WO2011009082A2 - Compositions comprenant des acides nucléiques modulant les rhbdf1 humains et procédés d’utilisation - Google Patents

Compositions comprenant des acides nucléiques modulant les rhbdf1 humains et procédés d’utilisation Download PDF

Info

Publication number
WO2011009082A2
WO2011009082A2 PCT/US2010/042336 US2010042336W WO2011009082A2 WO 2011009082 A2 WO2011009082 A2 WO 2011009082A2 US 2010042336 W US2010042336 W US 2010042336W WO 2011009082 A2 WO2011009082 A2 WO 2011009082A2
Authority
WO
WIPO (PCT)
Prior art keywords
nucleic acid
seq
acid molecule
acid molecules
cancer
Prior art date
Application number
PCT/US2010/042336
Other languages
English (en)
Other versions
WO2011009082A3 (fr
Inventor
Frank Y. Xie
Original Assignee
Intradigm Corporation
Priority date (The priority date is an assumption and is not a legal conclusion. Google has not performed a legal analysis and makes no representation as to the accuracy of the date listed.)
Filing date
Publication date
Application filed by Intradigm Corporation filed Critical Intradigm Corporation
Publication of WO2011009082A2 publication Critical patent/WO2011009082A2/fr
Publication of WO2011009082A3 publication Critical patent/WO2011009082A3/fr

Links

Classifications

    • C—CHEMISTRY; METALLURGY
    • C12—BIOCHEMISTRY; BEER; SPIRITS; WINE; VINEGAR; MICROBIOLOGY; ENZYMOLOGY; MUTATION OR GENETIC ENGINEERING
    • C12N—MICROORGANISMS OR ENZYMES; COMPOSITIONS THEREOF; PROPAGATING, PRESERVING, OR MAINTAINING MICROORGANISMS; MUTATION OR GENETIC ENGINEERING; CULTURE MEDIA
    • C12N15/00—Mutation or genetic engineering; DNA or RNA concerning genetic engineering, vectors, e.g. plasmids, or their isolation, preparation or purification; Use of hosts therefor
    • C12N15/09—Recombinant DNA-technology
    • C12N15/11—DNA or RNA fragments; Modified forms thereof; Non-coding nucleic acids having a biological activity
    • C12N15/113—Non-coding nucleic acids modulating the expression of genes, e.g. antisense oligonucleotides; Antisense DNA or RNA; Triplex- forming oligonucleotides; Catalytic nucleic acids, e.g. ribozymes; Nucleic acids used in co-suppression or gene silencing
    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61K—PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
    • A61K48/00—Medicinal preparations containing genetic material which is inserted into cells of the living body to treat genetic diseases; Gene therapy
    • C—CHEMISTRY; METALLURGY
    • C12—BIOCHEMISTRY; BEER; SPIRITS; WINE; VINEGAR; MICROBIOLOGY; ENZYMOLOGY; MUTATION OR GENETIC ENGINEERING
    • C12N—MICROORGANISMS OR ENZYMES; COMPOSITIONS THEREOF; PROPAGATING, PRESERVING, OR MAINTAINING MICROORGANISMS; MUTATION OR GENETIC ENGINEERING; CULTURE MEDIA
    • C12N2310/00—Structure or type of the nucleic acid
    • C12N2310/10—Type of nucleic acid
    • C12N2310/14—Type of nucleic acid interfering N.A.
    • C—CHEMISTRY; METALLURGY
    • C12—BIOCHEMISTRY; BEER; SPIRITS; WINE; VINEGAR; MICROBIOLOGY; ENZYMOLOGY; MUTATION OR GENETIC ENGINEERING
    • C12N—MICROORGANISMS OR ENZYMES; COMPOSITIONS THEREOF; PROPAGATING, PRESERVING, OR MAINTAINING MICROORGANISMS; MUTATION OR GENETIC ENGINEERING; CULTURE MEDIA
    • C12N2310/00—Structure or type of the nucleic acid
    • C12N2310/50—Physical structure
    • C12N2310/53—Physical structure partially self-complementary or closed
    • C12N2310/533—Physical structure partially self-complementary or closed having a mismatch or nick in at least one of the strands
    • C—CHEMISTRY; METALLURGY
    • C12—BIOCHEMISTRY; BEER; SPIRITS; WINE; VINEGAR; MICROBIOLOGY; ENZYMOLOGY; MUTATION OR GENETIC ENGINEERING
    • C12N—MICROORGANISMS OR ENZYMES; COMPOSITIONS THEREOF; PROPAGATING, PRESERVING, OR MAINTAINING MICROORGANISMS; MUTATION OR GENETIC ENGINEERING; CULTURE MEDIA
    • C12N2320/00—Applications; Uses
    • C12N2320/50—Methods for regulating/modulating their activity
    • C12N2320/52—Methods for regulating/modulating their activity modulating the physical stability, e.g. GC-content

