RNA interference mediated inhibition of adenosine A1 receptor (ADORA1) gene expression using short interfering RNA

Claim 2

1 . A short interfering RNA (siRNA) molecule that down regulates expression of an ADORA1 gene by RNA interference.

Claim 3

2 . The siRNA molecule of claim 1 , wherein said siRNA molecule is adapted for use to treat asthma.

Claim 4

3 . The siRNA molecule of claim 1 , wherein said siRNA molecule comprises a sense region and an antisense region and wherein said antisense region comprises sequence complementary to an RNA sequence encoding ADORA1 and the sense region comprises sequence complementary to the antisense region.

Claim 5

4 . The siRNA molecule of claim 3 , wherein said siRNA molecule is assembled from two nucleic acid fragments wherein one fragment comprises the sense region and the second fragment comprises the antisense region of said siRNA molecule.

Claim 6

5 . The siRNA molecule of claim 4 , wherein said sense region and antisense region are covalently connected via a linker molecule.

Claim 7

6 . The siRNA molecule of claim 5 , wherein said linker molecule is a polynucleotide linker.

Claim 8

7 . The siRNA molecule of claim 5 , wherein said linker molecule is a non-nucleotide linker.

Claim 9

8 . The siRNA molecule of claim 3 , wherein said antisense region comprises sequence complementary to sequence having any of SEQ ID NOs. 1-161.

Claim 10

9 . The siRNA molecule of claim 3 , wherein said antisense region comprises sequence having any of SEQ ID NOs. 162-322, 336, 338, 340, 342, 344, or 346.

Claim 11

10 . The siRNA molecule of claim 3 , wherein said sense region comprises sequence having any of SEQ ID NOs. 1-161, 335, 337, 339, 341, 343, or 345.

Claim 12

11 . The siRNA molecule of claim 3 , wherein said sense region comprises a sequence of SEQ ID NO. 323 and said antisense region comprises a sequence of SEQ ID NO. 324.

Claim 13

12 . The siRNA molecule of claim 3 , wherein said sense region comprises a sequence of SEQ ID NO. 325 and said antisense region comprises a sequence of SEQ ID NO. 326.

Claim 14

13 . The siRNA molecule of claim 3 , wherein said sense region comprises a sequence of SEQ ID NO. 327 and said antisense region comprises a sequence of SEQ ID NO. 328.

Claim 15

14 . The siRNA molecule of claim 3 , wherein said sense region comprises a sequence of SEQ ID NO. 329 and said antisense region comprises a sequence of SEQ ID NO. 330.

Claim 16

15 . The siRNA molecule of claim 3 , wherein said sense region comprises a sequence of SEQ ID NO. 331 and said antisense region comprises a sequence of SEQ ID NO. 332.

Claim 17

16 . The siRNA molecule of claim 3 , wherein said sense region comprises a sequence of SEQ ID NO. 333 and said antisense region comprises a sequence of SEQ ID NO. 334.

Claim 18

17 . The siRNA molecule of claim 3 , wherein said sense region comprises a 3′-terminal overhang and said antisense region comprises a 3′-terminal overhang.

Claim 19

18 . The siRNA molecule of claim 17 , wherein said 3′-terminal overhangs each comprise about 2 nucleotides.

Claim 20

19 . The siRNA molecule of claim 17 , wherein said antisense region 3′-terminal nucleotide overhang is complementary to RNA encoding ADORA1.

Claim 21

20 . The siRNA molecule of claim 3 , wherein said sense region comprises one or more 2′-O-methyl modified pyrimidine nucleotides.

Claim 22

21 . The siRNA molecule of claim 3 , wherein said sense region comprises a terminal cap moiety at the 5′-end, 3′-end, or both 5′ and 3′ ends of said sense region.

Claim 23

22 . The siRNA molecule of claim 3 , wherein said antisense region comprises one or more 2′-deoxy-2′-fluoro modified pyrimidine nucleotides.

