTGACTGTCTTGAGGGTTC-3' (SEQ ID NO: 85), 5'-GCGTGTCTGGAAGCTTCCTT-3' (SEQ ID NO: 86), 5'-GAGTCTCTGGAGCTTCCTCT-3' (SEQ ID NO: 87), 5'-AGTCGTAGTTGCTTCCTAAC-3' (SEQ ID NO: 88), 5'-GTCTTGGCTTCGTGGAGCAG-3' (SEQ ID NO: 89), 5'-TTGGCTCGGCTTGCCTACTT-3' (SEQ ID NO: 90), 5'-ACAGTGTTGAGATACTCGGG-3' (SEQ ID NO: 91), 5'-TCGCACTTCAGTCTGAGCAG-3' (SEQ ID NO: 92), 5'-GGTGTCCTTGCACGTGGCTT-3' (SEQ ID NO: 93), 5'-TTTGCACACTTCGTACCCAA-3' (SEQ ID NO: 94), 5'-GCTGACAAAGATTCACTGGT-3' (SEQ ID NO: 95), 5'-GCGGAGGTCTTGGCTTCGTG-3' (SEQ ID NO: 96), 5'-CCAAGATCAGCAGTCT-3' (SEQ ID NO: 97), 5'-CTTGAAGCATCGTATC-3' (SEQ ID NO: 98), 5'-GCACACTTCGTACCCA-3' (SEQ ID NO: 99), 5'-GATAGCACCTTCAGCA-3' (SEQ ID NO: 100), 5'-CGTATTATAGCCGATT-3' (SEQ ID NO: 101), 5'-GCAGGCTCAGTGATGT-3' (SEQ ID NO: 102), 5'-GAAAGGTTATGCAAGG-3' (SEQ ID NO: 103), 5'-ATGGCCTCCCATCTCC-3' (SEQ ID NO: 104), 5'-CGCTTTTCTGTCTGGT-3' (SEQ ID NO: 105), 5'-GTGTCTGGAAGCTTCC-3' (SEQ ID NO: 106), 5'-TGGCCTCCCATCTCCT-3' (SEQ ID NO: 107), 5'-ATCTGGCAGCCCATCA-3' (SEQ ID NO: 108), 5'-GAGGTCTTGGCTTCGT-3' (SEQ ID NO: 109), 5'-ACACTTCGTGGGGTCC-3' (SEQ ID NO: 110), 5'-TTCGTGGGGTCCTTTT-3' (SEQ ID NO: 111), 5'-CCACTTGGCAGACCAT-3' (SEQ ID NO: 112), 5'-CCATCCATGAGGTCCT-3' (SEQ ID NO: 113), 5'-TCCAACACTTCGTGGG-3' (SEQ ID NO: 114), 5'-TCTTCATCGGCCCTGC-3' (SEQ ID NO: 115), 5'-CCAGCAGGTCAGCAAA-3' (SEQ ID NO: 116), 5'-CGCTTTTCTCTCCGGT-3' (SEQ ID NO: 117), A motif having a sequence selected from the group consisting of 5'-GGUAUAGGACTCCAGATGUUUCC-3' (SEQ ID NO: 118), and and modifying the oligonucleotide to include a variant of that motif or sequence having at least about 75% sequence identity thereto (where U can be T and / or T can be U).

[0015] In one embodiment, the step of modifying the oligonucleotide comprises adding a sequence of nucleotides to the 5' and / or 3' end of the oligonucleotide such that the modified oligonucleotide comprises a motif.

[0016] In one particular embodiment, the step of modifying the oligonucleotide comprises adding a sequence of nucleotides, such as 5'-GGUATC-3' (SEQ ID NO: 119), 5'-GGUAUC-3' (SEQ ID NO: 120), or a fragment or portion thereof, to the 5' end of the oligonucleotide, such that the modified oligonucleotide comprises the motif.

[0017] In some embodiments, the step of modifying the oligonucleotide comprises adding a motif of 5'-GGUAUA-3' (SEQ ID NO: 4), 5'-GGUAU-3' (SEQ ID NO: 56), 5'-GGUA-3' (SEQ ID NO: 57), 5'-GUAU-3' (SEQ ID NO: 58), 5'-GGU-3' (SEQ ID NO: 59), 5'-GUA-3' (SEQ ID NO: 60), or a fragment or portion thereof (U can be T) to the 5' and / or 3' end of the oligonucleotide such that the modified oligonucleotide comprises the motif. More particularly, the step of modifying the oligonucleotide suitably comprises adding 5'-GGUAUA-3' (SEQ ID NO: 4), 5'-GGUAU-3' (SEQ ID NO: 56), 5'-GGUA-3' (SEQ ID NO: 57), 5'-GUAU-3' (SEQ ID NO: 58), 5'-GGU-3' (SEQ ID NO: 59), 5'-GUA-3' (SEQ ID NO: 60) or a fragment or portion thereof (U may be T) to the 5' end of the oligonucleotide such that the modified oligonucleotide comprises the motif.

[0018] In one embodiment, the method of this aspect further comprises testing the ability of the modified oligonucleotides to inhibit cGAS activity and selecting oligonucleotides that inhibit cGAS activity to a greater extent than unmodified oligonucleotides.

[0019] In one embodiment of the above aspect, the oligonucleotide does not bind or is not designed to bind to a transcript encoding cGAS or its complement.

[0020] In another embodiment of the above aspect, the oligonucleotide binds to, or is designed to bind to, a target transcript that does not encode cGAS or its complement.

[0021] In an alternative embodiment of the above aspect, the oligonucleotide binds to, or is designed to bind to, a target transcript encoding cGAS or its complement.

[0022] In yet another embodiment, the oligonucleotide does not bind or is not designed to bind to the target transcript.

[0023] In one embodiment of the above aspect, the motif is within 11 bases of the 5' and / or 3' end of the oligonucleotide.

[0024] In a further embodiment of the above aspect, the motif is within 8 bases of the 5' and / or 3' end of the oligonucleotide.

[0025] In still further embodiments of the above aspects, the motif is at or towards the 5' and / or 3' end of the oligonucleotide. By way of example, in embodiments where the motif is 5'-GGUAUA-3' (SEQ ID NO:4), 5'-GGUAU-3' (SEQ ID NO:56), 5'-GGUA-3' (SEQ ID NO:57), 5'-GUAU-3' (SEQ ID NO:58), 5'-GGU-3' (SEQ ID NO:59), 5'-GUA-3' (SEQ ID NO:60) (where U can be T), the motif is preferably at or towards the 5' end of the oligonucleotide.

[0026] Examples of motifs of the above embodiment include the sequences 5'-GGUAUC-3' (SEQ ID NO: 120), 5'-AGUCUC-3' (SEQ ID NO: 121), 5'-GGUCCC-3' (SEQ ID NO: 122), 5'-GGUCUC-3' (SEQ ID NO: 123), 5'-AAGCUC-3' (SEQ ID NO: 124), 5'-AGUCCC-3' (SEQ ID NO: 125), 5'-GGUAUA-3' (SEQ ID NO: 126), 5'-UGUUUC-3' (SEQ ID NO: 127), 5'-AAGCUC-3' (SEQ ID NO: 128), 5'-AGUCCC-3' (SEQ ID NO: 129), 5'-GGUAUA-3' (SEQ ID NO: 130), 5'-UGUUUC-3' (SEQ ID NO: 131), 5'-UGUUUC-3' (SEQ ID NO: 132), 5'-UGUUUC-3' (SEQ ID NO: 133), 5'-UGUUUC-3' (SEQ ID NO: 134), 5'-UGUUUC-3' (SEQ ID NO: 135), 5'-UGUUUC-3' (SEQ ID NO: 136), 5'-UGUUUC-3' (SEQ ID NO: 137), 5'-UGUUUC-3' (SEQ ID NO: 138), 5'-UGUUUC-3' (SEQ ID NO: 139), 5'-UGUUUC-3' (SEQ ID NO: 140), 5'-UGUUUC-3' (SEQ ID NO: 141), 5'-UGUUUC-3' (SEQ ID NO: 142), ' (SEQ ID NO: 5), 5'-UGUGUC-3' (SEQ ID NO: 6), 5'-CGUUUC-3' (SEQ ID NO: 7), 5'-CGUGUC-3' (SEQ ID NO: 8), 5'-GGUAU-3' (SEQ ID NO: 56), 5'-GGUA-3' (SEQ ID NO: 57), 5'-GUAU-3' (SEQ ID NO: 58), 5'-GGU-3' (SEQ ID NO: 59), 5'-GUA-3' (SEQ ID NO: 60), 5'-TGTCTG-3' (SEQ ID NO: 61) ), 5'-GTCT-3' (SEQ ID NO: 62), 5'-TCTCCG-3' (SEQ ID NO: 63), 5'-CTCC-3' (SEQ ID NO: 64), 5'-AAAGGTTA-3' (SEQ ID NO: 66), 5'-GAAGCTTC-3' (SEQ ID NO: 67), 5'-GCAGGCTC-3' (SEQ ID NO: 68), 5'-AGGGTT-3' (SEQ ID NO: 70), 5'-AAGGTT-3' (SEQ ID NO: 71), 5'-GGTT-3' (SEQ ID NO: 72), 5'-AGCTTCCT-3' (SEQ ID NO: 74), 5'-AGCTTCGA-3' (SEQ ID NO: 75), 5'-GGCTTCGT-3' (SEQ ID NO: 76), 5'-TGCTTCCT-3' (SEQ ID NO: 77), 5'-AGCTCTCT-3' (SEQ ID NO: 78), or 5'-GCTT-3' (SEQ ID NO: 80) (U may be T).More specifically, the motif of the above embodiment is preferably 5'-GGUAUC-3' (SEQ ID NO: 120), 5'-GGUATC-3' (SEQ ID NO: 119), 5'-AGUCTC-3' (SEQ ID NO: 126), 5'-AGTCTC-3' (SEQ ID NO: 127), 5'-GGUCCC-3' (SEQ ID NO: 122), 5'-GGUCTC-3' (SEQ ID NO: 128), 5'-AAGCUC-3' (SEQ ID NO: 124), 5'-AGTCCC-3' (SEQ ID NO: 129), 5'-GGUATA-3' (SEQ ID NO: 130), 5'-UGUTTC-3' (SEQ ID NO: 131), 5'-UGUGTC-3' (SEQ ID NO: 132), 5'-CGUTTC-3' (SEQ ID NO: 133), 5'-CGUGTC-3' (SEQ ID NO: 134), 5'-GGUAU-3' (SEQ ID NO: 56), 5'-GGUAT-3' (SEQ ID NO: 135), 5'-GGUA-3' (SEQ ID NO: 57), 5'-GUAU-3' (SEQ ID NO: 58), 5'-GUAT-3' (SEQ ID NO: 136), 5'-GGU-3' (SEQ ID NO: 59), 5'-GUA-3' (SEQ ID NO: 60), 5'-TGTCTG-3' (SEQ ID NO: 61), 5'-GTCT-3' (SEQ ID NO: 62), 5'-TCTCCG-3' (SEQ ID NO: 63), 5'-CTCC-3' (SEQ ID NO: 64), 5'-AAAGGTTA-3' (SEQ ID NO: 66), 5'-GAAGCTTC-3' (SEQ ID NO: 67), 5'-GCAGGCTC-3' (SEQ ID NO: 68), 5'-A GGGTT-3' (SEQ ID NO: 70), 5'-AAGGTT-3' (SEQ ID NO: 71), 5'-GGTT-3' (SEQ ID NO: 72), 5'-AGCTTCCT-3' (SEQ ID NO: 74), 5'-AGCTTCGA-3' (SEQ ID NO: 75), 5'-GGCTTCGT-3' (SEQ ID NO: 76), 5'-TGCTTCCT-3' (SEQ ID NO: 77), 5'-AGCTCTCT-3' (SEQ ID NO: 78) or 5'-GCTT-3' (SEQ ID NO: 80).

[0027] In one particular embodiment, the motif of the above aspect has the sequence 5'-GGUAUC-3' (SEQ ID NO: 120), 5'-GGUATC-3' (SEQ ID NO: 119), 5'-GCGGUATCCATGTCCCAGGC-3' (SEQ ID NO: 42), 5'-GCGGUAUCCATGTCCCAGGC-3' (SEQ ID NO: 137), 5'-GGUAUA-3' (SEQ ID NO: 4), 5'-GGUAU-3' (SEQ ID NO: 56), 5'-GGUA-3' (SEQ ID NO: 57), 5'-GUAU-3' (SEQ ID NO: 58), 5'-GGU-3' (SEQ ID NO: 59), 5'-GUA-3' (SEQ ID NO: 60), or a variant thereof having at least about 75% sequence identity thereto (U can be T and / or T can be U).

[0028] In one embodiment, one or more of the bases of the motif of the above aspect are modified bases and / or have a modified backbone.

[0029] In certain embodiments, the motif of the above aspect is 5'-mGmGmUATC-3', 5'-mGmGmUAUC-3', 5'-mAmGmUCTC-3', 5'-mAmGTCTC-3', 5'-mGmGmUmCmCC-3', 5'-mGmGmUmCTC-3', 5'-AAGCmUmC-3', 5'-AGTCCC-3' (SEQ ID NO: 129), 5'-mGmGmUATA-3', 5'-mUmGmUTT C-3', 5'-mUmGmUGTC-3', 5'-mCmGmUTTC-3', 5'-mCmGmUGTC-3', 5'-mGmGmUAU-3', 5'-mGmGmUAT-3', 5'-mGmGm UmAU-3', 5'-mGmGmUmAT-3', 5'-mGmGmUmAmU-3', 5'-mGmGmUA-3', 5'-mGmGmUmA-3', 5'-mGmGmU-3', 5'-mTmGTC TG-3', 5'-TGTCTmG-3', mTmGTCTG-3', 5'-mGTCT-3', 5'-GTCT-3' (SEQ ID NO: 62), 5'-TCTCCG-3' (SEQ ID NO: 63), 5'-TCTCCmG-3', 5'-CTCC-3' (SEQ ID NO: 64), 5'-mAmAAGGTTA-3', 5'-GAAGCTmTmC-3', 5'-mGmCmAGGCTC-3', 5'-mAAGGTT-3', The amino acid sequence has the sequence 5'-AGmGmGmTmT-3', 5'-AGCTmTmCmCmT-3', 5'-AGCTTmCmCmT-3', 5'-mAmGmCTTCGA-3', 5'-GGCTTmCmGmT-3', 5'-GGCTTCGT-3' (SEQ ID NO: 76), 5'-TGCTTCmCmT-3' or 5'-AGCmTmCmTmCmT-3' (wherein m is a modified base and / or has a modified backbone).

[0030] In one particular embodiment, the motif of the above aspect is 5'-mGmGmUATC-3', 5'-mGmGmUAUC-3', 5'-mGmCmGmGmUATCCATGTCCmCmAmGmGmC-3', 5'-mGmCmGmGmUAUCCATGTCCmCmAmGmGmC-3', 5'-mGmGmUmAmUmC-3', 5'-mGmCmGmGmUmAmUmCmCmAmUmGmUmCmCmCmAmAmGmGmC-3', 5'-mGmGmUATCCCCCCCCCCCCCCC-3', 5'-mGmGmUAU-3', 5'-mGmGmUAT-3', 5'-mGmGmUmAU-3', 5'-mGmGmUmAT-3', 5'-mGmGmUmAmU-3', 5'-mGmGmUA-3', 5'-mGmGmUmA-3', 5'-mGmUmAmU-3', 5'-mGmGmU-3', The sequence 5'-mGmUmA-3', or a variant thereof having at least about 75% sequence identity thereto (wherein U can be T and / or T can be U, m is a modified base and / or has a modified backbone).

[0031] In one embodiment of the above two aspects, the motif has the sequence: 5'-CGCTTTTCTGTCTGGT-3' (SEQ ID NO: 105), 5'-GAAAGGTTATGCAAGG-3' (SEQ ID NO: 103), 5'-GCAGGCTCAGTGATGT-3' (SEQ ID NO: 102), or 5'-GTGTCTGGAAGCTTCC-3' (SEQ ID NO: 106) (U can be T and / or T can be U).

[0032] In particular embodiments of the above two aspects, the motif has the sequence: 5'-mCmGmCTTTTCTGTCTmGmGmT-3', 5'-mGmAmAAGGTTATGCAmAmGmG-3', 5'-mGmCmAGGCTCAGTGAmTmGmT-3', or 5'-mGmTmGTCTGGAAGCTmTmCmC-3', (U may be T and / or T may be U, and m is a modified base and / or has a modified backbone). In such examples, m is suitably a 2'-LNA modified base.

[0033] In another embodiment of the above two aspects, the motif has the sequence: 5'-CGGAGGTCTTGGCTTCGTGG-3' (SEQ ID NO: 81), 5'-GGAGCTTCGAGGCCCCAGGC-3' (SEQ ID NO: 14), 5'-AGGTCTTGGCTTCGTGGAGC-3' (SEQ ID NO: 82), 5'-GGGAAAAGGTTATGCAAGGTC-3' (SEQ ID NO: 83), 5'-CTGTGATCTTGACATGCTGC-3' (SEQ ID NO: 84), 5'-ACTGACTGTCTTGAGGGTTC-3' (SEQ ID NO: 85), 5'-GCGTGTCTGGAAGCTTCCTT-3' (SEQ ID NO: 86), 5'-TCCGGCCTCGGAAGCTCTCT-3' (SEQ ID NO: 138), 5'-GAGTCTCTGGAGCTTCCTCT-3' (SEQ ID NO: 87), 5'-GGTCTTGGCTTCGTGGAGCA-3' (SEQ ID NO: 139), or 5'-AGTCGTAGTTGCTTCCTAAC-3' (SEQ ID NO: 88), (U can be T and / or T can be U).

[0034] In particular embodiments of the above two aspects, the motif has the sequence: 5'-mCmGmGmAmGGTCTTGGCTTmCmGmTmGmG-3', 5'-mGmGmAmGmCTTCGAGGCCCmCmAmGmGmC-3', 5'-mAmGmGmTmCTTGGCTTCGTmGmGmAmGmC-3', 5'-mGmGmGmAmAAGGTTATGCAmAmGmGmTmC-3', 5'-mCmTmGmTmGATCTTGACATmGmCmTmGmC-3', 5'-mAmCmTmGmACTGTCTTGAGmGmGmTmTmC-3', 5'-mGmCmGmTmGTCTGGAAGCTmTmCmCmTmT-3', 5'-mTmCmCmGmGCCTCGGAAGCmTmCmTmCmT-3', 5'-mGmAmGmTmCTCTGGAGCTTmCmCmTmCmT-3', 5'-mGmGmTmCmTTGGCTTCGTGmGmAmGmCmA-3', or 5'-mAmGmTmCmGTAGTTGCTTCmCmTmAmAmC-3', (U may be T and / or T may be U, and m is a modified base and / or has a modified backbone). In such examples, m is preferably a 2'-MOE modified base.

[0035] In another aspect, the invention provides a method for selecting or designing an oligonucleotide that does not inhibit cGAS activity, comprising: i) 5'-[C / U]CUUCU-3' (SEQ ID NO: 140), 5'-CACCCTTCTCTCTGGUCCCA-3' (SEQ ID NO: 141), 5'-CCUUCTCTCTGGTCCCAUCC-3' (SEQ ID NO: 142), 5'-UCUCUGGTCCCATCCCUUCU-3' (SEQ ID NO: 143), 5'-AUAUCTGCTGCCCACCUUCU-3' (SEQ ID NO: 144), 5'-GUCCCATCCCTTCTGCUGCC-3' (SEQ ID NO: 145), 5'-GUCUCCTCCACACCCUUCUC-3' (SEQ ID NO: 146), 5'-CAGGCCTCCAGTGTCUUCUC-3' (SEQ ID NO: 147), 5'-UCCCAACTCTTCTAACUCGU-3' (SEQ ID NO: 148), and scanning the polynucleotide or its complement for a region having a variant of that motif or sequence having at least about 75% sequence identity thereto (where U can be T and / or T can be U); ii) generating one or more candidate oligonucleotides containing the motif; iii) testing the ability of one or more candidate oligonucleotides to inhibit cGAS activity; and and iv) selecting an oligonucleotide that does not inhibit cGAS activity.

[0036] In a related aspect, the invention provides a method for reducing the cGAS inhibitory activity of an oligonucleotide by modifying the oligonucleotide, wherein the modified oligonucleotide comprises: 5'-[C / U]CUUCU-3' (SEQ ID NO: 140), 5'-CACCCTTCTCTCTGGUCCCA-3' (SEQ ID NO: 141), 5'-CCUUCTCTCTGGTCCCAUCC-3' (SEQ ID NO: 142), 5'-UCUCUGGTCCCATCCCUUCU-3' (SEQ ID NO: 143), 5'-AUAUCTGCTGCCCACCUUCU-3' (SEQ ID NO: 144), 5'-GUCCCATCCCTTCTGCUGCC-3' (SEQ ID NO: 145), 5'-GUCUCCTCCACACCCUUCUC-3' (SEQ ID NO: 146), 5'-CAGGCCTCCAGTGTCUUCUC-3' (SEQ ID NO: 147), 5'-UCCCAACTCTTCTAACUCGU-3' (SEQ ID NO: 148), and and modifying the oligonucleotide to include a variant of that motif or sequence having at least about 75% sequence identity thereto (U can be T and / or T can be U).

[0037] In one embodiment, the step of modifying the oligonucleotide comprises adding a sequence of nucleotides to the 5' and / or 3' end of the oligonucleotide such that the modified oligonucleotide comprises a motif.

[0038] In one embodiment, the method of the invention further comprises testing the ability of modified oligonucleotides to inhibit cGAS activity and selecting oligonucleotides that inhibit cGAS activity to a lesser extent than unmodified oligonucleotides. With reference to the two aforementioned embodiments, the motif is preferably within 13 bases of the 5' and / or 3' end of the oligonucleotide. More particularly, the motif is preferably within 9 bases of the 5' and / or 3' end of the oligonucleotide. Even more particularly, the motif is preferably at or towards the 5' and / or 3' end of the oligonucleotide.

[0039] In one embodiment of the two aforementioned aspects, the motif has the sequence 5'-CCUUCU-3' (SEQ ID NO: 149) or 5'-UCUUCU-3' (SEQ ID NO: 150) (U can also be T).

[0040] In one embodiment of the two aforementioned aspects, one or more of the bases of the motif are modified bases and / or have a modified backbone.

[0041] In one embodiment of the two aforementioned aspects, the motif has the sequence 5'-mCmCUUCU-3', 5'-mCmCmUmUmCU-3', 5'-CmCmUmUmCmU-3', 5'-CCUUCU-3' (SEQ ID NO: 149), 5'-CCmUmUmCmU-3', 5'-UCmUmUmCmU-3', 5'-UCUUCU-3' (SEQ ID NO: 150), 5'-UmCmUmUmCmU-3' or 5'-CmCmUmUmCmU-3' (U can be T and m is a modified base and / or has a modified backbone).

[0042] In embodiments of any of the above aspects, the motif comprises, consists essentially of, or consists of any of the sequences in Table 1, 1A, 2, 2A, 3, 3A, 4, 4A, 5, 5A, or 6, with or without the specific modifications defined, and is shown in the Examples to inhibit the activity of cGAS.

[0043] With respect to the above aspects, the method may include the further step of testing the ability of one or more candidate or modified oligonucleotides to inhibit TLR3, TLR7 and / or TLR9 activity, and optionally selecting oligonucleotides that inhibit or do not substantially inhibit TLR3, TLR7 and / or TLR9 activity.

[0044] In certain embodiments, the one or more candidate oligonucleotides or modified oligonucleotides inhibit cGAS and TLR7 activity. Examples of motifs for such embodiments include 5'-GCAGUCTCCATGTCCCAGGC-3' (SEQ ID NO: 30), 5'-GAUGGTTCCAGTCCCUCUUC-3' (SEQ ID NO: 38), 5'-AGCAGTCTCCATGTCCCAGG-3' (SEQ ID NO: 31), 5'-GGGUCTCCTCCACACCCUUC-3' (SEQ ID NO: 36), 5'-GGUGGCCACAGGCAACGUCA-3' (SEQ ID NO: 28), 5'-GCCGUTTCCATGTCCCAGGC-3' (SEQ ID NO: 46), 5'-GCGGUATCCATGTCCCAGGC-3' (SEQ ID NO: 42), 5'-GCGGUATACAGGTCCCAGGC-3' (SEQ ID NO: 43), 5'-GCUGUTTCCATGTCCCAGGC-3' (SEQ ID NO: 44), 5'-GCUGUGTCCATGTCCCAGGC-3' (SEQ ID NO: 45). sequence number 45), 5'-GCCGUGTCCATGTCCCAGGC-3' (SEQ ID NO: 47), 5'-GCGGUAUCCAUGUCCCAGGC-3' (SEQ ID NO: 151), 5'-GGUATCCCCCCCCCCCCCCC-3' (SEQ ID NO: 54), 5'-CCAUGTCCCAGGCCTCCAGU-3' (SEQ ID NO: 29), 5'-GCAAGGCAGAGAAACUCCAG-3' (SEQ ID NO: 37), 5'-GGAUUAAAACAGATTAAUAC-3' (SEQ ID NO: 55), 5'-AGCCGAACAGAAGGAGCGUC-3' (SEQ ID NO: 40), 5'-UCCGGCCTCGGAAGCUCUCU-3' (SEQ ID NO: 10), 5'-UCCGGCCTCGGAGTCUCCAU-3' (SEQ ID NO: 52), and 5'-GCGGUATCCATAGTCUCCAU-3' (SEQ ID NO: 48).

[0045] In further embodiments, one or more candidate oligonucleotides or modified oligonucleotides inhibit cGAS and TLR7 activity but do not substantially inhibit TLR9 activity. Exemplary motifs for such embodiments include 5'-GCAGUCTCCATGTCCCAGGC-3' (SEQ ID NO: 30), 5'-GAUGGTTCCAGTCCCUCUUC-3' (SEQ ID NO: 38), 5'-AGCAGTCTCCATGTCCCAGG-3' (SEQ ID NO: 31), 5'-GGGUCTCCTCCACACCCUUC-3' (SEQ ID NO: 36), 5'-GGUGGCCACAGGCAACGUCA-3' (SEQ ID NO: 28), 5'-GCCGUTTCCATGTCCCAGGC-3' (SEQ ID NO: 46), 5'-GCGGUATC CATGTCCCAGGC-3' (SEQ ID NO: 42), 5'-GCGGUATACAGGTCCCAGGC-3' (SEQ ID NO: 43), 5'-GCUGUTTCCATGTCCCAGGC-3' (SEQ ID NO: 44), 5'-GCUGUGTCCATGTCCCAGGC-3' (SEQ ID NO: 45), 5'-GCCGUGTCCATGTCCCAGGC-3' (SEQ ID NO: 47), 5'-GCGGUAUCCAUGUCCCAGGC-3' (SEQ ID NO: 151), and 5'-GGUATCCCCCCCCCCCCCCC-3' (SEQ ID NO: 54).

[0046] In other embodiments, one or more candidate oligonucleotides or modified oligonucleotides inhibit cGAS and TLR9 activity. Exemplary motifs for such embodiments include 5'-CUUGUGAAAAGATTAUCUUC-3' (SEQ ID NO:27), 5'-CCAUGTCCCAGGCCTCCAGU-3' (SEQ ID NO:29), 5'-GCAAGGCAGAGAAACUCCAG-3' (SEQ ID NO:37), 5'-GGAUUAAAACAGATTAAUAC-3' (SEQ ID NO:55), 5'-AGCCGAACAGAAGGAGCGUC-3' (SEQ ID NO:40), 5'-UCCGGCCTCGGAAGCUCUCU-3' (SEQ ID NO:10), 5'-UCCGGCCTCGGAGTCUCCAU-3' (SEQ ID NO:52), and 5'-GCGGUATCCATAGTCUCCAU-3' (SEQ ID NO:48).

[0047] In some embodiments, one or more candidate oligonucleotides or modified oligonucleotides inhibit cGAS and TLR9 activity but do not substantially inhibit TLR7 activity. An example motif for such embodiments includes 5'-CUUGUGAAAAGATTAUCUUC-3' (SEQ ID NO: 27).

[0048] In further embodiments, one or more candidate oligonucleotides or modified oligonucleotides inhibit cGAS, TLR7, and TLR9 activity. Exemplary motifs for such embodiments include 5'-CCAUGTCCCAGGCCTCCAGU-3' (SEQ ID NO:29), 5'-GCAAGGCAGAGAAACUCCAG-3' (SEQ ID NO:37), 5'-GGAUUAAAACAGATTAAUAC-3' (SEQ ID NO:55), 5'-AGCCGAACAGAAGGAGCGUC-3' (SEQ ID NO:40), 5'-UCCGGCCTCGGAAGCUCUCU-3' (SEQ ID NO:10), 5'-UCCGGCCTCGGAGTCUCCAU-3' (SEQ ID NO:52), and 5'-GCGGUATCCATAGTCUCCAU-3' (SEQ ID NO:48).

[0049] In alternative embodiments, one or more candidate oligonucleotides or modified oligonucleotides inhibit cGAS but do not substantially inhibit TLR7 and / or TLR9.

[0050] In alternative embodiments, one or more candidate or modified oligonucleotides that inhibit cGAS comprise, consist essentially of, or consist of any of the sequences in Tables 1, 1A, 2, 2A, 3, 3A, 4, 4A, 5, 5A, or 6, with or without the specific modifications defined, and are shown in the Examples to inhibit the activity of cGAS.

[0051] In alternative embodiments, one or more candidate or modified oligonucleotides that inhibit cGAS comprise or consist of any of the sequences in Tables 1, 1A, 2, 2A, 3, 3A, 4, 4A, 5, 5A or 6, with or without the specific modifications defined, and are shown in the Examples to inhibit the activity of cGAS, and the oligonucleotides inhibit or do not substantially inhibit TLR3, TLR7, TLR8 and / or TLR9 activity.

[0052] With respect to the above two aspects, the method may include the further step of testing the ability of one or more candidate or modified oligonucleotides to enhance TLR8 activity, and optionally selecting oligonucleotides that enhance or do not substantially enhance TLR8 activity. Accordingly, in some embodiments, one or more candidate or modified oligonucleotides inhibit cGAS activity and enhance TLR8 activity. In other embodiments, one or more candidate or modified oligonucleotides inhibit cGAS activity and do not substantially enhance TLR8 activity.

[0053] In any embodiment, the candidate oligonucleotide or modified oligonucleotide that inhibits cGAS activity is at least 15, 16, 17, 18, 19, or 20 nucleotides in length. In any embodiment, the candidate oligonucleotide or modified oligonucleotide that inhibits cGAS activity is at least 15 nucleotides in length but no more than 20 nucleotides in length.

[0054] In yet another aspect, the present invention provides a method for selecting or designing an oligonucleotide that inhibits TLR9 activity, comprising the steps of: i) 5'-G[G / C]CCT[C / G]-3' (SEQ ID NO: 152), 5'-CUU-3' (SEQ ID NO: 153), wherein the motif is within 10 bases of the 5' and / or 3' end of the oligonucleotide; 5'-CUUGUGAAAAGATTAUCUUC-3' (SEQ ID NO: 27), 5'-CUUCUCTCTGGTCCCAUCCC-3' (SEQ ID NO: 154), 5'-CCUUCTCTCTGGTCCCAUCC-3' (SEQ ID NO: 142), 5'-CCCUUCTCTCTGGTCCCAUC-3' (SEQ ID NO: 155), 5'-ACCCUTCTCTCTGGTCCCAU-3' (SEQ ID NO: 156), 5'-CUUCCACAATCAAGACAUUC-3' (SEQ ID NO: 157), 5'-CUUCGTGGGGTCCTTUUCAC-3' (SEQ ID NO: 158), 5'-CACUUCGTGGGGTCCUUUUC-3' (SEQ ID NO: 159), 5'-CCAACACTTCGTGGGGUCCU-3' (SEQ ID NO: 160), 5'-UCCGGCCTCGGAAGCUCUCU-3' (SEQ ID NO: 10), 5'-CCAUGTCCCAGGCCTCCAGU-3' (SEQ ID NO: 29), 5'-UUGGCCTGTGGATGCUUUGU-3' (SEQ ID NO: 161), 5'-AAAUGTCCTGGCCCTCACUG-3' (SEQ ID NO: 162), 5'-UCCGGCCTCGGAGTCUCCAU-3' (SEQ ID NO: 52), 5'-UCCGGCCTCGGCAGAUAUCG-3' (SEQ ID NO: 163), 5'-GGCCGAACTTTCCCGCCUUA-3' (SEQ ID NO: 12), 5'-GGUCUTGGCTTCGTGGAGCA-3' (SEQ ID NO: 13), 5'-GGAGCTTCGAGGCCCCAGGC-3' (SEQ ID NO: 14), 5'-GGUGGTCCACAACCCCUUUC-3' (SEQ ID NO: 15), 5'-CAUUAGGTGCAGAAAUCUUC-3' (SEQ ID NO: 16), 5'-UUCUGGGGACTTCCAGUUUA-3' (SEQ ID NO: 17), 5'-UGAUUCCAAAGCCAGGGUUA-3' (SEQ ID NO: 18), 5'-UCCGGGTCGTAGTTGCUUCC-3' (SEQ ID NO: 51), 5'-CCUAGAAAGAAGCAAAGAUU-3' (SEQ ID NO: 164), 5'-GAUUAAAACAGATTAAUACA-3' (SEQ ID NO: 165), 5'-GGAUUAAAACAGATTAAUAC-3' (SEQ ID NO: 55), 5'-AAUUUAAAGCATGAAUAUUA-3' (SEQ ID NO: 166), 5'-GCGUAGTTTCTCTTCCUCCC-3' (SEQ ID NO: 41), 5'-UGACAAAACAATAATAACAG-3' (SEQ ID NO: 167), 5'-ACA-3' (SEQ ID NO: 168), wherein the motif is at or towards the 5' end of the oligonucleotide; 5'-CAC-3' (SEQ ID NO: 169), wherein the motif is at or towards the 5' end of the oligonucleotide; 5'-ACACTTCGTGGGGTCCTTTT-3' (SEQ ID NO: 170), 5'-GCGTGTCTGGAAGCTTCCTT-3' (SEQ ID NO: 86), 5'-TCAAAGGACTGAGGAAAGGG-3' (SEQ ID NO: 171), 5'-ATCCAACACTTCGTGGGGTC-3' (SEQ ID NO: 172), 5'-GCCCATCCATGAGGTCCTGG-3' (SEQ ID NO: 173), 5'-GGGTATCGAAAGAGTCTGGA-3' (SEQ ID NO: 174), 5'-GGTTTTGGCTGGGATCAAGT-3' (SEQ ID NO: 175), 5'-GCGACTATACGCGCAATATG-3' (SEQ ID NO: 176), 5'-ACTGACTGTCTTGAGGGTTC-3' (SEQ ID NO: 85), 5'-AACACTTCGTGGGGTCCTTT-3' (SEQ ID NO: 177), 5'-GTCCAAGATCAGCAGTCTCA-3' (SEQ ID NO: 178), 5'-ACAGTGTTGAGATACTCGGG-3' (SEQ ID NO: 91), 5'-TGGGCTGGAATCCGAGTTAT-3' (SEQ ID NO: 179), 5'-CGGCATCCACCACGTCGTCC-3' (SEQ ID NO: 180), 5'-GCGTATTATAGCCGATTAAC-3' (SEQ ID NO: 181), 5'-GGAGGTCTTGGCTTCGTGGA-3' (SEQ ID NO: 182), 5'-TGGGTTACGGCTCAGTATGG-3' (SEQ ID NO: 183), 5'-CCGCCATGTTTCTTCTTGGA-3' (SEQ ID NO: 184), 5'-AGCTTCGAGGCCCCAG-3' (SEQ ID NO: 185), 5'-GCCATGTTTCTTCTTG-3' (SEQ ID NO: 186), 5'-CACTTCGTGGGGTCCT-3' (SEQ ID NO: 187), 5'-CGGCCTCGGAAGCTCT-3' (SEQ ID NO: 188), 5'-ACACTTCGTGGGGTCC-3' (SEQ ID NO: 110), 5'-TGCACACTTCGTACCC-3' (SEQ ID NO: 189), 5'-CCACATCCTGTGGCTC-3' (SEQ ID NO: 190), 5'-CTGCAGCTTCCTTGTC-3' (SEQ ID NO: 191), 5'-ACTTCGTGGGGTCCTT-3' (SEQ ID NO: 192), 5'-CCCACTTGGCAGACCA-3' (SEQ ID NO: 193), 5'-GTCCCCTGTTGACTGG-3' (SEQ ID NO: 194), 5'-ACGTTCAGTCCTGTCC-3' (SEQ ID NO: 195), 5'-GGTCATTACAATAGCT-3' (SEQ ID NO: 196), 5'-TCGTGGGGTCCTTTTC-3' (SEQ ID NO: 197), 5'-GTGTCTGGAAGCTTCC-3' (SEQ ID NO: 106), 5'-ATGGCCTCCCATCTCC-3' (SEQ ID NO: 104), 5'-TGCTCCTCGGTCTCCC-3' (SEQ ID NO: 198), 5'-GCATCCACCACGTCGT-3' (SEQ ID NO: 199), 5'-CTTCGTGGGGTCCTTT-3' (SEQ ID NO: 200), 5'-AGGCCCTTCGCACTTC-3' (SEQ ID NO: 201), 5'-GCGGUATCCATGTCCCAGGC-3', 5'-GCUGUTTCCATGTCCCAGGC-3', 5'-GCUGUGTCCATGTCCCAGGC-3' 5'-GCCGUTTCCATGTCCCAGGC-3', 5'-GCGGUATCC-3', 5'-GCUGUTTCC-3', 5'-GCUGUGTCC-3', 5'-GCCGUTTCC-3', 5'-CUUCGTGGGGTCCTTUUCAC, 5'-CUUCGTGGG-3', 5'-UCG-3', 5'-ACG-3', 5'-ACC-3', 5'-CGC-3', 5'-GAU-3', 5'-GGG-3', 5'-AGC-3', 5'-UUC-3', 5'-UUG-3', 5'-CAC-3', and scanning the polynucleotide or its complement for a region having a variant of that motif or sequence having at least about 75% sequence identity thereto (where U can be T and / or T can be U); ii) generating one or more candidate oligonucleotides containing the motif; iii) testing the ability of one or more candidate oligonucleotides to inhibit TLR9 activity; and iv) selecting an oligonucleotide that inhibits TLR9 activity.

[0055] In a related aspect, the invention provides a method for increasing the TLR9 inhibitory activity of an oligonucleotide by modifying the oligonucleotide, wherein the modified oligonucleotide comprises: 5'-G[G / C]CCT[C / G]-3' (SEQ ID NO: 152), 5'-CUU-3' (SEQ ID NO: 153), wherein the motif is within 10 bases of the 5' and / or 3' end of the oligonucleotide; 5'-CUUGUGAAAAGATTAUCUUC-3' (SEQ ID NO: 27), 5'-CUUCUCTCTGGTCCCAUCCC-3' (SEQ ID NO: 154), 5'-CCUUCTCTCTGGTCCCAUCC-3' (SEQ ID NO: 142), 5'-CCCUUCTCTCTGGTCCCAUC-3' (SEQ ID NO: 155), 5'-ACCCUTCTCTCTGGTCCCAU-3' (SEQ ID NO: 156), 5'-CUUCCACAATCAAGACAUUC-3' (SEQ ID NO: 157), 5'-CUUCGTGGGGTCCTTUUCAC-3' (SEQ ID NO: 158), 5'-CACUUCGTGGGGTCCUUUUC-3' (SEQ ID NO: 159), 5'-CCAACACTTCGTGGGGUCCU-3' (SEQ ID NO: 160), 5'-UCCGGCCTCGGAAGCUCUCU-3' (SEQ ID NO: 10), 5'-CCAUGTCCCAGGCCTCCAGU-3' (SEQ ID NO: 29), 5'-UUGGCCTGTGGATGCUUUGU-3' (SEQ ID NO: 161), 5'-AAAUGTCCTGGCCCTCACUG-3' (SEQ ID NO: 162), 5'-UCCGGCCTCGGAGTCUCCAU-3' (SEQ ID NO: 52), 5'-UCCGGCCTCGGCAGAUAUCG-3' (SEQ ID NO: 163), 5'-GGCCGAACTTTCCCGCCUUA-3' (SEQ ID NO: 12), 5'-GGUCUTGGCTTCGTGGAGCA-3' (SEQ ID NO: 13), 5'-GGAGCTTCGAGGCCCCAGGC-3' (SEQ ID NO: 14), 5'-GGUGGTCCACAACCCCUUUC-3' (SEQ ID NO: 15), 5'-CAUUAGGTGCAGAAAUCUUC-3' (SEQ ID NO: 16), 5'-UUCUGGGGACTTCCAGUUUA-3' (SEQ ID NO: 17), 5'-UGAUUCCAAAGCCAGGGUUA-3' (SEQ ID NO: 18), 5'-UCCGGGTCGTAGTTGCUUCC-3' (SEQ ID NO: 51), 5'-CCUAGAAAGAAGCAAAGAUU-3' (SEQ ID NO: 164), 5'-GAUUAAAACAGATTAAUACA-3' (SEQ ID NO: 165), 5'-GGAUUAAAACAGATTAAUAC-3' (SEQ ID NO: 55), 5'-AAUUUAAAGCATGAAUAUUA-3' (SEQ ID NO: 166), 5'-GCGUAGTTTCTCTTCCUCCC-3' (SEQ ID NO: 41), 5'-UGACAAAACAATAATAACAG-3' (SEQ ID NO: 167), 5'-ACA-3' (SEQ ID NO: 168), wherein the motif is at or towards the 5' end of the oligonucleotide; 5'-CAC-3' (SEQ ID NO: 169), wherein the motif is at or towards the 5' end of the oligonucleotide; 5'-ACACTTCGTGGGGTCCTTTT-3' (SEQ ID NO: 170), 5'-GCGTGTCTGGAAGCTTCCTT-3' (SEQ ID NO: 86), 5'-TCAAAGGACTGAGGAAAGGG-3' (SEQ ID NO: 171), 5'-ATCCAACACTTCGTGGGGTC-3' (SEQ ID NO: 172), 5'-GCCCATCCATGAGGTCCTGG-3' (SEQ ID NO: 173), 5'-GGGTATCGAAAGAGTCTGGA-3' (SEQ ID NO: 174), 5'-GGTTTTGGCTGGGATCAAGT-3' (SEQ ID NO: 175), 5'-GCGACTATACGCGCAATATG-3' (SEQ ID NO: 176), 5'-ACTGACTGTCTTGAGGGTTC-3' (SEQ ID NO: 85), 5'-AACACTTCGTGGGGTCCTTT-3' (SEQ ID NO: 177), 5'-GTCCAAGATCAGCAGTCTCA-3' (SEQ ID NO: 178), 5'-ACAGTGTTGAGATACTCGGG-3' (SEQ ID NO: 91), 5'-TGGGCTGGAATCCGAGTTAT-3' (SEQ ID NO: 179), 5'-CGGCATCCACCACGTCGTCC-3' (SEQ ID NO: 180), 5'-GCGTATTATAGCCGATTAAC-3' (SEQ ID NO: 181), 5'-GGAGGTCTTGGCTTCGTGGA-3' (SEQ ID NO: 182), 5'-TGGGTTACGGCTCAGTATGG-3' (SEQ ID NO: 183), 5'-CCGCCATGTTTCTTCTTGGA-3' (SEQ ID NO: 184), 5'-AGCTTCGAGGCCCCAG-3' (SEQ ID NO: 185), 5'-GCCATGTTTCTTCTTG-3' (SEQ ID NO: 186), 5'-CACTTCGTGGGGTCCT-3' (SEQ ID NO: 187), 5'-CGGCCTCGGAAGCTCT-3' (SEQ ID NO: 188), 5'-ACACTTCGTGGGGTCC-3' (SEQ ID NO: 110), 5'-TGCACACTTCGTACCC-3' (SEQ ID NO: 189), 5'-CCACATCCTGTGGCTC-3' (SEQ ID NO: 190), 5'-CTGCAGCTTCCTTGTC-3' (SEQ ID NO: 191), 5'-ACTTCGTGGGGTCCTT-3' (SEQ ID NO: 192), 5'-CCCACTTGGCAGACCA-3' (SEQ ID NO: 193), 5'-GTCCCCTGTTGACTGG-3' (SEQ ID NO: 194), 5'-ACGTTCAGTCCTGTCC-3' (SEQ ID NO: 195), 5'-GGTCATTACAATAGCT-3' (SEQ ID NO: 196), 5'-TCGTGGGGTCCTTTTC-3' (SEQ ID NO: 197), 5'-GTGTCTGGAAGCTTCC-3' (SEQ ID NO: 106), 5'-ATGGCCTCCCATCTCC-3' (SEQ ID NO: 104), 5'-TGCTCCTCGGTCTCCC-3' (SEQ ID NO: 198), 5'-GCATCCACCACGTCGT-3' (SEQ ID NO: 199), 5'-CTTCGTGGGGTCCTTT-3' (SEQ ID NO: 200), 5'-AGGCCCTTCGCACTTC-3' (SEQ ID NO: 201), 5'-GCGGUATCCATGTCCCAGGC-3', 5'-GCUGUTTCCATGTCCCAGGC-3', 5'-GCUGUGTCCATGTCCCAGGC-3' 5'-GCCGUTTCCATGTCCCAGGC-3', 5'-GCGGUATCC-3', 5'-GCUGUTTCC-3', 5'-GCUGUGTCC-3', 5'-GCCGUTTCC-3', 5'-CUUCGTGGGGTCCTTUUCAC, 5'-CUUCGTGGG-3', 5'-UCG-3', 5'-ACG-3', 5'-ACC-3', 5'-CGC-3', 5'-GAU-3', 5'-GGG-3', 5'-AGC-3', 5'-UUC-3', 5'-UUG-3', a motif comprising a sequence selected from the group consisting of: 5'-CAC-3'; and and modifying the oligonucleotide to include a variant of that motif or sequence having at least about 75% sequence identity thereto (U can be T and / or T can be U).

[0056] In one embodiment, the method of the invention further comprises testing the ability of the modified oligonucleotides to inhibit TLR9 activity and selecting those oligonucleotides that inhibit TLR9 activity to a greater extent than the unmodified oligonucleotides.

