RU2788030C2 - Mirna structures with high activity and reduced exposure outside the target - Google Patents

Abstract

FIELD: medicine.

SUBSTANCE: proposed group of inventions relates to medicine, namely to pharmaceuticals. An RNAi molecule has been proposed for downregulating target mRNA expression, consisting of a polynucleotide sense strand and a polynucleotide antisense strand. The molecule has a duplex region 15 to 30 nucleotides in length, in which one or more nucleotides in the antisense strand at positions 3 to 8 from the 5'-end of the duplex region are deoxynucleotides, where the 5'-end of the antisense strand corresponds to the 5'-terminus a duplex region where the antisense strand has deoxynucleotides at a plurality of positions in the primer region of said antisense strand. Proposed pharmaceutical composition for the treatment of malignant tumors, containing the specified molecule and a pharmaceutically acceptable carrier. Proposed cell for the expression of the RNAi molecule containing the specified molecule. A method for treating malignant tumors in a subject by means of gene silencing is proposed. Proposed is the use of a pharmaceutical composition for the treatment of malignant tumors in a subject and the use of the composition for the preparation or manufacture of a drug for the treatment of malignant tumors in a subject in need thereof.

EFFECT: proposed group of inventions provides an increase in gene knockdown activity through the use of an RNAi molecule in which one or more deoxynucleotides are located in the siRNA primer region.

27 cl, 15 dwg, 22 tbl, 22 ex

Description

TECHNICAL FIELD OF THE INVENTION

[0001] This invention relates to the field of biopharmaceutical and therapeutic agents, consisting of molecules based on nucleic acids. More specifically, this invention relates to the structures of compounds using RNA interference (RNAi) to modulate gene expression, and to their use.

SEQUENCE LIST

[0002] This application contains a sequence listing filed electronically as an ASCII file created on April 18, 2018 called Sequence Listings.txt, which is 183581 bytes in size and is hereby incorporated by reference in its entirety.

BACKGROUND OF THE INVENTION

[0003] RNA interference (RNAi) has been used in animals for post-transcriptional silencing of genes with a specific sequence mediated by small interfering RNAs (siRNAs). See, for example, Zamore et al., Cell, 2000, volume 101, pp. 25-33; Fire et al., Nature, 1998, volume 391, page 806811; Sharp, Genes & Development, 1999, Volume 13, pp. 139-141.

[0004] The RNAi response in cells can be triggered by double-stranded RNA (dsRNA), although the mechanism is not yet fully understood. In general, an siRNA can be about 21 to about 23 nucleotides in length and contain a base pair duplex region of about 19 nucleotides in length.

[0005] An endonuclease complex known as the RNA-induced silencing complex (RISC) is involved in RNAi. The siRNA has an antisense or guide strand that enters the RISC complex and mediates cleavage of a single-stranded target RNA having a sequence complementary to the antisense strand of the siRNA duplex. The other strand of miRNA is the accompanying strand. Cleavage of the target RNA occurs in the middle of the region complementary to the antisense strand of the miRNA duplex. See, for example, Elbashir et al., Genes & Development, 2001, Volume 15, pp. 188-200.

[0006] In order to improve the properties of siRNA compounds for use as drugs, the structure of siRNA compounds has been modified in various ways to improve stability and reduce off-target effects. Off-target siRNA activity involves the modulation of cellular nucleic acids other than the target RNA, which can occur through a variety of mechanisms.

[0007] Modifications of siRNA compounds include structural variations in their base, sugar, and/or backbone of any of the nucleotides. One way to modify miRNA nucleotides is to replace certain ribonucleotides in miRNA with deoxynucleotides. In particular, deoxynucleotides are often used at the cohesive ends or terminal nucleotides of an siRNA structure.

[0008] However, a disadvantage of using deoxynucleotides in an siRNA structure is that deoxynucleotides cannot be used in the siRNA primer region. This is due to the fact that such modification of miRNA is not favorable due to a decrease in the activity of gene silencing.

[0009] There is an urgent need for RNAi structures to modulate gene expression that reduce off-target activity without loss of activity.

[0010] In particular, RNAi-based therapeutics for the suppression of oncogenes and genes associated with malignant tumors require highly active and stable siRNA sequences and structures.

[0011] siRNA compounds with a highly active structure are needed to modulate gene expression.

SUMMARY

[0012] This invention relates to compounds, compositions and methods for modulating the expression of target genes using RNA interference. siRNA compounds and structures of this disclosure can be highly active in modulating gene expression.

[0013] The RNAi structures of this invention can be used to modulate gene expression with surprisingly high levels of activity and reduced off-target activity.

[0014] In some embodiments, the present invention provides gene silencing molecules via RNAi that reduce off-target activity without loss of activity.

[0015] In additional embodiments, the present invention provides gene silencing molecules via RNAi that reduce off-target activity with surprisingly high gene silencing activity.

[0016] The RNAi molecules of this invention can provide highly active and stable siRNA structures that can be used for gene suppression RNAi therapeutics.

[0017] In additional embodiments, the structures, molecules, and compositions of this invention can be used in methods for preventing or treating diseases or improving the symptoms of conditions or disorders associated with one or more genes.

[0018] Embodiments of this invention include the following:

[0019] A nucleic acid molecule, where:

a) the molecule has a polynucleotide sense strand and a polynucleotide antisense strand;

b) each strand of the molecule is 15 to 30 nucleotides in length;

c) a contiguous region of 15 to 30 nucleotides of the antisense strand is complementary to the mRNA sequence;

d) at least a portion of the sense strand is complementary to at least a portion of the antisense strand and the molecule has a duplex region of 15 to 30 nucleotides in length, wherein one or more nucleotides in the duplex region at positions 3 to 8 from the 5' end of the antisense strand are deoxynucleotides. The mRNA may be human mRNA.

[0020] In some embodiments, the antisense strand of a nucleic acid molecule may have deoxynucleotides in a plurality of positions, the plurality of positions being one of the following:

each of positions 4, 6 and 8 from the 5' end of the antisense strand;

each of positions 3, 5 and 7 from the 5' end of the antisense strand;

each of positions 1, 3, 5 and 7 from the 5' end of the antisense strand;

each of positions 3-8 from the 5' end of the antisense strand; and

each of positions 5-8 from the 5' end of the antisense strand.

[0021] Nucleic acid molecules can be RNAi molecules that are active in modulating mRNA expression.

[0022] In certain embodiments, nucleic acid molecules can be active in inhibiting the expression of a gene selected from protein-coding genes, proto-oncogenes, oncogenes, tumor suppressor genes, and cell signaling genes.

[0023] In additional embodiments, the mRNA may be human mRNA expressing any element or subelement of the human protein family, including SRY, β-globin, RAS, cytosolic GST, mitochondrial GST, MAPEG GST, GST-π, p16, p21, p53 , serum albumin, type VII collagen, complement C3, apolipoprotein B, phenylalanine hydroxylase, factor VIII, huntingtin, retinoblastoma protein RB1, CFTR, titin, utrophin, and dystrophin.

[0024] Nucleic acid molecules may have an IC ₅₀ for mRNA knockdown of less than 100 pM, or less than 50 pM, or less than 10 pM. In certain embodiments, nucleic acid molecules can inhibit mRNA by at least 25% in vivo after a single administration of the molecules.

[0025] In some embodiments, each strand of the molecule is 18 to 22 nucleotides in length. The duplex region can be 19 nucleotides in length. In certain embodiments, the polynucleotide sense strand and the polynucleotide antisense strand can be joined as one strand and form a duplex region connected at one end with a loop.

[0026] Nucleic acid molecules may have a blunt end. In certain embodiments, nucleic acid molecules may have one or more 3' sticky ends.

[0027] Embodiments of this invention include nucleic acid molecules having one or more nucleotides in the duplex region that are modified or chemically modified. Modified or chemically modified nucleotides may be 2'-O-alkyl-substituted nucleotides, 2'-deoxy-2'-fluoro-substituted nucleotides, phosphothioate nucleotides, closed nucleotides, or any combination thereof.

[0028] This invention further provides pharmaceutical compositions comprising nucleic acid molecules and a pharmaceutically acceptable carrier. The carrier may be a lipid molecule or a liposome.

[0029] Embodiments of this invention further include a vector or cell containing nucleic acid molecules.

[0030] This invention also provides methods for preventing, treating, or improving a disease in a subject in need of gene silencing by administering to the subject a composition comprising nucleic acid molecules. The disease may be a malignant tumor, sarcoma or carcinoma.

[0031] Embodiments of this invention include the use of a composition of nucleic acid molecules to prevent, improve or treat a disease or condition in a subject in need thereof.

[0032] In certain embodiments, the implementation of the composition according to this invention may be for use in medical therapy.

[0033] In additional embodiments, the implementation of the composition according to this invention may be for use in the treatment of the human or animal body.

[0034] In additional embodiments, a composition of this invention may be for the preparation or manufacture of a medicament for the prevention, amelioration, or treatment of a disease or condition in a subject in need thereof.

BRIEF DESCRIPTION OF THE FIGURES

[0035] FIG. 1: in Fig. 1 shows a marked reduction in orthotopic tumors from lung cancer in vivo with siRNAs of the invention directed to GST-π. GST-π siRNA was administered in a liposomal formulation at a dose of 2 mg/kg to athymic nude mice bearing orthotopic tumors from A549 lung cancer. Final primary tumor masses were measured at autopsy for the treatment group and the vehicle control group. GST-π siRNA showed a significant tumor-inhibiting effect from lung cancer in this 6-week study. As shown in FIG. 1, after 43 days, GST-π siRNA showed markedly favorable tumor inhibition, with a significantly reduced final average mass of primary tumors by 2.8 times compared to control.

[0036] FIG. 2: in Fig. 2 shows the in vivo tumor inhibition effect of GST-π siRNA. A cancer xenograft model using A549 cells was used at a relatively low siRNA dose of 0.75 mg/kg. GST-π siRNA showed favorable tumor inhibition within a few days. After 36 days, GST-π siRNA showed markedly favorable tumor inhibition, with a significantly reduced final mean tumor volumes of about 2-fold compared to control.

[0037] FIG. 3: in FIG. 3 shows the in vivo tumor inhibition effect of GST-π siRNA at the endpoint of FIG. 2. GST-π siRNA showed favorable tumor inhibition with more than 2-fold reduced average tumor masses.

[0038] FIG. 4: in FIG. 4 shows that the GST-π siRNA of the present invention significantly increased cancer cell death through in vitro apoptosis. GST-π siRNA caused upregulation of PUMA, a biological marker of apoptosis that is associated with cell viability loss. In FIG. 4, PUMA expression is significantly increased from days 2-6 post transfection with GST-π siRNA.

[0039] FIG. 5: in FIG. 5 shows that the GST-π siRNA of the invention provided a knockdown effect for A549 xenograft tumors in vivo. A dose-dependent knockdown of GST-π mRNA was observed in female athymic nude (nu/nu) mice (Charles River) using GST-π-targeted siRNA. As shown in FIG. 5, at a dose of 4 mg/kg, a significant decrease in GST-π mRNA was found by approximately 40% 24 hours after injection.

[0040] FIG. 6: in FIG. 6 shows that the GST-π siRNA of the invention inhibited xenograft tumors from pancreatic cancer in vivo. GST-π siRNA conferred gene silencing activity in vivo when administered as a liposomal formulation into pancreatic cancer xenograft tumors in female athymic nude mice aged 6 to 8 weeks. As shown in FIG. 6, dose effects were achieved using doses ranging from 0.375 mg/kg to 3 mg/kg of siRNA directed to GST-π. GST-π siRNA showed favorable tumor inhibition within a few days after injection, at the end point there was a decrease in tumor volume by approximately 2 times.

[0041] FIG. 7: in FIG. 7 shows that the GST-π siRNA of the invention showed improved serum stability. As shown in FIG. 7, the serum half-life (t _1/2 ) for both the sense strand (FIG. 7, top) and antisense strand (FIG. 7, bottom) of GST-π siRNA was approximately 100 minutes.

[0042] FIG. 8: in FIG. 8 shows that the GST-π siRNA of the present invention showed increased formulation stability in plasma. In FIG. 8 shows the incubation of the liposome composition of GST-π siRNA in 50% human serum in PBS and the detection of the remaining siRNA at various time points. As shown in FIG. 8, the plasma half-life (t _1/2 ) of the GST-π siRNA formulation significantly exceeded 100 hours.

[0043] FIG. 9: in FIG. 9 shows in vitro knockdown for the GST-π siRNA guide strand. As shown in FIG. 9, knockdown of the GST-π siRNA guide strand was approximately exponential compared to the shuffled sequence control, which showed no effect.

[0044] FIG. 10: in fig. 10 shows the in vitro knockdown for the GST-π siRNA follower strand of FIG. 9. As shown in FIG. 10, off-target knockdown of the GST-π siRNA follower strand is significantly reduced and essentially has no effect.

[0045] FIG. 11: in Fig. 11 shows in vitro knockdown for guide strands of several highly active GST-π siRNAs. As shown in FIG. 11, GST-π siRNA guide strand knockdown activities were approximately exponential.

[0046] FIG. 12: in FIG. 12 shows the in vitro knockdown for the follower strand from the GST-π siRNA of FIG. 11. As shown in FIG. 12, out-of-target follower strand knockdown activities for GST-π siRNA are significantly reduced, below about 500 pM.

[0047] FIG. 13: in FIG. 13 shows the in vitro guide strand knockdown of a highly active GST-π siRNA. As shown in FIG. 13, guide strand knockdown activity from GST-π siRNA was approximately exponential.

[0048] FIG. 14: in FIG. 14 shows the in vitro knockdown for the GST-π siRNA follower strand of FIG. 13. As shown in FIG. 14, out-of-target follower strand knockdown activity for GST-π siRNA is significantly reduced.

[0049] FIG. 15: in FIG. 15 shows the in vivo tumor inhibition effect of p21 siRNA. A cancer xenograft model using A549 cells was used at a relatively low siRNA dose of 0.75 mg/kg. p21 siRNA showed favorable tumor inhibition within a few days. After 30 days, GST-π siRNA showed markedly favorable tumor inhibition with significantly reduced final average tumor volumes by more than 2-fold compared to control.

DETAILED DESCRIPTION OF THE INVENTION

[0050] This invention relates to compounds, compositions and methods for nucleic acid therapeutics for modulating gene expression.

[0051] In some embodiments, the present invention provides RNAi-active molecules, as well as structures and compositions, that can provide silencing of gene expression.

[0052] The structures and compositions of this disclosure can be used in the prevention or treatment of various diseases.

[0053] In additional embodiments, this invention provides compositions for the delivery and capture of one or more therapeutic RNAi molecules according to this invention, as well as methods of using them. The RNA-based compositions of this invention can be used in methods for the prevention or treatment of cancers, such as cancers.

[0054] Therapeutic compositions of this invention include nucleic acid molecules that are active in RNA interference. Therapeutic nucleic acid molecules can be directed to different genes for gene silencing.

[0055] In various embodiments, this invention provides a number of molecules that can be active as small interfering RNA (siRNA) and can regulate or provide silencing gene expression.

[0056] Embodiments of the invention further provide a carrier, formulation, or formulation of lipid nanoparticles for delivering inventive siRNA to subjects in need of disease prevention or treatment. This invention further provides methods for administering siRNAs as therapeutic agents to mammals.

[0057] The therapeutic molecules and compositions of this invention can be used for RNA interference aimed at preventing or treating a disease by administering a compound or composition to a subject in need thereof.

[0058] This invention provides a number of RNAi molecules, where each molecule has a polynucleotide sense strand and a polynucleotide antisense strand; each strand of the molecule is 15 to 30 nucleotides in length; a contiguous region of 15 to 30 nucleotides of the antisense strand is at least partially complementary to the mRNA sequence; and at least a portion of the sense strand is complementary to at least a portion of the antisense strand, and the molecule has a duplex region of 15 to 30 nucleotides in length.

[0059] An RNAi molecule of the invention may have a contiguous region of 15 to 30 nucleotides of the antisense strand complementary to a human mRNA sequence that is located in the duplex region of the molecule.

[0060] In some embodiments, the RNAi molecule may have a contiguous region of 15 to 30 nucleotides of an antisense strand that is complementary to a human mRNA sequence.

[0061] Embodiments of the present invention may further provide methods for preventing, treating, or improving one or more symptoms of a disease or condition, or reducing the risk of developing a disease or condition, or delaying the onset of a disease or condition in a mammal in need thereof.

[0062] RNAi structures for universal genomic silencing

[0063] Embodiments of the present invention may provide RNAi molecules directed to any gene that exhibit high gene expression silencing activity.

[0064] The RNAi molecules of this disclosure can exhibit surprisingly high gene expression silencing activity while providing reduced off-target effects.

[0065] In some embodiments, the implementation of the RNAi molecule according to this invention contains one or more 2'-deoxynucleotides. In these embodiments, one or more 2'-deoxynucleotides can be located in the primer region of the siRNA.

[0066] One or more of the 2'-deoxynucleotides may be mono-deoxynucleotides.

[0067] As used herein, a deoxynucleotide refers to a mono-2'-deoxynucleotide.

[0068] The 2'-deoxynucleotide can be substituted at the 2'-position with halogen.

[0069] In certain embodiments, the RNAi molecule of this invention contains one or more 2'-deoxynucleotides in the antisense or guide strand of the siRNA. More specifically, one or more deoxynucleotides can be located at positions 1 to 8 from the 5' end of the siRNA antisense strand. In certain embodiments, one or more deoxynucleotides can be located at positions 2 to 8 from the 5' end of the siRNA antisense strand. In additional embodiments, one or more deoxynucleotides may be located at positions 3 to 8 from the 5' end of the antisense siRNA strand.

[0070] This invention provides siRNA structures that can have an antisense strand containing deoxynucleotides in multiple positions.

[0071] In some embodiments, the siRNA structure may have an antisense strand containing deoxynucleotides at each of positions 4, 6, and 8 from the 5' end of the antisense strand.

[0072] In further embodiments, the siRNA structure may have an antisense strand containing deoxynucleotides at each of positions 3, 5, and 7 from the 5' end of the antisense strand.

[0073] In further embodiments, the siRNA structure may have an antisense strand containing deoxynucleotides at each of positions 1, 3, 5, and 7 from the 5' end of the antisense strand.

[0074] In certain embodiments, the siRNA structure may have an antisense strand containing deoxynucleotides at each of positions 3 to 8 from the 5' end of the antisense strand.

[0075] In some embodiments, the siRNA structure may have an antisense strand containing deoxynucleotides at each of positions 5 to 8 from the 5' end of the antisense strand.

[0076] Any of these structures can be combined with one or more modified or chemically modified nucleotides at other positions.

[0077] The RNAi molecules of the invention can inhibit mRNA expression of a gene with a favorable IC _{50 of} less than about 200 pM. In certain embodiments, RNAi molecules of the invention can inhibit expression of an mRNA gene with a favorable IC _{50 of} less than about 100 pM, or less than about 50 pM, or less than about 30 pM, or less than about 20 pM, or less than about 10 pM, or less than about 5 pM, or less than about 1 pM.

[0078] In additional embodiments, the implementation of RNAi molecules according to this invention can inhibit the expression of mRNA gene at a certain level, at least 25% in vivo, after a single injection.

[0079] The siRNA of the invention can be directed to any gene.

[0080] Examples of genes to which siRNAs of the present invention can be targeted include those listed in the Gene Families Index of the HUGO Gene Nomenclature Committee.

[0081] Examples of genes to which siRNAs of the invention can be targeted include nuclear genes, mitochondrial genes, protein coding genes, proto-oncogenes, oncogenes, tumor suppressor genes, and cell signaling genes.

[0082] Examples of genes to which siRNAs of this invention can be directed include genes expressing any element or subelement of the human protein family, including SRY, β-globin, RAS, cytosolic GST, mitochondrial GST, MAPEG GST, GST-π, p16 , p21, p53, serum albumin, type VII collagen, complement C3, apolipoprotein B, phenylalanine hydroxylase, factor VIII, huntingtin, retinoblastoma protein RB1, CFTR, titin, utrophin, and dystrophin.

[0083] Modified and chemically modified siRNA

[0084] Embodiments of this invention encompass siRNA molecules that are modified or chemically modified to provide enhanced properties for therapeutic use, such as increased gene silencing activity. The present invention provides modified or chemically modified siRNA molecules that can have increased serum stability as well as reduced off-target effects without loss of siRNA molecule activity during gene modulation and gene silencing. In some aspects, the invention provides an siRNA that has modifications or chemical modifications in various combinations that enhance the stability and effect of the siRNA.

[0085] As used herein, the terms modified and chemically modified refer to changes made to the structure of a naturally occurring nucleotide or nucleic acid structure in miRNAs, which include miRNAs having one or more nucleotide analogs, altered nucleotides, non-standard nucleotides, non-naturally occurring nucleotides and their combinations.

[0086] In some embodiments, the implementation of the number of modified or chemically modified structures in the siRNA may include all structural components and/or all nucleotides of the siRNA molecule.

[0087] Examples of modified and chemically modified siRNA include an siRNA having a modification of the sugar group of a nucleotide, a modification of the nucleic base of a nucleotide, a modification of a backbone or bond in a nucleic acid, a modification of the structure of a nucleotide or nucleotides at the end of a siRNA strand, and combinations thereof.

[0088] Examples of modified and chemically modified siRNA include siRNA having a substituent modification on the 2' carbon of the sugar.

[0089] Examples of modified and chemically modified siRNA include siRNA having a modification at the 5' end, 3' end, or both ends of the strand.

[0090] Examples of modified and chemically modified siRNA include siRNA having modifications that create complementarity mismatches between strands.

[0091] Examples of modified and chemically modified siRNA include siRNA having a 5' propylamine terminal, a 5' phosphorylated terminal, a 3' puromycin terminal, or a 3' biotin terminal group.

[0092] Examples of modified and chemically modified siRNA include siRNA having a 2'-fluoro-substituted ribonucleotide, 2'-OMe-substituted ribonucleotide, 2'-deoxyribonucleotide, 2'-amino-substituted ribonucleotide, 2'-thio-substituted ribonucleotide.

[0093] Examples of modified and chemically modified siRNA include siRNA having one or more 5-halogenouridines, 5-halocytidines, 5-methylcytidines, ribothymidines, 2-aminopurines, 2,6-diaminopurines, 4-thiouridines, or 5-aminoallyluridines.

[0094] Examples of modified and chemically modified siRNA include siRNA having one or more phosphothioate groups.

[0095] Examples of modified and chemically modified siRNA include siRNA having one or more 2'-fluoro-substituted ribonucleotides, 2'-fluorouridines, 2'-fluorocytidines, 2'-deoxyribonucleotides, 2'-deoxyadenosines, or 2'-deoxyguanosines.

[0096] Examples of modified and chemically modified siRNA include siRNA having one or more phosphothioate linkages.

[0097] Examples of modified and chemically modified siRNA include siRNA having one or more alkylene diol bonds, oxyalkylthio bonds, or oxycarbonyloxy bonds.

[0098] Examples of modified and chemically modified siRNA include siRNA having one or more deoxyabasic groups, inosines, N3-methyluridines, N6,N6-dimethyladenosines, pseudouridines, purine ribonucleosides, and ribavirins.

[0099] Examples of modified and chemically modified siRNA include siRNA having one or more 3' or 5' inverted end groups.

[00100] Examples of modified and chemically modified siRNA include siRNA having one or more 5-(2-amino)propyluridines, 5-bromouridines, adenosines, 8-bromoguanosines, 7-deazaadenosines, or N6-methyladenosine.

[00101] GST-π and RNAi molecules

[00102] Various human cancer tissues have been found to correlate with the occurrence of the mutated KRAS gene. In some cases, the tissues also present an increased level of expression of glutathione-S-transferase pi (GST-π). (Miyanishi et al., Gastroenterology, 2001, volume 121:865-874, abstract) For example, elevated serum levels of GST-π have been observed in patients with various gastrointestinal malignancies. (Niitsu et al., Cancer, 1989, vol. 63, no. 2, pp. 317-323, abstract)

[00103] GST-π is a member of the GST family of enzymes that play a role in detoxification by catalyzing the conjugation of hydrophobic and electrophilic compounds to reduced glutathione. The expression of GST-π can be reduced in vitro using siRNA. (Niitsu et al., US 2014/0315975 A1). However, there are many disadvantages of existing siRNA agents such as lack of activity, off-target effects, lack of serum stability, and lack of in vivo activity or effect.

[00104] The nucleic acid sequence of the exemplary target human glutathione-S-transferase pi (human GST-π) mRNA is disclosed under Genbank Accession Number NM_000852.3 (hGSTP1) and is 986 nucleotides in length.

[00105] One skilled in the art will appreciate that the sequence shown may change over time and will include any necessary changes to the nucleic acid molecules herein as appropriate.

[00106] Embodiments of the invention may provide compositions and methods for silencing GST-π gene expression using small nucleic acid molecules. Examples of nucleic acid molecules include RNA interference active molecules (RNAi molecules), small interfering RNA (siRNA), double-stranded RNA (dsRNA), microRNA (miRNA), and short hairpin RNA (shRNA) molecules, as well as RNA directed by DNA (ddRNA), RNA interacting with Piwi (piRNA), and associated with miRNA repeats (rasiRNA). Such molecules are capable of mediating RNA interference against GST-π gene expression.

[00107] The composition and methods disclosed herein can also be used in the treatment of various types of cancer in a subject.

[00108] The nucleic acid molecules and methods of this invention can be used to down-regulate the expression of genes that encode GST-π.

[00109] The compositions and methods of this invention may include one or more nucleic acid molecules that, independently or in combination, can modulate or regulate the expression of a GST-π protein and/or genes encoding GST-π proteins, proteins and/or genes. encoding GST-π that are associated with the maintenance and/or development of diseases, conditions or disorders associated with GST-π, such as cancer.

[00110] The compositions and methods of this invention are described with reference to exemplary GST-π sequences. One skilled in the art will appreciate that various aspects and embodiments of the invention are directed to any related GST-π genes, sequences or variants, such as homologous genes and transcriptional variants, as well as polymorphisms, including single nucleotide polymorphism (SNP) associated with any GST-π genes.

[00111] In some embodiments, the compositions and methods of this invention may provide a double-stranded small interfering nucleic acid (siRNA) molecule that down-regulates the expression of a GST-π gene, such as human GST-π.

[00112] An RNAi molecule of the invention can be targeted to GST-π and any homologous sequences, e.g. using complementary sequences or by inserting non-canonical base pairs, e.g. mismatches and/or ambiguous base pairs, which may provide additional target sequences .

[00113] In cases where mismatches are identified, non-canonical base pairs, such as mismatches and/or ambiguous bases, can be used to generate nucleic acid molecules that target more than one gene sequence.

[00114] For example, non-canonical base pairs, such as UU and CC base pairs, can be used to design nucleic acid molecules that are capable of targeting sequences for different GST-π targets that share sequence homology. Thus, an RNAi molecule can be directed to a nucleotide sequence that is conserved between homologous genes, and a single RNAi molecule can be used to inhibit the expression of more than one gene.

[00115] In some aspects, the compositions and methods of this invention include RNAi molecules that are active against GST-π mRNA, where the RNAi molecule contains a sequence complementary to any mRNA encoding the GST-π sequence.

[00116] In some embodiments, an RNAi molecule of this disclosure may have activity against a GST-π RNA, wherein the RNAi molecule contains a sequence complementary to an RNA having a GST-π coding sequence variant, such as a mutant GST-π gene that, as known in the art to be associated with cancer.

[00117] In further embodiments, an RNAi molecule of the invention may comprise a nucleotide sequence that can interact with a GST-π gene nucleotide sequence and mediate silencing of GST-π gene expression.

[00118] Nucleic acid molecules for inhibiting GST-π expression may have a sense strand and an antisense strand in which the strands form a duplex region. Nucleic acid molecules may have one or more nucleotides in the duplex region that are modified or chemically modified, including such modifications as are known in the art. Any nucleotide at the sticky end of an siRNA can also be modified or chemically modified.

[00119] In some embodiments, the preferred modified or chemically modified nucleotides are 2'-deoxynucleotides. In additional embodiments, the modified or chemically modified nucleotides may include 2'-O-alkyl-substituted nucleotides, 2'-deoxy-2'-fluoro-substituted nucleotides, phosphothioate nucleotides, closed nucleotides, or any combination thereof.

[00120] In certain embodiments, a preferred structure may have an antisense strand containing deoxynucleotides in a plurality of positions, the plurality of positions being one of the following: each of positions 4, 6, and 8 from the 5' end of the antisense strand; each of positions 3, 5 and 7 from the 5' end of the antisense strand; each of positions 1, 3, 5 and 7 from the 5' end of the antisense strand; each of positions 3-8 from the 5' end of the antisense strand; and each of positions 5-8 from the 5' end of the antisense strand. Any of these structures can be combined with one or more 2'-deoxy-2'-fluoro-substituted nucleotides in the duplex region.

[00121] The nucleic acid molecules of the invention can inhibit the expression of GST-π mRNA with a favorable IC _{50 of} less than about 200 pM. In addition, nucleic acid molecules can inhibit the expression of GST-π mRNA at certain levels, at least 25% in vivo after a single administration.

[00122] This invention provides pharmaceutical compositions that may contain one or more miRNAs, as disclosed in the present description, in combination with a pharmaceutically acceptable carrier. Any suitable carrier can be used, including those known in the art, as well as lipid molecules, nanoparticles, or liposomes, any of which can encapsulate siRNA molecules.

[00123] Disclosed herein are methods for treating a disease associated with GST-π expression, which methods include administering to a subject in need thereof a composition comprising one or more siRNAs. The diseases to be treated may include cancer, cancer, cancer caused by cells expressing mutant KRAS, sarcoma, and carcinoma, among others.

[00124] Examples of RNAi molecules of the invention targeting GST-π mRNA are shown in Table 1.

Table 1: RNAi sequences of molecules for GST-π

ID Reference position SEQID No. MEANING CHAIN
(5'-->3')
SEQ ID ## from 1 to 65 SEQID No. ANTISENSE CIRCUIT
(5'-->3')
SEQ ID ## from 66 to 130 A1 652 1 UCCCAGAACCAGGGAGGCAtt 66 UGCCUCCCUGGUUCUGGGAca A10 635 2 CUUUUGAGACCCUGCUGUCtt 67 GACAGCAGGGUCUCAAAAGgc A11 649 3 CUGUCCCAGAACCAGGGAGtt 68 CUCCCUGGUUCUGGGACAGca A12 650 four UGUCCCAGAACCAGGGAGGtt 69 CCUCCCUGGUUCUGGGACAgc A13 631 five AAGCCUUUUGAGACCCUGCtt 70 GCAGGGUCUCAAAAGGCUUca A14 638 6 UUGAGACCCUGCUGUCCCAtt 71 UGGGACAGCAGGGUCUCAAAaa A15 636 7 UUUUGAGACCCUGCUGUCCtt 72 GGACAGCAGGGUCUCAAAAgg A16 640 8 GAGACCCUGCUGUCCCAGAtt 73 UCUGGGACAGCAGGGUCUCaa A17 332 nine GCUGGAAGGAGGAGGUGGUtt 74 ACCACCUCCUCCUUCCAGCtc A18 333 10 CUGGAAGGAGGAGGUGGUGtt 75 CACCACCUCCUCCUUCCAGct A19 321 eleven UCAGGGCCAGAGCUGGAAGtt 76 CUUCCAGCUCUGGCCCUGAtc A2 639 12 UGAGACCCUGCUGUCCCAGtt 77 CUGGGACAGCAGGGUCUCAaa A20 323 13 AGGGCCAGAGCUGGAAGGAtt 78 UCCUUCCAGCUCUGGCCCUga A21 331 fourteen AGCUGGAAGGAGGAGGUGGtt 79 CCACCUCCUCCUUCCAGCUct A22 641 15 AGACCCUGCUGUCCCAGAAtt 80 UUCUGGGACAGCAGGGUCUca A23 330 sixteen GAGCUGGAAGGAGGAGGUGtt 81 CACCUCCUCCUUCCAGCUCtg A25 647 17 UGCUGUCCCAGAACCAGGGtt 82 CCCUGGUUCUGGGACAGCAgg A26 653 eighteen CCCAGAACCAGGGAGGCAAtt 83 UUGCCUCCCUGGUUCUGGac A3 654 nineteen CCAGAACCAGGGAGGCAAGtt 84 CUUGCCUCCCUGGUUCUGGga A4 637 20 UUUGAGACCCUGCUGUCCCCtt 85 GGGACAGCAGGGUCUCAAAag A5 642 21 GACCCUGCUGUCCCAGAACtt 86 GUUCUGGGACAGCAGGGUCtc A6 319 22 GAUCAGGGCCAGAGCUGGAtt 87 UCCAGCUCUGGCCCUGAUCtg A7 632 23 AGCCUUUUGAGACCCUGCUtt 88 AGCAGGGUCUCAAAAGGCUtc A8 633 24 GCCUUUUGAGACCCUGCUGtt 89 CAGCAGGGUCUCAAAAGGCtt A9 634 25 CCUUUUGAGACCCUGCUGUtt 90 ACAGCAGGGUCUCAAAAGGct AG7 632 26 CGCCUUUUGAGACCCUGCAtt 91 UGCAGGGUCUCAAAAGGCGtc AK1 257 27 CCUACACCGUGGUCUAUUUtt 92 AAAUAGACCACGGUGUAGGgc AK10 681 28 UGUGGGAGACCAGAUCUCCtt 93 GGAGAUCUGGUCUCCCACAat AK11 901 29 GCGGGAGGCAGAGUUUGCCtt 94 GGCAAACUCUGCCUCCCGCtc AK12 922 thirty CCUUUCUCCAGGACCAAUAtt 95 UAUUGGUCCCUGGAGAAAGGaa AK13/A24 643 31 ACCCUGCUGUCCCAGAACCtt 96 GGUUCUGGGACAGCAGGGUct AK2 267 32 GGUCUAUUUCCCAGUUCGAtt 97 UCGAACUGGGAAAUAGACCac AK3 512 33 CCCUGGUGGACAUGGUGAAtt 98 UUCACCAUCCACCAGGGct AK4 560 34 ACAUCUCCCUCAUCUACACtt 99 GUGUAGAUGAGGGAGAUGUat AK5 593 35 GCAAGGAUGACUAUGUGAAtt one hundred UUCACAUAGUCAUCCUGCcc AK6 698 36 CCUUCGCUGACUACAACCUtt 101 AGGUUGUAGUCAGCGAAGGag AK7 313 37 CUGGCAGAUCAGGGCCAGAtt 102 UCUGGCCCUGAUCUGCCAGca AK8 421 38 GACGGAGACCCUCACCCUGUtt 103 ACAGGGUGAGGUCUCCGUCct AK9 590 39 CGGGCAAGGAUGACUAUGUtt 104 ACAUAGUCAUCCUUGCCCGcc AU10 635 40 CUUUUGAGACCCUGCUGUATtt 105 UACAGCAGGGUCUCAAAAGgc AU23 330 41 GAGCUGGAAGGAGGAGGUATtt 106 UACCUCCUCCUUCCAGCUCtg AU24 643 42 ACCCUGCUGUCCCAGAACATt 107 UGUUCUGGGACAGCAGGGUct AU25 648 43 UGCUGUCCCAGAACCAGGAtt 108 UCCUGGUUCUGGGACAGCAgg AU7 632 44 AGCCUUUUGAGACCCUGCAtt 109 UGCAGGGUCUCAAAAGGCUtc AU9 634 45 CCUUUUGAGACCCUGCUGAtt 110 UCAGCAGGGUCUCAAAAGGct B1 629 46 UGAAGCCUUUUGAGACCCUtt 111 AGGGUCUCAAAAGGCUUCAgt B10 627 47 ACUGAAGCCUUUUGAGACCtt 112 GGUCUCAAAAGGCUUCAGUtg B11 596 48 AGGAUGACUAUGUGAAGGCtt 113 GCCUUCACAUAGUCAUCCUtg B12 597 49 GGAUGACUAUGUGAAGGCAtt 114 UGCCUUCACAUAGUCAUCCtt B13 598 50 GAUGACUAUGUGAAGGCACtt 115 GUGCCUUCACAUAGUCAUCct B14 564 51 CUCCCUCAUCUACACCAACtt 116 GUUGGUGUAGAUGAGGGAGat B2 630 52 GAAGCCUUUUGAGACCCUGtt 117 CAGGGUCUCAAAAGGCUUCag B3 563 53 UCUCCCUCAUCUACACCAAtt 118 UUGGUGUAGAUGAGGGAGAtg B4 567 54 CCUCAUCUACACCAACUAUtt 119 AUAGUUGGUGUAGAUGAGGga B5 566 55 CCCUCAUCUACACCAACUAtt 120 UAGUUGGUGUAGAUGAGGag B6 625 56 CAACUGAAGCCUUUUGAGAtt 121 UCUCAAAAGGCUUCAGUUGcc B7 626 57 AACUGAAGCCUUUUGAGACtt 122 GUCUCAAAAGGCUUCAGUUgc B8 628 58 CUGAAGCCUUUUGAGACCCCtt 123 GGGUCUCAAAAGGCUUCAGtt B9 565 59 UCCCUCAUCUACACCAACUtt 124 AGUUGGUGUAGAUGAGGGAga BG3 563 60 GCUCCCUCAUCUACACCAAtt 125 UUGGUGUAGAUGAGGGAGCtg BU2 630 61 GAAGCCUUUUGAGACCCUAtt 126 UAGGGUCUCAAAAGGCUUCag BU10 627 62 ACUGAAGCCUUUUGAGACAtt 127 UGUCUCAAAAGGCUUCAGUtg BU14 565 63 CUCCCUCAUCUACACCAAAtt 128 UUUGGUGUAGAUGAGGGAGat BU4 567 64 CCUCAUCUACACCAACUAAtt 129 UUAGUUGGUGUAGAUGAGGga C1-934 934 65 ACCAUAAAAUUUCUAGAtt 130 UCUUAGAAAUUUUAUUGGUcc

[00125] Key to Table 1: A, G, C, and U in uppercase refer to ribo-A, ribo-G, ribo-C, and ribo-U, respectively. The lowercase letters a, u, g, c, t refer to 2'-deoxy-A, 2'-deoxy-U, 2'-deoxy-G, 2'-deoxy-C, and deoxythymidine, respectively.

[00126] Examples of RNAi molecules of the invention targeting GST-π mRNA are shown in Table 2.

