Pan-genotypic agents against influenza virus and methods of using the same

1. An oligonucleotide compound or salt thereof, comprising an oligonucleotide sequence comprising at least 8 nucleoside subunits complementary to a region of a Packaging Stein-Loop 2 (PSL2) motif of a PB2 viral RNA (vRNA) or mutant thereof, wherein the oligonucleotide compound inhibits virus production and the oligonucleotide sequence is selected from:

(SEQ ID NO:45)

5' ACCAAAAGAAT 3'; 

(SEQ ID NO:50)

5' CAGACACGACCAAAA 3'; 

(SEQ ID NO:51)

5' TACTTACTGACAGCC 3'; 

(SEQ ID NO:52)

5' AGACACGACCAAAAG 3'; 

(SEQ ID NO:56)

5' CGACCAAAAGAATTC 3'; 

(SEQ ID NO:58)

5' GATGGCCATCAATTA 3'; 

(SEQ ID NO:60)

5' TCTAGCATACTTACT 3'; 

(SEQ ID NO:63)

5' GGCCATCAATTAGTG 3'; 

(SEQ ID NO:64)

5' TTCGGATGGCCATCA 3'; 

(SEQ ID NO:66)

5' GACAGCCAGACAGCA 3'; 

(SEQ ID NO:91)

5' TACTTACTGACAGTC 3'; 

(SEQ ID NO:92)

5' TACTTACCGACAGCC 3'; and 

(SEQ ID NO:93) 

5' GGATTTCGGATGGCCA 3'.

2. The compound of claim 1 , wherein the oligonucleotide comprises an internucleoside linkage selected from: phosphorothioate, phosphorodithioate, phosphoramidate and thiophosphoramidate linkages.

3. The compound of claim 1 , wherein the oligonucleotide comprises a locked nucleic acid (LNA) nucleotide.

4. The compound of claim 1 , wherein the oligonucleotide comprises at least 5 deoxyribonucleotide units and is capable of recruiting an RNase.

5. The compound of claim 3 , wherein the oligonucleotide sequence comprises an LNA-containing sequence selected from:

(SEQ ID NO: 67)

LNA 1:

5′ AccAaaAGaaT 3′;

(SEQ ID NO: 68)

LNA 2:

5′ TggCcATcaaT 3′;

(SEQ ID NO: 69)

LNA 3:

5′ TagCAtActtA 3′;

(SEQ ID NO: 70)

LNA 4:

5′ CCAAAAGA 3′;

(SEQ ID NO: 71)

LNA 5:

5′ CATACTTA 3′;

(SEQ ID NO: 72)

LNA 6:

5′ CagaCaCGaCCaaAA 3′;

(SEQ ID NO: 73)

LNA 7:

5′ TAcTtaCTgaCagCC 3′;

(SEQ ID NO: 74)

LNA 8:

5′ AGACacgaccaAAAG 3′;

(SEQ ID NO: 75)

LNA 9:

5′ TACTtactgacaGCC 3′;

(SEQ ID NO: 76)

LNA9.2:

5′ TACttactgacAGCC 3′;

(SEQ ID NO: 77)

LNA10:

5′ACCaaaagAAT 3′;

(SEQ ID NO: 78)

LNA11:

5′ TGGccatcAAT 3′;

(SEQ ID NO: 79)

LNA12:

5′TAGcatacTTA 3′;

(SEQ ID NO: 80)

LNA13:

5′CgacCAaaAGaattC 3′;

(SEQ ID NO: 81)

LNA14:

5′CGACcaaaagaATTC 3′;

(SEQ ID NO: 82)

LNA15:

5′GaTGgCcATcaAttA 3′;

(SEQ ID NO: 83)

LNA16:

5′GATGgccatcaATTA 3′;

(SEQ ID NO: 84)

LNA17:

5′TcTAgCaTActTacT 3′;

(SEQ ID NO: 85)

LNA18:

5′TCTAgcatactTACT 3′;

(SEQ ID NO: 86)

LNA19:

5′GAAttcggatgGCCA 3′;

(SEQ ID NO: 87)

LNA20:

5′GGCCatcaattaGTG 3′;

(SEQ ID NO: 88)

LNA21:

5′TTCGgatggccaTCA 3′;

(SEQ ID NO: 89)

LNA22:

5′AGCCagacagCGA 3′;

(SEQ ID NO: 90)

LNA23:

5′GACAgccagacaGCA 3′;

(SEQ ID NO: 91)

LNA 9.G74C:

5′TACTtactgacaGTC 3′;

and

(SEQ ID NO: 92)

LNA 9.T80C:

5′TACTtaccgacaGCC 3′;

wherein capitalized letters denote LNA nucleotides and lowercase letters denote DNA nucleotides.

6. A method of inhibiting influenza A virus in a cell, the method comprising:

contacting a sample comprising viral RNA (vRNA) having a PSL2 motif with an effective amount of the oligonucleotide compound according to claim 1 .

7. The method of claim 6 , wherein contacting the sample with an agent results in at least 2 log 10 titer deficits of the virus or the agent disrupts the overall structure of the PSL2 motif of the vRNA.

8. The method of claim 6 , wherein the vRNA is present in a virion or a cell.

9. A method of treating or preventing influenza A virus infection in a subject, the method comprising:

administering to a subject in need thereof a pharmaceutical composition comprising an effective amount of the oligonucleotide compound according to claim 1 .

10. The method of claim 9 , wherein the subject is at risk of influenza A virus infection and the administering of the oligonucleotide compound protects the subject against infection for 1 week or more.

11. The method of claim 10 , wherein the administering comprises weekly, biweekly, or monthly administration of an effective dose of the oligonucleotide compound.