[0057] In one embodiment, the step of modifying the oligonucleotide comprises adding a sequence of nucleotides to the 5' and / or 3' end of the oligonucleotide such that the modified oligonucleotide comprises a motif.

[0058] In one embodiment, the step of modifying the oligonucleotide comprises adding a sequence of nucleotides to the 5' end of the oligonucleotide such that the modified oligonucleotide comprises the motif 5'-ACA-3' (SEQ ID NO: 168), 5'-CAC-3' (SEQ ID NO: 169), 5'-UCG-3', 5'-ACG-3', 5'-ACC-3', 5'-CGC-3', 5'-GAU-3', 5'-GGG-3', 5'-AGC-3', 5'-UUC-3', 5'-UUG-3', or 5'-CAC-3'. In some embodiments, the step of modifying the oligonucleotide comprises adding 5'-ACA-3' (SEQ ID NO: 168), 5'-CAC-3' (SEQ ID NO: 169), 5'-UCG-3', 5'-ACG-3', 5'-ACC-3', 5'-CGC-3', 5'-GAU-3', 5'-GGG-3', 5'-AGC-3', 5'-UUC-3', 5'-UUG-3', or 5'-CAC-3', or a portion or fragment thereof, to the 5' end of the oligonucleotide.

[0059] In one embodiment, modifying the oligonucleotide comprises adding a sequence of nucleotides to the 5' end of the oligonucleotide such that the modified oligonucleotide comprises the motif 5'-GCGGUATCC-3', 5'-GCUGUTTCC-3', 5'-GCUGUGTCC-3', 5'-GCCGUTTCC-3', or 5'-CUUCGTGGGGTCCTTUUCAC-3', or 5'-CUUCGTGGG-3'. In some embodiments, modifying the oligonucleotide comprises adding 5'-GCGGUATCC-3', 5'-GCUGUTTCC-3', 5'-GCUGUGTCC-3', 5'-GCCGUTTCC-3', 5'-CUUCGTGGGGTCCTTUUCAC-3', 5'-CUUCGTGGG-3', or a portion or fragment thereof, to the 5' end of the oligonucleotide.

[0060] In one embodiment of the two foregoing aspects, the oligonucleotide does not bind or is not designed to bind to a transcript encoding TLR9 or its complement.

[0061] In one embodiment of the two foregoing aspects, the oligonucleotide binds to, or is designed to bind to, a target transcript that does not encode TLR9 or its complement.

[0062] In alternative embodiments of the two foregoing aspects, the oligonucleotide binds to or is designed to bind to a target transcript encoding TLR9 or its complement.

[0063] For the two aforementioned embodiments, the motif is preferably within 10 bases of the 5' and / or 3' end of the oligonucleotide. More particularly, the motif is preferably within 5 bases of the 5' and / or 3' end of the oligonucleotide. Even more particularly, the motif is preferably at or toward the 5' and / or 3' end of the oligonucleotide. Even more particularly, the motif is preferably at or toward the 5' end of the oligonucleotide.

[0064] Examples of motifs of the above two embodiments include, but are not limited to, motifs having the sequence 5'-CUU-3' (SEQ ID NO: 153), 5'-CUT-3' (SEQ ID NO: 202), 5'-CTT-3' (SEQ ID NO: 203), 5'-UCG-3', 5'-ACG-3', 5'-ACC-3', 5'-CGC-3', 5'-GAU-3', 5'-GGG-3', 5'-AGC-3', 5'-UUC-3', 5'-UUG-3', or 5'-CAC-3'.

[0065] Further examples of motifs of the above two embodiments include, but are not limited to, motifs having the sequence 5'-GGCCTC-3' (SEQ ID NO: 204), 5'-GGCCTG-3' (SEQ ID NO: 205), or 5'-GCCCTC-3' (SEQ ID NO: 206) (T can also be U).

[0066] Additional examples of motifs according to the above two embodiments include, but are not limited to, motifs having the sequence 5'-ACA-3' (SEQ ID NO: 168) or 5'-CAC-3' (SEQ ID NO: 169).

[0067] In one particular embodiment, the motif of the above aspect has the sequence 5'-GGCCTC-3' (SEQ ID NO: 204), 5'-GGCCUC-3' (SEQ ID NO: 207), 5'-UCCGGCCTCGGAAGCUCUCU-3' (SEQ ID NO: 10), or a variant thereof having at least about 75% sequence identity thereto (U can be T and / or T can be U).

[0068] In one embodiment of the two aforementioned aspects, one or more of the bases of the motif are modified bases and / or have a modified backbone.

[0069] In one embodiment of the two foregoing aspects, the motif has the sequence 5'-mCmUmU-3', 5'-mCmUT-3', 5'-mUmCmG-3', 5'-mAmCmG-3', 5'-mAmCmC-3', 5'-mCmGmC-3', 5'-mGmAmU-3', 5'-mGmGmG-3', 5'-mAmGmC-3', 5'-mUmUmC-3', 5'-mUmUmG-3' or 5'-mCmAmC-3' (wherein m is a modified base and / or has a modified backbone). In such an embodiment, m is suitably a 2'-OMe modified base.

[0070] In one embodiment of the two aforementioned aspects, the motif has the sequence 5'-mGmGCCTC-3', 5'-GGCCTmC-3', 5'-mGmGmCCTG-3' or 5'-GCCCTmC-3' (T can be U and m is a modified base and / or has a modified backbone). In such an embodiment, m is suitably a 2'-OMe modified base.

[0071] In one embodiment of the above two aspects, the motif has the sequence 5'-mAmCmA-3' or 5'-mCmAmC-3', where m is a modified base and / or has a modified backbone. In such examples, m is suitably a 2'-LNA, 2'-MOE and / or 2'-OMe modified base.

[0072] In one particular embodiment, the motif of the above aspect has the sequence 5'-mGmGCCTC-3', 5'-mGmGCCUC-3', 5'-mUmCmCmGmGCCTCGGAAGCmUmCmUmCmU-3', or a variant thereof having at least about 75% sequence identity thereto (U can be T and / or T can be U, m is a modified base and / or has a modified backbone).

[0073] In one embodiment of the above two aspects, the motif has the sequence: 5'-CTTCGTGGGGTCCTTT-3' (SEQ ID NO: 200), 5'-CACTTCGTGGGGTCCT-3' (SEQ ID NO: 187), 5'-ACACTTCGTGGGGTCC-3' (SEQ ID NO: 110), 5'-TGCACACTTCGTACCC-3' (SEQ ID NO: 189), 5'-CGGCCTCGGAAGCTCT-3' (SEQ ID NO: 188), 5'-AGCTTCGAGGCCCCAG-3' (SEQ ID NO: 185), 5'-GCCATGTTTCTTCTTG-3' (SEQ ID NO: 186), or 5'-CTGCAGCTTCCTTGTC-3' (SEQ ID NO: 191), (U can be T and / or T can be U).

[0074] In particular embodiments of the above two aspects, the motif has the sequence: 5'-mCmTmTCGTGGGGGTCCmTmTmT-3', 5'-mCmAmCTTCGTGGGGGTmCmCmT-3', 5'-mAmCmACTTCGTGGGGmTmCmC-3', 5'-mTmGmCACACTTCGTAmCmCmC-3', 5'-mCmGmGCCTCGGAAGCmTmCmT-3', 5'-mAmGmCTTCGAGGCCCmCmAmG-3', 5'-mGmCmCATGTTTCTTCmTmTmG-3', or 5'-mCmTmGCAGCTTCCTTmGmTmC-3' (U may be T and / or T may be U, and m is a modified base and / or has a modified backbone). In such examples, m is suitably a 2'-LNA modified base.

[0075] In another embodiment of the above two aspects, the motif has the sequence: 5'-ACACTTCGTGGGGTCCTTTT-3' (SEQ ID NO: 170), 5'-AACACTTCGTGGGGTCCTTT-3' (SEQ ID NO: 177), 5'-CAACACTTCGTGGGGTCCTT-3' (SEQ ID NO: 208), 5'-CCAACACTTCGTGGGGTCCT-3' (SEQ ID NO: 209), 5'-GCGTGTCTGGAAGCTTCCTT-3' (SEQ ID NO: 86), 5'-GCCCATCCATGAGGTCCTGG-3' (SEQ ID NO: 173), 5'-GGGTATCGAAAGAGTCTGGA-3' (SEQ ID NO: 174), 5'-TGGGCTGGAATCCGAGTTAT-3' (SEQ ID NO: 179), 5'-GGTTTTGGCTGGGATCAAGT-3' (SEQ ID NO: 175), 5'-TCAAAGGACTGAGGAAAGGG-3' (SEQ ID NO: 171), 5'-TCCGGCCTCGGAAGCTCTCT-3' (SEQ ID NO: 138), 5'-GGTGGTCCACAACCCCTTTC-3' (SEQ ID NO: 210), 5'-ACTGACTGTCTTGAGGGTTC-3' (SEQ ID NO: 85), 5'-GCGTATTATAGCCGATTAAC-3' (SEQ ID NO: 181), or 5'-GCGACTATACGCGCAATATG-3' (SEQ ID NO: 176), (U can be T and / or T can be U).

[0076] In particular embodiments of the above two aspects, the motif has the sequence: 5'-mAmCmAmCmTTCGTGGGGTCmCmTmTmTmT-3', 5'-mAmAmCmAmCTTCGTGGGGTmCmCmTmTmT-3', 5'-mCmAmAmCmACTTCGTGGGGmTmCmCmTmT-3', 5'-mCmCmAmAmCACTTCGTGGGmGmTmCmCmT-3', 5'-mGmCmGmTmGTCTGGAAGCTmTmCmCmTmT-3', 5'-mGmCmCmCmATCCATGAGGTmCmCmTmGmG-3', 5'-mGmGmGmTmATCGAAAGAGTmCmTmGmGmA-3', 5'-mTmGmGmGmCTGGAATCCGAmGmTmTmAmT-3', 5'-mGmGmTmTmTTGGCTGGGATmCmAmAmGmT-3', 5'-mTmCmAmAmAGGACTGAGGAmAmAmGmGmG-3', 5'-mTmCmCmGmGCCTCGGAAGCmTmCmTmCmT-3', 5'-mGmGmTmGmGTCCACAACCCmCmTmTmTmC-3', 5'-mAmCmTmGmACTGTCTTGAGmGmGmTmTmC-3', 5'-mGmCmGmTmATTATAGCCGAmTmTmAmAmC-3', or 5'-mGmCmGmAmCTATACGCGCAmAmTmAmTmG-3' (U may be T and / or T may be U, and m is a modified base and / or has a modified backbone). In such examples, m is preferably a 2'-MOE modified base.

[0077] In particular embodiments of the above two aspects, the motif has the sequence: 5'-mUmCmCmGmGCCTCGGAAGCmUmCmUmCmU-3', 5'-mUmCmCmGmGCCTCGGAGTCmUmCmCmAmU-3', 5'-mUmCmCmGmGCCTCGGCAGAmUmAmUmCmG-3', 5'mGmCmGmGmUATCCATGTCCmCmAmGmGmC-3', 5'-mGmCmUmGmUTTCCATGTCCmCmAmGmGmC-3', 5'-mGmCmUmGmUGTCCATGTCCmCmAmGmGmC-3', 5'-mGmCmCmGmUTTCCATGTCCmCmAmGmGmC-3', 5'-mGmCmGmGmUATCC-3', 5'-mGmCmUmGmUTTCC-3', 5'-mGmCmUmGmUGTCC-3', 5'-mGmCmCmGmUTTCC-3', 5'-mCmUmUmCmGTGGGGTCCTTmUmUmCmAmC, and 5'-mCmUmUmCmGTGGG-3' (T may be U, and m is a modified base and / or has a modified backbone). Preferably, each base has a modified backbone, and the modified backbone is phosphorothioate.

[0078] In particular embodiments of the above two aspects, the motif has the sequence: 5'-mG * mC * mG * mG * mU * A * T * C* C * A * T * G * T * C * C * mC * mA * mG * mG * mC-3’、 5’-mU * mC * mC * mG * mG * C * C * T * C * G * G * A * A * G * C * mU * mC * mU * mC * mU-3’、 5’-mU * mC * mC * mG * mG * C * C * T * C * G * G * A * G * T * C * mU * mC * mC * mA * mU-3’、 5’-mU * mC * mC * mG * mG * C * C * T * C * G * G * C * A * G * A *mU * mA * mU * mC * mG-3’、 5’-mG * mC * mU * mG * mU * T * T * C * C * A * T * G * T * C * C * mC * mA * mG * mG * mC-3’、 5’-mG * mC * mU * mG * mU * G * T * C * C * A * T * G * T * C * C * mC * mA * mG * mG * mC-3’、 5’-mG * mC * mC * mG * mU * T * T * C * C * A * T * G * T * C * C * mC * mA * mG * mG * mC-3’、 5’-mG * mC *mG * mG * mU * A * T * C * C * -3’, 5’-mG * mC * mU * mG * mU * T * T * C * C * -3’, 5’-mG * mC * mU * mG * mU * G * T * C * C * -3’, 5’-mG * mC * mC * mG * mU * T * T * C * C * -3’, 5’-mC * mU * mU * mC * mG * T * G * G * G * G * T * C * C[[ID=1​​​​​​​​​​​​​​​​​​​​​​​​​​​​​​​* -3' (U can be T and / or T can be U, and "m" represents a 2'OMe base; * indicates a phosphorothioate backbone).

[0079] In yet another aspect, the present invention provides a method for selecting or designing an oligonucleotide that inhibits TLR9 activity, comprising the steps of: i) scanning the polynucleotide or its complement for regions having at least about 50% adenine bases; ii) generating one or more candidate oligonucleotides comprising a 5' region, a 3' region, and a central region comprising ribonucleic acid, deoxyribonucleic acid, or a combination thereof, optionally bases with a modified backbone, wherein one or both of the 5' region and the 3' region are modified and / or comprise bases with a modified backbone, and at least about 50% of the bases in the central region are adenine bases; iii) testing the ability of one or more candidate oligonucleotides to inhibit TLR9 activity; and and iv) selecting an oligonucleotide that inhibits TLR9 activity.

[0080] In one embodiment, the oligonucleotide comprises a motif having the sequence 5'-[T / G][A / T][G / A / T]AA[A / C][A / G][G / C / A]A[T / G][T / G / C]A[A / T]-3' (SEQ ID NO: 211), where T can also be U.

[0081] Preferably, the 5' and / or 3' regions are about 5 bases in length, the intermediate region is about 10 bases in length, and the intermediate region comprises at least five adenine bases, hi one embodiment, two, three, and / or four of the at least five adenine bases are in a contiguous sequence.

[0082] In one embodiment, the oligonucleotide does not bind or is not designed to bind to transcripts encoding TLR9 or its complement.

[0083] In one embodiment, the oligonucleotide binds to or is designed to bind to a target transcript that does not encode TLR9 or its complement.

[0084] In embodiments of any of the above aspects, the motif comprises, consists essentially of, or consists of any of the sequences in Table 1, 1A, 2, 2A, 3, 3A, 4, 4A, 5, 5A, or 6, with or without the specific modifications defined, and is shown in the Examples to inhibit the activity of TLR9.

[0085] With respect to the above three aspects, the method may include the further step of testing the ability of one or more candidate or modified oligonucleotides to inhibit TLR3, TLR7 and / or cGAS activity, and optionally selecting oligonucleotides that inhibit or do not substantially inhibit TLR3, TLR7 and / or cGAS activity.

[0086] In certain embodiments, one or more candidate oligonucleotides or modified oligonucleotides inhibit TLR9 and TLR7 activity. Exemplary motifs for such embodiments include 5'-CCAUGTCCCAGGCCTCCAGU-3' (SEQ ID NO:29), 5'-GCAAGGCAGAGAAACUCCAG-3' (SEQ ID NO:37), 5'-GGAUUAAAACAGATTAAUAC-3' (SEQ ID NO:55), 5'-AGCCGAACAGAAGGAGCGUC-3' (SEQ ID NO:40), 5'-UCCGGCCTCGGAAGCUCUCU-3' (SEQ ID NO:10), 5'-UCCGGCCTCGGAGTCUCCAU-3' (SEQ ID NO:52), 5'-GCGGUATCCATAGTCUCCAU-3' (SEQ ID NO:48), 5'-GAUUAAAACAGATTAAUACA-3' (SEQ ID NO:165), 5'-UGACAAAACAATAATAACAG-3' (SEQ ID NO:167), and 5'-CCAACACTTCGTGGGGUCCU-3' (SEQ ID NO:160).

[0087] In certain embodiments, one or more candidate oligonucleotides or modified oligonucleotides inhibit TLR9 and TLR7 activity but do not substantially inhibit cGAS activity. Exemplary motifs for such embodiments include 5'-GAUUAAAACAGATTAAUACA-3' (SEQ ID NO: 165) and 5'-UGACAAAACAATAATAACAG-3' (SEQ ID NO: 167).

[0088] In other embodiments, one or more candidate oligonucleotides or modified oligonucleotides inhibit TLR9 and cGAS activity. Exemplary motifs for such embodiments include 5'-CUUGUGAAAAGATTAUCUUC-3' (SEQ ID NO:27), 5'-CCAUGTCCCAGGCCTCCAGU-3' (SEQ ID NO:29), 5'-GCAAGGCAGAGAAACUCCAG-3' (SEQ ID NO:37), 5'-GGAUUAAAACAGATTAAUAC-3' (SEQ ID NO:55), 5'-AGCCGAACAGAAGGAGCGUC-3' (SEQ ID NO:40), 5'-UCCGGCCTCGGAAGCUCUCU-3' (SEQ ID NO:10), 5'-UCCGGCCTCGGAGTCUCCAU-3' (SEQ ID NO:52), and 5'-GCGGUATCCATAGTCUCCAU-3' (SEQ ID NO:48).

[0089] In other embodiments, one or more candidate oligonucleotides or modified oligonucleotides inhibit TLR9 and cGAS activity but do not substantially inhibit TLR7 activity. An example motif for such embodiments includes 5'-CUUGUGAAAAGATTAUCUUC-3' (SEQ ID NO: 27).

[0090] In further embodiments, one or more candidate oligonucleotides or modified oligonucleotides inhibit TLR9, TLR7, and cGAS activity. Exemplary motifs for such embodiments include 5'-CCAUGTCCCAGGCCTCCAGU-3' (SEQ ID NO:29), 5'-GCAAGGCAGAGAAACUCCAG-3' (SEQ ID NO:37), 5'-GGAUUAAAACAGATTAAUAC-3' (SEQ ID NO:55), 5'-AGCCGAACAGAAGGAGCGUC-3' (SEQ ID NO:40), 5'-UCCGGCCTCGGAAGCUCUCU-3' (SEQ ID NO:10), 5'-UCCGGCCTCGGAGTCUCCAU-3' (SEQ ID NO:52), and 5'-GCGGUATCCATAGTCUCCAU-3' (SEQ ID NO:48).

[0091] In alternative embodiments, one or more candidate oligonucleotides or modified oligonucleotides inhibit TLR9 but do not substantially inhibit TLR7 and / or cGAS. An exemplary motif for such embodiments includes 5'-CACUUCGTGGGGTCCUUUUC-3' (SEQ ID NO: 159).

[0092] In alternative embodiments, one or more candidate or modified oligonucleotides that inhibit TLR9 comprise or consist of any of the sequences in Tables 1, 1A, 2, 2A, 3, 3A, 4, 4A, 5, 5A or 6, with or without the specific modifications defined, and are shown in the Examples to inhibit TLR9 activity.

[0093] In alternative embodiments, one or more candidate or modified oligonucleotides that inhibit TLR9 comprise or consist of any of the sequences in Tables 1, 1A, 2, 2A, 3, 3A, 4, 4A, 5, 5A or 6, with or without the specific modifications defined, and are shown in the Examples to inhibit the activity of TLR9, and the oligonucleotides inhibit or do not substantially inhibit TLR3, TLR8, TLR7 and / or cGAS activity.

[0094] With respect to the above two aspects, the method may include the further step of testing the ability of one or more candidate or modified oligonucleotides to enhance TLR8 activity, and optionally selecting oligonucleotides that enhance or do not substantially enhance TLR8 activity. Accordingly, in some embodiments, one or more candidate or modified oligonucleotides inhibit TLR9 activity and enhance TLR8 activity. In other embodiments, one or more candidate or modified oligonucleotides inhibit TLR9 activity and do not substantially enhance TLR8 activity.

[0095] In any embodiment, the candidate or modified oligonucleotide that inhibits TLR9 activity is at least 3, 4, 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, 15, 16, 17, 18, 19, 20, 30, 40, or 50 nucleotides in length. In one embodiment, the candidate or modified oligonucleotide that inhibits TLR9 activity is at least 3 nucleotides in length but no more than 20 nucleotides in length, optionally at least 9 nucleotides in length but no more than 20 nucleotides in length.

[0096] In yet another aspect, the invention provides a method for selecting or designing an oligonucleotide that inhibits TLR7 activity, comprising the steps of: i) 5'-[A / G]GU[A / C][U / C]C-3' (SEQ ID NO: 1), 5'-A[G / A][U / G]C[U / C]C-3' (SEQ ID NO: 2), 5'-A[G / A][U / G]C[U / C]C[U / C][C / A]U-3' (SEQ ID NO: 212), 5'-GGUAUA-3' (SEQ ID NO: 4), 5'-UGUUUC-3' (SEQ ID NO: 5), 5'-UGUGUC-3' (SEQ ID NO: 6), 5'-CGUGUC-3' (SEQ ID NO: 8), 5'-GCAGUCTCCATGTCCCAGGC-3' (SEQ ID NO: 30), 5'-GAUGGTTCCAGTCCCUCUUC-3' (SEQ ID NO: 38), 5'-AGCAGTCTCCATGTCCCAGG-3' (SEQ ID NO: 31), 5'-GGGUTCCTCCACACCCUUC-3' (SEQ ID NO: 36), 5'-GGUGGCCACAGGCAACGUCA-3' (SEQ ID NO: 28), 5'-GCCGUTTCCATGTCCCAGGC-3' (SEQ ID NO: 46), 5'-GCGGUATCCATGTCCCAGGC-3' (SEQ ID NO: 42), 5'-GCGGUATACAGGTCCCAGGC-3' (SEQ ID NO: 43), 5'-GCUGUTTCCATGTCCCAGGC-3' (SEQ ID NO: 44), 5'-GCUGUGTCCATGTCCCAGGC-3' (SEQ ID NO: 45), 5'-GCCGUGTCCATGTCCCAGGC-3' (SEQ ID NO: 47), 5'-GCGGUAUCCAUGUCCCAGGC-3' (SEQ ID NO: 151), 5'-GGUATCCCCCCCCCCCCCCC-3' (SEQ ID NO: 54), 5'-GUCCCATCCCTTCTGCUGCC-3' (SEQ ID NO: 145), 5'-UUCUCTCTGGTCCCAUCCCU-3' (SEQ ID NO: 213), 5'-GUUCAGTCAGATCGCUGGGA-3' (SEQ ID NO: 214), 5'-AUGACATTTCGTGGCUCCUA-3' (SEQ ID NO: 215), 5'-UCUCCATGTCCCAGGCCUCC-3' (SEQ ID NO: 216), 5'-AGUCUCCATGTCCCAGGCCU-3' (SEQ ID NO: 217), 5'-CCAUGTCCCAGGCCTCCAGU-3' (SEQ ID NO: 29), 5'-GCAAGGCAGAGAAACUCCAG-3' (SEQ ID NO: 37), 5'-GGAUUAAAACAGATTAAUAC-3' (SEQ ID NO: 55), 5'-AGCCGAACAGAAGGAGCGUC-3' (SEQ ID NO: 40), 5'-UCCGGCCTCGGAAGCUCUCU-3' (SEQ ID NO: 10), 5'-UCCGGCCTCGGAGTCUCCAU-3' (SEQ ID NO: 52), 5'-GCGGUATCCATAGTCUCCAU-3' (SEQ ID NO: 48), 5'-GAUUAAAACAGATTAAUACA-3' (SEQ ID NO: 165), 5'-UGACAAAACAATAATAACAG-3' (SEQ ID NO: 167), 5'-CCAACACTTCGTGGGGUCCU-3' (SEQ ID NO: 160), 5'-GUGUCCTTCATGCTTUGGAU-3' (SEQ ID NO: 21), 5'-GUCCCAGGCCTCCAGUGUCU-3' (SEQ ID NO: 218), 5'-UUGGCCTGTGGATGCUUUGU-3' (SEQ ID NO: 161), 5'-GUCCGTACCTCCACCCACCG-3' (SEQ ID NO: 219), 5'-GUGUUTTTAATTTTGUAGAG-3' (SEQ ID NO: 220), 5'-GUCAAACCTAGAAAGAAGCA-3' (SEQ ID NO: 221), 5'-GGUCUCCTCCACACCCUUCU-3' (SEQ ID NO: 222), 5'-GUCUCCTCCACACCCUUCUC-3' (SEQ ID NO: 146), 5'-UGAUGATGCTTGCAGGAGGC-3' (SEQ ID NO: 223), 5'-UUAAATAATCTAGTTUGAAG-3' (SEQ ID NO: 20), 5'-AAAGCAGTCTCCATGUCCCA-3' (SEQ ID NO: 224), 5'-GCGUAGTTTCTCTTCCUCCC-3' (SEQ ID NO: 41), 5'-UCUGGTCCCATCCCTUCUGC-3' (SEQ ID NO: 35), 5'-UAUUUCCACATGCCCAGUGU-3' (SEQ ID NO: 225), 5'-UGUUUCCCCGGAGAGCAAUG-3' (SEQ ID NO: 39), 5'-UUAGCTCCTTGCCTCGUUCC-3' (SEQ ID NO: 226), 5'-AUUUCCACATGCCCAGUGUU-3' (SEQ ID NO: 33), 5'-UGGCGTAGTTTCTCTUCCUC-3' (SEQ ID NO: 227), 5'-UGACATTTCGTGGCTCCUAC-3' (SEQ ID NO: 228), 5'-GAAAAGATTATCTTCUUUUA-3' (SEQ ID NO: 25), 5'-UGUGAAAAGATTATCUUCUU-3' (SEQ ID NO: 26), 5'-UUGUGAAAAGATTATCUUCU-3' (SEQ ID NO: 229), 5'-UUUGAAATTCAGAAGAUUUG-3' (SEQ ID NO: 230), 5'-AAGCAGTCTCCATGTCCCAG-3' (SEQ ID NO: 231), 5'-AGGAUTAAAACAGATUAAUA-3' (SEQ ID NO: 232), 5'-AGAUUATCTTCTTTTAAUUU-3' (SEQ ID NO: 23), 5'-UCCCAACTCTTCTAACUCGU-3' (SEQ ID NO: 148), 5'-UAAAATAAGGGGAATAGGGG-3' (SEQ ID NO: 233), 5'-AGUGGCACATACCACACCCU-3' (SEQ ID NO: 32), 5'-AAGAUTATCTTCTTTUAAUU-3' (SEQ ID NO: 234), 5'-AGAAAGAAGCAAAGAUUCAA-3' (SEQ ID NO: 22), 5'-UCCCATCCCTTCTGCUGCCA-3' (SEQ ID NO: 235), 5'-ACCCUTCTCTCTGGTCCCAU-3' (SEQ ID NO: 156), 5'-AAUAUCTGCTGCCCACCUUC-3' (SEQ ID NO: 236), 5'-UCUCUCTGGTCCCATCCCUU-3' (SEQ ID NO: 237), 5'-AGGCCTCCAGTGTCTUCUCC-3' (SEQ ID NO: 238), 5'-CAAGCCCCAGCGTTCCUCCG-3' (SEQ ID NO: 239), 5'-AAAUGTCCTGGCCCTCACUG-3' (SEQ ID NO: 162), 5'-AAUUUAAAGCATGAAUAUUA-3' (SEQ ID NO: 166), 5'-AAAAGATTATCTTCTUUUAA-3' (SEQ ID NO: 24), 5'-AUAUCTGCTGCCCACCUUCU-3' (SEQ ID NO: 144), 5'-CAGUCTCCATGTCCCAGGCC-3' (SEQ ID NO: 240), 5'-AAAGATTATCTTCTTUUAAU-3' (SEQ ID NO: 241), 5'-CCCUUCTCTCTGGTCCCAUC-3' (SEQ ID NO: 155), 5'-AUGGCCTTTCCGTGCCAAGG-3' (SEQ ID NO: 9), 5'-GCAUUCCGTGCGGAAGCCUU-3' (SEQ ID NO: 11), 5'-GGCCGAACTTTCCCGCCUUA-3' (SEQ ID NO: 12), 5'-GGUCUTGGCTTCGTGGAGCA-3' (SEQ ID NO: 13), 5'-GGAGCTTCGAGGCCCCAGGC-3' (SEQ ID NO: 14), 5'-GGUGGTCCACAACCCCUUUC-3' (SEQ ID NO: 15), 5'-UUCUGGGGACTTCCAGUUUA-3' (SEQ ID NO: 17), 5'-UGAUUCCAAAGCCAGGGUUA-3' (SEQ ID NO: 18), 5'-GGGUATCGAAAGAGTCUGGA-3' (SEQ ID NO: 242), 5'-CUUGCACGTGGCTTCGUCUC-3' (SEQ ID NO: 243), 5'-GUGUCCTTGCACGTGGCUUC-3' (SEQ ID NO: 244), 5'-GUAAAAAGCTTTTGAAGUGA-3' (SEQ ID NO: 245), 5'-AUGCCATCCACTTGAUAGGC-3' (SEQ ID NO: 246), 5'-UGAAGTAAAAATCAAUAGCG-3' (SEQ ID NO: 247), 5'-AAGGCCCTTCGCACTUCUUA-3' (SEQ ID NO: 248), 5'-GUACUCGTCGGCATCCACCA-3' (SEQ ID NO: 249), 5'-GUCCUTGCACGTGGCUUCGU-3' (SEQ ID NO: 250), 5'-GCCCATCCATGAGGTCCUGG-3' (SEQ ID NO: 251), 5'-GUAAAAGGAGAAAACUAUCU-3' (SEQ ID NO: 252), 5'-UUGAAGTGAAGTAAAAGGAG-3' (SEQ ID NO: 253), 5'-GUUACTCGTGCCTTGGCAAA-3' (SEQ ID NO: 254), 5'-GUCCAAGATCAGCAGUCUCA-3' (SEQ ID NO: 255), 5'-UUCAATGGGAGAATAAAGCA-3' (SEQ ID NO: 256), 5'-GCAAGGCCCTTCGCACUUCU-3' (SEQ ID NO: 257), 5'-GGGUCCACCACTAGCCAGUA-3' (SEQ ID NO: 258), 5'-GUAGAGAAATTATTTUAGGA-3' (SEQ ID NO: 259), 5'-AUCCACCACGTCGTCCAUGU-3' (SEQ ID NO: 260), 5'-GGCAUCCACCACGTCGUCCA-3' (SEQ ID NO: 261), 5'-UUACUTTAAAAGCAAAAGGA-3' (SEQ ID NO: 262), 5'-UUUGAAGTGAAGTAAAAGGA-3' (SEQ ID NO: 263), 5'-GAAGUGAAGTAAAAGGAGAA-3' (SEQ ID NO: 264), 5'-GGCCATCTCTGCTTCUUGGU-3' (SEQ ID NO: 265), 5'-UGGGCTGGAATCCGAGUUAU-3' (SEQ ID NO: 266), 5'-GGAGATTTCAGAGCAGCUUC-3' (SEQ ID NO: 267), 5'-UUUACGGTTTTCAGAAUAUC-3' (SEQ ID NO: 268), 5'-GCGUGTCTGGAAGCTUCCUU-3' (SEQ ID NO: 269), 5'-GCUUATTTTAAGCATAUUAA-3' (SEQ ID NO: 270), 5'-UUAUUTTAAGCATATUAAAA-3' (SEQ ID NO: 271), 5'-UUCUGCAGCTTCCTTGUCCU-3' (SEQ ID NO: 272), 5'-AUUACTTTAAAAGCAAAAGG-3' (SEQ ID NO: 273), 5'-AUUUUAAGCATATTAAAAAG-3' (SEQ ID NO: 274), 5'-UGUGGCTTGTCCTCAGACAU-3' (SEQ ID NO: 275), 5'-AAAAGGAGAAAACTAUCUUC-3' (SEQ ID NO: 276), 5'-GGGUCCATACCCAAGGCAUC-3' (SEQ ID NO: 277), 5'-ACAGUGTTGAGATACUCGGG-3' (SEQ ID NO: 278), 5'-GGAUCTGCATGCCCTCAUCU-3' (SEQ ID NO: 279), 5'-AGUAAAAAGCTTTTGAAGUG-3' (SEQ ID NO: 280), 5'-GUCGUGGCAAATAGTCCUAG-3' (SEQ ID NO: 281), 5'-GGAGATCAGATGAGAGGAGC-3' (SEQ ID NO: 282), 5'-GUGGUTAAGTACATGAGCUC-3' (SEQ ID NO: 283), 5'-GGACACTTAGCTGTTCCUCG-3' (SEQ ID NO: 284), 5'-GUCUCTACTGTTACCUCUGA-3' (SEQ ID NO: 285), 5'-GAGUUCTTCGTAGGCUUCUG-3' (SEQ ID NO: 286), 5'-AAAGUCAAAAAGAAAAACUG-3' (SEQ ID NO: 287), 5'-AAAAGTGGGAAATAAAGGUU-3' (SEQ ID NO: 288), 5'-AGUUUATAGATTTCAAGUAG-3' (SEQ ID NO: 289), 5'-AAAAAGTGGGAAATAAAGGU-3' (SEQ ID NO: 290), 5'-UUUAUATTACAAAGCUACUU-3' (SEQ ID NO: 291), 5'-UGCUATTCATATTTTUAUUU-3' (SEQ ID NO: 292), 5'-GGUAU-3' (SEQ ID NO: 56), 5'-[G / A / C][G / A][G / A / C][T / A / C]TC-3' (SEQ ID NO: 293), 5'-[G / A / C]G[G / A / C][T / A / C]TC-3' (SEQ ID NO: 294), 5'-GGCTTC-3' (SEQ ID NO: 295), 5'-GGCATC-3' (SEQ ID NO: 296), 5'-AGCTTC-3' (SEQ ID NO: 297), 5'-GGAATC-3' (SEQ ID NO: 298), 5'-CACATC-3' (SEQ ID NO: 299), 5'-GGCCTC-3' (SEQ ID NO: 204), 5'-CACTTC-3' (SEQ ID NO: 300), 5'-AAGATC-3' (SEQ ID NO: 301), 5'-TGTCCTTGCACGTGGCTTCG-3' (SEQ ID NO: 302), 5'-TTTGCACACTTCGTACCCAA-3' (SEQ ID NO: 94), 5'-GTCCACATCCTGTGGCTCGT-3' (SEQ ID NO: 303), 5'-TGTGATGGCCTCCCATCTCC-3' (SEQ ID NO: 304), 5'-GGTTTTGGCTGGGATCAAGT-3' (SEQ ID NO: 175), 5'-GGTGTCCTTGCACGTGGCTT-3' (SEQ ID NO: 93), 5'-GGTCCATACCCAAGGCATCC-3' (SEQ ID NO: 305), 5'-GTGTCTTCATCGGCCCTGCC-3' (SEQ ID NO: 306), 5'-GTCTTGGCTTCGTGGAGCAG-3' (SEQ ID NO: 89), 5'-GCTGACAAAGATTCACTGGT-3' (SEQ ID NO: 95), 5'-GAAAGGTTATGCAAGG-3' (SEQ ID NO: 103), 5'-GACTATACGCGCAATA-3' (SEQ ID NO: 307), 5'-TGTGATGGCCTCCCAT-3' (SEQ ID NO: 308), 5'-TCCAACACTTCGTGGG-3' (SEQ ID NO: 114), 5'-GTGTCTGGAAGCTTCC-3' (SEQ ID NO: 106), 5'-CTTGAAGCATCGTATC-3' (SEQ ID NO: 98), 5'-TCGTAGTTGCTTCCTA-3' (SEQ ID NO: 309), 5'-CGCTTTTCTGTCTGGT-3' (SEQ ID NO: 105), 5'-GGCTGGAATCCGAGTT-3' (SEQ ID NO: 310), 5'-GATAGCACCTTCAGCA-3' (SEQ ID NO: 100), 5'-AGGACTCCAGATGTTT-3' (SEQ ID NO: 311), 5'-GTGATCTTGACATGCT-3' (SEQ ID NO: 312), 5'-AGATTTCAGAGCAGCT-3' (SEQ ID NO: 313), 5'-GGTTACGGCTCAGTAT-3' (SEQ ID NO: 314), 5'-GTTCAGTCCTGTCCAT-3' (SEQ ID NO: 315), 5'-AGGTCTTGGCTTCGTG-3' (SEQ ID NO: 316), 5'-CTGCAGCTTCCTTGTC-3' (SEQ ID NO: 191), 5'-GTCCTTGCACGTGGCT-3' (SEQ ID NO: 317), 5'-GTCTCTGGAGCTTCCT-3' (SEQ ID NO: 318), 5'-GGTCTTGGCTTCGTGG-3' (SEQ ID NO: 319), 5'-GGUA-3' (SEQ ID NO: 57), 5'-GUAU-3' (SEQ ID NO: 58), 5'-GGU-3' (SEQ ID NO: 59), 5'-GUA-3' (SEQ ID NO: 60), 5'-GUC-3', 5'-GUG-3', 5'-GUU-3', 5'-GGC-3', 5'-AUC-3', 5'-GAA-3', 5'-GAG-3', 5'-GGA-3', 5'-GAC-3', 5'-GAU-3', 5'-AUG-3', 5'-GCG-3', 5'-UUC-3', 5'-GCC-3', 5'-GGG-3', 5'-AUU-3', 5'-GCA-3', 5'-AGC-3', 5'-AAC-3', 5'-CCA-3', 5'-UGC-3', 5'-CAA-3', 5'-CGG-3', 5'-ACC-3', 5'-AGA-3', 5'-TTT-3', a motif comprising a sequence selected from the group consisting of: 5'-TCT-3'; and scanning the polynucleotide or its complement for a region having a variant of that motif or sequence having at least about 75% sequence identity thereto (where U can be T and / or T can be U); ii) generating one or more candidate oligonucleotides containing the motif; iii) testing the ability of one or more candidate oligonucleotides to inhibit TLR7 activity; and and iv) selecting an oligonucleotide that inhibits TLR7 activity.

[0097] In one embodiment, step i) comprises scanning the polynucleotide or its complement for a motif having the sequence 5'-GGUAU-3' (SEQ ID NO: 56), a portion or fragment thereof, or a variant having at least about 75% sequence identity thereto (U can be T). Suitably, the 5'-GGUAU-3' (SEQ ID NO: 56) motif is at or towards the 5' end of the oligonucleotide.

[0098] In some embodiments, step i) is selected from the group consisting of 5'-GGUAU-3' (SEQ ID NO: 56), 5'-GGUA-3' (SEQ ID NO: 57), 5'-GUAU-3' (SEQ ID NO: 58), 5'-GGU-3' (SEQ ID NO: 59), 5'-GUA-3' (SEQ ID NO: 60), 5'-GUC-3', 5'-GUG-3', 5'-GUU-3', 5'-GGC-3', 5'-AUC-3', 5'-GAA-3', 5'-GAG-3', 5'-GGA-3', 5'-GAC-3', 5'-GAU-3', 5'-AUG-3', 5'-GCG and scanning the polynucleotide or its complement for a motif having the sequence 5'-UUC-3', 5'-GCC-3', 5'-GGG-3', 5'-AUU-3', 5'-GCA-3', 5'-AGC-3', 5'-AAC-3', 5'-CCA-3', 5'-UGC-3', 5'-CAA-3', 5'-CGG-3', 5'-ACC-3', 5'-AGA-3', 5'-TTT-3', or a variant having at least about 75% sequence identity thereto (where U can be T). Preferably, the motif in such embodiments is at or towards the 5' end of the oligonucleotide.

