Vectors Conditionally Expressing Protein

Abstract

This invention relates to the field of therapeutics. Disclosed are methods of generating conditionally expressing erythropoietin under the control of an ecdysone receptor-based gene expression modulation system in the presence of activating ligand and uses for therapeutic purposes in animals. The methods of the invention cause an in vivo increase in the expression of erythropoietin and an increase in the hematocrit or volume percentage of red blood cells in blood after administration of the ligand.

1 - 6 . (canceled) 
     
     
         7 . A vector comprising a polynucleotide encoding a gene switch, wherein the polynucleotide comprises (1) at least one transcription factor sequence which is operably linked to a promoter, wherein the at least one transcription factor sequence encodes a ligand-dependent transcription factor, and (2) a polynucleotide encoding one or more proteins operably linked to a promoter which is activated by the ligand-dependent transcription factor, wherein the one or more proteins is selected from the group consisting of a C1 esterase inhibitor, a kallikrein inhibitor, a bradykinin B2 receptor inhibitor, a prostaglandin synthase, a glucagon-like peptide-1 (GLP-1), a glucagon-like peptide-2 (GLP-2), adiponectin, leptin, and cystic fibrosis transmembrane conductance regulator (CFTR). 
     
     
         8 . The vector of  claim 7 , wherein one or more of the proteins is a human protein. 
     
     
         9 . The vector of  claim 7 , wherein the vector is a viral vector. 
     
     
         10 . The vector of  claim 9 , wherein the viral vector is selected from the group consisting of an adenovirus, an adeno-associated virus, a retrovirus, a pox virus, a baculovirus, a vaccinia virus, a herpes simplex virus, an Epstein-Barr virus, a geminivirus, a pseudorabies virus, a parvovirus, and a caulimovirus virus vector. 
     
     
         11 . The vector of  claim 7 , wherein the gene switch is an ecdysone receptor (EcR)-based gene switch. 
     
     
         12 . The vector of  claim 7 , wherein the polynucleotide encoding a gene switch comprises a first transcription factor sequence under the control of a first promoter and a second transcription factor sequence under the control of a second promoter, wherein a first transcription factor encoded by the first transcription factor sequence and a second transcription factor encoded by the second transcription factor sequence interact to form a complex which functions as a ligand-dependent transcription factor. 
     
     
         13 . The vector of  claim 7 , wherein the polynucleotide encoding a gene switch comprises a first transcription factor sequence and a second transcription factor sequence under the control of a promoter, wherein a first transcription factor encoded by the first transcription factor sequence and a second transcription factor encoded by the second transcription factor sequence interact to form a complex which functions as a ligand-dependent transcription factor. 
     
     
         14 . The vector of  claim 13 , wherein the first transcription factor sequence and the second transcription factor sequence are connected by an EMCV internal ribosomal entry site (IRES). 
     
     
         15 . The vector of  claim 7 , wherein one of the one or more proteins comprises an amino acid sequence at least 80%, 85%, 90%, 95%, 96%, 97%, 98%, 99%, or 100% identical to the amino acid sequence encoded by SEQ ID NO: 9, 10, or 11; or wherein one of the one or more proteins comprises an amino acid sequence at least 80%, 85%, 90%, 95%, 96%, 97%, 98%, 99% or 100% identical to the amino acid sequence of SEQ ID NO: 12, 13, 14, 15, 17, 18, 19, 20 or 21. 
     
     
         16 - 22 . (canceled) 
     
     
         23 . A method of producing a population of cells expressing one or more proteins, wherein the method comprises modifying the cells with a vector according to  claim 7 . 
     
     
         24 - 30 . (canceled) 
     
     
         31 . A population of cells produced according to the method of  claim 23 . 
     
     
         32 - 69 . (canceled) 
     
     
         70 . A composition comprising the vector of  claim 7 , or the population of cells of  claim 31 , and a pharmaceutically acceptable carrier. 
     
     
         71 . The composition of  claim 70 , which is administered systemically, intravenously, intratumorally, orally, intraperitoneally, intramuscularly, intravertebrally, intracerdbrally, intrathecally, intradermally, or subcutaneously. 
     
     
         72 - 73 . (canceled) 
     
     
         74 . A kit comprising the vector of  claim 7 , or the population of cells of  claim 31 . 
     
     
         75 . (canceled) 
     
     
         76 . The vector of  claim 7 , wherein the ligand that activates the ligand-dependent transcription factor is a diacylhydrazine. 
     
     
         77 . The vector of  claim 76 , wherein the diacylhydrazine is RG-115819, RG-115830 or RG-115932. 
     
     
         78 . The vector of  claim 7 , wherein the ligand that activates the ligand-dependent transcription factor is an amidoketone or oxadiazoline. 
     
     
         79 - 81 . (canceled) 
     
     
         82 . A kit comprising the vector of  claim 7 , or the population of cells of  claim 31 , and a ligand. 
     
     
         83 . The kit of  claim 82 , wherein the ligand is a diacylhydrazine. 
     
     
         84 . The kit of  claim 83 , wherein the diacylhydrazine is RG-115819, RG-115830 or RG-115932. 
     
     
         85 . The kit and ligand of  claim 82 , wherein the ligand is an amidoketone or oxadiazoline.