Definitions

  • the present invention is in the field of molecular biology and medicine and relates to nucleic acid molecules, such as short interfering R]SIA (siRN ⁇ ) molecules, for modulating the expression of Human Rhomboid 5 Homo log- 1 (RHBDFl) as a novel therapeutic approach for the treatment of cancer and other diseases.
  • nucleic acid molecules such as short interfering R]SIA (siRN ⁇ ) molecules, for modulating the expression of Human Rhomboid 5 Homo log- 1 (RHBDFl) as a novel therapeutic approach for the treatment of cancer and other diseases.
  • Rhomboid 5 ⁇ lomolog-1 Homo sapiens Rhomboid 5 ⁇ lomolog-1.
  • hRHBDFl also known as EGFR-RS, EGF-AP, EGFR-RP, Rhomboid Family- 1 is published as GcnBank accession numbers ⁇ K026010, NM 022450. BC014425, AK056708, and M99624.
  • the human RHBDFl gene encodes an 855 amino acid protein (GenBank accession numbers BAB 15318,
  • hRHBDFl is a scven-transmcmbranc spanning protein that resides in the endoplasmic reticulum and Golgi, but not the plasma membrane (Nakagawa et al. 2005 Dev Dyn 233:1315 ⁇ i 33 I), Rhomboid 1 , the Drosophiia homolog of RHBDFi . has been shown to protcolytioally cleave Spitz, the Drosophiia homolog of TGF-alpha, into its soluble, active form.
  • hRHBDFl is not predicted to act as a serine protease because it lacks 2 of the 6 critical residues for protease activity.
  • hRI IBDFl knockdown results m a decrease of soluble TGF-alpha release in response to gastrin-rclcasing peptide (GRi 5 ) stimulation, while liRI IBDFl over-expression results in increased soluble TGF-alpha release.
  • J IxRI IBDF 1 was identified as a gene that was up-regulated in MD ⁇ -MB-435 breast cancer tumors treated with basic Fibroblast Growth Factors.
  • Data mining of NCBl's databases further revealed that hRHBDF 1 inRNA expression was increased m several different cancers, including human breast cancer, stomach cancer, prostate adenocarcinoma, bram glioblastoma, and other tumor types.
  • hRHBDF 1 is expressed at higher levels in stage 1 tumors than in other tumor stages, suggesting that it is involved in the early development of tumors.
  • siKN A duplexes have been used to knockdown hRHBDF 1 in MD A-MB-435 tumors.
  • the siRN A- mediated knockdown resulted m growth inhibition of the tumors.
  • Knockdown of hRITRDF! in MDA-MB-435 cells in culture resulted in an increase in apoptosis.
  • Knockdown of hRHBDF 1 in 1483 cells inhibited GRP-induced EGFR phosphorylation, as well as phosphorylation of downstream targets p44/42 MAPK and AKT.
  • RNA interference (RKAi) technology is an effective means for reducing the expression of specific gene products and may be uniquely useful in a number of therapeutic, diagnostic, and research applications for the modulation of expression of hRI IBDF 1.
  • the present invention provides compositions and methods for modulating expression of hRI IBDFl using RNAi technology.
  • compositions and methods for modulating the expression of hRI IBDFI as a therapeutic approach for the treatment of cancer and other diseases.
  • the present invention provides this and other advantages.
  • One aspect of the present invention provides a nucleic acid molecule that down regulates expression of a human Rhomboid 5 I Iom ⁇ log-1 (hRIIBDFl ) gene, wherein the nucleic acid molecule comprises a nucleotide sequence that targets ShRHBDFI mRNA. wherein the nucleic acid molecule comprises a nucleotide sequence that targets any one of the polynucleotide sequences set forth in SEQ) ID NOs: 1 -47 and 638. In some embodiments, the nucleic acid molecule consists of a nucleotide sequence that targets any one of the polynucleotide sequences set forth m SEQ [D NOs: 1-47 and 638.
  • the nucleic acid molecule is single-stranded. In some embodiments, the nucleic acid is double-stranded. In a particular embodiment, the nucleic acid is an siRN ⁇ molecule. In more particular embodiments, the nucleic acid molecule comprises any one of nucleotide sequences provided in SEQ ID NOs: 1 73-269 and 454-637. In some embodiments, the nucleic acid molecule is a double-stranded nucleic acid molecule comprising any pair of the sequences provided in l ablcs 1-8 (SEQ ID NQs: 1-637). In certain embodiments, the nucleic acid molecule down regulates expression of an hRHBDFl gene via RJS ⁇ interference (RJS ⁇ f).
  • RJS ⁇ f RJS ⁇ interference
  • the nucleic acid molecule is 15-50. 15-30. 16, 17, 18, 19, 20, 21, 22. 25, 24, or 25 nucleotides long. In some embodiments, the nucleic acid molecule is blunt-ended, while m other embodiments the nucleic acid molecule comprises one or two single-strand overhangs. In various embodiments, the nucleic acid molecule comprises both RNA and DNA. T ⁇ some embodiments, the nucleic acid molecule comprises modified bases, such as
  • the nucleic acid molecule decreases hRTIBDFl expression by at least j 0-100%. at least 25- 100%. at least 50-100%, at least 75-100%, or at least 90-100%, such as by at least 50%. 55%, 60%. 65%, 70%, 75%, 80%, 85%, 90%, 95%, 96%, 97%, 98 1 M), 99 1 M,, or 100%.
  • the invention provides tor an expression vector encoding a nucleic acid molecule that targets a polynucleotide comprising the nucleotide sequence set forth m any one of SEQ ID NOs: 1-47 and 63 S.
  • the present invention provides a composition comprising any one or more of the nucleic acid molecules of the invention.
  • the nucleic acid molecules of the invention are any one or more of the nucleic acid molecules of the invention.
  • composition may comprise 1 , 2, 3, 4, 5, 6, 7, 8, 9, 10, 1 1 , 12, 1 3, 14, 15, 16, 17, 18, I Q , 20, 21, 22, 23, 24, 25, 26, 27, 28, 29, 30, 35, 40, 45, 50, 55, 60, 65, 70, 75, SO, 85, 90, 95, 100 or more different nucleic acid molecules of the invention, wherein each different nucleic acid molecule targets a polynucleotide comprising the nucleotide sequence set forth in any one of SEQ I D NOs: 1 -47 and 638.
  • the composition comprises a nucleic acid of the invention and a pharmaceutically acceptable carrier.
  • the pharmaceutically acceptable carrier is a nucleic acid delivery vehicle.
  • the nucleic acid delivery vehicle further comprises a targeting moiety, a cationic copolymer, a hydrophilic polymer, or any combination of such components.
  • the cationic copolymer is a histidinc-lysine copolymer.
  • the composition comprises an additional therapeutic agent.
  • the present invention provides a method for reducing tumor growth in a human subject having or suspected of having an hRHBDFl expressing tumor, comprising administering to the subject a composition comprising a nucleic acid molecule of the invention, i.e. a nucleic acid molecule comprising any one of SEQ ID NOs: 173-269 and 454-637. thereby treating the cancer or preventing recurrence thereof.
  • the nucleic acid molecule is a double- stranded nucleic acid molecule comprising any pair of sequences provided in Tables 1-8 (SEQ ID NOs; 1-637).
  • the RHBDF-I expressing cancer may be selected from the group consisting of breast cancer, lung cancer, prostate cancer, colorectal cancer, brain cancer, esophageal cancer, stomach cancer, bladder cancer, pancreatic cancer, cervical cancer, head and neck cancer, kidney cancer, endometrial cancer, ovarian cancer, meningioma, melanoma, lymphoma, and glioblastoma.
  • the cancer may be a primary tumor, a benign tumor, a malignancy, a metastatic tumor or a recurrent tumor.
  • the present invention provides a method for reducing the synthesis or expression of hRHBDFl comprising delivering to a cell expressing hRHBDF l one or snore nucleic acid molecules, wherein the nucleic acid molecules target a polynucleotide comprising the nucleotide sequence set forth in any one of SEQ ID NOs; 1-47 and 638.
  • the nucleic acid molecules comprise a sequence as set forth in SEQ ID NOs: 173-269 and 454-637.
  • the nucleic acid molecules consist of a sequence as set forth in SEQ ID NOs: 1 73-269 and 454-637.
  • the nucleic acid molecules comprise a
  • double-stranded nucleic acid comprising any pair of sequences provided in Tables 1-8 (SEQ ID NOs: 1-637).
  • the present invention provides a method for reducing translation of an mRN ⁇ encoding hRHBDFl in a cell comprising delivering to the cell expressing hRIIBDFl one or more nucleic acid molecules, wherein the nucleic acid molecules target a polynucleotide comprising the nucleotide sequence set forth in any one of SEQ ID NOs: 1-47 and 638.
  • the nucleic acid molecules comprise a sequence as set forth in SEQ ID NOs: 173-269 and 454-637.
  • the nucleic acid molecules consist of a sequence as set forth in SEQ ) [D NOs: 173-269 and 454-637.
  • the nucleic acid molecules comprise a double-stranded nucleic acid comprising any pair of sequences provided in Tables 1-8 (SHQ ID NOs: 1-637).
  • the present invention provides a method for reducing the amount of an mRNA encoding hRHBDFl in a cell comprising delivering to the cell expressing hRHBDFl one or more nucleic acid molecules, wherein the nucleic acid molecules target a polynucleotide comprising the nucleotide sequence set forth in any one of S£Q ID NOs; 1-47 and 368.
  • the nucleic acid molecules comprise a sequence as set forth in SEQ ID NOs: 173-269 and 454-637.
  • the nucleic acid molecules consist of a sequence as set forth m SEQ ID NOs: 173-269 and 454-637.
  • the nucleic acid molecules comprise a double-stranded nucleic acid comprising any pair of sequences provided in Table 1-8 (S£Q ID NOs; 1-637).
  • the present invention provides a method for reducing the amount of hRHBDFl protein in a cell comprising delivering to the cell expressing hRHBDFl one or more nucleic acid molecules, wherein the nucleic acid molecules target a polynucleotide comprising the nucleotide sequence set forth in any one of SEQ LD NOs: 1 -47 and 638.
  • the nucleic acid molecules comprise a sequence as set forth in SEQ TD NOs: 173-269 and 454-637.
  • the nucleic acid molecules consist of a sequence as set forth in SEQ ID NOs: 173-269 and 454-637.
  • the nucleic acid molecules comprise a
  • double-stranded nucleic acid comprising any pair of sequences provided in Tables 1 -8 (SEQ ID NOs: 1-637).
  • the present invention provides a method for reducing hRHBDFl activity in a cell comprising delivering to the cell expressing hRHBDFl one or more nucleic acid molecules, wherein the nucleic acid molecules target a
  • nucleic acid molecules comprising the nucleotide sequence set forth in any one of SEQ ID NOs: 1 -47 and 638.
  • nucleic acid molecules comprise a sequence as set forth in SEQ) ID NOs: 173-269 and 454-637, In some embodiments, the nucleic acid molecules consist of a sequence as set forth in SEQ ID NOs: 173-269 and 454-637. In additional embodiments, the nucleic acid molecules comprise a
  • double-stranded nucleic acid comprising any pair of sequences provided in Tables 1 -8 (SEQ lD NOs: 1-637).
  • the present invention provides a method for reducing the cell growth or proliferation of a cell expressing hRHBDFl comprising delivering to the cell expressing hRIIBDFl one or more nucleic acid molecules, wherein the nucleic acid molecules target a polynucleotide comprising the nucleotide sequence set forth in any one of SEQ ID NOs: 1-47 and 638.
  • the nucleic acid molecules comprise a sequence as set forth in SEQ ID NQs: 173-269 and 454-637.
  • the nucleic acid molecules consist of a sequence as set forth in SEQ) [D NOs: 173-269 and 454-637.
  • the nucleic acid molecules comprise, consist essentially of or consist of a double-stranded nucleic acid comprising or consisting of any pair of sequences provided in Tables 1-8 (SEQ [D TNOS: 1-637),
  • the present invention provides a method for treating or preventing cardiovascular disease in a subject comprising the step of administering to the subject one or more nucleic acid molecules, wherein the nucleic acid molecules target a polynucleotide comprising the nucleotide sequence set forth in any one of SEQ ID NOs: 1-47 and 638.
  • the nucleic acid molecules comprise a sequence as set forth in SEQ TD NOs: 173-269 and 454-637.
  • the nucleic acid molecules consist of a sequence as set forth in SEQ TD NOs: 173-269 and 454-637.
  • the nucleic acid molecules comprise a
  • double-stranded nucleic acid comprising any pair of sequences provided in Tables 1 -8 (SEQ TD NOs: 1-637).
  • nucleic acid molecule of paragraph I wherein the nucleic acid is a short interfering RNA (siRNA) molecule.
  • nucleic acid molecule of paragraph I or the siRNA molecule of paragraph 2 wherein the molecule comprises the nucleotide sequence of any one of SEQ ID NOs: S 73-269 and 454-637.