[0099] In yet another aspect, the present invention provides a method for increasing the TLR7 inhibitory activity of an oligonucleotide by modifying the oligonucleotide, wherein the modified oligonucleotide comprises: 5'-[A / G]GU[A / C][U / C]C-3' (SEQ ID NO: 1), 5'-A[G / A][U / G]C[U / C]C-3' (SEQ ID NO: 2), 5'-A[G / A][U / G]C[U / C]C[U / C][C / A]U-3' (SEQ ID NO: 212), 5'-GGUAUA-3' (SEQ ID NO: 4), 5'-UGUUUC-3' (SEQ ID NO: 5), 5'-UGUGUC-3' (SEQ ID NO: 6), 5'-CGUGUC-3' (SEQ ID NO: 8), 5'-GCAGUCTCCATGTCCCAGGC-3' (SEQ ID NO: 30), 5'-GAUGGTTCCAGTCCCUCUUC-3' (SEQ ID NO: 38), 5'-AGCAGTCTCCATGTCCCAGG-3' (SEQ ID NO: 31), 5'-GGGUTCCTCCACACCCUUC-3' (SEQ ID NO: 36), 5'-GGUGGCCACAGGCAACGUCA-3' (SEQ ID NO: 28), 5'-GCCGUTTCCATGTCCCAGGC-3' (SEQ ID NO: 46), 5'-GCGGUATCCATGTCCCAGGC-3' (SEQ ID NO: 42), 5'-GCGGUATACAGGTCCCAGGC-3' (SEQ ID NO: 43), 5'-GCUGUTTCCATGTCCCAGGC-3' (SEQ ID NO: 44), 5'-GCUGUGTCCATGTCCCAGGC-3' (SEQ ID NO: 45), 5'-GCCGUGTCCATGTCCCAGGC-3' (SEQ ID NO: 47), 5'-GCGGUAUCCAUGUCCCAGGC-3' (SEQ ID NO: 151), 5'-GGUATCCCCCCCCCCCCCCC-3' (SEQ ID NO: 54), 5'-GUCCCATCCCTTCTGCUGCC-3' (SEQ ID NO: 145), 5'-UUCUCTCTGGTCCCAUCCCU-3' (SEQ ID NO: 213), 5'-GUUCAGTCAGATCGCUGGGA-3' (SEQ ID NO: 214), 5'-AUGACATTTCGTGGCUCCUA-3' (SEQ ID NO: 215), 5'-UCUCCATGTCCCAGGCCUCC-3' (SEQ ID NO: 216), 5'-AGUCUCCATGTCCCAGGCCU-3' (SEQ ID NO: 217), 5'-CCAUGTCCCAGGCCTCCAGU-3' (SEQ ID NO: 29), 5'-GCAAGGCAGAGAAACUCCAG-3' (SEQ ID NO: 37), 5'-GGAUUAAAACAGATTAAUAC-3' (SEQ ID NO: 55), 5'-AGCCGAACAGAAGGAGCGUC-3' (SEQ ID NO: 40), 5'-UCCGGCCTCGGAAGCUCUCU-3' (SEQ ID NO: 10), 5'-UCCGGCCTCGGAGTCUCCAU-3' (SEQ ID NO: 52), 5'-GCGGUATCCATAGTCUCCAU-3' (SEQ ID NO: 48), 5'-GAUUAAAACAGATTAAUACA-3' (SEQ ID NO: 165), 5'-UGACAAAACAATAATAACAG-3' (SEQ ID NO: 167), 5'-CCAACACTTCGTGGGGUCCU-3' (SEQ ID NO: 160), 5'-GUGUCCTTCATGCTTUGGAU-3' (SEQ ID NO: 21), 5'-GUCCCAGGCCTCCAGUGUCU-3' (SEQ ID NO: 218), 5'-UUGGCCTGTGGATGCUUUGU-3' (SEQ ID NO: 161), 5'-GUCCGTACCTCCACCCACCG-3' (SEQ ID NO: 219), 5'-GUGUUTTTAATTTTGUAGAG-3' (SEQ ID NO: 220), 5'-GUCAAACCTAGAAAGAAGCA-3' (SEQ ID NO: 221), 5'-GGUCUCCTCCACACCCUUCU-3' (SEQ ID NO: 222), 5'-GUCUCCTCCACACCCUUCUC-3' (SEQ ID NO: 146), 5'-UGAUGATGCTTGCAGGAGGC-3' (SEQ ID NO: 223), 5'-UUAAATAATCTAGTTUGAAG-3' (SEQ ID NO: 20), 5'-AAAGCAGTCTCCATGUCCCA-3' (SEQ ID NO: 224), 5'-GCGUAGTTTCTCTTCCUCCC-3' (SEQ ID NO: 41), 5'-UCUGGTCCCATCCCTUCUGC-3' (SEQ ID NO: 35), 5'-UAUUUCCACATGCCCAGUGU-3' (SEQ ID NO: 225), 5'-UGUUUCCCCGGAGAGCAAUG-3' (SEQ ID NO: 39), 5'-UUAGCTCCTTGCCTCGUUCC-3' (SEQ ID NO: 226), 5'-AUUUCCACATGCCCAGUGUU-3' (SEQ ID NO: 33), 5'-UGGCGTAGTTTCTCTUCCUC-3' (SEQ ID NO: 227), 5'-UGACATTTCGTGGCTCCUAC-3' (SEQ ID NO: 228), 5'-GAAAAGATTATCTTCUUUUA-3' (SEQ ID NO: 25), 5'-UGUGAAAAGATTATCUUCUU-3' (SEQ ID NO: 26), 5'-UUGUGAAAAGATTATCUUCU-3' (SEQ ID NO: 229), 5'-UUUGAAATTCAGAAGAUUUG-3' (SEQ ID NO: 230), 5'-AAGCAGTCTCCATGTCCCAG-3' (SEQ ID NO: 231), 5'-AGGAUTAAAACAGATUAAUA-3' (SEQ ID NO: 232), 5'-AGAUUATCTTCTTTTAAUUU-3' (SEQ ID NO: 23), 5'-UCCCAACTCTTCTAACUCGU-3' (SEQ ID NO: 148), 5'-UAAAATAAGGGGAATAGGGG-3' (SEQ ID NO: 233), 5'-AGUGGCACATACCACACCCU-3' (SEQ ID NO: 32), 5'-AAGAUTATCTTCTTTUAAUU-3' (SEQ ID NO: 234), 5'-AGAAAGAAGCAAAGAUUCAA-3' (SEQ ID NO: 22), 5'-UCCCATCCCTTCTGCUGCCA-3' (SEQ ID NO: 235), 5'-ACCCUTCTCTCTGGTCCCAU-3' (SEQ ID NO: 156), 5'-AAUAUCTGCTGCCCACCUUC-3' (SEQ ID NO: 236), 5'-UCUCUCTGGTCCCATCCCUU-3' (SEQ ID NO: 237), 5'-AGGCCTCCAGTGTCTUCUCC-3' (SEQ ID NO: 238), 5'-CAAGCCCCAGCGTTCCUCCG-3' (SEQ ID NO: 239), 5'-AAAUGTCCTGGCCCTCACUG-3' (SEQ ID NO: 162), 5'-AAUUUAAAGCATGAAUAUUA-3' (SEQ ID NO: 166), 5'-AAAAGATTATCTTCTUUUAA-3' (SEQ ID NO: 24), 5'-AUAUCTGCTGCCCACCUUCU-3' (SEQ ID NO: 144), 5'-CAGUCTCCATGTCCCAGGCC-3' (SEQ ID NO: 240), 5'-AAAGATTATCTTCTTUUAAU-3' (SEQ ID NO: 241), 5'-CCCUUCTCTCTGGTCCCAUC-3' (SEQ ID NO: 155), 5'-AUGGCCTTTCCGTGCCAAGG-3' (SEQ ID NO: 9), 5'-GCAUUCCGTGCGGAAGCCUU-3' (SEQ ID NO: 11), 5'-GGCCGAACTTTCCCGCCUUA-3' (SEQ ID NO: 12), 5'-GGUCUTGGCTTCGTGGAGCA-3' (SEQ ID NO: 13), 5'-GGAGCTTCGAGGCCCCAGGC-3' (SEQ ID NO: 14), 5'-GGUGGTCCACAACCCCUUUC-3' (SEQ ID NO: 15), 5'-UUCUGGGGACTTCCAGUUUA-3' (SEQ ID NO: 17), 5'-UGAUUCCAAAGCCAGGGUUA-3' (SEQ ID NO: 18), 5'-GGGUATCGAAAGAGTCUGGA-3' (SEQ ID NO: 242), 5'-CUUGCACGTGGCTTCGUCUC-3' (SEQ ID NO: 243), 5'-GUGUCCTTGCACGTGGCUUC-3' (SEQ ID NO: 244), 5'-GUAAAAAGCTTTTGAAGUGA-3' (SEQ ID NO: 245), 5'-AUGCCATCCACTTGAUAGGC-3' (SEQ ID NO: 246), 5'-UGAAGTAAAAATCAAUAGCG-3' (SEQ ID NO: 247), 5'-AAGGCCCTTCGCACTUCUUA-3' (SEQ ID NO: 248), 5'-GUACUCGTCGGCATCCACCA-3' (SEQ ID NO: 249), 5'-GUCCUTGCACGTGGCUUCGU-3' (SEQ ID NO: 250), 5'-GCCCATCCATGAGGTCCUGG-3' (SEQ ID NO: 251), 5'-GUAAAAGGAGAAAACUAUCU-3' (SEQ ID NO: 252), 5'-UUGAAGTGAAGTAAAAGGAG-3' (SEQ ID NO: 253), 5'-GUUACTCGTGCCTTGGCAAA-3' (SEQ ID NO: 254), 5'-GUCCAAGATCAGCAGUCUCA-3' (SEQ ID NO: 255), 5'-UUCAATGGGAGAATAAAGCA-3' (SEQ ID NO: 256), 5'-GCAAGGCCCTTCGCACUUCU-3' (SEQ ID NO: 257), 5'-GGGUCCACCACTAGCCAGUA-3' (SEQ ID NO: 258), 5'-GUAGAGAAATTATTTUAGGA-3' (SEQ ID NO: 259), 5'-AUCCACCACGTCGTCCAUGU-3' (SEQ ID NO: 260), 5'-GGCAUCCACCACGTCGUCCA-3' (SEQ ID NO: 261), 5'-UUACUTTAAAAGCAAAAGGA-3' (SEQ ID NO: 262), 5'-UUUGAAGTGAAGTAAAAGGA-3' (SEQ ID NO: 263), 5'-GAAGUGAAGTAAAAGGAGAA-3' (SEQ ID NO: 264), 5'-GGCCATCTCTGCTTCUUGGU-3' (SEQ ID NO: 265), 5'-UGGGCTGGAATCCGAGUUAU-3' (SEQ ID NO: 266), 5'-GGAGATTTCAGAGCAGCUUC-3' (SEQ ID NO: 267), 5'-UUUACGGTTTTCAGAAUAUC-3' (SEQ ID NO: 268), 5'-GCGUGTCTGGAAGCTUCCUU-3' (SEQ ID NO: 269), 5'-GCUUATTTTAAGCATAUUAA-3' (SEQ ID NO: 270), 5'-UUAUUTTAAGCATATUAAAA-3' (SEQ ID NO: 271), 5'-UUCUGCAGCTTCCTTGUCCU-3' (SEQ ID NO: 272), 5'-AUUACTTTAAAAGCAAAAGG-3' (SEQ ID NO: 273), 5'-AUUUUAAGCATATTAAAAAG-3' (SEQ ID NO: 274), 5'-UGUGGCTTGTCCTCAGACAU-3' (SEQ ID NO: 275), 5'-AAAAGGAGAAAACTAUCUUC-3' (SEQ ID NO: 276), 5'-GGGUCCATACCCAAGGCAUC-3' (SEQ ID NO: 277), 5'-ACAGUGTTGAGATACUCGGG-3' (SEQ ID NO: 278), 5'-GGAUCTGCATGCCCTCAUCU-3' (SEQ ID NO: 279), 5'-AGUAAAAAGCTTTTGAAGUG-3' (SEQ ID NO: 280), 5'-GUCGUGGCAAATAGTCCUAG-3' (SEQ ID NO: 281), 5'-GGAGATCAGATGAGAGGAGC-3' (SEQ ID NO: 282), 5'-GUGGUTAAGTACATGAGCUC-3' (SEQ ID NO: 283), 5'-GGACACTTAGCTGTTCCUCG-3' (SEQ ID NO: 284), 5'-GUCUCTACTGTTACCUCUGA-3' (SEQ ID NO: 285), 5'-GAGUUCTTCGTAGGCUUCUG-3' (SEQ ID NO: 286), 5'-AAAGUCAAAAAGAAAAACUG-3' (SEQ ID NO: 287), 5'-AAAAGTGGGAAATAAAGGUU-3' (SEQ ID NO: 288), 5'-AGUUUATAGATTTCAAGUAG-3' (SEQ ID NO: 289), 5'-AAAAAGTGGGAAATAAAGGU-3' (SEQ ID NO: 290), 5'-UUUAUATTACAAAGCUACUU-3' (SEQ ID NO: 291), 5'-UGCUATTCATATTTTUAUUU-3' (SEQ ID NO: 292), 5'-GGUAU-3' (SEQ ID NO: 56), 5'-[G / A / C][G / A][G / A / C][T / A / C]TC-3' (SEQ ID NO: 293), 5'-[G / A / C]G[G / A / C][T / A / C]TC-3' (SEQ ID NO: 294), 5'-GGCTTC-3' (SEQ ID NO: 295), 5'-GGCATC-3' (SEQ ID NO: 296), 5'-AGCTTC-3' (SEQ ID NO: 297), 5'-GGAATC-3' (SEQ ID NO: 298), 5'-CACATC-3' (SEQ ID NO: 299), 5'-GGCCTC-3' (SEQ ID NO: 204), 5'-CACTTC-3' (SEQ ID NO: 300), 5'-AAGATC-3' (SEQ ID NO: 301), 5'-TGTCCTTGCACGTGGCTTCG-3' (SEQ ID NO: 302), 5'-TTTGCACACTTCGTACCCAA-3' (SEQ ID NO: 94), 5'-GTCCACATCCTGTGGCTCGT-3' (SEQ ID NO: 303), 5'-TGTGATGGCCTCCCATCTCC-3' (SEQ ID NO: 304), 5'-GGTTTTGGCTGGGATCAAGT-3' (SEQ ID NO: 175), 5'-GGTGTCCTTGCACGTGGCTT-3' (SEQ ID NO: 93), 5'-GGTCCATACCCAAGGCATCC-3' (SEQ ID NO: 305), 5'-GTGTCTTCATCGGCCCTGCC-3' (SEQ ID NO: 306), 5'-GTCTTGGCTTCGTGGAGCAG-3' (SEQ ID NO: 89), 5'-GCTGACAAAGATTCACTGGT-3' (SEQ ID NO: 95), 5'-GAAAGGTTATGCAAGG-3' (SEQ ID NO: 103), 5'-GACTATACGCGCAATA-3' (SEQ ID NO: 307), 5'-TGTGATGGCCTCCCAT-3' (SEQ ID NO: 308), 5'-TCCAACACTTCGTGGG-3' (SEQ ID NO: 114), 5'-GTGTCTGGAAGCTTCC-3' (SEQ ID NO: 106), 5'-CTTGAAGCATCGTATC-3' (SEQ ID NO: 98), 5'-TCGTAGTTGCTTCCTA-3' (SEQ ID NO: 309), 5'-CGCTTTTCTGTCTGGT-3' (SEQ ID NO: 105), 5'-GGCTGGAATCCGAGTT-3' (SEQ ID NO: 310), 5'-GATAGCACCTTCAGCA-3' (SEQ ID NO: 100), 5'-AGGACTCCAGATGTTT-3' (SEQ ID NO: 311), 5'-GTGATCTTGACATGCT-3' (SEQ ID NO: 312), 5'-AGATTTCAGAGCAGCT-3' (SEQ ID NO: 313), 5'-GGTTACGGCTCAGTAT-3' (SEQ ID NO: 314), 5'-GTTCAGTCCTGTCCAT-3' (SEQ ID NO: 315), 5'-AGGTCTTGGCTTCGTG-3' (SEQ ID NO: 316), 5'-CTGCAGCTTCCTTGTC-3' (SEQ ID NO: 191), 5'-GTCCTTGCACGTGGCT-3' (SEQ ID NO: 317), 5'-GTCTCTGGAGCTTCCT-3' (SEQ ID NO: 318), 5'-GGTCTTGGCTTCGTGG-3' (SEQ ID NO: 319), 5'-GGUA-3' (SEQ ID NO: 57), 5'-GUAU-3' (SEQ ID NO: 58), 5'-GGU-3' (SEQ ID NO: 59), 5'-GUA-3' (SEQ ID NO: 60), 5'-GUC-3', 5'-GUG-3', 5'-GUU-3', 5'-GGC-3', 5'-AUC-3', 5'-GAA-3', 5'-GAG-3', 5'-GGA-3', 5'-GAC-3', 5'-GAU-3', 5'-AUG-3', 5'-GCG-3', 5'-UUC-3', 5'-GCC-3', 5'-GGG-3', 5'-AUU-3', 5'-GCA-3', 5'-AGC-3', 5'-AAC-3', 5'-CCA-3', 5'-UGC-3', 5'-CAA-3', 5'-CGG-3', 5'-ACC-3', 5'-AGA-3', 5'-TTT-3', a motif comprising a sequence selected from the group consisting of: 5'-TCT-3'; and and modifying the oligonucleotide to include a variant of that motif or sequence having at least about 75% sequence identity thereto (U can be T and / or T can be U).

[0100] In one embodiment, the step of modifying the oligonucleotide comprises adding a sequence of nucleotides to the 5' and / or 3' end of the oligonucleotide such that the modified oligonucleotide comprises a motif.

[0101] In certain embodiments, the step of modifying the oligonucleotide comprises modifying the oligonucleotide such that the modified oligonucleotide comprises the motif 5'-GGUAU-3' (SEQ ID NO: 56), 5'-GGUA-3' (SEQ ID NO: 57), 5'-GUAU-3' (SEQ ID NO: 58), 5'-GGU-3' (SEQ ID NO: 59) and 5'-GUA-3' (SEQ ID NO: 60), 5'-GUC-3', 5'-GUG-3', 5'-GUU-3', 5'-GGC-3', 5'-AUC-3', 5'-GAA-3', 5'-GAG-3', 5'-GGA-3', 5'-GAC-3', 5'-GAU- The invention comprises adding a motif selected from 3', 5'-AUG-3', 5'-GCG-3', 5'-UUC-3', 5'-GCC-3', 5'-GGG-3', 5'-AUU-3', 5'-GCA-3', 5'-AGC-3', 5'-AAC-3', 5'-CCA-3', 5'-UGC-3', 5'-CAA-3', 5'-CGG-3', 5'-ACC-3', 5'-AGA-3', 5'-TTT-3', or 5'-TCT-3', or a portion or fragment thereof (where U may be T), to the 5' and / or 3' end of the oligonucleotide. More particularly, the step of modifying the oligonucleotide preferably comprises modifying the oligonucleotide to include the motif 5'-GGUAU-3' (SEQ ID NO: 56), 5'-GGUA-3' (SEQ ID NO: 57), 5'-GUAU-3' (SEQ ID NO: 58), 5'-GGU-3' (SEQ ID NO: 59), 5'-GUA-3' (SEQ ID NO: 60), 5'-GUC-3', 5'-GUG-3', 5'-GUU-3', 5'-GUA-3', 5'-GGC-3', 5'-AUC-3', 5'-GAA-3', 5'-GAG-3', 5'-GGA-3', The method comprises adding a motif selected from 5'-GAC-3', 5'-GAU-3', 5'-AUG-3', 5'-GCG-3', 5'-UUC-3', 5'-GCC-3', 5'-GGG-3', 5'-AUU-3', 5'-GCA-3', 5'-AGC-3', 5'-AAC-3', 5'-CCA-3', 5'-UGC-3', 5'-CAA-3', 5'-CGG-3', 5'-ACC-3', 5'-AGA-3', 5'-TTT-3' or 5'-TCT-3' (U may be T) to the 5' end of the oligonucleotide.

[0102] In another embodiment, the method of the invention further comprises testing the ability of the modified oligonucleotides to inhibit TLR7 activity and selecting those oligonucleotides that inhibit TLR7 activity to a greater extent than the unmodified oligonucleotides.

[0103] Preferably, for the above two aspects, the oligonucleotide does not bind, or is not designed to bind, to transcripts encoding TLR7 or its complement.

[0104] Preferably, for the above two aspects, the oligonucleotide binds or is designed to bind to a target transcript that does not encode TLR7 or its complement.

[0105] In an alternative embodiment of the above aspect, the oligonucleotide binds to or is designed to bind to a target transcript encoding TLR7 or its complement.

[0106] In one embodiment of the above two aspects, the motif is within 11 bases of the 5' and / or 3' end of the oligonucleotide.

[0107] In one embodiment of the above two aspects, the motif is within 8 bases of the 5' and / or 3' end of the oligonucleotide.

[0108] In one embodiment of the above two aspects, the motif is at or towards the 5' and / or 3' end of the oligonucleotide.

[0109] In one embodiment of the above two aspects, the motif is selected from the group consisting of the sequences 5'-[A / G]GU[A / C][U / C]C-3' (SEQ ID NO: 1), 5'-A[G / A][U / G]C[U / C]C-3' (SEQ ID NO: 2), 5'-A[G / A][U / G]C[U / C]C[U / C][C / A]U-3' (SEQ ID NO: 212), 5'-GGUAUA-3' (SEQ ID NO: 4), 5'-UGUUUC-3' (SEQ ID NO: 5), 5'-UGUGUC-3' (SEQ ID NO: 6), 5'-CGUGUC-3' (SEQ ID NO: 8), 5'-GCAGUCTCCATGTCCCAGGC-3' (SEQ ID NO: 30). , 5'-GAUGGTTCCAGTCCCUCUUC-3' (SEQ ID NO: 38), 5'-AGCAGTCTCCATGTCCCAGG-3' (SEQ ID NO: 31), 5'-GGGUCTCCTCCACACCCUUC-3' (SEQ ID NO: 36), 5'-GGUGGCCACAGGCAACGUCA-3' (SEQ ID NO: 28), 5'-GCCGUTTCCATGTCCCAGGC-3' (SEQ ID NO: 46), 5'-GCGGUATCCATGTCCCAGGC-3' (SEQ ID NO: 42), 5'-GCGGUATACAGGTCCCAGGC-3' (SEQ ID NO: 43), 5' -GCUGUTTCCATGTCCCAGGC-3' (SEQ ID NO: 44), 5'-GCUGUGTCCATGTCCCAGGC-3' (SEQ ID NO: 45), 5'-GCCGUGTCCATGTCCCAGGC-3' (SEQ ID NO: 47), 5'-GCGGUAUCCAUGUCCCAGGC-3' (SEQ ID NO: 151), 5'-GGUATCCCCCCCCCCCCCCCC-3' (SEQ ID NO: 54), 5'-GUCCCATCCCTTCTGCUGCC-3' (SEQ ID NO: 145), 5'-UUCUCTCTGGTCCCAUCCCU-3' (SEQ ID NO: 213), 5 '-GUUCAGTCAGATCGCUGGGA-3' (SEQ ID NO: 214), 5'-AUGACATTTCGTGGCUCCUA-3' (SEQ ID NO: 215), 5'-UCUCCATGTCCCAGGCCUCC-3' (SEQ ID NO: 216), 5'-AGUCUCCATGTCCCAGGCCU-3' (SEQ ID NO: 217), 5'-CCAUGTCCCAGGCCTCCAGU-3' (SEQ ID NO: 29), 5'-GCAAGGCAGAGAAACUCCAG-3' (SEQ ID NO: 37), 5'-GGAUUAAAACAGATTAAUAC-3' (SEQ ID NO: 55),5'-AGCCGAACAGAAGGAGCGUC-3' (SEQ ID NO: 40), 5'-UCCGGCCTCGGAAGCUCUCU-3' (SEQ ID NO: 10), 5'-UCCGGCCTCGGAGTCUCCAU-3' (SEQ ID NO: 52), 5'-GCGGUATCCATAGTCUCCAU-3' (SEQ ID NO: 48), 5'-GAUUAAAACAGATTAAUACA-3' (SEQ ID NO: 165), 5'-UGACAAAACAATAATAACAG-3' (SEQ ID NO: 167), or 5'-CCAACACTTCGTGGGGUCCU-3' (SEQ ID NO: 160) (U may be T and / or T may be U).

[0110] In one embodiment of the above two aspects, the motif is selected from the sequences: 5'-GGCATCCACCACGTCGTCCA-3' (SEQ ID NO: 320), 5'-GTCCTTGCACGTGGCTTCGT-3' (SEQ ID NO: 321), 5'-TGTCCTTGCACGTGGCTTCG-3' (SEQ ID NO: 302), 5'-GGAGATTTCAGAGCAGCTTC-3' (SEQ ID NO: 322), 5'-TTCTGCAGCTTCCTTGTCCT-3' (SEQ ID NO: 323), 5' -TGGGCTGGAATCCGAGTTAT-3' (SEQ ID NO: 179), 5'-GTCCACATCCTGTGGCTCGT-3' (SEQ ID NO: 303), 5'-TGTGATGGCCTCCCATCTCC-3' (SEQ ID NO: 304), 5'-TTTGCACACTTCGTACCCAA-3' (SEQ ID NO: 94), or 5'-GTCCAAGATCAGCAGTCTCA (SEQ ID NO: 178) (U can be T and / or T can be U).

[0111] In particular embodiments of the above two aspects, the motif has the sequence: 5'-mGmGmCmAmTCCACCACGTCmGmTmCmCmA-3', 5'-mGmTmCmCmTTGCACGTGGCmTmTmCmGmT-3', 5'-mTmGmTmCmCTTGCAGTGGmCmTmTmCmG-3', 5'-mGmGmAmGmATTTCAGAGCAmGmCmTmTmC-3', 5'-mTmTmCmTmGCAGCTTCCTTmGmTmCmCmT-3', 5'-mTmGmGmGmCTGGAATCCGAmGmTmTmAmT-3', 5'-mGmTmCmCmACATCCTGTGGmCmTmCmGmT-3', 5'-mTmGmTmGmATGGCCTCCCAmTmCmTmCmC-3', 5'-mTmTmTmGmCACACTTCGTAmCmCmCmAmA-3', or 5'-mGmTmCmCmAAGATCAGCAGmTmCmTmCmA-3', (U may be T and / or T may be U, and m is a modified base and / or has a modified backbone). In such examples, m is preferably a 2'-MOE modified base.

[0112] In one embodiment of the above two aspects, the motif is selected from the group consisting of the sequences 5'-GGUAUC-3' (SEQ ID NO: 120), 5'-AGUCUC-3' (SEQ ID NO: 121), 5'-GGUCCC-3' (SEQ ID NO: 122), 5'-GGUCUC-3' (SEQ ID NO: 123), 5'-AAGCUC-3' (SEQ ID NO: 124), 5'-AGUCCC-3' (SEQ ID NO: 125), 5'-GGUAUA-3' (SEQ ID NO: 4), 5'-UGUU UC-3' (SEQ ID NO: 5), 5'-UGUGUC-3' (SEQ ID NO: 6), 5'-CGUUUC-3' (SEQ ID NO: 7), 5'-CGUGUC-3' (SEQ ID NO: 8), 5'-GGUAU-3' (SEQ ID NO: 56), 5'-GGUA-3' (SEQ ID NO: 57), 5'-GUAU-3' (SEQ ID NO: 58), 5'-GGU-3' (SEQ ID NO: 59), or 5'-GUA-3' (SEQ ID NO: 60) (U may be T).

[0113] In one embodiment of the above two aspects, the motif is 5'-GGUAUC-3' (SEQ ID NO: 120), 5'-GGUATC-3' (SEQ ID NO: 119), 5'-AGUCTC-3' (SEQ ID NO: 126), 5'-AGTCTC-3' (SEQ ID NO: 127), 5'-GGUCCC-3' (SEQ ID NO: 122), 5'-GGUCTC-3' (SEQ ID NO: 128), 5'-AAGCUC-3' (SEQ ID NO: 124), 5'-A GTCCC-3' (SEQ ID NO: 129), 5'-GGUATA-3' (SEQ ID NO: 130), 5'-UGUTTC-3' (SEQ ID NO: 131), 5'-UGUGTC-3' (SEQ ID NO: 132), 5'-CGUTTC-3' (SEQ ID NO: 133), 5'-CGUGTC-3' (SEQ ID NO: 134), 5'-GGUAT-3' (SEQ ID NO: 135), 5'-GGUAU-3' (SEQ ID NO: 56), 5'-GGUA-3' (SEQ ID NO: 137), Sequence number 57), 5'-GUAU-3' (SEQ ID NO: 58), 5'-GUAT-3' (SEQ ID NO: 136), 5'-GGU-3' (SEQ ID NO: 59) or 5'-GUA-3' (SEQ ID NO: 60), 5'-GUC-3', 5'-GUG-3', 5'-GUU-3', 5'-GGC-3', 5'-AUC-3', 5'-GAA-3', 5'-GAG-3', 5'-GGA-3', 5'-GAC-3', 5'-GAU- 3', 5'-AUG-3', 5'-GCG-3', 5'-UUC-3', 5'-GCC-3', 5'-GGG-3', 5'-AUU-3', 5'-GCA-3', 5'-AGC-3', 5'-AAC-3', 5'-CCA-3', 5'-UGC-3', 5'-CAA-3', 5'-CGG-3', 5'-ACC-3', 5'-AGA-3', 5'-TTT-3' or 5'-TCT-3'.

[0114] In one embodiment, the motif has the sequence 5'-GGUAU-3' (SEQ ID NO: 56), where U can be T, and the motif is at or towards the 5' end of the oligonucleotide.

[0115] In a further embodiment, the motif has the sequence 5'-GGUA-3' (SEQ ID NO: 57), where U can be T, and the motif is at or towards the 5' end of the oligonucleotide.

[0116] In a related embodiment, the motif has the sequence 5'-GUAU-3' (SEQ ID NO: 58), where U can be T, and the motif is at or towards the 5' end of the oligonucleotide.

[0117] In another embodiment, the motif has the sequence 5'-GGU-3' (SEQ ID NO: 59), where U can be T, and the motif is at or towards the 5' end of the oligonucleotide.

[0118] In one embodiment, the motif has the sequence 5'-GUA-3' (SEQ ID NO: 60), where U can be T, and the motif is at or towards the 5' end of the oligonucleotide.

[0119] In one embodiment of the above two aspects, one or more of the bases of the motif are modified bases and / or have a modified backbone.

[0120] In one embodiment of the above two aspects, the motif is 5'-mGmGmUATC-3', 5'-mAmGmUCTC-3', 5'-mAmGTCTC-3', 5'-mGmGmUmCmCC-3', 5'-mGmGmUmCTC-3', 5'-AAGCmUmC-3', 5'-AGTCCC-3' (SEQ ID NO: 129), 5'-mGmGmUATA-3', 5'-mUmGmUTTC-3', 5'-mUmGmUGTC-3', 5'-mCmGmUTTC- 3', 5'-mCmGmUGTC-3', 5'-mGmGmUAU-3', 5'-mGmGmUAT-3', 5'-mGmGmUmAU-3', 5'-mGmGmUmAT-3', 5'-mGmGmUmAmU-3', 5'-mGmGmUA-3', 5'-mGmGmUmA-3', 5'-mGmUmAmU-3', 5'-mGmGmU-3', or 5'-mGmUmA-3' (where m is a modified base and / or has a modified backbone).

[0121] In embodiments of any of the above aspects, the motif comprises, consists essentially of, or consists of any of the sequences in Table 1, 1A, 2, 2A, 3, 3A, 4, 4A, 5, 5A, or 6, with or without the specific modifications defined, and is shown in the Examples to inhibit the activity of TLR7.

[0122] With respect to the above three aspects, the method may include the further step of testing the ability of one or more candidate or modified oligonucleotides to inhibit TLR3, TLR8, TLR9 and / or cGAS activity, and optionally selecting oligonucleotides that inhibit or do not substantially inhibit TLR3, TLR8, TLR9 and / or cGAS activity.

[0123] In certain embodiments, one or more candidate oligonucleotides or modified oligonucleotides inhibit TLR7 and TLR9 activity. Exemplary motifs for such embodiments include 5'-CCAUGTCCCAGGCCTCCAGU-3' (SEQ ID NO:29), 5'-GCAAGGCAGAGAAACUCCAG-3' (SEQ ID NO:37), 5'-GGAUUAAAACAGATTAAUAC-3' (SEQ ID NO:55), 5'-AGCCGAACAGAAGGAGCGUC-3' (SEQ ID NO:40), 5'-UCCGGCCTCGGAAGCUCUCU-3' (SEQ ID NO:10), 5'-UCCGGCCTCGGAGTCUCCAU-3' (SEQ ID NO:52), 5'-GCGGUATCCATAGTCUCCAU-3' (SEQ ID NO:48), 5'-GAUUAAAACAGATTAAUACA-3' (SEQ ID NO:165), 5'-UGACAAAACAATAATAACAG-3' (SEQ ID NO:167), and 5'-CCAACACTTCGTGGGGUCCU-3' (SEQ ID NO:160).

[0124] In certain embodiments, one or more candidate oligonucleotides or modified oligonucleotides inhibit TLR7 and TLR9 activity but do not substantially inhibit cGAS activity. Exemplary motifs for such embodiments include 5'-GAUUAAAACAGATTAAUACA-3' (SEQ ID NO: 165) and 5'-UGACAAAACAATAATAACAG-3' (SEQ ID NO: 167).

[0125] In certain embodiments, one or more candidate oligonucleotides or modified oligonucleotides inhibit TLR7 and cGAS activity. Exemplary motifs for such embodiments include 5'-GCAGUCTCCATGTCCCAGGC-3' (SEQ ID NO: 30), 5'-GAUGGTTCCAGTCCCUCUUC-3' (SEQ ID NO: 38), 5'-AGCAGTCTCCATGTCCCAGG-3' (SEQ ID NO: 31), 5'-GGGUCTCCTCCACACCCUUC-3' (SEQ ID NO: 36), 5'-GGUGGCCACAGGCAACGUCA-3' (SEQ ID NO: 28), 5'-GCCGUT TCCATGTCCCAGGC-3' (SEQ ID NO: 46), 5'-GCGGUATCCATGTCCCAGGC-3' (SEQ ID NO: 42), 5'-GCGGUATACAGGTCCCAGGC-3' (SEQ ID NO: 43), 5'-GCUGUTTCCATGTCCCAGGC-3' (SEQ ID NO: 44), 5'-GCUGUGTCCATGTCCCAGGC-3' (SEQ ID NO: 45), 5'-GCCGUGTCCATGTCCCAGGC-3' (SEQ ID NO: 47), 5'-GC GGUAUCCAUGUCCCAGGC-3' (SEQ ID NO: 151), 5'-GGUATCCCCCCCCCCCCCCC-3' (SEQ ID NO: 54), 5'-CCAUGTCCCAGGCCTCCAGU-3' (SEQ ID NO: 29), 5'-GCAAGGCAGAGAAACUCCAG-3' (SEQ ID NO: 37), 5'-GGAUUAAAACAGATTAAUAC-3' (SEQ ID NO: 55), 5'-AGCCGAACAGAAGGAGCGUC-3' (SEQ ID NO: 40), Examples include 5'-UCCGGCCTCGGAAGCUCUCU-3' (SEQ ID NO: 10), 5'-UCCGGCCTCGGAGTCUCCAU-3' (SEQ ID NO: 52), 5'-GCGGUATCCATAGTCUCCAU-3' (SEQ ID NO: 48), 5'-GGUAU-3' (SEQ ID NO: 56), 5'-GGUA-3' (SEQ ID NO: 57), 5'-GUAU-3' (SEQ ID NO: 58), 5'-GGU-3' (SEQ ID NO: 59), and 5'-GUA-3' (SEQ ID NO: 60).

[0126] In further embodiments, one or more candidate oligonucleotides or modified oligonucleotides inhibit TLR7 and cGAS activity but do not substantially inhibit TLR9 activity. Exemplary motifs for such embodiments include 5'-GCAGUCTCCATGTCCCAGGC-3' (SEQ ID NO: 30), 5'-GAUGGTTCCAGTCCCUCUUC-3' (SEQ ID NO: 38), 5'-AGCAGTCTCCATGTCCCAGG-3' (SEQ ID NO: 31), 5'-GGGUCTCCTCCACACCCUUC-3' (SEQ ID NO: 36), 5'-GGUGGCCACAGGCAACGUCA-3' (SEQ ID NO: 28), 5'-GCCGUTTCCATGTCCCAGGC-3' (SEQ ID NO: 46), 5'-GCGGUATCCATGTCCCAGGC-3' (SEQ ID NO: 42), 5'-GCGGUATACAGGTCCCAGGC -3' (SEQ ID NO: 43), 5'-GCUGUTTCCATGTCCCAGGC-3' (SEQ ID NO: 44), 5'-GCUGUGTCCATGTCCCAGGC-3' (SEQ ID NO: 45), 5'-GCCGUGTCCATGTCCCAGGC-3' (SEQ ID NO: 47), 5'-GCGGUAUCCAUGUCCCAGGC-3' (SEQ ID NO: 151), and 5'-GGUATCCCCCCCCCCCCCCCC-3' (SEQ ID NO: 54), 5'-GGUAU-3' (SEQ ID NO: 56), 5'-GGUA-3' (SEQ ID NO: 57), 5'-GUAU-3' (SEQ ID NO: 58), 5'-GGU-3' (SEQ ID NO: 59), and 5'-GUA-3' (SEQ ID NO: 60).

[0127] In further embodiments, one or more candidate oligonucleotides or modified oligonucleotides inhibit TLR7, TLR9, and cGAS activity. Exemplary motifs for such embodiments include 5'-CCAUGTCCCAGGCCTCCAGU-3' (SEQ ID NO:29), 5'-GCAAGGCAGAGAAACUCCAG-3' (SEQ ID NO:37), 5'-GGAUUAAAACAGATTAAUAC-3' (SEQ ID NO:55), 5'-AGCCGAACAGAAGGAGCGUC-3' (SEQ ID NO:40), 5'-UCCGGCCTCGGAAGCUCUCU-3' (SEQ ID NO:10), 5'-UCCGGCCTCGGAGTCUCCAU-3' (SEQ ID NO:52), and 5'-GCGGUATCCATAGTCUCCAU-3' (SEQ ID NO:48).

[0128] In alternative embodiments, one or more candidate oligonucleotides or modified oligonucleotides inhibit TLR7 but do not substantially inhibit TLR9 and / or cGAS. Exemplary motifs for such embodiments include 5'-GUCCCATCCCTTCTGCUGCC-3' (SEQ ID NO: 145), 5'-UUCUCTCTGGTCCCAUCCCU-3' (SEQ ID NO: 213), 5'-GUUCAGTCAGATCGCUGGGA-3' (SEQ ID NO: 214), 5'-AUGACATTTCGTGGCUCCUA-3' (SEQ ID NO: 215), 5'-UCUCCATGTCCCAGGCCUCC-3' (SEQ ID NO: 216), and 5'-AGUCUCCATGTCCCAGGCCU-3' (SEQ ID NO: 217).

[0129] With respect to the above two aspects, the method may include the further step of testing the ability of one or more candidate or modified oligonucleotides to enhance TLR8 activity, and optionally selecting oligonucleotides that enhance or do not substantially enhance TLR8 activity. Accordingly, in some embodiments, one or more candidate or modified oligonucleotides inhibit TLR7 activity and enhance TLR8 activity. In other embodiments, one or more candidate or modified oligonucleotides inhibit TLR7 activity and do not substantially enhance TLR8 activity.

[0130] In yet another aspect, the present invention provides a method for selecting or designing an oligonucleotide that increases or enhances TLR8 activity, comprising the steps of: i) 5'-CUUCG-3', 5'-CUUCGTG-3', 5'-CUUCGTGGG-3', 5'-UCG-3', 5'-UCA-3', 5'-CGG-3', 5'-UGG-3', 5'-CGC-3', 5'-AGG-3', 5'-GGA-3', 5'-GGC-3', 5'-AGA-3', 5'-CGA-3', 5'-UAG-3', 5'-UCU-3', 5'-AGC-3', 5'-GGU-3', 5'-UGA-3', 5'-AGU-3', 5'-ACG-3', 5'-CGU-3', 5'-UCC-3', 5'-GCG-3', 5'-GGG-3', 5'-UGU-3', 5'-UCA-3', 5'-CUG-3', 5'-UUG-3', 5'-UUA-3', a motif comprising a sequence selected from the group consisting of: 5'-UGC-3'; and scanning the polynucleotide or its complement for a region having a variant of that motif or sequence having at least about 75% sequence identity thereto (where U can be T and / or T can be U); ii) generating one or more candidate oligonucleotides containing the motif; iii) testing the ability of one or more candidate oligonucleotides to increase or enhance TLR8 activity; iv) selecting an oligonucleotide that increases or enhances TLR8 activity.

[0131] In yet another aspect, the present invention provides a method for increasing or enhancing the TLR8 activity of an oligonucleotide, or for increasing the enhancing activity of an oligonucleotide, by modifying the oligonucleotide, wherein the modified oligonucleotide comprises: 5'-CUUCGTGGGGTCCTTUUCAC-3', 5'-CUUCG-3', 5'-CUUCGTG-3', 5'-CUUCGTGGG-3', 5'-UCG-3', 5'-UCA-3', 5'-CGG-3', 5'-UGG-3', 5'-CGC-3', 5'-AGG-3', 5'-GGA-3', 5'-GGC-3', 5'-AGA-3', 5'-CGA-3', 5'-UAG-3', 5'-UCU-3', 5'-AGC-3', 5'-GGU-3', 5'-UGA-3', 5'-AGU-3', 5'-ACG-3', 5'-CGU-3', 5'-UCC-3', 5'-GCG-3', 5'-GGG-3', 5'-UGU-3', 5'-UCA-3', 5'-CUG-3', 5'-UUG-3', 5'-UUA-3', a motif comprising a sequence selected from the group consisting of: 5'-UGC-3'; and and modifying the oligonucleotide to include a variant of that motif or sequence having at least about 75% sequence identity thereto (U can be T and / or T can be U).

[0132] In one embodiment, the step of modifying the oligonucleotide comprises adding a sequence of nucleotides to the 5' and / or 3' end of the oligonucleotide such that the modified oligonucleotide comprises a motif.

[0133] In certain embodiments, the step of modifying the oligonucleotide comprises modifying the oligonucleotide with 5'-CUUCG-3', 5'-CUUCGTG-3', 5'-CUUCGTGGG-3', 5'-UCG-3', 5'-CGG-3', 5'-UGG-3', 5'-CGC-3', 5'-AGG-3', 5'-GGA-3', 5'-GGC-3', 5'-AGA-3', 5'-CGA-3', 5'-UAG-3', 5'-UCU-3', 5'-AGC-3', 5'-GGU-3', 5'-UGA-3', 5'-CGU-3', 5'-UGG-3', 5'-CGC-3', 5'-AGG-3', 5'-GGA-3', 5'-GGC-3', 5'-AGA-3', 5'-CGA-3', 5'-UAG-3', 5'-UCU-3', 5'-AGC-3', 5'-GGU-3', 5'-UGA-3', 5'-UGG-3', 5'-UGG-3', 5'-UGG-3', 5'-UGG-3', 5'-UGG-3', 5'-UGA ... The method comprises adding a motif selected from 5'-AGU-3', 5'-ACG-3', 5'-CGU-3', 5'-UCC-3', 5'-GCG-3', 5'-GGG-3', 5'-UGU-3', 5'-UCA-3', 5'-CUG-3', 5'-UUG-3', 5'-UUA-3' and 5'-UGC-3', or a portion or fragment thereof (U may be T) to the 5' and / or 3' end of the oligonucleotide, such that the modified oligonucleotide comprises the motif. More particularly, the step of modifying the oligonucleotide preferably comprises modifying the oligonucleotide with 5'-CUUCG-3', 5'-CUUCGTG-3', 5'-CUUCGTGGG-3', 5'-UCG-3', 5'-CGG-3', 5'-UGG-3', 5'-CGC-3', 5'-AGG-3', 5'-GGA-3', 5'-GGC-3', 5'-AGA-3', 5'-CGA-3', 5'-UAG-3', 5'-UCU-3', 5'-AGC-3', 5'-GGU-3', The method comprises adding a motif selected from 5'-UGA-3', 5'-AGU-3', 5'-ACG-3', 5'-CGU-3', 5'-UCC-3', 5'-GCG-3', 5'-GGG-3', 5'-UGU-3', 5'-UCA-3', 5'-CUG-3', 5'-UUG-3', 5'-UUA-3' and 5'-UGC-3' (U may be T) to the 5' end of the oligonucleotide so that the modified oligonucleotide comprises the motif.

[0134] In another embodiment, the method of the invention further comprises testing the ability of modified oligonucleotides to increase or enhance TLR8 activity and selecting oligonucleotides that increase or enhance TLR8 activity to a greater extent than unmodified oligonucleotides.

[0135] Preferably, for the above two aspects, the oligonucleotide does not bind to, or is not designed to bind to, a transcript encoding TLR8 or its complement.

[0136] Preferably, for the above two aspects, the oligonucleotide binds or is designed to bind to a target transcript that does not encode TLR8 or its complement.

[0137] In an alternative embodiment of the above aspect, the oligonucleotide binds to or is designed to bind to a target transcript encoding TLR8 or its complement.

[0138] In one embodiment of the above two aspects, the motif is within 11 bases of the 5' and / or 3' end of the oligonucleotide.

[0139] In one embodiment of the above two aspects, the motif is within 8 bases of the 5' and / or 3' end of the oligonucleotide.

[0140] In one embodiment of the above two aspects, the motif is at or towards the 5' and / or 3' end of the oligonucleotide.

[0141] In one embodiment of the above two aspects, the motif has the sequence 5'-CUUCG-3', 5'-CUUCGTG-3', 5'-CUUCGTGGG-3', 5'-UCG-3, 5'-CGG-3', 5'-UGG-3', 5'-CGC-3', 5'-AGG-3' and 5'-GGA-3' (U can be T and / or T can be U).

[0142] In particular embodiments of the above two aspects, the motif has the sequence: 5'-CUUCG-3', 5'-CUUCGTG-3', 5'-CUUCGTGGG-3', 5'-UCG-3', 5'-CGG-3', 5'-UGG-3', 5'-CGC-3', 5'-AGG-3', or 5'-GGA-3' (one, two or more bases are modified bases and / or one or more internucleotide linkages have a modified backbone, preferably the modified bases are 2'OMe and the modified backbone is phosphorothioate).

[0143] In particular embodiments of the above two aspects, the motif has the sequence: 5'-mCmUmUmCmG-3', 5'-mCmUmUmCmGTG-3', 5'-mCmUmUmCmGTGGG-3', 5'-mUmCmG-3', 5'-mCmGmG-3', 5'-mUmGmG-3', 5'-mCmGmC-3', 5'-mAmGmG-3', or 5'-mGmGmA-3' (m is a modified base and / or has a modified backbone, preferably the modified base is 2'OMe and the modified backbone is phosphorothioate).

[0144] In particular embodiments of the above two aspects, the motif has the sequence: 5'-mC * mU * mU * C * G * T * G * G * G * G * T * C * C * T * T * mU * mU * mC * mA * mC-3', 5'-mC* mU * mU * mC * mG-3′, 5'-mC * mU * mU * mC * mG * T * G-3', 5'-mC * mU * mU * mC * mG * T * G * G * G-3', 5'-mU * mC * mG * -3', 5'-mC * mG * mG * -3', 5'-mU * mG * mG * -3', 5'-mC * mG * mC * -3', 5'-mA * mG * mG * -3', or 5'-mG * mG * mA * -3' (m is a 2'OMe modified base, T / G is a DNA base, * is a phosphorothioate modified backbone).

[0145] In one embodiment of the above two aspects, one or more of the bases of the motif are modified bases and / or have a modified backbone, in one embodiment not all bases are modified, e.g. one or more bases may be unmodified and one or more bases may be modified, e.g. modified with 2'OMe.

[0146] In embodiments of any of the above aspects, the motif comprises, consists essentially of, or consists of any of the sequences in Table 1, 1A, 2, 2A, 3, 3A, 4, 4A, 5, 5A, or 6, with or without the specific modifications defined, and is shown in the Examples to increase or enhance the activity of TLR8.

[0147] With respect to the above three aspects, the method may include the further step of testing the ability of one or more candidate or modified oligonucleotides to inhibit TLR3, TLR7, TLR9 and / or cGAS activity, and optionally selecting oligonucleotides that inhibit or do not substantially inhibit TLR3, TLR7, TLR9 and / or cGAS activity.

[0148] In one embodiment, the candidate oligonucleotide or modified oligonucleotide comprises, consists essentially of, or consists of any of the sequences in Table 1, 1A, 2, 2A, 3, 3A, 4, 4A, 5, 5A, or 6, with or without the specific modifications defined, and is shown in the Examples to increase or enhance the activity of TLR8.

[0149] In one embodiment, the candidate oligonucleotide or modified oligonucleotide comprises, consists essentially of, or consists of any of the sequences in Table 1, 1A, 2, 2A, 3, 3A, 4, 4A, 5, 5A, or 6, and is shown in the Examples to increase or enhance the activity of TLR8, and the oligonucleotide inhibits or does not substantially inhibit TLR3, TLR7, TLR9, and / or cGAS activity.

[0150] In yet another aspect, the present invention provides a method for selecting or designing an oligonucleotide that inhibits TLR8 activity, comprising the steps of: i) 5'-GAG-3', 5'-GAC-3', 5'-GAU-3', 5'-GAA-3', 5'-GUC-3', 5'-GUU-3', 5'-GUA-3', 5'-GUG-3', 5'-AUA-3', 5'-AUG-3', 5'-CUU-3', 5'-AAG-3', 5'-AUC-3', 5'-CCC-3', 5'-GCU-3', 5'-CCU-3', 5'-CUA-3', 5'-CUC-3', a motif comprising a sequence selected from the group consisting of: 5'-AAC-3'; and scanning the polynucleotide or its complement for a region having a variant of that motif or sequence having at least about 75% sequence identity thereto (where U can be T and / or T can be U); ii) generating one or more candidate oligonucleotides containing the motif; iii) testing the ability of one or more candidate oligonucleotides to inhibit TLR8 activity; and iv) selecting an oligonucleotide that inhibits TLR8 activity.

[0151] In yet another aspect, the present invention provides a method for increasing the TLR8 inhibitory activity of an oligonucleotide by modifying the oligonucleotide, wherein the modified oligonucleotide comprises: 5'-GAG-3', 5'-GAC-3', 5'-GAU-3', 5'-GAA-3', 5'-GUC-3', 5'-GUU-3', 5'-GUA-3', 5'-GUG-3', 5'-AUA-3', 5'-AUG-3', 5'-CUU-3', 5'-AAG-3', 5'-AUC-3', 5'-CCC-3', 5'-GCU-3', 5'-CCU-3', 5'-CUA-3', 5'-CUC-3', a motif comprising a sequence selected from the group consisting of: 5'-AAC-3'; and and modifying the oligonucleotide to include a variant of that motif or sequence having at least about 75% sequence identity thereto (U can be T and / or T can be U).

[0152] In one embodiment, the step of modifying the oligonucleotide comprises adding a sequence of nucleotides to the 5' and / or 3' end of the oligonucleotide such that the modified oligonucleotide comprises a motif.

[0153] In certain embodiments, the step of modifying the oligonucleotide comprises adding a motif selected from 5'-GAG-3', 5'-GAC-3', 5'-GAU-3', 5'-GAA-3', 5'-GUC-3', 5'-GUU-3', 5'-GUA-3', 5'-GUG-3', 5'-AUA-3', 5'-AUG-3', 5'-CUU-3', 5'-AAG-3', 5'-AUC-3', 5'-CCC-3', 5'-GCU-3', 5'-CCU-3', 5'-CUA-3' and 5'-CUC-3', 5'-AAC-3', or a portion or fragment thereof (wherein U can be T), to the 5' and / or 3' end of the oligonucleotide such that the modified oligonucleotide comprises the motif. More particularly, the step of modifying the oligonucleotide preferably comprises adding a motif selected from 5'-GAG-3', 5'-GAC-3', 5'-GAU-3', 5'-GAA-3', 5'-GUC-3', 5'-GUU-3', 5'-GUA-3', 5'-GUG-3', 5'-AUA-3', 5'-AUG-3', 5'-CUU-3', 5'-AAG-3', 5'-AUC-3', 5'-CCC-3', 5'-GCU-3', 5'-CCU-3', 5'-CUA-3', 5'-CUC-3' and 5'-AAC-3' (where U can be T) to the 5' end of the oligonucleotide such that the modified oligonucleotide comprises the motif.