Table 2: RNAi sequences of molecules for GST-π

ID SEQID No. MEANING CHAIN
(5'-->3')
SEQ ID ## 131 to 156 SEQID No. ANTISENSE CIRCUIT
(5'-->3')
SEQ ID ## 157 to 182 BU2' 131 GAAGCCUUUUGAGACCCUANN 157 UAGGGUCUCAAAAGGCUUCNN fourteen 132 GAAGCUUUUGAGACCCUA UU 158 UAGGGUCUCAAAAGGCUUC UU 15 133 GAAGCUUUUGAGACCCUA UU 159 uagggucuCAAAAGGCUUC UU sixteen 134 GAAGCUUUUGAGACCCUA UU 160 UagggucuCAAAAGGCUUC UU 17 135 GAAGCUUUUGAGACCCUA UU 161 UAgggucuCAAAAGGCUUC UU eighteen 136 GAAGCUUUUGAGACCCUA UU 162 UAGggucuCAAAAGGCUUC UU nineteen 137 GAAGCUUUUGAGACCCUA UU 163 UAGGgucuCAAAAGGCUUC UU 20 138 GAAGCUUUUGAGACCCUA UU 164 uAgGgUcUCAAAAGGCUUC UU 21 139 GAAGCUUUUGAGACCCUA UU 165 UAgGgUcUCAAAAGGCUUC UU 22 140 GAAGCUUUUGAGACCCUA UU 166 UaGgGuCuCAAAAGGCUUC UU 23 141 GAAGCUUUUGAGACCCUA UU 167 UAGgGuCuCAAAAGGCUUC UU 24 142 GAAGCCUUUUGAGACCCUAtt 168 UagggucuCAAAA G G C U U C UU 25 143 GAAGCUUUUGAGACCCUA UU 169 U AGGGUCUCAAAAGGCUUC UU 26 144 GAAGCUUUUGAGACCCUA UU 170 fUAGGGUCUCAAAAGGCUUC UU 27 145 GAAGCUUUUGAGACCCUA UU 171 uAGGGUCUCAAAAGGCUUC UU 28 146 GAAGCUUUUGAGACCCUA UU 172 UsAGGGUCUCAAAAGGCUUC UU 29 147 GAAGCCUUUUGAGACCCUfA UU 173 fUAGGGUCUfCAAAAGGCfUUC UU thirty 148 GAAGCCUUUUGAGfACCCUfA UU 174 fUAGGGUCUfCAfAfAAGGCfUUC UU 31 149 GAAGCCUUUUGAGACCCU AUU 175 U AGGGUCU C AAAAGGC U UC UU 31' 150 GAAGCCUUUUGAGACCCU AUU 176 fUAGGGUCU C AAAAGGC U UC UU 32 151 GAA GCCUUUUGAGACCCUA UU 177 UAGGGUCUCAAAAGGCUUC UU 39 152 GAA GCCU U U U GAGACCC U A UU 178 UAGgGuCuCAAAAGGCUUC UU 45 153 GAA GCCUUUUGAGACCCUA UU 179 U AGgGuCu C A AA AGGCU U C UU 46 154 GAA GCCU U U U GAGACCC U A UU 180 U AGgGuCu C A AA AGGCU U C UU 47 155 GAA GCCU U U U GAGACCC UAUU 181 U AGgGuCu C A AA AGGC UU C UU 48 156 GAA GCCU U U U GAGACCC UAUU 182 fUAGgGuCu C A AA AGGC UU C UU

[00127] Key to Table 2: Upper case A, G, C, and U refer to ribo-A, ribo-G, ribo-C, and ribo-U, respectively. The lowercase letters a, u, g, c, t refer to 2'-deoxy-A, 2'-deoxy-U, 2'-deoxy-G, 2'-deoxy-C, and deoxythymidine (dT=T= t), respectively. Underscore refers to 2'-OMe-substituted, e.g. U. The lowercase letter f refers to 2'-deoxy-2'-fluoro substitution, for example, fU is 2'-deoxy-2'-fluoro-U. N is A, C, G, U, U , a, c, g, u, t, or a modified, inverted, or chemically modified nucleotide.

[00128] Examples of RNAi molecules of the invention targeting GST-π mRNA are shown in Table 3.

Table 3: RNAi sequences of molecules for GST-π

ID SEQID No. MEANING CHAIN
(5'-->3')
SEQ ID ## 183 to 194 SEQID No. ANTISENSE CIRCUIT
(5'-->3')
SEQ ID ## 195 to 206 A9' 183 CCUUUUGAGACCCUGCUGUNN 195 ACAGCAGGGUCUCAAAAGGNN 1 184 CCUCAUCUACACCAACUAU UU 196 AUAGUUGGUGUAGAUGAGG UU 2 185 CCUCAUCUACACCAACUAU UU 197 auaguuggUGUAGAUGAGG UU 3 186 CCUCAUCUACACCAACUAU UU 198 AuaguuggUGUAGAUGAGG UU four 187 CCUCAUCUACACCAACUAU UU 199 AUaguuggUGUAGAUGAGG UU five 188 CCUCAUCUACACCAACUAU UU 200 AUAguuggUGUAGAUGAGG UU 6 189 CCUCAUCUACACCAACUAU UU 201 AUAGuuggUGUAGAUGAGG UU 7 190 CCUCAUCUACACCAACUAU UU 202 aUaGuUgGUGUAGAUGAGG UU 8 191 CCUCAUCUACACCAACUAU UU 203 AUaGuUgGUGUAGAUGAGG UU nine 192 CCUCAUCUACACCAACUAU UU 204 AuAgUuGgUGUAGAUGAGG UU 10 193 CCUCAUCUACACCAACUAU UU 205 AUAgUuGgUGUAGAUGAGG UU eleven 194 C C U C A U C U A CACCAACUAU UU 206 AuaguuggUGUAG A U G A G G UU

[00129] Key to Table 3: Upper case A, G, C, and U refer to ribo-A, ribo-G, ribo-C, and ribo-U, respectively. The lower case letters a, u, g, c, t refer to 2'-deoxy-A, 2'-deoxy-U, 2'-deoxy-G, 2'-deoxy-C and deoxythymidine (dT=T=t ), respectively. Underscore refers to 2'-OMe-substituted, e.g. U. The lowercase letter f refers to 2'-deoxy-2'-fluoro substitution, for example, fU is 2'-deoxy-2'-fluoro-U. N is A, C, G, U, U , a, c, g, u, t, or a modified, inverted, or chemically modified nucleotide.

[00130] Examples of RNAi molecules of the invention targeting GST-π mRNA are shown in Table 4.

Table 4: RNAi sequences of molecules for GST-π

ID SEQID No. MEANING CHAIN
(5'-->3')
SEQ ID ## 207 to 221 SEQID No. ANTISENSE CIRCUIT
(5'-->3')
SEQ ID ## 222 to 236 B13' 207 GAUGACUAUGUGAAGGCACNN 222 GUGCCUUCACAUAGUCUCNN four 208 GGAUGACUAUGUGAAGGCA UU 223 UGCCUUCACAUAGUCAUCC UU five 209 GGAUGACUAUGUGAAGGCA UU 224 ugccuucaCAUAGUCAUCC UU 6 210 GGAUGACUAUGUGAAGGCA UU 225 UgccuucaCAUAGUCAUCC UU 7 211 GGAUGACUAUGUGAAGGCA UU 226 UGccuucaCAUAGUCAUCC UU 8 212 GGAUGACUAUGUGAAGGCA UU 227 UGCcuucaCAUAGUCAUCC UU nine 213 GGAUGACUAUGUGAAGGCA UU 228 UGCCuucaCAUAGUCAUCC UU 10 214 GGAUGACUAUGUGAAGGCA UU 229 uGcCuUcACAUAGUCAUCC UU eleven 215 GGAUGACUAUGUGAAGGCA UU 230 UGcCuUcACAUAGUCAUCC UU 12 216 GGAUGACUAUGUGAAGGCA UU 231 UgCcUuCaCAUAGUCAUCC UU 13 217 GGAUGACUAUGUGAAGGCA UU 232 UGCcUuCaCAUAGUCAUCC UU fourteen 218 G G A U G A C UAUGUGAAGGCA UU 233 UgccuucaCAUAG U C A U C C UU 15 219 GGAUGACUAUfGUfGAAGGCA UU 234 UGCfCUUCACAUAGUCAUCC UU 17 220 GG AUGACUAUGUGAAGGCA UU 235 UGCCUUCA C A U A G U C A U C CUU eighteen 221 GG AUGACUAUGUGAAGGCA UU 236 UGCCUUCACA U A G U C A U C CUU

[00131] Key to Table 4: A, G, C, and U in uppercase refer to ribo-A, ribo-G, ribo-C, and ribo-U, respectively. The lower case letters a, u, g, c, t refer to 2'-deoxy-A, 2'-deoxy-U, 2'-deoxy-G, 2'-deoxy-C and deoxythymidine (dT=T=t ), respectively. Underscore refers to 2'-OMe-substituted, e.g. U. The lowercase letter f refers to 2'-deoxy-2'-fluoro substitution, for example, fU is 2'-deoxy-2'-fluoro-U. N is A, C, G, U, U , a, c, g, u, t, or a modified, inverted, or chemically modified nucleotide.

[00132] Examples of RNAi molecules of the invention targeting GST-π mRNA are shown in Table 5.

Table 5: RNAi sequences of molecules for GST-π

ID SEQID No. MEANING CHAIN
(5'-->3')
SEQ ID ## 237 to 248 SEQID No. ANTISENSE CIRCUIT
(5'-->3')
SEQ ID ## 249 to 260 B2' 237 GAAGCCUUUUGAGACCCUGNN 249 CAGGGUCUCAAAAGGCUUCNN 1 238 GAAGCCUUUUGAGACCCUG UU 250 CAGGGUCUCAAAAGGCUUC UU 2 239 GAAGCCUUUUGAGACCCUG UU 251 cagggucuCAAAAGGCUUC UU 3 240 GAAGCCUUUUGAGACCCUG UU 252 CagggucuCAAAAGGCUUC UU four 241 GAAGCCUUUUGAGACCCUG UU 253 CAgggucuCAAAAGGCUUC UU five 242 GAAGCCUUUUGAGACCCUG UU 254 CAGggucuCAAAAGGCUUC UU 6 243 GAAGCCUUUUGAGACCCUG UU 255 CAGGgucuCAAAAGGCUUC UU 7 244 GAAGCCUUUUGAGACCCUG UU 256 cAgGgUcUCAAAAGGCUUC UU 8 245 GAAGCCUUUUGAGACCCUG UU 257 CAgGgUcUCAAAAGGCUUCUU nine 246 GAAGCCUUUUGAGACCCUG UU 258 CaGgGuCuCAAAAGGCUUC UU 10 247 GAAGCCUUUUGAGACCCUG UU 259 CAGgGuCuCAAAAGGCUUC UU eleven 248 G A A G C C U U U UGAGACCCUG UU 260 CagggucuCAAAA G G C U U C UU

[00133] Key to Table 5: Upper case A, G, C, and U refer to ribo-A, ribo-G, ribo-C, and ribo-U, respectively. The lower case letters a, u, g, c, t refer to 2'-deoxy-A, 2'-deoxy-U, 2'-deoxy-G, 2'-deoxy-C and deoxythymidine (dT=T=t ), respectively. Underscore refers to 2'-OMe-substituted, e.g. U. The lowercase letter f refers to 2'-deoxy-2'-fluoro substitution, for example, fU is 2'-deoxy-2'-fluoro-U. N is A, C, G, U, U , a, c, g, u, t, or a modified, inverted, or chemically modified nucleotide.

[00134] Examples of RNAi molecules of the invention targeting GST-π mRNA are shown in Table 6.

Table 6: RNAi sequences of molecules for GST-π

ID SEQID No. MEANING CHAIN
(5'-->3')
SEQ ID ## 261 to 272 SEQID No. ANTISENSE CIRCUIT
(5'-->3')
SEQ ID ## 273 to 284 B4' 261 CCUCAUCUACACCAACUAUNN 273 AUAGUUGGUGUAGAUGAGGNN 1 262 CCUCAUCUACACCAACUAU UU 274 AUAGUUGGUGUAGAUGAGG UU 2 263 CCUCAUCUACACCAACUAU UU 275 auaguuggUGUAGAUGAGG UU 3 264 CCUCAUCUACACCAACUAU UU 276 AuaguuggUGUAGAUGAGG UU four 265 CCUCAUCUACACCAACUAU UU 277 AUaguuggUGUAGAUGAGG UU five 266 CCUCAUCUACACCAACUAU UU 278 AUAguuggUGUAGAUGAGG UU 6 267 CCUCAUCUACACCAACUAU UU 279 AUAGuuggUGUAGAUGAGG UU 7 268 CCUCAUCUACACCAACUAU UU 280 aUaGuUgGUGUAGAUGAGG UU 8 269 CCUCAUCUACACCAACUAU UU 281 AUaGuUgGUGUAGAUGAGG UU nine 270 CCUCAUCUACACCAACUAU UU 282 AuAgUuGgUGUAGAUGAGG UU 10 271 CCUCAUCUACACCAACUAU UU 283 AUAgUuGgUGUAGAUGAGG UU eleven 272 C C U C A U C U A CACCAACUAU UU 284 AuaguuggUGUAG A U G A G G UU

[00135] Key to Table 6: A, G, C, and U in uppercase refer to ribo-A, ribo-G, ribo-C, and ribo-U, respectively. The lower case letters a, u, g, c, t refer to 2'-deoxy-A, 2'-deoxy-U, 2'-deoxy-G, 2'-deoxy-C and deoxythymidine (dT=T=t ), respectively. Underscore refers to 2'-OMe-substituted, e.g. U. The lowercase letter f refers to 2'-deoxy-2'-fluoro substitution, for example, fU is 2'-deoxy-2'-fluoro-U. N is A, C, G, U, U , a, c, g, u, t, or a modified, inverted, or chemically modified nucleotide.

[00136] In some embodiments, the invention provides a series of nucleic acid molecules, in which: a) the molecule has a polynucleotide sense strand and a polynucleotide antisense strand; b) each strand of the molecule is 15 to 30 nucleotides in length; c) a contiguous region of 15 to 30 nucleotides of the antisense strand is complementary to the mRNA sequence encoding GST-π; d) at least part of the sense strand is complementary to at least part of the antisense strand and the molecule has a duplex region of 15 to 30 nucleotides in length.

[00137] In some embodiments, the implementation of the nucleic acid molecule may have a contiguous region of 15 to 30 nucleotides of the antisense strand, which is complementary to the mRNA sequence encoding GST-π, located in the duplex region of the molecule.

[00138] In further embodiments, the nucleic acid molecule may have a contiguous region of 15 to 30 nucleotides of an antisense strand that is complementary to the mRNA sequence encoding GST-π.

[00139] In certain embodiments, each strand of a nucleic acid molecule can be 18 to 22 nucleotides in length. The duplex region of a nucleic acid molecule can be 19 nucleotides in length.

[00140] In alternative forms, the nucleic acid molecule may have a polynucleotide sense strand and a polynucleotide antisense strand that are joined as one strand and form a duplex region connected at one end with a loop.

[00141] Some embodiments of the nucleic acid molecule of this disclosure may have a blunt end. In certain embodiments, the implementation of the nucleic acid molecule may have one or more nucleotides 3' sticky end.

[00142] This invention provides a number of nucleic acid molecules that are RNAi molecules active for gene silencing. Inventive nucleic acid molecules can be dsRNA, siRNA, miRNA, or shRNA active for gene silencing, as well as DNA-directed RNA (ddRNA), Piwi-interacting RNA (piRNA), or repeat-associated siRNA (rasiRNA). Nucleic acid molecules may be active to inhibit GST-π expression.

[00143] Embodiments of the invention further provide nucleic acid molecules having an IC ₅₀ for GST-π knockdown of less than 100 pM.

[00144] Additional embodiments of the present invention provide nucleic acid molecules having an IC ₅₀ for GST-π knockdown of less than 50 pM.

[00145] This invention further provides compositions comprising one or more inventive nucleic acid molecules, along with a pharmaceutically acceptable carrier. In certain embodiments, the carrier may be a lipid molecule or a liposome.

[00146] The compounds and compositions of this invention can be used in methods for preventing or treating a disease associated with GST-π by administering the compound or composition to a subject in need thereof.

[00147] The methods of this invention allow the use of inventive compounds in the prevention or treatment of cancer. Cancer can be present in various diseases, e.g., cancers associated with GST-π expression, cancers due to cells expressing mutant KRAS, sarcomas, fibrosarcoma, malignant fibrous histiocytoma, liposarcoma, rhabdomyosarcoma, leiomyosarcoma, angiosarcoma, Kaposi's sarcoma, lymphangiosarcoma, synovial sarcoma, chondrosarcoma, osteosarcoma, carcinomas, brain tumor, head and neck cancer, breast cancer, lung cancer, esophageal cancer, gastric cancer, duodenal cancer, appendix cancer, colon cancer colon, rectal cancer, liver cancer, pancreatic cancer, gallbladder cancer, bile duct cancer, pancreatic cancer mustache, kidney cancer, urethral cancer, bladder cancer, prostate cancer, testicular cancer, uterine cancer, ovarian cancer, skin cancer, leukemia, lymphoma cancer, epithelial cancer, and non-epithelial cancer.

[00148] P21 and RNAi molecules

[00149] p21 is a cell cycle regulation protein encoded by the CDKN1A gene and belonging to the CIP/KIP family. This protein has the function of inhibiting cell cycle progression in G1 phase and G2/M phase by inhibiting the effect of the cyclin-CDK complex through binding to this complex. In particular, the p21 gene is activated by p53, one of the tumor suppressor genes. It has been reported that when p53 is activated due to DNA damage or the like, p53 activates p21 so that the cell cycle is arrested in G1 phase and G2/M phase.

[00150] p21 is overexpressed in various human cancers, including prostate, cervical, breast, and squamous cell carcinomas, and, in many cases, upregulation of p21 is positively correlated with tumor grade, invasiveness, and aggressiveness. See, for example, Chang et al., Proc. Natl. Acad. sci. USA, 2000, Vol. 97, No. 8, pp. 4291-96. Upregulation of p21 has also been reported to be associated with oncogenicity and poor prognosis in many forms of cancer, including brain, prostate, ovarian, breast, and esophageal cell cancers. See, for example, Winters et al., Breast Cancer Research, 2003, vol. 5, no. 6, pp. R242-R249. Also, the disease may be age-related diseases, including atherosclerosis, Alzheimer's disease, amyloidosis, and arthritis. See, for example, Chang et al., Proc. Natl. Acad. sci. USA, 2000, Vol. 97, No. 8, pp. 4291-96.

[00151] p21 is present in various animals, including humans. Information about the sequence of the CDKN1A of a person (P21) can be found according to: NM_000389.4, NM_078467.2, NM_001291549.1, NM_001220778.1, NM_001220777.1 (NP_001207707.1, NP_001278478.1, NP_00120706.1, NP_510867.1, NP_000, NP_000. 1).

[00152] The target human p21 mRNA is disclosed under Genbank Accession Number NM_000389.4 (CDKN1A) and is 2175 bp in length.

[00153] One skilled in the art will appreciate that the sequence given may change over time and will include any necessary changes to the nucleic acid molecules herein as appropriate.

[00154] Embodiments of the invention may provide compositions and methods for silencing p21 gene expression using small nucleic acid molecules. Examples of nucleic acid molecules include RNA interference active molecules (RNAi molecules), small interfering RNA (siRNA), microRNA (miRNA), and short hairpin RNA (shRNA) molecules, as well as DNA-guided RNA (ddRNA), RNA , interacting with Piwi (piRNA), and associated with miRNA repeats (rasiRNA). Such molecules are capable of mediating RNA interference against p21 gene expression.

[00155] The composition and methods disclosed herein can also be used in the treatment of various types of cancer in a subject.

[00156] The nucleic acid molecules and methods of this invention can be used to down-regulate the expression of genes that encode p21.

[00157] The compositions and methods of this invention may contain one or more nucleic acid molecules that, independently or in combination, can modulate or regulate the expression of a p21 protein and/or genes encoding p21 proteins, proteins and/or genes encoding p21, associated with the maintenance and/or development of diseases, conditions or disorders associated with p21, such as cancer.

[00158] The compositions and methods of this invention are described with reference to exemplary p21 sequences. One skilled in the art will appreciate that various aspects and embodiments of the invention are directed to any related p21 genes, sequences, or variants, such as homologous genes and transcriptional variants and polymorphisms, including single nucleotide polymorphisms (SNPs) associated with any p21 genes.

[00159] In some embodiments, the compositions and methods of this invention may include a double-stranded small interfering nucleic acid (siRNA) molecule that down-regulates the expression of a p21 gene, such as human CDKN1A.

[00160] An RNAi molecule of the invention can be targeted to p21 and any homologous sequences, e.g., using complementary sequences or by inserting non-canonical base pairs, e.g., mismatches and/or ambiguous base pairs, which may provide additional target sequences.

[00161] In cases where mismatches are identified, non-canonical base pairs, such as mismatches and/or ambiguous base pairs, can be used to generate nucleic acid molecules that target more than one gene sequence.

[00162] For example, non-canonical base pairs, such as UU and CC base pairs, can be used to design nucleic acid molecules that are capable of targeting sequences for different p21 targets that share sequence homology. Thus, an RNAi molecule can be directed to a nucleotide sequence that is conserved between homologous genes, and a single RNAi molecule can be used to inhibit the expression of more than one gene.

[00163] In some aspects, the compositions and methods of this invention include RNAi molecules that are active against p21 mRNA, where the RNAi molecule contains a sequence complementary to any mRNA encoding the p21 sequence.

[00164] In some embodiments, an RNAi molecule of this disclosure may have activity against a p21 RNA, wherein the RNAi molecule contains a sequence complementary to an RNA having a variant p21 encoding sequence, such as a mutant p21 gene that is known in the art to be associated with a malignant tumor.

[00165] In further embodiments, an RNAi molecule of the invention may comprise a nucleotide sequence that can interact with a p21 gene nucleotide sequence and mediate silencing of p21 gene expression.

[00166] Nucleic acid molecules for inhibiting p21 expression may have a sense strand and an antisense strand in which the strands form a duplex region. Nucleic acid molecules may have one or more modified or chemically modified nucleotides in the duplex region, including such modifications as are known in the art. Any nucleotide at the sticky end of an siRNA can also be modified or chemically modified.

[00167] In some embodiments, the preferred modified or chemically modified nucleotides are 2'-deoxynucleotides. In additional embodiments, the modified or chemically modified nucleotides may include 2'-O-alkyl-substituted nucleotides, 2'-deoxy-2'-fluoro-substituted nucleotides, phosphothioate nucleotides, closed nucleotides, or any combination thereof.

[00168] In certain embodiments, a preferred structure may have an antisense strand containing deoxynucleotides in a plurality of positions, the plurality of positions being one of the following: each of positions 4, 6, and 8 from the 5' end of the antisense strand; each of positions 3, 5 and 7 from the 5' end of the antisense strand; each of positions 1, 3, 5 and 7 from the 5' end of the antisense strand; each of positions 3-8 from the 5' end of the antisense strand; and each of positions 5-8 from the 5' end of the antisense strand. Any of these structures can be combined with one or more 2'-deoxy-2'-fluoro-substituted nucleotides in the duplex region.

[00169] The nucleic acid molecules of the invention can inhibit the expression of p21 mRNA with a favorable IC _{50 of} less than about 200 pM. In addition, nucleic acid molecules can inhibit the expression of p21 mRNA at certain levels, at least 25% in vivo, after a single injection.

[00170] This invention provides pharmaceutical compositions that may contain one or more miRNAs, as disclosed in the present description, in combination with a pharmaceutically acceptable carrier. Any suitable carrier may be used, including those known in the art, as well as lipid molecules, nanoparticles, or liposomes, any of which can encapsulate siRNA molecules.

[00171] Disclosed herein are methods for treating a disease that may be associated with p21 expression, which methods include administering to a subject in need thereof a composition comprising one or more siRNAs. The diseases to be treated may include cancer, cancer caused by cells expressing mutant KRAS, sarcoma, and carcinoma, among others.

[00172] Examples of RNAi molecules of the invention targeting p21 mRNA are shown in Table 7.

Table 7: RNAi sequences of molecules for p21

Reference position SEQID No. MEANING CHAIN
(5'-->3')
SEQ ID ## 285 to 312 SEQID No. ANTISENSE CIRCUIT
(5'-->3')
SEQ ID ## 313 to 340 2085 285 CUUAGUGACUUUACUUGUAMUmU 313 UACAAGUAAAGUCACUAAGmUmU 500 286 CAGACCAGCAUGACAGAUUmUmU 314 AAUCUGUCUAUGCUGGUCUGmUmU 540 287 UGAUCUUCUCCAAGAGGAAmUmUmU 315 UUCCUCUUGGAGAAGAUCAmUmU 1706 288 GUUCAUUGCACUUUGAUUAmUmU 316 UAAUCAAAGUGCAAUGAACmUmU 1709 289 CAUUGCACUUUGAUUAGCAmUmU 317 UGCUAAUCAAAGUGCAAUGmUmU 210 290 AGCGAUGGAACUUCGACUUmUmU 318 AAGUCGAAGUUCCAUCCGCUmUmU 211 291 GCGAUGGAACUUCGACUUUmUmU 319 AAAGUCGAAGUUCCAUCGCmUmU 1473 292 GGGAAGGGACACACAAGAAmUmU 320 UUCUUGUGUGUCCCUUCCCmUmU 1507 293 UCUACCUAGGCAGCUCAAmUmU 321 UUGAGCUGCCUGAGGUAGAMUmU 2067 294 GGUGCUCAAUAAAUGAUUCmUmU 322 GAAUCAUUUAUUGAGCACCmUmU 1063 295 CAUCAUCAAAAACUUUGGAmUmU 323 UCCAAAGUUUUUGAUGAUGmUmU 1735 296 AAGGAGUCAGACAUUUUAAmUmU 324 UUAAAAUGUCUGACUCCUUmUmU 783 297 GUGCUGGGCAUUUUUAUUUmUmU 325 AAAUAAAAAUGCCCAGCACmUmUmU 869 298 GCCGGCUUCAUGCCAGCUAmUmU 326 UAGCUGGCAUGAAGCCGGCmUmU 1060 299 GGGCAUCAUCAAAAACUUUmUmU 327 AAAGUUUUUGAUGAUGCCCmUmU 1492 300 GAAGGGCACCCUAGUUCUAmUmU 328 UAGAACUAGGGUGCCCUUCmUmU 1704 301 CAGUUCAUUGCACUUUGAUmUmU 329 AUCAAAGUGCAAUGAACUGmUmU 1733 302 ACAAGGAGUCAGAACAUUUUmUmU 330 AAAAUGUCUGACUCCUUGUmUmU 1847 303 UGGAGGCACUGAAGUGCUUmUmU 331 AAGCACUUCAGUGCCUCCAmUmU 2000 304 GCAGGGACCACACCUGUAmUmU 332 UACAGGGUGUGGUCCCUGCmUmU 2014 305 CUGUACUGUUCUGUGUCUUmUmU 333 AAGACACAGAACAGUACAGmUmU 677 306 UUAAAACACCUCCUCAUGUAMUmU 334 UACAUGAGGAGGUGUUUAAmUmU 475 307 AGACUCUCAGGGUCGAAAAmUmU 335 UUUUCGACCCUGAGAGUCUmUmU 508 308 CAUGACAGAUUUCUACCACmUmU 336 GUGGUAGAAAUCUGUCAUGmUmU 514 309 AGAUUUCUACCACUCCAAAmUmU 337 UUUGGAGUGGUAGAAAUCUmUmU 549 310 CCAAGAGGAAGCCCUAAUCmUmU 338 GAUUAGGGCUUCCUCUUGGmUmU 382 311 GACAGCAGAGGAAGACCAUmUmU 339 AUGGUCUUCCUCUGCUGUCmUmU 2042 312 CUCCCACAAUGCUGAAUAUumUmU 340 AUAUUCAGCAUUGUGGGAGmUmU

[00173] Key to Table 7: A, G, C, and U in uppercase refer to ribo-A, ribo-G, ribo-C, and ribo-U, respectively. The lowercase letters a, g, c, t represent 2'-deoxy-A, 2'-deoxy-G, 2'-deoxy-C, and thymidine, respectively. mU is 2'-methoxy-U.

[00174] Examples of RNAi molecules of the invention targeting p21 mRNA are shown in Table 8.

Table 8: RNAi sequences of molecules for p21

Reference position SEQID No. MEANING CHAIN
(5'-->3')
SEQ ID ## 341 to 354 SEQID No. ANTISENSE CIRCUIT
(5'-->3')
SEQ ID ## 355 to 368 1735' 341 AAGGAGUCAGACAAUUUUAANN 355 UUAAAAUGUCUGACUCCUUNN 1 342 AAG GAGU C AGA C AUUU U AA UU 356 U U AaAa U gUC U GACUCCUU UU 2 343 AAG GAGU C AGA C A UU U U AA UU 357 U U AaAa U gUC U GACUCCUU UU 3 344 AAG GAGU C AGA C A UU U U AA UU 358 U U AaAaUgUCUGACUCCUU UU four 345 AAG GAGU C AGA C AUUU U AA UU 359 U U AaAaUgUCUGACUCCUU UU five 346 AAGGAGUCAGACAUUUUAA UU 360 UUaaaaugUCUGACUCCUU UU 6 347 AAGGAGUCAGACAUUUUAA UU 361 UUAAaaugUCUGACUCCUU UU 7 348 AAGGAGUCAGACAUUUUAA UU 362 uUaAaAuGUCUGACUCCUU UU 8 349 AAGGAGUCAGACAUUUUAA UU 363 UUaAaAuGUCUGACUCCUU UU nine 350 AAGGAGUCAGACAUUUUAA UU 364 UUAaAaUgUCUGACUCCUU UU 10 351 AAGGAGUCAGACAUUUUAA UU 365 U U AAAA U GUC U GACUCCUU UU eleven AAGGAGU C AGA C AUUU U AA UU 366 UUAAAAUGUCUGACUCCUU UU 12 353 AAG GAGUCAGACAUUUUAA UU 367 UUAAAAUGUCUGACUCCUU UU 13 354 AAG GAGU C AGA C AUUU U AA UU 368 UUAAAAUGUCUGACUCCUU UU

[00175] Key to Table 8: Upper case A, G, C, and U refer to ribo-A, ribo-G, ribo-C, and ribo-U, respectively. The lower case letters a, u, g, c, t refer to 2'-deoxy-A, 2'-deoxy-U, 2'-deoxy-G, 2'-deoxy-C and deoxythymidine (dT=T=t ), respectively. Underscore refers to 2'-OMe-substituted, e.g. U. N is A, C, G, U, U , a, c, g, u, t, or a modified, inverted, or chemically modified nucleotide.

[00176] Embodiments of this invention cover siRNA molecules from Tables 1-8 that are modified or chemically modified in accordance with the examples above.

[00177] In some embodiments, the invention provides a series of nucleic acid molecules, where a) the molecule has a polynucleotide sense strand and a polynucleotide antisense strand; b) each strand of the molecule is 15 to 30 nucleotides in length; c) a contiguous region of 15 to 30 nucleotides of the antisense strand is complementary to the mRNA sequence encoding p21; and d) at least a portion of the sense strand is complementary to at least a portion of the antisense strand and the molecule has a duplex region of 15 to 30 nucleotides in length.

[00178] In some embodiments, the nucleic acid molecule may have a contiguous region of 15 to 30 nucleotides of an antisense strand that is complementary to the mRNA sequence encoding p21 and located in the duplex region of the molecule.

[00179] In further embodiments, the nucleic acid molecule may have a contiguous region of 15 to 30 nucleotides of an antisense strand that is complementary to the mRNA sequence encoding p21.

[00180] In additional aspects, the nucleic acid molecule of this invention may have each strand of the molecule between 18 and 22 nucleotides in length. A nucleic acid molecule may have a duplex region of 19 nucleotides in length.

[00181] In certain embodiments, the nucleic acid molecule may have a polynucleotide sense strand and a polynucleotide antisense strand, which are connected as one strand and form a duplex region connected at one end with a loop.

[00182] The nucleic acid molecules of this invention may be blunt-ended, and may have one or more sticky 3' ends.

[00183] Nucleic acid molecules of the invention can be RNAi molecules that are active for gene silencing, such as dsRNA that is active for gene silencing, siRNA, miRNA, or shRNA that is active for gene silencing, as well as RNA directed by DNA (ddRNA), RNA interacting with Piwi (piRNA), and associated with miRNA repeats (rasiRNA).

[00184] This invention provides a number of nucleic acid molecules that are active to inhibit the expression of p21. In some embodiments, the nucleic acid molecule may have an IC ₅₀ for p21 knockdown of less than 100 pM.

[00185] In further embodiments, the nucleic acid molecule may have an IC ₅₀ for p21 knockdown of less than 50 pM.

[00186] This invention further provides compositions comprising one or more inventive nucleic acid molecules and a pharmaceutically acceptable carrier. The carrier may be a lipid molecule or a liposome.

[00187] The compounds and compositions of this invention can be used in methods for preventing or treating a disease associated with p21 by administering the compound or composition to a subject in need thereof.

[00188] In additional aspects, this invention includes methods for treating a disease associated with p21 expression by administering to a subject in need thereof a composition comprising one or more inventive nucleic acid molecules. The disease may be a cancer that may be present in a disease such as cancers associated with p21 expression, among others.

[00189] The methods of this invention allow the use of inventive compounds in the prevention or treatment of cancer. Cancer can be present in various diseases, e.g. p21 expression cancers, cancers due to cells expressing mutant KRAS, sarcomas, fibrosarcoma, malignant fibrous histiocytoma, liposarcoma, rhabdomyosarcoma, leiomyosarcoma, angiosarcoma, Kaposi's sarcoma, lymphangiosarcoma, synovial sarcoma, chondrosarcoma, osteosarcoma, carcinomas, brain tumor, head and neck cancer, breast cancer, lung cancer, esophageal cancer, gastric cancer, duodenal cancer, appendix cancer, colon cancer, rectal cancer, liver cancer, pancreatic cancer, gallbladder cancer, bile duct cancer, kidney cancer and, urethral cancer, bladder cancer, prostate cancer, testicular cancer, uterine cancer, ovarian cancer, skin cancer, leukemia, lymphoma cancer, epithelial cancers, and non-epithelial cancers.

[00190] Embodiments of the invention may provide RNAi molecules that can be used to down-regulate or inhibit gene and/or protein expression.

[00191] The RNAi molecules of this disclosure can be used alone or in combination with other siRNAs to modulate the expression of one or more genes.

[00192] The RNAi molecules of this disclosure can be used alone or in combination or in combination with other known drugs to prevent or treat diseases or improve the symptoms of conditions or disorders.

[00193] RNAi molecules according to this invention can be used to modulate or inhibit the expression of a gene with a specific sequence.

[00194] RNAi molecules of this disclosure may include a guide strand for which a number of contiguous nucleotides is at least partially complementary to human mRNA.

[00195] Embodiments of the present invention may include methods for preventing, treating, or improving symptoms of a disease or condition in a subject in need thereof.

[00196] In some embodiments, methods for preventing, treating, or improving symptoms of cancer in a subject may include administering to the subject an RNAi molecule of the invention in order to modulate gene expression in the subject or in an organism.

[00197] In some embodiments, this invention provides methods for down-regulating gene expression in a cell or organism by bringing the cell or organism into contact with an RNAi molecule of the invention.

[00198] RNAi molecules directed to Hsp47

[00199] In some embodiments, this invention may provide a range of RNAi molecules and compositions for modulating the expression of heat shock protein 47 (Hsp47), a collagen-specific molecular chaperone for intracellular transport and maturation.

[00200] Some examples of siRNAs for Hsp47 are given in US 8,710,209, which is hereby incorporated by reference in its entirety for all purposes.

[00201] Hsp47 or a gene sequence homologous to it is disclosed, for example, under GenBank accession number AB010273 (human), X60676 (mouse) or M69246 (rat, gp46).

[00202] Agents for suppressing Hsp47 are disclosed for inhibiting fibrosis. See, for example, US 8,173,170 B2, which is hereby incorporated by reference in its entirety for all purposes. However, there is limited information regarding the effect of Hsp47 inhibition on the development, progression and growth of malignant tumors.

[00203] In some embodiments, each siRNA strand of a molecule of the invention may be 15 to 60 nucleotides in length, or 15 to 40 nucleotides in length, or 19 to 25 nucleotides in length.

[00204] In certain embodiments, this invention provides a pharmaceutical composition containing RNAi molecules for the treatment of cancer, which are RNAi molecules directed to Hsp47.

[00205] Examples of RNAi molecules of this disclosure directed to Hsp47 mRNA are shown in Table 9.

Table 9: RNAi sequences of molecules for Hsp47

SEQID No. MEANING CHAIN
(5'-->3')
SEQ ID ## 369 to 389 SEQID No. ANTISENSE CIRCUIT
(5'-->3')
SEQ ID ## 390 to 410 mouse 369 CGAGAACAGUUUGUACAAGUU 390 CUUGUACAACUGUUCUCGUU 370 CAGGCCUCUACAACUACUATT 391 UAGUAGUUGUAGAGGCCUGTT 371 GAGCACUCCAAGAUCAACUUCCGCG 392 CGCGGAAGUUGAUCUUGGAGUGCUCUU 372 GGACAGGCCUCUACAACUATT 393 UAGUUGUAGAGGCCUGUCCTT 373 GAGCACUCCAAGAUCAACUTT 394 AGUUGAUCUUGGAGUGCUCTT 374 GAACACUCCAAGAUCAACUTT 395 AGUUGAUCUUGGAGUGUUCTT 375 CAGGCCUCUACAACUACUACGACGA 396 UCGUCGUAGUAGUUGUAGAGGCCUGUU 376 GAACACUCCAAGAUCAACUUCCGAG 397 CUCGGAAGUUGAUCUUGGAGUGUUCUU 377 GGACAGGCCUUACAACUACUACGA 398 UCGUAGUAGUUGUAGAGGCCUGUCCUU 378 CAGGCCUCUACAACUACUAdTdTdAdAdAdAdA 399 UAGUAGUUGUAGAGGCCUGdTdT 379 CAGGCCUCUACAACUACUA 400 UAGUAGUUGUAGAGGCCUGdTdT 380 CAGGCCUCUACAACUACUAdTdTdAdAdAdAdAdAdAdAdAdAdAdA 401 UAGUAGUUGUAGAGGCCUGdTdT 381 dAdAdAdAdACAGGCCUCUACAACUACUAdTdT 402 UAGUAGUUGUAGAGGCCUGdTdT 382 CAGGCCUCUACAACUACUAdTdTdAdAdAdAdAdAdAdA 403 UAGUAGUUGUAGAGGCCUGdTdT 383 dAdAdAdAdAdAdAdACAGGCCUCUACAACUACUAdTdT 404 UAGUAGUUGUAGAGGCCUGdTdT 384 CAGGCCUCUACAACUACUAdTdTdAdAdAdAdAdAdAdAdAdAdAdA 405 UAGUAGUUGUAGAGGCCUGdTdT 385 dAdAdAdAdAdAdAdAdAdAdAdAdAdAdAdACAGGCCUCUACAACUACUAdTdT 406 UAGUAGUUGUAGAGGCCUGdTdT 386 CAGGCCUCUACAACUACUAdTdTdAdAdAdAdAdAdAdAdAdAdAdAdAdAdAdA 407 UAGUAGUUGUAGAGGCCUGdTdT 387 CAGGCCUCUACAACUACUAdTdT 408 UAGUAGUUGUAGAGGCCUGdTdT mouse 388 GGACAGGCCUGUACAACUadTdT 409 UAGUUGUACAGGCCUGUCCdTdT human 389 GGACAGGCCUUACAACUAdTdT 410 UAGUUGUAGAGGCCUGUCCdTdT

[00206] Key to Table 9: Upper case A, G, C, and U refer to ribo-A, ribo-G, ribo-C, and ribo-U, respectively. lowercase d stands for "deoxy".