  • nucleic acid molecule of paragraph 1 or the siRN A molecule of paragraph 2 wherein the molecule comprises the nucleotide sequences of any pair set forth in Tables 1-8 (SEQ [D MOs: 1-637),
  • nucleic acid molecule of any of paragraphs 1-6 wherein at least one wobble basepair is formed between the 3' portion of the antisense strand (SEQ ID NO: 173-269 and 454-637) and the target polynucleotide.
  • nucleic acid molecule of paragraph 7 wherein at least two wobble basepairs are formed, wherein at least one wobble basepair is formed between the 5' portion of the antisense strand (SEQ ID NO: 173-269 and 454-637) and the target polynucleotide and at least one wobble basepair is formed between the 3' portion of the antisense strand (SEQ ID NO: 173-269 and 454-637) and the target polynucleotide. 12.
  • RNAi RNA interference interference
  • nucleic acid molecule of any one of paragraphs 4-15 wherein the nucleic acid molecule is blunt-ended.
  • nucleic acid molecule comprises one or two single-strand overhangs.
  • nucleic acid molecule of paragraph 19 wherein the modified bases are 2'-O-m ⁇ thylated nucleotides 21.
  • An expression vector encoding a nucleic acid molecule that, targets a polynucleotide comprising the nucleotide sequence set forth in any one of SEQ TD NOs: 1-47 and 638.
  • composition comprising the nucleic acid molecule, the siRNA ! 5 molecule, or the vector of any one of paragraphs 1-26.
  • composition comprising at least two different nucleic acid molecules, siRNA molecules, or vectors of any one of paragraphs 1-26.
  • composition paragraph 28 wherein each different molecule targets a different polynucleotide comprising the nucleotide sequence set forth in any one 0 of SEQ ID NOs; 1-47 and 638.
  • composition of any one of paragraphs 27-29 further comprising a targeting moiety.
  • composition of any one of paragraphs 27-30 further comprising a cationic copolymer. 5
  • the composition of any one of paragraphs 27-32 further comprising an additional therapeutic agent.
  • ⁇ method for treating or preventing a cancer in a subject with an RIIBDFI -expressing cancer and Slaving or at risk for having the cancer comprising administering to the subject the nucleic acid molecule of any one of paragraphs 1-25, the expression vector of paragraph 26 or the composition of any one of paragraphs 27-33, thereby treating or preventing the cancer.
  • the cancer is selected from the group consisting of breast cancer, lung cancer, prostate cancer, colorectal cancer, brain cancer, esophageal cancer, stomach cancer, bladder cancer, pancreatic cancer, cervical cancer, head and neck cancer, kidney cancer, endometrial cancer, ovarian cancer, meningioma, melanoma, lymphoma, and glioblastoma.
  • a method for reducing the synthesis or expression of hRf IBDFl in a ceil comprising introducing into the ceil one or more nucleic acid molecules, wherein the one or more nucleic acid molecules target a polynucleotide comprising the nucleotide sequence set forth in any one of SEQ ID NOs: 1-47 and 638.
  • a method for reducing the translation of mRNA encoding hRHBDFl in a ceil comprising introducing into the cell one or more nucleic acid molecules, wherein the one or more nucleic acid molecules target a polynucleotide comprising the nucleotide sequence set forth in any one of SEQ ID NOs: 1-47 and 638.
  • a method for reducing the amount of mRNA encoding hRI IBDFl in a cell comprising introducing into the cell one or more nucleic acid molecules.
  • a method for reducing the amount of hRI I BDFl protein in a cell comprising introducing into the ceil one or more nucleic acid molecules, wherein the one or more nucleic acid molecules target a polynucleotide comprising the nucleotide sequence set forth in any one of SEQ ID NOs: 1 -47 and 638.
  • a method for reducing hRIIBDFl activity in a cell comprising introducing into the cell one or more nucleic acid molecules, wherein the one or more nucleic acid molecules target a polynucleotide comprising the nucleotide sequence set forth in any one of SEQ ID NOs: 1-47 and 638, 41.
  • a method for reducing the cell growth or proliferation of a cell expressing liRHBDFl comprising introducing into the cell one or more nucleic acid molecules, wherein the one or more nucleic acid molecules target a polynucleotide comprising the nucleotide sequence set forth in any one of SEQ ID NOs: 1-47 and 638.
  • a method for treating or preventing a cardiovascular disease in a subject comprising the step of administering one or more nucleic acid molecules to the subject, wherein the one or more nucleic acid molecules target a polynucleotide comprising the nucleotide sequence set forth in any one of SEQ ID NOs: 1-47 and 638.
  • nucleic acid molecules comprise at least one of the nucleotide sequences set forth in SEQ ID NOs: 173-269 and 454-637.
  • nucleic acid molecules consists of at least one of the nucleotide sequences set forth in SEQ ID NOs: 173-269 and 454-637.
  • nucleic acid molecule of paragraph 3 wherein the nucleic acid molecule consists of a sequence as set forth in any one of SEQ ID NOs: 173-269 and 454-637.
  • Figure I depicts the rnRNA sequence for human RIIBDFl (GenBank Accession No. NM 022450.3, SEQ ID NO: 638). Nucleotides 1-138 are in the 5 '-untranslated region, and the protein coding region begins at nucleotide 139.
  • the present invention relates to nucleic acid molecules for modulating the expression of hRHBDFl .
  • the nucleic acid molecules are single stranded.
  • the nucleic acid molecules are double stranded, Tn various embodiments the nucleic acid is ribonucleic acid (RNA) or a mixture of RNA and deoxyribonucleic acid (DNA).
  • the nucleic acid molecules of the present invention such as si RNA, reduce expression of hRHBDFl.
  • the nucleic acid molecule is a DNA molecule encoding an RNA molecule, which reduces expression of hRHBDFl .
  • RNA sequence e.g., the sequence comprises uracil
  • RNA sequence e.g., the sequence comprises uracil
  • recitation of an RNA sequence also contemplates the corresponding DN ⁇ sequence.
  • the present invention relates to compounds, compositions, and methods for the study, diagnosis, and treatment of traits, diseases and conditions that respond to the modulation of hRHBDFl gene expression and/or activity.
  • the present invention is also directed to compounds, compositions, and methods relating to traits, diseases and conditions that respond to the modulation of expression and/or activity of genes involved in hRHBDFl pathways or other cellular processes that mediate the maintenance or development of such traits, diseases and conditions.
  • the invention relates to nucleic acid molecules including small nucleic acid molecules, such as short interfering nucleic acid (siNA), short interfering RNA (siRNA), double-stranded RNA (dsRNA).
  • micro-RNA miRNA
  • short hairpin RNA CsIiRNA RNA interference
  • the present invention also relates to small nucleic acid molecules, such as 8iNA, siRN A, and others that can inhibit the function of endogenous RNA molecules, such as endogenous micro-RN ⁇ (miRN ⁇ ) (e.g., miRNA inhibitors) or endogenous short interfering RN ⁇ (siRNA), (e.g., siRN ⁇ inhibitors) or that can inhibit the function of RISC (e.g.
  • RISC inhibitors to modulate hRHBDFl gene expression by interfering with the regulatory function of such endogenous RTsIAs or proteins associated with such endogenous R]SlAs (e.g. , RISC), including cocktails of such small nucleic acid molecules and nanoparticle formulations of such small nucleic acid molecules.
  • Such small nucleic acid molecules may be useful, for example, in providing compositions to inhibit, reduce or prevent recurrence of breast, lung, prostate, stomach, colorectal, brain, esophageal, bladder, pancreatic, cervical, head and neck, and ovarian cancer, melanoma, lymphoma, glioma, multidrug resistant cancers, and any other cancerous disease and/or other disease states, conditions, or traits associated with hRHBDFl gene expression or activity in a subject or organism.
  • inhibit or “down-regulate” it is meant that, the expression of the hRHBDFi gene, or level of rtiRNA encoding an hRHBDFl protein, levels of hRHBDFl protein, or activity of hRHBDFl is reduced below that observed in the absence of the nucleic acid molecules of the invention.
  • inhibition or down-regulation with the nucleic acid molecules of the invention is preferably below that, level observed in the presence of, for example, a nucleic acid with scrambled sequence or with mismatches.
  • inhibition or down-regulation of hRHBDF l with the nucleic acid molecule of the instant invention is greater in the presence of the nucleic acid molecule than in its absence.
  • the nucleic acid molecules of the invention can decrease the amount of soluble TGF-alpha released in response to GRP-stimulations. In some embodiments, the nucleic acid molecules of the invention can increase apoptosis. In some embodiments, the nucleic acid molecules of the invention can decrease
  • the nucleic acid molecules of the invention can decrease the phosphorylation of EGFR downstream targets. In some embodiments, the nucleic acid molecules of the invention can decrease phosphorylation of p44/42 MAPK or ⁇ KT.
  • module is meant that the expression of the hRHBDFl gene, or level of mRNA encoding an hRHBDFl protein, levels of hRHBDFl protein, or activity of hRHBDFl is greater than or less than that observed in the absence of the nucleic acid molecules of the invention.
  • double stranded RN ⁇ ⁇ r "dsRNA” is meant a double stranded RNA that comprises a strand that will hybridize to a target mRNA sequence and that is capable of activating cellular enzymes that degrade the messenger RN ⁇ .
  • dsRN ⁇ s are referred to as short interfering RN ⁇ (siRNA) and can be used to inhibit gene expression (see for example tlbashir et aL, 2001, Nature, 411, 494-498; and Bass, 2001 , Nature, 411 ,
  • double stranded RKA or "dsRNA” as used herein also refers to a double stranded RN ⁇ molecule capable of mediating RNA interference "RN ⁇ i”, including short interfering RNA "siRJS ' A” (see for example Bass, 2001, Nature, 411, 428-429; Eibashir et a!.. 2001, Nature, 41 1, 494-498: and Kreutzer et aL, International PCT Publication No. WO 00/44895; Zemicka-Goetz et aL. International PCT Publication No. WO 01/36646; Fire, International PCT Publication No.
  • WO 99/32619 Piaetinck et aL International PCT Publication No. WO 00/01846; Me Uo and Fire, International PCT Publication No. WO 01/29058; Dcschamps-Dcpaillette, International PCT Publication No. WO 99/07409; and Li et al, International PCT Publication No. WO 00/44914).
  • the dsRNA may be a 25-n ⁇ cr, The dsRNA may be blunt-ended or may comprise one or two smglc-strandcd overhangs.
  • nucleic acid that encodes an mRNA
  • nucleic acid sequences include but are not limited to structural genes encoding a polypeptide.
  • a nucleic acid that ''targets' a nucleic acid as described herein that comprises a nucleotide sequence that is complementary to and/or is capable of specifically hybridizing to a target sequence and thereby modulates the expression of a nucleic acid comprising the target sequence, including the level of mRNA transcribed, the level of a protein encoded by the mRNA, and/or activity ⁇ f the mRNA or protein.
  • “Complementarity” refers to the ability of a nucleic acid to form hydrogen bond(s) with another RNA sequence by either traditional Watson-Crick or other non-traditional types of basepairmg.
  • the binding free energy for a nucleic acid molecule with its target or complementary sequence is sufficient to allow the relevant function of the nucleic acid to proceed, e.g.. enzymatic nucleic acid cleavage, antisense or triple helix inhibition.
  • a percent complementarity indicates the percentage of residues in a nucleic acid molecule which can form hydrogen bonds ⁇ e.g., Watson-Crick basepairing) with a second nucleic acid sequence (e.g. , 5, 6, 7. 8, 9.
  • Perfectly complementary means that all the contiguous residues of a nucleic acid sequence will hydrogen bond with the same number of contiguous residues in a second nucleic acid sequence
  • RNA is meant a molecule comprising at least one ribonucleotide residue.
  • 'ribonucleotide or “2'-OH” is meant a nucleotide with a hydroxyl group at the 2" position of a ⁇ -D-ribo-furanose moiety.
  • RNA interference or "RNAi” is meant a biological process of inhibiting or down regulating gene expression in a cell as is generally known in the art and which is mediated by short interfering nucleic acid molecules (see for example Zamore and Haley. 2005, Science, 309, 1519-1524; Vaughn and Martienssen, 2005, Science, 309,