[0154] In another embodiment, the method of the invention further comprises testing the ability of the modified oligonucleotides to inhibit TLR8 activity and selecting those oligonucleotides that inhibit TLR8 activity to a greater extent than the unmodified oligonucleotides.

[0155] Preferably, for the above two aspects, the oligonucleotide does not bind to, or is not designed to bind to, a transcript encoding TLR8 or its complement.

[0156] Preferably, for the above two aspects, the oligonucleotide binds or is designed to bind to a target transcript that does not encode TLR8 or its complement.

[0157] In an alternative embodiment of the above aspect, the oligonucleotide binds to or is designed to bind to a target transcript encoding TLR8 or its complement.

[0158] In one embodiment of the above two aspects, the motif is within 11 bases of the 5' and / or 3' end of the oligonucleotide.

[0159] In one embodiment of the above two aspects, the motif is within 8 bases of the 5' and / or 3' end of the oligonucleotide.

[0160] In one embodiment of the above two aspects, the motif is at or towards the 5' and / or 3' end of the oligonucleotide.

[0161] In one embodiment of the above two aspects, the motif has the sequence 5'-GAG-3', 5'-GAC-3', 5'-GAU-3', 5'-GAA-3', 5'-GUC-3', 5'-GUU-3', 5'-GUA-3', 5'-GUG-3', 5'-AUA-3', 5'-AUG-3', 5'-CUU-3', 5'-AAG-3', 5'-AUC-3', 5'-CCC-3', 5'-GCU-3', 5'-CCU-3', 5'-CUA-3', 5'-CUC-3', 5'-AAC-3' (U can be T and / or T can be U).

[0162] In particular embodiments of the above two aspects, the motif has the sequence: 5'-GAX-3' or 5'-GUX-3' (wherein X is any nucleotide), 5'-GAG-3', 5'-GAC-3', 5'-GAU-3', 5'-GAA-3', 5'-GUC-3', 5'-GUU-3', 5'-GUA-3', 5'-GUG-3' (one, two or more bases are modified bases and / or one or more internucleotide linkages have a modified backbone, preferably the modified bases are 2'OMe and the modified backbone is phosphorothioate).

[0163] In particular embodiments of the above two aspects, the motif has the sequence: 5'-mGmAmX-3' or 5'-mGmUmX-3' (X is any nucleotide), 5'-mGmAmG-3', 5'-mGmAmC-3', 5'-mGmAmU-3', 5'-mGmAmA-3', 5'-mGmUmC-3', 5'-mGmUmU-3', 5'-mGmUmA-3', 5'-mGmUmG-3' (m is a modified base and / or has a modified backbone, preferably the modified base is 2'OMe and the modified backbone is phosphorothioate).

[0164] In one embodiment of the above two aspects, one or more of the bases of the motif are modified bases and / or have a modified backbone, in one embodiment not all bases are modified, e.g. one or more bases may be unmodified and one or more bases may be modified, e.g. modified with 2'OMe.

[0165] In embodiments of any of the above aspects, the motif comprises, consists essentially of, or consists of any of the sequences in Table 1, 1A, 2, 2A, 3, 3A, 4, 4A, 5, 5A, or 6, with or without the specific modifications defined, and is shown in the Examples to inhibit the activity of TLR8.

[0166] With respect to the above three aspects, the method may include the further step of testing the ability of one or more candidate or modified oligonucleotides to inhibit TLR3, TLR7, TLR9 and / or cGAS activity, and optionally selecting oligonucleotides that inhibit or do not substantially inhibit TLR3, TLR7, TLR9 and / or cGAS activity.

[0167] In one embodiment, the candidate oligonucleotide or modified oligonucleotide comprises, consists essentially of, or consists of any of the sequences in Table 1, 1A, 2, 2A, 3, 3A, 4, 4A, 5, 5A, or 6, and is shown in the Examples to inhibit the activity of TLR8.

[0168] In yet another aspect, the present invention provides a method for selecting or designing an oligonucleotide that inhibits TLR3 activity, comprising the steps of: i) 5'-TAC-3', 5'-CGC-3', 5'-GCA-3', 5'-UGA-3', 5'-CAG-3', 5'-UGG-3', 5'-UCA-3', 5'-TGA-3', 5'-CGT-3', 5'-GAC-3', 5'-CCA-3', 5'-TAG-3', 5'-TGG-3', 5'-TCA-3', 5'-TGC-3', 5'-CAC-3', 5'-CGG-3', 5'-CCC-3', 5'-ACT-3', 5'-GTA-3', 5'-GGA-3', 5'-AAG-3', 5'-ATA-3', 5'-GUC-3', 5'-UCC-3', 5'-AUC-3', 5'-CCG-3', 5'-CAA-3', 5'-GAU-3', a motif comprising a sequence selected from the group consisting of: 5'-CGA-3'; and scanning the polynucleotide or its complement for a region having a variant of that motif or sequence having at least about 75% sequence identity thereto (where U can be T and / or T can be U); ii) generating one or more candidate oligonucleotides containing the motif; iii) testing the ability of one or more candidate oligonucleotides to inhibit TLR3 activity; and iv) selecting an oligonucleotide that inhibits TLR3 activity.

[0169] In yet another aspect, the present invention provides a method for increasing the TLR3 inhibitory activity of an oligonucleotide by modifying the oligonucleotide, wherein the modified oligonucleotide comprises: 5'-TAC-3', 5'-CGC-3', 5'-GCA-3', 5'-UGA-3', 5'-CAG-3', 5'-UGG-3', 5'-UCA-3', 5'-TGA-3', 5'-CGT-3', 5'-GAC-3', 5'-CCA-3', 5'-TAG-3', 5'-TGG-3', 5'-TCA-3', 5'-TGC-3', 5'-CAC-3', 5'-CGG-3', 5'-CCC-3', 5'-ACT-3', 5'-GTA-3', 5'-GGA-3', 5'-AAG-3', 5'-ATA-3', 5'-GUC-3', 5'-UCC-3', 5'-AUC-3', 5'-CCG-3', 5'-CAA-3', 5'-GAU-3', a motif comprising a sequence selected from the group consisting of: 5'-CGA-3'; and and modifying the oligonucleotide to include a variant of that motif or sequence having at least about 75% sequence identity thereto (U can be T and / or T can be U).

[0170] In one embodiment, the step of modifying the oligonucleotide comprises adding a sequence of nucleotides to the 5' and / or 3' end of the oligonucleotide such that the modified oligonucleotide comprises a motif.

[0171] In certain embodiments, the step of modifying the oligonucleotide comprises adding a motif selected from 5'-TAC-3', 5'-CGC-3', 5'-GCA-3', 5'-UGA-3', 5'-CAG-3', 5'-UGG-3', 5'-UCA-3', or a portion or fragment thereof (where U can be T) to the 5' and / or 3' end of the oligonucleotide such that the modified oligonucleotide comprises the motif. More particularly, the step of modifying the oligonucleotide preferably comprises adding a motif selected from 5'-TAC-3', 5'-CGC-3', 5'-GCA-3', 5'-UGA-3', 5'-CAG-3', 5'-UGG-3', 5'-UCA-3' (where U can be T) to the 5' end of the oligonucleotide such that the modified oligonucleotide comprises the motif.

[0172] In another embodiment, the method of the invention further comprises testing the ability of the modified oligonucleotides to inhibit TLR3 activity and selecting those oligonucleotides that inhibit TLR3 activity to a greater extent than the unmodified oligonucleotides.

[0173] Preferably, for the above two aspects, the oligonucleotide does not bind or is not designed to bind to transcripts encoding TLR3 or its complement.

[0174] Preferably, for the above two aspects, the oligonucleotide binds or is designed to bind to a target transcript that does not encode TLR3 or its complement.

[0175] In an alternative embodiment of the above aspect, the oligonucleotide binds to or is designed to bind to a target transcript encoding TLR3 or its complement.

[0176] In one embodiment of the above two aspects, the motif is within 11 bases of the 5' and / or 3' end of the oligonucleotide.

[0177] In one embodiment of the above two aspects, the motif is within 8 bases of the 5' and / or 3' end of the oligonucleotide.

[0178] In one embodiment of the above two aspects, the motif is at or towards the 5' and / or 3' end of the oligonucleotide.

[0179] In one embodiment of the above two aspects, the motif has 5'-TAC-3', 5'-CGC-3', 5'-GCA-3', 5'-UGA-3', 5'-CAG-3', 5'-UGG-3', 5'-UCA-3' (U can be T and / or T can be U).

[0180] In particular embodiments of the above two aspects, the motif has the sequence: 5'-TAC-3', 5'-CGC-3', 5'-GCA-3', 5'-UGA-3', 5'-CAG-3', 5'-UGG-3', 5'-UCA-3' (one, two or more bases are modified bases and / or one or more internucleotide linkages have a modified backbone, preferably the modified bases are 2'OMe and the modified backbone is phosphorothioate).

[0181] In particular embodiments of the above two aspects, the motif has the sequence: 5'-mUmAmC-3', 5'-mCmGmC-3', 5'-mGmCmA-3', 5'-mUmGmA-3', 5'-mCmAmG-3', 5'-mUmGmG-3', 5'-mUmCmA-3' (m is a modified base and / or has a modified backbone, preferably the modified base is 2'OMe and the modified backbone is phosphorothioate).

[0182] In one embodiment of the above two aspects, one or more of the bases of the motif are modified bases and / or have a modified backbone.

[0183] In embodiments of any of the above aspects, the motif comprises, consists essentially of, or consists of any of the sequences in Table 1, 1A, 2, 2A, 3, 3A, 4, 4A, 5, 5A, or 6, with or without the specific modifications defined, and is shown in the Examples to inhibit the activity of TLR3.

[0184] With respect to the above three aspects, the method may include the further step of testing the ability of one or more candidate or modified oligonucleotides to inhibit TLR8, TLR7, TLR9 and / or cGAS activity, and optionally selecting oligonucleotides that inhibit or do not substantially inhibit TLR8, TLR7, TLR9 and / or cGAS activity.

[0185] In one embodiment, the candidate oligonucleotide or modified oligonucleotide comprises, consists essentially of, or consists of any of the sequences in Table 1, 1A, 2, 2A, 3, 3A, 4, 4A, 5, 5A, 6, or 7, with or without the specific modifications defined, and is shown in the Examples to inhibit the activity of TLR3.

[0186] In a further aspect, the invention pertains to oligonucleotides selected, designed or modified using the methods of the previous aspects.

[0187] In a still further aspect, the present invention provides a method for producing a pharmaceutical composition comprising: 5'-[A / G]GU[A / C][U / C]C-3' (SEQ ID NO: 1), 5'-A[G / A][U / G]C[U / C]C-3' (SEQ ID NO: 2), 5'-A[G / A][U / G]C[U / C]C[U / C][C / A]U-3' (SEQ ID NO: 3), 5'-GGUAUA-3' (SEQ ID NO: 4), 5'-UGUUUC-3' (SEQ ID NO: 5), 5'-UGUGUC-3' (SEQ ID NO: 6), 5'-CGUUUC-3' (SEQ ID NO: 7), 5'-CGUGUC-3' (SEQ ID NO: 8), 5'-AUGGCCTTTCCGTGCCAAGG-3' (SEQ ID NO: 9), 5'-UCCGGCCTCGGAAGCUCUCU-3' (SEQ ID NO: 10), 5'-GCAUUCCGTGCGGAAGCCUU-3' (SEQ ID NO: 11), 5'-GGCCGAACTTTCCCGCCUUA-3' (SEQ ID NO: 12), 5'-GGUCUTGGCTTCGTGGAGCA-3' (SEQ ID NO: 13), 5'-GGAGCTTCGAGGCCCCAGGC-3' (SEQ ID NO: 14), 5'-GGUGGTCCACAACCCCUUUC-3' (SEQ ID NO: 15), 5'-CAUUAGGTGCAGAAAUCUUC-3' (SEQ ID NO: 16), 5'-UUCUGGGGACTTCCAGUUUA-3' (SEQ ID NO: 17), 5'-UGAUUCCAAAGCCAGGGUUA-3' (SEQ ID NO: 18), 5'-CUUUAGTCGTAGTTGCUUCC-3' (SEQ ID NO: 19), 5'-UUAAATAATCTAGTTUGAAG-3' (SEQ ID NO: 20), 5'-GUGUCCTTCATGCTTUGGAU-3' (SEQ ID NO: 21), 5'-AGAAAGAAGCAAAGAUUCAA-3' (SEQ ID NO: 22), 5'-AGAUUATCTTCTTTTAAUUU-3' (SEQ ID NO: 23), 5'-AAAAGATTATCTTCTUUUAA-3' (SEQ ID NO: 24), 5'-GAAAAGATTATCTTCUUUUA-3' (SEQ ID NO: 25), 5'-UGUGAAAAGATTATCUUCUU-3' (SEQ ID NO: 26), 5'-CUUGUGAAAAGATTAUCUUC-3' (SEQ ID NO: 27), 5'-GGUGGCCACAGGCAACGUCA-3' (SEQ ID NO: 28), 5'-CCAUGTCCCAGGCCTCCAGU-3' (SEQ ID NO: 29), 5'-GCAGUCTCCATGTCCCAGGC-3' (SEQ ID NO: 30), 5'-AGCAGTCTCCATGTCCCAGG-3' (SEQ ID NO: 31), 5'-AGUGGCACATACCACACCCU-3' (SEQ ID NO: 32), 5'-AUUUCCACATGCCCAGUGUU-3' (SEQ ID NO: 33), 5'-GGUCCCATCCCTTCTGCUGC-3' (SEQ ID NO: 34), 5'-UCUGGTCCCATCCCTUCUGC-3' (SEQ ID NO: 35), 5'-GGGUTCCTCCACACCCUUC-3' (SEQ ID NO: 36), 5'-GCAAGGCAGAGAAACUCCAG-3' (SEQ ID NO: 37), 5'-GAUGGTTCCAGTCCCUCUUC-3' (SEQ ID NO: 38), 5'-UGUUUCCCCGGAGAGCAAUG-3' (SEQ ID NO: 39), 5'-AGCCGAACAGAAGGAGCGUC-3' (SEQ ID NO: 40), 5'-GCGUAGTTTCTCTTCCUCCC-3' (SEQ ID NO: 41), 5'-GCGGUATCCATGTCCCAGGC-3' (SEQ ID NO: 42), 5'-GCGGUATACAGGTCCCAGGC-3' (SEQ ID NO: 43), 5'-GCUGUTTCCATGTCCCAGGC-3' (SEQ ID NO: 44), 5'-GCUGUGTCCATGTCCCAGGC-3' (SEQ ID NO: 45), 5'-GCCGUTTCCATGTCCCAGGC-3' (SEQ ID NO: 46), 5'-GCCGUGTCCATGTCCCAGGC-3' (SEQ ID NO: 47), 5'-GCGGUATCCATAGTCUCCAU-3' (SEQ ID NO: 48), 5'-GCGGUATCCATCAGAUAUCG-3' (SEQ ID NO: 49), 5'-CUUUAGTCGTAGTTGUCUCU-3' (SEQ ID NO: 50), 5'-UCCGGGTCGTAGTTGCUUCC-3' (SEQ ID NO: 51), 5'-UCCGGCCTCGGAGTCUCCAU-3' (SEQ ID NO: 52), 5'-UCCGGCCTCGGGAGAUCUCU-3' (SEQ ID NO: 53), 5'-GGUATCCCCCCCCCCCCCCC-3' (SEQ ID NO: 54), 5'-GGAUUAAAACAGATTAAUAC-3' (SEQ ID NO: 55), 5'-GGUAU-3' (SEQ ID NO: 56), 5'-GGUA-3' (SEQ ID NO: 57), 5'-GUAU-3' (SEQ ID NO: 58), 5'-GGU-3' (SEQ ID NO: 59), 5'-GUA-3' (SEQ ID NO: 60), 5'-TGTCTG-3' (SEQ ID NO: 61), 5'-GTCT-3' (SEQ ID NO: 62), 5'-TCTCCG-3' (SEQ ID NO: 63), 5'-CTCC-3' (SEQ ID NO: 64), 5'-[G / A][A / C]AG[G / C][T / C]T[C / A] (SEQ ID NO: 65), 5'-AAAGGTTA-3' (SEQ ID NO: 66), 5'-GAAGCTTC-3' (SEQ ID NO: 67), 5'-GCAGGCTC-3' (SEQ ID NO: 68), 5'-A[G / A]GGTT-3' (SEQ ID NO: 69), 5'-AGGGTT-3' (SEQ ID NO: 70), 5'-AAGGTT-3' (SEQ ID NO: 71), 5'-GGTT-3' (SEQ ID NO: 72), 5'-[A / G]GCT[T / C][T / C][G / C][T / A]-3' (SEQ ID NO: 73), 5'-AGCTTCCT-3' (SEQ ID NO: 74), 5'-AGCTTCGA-3' (SEQ ID NO: 75), 5'-GGCTTCGT-3' (SEQ ID NO: 76), 5'-TGCTTCCT-3' (SEQ ID NO: 77), 5'-AGCTCTCT-3' (SEQ ID NO: 78), 5'-G[G / C]TT-3' (SEQ ID NO: 79), 5'-GCTT-3' (SEQ ID NO: 80), 5'-CGGAGGTCTTGGCTTCGTGG-3' (SEQ ID NO: 81), 5'-AGGTCTTGGCTTCGTGGAGC-3' (SEQ ID NO: 82), 5'-GGGAAAAGGTTATGCAAGGTC-3' (SEQ ID NO: 83), 5'-CTGTGATCTTGACATGCTGC-3' (SEQ ID NO: 84), 5'-ACTGACTGTCTTGAGGGTTC-3' (SEQ ID NO: 85), 5'-GCGTGTCTGGAAGCTTCCTT-3' (SEQ ID NO: 86), 5'-GAGTCTCTGGAGCTTCCTCT-3' (SEQ ID NO: 87), 5'-AGTCGTAGTTGCTTCCTAAC-3' (SEQ ID NO: 88), 5'-GTCTTGGCTTCGTGGAGCAG-3' (SEQ ID NO: 89), 5'-TTGGCTCGGCTTGCCTACTT-3' (SEQ ID NO: 90), 5'-ACAGTGTTGAGATACTCGGG-3' (SEQ ID NO: 91), 5'-TCGCACTTCAGTCTGAGCAG-3' (SEQ ID NO: 92), 5'-GGTGTCCTTGCACGTGGCTT-3' (SEQ ID NO: 93), 5'-TTTGCACACTTCGTACCCAA-3' (SEQ ID NO: 94), 5'-GCTGACAAAGATTCACTGGT-3' (SEQ ID NO: 95), 5'-GCGGAGGTCTTGGCTTCGTG-3' (SEQ ID NO: 96), 5'-CCAAGATCAGCAGTCT-3' (SEQ ID NO: 97), 5'-CTTGAAGCATCGTATC-3' (SEQ ID NO: 98), 5'-GCACACTTCGTACCCA-3' (SEQ ID NO: 99), 5'-GATAGCACCTTCAGCA-3' (SEQ ID NO: 100), 5'-CGTATTATAGCCGATT-3' (SEQ ID NO: 101), 5'-GCAGGCTCAGTGATGT-3' (SEQ ID NO: 102), 5'-GAAAGGTTATGCAAGG-3' (SEQ ID NO: 103), 5'-ATGGCCTCCCATCTCC-3' (SEQ ID NO: 104), 5'-CGCTTTTCTGTCTGGT-3' (SEQ ID NO: 105), 5'-GTGTCTGGAAGCTTCC-3' (SEQ ID NO: 106), 5'-TGGCCTCCCATCTCCT-3' (SEQ ID NO: 107), 5'-ATCTGGCAGCCCATCA-3' (SEQ ID NO: 108), 5'-GAGGTCTTGGCTTCGT-3' (SEQ ID NO: 109), 5'-ACACTTCGTGGGGTCC-3' (SEQ ID NO: 110), 5'-TTCGTGGGGTCCTTTT-3' (SEQ ID NO: 111), 5'-CCACTTGGCAGACCAT-3' (SEQ ID NO: 112), 5'-CCATCCATGAGGTCCT-3' (SEQ ID NO: 113), 5'-TCCAACACTTCGTGGG-3' (SEQ ID NO: 114), 5'-TCTTCATCGGCCCTGC-3' (SEQ ID NO: 115), 5'-CCAGCAGGTCAGCAAA-3' (SEQ ID NO: 116), 5'-CGCTTTTCTCTCCGGT-3' (SEQ ID NO: 117), 5'-GGUAUAGGACTCCAGATGUUUCC-3' (SEQ ID NO: 118), and and oligonucleotides comprising, consisting essentially of, or consisting of variants of that motif or sequence having at least about 75% sequence identity thereto (wherein U can be T and / or T can be U, and wherein the oligonucleotide inhibits cGAS activity when administered to a subject or in any assay described herein).

[0188] In one embodiment of the above aspect, the oligonucleotide comprises or consists of any of the sequences in Table 1, 1A, 2, 2A, 3, 3A, 4, 4A, 5, 5A or 6, with or without the specific modifications defined, and is shown in the Examples to inhibit the activity of cGAS.

[0189] In embodiments where the motif is 5'-GGUAUA-3' (SEQ ID NO:4), 5'-GGUAU-3' (SEQ ID NO:56), 5'-GGUA-3' (SEQ ID NO:57), 5'-GUAU-3' (SEQ ID NO:58), 5'-GGU-3' (SEQ ID NO:59), or 5'-GUA-3' (SEQ ID NO:60) (U can be T), the motif is suitably at or towards the 5' and / or 3' end of the oligonucleotide. In particular embodiments, the 5'-GGUAUA-3' (SEQ ID NO:4), 5'-GGUAU-3' (SEQ ID NO:56), 5'-GGUA-3' (SEQ ID NO:57), 5'-GUAU-3' (SEQ ID NO:58), 5'-GGU-3' (SEQ ID NO:59), or 5'-GUA-3' (SEQ ID NO:60) (U can be T) motif is at or towards the 5' end of the oligonucleotide.

[0190] In one embodiment, the oligonucleotides are 5'-GCGGUATCCATGTCCCAGGC-3' (SEQ ID NO: 42), 5'-GCGGUATACAGGTCCCAGGC-3' (SEQ ID NO: 43), 5'-GCUGUTTCCATGTCCCAGGC-3' (SEQ ID NO: 44), 5'-GCUGUGTCCATGTCCCAGGC-3' (SEQ ID NO: 45), 5'-GCCGUTTCCATGTCCCAGGC-3' (SEQ ID NO: 46), 5'-GCCGUGTCCATGTCCCAGGC-3' (SEQ ID NO: 47), 5'-GCGGUATCCATAGTCUCCAU-3' (SEQ ID NO: 48), 5'-GCGGUATCCATCAG AUAUCG-3' (SEQ ID NO: 49), 5'-CUUUAGTCGTAGTTGUCUCU-3' (SEQ ID NO: 50), 5'-UCCGGGTCGTAGTTGCUUCC-3' (SEQ ID NO: 51), 5'-UCCGGCCTCGGAGTCUCCAU-3' (SEQ ID NO: 52), 5'-UCCGGCCTCGGGAGAUCUCU-3' (SEQ ID NO: 53), 5'-GGUATCCCCCCCCCCCCCCCC-3' (SEQ ID NO: 54), or a variant thereof having at least about 75% sequence identity thereto (U may be T and / or T may be U).

[0191] In one embodiment, the oligonucleotide comprises, consists essentially of, or consists of any of the sequences in Table 1, 1A, 2, 2A, 3, 3A, 4, 4A, 5, 5A, or 6, with or without specific modifications as defined, and is shown in the Examples to inhibit the activity of cGAS.

[0192] In one embodiment, the oligonucleotide comprises, consists essentially of, or consists of any of the sequences in Table 1, 1A, 2, 2A, 3, 3A, 4, 4A, 5, 5A, or 6, with or without the specific modifications defined, and is shown in the Examples to inhibit the activity of cGAS, wherein the oligonucleotide inhibits or does not substantially inhibit TLR3, TLR7, and / or TLR9 activity.

[0193] In other embodiments, the oligonucleotide is 5'-mGmCmGmGmUATCCATGTCCmCmAmGmGmC-3', 5'-mGmCmGmGmUmAmUmCmCmAmUmGmUmCmCmCmAmAmGmGmC-3', 5'-mGmCmGmGmUATACAGGTCCmCmAmGmGmC-3', 5'-mGmCmUmGmUTTCCATGTCCmCmAmGmGmC-3', 5'-mGmCmUmGmUGTCCATGTCCmCmAmGmGmC-3', 5'-mGmCmCmGmUTTCCATGTCCmCmAmGmGmC-3', 5'-mGmCmCmGmUGTCCATGTCCmCmAmGmGmC-3', 5'-mGmCmGmGmUATCCATAGTCmUmCmCmAmU-3', 5'-mGmCmGmGmUATCCATCAGAmUmAmUmCmG-3', 5'-mCmUmUmUmAGTCGTAGTTGmUmCmUmCmU-3', 5'-mUmCmCmGmGGTCGTAGTTGmCmUmUmCmC-3', 5'-mUmCmCmGmGCCTCGGAGTCmUmCmCmAmU-3', 5'-mUmCmCmGmGCCTCGGGAGAmUmCmUmCmU-3', 5'-mGmGmUATCCCCCCCCCCCCCCC-3', 5'-mGmGmUmAmU-3', 5'-mGmGmUmA-3', 5'-mGmUmAmU-3', 5'-mGmGmU-3', The sequence 5'-mGmUmA-3', or a variant thereof having at least about 75% sequence identity thereto (wherein U can be T and / or T can be U, and m is a modified base and / or has a modified backbone).

[0194] In another embodiment, the oligonucleotide comprises, consists essentially of, or consists of the sequence 5'-GCGGUATCCATGTCCCAGGC-3' (SEQ ID NO: 42), or a variant thereof having at least about 75% sequence identity thereto (U can be T and / or T can be U).

[0195] In some embodiments, the oligonucleotide comprises, consists essentially of, or consists of the sequence 5'-GGUAU-3' (SEQ ID NO: 56) or a variant thereof having at least about 75% sequence identity thereto (wherein U can be T).

[0196] In other embodiments, the oligonucleotide comprises, consists essentially of, or consists of the sequence 5'-GGUA-3' (SEQ ID NO: 57) or a variant thereof having at least about 75% sequence identity thereto (wherein U can be T).

[0197] In some embodiments, the oligonucleotide comprises, consists essentially of, or consists of the sequence 5'-GUAU-3' (SEQ ID NO: 58) or a variant thereof having at least about 75% sequence identity thereto (wherein U can be T).

[0198] In certain embodiments, the oligonucleotide comprises, consists essentially of, or consists of the sequence 5'-GGU-3' (SEQ ID NO: 59) or a variant thereof having at least about 75% sequence identity thereto (wherein U can be T).

[0199] In some embodiments, the oligonucleotide comprises, consists essentially of, or consists of the sequence 5'-GUA-3' (SEQ ID NO: 60) or a variant thereof having at least about 75% sequence identity thereto (wherein U can be T).

[0200] In certain embodiments, the oligonucleotide comprises, consists essentially of, or consists of the sequence 5'-mGmCmGmGmUATCCATGTCCmCmAmGmGmC-3', or a variant thereof having at least about 75% sequence identity thereto (U can be T and / or T can be U, m is a modified base, and / or has a modified backbone).

[0201] In certain embodiments, the oligonucleotide comprises, consists essentially of, or consists of the sequence 5'-mGmCmGmGmUmAmUmCmCmAmUmGmUmCmCmCmAmGmGmC-3', or a variant thereof having at least about 75% sequence identity thereto (U can be T and / or T can be U, and m is a modified base and / or has a modified backbone).

[0202] In any embodiment of this aspect, the oligonucleotide that inhibits cGAS activity is at least 15, 16, 17, 18, 19, or 20 nucleotides in length.

[0203] In another aspect, the present invention provides a method for producing a pharmaceutical composition comprising: 5'[C / U]CUUCU-3' (SEQ ID NO: 140), 5'-CACCCTTCTCTCTGGUCCCA-3' (SEQ ID NO: 141), 5'-CCUUCTCTCTGGTCCCAUCC-3' (SEQ ID NO: 142), 5'-UCUCUGGTCCCATCCCUUCU-3' (SEQ ID NO: 143), 5'-AUAUCTGCTGCCCACCUUCU-3' (SEQ ID NO: 144), 5'-GUCCCATCCCTTCTGCUGCC-3' (SEQ ID NO: 145), 5'-GUCUCCTCCACACCCUUCUC-3' (SEQ ID NO: 146), 5'-CAGGCCTCCAGTGTCUUCUC-3' (SEQ ID NO: 147), 5'-UCCCAACTCTTCTAACUCGU-3' (SEQ ID NO: 148), and Oligonucleotides are provided that contain variants of that motif or sequence having at least about 75% sequence identity thereto (wherein U can be T and / or T can be U, and wherein the oligonucleotide does not inhibit or exhibits reduced inhibition of cyclic GMP-AMP synthase (cGAS) activity when administered to a subject or in any assay described herein).

[0204] In yet another aspect, the present invention provides an oligonucleotide comprising: 5'-G[G / C]CCT[C / G]-3' (SEQ ID NO: 152), 5'-CUU-3' (SEQ ID NO: 153), wherein the motif is within 10 bases of the 5' and / or 3' end of the oligonucleotide; 5'-CUUGUGAAAAGATTAUCUUC-3' (SEQ ID NO: 27), 5'-CUUCUCTCTGGTCCCAUCCC-3' (SEQ ID NO: 154), 5'-CCUUCTCTCTGGTCCCAUCC-3' (SEQ ID NO: 142), 5'-CCCUUCTCTCTGGTCCCAUC-3' (SEQ ID NO: 155), 5'-ACCCUTCTCTCTGGTCCCAU-3' (SEQ ID NO: 156), 5'-CUUCCACAATCAAGACAUUC-3' (SEQ ID NO: 157), 5'-CUUCGTGGGGTCCTTUUCAC-3' (SEQ ID NO: 158), 5'-CACUUCGTGGGGTCCUUUUC-3' (SEQ ID NO: 159), 5'-CCAACACTTCGTGGGGUCCU-3' (SEQ ID NO: 160), 5'-UCCGGCCTCGGAAGCUCUCU-3' (SEQ ID NO: 10), 5'-CCAUGTCCCAGGCCTCCAGU-3' (SEQ ID NO: 29), 5'-UUGGCCTGTGGATGCUUUGU-3' (SEQ ID NO: 161), 5'-AAAUGTCCTGGCCCTCACUG-3' (SEQ ID NO: 162), 5'-UCCGGCCTCGGAGTCUCCAU-3' (SEQ ID NO: 52), 5'-UCCGGCCTCGGCAGAUAUCG-3' (SEQ ID NO: 163), 5'-GGCCGAACTTTCCCGCCUUA-3' (SEQ ID NO: 12), 5'-GGUCUTGGCTTCGTGGAGCA-3' (SEQ ID NO: 13), 5'-GGAGCTTCGAGGCCCCAGGC-3' (SEQ ID NO: 14), 5'-GGUGGTCCACAACCCCUUUC-3' (SEQ ID NO: 15), 5'-CAUUAGGTGCAGAAAUCUUC-3' (SEQ ID NO: 16), 5'-UUCUGGGGACTTCCAGUUUA-3' (SEQ ID NO: 17), 5'-UGAUUCCAAAGCCAGGGUUA-3' (SEQ ID NO: 18), 5'-UCCGGGTCGTAGTTGCUUCC-3' (SEQ ID NO: 51), 5'-CCUAGAAAGAAGCAAAGAUU-3' (SEQ ID NO: 164), 5'-GAUUAAAACAGATTAAUACA-3' (SEQ ID NO: 165), 5'-GGAUUAAAACAGATTAAUAC-3' (SEQ ID NO: 55), 5'-AAUUUAAAGCATGAAUAUUA-3' (SEQ ID NO: 166), 5'-GCGUAGTTTCTCTTCCUCCC-3' (SEQ ID NO: 41), 5'-UGACAAAACAATAATAACAG-3' (SEQ ID NO: 167), 5'-ACA-3' (SEQ ID NO: 168), wherein the motif is at or towards the 5' end of the oligonucleotide; 5'-CAC-3' (SEQ ID NO: 169), wherein the motif is at or towards the 5' end of the oligonucleotide; 5'-ACACTTCGTGGGGTCCTTTT-3' (SEQ ID NO: 170), 5'-GCGTGTCTGGAAGCTTCCTT-3' (SEQ ID NO: 86), 5'-TCAAAGGACTGAGGAAAGGG-3' (SEQ ID NO: 171), 5'-ATCCAACACTTCGTGGGGTC-3' (SEQ ID NO: 172), 5'-GCCCATCCATGAGGTCCTGG-3' (SEQ ID NO: 173), 5'-GGGTATCGAAAGAGTCTGGA-3' (SEQ ID NO: 174), 5'-GGTTTTGGCTGGGATCAAGT-3' (SEQ ID NO: 175), 5'-GCGACTATACGCGCAATATG-3' (SEQ ID NO: 176), 5'-ACTGACTGTCTTGAGGGTTC-3' (SEQ ID NO: 85), 5'-AACACTTCGTGGGGTCCTTT-3' (SEQ ID NO: 177), 5'-GTCCAAGATCAGCAGTCTCA-3' (SEQ ID NO: 178), 5'-ACAGTGTTGAGATACTCGGG-3' (SEQ ID NO: 91), 5'-TGGGCTGGAATCCGAGTTAT-3' (SEQ ID NO: 179), 5'-CGGCATCCACCACGTCGTCC-3' (SEQ ID NO: 180), 5'-GCGTATTATAGCCGATTAAC-3' (SEQ ID NO: 181), 5'-GGAGGTCTTGGCTTCGTGGA-3' (SEQ ID NO: 182), 5'-TGGGTTACGGCTCAGTATGG-3' (SEQ ID NO: 183), 5'-CCGCCATGTTTCTTCTTGGA-3' (SEQ ID NO: 184), 5'-AGCTTCGAGGCCCCAG-3' (SEQ ID NO: 185), 5'-GCCATGTTTCTTCTTG-3' (SEQ ID NO: 186), 5'-CACTTCGTGGGGTCCT-3' (SEQ ID NO: 187), 5'-CGGCCTCGGAAGCTCT-3' (SEQ ID NO: 188), 5'-ACACTTCGTGGGGTCC-3' (SEQ ID NO: 110), 5'-TGCACACTTCGTACCC-3' (SEQ ID NO: 189), 5'-CCACATCCTGTGGCTC-3' (SEQ ID NO: 190), 5'-CTGCAGCTTCCTTGTC-3' (SEQ ID NO: 191), 5'-ACTTCGTGGGGTCCTT-3' (SEQ ID NO: 192), 5'-CCCACTTGGCAGACCA-3' (SEQ ID NO: 193), 5'-GTCCCCTGTTGACTGG-3' (SEQ ID NO: 194), 5'-ACGTTCAGTCCTGTCC-3' (SEQ ID NO: 195), 5'-GGTCATTACAATAGCT-3' (SEQ ID NO: 196), 5'-TCGTGGGGTCCTTTTC-3' (SEQ ID NO: 197), 5'-GTGTCTGGAAGCTTCC-3' (SEQ ID NO: 106), 5'-ATGGCCTCCCATCTCC-3' (SEQ ID NO: 104), 5'-TGCTCCTCGGTCTCCC-3' (SEQ ID NO: 198), 5'-GCATCCACCACGTCGT-3' (SEQ ID NO: 199), 5'-CTTCGTGGGGTCCTTT-3' (SEQ ID NO: 200), 5'-AGGCCCTTCGCACTTC-3' (SEQ ID NO: 201), 5'-GCGGUATCCATGTCCCAGGC-3', 5'-GCUGUTTCCATGTCCCAGGC-3', 5'-GCUGUGTCCATGTCCCAGGC-3' 5'-GCCGUTTCCATGTCCCAGGC-3', 5'-GCGGUATCC-3', 5'-GCUGUTTCC-3', 5'-GCUGUGTCC-3', 5'-GCCGUTTCC-3', 5'-CUUCGTGGGGTCCTTUUCAC-3', 5'-CUUCGTGGG-3', 5'-UCG-3', 5'-ACG-3', 5'-ACC-3', 5'-CGC-3', 5'-GAU-3', 5'-GGG-3', 5'-AGC-3', 5'-UUC-3', 5'-UUG-3', a motif or sequence selected from the group consisting of: 5'-CAC-3'; and and oligonucleotides comprising, consisting essentially of, or consisting of variants of that motif or sequence having at least about 75% sequence identity thereto (wherein U can be T and / or T can be U, and wherein the oligonucleotide inhibits TLR9 activity when administered to a subject or in any assay described herein).

[0205] In one embodiment, one, two or more bases of the oligonucleotide are modified bases or one, two or more internucleotide linkages have a modified backbone, preferably the modified bases are 2'OMe.

[0206] In one embodiment, the oligonucleotide comprises, consists essentially of, or consists of the sequence 5'-UCCGGCCTCGGAGTCUCCAU-3' (SEQ ID NO: 52), 5'-GCGGUATCCATAGTCUCCAU-3' (SEQ ID NO: 48), 5'-CCAACACTTCGTGGGGUCCU-3' (SEQ ID NO: 160), 5'-CACUUCGTGGGGTCCUUUUC-3' (SEQ ID NO: 159), 5'-UGACAAAACAATAATAACAG-3' (SEQ ID NO: 167), or a variant thereof having at least about 75% sequence identity thereto (U can be T and / or T can be U).

[0207] In one embodiment, the oligonucleotide comprises, consists essentially of, or consists of the sequence 5'-ACC-3', 5'-CGC-3', 5'-GAU-3', 5'-GGG-3', 5'-UCG-3' or 5'-ACG-3', preferably 5'-mAmCmC-3', 5'-mCmGmC-3', 5'-mGmAmU-3', 5'-mGmGmG-3', 5'-mUmCmG-3' or 5'-mAmCmG-3' (wherein m is a modified base and / or has a modified backbone, preferably the modified base is 2'OMe).

[0208] In one embodiment, the oligonucleotide comprises, consists essentially of, or consists of any of the sequences in Table 1, 1A, 2, 2A, 3, 3A, 4, 4A, 5, 5A, or 6, with or without specific modifications as defined, and is shown in the Examples to inhibit the activity of TLR9.

[0209] In one embodiment, the oligonucleotide comprises, consists essentially of, or consists of any of the sequences in Table 1, 1A, 2, 2A, 3, 3A, 4, 4A, 5, 5A, or 6, with or without the specific modifications defined, and is shown in the Examples to inhibit the activity of TLR9, and the oligonucleotide inhibits or does not substantially inhibit TLR3, TLR7, and / or cGAS activity.

[0210] In one embodiment, the oligonucleotide inhibits TLR9 activity and enhances TLR8 activity, e.g., the oligonucleotide comprises, consists essentially of, or consists of the sequence 5'-UCG-3' or 5'-CGC-3', preferably 5'-mUmCmG-3' or 5'-mCmGmC-3' (where m is a modified base and / or has a modified backbone, preferably the modified base is 2'OMe). In other embodiments, the oligonucleotide inhibits TLR9 activity and does not substantially enhance TLR8 activity, or inhibits TLR8 activity, e.g., the oligonucleotide comprises, consists essentially of, or consists of the sequence 5'-GAU-3', preferably 5'-mGmAmU-3' (where m is a modified base and / or has a modified backbone, preferably the modified base is 2'OMe).

[0211] In any embodiment, the oligonucleotide that inhibits TLR9 activity is at least 3, 4, 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, 15, 16, 17, 18, 19, 20, 30, 40, or 50 nucleotides in length. In one embodiment, the oligonucleotide that inhibits TLR9 activity is at least 3 nucleotides but not more than 20 nucleotides in length, optionally at least 9 nucleotides but not more than 20 nucleotides in length.

[0212] In other embodiments, the oligonucleotide is 5'-mUmCmCmGmGCCTCGGAGTCmUmCmCmAmU-3', 5'-mGmCmGmGmUATCCATAGTCmUmCmCmAmU-3', 5'-mCmCmAmAmCACTTCGTGGGmGmUmCmCmU-3', 5'-mCmAmCmUmUCGTGGGGTCCmUmUmUmUmC-3', 5'-mUmGmAmCmAAAACAATAATmAmAmCmAmG-3', 5'-mUmCmCmGmGCCTCGGAAGCmUmCmUmCmU-3', 5'-mUmCmCmGmGCCTCGGAGTCmUmCmCmAmU-3', The sequence is 5'-mUmCmCmGmGCCTCGGCAGAmUmAmUmCmG-3'. or a variant thereof having at least about 75% sequence identity thereto (wherein U can be T and / or T can be U, and m is a modified base and / or has a modified backbone).

[0213] In other embodiments, the oligonucleotide is 5'-mGmCmGmGmUATCCATGTCCmCmAmGmGmC-3', 5'-mGmCmUmGmUTTCCATGTCCmCmAmGmGmC-3', 5'-mGmCmUmGmUGTCCATGTCCmCmAmGmGmC-3', 5'-mGmCmCmGmUTTCCATGTCCmCmAmGmGmC-3', 5'-mGmCmGmGmUATCC-3', 5'-mGmCmUmGmUTTCC-3', 5'-mGmCmUmGmUGTCC-3', 5'-MGmCmCmGmUTTCC-3', 5'-mCmUmUmCmGTGGGGTCCTTmUmUmCmAmC-3', and The sequence is 5'-mCmUmUmCmGTGGG-3'. or a variant thereof having at least about 75% sequence identity thereto (wherein T can be U and m is a modified base and / or has a modified backbone). Preferably, each base has a modified backbone, and the modified backbone is phosphorothioate.

[0214] In other embodiments, the oligonucleotide is 5'-mG * mC * mG * mG * mU * A* T * C * C * A * T * G * T * C * C * mC * mA * mG * mG * mC-3’、 5’-mU * mC * mC * mG * mG * C * C * T * C * G * G * A * A * G * C * mU * mC * mU * mC * mU-3’、 5’-mU * mC * mC * mG * mG * C * C * T * C * G * G * A * G * T * C * mU * mC * mC * mA * mU-3’、 5’-mU * mC * mC * mG * mG * C * C * T * C * G * G * C * A * G* A * mU * mA * mU * mC * mG-3’、 5’-mG * mC * mU * mG * mU * T * T * C * C * A * T * G * T * C * C * mC * mA * mG * mG * mC-3’、 5’-mG * mC * mU * mG * mU * G * T * C * C * A * T * G * T * C * C * mC * mA * mG * mG * mC-3’、 5’-mG * mC * mC * mG * mU * T * T * C * C * A * T * G * T * C * C * mC * mA * mG * mG * mC-3’、 5’-mG* mC * mG * mG * mU * A * T * C * C * -3’、 5’-mG * mC * mU * mG * mU * T * T * C * C * -3’、 5’-mG * mC * mU * mG * mU * G * T * C * C * -3’、 5’-mG * mC * mC * mG * mU * T * T * C * C * -3’、 5’-mC * mU * mU * mC * mG * T * G * G * G * G * T * C * C * T * T * mU * mU * mC * mA * mC、 5’-mC * mU * mU * mC * mG * T * G * G* G * -3' sequence, (U can be T and / or T can be U, and "m" represents a 2'OMe base; * indicates a phosphorothioate backbone).

[0215] In another aspect, the invention provides an oligonucleotide comprising: a) a 5' region comprising bases that are modified and / or have modified backbones; b) an intermediate region comprising ribonucleic acid, deoxyribonucleic acid, or a combination thereof, optionally with a modified backbone, and bases, wherein at least about 50% of the bases in the intermediate region are adenine bases; c) a 3' region comprising bases that are modified and / or have modified backbones, and a 3' region that inhibits TLR9 activity when administered to a subject.

[0216] In one embodiment, the oligonucleotide is 5'-GAUUAAAACAGATTAAUACA-3' (SEQ ID NO: 165), 5'-GGAUUAAAACAGATTAAUAC-3' (SEQ ID NO: 55), 5'-CUUGUGAAAAGATTAUCUUC-3' (SEQ ID NO: 27), 5'-CCUAGAAAGAAGCAAAGAUU-3' (SEQ ID NO: 164), A motif or sequence selected from the group consisting of: 5'-AAUUUAAAGCATGAAUAUUA-3' (SEQ ID NO: 166), and It includes variants of that motif or sequence (U can be T and / or T can be U) that have at least about 75% sequence identity thereto.

[0217] In one embodiment, the oligonucleotide is 5'-GAUUAAAACAGATTAAUACA-3' (SEQ ID NO: 165), 5'-GGAUUAAAACAGATTAAUAC-3' (SEQ ID NO: 55), 5'-CUUGUGAAAAGATTAUCUUC-3' (SEQ ID NO: 27), 5'-CCUAGAAAGAAGCAAAGAUU-3' (SEQ ID NO: 164), A motif or sequence selected from the group consisting of: 5'-AAUUUAAAGCATGAAUAUUA-3' (SEQ ID NO: 166), and It includes variants of that motif or sequence (U can be T and / or T can be U) that have at least about 75% sequence identity thereto.