12. The method of claim 9 , wherein the pharmaceutical composition further comprises an additional active agent selected from a second oligonucleotide active agent and an antiviral drug.

13. The method of claim 9 , wherein the subject is diagnosed with or suspected of having an influenza A virus infection.

14. The compound of claim 1 , wherein the oligonucleotide sequence is selected from:

(SEQ ID NO:50)

5' CAGACACGACCAAAA 3'; 

(SEQ ID NO:51)

5' TACTTACTGACAGCC 3'; 

(SEQ ID NO:52)

5' AGACACGACCAAAAG 3'; 

(SEQ ID NO:56)

5' CGACCAAAAGAATTC 3'; 

(SEQ ID NO:58)

5' GATGGCCATCAATTA 3'; 

(SEQ ID NO:60)

5' TCTAGCATACTTACT 3'; 

(SEQ ID NO:63)

5' GGCCATCAATTAGTG 3'; 

(SEQ ID NO:64)

5' TTCGGATGGCCATCA 3'; 

(SEQ ID NO:66)

5' GACAGCCAGACAGCA 3'; 

(SEQ ID NO:91)

5' TACTTACTGACAGTC 3'; and 

(SEQ ID NO:92)

5' TACTTACCGACAGCC 3'.

15. An oligonucleotide compound or salt thereof, comprising an oligonucleotide sequence comprising at least 8 nucleoside subunits complementary to a region of a Packaging Stem-Loop 2 (PSL2) motif of a PB2 viral RNA (vRNA) or mutant thereof, wherein the oligonucleotide sequence is selected from:

(SEQ ID NO:51)

5' TACTTACTGACAGCC 3'; 

(SEQ ID NO:58)

5' GATGGCCATCAATTA 3'; 

(SEQ ID NO:60)

5' TCTAGCATACTTACT 3'; 

(SEQ ID NO:63)

5' GGCCATCAATTAGTG 3'; 

(SEQ ID NO:64)

5' TTCGGATGGCCATCA 3'; 

(SEQ ID NO:66)

5' GACAGCCAGACAGCA 3'; 

(SEQ ID NO:91)

5' TACTTACTGACAGTC 3'; and 

(SEQ ID NO:92)

5' TACTTACCGACAGCC 3'.

16. The method according to claim 6 , wherein the oligonucleotide sequence is selected from:

(SEQ ID NO:50)

5' CAGACACGACCAAAA 3'; 

(SEQ ID NO:51)

5' TACTTACTGACAGCC 3'; 

(SEQ ID NO:52)

5' AGACACGACCAAAAG 3'; 

(SEQ ID NO:56)

5' CGACCAAAAGAATTC 3'; 

(SEQ ID NO:58)

5' GATGGCCATCAATTA 3'; 

(SEQ ID NO:60)

5' TCTAGCATACTTACT 3'; 

(SEQ ID NO:63)

5' GGCCATCAATTAGTG 3'; 

(SEQ ID NO:64)

5' TTCGGATGGCCATCA 3'; 

(SEQ ID NO:66)

5' GACAGCCAGACAGCA 3'; 

(SEQ ID NO:91)

5' TACTTACTGACAGTC 3'; and 

(SEQ ID NO:92)

5' TACTTACCGACAGCC 3'.

17. The method according to claim 16 , wherein the oligonucleotide sequence is selected from:

(SEQ ID NO:51)

5' TACTTACTGACAGCC 3'; 

(SEQ ID NO:58)

5' GATGGCCATCAATTA 3'; 

(SEQ ID NO:60)

5' TCTAGCATACTTACT 3'; 

(SEQ ID NO:63)

5' GGCCATCAATTAGTG 3'; 

(SEQ ID NO:64)

5' TTCGGATGGCCATCA 3'; 

(SEQ ID NO:66)

5' GACAGCCAGACAGCA 3'; 

(SEQ ID NO:91)

5' TACTTACTGACAGTC 3'; and 

(SEQ ID NO:92)

5' TACTTACCGACAGCC 3'.

18. The method according to claim 9 , wherein the oligonucleotide sequence is selected from:

(SEQ ID NO:50)

5' CAGACACGACCAAAA 3'; 

(SEQ ID NO:51)

5' TACTTACTGACAGCC 3'; 

(SEQ ID NO:52)

5' AGACACGACCAAAAG 3'; 

(SEQ ID NO:56)

5' CGACCAAAAGAATTC 3'; 

(SEQ ID NO:58)

5' GATGGCCATCAATTA 3'; 

(SEQ ID NO:60)

5' TCTAGCATACTTACT 3'; 

(SEQ ID NO:63)

5' GGCCATCAATTAGTG 3'; 

(SEQ ID NO:64)

5' TTCGGATGGCCATCA 3'; 

(SEQ ID NO:66)

5' GACAGCCAGACAGCA 3'; 

(SEQ ID NO:91)

5' TACTTACTGACAGTC 3'; and 

(SEQ ID NO:92)

5' TACTTACCGACAGCC 3'.

19. The method according to claim 18 , wherein the oligonucleotide sequence is selected from:

(SEQ ID NO:51)

5' TACTTACTGACAGCC 3'; 

(SEQ ID NO:58)

5' GATGGCCATCAATTA 3'; 

(SEQ ID NO:60)

5' TCTAGCATACTTACT 3'; 

(SEQ ID NO:63)

5' GGCCATCAATTAGTG 3'; 

(SEQ ID NO:64)

5' TTCGGATGGCCATCA 3'; 

(SEQ ID NO:66)

5' GACAGCCAGACAGCA 3'; 

(SEQ ID NO:91)

5' TACTTACTGACAGTC 3'; and 

(SEQ ID NO:92)

5' TACTTACCGACAGCC 3'.