  • RNAi is meant to be equivalent to other terms used to describe sequence specific RNA interference, such as post transcriptional gene silencing, translational inhibition, transcriptional inhibition, or epigenetics.
  • siRNA molecules of the invention can be used to
  • epigenetic modulation of gene expression by siRNA molecules of the invention can result from siRNA mediated modification of chromatin structure or methylation patterns to alter gene expression (see, for example. Verd ⁇ l et a!. , 2004, Science, 303, 672-676; Pal-Bhadra et al , 2004, Science, 303, 669-672; ⁇ llshire, 2002, Science, 297, 1818-1819; Volpe et a!. , 2002, Science. 297, 1833-1837; J ⁇ nuwem, 2002, Science, 297, 221 5-2218; and Hal! et a!., 2002, Science, 297, 2232-2237).
  • modulation of gene expression by siR]M ⁇ molecules of the invention can result from siRNA mediated cleavage of RN ⁇ (either coding or non-coding RKA) via RISC, or alternately, translational inhibition as is known in the art.
  • modulation of gene expression by siRNA molecules of the invention can result from transcriptional inhibition (see for example Janowski et al , 2005. Nature Chemical Biology, 1, 216-222).
  • the nucleic acid modulators comprise sequences that are complementary to an hRHBDFl sequence, including variants thereof that have altered expression and/or activity, particularly variants associated with disease.
  • Variants of hRHBDF l include sequences having 70%, 75%, 80%, 85%, 90%, 91%. 92%. 93%, 94%, 95%, 96%, 97%, 98%, 99% or higher sequence identity to the wild type hRHBDFl (SEQ ID NO: 638), wherein such hRHBDFl variants may demonstrate altered (increased or decreased) hRHBDFl activity.
  • hRHBDFl sequences are available in any of a variety of public sequence databases including
  • the nucleic acid modulators (e g. , siRNA) of the invention comprise sequences complementary to the specific hRITRDFS target sequences provided in SEQ TD NOs: 1-47 and 638 (see Table 1 and Figure 1). Examples of such nucleic acid molecules also are shown m the Examples and provided m SEQ TD NOs: 173-269 and 454-637 (see Tables S -8). Tn one embodiment, the nucleic acid modulators of the invention consist of sequences complementary to the specific hRH BDFl target sequences provided in SEQ ID NOs: 1-47 and 638.
  • nucleic acid molecules of the invention consist of a sequence as set forth in SFQ ID NOs: 173-269 arid 454-637.
  • nucleic acid molecules of the invention are double-stranded and each double-stranded nucleic acid molecule consists of one pair nucleotide sequences set forth in Tables 1 -8 (SFQ ID NOs: 1 -637).
  • nucleic acid molecules of the invention target nucleic acid molecules comprising or consisting of one of the sequences provided in SEQ ID NOs: 1 -47 and 638.
  • '"vectors' * is meant any nucleic acid- and/or viral-based technique used to deliver a desired nucleic acid.
  • a subject is meant an organism which is a recipient of the nucleic acid molecules of the invention.
  • Subject also refers to an organism to which the nucleic acid molecules of the invention can be administered.
  • a subject is a mamma] or mammalian cells.
  • a subject is a human or human cell.
  • Nucleic acids can be synthesized using protocols known in the art as described in Camthers et al,, 1992, Methods in Enzymology 211, 3 19, Thompson el al,,
  • nucleic acids makes use of common nucleic acid protecting and coupling groups, such as dimethoxytrityl at the 5 '-end, and phosphoramidites at the 3 '-end.
  • nucleic acid protecting and coupling groups such as dimethoxytrityl at the 5 '-end, and phosphoramidites at the 3 '-end.
  • small scale syntheses arc conducted on a 394 Applied Biosystems, Inc.
  • oligonucleotide synthesis reagents for the 394 Applied Biosystems, Inc. synthesizer include; detritylation solution is 3% TCA in methylene chloride (ABI); capping is performed with 16% N-methylimidazoIe in TI IF ( ABI) and 10% acetic anhydride/10% 2,6-lutidine in TIIF (ABI); arid oxidation solution is 16.9 mM h, 49 mM pyridine, 9% water in TlTF. Burdick & Jackson Synthesis Grade acetom ' trile is used directly from the reagent bottle.
  • S-Ethyltetrazol ⁇ solution (0.25 M in acetonitrile) is made up from the solid obtained from American International Chemical, Inc.
  • Beaiicage reagent (3H-l,2-Benzodithiol-3-one 1,1 -dioxide, 0.05 M in acetonitrile) is used.
  • nucleotide is meant a heterocyclic nitrogenous base in N-glycosidic linkage with a phosphorylated sugar.
  • Nucleotides are recognized in the art to include natural bases (standard), and modified bases well known in the art. Such bases are generally located at the 1 " position of a nucleotide sugar moiety.
  • Nucleotides generally comprise a base, sugar and a phosphate group. The nucleotides can be unmodified or modified at the sugar, phosphate and/or base moiety, (also referred to interchangeably as nucleotide analogs, modified nucleotides, nan- natural nucleotides, non-standard nucleotides and other; see for example.
  • nucleic acid bases There are several examples of modified nucleic acid bases known in the art as summarized by Limbach et ai., 1 ⁇ 94, Nucleic Acids Res. 22, 2183.
  • Exemplary chemically modified and other natural nucleic acid bases that can be introduced into nucleic acids include, for example, inosinc. purine, pyridm-4-one, pyridin-2-one, phenyl, pscudouracil.
  • 6-a/apyrimidines or 6-alkylpyrimidincs e.g ⁇ -methyluridine
  • propyne quesosine
  • 2 -thio uridine 4-thiouridine
  • wybutosine wybutoxosinc.
  • 4-acetyltidirse
  • 5-carboxymethylaminometliyluridmc beta-D-galactosylqucosine. 1-methyladenosine, ! -methylinosine, 2,2-din ⁇ ethylgua ⁇ iosine, 3-i ⁇ ethylcytidine, 2-methyladcnosine.
  • nucleotide bases other than adenine, guanine, cytosine and uracil at 1' position or their equivalents; such bases can be used at any position, for example, within the catalytic core of an enzymatic nucleic acid molecule and/or in the substrate-binding regions of the nucleic acid molecule.
  • nucleoside is meant a heterocyclic nitrogenous base in N-glycosidic linkage with a sugar. Nucleosides are recognized in the art to include natural bases
  • Nucleosides generally comprise a base and sugar group.
  • the nucleosides can be unmodified or modified at the sugar, and/ or base moiety, (also referred to interchangeably as nucleoside analogs, modified nucleosides, non-natural nucleosides, non-standard nucleosides and othei; see for example, Usman and McSwiggcn, supra; Eckstein et aL, International PCT Publication No, WO 92 -'07065; Usman et aL International PCT Publication Mo.
  • nucleic acid bases There arc several examples of modified nucleic acid bases known m the art as summarized by Limbach et aL, 1994, Nucleic ⁇ cids Res. 22, 2183. Exemplary chemically modified and other natural nucleic acid bases that can be introduced into nucleic acids include, inosinc. purine, pyridm-4-onc.
  • 5-haiouridme e g . 5-bromouridiiic
  • 6-azapyrimidincs e g . 6-alkylpyrimidmcs
  • propyne e g . 2-thiou ⁇ dine
  • 4-thiouridinc e g . 4-thiouridinc.
  • nucleoside bases other than adenine, guanine, cytosme and uracil at 1' position or their equivalents; such bases can be used at any position, for example, within the catalytic core of an en/ymatic nucleic acid molecule and/or in the substrate-binding regions of the nucleic acid molecule.
  • the term "antisense strand" of an siR ⁇ I ⁇ or R ⁇ i agent refers to a strand that is substantially compjementaiy to a section of about 10-50 nucleotides, e.g., about 15-30, 16-25, 18-23 or 19-22 nucleotides of a hRHBDFl mRN ⁇ .
  • the antisense strand or "first strand” has sequence sufficiently complementary to the desired target mRN ⁇ sequence to direct target-specific RN ⁇ mteilerence ( RK ⁇ i;. e.g..
  • 'sense strand * ' or "second strand” of an siRN ⁇ or RN ⁇ i agent refers to a strand that is substantially complementary to the antisense strand or first strand.
  • guide strand refers to a strand of an RNAi agent, e.g., an antisense strand of an siRNA duplex, that enters into the RISC complex and directs cleavage of the target inRN ⁇ .
  • the term "5' portion * ' of a nucleic acid molecule refers the half of the molecule closer to the 5' terminal nucleotide. In a 25 basepair molecule, for example, the 5' portion would encompass the 13-5' terminal nucleotides.
  • the term "3' portion" of a nucleic acid molecule refers the half of the molecule closer to the 3' terminal nucleotide. In a 25 basepair molecule, for example, the 3' portion would encompass the 13-3' terminal nucleotides.
  • a nucleotide that is equidistant from the 5' and 3' terminal nucleotides ⁇ e.g. the 13th nucleotide of a 25 basepair nucleic acid molecule) will be considered to be in both the 5' portion and the 3' portion of the nucleic acid molecule.
  • the efficacy and specificity of gene/protein silencing by the siRNA nucleic acids of the present, invention may be enhanced using the methods described in US Patent Application Publications 2005/0186586, 2005/0181382,
  • double-stranded embodiments may comprise at least one blunt end and may comprise two blunt, ends.
  • the duplex comprises at least one overhang and may comprise two overhangs.
  • modifications can be made to the siRNA molecules of the invention to promote entry of a desired strand of an siRNA duplex into a RISC complex. This is achieved by enhancing the asymmetry of the siRNA duplex, such that entry of the desired strand is promoted. In this regard, the asymmetry is enhanced by lessening the basepair strength between the 5' portion of the desired strand and the 3' portion of a complementary strand of the duplex compared to the basepair strength between the 3' portion of the desired strand and the 5' portion of the complementary strand.
  • the base-pair strength is less due to fewer G: C bas ⁇ pairs between the 5' portion of the first or antisense strand and the 3' portion of the second or sense strand than between the 3' portion of the first or antisense strand and the 5' portion of the second or sense strand.
  • the basepair strength is less due to at least one mismatched basepair between the 5' portion of the first or antisense strand and the 3' portion of the second or sense strand.
  • the mismatched basepairs include but are not limited to G: ⁇ , C: ⁇ , C:U, G:G, ⁇ : ⁇ , C:C, Ii: Ii, C:T, and U:T.
  • the one or more mismatch basepairs are within 13, 12, 11. 10, 9, 8, 7, 6, 5, 4, 3, 2, or 1 nucleotides of the 5' terminal nucleotide of the anusense strand and the 3' terminal nucleotide of the sense strand.
  • a mismatch basepair occurs between the 5 " terminal nucleotide of the antisense strand and the 3' terminal nucleotide of the sense strand of a duplex.
  • the basepair strength is less due to at least one wobble basepair between the 5' portion of the first or antisense strand and the 3 " portion of the second or sense strand.
  • the wobble basepair may be G;U, G:T, LU, I: ⁇ , or 1;C.
  • the one or more wobble basepairs arc withm 13, 12, 11, 10. 9. 8. 7, 6, 5, 4, 3, 2, or 1 nucleotides of the 5 ' terminal nucleotide of the antisense strand and the 3 ' terminal nucleotide of the sense strand.
  • a wobble basepair occurs between the 5 " terminal nucleotide of the antisense strand and the 3 " terminal nucleotide of the sense strand of a duplex.
  • the basepair strength is less due to: (a) at least one mismatched basepair between the 5 * portion of the first or antisense strand and the 3' portion of the second or sense strand; and (b) at least one wobble basepair bet ween the 5 ' portion of the first or antisense strand and the 3' portion of the second or sense strand.
  • the mismatched basepair may be selected from the group consisting of G: A, C: A, C;U, G:G, A;A. C:C and U:U
  • the mismatched basepair is selected from the group consisting of G: A, C: ⁇ , C:T, G:G, A:A, C:C and U:T.
  • the wobble basepair is G:U, G:T, T;U. T:A. or LC.
  • the one or more mismatch basepairs and the one or snore wobble basepairs are within S 3, 12, 1 1, 10, 9, 8, 7, 6, 5, 4, 3, 2, or 1 nucleotides of the 5' terminal nucleotide of the antisense strand and the 3' terminal nucleotide of the sense strand.
  • a wobble basepair occurs between the 5' terminal nucleotide of the antisense strand and the 3 ' terminal nucleotide of the sense strand of a duplex.
  • a mismatch basepair occurs between the 5' terminal nucleotide of the antisense strand and the 3 ' terminal nucleotide of the sense strand of a duplex.