[00207] In Table 9, the antisense strand of any of the RNAi molecules directed to Hsp47 may alternatively have deoxynucleotides in a plurality of positions, the plurality of positions being one of the following: each of positions 4, 6, and 8 from the 5' end of the antisense strand; each of positions 3, 5 and 7 from the 5' end of the antisense strand; each of positions 1, 3, 5 and 7 from the 5' end of the antisense strand; each of positions 3-8 from the 5' end of the antisense strand; and each of positions 5-8 from the 5' end of the antisense strand.

[00208] Additional examples of RNAi molecules of this disclosure targeting Hsp47 mRNA are shown in Table 10.

Table 10: RNAi sequences of molecules and controls for Hsp47

SEQID No. Sequence Name 411 3'-C3-25rU-25rC-25rC-25rU-25rU-rC-rA-rA-rC-rU-rA-rG-rA-rA-rC-rC-rU-rC-rA-idAB-5' Semantic "A" 412 3'-C3-C3-mU-rG-mA-rG-mG-rU-mU-rC-mU-rA-rG-mU-25rU-mG-rA-mA-rG-mG-rA-5' Antisense "A" 413 3'-C3-rA-rG-mU-rG-rG-rG-mU-rA-mC-rA-25rC-rA-rG-rA-rG-mU-rC-rC-rU-idAB-5' Semantic "B" 414 3'-C3-C3-rA-rG-rG-rA-mC-rU-rC-rU-mG-rU-rG-mU-rA-25rC-rC-mC-rA-rC-mU-5' Antisense "B" 415 5'-idAB-rG-rA-rG-rA-rC-rA-rC-rA-rU-rG-rG-rG-rU-rG-25rC-25rU-25rA-25rU-25rA-C3-P-3' Semantic "C" 416 5'-mU-rA-mU-rA-mG-rC-25rA-rC-mC-rC-mA-rU-mG-rU-mG-rU-mC-rU-mC-C3-C3-3' Antisense "C" 417 5'-idAB-rC-mU-mU-rA-mC-rG-mC-mU-25rG-rA-rG-mU-rA-mC-mU-rU-mC-rG-rU-C3-3' semantic "D",
Negative control 418 5'-rA-mC-rG-rA-rA-25rG-mU-rA-rC-rU-mC-rA-rG-rC-rG-mU-rA-rA-rG-C3-C3-3' Antisense "D",
Negative control

[00209] Key to Table 10: Symbols: rX represents ribonucleotides, mX represents 2'-O-methyl ribonucleotides, 25rX represents ribonucleotides with 2'-5' bonds, C3 represents 1,3-propanediol spacer, idAB represents inverted 1,2 -dideoxy-D-ribose, P represents the phosphate group at the 3' end.

[00210] In Table 10, the antisense strand of any of the RNAi molecules directed to Hsp47 may alternatively have deoxynucleotides in a plurality of positions, the plurality of positions being one of the following: each of positions 4, 6, and 8 from the 5' end of the antisense strand; each of positions 3, 5 and 7 from the 5' end of the antisense strand; each of positions 1, 3, 5 and 7 from the 5' end of the antisense strand; each of positions 3-8 from the 5' end of the antisense strand; and each of positions 5-8 from the 5' end of the antisense strand.

[00211] RNAi molecules directed to MCL1

[00212] In some embodiments, the invention may provide a range of RNAi molecules and compositions for modulating the expression of Homo sapiens myeloid cell leukemia 1 (MCL1) mRNA transcriptional variant 2.

[00213] MCL1 is disclosed, for example, under Accession No. NM_182763.2 (human).

[00214] Examples of RNAi molecules of this disclosure directed to MCL1 mRNA are shown in Table 11.

Table 11: RNAi sequences of molecules for MCL1

SEQID No. MEANING CHAIN
(5'-->3')
SEQ ID ## 419 to 426 SEQID No. ANTISENSE CIRCUIT
(5'-->3')
SEQ ID ## 427 to 434 419 GCCUUCCAAGGAUGGGUUUGU 427 AAACCCAUCCUUGGAAGGCCG 420 GGAGUUCUUCCAUGUAGAGGA 428 CUCUACAUGGAAGAACUCCAC 421 CCAUGUAGAGGACCUAGAAGG 429 UUCUAGGUCCCUCUACAUGGAA 422 GCCUUCCAAGGAUGGGUUUGU 430 AAAcCcAuCCUUGGAAGGCCG 423 GCCUUCCAAGGAUGGGUUUGU 431 AAaCCCaUCCUUGGAAGGCCG 424 GCCUUCCAAGGAUGGGUUUGU 432 aAaCcCaUCCUUGGAAGGCCG 425 GCCUUCCAAGGAUGGGUUUGU 433 AAacccauCCUUGGAAGGCCG 426 GCCUUCCAAGGAUGGGUUUGU 434 AAACccauCCUUGGAAGGCCG

[00215] Key to Table 11: A, G, C, and U in uppercase refer to ribo-A, ribo-G, ribo-C, and ribo-U, respectively. lowercase d represents "deoxy".

[00216] In Table 11, the antisense strand of any of the RNAi molecules directed to MCL1 may alternatively have deoxynucleotides in a plurality of positions, the plurality of positions being one of the following: each of positions 4, 6, and 8 from the 5' end of the antisense strand; each of positions 3, 5 and 7 from the 5' end of the antisense strand; each of positions 1, 3, 5 and 7 from the 5' end of the antisense strand; each of positions 3-8 from the 5' end of the antisense strand; and each of positions 5-8 from the 5' end of the antisense strand.

[00217] RNAi molecules targeted to ARAF

[00218] In some embodiments, the invention may provide a range of RNAi molecules and compositions for modulating Homo sapiens serine/threonine kinase proto-oncogene A-Raf (ARAF) mRNA expression of transcriptional variant 1 gene.

[00219] ARAF is disclosed, for example, under accession number NM_001654.4 (human).

[00220] Examples of RNAi molecules of this disclosure targeting ARAF mRNA are shown in Table 12.

Table 12: RNAi sequences of molecules for ARAF

SEQID No. MEANING CHAIN
(5'-->3')
SEQ ID ## 435 to 442 SEQID No. ANTISENSE CIRCUIT
(5'-->3')
SEQ ID ## 443 to 450 435 GCUCAUUGUCGAGGUCCUUGA 443 AAGGACCUCGACAAUGAGCUC 436 GCCAAACCUGUGGCUACAAGU 444 UUGUAGCCACAGGUUUGGCAA 437 GGAAGACGCGACAUGUCAACA 445 UUGACAUGUCGCGUCUUCCUG 438 GCUCAUUGUCGAGGUCCUUGA 446 AAGgAcCuCGACAAUGAGCUC 439 GCUCAUUGUCGAGGUCCUUGA 447 AAgGaCCUCGACAAUGAGCUC 440 GCUCAUUGUCGAGGUCCUUGA 448 aAgGaCCUCGACAAUGAGCUC 441 GCUCAUUGUCGAGGUCCUUGA 449 AAggaccuCGACAAUGAGCUC 442 GCUCAUUGUCGAGGUCCUUGA 450 AAGGaccuCGACAAUGAGCUC

[00221] Key to Table 12: A, G, C, and U in uppercase refer to ribo-A, ribo-G, ribo-C, and ribo-U, respectively. lowercase d represents "deoxy".

[00222] In Table 12, the antisense strand of any of the RNAi molecules targeted to ARAF may alternatively have deoxynucleotides in a plurality of positions, the plurality of positions being one of the following: each of positions 4, 6, and 8 from the 5' end of the antisense strand; each of positions 3, 5 and 7 from the 5' end of the antisense strand; each of positions 1, 3, 5 and 7 from the 5' end of the antisense strand; each of positions 3-8 from the 5' end of the antisense strand; and each of positions 5-8 from the 5' end of the antisense strand.

[00223] RNA interference

[00224] In general, an siRNA can be about 21 to about 23 nucleotides in length and contain a base pair duplex region of about 19 nucleotides in length.

[00225] An endonuclease complex known as the RNA-induced silencing complex (RISC) is involved in RNAi. The siRNA has an antisense or guide strand that enters the RISC complex and mediates cleavage of a single-stranded target RNA having a sequence complementary to the antisense strand of the siRNA duplex. The other strand of miRNA is the accompanying strand. Cleavage of the target RNA occurs in the middle of the region complementary to the antisense strand of the miRNA duplex. See, for example, Elbashir et al., Genes & Development, 2001, Volume 15, pp. 188-200.

[00226] As used herein, the term "sense strand" refers to the nucleotide sequence of an siRNA molecule that is partially or fully complementary to at least a portion of the corresponding antisense strand of the siRNA molecule. The sense strand of an siRNA molecule may contain a nucleic acid sequence that shares homology with the target nucleic acid sequence.

[00227] As used herein, the term "antisense strand" refers to a nucleotide sequence of an siRNA molecule that is partially or fully complementary to at least a portion of a target nucleic acid sequence. The antisense strand of the siRNA molecule may contain a nucleic acid sequence that is complementary to at least a portion of the corresponding sense strand of the siRNA molecule.

[00228] RNAi molecules can down-regulate or knock down gene expression through the mediation of sequence-specific RNA interference. See, for example, Zamore et al., Cell, 2000, volume 101, pp. 25-33; Elbashir et al., Nature, 2001, vol. 411, pp. 494-498; Kreutzer et al., WO2000/044895; Zernicka-Goetz et al., WO2001/36646; Fire et al., WO1999/032619; Plaetinck et al., WO2000/01846; Mello et al., WO2001/029058.

[00229] As used herein, the terms "inhibit", "downregulate" or "reduce" in relation to gene expression means that there is a decrease in the expression of a gene or the level of mRNA molecules encoding one or more proteins, or the activity of one or more encoded proteins are lower than what is observed in the absence of the RNAi molecule or siRNA of the invention. For example, the expression level, mRNA level, or activity level of the encoded protein can be reduced by at least 1%, or at least 10%, or at least 20%, or at least 50%, or at least 90% or more of what is observed in the absence of the RNAi molecule or siRNA of the invention.

[00230] RNAi molecules can also be used to knock down viral gene expression and therefore influence viral replication.

[00231] RNAi molecules can be made from separate polynucleotide strands: a sense strand or accompanying strand and an antisense strand or guide strand. The guide and escort strands are at least partially complementary. The guide strand and the accompanying strand can form a duplex region having from about 15 to about 49 base pairs.

[00232] In some embodiments, the siRNA duplex region may have 17, 18, 19, 20, 21, 22, 23, 24, 25, 26, 27, 28, 29, 30, 31, 32, 33, 34, 35, 36, 37, 38, 39, 40, 41, 42, 43, 44, 45, 46, 47, 48, or 49 bp.

[00233] In certain embodiments, the RNAi molecule may be active in a RISC complex, wherein a segment of the duplex region is active for RISC.

[00234] In further embodiments, the RNAi molecule can be active as a dicer substrate for conversion to an RNAi molecule that can be active in a RISC complex.

[00235] In some aspects, an RNAi molecule may have complementary parts of the guide and accompanying sequences at opposite ends of the long molecule so that the molecule can form a duplex region from complementary sequence parts, and the strands are connected at one end of the duplex region by nucleotide or non-nucleotide linkers . For example, a hairpin structure or a stem and loop structure. Linker-chain interactions can be covalent bonds or non-covalent interactions.

[00236] An RNAi molecule of this disclosure may comprise a nucleotide, non-nucleotide, or mixed nucleotide/non-nucleotide linker that connects the sense region of the nucleic acid to the antisense region of the nucleic acid. The nucleotide linker may be a linker ≥2 nucleotides in length, for example about 3, 4, 5, 6, 7, 8, 9 or 10 nucleotides in length. The nucleotide linker may be a nucleic acid aptamer. By "aptamer" or "nucleic acid aptamer" as used herein, is meant a nucleic acid molecule that binds specifically to a target molecule, wherein the nucleic acid molecule has a sequence that contains a sequence recognized by the target molecule in its natural environment. Alternatively, an aptamer may be a nucleic acid molecule that binds to a target molecule, where the target molecule does not naturally bind to the nucleic acid. For example, an aptamer can be used to bind to the ligand-binding domain of a protein, thereby preventing the naturally occurring ligand from interacting with the protein. See, for example, Gold et al., Annu Rev Biochem, 1995, vol. 64, pp. 763-797; Brody et al., J. Biotechnol., 2000, vol. 74, pp. 5-13; Hermann et al., Science, 2000, volume 287, pp. 820-825.

[00237] Examples of a non-nucleotide linker include an abasic nucleotide, a polyether, a polyamine, a polyamide, a peptide, a carbohydrate, a lipid, a polyhydrocarbon, or other polymeric compounds such as polyethylene glycols, such as those having 2 to 100 ethylene glycol units. Some examples are described in Seela et al., Nucleic Acids Research, 1987, volume 15, pp. 3113-3129; Cload et al., J. Am. Chem. Soc., 1991, vol. 113, pp. 6324-6326; Jaeschke et al., Tetrahedron Lett., 1993, vol. 34, p. 301; Arnold et al., WO1989/002439; Usman et al., WO1995/006731; Dudycz et al., WO1995/011910 and Ferentz et al., J. Am. Chem. Soc., 1991, volume 113, pp. 4000-4002.

[00238] The RNAi molecule may have one or more sticky ends from the duplex region. Sticky ends, which are single-stranded regions of unpaired bases, can be from one to eight nucleotides in length or longer. The sticky end may be a 3' sticky end, where the 3' end of the strand is a single stranded region of one to eight nucleotides. The sticky end may be a 5' end sticky end, where the 5' end of the strand has a single stranded region of one to eight nucleotides.

[00239] The sticky ends of an RNAi molecule may be of the same length or may be of different lengths.

[00240] An RNAi molecule may have one or more blunt end nucleotides where the duplex region does not terminate in an overhang and the bases of the strands are paired to the end of the duplex region.

[00241] An RNAi molecule of this disclosure may have one or more blunt ends, or may have one or more sticky ends, or may have a combination of a blunt end and a sticky end.

[00242] The 5' end of the RNAi chain of the molecule may be at the blunt end or may be at the sticky end. The 3' end of the RNAi chain of the molecule may be at the blunt end or may be at the sticky end.

[00243] The 5' end of the RNAi chain of the molecule may be at the blunt end, while the 3' end is at the sticky end. The 3' end of the RNAi chain of the molecule may be at the blunt end, while the 5' end is at the sticky end.

[00244] In some embodiments, both ends of the RNAi molecule are blunt ends.

[00245] In additional embodiments, both ends of the RNAi molecule have a sticky end.

[00246] The sticky ends at the 5' and 3' ends can be of various lengths.

[00247] In certain embodiments, the RNAi molecule may have a blunt end, where the 5' end of the antisense strand and the 3' end of the sense strand do not have any sticky end nucleotides.

[00248] In further embodiments, the RNAi molecule may have a blunt end, where the 3' end of the antisense strand and the 5' end of the sense strand do not have any sticky end nucleotides.

[00249] An RNAi molecule may have mismatches in base pairing in the duplex region.

[00250] Any nucleotide at the sticky end of the RNAi molecule can be a deoxyribonucleotide or a ribonucleotide.

[00251] One or more deoxyribonucleotides may be at the 5' end, where the 3' end of the other strand of the RNAi molecule may not have an overhang or may not have a deoxyribonucleotide overhang.

[00252] One or more deoxyribonucleotides may be at the 3' end, where the 5' end of the other strand of the RNAi molecule may not have an overhang or may not have a deoxyribonucleotide overhang.

[00253] In some embodiments, one or more or all of the cohesive end nucleotides of the RNAi molecule may be 2'-deoxyribonucleotides.

[00254] Dicer substrate RNAi molecules

[00255] In some aspects, the RNAi molecule may be of a suitable length as a dicer substrate that can be processed to produce a RISC active RNAi molecule. See, for example, Rossi et al., US2005/0244858.

[00256] The double-stranded RNA (dsRNA) that is the substrate of the dicer may be of sufficient length that it is processed by the dicer to produce an active RNAi molecule, and may additionally have one or more of the following properties: (i) the dsRNA of the dicer substrate may be asymmetric , for example, having a sticky 3' end on the antisense strand, and (ii) the dsRNA of the dicer substrate may have a modified 3' end on the sense strand in order to determine the orientation of the dicer that binds and processes the dsRNA into an active RNAi molecule.

[00257] In certain embodiments, the longest strand in the dsRNA of a dicer substrate may be 24-30 nucleotides in length.

[00258] The dicer substrate dsRNA can be symmetrical or asymmetric.

[00259] In some embodiments, the Dicer substrate dsRNA may have a sense strand of 22-28 nucleotides and an antisense strand of 24-30 nucleotides.

[00260] In certain embodiments, the Dicer substrate dsRNA may have a sticky end at the 3' end of the antisense strand.

[00261] In additional embodiments, the Dicer substrate dsRNA may have a sense strand of 25 nucleotides in length and an antisense strand of 27 nucleotides in length, with a sticky 3′ end of 2 bases. The sticky end can be 1, 2 or 3 nucleotides in length. The sense strand may also have a 5'-phosphate.

[00262] A dicer substrate asymmetric dsRNA may have two deoxyribonucleotides at the 3' end of the sense strand instead of two ribonucleotides.

[00263] The sense strand of a dsRNA dicer substrate can be from about 22 to about 30, or from about 22 to about 28; or from about 24 to about 30; or from about 25 to about 30; or from about 26 to about 30; or about 26 and 29; or about 27 to about 28 nucleotides in length.

[00264] The sense strand of a dsRNA dicer substrate can be 22, 23, 24, 25, 26, 27, 28, 29, or 30 nucleotides in length.

[00265] In certain embodiments, the Dicer substrate dsRNA may have sense and antisense strands that are at least about 25 nucleotides in length and no more than about 30 nucleotides in length.

[00266] In certain embodiments, the Dicer substrate dsRNA may have sense and antisense strands that are 26 to 29 nucleotides in length.

[00267] In certain embodiments, a dicer substrate dsRNA may have sense and antisense strands that are 27 nucleotides in length.

[00268] The sense and antisense strands of a dsRNA substrate of a dicer may be the same length, as in blunt ends, or different lengths, as in sticky ends, or may have a blunt end and a sticky end.

[00269] A dicer substrate dsRNA may have a duplex region of 19, 20, 21, 22, 23, 24, 25, 26, or 27 nucleotides in length.

[00270] The antisense strand of a dsRNA dicer substrate may have any sequence that anneals to at least a portion of the sense strand sequence under biological conditions, such as in the cytoplasm of a eukaryotic cell.

[00271] The dicer substrate with the sense and antisense strands can be linked by a third structure such as a linker group or a linker oligonucleotide. A linker connects two strands in a dsRNA, for example, to form a hairpin upon annealing.

[00272] The sense and antisense strands of a dicer substrate are generally complementary, but may have mismatches in base pairing.

[00273] In some embodiments, the dsRNA of the dicer substrate may be asymmetric such that the sense strand is 22-28 nucleotides and the antisense strand is 24-30 nucleotides.

[00274] A region of one of the strands, in particular the antisense strand, of the dsRNA of the dicer substrate may have a sequence stretch of at least 19 nucleotides, where these nucleotides are in a 21 nucleotide region adjacent to the 3' end of the antisense strand and are sufficiently complementary the nucleotide sequence of the RNA obtained from the target gene.

[00275] The antisense dsRNA strand of a dicer substrate may have 1 to 9 ribonucleotides at the 5' end to give a length of 22-28 nucleotides. When the antisense strand is 21 nucleotides long, then 1-7 ribonucleotides, or 2-5 ribonucleotides, or 4 ribonucleotides can be added at the 3' end. The added ribonucleotides may be of any sequence.

[00276] The sense strand of a dsRNA dicer substrate may be 24-30 nucleotides. The sense strand may be substantially complementary to the antisense strand for annealing to the antisense strand under biological conditions.

[00277] Methods of using RNAi molecules

[00278] The nucleic acid molecules and RNAi molecules of this invention can be delivered to a cell or tissue by direct application of the molecule or by using the molecules in combination with a carrier or diluent.

[00279] Nucleic acid molecules and RNAi molecules of this invention can be delivered or introduced into a cell, tissue, organ, or subject by direct application of the molecules with a carrier or diluent, or any other delivery vehicle that acts to support, enhance, or facilitate entry into the cell , for example, viral sequences, viral substance, or lipid or liposomal formulations.

[00280] The nucleic acid molecules and RNAi molecules of this invention may be complexed with cationic lipids, may be packaged in liposomes, or may otherwise be delivered to target cells or tissues. The nucleic acid or nucleic acid complexes can be locally administered to relevant tissues ex vivo or in vivo via direct application to the skin, transdermal application or injection.

[00281] Delivery systems may include, for example, aqueous and non-aqueous gels, creams, emulsions, microemulsions, liposomes, ointments, aqueous and non-aqueous solutions, lotions, aerosols, hydrocarbon bases, and powders, and may contain excipients such as solubilizers and penetration enhancers.

[00282] The compositions and methods of this disclosure may include an expression vector that contains a nucleic acid sequence encoding at least one RNAi molecule of this invention in a manner that allows expression of the nucleic acid molecule.

[00283] The nucleic acid molecules and RNAi molecules of this invention can be expressed from transcription units inserted into DNA or RNA vectors. Recombinant vectors can be plasmid DNA or viral vectors. You can use viral vectors that provide transient expression of nucleic acid molecules.

[00284] For example, the vector may contain sequences encoding both strands of the RNAi duplex molecule, or one nucleic acid molecule that is self-complementary and thus forms an RNAi molecule. The expression vector may contain a nucleic acid sequence encoding two or more nucleic acid molecules.

[00285] A nucleic acid molecule can be expressed in cells from eukaryotic promoters. Those skilled in the art will appreciate that any nucleic acid can be expressed from a suitable DNA/RNA vector in eukaryotic cells.

[00286] In some aspects, a viral construct can be used to introduce an expression construct into a cell for transcription of the dsRNA construct encoded by the expression construct.

[00287] Lipid formulations can be administered to animals by intravenous, intramuscular, or intraperitoneal injection, or orally or by inhalation, or by other means known in the art.

[00288] Pharmaceutically acceptable formulations for administering oligonucleotides are known and can be used.

[00289] Methods of treating a disease

[00290] Examples of diseases include cancer, sarcomas, fibrosarcoma, malignant fibrous histiocytoma, liposarcoma, rhabdomyosarcoma, leiomyosarcoma, angiosarcoma, Kaposi's sarcoma, lymphangiosarcoma, synovial sarcoma, chondrosarcoma, osteosarcoma, and carcinomas.

[00291] In certain aspects, the methods of this invention may use inventive compounds to prevent or treat cancers and cancers in any organ or tissue, including, for example, brain tumors, head and neck cancers, breast cancers. lung cancer, esophageal cancer, gastric cancer, duodenal cancer, colon cancer, liver cancer, pancreatic cancer, gallbladder cancer, bile duct cancer, kidney cancer, urethra cancer, bladder cancer, prostate cancer, testicular cancer, uterine cancer, ovarian cancer, skin cancer, leukemia, malignant venous lymphoma, epithelial malignant tumors and non-epithelial malignant tumors.

[00292] Example of an in vitro knockdown protocol

[00293] One day prior to transfection, cells were seeded in a 96-well plate at 2×10 ³ cells per well with 100 μl of DMEM (HyClone Cat # SH30243.01) containing 10% FBS and cultured in a 37°C incubator, containing a humidified atmosphere of 5% CO ₂ in the air. Before transfection, the medium was replaced with 90 μl Opti-MEM I Reduced Serum Medium (Life Technologies, Cat # 31985-070) containing 2% FBS. Then 0.2 µl of Lipofectamine RNAiMax (Life Technologies, cat. no. 13778-100) was mixed with 4.8 µl of Opti-MEM I for 5 minutes at room temperature. Then 1 µl of siRNA was mixed with 4 µl of Opti-MEM I and combined with LF2000 solution and mixed gently without swirling. After 5 minutes at room temperature, the mixture was incubated for an additional 10 minutes at room temperature in order to allow the formation of RNA-RNAiMax complexes. In addition, 10 μl of RNA-RNAiMax complexes were added to the well and the plate was gently shaken by hand. Cells were incubated in a 37° C. incubator containing a humidified atmosphere of 5% CO ₂ in air for 2 hours. The medium was changed to fresh Opti-MEM I Reduced Serum Medium containing 2% FBS. 24 hours after transfection, cells were washed in ice-cold PBS once. Cells were lysed using 50 μl of Cell-to-Ct Lysis Buffer (Life Technologies, Cat # 4391851 C) for 5-30 minutes at room temperature. Added 5 μl Stop Solution and incubated for 2 minutes at room temperature. mRNA levels were immediately measured by RT-qPCR with TAQMAN. Samples could be frozen at -80°C and analyzed further.

[00294] Exemplary Protocol for Serum Stability

[00295] 0.2 mg/ml siRNA was incubated with 10% human serum at 37°C. At certain time points (0, 5, 15 and 30 min), a 200 μl aliquot of the sample was taken and extracted using 200 μl of extraction solvent (chloroform:phenol:isoamyl alcohol=24:25:1). The sample was vigorously mixed and centrifuged at 13,000 rpm for 10 min at RT, then the top layer solution was transferred and filtered through a 0.45 µm filter. The filtrate was transferred to a 300 μl HPLC injection vial. For LCMS, the mobile phase was MPA: 100 mM HFIP+7 mM TEA in H ₂ O, MPB: 50% methanol+50% acetonitrile. Column: Waters Acquity OST 2.1×50 mm, 1.7 µm.

[00296] EXAMPLES

[00297] Example 1: The siRNA of the invention directed to GST-π was found to be active for gene silencing in vitro. Dose dependent GST-π siRNA activities for gene knockdown were found to occur at IC _50s below about 250 picomoles (pM) and as low as 1 pM.

[00298] In vitro transfection was performed in the A549 cell line in order to determine the effect of siRNA knockdown. A dose-dependent knockdown for GST-π mRNA was observed using the siRNA from Table 1 as shown in Table 13.

Table 13: Dose-dependent knockdown for GST-π mRNA in the A549 cell line

miRNA structure IC ₅₀ (pM) A9 (SEQ ID Nos. 25 and 90) 24 B2 (SEQ ID Nos. 52 and 117) 121 B3 (SEQ ID Nos. 53 and 118) 235 B4 (SEQ ID Nos. 54 and 119) 229 B13 (SEQ ID Nos. 50 and 115) 17 BU2 (SEQ ID Nos. 61 and 126) 31

[00299] As shown in Table 13, the GST-π siRNA activities of Table 1 were in the range of 17-235 pM, which is suitable for many uses, including as a drug to be used in vivo.

[00300] Example 2: The GST-π siRNA structure of the present invention having deoxynucleotides located in the primer region of the antisense strand of the siRNA provided unexpectedly and beneficially increased in vitro gene knockdown activity.

[00301] In vitro transfection was performed in the A549 cell line in order to determine the knockdown effect for GST-π siRNA based on the BU2' structure (SEQ ID Nos. 131 and 157). Dose-dependent knockdown of GST-π mRNA was observed using GST-π siRNA based on the BU2' structure, as shown in Table 14.

Table 14: Dose-dependent knockdown of GST-π mRNA in cell line A549 for GST-π siRNA based on the BU2' structure

Structure of GST-π miRNA IC ₅₀ (pM) BU2 without deoxynucleotides in the duplex region (SEQ ID Nos. 61 and 126) 31 BU2 with deoxynucleotides at positions 3, 5 and 7 of the primer region of the antisense strand (SEQ ID Nos. 139 and 165) five BU2 with deoxynucleotides at positions 4, 6 and 8 of the primer region of the antisense strand (SEQ ID Nos. 141 and 167) 8 BU2 with deoxynucleotides at positions 4, 6 and 8 of the primer region of the antisense strand (SEQ ID Nos. 156 and 182) five

[00302] As shown in Table 14, the activities of GST-π siRNAs based on the BU2' structure having three deoxynucleotides in the primer region of the antisense strand are surprisingly and unexpectedly increased up to 6-fold compared to GST-π siRNAs without deoxynucleotides in the duplex region .

[00303] These data show that a GST-π siRNA having a structure with three deoxynucleotides located at positions 3, 5, and 7, or at positions 4, 6, and 8 in the primer region of the antisense strand, provided surprisingly increased gene knockdown activity compared to with GST-π siRNA without deoxynucleotides in the duplex region.

[00304] The activities shown in Table 14 for GST-π siRNAs having three deoxynucleotides in the primer region of the antisense strand were in the range of 5 to 8 pM, which is exceptionally suitable for many uses, including as a drug to be used in vivo.

[00305] Example 3: The GST-π siRNA structure of the present invention having deoxynucleotides located in the primer region of the antisense strand of the siRNA provided unexpectedly and beneficially increased in vitro gene knockdown activity.

[00306] In vitro transfection was performed in the A549 cell line in order to determine the knockdown effect for GST-π siRNA based on the A9' structure (SEQ ID Nos. 183 and 195). Dose-dependent knockdown of GST-π mRNA was observed using GST-π siRNA based on the A9' structure, as shown in Table 15.

Table 15: Dose-dependent knockdown of GST-π mRNA in the A549 cell line for GST-π siRNA based on the structure of structures* A9'

Structure of GST-π miRNA IC ₅₀ (pM) A9 without deoxynucleotides in the duplex region (SEQ ID Nos. 25 and 90) 24 A9 with deoxynucleotides at positions 4, 6 and 8 of the primer region of the antisense strand (SEQ ID Nos. 193 and 205) 1 A9 with deoxynucleotides at positions 1, 3, 5 and 7 of the primer region of the antisense strand (SEQ ID Nos. 190 and 202) five A9 with deoxynucleotides at positions 3-8 of the primer region of the antisense strand (SEQ ID Nos. 187 and 199) 6 A9 with deoxynucleotides at positions 5-8 of the primer region of the antisense strand (SEQ ID Nos. 189 and 201) 7 A9 with deoxynucleotides at positions 3, 5 and 7 of the primer region of the antisense strand (SEQ ID Nos. 191 and 203) 15

[00307] As shown in Table 15, the activities of GST-π siRNAs based on the A9' structure having three to six deoxynucleotides in the primer region of the antisense strand are surprisingly increased up to 24-fold compared to GST-π siRNAs without deoxynucleotides in the region duplex.

[00308] These data show that a GST-π siRNA having a structure with 3-6 deoxynucleotides located at positions 4, 6 and 8, or at positions 1, 3, 5 and 7, or at positions 3-8, or at positions 5-8, or positions 3, 5 and 7 in the primer region of the antisense strand, provided unexpectedly increased gene knockdown activity compared to GST-π siRNA without deoxynucleotides in the duplex region.

[00309] The activity reported in Table 15 for GST-π siRNA having three to six deoxynucleotides in the primer region of the antisense strand ranged from 1 to 15 pM, which is exceptionally suitable for many uses, including as a drug to be used in vivo.

[00310] Example 4: The GST-π siRNA structure having deoxynucleotides located in the primer region of the siRNA antisense strand provided unexpectedly and beneficially increased in vitro gene knockdown activity.

[00311] In vitro transfection was performed in the A549 cell line in order to determine the knockdown effect for GST-π siRNA based on the B13' structure (SEQ ID Nos. 207 and 222). Dose-dependent knockdown of GST-π mRNA was observed using GST-π siRNA based on the B13' structure, as shown in Table 16.

Table 16: Dose-dependent knockdown of GST-π mRNA in the A549 cell line for GST-π siRNA based on the B13' structure

Structure of GST-π miRNA IC ₅₀ (pM) B13 without deoxynucleotides in the duplex region (SEQ ID Nos. 50 and 115) 17 B13 with deoxynucleotides at positions 4, 6 and 8 of the primer region of the antisense strand (SEQ ID Nos. 217 and 232) eleven

[00312] As shown in Table 16, the activity of GST-π siRNA based on the B13' structure having three deoxynucleotides in the primer region of the antisense strand is unexpectedly increased compared to GST-π siRNA without deoxynucleotides in the duplex region.

[00313] These data demonstrate that a GST-π siRNA having a structure with three deoxynucleotides located at positions 4, 6, and 8 in the primer region of the antisense strand provided unexpectedly increased gene knockdown activity compared to a GST-π siRNA without deoxynucleotides in the antisense strand region. duplex.

[00314] The activity shown in Table 16 for GST-π siRNA having three deoxynucleotides in the primer region of the antisense strand was in the picomolar range and was 11 pM, which is exceptionally suitable for many uses, including as a drug to be used in vivo.

[00315] Example 5: The GST-π siRNA structure having deoxynucleotides located in the primer region of the antisense strand of the siRNA provided unexpectedly and beneficially increased in vitro gene knockdown activity.

[00316] In vitro transfection was performed in the A549 cell line in order to determine the knockdown effect for GST-π siRNA based on the B4' structure (SEQ ID Nos. 261 and 273). Dose-dependent knockdown of GST-π mRNA was observed using GST-π siRNA based on the B4' structure, as shown in Table 17.

Table 17: Dose-dependent knockdown of GST-π mRNA in the A549 cell line for GST-π siRNA based on the B4' structure

Structure of GST-π miRNA IC ₅₀ (pM) B4 without deoxynucleotides in the duplex region (SEQ ID Nos. 54 and 119) 229 B4 with deoxynucleotides at positions 3-8 of the primer region of the antisense strand (SEQ ID Nos. 265 and 277) 113

[00317] As shown in Table 17, the activities of GST-π siRNA based on the B4' structure having six deoxynucleotides in the primer region of the antisense strand are unexpectedly increased by more than 2-fold compared to GST-π siRNA without deoxynucleotides in the duplex region.

[00318] These data show that GST-π siRNA having a structure with six deoxynucleotides located at positions 3-8 in the primer region of the antisense strand provided surprisingly increased gene knockdown activity compared to GST-π siRNA without deoxynucleotides in the duplex region. .

[00319] The activity shown in Table 17 for GST-π siRNA having six deoxynucleotides in the primer region of the antisense strand was in the picomolar range and was 113 pM, which is exceptionally suitable for many uses, including as a drug to be used in vivo.

[00320] Example 6: The GST-π siRNA structure having deoxynucleotides located in the primer region of the siRNA antisense strand provided unexpectedly and beneficially increased in vitro gene knockdown activity.

[00321] In vitro transfection was performed in the A549 cell line in order to determine the knockdown effect for GST-π siRNA based on the B2' structure (SEQ ID Nos. 237 and 249). Dose-dependent knockdown of GST-π mRNA was observed using GST-π siRNA based on the B2' structure, as shown in Table 18.

Table 18: Dose-dependent knockdown of GST-π mRNA in the A549 cell line for GST-π siRNA based on the B2' structure

Structure of GST-π miRNA IC ₅₀ (pM) B2 without deoxynucleotides in the duplex region (SEQ ID Nos. 52 and 117) 121 B2 with deoxynucleotides at positions 5-8 of the primer region of the antisense strand (SEQ ID Nos. 243 and 255) thirty B2 with deoxynucleotides at positions 1, 3, 5 and 7 of the primer region of the antisense strand (SEQ ID Nos. 244 and 256) 50 B2 with deoxynucleotides at positions 3, 5 and 7 of the primer region of the antisense strand (SEQ ID Nos. 245 and 257) one hundred

[00322] As shown in Table 18, the activities of GST-π siRNA based on the B2' structure having three to four deoxynucleotides in the primer region of the antisense strand are surprisingly increased up to 4-fold compared to GST-π siRNAs without deoxynucleotides in the region duplex.

[00323] These data show that a GST-π siRNA having a structure with 3-4 deoxynucleotides located at positions 5-8, or at positions 1, 3, 5 and 7, or at positions 3, 5 and 7 in the primer region antisense strand provided unexpectedly increased gene knockdown activity compared to GST-π siRNA without deoxynucleotides in the duplex region.

[00324] The activities reported in Table 18 for GST-π siRNA having three to four deoxynucleotides in the primer region of the antisense strand were in the range of 30-100 pM, which is exceptionally suitable for many uses, including as a drug, to be used in vivo.

[00325] Example 7: A GST-π siRNA structure containing one or more 2'-deoxy-2'-fluoro-substituted nucleotides provided unexpectedly increased in vitro gene knockdown activity.

[00326] In vitro transfection was performed in the A549 cell line in order to determine the knockdown effect for GST-π siRNA based on the BU2' structure (SEQ ID Nos. 131 and 157). Dose-dependent knockdown of GST-π mRNA was observed using GST-π siRNA based on the BU2' structure, as shown in Table 19.

Table 19: Dose-dependent knockdown of GST-π mRNA in cell line A549 for GST-π siRNA based on the BU2' structure

Structure of GST-π miRNA IC ₅₀ (pM) BU2 without 2'-F-deoxynucleotides
(SEQ ID Nos. 61 and 126) 31 BU2 with seven 2'-F-deoxynucleotides, one at position 1 at the 3' end of the antisense strand (SEQ ID Nos. 148 and 174) 3 BU2 with four 2'-F-deoxynucleotides, one at position 1 at the 3' end of the antisense strand (SEQ ID Nos. 147 and 173) eleven BU2 with one 2'-F-deoxynucleotide at position 1 at the 3' end of the antisense strand (SEQ ID Nos. 144 and 170) 13

[00327] As shown in Table 19, the activities of GST-π siRNA based on the BU2' structure having one or more 2'-F-deoxynucleotides are surprisingly increased by up to 10-fold compared to GST-π siRNA without 2'-F -deoxynucleotides.

[00328] These data show that GST-π siRNA having a structure with one or more 2'-F-deoxynucleotides provided unexpectedly increased gene knockdown activity compared to GST-π siRNA without a 2'-F-deoxynucleotide.