[0218] In a further aspect, the present invention provides an oligonucleotide comprising: 5'-[A / G]GU[A / C][U / C]C-3' (SEQ ID NO: 1), 5'-A[G / A][U / G]C[U / C]C-3' (SEQ ID NO: 2), 5'-A[G / A][U / G]C[U / C]C[U / C][C / A]U-3' (SEQ ID NO: 212), 5'-GGUAUA-3' (SEQ ID NO: 4), 5'-UGUUUC-3' (SEQ ID NO: 5), 5'-UGUGUC-3' (SEQ ID NO: 6), 5'-CGUGUC-3' (SEQ ID NO: 8), 5'-GCAGUCTCCATGTCCCAGGC-3' (SEQ ID NO: 30), 5'-GAUGGTTCCAGTCCCUCUUC-3' (SEQ ID NO: 38), 5'-AGCAGTCTCCATGTCCCAGG-3' (SEQ ID NO: 31), 5'-GGGUTCCTCCACACCCUUC-3' (SEQ ID NO: 36), 5'-GGUGGCCACAGGCAACGUCA-3' (SEQ ID NO: 28), 5'-GCCGUTTCCATGTCCCAGGC-3' (SEQ ID NO: 46), 5'-GCGGUATCCATGTCCCAGGC-3' (SEQ ID NO: 42), 5'-GCGGUATACAGGTCCCAGGC-3' (SEQ ID NO: 43), 5'-GCUGUTTCCATGTCCCAGGC-3' (SEQ ID NO: 44), 5'-GCUGUGTCCATGTCCCAGGC-3' (SEQ ID NO: 45), 5'-GCCGUGTCCATGTCCCAGGC-3' (SEQ ID NO: 47), 5'-GCGGUAUCCAUGUCCCAGGC-3' (SEQ ID NO: 151), 5'-GGUATCCCCCCCCCCCCCCC-3' (SEQ ID NO: 54), 5'-GUCCCATCCCTTCTGCUGCC-3' (SEQ ID NO: 145), 5'-UUCUCTCTGGTCCCAUCCCU-3' (SEQ ID NO: 213), 5'-GUUCAGTCAGATCGCUGGGA-3' (SEQ ID NO: 214), 5'-AUGACATTTCGTGGCUCCUA-3' (SEQ ID NO: 215), 5'-UCUCCATGTCCCAGGCCUCC-3' (SEQ ID NO: 216), 5'-AGUCUCCATGTCCCAGGCCU-3' (SEQ ID NO: 217), 5'-CCAUGTCCCAGGCCTCCAGU-3' (SEQ ID NO: 29), 5'-GCAAGGCAGAGAAACUCCAG-3' (SEQ ID NO: 37), 5'-GGAUUAAAACAGATTAAUAC-3' (SEQ ID NO: 55), 5'-AGCCGAACAGAAGGAGCGUC-3' (SEQ ID NO: 40), 5'-UCCGGCCTCGGAAGCUCUCU-3' (SEQ ID NO: 10), 5'-UCCGGCCTCGGAGTCUCCAU-3' (SEQ ID NO: 52), 5'-GCGGUATCCATAGTCUCCAU-3' (SEQ ID NO: 48), 5'-GAUUAAAACAGATTAAUACA-3' (SEQ ID NO: 165), 5'-UGACAAAACAATAATAACAG-3' (SEQ ID NO: 167), 5'-CCAACACTTCGTGGGGUCCU-3' (SEQ ID NO: 160), 5'-GUGUCCTTCATGCTTUGGAU-3' (SEQ ID NO: 21), 5'-GUCCCAGGCCTCCAGUGUCU-3' (SEQ ID NO: 218), 5'-UUGGCCTGTGGATGCUUUGU-3' (SEQ ID NO: 161), 5'-GUCCGTACCTCCACCCACCG-3' (SEQ ID NO: 219), 5'-GUGUUTTTAATTTTGUAGAG-3' (SEQ ID NO: 220), 5'-GUCAAACCTAGAAAGAAGCA-3' (SEQ ID NO: 221), 5'-GGUCUCCTCCACACCCUUCU-3' (SEQ ID NO: 222), 5'-GUCUCCTCCACACCCUUCUC-3' (SEQ ID NO: 146), 5'-UGAUGATGCTTGCAGGAGGC-3' (SEQ ID NO: 223), 5'-UUAAATAATCTAGTTUGAAG-3' (SEQ ID NO: 20), 5'-AAAGCAGTCTCCATGUCCCA-3' (SEQ ID NO: 224), 5'-GCGUAGTTTCTCTTCCUCCC-3' (SEQ ID NO: 41), 5'-UCUGGTCCCATCCCTUCUGC-3' (SEQ ID NO: 35), 5'-UAUUUCCACATGCCCAGUGU-3' (SEQ ID NO: 225), 5'-UGUUUCCCCGGAGAGCAAUG-3' (SEQ ID NO: 39), 5'-UUAGCTCCTTGCCTCGUUCC-3' (SEQ ID NO: 226), 5'-AUUUCCACATGCCCAGUGUU-3' (SEQ ID NO: 33), 5'-UGGCGTAGTTTCTCTUCCUC-3' (SEQ ID NO: 227), 5'-UGACATTTCGTGGCTCCUAC-3' (SEQ ID NO: 228), 5'-GAAAAGATTATCTTCUUUUA-3' (SEQ ID NO: 25), 5'-UGUGAAAAGATTATCUUCUU-3' (SEQ ID NO: 26), 5'-UUGUGAAAAGATTATCUUCU-3' (SEQ ID NO: 229), 5'-UUUGAAATTCAGAAGAUUUG-3' (SEQ ID NO: 230), 5'-AAGCAGTCTCCATGTCCCAG-3' (SEQ ID NO: 231), 5'-AGGAUTAAAACAGATUAAUA-3' (SEQ ID NO: 232), 5'-AGAUUATCTTCTTTTAAUUU-3' (SEQ ID NO: 23), 5'-UCCCAACTCTTCTAACUCGU-3' (SEQ ID NO: 148), 5'-UAAAATAAGGGGAATAGGGG-3' (SEQ ID NO: 233), 5'-AGUGGCACATACCACACCCU-3' (SEQ ID NO: 32), 5'-AAGAUTATCTTCTTTUAAUU-3' (SEQ ID NO: 234), 5'-AGAAAGAAGCAAAGAUUCAA-3' (SEQ ID NO: 22), 5'-UCCCATCCCTTCTGCUGCCA-3' (SEQ ID NO: 235), 5'-ACCCUTCTCTCTGGTCCCAU-3' (SEQ ID NO: 156), 5'-AAUAUCTGCTGCCCACCUUC-3' (SEQ ID NO: 236), 5'-UCUCUCTGGTCCCATCCCUU-3' (SEQ ID NO: 237), 5'-AGGCCTCCAGTGTCTUCUCC-3' (SEQ ID NO: 238), 5'-CAAGCCCCAGCGTTCCUCCG-3' (SEQ ID NO: 239), 5'-AAAUGTCCTGGCCCTCACUG-3' (SEQ ID NO: 162), 5'-AAUUUAAAGCATGAAUAUUA-3' (SEQ ID NO: 166), 5'-AAAAGATTATCTTCTUUUAA-3' (SEQ ID NO: 24), 5'-AUAUCTGCTGCCCACCUUCU-3' (SEQ ID NO: 144), 5'-CAGUCTCCATGTCCCAGGCC-3' (SEQ ID NO: 240), 5'-AAAGATTATCTTCTTUUAAU-3' (SEQ ID NO: 241), 5'-CCCUUCTCTCTGGTCCCAUC-3' (SEQ ID NO: 155), 5'-AUGGCCTTTCCGTGCCAAGG-3' (SEQ ID NO: 9), 5'-GCAUUCCGTGCGGAAGCCUU-3' (SEQ ID NO: 11), 5'-GGCCGAACTTTCCCGCCUUA-3' (SEQ ID NO: 12), 5'-GGUCUTGGCTTCGTGGAGCA-3' (SEQ ID NO: 13), 5'-GGAGCTTCGAGGCCCCAGGC-3' (SEQ ID NO: 14), 5'-GGUGGTCCACAACCCCUUUC-3' (SEQ ID NO: 15), 5'-UUCUGGGGACTTCCAGUUUA-3' (SEQ ID NO: 17), 5'-UGAUUCCAAAGCCAGGGUUA-3' (SEQ ID NO: 18), 5'-GGGUATCGAAAGAGTCUGGA-3' (SEQ ID NO: 242), 5'-CUUGCACGTGGCTTCGUCUC-3' (SEQ ID NO: 243), 5'-GUGUCCTTGCACGTGGCUUC-3' (SEQ ID NO: 244), 5'-GUAAAAAGCTTTTGAAGUGA-3' (SEQ ID NO: 245), 5'-AUGCCATCCACTTGAUAGGC-3' (SEQ ID NO: 246), 5'-UGAAGTAAAAATCAAUAGCG-3' (SEQ ID NO: 247), 5'-AAGGCCCTTCGCACTUCUUA-3' (SEQ ID NO: 248), 5'-GUACUCGTCGGCATCCACCA-3' (SEQ ID NO: 249), 5'-GUCCUTGCACGTGGCUUCGU-3' (SEQ ID NO: 250), 5'-GCCCATCCATGAGGTCCUGG-3' (SEQ ID NO: 251), 5'-GUAAAAGGAGAAAACUAUCU-3' (SEQ ID NO: 252), 5'-UUGAAGTGAAGTAAAAGGAG-3' (SEQ ID NO: 253), 5'-GUUACTCGTGCCTTGGCAAA-3' (SEQ ID NO: 254), 5'-GUCCAAGATCAGCAGUCUCA-3' (SEQ ID NO: 255), 5'-UUCAATGGGAGAATAAAGCA-3' (SEQ ID NO: 256), 5'-GCAAGGCCCTTCGCACUUCU-3' (SEQ ID NO: 257), 5'-GGGUCCACCACTAGCCAGUA-3' (SEQ ID NO: 258), 5'-GUAGAGAAATTATTTUAGGA-3' (SEQ ID NO: 259), 5'-AUCCACCACGTCGTCCAUGU-3' (SEQ ID NO: 260), 5'-GGCAUCCACCACGTCGUCCA-3' (SEQ ID NO: 261), 5'-UUACUTTAAAAGCAAAAGGA-3' (SEQ ID NO: 262), 5'-UUUGAAGTGAAGTAAAAGGA-3' (SEQ ID NO: 263), 5'-GAAGUGAAGTAAAAGGAGAA-3' (SEQ ID NO: 264), 5'-GGCCATCTCTGCTTCUUGGU-3' (SEQ ID NO: 265), 5'-UGGGCTGGAATCCGAGUUAU-3' (SEQ ID NO: 266), 5'-GGAGATTTCAGAGCAGCUUC-3' (SEQ ID NO: 267), 5'-UUUACGGTTTTCAGAAUAUC-3' (SEQ ID NO: 268), 5'-GCGUGTCTGGAAGCTUCCUU-3' (SEQ ID NO: 269), 5'-GCUUATTTTAAGCATAUUAA-3' (SEQ ID NO: 270), 5'-UUAUUTTAAGCATATUAAAA-3' (SEQ ID NO: 271), 5'-UUCUGCAGCTTCCTTGUCCU-3' (SEQ ID NO: 272), 5'-AUUACTTTAAAAGCAAAAGG-3' (SEQ ID NO: 273), 5'-AUUUUAAGCATATTAAAAAG-3' (SEQ ID NO: 274), 5'-UGUGGCTTGTCCTCAGACAU-3' (SEQ ID NO: 275), 5'-AAAAGGAGAAAACTAUCUUC-3' (SEQ ID NO: 276), 5'-GGGUCCATACCCAAGGCAUC-3' (SEQ ID NO: 277), 5'-ACAGUGTTGAGATACUCGGG-3' (SEQ ID NO: 278), 5'-GGAUCTGCATGCCCTCAUCU-3' (SEQ ID NO: 279), 5'-AGUAAAAAGCTTTTGAAGUG-3' (SEQ ID NO: 280), 5'-GUCGUGGCAAATAGTCCUAG-3' (SEQ ID NO: 281), 5'-GGAGATCAGATGAGAGGAGC-3' (SEQ ID NO: 282), 5'-GUGGUTAAGTACATGAGCUC-3' (SEQ ID NO: 283), 5'-GGACACTTAGCTGTTCCUCG-3' (SEQ ID NO: 284), 5'-GUCUCTACTGTTACCUCUGA-3' (SEQ ID NO: 285), 5'-GAGUUCTTCGTAGGCUUCUG-3' (SEQ ID NO: 286), 5'-AAAGUCAAAAAGAAAAACUG-3' (SEQ ID NO: 287), 5'-AAAAGTGGGAAATAAAGGUU-3' (SEQ ID NO: 288), 5'-AGUUUATAGATTTCAAGUAG-3' (SEQ ID NO: 289), 5'-AAAAAGTGGGAAATAAAGGU-3' (SEQ ID NO: 290), 5'-UUUAUATTACAAAGCUACUU-3' (SEQ ID NO: 291), 5'-UGCUATTCATATTTTUAUUU-3' (SEQ ID NO: 292), 5'-GGUAU-3' (SEQ ID NO: 56), 5'-[G / A / C][G / A][G / A / C][T / A / C]TC-3' (SEQ ID NO: 293), 5'-[G / A / C]G[G / A / C][T / A / C]TC-3' (SEQ ID NO: 294), 5'-GGCTTC-3' (SEQ ID NO: 295), 5'-GGCATC-3' (SEQ ID NO: 296), 5'-AGCTTC-3' (SEQ ID NO: 297), 5'-GGAATC-3' (SEQ ID NO: 298), 5'-CACATC-3' (SEQ ID NO: 299), 5'-GGCCTC-3' (SEQ ID NO: 204), 5'-CACTTC-3' (SEQ ID NO: 300), 5'-AAGATC-3' (SEQ ID NO: 301), 5'-TGTCCTTGCACGTGGCTTCG-3' (SEQ ID NO: 302), 5'-TTTGCACACTTCGTACCCAA-3' (SEQ ID NO: 94), 5'-GTCCACATCCTGTGGCTCGT-3' (SEQ ID NO: 303), 5'-TGTGATGGCCTCCCATCTCC-3' (SEQ ID NO: 304), 5'-GGTTTTGGCTGGGATCAAGT-3' (SEQ ID NO: 175), 5'-GGTGTCCTTGCACGTGGCTT-3' (SEQ ID NO: 93), 5'-GGTCCATACCCAAGGCATCC-3' (SEQ ID NO: 305), 5'-GTGTCTTCATCGGCCCTGCC-3' (SEQ ID NO: 306), 5'-GTCTTGGCTTCGTGGAGCAG-3' (SEQ ID NO: 89), 5'-GCTGACAAAGATTCACTGGT-3' (SEQ ID NO: 95), 5'-GAAAGGTTATGCAAGG-3' (SEQ ID NO: 103), 5'-GACTATACGCGCAATA-3' (SEQ ID NO: 307), 5'-TGTGATGGCCTCCCAT-3' (SEQ ID NO: 308), 5'-TCCAACACTTCGTGGG-3' (SEQ ID NO: 114), 5'-GTGTCTGGAAGCTTCC-3' (SEQ ID NO: 106), 5'-CTTGAAGCATCGTATC-3' (SEQ ID NO: 98), 5'-TCGTAGTTGCTTCCTA-3' (SEQ ID NO: 309), 5'-CGCTTTTCTGTCTGGT-3' (SEQ ID NO: 105), 5'-GGCTGGAATCCGAGTT-3' (SEQ ID NO: 310), 5'-GATAGCACCTTCAGCA-3' (SEQ ID NO: 100), 5'-AGGACTCCAGATGTTT-3' (SEQ ID NO: 311), 5'-GTGATCTTGACATGCT-3' (SEQ ID NO: 312), 5'-AGATTTCAGAGCAGCT-3' (SEQ ID NO: 313), 5'-GGTTACGGCTCAGTAT-3' (SEQ ID NO: 314), 5'-GTTCAGTCCTGTCCAT-3' (SEQ ID NO: 315), 5'-AGGTCTTGGCTTCGTG-3' (SEQ ID NO: 316), 5'-CTGCAGCTTCCTTGTC-3' (SEQ ID NO: 191), 5'-GTCCTTGCACGTGGCT-3' (SEQ ID NO: 317), 5'-GTCTCTGGAGCTTCCT-3' (SEQ ID NO: 318), 5'-GGTCTTGGCTTCGTGG-3' (SEQ ID NO: 319), 5'-GGUA-3' (SEQ ID NO: 57), 5'-GUAU-3' (SEQ ID NO: 58), 5'-GGU-3' (SEQ ID NO: 59), 5'-GUA-3' (SEQ ID NO: 60), 5'-GUC-3', 5'-GUG-3', 5'-GUU-3', 5'-GGC-3', 5'-AUC-3', 5'-GAG-3', 5'-GGA-3', 5'-TTT-3', 5'-TCT-3', 5'-GAA-3', 5'-GAC-3', 5'-GAU-3', 5'-AUG-3', 5'-GCG-3', 5'-UUC-3', 5'-GCC-3', 5'-GGG-3', 5'-AUU-3', 5'-GCA-3', 5'-AGC-3', 5'-AAC-3', 5'-CCA-3', 5'-UGC-3', 5'-CAA-3', 5'-CGG-3', 5'-ACC-3', 5'-AGA-3', 5'-TTT-3', 5'-TCT-3', and Oligonucleotides are provided that comprise, consist essentially of, or consist of variants of that motif or sequence having at least about 75% sequence identity thereto (wherein U can be T and / or T can be U, and wherein the oligonucleotide inhibits TLR7 activity when administered to a subject or in any assay described herein).

[0219] In one embodiment, the oligonucleotide is 5'-[A / G]GU[A / C][U / C]C-3' (SEQ ID NO: 1), 5'-A[G / A][U / G]C[U / C]C-3' (SEQ ID NO: 2), 5'-A[G / A][U / G]C[U / C]C[U / C][C / A]U-3' (SEQ ID NO: 212), 5'-GGUAUA-3' (SEQ ID NO: 4), 5'-UGUUUC-3' (SEQ ID NO: 5), 5'-UGUGUC-3' (SEQ ID NO: 6), 5'-CGUGUC-3' (SEQ ID NO: 8), 5'-GCAGUCTCCATGTCCCAGGC-3' (SEQ ID NO: 30), 5'- GAUGGTTCCAGTCCCUCUUC-3' (SEQ ID NO: 38), 5'-AGCAGTCTCCATGTCCCAGG-3' (SEQ ID NO: 31), 5'-GGGUCTCCTCCACACCCUUC-3' (SEQ ID NO: 36), 5'-GGUGGCCACAGGCAACGUCA-3' (SEQ ID NO: 28), 5'-GCCGUTTCCATGTCCCAGGC-3' (SEQ ID NO: 46), 5'-GCGGUATCCATGTCCCAGGC-3' (SEQ ID NO: 42), 5'-GCGGUATACAGGTCCCAGGC-3' (SEQ ID NO: 43), 5'-G CUGUTTCCATGTCCCAGGC-3' (SEQ ID NO: 44), 5'-GCUGUGTCCATGTCCCAGGC-3' (SEQ ID NO: 45), 5'-GCCGUGTCCATGTCCCAGGC-3' (SEQ ID NO: 47), 5'-GCGGUAUCCAUGUCCCAGGC-3' (SEQ ID NO: 151), 5'-GGUATCCCCCCCCCCCCCCCC-3' (SEQ ID NO: 54), 5'-GUCCCATCCCTTCTGCUGCC-3' (SEQ ID NO: 145), 5'-UUCUCTCTGGTCCCAUCCCU-3' (SEQ ID NO: 213), 5' -GUUCAGTCAGATCGCUGGGA-3' (SEQ ID NO: 214), 5'-AUGACATTTCGTGGCUCCUA-3' (SEQ ID NO: 215), 5'-UCUCCATGTCCCAGGCCUCC-3' (SEQ ID NO: 216), 5'-AGUCUCCATGTCCCAGGCCU-3' (SEQ ID NO: 217), 5'-CCAUGTCCCAGGCCTCCAGU-3' (SEQ ID NO: 29), 5'-GCAAGGCAGAGAAACUCCAG-3' (SEQ ID NO: 37), 5'-GGAUUAAAACAGATTAAUAC-3' (SEQ ID NO: 55),The nucleic acid sequence of the present invention comprises, consists essentially of, or consists of the sequence of 5'-AGCCGAACAGAAGGAGCGUC-3' (SEQ ID NO: 40), 5'-UCCGGCCTCGGAAGCUCUCU-3' (SEQ ID NO: 10), 5'-UCCGGCCTCGGAGTCUCCAU-3' (SEQ ID NO: 52), 5'-GCGGUATCCATAGTCUCCAU-3' (SEQ ID NO: 48), 5'-GAUUAAAACAGATTAAUACA-3' (SEQ ID NO: 165), 5'-UGACAAAACAATAATAACAG-3' (SEQ ID NO: 167), 5'-CCAACACTTCGTGGGGUCCU-3' (SEQ ID NO: 160), or a variant thereof having at least about 75% sequence identity thereto (U can be T and / or T can be U).

[0220] In one embodiment, the oligonucleotide is 5'-GUX-3' or 5'-GAX-3' (X is any nucleotide), 5'-GUC-3', 5'-GUG-3', 5'-GUA-3', 5'-GUU-3', 5'-GGC-3', 5'-AUC-3', 5'-GAG-3', 5'-GGA-3', 5'-TTT-3', 5'-TCT-3', 5'-GAA-3', 5'-GAC-3', 5'-GAU-3', 5'-AUG-3', 5'-GCG-3', 5'-UUC-3', 5'-GCC-3', 5'-GGG-3', 5'- AUU-3', 5'-GCA-3', 5'-AGC-3', 5'-AAC-3', 5'-CCA-3', 5'-UGC-3', 5'-CAA-3', 5'-CGG-3', 5'-ACC-3', 5'-AGA-3', 5'-TTT-3' or 5'-TCT-3' (preferably in which one or two bases are modified and one or more internucleotide linkages are modified backbone), preferably 5'-mGmUmX-3' or 5'-mGmAmX-3' (X is any nucleotide), 5'-mGmUmC-3', 5'-mGmUmG-3' ', 5'-mGmUmA-3', 5'-mGmUmU-3', 5'-mGmGmC-3', 5'-mAmUmC-3', 5'-mGmAmG-3', 5'-mGmGmA-3', 5'-mTmTmT-3' or 5-mTmCmT-3', 5'-mGmAmA-3', 5 '-mGmAmC-3', 5'-mGmAmU-3', 5'-mAmUmG-3', 5'-mGmCmG-3', 5'-mUmUmC-3', 5'-mGmCmC-3', 5'-mGmGmG-3', 5'-mAmUmU-3', 5'-mGmCmA-3', 5'-m The nucleic acid sequence of the present invention may comprise, consist essentially of, or consist of the sequence AmGmC-3', 5'-mAmAmC-3', 5'-mCmCmA-3', 5'-mUmGmC-3', 5'-mCmAmA-3', 5'-mCmGmG-3', 5'-mAmCmC-3', 5'-mAmGmA-3', 5'-mUmUmU-3', 5'mUmCmU-3', 5'mTmTm-3', or 5'-mTmCmT-3' (wherein m is a modified base and / or has a modified backbone, preferably the modified base is 2'OMe and the modified backbone is phosphorothioate).

[0221] In one embodiment, the oligonucleotide comprises, consists essentially of, or consists of any of the sequences in Table 1, 1A, 2, 2A, 3, 3A, 4, 4A, 5, 5A, or 6, with or without specific modifications as defined, and is shown in the Examples to inhibit the activity of TLR7.

[0222] In one embodiment, the oligonucleotide comprises, consists essentially of, or consists of any of the sequences in Table 1, 1A, 2, 2A, 3, 3A, 4, 4A, 5, 5A, or 6, with or without the specific modifications defined, and is shown in the Examples to inhibit the activity of TLR7, and the oligonucleotide inhibits or does not substantially inhibit TLR3, TLR9, and / or cGAS activity.

[0223] In one embodiment, the oligonucleotide inhibits TLR7 activity and inhibits TLR8 activity, for example, the oligonucleotide is 5'-GUX-3' or 5'-GAX-3' (X is any nucleotide), 5'-GUC-3', 5'-GUG-3', 5'-GUA-3', 5'-GUU-3', or 5'-GAG-3', 5'-GAC-3', 5'-GAU-3', 5'-GAA-3', preferably 5'-mGmUmX-3' or 5'-mGmAmX-3' (X is any nucleotide). is any nucleotide), 5'-mGmUmC-3', 5'-mGmUmG-3', 5'-mGmUmA-3', 5'-mGmUmU-3', 5'-mGmAmG-3', 5'-mGmAmC-3', 5'-mGmAmU-3', 5'-mGmAmA-3' (where m is a modified base and / or has a modified backbone, preferably the modified base is 2'OMe and the modified backbone is phosphorothioate).

[0224] With reference to the previous embodiment, the oligonucleotide may comprise: a) a 5' region comprising bases that are modified and / or have modified backbones; b) an intermediate region comprising bases of ribonucleic acid, deoxyribonucleic acid, or a combination thereof, optionally with a modified backbone; c) a 3' region that includes bases that are modified and / or have modified backbones.

[0225] In one embodiment, the intermediate region is about 10 bases in length.

[0226] In one embodiment, the 5' and / or 3' regions are (a) about 3 bases in length, or (b) about 5 bases in length. In embodiments where the 5' and / or 3' regions are about 5 bases in length, the bases are preferably 2'-OMe and / or 2'-MOE modified. In embodiments where the 5' and / or 3' regions are about 3 bases in length, the bases are preferably 2'-LNA modified.

[0227] In embodiments where the motif is 5'-GGUAUA-3' (SEQ ID NO:4), 5'-GGUAU-3' (SEQ ID NO:56), 5'-GGUA-3' (SEQ ID NO:57), 5'-GUAU-3' (SEQ ID NO:58), 5'-GGU-3' (SEQ ID NO:59), or 5'-GUA-3' (SEQ ID NO:60) (U can be T), the motif is suitably at or towards the 5' and / or 3' end of the oligonucleotide. In particular embodiments, the 5'-GGUAUA-3' (SEQ ID NO:4), 5'-GGUAU-3' (SEQ ID NO:56), 5'-GGUA-3' (SEQ ID NO:57), 5'-GUAU-3' (SEQ ID NO:58), 5'-GGU-3' (SEQ ID NO:59), or 5'-GUA-3' (SEQ ID NO:60) (U can be T) motif is at or towards the 5' end of the oligonucleotide.

[0228] In some embodiments, the oligonucleotide comprises, consists essentially of, or consists of the sequence 5'-GGUAU-3' (SEQ ID NO: 56) or a variant thereof having at least about 75% sequence identity thereto (wherein U can be T).

[0229] In other embodiments, the oligonucleotide comprises, consists essentially of, or consists of the sequence 5'-GGUA-3' (SEQ ID NO: 57) or a variant thereof having at least about 75% sequence identity thereto (wherein U can be T).

[0230] In some embodiments, the oligonucleotide comprises, consists essentially of, or consists of the sequence 5'-GUAU-3' (SEQ ID NO: 58) or a variant thereof having at least about 75% sequence identity thereto (wherein U can be T).

[0231] In certain embodiments, the oligonucleotide comprises, consists essentially of, or consists of the sequence 5'-GGU-3' (SEQ ID NO: 59) or a variant thereof having at least about 75% sequence identity thereto (wherein U can be T).

[0232] In some embodiments, the oligonucleotide comprises, consists essentially of, or consists of the sequence 5'-GUA-3' (SEQ ID NO: 60) or a variant thereof having at least about 75% sequence identity thereto (wherein U can be T).

[0233] In other embodiments, the oligonucleotide is 5'-mGmGmUmAmU-3', 5'-mGmGmUmA-3', 5'-mGmUmAmU-3', 5'-mGmGmU-3', The sequence 5'-mGmUmA-3', or a variant thereof having at least about 75% sequence identity thereto (wherein U can be T and / or T can be U, and m is a modified base and / or has a modified backbone).

[0234] In a further aspect, the present invention provides an oligonucleotide comprising: 5'-CUUCGTGGGGTCCTTUUCAC-3', 5'-CUUCG-3', 5'-CUUCGTG-3', 5'-CUUCGTGGG-3', 5'-UCG-3', 5'-UCA-3', 5'-CGG-3', 5'-UGG-3', 5'-CGC-3', 5'-AGG-3', 5'-GGA-3', 5'-GGC-3', 5'-AGA-3', 5'-CGA-3', 5'-UAG-3', 5'-UCU-3', 5'-AGC-3', 5'-GGU-3', 5'-UGA-3', 5'-AGU-3', 5'-ACG-3', 5'-CGU-3', 5'-UCC-3', 5'-GCG-3', 5'-GGG-3', 5'-UGU-3', 5'-UCA-3', 5'-CUG-3', 5'-UUG-3', 5'-UUA-3', 5'-UGC-3', a motif or sequence selected from the group consisting of: and Oligonucleotides are provided that include variants of that motif or sequence having at least about 75% sequence identity thereto (wherein U can be T and / or T can be U, and wherein the oligonucleotide enhances TLR8 activity when administered to a subject or in any assay described herein).

[0235] In one embodiment, the oligonucleotide is 5'-CGX-3', 5'-AGX-3', 5'GGX-3' (wherein X is any nucleotide), 5'-UGG-3', 5'-UCA-3', 5'-CGG-3', 5'-UGG-3', 5'-CGC-3', 5'-AGG-3', 5'-GGA-3', 5'-GGC-3', 5'-AGA-3', 5'-CGA-3', 5'-UAG-3', 5'-UCU-3', 5'-AGC-3', 5'-GGU-3', 5'-U GA-3', 5'-AGU-3', 5'-ACG-3', 5'-CGU-3', 5'-UCC-3', 5'-GCG-3', 5'-GGG-3', 5'-UGU-3', 5'-UCA-3', 5'-CUG-3', 5'-UUG-3', 5'-UUA-3' or 5'-UGC-3' (preferably one or two bases are modified bases and / or one or more internucleotide linkages have a modified backbone), more preferably 5'-mUmCmG-3', 5'-mUm CmA-3', 5'-mCmGmG-3', 5'-mUmGmG-3', 5'-mCmGmC-3', 5'-mAmGmG-3', 5'-mGmGmA-3', 5'-mGmGmC-3', 5'-mAmGmA-3', 5'-mCmG mA-3', 5'-mUmAmG-3', 5'-mUmCmU-3', 5'-mAmGmC-3', 5'-mGmGmU-3', 5'-mUmGmA-3'5'-mAmGmU-3', 5'-mAmCmG-3', 5'-mCmGmU- 3', 5'-mUmCmC-3', 5'-mGmCmG-3', 5'-mGmGmG-3', 5'-mUmGmU-3', 5'-mUmCmA-3', 5'-mCmUmG-3', 5'-mUmUmG-3', 5'-mUmUmA-3' or 5'-mUmGmC-3' (where m is a modified base and / or has a modified backbone, preferably the modified base is 2'OMe and the modified backbone is phosphorothioate).

[0236] In one embodiment, the oligonucleotide is mC * mU * mU * C * G * T * G *G * G * G * T * C * C * T * T * mU * mU * mC * mA * mC (m is a modified base, 2'OMe, * is a modified backbone phosphorothioate). The oligonucleotide may have an LNA in the first CG.

[0237] In one embodiment, the oligonucleotide comprises, consists essentially of, or consists of any of the sequences in Table 1, 1A, 2, 2A, 3, 3A, 4, 4A, 5, 5A, or 6, with or without specific modifications as defined, and is shown in the Examples to enhance the activity of TLR8.

[0238] In one embodiment, the oligonucleotide comprises, consists essentially of, or consists of any of the sequences in Table 1, 1A, 2, 2A, 3, 3A, 4, 4A, 5, 5A, or 6, with or without the specific modifications defined, and is shown in the Examples to enhance the activity of TLR8, and the oligonucleotide inhibits or does not substantially inhibit TLR3, TLR7, TLR9, and / or cGAS activity.

[0239] In a further aspect, the present invention provides an oligonucleotide comprising: 5'-GAG-3', 5'-GAC-3', 5'-GAU-3', 5'-GAA-3', 5'-GUC-3', 5'-GUU-3', 5'-GUA-3', 5'-GUG-3', 5'-AUA-3', 5'-AUG-3', 5'-CUU-3', 5'-AAG-3', 5'-AUC-3', 5'-CCC-3', 5'-GCU-3', 5'-CCU-3', 5'-CUA-3', 5'-CUC-3', a motif or sequence selected from the group consisting of: 5'-AAC-3'; and (U can be T and / or T can be U, and wherein the oligonucleotide inhibits TLR8 activity when administered to a subject or in any assay described herein).

[0240] In one embodiment, the oligonucleotide is 5'-GAX-3' or 5'-GUX-3' (X is any nucleotide), 5'-GAG-3', 5'-GAC-3', 5'-GAU-3', 5'-GAA-3', 5'-GUC-3', 5'-GUU-3', 5'-GUA-3', 5'-GUG-3', 5'-AUA-3', 5'-AUG-3', 5'-CUU-3', 5'-AA G-3', 5'-AUC-3', 5'-CCC-3', 5'-GCU-3', 5'-CCU-3', 5'-CUA-3, 5'-CUC-3 or 5'-AAC-3' (preferably one or two bases are modified bases and / or one or more internucleotide linkages have a modified backbone), more preferably 5'-mGmAmX-3' or 5'-mGmUmX-3' (X is any nucleoside). ), 5'-mGmAmG-3', 5'-mGmAmC-3', 5'-mGmAmU-3', 5'-mGmAmA-3', 5'-mGmUmC-3', 5'-mGmUmU-3', 5'-mGmUmA-3', 5'-mGmUmG-3', 5'-mAmUmA-3', 5'-mAmUmG-3', 5'-mCmUmU-3', 5'-mAmAmG-3', 5'-mAmUm C-3', 5'-mCmCmC-3', 5'-mGmCmU-3', 5'-mCmCmU-3', 5'-mCmUmA-3, 5'-mCmUmC-3 or 5'-mAmAmC-3' (where m is a modified base and / or has a modified backbone, preferably the modified base is 2'OMe and the modified backbone is phosphorothioate).

[0241] In one embodiment, the oligonucleotide comprises, consists essentially of, or consists of any of the sequences in Table 1, 1A, 2, 2A, 3, 3A, 4, 4A, 5, 5A, or 6, with or without specific modifications as defined, and is shown in the Examples to inhibit the activity of TLR8.

[0242] In one embodiment, the oligonucleotide comprises, consists essentially of, or consists of any of the sequences in Table 1, 1A, 2, 2A, 3, 3A, 4, 4A, 5, 5A, or 6, with or without the specific modifications defined, and is shown in the Examples to inhibit the activity of TLR8, and the oligonucleotide inhibits or does not substantially inhibit TLR3, TLR7, TLR9, and / or cGAS activity.

[0243] In a further aspect, the present invention provides an oligonucleotide comprising: 5'-TAC-3', 5'-CGC-3', 5'-GCA-3', 5'-UGA-3', 5'-CAG-3', 5'-UGG-3', 5'-UCA-3', 5'-TGA-3', 5'-CGT-3', 5'-GAC-3', 5'-CCA-3', 5'-TAG-3', 5'-TGG-3', 5'-TCA-3', 5'-TGC-3', 5'-CAC-3', 5'-CGG-3', 5'-CCC-3', 5'-ACT-3', 5'-GTA-3', 5'-GGA-3', 5'-AAG-3', 5'-ATA-3', 5'-GUC-3', 5'-UCC-3', 5'-AUC-3', 5'-CCG-3', 5'-CAA-3', 5'-GAU-3', a motif or sequence selected from the group consisting of: 5'-CGA-3'; and (U can be T and / or T can be U, wherein the oligonucleotide inhibits TLR3 activity when administered to a subject or in any assay described herein).

[0244] In one embodiment, the oligonucleotide comprises, consists essentially of, or consists of the sequence 5'-TAC-3', 5'-CGC-3', 5'-GCA-3', 5'-UGA-3', 5'-CAG-3', 5'-UGG-3', or 5'-UCA-3', preferably 5'-mTmAmC-3', 5'-mCmGmC-3', 5'-mGmCmA-3', 5'-mUmGmA-3', 5'-mCmAmG-3', 5'-mUmGmG-3', 5'-mUmCmA-3' (wherein m is a modified base and / or has a modified backbone, preferably the modified base is 2'OMe and the modified backbone is phosphorothioate).

[0245] In one embodiment, the oligonucleotide comprises, consists essentially of, or consists of any of the sequences in Table 1, 1A, 2, 2A, 3, 3A, 4, 4A, 5, 5A, 6, or 7, with or without specific modifications as defined, and is shown in the Examples to inhibit the activity of TLR3.

[0246] In one embodiment, the oligonucleotide comprises, consists essentially of, or consists of any of the sequences in Table 1, 1A, 2, 2A, 3, 3A, 4, 4A, 5, 5A, 6, or 7, with or without the specific modifications defined, and is shown in the Examples to inhibit the activity of TLR3, and the oligonucleotide inhibits or does not substantially inhibit TLR8, TLR7, TLR9, and / or cGAS activity.

[0247] In yet another aspect, the invention provides a composition comprising, consisting essentially of, or consisting of an oligonucleotide of any of the above aspects or embodiments.

[0248] In one embodiment, the composition further comprises a pharmaceutically or physiologically acceptable carrier.

[0249] In one embodiment, the composition consists essentially of the oligonucleotide of any of the above aspects or embodiments and a pharmaceutically acceptable carrier.

[0250] In one embodiment, the composition further comprises a non-toxic pharmaceutically or physiologically acceptable carrier.

[0251] In one embodiment, the only active pharmaceutical ingredient present in the composition is the oligonucleotide of the above aspect or embodiment. Preferably, prior to use, i.e., prior to administration to a subject or contact with a cell, the only active pharmaceutical ingredient present in the composition is the oligonucleotide of the above aspect or embodiment. In any embodiment, at least about 10%, at least about 15%, at least about 20%, at least about 25%, at least about 30%, at least about 35%, at least about 40%, at least about 45%, at least about 50%, at least about 55%, at least about 60%, at least about 65%, at least about 70%, at least about 75%, at least about 80%, at least about 85%, at least about 90%, at least about 95%, or at least about 100% of the oligonucleotide content of the composition is the oligonucleotide of any of the above aspects or embodiments. Preferably, the composition has the above oligonucleotide content before use, i.e., before administration to a subject or contact with a cell.

[0252] In any embodiment, 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%, at least 65%, at least 70%, at least 75%, at least 80%, at least 85%, at least 90%, at least 95%, or at least 100% of the oligonucleotide content of the composition is an oligonucleotide of any of the above aspects or embodiments. Preferably, the composition has the above oligonucleotide content prior to use, i.e., before administration to a subject or contact with a cell.

[0253] In any embodiment, at least about 10%, at least about 15%, at least about 20%, at least about 25%, at least about 30%, at least about 35%, at least about 40%, at least about 45%, at least about 50%, at least about 55%, at least about 60%, at least about 65%, at least about 70%, at least about 75%, at least about 80%, at least about 85%, at least about 90%, at least about 95%, or at least about 100% of the active pharmaceutical ingredient in the composition is an oligonucleotide of any of the above aspects or embodiments.

[0254] In a further aspect, the invention relates to a method for reducing expression of a target gene in a cell, comprising contacting the cell with an oligonucleotide or composition of the above aspect.

[0255] In yet another aspect, the invention pertains to a method of treating or preventing a disease, disorder, or condition in a subject, comprising administering to the subject a therapeutically effective amount of an oligonucleotide or composition of the above aspect, thereby treating or preventing the disease, disorder, or condition in the subject.

[0256] In a related aspect, the invention provides the use of an oligonucleotide or composition of the above aspect in the manufacture of a medicament for treating or preventing a disease, disorder or condition in a subject.

[0257] In another related aspect, the invention relates to an oligonucleotide or composition of the above aspect for use in treating or preventing a disease, disorder or condition in a subject.

[0258] In one embodiment of the above three aspects, the oligonucleotide or composition reduces expression of a target gene involved in the disease, disorder or condition.

[0259] Suitably, the disease, disorder or condition of the three aforementioned aspects exhibits increased, excessive or abnormal cGAS expression, activity and / or signalling.

[0260] In one embodiment, the disease, disorder or condition is selected from the group consisting of Huntington's disease, Parkinson's disease, motor neuron disease (MND), amyotrophic lateral sclerosis (ALS), prion disease, frontotemporal dementia, traumatic brain injury, Alzheimer's disease, acute pancreatitis, silica-induced fibrosis, age-related macular degeneration, Aicardi syndrome, myocardial infarction, heart failure, polyarthritis / fetal and neonatal anemia, systemic lupus erythematosus, acute kidney injury, alcohol-related liver disease, non-alcoholic fatty liver disease, silica-driven pulmonary inflammation, chronic obstructive pulmonary disease, brain damage after ischemic stroke, sepsis, non-alcoholic steatohepatitis (NASH), cancer, sickle cell disease, inflammatory bowel disease, type 2 diabetes mellitus, overnutrition-induced obesity, COVID-19, chronic obstructive pulmonary disorder (COPD), hematopoietic disorders, aging-associated inflammation, Cutibacterium acnes, and the like. acnes infection, hepatitis B, posterior segment eye disease, arthritis, rheumatoid arthritis, emphysema, colorectal cancer, skin cancer, metastasis, and breast cancer.

[0261] In some embodiments, the disease, disorder, or condition of any of the three foregoing aspects is an age-related disease, disorder, or condition, such as a senescence and / or senescence-related disease, disorder, or condition. In this regard, the oligonucleotide preferably inhibits cGAS activity when administered to a subject.

[0262] Suitably, the disease, disorder or condition of the three aforementioned aspects exhibits increased, excessive or abnormal TLR9 expression, activity and / or signalling. In one embodiment, the disease, disorder or condition is selected from the group consisting of psoriasis, rheumatoid arthritis, alopecia universalis, acute disseminated encephalomyelitis, Addison's disease, allergy, ankylosing spondylitis, antiphospholipid syndrome, arteriosclerosis, atherosclerosis, autoimmune hemolytic anemia, autoimmune hepatitis, bullous pemphigoid, Chagas' disease, chronic obstructive pulmonary disease, celiac disease, cutaneous lupus erythematosus (CLE), dermatomyositis, diabetes, dilated cardiomyopathy (DC), uterine adenomyosis, Goodpasture's syndrome, Graves' disease, Guillain-Barré syndrome, Hashimoto's disease, hidradenitis suppurativa, idiopathic thrombocytopenic purpura, inflammatory bowel disease, interstitial pneumonia, and the like. In some embodiments, the therapeutic agent is selected from the group consisting of: chronic cystitis, moles, multiple sclerosis (MS), myasthenia gravis, myocarditis, narcolepsy, neuromyotonia, pemphigus, pernicious anemia, polymyositis, primary biliary cirrhosis, rheumatoid arthritis (RA), schizophrenia, Sjogren's syndrome, systemic lupus erythematosus (SLE), systemic sclerosis, temporal arteritis, vasculitis, vitiligo, vulvodynia, Wegener's granulomatosis, trauma pain, neuropathic pain, and acetaminophen poisoning, breast cancer, cervical squamous cell carcinoma, gastric cancer, glioma, hepatocellular carcinoma, lung cancer, melanoma, prostate cancer, recurrent glioblastoma, recurrent non-Hodgkin's lymphoma, and colorectal cancer.

[0263] In one embodiment, the disease, disorder or condition exhibits increased, excessive or abnormal TLR7 expression, activity and / or signaling.

[0264] In one embodiment, the disease, disorder or condition of the three aforementioned aspects exhibits increased, excessive or abnormal TLR8 expression, activity and / or signaling.

[0265] In one embodiment, the disease, disorder or condition of the three aforementioned aspects exhibits increased, excessive or abnormal TLR3 expression, activity and / or signaling.

[0266] In other embodiments, the disease, disorder, or condition of any of the three foregoing aspects is an inflammatory disease, disorder, or condition associated with administration of a therapeutic oligonucleotide to a subject. In this regard, the inflammatory disease, disorder, or condition is associated, at least in part, with activation of one or more nucleic acid sensors, such as cGAS, TLR3, TLR8, TLR9, and / or TLR7, following administration of the therapeutic oligonucleotide. In one embodiment, the inflammatory disease, disorder, or condition comprises liver inflammation.

[0267] In a further aspect, the invention pertains to a method of inhibiting cGAS in a subject, comprising administering to the subject an effective amount of an oligonucleotide or composition of the above aspects.

[0268] In a related aspect, the invention pertains to a method of inhibiting cGAS in a cell, comprising contacting the cell with an effective amount of an oligonucleotide or composition of the above aspect.

[0269] In particular embodiments of the above two aspects, the oligonucleotide comprises, consists of, or consists essentially of a motif selected from 5'-GGUAUA-3' (SEQ ID NO:4), 5'-GGUAU-3' (SEQ ID NO:56), 5'-GGUA-3' (SEQ ID NO:57), 5'-GUAU-3' (SEQ ID NO:58), 5'-GUA-3' (SEQ ID NO:60), and 5'-GGU-3' (SEQ ID NO:59), where U can be T. In such instances, the motif is suitably at or towards the 5' end of the oligonucleotide.

[0270] In one embodiment, the oligonucleotide inhibits or prevents aging in cells.

[0271] In some embodiments, the cell is an immune cell.

[0272] In one embodiment, the cells are in cell culture. In one embodiment, the method comprises culturing the cells. In one embodiment, the method produces more viable cells than cells cultured under the same conditions but lacking oligonucleotides. Thus, the method can be used to produce a given number of cells with fewer passages.

[0273] In one embodiment of the above aspect, the oligonucleotide comprises: 5'-GCAGUCTCCATGTCCCAGGC-3' (SEQ ID NO: 30), 5'-GAUGGTTCCAGTCCCUCUUC-3' (SEQ ID NO: 38), 5'-AGCAGTCTCCATGTCCCAGG-3' (SEQ ID NO: 31), 5'-GGGUTCCTCCACACCCUUC-3' (SEQ ID NO: 36), 5'-GGUGGCCACAGGCAACGUCA-3' (SEQ ID NO: 28), 5'-GCCGUTTCCATGTCCCAGGC-3' (SEQ ID NO: 46), 5'-GCGGUATCCATGTCCCAGGC-3' (SEQ ID NO: 42), 5'-GCGGUATACAGGTCCCAGGC-3' (SEQ ID NO: 43), 5'-GCUGUTTCCATGTCCCAGGC-3' (SEQ ID NO: 44), 5'-GCUGUGTCCATGTCCCAGGC-3' (SEQ ID NO: 45), 5'-GCCGUGTCCATGTCCCAGGC-3' (SEQ ID NO: 47), 5'-GCGGUAUCCAUGUCCCAGGC-3' (SEQ ID NO: 151), 5'-GGUATCCCCCCCCCCCCCCC-3' (SEQ ID NO: 54), 5'-CUUGUGAAAAGATTAUCUUC-3' (SEQ ID NO: 27), 5'-CCAUGTCCCAGGCCTCCAGU-3' (SEQ ID NO: 29), 5'-GCAAGGCAGAGAAACUCCAG-3' (SEQ ID NO: 37), 5'-GGAUUAAAACAGATTAAUAC-3' (SEQ ID NO: 55), 5'-AGCCGAACAGAAGGAGCGUC-3' (SEQ ID NO: 40), 5'-UCCGGCCTCGGAAGCUCUCU-3' (SEQ ID NO: 10), 5'-UCCGGCCTCGGAGTCUCCAU-3' (SEQ ID NO: 52), The sequence of 5'-GCGGUATCCATAGTCUCCAU-3' (SEQ ID NO: 48), or a variant thereof having at least about 75% sequence identity thereto (U may be T and / or T may be U).