  • the basepair strength is reduced due to at least one basepair comprising a rare nucleotide such as inosine, 1 -methyl inosine, pseudouridine, 5,6-dihydrouridinc. ⁇ bothymidine, 2N-methylguanosme and 2,2N,N-dimcthylguanosine; or a modified nucleotide, such as 2-ammo-G, 2-amino-A. 2,6-diamino-G. and
  • the efficacy of the siRNA molecule to silence a target gene is enhanced by enhancing the recycling of the guide strand siRN ⁇ from the RISC. This is achieved by lessening the basepair strength between the guide strand and its target mRNA by introducing wobble base-pairing (e g., G:U, 1:U, 1:A. and 1:C) in the 5' or 3' portion of the guide strand of siRN ⁇ ,
  • wobble base-pairing e g., G:U, 1:U, 1:A. and 1:C
  • guide strand recycle is enhanced due to at least one wobble basepair between the guide strand and the target mRN ⁇ .
  • at least one wobble basepair is in the 5' portion of the guide strand.
  • the at lease one wobble basepair is within 13, 12, 11, 10. 9, 8, 7, 6, 5, 4 , 3, 2, or 1 nucleotides of the 5 ' terminal nucleotide of the guide strand.
  • a wobble basepair occurs between the 5' terminal nucleotide of the guide strand and the target mRNA.
  • at least one wobble basepair is in the 3' portion of the guide strand.
  • the at least one wobble basepair is within 13. 12, 11, 10, 9, 8, 7, 6, 5, 4. 3. 2. or 1 nucleotides of the 3' terminal nucleotide of the guide strand.
  • a wobble basepair occurs between the 3' terminal nucleotide of the guide strand and the target mRNA.
  • at least one wobble basepair is in the 5' portion of the guide strand and at least one wobble basepair is in the 3' portion of the guide strand.
  • at least one wobble basepair is within 13, 12, 11, 10, 9, 8, 7. 6, 5, 4, 3, 2, or 1 nucleotides of the 5 ' terminal nucleotide of the guide strand
  • at least one wobble bascpairs is within 1 3. 12, 1 1 , 10, 9, 8, 7, 6, 5, 4, 3. 2, or 1 nucleotides of the 3 ' terminal nucleotide of the guide strand.
  • a wobble basepair occurs at the 5' terminal nucleotide of the guide strand and/or at the 3' terminal nucleotide of the guide strand.
  • siRN ⁇ molecules of the invention are not necessary for effective silencing to occur.
  • three or four mismatches between a guide strand of an siRN ⁇ duplex and its target RNA, placed so as to still permit mRNA cleavage facilitates the release of cleaved target RNA from the RISC complex, thereby increasing the rate of enzyme turnover.
  • the efficiency of cleavage is greater when a G:U wobble basepair is present in the 5' or 3' portion of the complex termed between the siRN A and the target.
  • At least one terminal nucleotide of the RNA molecules described herein can be substituted with a nucleotide that does not form a Watson-Crick basepair with the corresponding nucleotide in a target mRN ⁇ .
  • a nucleic acid that reduces hRJIBDf 1 1 expression and/or activity can be expressed within cells from a nucleic acid encoding it under the control of a eukaryotic promoter ⁇ e.g., Izant and Weintraub. 1 ⁇ 85, Science. 22 ⁇ , 345; McGarry and Lmdquist, 19S6, Proc. Natl. Acad. Sci.. USA 83, 399; Scanlon et al., 1 991, Pr ⁇ c. Natl. ⁇ cad. Sci. IJSA, 88, 10591 -5; Kashani-Sabct el al, 1992, Antiscnse Res. Dev..
  • a eukaryotic promoter ⁇ e.g., Izant and Weintraub. 1 ⁇ 85, Science. 22 ⁇ , 345; McGarry and Lmdquist, 19S6, Proc. Natl. Acad. Sci.. USA 83,
  • nucleic acid can be expressed in eukaryotic cells tram the appropriate DN ⁇ /RN ⁇ vector. 1 he activity of such nucleic acids can be augmented by their release from the primary transcript by an enzymatic nucleic acid (Draper et al., PCT WO 93/23569, and Sullivan et al.. PCT WO 94/02595; Ohkawa et al., 1992, Nucleic ⁇ cids Symp.
  • an enzymatic nucleic acid Draper et al., PCT WO 93/23569, and Sullivan et al.. PCT WO 94/02595; Ohkawa et al., 1992, Nucleic ⁇ cids Symp.
  • nucleic acid molecules of the present invention are expressed from transcription units (see for example Couture ei aL, 1996, TIG., 12, 510) inserted into DNA or RNA vectors.
  • the recombinant vectors are preferably DNA plasm ids or viral vectors.
  • RNA expressing viral vectors can be constructed based on, but not limited to, adeno-associated virus, retrovirus, adenovirus, or alpha virus.
  • the recombinant vectors capable of expressing the nucleic acid molecules are delivered as described above, and persist in target cells.
  • nucleic acid molecules expressing vectors can be systemic, such as by intravenous or intramuscular administration, by administration to target cells ex-planted from the patient or subject followed by ⁇ introduction into the patient or subject, or by any other means that would allow for introduction into the desired target cell (for a review sec Couture et aL, 1996. TIG., 12, 510).
  • the invention features an expression vector comprising a nucleic acid sequence encoding at least one of the nucleic acid molecules of the instant invention is disclosed.
  • the nucleic acid sequence encoding the nucleic acid molecule of the instant invention is ⁇ pcrably linked in a manner which allows expression of that nucleic acid molecule.
  • the invention features an expression vector comprising: a) a transcription initiation region ⁇ e.g. , cukaryotic pol L II or III initiation region); b) a transcription termination region (e.g..
  • the vector can optionally include an open reading frame (ORF) for a protein operably linked on the 5' side or the 3 '-side of the sequence encoding the nucleic acid catalyst of the invention; and/or an mtron (intervening sequences).
  • ORF open reading frame
  • polymerase LIl polymerase LIl
  • Transcripts from pol Ll or poi 111 promoters arc expressed at high levels in all cells: the levels of a given pol II promoter in a given cell type depends on the nature of the gene regulatory sequences (enhancers, silencers, etc.) present nearby.
  • Prokaryotic RNA polymerase promoters are also used, providing that the prokaryotic RNA polymerase enzyme is expressed m the appropriate cells (Elroy- Stein and Moss. M Q O, Proc. Natl. Acad. Sci. USA. 87. 6743-7; Gao and Huang 1993, Nucleic Acids Res.. 2J . 2867-72; Lieber et ai, 1993. Methods En/ymoL, 21 7, 47-66; Zhou et ai, 1990, MoL Ceil BK)]., 10, 4529-37).
  • nucleic acid molecules, such as ribo/ymes expressed from such promoters can function in mammalian cells ⁇ e.g .
  • transcription units such as the ones derived from genes encoding U6 small nuclear (snRKA), transfer RKA (tRNA) and adenovirus VA RNA are useful in generating high concentrations of desired RNA molecules such as ribozymes in cells (Thompson et ai, supra; Couture and Stinchcomb, 19%, supra; Noonberg et ai, 1994, Nucleic Acid Res.. 22, 2830; Noonberg et ai, U.S. Pat. No.
  • ribozyme transcription units can be incorporated into a variety of vectors for introduction into mammalian cells, including but not restricted to. plasmid DNA vectors, viral DNA vectors (such as adenovirus or adeno-associated virus vectors), or viral RNA vectors (such as retroviral or alpbavirus vectors) (for a review see Couture and Stinchcomb, 1996, supra).
  • plasmid DNA vectors such as adenovirus or adeno-associated virus vectors
  • viral RNA vectors such as retroviral or alpbavirus vectors
  • the invention features an expression vector comprising nucleic acid sequence encoding at least one of the nucleic acid molecules of the invention, in a manner which allows expression of that nucleic acid molecule.
  • the expression vector comprises in one embodiment; a) a transcription initiation region; b) a transcription termination region; c) a nucleic acid sequence encoding at least one said nucleic acid molecule; and wherein said sequence is operably linked to said initiation region and said termination region, in a manner which allows expression and/or delivery of said nucleic acid molecule.
  • the expression vector comprises; a) a transcription initiation region: b) a transcription termination region; c) an open reading frame; d) a nucleic acid sequence encoding at least one said nucleic acid molecule, wherein said sequence is operably linked to the 3 '-end of said open reading frame; and wherein said sequence is operably linked to said initiation region, said open reading frame and said termination region, in a manner which allows expression and/or delivery of said nucleic acid molecule.
  • the expression vector comprises: a) a transcription initiation region; b) a transcription termination region; c) an intron; d) a nucleic acid sequence encoding at least one said nucleic acid molecule; and wherein said sequence is operably linked to said initiation region, said intron and said termination region, in a manner which allows expression and/or delivery of said nucleic acid molecule.
  • the expression vector comprises: a) a transcription initiation region; b) a transcription termination region; c) an intron; d) an open reading frame; e) a nucleic acid sequence encoding at least one said nucleic acid molecule, wherein said sequence is operably linked to the 3 '-end of said open reading frame; and wherein said sequence is operably linked to said initiation region, said intron, said open reading frame and said termination region, in a manner which allows expression arid/or delivery of said nucleic acid molecule.
  • nucleic acid molecules Methods for the delivery of nucleic acid molecules are described in Akhtar et a!., 1992, Trends Cell Bio., 2, 139; arid Delivery Strategies for Antisens ⁇ Oligonucleotide Therapeutics, ed. Akhtar; Sullivan el a!., PCT WO 94/02595, further describes the general methods for delivery of enzymatic RN ⁇ molecules. These protocols can be utilized for the delivery of virtually any nucleic acid molecule.
  • Nucleic acid molecules can be administered to cells by a variety of methods known to those familiar to the art, including, but not restricted to, encapsulation in liposomes, forming nanoplex with positively charged polymers ⁇ e.g , Histidine-Lysine copolymer), by iontophoresis, or by incorporation into other vehicles, such as hydrogels, cyclodextrins. biodegradable nanocapsules, and bioadhcsivc microspheres.
  • the nucleic acid/vehicle combination is locally delivered by direct injection or by use of an infusion pump.
  • the nucleic acids and compositions of the invention may be administered directly into a tumor.
  • routes of delivery include, but are not limited to oral (tablet or pill form) and/or intrathecal delivery (Gold, 1997. Ncurosciencc. 76, 1153-1158).
  • Other approaches include the use of various transport and carrier systems, for example, through the use of conjugates and biodegradable polymers.
  • o ⁇ drug delivery strategies including CNS delivery, see TIo et aL, I QQQ , Curr. Opin. MoI. Ther., 1 , 336-343 and Jain, Drug Delivery Systems: Technologies and Commercial
  • the molecules of the instant invention can be used as pharmaceutical agents.
  • Pharmaceutical agents prevent, inhibit the occurrence of, or treat (alleviate a symptom to some extent, preferably all of the symptoms of) a disease state in a subject.
  • the negatively charged polynucleotides of the invention can be administered and introduced into a subject by any standard means, with or without stabilizers, buffers, and the like, to form a pharmaceutical composition.
  • standard protocols for formation of liposomes can be followed.
  • the compositions of the present invention can also be formulated and used as tablets, capsules or elixirs for oral administration; suppositories for rectal administration; sterile solutions; suspensions tor injectable administration; and the other compositions known in the art.
  • the present invention also includes pharmaceutically acceptable formulations of the compounds described. These formulations include salts of the above compounds, e g. , acid addition salts, for example, salts of hydrochloric, hydrobromic, acetic acid, and benzene sulfonic acid.
  • a pharmacological composition or formulation refers to a composition or formulation m a form suitable for administration, e.g., systemic administration, into a cell or subject, preferably a human. Suitable forms, in part, depend upon the use or the route of entry, for example oral, transdermal, or by injection. Such forms should not prevent the composition or formulation from reaching a target cell. For example,
  • pharmacological compositions injected into the blood stream should be soluble.
  • Other factors include considerations such as toxicity and forms which prevent the composition or formulation from exerting its effect.