[00329] The activities shown in Table 19 for GST-π siRNA having one or more 2'-F-deoxynucleotides ranged from 3 to 13 pM, which is exceptionally suitable for many uses, including as a drug, to be used in vivo.

[00330] Example 8: A GST-π siRNA structure containing one or more 2'-deoxy-2'-fluoro-substituted nucleotides provided unexpectedly increased in vitro gene knockdown activity.

[00331] In vitro transfection was performed in the A549 cell line in order to determine the knockdown effect for GST-π siRNA based on the B13' structure (SEQ ID Nos. 207 and 222). A dose-dependent knockdown of GST-π mRNA was observed using GST-π siRNA based on the B13' structure, as shown in Table 20.

Table 20: Dose-dependent knockdown of GST-π mRNA in the A549 cell line for GST-π siRNA based on the B13' structure

Structure of GST-π miRNA IC ₅₀ (pM) B13 without 2'-F-deoxynucleotides
(SEQ ID Nos. 50 and 115) 17 B13 with three 2'-F-deoxynucleotides located at positions outside the sticky ends (SEQ ID Nos. 219 and 234) 6

[00332] As shown in Table 20, the activity of GST-π siRNA based on the B13' structure having three 2'-F-deoxynucleotides located at positions outside the sticky ends is surprisingly increased approximately 3-fold compared to GST-π siRNA without 2'-F-deoxynucleotides.

[00333] These data show that GST-π siRNA having a structure with one or more 2'-F-deoxynucleotides provided unexpectedly increased gene knockdown activity compared to GST-π siRNA without a 2'-F-deoxynucleotide.

[00334] The activity presented in Table 20 for GST-π siRNA having one or more 2'-F-deoxynucleotides was in the picomolar range and was 6 pM, which is exceptionally suitable for many uses, including as a drug, to be used in vivo.

[00335] Example 9: A549 Orthotopic Lung Cancer Mouse Model. The GST-π siRNA of the invention can exhibit a marked reduction in orthotopic tumors from lung cancer in vivo. In this example, GST-π siRNA conferred in vivo gene knockdown activity when administered as a liposomal formulation to orthotopic lung cancer tumors in athymic nude mice.

[00336] In general, an orthotopic tumor model can exhibit immediate clinical relevance to drug effect and activity, as well as improved predictive ability. In the orthotopic tumor model, tumor cells are implanted directly into the specific organ from which the cells originate.

[00337] The antitumor effect of the siRNA formulation against human lung cancer A549 was assessed by comparing the final mass of primary tumors, measured at autopsy, for the treatment group and the vehicle control group.

[00338] FIG. 1 shows in vivo orthotopic tumor inhibition from lung cancer for GST-π siRNA based on the BU2 structure (SEQ ID Nos. 61 and 126). The orthotopic A549 lung cancer mouse model was used at a relatively low dose of 2 mg/kg siRNA directed to GST-π.

[00339] GST-π siRNA showed a significant and surprisingly beneficial lung tumor inhibitory effect in this 6 week study. As shown in FIG. 1, after 43 days GST-π siRNA showed a markedly favorable effect of tumor inhibition with a significant reduction in the final average masses of tumors by 2.8 times compared with the control.

[00340] Male NCr nu/nu mice aged 5-6 weeks were used for this study. The experimental animals were kept in a HEPA-filtered medium during the experimental period. Prior to use, the siRNA formulations were stored at 4° C. and warmed to room temperature 10 minutes prior to mouse injection.

[00341] For this orthotopic A549 human lung cancer model, on the day of surgical orthotopic implantation (SOI), stock tumors were harvested from the subcutaneous location of animals bearing an A549 tumor xenograft and placed in RPMI-1640 medium. Necrotic tissue was removed and viable tissue was cut into pieces of 1.5-2 mm ³ . Animals were anesthetized with isoflurane inhalation and the surgical field was sterilized with iodine and alcohol. An approximately 1.5 cm long transverse incision was made in the left chest wall of the mouse using a pair of surgical scissors. An intercostal incision was made between the third and fourth ribs and the lung was exposed. One A549 tumor fragment was transplanted to the lung surface using 8-0 surgical suture (nylon). The chest was closed using 6-0 surgical suture (silk). The lung was again inflated by intrathoracic puncture using a 3 cm ³ syringe with a 25 G × 1 1/2 needle in order to evacuate the air remaining in the chest cavity. The chest wall was closed with 6-0 surgical silk suture. All procedures of the operation described above were performed under a microscope at 7× magnification in a laminar flow cabinet with HEPA filtration.

[00342] Three days after tumor implantation, the model tumor bearing mice were randomly divided into groups of 10 mice per group. For the group of interest, treatment of 10 mice was started 3 days after tumor implantation.

[00343] For the group of interest, the composition was (ionizable lipid: cholesterol: DOPE: DOPC: DPPE-PEG-2K: DSPE-PEG-2K), liposome composition. Liposomes encapsulated GST-π siRNA.

[00344] At the end point of the study, experimental mice were sacrificed 42 days after the start of treatment. Primary tumors were excised and weighed on an electronic balance for further analysis.

[00345] To assess the toxicity of the compounds, the average body weight of mice in the treatment and control groups remained in the normal range during the entire experimental period. No other symptoms of toxicity were observed in mice.

[00346] Example 10: The GST-π siRNA of the present invention showed a marked reduction in xenograft cancers in vivo. GST-π siRNA provided gene knockdown activity in vivo when administered in a liposomal formulation into xenograft malignant tumors.

[00347] FIG. 2 shows the tumor inhibitory effect of GST-π siRNA (SEQ ID Nos. 156 and 182). A cancer xenograft model was used at a relatively low dose of 0.75 mg/kg siRNA directed to GST-π.

[00348] GST-π siRNA showed a significant and unexpectedly beneficial effect of tumor inhibition within a few days after administration. After 36 days, GST-π siRNA showed a markedly favorable effect of tumor inhibition with a 2-fold reduction in tumor volume compared to control.

[00349] As shown in FIG. 3, GST-π siRNA showed a significant and surprisingly beneficial effect of tumor inhibition on endpoint day. In particular, there was a decrease in tumor mass by more than 2 times.

[00350] GST-π siRNA was administered with two injections (days 1 and 15) of a liposomal formulation having the composition (ionizable lipid:cholesterol:DOPE:DOPC:DPPE-PEG-2K) (25:30:20:20:5).

[00351] For the cancer xenograft model, the A549 cell line was derived from ATCC. Cells were maintained in culture medium supplemented with 10% fetal bovine serum and 100 U/ml penicillin and 100 μg/ml streptomycin. Cells were separated 48 hours prior to inoculation so that the cells were in logarithmic growth phase when harvested. Cells were lightly trypsinized using trypsin-EDTA and harvested from tissue culture. The number of viable cells was counted and determined in a hemocytometer in the presence of trypan blue (only viable cells were counted). Cells were resuspended to a concentration of 5×10 ⁷ /ml in media without serum. The cell suspension was then mixed with ice-cold thawed BD Matrigel in a 1:1 ratio for injection.

[00352] Mice - female athymic nude (nu/nu) mice from Charles River Laboratory, immunodeficient, 6-8 weeks old, 7-8 mice per group.

[00353] To obtain a tumor model, each mouse was inoculated subcutaneously in the right flank with 0.1 ml of 2.5×10 ⁶ A549 cell inoculum using a 25 G needle and syringe, one inoculum per mouse. For inoculation, mice were anesthetized.

[00354] For tumor volume measurement and randomization, tumor size was measured to the nearest 0.1 mm. Tumor volume was calculated using the formula: tumor volume=length×width ^2/2 . When developed tumors reached approximately 120-175 mm ³ , the average tumor volume was approximately 150 mm ³ , mice were assigned to different vehicle and treatment groups such that the average tumor volumes in the treatment groups were within 10% of the average tumor volume in the control group carrier, ideally, CV% of tumor volume was less than 25%. On the same day, the test products and control vehicle were administered according to the dosing schedule. Monitoring of tumor volumes was carried out three times during week 1, twice during the remaining weeks, including on the day of completion of the study.

[00355] For dosing on dosing day, test articles were removed from the -80°C refrigerator and thawed on ice. Prior to transfer to syringes, the bottle containing the formulation was inverted by hand several times. All test articles were dosed at 0.75 mg/kg IV, q2w × 2, at 10 ml/kg.

[00356] For body weight, mice were weighed to the nearest 0.1 g. Body weight was monitored and recorded daily for 7 days after dosing the first dose. Body weight monitoring was carried out and it was recorded twice during the weeks, during the remaining weeks, including on the day of completion of the study.

[00357] When collecting tumors, 28 days after the first dosing, the volume of the tumor was measured, the tumor was excised for mass measurement and stored for PD study of biological markers. The mass of the tumor was recorded.

[00358] Example 11: The GST-π siRNA of the present invention showed increased cancer cell death by apoptosis of cancer cells in vitro. GST-π siRNA mediated GST-π knockdown, which led to upregulation of PUMA, a biological marker of apoptosis, and which is associated with loss of cell viability.

[00359] GST-π siRNA SEQ ID Nos. 156 and 182, which contained a combination of deoxynucleotides in the primer region, 2'-F-substituted deoxynucleotide, and 2'-OMe-substituted ribonucleotides, provided unexpectedly increased apoptosis of cancer cells.

[00360] The expression level of PUMA for GST-π siRNA SEQ ID NOs 156 and 182 was measured as shown in FIG. 4. In FIG. 4, PUMA expression is significantly reduced from days 2-4 post transfection with GST-π siRNA.

[00361] These data show that the structure of the GST-π siRNA containing a combination of deoxynucleotides in the primer region, 2'-F-substituted deoxynucleotide and 2'-OMe-substituted ribonucleotides, provided unexpectedly increased apoptosis of cancer cells.

[00362] The protocol for the biological marker PUMA was as follows. One day prior to transfection, cells were seeded in a 96-well plate at 2×10 ³ cells per well with 100 μl of DMEM (HyClone, catalog # SH30243.01) containing 10% FBS and cultured in a 37°C incubator containing humidified atmosphere 5% CO ₂ in the air. The next day, before transfection, the medium was changed to 90 μl Opti-MEM I Reduced Serum Medium (Life Technologies, Cat # 31985-070) containing 2% FBS. Then 0.2 µl of Lipofectamine RNAiMAX (Life Technologies, Cat # 13778-100) was mixed with 4.8 µl of Opti-MEM I for 5 minutes at room temperature. 1 µl of GST-π siRNA (stock conc. 1 µM) was mixed with 4 µl of Opti-MEM I and combined with the RNAiMAX solution and then mixed gently. The mixture was incubated for 10 minutes at room temperature in order to allow the formation of RNA-RNAiMAX complexes. 10 μl of RNA-RNAiMAX complexes were added to each well, to a final siRNA concentration of 10 nM. Cells were incubated for 2 hours and the medium was changed to fresh Opti-MEM I Reduced Serum Medium containing 2% FBS. On days 1, 2, 3, 4, and 6 post-transfection, cells were washed with ice-cold PBS once, and then lysed using 50 µl Cell-to-Ct Lysis Buffer (Life Technologies, cat# 4391851 C) for 5-30 minutes at room temperature. Added 5 μl Stop Solution and incubated for 2 minutes at room temperature. PUMA mRNA levels (BBC3, catalog # Hs00248075, Life Technologies) were measured by qPCR using TAQMAN.

[00363] Example 12: The GST-π siRNA of the present invention can demonstrate a marked reduction in xenograft cancers in vivo. GST-π siRNA can provide gene knockdown activity in vivo when administered in a liposome formulation to xenograft malignant tumors.

[00364] FIG. 5 shows the tumor inhibition effect of GST-π siRNA (SEQ ID Nos. 61 and 126). Dose-dependent knockdown of GST-π mRNA was observed in vivo using siRNA directed to GST-π. A cancer xenograft model was used at a relatively low dose of 0.75 mg/kg siRNA directed to GST-π.

[00365] GST-π siRNA showed a significant and unexpectedly beneficial effect of tumor inhibition within a few days after administration. As shown in FIG. 5, treatment with GST-π siRNA resulted in a significant decrease in GST-π mRNA expression at day 4 post-injection in the lipid formulation. At the higher dose of 4 mg/kg, a significant reduction of approximately 40% was found 24 hours after injection.

[00366] GST-π siRNA was administered with a single injection of 10 ml/kg of a liposomal formulation having the composition (ionizable lipid:cholesterol:DOPE:DOPC:DPPE-PEG-2K) (25:30:20:20:5).

[00367] For the cancer xenograft model, the A549 cell line was derived from ATCC. Cells were maintained in RPMI-1640 supplemented with 10% fetal bovine serum and 100 U/ml penicillin and 100 μg/ml streptomycin. Cells were separated 48 hours prior to inoculation so that the cells were in logarithmic growth phase when harvested. Cells were lightly trypsinized using trypsin-EDTA and harvested from tissue culture. The number of viable cells was counted and determined in a hemocytometer in the presence of trypan blue (only viable cells were counted). Cells were resuspended to a concentration of 4×10 ⁷ /ml in RPMI media without serum. The cell suspension was then well mixed with ice-cold thawed BD Matrigel in a 1:1 ratio for injection.

[00368] Mice - female athymic nude (nu/nu) mice from Charles River Laboratory, immunodeficient, 6-8 weeks old, 3 mice per group.

[00369] To obtain a tumor model, each mouse was inoculated subcutaneously in the right flank with 0.1 ml of 2x10 ⁶ A549 cell inoculum using a 25 G needle and syringe, one inoculum per mouse. For inoculation, mice were anesthetized.

[00370] For tumor volume measurement and randomization, tumor size was measured to the nearest 0.1 mm. Tumor volume was calculated using the formula: tumor volume=length×width ^2/2 . Tumor volume was monitored twice a week. When developed tumors reached approximately 350-600 mm ³ , mice were assigned to groups with different time points. On the same day, the test products were administered according to the dosing schedule.

[00371] For dosing, on the day when the developed tumors reached approximately 350-600 mm ³ , the test articles were taken from the refrigerator at 4°C. Before being transferred to syringes, the bottle containing the formulation was inverted by hand several times to obtain a homogeneous solution.

[00372] For body weight, mice were weighed to the nearest 0.1 g. Body weight was monitored and recorded twice during the weeks, during the remaining weeks, including the day of completion of the study.

[00373] When harvesting tumors, animals were sacrificed by CO ₂ overdose and tumors were excised at 0, 24, 48, 72, 96 (optional) and 168 hours post dosing. First, the wet weight of the tumors was determined, and then they were divided into three parts for the analysis of KD, distribution and biological markers. Samples were flash frozen in liquid nitrogen and stored at -80°C until ready for processing.

[00374] Example 13: The GST-π siRNA of the present invention inhibited xenograft tumors from pancreatic cancer in vivo. GST-π siRNA conferred in vivo gene knockdown activity when administered as a liposomal formulation into pancreatic cancer xenograft tumors.

[00375] In this xenograft model, each mouse was inoculated subcutaneously in the right flank with 0.1 ml of 2.5×10 ⁶ PANC-1 cell inoculum. Female athymic nude mice, 6-8 weeks old, Charles River were used. Tumor size was measured to the nearest 0.1 mm. When developed tumors reached approximately 150-250 mm ³ (average tumor volume approximately 200 mm ³ ), mice were assigned to different vehicle and treatment groups such that the average tumor volumes in the treatment groups were within 10% of the average tumor volume in the control group. carrier. On the same day, the test products and control vehicle were administered according to the dosing schedule. Monitoring of tumor volumes was carried out three times during week 1, twice during the remaining weeks, including on the day of completion of the study.

[00376] FIG. 6 shows the tumor inhibition effect of GST-π siRNA (SEQ ID Nos. 61 and 126). As shown in FIG. 6, dose effects were obtained at doses ranging from 0.375 mg/kg to 3 mg/kg of siRNA directed to GST-π. GST-π siRNA showed a significant and surprisingly beneficial tumor inhibition effect within days of administration. Thus, GST-π siRNA showed a significant and surprisingly beneficial endpoint tumor inhibition effect.

[00377] GST-π siRNA was administered in a liposomal formulation having the composition (ionizable lipid:cholesterol:DOPE:DOPC:DPPE-PEG-2K) (25:30:20:20:5).

[00378] Example 14: The GST-π siRNA of the invention showed increased serum stability.

[00379] FIG. 7 shows incubation in human serum and detection of remaining siRNA at various time points by HPLS/LCMS. As shown in FIG. 7, the serum half-life (t _1/2 ) for both the sense strand (FIG. 7, top) and antisense strand (FIG. 7, bottom) of GST-π siRNA (SEQ ID Nos. 61 and 126) was approximately 100 minutes.

[00380] Example 15: The GST-π siRNA of the present invention showed increased formulation stability in plasma.

[00381] FIG. 8 shows the incubation of the composition in plasma and the detection of the remaining siRNA at various time points. As shown in FIG. 8, the plasma half-life (t _1/2 ) of the GST-π siRNA formulation (SEQ ID Nos. 61 and 126) was well over 100 hours.

[00382] GST-π siRNA was prepared in a liposomal formulation having the composition (ionizable lipid:cholesterol:DOPE:DOPC:DPPE-PEG-2K) (25:30:20:20:5). The z-averaged size of the liposomal nanoparticles was 40.0 nm and the siRNA was 91% encapsulated.

[00383] The formulation was incubated in 50% human serum in PBS for 40 min, 1.5 h, 3 h, 24 h, and 96 h. GST-π siRNA was quantified by ELISA-based assay.

[00384] Example 16: The GST-π siRNA of the invention showed reduced off-target effects with the accompanying strand.

[00385] For GST-π siRNA (SEQ ID Nos. 156 and 182) in FIG. 9 shows that in vitro guide strand knockdown was approximately exponential compared to the shuffled sequence control, which showed no effect. The IC ₅₀ of this siRNA was measured to be 5 pM. In FIG. 10 shows in vitro knockdown for the accompanying strand of the same GST-π siRNA. As shown in FIG. 10, knockdown of the off-target follower strand for GST-π siRNA is significantly reduced, more than 100-fold.

[00386] For GST-π siRNAs (SEQ ID Nos. 187 and 199), (SEQ ID Nos. 189 and 201), and (SEQ ID Nos. 190 and 202) in FIG. 11 shows that in vitro knockdowns for guide chains were approximately exponential. The IC ₅₀ of these siRNAs were measured to be 6, 7 and 5 pM, respectively. As shown in FIG. 12, in vitro knockdowns for the follower strands of these GST-π siRNAs were significantly reduced by at least 10-fold. All of these GST-π siRNAs had deoxynucleotides in the primer region of the duplex region, with no other modifications in the duplex region.

[00387] For GST-π siRNA (SEQ ID Nos. 217 and 232) in FIG. 13 shows that the in vitro knockdown for the guide strand of this highly active GST-π siRNA was approximately exponential. The IC ₅₀ of this siRNA was measured to be 11 pM. As shown in FIG. 14, in vitro knockdown for the follower strand of this GST-π siRNA was significantly reduced by more than 100-fold. This GST-π siRNA had deoxynucleotides in the primer region of the duplex region, with no other modifications in the duplex region.

[00388] Off-target effects were determined using the psiCHECK-2 expression reporter plasmid, which encodes the Renilla luciferase gene. (Dual-Luciferase Reporter Assay System, Promega, Cat #: E1960). The siRNA concentration was usually 50 pM. Protocol: day 1, HeLa cells were seeded at 5-7.5×10 ³ /100 μl/well. Day 2, co-transfection at approximately 80% cell confluence. Day 3, cells were harvested to measure luciferase activity. Luciferase activity was measured using Promega's Luciferase Assay System (E4550), following the manufacturer's protocol.

[00389] The psiCHECK-2 vector allowed monitoring of changes in the expression of a target gene fused to the Renilla luciferase reporter gene. The siRNA construct was cloned into a multiple cloning site and the vector was co-transfected with the siRNA into HeLa cells. If a specific siRNA binds to a target mRNA and initiates the RNAi process, cleavage and subsequent degradation of the Renilla luciferase-mRNA fusion occurs, reducing the Renilla luciferase signal.

[00390] For example, BU2' siRNA plasmid inserts were as follows:

[00391] Insert psiCHECK-2 plasmid (F):

SEQ ID NO: 451

ctcgag gggcaacTGAAGCCTTTTGAGACCCTGcTgTcccag gcggccgc

[00392] Insert psiCHECK-2 (R) plasmid:

SEQ ID NO: 452

ctcgag cTgggacagCAGGGTCTCAAAAAGGCTTCagTTgccc gcggccgc

[00393] Example 17: The GST-π siRNAs of the present invention exhibited beneficially reduced off-target miRNA-like effects, which are primer-dependent off-target gene silencing.

[00394] For GST-π siRNAs (SEQ ID Nos. 156 and 182), (SEQ ID Nos. 187 and 199), (SEQ ID Nos. 189 and 201), (SEQ ID Nos. 190 and 202) and (SEQ ID Nos. 217 and 232) it was found that the activity of the mimic miRNA outside the target is essentially unimportant. The primer-dependent off-target unintended gene silencing for these GST-π siRNAs was at least 10-100 times less than the guide strand activity on the target.

[00395] To test miRNA-dependent off-target effects, one to four repeats of primed target sequences that are complementary to the entire primed-containing region, positions 1-8 from the 5' end of the antisense strand, but not to the remainder of the off-priming region, positions 9-21 were introduced into the region corresponding to the 3'UTR of the luciferase mRNA to determine the potency of seed-dependent off-target side effects. Plasmid inserts were used to mimic miRNAs with full matches in the primer region and mismatches (loops) outside the primer region.

[00396] For example, BU2' siRNA plasmid inserts were as follows:

[00397] Insert plasmid psiCHECK-2 (Fmi1):

SEQ ID NO: 453

ctcgag gggcaacTCTACGCAAAACAGACCCTGcTgTcccag gcggccgc

[00398] Insert plasmid psiCHECK-2 (Fmi2):

SEQ ID NO: 454

ctcgag gggcaacTCTACGCAAAACAGACCCTGcT CTACGCAAAACAGACCCTGcT gTcccag gcggccgc

[00399] Insert plasmid psiCHECK-2 (Fmi3):

SEQ ID NO: 455

ctcgag gggcaacTCTACGCAAAACAGACCCTGcT CTACGCAAAACAGACCCTGcT CTACGCAAAACAGACCCTGcT gTcccag gcggccgc

[00400] Insert plasmid psiCHECK-2 (Fmi4):

SEQ ID NO: 456

ctcgag gggcaacTCTACGCAAAACAGACCCTGcT CTACGCAAAACAGACCCTGcT CTACGCAAAACAGACCCTGcT CTACGCAAAACAGACCCTGcT gTcccag gcggccgc

[00401] The embodiments described herein are not limiting, and one of ordinary skill in the art will readily appreciate that specific combinations of the modifications described herein can be tested without undue experimentation towards identifying an improved RNAi nucleic acid molecule. activity.

[00402] Example 18: siRNAs of the invention directed to p21 were found to be active for gene silencing in vitro. The dose-dependent p21 siRNA activities for gene knockdown were found to exhibit an IC ₅₀ of less than about 3 picomoles (pM) and as little as 1 pM.

[00403] In vitro transfection was performed in the A549 cell line to determine the siRNA knockdown effect. A dose dependent knockdown for p21 mRNA was observed using the siRNA from Table 7 as shown in Table 21.

Table 21: Dose-dependent knockdown of p21 mRNA in the A549 cell line

p21 miRNA structure IC ₅₀ (pM) 1735 (SEQ ID Nos. 296 and 324) 0.3 2042 (SEQ ID Nos. 312 and 340) 10

[00404] As shown in Table 21, the p21 siRNA activities of Table 7 were in the range of 0.3-10 pM, which is suitable for many uses, including as a drug to be used in vivo.

[00405] Example 19: The p21 siRNA structure of the invention, having deoxynucleotides located in the primer region of the antisense strand of the siRNA, provided unexpectedly and beneficially increased gene knockdown activity.

[00406] In vitro transfection was performed in the A549 cell line in order to determine the knockdown effect for p21 siRNA based on the 1735' structure (SEQ ID Nos. 341 and 355). A dose-dependent knockdown of p21 mRNA was observed using p21 siRNA based on the 1735' structure, as shown in Table 22.

Table 22: Dose-dependent knockdown of p21 mRNA in the A549 cell line for p21 siRNA based on the 1735' structure

Structure of P21 siRNA IC ₅₀ (pM) 1735 without deoxynucleotides in the duplex region (SEQ ID Nos. 296 and 324) 0.3 1735 with deoxynucleotides at positions 4, 6, and 8 of the antisense strand primer region and additional 2'-Ome nucleotides (SEQ ID Nos. 342 and 356) 0.05 1735 with deoxynucleotides at positions 4, 6, and 8 of the antisense primer region and additional 2'-Ome nucleotides (SEQ ID NOs. 343 and 357) 0.001 1735 with deoxynucleotides at positions 4, 6, and 8 of the antisense primer region and additional 2'-ome nucleotides (SEQ ID NOs. 344 and 358) 0.1

[00407] As shown in Table 22, the activities of p21 siRNA based on structure 1735' having three deoxynucleotides in the primer region of the antisense strand are surprisingly and unexpectedly increased up to 300-fold compared to p21 siRNA without deoxynucleotides in the duplex region.

[00408] These data show that p21 siRNA having a structure with deoxynucleotides in the primer region of the antisense strand provided surprisingly increased gene knockdown activity compared to p21 siRNA without deoxynucleotides in the duplex region.

[00409] The activities shown in Table 22 for p21 siRNA having three deoxynucleotides in the primer region of the antisense strand ranged from 0.001 to 0.1 pM, which is exceptionally suitable for many uses, including as a drug subject to use in vivo.

[00410] Example 20: The p21 siRNA of the invention can demonstrate a marked reduction in xenograft cancers in vivo. p21 siRNA can provide gene knockdown activity in vivo when administered in a liposomal formulation to xenograft malignant tumors.

[00411] FIG. 15 shows the tumor inhibition effect for p21 siRNA (SEQ ID NOs: 341 and 355 where N=U). A cancer xenograft model was used at a relatively low dose of 0.75 mg/kg siRNA directed to p21.

[00412] p21 siRNA showed a significant and unexpectedly beneficial effect of tumor inhibition within a few days after injection. After 30 days, p21 siRNA showed a markedly favorable effect of tumor inhibition with a reduction in tumor volume of more than 2-fold compared to the control.

[00413] p21 siRNA was administered at a dose of 0.75 mg/kg by four injections of 10 ml/kg (days 1, 8, 15 and 22) of a liposomal formulation having the composition (ionizable lipid:cholesterol:DOPE:DOPC:DPPE-PEG-2K ) (25:30:20:20:5).

[00414] For the cancer xenograft model, the A549 cell line was derived from ATCC. Cells were maintained in culture medium supplemented with 10% fetal bovine serum and 100 U/ml penicillin and 100 μg/ml streptomycin. Cells were separated 48 hours prior to inoculation so that the cells were in logarithmic growth phase when harvested. Cells were lightly trypsinized using trypsin-EDTA and harvested from tissue culture. The number of viable cells was counted and determined in a hemocytometer in the presence of trypan blue (only viable cells were counted). Cells were resuspended to a concentration of 5×10 ⁷ /ml in media without serum. The cell suspension was then thoroughly mixed with ice-cold thawed BD Matrigel in a 1:1 ratio for injection.

[00415] Mice - female athymic nude mice (nu/nu) from Charles River Laboratory, immunodeficient, 6-8 weeks old, 7-8 mice per group.

[00416] To obtain a tumor model, each mouse was inoculated subcutaneously in the right flank with 0.1 ml of 2.5×10 ⁶ A549 cell inoculum using a 25 G needle and syringe, one inoculum per mouse. For inoculation, mice were anesthetized.

[00417] For tumor volume measurement and randomization, tumor size was measured to the nearest 0.1 mm. Tumor volume was calculated using the formula: tumor volume=length×width ^2/2 . When developed tumors reached approximately 120-175 mm ³ , the average tumor volume was approximately 150 mm ³ , mice were assigned to different vehicle and treatment groups such that the average tumor volumes in the treatment groups were within 10% of the average tumor volume in the control group carrier, ideally, CV% of tumor volume was less than 25%. On the same day, the test products and control vehicle were administered according to the dosing schedule. Monitoring of tumor volumes was carried out three times during week 1, twice during the remaining weeks, including on the day of completion of the study.

[00418] For dosing on dosing day, test articles were removed from the -80°C refrigerator and thawed on ice. Prior to transfer to syringes, the bottle containing the formulation was inverted by hand several times. All test articles were dosed at 0.75 mg/kg intravenously.

[00419] For body weight, mice were weighed to the nearest 0.1 g. Body weight was monitored and recorded daily for 7 days after dosing the first dose. Body weight monitoring was carried out and it was recorded twice during the weeks, during the remaining weeks, including on the day of completion of the study.

[00420] When collecting tumors, 28 days after the first dosing, the volume of the tumor was measured, the tumor was excised for mass measurement and stored for PD study of biological markers. The mass of the tumor was recorded.

[00421] The embodiments described herein are not limiting, and one of ordinary skill in the art will readily appreciate that specific combinations of the modifications described herein can be tested without undue experimentation towards identifying an improved RNAi nucleic acid molecule. activity.

[00422] Example 21: MCL1 directed siRNAs of the invention were prepared and found to be active for gene silencing in vitro. Dose-dependent MCL1 siRNA activities for gene knockdown were found to exhibit an IC _{50 of} less than about 100 picomoles (pM).

[00423] In vitro transfection was performed in the A549 cell line in order to determine the siRNA knockdown effect. Dose-dependent knockdown for MCL1 mRNA was observed using siRNA from Table 11.

[00424] Example 22: ARAF-targeted siRNAs of the invention were prepared and found to be active for gene silencing in vitro. Dose dependent ARAF siRNA activities for gene knockdown were found to exhibit an IC _{50 of} less than about 100 picomoles (pM).

[00425] In vitro transfection was performed in the A549 cell line in order to determine the siRNA knockdown effect. Dose-dependent knockdown for ARAF mRNA was observed using siRNA from Table 12.

[00426] All publications, patents and references specifically mentioned in the present description are incorporated into this description by reference in their entirety for all purposes.

[00427] It is understood that this invention is not limited to the specific methods, protocols, materials, and reagents described, as they may vary. It should also be understood that the terminology used in the present description is only for the purpose of describing specific embodiments and is not intended to limit the scope of the present invention. One skilled in the art will readily appreciate that various substitutions and modifications can be made to the description disclosed herein without departing from the spirit and scope of the description, and that these embodiments are within the scope of this description and the appended claims.

[00428] It should be noted that, as used herein and in the appended claims, the singular forms include the plural unless the context clearly dictates otherwise. Also, the singular forms and the terms "one or more" and "at least one" can be used interchangeably in the present description. It should also be noted that the terms "comprises", "comprising", "including" and "having" can be used interchangeably, and they are open reading and without limitation.

[00429] The listing of ranges of values in this specification is only intended as a shorthand for individually specifying each individual value falling within the range, unless otherwise noted herein, and each individual value is included in the specification as if it were individually specified in the present description. For Markush groups, those skilled in the art will appreciate that this description includes individual elements as well as subgroups of elements from the Markush group.

[00430] Without further elaboration, it is believed that a person skilled in the art can, based on the above description, use the present invention to its fullest extent. Therefore, the following specific embodiments are to be construed as illustrative only and not limiting the rest of the disclosure in any way at all.

[00431] All features disclosed in this description can be combined in any combination. Each feature disclosed in this specification can be replaced with an alternative feature that serves the same, equivalent, or similar purpose.

