Nicked or gapped nucleic acid molecules and uses thereof

Abstract

The present disclosure provides meroduplex (nicked or gapped) ribonucleic acid molecules (mdRNA) that decreases or silences target gene expression. An mdRNA of this disclosure comprises at least three strands that combine to form at least two non-overlapping double-stranded regions separated by a nick or gap wherein one strand is complementary to a target gene RNA. In addition, the meroduplex may have one or more modifications or substitutions, such as nucleotide base, sugar, terminal cap structure, internucleotide linkage, or any combination of such modifications. Also provided are methods of decreasing expression of a target gene in a cell or in a subject to treat a disease related to altered expression of a target gene.

1 .- 64 . (canceled) 
     
     
         65 . A method for reducing expression of a target gene in a cell, said method comprising:
 contacting a target gene expressing cell with a composition comprising a 19 to 30 base pair double-stranded ribonucleic acid, wherein said target gene expression is reduced as a consequence of contacting said cell with said composition, said ribonucleic acid comprising   (a) an A strand having a length of 15 to 30 nucleotides and a nucleotide sequence that is complementary to a nucleotide sequence in said target gene;   (b) an S1 strand having a length of 5 to 25 nucleotides, wherein said S1 strand anneals to said A strand thereby forming a first double-stranded region of 5 to 15 base pairs; and   (c) an S2 strand having a length of 5 to 25 nucleotides, wherein said S2 strand anneals to said A strand thereby forming a second double-stranded region of 3 to 25 base pairs and wherein said annealed S2 strand is separated from said annealed S1 strand by a nick or a gap.   
     
     
         66 . The method of  claim 65  wherein said composition comprises a liposome, a hydrogel, a cyclodextrin, a biodegradable nanocapsule, a bioadhesive microsphere, or a proteinaceous vector. 
     
     
         67 . The method of  claim 66  wherein said liposome is a surface-modified liposome. 
     
     
         68 . The method of  claim 67  wherein said liposome comprises one or more lipids selected from the group consisting of a non-cationic lipid and a cationic lipid. 
     
     
         69 . The method of  claim 65  wherein said double-stranded ribonucleic acid is combined, complexed, or conjugated with a peptide. 
     
     
         70 . The method of  claim 69  wherein said peptide facilitates delivery of said double-stranded ribonucleic acid into said target cell. 
     
     
         71 . The method of  claim 69  wherein said peptide is selected from the group consisting of PN27, PN28, PN29, PN58, PN61, PN73, PN158, PN159, PN173, PN182, PN202, PN204, PN250, PN361, PN365, PN404, PN453, and PN509. 
     
     
         72 . The method of  claim 69  wherein said peptide comprises an N-terminal protein transduction domain from HIV TAT. 
     
     
         73 . The method of  claim 65  wherein overexpression or inappropriate expression of said target gene is associated with one or more disease or disorder. 
     
     
         74 . The method of  claim 73  wherein said disease or disorder is selected from the group consisting of a hyperproliferative disease or disorder, an angiogenic disease or disorder, a metabolic disease or disorder, and an inflammatory disease or disorder. 
     
     
         75 . The method of  claim 73  wherein said disease or disorder is selected from the group consisting of a cardiac disease, a pulmonary disease, a neovascularization, an ischemia, age-related macular degeneration, diabetic retinopathy, glomerulonephritis, diabetes, asthma, chronic obstructive pulmonary disease, chronic bronchitis, lymphangiogenesis, and atherosclerosis. 
     
     
         76 . The method of  claim 74  wherein said hyperproliferative disease or disorder is selected from the group consisting of a neoplasm, a carcinoma, a sarcoma, a tumor, and a cancer. 
     
     
         77 . The method of  claim 74  wherein said hyperproliferative disease or disorder is selected from the group consisting of an oral cancer, a throat cancer, a laryngeal cancer, an esophageal cancer, a pharyngeal cancer, a nasopharyngeal cancer, an oropharyngeal cancer, a gastrointestinal tract cancer, a gastrointestinal stromal tumor (GIST), a small intestine cancer, a colon cancer, a rectal cancer, a colorectal cancer, an anal cancer, a pancreatic cancer, a breast cancer, a cervical cancer, uterine cancer, a vulvar cancer, vaginal cancer, urinary tract cancer, bladder cancer, kidney cancer, adrenocortical cancer, islet cell carcinoma, gallbladder cancer, stomach cancer, prostate cancer, ovarian cancer, endometrial cancer, trophoblastic tumor, testicular cancer, penial cancer, bone cancer, osteosarcoma, liver cancer, extrahepatic bile duct cancer, skin cancer, basal cell carcinoma (BCC), lung cancer, small cell lung cancer, non-small cell lung cancer (NSCLC), brain cancer, melanoma, Kaposi's sarcoma, eye cancer, head and neck cancer, squamous cell carcinoma of head and neck, tymoma, thymic carcinoma, thyroid cancer, parathyroid cancer, Rippel-Linda syndrome, leukemia, acute myeloid leukemia, chronic myelogenous leukemia, acute lymphoblastic leukemia, hairy cell leukemia, lymphoma, non-Hodgkin's lymphoma, Burkitt's lymphoma, T-cell lymphoma, multiple myeloma, malignant pleural mesothelioma, Barrett's adenocarcinoma, and Wilm's tumor. 
     
     
         78 . The method of  claim 74  wherein said inflammatory disease or disorder is selected from the group consisting of diabetes mellitus, rheumatoid arthritis, pannus growth in inflamed synovial lining, collagen-induced arthritis, spondylarthritis, ankylosing spondylitis, multiple sclerosis, encephalomyelitis, inflammatory bowel disease, Chron's disease, psoriasis or psoriatic arthritis, myasthenia gravis, systemic lupus erythematosis, graft-versus-host disease, atherosclerosis, and an allergy.