  • systemic administration in vivo systemic absorption or accumulation of drugs in the blood stream followed by distribution throughout the entire body.
  • Administration routes which lead to systemic absorption include, without limitations: intravenous, subcutaneous, intraperitoneal, inhalation, oral, in ⁇ rapulmonary and intramuscular.
  • Each of these administration routes exposes the desired negatively charged nucleic acids, to an accessible diseased tissue.
  • the rate of entry of a drug into the circulation has been shown to be a function of molecular weight or si/e.
  • the use of a liposome or other drug carrier comprising the compounds of the instant invention can potentially localize the drug, for example, in certain tissue types, such as the tissues of the reticular endothelial system (RES).
  • RES reticular endothelial system
  • a liposome formulation which can facilitate the association of drug with the surface of cells, such as, lymphocytes and macrophages is also useful. This approach can provide enhanced delivery of the drug to target cells by taking advantage of the specificity of macrophage and lymphocyte immune recognition of abnormal cells, such as cancer cells.
  • compositions or formulation that allows for the effective distribution of the nucleic acid molecules of the instant invention m the physical location most suitable for their desired activity.
  • agents suitable for formulation with the nucleic acid molecules of the instant invention include: S 3 EG conjugated nucleic acids, phospholipid conjugated nucleic acids, nucleic acids containing lipophilic moieties, phosphorothioates,
  • P-glycopr ⁇ tem inhibitors such as Pluronic P85 which can enhance entry of drugs into various tissues: biodegradable polymers, such as ⁇ listidine-Lysine copolymer and poly (DL-lactidc-coglycolide) microspheres for sustained release delivery after implantation (Emerich. DF et uL I QQQ , Cell Transplant, 8. 47-58) Alkemies. Inc. Cambridge. Mass.; and loaded nanoparticles, such as those made of polybutylcyanoacrylatc, which can deliver drugs across the blood brain barrier and can alter neuronal uptake mechanisms (Prog Neuropsychopharmacol Biol Psychiatry. 23, 941-949, 1999).
  • biodegradable polymers such as ⁇ listidine-Lysine copolymer and poly (DL-lactidc-coglycolide) microspheres for sustained release delivery after implantation (Emerich. DF et uL I QQQ , Cell Transplant, 8.
  • composition comprising
  • Nucleic acid molecules of the invention can also comprise covalently attached PEG molecules of various molecular weights. These formulations offer a method for increasing the accumulation of drugs in target tissues. This class of drug carriers resists opsonization and elimination by the mononuclear phagocytic system (MPS or RES), thereby enabling longer blood circulation times and enhanced tissue exposure for the encapsulated drug (Lasie et al. Chem. Rev, 1995, 95, 2601-2627: Ishiwata ei ai, Chem.
  • MPS or RES mononuclear phagocytic system
  • Liposomes are also likely to protect drugs from nuclease degradation to a greater extent compared to cationic liposomes, based on their ability to avoid
  • the present invention includes nucleic acid
  • compositions such as si RNA compositions, prepared as described in US 2003/0166601.
  • the present invention provides a composition comprising a nucleic acid ⁇ e.g., siRN ⁇ ), complcxcd with a cationic polymer, especially a I listidme- Lysine co-polymer, a hydrophilic polymer, such as PEG, and optionally further comprising a targeting moiety.
  • a targeting moiety as described above is utilized to target the desired nucleic acid molecule(s) to a ceil of interest
  • compositions comprising the nucleic acid molecules of the present invention include at least one targeting moiety, such as a ligand for a cell surface receptor or other cell surface marker that permits highly specific interaction of the composition comprising the nucleic acid molecule (the "'vector") with the target tissue or ceil.
  • the vector preferably will include an unshielded ligand or a shielded ligand.
  • the vector may include two or more targeting moieties, depending on the cell type that is to be targeted. Use of multiple (two or more) targeting moieties can provide additional selectivity in cell targeting, and also can contribute to higher affinity and/or avidity of binding of the vector to the target cell.
  • the relative molar ratio of the targeting moieties may be varied to provide optimal targeting efficiency.
  • Methods for optimizing ceil binding and selectivity in this fashion arc known in the art.
  • assays for measuring cell selectivity and affinity and efficiency of binding arc known in the art and can be used to optimize the nature and quantity of the targeting ligand(s).
  • Suitable ligaiids include, but arc not limited to; the recognition sequence for integrin binding to extracellular matrix proteins (RGD) and monoclonal antibodies against receptors o ⁇ the surface of tumor cells or epithelial cells.
  • RGD extracellular matrix proteins
  • a targeting moiety is sialyl-Lcwis x , where the composition is intended for treating a region of inflammation.
  • Other peptide Kenya may be identified using methods such as phage display (F. Bartoli et a!., Isolation of peptide ligands for tissue-specific cell surface receptors, in Vector Targeting Strategies for Therapeutic Gene Delivery (Abstracts form Cold Spring Harbor Laboratory 1999 meeting), 1999, p4) and microbial display (Georgiou et ⁇ /., Ultra-High Affinity Antibodies from Libraries Displayed on the Surface of Microorganisms and Screened by F ⁇ CS, in Vector Targeting Strategies for Therapeutic Gene Delivery (Abstracts form Cold Spring Harbor Laboratory 1999 meeting), 1999, p 3.). Ligands identified in this manner are suitable for use in the present invention.
  • Li VE ⁇ Direct selection of Targeting Ligands from Combinatorial Libraries, in Vector Targeting Strategies for Therapeutic Gene Delivery (Abstracts form Cold Spring Harbor Laboratory 1999 meeting), 1999. p8, ⁇ .
  • the present invention includes nucleic acid
  • the present invention provides a nucleic acid of the present invention in a composition comprising the histidine- lysine copolymers (also referred to herein as PolyTranTM) as described in US Patents 7,163,695, 7,070,807 and 6.692,911 either alone or in combination with PEG (e.g., branched or unbranched PEG or a mixture of both) or in combination with PEG and a targeting moiety.
  • PEG e.g., branched or unbranched PEG or a mixture of both
  • compositions prepared for storage or administration which include a pharmaceutically effective amount, of the desired compounds in a pharmaceutically acceptable carrier or diluent.
  • Acceptable carriers or diluents for therapeutic use are well known in the pharmaceutical art, and are described, for example, in Remington: The Science and Practice of Pharmacy, 20th Edition.
  • preservatives such as sodium benzoate, sorbic acid and esters of p-hydroxybenzoic acid.
  • antioxidants and suspending agents can be used.
  • a pharmaceutically effective dose is that dose required to prevent, inhibit the occurrence, or treat (alleviate a symptom to some extent, preferably all of the symptoms) of a disease state.
  • the pharmaceutically effective dose depends on the type of disease, the composition used, the route of administration, the type of mammal being treated, the physical characteristics of the specific mammal under consideration, concurrent medication, and other factors which those skilled in the medical arts will recognize.
  • an amount between 0.1 mg/kg and 100 mg/kg body weight/day of active ingredients is administered dependent upon potency of the negatively charged polymer.
  • nucleic acid molecules of the invention and formulations thereof can be administered orally, topically, parenterally, by inhalation or spray or rectaliy in dosage unit formulations containing conventional non-toxic pharmaceutically acceptable carriers, adjuvants and vehicles.
  • parenteral as used herein includes percutaneous. subcutaneous, intravascular (e.g., intravenous), intramuscular, or intrathecal injection or infusion techniques and the like.
  • a pharmaceutical formulation comprising a nucleic acid molecule of the invention and a pharmaceutically acceptable carrier.
  • One or more nucleic acid molecules of the invention can be present in association with one or more non-toxic pharmaceutically acceptable carriers and/or diluents and/or adjuvants, and if desired other active ingredients.
  • compositions containing nucleic acid molecules of the invention can be in a form suitable for oral use, for example, as tablets, troches, lozenges, aqueous or oily suspensions, dispersible powders or granules, emulsion, hard or soft capsules, or syrups or elixirs.
  • nucleic acid compositions of the invention can be used in combination with other nucleic acid compositions that target the same or different areas of the target gene ⁇ e.g., hRHBDFl). or that target other genes of interest.
  • the nucleic acid compositions of the invention can also be used in combination with any of a variety of treatment modalities, such as chemotherapy, surgery, radiation therapy, or small molecule regimens.
  • compositions intended for oral use can be prepared according to any method known to the art for the manufacture of pharmaceutical compositions and such compositions can contain one or more such sweetening agents, flavoring agents, coloring agents or preservative agents in order to provide pharmaceutically elegant and palatable preparations.
  • Tablets contain the active ingredient in admixture with non-toxic pharmaceutically acceptable excipients that are suitable for the manufacture of tablets. These excipients can be for example, inert diluents, such as calcium carbonate, sodium carbonate, lactose, calcium phosphate or sodium phosphate; granulating and
  • disintegrating agents for example, corn starch, or alginic acid
  • binding agents for example starch, gelatin or acacia
  • lubricating agents for example magnesium stearat ⁇ , stearic acid or talc.
  • the tablets can be uncoated or they can be coated by known techniques. In some cases such coatings can be prepared by known techniques to delay disintegration and absorption in the gastrointestinal tract and thereby provide a sustained action over a longer period.
  • a time delay material such as glyceryl monosterate or glyceryl dist ⁇ arat ⁇ can be employed.
  • Formulations for oral use can also be presented as hard gelatin capsules wherein the active ingredient is mixed with an inert solid diluent, for example, calcium carbonate, calcium phosphate or kaolin, or as soft gelatin capsules wherein the active ingredient is mixed with water or an oil medium, for example peanut oil, liquid paraffin or olive oil.
  • Aqueous suspensions contain the active materials in admixture with excipierits suitable for the manufacture of aqueous suspensions.
  • excipients are suspending agents, for example sodium carboxym ⁇ thylcellulos ⁇ , methylcellulose,
  • hydropropyl-methylcellulose, sodium alginate, polyvinylpyrrolidone, gum tragacanth and gum acacia; dispersing or wetting agents can be a naturally-occurring phosphatide, for example, lecithin, or condensation products of an alkyl ⁇ ne oxide with fatty acids, for example poiyoxycthylcnc stearate, or condensation products of ethylene oxide with long chain aliphatic alcohols, for example heptadeca ⁇ thyi ⁇ neoxycetanol, or condensation products of ethylene oxide with partial esters derived from fatty acids and a hcxitol such as poiyoxycthylcnc sorbitol monooleatc. or condensation products of ethylene oxide with partial esters derived from fatty acids and hexitol anhydrides, for example polyethylene sorbitan monoolcate.
  • the aqueous suspensions can also contain one or more
  • preservatives for example ethyl, or n-propyl p-hydroxybenzoate, one or more coloring agents, one or snore flavoring agents, and one or more sweetening agents, such as sucrose or saccharin.
  • Oily suspensions can be formulated by suspending the active ingredients in a vegetable oil, for example arachis oil, olive oil, sesame oil or coconut oil, or in a mineral oil such as liquid paraffin.
  • the oily suspensions can contain a thickening agent, for example beeswax, hard paraffin or cetyl alcohol.
  • Sweetening agents and flavoring agents can be added to provide palatable oral preparations.