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SEQUENCE LIST

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<120> MIRNA STRUCTURES WITH HIGH ACTIVITY AND REDUCED IMPACT

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oligonucleotide

<220>

<223> Description of the combined DNA/RNA molecule: synthetic

oligonucleotide

<400> 25

ccuuuugaga cccugcugut t 21

<210> 26

<211> 21

<212> DNA

<213> Artificial sequence

<220>

<223> Artificial sequence description: synthetic

oligonucleotide

<220>

<223> Description of the combined DNA/RNA molecule: synthetic

oligonucleotide

<400> 26

cgccuuuuga gacccugcat t 21

<210> 27

<211> 21

<212> DNA

<213> Artificial sequence

<220>

<223> Artificial sequence description: synthetic

oligonucleotide

<220>

<223> Description of the combined DNA/RNA molecule: synthetic

oligonucleotide

<400> 27

ccuacaccgu ggucuauuut t 21

<210> 28

<211> 21

<212> DNA

<213> Artificial sequence

<220>

<223> Artificial sequence description: synthetic

oligonucleotide

<220>

<223> Description of the combined DNA/RNA molecule: synthetic

oligonucleotide

<400> 28

uggggagac cagaucucct t 21

<210> 29

<211> 21

<212> DNA

<213> Artificial sequence

<220>

<223> Artificial sequence description: synthetic

oligonucleotide

<220>

<223> Description of the combined DNA/RNA molecule: synthetic

oligonucleotide

<400> 29

gcgggaggca gaguuugcct t 21

<210> 30

<211> 21

<212> DNA

<213> Artificial sequence

<220>

<223> Artificial sequence description: synthetic

oligonucleotide

<220>

<223> Description of the combined DNA/RNA molecule: synthetic

oligonucleotide

<400> 30

ccuuucucca ggaccaauat t 21

<210> 31

<211> 21

<212> DNA

<213> Artificial sequence

<220>

<223> Artificial sequence description: synthetic

oligonucleotide

<220>

<223> Description of the combined DNA/RNA molecule: synthetic

oligonucleotide

<400> 31

acccugucugu cccagaacct t 21

<210> 32

<211> 21

<212> DNA

<213> Artificial sequence

<220>

<223> Artificial sequence description: synthetic

oligonucleotide

<220>

<223> Description of the combined DNA/RNA molecule: synthetic

oligonucleotide

<400> 32

ggucuauuuc ccaguucgat t 21

<210> 33

<211> 21

<212> DNA

<213> Artificial sequence

<220>

<223> Artificial sequence description: synthetic

oligonucleotide

<220>

<223> Description of the combined DNA/RNA molecule: synthetic

oligonucleotide

<400> 33

ccggugga cauggugaat t 21

<210> 34

<211> 21

<212> DNA

<213> Artificial sequence

<220>

<223> Artificial sequence description: synthetic

oligonucleotide

<220>

<223> Description of the combined DNA/RNA molecule: synthetic

oligonucleotide

<400> 34

acaucucccu caucuacact t 21

<210> 35

<211> 21

<212> DNA

<213> Artificial sequence

<220>

<223> Artificial sequence description: synthetic

oligonucleotide

<220>

<223> Description of the combined DNA/RNA molecule: synthetic

oligonucleotide

<400> 35

gcaaggauga cuaugugaat t 21

<210> 36

<211> 21

<212> DNA

<213> Artificial sequence

<220>

<223> Artificial sequence description: synthetic

oligonucleotide

<220>

<223> Description of the combined DNA/RNA molecule: synthetic

oligonucleotide

<400> 36

ccuucgcuga cuacaaccut t 21

<210> 37

<211> 21

<212> DNA

<213> Artificial sequence

<220>

<223> Artificial sequence description: synthetic

oligonucleotide

<220>

<223> Description of the combined DNA/RNA molecule: synthetic

oligonucleotide

<400> 37

cuggcagauc agggccagat t 21

<210> 38

<211> 21

<212> DNA

<213> Artificial sequence

<220>

<223> Artificial sequence description: synthetic

oligonucleotide

<220>

<223> Description of the combined DNA/RNA molecule: synthetic

oligonucleotide

<400> 38

gacggagacc ucacccugut t 21

<210> 39

<211> 21

<212> DNA

<213> Artificial sequence

<220>

<223> Artificial sequence description: synthetic

oligonucleotide

<220>

<223> Description of the combined DNA/RNA molecule: synthetic

oligonucleotide

<400> 39

cgggcaagga ugacuaugut t 21

<210> 40

<211> 21

<212> DNA

<213> Artificial sequence

<220>

<223> Artificial sequence description: synthetic

oligonucleotide

<220>

<223> Description of the combined DNA/RNA molecule: synthetic

oligonucleotide

<400> 40

cuuuugagac ccugcuguat t 21

<210> 41

<211> 21

<212> DNA

<213> Artificial sequence

<220>

<223> Artificial sequence description: synthetic

oligonucleotide

<220>

<223> Description of the combined DNA/RNA molecule: synthetic

oligonucleotide

<400> 41

gagcuggaag gaggagguat t 21

<210> 42

<211> 21

<212> DNA

<213> Artificial sequence

<220>

<223> Artificial sequence description: synthetic

oligonucleotide

<220>

<223> Description of the combined DNA/RNA molecule: synthetic

oligonucleotide

<400> 42

acccugucugu cccagaacat t 21

<210> 43

<211> 21

<212> DNA

<213> Artificial sequence

<220>

<223> Artificial sequence description: synthetic

oligonucleotide

<220>

<223> Description of the combined DNA/RNA molecule: synthetic

oligonucleotide

<400> 43

ugcugucca gaaccaggat t 21

<210> 44

<211> 21

<212> DNA

<213> Artificial sequence

<220>

<223> Artificial sequence description: synthetic

oligonucleotide

<220>

<223> Description of the combined DNA/RNA molecule: synthetic

oligonucleotide

<400> 44

agccuuuuga gacccugcat t 21

<210> 45

<211> 21

<212> DNA

<213> Artificial sequence

<220>

<223> Artificial sequence description: synthetic

oligonucleotide

<220>

<223> Description of the combined DNA/RNA molecule: synthetic

oligonucleotide

<400> 45

ccuuuugaga cccugcugat t 21

<210> 46

<211> 21

<212> DNA

<213> Artificial sequence

<220>

<223> Artificial sequence description: synthetic

oligonucleotide

<220>

<223> Description of the combined DNA/RNA molecule: synthetic

oligonucleotide

<400> 46

ugaagccuuu ugagacccut t 21

<210> 47

<211> 21

<212> DNA

<213> Artificial sequence

<220>

<223> Artificial sequence description: synthetic

oligonucleotide

<220>

<223> Description of the combined DNA/RNA molecule: synthetic

oligonucleotide

<400> 47

acugaagccu uuugagacct t 21

<210> 48

<211> 21

<212> DNA

<213> Artificial sequence

<220>

<223> Artificial sequence description: synthetic

oligonucleotide

<220>

<223> Description of the combined DNA/RNA molecule: synthetic

oligonucleotide

<400> 48

aggaugacua ugugaaggct t 21

<210> 49

<211> 21

<212> DNA

<213> Artificial sequence

<220>

<223> Artificial sequence description: synthetic

oligonucleotide

<220>

<223> Description of the combined DNA/RNA molecule: synthetic

oligonucleotide

<400> 49

ggaugacuau gugaaggcat t 21

<210> 50

<211> 21

<212> DNA

<213> Artificial sequence

<220>

<223> Artificial sequence description: synthetic

oligonucleotide

<220>

<223> Description of the combined DNA/RNA molecule: synthetic

oligonucleotide

<400> 50

gaugacuaug ugaaggcact t 21

<210> 51

<211> 21

<212> DNA

<213> Artificial sequence

<220>

<223> Artificial sequence description: synthetic

oligonucleotide

<220>

<223> Description of the combined DNA/RNA molecule: synthetic

oligonucleotide

<400> 51

cucccucauc uacaccaact t 21

<210> 52

<211> 21

<212> DNA

<213> Artificial sequence

<220>

<223> Artificial sequence description: synthetic

oligonucleotide

<220>

<223> Description of the combined DNA/RNA molecule: synthetic

oligonucleotide

<400> 52

gaagccuuuu gagacccugt t 21

<210> 53

<211> 21

<212> DNA

<213> Artificial sequence

<220>

<223> Artificial sequence description: synthetic

oligonucleotide

<220>

<223> Description of the combined DNA/RNA molecule: synthetic

oligonucleotide

<400> 53

ucucccucau cuacaccaat t 21

<210> 54

<211> 21

<212> DNA

<213> Artificial sequence

<220>

<223> Artificial sequence description: synthetic

oligonucleotide

<220>

<223> Description of the combined DNA/RNA molecule: synthetic

oligonucleotide

<400> 54

ccucaucuac accaacuaut t 21

<210> 55

<211> 21

<212> DNA

<213> Artificial sequence

<220>

<223> Artificial sequence description: synthetic

oligonucleotide

<220>

<223> Description of the combined DNA/RNA molecule: synthetic

oligonucleotide

<400> 55

cccucaucua caccaacuat t 21

<210> 56

<211> 21

<212> DNA

<213> Artificial sequence

<220>

<223> Artificial sequence description: synthetic

oligonucleotide

<220>

<223> Description of the combined DNA/RNA molecule: synthetic

oligonucleotide

<400> 56

caacugaagc cuuuugagat t 21

<210> 57

<211> 21

<212> DNA

<213> Artificial sequence

<220>

<223> Artificial sequence description: synthetic

oligonucleotide

<220>

<223> Description of the combined DNA/RNA molecule: synthetic

oligonucleotide

<400> 57

aacugaagcc uuuugagact t 21

<210> 58

<211> 21

<212> DNA

<213> Artificial sequence

<220>

<223> Artificial sequence description: synthetic

oligonucleotide

<220>

<223> Description of the combined DNA/RNA molecule: synthetic

oligonucleotide

<400> 58

cugaagccuu uugagaccct t 21

<210> 59

<211> 21

<212> DNA

<213> Artificial sequence

<220>

<223> Artificial sequence description: synthetic

oligonucleotide

<220>

<223> Description of the combined DNA/RNA molecule: synthetic

oligonucleotide

<400> 59

ucccucaucu acaccaacut t 21

<210> 60

<211> 21

<212> DNA

<213> Artificial sequence

<220>

<223> Artificial sequence description: synthetic

oligonucleotide

<220>

<223> Description of the combined DNA/RNA molecule: synthetic

oligonucleotide

<400> 60

gcucccucau cuacaccaat t 21

<210> 61

<211> 21

<212> DNA

<213> Artificial sequence

<220>

<223> Artificial sequence description: synthetic

oligonucleotide

<220>

<223> Description of the combined DNA/RNA molecule: synthetic

oligonucleotide

<400> 61

gaagccuuuu gagacccuat t 21

<210> 62

<211> 21

<212> DNA

<213> Artificial sequence

<220>

<223> Artificial sequence description: synthetic

oligonucleotide

<220>

<223> Description of the combined DNA/RNA molecule: synthetic

oligonucleotide

<400> 62

acugaagccu uuugagacat t 21

<210> 63

<211> 21

<212> DNA

<213> Artificial sequence

<220>

<223> Artificial sequence description: synthetic

oligonucleotide

<220>

<223> Description of the combined DNA/RNA molecule: synthetic

oligonucleotide

<400> 63

cucccucauc uacaccaaat t 21

<210> 64

<211> 21

<212> DNA

<213> Artificial sequence

<220>

<223> Artificial sequence description: synthetic

oligonucleotide

<220>

<223> Description of the combined DNA/RNA molecule: synthetic

oligonucleotide

<400> 64

ccucaucuac accaacuaatt 21

<210> 65

<211> 21

<212> DNA

<213> Artificial sequence

<220>

<223> Artificial sequence description: synthetic

oligonucleotide

<220>

<223> Description of the combined DNA/RNA molecule: synthetic

oligonucleotide

<400> 65

accaauaaaa uuucuaagat t 21

<210> 66

<211> 21

<212> DNA

<213> Artificial sequence

<220>

<223> Artificial sequence description: synthetic

oligonucleotide

<220>

<223> Description of the combined DNA/RNA molecule: synthetic

oligonucleotide

<400> 66

ugccucccug guucugggac a 21

<210> 67

<211> 21

<212> DNA

<213> Artificial sequence

<220>

<223> Artificial sequence description: synthetic

oligonucleotide

<220>

<223> Description of the combined DNA/RNA molecule: synthetic

oligonucleotide

<400> 67

gacagcaggg ucucaaaagg c 21

<210> 68

<211> 21

<212> DNA

<213> Artificial sequence

<220>

<223> Artificial sequence description: synthetic

oligonucleotide

<220>

<223> Description of the combined DNA/RNA molecule: synthetic

oligonucleotide

<400> 68

cucccugguu cugggacagc a 21

<210> 69

<211> 21

<212> DNA

<213> Artificial sequence

<220>

<223> Artificial sequence description: synthetic

oligonucleotide

<220>

<223> Description of the combined DNA/RNA molecule: synthetic

oligonucleotide

<400> 69

ccucccuggu ucugggacag c 21

<210> 70

<211> 21

<212> DNA

<213> Artificial sequence

<220>

<223> Artificial sequence description: synthetic

oligonucleotide

<220>

<223> Description of the combined DNA/RNA molecule: synthetic

oligonucleotide

<400> 70

gcagggucuc aaaaggcuuc a 21

<210> 71

<211> 21

<212> DNA

<213> Artificial sequence

<220>

<223> Artificial sequence description: synthetic

oligonucleotide

<220>

<223> Description of the combined DNA/RNA molecule: synthetic

oligonucleotide

<400> 71

ugggacagca gggucucaaa a 21

<210> 72

<211> 21

<212> DNA

<213> Artificial sequence

<220>

<223> Artificial sequence description: synthetic

oligonucleotide

<220>

<223> Description of the combined DNA/RNA molecule: synthetic

oligonucleotide

<400> 72

ggacagcagg gucucaaaag g 21

<210> 73

<211> 21

<212> DNA

<213> Artificial sequence

<220>

<223> Artificial sequence description: synthetic

oligonucleotide

<220>

<223> Description of the combined DNA/RNA molecule: synthetic

oligonucleotide

<400> 73

ucugggacag cagggucuca a 21

<210> 74

<211> 21

<212> DNA

<213> Artificial sequence

<220>

<223> Artificial sequence description: synthetic

oligonucleotide

<220>

<223> Description of the combined DNA/RNA molecule: synthetic

oligonucleotide

<400> 74

accaccuccu ccuuccagct c 21

<210> 75

<211> 21

<212> DNA

<213> Artificial sequence

<220>

<223> Artificial sequence description: synthetic

oligonucleotide

<220>

<223> Description of the combined DNA/RNA molecule: synthetic

oligonucleotide

<400> 75

caccaccucc uccuuccagc t 21

<210> 76

<211> 21

<212> DNA

<213> Artificial sequence

<220>

<223> Artificial sequence description: synthetic

oligonucleotide

<220>

<223> Description of the combined DNA/RNA molecule: synthetic

oligonucleotide

<400> 76

cuuccagcuc uggccugat c 21

<210> 77

<211> 21

<212> DNA

<213> Artificial sequence

<220>

<223> Artificial sequence description: synthetic

oligonucleotide

<220>

<223> Description of the combined DNA/RNA molecule: synthetic

oligonucleotide

<400> 77

cugggacagc agggucucaa a 21

<210> 78

<211> 21

<212> DNA

<213> Artificial sequence

<220>

<223> Artificial sequence description: synthetic

oligonucleotide

<220>

<223> Description of the combined DNA/RNA molecule: synthetic

oligonucleotide

<400> 78

uccuuccagc ucuggccug a 21

<210> 79

<211> 21

<212> DNA

<213> Artificial sequence

<220>

<223> Artificial sequence description: synthetic

oligonucleotide

<220>

<223> Description of the combined DNA/RNA molecule: synthetic

oligonucleotide

<400> 79

caccuccuc cuccagcuc t 21

<210> 80

<211> 21

<212> DNA

<213> Artificial sequence

<220>

<223> Artificial sequence description: synthetic

oligonucleotide

<220>

<223> Description of the combined DNA/RNA molecule: synthetic

oligonucleotide

<400> 80

uucugggaca gcagggucuc a 21

<210> 81

<211> 21

<212> DNA

<213> Artificial sequence

<220>

<223> Artificial sequence description: synthetic

oligonucleotide

<220>

<223> Description of the combined DNA/RNA molecule: synthetic

oligonucleotide

<400> 81

caccuccucc uuccagcuct g 21

<210> 82

<211> 21

<212> DNA

<213> Artificial sequence

<220>

<223> Artificial sequence description: synthetic

oligonucleotide

<220>

<223> Description of the combined DNA/RNA molecule: synthetic

oligonucleotide

<400> 82

cccugguucu gggacagcag g 21

<210> 83

<211> 21

<212> DNA

<213> Artificial sequence

<220>

<223> Artificial sequence description: synthetic

oligonucleotide

<220>

<223> Description of the combined DNA/RNA molecule: synthetic

oligonucleotide

<400> 83

uugccucccu gguucuggga c 21

<210> 84

<211> 21

<212> DNA

<213> Artificial sequence

<220>

<223> Artificial sequence description: synthetic

oligonucleotide

<220>

<223> Description of the combined DNA/RNA molecule: synthetic

oligonucleotide

<400> 84

cuugcccccc ugguucuggg a 21

<210> 85

<211> 21

<212> DNA

<213> Artificial sequence

<220>

<223> Artificial sequence description: synthetic

oligonucleotide

<220>

<223> Description of the combined DNA/RNA molecule: synthetic

oligonucleotide

<400> 85

gggacagcag ggucucaaaa g 21

<210> 86

<211> 21

<212> DNA

<213> Artificial sequence

<220>

<223> Artificial sequence description: synthetic

oligonucleotide

<220>

<223> Description of the combined DNA/RNA molecule: synthetic

oligonucleotide

<400> 86

guucugggac agcaggguct c 21

<210> 87

<211> 21

<212> DNA

<213> Artificial sequence

<220>

<223> Artificial sequence description: synthetic

oligonucleotide

<220>

<223> Description of the combined DNA/RNA molecule: synthetic

oligonucleotide

<400> 87

uccagcucug gcccugauct g 21

<210> 88

<211> 21

<212> DNA

<213> Artificial sequence

<220>

<223> Artificial sequence description: synthetic

oligonucleotide

<220>

<223> Description of the combined DNA/RNA molecule: synthetic

oligonucleotide

<400> 88

agcagggucu caaaaggcut c 21

<210> 89

<211> 21

<212> DNA

<213> Artificial sequence

<220>

<223> Artificial sequence description: synthetic

oligonucleotide

<220>

<223> Description of the combined DNA/RNA molecule: synthetic

oligonucleotide

<400> 89

cagcaggguc ucaaaaggct t 21

<210> 90

<211> 21

<212> DNA

<213> Artificial sequence

<220>

<223> Artificial sequence description: synthetic

oligonucleotide

<220>

<223> Description of the combined DNA/RNA molecule: synthetic

oligonucleotide

<400> 90

acagcagggu cucaaaaggc t 21

<210> 91

<211> 21

<212> DNA

<213> Artificial sequence

<220>

<223> Artificial sequence description: synthetic

oligonucleotide

<220>

<223> Description of the combined DNA/RNA molecule: synthetic

oligonucleotide

<400> 91

ugcagggucu caaaaggcgt c 21

<210> 92

<211> 21

<212> DNA

<213> Artificial sequence

<220>

<223> Artificial sequence description: synthetic

oligonucleotide

<220>

<223> Description of the combined DNA/RNA molecule: synthetic

oligonucleotide

<400> 92

aaauagacca cgguguaggg c 21

<210> 93

<211> 21

<212> DNA

<213> Artificial sequence

<220>

<223> Artificial sequence description: synthetic

oligonucleotide

<220>

<223> Description of the combined DNA/RNA molecule: synthetic

oligonucleotide

<400> 93

ggagaucugg ucucccacaa t 21

<210> 94

<211> 21

<212> DNA

<213> Artificial sequence

<220>

<223> Artificial sequence description: synthetic

oligonucleotide

<220>

<223> Description of the combined DNA/RNA molecule: synthetic

oligonucleotide

<400> 94

ggcaaacucu gccucccgct c 21

<210> 95

<211> 21

<212> DNA

<213> Artificial sequence

<220>

<223> Artificial sequence description: synthetic

oligonucleotide

<220>

<223> Description of the combined DNA/RNA molecule: synthetic

oligonucleotide

<400> 95

uauugguccu ggagaaagga a 21

<210> 96

<211> 21

<212> DNA

<213> Artificial sequence

<220>

<223> Artificial sequence description: synthetic

oligonucleotide

<220>

<223> Description of the combined DNA/RNA molecule: synthetic

oligonucleotide

<400> 96

gguucuggga cagcaggguc t 21

<210> 97

<211> 21

<212> DNA

<213> Artificial sequence

<220>

<223> Artificial sequence description: synthetic

oligonucleotide

<220>

<223> Description of the combined DNA/RNA molecule: synthetic

oligonucleotide

<400> 97

ucgaacuggg aaauagacca c 21

<210> 98

<211> 21

<212> DNA

<213> Artificial sequence

<220>

<223> Artificial sequence description: synthetic

oligonucleotide

<220>

<223> Description of the combined DNA/RNA molecule: synthetic

oligonucleotide

<400> 98

uucaccaugu ccaccagggc t 21

<210> 99

<211> 21

<212> DNA

<213> Artificial sequence

<220>

<223> Artificial sequence description: synthetic

oligonucleotide

<220>

<223> Description of the combined DNA/RNA molecule: synthetic

oligonucleotide

<400> 99

guguagauga gggagaugua t 21

<210> 100

<211> 21

<212> DNA

<213> Artificial sequence

<220>

<223> Artificial sequence description: synthetic

oligonucleotide

<220>

<223> Description of the combined DNA/RNA molecule: synthetic

oligonucleotide

<400> 100

uucacauagu cauccuugcc c 21

<210> 101

<211> 21

<212> DNA

<213> Artificial sequence

<220>

<223> Artificial sequence description: synthetic

oligonucleotide

<220>

<223> Description of the combined DNA/RNA molecule: synthetic

oligonucleotide

<400> 101

agguuguagu cagcgaagga g 21

<210> 102

<211> 21

<212> DNA

<213> Artificial sequence

<220>

<223> Artificial sequence description: synthetic

oligonucleotide

<220>

<223> Description of the combined DNA/RNA molecule: synthetic

oligonucleotide

<400> 102

ucuggccug aucugccagc a 21

<210> 103

<211> 21

<212> DNA

<213> Artificial sequence

<220>

<223> Artificial sequence description: synthetic

oligonucleotide

<220>

<223> Description of the combined DNA/RNA molecule: synthetic

oligonucleotide

<400> 103

acagggugag gucuccgucc t 21

<210> 104

<211> 21

<212> DNA

<213> Artificial sequence

<220>

<223> Artificial sequence description: synthetic

oligonucleotide

<220>

<223> Description of the combined DNA/RNA molecule: synthetic

oligonucleotide

<400> 104

acauagucau ccuugcccgc c 21

<210> 105

<211> 21

<212> DNA

<213> Artificial sequence

<220>

<223> Artificial sequence description: synthetic

oligonucleotide

<220>

<223> Description of the combined DNA/RNA molecule: synthetic

oligonucleotide

<400> 105

uacagcaggg ucucaaaagg c 21

<210> 106

<211> 21

<212> DNA

<213> Artificial sequence

<220>

<223> Artificial sequence description: synthetic

oligonucleotide

<220>

<223> Description of the combined DNA/RNA molecule: synthetic

oligonucleotide

<400> 106

uaccuccucc uuccagcuct g 21

<210> 107

<211> 21

<212> DNA

<213> Artificial sequence

<220>

<223> Artificial sequence description: synthetic

oligonucleotide

<220>

<223> Description of the combined DNA/RNA molecule: synthetic

oligonucleotide

<400> 107

uguucuggga cagcaggguc t 21

<210> 108

<211> 21

<212> DNA

<213> Artificial sequence

<220>

<223> Artificial sequence description: synthetic

oligonucleotide

<220>

<223> Description of the combined DNA/RNA molecule: synthetic

oligonucleotide

<400> 108

uccugguucu gggacagcag g 21

<210> 109

<211> 21

<212> DNA

<213> Artificial sequence

<220>

<223> Artificial sequence description: synthetic

oligonucleotide

<220>

<223> Description of the combined DNA/RNA molecule: synthetic

oligonucleotide

<400> 109

ugcagggucu caaaaggcut c 21

<210> 110

<211> 21

<212> DNA

<213> Artificial sequence

<220>

<223> Artificial sequence description: synthetic

oligonucleotide

<220>

<223> Description of the combined DNA/RNA molecule: synthetic

oligonucleotide

<400> 110

ucagcagggu cucaaaaggc t 21

<210> 111

<211> 21

<212> DNA

<213> Artificial sequence

<220>

<223> Artificial sequence description: synthetic

oligonucleotide

<220>

<223> Description of the combined DNA/RNA molecule: synthetic

oligonucleotide

<400> 111

agggucucaa aaggcuucag t 21

<210> 112

<211> 21

<212> DNA

<213> Artificial sequence

<220>

<223> Artificial sequence description: synthetic

oligonucleotide

<220>

<223> Description of the combined DNA/RNA molecule: synthetic

oligonucleotide

<400> 112

ggucucaaaa ggcuucagut g 21

<210> 113

<211> 21

<212> DNA

<213> Artificial sequence

<220>

<223> Artificial sequence description: synthetic

oligonucleotide

<220>

<223> Description of the combined DNA/RNA molecule: synthetic

oligonucleotide

<400> 113

gccuucacau agucauccut g 21

<210> 114

<211> 21

<212> DNA

<213> Artificial sequence

<220>

<223> Artificial sequence description: synthetic

oligonucleotide

<220>

<223> Description of the combined DNA/RNA molecule: synthetic

oligonucleotide

<400> 114

ugccuucaca uagucaucct t 21

<210> 115

<211> 21

<212> DNA

<213> Artificial sequence

<220>

<223> Artificial sequence description: synthetic

oligonucleotide

<220>

<223> Description of the combined DNA/RNA molecule: synthetic

oligonucleotide

<400> 115

gugccuucac auagucaucc t 21

<210> 116

<211> 21

<212> DNA

<213> Artificial sequence

<220>

<223> Artificial sequence description: synthetic

oligonucleotide

<220>

<223> Description of the combined DNA/RNA molecule: synthetic

oligonucleotide

<400> 116

guugguguag augagggaga t 21

<210> 117

<211> 21

<212> DNA

<213> Artificial sequence

<220>

<223> Artificial sequence description: synthetic

oligonucleotide

<220>

<223> Description of the combined DNA/RNA molecule: synthetic

oligonucleotide

<400> 117

cagggucuca aaaggcuuca g 21

<210> 118

<211> 21

<212> DNA

<213> Artificial sequence

<220>

<223> Artificial sequence description: synthetic

oligonucleotide

<220>

<223> Description of the combined DNA/RNA molecule: synthetic

oligonucleotide

<400> 118

uugguguaga ugagggat g 21

<210> 119

<211> 21

<212> DNA

<213> Artificial sequence

<220>

<223> Artificial sequence description: synthetic

oligonucleotide

<220>

<223> Description of the combined DNA/RNA molecule: synthetic

oligonucleotide

<400> 119

auaguuggug uagaugaggg a 21

<210> 120

<211> 21

<212> DNA

<213> Artificial sequence

<220>

<223> Artificial sequence description: synthetic

oligonucleotide

<220>

<223> Description of the combined DNA/RNA molecule: synthetic

oligonucleotide

<400> 120

uaguuggugu agaugaggga g 21

<210> 121

<211> 21

<212> DNA

<213> Artificial sequence

<220>

<223> Artificial sequence description: synthetic

oligonucleotide

<220>

<223> Description of the combined DNA/RNA molecule: synthetic

oligonucleotide

<400> 121

ucucaaaagg cuucaguugc c 21

<210> 122

<211> 21

<212> DNA

<213> Artificial sequence

<220>

<223> Artificial sequence description: synthetic

oligonucleotide

<220>

<223> Description of the combined DNA/RNA molecule: synthetic

oligonucleotide

<400> 122

gucucaaaag gcuucaguug c 21

<210> 123

<211> 21

<212> DNA

<213> Artificial sequence

<220>

<223> Artificial sequence description: synthetic

oligonucleotide

<220>

<223> Description of the combined DNA/RNA molecule: synthetic

oligonucleotide

<400> 123

gggucucaaa aggcuucagt t 21

<210> 124

<211> 21

<212> DNA

<213> Artificial sequence

<220>

<223> Artificial sequence description: synthetic

oligonucleotide

<220>

<223> Description of the combined DNA/RNA molecule: synthetic

oligonucleotide

<400> 124

aguuggugua gaugagggag a 21

<210> 125

<211> 21

<212> DNA

<213> Artificial sequence

<220>

<223> Artificial sequence description: synthetic

oligonucleotide

<220>

<223> Description of the combined DNA/RNA molecule: synthetic

oligonucleotide

<400> 125

uugguguaga ugagggagct g 21

<210> 126

<211> 21

<212> DNA

<213> Artificial sequence

<220>

<223> Artificial sequence description: synthetic

oligonucleotide

<220>

<223> Description of the combined DNA/RNA molecule: synthetic

oligonucleotide

<400> 126

uagggucuca aaaggcuuca g 21

<210> 127

<211> 21

<212> DNA

<213> Artificial sequence

<220>

<223> Artificial sequence description: synthetic

oligonucleotide

<220>

<223> Description of the combined DNA/RNA molecule: synthetic

oligonucleotide

<400> 127

ugucucaaaa ggcuucagut g 21

<210> 128

<211> 21

<212> DNA

<213> Artificial sequence

<220>

<223> Artificial sequence description: synthetic

oligonucleotide

<220>

<223> Description of the combined DNA/RNA molecule: synthetic

oligonucleotide

<400> 128

uuugguguag augagggaga t 21

<210> 129

<211> 21

<212> DNA

<213> Artificial sequence

<220>

<223> Artificial sequence description: synthetic

oligonucleotide

<220>

<223> Description of the combined DNA/RNA molecule: synthetic

oligonucleotide

<400> 129

uuaguuggug uagaugaggg a 21

<210> 130

<211> 21

<212> DNA

<213> Artificial sequence

<220>

<223> Artificial sequence description: synthetic

oligonucleotide

<220>

<223> Description of the combined DNA/RNA molecule: synthetic

oligonucleotide

<400> 130

ucuuagaaau uuuauugguc c 21

<210> 131

<211> 21

<212> DNA

<213> Artificial sequence

<220>

<223> Artificial sequence description: synthetic

oligonucleotide

<220>

<223> Description of the combined DNA/RNA molecule: synthetic

oligonucleotide

<220>

<221> modified base

<222> (20)..(21)

<223> a, c, t, g, u, unknown or other

<400> 131

gaagccuuuu gagacccuan n 21

<210> 132

<211> 21

<212> RNA

<213> Artificial sequence

<220>

<223> Artificial sequence description: synthetic

oligonucleotide

<220>

<221> modified base

<222> (20)..(21)

<223> 2'-OMe-nucleotide

<400> 132

gaagccuuuu gagacccuau u 21

<210> 133

<211> 21

<212> RNA

<213> Artificial sequence

<220>

<223> Artificial sequence description: synthetic

oligonucleotide

<220>

<221> modified base

<222> (20)..(21)

<223> 2'-OMe-nucleotide

<400> 133

gaagccuuuu gagacccuau u 21

<210> 134

<211> 21

<212> RNA

<213> Artificial sequence

<220>

<223> Artificial sequence description: synthetic

oligonucleotide

<220>

<221> modified base

<222> (20)..(21)

<223> 2'-OMe-nucleotide

<400> 134

gaagccuuuu gagacccuau u 21

<210> 135

<211> 21

<212> RNA

<213> Artificial sequence

<220>

<223> Artificial sequence description: synthetic

oligonucleotide

<220>

<221> modified base

<222> (20)..(21)

<223> 2'-OMe-nucleotide

<400> 135

gaagccuuuu gagacccuau u 21

<210> 136

<211> 21

<212> RNA

<213> Artificial sequence

<220>

<223> Artificial sequence description: synthetic

oligonucleotide

<220>

<221> modified base

<222> (20)..(21)

<223> 2'-OMe-nucleotide

<400> 136

gaagccuuuu gagacccuau u 21

<210> 137

<211> 21

<212> RNA

<213> Artificial sequence

<220>

<223> Artificial sequence description: synthetic

oligonucleotide

<220>

<221> modified base

<222> (20)..(21)

<223> 2'-OMe-nucleotide

<400> 137

gaagccuuuu gagacccuau u 21

<210> 138

<211> 21

<212> RNA

<213> Artificial sequence

<220>

<223> Artificial sequence description: synthetic

oligonucleotide

<220>

<221> modified base

<222> (20)..(21)

<223> 2'-OMe-nucleotide

<400> 138

gaagccuuuu gagacccuau u 21

<210> 139

<211> 21

<212> RNA

<213> Artificial sequence

<220>

<223> Artificial sequence description: synthetic

oligonucleotide

<220>

<221> modified base

<222> (20)..(21)

<223> 2'-OMe-nucleotide

<400> 139

gaagccuuuu gagacccuau u 21

<210> 140

<211> 21

<212> RNA

<213> Artificial sequence

<220>

<223> Artificial sequence description: synthetic

oligonucleotide

<220>

<221> modified base

<222> (20)..(21)

<223> 2'-OMe-nucleotide

<400> 140

gaagccuuuu gagacccuau u 21

<210> 141

<211> 21

<212> RNA

<213> Artificial sequence

<220>

<223> Artificial sequence description: synthetic

oligonucleotide

<220>

<221> modified base

<222> (20)..(21)

<223> 2'-OMe-nucleotide

<400> 141

gaagccuuuu gagacccuau u 21

<210> 142

<211> 21

<212> DNA

<213> Artificial sequence

<220>

<223> Artificial sequence description: synthetic

oligonucleotide

<220>

<223> Description of the combined DNA/RNA molecule: synthetic

oligonucleotide

<400> 142

gaagccuuuu gagacccuat t 21

<210> 143

<211> 21

<212> RNA

<213> Artificial sequence

<220>

<223> Artificial sequence description: synthetic

oligonucleotide

<220>

<221> modified base

<222> (20)..(21)

<223> 2'-OMe-nucleotide

<400> 143

gaagccuuuu gagacccuau u 21

<210> 144

<211> 21

<212> RNA

<213> Artificial sequence

<220>

<223> Artificial sequence description: synthetic

oligonucleotide

<220>

<221> modified base

<222> (20)..(21)

<223> 2'-OMe-nucleotide

<400> 144

gaagccuuuu gagacccuau u 21

<210> 145

<211> 21

<212> RNA

<213> Artificial sequence

<220>

<223> Artificial sequence description: synthetic

oligonucleotide

<220>

<221> modified base

<222> (20)..(21)

<223> 2'-OMe-nucleotide

<400> 145

gaagccuuuu gagacccuau u 21

<210> 146

<211> 21

<212> RNA

<213> Artificial sequence

<220>

<223> Artificial sequence description: synthetic

oligonucleotide

<220>

<221> modified base

<222> (20)..(21)

<223> 2'-OMe-nucleotide

<400> 146

gaagccuuuu gagacccuau u 21

<210> 147

<211> 21

<212> RNA

<213> Artificial sequence

<220>

<223> Artificial sequence description: synthetic

oligonucleotide

<220>

<221> modified base

<222> (19)..(19)

<223> 2'-deoxy-2'-fluoronucleotide

<220>

<221> modified base

<222> (20)..(21)

<223> 2'-OMe-nucleotide

<400> 147

gaagccuuuu gagacccuau u 21

<210> 148

<211> 21

<212> RNA

<213> Artificial sequence

<220>

<223> Artificial sequence description: synthetic

oligonucleotide

<220>

<221> modified base

<222> (14)..(14)

<223> 2'-deoxy-2'-fluoronucleotide

<220>

<221> modified base

<222> (19)..(19)

<223> 2'-deoxy-2'-fluoronucleotide

<220>

<221> modified base

<222> (20)..(21)

<223> 2'-OMe-nucleotide

<400> 148

gaagccuuuu gagacccuau u 21

<210> 149

<211> 21

<212> RNA

<213> Artificial sequence

<220>

<223> Artificial sequence description: synthetic

oligonucleotide

<220>

<221> modified base

<222> (19)..(21)

<223> 2'-OMe-nucleotide

<400> 149

gaagccuuuu gagacccuau u 21

<210> 150

<211> 21

<212> RNA

<213> Artificial sequence

<220>

<223> Artificial sequence description: synthetic

oligonucleotide

<220>

<221> modified base

<222> (19)..(21)

<223> 2'-OMe-nucleotide

<400> 150

gaagccuuuu gagacccuau u 21

<210> 151

<211> 21

<212> RNA

<213> Artificial sequence

<220>

<223> Artificial sequence description: synthetic

oligonucleotide

<220>

<221> modified base

<222> (1)..(3)

<223> 2'-OMe-nucleotide

<220>

<221> modified base

<222> (20)..(21)

<223> 2'-OMe-nucleotide

<400> 151

gaagccuuuu gagacccuau u 21

<210> 152

<211> 21

<212> RNA

<213> Artificial sequence

<220>

<223> Artificial sequence description: synthetic

oligonucleotide

<220>

<221> modified base

<222> (1)..(3)

<223> 2'-OMe-nucleotide

<220>

<221> modified base

<222>(8)..(8)

<223> 2'-OMe-nucleotide

<220>

<221> modified base

<222> (10)..(10)

<223> 2'-OMe-nucleotide

<220>

<221> modified base

<222> (18)..(18)

<223> 2'-OMe-nucleotide

<220>

<221> modified base

<222> (20)..(21)

<223> 2'-OMe-nucleotide

<400> 152

gaagccuuuu gagacccuau u 21

<210> 153

<211> 21

<212> RNA

<213> Artificial sequence

<220>

<223> Artificial sequence description: synthetic

oligonucleotide

<220>

<221> modified base

<222> (1)..(3)

<223> 2'-OMe-nucleotide

<220>

<221> modified base

<222> (20)..(21)

<223> 2'-OMe-nucleotide

<400> 153

gaagccuuuu gagacccuau u 21

<210> 154

<211> 21

<212> RNA

<213> Artificial sequence

<220>

<223> Artificial sequence description: synthetic

oligonucleotide

<220>

<221> modified base

<222> (1)..(3)

<223> 2'-OMe-nucleotide

<220>

<221> modified base

<222>(8)..(8)

<223> 2'-OMe-nucleotide

<220>

<221> modified base

<222> (10)..(10)

<223> 2'-OMe-nucleotide

<220>

<221> modified base

<222> (18)..(18)

<223> 2'-OMe-nucleotide

<220>

<221> modified base

<222> (20)..(21)

<223> 2'-OMe-nucleotide

<400> 154

gaagccuuuu gagacccuau u 21

<210> 155

<211> 21

<212> RNA

<213> Artificial sequence

<220>

<223> Artificial sequence description: synthetic

oligonucleotide

<220>

<221> modified base

<222> (1)..(3)

<223> 2'-OMe-nucleotide

<220>

<221> modified base

<222>(8)..(8)

<223> 2'-OMe-nucleotide

<220>

<221> modified base

<222> (10)..(10)

<223> 2'-OMe-nucleotide

<220>

<221> modified base

<222> (18)..(21)

<223> 2'-OMe-nucleotide

<400> 155

gaagccuuuu gagacccuau u 21

<210> 156

<211> 21

<212> RNA

<213> Artificial sequence

<220>

<223> Artificial sequence description: synthetic

oligonucleotide

<220>

<221> modified base

<222> (1)..(3)

<223> 2'-OMe-nucleotide

<220>

<221> modified base

<222>(8)..(8)

<223> 2'-OMe-nucleotide

<220>

<221> modified base

<222> (10)..(10)

<223> 2'-OMe-nucleotide

<220>

<221> modified base

<222> (18)..(21)

<223> 2'-OMe-nucleotide

<400> 156

gaagccuuuu gagacccuau u 21

<210> 157

<211> 21

<212> DNA

<213> Artificial sequence

<220>

<223> Artificial sequence description: synthetic

oligonucleotide

<220>

<223> Description of the combined DNA/RNA molecule: synthetic

oligonucleotide

<220>

<221> modified base

<222> (20)..(21)

<223> a, c, t, g, u, unknown or other

<400> 157

uagggucuca aaaggcuucn n 21

<210> 158

<211> 21

<212> RNA

<213> Artificial sequence

<220>

<223> Artificial sequence description: synthetic

oligonucleotide

<220>

<221> modified base

<222> (20)..(21)

<223> 2'-OMe-nucleotide

<400> 158

uagggucuca aaaggcuucu u 21

<210> 159

<211> 21

<212> DNA

<213> Artificial sequence

<220>

<223> Artificial sequence description: synthetic

oligonucleotide

<220>

<223> Description of the combined DNA/RNA molecule: synthetic

oligonucleotide

<220>

<221> modified base

<222> (1)..(1)

<223> 2'-deoxynucleotide

<220>

<221> modified base

<222>(6)..(6)

<223> 2'-deoxynucleotide

<220>

<221> modified base

<222>(8)..(8)

<223> 2'-deoxynucleotide

<220>

<221> modified base

<222> (20)..(21)

<223> 2'-OMe-nucleotide

<400> 159

uagggucuca aaaggcuucu u 21

<210> 160

<211> 21

<212> DNA

<213> Artificial sequence

<220>

<223> Artificial sequence description: synthetic

oligonucleotide

<220>

<223> Description of the combined DNA/RNA molecule: synthetic

oligonucleotide

<220>

<221> modified base

<222>(6)..(6)

<223> 2'-deoxynucleotide

<220>

<221> modified base

<222>(8)..(8)

<223> 2'-deoxynucleotide

<220>

<221> modified base

<222> (20)..(21)

<223> 2'-OMe-nucleotide

<400> 160

uagggucuca aaaggcuucu u 21

<210> 161

<211> 21

<212> DNA

<213> Artificial sequence

<220>

<223> Artificial sequence description: synthetic

oligonucleotide

<220>

<223> Description of the combined DNA/RNA molecule: synthetic

oligonucleotide

<220>

<221> modified base

<222>(6)..(6)

<223> 2'-deoxynucleotide

<220>

<221> modified base

<222>(8)..(8)

<223> 2'-deoxynucleotide

<220>

<221> modified base

<222> (20)..(21)

<223> 2'-OMe-nucleotide

<400> 161

uagggucuca aaaggcuucu u 21

<210> 162

<211> 21

<212> DNA

<213> Artificial sequence

<220>

<223> Artificial sequence description: synthetic

oligonucleotide

<220>

<223> Description of the combined DNA/RNA molecule: synthetic

oligonucleotide

<220>

<221> modified base

<222>(6)..(6)

<223> 2'-deoxynucleotide

<220>

<221> modified base

<222>(8)..(8)

<223> 2'-deoxynucleotide

<220>

<221> modified base

<222> (20)..(21)

<223> 2'-OMe-nucleotide

<400> 162

uagggucuca aaaggcuucu u 21

<210> 163

<211> 21

<212> DNA

<213> Artificial sequence

<220>

<223> Artificial sequence description: synthetic

oligonucleotide

<220>

<223> Description of the combined DNA/RNA molecule: synthetic

oligonucleotide

<220>

<221> modified base

<222>(6)..(6)

<223> 2'-deoxynucleotide

<220>

<221> modified base

<222>(8)..(8)

<223> 2'-deoxynucleotide

<220>

<221> modified base

<222> (20)..(21)

<223> 2'-OMe-nucleotide

<400> 163

uagggucuca aaaggcuucu u 21

<210> 164

<211> 21

<212> DNA

<213> Artificial sequence

<220>

<223> Artificial sequence description: synthetic

oligonucleotide

<220>

<223> Description of the combined DNA/RNA molecule: synthetic

oligonucleotide

<220>

<221> modified base

<222> (1)..(1)

<223> 2'-deoxynucleotide

<220>

<221> modified base

<222> (20)..(21)

<223> 2'-OMe-nucleotide

<400> 164

uagggucuca aaaggcuucu u 21

<210> 165

<211> 21

<212> DNA

<213> Artificial sequence

<220>

<223> Artificial sequence description: synthetic

oligonucleotide

<220>

<223> Description of the combined DNA/RNA molecule: synthetic

oligonucleotide

<220>

<221> modified base

<222> (20)..(21)

<223> 2'-OMe-nucleotide

<400> 165

uagggucuca aaaggcuucu u 21

<210> 166

<211> 21

<212> DNA

<213> Artificial sequence

<220>

<223> Artificial sequence description: synthetic

oligonucleotide

<220>

<223> Description of the combined DNA/RNA molecule: synthetic

oligonucleotide

<220>

<221> modified base

<222>(6)..(6)

<223> 2'-deoxynucleotide

<220>

<221> modified base

<222>(8)..(8)

<223> 2'-deoxynucleotide

<220>

<221> modified base

<222> (20)..(21)

<223> 2'-OMe-nucleotide

<400> 166

uagggucuca aaaggcuucu u 21

<210> 167

<211> 21

<212> DNA

<213> Artificial sequence

<220>

<223> Artificial sequence description: synthetic

oligonucleotide

<220>

<223> Description of the combined DNA/RNA molecule: synthetic

oligonucleotide

<220>

<221> modified base

<222>(6)..(6)

<223> 2'-deoxynucleotide

<220>

<221> modified base

<222>(8)..(8)

<223> 2'-deoxynucleotide

<220>

<221> modified base

<222> (20)..(21)