[0274] In one embodiment, the oligonucleotide comprises, consists essentially of, or consists of the sequence 5'-GCGGUATCCATGTCCCAGGC-3' (SEQ ID NO: 42), or a variant thereof having at least about 75% sequence identity thereto (U can be T and / or T can be U).

[0275] In another embodiment, the oligonucleotide comprises, consists essentially of, or consists of the sequence 5'-mGmCmGmGmUATCCATGTCCmCmAmGmGmC-3', or a variant thereof having at least about 75% sequence identity thereto (U can be T and / or T can be U, and m is a modified base and / or has a modified backbone).

[0276] In another embodiment, the oligonucleotide comprises, consists essentially of, or consists of the sequence 5'-mGmCmGmGmUmAmTmCmCmAmTmGmTmCmCmCmAmGmGmC-3', or a variant thereof having at least about 75% sequence identity thereto (U can be T and / or T can be U, and m is a modified base and / or has a modified backbone).

[0277] In a still further aspect, the present invention provides a method of inhibiting TLR9 in a subject, the method comprising administering to the subject an effective amount of an oligonucleotide or composition of the above aspect.

[0278] In a related aspect, the invention pertains to a method of inhibiting TLR9 in a cell, comprising contacting the cell with an effective amount of an oligonucleotide or composition of the above aspect.

[0279] With respect to the above two aspects, the oligonucleotide preferably inhibits TLR9 activation in a cell or subject in response to an RNA molecule, such as an exogenous RNA molecule.

[0280] In one embodiment of the above aspect, the oligonucleotide comprises, consists essentially of, or consists of the sequence 5'-UCCGGCCTCGGAGTCUCCAU-3' (SEQ ID NO: 52), 5'-GCGGUATCCATAGTCUCCAU-3' (SEQ ID NO: 48), 5'-CCAACACTTCGTGGGGUCCU-3' (SEQ ID NO: 160), 5'-CACUUCGTGGGGTCCUUUUC-3' (SEQ ID NO: 159), 5'-UCCGGCCTCGGAAGCUCUCU-3' (SEQ ID NO: 10), 5'-GAUUAAAACAGATTAAUACA-3' (SEQ ID NO: 165), 5'-UGACAAAACAATAATAACAG-3' (SEQ ID NO: 167), 5'-UCCGGCCTCGGAAGCUCUCU-3' (SEQ ID NO: 10), or a variant thereof having at least about 75% sequence identity thereto (U can be T and / or T can be U).

[0281] In one embodiment, the oligonucleotide comprises, consists essentially of, or consists of the sequence 5'-UCCGGCCTCGGAAGCUCUCU-3' (SEQ ID NO: 10), or a variant thereof having at least about 75% sequence identity thereto (U can be T and / or T can be U).

[0282] In another embodiment, the oligonucleotide comprises, consists essentially of, or consists of the sequence 5'-mUmCmCmGmGCCTCGGAAGCmUmCmUmCmU-3', or a variant thereof having at least about 75% sequence identity thereto (U can be T and / or T can be U, and m is a modified base and / or has a modified backbone).

[0283] In another aspect, the invention pertains to a method of inhibiting TLR7 in a subject, comprising administering to the subject an effective amount of an oligonucleotide or composition of the above aspects.

[0284] In a related aspect, the invention provides a method of inhibiting TLR7 in a cell, comprising contacting the cell with an effective amount of an oligonucleotide or composition of the above aspect.

[0285] With respect to the above two aspects, the oligonucleotide or composition preferably inhibits TLR7 activation in a cell or subject in response to an RNA molecule, such as an exogenous RNA molecule, or a TLR7 agonist, such as a small molecule.

[0286] In another related aspect, the invention relates to a method for preventing or inhibiting TLR7 activation by an RNA molecule in a cell, comprising contacting the cell with an effective amount of an oligonucleotide of the above aspect.

[0287] In a further related aspect, the invention pertains to a method for preventing or inhibiting TLR7 activation by an RNA molecule or a TLR7 agonist in a subject, the method comprising administering to the subject an effective amount of an oligonucleotide of the above aspect.

[0288] In a still further aspect, the present invention provides a method of inhibiting TLR8 in a subject, the method comprising administering to the subject an effective amount of an oligonucleotide of the above aspect.

[0289] In a related aspect, the invention pertains to a method of inhibiting TLR8 in a cell, comprising contacting the cell with an effective amount of an oligonucleotide of the above aspect.

[0290] With respect to the above two aspects, the oligonucleotide or composition preferably inhibits TLR8 activation in a cell or subject in response to an RNA molecule, such as an exogenous RNA molecule, or a small molecule, e.g., a TLR8 agonist, such as motolimod.

[0291] In another related aspect, the invention relates to a method for preventing or inhibiting TLR8 activation by an RNA molecule or a TLR8 agonist in a cell, comprising contacting the cell with an effective amount of an oligonucleotide or composition of the above aspect.

[0292] In a further related aspect, the invention pertains to a method for preventing inhibition of TLR8 activation by an RNA molecule in a subject, the method comprising administering to the subject an effective amount of an oligonucleotide or composition of the above aspect.

[0293] In a still further aspect, the present invention provides a method of inhibiting TLR3 in a subject, the method comprising administering to the subject an effective amount of an oligonucleotide of the above aspect.

[0294] In a related aspect, the invention pertains to a method of inhibiting TLR3 in a cell, comprising contacting the cell with an effective amount of an oligonucleotide of the above aspect.

[0295] With respect to the above two aspects, the oligonucleotide or composition preferably inhibits TLR3 activation in a cell or subject in response to an RNA molecule, such as an exogenous RNA molecule, or a TLR3 agonist, such as a small molecule.

[0296] In another related aspect, the invention relates to a method for preventing or inhibiting TLR3 activation by an RNA molecule or a TLR3 agonist in a cell, comprising contacting the cell with an effective amount of an oligonucleotide or composition of the above aspect.

[0297] In a further related aspect, the invention pertains to a method for preventing inhibition of TLR3 activation by an RNA molecule or a TLR3 agonist in a subject, comprising administering to the subject an effective amount of an oligonucleotide or composition of the above aspect.

[0298] In a still further aspect, the present invention provides a method of increasing the activity of or enhancing TLR8 in a subject, the method comprising the step of administering to the subject an effective amount of the oligonucleotide of the above aspect.

[0299] In a related aspect, the invention pertains to a method of increasing the activity of or enhancing TLR8 in a cell, comprising contacting the cell with an effective amount of an oligonucleotide of the above aspect.

[0300] In the two embodiments described above, the oligonucleotide or composition preferably increases TLR8 activation in a cell or subject in response to an RNA molecule, such as an exogenous RNA molecule, or a small molecule, such as a TLR8 agonist, such as motolimod. Thus, enhancing TLR8 activity with the oligonucleotide of the present invention in the presence of a co-administered TLR8 agonist allows for a lower dose of the TLR8 agonist to be administered in the present invention. Other TLR7 / 8 agonists that can be enhanced include R848, loxoribine, gardikimod, isatoribine, imiquimod, CL075, CL097, CL264, CL307, 852A, or TL8-506.

[0301] In another related aspect, the invention relates to a method for increasing or enhancing TLR8 activation by an RNA molecule in a cell, comprising contacting the cell with an effective amount of an oligonucleotide or composition of the above aspect.

[0302] In a further related aspect, the invention pertains to a method for increasing or enhancing TLR8 activation by an RNA molecule in a subject, comprising administering to the subject an effective amount of an oligonucleotide or composition of the above aspect.

[0303] In a particular embodiment, the RNA molecule is a messenger RNA (mRNA) molecule. Suitably, the mRNA molecule is a component of or is included in an immunogenic composition, such as an mRNA vaccine composition.

[0304] In another aspect, the present invention provides an immunogenic composition comprising an RNA molecule and an oligonucleotide according to the above aspects.

[0305] Preferably, the immunogenic composition is an mRNA vaccine composition.

[0306] In one embodiment of the above aspect, the oligonucleotides are 5'-GCAGUCTCCATGTCCCAGGC-3' (SEQ ID NO: 30), 5'-GAUGGTTCCAGTCCCUCUUC-3' (SEQ ID NO: 38), 5'-AGCAGTCTCCATGTCCCAGG-3' (SEQ ID NO: 31), 5'-GGGUCTCCTCCACACCCUUC-3' (SEQ ID NO: 36), 5'-GGUGGCCACAGGCAACGUCA-3' (SEQ ID NO: 28), 5'-GCCGUTTCCATGTCCCAGGC-3' (SEQ ID NO: 46), 5'-GCGGUATCCATGTCCCAGGC-3' (SEQ ID NO: 42), 5'-GCGGUATACAGGTCCCAGGC-3' (SEQ ID NO: 43), 5'-GCUGUTTCCATGTCCCAGGC-3' (SEQ ID NO: 44), 5'-GCUGUGTCCATGTCCCAGGC-3' (SEQ ID NO: 45), 5'-GCCGUGTCCATGTCC CAGGC-3' (SEQ ID NO: 47), 5'-GCGGUAUCCAUGUCCCAGGC-3' (SEQ ID NO: 151), 5'-GGUATCCCCCCCCCCCCCCC-3' (SEQ ID NO: 54), 5'-GUCCCATCCCTTCTGCUGCC-3' (SEQ ID NO: 145), 5'-UUCUCTCTGGTCCCAUCCCU-3' (SEQ ID NO: 213), 5'-GUUCAGTCAGATCGCUGGGA-3' (SEQ ID NO: 214), 5'-AUGACATTTCGTGGCUCCUA-3' (SEQ ID NO: 215), 5'-UCUCCATGTCCCAGGCCUCC-3' (SEQ ID NO: 216), 5'-AGUCUCCATGTCCCAGGCCU-3' (SEQ ID NO: 217), or a variant thereof having at least about 75% sequence identity thereto (U can be T and / or T can be U).

[0307] In particular embodiments of the above five aspects, the oligonucleotide comprises, consists of, or consists essentially of a motif selected from 5'-GGUAUA-3' (SEQ ID NO:4), 5'-GGUAU-3' (SEQ ID NO:56), 5'-GGUA-3' (SEQ ID NO:57), 5'-GUAU-3' (SEQ ID NO:58), 5'-GUA-3' (SEQ ID NO:60), and 5'-GGU-3' (SEQ ID NO:59), where U can be T. In such instances, the motif is preferably at or towards the 5' end of the oligonucleotide.

[0308] Suitably for the above embodiments, one or more of the bases of the motif are modified bases and / or have modified backbones such as those described herein.

[0309] Examples of modified bases useful in the present invention include, but are not limited to, those containing 2'-O-methyl, 2'-O-methoxyethoxy, 2'-fluoro, 2'-allyl, 2'-O-[2-(methylamino)-2-oxoethyl], 4'-thio, 4'-CH2-O-2'-bridged, 4'-(CH2)2-O-2'-bridged, 2'-LNA, 2'-amino, fluoroarabinonucleotide, threose nucleic acid, or 2'-O--(N-methylcarbamate).

[0310] With reference to the above embodiment, the modified backbone preferably comprises a phosphorothioate, a non-bridging oxygen atom replacing a sulfur atom, a phosphonate such as a methylphosphonate, a phosphodiester, a phosphoromorpholidate, a phosphoropiperazidate, an amide, a methylene (methylamino), a formacetal, a thioformacetal, a peptide nucleic acid or a phosphoramidate such as a morpholinophosphorodiamidate (PMO), an N3'-P5' phosphoramidite or a thiophosphoramidite.

[0311] With respect to the above embodiments, at least a portion of the oligonucleotide is preferably a ribonucleic acid, a deoxyribonucleic acid, a DNA phosphorothioate, an RNA phosphorothioate, a 2'-O-methyl-oligonucleotide, a 2'-O-methyl-oligodeoxyribonucleotide, a 2'-O-hydrocarbyl ribonucleic acid, a 2'-O-hydrocarbyl DNA, a 2'-O-hydrocarbyl RNA phosphorothioate, a 2'-O-hydrocarbyl DNA phosphorothioate, a 2'-F-phosphorothioate, a 2'-F-phosphodiester, a 2'-methoxyethyl phosphorothioate, a 2-methoxyethyl phosphodiester. The nucleic acid may comprise / be any of a 3'-amino acid, a 2'-amino acid, ...

[0312] In one embodiment of the above aspect, the modified base is (a) 2'O-methyl and phosphorothioate backbones, (b) 2'-LNA and a phosphorothioate backbone, or (c) Containing 2'-O-methoxyethoxy and phosphorothioate backbones.

[0313] In one embodiment of the above aspect, at least one of the bases of the oligonucleotide does not hybridize to the target polynucleotide.

[0314] Preferably, the oligonucleotide of the above aspect is an antisense oligonucleotide, such as a gapmer antisense oligonucleotide, or a double-stranded oligonucleotide for gene silencing, such as an siRNA or shRNA. In certain embodiments, one or more bases of the motif or oligonucleotide are removed in vivo by an endonuclease.

[0315] In another embodiment, the oligonucleotide of the invention is a synthetic oligonucleotide. In this regard, the oligonucleotide is designed so that it does not bind to or hybridize to a target polynucleotide, such as a cellular or naturally occurring transcript.

[0316] Any embodiment herein shall be construed to apply mutatis mutandis to any other embodiment unless specifically stated otherwise.

[0317] The present invention is not to be limited in scope by the specific embodiments described herein, which are intended for the purpose of illustration only. Functionally equivalent products, compositions and methods are clearly within the scope of the invention as described herein.

[0318] Throughout this specification, unless specifically stated otherwise or the context requires otherwise, references to a single step, composition of matter, group of steps, or composition of matter shall be interpreted as encompassing one and more (i.e., one or more) of that step, composition of matter, group of steps, or composition of matter.

[0319] The invention will now be described by way of the following non-limiting examples and with reference to the accompanying drawings. [Brief explanation of the drawings]

[0320] [Figure 1A]Sequence-dependent inhibition of cGAS sensing by ASOs. (A) HeLa and HT-29 cells were transfected with 20 nM of the indicated ASOs targeting cGAS (Table 1) for 24 h before RNA purification and RT-qPCR analysis. cGAS levels are reported relative to 18S and normalized to mock conditions. Data shown represent the median of two independent experiments for each cell line. (B) THP-1 cells pretreated overnight with 100 nM of the indicated ASOs were transfected with 2.5 μg / ml ISD70 or not transfected (non-treated [NT]) for 8.5 h, and IP-10 levels in the supernatants were determined by ELISA. Data shown are averages from two independent experiments with biological triplicates (±s.e.m. and ordinary one-way ANOVA with Tukey's multiple comparison test for the "ISD70 only" condition or other indicated set of conditions). There was no basal effect of ASOs on NT cells (Alharbi et al., 2020). (C) HT-29 cells pretreated overnight with 125, 250, or 500 nM of the indicated ASOs were transfected with 2.5 μg / ml ISD70 for 24 hours or were not transfected (non-treated [NT]), and IP-10 levels in the supernatants were determined by ELISA. IP-10 levels were normalized to the "ISD70 only" condition after background correction for the NT condition. Data shown are averages from two independent experiments with biological triplicates (±s.e.m. and Mann-Whitney U test for the "ISD70 only" condition). (D, E) THP-1 cells pretreated overnight with 100 nM of the indicated ASOs were transfected with 2.5 μg / ml ISD70 for 7–8 hours, and IP-10 levels in the supernatants were determined by ELISA. (D) Stimulation and ELISA were performed on two independent plates, and the results are shown on each axis (with a correlation r = 0.7716, P < 0.0001). Average values ​​from both plates are shown in Table 2. The ISD70-only condition is shown in blue. (E) IP-10 levels were normalized to the "ISD70-only" condition after background correction with the NT condition.Data shown are averages from three independent experiments with biological triplicates (±sem and ordinary one-way ANOVA with Tukey's multiple comparison test for the condition "ISD70 only" or other indicated set of conditions). (F) HT-29 cells pretreated overnight with 187.5 nM of the indicated ASO were transfected with or without 2.5 μg / ml ISD70 for 24 hours, and IP-10 levels in the supernatants were determined by ELISA. IP-10 levels were normalized to the "ISD70 only" condition after background correction with the NT condition. Data shown are averages from three independent experiments with biological triplicates (±sem and ordinary one-way ANOVA with Dunnett's multiple comparison test for the condition "ISD70 only"). G) cGAS- / -, UNC93B1- / -, and matched control (UNC93B1WT) THP-1 cells with rescued UNC93B1 expression were pretreated with 100 or 250 nM ASO for 6 hours and transfected overnight with 2.5 μg / ml ISD70. GSK (100 nM) and ODN2006 (500 nM) were used as human STING and TLR9 agonists, respectively. IP-10 levels in the supernatants were determined by ELISA and normalized to the "GSK" condition after background correction with the NT condition. Data shown are averages from three independent experiments with biological triplicates (±s.e.m. and ordinary two-way ANOVA with Tukey's multiple comparison test against the "ISD70 only" condition). *P ≤ 0.05, **P ≤ 0.01, ***P ≤ 0.001, ****P ≤ 0.0001; ns: not significant. [Figure 1B]Sequence-dependent inhibition of cGAS sensing by ASOs. (A) HeLa and HT-29 cells were transfected with 20 nM of the indicated ASOs targeting cGAS (Table 1) for 24 h before RNA purification and RT-qPCR analysis. cGAS levels are reported relative to 18S and normalized to mock conditions. Data shown represent the median of two independent experiments for each cell line. (B) THP-1 cells pretreated overnight with 100 nM of the indicated ASOs were transfected with 2.5 μg / ml ISD70 or not transfected (non-treated [NT]) for 8.5 h, and IP-10 levels in the supernatants were determined by ELISA. Data shown are averages from two independent experiments with biological triplicates (±s.e.m. and ordinary one-way ANOVA with Tukey's multiple comparison test for the "ISD70 only" condition or other indicated set of conditions). There was no basal effect of ASOs on NT cells (Alharbi et al., 2020). (C) HT-29 cells pretreated overnight with 125, 250, or 500 nM of the indicated ASOs were transfected with 2.5 μg / ml ISD70 for 24 hours or were not transfected (non-treated [NT]), and IP-10 levels in the supernatants were determined by ELISA. IP-10 levels were normalized to the "ISD70 only" condition after background correction for the NT condition. Data shown are averages from two independent experiments with biological triplicates (±s.e.m. and Mann-Whitney U test for the "ISD70 only" condition). (D, E) THP-1 cells pretreated overnight with 100 nM of the indicated ASOs were transfected with 2.5 μg / ml ISD70 for 7–8 hours, and IP-10 levels in the supernatants were determined by ELISA. (D) Stimulation and ELISA were performed on two independent plates, and the results are shown on each axis (with a correlation r = 0.7716, P < 0.0001). Average values ​​from both plates are shown in Table 2. The ISD70-only condition is shown in blue. (E) IP-10 levels were normalized to the "ISD70-only" condition after background correction with the NT condition.Data shown are averages from three independent experiments with biological triplicates (±sem and ordinary one-way ANOVA with Tukey's multiple comparison test for the condition "ISD70 only" or other indicated set of conditions). (F) HT-29 cells pretreated overnight with 187.5 nM of the indicated ASO were transfected with or without 2.5 μg / ml ISD70 for 24 hours, and IP-10 levels in the supernatants were determined by ELISA. IP-10 levels were normalized to the "ISD70 only" condition after background correction with the NT condition. Data shown are averages from three independent experiments with biological triplicates (±sem and ordinary one-way ANOVA with Dunnett's multiple comparison test for the condition "ISD70 only"). G) cGAS- / -, UNC93B1- / -, and matched control (UNC93B1WT) THP-1 cells with rescued UNC93B1 expression were pretreated with 100 or 250 nM ASO for 6 hours and transfected overnight with 2.5 μg / ml ISD70. GSK (100 nM) and ODN2006 (500 nM) were used as human STING and TLR9 agonists, respectively. IP-10 levels in the supernatants were determined by ELISA and normalized to the "GSK" condition after background correction with the NT condition. Data shown are averages from three independent experiments with biological triplicates (±s.e.m. and ordinary two-way ANOVA with Tukey's multiple comparison test against the "ISD70 only" condition). *P ≤ 0.05, **P ≤ 0.01, ***P ≤ 0.001, ****P ≤ 0.0001; ns: not significant. [Figure 1C]Sequence-dependent inhibition of cGAS sensing by ASOs. (A) HeLa and HT-29 cells were transfected with 20 nM of the indicated ASOs targeting cGAS (Table 1) for 24 h before RNA purification and RT-qPCR analysis. cGAS levels are reported relative to 18S and normalized to mock conditions. Data shown represent the median of two independent experiments for each cell line. (B) THP-1 cells pretreated overnight with 100 nM of the indicated ASOs were transfected with 2.5 μg / ml ISD70 or not transfected (non-treated [NT]) for 8.5 h, and IP-10 levels in the supernatants were determined by ELISA. Data shown are averages from two independent experiments with biological triplicates (±s.e.m. and ordinary one-way ANOVA with Tukey's multiple comparison test for the "ISD70 only" condition or other indicated set of conditions). There was no basal effect of ASOs on NT cells (Alharbi et al., 2020). (C) HT-29 cells pretreated overnight with 125, 250, or 500 nM of the indicated ASOs were transfected with 2.5 μg / ml ISD70 for 24 hours or were not transfected (non-treated [NT]), and IP-10 levels in the supernatants were determined by ELISA. IP-10 levels were normalized to the "ISD70 only" condition after background correction for the NT condition. Data shown are averages from two independent experiments with biological triplicates (±s.e.m. and Mann-Whitney U test for the "ISD70 only" condition). (D, E) THP-1 cells pretreated overnight with 100 nM of the indicated ASOs were transfected with 2.5 μg / ml ISD70 for 7–8 hours, and IP-10 levels in the supernatants were determined by ELISA. (D) Stimulation and ELISA were performed on two independent plates, and the results are shown on each axis (with a correlation r = 0.7716, P < 0.0001). Average values ​​from both plates are shown in Table 2. The ISD70-only condition is shown in blue. (E) IP-10 levels were normalized to the "ISD70-only" condition after background correction with the NT condition.Data shown are averages from three independent experiments with biological triplicates (±sem and ordinary one-way ANOVA with Tukey's multiple comparison test for the condition "ISD70 only" or other indicated set of conditions). (F) HT-29 cells pretreated overnight with 187.5 nM of the indicated ASO were transfected with or without 2.5 μg / ml ISD70 for 24 hours, and IP-10 levels in the supernatants were determined by ELISA. IP-10 levels were normalized to the "ISD70 only" condition after background correction with the NT condition. Data shown are averages from three independent experiments with biological triplicates (±sem and ordinary one-way ANOVA with Dunnett's multiple comparison test for the condition "ISD70 only"). G) cGAS- / -, UNC93B1- / -, and matched control (UNC93B1WT) THP-1 cells with rescued UNC93B1 expression were pretreated with 100 or 250 nM ASO for 6 hours and transfected overnight with 2.5 μg / ml ISD70. GSK (100 nM) and ODN2006 (500 nM) were used as human STING and TLR9 agonists, respectively. IP-10 levels in the supernatants were determined by ELISA and normalized to the "GSK" condition after background correction with the NT condition. Data shown are averages from three independent experiments with biological triplicates (±s.e.m. and ordinary two-way ANOVA with Tukey's multiple comparison test against the "ISD70 only" condition). *P ≤ 0.05, **P ≤ 0.01, ***P ≤ 0.001, ****P ≤ 0.0001; ns: not significant. [Figure 1D]Sequence-dependent inhibition of cGAS sensing by ASOs. (A) HeLa and HT-29 cells were transfected with 20 nM of the indicated ASOs targeting cGAS (Table 1) for 24 h before RNA purification and RT-qPCR analysis. cGAS levels are reported relative to 18S and normalized to mock conditions. Data shown represent the median of two independent experiments for each cell line. (B) THP-1 cells pretreated overnight with 100 nM of the indicated ASOs were transfected with 2.5 μg / ml ISD70 or not transfected (non-treated [NT]) for 8.5 h, and IP-10 levels in the supernatants were determined by ELISA. Data shown are averages from two independent experiments with biological triplicates (±s.e.m. and ordinary one-way ANOVA with Tukey's multiple comparison test for the "ISD70 only" condition or other indicated set of conditions). There was no basal effect of ASOs on NT cells (Alharbi et al., 2020). (C) HT-29 cells pretreated overnight with 125, 250, or 500 nM of the indicated ASOs were transfected with 2.5 μg / ml ISD70 for 24 hours or were not transfected (non-treated [NT]), and IP-10 levels in the supernatants were determined by ELISA. IP-10 levels were normalized to the "ISD70 only" condition after background correction for the NT condition. Data shown are averages from two independent experiments with biological triplicates (±s.e.m. and Mann-Whitney U test for the "ISD70 only" condition). (D, E) THP-1 cells pretreated overnight with 100 nM of the indicated ASOs were transfected with 2.5 μg / ml ISD70 for 7–8 hours, and IP-10 levels in the supernatants were determined by ELISA. (D) Stimulation and ELISA were performed on two independent plates, and the results are shown on each axis (with a correlation r = 0.7716, P < 0.0001). Average values ​​from both plates are shown in Table 2. The ISD70-only condition is shown in blue. (E) IP-10 levels were normalized to the "ISD70-only" condition after background correction with the NT condition.Data shown are averages from three independent experiments with biological triplicates (±sem and ordinary one-way ANOVA with Tukey's multiple comparison test for the condition "ISD70 only" or other indicated set of conditions). (F) HT-29 cells pretreated overnight with 187.5 nM of the indicated ASO were transfected with or without 2.5 μg / ml ISD70 for 24 hours, and IP-10 levels in the supernatants were determined by ELISA. IP-10 levels were normalized to the "ISD70 only" condition after background correction with the NT condition. Data shown are averages from three independent experiments with biological triplicates (±sem and ordinary one-way ANOVA with Dunnett's multiple comparison test for the condition "ISD70 only"). G) cGAS- / -, UNC93B1- / -, and matched control (UNC93B1WT) THP-1 cells with rescued UNC93B1 expression were pretreated with 100 or 250 nM ASO for 6 hours and transfected overnight with 2.5 μg / ml ISD70. GSK (100 nM) and ODN2006 (500 nM) were used as human STING and TLR9 agonists, respectively. IP-10 levels in the supernatants were determined by ELISA and normalized to the "GSK" condition after background correction with the NT condition. Data shown are averages from three independent experiments with biological triplicates (±s.e.m. and ordinary two-way ANOVA with Tukey's multiple comparison test against the "ISD70 only" condition). *P ≤ 0.05, **P ≤ 0.01, ***P ≤ 0.001, ****P ≤ 0.0001; ns: not significant. [Figure 1E]Sequence-dependent inhibition of cGAS sensing by ASOs. (A) HeLa and HT-29 cells were transfected with 20 nM of the indicated ASOs targeting cGAS (Table 1) for 24 h before RNA purification and RT-qPCR analysis. cGAS levels are reported relative to 18S and normalized to mock conditions. Data shown represent the median of two independent experiments for each cell line. (B) THP-1 cells pretreated overnight with 100 nM of the indicated ASOs were transfected with 2.5 μg / ml ISD70 or not transfected (non-treated [NT]) for 8.5 h, and IP-10 levels in the supernatants were determined by ELISA. Data shown are averages from two independent experiments with biological triplicates (±s.e.m. and ordinary one-way ANOVA with Tukey's multiple comparison test for the "ISD70 only" condition or other indicated set of conditions). There was no basal effect of ASOs on NT cells (Alharbi et al., 2020). (C) HT-29 cells pretreated overnight with 125, 250, or 500 nM of the indicated ASOs were transfected with 2.5 μg / ml ISD70 for 24 hours or were not transfected (non-treated [NT]), and IP-10 levels in the supernatants were determined by ELISA. IP-10 levels were normalized to the "ISD70 only" condition after background correction for the NT condition. Data shown are averages from two independent experiments with biological triplicates (±s.e.m. and Mann-Whitney U test for the "ISD70 only" condition). (D, E) THP-1 cells pretreated overnight with 100 nM of the indicated ASOs were transfected with 2.5 μg / ml ISD70 for 7–8 hours, and IP-10 levels in the supernatants were determined by ELISA. (D) Stimulation and ELISA were performed on two independent plates, and the results are shown on each axis (with a correlation r = 0.7716, P < 0.0001). Average values ​​from both plates are shown in Table 2. The ISD70-only condition is shown in blue. (E) IP-10 levels were normalized to the "ISD70-only" condition after background correction with the NT condition.Data shown are averages from three independent experiments with biological triplicates (±sem and ordinary one-way ANOVA with Tukey's multiple comparison test for the condition "ISD70 only" or other indicated set of conditions). (F) HT-29 cells pretreated overnight with 187.5 nM of the indicated ASO were transfected with or without 2.5 μg / ml ISD70 for 24 hours, and IP-10 levels in the supernatants were determined by ELISA. IP-10 levels were normalized to the "ISD70 only" condition after background correction with the NT condition. Data shown are averages from three independent experiments with biological triplicates (±sem and ordinary one-way ANOVA with Dunnett's multiple comparison test for the condition "ISD70 only"). G) cGAS- / -, UNC93B1- / -, and matched control (UNC93B1WT) THP-1 cells with rescued UNC93B1 expression were pretreated with 100 or 250 nM ASO for 6 hours and transfected overnight with 2.5 μg / ml ISD70. GSK (100 nM) and ODN2006 (500 nM) were used as human STING and TLR9 agonists, respectively. IP-10 levels in the supernatants were determined by ELISA and normalized to the "GSK" condition after background correction with the NT condition. Data shown are averages from three independent experiments with biological triplicates (±s.e.m. and ordinary two-way ANOVA with Tukey's multiple comparison test against the "ISD70 only" condition). *P ≤ 0.05, **P ≤ 0.01, ***P ≤ 0.001, ****P ≤ 0.0001; ns: not significant. [Figure 1F]Sequence-dependent inhibition of cGAS sensing by ASOs. (A) HeLa and HT-29 cells were transfected with 20 nM of the indicated ASOs targeting cGAS (Table 1) for 24 h before RNA purification and RT-qPCR analysis. cGAS levels are reported relative to 18S and normalized to mock conditions. Data shown represent the median of two independent experiments for each cell line. (B) THP-1 cells pretreated overnight with 100 nM of the indicated ASOs were transfected with 2.5 μg / ml ISD70 or not transfected (non-treated [NT]) for 8.5 h, and IP-10 levels in the supernatants were determined by ELISA. Data shown are averages from two independent experiments with biological triplicates (±s.e.m. and ordinary one-way ANOVA with Tukey's multiple comparison test for the "ISD70 only" condition or other indicated set of conditions). There was no basal effect of ASOs on NT cells (Alharbi et al., 2020). (C) HT-29 cells pretreated overnight with 125, 250, or 500 nM of the indicated ASOs were transfected with 2.5 μg / ml ISD70 for 24 hours or were not transfected (non-treated [NT]), and IP-10 levels in the supernatants were determined by ELISA. IP-10 levels were normalized to the "ISD70 only" condition after background correction for the NT condition. Data shown are averages from two independent experiments with biological triplicates (±s.e.m. and Mann-Whitney U test for the "ISD70 only" condition). (D, E) THP-1 cells pretreated overnight with 100 nM of the indicated ASOs were transfected with 2.5 μg / ml ISD70 for 7–8 hours, and IP-10 levels in the supernatants were determined by ELISA. (D) Stimulation and ELISA were performed on two independent plates, and the results are shown on each axis (with a correlation r = 0.7716, P < 0.0001). Average values ​​from both plates are shown in Table 2. The ISD70-only condition is shown in blue. (E) IP-10 levels were normalized to the "ISD70-only" condition after background correction with the NT condition.Data shown are averages from three independent experiments with biological triplicates (±sem and ordinary one-way ANOVA with Tukey's multiple comparison test for the condition "ISD70 only" or other indicated set of conditions). (F) HT-29 cells pretreated overnight with 187.5 nM of the indicated ASO were transfected with or without 2.5 μg / ml ISD70 for 24 hours, and IP-10 levels in the supernatants were determined by ELISA. IP-10 levels were normalized to the "ISD70 only" condition after background correction with the NT condition. Data shown are averages from three independent experiments with biological triplicates (±sem and ordinary one-way ANOVA with Dunnett's multiple comparison test for the condition "ISD70 only"). G) cGAS- / -, UNC93B1- / -, and matched control (UNC93B1WT) THP-1 cells with rescued UNC93B1 expression were pretreated with 100 or 250 nM ASO for 6 hours and transfected overnight with 2.5 μg / ml ISD70. GSK (100 nM) and ODN2006 (500 nM) were used as human STING and TLR9 agonists, respectively. IP-10 levels in the supernatants were determined by ELISA and normalized to the "GSK" condition after background correction with the NT condition. Data shown are averages from three independent experiments with biological triplicates (±s.e.m. and ordinary two-way ANOVA with Tukey's multiple comparison test against the "ISD70 only" condition). *P ≤ 0.05, **P ≤ 0.01, ***P ≤ 0.001, ****P ≤ 0.0001; ns: not significant. [Figure 1G]Sequence-dependent inhibition of cGAS sensing by ASOs. (A) HeLa and HT-29 cells were transfected with 20 nM of the indicated ASOs targeting cGAS (Table 1) for 24 h before RNA purification and RT-qPCR analysis. cGAS levels are reported relative to 18S and normalized to mock conditions. Data shown represent the median of two independent experiments for each cell line. (B) THP-1 cells pretreated overnight with 100 nM of the indicated ASOs were transfected with 2.5 μg / ml ISD70 or not transfected (non-treated [NT]) for 8.5 h, and IP-10 levels in the supernatants were determined by ELISA. Data shown are averages from two independent experiments with biological triplicates (±s.e.m. and ordinary one-way ANOVA with Tukey's multiple comparison test for the "ISD70 only" condition or other indicated set of conditions). There was no basal effect of ASOs on NT cells (Alharbi et al., 2020). (C) HT-29 cells pretreated overnight with 125, 250, or 500 nM of the indicated ASOs were transfected with 2.5 μg / ml ISD70 for 24 hours or were not transfected (non-treated [NT]), and IP-10 levels in the supernatants were determined by ELISA. IP-10 levels were normalized to the "ISD70 only" condition after background correction for the NT condition. Data shown are averages from two independent experiments with biological triplicates (±s.e.m. and Mann-Whitney U test for the "ISD70 only" condition). (D, E) THP-1 cells pretreated overnight with 100 nM of the indicated ASOs were transfected with 2.5 μg / ml ISD70 for 7–8 hours, and IP-10 levels in the supernatants were determined by ELISA. (D) Stimulation and ELISA were performed on two independent plates, and the results are shown on each axis (with a correlation r = 0.7716, P < 0.0001). Average values ​​from both plates are shown in Table 2. The ISD70-only condition is shown in blue. (E) IP-10 levels were normalized to the "ISD70-only" condition after background correction with the NT condition.Data shown are averages from three independent experiments with biological triplicates (±sem and ordinary one-way ANOVA with Tukey's multiple comparison test for the condition "ISD70 only" or other indicated set of conditions). (F) HT-29 cells pretreated overnight with 187.5 nM of the indicated ASO were transfected with or without 2.5 μg / ml ISD70 for 24 hours, and IP-10 levels in the supernatants were determined by ELISA. IP-10 levels were normalized to the "ISD70 only" condition after background correction with the NT condition. Data shown are averages from three independent experiments with biological triplicates (±sem and ordinary one-way ANOVA with Dunnett's multiple comparison test for the condition "ISD70 only"). G) cGAS- / -, UNC93B1- / -, and matched control (UNC93B1WT) THP-1 cells with rescued UNC93B1 expression were pretreated with 100 or 250 nM ASO for 6 hours and transfected overnight with 2.5 μg / ml ISD70. GSK (100 nM) and ODN2006 (500 nM) were used as human STING and TLR9 agonists, respectively. IP-10 levels in the supernatants were determined by ELISA and normalized to the "GSK" condition after background correction with the NT condition. Data shown are averages from three independent experiments with biological triplicates (±s.e.m. and ordinary two-way ANOVA with Tukey's multiple comparison test against the "ISD70 only" condition). *P ≤ 0.05, **P ≤ 0.01, ***P ≤ 0.001, ****P ≤ 0.0001; ns: not significant.