Claim 24

23 . The siRNA molecule of claim 3 , wherein said antisense region comprises a phosphorothioate internucleotide linkage at the 3′ end of said antisense region.

Claim 25

24 . The siRNA molecule of claim 3 , wherein said antisense region comprises between about one and about five phosphorothioate internucleotide linkages at the 5′ end of said antisense region.

Claim 26

25 . The siRNA molecule of claim 17 , wherein said 3′-terminal nucleotide overhangs comprise ribonucleotides that are chemically modified at a nucleic acid sugar, base, or backbone.

Claim 27

26 . The siRNA molecule of claim 17 , wherein said 3′-terminal nucleotide overhangs comprise deoxyribonucleotides that are chemically modified at a nucleic acid sugar, base, or backbone.

Claim 28

27 . The siRNA molecule of claim 17 , wherein said 3′-terminal nucleotide overhangs comprise one or more universal base ribonucleotides.

Claim 29

28 . The siRNA molecule of claim 17 , wherein said 3′-terminal nucleotide overhangs comprise one or more acyclic nucleotides.

Claim 30

29 . The siRNA molecule of claim 17 , wherein said 3′-terminal nucleotide overhangs comprise nucleotides comprising internucleotide linkages having Formula I: wherein each R1 and R2 is independently any nucleotide, non-nucleotide, or polynucleotide which can be naturally occurring or chemically modified, each X and Y is independently O, S, N, alkyl, or substituted alkyl, each Z and W is independently O, S, N, alkyl, substituted alkyl, O-alkyl, S-alkyl, alkaryl, or aralkyl, and wherein W, X, Y and Z are not all O.

Claim 31

30 . The siRNA molecule of claim 17 , wherein said 3′-terminal nucleotide overhangs comprise nucleotides or non-nucleotides having Formula II: wherein each R3, R4, R5, R6, R7, R8, R10, R11 and R12 is independently H, OH, alkyl substituted alkyl, alkaryl or aralkyl, F, Cl, Br, CN, CF3, OCF3, OCN, O-alkyl, S-alkyl, N-alkyl, O-alkenyl, S-alkenyl, N-alkenyl, SO-alkyl, alkyl-OSH, alkyl-OH, O-alkyl-OH, O-alkyl-SH, S-alkyl-OH, S-alkyl-SH, alkyl-S-alkyl, alkyl-O-alkyl, ONO2, NO2, N3, NH2, aminoalkyl, aminoacid, aminoacyl, ONH2, O-aminoalkyl, O-aminoacid, O-aminoacyl, heterocycloalkyl, heterocycloalkaryl, aminoalkylamino, polyalklylamino, substituted silyl, or group having Formula I; R9 is O, S, CH2, S═O, CHF, or CF2, and B is a nucleosidic base or any other non-naturally occurring base that can be complementary or non-complementary to ADORA1 RNA or a non-nucleosidic base or any other non-naturally occurring universal base that can be complementary or non-complementary to ADORA1 RNA.

Claim 32

31 . An expression vector comprising a nucleic acid sequence encoding at least one siRNA molecule of claim 1 in a manner that allows expression of the nucleic acid molecule.

Claim 33

32 . A mammalian cell comprising an expression vector of claim 31 .

Claim 34

33 . The mammalian cell of claim 32 , wherein said mammalian cell is a human cell.

Claim 35

34 . The expression vector of claim 31 , wherein said siRNA molecule comprises a sense region and an antisense region and wherein said antisense region comprises sequence complementary to an RNA sequence encoding ADORA1 and the sense region comprises sequence complementary to the antisense region.

Claim 36

35 . The expression vector of claim 34 , wherein said siRNA molecule comprises two distinct strands having complementarity sense and antisense regions.

Claim 37

36 . The expression vector of claim 34 , wherein said siRNA molecule comprises a single strand having complementary sense and antisense regions.