<223> 2'-OMe-nucleotide

<400> 167

uagggucuca aaaggcuucu u 21

<210> 168

<211> 21

<212> DNA

<213> Artificial sequence

<220>

<223> Artificial sequence description: synthetic

oligonucleotide

<220>

<223> Description of the combined DNA/RNA molecule: synthetic

oligonucleotide

<220>

<221> modified base

<222>(6)..(6)

<223> 2'-deoxynucleotide

<220>

<221> modified base

<222>(8)..(8)

<223> 2'-deoxynucleotide

<220>

<221> modified base

<222> (14)..(14)

<223> 2'-OMe-nucleotide

<220>

<221> modified base

<222> (16)..(16)

<223> 2'-OMe-nucleotide

<220>

<221> modified base

<222> (18)..(18)

<223> 2'-OMe-nucleotide

<220>

<221> modified base

<222> (20)..(21)

<223> 2'-OMe-nucleotide

<400> 168

uagggucuca aaaggcuucu u 21

<210> 169

<211> 21

<212> RNA

<213> Artificial sequence

<220>

<223> Artificial sequence description: synthetic

oligonucleotide

<220>

<221> modified base

<222> (1)..(1)

<223> 2'-OMe-nucleotide

<220>

<221> modified base

<222> (20)..(21)

<223> 2'-OMe-nucleotide

<400> 169

uagggucuca aaaggcuucu u 21

<210> 170

<211> 21

<212> RNA

<213> Artificial sequence

<220>

<223> Artificial sequence description: synthetic

oligonucleotide

<220>

<221> modified base

<222> (1)..(1)

<223> 2'-deoxy-2'-fluoronucleotide

<220>

<221> modified base

<222> (20)..(21)

<223> 2'-OMe-nucleotide

<400> 170

uagggucuca aaaggcuucu u 21

<210> 171

<211> 21

<212> DNA

<213> Artificial sequence

<220>

<223> Artificial sequence description: synthetic

oligonucleotide

<220>

<223> Description of the combined DNA/RNA molecule: synthetic

oligonucleotide

<220>

<221> modified base

<222> (1)..(1)

<223> 2'-deoxynucleotide

<220>

<221> modified base

<222> (20)..(21)

<223> 2'-OMe-nucleotide

<400> 171

uagggucuca aaaggcuucu u 21

<210> 172

<211> 21

<212> RNA

<213> Artificial sequence

<220>

<223> Artificial sequence description: synthetic

oligonucleotide

<220>

<221> modified base

<222> (1)..(2)

<223> Phosphothioate bond

<220>

<221> modified base

<222> (20)..(21)

<223> 2'-OMe-nucleotide

<400> 172

uagggucuca aaaggcuucu u 21

<210> 173

<211> 21

<212> RNA

<213> Artificial sequence

<220>

<223> Artificial sequence description: synthetic

oligonucleotide

<220>

<221> modified base

<222> (1)..(1)

<223> 2'-deoxy-2'-fluoronucleotide

<220>

<221> modified base

<222>(9)..(9)

<223> 2'-deoxy-2'-fluoronucleotide

<220>

<221> modified base

<222> (17)..(17)

<223> 2'-deoxy-2'-fluoronucleotide

<220>

<221> modified base

<222> (20)..(21)

<223> 2'-OMe-nucleotide

<400> 173

uagggucuca aaaggcuucu u 21

<210> 174

<211> 21

<212> RNA

<213> Artificial sequence

<220>

<223> Artificial sequence description: synthetic

oligonucleotide

<220>

<221> modified base

<222> (1)..(1)

<223> 2'-deoxy-2'-fluoronucleotide

<220>

<221> modified base

<222>(9)..(9)

<223> 2'-deoxy-2'-fluoronucleotide

<220>

<221> modified base

<222> (11)..(12)

<223> 2'-deoxy-2'-fluoronucleotide

<220>

<221> modified base

<222> (17)..(17)

<223> 2'-deoxy-2'-fluoronucleotide

<220>

<221> modified base

<222> (20)..(21)

<223> 2'-OMe-nucleotide

<400> 174

uagggucuca aaaggcuucu u 21

<210> 175

<211> 21

<212> RNA

<213> Artificial sequence

<220>

<223> Artificial sequence description: synthetic

oligonucleotide

<220>

<221> modified base

<222> (1)..(1)

<223> 2'-OMe-nucleotide

<220>

<221> modified base

<222>(9)..(9)

<223> 2'-OMe-nucleotide

<220>

<221> modified base

<222> (17)..(17)

<223> 2'-OMe-nucleotide

<220>

<221> modified base

<222> (20)..(21)

<223> 2'-OMe-nucleotide

<400> 175

uagggucuca aaaggcuucu u 21

<210> 176

<211> 21

<212> RNA

<213> Artificial sequence

<220>

<223> Artificial sequence description: synthetic

oligonucleotide

<220>

<221> modified base

<222> (1)..(1)

<223> 2'-deoxy-2'-fluoronucleotide

<220>

<221> modified base

<222>(9)..(9)

<223> 2'-OMe-nucleotide

<220>

<221> modified base

<222> (17)..(17)

<223> 2'-OMe-nucleotide

<220>

<221> modified base

<222> (20)..(21)

<223> 2'-OMe-nucleotide

<400> 176

uagggucuca aaaggcuucu u 21

<210> 177

<211> 21

<212> RNA

<213> Artificial sequence

<220>

<223> Artificial sequence description: synthetic

oligonucleotide

<220>

<221> modified base

<222> (20)..(21)

<223> 2'-OMe-nucleotide

<400> 177

uagggucuca aaaggcuucu u 21

<210> 178

<211> 21

<212> DNA

<213> Artificial sequence

<220>

<223> Artificial sequence description: synthetic

oligonucleotide

<220>

<223> Description of the combined DNA/RNA molecule: synthetic

oligonucleotide

<220>

<221> modified base

<222>(6)..(6)

<223> 2'-deoxynucleotide

<220>

<221> modified base

<222>(8)..(8)

<223> 2'-deoxynucleotide

<220>

<221> modified base

<222> (20)..(21)

<223> 2'-OMe-nucleotide

<400> 178

uagggucuca aaaggcuucu u 21

<210> 179

<211> 21

<212> DNA

<213> Artificial sequence

<220>

<223> Artificial sequence description: synthetic

oligonucleotide

<220>

<223> Description of the combined DNA/RNA molecule: synthetic

oligonucleotide

<220>

<221> modified base

<222> (1)..(1)

<223> 2'-OMe-nucleotide

<220>

<221> modified base

<222>(6)..(6)

<223> 2'-deoxynucleotide

<220>

<221> modified base

<222>(8)..(8)

<223> 2'-deoxynucleotide

<220>

<221> modified base

<222>(9)..(9)

<223> 2'-OMe-nucleotide

<220>

<221> modified base

<222> (11)..(12)

<223> 2'-OMe-nucleotide

<220>

<221> modified base

<222> (18)..(18)

<223> 2'-OMe-nucleotide

<220>

<221> modified base

<222> (20)..(21)

<223> 2'-OMe-nucleotide

<400> 179

uagggucuca aaaggcuucu u 21

<210> 180

<211> 21

<212> DNA

<213> Artificial sequence

<220>

<223> Artificial sequence description: synthetic

oligonucleotide

<220>

<223> Description of the combined DNA/RNA molecule: synthetic

oligonucleotide

<220>

<221> modified base

<222> (1)..(1)

<223> 2'-OMe-nucleotide

<220>

<221> modified base

<222>(6)..(6)

<223> 2'-deoxynucleotide

<220>

<221> modified base

<222>(8)..(8)

<223> 2'-deoxynucleotide

<220>

<221> modified base

<222>(9)..(9)

<223> 2'-OMe-nucleotide

<220>

<221> modified base

<222> (11)..(12)

<223> 2'-OMe-nucleotide

<220>

<221> modified base

<222> (18)..(18)

<223> 2'-OMe-nucleotide

<220>

<221> modified base

<222> (20)..(21)

<223> 2'-OMe-nucleotide

<400> 180

uagggucuca aaaggcuucu u 21

<210> 181

<211> 21

<212> DNA

<213> Artificial sequence

<220>

<223> Artificial sequence description: synthetic

oligonucleotide

<220>

<223> Description of the combined DNA/RNA molecule: synthetic

oligonucleotide

<220>

<221> modified base

<222> (1)..(1)

<223> 2'-OMe-nucleotide

<220>

<221> modified base

<222>(6)..(6)

<223> 2'-deoxynucleotide

<220>

<221> modified base

<222>(8)..(8)

<223> 2'-deoxynucleotide

<220>

<221> modified base

<222>(9)..(9)

<223> 2'-OMe-nucleotide

<220>

<221> modified base

<222> (11)..(12)

<223> 2'-OMe-nucleotide

<220>

<221> modified base

<222> (17)..(18)

<223> 2'-OMe-nucleotide

<220>

<221> modified base

<222> (20)..(21)

<223> 2'-OMe-nucleotide

<400> 181

uagggucuca aaaggcuucu u 21

<210> 182

<211> 21

<212> DNA

<213> Artificial sequence

<220>

<223> Artificial sequence description: synthetic

oligonucleotide

<220>

<223> Description of the combined DNA/RNA molecule: synthetic

oligonucleotide

<220>

<221> modified base

<222> (1)..(1)

<223> 2'-deoxy-2'-fluoronucleotide

<220>

<221> modified base

<222>(6)..(6)

<223> 2'-deoxynucleotide

<220>

<221> modified base

<222>(8)..(8)

<223> 2'-deoxynucleotide

<220>

<221> modified base

<222>(9)..(9)

<223> 2'-OMe-nucleotide

<220>

<221> modified base

<222> (11)..(12)

<223> 2'-OMe-nucleotide

<220>

<221> modified base

<222> (17)..(18)

<223> 2'-OMe-nucleotide

<220>

<221> modified base

<222> (20)..(21)

<223> 2'-OMe-nucleotide

<400> 182

uagggucuca aaaggcuucu u 21

<210> 183

<211> 21

<212> DNA

<213> Artificial sequence

<220>

<223> Artificial sequence description: synthetic

oligonucleotide

<220>

<223> Description of the combined DNA/RNA molecule: synthetic

oligonucleotide

<220>

<221> modified base

<222> (20)..(21)

<223> a, c, t, g, u, unknown or other

<400> 183

ccuuuugaga cccugcugun n 21

<210> 184

<211> 21

<212> RNA

<213> Artificial sequence

<220>

<223> Artificial sequence description: synthetic

oligonucleotide

<220>

<221> modified base

<222> (20)..(21)

<223> 2'-OMe-nucleotide

<400> 184

ccucaucuac accaacuauu u 21

<210> 185

<211> 21

<212> RNA

<213> Artificial sequence

<220>

<223> Artificial sequence description: synthetic

oligonucleotide

<220>

<221> modified base

<222> (20)..(21)

<223> 2'-OMe-nucleotide

<400> 185

ccucaucuac accaacuauu u 21

<210> 186

<211> 21

<212> RNA

<213> Artificial sequence

<220>

<223> Artificial sequence description: synthetic

oligonucleotide

<220>

<221> modified base

<222> (20)..(21)

<223> 2'-OMe-nucleotide

<400> 186

ccucaucuac accaacuauu u 21

<210> 187

<211> 21

<212> RNA

<213> Artificial sequence

<220>

<223> Artificial sequence description: synthetic

oligonucleotide

<220>

<221> modified base

<222> (20)..(21)

<223> 2'-OMe-nucleotide

<400> 187

ccucaucuac accaacuauu u 21

<210> 188

<211> 21

<212> RNA

<213> Artificial sequence

<220>

<223> Artificial sequence description: synthetic

oligonucleotide

<220>

<221> modified base

<222> (20)..(21)

<223> 2'-OMe-nucleotide

<400> 188

ccucaucuac accaacuauu u 21

<210> 189

<211> 21

<212> RNA

<213> Artificial sequence

<220>

<223> Artificial sequence description: synthetic

oligonucleotide

<220>

<221> modified base

<222> (20)..(21)

<223> 2'-OMe-nucleotide

<400> 189

ccucaucuac accaacuauu u 21

<210> 190

<211> 21

<212> RNA

<213> Artificial sequence

<220>

<223> Artificial sequence description: synthetic

oligonucleotide

<220>

<221> modified base

<222> (20)..(21)

<223> 2'-OMe-nucleotide

<400> 190

ccucaucuac accaacuauu u 21

<210> 191

<211> 21

<212> RNA

<213> Artificial sequence

<220>

<223> Artificial sequence description: synthetic

oligonucleotide

<220>

<221> modified base

<222> (20)..(21)

<223> 2'-OMe-nucleotide

<400> 191

ccucaucuac accaacuauu u 21

<210> 192

<211> 21

<212> RNA

<213> Artificial sequence

<220>

<223> Artificial sequence description: synthetic

oligonucleotide

<220>

<221> modified base

<222> (20)..(21)

<223> 2'-OMe-nucleotide

<400> 192

ccucaucuac accaacuauu u 21

<210> 193

<211> 21

<212> RNA

<213> Artificial sequence

<220>

<223> Artificial sequence description: synthetic

oligonucleotide

<220>

<221> modified base

<222> (20)..(21)

<223> 2'-OMe-nucleotide

<400> 193

ccucaucuac accaacuauu u 21

<210> 194

<211> 21

<212> RNA

<213> Artificial sequence

<220>

<223> Artificial sequence description: synthetic

oligonucleotide

<220>

<221> modified base

<222> (1)..(1)

<223> 2'-OMe-nucleotide

<220>

<221> modified base

<222> (3)..(3)

<223> 2'-OMe-nucleotide

<220>

<221> modified base

<222> (5)..(5)

<223> 2'-OMe-nucleotide

<220>

<221> modified base

<222>(7)..(7)

<223> 2'-OMe-nucleotide

<220>

<221> modified base

<222>(9)..(9)

<223> 2'-OMe-nucleotide

<220>

<221> modified base

<222> (20)..(21)

<223> 2'-OMe-nucleotide

<400> 194

ccucaucuac accaacuauu u 21

<210> 195

<211> 21

<212> DNA

<213> Artificial sequence

<220>

<223> Artificial sequence description: synthetic

oligonucleotide

<220>

<223> Description of the combined DNA/RNA molecule: synthetic

oligonucleotide

<220>

<221> modified base

<222> (20)..(21)

<223> a, c, t, g, u, unknown or other

<400> 195

acagcagggu cucaaaaggn n 21

<210> 196

<211> 21

<212> RNA

<213> Artificial sequence

<220>

<223> Artificial sequence description: synthetic

oligonucleotide

<220>

<221> modified base

<222> (20)..(21)

<223> 2'-OMe-nucleotide

<400> 196

auaguuggug uagaugaggu u 21

<210> 197

<211> 21

<212> DNA

<213> Artificial sequence

<220>

<223> Artificial sequence description: synthetic

oligonucleotide

<220>

<223> Description of the combined DNA/RNA molecule: synthetic

oligonucleotide

<220>

<221> modified base

<222> (2)..(2)

<223> 2'-deoxynucleotide

<220>

<221> modified base

<222> (5)..(6)

<223> 2'-deoxynucleotide

<220>

<221> modified base

<222> (20)..(21)

<223> 2'-OMe-nucleotide

<400> 197

auaguuggug uagaugaggu u 21

<210> 198

<211> 21

<212> DNA

<213> Artificial sequence

<220>

<223> Artificial sequence description: synthetic

oligonucleotide

<220>

<223> Description of the combined DNA/RNA molecule: synthetic

oligonucleotide

<220>

<221> modified base

<222> (2)..(2)

<223> 2'-deoxynucleotide

<220>

<221> modified base

<222> (5)..(6)

<223> 2'-deoxynucleotide

<220>

<221> modified base

<222> (20)..(21)

<223> 2'-OMe-nucleotide

<400> 198

auaguuggug uagaugaggu u 21

<210> 199

<211> 21

<212> DNA

<213> Artificial sequence

<220>

<223> Artificial sequence description: synthetic

oligonucleotide

<220>

<223> Description of the combined DNA/RNA molecule: synthetic

oligonucleotide

<220>

<221> modified base

<222> (5)..(6)

<223> 2'-deoxynucleotide

<220>

<221> modified base

<222> (20)..(21)

<223> 2'-OMe-nucleotide

<400> 199

auaguuggug uagaugaggu u 21

<210> 200

<211> 21

<212> DNA

<213> Artificial sequence

<220>

<223> Artificial sequence description: synthetic

oligonucleotide

<220>

<223> Description of the combined DNA/RNA molecule: synthetic

oligonucleotide

<220>

<221> modified base

<222> (5)..(6)

<223> 2'-deoxynucleotide

<220>

<221> modified base

<222> (20)..(21)

<223> 2'-OMe-nucleotide

<400> 200

auaguuggug uagaugaggu u 21

<210> 201

<211> 21

<212> DNA

<213> Artificial sequence

<220>

<223> Artificial sequence description: synthetic

oligonucleotide

<220>

<223> Description of the combined DNA/RNA molecule: synthetic

oligonucleotide

<220>

<221> modified base

<222> (5)..(6)

<223> 2'-deoxynucleotide

<220>

<221> modified base

<222> (20)..(21)

<223> 2'-OMe-nucleotide

<400> 201

auaguuggug uagaugaggu u 21

<210> 202

<211> 21

<212> DNA

<213> Artificial sequence

<220>

<223> Artificial sequence description: synthetic

oligonucleotide

<220>

<223> Description of the combined DNA/RNA molecule: synthetic

oligonucleotide

<220>

<221> modified base

<222> (5)..(5)

<223> 2'-deoxynucleotide

<220>

<221> modified base

<222> (20)..(21)

<223> 2'-OMe-nucleotide

<400> 202

auaguuggug uagaugaggu u 21

<210> 203

<211> 21

<212> DNA

<213> Artificial sequence

<220>

<223> Artificial sequence description: synthetic

oligonucleotide

<220>

<223> Description of the combined DNA/RNA molecule: synthetic

oligonucleotide

<220>

<221> modified base

<222> (5)..(5)

<223> 2'-deoxynucleotide

<220>

<221> modified base

<222> (20)..(21)

<223> 2'-OMe-nucleotide

<400> 203

auaguuggug uagaugaggu u 21

<210> 204

<211> 21

<212> DNA

<213> Artificial sequence

<220>

<223> Artificial sequence description: synthetic

oligonucleotide

<220>

<223> Description of the combined DNA/RNA molecule: synthetic

oligonucleotide

<220>

<221> modified base

<222> (2)..(2)

<223> 2'-deoxynucleotide

<220>

<221> modified base

<222>(6)..(6)

<223> 2'-deoxynucleotide

<220>

<221> modified base

<222> (20)..(21)

<223> 2'-OMe-nucleotide

<400> 204

auaguuggug uagaugaggu u 21

<210> 205

<211> 21

<212> DNA

<213> Artificial sequence

<220>

<223> Artificial sequence description: synthetic

oligonucleotide

<220>

<223> Description of the combined DNA/RNA molecule: synthetic

oligonucleotide

<220>

<221> modified base

<222>(6)..(6)

<223> 2'-deoxynucleotide

<220>

<221> modified base

<222> (20)..(21)

<223> 2'-OMe-nucleotide

<400> 205

auaguuggug uagaugaggu u 21

<210> 206

<211> 21

<212> DNA

<213> Artificial sequence

<220>

<223> Artificial sequence description: synthetic

oligonucleotide

<220>

<223> Description of the combined DNA/RNA molecule: synthetic

oligonucleotide

<220>

<221> modified base

<222> (2)..(2)

<223> 2'-deoxynucleotide

<220>

<221> modified base

<222> (5)..(6)

<223> 2'-deoxynucleotide

<220>

<221> modified base

<222> (14)..(14)

<223> 2'-OMe-nucleotide

<220>

<221> modified base

<222> (16)..(16)

<223> 2'-OMe-nucleotide

<220>

<221> modified base

<222> (18)..(18)

<223> 2'-OMe-nucleotide

<220>

<221> modified base

<222> (20)..(21)

<223> 2'-OMe-nucleotide

<400> 206

auaguuggug uagaugaggu u 21

<210> 207

<211> 21

<212> DNA

<213> Artificial sequence

<220>

<223> Artificial sequence description: synthetic

oligonucleotide

<220>

<223> Description of the combined DNA/RNA molecule: synthetic

oligonucleotide

<220>

<221> modified base

<222> (20)..(21)

<223> a, c, t, g, u, unknown or other

<400> 207

gaugacuaug ugaaggcacn n 21

<210> 208

<211> 21

<212> RNA

<213> Artificial sequence

<220>

<223> Artificial sequence description: synthetic

oligonucleotide

<220>

<221> modified base

<222> (20)..(21)

<223> 2'-OMe-nucleotide

<400> 208

ggaugacuau gugaaggcau u 21

<210> 209

<211> 21

<212> RNA

<213> Artificial sequence

<220>

<223> Artificial sequence description: synthetic

oligonucleotide

<220>

<221> modified base

<222> (20)..(21)

<223> 2'-OMe-nucleotide

<400> 209

ggaugacuau gugaaggcau u 21

<210> 210

<211> 21

<212> RNA

<213> Artificial sequence

<220>

<223> Artificial sequence description: synthetic

oligonucleotide

<220>

<221> modified base

<222> (20)..(21)

<223> 2'-OMe-nucleotide

<400> 210

ggaugacuau gugaaggcau u 21

<210> 211

<211> 21

<212> RNA

<213> Artificial sequence

<220>

<223> Artificial sequence description: synthetic

oligonucleotide

<220>

<221> modified base

<222> (20)..(21)

<223> 2'-OMe-nucleotide

<400> 211

ggaugacuau gugaaggcau u 21

<210> 212

<211> 21

<212> RNA

<213> Artificial sequence

<220>

<223> Artificial sequence description: synthetic

oligonucleotide

<220>

<221> modified base

<222> (20)..(21)

<223> 2'-OMe-nucleotide

<400> 212

ggaugacuau gugaaggcau u 21

<210> 213

<211> 21

<212> RNA

<213> Artificial sequence

<220>

<223> Artificial sequence description: synthetic

oligonucleotide

<220>

<221> modified base

<222> (20)..(21)

<223> 2'-OMe-nucleotide

<400> 213

ggaugacuau gugaaggcau u 21

<210> 214

<211> 21

<212> RNA

<213> Artificial sequence

<220>

<223> Artificial sequence description: synthetic

oligonucleotide

<220>

<221> modified base

<222> (20)..(21)

<223> 2'-OMe-nucleotide

<400> 214

ggaugacuau gugaaggcau u 21

<210> 215

<211> 21

<212> RNA

<213> Artificial sequence

<220>

<223> Artificial sequence description: synthetic

oligonucleotide

<220>

<221> modified base

<222> (20)..(21)

<223> 2'-OMe-nucleotide

<400> 215

ggaugacuau gugaaggcau u 21

<210> 216

<211> 21

<212> RNA

<213> Artificial sequence

<220>

<223> Artificial sequence description: synthetic

oligonucleotide

<220>

<221> modified base

<222> (20)..(21)

<223> 2'-OMe-nucleotide

<400> 216

ggaugacuau gugaaggcau u 21

<210> 217

<211> 21

<212> RNA

<213> Artificial sequence

<220>

<223> Artificial sequence description: synthetic

oligonucleotide

<220>

<221> modified base

<222> (20)..(21)

<223> 2'-OMe-nucleotide

<400> 217

ggaugacuau gugaaggcau u 21

<210> 218

<211> 21

<212> RNA

<213> Artificial sequence

<220>

<223> Artificial sequence description: synthetic

oligonucleotide

<220>

<221> modified base

<222> (1)..(1)

<223> 2'-OMe-nucleotide

<220>

<221> modified base

<222> (3)..(3)

<223> 2'-OMe-nucleotide

<220>

<221> modified base

<222> (5)..(5)

<223> 2'-OMe-nucleotide

<220>

<221> modified base

<222>(7)..(7)

<223> 2'-OMe-nucleotide

<220>

<221> modified base

<222> (20)..(21)

<223> 2'-OMe-nucleotide

<400> 218

ggaugacuau gugaaggcau u 21

<210> 219

<211> 21

<212> RNA

<213> Artificial sequence

<220>

<223> Artificial sequence description: synthetic

oligonucleotide

<220>

<221> modified base

<222> (11)..(11)

<223> 2'-deoxy-2'-fluoronucleotide

<220>

<221> modified base

<222> (13)..(13)

<223> 2'-deoxy-2'-fluoronucleotide

<220>

<221> modified base

<222> (20)..(21)

<223> 2'-OMe-nucleotide

<400> 219

ggaugacuau gugaaggcau u 21

<210> 220

<211> 21

<212> RNA

<213> Artificial sequence

<220>

<223> Artificial sequence description: synthetic

oligonucleotide

<220>

<221> modified base

<222> (1)..(2)

<223> 2'-OMe-nucleotide

<220>

<221> modified base

<222> (20)..(21)

<223> 2'-OMe-nucleotide

<400> 220

ggaugacuau gugaaggcau u 21

<210> 221

<211> 21

<212> RNA

<213> Artificial sequence

<220>

<223> Artificial sequence description: synthetic

oligonucleotide

<220>

<221> modified base

<222> (1)..(2)

<223> 2'-OMe-nucleotide

<220>

<221> modified base

<222> (20)..(21)

<223> 2'-OMe-nucleotide

<400> 221

ggaugacuau gugaaggcau u 21

<210> 222

<211> 21

<212> DNA

<213> Artificial sequence

<220>

<223> Artificial sequence description: synthetic

oligonucleotide

<220>

<223> Description of the combined DNA/RNA molecule: synthetic

oligonucleotide

<220>

<221> modified base

<222> (20)..(21)

<223> a, c, t, g, u, unknown or other

<400> 222

gugccuucac auagucaucn n 21

<210> 223

<211> 21

<212> RNA

<213> Artificial sequence

<220>

<223> Artificial sequence description: synthetic

oligonucleotide

<220>

<221> modified base

<222> (20)..(21)

<223> 2'-OMe-nucleotide

<400> 223

ugccuucaca uagucauccu u 21

<210> 224

<211> 21

<212> DNA

<213> Artificial sequence

<220>

<223> Artificial sequence description: synthetic

oligonucleotide

<220>

<223> Description of the combined DNA/RNA molecule: synthetic

oligonucleotide

<220>

<221> modified base

<222> (1)..(1)

<223> 2'-deoxynucleotide

<220>

<221> modified base

<222> (5)..(6)

<223> 2'-deoxynucleotide

<220>

<221> modified base

<222> (20)..(21)

<223> 2'-OMe-nucleotide

<400> 224

ugccuucaca uagucauccu u 21

<210> 225

<211> 21

<212> DNA

<213> Artificial sequence

<220>

<223> Artificial sequence description: synthetic

oligonucleotide

<220>

<223> Description of the combined DNA/RNA molecule: synthetic

oligonucleotide

<220>

<221> modified base

<222> (5)..(6)

<223> 2'-deoxynucleotide

<220>

<221> modified base

<222> (20)..(21)

<223> 2'-OMe-nucleotide

<400> 225

ugccuucaca uagucauccu u 21

<210> 226

<211> 21

<212> DNA

<213> Artificial sequence

<220>

<223> Artificial sequence description: synthetic

oligonucleotide

<220>

<223> Description of the combined DNA/RNA molecule: synthetic

oligonucleotide

<220>

<221> modified base

<222> (5)..(6)

<223> 2'-deoxynucleotide

<220>

<221> modified base

<222> (20)..(21)

<223> 2'-OMe-nucleotide

<400> 226

ugccuucaca uagucauccu u 21

<210> 227

<211> 21

<212> DNA

<213> Artificial sequence

<220>

<223> Artificial sequence description: synthetic

oligonucleotide

<220>

<223> Description of the combined DNA/RNA molecule: synthetic

oligonucleotide

<220>

<221> modified base

<222> (5)..(6)

<223> 2'-deoxynucleotide

<220>

<221> modified base

<222> (20)..(21)

<223> 2'-OMe-nucleotide

<400> 227

ugccuucaca uagucauccu u 21

<210> 228

<211> 21

<212> DNA

<213> Artificial sequence

<220>

<223> Artificial sequence description: synthetic

oligonucleotide

<220>

<223> Description of the combined DNA/RNA molecule: synthetic

oligonucleotide

<220>

<221> modified base

<222> (5)..(6)

<223> 2'-deoxynucleotide

<220>

<221> modified base

<222> (20)..(21)

<223> 2'-OMe-nucleotide

<400> 228

ugccuucaca uagucauccu u 21

<210> 229

<211> 21

<212> DNA

<213> Artificial sequence

<220>

<223> Artificial sequence description: synthetic

oligonucleotide

<220>

<223> Description of the combined DNA/RNA molecule: synthetic

oligonucleotide

<220>

<221> modified base

<222> (1)..(1)

<223> 2'-deoxynucleotide

<220>

<221> modified base

<222> (5)..(5)

<223> 2'-deoxynucleotide

<220>

<221> modified base

<222> (20)..(21)

<223> 2'-OMe-nucleotide

<400> 229

ugccuucaca uagucauccu u 21

<210> 230

<211> 21

<212> DNA

<213> Artificial sequence

<220>

<223> Artificial sequence description: synthetic

oligonucleotide

<220>

<223> Description of the combined DNA/RNA molecule: synthetic

oligonucleotide

<220>

<221> modified base

<222> (5)..(5)

<223> 2'-deoxynucleotide

<220>

<221> modified base

<222> (20)..(21)

<223> 2'-OMe-nucleotide

<400> 230

ugccuucaca uagucauccu u 21

<210> 231

<211> 21

<212> DNA

<213> Artificial sequence

<220>

<223> Artificial sequence description: synthetic

oligonucleotide

<220>

<223> Description of the combined DNA/RNA molecule: synthetic

oligonucleotide

<220>

<221> modified base

<222>(6)..(6)

<223> 2'-deoxynucleotide

<220>

<221> modified base

<222> (20)..(21)

<223> 2'-OMe-nucleotide

<400> 231

ugccuucaca uagucauccu u 21

<210> 232

<211> 21

<212> DNA

<213> Artificial sequence

<220>

<223> Artificial sequence description: synthetic

oligonucleotide

<220>

<223> Description of the combined DNA/RNA molecule: synthetic

oligonucleotide

<220>

<221> modified base

<222>(6)..(6)

<223> 2'-deoxynucleotide

<220>

<221> modified base

<222> (20)..(21)

<223> 2'-OMe-nucleotide

<400> 232

ugccuucaca uagucauccu u 21

<210> 233

<211> 21

<212> DNA

<213> Artificial sequence

<220>

<223> Artificial sequence description: synthetic

oligonucleotide

<220>

<223> Description of the combined DNA/RNA molecule: synthetic

oligonucleotide

<220>

<221> modified base

<222> (5)..(6)

<223> 2'-deoxynucleotide

<220>

<221> modified base

<222> (14)..(14)

<223> 2'-OMe-nucleotide

<220>

<221> modified base

<222> (16)..(16)

<223> 2'-OMe-nucleotide

<220>

<221> modified base

<222> (18)..(18)

<223> 2'-OMe-nucleotide

<220>

<221> modified base

<222> (20)..(21)

<223> 2'-OMe-nucleotide

<400> 233

ugccuucaca uagucauccu u 21

<210> 234

<211> 21

<212> RNA

<213> Artificial sequence

<220>

<223> Artificial sequence description: synthetic

oligonucleotide

<220>

<221> modified base

<222> (4)..(4)

<223> 2'-deoxy-2'-fluoronucleotide

<220>

<221> modified base

<222> (20)..(21)

<223> 2'-OMe-nucleotide

<400> 234

ugccuucaca uagucauccu u 21

<210> 235

<211> 21

<212> RNA

<213> Artificial sequence

<220>

<223> Artificial sequence description: synthetic

oligonucleotide

<220>

<221> modified base

<222>(9)..(9)

<223> 2'-OMe-nucleotide

<220>

<221> modified base

<222> (11)..(11)

<223> 2'-OMe-nucleotide

<220>

<221> modified base

<222> (13)..(13)

<223> 2'-OMe-nucleotide

<220>

<221> modified base

<222> (15)..(15)

<223> 2'-OMe-nucleotide

<220>

<221> modified base

<222> (17)..(17)

<223> 2'-OMe-nucleotide

<220>

<221> modified base

<222> (19)..(21)

<223> 2'-OMe-nucleotide

<400> 235

ugccuucaca uagucauccu u 21

<210> 236

<211> 21

<212> RNA

<213> Artificial sequence

<220>

<223> Artificial sequence description: synthetic

oligonucleotide

<220>

<221> modified base

<222> (11)..(11)

<223> 2'-OMe-nucleotide

<220>

<221> modified base

<222> (13)..(13)

<223> 2'-OMe-nucleotide

<220>

<221> modified base

<222> (15)..(15)

<223> 2'-OMe-nucleotide

<220>

<221> modified base

<222> (17)..(17)

<223> 2'-OMe-nucleotide

<220>

<221> modified base

<222> (19)..(21)

<223> 2'-OMe-nucleotide

<400> 236

ugccuucaca uagucauccu u 21

<210> 237

<211> 21

<212> DNA

<213> Artificial sequence

<220>

<223> Artificial sequence description: synthetic

oligonucleotide

<220>

<223> Description of the combined DNA/RNA molecule: synthetic

oligonucleotide

<220>

<221> modified base

<222> (20)..(21)

<223> a, c, t, g, u, unknown or other

<400> 237

gaagccuuuu gagacccugn n 21

<210> 238

<211> 21

<212> RNA

<213> Artificial sequence

<220>

<223> Artificial sequence description: synthetic

oligonucleotide

<220>

<221> modified base

<222> (20)..(21)

<223> 2'-OMe-nucleotide

<400> 238

gaagccuuuu gagacccugu u 21

<210> 239

<211> 21

<212> RNA

<213> Artificial sequence

<220>

<223> Artificial sequence description: synthetic

oligonucleotide

<220>

<221> modified base

<222> (20)..(21)

<223> 2'-OMe-nucleotide

<400> 239

gaagccuuuu gagacccugu u 21

<210> 240

<211> 21

<212> RNA

<213> Artificial sequence

<220>

<223> Artificial sequence description: synthetic

oligonucleotide

<220>

<221> modified base

<222> (20)..(21)

<223> 2'-OMe-nucleotide

<400> 240

gaagccuuuu gagacccugu u 21

<210> 241

<211> 21

<212> RNA

<213> Artificial sequence

<220>

<223> Artificial sequence description: synthetic

oligonucleotide

<220>

<221> modified base

<222> (20)..(21)

<223> 2'-OMe-nucleotide

<400> 241

gaagccuuuu gagacccugu u 21

<210> 242

<211> 21

<212> RNA

<213> Artificial sequence

<220>

<223> Artificial sequence description: synthetic

oligonucleotide

<220>

<221> modified base

<222> (20)..(21)

<223> 2'-OMe-nucleotide

<400> 242

gaagccuuuu gagacccugu u 21

<210> 243

<211> 21

<212> RNA

<213> Artificial sequence

<220>

<223> Artificial sequence description: synthetic

oligonucleotide

<220>

<221> modified base

<222> (20)..(21)

<223> 2'-OMe-nucleotide

<400> 243

gaagccuuuu gagacccugu u 21

<210> 244

<211> 21

<212> RNA

<213> Artificial sequence

<220>

<223> Artificial sequence description: synthetic

oligonucleotide

<220>

<221> modified base

<222> (20)..(21)

<223> 2'-OMe-nucleotide

<400> 244

gaagccuuuu gagacccugu u 21

<210> 245

<211> 21

<212> RNA

<213> Artificial sequence

<220>

<223> Artificial sequence description: synthetic

oligonucleotide

<220>

<221> modified base

<222> (20)..(21)

<223> 2'-OMe-nucleotide

<400> 245

gaagccuuuu gagacccugu u 21

<210> 246

<211> 21

<212> RNA

<213> Artificial sequence

<220>

<223> Artificial sequence description: synthetic

oligonucleotide

<220>

<221> modified base

<222> (20)..(21)

<223> 2'-OMe-nucleotide

<400> 246

gaagccuuuu gagacccugu u 21

<210> 247

<211> 21

<212> RNA

<213> Artificial sequence

<220>

<223> Artificial sequence description: synthetic

oligonucleotide

<220>

<221> modified base

<222> (20)..(21)

<223> 2'-OMe-nucleotide

<400> 247

gaagccuuuu gagacccugu u 21

<210> 248

<211> 21

<212> RNA

<213> Artificial sequence

<220>

<223> Artificial sequence description: synthetic

oligonucleotide

<220>

<221> modified base

<222> (1)..(1)

<223> 2'-OMe-nucleotide

<220>

<221> modified base

<222> (3)..(3)

<223> 2'-OMe-nucleotide

<220>

<221> modified base

<222> (5)..(5)

<223> 2'-OMe-nucleotide

<220>

<221> modified base

<222>(7)..(7)

<223> 2'-OMe-nucleotide

<220>

<221> modified base

<222>(9)..(9)

<223> 2'-OMe-nucleotide

<220>

<221> modified base

<222> (20)..(21)

<223> 2'-OMe-nucleotide

<400> 248

gaagccuuuu gagacccugu u 21

<210> 249

<211> 21

<212> DNA

<213> Artificial sequence

<220>

<223> Artificial sequence description: synthetic

oligonucleotide

<220>

<223> Description of the combined DNA/RNA molecule: synthetic

oligonucleotide

<220>

<221> modified base

<222> (20)..(21)

<223> a, c, t, g, u, unknown or other

<400> 249

cagggucuca aaaggcuucn n 21

<210> 250

<211> 21

<212> RNA

<213> Artificial sequence

<220>

<223> Artificial sequence description: synthetic

oligonucleotide

<220>

<221> modified base

<222> (20)..(21)

<223> 2'-OMe-nucleotide

<400> 250

cagggucuca aaaggcuucu u 21

<210> 251

<211> 21

<212> DNA

<213> Artificial sequence

<220>

<223> Artificial sequence description: synthetic

oligonucleotide

<220>

<223> Description of the combined DNA/RNA molecule: synthetic

oligonucleotide

<220>

<221> modified base

<222>(6)..(6)

<223> 2'-deoxynucleotide

<220>

<221> modified base

<222>(8)..(8)

<223> 2'-deoxynucleotide

<220>

<221> modified base

<222> (20)..(21)

<223> 2'-OMe-nucleotide

<400> 251

cagggucuca aaaggcuucu u 21

<210> 252

<211> 21

<212> DNA

<213> Artificial sequence

<220>

<223> Artificial sequence description: synthetic

oligonucleotide

<220>

<223> Description of the combined DNA/RNA molecule: synthetic

oligonucleotide

<220>

<221> modified base

<222>(6)..(6)