[0321] [Figure 2A]Identification of highly potent inhibitors of cGAS sensing. (A) Top: Sequence alignment of cGAS inhibitors and identification of key bases predicted by MEME analysis highlighted by arrows (Figure 6A). Bottom: Design of C2 and ASO2 mutants incorporating point mutations at selected positions in the MEME motif (mutations highlighted in yellow). (B, D) HT-29 cells pretreated overnight with the indicated doses of ASO (500, 250, 125, or 62.5 nM) were transfected with or without 2.5 μg / ml ISD70 for 24 hours, and IP-10 levels in the supernatants were determined by ELISA. IP-10 levels were normalized to the "ISD70 only" condition after background correction with the NT condition. Data shown are averages from two (D) or three (B) independent experiments in biological triplicates (±s.e.m. and ordinary two-way ANOVA with Tukey's multiple comparison test for C2 [B] or ASO2 [D] conditions; comparisons were not significant between 250 and 500 nM ASO). (C) Mouse LL171 cells were treated with the indicated amounts of ASO (200, 400, or 600 nM) for 6 hours and then stimulated overnight with 2.5 μg / ml ISD45. The next day, cells were lysed, and ISRE-Luc levels were analyzed by luciferase assay. ISRE-luciferase levels were normalized to the "ISD45 only" condition after background correction with the NT condition. Data shown are averages from two independent experiments in biological triplicates (±s.e.m. and Mann-Whitney U test). (E) Top: Sequence alignment of C2-Mut1 mutants, C2-ASO2-A and C2-ASO2-B, with the 3' ends from ASO2 up (ASO2up) and ASO2 down (ASO2down) underlined. Bottom: Homopolymer dC20 mutants, with the 2'OMe bases in pink and the cGAS inhibitory motif underlined. (F) HT-29 cells pretreated overnight with 187.5 nM of the indicated ASO were transfected or not with 2.5 μg / ml ISD70 for 24 h, and IP-10 levels in the supernatants were determined by ELISA.(G) THP-1 cells pretreated overnight with 100 nM of the indicated ASO were transfected with 2.5 μg / ml of ISD70 for 24 hours, and IP-10 levels in the supernatants were determined by ELISA. (F, G) IP-10 levels were normalized to the "ISD70 only" condition after background correction with NT conditions. Data shown are averages from three independent experiments with biological triplicates (±s.e.m. and ordinary one-way ANOVA with Tukey's multiple comparison test for the condition "ISD70 only" or other indicated pair of conditions). (H) LL171 cells were treated with 200 nM of ASO for 6 hours and then stimulated with 2.5 μg / ml of ISD45 overnight. The next day, cells were lysed, and ISRE-Luc levels were analyzed by luciferase assay. ISRE-luciferase levels were normalized to the "ISD45 only" condition after background correction with NT conditions. Data shown are averages from three independent experiments with biological triplicates (±s.e.m. and ordinary one-way ANOVA with Dunnett's multiple comparison test against the "ISD70 only" condition). (I) THP-1 cells were treated with 100 nM C2-Mut1 for 6 h or transfected with 100, 250, or 500 nM C2-Mut1-PS and 2.5 μg / ml ISD70 overnight. IP-10 levels were normalized to the "ISD70 only" condition after background correction with the NT condition. Data shown are averages from two independent experiments with biological triplicates (±s.e.m. and Mann-Whitney U test against the "ISD70 only" condition). (J) Sequences of [LINC-PINT] ASO101–116, showing the location of the inhibitory GGUCCC motif (pink) from ASO103 identified by MEME (see D and Figure 6B). Yellow regions highlight the gapmer DNA portion. (K) THP-1 cells pretreated overnight with 100 nM of the indicated [LINC-PINT] ASO were transfected with 2.5 μg / ml ISD70 for 7.5 hours, and IP-10 levels in the supernatants were determined by ELISA. IP-10 levels were normalized to the "ISD70 only" condition after background correction with the NT condition.Data shown are averages from three independent experiments with biological triplicates (±s.e.m. and one-way ANOVA with Dunnett's multiple comparison test against the "ISD70 only" condition). (L, M) LL171 cells were treated with 200 nM ASO for 6 h or 20 min, then "washed" or not (L), or treated with 50, 100, or 200 nM ASO for 20 min (M), and stimulated overnight with 2.5 μg / ml ISD45. The next day, cells were lysed, and ISRE-Luc levels were analyzed by luciferase assay. ISRE-luciferase levels were normalized to the "ISD45 only" condition after background correction with the NT condition. Data shown are averages from two (L) or three (M) independent experiments with biological triplicates (±s.e.m. and one-way ANOVA with Tukey's multiple comparison test (L) for the "ISD45 only" condition, or Mann-Whitney U test (M) for "ISD45 only," or other indicated pair of conditions). (N) THP-1 cells pretreated with 250 nM of the indicated ASO for 40 min were transfected with 2.5 μg / ml ISD70 overnight, and IP-10 levels in the supernatants were determined by ELISA. Data shown are averages from three independent experiments with biological triplicates (±s.e.m. and one-way ANOVA with Dunnett's multiple comparison test for the "ISD70 only" condition, and Mann-Whitney U test comparing Mut1-dC and dC20). *P ≤ 0.05, **P ≤ 0.01, ***P ≤ 0.001, ****P ≤ 0.0001, ns: not significant. [Figure 2B]Identification of highly potent inhibitors of cGAS sensing. (A) Top: Sequence alignment of cGAS inhibitors and identification of key bases predicted by MEME analysis highlighted by arrows (Figure 6A). Bottom: Design of C2 and ASO2 mutants incorporating point mutations at selected positions in the MEME motif (mutations highlighted in yellow). (B, D) HT-29 cells pretreated overnight with the indicated doses of ASO (500, 250, 125, or 62.5 nM) were transfected with or without 2.5 μg / ml ISD70 for 24 hours, and IP-10 levels in the supernatants were determined by ELISA. IP-10 levels were normalized to the "ISD70 only" condition after background correction with the NT condition. Data shown are averages from two (D) or three (B) independent experiments in biological triplicates (±s.e.m. and ordinary two-way ANOVA with Tukey's multiple comparison test for C2 [B] or ASO2 [D] conditions; comparisons were not significant between 250 and 500 nM ASO). (C) Mouse LL171 cells were treated with the indicated amounts of ASO (200, 400, or 600 nM) for 6 hours and then stimulated overnight with 2.5 μg / ml ISD45. The next day, cells were lysed, and ISRE-Luc levels were analyzed by luciferase assay. ISRE-luciferase levels were normalized to the "ISD45 only" condition after background correction with the NT condition. Data shown are averages from two independent experiments in biological triplicates (±s.e.m. and Mann-Whitney U test). (E) Top: Sequence alignment of C2-Mut1 mutants, C2-ASO2-A and C2-ASO2-B, with the 3' ends from ASO2 up (ASO2up) and ASO2 down (ASO2down) underlined. Bottom: Homopolymer dC20 mutants, with the 2'OMe bases in pink and the cGAS inhibitory motif underlined. (F) HT-29 cells pretreated overnight with 187.5 nM of the indicated ASO were transfected or not with 2.5 μg / ml ISD70 for 24 h, and IP-10 levels in the supernatants were determined by ELISA.(G) THP-1 cells pretreated overnight with 100 nM of the indicated ASO were transfected with 2.5 μg / ml of ISD70 for 24 hours, and IP-10 levels in the supernatants were determined by ELISA. (F, G) IP-10 levels were normalized to the "ISD70 only" condition after background correction with NT conditions. Data shown are averages from three independent experiments with biological triplicates (±s.e.m. and ordinary one-way ANOVA with Tukey's multiple comparison test for the condition "ISD70 only" or other indicated pair of conditions). (H) LL171 cells were treated with 200 nM of ASO for 6 hours and then stimulated with 2.5 μg / ml of ISD45 overnight. The next day, cells were lysed, and ISRE-Luc levels were analyzed by luciferase assay. ISRE-luciferase levels were normalized to the "ISD45 only" condition after background correction with NT conditions. Data shown are averages from three independent experiments with biological triplicates (±s.e.m. and ordinary one-way ANOVA with Dunnett's multiple comparison test against the "ISD70 only" condition). (I) THP-1 cells were treated with 100 nM C2-Mut1 for 6 h or transfected with 100, 250, or 500 nM C2-Mut1-PS and 2.5 μg / ml ISD70 overnight. IP-10 levels were normalized to the "ISD70 only" condition after background correction with the NT condition. Data shown are averages from two independent experiments with biological triplicates (±s.e.m. and Mann-Whitney U test against the "ISD70 only" condition). (J) Sequences of [LINC-PINT] ASO101–116, showing the location of the inhibitory GGUCCC motif (pink) from ASO103 identified by MEME (see D and Figure 6B). Yellow regions highlight the gapmer DNA portion. (K) THP-1 cells pretreated overnight with 100 nM of the indicated [LINC-PINT] ASO were transfected with 2.5 μg / ml ISD70 for 7.5 hours, and IP-10 levels in the supernatants were determined by ELISA. IP-10 levels were normalized to the "ISD70 only" condition after background correction with the NT condition.Data shown are averages from three independent experiments with biological triplicates (±s.e.m. and one-way ANOVA with Dunnett's multiple comparison test against the "ISD70 only" condition). (L, M) LL171 cells were treated with 200 nM ASO for 6 h or 20 min, then "washed" or not (L), or treated with 50, 100, or 200 nM ASO for 20 min (M), and stimulated overnight with 2.5 μg / ml ISD45. The next day, cells were lysed, and ISRE-Luc levels were analyzed by luciferase assay. ISRE-luciferase levels were normalized to the "ISD45 only" condition after background correction with the NT condition. Data shown are averages from two (L) or three (M) independent experiments with biological triplicates (±s.e.m. and one-way ANOVA with Tukey's multiple comparison test (L) for the "ISD45 only" condition, or Mann-Whitney U test (M) for "ISD45 only," or other indicated pair of conditions). (N) THP-1 cells pretreated with 250 nM of the indicated ASO for 40 min were transfected with 2.5 μg / ml ISD70 overnight, and IP-10 levels in the supernatants were determined by ELISA. Data shown are averages from three independent experiments with biological triplicates (±s.e.m. and one-way ANOVA with Dunnett's multiple comparison test for the "ISD70 only" condition, and Mann-Whitney U test comparing Mut1-dC and dC20). *P ≤ 0.05, **P ≤ 0.01, ***P ≤ 0.001, ****P ≤ 0.0001, ns: not significant. [Figure 2C]Identification of highly potent inhibitors of cGAS sensing. (A) Top: Sequence alignment of cGAS inhibitors and identification of key bases predicted by MEME analysis highlighted by arrows (Figure 6A). Bottom: Design of C2 and ASO2 mutants incorporating point mutations at selected positions in the MEME motif (mutations highlighted in yellow). (B, D) HT-29 cells pretreated overnight with the indicated doses of ASO (500, 250, 125, or 62.5 nM) were transfected with or without 2.5 μg / ml ISD70 for 24 hours, and IP-10 levels in the supernatants were determined by ELISA. IP-10 levels were normalized to the "ISD70 only" condition after background correction with the NT condition. Data shown are averages from two (D) or three (B) independent experiments in biological triplicates (±s.e.m. and ordinary two-way ANOVA with Tukey's multiple comparison test for C2 [B] or ASO2 [D] conditions; comparisons were not significant between 250 and 500 nM ASO). (C) Mouse LL171 cells were treated with the indicated amounts of ASO (200, 400, or 600 nM) for 6 hours and then stimulated overnight with 2.5 μg / ml ISD45. The next day, cells were lysed, and ISRE-Luc levels were analyzed by luciferase assay. ISRE-luciferase levels were normalized to the "ISD45 only" condition after background correction with the NT condition. Data shown are averages from two independent experiments in biological triplicates (±s.e.m. and Mann-Whitney U test). (E) Top: Sequence alignment of C2-Mut1 mutants, C2-ASO2-A and C2-ASO2-B, with the 3' ends from ASO2 up (ASO2up) and ASO2 down (ASO2down) underlined. Bottom: Homopolymer dC20 mutants, with the 2'OMe bases in pink and the cGAS inhibitory motif underlined. (F) HT-29 cells pretreated overnight with 187.5 nM of the indicated ASO were transfected or not with 2.5 μg / ml ISD70 for 24 h, and IP-10 levels in the supernatants were determined by ELISA.(G) THP-1 cells pretreated overnight with 100 nM of the indicated ASO were transfected with 2.5 μg / ml of ISD70 for 24 hours, and IP-10 levels in the supernatants were determined by ELISA. (F, G) IP-10 levels were normalized to the "ISD70 only" condition after background correction with NT conditions. Data shown are averages from three independent experiments with biological triplicates (±s.e.m. and ordinary one-way ANOVA with Tukey's multiple comparison test for the condition "ISD70 only" or other indicated pair of conditions). (H) LL171 cells were treated with 200 nM of ASO for 6 hours and then stimulated with 2.5 μg / ml of ISD45 overnight. The next day, cells were lysed, and ISRE-Luc levels were analyzed by luciferase assay. ISRE-luciferase levels were normalized to the "ISD45 only" condition after background correction with NT conditions. Data shown are averages from three independent experiments with biological triplicates (±s.e.m. and ordinary one-way ANOVA with Dunnett's multiple comparison test against the "ISD70 only" condition). (I) THP-1 cells were treated with 100 nM C2-Mut1 for 6 h or transfected with 100, 250, or 500 nM C2-Mut1-PS and 2.5 μg / ml ISD70 overnight. IP-10 levels were normalized to the "ISD70 only" condition after background correction with the NT condition. Data shown are averages from two independent experiments with biological triplicates (±s.e.m. and Mann-Whitney U test against the "ISD70 only" condition). (J) Sequences of [LINC-PINT] ASO101–116, showing the location of the inhibitory GGUCCC motif (pink) from ASO103 identified by MEME (see D and Figure 6B). Yellow regions highlight the gapmer DNA portion. (K) THP-1 cells pretreated overnight with 100 nM of the indicated [LINC-PINT] ASO were transfected with 2.5 μg / ml ISD70 for 7.5 hours, and IP-10 levels in the supernatants were determined by ELISA. IP-10 levels were normalized to the "ISD70 only" condition after background correction with the NT condition.Data shown are averages from three independent experiments with biological triplicates (±s.e.m. and one-way ANOVA with Dunnett's multiple comparison test against the "ISD70 only" condition). (L, M) LL171 cells were treated with 200 nM ASO for 6 h or 20 min, then "washed" or not (L), or treated with 50, 100, or 200 nM ASO for 20 min (M), and stimulated overnight with 2.5 μg / ml ISD45. The next day, cells were lysed, and ISRE-Luc levels were analyzed by luciferase assay. ISRE-luciferase levels were normalized to the "ISD45 only" condition after background correction with the NT condition. Data shown are averages from two (L) or three (M) independent experiments with biological triplicates (±s.e.m. and one-way ANOVA with Tukey's multiple comparison test (L) for the "ISD45 only" condition, or Mann-Whitney U test (M) for "ISD45 only," or other indicated pair of conditions). (N) THP-1 cells pretreated with 250 nM of the indicated ASO for 40 min were transfected with 2.5 μg / ml ISD70 overnight, and IP-10 levels in the supernatants were determined by ELISA. Data shown are averages from three independent experiments with biological triplicates (±s.e.m. and one-way ANOVA with Dunnett's multiple comparison test for the "ISD70 only" condition, and Mann-Whitney U test comparing Mut1-dC and dC20). *P ≤ 0.05, **P ≤ 0.01, ***P ≤ 0.001, ****P ≤ 0.0001, ns: not significant. [Figure 2D]Identification of highly potent inhibitors of cGAS sensing. (A) Top: Sequence alignment of cGAS inhibitors and identification of key bases predicted by MEME analysis highlighted by arrows (Figure 6A). Bottom: Design of C2 and ASO2 mutants incorporating point mutations at selected positions in the MEME motif (mutations highlighted in yellow). (B, D) HT-29 cells pretreated overnight with the indicated doses of ASO (500, 250, 125, or 62.5 nM) were transfected with or without 2.5 μg / ml ISD70 for 24 hours, and IP-10 levels in the supernatants were determined by ELISA. IP-10 levels were normalized to the "ISD70 only" condition after background correction with the NT condition. Data shown are averages from two (D) or three (B) independent experiments in biological triplicates (±s.e.m. and ordinary two-way ANOVA with Tukey's multiple comparison test for C2 [B] or ASO2 [D] conditions; comparisons were not significant between 250 and 500 nM ASO). (C) Mouse LL171 cells were treated with the indicated amounts of ASO (200, 400, or 600 nM) for 6 hours and then stimulated overnight with 2.5 μg / ml ISD45. The next day, cells were lysed, and ISRE-Luc levels were analyzed by luciferase assay. ISRE-luciferase levels were normalized to the "ISD45 only" condition after background correction with the NT condition. Data shown are averages from two independent experiments in biological triplicates (±s.e.m. and Mann-Whitney U test). (E) Top: Sequence alignment of C2-Mut1 mutants, C2-ASO2-A and C2-ASO2-B, with the 3' ends from ASO2 up (ASO2up) and ASO2 down (ASO2down) underlined. Bottom: Homopolymer dC20 mutants, with the 2'OMe bases in pink and the cGAS inhibitory motif underlined. (F) HT-29 cells pretreated overnight with 187.5 nM of the indicated ASO were transfected or not with 2.5 μg / ml ISD70 for 24 h, and IP-10 levels in the supernatants were determined by ELISA.(G) THP-1 cells pretreated overnight with 100 nM of the indicated ASO were transfected with 2.5 μg / ml of ISD70 for 24 hours, and IP-10 levels in the supernatants were determined by ELISA. (F, G) IP-10 levels were normalized to the "ISD70 only" condition after background correction with NT conditions. Data shown are averages from three independent experiments with biological triplicates (±s.e.m. and ordinary one-way ANOVA with Tukey's multiple comparison test for the condition "ISD70 only" or other indicated pair of conditions). (H) LL171 cells were treated with 200 nM of ASO for 6 hours and then stimulated with 2.5 μg / ml of ISD45 overnight. The next day, cells were lysed, and ISRE-Luc levels were analyzed by luciferase assay. ISRE-luciferase levels were normalized to the "ISD45 only" condition after background correction with NT conditions. Data shown are averages from three independent experiments with biological triplicates (±s.e.m. and ordinary one-way ANOVA with Dunnett's multiple comparison test against the "ISD70 only" condition). (I) THP-1 cells were treated with 100 nM C2-Mut1 for 6 h or transfected with 100, 250, or 500 nM C2-Mut1-PS and 2.5 μg / ml ISD70 overnight. IP-10 levels were normalized to the "ISD70 only" condition after background correction with the NT condition. Data shown are averages from two independent experiments with biological triplicates (±s.e.m. and Mann-Whitney U test against the "ISD70 only" condition). (J) Sequences of [LINC-PINT] ASO101–116, showing the location of the inhibitory GGUCCC motif (pink) from ASO103 identified by MEME (see D and Figure 6B). Yellow regions highlight the gapmer DNA portion. (K) THP-1 cells pretreated overnight with 100 nM of the indicated [LINC-PINT] ASO were transfected with 2.5 μg / ml ISD70 for 7.5 hours, and IP-10 levels in the supernatants were determined by ELISA. IP-10 levels were normalized to the "ISD70 only" condition after background correction with the NT condition.Data shown are averages from three independent experiments with biological triplicates (±s.e.m. and one-way ANOVA with Dunnett's multiple comparison test against the "ISD70 only" condition). (L, M) LL171 cells were treated with 200 nM ASO for 6 h or 20 min, then "washed" or not (L), or treated with 50, 100, or 200 nM ASO for 20 min (M), and stimulated overnight with 2.5 μg / ml ISD45. The next day, cells were lysed, and ISRE-Luc levels were analyzed by luciferase assay. ISRE-luciferase levels were normalized to the "ISD45 only" condition after background correction with the NT condition. Data shown are averages from two (L) or three (M) independent experiments with biological triplicates (±s.e.m. and one-way ANOVA with Tukey's multiple comparison test (L) for the "ISD45 only" condition, or Mann-Whitney U test (M) for "ISD45 only," or other indicated pair of conditions). (N) THP-1 cells pretreated with 250 nM of the indicated ASO for 40 min were transfected with 2.5 μg / ml ISD70 overnight, and IP-10 levels in the supernatants were determined by ELISA. Data shown are averages from three independent experiments with biological triplicates (±s.e.m. and one-way ANOVA with Dunnett's multiple comparison test for the "ISD70 only" condition, and Mann-Whitney U test comparing Mut1-dC and dC20). *P ≤ 0.05, **P ≤ 0.01, ***P ≤ 0.001, ****P ≤ 0.0001, ns: not significant. [Figure 2E]Identification of highly potent inhibitors of cGAS sensing. (A) Top: Sequence alignment of cGAS inhibitors and identification of key bases predicted by MEME analysis highlighted by arrows (Figure 6A). Bottom: Design of C2 and ASO2 mutants incorporating point mutations at selected positions in the MEME motif (mutations highlighted in yellow). (B, D) HT-29 cells pretreated overnight with the indicated doses of ASO (500, 250, 125, or 62.5 nM) were transfected with or without 2.5 μg / ml ISD70 for 24 hours, and IP-10 levels in the supernatants were determined by ELISA. IP-10 levels were normalized to the "ISD70 only" condition after background correction with the NT condition. Data shown are averages from two (D) or three (B) independent experiments in biological triplicates (±s.e.m. and ordinary two-way ANOVA with Tukey's multiple comparison test for C2 [B] or ASO2 [D] conditions; comparisons were not significant between 250 and 500 nM ASO). (C) Mouse LL171 cells were treated with the indicated amounts of ASO (200, 400, or 600 nM) for 6 hours and then stimulated overnight with 2.5 μg / ml ISD45. The next day, cells were lysed, and ISRE-Luc levels were analyzed by luciferase assay. ISRE-luciferase levels were normalized to the "ISD45 only" condition after background correction with the NT condition. Data shown are averages from two independent experiments in biological triplicates (±s.e.m. and Mann-Whitney U test). (E) Top: Sequence alignment of C2-Mut1 mutants, C2-ASO2-A and C2-ASO2-B, with the 3' ends from ASO2 up (ASO2up) and ASO2 down (ASO2down) underlined. Bottom: Homopolymer dC20 mutants, with the 2'OMe bases in pink and the cGAS inhibitory motif underlined. (F) HT-29 cells pretreated overnight with 187.5 nM of the indicated ASO were transfected or not with 2.5 μg / ml ISD70 for 24 h, and IP-10 levels in the supernatants were determined by ELISA.(G) THP-1 cells pretreated overnight with 100 nM of the indicated ASO were transfected with 2.5 μg / ml of ISD70 for 24 hours, and IP-10 levels in the supernatants were determined by ELISA. (F, G) IP-10 levels were normalized to the "ISD70 only" condition after background correction with NT conditions. Data shown are averages from three independent experiments with biological triplicates (±s.e.m. and ordinary one-way ANOVA with Tukey's multiple comparison test for the condition "ISD70 only" or other indicated pair of conditions). (H) LL171 cells were treated with 200 nM of ASO for 6 hours and then stimulated with 2.5 μg / ml of ISD45 overnight. The next day, cells were lysed, and ISRE-Luc levels were analyzed by luciferase assay. ISRE-luciferase levels were normalized to the "ISD45 only" condition after background correction with NT conditions. Data shown are averages from three independent experiments with biological triplicates (±s.e.m. and ordinary one-way ANOVA with Dunnett's multiple comparison test against the "ISD70 only" condition). (I) THP-1 cells were treated with 100 nM C2-Mut1 for 6 h or transfected with 100, 250, or 500 nM C2-Mut1-PS and 2.5 μg / ml ISD70 overnight. IP-10 levels were normalized to the "ISD70 only" condition after background correction with the NT condition. Data shown are averages from two independent experiments with biological triplicates (±s.e.m. and Mann-Whitney U test against the "ISD70 only" condition). (J) Sequences of [LINC-PINT] ASO101–116, showing the location of the inhibitory GGUCCC motif (pink) from ASO103 identified by MEME (see D and Figure 6B). Yellow regions highlight the gapmer DNA portion. (K) THP-1 cells pretreated overnight with 100 nM of the indicated [LINC-PINT] ASO were transfected with 2.5 μg / ml ISD70 for 7.5 hours, and IP-10 levels in the supernatants were determined by ELISA. IP-10 levels were normalized to the "ISD70 only" condition after background correction with the NT condition.Data shown are averages from three independent experiments with biological triplicates (±s.e.m. and one-way ANOVA with Dunnett's multiple comparison test against the "ISD70 only" condition). (L, M) LL171 cells were treated with 200 nM ASO for 6 h or 20 min, then "washed" or not (L), or treated with 50, 100, or 200 nM ASO for 20 min (M), and stimulated overnight with 2.5 μg / ml ISD45. The next day, cells were lysed, and ISRE-Luc levels were analyzed by luciferase assay. ISRE-luciferase levels were normalized to the "ISD45 only" condition after background correction with the NT condition. Data shown are averages from two (L) or three (M) independent experiments with biological triplicates (±s.e.m. and one-way ANOVA with Tukey's multiple comparison test (L) for the "ISD45 only" condition, or Mann-Whitney U test (M) for "ISD45 only," or other indicated pair of conditions). (N) THP-1 cells pretreated with 250 nM of the indicated ASO for 40 min were transfected with 2.5 μg / ml ISD70 overnight, and IP-10 levels in the supernatants were determined by ELISA. Data shown are averages from three independent experiments with biological triplicates (±s.e.m. and one-way ANOVA with Dunnett's multiple comparison test for the "ISD70 only" condition, and Mann-Whitney U test comparing Mut1-dC and dC20). *P ≤ 0.05, **P ≤ 0.01, ***P ≤ 0.001, ****P ≤ 0.0001, ns: not significant. [Figure 2F]Identification of highly potent inhibitors of cGAS sensing. (A) Top: Sequence alignment of cGAS inhibitors and identification of key bases predicted by MEME analysis highlighted by arrows (Figure 6A). Bottom: Design of C2 and ASO2 mutants incorporating point mutations at selected positions in the MEME motif (mutations highlighted in yellow). (B, D) HT-29 cells pretreated overnight with the indicated doses of ASO (500, 250, 125, or 62.5 nM) were transfected with or without 2.5 μg / ml ISD70 for 24 hours, and IP-10 levels in the supernatants were determined by ELISA. IP-10 levels were normalized to the "ISD70 only" condition after background correction with the NT condition. Data shown are averages from two (D) or three (B) independent experiments in biological triplicates (±s.e.m. and ordinary two-way ANOVA with Tukey's multiple comparison test for C2 [B] or ASO2 [D] conditions; comparisons were not significant between 250 and 500 nM ASO). (C) Mouse LL171 cells were treated with the indicated amounts of ASO (200, 400, or 600 nM) for 6 hours and then stimulated overnight with 2.5 μg / ml ISD45. The next day, cells were lysed, and ISRE-Luc levels were analyzed by luciferase assay. ISRE-luciferase levels were normalized to the "ISD45 only" condition after background correction with the NT condition. Data shown are averages from two independent experiments in biological triplicates (±s.e.m. and Mann-Whitney U test). (E) Top: Sequence alignment of C2-Mut1 mutants, C2-ASO2-A and C2-ASO2-B, with the 3' ends from ASO2 up (ASO2up) and ASO2 down (ASO2down) underlined. Bottom: Homopolymer dC20 mutants, with the 2'OMe bases in pink and the cGAS inhibitory motif underlined. (F) HT-29 cells pretreated overnight with 187.5 nM of the indicated ASO were transfected or not with 2.5 μg / ml ISD70 for 24 h, and IP-10 levels in the supernatants were determined by ELISA.(G) THP-1 cells pretreated overnight with 100 nM of the indicated ASO were transfected with 2.5 μg / ml of ISD70 for 24 hours, and IP-10 levels in the supernatants were determined by ELISA. (F, G) IP-10 levels were normalized to the "ISD70 only" condition after background correction with NT conditions. Data shown are averages from three independent experiments with biological triplicates (±s.e.m. and ordinary one-way ANOVA with Tukey's multiple comparison test for the condition "ISD70 only" or other indicated pair of conditions). (H) LL171 cells were treated with 200 nM of ASO for 6 hours and then stimulated with 2.5 μg / ml of ISD45 overnight. The next day, cells were lysed, and ISRE-Luc levels were analyzed by luciferase assay. ISRE-luciferase levels were normalized to the "ISD45 only" condition after background correction with NT conditions. Data shown are averages from three independent experiments with biological triplicates (±s.e.m. and ordinary one-way ANOVA with Dunnett's multiple comparison test against the "ISD70 only" condition). (I) THP-1 cells were treated with 100 nM C2-Mut1 for 6 h or transfected with 100, 250, or 500 nM C2-Mut1-PS and 2.5 μg / ml ISD70 overnight. IP-10 levels were normalized to the "ISD70 only" condition after background correction with the NT condition. Data shown are averages from two independent experiments with biological triplicates (±s.e.m. and Mann-Whitney U test against the "ISD70 only" condition). (J) Sequences of [LINC-PINT] ASO101–116, showing the location of the inhibitory GGUCCC motif (pink) from ASO103 identified by MEME (see D and Figure 6B). Yellow regions highlight the gapmer DNA portion. (K) THP-1 cells pretreated overnight with 100 nM of the indicated [LINC-PINT] ASO were transfected with 2.5 μg / ml ISD70 for 7.5 hours, and IP-10 levels in the supernatants were determined by ELISA. IP-10 levels were normalized to the "ISD70 only" condition after background correction with the NT condition.Data shown are averages from three independent experiments with biological triplicates (±s.e.m. and one-way ANOVA with Dunnett's multiple comparison test against the "ISD70 only" condition). (L, M) LL171 cells were treated with 200 nM ASO for 6 h or 20 min, then "washed" or not (L), or treated with 50, 100, or 200 nM ASO for 20 min (M), and stimulated overnight with 2.5 μg / ml ISD45. The next day, cells were lysed, and ISRE-Luc levels were analyzed by luciferase assay. ISRE-luciferase levels were normalized to the "ISD45 only" condition after background correction with the NT condition. Data shown are averages from two (L) or three (M) independent experiments with biological triplicates (±s.e.m. and one-way ANOVA with Tukey's multiple comparison test (L) for the "ISD45 only" condition, or Mann-Whitney U test (M) for "ISD45 only," or other indicated pair of conditions). (N) THP-1 cells pretreated with 250 nM of the indicated ASO for 40 min were transfected with 2.5 μg / ml ISD70 overnight, and IP-10 levels in the supernatants were determined by ELISA. Data shown are averages from three independent experiments with biological triplicates (±s.e.m. and one-way ANOVA with Dunnett's multiple comparison test for the "ISD70 only" condition, and Mann-Whitney U test comparing Mut1-dC and dC20). *P ≤ 0.05, **P ≤ 0.01, ***P ≤ 0.001, ****P ≤ 0.0001, ns: not significant. [Figure 2G]Identification of highly potent inhibitors of cGAS sensing. (A) Top: Sequence alignment of cGAS inhibitors and identification of key bases predicted by MEME analysis highlighted by arrows (Figure 6A). Bottom: Design of C2 and ASO2 mutants incorporating point mutations at selected positions in the MEME motif (mutations highlighted in yellow). (B, D) HT-29 cells pretreated overnight with the indicated doses of ASO (500, 250, 125, or 62.5 nM) were transfected with or without 2.5 μg / ml ISD70 for 24 hours, and IP-10 levels in the supernatants were determined by ELISA. IP-10 levels were normalized to the "ISD70 only" condition after background correction with the NT condition. Data shown are averages from two (D) or three (B) independent experiments in biological triplicates (±s.e.m. and ordinary two-way ANOVA with Tukey's multiple comparison test for C2 [B] or ASO2 [D] conditions; comparisons were not significant between 250 and 500 nM ASO). (C) Mouse LL171 cells were treated with the indicated amounts of ASO (200, 400, or 600 nM) for 6 hours and then stimulated overnight with 2.5 μg / ml ISD45. The next day, cells were lysed, and ISRE-Luc levels were analyzed by luciferase assay. ISRE-luciferase levels were normalized to the "ISD45 only" condition after background correction with the NT condition. Data shown are averages from two independent experiments in biological triplicates (±s.e.m. and Mann-Whitney U test). (E) Top: Sequence alignment of C2-Mut1 mutants, C2-ASO2-A and C2-ASO2-B, with the 3' ends from ASO2 up (ASO2up) and ASO2 down (ASO2down) underlined. Bottom: Homopolymer dC20 mutants, with the 2'OMe bases in pink and the cGAS inhibitory motif underlined. (F) HT-29 cells pretreated overnight with 187.5 nM of the indicated ASO were transfected or not with 2.5 μg / ml ISD70 for 24 h, and IP-10 levels in the supernatants were determined by ELISA.(G) THP-1 cells pretreated overnight with 100 nM of the indicated ASO were transfected with 2.5 μg / ml of ISD70 for 24 hours, and IP-10 levels in the supernatants were determined by ELISA. (F, G) IP-10 levels were normalized to the "ISD70 only" condition after background correction with NT conditions. Data shown are averages from three independent experiments with biological triplicates (±s.e.m. and ordinary one-way ANOVA with Tukey's multiple comparison test for the condition "ISD70 only" or other indicated pair of conditions). (H) LL171 cells were treated with 200 nM of ASO for 6 hours and then stimulated with 2.5 μg / ml of ISD45 overnight. The next day, cells were lysed, and ISRE-Luc levels were analyzed by luciferase assay. ISRE-luciferase levels were normalized to the "ISD45 only" condition after background correction with NT conditions. Data shown are averages from three independent experiments with biological triplicates (±s.e.m. and ordinary one-way ANOVA with Dunnett's multiple comparison test against the "ISD70 only" condition). (I) THP-1 cells were treated with 100 nM C2-Mut1 for 6 h or transfected with 100, 250, or 500 nM C2-Mut1-PS and 2.5 μg / ml ISD70 overnight. IP-10 levels were normalized to the "ISD70 only" condition after background correction with the NT condition. Data shown are averages from two independent experiments with biological triplicates (±s.e.m. and Mann-Whitney U test against the "ISD70 only" condition). (J) Sequences of [LINC-PINT] ASO101–116, showing the location of the inhibitory GGUCCC motif (pink) from ASO103 identified by MEME (see D and Figure 6B). Yellow regions highlight the gapmer DNA portion. (K) THP-1 cells pretreated overnight with 100 nM of the indicated [LINC-PINT] ASO were transfected with 2.5 μg / ml ISD70 for 7.5 hours, and IP-10 levels in the supernatants were determined by ELISA. IP-10 levels were normalized to the "ISD70 only" condition after background correction with the NT condition.Data shown are averages from three independent experiments with biological triplicates (±s.e.m. and one-way ANOVA with Dunnett's multiple comparison test against the "ISD70 only" condition). (L, M) LL171 cells were treated with 200 nM ASO for 6 h or 20 min, then "washed" or not (L), or treated with 50, 100, or 200 nM ASO for 20 min (M), and stimulated overnight with 2.5 μg / ml ISD45. The next day, cells were lysed, and ISRE-Luc levels were analyzed by luciferase assay. ISRE-luciferase levels were normalized to the "ISD45 only" condition after background correction with the NT condition. Data shown are averages from two (L) or three (M) independent experiments with biological triplicates (±s.e.m. and one-way ANOVA with Tukey's multiple comparison test (L) for the "ISD45 only" condition, or Mann-Whitney U test (M) for "ISD45 only," or other indicated pair of conditions). (N) THP-1 cells pretreated with 250 nM of the indicated ASO for 40 min were transfected with 2.5 μg / ml ISD70 overnight, and IP-10 levels in the supernatants were determined by ELISA. Data shown are averages from three independent experiments with biological triplicates (±s.e.m. and one-way ANOVA with Dunnett's multiple comparison test for the "ISD70 only" condition, and Mann-Whitney U test comparing Mut1-dC and dC20). *P ≤ 0.05, **P ≤ 0.01, ***P ≤ 0.001, ****P ≤ 0.0001, ns: not significant. [Figure 2H]Identification of highly potent inhibitors of cGAS sensing. (A) Top: Sequence alignment of cGAS inhibitors and identification of key bases predicted by MEME analysis highlighted by arrows (Figure 6A). Bottom: Design of C2 and ASO2 mutants incorporating point mutations at selected positions in the MEME motif (mutations highlighted in yellow). (B, D) HT-29 cells pretreated overnight with the indicated doses of ASO (500, 250, 125, or 62.5 nM) were transfected with or without 2.5 μg / ml ISD70 for 24 hours, and IP-10 levels in the supernatants were determined by ELISA. IP-10 levels were normalized to the "ISD70 only" condition after background correction with the NT condition. Data shown are averages from two (D) or three (B) independent experiments in biological triplicates (±s.e.m. and ordinary two-way ANOVA with Tukey's multiple comparison test for C2 [B] or ASO2 [D] conditions; comparisons were not significant between 250 and 500 nM ASO). (C) Mouse LL171 cells were treated with the indicated amounts of ASO (200, 400, or 600 nM) for 6 hours and then stimulated overnight with 2.5 μg / ml ISD45. The next day, cells were lysed, and ISRE-Luc levels were analyzed by luciferase assay. ISRE-luciferase levels were normalized to the "ISD45 only" condition after background correction with the NT condition. Data shown are averages from two independent experiments in biological triplicates (±s.e.m. and Mann-Whitney U test). (E) Top: Sequence alignment of C2-Mut1 mutants, C2-ASO2-A and C2-ASO2-B, with the 3' ends from ASO2 up (ASO2up) and ASO2 down (ASO2down) underlined. Bottom: Homopolymer dC20 mutants, with the 2'OMe bases in pink and the cGAS inhibitory motif underlined. (F) HT-29 cells pretreated overnight with 187.5 nM of the indicated ASO were transfected or not with 2.5 μg / ml ISD70 for 24 h, and IP-10 levels in the supernatants were determined by ELISA.(G) THP-1 cells pretreated overnight with 100 nM of the indicated ASO were transfected with 2.5 μg / ml of ISD70 for 24 hours, and IP-10 levels in the supernatants were determined by ELISA. (F, G) IP-10 levels were normalized to the "ISD70 only" condition after background correction with NT conditions. Data shown are averages from three independent experiments with biological triplicates (±s.e.m. and ordinary one-way ANOVA with Tukey's multiple comparison test for the condition "ISD70 only" or other indicated pair of conditions). (H) LL171 cells were treated with 200 nM of ASO for 6 hours and then stimulated with 2.5 μg / ml of ISD45 overnight. The next day, cells were lysed, and ISRE-Luc levels were analyzed by luciferase assay. ISRE-luciferase levels were normalized to the "ISD45 only" condition after background correction with NT conditions. Data shown are averages from three independent experiments with biological triplicates (±s.e.m. and ordinary one-way ANOVA with Dunnett's multiple comparison test against the "ISD70 only" condition). (I) THP-1 cells were treated with 100 nM C2-Mut1 for 6 h or transfected with 100, 250, or 500 nM C2-Mut1-PS and 2.5 μg / ml ISD70 overnight. IP-10 levels were normalized to the "ISD70 only" condition after background correction with the NT condition. Data shown are averages from two independent experiments with biological triplicates (±s.e.m. and Mann-Whitney U test against the "ISD70 only" condition). (J) Sequences of [LINC-PINT] ASO101–116, showing the location of the inhibitory GGUCCC motif (pink) from ASO103 identified by MEME (see D and Figure 6B). Yellow regions highlight the gapmer DNA portion. (K) THP-1 cells pretreated overnight with 100 nM of the indicated [LINC-PINT] ASO were transfected with 2.5 μg / ml ISD70 for 7.5 hours, and IP-10 levels in the supernatants were determined by ELISA. IP-10 levels were normalized to the "ISD70 only" condition after background correction with the NT condition.Data shown are averages from three independent experiments with biological triplicates (±s.e.m. and one-way ANOVA with Dunnett's multiple comparison test against the "ISD70 only" condition). (L, M) LL171 cells were treated with 200 nM ASO for 6 h or 20 min, then "washed" or not (L), or treated with 50, 100, or 200 nM ASO for 20 min (M), and stimulated overnight with 2.5 μg / ml ISD45. The next day, cells were lysed, and ISRE-Luc levels were analyzed by luciferase assay. ISRE-luciferase levels were normalized to the "ISD45 only" condition after background correction with the NT condition. Data shown are averages from two (L) or three (M) independent experiments with biological triplicates (±s.e.m. and one-way ANOVA with Tukey's multiple comparison test (L) for the "ISD45 only" condition, or Mann-Whitney U test (M) for "ISD45 only," or other indicated pair of conditions). (N) THP-1 cells pretreated with 250 nM of the indicated ASO for 40 min were transfected with 2.5 μg / ml ISD70 overnight, and IP-10 levels in the supernatants were determined by ELISA. Data shown are averages from three independent experiments with biological triplicates (±s.e.m. and one-way ANOVA with Dunnett's multiple comparison test for the "ISD70 only" condition, and Mann-Whitney U test comparing Mut1-dC and dC20). *P ≤ 0.05, **P ≤ 0.01, ***P ≤ 0.001, ****P ≤ 0.0001, ns: not significant. [Figure 2I]Identification of highly potent inhibitors of cGAS sensing. (A) Top: Sequence alignment of cGAS inhibitors and identification of key bases predicted by MEME analysis highlighted by arrows (Figure 6A). Bottom: Design of C2 and ASO2 mutants incorporating point mutations at selected positions in the MEME motif (mutations highlighted in yellow). (B, D) HT-29 cells pretreated overnight with the indicated doses of ASO (500, 250, 125, or 62.5 nM) were transfected with or without 2.5 μg / ml ISD70 for 24 hours, and IP-10 levels in the supernatants were determined by ELISA. IP-10 levels were normalized to the "ISD70 only" condition after background correction with the NT condition. Data shown are averages from two (D) or three (B) independent experiments in biological triplicates (±s.e.m. and ordinary two-way ANOVA with Tukey's multiple comparison test for C2 [B] or ASO2 [D] conditions; comparisons were not significant between 250 and 500 nM ASO). (C) Mouse LL171 cells were treated with the indicated amounts of ASO (200, 400, or 600 nM) for 6 hours and then stimulated overnight with 2.5 μg / ml ISD45. The next day, cells were lysed, and ISRE-Luc levels were analyzed by luciferase assay. ISRE-luciferase levels were normalized to the "ISD45 only" condition after background correction with the NT condition. Data shown are averages from two independent experiments in biological triplicates (±s.e.m. and Mann-Whitney U test). (E) Top: Sequence alignment of C2-Mut1 mutants, C2-ASO2-A and C2-ASO2-B, with the 3' ends from ASO2 up (ASO2up) and ASO2 down (ASO2down) underlined. Bottom: Homopolymer dC20 mutants, with the 2'OMe bases in pink and the cGAS inhibitory motif underlined. (F) HT-29 cells pretreated overnight with 187.5 nM of the indicated ASO were transfected or not with 2.5 μg / ml ISD70 for 24 h, and IP-10 levels in the supernatants were determined by ELISA.(G) THP-1 cells pretreated overnight with 100 nM of the indicated ASO were transfected with 2.5 μg / ml of ISD70 for 24 hours, and IP-10 levels in the supernatants were determined by ELISA. (F, G) IP-10 levels were normalized to the "ISD70 only" condition after background correction with NT conditions. Data shown are averages from three independent experiments with biological triplicates (±s.e.m. and ordinary one-way ANOVA with Tukey's multiple comparison test for the condition "ISD70 only" or other indicated pair of conditions). (H) LL171 cells were treated with 200 nM of ASO for 6 hours and then stimulated with 2.5 μg / ml of ISD45 overnight. The next day, cells were lysed, and ISRE-Luc levels were analyzed by luciferase assay. ISRE-luciferase levels were normalized to the "ISD45 only" condition after background correction with NT conditions. Data shown are averages from three independent experiments with biological triplicates (±s.e.m. and ordinary one-way ANOVA with Dunnett's multiple comparison test against the "ISD70 only" condition). (I) THP-1 cells were treated with 100 nM C2-Mut1 for 6 h or transfected with 100, 250, or 500 nM C2-Mut1-PS and 2.5 μg / ml ISD70 overnight. IP-10 levels were normalized to the "ISD70 only" condition after background correction with the NT condition. Data shown are averages from two independent experiments with biological triplicates (±s.e.m. and Mann-Whitney U test against the "ISD70 only" condition). (J) Sequences of [LINC-PINT] ASO101–116, showing the location of the inhibitory GGUCCC motif (pink) from ASO103 identified by MEME (see D and Figure 6B). Yellow regions highlight the gapmer DNA portion. (K) THP-1 cells pretreated overnight with 100 nM of the indicated [LINC-PINT] ASO were transfected with 2.5 μg / ml ISD70 for 7.5 hours, and IP-10 levels in the supernatants were determined by ELISA. IP-10 levels were normalized to the "ISD70 only" condition after background correction with the NT condition.Data shown are averages from three independent experiments with biological triplicates (±s.e.m. and one-way ANOVA with Dunnett's multiple comparison test against the "ISD70 only" condition). (L, M) LL171 cells were treated with 200 nM ASO for 6 h or 20 min, then "washed" or not (L), or treated with 50, 100, or 200 nM ASO for 20 min (M), and stimulated overnight with 2.5 μg / ml ISD45. The next day, cells were lysed, and ISRE-Luc levels were analyzed by luciferase assay. ISRE-luciferase levels were normalized to the "ISD45 only" condition after background correction with the NT condition. Data shown are averages from two (L) or three (M) independent experiments with biological triplicates (±s.e.m. and one-way ANOVA with Tukey's multiple comparison test (L) for the "ISD45 only" condition, or Mann-Whitney U test (M) for "ISD45 only," or other indicated pair of conditions). (N) THP-1 cells pretreated with 250 nM of the indicated ASO for 40 min were transfected with 2.5 μg / ml ISD70 overnight, and IP-10 levels in the supernatants were determined by ELISA. Data shown are averages from three independent experiments with biological triplicates (±s.e.m. and one-way ANOVA with Dunnett's multiple comparison test for the "ISD70 only" condition, and Mann-Whitney U test comparing Mut1-dC and dC20). *P ≤ 0.05, **P ≤ 0.01, ***P ≤ 0.001, ****P ≤ 0.0001, ns: not significant. [Figure 2J]Identification of highly potent inhibitors of cGAS sensing. (A) Top: Sequence alignment of cGAS inhibitors and identification of key bases predicted by MEME analysis highlighted by arrows (Figure 6A). Bottom: Design of C2 and ASO2 mutants incorporating point mutations at selected positions in the MEME motif (mutations highlighted in yellow). (B, D) HT-29 cells pretreated overnight with the indicated doses of ASO (500, 250, 125, or 62.5 nM) were transfected with or without 2.5 μg / ml ISD70 for 24 hours, and IP-10 levels in the supernatants were determined by ELISA. IP-10 levels were normalized to the "ISD70 only" condition after background correction with the NT condition. Data shown are averages from two (D) or three (B) independent experiments in biological triplicates (±s.e.m. and ordinary two-way ANOVA with Tukey's multiple comparison test for C2 [B] or ASO2 [D] conditions; comparisons were not significant between 250 and 500 nM ASO). (C) Mouse LL171 cells were treated with the indicated amounts of ASO (200, 400, or 600 nM) for 6 hours and then stimulated overnight with 2.5 μg / ml ISD45. The next day, cells were lysed, and ISRE-Luc levels were analyzed by luciferase assay. ISRE-luciferase levels were normalized to the "ISD45 only" condition after background correction with the NT condition. Data shown are averages from two independent experiments in biological triplicates (±s.e.m. and Mann-Whitney U test). (E) Top: Sequence alignment of C2-Mut1 mutants, C2-ASO2-A and C2-ASO2-B, with the 3' ends from ASO2 up (ASO2up) and ASO2 down (ASO2down) underlined. Bottom: Homopolymer dC20 mutants, with the 2'OMe bases in pink and the cGAS inhibitory motif underlined. (F) HT-29 cells pretreated overnight with 187.5 nM of the indicated ASO were transfected or not with 2.5 μg / ml ISD70 for 24 h, and IP-10 levels in the supernatants were determined by ELISA.(G) THP-1 cells pretreated overnight with 100 nM of the indicated ASO were transfected with 2.5 μg / ml of ISD70 for 24 hours, and IP-10 levels in the supernatants were determined by ELISA. (F, G) IP-10 levels were normalized to the "ISD70 only" condition after background correction with NT conditions. Data shown are averages from three independent experiments with biological triplicates (±s.e.m. and ordinary one-way ANOVA with Tukey's multiple comparison test for the condition "ISD70 only" or other indicated pair of conditions). (H) LL171 cells were treated with 200 nM of ASO for 6 hours and then stimulated with 2.5 μg / ml of ISD45 overnight. The next day, cells were lysed, and ISRE-Luc levels were analyzed by luciferase assay. ISRE-luciferase levels were normalized to the "ISD45 only" condition after background correction with NT conditions. Data shown are averages from three independent experiments with biological triplicates (±s.e.m. and ordinary one-way ANOVA with Dunnett's multiple comparison test against the "ISD70 only" condition). (I) THP-1 cells were treated with 100 nM C2-Mut1 for 6 h or transfected with 100, 250, or 500 nM C2-Mut1-PS and 2.5 μg / ml ISD70 overnight. IP-10 levels were normalized to the "ISD70 only" condition after background correction with the NT condition. Data shown are averages from two independent experiments with biological triplicates (±s.e.m. and Mann-Whitney U test against the "ISD70 only" condition). (J) Sequences of [LINC-PINT] ASO101–116, showing the location of the inhibitory GGUCCC motif (pink) from ASO103 identified by MEME (see D and Figure 6B). Yellow regions highlight the gapmer DNA portion. (K) THP-1 cells pretreated overnight with 100 nM of the indicated [LINC-PINT] ASO were transfected with 2.5 μg / ml ISD70 for 7.5 hours, and IP-10 levels in the supernatants were determined by ELISA. IP-10 levels were normalized to the "ISD70 only" condition after background correction with the NT condition.Data shown are averages from three independent experiments with biological triplicates (±s.e.m. and one-way ANOVA with Dunnett's multiple comparison test against the "ISD70 only" condition). (L, M) LL171 cells were treated with 200 nM ASO for 6 h or 20 min, then "washed" or not (L), or treated with 50, 100, or 200 nM ASO for 20 min (M), and stimulated overnight with 2.5 μg / ml ISD45. The next day, cells were lysed, and ISRE-Luc levels were analyzed by luciferase assay. ISRE-luciferase levels were normalized to the "ISD45 only" condition after background correction with the NT condition. Data shown are averages from two (L) or three (M) independent experiments with biological triplicates (±s.e.m. and one-way ANOVA with Tukey's multiple comparison test (L) for the "ISD45 only" condition, or Mann-Whitney U test (M) for "ISD45 only," or other indicated pair of conditions). (N) THP-1 cells pretreated with 250 nM of the indicated ASO for 40 min were transfected with 2.5 μg / ml ISD70 overnight, and IP-10 levels in the supernatants were determined by ELISA. Data shown are averages from three independent experiments with biological triplicates (±s.e.m. and one-way ANOVA with Dunnett's multiple comparison test for the "ISD70 only" condition, and Mann-Whitney U test comparing Mut1-dC and dC20). *P ≤ 0.05, **P ≤ 0.01, ***P ≤ 0.001, ****P ≤ 0.0001, ns: not significant. [Figure 2K]Identification of highly potent inhibitors of cGAS sensing. (A) Top: Sequence alignment of cGAS inhibitors and identification of key bases predicted by MEME analysis highlighted by arrows (Figure 6A). Bottom: Design of C2 and ASO2 mutants incorporating point mutations at selected positions in the MEME motif (mutations highlighted in yellow). (B, D) HT-29 cells pretreated overnight with the indicated doses of ASO (500, 250, 125, or 62.5 nM) were transfected with or without 2.5 μg / ml ISD70 for 24 hours, and IP-10 levels in the supernatants were determined by ELISA. IP-10 levels were normalized to the "ISD70 only" condition after background correction with the NT condition. Data shown are averages from two (D) or three (B) independent experiments in biological triplicates (±s.e.m. and ordinary two-way ANOVA with Tukey's multiple comparison test for C2 [B] or ASO2 [D] conditions; comparisons were not significant between 250 and 500 nM ASO). (C) Mouse LL171 cells were treated with the indicated amounts of ASO (200, 400, or 600 nM) for 6 hours and then stimulated overnight with 2.5 μg / ml ISD45. The next day, cells were lysed, and ISRE-Luc levels were analyzed by luciferase assay. ISRE-luciferase levels were normalized to the "ISD45 only" condition after background correction with the NT condition. Data shown are averages from two independent experiments in biological triplicates (±s.e.m. and Mann-Whitney U test). (E) Top: Sequence alignment of C2-Mut1 mutants, C2-ASO2-A and C2-ASO2-B, with the 3' ends from ASO2 up (ASO2up) and ASO2 down (ASO2down) underlined. Bottom: Homopolymer dC20 mutants, with the 2'OMe bases in pink and the cGAS inhibitory motif underlined. (F) HT-29 cells pretreated overnight with 187.5 nM of the indicated ASO were transfected or not with 2.5 μg / ml ISD70 for 24 h, and IP-10 levels in the supernatants were determined by ELISA.(G) THP-1 cells pretreated overnight with 100 nM of the indicated ASO were transfected with 2.5 μg / ml of ISD70 for 24 hours, and IP-10 levels in the supernatants were determined by ELISA. (F, G) IP-10 levels were normalized to the "ISD70 only" condition after background correction with NT conditions. Data shown are averages from three independent experiments with biological triplicates (±s.e.m. and ordinary one-way ANOVA with Tukey's multiple comparison test for the condition "ISD70 only" or other indicated pair of conditions). (H) LL171 cells were treated with 200 nM of ASO for 6 hours and then stimulated with 2.5 μg / ml of ISD45 overnight. The next day, cells were lysed, and ISRE-Luc levels were analyzed by luciferase assay. ISRE-luciferase levels were normalized to the "ISD45 only" condition after background correction with NT conditions. Data shown are averages from three independent experiments with biological triplicates (±s.e.m. and ordinary one-way ANOVA with Dunnett's multiple comparison test against the "ISD70 only" condition). (I) THP-1 cells were treated with 100 nM C2-Mut1 for 6 h or transfected with 100, 250, or 500 nM C2-Mut1-PS and 2.5 μg / ml ISD70 overnight. IP-10 levels were normalized to the "ISD70 only" condition after background correction with the NT condition. Data shown are averages from two independent experiments with biological triplicates (±s.e.m. and Mann-Whitney U test against the "ISD70 only" condition). (J) Sequences of [LINC-PINT] ASO101–116, showing the location of the inhibitory GGUCCC motif (pink) from ASO103 identified by MEME (see D and Figure 6B). Yellow regions highlight the gapmer DNA portion. (K) THP-1 cells pretreated overnight with 100 nM of the indicated [LINC-PINT] ASO were transfected with 2.5 μg / ml ISD70 for 7.5 hours, and IP-10 levels in the supernatants were determined by ELISA. IP-10 levels were normalized to the "ISD70 only" condition after background correction with the NT condition.Data shown are averages from three independent experiments with biological triplicates (±s.e.m. and one-way ANOVA with Dunnett's multiple comparison test against the "ISD70 only" condition). (L, M) LL171 cells were treated with 200 nM ASO for 6 h or 20 min, then "washed" or not (L), or treated with 50, 100, or 200 nM ASO for 20 min (M), and stimulated overnight with 2.5 μg / ml ISD45. The next day, cells were lysed, and ISRE-Luc levels were analyzed by luciferase assay. ISRE-luciferase levels were normalized to the "ISD45 only" condition after background correction with the NT condition. Data shown are averages from two (L) or three (M) independent experiments with biological triplicates (±s.e.m. and one-way ANOVA with Tukey's multiple comparison test (L) for the "ISD45 only" condition, or Mann-Whitney U test (M) for "ISD45 only," or other indicated pair of conditions). (N) THP-1 cells pretreated with 250 nM of the indicated ASO for 40 min were transfected with 2.5 μg / ml ISD70 overnight, and IP-10 levels in the supernatants were determined by ELISA. Data shown are averages from three independent experiments with biological triplicates (±s.e.m. and one-way ANOVA with Dunnett's multiple comparison test for the "ISD70 only" condition, and Mann-Whitney U test comparing Mut1-dC and dC20). *P ≤ 0.05, **P ≤ 0.01, ***P ≤ 0.001, ****P ≤ 0.0001, ns: not significant. [Figure 2L]Identification of highly potent inhibitors of cGAS sensing. (A) Top: Sequence alignment of cGAS inhibitors and identification of key bases predicted by MEME analysis highlighted by arrows (Figure 6A). Bottom: Design of C2 and ASO2 mutants incorporating point mutations at selected positions in the MEME motif (mutations highlighted in yellow). (B, D) HT-29 cells pretreated overnight with the indicated doses of ASO (500, 250, 125, or 62.5 nM) were transfected with or without 2.5 μg / ml ISD70 for 24 hours, and IP-10 levels in the supernatants were determined by ELISA. IP-10 levels were normalized to the "ISD70 only" condition after background correction with the NT condition. Data shown are averages from two (D) or three (B) independent experiments in biological triplicates (±s.e.m. and ordinary two-way ANOVA with Tukey's multiple comparison test for C2 [B] or ASO2 [D] conditions; comparisons were not significant between 250 and 500 nM ASO). (C) Mouse LL171 cells were treated with the indicated amounts of ASO (200, 400, or 600 nM) for 6 hours and then stimulated overnight with 2.5 μg / ml ISD45. The next day, cells were lysed, and ISRE-Luc levels were analyzed by luciferase assay. ISRE-luciferase levels were normalized to the "ISD45 only" condition after background correction with the NT condition. Data shown are averages from two independent experiments in biological triplicates (±s.e.m. and Mann-Whitney U test). (E) Top: Sequence alignment of C2-Mut1 mutants, C2-ASO2-A and C2-ASO2-B, with the 3' ends from ASO2 up (ASO2up) and ASO2 down (ASO2down) underlined. Bottom: Homopolymer dC20 mutants, with the 2'OMe bases in pink and the cGAS inhibitory motif underlined. (F) HT-29 cells pretreated overnight with 187.5 nM of the indicated ASO were transfected or not with 2.5 μg / ml ISD70 for 24 h, and IP-10 levels in the supernatants were determined by ELISA.(G) THP-1 cells pretreated overnight with 100 nM of the indicated ASO were transfected with 2.5 μg / ml of ISD70 for 24 hours, and IP-10 levels in the supernatants were determined by ELISA. (F, G) IP-10 levels were normalized to the "ISD70 only" condition after background correction with NT conditions. Data shown are averages from three independent experiments with biological triplicates (±s.e.m. and ordinary one-way ANOVA with Tukey's multiple comparison test for the condition "ISD70 only" or other indicated pair of conditions). (H) LL171 cells were treated with 200 nM of ASO for 6 hours and then stimulated with 2.5 μg / ml of ISD45 overnight. The next day, cells were lysed, and ISRE-Luc levels were analyzed by luciferase assay. ISRE-luciferase levels were normalized to the "ISD45 only" condition after background correction with NT conditions. Data shown are averages from three independent experiments with biological triplicates (±s.e.m. and ordinary one-way ANOVA with Dunnett's multiple comparison test against the "ISD70 only" condition). (I) THP-1 cells were treated with 100 nM C2-Mut1 for 6 h or transfected with 100, 250, or 500 nM C2-Mut1-PS and 2.5 μg / ml ISD70 overnight. IP-10 levels were normalized to the "ISD70 only" condition after background correction with the NT condition. Data shown are averages from two independent experiments with biological triplicates (±s.e.m. and Mann-Whitney U test against the "ISD70 only" condition). (J) Sequences of [LINC-PINT] ASO101–116, showing the location of the inhibitory GGUCCC motif (pink) from ASO103 identified by MEME (see D and Figure 6B). Yellow regions highlight the gapmer DNA portion. (K) THP-1 cells pretreated overnight with 100 nM of the indicated [LINC-PINT] ASO were transfected with 2.5 μg / ml ISD70 for 7.5 hours, and IP-10 levels in the supernatants were determined by ELISA. IP-10 levels were normalized to the "ISD70 only" condition after background correction with the NT condition.Data shown are averages from three independent experiments with biological triplicates (±s.e.m. and one-way ANOVA with Dunnett's multiple comparison test against the "ISD70 only" condition). (L, M) LL171 cells were treated with 200 nM ASO for 6 h or 20 min, then "washed" or not (L), or treated with 50, 100, or 200 nM ASO for 20 min (M), and stimulated overnight with 2.5 μg / ml ISD45. The next day, cells were lysed, and ISRE-Luc levels were analyzed by luciferase assay. ISRE-luciferase levels were normalized to the "ISD45 only" condition after background correction with the NT condition. Data shown are averages from two (L) or three (M) independent experiments with biological triplicates (±s.e.m. and one-way ANOVA with Tukey's multiple comparison test (L) for the "ISD45 only" condition, or Mann-Whitney U test (M) for "ISD45 only," or other indicated pair of conditions). (N) THP-1 cells pretreated with 250 nM of the indicated ASO for 40 min were transfected with 2.5 μg / ml ISD70 overnight, and IP-10 levels in the supernatants were determined by ELISA. Data shown are averages from three independent experiments with biological triplicates (±s.e.m. and one-way ANOVA with Dunnett's multiple comparison test for the "ISD70 only" condition, and Mann-Whitney U test comparing Mut1-dC and dC20). *P ≤ 0.05, **P ≤ 0.01, ***P ≤ 0.001, ****P ≤ 0.0001, ns: not significant. [Figure 2M]Identification of highly potent inhibitors of cGAS sensing. (A) Top: Sequence alignment of cGAS inhibitors and identification of key bases predicted by MEME analysis highlighted by arrows (Figure 6A). Bottom: Design of C2 and ASO2 mutants incorporating point mutations at selected positions in the MEME motif (mutations highlighted in yellow). (B, D) HT-29 cells pretreated overnight with the indicated doses of ASO (500, 250, 125, or 62.5 nM) were transfected with or without 2.5 μg / ml ISD70 for 24 hours, and IP-10 levels in the supernatants were determined by ELISA. IP-10 levels were normalized to the "ISD70 only" condition after background correction with the NT condition. Data shown are averages from two (D) or three (B) independent experiments in biological triplicates (±s.e.m. and ordinary two-way ANOVA with Tukey's multiple comparison test for C2 [B] or ASO2 [D] conditions; comparisons were not significant between 250 and 500 nM ASO). (C) Mouse LL171 cells were treated with the indicated amounts of ASO (200, 400, or 600 nM) for 6 hours and then stimulated overnight with 2.5 μg / ml ISD45. The next day, cells were lysed, and ISRE-Luc levels were analyzed by luciferase assay. ISRE-luciferase levels were normalized to the "ISD45 only" condition after background correction with the NT condition. Data shown are averages from two independent experiments in biological triplicates (±s.e.m. and Mann-Whitney U test). (E) Top: Sequence alignment of C2-Mut1 mutants, C2-ASO2-A and C2-ASO2-B, with the 3' ends from ASO2 up (ASO2up) and ASO2 down (ASO2down) underlined. Bottom: Homopolymer dC20 mutants, with the 2'OMe bases in pink and the cGAS inhibitory motif underlined. (F) HT-29 cells pretreated overnight with 187.5 nM of the indicated ASO were transfected or not with 2.5 μg / ml ISD70 for 24 h, and IP-10 levels in the supernatants were determined by ELISA.(G) THP-1 cells pretreated overnight with 100 nM of the indicated ASO were transfected with 2.5 μg / ml of ISD70 for 24 hours, and IP-10 levels in the supernatants were determined by ELISA. (F, G) IP-10 levels were normalized to the "ISD70 only" condition after background correction with NT conditions. Data shown are averages from three independent experiments with biological triplicates (±s.e.m. and ordinary one-way ANOVA with Tukey's multiple comparison test for the condition "ISD70 only" or other indicated pair of conditions). (H) LL171 cells were treated with 200 nM of ASO for 6 hours and then stimulated with 2.5 μg / ml of ISD45 overnight. The next day, cells were lysed, and ISRE-Luc levels were analyzed by luciferase assay. ISRE-luciferase levels were normalized to the "ISD45 only" condition after background correction with NT conditions. Data shown are averages from three independent experiments with biological triplicates (±s.e.m. and ordinary one-way ANOVA with Dunnett's multiple comparison test against the "ISD70 only" condition). (I) THP-1 cells were treated with 100 nM C2-Mut1 for 6 h or transfected with 100, 250, or 500 nM C2-Mut1-PS and 2.5 μg / ml ISD70 overnight. IP-10 levels were normalized to the "ISD70 only" condition after background correction with the NT condition. Data shown are averages from two independent experiments with biological triplicates (±s.e.m. and Mann-Whitney U test against the "ISD70 only" condition). (J) Sequences of [LINC-PINT] ASO101–116, showing the location of the inhibitory GGUCCC motif (pink) from ASO103 identified by MEME (see D and Figure 6B). Yellow regions highlight the gapmer DNA portion. (K) THP-1 cells pretreated overnight with 100 nM of the indicated [LINC-PINT] ASO were transfected with 2.5 μg / ml ISD70 for 7.5 hours, and IP-10 levels in the supernatants were determined by ELISA. IP-10 levels were normalized to the "ISD70 only" condition after background correction with the NT condition.Data shown are averages from three independent experiments with biological triplicates (±s.e.m. and one-way ANOVA with Dunnett's multiple comparison test against the "ISD70 only" condition). (L, M) LL171 cells were treated with 200 nM ASO for 6 h or 20 min, then "washed" or not (L), or treated with 50, 100, or 200 nM ASO for 20 min (M), and stimulated overnight with 2.5 μg / ml ISD45. The next day, cells were lysed, and ISRE-Luc levels were analyzed by luciferase assay. ISRE-luciferase levels were normalized to the "ISD45 only" condition after background correction with the NT condition. Data shown are averages from two (L) or three (M) independent experiments with biological triplicates (±s.e.m. and one-way ANOVA with Tukey's multiple comparison test (L) for the "ISD45 only" condition, or Mann-Whitney U test (M) for "ISD45 only," or other indicated pair of conditions). (N) THP-1 cells pretreated with 250 nM of the indicated ASO for 40 min were transfected with 2.5 μg / ml ISD70 overnight, and IP-10 levels in the supernatants were determined by ELISA. Data shown are averages from three independent experiments with biological triplicates (±s.e.m. and one-way ANOVA with Dunnett's multiple comparison test for the "ISD70 only" condition, and Mann-Whitney U test comparing Mut1-dC and dC20). *P ≤ 0.05, **P ≤ 0.01, ***P ≤ 0.001, ****P ≤ 0.0001, ns: not significant. [Figure 2N]Identification of highly potent inhibitors of cGAS sensing. (A) Top: Sequence alignment of cGAS inhibitors and identification of key bases predicted by MEME analysis highlighted by arrows (Figure 6A). Bottom: Design of C2 and ASO2 mutants incorporating point mutations at selected positions in the MEME motif (mutations highlighted in yellow). (B, D) HT-29 cells pretreated overnight with the indicated doses of ASO (500, 250, 125, or 62.5 nM) were transfected with or without 2.5 μg / ml ISD70 for 24 hours, and IP-10 levels in the supernatants were determined by ELISA. IP-10 levels were normalized to the "ISD70 only" condition after background correction with the NT condition. Data shown are averages from two (D) or three (B) independent experiments in biological triplicates (±s.e.m. and ordinary two-way ANOVA with Tukey's multiple comparison test for C2 [B] or ASO2 [D] conditions; comparisons were not significant between 250 and 500 nM ASO). (C) Mouse LL171 cells were treated with the indicated amounts of ASO (200, 400, or 600 nM) for 6 hours and then stimulated overnight with 2.5 μg / ml ISD45. The next day, cells were lysed, and ISRE-Luc levels were analyzed by luciferase assay. ISRE-luciferase levels were normalized to the "ISD45 only" condition after background correction with the NT condition. Data shown are averages from two independent experiments in biological triplicates (±s.e.m. and Mann-Whitney U test). (E) Top: Sequence alignment of C2-Mut1 mutants, C2-ASO2-A and C2-ASO2-B, with the 3' ends from ASO2 up (ASO2up) and ASO2 down (ASO2down) underlined. Bottom: Homopolymer dC20 mutants, with the 2'OMe bases in pink and the cGAS inhibitory motif underlined. (F) HT-29 cells pretreated overnight with 187.5 nM of the indicated ASO were transfected or not with 2.5 μg / ml ISD70 for 24 h, and IP-10 levels in the supernatants were determined by ELISA.(G) THP-1 cells pretreated overnight with 100 nM of the indicated ASO were transfected with 2.5 μg / ml of ISD70 for 24 hours, and IP-10 levels in the supernatants were determined by ELISA. (F, G) IP-10 levels were normalized to the "ISD70 only" condition after background correction with NT conditions. Data shown are averages from three independent experiments with biological triplicates (±s.e.m. and ordinary one-way ANOVA with Tukey's multiple comparison test for the condition "ISD70 only" or other indicated pair of conditions). (H) LL171 cells were treated with 200 nM of ASO for 6 hours and then stimulated with 2.5 μg / ml of ISD45 overnight. The next day, cells were lysed, and ISRE-Luc levels were analyzed by luciferase assay. ISRE-luciferase levels were normalized to the "ISD45 only" condition after background correction with NT conditions. Data shown are averages from three independent experiments with biological triplicates (±s.e.m. and ordinary one-way ANOVA with Dunnett's multiple comparison test against the "ISD70 only" condition). (I) THP-1 cells were treated with 100 nM C2-Mut1 for 6 h or transfected with 100, 250, or 500 nM C2-Mut1-PS and 2.5 μg / ml ISD70 overnight. IP-10 levels were normalized to the "ISD70 only" condition after background correction with the NT condition. Data shown are averages from two independent experiments with biological triplicates (±s.e.m. and Mann-Whitney U test against the "ISD70 only" condition). (J) Sequences of [LINC-PINT] ASO101–116, showing the location of the inhibitory GGUCCC motif (pink) from ASO103 identified by MEME (see D and Figure 6B). Yellow regions highlight the gapmer DNA portion. (K) THP-1 cells pretreated overnight with 100 nM of the indicated [LINC-PINT] ASO were transfected with 2.5 μg / ml ISD70 for 7.5 hours, and IP-10 levels in the supernatants were determined by ELISA. IP-10 levels were normalized to the "ISD70 only" condition after background correction with the NT condition.Data shown are averages from three independent experiments with biological triplicates (±s.e.m. and one-way ANOVA with Dunnett's multiple comparison test against the "ISD70 only" condition). (L, M) LL171 cells were treated with 200 nM ASO for 6 h or 20 min, then "washed" or not (L), or treated with 50, 100, or 200 nM ASO for 20 min (M), and stimulated overnight with 2.5 μg / ml ISD45. The next day, cells were lysed, and ISRE-Luc levels were analyzed by luciferase assay. ISRE-luciferase levels were normalized to the "ISD45 only" condition after background correction with the NT condition. Data shown are averages from two (L) or three (M) independent experiments with biological triplicates (±s.e.m. and one-way ANOVA with Tukey's multiple comparison test (L) for the "ISD45 only" condition, or Mann-Whitney U test (M) for "ISD45 only," or other indicated pair of conditions). (N) THP-1 cells pretreated with 250 nM of the indicated ASO for 40 min were transfected with 2.5 μg / ml ISD70 overnight, and IP-10 levels in the supernatants were determined by ELISA. Data shown are averages from three independent experiments with biological triplicates (±s.e.m. and one-way ANOVA with Dunnett's multiple comparison test for the "ISD70 only" condition, and Mann-Whitney U test comparing Mut1-dC and dC20). *P ≤ 0.05, **P ≤ 0.01, ***P ≤ 0.001, ****P ≤ 0.0001, ns: not significant.