  • These compositions can be preserved by the addition of an anti-oxidant such as ascorbic acid,
  • Dispersible powders and granules suitable for preparation of an aqueous suspension by the addition of water provide the active ingredient in admixture with a dispersing or wetting agent, suspending agent and one or more preservatives.
  • a dispersing or wetting agent for example sweetening, flavoring and coloring agents, can also be present.
  • compositions of the invention can also be in the form of oif-in-water emulsions.
  • the oily phase can be a vegetable oil or a mineral oil or mixtures of these.
  • Suitable emulsifying agents can be naturally-occurring gums, for example gum acacia or gum tragacanth, naturally-occurring phosphatides, for example soy bean, lecithin, and esters or partial esters derived from fatty acids and hexitol. anhydrides, for example sorbitan monooleate, and condensation products of the said partial esters with ethylene oxide, for example polyoxyethylene sorbitan monool ⁇ ate.
  • the emulsions can also contain sweetening and flavoring agents,
  • Syrups and elixirs can be formulated with sweetening agents, for example glycerol, propylene glycol, sorbitol, glucose or sucrose. Such formulations can also contain a demulcent, a preservative and flavoring and coloring agents.
  • sweetening agents for example glycerol, propylene glycol, sorbitol, glucose or sucrose.
  • Such formulations can also contain a demulcent, a preservative and flavoring and coloring agents.
  • compositions can be in the form of a sterile injectable aqueous or oleaginous suspension.
  • This suspension can be formulated according to the known art using those suitable dispersing or wetting agents and suspending agents that have been mentioned above.
  • the sterile injectable preparation can also be a sterile injectable solution or suspension in a non-toxic parentally acceptable diluent or solvent, for example as a solution in 1,3-butancdiol.
  • the acceptable vehicles and solvents that can be employed are water. Ringer's solution and isotonic sodium chloride solution.
  • sterile, fixed oils arc conventionally employed as a solvent or suspending medium.
  • any bland fixed oil can be employed including synthetic mono-or diglycerides.
  • fatty acids such as oleic acid find use in the preparation of injcctables.
  • the nucleic acid molecules of the invention can also be administered in the form of suppositories, e.g., for rectal administration of the drug.
  • suppositories e.g., for rectal administration of the drug.
  • These compositions can be prepared by mixing the drug with a suitable non-irritating ⁇ xcipient that is solid at ordinary temperatures but liquid at, the rectal temperature and will therefore melt in the rectum to release the drug.
  • suitable non-irritating ⁇ xcipient that is solid at ordinary temperatures but liquid at, the rectal temperature and will therefore melt in the rectum to release the drug.
  • Such materials include cocoa butter and polyethylene glycols.
  • Nucleic acid molecules of the invention can be administered parenterally in a sterile medium.
  • the drug depending on the vehicle and concentration used, can either be suspended or dissolved in the vehicle.
  • adjuvants such as local anesthetics, preservatives and buffering agents can be dissolved in the vehicle.
  • Dosage levels of the order of from about 0.01 mg to about 140 mg per kilogram of body weight per day are useful in the treatment of the disease conditions described herein (about 0.5 mg to about 7 g per patient or subject per day).
  • the amount of active ingredient that can be combined with the carrier materials to produce a single dosage form varies depending upon the host treated and the particular mode of administration.
  • Dosage unit forms generally contain between from about 1 mg to about 500 mg of an active ingredient,
  • the specific dose level for any particular patient or subject depends upon a variety of factors including the activity of the specific compound employed, the age, body weight, general health, sex, diet, time of administration, route of administration, and rate of excretion, drag combination and the severity of the particular disease undergoing therapy.
  • the composition can also be added to the animal feed or drinking water. It can be convenient to formulate the animal feed and drinking water compositions so that the animal takes in a therapeutically appropriate quantity of the composition along with its diet. It can also be convenient to present the composition as a prcmix for addition to the feed or drinking water.
  • nucleic acid molecules of the present invention can also be administered to a subject in combination with other therapeutic compounds to increase the overall therapeutic effect.
  • the use of multiple compounds to treat an indication can increase the beneficial effects while reducing the presence of side effects.
  • nucleic acid-based modulators of the invention are added directly as " 'naked' " nucleic acids, or can be complcxed with cationic polymers, including cationic lipids, packaged within liposomes, or otherwise 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 through injection or infusion pump, with or without their incorporation in biopolymers.
  • nucleic acid molecules of the instant invention may be used in compositions comprising multiple nucleic acid molecules (e g. , siRNAs) targeting different target sequences within the liRHBDF i gene or targeting sequences within other genes.
  • multiple nucleic acid molecules e g. , siRNAs
  • nucleic acid molecules of the instant invention can be used to treat diseases or conditions associated with altered expression and-'or activity of hRIIBDFl .
  • the small nucleic acid molecules described herein may be useful, for example, m providing compositions to inhibit, reduce or prevent recurrence of breast, lung, prostate, stomach, colorectal, brain, esophageal, bladder, pancreatic, cervical, head and neck, and ovarian cancer, melanoma, lymphoma, glioma, multidrug resistant cancers, arid any other cancerous diseases and/or other disease states, conditions, or traits associated with liRHBDFl gene expression or activity in a subject or organism.
  • nucleic acid molecules of the instant invention can also be used to prevent diseases or conditions associated with altered activity and/or expression of hRIIBDFl in individuals that are at risk for developing such a disease or condition.
  • t ⁇ treat or prevent a disease or condition associated with the expression levels of hRI IBDFl, the subject having the disease or condition, or at risk for developing the disease or condition, can be treated, as is evident to those skilled in the art, individually or in combination with one or more therapeutic agents under conditions suitable for the treatment.
  • the present invention provides methods for treating or preventing diseases or conditions that respond to the modulation of hRI IBDf 1 I expression comprising administering to a subject in need thereof an effective amount of a
  • composition comprising one or more of the nucleic acid molecules of the invention, such as those set forth in SEQ LD NOs: 173-269 and 454-637.
  • the present invention provides methods for treating or preventing diseases associated with expression of hRHBDFl comprising administering to a subject in need thereof an effective amount of any one or more of the nucleic acid molecules of the invention, such as those provided in SEQ ID KOs: 173-269 and 454-637, such that the expression of hRHBDFl in the subject is decreased, thereby treating or preventing the disease associated with expression of hRHBDF l .
  • the present invention provides methods for treating or preventing disease associated with expression of hRHBDFl comprising administering to a subject in need thereof an effective amount of a double stranded nucleic acid molecule of the invention, such as the pairs provided in Tables 1 ⁇ 8 (SEQ ID NOs: S -637).
  • the administered nucleic acid modulators of the invention consist of sequences complementary to the specific liRITBDFl target sequences provided in SEQ ID NOs: S -47 and 638.
  • the administered nucleic acid molecules of the invention consist of a sequence as set forth m SEQ ID NOs: 173-269 and 454-637.
  • the administered nucleic acid molecules of the invention are double-stranded and each double-stranded nucleic acid molecule consists of one pair nucleotide sequences set forth in Tables 1-8 (SEQ ID NOs: 1-637).
  • the compositions of the invention may be useful in methods for treating or preventing breast, lung, prostate, stomach, colorectal, brain, esophageal, bladder, pancreatic, cervical, head and neck, meningioma, kidney, endometrial, and ovarian cancer, melanoma, lymphoma, glioblastoma, multidrug resistant cancers, and any other cancerous diseases, or other conditions which respond to the modulation of liRi IBDFl expression.
  • the nucleic acid molecules of the invention can be used in combination with other known treatments to treat conditions or diseases discussed herein.
  • the described molecules may be useful in combination with one or more known therapeutic or diagnostic agents to treat breast, lung, prostate, stomach, colorectal, brain, esophageal, bladder, pancreatic, cervical, head and neck, meningioma, kidney, endometrial, and ovarian cancer, melanoma, lymphoma, glioblastoma, multidrug resistant cancers, and any other cancerous diseases or other conditions which respond to the modulation of hRHBDFl expression.
  • the nucleic acid molecules of the present invention may be useful to treat lung cancer, kidney cancer, pancreas cancer, breast cancer, head and neck cancer, stomach cancer or colon cancer.
  • therapeutic agents that may be useful in conjunction with the nucleic acid molecules of the present invention to treat disease as described herein include, but are not limited to chcmothcrape ⁇ tic agents, radiation therapy, immunosuppressive agents (such as cyclosporin, azathioprinc. methotrexate.
  • mycophcnolatc. and FK5U6. antibodies or immimoablativc agents (such as CAMP ATH, anti-CD3 antibodies or other antibody therapies, cytoxin, fludaribinc. cyclosporin, FK506, rapamycin, inycopbenolic acid, steroids, cytokines, and irradiation).
  • immimoablativc agents such as CAMP ATH, anti-CD3 antibodies or other antibody therapies, cytoxin, fludaribinc. cyclosporin, FK506, rapamycin, inycopbenolic acid, steroids, cytokines, and irradiation).
  • These agents inhibit either the calcium dependent phosphatase calcineurin (cyclosporinc and FK506) or inhibit the p7QS ⁇ kinase that is important for growth factor induced signaling (rapamycin).
  • nucleic acid molecules of the present invention are administered to a patient m
  • the cell compositions of the present invention are administered following B-cell ablative therapy such as agents that react with CD20, e.g., Riiuxa ⁇
  • compositions and methods are known in the art for identifying subjects having, or at risk for having the diseases or disorders associated with expression of hRHBDFl as described herein.
  • wobble basepair e.g., G/U, L'U, I/ ⁇ , and I/O between the antiscnsc strand of the duplex and its complementary target mRN ⁇ , among other things, is believed to increase RISC turnover.
  • Tables 4, 5, 7 and 8 list human RIlBDFl targeting siRNA molecules that create such wobble bascpairs.
  • Table 8 List of 25-mer siRNA targeting human RIIBDFl that create 2 wobble basepairs between the antiscnsc strand and the target niRNA. wherein the wobble creating nucleotides arc in the 3' portion of the antiscnse strand.
  • the nucleic acid duplexes described in this Example can be used for reducing the expression ofhRHBDFl and may be useful in a variety of therapeutic sellings, for example, in the treatment of cardiovascular disorders such as aortic valve disease and cancers including but not limited to breast, lung, prostate, stomach, colorectal, brain, esophageal, bladder, pancreatic, cervical, head and neck, meningioma, kidney, endometrial, and ovarian cancer, melanoma, lymphoma, glioblastoma, multidrug resistant cancers, and any other cancerous diseases, and/or other disease states, conditions, or traits associated with hRJ ⁇ BDFl gene expression or activity in a subject or organism.
  • cardiovascular disorders such as aortic valve disease and cancers including but not limited to breast, lung, prostate, stomach, colorectal, brain, esophageal, bladder, pancreatic, cervical, head and neck, meningioma, kidney, endometrial, and