<223> 2'-deoxynucleotide

<220>

<221> modified base

<222>(8)..(8)

<223> 2'-deoxynucleotide

<220>

<221> modified base

<222> (20)..(21)

<223> 2'-OMe-nucleotide

<400> 252

cagggucuca aaaggcuucu u 21

<210> 253

<211> 21

<212> DNA

<213> Artificial sequence

<220>

<223> Artificial sequence description: synthetic

oligonucleotide

<220>

<223> Description of the combined DNA/RNA molecule: synthetic

oligonucleotide

<220>

<221> modified base

<222>(6)..(6)

<223> 2'-deoxynucleotide

<220>

<221> modified base

<222>(8)..(8)

<223> 2'-deoxynucleotide

<220>

<221> modified base

<222> (20)..(21)

<223> 2'-OMe-nucleotide

<400> 253

cagggucuca aaaggcuucu u 21

<210> 254

<211> 21

<212> DNA

<213> Artificial sequence

<220>

<223> Artificial sequence description: synthetic

oligonucleotide

<220>

<223> Description of the combined DNA/RNA molecule: synthetic

oligonucleotide

<220>

<221> modified base

<222>(6)..(6)

<223> 2'-deoxynucleotide

<220>

<221> modified base

<222>(8)..(8)

<223> 2'-deoxynucleotide

<220>

<221> modified base

<222> (20)..(21)

<223> 2'-OMe-nucleotide

<400> 254

cagggucuca aaaggcuucu u 21

<210> 255

<211> 21

<212> DNA

<213> Artificial sequence

<220>

<223> Artificial sequence description: synthetic

oligonucleotide

<220>

<223> Description of the combined DNA/RNA molecule: synthetic

oligonucleotide

<220>

<221> modified base

<222>(6)..(6)

<223> 2'-deoxynucleotide

<220>

<221> modified base

<222>(8)..(8)

<223> 2'-deoxynucleotide

<220>

<221> modified base

<222> (20)..(21)

<223> 2'-OMe-nucleotide

<400> 255

cagggucuca aaaggcuucu u 21

<210> 256

<211> 21

<212> DNA

<213> Artificial sequence

<220>

<223> Artificial sequence description: synthetic

oligonucleotide

<220>

<223> Description of the combined DNA/RNA molecule: synthetic

oligonucleotide

<220>

<221> modified base

<222> (20)..(21)

<223> 2'-OMe-nucleotide

<400> 256

cagggucuca aaaggcuucu u 21

<210> 257

<211> 21

<212> DNA

<213> Artificial sequence

<220>

<223> Artificial sequence description: synthetic

oligonucleotide

<220>

<223> Description of the combined DNA/RNA molecule: synthetic

oligonucleotide

<400> 257

cagggucuca aaaggcuucu u 21

<210> 258

<211> 21

<212> DNA

<213> Artificial sequence

<220>

<223> Artificial sequence description: synthetic

oligonucleotide

<220>

<223> Description of the combined DNA/RNA molecule: synthetic

oligonucleotide

<220>

<221> modified base

<222>(6)..(6)

<223> 2'-deoxynucleotide

<220>

<221> modified base

<222>(8)..(8)

<223> 2'-deoxynucleotide

<220>

<221> modified base

<222> (20)..(21)

<223> 2'-OMe-nucleotide

<400> 258

cagggucuca aaaggcuucu u 21

<210> 259

<211> 21

<212> DNA

<213> Artificial sequence

<220>

<223> Artificial sequence description: synthetic

oligonucleotide

<220>

<223> Description of the combined DNA/RNA molecule: synthetic

oligonucleotide

<220>

<221> modified base

<222>(6)..(6)

<223> 2'-deoxynucleotide

<220>

<221> modified base

<222>(8)..(8)

<223> 2'-deoxynucleotide

<220>

<221> modified base

<222> (20)..(21)

<223> 2'-OMe-nucleotide

<400> 259

cagggucuca aaaggcuucu u 21

<210> 260

<211> 21

<212> DNA

<213> Artificial sequence

<220>

<223> Artificial sequence description: synthetic

oligonucleotide

<220>

<223> Description of the combined DNA/RNA molecule: synthetic

oligonucleotide

<220>

<221> modified base

<222>(6)..(6)

<223> 2'-deoxynucleotide

<220>

<221> modified base

<222>(8)..(8)

<223> 2'-deoxynucleotide

<220>

<221> modified base

<222> (14)..(14)

<223> 2'-OMe-nucleotide

<220>

<221> modified base

<222> (16)..(16)

<223> 2'-OMe-nucleotide

<220>

<221> modified base

<222> (18)..(18)

<223> 2'-OMe-nucleotide

<220>

<221> modified base

<222> (20)..(21)

<223> 2'-OMe-nucleotide

<400> 260

cagggucuca aaaggcuucu u 21

<210> 261

<211> 21

<212> DNA

<213> Artificial sequence

<220>

<223> Artificial sequence description: synthetic

oligonucleotide

<220>

<223> Description of the combined DNA/RNA molecule: synthetic

oligonucleotide

<220>

<221> modified base

<222> (20)..(21)

<223> a, c, t, g, u, unknown or other

<400> 261

ccucaucuac accaacuaun n 21

<210> 262

<211> 21

<212> RNA

<213> Artificial sequence

<220>

<223> Artificial sequence description: synthetic

oligonucleotide

<220>

<221> modified base

<222> (20)..(21)

<223> 2'-OMe-nucleotide

<400> 262

ccucaucuac accaacuauu u 21

<210> 263

<211> 21

<212> RNA

<213> Artificial sequence

<220>

<223> Artificial sequence description: synthetic

oligonucleotide

<220>

<221> modified base

<222> (20)..(21)

<223> 2'-OMe-nucleotide

<400> 263

ccucaucuac accaacuauu u 21

<210> 264

<211> 21

<212> RNA

<213> Artificial sequence

<220>

<223> Artificial sequence description: synthetic

oligonucleotide

<220>

<221> modified base

<222> (20)..(21)

<223> 2'-OMe-nucleotide

<400> 264

ccucaucuac accaacuauu u 21

<210> 265

<211> 21

<212> RNA

<213> Artificial sequence

<220>

<223> Artificial sequence description: synthetic

oligonucleotide

<220>

<221> modified base

<222> (20)..(21)

<223> 2'-OMe-nucleotide

<400> 265

ccucaucuac accaacuauu u 21

<210> 266

<211> 21

<212> RNA

<213> Artificial sequence

<220>

<223> Artificial sequence description: synthetic

oligonucleotide

<220>

<221> modified base

<222> (20)..(21)

<223> 2'-OMe-nucleotide

<400> 266

ccucaucuac accaacuauu u 21

<210> 267

<211> 21

<212> RNA

<213> Artificial sequence

<220>

<223> Artificial sequence description: synthetic

oligonucleotide

<220>

<221> modified base

<222> (20)..(21)

<223> 2'-OMe-nucleotide

<400> 267

ccucaucuac accaacuauu u 21

<210> 268

<211> 21

<212> RNA

<213> Artificial sequence

<220>

<223> Artificial sequence description: synthetic

oligonucleotide

<220>

<221> modified base

<222> (20)..(21)

<223> 2'-OMe-nucleotide

<400> 268

ccucaucuac accaacuauu u 21

<210> 269

<211> 21

<212> RNA

<213> Artificial sequence

<220>

<223> Artificial sequence description: synthetic

oligonucleotide

<220>

<221> modified base

<222> (20)..(21)

<223> 2'-OMe-nucleotide

<400> 269

ccucaucuac accaacuauu u 21

<210> 270

<211> 21

<212> RNA

<213> Artificial sequence

<220>

<223> Artificial sequence description: synthetic

oligonucleotide

<220>

<221> modified base

<222> (20)..(21)

<223> 2'-OMe-nucleotide

<400> 270

ccucaucuac accaacuauu u 21

<210> 271

<211> 21

<212> RNA

<213> Artificial sequence

<220>

<223> Artificial sequence description: synthetic

oligonucleotide

<220>

<221> modified base

<222> (20)..(21)

<223> 2'-OMe-nucleotide

<400> 271

ccucaucuac accaacuauu u 21

<210> 272

<211> 21

<212> RNA

<213> Artificial sequence

<220>

<223> Artificial sequence description: synthetic

oligonucleotide

<220>

<221> modified base

<222> (1)..(1)

<223> 2'-OMe-nucleotide

<220>

<221> modified base

<222> (3)..(3)

<223> 2'-OMe-nucleotide

<220>

<221> modified base

<222> (5)..(5)

<223> 2'-OMe-nucleotide

<220>

<221> modified base

<222>(7)..(7)

<223> 2'-OMe-nucleotide

<220>

<221> modified base

<222>(9)..(9)

<223> 2'-OMe-nucleotide

<220>

<221> modified base

<222> (20)..(21)

<223> 2'-OMe-nucleotide

<400> 272

ccucaucuac accaacuauu u 21

<210> 273

<211> 21

<212> DNA

<213> Artificial sequence

<220>

<223> Artificial sequence description: synthetic

oligonucleotide

<220>

<223> Description of the combined DNA/RNA molecule: synthetic

oligonucleotide

<220>

<221> modified base

<222> (20)..(21)

<223> a, c, t, g, u, unknown or other

<400> 273

auaguuggug uagaugaggn n 21

<210> 274

<211> 21

<212> RNA

<213> Artificial sequence

<220>

<223> Artificial sequence description: synthetic

oligonucleotide

<220>

<221> modified base

<222> (20)..(21)

<223> 2'-OMe-nucleotide

<400> 274

auaguuggug uagaugaggu u 21

<210> 275

<211> 21

<212> DNA

<213> Artificial sequence

<220>

<223> Artificial sequence description: synthetic

oligonucleotide

<220>

<223> Description of the combined DNA/RNA molecule: synthetic

oligonucleotide

<220>

<221> modified base

<222> (2)..(2)

<223> 2'-deoxynucleotide

<220>

<221> modified base

<222> (5)..(6)

<223> 2'-deoxynucleotide

<220>

<221> modified base

<222> (20)..(21)

<223> 2'-OMe-nucleotide

<400> 275

auaguuggug uagaugaggu u 21

<210> 276

<211> 21

<212> DNA

<213> Artificial sequence

<220>

<223> Artificial sequence description: synthetic

oligonucleotide

<220>

<223> Description of the combined DNA/RNA molecule: synthetic

oligonucleotide

<220>

<221> modified base

<222> (2)..(2)

<223> 2'-deoxynucleotide

<220>

<221> modified base

<222> (5)..(6)

<223> 2'-deoxynucleotide

<220>

<221> modified base

<222> (20)..(21)

<223> 2'-OMe-nucleotide

<400> 276

auaguuggug uagaugaggu u 21

<210> 277

<211> 21

<212> DNA

<213> Artificial sequence

<220>

<223> Artificial sequence description: synthetic

oligonucleotide

<220>

<223> Description of the combined DNA/RNA molecule: synthetic

oligonucleotide

<220>

<221> modified base

<222> (5)..(6)

<223> 2'-deoxynucleotide

<220>

<221> modified base

<222> (20)..(21)

<223> 2'-OMe-nucleotide

<400> 277

auaguuggug uagaugaggu u 21

<210> 278

<211> 21

<212> DNA

<213> Artificial sequence

<220>

<223> Artificial sequence description: synthetic

oligonucleotide

<220>

<223> Description of the combined DNA/RNA molecule: synthetic

oligonucleotide

<220>

<221> modified base

<222> (5)..(6)

<223> 2'-deoxynucleotide

<220>

<221> modified base

<222> (20)..(21)

<223> 2'-OMe-nucleotide

<400> 278

auaguuggug uagaugaggu u 21

<210> 279

<211> 21

<212> DNA

<213> Artificial sequence

<220>

<223> Artificial sequence description: synthetic

oligonucleotide

<220>

<223> Description of the combined DNA/RNA molecule: synthetic

oligonucleotide

<220>

<221> modified base

<222> (5)..(6)

<223> 2'-deoxynucleotide

<220>

<221> modified base

<222> (20)..(21)

<223> 2'-OMe-nucleotide

<400> 279

auaguuggug uagaugaggu u 21

<210> 280

<211> 21

<212> DNA

<213> Artificial sequence

<220>

<223> Artificial sequence description: synthetic

oligonucleotide

<220>

<223> Description of the combined DNA/RNA molecule: synthetic

oligonucleotide

<220>

<221> modified base

<222> (5)..(5)

<223> 2'-deoxynucleotide

<220>

<221> modified base

<222> (20)..(21)

<223> 2'-OMe-nucleotide

<400> 280

auaguuggug uagaugaggu u 21

<210> 281

<211> 21

<212> DNA

<213> Artificial sequence

<220>

<223> Artificial sequence description: synthetic

oligonucleotide

<220>

<223> Description of the combined DNA/RNA molecule: synthetic

oligonucleotide

<220>

<221> modified base

<222> (5)..(5)

<223> 2'-deoxynucleotide

<220>

<221> modified base

<222> (20)..(21)

<223> 2'-OMe-nucleotide

<400> 281

auaguuggug uagaugaggu u 21

<210> 282

<211> 21

<212> DNA

<213> Artificial sequence

<220>

<223> Artificial sequence description: synthetic

oligonucleotide

<220>

<223> Description of the combined DNA/RNA molecule: synthetic

oligonucleotide

<220>

<221> modified base

<222> (2)..(2)

<223> 2'-deoxynucleotide

<220>

<221> modified base

<222>(6)..(6)

<223> 2'-deoxynucleotide

<220>

<221> modified base

<222> (20)..(21)

<223> 2'-OMe-nucleotide

<400> 282

auaguuggug uagaugaggu u 21

<210> 283

<211> 21

<212> DNA

<213> Artificial sequence

<220>

<223> Artificial sequence description: synthetic

oligonucleotide

<220>

<223> Description of the combined DNA/RNA molecule: synthetic

oligonucleotide

<220>

<221> modified base

<222>(6)..(6)

<223> 2'-deoxynucleotide

<220>

<221> modified base

<222> (20)..(21)

<223> 2'-OMe-nucleotide

<400> 283

auaguuggug uagaugaggu u 21

<210> 284

<211> 21

<212> DNA

<213> Artificial sequence

<220>

<223> Artificial sequence description: synthetic

oligonucleotide

<220>

<223> Description of the combined DNA/RNA molecule: synthetic

oligonucleotide

<220>

<221> modified base

<222> (2)..(2)

<223> 2'-deoxynucleotide

<220>

<221> modified base

<222> (5)..(6)

<223> 2'-deoxynucleotide

<220>

<221> modified base

<222> (14)..(14)

<223> 2'-OMe-nucleotide

<220>

<221> modified base

<222> (16)..(16)

<223> 2'-OMe-nucleotide

<220>

<221> modified base

<222> (18)..(18)

<223> 2'-OMe-nucleotide

<220>

<221> modified base

<222> (20)..(21)

<223> 2'-OMe-nucleotide

<400> 284

auaguuggug uagaugaggu u 21

<210> 285

<211> 21

<212> RNA

<213> Artificial sequence

<220>

<223> Artificial sequence description: synthetic

oligonucleotide

<220>

<221> modified base

<222> (20)..(21)

<223> 2'-methoxynucleotide

<400> 285

cuuagugacu uuacuuguau u 21

<210> 286

<211> 21

<212> RNA

<213> Artificial sequence

<220>

<223> Artificial sequence description: synthetic

oligonucleotide

<220>

<221> modified base

<222> (20)..(21)

<223> 2'-methoxynucleotide

<400> 286

cagaccagca ugacagauuu u 21

<210> 287

<211> 21

<212> RNA

<213> Artificial sequence

<220>

<223> Artificial sequence description: synthetic

oligonucleotide

<220>

<221> modified base

<222> (20)..(21)

<223> 2'-methoxynucleotide

<400> 287

ugaucuucuc caagaggaau u 21

<210> 288

<211> 21

<212> RNA

<213> Artificial sequence

<220>

<223> Artificial sequence description: synthetic

oligonucleotide

<220>

<221> modified base

<222> (20)..(21)

<223> 2'-methoxynucleotide

<400> 288

guucauugca cuuugauuau u 21

<210> 289

<211> 21

<212> RNA

<213> Artificial sequence

<220>

<223> Artificial sequence description: synthetic

oligonucleotide

<220>

<221> modified base

<222> (20)..(21)

<223> 2'-methoxynucleotide

<400> 289

cauugcacuu ugauuagcau u 21

<210> 290

<211> 21

<212> RNA

<213> Artificial sequence

<220>

<223> Artificial sequence description: synthetic

oligonucleotide

<220>

<221> modified base

<222> (20)..(21)

<223> 2'-methoxynucleotide

<400> 290

agcgauggaa cuucgacuuu u 21

<210> 291

<211> 21

<212> RNA

<213> Artificial sequence

<220>

<223> Artificial sequence description: synthetic

oligonucleotide

<220>

<221> modified base

<222> (20)..(21)

<223> 2'-methoxynucleotide

<400> 291

gcgauggaac uucgacuuuu u 21

<210> 292

<211> 21

<212> RNA

<213> Artificial sequence

<220>

<223> Artificial sequence description: synthetic

oligonucleotide

<220>

<221> modified base

<222> (20)..(21)

<223> 2'-methoxynucleotide

<400> 292

gggaagggac acacaagaau u 21

<210> 293

<211> 21

<212> RNA

<213> Artificial sequence

<220>

<223> Artificial sequence description: synthetic

oligonucleotide

<220>

<221> modified base

<222> (20)..(21)

<223> 2'-methoxynucleotide

<400> 293

ucuaccucag gcagcucaau u 21

<210> 294

<211> 21

<212> RNA

<213> Artificial sequence

<220>

<223> Artificial sequence description: synthetic

oligonucleotide

<220>

<221> modified base

<222> (20)..(21)

<223> 2'-methoxynucleotide

<400> 294

ggugcucaau aaaugauucu u 21

<210> 295

<211> 21

<212> RNA

<213> Artificial sequence

<220>

<223> Artificial sequence description: synthetic

oligonucleotide

<220>

<221> modified base

<222> (20)..(21)

<223> 2'-methoxynucleotide

<400> 295

caucaucaaa aacuuuggau u 21

<210> 296

<211> 21

<212> RNA

<213> Artificial sequence

<220>

<223> Artificial sequence description: synthetic

oligonucleotide

<220>

<221> modified base

<222> (20)..(21)

<223> 2'-methoxynucleotide

<400> 296

aaggagucag acauuuuaau u 21

<210> 297

<211> 21

<212> RNA

<213> Artificial sequence

<220>

<223> Artificial sequence description: synthetic

oligonucleotide

<220>

<221> modified base

<222> (20)..(21)

<223> 2'-methoxynucleotide

<400> 297

gugcugggca uuuuuauuuu u 21

<210> 298

<211> 21

<212> RNA

<213> Artificial sequence

<220>

<223> Artificial sequence description: synthetic

oligonucleotide

<220>

<221> modified base

<222> (20)..(21)

<223> 2'-methoxynucleotide

<400> 298

gccggcuuca ugccagcuau u 21

<210> 299

<211> 21

<212> RNA

<213> Artificial sequence

<220>

<223> Artificial sequence description: synthetic

oligonucleotide

<220>

<221> modified base

<222> (20)..(21)

<223> 2'-methoxynucleotide

<400> 299

gggcaucauc aaaaacuuuu u 21

<210> 300

<211> 21

<212> RNA

<213> Artificial sequence

<220>

<223> Artificial sequence description: synthetic

oligonucleotide

<220>

<221> modified base

<222> (20)..(21)

<223> 2'-methoxynucleotide

<400> 300

gaagggcacc cuaguucuau u 21

<210> 301

<211> 21

<212> RNA

<213> Artificial sequence

<220>

<223> Artificial sequence description: synthetic

oligonucleotide

<220>

<221> modified base

<222> (20)..(21)

<223> 2'-methoxynucleotide

<400> 301

caguucauug cacuuugauu u 21

<210> 302

<211> 21

<212> RNA

<213> Artificial sequence

<220>

<223> Artificial sequence description: synthetic

oligonucleotide

<220>

<221> modified base

<222> (20)..(21)

<223> 2'-methoxynucleotide

<400> 302

acaaggaguc agacauuuuu u 21

<210> 303

<211> 21

<212> RNA

<213> Artificial sequence

<220>

<223> Artificial sequence description: synthetic

oligonucleotide

<220>

<221> modified base

<222> (20)..(21)

<223> 2'-methoxynucleotide

<400> 303

uggaggcacu gaagugcuuu u 21

<210> 304

<211> 21

<212> RNA

<213> Artificial sequence

<220>

<223> Artificial sequence description: synthetic

oligonucleotide

<220>

<221> modified base

<222> (20)..(21)

<223> 2'-methoxynucleotide

<400> 304

gcagggacca cacccuguau u 21

<210> 305

<211> 21

<212> RNA

<213> Artificial sequence

<220>

<223> Artificial sequence description: synthetic

oligonucleotide

<220>

<221> modified base

<222> (20)..(21)

<223> 2'-methoxynucleotide

<400> 305

cuguacuguu cugugucuuuuu 21

<210> 306

<211> 21

<212> RNA

<213> Artificial sequence

<220>

<223> Artificial sequence description: synthetic

oligonucleotide

<220>

<221> modified base

<222> (20)..(21)

<223> 2'-methoxynucleotide

<400> 306

uuaaacaccu ccucauguau u 21

<210> 307

<211> 21

<212> RNA

<213> Artificial sequence

<220>

<223> Artificial sequence description: synthetic

oligonucleotide

<220>

<221> modified base

<222> (20)..(21)

<223> 2'-methoxynucleotide

<400> 307

agacucucag ggucgaaaau u 21

<210> 308

<211> 21

<212> RNA

<213> Artificial sequence

<220>

<223> Artificial sequence description: synthetic

oligonucleotide

<220>

<221> modified base

<222> (20)..(21)

<223> 2'-methoxynucleotide

<400> 308

caugacagau uucuaccacu u 21

<210> 309

<211> 21

<212> RNA

<213> Artificial sequence

<220>

<223> Artificial sequence description: synthetic

oligonucleotide

<220>

<221> modified base

<222> (20)..(21)

<223> 2'-methoxynucleotide

<400> 309

agauuucuac cacuccaaau u 21

<210> 310

<211> 21

<212> RNA

<213> Artificial sequence

<220>

<223> Artificial sequence description: synthetic

oligonucleotide

<220>

<221> modified base

<222> (20)..(21)

<223> 2'-methoxynucleotide

<400> 310

ccaagaggaa gcccuaaucu u 21

<210> 311

<211> 21

<212> RNA

<213> Artificial sequence

<220>

<223> Artificial sequence description: synthetic

oligonucleotide

<220>

<221> modified base

<222> (20)..(21)

<223> 2'-methoxynucleotide

<400> 311

gacagcagag gaagaccauu u 21

<210> 312

<211> 21

<212> RNA

<213> Artificial sequence

<220>

<223> Artificial sequence description: synthetic

oligonucleotide

<220>

<221> modified base

<222> (20)..(21)

<223> 2'-methoxynucleotide

<400> 312

cucccacaau gcugaauauu u 21

<210> 313

<211> 21

<212> RNA

<213> Artificial sequence

<220>

<223> Artificial sequence description: synthetic

oligonucleotide

<220>

<221> modified base

<222> (20)..(21)

<223> 2'-methoxynucleotide

<400> 313

uacaaguaaa gucacuaagu u 21

<210> 314

<211> 21

<212> RNA

<213> Artificial sequence

<220>

<223> Artificial sequence description: synthetic

oligonucleotide

<220>

<221> modified base

<222> (20)..(21)

<223> 2'-methoxynucleotide

<400> 314

aaucugucuau gcuggucugu u 21

<210> 315

<211> 21

<212> RNA

<213> Artificial sequence

<220>

<223> Artificial sequence description: synthetic

oligonucleotide

<220>

<221> modified base

<222> (20)..(21)

<223> 2'-methoxynucleotide

<400> 315

uuccucuugg agaagaucau u 21

<210> 316

<211> 21

<212> RNA

<213> Artificial sequence

<220>

<223> Artificial sequence description: synthetic

oligonucleotide

<220>

<221> modified base

<222> (20)..(21)

<223> 2'-methoxynucleotide

<400> 316

uaaucaaagu gcaaugaacu u 21

<210> 317

<211> 21

<212> RNA

<213> Artificial sequence

<220>

<223> Artificial sequence description: synthetic

oligonucleotide

<220>

<221> modified base

<222> (20)..(21)

<223> 2'-methoxynucleotide

<400> 317

ugcuaucaa agugcaaugu u 21

<210> 318

<211> 21

<212> RNA

<213> Artificial sequence

<220>

<223> Artificial sequence description: synthetic

oligonucleotide

<220>

<221> modified base

<222> (20)..(21)

<223> 2'-methoxynucleotide

<400> 318

aagucgaagu uccaucgcuu u 21

<210> 319

<211> 21

<212> RNA

<213> Artificial sequence

<220>

<223> Artificial sequence description: synthetic

oligonucleotide

<220>

<221> modified base

<222> (20)..(21)

<223> 2'-methoxynucleotide

<400> 319

aaagucgaag uuccaucgcu u 21

<210> 320

<211> 21

<212> RNA

<213> Artificial sequence

<220>

<223> Artificial sequence description: synthetic

oligonucleotide

<220>

<221> modified base

<222> (20)..(21)

<223> 2'-methoxynucleotide

<400> 320

uucuugugug ucccuucccu u 21

<210> 321

<211> 21

<212> RNA

<213> Artificial sequence

<220>

<223> Artificial sequence description: synthetic

oligonucleotide

<220>

<221> modified base

<222> (20)..(21)

<223> 2'-methoxynucleotide

<400> 321

uugagcugcc ugagguagau u 21

<210> 322

<211> 21

<212> RNA

<213> Artificial sequence

<220>

<223> Artificial sequence description: synthetic

oligonucleotide

<220>

<221> modified base

<222> (20)..(21)

<223> 2'-methoxynucleotide

<400> 322

gaaucauuua uugagcaccu u 21

<210> 323

<211> 21

<212> RNA

<213> Artificial sequence

<220>

<223> Artificial sequence description: synthetic

oligonucleotide

<220>

<221> modified base

<222> (20)..(21)

<223> 2'-methoxynucleotide

<400> 323

uccaaaguuu uugaugaugu u 21

<210> 324

<211> 21

<212> RNA

<213> Artificial sequence

<220>

<223> Artificial sequence description: synthetic

oligonucleotide

<220>

<221> modified base

<222> (20)..(21)

<223> 2'-methoxynucleotide

<400> 324

uuaaaauguc ugacuccuuu u 21

<210> 325

<211> 21

<212> RNA

<213> Artificial sequence

<220>

<223> Artificial sequence description: synthetic

oligonucleotide

<220>

<221> modified base

<222> (20)..(21)

<223> 2'-methoxynucleotide

<400> 325

aaauaaaaau gccagcacu u 21

<210> 326

<211> 21

<212> RNA

<213> Artificial sequence

<220>

<223> Artificial sequence description: synthetic

oligonucleotide

<220>

<221> modified base

<222> (20)..(21)

<223> 2'-methoxynucleotide

<400> 326

uagcuggcau gaagccggcu u 21

<210> 327

<211> 21

<212> RNA

<213> Artificial sequence

<220>

<223> Artificial sequence description: synthetic

oligonucleotide

<220>

<221> modified base

<222> (20)..(21)

<223> 2'-methoxynucleotide

<400> 327

aaaguuuuug augaugcccu u 21

<210> 328

<211> 21

<212> RNA

<213> Artificial sequence

<220>

<223> Artificial sequence description: synthetic

oligonucleotide

<220>

<221> modified base

<222> (20)..(21)

<223> 2'-methoxynucleotide

<400> 328

uagaacuagg gugcccuucu u 21

<210> 329

<211> 21

<212> RNA

<213> Artificial sequence

<220>

<223> Artificial sequence description: synthetic

oligonucleotide

<220>

<221> modified base

<222> (20)..(21)

<223> 2'-methoxynucleotide

<400> 329

aucaaagugc aaugaacugu u 21

<210> 330

<211> 21

<212> RNA

<213> Artificial sequence

<220>

<223> Artificial sequence description: synthetic

oligonucleotide

<220>

<221> modified base

<222> (20)..(21)

<223> 2'-methoxynucleotide

<400> 330

aaaaugucug acuccuuguuu 21

<210> 331

<211> 21

<212> RNA

<213> Artificial sequence

<220>

<223> Artificial sequence description: synthetic

oligonucleotide

<220>

<221> modified base

<222> (20)..(21)

<223> 2'-methoxynucleotide

<400> 331

aagcacuuca gugccuccau u 21

<210> 332

<211> 21

<212> RNA

<213> Artificial sequence

<220>

<223> Artificial sequence description: synthetic

oligonucleotide

<220>

<221> modified base

<222> (20)..(21)

<223> 2'-methoxynucleotide

<400> 332

uacagggugu ggucccugcu u 21

<210> 333

<211> 21

<212> RNA

<213> Artificial sequence

<220>

<223> Artificial sequence description: synthetic

oligonucleotide

<220>

<221> modified base

<222> (20)..(21)

<223> 2'-methoxynucleotide

<400> 333

aagacacaga acaguacagu u 21

<210> 334

<211> 21

<212> RNA

<213> Artificial sequence

<220>

<223> Artificial sequence description: synthetic

oligonucleotide

<220>

<221> modified base

<222> (20)..(21)

<223> 2'-methoxynucleotide

<400> 334

uacaugagga gguguuuaau u 21

<210> 335

<211> 21

<212> RNA

<213> Artificial sequence

<220>

<223> Artificial sequence description: synthetic

oligonucleotide

<220>

<221> modified base

<222> (20)..(21)

<223> 2'-methoxynucleotide

<400> 335

uuuucgaccc ugagagucuu u 21

<210> 336

<211> 21

<212> RNA

<213> Artificial sequence

<220>

<223> Artificial sequence description: synthetic

oligonucleotide

<220>

<221> modified base

<222> (20)..(21)

<223> 2'-methoxynucleotide

<400> 336

gugguagaaa ucugucaugu u 21

<210> 337

<211> 21

<212> RNA

<213> Artificial sequence

<220>

<223> Artificial sequence description: synthetic

oligonucleotide

<220>

<221> modified base

<222> (20)..(21)

<223> 2'-methoxynucleotide

<400> 337

uuuggagugg uagaaaucuu u 21

<210> 338

<211> 21

<212> RNA

<213> Artificial sequence

<220>

<223> Artificial sequence description: synthetic

oligonucleotide

<220>

<221> modified base

<222> (20)..(21)

<223> 2'-methoxynucleotide

<400> 338

gauuagggcu uccucuuggu u 21

<210> 339

<211> 21

<212> RNA

<213> Artificial sequence

<220>

<223> Artificial sequence description: synthetic

oligonucleotide

<220>

<221> modified base

<222> (20)..(21)

<223> 2'-methoxynucleotide

<400> 339

auggucuucc ucugcugucu u 21

<210> 340

<211> 21

<212> RNA

<213> Artificial sequence

<220>

<223> Artificial sequence description: synthetic

oligonucleotide

<220>

<221> modified base

<222> (20)..(21)

<223> 2'-methoxynucleotide

<400> 340

auauucagca uuggggagu u 21

<210> 341

<211> 21

<212> DNA

<213> Artificial sequence

<220>

<223> Artificial sequence description: synthetic

oligonucleotide

<220>

<223> Description of the combined DNA/RNA molecule: synthetic

oligonucleotide

<220>

<221> modified base

<222> (20)..(21)

<223> a, c, t, g, u, unknown or other

<400> 341

aaggagucag acauuuuaann 21

<210> 342

<211> 21

<212> RNA

<213> Artificial sequence

<220>

<223> Artificial sequence description: synthetic

oligonucleotide

<220>

<221> modified base

<222> (1)..(3)

<223> 2'-OMe-nucleotide

<220>

<221> modified base

<222>(8)..(8)

<223> 2'-OMe-nucleotide

<220>

<221> modified base

<222> (12)..(12)

<223> 2'-OMe-nucleotide

<220>

<221> modified base

<222> (17)..(17)

<223> 2'-OMe-nucleotide

<220>

<221> modified base

<222> (20)..(21)

<223> 2'-OMe-nucleotide

<400> 342

aaggagucag acauuuuaau u 21

<210> 343

<211> 21

<212> RNA

<213> Artificial sequence

<220>

<223> Artificial sequence description: synthetic

oligonucleotide

<220>

<221> modified base

<222> (1)..(3)

<223> 2'-OMe-nucleotide

<220>

<221> modified base

<222>(8)..(8)

<223> 2'-OMe-nucleotide

<220>

<221> modified base

<222> (12)..(12)

<223> 2'-OMe-nucleotide

<220>

<221> modified base

<222> (14)..(15)

<223> 2'-OMe-nucleotide

<220>

<221> modified base

<222> (17)..(17)

<223> 2'-OMe-nucleotide

<220>

<221> modified base

<222> (20)..(21)

<223> 2'-OMe-nucleotide

<400> 343

aaggagucag acauuuuaau u 21

<210> 344

<211> 21

<212> RNA

<213> Artificial sequence

<220>

<223> Artificial sequence description: synthetic

oligonucleotide

<220>

<221> modified base

<222> (1)..(3)

<223> 2'-OMe-nucleotide

<220>

<221> modified base

<222>(8)..(8)

<223> 2'-OMe-nucleotide

<220>

<221> modified base

<222> (12)..(12)

<223> 2'-OMe-nucleotide

<220>

<221> modified base

<222> (14)..(15)

<223> 2'-OMe-nucleotide

<220>

<221> modified base

<222> (17)..(17)

<223> 2'-OMe-nucleotide

<220>

<221> modified base

<222> (20)..(21)

<223> 2'-OMe-nucleotide

<400> 344

aaggagucag acauuuuaau u 21

<210> 345

<211> 21

<212> RNA

<213> Artificial sequence

<220>

<223> Artificial sequence description: synthetic

oligonucleotide

<220>

<221> modified base

<222> (1)..(3)

<223> 2'-OMe-nucleotide

<220>

<221> modified base

<222>(8)..(8)

<223> 2'-OMe-nucleotide

<220>

<221> modified base

<222> (12)..(12)

<223> 2'-OMe-nucleotide

<220>

<221> modified base

<222> (17)..(17)

<223> 2'-OMe-nucleotide

<220>

<221> modified base

<222> (20)..(21)

<223> 2'-OMe-nucleotide

<400> 345

aaggagucag acauuuuaau u 21

<210> 346

<211> 21

<212> RNA

<213> Artificial sequence

<220>

<223> Artificial sequence description: synthetic

oligonucleotide

<220>

<221> modified base

<222> (20)..(21)

<223> 2'-OMe-nucleotide

<400> 346

aaggagucag acauuuuaau u 21

<210> 347

<211> 21

<212> RNA

<213> Artificial sequence

<220>

<223> Artificial sequence description: synthetic

oligonucleotide

<220>

<221> modified base

<222> (20)..(21)

<223> 2'-OMe-nucleotide

<400> 347

aaggagucag acauuuuaau u 21

<210> 348

<211> 21

<212> RNA

<213> Artificial sequence

<220>

<223> Artificial sequence description: synthetic

oligonucleotide

<220>

<221> modified base

<222> (20)..(21)

<223> 2'-OMe-nucleotide

<400> 348

aaggagucag acauuuuaau u 21

<210> 349

<211> 21

<212> RNA

<213> Artificial sequence

<220>

<223> Artificial sequence description: synthetic

oligonucleotide

<220>

<221> modified base

<222> (20)..(21)

<223> 2'-OMe-nucleotide

<400> 349

aaggagucag acauuuuaau u 21

<210> 350

<211> 21

<212> RNA

<213> Artificial sequence

<220>

<223> Artificial sequence description: synthetic

oligonucleotide

<220>

<221> modified base

<222> (20)..(21)

<223> 2'-OMe-nucleotide

<400> 350

aaggagucag acauuuuaau u 21

<210> 351

<211> 21

<212> RNA

<213> Artificial sequence

<220>

<223> Artificial sequence description: synthetic

oligonucleotide

<220>

<221> modified base

<222> (20)..(21)

<223> 2'-OMe-nucleotide

<400> 351

aaggagucag acauuuuaau u 21

<210> 352

<211> 21

<212> RNA

<213> Artificial sequence

<220>

<223> Artificial sequence description: synthetic

oligonucleotide

<220>

<221> modified base

<222>(8)..(8)

<223> 2'-OMe-nucleotide

<220>

<221> modified base

<222> (12)..(12)

<223> 2'-OMe-nucleotide

<220>

<221> modified base

<222> (17)..(17)

<223> 2'-OMe-nucleotide

<220>

<221> modified base

<222> (20)..(21)

<223> 2'-OMe-nucleotide

<400> 352

aaggagucag acauuuuaau u 21

<210> 353

<211> 21

<212> RNA

<213> Artificial sequence

<220>

<223> Artificial sequence description: synthetic

oligonucleotide

<220>

<221> modified base

<222> (1)..(3)

<223> 2'-OMe-nucleotide

<220>

<221> modified base

<222> (20)..(21)

<223> 2'-OMe-nucleotide

<400> 353

aaggagucag acauuuuaau u 21

<210> 354

<211> 21

<212> RNA

<213> Artificial sequence

<220>

<223> Artificial sequence description: synthetic

oligonucleotide

<220>

<221> modified base

<222> (1)..(3)

<223> 2'-OMe-nucleotide

<220>

<221> modified base

<222>(8)..(8)

<223> 2'-OMe-nucleotide

<220>

<221> modified base

<222> (12)..(12)

<223> 2'-OMe-nucleotide

<220>

<221> modified base

<222> (17)..(17)

<223> 2'-OMe-nucleotide

<220>

<221> modified base

<222> (20)..(21)

<223> 2'-OMe-nucleotide

<400> 354

aaggagucag acauuuuaau u 21

<210> 355

<211> 21

<212> DNA

<213> Artificial sequence

<220>

<223> Artificial sequence description: synthetic

oligonucleotide

<220>

<223> Description of the combined DNA/RNA molecule: synthetic

oligonucleotide

<220>

<221> modified base

<222> (20)..(21)

<223> a, c, t, g, u, unknown or other

<400> 355

uuaaaauguc ugacuccuun n 21

<210> 356

<211> 21

<212> DNA

<213> Artificial sequence

<220>

<223> Artificial sequence description: synthetic

oligonucleotide

<220>

<223> Description of the combined DNA/RNA molecule: synthetic

oligonucleotide

<220>

<221> modified base

<222> (2)..(2)

<223> 2'-OMe-nucleotide

<220>

<221> modified base

<222>(7)..(7)