[0322] [Figure 3A]Sequence-dependent inhibition of cGAS function. (A, B) THP-1 cells pretreated for 6 hours with the indicated doses (500, 250, 125, 62.5, 31.25 nM) (A) or 125 nM (B) of C2-Mut1 or A151 oligonucleotides were transfected overnight with 2.5 μg / ml ISD70, and IP-10 (A) or IFN-β (B) levels in the supernatants were determined by ELISA. IP-10 (A) and IFN-β (B) levels were normalized to the "ISD70 only" condition after background correction for the NT condition. (A, B) Data shown are averages from three independent experiments with biological triplicates (±s.e.m. and ordinary two-way ANOVA with Sidak's multiple comparison test for A151 [A] or ordinary one-way ANOVA with Tukey's multiple comparison test for the condition "NT" or other indicated set of conditions [B]). MG-63 (C) or mouse immortalized BMDM (D) were pretreated with or without 500 nM ISD for 6 hours, followed by overnight stimulation with 2.5 μg / ml of ISD (ISD70 for MG-63 and ISD45 for BMDM), LPS (1 μg / ml), GSK (100 nM), PAM3C (100 ng / ml), ODN1826 (500 nM), or DMXAA (50 μg / ml). Supernatant IP-10 (C and D) and TNF-α (D) levels were determined by ELISA. (C) Data were background corrected for NT conditions and then normalized to the "GSK" condition. Data shown are averages from two independent experiments with biological triplicates (±s.e.m. and Mann-Whitney U test). (D) IP-10 levels were normalized to the DMXAA condition, while TNF-α was normalized to the LPS condition. Data shown are averages from three independent experiments with biological triplicates (±s.e.m. and Mann-Whitney U test). (E) Recombinant cGAS protein was incubated in vitro with 2.3 μM ISD70 with or without increasing concentrations of ASO (0.5, 2, and 10 μM) for 40 min. The reaction was stopped with EDTA, and cGAMP levels were analyzed by ELISA. Data were normalized to condition A151 2 μM.Data shown are averages from three independent experiments performed in technical duplicate by ELISA (±s.e.m. and Mann-Whitney U test). (F) Prior to RNA purification, BJ hTERT SV40T cells were transfected overnight with increasing amounts of the indicated ASO (20, 50, 100 nM) or 2 mM aspirin (ASP). Expression of a panel of three human IFN-driven genes was analyzed by RT-qPCR. Expression of the indicated genes was reported relative to 18S expression and further normalized to the mean of the "mock" condition. Data shown represent the mean of three independent experiments with biological duplicates (±s.e.m. and Mann-Whitney U test). (G) Prior to RNA purification, Trex1 mutant primary BMDMs from three different mice were transfected with 50 nM ASO (or Lipofectamine alone, "mock") for 20 h. Expression of a panel of three mouse IFN-driven genes was analyzed by RT-qPCR. Expression of the indicated genes was reported relative to 18S expression and further normalized to the mean of the "mock" condition. Data shown represent the mean of three mice performed in biological duplicates (±s.e.m. and Mann-Whitney U test shown). *P≦0.05, **P≦0.01, ***P≦0.001, ****P≦0.0001, ns: not significant. [Figure 3B]Sequence-dependent inhibition of cGAS function. (A, B) THP-1 cells pretreated for 6 hours with the indicated doses (500, 250, 125, 62.5, 31.25 nM) (A) or 125 nM (B) of C2-Mut1 or A151 oligonucleotides were transfected overnight with 2.5 μg / ml ISD70, and IP-10 (A) or IFN-β (B) levels in the supernatants were determined by ELISA. IP-10 (A) and IFN-β (B) levels were normalized to the "ISD70 only" condition after background correction for the NT condition. (A, B) Data shown are averages from three independent experiments with biological triplicates (±s.e.m. and ordinary two-way ANOVA with Sidak's multiple comparison test for A151 [A] or ordinary one-way ANOVA with Tukey's multiple comparison test for the condition "NT" or other indicated set of conditions [B]). MG-63 (C) or mouse immortalized BMDM (D) were pretreated with or without 500 nM ISD for 6 hours, followed by overnight stimulation with 2.5 μg / ml of ISD (ISD70 for MG-63 and ISD45 for BMDM), LPS (1 μg / ml), GSK (100 nM), PAM3C (100 ng / ml), ODN1826 (500 nM), or DMXAA (50 μg / ml). Supernatant IP-10 (C and D) and TNF-α (D) levels were determined by ELISA. (C) Data were background corrected for NT conditions and then normalized to the "GSK" condition. Data shown are averages from two independent experiments with biological triplicates (±s.e.m. and Mann-Whitney U test). (D) IP-10 levels were normalized to the DMXAA condition, while TNF-α was normalized to the LPS condition. Data shown are averages from three independent experiments with biological triplicates (±s.e.m. and Mann-Whitney U test). (E) Recombinant cGAS protein was incubated in vitro with 2.3 μM ISD70 with or without increasing concentrations of ASO (0.5, 2, and 10 μM) for 40 min. The reaction was stopped with EDTA, and cGAMP levels were analyzed by ELISA. Data were normalized to condition A151 2 μM.Data shown are averages from three independent experiments performed in technical duplicate by ELISA (±s.e.m. and Mann-Whitney U test). (F) Prior to RNA purification, BJ hTERT SV40T cells were transfected overnight with increasing amounts of the indicated ASO (20, 50, 100 nM) or 2 mM aspirin (ASP). Expression of a panel of three human IFN-driven genes was analyzed by RT-qPCR. Expression of the indicated genes was reported relative to 18S expression and further normalized to the mean of the "mock" condition. Data shown represent the mean of three independent experiments with biological duplicates (±s.e.m. and Mann-Whitney U test). (G) Prior to RNA purification, Trex1 mutant primary BMDMs from three different mice were transfected with 50 nM ASO (or Lipofectamine alone, "mock") for 20 h. Expression of a panel of three mouse IFN-driven genes was analyzed by RT-qPCR. Expression of the indicated genes was reported relative to 18S expression and further normalized to the mean of the "mock" condition. Data shown represent the mean of three mice performed in biological duplicates (±s.e.m. and Mann-Whitney U test shown). *P≦0.05, **P≦0.01, ***P≦0.001, ****P≦0.0001, ns: not significant. [Figure 3C]Sequence-dependent inhibition of cGAS function. (A, B) THP-1 cells pretreated for 6 hours with the indicated doses (500, 250, 125, 62.5, 31.25 nM) (A) or 125 nM (B) of C2-Mut1 or A151 oligonucleotides were transfected overnight with 2.5 μg / ml ISD70, and IP-10 (A) or IFN-β (B) levels in the supernatants were determined by ELISA. IP-10 (A) and IFN-β (B) levels were normalized to the "ISD70 only" condition after background correction for the NT condition. (A, B) Data shown are averages from three independent experiments with biological triplicates (±s.e.m. and ordinary two-way ANOVA with Sidak's multiple comparison test for A151 [A] or ordinary one-way ANOVA with Tukey's multiple comparison test for the condition "NT" or other indicated set of conditions [B]). MG-63 (C) or mouse immortalized BMDM (D) were pretreated with or without 500 nM ISD for 6 hours, followed by overnight stimulation with 2.5 μg / ml of ISD (ISD70 for MG-63 and ISD45 for BMDM), LPS (1 μg / ml), GSK (100 nM), PAM3C (100 ng / ml), ODN1826 (500 nM), or DMXAA (50 μg / ml). Supernatant IP-10 (C and D) and TNF-α (D) levels were determined by ELISA. (C) Data were background corrected for NT conditions and then normalized to the "GSK" condition. Data shown are averages from two independent experiments with biological triplicates (±s.e.m. and Mann-Whitney U test). (D) IP-10 levels were normalized to the DMXAA condition, while TNF-α was normalized to the LPS condition. Data shown are averages from three independent experiments with biological triplicates (±s.e.m. and Mann-Whitney U test). (E) Recombinant cGAS protein was incubated in vitro with 2.3 μM ISD70 with or without increasing concentrations of ASO (0.5, 2, and 10 μM) for 40 min. The reaction was stopped with EDTA, and cGAMP levels were analyzed by ELISA. Data were normalized to condition A151 2 μM.Data shown are averages from three independent experiments performed in technical duplicate by ELISA (±s.e.m. and Mann-Whitney U test). (F) Prior to RNA purification, BJ hTERT SV40T cells were transfected overnight with increasing amounts of the indicated ASO (20, 50, 100 nM) or 2 mM aspirin (ASP). Expression of a panel of three human IFN-driven genes was analyzed by RT-qPCR. Expression of the indicated genes was reported relative to 18S expression and further normalized to the mean of the "mock" condition. Data shown represent the mean of three independent experiments with biological duplicates (±s.e.m. and Mann-Whitney U test). (G) Prior to RNA purification, Trex1 mutant primary BMDMs from three different mice were transfected with 50 nM ASO (or Lipofectamine alone, "mock") for 20 h. Expression of a panel of three mouse IFN-driven genes was analyzed by RT-qPCR. Expression of the indicated genes was reported relative to 18S expression and further normalized to the mean of the "mock" condition. Data shown represent the mean of three mice performed in biological duplicates (±s.e.m. and Mann-Whitney U test shown). *P≦0.05, **P≦0.01, ***P≦0.001, ****P≦0.0001, ns: not significant. [Figure 3D]Sequence-dependent inhibition of cGAS function. (A, B) THP-1 cells pretreated for 6 hours with the indicated doses (500, 250, 125, 62.5, 31.25 nM) (A) or 125 nM (B) of C2-Mut1 or A151 oligonucleotides were transfected overnight with 2.5 μg / ml ISD70, and IP-10 (A) or IFN-β (B) levels in the supernatants were determined by ELISA. IP-10 (A) and IFN-β (B) levels were normalized to the "ISD70 only" condition after background correction for the NT condition. (A, B) Data shown are averages from three independent experiments with biological triplicates (±s.e.m. and ordinary two-way ANOVA with Sidak's multiple comparison test for A151 [A] or ordinary one-way ANOVA with Tukey's multiple comparison test for the condition "NT" or other indicated set of conditions [B]). MG-63 (C) or mouse immortalized BMDM (D) were pretreated with or without 500 nM ISD for 6 hours, followed by overnight stimulation with 2.5 μg / ml of ISD (ISD70 for MG-63 and ISD45 for BMDM), LPS (1 μg / ml), GSK (100 nM), PAM3C (100 ng / ml), ODN1826 (500 nM), or DMXAA (50 μg / ml). Supernatant IP-10 (C and D) and TNF-α (D) levels were determined by ELISA. (C) Data were background corrected for NT conditions and then normalized to the "GSK" condition. Data shown are averages from two independent experiments with biological triplicates (±s.e.m. and Mann-Whitney U test). (D) IP-10 levels were normalized to the DMXAA condition, while TNF-α was normalized to the LPS condition. Data shown are averages from three independent experiments with biological triplicates (±s.e.m. and Mann-Whitney U test). (E) Recombinant cGAS protein was incubated in vitro with 2.3 μM ISD70 with or without increasing concentrations of ASO (0.5, 2, and 10 μM) for 40 min. The reaction was stopped with EDTA, and cGAMP levels were analyzed by ELISA. Data were normalized to condition A151 2 μM.Data shown are averages from three independent experiments performed in technical duplicate by ELISA (±s.e.m. and Mann-Whitney U test). (F) Prior to RNA purification, BJ hTERT SV40T cells were transfected overnight with increasing amounts of the indicated ASO (20, 50, 100 nM) or 2 mM aspirin (ASP). Expression of a panel of three human IFN-driven genes was analyzed by RT-qPCR. Expression of the indicated genes was reported relative to 18S expression and further normalized to the mean of the "mock" condition. Data shown represent the mean of three independent experiments with biological duplicates (±s.e.m. and Mann-Whitney U test). (G) Prior to RNA purification, Trex1 mutant primary BMDMs from three different mice were transfected with 50 nM ASO (or Lipofectamine alone, "mock") for 20 h. Expression of a panel of three mouse IFN-driven genes was analyzed by RT-qPCR. Expression of the indicated genes was reported relative to 18S expression and further normalized to the mean of the "mock" condition. Data shown represent the mean of three mice performed in biological duplicates (±s.e.m. and Mann-Whitney U test shown). *P≦0.05, **P≦0.01, ***P≦0.001, ****P≦0.0001, ns: not significant. [Figure 3E]Sequence-dependent inhibition of cGAS function. (A, B) THP-1 cells pretreated for 6 hours with the indicated doses (500, 250, 125, 62.5, 31.25 nM) (A) or 125 nM (B) of C2-Mut1 or A151 oligonucleotides were transfected overnight with 2.5 μg / ml ISD70, and IP-10 (A) or IFN-β (B) levels in the supernatants were determined by ELISA. IP-10 (A) and IFN-β (B) levels were normalized to the "ISD70 only" condition after background correction for the NT condition. (A, B) Data shown are averages from three independent experiments with biological triplicates (±s.e.m. and ordinary two-way ANOVA with Sidak's multiple comparison test for A151 [A] or ordinary one-way ANOVA with Tukey's multiple comparison test for the condition "NT" or other indicated set of conditions [B]). MG-63 (C) or mouse immortalized BMDM (D) were pretreated with or without 500 nM ISD for 6 hours, followed by overnight stimulation with 2.5 μg / ml of ISD (ISD70 for MG-63 and ISD45 for BMDM), LPS (1 μg / ml), GSK (100 nM), PAM3C (100 ng / ml), ODN1826 (500 nM), or DMXAA (50 μg / ml). Supernatant IP-10 (C and D) and TNF-α (D) levels were determined by ELISA. (C) Data were background corrected for NT conditions and then normalized to the "GSK" condition. Data shown are averages from two independent experiments with biological triplicates (±s.e.m. and Mann-Whitney U test). (D) IP-10 levels were normalized to the DMXAA condition, while TNF-α was normalized to the LPS condition. ...

Claims

【1】 オリゴヌクレオチドであって、 (1)5[A / 1(7[[A / C44! / 444-3′、 ()5-AY / / A4U / / 44[5 / 44---3'、 (3)5'A!! / / A[U / / 44[5 / C)4[5 / !4!!!!!!!!! (4)5'-175252-3'、 (5)5'------------------------------- (6)5'----------------------------------- (7)5'-3-3'、 (8)5'-13321534433241443333'、 (9)55'-1125114444333214433353533'、 (100)5'-21332144344332414433333'. (11)4'-111115344433444332323335533'、 (12)-'-11157133232323'. (13)5'--133315444433247443332133'、 (14)5'-1433324744333'、 (15)5'--13-3-3' (16)5'-1444433247444333'. (17)5'-1433324744333'、 (18)5'-133315744433247443332133'、 (19)5'-11317523332157333'、 (200)5'-111522433333333'. (21)44'-1453332443344444735733'、 (22)53444447144332333'. (203)53232323232177733'、 (24)53'-244444443533'、 (255)533244443333333'. (22)53533244433333'. (27)5'--3325144333321113344433215-3'、 (208)4'--13222171332313333'、 (29)5'-111125552222233112444442553'、 (30)5'-211333722232171221117131533'、 (31)5'-5333111334431172235353'、 (32)5'3533' (33)5'-113177524433324214435333'、 (34) 5'-GAUU (35) 5' - UGAC (36)5'-AAAOO-3'、 - (38)5'-GGGGGGGGTGAGGGGGMM'、 (39)5'-22GAATGTGATG22222-3'、 (40)5'--22--3'、 (41)5'-EEEUUTTTAATTTTGUAGAG-3'、 (42) 5'-GUSHAAA (43)5'-3--3--3'、 - - (46)5'-UUAAATAATCTAGTTUGAAG-3'、 (47)5'-AAGLAGTATAAATGUAAA-3'、 (48)5'-GGEUAGTTTATATTALMAM-3'、 (49)5'-EEEGOEEEE (50)5'-UAUU:AAATGA0AGUG-3'、 (51) 5'-Uggh - (53)5'-BUUCCACATGCCAGC - - (56)5'-AAAAGATTATCTTCUUUUA-3'、 (57)5'-UGUGAAAGATTATCUUPUU-3'、 (58)5'-UUGUAAAAATTATCUUPO-3'、 (59)5'-UUUGAATTCAAACUUG-3', (60)5'-AGAAGTCTAAATGTAAAG-3'、 (61)5'-AGAUTAAAAGATUAAUA-3'、 (62)5'-AGAUUATCTTCTTTTAA UUU-3', (63)5'-------3'、 (64) 5' - 3', (65)5'-AGUGGAAOO-3'、 (66)5’-AAGAUTATCTTCTTTUAAUU-3’、 (67)5’-AGAAAGAAGCAAAGAUUCAA-3’、 (68)5’-UCCCATCCCTTCTGCUGCCA-3’、 (69)5’-ACCCUTCTCTCTGGTCCCAU-3’、 (70)5’-AAUAUCTGCTGCCCACCUUC-3’、 (71)5’-UCUCUCTGGTCCCATCCCUU-3’、 (72)5’-AGGCCTCCAGTGTCTUCUCC-3’、 (73)5’-CAAGCCCCAGCGTTCCUCCG-3’、 (74)5’-AAAUGTCCTGGCCCTCACUG-3’、 (75)5’-AAUUUAAAGCATGAAUAUUA-3’、 (76)5’-AAAAGATTATCTTCTUUUAA-3’、 (77)5’-AUAUCTGCTGCCCACCUUCU-3’、 (78)5’-CAGUCTCCATGTCCCAGGCC-3’、 (79)5’-AAAGATTATCTTCTTUUAAU-3’、 (80)5’-CCCUUCTCTCTGGTCCCAUC-3’、 (81)5’-AUGGCCTTTCCGTGCCAAGG-3’、 (82)5’-GCAUUCCGTGCGGAAGCCUU-3’、 (83)5’-GGCCGAACTTTCCCGCCUUA-3’、 (84)5’-GGUCUTGGCTTCGTGGAGCA-3’、 (85)5’-GGAGCTTCGAGGCCCCAGGC-3’、 (86)5’-GGUGGTCCACAACCCCUUUC-3’、 (87)5’-UUCUGGGGACTTCCAGUUUA-3’、 (88)5’-UGAUUCCAAAGCCAGGGUUA-3’、 (89)5’-GGGUATCGAAAGAGTCUGGA-3’、 (90)5’-CUUGCACGTGGCTTCGUCUC-3’、 (91)5’-GUGUCCTTGCACGTGGCUUC-3’、 (92)5’-GUAAAAAGCTTTTGAAGUGA-3’、 (93)5’-AUGCCATCCACTTGAUAGGC-3’、 (94)5’-UGAAGTAAAAATCAAUAGCG-3’、 (95)5’-AAGGCCCTTCGCACTUCUUA-3’、 (96)5’-GUACUCGTCGGCATCCACCA-3’、 (97)5’-GUCCUTGCACGTGGCUUCGU-3’、 (98)5'---------' (99)5'-OAAAAGAAAAA--3'、 (100)5'-22GAAGTGAAGTAAAGGAG-3'、 (101)5'-2222 (102)5'-CCAC (103)5'-UCATGGAATAAAM (104)5'-GAAAGGAATTCGAAUMO-3'、 (105)5'--3--3--3'、 (106)5'-EUAGGAAATTATTTUAGG-3'、 (107)5'-AAALACCAGTAGTALAUG-3'、 - (109)5'-UUACUTTA�B', (110)5'-222GAGTGAAGTAAAGGG3'、 (111)5'-3U33T3 (112)5'-GGAATLEAGGTTLUGG-3'、 (1113)5'-UGFTGGATFGAGUUCMU - (115)5'-UUUAGTTTTCAGAAUAL-3'、 (116)5'---------- (117)5'-GCUUATTTTAAGLATAUUAA-3'、 (118)5'-UUAUUTTAAGCATATUAAAA-3'、 (119)5'-UECUGAGCTTCCTTGECO-3'、 (120)5'-AUUATTTAAAAGAAAAG-3'、 (121)5'-BUUUCATCATCAAC - (123)5'-AAAAGAGAAAAACTA#CU"-3'、 (124)5'-GGGAATAAAGGAAL-3'、 (125)5'-----------------'- (126)5'-GGALOGAOGGAMO-3'、 (127)5'-AGUAAAAGCTTTTGAAGU-3'、 (128)5'-GGGGGGAAATAGTGGLG-3'、 (1219)4'-11112172443211247713-3'、 (1300)4'-15775442241423344721353-3'、 (1311)4'-11123323444421347444444335313' (132)4'-153534423444444235353573' (133)4'-112444444553573' (134)4'--2-3-3'. (135)4'--2-2-2-3-3'. (1136)4'-215552442112444433'. (137)4'--2-2-2-3'. (1308)4'-155522444423322213533'. (139)5'-5135244432444444445! 555-3'、 (1400)4'-17525-3'、 (14Y)5[Y / A / C\\1 / A47 / A / 44\\\ / 44\MMMMMQ (14\)5'[Q1 / A / C)WW / A / 44\4 / 4 / 44\4\!!!!! (143)4'-1-13-444-3'、 (144)4'-1-132-43-3'、 (145)4'-2-134443-3'、 (146)4'-1172243-3'、 (147)4'-3-3'、 (148)4'-1-13-44-3'、 (149)44'-3-3'、 (1500)4'-2-3-3'、 (1511)4'-4414433344444132374411134444313' (152)5'--4444473333234443744333'. (153)4'-1444332323443447471344344343' (154)5'-4414472441713344333333'. (155)5'-144444441113447177244323'. (156)44'-1114444443344417323144713443' (157)4'-14443332442333223333'. (1548)4'-144444344443244317733344433' (159)5'-1444444411113444431411117121333' (1600)-4'-134472332221444433344443' (161)4'-11444444133'、 (162)4'------------------------------------------------------------------------- (163)4'-4-4-4-4-3' (164) 5'-TCCAACACTTCGTGGG-3', (165) 5'-GTGTCTGGAAGCTTCC-3', (166) 5'-CTTGAAGCATCGTATC-3', (167) 5'-TCGTAGTTGCTTCCTA-3', (168) 5'-CGCTTTTCTGTCTGGT-3', (169) 5'-GGCTGGATCCGAGTT-3', (170) 5'-GATAGACCCTTCAGCA-3', (171) 5'-AGGACTCAGATGTTT-3', (172) 5'-GTGATCTTGACATGCT-3', (173) 5'-AGATTTTCAGAGCAGCT-3', (174) 5'-GGTTACGGCTCACGTA-3', (175) 5'-GTTAGTCCCTGTCCAT-3', (176) 5'-AGGTTGGCTTCGTG-3', (177) 5'-CTGCAGCTTCCCTTC-3', (178) 5'-GTCCCTGCACGTGGC-3', (179) 5'-GTCTCTGGAGCTTCC-3', (180) 5'-GGTTGGCTTCGTGG-3', (181) 5'-GGUA-3', (182) 5'-GUAU-3', (183) 5'-GGU-3', (184) 5'-GUA-3', (185) 5'-GUC-3', (186) 5'-GUG-3', (187) 5'-GUU-3', (188) 5'-GUA-3', (189) 5'-GGC-3', (190) 5'-AUC-3', (191) 5'-GAA-3', (192) 5'-GAG-3', (193) 5'-GGA-3', (194) 5'-GAC-3', (195) 5'-GAU-3', (196) 5'-AUG-3', (197) 5'-GCG-3', (198) 5'-UUC-3', (199) 5'-GCC-3', (200) 5'-GGG-3', (201) 5'-AUU-3', (202) 5'-GCA-3', (203) 5'-AGC-3', (204) 5'-AAC-3', (205) 5'-CCA-3', (206) 5'-UGC-3', (207) 5'-CAA-3', (208) 5'-CGG-3', (209) 5'-ACC-3', (210) 5'-AGA-3', (211) 5'-TTT-3', and (212) An oligonucleotide comprising or consisting of a motif or sequence selected from the group consisting of 5'-TCT-3', one or more nucleotides of the motif are modified, each of the modified nucleotides independently comprising a modified sugar and / or a modified internucleotide bridge; Unmodified nucleotides are either RNA or DNA nucleotides containing phosphodiester internucleotide bridges; U may be T and / or T may be U; and An oligonucleotide, wherein the oligonucleotide inhibits TLR7 activity.

2. 2. The oligonucleotide of claim 1, wherein the oligonucleotide does not bind or is not designed to bind to a transcript encoding TLR7 or its complement.

3. 2. The oligonucleotide of claim 1, wherein the oligonucleotide binds or is designed to bind to a target transcript that does not encode TLR7 or its complement.

4. the oligonucleotide comprises a 5' region, an intermediate region, and a 3' region; a) the 5' region comprises one or more modified nucleotides, each of which independently comprises a modified sugar and / or a modified internucleotide bridge; b) said intermediate region comprises ribonucleic acid, deoxyribonucleic acid, or a combination thereof, optionally with modified internucleotide bridges; and 2. The oligonucleotide of claim 1, wherein c) the 3' region comprises one or more modified nucleotides, each of which independently comprises a modified sugar and / or a modified internucleotide bridge.

5. The motif may be selected from the sequence 5'-GGUAUC-3', 5'-AGUCUC-3', 5'-GGUCCC-3', 5'-GGUCUC-3', 5'-AAGCUC-3', 5'-AGUCCC-3', 5'-GGUAUA-3', 5'-UGUUUC-3', 5'-UGUGUC-3', 5'-CGUUUC-3', 5'-CGUGUC-3', 5'-GGUAU-3', 5'-GGUA-3', 5'-GUAU-3', 5'-GGU-3', 5'-GUA-3', 5'-GUC-3', 5'-GUG-3', 5'-GUU-3', 5'-GUA-3', 5'-GU 2. The oligonucleotide of claim 1, comprising: GGC-3', 5'-AUC-3', 5'-GAA-3', 5'-GAG-3', 5'-GGA-3', 5'-GAC-3', 5'-GAU-3', 5'-AUG-3', 5'-GCG-3', 5'-UUC-3', 5'-GCC-3', 5'-GGG-3', 5'-AUU-3', 5'-GCA-3', 5'-AGC-3', 5'-AAC-3', 5'-CCA-3', 5'-UGC-3', 5'-CAA-3', 5'-CGG-3', 5'-ACC-3', or 5'-AGA-3', and U may be T.

6. The motif has the sequence 5'-GGUAUC-3', 5'-GGUATC-3', 5'-AGUCTC-3', 5'-AGTCTC-3', 5'-GGUCCC-3', 5'-GGUCTC-3', 5'-AAGCUC-3', 5'-AGTCCC-3', 5'-GGUATA-3', 5'-UGUTTC-3', 2. The oligonucleotide of claim 1, having the following structure: 5'-UGUGTC-3', 5'-CGUTTC-3', 5'-CGUGTC-3', 5'-GGUAU-3', 5'-GGUAT-3', 5'-GGUA-3', 5'-GUAU-3', 5'-GUAT-3', 5'-GGU-3', or 5'-GUA-3'.

7. The oligonucleotide of claim 1, wherein the motif has the sequence 5'-GUC-3'.

8. 2. The oligonucleotide of claim 1, wherein the modified sugar comprises 2'-O-methyl, 2'-O-methoxyethoxy, 2'-fluoro, 2'-allyl, 2'-O-[2-(methylamino)-2-oxoethyl], 4'-thio, 4'-CH2-O-2'-bridged, 4'-(CH2)2-O-2'-bridged, 2'-LNA, 2'-amino, fluoroarabinonucleotide, threose nucleic acid, or 2'-O--(N-methylcarbamate).

9. 2. The oligonucleotide of claim 1, wherein the modified internucleotide bridge comprises a phosphorothioate, a non-bridging oxygen atom replacing a sulfur atom, a phosphonate such as a methylphosphonate, a phosphodiester, a phosphoromorpholidate, a phosphoropiperazidate, an amide, a methylene (methylamino), a formacetal, a thioformacetal, a peptide nucleic acid or a phosphoramidate such as a morpholinophosphorodiamidate (PMO), an N3'-P5' phosphoramidite, or a thiophosphoramidite.

10. the modified sugar and modified internucleotide bridge (a) 2'O-methyl and phosphorothioate internucleotide crosslinks; (b) 2'-LNA and phosphorothioate internucleotide bridges; or (c) 2'-O-methoxyethoxy and phosphorothioate internucleotide crosslinks; The oligonucleotide of claim 1 , comprising:

11. A composition comprising the oligonucleotide according to any one of claims 1 to 10.

12. The composition of claim 11 , wherein the composition is an immunogenic composition comprising an RNA molecule and the oligonucleotide.

13. The composition of claim 12, wherein the immunogenic composition is an RNA composition or an mRNA vaccine composition comprising the oligonucleotide.

14. A pharmaceutical composition for treating or preventing a disease, disorder, or condition associated with increased, excessive, or abnormal TLR7 expression, activity, and / or signaling in a subject, comprising the oligonucleotide of any one of claims 1 to 10.

15. The disease, disorder, or condition is selected from the group consisting of Huntington's disease, Parkinson's disease, motor neuron disease (MND), amyotrophic lateral sclerosis (ALS), prion disease, frontotemporal dementia, traumatic brain injury, Alzheimer's disease, acute pancreatitis, silica-induced fibrosis, age-related macular degeneration, Aicardi syndrome, myocardial infarction, heart failure, polyarthritis / fetal and neonatal anemia, systemic lupus erythematosus, acute kidney injury, alcohol-related liver disease, non-alcoholic fatty liver disease, silica-driven pulmonary inflammation, chronic obstructive pulmonary disease, brain damage following ischemic stroke, sepsis, non-alcoholic steatohepatitis (NASH), cancer, sickle cell disease, inflammatory bowel disease, type 2 diabetes mellitus, overnutrition-induced obesity, COVID-19, chronic obstructive pulmonary disorder (COPD), hematopoietic disorders, aging-related inflammation, Cutibacterium acnes, 15. The pharmaceutical composition of claim 14, wherein the therapeutic agent is selected from the group consisting of H. acnes infection, hepatitis B, posterior eye disease, arthritis, rheumatoid arthritis, emphysema, colorectal cancer, skin cancer, metastasis, and breast cancer.

16. The pharmaceutical composition of claim 14, wherein the oligonucleotide is capable of reducing TLR7 expression or activity when administered to a subject.

17. 17. The pharmaceutical composition of claim 16, wherein the disease, disorder, or condition is an inflammatory disease, disorder, or condition associated with administration of a therapeutic oligonucleotide to the subject.

18. 18. The pharmaceutical composition of claim 17, wherein the inflammatory disease, disorder, or condition comprises liver inflammation.

19. A pharmaceutical composition for inhibiting TLR7 in a subject, comprising an oligonucleotide according to any one of claims 1 to 10.

20. 20. The pharmaceutical composition of claim 19, wherein the oligonucleotide at least partially inhibits TLR7 activation in a cell in response to an RNA molecule.

21. The pharmaceutical composition of claim 20 , wherein the RNA molecule is an exogenous mRNA molecule.

22. 11. An in vitro method of inhibiting TLR7 in a cell comprising the step of contacting the cell with an effective amount of an oligonucleotide according to any one of claims 1 to 10.