Landscapes

  • Health & Medical Sciences (AREA)
  • Life Sciences & Earth Sciences (AREA)
  • Engineering & Computer Science (AREA)
  • Genetics & Genomics (AREA)
  • Biomedical Technology (AREA)
  • Chemical & Material Sciences (AREA)
  • Molecular Biology (AREA)
  • Organic Chemistry (AREA)
  • Biotechnology (AREA)
  • General Engineering & Computer Science (AREA)
  • Zoology (AREA)
  • Bioinformatics & Cheminformatics (AREA)
  • Wood Science & Technology (AREA)
  • Microbiology (AREA)
  • Plant Pathology (AREA)
  • Physics & Mathematics (AREA)
  • Biochemistry (AREA)
  • General Health & Medical Sciences (AREA)
  • Biophysics (AREA)
  • Medicines That Contain Protein Lipid Enzymes And Other Medicines (AREA)
  • Pharmaceuticals Containing Other Organic And Inorganic Compounds (AREA)

Abstract

La présente invention a pour objet des molécules d’acide nucléique qui inhibent l’expression des RHBDF1. La présente invention concerne également des procédés d’utilisation des molécules d’acide nucléique.
PCT/US2010/042336 2009-07-17 2010-07-16 Compositions comprenant des acides nucléiques modulant les rhbdf1 humains et procédés d’utilisation WO2011009082A2 (fr)

Applications Claiming Priority (2)

Application Number Priority Date Filing Date Title
US22649109P 2009-07-17 2009-07-17
US61/226,491 2009-07-17

Publications (2)

Publication Number Publication Date
WO2011009082A2 true WO2011009082A2 (fr) 2011-01-20
WO2011009082A3 WO2011009082A3 (fr) 2011-03-31

Family

ID=43450252

Family Applications (1)

Application Number Title Priority Date Filing Date
PCT/US2010/042336 WO2011009082A2 (fr) 2009-07-17 2010-07-16 Compositions comprenant des acides nucléiques modulant les rhbdf1 humains et procédés d’utilisation

Country Status (1)

Country Link
WO (1) WO2011009082A2 (fr)

Cited By (1)

* Cited by examiner, † Cited by third party
Publication number Priority date Publication date Assignee Title
CN104877030A (zh) * 2015-05-22 2015-09-02 南开大学 抑制HIF1α稳定性的多肽及其应用

Citations (2)

* Cited by examiner, † Cited by third party
Publication number Priority date Publication date Assignee Title
US20070003519A1 (en) * 2003-04-01 2007-01-04 Lu Patrick Y Targets for tumor growth inhibition
WO2009032930A2 (fr) * 2007-09-04 2009-03-12 Intradigm Corporation Compositions comprenant de l'arnsi de kinase lié à une intégrine humaine et leur procédé d'utilisation

Patent Citations (2)

* Cited by examiner, † Cited by third party
Publication number Priority date Publication date Assignee Title
US20070003519A1 (en) * 2003-04-01 2007-01-04 Lu Patrick Y Targets for tumor growth inhibition
WO2009032930A2 (fr) * 2007-09-04 2009-03-12 Intradigm Corporation Compositions comprenant de l'arnsi de kinase lié à une intégrine humaine et leur procédé d'utilisation

Non-Patent Citations (2)

* Cited by examiner, † Cited by third party
Title
DATABASE GENBANK [Online] 17 November 2007 'Homo sapiens mRNA for rhomboid family 1 variant, clone: HRC06404.' Database accession no. AK225408.1. *
YAN ET AL.: 'Human rhomboid family-1 gene silencing causes apoptosis or autophagy to epithelial cancer cells and inhibits xenograft tumor growth.' MOL CANCER THER. vol. 7, 2008, pages 1355 - 1364 *

Cited By (1)

* Cited by examiner, † Cited by third party
Publication number Priority date Publication date Assignee Title
CN104877030A (zh) * 2015-05-22 2015-09-02 南开大学 抑制HIF1α稳定性的多肽及其应用

Also Published As

Publication number Publication date
WO2011009082A3 (fr) 2011-03-31

Similar Documents

Publication Publication Date Title
CN107075515B (zh) C/EBPα组合物和使用方法
US20110046067A1 (en) COMPOSITIONS COMPRISING HUMAN EGFR-siRNA AND METHODS OF USE
US20030153521A1 (en) Nucleic acid treatment of diseases or conditions related to levels of Ras
US20100286241A1 (en) Compositions comprising k-ras sirna and methods of use
US20110065644A1 (en) Compositions comprising human pcsk9 and apolipoprotein b sirna and methods of use
US20100298409A1 (en) Compositions comprising stat3 sirna and methods of use thereof
US20110053862A1 (en) Compositions comprising survivin sirna and methods of use thereof
JP2005500025A (ja) 女性生殖疾病及び状態の核酸に基づく調節
US20110053861A1 (en) Compositions comprising nuclear factor-kappa b (nf-kb) sirna and methods of use
JP2016180011A (ja) がんを治療するための試薬および方法
EP1758999B1 (fr) Procedes pour inhiber la proliferation des cellules tumorales avec foxm1 arnsi
US20110105588A1 (en) Compositions comprising notch1 sirna and methods of use thereof
US20100280097A1 (en) Compositions comprising hif-1 alpha sirna and methods of use thereof
US20100279919A1 (en) Compositions comprising human integrin-linked kinase-sirna and methods of use thereof
WO2009143277A2 (fr) Compositions comprenant un arnsi hscn9a et leurs procédés d'utilisation
WO2011009082A2 (fr) Compositions comprenant des acides nucléiques modulant les rhbdf1 humains et procédés d’utilisation
US20030073207A1 (en) Enzymatic nucleic acid treatment of diseases or conditions related to levels of epidermal growth factor receptors
US20100273858A1 (en) Compositions comprising stat5 sirna and methods of use thereof
JP2020537636A (ja) 敗血症関連障害の処置のための組成物及び方法

Legal Events

Date Code Title Description
121 Ep: the epo has been informed by wipo that ep was designated in this application

Ref document number: 10800632

Country of ref document: EP

Kind code of ref document: A2

NENP Non-entry into the national phase in:

Ref country code: DE

122 Ep: pct application non-entry in european phase

Ref document number: 10800632

Country of ref document: EP

Kind code of ref document: A2