<223> 2'-OMe-nucleotide

<220>

<221> modified base

<222> (11)..(11)

<223> 2'-OMe-nucleotide

<220>

<221> modified base

<222> (20)..(21)

<223> 2'-OMe-nucleotide

<400> 356

uuaaaauguc ugacuccuuu u 21

<210> 357

<211> 21

<212> DNA

<213> Artificial sequence

<220>

<223> Artificial sequence description: synthetic

oligonucleotide

<220>

<223> Description of the combined DNA/RNA molecule: synthetic

oligonucleotide

<220>

<221> modified base

<222> (2)..(2)

<223> 2'-OMe-nucleotide

<220>

<221> modified base

<222>(7)..(7)

<223> 2'-OMe-nucleotide

<220>

<221> modified base

<222> (11)..(11)

<223> 2'-OMe-nucleotide

<220>

<221> modified base

<222> (20)..(21)

<223> 2'-OMe-nucleotide

<400> 357

uuaaaauguc ugacuccuuu u 21

<210> 358

<211> 21

<212> DNA

<213> Artificial sequence

<220>

<223> Artificial sequence description: synthetic

oligonucleotide

<220>

<223> Description of the combined DNA/RNA molecule: synthetic

oligonucleotide

<220>

<221> modified base

<222> (2)..(2)

<223> 2'-OMe-nucleotide

<220>

<221> modified base

<222> (20)..(21)

<223> 2'-OMe-nucleotide

<400> 358

uuaaaauguc ugacuccuuu u 21

<210> 359

<211> 21

<212> DNA

<213> Artificial sequence

<220>

<223> Artificial sequence description: synthetic

oligonucleotide

<220>

<223> Description of the combined DNA/RNA molecule: synthetic

oligonucleotide

<220>

<221> modified base

<222> (2)..(2)

<223> 2'-OMe-nucleotide

<220>

<221> modified base

<222> (20)..(21)

<223> 2'-OMe-nucleotide

<400> 359

uuaaaauguc ugacuccuuu u 21

<210> 360

<211> 21

<212> DNA

<213> Artificial sequence

<220>

<223> Artificial sequence description: synthetic

oligonucleotide

<220>

<223> Description of the combined DNA/RNA molecule: synthetic

oligonucleotide

<220>

<221> modified base

<222>(7)..(7)

<223> 2'-deoxynucleotide

<220>

<221> modified base

<222> (20)..(21)

<223> 2'-OMe-nucleotide

<400> 360

uuaaaauguc ugacuccuuu u 21

<210> 361

<211> 21

<212> DNA

<213> Artificial sequence

<220>

<223> Artificial sequence description: synthetic

oligonucleotide

<220>

<223> Description of the combined DNA/RNA molecule: synthetic

oligonucleotide

<220>

<221> modified base

<222>(7)..(7)

<223> 2'-deoxynucleotide

<220>

<221> modified base

<222> (20)..(21)

<223> 2'-OMe-nucleotide

<400> 361

uuaaaauguc ugacuccuuu u 21

<210> 362

<211> 21

<212> DNA

<213> Artificial sequence

<220>

<223> Artificial sequence description: synthetic

oligonucleotide

<220>

<223> Description of the combined DNA/RNA molecule: synthetic

oligonucleotide

<220>

<221> modified base

<222> (1)..(1)

<223> 2'-deoxynucleotide

<220>

<221> modified base

<222>(7)..(7)

<223> 2'-deoxynucleotide

<220>

<221> modified base

<222> (20)..(21)

<223> 2'-OMe-nucleotide

<400> 362

uuaaaauguc ugacuccuuu u 21

<210> 363

<211> 21

<212> DNA

<213> Artificial sequence

<220>

<223> Artificial sequence description: synthetic

oligonucleotide

<220>

<223> Description of the combined DNA/RNA molecule: synthetic

oligonucleotide

<220>

<221> modified base

<222>(7)..(7)

<223> 2'-deoxynucleotide

<220>

<221> modified base

<222> (20)..(21)

<223> 2'-OMe-nucleotide

<400> 363

uuaaaauguc ugacuccuuu u 21

<210> 364

<211> 21

<212> DNA

<213> Artificial sequence

<220>

<223> Artificial sequence description: synthetic

oligonucleotide

<220>

<223> Description of the combined DNA/RNA molecule: synthetic

oligonucleotide

<220>

<221> modified base

<222> (20)..(21)

<223> 2'-OMe-nucleotide

<400> 364

uuaaaauguc ugacuccuuu u 21

<210> 365

<211> 21

<212> RNA

<213> Artificial sequence

<220>

<223> Artificial sequence description: synthetic

oligonucleotide

<220>

<221> modified base

<222> (2)..(2)

<223> 2'-OMe-nucleotide

<220>

<221> modified base

<222>(7)..(7)

<223> 2'-OMe-nucleotide

<220>

<221> modified base

<222> (11)..(11)

<223> 2'-OMe-nucleotide

<220>

<221> modified base

<222> (20)..(21)

<223> 2'-OMe-nucleotide

<400> 365

uuaaaauguc ugacuccuuu u 21

<210> 366

<211> 21

<212> RNA

<213> Artificial sequence

<220>

<223> Artificial sequence description: synthetic

oligonucleotide

<220>

<221> modified base

<222> (20)..(21)

<223> 2'-OMe-nucleotide

<400> 366

uuaaaauguc ugacuccuuu u 21

<210> 367

<211> 21

<212> RNA

<213> Artificial sequence

<220>

<223> Artificial sequence description: synthetic

oligonucleotide

<220>

<221> modified base

<222> (20)..(21)

<223> 2'-OMe-nucleotide

<400> 367

uuaaaauguc ugacuccuuu u 21

<210> 368

<211> 21

<212> RNA

<213> Artificial sequence

<220>

<223> Artificial sequence description: synthetic

oligonucleotide

<220>

<221> modified base

<222> (20)..(21)

<223> 2'-OMe-nucleotide

<400> 368

uuaaaauguc ugacuccuuu u 21

<210> 369

<211> 21

<212> RNA

<213> Artificial sequence

<220>

<223> Artificial sequence description: synthetic

oligonucleotide

<400> 369

cgagaacagu uuguacaagu u 21

<210> 370

<211> 21

<212> DNA

<213> Artificial sequence

<220>

<223> Artificial sequence description: synthetic

oligonucleotide

<220>

<223> Description of the combined DNA/RNA molecule: synthetic

oligonucleotide

<400> 370

caggccucua caacuacuat t 21

<210> 371

<211> 25

<212> RNA

<213> Artificial sequence

<220>

<223> Artificial sequence description: synthetic

oligonucleotide

<400> 371

gagcacucca agaucaacuu ccgcg 25

<210> 372

<211> 21

<212> DNA

<213> Artificial sequence

<220>

<223> Artificial sequence description: synthetic

oligonucleotide

<220>

<223> Description of the combined DNA/RNA molecule: synthetic

oligonucleotide

<400> 372

ggacaggccu cuacaacuat t 21

<210> 373

<211> 21

<212> DNA

<213> Artificial sequence

<220>

<223> Artificial sequence description: synthetic

oligonucleotide

<220>

<223> Description of the combined DNA/RNA molecule: synthetic

oligonucleotide

<400> 373

gagcacucca agaucaacut t 21

<210> 374

<211> 21

<212> DNA

<213> Artificial sequence

<220>

<223> Artificial sequence description: synthetic

oligonucleotide

<220>

<223> Description of the combined DNA/RNA molecule: synthetic

oligonucleotide

<400> 374

gaacacucca agaucacut t 21

<210> 375

<211> 25

<212> RNA

<213> Artificial sequence

<220>

<223> Artificial sequence description: synthetic

oligonucleotide

<400> 375

caggccucua caacuacuac gacga 25

<210> 376

<211> 25

<212> RNA

<213> Artificial sequence

<220>

<223> Artificial sequence description: synthetic

oligonucleotide

<400> 376

gaacacucca agaucaacuu ccgag 25

<210> 377

<211> 25

<212> RNA

<213> Artificial sequence

<220>

<223> Artificial sequence description: synthetic

oligonucleotide

<400> 377

ggacaggccu cuacaacuac uacga 25

<210> 378

<211> 26

<212> DNA

<213> Artificial sequence

<220>

<223> Artificial sequence description: synthetic

oligonucleotide

<220>

<223> Description of the combined DNA/RNA molecule: synthetic

oligonucleotide

<400> 378

caggccucua caacuacuat taaaaa 26

<210> 379

<211> 19

<212> RNA

<213> Artificial sequence

<220>

<223> Artificial sequence description: synthetic

oligonucleotide

<400> 379

caggccucua caacuacua 19

<210> 380

<211> 33

<212> DNA

<213> Artificial sequence

<220>

<223> Artificial sequence description: synthetic

oligonucleotide

<220>

<223> Description of the combined DNA/RNA molecule: synthetic

oligonucleotide

<400> 380

caggccucua caacuacuat taaaaaaaaa aaa 33

<210> 381

<211> 26

<212> DNA

<213> Artificial sequence

<220>

<223> Artificial sequence description: synthetic

oligonucleotide

<220>

<223> Description of the combined DNA/RNA molecule: synthetic

oligonucleotide

<400> 381

aaaaacaggc cucuacaacu acuatt 26

<210> 382

<211> 29

<212> DNA

<213> Artificial sequence

<220>

<223> Artificial sequence description: synthetic

oligonucleotide

<220>

<223> Description of the combined DNA/RNA molecule: synthetic

oligonucleotide

<400> 382

caggccucua caacuacuat taaaaaaaa 29

<210> 383

<211> 29

<212> DNA

<213> Artificial sequence

<220>

<223> Artificial sequence description: synthetic

oligonucleotide

<220>

<223> Description of the combined DNA/RNA molecule: synthetic

oligonucleotide

<400> 383

aaaaaaaaca ggccucuaca acuacuatt 29

<210> 384

<211> 33

<212> DNA

<213> Artificial sequence

<220>

<223> Artificial sequence description: synthetic

oligonucleotide

<220>

<223> Description of the combined DNA/RNA molecule: synthetic

oligonucleotide

<400> 384

caggccucua caacuacuat taaaaaaaaa aaa 33

<210> 385

<211> 37

<212> DNA

<213> Artificial sequence

<220>

<223> Artificial sequence description: synthetic

oligonucleotide

<220>

<223> Description of the combined DNA/RNA molecule: synthetic

oligonucleotide

<400> 385

aaaaaaaaaa aaaaaacagg ccucuacaac uacuatt 37

<210> 386

<211> 37

<212> DNA

<213> Artificial sequence

<220>

<223> Artificial sequence description: synthetic

oligonucleotide

<220>

<223> Description of the combined DNA/RNA molecule: synthetic

oligonucleotide

<400> 386

caggccucua caacuacuat taaaaaaaaa aaaaaaa 37

<210> 387

<211> 21

<212> DNA

<213> Artificial sequence

<220>

<223> Artificial sequence description: synthetic

oligonucleotide

<220>

<223> Description of the combined DNA/RNA molecule: synthetic

oligonucleotide

<400> 387

caggccucua caacuacuat t 21

<210> 388

<211> 21

<212> DNA

<213> Artificial sequence

<220>

<223> Artificial sequence description: synthetic

oligonucleotide

<220>

<223> Description of the combined DNA/RNA molecule: synthetic

oligonucleotide

<400> 388

ggacaggccu guacaacuat t 21

<210> 389

<211> 21

<212> DNA

<213> Artificial sequence

<220>

<223> Artificial sequence description: synthetic

oligonucleotide

<220>

<223> Description of the combined DNA/RNA molecule: synthetic

oligonucleotide

<400> 389

ggacaggccu cuacaacuat t 21

<210> 390

<211> 21

<212> RNA

<213> Artificial sequence

<220>

<223> Artificial sequence description: synthetic

oligonucleotide

<400> 390

cuuguacaaa cuguucucgu u 21

<210> 391

<211> 21

<212> DNA

<213> Artificial sequence

<220>

<223> Artificial sequence description: synthetic

oligonucleotide

<220>

<223> Description of the combined DNA/RNA molecule: synthetic

oligonucleotide

<400> 391

uaguaguugu agaggccugt t 21

<210> 392

<211> 27

<212> RNA

<213> Artificial sequence

<220>

<223> Artificial sequence description: synthetic

oligonucleotide

<400> 392

cgcggaaguu gaucuuggag ugcucuu 27

<210> 393

<211> 21

<212> DNA

<213> Artificial sequence

<220>

<223> Artificial sequence description: synthetic

oligonucleotide

<220>

<223> Description of the combined DNA/RNA molecule: synthetic

oligonucleotide

<400> 393

uaguuguaga ggccugucct t 21

<210> 394

<211> 21

<212> DNA

<213> Artificial sequence

<220>

<223> Artificial sequence description: synthetic

oligonucleotide

<220>

<223> Description of the combined DNA/RNA molecule: synthetic

oligonucleotide

<400> 394

aguugaucuu ggagugcuct t 21

<210> 395

<211> 21

<212> DNA

<213> Artificial sequence

<220>

<223> Artificial sequence description: synthetic

oligonucleotide

<220>

<223> Description of the combined DNA/RNA molecule: synthetic

oligonucleotide

<400> 395

aguugaucuu ggaguguuct t 21

<210> 396

<211> 27

<212> RNA

<213> Artificial sequence

<220>

<223> Artificial sequence description: synthetic

oligonucleotide

<400> 396

ucgucguagu aguuguagag gccuguu 27

<210> 397

<211> 27

<212> RNA

<213> Artificial sequence

<220>

<223> Artificial sequence description: synthetic

oligonucleotide

<400> 397

cucggaaguu gaucuuggag uguucuu 27

<210> 398

<211> 27

<212> RNA

<213> Artificial sequence

<220>

<223> Artificial sequence description: synthetic

oligonucleotide

<400> 398

ucguaguagu ugugaggcc uguccuu 27

<210> 399

<211> 21

<212> DNA

<213> Artificial sequence

<220>

<223> Artificial sequence description: synthetic

oligonucleotide

<220>

<223> Description of the combined DNA/RNA molecule: synthetic

oligonucleotide

<400> 399

uaguaguugu agaggccugt t 21

<210> 400

<211> 21

<212> DNA

<213> Artificial sequence

<220>

<223> Artificial sequence description: synthetic

oligonucleotide

<220>

<223> Description of the combined DNA/RNA molecule: synthetic

oligonucleotide

<400> 400

uaguaguugu agaggccugt t 21

<210> 401

<211> 21

<212> DNA

<213> Artificial sequence

<220>

<223> Artificial sequence description: synthetic

oligonucleotide

<220>

<223> Description of the combined DNA/RNA molecule: synthetic

oligonucleotide

<400> 401

uaguaguugu agaggccugt t 21

<210> 402

<211> 21

<212> DNA

<213> Artificial sequence

<220>

<223> Artificial sequence description: synthetic

oligonucleotide

<220>

<223> Description of the combined DNA/RNA molecule: synthetic

oligonucleotide

<400> 402

uaguaguugu agaggccugt t 21

<210> 403

<211> 21

<212> DNA

<213> Artificial sequence

<220>

<223> Artificial sequence description: synthetic

oligonucleotide

<220>

<223> Description of the combined DNA/RNA molecule: synthetic

oligonucleotide

<400> 403

uaguaguugu agaggccugt t 21

<210> 404

<211> 21

<212> DNA

<213> Artificial sequence

<220>

<223> Artificial sequence description: synthetic

oligonucleotide

<220>

<223> Description of the combined DNA/RNA molecule: synthetic

oligonucleotide

<400> 404

uaguaguugu agaggccugt t 21

<210> 405

<211> 21

<212> DNA

<213> Artificial sequence

<220>

<223> Artificial sequence description: synthetic

oligonucleotide

<220>

<223> Description of the combined DNA/RNA molecule: synthetic

oligonucleotide

<400> 405

uaguaguugu agaggccugt t 21

<210> 406

<211> 21

<212> DNA

<213> Artificial sequence

<220>

<223> Artificial sequence description: synthetic

oligonucleotide

<220>

<223> Description of the combined DNA/RNA molecule: synthetic

oligonucleotide

<400> 406

uaguaguugu agaggccugt t 21

<210> 407

<211> 21

<212> DNA

<213> Artificial sequence

<220>

<223> Artificial sequence description: synthetic

oligonucleotide

<220>

<223> Description of the combined DNA/RNA molecule: synthetic

oligonucleotide

<400> 407

uaguaguugu agaggccugt t 21

<210> 408

<211> 21

<212> DNA

<213> Artificial sequence

<220>

<223> Artificial sequence description: synthetic

oligonucleotide

<220>

<223> Description of the combined DNA/RNA molecule: synthetic

oligonucleotide

<400> 408

uaguaguugu agaggccugt t 21

<210> 409

<211> 21

<212> DNA

<213> Artificial sequence

<220>

<223> Artificial sequence description: synthetic

oligonucleotide

<220>

<223> Description of the combined DNA/RNA molecule: synthetic

oligonucleotide

<400> 409

uaguuguaca ggccugucct t 21

<210> 410

<211> 21

<212> DNA

<213> Artificial sequence

<220>

<223> Artificial sequence description: synthetic

oligonucleotide

<220>

<223> Description of the combined DNA/RNA molecule: synthetic

oligonucleotide

<400> 410

uaguuguaga ggccugucct t 21

<210> 411

<211> 19

<212> RNA

<213> Artificial sequence

<220>

<223> Artificial sequence description: synthetic

oligonucleotide

<220>

<223> 5'-inverted 1,2-dideoxy-D-ribose

<220>

<221> mixed features

<222> (15)..(19)

<223> 2'-5' bond between nucleotides

<220>

<223> 3'-C3 spacer

<400> 411

acuccaagau caacuuccu 19

<210> 412

<211> 19

<212> RNA

<213> Artificial sequence

<220>

<223> Artificial sequence description: synthetic

oligonucleotide

<220>

<221> modified base

<222> (2)..(2)

<223> 2'-O-methylribonucleotide

<220>

<221> modified base

<222> (4)..(4)

<223> 2'-O-methylribonucleotide

<220>

<221> modified base

<222>(6)..(6)

<223> 2'-O-methylribonucleotide

<220>

<221> mixed features

<222> (7)..(8)

<223> 2'-5' bond between nucleotides

<220>

<221> modified base

<222>(8)..(8)

<223> 2'-O-methylribonucleotide

<220>

<221> modified base

<222> (11)..(11)

<223> 2'-O-methylribonucleotide

<220>

<221> modified base

<222> (13)..(13)

<223> 2'-O-methylribonucleotide

<220>

<221> modified base

<222> (15)..(15)

<223> 2'-O-methylribonucleotide

<220>

<221> modified base

<222> (17)..(17)

<223> 2'-O-methylribonucleotide

<220>

<221> modified base

<222> (19)..(19)

<223> 2'-O-methylribonucleotide

<220>

<223> 3'-C3-C3 spacer

<400> 412

aggaaguuga ucuuggagu 19

<210> 413

<211> 19

<212> RNA

<213> Artificial sequence

<220>

<223> Artificial sequence description: synthetic

oligonucleotide

<220>

<223> 5'-inverted 1,2-dideoxy-D-ribose

<220>

<221> modified base

<222> (4)..(4)

<223> 2'-O-methylribonucleotide

<220>

<221> mixed features

<222> (9)..(10)

<223> 2'-5' bond between nucleotides

<220>

<221> modified base

<222> (11)..(11)

<223> 2'-O-methylribonucleotide

<220>

<221> modified base

<222> (13)..(13)

<223> 2'-O-methylribonucleotide

<220>

<221> modified base

<222> (17)..(17)

<223> 2'-O-methylribonucleotide

<220>

<223> 3'-C3 spacer

<400> 413

uccugagaca cauggguga 19

<210> 414

<211> 19

<212> RNA

<213> Artificial sequence

<220>

<223> Artificial sequence description: synthetic

oligonucleotide

<220>

<221> modified base

<222> (1)..(1)

<223> 2'-O-methylribonucleotide

<220>

<221> modified base

<222> (4)..(4)

<223> 2'-O-methylribonucleotide

<220>

<221> mixed features

<222> (6)..(7)

<223> 2'-5' bond between nucleotides

<220>

<221> modified base

<222>(8)..(8)

<223> 2'-O-methylribonucleotide

<220>

<221> modified base

<222> (11)..(11)

<223> 2'-O-methylribonucleotide

<220>

<221> modified base

<222> (15)..(15)

<223> 2'-O-methylribonucleotide

<220>

<223> 3'-C3-C3 spacer

<400> 414

ucacccaugu gucucagga 19

<210> 415

<211> 19

<212> RNA

<213> Artificial sequence

<220>

<223> Artificial sequence description: synthetic

oligonucleotide

<220>

<223> 5'-inverted 1,2-dideoxy-D-ribose

<220>

<221> mixed features

<222> (15)..(19)

<223> 2'-5' bond between nucleotides

<220>

<223> 3'-C3-spacer-phosphate

<400> 415

gagacacaugggugcuaua 19

<210> 416

<211> 19

<212> RNA

<213> Artificial sequence

<220>

<223> Artificial sequence description: synthetic

oligonucleotide

<220>

<221> modified base

<222> (1)..(1)

<223> 2'-O-methylribonucleotide

<220>

<221> modified base

<222> (3)..(3)

<223> 2'-O-methylribonucleotide

<220>

<221> modified base

<222> (5)..(5)

<223> 2'-O-methylribonucleotide

<220>

<221> mixed features

<222> (7)..(8)

<223> 2'-5' bond between nucleotides

<220>

<221> modified base

<222>(9)..(9)

<223> 2'-O-methylribonucleotide

<220>

<221> modified base

<222> (11)..(11)

<223> 2'-O-methylribonucleotide

<220>

<221> modified base

<222> (13)..(13)

<223> 2'-O-methylribonucleotide

<220>

<221> modified base

<222> (15)..(15)

<223> 2'-O-methylribonucleotide

<220>

<221> modified base

<222> (17)..(17)

<223> 2'-O-methylribonucleotide

<220>

<221> modified base

<222> (19)..(19)

<223> 2'-O-methylribonucleotide

<220>

<223> 3'-C3-C3-spacer

<400> 416

uauagcaccc augugucuc 19

<210> 417

<211> 19

<212> RNA

<213> Artificial sequence

<220>

<223> Artificial sequence description: synthetic

oligonucleotide

<220>

<223> 5'-inverted 1,2-dideoxy-D-ribose

<220>

<221> modified base

<222> (2)..(3)

<223> 2'-O-methylribonucleotide

<220>

<221> modified base

<222> (5)..(5)

<223> 2'-O-methylribonucleotide

<220>

<221> modified base

<222> (7)..(8)

<223> 2'-O-methylribonucleotide

<220>

<221> mixed features

<222> (9)..(10)

<223> 2'-5' bond between nucleotides

<220>

<221> modified base

<222> (12)..(12)

<223> 2'-O-methylribonucleotide

<220>

<221> modified base

<222> (14)..(15)

<223> 2'-O-methylribonucleotide

<220>

<221> modified base

<222> (17)..(17)

<223> 2'-O-methylribonucleotide

<220>

<223> 3'-C3-spacer

<400> 417

cuuacgcuga guacuucgu 19

<210> 418

<211> 19

<212> RNA

<213> Artificial sequence

<220>

<223> Artificial sequence description: synthetic

oligonucleotide

<220>

<221> modified base

<222> (2)..(2)

<223> 2'-O-methylribonucleotide

<220>

<221> mixed signs

<222> (6)..(7)

<223> 2'-5' bond between nucleotides

<220>

<221> modified base

<222>(7)..(7)

<223> 2'-O-methylribonucleotide

<220>

<221> modified base

<222> (11)..(11)

<223> 2'-O-methylribonucleotide

<220>

<221> modified base

<222> (16)..(16)

<223> 2'-O-methylribonucleotide

<220>

<223> 3'-C3-C3-spacer

<400> 418

acgaaguacu cagcguaag 19

<210> 419

<211> 21

<212> RNA

<213> Artificial sequence

<220>

<223> Artificial sequence description: synthetic

oligonucleotide

<400> 419

gccuuccaag gauggguuug u 21

<210> 420

<211> 21

<212> RNA

<213> Artificial sequence

<220>

<223> Artificial sequence description: synthetic

oligonucleotide

<400> 420

ggaguucuuc cauguagagg a 21

<210> 421

<211> 21

<212> RNA

<213> Artificial sequence

<220>

<223> Artificial sequence description: synthetic

oligonucleotide

<400> 421

ccauguagag gaccuagaag g 21

<210> 422

<211> 21

<212> RNA

<213> Artificial sequence

<220>

<223> Artificial sequence description: synthetic

oligonucleotide

<400> 422

gccuuccaag gauggguuug u 21

<210> 423

<211> 21

<212> RNA

<213> Artificial sequence

<220>

<223> Artificial sequence description: synthetic

oligonucleotide

<400> 423

gccuuccaag gauggguuug u 21

<210> 424

<211> 21

<212> RNA

<213> Artificial sequence

<220>

<223> Artificial sequence description: synthetic

oligonucleotide

<400> 424

gccuuccaag gauggguuug u 21

<210> 425

<211> 21

<212> RNA

<213> Artificial sequence

<220>

<223> Artificial sequence description: synthetic

oligonucleotide

<400> 425

gccuuccaag gauggguuug u 21

<210> 426

<211> 21

<212> RNA

<213> Artificial sequence

<220>

<223> Artificial sequence description: synthetic

oligonucleotide

<400> 426

gccuuccaag gauggguuug u 21

<210> 427

<211> 21

<212> RNA

<213> Artificial sequence

<220>

<223> Artificial sequence description: synthetic

oligonucleotide

<400> 427

aaacccaucc uuggaaggcc g 21

<210> 428

<211> 21

<212> RNA

<213> Artificial sequence

<220>

<223> Artificial sequence description: synthetic

oligonucleotide

<400> 428

cucuacaugg aagaacucca c 21

<210> 429

<211> 21

<212> RNA

<213> Artificial sequence

<220>

<223> Artificial sequence description: synthetic

oligonucleotide

<400> 429

uucuaggucc ucuacaugga a 21

<210> 430

<211> 21

<212> DNA

<213> Artificial sequence

<220>

<223> Artificial sequence description: synthetic

oligonucleotide

<220>

<223> Description of the combined DNA/RNA molecule: synthetic

oligonucleotide

<220>

<221> modified base

<222>(8)..(8)

<223> 2'-deoxynucleotide

<400> 430

aaacccaucc uuggaaggcc g 21

<210> 431

<211> 21

<212> DNA

<213> Artificial sequence

<220>

<223> Artificial sequence description: synthetic

oligonucleotide

<220>

<223> Description of the combined DNA/RNA molecule: synthetic

oligonucleotide

<400> 431

aaacccaucc uuggaaggcc g 21

<210> 432

<211> 21

<212> DNA

<213> Artificial sequence

<220>

<223> Artificial sequence description: synthetic

oligonucleotide

<220>

<223> Description of the combined DNA/RNA molecule: synthetic

oligonucleotide

<400> 432

aaacccaucc uuggaaggcc g 21

<210> 433

<211> 21

<212> DNA

<213> Artificial sequence

<220>

<223> Artificial sequence description: synthetic

oligonucleotide

<220>

<223> Description of the combined DNA/RNA molecule: synthetic

oligonucleotide

<220>

<221> modified base

<222>(8)..(8)

<223> 2'-deoxynucleotide

<400> 433

aaacccaucc uuggaaggcc g 21

<210> 434

<211> 21

<212> DNA

<213> Artificial sequence

<220>

<223> Artificial sequence description: synthetic

oligonucleotide

<220>

<223> Description of the combined DNA/RNA molecule: synthetic

oligonucleotide

<220>

<221> modified base

<222>(8)..(8)

<223> 2'-deoxynucleotide

<400> 434

aaacccaucc uuggaaggcc g 21

<210> 435

<211> 21

<212> RNA

<213> Artificial sequence

<220>

<223> Artificial sequence description: synthetic

oligonucleotide

<400> 435

gcucauuguc gagguccuug a 21

<210> 436

<211> 21

<212> RNA

<213> Artificial sequence

<220>

<223> Artificial sequence description: synthetic

oligonucleotide

<400> 436

gccaaaccug uggcuacaag u 21

<210> 437

<211> 21

<212> RNA

<213> Artificial sequence

<220>

<223> Artificial sequence description: synthetic

oligonucleotide

<400> 437

ggaagacgcg acaugucaac a 21

<210> 438

<211> 21

<212> RNA

<213> Artificial sequence

<220>

<223> Artificial sequence description: synthetic

oligonucleotide

<400> 438

gcucauuguc gagguccuug a 21

<210> 439

<211> 21

<212> RNA

<213> Artificial sequence

<220>

<223> Artificial sequence description: synthetic

oligonucleotide

<400> 439

gcucauuguc gagguccuug a 21

<210> 440

<211> 21

<212> RNA

<213> Artificial sequence

<220>

<223> Artificial sequence description: synthetic

oligonucleotide

<400> 440

gcucauuguc gagguccuug a 21

<210> 441

<211> 21

<212> RNA

<213> Artificial sequence

<220>

<223> Artificial sequence description: synthetic

oligonucleotide

<400> 441

gcucauuguc gagguccuug a 21

<210> 442

<211> 21

<212> RNA

<213> Artificial sequence

<220>

<223> Artificial sequence description: synthetic

oligonucleotide

<400> 442

gcucauuguc gagguccuug a 21

<210> 443

<211> 21

<212> RNA

<213> Artificial sequence

<220>

<223> Artificial sequence description: synthetic

oligonucleotide

<400> 443

aaggaccucg acaaugagcu c 21

<210> 444

<211> 21

<212> RNA

<213> Artificial sequence

<220>

<223> Artificial sequence description: synthetic

oligonucleotide

<400> 444

uuguagccac agguuuggca a 21

<210> 445

<211> 21

<212> RNA

<213> Artificial sequence

<220>

<223> Artificial sequence description: synthetic

oligonucleotide

<400> 445

uugacauguc gcgucuuccu g 21

<210> 446

<211> 21

<212> DNA

<213> Artificial sequence

<220>

<223> Artificial sequence description: synthetic

oligonucleotide

<220>

<223> Description of the combined DNA/RNA molecule: synthetic

oligonucleotide

<220>

<221> modified base

<222>(8)..(8)

<223> 2'-deoxynucleotide

<400> 446

aaggaccucg acaaugagcu c 21

<210> 447

<211> 21

<212> DNA

<213> Artificial sequence

<220>

<223> Artificial sequence description: synthetic

oligonucleotide

<220>

<223> Description of the combined DNA/RNA molecule: synthetic

oligonucleotide

<400> 447

aaggaccucg acaaugagcu c 21

<210> 448

<211> 21

<212> DNA

<213> Artificial sequence

<220>

<223> Artificial sequence description: synthetic

oligonucleotide

<220>

<223> Description of the combined DNA/RNA molecule: synthetic

oligonucleotide

<400> 448

aaggaccucg acaaugagcu c 21

<210> 449

<211> 21

<212> DNA

<213> Artificial sequence

<220>

<223> Artificial sequence description: synthetic

oligonucleotide

<220>

<223> Description of the combined DNA/RNA molecule: synthetic

oligonucleotide

<220>

<221> modified base

<222>(8)..(8)

<223> 2'-deoxynucleotide

<400> 449

aaggaccucg acaaugagcu c 21

<210> 450

<211> 21

<212> DNA

<213> Artificial sequence

<220>

<223> Artificial sequence description: synthetic

oligonucleotide

<220>

<223> Description of the combined DNA/RNA molecule: synthetic

oligonucleotide

<220>

<221> modified base

<222>(8)..(8)

<223> 2'-deoxynucleotide

<400> 450

aaggaccucg acaaugagcu c 21

<210> 451

<211> 50

<212> DNA

<213> Artificial sequence

<220>

<223> Artificial sequence description: synthetic

oligonucleotide

<220>

<223> Description of the combined DNA/RNA molecule: synthetic

oligonucleotide

<400> 451

ctcgaggggc aactgaagcc ttttgagacc ctgctgtccc aggcggccgc 50

<210> 452

<211> 50

<212> DNA

<213> Artificial sequence

<220>

<223> Artificial sequence description: synthetic

oligonucleotide

<220>

<223> Description of the combined DNA/RNA molecule: synthetic

oligonucleotide

<400> 452

ctcgagctgg gacagcaggg tctcaaaagg cttcagttgc ccgcggccgc 50

<210> 453

<211> 50

<212> DNA

<213> Artificial sequence

<220>

<223> Artificial sequence description: synthetic

oligonucleotide

<220>

<223> Description of the combined DNA/RNA molecule: synthetic

oligonucleotide

<400> 453

ctcgaggggc aactctacgc aaaacagacc ctgctgtccc aggcggccgc 50

<210> 454

<211> 71

<212> DNA

<213> Artificial sequence

<220>

<223> Artificial sequence description: synthetic

oligonucleotide

<220>

<223> Description of the combined DNA/RNA molecule: synthetic

oligonucleotide

<400> 454

ctcgaggggc aactctacgc aaaacagacc ctgctctacg caaaacagac cctgctgtcc 60

caggcggccg c 71

<210> 455

<211> 92

<212> DNA

<213> Artificial sequence

<220>

<223> Artificial sequence description: synthetic

oligonucleotide

<220>

<223> Description of the combined DNA/RNA molecule: synthetic

oligonucleotide

<400> 455

ctcgaggggc aactctacgc aaaacagacc ctgctctacg caaaacagac cctgctctac 60

gcaaaacaga ccctgctgtc ccaggcggcc gc 92

<210> 456

<211> 113

<212> DNA

<213> Artificial sequence

<220>

<223> Artificial sequence description: synthetic

oligonucleotide

<220>

<223> Description of the combined DNA/RNA molecule: synthetic

oligonucleotide

<400> 456

ctcgaggggc aactctacgc aaaacagacc ctgctctacg caaaacagac cctgctctac 60

gcaaaacaga ccctgctcta cgcaaaacag accctgctgt cccaggcggc cgc 113

<---

Claims (36)

1. An RNAi molecule for downregulating target mRNA expression, where: a) the molecule has a polynucleotide sense strand and a polynucleotide antisense strand; b) each strand of the molecule is 15 to 30 nucleotides in length; c) a contiguous region of 15 to 30 nucleotides of the antisense strand is complementary to the mRNA sequence; d) at least a portion of the sense strand is complementary to at least a portion of the antisense strand and the molecule has a duplex region 15 to 30 nucleotides in length in which one or more nucleotides in the antisense strand at positions 3 to 8 from the 5' end of the region duplex are deoxynucleotides, where the 5' end of the antisense strand corresponds to the 5' end of the duplex region, where the antisense strand has deoxynucleotides at a plurality of positions in the primer region of said antisense strand, where the plurality of positions is one of the following: each of positions 4, 6 and 8 from the 5' end of the antisense strand; each of positions 3, 5 and 7 from the 5' end of the antisense strand; each of positions 1, 3, 5 and 7 from the 5' end of the antisense strand; each of positions 3-8 from the 5' end of the antisense strand; and each of positions 5-8 from the 5' end of the antisense strand. 2. The molecule according to claim 1, wherein the modulation of mRNA expression is the inhibition of expression of a gene selected from protein-coding genes, proto-oncogenes, oncogenes, tumor suppressor genes, and cell signaling genes. 3. A molecule according to claim 1, wherein the mRNA is human mRNA. 4. The molecule according to claim 1, wherein the mRNA is a human mRNA expressing any element or subelement of the human protein family, including SRY, β-globin, RAS, cytosolic GST, mitochondrial GST, MAPEG GST, GST-π, p16, p21 , p53, serum albumin, type VII collagen, complement C3, apolipoprotein B, phenylalanine hydroxylase, factor VIII, huntingtin, retinoblastoma protein RB1, CFTR, titin, atrophin, and dystrophin. 5. A molecule according to claim 1, wherein the molecule has an IC ₅₀ for mRNA knockdown of less than 100 pM. 6. A molecule according to claim 1, wherein the molecule has an IC ₅₀ for mRNA knockdown of less than 50 pM. 7. A molecule according to claim 1, wherein the molecule has an IC ₅₀ for mRNA knockdown of less than 10 pM. 8. A molecule according to claim 1, wherein a single administration of the molecule inhibits mRNA by at least 25% in vivo. 9. A molecule according to claim 1, wherein each strand of the molecule is 18 to 22 nucleotides in length. 10. A molecule according to claim 1, wherein the duplex region is 19 nucleotides in length. 11. A molecule according to claim 1, wherein the polynucleotide sense strand and the polynucleotide antisense strand are joined as a single strand and form a duplex region connected at one end by a loop. 12. Molecule according to claim 1, where the molecule has a blunt end. 13. A molecule according to claim 1, wherein the molecule has one or more 3' sticky ends. 14. A molecule according to claim 1, wherein one or more nucleotides in the duplex region are modified or chemically modified. 15. A molecule according to claim 14, wherein the modified or chemically modified nucleotides are 2'-O-alkyl-substituted nucleotides, 2'-deoxy-2'-fluoro-substituted nucleotides, phosphothioate nucleotides, closed nucleotides, or any combination thereof. 16. A molecule according to claim 1, wherein the base sequence of the antisense strand is SEQ ID NO: 157 and the base sequence of the sense strand is SEQ ID NO: 131. 17. A molecule according to claim 1, wherein the antisense strand is SEQ ID NO: 182 and the sense strand is SEQ ID NO: 156. 18. A molecule according to claim 1, wherein the base sequence of the antisense strand is SEQ ID NO: 195 and the base sequence of the sense strand is SEQ ID NO: 183. 19. A molecule according to claim 1, wherein the antisense strand is SEQ ID NO: 205 and the sense strand is SEQ ID NO: 193. 20. A molecule according to claim 1, wherein the antisense strand is SEQ ID NO: 357 and the sense strand is SEQ ID NO: 343. 21. A molecule according to claim 1, wherein the antisense strand is SEQ ID NO: 356 and the sense strand is SEQ ID NO: 342. 22. Pharmaceutical composition for the treatment of malignant tumors, containing a molecule according to any one of paragraphs. 1-21 and a pharmaceutically acceptable carrier. 23. The pharmaceutical composition of claim 22, wherein the carrier comprises a lipid molecule or liposome. 24. Cell for the expression of the RNAi molecule, containing the molecule according to any one of paragraphs. 1-21. 25. A method of treating malignant tumors in a subject in need thereof by means of gene silencing, wherein the method comprises administering to the subject a composition according to claim 22. 26. The use of a composition according to claim 22 for the treatment of malignant tumors in a subject in need thereof. 27. The use of a composition according to claim 22 for the preparation or manufacture of a medicament for the treatment of cancer in a subject in need thereof.

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